EP1377287A2 - Proanthocyanidines destinees au traitement des amyloses et des maladies induites par alpha-synucleine - Google Patents

Proanthocyanidines destinees au traitement des amyloses et des maladies induites par alpha-synucleine

Info

Publication number
EP1377287A2
EP1377287A2 EP02719009A EP02719009A EP1377287A2 EP 1377287 A2 EP1377287 A2 EP 1377287A2 EP 02719009 A EP02719009 A EP 02719009A EP 02719009 A EP02719009 A EP 02719009A EP 1377287 A2 EP1377287 A2 EP 1377287A2
Authority
EP
European Patent Office
Prior art keywords
epicatechin
amyloid
proanthocyanidin
disease
proanthocyanidins
Prior art date
Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
Withdrawn
Application number
EP02719009A
Other languages
German (de)
English (en)
Other versions
EP1377287A4 (fr
Inventor
Alan D. Snow
Geraldo M. Castillo
Paula Y. Choi
Beth P. Nguyen
Current Assignee (The listed assignees may be inaccurate. Google has not performed a legal analysis and makes no representation or warranty as to the accuracy of the list.)
ProteoTech Inc
Original Assignee
ProteoTech Inc
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Priority claimed from US09/938,987 external-priority patent/US6607758B2/en
Priority claimed from US10/053,625 external-priority patent/US6929808B2/en
Application filed by ProteoTech Inc filed Critical ProteoTech Inc
Publication of EP1377287A2 publication Critical patent/EP1377287A2/fr
Publication of EP1377287A4 publication Critical patent/EP1377287A4/fr
Withdrawn legal-status Critical Current

Links

Classifications

    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/21—Esters, e.g. nitroglycerine, selenocyanates
    • A61K31/215—Esters, e.g. nitroglycerine, selenocyanates of carboxylic acids
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/33—Heterocyclic compounds
    • A61K31/335—Heterocyclic compounds having oxygen as the only ring hetero atom, e.g. fungichromin
    • A61K31/35—Heterocyclic compounds having oxygen as the only ring hetero atom, e.g. fungichromin having six-membered rings with one oxygen as the only ring hetero atom
    • A61K31/352—Heterocyclic compounds having oxygen as the only ring hetero atom, e.g. fungichromin having six-membered rings with one oxygen as the only ring hetero atom condensed with carbocyclic rings, e.g. methantheline 
    • A61K31/353—3,4-Dihydrobenzopyrans, e.g. chroman, catechin
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P25/00—Drugs for disorders of the nervous system
    • A61P25/14—Drugs for disorders of the nervous system for treating abnormal movements, e.g. chorea, dyskinesia
    • A61P25/16—Anti-Parkinson drugs
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P25/00—Drugs for disorders of the nervous system
    • A61P25/28—Drugs for disorders of the nervous system for treating neurodegenerative disorders of the central nervous system, e.g. nootropic agents, cognition enhancers, drugs for treating Alzheimer's disease or other forms of dementia
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P29/00—Non-central analgesic, antipyretic or antiinflammatory agents, e.g. antirheumatic agents; Non-steroidal antiinflammatory drugs [NSAID]
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P3/00—Drugs for disorders of the metabolism
    • A61P3/08—Drugs for disorders of the metabolism for glucose homeostasis
    • A61P3/10—Drugs for disorders of the metabolism for glucose homeostasis for hyperglycaemia, e.g. antidiabetics
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P35/00—Antineoplastic agents
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P35/00—Antineoplastic agents
    • A61P35/02—Antineoplastic agents specific for leukemia

