JP7528065B2 - Methods and compositions for treating age-related dysfunction using CCR3 inhibitors - Google Patents

Description

The following is provided as background information only and is not admitted to be prior art to the present invention.

Aging is a significant risk factor for multiple human diseases, including cognitive impairment, cancer, arthritis, blindness, osteoporosis, diabetes, cardiovascular disease, and stroke. Synapse loss, in addition to the normal synapse loss during natural aging, is an early pathological event common to many neurodegenerative conditions and is the most highly correlated neuronal and cognitive dysfunction associated with these conditions. Thus, aging remains the single most dominant risk factor for dementia-related neurodegenerative diseases such as Alzheimer's disease (AD) (Bishop, N.A. et al., Neural mechanisms of aging and cognitive decline. Nature 464(7288), 529-535(2010); Heeden, T. et al., Insights into the aging mind: a view from cognitive neuroscience. Nat. Rev. Neurosci. 5(2), 87-96(2004); Mattson, M.P., et al., Aging and neuronal vulnerability. Nat. Rev. Neurosci. 7(4), 278-294(2006)). Aging affects all tissues and functions of the body, including the central nervous system, and declines in functions such as cognition and motor activity can have a serious impact on quality of life.

Treatment of cognitive and motor decline associated with neurodegeneration has had limited success in preventing and reversing impairment. It is therefore important to identify new therapies to preserve cognitive integrity by protecting against, preventing or reversing the effects of aging. Unfortunately, pharmacological treatment of cognitive and motor dysfunction faces significant hurdles. For example, therapeutic agents such as small molecules that are effective in treating cognitive decline and motor activity would need to cross the blood-brain barrier (BBB). Passing through the BBB is an exception, and 98% of all small molecules do not pass through it (Pardridge, William M., The Blood-Brain Barrier: Bottleneck in Brain Drug Development, NeuroRx: The J. of the Am. Society for Experimental NeuroTherapeutics, 2:3-14, 2005).

As a result, there are limited therapies for neurodegenerative diseases such as Alzheimer's disease, Huntington's disease, Parkinson's disease, and amyotrophic lateral sclerosis (ALS), and in fact, Alzheimer's disease has a very high failure rate in clinical development across all disease areas (99.6% failure rate over the last 20 years) (see above and Burke, M., Renewed focus on dementia checked by drug challenges, Chemistry World, Jul. 3, 2014). BBB-permeable drugs approved for treating dementias such as Alzheimer's disease, such as donepezil, are temporarily effective but do not work in all patients (Banks, W.A., Drug Delivery to the Brain in Alzheimer's Disease: Consideration of the Blood-brain Barrier, Adv. Drug. Deliv. Rev. 64(7):629-39 (2012) and Burke, M., supra). Similarly, levodopa, which treats Parkinson's disease symptoms such as motor deficits and which crosses the BBB, eventually loses its efficacy. An additional hurdle for therapeutic agents crossing the BBB is that increasing the lipophilicity of a compound (which tends to increase the compound's BBB permeability) often results in greater clearance from the blood, leading to reduced bioavailability; increased lipophilicity offsets the plasma area under the curve (AUC) and minimizes brain uptake (Pardridge, William M., supra).

The present invention overcomes these problems. For example, the compound of the present invention, kagoubutu1, not only exhibits resistance to crossing the BBB at significant concentrations, but also unexpectedly improves symptoms associated with age-related neurodegeneration and age-related motor decline. Thus, the present invention can function peripherally without the actions that have been deemed necessary for effective cognitive drugs, i.e., without acting directly on the central nervous system. Also, certain embodiments of the present invention can cross the BBB at significant concentrations, but provide an alternative mechanism of action to current therapies for cognitive and motor disorders through their ability to antagonize the CCL11/CCR3 pathway.

The compounds of the present invention act as antagonists of c-c motif chemokine receptor 3 (CCR3), the receptor for eotaxin-1. Eotaxin-1 (CCL11) is a protein whose plasma levels increase with age and has been shown to impair cognitive function (Villeda et al., The aging systemic milieu negatively regulates neurogenesis and cognitive function, Nature, 477(7362):90-94 (2011)). Eotaxin/CC11 acts primarily on the G protein-coupled receptor CCR3 expressed on eosinophils in the periphery, as well as on neurons and glial cells in the central nervous system (Xia, M, et al., Immunohistochemical Study of the β-Chemokine Receptors CCR3 and CCR5 and Their Ligands in the Normal and Alzheimer's Disease Brains, Am. J. Pathol. 153(1); 31-37 (1998)). In Alzheimer's disease, CCR3 levels have been observed to be elevated in the CNS (ibid.). Thus, the cognitive effects of antagonizing CCR3 in Alzheimer's disease and related disorders are expected to be mediated by the eotaxin-1/CCR3 interaction found in the central nervous system. However, compound 1 (a compound of the present invention) unexpectedly improves cognitive function and stimulates neurogenesis, despite its inability to cross the BBB in significant concentrations.

Methods are provided for treating age-related dysfunction, neurodegenerative diseases, and associated cognitive and motor decline in a patient, including, by way of example and not limitation, Alzheimer's disease, Parkinson's disease, dementia with Lewy bodies, frontotemporal dementia, Huntington's disease, amyotrophic lateral sclerosis, multiple sclerosis, glaucoma, myotonic dystrophy, vascular dementia, progressive supranuclear palsy, and the like. Aspects of the method include modulating CCR3, the primary receptor for CCL11/eotaxin-1, through administration of a therapeutically effective amount of a CCR3 antagonist of the invention. The method includes administering a therapeutically effective amount of a CCR3 antagonist to a subject or patient and monitoring for a predetermined clinical endpoint. Also included are methods of treating neurodegenerative diseases and associated cognitive and motor decline with peripherally acting agents, i.e., agents that do not cross the blood-brain barrier in significant concentrations but that ameliorate the disease (e.g., improve cognitive or motor activity).

Methods of using diagnostic or companion diagnostic tests in connection with the described age-related disorders are also provided. By way of example, such diagnostic or companion diagnostics include, but are not limited to, devices capable of determining or detecting the presence of a subset of white blood cells from a subject. Diagnostic or companion diagnostic devices may also determine or detect the presence, relative or absolute concentration, relative or absolute number of eosinophils from the blood or tissue of a subject. Such diagnostic or companion diagnostic devices may be used in combination.

INCORPORATION BY REFERENCE All publications, patents, and patent applications mentioned in this specification are incorporated by reference in their entirety to the same extent as if each individual publication or patent application was specifically and individually indicated to be incorporated by reference.

Figure 1 shows the total number of doublecortin (DCX) positive cells in the hippocampus of 2-month-old and 18-month-old C57B1/6 mice treated with IgG (n=18 in each group), 18-month-old mice treated with either high dose (n=16) or low dose (n=31) of Compound 1 for 2 weeks, and 18-month-old mice treated with low dose (n=16) of Compound 1 ("Cmpd1") for 4 weeks. Data shown are mean ± SEM, ^* P<0.05, ^*** P<0.001. Figure 1 shows the total number of 5-bromo-2'-deoxyuridine (BrdU) positive cells in the hippocampus of 2- and 18-month-old C57B1/6 mice treated with IgG (n=19 in both groups) and 18-month-old mice treated for 2 weeks with either high (n=16) or low dose (n=24) of Compound 1. Data shown are mean ± SEM, ^*** P<0.001. Figure 1 shows the effect of Compound 1 on the time spent in novel (N) versus familiar/old ("F" or "O") arms in a Y-maze test in young and old C57B1/6 mice. Two-month-old mice were treated with IgG for 2 weeks (n=18). Eighteen-month-old mice were treated with IgG for 2 weeks (n=18), high dose Compound 1 for 2 weeks (n=15), low dose Compound 1 for 2 weeks (n=31), or low dose Compound 1 for 4 weeks (n=16). Data shown are mean ± SEM, ^** P<0.05, ^*** P<0.01. Figure 1 shows the effect of Compound 1 on the total number of visits to novel (N) versus familiar (F) arms in the Y-maze test in young and old C57B1/6 mice. Two-month-old mice were treated with IgG for 2 weeks (n=18). 18-month-old mice were treated with IgG for 2 weeks (n=18), high dose Compound 1 for 2 weeks (n=15), low dose Compound 1 for 2 weeks (n=31), or low dose Compound 1 for 4 weeks (n=16). Data shown are mean ± SEM, ^* P<0.05, ^** P<0.01, ^*** P<0.001. Figure 1 shows the effect of Compound 1 on the number of novel and familiar arm entries ("visits") by C57B1/6 mice during the Y-maze test. The visits to each arm were plotted by treatment group and paired t-tests were performed. Three-month-old mice (n=15) were subcutaneously infused with vehicle (veh) via an Alzet minipump (0.5 μL/hr) for 4 weeks with one supplement. 16.5-month-old mice were subcutaneously infused with either vehicle (veh, n=16) or 50 mg/mL Compound 1 (n=16) via an Alzet minipump (0.5 μL/hr) for 4 weeks with one supplement. Data shown are mean ± SEM, ^* P<0.05. Figure 1 shows the effect of Compound 1 on the mean time taken by C57B1/6 mice to find the target hole in the Barnes maze test. Mean times are plotted by treatment group. Three-month-old mice (n=18) were infused subcutaneously with vehicle (veh) via Alzet minipumps (0.5 μL/hr) for 4 weeks with one supplement. 16.5-month-old mice were infused subcutaneously with either vehicle (veh, n=15) or 50 mg/mL Compound 1 (n=15) via Alzet minipumps (0.5 μL/hr) for 4 weeks with one supplement. Data shown are mean ± SEM. Figure 1 shows the effect of Compound 1 on the difference in latency to find the target hole between the last trial and the first trial on the last day of the Barnes maze test in C57B1/6 mice. The differences were plotted by treatment group and the data were subjected to an unpaired t-test. Three-month-old mice (n=18) were infused subcutaneously with vehicle (veh) via Alzet minipumps (0.5 μL/hr) for 4 weeks with one supplement. 16.5-month-old mice were infused subcutaneously with either vehicle (veh, n=15) or 50 mg/mL Compound 1 (n=15) via Alzet minipumps (0.5 μL/hr) for 4 weeks with one supplement. Data shown are mean ± SEM. Figure 1 shows the effect of Compound 1 on neurogenesis in C57B1/6 mice by detecting BrdU positive cells in whole sections of the dentate gyrus. The number of BrdU positive cells from the dentate gyrus was plotted by treatment group, and the data was subjected to an unpaired t-test between 16.5 month old groups. Three month old mice (n=17) were subcutaneously infused with vehicle (veh) via Alzet minipump (0.5 μL/hr) for 4 weeks with one supplement. 16.5 month old mice were subcutaneously infused with either vehicle (veh, n=17) or 50 mg/mL Compound 1 (n=17) via Alzet minipump (0.5 μL/hr) for 4 weeks with one supplement. Data shown are mean ± SEM, ^* P<0.05. The levels of Compound 1 detected in the CSF of both young (n=2) and aged (n=3) C57Bl/6 mice (all levels were <10 nM) are shown. Compound 1 levels were detected using mass spectrometry. Figure 1 shows the time course distribution of Compound 1 in tissues of C57BL/6JOlaHsd mice, measured as area under the curve (AUC). Compounds were tagged with carbon-14 label and measurements were taken at 1 hour, 24 hours and 168 hours. Figure 1 shows the pharmacokinetic profile of Compound 1 following oral (P.O.) dosing. Compound 1 was measured in plasma from male 2-month-old C57Bl/6 mice dosed at either 30 mg/kg or 150 mg/kg by oral gavage at 20 minutes, 2 hours, 8 hours, and 12 hours post-dose. Plasma concentrations following drug administration were plotted over time. Figure 1 shows the effect of Compound 1 on exploration preference during the open field test in young C57B1/6 mice. Exploratory preference was plotted as the ratio of time spent in the periphery to time spent in the center, and data were subjected to one-way ANOVA. Two-month-old mice were orally dosed with vehicle twice daily (n=11), or with Compound 1 orally at 30 mg/kg twice daily (n=12), or with vehicle orally twice daily and recombinant murine CCL11 (eotaxin-1, or "rmE") intraperitoneally (n=14), or with Compound 1 orally at 30 mg/kg twice daily and recombinant murine CCL11 (n=14). Data shown are mean ± SEM, ^** P<0.01. Figure 1 shows the effect of Compound 1 on time spent in novel versus familiar arms in the Y-maze test in young C57B1/6 mice. Time spent in novel versus familiar arms was plotted for each treatment group and data were subjected to paired t-tests. Two-month-old mice were orally administered vehicle twice daily (n=13), orally administered Compound 1 at 30 mg/kg twice daily (n=14), orally administered vehicle twice daily and intraperitoneally administered recombinant murine CCL11 (eotaxin-1) (n=15), or orally administered Compound 1 at 30 mg/kg twice daily and intraperitoneally administered recombinant murine CCL11 (n=15). Data shown are mean ± SEM, ^** P<0.01. Figure 1 shows the effect of Compound 1 on the ratio of time spent in novel versus familiar arms in the Y-maze test in young C57B1/6 mice. Proportions were plotted by treatment group and data were subjected to ANOVA, Kruskal-Wallis test, and post-hoc. Two-month-old mice were orally administered vehicle twice daily (n=12), or Compound 1 orally at 30 mg/kg twice daily (n=14), orally administered vehicle twice daily and intraperitoneally administered recombinant murine CCL11 (eotaxin-1) (n=15), or orally administered Compound 1 orally at 30 mg/kg twice daily and intraperitoneally administered recombinant murine CCL11 (n=15). Data shown are mean ± SEM, ^* P<0.05. Figure 1 shows the effect of Compound 1 on the ratio of novel vs. familiar arm entries in the Y-maze test in young C57B1/6 mice. Ratios were plotted by treatment group and data were t-tested between vehicle and recombinant murine CCL11 treatment. Two-month-old mice were orally administered vehicle twice daily (n=13), orally administered Compound 1 at 30 mg/kg twice daily (n=14), orally administered vehicle twice daily and intraperitoneally administered recombinant murine CCL11 (eotaxin-1) (n=15), or orally administered Compound 1 at 30 mg/kg twice daily and intraperitoneally administered recombinant murine CCL11 (n=15). Data shown are mean ± SEM, ^* P<0.05. Figure 1 shows the effect of Compound 1 on the mean latency to find the target hole during the Barnes maze test in young C57B1/6 mice. Mean latency is plotted as units of time for each trial and for each treatment group. Two-month-old mice were orally dosed with vehicle twice daily (n=13), or with Compound 1 orally at 30 mg/kg twice daily (n=14), or with vehicle orally twice daily and recombinant murine CCL11 (eotaxin-1) intraperitoneally (n=15), or with Compound 1 orally at 30 mg/kg twice daily and recombinant murine CCL11 (n=15). Data shown are mean ± SEM. Figure 1 shows the effect of Compound 1 on the latency to find the target hole during the Barnes maze test in young C57B1/6 mice. For each treatment group, averaged latencies over the last three trials were plotted and data were subjected to unpaired t-tests. Two-month-old mice were orally administered vehicle twice daily (n=13), or Compound 1 orally at 30 mg/kg twice daily (n=14), orally administered vehicle twice daily and intraperitoneally administered recombinant murine CCL11 (eotaxin-1) (n=15), or orally administered Compound 1 orally at 30 mg/kg twice daily and intraperitoneally administered recombinant murine CCL11 (n=15). Data shown are mean ± SEM, ^* P<0.05. The effect of Compound 1 on memory for novel arms in the Y-maze is shown by the number of novel arm entries in total entries. Compound 1 was orally administered to 24-month-old mice twice daily at 30 mg/kg or vehicle. Data shown are mean ± SEM, ^** P<0.01. The effect of Compound 1 on total distance traveled in a Y-maze as a measure of locomotor activity is shown. Twenty-four-month-old mice were orally administered Compound 1 at 30 mg/kg twice daily or vehicle. Data shown are mean ± SEM, ^* P<0.05. Figure 1 shows the effect of Compound 1 on the number of entries into novel and familiar arms ("visits") by C57B1/6 mice in the Y-maze test at 24 months of age. The number of visits to each arm was plotted by treatment group and paired t-tests were performed. Mice aged 23 months were subcutaneously administered either vehicle control or Compound 1 BID (twice daily) for 21 days. After 3 weeks, mice were subjected to the Y-maze test. All mice were injected intraperitoneally with BrdU at 50 mg/kg for 5 consecutive days immediately prior to the start of treatment. Data shown are mean ± SEM, ^* P<0.05, ^** P<0.01, ^*** P<0.001 (by paired Student's t-test between novel and familiar arms). Figure 1 shows the effect of Compound 1 on the ratio of novel vs. familiar arm entries in the Y-maze test in 24-month-old mice. Ratios were plotted by treatment group and t-tests were performed on the data between vehicle and Compound 1 treatment. Mice aged 23 months were subcutaneously administered either vehicle control or Compound 1 BID (twice daily) for 21 days. After 3 weeks, mice were subjected to the Y-maze test. All mice were injected intraperitoneally with BrdU at 50 mg/kg for 5 consecutive days immediately prior to the start of treatment. Data shown are mean ± SEM, ^* P<0.05, ^** P<0.01, ^*** P<0.001 compared to control by Student's t-test. Figure 1 shows the effect of Compound 1 on time spent in novel versus familiar arms in the Y-maze test in 24-month-old C57B1/6 mice. Time spent in novel versus familiar arms was plotted by treatment group and data were subjected to paired t-test. 23-month-old mice were subcutaneously administered either vehicle control or Compound 1 BID (twice daily) for 21 days. After 3 weeks, mice were subjected to the Y-maze test. All mice were injected intraperitoneally with BrdU at 50 mg/kg for 5 consecutive days immediately prior to the start of treatment. Data shown are mean ± SEM, ^* P<0.05, ^** P<0.01, ^*** P<0.001 (by paired Student's t-test between novel and familiar arms). Figure 1 shows the effect of Compound 1 on the ratio of time spent in novel versus familiar arms (duration) in the Y-maze test in 24-month-old C57B1/6 mice. Ratios were plotted by treatment group and t-tests were performed on the data. 23-month-old mice were subcutaneously administered either vehicle control or Compound 1 BID (twice daily) for 21 days. After 3 weeks, mice were subjected to the Y-maze test. Immediately prior to the start of treatment, all mice were injected intraperitoneally with BrdU at 50 mg/kg for 5 consecutive days. Data shown are mean ± SEM, ^* P<0.05, ^** P<0.01, ^*** P<0.001 (compared to control by Student's t-test). Figure 1 shows the effect of Compound 1 on the mean speed during the Y-maze test in 24-month-old C57B1/6 mice. Mean speed was plotted by treatment group and t-tests were performed on the data. 23-month-old mice were administered either vehicle control or Compound 1 subcutaneously BID (twice daily) for 21 days. After 3 weeks, mice were subjected to the Y-maze test. All mice were injected intraperitoneally with BrdU at 50 mg/kg for 5 consecutive days immediately prior to the start of treatment. Data shown are mean ± SEM, ^* P<0.05, ^** P<0.01, ^*** P<0.001 compared to control by Student's t-test. Figure 1 shows the effect of Compound 1 on the mean time taken by 24-month-old C57B1/6 mice to find the target hole in the Barnes maze test. Mean times are plotted by treatment group. 23-month-old mice were subcutaneously administered either vehicle control or Compound 1 BID (twice daily) for 21 days. After 3 weeks, mice were subjected to the Barnes maze test. Data shown are mean ± SEM, ^* P<0.05 compared to control by Student's t-test. Figure 1 shows the effect of Compound 1 on the speed averaged over all trials for each treatment group in the Barnes maze test in 24-month-old C57B1/6 mice. 23-month-old mice were subcutaneously administered either vehicle control or Compound 1 BID (twice daily) for 21 days. After 3 weeks, mice were subjected to the Barnes maze test. Data shown are mean ± SEM, ^* P<0.05 (compared to control by Student's t-test). Figure 1 shows the effect of Compound 1 on distance traveled in the open field test in 24-month-old C57B1/6 mice. Mean distance traveled is plotted for each treatment group. 23-month-old mice were subcutaneously dosed BID (twice daily) with either vehicle control or Compound 1 for 21 days. After 3 weeks, mice were subjected to the open field test. Data shown are mean ± SEM. Figure 1 shows the effect of Compound 1 on speed in the open field test in 24-month-old C57B1/6 mice. The average speed of the mice is plotted for each treatment group. 23-month-old mice were subcutaneously dosed BID (twice daily) with either vehicle control or Compound 1 for 21 days. After 3 weeks, mice were subjected to the open field test. Data shown are the mean ± SEM. Figure 1 shows the effect of Compound 1 on plasma cytokine TNFα levels in 24-month-old C57B1/6 mice. 23-month-old mice were subcutaneously dosed BID (twice daily) with either vehicle control or Compound 1 for 21 days. Compound 1-treated mice tended to have lower levels of this proinflammatory cytokine than vehicle-treated mice. Figure 1 shows the effect of Compound 1 on plasma IL6 cytokine levels in 24-month-old C57B1/6 mice. 23-month-old mice were subcutaneously dosed BID (twice daily) with either vehicle control or Compound 1 for 21 days. Compound 1-treated mice tended to have lower levels of this proinflammatory cytokine than vehicle-treated mice. Figure 1 shows the effect of Compound 1 on plasma IL1β cytokine levels in 24-month-old C57B1/6 mice. 23-month-old mice were subcutaneously dosed BID (twice daily) with either vehicle control or Compound 1 for 21 days. Compound 1-treated mice tended to have lower levels of this proinflammatory cytokine than vehicle-treated mice. Figure 1 shows the effect of Compound 1 on plasma IL5 cytokine levels in 24-month-old C57B1/6 mice. 23-month-old mice were subcutaneously dosed BID (twice daily) with either vehicle control or Compound 1 for 21 days. Compound 1-treated mice tended to have lower levels of this proinflammatory cytokine than vehicle-treated mice. Figure 1 shows the effect of Compound 1 on plasma IL17 cytokine levels in 24-month-old C57B1/6 mice. 23-month-old mice were subcutaneously dosed BID (twice daily) with either vehicle control or Compound 1 for 21 days. Compound 1-treated mice tended to have lower levels of this proinflammatory cytokine than vehicle-treated mice. 1 shows the effect of Compound 1 on activated microglia in 24-month-old C57B1/6 mice. 23-month-old mice were subcutaneously dosed BID (twice daily) with either vehicle control or Compound 1 for 21 days. Compound 1-treated mice tended to have lower levels of CD68+ activated microglia than vehicle-treated mice. Figure 1 shows the effect of Compound 1 on the percentage of eosinophils to total white blood cell (WBC) counts in 24-month-old C57BI/6 mice subcutaneously dosed BID with either saline control or 30 mg/kg Compound 1 for 3 weeks. Compound 1 reduced endogenous age-related elevation of blood eosinophils. FIG. 1 shows the effect of Compound 1 on the percentage of eosinophils relative to total white blood cell (WBC) count in 3-month-old hairless mice orally dosed BID with either saline control or 30 mg/kg Compound 1 for 2 weeks. Compound 1 reduced the oxazolone-induced increase in blood eosinophils. Figure 1 shows the results of Compound 1 in the rotarod test for motor coordination. Compound 1 or vehicle was continuously infused by osmotic pumps into 24-month-old C57 mice for 4 weeks. Treated mice had a higher success rate than vehicle-treated mice in a binomial test. ^* P<0.05. Figure 1 shows the results of Compound 1 in the T-maze memory test. Compound 1 or vehicle was continuously infused by osmotic pumps into 24-month-old C57 mice for 4 weeks. Treated mice had a higher success rate than vehicle-treated mice in a binomial test. ^* P<0.05. The results of Compound 1 on fecal volume overnight are shown. Compound 1 or vehicle was continuously infused by osmotic pump to 24-month-old C57 mice for 4 weeks. The weight of fecal pellets was measured overnight. The fecal volume was significantly lower in Compound 1-treated mice than in vehicle-treated mice (by Student's t-test). ^* P<0.05. The results of Compound 1 on overnight water intake are shown. Compound 1 or vehicle was continuously infused by osmotic pump to 24-month-old C57 mice for 4 weeks. Total water intake was measured overnight. Compound 1-treated mice consumed significantly more water than vehicle-treated mice (by Student's t-test). ^* P<0.05. The results of Compound 1 on overnight food intake are shown. Compound 1 or vehicle was continuously infused by osmotic pump to 24-month-old C57 mice for 4 weeks. Total food intake was measured overnight. Total food intake overnight was not different between the two groups. 1 shows the effect of Compound 1 (Cmpd1) on the number of CD68 positive (CD68+) activated microglia in the brains of 3-month-old mice treated with LPS and then for 18 days with Compound 1. Compound 1-treated mice showed a decrease in CD68+ immunoreactivity, i.e., reduced gliosis. The effect of Compound 1 on anxiety in an open field test in 3-month-old mice that had been intraperitoneally administered LPS for 4 weeks and orally administered Compound 1 BID (twice a day) for 1 week was shown. Treatment with LPS increased preference for the perimeter of the open field, indicating increased anxiety. Treatment with Compound 1 reduced LPS-induced anxiety in the open field. Data shown are mean ± SEM, ^* P<0.05 (by Student's t-test). Figure 1 shows the effect of Compound 1 on the number of entries into novel and familiar arms in the Y-maze test in 3-month-old C57B1/6 mice treated with LPS. The number of visits to each arm was plotted for each treatment group and paired t-tests were performed. Three-month-old mice were intraperitoneally administered either vehicle or LPS for 6 weeks and orally administered vehicle control or Compound 1 BID (twice a day) for 3 weeks. Compound 1-treated mice showed a significant preference for the novel arm, but vehicle-treated mice did not. Data shown are mean ± SEM, ^** P<0.01, ^* P<0.05 (by paired Student's t-test between novel and familiar arms). Figure 1 shows the effect of Compound 1 on IL1β mRNA in the brains of 3-month-old C57B1/6 mice treated with LPS and/or Compound 1. Mice were administered either vehicle control or LPS intraperitoneally for 7 weeks and vehicle or Compound 1 orally administered BID (twice daily) for 4 weeks. Tissues were harvested and RNA was prepared from brain cortical tissue. IL1β mRNA levels were measured by qPCR and data are shown relative to vehicle control. Treatment with LPS showed a trend toward increased IL1β mRNA levels, while treatment with Compound 1 showed a significant decrease. Data shown are mean ± SEM, ^* P<0.05 by Student's t-test. Figure 1 shows the effect of Compound 1 on microglial activation in the hippocampus of 3-month-old C57B1/6 mice treated with LPS and/or Compound 1. Mice were administered either vehicle control or LPS intraperitoneally for 7 weeks and vehicle or Compound 1 orally administered BID (twice daily) for 4 weeks. Tissues were harvested and brain sections were subjected to immunohistochemistry for CD68, a marker of activated microglia. There was a trend towards increased CD68 levels with treatment with LPS and a strong trend towards decreased levels with treatment with Compound 1. Data shown are means ± SEM. Figure 1 shows the effect of Compound 1 on total microglia in the hippocampus of 3-month-old C57B1/6 mice treated with LPS and/or Compound 1. Mice were administered either vehicle control or LPS intraperitoneally for 7 weeks and vehicle or Compound 1 orally administered BID (twice daily) for 4 weeks. Tissues were removed and brain sections were subjected to immunohistochemistry for the microglial marker Iba1. A significant increase in Iba1 was observed when treated with LPS, whereas a tendency for a decrease was observed when treated with Compound 1. Data shown are mean ± SEM, ^*** P<0.001, ^* P<0.05 (by Student's t-test). Figure 1 shows the effect of Compound 1 on total astrocyte number in the hippocampus of 3-month-old C57B1/6 mice treated with LPS and/or Compound 1. Mice were administered either vehicle control or LPS intraperitoneally for 7 weeks and vehicle or Compound 1 orally administered BID (twice daily) for 4 weeks. Tissues were harvested and brain sections were subjected to immunohistochemistry for GFAP (a marker for astrocytes). A trend towards reduction was observed when treated with Compound 1. Data shown are mean ± SEM. Shown are the dosing regimens of three groups of C57BL/6 mice treated with (1) control for both LPS and Compound 1, (2) vehicle control for LPS plus Compound 1, or (3) control for both LPS plus Compound 1. All three groups were treated with control, LPS, or Compound 1 on the same day for three consecutive days. The graph shows the results of histological analysis of mouse brains in an acute model of LPS-induced inflammation. Microgliosis was measured by determining the percentage of Iba-1 positive area in the hippocampus. Data shown are mean ± SEM, ^* P<0.05, ^*** P<0.001 and were tested for statistical significance between treatment groups using ordinary one-way ANOVA with Dunnett's post hoc multiple comparison test. Dosing regimens are shown for three groups of C57BL/6 mice treated with vehicle control, vehicle-treated LPS, or Compound 1. The graph shows the results of histological analysis of mouse brains in an acute model of LPS-induced inflammation. Microgliosis was measured by determining the percentage of Iba-1 positive area in the hippocampus. Data shown are mean ± SEM, ^* P<0.05, ^*** P<0.0001, and statistical significance was tested between treatment groups using ordinary one-way ANOVA with Dunnett's post hoc multiple comparison test. 1 shows a treatment study timeline in the mouse MPTP model of Parkinson's disease. The study had two arms, the first arm tested gait and fine motor function after 10 days of treatment, and the second arm tested immune cell infiltration after 3 days of treatment. Correlation heatmap showing the degree of correlation for 97 gait parameters in gait and fine motor tests of C57Bl/6J mice after 10 days of Compound 1 treatment on day 11 of the test. Ten principal components are shown, showing how the original parameters are correlated in the data set. The stronger the intensity for each parameter, the stronger the parameter is related to the corresponding principal component. For the ten individual principal components (x-axis), red is positive correlation and blue is negative correlation. The left Y-axis shows the overall grouping of the individual variables on the right Y-axis. For example, "ILC" refers to intralimb orientation and "tail B" refers to tail base. Figure 1 shows the fine motor skills and gait characteristics of C57Bl/6J mice after 10 days of Compound 1 treatment, on study day 11. This is shown as the overall gait analysis score for MPTP-treated C57Bl/6J mice. Differences between treatment groups in each of the 10 major components were combined into a composite score that represents the difference from the vehicle-treated control group. There was a significant difference in overall gait with MPTP treatment ( ^* P<0.05), which was abolished by Compound 1 treatment. Figure 1 shows the forepaw toe clearance (one of the gait characteristics assessed by principal component analysis) of C57Bl/6J mice on study day 11, 10 days after treatment with Compound 1. C57Bl/6J mice were tested on study day 11, 10 days after treatment with Compound 1. Data are mean + SEM (Group 1: Vehicle + Vehicle, n = 15; Group 2: MPTP + Vehicle, n = 14; Group 3: MPTP + Compound 1 (30 mg/kg), n = 13). Statistical significance is ^* P<0.05, Group 2: MPTP + Vehicle vs. Group 1: Vehicle + Vehicle (paired t-test). Figure 1 shows the front paw swing velocity (one of the gait characteristics assessed by principal components analysis) of C57Bl/6J mice on study day 11 after 10 days of Compound 1 treatment. Data are mean + SEM (Group 1: Vehicle + Vehicle, n = 15; Group 2: MPTP + Vehicle, n = 14; Group 3: MPTP + Compound 1 (30 mg/kg), n = 13). Statistical significance is ^* P<0.05, Group 2: MPTP + Vehicle vs. Group 1: Vehicle + Vehicle (paired t-test). Figure 1 shows the range of ankle movement (one of the gait characteristics assessed by principal components analysis) of C57Bl/6J mice on study day 11 after 10 days of Compound 1 treatment. Data are mean + SEM (Group 1: Vehicle + Vehicle, n = 15; Group 2: MPTP + Vehicle, n = 14; Group 3: MPTP + Compound 1 (30 mg/kg), n = 13). Statistical significance is ^* P<0.05, Group 2: MPTP + Vehicle vs. Group 1: Vehicle + Vehicle (paired t-test). Figure 1 shows the acute effect of Compound 1 on T cell trafficking to the brain 3 days after MPTP and Compound 1 treatment. Total number of CD3 positive T cells counted in the substantia nigra from 30 μm sections per mouse is shown. Data shown are mean ± SEM, ^*** P<0.001, one-way ANOVA, Sidac® post hoc multiple comparison test. Figure 1 shows the acute effect of Compound 1 on microgliosis 3 days after MPTP and Compound 1 treatment, showing the extent of CD68 positive area measured in the striatum (n=7 for saline + vehicle, n=8 for MPTP + vehicle, n=7 for MPTP + Compound 1). Data shown are mean ± SEM, ^** P<0.01, ^*** P<0.001, ^*** P<0.0001, One-way ANOVA, Sidac post-hoc multiple comparison test. Figure 1 shows the acute effect of Compound 1 on microgliosis 3 days after MPTP and Compound 1 treatment, measured in the substantia nigra as the extent of CD68 positive area (n=7 for saline + vehicle, n=8 for MPTP + vehicle, n=8 for MPTP + Compound 1). Data shown are mean ± SEM, ^** P<0.01, ^*** P<0.001, ^*** P<0.0001, One-way ANOVA, Sidac post-hoc multiple comparison test. Figure 1 shows plasma eotaxin-1 levels in 6-month-old LINE61 transgenic mice overexpressing synuclein. Eotaxin-1 levels (CCL11) in non-transgenic (NTg) Line61 mice and transgenic Line61 synuclein mice (Tg) are plotted as pg/mL concentration. Figure 1 shows the results of wire suspension test in Line61 synuclein mice (age-matched littermates of transgenic (Tg) and non-transgenic (nTg)). Mean wire suspension time per group is shown, with animals in group C (non-transgenic, vehicle-treated) showing significantly higher wire suspension time. Animals in group A (transgenic, Compound-1-treated) showed significantly higher wire suspension time compared to group B (transgenic, vehicle-treated). Data are mean + SEM of all animals per group, ^*** P<0.001, Dunn's post-hoc test, ^* P<0.05 (shown as Mann-Whitney test for groups A and B). Figure 1 shows the results of grip strength testing in Line61 synuclein mice (transgenic (Tg) and non-transgenic (nTg) age-matched littermates). Mean maximum grip force [g] per group is shown. Groups A and C animals (transgenic Compound 1-treated and non-transgenic vehicle-treated, respectively) showed significantly higher grip force compared to group B (transgenic vehicle-treated). Data are presented as mean + standard error of all animals per group. Groups were compared to vehicle-treated transgenic animals (group B) using one-way ANOVA followed by Bonferroni post-hoc test. The number of mice from each treatment group that successfully traversed the beam in the beam-walk test is shown. N=15 mice in group A, 14 mice in group B, and 15 mice in group C (groups listed in FIG. 40). All mice were able to traverse the easiest beam in trial 1, but no mice in treatment group B were able to traverse the most difficult beam in trial 5 (trial 1=13 mm rectangular beam, trial 2=10 mm rectangular beam, trial 3=28 mm cylindrical beam, trial 4=16 mm cylindrical beam, trial 5=11 mm cylindrical beam). Results of 5 trials of beam-walk slips for 3 groups of mice (group A, transgenic compound 1-treated; group B, transgenic vehicle-treated; and group C, non-transgenic vehicle-treated) are shown. Only mice that completely traversed the beam were included in the analysis. Graphs represent the mean number of slips [n] per group, with each graph representing one trial (1-5). Data are the mean + standard error of all animals per group. Groups were compared with group B by one-way ANOVA followed by Bonferroni post-hoc test. Statistics for trial 5 could not be performed as mice in group B were unable to traverse the beam. Figure 1 shows the number of eosinophils from peripheral blood. The percentage of eosinophils (out of total white blood cells) in peripheral blood from three groups of mice (Tg Cmpd1 = transgenic treated with Compound 1, Tg Veh = transgenic treated with vehicle, nTGVeh = non-transgenic treated with vehicle) is shown. Data were compared by t-test. Figure 1 shows the number of eosinophils from peripheral blood. Absolute number of eosinophils in peripheral blood from three groups of mice (Tg Cmpd1 = transgenic treated with Compound 1, Tg Veh = transgenic treated with vehicle, nTGVeh = non-transgenic treated with vehicle). Data were compared by t-test. Figure 1 shows the effect of Compound 1 on neuroinflammation. Shown is the CD68 positive area quantified in the hippocampus of non-transgenic vehicle-treated mice, transgenic vehicle-treated mice, and transgenic Compound 1-treated mice (n=14, 12, and 15, respectively). Figure 1 shows the effect of Compound 1 on neuroinflammation. Shown is the CD68 positive area quantified in the striatum of non-transgenic vehicle-treated mice, transgenic vehicle-treated mice, and transgenic Compound 1-treated mice (n=15, 11, and 16, respectively). Figure 1 shows the effect of Compound 1 on neuroinflammation. Iba-1 positive area quantified in the hippocampus of non-transgenic vehicle-treated, transgenic vehicle-treated, and transgenic Compound 1-treated mice (n=14, 13, and 16, respectively). Figure 1 shows the effect of Compound 1 on neuroinflammation. Iba1 positive area quantified in the striatum of non-transgenic vehicle-treated, transgenic vehicle-treated, and transgenic Compound 1-treated mice (n=15, 11, and 16, respectively). Data are mean +/- SEM, ^* P<0.05. Figure 1 shows the effect of Compound 1 on neuroinflammation. GFAP-positive areas quantified in the hippocampus of non-transgenic vehicle-treated mice, transgenic vehicle-treated mice, and transgenic Compound 1-treated mice (n=14, 13, and 15, respectively) are shown. Figure 1 shows the effect of Compound 1 on neuroinflammation. GFAP-positive areas quantified in the striatum of non-transgenic vehicle-treated mice, transgenic vehicle-treated mice, and transgenic Compound 1-treated mice (n=15, 13, and 15, respectively) are shown. Figure 1 shows the effect of Compound 1 on neuroinflammation. Iba-1 positive area quantified in the substantia nigra pars compacta of non-transgenic vehicle-treated, transgenic vehicle-treated, and transgenic Compound 1-treated mice. Data are mean +/- SEM, ^* P<0.05. Figure 1 shows the effect of Compound 1 on circulating levels of IL-4 and IL-6 cytokines in non-transgenic vehicle-treated, transgenic vehicle-treated, and transgenic Compound 1-treated mice (n=14, 15, and 17, respectively). Levels of IL-4 measured in peripheral heart plasma from all three groups are shown. Data shown are mean ± SEM, ^* P<0.05, ^** P<0.01, one-way ANOVA, post-hoc Dunnett's multiple comparison test. Figure 1 shows the effect of Compound 1 on circulating levels of IL-4 and IL-6 cytokines in non-transgenic vehicle-treated, transgenic vehicle-treated, and transgenic Compound 1-treated mice (n=14, 15, and 17, respectively). Levels of IL-6 measured in peripheral heart plasma from all three groups are shown. Data shown are mean ± SEM, ^* P<0.05, ^** P<0.01, one-way ANOVA, post-hoc Dunnett's multiple comparison test. Figure 1 shows the effect of Compound 1 on EAE-induced markers in the cerebellum of C57BL/6 mice. EAE resulted in an increase in CD3 and CD8 positive infiltrating T cells in the cerebellum. CD3 positive infiltrating T cells in the cerebellum were shown to be increased and significantly decreased after 9 days of treatment with Compound 1. Data shown are mean ± SEM, ^* P<0.05, ^** P<0.01, one-way ANOVA, post-hoc Dunnett's multiple comparison test. Figure 1 shows the effect of Compound 1 on EAE-induced markers in the cerebellum of C57BL/6 mice. EAE resulted in an increase in CD3 and CD8 positive infiltrating T cells in the cerebellum. CD8 positive infiltrating T cells in the cerebellum were shown to be increased and significantly decreased after 9 days of treatment with Compound 1. Data shown are mean ± SEM, ^* P<0.05, ^** P<0.01, one-way ANOVA, post-hoc Dunnett's multiple comparison test. Figure 1 shows the effect of Compound 1 on EAE-induced markers in the cerebellum of C57BL/6 mice. EAE resulted in an increase in CD3 and CD8 positive infiltrating T cells in the cerebellum. A significant increase in Iba-1 positive area was shown in the cerebellum after EAE and was significantly decreased in the cerebellum after 9 days of treatment. Data shown are mean ± SEM, ^* P<0.05, ^** P<0.01, one-way ANOVA, post-hoc Dunnett's multiple comparison test. Figure 1 shows the effect of Compound 1 on EAE-induced markers in the cerebellum of C57BL/6 mice. EAE resulted in an increase in CD3 and CD8 positive infiltrating T cells in the cerebellum. A significant increase in CD68 positive area in the cerebellum after EAE was shown, which was significantly decreased in the cerebellum after 9 days of treatment. Data shown are mean ± SEM, ^* P<0.05, ^** P<0.01, one-way ANOVA, post-hoc Dunnett's multiple comparison test. Concentrations of human eotaxin-1 in the proteomic screen are shown. Relative human eotaxin-1 concentrations were measured with a commercial affinity-based assay (SomaLogic). Plasma samples from donors aged 18, 30, 45, 55 and 66 years were plotted. Figure 1 shows the effect of Compound 1 on inhibiting eosinophil morphological changes. Whole blood from humans treated with Compound 1 was incubated with recombinant eotaxin to induce eosinophil morphological changes. Percent inhibition of morphological changes was plotted against plasma concentration of Compound 1. 1 shows the effect of compound 1 on CCR3 internalization. To induce CCR3 internalization, whole blood from humans treated with compound 1 was incubated with recombinant eotaxin and labeled with anti-CCR3 antibody. Inhibition of CCR3 internalization by compound 1 was plotted against the plasma concentration of compound 1.

Aspects of the present invention include methods for treating age-related dysfunction/neurodegenerative diseases. Age-related dysfunction can manifest in many different ways, for example, as age-related cognitive and/or physiological dysfunction, including, but not limited to, cell injury, tissue damage, organ dysfunction, age-related shortening of life span, and carcinogenesis, in the form of damage to central or peripheral organs of the body (in which specific organs and tissues of interest include, but are not limited to, skin, neurons, muscle, pancreas, brain, kidney, lung, stomach, intestine, spleen, heart, adipose tissue, testes, ovaries, uterus, liver, and bone), reduced neurogenesis, etc.

