WO2018029685A1

WO2018029685A1 - Compositions and methods for treating a fear memory

Info

Publication number: WO2018029685A1
Application number: PCT/IL2017/050880
Authority: WO
Inventors: Asya Rolls; Hilla AZULAY-DEBBY; Fahed HAKIM; BOSHNAK (THAWHO), Nadia
Original assignee: Technion Research & Development Foundation Limited
Priority date: 2016-08-11
Filing date: 2017-08-09
Publication date: 2018-02-15

Abstract

A method for inhibiting a formation of an extracellular matrix which includes a chondroitin sulfate proteoglycan, by contacting the extracellular matrix with xyloside, matrix metalloproteinase (MMP) inhibitor, or a combination of xyloside and a MMP inhibitor, is provided.

Description

COMPOSITIONS AND METHODS FOR TREATING A FEAR MEMORY

FIELD OF THE INVENTION

[001] A composition of xyloside, matrix metalloproteinase (MMP) inhibitor, or a combination of xyloside and a MMP inhibitor for inhibiting a formation of an extracellular matrix which includes a chondroitin sulfate proteoglycan, is provided.

BACKGROUND OF THE INVENTION

[002] Post-traumatic stress disorder (PTSD) may develop in people who have been through a traumatic event. A traumatic event can be a life/safety threatening event. As a result of such an event a person may experience feelings of intense fear, helplessness or horror.

[003] People suffering from PTSD may experience feelings of panic or extreme fear, similar to the fear they felt during the traumatic event. A person with PTSD experiences four main types of difficulties. Re-living the traumatic event - The person relives the event through unwanted and recurring memories, often in the form of vivid images and nightmares. There may be intense emotional or physical reactions, such as sweating, heart palpitations or panic when reminded of the event. Being overly alert or wound up - a person may experience sleeping difficulties, irritability and lack of concentration, becoming easily startled and constantly on the lookout for signs of danger. Avoiding reminders of the event - a person may deliberately avoid activities, places, people, thoughts or feelings associated with the event because they bring back painful memories. Feeling emotionally numb - a person loses interest in day-to-day activities, feels cut off and detached from friends and family, or feels emotionally flat and numb. [004] It's common for people with PTSD to experience other mental health problems at the same time. These may have developed directly in response to the traumatic event or have followed the PTSD. These additional problems, most commonly depression, anxiety, and alcohol or drug use, are more likely to occur if PTSD has persisted for a long time.

[005] Unfortunately, PTSD is the fourth-most common psychiatric disorder, affecting about 7.7 million American adults. From a clinical perspective, improving treatment regimens and identifying prevention measures is of critical importance. The existing pharmacological and psychotherapeutic treatments for fear-related disorders include antidepressant, anxiolytic drugs as well as enhancement of gamma-aminobutyric acid (GABA) activity. These agents are not specific in their actions, can trigger severe side effects and are only effective in some cases.

[006] In many cases of fear-related disorders, particularly PTSD, the traumatic incident can be identified and early intervention may prevent the development of PTSD. Perineuronal nets (PNNs) are specialized extracellular matrix structures around interneurons in the brain and spinal cord. Chondroitin sulfate proteoglycans (CSPGs) are crucial components of PNNs.

[007] The absence of PNNs have been suggested to be a permissive factor that allows synaptic plasticity in the adolescent. It has been shown that the degradation of PNNs induced by chondroitinase ABC (ChABC) converts the plasticity of the adult CNS to an adolescent phenotype.

SUMMARY OF THE INVENTION

[008] In one embodiment, this invention provides a method for inhibiting a formation of an extracellular matrix comprising a chondroitin sulfate proteoglycan in a subject in need thereof, comprising, administering to the subject a composition comprising xyloside, matrix metalloproteinase (MMP) inhibitor, or a combination of xyloside and a MMP inhibitor, thereby inhibiting the formation of an extracellular matrix comprising a chondroitin sulfate proteoglycan in a subject in need thereof.

[009] In one embodiment, this invention further provides a method for inhibiting fear memory, trauma or an anxiety disorder in a subject in need thereof, comprising administering to said subject a chondroitin sulfate proteoglycan inhibitor, matrix metalloproteinase (MMP) inhibitor, or a combination of a chondroitin sulfate proteoglycan inhibitor and a MMP inhibitor, thereby inhibiting fear memory, trauma or an anxiety disorder in a subject in need thereof.

[010] In one embodiment, this invention further provides a method for inhibiting fear memory in a subject in need thereof, comprising inhibiting a formation of an extracellular matrix comprising a chondroitin sulfate proteoglycan in the brain of the subject, thereby inhibiting fear memory in a subject in need thereof.

[011] In another embodiment, formation of an extracellular matrix comprising a chondroitin sulfate proteoglycan is within the brain. In another embodiment, formation of an extracellular matrix comprising a chondroitin sulfate proteoglycan is within the the amygdala, hippocampus, or both. In another embodiment, inhibiting the formation of the extracellular matrix comprising a chondroitin sulfate proteoglycan is affecting memory. In another embodiment, inhibiting the formation of the extracellular matrix comprising a chondroitin sulfate proteoglycan is inhibiting fear memory, trauma, or an anxiety disorder. In another embodiment, inhibiting the formation of the extracellular matrix comprising a chondroitin sulfate proteoglycan is reducing the risk for developing a posttraumatic stress disorder. [012] In another embodiment, inhibiting fear memory is inhibiting the formation of the extracellular matrix comprising a chondroitin sulfate proteoglycan in the brain. In another embodiment, inhibiting fear memory is reducing the risk for developing a fear memory, trauma, or an anxiety disorder.

[013] In one embodiment, this invention further provides a composition comprising a chondroitin sulfate proteoglycan inhibitor and a MMP inhibitor.

BRIEF DESCRIPTION OF THE DRAWINGS

[014] Figure 1. A bar graph showing attenuation of fear memory. Percent of freezing duration in response to the tone presentation. Two conditions were compared: (1) mice treated with IP injections of Xyloside (2) mice injections saline and received control group. The groups showed no significant (***/?<0.001) difference in terms.