Definitions

  • the invention relates to methods and compositions for treatment and prevention of a ⁇ rjU ⁇ id, NAC (i.e. non-amyloid component) and ⁇ -synuclein diseases, such as Alzheimer's disease and Parkinson ' s disease, and to method of isolation of new compounds for the same; particularly it relates to polyphenolic compositions and methods of using same to treat these same diseases; more particularly it relates to proanthocyanidin and related compounds for treatment and prevention of amyloid, NAC and ⁇ -synuclein diseases.
  • NAC i.e. non-amyloid component
  • ⁇ -synuclein diseases such as Alzheimer's disease and Parkinson ' s disease
  • Alzheimer's disease is characterized by the accumulation of a 39-43 amino acid peptide termed the beta-amyloid protein or A ⁇ , in a fibrillar form, existing as extracellular amyloid plaques and as amyloid within the walls of cerebral blood vessels.
  • a ⁇ beta-amyloid protein
  • Fibrillar A ⁇ amyloid deposition in Alzheimer's disease is believed to be detrimental to the patient and eventually leads to toxicity and neuronal cell death, characteristic hallmarks of Alzheimer's disease.
  • Accumulating evidence implicates amyloid, and more specifically, the formation, deposition, accumulation and/or persistence of A ⁇ fibrils, as a major causative factor of Alzheimer's disease pathogenesis.
  • Alzheimer's disease a number of other amyloid diseases involve accumulation of A ⁇ fibrils, including Down's syndrome, disorders involving congophilic angiopathy, hereditary cerebral hemorrhage of the Dutch type, and inclusion body myositosis.
  • Parkinson's disease is another human disorder characterized by the formation, deposition, accumulation and/or persistence of abnormal fibrillar protein deposits that demonstrate many of the characteristics of amyloid.
  • an accumulation of cytoplasmic Lewy bodies consisting of filaments of ⁇ -synuclein/NAC are believed important in the pathogenesis and as therapeutic targets.
  • New agents or compounds able to inhibit ⁇ - synuclein/NAC formation, deposition, accumulation and/or persistence, or disrupt pre-formed ⁇ -synuclein/NAC fibrils (or portions thereof) are regarded as potential therapeutics for the treatment of Parkinson's disease.
  • amyloid deposition A variety of other human diseases also demonstrate amyloid deposition and usually involve systemic organs (i.e. organs or tissues lying outside the central nervous system), with the amyloid accumulation leading to organ dysfunction or failure. These amyloid diseases (discussed below) leading to marked amyloid accumulation in a number of different organs and tissues are known as systemic amyloidoses.
  • systemic amyloidoses single organs may be affected such as the pancreas in 90% of patients with type 2 diabetes.
  • the beta-cells in the islets of Langerhans in pancreas are believed to be destroyed by the accumulation of fibrillar amyloid deposits consisting primarily of a protein known as islet amyloid polypeptide (IAPP).
  • IAPP islet amyloid polypeptide
  • amyloid accumulation is believed to lead to new effective treatments for type 2 diabetes.
  • Alzheimer's disease, Parkinson's and "systemic" amyloid diseases there is currently no cure or effective treatment, and the patient usually dies within 3 to 10 years from disease onset.
  • amyloid diseases include, but are not limited to, the amyloid associated with Alzheimer's disease, Down's syndrome, hereditary cerebral hemorrhage with amyloidosis of the Dutch type, and inclusion body myositosis (Askanas et al, Ann. Neurol.
  • beta-amyloid protein or A ⁇ amyloid protein
  • a ⁇ amyloid associated with chronic inflammation
  • various forms of malignancy and Familial Mediterranean Fever wherein the specific amyloid is referred to as AA amyloid or inflammation-associated amyloidosis
  • AL amyloid amyloid associated with multiple myeloma and other B-cell dyscrasias
  • AL amyloid amyloid associated with type II diabetes
  • the specific amyloid protein is referred to as amylin or islet amyloid polypeptide
  • the amyloid associated with the prion diseases including Creutzfeldt- Jakob disease, Gerstmann-Straussler syndrome, kuru and animal scrapie wherein the specific amyloid is referred to as PrP amyloid
  • the amyloid associated with long-term hemodialysis and carpal tunnel syndrome wherein the specific amyloid is referred to as beta 2
  • the ⁇ -synuclein protein which forms fibrils, and is Congo red and Thioflavin S positive is found as part of Lewy bodies in the brains of patients with Parkinson's disease, Lewy body disease (Lewy in Handbuch der Neurologie. M. Lewandowski, ed., Springer, Berline pp.920-933, 1912; Pollanen et al, J. Neuropath. Exp. Neurol. 52:183-191, 1993; Spillantini et al, Proc. Natl. Acad. Sci. USA 95:6469-6473, 1998; Arai et al, Neurosc. Lett.
  • Parkinson's disease due to the fact that fibrils develop in the brains of patients with this disease (which are Congo red and Thioflavin S positive, and which contain predominant beta-pleated sheet secondary structure), should be regarded as a disease that also displays the characteristics of an amyloid-like disease.
  • Alzheimer's disease, Parkinson's disease, type II diabetes, systemic AA amyloidosis, and other amyloidoses are urgent sought.
  • Polyphenols are an incredibly diverse group of compounds (Ferreira et al, Tetrahedron 48:1743-1803,1992) that widely occur in a variety of plants, some of which enter into our food chain. Although some of the polyphenols are considered to be nonnutritive, interest in these compounds has arisen because of their possible beneficial effects for health.
  • quercetin a flavanoid
  • Catechin and epicatechin have been shown to inhibit Leukemia virus reverse transcriptase activity (Chu et al, J.
  • proanthocyanidins, and procyanidins particularly epicatechin- epicatechin dimers or trimers or other oligomers, epicatechin-catechin dimers or the like, or analogs or derivatives thereof, have any benefit for the inhibition of amyloid or ⁇ -synuclein/ NAC fibril formation, and/or cause a disruption of pre-formed amyloid or ⁇ -synuclein/NAC fibrils.
  • proanthocyanidins are potent inhibitors of amyloid and ⁇ -synuclein/NAC fibrillogenesis, and cause a potent disruption disassembly of pre-formed fibrils for a variety of amyloid and ⁇ -synuclein diseases are disclosed.
  • Exemplary compounds are identified to serve as potent amyloid fibril inhibiting agents, including procyanidins, such as epicatechin-epicatechin, catechin-epicatechin dimers, epiafzelechin-epicatechin dimers, epicatechin-epicatechin-epicatechin trimers, as well as other epicatechin and/or catechin oligomers for the treatment of amyloid diseases including, but not limited to, Alzheimer's disease, type II diabetes, and systemic AA amyloidosis, as well as inhibiting ⁇ -synuclein or non-amyloid component (NAC) fibril formation for the treatment of Parkinson's and Lewy body disease.
  • procyanidins such as epicatechin-epicatechin, catechin-epicatechin dimers, epiafzelechin-epicatechin dimers, epicatechin-epicatechin-epicatechin trimers, as well as other epicatechin and/or catechin oligomers for the treatment of amyloid diseases including
  • This invention is also directed to methods for inhibiting or eliminating amyloid fibril formation, deposition, accumulation and/or persistence in a number of different amyloid diseases by treatment of patients with proanthocyanidins, such as procyanidins of the A, B and C types, or other monomers, dimers, trimers and multimers of epicatechin and catechin.
  • Exemplary compounds are substituted epicatechin-epicatechin or catechin-epicatechin dimers, such as epicatechin-4 ⁇ — 8-epicatechin or catechin-4 ⁇ — >8-epicatechin, and epiafzelechin-4 ⁇ -»8- epicatechin, or other proanthocyanidin oligomers.
  • amyloid-inhibiting compounds derived from plant material for the therapeutic intervention of Alzheimer's disease, type II diabetes, Parkinson's disease, systemic AA amyloidosis and other disorders involving amyloid fibril accumulation; more particularly, it relates to methods of isolating amyloid- inhibiting compounds from Uncaria tomentosa and related plants, and other known proanthocyanidin producing plants, and to the use of those compounds.
  • Fraction F chlorogenic acid
  • epicatechin C 15 H 14 O 6 ; FW 290.27
  • Fraction J epicatechin
  • fraction H isolated from PTI-777 was a most potent inhibitor of amyloid fibrillogenesis.
  • PTI-777 has the ability to enter the brain as demonstrated by radiolabelling experiments, indicating that it has the potential to be useful as a therapeutic agent for Alzheimer's disease, Parkinson's disease, and other central nervous system disorders involving deposition and accumulation of fibrillar proteins, such as type 2 diabetes and systemic AA amyloidosis.
  • Compound K2 a component of PTI-777, was purified and identified as epicatechin-4 ⁇ — >8- epicatechin-4 ⁇ — >8-epicatechin, also known as procyanidin Cl.
  • Compound Kl a component of PTI-777, was purified and identified as epiafzelechin-4 ⁇ - ⁇ 8-epicatechin.
  • proanthocyanidins as a potent inhibitor of Alzheimer's A ⁇ amyloidosis, Parkinson's disease ⁇ -synuclein/NAC fibrillogenesis, and type II diabetes IAPP fibrillogenesis, is disclosed herein, and supports the conclusion that procyanidins in particular, and proanthocyanidins in general, are useful compounds for the treatment of amyloidosis and related fibrillogenesis associated with Alzheimer's disease, Parkinson's disease, type 2 diabetes, systemic AA amyloidosis and other amyloid diseases.
  • a method of treating a human or other a mammal suffering from, or subject to, an amyloid disease, or any disease characterized by ⁇ -synuclein or NAC fibrillogenesis is disclosed. Any mammal may be the subject or the disease or condition, or simply be a mammal that is subject to the disease or condition.
  • Amyloid disease as used herein includes but is not limited to the various known and disclosed amyloidoses discussed herein.
  • a disclosed treatment for an amyloid disease is intended to cover a like treatment for the corresponding amyloidosis, and vice-versa. The same is true for ⁇ -synuclein diseases.
  • fibrillogenesis refers to the fibril, plaque and tangle-like forming propensities of the various substituent proteins and/or precursor proteins disclosed herein, whether or not any particular degree of fibrillogenesis has progressed, or is expected to progress, to any particular recognized amyloidosis or to an amyloid or ⁇ -synuclein disease.
  • treatment of fibrillogenesis as disclosed is intended to include and cover treatment of any amyloidosis or any amyloid or ⁇ -synuclein disease corresponding to, following from, otherwise related to, that fibrillogenesis.
  • Treatment is also intended in every possible instance to include and cover “in vitro treatment", whether for experimental or screening purposes and the like, and whether or not the in vitro treatment leads to, or is ever intended to lead to, treatment of a like fibrillogenesis, or any amyloid or ⁇ -synuclein disease corresponding to that fibrillogenesis, in a mammalian subject.
  • the method includes administering to the mammal a therapeutically effective amount of any proanthocyanidin, or proanthocyanidin compound, that may be found in the group of proanthocyanidin and proanthocyanidin compounds characterized by either Formula I or Formula II, or both (see Figures 54-56).
  • the group also includes proanthocyanidins characterized by oligomeric combinations of Formula I and Formula II (see Figure 56), and also includes any pharmaceutically acceptable salt of any of the foregoing proanthocyanidins.
  • proanthocyanidins also referred to herein as PA
  • PA include a variety of structural shapes and oligomeric forms.
  • Formulae I and II are intended each to represent one general form of an oligomeric unit effective to make up the various disclosed oligomers.
  • some proanthocyanidin oligomers are well characterized by Formula I, which is to say that the general structure stated by Formula I is a valid generalization of each unit of the olig ⁇ rner.
  • An example of a proanthocyanidin characterized by Formula I is epicatechin-4 ⁇ — >8-epicatechin, a dimer where two epicatechin units, each a particular instance of, and conforming to, Formula I, are joined from the 4 carbon atom of one unit, to the 8 carbon atom of the other unit, thus effecting a so-called 4-8 linkage.
  • proanthocyanidin oligomers entirely characterized by Formula II will have all their units joined from the 4 carbon atom of one unit to the 6 carbon atom of the adjacent unit, thus effecting a so-called 4-6 linkage, and so on.
  • Some proanthocyanidins can not be well characterized by only Formula I or Formula IJ, in fact presenting in one unit a Formula I configuration and in another unit a Formula II configuration.
  • a proanthocyanidin having a unit in a 4-6 linkage to a second unit which itself has a 4-8 linkage to a third unit is not strictly either a Formula I or a Formula JI compound, but is actually an oligomeric combination of Formula I and Formula ⁇ , where among other characteristics, each unit may be susceptible of more than one characterization (viz. Formula I or Formula U).
  • the middle of the three units has both it's 4 carbon and 6 carbon link sites filled and is therefore a Formula II unit (it may not be possible to specify with particularity what the first unit is, since as terminal unit it has only one carbon linked); the third unit however has it's 8 carbon link site filled and whether or not it is a terminal unit, it will most likely conform to Formula I (except in the relatively rare occurrence of the third unit itself linking at it's 6 carbon site to a fourth unit, making it a unit with it's 6 and 8 carbon link sites filled - which fits neither formula, though it may be nonetheless a useful compound for treatment).
  • an oligomer with some units 4-6 (or 6-4) linked and some 4-8 (or 8-4 linked) will show units in both Formula I and Formula U configurations, but be neither a Formula I nor a Formula II compound, but will be instead an oligomeric combination of Formula I and Formula II.
  • proanthocyanidins characterized as above, or elsewhere herein, particularly with oligomer units numbering in the range of 2 - 20 that is, where n in either Formula is an integer value from 2 to 20
  • proanthocyanidins not precisely adhering to the above formulaic prescriptions such as the presence of one or more variant linkages, like the aforementioned 6-8 or 8-6 unit linkage, or even a 6-6 unit linkage
  • proanthocyanidins not precisely adhering to the above formulaic prescriptions such as the presence of one or more variant linkages, like the aforementioned 6-8 or 8-6 unit linkage, or even a 6-6 unit linkage
  • R ⁇ and R 2 are independently selected from hydrogen and hydroxy;
  • R 3 is selected from the group consisting of hydrogen, optionally substituted O- glycosyl, -C(O)-(optionally substituted aryl), and -C(O)-(optionally substituted heteroaryl);
  • R 4 is selected from the group consisting of hydrogen, catechin, epicatechin, epiafzelechin, and gallates of catechin and epicatechin.
  • the lines at the 2-, 3- and 4-position denote optional R and S (sometimes and alternatively referred to as ⁇ and ⁇ ) stereochemical configurations. Generally the configuration at the 4-position is trans to the configuration at the 3-position.
  • the lines at the 4- and 8-positions in Formula I and at the 4- and 6- positions in Formula II denote possible oligomer bonds between individual units as earlier discussed, and the each of the substitutions at R l5 R 2 , R 3 , and R 4 , and each of the configurations at the 2-, 3-, and 4-positions, and each of the oligomer bond configurations of 4-8 and 4-6 are independently selected for each individual unit and may be different for unit in the oligomer series of units, though often the units of shorter oligomers are homogenous with one another.
  • Preferred oligomers with have from 2 to 5 units, or even 2-3 units.
  • the chiral configuration at each 2- carbon position is preferably R as opposed to S.
  • some or all of the units have an R 3 that is either hydrogen, 2,3-dihydroxybenzoyl, 3,4- dihydroxybenzoyl; 2,3,4-trihydroxybenzoyl, or 3,4,5-trihydroxybenzoyl, and preferably each R 3 is hydrogen and each R t is hydroxy and each R 2 is hydrogen.
  • R 3 may also be an optionally substituted O-glycosyl.
  • the method includes the step of administering to the subject a therapeutically effective amount of a proanthocyanidin.
  • the proanthocyanidin is preferably a procyanidin oligomer having from 2 to 20, and more preferably 2-5, flavanoid units.
  • Each flavanoid unit can advantageously be one of the catechins, including catechin, epicatechin, epiafzelechin, gallocatechin, galloepicatechin, epigallocatechin and the gallates of the catechins.
  • the flavanoid unit can also be one of the flavanols, flavonols, flavandiols, leucocyanidins, or anthocyanidins.
  • amyloid disease to be treated can be Alzheimer's disease, Down's syndrome, hereditary cerebral hemorrhage with amyloidosis of the Dutch type, inclusion body myositosis, the amyloidosis of chronic inflammation, the amyloidosis of malignancy and
  • Familial Mediterranean Fever the amyloidosis of multiple myeloma and B-cell dyscraisa, the amyloidosis of type 2 diabetes, the amyloidosis of prion diseases, Creutzfeldt- Jakob disease, Gerstmann-Straussler syndrome, kuru, scrapie, mad cow disease, the amyloidosis associated with long-term hemodialysis, the amyloidosis with carpal tunnel syndrome, senile cardiac amyloidosis, Familial Amyloidotic Polyneuropathy, the amyloidosis associated with endocrine tumors, systemic AA amyloidosis, AL amyloidosis, A ⁇ amyloidosis or PrP amyloidosis, but particularly Alzheimer's disease.
  • the particular ⁇ -synuclein or NAC fibrillogenesis to be treated can be the fibrillogenesis associated with Lewy body disease, Parkinson's disease or multiple system atrophy.
  • a method of treatment of amyloid, ⁇ -synuclein or NAC fibrillogenesis, in an in vitro environment is also disclosed.
  • the method includes the step of administering into the in vitro environment a therapeutically effective amount of a proanthocyanidin.
  • the proanthocyanidin is a procyanidin which is an oligomer of any or all of epicatechin, catechin, epiafzelechin, epicatechin gallates or catechin gallates.
  • the procyanidin may advantageously be a procyanidin that is an A, B or C type procyanidin.
  • the procyanidin is preferably a dimer or trimer of epicatechin and/or catechin units, such as the dimers of the type Bl, B2, B3, B4, B5, B6, B7, and B8 type procyanidins.
  • the procyanidin dimer is epicatechin-4 ⁇ 8-epicatechin; in another embodiment, the procyanidin dimer is catechin-4 ⁇ — >8-epicatechin; in still another, the procyanidin is the epicatechin trimer epicatechin-4 ⁇ — >8-epicatechin-4 ⁇ — >8-epicatechin; in yet another embodiment the procyanidin is the dimer epiafzelechin-4 ⁇ 8-epicatechin.
  • the method may also include an administration step to deliver the procyanidin to the subject by way of oral administration, parenteral injection, intraperitoneal injection, intravenous injection, subcutaneous injection, intramuscular injection, topical administration, or aerosol spray administration.
  • a pharmaceutical composition or agent is also disclosed. It is a therapeutically effective amount of a proanthocyanidin (PA) together with a pharmaceutically acceptable carrier, diluent, or excipient, or the like.
  • PA proanthocyanidin
  • the therapeutic amount of the PA is selected for efficacy in treating an amyloid, ⁇ -synuclein or NAC fibrillogenesis in a mammalian subject.
  • Disclosed compositions are delivered in therapeutic dosages in the range of about 10 mg/kg to 1,000 mg/kg of body weight of the subject, and preferably in the range of about 10 mg/kg to 100 mg/kg of body weight of the subject.
  • the proanthocyanidin is preferably epicatechin or one or more of the dimers and trimers of epicatechin and catechin, or a mixture thereof, as well as the pharmaceutically acceptable analogs and derivatives thereof.
