Pharmaceuticals

15 pages, 2867 KiB

Open AccessArticle

A Comparative Survey of Anti-Melanoma and Anti-Inflammatory Potential of Usnic Acid Enantiomers—A Comprehensive In Vitro Approach

by Agnieszka Galanty, Paweł Zagrodzki, Joanna Gdula-Argasińska, Karolina Grabowska, Paulina Koczurkiewicz-Adamczyk, Dagmara Wróbel-Biedrawa, Irma Podolak, Elżbieta Pękala and Paweł Paśko

Pharmaceuticals 2021, 14(9), 945; https://doi.org/10.3390/ph14090945 - 21 Sep 2021

Cited by 15 | Viewed by 2887

Abstract

Usnic acid (UA) is a chiral lichen metabolite with an interesting pharmacological profile. The aim of this study was to compare the anti-melanoma effect of (+)-UA and (−)-UA in an in vitro model by studying their impact on the cells as well as [...] Read more.

Usnic acid (UA) is a chiral lichen metabolite with an interesting pharmacological profile. The aim of this study was to compare the anti-melanoma effect of (+)-UA and (−)-UA in an in vitro model by studying their impact on the cells as well as the processes associated with cancer progression. The effect of UA enantiomers on the viability, proliferation, and invasive potential of three melanoma cell lines (HTB140, A375, WM793) was evaluated. Their interaction with a chemotherapeutic drug—doxorubicin was assessed by isobolographic analysis. Anti-inflammatory and anti-tyrosinase properties of (+)-UA and (−)-UA were also examined. Both UA enantiomers dose- and time-dependently decreased the viability of all three melanoma cell lines. Their synergistic effect with doxorubicin was observed on A375 cells. (+)-Usnic acid at a sub-cytotoxic dose strongly inhibited melanoma cells migration. Both UA enantiomers decreased the release of pro-inflammatory mediators. The cytotoxic effect of (+)-UA and (−)-UA depends greatly on the melanoma cell type; however, the overall anti-melanoma potential is perspective. Our results indicate that the strategy of combining usnic acid enantiomers with cytostatic drugs may be an interesting option to consider in combating melanoma; however, further studies are required. Full article

(This article belongs to the Special Issue Antiradical, Chemopreventive and Antimicrobial Analysis of Bioactive Substances)

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Figure 1

Figure 1
Isobolograms for the combinations of usnic acid enantiomers with doxorubicin to HTB140 and A375 melanoma cells, after 24 h (A) and 48 h (B) of incubation. Points on x- and y-axes, connected by the line, represent median doses (Fa = 0.50 means IC50, Fa = 0.75 means IC75, Fa = 0.90 means IC90, Fa = 0.95 means IC95, Fa = 0.97 means IC97) of the substances given alone, whereas the dots represent appropriate combinations of both substances, respectively. Dots below the lines of the same colour indicate synergy, dots on the line of the same colour indicate additive effect, whereas the dots over the lines of the same colour indicate antagonism of the interaction. Full article ">Figure 2
The effect of (+)- and (−)-usnic acid on proliferation of HTB140 (A), A375 (B) and WM793 (C) human melanoma cells. Full article ">Figure 2 Cont.
The effect of (+)- and (−)-usnic acid on proliferation of HTB140 (A), A375 (B) and WM793 (C) human melanoma cells. Full article ">Figure 3
The effect of (+)- and (−)-usnic acid on migration ability of HTB140, A375 and WM793 human melanoma cells. Cells were treated or not (untreated) with 10 µg/mL of (+)-usnic acid (+UA) or (−)-usnic acid (−UA) for 24 h. Values are presented as the mean ± SD (standard deviation) of 10 randomly selected fields of view. Experiments were run twice in a blindfolded manner. Statistical analyses were carried out by using one-way ANOVA and Tukey post-hoc test. Significant differences (* p < 0.05 and ** p < 0.01) between usnic acid enantiomers were marked by upper black line. Full article ">Figure 4
The effect of (+)- and (−)-usnic acid on tumour necrosis factor-alpha (TNF-α), interleukin-6 (IL-6) and nitric oxide (NO) release in LPS-stimulated RAW 264.7 macrophages. RAW cells were pre-treated with 10 (UA10) and 25 (UA25) µg/mL of usnic acid enantiomers for 1 h, afterwards cells were incubated with (10 ng/mL) or without LPS (untreated) for the next 24 h. Dexamethasone (DEX) was used as a reference. Values are presented as the mean ± SD (standard deviation) of three independent experiments in triplicate. Statistical analyses were carried out by using one-way ANOVA and Tukey post-hoc test with * p < 0.05 and *** p < 0.001 against the LPS-stimulated cells. Full article ">Figure 5
The effect of (+)- and (−)-usnic acid on toll-like receptor 4 (TLR4), cytosolic phospholipase A2 (cPLA2), cyclooxygenase-1 (COX-1) and cyclooxygenase-2 (COX-2) release in LPS-stimulated RAW 264.7 macrophages. RAW cells were treated with 10 (UA10) and 25 (UA25) µg/mL of usnic acid enantiomers and 10 ng/mL of LPS for 24 h. Control cells were not treated with the tested compounds nor LPS (untreated). Densitometric evaluation of the bands TLR4, cPLA2, COX-1, COX-2 from Western blots in relation to beta-actin is shown. Values are presented as the mean ± SD (standard deviation) of three independent experiments in triplicate. Statistical analyses were carried out by using one-way ANOVA and Tukey post-hoc test with * p < 0.05, ** p < 0.01 and *** p < 0.001 against the LPS-stimulated cells. Significant differences (a: p < 0.01 and b: p < 0.001) between usnic acid enantiomers and doses used in the experiment were marked by upper black line. Full article ">

23 pages, 2031 KiB

Open AccessArticle

Less Polar Compounds and Targeted Antioxidant Potential (In Vitro and In Vivo) of Codium adhaerens C. Agardh 1822

by Sanja Radman, Ana-Marija Cikoš, Ivana Flanjak, Sanja Babić, Lara Čižmek, Drago Šubarić, Rozelindra Čož-Rakovac, Stela Jokić and Igor Jerković

Pharmaceuticals 2021, 14(9), 944; https://doi.org/10.3390/ph14090944 - 21 Sep 2021

Cited by 15 | Viewed by 2856

Abstract

Codium adhaerens from the Adriatic Sea (Croatia) was comprehensively investigated regarding less polar compounds for the first time. Although there are several phytochemical studies on C. adhaerens from other regions, this is the first report on volatile organic compounds (VOCs) from fresh (FrCa) [...] Read more.

Codium adhaerens from the Adriatic Sea (Croatia) was comprehensively investigated regarding less polar compounds for the first time. Although there are several phytochemical studies on C. adhaerens from other regions, this is the first report on volatile organic compounds (VOCs) from fresh (FrCa) and air-dried (DrCa) samples. The novelty is also related to its targeted antioxidant potential in vitro and in vivo. The main aims were to: (a) identify and compare VOCs of FrCa and DrCa obtained by headspace solid-phase microextraction (HS-SPME) and hydrodistillation (HD); (b) determine fatty acid (FA) composition of freeze-dried sample (FdCa); (c) determine the composition of less polar fractions of FdCa by high-performance liquid chromatography–high-resolution mass spectrometry with electrospray ionisation (UHPLC-ESI-HRMS); and (d) comprehensively evaluate the antioxidant activity of the fractions by four in vitro assays and in vivo zebrafish model (including embryotoxicity). Significant changes of VOCs were found after air drying. ω6 FAs were present in higher content than ω3 FAs indicating C. adhaerens as a good source of dietary polyunsaturated FAs. The results obtained in vivo correlate well with in vitro methods and both fractions exerted similar antioxidative responses which is in agreement with the high abundance of present biomolecules with known antioxidant properties (e.g., fucoxanthin, pheophytin a, and pheophorbide a). These results suggest that C. adhaerens might be a potent source of natural antioxidants that could be further used in the research of oxidative stress-related diseases. Full article

(This article belongs to the Special Issue Chemistry and Biomedical Potential of Marine Natural Products)

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Graphical abstract

Graphical abstract
Full article ">Figure 1
The VOCs from C. adhaerens sorted by structural groups extracted by HS-SPME. Full article ">Figure 2
The VOCs from C. adhaerens obtained by HD sorted by structural groups. Full article ">Figure 3
Total ion chromatogram (TIC) of the fractions (a) F3 and (b) F4. Full article ">Figure 4
Extracted ion chromatograms (XIC) of most abundant ions in the fractions (a) F3 and (b) F4 (zoomed). Full article ">Figure 5
Antioxidant activity of C. adhaerens less polar fractions (F3 and F4) obtained using 2,2-diphenyl-1-picryl-hydrazyl-hydrate (DPPH) and ferric reducing antioxidant power (FRAP) in vitro assays (mean ± SD; n = 3). An asterisk indicates a significant difference between F3 and F4 (*** p < 0.01; **** p < 0.001). Full article ">Figure 6
ROS scavenging potential of C. adhaerens fractions F3 and F4 in zebrafish. Left: Representative fluorescence images of H2O2 treated and the fractions co-treated zebrafish larvae. Right: Mean fluorescent intensity of DCF in the whole larvae calculated using Image J program. Sign # represents significant difference when compared with H2O2-treated group (# p < 0.05; ### p < 0.001). Each treatment was normalised relative to non-treated controls (100%). Full article ">

20 pages, 5101 KiB

Open AccessArticle

Development and Validation of a New Storage Procedure to Extend the In-Use Stability of Azacitidine in Pharmaceutical Formulations

by Antonella Iudicello, Filippo Genovese, Valentina Strusi, Massimo Dominici and Barbara Ruozi

Pharmaceuticals 2021, 14(9), 943; https://doi.org/10.3390/ph14090943 - 21 Sep 2021

Cited by 3 | Viewed by 4518

Abstract

Stability studies performed by the pharmaceutical industry are principally designed to fulfill licensing requirements. Thus, post-dilution or post-reconstitution stability data are frequently limited to 24 h only for bacteriological reasons, regardless of the true physicochemical stability which could, in many cases, be longer. [...] Read more.

Stability studies performed by the pharmaceutical industry are principally designed to fulfill licensing requirements. Thus, post-dilution or post-reconstitution stability data are frequently limited to 24 h only for bacteriological reasons, regardless of the true physicochemical stability which could, in many cases, be longer. In practice, the pharmacy-based centralized preparation may require preparation in advance for administration, for example, on weekends, holidays, or in general when pharmacies may be closed. We report an innovative strategy for storing resuspended solutions of azacitidine, a well-known chemotherapic agent, for which the manufacturer lists maximum stability of 22 h. By placing the syringe with the azacitidine reconstituted suspension between two refrigerant gel packs and storing it at 4 °C, we found that the concentration of azacitidine remained above 98% of the initial concentration for 48 h, and no change in color nor the physicochemical properties of the suspension were observed throughout the study period. The physicochemical and microbiological properties were evaluated by HPLC–UV and UHPLC-HRMS analysis, FTIR spectroscopy, pH determination, visual and subvisual examination, and sterility assay. The HPLC-UV method used for evaluating the chemical stability of azacitidine was validated according to ICH. Precise control of storage temperature was obtained by a digital data logger. Our study indicates that by changing the storage procedure of azacitidine reconstituted suspension, the usage window of the drug can be significantly extended to a time frame that better copes with its use in the clinical environment. Full article

(This article belongs to the Section Pharmaceutical Technology)

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11 pages, 1235 KiB

Open AccessArticle

The Synthetic Curcumin Analog HO-3867 Rescues Suppression of PLAC1 Expression in Ovarian Cancer Cells

by Eric J. Devor, Brandon M. Schickling, Jace R. Lapierre, David P. Bender, Jesus Gonzalez-Bosquet and Kimberly K. Leslie

Pharmaceuticals 2021, 14(9), 942; https://doi.org/10.3390/ph14090942 - 21 Sep 2021

Cited by 7 | Viewed by 2818

Abstract

Elevated expression of placenta-specific protein 1 (PLAC1) is associated with the increased proliferation and invasiveness of a variety of human cancers, including ovarian cancer. Recent studies have shown that the tumor suppressor p53 directly suppresses PLAC1 transcription. However, mutations in p53 lead to [...] Read more.