[015] Figure 2. A bar graph showing that attenuation of fear memory is time dependent. Percent of freezing duration in response to the tone presentation. When the injection of Xyloside was done immediately before the mice were reintroduced to the fear conditioning chamber: (1) mice treated with IP injections of Xyloside (2) mice injections saline and received control group.

[016] Figure 3. A graph showing the percent of freezing duration in response to the tone presentation.

[017] Figures 4A-B. Are graphs showing the effect of an open field test after 48 hours from the injection. Results of neurobehavioral OFT testing in mice that were exposed to fear-conditioning and injected them with xyloside or saline directly into the amygdala. (Figure 4A) Time in the center field (Figure 4B) Time in border field. The groups showed no significant difference in terms of the length of time spent in the central and border squares. DETAILED DESCRIPTION OF THE INVENTION

[018] The present invention provides, in one embodiment, a method for inhibiting a formation of an extracellular matrix comprising a chondroitin sulfate proteoglycan in a subject in need thereof, comprising, administering to the subject a composition comprising xyloside, matrix metalloproteinase (MMP) inhibitor, or a combination of xyloside and a MMP inhibitor, thereby inhibiting the formation of an extracellular matrix comprising a chondroitin sulfate proteoglycan in a subject in need thereof. In another embodiment, xyloside is beta-xyloside and/or beta-D- xyloside.

[019] In one embodimentxyloside is: beta-D-xyloside, beta-xyloside, 4-Nitrophenyl- beta-D-xyloside, 3-D-xyloside, 3-O-beta-D-xyloside, 7-O-beta-D-xyloside, alpha-D- xyloside, 3-o-xyloside, or any combination thereof.

[020] In another embodiment, the invention is directed at inhibiting the formation of an extracellular matrix comprising a chondroitin sulfate proteoglycan. In another embodiment, the invention is directed at inhibiting the formation of an extracellular matrix complex comprising a chondroitin sulfate proteoglycan. In another embodiment, chondroitin sulfate proteoglycan comprises neurocan. In another embodiment, chondroitin sulfate proteoglycan comprises phosphocan. In another embodiment, chondroitin sulfate proteoglycan comprises both phosphocan and neurocan. In another embodiment, chondroitin sulfate proteoglycan comprises both phosphocan, versican, NG2, aggrecan, lectican, neurocan, or any combination thereof. In another embodiment, an extracellular matrix comprises receptor protein tyrosine phosphatase beta.

[021] In another embodiment, inhibiting the formation of an extracellular matrix comprising a chondroitin sulfate proteoglycan is inhibiting the attachment of glycosaminoglycans to core proteins of proteoglycans in the extracellular matrix. In another embodiment, inhibiting the formation of an extracellular matrix is attenuating fear memory.

[022] In one embodiment, provided herein that a method for inhibiting a formation of an extracellular matrix comprising a chondroitin sulfate proteoglycan in a subject in need thereof is a method for inhibiting a formation of an extracellular matrix comprising a chondroitin sulfate proteoglycan in the brain of a subject in need thereof. In another embodiment, inhibiting a formation of an extracellular matrix comprising a chondroitin sulfate proteoglycan in the brain is inhibiting a formation of an extracellular matrix comprising a chondroitin sulfate proteoglycan in the amygdala, hippocampus, or both. In one embodiment, provided herein that a method for inhibiting a formation of an extracellular matrix comprising a chondroitin sulfate proteoglycan in the amygdala, hippocampus, or both.

[023] In one embodiment, inhibiting a formation of an extracellular matrix is attenuating a formation of an extracellular matrix. In one embodiment, inhibiting a formation of an extracellular matrix is inhibiting a formation of an extracellular matrix in the brain of a subject after the subject experienced a fear episode. In one embodiment, inhibiting a formation of an extracellular matrix is inhibiting a formation of an extracellular matrix in the brain of a subject after the subject experienced a traumatic episode or trauma. In one embodiment, inhibiting a formation of an extracellular matrix is inhibiting a formation of an extracellular matrix in the brain of a subject after the subject experienced a psychological trauma.

[024] In one embodiment, inhibiting a formation of an extracellular matrix comprising a chondroitin sulfate proteoglycan after fear conditioning or a traumatic event, prevents the manifestation of fear-related behavior or trauma 1 to 48 hours later. In one embodiment, inhibiting a formation of an extracellular matrix comprising a chondroitin sulfate proteoglycan after fear conditioning or a traumatic event, prevents the manifestation of fear-related behavior or trauma 2 to 24 hours later. In one embodiment, inhibiting a formation of an extracellular matrix comprising a chondroitin sulfate proteoglycan after fear conditioning or a traumatic event, prevents the manifestation of fear-related behavior or trauma 5 to 36 hours later.

[025] In one embodiment, inhibiting a formation of an extracellular matrix is inhibiting a formation of an extracellular matrix in the brain of a subject at risk of experiencing a fear episode. In one embodiment, inhibiting a formation of an extracellular matrix is inhibiting a formation of an extracellular matrix in the brain of a subject at risk of experiencing a traumatic episode, trauma or an anxiety disorder. In one embodiment, inhibiting a formation of an extracellular matrix is inhibiting a formation of an extracellular matrix in the brain of a subject at risk of experiencing a psychological trauma or an anxiety disorder. In one embodiment, a subject at risk of experiencing a psychological trauma, a fear episode, anxiety disorder, or trauma is a mentally/psychologically healthy subject within a stressful or trauma inducing environment (such as but not limited to a war zone). In one embodiment, a subject at risk of experiencing a psychological trauma, anxiety disorder, a fear episode, or trauma is a mentally/psychologically healthy subject about to enter stressful or trauma inducing environment (such as but not limited to a war zone). [026] In another embodiment, xyloside, matrix metalloproteinase (MMP) inhibitor, or a combination of xyloside and a MMP inhibitor are used for the preparation of a medicament for inhibiting a formation of an extracellular matrix in a subject in need thereof as described herein.