In some embodiments, the age-related impairment is an age-related impairment in the cognitive abilities of an individual, i.e., age-related cognitive impairment. Cognitive ability or "cognition" refers to mental processes including attention and concentration, learning complex tasks and concepts, memory (acquiring, retaining and recalling new information in the short and/or long term), information processing (processing information collected by the senses), visuospatial functions (seeing, depth perception, using mental imagery, copying pictures, constructing objects or shapes), language production and comprehension, verbal fluency (finding words), problem solving, decision making, and executive functions (planning and prioritizing). "Cognitive decline" refers to the progressive decline of one or more of these abilities, e.g., memory, language, thinking, judgment, etc. "Cognitive impairment" and "cognitive impairment" refer to a decline in cognitive ability compared to a healthy individual, e.g., an age-matched healthy individual, or compared to the individual's ability at a previous time, e.g., 2 weeks, 1 month, 2 months, 3 months, 6 months, 1 year, 2 years, 5 years, 10 years or more ago. Age-related cognitive impairment includes cognitive impairments typically associated with aging, such as cognitive impairments associated with the natural aging process, e.g., mild cognitive impairment (MCI), and age-related disorders, i.e., disorders that are seen to increase in frequency with the progression of aging, e.g., cognitive impairments associated with neurodegenerative conditions such as Alzheimer's disease, Parkinson's disease, dementia with Lewy bodies, frontotemporal dementia, Huntington's disease, amyotrophic lateral sclerosis, multiple sclerosis, glaucoma, myotonic dystrophy, vascular dementia, etc.

"Treatment" refers to at least improving one or more symptoms associated with an age-related dysfunction from which the subject suffers, where improvement is used in a broad sense to refer to at least reducing the parameter, e.g., the severity of a symptom associated with the dysfunction being treated. Treatment thus also includes situations in which a pathological condition or at least symptoms associated with the condition are completely suppressed, e.g., their occurrence is prevented or inhibited, e.g., arrested, such that an adult mammal is free of the dysfunction or at least symptoms that characterize the dysfunction. In some cases, "treatment," "treating," and the like refer to obtaining a desired pharmacological and/or physiological effect. The effect may be a prophylactic effect, in terms of completely or partially preventing the disease or its symptoms, and/or a therapeutic effect, in terms of a partial or complete cure of the disease and/or the deleterious effects caused by the disease. "Treatment" can be any treatment of a disease in a subject, including (a) preventing the onset of the disease in a subject who may have a predisposition to the disease but has not yet been diagnosed with the disease, (b) inhibiting the disease, i.e., arresting the onset of the disease, or (c) relieving the disease, i.e., causing the disease to regress. Treatment can result in a wide variety of physical manifestations, such as modulating gene expression, increasing neurogenesis, rejuvenating tissues or organs, and the like. In some embodiments, treatment of ongoing disease is performed to stabilize or reduce undesirable clinical symptoms in the patient. Such treatment may occur before complete loss of function of the affected tissue. The therapy may be performed during the symptomatic phase of the disease, or in some cases, after the symptomatic phase of the disease.

In some cases where the age-related impairment is age-related cognitive decline, treatment with the disclosed method slows or reduces the progression of age-related cognitive decline. In other words, the cognitive ability of the individual, if any, declines after treatment with the disclosed method, but at a slower rate than before treatment with the disclosed method or in the absence of treatment with the disclosed method. In some cases, treatment with the disclosed method stabilizes the cognitive ability of the individual. For example, after treatment with the disclosed method, the progression of cognitive decline in an individual exhibiting age-related cognitive decline is halted. As another example, after treatment with the disclosed method, cognitive decline is prevented in an individual, for example, an individual over the age of 40 who is predicted to develop age-related cognitive decline. In other words, cognitive impairment (worsening cognitive impairment) is not observed. In some cases, treatment with the disclosed method reduces or reverses cognitive impairment, as observed, for example, by improvement in cognitive ability in an individual exhibiting age-related cognitive decline. In other words, after treatment with the disclosed methods, the cognitive ability of an individual exhibiting age-related cognitive decline is improved, i.e., improved by treatment, than before treatment with the disclosed methods. In some cases, treatment with the disclosed methods prevents cognitive impairment. In other words, after treatment with the disclosed methods, the cognitive ability of an individual exhibiting age-related cognitive decline is restored, e.g., to a level that the individual exhibiting age-related cognitive decline had when the individual was about 40 years old or younger, as evidenced by, e.g., improved cognitive ability of the individual exhibiting age-related cognitive decline.

In some cases where the age-related dysfunction is age-related motor dysfunction or motor decline, treatment with the disclosed method slows or reduces the progression of the age-related dysfunction or decline. In other words, even if the motor capacity of the individual declines after treatment with the disclosed method, the decline is slower than before treatment with the disclosed method or in the absence of treatment with the disclosed method. In some cases, treatment with the disclosed method stabilizes the motor capacity of the individual. For example, after treatment with the disclosed method, the progression of motor decline in an individual exhibiting age-related motor decline is halted. As another example, after treatment with the disclosed method, motor decline is prevented in an individual, for example, an individual over the age of 40 who is predicted to develop age-related motor decline. In other words, motor impairment (worsening of motor impairment) is not observed. In some cases, treatment with the disclosed methods reduces or reverses motor dysfunction, e.g., as observed by an improvement in motor coordination or motor performance of an individual exhibiting age-related motor decline. In other words, after treatment with the disclosed methods, the motor performance of an individual exhibiting age-related motor decline is better than before treatment with the disclosed methods, i.e., improved by treatment. In some cases, treatment with the disclosed methods prevents motor dysfunction. In other words, after treatment with the disclosed methods, e.g., as evidenced by an improvement in motor coordination or motor performance of an individual exhibiting age-related motor decline, e.g., the motor coordination or motor performance of the individual exhibiting age-related motor decline is restored to a level that the individual exhibiting age-related motor decline had when the individual was about 40 years old or younger.

In some cases, treatment of adult mammals according to the methods of the present invention results in changes in central nervous system organs, such as the brain, spinal cord, etc., which may be manifested in a wide variety of ways, including, but not limited to, molecular, structural and/or functional changes in the form of increased neurogenesis, e.g., as described in more detail below.

A method for treating dysfunction caused by a neurodegenerative disease is provided, comprising administering a compound of the formula discussed below. An embodiment of the invention includes a method for improving cognitive or motor activity in a subject with a brain-related, cognition-related or motor disease, comprising administering a therapeutically effective amount of a compound of the formula discussed below. An additional embodiment of the invention includes a method for increasing neurogenesis in a subject with a brain-related or cognition-related disease, comprising administering a therapeutically effective amount of a compound of the formula discussed below. An additional embodiment of the invention includes a method for alleviating or treating symptoms of a brain-related or cognition-related disease, comprising administering a therapeutically effective amount of a compound of the formula discussed below. An additional embodiment of the invention includes a method for alleviating symptoms of a central nervous system-related disease, comprising administering a therapeutically effective amount of a drug of the formula discussed below that acts primarily peripherally. Yet an additional embodiment of the invention includes a method for improving motor activity in a subject with age-related motor dysfunction, comprising administering a therapeutically effective amount of a compound of the formula discussed below. The methods of the invention may also include monitoring improvement of age-related diseases, including, for example, improvement in cognition, motor activity, neurogenesis, etc., in a subject diagnosed with one or more age-related diseases or dysfunctions.

Additional embodiments of the invention include administering a therapeutically effective amount of a compound, where the compound is in the form of a co-crystal or salt of the formula discussed below. Further embodiments of the invention include administering a therapeutically effective amount of a compound, where the compound is in the form of an individual optical isomer, a mixture of individual enantiomers, a racemate, or an enantiomerically pure compound. Additional embodiments of the invention also include administering a therapeutically effective amount of a compound, where the compound is in the form of a pharmaceutical composition and pharmaceutical formulation, discussed further below.

Additional embodiments of the present invention for treating age-related motor dysfunction or motor decline include modifiers that inhibit the eotaxin/CCR3 pathway. Such modifiers include compounds of the formulas discussed below, as well as other eotaxin inhibitors and CCR3 inhibitors. Contemplated modifiers include, by way of example and without limitation, compounds of the formulas discussed below, as well as other CCR3 small molecule inhibitors (e.g., bipiperidine derivatives, e.g., as described in U.S. Pat. No. 7,705,153, cyclic amine derivatives, e.g., as described in U.S. Pat. No. 7,576,117, and cyclic amine derivatives, e.g., as described in Pease JE and Horuk R, Expert Opin Drug Discov (2014) 9(5):467-83 (all of which are incorporated herein by reference in their entirety), anti-eotaxin antibodies, anti-CCR3 antibodies, aptamers that inhibit either the expression or function of eotaxin or CCR3 (methods for producing such aptamers include U.S. Pat. Nos. 5,270,163, 5,840,867, and 6,180,348), antisense oligonucleotides or siRNAs that inhibit either the expression or function of eotaxin or CCR3 (e.g., those described in U.S. Pat. No. 6,822,087), and soluble CCR3 receptor proteins (e.g., decoys).

Methods of using diagnostic tests or companion diagnostic tests in connection with the described age-related disorders are also provided. One embodiment of such a diagnostic test or companion diagnostic test is an in vitro diagnostic test. An embodiment of an in vitro diagnostic test is a companion device used in conjunction with a particular therapeutic agent. Particular therapeutic agent embodiments include, but are not limited to, for example, CCR3 small molecule inhibitors, anti-CCR3 antibodies, small molecule inhibitors against the CCR3 ligand eotaxin-1, anti-eotaxin-1 antibodies, and antisense RNA against either eotaxin-1 or CCR3.

By way of example, such a diagnosis or companion diagnostic includes, but is not limited to, determining or detecting the presence of a subset of white blood cells from a subject. A diagnosis or companion diagnostic may also be the determination of the presence, relative or absolute concentration, relative or absolute number of eosinophils from the subject's blood, tissue, or other such sample. Blood may be obtained, for example, by venipuncture or other similar methods. Other samples include, by way of example, but are not limited to, sputum, cerebrospinal fluid, or tissue biopsy.

Methods for detecting or determining the presence, absolute or relative concentration, or relative or absolute number of white blood cells, such as eosinophils, neutrophils, lymphocytes, basophils, and monocytes, are known to those of skill in the art. Methods for determining the presence, absolute or relative concentration, or relative or absolute number of eosinophils are also known to those of skill in the art. See Walsh GM, Eosinophils-Methods and Protocols ISBN: 9781493910151, which is incorporated herein by reference in its entirety. Such methods include, by way of example and without limitation, (1) a five-part complete blood count using a hematology analyzer (see, e.g., O'Neil et al., Laboratory Hematology 7:116-124 (2001) and The U.S. Centers for Disease Control laboratory, Journal of Clinical Oncology 2002, 14:131-132 (2002)). (2) quantification of eosinophil counts in sputum using hematoxylin and eosin (H and E) stained smear preparations (see, e.g., Bandyopadhyay A, et al. Lung India. 2013 Apr-Jun; 30 (2):117-123); (3) CD45, CD125, CD193, F4/80, and Siglec-8 (Abcam, San Diego, CA, USA); (4) flow cytometry quantification of eosinophils using antibodies specific for various immune cells such as eosinophils (see, e.g., Yu Y-R A et al., Am J Respir Cell Mol Biol. 2016 Jan; 54 (1):13-24 and Cossarizza A, et al., Eur. J. Immunol. 2017 47:1584-1797), (5) eosinophil autofluorescence quantified using flow cytometry (see, e.g., Guenther G et al., J Immunol May 1, 2015, 194 (1 Supplement) 206.12, and Thurau A M et al. Cytometry 23:159-58 (1996 )); (5) isolation of eosinophils from whole blood using dextran 70, Ficoll-Paque, and antibody-based magnetic colloid cell separation (see, e.g., Munoz NM, et al., Nature Protocols, 2613-20 (2006); Akuthota P, et al., Curr Protoc Immunol 98 (1):pp. 7.31.1-7.31.8, 2012; and Akuthota P, et al. Methods Mol Biol 1178:13-20 (2014)); (6) histological staining (see, e.g., Koller DY et al., Allergy 54 (10):1094-9 (1999), and Akuthota P., Capron K., Weller P.F. (2014) Eosinophil Purification from Peripheral Blood Cells, 1999) by general staining (eosin or similar dye) or by antibody-based staining for eosinophil-specific proteins (major basic protein (MBP), eosinophil cationic protein (ECP), eosinophil peroxidase (EPO), eosinophil-derived neurotoxin (EDN), Siglec F (or Siglec 8) among others). Blood. In: Walsh G. (eds) Eosinophils. Methods in Molecular Biology (Methods and Protocols), vol 1178. Humana Press, New York, NY, etc.), (7) histological staining of processes spatially associated with eosinophils, such as degranulation, eosinophil extracellular traps (EETs), and/or specific proteins therein (e.g., galectin 10).

A diagnostic or companion diagnostic device may incorporate such methods to determine the presence, absolute or relative concentration, or relative or absolute number of eosinophils. By determining the presence, absolute or relative concentration, or relative or absolute number of eosinophils, the device may be used with other methods of the invention. By way of example and without limitation, such other methods may include methods of treating age-related dysfunction/neurodegenerative diseases as described herein. In some embodiments, the age-related dysfunction is an age-related dysfunction in the cognitive abilities of an individual, i.e., an age-related dysfunction. Such age-related dysfunction may include, by way of example and without limitation, neurodegenerative conditions such as Alzheimer's disease, Parkinson's disease, Lewy body dementia, frontotemporal dementia, Huntington's disease, amyotrophic lateral sclerosis, multiple sclerosis, glaucoma, myotonic dystrophy, vascular dementia, etc. Such age-related dysfunction may include cognitive and/or motor dysfunction or decline.

An embodiment of the invention includes treating a subject diagnosed with age-related dysfunction with a therapeutically effective amount of at least one of the compounds of the invention disclosed herein in combination with a diagnostic or companion diagnostic device. Another embodiment of the invention includes administering a therapeutically effective amount of compound 1 disclosed herein in combination with a diagnostic device. Such an embodiment of the device can be used, for example, to determine or detect the presence, absolute or relative concentration, or absolute or relative number of eosinophils in a blood or tissue sample of the subject. Another embodiment performs a step of detecting or determining the presence, absolute or relative concentration, or absolute or relative number of eosinophils in a blood or tissue sample of the subject before, during, or after treatment.

Experimental figures herein show that a mammalian model of Parkinson's disease with synacrein overexpression results in a reduction in eosinophils that are restored to levels in non-transgenic mice with Compound 1 treatment, suggesting beneficial immune modulation in this model of Parkinson's disease. This also suggests that determining eosinophil levels in Parkinson's disease patients treated with Compound 1 may be a biomarker in the disease, including determining the level of disease progression, stagnation, or regression, as well as treatment efficacy (see, e.g., Figures 44A and 44B).

Another embodiment performs a step of detecting or determining the presence, absolute or relative concentration, or absolute or relative number of eosinophils in a blood or tissue sample of a subject diagnosed with Parkinson's disease before, during, or after treatment. A further embodiment includes determining the presence, absolute or relative concentration, or absolute or relative number of eosinophils in a blood or tissue sample of a subject diagnosed with Parkinson's disease before being treated with a compound of the invention, such as Compound 1, to obtain a baseline concentration or number. A further embodiment then performs a step of detecting or determining the level of eosinophils after treatment and comparing such level with the baseline concentration or number of eosinophils to monitor the effectiveness of the treatment. Such a comparison of levels may indicate an increase or decrease in the number or concentration of eosinophils compared to the baseline. Another embodiment of the invention includes a step of monitoring the progression of the disease or dysfunction by comparing the number or concentration of eosinophils, where if the number increases after treatment, there is an improvement in the progression of the disease. In another embodiment, the improvement is a 1-5% increase, a 5-10% increase, an 11-15% increase, a 16-20% increase, a 21-25% increase, a 26-30% increase, a 31-35% increase, a 36-40% increase, a 41-45% increase, a 46-50% increase, a 51-55% increase, a 56-60% increase, a 61-65% increase, a 66-70% increase, a 71-75% increase, a 76-80% increase over baseline in blood or tissue eosinophil count or concentration in the subject. may include an increase of 81-85%, an increase of 86-90%, an increase of 91-95%, an increase of 96-100%, as well as 1-1.5x, 1.5-2x, 2-2.5x, 2.5-3x, 3-3.5x, 3.5-4x, 4-4.5x, 4.5-5x, 5-5.5x, 5.5-6x, 6-6.5x, 6.5-7x, 7-7.5x, 7.5-8x, 8-8.5x, 8.5-9x, 9-9.5x, 9.5-10x increase, and an increase of more than 10x.

Another embodiment of the present invention includes diagnosing Parkinson's disease by determining a baseline in the number or concentration of eosinophils in the blood or tissues of a subject and comparing it to a standard number or concentration of eosinophils in a population without Parkinson's disease that has a normal eosinophil count. Eosinophil count is known in the art to be the number of eosinophils in the body. In adults, the eosinophil count is between 30 and 350, but can be up to 500 cubic millimeters (mm ³ ) in the blood. (See Medical News Today, Dec 2018, available at https://www.medicalnewstoday.com/articles/323868.php, which is incorporated herein by reference in its entirety.) A count of more than 500 mm ³ in the blood is considered eosinophilia. Lower than normal eosinophil counts occur in several diseases, such as alcoholism and overprotection of cortisol (ibid.). Aspects of the present invention detect or determine the number or concentration of eosinophils in a subject suspected of having Parkinson's disease. If the number or concentration of eosinophils in a subject is lower than normal (e.g., 30 ^mm3 in blood), the results can be used by caregivers to determine a diagnosis of the disease.

a. Compounds The methods of the invention further include administration of the following compounds to a subject: In the groups, radicals or moieties defined in this Compounds section, the number of carbon atoms is often specified preceding the group, e.g., C _1-6 alkyl means an alkyl group or alkyl radical having 1 to 6 carbon atoms. Generally, in groups disclosed in this Compounds section that contain more than one subgroup, the last listed group is the point of attachment of the radical, e.g., "thioalkyl" means a monovalent radical of the formula HS-Alk-. Unless otherwise specified below, in all formulas and groups, conventional definitions of terms apply and conventional stable atom valences are presumed and achieved.

Embodiments of the invention further include administering to a subject a compound of formula 1,

In the formula,
A is CH ₂ , O or N—C _1-6 alkyl;
^R1 is
・NHR ^1.1 , NMeR ^1.1 ,
・NHR ^1.2 , NMeR ^1.2 ,
・NHCH ₂ -R ^1.3 ,
NH-C _3-6 cycloalkyl, optionally one carbon atom of which is replaced by a nitrogen atom and the ring is optionally substituted with one or two residues selected from the group consisting of C _1-6 alkyl, O-C _1-6 alkyl, NHSO ₂ -phenyl, NHCONH _- phenyl, halogen, CN, SO ₂ -C _1-6 alkyl, COO-C _1-6 alkyl,
a C _9,10 bicycle, one or two carbon atoms of which are replaced by a nitrogen atom, the ring system being linked to the basic structure of formula 1 via the nitrogen atom, the ring system being optionally substituted with one or two residues selected from the group consisting of C _1-6 alkyl, COO-C _1-6 alkyl, C _1-6 haloalkyl, O-C _1-6 alkyl, NO ₂ , halogen, CN, NHSO _{2 -C 1-6} alkyl, methoxyphenyl,
a group selected from NHCH(pyridinyl)CH ₂ COO—C _1-6 alkyl, NHCH(CH ₂ O—C _1-6 alkyl)-benzimidazolyl, optionally substituted with halogen or CN, or 1-aminocyclopentyl, optionally substituted with methyl-oxadiazole,
R ^1.1 is phenyl, optionally C _1-6 alkyl, C _2-6 alkenyl, C _2-6 alkynyl, C _1-6 haloalkyl, C _1-6 alkylene-OH, C 2-6 alkenylene- _{OH, C 2-6 alkynylene-OH, CH 2 CON(C 1-6 alkyl) 2 , CH 2 NHCONH-C 3-6 cycloalkyl , CN , CO -} pyridinyl, CONR ^1.1.1 R ^1.1.2 , COO-C _1-6 alkyl, N(SO ₂ -C _1-6 alkyl)(CH ₂ CON(C _1-4 alkyl) ₂ ), O-C _1-6 alkyl, O-pyridinyl, SO ₂ -C _1-6 alkyl, SO ₂ -C _1-6 alkylene-OH, SO ₂ -C _{3-6cycloalkyl} , SO ₂ -piperidinyl, SO ₂ NH-C _1-6alkyl , SO ₂ N(C _1-6alkyl ) ₂ , halogen, CN, CO-morpholinyl, CH ₂ -pyridinyl or a heterocycle optionally substituted with 1 or 2 residues selected from the group consisting of C _1-6alkyl , NHC _1-6alkyl and ═O,
R ^1.1.1 is H, C _1-6 alkyl, C _3-6 cycloalkyl, C _1-6 haloalkyl, CH ₂ CON(C _1-6 alkyl) ₂ , CH ₂ CO-azetindinyl, C _1-6 alkylene-C _3-6 cycloalkyl, CH ₂ -pyranyl, CH ₂ -tetrahydrofuranyl, CH ₂ -furanyl, C _1-6 alkylene-OH or thiadiazolyl, optionally substituted with C _1-6 alkyl;
R ^1.1.2 is H, C _1-6 alkyl, SO ₂ C _1-6 alkyl;
or R ^1.1.1 and R ^1.1.2 together form a 4-, 5- or 6-membered carbocyclic ring, which optionally contains one N or O, replacing a carbon atom of the ring, and optionally substituted with one or two residues selected from the group consisting of C _1-6 alkyl, C _1-4 alkylene-OH, OH, ═O;
or,
R ^1.1 is phenyl, two adjacent residues of which together form a 5- or 6-membered carbocyclic aromatic or non-aromatic ring, which ring optionally contains, independently of each other, one or two N, S or SO ₂ , replacing a carbon atom of the ring, which ring is optionally substituted with C _1-4 alkyl or ═O;
R ^1.2 is
heteroaryl optionally substituted with one or two residues selected from the group consisting of C _1-6 alkyl, C _2-6 alkenyl, C _2-6 alkynyl, C _3-6 cycloalkyl, CH ₂ COO-C _1-6 alkyl, CONR ^1.2.1 R ^1.2.2 , COR ^1.2.3 , COO-C _1-6 alkyl, CONH ₂ , O-C _1-6 alkyl, halogen, CN, SO ₂ N(C _1-6 alkyl) ₂ , or heteroaryl optionally substituted with one or two residues selected from the group consisting of C _1-6 alkyl,
Heteroaryl optionally substituted with a 5- or 6-membered carbocyclic non-aromatic ring, which ring contains, independently of each other, two N, O, S or _SO2 , replacing the carbon atoms of the ring;
- aromatic or non-aromatic C _9,10 bicycles, one or two of whose carbon atoms are replaced by N, O or S, each optionally substituted by one or two residues selected from the group consisting of N(C _1-6 alkyl) ₂ , CONH-C _{1-6 alkyl} , ═O;
- optionally pyridinyl-substituted heterocyclic non-aromatic rings,
4,5-dihydro-naphtho[2,1-d]thiazole optionally substituted with NHCO-C _1-6 alkyl,
R ^1.2.1 is H, C _1-6 alkyl, C _1-6 alkylene-C _3-6 cycloalkyl, C _1-4 alkylene-phenyl, C _1-4 alkylene-furanyl, C _3-6 cycloalkyl, C _1-4 alkylene-O-C _1-4 alkyl, C _1-6 haloalkyl, or a 5- or 6-membered carbocyclic non-aromatic ring which optionally contains, independently of each other, one or two N, O, S or SO ₂ , the ring carbon atoms of which are replaced and which is optionally substituted with 4-cyclopropylmethyl-piperazinyl;
R ^1.2.2 is H, C _1-6 alkyl;
^R1.2.3 is a 5- or 6-membered carbocyclic non-aromatic ring, which ring optionally contains, independently of each other, one or two N, O, S or _SO2 , replacing the carbon atoms of the ring;
R ^1.3 is selected from phenyl, heteroaryl or indolyl, each optionally substituted with one or two residues selected from the group consisting of C _1-6 alkyl, C _3-6 cycloalkyl, O-C _1-6 alkyl, O-C _1-6 haloalkyl, phenyl, heteroaryl;
R ² is selected from the group consisting of C _1-6 alkylene-phenyl, C _1-6 alkylene-naphthyl and C _1-6 alkylene-heteroaryl, each of which is optionally substituted with 1, 2 or 3 residues selected from the group consisting of C _1-6 alkyl, C _1-6 haloalkyl, O-C _1-6 alkyl, O-C _1-6 haloalkyl, halogen;
^R3 is H, _C1-6 alkyl;
R ⁴ is H, C _1-6 alkyl, or
Or R ³ and R ⁴ together form a CH ₂ -CH ₂ group.

Another embodiment of the present invention further comprises a compound of formula 1 (above):
In the formula,
A is CH ₂ , O or N—C _1-4 alkyl;
^R1 is
・NHR ^1.1 , NMeR ^1.1 ,
・NHR ^1.2 , NMeR ^1.2 ,
・NHCH ₂ -R ^1.3
is selected from
R ^1.1 is phenyl, optionally C _1-6 alkyl, C _2-6 alkenyl, C _2-6 alkynyl, C _1-6 haloalkyl, C _1-6 alkylene-OH, C 2-6 alkenylene- _{OH, C 2-6 alkynylene-OH, CH 2 CON(C 1-6 alkyl) 2 , CH 2 NHCONH-C 3-6 cycloalkyl , CN , CO -} pyridinyl, CONR ^1.1.1 R ^1.1.2 , COO-C _1-6 alkyl, N(SO ₂ -C _1-6 alkyl)(CH ₂ CON(C _1-4 alkyl) ₂ ), O-C _1-6 alkyl, O-pyridinyl, SO ₂ -C _1-6 alkyl, SO ₂ -C _1-6 alkylene-OH, SO ₂ -C _{3-6cycloalkyl} , SO ₂ -piperidinyl, SO ₂ NH-C _1-6alkyl , SO ₂ N(C _1-6alkyl ) ₂ , halogen, CN, CO-morpholinyl, CH ₂ -pyridinyl or a heterocycle optionally substituted with 1 or 2 residues selected from the group consisting of C _1-6alkyl , NHC _1-6alkyl , ═O,
R ^1.1.1 is H, C _1-6 alkyl, C _3-6 cycloalkyl, C _1-6 haloalkyl, CH ₂ CON(C _1-6 alkyl) ₂ , CH ₂ CO-azetindinyl, C _1-6 alkylene-C _3-6 cycloalkyl, CH ₂ -pyranyl, CH ₂ -tetrahydrofuranyl, CH ₂ -furanyl, C _1-6 alkylene-OH or thiadiazolyl, optionally substituted with C _1-6 alkyl;
R ^1.1.2 is H, C _1-6 alkyl, SO ₂ C _1-6 alkyl;
or R ^1.1.1 and R ^1.1.2 together form a 4-, 5- or 6-membered carbocyclic ring, which optionally contains one N or O, replacing a carbon atom of the ring, and optionally substituted with one or two residues selected from the group consisting of C _1-6 alkyl, C _1-4 alkylene-OH, OH, ═O;
or,
R ^1.1 is phenyl, two adjacent residues of which together form a 5- or 6-membered carbocyclic aromatic or non-aromatic ring, which ring optionally contains, independently of each other, one or two N, S or SO ₂ , replacing a carbon atom of the ring, which ring is optionally substituted with C _1-4 alkyl or ═O;
R ^1.2 is
heteroaryl optionally substituted with one or two residues selected from the group consisting of C _1-6 alkyl, C _2-6 alkenyl, C _2-6 alkynyl, C _3-6 cycloalkyl, CH ₂ COO-C _1-6 alkyl, CONR ^1.2.1 R ^1.2.2 , COR ^1.2.3 , COO-C _1-6 alkyl, CONH ₂ , O-C _1-6 alkyl, halogen, CN, SO ₂ N(C _1-4 alkyl) ₂ , or heteroaryl optionally substituted with one or two residues selected from the group consisting of C _1-6 alkyl,
Heteroaryl optionally substituted with a 5- or 6-membered carbocyclic non-aromatic ring, which ring contains, independently of each other, two N, O, S or _SO2 , replacing the carbon atoms of the ring,
R ^1.2.1 is H, C _1-6 alkyl, C _1-6 alkylene-C _3-6 cycloalkyl, C _1-4 alkylene-phenyl, C _1-4 alkylene-furanyl, C _3-6 cycloalkyl, C _1-4 alkylene-O-C _1-4 alkyl, C _1-6 haloalkyl, or a 5- or 6-membered carbocyclic non-aromatic ring which optionally contains, independently of each other, 1 or 2 N, O, S or SO ₂ , which carbon atoms of the ring are replaced, and which is optionally substituted with 4-cyclopropylmethyl-piperazinyl;
R ^1.2.2 is H, C _1-6 alkyl;
^R1.2.3 is a 5- or 6-membered carbocyclic non-aromatic ring, which ring optionally contains, independently of each other, one or two N, O, S or _SO2 , replacing the carbon atoms of the ring;
R ^1.3 is selected from phenyl, heteroaryl, or indolyl, each optionally substituted with one or two residues selected from the group consisting of C _1-6 alkyl, C _3-6 cycloalkyl, O-C _1-6 alkyl, O-C _1-6 haloalkyl, phenyl, heteroaryl; and in some examples R ^1.3 is selected from phenyl, pyrazolyl, isoxazolyl, pyridinyl, pyrimidinyl, indolyl, or oxadiazolyl, each optionally substituted with one or two residues selected from the group consisting of C _1-6 alkyl, C _3-6 cycloalkyl, O-C _1-6 alkyl, O-C _1-6 haloalkyl, phenyl, pyrrolidinyl;
R ² is selected from the group consisting of C _1-6 alkylene-phenyl, C _1-6 alkylene-naphthyl and C _1-6 alkylene-thiophenyl, each of which is optionally substituted with 1, 2 or 3 residues selected from the group consisting of C _1-6 alkyl, C _1-6 haloalkyl, O-C _1-6 alkyl, O-C _1-6 haloalkyl, halogen;
^R3 is H, _C1-4 alkyl;
R ⁴ is H, C _1-4 alkyl, or R ³ and R ⁴ together form a CH ₂ -CH ₂ group;
The method includes administering the compound to a subject.

Another embodiment of the present invention further comprises a compound of formula 1 (above):
In the formula,
A is CH ₂ , O or N—C _1-4 alkyl;
^R1 is
selected from NHR ^1.1 , NMeR ^1.1 ,
R ^1.1 is phenyl, optionally C _1-6 alkyl, C _2-6 alkenyl, C _2-6 alkynyl, C _1-6 haloalkyl, C _1-6 alkylene-OH, C 2-6 alkenylene- _{OH, C 2-6 alkynylene-OH, CH 2 CON(C 1-6 alkyl) 2 , CH 2 NHCONH-C 3-6 cycloalkyl , CN , CO -} pyridinyl, CONR ^1.1.1 R ^1.1.2 , COO-C _1-6 alkyl, N(SO ₂ -C _1-6 alkyl)(CH ₂ CON(C _1-4 alkyl) ₂ ), O-C _1-6 alkyl, O-pyridinyl, SO ₂ -C _1-6 alkyl, SO ₂ -C _1-6 alkylene-OH, SO ₂ -C _{3-6cycloalkyl} , SO ₂ -piperidinyl, SO ₂ NH-C _1-6alkyl , SO ₂ N(C _1-6alkyl ) ₂ , halogen, CN, CO-morpholinyl, CH ₂ -pyridinyl or a heterocycle optionally substituted with 1 or 2 residues selected from the group consisting of C _1-6alkyl , NHC _1-6alkyl , ═O,
R ^1.1.1 is H, C _1-6 alkyl, C _3-6 cycloalkyl, C _1-6 haloalkyl, CH ₂ CON(C _1-6 alkyl) ₂ , CH ₂ CO-azetindinyl, C _1-6 alkylene-C _3-6 cycloalkyl, CH ₂ -pyranyl, CH ₂ -tetrahydrofuranyl, CH ₂ -furanyl, C _1-6 alkylene-OH or thiadiazolyl, optionally substituted with C _1-6 alkyl;
R ^1.1.2 is H, C _1-6 alkyl, SO ₂ C _1-6 alkyl, or R ^1.1.1 and R ^1.1.2 together form a 4-, 5- or 6-membered carbocyclic ring optionally containing one N or O replacing a carbon atom of the ring and optionally substituted with one or two residues selected from the group consisting of C _1-6 alkyl, C _1-4 alkylene-OH, OH, ═O;
or,
R ^1.1 is phenyl, two adjacent residues of which together form a 5- or 6-membered carbocyclic aromatic or non-aromatic ring, which optionally contains, independently of each other, one or two N, S or SO ₂ , replacing the carbon atoms of the ring, which is optionally substituted with C _1-4 alkyl or ═O;
R ² is selected from the group consisting of C _1-6 alkylene-phenyl, C _1-6 alkylene-naphthyl and C _1-6 alkylene-thiophenyl, each of which is optionally substituted with 1, 2 or 3 residues selected from the group consisting of C _1-6 alkyl, C _1-6 haloalkyl, O-C _1-6 alkyl, O-C _1-6 haloalkyl, halogen;
^R3 is H, _C1-4 alkyl;
R ⁴ is H, C _1-4 alkyl, or R ³ and R ⁴ together form a CH ₂ -CH ₂ group;
The method includes administering the compound to a subject.

Another embodiment of the present invention further comprises a compound of formula 1,
In the formula,
A is CH ₂ , O or N—C _1-4 alkyl;
^R1 is
selected from NHR ^1.2 , NMeR ^1.2 ,
R ^1.2 is
heteroaryl optionally substituted with one or two residues selected from the group consisting of C _1-6 alkyl, C _2-6 alkenyl, C _2-6 alkynyl, C _3-6 cycloalkyl, CH ₂ COO-C _1-6 alkyl, CONR ^1.2.1 R ^1.2.2 , COR ^1.2.3 , COO-C _1-6 alkyl, CONH ₂ , O-C _1-6 alkyl, halogen, CN, SO ₂ N(C _1-4 alkyl) ₂ , or heteroaryl optionally substituted with one or two residues selected from the group consisting of C _1-6 alkyl,
Heteroaryl optionally substituted with a 5- or 6-membered carbocyclic non-aromatic ring, which ring contains, independently of each other, two N, O, S or _SO2 , replacing the carbon atoms of the ring;
- benzothiazolyl, indazolyl, dihydroindolyl, indanyl, tetrahydroquinolinyl, each of which is optionally substituted with one or two residues selected from the group consisting of N(C _1-6 alkyl) ₂ , CONH-C _1-6 alkyl, ═O;
piperidinyl optionally substituted with pyridinyl,
4,5-dihydro-naphtho[2,1-d]thiazole optionally substituted with NHCO-C _1-6 alkyl,
R ^1.2.1 is H, C _1-6 alkyl, C _1-6 alkylene-C _3-6 cycloalkyl, C _1-4 alkylene-phenyl, C _1-4 alkylene-furanyl, C _3-6 cycloalkyl, C _1-4 alkylene-O-C _1-4 alkyl, C _1-6 haloalkyl, or a 5- or 6-membered carbocyclic non-aromatic ring which optionally contains, independently of each other, one or two N, O, S or SO ₂ , the ring carbon atoms of which are replaced and which is optionally substituted with 4-cyclopropylmethyl-piperazinyl;
R ^1.2.2 is H, C _1-6 alkyl;
^R1.2.3 is a 5- or 6-membered carbocyclic non-aromatic ring, optionally containing, independently of each other, one or two N, O, S or _SO2 , replacing a carbon atom of the ring;
R ² is selected from the group consisting of C _1-6 alkylene-phenyl, C _1-6 alkylene-naphthyl and C _1-6 alkylene-thiophenyl, each of which is optionally substituted with 1, 2 or 3 residues selected from the group consisting of C _1-6 alkyl, C _1-6 haloalkyl, O-C _1-6 alkyl, O-C _1-6 haloalkyl, halogen;
^R3 is H, _C1-4 alkyl;
R ⁴ is H, C _1-4 alkyl, or R ³ and R ⁴ together form a CH ₂ -CH ₂ group;
The method includes administering the compound to a subject.

Another embodiment of the present invention further comprises a compound of formula 1 (above):
In the formula,
A is CH ₂ , O or N—C _1-4 alkyl;
^R1 is
selected from NHR ^1.2 , NMeR ^1.2 ,
R ^1.2 is
heteroaryl optionally substituted with one or two residues selected from the group consisting of C _1-6 alkyl, C _2-6 alkenyl, C _2-6 alkynyl, C _3-6 cycloalkyl, CH ₂ COO-C _1-6 alkyl, CONR ^1.2.1 R ^1.2.2 , COR ^1.2.3 , COO-C _1-6 alkyl, CONH ₂ , O-C _1-6 alkyl, halogen, CN, SO ₂ N(C _1-4 alkyl) ₂ , or heteroaryl optionally substituted with one or two residues selected from the group consisting of C _1-6 alkyl,
Heteroaryl optionally substituted with a 5- or 6-membered carbocyclic non-aromatic ring, which ring contains, independently of each other, two N, O, S or _SO2 , replacing the carbon atoms of the ring,
R ^1.2.1 is H, C _1-6 alkyl, C _1-6 alkylene-C _3-6 cycloalkyl, C _1-4 alkylene-phenyl, C _1-4 alkylene-furanyl, C _3-6 cycloalkyl, C _1-4 alkylene-O-C _1-4 alkyl, C _1-6 haloalkyl, or a 5- or 6-membered carbocyclic non-aromatic ring which optionally contains, independently of each other, 1 or 2 N, O, S or SO ₂ , which carbon atoms of the ring are replaced, and which is optionally substituted with 4-cyclopropylmethyl-piperazinyl;
R ^1.2.2 is H, C _1-6 alkyl;
^R1.2.3 is a 5- or 6-membered carbocyclic non-aromatic ring, optionally containing, independently of each other, one or two N, O, S or _SO2 , replacing a carbon atom of the ring;
R ² is selected from the group consisting of C _1-6 alkylene-phenyl, C _1-6 alkylene-naphthyl and C _1-6 alkylene-thiophenyl, each of which is optionally substituted with 1, 2 or 3 residues selected from the group consisting of C _1-6 alkyl, C _1-6 haloalkyl, O-C _1-6 alkyl, O-C _1-6 haloalkyl, halogen;
^R3 is H, _C1-4 alkyl;
R ⁴ is H, C _1-4 alkyl, or R ³ and R ⁴ together form a CH ₂ -CH ₂ group;
The method includes administering the compound to a subject.

Another embodiment of the present invention further comprises a compound of formula 1,
In the formula,
A is CH ₂ , O or N—C _1-4 alkyl;
^R1 is
- selected from NHCH ₂ -R ^1.3 ;
R ^1.3 is selected from phenyl, pyrazolyl, isoxazolyl, pyridinyl, pyrimidinyl, indolyl or oxadiazolyl, each optionally substituted with one or two residues selected from the group consisting of C _1-6 alkyl, C _3-6 cycloalkyl, O-C _1-6 alkyl, O-C _1-6 haloalkyl, phenyl, pyrrolidinyl;
R ² is selected from the group consisting of C _1-6 alkylene-phenyl, C _1-6 alkylene-naphthyl and C _1-6 alkylene-thiophenyl, each of which is optionally substituted with 1, 2 or 3 residues selected from the group consisting of C _1-6 alkyl, C _1-6 haloalkyl, O-C _1-6 alkyl, O-C _1-6 haloalkyl, halogen;
^R3 is H, _C1-4 alkyl;
R ⁴ is H, C _1-4 alkyl, or R ³ and R ⁴ together form a CH ₂ -CH ₂ group;
The method includes administering a compound to a subject.

Another embodiment of the present invention further comprises a compound of formula 1,
In the formula,
A is CH ₂ , O or N—C _1-4 alkyl;
^R1 is
・NHR ^1.1 , NMeR ^1.1 ,
・NHR ^1.2 , NMeR ^1.2 ,
・NHCH ₂ -R ^1.3 ,
NH-C _3-6 cycloalkyl, optionally one carbon atom of which is replaced by a nitrogen atom and the ring is optionally substituted with one or two residues selected from the group consisting of C _1-6 alkyl, O-C _1-6 alkyl, NHSO ₂ -phenyl, NHCONH _- phenyl, halogen, CN, SO ₂ -C _1-6 alkyl, COO-C _1-6 alkyl,
a C _9,10 bicyclic ring, one or two carbon atoms of which are replaced by a nitrogen atom, the ring system being linked to the basic structure of formula 1 via the nitrogen atom, the ring system being optionally substituted with one or two residues selected from the group consisting of C _1-6 alkyl, COO-C _1-6 alkyl, C _1-6 haloalkyl, O-C _1-6 alkyl, NO ₂ , halogen, CN, NHSO ₂ -C _1-6 alkyl, m _- methoxyphenyl,
a radical selected from NHCH(pyridinyl)CH ₂ COO—C _1-6 alkyl, NHCH(CH ₂ O—C _1-6 alkyl)-benzimidazolyl, optionally substituted with Cl, or a radical selected from 1-aminocyclopentyl, optionally substituted with methyloxadiazolyl,
R ^1.1 is phenyl, optionally C _1-6 alkyl, C _1-6 haloalkyl, CH ₂ CON(C _1-6 alkyl) ₂ , CH ₂ NHCONH-C _3-6 cycloalkyl, CN, CONR ^1.1.1 R ^1.1.2 , COO-C _1-6 alkyl, O-C _1-6 alkyl, SO ₂ -C _1-6 alkyl, SO ₂ -C _1-6 alkylene-OH, SO ₂ -C _3-6 cycloalkyl, SO ₂ -piperidinyl, SO ₂ NH-C _1-6 alkyl, SO ₂ N(C _1-6 alkyl) ₂ , halogen, CN, CO-morpholinyl, CH ₂ -pyridinyl, or optionally C _1-6 alkyl, NHC _1-6 alkyl, ═O;
R ^1.1.1 is H, C _1-6 alkyl, C _3-6 cycloalkyl, C _1-6 haloalkyl, CH ₂ CON(C _1-6 alkyl) ₂ , CH ₂ CO-azetindinyl, C _1-6 alkylene-C _3-6 cycloalkyl, CH ₂ -pyranyl, CH ₂ -tetrahydrofuranyl, CH ₂ -furanyl, C _1-6 alkylene-OH or thiadiazolyl, optionally substituted with C _1-6 alkyl;
R ^1.1.2 is H, C _1-6 alkyl, SO ₂ C _1-6 alkyl, or R ^1.1.1 and R ^1.1.2 together form a 4-, 5- or 6-membered carbocyclic ring, which optionally contains one O, replacing a carbon atom of the ring, and optionally substituted with one or two residues selected from the group consisting of CH ₂ OH;
R ^1.2 is
heteroaryl optionally substituted with one or two residues selected from the group consisting of C _1-6 alkyl, C _3-6 cycloalkyl, CH ₂ COO-C _1-6 alkyl, CONR ^1.2.1 R ^1.2.2 , COO-C _1-6 alkyl, CONH ₂ , O-C _1-6 alkyl, halogen, CN, CO-pyrrolidinyl, CO-morpholinyl, or heteroaryl optionally substituted with one or two residues selected from the group consisting of C _1-6 alkyl,
- benzothiazolyl, indazolyl, dihydroindolyl, indanyl, tetrahydroquinolinyl, each of which is optionally substituted with one or two residues selected from the group consisting of N(C _1-6 alkyl) ₂ , CONH-C _1-6 alkyl, ═O;
piperidinyl optionally substituted with pyridinyl,
4,5-dihydro-naphtho[2,1-d]thiazole optionally substituted with NHCO-C _1-6 alkyl,
R ^1.2.1 is H, C _1-6 alkyl;
R ^1.2.2 is H, C _1-6 alkyl;
R ^1.3 is selected from phenyl, pyrazolyl, isoxazolyl, pyrimidinyl, indolyl, or oxadiazolyl, each optionally substituted with one or two residues selected from the group consisting of C _1-6 alkyl, C _3-6 cycloalkyl, O-C _1-6 alkyl, O-C _1-6 haloalkyl;
R ² is selected from C _{1-6 alkylene-phenyl or C 1-6 alkylene-naphthyl, each optionally substituted with one or two residues selected from the group consisting of C 1-6 alkyl , C 1-6} haloalkyl, O—C _1-6 alkyl, O—C _1-6 haloalkyl, halogen, or CH ₂ -thiophenyl, optionally substituted with one or two residues selected from the group consisting of halogen _;
^R3 is H, _C1-4 alkyl;
R ⁴ is H, C _1-4 alkyl, or R ³ and R ⁴ together form a CH ₂ -CH ₂ group;
The method includes administering a compound to a subject.

