WO2010101949A1 - Quinoléines substituées en 8 et analogues associés utilisés comme modulateurs de sirtuine - Google Patents

Quinoléines substituées en 8 et analogues associés utilisés comme modulateurs de sirtuine Download PDF

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Publication number
WO2010101949A1
WO2010101949A1 PCT/US2010/025963 US2010025963W WO2010101949A1 WO 2010101949 A1 WO2010101949 A1 WO 2010101949A1 US 2010025963 W US2010025963 W US 2010025963W WO 2010101949 A1 WO2010101949 A1 WO 2010101949A1
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Prior art keywords
alkyl
substituted
fluoro
heterocycle
independently selected
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PCT/US2010/025963
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English (en)
Inventor
Chi B. Vu
Rebecca L. Casaubon
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Sirtris Pharmaceuticals, Inc.
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Priority to BRPI1011477A priority Critical patent/BRPI1011477A2/pt
Priority to EP10749227A priority patent/EP2403833A4/fr
Priority to CN2010800194505A priority patent/CN102414180A/zh
Priority to AU2010221417A priority patent/AU2010221417A1/en
Priority to CA2754058A priority patent/CA2754058A1/fr
Priority to JP2011553053A priority patent/JP2012519211A/ja
Priority to MX2011009213A priority patent/MX2011009213A/es
Priority to EA201171098A priority patent/EA201171098A1/ru
Publication of WO2010101949A1 publication Critical patent/WO2010101949A1/fr
Priority to IL214943A priority patent/IL214943A0/en

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    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D471/00—Heterocyclic compounds containing nitrogen atoms as the only ring hetero atoms in the condensed system, at least one ring being a six-membered ring with one nitrogen atom, not provided for by groups C07D451/00 - C07D463/00
    • C07D471/02—Heterocyclic compounds containing nitrogen atoms as the only ring hetero atoms in the condensed system, at least one ring being a six-membered ring with one nitrogen atom, not provided for by groups C07D451/00 - C07D463/00 in which the condensed system contains two hetero rings
    • C07D471/04—Ortho-condensed systems
    • 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
    • A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
    • A61K31/435—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with one nitrogen as the only ring hetero atom
    • A61K31/47—Quinolines; Isoquinolines
    • A61K31/4709—Non-condensed quinolines and containing further heterocyclic rings
    • 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
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    • 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
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    • A61P3/00—Drugs for disorders of the metabolism
    • 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
    • A61P3/04—Anorexiants; Antiobesity agents
    • 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
    • A—HUMAN NECESSITIES
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    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P43/00—Drugs for specific purposes, not provided for in groups A61P1/00-A61P41/00
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P5/00—Drugs for disorders of the endocrine system
    • A61P5/48—Drugs for disorders of the endocrine system of the pancreatic hormones
    • A61P5/50—Drugs for disorders of the endocrine system of the pancreatic hormones for increasing or potentiating the activity of insulin
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P7/00—Drugs for disorders of the blood or the extracellular fluid
    • A61P7/02—Antithrombotic agents; Anticoagulants; Platelet aggregation inhibitors
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
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    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D215/00—Heterocyclic compounds containing quinoline or hydrogenated quinoline ring systems
    • C07D215/02—Heterocyclic compounds containing quinoline or hydrogenated quinoline ring systems having no bond between the ring nitrogen atom and a non-ring member or having only hydrogen atoms or carbon atoms directly attached to the ring nitrogen atom
    • C07D215/16—Heterocyclic compounds containing quinoline or hydrogenated quinoline ring systems having no bond between the ring nitrogen atom and a non-ring member or having only hydrogen atoms or carbon atoms directly attached to the ring nitrogen atom with hetero atoms or with carbon atoms having three bonds to hetero atoms with at the most one bond to halogen, e.g. ester or nitrile radicals, directly attached to ring carbon atoms
    • C07D215/48—Carbon atoms having three bonds to hetero atoms with at the most one bond to halogen
    • C—CHEMISTRY; METALLURGY
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    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D401/00—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, at least one ring being a six-membered ring with only one nitrogen atom
    • C07D401/02—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, at least one ring being a six-membered ring with only one nitrogen atom containing two hetero rings
    • C07D401/12—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, at least one ring being a six-membered ring with only one nitrogen atom containing two hetero rings linked by a chain containing hetero atoms as chain links
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D401/00—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, at least one ring being a six-membered ring with only one nitrogen atom
    • C07D401/14—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, at least one ring being a six-membered ring with only one nitrogen atom containing three or more hetero rings
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D403/00—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, not provided for by group C07D401/00
    • C07D403/02—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, not provided for by group C07D401/00 containing two hetero rings
    • C07D403/12—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, not provided for by group C07D401/00 containing two hetero rings linked by a chain containing hetero atoms as chain links
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D405/00—Heterocyclic compounds containing both one or more hetero rings having oxygen atoms as the only ring hetero atoms, and one or more rings having nitrogen as the only ring hetero atom
    • C07D405/02—Heterocyclic compounds containing both one or more hetero rings having oxygen atoms as the only ring hetero atoms, and one or more rings having nitrogen as the only ring hetero atom containing two hetero rings
    • C07D405/12—Heterocyclic compounds containing both one or more hetero rings having oxygen atoms as the only ring hetero atoms, and one or more rings having nitrogen as the only ring hetero atom containing two hetero rings linked by a chain containing hetero atoms as chain links
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D405/00—Heterocyclic compounds containing both one or more hetero rings having oxygen atoms as the only ring hetero atoms, and one or more rings having nitrogen as the only ring hetero atom
    • C07D405/14—Heterocyclic compounds containing both one or more hetero rings having oxygen atoms as the only ring hetero atoms, and one or more rings having nitrogen as the only ring hetero atom containing three or more hetero rings
    • C—CHEMISTRY; METALLURGY
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    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D413/00—Heterocyclic compounds containing two or more hetero rings, at least one ring having nitrogen and oxygen atoms as the only ring hetero atoms
    • C07D413/14—Heterocyclic compounds containing two or more hetero rings, at least one ring having nitrogen and oxygen atoms as the only ring hetero atoms containing three or more hetero rings
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    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D417/00—Heterocyclic compounds containing two or more hetero rings, at least one ring having nitrogen and sulfur atoms as the only ring hetero atoms, not provided for by group C07D415/00
    • C07D417/02—Heterocyclic compounds containing two or more hetero rings, at least one ring having nitrogen and sulfur atoms as the only ring hetero atoms, not provided for by group C07D415/00 containing two hetero rings
    • C07D417/04—Heterocyclic compounds containing two or more hetero rings, at least one ring having nitrogen and sulfur atoms as the only ring hetero atoms, not provided for by group C07D415/00 containing two hetero rings directly linked by a ring-member-to-ring-member bond
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D417/00—Heterocyclic compounds containing two or more hetero rings, at least one ring having nitrogen and sulfur atoms as the only ring hetero atoms, not provided for by group C07D415/00
    • C07D417/02—Heterocyclic compounds containing two or more hetero rings, at least one ring having nitrogen and sulfur atoms as the only ring hetero atoms, not provided for by group C07D415/00 containing two hetero rings
    • C07D417/12—Heterocyclic compounds containing two or more hetero rings, at least one ring having nitrogen and sulfur atoms as the only ring hetero atoms, not provided for by group C07D415/00 containing two hetero rings linked by a chain containing hetero atoms as chain links
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D417/00—Heterocyclic compounds containing two or more hetero rings, at least one ring having nitrogen and sulfur atoms as the only ring hetero atoms, not provided for by group C07D415/00
    • C07D417/14—Heterocyclic compounds containing two or more hetero rings, at least one ring having nitrogen and sulfur atoms as the only ring hetero atoms, not provided for by group C07D415/00 containing three or more hetero rings

Definitions

  • the Silent Information Regulator (SIR) family of genes represents a highly conserved group of genes present in the genomes of organisms ranging from archaebacteria to a eukaryotes.
  • the encoded SIR proteins are involved in diverse processes from regulation of gene silencing to DNA repair.
  • the proteins encoded by members of the SIR gene family show high sequence conservation in a 250 amino acid core domain.
  • a well-characterized gene in this family is S. cerevisiae SIR2, which is involved in silencing HM loci that contain information specifying yeast mating type, telomere position effects and cell aging.
  • the yeast Sir2 protein belongs to a family of histone deacetylases.
  • the Sir2 homolog, CobB, in Salmonella typhimurium functions as an NAD (nicotinamide adenine dinucleo tide) -dependent ADP-ribosyl transferase.
  • the Sir2 protein is a class III deacetylase which uses NAD as a cosubstrate. Unlike other deacetylases, many of which are involved in gene silencing, Sir2 is insensitive to class I and II histone deacetylase inhibitors like trichostatin A (TSA). Deacetylation of acetyl-lysine by Sir2 is tightly coupled to NAD hydrolysis, producing nicotinamide and a novel acetyl-ADP ribose compound.
  • TSA trichostatin A
  • the NAD- dependent deacetylase activity of Sir2 is essential for its functions which can connect its biological role with cellular metabolism in yeast. Mammalian Sir2 homologs have NAD-dependent histone deacetylase activity.
  • SIRT1-SIRT7 are seven Sir2-like genes that share the conserved catalytic domain of Sir2.
  • SIRTl is a nuclear protein with the highest degree of sequence similarity to Sir2.
  • SIRTl regulates multiple cellular targets by deacetylation including the tumor suppressor p53, the cellular signaling factor NF- kB, and the FOXO transcription factor.
  • SIRT3 is a homolog of SIRTl that is conserved in prokaryotes and eukaryotes.
  • the SIRT3 protein is targeted to the mitochondrial cristae by a unique domain located at the N-terminus.
  • SIRT3 has NAD+-dependent protein deacetylase activity and is ubiquitously expressed, particularly in metabolically active tissues. Upon transfer to the mitochondria, SIRT3 is believed to be cleaved into a smaller, active form by a mitochondrial matrix processing peptidase (MPP).
  • MPP mitochondrial matrix processing peptidase
  • Caloric restriction has been known for over 70 years to improve the health and extend the lifespan of mammals.
  • Yeast life span like that of metazoans, is also extended by interventions that resemble caloric restriction, such as low glucose.
  • the discovery that both yeast and flies lacking the SIR2 gene do not live longer when calorically restricted provides evidence that SIR2 genes mediate the beneficial health effects of a restricted calorie diet.
  • mutations that reduce the activity of the yeast glucose-responsive cAMP (adenosine 3',5'-monophosphate)- dependent (PKA) pathway extend life span in wild type cells but not in mutant sir2 strains, demonstrating that SIR2 is likely to be a key downstream component of the caloric restriction pathway.
  • novel sirtuin-modulating compounds and methods of use thereof are novel sirtuin-modulating compounds and methods of use thereof.
  • the invention provides sirtuin-modulating compounds of
  • the invention provides methods for using sirtuin- modulating compounds, or compositions comprising sirtuin-modulating compounds.
  • sirtuin-modulating compounds that increase the level and/or activity of a sirtuin protein may be used for a variety of therapeutic applications including, for example, increasing the lifespan of a cell, and treating and/or preventing a wide variety of diseases and disorders including, for example, diseases or disorders related to aging or stress, diabetes, obesity, neurodegenerative diseases, chemotherapeutic induced neuropathy, neuropathy associated with an ischemic event, ocular diseases and/or disorders, cardiovascular disease, blood clotting disorders, inflammation, and/or flushing, etc.
  • Sirtuin-modulating compounds that increase the level and/or activity of a sirtuin protein may also be used for treating a disease or disorder in a subject that would benefit from increased mitochondrial activity, for enhancing muscle performance, for increasing muscle ATP levels, or for treating or preventing muscle tissue damage associated with hypoxia or ischemia.
  • sirtuin-modulating compounds that decrease the level and/or activity of a sirtuin protein may be used for a variety of therapeutic applications including, for example, increasing cellular sensitivity to stress, increasing apoptosis, treatment of cancer, stimulation of appetite, and/or stimulation of weight gain, etc.
  • the methods comprise administering to a subject in need thereof a pharmaceutically effective amount of a sirtuin-modulating compound.
  • the sirtuin-modulating compounds may be administered alone or in combination with other compounds, including other sirtuin-modulating compounds, or other therapeutic agents.
  • DETAILED DESCRIPTION 1. Definitions
  • agent is used herein to denote a chemical compound, a mixture of chemical compounds, a biological macromolecule (such as a nucleic acid, an antibody, a protein or portion thereof, e.g., a peptide), or an extract made from biological materials such as bacteria, plants, fungi, or animal (particularly mammalian) cells or tissues.
  • a biological macromolecule such as a nucleic acid, an antibody, a protein or portion thereof, e.g., a peptide
  • an extract made from biological materials such as bacteria, plants, fungi, or animal (particularly mammalian) cells or tissues.
  • the activity of such agents may render it suitable as a "therapeutic agent” which is a biologically, physiologically, or pharmacologically active substance (or substances) that acts locally or systemically in a subject.
  • bioavailable when referring to a compound is art-recognized and refers to a form of a compound that allows for it, or a portion of the amount of compound administered, to be absorbed by, incorporated into, or otherwise physiologically available to a subject or patient to whom it is administered.
  • Biologically active portion of a sirtuin refers to a portion of a sirtuin protein having a biological activity, such as the ability to deacetylate.
  • Biologically active portions of a sirtuin may comprise the core domain of sirtuins.
  • Biologically active portions of SIRTl having GenBank Accession No. NP_036370 that encompass the NAD+ binding domain and the substrate binding domain may include without limitation, amino acids 62-293 of GenBank Accession No. NP_036370, which are encoded by nucleotides 237 to 932 of GenBank Accession No. NM_012238. Therefore, this region is sometimes referred to as the core domain.