[027] In some embodiments, inhibiting the formation of the extracellular matrix comprising a chondroitin sulfate proteoglycan is affecting memory. In some embodiments, inhibiting the formation of the extracellular matrix comprising a chondroitin sulfate proteoglycan is affecting fear memory and/or trauma. In some embodiments, inhibiting the formation of the extracellular matrix comprising a chondroitin sulfate proteoglycan is inhibiting or attenuating trauma or an anxiety disorder in a subject having a fear memory or at risk of having a fear memory. In some embodiments, inhibiting the formation of the extracellular matrix comprising a chondroitin sulfate proteoglycan is inhibiting the formation of fear memory and/or trauma. In some embodiments, inhibiting the formation of the extracellular matrix comprising a chondroitin sulfate proteoglycan is reducing the risk of trauma or anxiety disorder in a subject having a fear memory.

[028] In another embodiment, inhibiting the formation of the extracellular matrix comprising a chondroitin sulfate proteoglycan is reducing the risk for developing a fear memory, trauma or an anxiety disorder. In another embodiment, inhibiting the formation of the extracellular matrix comprising a chondroitin sulfate proteoglycan is reducing the risk for developing a fear memory, trauma or an anxiety disorder after induction. In another embodiment, induction includes exposure to a traumatic event. In another embodiment, a traumatic event is an experience that causes physical, emotional, psychological distress, or harm. In another embodiment, a traumatic event is an event that is perceived and experienced as a threat to one's safety or to the stability of one's world. [029] In another embodiment, the methods of the present invention are utilized after a traumatic event. In another embodiment, the methods of the present invention are utilized after a traumatic event for reducing the risks of psychiatric deterioration. In another embodiment, the methods of the present invention are utilized before a traumatic event for reducing the traumatic impact and a psychiatric pathology associated with the traumatic event. In another embodiment, the methods of the present invention are utilized before a traumatic event for reducing the traumatic impact and/or the development of a psychiatric pathology associated with the traumatic event.

[030] In one embodiment, a subject in need thereof or a subject is a subject that experienced a trauma or a traumatic event in the past 48 hours. In one embodiment, a subject in need thereof or a subject is a subject that experienced a trauma or a traumatic event in the past 36 hours. In one embodiment, a subject in need thereof or a subject is a subject that experienced a trauma or a traumatic event in the past 24 hours. In one embodiment, a subject in need thereof or a subject is a subject that experienced a trauma or a traumatic event in the past 18 hours. In one embodiment, a subject in need thereof or a subject is a subject that experienced a trauma or a traumatic event in the past 12 hours. In one embodiment, a subject in need thereof or a subject is a subject that experienced a trauma or a traumatic event in the past 8 hours.

[031] In one embodiment, a subject in need thereof or a subject is a subject afflicted with a post-traumatic stress disorder. In one embodiment, a subject in need thereof or a subject is a subject at risk of developing a trauma or a post-traumatic stress disorder within 48 hours. In one embodiment, a subject in need thereof or a subject is a subject at risk of developing a trauma or a post-traumatic stress disorder within 36 hours. In one embodiment, a subject in need thereof or a subject is a subject at risk of developing a trauma or a post-traumatic stress disorder within 24 hours. In one embodiment, a subject in need thereof or a subject is a subject at risk of developing a trauma or a post-traumatic stress disorder within 18 hours. In one embodiment, a subject in need thereof or a subject is a subject at risk of developing a trauma or a post-traumatic stress disorder within 12 hours. In one embodiment, a subject in need thereof or a subject is a subject at risk of developing a trauma or a post-traumatic stress disorder within 8 hours. [032] In another embodiment, the methods of the present invention are utilized 0.1 to 48 hours after a traumatic event or after fear conditioning. In another embodiment, the methods of the present invention are utilized 0.1 to 30 hours after a traumatic event or after fear conditioning. In another embodiment, the methods of the present invention are utilized 0.1 to 24 hours after a traumatic event or after fear conditioning. In another embodiment, the methods of the present invention are utilized 0.1 to 12 hours after a traumatic event or after fear conditioning. In another embodiment, the methods of the present invention are utilized 0.1 to 10 hours after a traumatic event or after fear conditioning. In another embodiment, the methods of the present invention are utilized 0.1 to 6 hours after a traumatic event or after fear conditioning. In another embodiment, the methods of the present invention comprise the administration of xyloside.

[033] In one embodiment, inhibiting the formation of the extracellular matrix comprising a chondroitin sulfate proteoglycan is inhibiting, treating, ameliorating, or any combination thereof of a posttraumatic stress disorder or an anxiety disorder. In one embodiment, inhibiting the formation of the extracellular matrix comprising a chondroitin sulfate proteoglycan is reducing the risk of a posttraumatic stress disorder or an anxiety disorder. In one embodiment, inhibiting the formation of the extracellular matrix comprising a chondroitin sulfate proteoglycan is delaying the onset of posttraumatic stress disorder or an anxiety disorder. In one embodiment, inhibiting the formation of the extracellular matrix comprising a chondroitin sulfate proteoglycan is reducing the risk for developing a posttraumatic stress disorder or an anxiety disorder.

[034] In one embodiment, a subject is an aid worker, a medical worker, or a combatant experiencing a traumatic experience. In one embodiment, a subject is an aid worker, a medical worker, or a combatant recently experienced a traumatic experience. In one embodiment, a subject is an aid worker, a medical worker, or a combatant recently destined to experience a traumatic experience. In one embodiment, a subject is an a human or an animal. In one embodiment, a subject is a subject is at risk of developing a fear memory or an anxiety disorder.

[035] In one embodiment, the MMP inhibitor is MMP-2 inhibitor, MMP-9 inhibitor, or a combination thereof. In one embodiment, the MMP inhibitor is GM6001. In one embodiment, the MMP inhibitor is TIMP-1. In one embodiment, the MMP inhibitor is Batimastat (BB-94). In one embodiment, the MMP inhibitor is marimastat (BB-2516). In one embodiment, the MMP inhibitor is an agent which inhibits MMP production. In one embodiment, the MMP inhibitor is an agent which inhibits MMP activity. In one embodiment, the MMP inhibitor is a macrocyclic lactone. In one embodiment, the MMP inhibitor is a Bryostatin. In one embodiment, the MMP inhibitor is bryostatin-1. In one embodiment, the MMP inhibitor is a selective MT1-MMP cyclic peptide inhibitor.