  • Preferred proanthocyanidins are the procyanidin dimer epicatechin-4 ⁇ 8-epicatechin, the procyanidin dimer catechin-4 ⁇ 8-epicatechin, the procyanidin dimer epiafzelechin-4 ⁇ — >8-epicatechin, and the procyanidin trimer epicatechin- 4 ⁇ -»8-epicatechin-4 ⁇ — >8-epicatechin.
  • the composition is a mixture of two or more proanthocyanidins
  • they may also be advantageously selected from epicatechin and the dimers and trimers of epicatechin and catechin and the pharmaceutically acceptable analogs and derivatives of these compounds.
  • a mixture of two or more procyanidins such as the dimers and trimers of epicatechin and catechin and/or their pharmaceutically acceptable analogs and derivatives may be employed to therapeutic advantage, and in particular, a mixture of two or more proanthocyanidins such as epicatechin-4 ⁇ 8-epicatechin, catechin-4 ⁇ 8-epicatechin, epiafzelechin-4fi— >8-epicatechin, and epicatechin-4 ⁇ —>8-epicatechin-4 ⁇ - ⁇ 8-epicatechin. It is believed that a mixture of substantially pure proanthocyanidins as a pharmaceutical composition is especially advantageous and has not been earlier suggested in the art.
  • compositions contain one or more proanthocyanidins, each proanthocyanidin present in the composition in a proportion percentage or percentage purity that "significantly exceeds" a proportion percentage of the same proanthocyanidin' s natural presence in a plant, or in an extract from the plant. For example, suppose that a particular proanthocyanidin is present in a plant in a percentage by weight of 0.01 percent, and is present in an extract of the plant in a percentage by weight of 1.0 percent. In a disclosed composition then, the same proanthocyanidin is present in the composition in a percentage by weight that is significantly greater than 0.01 percent or 1.0 percent, say 10 percent.
  • a PA is present in a tablet to be delivered orally in accordance with the disclosure herein.
  • the PA is an isolated PA present in a percentage purity of 98.5% (that is, the PA is 98.5% pure, as measured by conventional purity indicia, such as for example the characteristic single sharp peak band on an HPLC).
  • the particular PA is however only a 15% ingredient by weight in the tablet.
  • the PA is known to be present in a fruit in a dry weight percentage of 0.06, while certain fruit extracts are known to contain up to 0.75 percent dry weight percent of the same PA.
  • the PA is proportionally more present in the tablet than in the extract by a ratio of 20: 1, and this is one measure of significantly exceeding the natural proportion percentage presence in a plant or extract of the plant.
  • a PA present in a therapeutically administered dose form that has a percentage of the PA (by weight, dry weight, volume, or purity) that is 10 times (or more) greater than the natural percentage presence of the same PA in a plant is a percentage that "significantly exceeds" the natural percentage presence of the PA in the plant.
  • extracts of a plant it should be noted that only conventional or natural extracts are to be considered (juices, concentrates and the like, or extracts known and used for other purposes), not new extracts prepared after the priority date of this disclosure the effect of which is to concentrate the particular PA so as to negative a finding of "significantly exceeding", as just defined. It should also be noted that in some cases, a finding of
  • the purpose of the disclosed standard of measurement is to set forth a fair margin by which a claimed composition exceeds reading on the active ingredients' natural occurrence in plants and conventional extracts of plants.
  • compositions will contain proanthocyanidin that is at least substantially pure.
  • Proanthocyanidin that is in substantially pure isolated or synthetic form may be advantageously employed as well.
  • pure means better than 95% pure
  • substantially pure PA means a PA purified by extraction or other known means or means disclosed herein such that the PA is present in the therapeutic dosage with only those impurities that can not readily nor reasonably be removed by the extraction or purification processes.
  • isolated means that the PA in question is not accompanied in the therapeutic form by significant quantities of other PA's.
  • An “isolated pure” compound is a compound in isolated purified form such as is conventional for active ingredients in the pharmaceutical industry.
  • One method includes a) dissolving the plant material with methanol or the like non-polar solvent, b) loading the methanol-extracted plant material onto a silica gel column, c) eluting the column with a series of increasing proportions of methanol in chloroform to elute the proanthocyanidins, d) separating the proanthocyanidins in the extract by reverse phase HPLC, and e) collecting and freeze drying the separated and isolated proanthocyanidin, now deemed thereby to be "pure”.
  • the series of methanol in chloroform elutions will beneficially include at least elutions of 10% methanol in chloroform, 20% methanol in chloroform, 40% methanol in chloroform, 50% methanol, and 100% methanol in chloroform.
  • a preferred plant material is derived from Uncaria tomentosa.
  • a proanthocyanidin composition made from the disclosed isolation process is also disclosed.
  • composition eluted from the silica gel column with the 20% methanol in chloroform step of the series will contain primarily procyanidin dimers and trimers;
  • proanthocyanidin composition eluted from the silica gel column with the 40% methanol in chloroform step will contain primarily procyanidin trimers and tetramers;
  • the composition eluted from the silica gel column with the 50% methanol in chloroform step will contain primarily procyanidin trimers, tetramers, pentamers, and hexamers;
  • the composition eluted from the silica gel column with the 100% methanol in chloroform step will contain primarily procyanidins tetramers, pentamers, hexamers, and oligomers of greater than six units.
  • a second isolation method includes a) dissolving the plant material with ethanol or the like non-polar solvent, b) loading the ethanol-extracted plant material onto a LH20 column, c) eluting the column with ethanol, followed by a series of increasing proportions of acetone in ethanol (and/or methanol) to elute the proanthocyanidins, d) separating the proanthocyanidins in the extract by reverse phase HPLC, and e) collecting and freeze drying the separated and isolated proanthocyanidin, now deemed thereby to be "pure".
  • the series of acetone in ethanol (and/or methanol) elutions will beneficially include at least elutions of 5% acetone in ethanol, 10% acetone in ethanol, 50% acetone in ethanol, 50% acetone in methanol and 100% methanol.
  • a preferred plant material is derived from Uncaria tomentosa.
  • a proanthocyanidin composition made from the second disclosed isolation process is also disclosed.
  • the composition eluted from the LH 20 column with ethanol will contain primarily procyanidin dimers and trimers; the proanthocyanidin composition eluted from the LH20 column with the 5% acetone in ethanol step will contain primarily procyanidin dimers and trimers; the proanthocyanidin composition eluted from the LH20 column with the 10% acetone in ethanol step will contain primarily procyanidin dimers and trimers; the proanthocyanidin composition eluted from the LH20 column with the 50% acetone in ethanol step will contain primarily procyanidin dimers, trimers, and tetramers; the proanthocyanidin composition eluted from the LH20 column with the 50% acetone in methanol step will contain primarily procyanidin trimers, tetramers, pentamers, hexamers and oligomers of greater than six units; and the pro
  • a further method of treatment of an amyloid disease, or a disease characterized by ⁇ - synuclein or NAC fibrillogenesis, in a mammalian subject is also disclosed.
  • the method includes administering to the subject a therapeutically effective amount of the proanthocyanidin isolated by way of the disclosed isolation process.
  • the amyloid disease for treatment is selected from the group consisting of amyloid diseases associated with Alzheimer's disease, Down's syndrome, hereditary cerebral hemorrhage with amyloidosis of the Dutch type, inclusion body myositosis, the amyloidosis associated with type 2 diabetes, the amyloidosis associated with chronic inflammation, various forms of malignancy, and Familial Mediterranean Fever, the amyloidosis associated with multiple myeloma and other B-cell dyscrasias, the amyloidosis associated with the prion diseases including Creutzfeldt- Jakob disease, Gerstmann-Strausller syndrome, kuru, animal scrapie, and mad cow disease, the amyloidosis associated with long- term hemodialysis and carpal tunnel syndrome, the amyloidosis associated with endocrine tumors such as medullary carcinoma of the thyroid, and the ⁇ -synuclein disease is selected from the group consisting of Parkinson's
  • compositions include a pharmaceutically acceptable carrier, diluent, or excipient, or the like, and a proanthocyanidin (PA), or proanthocyanidin compound, that may be found in the group of proanthocyanidin and proanthocyanidin compounds characterized by either Formula I or Formula II, or both (see Figures 54-56).
  • PA proanthocyanidin
  • the group also includes proanthocyanidins characterized by oligomeric combinations of Formula I and Formula II (see Figure 56), and also includes any pharmaceutically acceptable salt of any of the foregoing proanthocyanidins.
  • n is an integer in the range of 2 to 20 and preferably 2-5 or even 2-3.
  • the PA is selectably present in the composition in an amount effective to treat an amyloid disease, or a disease characterized by ⁇ -synuclein or NAC fibrillogenesis, in a mammalian subject.
  • Rj and R 2 are independently selected from hydrogen and hydroxy;
  • R 3 is selected from the group consisting of hydrogen, optionally substituted O- glycosyl, -C(O)-(optionally substituted aryl), and -C(O)-(optionally substituted heteroaryl);
  • R 4 is selected from the group consisting of hydrogen, catechin, epicatechin, and gallates of catechin and epicatechin.
  • the lines at the 2-, 3- and 4-position denote optional R and S (sometimes and alternatively referred to as ⁇ and ⁇ ) stereochemical configurations. Generally the configuration at the 4-position is trans to the configuration at the 3-position.
  • the lines at the 4- and 8-positions in Formula I and at the 4- and 6- positions in Formula U denote possible oligomer bonds between individual units as earlier discussed, and the each of the substitutions at Rj, R 2 , R 3 , and R 4 , and each of the configurations at the 2-, 3-, and 4-positions, and each of the oligomer bond configurations of 4-8 and 4-6 are independently selected for each individual unit and may be different for unit in the oligomer series of units, though often the units of shorter oligomers are homogenous with one another.
  • the invention is described with reference to specific embodiments, plant species and parts, methods, procedures and the like.
  • polyphenols including flavanoids, procyanidins and proanthocyanidins can be isolated and/or purified from plant materials by a number of different methods. It will further be recognized that these alternate methods, and consequent changes in other steps of the method, such as use of different solvents or different columns for purification, and of procyanidins and proanthocyanidins from a composition of partially purified polyphenols, fall within the scope of the presently disclosed plant-derived extracts, and compounds derived thereof.
  • amyloid diseases include, but are not limited to, the amyloid associated with Alzheimer's disease, Down's syndrome, hereditary cerebral hemorrhage with amyloidosis of the Dutch type, inclusion body myositosis (wherein the specific amyloid is referred to as beta-amyloid protein or A ⁇ ), the amyloid associated with chronic inflammation, various forms of malignancy and Familial Mediterranean Fever (wherein the specific amyloid is referred to as AA amyloid or inflammation-associated amyloidosis), the amyloid associated with multiple myeloma and other B-cell dyscrasias (wherein the specific amyloid is referred to as AL amyloid), the amyloid associated with type U diabetes (wherein the specific amyloid protein is referred to as amylin or islet amyloid polypeptide), the amyloid associated with the prion diseases including Creutzfeldt- Jakob disease, Gerstmann-Straus
  • ⁇ - synuclein protein which forms fibrils, and is Congo red and Thioflavin S positive, is found as part of Lewy bodies in the brains of patients with Parkinson's disease, Lewy body disease (Lewy in Handbuch der Neurologie. M. Lewandowski, ed., Springer, Berline pp.920-933, 1912; Pollanen et al, J. Neuropath. Exp. Neurol. 52:183-191, 1993; Spillantini et al, Proc. Natl. Acad. Sci. USA 95:6469-6473. 1998; Arai et al, Neurosc. Lett. 259:83-86, 1999), and multiple system atrophy.
  • Parkinson's disease due to the fact that fibrils develop in the brains of patients with this disease (which are Congo red and Thioflavin S positive, and which contain predominant beta-pleated sheet secondary structure), should be regarded as a disease that also displays the characteristics of an amyloid-like disease.
  • Use of the inner bark and/or roots from Uncaria tomentosa also referred to as Una de
  • Gato or Cat's claw to isolate and use the amyloid inhibiting compounds for the treatment of amyloid formation, deposition, accumulation and/or persistence in Alzheimer's disease, type IT diabetes other amyloidoses, and Parkinson's disease are disclosed.
  • Una de Gato or Cat's claw is also referred to as, but not limited to, Paraguayo, Garabato, Garbato casha, Tambor huasca, Una de gavilan, Hawk's claw, Nail of Cat, and Nail of Cat Schuler.
  • Rubiciaceae family which includes but is not limited to the Uncaria genus, for the treatment of amyloid formation, deposition, accumulation and/or persistence in Alzheimer's disease, type II diabetes, other amyloidoses and Parkinson's disease are disclosed.
  • Uncaria species which may include but not limited to, Uncaria tomentosa, Uncaria attenuata, Uncaria elliptica, Uncaria guianensis, Uncaria pteropoda, Uncaria bernaysli, Uncaria ferra DC, Uncaria kawakamii, Uncaria rhyncophylla, Uncaria calophylla, Uncaria gambir, and Uncaria orientalis are also disclosed.
  • Uncaria tomentosa and related plant materials for the treatment of amyloid formation, deposition, accumulation and or persistence in Alzheimer's disease, type II diabetes, other amyloidoses and Parkinson's disease are disclosed.
  • Uncaria tomentosa extracts and individual compounds derived thereof The extract preferably comprises polyphenols(s), such as polyphenols of a least one proanthocyanidin selected from, but not limited to, epicatechin, catechin, epiafzelechin, procyanidin B2, procyanidin oligomers 2 though 10, preferably 2 through 5 or 4 through 10, procyanidin B4, procyanidin Cl, and derivatives thereof.
  • polyphenols(s) such as polyphenols of a least one proanthocyanidin selected from, but not limited to, epicatechin, catechin, epiafzelechin, procyanidin B2, procyanidin oligomers 2 though 10, preferably 2 through 5 or 4 through 10, procyanidin B4, procyanidin Cl, and derivatives thereof.
  • proanthocyanidins contained within Uncaria tomentosa and related plant materials for the treatment of amyloid formation, deposition, accumulation and/or persistence in Alzheimer's disease, type II diabetes, other amyloidoses and Parkinson's disease are disclosed.
  • Alzheimer's disease type II diabetes, other amyloidoses and Parkinson's disease are disclosed.
  • epicatechin-4 ⁇ - 8-epicatechin also known as procyanidin or proanthocyanidin B2
  • procyanidin or proanthocyanidin B2 for the treatment of amyloid formation, deposition, accumulation and/or persistence in Alzheimer's disease, type II diabetes, other amyloidoses and Parkinson's disease
  • catechin-4 ⁇ — 8-epicatechin also known as procyanidin or proanthocyanidin B4
  • amyloid formation, deposition, accumulation and/or persistence in Alzheimer's disease, type II diabetes, other amyloidoses and Parkinson's disease is disclosed.
  • epicatechin-4 ⁇ —»8-epicatechin-4 ⁇ — >8-epicatechin also known as procyanidin or proanthocyanidin Cl
  • procyanidin or proanthocyanidin Cl for the treatment of amyloid formation, deposition, accumulation and/or persistence in Alzheimer's disease, type II diabetes, other amyloidoses and Parkinson's disease is disclosed.
  • epiafzelechin-4 ⁇ — >8-epicatechin for the treatment of amyloid formation, deposition, accumulation and/or persistence in Alzheimer's disease, type II diabetes, other amyloidoses and Parkinson's disease is also disclosed.
  • Uncaria tomentosa and related plant materials for use as potent agents which inhibit amyloid formation, amyloid deposition, amyloid accumulation, amyloid persistence, amyloid protein- amyloid protein interactions, and/or cause a dissolution/disruption of pre-formed or pre- deposited amyloid fibrils in Alzheimer's disease, type II diabetes, systemic AA amyloidosis, other amyloidoses and Parkinson's disease are also disclosed.
  • compositions and methods involving administering to a subject a therapeutic dose of proanthocyanidins, epicatechin-4 ⁇ — >8-epicatechin, catechin-4 ⁇ -»8-epicatechin, epiafzelechin- 4 ⁇ — »8-epicatechin, epicatechin-4 ⁇ — >8-epicatechin-4 ⁇ — >8-epicatechin or analogs or derivatives thereof (as disclosed herein) that inhibits amyloid deposition are disclosed. Accordingly, the compositions and methods of the invention are useful for inhibiting amyloidosis in disorders in which amyloid deposition occurs.
  • the compounds of the invention can be used therapeutically to treat amyloidosis or can be used prophylactically in a subject susceptible to amyloidosis.
  • the methods of the invention are based, at least in part, in directly inhibiting amyloid fibril formation, inhibiting amyloid fibril growth, and/or causing dissolution/disruption of preformed amyloid fibrils.
  • Pharmaceutical compositions for treating amyloidosis are disclosed.
  • the pharmaceutical compositions include a therapeutic compound of the invention in an amount effective to inhibit amyloid deposition and a pharmaceutically acceptable vehicle.
  • the proanthocyanidin composition of the invention which can be administered as a pharmaceutical composition is disclosed.
  • the pharmaceutical composition may include, but is not limited to, a proanthocyanidin extract or purified compound, and a pharmaceutically acceptable carrier, such as lactose, cellulose, or equivalent, or contained within a pharmaceutical dosage, such as a capsule or tablet.
  • procyanidin B2 epicatechin-4 ⁇ ->8-epicatechin
  • catechin-4 ⁇ — >8- epicatechin i.e. procyanidin B4
  • epicatechin-4 ⁇ — >8-epicatechin-4 ⁇ — >8-epicatechin i.e.
  • procyanidin Cl epiafzelechin-4 ⁇ — >8-epicatechin, or analogs or derivatives thereof, including proanthocyanidins Bl, B2, B3, B4, B5, B6, B7, B8, Cl or C2, for use as potent agents which inhibit amyloid formation, amyloid deposition, amyloid accumulation, amyloid persistence, amyloid protein-amyloid protein interactions, and/or cause a dissolution/ disruption of pre- formed or pre-deposited amyloid fibrils in Alzheimer's disease, type II diabetes, systemic AA amyloidosis, other amyloidoses and Parkinson's disease is disclosed.
  • Preventing or treating amyloidosis in a mammal by administering a proanthocyanidin composition which may include but not limited to, a proanthocyanidin extract, a proanthocyanidin compound, a proanthocyanidin polymer or mixture thereof, to the mammal in an amount and for a time sufficient to prevent, reduce, or eliminate amyloid formation, deposition, accumulation and/or persistence, and thereby lead to effective treatments for Alzheimer's disease, Parkinson's disease, type 2 diabetes, systemic AA amyloidosis, and other amyloid disorders is disclosed.