Another embodiment of the present invention further comprises a compound of formula 1,
In the formula,
A is _CH2 , O or NMe;
^R1 is
・NHR ^1.1 , NMeR ^1.1 ,
・NHR ^1.2 , NMeR ^1.2 ,
・NHCH ₂ -R ^1.3 ,
NH-cyclohexyl optionally substituted with one or two residues selected from the group consisting of C _1-4 alkyl, NHSO ₂ -phenyl, NHCONH-phenyl, halogen;
NH-pyrrolidinyl optionally substituted with one or two residues selected from the group consisting of SO ₂ -C _1-4 alkyl, COO-C _1-4 alkyl,
piperidinyl optionally substituted with one or two residues selected from the group consisting of NHSO ₂ -C _1-4 alkyl, m-methoxyphenyl;
dihydroindolyl, dihydroisoindolyl, tetrahydroquinolinyl or tetrahydroisoquinolinyl optionally substituted with one or two residues selected from the group consisting of C _1-4 alkyl, COO-C _1-4 alkyl, C _1-4 haloalkyl, O-C _1-4 alkyl, NO ₂ , halogen,
a radical selected from NHCH(pyridinyl)CH ₂ COO—C _1-4 alkyl, NHCH(CH ₂ O—C _1-4 alkyl)-benzimidazolyl, optionally substituted with Cl, or a radical selected from 1-aminocyclopentyl, optionally substituted with methyloxadiazolyl,
R ^1.1 is phenyl, optionally C _1-4 alkyl, C _1-4 haloalkyl, CH ₂ CON(C _1-4 alkyl) ₂ , CH ₂ NHCONH-C _3-6 cycloalkyl, CN, CONR ^1.1.1 R ^1.1.2 , COO-C _1-4 alkyl, O-C _1-4 alkyl, SO ₂ -C _1-4 alkyl, SO ₂ -C _1-4 alkylene-OH, SO ₂ -C _3-6 cycloalkyl, SO ₂ -piperidinyl, SO ₂ NH-C _1-4 alkyl, SO ₂ N(C _1-4 alkyl) ₂ , halogen, CO-morpholinyl, CH ₂ -pyridinyl, or each optionally C _1-4 alkyl, NHC _1-4 alkyl, ═O; substituted with 1 or 2 residues selected from the group consisting of imidazolidinyl, piperidinyl, oxazinanyl, pyrazolyl, triazolyl, tetrazolyl, oxazolyl, oxadiazolyl, thiazolyl, pyridinyl, pyrimidinyl;
R ^1.1.1 is H, C _1-6 alkyl, C _3-6 cycloalkyl, C _1-4 haloalkyl, CH ₂ CON(C _1-4 alkyl) ₂ , CH ₂ CO-azetindinyl, C _1-4 alkylene-C _3-6 cycloalkyl, CH ₂ -pyranyl, CH ₂ -tetrahydrofuranyl, CH ₂ -furanyl, C _1-4 alkylene-OH or thiadiazolyl, optionally substituted with C _1-4 alkyl;
R ^1.1.2 is H, C _1-4 alkyl, SO ₂ C _1-4 alkyl, or R ^1.1.1 and R ^1.1.2 together form a 4-, 5- or 6-membered carbocyclic ring, which optionally contains one O, replacing a carbon atom of the ring, and optionally substituted with one or two residues selected from the group consisting of CH ₂ OH;
R ^1.2 is
pyridinyl, pyridazinyl, pyrrolyl _, pyrazolyl, isoxazolyl, thiazolyl, thiadiazolyl, optionally substituted by one or two residues selected from the group consisting of C _1-4 alkyl, C _3-6 cycloalkyl, CH ₂ COO-C _1-4 alkyl, CONR ^1.2.1 R ^1.2.2 , COO-C _1-4 alkyl, CONH ₂ , O-C _1-4 alkyl, halogen, CO-pyrrolidinyl, CO-morpholinyl or pyrazolyl, triazolyl, tetrazolyl, isoxazolyl, oxadiazolyl, each optionally substituted by one or two residues selected from the group consisting of C 1-4 alkyl,
- benzothiazolyl, indazolyl, dihydroindolyl, indanyl, tetrahydroquinolinyl, each optionally substituted with one or two residues selected from the group consisting of N(C _1-4 alkyl) ₂ , CONH-C _1-4 alkyl, ═O;
piperidinyl optionally substituted with pyridinyl,
4,5-dihydro-naphtho[2,1-d]thiazoles optionally substituted with NHCO-C _1-4 alkyl,
R ^1.2.1 is H, C _1-4 alkyl;
R ^1.2.2 is H, C _1-4 alkyl;
R ^1.3 is selected from phenyl, pyrazolyl, isoxazolyl, pyrimidinyl, indolyl or oxadiazolyl, each optionally substituted with one or two residues selected from the group consisting of C _1-4 alkyl, C _3-6 cycloalkyl, O-C _1-4 alkyl, O-C _1-4 haloalkyl;
R ² is selected from C _1-6 alkylene-phenyl or C 1-6 alkylene-naphthyl, each optionally substituted with one or two residues selected from _the group consisting of C _1-4 alkyl, C _1-4 haloalkyl, O—C _1-4 haloalkyl, halogen, or CH ₂ -thiophenyl, optionally substituted with one or two residues selected from the group consisting of halogen;
^R3 is H;
R ⁴ is H or R ³ and R ⁴ together form a CH ₂ -CH ₂ group;
The method includes administering the compound to a subject.

Another embodiment of the present invention further comprises a compound of formula 1,
In the formula,
A is _CH2 , O or NMe;
^R1 is
・NHR ^1.1 , NMeR ^1.1 ,
・NHR ^1.2 , NMeR ^1.2 ,
・NHCH ₂ -R ^1.3 ,
NH-piperidinyl optionally substituted with pyridinyl,
NH-cyclohexyl, optionally substituted with one or two residues selected from the group consisting of t-Bu, NHSO ₂ -phenyl, NHCONH-phenyl, F;
NH-pyrrolidinyl, optionally substituted with one or two residues selected from the group consisting of SO ₂ Me, COO-t-Bu,
piperidinyl optionally substituted with one or two residues selected from the group consisting of NHSO ₂ -n-Bu, m-methoxyphenyl;
dihydroindolyl, dihydroisoindolyl, tetrahydroquinolinyl or tetrahydroisoquinolinyl, optionally substituted with one or two residues selected from the group consisting of Me, COOMe, CF ₃ , OMe, NO ₂ , F, Br,
a radical selected from NHCH(pyridinyl)CH ₂ COOMe, NHCH(CH ₂ OMe)-benzimidazolyl, optionally substituted by Cl, or 1-aminocyclopentyl, optionally substituted by methyloxadiazolyl,
R ^1.1 is phenyl, optionally Me, Et, t-Bu, CF ₃ , CH ₂ CONMe ₂ , CH ₂ NHCONH-cyclohexyl, CN, CONR ^1.1.1 R ^1.1.2 , COOMe, COOEt, OMe, SO ₂ Me, SO ₂ CH ₂ CH ₂ OH, SO ₂ Et, SO ₂ -cyclopropyl, SO ₂ -piperidinyl, SO ₂ NHEt, SO ₂ NMeEt, F, Cl, CO-morpholinyl, CH ₂ -pyridinyl or one or two residues selected from the group consisting of imidazolidinyl, piperidinyl, oxazinanyl, pyrazolyl, triazolyl, tetrazolyl, oxazolyl, oxadiazolyl, thiazolyl, pyridinyl, pyrimidinyl, each of which is optionally substituted with one or two residues selected from the group consisting of Me, NHMe, =O,
R ^1.1.1 is H, Me, Et, t-Bu, i-Pr, cyclopropyl, CH ₂ -i-Pr, CH ₂ -t-Bu, CH(CH ₃ )CH ₂ CH ₃ , CH ₂ CHF ₂ , CH ₂ CONMe ₂ , CH ₂ CO-azetindinyl, CH ₂ -cyclopropyl, CH ₂ -cyclobutyl, CH ₂ -pyranyl, CH ₂ -tetrahydrofuranyl, CH ₂ -furanyl, CH ₂ CH ₂ OH or thiadiazolyl, optionally substituted with Me;
R ^1.1.2 is H, Me, Et, SO ₂ Me, SO ₂ Et, or R ^1.1.1 and R ^1.1.2 together form a 4-, 5- or 6-membered carbocyclic ring, which optionally contains one O, replacing the ring carbon atoms, and optionally substituted with one or two residues selected from the group consisting of CH ₂ OH;
R ^1.2 is
- optionally with Me, Et, Pr, Bu, cyclopropyl, CH ₂ COOEt, CONR ^1.2.1 R ^1.2.2 , COOMe, COOEt, CONH ₂ , OMe, Cl, Br, CO-pyrrolidinyl, CO-morpholinyl or pyridinyl, pyrrolyl, pyrazolyl, isoxazolyl, thiazolyl, thiadiazolyl, each optionally substituted with Me by one or two residues selected from the group consisting of pyrazolyl, triazolyl, tetrazolyl, isoxazolyl, oxadiazolyl,
- benzothiazolyl, indazolyl, dihydroindolyl, indanyl, tetrahydroquinolinyl, each optionally substituted with one or two residues selected from the group consisting of _NMe2 , CONHMe, =O;
4,5-dihydro-naphtho[2,1-d]thiazoles optionally substituted with NHCOMe,
R ^1.2.1 is H, Me;
R ^1.2.2 is H, Me;
^R1.3 is selected from phenyl, pyrazolyl, isoxazolyl, pyrimidinyl, indolyl or oxadiazolyl, each optionally substituted with one or two residues selected from the group consisting of Me, Et, Pr, cyclopentyl, OMe, _OCHF2 ;
^R2 is selected from CH2-phenyl or CH2-naphthyl, each optionally substituted with one or two residues selected from the group consisting of _CH3 , _CF3 , _OCF3 , F, _Cl , Br, Et, or _CH2 -thiophenyl, optionally substituted with one or two residues selected from the group consisting of _Cl , Br;
^R3 is H;
R ⁴ is H or R ³ and R ⁴ together form a CH ₂ -CH ₂ group;
The method includes administering the compound to a subject.

Another embodiment of the present invention further comprises a compound of formula 1,
In the formula,
A is _CH2 , O or NMe;
^R1 is
・NHR ^1.1 ,
・NHR ^1.2
is selected from
R ^1.1 is phenyl, optionally Me, Et, Pr, Bu, CF ₃ , CH ₂ CONMe ₂ , CH ₂ NHCONH-cyclohexyl, CN, CONR ^1.1.1 R ^1.1.2 , COOMe, COOEt, OMe, SO ₂ Me, SO ₂ CH ₂ CH ₂ OH, SO ₂ Et, SO ₂ -cyclopropyl, SO ₂ -piperidinyl, SO ₂ NHEt, SO ₂ NMeEt, F, Cl, CO-morpholinyl, CH ₂ -pyridinyl or one or two residues selected from the group consisting of imidazolidinyl, piperidinyl, oxazinanyl, pyrazolyl, triazolyl, tetrazolyl, oxazolyl, oxadiazolyl, thiazolyl, pyridinyl, pyrimidinyl, each of which is optionally substituted with one or two residues selected from the group consisting of Me, NHMe, =O,
R ^1.1.1 is H, Me, Et, t-Bu, i-Pr, cyclopropyl, CH ₂ -i-Pr, CH ₂ -t-Bu, CH(CH ₃ )CH ₂ CH ₃ , CH ₂ CHF ₂ , CH ₂ CONMe ₂ , CH ₂ CO-azetindinyl, CH ₂ -cyclopropyl, CH ₂ -cyclobutyl, CH ₂ -pyranyl, CH ₂ -tetrahydrofuranyl, CH ₂ -furanyl, CH ₂ CH ₂ OH or thiadiazolyl, optionally substituted with Me;
R ^1.1.2 is H, Me, Et, SO ₂ Me, SO ₂ Et, or R ^1.1.1 and R ^1.1.2 together form a 4-, 5- or 6-membered carbocyclic ring, which optionally contains one O, replacing the ring carbon atoms, and optionally substituted with one or two residues selected from the group consisting of CH ₂ OH;
R ^1.2 is
- optionally with Me, Et, Pr, Bu, cyclopropyl, CH ₂ COOEt, CONR ^1.2.1 R ^1.2.2 , COOMe, COOEt, CONH ₂ , OMe, Cl, Br, CO-pyrrolidinyl, CO-morpholinyl or pyridinyl, pyrrolyl, pyrazolyl, isoxazolyl, thiazolyl, thiadiazolyl, each optionally substituted with Me by one or two residues selected from the group consisting of pyrazolyl, triazolyl, tetrazolyl, isoxazolyl, oxadiazolyl,
- benzothiazolyl, indazolyl, dihydroindolyl, indanyl, tetrahydroquinolinyl, each optionally substituted with one or two residues selected from the group consisting of _NMe2 , CONHMe, =O;
4,5-dihydro-naphtho[2,1-d]thiazoles optionally substituted with NHCOMe,
R ^1.2.1 is H, Me;
R ^1.2.2 is H, Me;
^R2 is selected from _CH2 -phenyl or _CH2 -naphthyl, each of which is optionally substituted with one or two residues selected from the group consisting of _CH3 , _CF3 , _OCF3 , F, Cl, Br, Et;
^R3 is H;
^R4 is H;
The method includes administering a compound to a subject.

Another embodiment of the present invention further comprises a compound of formula 1,
In the formula,
A is _CH2 , O or NMe;
^R1 is
・NHR ^1.1 , NMeR ^1.1 ,
・NHR ^1.2 , NMeR ^1.2 ,
・NHCH ₂ -R ^1.3
is selected from
R ^1.1 is phenyl, optionally Me, Et, Pr, Bu, CF ₃ , CH ₂ CONMe ₂ , CH ₂ NHCONH-cyclohexyl, CN, CONR ^1.1.1 R ^1.1.2 , COOMe, COOEt, OMe, SO ₂ Me, SO ₂ CH ₂ CH ₂ OH, SO ₂ Et, SO ₂ -cyclopropyl, SO ₂ -piperidinyl, SO ₂ NHEt, SO ₂ NMeEt, F, Cl, CO-morpholinyl, CH ₂ -pyridinyl or one or two residues selected from the group consisting of imidazolidinyl, piperidinyl, oxazinanyl, pyrazolyl, triazolyl, tetrazolyl, oxazolyl, oxadiazolyl, thiazolyl, pyridinyl, pyrimidinyl, each of which is optionally substituted with one or two residues selected from the group consisting of Me, NHMe, =O,
R ^1.1.1 is H, Me, Et, Pr, Bu, cyclopropyl, CH ₂ -Pr, CH ₂ -Bu, CH(CH ₃ )CH ₂ CH ₃ , CH ₂ CHF ₂ , CH ₂ CONMe ₂ , CH ₂ CO-azetindinyl, CH ₂ -cyclopropyl, CH ₂ -cyclobutyl, CH ₂ -pyranyl, CH ₂ -tetrahydrofuranyl, CH ₂ -furanyl, CH ₂ CH ₂ OH or thiadiazolyl, optionally substituted with Me;
R ^1.1.2 is H, Me, Et, SO ₂ Me, SO ₂ Et, or R ^1.1.1 and R ^1.1.2 together form a 4-, 5- or 6-membered carbocyclic ring, which optionally contains one O, replacing the ring carbon atoms, and optionally substituted with one or two residues selected from the group consisting of CH ₂ OH;
R ^1.2 is
- optionally with Me, Et, Pr, Bu, cyclopropyl, CH ₂ COOEt, CONR ^1.2.1 R ^1.2.2 , COOMe, COOEt, CONH ₂ , OMe, Cl, Br, CO-pyrrolidinyl, CO-morpholinyl or pyridinyl, pyrrolyl, pyrazolyl, isoxazolyl, thiazolyl, thiadiazolyl, each optionally substituted with Me by one or two residues selected from the group consisting of pyrazolyl, triazolyl, tetrazolyl, isoxazolyl, oxadiazolyl,
- benzothiazolyl, indazolyl, dihydroindolyl, indanyl, tetrahydroquinolinyl, each optionally substituted with one or two residues selected from the group consisting of _NMe2 , CONHMe, =O;
4,5-dihydro-naphtho[2,1-d]thiazoles optionally substituted with NHCOMe,
R ^1.2.1 is H, Me;
R ^1.2.2 is H, Me;
^R1.3 is selected from phenyl, pyrazolyl, isoxazolyl, pyrimidinyl, indolyl or oxadiazolyl, each optionally substituted with one or two residues selected from the group consisting of Me, Et, Pr, cyclopentyl, OMe, _OCHF2 ;
^R2 is selected from CH2-phenyl or CH2-naphthyl, each optionally substituted with one or two residues selected from the group consisting of _CH3 , _CF3 , _OCF3 , F, _Cl , Br, Et, or _CH2 -thiophenyl, optionally substituted with one or two residues selected from the group consisting of _Cl , Br;
^R3 is H;
R ⁴ is H or R ³ and R ⁴ together form a CH ₂ -CH ₂ group;
The method includes administering the compound to a subject.

Another embodiment of the present invention further comprises a compound of formula 1,
In the formula,
A is _CH2 , O or NMe;
^R1 is
selected from NHR ^1.1 , NMeR ^1.1 ,
R ^1.1 is phenyl, optionally Me, Et, t-Bu, CF ₃ , CH ₂ CONMe ₂ , CH ₂ NHCONH-cyclohexyl, CN, CONR ^1.1.1 R ^1.1.2 , COOMe, COOEt, OMe, SO ₂ Me, SO ₂ CH ₂ CH ₂ OH, SO ₂ Et, SO ₂ -cyclopropyl, SO ₂ -piperidinyl, SO ₂ NHEt, SO ₂ NMeEt, F, Cl, CO-morpholinyl, CH ₂ -pyridinyl or one or two residues selected from the group consisting of imidazolidinyl, piperidinyl, oxazinanyl, pyrazolyl, triazolyl, tetrazolyl, oxazolyl, oxadiazolyl, thiazolyl, pyridinyl, pyrimidinyl, each of which is optionally substituted with one or two residues selected from the group consisting of Me, NHMe, =O,
R ^1.1.1 is H, Me, Et, Bu, Pr, cyclopropyl, CH ₂ -Pr, CH ₂ -Bu, CH(CH ₃ )CH ₂ CH ₃ , CH ₂ CHF ₂ , CH ₂ CONMe ₂ , CH ₂ CO-azetindinyl, CH ₂ -cyclopropyl, CH ₂ -cyclobutyl, CH ₂ -pyranyl, CH ₂ -tetrahydrofuranyl, CH ₂ -furanyl, CH ₂ CH ₂ OH or thiadiazolyl, optionally substituted with Me;
R ^1.1.2 is H, Me, Et, SO ₂ Me, SO ₂ Et, or R ^1.1.1 and R ^1.1.2 together form a 4-, 5- or 6-membered carbocyclic ring, which optionally contains one O, replacing the ring carbon atoms, and optionally substituted with one or two residues selected from the group consisting of CH ₂ OH;
^R2 is selected from CH2-phenyl or CH2-naphthyl, each optionally substituted with one or two residues selected from the group consisting of _CH3 , _CF3 , _OCF3 , F, _Cl , Br, Et, or _CH2 -thiophenyl, optionally substituted with one or two residues selected from the group consisting of _Cl , Br;
^R3 is H;
R ⁴ is H or R ³ and R ⁴ together form a CH ₂ -CH ₂ group;
The method includes administering a compound to a subject.

Another embodiment of the present invention further comprises a compound of formula 1,
In the formula,
A is _CH2 , O or NMe;
^R1 is
selected from NHR ^1.1 , NMeR ^1.1 ,
R ^1.1 is phenyl, optionally Me, Et, t-Bu, CF ₃ , CH ₂ CONMe ₂ , CH ₂ NHCONH-cyclohexyl, CN, CONR ^1.1.1 R ^1.1.2 , COOMe, COOEt, OMe, SO ₂ Me, SO ₂ CH ₂ CH ₂ OH, SO ₂ Et, SO ₂ -cyclopropyl, SO ₂ -piperidinyl, SO ₂ NHEt, SO ₂ NMeEt, F, Cl, CO-morpholinyl, CH ₂ -pyridinyl or one or two residues selected from the group consisting of imidazolidinyl, piperidinyl, oxazinanyl, pyrazolyl, triazolyl, tetrazolyl, oxazolyl, oxadiazolyl, thiazolyl, pyridinyl, pyrimidinyl, each of which is optionally substituted with one or two residues selected from the group consisting of Me, NHMe, =O,
R ^1.1.1 is H, Me, Et, Bu, Pr, cyclopropyl, CH ₂ -Pr, CH ₂ -Bu, CH(CH ₃ )CH ₂ CH ₃ , CH ₂ CHF ₂ , CH ₂ CONMe ₂ , CH ₂ CO-azetindinyl, CH ₂ -cyclopropyl, CH ₂ -cyclobutyl, CH ₂ -pyranyl, CH ₂ -tetrahydrofuranyl, CH ₂ -furanyl, CH ₂ CH ₂ OH or thiadiazolyl, optionally substituted with Me;
R ^1.1.2 is H, Me, Et, SO ₂ Me, SO ₂ Et, or R ^1.1.1 and R ^1.1.2 together form a 4-, 5- or 6-membered carbocyclic ring, which optionally contains one O, replacing the ring carbon atoms, and optionally substituted with one or two residues selected from the group consisting of CH ₂ OH;
^R2 is defined as shown in Table 1 below,
^R3 is H;
^R4 is H.

Another embodiment of the present invention further comprises a compound of formula 1,
In the formula,
A is _CH2 , O or NMe;
^R1 is
selected from NHR ^1.1 , NMeR ^1.1 ,
R ^1.1 is phenyl, optionally Me, Et, t-Bu, CF ₃ , CH ₂ CONMe ₂ , CH ₂ NHCONH-cyclohexyl, CN, CONR ^1.1.1 R ^1.1.2 , COOMe, COOEt, OMe, SO ₂ Me, SO ₂ CH ₂ CH ₂ OH, SO ₂ Et, SO ₂ -cyclopropyl, SO ₂ -piperidinyl, SO ₂ NHEt, SO ₂ NMeEt, F, Cl, CO-morpholinyl, CH ₂ -pyridinyl or one or two residues selected from the group consisting of imidazolidinyl, piperidinyl, oxazinanyl, pyrazolyl, triazolyl, tetrazolyl, oxazolyl, oxadiazolyl, thiazolyl, pyridinyl, pyrimidinyl, each of which is optionally substituted with one or two residues selected from the group consisting of Me, NHMe, =O,
R ^1.1.1 and R ^1.1.2 together form a 4-, 5- or 6-membered carbocyclic ring, which optionally contains one O, replacing the carbon atoms of the ring, and optionally substituted with one or two residues selected from the group consisting of CH ₂ OH;
^R2 is defined as shown in Table 1 below,
^R3 is H;
^R4 is H.

Another embodiment of the present invention further comprises a compound of formula 1,
In the formula,
A is _CH2 , O or NMe;
^R1 is
selected from NHR ^1.1 , NMeR ^1.1 ,
R ^1.1 is phenyl, optionally substituted with one or two residues selected from the group consisting of Me, Et, t-Bu, CF ₃ , CH ₂ CONMe ₂ , CH ₂ NHCONH-cyclohexyl, CN, CONR ^1.1.1 R ^1.1.2 , COOMe, COOEt, OMe, F, Cl;
R ^1.1.1 is H, Me, Et, Bu, Pr, cyclopropyl, CH ₂ -Pr, CH ₂ -Bu, CH(CH ₃ )CH ₂ CH ₃ , CH ₂ CHF ₂ , CH ₂ CONMe ₂ , CH ₂ CO-azetindinyl, CH ₂ -cyclopropyl, CH ₂ -cyclobutyl, CH ₂ -pyranyl, CH ₂ -tetrahydrofuranyl, CH ₂ -furanyl, CH ₂ CH ₂ OH or thiadiazolyl, optionally substituted with Me;
^R1.1.2 is H, Me, Et, _SO2Me , _SO2Et ;
^R2 is defined as shown in Table 1 below,
^R3 is H;
^R4 is H.

Another embodiment of the invention further comprises administering to a subject a compound of formula 1,
In the formula,
A is _CH2 , O or NMe;
^R1 is
selected from NHR ^1.1 , NMeR ^1.1 ,
R ^1.1 is phenyl, optionally substituted with one or two residues selected from the group consisting of SO ₂ Me, SO ₂ CH ₂ CH ₂ OH, SO ₂ Et, SO ₂ -cyclopropyl, SO ₂ -piperidinyl, SO ₂ NHEt, SO ₂ NMeEt;
R ^1.1.1 is H, Me, Et, Bu, Pr, cyclopropyl, CH ₂ -Pr, CH ₂ -Bu, CH(CH ₃ )CH ₂ CH ₃ , CH ₂ CHF ₂ , CH ₂ CONMe ₂ , CH ₂ CO-azetindinyl, CH ₂ -cyclopropyl, CH ₂ -cyclobutyl, CH ₂ -pyranyl, CH ₂ -tetrahydrofuranyl, CH ₂ -furanyl, CH ₂ CH ₂ OH or thiadiazolyl, optionally substituted with Me;
^R1.1.2 is H, Me, Et, _SO2Me , _SO2Et ;
^R2 is defined as shown in Table 1 below,
^R3 is H;
^R4 is H.

Another embodiment of the invention further comprises administering to a subject a compound of formula 1, wherein
A is _CH2 , O or NMe;
^R1 is
selected from NHR ^1.1 , NMeR ^1.1 ,
R ^1.1 is phenyl, optionally substituted with one residue selected from the group consisting of Me, Et, t-Bu, CF ₃ , CH ₂ CONMe ₂ , CH ₂ NHCONH-cyclohexyl, CN, CONR ^1.1.1 R ^1.1.2 , COOMe, COOEt, OMe, SO ₂ Me, SO ₂ CH ₂ CH ₂ OH, SO ₂ Et, SO ₂ cyclopropyl, SO ₂ piperidinyl, SO ₂ NHEt, SO ₂ NMeEt, F, Cl, and further substituted with one residue selected from the group consisting of CO-morpholinyl, CH ₂ pyridinyl, or one residue selected from the group consisting of imidazolidinyl, piperidinyl, oxazinanyl, pyrazolyl, triazolyl, tetrazolyl, oxazolyl, oxadiazolyl, thiazolyl, pyridinyl, pyrimidinyl, pyrimidinyl, each of which is optionally substituted with one or two residues selected from the group consisting of Me, NHMe, =O,
R ^1.1.1 is H, Me, Et, Bu, Pr, cyclopropyl, CH ₂ -Pr, CH ₂ -Bu, CH(CH ₃ )CH ₂ CH ₃ , CH ₂ CHF ₂ , CH ₂ CONMe ₂ , CH ₂ CO-azetindinyl, CH ₂ -cyclopropyl, CH ₂ -cyclobutyl, CH ₂ -pyranyl, CH ₂ -tetrahydrofuranyl, CH ₂ -furanyl, CH ₂ CH ₂ OH or thiadiazolyl, optionally substituted with Me;
^R1.1.2 is H, Me, Et, _SO2Me , _SO2Et ;
^R2 is defined as shown in Table 1 below,
^R3 is H;
^R4 is H.

Another embodiment of the present invention further comprises a compound of formula 1,
In the formula,
A is _CH2 , O or NMe;
^R1 is
selected from NHR ^1.2 , NMeR ^1.2 ,
R ^1.2 is
- optionally with Me, Et, Pr, Bu, cyclopropyl, CH ₂ COOEt, CONR ^1.2.1 R ^1.2.2 , COOMe, COOEt, CONH ₂ , OMe, Cl, Br, CO-pyrrolidinyl, CO-morpholinyl or pyridinyl, pyridazinyl, pyrrolyl, pyrazolyl, isoxazolyl, thiazolyl, thiadiazolyl, each of which is optionally substituted with Me by one or two residues selected from the group consisting of pyrazolyl, triazolyl, tetrazolyl, isoxazolyl, oxadiazolyl,
- benzothiazolyl, indazolyl, dihydroindolyl, indanyl, tetrahydroquinolinyl, each optionally substituted with one or two residues selected from the group consisting of _NMe2 , CONHMe, =O;
4,5-dihydro-naphtho[2,1-d]thiazoles optionally substituted with NHCOMe,
R ^1.2.1 is H, Me;
R ^1.2.2 is H, Me;
^R2 is selected from CH2-phenyl or CH2-naphthyl, each optionally substituted with one or two residues selected from the group consisting of _CH3 , _CF3 , _OCF3 , F, _Cl , Br, Et, or _CH2 -thiophenyl, optionally substituted with one or two residues selected from the group consisting of _Cl , Br;
^R3 is H;
R ⁴ is H or R ³ and R ⁴ together form a CH ₂ -CH ₂ group;
The method includes administering the compound to a subject.

Another embodiment of the present invention further comprises a compound of formula 1,
In the formula,
A is _CH2 , O or NMe;
^R1 is
selected from NHR ^1.2 , NMeR ^1.2 ,
R ^1.2 is selected from pyridinyl, pyridazinyl, pyrrolyl, pyrazolyl, isoxazolyl, thiazolyl, thiadiazolyl, optionally substituted by one or two residues selected from the group consisting of Me, Et, n-Pr, i-Pr, Bu, cyclopropyl, CH ₂ COOEt, CONR ^1.2.1 R ^1.2.2 , COOMe, COOEt, CONH ₂ , OMe, Cl, Br, CO-pyrrolidinyl, CO-morpholinyl or pyrazolyl, triazolyl, tetrazolyl, isoxazolyl, oxadiazolyl, each optionally substituted by Me;
R ^1.2.1 is H, Me;
R ^1.2.2 is H, Me;
^R2 is selected from CH2-phenyl or CH2-naphthyl, each optionally substituted with one or two residues selected from the group consisting of _CH3 , _CF3 , _OCF3 , F, _Cl , Br, Et, or _CH2 -thiophenyl, optionally substituted with one or two residues selected from the group consisting of _Cl , Br;
^R3 is H;
R ⁴ is H or R ³ and R ⁴ together form a CH ₂ -CH ₂ group;
The method includes administering a compound to a subject.

Another embodiment of the invention further comprises administering to a subject a compound of formula 1,
In the formula,
A is _CH2 , O or NMe;
^R1 is
- selected from NHCH ₂ -R ^1.3 ;
^R1.3 is selected from phenyl, pyrazolyl, isoxazolyl, pyrimidinyl, indolyl or oxadiazolyl, each optionally substituted with one or two residues selected from the group consisting of Me, Et, Pr, cyclopentyl, OMe, _OCHF2 ;
^R2 is selected from CH2-phenyl or CH2-naphthyl, each optionally substituted with one or two residues selected from the group consisting of _CH3 , _CF3 , _OCF3 , F, _Cl , Br, Et, or _CH2 -thiophenyl, optionally substituted with one or two residues selected from the group consisting of _Cl , Br;
^R3 is H;
R ⁴ is H, or R ³ and R ⁴ together form a CH ₂ -CH ₂ group.

Another embodiment of the invention further comprises administering to the subject a compound of formula 1,
In the formula,
A is _CH2 , O or NMe;
^R1 is
NH-piperidinyl optionally substituted with pyridinyl,
NH-cyclohexyl, optionally substituted with one or two residues selected from the group consisting of t-Bu, NHSO ₂ -phenyl, NHCONH-phenyl, F;
NH-pyrrolidinyl, optionally substituted with one or two residues selected from the group consisting of SO ₂ Me, COO-t-Bu,
piperidinyl optionally substituted with one or two residues selected from the group consisting of NHSO ₂ -n-Bu, m-methoxyphenyl;
dihydroindolyl, dihydroisoindolyl, tetrahydroquinolinyl or tetrahydroisoquinolinyl, optionally substituted with one or two residues selected from the group consisting of Me, COOMe, CF ₃ , OMe, NO ₂ , F, Br,
a group selected from NHCH(pyridinyl)CH ₂ COOMe, NHCH(CH ₂ OMe)-benzimidazolyl, optionally substituted with Cl,
or 1-aminocyclopentyl optionally substituted with methyloxadiazolyl,
^R2 is selected from CH2-phenyl or CH2-naphthyl, each optionally substituted with one or two residues selected from the group consisting of _CH3 , _CF3 , _OCF3 , F, _Cl , Br, Et, or _CH2 -thiophenyl, optionally substituted with one or two residues selected from the group consisting of _Cl , Br;
^R3 is H;
R ⁴ is H, or R ³ and R ⁴ together form a CH ₂ -CH ₂ group.

Another embodiment of the present invention further comprises a compound of formula 1,
In the formula,
A is _CH2 , O or NMe;
^R1 is selected from ^NHR1.2 , ^NMeR1.2 ;
^R2 is defined as shown in Table 1 below,
^R3 is H;
^R4 and ^R1.2 are
- optionally substituted by one or two residues selected from the group consisting of Me, Et, i-Pr, n-Bu, cyclopropyl, CONR ^1.2.1 R ^1.2.2 , COOMe, COOEt, CONH ₂ , OMe, Cl, Br, CO-pyrrolidinyl, CO-morpholinyl or pyridinyl, each optionally substituted by Me,
pyrrolyl, optionally substituted with one or two residues selected from the group consisting of Me, Et, COOMe, COOEt,
pyrazolyl optionally substituted with one or two residues selected from the group consisting of Me, Et, cyclopropyl, COOEt, CO-pyrrolidinyl,
isoxazolyl optionally substituted with one or two residues selected from the group consisting of t-Bu, COOEt,
thiazolyl optionally substituted with one or two residues selected from the group consisting of Me, n-Pr, i-Pr, Bu, COOMe, COOEt, CH ₂ COOEt, CONR ^1.2.1 R ^1.2.2 ,
thiadiazolyl optionally substituted with one or two residues selected from the group consisting of COOEt,
- benzothiazolyl, indazolyl, dihydroindolyl, indanyl, tetrahydroquinolinyl, each optionally substituted with one or two residues selected from the group consisting of _NMe2 , CONHMe, =O;
4,5-dihydro-naphtho[2,1-d]thiazoles optionally substituted with NHCOMe,
R ^1.2.1 is H or Me;
R ^1.2.2 is H or Me;
The method includes administering a compound to a subject.

Another embodiment of the present invention further comprises a compound of formula 1,
In the formula,
A is _CH2 , O or NMe;
^R1 is selected from ^NHR1.2 , ^NMeR1.2 ;
^R2 is defined as shown in Table 1 below,
^R3 is H;
^R4 and ^R1.2 are
pyridinyl optionally substituted with one or two residues selected from the group consisting of Me, Et, i-Pr, n-Bu, CONR ^1.2.1 R ^1.2.2 , COOMe, COOEt, CONH ₂ , OMe, Cl, Br,
pyrrolyl, optionally substituted with one or two residues selected from the group consisting of Me, Et, COOMe, COOEt,
pyrazolyl optionally substituted with one or two residues selected from the group consisting of Me, Et, cyclopropyl, COOEt, CO-pyrrolidinyl,
isoxazolyl optionally substituted with one or two residues selected from the group consisting of t-Bu, COOEt,
thiazolyl optionally substituted with one or two residues selected from the group consisting of Me, n-Pr, i-Pr, Bu, COOMe, COOEt, CONR ^1.2.1 R ^1.2.2 ,
thiadiazolyl optionally substituted with one or two residues selected from the group consisting of COOEt,
- benzothiazolyl, indazolyl, dihydroindolyl, indanyl, tetrahydroquinolinyl, each of which is optionally substituted with one or two residues selected from the group consisting of _NMe2 , CONHMe, =O,
R ^1.2.1 is H or Me;
R ^1.2.2 is H or Me;
The method includes administering a compound to a subject.

Another embodiment of the present invention further comprises a compound of formula 1,
In the formula,
A is CH ₂ , O or NMe, R ¹ is selected from NHR ^1.2 , NMeR ^1.2 , R ² is defined as in table 1 shown below, R ³ is H, R ⁴ is H, R ^1.2 is pyridinyl optionally substituted with one or two residues selected from the group consisting of Me, Et, i-Pr, n-Bu, CONR ^1.2.1 R ^1.2.2 , COOMe, COOEt, CONH ₂ , OMe, Cl, Br, R ^1.2.1 is H or Me, R ^1.2.2 is H or Me,
A is CH ₂ , O or NMe, R ¹ is selected from NHR ^1.2 , NMeR ^1.2 , R ² is defined as in table 1 shown below, R ³ is H, R ⁴ is H, R ^1.2 is pyrrolyl optionally substituted with one or two residues selected from the group consisting of Me, Et, COOMe, COOEt, R ^1.2.1 is H or Me, R ^1.2.2 is H or Me,
A is CH ₂ , O or NMe, R ¹ is selected from NHR ^1.2 , NMeR ^1.2 , R ² is defined as in table 1 shown below, R ³ is H, R ⁴ is H, R ^1.2 is pyrazolyl optionally substituted with one or two residues selected from the group consisting of Me, Et, cyclopropyl, COOEt, CO-pyrrolidinyl, R ^1.2.1 is H or Me, R ^1.2.2 is H or Me,
A is CH ₂ , O or NMe, R ¹ is selected from NHR ^1.2 , NMeR ^1.2 , R ² is defined as in Table 1 shown below, R ³ is H, R ⁴ is H, R ^1.2 is isoxazolyl optionally substituted with one or two residues selected from the group consisting of t-Bu, COOE, R ^1.2.1 is H or Me, R ^1.2.2 is H or Me,
A is CH ₂ , O or NMe, R ¹ is selected from NHR ^1.2 , NMeR ^1.2 , R ² is defined as in table 1 shown below, R ³ is H, R ⁴ is H, R ^1.2 is thiazolyl optionally substituted with one or two residues selected from the group consisting of Me, n-Pr, i-Pr, Bu, COOMe, COOEt, CONR ^1.2.1 R ^1.2.2 , R ^1.2.1 is H or Me, R ^1.2.2 is H or Me,
A is CH ₂ , O or NMe, R ¹ is selected from NHR ^1.2 , NMeR ^1.2 , R ² is defined as in table 1 shown below, R ³ is H, R ⁴ is H, R ^1.2 is thiadiazolyl optionally substituted with one or two residues selected from the group consisting of COOEt, R ^1.2.1 is H or Me, R ^1.2.2 is H or Me,
A is CH ₂ , O or NMe, R ¹ is selected from NHR ^1.2 , NMeR ^1.2 , R ² is defined as in table 1 shown below, R ³ is H, R ⁴ is H, R ^1.2 is benzothiazolyl, indazolyl, dihydroindolyl, indanyl, tetrahydroquinolinyl, each of which is optionally substituted with one or two residues selected from the group consisting of NMe ₂ , CONHMe, ═O, R ^1.2.1 is H or Me, and R ^1.2.2 is H or Me,
The method includes administering the compound to a subject.

Another embodiment of the present invention further comprises a compound of formula 1,
In the formula,
R ^1.3 is
phenyl optionally substituted with _OCHF2 ,
pyrazolyl optionally substituted with Me or Et,
isoxazolyl optionally substituted with Pr,
pyrimidinyl optionally substituted with two OMe,
Indolyl,
- oxadiazolyl optionally substituted with cyclopentyl

Another embodiment of the present invention further includes administering to a subject a compound of formula 1, wherein all groups are as defined above except that A is _CH2 .

Another embodiment of the invention further includes administering to a subject a compound of formula 1, wherein all groups are as defined above except that A is O.

Another embodiment of the invention further includes administering to a subject a compound of formula 1, wherein all groups are as defined above, except that A is NMe.

Another embodiment of the invention is a compound of formula 1:
In the formula,
A is _CH2 , O or NMe;
^R1 is

is selected from
^R2 is

is selected from
^R3 is H;
R ⁴ is H, or R ³ and R ⁴ together form a CH ₂ -CH ₂ group.

Another embodiment of the invention is a compound of formula 1,
In the formula,
A is as defined above,
^R3 is H;
^R4 is H;
^R2 is defined as shown in Table 1 below,
R ¹ is a compound selected from the following:

Another embodiment of the present invention further comprises a compound of formula 1,
In the formula,
A is as defined above,
^R3 is H;
^R4 is H;
^R2 is defined as shown in Table 1 below,
^R1 is

Selected from:
The method includes administering a compound to a subject.