Elevated expression of placenta-specific protein 1 (PLAC1) is associated with the increased proliferation and invasiveness of a variety of human cancers, including ovarian cancer. Recent studies have shown that the tumor suppressor p53 directly suppresses PLAC1 transcription. However, mutations in p53 lead to the loss of PLAC1 transcriptional suppression. Small molecules that structurally convert mutant p53 proteins to wild-type conformations are emerging. Our objective was to determine whether the restoration of the wild-type function of mutated p53 could rescue PLAC1 transcriptional suppression in tumors harboring certain TP53 mutations. Ovarian cancer cells OVCAR3 and ES-2, both harboring TP53 missense mutations, were treated with the p53 reactivator HO-3867. Treatment with HO-3867 successfully rescued PLAC1 transcriptional suppression. In addition, cell proliferation was inhibited and cell death through apoptosis was increased in both cell lines. We conclude that the use of HO-3867 as an adjuvant to conventional therapeutics in ovarian cancers harboring TP53 missense mutations could improve patient outcomes. Validation of this conclusion must, however, come from an appropriately designed clinical trial. Full article

(This article belongs to the Section Pharmacology)

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Total PLAC1 and PLAC1P1-specific expression in OVCAR3 ovarian cancer cells treated with 2 µM and 5 µM HO-3867 for 24 h and 48 h. Full article ">Figure 2
Total PLAC1 and PLAC1P1-specific expression in ES-2 ovarian cancer cells treated with 2 µM and 5 µM HO-3867 for 24 h. ES-2 cells exposed to 2 µM HO-3867 for 48 h did not yield RNA of sufficient quality to perform qPCR, and ES-2 cells exposed to 5 µM HO-3867 for 48 h did not survive. The 24 h results for OVCAR3 cells are shown on the left for comparison. Full article ">Figure 3
Proliferation and apoptosis in OVCAR3 (A) and ES-2 (B) cells. Cells were treated with increasing amounts of HO-3867, as shown. Proliferation was measured by phase contrast, and apoptosis was assessed by caspase 3/7 activation. Cells were monitored hourly for 48 h using the Incucyte S3 live-cell imaging platform. Area under the curve (AUC) values were calculated automatically by the Incucyte Software and analyzed via ANOVA. Significance values are indicated as ** p < 0.01; *** p < 0.001; **** p < 0.0001. Full article ">Figure 4
A model of the effect of HO-3867 on p53 binding in the P1 promoter of PLAC1. In the top panel (A), wild-type p53 freely binds to its binding site, sterically hindering access of the P1 transcription factors RXRα and LXRβ to their binding sites. This has the effect of suppressing PLAC1 transcription. However, when p53 is mutated, as in the middle panel (B), p53 is unable to bind to its binding site, thus freeing the P1 transcription factors RXRα and LXRβ to bind to theirs and promoting PLAC1 transcription. When exposed to HO-3867, as in the lower panel (C), certain p53 mutants are reactivated and will bind to the p53 binding site, once again sterically hindering access of the P1 transcription factors RXRα and LXRβ to their binding sites and hindering PLAC1 transcription. TSS is the transcription start site. Full article ">

20 pages, 1619 KiB

Open AccessArticle

Chicory Extracts and Sesquiterpene Lactones Show Potent Activity against Bacterial and Fungal Pathogens

by Suvi T. Häkkinen, Marina Soković, Liisa Nohynek, Ana Ćirić, Marija Ivanov, Dejan Stojković, Irina Tsitko, Melanie Matos, João P. Baixinho, Viktoriya Ivasiv, Naiara Fernández, Claudia Nunes dos Santos and Kirsi-Marja Oksman-Caldentey

Pharmaceuticals 2021, 14(9), 941; https://doi.org/10.3390/ph14090941 - 20 Sep 2021

Cited by 29 | Viewed by 6326

Abstract

Chicory (Cichorium intybus L.) is an important industrial crop cultivated mainly to extract the dietary fiber inulin. However, chicory also contains bioactive compounds such as sesquiterpene lactones and certain polyphenols, which are currently discarded as waste. Plants are an important source of [...] Read more.

Chicory (Cichorium intybus L.) is an important industrial crop cultivated mainly to extract the dietary fiber inulin. However, chicory also contains bioactive compounds such as sesquiterpene lactones and certain polyphenols, which are currently discarded as waste. Plants are an important source of active pharmaceutical ingredients, including novel antimicrobials that are urgently needed due to the global spread of drug-resistant bacteria and fungi. Here, we tested different extracts of chicory for a range of bioactivities, including antimicrobial, antifungal and cytotoxicity assays. Antibacterial and antifungal activities were generally more potent in ethyl acetate extracts compared to water extracts, whereas supercritical fluid extracts showed the broadest range of bioactivities in our assays. Remarkably, the chicory supercritical fluid extract and a purified fraction thereof inhibited both methicillin-resistant Staphylococcus aureus (MRSA) and ampicillin-resistant Pseudomonas aeruginosa IBRS P001. Chicory extracts also showed higher antibiofilm activity against the yeast Candida albicans than standard sesquiterpene lactone compounds. The cytotoxicity of the extracts was generally low. Our results may thus lead to the development of novel antibacterial and antifungal preparations that are both effective and safe for human use. Full article

(This article belongs to the Special Issue Natural Pharmacons: Biologically Active Plant Based Pharmaceuticals)

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Growth inhibition by Ci_SFEopt and Ci_SFEpur at a concentration of 5 mg/mL against (A) S. aureus, and (B) S. aureus MRSA. Green, Ci_SFEpur; orange, Ci_SFEopt, black, microbial control. Full article ">Figure 2
Bioautographic assay of Ci_SFEopt and Ci_SFEpur developed in 85:15 (v/v) dichloromethane:methanol as the solvent. (A) Antibacterial activity against Staphylococcus aureus (oral isolate), S. aureus ATCC (left); and Pseudomonas aeruginosa (right). (B) Antifungal activity against Candida albicans (left) and C. krusei (right). Spots on different Rf values: 1. Rf = 0.18 (activity only against S. aureus ATCC); 2. Rf = 0.20 (activity only against P. aeruginosa); 3. Rf = 0.65 (activity against S. aureus ATCC and C. albicans); 4. Rf = 0.71 (activity against all tested bacteria and fungi); 5. Rf = 0.76 (activity against all tested bacteria and fungi, except C. krusei); 6. Rf = 0.82 (activity against all tested microorganisms, except P. aeruginosa and C. krusei); 7. Rf = 0.88 (activity against all tested bacteria and fungi, except C. krusei). Full article ">Figure 3
Percentage inhibition of biofilm formation in (A) C. albicans 475/15 and (B) P. aeruginosa IBRS P001 after treatment with chicory extracts and pure compounds. Results are presented as a mean (± stdev) of triplicates. Full article ">Figure 4
Modes of antibiofilm activity against P. aeruginosa IBRS P001 expressed as percentage inhibition compared to the untreated control. (A) Inhibition of biofilm cell viability determined by MTT assay. (B) Inhibition of biofilm exopolysaccharide content. (C) Inhibition of biofilm eDNA. (D) Inhibition of biofilm CFU formed on urinary catheters. Results are expressed as mean (±stdev) of triplicates. Full article ">Figure 5
Cytotoxic profile of chicory extracts in Caco-2 cells after incubation for 4 h. (A) Wit_EtOAc. (B) Wit_H2O. (C) Ci_EtOAc. (D) Ci_SFEopt. (E) Ci_SFEpur. Results are presented as mean (±stdev) of three independent experiments. Full article ">

16 pages, 6333 KiB

Open AccessArticle

Antioxidant and Anti-Inflammatory Potential of Thymoquinone and Lycopene Mitigate the Chlorpyrifos-Induced Toxic Neuropathy

by Mohamed Aboubakr, Said M. Elshafae, Ehab Y. Abdelhiee, Sabreen E. Fadl, Ahmed Soliman, Afaf Abdelkader, Mohamed M. Abdel-Daim, Khaled A. Bayoumi, Roua S. Baty, Enas Elgendy, Amira Elalfy, Bodour Baioumy, Samah F. Ibrahim and Ahmed Abdeen

Pharmaceuticals 2021, 14(9), 940; https://doi.org/10.3390/ph14090940 - 20 Sep 2021

Cited by 51 | Viewed by 4154

Abstract

CPF (chlorpyrifos) is an organophosphate pesticide used in agricultural and veterinary applications. Our experiment aimed to explore the effects of thymoquinone (TQ) and/or lycopene (LP) against CPF-induced neurotoxicity. Wistar rats were categorized into seven groups: first group served as a control (corn oil [...] Read more.

CPF (chlorpyrifos) is an organophosphate pesticide used in agricultural and veterinary applications. Our experiment aimed to explore the effects of thymoquinone (TQ) and/or lycopene (LP) against CPF-induced neurotoxicity. Wistar rats were categorized into seven groups: first group served as a control (corn oil only); second group, TQ (10 mg/kg); third group, LP (10 mg/kg); fourth group, CPF (10 mg/kg) and deemed as CPF toxic control; fifth group, TQ + CPF; sixth group, (LP + CPF); and seventh group, (TQ + LP + CPF). CPF intoxication inhibited acetylcholinesterase (AchE), decreased glutathione (GSH) content, and increased levels of malondialdehyde (MDA), an oxidative stress biomarker. Furthermore, CPF impaired the activity of antioxidant enzymes including superoxide dismutase (SOD) and catalase (CAT) along with enhancement of the level of inflammatory mediators such as tumor necrosis factor-α (TNF-α), interleukin (IL)-6, and IL-1β. CPF evoked apoptosis in brain tissue. TQ or LP treatment of CPF-intoxicated rats greatly improved AchE activity, oxidative state, inflammatory responses, and cell death. Co-administration of TQ and LP showed better restoration than their sole treatment. In conclusion, TQ or LP supplementation may alleviate CPF-induced neuronal injury, most likely due to TQ or LPs’ antioxidant, anti-inflammatory, and anti-apoptotic effects. Full article

(This article belongs to the Special Issue Natural Bioactive Compounds and Neurological Disorders: Exploiting Resources from Nature)

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Figure 1
Dot plot of AchE activity and inflammatory cytokines after treatment with CPF, TQ, and LP. AchE, acetylcholinesterase; CPF, chlorpyrifos; IL-6, interleukin-6; IL-1β, interleukin-1β; LP, lycopene; TNF-α, tumor necrosis factor-α; TQ, thymoquinone. Values proffered as the mean ± SE (n = 7). ** p ≤ 0.01 and *** p ≤ 0.001. Full article ">Figure 2
Dot plot of oxidative/antioxidative status after treatment with CPF, TQ, and LP. CAT, catalase; CPF, chlorpyrifos; GSH, reduced glutathione; LP, lycopene; MDA, malondialdehyde; SOD, superoxide dismutase; TQ, thymoquinone. Values are proffered as the mean ± SE (n = 7). *** p ≤ 0.001. Full article ">Figure 3
Histopathology of the cerebrum in control and CPF, TQ, and LP treated groups (H&E stain). Apparently, normal neurons were observed in control (A), TQ (B) and LP (C) treated rats with null to minimal apoptotic neurons. (D–F) Cerebral cortex of CPF-treated rats showing neuronal degeneration (tigrolysis; black arrow) (D), central chromatolysis (red arrow) (E) and neuropile vacuolation (black arrowhead) (F). A low number of shrunken apoptotic neurons (yellow arrows) was observed in TQ (G), LP (H) and combination (I) groups co-treated with CPF. Full article ">Figure 4
Histopathology of the cerebellum in control and CPF, TQ, and LP treated groups (H&E stain). Nearly normal Purkinje neurons were observed in control (A), TQ- (B) and LP- (C) treated rats with null to mild degeneration of few neurons. (D–F) Cerebellum of CPF treated rats showing Purkinje cell apoptosis (black arrows) (D), loss (black arrowheads) (E) and disorganization with molecular cell layer (F). Marked decreases in degenerated/apoptotic Purkinje cells were observed in TQ (G), LP (H) and combination (I) groups co-treated with CPF. Full article ">Figure 5
Immunohistochemical staining of cleaved caspase 3 in cerebral sections. (A–C) Almost all the neurons in control (A), TQ- (B) and LP- (C) treated groups had no cleaved caspase 3 staining. (D,E) Many neurons displayed strong staining for cleaved caspase 3 in CPF intoxicated rats. (F–H) Few neurons were weakly to moderately positive for cleaved caspase 3 in CPF intoxicated rats co-treated with TQ (F), LP (G) or both (H). (I) Bar graph represents the relative cleaved caspase 3 positivity in different groups in contrast to CPF intoxicated group. Substantial differences are exhibited as: * p ≤ 0.05, ** p ≤ 0.01. Full article ">Figure 6
Immunohistochemical staining of cleaved caspase 3 in the cerebellum. All the neurons in molecular, Purkinje and granular cell layers displayed no caspase 3 staining in control (A), TQ- (B) and LP- (C) treated rats. (D,E) Most Purkinje cells and many neurons in granular and molecular cell layers showed strong caspase 3 staining in CPF intoxicated rats. (F) Most neurons in all the cerebellar layers had no caspase 3 staining in TQ + CPF treated rats. (G) Certain Purkinje cells were vacuolated and moderately stained with cleaved caspase 3 in LP + CPF treated rats. (H) All the neurons in the cerebellar sections were devoid of caspase 3 staining in TQ + LP + CPF treated rats. (I) Bar graph represents the relative cleaved caspase 3 positivity in cerebellar sections of different groups compared to CPF intoxicated group. Substantial differences are exhibited as: * p ≤ 0.05, ** p ≤ 0.01, and *** p ≤ 0.001. Full article ">Figure 7
Schematic diagram summarizes the mechanistic insights behinds the protective potential of TQ and LP during CPF-induced neurotoxicity. AchE, acetylcholinesterase; CAT, catalase; CPF, chlorpyrifos; GSH, reduced glutathione; IL- 6, interleukin-6; IL-1β, interleukin-1β; LP, lycopene; MDA, malondialdehyde; ROS, reactive oxygen species; SOD, superoxide dismutase; TNF-α, tumor necrosis factor-α; TQ, thymoquinone. Full article ">

20 pages, 5178 KiB

Open AccessArticle

Preparation of Soluble Complex of Curcumin for the Potential Antagonistic Effects on Human Colorectal Adenocarcinoma Cells

by Jamal Moideen Muthu Mohamed, Ali Alqahtani, Barkat A. Khan, Adel Al Fatease, Taha Alqahtani, Krishnaraju Venkatesan, Fazil Ahmad, Bashar I. Alzghoul and Ali Alamri

Pharmaceuticals 2021, 14(9), 939; https://doi.org/10.3390/ph14090939 - 19 Sep 2021

Cited by 19 | Viewed by 3035

Abstract

This study was designed to investigate the effects of curcumin (CMN) soluble complex (SC) prepared by melt casting (HM) and hot-melt extrusion (HME) technology. Phase solubility (PS) study, in silico molecular modeling, aqueous solubility, drug release, and physicochemical investigation including a novel dyeing [...] Read more.