  • a proanthocyanidin composition which may include but not limited to, a proanthocyanidin extract, a proanthocyanidin compound, a proanthocyanidin polymer or mixture thereof
  • a method of isolation to purify and identify the procyanidins, proanthocyanidins, epicatechin-4 ⁇ -8-epicatechin (i.e. procyanidin B2), catechin-4 ⁇ — >8-epicatechin (i.e. procyanidin B4), epicatechin-4 ⁇ — >8-epicatechin-4 ⁇ -»8-epicatechin (i.e. procyanidin Cl) and epiafzelechin-4 ⁇ — >8-epicatechin, or analogs or derivatives thereof from Uncaria tomentosa are disclosed.
  • an extract prepared to produce commercially obtained pills, tablets, caplets, soft and hard gelatin capsules, lozenges, sachets, cachets, vegicaps, liquid drops, elixers, suspensions, emulsions, solutions, syrups, tea bags, aerosols (as a solid or in a liquid medium), suppositories, sterile injectable solutions, sterile packaged powders, bark bundles and/or bark powder, using the methods described in the present invention.
  • Amyloid is a generic term referring to a group of diverse but specific extracellular protein deposits which all have common morphological properties, staining characteristics, and X-ray diffraction spectra. Regardless of the nature of the amyloid protein deposited all amyloids have the following characteristics: 1) showing an amorphous appearance at the light microscopic level, appearing eosinophilic using hematoxylin and eosin stains; 2) staining with Congo red and demonstrating a red/green birefringence as viewed under polarized light (Puchtler et al., J. Histochem. Cytochem.
  • Amyloidoses and "amyloid diseases" today are classified according to the specific amyloid protein deposited.
  • amyloids include, but are not limited to, the amyloid associated with Alzheimer's disease, Down's syndrome, hereditary cerebral hemorrhage with amyloidosis of the Dutch type and inclusion body myositosis (where the specific amyloid is referred to as beta-amyloid protein or A ⁇ ), the amyloid associated with chronic inflammation, various forms of malignancy and familial Mediterranean fever (where the specific amyloid is referred to as AA amyloid or inflammation- associated amyloid), the amyloid associated with multiple myeloma and other B-cell dyscrasias (where the specific amyloid is referred to as AL amyloid), the amyloid associated with type U diabetes (where the specific amyloid is referred to as amylin or islet amyloid), the amyloid associated with the prion diseases including Creutzfeldt- Jakob disease, Gerstmann-Straussler syndrome, kuru, and scrapie (where the specific amyloid is referred to as PrP am
  • amyloid deposits in clinical conditions share common physical properties relating to the presence of a beta-pleated sheet conformation
  • many different chemical types exist and additional ones are likely to be described in the future.
  • a circulating precursor protein may result from overproduction of either intact or aberrant molecules (for example, in plasma cell dyscrasias), reduced degradation or excretion (serum amyloid A in some secondary amyloid syndromes and beta 2 -microglobulin in long-term hemodialysis), or genetic abnormalities associated with variant proteins (for example, familial amyloidotic polyneuropathy).
  • Proteolysis of a larger protein precursor molecule occurs in many types of amyloidosis, resulting in the production of lower molecular weight fragments that polymerize and assume a beta-pleated sheet conformation as tissue deposits, usually in an extracellular location.
  • the precise mechanisms involved and the aberrant causes leading to changes in proteolytic processing and/or translational modification are not known in most amyloids.
  • Systemic amyloid diseases which include the amyloid associated with chronic inflammation, various forms of malignancy and familial Mediterranean fever (i.e. AA amyloid or inflammation-associated amyloidosis) (Benson and Cohen, Arth. Rheum. 22:36-42, 1979; Kamei et al, Acta Path. Jpn. 32:123-133, 1982; McAdam et al., Lancet 2:572-573, 1975; Metaxas, Kidney Int. 20:676-685, 1981), and the amyloid associated with multiple myeloma and other B-cell dyscrasias (i.e. AL amyloid) (Harada et al., J. Histochem. Cytochem.
  • amyloid deposition in these diseases may occur, for example, in liver, heart, spleen, gastrointestinal tract, kidney, skin, and/or lungs (Johnson et al, N. Engl. J. Med. 321:513-518, 1989).
  • amyloid deposition in the kidney may lead to renal failure
  • amyloid deposition in the heart may lead to heart failure.
  • amyloid accumulation in systemic organs leads to eventual death generally within 3-5 years.
  • amyloidoses may affect a single organ or tissue such as observed with the A ⁇ amyloid deposits found in the brains of patients with Alzheimer's disease and Down's syndrome: the PrP amyloid deposits found in the brains of patients with Creutzfeldt- Jakob disease, Gerstmann-Straussler syndrome, and kuru; the islet amyloid (amylin) deposits found in the islets of Langerhans in the pancreas of 90% of patients with type II diabetes (Johnson et al, N. Engl. J. Med. 321:513-518, 1989; Lab. Invest.
  • beta 2 -microglobulin amyloid deposits in the medial nerve leading to carpal tunnel syndrome as observed in patients undergoing long-term hemodialysis (Geyjo et al, Biochem. Biophys. Res. Comm. 129:701-706, 1985; Kidney InL 30:385-390, 1986); the prealbumin/transthyretin amyloid observed in the hearts of patients with senile cardiac amyloid; and the prealbumin/transthyretin amyloid observed in peripheral nerves of patients who have familial amyloidotic polyneuropathy (Skinner and Cohen, Biochem. Biophys. Res. Comm. 99:1326- 1332, 1981; Saraiva et al, J.
  • Alzheimer's disease is a leading cause of dementia in the elderly, affecting 5-10% of the population over the age of 65 years (A Guide to Understanding Alzheimer's Disease and Related Disorders, Jorm, ed., New York University Press, New York, 1987).
  • Alzheimer's disease the parts of the brain essential for cognitive processes such as memory, attention, language, and reasoning degenerate, robbing victims of much that makes us human, including independence.
  • onset is in middle age, but more commonly, symptoms appear from the mid-60' s onward.
  • Alzheimer's disease today affects 4-5 million Americans, with slightly more than half of these people receiving care at home, while the others are in many different health care institutions.
  • Alzheimer's disease and other dementias doubles every 5 years beyond the age of 65, and recent studies indicate that nearly 50% of all people age 85 and older have symptoms of Alzheimer's disease (2000 Progress Report on Alzheimer's Disease, National Institute on Aging/National Institute of Health). 13% (33 million people) of the total population of the United States are age 65 and older, and this percentage will climb to 20% by the year 2025 (2000 Progress Report on Alzheimer's Disease).
  • Alzheimer's disease also puts a heavy economic burden on society.
  • a recent study estimated that the cost of caring for one Alzheimer's disease patient with severe cognitive impairments at home or in a nursing home, is more than $47,000 per year (A Guide to Understanding Alzheimer's Disease and Related Disorders).
  • the annual economic toll of Alzheimer's disease in the United States in terms of health care expenses and lost wages of both patients and their caregivers is estimated at $80 to $100 billion (2000 Progress Report on Alzheimer's Disease).
  • Tacrine hydrochloride (“Cognex”) the first FDA approved drug for Alzheimer's disease, is a acetylcholinesterase inhibitor (Cutler and Sramek, N.
  • Amyloid as a Therapeutic Target for Alzheimer's Disease is characterized by the deposition and accumulation of a 39-43 amino acid peptide termed the beta-amyloid protein, A ⁇ or ⁇ /A4 (Glenner and Wong, Biochem. Biophys. Res. Comm. 120:885-890, 1984; Masters et al., Proc. Natl. Acad. Sci. USA 82:4245-4249, 1985; Husby et al., Bull. WHO 71:105-108, 1993).
  • a ⁇ is derived by protease cleavage from larger precursor proteins termed beta-amyloid precursor proteins (or ⁇ PPs) of which there are several alternatively spliced variants.
  • BPPs proteins consisting of 695, 751 and 770 amino acids (Tanzi et al., Nature 331:528-530. 1988; Kitaguchi et al., Nature 331:530-532, 1988; Ponte et al., Nature 331:525-527, 1988).
  • the small A ⁇ peptide is a major component which makes up the amyloid deposits of "plaques” in the brains of patients with Alzheimer's disease.
  • Alzheimer's disease is characterized by the presence of numerous neurofibrillary "tangles", consisting of paired helical filaments which abnormally accumulate in the neuronal cytoplasm (Grundke-Iqbal et al., Proc. Natl. Acad. Sci. USA 83:4913-4917, 1986; Kosik et al., Proc. Natl. Acad. Sci. USA 83:4044-4048, 1986; Lee et al., Science 251:675-678, 1991).
  • the pathological hallmark of Alzheimer's disease is therefore the presence of "plaques” and "tangles", with amyloid being deposited in the central core of the plaques.
  • Alzheimer's disease brain is the accumulation of amyloid in the walls of blood vessels, both within the brain parenchyma and in the walls of meningeal vessels that lie outside the brain.
  • the amyloid deposits localized to the walls of blood vessels are referred to as cerebrovascular amyloid or congophilic angiopathy (Mandybur, J. Neuropath. Exp. Neurol. 45:79-90, 1986; Pardridge et al., J. Neurochem. 49:1394-1401, 1987).
  • Alzheimer's A ⁇ protein in cell culture has been shown to cause degeneration of nerve cells within short periods of time (Pike et al., Br. Res. 563:311-314, 1991; J. Neurochem. 64:253-265, 1995).
  • a ⁇ has also been found to be neurotoxic in slice cultures of hippocampus (Harrigan et al., Neurobiol. Aging 16:779-789, 1995) and induces nerve cell death in transgenic mice (Games et al., Nature 373:523-527, 1995; Hsiao et al, Science 274:99-102, 1996). Injection of the Alzheimer's A ⁇ into rat brain also causes memory impairment and neuronal dysfunction (Flood et al., Proc. Natl. Acad ' . Sci. USA 88:3363-3366, 1991: Br. Res. 663:271-276, 1994).
  • a ⁇ amyloid is directly involved in the pathogenesis of Alzheimer's disease. It has been discovered that the production of A ⁇ can result from mutations in the gene encoding, its precursor, beta amyloid precursor protein (Van Broeckhoven et al., Science 248:1120-1122, 1990; Murrell et al., Science 254:97-99, 1991; Haass et al., Nature Med. 1:1291-1296, 1995). The identification of mutations in the beta-amyloid precursor protein gene which causes early onset familial Alzheimer's disease is the strongest argument that amyloid is central to the pathogenetic process underlying this disease.
  • Parkinson's disease is a neurodegenerative disorder that is pathologically characterized by the presence of intracytoplasmic Lewy bodies (Lewy in Handbuch der Neurologic M. Lewandowski, ed., Springer, Berlin, pp. 920-933, 1912; Pollanen et al., J. Neuropath. Exp. Neurol. 52:183-191, 1993), the major components of which are filaments consisting of ⁇ - synuclein (Spillantini et al., Proc. Natl. Acad. Sci. t SA_95:6469-6473, 1998; Arai et al.,
  • ⁇ -Synuclein aggregation and fibril formation fulfills of the criteria of a nucleation-dependent polymerization process (Wood et al., J. Biol. Chem. 274:19509-19512, 1999).
  • ⁇ - synuclein fibril formation resembles that of Alzheimer's beta-amyloid protein (A ⁇ ) fibrils, ⁇ - Synuclein recombinant protein, and non-amyloid component (known as NAC), which is a 35- amino acid peptide fragment of ⁇ -synuclein, both have the ability to form fibrils when incubated at 37°C, and are positive with amyloid stains such as Congo red (demonstrating a red/green birefringence when viewed under polarized light) and Thioflavin S (demonstrating positive fluorescence) (Hashimoto et al., Brain Res. 799:301-306, 1998; Ueda et al., Proc. Natl. Acad. Sci. U
  • ⁇ -synuclein/NAC accumulation of ⁇ -synuclein/NAC is also a cytopathological feature common to Lewy body disease and multiple system atrophy (Wakabayashi et al, Acta Neuropath. 96:445-452, 1998; Piao et al, Acta Neuropath. 101:285-293. 2001).
  • Multiple system atrophy is a sporadic neurodegenerative disease in adults characterized by neuronal and glial cytoplasmic inclusions, containing ⁇ -synuclein/NAC.
  • Parkinson's disease ⁇ -synuclein NAC fibrils like the A ⁇ fibrils of Alzheimer's disease, also consist of a predominant beta-pleated sheet structure. It is therefore believed that compounds found to inhibit Alzheimer's disease A ⁇ amyloid fibril formation can also be anticipated to be effective in the inhibition of ⁇ -synuclein and/or NAC fibril formation. These compounds would therefore also serve as therapeutics for Parkinson's disease, in addition to having efficacy as a therapeutic for Alzheimer's disease and other amyloid disorders.
  • Islet amyloid deposits are observed in ⁇ 90% of patients with well-established type 2 diabetes and would appear to be a characteristic feature of the disease process (Westermark, Med. Sci. 77:91-94,1972; Clark et al, Diabetes Res. 9:151-159,1988). In many patients the deposits are widespread and affect many islets. The degree of islet (predominantly ⁇ -cell) mass that has been replaced by amyloid may be a marker for the severity of the diabetic disease process, with those individuals requiring insulin treatment having the greatest islet mass reduction and amyloid formation (Westermark, Amyloid: Int. J. Exp. Clin. Invest. 1:47- 60,1994).
  • islet amyloid Since islet amyloid has been observed in autopsy samples obtained from different populations, it appears to be a phenomenon common to the disease rather than to a subpopulation of individuals with the syndrome (Westermark, J. Med. Sci. 77:91-94,1972; Clark et al, Diabetes Res. 9:151-159,1988). The prevalence of islet amyloid deposits increases with age (Bell, Am. J. Path. 35:801-805, 1959), which is not surprising because normal aging is associated with a deterioration in glucose tolerance and an increased prevalence of type 2 diabetes (Davidson, Metabolism 28:687-705, 1979).
  • IAPP islet amyloid polypeptide
  • amylin The major protein in islet amyloid is a 37-amino acid peptide known as islet amyloid polypeptide (IAPP) or amylin.
  • IAPP is a known normal secretory product of the pancreatic ⁇ - cells (Kanh et al, Diabetes 39:634-638,1990) that is stored in insulin-bearing cytoplasmic granules (Clark et al, Cell Tissue Res. 257:179-185, 1989). It has long been questioned whether the deposition of islet amyloid is involved in or merely a consequence of the pathogenesis of type 2 diabetes.
  • islet amyloid formation, deposition and persistence may be an important primary factor leading to ⁇ -cell dysfunction and cell death, hyperglycemia, and in the development of type 2 diabetes.
  • IAPP has been hypothesized to have an important role in the pathogenesis of type 2 diabetes through its impairment of ⁇ -cell function and reduction of ⁇ -cell mass (Johnson et al, N. Engl. J. Med. 321:513-518,1989). Besides being able to form islet amyloid deposits that replace ⁇ -cell mass, amyloid fibrils appear to damage islets directly.
  • Uncaria tomentosa also known as "Una de Gato” (in Spanish) or "Cat's claw” (in English) refers to a woody vine that grows within the Peruvian Amazon rain forest. This slow growing vine takes 20 years to reach maturity, and can grow over 100 feet in length as it attaches and wraps itself around the native trees. It is found abundantly in the foothills, at elevations of two to eight thousand feet. The vine is referred to as "Cat's claw” because of its distinctive curved claw-like thorns that project from the base of its leaves.
  • the native Indian tribes traditionally have boiled the inner bark and root of the herb to make a tea decoction and regard Uncaria tomentosa as a sacred medicinal plant.
  • Uncaria tomentosa The highly effective properties contained within the inner bark of this plant are believed to have a profound and positive influence on the body, although scientific medical data is generally lacking on its potential benefits in humans.
  • the alkaloids and phytochemicals in the inner bark of Uncaria tomentosa are almost identical to those found in the root, and harvesting this way preserves the plant and provides for the future of the rainforest.
  • Some of the active substances present in Uncaria tomentosa are alkaloids, which occur in the plant and its watery extract as a complex bound to tannins. In this form, only little of them can be activated. The complexes get split by the acid milieu of the stomach; the alkaloids get transformed into their hydrochloride form, and in this way, get well absorbed.
  • Uncaria tomentosa extract means more tannin is present and beneficial alkaloids are locked up with the tannins, which have formed a non-bioavailable and poorly absorbed complex.
  • a light golden color of Uncaria tomentosa suggests that there are less tannins, and more alkaloids available in the extract.
  • Uncaria tomentosa is one of the most important plants in the South American Peruvian rainforest. A number of oxindole alkaloids have already been isolated from the inner bark of this plant. Two US patents (US patent #4,844,901 and US patent #4,940,725) describe the isolation and use of six oxindole alkaloids from Uncaria tomentosa, which are believed to be "suitable for the unspecified stimulation of the immunologic system". These oxindole alkaloids are believed to provide a general boost to the immune system as well as have a profound effect on the ability of white blood cells and macrophages to phagocytize harmful microorganisms and foreign matter. The most immunologically active alkaloid appears to be alloisopteropodine, isomer A, a pentacyclic oxindole alkaloid (US patent #4,940,725).
  • Uncaria tomentosa may be used to treat a variety of ailments, nowhere has there been any use or suggestion of use, of this plant or extracts thereof, or compounds derived thereof, for the treatment of amyloid formation, deposition, accumulation and/or persistence, such as that which occurs in the amyloidoses, including Alzheimer's disease and Parkinson's disease.
  • the present invention clearly demonstrates the effectiveness of Uncaria tomentosa derived compounds, including procyanidins and proanthocyanidins, for the treatment of amyloidosis associated with Alzheimer's disease, type 2 diabetes, systemic A A amyloidosis, and other amyloid diseases, as well as for the treatment of ⁇ -synuclein fibril formation and accumulation, such as that observed in patients with Parkinson's disease.
  • Uncaria tomentosa derived compounds including procyanidins and proanthocyanidins
  • Proanthocyanidins Procyanidins. Flavanoids and Tannins
  • Proanthocyanidins are polyphenolic molecules occurring naturally in fruits, berries and other plant material. These molecules belong to the flavanoid family of compounds.
  • the flavanoid polyphenolics include the catechins, anthocyanins, and proanthocyanidins.
  • Proanthocyanidins are also known in the art as condensed tannins, leucoanthocyanidins, leucodelphinins, leucocyanins, anthocyanogens, epicatechin-catechin polymers or procyanidins.
  • procyanidins and proanthocyanidins see Santos-Buelga and Scalbert, J. Sc. Food Agri.
  • Proanthocyanidin oligomers or polymers useful for the present anti-amyloid activity are comprised of monomeric units of leucoanthocyanidins.
  • Leucoanthocyanidins are generally monomeric flavanoids which include catechins, epicatechins, gallocatechins, galloepicatechins, flavanols, flavonols, and flavan-3,4-diols, leucocyanidins and anthocyanidins.
  • the therapeutically effective proanthocyanidin polymers have from 2 to 20 flavanoid units, and more preferably from 2 to 11 flavanoid units.
  • Proanthocyanidins polymers or oligomers are known to have varying numbers of flavanoid units, and have been reported for example in Mattice et al, Phytochem. 23:1309- 1311, 1984; Czochanska et al, J.C. S. Chem. Comm. 375, 1979; Jones et al, Photochemistry, 15:1407-1409, 1976.
  • Proanthocyanidin oligomers having the recited ranges of flavanoid units and described in these references are incorporated herein by reference as if their disclosure was fully set forth herein.