Another embodiment of the present invention further comprises a compound of formula 1,
In the formula,
A is as defined above,
^R3 is H;
^R4 is H;
^R2 is defined as shown in Table 1 below,
^R1 is

Selected from:
The method includes administering the compound to a subject.

Another embodiment of the present invention further comprises a compound of formula 1,
In the formula,
A is as defined above,
^R3 is H;
^R4 is H;
^R2 is defined as shown in Table 1 below,
^R1 is

Selected from:
The method includes administering the compound to a subject.

Another embodiment of the present invention further comprises a compound of formula 1,
wherein A is as defined above;
^R3 is H;
^R4 is H;
^R2 is defined as shown in Table 1 below,
^R1 is

Selected from:
The method includes administering a compound to a subject.

Another embodiment of the invention further includes administration of a compound of formula 1 to a subject, wherein the compound of formula 1 is present in the form of an individual optical isomer, a mixture of individual enantiomers, or a racemate, e.g., an enantiomerically pure compound.

Another embodiment of the present invention further includes administration of a compound of formula 1 to a subject, wherein the compound of formula 1 is in the form of an acid addition salt with a pharmacologically acceptable acid, and optionally in the form of a solvate and/or hydrate.

b. Co-Crystals and Salts Additional embodiments of the invention further include administering to a subject a co-crystal of a compound of Formula 2 (below). Generally, for groups in this "Co-Crystals and Salts" section that contain more than one subgroup, the first subgroup listed is the radical attachment point, e.g., the substituent "C _1-3 alkyl-aryl" means an aryl group that is attached to a C 1-3 alkyl group, where the alkyl group is attached to a parent nucleus or group to which the substituent is attached.

In the formula,
R ¹ is C _1-6 alkyl, C _1-6 haloalkyl, O—C _1-6 haloalkyl, halogen;
m is 1, 2, or 3, and in some cases, 1 or 2;
R ^2a and R ^2b are each independently selected from H, C _1-6 alkyl, C _1-6 alkenyl, C _1-6 alkynyl, C _3-6 cycloalkyl, COO-C _1-6 alkyl, O-C _1-6 alkyl, CONR ^2b.1 R ^2b.2 and halogen;
R ^2b.1 is H, C _1-6 alkyl, C _0-4 alkyl-C _3-6 cycloalkyl, C _1-6 haloalkyl;
R ^2b.2 is H, C _1-6 alkyl or a C _3-6 alkylene group in which R ^2b.1 and R ^2b.2 together form a heterocyclic ring with the nitrogen atom, optionally one carbon atom of the ring being replaced by an oxygen atom;
^R3 is H, _C1-6 alkyl;
X is an anion selected from the group consisting of chloride, bromide, iodide, sulfate, phosphate, methanesulfonate, nitrate, maleate, acetate, benzoate, citrate, salicylate, fumarate, tartrate, dibenzoyltartrate, oxalate, succinate, benzoate and p-toluenesulfonate, in some cases chloride or dibenzoyltartrate;
j is 0, 0.5, 1, 1.5 or 2, and in some cases, 1 or 2;
The co-crystal former is selected from the group consisting of orotic acid, hippuric acid, L-pyroglutamic acid, D-pyroglutamic acid, nicotinic acid, L-(+)-ascorbic acid, saccharin, piperazine, 3-hydroxy-2-naphthoic acid, mucic acid (galactaric acid), pamoic acid (embonic acid), stearic acid, cholic acid, deoxycholic acid, nicotinamide, isonicotinamide, succinamide, uracil, L-lysine, L-proline, D-valine, L-arginine, glycine, and in some cases ascorbic acid, mucic acid, pamoic acid, succinamide, nicotinic acid, nicotinamide, isonicotinamide, l-lysine, l-proline.

Another aspect of the invention is further a co-crystal of a compound of formula 2,
In the formula,
R ^2a is H, C _1-6 alkyl, C _1-6 alkenyl, C _1-6 alkynyl, C _3-6 cycloalkyl, O—C _1-6 alkyl, CONR ^2a.1 R ^2a.2 ;
R ^2a.1 is H, C _1-6 alkyl, C _1-6 haloalkyl;
R ^2a.2 is H, C _1-6 alkyl;
R ^2b is H, C _1-6 alkyl, C _1-6 alkenyl, C _1-6 alkynyl, C _3-6 cycloalkyl, COO-C _1-6 alkyl, O-C _1-6 alkyl, CONR ^2b.1 R ^2b.2 , halogen;
R ^2b.1 is H, C _1-6 alkyl, C _0-4 alkyl-C _3-6 cycloalkyl, C _1-6 haloalkyl;
R ^2b.2 is H, C _1-6 alkyl or a C _3-6 alkylene group in which R ^2b.1 and R ^2b.2 together form a heterocyclic ring with the nitrogen atom, optionally one carbon atom of the ring is replaced by an oxygen atom and the remaining residues are as defined above;
The cocrystal is administered to a subject.

Another aspect of the invention is further a co-crystal of a compound of formula 2,
In the formula,
R ^2a is H, C _1-6 alkyl, C _1-6 alkynyl, C _3-6 cycloalkyl, O—C _1-6 alkyl, CONR ^2a.1 R ^2a.2 ;
R ^2a.1 is C _1-6 alkyl;
R ^2a.2 is H;
R ^2b is H, C _1-6 alkyl, O—C _1-6 alkyl, CONR ^2b.1 R ^2b.2 ;
R ^2b.1 is C _1-6 alkyl, C _0-4 alkyl-C _3-6 cycloalkyl, C _1-6 haloalkyl;
R ^2b.2 is H, C _1-6 alkyl or a C _3-6 alkylene group in which R ^2b.1 and R ^2b.2 together form a heterocyclic ring with the nitrogen atom, optionally one carbon atom of the ring being replaced by an oxygen atom and the remaining residues being as defined above;
The cocrystal is administered to a subject.

Another aspect of the invention is further a co-crystal of a compound of formula 2,
In the formula,
R ^2a is H, C _1-4 alkyl, C _1-4 alkynyl, C _3-6 cycloalkyl, O—C _1-4 alkyl, CONR ^2a.1 R ^2a.2 ;
R ^2a.1 is C _1-4 alkyl;
R ^2a.2 is H;
R ^2b is H, C _1-4 alkyl, O—C _1-4 alkyl, CONR ^2b.1 R ^2b.2 ;
R ^2b.1 is C _1-4 alkyl, C _0-4 alkyl-C _3-6 cycloalkyl, C _1-4 haloalkyl;
R ^2b.2 is H, C _1-4 alkyl or a C _3-6 alkylene group in which R ^2b.1 and R ^2b.2 together form a heterocyclic ring with the nitrogen atom, optionally one carbon atom of the ring being replaced by an oxygen atom and the remaining residues being as defined above;
The cocrystal is administered to a subject.

Another aspect of the invention is further a co-crystal of a compound of formula 2,
In the formula,
R ^2a is H, C _1-4 alkyl;
R ^2b is H, CONR ^2b.1 R ^2b.2 ;
R ^2b.1 is C _1-4 alkyl, C _0-4 alkyl-C _3-6 cycloalkyl, C _1-4 haloalkyl;
R ^2b.2 is H, C _1-4 alkyl or a C _3-6 alkylene group in which R ^2b.1 and R ^2b.2 together form a heterocyclic ring with the nitrogen atom, optionally one carbon atom of the ring being replaced by an oxygen atom and the remaining residues being as defined above;
The cocrystal is administered to a subject.

Another aspect of the invention is further a co-crystal of a compound of formula 2,
In the formula,
R ¹ is C _1-6 alkyl, C _1-6 haloalkyl, O—C _1-6 haloalkyl, halogen;
m is 1 or 2;
R ^2a is H, C _1-4 alkyl;
R ^2b is H, CONR ^2b.1 R ^2b.2 ;
R ^2b.1 is C _1-4 alkyl, C _0-4 alkyl-C _3-6 cycloalkyl, C _1-4 haloalkyl;
R ^2b.2 is H, C _1-4 alkyl, or
or R ^2b.1 and R ^2b.2 together are a C _3-6 alkylene group forming a heterocyclic ring with the nitrogen atom, optionally one carbon atom of the ring being replaced by an oxygen atom;
^R3 is H, _C1-6 alkyl;
X is an anion selected from the group consisting of chloride or dibenzoyltartrate;
j is 1 or 2;
The cocrystal is administered to a subject.

Another aspect of the invention is further a co-crystal of a compound of formula 2,
In the formula,
R ^2a is H, C _1-4 alkyl, and in some cases methyl, ethyl, propyl;
R ^2b is H, CONR ^2b.1 R ^2b.2 ;
R ^2b.1 is C _1-4 alkyl, in some cases methyl, ethyl, propyl;
R ^2b.2 is C _1-4 alkyl, in some cases methyl, ethyl, propyl;
The remaining residues are as defined above.
The cocrystal is administered to a subject.

Another aspect of the invention is further a co-crystal of a compound of formula 2,
In the formula,
R ^2a is H, C _1-4 alkyl, and in some cases methyl, ethyl, propyl;
R ^2b is H, CONR ^2b.1 R ^2b.2 ;
R ^2b.1 is C _0-4 alkyl-C _3-6 cycloalkyl;
R ^2b.2 is H, C _1-4 alkyl, in some cases H, methyl, ethyl, propyl;
The remaining residues are as defined above.
The method includes administering the cocrystal to a subject.

Another aspect of the invention is further a co-crystal of a compound of formula 2,
In the formula,
R ^2a is H, C _1-4 alkyl, and in some cases methyl, ethyl, propyl;
R ^2b is H, CONR ^2b.1 R ^2b.2 ;
R ^2b.1 is C _1-4 haloalkyl;
R ^2b.2 is H, C _1-4 alkyl, in some cases H, methyl, ethyl, propyl;
The remaining residues are as defined above.
The method includes administering the cocrystal to a subject.

Another aspect of the invention is further a co-crystal of a compound of formula 2,
In the formula,
R ^2b.1 and R ^2b.2 together are a C _3-6 alkylene group forming a heterocyclic ring with the nitrogen atom, optionally one carbon atom of the ring is replaced by an oxygen atom, and the remaining residues are as defined above;
The cocrystal is administered to a subject.

Another aspect of the invention is further a co-crystal of a compound of formula 2,
In the formula,
R ¹ , m, R ^2a , R ^2b , R ³ , X and j are as defined above and the co-crystal former is selected from the group consisting of ascorbic acid, mucic acid, pamoic acid, succinamide, nicotinic acid, nicotinamide, isonicotinamide, l-lysine, l-proline, or a hydrate or hydrochloride thereof;
The cocrystal is administered to a subject.

Another aspect of the present invention is a co-crystal of a compound of formula 2a:
wherein R ^2a , R ^2b , R ³ , X and j are as defined above.
The cocrystal is administered to a subject.

Another aspect of the invention is further a co-crystal of a compound of formula 2a,
In the formula,
R ^2a is H, C _1-4 alkyl, and in some cases methyl, ethyl, propyl;
R ^2b is H, CONR ^2b.1 R ^2b.2 ;
R ^2b.1 is C _1-4 alkyl, in some cases methyl, ethyl, propyl;
R ^2b.2 is C _1-4 alkyl, in some cases methyl, ethyl, propyl;
The remaining residues are as defined above.
The cocrystal is administered to a subject.

Another aspect of the invention is further a co-crystal of a compound of formula 2a,
In the formula,
R ^2a is H, C _1-4 alkyl, and in some cases methyl, ethyl, propyl;
R ^2b is H, CONR ^2b.1 R ^2b.2 ;
R ^2b.1 is C _0-4 alkyl-C _3-6 cycloalkyl;
R ^2b.2 is H, C _1-4 alkyl, in some cases H, methyl, ethyl, propyl;
The remaining residues are as defined above.
The cocrystal is administered to a subject.

Another aspect of the invention is further a co-crystal of a compound of formula 2a,
In the formula,
R ^2a is H, C _1-4 alkyl, and in some cases methyl, ethyl, propyl;
R ^2b is H, CONR ^2b.1 R ^2b.2 ;
R ^2b.1 is C _1-4 haloalkyl;
R ^2b.2 is H, C _1-4 alkyl, in some cases H, methyl, ethyl, propyl;
The remaining residues are as defined above.
The cocrystal is administered to a subject.

Another aspect of the invention is further a co-crystal of a compound of formula 2a,
In the formula,
R ^2b.1 and R ^2b.2 together are a C _3-6 alkylene group forming a heterocycle with the nitrogen atom, optionally one carbon atom of the ring being replaced by an oxygen atom;
The remaining residues are as defined above.
The cocrystal is administered to a subject.

The free base of the compound of formula 2 (j=0) is often amorphous and is used in the process of making the co-crystal, while in some cases, a salt of the compound of formula 2 is used in the process of making the co-crystal. Thus, another aspect of the invention is a salt of the compound of formula 2, wherein R ¹ , m, R ^2a , R ^2b , R ³ are as defined above for the co-crystal, X is an anion selected from the group consisting of chloride, bromide, iodide, sulfate, phosphate, methanesulfonate, nitrate, maleate, acetate, benzoate, citrate, salicylate, fumarate, tartrate, dibenzoyltartrate, oxalate, succinate, benzoate and p-toluenesulfonate, in some cases chloride or dibenzoyltartrate, and j is 0, 0.5, 1, 1.5 or 2, in some cases 1 or 2.

Another aspect of the invention further includes administering to a subject a co-crystal of a compound of formula 2, wherein R ¹ , m, R ^2a , R ^2b , and R ³ are as defined above for the co-crystal, X is an anion selected from the group consisting of chloride or dibenzoyltartrate, and j is 1 or 2.

Another aspect of the invention further includes administering to a subject a salt of a compound of formula 2, wherein R ¹ , m, R ^2a , R ^2b , and R ³ are as defined above for salts, X is chloride, and j is 2.

Another aspect of the invention further includes administering to a subject a salt of a compound of formula 2, wherein R ¹ , m, R ^2a , R ^2b , and R ³ are as defined above for salts, X is dibenzoyl tartrate, and j is 1.

Another aspect of the invention further includes administration to a subject of a salt of a compound of formula 2a below, wherein R ^2a , R ^2b , R ³ , X and j are as defined above.

Another aspect of the invention is further a salt of a compound of formula 2a,
In the formula,
R ^2a is H, C _1-4 alkyl, and in some cases methyl, ethyl, propyl;
R ^2b is H, CONR ^2b.1 R ^2b.2 ;
R ^2b.1 is C _1-4 alkyl, in some cases methyl, ethyl, propyl;
R ^2b.2 is C _1-4 alkyl, in some cases methyl, ethyl, propyl;
The remaining residues are as defined above.
The method includes administering a salt to a subject.

Another aspect of the invention is further a salt of a compound of formula 2a,
In the formula,
R ^2a is H, C _1-4 alkyl, and in some cases methyl, ethyl, propyl;
R ^2b is H, CONR ^2b.1 R ^2b.2 ;
R ^2b.1 is C _0-4 alkyl-C _3-6 cycloalkyl;
R ^2b.2 is H, C _1-4 alkyl, in some cases H, methyl, ethyl, propyl;
The remaining residues are as defined above.
The method includes administering a salt to a subject.

Another aspect of the invention is further a salt of a compound of formula 2a,
In the formula,
R ^2a is H, C _1-4 alkyl, and in some cases methyl, ethyl, propyl;
R ^2b is H, CONR ^2b.1 R ^2b.2 ;
R ^2b.1 is C _1-4 haloalkyl;
R ^2b.2 is H, C _1-4 alkyl, in some cases H, methyl, ethyl, propyl;
The remaining residues are as defined above.
The method includes administering a salt to a subject.

Another aspect of the invention is further a salt of a compound of formula 2a,
wherein R ^2b.1 and R ^2b.2 together are a C _3-6 alkylene group forming a heterocyclic ring with the nitrogen atom, optionally with one carbon atom of the ring replaced by an oxygen atom, and the remaining residues are as defined above, comprising administering to a subject a salt thereof.

Another aspect of the invention further includes administering to a subject a salt of a compound of formula 2a, wherein R ¹ , m, R ^2a , R ^2b , and R ³ are as defined above for that salt, X is chloride, and j is 2.

Another aspect of the invention further includes administration to a subject of a salt of a compound of formula 2a, wherein R ¹ , m, R ^2a , R ^2b , R ³ are as defined above for said salts, X is dibenzoyl tartrate, and j is 1. Another aspect of the invention is a salt of a compound of formula 2a, wherein R ¹ , m, R ^2a , R ^2b , R ³ are as defined above for said salts, X is (S)-(S)-(+)-2,3-dibenzoyl-tartrate, and j is 1.

c. Formulations Additional embodiments of the present invention further include pharmaceutical compositions comprising a compound of formula 3:

In the formula,
R ¹ is H, C _1-6 alkyl, C _0-4 alkyl-C _3-6 cycloalkyl, C _1-6 haloalkyl;
^R2 is H, _C1-6 alkyl;
X is an anion selected from the group consisting of chloride or 1/2 dibenzoyltartrate;
j is 1 or 2,
This includes administration to a subject.

An embodiment of the present invention is a pharmaceutical composition comprising a compound of formula 3,
In the formula,
^R1 is H, _C1-6 alkyl;
^R2 is H, _C1-6 alkyl;
X is an anion selected from the group consisting of chloride or 1/2 dibenzoyltartrate;
j is 1 or 2,
This includes administration to a subject.

An embodiment of the present invention is a pharmaceutical composition comprising a compound of formula 3,
In the formula,
R ¹ is H, methyl, ethyl, propyl, butyl;
^R2 is H, methyl, ethyl, propyl, butyl;
X is an anion selected from the group consisting of chloride or 1/2 dibenzoyltartrate, such as chloride;
j is 1 or 2, and in some cases, 2, of the pharmaceutical composition;
This includes administration to a subject.

An embodiment of the present invention is a pharmaceutical composition comprising a compound of formula 3,
In the formula,
R ¹ is H, methyl, ethyl, propyl, butyl;
^R2 is H, methyl;
X is an anion selected from the group consisting of chloride or 1/2 dibenzoyltartrate, such as chloride;
j is 1 or 2, and in some cases, 2, of the pharmaceutical composition;
This includes administration to a subject.

An embodiment of the present invention is a pharmaceutical composition comprising a compound of formula 3,
In the formula,
^R1 is H, methyl;
^R2 is H, methyl;
X is an anion selected from the group consisting of chloride or 1/2 dibenzoyltartrate, such as chloride;
j is 1 or 2, and in some cases, 2, of the pharmaceutical composition;
This includes administration to a subject.

Embodiments of the invention further include administering to a subject a pharmaceutical composition comprising a compound as a hydrochloride salt, as set forth in Table 2. Additional embodiments of the invention further include administering to a subject a pharmaceutical composition comprising a compound as a dihydrochloride salt, as set forth in Table 2.

Another object of the present invention is the administration to a subject of a pharmaceutical dosage form of the above compound, the pharmaceutical dosage form being an orally deliverable dosage form.

Another object of the present invention is the administration to a subject of a pharmaceutical dosage form of the above compound, the pharmaceutical dosage form being in the form of a tablet, capsule, pellet, powder or granule.

Another object of the present invention is the administration of the above pharmaceutical dosage form for use as a medicine to a subject.

Another object of the present invention is the use of the above pharmaceutical dosage form in the preparation of a medicament for treating a neurodegenerative disease or condition selected from Alzheimer's disease, Parkinson's disease, frontotemporal dementia, Huntington's disease, amyotrophic lateral sclerosis, multiple sclerosis, glaucoma, myotonic dystrophy, vascular dementia and progressive supranuclear palsy.

Another object of the present invention is a process for the treatment and/or prevention of a disease or condition selected from neurodegenerative diseases such as Alzheimer's disease, Parkinson's disease, frontotemporal dementia, Huntington's disease, amyotrophic lateral sclerosis, multiple sclerosis, glaucoma, myotonic dystrophy, vascular dementia and progressive supranuclear palsy, characterized in that an effective amount of a pharmaceutical dosage form as defined above is orally administered to a subject or patient once, twice, three times or several times per day.

d. Dosage Forms/Ingredients Solid pharmaceutical compositions ready for use/ingestion made from the compound of formula 3 include powders, granules, pellets, tablets, capsules, chewable tablets, dispersible tablets, troches and lozenges.
Capsule formulations according to the present invention include powdered intermediates of the compound of formula 3, intermediate blends containing the powdered intermediates, pellets or granules obtained by conventional wet granulation, dry granulation, hot melt granulation, hot melt extrusion or spray drying of suitable intermediate blends, which are filled into conventional capsules, e.g. hard gelatin capsules or HPMC capsules.
The capsule formulation obtained above may also contain a powdered intermediate of the compound of formula 3 in a compacted form.
Capsule formulations according to the present invention comprise a compound of formula 3 suspended or diluted in a liquid or mixture of liquids.
Tablet formulations according to the invention include tablets as obtained by direct compression of a suitable final blend or by tableting pellets or granules obtained by conventional wet granulation, dry granulation, hot melt granulation, hot melt extrusion or spray drying of a suitable intermediate blend.

Another object of the present invention is a dosage form to which a pH adjusting or buffering agent is added for improving the stability of the active ingredient. The pH adjusting/buffering agent can be a basic amino acid with an amino group and alkaline character (isoelectric point pI: 7.59-10.76), such as, for example, L-arginine, L-lysine or L-histidine. A buffering agent within the meaning of the present invention is L-arginine. L-arginine has a particularly favorable stabilizing effect on the composition of the present invention, for example by inhibiting the chemical decomposition of the compound of formula 3.

Thus, in an embodiment, the invention is directed to a pharmaceutical composition (e.g., an oral solid dosage form, particularly a tablet) comprising a compound of formula 3, and L-arginine, particularly for stabilizing the pharmaceutical composition against chemical degradation, and one or more pharmaceutical excipients.

Preferably, the pharmaceutical excipients used in the present invention are conventional substances such as cellulose and its derivatives, D-mannitol, corn starch, pregelatinized starch as fillers, copovidone as binders, crospovidone as disintegrants, magnesium stearate as lubricants, colloidal anhydrous silica as flow agents, hypromellose as film coating agents, polyethylene glycol as plasticizers, titanium dioxide, iron oxide red/yellow as pigments, and talc, etc.

In particular, the pharmaceutical excipients can be first and second diluents, binders, disintegrants, and lubricants, with additional disintegrants and additional glidants being further options.
- Diluents suitable for the pharmaceutical compositions according to the invention are cellulose powder, microcrystalline cellulose, lactose in various crystalline modifications, anhydrous dibasic calcium phosphate, dibasic calcium phosphate dihydrate , erythritol, low-substituted hydroxypropylcellulose, mannitol, starch or modified starch (e.g. pregelatinized starch or partially hydrolyzed starch) or xylitol. Among these diluents, mannitol and microcrystalline cellulose are employed in some cases.
- Diluents useful as the second diluent are the diluents mentioned above: mannitol and microcrystalline cellulose.
Suitable lubricants for the pharmaceutical compositions according to the invention are talc, polyethylene glycol, calcium behenate, calcium stearate, sodium stearyl fumarate, hydrogenated castor oil or magnesium stearate. The lubricant is in some cases magnesium stearate.
Suitable binders for the pharmaceutical composition according to the invention are copovidone (copolymer of vinylpyrrolidone with other vinyl derivatives), hydroxypropylmethylcellulose (HPMC), hydroxypropylcellulose (HPC), polyvinylpyrrolidone (povidone), pregelatinized starch, stearic acid-palmitic acid, low-substituted hydroxypropylcellulose (L-HPC), copovidone and pregelatinized starch, which are employed in some formulations. The abovementioned binders pregelatinized starch and L-HPC exhibit additional diluent and disintegrant properties and can also be used as a second diluent or second disintegrant.
Disintegrants suitable for the pharmaceutical composition according to the invention are corn starch, crospovidone, polacrilin potassium, croscarmellose sodium, low-substituted hydroxypropyl cellulose (L-HPC) or pregelatinized starch (such as croscarmellose sodium).
- Colloidal silicon dioxide can be used as an optional flow agent.

An exemplary composition according to the present invention includes the diluent mannitol, microcrystalline cellulose as a diluent with additional disintegration properties, the binder copovidone, the disintegrant croscarmellose sodium, and magnesium stearate as a lubricant.

Exemplary pharmaceutical compositions include:
Active ingredient: 10-50%
Diluent 1: 20-88%
Diluent 2: 5-50%
1-5% binder,
It contains 1-15% disintegrant and 0.1-5% lubricant (by weight).

The pharmaceutical composition according to some embodiments comprises:
Active ingredient: 10-50%
Diluent 1: 20-75%
Diluent 2: 5-30%
2-30% binder,
It contains 1-12% disintegrant and 0.1-3% lubricant (by weight).

The pharmaceutical composition according to some embodiments comprises:
Active ingredients: 10-90%
Diluent 1: 5-70%
Diluent 2: 5-30%
Binder: 0-30%
It contains 1-12% disintegrant and 0.1-3% lubricant (by weight).

The pharmaceutical composition according to some embodiments comprises:
Active ingredient: 10-50%
Diluent 1: 20-75%
Diluent 2: 5-30%
2-30% binder,
Buffer 0.5-20%
It contains 1-12% disintegrant and 0.1-3% lubricant (by weight).

The pharmaceutical composition according to some embodiments comprises:
30-70% active ingredient,
Diluent 1: 20-75%
Diluent 2: 5-30%
2-30% binder,
Buffer 0.5-20%
It contains 1-12% disintegrant and 0.1-3% lubricant (by weight).

In some cases, pharmaceutical compositions are employed that contain 10-90% active ingredient (e.g., 30-70% active ingredient) (by weight).

Tablet formulations according to the invention may be uncoated or coated, e.g., film coated, with a suitable coating known not to adversely affect the dissolution properties of the final formulation. For example, tablets may be seal coated for the protection of the patient's environment and clinical personnel, as well as for moisture protection purposes, by dissolving a high molecular weight polymer such as polyvinylpyrrolidone or hydroxypropyl-methylcellulose in water or an organic solvent such as acetone, along with a plasticizer, lubricant, and optionally a pigment and surfactant, and applying this mixture to the tablet core in a coating apparatus such as a pan coater or a fluid bed coater with a Wurster insert.

Additionally, agents such as beeswax, shellac, cellulose acetate phthalate, polyvinyl acetate phthalate, zein, film-forming polymers (such as hydroxypropylcellulose, ethylcellulose, and polymeric methacrylates) can be applied to the tablet, provided that the coating has no substantial effect on the disintegration/dissolution of the dosage form and that the stability of the coated dosage form is not affected.

After the dosage form is film coated, a sugar coating may be applied over the sealed pharmaceutical dosage form. The sugar coating may include sucrose, dextrose, sorbitol, etc., or mixtures thereof. If desired, colorants or opacifying agents may be added to the sugar solution.

The solid formulations of the present invention tend to be hygroscopic. They may be packaged using moisture-resistant packaging materials such as PVC-blisters, PVDC-blisters, or aluminum foil blister packs, alu/alu blisters, clear or opaque polymer blisters with pouches, polypropylene tubes, glass bottles and HDPE bottles, optionally including child-resistant features, or may be tamper-proof. The primary packaging material may contain desiccants such as molecular sieves or silica gel to improve the chemical stability of the API. Opaque packaging such as colored blister materials, tubes, amber glass bottles, etc. may be used to extend the shelf life of the API by reducing light degradation.

e. Dosage The dosage range of the compound of formula 3 is typically 100-1000 mg, particularly 200-900 mg, 300-900 mg, 350-850 mg or 390-810 mg. One or two tablets, in some cases two tablets, can be given at oral dosages of 100 mg, 200 mg, 300 mg, 350 mg, 400 mg, 450 mg, 500 mg, 550 mg, 600 mg, 650 mg, 700 mg, 750 mg, 800 mg, 850 mg, 900 mg per day, and in some cases 350 mg, 400 mg, 450 mg, 750 mg, 800 mg, 850 mg are employed.

The dosage range can be achieved by one tablet or two tablets, and in some cases two tablets are administered, each containing half the dose.

The active ingredient may be applied up to three times daily (such as once or twice daily). Specific dosage strengths are 400 mg or 800 mg.

f. Terms and Definitions Used Terms not specifically defined herein are to be given the meaning that would be given to them by one of ordinary skill in the art in light of the disclosure and context. However, as used herein, unless otherwise specified to the contrary, the following terms have the meanings indicated and are subject to the following conventions:

The term "about" means ±5% of the indicated value. Thus, about 100 minutes can also be read as 95-105 minutes.

The compounds of the invention are shown in the form of chemical names and as chemical formulas, with the chemical formulas taking precedence in the event of any discrepancy. In the subordinate chemical formulas, asterisks may be used to indicate bonds connecting the compounds to the parent molecule, as defined.

Unless otherwise indicated, throughout this specification and the appended claims, a given chemical formula or name is intended to include tautomers, all stereoisomers, optical isomers and geometric isomers (e.g., enantiomers, diastereomers, E/Z isomers, etc.), their racemates, mixtures in which the separate enantiomers exist in various ratios, mixtures of diastereomers, or mixtures of any of the above forms in which such isomers and enantiomers exist, as well as salts, including pharma- ceutically acceptable salts, and solvates thereof, such as, for example, hydrates (including solvates of the free compounds and solvates of salts of the compounds).

The term "substituted," as used herein, means that one or more hydrogens on any of the designated atoms have been replaced with one selected from the indicated group, provided that the normal valence of the designated atom is not exceeded and that the substitution results in a stable compound.

The term "optionally substituted" means within the scope of the present invention a group as mentioned above, optionally substituted by a lower molecular group. Examples of lower molecular groups that are considered chemically meaningful are groups consisting of 1-200 atoms. Applicable are those groups that do not adversely affect the pharmacological efficacy of the compounds of the present invention. For example, such groups may include:
- Straight or branched carbon chains optionally interrupted by heteroatoms and optionally substituted by rings, heteroatoms or other common functional groups.
- An aromatic or non-aromatic ring system consisting of carbon atoms and optionally heteroatoms, which may further be substituted by functional groups.
- A substantial number of aromatic or non-aromatic ring systems consisting of carbon atoms and, optionally, heteroatoms, which may be linked by one or more carbon chains, optionally interrupted by heteroatoms and optionally substituted by heteroatoms or other common functional groups.

The compounds disclosed herein can exist as therapeutically acceptable salts. The invention includes the compounds listed above in the form of salts, including acid addition salts. Suitable salts include salts formed with both organic and inorganic acids. Such acid addition salts will usually be pharma-ceutically acceptable. However, salts that are not pharma-ceutically acceptable may be useful in the preparation and purification of the compounds. Base addition salts may also be formed and may be pharma-ceutically acceptable. For a more complete discussion of salt preparation and selection, see Properties, Selection, and Use (Stahl, P. Heinrich. Wiley-VCHA, Zurich, Switzerland, 2002).

The term "therapeutically acceptable salts," as used herein, refers to salts or zwitterionic forms of the compounds disclosed herein that are water-soluble, oil-soluble, water-dispersible, or oil-dispersible and are therapeutically acceptable as defined herein. The salts can be prepared during the final isolation and purification of the compounds of the invention, or separately, by reacting the appropriate compound in the form of the free base with a suitable acid. Representative acid addition salts include acetate, adipate, alginate, L-ascorbate, aspartate, benzoate, benzenesulfonate (besylate), bisulfate, butyrate, camphorate, camphorsulfonate, citrate, digluconate, formate, fumarate, gentisate, glutarate, glycerophosphate, glycolate, hemisulfate, heptanoate, hexanoate, hippurate, hydrochloride, hydrobromide, hydroiodide, 2-hydroxyethanesulfonate (isethionate), lactate, maleate, and malonate. salts, DL-mandelate, mesitylenesulfonate, methanesulfonate, naphthylenesulfonate, nicotinate, 2-naphthalenesulfonate, oxalate, pamoate, pectinate, persulfate, 3-phenylpropionate, phosphonate, picrate, pivalate, propionate, pyroglutamate, succinate, sulfonate, tartrate, L-tartrate, trichloroacetate, trifluoroacetate, phosphate, glutamate, bicarbonate, para-toluenesulfonate (p-tosylate) and undecanoate. The basic groups of the compounds disclosed herein can also be quaternized with methyl, ethyl, propyl and butyl chlorides, bromides and iodides, dimethyl, diethyl, dibutyl and diamyl sulfates, decyl, lauryl, myristyl and steryl chlorides, bromides and iodides, and benzyl and phenethyl bromides. Examples of acids that can be used to form therapeutically acceptable addition salts include inorganic acids such as hydrochloric acid, hydrobromic acid, sulfuric acid, and phosphoric acid, and organic acids such as oxalic acid, maleic acid, succinic acid, and citric acid. Salts can also be formed by coordinating the compounds with alkali metal or alkaline earth ions. Thus, the present invention contemplates sodium, potassium, magnesium, calcium, and the like salts of the compounds disclosed herein.

Base addition salts can be prepared during the final isolation and purification of the compounds of the invention by reacting the carboxyl group with a suitable base, such as the hydroxide, carbonate or bicarbonate of a metal cation, or ammonia, an organic primary amine, secondary amine or tertiary amine. Therapeutically acceptable salt cations include lithium, sodium, potassium, calcium, magnesium and aluminum, as well as non-toxic quaternary amine cations such as ammonium, tetramethylammonium, tetraethylammonium, methylamine, dimethylamine, trimethylamine, triethylamine, diethylamine, ethylamine, tributylamine, pyridine, N,N-dimethylaniline, N-methylpiperidine, N-methylmorpholine, dicyclohexylamine, procaine, dibenzylamine, N,N-dibenzylphenethylamine, 1-ephenamine and N,P-dibenzylethylenediamine. Other representative organic amines useful for the formation of base addition salts include ethylenediamine, ethanolamine, diethanolamine, piperidine and piperazine.

Although it may be possible for the compounds of the present invention to be administered as the raw chemical, they may also be provided as pharmaceutical formulations. Thus, what is provided herein is a pharmaceutical formulation comprising one or more specific compounds disclosed herein, or one or more pharma- ceutically acceptable salts, esters, prodrugs, amides, or solvates thereof, together with one or more pharma- ceutically acceptable carriers thereof, and optionally one or more other therapeutic ingredients. The carrier(s) must be "acceptable" in the sense of being compatible with the other ingredients of the formulation and not deleterious to the recipient thereof. Appropriate formulations will depend on the chosen route of administration. Any of the known techniques, carriers, and excipients may be used as suitable and understood in the art, e.g., Remington's Pharmaceutical Sciences. The pharmaceutical compositions disclosed herein may be manufactured in any form known in the art, e.g., by conventional mixing, dissolving, granulating, dragee-making, triturating, emulsifying, encapsulating, entrapping, or compressing processes.

"Heterocycle" ("het") includes 5-, 6-, or 7-membered saturated or unsaturated heterocycles, or 5-10 membered bicyclic heterocycles, which may contain 1, 2, or 3 heteroatoms selected from oxygen, sulfur, and nitrogen, and the ring may be bonded to the molecule by a carbon atom, or, if present, a nitrogen atom. Below are examples of 5-, 6-, or 7-membered saturated or unsaturated heterocycles:

Unless otherwise specified, the heterocycle may contain a keto group. Examples include:

Examples of 5-10 membered bicyclic heterocycles are pyrrolidine, indole, indolizine, isoindole, indazole, purine, quinoline, isoquinoline, benzimidazole, benzofuran, benzopyran, benzothiazole, benzisothiazole, pyridopyrimidine, pteridine, pyrimidopyrimidine.

The term "heterocycle" includes heteroaromatic groups, while the term "heteroaromatic group" ("hetaryl") refers to a 5- or 6-membered heteroaromatic group, which may contain 1, 2 or 3 heteroatoms selected from oxygen, sulfur and nitrogen, or a 5- to 10-membered bicyclic hetaryl ring, which contains sufficient conjugated double bonds that an aromatic system is formed. The ring may be bonded to the molecule by a carbon atom or, if present, a nitrogen atom. The following are examples of 5- or 6-membered heteroaromatic groups:

Examples of 5- to 10-membered bicyclic hetaryl rings include pyrrolidine, indole, indolizine, isoindole, indazole, purine, quinoline, isoquinoline, benzimidazole, benzofuran, benzopyran, benzothiazole, benzisothiazole, pyridopyrimidine, pteridine, and pyrimidopyrimidine.

The term "halogen" as used herein means a halogen substituent selected from fluoro, chloro, bromo, or iodo.

The term "C _1-6 alkyl" (including those which are part of other groups) refers to branched and unbranched alkyl groups having 1 to 6 carbon atoms, and the term "C _1-4 alkyl" refers to branched and unbranched alkyl groups having 1 to 4 carbon atoms. In some cases, alkyl groups having 1 to 4 carbon atoms are present. Examples of these include methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, sec-butyl, tert-butyl, n-pentyl, isopentyl, neopentyl or hexyl. For the abovementioned groups, the abbreviations Me, Et, n-Pr, i-Pr, n-Bu, i-Bu, t-Bu, etc. may also be used optionally. Unless otherwise stated, the definitions propyl, butyl, pentyl and hexyl include all possible isomeric forms of the groups in question. Thus, for example, propyl includes n-propyl and isopropyl, butyl includes isobutyl, sec-butyl and tert-butyl, etc.

The term "C _1-6 alkylene" (including those which are part of other groups) means branched and unbranched alkylene groups having 1 to 6 carbon atoms, and the term "C _1-4 alkylene" means branched and unbranched alkylene groups having 1 to 4 carbon atoms. In some cases, alkylene groups having 1 to 4 carbon atoms are present. Examples include methylene, ethylene, propylene, 1-methylethylene, butylene, 1-methylpropylene, 1,1-dimethylethylene, 1,2-dimethylethylene, pentylene, 1,1-dimethylpropylene, 2,2-dimethylpropylene, 1,2-dimethylpropylene, 1,3-dimethylpropylene or hexylene. Unless otherwise stated, the definitions propylene, butylene, pentylene and hexylene also include all possible isomers of the related groups with the same number of carbons. Thus, for example, propyl also includes 1-methylethylene, butylene includes 1-methylpropylene, 1,1-dimethylethylene, and 1,2-dimethylethylene.

The term "C _2-6 alkenyl" (including those which are part of other groups) denotes branched and unbranched alkenyl groups having 2 to 6 carbon atoms, and the term "C _2-4 alkenyl" denotes branched and unbranched alkenyl groups having 2 to 4 carbon atoms, provided that they have at least one double bond. In some cases, what is used is an alkenyl group having 2 to 4 carbon atoms. Examples include ethenyl, i.e. vinyl, propenyl, butenyl, pentenyl or hexenyl. Unless otherwise stated, the definitions of propenyl, butenyl, pentenyl and hexenyl include all possible isomeric forms of the groups in question. Thus, for example, propenyl includes 1-propenyl and 2-propenyl, butenyl includes 1-butenyl, 2-butenyl, 3-butenyl, 1-methyl-1-propenyl, 1-methyl-2-propenyl, etc.

The term "C _2-6 alkenylene" (including those which are part of other groups) means branched and unbranched alkenylene groups having 2 to 6 carbon atoms, and the term "C _2-4 alkenylene" means branched and unbranched alkylene groups having 2 to 4 carbon atoms. In some cases, alkenylene groups having 2 to 4 carbon atoms are present. Examples include ethenylene, propenylene, 1-methylethenylene, butenylene, 1-methylpropenylene, 1,1-dimethylethenylene, 1,2-dimethylethenylene, pentenylene, 1,1-dimethylpropenylene, 2,2-dimethylpropenylene, 1,2-dimethylpropenylene, 1,3-dimethylpropenylene or hexenylene. Unless stated otherwise, the definitions propenylene, butenylene, pentenylene and hexenylene include all possible isomers of the respective groups with the same number of carbons. Thus, for example, propenyl also includes 1-methylethenylene, and butenylene includes 1-methylpropenylene, 1,1-dimethylethenylene and 1,2-dimethylethenylene.

The term "C _2-6 alkynyl" (including those that are part of other groups) refers to branched and unbranched alkynyl groups having 2 to 6 carbon atoms, and the term "C _2-4 alkynyl" refers to branched and unbranched alkynyl groups having 2 to 4 carbon atoms, provided that they have at least one triple bond. In some cases, alkynyl groups having 2 to 4 carbon atoms are present. Examples include ethynyl, propynyl, butynyl, pentynyl or hexynyl. Unless otherwise stated, the definitions of propynyl, butynyl, pentynyl and hexynyl include all possible isomeric forms of the respective groups. Thus, for example, propynyl includes 1-propynyl and 2-propynyl, butynyl includes 1-butynyl, 2-butynyl, 3-butynyl, 1-methyl-1-propynyl, 1-methyl-2-propynyl, etc.

The term "C _2-6 alkynylene" (including those which are part of other groups) means branched and unbranched alkynylene groups having 2 to 6 carbon atoms, and the term "C _2-4 alkynylene" means branched and unbranched alkynylene groups having 2 to 4 carbon atoms. In some cases, alkynylene groups having 2 to 4 carbon atoms are present. Examples include ethynylene, propynylene, 1-methylethynylene, butynylene, 1-methylpropynylene, 1,1-dimethylethynylene, 1,2-dimethylethynylene, pentynylene, 1,1-dimethylpropynylene, 2,2-dimethylpropynylene, 1,2-dimethylpropynylene, 1,3-dimethylpropynylene or hexynylene. Unless stated otherwise, the definitions propynylene, butynylene, pentynylene and hexynylene include all possible isomers of the respective groups with the same number of carbons. Thus, for example, propynyl also includes 1-methylethynylene, butynylene includes 1-methylpropynylene, 1,1-dimethylethynylene and 1,2-dimethylethynylene.

The term "C _3-6 cycloalkyl" (including those that are part of other groups) as used herein means cyclic alkyl groups having from 3 to 8 carbon atoms, and in some cases such groups are cyclic alkyl groups having from 5 to 6 carbon atoms. Examples include cyclopropyl, cyclobutyl, cyclopentyl or cyclohexyl.

The term "C _1-6 haloalkyl" (including those which are part of other groups) refers to branched and unbranched alkyl groups having 1 to 6 carbon atoms in which one or more hydrogen atoms have been replaced by halogen atoms selected from fluorine, chlorine or bromine (such as fluorine and chlorine, e.g. fluorine). Correspondingly, "C _1-4 haloalkyl" refers to branched and unbranched alkyl groups having 1 to 4 carbon atoms in which one or more hydrogen atoms have been replaced by the atoms mentioned above. In some cases, C _1-4 haloalkyls are present. Examples include CH ₂ F, CHF ₂ , CF ₃ .