  • SIRTl also sometimes referred to as core domains
  • core domains include about amino acids 261 to 447 of GenBank Accession No. NP_036370, which are encoded by nucleotides 834 to 1394 of GenBank Accession No. NM_012238; about amino acids 242 to 493 of GenBank Accession No. NP_036370, which are encoded by nucleotides 777 to 1532 of GenBank Accession No. NM_012238; or about amino acids 254 to 495 of GenBank Accession No. NP_036370, which are encoded by nucleotides 813 to 1538 of GenBank Accession No. NM_012238.
  • cat(s) refers to a feline animal including domestic cats and other members of the family Felidae, genus Felis.
  • Diabetes refers to high blood sugar or ketoacidosis, as well as chronic, general metabolic abnormalities arising from a prolonged high blood sugar status or a decrease in glucose tolerance. “Diabetes” encompasses both the type I and type II (Non Insulin Dependent Diabetes Mellitus or NIDDM) forms of the disease.
  • the risk factors for diabetes include the following factors: waistline of more than 40 inches for men or 35 inches for women, blood pressure of 130/85 mmHg or higher, triglycerides above 150 mg/dl, fasting blood glucose greater than 100 mg/dl or high- density lipoprotein of less than 40 mg/dl in men or 50 mg/dl in women.
  • ED 50 refers to the art-recognized measure of effective dose In certain embodiments, ED 50 means the dose of a drug which produces 50% of its maximum response or effect, or alternatively, the dose which produces a predetermined response in 50% of test subjects or preparations.
  • LD 50 refers to the art-recognized measure of lethal dose. In certain embodiments, LD 50 means the dose of a drug which is lethal in 50% of test subjects.
  • therapeutic index is an art-recognized term which refers to the therapeutic index of a drug, defined as LD50/ED50.
  • hyperinsulinemia refers to a state in an individual in which the level of insulin in the blood is higher than normal.
  • insulin resistance refers to a state in which a normal amount of insulin produces a subnormal biologic response relative to the biological response in a subject that does not have insulin resistance.
  • insulin resistance disorder refers to any disease or condition that is caused by or contributed to by insulin resistance. Examples include: diabetes, obesity, metabolic syndrome, insulin-resistance syndromes, syndrome X, insulin resistance, high blood pressure, hypertension, high blood cholesterol, dyslipidemia, hyperlipidemia, atherosclerotic disease including stroke, coronary artery disease or myocardial infarction, hyperglycemia, hyperinsulinemia and/or hyperproinsulinemia, impaired glucose tolerance, delayed insulin release, diabetic complications, including coronary heart disease, angina pectoris, congestive heart failure, stroke, cognitive functions in dementia, retinopathy, peripheral neuropathy, nephropathy, glomerulonephritis, glomerulosclerosis, nephrotic syndrome, hypertensive nephrosclerosis some types of cancer (such as endometrial, breast, prostate, and colon), complications of pregnancy, poor female reproductive health (such as menstrual irregularities, infertility, irregular ovulation, polycystic
  • livestock animals refers to domesticated quadrupeds, which includes those being raised for meat and various byproducts, e.g., a bovine animal including cattle and other members of the genus Bos, a porcine animal including domestic swine and other members of the genus Sus, an ovine animal including sheep and other members of the genus Ovis, domestic goats and other members of the genus Capra; domesticated quadrupeds being raised for specialized tasks such as use as a beast of burden, e.g., an equine animal including domestic horses and other members of the family Equidae, genus Equus.
  • mammals include humans, primates, livestock animals (including bovines, porcines, etc.), companion animals (e.g., canines, felines, etc.) and rodents (e.g., mice and rats).
  • livestock animals including bovines, porcines, etc.
  • companion animals e.g., canines, felines, etc.
  • rodents e.g., mice and rats.
  • Obese individuals or individuals suffering from obesity are generally individuals having a body mass index (BMI) of at least 25 or greater. Obesity may or may not be associated with insulin resistance.
  • BMI body mass index
  • parenteral administration and “administered parenterally” are art-recognized and refer to modes of administration other than enteral and topical administration, usually by injection, and includes, without limitation, intravenous, intramuscular, intraarterial, intrathecal, intracapsular, intraorbital, intracardiac, intradermal, intraperitoneal, transtracheal, subcutaneous, subcuticular, intraarticular, subcapsular, subarachnoid, intraspinal, and intrasternal injection and infusion.
  • a “patient”, “subject”, “individual” or “host” refers to either a human or a non-human animal.
  • pharmaceutically acceptable carrier refers to a pharmaceutically-acceptable material, composition or vehicle, such as a liquid or solid filler, diluent, excipient, solvent or encapsulating material, involved in carrying or transporting any subject composition or component thereof.
  • a pharmaceutically-acceptable material such as a liquid or solid filler, diluent, excipient, solvent or encapsulating material, involved in carrying or transporting any subject composition or component thereof.
  • Each carrier must be “acceptable” in the sense of being compatible with the subject composition and its components and not injurious to the patient.
  • materials which may serve as pharmaceutically acceptable carriers include: (1) sugars, such as lactose, glucose and sucrose; (2) starches, such as corn starch and potato starch; (3) cellulose, and its derivatives, such as sodium carboxymethyl cellulose, ethyl cellulose and cellulose acetate; (4) powdered tragacanth; (5) malt; (6) gelatin; (7) talc; (8) excipients, such as cocoa butter and suppository waxes; (9) oils, such as peanut oil, cottonseed oil, safflower oil, sesame oil, olive oil, corn oil and soybean oil; (10) glycols, such as propylene glycol; (11) polyols, such as glycerin, sorbitol, mannitol and polyethylene glycol; (12) esters, such as ethyl oleate and ethyl laurate; (13) agar; (14) buffering agents, such as magnesium hydroxide and aluminum hydroxide;
  • preventing is art-recognized, and when used in relation to a condition, such as a local recurrence (e.g., pain), a disease such as cancer, a syndrome complex such as heart failure or any other medical condition, is well understood in the art, and includes administration of a composition which reduces the frequency of, or delays the onset of, symptoms of a medical condition in a subject relative to a subject which does not receive the composition.
  • a condition such as a local recurrence (e.g., pain)
  • a disease such as cancer
  • a syndrome complex such as heart failure or any other medical condition
  • prevention of cancer includes, for example, reducing the number of detectable cancerous growths in a population of patients receiving a prophylactic treatment relative to an untreated control population, and/or delaying the appearance of detectable cancerous growths in a treated population versus an untreated control population, e.g., by a statistically and/or clinically significant amount.
  • Prevention of an infection includes, for example, reducing the number of diagnoses of the infection in a treated population versus an untreated control population, and/or delaying the onset of symptoms of the infection in a treated population versus an untreated control population.
  • Prevention of pain includes, for example, reducing the magnitude of, or alternatively delaying, pain sensations experienced by subjects in a treated population versus an untreated control population.
  • prophylactic or therapeutic treatment refers to administration of a drug to a host. If it is administered prior to clinical manifestation of the unwanted condition (e.g., disease or other unwanted state of the host animal) then the treatment is prophylactic, i.e., it protects the host against developing the unwanted condition, whereas if administered after manifestation of the unwanted condition, the treatment is therapeutic (i.e., it is intended to diminish, ameliorate or maintain the existing unwanted condition or side effects therefrom).
  • pyrogen-free refers to a composition that does not contain a pyrogen in an amount that would lead to an adverse effect (e.g., irritation, fever, inflammation, diarrhea, respiratory distress, endotoxic shock, etc.) in a subject to which the composition has been administered.
  • an adverse effect e.g., irritation, fever, inflammation, diarrhea, respiratory distress, endotoxic shock, etc.
  • the term is meant to encompass compositions that are free of, or substantially free of, an endotoxin such as, for example, a lipopoly saccharide (LPS).
  • LPS lipopoly saccharide
  • Replicative lifespan of a cell refers to the number of daughter cells produced by an individual "mother cell.”
  • Increasing the lifespan of a cell or “extending the lifespan of a cell,” as applied to cells or organisms, refers to increasing the number of daughter cells produced by one cell; increasing the ability of cells or organisms to cope with stresses and combat damage, e.g., to DNA, proteins; and/or increasing the ability of cells or organisms to survive and exist in a living state for longer under a particular condition, e.g., stress (for example, heatshock, osmotic stress, high energy radiation, chemically-induced stress, DNA damage, inadequate salt level, inadequate nitrogen level, or inadequate nutrient level). Lifespan can be increased by at least about 10%, 20%, 30%, 40%, 50%, 60% or between 20% and 70%, 30% and 60%, 40% and 60% or more using methods described herein.
  • sirtuin-activating compound refers to a compound that increases the level of a sirtuin protein and/or increases at least one activity of a sirtuin protein.
  • a sirtuin-activating compound may increase at least one biological activity of a sirtuin protein by at least about 10%, 25%, 50%, 75%, 100%, or more.
  • Exemplary biological activities of sirtuin proteins include deacetylation, e.g., of histones and p53; extending lifespan; increasing genomic stability; silencing transcription; and controlling the segregation of oxidized proteins between mother and daughter cells.
  • sirtuin protein refers to a member of the sirtuin deacetylase protein family, or preferably to the sir2 family, which include yeast Sir2 (GenBank Accession No. P53685), C. elegans Sir-2.1 (GenBank Accession No. NP_501912), and human SIRTl (GenBank Accession No. NM_012238 and NP_036370 (or AF083106)) and SIRT2 (GenBank Accession No. NM_012237, NM_030593, NP_036369, NP_085096, and AF083107) proteins.
  • HST genes additional yeast Sir2-like genes termed "HST genes” (homologues of Sir two) HSTl , HST2, HST3 and HST4, and the five other human homologues hSIRT3, hSIRT4, hSIRT5, hSIRT6 and hSIRT7 (Brachmann et al. (1995) Genes Dev. 9:2888 and Frye et al. (1999) BBRC 260:273).
  • HST genes homologues of Sir two HSTl , HST2, HST3 and HST4
  • hSIRT3, hSIRT4, hSIRT5, hSIRT6 and hSIRT7 Bos et al. (1995) Genes Dev. 9:2888 and Frye et al. (1999) BBRC 260:273
  • Preferred sirtuins are those that share more similarities with SIRTl, i.e., hSIRTl, and/or Sir2 than with SIRT2, such as those members having at least part of the N-terminal sequence present in SIRTl and absent in SIRT2 such as SIRT3 has.
  • SIRTl protein refers to a member of the sir2 family of sirtuin deacetylases.
  • a SIRTl protein includes yeast Sir2 (GenBank Accession No. P53685), C. elegans Sir-2.1 (GenBank Accession No. NP_501912), human SIRTl (GenBank Accession No. NM_012238 or NP_036370 (or AF083106)), and equivalents and fragments thereof.
  • a SIRTl protein includes a polypeptide comprising a sequence consisting of, or consisting essentially of, the amino acid sequence set forth in GenBank Accession Nos.
  • SIRTl proteins include polypeptides comprising all or a portion of the amino acid sequence set forth in GenBank Accession Nos. NP_036370, NP_501912, NP_085096, NP_036369, or P53685; the amino acid sequence set forth in GenBank Accession Nos.
  • Polypeptides of the invention also include homologs (e.g., orthologs and paralogs), variants, or fragments, of GenBank Accession Nos. NP_036370, NP_501912, NP_085096, NP_036369, or P53685.
  • SIRT2 protein As used herein "SIRT2 protein”, “SIRT3 protein”, “SIRT4 protein”, SIRT 5 protein”, “SIRT6 protein”, and “SIRT7 protein” refer to other mammalian, e.g. human, sirtuin deacetylase proteins that are homologous to SIRTl protein, particularly in the approximately 275 amino acid conserved catalytic domain.
  • SIRT3 protein refers to a member of the sirtuin deacetylase protein family that is homologous to SIRTl protein.
  • a SIRT3 protein includes human SIRT3 (GenBank Accession No.
  • a SIRT3 protein includes a polypeptide comprising a sequence consisting of, or consisting essentially of, the amino acid sequence set forth in GenBank Accession Nos. AAH01042, NP_036371, NP_001017524, or NP_071878.
  • SIRT3 proteins include polypeptides comprising all or a portion of the amino acid sequence set forth in GenBank Accession Nos.
  • Polypeptides of the invention also include homologs (e.g., orthologs and paralogs), variants, or fragments, of GenBank Accession Nos. AAH01042, NP_036371, NP_001017524, or NP_071878.
  • a SIRT3 protein includes a fragment of SIRT3 protein that is produced by cleavage with a mitochondrial matrix processing peptidase (MPP) and/or a mitochondrial intermediate peptidase (MIP).
  • MPP mitochondrial matrix processing peptidase
  • MIP mitochondrial intermediate peptidase
  • systemic administration refers to the administration of a subject composition, therapeutic or other material other than directly into the central nervous system, such that it enters the patient's system and, thus, is subject to metabolism and other like processes.
  • tautomer as used herein is art-regcognized and refers to the formal migration of a hydrogen atom, i.e., proton, accompanied by a swap of a single bond and adjacent double bond.
  • tautomer When used herein to describe a compound or genus of compounds, tautomer includes any portion of a compound or the entire compound such as a single substituent of a compound, multiple substiutents of a compound or, for example, the entire compound.
  • the tautomer of a compound that includes a hydroxyl- substituted pyridine ring (A) may also include within the scope of the invention the tautomerized form of the substituted ring (B):
  • a B The term "therapeutic agent” is art-recognized and refers to any chemical moiety that is a biologically, physiologically, or pharmacologically active substance that acts locally or systemically in a subject.
  • the term also means any substance intended for use in the diagnosis, cure, mitigation, treatment or prevention of disease or in the enhancement of desirable physical or mental development and/or conditions in an animal or human.
  • therapeutic effect is art-recognized and refers to a local or systemic effect in animals, particularly mammals, and more particularly humans caused by a pharmacologically active substance.
  • therapeutically- effective amount means that amount of such a substance that produces some desired local or systemic effect at a reasonable benefit/risk ratio applicable to any treatment.
  • the therapeutically effective amount of such substance will vary depending upon the subject and disease condition being treated, the weight and age of the subject, the severity of the disease condition, the manner of administration and the like, which can readily be determined by one of ordinary skill in the art.