This study was designed to investigate the effects of curcumin (CMN) soluble complex (SC) prepared by melt casting (HM) and hot-melt extrusion (HME) technology. Phase solubility (PS) study, in silico molecular modeling, aqueous solubility, drug release, and physicochemical investigation including a novel dyeing test was performed to obtain an optimized complex by a central composite design (CCD). The results show that the HME-SC produces better improvements towards solubility (0.852 ± 0.02), dissolution (91.87 ± 0.21% at 30 min), with an ideal stability constant (309 and 377 M⁻¹ at 25 and 37 °C, respectively) and exhibits A_L type of isotherm indicating 1:1 stoichiometry. Intermolecular hydrogen bonding involves the formation of SC, which does not undergo any chemical modification, followed by the complete conversion of the amorphous form which was identified by XRD. The in vitro cytotoxicity showed that IC₅₀ was achieved in the SW480 (72 µM.mL⁻¹) and Caco-2 (40 µM.mL⁻¹) cells while that of pure CMN ranged from 146 to 116 µM/mL⁻¹. Apoptosis studies showed that cell death is primarily due to apoptosis, with a low rate of necrosis. In vivo toxicity, confirmed by the zebrafish model, exhibited the safety of the HME-SC. In conclusion, the HME-SC potentially enhances the solubility and cytotoxicity to the treatment of colorectal cancer (CRC). Full article

(This article belongs to the Topic Compounds with Medicinal Value)

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14 pages, 5534 KiB

Open AccessArticle

Asymmetric Total Syntheses of Both Enantiomers of Plymuthipyranone B and Its Unnatural Analogues: Evaluation of anti-MRSA Activity and Its Chiral Discrimination

by Mizuki Moriyama, Xiaoxi Liu, Yuki Enoki, Kazuaki Matsumoto and Yoo Tanabe

Pharmaceuticals 2021, 14(9), 938; https://doi.org/10.3390/ph14090938 - 19 Sep 2021

Cited by 2 | Viewed by 2536

Abstract

Chiral total syntheses of both enantiomers of the anti-MRSA active plymuthipyranone B and all of the both enantiomers of three unnatural and synthetic analogues were performed. These two pairs of four chiral compounds are composed of the same 3-acyl-5,6-dihydro-2H-pyran-2-one structure. [...] Read more.

Chiral total syntheses of both enantiomers of the anti-MRSA active plymuthipyranone B and all of the both enantiomers of three unnatural and synthetic analogues were performed. These two pairs of four chiral compounds are composed of the same 3-acyl-5,6-dihydro-2H-pyran-2-one structure. The starting synthetic step utilized a privileged asymmetric Mukaiyama aldol addition using Ti(OiPr)₄/(S)-BINOL or Ti(OiPr)₄/(R)-BINOL catalysis to afford the corresponding (R)- and (S)-δ-hydroxy-β-ketoesters, respectively, with highly enantiomeric excess (>98%). Conventional lactone formation and successive EDCI-mediated C-acylation produced the desired products, (R)- and (S)-plymuthipyranones B and three (R)- and (S)- synthetic analogues, with an overall yield of 42–56% with a highly enantiomeric excess (95–99%). A bioassay of the anti-MRSA activity against ATCC 43300 and 33591 revealed that (i) the MICs of the synthetic analogues against ATCC 43300 and ATCC 33591 were between 2 and 16 and 4 and 16 μg/mL, respectively, and those of vancomycin (reference) were 1 μg/mL. (ii) The natural (S)-plymuthipyranone B exhibited significantly higher activity than the unnatural (R)-antipode against both AACCs. (iii) The natural (R)-plymuthipyranone B and (R)-undecyl synthetic analogue at the C6 position exhibited the highest activity. The present work is the first investigation of the SAR between chiral R and S forms of this chemical class. Full article

(This article belongs to the Special Issue Chirality in Drug Discovery)

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19 pages, 6152 KiB

Open AccessArticle

High-Throughput Screening and Molecular Dynamics Simulation of Natural Product-like Compounds against Alzheimer’s Disease through Multitarget Approach

by Danish Iqbal, Md Tabish Rehman, Abdulaziz Bin Dukhyil, Syed Mohd Danish Rizvi, Mohamed F. Al Ajmi, Bader Mohammed Alshehri, Saeed Banawas, M. Salman Khan, Wael Alturaiki and Mohammed Alsaweed

Pharmaceuticals 2021, 14(9), 937; https://doi.org/10.3390/ph14090937 - 18 Sep 2021

Cited by 36 | Viewed by 4410

Abstract

Alzheimer’s disease (AD) is a progressive neurological disorder that affects 50 million people. Despite this, only two classes of medication have been approved by the FDA. Therefore, we have planned to develop therapeutics by multitarget approach. We have explored the library of 2029 [...] Read more.

Alzheimer’s disease (AD) is a progressive neurological disorder that affects 50 million people. Despite this, only two classes of medication have been approved by the FDA. Therefore, we have planned to develop therapeutics by multitarget approach. We have explored the library of 2029 natural product-like compounds for their multi-targeting potential against AD by inhibiting AChE, BChE (cholinergic pathway) MAO-A, and MOA-B (oxidative stress pathway) through in silico high-throughput screening and molecular dynamics simulation. Based on the binding energy of these target enzymes, approximately 189 compounds exhibited a score of less than −10 kcal/mol against all targets. However, none of the control inhibitors exhibited a binding affinity of less than −10 kcal/mol. Among these, the top 10 hits of compounds against all four targets were selected for ADME-T analysis. As a result, only F0850-4777 exhibited an acceptable range of physicochemical properties, drug-likeness, pharmacokinetics, and suitability for BBB permeation with high GI-A and non-toxic effects. The molecular dynamics study confirmed that F0850-4777 remained inside the binding cavity of targets in a stable conformation throughout the simulation and Prime-MM/GBSA study revealed that van der Waals’ energy (ΔG_vdW) and non-polar solvation or lipophilic energy (ΔG_{Sol_Lipo}) contribute favorably towards the formation of a stable protein–ligand complex. Thus, F0850-4777 could be a potential candidate against multiple targets of two pathophysiological pathways of AD and opens the doors for further confirmation through in vitro and in vivo systems. Full article

(This article belongs to the Special Issue In Silico Approaches in Drug Design)

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26 pages, 11394 KiB

Open AccessReview

The Endocannabinoid System and Cannabidiol: Past, Present, and Prospective for Cardiovascular Diseases

by Martina Rabino, Sara Mallia, Elisa Castiglioni, Davide Rovina, Giulio Pompilio and Aoife Gowran

Pharmaceuticals 2021, 14(9), 936; https://doi.org/10.3390/ph14090936 - 17 Sep 2021

Cited by 13 | Viewed by 5889

Abstract

In the past, cannabis was commonly associated with mysticism and illegality. Fortunately, in recent years perspectives and discourses have changed. More prominence has been given to the rigorous scientific effort that led to the discovery of cannabis’ many physiological actions and endogenous signalling [...] Read more.

In the past, cannabis was commonly associated with mysticism and illegality. Fortunately, in recent years perspectives and discourses have changed. More prominence has been given to the rigorous scientific effort that led to the discovery of cannabis’ many physiological actions and endogenous signalling mechanisms. The endocannabinoid system is a complex and heterogeneous pro-homeostatic network comprising different receptors with several endogenous ligands, numerous metabolic enzymes and regulatory proteins. Therefore, it is not surprising that alterations and dysfunctions of the endocannabinoid system are observed in almost every category of disease. Such high degree of pathophysiological involvement suggests the endocannabinoid system is a promising therapeutic target and prompted the translation of resurgent scientific findings into clinical therapies. Shifting attitudes toward cannabis also raised other matters such as increased patient awareness, prescription requests, self-medication, recreational use, recognition of new knowledge gaps, renewed scientific activity, and seemingly exponential growth of the cannabis industry. This review, following a general overview of cannabis and the endocannabinoid system, assiduously describes its role within the context of cardiovascular diseases, paying particular attention to the Janus influence that endocannabinoid system modulators can have on the cardiovascular system. Full article

(This article belongs to the Special Issue Cannabidiol: Advances in Therapeutic Applications and Future Perspectives)

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Graphical abstract

Graphical abstract
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A schematic summary of the main endocannabinoid metabolic pathways. The biosynthesis of the two most researched eCBs, arachidonoyl ethanolamide (AEA) and 2-arachidonoylglycerol (2-AG), starts with the catabolism of phospholipid precursors via the action of two respective enzymes, namely, NAPE phospholipase D (NAPE-PLD) and the alpha or beta isoform of diacylglycerol lipase (DAGL). Despite AEA and 2-AG both possessing an AA chain structural motif, their catabolic pathways are different, and thus are respectively catabolized by fatty acid amide hydrolase (FAAH) and monoacylglycerol lipase (MAGL) to form arachidonic acid (AA) and ethanolamine following AEA degradation, or AA and glycerol in the case of 2-AG breakdown. Both products are rapidly re-incorporated into membrane phospholipids. Blue arrows signify enzymatic reactions; black arrow indicates the passage of ions through the ion channel; ↑ i[Ca2+] represents elevated intracellular calcium. Full article ">Figure 2
Contribution of the endocannabinoid system (ECS) to cardiovascular pathophysiology and cardioprotection. Schematic representation of the known interaction between the ECS and the bone marrow niche, immune cells, and other cells or cellular cargo that mediate pathophysiological mechanisms in CVDs, e.g., mesenchymal stem cells (MSCs), haematopoietic stem cells (HSCs), adipocytes, and extracellular vesicles (EVs). Comprehensive understanding of the dynamic alterations in the ECS during damage to the myocardium represents a crucial point to establish novel cannabinoid-based therapeutic approaches. For instance, during injury or pathological conditions the ECS becomes dysfunctional and loses specificity of activity that can further drive pathological events. Alterations including increased CB1 expression and anandamide (AEA) during these states activate various molecular signalling such as inflammation and fibrotic signalling. Thus targeting the ECS by inhibiting or enhancing relevant components, e.g., receptors, eCB metabolism etc., could be of potential benefit to improve current and future treatment strategies aimed at cardioprotection. Full article ">

18 pages, 1695 KiB

Open AccessFeature PaperArticle

The Metabolomic Approach Reveals the Alteration in Human Serum and Cerebrospinal Fluid Composition in Parkinson’s Disease Patients

by Szymon Plewa, Karolina Poplawska-Domaszewicz, Jolanta Florczak-Wyspianska, Agnieszka Klupczynska-Gabryszak, Bartosz Sokol, Wojciech Miltyk, Roman Jankowski, Wojciech Kozubski, Zenon J. Kokot and Jan Matysiak

Pharmaceuticals 2021, 14(9), 935; https://doi.org/10.3390/ph14090935 - 17 Sep 2021

Cited by 20 | Viewed by 3861

Abstract

Parkinson’s disease (PD) is a major public health problem. Since currently there are no reliable diagnostic tools to reveal the early steps of PD, new methods should be developed, including those searching the variations in human metabolome. Alterations in human metabolites could help [...] Read more.