The term "C _1-n alkyl", alone or in combination with other radicals, where n is an integer from 2 to n, refers to an acyclic branched or straight-chain saturated hydrocarbon radical having 1 to n carbon atoms. For example, the term "C _1-5 alkyl" includes H ₃ C-, H ₃ C-CH ₂ -, H ₃ C-CH ₂ -CH ₂ -, H ₃ C-CH(CH ₃ )-, H ₃ C-CH ₂ -CH ₂ -CH ₂ -, H ₃ C-CH ₂ -CH(CH ₃ )-, H ₃ C-CH(CH3) _-CH2- , H3C _- C( _CH3 ) _2- , _{H3C - CH2} - _CH2- CH2 _{-CH2- ,} H3C _{-CH2-CH2-CH(CH3)-, H3C-CH2-CH(CH3)-CH2 -, H 3 C - CH ( CH 3 ) -CH 2 -CH 2} These include the radicals --, _H3C -- _CH2 --C( _CH3 ) _2-- , _H3C --C( _CH3 ) ₂ -- _CH2-- , _H3C --CH( _CH3 )--CH( _CH3 )-- and _H3C -- _CH2 -- _CH ( _CH2CH3 )--.

The term "C _1-n haloalkyl", where n is an integer from 2 to n, alone or in combination with another radical, refers to an acyclic branched or straight chain saturated hydrocarbon radical having 1 to n carbon atoms in which one or more hydrogen atoms are replaced by a halogen atom selected from fluorine, chlorine or bromine (such as fluorine and chlorine, e.g., fluorine). Examples include CH ₂ F, CHF ₂ , CF ₃ .

The term "C _1-n alkylene", alone or in combination with other radicals, refers to an acyclic straight or branched chain divalent alkyl radical containing 1 to n carbon atoms, where n is an integer from 2 to n. For example, the term "C _1-4 alkylene" includes -CH ₂ -, -CH ₂ -CH ₂ -, -CH(CH ₃ )-, -CH ₂ -CH ₂ -CH ₂ -, -C(CH ₃ ) ₂ -, -CH(CH ₂ CH ₃ ) _- , -CH(CH ₃ )-CH ₂ -, -CH ₂ -CH ₍ CH3)-, -CH2 _{- CH2} -CH2 _- CH2-, -CH2 _- CH2-CH( _CH3 )-, _-CH2 - _{CH3 ) -CH2} -CH2-, -CH2-CH _{( CH3} )-CH2- _{, -CH2} -C(CH ₃ ) ₂ -, -C(CH ₃ ) ₂ -CH ₂ -, -CH(CH ₃ )-CH(CH ₃ )-, -CH ₂ -CH(CH ₂ CH ₃ )-, -CH(CH ₂ CH ₃ )-CH ₂ -, -CH(CH ₂ CH ₂ CH ₃ )-, -CH(CH(CH ₃ )) ₂ - and -C(CH ₃ )(CH ₂ CH ₃ )- is included.

The term "C2 _-n alkenyl" is used for groups defined similarly to "C1 _-n alkyl" having at least two carbon atoms, where at least two of the carbon atoms of the group are joined to each other by a double bond.

The term "C _2-n alkynyl" is used to refer to groups defined similarly to the definition of "C _1-n alkyl" having at least two carbon atoms, where at least two of those carbon atoms of the group are joined to each other by a triple bond.

The term "C _3-n cycloalkyl," where n is an integer from 4 to n, alone or in combination with other radicals, refers to a cyclic, unbranched, saturated hydrocarbon radical having from 3 to n carbon atoms. For example, the term C _3-7 cycloalkyl includes cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl and cycloheptyl.

It should be noted that, as used in this specification and the appended claims, the singular forms "a," "an," and "the" include plural references unless the context clearly dictates otherwise. Thus, for example, a reference to a "cell" includes a plurality of such cells, and a reference to a "peptide" includes a reference to one or more peptides and equivalents thereof known to those of skill in the art, such as polypeptides.

By "individual suffering from or at risk of suffering from age-related cognitive impairment" is meant an individual who is greater than about 50% of their life expectancy (such as greater than 60% of their life expectancy, e.g., greater than 70%, greater than 75%, greater than 80%, greater than 85%, greater than 90%, greater than 95% or even greater than 99%). The age of the individual will depend on the species in question. Thus, this percentage is based on the predicted life expectancy of the species in question. For example, in humans, such individuals are 50 years of age or older, e.g., 60 years of age or older, 70 years of age or older, 80 years of age or older, 90 years of age or older, and usually up to 100 years of age (such as 90 years of age), i.e., between about 50 and 100 years of age, e.g., 50 years, 51 years, 52 years, 53 years, 54 years, 55 years, 56 years, 57 years, 58 years, 60 years, 61 years, 62 years, 63 years, or older. , 64, 65, 66, 67, 68, 69, 70, 71, 72, 73, 74, 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, 85, 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, 100 years old or older, or any age between 50 and 100 years old, who have not yet begun to show symptoms of an age-related condition, such as cognitive impairment 50 years or older, e.g., 60 years or older, 70 years or older, 80 years or older, 90 years or older, and usually 100 years or younger, i.e., about 50 to 100 years old, e.g., 50, 51, 52, 53, 54, 55, 56, 57, 58, 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70, 71, 72, 73, 74, 75, 76, 77, 78, 79, 80, 81, 82, Individuals who are 83, 84, 85, 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, 100 years old, individuals of any age who suffer from cognitive impairment due to an age-related disease as further described below, and individuals of any age who have been diagnosed with an age-related disease typically associated with cognitive impairment, but have not yet begun to show symptoms of cognitive impairment. The corresponding ages for non-human subjects are known and are intended to apply in the present invention.

As generally outlined elsewhere, in some cases, the subject is a mammal. Mammalian species that may be treated with the methods of the invention include canines and felines, equines, bovines, sheep, etc., as well as primates, including humans. The methods, compositions, and reagents of the invention may also be applied to animal models, including small mammals, e.g., murines, lagomorphs, etc., for example, in experimental investigations.

As used herein and as described above, "treatment" refers to either (i) the prevention of a disease or disorder, or (ii) the reduction or elimination of symptoms of a disease or disorder. Treatment may be performed as a prophylaxis (before the onset of the disease) or as a treatment (after the onset of the disease). The action may be prophylactic in that it completely or partially prevents the disease or its symptoms, and/or it may be therapeutic in that it partially or completely cures the disease and/or the adverse effects caused by the disease. Thus, the term "treatment" as used herein encompasses the treatment of any age-related disease or disorder in a mammal, including (a) preventing the onset of the disease in a subject who may have a predisposition to the disease but has not yet been diagnosed as suffering from the disease, (b) inhibiting the disease, i.e., arresting the onset of the disease, or (c) relieving the disease, i.e., regressing the disease. Treatment may result in a wide variety of physical manifestations, such as modulation of gene expression, rejuvenation of tissues or organs, etc. The therapeutic agent of the present invention may be administered before, during, or after the onset of the disease. Of particular interest is the treatment of ongoing disease, which stabilizes or alleviates undesirable clinical symptoms in the patient. Such treatment may be administered prior to complete loss of function of affected tissues. Applicable therapies may be administered during, and in some cases after, the symptomatic phase of the disease.

In some embodiments, the condition to be treated is age-related impairment of cognitive abilities of an individual. Cognitive abilities or "cognition" refers to mental processes including attention and concentration, learning complex tasks and concepts, memory (acquiring, retaining and recalling new information in the short and/or long term), information processing (processing information collected by the senses), visuospatial functions (seeing, depth perception, using mental imagery, copying pictures, constructing objects or shapes), language production and comprehension, verbal fluency (finding words), problem solving, decision making, and executive functions (planning and prioritization). "Cognitive decline" refers to a progressive decline in one or more of these abilities, e.g., memory, language, thinking, judgment, etc. "Cognitive impairment" and "cognitive impairment" refer to a decline in cognitive abilities compared to a healthy individual, e.g., an age-matched healthy individual, or compared to the individual's abilities at a previous time, e.g., 2 weeks, 1 month, 2 months, 3 months, 6 months, 1 year, 2 years, 5 years, 10 years or more ago. "Age-related cognitive impairment" refers to cognitive impairments typically associated with aging, including, for example, cognitive impairments associated with the natural aging process, such as mild cognitive impairment (MCI), and age-related disorders, i.e. disorders whose incidence increases with the progression of aging, such as cognitive impairments associated with neurodegenerative conditions, such as Alzheimer's disease, Parkinson's disease, frontotemporal dementia, Huntington's disease, amyotrophic lateral sclerosis, multiple sclerosis, glaucoma, myotonic dystrophy, vascular dementia, progressive supranuclear palsy, ataxia, and associated frailty.

g. Combinations The compounds of formula 1 may be used alone or in combination with other active substances of formula 1 according to the present invention. The compounds of formula 1 may also be optionally combined with other pharmacologically active substances. These include: beta2 adrenoceptor agonists (short and long acting), anticholinergics (short and long acting), anti-inflammatory steroids (oral and topical corticosteroids), cromoglycates, methylxanthines, dissociating glucocorticoid mimetics, PDE3 inhibitors, PDE4 inhibitors, PDE7 inhibitors, LTD4 antagonists, EGFR inhibitors, dopamine agonists, PAF antagonists, lipoxin A4 derivatives, FPRL1 modulators, LTB4 receptor (BLT1, BLT2) antagonists, histamine H1 receptor antagonists, histamine H4 receptor antagonists, dual histamine H1/H3 receptor antagonists, PI3 kinase inhibitors, inhibitors of non-receptor tyrosine kinases (e.g. LYN, LCK, SYK, ZAP-70, FYN, BTK or ITK), MAP kinases (e.g. p3 8, ERK1, ERK2, JNK1, JNK2, JNK3, or SAP), NF-κB signaling inhibitors (e.g., IKK2 inhibitors), iNOS inhibitors, MRP4 inhibitors, leukotriene biosynthesis inhibitors (e.g., 5-lipoxygenase (5-LO) inhibitors), cPLA2 inhibitors, leukotriene A4 hydrolase inhibitors or FLAP inhibitors, nonsteroidal anti-inflammatory drugs (NSAIDs), CR TH2 antagonists, DP1 receptor modulators, thromboxane receptor antagonists, additional CCR3 antagonists, CCR4 antagonists, CCR1 antagonists, CCR5 antagonists, CCR6 antagonists, CCR7 antagonists, CCR8 antagonists, CCR9 antagonists, CCR30 antagonists, CXCR3 antagonists, CXCR4 antagonists, CXCR ² antagonists, CXCR1 antagonists, CXCR5 antagonists, CXCR6 antagonists, CX3CR3 antagonists, neurokinin (NK1, NK2) antagonists, sphingosine 1-phosphate receptor modulators, sphingosine 1-phosphate lyase inhibitors, adenosine receptor inhibitors (e.g., A2a agonists), purinergic receptor modulators (e.g., P2X7 inhibitors), histone deacetylase (HDAC) activators, bradykinin (BK1, BK2) antagonists, TACE inhibitors, PPAR gamma modulators, Rho kinase inhibitors, interleukin 1-beta converting enzyme (ICE) inhibitors, Toll-like receptor (TLR) modulators, HMG-CoA reductase inhibitors anti-inflammatory drugs, VLA-4 antagonists, ICAM-1 inhibitors, SHIP agonists, GABAa receptor antagonists, ENaC inhibitors, melanocortin receptor (MC1R, MC2R, MC3R, MC4R, MC5R) modulators, CGRP antagonists, endothelin antagonists, TNFα antagonists, anti-TNF antibodies, anti-GM-CSF antibodies, anti-CD46 antibodies, anti-IL-1 antibodies, anti-IL-2 antibodies, anti-IL-4 antibodies, anti-IL-5 antibodies, anti-IL-13 antibodies, anti-IL-4/IL-13 antibodies, anti-TSLP antibodies, anti-OX40 antibodies, mucoregulators, immunotherapeutic agents, compounds against airway swelling, compounds against cough, VEGF inhibitors, and also combinations of two or three active substances.

In some embodiments, the other active agents are betamimetics, anticholinergics, corticosteroids, PDE4 inhibitors, LTD4 antagonists, EGFR inhibitors, CRTH2 inhibitors, 5-LO inhibitors, histamine receptor antagonists and SYK inhibitors, as well as combinations of two or three active agents, i.e.,
- combinations of betamimetics with corticosteroids, PDE4 inhibitors, CRTH2 inhibitors or LTD4 antagonists; - combinations of anticholinergics with betamimetics, corticosteroids, PDE4 inhibitors, CRTH2 inhibitors or LTD4 antagonists; - combinations of corticosteroids with PDE4 inhibitors, CRTH2 inhibitors or LTD4 antagonists; - combinations of PDE4 inhibitors with CRTH2 inhibitors or LTD4 antagonists; - combinations of CRTH2 inhibitors with LTD4 antagonists.

In these embodiments, the compounds that make up the combination are co-administered to the subject. The terms "co-administration" and "in combination with" include administration of two or more therapeutic agents simultaneously, simultaneously, or sequentially, without particular time limitations. In one embodiment, the agents are present simultaneously in a cell or in the subject's body, or exert their biological or therapeutic actions simultaneously. In one embodiment, the therapeutic agents are present in the same composition or unit dosage form. In another embodiment, the therapeutic agents are present in separate compositions or separate unit dosage forms. In certain embodiments, a first therapeutic agent can be administered prior to (e.g., minutes, 15 minutes, 30 minutes, 45 minutes, 1 hour, 2 hours, 4 hours, 6 hours, 12 hours, 24 hours, 48 hours, 72 hours, 96 hours, 1 week, 2 weeks, 3 weeks, 4 weeks, 5 weeks, 6 weeks, 8 weeks, or 12 weeks before) administration of a second therapeutic agent, in conjunction with administration of the second therapeutic agent, or after (e.g., 5 minutes, 15 minutes, 30 minutes, 45 minutes, 1 hour, 2 hours, 4 hours, 6 hours, 12 hours, 24 hours, 48 hours, 72 hours, 96 hours, 1 week, 2 weeks, 3 weeks, 4 weeks, 5 weeks, 6 weeks, 8 weeks, or 12 weeks after) administration of the second therapeutic agent. "Concomitant administration" of a known therapeutic agent and a pharmaceutical composition of the present disclosure means administering a compound of the present invention and a second agent at a time such that both the known agent and the composition of the present disclosure have a therapeutic effect. Such concomitant administration may include administering the agent simultaneously (i.e., simultaneously) with the administration of the compound of the present invention, before the administration of the compound of the present invention, or after the administration of the compound of the present invention. The routes of administration of the two agents may vary, and representative routes of administration are described in further detail below. A person skilled in the art would have no difficulty in determining the appropriate timing, sequence, and dosage of administration of a particular drug and a compound of the present disclosure. In some embodiments, the compounds (e.g., a compound of the present invention and at least one additional compound) are administered to a subject within 24 hours of administration of the other (e.g., within 12 hours of administration of the other, within 6 hours of administration of the other, within 3 hours of administration of the other, or within 1 hour of administration of the other). In certain embodiments, the compounds are administered within 1 hour of administration of the other. In certain embodiments, the compounds are administered substantially simultaneously. By substantially simultaneous administration, it is meant that the compounds are administered to the subject within about 10 minutes of one another (e.g., within 5 minutes or within 1 minute of the other).

"Companion diagnostic" or "companion diagnostic device" means an in vitro diagnostic device or imaging tool that provides information essential to the safe and effective use of a corresponding therapeutic product. The use of an in vitro diagnostic companion device with a particular therapeutic product is specified in the instructions for use in the labeling of both the device and the corresponding therapeutic product, as well as in the labeling of any generic equivalents and biologically similar equivalents of the therapeutic product.

Companion diagnostic tests can be in several forms, including, by way of example only, but not limited to, tests that screen for familial genetic patterns and hard-to-diagnose conditions, prognostic tests that predict the future course of a disease, theranostic tests that indicate a patient's response to a prescribed therapy, monitoring tests that evaluate the efficacy and proper dosing of a prescribed therapy, and recurrence tests that analyze a patient's risk of disease recurrence. See Agarwal A, et al., Pharmgenomics Pers Med. 8:99-110 (2015), which is incorporated herein by reference in its entirety.

h. Pharmaceutical Forms Suitable preparations for administering the compounds of Formula 1 and the co-crystal or salt forms of Formula 2 and Formula 2a include, for example, tablets, capsules, suppositories, liquids and powders. The content of the pharma- ceutical active compound(s) should be in the range of 0.05-90% by weight (such as 0.1-50% by weight) of the total composition. Suitable tablets may be obtained, for example, by mixing the active substance(s) with known excipients, for example, inert diluents such as calcium carbonate, calcium phosphate or lactose, disintegrating agents such as corn starch or alginic acid, binding agents such as starch or gelatin, lubricants such as magnesium stearate or talc, and/or agents that delay release, such as carboxymethylcellulose, cellulose acetate phthalate or polyvinyl acetate. The tablet may also include several layers.

Thus, coated tablets may be prepared by coating a tablet core, prepared similarly to a tablet, with a substance usually used for tablet coating, such as Kollidon or shellac, gum arabic, talc, titanium dioxide or sugar. The tablet core may also consist of a number of layers, to provide delayed release or to prevent incompatibilities. Similarly, the tablet coating may consist of a number of layers, optionally with the excipients mentioned above for the tablets, to provide delayed release.

Syrups or elixirs containing the active substances or combinations thereof according to the invention may further contain sweeteners such as saccharin, cyclamate, glycerol or sugar, and flavour enhancers, for example flavourings such as vanillin or orange extract. These preparations may also contain suspension adjuvants, thickeners such as sodium carboxymethylcellulose, wetting agents, for example condensates of fatty acid alcohols with ethylene oxide, or preservatives such as p-hydroxybenzoates.

The solutions are prepared in the usual manner, for example with the addition of an isotonicity agent, a preservative such as p-hydroxybenzoate or a stabilizer such as an alkali metal salt of ethylenediaminetetraacetic acid, and optionally with an emulsifier and/or dispersant, while, for example, an organic solvent may optionally be used as a solubilizer or dissolution aid when water is used as the diluent, and the solutions may be transferred into injection vials, injection ampoules or infusion bottles.

Capsules containing one or more active substances or combinations of active substances can be prepared, for example, by mixing the active substances with an inert carrier such as lactose or sorbitol and encapsulating the mixture in a gelatin capsule.

Suitable suppositories can be prepared, for example, by mixing with carriers provided for suppositories, such as neutral fats or polyethylene glycol or their derivatives.

Excipients that may be used include, for example, water, pharma- ceutically acceptable organic solvents such as paraffins (e.g., petroleum fractions), vegetable oils (e.g., peanut oil or sesame oil), monofunctional or polyfunctional alcohols (e.g., ethanol or glycerol), carriers such as natural mineral powders (e.g., kaolin, clay, talc, chalk), synthetic mineral powders (e.g., highly dispersed silicic acid and silicates), sugars (e.g., cane sugar, lactose and glucose), emulsifiers (e.g., lignin, spent sulfite liquor, methylcellulose, starch and polyvinylpyrrolidone), and lubricants (e.g., magnesium stearate, talc, stearic acid and sodium lauryl sulfate).

For oral use, tablets may of course contain, in addition to the specified carriers, additives such as sodium citrate, calcium carbonate and dicalcium phosphate, together with various additional substances such as starches, e.g. potato starch, gelatin, and the like. Lubricants such as magnesium stearate, sodium lauryl sulfate and talc may also be used to prepare tablets. In the case of aqueous suspensions, the active substance may be combined with various flavor enhancers or colorings, in addition to the above excipients.

For administration of the compounds of formula 1, or the co-crystal or salt forms of formula 2 and formula 2a, preparations or pharmaceutical formulations suitable for inhalation may be used. Inhalable preparations include inhalable powders, propellant-containing metered dose aerosols or propellant-free inhalable solutions. Within the scope of the present invention, the term propellant-free inhalable solutions also includes concentrates or sterile inhalable solutions ready for use. Formulations that may be used within the scope of the present invention are described in more detail in the following sections of this specification.

Inhalable powders that may be used according to the invention may contain the compound of formula 1, or the co-crystal or salt forms of formula 2 and formula 2a, either alone or in admixture with suitable physiologically acceptable excipients.

If the active substance of formula 1 or the co-crystal or salt form of formula 2 and formula 2a is present in admixture with a physiologically acceptable excipient, these inhalable powders according to the invention may be prepared using physiologically acceptable excipients such as monosaccharides (e.g. glucose or arabinose), disaccharides (e.g. lactose, saccharose, maltose), oligosaccharides and polysaccharides (e.g. dextran), polyhydric alcohols (e.g. sorbitol, mannitol, xylitol), salts (e.g. sodium chloride, calcium carbonate) or mixtures of these excipients. In some cases, mono- or disaccharides such as lactose or glucose are used, for example in the form of their hydrates, e.g. lactose as lactose monohydrate.

Within the scope of the inhalable powders according to the invention, the maximum average particle size of the excipients is at most 250 μm (such as 10-150 μm), including 15-80 μm. It may also seem appropriate to add finer excipient portions with an average particle size of 1-9 μm to the above-mentioned excipients. These finer excipients are also selected from the group of possible excipients listed above. Then, to prepare the inhalable powders according to the invention, a micronized active substance of the compound of formula 1 or of the co-crystal or salt form of formula 2 and formula 2a, such as having an average particle size of 0.5-10 μm (including 1-5 μm), is added to the excipient mixture. The processes of making the inhalable powders according to the invention by grinding and micronization and finally mixing the components together are known from the prior art.

The inhalable powders according to the invention can be administered using inhalers known from the prior art.

The inhalation aerosols containing a gas propellant according to the invention may contain the compound of formula 1 or the co-crystal or salt form of formula 2 and formula 2a dissolved or in dispersed form in the gas propellant. The compound of formula 1 or the co-crystal or salt form of formula 2 and formula 2a may be contained in separate formulations or in a common formulation, either with both components dissolved, either with both components dispersed, or in each case only one component dissolved and the other dispersed. Gas propellants which may be used to prepare the inhalation aerosols are known from the prior art. Suitable gas propellants are selected from hydrocarbons such as n-propane, n-butane or isobutane, and halohydrocarbons such as fluorinated derivatives of methane, ethane, propane, butane, cyclopropane or cyclobutane. The above gas propellants may be used alone or in mixtures. In some cases the gas propellant is a halogenated alkane derivative selected from TG134a and TG227, and mixtures thereof.

Inhalation aerosols utilizing propellants may also contain other ingredients such as co-solvents, stabilizers, surfactants, antioxidants, lubricants and pH adjusters, all of which are known in the art.

The propellant-based inhalation aerosols according to the invention described above may be administered using inhalers known in the art (MDI = metered dose inhaler). Furthermore, the active substances of the compounds of formula 1 or the co-crystal or salt forms of formula 2 and formula 2a according to the invention may be administered in the form of propellant-free inhalable solutions and suspensions. The solvent used may be an aqueous solvent or an alcoholic solvent such as an ethanol solution. The solvent may be water alone or a mixture of water and ethanol. There is no limit to the relative proportion of ethanol to water, but in some cases the maximum is up to 70 volume percent (including up to 30 volume percent, such as up to 60 volume percent). The remaining volume is made up of water. The solutions or suspensions containing the compounds of formula 1 or the co-crystal or salt forms of formula 2 and formula 2a are adjusted to a pH of 2-7 (such as 2-5) using a suitable acid. The pH may be adjusted using an acid selected from inorganic or organic acids. Examples of particularly suitable inorganic acids include hydrochloric acid, hydrobromic acid, nitric acid, sulfuric acid and/or phosphoric acid. Particularly suitable examples of organic acids include ascorbic acid, citric acid, malic acid, tartaric acid, maleic acid, succinic acid, fumaric acid, acetic acid, formic acid and/or propionic acid. In some cases, the inorganic acid is hydrochloric acid and sulfuric acid. It is also possible to use an acid that already forms an acid addition salt with one of the active substances of the invention. Among the organic acids, in some cases, ascorbic acid, fumaric acid and citric acid are used. If desired, mixtures of the above-mentioned acids may be used, especially in the case of acids that, in addition to their acidifying properties, have other properties, for example as flavoring agents, antioxidants or complexing agents, such as citric acid or ascorbic acid. According to the invention, in some cases, hydrochloric acid is used to adjust the pH.

If desired, these formulations may omit the addition of edetic acid (EDTA) or one of its known salts, sodium edetate, as a stabilizer or complexing agent. Other embodiments may include this or these compounds. In embodiments, the ratio to sodium edetate is less than 100 mg/100 ml (such as less than 50 mg/100 ml, including less than 20 mg/100 ml). In some cases, inhalable solutions are used that have a ratio of sodium edetate between 0 and 10 mg/100 ml. Propellant-free inhalable solutions may contain cosolvents and/or other excipients, such as alcohols, particularly isopropyl alcohol, glycols, particularly propylene glycol, polyethylene glycol, polypropylene glycol, glycol ethers, glycerol, polyoxyethylene alcohols and polyoxyethylene fatty acid esters. The term excipients and additives in this context refers to any pharmacologically acceptable substance that is not an active substance but can be formulated with an active substance or active substances in a physiologically suitable solvent to improve the qualitative properties of the active substance formulation. In some embodiments, these substances have no pharmacological effect or no appreciable or at least no undesirable pharmacological effect in relation to the desired therapy. These excipients and additives include, for example, surfactants such as soy lecithin, oleic acid, sorbitan esters (such as polysorbates), polyvinylpyrrolidone, other stabilizers, complexing agents, antioxidants and/or preservatives that ensure or extend the shelf life of the final pharmaceutical formulation, flavoring agents, vitamins and/or other additives known in the art. Additives also include pharmacologically acceptable salts such as sodium chloride as isotonicity agents.

In some embodiments, excipients include antioxidants such as ascorbic acid (e.g., provided that it is not already being used to adjust pH), vitamin A, vitamin E, tocopherols, and vitamins and provitamins similar to those found in the human body.

Preservatives may be used to protect the formulation from contamination with pathogens. Suitable preservatives are those known in the art, in particular acetylpyridinium chloride, benzalkonium chloride, or benzoic acid or benzoate salts (such as sodium benzoate) in concentrations known from the prior art. The above preservatives may be present in a concentration of up to 50 mg/100 ml (such as 5-20 mg/100 ml). In some embodiments, the formulations of the invention contain only benzalkonium chloride and sodium edetate in addition to the solvent water and the compound of formula 1, or the co-crystal or salt form of formulas 2 and 2a. In an embodiment, sodium edetate is not present.

The dosage of the compounds according to the invention will of course depend to a large extent on the method of administration and the condition being treated. When administered by inhalation, the compounds of formula 1, or the co-crystal or salt forms of formula 2 and formula 2a, are characterized by high efficacy even at doses in the μg range. The compounds of formula 1, or the co-crystal or salt forms of formula 2 and formula 2a, may also be used effectively in the μg range. Thus, dosages may be, for example, in the gram range.

In another aspect, the present invention relates to a pharmaceutical formulation as described above, characterized in that it comprises a compound of formula 1, or a co-crystal or salt form of formula 2 and formula 2a, in particular a pharmaceutical formulation as described above that can be administered by inhalation.

The following formulation examples are intended to illustrate the present invention without limiting its scope.

i. Examples of pharmaceutical preparations A) Tablets 100 mg of active substance 1, 2 or 2a per tablet
Lactose 140mg
Corn starch 240mg
Polyvinylpyrrolidone 15mg
Magnesium stearate 5mg
500mg

The finely milled active substance, lactose and part of the corn starch are mixed together. The mixture is sieved, then moistened with a solution of polyvinylpyrrolidone in water, kneaded, wet granulated and dried. The granules, the remaining corn starch and magnesium stearate are sieved and mixed together. The mixture is compressed into tablets of suitable shape and size.

B) Tablets: 80 mg of active substance 1, 2 or 2a per tablet
Lactose 55mg
Corn starch 190mg
Microcrystalline cellulose 35mg
Polyvinylpyrrolidone 15mg
Sodium carboxymethyl starch 23mg
Magnesium stearate 2mg
400mg

The finely ground active substance, a portion of the corn starch, lactose, microcrystalline cellulose and polyvinylpyrrolidone are mixed together, the mixture is sieved and processed with the remaining corn starch and water to form granules, which are dried and sieved. Sodium carboxymethyl starch and magnesium stearate are added and mixed, and the mixture is compressed to form tablets of suitable size.

C) Liquid ampoule containing 50 mg of active substance 1, 2 or 2a
Sodium chloride 50mg
Water for injection 5ml

The active substance is dissolved in water at its original pH or optionally at pH 5.5-6.5 and sodium chloride is added to make an isotonic solution. The resulting solution is filtered to remove pyrogens and the filtrate is transferred under aseptic conditions into ampoules which are then sterilized and heat sealed. The ampoules contain 5 mg, 25 mg and 50 mg of active substance.

D) Metered dose aerosol Active substance 1, 2 or 2a 0.005
Sorbitan trioleate 0.1
Monofluorotrichloromethane and TG134a:TG227 (2:1) added to 100

The suspension is placed into a conventional aerosol container equipped with a metered dose valve. Preferably, each actuation releases 50 μl of suspension. If desired, higher doses (e.g., 0.02% by weight) of the active agent may be released.

E) Liquid (unit: mg/100 ml)
Active substance 1, 2 or 2a 333.3 mg
Benzalkonium chloride 10.0 mg
EDTA 50.0mg
Add HCl (1N) until pH is 2.4. This solution can be prepared in the usual way.

F) Inhalable powder 12 μg of active substance 1, 2 or 2a
Lactose monohydrate 25 mg added Inhalable powders are prepared in the usual manner by mixing the individual components.

j. Indications Provided are methods of improving cognition or other symptoms of dementia through treating a subject/patient diagnosed with a cognitive-related disease. Aspects of the method include modulating CCR3, e.g., by a CCR3 modulator, in a manner sufficient to treat a patient with a cognitive-related disease. The method includes treating the cognitive-related disease with an orally administrable and bioavailable composition (including a compound of Formula 1, a co-crystal or salt composition of Formula 2 or Formula 2a), or a formulation of Formula 3 as described above. As generally described above and in further detail below, various age-related dysfunctions, e.g., cognitive-related diseases, may be treated by embodiments of the present invention. In some cases, the target condition is a cognitive-related disease condition associated with neurodegeneration, e.g., evidenced by neuronal insufficiency, such as reduced neurogenesis (e.g., as evidenced by reduced numbers of BrdU or EdU positive cells, Ki67 positive cells, and Dcx positive cells compared to non-diseased tissue). Compositions that modulate CCR3 can be administered to patients/subjects diagnosed with a cognition-related disease such as mild cognitive impairment (MCI), Alzheimer's disease, Parkinson's disease, frontotemporal dementia (FTD), Huntington's disease, amyotrophic lateral sclerosis (ALS), multiple sclerosis (MS), glaucoma, myotonic dystrophy, dementia, progressive supranuclear palsy (PSP), ataxia, multiple system atrophy and frailty, examples of which are not limited to those described further below. The methods of the invention can further comprise monitoring the improvement of the progression of the neurodegenerative disease through measuring improvements in cognition or physical function.

A method is provided for improving motor coordination, motor function, or other symptoms of a movement disorder through treating a subject/patient diagnosed with a movement disorder. Aspects of the method include modulating CCR3, e.g., by a CCR3 modulator, in a manner sufficient to treat a patient with a movement disorder. The method includes treating the movement disorder with an orally administrable and bioavailable composition (including a composition of a compound of Formula 1, a cocrystal or salt of Formula 2 or Formula 2a), or a formulation of Formula 3 as described above. As generally described above and in further detail below, various age-related dysfunctions, e.g., movement disorders, may be treated by embodiments of the present invention. In some cases, the target condition is a movement disorder associated with neurodegeneration, e.g., as evidenced by neuronal insufficiency, such as reduced neurogenesis (e.g., as evidenced by reduced numbers of BrdU or EdU positive cells, Ki67 positive cells, and Dcx positive cells compared to non-affected tissue). The CCR3 modulating composition can be administered to a patient/subject diagnosed with a movement disorder such as Parkinson's disease, parkinsonism, dementia with Lewy bodies, ataxia, dystonia, cervical dystonia, chorea, Huntington's disease, multiple system atrophy, spasticity, progressive supranuclear palsy, tardive dyskinesia, Tourette's syndrome, and tremor, which are examples, but are not limited to, as described further below. The method of the invention can further include monitoring the improvement of the progression of the neurodegenerative disease through measuring improvements in cognitive or physical function.

Mild cognitive impairment (MCI) is a mild disruption of cognition that manifests as problems with memory, or other mental functions such as planning, understanding instructions, or decision-making, that worsen over time, without a decline in overall mental function and daily activities. Thus, while significant neuronal death typically does not occur, neurons in the aging brain are susceptible to sublethal age-related changes in structure, synaptic integrity, and molecular processing at the synapses, all of which reduce cognitive function. Individuals suffering from or at risk of developing age-related cognitive impairment who would benefit from treatment with the compounds of the invention, e.g., the methods disclosed herein, include individuals of any age suffering from cognitive impairment due to age-related disorders, and individuals of any age who have been diagnosed with an age-related disorder typically associated with cognitive impairment, but who have not yet begun to show symptoms of cognitive impairment. Examples of such age-related disorders include, but are not limited to, those listed below.

Alzheimer's disease: Alzheimer's disease is characterized by progressive and irreversible cognitive loss associated with an excessive number of senile plaques in the cerebral cortex and subcortical gray matter, as well as excess beta-amyloid and neurofibrillary tangles composed of tau protein. Its most common form affects people over the age of 60, and its prevalence increases with age. Alzheimer's disease accounts for more than 65% of dementia in older adults.

The cause of Alzheimer's disease is unknown. The disease is familial in about 15-20% of cases. The remaining, so-called sporadic cases, have some genetic link. The disease has an autosomal dominant inheritance pattern in most early-onset cases and in some late-onset cases, with variable penetrance in later stages. Environmental factors are the focus of ongoing research.

During the course of the disease, synaptic and ultimately neuronal loss is seen in the cerebral cortex, hippocampus and subcortical structures (including selective cell loss in the nucleus basalis of Meynert), locus coeruleus and dorsal raphe nucleus. Cerebral glucose utilization and perfusion is decreased in some brain regions (parietal and temporal cortex in early disease and prefrontal cortex in late disease). Neuritic or senile plaques (composed of neurites, astrocytes and glial cells around an amyloid core) and neurofibrillary tangles (composed of paired helical fibrils) play important roles in the pathogenesis of Alzheimer's disease. Senile plaques and neurofibrillary tangles occur during normal aging but are much more common in people with Alzheimer's disease.

Parkinson's Disease: Parkinson's disease (PD) is an idiopathic, slowly progressive, degenerative CNS disorder characterized by slow and reduced movement (bradykinesia), muscle rigidity, resting tremor (dystonia), muscle rigidity and postural instability. PD was initially viewed as primarily a movement disorder, but it is now recognized that it also causes depressive symptoms and emotional changes. PD can also affect cognition, behavior, sleep, autonomic function and sensory function. The most common cognitive impairments include impairments in attention and concentration, working memory, executive function, language production and visuospatial function. PD is characterized by symptoms associated with decreased motor function that usually precede symptoms associated with cognitive impairment, which aids in diagnosing the disease.

Primary Parkinson's disease involves the loss of pigmented neurons in the substantia nigra, locus coeruleus, and other brainstem dopaminergic cell groups. The cause is unknown. Loss of nigral neurons radiating to the caudate nucleus and putamen leads to depletion of the neurotransmitter dopamine in these areas. Onset is generally after age 40, with an increased prevalence in older age groups.

Approximately 60,000 Americans are newly diagnosed with Parkinson's disease each year, and approximately 1 million Americans are currently affected. While PD itself is not fatal, its complications are the 14th leading cause of death in the United States. Currently, PD is incurable, and treatments are generally prescribed to control symptoms and, in late-stage, severe cases, surgery.

Treatment options for PD include the administration of medications to help manage motor deficits. These options increase or replace the neurotransmitter dopamine, which is at low levels in the brain in PD patients. These include carbidopa/levodopa (increases dopamine production in the brain), apomorphine, pramipexole, ropinirole and rotigotine (dopamine agonists), selegiline and rasagiline (MAO-B inhibitors that prevent the breakdown of dopamine), entacapone and tolcapone (catechol-O-methyltransferase [COMT] inhibitors that allow more levodopa to be utilized in the brain), benztropine and trihexyphenidyl (anticholinergics), and amantadine (controls tremors and rigidity). Exercise/physical therapy is also commonly used to help maintain physical and mental function.

However, current treatment options treat the symptoms of PD, are not curative, and do not prevent disease progression. In addition, current drugs tend to lose efficacy in late-stage PD. The most commonly prescribed drug, levodopa, typically produces adverse effects within 5-10 years of taking it. These adverse effects can be significant, resulting in motor fluctuations between doses, unpredictable fluctuations in motor control, and difficult to manage, as well as disabling jerks/twitching movements (dyskinesias) similar to the symptoms of PD itself. Thus, there remains a need for new therapeutic agents with new mechanisms of action that can be administered alone or in combination with current PD drugs.

Parkinsonism: Secondary parkinsonism (also called atypical parkinsonism or parkinson plus) results from loss or disruption of dopamine action in the basal ganglia due to other idiopathic degenerative diseases, drugs, or exogenous toxins. The most common cause of secondary parkinsonism is the ingestion of antipsychotics or reserpine, which block dopamine receptors and thus produce parkinsonism. Less common causes include carbon monoxide poisoning, manganese poisoning, hydrocephalus, structural lesions (tumors, infarctions affecting the midbrain or basal ganglia), subdural hematomas, and degenerative disorders including striatonigral degeneration. Certain disorders such as progressive supranuclear palsy (PSP), multiple system atrophy (MSA), corticobasal degeneration (CBD) and dementia with Lewy bodies (DLB) may be classified as "parkinsonism" because they may present with symptoms of parkinsonism before the presenting symptoms required for a specific diagnosis can appear.

Assessment of PD progression Several assessment scales have been used to assess the progression of PD. The most widely used scales include the Unified Parkinson's Disease Rating Scale (UPDRS, introduced in 1987) (J. Rehabil Res. Dev., 2012 49(8)1269-76) and the Horn-Yahr scale (Neurology, 1967 17(5):427-42). Additional scales include the Movement Disorder Society (MDS) updated UPDRS scale (MDS-UPDRS) and the Schwab England Activities of Daily Living (ADL) scale.

The UPDRS scale assesses 31 items divided into three subscales: (1) mental function, behavior, and mood, (2) activities of daily living, and (3) motor function tests. The Horn-Yahr scale categorizes PD into five stages with substages: 0 (no signs of disease), 1 (unilateral symptoms), 1.5 (unilateral symptoms, but also cervical and spinal), 2 (bilateral symptoms, but no balance impairment), 2.5 (mild symptoms on both sides, able to bounce back when performing a "pull" test), 3 (mild to moderate disease with balance impairment), 4 (severely impaired, but able to walk or stand without assistance), and 5 (requires wheelchair or is bedridden without assistance). The Schwab-England scale categorizes PD into several percentages, from 100% (fully independent) to 10% (fully dependent).

Frontotemporal Dementia: Frontotemporal dementia (FTD) is a condition resulting from the progressive deterioration of the frontal lobes of the brain. Over time, atrophy can progress to the temporal lobes. FTD is second only to Alzheimer's Disease (AD) in prevalence and accounts for 20% of presenile dementia cases. Symptoms are classified into three groups based on which frontal and temporal lobe functions are affected:

Behavioral-type FTD (bvFTD), with symptoms including lethargy and loss of spontaneity on the one hand and disinhibition on the other; progressive non-fluent aphasia (PNFA), where speech fluency disorders due to articulation difficulties, phonological paraphasias and/or syntactic errors are observed but word comprehension is preserved; and semantic dementia (SD), where patients retain normal phonological and syntactic fluency but develop worsening naming and word comprehension disorders. Other cognitive symptoms common to all FTD patients include executive dysfunction and concentration disorders. Other cognitive abilities including perception, spatial abilities, memory and executive typically remain intact. FTD can be diagnosed by observing atrophy of the frontal and/or anterior temporal lobes on structural MRI scans.

There are numerous forms of FTD, any of which may be treated or prevented using the methods and compositions of the present invention. For example, one form of frontotemporal dementia is semantic dementia (SD). SD is characterized by loss of semantic memory in both verbal and non-verbal domains. Patients with SD often present with word-finding difficulties. Clinical signs include fluent aphasia, naming aphasia, impaired word comprehension, and associative visual agnosia (inability to match semantically related images or objects). Cases of "pure" semantic dementia have been described that have few late behavioral symptoms, but as the disease progresses, behavioral and personality changes similar to those seen in frontotemporal dementia are often seen. Structural MRI imaging shows a characteristic pattern of atrophy in the temporal lobe (predominantly on the left side), with greater inferior than superior temporal lobe damage, and greater anterior than posterior temporal lobe atrophy.

As another example, another form of frontotemporal dementia is Pick's disease (PiD, also known as PcD). The defining feature of this disease is the accumulation of tau protein in neurons, which accumulates in silver-staining, spherical aggregates known as "Pick's globules." Symptoms include loss of speech (aphasia) and dementia. Patients with orbitofrontal dysfunction can become aggressive and socially inept.

Patients with Pick's disease may steal or engage in compulsive or repetitive stereotyped behaviors. Patients with dorsomedial or dorsolateral frontal dysfunction may exhibit apathy, blunted affect, or reduced spontaneity. Patients may exhibit lack of self-monitoring, abnormal self-awareness, and an inability to grasp meaning.

Patients with loss of gray matter in the bilateral posterolateral orbitofrontal cortex and right anterior insula may exhibit altered eating behaviors, such as a pathological sweet preference. Patients with more focal loss of gray matter in the anterior lateral orbitofrontal cortex may develop hyperphagia. Although some symptoms may be alleviated initially, the disease progresses and patients often die within 2 to 10 years.