  • certain compositions described herein may be administered in a sufficient amount to produce a desired effect at a reasonable benefit/risk ratio applicable to such treatment.
  • Treating" a condition or disease refers to curing as well as ameliorating at least one symptom of the condition or disease.
  • the term “vision impairment” refers to diminished vision, which is often only partially reversible or irreversible upon treatment (e.g., surgery). Particularly severe vision impairment is termed “blindness” or “vision loss”, which refers to a complete loss of vision, vision worse than 20/200 that cannot be improved with corrective lenses, or a visual field of less than 20 degrees diameter (10 degrees radius).
  • the invention provides novel sirtuin-modulating compounds for treating and/or preventing a wide variety of diseases and disorders including, for example, diseases or disorders related to aging or stress, diabetes, obesity, neurodegenerative diseases, ocular diseases and disorders, cardiovascular disease, blood clotting disorders, inflammation, cancer, and/or flushing, etc.
  • Sirtuin- modulating compounds that increase the level and/or activity of a sirtuin protein may also be used for treating a disease or disorder in a subject that would benefit from increased mitochondrial activity, for enhancing muscle performance, for increasing muscle ATP levels, or for treating or preventing muscle tissue damage associated with hypoxia or ischemia.
  • Other compounds disclosed herein may be suitable for use in a pharmaceutical composition and/or one or more methods disclosed herein.
  • sirtuin-modulating compound of the invention are represented by structural formula (I):
  • each of Z 1 , Z 2 , Z 3 , Z 4 , and Z 5 are independently selected from N and CR, wherein: each of Z 3 , Z 4 and Z 5 is independently CR, or at least one of Z 1 or Z 2 is N and no more than two of Z 1 -Z 5 are simultaneously N, or two of Z 3 , Z 4 and Z 5 are N and each other of Z 1 -Z 5 is independently CR; each R is independently selected from hydrogen, halo, -OH, -C ⁇ N, fluoro-substituted C 1 -C 2 alkyl, -0-(Ci-C 2 ) fluoro-substituted alkyl, -S-(Ci-C 2 ) fluoro-substituted alkyl, Ci-C 4 alkyl, -0-(Ci-C 4 ) alkyl, -S-(Ci-C 4 ) alkyl, C 3 -C 7 cycloalky
  • any heterocycle or saturated heterocycle substituent of R 1 is optionally and independently substituted at any substitutable nitrogen atom with Ci-C 4 alkyl, fluoro- or chloro-substituted Ci-C 4 alkyl or -(Ci-C 4 alkyl)- 0-(Ci-C 4 alkyl);
  • each R 3 is independently selected from hydrogen and -Ci-C 4 alkyl, wherein the alkyl is optionally substituted with one or more of -OH, fluoro, -NH 2 ,
  • X is -C(O)-NR 6 -f . In certain embodiments, where each of Z 3 , Z 4 and Z 5 is CR, X is -C(O)-NR 6 - f. In particular embodiments, X is -C(O)-NH-I.
  • each R is independently selected from hydrogen, halo, -OH, -C ⁇ N, fluoro-substituted C 1 -C 2 alkyl, -O-(C r C 2 ) fluoro-substituted alkyl, -S-(Ci-C 2 ) fluoro-substituted alkyl, Ci-C 4 alkyl, -0-(Ci-C 4 ) alkyl, -S-(Ci-C 4 ) alkyl and C 3 -C 7 cycloalkyl;
  • R is selected from a carbocycle and a heterocycle other than piperazine, wherein R 2 is optionally substituted with one to two substitutents independently selected from halo, -C ⁇ N, C 1 -C 4 alkyl, C 3 -C 7 cycloalkyl, C 1 -C 2 fluoro-substituted alkyl, -O-R 3 , -S-R 3 , -(C 1 -C 4 alkyl)-N(R 3 )(R 3 ), -N(R 3 )(R 3 ), -0-(Ci-C 4 alkyl)-N(R 3 )(R 3 ), -(Ci-C 4 alkyl)-O-(Ci-C 4 alkyl)-N(R 3 )(R 3 ), -C(O)-N(R 3 XR 3 ), -(Ci-C 4 alkyl)-C(O)-N(R 3 )(R 3 ),
  • each of Z 3 , Z 4 and Z 5 is CR in the compound represented by Structural Formula (I). In certain such embodiments, each of Z x -Z 5 is CR.
  • R 1 is optionally substituted aryl, heteroaryl or saturated heterocyclyl.
  • R 1 is selected from optionally substituted thiazole, pyridine, pyrazine, and phenyl.
  • Suitable values of R 2 include phenyl, pyridyl and benzomorpholine, such as when R 1 has the values indicated previously.
  • each of Z x -Z 5 is CR
  • R 1 is selected from thiazole, pyridine, pyrazine and phenyl
  • R 2 is selected from phenyl, pyridyl, and benzomorpholine
  • X is -C(O)-NH-I
  • sirtuin-modulating compounds of the invention are represented by structural formula (II):
  • each of Z 3 , Z 4 , and Z 5 are independently selected from N and CR, wherein only one of Z 3 , Z 4 , and Z 5 is N, wherein: each R is independently selected from hydrogen, halo, -OH, -C ⁇ N, fluoro-substituted C 1 -C 2 alkyl, -O-(C r C 2 ) fluoro-substituted alkyl, -S-(C 1 -C 2 ) fluoro-substituted alkyl, C 1 -C 4 alkyl, -0-(C 1 -C 4 ) alkyl, -S-(C 1 -C 4 ) alkyl, C 3 -C 7 cycloalkyl, -(Ci-C 2 ) alkyl-N(R 3 )(R 3 ), hydroxy- substituted Ci-C 4 alkoxy, -(C 1 -GO-O- saturated heterocycle,
  • R 1 is selected from a carbocycle and a heterocycle, wherein R 1 is optionally substituted with one or more substitutents independently selected from halo, -C ⁇ N, C 1 -C 4 alkyl, hydroxy-substituted C 1 -C 4 alkoxy, C 3 -C 7 cycloalkyl, fluoro-substituted Ci-C 2 alkyl, -O-R 3 , -S-R 3 , -(C 1 -C 4 alkyl) -N(R 3 ) (R 3 ), -N(R 3 )(R 3 ), -0-(C 1 -C 4 alkyl)-N(R 3 )(R 3 ), -(Ci-C 4 alkyl)-O-(Ci-C 4 alkyl)-N(R 3 )(R 3 ), -C(O)-N(R 3 )(R 3 ), and -(Ci-C 4 alkyl)-
  • each R is independently selected from hydrogen, halo, -OH, -C ⁇ N, fluoro-substituted C 1 -C 2 alkyl, -O-(C r C 2 ) fluoro-substituted alkyl, -S-(Ci-C 2 ) fluoro-substituted alkyl, Ci-C 4 alkyl, -0-(Ci-C 4 ) alkyl, -S-(Ci-C 4 ) alkyl and C 3 -C 7 cycloalkyl; R 1 is selected from a carbocycle and a heterocycle, wherein R 1 is optionally substituted with one to two substitutents independently selected from halo, -C ⁇ N, Ci-C 4
  • R is selected from a carbocycle and a heterocycle other than piperazine, wherein R 2 is optionally substituted with one to two substitutents independently selected from halo, -C ⁇ N, C 1 -C 4 alkyl, C 3 -C 7 cycloalkyl, C 1 -C 2 fluoro-substituted alkyl, -O-R 3 , -S-R 3 , -(C 1 -C 4 alkyl)-N(R 3 )(R 3 ), -N(R 3 )(R 3 ), -0-(Ci-C 4 alkyl)-N(R 3 )(R 3 ), -(Ci-C 4 alkyl)-O-(Ci-C 4 alkyl)-N(R 3 )(R 3 ), -C(O)-N(R 3 XR 3 ), -(Ci-C 4 alkyl)-C(O)-N(R 3 )(R 3 ),
  • R 1 is optionally substituted aryl, heteroaryl or saturated heterocyclyl.
  • X is -C(O)-NH-f .
  • sirtuin-modulating compounds of the invention are represented by Structural Formula (III):
  • each R is independently selected from hydrogen, halo, -OH, -C ⁇ N, fluoro-substituted Ci-C 2 alkyl, -0-(Ci-C 2 ) fluoro-substituted alkyl, -S-(Ci-C 2 ) fluoro-substituted alkyl, Ci-C 4 alkyl, -0-(Ci-C 4 ) alkyl, -S-(Ci-C 4 ) alkyl C 3 -C 7 cycloalkyl, -(Ci-C 2 ) alkyl-N(R 3 )(R 3 ), hydroxy- substituted Ci-C 4 alkoxy, -(C 1 -C-O-O- saturated heterocycle, -0-(C 1 -C 3 ) alkyl-N(R 3 )(R 3 ), and -N(R 3 )(R 3 );
  • R 1 is selected from a carbocycle and a heterocycle, wherein R 1 is optionally substituted with one or more substitutents independently selected from halo, -C ⁇ N, Ci-C 4 alkyl, hydroxy-substituted Ci-C 4 alkoxy, C 3 -C 7 cycloalkyl, fluoro-substituted Ci-C 2 alkyl, -O-R 3 , -S-R 3 , -(C 1 -C 4 alkyl) -N(R 3 ) (R 3 ), -N(R 3 )(R 3 ), -0-(C 1 -C 4 alkyl)-N(R 3 )(R 3 ), -(C 1 -C 4 alkyl)-O-(C r C 4 alkyl)-N(R 3 )(R 3 ), -C(O)-N(R 3 )(R 3 ), and -(Ci-C 4 alkyl)-C(O
  • any heterocycle or saturated heterocycle substituent of R 1 is optionally and independently substituted at any substitutable nitrogen atom with Ci-C 4 alkyl fluoro- or chloro-substituted Ci-C 4 alkyl or -(Ci-C 4 alkyl)- 0-(Ci-C 4 alkyl);
  • each R is independently selected from hydrogen, halo, -OH, -C ⁇ N, fluoro-substituted Ci-C 2 alkyl, -0-(Ci-C 2 ) fluoro-substituted alkyl, -S-(Ci-C 2 ) fluoro-substituted alkyl, C 1 -C 4 alkyl, -0-(C r C 4 ) alkyl, -S-(C 1 -C 4 ) alkyl and C 3 -C 7 cycloalkyl;
  • R 1 is selected from a carbocycle and a heterocycle, wherein R 1 is optionally substituted with one to two substitutents independently selected from halo, -C ⁇ N, C 1 -C 4 alkyl, hydroxy-substituted C 1 -C 4 alkoxy, C 3 -C 7 cycloalkyl, fluoro-substituted Ci-C 2 alkyl, -O-R 3 , -S-R 3 , -(C 1 -C 4 alkyl) -N(R 3 ) (R 3 ), -N(R 3 )(R 3 ), -0-(C 1 -C 4 alkyl)-N(R 3 )(R 3 ), -(Ci-C 4 alkyl)-O-(Ci-C 4 alkyl)-N(R 3 )(R 3 ), -C(O)-N(R 3 )(R 3 ), and -(Ci-C 4 alkyl)-
  • R 2 is selected from a carbocycle and a heterocycle other than piperazine, wherein R 2 is optionally substituted with one to two substitutents independently selected from halo, -C ⁇ N, C 1 -C 4 alkyl, C 3 -C 7 cycloalkyl, C 1 -C 2 fluoro-substituted alkyl, -O-R 3 , -S-R 3 , -(C 1 -C 4 alkyl)-N(R 3 )(R 3 ), -N(R 3 )(R 3 ), -0-(Ci-C 4 alkyl)-N(R 3 )(R 3 ), -(Ci-C 4 alkyl)-O-(Ci-C 4 alkyl)-N(R 3 )(R 3 ), -C(O)-N(R 3 )(R 3 ), -C(O)-N(R 3 )(R 3 ), -(O)-N(R
  • R 1 is optionally substituted aryl, heteroaryl or saturated heterocyclyl.
  • X is -C(O)-NH-f .
  • compounds of the invention are a subset of the compounds of Structural Formula (III) represented by Structural Formula (IV):
  • each R is independently selected from hydrogen, halo, -OH, -C ⁇ N, fluoro-substituted Ci-C 2 alkyl, -0-(Ci-C 2 ) fluoro-substituted alkyl, -S-(Ci-C 2 ) fluoro-substituted alkyl, C 1 -C 4 alkyl, -0-(C r C 4 ) alkyl, -S-(C 1 -C 4 ) alkyl, C 3 -C 7 cycloalkyl, -(C 1 -C 2 ) alkyl-N(R 3 )(R 3 ), hydroxy- substituted C 1 -C 4 alkoxy, -(Ci-C 4 )-0- saturated heterocycle, -0-(Ci-C 3 ) alkyl-N(R 3 )(R 3 ), and -N(R 3 )(R 3 );
  • R 1 is optionally and independently substituted at any substitutable nitrogen atom with Ci-C 4 alkyl fluoro- or chloro-substituted Ci-C 4 alkyl and -(Ci-C 4 alkyl)-O-(C r C 4 alkyl);
  • R 2 is selected from a carbocycle and a heterocycle, wherein R 2 is optionally substituted with one or more substitutents independently selected from halo, -C ⁇ N, C 1 -C 4 alkyl, C 3 -C 7 cycloalkyl, fluoro- substituted C 1 -C 2 alkyl, hydroxy- substituted Ci-C 4 alkoxy, -O-R 3 , -S-R 3 , -SO
  • R 2 when R 2 is phenyl, R 2 is also optionally substituted with
  • any phenyl, saturated heterocycle, second heterocycle or cycloalkyl substituent of R 2 is optionally substituted at any substitutable carbon atom one or more substituents independently selected from halo, -C ⁇ N, Ci-C 4 alkyl, fluoro- or chloro-substituted C 1 -C 2 alkyl, -O-(fluoro-substituted Ci-C 2 alkyl), -0-(Ci-C 4 ) alkyl, -S-(Ci-C 4 ) alkyl, -S-(fluoro-substituted Ci-C 2 alkyl), -NH-(Ci-C 4 ) alkyl, and -N-
  • each R is independently selected from hydrogen, halo, -OH, -C ⁇ N, fluoro-substituted C 1 -C 2 alkyl, -0-(C 1 -C 2 ) fluoro-substituted alkyl, -S-(Ci-C 2 ) fluoro-substituted alkyl, C 1 -C 4 alkyl, -0-(C 1 -C 4 ) alkyl, -S-(C 1 -C 4 ) alkyl and C 3 -C 7 cycloalkyl;
  • R 1 is phenyl, R 1 is also optionally substituted with O-(saturated heterocycle), fluoro-substituted -O-(saturated heterocycle),
  • R 2 is selected from a carbocycle and a heterocycle, wherein R 2 is optionally substituted with one to two substitutents independently selected from halo, -C ⁇ N, Ci-C 4 alkyl, C 3 -C 7 cycloalkyl, Ci-C 2 fluoro-substituted alkyl, -O-R 3 , -S-R 3 , -(Ci-C 4 alkyl)-N(R 3 )(R 3 ), -N(R 3 )(R 3 ), -0-(C 1 -C 4 alkyl)-N(R 3 )(R 3 ), -(C 1 -C 4 alkyl)-O-(Ci-C 4 alkyl)-N(R 3 )(R 3 ), -C(O)-N(R 3 )(R 3 ), -C(O)-N(R 3 )(R 3 ), -C(O)-N(R 3 )(R 3
  • R 1 is optionally substituted aryl, heteroaryl or saturated heterocyclyl.