Parkinson’s disease (PD) is a major public health problem. Since currently there are no reliable diagnostic tools to reveal the early steps of PD, new methods should be developed, including those searching the variations in human metabolome. Alterations in human metabolites could help to establish an earlier and more accurate diagnosis. The presented research shows a targeted metabolomics study of both of the serum and CSF from PD patients, atypical parkinsonian disorders (APDs) patients, and the control. The use of the LC-MS/MS system enabled to quantitate 144 analytes in the serum and 51 in the CSF. This information about the concentration enabled for selection of the metabolites useful for differentiation between the studied group of patients, which should be further evaluated as candidates for markers of screening and differential diagnosis of PD and APDs. Among them, the four compounds observed to be altered in both the serum and CSF seem to be the most important: tyrosine, putrescine, trans-4-hydroxyproline, and total dimethylarginine. Furthermore, we indicated the metabolic pathways potentially related to neurodegeneration processes. Our studies present evidence that the proline metabolism might be related to neurodegeneration processes underlying PD and APDs. Further studies on the proposed metabolites and founded metabolic pathways may significantly contribute to understanding the molecular background of PD and improving the diagnostics and treatment in the future. Full article

(This article belongs to the Section Pharmacology)

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Figure 1
Box plots showing the distributions of the serum metabolites—amino acids and biogenic amines—found as altered between the studied groups: C—control group, PD—Parkinson’s disease, APDs—atypical parkinsonian disorders. Boxes extend from 25th to 75th percentiles; whiskers denote a range of measured concentrations; line and plus represent the median concentration and the mean of concentration in a group, respectively; dots, squares, and triangles show the measured concentrations in each group. * p ≤ 0.05, ** p ≤ 0.01, and *** p ≤ 0.001; ns—not significant. Full article ">Figure 2
Box plots showing the distributions of the serum metabolites—in the class of lipids—found as altered between the studied groups: C—control group, PD—Parkinson’s disease, APDs—atypical parkinsonian disorders. Boxes extend from 25th to 75th percentiles; whiskers denote a range of measured concentrations; line and plus represent the median concentration and the mean of concentration in a group, respectively; dots, squares, and triangles show the measured concentrations in each group. * p ≤ 0.05, ** p ≤ 0.01; ns—not significant. Full article ">Figure 3
Box plots showing the distributions of the CSF metabolites altered between the studied groups: C—control group, PD—Parkinson’s disease, APDs—atypical parkinsonian disorders. Boxes extend from 25th to 75th percentiles; whiskers denote a range of measured concentrations; line and plus represent the median concentration and the mean of concentration in a group, respectively; dots, squares, and triangles show the measured concentrations in each group. * p ≤ 0.05, ** p ≤ 0.01, and *** p ≤ 0.001; ns—not significant. Full article ">Figure 4
Metabolic pathways found to be potentially related to neurodegenerative processes, based on the identified potential biomarkers (left); the scheme of the arginine and proline metabolism pathway with significantly altered metabolites (red) between the studied groups of patients (right). Full article ">Figure 5
The hypothetical interlinks between the intermediates of proline–hydroxyproline conversion. Full article ">

16 pages, 2068 KiB

Open AccessReview

Manufacturing Bacteriophages (Part 1 of 2): Cell Line Development, Upstream, and Downstream Considerations

by Tayfun Tanir, Marvin Orellana, Aster Escalante, Carolina Moraes de Souza and Michael S. Koeris

Pharmaceuticals 2021, 14(9), 934; https://doi.org/10.3390/ph14090934 - 17 Sep 2021

Cited by 9 | Viewed by 5626

Abstract

Within this first part of the two-part series on phage manufacturing, we will give an overview of the process leading to bacteriophages as a drug substance, before covering the formulation into a drug product in the second part. The principal goal is to [...] Read more.

Within this first part of the two-part series on phage manufacturing, we will give an overview of the process leading to bacteriophages as a drug substance, before covering the formulation into a drug product in the second part. The principal goal is to provide the reader with a comprehensive framework of the challenges and opportunities that present themselves when developing manufacturing processes for bacteriophage-based products. We will examine cell line development for manufacture, upstream and downstream processes, while also covering the additional opportunities that engineered bacteriophages present. Full article

(This article belongs to the Special Issue Bacteriophages as Therapeutic Delivery Vehicles)

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Figure 1
A simplified process flow diagram for phage production in batch operation mode. Full article ">Figure 2
A simplified process flow diagram of the proposed SCF/SCI system where SCF and SCI are fermenters for bacteria and phage amplification, respectively. All the control systems are excluded to provide a simple depiction of the system. Adapted from [<a href="#B69-pharmaceuticals-14-00934" class="html-bibr">69</a>]. Full article ">Figure 3
A simplified PFD of a continuous phage manufacturing system. As before, all the control systems are excluded to provide a simple depiction of the system. Full article ">Figure 4
It is important to realize that crude lysate starts becoming DS and needs to be treated as such. The process developer should consider the critical quality attributes that are measured along the downstream separation cascade. Full article ">Figure 5
Generalized process flow for phage purification. From harvest, pretreatment is required to collect the phages from the lysate. Crude lysate can be processed using a combination of centrifugation, microfiltration, and precipitation. Once the lysate has been concentrated and the host cell debris reduced, more sensitive procedures may be implemented, such as chromatography and specific types of filtrations. Polishing procedures may be required to further isolate and purify the phage before moving on to fill-finish. Full article ">

18 pages, 1700 KiB

Open AccessReview

Neuropsychiatric Disorders and COVID-19: What We Know So Far

by Fernanda Majolo, Guilherme Liberato da Silva, Lucas Vieira, Cetin Anli, Luís Fernando Saraiva Macedo Timmers, Stefan Laufer and Márcia Inês Goettert

Pharmaceuticals 2021, 14(9), 933; https://doi.org/10.3390/ph14090933 - 17 Sep 2021

Cited by 15 | Viewed by 4350

Abstract

SARS-CoV-2 (Severe Acute Respiratory Syndrome Coronavirus-2) affects the central nervous system (CNS), which is shown in a significant number of patients with neurological events. In this study, an updated literature review was carried out regarding neurological disorders in COVID-19. Neurological symptoms are more [...] Read more.

SARS-CoV-2 (Severe Acute Respiratory Syndrome Coronavirus-2) affects the central nervous system (CNS), which is shown in a significant number of patients with neurological events. In this study, an updated literature review was carried out regarding neurological disorders in COVID-19. Neurological symptoms are more common in patients with severe infection according to their respiratory status and divided into three categories: (1) CNS manifestations; (2) cranial and peripheral nervous system manifestations; and (3) skeletal muscle injury manifestations. Patients with pre-existing cerebrovascular disease are at a higher risk of admission to the intensive care unit (ICU) and mortality. The neurological manifestations associated with COVID-19 are of great importance, but when life-threatening abnormal vital signs occur in severely ill COVID-19 patients, neurological problems are usually not considered. It is crucial to search for new treatments for brain damage, as well as for alternative therapies that recover the damaged brain and reduce the inflammatory response and its consequences for other organs. In addition, there is a need to diagnose these manifestations as early as possible to limit long-term consequences. Therefore, much research is needed to explain the involvement of SARS-CoV-2 causing these neurological symptoms because scientists know zero about it. Full article

(This article belongs to the Special Issue COVID-19 in Pharmaceuticals)

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Figure 1
Pathways through which viruses can infect the peripheral nervous system (PNS) or central nervous system (CNS): (1) nerve endings were accessed in the tissues by the infection using axonal transport machinery to gain access to the CNS; or (2) by the infected cells in a circulatory system, which carries the infection through the blood–brain barrier into the CNS. Adapted from Yachou et al., 2020 [<a href="#B98-pharmaceuticals-14-00933" class="html-bibr">98</a>]. Full article ">Figure 2
SARS-CoV-2 infecting cells in lung tissue through the interaction with angiotensin-converting enzyme 2 (ACE2), promoting the generation of pro-inflammatory cytokines (cytokine storm). This process attracts “defense” cells such as macrophages, monocytes, and T cells to the site of infection, triggering the inflammation process. The result of this inflammation is lung tissue damage. Adapted from Tay et al., 2020 [<a href="#B102-pharmaceuticals-14-00933" class="html-bibr">102</a>]. Full article ">Figure 3
Microglia and astrocytes are activated by ATP expressed during the release of pro-inflammatory cytokines. NMDA receptors expressed by Glutamate activation allow the Ca2+-dependent exocytosis of ATP, thus releasing more Glutamate. A massive release of these neurotransmitters increases cell death and excitotoxicity, since postsynaptic neuron increased the Ca2+-calmodulin (CaM) complex formation and consequent nNOS activation. Neurotoxicity is mediated by NO production, which interacts with the iron–sulfur centers in the mitochondrial electron transport chain, and produces reactive oxygen species (ROS), impairing cellular energy production. Still, a reaction of O2− (superoxide ion) and NO forms peroxynitrite (ONOO−) and peroxynitrous acid (ONOOH). Such formation leads to oxidative stress, which includes DNA damage, lipid peroxidation, tyrosine nitration, and excess S-nitrosylation, causing neuronal impairment and/or death. The activation of caspase-1 is mediated by the NLRP3 inflammasome by the cleavage of pro-IL-1β and pro-IL-18 in IL-1β and IL-18, respectively. The mature forms of cytokines are secreted worsening the neuroinflammatory process established. Then, the neuroinflammatory process established can be worsened by these mature cytokines. Adapted from Ribeiro et al., 2021 [<a href="#B105-pharmaceuticals-14-00933" class="html-bibr">105</a>]. Full article ">

25 pages, 5534 KiB

Open AccessArticle

Engineering Mitochondriotropic Carbon Dots for Targeting Cancer Cells

by Archontia Kaminari, Eleni Nikoli, Alexandros Athanasopoulos, Elias Sakellis, Zili Sideratou and Dimitris Tsiourvas

Pharmaceuticals 2021, 14(9), 932; https://doi.org/10.3390/ph14090932 - 16 Sep 2021

Cited by 16 | Viewed by 3837

Abstract

Aiming to understand and enhance the capacity of carbon dots (CDs) to transport through cell membranes and target subcellular organelles—in particular, mitochondria—a series of nitrogen-doped CDs were prepared by the one-step microwave-assisted pyrolysis of citric acid and ethylenediamine. Following optimization of the reaction [...] Read more.

Aiming to understand and enhance the capacity of carbon dots (CDs) to transport through cell membranes and target subcellular organelles—in particular, mitochondria—a series of nitrogen-doped CDs were prepared by the one-step microwave-assisted pyrolysis of citric acid and ethylenediamine. Following optimization of the reaction conditions for maximum fluorescence, functionalization at various degrees with alkylated triphenylphosphonium functional groups of two different alkyl chain lengths afforded a series of functionalized CDs that exhibited either lysosome or mitochondria subcellular localization. Further functionalization with rhodamine B enabled enhanced fluorescence imaging capabilities in the visible spectrum and allowed the use of low quantities of CDs in relevant experiments. It was thus possible, by the appropriate selection of the alkyl chain length and degree of functionalization, to attain successful mitochondrial targeting, while preserving non-toxicity and biocompatibility. In vitro cell experiments performed on normal as well as cancer cell lines proved their non-cytotoxic character and imaging potential, even at very low concentrations, by fluorescence microscopy. Precise targeting of mitochondria is feasible with carefully designed CDs that, furthermore, are specifically internalized in cells and cell mitochondria of high transmembrane potential and thus exhibit selective uptake in malignant cells compared to normal cells. Full article

(This article belongs to the Special Issue Nano Drug Carriers 2021)