[036] In one embodiment, the invention provides a method for inhibiting fear memory or an anxiety disorder in a subject in need thereof, comprising administering to the subject a chondroitin sulfate proteoglycan inhibitor, matrix metalloproteinase (MMP) inhibitor, or a combination of a chondroitin sulfate proteoglycan inhibitor and a MMP inhibitor, thereby inhibiting fear memory or an anxiety disorder in a subject in need thereof. In one embodiment, inhibiting fear memory is avoiding anxiety disorder. In one embodiment, inhibiting fear memory is avoiding posttraumatic stress disorder. In one embodiment, chondroitin sulfate proteoglycan inhibitor inhibits the expression of a chondroitin sulfate proteoglycan. In one embodiment, chondroitin sulfate proteoglycan inhibitor inhibits the expression of a chondroitin sulfate proteoglycan and has the ability to penetrate or permeate the blood-brain-barrier.

[037] In one embodiment, inhibiting fear memory is inhibiting the formation of the extracellular matrix comprising a chondroitin sulfate proteoglycan in the brain. In one embodiment, inhibiting fear memory is inhibiting the risks associated with fear memory such as but not limited to an anxiety disorder and posttraumatic stress disorder. In one embodiment, inhibiting fear memory is reducing the risk for developing a fear memory or an anxiety disorder.

[038] In one embodiment, provided herein is a method for inhibiting fear memory in a subject in need thereof, comprising inhibiting a formation of an extracellular matrix comprising a chondroitin sulfate proteoglycan in the brain of the subject, thereby inhibiting fear memory in a subject in need thereof.

[039] In one embodiment, Xylosine as described herein is protoglican that blocks the attachment of the GAGs to the CSPG's central core protein, which was the primary inhibitory element of CSPGs. In one embodiment, the deposition of CSPGs without GAG side chains inhibited the formation of the CSPG and therefore inhibit the formation of the extracellular matrix as described herein.

[040] In one embodiment, the present invention provides that in mature brain fearful experience can alter the composition of PNNs, especially, chondroitin sulfate proteoglycans (CSPGs) in the amygdala. In one embodiment, the present invention provides that inhibition of CSPG formation using xyloside, provided systemically (such as but not limited to intraperitoneally), significantly inhibited the formation of fear response. In one embodiment, these findings have direct implications in the prevention and treatment anxiety disorders, specifically PTSD.

[041] In one embodiment, manipulation of matrix formation with xyloside (crossed the BBB) immediately after experiencing an event of fear and/or trauma, prevents the recurrence of fear. In one embodiment, application of xyloside is effective after the trauma and not after the memory has been consolidated. In one embodiment, xyloside is: beta-D-xyloside, beta-xyloside, 4-Nitrophenyl-beta-D-xyloside, 3-D-xyloside, 3-O-beta- D-xyloside, 7-O-beta-D-xyloside, alpha-D-xyloside, 3-o-xyloside, or any combination thereof.

[042] In one embodiment, xyloside, according to the methods described herein, is administered at a dose of 0.25 to 50 mg per kg body weight. In one embodiment, xyloside, according to the methods described herein, is administered at a dose of 0. 5 to 20 mg per kg body weight. In one embodiment, xyloside, according to the methods described herein, is administered at a dose of 1 to 15 mg per kg body weight. In one embodiment, xyloside, according to the methods described herein, is administered at a dose of 2 to 10 mg per kg body weight. In one embodiment, xyloside dose as described herein is a daily dose. In one embodiment, xyloside dose as described herein is a single dose. In one embodiment, xyloside dose as described herein is administered 1 to 5 times. In one embodiment, xyloside dose as described herein is administered 1 to 3 times a day.

[043] In one embodiment, provided herein a composition comprising a chondroitin sulfate proteoglycan inhibitor and a MMP inhibitor. In one embodiment, a composition is a pharmaceutical composition. In one embodiment, a "pharmaceutical composition" refers to a preparation of one or more of xyloside, MMP inhibitor or both with other chemical components such as physiologically suitable carriers and excipients. The purpose of a pharmaceutical composition is to facilitate administration of xyloside, MMP inhibitor or both to an organism.

[044] In one embodiment, the present invention provides combined preparations. In one embodiment, "a combined preparation" defines especially a "kit of parts" in the sense that the combination partners (xyloside, MMP inhibitor) as defined above can be dosed independently or by use of different fixed combinations with distinguished amounts of the combination partners i.e., simultaneously, concurrently, separately or sequentially. In some embodiments, the parts of the kit of parts can then, e.g., be administered simultaneously or chronologically staggered, that is at different time points and with equal or different time intervals for any part of the kit of parts. The ratio of the total amounts of the combination partners, in some embodiments, can be administered in the combined preparation. In one embodiment, the combined preparation can be varied, e.g., in order to cope with the needs of a patient subpopulation to be treated or the needs of the single patient which different needs can be due to a particular event/disease, severity, age, sex, or body weight as can be readily made by a person skilled in the art.

[045] In one embodiment, the phrases "physiologically acceptable carrier" and "pharmaceutically acceptable carrier" which be interchangeably used refer to a carrier or a diluent that does not cause significant irritation to an organism and does not abrogate the biological activity and properties of the administered compound. An adjuvant is included under these phrases. In one embodiment, one of the ingredients included in the pharmaceutically acceptable carrier can be for example polyethylene glycol (PEG), a biocompatible polymer with a wide range of solubility in both organic and aqueous media (Mutter et al. (1979).

[046] In one embodiment, "excipient" refers to an inert substance added to a pharmaceutical composition to further facilitate administration of xyloside, MMP inhibitor or both. In one embodiment, excipients include calcium carbonate, calcium phosphate, various sugars and types of starch, cellulose derivatives, gelatin, vegetable oils and polyethylene glycols.

[047] Techniques for formulation and administration of drugs are found in "Remington's Pharmaceutical Sciences" Mack Publishing Co., Easton, PA, latest edition, which is incorporated herein by reference. [048] In one embodiment, suitable routes of administration, for example, include oral, rectal, transmucosal, transnasal, intestinal or parenteral delivery, including intramuscular, subcutaneous and intramedullary injections as well as intrathecal, direct intraventricular, intravenous, intraperitoneal, intranasal, or intraocular injections.