  • X is -C(O)-NH-f .
  • sirtuin-modulating compounds of the invention are represented by structural formula (V):
  • each R is independently selected from hydrogen, halo, -OH, -C ⁇ N, fluoro-substituted C 1 -C 2 alkyl, -0-(Ci-C 2 ) fluoro-substituted alkyl, -S-(Ci-C 2 ) fluoro-substituted alkyl, C 1 -C 4 alkyl, -0-(C 1 -C 4 ) alkyl, -S-(C 1 -C 4 ) alkyl and C 3 -C 7 cycloalkyl, -(C 1 -C 2 ) alkyl-N(R 3 )(R 3 ), hydroxy- substituted C 1 -C 4 alkoxy, -(C 1 -GO-O- saturated heterocycle, -0-(Ci-C 3 ) alkyl-N(R 3 )(R 3 ),
  • Ci-C 2 alkyl -0-(C r C 4 ) alkyl, -S-(C 1 -C 4 ) alkyl, -S-(fluoro-substituted Ci-C 2 alkyl), -NH-(C 1 -C 4 ) alkyl, and -N-(C r C 4 ) 2 alkyl; and any second heterocycle or saturated heterocycle substituent of R 2 is optionally and independently substituted at any substitutable nitrogen atom with Ci-C 4 alkyl, fluoro- or chloro-substituted C 1 -C 4 alkyl Or-(C 1 -C 4 alkyl)-
  • each R is independently selected from hydrogen, halo, -OH, -C ⁇ N, fluoro-substituted C 1 -C 2 alkyl, -0-(C 1 -C 2 ) fluoro-substituted alkyl, -S-(Ci-C 2 ) fluoro-substituted alkyl, C 1 -C 4 alkyl, -0-(C 1 -C 4 ) alkyl, -S-(C 1 -C 4 ) alkyl and C 3 -C 7 cycloalkyl;
  • R 2 is selected from a carbocycle and a heterocycle, wherein R 2 is optionally substituted with one to two substitutents independently selected from halo, -C ⁇ N, Ci-C 4 alkyl, C 3 -C 7 cycloalkyl, C 1 -C 2 fluoro-substituted alkyl, -O-R 3 , -S-R 3 , -(C 1 -C 4 alkyl)-N(R 3 )(R 3 ), -N(R 3 )(R 3 ), -0-(C 1 -C 4 alkyl)-N(R 3 )(R 3 ), -(C 1 -C 4 alkyl)-O-(C r C 4 alkyl)-N(R 3 )(R 3 ), -C(O)-N(R 3 )(R 3 ), -(O)-N(R 3 )(R 3 ), -(O)-N(R 3 )(R 3
  • compounds of the invention are a subset of the compounds of Structural Formula (V) represented by Structural Formula (VI):
  • each R is independently selected from hydrogen, halo, -OH, -C ⁇ N, fluoro-substituted Ci-C 2 alkyl, -0-(Ci-C 2 ) fluoro-substituted alkyl, -S-(Ci-C 2 ) fluoro-substituted alkyl, Ci-C 4 alkyl, -0-(Ci-C 4 ) alkyl, -S-(Ci-C 4 ) alkyl and C 3 -C 7 cycloalkyl, -(C 1 -C 2 ) alkyl-N(R 3 )(R 3 ), hydroxy- substituted C 1 -C 4 alkoxy, -(C 1 -C-O-O- saturated heterocycle, -0-(C 1 -C 3 ) alkyl-N(R 3 )(R 3 ), and
  • any second heterocycle or saturated heterocycle substituent of R 2 is optionally and independently substituted at any substitutable nitrogen atom with Ci-C 4 alkyl, fluoro- or chloro-substituted C 1 -C 4 alkyl Or-(Ci-C 4 alkyl)- 0-(Ci-C 4 alkyl); each R 3 is independently selected from hydrogen and -Ci-C 4 alkyl, wherein the alkyl is optionally substituted with one or more of -OH, fluoro, -NH 2 , -NH(Ci-C 4
  • R 1 is optionally substituted aryl, heteroaryl or saturated heterocyclyl.
  • R 1 for any of structural formulas (I) to (VI) is selected from:
  • R 1 is selected from
  • R 1 is selected from: and .
  • R 1 is selected from:
  • R of any of structural formulas (I) to (VI) is selected from optionally substituted aryl and optionally substituted heteroaryl.
  • R is selected from: 5 and wherein R is optionally substituted with one or more groups independently selected from halo, C 1 -C 4 alkyl, -(C 1 -C 4 alkyl)-N(R 3 )(R 3 ), Q-C 2 fluoro-substituted alkyl, -O-R 3 , -SO 2 -R 3 , -N(R 3 )(R 3 ), and -0-(Ci-C 4 alkyl)-N(R 3 )(R 3 ).
  • R 2 is meta-substituted relative to the attachment of R 2 to the rest of the compound, and wherein R 2 is optionally
  • R 2 is selected from: ⁇ — ' ,
  • R 2 is selected from optionally substituted aryl and optionally substituted heteroaryl. In certain such embodiments, R 2 is selected from:
  • R 2 is meta-substituted relative to the attachment of R 2 to the rest of the compound, and R 2 is optionally further substituted.
  • R is
  • the embodiments described below apply to compounds of any of Structural Formulas (I)-(VI).
  • the compound of any of structures (I)-(VI) is a free base.
  • Sirtuin-modulating compounds of the invention advantageously modulate the level and/or activity of a sirtuin protein, particularly the deacetylase activity of the sirtuin protein.
  • sirtuin-modulating compounds of the invention do not substantially have one or more of the following activities: inhibition of PI3-kinase, inhibition of aldoreductase, inhibition of tyrosine kinase, transactivation of EGFR tyrosine kinase, coronary dilation, or spasmolytic activity, at concentrations of the compound that are effective for modulating the deacetylation activity of a sirtuin protein (e.g., such as a SIRTl and/or a SIRT3 protein).
  • a sirtuin protein e.g., such as a SIRTl and/or a SIRT3 protein.
  • An alkyl group is a straight chained or branched non-aromatic hydrocarbon which is completely saturated.
  • a straight chained or branched alkyl group has from 1 to about 20 carbon atoms, preferably from 1 to about 10.
  • Examples of straight chained and branched alkyl groups include methyl, ethyl, n-propyl, iso- propyl, n-butyl, sec-butyl, tert-butyl, pentyl, hexyl, pentyl and octyl.
  • a C 1 -C 4 straight chained or branched alkyl group is also referred to as a "lower alkyl" group.
  • alkenyl and alkynyl refer to unsaturated aliphatic groups analogous in length and possible substitution to the alkyl groups described above, but that contain at least one double or triple bond respectively.
  • alkoxyl or alkoxy as used herein refers to an alkyl group having an oxygen radical attached thereto. Representative alkoxyl groups include methoxy, ethoxy, propyloxy, tert-butoxy and the like.
  • a cycloalkyl group is a cyclic hydrocarbon which is completely saturated. Typically, a cycloalkyl group has from 3 to about 10 carbon atoms, more typically 3 to 8 carbon atoms.
  • heterocycle refers to a saturated or unsaturated ring comprising one or more heteroatoms selected from, for example, N, O, and S atoms.
  • Heterocycles include 4-7 membered monocyclic and 8- 12 membered bicyclic rings. Each ring of a bicyclic heterocycle may be selected from saturated, unsaturated and aromatic rings.
  • an aromatic ring e.g., pyridyl
  • a saturated or unsaturated ring e.g., cyclohexane, cyclopentane, or cyclohexene.
  • heterocyclyl and heterocyclic also include polycyclic ring systems having two or more cyclic rings in which two or more carbons or heteroatoms are common to two adjoining rings wherein at least one of the rings is heterocyclic, e.g., the other cyclic rings can be cycloalkyls, cycloalkenyls, cycloalkynyls, aryls, heteroaryls, and/or heterocycloalkyls.
  • Heterocyclyl groups include, for example, piperidine, piperazine, pyrrolidine, morpholine, lactones, and lactams.
  • heteroaryl includes substituted or unsubstituted aromatic single ring structures, preferably 5- to 7-membered rings, more preferably 5- to 6- membered rings, whose ring structures include at least one heteroatom, preferably one to four heteroatoms, more preferably one or two heteroatoms.
  • heteroaryl and “hetaryl” also include polycyclic ring systems having two or more cyclic rings in which two or more carbons or heteroatoms are common to two adjoining rings wherein at least one of the rings is heteroaromatic, e.g., the other cyclic rings can be cycloalkyls, cycloalkenyls, cycloalkynyls, aryls, heteroaryls, and/or heterocyclyls.
  • Heteroaryl groups include, for example, pyrrole, furan, thiophene, imidazole, oxazole, thiazole, pyrazole, pyridine, pyrazine, pyridazine, and pyrimidine.
  • Monocyclic rings include 5-7 membered aryl or heteroaryl, 3-7 membered cycloalkyl, and 5-7 membered non-aromatic heterocyclyl. Monocyclic rings are optionally substituted with one or more substituents such as halo, cyano, lower alkoxy, lower alkyl, hydroxyl, amino, lower alkylamino and lower dialkylamino.
  • Exemplary monocyclic groups include substituted or unsubstituted heterocycles or carbocycles such as thiazolyl, oxazolyl, oxazinyl, thiazinyl, dithianyl, dioxanyl, isoxazolyl, isothiazolyl, triazolyl, furanyl, tetrahydrofuranyl, dihydrofuranyl, pyranyl, tetrazolyl, pyrazolyl, pyrazinyl, pyridazinyl, imidazolyl, pyridinyl, pyrrolyl, dihydropyrrolyl, pyrrolidinyl, piperidinyl, piperazinyl, pyrimidinyl, morpholinyl, tetrahydrothiophenyl, thiophenyl, cyclohexyl, cyclopentyl, cyclopropyl, cyclobutyl, cycloheptany
  • Aromatic (aryl) groups include carbocyclic aromatic groups such as phenyl, naphthyl, and anthracyl, and heteroaryl groups such as imidazolyl, thienyl, furyl, pyridyl, pyrimidyl, pyranyl, pyrazolyl, pyrrolyl, pyrazinyl, thiazolyl, oxazolyl, and tetrazolyl.
  • Aromatic groups also include fused polycyclic aromatic ring systems in which a carbocyclic aromatic ring or heteroaryl ring is fused to one or more other heteroaryl rings.
  • Azabicyclo refers to a bicyclic molecule that contains a nitrogen atom in the ring skeleton.
  • the two rings of the bicycle may be fused, at two mutually bonded atoms, e.g., indole, across a sequence of atoms, e.g., azabicyclo[2.2.1]heptane, or at a single atom, e.g., spirocycle.
  • Bridged azabicyclo refers to a bicyclic molecule that contains a nitrogen atom and two fused rings wherein the fusion occurs across a sequence of atoms, i.e., bridgehead atoms.
  • Bridged bicyclo compounds comprise at least one bridge of one or more atoms connecting two bridgehead atoms.
  • Suitable substituents on a heterocyclyl or heterocyclylmethyl group include - OH, halogen (-Br, -Cl, -I and -F), -OR a , -O-COR a , -COR a , -C(O)R a , -CN, -NO 2 , - COOH, -COOR a , -OCO 2 R a , -C(0)NR a R b , -0C(0)NR a R b , -SO 3 H, -NH 2 , -NHR a , - N(R a R b ), -COOR a , -CHO, -CONH 2 , -C0NHR a , -C0N(R a R b ), -NHC0R a , -NRC0R a , -NHCONH 2 , -NHCONR ⁇ , -NHC
  • R a -R d are each independently an optionally substituted group selected from an aliphatic, benzyl, or aromatic group, preferably an alkyl, benzylic or aryl group.
  • Optional substituents on R a -R d are selected from NH 2 , NH(Ci_ 4 aliphatic), N(Q.
  • Ci_ 4 aliphatic 2 , halogen, Ci_ 4 aliphatic, OH, O(Ci_ 4 aliphatic), NO 2 , CN, CO 2 H, CO 2 (Ci. 4 aliphatic), O(haloCi_ 4 aliphatic), or haloCi_ 4 aliphatic, wherein each of the foregoing Ci_ 4 aliphatic groups of is unsubstituted.
  • -NR a R b taken together, can also form a substituted or unsubstituted non-aromatic heterocyclic group.
  • a substituted aliphatic or substituted aryl group can have more than one substituent.
  • Halo-substituted includes from one halo substituent up to per-halo substitution.