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Graphical abstract

Graphical abstract
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(A) Hydrodynamic size distribution of N-doped CDs. (B) TEM bright-field images of N-doped CDs; scale bar: 20 nm. (C,D) High-resolution TEM (HRTEM) images of N-doped CDs. The insets are the corresponding fast Fourier transform (FFT) patterns. Full article ">Figure 2
(A) Excitation (blue line) and emission spectra (red line, λex = 352 nm) of N-doped CDs in water. Inset: photographs of their solutions under ambient light (left) and under a 365 nm UV lamp (right). (B) Emission spectra at progressively increasing excitation wavelengths (320 nm–420 nm). Inset: the corresponding normalized emission spectra at various excitation wavelengths. (C) Emission spectra of CDs and of their alkyl-TPP derivatives (λex = 352 nm). (D) Normalized UV–Vis spectra of CDs and of their alkyl-TPP derivatives. Full article ">Figure 3
CD•Rh and CD-C4TPP•Rh internalization in live MDA-MB-231 cells. (A,B) Cells were treated with 25 μg/mL of CD•Rh and CD-C4TPP•Rh for 24 h followed by treatment for 15 min with 100 nM of MitoTracker Green FM and LysoTracker Green DND-26. Overlay depicts co-localization of green and red fluorescence. Plots depict fluorescence intensity profiles along straight white lines. Pearson’s correlation coefficient for CD•Rh was calculated at 0.36 for MitoTracker and 0.63 for LysoTracker (A), whereas for CD-C4TPP•Rh at 0.68 for MitoTracker and 0.57 for Lysotracker (B); scale bar: 10 μm. (C) Cytotoxicity of CD and its derivatives in MDA-MB-231 cells. Cells were treated with increasing concentrations (125–500 μg/mL) of CD, CD-C4TPP and CD-C10TPP for 24 h. Cell viability was assessed by the MTT assay. Results are expressed as the mean ± standard error for at least three independent experiments and were analyzed using the Student t-test. Significance is relative to respective controls. * p > 0.05, ** p > 0.01, *** p > 0.001. Full article ">Figure 4
CD-C4TPP•Rh and CD-C10TPP•Rh internalization in live MDA-MB-231 cells. (A,B) Cells were treated with 50 μg/mL of CD-C4TPP•Rh and CD-C10TPP•Rh for 3 h followed by treatment for 15 min with 100 nM of MitoTracker GreenFM and LysoTracker Green DND-26. Overlay depicts co-localization of green and red fluorescence. Plots depict fluorescence intensity profiles along straight white lines. Pearson’s correlation coefficient for CD-C4TPP•Rh was calculated at 0.85 for MitoTracker and 0.51 for LysoTracker (A), whereas for CD-C10TPP•Rh at 0.41 for MitoTracker and 0.63 for LysoTracker (B); scale bar: 10 μm. Full article ">Figure 5
Internalization and retention patterns of CD-C4TPP•Rh in live MDA-MB-231 cells. Cells were treated with: (A) CD-C4TPP•Rh 50 μg/mL for 1, 3 and 24 h, (B) 50 μg/mL CD-C4TPP•Rh (RhoB 350 nM) and with plain RhoB (350 nM) for 3 h, (C) 50 and 10 μg/mL CD-C4TPP•Rh for 24 h, (D) 50 μg/mL CD-C4TPP•Rh for 3 h, 20 nM TMRM and 700 nM RhoB for 30 min. Following the above-mentioned incubation times, the compounds were then removed and cells were left in culture for 24 h. Respective graphs depict % quantification of fluorescence intensities. Overlay depicts bright field and red channel. Scale bar: 10 and 20 μm. Results are expressed as the mean ± standard error for at least three independent experiments and were analyzed using the Student t-test. Significance is relative to respective controls. *** p > 0.001. Full article ">Figure 6
Photostability of CD-C4TPP•Rh vs. TMRM and RhoB in live MDA-MB-231 cells and the corresponding fluorescence quantification (%) over time. Cells were treated with 50 μg/mL CD-C4TPP•Rh for 3 h, and TMRM 10 nM or RhoB 700 nM for 30 min. Photostability was assessed under continuous laser irradiation with 5 min intervals. Scale bar: 20 μm. Full article ">Figure 7
Staining patterns of CD-C4TPP•Rh in live MDA-MB-231 cells under various treatments and the corresponding fluorescence quantification over time. (A) Cells were treated with CD-C4TPP•Rh 50 μg/mL for 3 h, and TMRM 10 nM for 30 min following treatment with 2 μM of staurosporine (STS) for 2, 6 and 16 h. The respective graph depicts % quantification of fluorescence intensities over time. (B) Cells were treated with CD-C4TPP•Rh 50 μg/mL for 3 h following treatment with 10 and 30 μM of CCCP for 30 min. Respective graphs depict % quantification of fluorescence intensities. Overlay depicts bright field and red channel. Scale bar: 10 μm. Results are expressed as the mean ± standard error for at least three independent experiments and were analyzed using the Student t-test. Significance is relative to respective controls. ** p > 0.01, *** p > 0.001. Full article ">Figure 8
(A) Comparison of CD-C4TPP•Rh internalization in live COS-7 and MDA-MB-231 cells, as well as in co-culture of both cell lines. Cells were treated with 50 μg/mL of CD-C4TPP•Rh for 24 h. Overlay depicts bright field and red channel. Scale bar: 20 μm. (B) Fluorescence quantification (%) of different internalization intensities between cell lines. (C) Cytotoxicity of CD-C4TPP•Rh in MDA-MB-231 cells and COS-7 cells. Cells were treated with increasing concentrations (50–500 μg/mL) of CD-C4TPP•Rh for 24 h. Cell viability was assessed by the MTT assay. Results are expressed as the mean ± standard error for at least three independent experiments and were analyzed using the Student t-test. Significance is relative to respective controls. *** p > 0.001. Full article ">Scheme 1
Microwave synthesis of N-doped carbon dots and their subsequent functionalization with alkyl-TPP groups and rhodamine B isothiocyanate. Full article ">

11 pages, 1808 KiB

Open AccessArticle

Pilot Sub-Study of the Effect of Hepatitis C Cure by Glecaprevir/Pibrentasvir on the Gut Microbiome of Patients with Chronic Hepatitis C Genotypes 1 to 6 in the Mythen Study

by Bahtiyar Yilmaz, Lisa Ruckstuhl, Beat Müllhaupt, Lorenzo Magenta, Melanie Harrer Kuster, Olivier Clerc, Ralph Torgler and Nasser Semmo

Pharmaceuticals 2021, 14(9), 931; https://doi.org/10.3390/ph14090931 - 16 Sep 2021

Cited by 5 | Viewed by 2399

Abstract

In this small pilot sub-study, longitudinal gut microbiota composition changes, after successful treatment of hepatitis C virus (HCV) with the co-formulated glecaprevir/pibrentasvir (GLE/PIB), were analyzed before treatment (baseline) and 12 weeks post-treatment. Participating patients provided a fresh stool sample the week before their [...] Read more.

In this small pilot sub-study, longitudinal gut microbiota composition changes, after successful treatment of hepatitis C virus (HCV) with the co-formulated glecaprevir/pibrentasvir (GLE/PIB), were analyzed before treatment (baseline) and 12 weeks post-treatment. Participating patients provided a fresh stool sample the week before their study visit, from which microbial DNA was extracted and sequenced for the 16S rRNA region in an Illumina MiSeq2 platform. Microbial and statistical analyses were conducted to determine the alpha-diversity (number of different taxa within a sample) and beta-diversity (number of overlapping taxa between samples). Stool samples from 58 patients were eligible for analysis. There were 27 patients with HCV genotype 1, 10 with genotype 2, 16 with genotype 3, and 5 with genotype 4. No statistically significant differences in gut microbiota diversity, species richness, or microbial community pattern were found at baseline and at post-treatment Week 12. Lack of statistically significant differences remained consistent in further analysis by demographic and baseline disease characteristics. Surprisingly, no statistically significant changes in alpha- and beta-diversity were seen in the microbiota after GLE/PIB treatment, though there was a trend toward less richness over time. Further investigation is needed into this unexpected outcome to better understand the role of HCV treatment and the gut microbiota. Full article

(This article belongs to the Section Pharmacology)

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Figure 1
Microbial differences before treatment (baseline) vs. SVR12 groups. Taxonomy profile of patients at baseline and 12 weeks after the end of the treatment are shown (A) at phylum level and (B) at family level. (C) Microbial clustering is shown based on generalized UniFrac metrics using fecal DNA samples at baseline and SVR12. Non-parametric analysis of variance (Adonis) was used to test significant differences between groups on the PCoA plot, with a result of p > 0.05. The ellipses represent a 95% CI surrounding each disease group. B = baseline; E = end of treatment; PC = principal component; PCoA = principal coordinate analysis; SVR12 = sustained virologic response 12 weeks after the end of the treatment. Full article ">Figure 2
Overall species richness comparison of before treatment (baseline) and SVR12 groups. In this analysis, p < 0.05 was considered significant. Box-and-whisker plots display quartiles and range. SVR12 = sustained virologic response 12 weeks after the end of the treatment. Full article ">Figure 3
Overall beta-diversity differences before treatment (baseline) vs. SVR12 groups by patient and HCV genotype. Notable outliers with considerable changes are circled in red. Examples of minimal change are circled in blue for comparison. Non-parametric analysis of variance (Adonis) was used to test significant differences between groups on the PCoA (principal coordinate analysis) plot with a result of p > 0.05. SVR12 = sustained virologic response 12 weeks after the end of treatment. Full article ">

15 pages, 1245 KiB

Open AccessReview

Antidepressants in Alzheimer’s Disease: A Focus on the Role of Mirtazapine

by Ana Salomé Correia and Nuno Vale

Pharmaceuticals 2021, 14(9), 930; https://doi.org/10.3390/ph14090930 - 16 Sep 2021

Cited by 24 | Viewed by 10131

Abstract

Mirtazapine belongs to the category of antidepressants clinically used mainly in major depressive disorder but also used in obsessive-compulsive disorders, generalized anxiety, and sleep disturbances. This drug acts mainly by antagonizing the adrenergic α2, and the serotonergic 5-HT2 and 5-HT3 receptors. Neuropsychiatric symptoms, [...] Read more.

Mirtazapine belongs to the category of antidepressants clinically used mainly in major depressive disorder but also used in obsessive-compulsive disorders, generalized anxiety, and sleep disturbances. This drug acts mainly by antagonizing the adrenergic α2, and the serotonergic 5-HT2 and 5-HT3 receptors. Neuropsychiatric symptoms, such as depression and agitation, are strongly associated with Alzheimer’s disease, reducing the life quality of these patients. Thus, it is crucial to control depression in Alzheimer’s patients. For this purpose, drugs such as mirtazapine are important in the control of anxiety, agitation, and other depressive symptoms in these patients. Indeed, despite some contradictory studies, evidence supports the role of mirtazapine in this regard. In this review, we will focus on depression in Alzheimer’s disease, highlighting the role of mirtazapine in this context. Full article

(This article belongs to the Special Issue Treatment of Alzheimer Disease)

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Figure 1
The hallmarks of AD are the presence of a Aβ plaques and neurofibrillary tangles. Reprinted from “Alzheimer’s Brain (Disintegrating Microtubule)”, by BioRender.com (2021). Retrieved from [<a href="#B18-pharmaceuticals-14-00930" class="html-bibr">18</a>]. Available online: <a href="https://app.biorender.com/biorender-templates" target="_blank">https://app.biorender.com/biorender-templates</a> (accessed on 16 August 2021). Full article ">Figure 2
Findings such as decreased levels of 5-HT, NE, and BDNF, as well as dysregulation of the HPA axis and pro-inflammatory pathways, are associated with depression, contributing to the increase in neurodegeneration phenomena present in AD, such as the presence of Aβ plaques and brain atrophy. Adapted with permission from ref. [<a href="#B48-pharmaceuticals-14-00930" class="html-bibr">48</a>]. Copyright 2019 Frontiers Media S.A. Created with BioRender.com [<a href="#B49-pharmaceuticals-14-00930" class="html-bibr">49</a>]. Available online: <a href="https://biorender.com/" target="_blank">https://biorender.com/</a> (accessed on 16 August 2021). Full article ">Figure 3
Summary of the mechanism of action of mirtazapine. This drug is an antagonist of 5-HT2A, 5-HT2C, 5-HT3, H1, and α2 receptors, resulting in antidepressant and sedative effects [<a href="#B107-pharmaceuticals-14-00930" class="html-bibr">107</a>]. Image for illustrative purposes only. Created with BioRender.com [<a href="#B49-pharmaceuticals-14-00930" class="html-bibr">49</a>]. Available online: <a href="https://biorender.com/" target="_blank">https://biorender.com/</a> (accessed on 16 August 2021). Full article ">

15 pages, 1518 KiB

Open AccessArticle

In Vitro and In Vivo Evaluation of Oral Controlled Release Formulation of BCS Class I Drug Using Polymer Matrix System

by Mosab Arafat, Muhammad Sarfraz, Mohammad F. Bostanudin, Anna Esmaeil, Aisha Salam and Salahdein AbuRuz

Pharmaceuticals 2021, 14(9), 929; https://doi.org/10.3390/ph14090929 - 16 Sep 2021

Cited by 8 | Viewed by 4777

Abstract

Diltiazem hydrochloride is a calcium channel blocker, which belongs to the family of benzothiazepines. It is commonly used to treat hypertension and atrial fibrillation. Even though the drug has high solubility, its high permeability and rapid metabolism in the liver can limit the [...] Read more.