[049] Various embodiments of dosage ranges are contemplated by this invention. The dosage of the polypeptide of the present invention, in one embodiment, is in the range of 0.05-80 mg/day. In another embodiment, the dosage is in the range of 0.05-50 mg/day. In another embodiment, the dosage is in the range of 0.1-20 mg/day. In another embodiment, the dosage is in the range of 0.1-10 mg/day. In another embodiment, the dosage is in the range of 0.1-5 mg/day. In another embodiment, the dosage is in the range of 0.5-5 mg/day. In another embodiment, the dosage is in the range of 0.5-50 mg/day. In another embodiment, the dosage is in the range of 5-80 mg/day. In another embodiment, the dosage is in the range of 35-65 mg/day. In another embodiment, the dosage is in the range of 35-65 mg/day. In another embodiment, the dosage is in the range of 20-60 mg/day. In another embodiment, the dosage is in the range of 40-60 mg/day. In another embodiment, the dosage is in a range of 45-60 mg/day. In another embodiment, the dosage is in the range of 40-60 mg/day. In another embodiment, the dosage is in a range of 60-120 mg/day. In another embodiment, the dosage is in the range of 120-240 mg/day. In another embodiment, the dosage is in the range of 40-60 mg/day. In another embodiment, the dosage is in a range of 240-400 mg/day. In another embodiment, the dosage is in a range of 45-60 mg/day. In another embodiment, the dosage is in the range of 15-25 mg/day. In another embodiment, the dosage is in the range of 5-10 mg/day. In another embodiment, the dosage is in the range of 55-65 mg/day.

[050] In one embodiment, the dosage is 20 mg/day. In another embodiment, the dosage is 30 mg/day. In another embodiment, the dosage is 40 mg/day. In another embodiment, the dosage is 50 mg/day. In another embodiment, the dosage is 60 mg/day. In another embodiment, the dosage is 70 mg/day. In another embodiment, the dosage is 80 mg/day. In another embodiment, the dosage is 90 mg/day. In another embodiment, the dosage is 100 mg/day.

[051] In one embodiment, dosage is a MMP inhibitor dosage. In one embodiment, dosage is xyloside dosage, matrix metalloproteinase (MMP) inhibitor dosage, or a combination of xyloside and a MMP inhibitor dosage.

[052] Oral administration, in one embodiment, comprises a unit dosage form comprising tablets, capsules, lozenges, chewable tablets, suspensions, emulsions and the like. Such unit dosage forms comprise a safe and effective amount of the desired compound, or compounds, each of which is in one embodiment, from about 0.7 or 3.5 mg to about 280 mg/70 kg, or in another embodiment, about 0.5 or 10 mg to about 210 mg/70 kg. The pharmaceutically-acceptable carriers suitable for the preparation of unit dosage forms for peroral administration are well-known in the art. In some embodiments, tablets typically comprise conventional pharmaceutically-compatible adjuvants as inert diluents, such as calcium carbonate, sodium carbonate, mannitol, lactose and cellulose; binders such as starch, gelatin and sucrose; disintegrants such as starch, alginic acid and croscarmelose; lubricants such as magnesium stearate, stearic acid and talc. In one embodiment, glidants such as silicon dioxide can be used to improve flow characteristics of the powder-mixture. In one embodiment, coloring agents, such as the FD&C dyes, can be added for appearance. Sweeteners and flavoring agents, such as aspartame, saccharin, menthol, peppermint, and fruit flavors, are useful adjuvants for chewable tablets. Capsules typically comprise one or more solid diluents disclosed above. In some embodiments, the selection of carrier components depends on secondary considerations like taste, cost, and shelf stability, which are not critical for the purposes of this invention, and can be readily made by a person skilled in the art.

[053] In one embodiment, the oral dosage form comprises predefined release profile. In one embodiment, the oral dosage form of the present invention comprises an extended release tablets, capsules, lozenges or chewable tablets. In one embodiment, the oral dosage form of the present invention comprises a slow release tablets, capsules, lozenges or chewable tablets. In one embodiment, the oral dosage form of the present invention comprises an immediate release tablets, capsules, lozenges or chewable tablets. In one embodiment, the oral dosage form is formulated according to the desired release profile of the xyloside, MMP inhibitor or both as known to one skilled in the art.

[054] Peroral compositions, in some embodiments, comprise liquid solutions, emulsions, suspensions, and the like. In some embodiments, pharmaceutically-acceptable carriers suitable for preparation of such compositions are well known in the art. In some embodiments, liquid oral compositions comprise from about 0.012% to about 0.933% of the desired compound or compounds, or in another embodiment, from about 0.033% to about 0.7%.

[055] In some embodiments, compositions for use in the methods of this invention comprise solutions or emulsions, which in some embodiments are aqueous solutions or emulsions comprising a safe and effective amount of the compounds of the present invention and optionally, other compounds, intended for topical intranasal administration. In some embodiments, h compositions comprise from about 0.01% to about 10.0% w/v of a subject compound, more preferably from about 0.1% to about 2.0, which is used for systemic delivery of the compounds by the intranasal route. [056] In another embodiment, the pharmaceutical compositions are administered by intravenous, intra-arterial, or intramuscular injection of a liquid preparation. In some embodiments, liquid formulations include solutions, suspensions, dispersions, emulsions, oils and the like. In one embodiment, the pharmaceutical compositions are administered intravenously, and are thus formulated in a form suitable for intravenous administration. In another embodiment, the pharmaceutical compositions are administered intra- arterially, and are thus formulated in a form suitable for intra-arterial administration. In another embodiment, the pharmaceutical compositions are administered intramuscularly, and are thus formulated in a form suitable for intramuscular administration. [057] Further, in another embodiment, the pharmaceutical compositions are administered topically to body surfaces, and are thus formulated in a form suitable for topical administration. Suitable topical formulations include gels, ointments, creams, lotions, drops and the like. For topical administration, the compounds of the present invention are combined with an additional appropriate therapeutic agent or agents, prepared and applied as solutions, suspensions, or emulsions in a physiologically acceptable diluent with or without a pharmaceutical carrier.