  • Exemplary halo substituted Q-C 2 alkyl includes -CIH 2 , CF 2 H, -CCI 3 , -CH 2 CH 2 Br, -CH 2 CHCl 2 , -CHBrCH 2 Br, and -CF 2 CHCl 2 .
  • Per-halo-substituted Q-C 2 alkyl for example, includes -CCI 3 and -CCl 2 CF 3 .
  • Fluoro-substituted includes from one fluoro substituent up to per-fluoro- substitution.
  • Exemplary fluoro-substituted Q-C 2 alkyl includes -CFH 2 , CF 2 H, -CF 3 , -CH 2 CH 2 F, -CH 2 CHF 2 , -CHFCH 3 , and -CF 2 CHF 2 .
  • Per-fluoro-substituted Q-C 2 alkyl for example, includes -CF 3 and -CF 2 CF 3 .
  • the compounds disclosed herein also include partially and fully deuterated variants.
  • deuterated variants may be used for kinetic studies.
  • One of ordinary skill in the art can select the sites at which such deuterium atoms are present.
  • salts, particularly pharmaceutically acceptable salts, of the sirtuin-modulating compounds described herein are also included in the present invention.
  • the compounds of the present invention that possess a sufficiently acidic, a sufficiently basic, or both functional groups can react with any of a number of inorganic bases, and inorganic and organic acids, to form a salt.
  • compounds that are inherently charged, such as those with a quaternary nitrogen can form a salt with an appropriate counterion (e.g., a halide such as bromide, chloride, or fluoride, particularly bromide).
  • Acids commonly employed to form acid addition salts are inorganic acids such as hydrochloric acid, hydrobromic acid, hydroiodic acid, sulfuric acid, phosphoric acid, and the like, and organic acids such as p-toluenesulfonic acid, methanesulfonic acid, oxalic acid, p-bromophenyl-sulfonic acid, carbonic acid, succinic acid, citric acid, benzoic acid, acetic acid, and the like.
  • inorganic acids such as hydrochloric acid, hydrobromic acid, hydroiodic acid, sulfuric acid, phosphoric acid, and the like
  • organic acids such as p-toluenesulfonic acid, methanesulfonic acid, oxalic acid, p-bromophenyl-sulfonic acid, carbonic acid, succinic acid, citric acid, benzoic acid, acetic acid, and the like.
  • salts include the sulfate, pyrosulfate, bisulfate, sulfite, bisulfite, phosphate, monohydrogenphosphate, dihydrogenphosphate, metaphosphate, pyrophosphate, chloride, bromide, iodide, acetate, propionate, decanoate, caprylate, acrylate, formate, isobutyrate, caproate, heptanoate, propiolate, oxalate, malonate, succinate, suberate, sebacate, fumarate, maleate, butyne-l,4-dioate, hexyne-l,6-dioate, benzoate, chlorobenzoate, methylbenzoate, dinitrobenzoate, hydroxybenzoate, methoxybenzoate, phthalate, sulfonate, xylenesulfonate, phenylacetate, phenylpropionate, phenylbut
  • Base addition salts include those derived from inorganic bases, such as ammonium or alkali or alkaline earth metal hydroxides, carbonates, bicarbonates, and the like.
  • bases useful in preparing the salts of this invention thus include sodium hydroxide, potassium hydroxide, ammonium hydroxide, potassium carbonate, and the like.
  • the present invention provides methods of producing the above-defined sirtuin-modulating compounds.
  • the compounds may be synthesized using conventional techniques.
  • these compounds are conveniently synthesized from readily available starting materials.
  • Synthetic chemistry transformations and methodologies useful in synthesizing the sirtuin-modulating compounds described herein are known in the art and include, for example, those described in R. Larock, Comprehensive Organic Transformations (1989); T. W. Greene and P. G. M. Wuts, Protective Groups in Organic Synthesis, 2d. Ed. (1991); L. Fieser and M. Fieser, Fieser and Fieser's Reagents for Organic Synthesis (1994); and L. Paquette, ed., Encyclopedia of Reagents for Organic Synthesis (1995).
  • a sirtuin-modulating compound may traverse the cytoplasmic membrane of a cell.
  • a compound may have a cell- permeability of at least about 20%, 50%, 75%, 80%, 90% or 95%.
  • Sirtuin-modulating compounds described herein may also have one or more of the following characteristics: the compound may be essentially non-toxic to a cell or subject; the sirtuin-modulating compound may be an organic molecule or a small molecule of 2000 amu or less, 1000 amu or less; a compound may have a half-life under normal atmospheric conditions of at least about 30 days, 60 days, 120 days, 6 months or 1 year; the compound may have a half- life in solution of at least about 30 days, 60 days, 120 days, 6 months or 1 year; a sirtuin-modulating compound may be more stable in solution than resveratrol by at least a factor of about 50%, 2 fold, 5 fold, 10 fold, 30 fold, 50 fold or 100 fold; a sirtuin-modulating compound may promote deacetylation of the DNA repair factor Ku70; a sirtuin- modulating compound may promote deacetylation of RelA/p65; a compound may increase general turnover rates and enhance the sensitivity of
  • a sirtuin-modulating compound does not have any substantial ability to inhibit a histone deacetylase (HDACs) class I, a HDAC class II, or HDACs I and II, at concentrations (e.g., in vivo) effective for modulating the deacetylase activity of the sirtuin.
  • HDACs histone deacetylase
  • the sirtuin-modulating compound is a sirtuin-activating compound and is chosen to have an EC 50 for activating sirtuin deacetylase activity that is at least 5 fold less than the EC 50 for inhibition of an HDAC I and/or HDAC II, and even more preferably at least 10 fold, 100 fold or even 1000 fold less.
  • HDAC I and/or HDAC II activity are well known in the art and kits to perform such assays may be purchased commercially. See e.g., Bio Vision, Inc. (Mountain View, CA; world wide web at biovision.com) and Thomas Scientific (Swedesboro, NJ; world wide web at tomassci.com).
  • a sirtuin-modulating compound does not have any substantial ability to modulate sirtuin homologs.
  • an activator of a human sirtuin protein may not have any substantial ability to activate a sirtuin protein from lower eukaryotes, particularly yeast or human pathogens, at concentrations (e.g., in vivo) effective for activating the deacetylase activity of human sirtuin.
  • a sirtuin-activating compound may be chosen to have an EC 50 for activating a human sirtuin, such as SIRTl and/or SIRT3, deacetylase activity that is at least 5 fold less than the EC 50 for activating a yeast sirtuin, such as Sir2 (such as Candida, S. cerevisiae, etc.), and even more preferably at least 10 fold, 100 fold or even 1000 fold less.
  • a human sirtuin such as SIRTl and/or SIRT3
  • deacetylase activity that is at least 5 fold less than the EC 50 for activating a yeast sirtuin, such as Sir2 (such as Candida, S. cerevisiae, etc.)
  • Sir2 such as Candida, S. cerevisiae, etc.
  • an inhibitor of a sirtuin protein from lower eukaryotes, particularly yeast or human pathogens does not have any substantial ability to inhibit a sirtuin protein from humans at concentrations (e.g., in vivo) effective for inhibiting the deacetylase activity of a sirtuin protein from a lower eukaryote.
  • a sirtuin-inhibiting compound may be chosen to have an IC 50 for inhibiting a human sirtuin, such as SIRTl and/or SIRT3, deacetylase activity that is at least 5 fold less than the IC 50 for inhibiting a yeast sirtuin, such as Sir2 (such as Candida, S. cerevisiae, etc.), and even more preferably at least 10 fold, 100 fold or even 1000 fold less.
  • a sirtuin-modulating compound may have the ability to modulate one or more sirtuin protein homologs, such as, for example, one or more of human SIRTl, SIRT2, SIRT3, SIRT4, SIRT5, SIRT6, or SIRT7.
  • a sirtuin-modulating compound has the ability to modulate both a SIRTl and a SIRT3 protein.
  • a SIRTl modulator does not have any substantial ability to modulate other sirtuin protein homologs, such as, for example, one or more of human SIRT2, SIRT3, SIRT4, SIRT5, SIRT6, or SIRT7, at concentrations (e.g., in vivo) effective for modulating the deacetylase activity of human SIRTl .
  • a sirtuin-modulating compound may be chosen to have an ED 50 for modulating human SIRTl deacetylase activity that is at least 5 fold less than the ED 50 for modulating one or more of human SIRT2, SIRT3, SIRT4, SIRT5, SIRT6, or SIRT7, and even more preferably at least 10 fold, 100 fold or even 1000 fold less.
  • a SIRTl modulator does not have any substantial ability to modulate a SIRT3 protein.
  • a SIRT3 modulator does not have any substantial ability to modulate other sirtuin protein homologs, such as, for example, one or more of human SIRTl, SIRT2, SIRT4, SIRT5, SIRT6, or SIRT7, at concentrations (e.g., in vivo) effective for modulating the deacetylase activity of human SIRT3.
  • a sirtuin-modulating compound may be chosen to have an ED 50 for modulating human SIRT3 deacetylase activity that is at least 5 fold less than the ED 50 for modulating one or more of human SIRTl, SIRT2, SIRT4, SIRT5, SIRT6, or SIRT7, and even more preferably at least 10 fold, 100 fold or even 1000 fold less.
  • a SIRT3 modulator does not have any substantial ability to modulate a SIRTl protein.
  • a sirtuin-modulating compound may have a binding affinity for a sirtuin protein of about 10 "9 M, 10 "10 M, 10 "11 M, 10 "12 M or less.
  • a sirtuin-modulating compound may reduce (activator) or increase (inhibitor) the apparent Km of a sirtuin protein for its substrate or NAD+ (or other cofactor) by a factor of at least about 2, 3, 4, 5, 10, 20, 30, 50 or 100.
  • Km values are determined using the mass spectrometry assay described herein.
  • Preferred activating compounds reduce the Km of a sirtuin for its substrate or cofactor to a greater extent than caused by resveratrol at a similar concentration or reduce the Km of a sirtuin for its substrate or cofactor similar to that caused by resveratrol at a lower concentration.
  • a sirtuin-modulating compound may increase the Vmax of a sirtuin protein by a factor of at least about 2, 3, 4, 5, 10, 20, 30, 50 or 100.
  • a sirtuin-modulating compound may have an ED50 for modulating the deacetylase activity of a SIRTl and/or SIRT3 protein of less than about 1 nM, less than about 10 nM, less than about 100 nM, less than about 1 ⁇ M, less than about 10 ⁇ M, less than about 100 ⁇ M, or from about 1-10 nM, from about 10-100 nM, from about 0.1-1 ⁇ M, from about 1-10 ⁇ M or from about 10-100 ⁇ M.
  • a sirtuin- modulating compound may modulate the deacetylase activity of a SIRTl and/or SIRT3 protein by a factor of at least about 5, 10, 20, 30, 50, or 100, as measured in a cellular assay or in a cell based assay.
  • a sirtuin- activating compound may cause at least about 10%, 30%, 50%, 80%, 2 fold, 5 fold, 10 fold, 50 fold or 100 fold greater induction of the deacetylase activity of a sirtuin protein relative to the same concentration of resveratrol.
  • a sirtuin-modulating compound may have an ED50 for modulating SIRT5 that is at least about 10 fold, 20 fold, 30 fold, 50 fold greater than that for modulating SIRTl and/or SIRT3. 3. Exemplary Uses
  • the invention provides methods for modulating the level and/or activity of a sirtuin protein and methods of use thereof. In certain embodiments, the invention provides methods for increasing sirtuin- 1 activity in a cell comprising the step of contacting the cell with a compound represented by Structural Formula (VII):
  • each of Z 1 -Z 5 is independently selected from N and CR, wherein no more than two of Z x -Z 5 are simultaneously N; each R is independently selected from hydrogen, halo, -OH, -C ⁇ N, fluoro-substituted Ci-C 2 alkyl, -0-(Ci-C 2 ) fluoro-substituted alkyl, -S-(Ci-C 2 ) fluoro-substituted alkyl, C 1 -C 4 alkyl, -0-(C 1 -C 4 ) alkyl, -S-(C 1 -C 4 ) alkyl and C 3 -C 7 cycloalkyl, -(C 1 -C 2 ) alkyl-N(R 3 )(R 3 ), hydroxy-substituted C 1 -C 4 alkoxy, -(C 1 -C-O-O- saturated heterocycle, -0-(Ci
  • any heterocycle or saturated heterocycle substituent of R 1 is optionally and independently substituted at any substitutable nitrogen atom with C 1 -C 4 alkyl, fluoro- or chloro-substituted C 1 -C 4 alkyl or -(Ci-C 4 alkyl)- 0-(Ci-C 4 alkyl);
  • Ci-C 4 alkyl fluoro- or chloro-substituted Ci-C 2 alkyl, -O-( fluoro-substituted Ci-C 2 alkyl), -0-(Ci-C 4 ) alkyl, -S-(Ci-C 4 ) alkyl, -S-(fluoro-substituted Ci-C 2 alkyl), -NH-(C 1 -C 4 ) alkyl, and -N-(C r C 4 ) 2 alkyl; and any second heterocycle or saturated heterocycle substituent of R is optionally and independently substituted at any substitutable nitrogen atom with Ci-C 4 alkyl fluoro- or chloro-substituted Ci-C 4 alkyl Or-(Ci-C 4 alkyl)- 0-(Ci-C 4 alkyl); each R 3 is independently selected from hydrogen, and -Ci -C 4 alkyl, wherein the alkyl is optionally substituted with one or
  • each R is independently selected from hydrogen, halo, -OH, -C ⁇ N, fluoro-substituted Ci-C 2 alkyl, -0-(Ci-C 2 ) fluoro-substituted alkyl, -S-(Ci-C 2 ) fluoro-substituted alkyl, Ci-C 4 alkyl, -0-(Ci-C 4 ) alkyl, -S-(Ci-C 4 ) alkyl and C 3 -C 7 cycloalkyl;
  • R 1 when R 1 is phenyl, R 1 is also optionally substituted with O-(saturated heterocycle), fluoro-substituted -O-(saturated heterocycle), C 1 -C 4 alkyl- substituted unsaturated heterocycle), 3,4-methylenedioxy, fluoro-substituted
  • each R 3 is independently selected from hydrogen, and -Ci -C 4 alkyl, wherein the alkyl is optionally substituted with one or more of -OH, fluoro, -NH 2 ,
  • the invention provides methods for treating a subject suffering from or susceptible to insulin resistance, a metabolic syndrome, diabetes, or complications thereof, or for increasing insulin sensitivity in a subject, comprising administering to the subject in need thereof a compound represented by Structural Formula (VIII):
  • each of Z 1 -Z 5 is independently selected from N and CR, wherein no more than two of Z x -Z 5 are simultaneously N; each R is independently selected from hydrogen, halo, -OH, -C ⁇ N, fluoro-substituted Ci-C 2 alkyl, -0-(Ci-C 2 ) fluoro-substituted alkyl,
  • -S-(Ci-C 2 ) fluoro-substituted alkyl Ci-C 4 alkyl, -0-(Ci-C 4 ) alkyl, -S-(Ci-C 4 ) alkyl and C 3 -C 7 cycloalkyl, -(C 1 -C 2 ) alkyl-N(R 3 )(R 3 ), hydroxy-substituted C 1 -C 4 alkoxy, -(C 1 -C-O-O- saturated heterocycle, -0-(C 1 -C 3 ) alkyl-N(R 3 )(R 3 ), and -N(R 3 )(R 3 );
  • R 1 when R 1 is phenyl, R 1 is also optionally substituted with -(aryl), -(heterocycle), O-(heterocycle), -O-(carbocycle), methylenedioxy, fluoro-substituted-methylenedioxy, ethylenedioxy, or fluoro-substituted-ethylenedioxy, wherein: any aryl, cycloalkyl, carbocycle, saturated heterocycle, or heterocycle substituent of R 1 is optionally substituted at any substitutable carbon atom with one or more substituents independently selected from -OH, -C 1 -C 4 alkyl, fluoro, fluoro- or chloro-substituted C 1 -C 4 alkyl, -NH 2 , -NH(Ci-C 4 alkyl
  • Ci-C 4 alkyl, and halo-substituted C 1 -C 4 alkyl and each R 6 is independently selected from hydrogen, C 1 -C 4 alkyl, halo- substituted Ci-C 4 alkyl.