Diltiazem hydrochloride is a calcium channel blocker, which belongs to the family of benzothiazepines. It is commonly used to treat hypertension and atrial fibrillation. Even though the drug has high solubility, its high permeability and rapid metabolism in the liver can limit the bioavailability and increase the dose frequencies for up to four times per day. This study focused on a polymer matrix system not only to control the drug release but also to prolong the duration of bioavailability. The polymer matrices were prepared using different ratios of poloxamer-188, hydroxypropyl methylcellulose, and stearyl alcohol. In vitro and in vivo assessments took place using 24 rabbits and the results were compared to commercially available product Tildiem^® (60 mg tablet) as reference. Overall, the rate of drug release was sustained with the gradual increase of poloxamer-188 incorporated with hydroxypropyl methylcellulose and stearyl alcohol in the matrix system, achieving a maximum release period of 10 h. The oral bioavailability and pharmacokinetic parameters of diltiazem hydrochloride incorporated in polymer matrix system were similar to commercial reference Tildiem^®. In conclusion, the combination of polymers can have a substantial effect on controlling and prolonging the drug release pattern. The outcomes showed that poloxamer-188 combined with hydroxypropyl methylcellulose and stearyl alcohol is a powerful matrix system for controlling release of diltiazem hydrochloride. Full article

(This article belongs to the Section Pharmaceutical Technology)

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DLZ calibration curve in distilled water at 37 °C, (n = 6), values are means ± S.D. (n = 6). Full article ">Figure 2
The rate of DLZ-controlled release from polymer matrices of various drug to polymer percentages and Tildiem® over 600 min, using distilled water at 37 °C, values are means ± S.D. (n = 6). Full article ">Figure 3
The rate of DLZ controlled release from polymer matrices containing constant amount of drug and various polymer mixture percentages and Tildiem® over 600 min, using distilled water at 37 °C, values are means ± S.D. (n = 6). Full article ">Figure 4
The rate of DLZ controlled release from polymer matrices of test formulation (F8) over 600 min, using different simulated physiological media namely: FaSSIF, FeSSIF and SGF in comparison to distilled water (DW) at 37 °C, values are means ± S.D. (n = 6). Full article ">Figure 5
DSC Thermal profile of DLZ dispersion in test formulation and in pure crystalline powder. Full article ">Figure 6
Chromatograms of DLZ obtained from (A) plasma free of DLZ and (B) plasma spiked with10 µg/mL DLZ (Reprinted from ref. [<a href="#B46-pharmaceuticals-14-00929" class="html-bibr">46</a>]). Full article ">Figure 7
Plasma concentration of DLZ after oral administration of the commercial reference Tildiem® (60 mg tablet) and test formulation (F8), each point represents the mean ± SD, n = 12. Full article ">

22 pages, 1663 KiB

Open AccessReview

State of the Art on Approved Cystic Fibrosis Transmembrane Conductance Regulator (CFTR) Modulators and Triple-Combination Therapy

by Aniello Meoli, Valentina Fainardi, Michela Deolmi, Giulia Chiopris, Francesca Marinelli, Caterina Caminiti, Susanna Esposito and Giovanna Pisi

Pharmaceuticals 2021, 14(9), 928; https://doi.org/10.3390/ph14090928 - 15 Sep 2021

Cited by 24 | Viewed by 5937

Abstract

Cystic fibrosis (CF) is the most common life-limiting inherited disease in Caucasian populations, affecting approximately 80,000 people worldwide. CF is a complex multi-organ monogenic autosomal recessive disorder caused by a mutation in cystic fibrosis transmembrane conductance regulator (CFTR) gene. Since the [...] Read more.

Cystic fibrosis (CF) is the most common life-limiting inherited disease in Caucasian populations, affecting approximately 80,000 people worldwide. CF is a complex multi-organ monogenic autosomal recessive disorder caused by a mutation in cystic fibrosis transmembrane conductance regulator (CFTR) gene. Since the discovery of the CFTR gene in 1989, more than 2000 mutations have been identified so far and about 240 can cause CF. Until recently, the treatment for CF was aimed to prevent and manage the manifestations of CFTR dysfunction, primarily recurrent pulmonary infections and pancreatic exocrine failure. Over the past few decades, the therapeutic approach to CF has been revolutionized by the development of a new class of small molecules called CFTR modulators that target specific defects caused by mutations in the CFTR gene. CFTR modulators have been shown to change profoundly the clinical course of the CF, leading to meaningful improvements in the lives of a large proportion of people of CF heterozygous for F508del, especially if started in young children. Further studies are needed to extend the use of triple CFTR modulation therapy also for young children in order to prevent the irreversible effects of the disease and for patients with very rare mutations with a personalized approach to treatment. Full article

(This article belongs to the Special Issue Prevention, Diagnosis and Treatment of Pediatric Diseases)

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14 pages, 2129 KiB

Open AccessArticle

Population Pharmacokinetic Analysis of Pazopanib in Patients and Determination of Target AUC

by Agustos Cetin Ozbey, David Combarel, Vianney Poinsignon, Christine Lovera, Esma Saada, Olivier Mir and Angelo Paci

Pharmaceuticals 2021, 14(9), 927; https://doi.org/10.3390/ph14090927 - 15 Sep 2021

Cited by 7 | Viewed by 3090

Abstract

Pazopanib is a potent multi-targeted kinase inhibitor approved for the treatment of advanced renal cell carcinoma and soft tissue sarcoma. The pharmacokinetics of pazopanib is characterized by a significant inter- and intra-patient variability and a target through plasma concentration of 20.5 mg·L⁻¹ [...] Read more.

Pazopanib is a potent multi-targeted kinase inhibitor approved for the treatment of advanced renal cell carcinoma and soft tissue sarcoma. The pharmacokinetics of pazopanib is characterized by a significant inter- and intra-patient variability and a target through plasma concentration of 20.5 mg·L⁻¹. However, routine monitoring of trough plasma concentrations at fixed hours is difficult in daily practice. Herein, we aimed to characterize the pharmacokinetic (PK) profile of pazopanib and to identify a target area under the curve (AUC) more easily extrapolated from blood samples obtained at various timings after drug intake. A population pharmacokinetic (popPK) model was constructed to analyze pazopanib PK and to estimate the pazopanib clearance of a patient regardless of the time of sampling. Data from the therapeutic drug monitoring (TDM) of patients with cancer at Institute Gustave Roussy and a clinical study (phase I/II) that evaluates the tolerance to pazopanib were used. From the individual clearance, it is then possible to obtain the patient’s AUC. A target AUC for maximum efficacy and minimum side effects of 750 mg·h·L⁻¹ was determined. The comparison of the estimated AUC with the target AUC would enable us to determine whether plasma exposure is adequate or whether it would be necessary to propose therapeutic adjustments. Full article

(This article belongs to the Special Issue Precision Medicine in Oncology: Controlling the Pharmacokinetics Variability)

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27 pages, 2280 KiB

Open AccessReview

New Strategies for the Treatment of Atrial Fibrillation

by Norbert Jost, Torsten Christ and János Magyar

Pharmaceuticals 2021, 14(9), 926; https://doi.org/10.3390/ph14090926 - 15 Sep 2021

Cited by 7 | Viewed by 5556

Abstract

Atrial fibrillation (AF) is the most prevalent cardiac arrhythmia in the clinical practice. It significantly contributes to the morbidity and mortality of the elderly population. Over the past 25–30 years intense effort in basic research has advanced the understanding of the relationship between [...] Read more.

Atrial fibrillation (AF) is the most prevalent cardiac arrhythmia in the clinical practice. It significantly contributes to the morbidity and mortality of the elderly population. Over the past 25–30 years intense effort in basic research has advanced the understanding of the relationship between the pathophysiology of AF and atrial remodelling. Nowadays it is clear that the various forms of atrial remodelling (electrical, contractile and structural) play crucial role in initiating and maintaining the persistent and permanent types of AF. Unlike in ventricular fibrillation, in AF rapid ectopic firing originating from pulmonary veins and re-entry mechanism may induce and maintain (due to atrial remodelling) this complex cardiac arrhythmia. The present review presents and discusses in detail the latest knowledge on the role of remodelling in AF. Special attention is paid to novel concepts and pharmacological targets presumably relevant to the drug treatment of atrial fibrillation. Full article

(This article belongs to the Special Issue Ion Channels: Current Pharmacological Challenges)

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General scheme presenting the main factors, which are implicated in provoking and sustaining atrial fibrillation (AF). AF is usually initiated by spontaneous ectopic beats (extrasystole) originating from a pacemaker region (PVs, left upper pulmonary vein) and later is maintained by mechanism of atrial remodelling. The atrial fibrillation forms (lone, persistent and permanent) are promoted by single/mother wave and/or multiple wave/circuit re-entries. The main factors that may promote reentry are atrial ischaemia and inflammation, while the structural/morphological remodellings are sustained by atrial (micro)fibrosis and left atrial dilation. DAD and EAD—delayed and early afterdepolarization; PVs—pulmonary veins. Full article ">Figure 2
The most important transmembrane ionic inward and outward currents determining atrial action potential in sinus rhythm (SR, A) and in permanent atrial fibrillation (AF, B). The ion channel/current (functional elements) density changes are presented in bottom left column, while the expression level current subunit forming proteins and mRNA genes, respectively, are depicted in middle and right columns. The corresponding pictograms near each ion current name present the magnitude and time course of the current reflection of relative real sizes. ↓—decreased current or downregulated protein/gene, respectively; ↑—increased current or upregulated protein/gene, respectively; ↔—unchanged current, protein or gene, respectively; n.d.—no data. Full article ">Figure 3
Three proposed positive feedback loops to present the progredient development of atrial remodelling in AF. The electric and contractile remodelling are associated with the downregulation of L-type Ca2+-channels which as a result will cause the alteration of the whole Ca2+-homeostasis process. The loss of contractility and increase in compliance will induce the stretch of the atrial myocardium, which is considered to be the main inducing factor of the structural remodelling. The resulting intra-atrial circuits due to a reduction in wavelength and increased non-uniform tissue anisotropy and even zig-zag conduction (anatomical basis of re-entry or rotors). Reprinted with permission from ref [<a href="#B10-pharmaceuticals-14-00926" class="html-bibr">10</a>]. Copyright year 2002, Oxford Academic. Full article ">Figure 4
Possible pharmacological investigational strategies for developing new antiarrhythmic drugs for treating AF. Full article ">Figure 5
The chemical structures of the reviewed drugs in the present paper (specific-, multiple-, atrial selective ion channel blocker drugs and gap-junction modulator antiarrhythmic peptides. Full article ">

12 pages, 7576 KiB

Open AccessArticle

Incident Type 2 Diabetes Risk of Selective Estrogen Receptor Modulators in Female Patients with Breast Cancer

by Yeo-Jin Choi, Keunhyeong Bak, Yoon Yeo, Yongwon Choi and Sooyoung Shin

Pharmaceuticals 2021, 14(9), 925; https://doi.org/10.3390/ph14090925 - 14 Sep 2021

Cited by 8 | Viewed by 2939

Abstract

Accumulating evidence indicates a link between diabetes and cancer. Selective estrogen receptor modulators (SERMs) may increase diabetes risk via antiestrogen effects. This study investigated incident diabetes risk of SERM treatment and its effects on metastatic cancer and death prevention in breast cancer survivors. [...] Read more.

Accumulating evidence indicates a link between diabetes and cancer. Selective estrogen receptor modulators (SERMs) may increase diabetes risk via antiestrogen effects. This study investigated incident diabetes risk of SERM treatment and its effects on metastatic cancer and death prevention in breast cancer survivors. This retrospective cohort study included female patients with early-stage breast cancer, treated with or without SERMs, between 2008 and 2020 in a tertiary care hospital in Korea. Four propensity score-matched comparison pairs were designed: SERM use versus non-use, long-term use (≥1500 days) versus non-use, tamoxifen use versus non-use, and toremifene use versus non-use; then, logistic regression analysis was performed for risk analysis. SERMs in general were not associated with an elevated risk of diabetes; however, when used for ≥1500 days, SERMs—especially toremifene—substantially increased diabetes risk in breast cancer patients (OR 1.63, p = 0.048). Meanwhile, long-term SERM treatment was effective at preventing metastatic cancer (OR 0.20, p < 0.001) and death (OR 0.13, p < 0.001). SERM treatment, albeit generally safe and effective, may increase diabetes risk with its long-term use in women with breast cancer. Further studies are required to verify the association between toremifene treatment and incident diabetes. Full article

(This article belongs to the Special Issue Adverse Drug Reactions and Gender Differences)

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10 pages, 1297 KiB

Open AccessPerspective

Progress in the Treatment of Migraine Attacks: From Traditional Approaches to Eptinezumab

by Damiana Scuteri and Giacinto Bagetta

Pharmaceuticals 2021, 14(9), 924; https://doi.org/10.3390/ph14090924 - 13 Sep 2021

Cited by 7 | Viewed by 3073

Abstract

Migraine is the second cause of disability and of lost years of healthy life worldwide. Migraine is characterized by recurrent headache attacks and accompanying disabling symptoms lasting 4–48 h. In episodic migraine, attacks occur in less than 15 days per month and in [...] Read more.