[058] In one embodiment, pharmaceutical compositions of the present invention are manufactured by processes well known in the art, e.g., by means of conventional mixing, dissolving, granulating, dragee-making, levigating, emulsifying, encapsulating, entrapping or lyophilizing processes.

[059] In one embodiment, pharmaceutical compositions for use in accordance with the present invention is formulated in conventional manner using one or more physiologically acceptable carriers comprising excipients and auxiliaries, which facilitate processing of xyloside, MMP inhibitor or both into preparations which, can be used pharmaceutically. In one embodiment, formulation is dependent upon the route of administration chosen. [060] In one embodiment, injectables, of the invention are formulated in aqueous solutions. In one embodiment, injectables, of the invention are formulated in physiologically compatible buffers such as Hank's solution, Ringer's solution, or physiological salt buffer. In some embodiments, for transmucosal administration, penetrants appropriate to the barrier to be permeated are used in the formulation. Such penetrants are generally known in the art.

[061] In one embodiment, the preparations described herein are formulated for parenteral administration, e.g., by bolus injection or continuous infusion. In some embodiments, formulations for injection are presented in unit dosage form, e.g., in ampoules or in multidose containers with optionally, an added preservative. In some embodiments, compositions are suspensions, solutions or emulsions in oily or aqueous vehicles, and contain formulatory agents such as suspending, stabilizing and/or dispersing agents.

[062] The compositions also comprise, in some embodiments, preservatives, such as benzalkonium chloride and thimerosal and the like; chelating agents, such as edetate sodium and others; buffers such as phosphate, citrate and acetate; tonicity agents such as sodium chloride, potassium chloride, glycerin, mannitol and others; antioxidants such as ascorbic acid, acetylcystine, sodium metabisulfote and others; aromatic agents; viscosity adjusters, such as polymers, including cellulose and derivatives thereof; and polyvinyl alcohol and acid and bases to adjust the pH of these aqueous compositions as needed. The compositions also comprise, in some embodiments, local anesthetics or other actives. The compositions can be used as sprays, mists, drops, and the like.

[063] In some embodiments, pharmaceutical compositions for parenteral administration include aqueous solutions of the xyloside, MMP inhibitor or both in water-soluble form. Additionally, suspensions of xyloside, MMP inhibitor or both, in some embodiments, are prepared as appropriate oily or water based injection suspensions. Suitable lipophilic solvents or vehicles include, in some embodiments, fatty oils such as sesame oil, or synthetic fatty acid esters such as ethyl oleate, triglycerides or liposomes. Aqueous injection suspensions contain, in some embodiments, substances, which increase the viscosity of the suspension, such as sodium carboxymethyl cellulose, sorbitol or dextran. In another embodiment, the suspension also contains suitable stabilizers or agents which increase the solubility of xyloside, MMP inhibitor or both to allow for the preparation of highly concentrated solutions.

[064] In another embodiment, the pharmaceutical composition delivered in a controlled release system is formulated for intravenous infusion, implantable osmotic pump, transdermal patch, liposomes, or other modes of administration. In one embodiment, a pump is used (see Langer, supra; Sefton, CRC Crit. Ref. Biomed. Eng. 14:201 (1987); Buchwald et al., Surgery 88:507 (1980); Saudek et al., N. Engl. J. Med. 321:574 (1989). In another embodiment, polymeric materials can be used. In yet another embodiment, a controlled release system can be placed in proximity to the therapeutic target, i.e., the brain, thus requiring only a fraction of the systemic dose (see, e.g., Goodson, in Medical Applications of Controlled Release, supra, vol. 2, pp. 115-138 (1984). Other controlled release systems are discussed in the review by Langer (Science 249: 1527-1533 (1990).

[065] In some embodiments, xyloside, MMP inhibitor or both is/are in powder form for constitution with a suitable vehicle, e.g., sterile, pyrogen-free water based solution, before use. Compositions are formulated, in some embodiments, for atomization and inhalation administration. In another embodiment, compositions are contained in a container with attached atomizing means.

[066] In one embodiment, the preparation of the present invention is formulated in rectal compositions such as suppositories or retention enemas, using, e.g., conventional suppository bases such as cocoa butter or other glycerides. [067] In some embodiments, pharmaceutical compositions suitable for use in context of the present invention include compositions wherein xyloside, MMP inhibitor or both are contained in an amount effective to achieve the intended purpose. In some embodiments, a therapeutically effective amount means an amount of xyloside, MMP inhibitor or both effective to prevent, alleviate or ameliorate symptoms of disease or prolong the survival of the subject being treated.

[068] In one embodiment, determination of a therapeutically effective amount is well within the capability of those skilled in the art.

[069] In some embodiments, compounds modified by the covalent attachment of water- soluble polymers such as polyethylene glycol, copolymers of polyethylene glycol and polypropylene glycol, carboxymethyl cellulose, dextran, polyvinyl alcohol, polyvinylpyrrolidone or polyproline. In another embodiment, the modified compounds exhibit substantially longer half -lives in blood following intravenous injection than do the corresponding unmodified compounds. In one embodiment, modifications also increase the compound's solubility in aqueous solution, eliminate aggregation, enhance the physical and chemical stability of the compound, and greatly reduce the immunogenicity and reactivity of the compound. In another embodiment, the desired in vivo biological activity is achieved by the administration of such polymer-compound abducts less frequently or in lower doses than with the unmodified compound. [070] In one embodiment, toxicity and therapeutic efficacy of xyloside, MMP inhibitor or both described herein can be determined by standard pharmaceutical procedures in vitro, in cell cultures or experimental animals. In one embodiment, the data obtained from these in vitro and cell culture assays and animal studies can be used in formulating a range of dosage for use in human. In one embodiment, the dosages vary depending upon the dosage form employed and the route of administration utilized. In one embodiment, the exact formulation, route of administration and dosage can be chosen by the individual physician in view of the patient's condition. [See e.g., Fingl, et al., (1975) "The Pharmacological Basis of Therapeutics", Ch. 1 p. l].