  • each R is independently selected from hydrogen, halo, -OH, -C ⁇ N, fluoro-substituted Ci-C 2 alkyl, -0-(Ci-C 2 ) fluoro-substituted alkyl, -S-(Ci-C 2 ) fluoro-substituted alkyl, Ci-C 4 alkyl, -0-(Ci-C 4 ) alkyl, -S-(Ci-C 4 ) alkyl and C 3 -C 7 cycloalkyl;
  • R 2 is selected from a carbocycle and a heterocycle, wherein R 2 is optionally substituted with one to two substitutents independently selected from halo, -C ⁇ N, Ci-C 4 alkyl, C 3 -C 7 cycloalkyl, C 1 -C 2 fluoro-substituted alkyl, -O-R 3 , -S-R 3 , -(C 1 -C 4 alkyl)-N(R 3 )(R 3 ), -N(R 3 )(R 3 ), -0-(C r C 4 alkyl)-N(R 3 )(R 3 ), -(C 1 -C 4 alkyl)-O-(C r C 4 alkyl)-N(R 3 )(R 3 ), -C(O)-N(R 3 )(R 3 ), -C(O)-N(R 3 )(R 3 ), -(O)-N(R 3 )(R 3
  • the invention provides methods for using sirtuin- modulating compounds wherein the sirtuin-modulating compounds activate a sirtuin protein, e.g., increase the level and/or activity of a sirtuin protein.
  • Sirtuin- modulating compounds that increase the level and/or activity of a sirtuin protein may be useful for a variety of therapeutic applications including, for example, increasing the lifespan of a cell, and treating and/or preventing a wide variety of diseases and disorders including, for example, diseases or disorders related to aging or stress, diabetes, obesity, neurodegenerative diseases, cardiovascular disease, blood clotting disorders, inflammation, cancer, and/or flushing, etc.
  • the methods comprise administering to a subject in need thereof a pharmaceutically effective amount of a sirtuin-modulating compound, e.g., a sirtuin-activating compound.
  • activators of the instant invention may interact with a sirtuin at the same location within the sirtuin protein (e.g., active site or site affecting the Km or Vmax of the active site). It is believed that this is the reason why certain classes of sirtuin activators and inhibitors can have substantial structural similarity.
  • the sirtuin-modulating compounds described herein may be taken alone or in combination with other compounds.
  • a mixture of two or more sirtuin-modulating compounds may be administered to a subject in need thereof.
  • a sirtuin-modulating compound that increases the level and/or activity of a sirtuin protein may be administered with one or more of the following compounds: resveratrol, butein, fisetin, piceatannol, or quercetin.
  • a sirtuin-modulating compound that increases the level and/or activity of a sirtuin protein may be administered in combination with nicotinic acid.
  • a sirtuin-modulating compound that decreases the level and/or activity of a sirtuin protein may be administered with one or more of the following compounds: nicotinamide (NAM), suramin; NF023 (a G-protein antagonist); NF279 (a purinergic receptor antagonist); Trolox (6-hydroxy-2,5,7,8,tetramethylchroman-2-carboxylic acid); (-)- epigallocatechin (hydroxy on sites 3,5,7,3',4', 5'); (-)-epigallocatechin gallate (Hydroxy sites 5,7,3',4',5' and gallate ester on 3); cyanidin chloride (3,5,7,3',4'- pentahydroxyflavylium chloride); delphinidin chloride (3,5,7,3',4',5'- hexahydroxyflavylium chloride); myricetin (cannabiscetin; 3,5,7,3',4',
  • one or more sirtuin-modulating compounds may be administered with one or more therapeutic agents for the treatment or prevention of various diseases, including, for example, cancer, diabetes, neurodegenerative diseases, cardiovascular disease, blood clotting, inflammation, flushing, obesity, aging, stress, etc.
  • combination therapies comprising a sirtuin-modulating compound may refer to (1) pharmaceutical compositions that comprise one or more sirtuin-modulating compounds in combination with one or more therapeutic agents (e.g., one or more therapeutic agents described herein); and (2) co-administration of one or more sirtuin-modulating compounds with one or more therapeutic agents wherein the sirtuin-modulating compound and therapeutic agent have not been formulated in the same compositions (but may be present within the same kit or package, such as a blister pack or other multi-chamber package; connected, separately sealed containers (e.g., foil pouches) that can be separated by the user; or a kit where the sirtuin modulating compound(s) and other therapeutic agent(s) are in separate vessels).
  • the sirtuin-modulating compound may be administered at the same, intermittent, staggered, prior to, subsequent to, or combinations thereof, with the administration of another therapeutic agent.
  • methods for reducing, preventing or treating diseases or disorders using a sirtuin-modulating compound may also comprise increasing the protein level of a sirtuin, such as human SIRTl, SIRT2 and/or SIRT3, or homologs thereof.
  • Increasing protein levels can be achieved by introducing into a cell one or more copies of a nucleic acid that encodes a sirtuin.
  • the level of a sirtuin can be increased in a mammalian cell by introducing into the mammalian cell a nucleic acid encoding the sirtuin, e.g., increasing the level of SIRTl by introducing a nucleic acid encoding the amino acid sequence set forth in GenBank Accession No. NP_036370 and/or increasing the level of SIRT3 by introducing a nucleic acid encoding the amino acid sequence set forth in GenBank Accession No. AAH01042.
  • a nucleic acid that is introduced into a cell to increase the protein level of a sirtuin may encode a protein that is at least about 80%, 85%, 90%, 95%, 98%, or 99% identical to the sequence of a sirtuin, e.g., SIRTl and/or SIRT3 protein.
  • the nucleic acid encoding the protein may be at least about 80%, 85%, 90%, 95%, 98%, or 99% identical to a nucleic acid encoding a SIRTl (e.g. GenBank Accession No. NM_012238) and/or SIRT3 (e.g., GenBank Accession No. BC001042) protein.
  • the nucleic acid may also be a nucleic acid that hybridizes, preferably under stringent hybridization conditions, to a nucleic acid encoding a wild-type sirtuin, e.g., SIRTl and/or SIRT3 protein.
  • Stringent hybridization conditions may include hybridization and a wash in 0.2 x SSC at 65 0 C.
  • the protein is preferably biologically active, e.g., is capable of deacetylation.
  • a protein that differs from wild-type SIRTl having GenBank Accession No. NP_036370 preferably contains the core structure thereof.
  • the core structure sometimes refers to amino acids 62-293 of GenBank Accession No. NP_036370, which are encoded by nucleotides 237 to 932 of GenBank Accession No. NM_012238, which encompasses the NAD binding as well as the substrate binding domains.
  • the core domain of SIRTl may also refer to about amino acids 261 to 447 of GenBank Accession No. NP_036370, which are encoded by nucleotides 834 to 1394 of GenBank Accession No.
  • NM_012238 to about amino acids 242 to 493 of GenBank Accession No. NP_036370, which are encoded by nucleotides 777 to 1532 of GenBank Accession No. NM_012238; or to about amino acids 254 to 495 of
  • methods for reducing, preventing or treating diseases or disorders using a sirtuin-modulating compound may also comprise decreasing the protein level of a sirtuin, such as human SIRTl, SIRT2 and/or SIRT3, or homologs thereof. Decreasing a sirtuin protein level can be achieved according to methods known in the art.
  • an siRNA, an antisense nucleic acid, or a ribozyme targeted to the sirtuin can be expressed in the cell.
  • a dominant negative sirtuin mutant e.g., a mutant that is not capable of deacetylating, may also be used.
  • mutant H363Y of SIRTl described, e.g., in Luo et al. (2001) Cell 107:137 can be used.
  • agents that inhibit transcription can be used.
  • Methods for modulating sirtuin protein levels also include methods for modulating the transcription of genes encoding sirtuins, methods for stabilizing/destabilizing the corresponding mRNAs, and other methods known in the art.
  • the invention provides a method extending the lifespan of a cell, extending the proliferative capacity of a cell, slowing aging of a cell, promoting the survival of a cell, delaying cellular senescence in a cell, mimicking the effects of calorie restriction, increasing the resistance of a cell to stress, or preventing apoptosis of a cell, by contacting the cell with a sirtuin-modulating compound of the invention that increases the level and/or activity of a sirtuin protein.
  • the methods comprise contacting the cell with a sirtuin-activating compound.
  • the methods described herein may be used to increase the amount of time that cells, particularly primary cells (i.e., cells obtained from an organism, e.g., a human), may be kept alive in a cell culture.
  • Embryonic stem (ES) cells and pluripotent cells, and cells differentiated therefrom may also be treated with a sirtuin-modulating compound that increases the level and/or activity of a sirtuin protein to keep the cells, or progeny thereof, in culture for longer periods of time.
  • ES Embryonic stem
  • Such cells can also be used for transplantation into a subject, e.g., after ex vivo modification.
  • cells that are intended to be preserved for long periods of time may be treated with a sirtuin-modulating compound that increases the level and/or activity of a sirtuin protein.
  • the cells may be in suspension (e.g., blood cells, serum, biological growth media, etc.) or in tissues or organs.
  • blood collected from an individual for purposes of transfusion may be treated with a sirtuin-modulating compound that increases the level and/or activity of a sirtuin protein to preserve the blood cells for longer periods of time.
  • blood to be used for forensic purposes may also be preserved using a sirtuin-modulating compound that increases the level and/or activity of a sirtuin protein.
  • Other cells that may be treated to extend their lifespan or protect against apoptosis include cells for consumption, e.g., cells from non-human mammals (such as meat) or plant cells (such as vegetables).
  • Sirtuin-modulating compounds that increase the level and/or activity of a sirtuin protein may also be applied during developmental and growth phases in mammals, plants, insects or microorganisms, in order to, e.g., alter, retard or accelerate the developmental and/or growth process.
  • sirtuin-modulating compounds that increase the level and/or activity of a sirtuin protein may be used to treat cells useful for transplantation or cell therapy, including, for example, solid tissue grafts, organ transplants, cell suspensions, stem cells, bone marrow cells, etc.
  • the cells or tissue may be an autograft, an allograft, a syngraft or a xenograft.
  • the cells or tissue may be treated with the sirtuin-modulating compound prior to administration/implantation, concurrently with administration/implantation, and/or post administration/implantation into a subject.
  • the cells or tissue may be treated prior to removal of the cells from the donor individual, ex vivo after removal of the cells or tissue from the donor individual, or post implantation into the recipient.
  • the donor or recipient individual may be treated systemically with a sirtuin-modulating compound or may have a subset of cells/tissue treated locally with a sirtuin-modulating compound that increases the level and/or activity of a sirtuin protein.
  • the cells or tissue may additionally be treated with another therapeutic agent useful for prolonging graft survival, such as, for example, an immunosuppressive agent, a cytokine, an angiogenic factor, etc.
  • cells may be treated with a sirtuin-modulating compound that increases the level and/or activity of a sirtuin protein in vivo, e.g., to increase their lifespan or prevent apoptosis.
  • skin can be protected from aging (e.g., developing wrinkles, loss of elasticity, etc.) by treating skin or epithelial cells with a sirtuin-modulating compound that increases the level and/or activity of a sirtuin protein.
  • skin is contacted with a pharmaceutical or cosmetic composition comprising a sirtuin-modulating compound that increases the level and/or activity of a sirtuin protein.
  • exemplary skin afflictions or skin conditions that may be treated in accordance with the methods described herein include disorders or diseases associated with or caused by inflammation, sun damage or natural aging.