Huntington's Disease: Huntington's disease (HD) is an inherited progressive neurodegenerative disorder characterized by the development of emotional, behavioral, and psychiatric disorders, loss of intellectual or cognitive function, and movement abnormalities (dyskinetic movement). Classical signs of HD include the development of chorea (involuntary rapid and irregular jerking movements that may affect the face, arms, legs, or trunk) and cognitive decline, including gradual loss of thought processing and acquired intellectual abilities. Impairment of memory, abstract thinking, and judgment, loss of perception of time, place, or identity (disorientation), increased agitation, and personality changes (disorganization) may also be present. Symptoms typically become evident during the fourth or fifth decade, but age of onset is variable, ranging from early childhood to late adulthood (e.g., the seventh or eighth decade).

HD is inherited in families as an autosomal dominant trait. The disorder results from an abnormally long sequence of coded instructions, or "repeats," in a gene on chromosome 4 (4p16.3). The progressive loss of nervous system function associated with HD results from the loss of neurons in specific areas of the brain, including the basal ganglia and cerebral cortex.

Amyotrophic Lateral Sclerosis: Amyotrophic Lateral Sclerosis (ALS) is a rapidly progressive, universally fatal neurological disease that attacks motor neurons. Muscle weakness and atrophy, as well as signs of anterior horn cell dysfunction, are most often seen early in the hands and less commonly in the feet. Site of attack is random and progression is asymmetric. Seizures are common and may precede weakness. Patients rarely survive 30 years; 50% die within 3 years of onset, 20% survive 5 years, and 10% survive 10 years.

Diagnostic features include onset in mid- or late adulthood and progressive generalized movement disorder without sensory abnormalities. Nerve conduction velocities are normal until late in the disease. Recent studies have also documented a picture of cognitive dysfunction, particularly declines in immediate verbal memory, visual memory, language, and executive function.

Reductions in cell body area, number of synapses, and total synaptic length have been reported in neurons that appear normal in patients with ALS. It has been suggested that functional impairment may occur due to continued loss of synapses once active zone plasticity reaches its limit. Promotion of new synapse formation or prevention of synapse loss may preserve neuronal function in these patients.

Multiple Sclerosis: Multiple sclerosis (MS) is characterized by a variety of symptoms and signs of CNS dysfunction with remissions and relapsing exacerbations. The most commonly seen symptoms are paresthesias in one or more limbs, the trunk, or one side of the face, weakness or clumsiness of the legs or hands, or visual impairment, e.g., partial blindness and pain in one eye (retrobulbar optic neuritis), blurred vision, or scotoma. Common cognitive dysfunction includes impairments in memory (acquisition, retention, and recall of new information), attention and concentration (especially divided attention), information processing, executive function, visuospatial function, and verbal fluency. Common early symptoms are ophthalmoparesis leading to double vision (double vision), transient weakness of one or more limbs, slight rigidity or unusual tiredness of the limbs, slight gait disturbance, difficulty with bladder control, dizziness, and mild affective disturbance, all of which indicate diffuse CNS dysfunction and often occur months or years before the disease is recognized. Excess heat can worsen symptoms and signs.

The course is highly variable and unpredictable, and remittent in most patients. Initially, there may be months or years of remission between episodes of symptoms, especially when the disease begins with retrobulbar optic neuritis. However, a minority of patients have frequent and rapidly incapacitating attacks, and the course may progress rapidly.

Glaucoma: Glaucoma is a common neurodegenerative disease that affects retinal ganglion cells (RGCs). Evidence supports the existence of a compartmentalized degenerative program in synapses and dendrites, including those of RGCs. Recent evidence also shows a correlation between cognitive impairment and glaucoma in older adults (Yochim BP, et al. Prevalence of cognitive impairment, depression, and anxiety symptoms among older adults with glaucoma. J Glaucoma. 2012; 21(4): 250-254).

Myotonic dystrophy: Myotonic dystrophy (DM) is an autosomal dominant, multisystem disorder characterized by dystrophic muscle weakness and myotonia. The molecular defect is an expansion of a trinucleotide repeat (CTG) in the 3' untranslated region of the myotonin protein kinase gene on chromosome 19. Symptoms can occur at any age and range in clinical severity. Myotonia is prominent in the hand muscles, and even in milder cases, ptosis is common. In severe cases, there is marked peripheral muscle weakness, often accompanied by cataracts, premature balding, axe-like facies, arrhythmias, testicular atrophy, and endocrine abnormalities (e.g. diabetes mellitus). In severe congenital forms, mental retardation is common, whereas in milder adult forms of the disorder, age-related decline in frontal and temporal cognitive functions, especially language and executive functions, is observed. Severely affected individuals die by their early 50s.

Dementia: Dementia is a type of disorder in which symptoms affect thinking and social skills severely enough to interfere with daily functioning. In addition to the dementia observed in the later stages of age-related disorders discussed above, other cases of dementia include vascular dementia and dementia with Lewy bodies, which are described below.

In vascular dementia or "multi-infarct dementia," cognitive impairment is due to problems with the blood supply to the brain, typically resulting from a series of minor strokes, or occasionally one major stroke preceded and followed by other, less severe strokes. Vascular pathology may be due to diffuse cerebrovascular disease, such as small vessel disease, or focal pathology, or both. Patients with vascular dementia present with acute or subacute cognitive impairment following an acute cerebrovascular event, after which progressive cognitive decline is observed. Cognitive impairment is similar to that observed in Alzheimer's disease, including impairments in language, memory, complex visual processing, or executive functions, but the associated brain changes are due to a chronic reduction in blood flow in the brain, rather than to AD pathology, which ultimately leads to dementia. Neuroimaging with single photon emission computed tomography (SPECT) and positron emission tomography (PET) may be used in conjunction with an evaluation with mental status tests to confirm the diagnosis of multi-infarct dementia.

Dementia with Lewy Bodies: Dementia with Lewy Bodies (DLB), also known by a variety of other names including dementia with Lewy bodies, diffuse Lewy body disease, cortical Lewy body disease, and senile dementia of the Lewy type, is a type of dementia characterized anatomically by the presence of Lewy bodies (clusters of alpha-synuclein and ubiquitin proteins) in neurons that are detectable on postmortem brain histology. Its main feature is cognitive decline, particularly of executive functions. Alertness and short-term memory can fluctuate.

Persistent or recurrent visual hallucinations with vivid and detailed images are often early diagnostic symptoms. DLB is often confused with Alzheimer's disease and/or vascular dementia in its early stages, but Alzheimer's disease usually begins in a fairly gradual manner, whereas DLB often has a rapid or acute onset. DLB symptoms also include motor symptoms similar to those of Parkinson's disease. DLB is differentiated from dementia, which can occur in Parkinson's disease, by the time frame in which the dementia symptoms appear compared to Parkinson's symptoms. Parkinson's disease with dementia (PDD) is diagnosed when the onset of dementia occurs more than one year after the onset of Parkinson's symptoms. DLB is diagnosed when cognitive symptoms begin at the same time as or within one year of Parkinson's symptoms.

Treatment of DLB is a complex process and requires a multifaceted approach. (Neurology, 2017 89:1-13). Typical parkinsonism therapeutic agents such as dopaminergic and anticholinergic drugs can worsen cognitive and behavioral symptoms. Optimal treatment generally uses both pharmacological approaches (exercise, cognitive training and caregiver training) as well as non-pharmacological approaches. For cognitive symptoms, acetylcholinesterase inhibitors (e.g., rivastigmine, donepezil) can be administered, as can memantine, an NMDA receptor antagonist. For neuropsychiatric symptoms, acetylcholinesterase inhibitors can improve affective blunting and hallucinations. Antipsychotics unfortunately increase the risk of death in DLB patients. In DLB patients, motor symptoms are less responsive to dopaminergic treatment and may worsen the risk of psychosis. Levodopa can be used, but only at low threshold doses, so there is a clear need in the art for new agents to treat DLB.

Progressive Supranuclear Palsy: Progressive supranuclear palsy (PSP) is a brain disorder that causes serious progressive problems with controlling walking and balance, along with complex eye movements and thinking problems. One of the classic signs of the disease is the inability to properly aim the eyes, which occurs due to lesions in the brain areas that coordinate eye movements. Some individuals describe this effect as blurred vision. Affected individuals often exhibit mood and behavior changes, including depression and emotional blunting, as well as progressive mild dementia. The disorder's long name indicates that the disease begins slowly, continues to worsen (progressive), and causes weakness (paralysis) by damaging specific brain parts (supranuclear) above pea-sized structures called nuclei that control eye movements. PSP was first described as a distinct disorder in 1964, when three scientists published a paper distinguishing the condition from Parkinson's disease. PSP is sometimes called Steele-Richardson-Olszewski syndrome, reflecting a combination of the names of the scientists who defined the disorder. PSP progressively worsens, but no one is killed by it.

Ataxia: People with ataxia have problems with coordination because the parts of the nervous system that control movement and balance are affected. Ataxia can affect fingers, hands, arms, legs, body, speech, and eye movements. The term ataxia is often used to describe a condition of incoordination that may be associated with infection, injury, other disease, or degenerative changes in the central nervous system. Ataxia is also used to describe a group of specific degenerative disorders of the nervous system called hereditary and sporadic ataxias, which are the focus of the National Ataxia Foundation.

Multiple System Atrophy: Multiple System Atrophy (MSA) is a neurodegenerative disorder. MSA is associated with the degeneration of nerve cells in certain areas of the brain. This cell degeneration causes problems with movement, balance and other autonomic functions (such as bladder control or blood pressure regulation).

The cause of MSA is unknown, and no specific risk factors have been identified. Approximately 55% of cases occur in men, with the typical age of onset being in the late 50s to early 60s. MSA often presents with some of the same symptoms as Parkinson's disease; however, people with MSA generally respond minimally, if at all, to dopamine medications used for Parkinson's disease.

Dystonia: Dystonia is a condition involving sustained involuntary muscle contractions. These contractions may manifest as twisting, repetitive movements. The disorder may affect the entire body or specific body parts, and is referred to as generalized or focal dystonia (respectively). Cervical dystonia may cause long-lasting or intermittent contractions in the neck muscles. There is no cure for dystonia. Current therapies include carbidopa-levodopa, trihexyphenidyl, benztropine, tetrabenazine, diazepam, clonazepam, baclofen, physical therapy, speech therapy, stretching, massage, and invasive surgery.

Frailty: Frailty syndrome ("frailty") is a geriatric syndrome characterized by functional and physical decline, including reduced mobility, muscle weakness, physical retardation, lack of stamina, low physical activity, malnutrition and involuntary weight loss. Such decline is often associated with cognitive dysfunction and disease, such as cancer. However, frailty can occur in the absence of disease. Individuals affected by frailty are at increased risk of poor fracture outcomes, falls, disability, comorbidities and premature death (C. Buigües, et al. Effect of a Prebiotic Formulation on Frailty Syndrome: A Randomized, Double-Blind Clinical Trial, Int. J. Mol. Sci. 2016, 17, 932). In addition, individuals suffering from frailty have a higher incidence of high medical costs (ibid.).

General symptoms of frailty can be determined by certain types of tests. For example, involuntary weight loss involves a loss of at least 10 pounds or more than 5% of the previous year's weight, muscle weakness can be determined by a decrease in grip strength to the bottom 20% at baseline (adjusted for sex and BMI), physical retardation can be based on the time required to walk a 15-foot distance, lack of endurance can be determined by an individual's self-report of fatigue, and low physical activity can be measured using standardized questionnaires (Z. Palace et al., The Frailty Syndrome, Today's Geriatric Medicine 7(1), at 18 (2014)).

In some embodiments, the methods and compositions of the invention find use in slowing the progression of age-related cognitive, motor, or other age-related impairments. In other words, after treatment with the disclosed methods, the individual's cognitive, motor, or other abilities will decline more slowly than before treatment with the disclosed methods or in the absence of treatment with the disclosed methods. In some such cases, the treatment methods of the invention include measuring the progression of cognitive, motor, or other age-related decline after treatment and determining that the progression of decline has been reduced. In some such cases, the determination is made by comparison to a control, e.g., the rate of decline of the individual before treatment (e.g., as determined by prior measurement of cognitive, motor, or other age-related abilities at two or more time points prior to administration of a blood product of the invention).

The methods and compositions of the invention are also useful for stabilizing the cognitive, motor or other capabilities of an individual, e.g., an individual exhibiting or at risk of exhibiting age-related cognitive decline. For example, the individual may exhibit an age-related cognitive impairment, and the progression of cognitive impairment observed prior to treatment with the disclosed methods will be halted following treatment with the disclosed methods. As another example, the individual may be at risk for developing age-related cognitive decline (e.g., the individual may be over 50 years old or may have been diagnosed with an age-related disorder), and the individual's cognitive performance is substantially unchanged following treatment with the disclosed methods compared to prior to treatment with the disclosed methods. That is, no cognitive decline is detectable.

The methods and compositions of the invention are also useful for alleviating cognitive, motor or other age-related impairments in individuals suffering from age-related impairments. In other words, the affected ability is improved in the individual following treatment with the methods of the invention. For example, the cognitive ability of the individual is increased, for example, by 2-fold or more, 5-fold or more, 10-fold or more, 15-fold or more, 20-fold or more, 30-fold or more, or 40-fold or more (including 50-fold or more, 60-fold or more, 70-fold or more, 80-fold or more, 90-fold or more, or 100-fold or more) following treatment with the methods of the invention, compared to the cognitive ability observed in the individual prior to treatment with the methods of the invention.

In some cases, treatment with the methods and compositions of the invention restores cognitive, motor or other capabilities of an individual exhibiting age-related cognitive or motor decline, e.g., to the level the individual had when he or she was about 40 years of age or younger. In other words, inhibits cognitive or motor impairment.

k. Methods of Diagnosis and Monitoring Improvement of Neurodegenerative Diseases Those skilled in the art will recognize that the following types of assessments can be used alone or in combination among various methods for diagnosing and monitoring the progression and improvement of neurodegenerative diseases in subjects suffering from neurodegenerative diseases. The following types of methods are provided as examples and are not limited to the methods provided. Those skilled in the art will recognize that other disease monitoring methods will also be useful in practicing the present invention. These methods are also contemplated by the methods of the present invention.

i. General Cognition The methods of the present invention further include a method of monitoring the effect of a medication or treatment for treating cognitive impairment and/or age-related dementia on a subject, comprising comparing cognitive function before and after treatment. Those skilled in the art will recognize that there are known methods for assessing cognitive function. For example, but not limited to, the methods may include assessment of cognitive function based on medical history, family history, physical and neurological examination by a clinician specialized in dementia and cognitive function, laboratory testing, and neuropsychological evaluation. Additional embodiments contemplated by the present invention include assessment of consciousness, such as using the Glasgow Coma Scale (EMV), mental status testing, including Abbreviated Mental Test Score (AMTS) or Mini-Mental State Examination (MMSE) (Folstein et al., J. Psychiatr. Res 1975; 12: 1289-198), global assessment of higher order function, estimation of intracranial pressure, such as by fundus examination.

In one embodiment, peripheral nervous system testing may be utilized to assess cognitive function, including any one of the following: smell, visual field and vision, eye movements and pupils (sympathetic and parasympathetic), facial sensory function, strength of the muscles of the face and shoulder girdle, hearing, taste, pharyngeal movement and reflexes, tongue movement (which may be tested individually (e.g., visual acuity may be tested with a Snellen chart, and a reflex hammer may be used to test reflexes including masseter reflex, biceps tendon reflex, triceps tendon reflex, knee tendon reflex, Achilles tendon reflex, and plantar reflex (i.e., Babinski sign)), muscle strength (often on an MRC scale of 1-5), muscle tone, and signs of rigidity.

ii. Multiple Sclerosis In addition to monitoring the improvement of symptoms associated with cognition, techniques known to those skilled in the art can be used to monitor the progression or improvement of neurodegeneration associated with multiple sclerosis (MS). By way of example and not limitation, monitoring can be performed through techniques such as cerebrospinal fluid (CSF) monitoring, magnetic resonance imaging (MRI) to detect the appearance of lesions and demyelinating plaques, evoked potential testing, and ambulatory monitoring.

CSF analysis may be performed, for example via lumbar puncture, to obtain pressure, appearance, and CSF contents. Normal values are typically: pressure 70-180 mm _H2O , appearance clear and colorless, total protein 15-60 mg/100 mL, IgG 3-12% of total protein, glucose 50-80 mg/100 mL, cell count 0-5 white blood cells, no red blood cells, chloride 110-125 mEq/L. Abnormal results may indicate the presence or progression of MS.

MRI is another technique that may be performed to monitor disease progression and improvement. Typical criteria for monitoring MS with MRI include the appearance of patchy areas of abnormal white matter in the cerebral hemispheres and periventricular areas, lesions in the cerebellum and/or brain stem, and cervical or thoracic spinal cord regions.

Evoked potentials may be used to monitor the progression and improvement of a subject's MS. Evoked potentials measure the delay of electrical impulses, such as in visual evoked responses (VER), brainstem auditory evoked responses (BAER), and somatosensory evoked responses (SSER). Abnormal responses help indicate that there is a decrease in conduction velocity in the central sensory pathways.

Ambulatory monitoring can also be used to monitor disease progression and improvement in subjects with MS. MS is often accompanied by motor dysfunction and gait abnormalities, due in part to fatigue. Monitoring can be performed, for example, by using a portable monitoring device worn by the subject (Moon, Y., et al., Monitoring gait in multiple sclerosis with novel wearable motion sensors, PLOS One, 12(2): e0171346 (2017)).

iii. Huntington's Disease Techniques known to those skilled in the art can be used to monitor the progression or amelioration of neurodegeneration associated with Huntington's disease (HD), in addition to monitoring the improvement of symptoms associated with cognition. Monitoring can be performed through techniques such as, by way of example and not limitation, motor function assessment, behavioral assessment, functional assessment, and imaging.

Examples of motor functions that may be monitored as indicators of disease progression or improvement include chorea and dystonia, rigidity, bradykinesia, oculomotor dysfunction, and changes in gait/balance. Techniques for monitoring these indicators are known to those skilled in the art (see Tang C, et al., Monitoring Huntington's disease progression through preclinical and early stages, Neurodegenerator Dis Manag 2(4):421-35 (2012)).

The psychological effects of HD provide an opportunity to monitor disease progression and improvement. For example, a psychiatric evaluation may be performed to determine whether a subject is suffering from psychosis accompanied by depression, irritability, agitation, anxiety, apathy, and paranoia. (Ibid.)

Functional assessment may be used to monitor disease progression or improvement. Total functional scores have been reported (see references above), and in some HD groups, they often decline by one point per year.

MRI or PET may be used to monitor disease progression or improvement.For example, in HD, there is loss of striatal projection neurons, and the number of these neurons may be monitored in subjects.The technique for determining the neuronal changes in HD subjects includes imaging the binding of dopamine _D2 receptors (see above).

iv. ALS
Techniques known to those skilled in the art can be used to monitor the progression or amelioration of neurodegeneration associated with amyotrophic lateral sclerosis (ALS), in addition to monitoring the improvement of symptoms associated with cognition, through techniques such as, by way of example and not limitation, functional assessment, determining muscle strength, measuring respiratory function, measuring lower motor neuron (LMN) loss, and measuring upper motor neuron (UMN) dysfunction.

Functional assessment can be performed using functional scales known to those skilled in the art, such as the ALS Functional Rating Scale (ALSFRS-R), which assesses symptoms related to ocular, limb, and respiratory function. The rate of change is useful for predicting survival and disease progression or improvement. Another scale is the Combined Assessment of Function and Survival (CAFS), which ranks the clinical outcome of subjects by combining survival and changes in the ALSFRS-R (Simon NG, et al., Quantifying Disease Progression in Amyotrophic Lateral Sclerosis, Ann Neurol 76:643-57 (2014)).

Muscle strength can be tested and quantified through the use of multiple manual muscle testing (MMT) scoring, which involves averaging measurements from several muscle groups using the Medical Research Council (MRC) muscle strength rating scale (ibid.). Among other techniques, handheld dynamometers (HHDs) may also be used (ibid.).

A respiratory function assessment can be performed using a portable spirometry unit, which can be used to obtain baseline forced vital capacity (FVC) to predict disease progression or improvement. In addition, maximum expiratory pressure, nasal inspiratory pressure (SNIP) and supine FVC can be obtained and used to monitor disease progression/improvement (ibid.).

Loss of lower motor neurons is another indicator in ALS that can be used to monitor disease progression or improvement. Neurophysiological indicators can be determined by measuring the compound muscle action potential (CMAP) in motor nerve conduction studies, parameters of which include CMAP amplitude and F-wave frequency (see above and de Carvalho M, et al., Nerve conduction studies in amyotrophic lateral sclerosis. Muscle Nerve 23:344-352, (2000)). Lower motor neuron unit number (MUNE) can also be estimated. In MUNE, the number of remaining motor axons transmitting to muscles is estimated through the estimation of the contribution of individual motor units to the maximum CMAP response, and this number is used to determine disease progression or improvement (Simon NG et al., supra). Additional techniques to assess LMN loss include nerve excitability tests, electrical impedance myography, and muscle ultrasound to detect changes in muscle thickness (see references above).

Upper motor neuron dysfunction is another indicator in ALS that can be used to monitor disease progression or improvement. Techniques to determine dysfunction include MRI or PET scans of the brain and spinal cord, transcranial magnetic stimulation, and determining levels of biomarkers in cerebrospinal fluid (CSF).

v. Glaucoma Using techniques known to those skilled in the art, the progression or improvement of neurodegeneration associated with glaucoma can be monitored in addition to monitoring the improvement of symptoms associated with cognition.Monitoring can be performed through techniques such as, but not limited to, techniques for determining intraocular pressure, evaluation of damage to the optic disc or optic disc, visual field testing for peripheral visual field defects, and techniques for imaging the optic disc and retina for topographical analysis.

vi. Progressive supranuclear palsy (PSP)
Techniques known to those skilled in the art can be used to monitor the progression or amelioration of neurodegeneration associated with Progressive Supranuclear Palsy (PSP), in addition to monitoring the improvement of symptoms associated with cognition. This can be done through techniques such as, but not limited to, functional assessment (activities of daily living, i.e., ADLs), motor assessment, psychiatric symptom assessment, volumetric magnetic resonance imaging (MRI), and functional MRI. .

A subject's level of functioning, whether independent, partially dependent on others, or completely dependent, can be useful for judging the progression or improvement of the disease (see Duff, K, et al., Functional impairment in progressive supranuclear palsy, Neurology 80:380-84, (2013)). The Progressive Supranuclear Palsy Rating Scale (PSPRS) is a rating scale that includes 28 indicators in six categories: daily activities (by history), behavior, eye, eye movement, limb movement, and gait/trunk. The result is a score ranging from 0 to 100. Six items are scored from 0 to 2 and 22 items are scored from 0 to 4, resulting in a total possible score of 100. The PSPRS score is a practical tool and a robust predictor of patient survival. The PSPRS score is sensitive to disease progression and is useful for monitoring disease progression or improvement (Golbe LI, et al., A clinical rating scale for progressive supranuclear palsy, Brain 130:1552-65, (2007)).

The ADL portion of the Unified Parkinson's Disease Rating Scale (UPDRS) can also be used to quantify functional activity in PSP subjects (Duff K et al., supra). Similarly, the Schwab England Activities of Daily Living (SE-ADL) can be used to assess independence (supra). In addition, the motor function portion of the UPDRS is useful as a reliable measure to assess disease progression in PSP patients. The motor portion can include, for example, 27 different scales to quantify motor function in PSP patients. Examples of these scales include resting tremor, rigidity, finger tapping, posture and gait. The subject's disease progression or improvement can be assessed by performing a baseline neuropsychological assessment performed by a trained medical professional, using the Neuropsychiatric Inventory (NPI) to determine the frequency and severity of abnormal behaviors (e.g., delusions, hallucinations, agitation, depression, anxiety, euphoria, apathy, disinhibition, irritability and abnormal motor behaviors) (supra).

Functional MRI (fMRI) can also be employed to monitor disease progression and improvement. fMRI is a technique that uses MRI to measure changes in brain activity in specific brain regions, usually based on blood flow to those regions. Blood flow is considered to correlate with activation of brain regions. Physical and mental testing can be performed on patients with neurodegenerative disorders such as PSP before or during their scan in the MRI scanner. By way of example, and not limitation, the test can be a well-established force control paradigm in which the patient is asked to exert force on the hand most affected by the PSP, and the maximum voluntary contraction (MVC) is measured by fMRI immediately after the test (Burciu, RG, et al., Distinct patterns of brain activity in progressive supranuclear palsy and Parkinson's disease, Mov. Disord. 30(9):1248-58 (2015)).

Volumetric MRI is a technique that uses an MRI scanner to determine volumetric differences in regional brain volumes. This may be done, for example, by contrasting different disorders or by determining volumetric differences in brain regions of a patient over time. Volumetric MRI may also be used in neurodegenerative disorders such as PSP to determine disease progression or improvement. This technique is known to those skilled in the art (Messina D, et al., Patterns of brain atrophy in Parkinson's disease, progressive supranuclear palsy and multiple system atrophy, Parkinsonism and Related Disorders, 17(3):172-76 (2011)). Examples of cerebral regions that may be measured include, but are not limited to, the intracranial volume, cerebral cortex, cerebellar cortex, thalamus, caudate nucleus, putamen, globus pallidus, hippocampus, amygdala, lateral ventricle, third ventricle, fourth ventricle, and brain stem.

vii. Neurogenesis A non-invasive technique for assessing neurogenesis has been reported (Tamura Y. et al., J. Neurosci. (2016) 36 (31): 8123-31). Positron Emission Tomography (PET) using the tracer [ ¹⁸ F]FLT in combination with the BBB transporter inhibitor probenecid allows the accumulation of the tracer in neurogenic regions of the brain. This imaging allows neurogenesis to be assessed in patients undergoing treatment for neurodegenerative diseases.

viii. Parkinson's Disease and Motor Function Several rating scales are used to assess the progression of PD. The most widely used scales include the Unified Parkinson's Disease Rating Scale (UPDRS, introduced in 1987) (J. Rehabil. Res. Dev., 2012 49(8):1269-76) and the Horn-Yahr Scale (Neurology, 1967 17(5):427-42). Additional scales include the Movement Disorder Society (MDS) updated UPDRS scale (MDS-UPDRS) and the Schwab England Activities of Daily Living (ADL) scale.

The UPDRS scale assesses 31 items divided into three subscales: (1) mental function, behavior, and mood, (2) activities of daily living, and (3) motor function tests. The Horn-Yahr scale categorizes PD into five stages with substages: 0 (no signs of disease), 1 (unilateral symptoms), 1.5 (unilateral symptoms, but also cervical and spinal), 2 (bilateral symptoms, but no balance impairment), 2.5 (mild symptoms on both sides, able to bounce back during "pull" test), 3 (mild to moderate disease with balance impairment), 4 (severely impaired, but able to walk or stand without assistance), and 5 (requires wheelchair or is bedridden without assistance). The Schwab-England scale categorizes PD into several percentages, from 100% (fully independent) to 10% (fully dependent).

Gross motor function can be assessed using widely used scales, including the Gross Motor Function Scale (GMF), which tests three items: dependency, pain, and instability (Aberg A.C., et al. (2003) Disabil. Rehabil. 2003 May 6;25(9):462-72.). Motor function can also be assessed using home monitoring or wearable sensors. For example, gait (locomotor speed, variability, leg rigidity) can be sensed by accelerometers, posture (trunk tilt) by gyroscopes, leg movements by accelerometers, hand movements by accelerometers and gyroscopes, tremors (amplitude, frequency, duration, laterality) by accelerometers, falls by accelerometers, freezing of gait by accelerometers, dyskinesias by accelerometers, gyroscopes and inertial sensors, bradykinesia (duration and frequency) by accelerometers and gyroscopes, and aphasia (pitch) can be sensed using microphones (Pastorino M, et al., Journal of Physics: Conference Series 450 (2013) 012055).

l. Reagents, Devices, and Kits Reagents, devices, and kits thereof for carrying out one or more of the above methods are also provided. The reagents, devices, and kits thereof of the present invention can vary widely. Applicable reagents and devices include those described above for the method of administering a compound of formula 1 in a subject.

In addition to the above components, the kits of the invention will further include instructions for practicing the methods of the invention. These instructions may be provided in the kits of the invention in a variety of forms, one or more of which may be provided within the kit. One form in which these instructions may be provided is as information printed on a suitable medium or substrate, such as one or more sheets of paper on which the information is printed, within the kit packaging, within an insert, etc. Yet another means is a computer readable medium on which the information is recorded, such as a diskette, CD, portable flash drive, etc. Yet another means that may be provided is a website address that may be used via the internet to access the information at a remote site. Any convenient means may be provided in the kit.

VII. EXAMPLES The following examples are offered by way of illustration and not by way of limitation.

Pharmaceutical Preparations Pharmaceutical compositions for administration to subjects with dementia or neurodegenerative diseases, comprising the above compounds, cocrystals and salts, can be synthesized, prepared and formulated using the examples disclosed in U.S. Patent Application Publication Nos. 2013/0266646, 2016/0081998, U.S. Patent Nos. 8,278,302, 8,653,075, RE45323, 8,742,115, 9,233,950 and 8,680,280, which are incorporated herein by reference in their entirety.Furthermore, the pharmaceutical compositions can be prepared as described in the following examples.

1. Tablet formulation - Wet granulation Copovidone is dissolved in ethanol at ambient temperature to make the granulation liquid. The active CCR3 antagonist ingredient, lactose and a portion of the crospovidone are blended in a suitable mixer to make a premix. The premix is moistened with the granulation liquid and then granulated. The moist granules are optionally sieved through a sieve with 1.6-3.0 mm openings. The granules are dried in a suitable dryer at 45°C to a residual moisture content of 1-3% moisture lost upon drying. The dried granules are sieved through a sieve with 1.0 mm openings. The granules are blended with a portion of the crospovidone and microcrystalline cellulose in a suitable mixer. Magnesium stearate is added to the blend after sieving through a 1.0 mm screen for sizing. The final blend is then made by final blending in a suitable mixer and compressed into tablets. The following tablet compositions can be obtained:

2. Tablet formulation - Melt granulation The active CCR3 antagonist ingredient, lactose, a portion of mcc, polyethylene glycol, lactose and a portion of crospovidone are blended in a suitable mixer to make a premix. The premix is heated in a high shear mixer and then granulated. The hot granules are cooled to room temperature and sieved through a 1.0 mm mesh screen. The granules are blended with a portion of crospovidone and microcrystalline cellulose in a suitable mixer. Magnesium stearate is added to this blend after sieving through a 1.0 mm mesh screen for sizing. The final blend is then made by final blending in a suitable mixer and compressed into tablets. The following tablet compositions can be obtained:

3. Tablet formulation - Hot melt granulation The active CCR3 antagonist ingredient, mannitol, polyethylene glycol and a portion of the crospovidone are blended in a suitable mixer to make a premix. The premix is heated in a high shear mixer and then granulated. The hot granules are cooled to room temperature and sieved through a 1.0 mm mesh screen. The granules are blended with a portion of the crospovidone and mannitol in a suitable mixer. Magnesium stearate is added to this blend after sieving through a 1.0 mm mesh screen for sizing. The final blend is then made by final blending in a suitable mixer and compressed into tablets. The following tablet compositions can be obtained:

4. Tablet formulation - Hot melt extrusion process The active CCR3 antagonist ingredient and stearic-palmitic acid are blended in a suitable mixer to make a premix. The premix is extruded in a twin screw extruder and then granulated. The granules are sieved through a 1.0 mm mesh screen. The granules are blended with mannitol and crospovidone in a suitable mixer. Magnesium stearate is added to this blend after sieving through a 1.0 mm mesh screen for sizing. The final blend is then made by final blending in a suitable mixer and compressed into tablets. The following tablet compositions can be obtained:

5. Tablet formulation - Hot melt extrusion process The active CCR3 antagonist ingredient and stearic-palmitic acid are blended in a suitable mixer to produce a premix. The premix is extruded in a twin screw extruder and then granulated. The granules are sieved through a 1.0 mm mesh screen. The granules are directly filled into hard capsules. The following capsule compositions can be obtained:

6. Tablet formulation - roller compaction The active CCR3 antagonist ingredient, mannitol and a portion of the crospovidone, and magnesium stearate are blended in a suitable mixer to make a premix. The premix is compressed in a roller compactor and then granulated. Optionally, the granules are sieved through a 0.8 mm mesh screen. The granules are blended with mannitol and a portion of the crospovidone in a suitable mixer. Magnesium stearate is added to this blend after sieving through a 1.0 mm mesh screen for sizing. The final blend is then made by final blending in a suitable mixer and compressed into tablets. The following tablet compositions can be obtained:

7. Tablet formulation - roller compaction The active CCR3 antagonist ingredient and magnesium stearate are blended in a suitable mixer to make a premix. The premix is compressed in a roller compactor and then granulated. Optionally, the granules are sieved through a 0.8 mm mesh screen. The granules are blended with mannitol and croscarmellose sodium in a suitable mixer. Magnesium stearate is added to this blend after sieving through a 1.0 mm mesh screen for sizing. The final blend is then made by final blending in a suitable mixer and compressed into tablets. The following tablet compositions can be obtained:

8. Tablet formulation - roller compaction The active CCR3 antagonist ingredient and magnesium stearate are blended in a suitable mixer to make a premix. The premix is compressed in a roller compactor and then granulated. Optionally, the granules are sieved through a 0.8 mm mesh screen. The granules are blended with microcrystalline cellulose and crospovidone in a suitable mixer. Magnesium stearate is added to this blend after sieving through a 1.0 mm mesh screen for sizing. The final blend is then made by final blending in a suitable mixer and compressed into tablets. The following tablet compositions can be obtained:

9. Coated Tablet Formulation Film-coated tablets can be prepared using the tablet cores according to the above formulation. Hydroxypropylmethylcellulose, polyethylene glycol, talc, titanium dioxide and iron oxide are suspended in purified water in a suitable mixer at ambient temperature to prepare a coating suspension. The tablet cores are coated with the coating suspension to obtain a weight gain of about 3% to prepare film-coated tablets. The following film coating compositions can be obtained:

b. Drug Formulation and Administration There is provided an investigational product of the present invention (Compound 1) which corresponds to the chemical structure below.

Those skilled in the art will recognize that the compounds, cocrystals, salts and formulations previously described in the sections above can also be used in these examples.

Compound 1 was provided as 100 mg, 200 mg, and 400 mg film-coated tablets that were biconvex, round or oval, and dark red in color. The tablets were made by a dry granulation process and contained the following inactive ingredients: microcrystalline cellulose, hydrogen phosphate, croscarmellose sodium, magnesium stearate, polyvinyl alcohol, titanium dioxide, polyethylene glycol, talc, iron oxide red, and iron oxide yellow. Corresponding placebo tablets were made by a direct compression process and contained the same inactive ingredients.

c. Preclinical Examples 1. Materials and Methods (a) Subcutaneously Implanted Osmotic Pumps Alzet minipumps were filled and prepared the day before implantation to allow priming at 37°C and to allow blinding of treatment, and P-numbered by mouse ID. Pumps were implanted on the back, slightly posterior to the scapula and slightly lateral to the trunk. Mice were anesthetized with 3-5% isoflurane using a vaporizer and regulator in an induction chamber, then moved to the procedure area, fitted with a nose cone, and anesthesia maintained with 1-3% isoflurane. Ophthalmic ointment was applied to the eyes to prevent dryness. Mice were injected subcutaneously with 5 mg/kg meloxicam. The hair in the incision area was clipped using small sharp scissors, and the area was disinfected by alternating applications of 70% isopropanol and betadine. A 0.5-1 cm incision was made and a hemostat was inserted to spread the subcutaneous tissue to create space for the pump. A pump was inserted into the space and the wound was closed with wound clips. All surgical tools were autoclaved before first use on the day of surgery. Instruments were then sterilized in a glass bead sterilizer between mice. Mice were placed in clean recovery cages partially placed on a heating pad until they were fully recovered and showed ambulation. Mice were checked for induction of anesthesia by toe pinch method and respiratory monitoring. Mice were monitored every 15 minutes after surgery until they recovered. The next day, mice were administered a second dose of meloxicam. If signs of infection were observed, mice were administered Baytril 5 mg/kg subcutaneously until the infection cleared.

(b) Open Field Test The open field is used to assess overall locomotor activity and exploratory behavior in a novel environment. Before testing, mice are placed in the experimental room for at least 30 min to acclimate to the laboratory conditions (dim lighting). The test arena consists of a 50 cm x 50 cm square arena. Mice are placed in the center of the arena and followed for 15 min. The time spent in the periphery and center are analyzed along with rearing behavior. All surfaces are disinfected with 70% ethanol between trials.

(c) Y-maze The large Y-maze test evaluates short-term memory of grasping a particular context. Before testing, mice are placed in the experiment room for at least 30 minutes to acclimate to the laboratory conditions (dim lighting). In the first training trial, the mouse is placed at the end of one arm of the large Y-maze, designated as the "start arm" (arm length: 15 inches). The third arm of the maze is closed and the mouse is allowed to freely explore two of the three arms (the "start arm" and the "familiar arm") for 5 minutes. Each arm is equipped with spatial cues. After 1 hour, the mouse is returned to the "start arm" of the maze and allowed to explore all three arms, including the open third arm (the "novel arm"). Movement into and out of each arm is tracked using automated tracking software (CleverSys). Testing is performed under dim lighting and the apparatus is disinfected with 70% ethanol between trials.

(d) Barnes maze A modified Barnes maze was used to assess spatial tasks/episodic learning and memory. The Barnes maze apparatus consists of a circular platform with a diameter of 122 cm with 40 escape holes, each of which is 5 cm in diameter and arranged along three rings at different distances from the center of the platform. An escape box is attached to one of the holes, and all holes are left open. A bright light and an electric fan are directed onto the maze to provide aversive stimuli and encourage escape. Visual cues are placed on all four ends of the maze. The mouse is given a series of four or five trials, with an interval of about 10 minutes between trials, and the maximum duration of each trial is 90 or 120 seconds. During each trial, the mouse is placed in the center of the maze. After 10 seconds, the mouse is allowed to explore, and if the mouse finds and enters the escape box, the trial is terminated before the end time of the trial. Mice that cannot find the escape box are guided to the escape box and allowed to enter it for 30 seconds before being returned to their home cage. Training is carried out for 4 days. Data recorded and analyzed include speed, escape latency, and distance traveled.

Divide mice into groups of 4-5 mice per group, with treatment groups balanced. For example, perform 4 trials with mice in group 1, then 4 trials with mice in group 2, and so on until all groups have been tested. Disinfect the arena with 70% ethanol between trials.

(e) Rotarod Mice were trained on the rotarod for three trials, each lasting up to 100 seconds (rotarod is a test of motor coordination). In the last trial, success or failure was recorded (a fall latency of more than 90 seconds was considered successful). A binomial test was performed to compare the success rates of control and compound-treated mice.

(f) T-maze The water maze was filled with water at least 24 hours prior to testing and allowed to reach room temperature. The water was stained with white latex paint to allow the mice to see during tracking and to allow the use of a hidden platform. Two distinct visual cues were placed at either end of the T-arm of the inserted T-maze. On day 1, mice were given 4 trials, each trial using a visible platform, with a 30-minute interval between trials. Mice were given 60 seconds to reach the platform. If they did not reach the platform within that time, they were guided to the platform and allowed to remain there for 5 seconds before being removed from the tank. After the third mouse, goal arms were switched each time, ensuring that both treatment groups had the same number of goal arms on the right and left turns. After each trial, mice were placed in an empty cage with blue padding and allowed to dry under a red light lamp before being returned to their home cage. On the second day, which was the test day, the mice were given four trials of the same test except that they used the hidden platform, with a 30-minute interval between trials. Mice were scored for correct or incorrect choices and latency to reach the platform, and a binomial test was performed to compare success rates between control and compound-treated mice. All trials were recorded using TopScan.

(g) Protein quantification Mouse plasma CCL11 protein levels were measured by sandwich ELISA. For this assay, plasma was diluted 1:10 (Mouse CCL11/Eotaxin Duo Set ELISA Kit, R&D Systems, Minneapolis, Minn.).

Human CCL11 from human plasma was measured by SomaLogic's aptamer-based assay (SOMAscan, SomaLogic, Inc., Boulder, CO) (Gold L, et al. (2010). Aptamer-Based Multiplexed Proteomic Technology for Biomarker Discovery. PLOS ONE 5(12):e15004).

A panel of circulating cytokines from mouse plasma was analyzed by Luminex. Luminex Assay Service was performed by Eve Technologies (Calgary, Alberta, Canada).

(h) Quantification of mRNA. Snap-frozen hemibrains were dissected into cortex, hippocampus, striatum and thalamus. The cortex was divided into three homogenous parts, and one part was used for RNA isolation. RNA was isolated using Prolink RNA Mini Kit (Thermo Fisher Scientific #12183025). cDNA was generated using Taqman RT Kit (Thermo Fisher Sci #N8080234). qPCR was performed on Quant Studio 6 using custom-made Taqman Multiplex Primers for IL-1β and GAPDH. All samples were run on a single plate and analyzed for ddCT first compared to their endogenous control and then compared to the control group.

(i) Quantitation of Compound 1 by HPLC/MS Compound levels were measured by MS/MS by Quintara (Hayward, Calif.).

(j) Flow Analysis Eosinophil Counts. 200 uL of whole blood was collected during perfusion and sent to Charles River Clinical Pathology Services (Shrewsbury, Mass.) for hematological analysis of eosinophil counts by FACS (fluorescence activated cell sorter).

Morphological changes in eosinophils (ESC)
ESCs are determined by the morphological changes in human eosinophils activated with human eotaxin-1 (PreProTech, Rocky Hill, NJ) compared to native eosinophils. This change is detected as a change in forward scatter measured by FACS (fluorescence activated cell sorter). The mean forward scatter of the autofluorescent (eosinophil) population in each sample was determined along with the mean of each triplicate sample group. Methods for determining ESCs have been previously described and are known in the art.

CCR3 Receptor Internalization The CCR3 receptor internalization assay is FACS-based and uses human eotaxin-1 (PreProTech, Rocky Hill, NJ) and APC-labeled anti-human CCR3 antibody (R&D Systems, Minneapolis, Minnesota) or isotype control IgG ₂ A antibody to monitor human eotaxin-1-induced CCR3 receptor internalization in human eosinophils compared to naive eosinophils. The median APC fluorescence intensity units for each sample were determined along with the mean for each triplicate sample group. Methods for monitoring CCR3 receptor internalization have been previously described and are known in the art.