[071] In one embodiment, depending on the severity and responsiveness of the condition to be treated, dosing can be of a single or a plurality of administrations, with course of treatment lasting from several days to several weeks or until cure is effected or diminution of the disease state is achieved.

[072] In one embodiment, the amount of a composition to be administered will, of course, be dependent on the subject being treated, the severity of the affliction, the manner of administration, the judgment of the prescribing physician, etc.

[073] In one embodiment, compositions including the preparation of the present invention formulated in a compatible pharmaceutical carrier are also be prepared, placed in an appropriate container, and labeled for treatment of an indicated condition.

[074] In one embodiment, compositions of the present invention are presented in a pack or dispenser device, such as an FDA approved kit, which contains one or more unit dosage forms containing xyloside, MMP inhibitor or both. In one embodiment, the pack, for example, comprise metal or plastic foil, such as a blister pack. In one embodiment, the pack or dispenser device is accompanied by instructions for administration. In one embodiment, the pack or dispenser is accommodated by a notice associated with the container in a form prescribed by a governmental agency regulating the manufacture, use or sale of pharmaceuticals, which notice is reflective of approval by the agency of the form of the compositions or human or veterinary administration. Such notice, in one embodiment, is labeling approved by the U.S. Food and Drug Administration for prescription drugs or of an approved product insert. EXAMPLES

Material and methods

[075] Subjects: Male C57BL/6J mice, weighing 24-28 g upon arrival, all experiments approved and carried out in compliance with guidelines established by the Technion Animal Care and Use Committee (ACUC).

[076] Xyloside: Xyloside (4-methylumbelliferyl-P-D-xylopyranoside; Sigma- Aldrich, M7008) was dissolved in Saline. Prior to injection, the xyloside solution was diluted to a final concentration of 133.3ηι§/100μ1/πιου86 in saline. Beta-D- xyloside was used to inhibit CSPG-matrix formation. [077] Beta-D- xyloside crossed the blood-brain barrier (BBB) and thus was administered systemically. Xylosine is protoglican that blocks the attachment of the GAGs to the CSPG's central core protein, which is the primary inhibitory element of CSPGs. Thus, the deposition of CSPGs without GAG side chains inhibited the formation of the CSPG and therefore inhibit the formation of the matrix. [078] Fear-conditioning: Analysis of fear memory and extinction learning was based on freezing responses observed after the mice are returned to the conditioning chamber. Mice were placed in a different chamber and exposed to the conditioning tone. The freezing duration during tone presentation was calculated and compared to freezing during the period in the absence of the tone. This represented the cue conditioning. To evaluate extinction, the mice were repeatedly exposed to the tone and the decrease in the freezing duration was monitored. Mice were re-exposed to the conditioning chamber without shock presentation, and freezing after several exposures to the chamber was monitored. This setup was used as an indication of the stability of the memory.

[079] Experimental design: On day 1, C57BL/6 mice were placed in a conditioning chamber. After 1 min of habituation the CS tone was activated for 8 seconds. 2 second later a footshock (0.25 mA, 2 seconds), followed. This procedure was repeated for a total of three shocks. Then Control mice received an injection of saline and study mice received the Xyloside injection. The mice were then returned to their home cages.

[080] On day 2, mice were placed in a testing chamber. After 1 minute of habituation (baseline freezing was not different between the control and the Xyloside group), followed by 8 seconds of tone, then repeated twice more, for a total of three cycles. A video camera recorded all behavior during this test for off-line scoring at a later time.

EXAMPLE 1: Fearful experience can alter the composition of PNNs

[081] The formation of perineuronal nets (PNNs) in the mature brain is crucial for stabilization of memory. In this experiment, it was shown that even in mature brain fearful experience can alter the composition of PNNs, especially, chondroitin sulfate proteoglycans (CSPGs) in the amygdala. It was further demonstrated that inhibition of CSPG formation using xyloside, injected even to the periphery (intraperitonealy), significantly inhibited the formation of fear response. These findings have direct implications in the prevention and treatment anxiety disorders, specifically PTSD.

[082] The changes in CSPG levels were demonstrated at the protein levels (Mass spectrometry). However, the functional relevance of these changes was further explored. Mice were exposed to fear-conditioning and injected them with xyloside to the periphery (intraperitonealy). Xyloside inhibits CSPG formation immediately following fear conditioning but it is not expected to affect existing CSPGs.

[083] It was discovered that manipulation of matrix formation immediately after fear conditioning, prevents the recurrence of fear. To inhibit matrix formation, the CSPG formation inhibitor (beta-D- xyloside) was used. Interestingly, beta-D-Xyloside crossed the BBB and therefore was administered systemically. Surprisingly, animals treated with beta-D-xyloside administered immediately after fear induction (conditioning session), did not show fear response (freezing) on the following day (see Fig. 1).

[084] An additional experiment, included beta-D- xyloside administration 24 hours after the fear conditioning or immediately before the mice were reintroduced to fear. It was found that these treatments with beta-D- xyloside resulted with no effect on the fear response and mice in both groups presented similar levels of freezing. Thus, application of beta-D- xyloside was effective only after the trauma and not after the memory has been consolidated, at this later point, beta-D- xyloside did not have any effect on behavior. This further supports that the effects of beta-D- xyloside on the attenuation of fear response is during the consolidation process.

[085] In conclusion: (1) fear conditioning affects matrix formation in a sleep independent manner; and (2) inhibition of matrix formation immediately after the fear conditioning, prevents the manifestation of fear-related behavior (freezing) twenty-four hours later.

EXAMPLE 2: Xylosie injection directly into the amygdala

[086] This experiment was conducted on 10 mice. The control group was injected with saline (ImicroL) and the study group was injected with Xyloside at concentration 5mg/microL. The injections were done via a cannula implanted in the Amygdala and allowed to deliver the compound directly to the amygdala in behaving mice.

[087] Saline / Xyloside injection was performed half an hour after the mice was exposed to the fear conditioning. The cannula guide tubes were placed above the left and right BLA (AP, 1.22 mm; ML, 2.7 mm; DV, 4.1 mm).