  • compositions find utility in the prevention or treatment of contact dermatitis (including irritant contact dermatitis and allergic contact dermatitis), atopic dermatitis (also known as allergic eczema), actinic keratosis, keratinization disorders (including eczema), epidermolysis bullosa diseases (including pemphigus), exfoliative dermatitis, seborrheic dermatitis, erythemas (including erythema multiforme and erythema nodosum), damage caused by the sun or other light sources, discoid lupus erythematosus, dermatomyositis, psoriasis, skin cancer and the effects of natural aging.
  • contact dermatitis including irritant contact dermatitis and allergic contact dermatitis
  • atopic dermatitis also known as allergic eczema
  • actinic keratosis also known as allergic eczema
  • sirtuin-modulating compounds that increase the level and/or activity of a sirtuin protein may be used for the treatment of wounds and/or burns to promote healing, including, for example, first-, second- or third- degree burns and/or thermal, chemical or electrical burns.
  • the formulations may be administered topically, to the skin or mucosal tissue.
  • Topical formulations comprising one or more sirtuin-modulating compounds that increase the level and/or activity of a sirtuin protein may also be used as preventive, e.g., chemopreventive, compositions. When used in a chemopreventive method, susceptible skin is treated prior to any visible condition in a particular individual.
  • Sirtuin-modulating compounds may be delivered locally or systemically to a subject.
  • a sirtuin-modulating compound is delivered locally to a tissue or organ of a subject by injection, topical formulation, etc.
  • a sirtuin-modulating compound that increases the level and/or activity of a sirtuin protein may be used for treating or preventing a disease or condition induced or exacerbated by cellular senescence in a subject; methods for decreasing the rate of senescence of a subject, e.g., after onset of senescence; methods for extending the lifespan of a subject; methods for treating or preventing a disease or condition relating to lifespan; methods for treating or preventing a disease or condition relating to the proliferative capacity of cells; and methods for treating or preventing a disease or condition resulting from cell damage or death.
  • the method does not act by decreasing the rate of occurrence of diseases that shorten the lifespan of a subject.
  • a method does not act by reducing the lethality caused by a disease, such as cancer.
  • a sirtuin-modulating compound that increases the level and/or activity of a sirtuin protein may be administered to a subject in order to generally increase the lifespan of its cells and to protect its cells against stress and/or against apoptosis. It is believed that treating a subject with a compound described herein is similar to subjecting the subject to hormesis, i.e., mild stress that is beneficial to organisms and may extend their lifespan.
  • Sirtuin-modulating compounds that increase the level and/or activity of a sirtuin protein may be administered to a subject to prevent aging and aging-related consequences or diseases, such as stroke, heart disease, heart failure, arthritis, high blood pressure, and Alzheimer's disease.
  • Other conditions that can be treated include ocular disorders, e.g., associated with the aging of the eye, such as cataracts, glaucoma, and macular degeneration.
  • Sirtuin-modulating compounds that increase the level and/or activity of a sirtuin protein can also be administered to subjects for treatment of diseases, e.g., chronic diseases, associated with cell death, in order to protect the cells from cell death.
  • Exemplary diseases include those associated with neural cell death, neuronal dysfunction, or muscular cell death or dysfunction, such as Parkinson's disease, Alzheimer's disease, multiple sclerosis, amyotrophic, amniotropic lateral sclerosis, and muscular dystrophy; AIDS; fulminant hepatitis; diseases linked to degeneration of the brain, such as Creutzfeld- Jakob disease, retinitis pigmentosa and cerebellar degeneration; myelodysplasia such as aplastic anemia; ischemic diseases such as myocardial infarction and stroke; hepatic diseases such as alcoholic hepatitis, hepatitis B and hepatitis C; joint-diseases such as osteoarthritis; atherosclerosis; alopecia; damage to the skin due to UV light; lichen planus; atrophy of the skin; cataract; and graft rejections.
  • Parkinson's disease Alzheimer's disease, multiple sclerosis, amyotrophic, amniotropic
  • Sirtuin-modulating compounds that increase the level and/or activity of a sirtuin protein can also be administered to a subject suffering from an acute disease, e.g., damage to an organ or tissue, e.g., a subject suffering from stroke or myocardial infarction or a subject suffering from a spinal cord injury. Sirtuin- modulating compounds that increase the level and/or activity of a sirtuin protein may also be used to repair an alcoholic's liver. Cardiovascular Disease
  • the invention provides a method for treating and/or preventing a cardiovascular disease by administering to a subject in need thereof a sirtuin-modulating compound that increases the level and/or activity of a sirtuin protein.
  • Cardiovascular diseases that can be treated or prevented using the sirtuin- modulating compounds that increase the level and/or activity of a sirtuin protein include cardiomyopathy or myocarditis; such as idiopathic cardiomyopathy, metabolic cardiomyopathy, alcoholic cardiomyopathy, drug-induced cardiomyopathy, ischemic cardiomyopathy, and hypertensive cardiomyopathy.
  • cardiomyopathy or myocarditis such as idiopathic cardiomyopathy, metabolic cardiomyopathy, alcoholic cardiomyopathy, drug-induced cardiomyopathy, ischemic cardiomyopathy, and hypertensive cardiomyopathy.
  • atheromatous disorders of the major blood vessels such as the aorta, the coronary arteries, the carotid arteries, the cerebrovascular arteries, the renal arteries, the iliac arteries, the femoral arteries, and the popliteal arteries.
  • vascular diseases that can be treated or prevented include those related to platelet aggregation, the retinal arterioles, the glomerular arterioles, the vasa nervorum, cardiac arterioles, and associated capillary beds of the eye, the kidney, the heart, and the central and peripheral nervous systems.
  • the sirtuin-modulating compounds that increase the level and/or activity of a sirtuin protein may also be used for increasing HDL levels in plasma of an individual.
  • a sirtuin-modulating compound that increases the level and/or activity of a sirtuin protein may be administered as part of a combination therapeutic with another cardiovascular agent.
  • a sirtuin- modulating compound that increases the level and/or activity of a sirtuin protein may be administered as part of a combination therapeutic with an anti- arrhythmia agent.
  • a sirtuin-modulating compound that increases the level and/or activity of a sirtuin protein may be administered as part of a combination therapeutic with another cardiovascular agent.
  • Sirtuin-modulating compounds that increase the level and/or activity of a sirtuin protein may be administered to subjects who have recently received or are likely to receive a dose of radiation or toxin.
  • the dose of radiation or toxin is received as part of a work-related or medical procedure, e.g., administered as a prophylactic measure.
  • the radiation or toxin exposure is received unintentionally.
  • the compound is preferably administered as soon as possible after the exposure to inhibit apoptosis and the subsequent development of acute radiation syndrome.
  • Sirtuin-modulating compounds may also be used for treating and/or preventing cancer.
  • sirtuin-modulating compounds that increase the level and/or activity of a sirtuin protein may be used for treating and/or preventing cancer.
  • Calorie restriction has been linked to a reduction in the incidence of age-related disorders including cancer.
  • an increase in the level and/or activity of a sirtuin protein may be useful for treating and/or preventing the incidence of age-related disorders, such as, for example, cancer.
  • Exemplary cancers that may be treated using a sirtuin-modulating compound are those of the brain and kidney; hormone-dependent cancers including breast, prostate, testicular, and ovarian cancers; lymphomas, and leukemias.
  • a modulating compound may be administered directly into the tumor.
  • Cancer of blood cells e.g., leukemia
  • Benign cell growth e.g., warts
  • Other diseases that can be treated include autoimmune diseases, e.g., systemic lupus erythematosus, scleroderma, and arthritis, in which autoimmune cells should be removed.
  • Viral infections such as herpes, HIV, adenovirus, and HTLV-I associated malignant and benign disorders can also be treated by administration of sirtuin-modulating compound.
  • cells can be obtained from a subject, treated ex vivo to remove certain undesirable cells, e.g., cancer cells, and administered back to the same or a different subject.
  • Chemotherapeutic agents may be co-administered with modulating compounds described herein as having anti-cancer activity, e.g., compounds that induce apoptosis, compounds that reduce lifespan or compounds that render cells sensitive to stress. Chemotherapeutic agents may be used by themselves with a sirtuin-modulating compound described herein as inducing cell death or reducing lifespan or increasing sensitivity to stress and/or in combination with other chemotherapeutics agents. In addition to conventional chemotherapeutics, the sirtuin-modulating compounds described herein may also be used with antisense RNA, RNAi or other polynucleotides to inhibit the expression of the cellular components that contribute to unwanted cellular proliferation.
  • Combination therapies comprising sirtuin-modulating compounds and a conventional chemotherapeutic agent may be advantageous over combination therapies known in the art because the combination allows the conventional chemotherapeutic agent to exert greater effect at lower dosage.
  • the effective dose (ED 50 ) for a chemo therapeutic agent, or combination of conventional chemotherapeutic agents, when used in combination with a sirtuin-modulating compound is at least 2 fold less than the ED 50 for the chemotherapeutic agent alone, and even more preferably at 5 fold, 10 fold or even 25 fold less.
  • the therapeutic index (TI) for such chemotherapeutic agent or combination of such chemotherapeutic agent when used in combination with a sirtuin-modulating compound described herein can be at least 2 fold greater than the TI for conventional chemotherapeutic regimen alone, and even more preferably at 5 fold, 10 fold or even 25 fold greater.
  • sirtuin-modulating compounds that increase the level and/or activity of a sirtuin protein can be used to treat patients suffering from neurodegenerative diseases, and traumatic or mechanical injury to the central nervous system (CNS), spinal cord or peripheral nervous system (PNS).
  • Neurodegenerative disease typically involves reductions in the mass and volume of the human brain, which may be due to the atrophy and/or death of brain cells, which are far more profound than those in a healthy person that are attributable to aging.
  • Neurodegenerative diseases can evolve gradually, after a long period of normal brain function, due to progressive degeneration (e.g., nerve cell dysfunction and death) of specific brain regions.
  • neurodegenerative diseases can have a quick onset, such as those associated with trauma or toxins.
  • neurodegenerative diseases include, but are not limited to, Alzheimer's disease (AD), Parkinson's disease (PD), Huntington's disease (HD), amyotrophic lateral sclerosis (ALS; Lou Gehrig's disease), diffuse Lewy body disease, chorea- acanthocytosis, primary lateral sclerosis, ocular diseases (ocular neuritis), chemotherapy-induced neuropathies (e.g., from vincristine, paclitaxel, bortezomib), diabetes-induced neuropathies and Friedreich's ataxia.
  • Sirtuin-modulating compounds that increase the level and/or activity of a sirtuin protein can be used to treat these disorders and others as described below.
  • AD is a CNS disorder that results in memory loss, unusual behavior, personality changes, and a decline in thinking abilities. These losses are related to the death of specific types of brain cells and the breakdown of connections and their supporting network (e.g. glial cells) between them. The earliest symptoms include loss of recent memory, faulty judgment, and changes in personality.
  • PD is a CNS disorder that results in uncontrolled body movements, rigidity, tremor, and dyskinesia, and is associated with the death of brain cells in an area of the brain that produces dopamine.
  • ALS motor neuron disease
  • ALS motor neuron disease
  • HD is another neurodegenerative disease that causes uncontrolled movements, loss of intellectual faculties, and emotional disturbance.
  • Tay-Sachs disease and Sandhoff disease are glycolipid storage diseases where GM2 ganglioside and related glycolipidssubstrates glycolipids substrates for ⁇ -hexosaminidase accumulate in the nervous system and trigger acute neurodegeneration.
  • HIV-I also induces neurological disease, which can be treated with sirtuin-modulating compounds of the invention.
  • Neuronal loss is also a salient feature of prion diseases, such as Creutzfeldt- Jakob disease in human, BSE in cattle (mad cow disease), Scrapie Disease in sheep and goats, and feline spongiform encephalopathy (FSE) in cats.
  • Sirtuin-modulating compounds that increase the level and/or activity of a sirtuin protein may be useful for treating or preventing neuronal loss due to these prior diseases.
  • a sirtuin-modulating compound that increases the level and/or activity of a sirtuin protein may be used to treat or prevent any disease or disorder involving axonopathy.
  • Distal axonopathy is a type of peripheral neuropathy that results from some metabolic or toxic derangement of peripheral nervous system (PNS) neurons. It is the most common response of nerves to metabolic or toxic disturbances, and as such may be caused by metabolic diseases such as diabetes, renal failure, deficiency syndromes such as malnutrition and alcoholism, or the effects of toxins or drugs.
  • PNS peripheral nervous system
  • Those with distal axonopathies usually present with symmetrical glove- stocking sensori-motor disturbances.
  • Diabetic neuropathies are neuropathic disorders that are associated with diabetes mellitus. Relatively common conditions which may be associated with diabetic neuropathy include third nerve palsy; mononeuropathy; mononeuritis multiplex; diabetic amyotrophy; a painful polyneuropathy; autonomic neuropathy; and thoracoabdominal neuropathy.
  • Peripheral neuropathy is the medical term for damage to nerves of the peripheral nervous system, which may be caused either by diseases of the nerve or from the side-effects of systemic illness.
  • Major causes of peripheral neuropathy include seizures, nutritional deficiencies, and HIV, though diabetes is the most likely cause.
  • a sirtuin-modulating compound that increases the level and/or activity of a sirtuin protein may be used to treat or prevent multiple sclerosis (MS), including relapsing MS and monosymptomatic MS, and other demyelinating conditions, such as, for example, chromic inflammatory demyelinating polyneuropathy (CIDP), or symptoms associated therewith.
  • MS multiple sclerosis
  • CIDP chromic inflammatory demyelinating polyneuropathy
  • a sirtuin-modulating compound that increases the level and/or activity of a sirtuin protein may be used to treat trauma to the nerves, including, trauma due to disease, injury (including surgical intervention), or environmental trauma (e.g., neurotoxins, alcoholism, etc.).
  • Sirtuin-modulating compounds that increase the level and/or activity of a sirtuin protein may also be useful to prevent, treat, and alleviate symptoms of various PNS disorders.