(k) Histology Mice were dissected the day after the end of behavioral testing. Anesthesia was induced with 2,2,2-tribromoethanol, after which mice were perfused transcardially with 0.9% saline. Brains were dissected and cut sagittally into two halves. One half was snap frozen on dry ice for later use, and the other half was fixed in 4% paraformaldehyde in PBS for immunohistochemical analysis. Two days after fixation, the hemibrain was transferred to 30% sucrose in PBS solution, which changed after 2 days. The hemibrain was sectioned at 30 μm on a microtome at −22° C. Brain sections were stored at −20° C. in cryoprotectant medium until required for staining. Blocking was performed on free-floating sections in the appropriate serum of 10% serum in PBST 0.5%. Primary antibodies were incubated overnight at 4C. For light microscopy, antibodies DCX, 1:200, Santa Cruz BioTech, CD68, 1:1000. AbD Serotec were used at the indicated concentrations. The next day, secondary biotinylated antibodies were applied at a concentration of 1:300. Visualization by staining was performed by reaction with ABC Kit (Vector) and diaminobenzidine (Sigma). Dehydration of mounted slides was performed by immersion in ethanol and xylene. Images were obtained at 5x magnification on a Leica light microscope.

For fluorescence microscopy, the antibodies GFAP, 1:500, DAKO; Iba1, 1:1000, Wako, and BrdU, 1:500, AbCam were used at the indicated concentrations. For BrdU, an antigen retrieval protocol was required before blocking (2N HCl, 37C, 30 min). The following day, the appropriate fluorescent secondary antibody was applied at a concentration of 1:300 for 1 h at room temperature. Slides were coverslipped using Prolong Gold Mounting Media. Images were obtained under a light microscope at 5x magnification.

2. Experimental Groups First experimental group (see Figures 1-2): 2-month-old or 18-month-old C57B1/6 mice were administered either IgG antibody control by intraperitoneal injection or Compound 1 by subcutaneous administration via an Alzet osmotic pump for either 2 or 4 weeks. During the last week of treatment, mice were subjected to behavioral testing before perfusion on the last day of treatment. Just before the start of treatment, all mice were injected intraperitoneally with BrdU at 150 mg/kg for 5 consecutive days.

Drug formulations: Control rat IgG2A clone 54447 (MAB006, R&D Systems) was administered at 50ug/kg in sterile saline. Compound 1 was formulated in 40% HP-β-cyclodextrin and adjusted to pH 6.5 with NaOH (1M). Solutions were prepared fresh weekly and stored at 4°C.

Treatment Groups:
Treatment Group 1 (Young Control): Young C57BL/6 mice (n=18), 1-2 months of age, received 5 intraperitoneal (IP) injections of control IgG (injections given every 3 days for 14 days).
Treatment group 2 (aged control): aged C57BL/6 mice (n=18) aged 18 months were given five intraperitoneal (IP) injections of control IgG (injected once every 3 days for 14 days).
Treatment group 3 (Compound 1 (dose 1) in aged group): 18-month-old aged C57BL/6 mice (n=16) were infused with approximately 50 mg/ml of Compound 1 via an Alzet minipump model 2001 (1 uL/hr) for 2 weeks with one supplement.
Treatment group 4 (compound 1 (dose 2) in the aged group): 18-month-old aged C57BL/6 mice (n=16) were infused with approximately 50 mg/ml of compound 1 via an Alzet minipump model 2002 (0.5 uL/hr) for 2 weeks.
Treatment group 5 (Compound 1 (dose 2) in aged group): 18 month old aged C57BL/6 mice (n=16) were infused with approximately 50 mg/ml of Compound 1 via an Alzet minipump model 2002 (0.5 uL/hr) for 4 weeks with one supplement.

Compound 1 administered subcutaneously for 4 weeks to 18-month-old mice increased the number of doublecortin-positive cells (an indicator of neurogenesis) in the hippocampus (see Figure 1A). Compound 1 administered at a high dose for 2 weeks tended to increase BrdU-positive cells (a cell proliferation marker) in the hippocampus (see Figure 1B). Compound 1 infusions at low or high doses for 2 or 4 weeks improved the cued Y-maze (a test of memory) (see Figure 2, where the percentage of time normalized to total interaction time and the number of visits are recorded. Similar effects were seen when total time spent was assessed). Comparisons were also made with control groups (treatment groups 1 and 2) treated with IgG antibodies instead of the compound, and treatment and behavior were performed in parallel.

Thus, administration of Compound 1 increased the number of Dcx-positive cells and BrdU-positive cells, indicating that Compound 1 increased neurogenesis and cell survival, respectively. Compound 1 was also able to improve memory (cognition) as evidenced by behavior in the Y-maze test.

Second experimental group (see Figures 3-6): 3-month-old or 16.5-month-old C57B1/6 mice were administered either vehicle control or Compound 1 subcutaneously via Alzet osmotic pumps for 4 weeks. During the last week of treatment, mice were subjected to behavioral testing before perfusion on the last day of treatment. Just before the start of treatment, all mice were injected intraperitoneally with BrdU at 150 mg/kg for 5 consecutive days.

Drug preparation: Compound 1 was formulated in 40% HP-β-cyclodextrin and adjusted to pH 6.5 with NaOH (1M). Vehicle solutions were prepared and pH adjusted similarly. Solutions were prepared fresh weekly and stored at 4°C.

Treatment Groups:
Treatment Group 1 (Young Control): Young C57BL/6 mice (n=19) aged 3 months were infused with vehicle via Alzet minipump model 2002 (0.5 uL/hr) for 4 weeks with one refill.
Treatment group 2 (aged controls): aged C57BL/6 mice (n=19) aged 16 months were infused with vehicle via Alzet minipump model 2002 (0.5 uL/hr) for 4 weeks with one supplement.
Treatment group 3 (Compound 1 (dose 2) in aged group): 16-month-old aged C57BL/6 mice (n=19) were infused with approximately 50 mg/ml of Compound 1 via an Alzet minipump model 2002 (0.5 uL/hr) for 4 weeks with one supplement.

Compound 1 administered subcutaneously for 4 weeks to 16.5-month-old mice improved cognitive behavior in a cued Y-maze test of memory (see Figure 3). Compound 1 administered for 4 weeks also tended to improve behavior in the Barnes maze (a test of hippocampal-dependent spatial memory) (see Figure 4). A trend toward improved memory was observed when analyzing the mean latency for the 4th trial and also in the difference in latency between the 13th and 16th trials. Compound 1 administered for 4 weeks also significantly increased the number of BrdU-positive cells (an indicator of neurogenesis) in the hippocampus (see Figure 5). Thus, Compound 1 was able to achieve both improved cell survival and improved memory (cognition) as evidenced by the results obtained using the Y-maze and Barnes maze tests.

Cerebrospinal Fluid Levels of Compounds in Mice Cerebrospinal fluid (CSF) was collected from both the "young" group at 2 months of age and the "old" group at 16.5 months of age, and Compound 1 levels were determined by mass spectrometry. Figure 6 shows the levels of the compound of the present invention detected in the CSF of both young and old mice (all below 10 nM). These CSF levels are far from the Ki of the compound in mice (124 nM, as determined by receptor binding in cell lines), and therefore do not cross the blood-brain barrier (BBB) in significant concentrations. For comparison, levels of the compounds of the invention measured in the plasma of young (2 month old) and old (18 month old) mice perfused with a 50 mg/mL solution at 0.5 μL/hr for 2 and 4 weeks, respectively, were significantly higher (352±31 nM at 2 weeks and 355±43 nM at 4 weeks (values are mean±standard error)), further indicating that the compounds of the invention are unable to cross the BBB in significant concentrations.

This data indicates that Compound 1 does not act directly in the CNS, but rather acts peripherally. In addition, Compound 1 crosses the BBB in insufficient concentrations to have an effect. Furthermore, there is no difference in BBB crossing between young and old mice, indicating that the effects of Compound 1 on cognition and neurodegeneration are not due to differences in the BBB between the two groups.

Compound tissue distribution levels Compound 1 was distributed in mouse tissues after oral administration of [ ¹⁴ C]-radiolabeled Compound 1 ("labeled compound") to male pigmented C57BL/6JOlaHsd mice (Harlan Labs, BV). The labeled compound was administered at 10 mg/kg body weight (equivalent to 17 μmol/kg). One mouse was sacrificed at 1, 24 and 168 hours after administration. Blood, plasma and eye radioactivity concentrations were measured using liquid scintillation counting (LSC). Tissue and organ concentrations were determined by whole body autoradiography (QWBA). Preparation of whole-body sections of mice was performed using a cryostat microtome Reichert-Jung CRYO MACROCUT or CRYO MACROCUT LEICA CM 3600 ³ according to known techniques (see S. Ullberg, et al., Autoradiography in Pharmacology in: The Int. Encyclopedia of Pharmacology, J. Cohen (Ed.), 1(78):221-39 (1971)).

To allow quantitative evaluation of radioactivity, sections of the adrenal gland, blood, bone marrow, brain, eye (lens), epididymis, fat (white and brown), Harderian gland, heart, kidney, liver, lung, muscle, pituitary gland, pancreas, prostate, spinal cord, spleen, salivary gland, skin, testis, thyroid gland, thymus, uveal tract, and whole body sections selected in 5-7 horizontal planes were taken from the embedded mice. Two sections were taken from each selected horizontal plane per mouse and the sections were freeze-dried on a microtome at -20 to -25°C for a minimum of 48 hours.

Figure 7 shows a table of recorded area under the curve (AUC) values at all three time points, quantifying the exposure of each tissue to the labeled POI following compound administration. Figure 7 shows that Compound 1 does not penetrate the blood-brain barrier (BBB) at significant levels.

These results also indicate that Compound 1 acts peripherally, as it is unable to cross the BBB at significant levels. Thus, Compound 1's effects are not directly directed at the central nervous system, overcoming a difficult challenge that has caused the failure of many potential CNS disease-targeting drugs.

Pharmacokinetic Profile of Oral Administration Compound 1 concentrations were measured in plasma from 2-month-old male C57B1/6 mice at 20 minutes, 2 hours, 8 hours, and 12 hours after oral gavage at two doses of 30 mg/kg and 150 mg/kg. The 30 mg/kg dose was found to be sufficient to achieve compound 1 concentrations of >100 nM for 8 hours (Figure 8).

Third experimental group (see Figures 9-11): Two-month-old C57Bl/6 mice were administered either vehicle control or Compound 1 by oral gavage twice daily for 18 days. During the final week of treatment, mice were subjected to behavioral testing before perfusion on the final day of treatment. Just prior to the start of treatment, all mice were injected intraperitoneally with BrdU at 150 mg/kg for five consecutive days.

Drug preparation: Compound 1 was formulated in 40% HP-β-cyclodextrin and adjusted to pH 6.5 with NaOH (1M). Vehicle solutions were prepared and pH adjusted similarly. Solutions were prepared fresh weekly and stored at 4°C.

Treatment Groups:
Treatment Group 1a (vehicle treatment): Young C57BL/6 mice (n=15) aged 7 weeks were orally (PO) dosed with vehicle solution twice daily (BID) for 18 days, with one injection on the last day, for a total of 35 doses.
Treatment group 1b (treatment with Compound 1): Young C57BL/6 mice (n=15), 7 weeks of age, were orally (PO) dosed with Compound 1 at 30 mg/kg twice daily (BID) for 18 days, with one injection on the last day, for a total of 35 doses.
Treatment group 2a (combined treatment with rmCCL11 with vehicle): Young C57BL/6 mice (n=15) aged 7 weeks were orally (PO) dosed with vehicle solution twice daily (BID) for 18 days, with one injection on the last day, for a total of 35 injections. Mice were simultaneously given rmCCL11 via peripheral injection (IP) every 3 days starting on the first day of treatment, for a total of 5 injections.
Treatment group 2b (combined treatment with Compound 1 and rmCCL11): Young C57BL/6 mice (n=15) aged 7 weeks were orally (PO) dosed with Compound 1 at 30 mg/kg twice daily (BID) for 18 days, with one injection on the last day, for a total of 35 doses. Mice were simultaneously given rmCCL11 via peripheral injection (IP) every 3 days starting on the first day of treatment, for a total of 5 injections.

Treatment with recombinant mouse CCL11 ("rmE") significantly worsened anxiety in the open field, whereas oral administration of Compound 1 twice daily for 2 weeks improved anxiety (see FIG. 9). rmCCL11 impaired memory in the Y-maze, but Compound 1-treated mice were no longer significantly different from control mice (see FIG. 10). rmCCL11 also impaired memory in the Barnes maze, and treatment with Compound 1 significantly improved memory behavior (see FIG. 11). Thus, rmE impaired both anxiety in the open field test and memory as evidenced by behavior measured in the Y-maze and Barnes maze tests. However, treatment with Compound 1 was able to attenuate these effects.

Fourth experimental group (see Figure 12): 23-month-old C57B1/6 mice were administered either vehicle control or Compound 1 by oral gavage twice daily for 19 days. After 11 days of treatment, the mice were subjected to the Y-maze test.

Drug preparation: Compound 1 was prepared in 40% HP-β-cyclodextrin and adjusted to pH 6.5 with NaOH (1M). Vehicle solutions were prepared and pH adjusted similarly. Kolliphor was added weekly to each solution at 10% and stored at 4°C.

Treatment Groups:
Treatment group 1 (vehicle treatment): 23-month-old aged C57B1/6 mice (n=8) were orally gavaged (PO) with vehicle solution twice daily (BID) for 19 days.
Treatment group 2 (treatment with Compound 1): 23-month-old aged C57B1/6 mice (n=11) were administered Compound 1 by oral gavage (PO) at 30 mg/kg twice daily (BID) for 19 days.

Treatment with compound 1 significantly improved memory in the Y-maze. Mice treated with compound 1 showed normal memory for the novel arm as measured by the number of visits (Figure 12A). Treatment with compound 1 also significantly increased the distance traveled by mice during the test (Figure 12B).

Fifth experimental group (see Figures 13-17): 23-month-old C57B1/6 mice were subcutaneously administered either vehicle control or Compound 1 twice daily for 21 days. After 3 weeks, the mice were subjected to behavioral testing and sacrificed the day after the last behavioral test.

Drug preparation: Compound 1 was prepared in 40% HP-β-cyclodextrin and adjusted to pH 6.5 with NaOH (1M). Vehicle solutions were prepared and pH adjusted similarly. Kolliphor was added weekly to each solution at 10% and stored at 4°C.

Treatment Groups:
Treatment group 1b (vehicle treatment): 23 month old aged C57B1/6 mice (n=9) were administered vehicle solution subcutaneously (SQ) twice daily (BID) for 21 days.
Treatment group 4 (treatment with Compound 1): 23-month-old aged C57B1/6 mice (n=17) were administered Compound 1 subcutaneously (SQ) at 30 mg/kg twice daily (BID) for 21 days.

Treatment with Compound 1 significantly improved memory in the Y-maze and Barnes maze. Mice treated with Compound 1 showed normal memory for the novel arms, both in terms of the number of visits (Fig. 13A-B) and time spent in the novel arms (Fig. 13C-D). Treatment with Compound 1 also significantly increased the speed of the mice during the test (Fig. 13E). Mice treated with Compound 1 performed significantly better in terms of spatial memory in the Barnes maze (Fig. 14A) and increased their speed during the test (Fig. 14B). Locomotor activity in the open field also improved, with a strong tendency toward improvement in both distance traveled (Fig. 15A) and speed (Fig. 15B).

Levels of inflammatory cytokines were measured in mouse plasma by Luminex assay (Figure 16). There was a strong trend towards a decrease in several inflammatory markers, including TNFa, IL6, IL1β, IL5 and IL17. Activated microglia were also quantified in the mouse hippocampus by IHC staining (Figure 17). Treatment with Compound 1 strongly trended towards a decrease in microgliosis.

Sixth experimental group (see Figure 18): 23-month-old C57B1/6 mice were subcutaneously injected with either vehicle control or Compound 1 twice daily for 30 days and then sacrificed on the day of the last injection.

Drug preparation: Compound 1 was prepared in 40% HP-β-cyclodextrin and adjusted to pH 6.5 with NaOH (1M). Vehicle solutions were prepared and pH adjusted similarly. Kolliphor was added weekly to each solution at 10% and stored at 4°C.

Treatment Groups:
Treatment group 1a (vehicle treatment): 23 month old aged C57B1/6 mice (n=9) were administered vehicle solution subcutaneously (SQ) twice daily (BID) for 30 days.
Treatment group 3 (treatment with Compound 1): 23-month-old aged C57B1/6 mice (n=18) were administered Compound 1 subcutaneously (SQ) at 30 mg/kg twice daily (BID) for 30 days.

When treated with compound 1, a subset of mice (n=2, 5) showed a strong tendency toward a decrease in blood eosinophil counts as determined by FACS analysis (Figure 18).

Seventh experimental group (see Figure 19): Three-month-old hairless mice were treated with oxazolone (Ox) every other day and orally administered either saline control or 30 mg/kg of Compound 1 twice daily for two weeks.

Drug preparation: Compound 1 was prepared in 40% HP-β-cyclodextrin and adjusted to pH 6.5 with NaOH (1M). Vehicle solutions were prepared and pH adjusted similarly. Kolliphor was added weekly to each solution at 10% and stored at 4°C.

Treatment Groups:
Treatment group 1 (vehicle treatment): 3-month-old hairless mice (n=3) were administered vehicle solution by oral gavage (PO) twice daily (BID) for 2 weeks.
- Treatment group 2 (vehicle treatment and oxazolone treatment): 3-month-old hairless mice (n=6) were administered vehicle solution by oral gavage (PO) twice daily (BID) for 2 weeks.
Treatment group 3 (compound 1 treatment and oxazolone treatment): 3-month-old hairless mice (n=6) were administered compound 1 by oral gavage (PO) twice daily (BID) at 30 mg/kg for 2 weeks.

Compound 1 reduced the oxazolone-induced increase in blood eosinophils by complete blood count (CBC) analysis. N was 3, 6, 8. ^* P<0.05, ^** P<0.01. In the oxazolone-induced model of eosinophilia, compound 1 significantly reduced blood eosinophil counts (Figure 19). This demonstrates that inhibition of CCR3 may be sufficient to normalize eosinophil levels and function, especially in diseased states, and demonstrates a second mechanism (in addition to reducing brain inflammation) by which compound 1 may be effective in treating neuronal loss and associated motor and cognitive deficits.

Eighth experimental group (see Figures 20-22): Compound 1 or vehicle was continuously infused into 24-month-old C57 mice for 4 weeks via Alzet osmotic pumps implanted to continuously deliver Compound 1 or vehicle.

Drug preparation: Compound 1 was prepared in 40% HP-β-cyclodextrin and adjusted to pH 6.5 with NaOH (1M). Vehicle solutions were prepared and pH adjusted similarly. Kolliphor was added weekly to each solution at 10% and stored at 4°C.

Treatment Groups:
Treatment Group 1 (Aged Control): Aged C57BL/6 mice (n=15) aged 23 months were infused with vehicle via an Alzet minipump model 2002 (0.5 uL/hr) with one supplement for 4 weeks.
Treatment group 2 (Compound 1 (dose 2) in aged group): 23 month old aged C57BL/6 mice (n=15) were infused with approximately 50 mg/mL of Compound 1 via an Alzet minipump model 2002 (0.5 uL/hr) for 4 weeks with one supplement.

Mice were tested for motor coordination on a rotarod. Success or failure was recorded on the last trial, with a fall latency of more than 90 seconds considered successful. Compound 1-treated mice were more successful than vehicle-treated mice by binomial test. N = 15, ^* P<0.05. After three consecutive trials, a higher percentage of Compound 1-treated mice (47%) were able to stay on the rod for more than 90 seconds (threshold), while only 20% of control-treated mice were able to stay on the rod (Figure 20). These results suggest that Compound 1 has a consistent effect on motor function in the eotaxin model.

Mice were tested for cognitive function in a T-maze (Figure 21). The number of successful or unsuccessful attempts to enter the correct arm was recorded. Compound 1-treated mice were more successful than vehicle-treated mice by binomial test. ^* P<0.05.

Fecal mass was measured overnight by weighing the dry weight of fecal pellets (Figure 22). Gastrointestinal function is a known symptom of Parkinson's disease. Water and food intake was measured over the same period. Compound 1-treated mice had significantly less fecal mass compared to control mice. Also, overnight water intake was significantly higher compared to control mice. ^* P<0.05. These results indicate that treatment with Compound 1 can modify the deficit in gastrointestinal function.

Ninth experimental group (see Figure 23): Three-month-old C57 mice were intraperitoneally administered lipopolysaccharide (LPS) at 10 mg/kg once to induce inflammation, and then treated with Compound 1 for 18 days.

Drug preparation: Compound 1 was prepared in 40% HP-β-cyclodextrin and adjusted to pH 6.5 with NaOH (1M). Vehicle solutions were prepared and pH adjusted similarly. Kolliphor was added weekly to each solution at 10% and stored at 4°C.

Treatment Groups:
Treatment group 1 (vehicle treatment): Young C57BL/6 mice (n=15) aged 3 months were injected once with LPS and gavaged with vehicle twice daily for 18 days.
Treatment group 2 (treatment with Compound 1): Young C57BL/6 mice (n=15) aged 3 months were injected with LPS once and administered Compound 1 by oral gavage at 30 mg/kg twice daily for 18 days.

Brain sections were immunostained to detect CD68+ activated microglia. Compound 1-treated mice showed a significant decrease in CD68+ immunoreactivity (reduction in activated microglia) in contrast to vehicle (saline)-treated mice (Figure 23). N = 9, 10, 8. ^* P<0.05 (by one-way ANOVA).

These data demonstrate the potent anti-neuroinflammatory activity of compound 1 and its potential therapeutic potential to reduce neuroinflammation-induced neuronal toxicity in diseases manifested by neurodegeneration, cognitive or motor dysfunction.

Experimental group 10 (see Figures 24-29): Three-month-old C57 mice were intraperitoneally administered lipopolysaccharide (LPS) at 0.5 mg/kg daily to induce inflammation and were chronically treated with Compound 1 for up to four weeks.

Drug preparation: Compound 1 was prepared in 40% HP-β-cyclodextrin and adjusted to pH 6.5 with NaOH (1M). Vehicle solutions were prepared and pH adjusted similarly. Kolliphor was added weekly to each solution at 10% and stored at 4°C.

Treatment Groups:
Treatment group 1 (vehicle treatment): 3-month-old C57 mice (n=10) were injected intraperitoneally with LPS daily for 7 weeks and orally gavaged (PO) with vehicle solution twice daily (BID) for up to 4 weeks.
Treatment group 2 (vehicle-treated and oxazolone-treated): 3-month-old C57 mice (n=10) were injected intraperitoneally with LPS daily for 7 weeks and orally gavaged (PO) twice daily (BID) with vehicle solution for up to 4 weeks.
Treatment group 3 (compound 1 treatment and oxazolone treatment): 3-month-old C57 mice (n=9) were injected intraperitoneally with LPS daily for 7 weeks and administered compound 1 by oral gavage (PO) at 30 mg/kg twice daily (BID) for up to 4 weeks.

Figure 24A illustrates the dosing scheme for the tenth experimental group. The frame highlighted with a box represents the time point when the assay in Figure 24B was performed. One week after treatment with Compound 1, anxiety was tested in the open field test (Figure 24B). Treatment with LPS significantly increased anxiety in the open field, and Compound 1 significantly reduced the increase in anxiety ( ^* P<0.05).

Figure 25A illustrates the dosing scheme for the tenth experimental group. The frame highlighted with a box represents the time point at which the assay in Figure 25B was performed. Three weeks after treatment with Compound 1, cognition was tested in a Y-maze (Figure 25B). Mice treated with LPS showed no significant preference for the novel arm. However, Compound 1-treated mice showed a significant preference for the novel arm, similar to vehicle-treated mice ( ^* P<0.05, ^** P<0.01).

Figure 26A illustrates the dosing scheme for experimental group 10. The areas of the scheme highlighted with boxes represent the time points at which the assays in Figure 26B were performed. After 4 weeks of treatment with Compound 1, mRNA levels of the proinflammatory cytokine IL-1β were measured by quantitative PCR (Figure 26B). There was a trend towards increased IL-1β levels in LPS-treated mice, and a significant decrease in IL-1β expression was observed in Compound 1-treated mice ( ^* P<0.05).

Figure 27A illustrates the dosing scheme for the tenth experimental group. The frame highlighted with a box represents the time point at which the assay in Figure 27B was performed. Brain sections were immunostained for detection of CD68+ activated microglia. Compound 1-treated mice showed a significant reduction in CD68+ immunoreactivity (reduction in activated microglia) in contrast to mice treated with LPS alone (Figure 27B).

Figure 28A illustrates the dosing scheme for the tenth experimental group. The frame highlighted with a box represents the time point at which the assay in Figure 28B was performed. Brain sections were immunostained for detection of Iba1 positive microglia. Compound 1-treated mice showed a significant reduction in Iba1+ immunoreactivity (reduced total microglia numbers) in contrast to mice treated with LPS alone (Figure 28B).

Figure 29A illustrates the dosing scheme for experimental group 10. The frame highlighted with a box represents the time point at which the assay in Figure 29B was performed. Brain sections were immunostained for detection of GFAP-positive astrocytes. Compound 1-treated mice showed a significant reduction in GFAP+ immunoreactivity (reduced total astrocyte numbers) in contrast to mice treated with LPS alone (Figure 29B).

These data demonstrate the potent anti-neuroinflammatory activity of compound 1 and its potential therapeutic potential to reduce neuroinflammation-induced neuronal toxicity in diseases manifested by neurodegeneration, cognitive or motor dysfunction.

11th experimental group (see FIG. 30): Forty-five 2-month-old C57BL/6 mice (42) were assigned to three groups: vehicle-treated control, vehicle-treated LPS mice, and Compound 1-treated LPS mice. Vehicle or compound was administered orally twice daily (PO BID) for a total of six doses. Histological evaluation was performed in the hippocampus using the microglial marker Iba-1.

Drug preparation: Compound 1 was formulated in 40% HP-β-cyclodextrin and adjusted to pH 6.5 with NaOH (1M). Vehicle solutions were prepared and pH adjusted similarly. Solutions were prepared fresh weekly and stored at 4°C.

LPS preparation: LPS was prepared in saline at 0.55 mg/ml and administered at 5 mg/kg.

Treatment Groups:
Treatment Group 1, vehicle-treated control: Young C57BL/6 mice (n=13) aged 2 months were given one IP injection of saline followed by vehicle PO BID for 3 days for a total of 6 injections.
Treatment group 2, LPS vehicle-treated: Young C57BL/6 mice (n=19) aged 2 months were given a single IP injection of LPS (5 mg/kg) followed by vehicle injections PO BID for 3 days for a total of 6 injections.
Treatment group 3, Compound 1-treated LPS: Young C57BL/6 mice (n=18) aged 2 months were given a single IP injection of LPS (5 mg/kg) followed by Compound 1 (30 mg/kg) PO BID for 3 days for a total of 6 injections.

Tissue processing and histology: Mouse hemibrains were sectioned at 30 μm on a microtome at −22°C. Brain slices were collected sequentially in 12 tubes, with every 12th section of the hippocampus being represented in a given tube. Brain sections were stored at −20°C in cryoprotective media until required for staining. Free-floating sections were blocked with appropriate serum in 10% serum in PBST 0.5%. Primary antibody Iba1 (Wako, 1:1000) was incubated overnight at 4°C. The next day, the appropriate fluorescent secondary antibody was applied at a concentration of 1:300 for 1 hour at room temperature. Slides were coverslipped using Prolong Gold Mounting Media.

Quantification of neuroinflammation: Iba-1 positive area was quantified as a percentage of the ROI around the entire hippocampus using thresholding in Image Pro Premier v9.2 software. Iba-1 positive percent area was averaged from approximately 5 sections for each mouse. Regular one-way ANOVA was used to test for statistical significance with Dunnett's post hoc multiple comparison test between treatment groups.

Figure 30 shows histological analysis of mouse brain in an acute model of LPS-induced inflammation. It reveals a significant reduction in LPS-induced microgliosis in the hippocampus of mice treated with Compound 1 for 3 days when both Compound 1 and LPS treatments were initiated on the same day. Microgliosis was measured by determining the percentage of Iba-1 positive area in the hippocampus. Ordinary one-way ANOVA was used to test for statistical significance with Dunnett's post hoc multiple comparison test between treatment groups ( ^* P<0.05, ^*** P<0.001).

12th experimental group (see FIG. 31): Fifteen (15) 2-month-old C57BL/6 mice were assigned to three groups: vehicle-treated control, vehicle-treated LPS mice, and Compound 1-treated LPS mice. Vehicle or compound was administered orally twice daily (PO BID) for a total of six doses. Histological evaluation was performed in the hippocampus using the microglial marker Iba-1.

Drug preparation: Compound 1 was formulated in 40% HP-β-cyclodextrin and adjusted to pH 6.5 with NaOH (1M). Vehicle solutions were prepared and pH adjusted similarly. Solutions were prepared fresh weekly and stored at 4°C.

LPS preparation: LPS was prepared in saline at 0.55 mg/ml and administered at 5 mg/kg.

Treatment Groups:
Treatment Group 1, vehicle-treated controls: Young C57BL/6 mice (n=13) aged 2 months were given one IP injection of saline followed by a 72 hour delay and vehicle injections PO BID over 3 days for a total of 6 injections.
Treatment group 2, vehicle-treated LPS: Young C57BL/6 mice (n=19) aged 2 months were given a single IP injection of LPS (5 mg/kg) followed by a 72 hour delay and vehicle injections PO BID over 3 days for a total of 6 injections.
Treatment group 3, Compound 1-treated LPS: Young C57BL/6 mice (n=18) aged 2 months were given a single IP injection of LPS (5 mg/kg) followed by a 72 hour delay and Compound 1 (30 mg/kg) PO BID for 3 days for a total of 6 injections.

Tissue processing and histology: Mouse hemibrains were sectioned at 30 μm on a microtome at −22°C. Brain slices were collected sequentially in 12 tubes, with every 12th section of the hippocampus being represented in a given tube. Brain sections were stored at −20°C in cryoprotective media until required for staining. Free-floating sections were blocked with appropriate serum in 10% serum in PBST 0.5%. Primary antibody Iba1 (Wako, 1:1000) was incubated overnight at 4°C. The next day, the appropriate fluorescent secondary antibody was applied at a concentration of 1:300 for 1 hour at room temperature. Slides were coverslipped using Prolong Gold Mounting Media.

Quantification of neuroinflammation: Iba-1 positive area was quantified as a percentage of the ROI around the entire hippocampus using thresholding in Image Pro Premier v9.2 software. Iba-1 positive percent area was averaged from approximately 5 sections for each mouse. Regular one-way ANOVA was used to test for statistical significance with Dunnett's post hoc multiple comparison test between treatment groups.

Figure 31 shows the results of histological analysis of mouse brain in an acute model of LPS-induced inflammation. It reveals a significant reduction in LPS-induced microgliosis in the hippocampus of mice treated with Compound 1 for 3 days when Compound 1 administration was initiated after LPS had been administered for the previous 3 days. Microgliosis was measured by determining the percentage of Iba-1 positive area in the hippocampus. Ordinary one-way ANOVA was used to test for statistical significance with Dunnett's post hoc multiple comparison test between treatment groups. ( ^* P<0.05, ^*** P<0.001).

3. Mouse MPTP Model of Parkinson's Disease A mouse model with MPTP (1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine) was used to determine the level of amelioration of MPTP-induced Parkinsonian-like deficits. MPTP is a prodrug of MPP+, which can cause permanent Parkinsonian symptoms. MPP+ acts by killing dopaminergic neurons in the substantia nigra region of the brain.

A total of 74 8-week-old male C57Bl/6J mice were used in the experiment. The animals were divided into two separate test arms, the 12-day and 4-day test arms, and treated similarly in both arms. Mice were divided into three groups within the test arms, one group received MPTP vehicle and compound vehicle as a control, and the other two groups received MPTP twice daily on days 1 and 2, plus compound vehicle or compound 1 daily (30 mg/kg) on days 1-12, with only a single dose on day 12 (Figure 32). After 10 days of treatment, on day 11 of the study, the motor function of the 12-day test arm mice was assessed with microkinetic analysis. Endpoint tissue processing was performed on days 12 and 4 of the study. Samples were processed for hematology, immunohistochemistry, and HPLC measurements. Striatal levels of dopamine (DA), 3,2-dihydroxyphenylacetic acid (DOPAC), and homovanic acid (HVA) were assessed by HPLC on day 12. Tyrosine hydroxylase (TH) immunoreactivity was assessed from brain sections collected through the substantia nigra.

Due to the large number of 97 individual parameters, a principal component analysis (PCA) was performed. PCA combined all parameter data together, revealed correlations between different parameters, and provided a global view of fine motor and gait characteristics. Figure 33A illustrates the visualization of the 10 principal components or PCs, showing how the original parameters are correlated within the data set. Strong intensity (blue or red) for each parameter indicates that the particular parameter is strongly involved in the corresponding PC. A heat map is formed from the PCA of all available parameter data. PC scores are presented as an overall gait analysis score in Figure 33B based on the difference between Group 2: MPTP+vehicle and Group 1: vehicle+vehicle mice in all PC scores.

Figure 34 shows the ambulatory characteristics of forepaw toe clearance on day 11 of the study. The MPTP + vehicle treatment group showed a statistically significant change in forepaw toe clearance compared to the vehicle + vehicle treatment group. The MPTP + Compound 1 treatment group showed a trend toward efficacy compared to the MPTP + vehicle treatment group. Data are mean + SEM (Group 1: Vehicle + Vehicle, n = 15 vs. Group 2: MPTP + Vehicle, n = 14 vs. Group 3: MPTP + Compound 1, n = 13). Statistical significance is ^* P < 0.05 for Group 2: MPTP + Vehicle vs. Group 1: Vehicle + Vehicle (unpaired t-test).

Figure 35 shows the gait characteristics of front paw swing velocity on day 11 of the study. The MPTP + vehicle treatment group showed a statistically significant change in front paw swing velocity compared to the vehicle + vehicle treatment group. The MPTP + Compound 1 treatment group showed a trend toward efficacy compared to the MPTP + vehicle treatment group. Data are mean + SEM (Group 1: Vehicle + Vehicle, n = 15 vs. Group 2: MPTP + Vehicle, n = 14 vs. Group 3: MPTP + Compound 1 (30 mg/kg), n = 13). Statistical significance is ^* P<0.05 for Group 2: MPTP + Vehicle vs. Group 1: Vehicle + Vehicle (unpaired t-test).

Figure 36 shows the ambulatory characteristics of ankle range of motion on day 11 of the study. The MPTP + vehicle treatment group showed a statistically significant change in ankle range of motion compared to the vehicle + vehicle treatment group. The MPTP + Compound 1 treatment group showed a trend toward efficacy compared to the MPTP + vehicle treatment group. Data are mean + SEM (Group 1: Vehicle + Vehicle, n = 15 vs. Group 2: MPTP + Vehicle, n = 14 vs. Group 3: MPTP + Compound 1 (30 mg/kg), n = 13). Statistical significance is ^* P<0.05 for Group 2: MPTP + Vehicle vs. Group 1: Vehicle + Vehicle (unpaired t-test).

Figure 37 shows the short-term effects of MPTP and Compound 1 on T cell infiltration into the brain. The total number of CD3 positive T cells counted in the substantia nigra from three 30 μm thick sections per mouse on day 4 of the study is shown. Data shown are mean ± SEM, ^*** P<0.001, one-way ANOVA, Sidak's post hoc multiple comparison test. Mice treated with MPTP + Compound 1 showed a trend towards fewer CD3 positive cells, indicating less T cell infiltration into the brain.

Figure 38 shows the short-term effects of MPTP and Compound 1 on microgliosis on day 4 of the study. Figure 38A shows the extent of CD68-positive area measured in the striatum from three sections of 30 μm thickness for each mouse on day 4 of the study. Figure 38B shows the extent of CD68-positive area measured in the substantia nigra pars compacta from three sections of 30 μm thickness for each mouse from day 4 of the study. These data demonstrate the presence of prominent microgliosis in the substantia nigra, the primary site of neuronal loss observed in Parkinson's disease, but not in the striatum, the main brain region to which neurons from the substantia nigra project, suggesting that the treatment affects the central site of disease pathology but not the secondary projection areas.

4. Synuclein transgenic mouse model of Parkinson's disease The synuclein transgenic mouse model was used to determine the effect of long-term CCR3 inhibition by compound 1 and its ability to slow, stop, or reverse the progression of Parkinson's disease-like symptoms. This model overexpressing human α-synuclein allows testing whether interventional treatments can prevent α-synuclein-induced behavioral and pathological effects. To select the optimal synuclein model for testing, plasma eotaxin levels were measured as a biomarker in multiple synuclein mouse models of different ages. These models included A53T, DxJ9M, and Line61. Plasma from 6-month-old Line61 synuclein transgenic mice was measured by ELISA assay to detect plasma eotaxin levels as a biomarker. Six-month-old 61LiNE mice (QPS Neuro, Grambach, Austria) showed a transgene-induced increase in eotaxin levels ( FIG. 39 ), indicating that plasma eotaxin levels may serve as a suitable clinical biomarker for selecting treatment populations.

Thirty-three 4.5-month-old male α-synonuclein transgenic mice (Line 61) were assigned to two groups (group A, n=17, compound 11 mg/mL; group B, n=16, vehicle) and a group of 15 non-transgenic matched littermates (group C, vehicle) were treated via their drinking water for 6 weeks. Animals received either vehicle or compound 1. Animal behavior was evaluated at the end of the treatment period. Tissue processing was then performed.

Figure 40 shows the results of the wire suspension test. The mean wire suspension time per group is shown, with animals in group C (non-transgenic, vehicle-treated) showing significantly higher wire suspension times. Animals in group A (transgenic, compound 1-treated) showed significantly higher wire suspension times compared to group B (transgenic, vehicle-treated). Data are presented as the mean + standard error of all animals per group, ^*** P<0.001, Dunn's post-hoc test, ^* P<0.05, Mann-Whitney test for group A vs. group B). These data demonstrate a dramatic loss of function in the ability to hang on the wire in synacrein mice, which was at least partially rescued by compound 1 treatment.

Figure 41 shows the results of the grip strength test. The mean maximum grip force [g] per group is shown. Animals in groups A and C (transgenic, compound 1-treated and non-transgenic, vehicle-treated, respectively) showed significantly higher grip strength compared to group B (transgenic, vehicle-treated). Data are presented as the mean + standard error of all animals per group. Groups were compared to vehicle-treated transgenic animals (group B) by one-way ANOVA followed by Bonferroni's post-hoc test. These data demonstrate a statistically significant deficit in synacrein mice in forelimb strength, which was completely rescued by compound 1 treatment.

Figure 42 shows the results of the beam-walk test. The number of animals per group per trial that were able to completely traverse the beam is shown. Group B animals (transgenic, vehicle-treated) were unable to traverse the fifth beam (the most difficult), and Group A transgenic mice treated with Compound 1 performed better than Group B transgenic mice treated with vehicle, indicating that performance in the beam-walk test approached that of non-transgenic controls.

Figure 43 shows the results of 5 trials of beam walking for three groups of mice (group A: transgenic compound 1-treated, group B: transgenic vehicle-treated, and group C: non-transgenic vehicle-treated) with slips or gaits performed in each trial. Graphs represent the mean number of slips [n] per group, and each graph represents one trial (1-5). Data are the mean + standard error of all animals per group. Each group was compared to group B by one-way ANOVA followed by Bonferroni post-hoc test. These data clearly show that synacrein-overexpressing mice are highly impaired in their ability to traverse the beam and that treatment with compound 1 at least partially rescues this effect, indicating a dramatic improvement in motor function.

Figure 44 shows eosinophil counts from peripheral blood. Figure 44A shows the percentage of eosinophils in peripheral blood from three mouse groups (Tg Cmpd1 = transgenic Compound 1-treated, Tg Veh = transgenic vehicle-treated, nTG Veh = non-transgenic vehicle-treated). Data were compared by t-test. Figure 44B shows absolute numbers of eosinophils in peripheral blood from three mouse groups (Tg Cmpd1 = transgenic Compound 1-treated, Tg Veh = transgenic vehicle-treated, nTG Veh = non-transgenic vehicle-treated). Data were compared by t-test. These data demonstrate that a Parkinson's disease model of synakrein overexpression results in a reduction in eosinophils that are restored to levels in non-transgenic mice with Compound 1 treatment, suggesting beneficial immune modulation in this model of Parkinson's disease. This also suggests that determining eosinophil levels in Parkinson's disease patients treated with Compound 1 could be a biomarker for the disease, including determining the level of disease progression, stagnation, or regression, as well as the efficacy of treatment.

Figure 45 shows the effect of Compound 1 on neuroinflammation. Figure 45A shows CD68 positive area quantified in the hippocampus of non-transgenic vehicle-treated mice, transgenic vehicle-treated mice, and transgenic Compound 1-treated mice (n=14, 12, and 15, respectively). Figure 45B shows CD68 positive area quantified in the striatum of non-transgenic vehicle-treated mice, transgenic vehicle-treated mice, and transgenic Compound 1-treated mice (n=15, 11, and 16, respectively). Figure 45C shows Iba1 positive area quantified in the hippocampus of non-transgenic vehicle-treated mice, transgenic vehicle-treated mice, and transgenic Compound 1-treated mice (n=14, 13, and 16, respectively). Figure 45D shows Iba1 positive area quantified in the striatum of non-transgenic vehicle-treated, transgenic vehicle-treated, and transgenic compound 1-treated mice (n=15, 11, and 16, respectively). Data are mean +/- SEM, ^* P<0.05. Figure 45E shows GFAP positive astrocytes quantified in the hippocampus of non-transgenic vehicle-treated, transgenic vehicle-treated, and transgenic compound 1-treated mice (n=14, 13, and 15, respectively). Figure 45F shows GFAP positive area quantified in the striatum of non-transgenic vehicle-treated, transgenic vehicle-treated, and transgenic compound 1-treated mice (n=15, 13, and 15, respectively). Data are mean +/- SEM, ^* P<0.05. FIG. 45G shows Iba-1 positive area quantified in the substantia nigra pars compacta of non-transgenic vehicle-treated, transgenic vehicle-treated, and transgenic Compound 1-treated mice.