Migraine is the second cause of disability and of lost years of healthy life worldwide. Migraine is characterized by recurrent headache attacks and accompanying disabling symptoms lasting 4–48 h. In episodic migraine, attacks occur in less than 15 days per month and in chronic migraine, in more than 15 monthly days. Whilst successful translation of pharmacological discoveries into efficacious therapeutics has been achieved in the preventative therapy of chronic migraine, treatment of acute migraine suffers the lack of effective advancements. An effective treatment affords complete freedom from pain two hours after therapy and provides the absence of the most bothersome symptom (MBS) associated with migraine after 2 h. However, available anti-migraine abortive treatments for acute attacks do not represent an effective and safe treatment for all the populations treated. In particular, the most used specific treatment is represented by triptans that offer 2-h sustained freedom from pain achieved in 18–50% of patients but they are contraindicated in coronary artery disease, stroke and peripheral vascular disease due to the vasoconstriction at the basis of their pharmacologic action. The most novel therapies, i.e., gepants and ditans, are without sufficient post-marketing data for secure use. Here, an attempt is proposed to analyse the rational basis and evidence in favour of investigating the efficacy and safety in acute migraine attacks of eptinezumab, i.e., monoclonal antibody (mAb) directed towards calcitonin gene-related peptide (CGRP) unique for intravenous infusion administration. Full article

(This article belongs to the Special Issue Monoclonal Antibodies for Migraine Prevention)

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19 pages, 1129 KiB

Open AccessReview

Osteosarcoma in Children: Not Only Chemotherapy

by Maura Argenziano, Chiara Tortora, Elvira Pota, Alessandra Di Paola, Martina Di Martino, Caterina Di Leva, Daniela Di Pinto and Francesca Rossi

Pharmaceuticals 2021, 14(9), 923; https://doi.org/10.3390/ph14090923 - 13 Sep 2021

Cited by 24 | Viewed by 3995

Abstract

Osteosarcoma (OS) is the most severe bone malignant tumor, responsible for altered osteoid deposition and with a high rate of metastasis. It is characterized by heterogeneity, chemoresistance and its interaction with bone microenvironment. The 5-year survival rate is about 67% for patients with [...] Read more.

Osteosarcoma (OS) is the most severe bone malignant tumor, responsible for altered osteoid deposition and with a high rate of metastasis. It is characterized by heterogeneity, chemoresistance and its interaction with bone microenvironment. The 5-year survival rate is about 67% for patients with localized OS, while it remains at 20% in case of metastases. The standard therapy for OS patients is represented by neoadjuvant chemotherapy, surgical resection, and adjuvant chemotherapy. The most used chemotherapy regimen for children is the combination of high-dose methotrexate, doxorubicin, and cisplatin. Considered that the necessary administration of high-dose chemotherapy is responsible for a lot of acute and chronic side effects, the identification of novel therapeutic strategies to ameliorate OS outcome and the patients’ life expectancy is necessary. In this review we provide an overview on new possible innovative therapeutic strategies in OS. Full article

(This article belongs to the Special Issue Osteosarcomas: Treatment Strategies)

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17 pages, 3367 KiB

Open AccessArticle

Development and Characterisation of Antibody-Based Optical Imaging Probes for Inflammatory Bowel Disease

by Matthijs David Linssen, Wouter Tjerk Rudolph Hooghiemstra, Annelies Jorritsma-Smit, Derk Pieter Allersma, Gerard Dijkstra and Wouter Bastiaan Nagengast

Pharmaceuticals 2021, 14(9), 922; https://doi.org/10.3390/ph14090922 - 13 Sep 2021

Cited by 5 | Viewed by 3168

Abstract

Monoclonal antibodies are an important addition to the medicinal treatment paradigm for IBD patients. While effective, these agents show a high degree of primary and secondary non-response, and methods to predict response are highly desired. Information on drug distribution at the target level [...] Read more.

Monoclonal antibodies are an important addition to the medicinal treatment paradigm for IBD patients. While effective, these agents show a high degree of primary and secondary non-response, and methods to predict response are highly desired. Information on drug distribution at the target level is often lacking. Fluorescent endoscopic imaging using labelled antibody drugs may provide insight regarding drug distribution, target engagement and drug response, but these assessments require stable and functional fluorescently-conjugated probes. Infliximab, vedolizumab, adalimumab and ustekinumab were conjugated to IRDye 800CW, IRDye 680LT and ZW800-1. The resulting 12 tracer candidates were analysed and characterised on SE-HPLC, SDS-PAGE, iso-electric focussing (IEF) and ELISA in order to evaluate their feasibility as candidate clinical tracers for cGMP development. Major differences in the conjugation results could be seen for each conjugated drug. For Infliximab, 2 conjugates (800CW and 680LT) showed formation of aggregates, while conjugates of all drugs with ZW800-1 showed reduced fluorescent brightness, reduced purification yield and formation of fragments. All 6 of these candidates were considered unfeasible. From the remaining 6, ustekinumab-680LT showed reduced binding to IL23, and was therefore considered unfeasible. Out of 12 potential tracer candidates, 5 were considered feasible for further development: vedolizumab-800CW, vedolizumab-680LT, adalimumab-800CW, adalimumab-680LT and ustekinumab-800CW. Infliximab-680LT and ustekinumab-680LT failed to meet the standards for this panel, but may be rendered feasible if tracer production methods were further optimized. Full article

(This article belongs to the Special Issue Hybrid Agents for Multimodal Imaging)

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Fluorescent emission of the 12 tracer candidates on Odyssey CLX, grouped by (A) IRDye 800CW, (B) IRDye 680LT and (C) ZW800-1. All antibodies conjugated to 680LT showed fluorescence counts comparable to or higher than 800CW. ZW800 conjugates are approximately half as bright as 800CW conjugates. Higher brightness is a desirable factor for a tracer as a lower dose is required for sufficient signal. Equal aliquots of each tracer batch were measured in a single plate. Data are depicted as the mean and standard error of mean of three tracer batches for each candidate. Full article ">Figure 2
Coomassie-stained white light and Odyssey fluorescence images of sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) gels. Each image row represents a single antibody. Conjugated samples of all 3 dyes both intact and reduced were run on the same gel both intact and after reduction by β-mercaptoethanol. PL represents the protein ladder, molecular weights for the ladder are depicted left of the coomassie stained bands. Intact antibodies showed clearly defined single bands around 150 kDa, in line with the unmodified reference antibody (ST) lane. Reduced gels showed 2 clearly defined bands, at 50 kDa for heavy chain and 25 kDa for light chain. Fluorescence shows comparable results for all intact antibodies, which show up as clear single bands in their respective fluorescent wavelength (Green: 800 nm, red: 700 nm). After reduction, fluorescent bands can be seen at the 50 kDa and 25 kDa locations for all antibodies, except when conjugated to ZW800. A poorly defined band can be observed at the very bottom of the gel for lanes of reduced ZW800 tracers, while negligible signal is detectable at the reduced band locations. Abbreviations: Fluo: Odyssey CLX fluorescence scan; PL: protein ladder; ST: unconjugated protein reference; 800CW: lane with antibody conjugated to IRDye 800CW; 680LT: lane with antibody conjugated to IRDye 680LT; ZW800: lane with antibody conjugated to ZW800-1. Full article ">Figure 3
Fluorescent intensity sum of heavy and light protein chains compared to the intensity of the intact antibody (IgG), on sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE). In all antibodies, the sum of mean fluorescent signals on heavy and light chains is comparable to full antibody when conjugated to 800CW (range: 82.0–95.4%) and 680LT (range: 106.6–121.5%). ZW800 conjugates do not show any relevant amounts of fluorescence on the reduced chains, instead a diffuse band at the fluid front of the gel can be seen that matches or exceed the fluorescence intensity of the intact IgG (125.7% (A), 116.5% (B), 159.8% (C) and 167.2% (D)). A sum of the heavy and light chain fluorescence signal over 100% of the intact antibody suggests the occurrence of fluorescence quenching in the intact molecules. Data are displayed as the mean and standard error of mean of three measurements. Full article ">Figure 4
Relative distribution of dye signals across heavy and light chains measured on sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE). ZW800 conjugates did not have visible heavy and light chain signal, data displayed from regions of interest placed at the estimated band position based on coomassie. Adalimumab (C) and ustekinumab (D) show similar distribution patterns, where dye is mostly on the heavy chain (range adalimumab: 73.1–78.2%, ustekinumab 75.3–82.5%). Infliximab-800CW (B) shows a very low amount of heavy chain binding (17.4%). Light chain binding is also seen to a lesser extent in other infliximab conjugates (680LT: 62.8% HC; ZW800: 63.1% HC). Vedolizumab (A) shows mostly heavy chain binding, but shows more per-dye fluctuation than other antibodies (800CW: 72.0% HC; 680LT: 75.9% HC; ZW800-1: 47.3% HC). Increased light chain binding could be related to tracer instability. Full article ">Figure 5
Coomassie blue stain and fluorescent imaging results of completed immobilized pH gradient (IPG) strips. Unmodified reference (ST) and samples from each conjugate (800CW, 680LT, ZW800) were run on separate IPG strips. pI bands and patterns for each reference antibody are highlighted with a solid blue outline. In tracer strips with coomassie, bands that correspond to the reference antibody are given a dotted blue outline, while bands that are seen only after conjugation are given a solid orange outline. In tracer fluorescence, green solid outlines correspond to bands and patterns visible on the fluorescent scans but not on the coomassie-stained strips. Dotted blue and orange outlines correspond to bands marked before with solid blue and orange outlines in reference antibody or coomassie-stained conjugated samples, respectively. The dashed yellow outline highlights areas of the gel that are aspecifically stained as a result of antibody buffer components. Infliximab (B) and Ustekinumab (D) showed the best retention of their native bands, but all conjugations (vedolizumab (A), infliximab (B), adalimumab (C), ustekinumab (D)) resulted in shifts in primary band pI and formation of new bands. Infliximab showed much more acidic bands both natively and especially after conjugation to dyes. Extensive acidic shift may be related to instability in the conjugated protein. The scale of the gels is shown in cm and in pI. pI scale was based on a pI standard ladder (not shown) run on the same system alongside the samples. pI’s of sample bands are interpolated based on the position on the gel and the distribution of pI’s in the pI standard. Full article ">Figure 6
Indirect enzyme-linked immunosorbent assay (ELISA) results for the determination of the affinity of vedolizumab (A), infliximab (B), adalimumab (C) and ustekinumab (D) conjugates to their target. Each panel (A–D) shows overlayed results of one antibody’s reference standard and conjugates of that antibody to 3 different dyes. Serially diluted binding curves were compared to an unmodified reference on the same plate. Affinity was calculated as the ratio between the EC50 values for reference and sample curves. Comparable binding affinities were shown for all conjugates except infliximab-800CW, infliximab-680LT and ustekinumab-680LT Full article ">Figure 7
Feasibility assessment radar graphs. A score is calculated in 6 critical parameters for each tracer candidate. Each parameter is scored on a 0–100 scale and plotted on the radar graph for performance comparison within and between antibodies. All six parameter scores were added up for a total feasibility score (<a href="#pharmaceuticals-14-00922-t001" class="html-table">Table 1</a>) to determine the most feasible antibody-dye combination. Int: Antibody Integrity; LE: label efficiency; Bind: Target binding affinity; Bright: Fluorescent brightness; Link: Link stability. Full article ">

15 pages, 4315 KiB

Open AccessArticle

Use of 3D Printing for the Development of Biodegradable Antiplatelet Materials for Cardiovascular Applications

by Juan Domínguez-Robles, Luis Diaz-Gomez, Emilia Utomo, Tingjun Shen, Camila J. Picco, Carmen Alvarez-Lorenzo, Angel Concheiro, Ryan F. Donnelly and Eneko Larrañeta

Pharmaceuticals 2021, 14(9), 921; https://doi.org/10.3390/ph14090921 - 11 Sep 2021

Cited by 37 | Viewed by 5033

Abstract

Small-diameter synthetic vascular grafts are required for surgical bypass grafting when there is a lack of suitable autologous vessels due to different reasons, such as previous operations. Thrombosis is the main cause of failure of small-diameter synthetic vascular grafts when used for this [...] Read more.

Small-diameter synthetic vascular grafts are required for surgical bypass grafting when there is a lack of suitable autologous vessels due to different reasons, such as previous operations. Thrombosis is the main cause of failure of small-diameter synthetic vascular grafts when used for this revascularization technique. Therefore, the development of biodegradable vascular grafts capable of providing a localized and sustained antithrombotic drug release mark a major step forward in the fight against cardiovascular diseases, which are the leading cause of death globally. The present paper describes the use of an extrusion-based 3D printing technology for the production of biodegradable antiplatelet tubular grafts for cardiovascular applications. For this purpose, acetylsalicylic acid (ASA) was chosen as a model molecule due to its antiplatelet activity. Poly(caprolactone) and ASA were combined for the fabrication and characterization of ASA-loaded tubular grafts. Moreover, rifampicin (RIF) was added to the formulation containing the higher ASA loading, as a model molecule that can be used to prevent vascular prosthesis infections. The produced tubular grafts were fully characterized through multiple techniques and the last step was to evaluate their drug release, antiplatelet and antimicrobial activity and cytocompatibility. The results suggested that these materials were capable of providing a sustained ASA release for periods of up to 2 weeks. Tubular grafts containing 10% (w/w) of ASA showed lower platelet adhesion onto the surface than the blank and grafts containing 5% (w/w) of ASA. Moreover, tubular grafts scaffolds containing 1% (w/w) of RIF were capable of inhibiting the growth of Staphylococcus aureus. Finally, the evaluation of the cytocompatibility of the scaffold samples revealed that the incorporation of ASA or RIF into the composition did not compromise cell viability and proliferation at short incubation periods (24 h). Full article

(This article belongs to the Special Issue Recent Advances in Polymers as Matrices for Drug Delivery Applications)

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6 pages, 638 KiB

Open AccessOpinion

The Mediterranean Diet as a Source of Natural Compounds: Does It Represent a Protective Choice against Cancer?

by Giuseppina Augimeri and Daniela Bonofiglio

Pharmaceuticals 2021, 14(9), 920; https://doi.org/10.3390/ph14090920 - 11 Sep 2021

Cited by 16 | Viewed by 3699

Abstract

The Mediterranean diet (MD), characterized by a high intake of fruits, vegetables, legumes, nuts and grains, a moderate intake of red wine and a reduced consumption of meat, has been considered one of the healthiest dietary patterns worldwide. Growing evidence suggests an inverse [...] Read more.