  • Procyanidins also referred to as proanthocyanidins, are polymeric or oligomeric compounds composed of epicatechin and catechin residues. Disclosed compounds include dimers of epicatechin and catechin residues, and trimers of epicatechin. Catechin and epicatechin residues may be combined in all possible combinations in polymeric procyanidins up to molecular weights of up to about 10,000 daltons.
  • Proanthocyanidin polymers are known to have a varying number of flavanoid units. The polymers preferably contain two to fifteen monomeric flavanoid subunits, most preferably two to ten subunits.
  • Tannins are classically divided into 2 groups.
  • Hydrolysable tannins are esters of phenolic acids and a polyol, usually glucose.
  • the phenolic acids are either gallic acid in gallotannins or other phenolic acids derived from the oxidation of galloyl residues in ellagitannins.
  • Proanthocyanidins forming the second group of tannins, are far more common in our diet. They are polymers made of elementary flavan-3-ol units.
  • a key feature of proanthocyanidins is that they yield anthocyanidins upon heating in acidic media, hence their name (reviewed in Santos-Buelga and Scalbert, J. Sc. Food Agri. 80:1094-1117,2000).
  • tannins possess 12-16 phenolic groups and 5-7 aromatic rings per 1000 units of relative molecular mass (E. Haslam, Practical Polyphenoics-from Structure to Molecular Recognition and Physiological Action. Cambridge University Press, Cambridge, 1998). This feature, together with their high molecular weight, clearly makes the tannins and similar phenolic polymers found in processed products such as red wine or black tea different both in structure and properties from the low-molecular-weight phenolic acids and monomeric flavanoids.
  • the phenolic polymers formed by enzymatic and/or chemical transformation of simple flavanols, proanthocyanidins and other phenolic compounds, are called tannin-like compounds.
  • Proanthocyanidins are polymeric flavan-3-ols whose elementary units are linked by C- C and occasionally C-O-C bonds.
  • the flavan-3-ol units have the typical C6-C3-C6 flavanoid skeleton.
  • the three rings are distinguished by the letters A, B and C (see Figure 1). They differ structurally according to the number of hydroxyl groups on both aromatic rings and the stereochemistry of the asymmetric carbons on the heterocycle.
  • the most common proanthocyanidins in food are procyanidins with a 3', 4'-dihydroxy substitution on the B ring and prodelphinidins with a 3', 4', 5'-trihydroxy substitution.
  • Procyanidins or mixed procyanidins/prodelphinidins are most common in food.
  • Propelargonidins with 4'-hydroxy firings are relatively rare in food sources, bit notably disclosed herein in the form of epiafzelechin.
  • the three carbons C2, C3, C4 of the flavanol heterocycle are asymmetric and may occur in different configurations. With some very rare exceptions, the configuration of C2 is R.
  • Flavan-3-ol units with the 2S configuration are distinguished by the prefix enantio(ent-).
  • the stereochemistry of the C2-C3 linkage may be either trans (2R, 3S) or cis (2R, 3R) as in (+)-(gallo)catechin and (-)-epi(gallo)catechin polymers respectively.
  • the interflavan bond at C4 is always trans with respect to the hydroxy group at C3 (E. Haslam, Practical Polyphenoics- from Structure to Molecular Recognition and Physiological Action. Cambridge University Press, Cambridge, 1998).
  • the most usual interflavanol linkages are C-C bonds established between the C4 of one flavanoid unit ("extension or upper unit").
  • proanthocyanidins belong to the so-called B-type (dimeric) and C type (trimeric) proanthocyanidins.
  • oligomeric proanthocyanidins were named by an alpha-numeric system, with a letter A, B or C to describe the type of interflavanol linkage; a number was added to the letter as they were detected (Thompson et al, J. Chem. Soc. Perkins Trans. 1: 1387-1399, 1972).
  • a new nomenclature was later introduced to name an increasing number of new structures. It is based on that utilized for the polysaccharides (Hemingway et al, J. Chem. Soc. Perkins Trans. 1:1387-1399, 1972). In this nomenclature, the elementary units of the oligomers are designated with the name of the corresponding flavan-3-ol monomers.
  • Flavanol units can bear various acyl or glycosyl substituents.
  • the most common acyl substiuent is gallic acid which forms an ester with the hydroxyl n the C3 position, as in tea (Nonaka et al, Chem. Pharmaceutic. Bull. 31:3906-3914, 1983) and wine (Prieur et al,
  • glycosylated proanthocyanidin oligomers have also been characterized.
  • the sugar is generally linked to the hydroxyl group at the C3 position (Ishimaru et al, Phytochemistrv 26:1167-1170, 1987; Zhang et al, Phytochemistry 27:3277- 3280, 1988), but also at the C5 position (Gujer et al, Phytochemistrv 25:1431-1436, 1986).
  • proanthocyanidins heterosides are less frequently reported than other flavanoid glycosides, their occurrence may be underestimated, as sugars are frequently associated with purified proanthocyanidin polymers (Porter et al, Phytochemistry 24:567-569, 1985; Mathews et al, J. Agric. Food Chem. 45:1195-1201, 1997).
  • Such variations, and other variations disclosed herein are included with the scope of disclosure of the disclosed proanthocyanidins. More recently, the introduction of electrospray mass spectrometry techniques coupled to liquid chromatography led to a more detailed characterization of proanthocyanidin polymers. Such methods were employed in the present invention to identify procyanidins and proanthocyanidins derived from Uncaria tomentosa which demonstrate potent anti-amyloid and anti- ⁇ -synuclein/NAC activity.
  • FIGURE 2 is a HPLC tracing using method 1 (see Example 2, Table 1 for details) demonstrating the separation of PTI-777. Using this method, there is a good separation of HI and H2 peaks.
  • FIGURE 3 is a HPLC tracing using method 2 (see Example 2, Table 1 for details) demonstrating the separation of HI and H2 from PTI-777 following silica gel chromatography and elution with 20% methanol in chloroform.
  • FIGURE 4 is a HPLC tracing using method 1 (see Example 2, Table 1 for details) demonstrating the separation of mostly HI (with less H2) after fractioning PTI-777 using silica gel chromatography, followed by HPLC.
  • FIGURE 5 is a HPLC tracing using method 1 (see Example 2, Table 1 for details) demonstrating the isolation of pure H2 from PTI-777, after fractionating PTI-777 using silica gel chromatography, followed by HPLC.
  • FIGURE 6 is a ⁇ NMR spectrum of peak H2 derived from PTI-777.
  • FIGURE 7 is a 13 C NMR spectrum of peak H2 derived from PTI-777.
  • FIGURE 8 is a 1 C NMR spectrum of peak H2 in deteroacetone (instead of deuteromethanol).
  • FIGURE 9 is a 1H NMR spectrum of peak H2 in deteroacetone (instead of deuteromethanol).
  • FIGURE 10 is the peracetate structure of a sample of pure H2 following acetylation.
  • FIGURE 11 is the ⁇ NMR spectrum of the H2 peracetate in CDC1 3 .
  • FIGURE 12 is the 13 C NMR spectrum of the H2 peracetate in CDC1 3 .
  • FIGURE 13 is the CIGAR 'H- ⁇ C correlation spectrum (low resolution) of the H2 peracetate.
  • FIGURE 14 is the CIGAR ⁇ - 13 C correlation spectrum (high resolution) of the H2 peracetate.
  • FIGURE 15 is the NOESY correlation spectrum of the H2 peracetate.
  • FIGURE 16 is the NOESY correlation spectrum of the H2 peracetate.
  • FIGURE 17 is the NOESY correlation spectrum of the H2 peracetate.
  • FIGURE 18 is the structure of H2 identified to be epicatechin-4 ⁇ -»8-epicatechin.
  • FIGURE 19 is the ⁇ NMR spectrum of peak HI .
  • FIGURE 20 is the 13 C NMR spectrum of peak HI.
  • FIGURE 21 is the peracetate structure of a sample of pure HI following acetylation.
  • FIGURE 22 is the ⁇ NMR spectrum of the HI peracetate.
  • FIGURE 23 is the 13 C NMR spectrum of the HI peracetate.
  • FIGURE 24 is the CIGAR l H - 13 C correlation spectrum (low resolution) of the HI peracetate.
  • FIGURE 25 is the CIGAR ⁇ - 13 C correlation spectrum (high resolution) of the HI peracetate.
  • FIGURE 26 is the structure of HI identified to be catechin-4 ⁇ -»8-epicatechin.
  • FIGURE 27 is a HPLC tracing demonstrating separation of peak K2.
  • FIGURE 28 is a HPLC tracing demonstrating separation of a pure peak K2.
  • FIGURE 29 is a -ve ion electrospray mass spectrum of K2.
  • FIGURE 30 is the ⁇ NMR spectrum of K2.
  • FIGURE 31 is the ⁇ NMR spectrum of the K2 peracetate.
  • FIGURE 32 is the 13 C NMR spectrum of the K2 peracetate.
  • FIGURE 33 is a CIGAR l H- C correlation spectrum (low resolution) of the K2 peracetate.
  • FIGURE 34 is a CIGAR ⁇ - 13 C correlation spectrum (high resolution) of the K2 peracetate.
  • FIGURE 35 is the peracetate structure of K2.
  • FIGURE 36 is the structure of K2 determined to be epicatechin-4 ⁇ — >8-epicatechin-4 ⁇ — >8- epicatechin.
  • FIGURE 37 is a graph of a Thioflavin T fluorometry assay demonstrating the dose-dependent disruption/disassembly of pre-formed A ⁇ 1-42 fibrils by proanthocyanidins (compounds H2, Hl and K2).
  • FIGURE 38 is a black and white figure of a SDS-PAGE/Western blot further demonstrating the disruption of A ⁇ 1-42 fibrils, even in monomeric form by proanthocyanidins (compounds).
  • FIGURE 39 is a graph of a circular dichroism spectroscopy assay demonstrating compound H2 (referred to as PTC38 in this figure) causes a marked disruption/disassembly of ⁇ -sheet structure in A ⁇ 1-42 fibrils at 7 days following incubation.
  • FIGURE 40 is a graph of a circular dichroism spectroscopy assay demonstrating compound
  • FIGURE 41 is a graph of a Thioflavin T fluorometry assay demonstrating the dose-dependent disruption/disassembly of pre-formed NAC fibrils by proanthocyanidins (compounds H2, HI and K2).
  • FIGURE 42 is a graph of a Thioflavin T fluorometry assay demonstrating the dose-dependent disruption/disassembly of pre-formed IAPP fibrils by proanthocyanidins (compounds H2, HI and K2).
  • FIGURE 43 is the peracetate structure of Kl.
  • FIGURE 44 is the structure of Kl determined to be epiafzelechin-4 ⁇ — »8-epicatechin.
  • FIGURE 45 is the " ve ion electrospray mass spectrum of Kl.
  • FIGURE 46 is the 13 C NMR spectrum of Kl.
  • FIGURE 47 is the ⁇ NMR spectrum of Kl .
  • FIGURE 48 is the ⁇ NMR spectrum of the Kl peracetate.
  • FIGURE 49 is the I3 C NMR spectrum of the Kl peracetate.
  • FIGURE 50 is the CIGAR 'H - 13 C correlation spectrum (low resolution) of the Kl peracetate.
  • FIGURE 51 is the CIGAR 'H - 13 C correlation spectrum (medium resolution) of the Kl peracetate.
  • FIGURE 52 is the CIGAR H - 13 C correlation spectrum (high resolution) of the Kl peracetate.
  • FIGURE 53 is the CIGAR ! H - 13 C correlation spectrum (high resolution) of the Kl peracetate.
  • FIGURE 54 is an illustration of general Formula I for the structure of proanthocyanidins.
  • FIGURE 55 is an illustration of general Formula U for the structure of proanthocyanidins.
  • FIGURE 56 is an alternate example of proanthocyanidin structure.
  • FIGURE 57 is a flowchart of an isolation process for proanthocyanidins.
  • Proanthocyanidins includes “procyanidins”; “procyanidins” are a specific class of
  • “Mammal” and “mammalian subject” includes, but is not limited to, humans and non- human mammals, such as companion animals (cats, dogs, and the like), lab animals (such as mice, rats, guinea pigs, and the like) and farm animals (cattle, horses, sheep, goats, swine, and the like).
  • “Pharmaceutically acceptable excipient” means an excipient that is useful in preparing a pharmaceutical composition that is generally safe, non-toxic, and desirable, and includes excipients that are acceptable for veterinary use as well as for human pharmaceutical use. Such excipients may be solid, liquid, semisolid, or, in the case of an aerosol composition, gaseous.
  • “Pharmaceutically acceptable salts” means salts that are pharmaceutically acceptable and have the desired pharmacological properties. Such salts include salts that may be formed where acidic protons present in the compounds are capable of reacting with inorganic or organic bases. Suitable inorganic salts include those formed with the alkali metals, e.g. sodium and potassium, magnesium, calcium, and aluminum.
  • Suitable organic salts include those formed with organic bases such as the amine bases, e.g. ethanolamine, diethanolamine, triethanolamine, tromethamine, N-methylglucamine, and the like. Such salts also include acid addition salts formed with inorganic acids (e.g. hydrochloric and hydrobromic acids) and organic acids (e.g. acetic acid, citric acid, maleic acid, and the alkane- and arene-sulfonic acids such as methanesulfonic acid and benzenesulfonic acid). When there are two acidic groups present, a pharmaceutically acceptable salt may be a mono-acid-mono-salt or a di-salt; and similarly where there are more than two acidic groups present, some or all of such groups can be salified.
  • organic bases such as the amine bases, e.g. ethanolamine, diethanolamine, triethanolamine, tromethamine, N-methylglucamine, and the like.
  • Such salts also include acid
  • a “therapeutically effective amount” in general means the amount that, when administered to a subject or animal for treating a disease, is sufficient to effect the desired degree of treatment for the disease.
  • a “therapeutically effective amount” or a “therapeutically effective dosage” preferably inhibits, reduces, disrupts, disassembles amyloidosis, fibril formation, deposition, accumulation and/or persistence, or a disease associated with ⁇ - synuclein/NAC fibril formation in a patient by at least 20, more preferably by at least 40%, even more preferably by at least 60%, and still more preferably by at least 80%, relative to untreated subjects.
  • Effective amounts of a proanthocyanidin or procyanidin, or other disclosed compositions for treatment of a mammalian subject are about 1 mg to about 10,000 mg/kg of body weight of the subject, but more preferably from about 10 mg/kg/body weight to 100 mg/kg body weight.
  • a broad range of disclosed composition dosages are believed to be both safe and effective.
  • Treating" or “treatment” of a disease includes preventing the disease from occurring in a mammal that may be predisposed to the disease but does not yet experience or exhibit symptoms of the disease (prophylactic treatment), inhibiting the disease (slowing or arresting its development), providing relief from the symptoms or side-effects of the disease (including palliative treatment), and relieving the disease (causing regression of the disease).
  • Treating" amyloidosis or “amyloid diseases” includes any one or more of the following: preventing, inhibiting, reducing, disassembling, disrupting, and disaggregating amyloid fibrils and amyloid protein deposits, such as A ⁇ and the other amyloids referred to herein.
  • Treating" an ⁇ -synuclein disease or “treating ⁇ -synuclein or NAC fibrillogenesis” includes any one or more of the following: preventing, inhibiting, reducing, disassembling, disrupting, and disaggregating ⁇ -synuclein/NAC fibrils and ⁇ -synuclein/NAC-associated protein deposits, such as those in Lewy body disease, Parkinson's disease and multiple system atrophy.
  • NAC non-amyloid component
  • ⁇ -synuclein is a 35-amino acid peptide fragment of ⁇ -synuclein, which also, like ⁇ -synuclein, has the ability to form amyloid-like fibrils when incubated at 37°C, and are positive with amyloid stains such as Congo red (demonstrating a red/green birefringence when viewed under polarized light) and Thioflavin S (demonstrating positive fluorescence) (Hashimoto et al., Brain Res. 799:301-306, 1998; Ueda et al., Proc. Natl. Acad. Sci. U.S.A 90:11282-11286, 1993).
  • Fibrillogenesis refers to the presence of amyloid fibrils or fibrils formed containing ⁇ -synuclein and/or NAC. Inhibition of such fibrillogenesis with a therapeutic compound may include, but not limited to, treating, inhibiting, preventing, or managing such amyloid, amyloid fibril, ⁇ -synuclein and/or NAC fibril formation, deposition, accumulation, aggregation and/or persistence in a mammalian subject.
  • a pharmaceutical agent or “pharmacological agent” or “pharmaceutical composition” refers to a compound or combination of compounds used for treatment, preferably in a pure or near pure form.
  • pharmaceutical or pharmacological agents include the proanthocyanidins and procyanidins as examples.
  • Disclosed pharmaceutical or pharmacological compounds or compounds in compositions are purified to 80% homogeneity, and preferably 90% homogeneity. Compounds and compositions purified to 99.9% homogeneity are believed to be advantageous. As a test or confirmation, a pure compound on HPLC would yield a single sharp-peak band.
  • the disclosed compounds and compositions may possess one or more chiral centers, and can therefore be produced as individual stereoisomers or as mixtures of stereoisomers, depending on whether individual stereoisomers or mixtures of stereoisomers of the starting materials are used. Unless indicated otherwise, the description or naming of a compound or group of compounds is intended to include both the individual stereoisomers and mixtures (racemic or otherwise) of stereoisomers. Methods for the determination of stereochemistry and the separation of stereoisomers are well known to a person of ordinary skill in the art [see the discussion in Chapter 4 of March J: Advanced Organic Chemistry. 4th ed. John Wiley and Sons, New York, NY, 1992].
  • Optionally substituted glycosyl is glycosyl optionally substituted with up to three anionic substituents selected from sulfate, sulfonate, phosphate, phosphonate, and carboxylate, each optionally esterified with optionally substituted alkyl, optionally substituted aryl, or optionally substituted heteroaryl; examples include glucosyl, galactosyl, rhamnogalactosyl, and the like.
  • Aryl is a cyclic (monocyclic, condensed bicyclic, or linked bicyclic) group having from 5 to 12 ring carbon atoms, and sufficient ring unsaturation that the group is "aromatic” as that term is conventionally used, e.g. phenyl, naphthyl, biphenylyl, and the like.
  • a “heteroaryl” group is an “aryl” group as just defined in which from 1 to 4 of the ring carbon atoms have been replaced by O, S or NR (where R is hydrogen or Cj. 6 alkyl), e.g. pyrrolyl, furanyl, thienyl, benzofuranyl, and the like.
  • a "substituted aryl or heteroaryl” is an aryl or heteroaryl group as just defined substituted by 1 to 3, preferably adjacent, hydroxyl groups, and up to 5 non- interfering substituents.
  • a non-interfering substituent is a substituent that does not adversely affect the pharmacological activity of the compound and is not otherwise pharmacologically undesirable. Suitable non-interfering substituents include halogen, and C ⁇ alkyl and C ⁇ alkoxy, each optionally substituted with up to five halogen atoms.
  • Disclosed compounds for pharmacological or pharmaceutical treatment of an amyloid disease, or for treatment of ⁇ -synuclein/NAC fibrillogenesis will include and not be limited to proanthocyanidins, procyanidins, anthocyanins, condensed tannins, leucoanthocyanidins, leucocyanins, anthocyanogens, epicatechin-catechin polymers or oligomers, flavanoids, flavan- 3,4-diols, propelargonidins, and A-type, B-type and C-type procyanidins.
  • Uncaria tomentosa compounds and extracts exhibit potent anti-amyloid activity.
  • the individual compounds found to exhibit such activity belong to the general class of compounds known as polyphenols, and more specifically procyanidins and proanthocyanidins.
  • the extracts and compounds having anti-amyloid and anti- ⁇ -synuclein/NAC inhibitory activity can be purified by a variety of methods disclosed herein, including solvent extraction techniques, gel permeation chrQmatography, preparative high performance liquid chromatography or a combination of such techniques.