These data demonstrate that while synacrein overexpression does not result in dramatic microgliosis as evidenced by CD68 and Iba1, or astrogliosis as evidenced by GFAP immunoreactivity, treatment with Compound 1 reduces microgliosis by both markers and astrogliosis by GFAP, potentially demonstrating a global inflammatory effect, i.e., not just on one inflammatory cell type. Furthermore, the striatum was more affected than the hippocampus, demonstrating that this region associated with Parkinson's disease was specifically reduced in microgliosis and astrogliosis markers. This is further evidenced by the effect of Compound 1 in reversing microgliosis in the pars compacta.

Figure 46 shows the effect of Compound 1 on circulating levels of IL-4 and IL-6 cytokines in non-transgenic vehicle-treated mice, transgenic vehicle-treated mice, and transgenic Compound 1-treated mice (n=14, 15, and 17, respectively). Figure 46A shows the levels of IL-4 measured in peripheral heart plasma of all three groups. Figure 46B shows the levels of IL-6 measured in peripheral heart plasma of all three groups. Data shown are mean ± SEM, ^* P<0.05, ^** P<0.01, one-way ANOVA, Dunnett's post hoc multiple comparison test. These changes in key cytokines indicate a general effect of Compound 1 treatment on proteins involved in immune function and inflammation.

5. T cell infiltration into the brain (a) Tissue processing and histology Mice were sacrificed 2 hours after the last PO dose the day after 9 days of dosing. Anesthesia was induced with 2,2,2-tribromoethanol. Mice were then perfused transcardially with 1% EDTA in PBS followed by 4% PFA in PBS. Brains were dissected, cut sagittally, bisected, and drop-fixed with 4% PFA in PBS. Two days after fixation, one brain was transferred to 30% sucrose in PBS solution, which changed after 2 days. Plasma was collected and stored on dry ice.

Hemibrains were sectioned sagittally at 35 μm on a microtome at −22°C. Brain slices were collected sequentially in 12 tubes, with every 12th section of the brain represented in a given tube. Brain sections were stored at −20°C in cryoprotective media until required for staining. Free-floating sections were blocked with the appropriate serum: 10% serum in PBST 0.5%. Primary antibodies were incubated overnight at 4°C or room temperature as described below.

Rat anti-CD8a (63-0081-80, Thermo Fisher Scientific) was used at a concentration of 1:100 at room temperature, rat anti-CD3 (555273, BD Biosciences) was used at a concentration of 1:100 at room temperature, rabbit anti-Iba-1 (016-26721, Wako) was used at a concentration of 1:1000 at 4°C, rat anti-CD68 (MCA1957, Bio-Rad) was used at a concentration of 1:1000 at 4°C, and Dilight 488/594-labeled Lycopersicon Esculentum (tomato) lectin (DL-1177, Fisher Scientific) was used at a concentration of 1:200 at room temperature. A suitable fluorescent secondary antibody (Alexa-488/555/647, Invitrogen) was applied the following day at a concentration of 1:300 for 1 hour at room temperature. Slides were coverslipped using Prolong Gold Mounting Media. Images were acquired at 20x on a Hamamatsu Nanozoomer 2.0HT slide scanner.

(b) T cell infiltration quantification CD3 and CD8 positive cells were counted from images acquired from the cerebellum on a Hamamatsu slide scanner. The total number of CD3 and CD8 positive cells in the cerebellum was summed from approximately 5 sections of 35 μm each. Regular one-way ANOVA was used to test for statistical significance with Dunnett's post hoc multiple comparison test between treatment groups.

(c) Neuroinflammation quantification. Iba-1 and CD68 positive areas were quantified as a percentage of an ROI around the entire cerebellum using thresholding in Image Pro Premier v9.2 software. Iba-1 and CD68 positive percent areas were averaged from approximately 5 sections per mouse. Regular one-way ANOVA was used to test for statistical significance with Dunnett's post hoc multiple comparison test between treatment groups.

Treatment groups: Two-month-old C57Bl/6 mice were divided into three treatment groups: vehicle control, vehicle-treated EAE (experimental autoimmune encephalomyelitis, Methods Mol Biol. 2012;900:381-401, incorporated herein by reference in its entirety), and compound 1-treated EAE. EAE was induced on day 0 with a subcutaneous (SQ) injection of MOG+CFA emulsion and an intravenous (IV) injection of Pertussis toxin (PT). A booster injection of PT occurred on day 2. Vehicle or compound 1 administration was administered by oral gavage (PO) on day 0 and continued twice daily (BID) until day 9. Mice were sacrificed 2 hours after the last oral dose.

Drug preparation: Compound 1 was formulated in 40% HP-β-cyclodextrin and adjusted to pH 6.5 with NaOH (1M). Vehicle solutions were prepared and pH adjusted similarly. Solutions were prepared fresh weekly and stored at 4°C.

EAE Emulsion: MOG35-55 (Anaspec AS-60130-5) was reconstituted in PBS at 4 mg/ml. M. tuberculosis H37 Ra (Fisher Scientific DF3114-33-8) was dissolved in Freud's incomplete adjuvant at 4 mg/ml. These two solutions were then emulsified using a glass syringe (Thermo Scientific Male Luer-LOK Priming Syringe 03-170-301) and a 3-way stopcock (Cadence 6001). The solution was prepared immediately prior to dosing.

Pertussis toxin: Pertussis toxin (Sigma P7208-50UG) was dissolved in saline at 0.002mg/ml and injected 100ul IV on the day of induction and again 2 days later.

Treatment Groups:
Treatment Group 1 (vehicle treatment): Young C57BL/6 mice (n=5), 2.5 months of age, received 100 ul of vehicle PO BID for 9 days starting 2 hours after saline SQ injection, with only one treatment injection on the first and last day for a total of 19 treatment injections.
Treatment group 2 (vehicle treatment with EAE induction): Young C57BL/6 mice (n=10) aged 2.5 months were administered 100 ul of vehicle PO BID for 9 days starting 2 hours after EAE induction, with only one treatment injection on the first and last day for a total of 19 treatment injections.
Treatment group 3 (treatment with Compound 1): Young C57BL/6 mice (n=10), 2.5 months of age, were administered 100 ul of Compound 1 (30 mg/kg) PO BID for 9 days, starting 2 hours after EAE induction, with only one treatment injection on the first and last day, for a total of 19 treatment injections.

EAE induction resulted in an increase in CD3 and CD8 positive infiltrating T cells in the cerebellum. These T cell numbers were significantly reduced with treatment with compound 1. EAE induction also resulted in a significant increase in microgliosis in the cerebellum, which was also significantly rescued by compound 1 treatment after 9 days. Figure 47A shows an increase in CD3 positive infiltrating T cells in the cerebellum, which was significantly reduced after 9 days of treatment with compound 1. Figure 47B shows an increase in CD8 positive infiltrating T cells in the cerebellum, which was significantly reduced after 9 days of treatment with compound 1. Figure 47C shows a significant increase in Iba-1 positive area in the cerebellum after EAE, which was significantly reduced in the cerebellum after 9 days of treatment. Figure 47D shows a significant increase in CD68 positive area in the cerebellum after EAE, which was significantly reduced in the cerebellum after 9 days of treatment.

d. Example 1. Eotaxin Levels and Aging in Humans Human eotaxin-1 levels were determined using a commercially available affinity-based assay (SOMAscan, SomaLogic, Inc., Boulder, Colorado). Plasma samples were collected from donors aged 18, 30, 45, 55, and 66 years for testing by SomaLogic using SOMAscan, an aptamer-based affinity assay that tests for relative levels of human eotaxin-1, among others. Eotaxin-1 levels were determined and plotted by age group (FIG. 45). Relative eotaxin-1 concentrations increased with age, indicating that the eotaxin-1 pathway, including its primary receptor CCR3, is a therapeutic target for age-related diseases such as neurodegenerative diseases and cognitive decline.

2. Human Biomarker Assays Whole blood from humans treated with Compound 1 was incubated with human recombinant eotaxin-1 to induce eosinophil morphological changes (FIG. 48A) or CCR3 receptor internalization (FIG. 49B).

Both assays demonstrated that compound 1 exerted a potent concentration-dependent effect on the readings of each functional biomarker.

Together, the results from ESC and CCR3 receptor internalization assays confirm that compound 1 functions as an inhibitor of the human eotaxin-1 pathway. In particular, compound 1 can function as a potent inhibitor of CCR3 by binding to that receptor.

Furthermore, the data obtained in Figure 44 show that in mammalian transgenic Parkinson's disease models, eosinophils were expressed at lower levels than in non-transgenic animals and were reversed by administration of Compound 1. Thus, this data, when applied to Parkinson's disease patients, may be useful in diagnosing, monitoring, and determining the prognosis of the disease.

3. Treatment of subjects with Parkinson's disease using Compound 1 Subjects diagnosed with Parkinson's disease are orally administered 400 mg of compound or placebo twice daily (BID). Subjects are treated for 12 weeks, with follow-up for 2 weeks thereafter. Subjects are evaluated for the effect of Compound 1 on motor function in a practically defined drug-free state where levodopa is discontinued for 12 hours or more. Subjects are also evaluated by flow cytometry, pharmacogenomics, and biomarker analysis performed on blood and plasma samples, including eosinophil levels. Gait analysis is evaluated using Zeno Walkway. Bradykinesia, tremor, general activity, and sleep are evaluated using wearable devices.

Changes in baseline (day 1) motor function during the pragmatically defined drug-free state at week 12 will be determined using the Movement Disorder Society Unified Parkinson's Disease Rating Scale (MDS-UPDRS) part 3. Changes in baseline (day 1) clinical function, motor function, and activities of daily living during the drug-free state at week 12 will be assessed with the MDS-UPDRS parts 1-4, Montreal Cognitive Assessment (MoCA), Schwab and England Activities of Daily Living (SE-ADL) scale, Clinical Impression of Severity Index-PD (CISI-PD), PD Quality of Life Questionnaire-39 (PDQ-39), Sheehan-Suicidality Tracking Scale (S-STS), and the 10-meter timed walk (also assessed in the drug-free state).

It is to be understood that the invention is not limited to the specific embodiments described, as these may vary. It is also to be understood that the terminology used herein is for the purpose of describing the specific embodiments only, and is not intended to be limiting, since the scope of the invention will be limited only by the appended claims.

When a range of values is given, it is understood that each intermediate value between the upper and lower limits of the range, to the nearest tenth of the lower limit of the range, and any other stated or intermediate value in the stated range, is included in the invention, unless the context clearly indicates otherwise. The upper and lower limits of these narrower ranges may be independently included in the narrower range, and are also included in the invention, subject to any specifically excluded limit in the stated range. When a stated range includes one or both of the limits, ranges excluding either or both of those included limits are also included in the invention.

Unless otherwise specified, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention belongs. Although any methods and materials similar or equivalent to those described herein can be used in practicing or testing the present invention, representative illustrative methods and materials are described above.

All publications and patents cited herein are incorporated by reference as if each individual publication or patent was specifically and individually indicated to be incorporated by reference, and are incorporated by reference herein for the purpose of disclosing and describing the methods and/or materials in connection with which the publication is cited. The citation of any publication is for its disclosure prior to the filing date and should not be construed as an admission that the present invention is not entitled to antedate such publication by virtue of prior invention. Further, the publication dates provided may be different from the actual publication dates, which may need to be independently confirmed.

As used herein and in the appended claims, it should be noted that the singular forms "a," "an," and "the" include plural referents unless the context clearly dictates otherwise. It should be further noted that the claims may be written to the exclusion of any optional element. As such, this statement is intended to serve as an underlying antecedent for the use of exclusive language such as "solely," "only," and the like, or for the use of "negative" limitations in connection with the recitation of claim elements.

As will be apparent to one of ordinary skill in the art upon reading this disclosure, each of the individual embodiments described and illustrated herein has distinct components and features which may be readily separated from or combined with the features of any of the other embodiments without departing from the scope or spirit of the invention. Any method shown can be carried out in the order of events shown or in any other order which is logically possible.

Although the foregoing invention has been described in some detail by way of illustrations and examples for purposes of clarity of understanding, it will be readily apparent to those skilled in the art in light of the teachings of the present invention that certain changes and modifications may be made thereto without departing from the spirit or scope of the appended claims.

The foregoing is thus merely illustrative of the principles of the present invention. It is clear that those skilled in the art will be able to devise various configurations which embody the principles of the present invention and are within its spirit and scope, although not expressly described or shown herein. Furthermore, any examples and specific terminology presented herein are intended, in principle, to aid the reader in understanding the principles and concepts of the present invention which the inventors have contributed to the advancement of the art, and should not be construed as being limited to the examples and conditions so specifically shown. Furthermore, any statements herein presenting principles, aspects and embodiments of the present invention, as well as specific examples thereof, are intended to include both structural and functional equivalents thereof.

Additionally, it is intended that such equivalents include both currently known equivalents and equivalents developed in the future, i.e., any elements developed that perform the same function, regardless of structure. Thus, the scope of the present invention is not intended to be limited to the exemplary embodiments shown and described herein. Rather, the scope and spirit of the present invention is embodied by the appended claims.

CROSS-REFERENCE TO RELATED APPLICATIONS This application claims priority pursuant to 35 U.S.C. § 119(e) to the filing date of U.S. Provisional Patent Application No. 62/737,017, filed September 26, 2018, the disclosure of which is incorporated herein by reference.

Claims (18)

A pharmaceutical composition for use in a method for systemically treating age-related neurodegeneration, age-related cognitive decline, or age-related motor decline in a subject diagnosed with said age-related neurodegeneration, age-related cognitive decline, or age-related motor decline, comprising:
A compound of formula 1 below

In the formula,
A is CH ₂ , O or N—C _1-6 alkyl;
^R1 is
・NHR ^1.1 , NMeR ^1.1 ,
・NHR ^1.2 , NMeR ^1.2 ,
・NHCH ₂ -R ^1.3 ,
NH-C _3-6 cycloalkyl, optionally in which one carbon atom is replaced by a nitrogen atom and the ring is optionally substituted with one or two residues selected from the group consisting of C _1-6 alkyl, O-C _1-6 alkyl, NHSO ₂ -phenyl, NHCONH _- phenyl, halogen, CN, SO ₂ -C _1-6 alkyl, COO-C _1-6 alkyl,
a C _9-10 bicyclic ring in which one or two carbon atoms are replaced by a nitrogen atom, the ring system being linked to the basic structure of formula 1 via the nitrogen atom, said ring system being optionally substituted with one or two residues selected from the group consisting of C _{1-6 alkyl} , COO-C _1-6 alkyl, C _1-6 haloalkyl, O-C _1-6 alkyl, NO ₂ , halogen, CN, NHSO ₂ -C _1-6 alkyl, methoxyphenyl;
- a group selected from NHCH(pyridinyl)CH ₂ COO-C _1-6 alkyl, NHCH(CH ₂ O-C _1-6 alkyl)-benzimidazolyl, optionally substituted with halogen or CN ,
1-aminocyclopentyl optionally substituted with methyl-oxadiazole , or
piperidinyl optionally substituted with one or two residues selected from the group consisting of NHSO ₂ -C _1-4 alkyl, m-methoxyphenyl;
is selected from
R ^1.1 is phenyl, optionally C _1-6 alkyl, C _2-6 alkenyl, C _2-6 alkynyl, C _1-6 haloalkyl, C _1-6 alkylene-OH, C 2-6 alkenylene- _{OH, C 2-6 alkynylene-OH, CH 2 CON(C 1-6 alkyl) 2 , CH 2 NHCONH-C 3-6 cycloalkyl , CN , CO -} pyridinyl, CONR ^1.1.1 R ^1.1.2 , COO-C _1-6 alkyl, N(SO ₂ -C _1-6 alkyl)(CH ₂ CON(C _1-4 alkyl) ₂ )O-C _1-6 alkyl, O-pyridinyl, SO ₂ -C _1-6 alkyl, SO ₂ -C _1-6 alkylene-OH, SO ₂ -C _{3-6cycloalkyl} , SO ₂ -piperidinyl, SO ₂ NH-C _1-6alkyl , SO ₂ N(C _1-6alkyl ) ₂ , halogen, CN, CO-morpholinyl, CH ₂ -pyridinyl or a heterocycle optionally substituted with 1 or 2 residues selected from the group consisting of C _1-6alkyl , NHC _1-6alkyl and ═O,
R ^1.1.1 is H, C _1-6 alkyl, C _3-6 cycloalkyl, C _1-6 haloalkyl, CH ₂ CON(C _1-6 alkyl) ₂ , CH ₂ CO-azetindinyl, C _1-6 alkylene-C _3-6 cycloalkyl, CH ₂ -pyranyl, CH ₂ -tetrahydrofuranyl, CH ₂ -furanyl, C _1-6 alkylene-OH or thiadiazolyl, optionally substituted with C _1-6 alkyl;
R ^1.1.2 is H, C _1-6 alkyl, SO ₂ C _1-6 alkyl;
or R ^1.1.1 and R ^1.1.2 together form a 4-, 5- or 6-membered carbocyclic ring, which optionally contains one N or O, replacing a carbon atom of said ring, optionally substituted with one or two residues selected from the group consisting of C _1-6 alkyl, C _1-4 alkylene-OH, OH, ═O;
or,
R ^1.1 is phenyl and two adjacent residues together form a 5- or 6-membered carbocyclic aromatic or non-aromatic ring, said ring optionally containing, independently of each other, one or two N, S or SO ₂ , replacing a carbon atom of said ring, said ring optionally being substituted with C _1-4 alkyl or ═O;
R ^1.2 is
heteroaryl optionally substituted with one or two residues selected from the group consisting of C _1-6 alkyl, C _2-6 alkenyl, C _2-6 alkynyl, C _3-6 cycloalkyl, CH ₂ COO-C _1-6 alkyl, CONR ^1.2.1 R ^1.2.2 , COR ^1.2.3 , COO-C _1-6 alkyl, CONH ₂ , O-C _1-6 alkyl, halogen, CN, SO ₂ N(C _1-6 alkyl) ₂ , or heteroaryl optionally substituted with one or two residues selected from the group consisting of C _1-6 alkyl,
Heteroaryl optionally substituted with a 5- or 6-membered carbocyclic non-aromatic ring, said ring containing, independently of each other, two N, O, S or _SO2 , replacing the carbon atoms of said ring;
an aromatic or non-aromatic C _9-10 bicycle in which one or two carbon atoms are replaced by N, O or S, each optionally substituted by one or two residues selected from the group consisting of N(C _1-6 alkyl) ₂ , CONH- C _{1-6 alkyl} , ═O;
- optionally pyridinyl-substituted heterocyclic non-aromatic rings,
4,5-dihydro-naphtho[2,1-d]thiazole optionally substituted with NHCO-C _1-6 alkyl,
R ^1.2.1 is H, C _1-6 alkyl, C _1-6 alkylene-C _3-6 cycloalkyl, C _1-4 alkylene-phenyl, C _1-4 alkylene-furanyl, C _3-6 cycloalkyl, C _1-4 alkylene-O-C _1-4 alkyl, C _1-6 haloalkyl, or a 5- or 6-membered carbocyclic non-aromatic ring which optionally contains, independently of each other, one or two N, O, S or SO ₂ , said ring carbon atoms being replaced and optionally substituted with 4-cyclopropylmethyl-piperazinyl;
R ^1.2.2 is H, C _1-6 alkyl;
^R1.2.3 is a 5- or 6-membered carbocyclic non-aromatic ring, said ring optionally containing, independently of each other, one or two N, O, S or _SO2 , replacing a carbon atom of said ring;
R ^1.3 is selected from phenyl, heteroaryl or indolyl, each optionally substituted with one or two residues selected from the group consisting of C _1-6 alkyl, C _3-6 cycloalkyl, O-C _1-6 alkyl, O-C _1-6 haloalkyl, phenyl, heteroaryl;
R ² is selected from the group consisting of C _1-6 alkylene-phenyl, C _1-6 alkylene-naphthyl and C _1-6 alkylene-heteroaryl, each of which is optionally substituted with 1, 2 or 3 residues selected from the group consisting of C _1-6 alkyl, C _1-6 haloalkyl, O—C _1-6 alkyl, O—C _1-6 haloalkyl, halogen;
^R3 is H, _C1-6 alkyl;
R ⁴ is H, C _1-6 alkyl, or
or R ³ and R ⁴ together form a CH ₂ -CH ₂ group;
A therapeutically effective amount of
Pharmaceutical compositions. In the compound of formula 1,
A is CH ₂ , O or N—C _1-4 alkyl;
^R1 is
・NHR ^1.1 , NMeR ^1.1 ,
・NHR ^1.2 , NMeR ^1.2 ,
・NHCH ₂ -R ^1.3 ,
NH-C _3-6 cycloalkyl, optionally in which one carbon atom is replaced by a nitrogen atom and the ring is optionally substituted with one or two residues selected from the group consisting of C _1-6 alkyl, O-C _1-6 alkyl, NHSO ₂ -phenyl, NHCONH _- phenyl, halogen, CN, SO ₂ -C _1-6 alkyl, COO-C _1-6 alkyl,
a C _9-10 bicyclic ring in which one or two carbon atoms are replaced by a nitrogen atom, the ring system being linked to the basic structure of formula 1 via the nitrogen atom, said ring system being optionally substituted with one or two residues selected from the group consisting of C _1-6 alkyl, COO-C _1-6 alkyl, C _1-6 haloalkyl, O-C _1-6 alkyl, NO ₂ , halogen, CN, NHSO ₂ -C _1-6 alkyl, m-methoxyphenyl _;
a radical selected from NHCH(pyridinyl)CH ₂ COO—C _1-6 alkyl, NHCH(CH ₂ O—C _1-6 alkyl)-benzimidazolyl, optionally substituted with Cl, or a radical selected from 1-aminocyclopentyl, optionally substituted with methyloxadiazolyl,
R ^1.1 is phenyl, optionally C _1-6 alkyl, C _1-6 haloalkyl, CH ₂ CON(C _1-6 alkyl) ₂ , CH ₂ NHCONH-C _3-6 cycloalkyl, CN, CONR ^1.1.1 R ^1.1.2 , COO-C _1-6 alkyl, O-C _1-6 alkyl, SO ₂ -C _1-6 alkyl, SO ₂ -C _1-6 alkylene-OH, SO ₂ -C _3-6 cycloalkyl, SO ₂ -piperidinyl, SO ₂ NH-C _1-6 alkyl, SO ₂ N(C _1-6 alkyl) ₂ , halogen, CN, CO-morpholinyl, CH ₂ -pyridinyl, or optionally C _1-6 alkyl, NHC _1-6 alkyl, ═O;
R ^1.1.1 is H, C _1-6 alkyl, C _3-6 cycloalkyl, C _1-6 haloalkyl, CH ₂ CON(C _1-6 alkyl) ₂ , CH ₂ CO-azetindinyl, C _1-6 alkylene-C _3-6 cycloalkyl, CH ₂ -pyranyl, CH ₂ -tetrahydrofuranyl, CH ₂ -furanyl, C _1-6 alkylene-OH or thiadiazolyl, optionally substituted with C _1-6 alkyl;
R ^1.1.2 is H, C _1-6 alkyl, SO ₂ C _1-6 alkyl;
or R ^1.1.1 and R ^1.1.2 together form a 4-, 5- or 6-membered carbocyclic ring, which optionally contains one O, and which replaces a carbon atom of the ring, optionally substituted with one or two residues selected from the group consisting of CH ₂ OH;
R ^1.2 is
heteroaryl optionally substituted with one or two residues selected from the group consisting of C _1-6 alkyl, C _3-6 cycloalkyl, CH ₂ COO-C _1-6 alkyl, CONR ^1.2.1 R ^1.2.2 , COO-C _1-6 alkyl, CONH ₂ , O-C _1-6 alkyl, halogen, CN, CO-pyrrolidinyl, CO-morpholinyl, or heteroaryl optionally substituted with one or two residues selected from the group consisting of C _1-6 alkyl,
- benzothiazolyl, indazolyl, dihydroindolyl, indanyl, tetrahydroquinolinyl, each of which is optionally substituted with one or two residues selected from the group consisting of N(C _1-6 alkyl) ₂ , CONH-C _1-6 alkyl, ═O;
piperidinyl optionally substituted with pyridinyl,
4,5-dihydro-naphtho[2,1-d]thiazole optionally substituted with NHCO-C _1-6 alkyl,
R ^1.2.1 is H, C _1-6 alkyl;
R ^1.2.2 is H, C _1-6 alkyl;
R ^1.3 is selected from phenyl, pyrazolyl, isoxazolyl, pyrimidinyl, indolyl or oxadiazolyl, each optionally substituted with one or two residues selected from the group consisting of C _1-6 alkyl, C _3-6 cycloalkyl, O-C _1-6 alkyl, O-C _1-6 haloalkyl;
R ² is selected from CH 2 -phenyl or CH 2 -naphthyl, each optionally substituted with one or two residues selected from the group consisting of C _1-6 alkyl, C _1-6 haloalkyl, O—C _1-6 alkyl, O—C _1-6 haloalkyl, halogen, or CH ₂ -thiophenyl, optionally substituted with _one or two residues selected from the group consisting of halogen _;
^R3 is H, _C1-4 alkyl;
R ⁴ is H, C _1-4 alkyl, or
or R ³ and R ⁴ together form a CH ₂ -CH ₂ group;
The pharmaceutical composition of claim 1. In the compound of formula 1,
A is _CH2 , O or NMe;
^R1 is
・NHR ^1.1 , NMeR ^1.1 ,
・NHR ^1.2 , NMeR ^1.2 ,
・NHCH ₂ -R ^1.3 ,
NH-cyclohexyl optionally substituted with one or two residues selected from the group consisting of C _1-4 alkyl, NHSO ₂ -phenyl, NHCONH-phenyl, halogen;
NH-pyrrolidinyl optionally substituted with one or two residues selected from the group consisting of SO ₂ -C _1-4 alkyl, COO-C _1-4 alkyl,
piperidinyl optionally substituted with one or two residues selected from the group consisting of NHSO ₂ -C _1-4 alkyl, m-methoxyphenyl;
dihydroindolyl, dihydroisoindolyl, tetrahydroquinolinyl or tetrahydroisoquinolinyl optionally substituted with one or two residues selected from the group consisting of C _1-4 alkyl, COO-C _1-4 alkyl, C _1-4 haloalkyl, O-C _1-4 alkyl, NO ₂ , halogen,
a radical selected from NHCH(pyridinyl)CH ₂ COO—C _1-4 alkyl, NHCH(CH ₂ O—C _1-4 alkyl)-benzimidazolyl, optionally substituted with Cl, or a radical selected from 1-aminocyclopentyl, optionally substituted with methyloxadiazolyl,
R ^1.1 is phenyl, optionally C _1-4 alkyl, C _1-4 haloalkyl, CH ₂ CON(C _1-4 alkyl) ₂ , CH ₂ NHCONH-C _3-6 cycloalkyl, CN, CONR ^1.1.1 R ^1.1.2 , COO-C _1-4 alkyl, O-C _1-4 alkyl, SO ₂ -C _1-4 alkyl, SO ₂ -C _1-4 alkylene-OH, SO ₂ -C _3-6 cycloalkyl, SO ₂ -piperidinyl, SO ₂ NH-C _1-4 alkyl, SO ₂ N(C _1-4 alkyl) ₂ , halogen, CO-morpholinyl, CH ₂ -pyridinyl, or each optionally C _1-4 alkyl, NHC _1-4 alkyl, ═O; substituted with 1 or 2 residues selected from the group consisting of imidazolidinyl, piperidinyl, oxazinanyl, pyrazolyl, triazolyl, tetrazolyl, oxazolyl, oxadiazolyl, thiazolyl, pyridinyl, pyrimidinyl;
R ^1.1.1 is H, C _1-6 alkyl, C _3-6 cycloalkyl, C _1-4 haloalkyl, CH ₂ CON(C _1-4 alkyl) ₂ , CH ₂ CO-azetindinyl, C _1-4 alkylene-C _3-6 cycloalkyl, CH ₂ -pyranyl, CH ₂ -tetrahydrofuranyl, CH ₂ -furanyl, C _1-4 alkylene-OH or thiadiazolyl, optionally substituted with C _1-4 alkyl;
R ^1.1.2 is H, C _1-4 alkyl, SO ₂ C _1-4 alkyl, or
or R ^1.1.1 and R ^1.1.2 together form a 4-, 5- or 6-membered carbocyclic ring, which optionally contains one O, and which replaces a carbon atom of the ring, optionally substituted with one or two residues selected from the group consisting of CH ₂ OH;
R ^1.2 is
pyridinyl, pyridazinyl, pyrrolyl _, pyrazolyl, isoxazolyl, thiazolyl, thiadiazolyl, optionally substituted by one or two residues selected from the group consisting of C _1-4 alkyl, C _3-6 cycloalkyl, CH ₂ COO-C _1-4 alkyl, CONR ^1.2.1 R ^1.2.2 , COO-C _1-4 alkyl, CONH ₂ , O-C _1-4 alkyl, halogen, CO-pyrrolidinyl, CO-morpholinyl or pyrazolyl, triazolyl, tetrazolyl, isoxazolyl, oxadiazolyl, each optionally substituted by one or two residues selected from the group consisting of C 1-4 alkyl,
- benzothiazolyl, indazolyl, dihydroindolyl, indanyl, tetrahydroquinolinyl, each optionally substituted with one or two residues selected from the group consisting of N(C _1-4 alkyl) ₂ , CONH-C _1-4 alkyl, ═O;
piperidinyl optionally substituted with pyridinyl,
4,5-dihydro-naphtho[2,1-d]thiazoles optionally substituted with NHCO-C _1-4 alkyl,
R ^1.2.1 is H, C _1-4 alkyl;
R ^1.2.2 is H, C _1-4 alkyl;
R ^1.3 is selected from phenyl, pyrazolyl, isoxazolyl, pyrimidinyl, indolyl or oxadiazolyl, each optionally substituted with one or two residues selected from the group consisting of C _1-4 alkyl, C _3-6 cycloalkyl, O-C _1-4 alkyl, O-C _1-4 haloalkyl;
R ² is selected from CH 2 -phenyl or CH ₂ -naphthyl, each optionally substituted with one or two residues selected from the group consisting of C _1-4 alkyl, C _1-4 haloalkyl, O—C _1-4 haloalkyl, halogen, or CH ₂ -thiophenyl, optionally substituted with _one or two residues selected from the group consisting of halogen;
^R3 is H;
^R4 is H or
or R ³ and R ⁴ together form a CH ₂ -CH ₂ group;
The pharmaceutical composition of claim 1. In Formula 1,
A is _CH2 , O or NMe;
^R1 is
・NHR ^1.1 , NMeR ^1.1 ,
・NHR ^1.2 , NMeR ^1.2 ,
・NHCH ₂ -R ^1.3 ,
NH-piperidinyl optionally substituted with pyridinyl,
NH-cyclohexyl, optionally substituted with one or two residues selected from the group consisting of t-Bu, NHSO ₂ -phenyl, NHCONH-phenyl, F;
NH-pyrrolidinyl, optionally substituted with one or two residues selected from the group consisting of SO ₂ Me, COO-t-Bu,
piperidinyl optionally substituted with one or two residues selected from the group consisting of NHSO ₂ -n-Bu, m-methoxyphenyl;
dihydroindolyl, dihydroisoindolyl, tetrahydroquinolinyl or tetrahydroisoquinolinyl, optionally substituted with one or two residues selected from the group consisting of Me, COOMe, CF ₃ , OMe, NO ₂ , F, Br,
a radical selected from NHCH(pyridinyl)CH ₂ COOMe, NHCH(CH ₂ OMe)-benzimidazolyl, optionally substituted by Cl, or 1-aminocyclopentyl, optionally substituted by methyloxadiazolyl,
R ^1.1 is phenyl, optionally Me, Et, t-Bu, CF ₃ , CH ₂ CONMe ₂ , CH ₂ NHCONH-cyclohexyl, CN, CONR ^1.1.1 R ^1.1.2 , COOMe, COOEt, OMe, SO ₂ Me, SO ₂ CH ₂ CH ₂ OH, SO ₂ Et, SO ₂ -cyclopropyl, SO ₂ -piperidinyl, SO ₂ NHEt, SO ₂ NMeEt, F, Cl, CO-morpholinyl, CH ₂ -pyridinyl or one or two residues selected from the group consisting of imidazolidinyl, piperidinyl, oxazinanyl, pyrazolyl, triazolyl, tetrazolyl, oxazolyl, oxadiazolyl, thiazolyl, pyridinyl, pyrimidinyl, each of which is optionally substituted with one or two residues selected from the group consisting of Me, NHMe, =O,
R ^1.1.1 is H, Me, Et, t-Bu, i-Pr, cyclopropyl, CH ₂ -i-Pr, CH ₂ -t-Bu, CH(CH ₃ )CH ₂ CH ₃ , CH ₂ CHF ₂ , CH ₂ CONMe ₂ , CH ₂ CO-azetindinyl, CH ₂ -cyclopropyl, CH ₂ -cyclobutyl, CH ₂ -pyranyl, CH ₂ -tetrahydrofuranyl, CH ₂ -furanyl, CH ₂ CH ₂ OH or thiadiazolyl, optionally substituted with Me;
R ^1.1.2 is H, Me, Et, SO ₂ Me, SO ₂ Et;
or R ^1.1.1 and R ^1.1.2 together form a 4-, 5- or 6-membered carbocyclic ring, which optionally contains one O, and which replaces a carbon atom of the ring, optionally substituted with one or two residues selected from the group consisting of CH ₂ OH;
R ^1.2 is
- optionally with Me, Et, Pr, Bu, cyclopropyl, CH ₂ COOEt, CONR ^1.2.1 R ^1.2.2 , COOMe, COOEt, CONH ₂ , OMe, Cl, Br, CO-pyrrolidinyl, CO-morpholinyl or pyridinyl, pyrrolyl, pyrazolyl, isoxazolyl, thiazolyl, thiadiazolyl, each optionally substituted with Me by one or two residues selected from the group consisting of pyrazolyl, triazolyl, tetrazolyl, isoxazolyl, oxadiazolyl,
- benzothiazolyl, indazolyl, dihydroindolyl, indanyl, tetrahydroquinolinyl, each optionally substituted with one or two residues selected from the group consisting of _NMe2 , CONHMe, =O;
4,5-dihydro-naphtho[2,1-d]thiazoles optionally substituted with NHCOMe,
R ^1.2.1 is H, Me;
R ^1.2.2 is H, Me;
^R1.3 is selected from phenyl, pyrazolyl, isoxazolyl, pyrimidinyl, indolyl or oxadiazolyl, each optionally substituted with one or two residues selected from the group consisting of Me, Et, Pr, cyclopentyl, OMe, _OCHF2 ;
^R2 is selected from CH2-phenyl or CH2-naphthyl, each optionally substituted with one or two residues selected from the group consisting of _CH3 , _CF3 , _OCF3 , F, _Cl , Br, Et, or _CH2 -thiophenyl, optionally substituted with one or two residues selected from the group consisting of _Cl , Br;
^R3 is H;
^R4 is H or
or R ³ and R ⁴ together form a CH ₂ -CH ₂ group;
The pharmaceutical composition of claim 1. In Formula 1,
A is _CH2 , O or NMe;
^R1 is
・NHR ^1.1 ,
・NHR ^1.2
is selected from
R ^1.1 is phenyl, optionally Me, Et, Bu, CF ₃ , CH ₂ CONMe ₂ , CH ₂ NHCONH-cyclohexyl, CN, CONR ^1.1.1 R ^1.1.2 , COOMe, COOEt, OMe, SO ₂ Me, SO ₂ CH ₂ CH ₂ OH, SO ₂ Et, SO ₂ -cyclopropyl, SO ₂ -piperidinyl, SO ₂ NHEt, SO ₂ NMeEt, F, Cl, CO-morpholinyl, CH ₂ -pyridinyl or one or two residues selected from the group consisting of imidazolidinyl, piperidinyl, oxazinanyl, pyrazolyl, triazolyl, tetrazolyl, oxazolyl, oxadiazolyl, thiazolyl, pyridinyl, pyrimidinyl, each of which is optionally substituted with one or two residues selected from the group consisting of Me, NHMe, =O,
R ^1.1.1 is H, Me, Et, t-Bu, i-Pr, cyclopropyl, CH ₂ -i-Pr, CH ₂ -t-Bu, CH(CH ₃ )CH ₂ CH ₃ , CH ₂ CHF ₂ , CH ₂ CONMe ₂ , CH ₂ CO-azetindinyl, CH ₂ -cyclopropyl, CH ₂ -cyclobutyl, CH ₂ -pyranyl, CH ₂ -tetrahydrofuranyl, CH ₂ -furanyl, CH ₂ CH ₂ OH or thiadiazolyl, optionally substituted with Me;
R ^1.1.2 is H, Me, Et, SO ₂ Me, SO ₂ Et;
or R ^1.1.1 and R ^1.1.2 together form a 4-, 5- or 6-membered carbocyclic ring, which optionally contains one O, and which replaces a carbon atom of the ring, optionally substituted with one or two residues selected from the group consisting of CH ₂ OH;
R ^1.2 is
- optionally with Me, Et, Pr, Bu, cyclopropyl, CH ₂ COOEt, CONR ^1.2.1 R ^1.2.2 , COOMe, COOEt, CONH ₂ , OMe, Cl, BrCO-pyrrolidinyl, CO-morpholinyl or pyridinyl, pyrrolyl, pyrazolyl, isoxazolyl, thiazolyl, thiadiazolyl, each optionally substituted with Me by one or two residues selected from the group consisting of pyrazolyl, triazolyl, tetrazolyl, isoxazolyl, oxadiazolyl,
- benzothiazolyl, indazolyl, dihydroindolyl, indanyl, tetrahydroquinolinyl, each optionally substituted with one or two residues selected from the group consisting of _NMe2 , CONHMe, =O;
4,5-dihydro-naphtho[2,1-d]thiazoles optionally substituted with NHCOMe,
R ^1.2.1 is H, Me;
R ^1.2.2 is H, Me;
^R2 is selected from _CH2 -phenyl or _CH2 -naphthyl, each of which is optionally substituted with one or two residues selected from the group consisting of _CH3 , _CF3 , _OCF3 , F, Cl, Br, Et;
^R3 is H;
^R4 is H;
The pharmaceutical composition of claim 1. In Formula 1,
A is _CH2 , O or NMe;
R ¹ is selected from the following:

^R2 is selected from the following:

^R3 is H;
^R4 is H or
or R ³ and R ⁴ together form a CH ₂ -CH ₂ group;
The pharmaceutical composition of claim 1. The compound is a co-crystal of the formula:

In the formula,
R ¹ is C _1-6 alkyl, C _1-6 haloalkyl, O—C _1-6 haloalkyl, halogen;
m is 1, 2 or 3;
R ^2a and R ^2b are each independently selected from H, C _1-6 alkyl, C _1-6 alkenyl, C _1-6 alkynyl, C _3-6 cycloalkyl, COO-C _1-6 alkyl, O-C _1-6 alkyl, CONR ^2b.1 R ^2b.2 and halogen;
R ^2b.1 is H, C _1-6 alkyl, C _0-4 alkyl-C _3-6 cycloalkyl, C _1-6 haloalkyl;
R ^2b.2 is H, C _1-6 alkyl, or
or R ^2b.1 and R ^2b.2 together are a C _3-6 alkylene group forming a heterocyclic ring with said nitrogen atom, optionally one carbon atom of said ring being replaced by an oxygen atom;
^R3 is H, _C1-6 alkyl;
X is an anion selected from the group consisting of chloride, bromide, iodide, sulfate, phosphate, methanesulfonate, nitrate, maleate, acetate, benzoate, citrate, salicylate, fumarate, tartrate, dibenzoyltartrate, oxalate, succinate, benzoate and p-toluenesulfonate;
j is 0, 0.5, 1, 1.5 or 2;
with a co-crystal former selected from the group consisting of orotic acid, hippuric acid, L-pyroglutamic acid, D-pyroglutamic acid, nicotinic acid, L-(+)-ascorbic acid, saccharin, piperazine, 3-hydroxy-2-naphthoic acid, mucic acid (galactaric acid), pamoic acid (embonic acid), stearic acid, cholic acid, deoxycholic acid, nicotinamide, isonicotinamide, succinamide, uracil, L-lysine, L-proline, D-valine, L-arginine, glycine;
The pharmaceutical composition of claim 1. 2. The pharmaceutical composition of claim 1, wherein the compound is a crystalline salt of the following formula:

The pharmaceutical composition of claim 1, wherein the compound comprises at least one cocrystal of a compound of the formula of claim 7 and a pharma- ceutical acceptable carrier. The pharmaceutical composition of claim 1, wherein the compound of formula 1 is administered in the form of an individual optical isomer, a mixture of individual enantiomers, a racemate, or an enantiomerically pure compound. The compound has as an active ingredient a compound of the following formula:

In the formula,
R ¹ is H, C _1-6 alkyl, C _0-4 alkyl-C _3-6 cycloalkyl, C _1-6 haloalkyl;
^R2 is H, _C1-6 alkyl;
X is an anion selected from the group consisting of chloride or 1/2 dibenzoyltartrate;
j is 1 or 2;
comprising one or more compounds,
A first diluent, a second diluent, a binder, a disintegrant and a lubricant,
The pharmaceutical composition of claim 1. The pharmaceutical composition of claim 11, further comprising an additional disintegrant. The pharmaceutical composition of claim 11 or 12, further comprising an additional fluidizing agent. The pharmaceutical composition according to any one of claims 11 to 13, wherein the diluent of the pharmaceutical composition further comprises cellulose powder, dibasic calcium phosphate, dibasic calcium phosphate dihydrate, erythritol, low-substituted hydroxypropylcellulose, mannitol, pregelatinized starch or xylitol. The pharmaceutical composition according to any one of claims 1 to 14, wherein the age-related neurodegeneration, age-related cognitive decline or age-related motor decline is selected from the group consisting of Alzheimer's disease, Parkinson's disease, frontotemporal dementia, Huntington's disease, amyotrophic lateral sclerosis, glaucoma, myotonic dystrophy, vascular dementia and progressive supranuclear palsy. 13. A pharmaceutical composition comprising a therapeutically effective amount of a compound of formula 1 according to claim 1 for use in a method for the systemic treatment of age-related neurodegeneration. A pharmaceutical composition comprising a therapeutically effective amount of a compound of formula 1 according to claim 1 for use in a method for the systemic treatment of age-related cognitive decline. A pharmaceutical composition comprising a therapeutically effective amount of a compound of formula 1 according to claim 1 for use in a method for the systemic treatment of age-related motor function decline.

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