The Mediterranean diet (MD), characterized by a high intake of fruits, vegetables, legumes, nuts and grains, a moderate intake of red wine and a reduced consumption of meat, has been considered one of the healthiest dietary patterns worldwide. Growing evidence suggests an inverse relationship between high adherence to the MD and cancer, as well as other chronic degenerative diseases. The beneficial effects elicited by the MD pattern on cancer are due to the high contents of bioactive compounds contained in many foods of MD, which protect cells by oxidative and inflammatory processes and inhibit carcinogenesis by targeting the various hallmarks of cancer with different mechanisms of action. Although over the past decades numerous dietary and phytochemical compounds from Mediterranean food that have anticancer potential have been identified, a clear association between the MD eating pattern and cancer needs to be established. While we wait for answers to this question from well-conducted research, the empowering of the MD as a protective choice against cancer should represent the priority for public health policies. Full article

(This article belongs to the Special Issue Anticancer Compounds in Medicinal Plants)

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9 pages, 544 KiB

Open AccessCommunication

Testing the Stability of Drug Resistance on Cryopreserved, Gene-Engineered Human Induced Pluripotent Stem Cells

by Dilaware Khan, Ann-Christin Nickel, Sebastian Jeising, Constanze Uhlmann, Sajjad Muhammad, Daniel Hänggi, Igor Fischer and Ulf Dietrich Kahlert

Pharmaceuticals 2021, 14(9), 919; https://doi.org/10.3390/ph14090919 - 11 Sep 2021

Cited by 1 | Viewed by 2671

Abstract

Human induced pluripotent stem cells (hiPSCs) have emerged as a powerful tool for in vitro modelling of diseases with broad application in drug development or toxicology testing. These assays usually require large quantities of hiPSC, which can entail long-term storage via cryopreservation of [...] Read more.

Human induced pluripotent stem cells (hiPSCs) have emerged as a powerful tool for in vitro modelling of diseases with broad application in drug development or toxicology testing. These assays usually require large quantities of hiPSC, which can entail long-term storage via cryopreservation of the same cell charges. However, it is essential that cryopreservation does not oppose durable changes on the cells. In this project, we characterize one parameter of functionality of one that is well established in the field, in a different research context, an applied hiPSC line (iPS11), namely their resistance to a medium size library of chemo interventions (>160 drugs). We demonstrate that cells, before and after cryopreservation, do not change their relative overall drug response phenotypes, as defined by identification of the top 20 interventions causing dose-dependent reduction of cell growth. Importantly, also frozen cells that are exogenously enforced for stable overexpression of oncogenes myelocytomatosis (cMYC) or tumor protein 53 mutation (TP53R175H), respectively, are not changed in their relative top 20 drugs response compared to their non-frozen counterparts. Taken together, our results support iPSCs as a reliable in vitro platform for in vitro pharmacology, further raising hopes that this technology supports biomarker-associated drug development. Given the general debate on ethical and economic problems associated with the reproducibly crisis in biomedicine, our results may be of interest to a wider audience beyond stem cell research. Full article

(This article belongs to the Special Issue Drug Screening or Drug Designing Based on Stem Cell Models)

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14 pages, 3380 KiB

Open AccessArticle

Gene Dosage Analysis on the Single-Cell Transcriptomes Linking Cotranslational Protein Targeting to Metastatic Triple-Negative Breast Cancer

by Yining Liu and Min Zhao

Pharmaceuticals 2021, 14(9), 918; https://doi.org/10.3390/ph14090918 - 10 Sep 2021

Cited by 3 | Viewed by 2568

Abstract

Many recent efforts have been put into the association between expression heterogeneity and different cell types and states using single-cell RNA transcriptome analysis. There is only limited understanding of gene dosage effects for the genetic heterogeneity at the single-cell level. By focusing on [...] Read more.

Many recent efforts have been put into the association between expression heterogeneity and different cell types and states using single-cell RNA transcriptome analysis. There is only limited understanding of gene dosage effects for the genetic heterogeneity at the single-cell level. By focusing on concordant copy number variation (CNV) and expression, we presented a computational framework to explore dosage effect for aggressive metastatic triple-negative breast cancer (TNBC) at the single-cell level. In practice, we collected CNV and single-cell expression data from the same patients with independent technologies. By focusing on 47,198 consistent copy number gains (CNG) and gene up-regulation from 1145 single cells, ribosome proteins with important roles in protein targeting were enriched. Independent validation in another metastatic TNBC dataset further prioritized signal recognition particle-dependent protein targeting as the top functional module. More interesting, the increased ribosome gene copies in TNBC may associate with their enhanced stemness and metastatic potential. Indeed, the prioritization of a well-upregulated functional module confirmed by high copy numbers at the single-cell level and contributing to patient survival may indicate the possibility of targeted therapy based on ribosome proteins for TNBC. Full article

(This article belongs to the Special Issue Potential Molecular Targets and Therapeutics in Triple-Negative Breast Cancer)

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The workflow to explore the global CNV patterns across multiple cell types with gene expression changes at the single-cell level. This workflow starts from the CNV and expression profiling at the single-cell level from the same tissue samples. By mapping the CNV and gene expression changes in the same cells, this workflow will identify concordant CNV and expression changes. By running the gene–gene interaction network analysis, the workflow will build highly connected functional modules to prioritize the key genes with significant gene dosage effects. We also recommend validating the whole process by integrating other independent datasets with both CNV and expression data at the single-cell level. The final validated functional modules in multiple cells will be highly reliable for further clinical feature evaluation and experimental validation. Full article ">Figure 2
Functional analysis of two top-ranked gene lists with concordant copy number gain and upregulation (CNG-UP) genes. (A) The bar plot shows the gene ontology (GO) cluster representatives for the 94 CNG-UP genes from the primary scRNAseq dataset. The x-axis indicates the log of corrected p-values. (B) The five functional modules-based gene–gene interactions for the 94 CNG-UP genes from the primary scRNAseq dataset. The different colors represent the five identified functional modules. The top ranked modules related to cotranslational protein targeting membranes are depicted in red nodes. Four other identified modules are purine ribonucleoside triphosphate biosynthetic processes (blue nodes), SCF(Skp2)-mediated degradation of p27/p21 (green nodes), apoptosis (purple nodes), and neutrophil degranulation (orange nodes). (C) The representative GO clusters for 86 CNG-UP genes from the validating dataset. The x-axis indicates the log of corrected p-values. (D) Three functional modules summarized from gene–gene interactions for the 86 CNG-UP genes from the validating dataset. The top three ranked modules are SRP-dependent cotranslational protein targeting to membranes (in red), the interferon-gamma-mediated signaling pathway (in blue), and cellular responses to stress (in green). (E) Overlapping of the top-ranked modules associated with protein targeting from primary and validating datasets. Seven genes are shared in both modules. (F) The representative GO clusters for 33 CNG-UP genes related to SRP-dependent cotranslational protein targeting to membranes combined from the primary and validating datasets. The x-axis indicates the log of corrected p-values. Full article ">Figure 3
The overall mutational and prognostic features for 33 genes with increased gene expression induced by CNGs. (A) The gene–gene interaction network showing the mutational frequency in 6688 breast cancer samples combined from 12 studies. The size and color of each node is correlated with the number of connections. (B) The mutational summary for twelve breast cancer datasets; (C) The overall survival plot shows the median survival months for patients with or without mutations on the 33 genes. Full article ">Figure 4
Clinical features summary for the 33 ribosome genes on (A) race category; (B) neoplasm histological grade; (C) tumor stage; (D) chemotherapy treatment; (E) diagnosis age; (F) aneuploidy score; and (G) ragnum hypoxia score. The p-values are the statistical tests between patients with or without any genetic mutations on the 33 genes. Full article ">Figure 5
Intratumor heterogeneity and its relationship with five key cellular states. (A) The diversity based on the Shannon–Weiner index for six patients with hundreds of cells. (B) The correlation between Simpson index and Shannon–Weiner index in six patients; (C) The t-SNE biplots of cells (red) and genes (purple). The teal lines and labels correspond to the cell states’ vector. The orange circles represent cells and purple “+” are the genes ranked with the top 10% of variations. Full article ">

17 pages, 889 KiB

Open AccessArticle

Antifungal Activity of Extracts, Fractions, and Constituents from Coccoloba cowellii Leaves

by Daniel Méndez, Julio C. Escalona-Arranz, Enrique Molina Pérez, Kenn Foubert, An Matheeussen, Emmy Tuenter, Ann Cuypers, Paul Cos and Luc Pieters

Pharmaceuticals 2021, 14(9), 917; https://doi.org/10.3390/ph14090917 - 10 Sep 2021

Cited by 4 | Viewed by 3426

Abstract

Coccoloba cowellii Britton (Polygonaceae, order Caryophyllales) is an endemic and critically endangered plant species that only grows in the municipality of Camagüey, a province of Cuba. A preliminary investigation of its total methanolic extract led to the discovery of promising antifungal activity. In [...] Read more.

Coccoloba cowellii Britton (Polygonaceae, order Caryophyllales) is an endemic and critically endangered plant species that only grows in the municipality of Camagüey, a province of Cuba. A preliminary investigation of its total methanolic extract led to the discovery of promising antifungal activity. In this study, a bioassay-guided fractionation allowed the isolation of quercetin and four methoxyflavonoids: 3-O-methylquercetin, myricetin 3,3′,4′-trimethyl ether, 6-methoxymyricetin 3,4′-dimethyl ether, and 6-methoxymyricetin 3,3′,4′-trimethyl ether. The leaf extract, fractions, and compounds were tested against various fungi and showed strong in vitro antifungal activity against Cryptococcus neoformans and various Candida spp. with no cytotoxicity (CC₅₀ > 64.0 µg/mL) on MRC-5 SV2 cells, determined by a resazurin assay. A Candida albicans SC5314 antibiofilm assay indicated that the antifungal activity of C. cowellii extracts and constituents is mainly targeted to planktonic cells. The total methanolic extract showed higher and broader activity compared with the fractions and mixture of compounds. Full article

(This article belongs to the Special Issue Natural Pharmacons: Biologically Active Plant Based Pharmaceuticals)

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16 pages, 2398 KiB

Open AccessArticle

Chemoreversal Agents from Taiwanofungus Genus and Their More Potent Methyl Derivatives Targeting Signal Transducer and Activator of Transcription 3 (STAT3) Phosphorylation

by Ko-Hua Yu, Chin-Chuan Hung, Tian-Shung Wu, Chin-Fu Chen, I-Ting Wu, Ping-Chung Kuo, Sio-Hong Lam and Hsin-Yi Hung

Pharmaceuticals 2021, 14(9), 916; https://doi.org/10.3390/ph14090916 - 10 Sep 2021

Cited by 2 | Viewed by 2077

Abstract

Multidrug resistance (MDR), for which the mechanisms are not yet fully clear, is one of the major obstacles to cancer treatment. In recent years, signal transducer and activator of transcription 3 (STAT3) were found to be one of the important MDR mechanism pathways. [...] Read more.

Multidrug resistance (MDR), for which the mechanisms are not yet fully clear, is one of the major obstacles to cancer treatment. In recent years, signal transducer and activator of transcription 3 (STAT3) were found to be one of the important MDR mechanism pathways. Based on the previous research, zhankuic acid A, B, and C were found to have collateral sensitivity effects on MDR cancer cells, and MDR inhibitory activity of zhankuic acid methyl ester was found to be better than that of its acid. Therefore, we executed a systematic examination of the structure–activity relationship of zhankuic acid methyl ester derivatives to collateral sensitivity in MDR cancer cells. The results showed that compound 12 is the best in terms of chemoreversal activity, where the reversal fold was 692, and the IC₅₀ value of paclitaxel combined with 10 μM compound 12 treatment was 1.69 nM in MDR KBvin cells. Among all the derivatives, methyl ester compounds were found to be better than their acids, and a detailed discussion of the structure–activity relationships of all of the derivatives is provided in this work. In addition, compounds 8, 12, and 26 were shown to influence the activation of STAT3 in KBvin cells, accounting for part of their chemoreversal effects. Our results may provide a new combined therapy with paclitaxel to treat multidrug-resistant cancers and provide a new therapy option for patients. Full article

(This article belongs to the Special Issue Identification of Novel Compounds against Multidrug-Resistant Cancer Cells)

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