  • Anti-amyloid and anti- ⁇ -synuclein/NAC compositions and compounds containing the proanthocyanidins can be prepared in accordance with standard techniques well known to those skilled in the pharmaceutical art. Such compositions can be administered in dosages and by techniques well known to those skilled in the art taking into consideration such factors such as age, sex, weight, and condition of the particular patient, and the route of administration. The compositions can be co-administered or sequentially administered with other potential anti- amyloid agents, or anti- ⁇ -synuclein/NAC agents; again taking into consideration such factors as the age, sex, weight and condition of the particular patient, and the route of administration.
  • compositions useful to effect the disclosed aims include solid compositions for oral administration such as capsules, tablets, pills, and the like, as well as chewable solid formulations, to which the present invention may be well suited; liquid preparations for orifice, e.g. oral, nasal, administration such as suspensions, syrups or elixers; and preparations for parental, subcutaneous, intradermal, intramuscular or intravenous administration (e.g. injectable administration) such as sterile suspensions or emulsions.
  • the active proanthocyanidin compound may be in admixture with a suitable carrier, diluent, or excipient such as sterile water, physiological saline or the like.
  • the active anti-amyloid compounds of the invention can be provided in lyophilized form for reconstituting, for instance, in isotonic, aqueous, saline buffer.
  • These compounds can be purified, e.g., compounds or combinations thereof can be substantially pure; for instance, purified to apparent homogeneity. Purity is a relative concept, and the numerous Examples demonstrate isolation of inventive compounds or combinations thereof, as well as purification thereof, such by the methods exemplified a skilled artisian can obtain a substantially pure compound or combination thereof, or purify them to apparent homogeneity (e.g., purity by HPLC; observation of a single chromatographic peak).
  • a substantially pure compound or combination of compounds is at least about 70% pure, more advantageously at least 80-% pure, at least 90% pure, more preferably greater than 90% pure, e.g., at least 90-95% pure, or even purer such as greater than 95% pure, e.g., 99.99% pure.
  • Polyphenols, (+)-catechin and (-)-epicatechin are used herein to exemplify the types of polyphenol oligomers that may be prepared by the method of the present invention.
  • the linkages between adjacent, the polyphenol monomers, (+)-catechin and (-)-epicatechin are from position 4 to position 6 or from position 4 to position 8; and this linkage between position 4 of a monomer and position 6 and 8 of the adjacent monomeric units is designated herein as (4->6) or (4 ⁇ 8).
  • stereoisomers of the oligomers are encompassed within the scope of the invention.
  • the stereochemistry of the substituents on a flavanoid monomer of the oligomer may be described in terms of their relative stereochemistry, "alpha/beta” or “cis/trans”, or in the terms of the absolute stereochemistry, R/S.
  • alpha indicates that the substituent is oriented below the plane of the flavan ring
  • beta indicates that the substituent is oriented above the plane of the ring.
  • cis indicates that the two substituents are oriented on the same face of the ring
  • trans indicates that the two substituents are oriented on opposite faces of the ring. .
  • R and S are used to denote the arrangement of the substituents about a sterogenic or "chiral" center, based on the ranking of the groups according to the atomic number of the atoms directly attached to that stereogenic center.
  • the polyphenol, (+)-catechin may be defined as (2R, trans)-2-(3',4'- dihydroxyphenyl)-3,4-dihydo-2H-l-benzopyran-3,5,7-triol, or as (2R, 3S)-flavan-3,3', 4', 5,7- pentaol.
  • Interflavan (polyphenol-polyphenol) bonding is often characterized using the relative terms ⁇ / ⁇ or cis/trans; ⁇ / ⁇ is used herein to designate the relative stereochemistry of the interflavan bonding.
  • stereochemcial linkages between position 4 of a monomer and position 6 and 8 of the adjacent monomer there are multiple stereochemical linkages between position 4 of a monomer and position 6 and 8 of the adjacent monomer; and the stereochemcial linkages between monomeric units is designated as (4 ⁇ ->6) or (4 ⁇ — >6) or (4 ⁇ — >8) or (4 ⁇ — 8) for linear oligomers.
  • catechin is linked to another catechin or epicatechin
  • the linkages are advantageously (4 ⁇ — 6) or (4 ⁇ 8).
  • epicatechin is linked to catechin or another epicatechin
  • the linkages are advantageously (4 ⁇ — >6) or (4 ⁇ — >8).
  • a bond to carbon position 2 has alpha or beta stereochemistry
  • a bond to carbon position 3 has alpha or beta stereochemistry (e.g., (-)- epicatechin or (+)-catechin).
  • Examples of preferred compounds include, but are not limited to, dimers, epicatechin- 4 ⁇ — >8-epicatechin and epicatechin-4 ⁇ — >6-epicatechin, wherein epicatechin-4 ⁇ — >8-epicatechin is preferred; trimers, [epicatechin-(4 ⁇ - 8)] 2 -epicatechin, [epicatechin-(4 ⁇ 8)] 2 -catechin and [epicatechin-(4 ⁇ — 6)] 2 -epicatechin, wherein [epicatechin-(4 ⁇ — >8)] 2 -epicatechin is preferred; tetramers, [epicatechin-(4 ⁇ — >8)] 3 -e ⁇ icatechin; [epicatechin-(4 ⁇ — >8)] 3 -catechin; and [epicatechin-(4 ⁇ — >8)] 2 -epicatechin-(4 ⁇ 6)-catechin, wherein [epicatechin-(4 ⁇ 8)] 3 - epicatechin is preferred; and pentamers,
  • Proanthocyanidins can not only be extracted and purified from Uncaria tomentosa as described in the present invention, but also from other various plants such as grape, kaki (Japanese persimmon), betel palm, apple, barley, cocoa leaf, cocoa liqueur, dark chocolate, Nest-leaf, rhubarb, cinnamon, adzuki bean, raspberry, etc. They can also be obtained by conventional chemical synthesis.
  • procyanidins or proanthocyanidins derived from Uncaria tomentosa are disclosed, persons skilled in the art will appreciate by means of this disclosure and envision synthetic and alternate extraction routes to obtain the active compounds. Accordingly, synthetic polyphenols or procyanidins or proanthocyanidins or their derivatives which include, but are not limited to glycosides, gallates, esters, and the like are included within the scope of the invention.
  • proanthocyanidins are potent inhibitors of amyloid and ⁇ -synuclein/NAC fibrillogenesis, and cause a potent disruption/disassembly of pre-formed fibrils for a variety of amyloid and ⁇ -synuclein diseases.
  • Exemplary compounds identified to serve as potent amyloid fibril inhibiting agents include procyanidins, such as epicatechin-epicatechin, catechin-epicatechin, epiafzelechin-epicatechin dimers, epicatechin-epicatechin-epicatechin trimers, as well as other epicatechin and/or catechin oligomers for the treatment of amyloid diseases including, but not limited to, Alzheimer's disease, type II diabetes, and systemic AA amyloidosis, as well as inhibiting ⁇ -synuclein or non-amyloid component (NAC) fibril formation for the treatment of Parkinson's and Lewy body disease.
  • procyanidins such as epicatechin-epicatechin, catechin-epicatechin, epiafzelechin-epicatechin dimers, epicatechin-epicatechin-epicatechin trimers, as well as other epicatechin and/or catechin oligomers for the treatment of amyloid diseases including, but not limited to,
  • This invention is also directed to methods for inhibiting or eliminating amyloid fibril formation, deposition, accumulation and/or persistence in a number of different amyloid diseases by treatment of patients with proanthocyanidins of the A, B and C types, including monomers, dimers, trimers and multimers of epicatechin and catechin.
  • An exemplary procyanidin compound is a substituted epicatechin-epicatechin or catechin-epicatechin dimer, such as epicatechin-4 ⁇ — »8-epicatechin, catechin-4 ⁇ 8-epicatechin, or epiafzelechin-4 ⁇ -»8- epicatechin, or other oligomers.
  • amyloid-inhibiting compounds derived from plant material are disclosed for the therapeutic intervention of Alzheimer's disease, type 2 diabetes, Parkinson's disease, systemic AA amyloidosis and other diseases involving amyloid fibril formation and accumulation, especially methods of isolating amyloid inhibiting compounds from Uncaria tomentosa and related plants, and to the use of those compounds.
  • the disclosed compounds act to inhibit or prevent amyloid fibril formation, inhibit or prevent amyloid fibril growth, and/or cause disassembly, disruption, and/or disaggregation of preformed amyloid fibrils and amyloid protein deposits.
  • Their activity can be measured in vitro by methods such as those discussed in Examples 4 through 7 while their activity in vivo against amyloidoses can be measured in animal models, such as those of Alzheimer's disease and in humans by a method such as that discussed in Example 11.
  • the disclosed compounds also act to inhibit or prevent ⁇ -synuclein/NAC fibril formation, inhibit or prevent ⁇ -synuclein/NAC fibril growth, and/or cause disassembly, disruption, and/or disaggregation of preformed ⁇ -synuclein/NAC fibrils and ⁇ - synuclein/NAC-associated protein deposits.
  • Their activity can be measured in vitro by methods similar to those discussed in Examples 4 through 7 below.
  • the therapeutic ratio of a compound can be determined, for example, by comparing the dose that gives effective anti-fibril (anti-amyloid or anti- ⁇ -synuclein NAC activity in a suitable in vivo model in a suitable animal species such as the mouse, with the dose that gives significant weight loss (or other observable side-effects) in the test animal species.
  • compounds will be administered in pure isolated form in therapeutically effective amounts by any of the usual modes known in the art, either singly or in combination with at least one other compound of this invention and/or at least one other conventional therapeutic agent for the disease being treated.
  • a therapeutically effective amount may vary widely depending on the disease, its severity, the age and relative health of the animal being treated, the potency of the compound(s), and other factors.
  • therapeutically effective amounts of compounds of this invention may range from 1-1000 mg/Kg body weight; for example, 10-100 mg/Kg.
  • a person of ordinary skill in the art will be conventionally able, and without undue experimentation, having regard to that skill and to this disclosure, to determine a therapeutically effective amount of a compound for the treatment of amyloidosis or ⁇ -synuclein/NAC fibril formation.
  • compositions will be administered as pharmaceutical compositions by one of the following routes: oral, topical, systemic (e.g. transdermal, intranasal, or by suppository), or parenteral (e.g. intramuscular, subcutaneous, or intravenous injection).
  • routes e.g. oral, topical, systemic (e.g. transdermal, intranasal, or by suppository), or parenteral (e.g. intramuscular, subcutaneous, or intravenous injection).
  • routes e.g. intramuscular, subcutaneous, or intravenous injection.
  • routes e.g. intramuscular, subcutaneous, or intravenous injection.
  • routes e.g. intramuscular, subcutaneous, or intravenous injection.
  • Compositions may take the form of tablets, pills, capsules, semisolids, powders, sustained release formulations, solutions, suspensions, elixirs, aerosols, or any other appropriate compositions; and comprise at least one compound of this invention in combination with at
  • Suitable excipients are well known to persons of ordinary skill in the art, and they, and the methods of formulating the compositions, may be found in such standard references as Alfonso AR: Remington's Pharmaceutical Sciences. 17th ed., Mack Publishing Company, Easton PA, 1985.
  • Suitable liquid carriers, especially for injectable solutions include water, aqueous saline solution, aqueous dextrose solution, and glycols.
  • the compound(s) optimally only one such compound is administered in any particular dosage form — can be administered, orally, for example, as tablets, troches, lozenges, aqueous or oily suspension, dispersible powders or granules, emulsions, hard or soft capsules, or syrups or elixirs.
  • Compositions intended for oral use may be prepared according to any method known in the art for the manufacture of pharmaceutical compositions and such compositions may contain one or more agents selected from the group consisting of sweetening agents, flavoring agents, coloring agents and preserving agents in order to provide pharmaceutically elegant and palatable preparations.
  • Tablets contain the compound in admixture with non-toxic pharmaceutically acceptable excipients that are suitable for the manufacture of tablets.
  • excipients may be for example, inert diluents, such as calcium carbonate, sodium carbonate, lactose, calcium phosphate or sodium phosphate; granulating and disintegrating agents, for example, maize starch or alginic acid; binding agents, for example, maize starch, gelatin or acacia, and lubricating agents, for example, magnesium stearate or stearic acid or tale.
  • the tablets may be uncoated or they may be coated by known techniques to delay disintegration and absorption in the gastrointestinal tract and thereby provide a sustained action over a longer period.
  • a time delay material such as glycerol monostearate or glycerol distearate may be employed.
  • Formulations for oral use may also be presented as hard gelatin capsules wherein the compound is mixed with an inert solid diluent, for example, calcium carbonate, calcium phosphate or kaolin, or as soft gelatin capsules wherein the active ingredient is mixed with water or an oil medium, for example, peanut oil, liquid paraffin or olive oil.
  • Aqueous suspensions contain the compound in admixture with excipients suitable for the manufacture of aqueous suspensions.
  • excipients are suspending agents, for example, sodium carboxymethylcellulose, methylcellulose, hydroxypropylmethyl cellulose, sodium alginate, polyvinylpyrrolidone, gum tragacanth and gum acacia; dispersing or wetting agents may be naturally occurring phosphatides, for example lecithin, or condensation products of an alkylene oxide with fatty acids, for example polyoxyethylene stearate, or condensation products of ethylene oxide with long chain aliphatic alcohols, for example, heptadecaethyleneoxycetanol, or condensation products of ethylene oxide with partial esters derived from fatty acids such as hexitol such as polyoxyethylene sorbitol monooleate, or condensation products of ethylene oxide with partial esters from fatty acids and a hexitol annhydrides, for example, polyethylene sorbitan monooleate.
  • dispersing or wetting agents may be naturally occurring phosphatides, for example lecithin, or condensation products of an alkylene oxide
  • the aqueous suspensions may also contain one or more preservatives, for example, ethyl or n-propyl p-hydroxybenzoate, one or more coloring agents, one or more flavoring agents, or one or more sweetening agents, such as sucrose or saccharin.
  • preservatives for example, ethyl or n-propyl p-hydroxybenzoate, one or more coloring agents, one or more flavoring agents, or one or more sweetening agents, such as sucrose or saccharin.
  • Oily suspensions may be formulated by suspending the compound in a vegetable oil, for example arachis oil, olive oil, sesame oil, or coconut oil or in a mineral oil such as liquid paraffin.
  • the oily suspensions may contain a thickening agent, for example beeswax, hard paraffin or cetyl alcohol.
  • Sweetening agents, such as those set forth below, and flavoring agents may be added to provide a palatable oral preparation. These compositions may be preserved by the addition of an antioxidant such as ascorbic acid.
  • Dispersible powders and granules suitable for preparation of an aqueous suspension by the addition of water provide the active ingredient in admixture with a dispersing or wetting agent, a suspending agent and one or more preservatives. Suitable dispersing or wetting agents and suspending agents are exemplified by those already described above. Additional excipients, for example sweetening, flavoring and agents, may also be present.
  • the compounds may also be in the form of oil-in-water emulsions.
  • the oily phase may be a vegetable oil, for example olive oil or arachis oils, or a mineral oil, for example liquid paraffin or mixtures of these.
  • Suitable emulsifying agents may be naturally-occurring gums, for example gum acacia or gum tragacanth, naturally occurring phosphatides, for example soy bean, lecithin, and occurring phosphatides, for example soy bean, lecithin, and esters or partial esters derived from fatty acids and hexitol anhydrides, for example sorbitan monooleate, and condensation products of the said partial esters with ethylene oxide, for example polyoxyethylene sorbitan monooleate.
  • the emulsion may also contain sweetening and flavoring agents.
  • Syrups and elixirs may be formulated with sweetening agents, for example, glycerol, sorbitol or sucrose. Such formulations may also contain a demulcent, a preservative and flavoring and coloring agents.
  • the compound can also be administered by injection or infusion, either subcutaneously or intravenously, or intramuscularly, or intrasternally, or intranasally, or by infusion techniques in the form of sterile injectable or oleaginous suspension.
  • the compound may be in the form of a sterile injectable aqueous or oleaginous suspensions.
  • the sterile injectable preparation may also be a sterile injectable solution or suspension in a non-toxic parenterally-acceptable diluent or solvent for example, as a solution in 1,3-butanediol.
  • acceptable vehicles and solvents that may be employed are water, Ringer's solution and isotonic sodium chloride solution.
  • sterile, fixed oils are conventionally employed as a solvent or suspending medium.
  • any bland fixed oils may be conventionally employed including synthetic mono- or diglycerides.
  • fatty acids such as oleic acid find use in the preparation of injectables. Dosage regimens can be adjusted to provide the optimum therapeutic response. For example, several divided dosages may be administered daily or the dosage may be proportionally reduced as indicated by the exigencies of the therapeutic situation.
  • Dosage unit form refers to physically discrete units suited as unitary dosages for the subjects to be treated; each containing a therapeutically effective quantity of the compound and at least one pharmaceutical excipient.
  • a drug product will comprise a dosage unit form within a container that is labeled or accompanied by a label indicating the intended method of treatment, such as the treatment of an amyloid disease, such as Alzheimer's disease, or of a disease associated with ⁇ - synuclein/NAC fibril formation, such as Parkinson's disease.
  • PTI-777 represents a group of approximately 11 major fractions referred to as fraction F, fraction G, fraction H, fraction I, fraction J, fraction Kl, fraction K2, fraction L, fraction M, fraction N and fraction O, which were isolated from the powdered bark of Uncaria tomentosa. Some of these fractions, as demonstrated in the present invention, were further purified to one or two major components as was done with fraction H (now found to contain 2 major components referred to as HI and H2 as described below). Using PTI-777 as a starting point, as described in pending US patent application Serial No. 60/271,777 filed 2/27/2001, PTI- 777 represents a group of approximately 11 major fractions referred to as fraction F, fraction G, fraction H, fraction I, fraction J, fraction Kl, fraction K2, fraction L, fraction M, fraction N and fraction O, which were isolated from the powdered bark of Uncaria tomentosa. Some of these fractions, as demonstrated in the present invention, were further purified to one or two major components as was done
  • fraction 9 was almost pure epicatechin
  • UV ultraviolet
  • IR infra-red