Freezing behavior test after fear conditioning- attenuating consolidated fear memory.

[088] Percent of freezing duration in response to the tone presentation. Two conditions were compared: (1) mice treated with directly injections of Xyloside into the Amygdala (2) mice injections saline and received control group. The groups showed significant (*p<0.005) difference in terms (see table 1 and figure 3).

Table 1

Open field test after 48 hours from the injection

[089] Results of neurobehavioral OFT testing in mice that were exposed to fear- conditioning and injected them with xyloside or saline directly into the amygdala, (figure 4 A) Time in the center field (figure 4B) Time in border field. The groups showed no significant difference in terms of the length of time spent in the central and border square (see table 2 and 3, respectively).

Table 2

Table 3

Claims

CLAIMS What is claimed is:

1. A method for inhibiting a formation of an extracellular matrix comprising a chondroitin sulfate proteoglycan in a subject in need thereof, comprising, administering to said subject a composition comprising xyloside, matrix metalloproteinase (MMP) inhibitor, or a combination of xyloside and a MMP inhibitor, thereby inhibiting the formation of an extracellular matrix comprising a chondroitin sulfate proteoglycan in a subject in need thereof.

2. The method of claim 1, wherein saidxyloside is: beta-D-xyloside, beta-xyloside, 4- Nitrophenyl-beta-D-xyloside, 3-D-xyloside, 3-O-beta-D-xyloside, 7-O-beta-D-xyloside, alpha-D-xyloside, 3-o-xyloside, or any combination thereof.

3. The method of anyone of claims 1 and 2, wherein said formation is within the brain amygdala, hippocampus, or any combination thereof.

4. The method of anyone of claims 1 to 3, wherein said inhibiting the formation of the extracellular matrix comprising a chondroitin sulfate proteoglycan is affecting memory.

5. The method of anyone of claims 1 to 4, wherein said inhibiting the formation of the extracellular matrix comprising a chondroitin sulfate proteoglycan is inhibiting fear memory or an anxiety disorder.

6. The method of anyone of claims 1 to 4, wherein said inhibiting the formation of the extracellular matrix comprising a chondroitin sulfate proteoglycan is reducing the risk for developing a fear memory or an anxiety disorder.

7. The method of anyone of claims 1 to 4, wherein said inhibiting the formation of the extracellular matrix comprising a chondroitin sulfate proteoglycan is inhibiting, treating, ameliorating, or any combination thereof of a posttraumatic stress disorder.

8. The method of anyone of claims 1 to 4, wherein said inhibiting the formation of the extracellular matrix comprising a chondroitin sulfate proteoglycan is reducing the risk for developing a posttraumatic stress disorder.

9. The method of anyone of claims 1 to 6, wherein said subject is at risk of developing a fear memory or an anxiety disorder.

10. The method of anyone of claims 1 to 9, wherein said MMP inhibitor is MMP-2 inhibitor, MMP-9 inhibitor, or a combination thereof.

11. The method of anyone of claims 1 to 10, wherein said MMP inhibitor is GM6001.

12. A method for inhibiting fear memory, trauma or an anxiety disorder in a subject in need thereof, comprising administering to said subject a chondroitin sulfate proteoglycan inhibitor, matrix metalloproteinase (MMP) inhibitor, or a combination of a chondroitin sulfate proteoglycan inhibitor and a MMP inhibitor, thereby inhibiting fear memory, trauma or an anxiety disorder in a subject in need thereof.

13. The method of claim 12, wherein said chondroitin sulfate proteoglycan inhibitor inhibits the expression of a chondroitin sulfate proteoglycan.

14. The method of anyone of claims 12 and 13, wherein said inhibiting fear memory is inhibiting the formation of the extracellular matrix comprising a chondroitin sulfate proteoglycan in the brain.

15. The method of anyone of claim 14, wherein said brain is amygdala, hippocampus, or both.

16. The method of anyone of claims 12 to 15, wherein said inhibiting fear memory is reducing the risk for developing a fear memory or an anxiety disorder.

17. The method of anyone of claims 12 to 15, wherein said inhibiting fear memory is inhibiting, treating, ameliorating, or any combination thereof of a posttraumatic stress disorder.

18. The method of claim 17, wherein said inhibiting a posttraumatic stress disorder is reducing the risk for developing a posttraumatic stress disorder.

19. The method of anyone of claims 12 to 15, wherein said subject is at risk of developing a fear memory or an anxiety disorder.

20. The method of anyone of claims 12 to 19, wherein said MMP inhibitor is MMP-2 inhibitor, MMP-9 inhibitor, or a combination thereof.

21. The method of anyone of claims 12 to 19, wherein said MMP inhibitor is GM6001.

22. The method of anyone of claims 12 to 21, wherein said chondroitin sulfate proteoglycan inhibitor is xyloside.

23. The method of claim 22, wherein saidxyloside is: beta-D-xyloside, beta-xyloside, 4- Nitrophenyl-beta-D-xyloside, 3-D-xyloside, 3-O-beta-D-xyloside, 7-O-beta-D-xyloside, alpha-D-xyloside, 3-o-xyloside, or any combination thereof.

24. A method for inhibiting fear memory in a subject in need thereof, comprising inhibiting a formation of an extracellular matrix comprising a chondroitin sulfate proteoglycan in the brain of said subject, thereby inhibiting fear memory in a subject in need thereof.

25. A composition comprising a chondroitin sulfate proteoglycan inhibitor and a MMP inhibitor.

26. The composition of claim 24, wherein said MMP inhibitor is MMP-2 inhibitor, MMP-9 inhibitor, or a combination thereof.

27. The composition of anyone of claims 24 and 25, wherein said MMP inhibitor is GM6001.

28. The composition of anyone of claims 24 to 26, wherein said chondroitin sulfate proteoglycan inhibitor is xyloside.

29. The composition of claim 28, wherein saidxyloside is: beta-D-xyloside, beta- xyloside, 4-Nitrophenyl-beta-D-xyloside, 3-D-xyloside, 3-O-beta-D-xyloside, 7-O-beta- D-xyloside, alpha-D-xyloside, 3-o-xyloside, or any combination thereof.