  • the analytical HPLC equipment consisted of a Waters 717 autosampler, 600 pump and controller, and a 2487 UV detector controlled by Omega software. Samples were analyzed by using an RP-18 semi-preparative column (Phenomenex Jupiter 5 ⁇ m C18 300A, 250 x 10 mm) with a guard column (Phenomenex SecurityGuard cartridge containing a C18 ODS 4 x 3 mm, 5 ⁇ m column) fitted at 30°C.Samples (5 ⁇ l) were analyzed using a mobile phase flow rate of 5.0 rnL/min, with UV detection at 280 nm. Solvent A - CH 3 CN containing 0.1% TFA Solvent B - H 2 O containing 0.1% TFA
  • Epicatechin-4 ⁇ 8-epicatechin is also known as procyanidin B2 or proanthocyanidin B2.
  • Our NMR data of H2 match partial NMR data published on procyanidin B2 (Kashiwada et al, Chem. Pharm. Bull. 38:888-893, 1990; Porter et al, J. Chem. Soc. Perkin 1:1217-1221, 1982), and our data of the H2 peracetate ( Figure 10; Table 2) exactly matched the published data on peracetylated procyanidin B2 (Franck et al, ACH Models in Chemistry 136:511-517, 1999).
  • the optical rotation of +29.0° compared to a literature 5 value of +25° showed the absolute stereochemistry to be the same as found previously.
  • the minor component of peak H, referred to as HI, of the PTI-777 extract was also isolated by a series of chromatographic techniques, monitored by HPLC (see Example 1, Experimental Procedures for details).
  • HPLC HPLC tracing
  • Figure 2 The minor component of peak H, referred to as HI, of the PTI-777 extract was also isolated by a series of chromatographic techniques, monitored by HPLC (see Example 1, Experimental Procedures for details).
  • HPLC tracing, Figure 2 The minor component of peak H, referred to as HI, of the PTI-777 extract was also isolated by a series of chromatographic techniques, monitored by HPLC (see Example 1, Experimental Procedures for details).
  • HPLC tracing, Figure 3 An HPLC method was developed to separate the two main components of peak H on a preparative scale)(see HPLC tracing, Figure 3), to give us a mostly pure HI (16 mg)(see HPLC tracing, Figure 4) and pure H2 (23 mg).
  • the structure of the natural product HI was therefore determined to be catechin-4oc ⁇ 8- epicatechin, also known as procyanidin B4 or proanthocyanidin B4 ( Figure 26).
  • Our NMR data on compound HI matched partial NMR data published on procyanidin B4 (Thompson et al, JCS Perkin 1:1387-1399, 1972; Fletcher et al, JCS Perkin 1:1628-1637, 1977) and our data of acetylated compound HI ( Figure 21; Table 3) matched partial NMR data published on peracetylated procyanidin B4 (Thompson et al, JCS Perkin 1:1387-1399, 1972; Fletcher et al, JCS Perkin 1:1628-1637, 1977).
  • a further fraction from the silica gel column which was rich in peak K2 was acetylated as before (in Examples 1 and 2) to enable us to obtain more material for structure elucidation.
  • the peracetate of K2 was purified by column chromatography over silica gel.
  • the positions of the 13 C signals and the small couplings of the H signals of the lower unit was typical of epicatechin, and the positions of the 13 C signals and the small couplings of the 'H signals of the other two units were typical of coupled epicatechin (Fletcher et al, J.C.S. Perkin 1:1628-1637, 1977).
  • the presence of three flavan-3-ol units could be seen from the six 13 C signals in the 60-80 region, as well as a signal at 26.39 for the free C-4 position of the lower unit and signals at 34.36 and 35.04 for the coupled C-4's of the other units.
  • Thioflavin T fluorometry was used to determine the effects of H2, HI, K2 and EDTA (as a negative control) on disassembly/dissolution of pre-formed A ⁇ 1-42 fibrils ( Figure 37).
  • Thioflavin T binds specifically to fibrillar amyloid, and this binding produces a fluorescence enhancement at 485 nm that is directly proportional to the amount of amyloid fibrils formed. The higher the fluorescence the greater the amount of amyloid fibrils formed (Naki et al, Lab. Invest. 65:104-110, 1991; Levine m, Protein Sc. 2:404-410, 1993; Amyloid Int. J. Exp. Clin. Invest. 2:1-6. 1995).
  • pre-fibrillized A ⁇ 1-42 (Bachem Inc) was incubated at 37°C for 1 week either alone, or in the presence of EDTA, H2, HI, or K2 at an A ⁇ :test compound weight ratios of 1:0.1, 1:0.01, 1:0.001 or 1:0.0001.
  • 50 ⁇ l of each incubation mixture was transferred into a 96- well microtiter plate containing 150 ⁇ l of distilled water and 50 ⁇ l of a Thioflavin T solution (i.e. 500mM Thioflavin T in 250 mM phosphate buffer)(pH 6.8).
  • the fluorescence was read at 485 nm (444 nm excitation wavelength) using an ELISA plate fluorometer after subtraction with buffer alone or compound alone, as blank.
  • compound HI caused a dose-dependent disruption/ disassembly of preformed A ⁇ 1-42 fibrils, with a significant 16 +/- 3% disruption when used at an A ⁇ :Hl wt/wt ratio of 1:0.001; a significant 33 +/- 6% disruption when used at an AB:H1 wt/wt ratio of 1:0.01; and a significant 54 +/- 8% disruption when used at an A ⁇ :Hl wt/wt ratio of 1:0.1 (i.e. 1:1 molar ratio).
  • Compound K2 also caused a dose-dependent disruption/disassembly of preformed A ⁇ 1-42 fibrils, with a significant 27 +/- 4% disruption when used at an A ⁇ :K2 wt/wt ratio of 1:0.01; and a significant 60 +/- 19% disruption when used at an AB:K2 wt/wt ratio of 1:0.1 (i.e. 1:1 molar ratio).
  • Circular dichroism (CD) spectroscopy is a method used to determine the effects of test compounds to disrupt pre-formed amyloid fibrils.
  • circular dichroism spectroscopy was used to determine the effects of pure compound H2 (i.e. (epicatechin-4 ⁇ — >8-epicatechin) on disruption of ⁇ -pleated sheet structure of pre-formed A ⁇ 1- 42 and 1-40 fibrils of the types found in the brains of patients with Alzheimer's and related disorders.
  • H2 i.e. (epicatechin-4 ⁇ — >8-epicatechin)
  • a ⁇ 1-42 or A ⁇ 1-40 peptides (Bachem Inc., Torrance, CA) were first dissolved in 2mM NaOH solution, maintaining the pH of these solutions above 10.
  • peptides were then dissolved in PBS containing 10% TFE, and the pH was adjusted to 7.2.
  • a ⁇ 1-40 or A ⁇ 1-42 was incubated in the absence or presence of compound H2 at an AB:H2 weight/weight ratio of 1:0.1 (i.e. molar ratio of 1:1).
  • CD spectra were recorded on a AVIV 202 spectropolarimeter with 50 ⁇ M of A ⁇ and H2 compound mixtures. All spectra were collected with 0.1 cm quartz cell using a thermstated cuvette holder.
  • Wavelength traces were scanned from 260-195 nm at 0.5 nm increments with a bandwidth of 1 nm and averaged over a time of 5 seconds; the temperature was held constant at 25°C. All spectra reported are an average of 4 scans. As shown in Figure 39, A ⁇ 1-42 alone in 10% TFE PBS buffer showed the typical CD spectra of an amyloid protein with significant ⁇ -sheet structure, as demonstrated by the sharp minima observed at 218 nm.
  • Parkinson's disease is a neurodegenerative disorder that is pathologically characterized by the presence of intracytoplasmic Lewy bodies (Lewy in Handbuch der Neurologie. M. Lewandowski, ed., Springer, Berlin, pp. 920-933, 1912; Pollanen et al., J. Neuropath. Exp. Neurol. 52:183-191, 1993), the major components of which are filaments consisting of ⁇ - synuclein (Spillantini et al., Proc. Natl. Acad. Sci. USA_95: 6469-6473, 1998; Arai et al., Neurosc. Lett.
  • ⁇ -Synuclein recombinant protein, and non-amyloid component (known as NAC), which is a 35-amino acid peptide fragment of ⁇ -synuclein, both have the ability to form fibrils when incubated at 37°C, and are positive with amyloid stains such as Congo red (demonstrating a red/green birefringence when viewed under polarized light) and Thioflavin S (demonstrating positive fluorescence) (Hashimoto et al, Brain Res.
  • each incubation mixture was transferred into a 96-well microtiter plate containing 150 ⁇ l of distilled water and 50 ⁇ l of a Thioflavin T solution (i.e. 500mM Thioflavin T in 250 mM phosphate buffer (pH 6.8).
  • the fluorescence was read at 485 nm (444 nm excitation wavelength) using an ELISA plate fluorometer after subtraction of buffer alone as blank.
  • compound HI caused a dose-dependent disruption/ disassembly of preformed NAC fibrils, with a significant 31 +/- 16% disruption when used at a NAC:H1 wt wt ratio of 1:0.01; and a significant 64 +/- 3% disruption when used at a NAC:H1 wt/wt ratio of 0.1 (i.e. 1:1 molar ratio).
  • Compound K2 also caused a dose-dependent disruption/disassembly of preformed NAC fibrils, with a significant 20 +/- 27% disruption when used at an NAC:K2 wt/wt ratio of 1:0.01; and a significant 39 +/- 12% disruption when used at an NAC:K2 wt/wt ratio of 1:0.1 (i.e. 1:1 molar ratio).
  • This study indicated that the proanthocyanidins H2, HI and K2 were also potent disruptors of NAC fibrils, and exerted their effect in a dose-dependent manner. It is expected that similar efficacy of these proanthocyanidins will be also observed for disruption/disassembly of ⁇ -synuclein fibrils.
  • Example 8 Efficacy of Proanthocyanidins H2, HI and K2 as Disruptors of Type 2 Diabetes Amyloid Fibrils
  • Islet amyloid deposits are observed in -90% of patients with well-established type 2 diabetes and would appear to be a characteristic feature of the disease process (Westermark, Med. Sci. 77:91-94,1972; Clark et al, Diabetes Res. 9:151-159,1988). In many patients, the deposits are widespread and affect many islets. The degree of islet (predominantly ⁇ -cell) mass that has been replaced by amyloid may be a marker for the severity of the diabetic disease process, with those individuals requiring insulin treatment having the greatest islet mass reduction and amyloid formation (Westermark, Amyloid: Int. J. Exp. Clin. Invest. 1:47- 60,1994).
  • IAPP islet amyloid polypeptide
  • amylin The major protein in type 2 diabetes islet amyloid is a 37-amino acid peptide known as islet amyloid polypeptide (IAPP) or amylin.
  • IAPP is a known normal secretory product of the pancreatic ⁇ -cells (Kanh et al, Diabetes 39:634-638,1990) that is stored in insulin-bearing cytoplasmic granules (Clark et al, Cell Tissue Res. 257:179-185, 1989).
  • IAPP has been hypothesized to have an important role in the pathogenesis of type 2 diabetes through its impairment of ⁇ -cell function and reduction of ⁇ -cell mass (Johnson et al, N. Engl. J. Med.
  • IAPP islet amyloid
  • compound H2 caused a dose-dependent disruption/disassembly of preformed IAPP fibrils, with a significant (p ⁇ 0.01) 36 +/- 5% disruption when used at a IAPP:H2 wt/wt ratio of 1:0.01; and a significant (p ⁇ 0.01) 83 +/- 1% disruption when used at a IAPP:H2 wt/wt ratio of 1:0.1 (i.e. 1:1 molar ratio).
  • compound HI caused a dose-dependent disruption/ disassembly of preformed IAPP fibrils, with a significant 35 +/- 4% disruption when used at a IAPP:H1 wt/wt ratio of 1:0.01; and a significant 79 +/- 1% disruption when used at a IAPP:H1 wt/wt ratio of 0.1 (i.e. 1 : 1 molar ratio).
  • Compound K2 also caused a dose-dependent disruption/disassembly of preformed IAPP fibrils, with a significant 26 +/- 4% disruption when used at an IAPP:K2 wt/wt ratio of 1:0.01; and a significant 62 +/- 1% disruption when used at an IAPP:K2 wt/wt ratio of 1:0.1 (i.e. 1:1 molar ratio).
  • This study indicated that the proanthocyanidins H2, HI and K2 were also potent disruptors of IAPP fibrils, and exerted their effect in a dose-dependent manner.
  • proanthocyanidins are expected to be useful for the treatment of IAPP amyloidosis in type 2 diabetes.
  • Fractions 38 to 42 contained compound Kl (22 mg) as a pale brown gum.
  • the retention time of this Kl peak was 15.0 minutes as monitored by HPLC Method 1.
  • acetylation of Kl to help determine the structure, a sample of Kl (15 mg) was dissoved in a mixture of acetic anhydride (0.5 ml) and pyridine (0.5 ml). The mixture stood at room temperature for 18 hours, then the solvents were removed in vacuo to give the Kl peracetate (16 mg) as a colourless gum.
  • the NMR data is shown in Table 6 below. Identification of Kl and the Kl peracetate;
  • Kl The minor component of peak K, called Kl, of the PTI-777 extract was isolated by column chromatography over sephadex LH20, monitored by HPLC. Elution with 95% ethanol followed by increasing amounts of acetone and water, followed by methanol, gave pure peak Kl in fractions 38 to 42 (see Table 5).
  • the structure of the Kl peracetate is shown in Figure 43, whereas the structure of Kl is shown in Figure 44. To arrive at these structures, the following analysis and results were obtained.
  • the lower flavan-3-ol unit was shown to be epicatechin by CIGAR correlations from H-2(l) to C-2' and C-6' signals of a 3',4'-dioxygenated aromatic ring.
  • the upper flavan-3-ol unit was identified by CIGAR correlations from H-2(u) to equivalent C-27C-6' signals of a 4'-oxygenated ring. This constitutes an epiafzelechin unit.
  • the structure of the natural product Kl was therefore assigned to be epiafzelechin-4 ⁇ — >8-epicatechin. This compound is a known compound
  • Proanthocyanidins act as potent inhibitors/disruptors and/or causing disassembly of amyloid fibrils (regardless of the type of amyloid protein present. Examples are shown for A ⁇ , NAC and IAPP fibrils), as well as a potent inhibitor/disruptor of ⁇ -synuclein/NAC fibrils. Both procyanidin dimers and trimers are shown specifically to inhibit such fibrillogenesis, and our ongoing studies suggest that procyanidin tetramers and oligomers (greater than tetramers) are also able to exert such amyloid fibril inhibiting effects. Thus, preferred therapeutic applications include the use of proanthocyanidins and procyanidins for the treatment of amyloid diseases, and diseases which include ⁇ -synuclein/NAC fibrillogenesis.
  • proanthocyanidins of the present invention were discovered, isolated and identified from the plant Uncaria tomentosa. However, it is probable that similar amyloid/ ⁇ -synuclein/ NAC inhibitory activity is observed with any proanthocyanidin regardless of the source (i.e. plant or food), and will include proanthocyanidins that can be synthesized by methods known to those knowledgeable and skilled in the art. Preparations of proanthocyanidins compounds for parenteral administration include sterile aqueous or non-aqueous solutions, suspensions, emulsions, which may contain axillary agents or excipients which are known in the art.
  • compositions such as tablets, pills, caplets, soft and hard gelatin capsules, lozenges, sachets, cachets, vegicaps, liquid drops, elixers, suspensions, emulsions, solutions, syrups, tea bags, aerosols (as a solid or in a liquid medium), suppositories, sterile injectible solutions, sterile packaged powders, can be prepared according to routine method and are known in the art.
  • Proanthocyanidins of the present invention may be administered by any means that achieve their intended purpose, for example to treat amyloid diseases, such as Alzheimer's disease or type 2 diabetes, or other pathologies involving ⁇ -synuclein/NAC fibrillogenesis, using a proanthocyanidin described herein, in the form of a pharmaceutical or pharmacological composition.
  • amyloid diseases such as Alzheimer's disease or type 2 diabetes
  • proanthocyanidin described herein in the form of a pharmaceutical or pharmacological composition.
  • administration of such a composition may be by various parenteral routes such as subcutaneous, intravenous, intradermal, intramuscular, intraperitoneal, intranasal, transdermal or buccal routes.
  • parenteral routes such as subcutaneous, intravenous, intradermal, intramuscular, intraperitoneal, intranasal, transdermal or buccal routes.
  • administration may be by the oral route.
  • Parenteral administration can be by bolus injection or by gradual perfusion over time.
  • a preferred mode of using a proanthocyanidin pharmaceutical composition of the present invention is by oral administration or intravenous application.
  • a typical regimen for preventing, suppressing or treating amyloid pathologies, such as Alzheimer's disease amyloidosis comprises administration of an effective amount of a proanthocyanidin over a period of one or several days, up to and including between one week to about 10 years.
  • the dosage of the proanthocyanidin of the present invention administered in vivo or in vitro will be dependent upon the age, sex, health, and weight of the recipient, kind of concurrent treatment, if any, frequency of treatment, and the nature of the effect desired.
  • the most preferred dosage will be tailored to the individual subject, as is understood and determinable by one of skill in the art, without undue experimentation.
  • the total dose required for each treatment may be administered by multiple doses or in a single dose.
  • a proanthocyanidin or procyanidin compound may be administered alone or in conjunction with other therapeutics directed to amyloid disease or ⁇ -synuclein/NAC fibrillogenesis, such as Alzheimer's disease or Parkinson's disease, as described herein.
  • Effective amounts of a proanthocyanidin compound for treatment are about 10 mg to about 1,000 mg/kg body weight, and preferably from about 10 mg to 100 mg/kg body weight, such as 10, 20, 30, 40, 50, 60, 70, 80, 90 or 100 mg/kg body weight.
  • Preparations for parenteral administration include sterile aqueous or non-aqueous solutions, suspensions and emulsions, which may contain axillary agents or excipients that are known in the art.
  • Pharmaceutical compositions containing a proanthocyanidin of the present invention may include all compositions where the proanthocyanidin is contained in an amount effective to achieve its intended purpose.
  • a pharmaceutical composition may contain suitable pharmaceutically acceptable carriers, such as excipients, carriers and/or axillaries that facilitate processing of the active compounds into preparations which can be used pharmaceutically.
  • compositions comprise at least one proanthocyanidin compound may also include solutions for administration intravenously, subcutaneously, dermally, orally, mucosally, rectally, or may be by injection or orally, and contain from about 0.01 to 100 %, preferably about 95-100% of active compound together with the excipient.
  • Pharmaceutical compositions for oral administration include pills, tablets, caplets, soft and hard gelatin capsules, lozenges, sachets, cachets, vegicaps, liquid drops, elixers, suspensions, emulsions, solutions and syrups.
  • the proanthocyanidin compounds for Alzheimer's and Parkinson's disease, and other central nervous system disorders may be optimized to cross the blood-brain barrier.
  • Methods of introductions include but are not limited to systemic administration, parenteral administration, i.e. via an intraperitoneal, intravenous, perioral, subcutaneous, intramuscular, intraarterial, intradermal, intramuscular, intranasal, epidural or oral routes.
  • a proanthocyanidin compound may be directly administered to the cerebrospinal fluid by intraventricular injection.
  • a proanthocyanidin compound may be administered locally to the areas of tissue in need of treatment; this may be achieved by, for example, and not by way of limitation, local infusion during surgery, topical application, by injection, by infusing a cannulae with osmotic pump, by means of a catheter, by means of a suppository, or by means of an implant.
  • a proanthocyanidin compound may be delivered in a controlled release system, such as an osmotic pump.
  • 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.
  • Exam le 11; Clinical Testing in Alzheimer's Patients for Example Five to fifty women are selected for a clinical study.
  • the women are post-menopausal, i.e. have ceased menstruating for between 6 and 12 months prior to the study's initiation, have been diagnosed with early stage Alzheimer's disease, and expected to have worsening symptoms of Alzheimer's disease within the study period, but are in good general health otherwise.
  • the study has a placebo group, i.e. the women are divided into two groups, one of which receives the compound of this invention and the other receives a placebo.
  • the patients are benchmarked as to memory, cognition, reasoning, and other symptoms associated with Alzheimer's disease. Women in the test group receive a therapeutic dose of the compound by the oral route. They continue this therapy for 6-36 months.

Landscapes

  • Health & Medical Sciences (AREA)
  • Animal Behavior & Ethology (AREA)
  • Chemical & Material Sciences (AREA)
  • Veterinary Medicine (AREA)
  • Medicinal Chemistry (AREA)
  • Public Health (AREA)
  • General Health & Medical Sciences (AREA)
  • Pharmacology & Pharmacy (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • General Chemical & Material Sciences (AREA)
  • Organic Chemistry (AREA)
  • Nuclear Medicine, Radiotherapy & Molecular Imaging (AREA)
  • Chemical Kinetics & Catalysis (AREA)
  • Engineering & Computer Science (AREA)
  • Bioinformatics & Cheminformatics (AREA)
  • Neurosurgery (AREA)
  • Biomedical Technology (AREA)
  • Neurology (AREA)
  • Diabetes (AREA)
  • Hematology (AREA)
  • Emergency Medicine (AREA)
  • Epidemiology (AREA)
  • Obesity (AREA)
  • Oncology (AREA)
  • Endocrinology (AREA)
  • Psychology (AREA)
  • Hospice & Palliative Care (AREA)
  • Psychiatry (AREA)
  • Pain & Pain Management (AREA)
  • Rheumatology (AREA)
  • Pharmaceuticals Containing Other Organic And Inorganic Compounds (AREA)
  • Medicines Containing Plant Substances (AREA)
  • Pyrane Compounds (AREA)

Abstract

La présente invention concerne une technique de traitement d'une amylose ou d'une maladie caractérisée par l'α-synucléine ou par une fibrillogenèse NAC (à composante non amyloïde) chez un mammifère. Cette technique consiste à administrer à ce mammifère une quantité thérapeutiquement efficace de diverses proanthocyanidines de cette invention ou d'une proanthocyanidine caractérisée par les formules générales données dans les spécifications. Cette invention concerne aussi une composition pharmaceutique comprenant une quantité thérapeutiquement efficace de proanthocyanidine et un excipient répondant aux normes pharmaceutiques. Cette quantité thérapeutique de proanthocyanidine est sélectionnée pour son efficacité dans le traitement des amyloses, des maladies induites par l'α-synucléine ou par des fibrillogenèses NAC chez un mammifère.
EP02719009A 2001-03-15 2002-02-15 Proanthocyanidines destinees au traitement des amyloses et des maladies induites par alpha-synucleine Withdrawn EP1377287A4 (fr)

Applications Claiming Priority (15)

Application Number Priority Date Filing Date Title
US27686601P 2001-03-15 2001-03-15
US276866P 2001-03-15
US938987 2001-08-24
US09/938,987 US6607758B2 (en) 1997-05-15 2001-08-24 Methods for inhibiting and reducing amyloid fibril formation associated with Alzheimer's Disease and other amyloidoses
US10/053,625 US6929808B2 (en) 2000-11-03 2001-11-02 Methods of isolating amyloid-inhibiting compounds and use of compounds isolated from Uncaria tomentosa and related plants
WOPCT/US05/01131 2001-11-02
US53625 2001-11-02
PCT/US2001/051131 WO2002042429A2 (fr) 2000-11-03 2001-11-02 Procedes d'isolement de composes inhibiteurs d'amyloide, et utilisation de composes utilises a partir d'uncaria tomentosa et de plantes parentes
US33872101P 2001-12-04 2001-12-04
US338721P 2001-12-04
US33903301P 2001-12-10 2001-12-10
US33896901P 2001-12-10 2001-12-10
US339033P 2001-12-10
US338969P 2001-12-10
PCT/US2002/004764 WO2002076381A2 (fr) 2001-03-15 2002-02-15 Proanthocyanidines destinees au traitement des amyloses et des maladies induites par alpha-synucleine

Publications (2)

Publication Number Publication Date
EP1377287A2 true EP1377287A2 (fr) 2004-01-07
EP1377287A4 EP1377287A4 (fr) 2007-11-14

Family

ID=56290250

Family Applications (1)

Application Number Title Priority Date Filing Date
EP02719009A Withdrawn EP1377287A4 (fr) 2001-03-15 2002-02-15 Proanthocyanidines destinees au traitement des amyloses et des maladies induites par alpha-synucleine

Country Status (6)

Country Link
EP (1) EP1377287A4 (fr)
JP (1) JP4380991B2 (fr)
AU (1) AU2002250117B2 (fr)
CA (1) CA2441099C (fr)
NZ (1) NZ528322A (fr)
WO (1) WO2002076381A2 (fr)

Families Citing this family (12)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US20070208087A1 (en) 2001-11-02 2007-09-06 Sanders Virginia J Compounds, compositions and methods for the treatment of inflammatory diseases
US7514107B2 (en) * 2002-03-21 2009-04-07 Mars, Incorporated Treatment of diseases involving defective gap junctional communication
SI2527315T1 (sl) 2002-05-31 2014-06-30 Proteotech Inc., Spojine, sestavki in metode za zdravljenje amiloidnih bolezni in sinukleinopatij, kot so Alzheimerjeva bolezen, diabetes tipa 2 in Parkinsonova bolezen
EP1790648A1 (fr) * 2002-10-11 2007-05-30 Proteotech, Inc. Utilisation de procyanidine B2 pour la preparation de médicament pour le traitement des maladies amyloides et synucleines
JP2006232670A (ja) * 2003-05-20 2006-09-07 Ajinomoto Co Inc 神経障害用薬剤
WO2005082351A1 (fr) * 2004-03-02 2005-09-09 Eisai R & D Management Co., Ltd. Agent thérapeutique pour un trouble neurodégénératif
US8263589B2 (en) * 2006-03-29 2012-09-11 Wista Laboratories Ltd. Inhibitors of protein aggregation
WO2009104556A1 (fr) * 2008-02-19 2009-08-27 株式会社岐阜セラツク製造所 Composition
CN105920216A (zh) * 2008-05-09 2016-09-07 西奈山医学院 用于预防和治疗神经退行性疾病的方法
US20140179774A1 (en) * 2012-12-26 2014-06-26 Industrial Technology Research Institute Methods for inhibition of shc-1/p66 to combat aging-related diseases
CA2983091A1 (fr) 2015-05-11 2016-11-17 Meiji Co., Ltd. Composition pour faciliter la production du facteur neurotrophique derive du cerveau
KR102441381B1 (ko) * 2016-07-18 2022-09-07 (주)아모레퍼시픽 3-O-갈로일-3,3',5,5',7-펜타하이드록시플라반(3-O-galloyl-3,3',5,5',7- pentahydroxyflavan)을 포함하는 인지 기능 개선용 조성물

Citations (4)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US5922756A (en) * 1998-02-14 1999-07-13 Chan; Marion Man-Ying Method of inhibiting nitric oxide synthase
WO1999045797A1 (fr) * 1998-03-12 1999-09-16 Mars, Incorporated Produits contenant du ou des polyphenols et de l'arginine-l favorisant la production d'oxyde nitrique
WO2000030666A1 (fr) * 1998-11-24 2000-06-02 University Of Washington Composition et procedes pour inhiber la formation de depots de substances amyloides dans le cerveau
WO2001049307A1 (fr) * 1999-12-30 2001-07-12 Proteotech, Inc. Catechines et extrait de the vert destines au traitement de l'amyloidose dans la maladie d'alzheimer et d'autres amyloidoses

Family Cites Families (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US4698360B1 (en) * 1985-04-09 1997-11-04 D Investigations Pharmacologiq Plant extract with a proanthocyanidins content as therapeutic agent having radical scavenger effect and use thereof
CA2289531C (fr) * 1997-05-15 2009-09-29 University Of Washington Composition et procedes de traitement de la maladie d'alzheimer et autres amyloidoses
WO2000057707A1 (fr) * 1999-03-15 2000-10-05 Proteotech, Inc. Methodes de traitement de la maladie d'alzheimer et d'autres amyloses avec l'hypericum perforatum et ses derives

Patent Citations (4)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US5922756A (en) * 1998-02-14 1999-07-13 Chan; Marion Man-Ying Method of inhibiting nitric oxide synthase
WO1999045797A1 (fr) * 1998-03-12 1999-09-16 Mars, Incorporated Produits contenant du ou des polyphenols et de l'arginine-l favorisant la production d'oxyde nitrique
WO2000030666A1 (fr) * 1998-11-24 2000-06-02 University Of Washington Composition et procedes pour inhiber la formation de depots de substances amyloides dans le cerveau
WO2001049307A1 (fr) * 1999-12-30 2001-07-12 Proteotech, Inc. Catechines et extrait de the vert destines au traitement de l'amyloidose dans la maladie d'alzheimer et d'autres amyloidoses

Non-Patent Citations (1)

* Cited by examiner, † Cited by third party
Title
See also references of WO02076381A2 *

Also Published As

Publication number Publication date
AU2002250117B2 (en) 2007-11-29
WO2002076381A3 (fr) 2002-11-21
NZ528322A (en) 2005-03-24
CA2441099C (fr) 2011-04-19
EP1377287A4 (fr) 2007-11-14
WO2002076381A2 (fr) 2002-10-03
JP2005505497A (ja) 2005-02-24
JP4380991B2 (ja) 2009-12-09
CA2441099A1 (fr) 2002-10-03
AU2002250117B8 (en) 2002-10-08

Similar Documents

Publication Publication Date Title
US20100010079A1 (en) Proanthocyanidins for the Treatment of Amyloid and Alpha-Synuclein Diseases
US9133176B2 (en) Isolation, purification and synthesis of procyanidin B2 and uses thereof
US6929808B2 (en) Methods of isolating amyloid-inhibiting compounds and use of compounds isolated from Uncaria tomentosa and related plants
Snow et al. The Amazon rain forest plant Uncaria tomentosa (cat’s claw) and its specific proanthocyanidin constituents are potent inhibitors and reducers of both brain plaques and tangles
US6939570B1 (en) Composition and methods for treating Alzheimer's disease and other amyloidoses
AU2002239764A1 (en) Methods of isolating amyloid-inhibiting compounds and use of compounds isolated from uncaria tomentosa and related plants
US6264994B1 (en) Compositions for treating alzheimer's disease and other amyloidoses
US6346280B1 (en) Composition and methods for inhibiting the formation of brain amyloid deposits
CA2441099C (fr) Proanthocyanidines destinees au traitement des amyloses et des maladies induites par alpha-synucleine
AU2002250117A1 (en) Proanthocyanidins for the treatment of amyloid and alpha-synuclein diseases
US20100197778A1 (en) Proanthocyanidins for the treatment of amyloid and alpha-synuclein diseases
EP1921076A1 (fr) synthèse de procyanidine B2
Kastin et al. the Amazon rain forest plant Uncaria tomentosa (cat’s claw) and its specific proanthocyanidin constituents are potent inhibitors and reducers of both brain plaques

Legal Events

Date Code Title Description
PUAI Public reference made under article 153(3) epc to a published international application that has entered the european phase

Free format text: ORIGINAL CODE: 0009012

17P Request for examination filed

Effective date: 20030924

AK Designated contracting states

Kind code of ref document: A2

Designated state(s): AT BE CH CY DE DK ES FI FR GB GR IE IT LI LU MC NL PT SE TR

AX Request for extension of the european patent

Extension state: AL LT LV MK RO SI

RIN1 Information on inventor provided before grant (corrected)

Inventor name: NGUYEN, BETH, P.

Inventor name: CHOI, PAULA, Y.

Inventor name: CASTILLO, GERALDO, M.

Inventor name: SNOW, ALAN, D.

RIC1 Information provided on ipc code assigned before grant

Ipc: A61P 3/10 20060101ALI20070717BHEP

Ipc: A61P 25/28 20060101ALI20070717BHEP

Ipc: A61P 25/16 20060101ALI20070717BHEP

Ipc: A61K 31/353 20060101ALI20070717BHEP

Ipc: A61K 31/352 20060101ALI20070717BHEP

Ipc: A61K 31/35 20060101AFI20070717BHEP

A4 Supplementary search report drawn up and despatched

Effective date: 20071016

17Q First examination report despatched

Effective date: 20090618

STAA Information on the status of an ep patent application or granted ep patent

Free format text: STATUS: THE APPLICATION IS DEEMED TO BE WITHDRAWN

18D Application deemed to be withdrawn

Effective date: 20120925