  • the term "peripheral neuropathy” encompasses a wide range of disorders in which the nerves outside of the brain and spinal cord — peripheral nerves — have been damaged. Peripheral neuropathy may also be referred to as peripheral neuritis, or if many nerves are involved, the terms polyneuropathy or polyneuritis may be used.
  • PNS diseases treatable with sirtuin-modulating compounds that increase the level and/or activity of a sirtuin protein include: diabetes, leprosy, Charcot-Marie- Tooth disease, Guillain-Barre syndrome and Brachial Plexus Neuropathies (diseases of the cervical and first thoracic roots, nerve trunks, cords, and peripheral nerve components of the brachial plexus.
  • a sirtuin activating compound may be used to treat or prevent a polyglutamine disease.
  • Exemplary polyglutamine diseases include Spinobulbar muscular atrophy (Kennedy disease), Huntington's Disease (HD), Dentatorubral-pallidoluysian atrophy (Haw River syndrome), Spinocerebellar ataxia type 1, Spinocerebellar ataxia type 2, Spinocerebellar ataxia type 3 (Machado- Joseph disease), Spinocerebellar ataxia type 6, Spinocerebellar ataxia type 7, and Spinocerebellar ataxia type 17.
  • the invention provides a method to treat a central nervous system cell to prevent damage in response to a decrease in blood flow to the cell.
  • the severity of damage that may be prevented will depend in large part on the degree of reduction in blood flow to the cell and the duration of the reduction.
  • apoptotic or necrotic cell death may be prevented.
  • ischemic-mediated damage such as cytoxic edema or central nervous system tissue anoxemia, may be prevented.
  • the central nervous system cell may be a spinal cell or a brain cell.
  • the ischemic condition is a stroke that results in any type of ischemic central nervous system damage, such as apoptotic or necrotic cell death, cytoxic edema or central nervous system tissue anoxia.
  • the stroke may impact any area of the brain or be caused by any etiology commonly known to result in the occurrence of a stroke.
  • the stroke is a brain stem stroke.
  • the stroke is a cerebellar stroke.
  • the stroke is an embolic stroke.
  • the stroke may be a hemorrhagic stroke.
  • the stroke is a thrombotic stroke.
  • a sirtuin activating compound may be administered to reduce infarct size of the ischemic core following a central nervous system ischemic condition. Moreover, a sirtuin activating compound may also be beneficially administered to reduce the size of the ischemic penumbra or transitional zone following a central nervous system ischemic condition.
  • a combination drug regimen may include drugs or compounds for the treatment or prevention of neurodegenerative disorders or secondary conditions associated with these conditions.
  • a combination drug regimen may include one or more sirtuin activators and one or more anti- neurodegeneration agents.
  • sirtuin-modulating compounds that increase the level and/or activity of a sirtuin protein can be used to treat or prevent blood coagulation disorders (or hemostatic disorders).
  • blood coagulation disorders or hemostatic disorders
  • the terms “hemostasis”, “blood coagulation,” and “blood clotting” refer to the control of bleeding, including the physiological properties of vasoconstriction and coagulation. Blood coagulation assists in maintaining the integrity of mammalian circulation after injury, inflammation, disease, congenital defect, dysfunction or other disruption. Further, the formation of blood clots does not only limit bleeding in case of an injury (hemostasis), but may lead to serious organ damage and death in the context of atherosclerotic diseases by occlusion of an important artery or vein. Thrombosis is thus blood clot formation at the wrong time and place.
  • the present invention provides anticoagulation and antithrombotic treatments aiming at inhibiting the formation of blood clots in order to prevent or treat blood coagulation disorders, such as myocardial infarction, stroke, loss of a limb by peripheral artery disease or pulmonary embolism.
  • blood coagulation disorders such as myocardial infarction, stroke, loss of a limb by peripheral artery disease or pulmonary embolism.
  • modulating or modulation of hemostasis includes the induction (e.g., stimulation or increase) of hemostasis, as well as the inhibition (e.g., reduction or decrease) of hemostasis.
  • the invention provides a method for reducing or inhibiting hemostasis in a subject by administering a sirtuin-modulating compound that increases the level and/or activity of a sirtuin protein.
  • a sirtuin-modulating compound that increases the level and/or activity of a sirtuin protein.
  • the compositions and methods disclosed herein are useful for the treatment or prevention of thrombotic disorders.
  • thrombotic disorder includes any disorder or condition characterized by excessive or unwanted coagulation or hemostatic activity, or a hypercoagulable state.
  • Thrombotic disorders include diseases or disorders involving platelet adhesion and thrombus formation, and may manifest as an increased propensity to form thromboses, e.g., an increased number of thromboses, thrombosis at an early age, a familial tendency towards thrombosis, and thrombosis at unusual sites.
  • a combination drug regimen may include drugs or compounds for the treatment or prevention of blood coagulation disorders or secondary conditions associated with these conditions.
  • a combination drug regimen may include one or more sirtuin-modulating compounds that increase the level and/or activity of a sirtuin protein and one or more anti-coagulation or anti- thrombosis agents.
  • sirtuin-modulating compounds that increase the level and/or activity of a sirtuin protein may be used for treating or preventing weight gain or obesity in a subject.
  • sirtuin-modulating compounds that increase the level and/or activity of a sirtuin protein may be used, for example, to treat or prevent hereditary obesity, dietary obesity, hormone related obesity, obesity related to the administration of medication, to reduce the weight of a subject, or to reduce or prevent weight gain in a subject.
  • a subject in need of such a treatment may be a subject who is obese, likely to become obese, overweight, or likely to become overweight.
  • Subjects who are likely to become obese or overweight can be identified, for example, based on family history, genetics, diet, activity level, medication intake, or various combinations thereof.
  • sirtuin-modulating compounds that increase the level and/or activity of a sirtuin protein may be administered to subjects suffering from a variety of other diseases and conditions that may be treated or prevented by promoting weight loss in the subject.
  • diseases include, for example, high blood pressure, hypertension, high blood cholesterol, dyslipidemia, type 2 diabetes, insulin resistance, glucose intolerance, hyperinsulinemia, coronary heart disease, angina pectoris, congestive heart failure, stroke, gallstones, cholecystitis and cholelithiasis, gout, osteoarthritis, obstructive sleep apnea and respiratory problems, some types of cancer (such as endometrial, breast, prostate, and colon), complications of pregnancy, poor female reproductive health (such as menstrual irregularities, infertility, irregular ovulation), bladder control problems (such as stress incontinence); uric acid nephrolithiasis; psychological disorders (such as depression, eating disorders, distorted body image,
  • sirtuin-modulating compounds that increase the level and/or activity of a sirtuin protein may be used for inhibiting adipogenesis or fat cell differentiation, whether in vitro or in vivo. Such methods may be used for treating or preventing obesity.
  • sirtuin-modulating compounds that increase the level and/or activity of a sirtuin protein may be used for reducing appetite and/or increasing satiety, thereby causing weight loss or avoidance of weight gain.
  • a subject in need of such a treatment may be a subject who is overweight, obese or a subject likely to become overweight or obese.
  • the method may comprise administering daily or, every other day, or once a week, a dose, e.g., in the form of a pill, to a subject.
  • the dose may be an "appetite reducing dose.”
  • sirtuin-modulating compounds that increase the level and/or activity of a sirtuin protein may be administered as a combination therapy for treating or preventing weight gain or obesity.
  • one or more sirtuin-modulating compounds that increase the level and/or activity of a sirtuin protein may be administered in combination with one or more anti-obesity agents.
  • sirtuin-modulating compounds that increase the level and/or activity of a sirtuin protein may be administered to reduce drug-induced weight gain.
  • a sirtuin-modulating compound that increases the level and/or activity of a sirtuin protein may be administered as a combination therapy with medications that may stimulate appetite or cause weight gain, in particular, weight gain due to factors other than water retention.
  • sirtuin-modulating compounds that increase the level and/or activity of a sirtuin protein may be used for treating or preventing a metabolic disorder, such as insulin-resistance, a pre-diabetic state, type II diabetes, and/or complications thereof.
  • Administration of a sirtuin-modulating compounds that increases the level and/or activity of a sirtuin protein may increase insulin sensitivity and/or decrease insulin levels in a subject.
  • a subject in need of such a treatment may be a subject who has insulin resistance or other precursor symptom of type II diabetes, who has type II diabetes, or who is likely to develop any of these conditions.
  • the subject may be a subject having insulin resistance, e.g., having high circulating levels of insulin and/or associated conditions, such as hyperlipidemia, dyslipogenesis, hypercholesterolemia, impaired glucose tolerance, high blood glucose sugar level, other manifestations of syndrome X, hypertension, atherosclerosis and lipodystrophy.
  • insulin resistance e.g., having high circulating levels of insulin and/or associated conditions, such as hyperlipidemia, dyslipogenesis, hypercholesterolemia, impaired glucose tolerance, high blood glucose sugar level, other manifestations of syndrome X, hypertension, atherosclerosis and lipodystrophy.
  • sirtuin-modulating compounds that increase the level and/or activity of a sirtuin protein may be administered as a combination therapy for treating or preventing a metabolic disorder.
  • one or more sirtuin-modulating compounds that increase the level and/or activity of a sirtuin protein may be administered in combination with one or more anti-diabetic agents.
  • sirtuin-modulating compounds that increase the level and/or activity of a sirtuin protein can be used to treat or prevent a disease or disorder associated with inflammation.
  • Sirtuin-modulating compounds that increase the level and/or activity of a sirtuin protein may be administered prior to the onset of, at, or after the initiation of inflammation.
  • the compounds are preferably provided in advance of any inflammatory response or symptom. Administration of the compounds may prevent or attenuate inflammatory responses or symptoms.
  • sirtuin-modulating compounds that increase the level and/or activity of a sirtuin protein may be used to treat or prevent allergies and respiratory conditions, including asthma, bronchitis, pulmonary fibrosis, allergic rhinitis, oxygen toxicity, emphysema, chronic bronchitis, acute respiratory distress syndrome, and any chronic obstructive pulmonary disease (COPD).
  • the compounds may be used to treat chronic hepatitis infection, including hepatitis B and hepatitis C.
  • sirtuin-modulating compounds that increase the level and/or activity of a sirtuin protein may be used to treat autoimmune diseases, and/or inflammation associated with autoimmune diseases such as arthritis, including rheumatoid arthritis, psoriatic arthritis, and ankylosing spondylitis, as well as organ-tissue autoimmune diseases (e.g., Raynaud's syndrome), ulcerative colitis, Crohn's disease, oral mucositis, scleroderma, myasthenia gravis, transplant rejection, endotoxin shock, sepsis, psoriasis, eczema, dermatitis, multiple sclerosis, autoimmune thyroiditis, uveitis, systemic lupus erythematosis, Addison's disease, autoimmune polyglandular disease (also known as autoimmune polyglandular syndrome), and Grave's disease.
  • autoimmune diseases such as arthritis, including rheumatoid arthritis,
  • one or more sirtuin-modulating compounds that increase the level and/or activity of a sirtuin protein may be taken alone or in combination with other compounds useful for treating or preventing inflammation. Flushing In another aspect, sirtuin-modulating compounds that increase the level and/or activity of a sirtuin protein may be used for reducing the incidence or severity of flushing and/or hot flashes which are symptoms of a disorder.
  • the subject method includes the use of sirtuin-modulating compounds that increase the level and/or activity of a sirtuin protein, alone or in combination with other agents, for reducing incidence or severity of flushing and/or hot flashes in cancer patients.
  • the method provides for the use of sirtuin-modulating compounds that increase the level and/or activity of a sirtuin protein to reduce the incidence or severity of flushing and/or hot flashes in menopausal and postmenopausal woman.
  • sirtuin-modulating compounds that increase the level and/or activity of a sirtuin protein may be used as a therapy for reducing the incidence or severity of flushing and/or hot flashes which are side-effects of another drug therapy, e.g., drug-induced flushing.
  • a method for treating and/or preventing drug-induced flushing comprises administering to a patient in need thereof a formulation comprising at least one flushing inducing compound and at least one sirtuin-modulating compound that increases the level and/or activity of a sirtuin protein.
  • a method for treating drug induced flushing comprises separately administering one or more compounds that induce flushing and one or more sirtuin-modulating compounds, e.g., wherein the sirtuin-modulating compound and flushing inducing agent have not been formulated in the same compositions.
  • the sirtuin-modulating compound may be administered (1) at the same as administration of the flushing inducing agent, (2) intermittently with the flushing inducing agent, (3) staggered relative to administration of the flushing inducing agent, (4) prior to administration of the flushing inducing agent, (5) subsequent to administration of the flushing inducing agent, and (6) various combination thereof.
  • flushing inducing agents include, for example, niacin, raloxifene, antidepressants, anti-psychotics, chemotherapeutics, calcium channel blockers, and antibiotics.
  • sirtuin-modulating compounds that increase the level and/or activity of a sirtuin protein may be used to reduce flushing side effects of a vasodilator or an antilipemic agent (including anticholesteremic agents and lipotropic agents).
  • a sirtuin-modulating compound that increases the level and/or activity of a sirtuin protein may be used to reduce flushing associated with the administration of niacin.
  • the invention provides a method for treating and/or preventing hyperlipidemia with reduced flushing side effects.
  • the method involves the use of sirtuin-modulating compounds that increase the level and/or activity of a sirtuin protein to reduce flushing side effects of raloxifene.
  • the method involves the use of sirtuin-modulating compounds that increase the level and/or activity of a sirtuin protein to reduce flushing side effects of antidepressants or anti-psychotic agent.
  • sirtuin-modulating compounds that increase the level and/or activity of a sirtuin protein can be used in conjunction (administered separately or together) with a serotonin reuptake inhibitor, or a 5HT2 receptor antagonist.
  • sirtuin-modulating compounds that increase the level and/or activity of a sirtuin protein may be used as part of a treatment with a serotonin reuptake inhibitor (SRI) to reduce flushing.
  • SRI serotonin reuptake inhibitor
  • sirtuin-modulating compounds that increase the level and/or activity of a sirtuin protein may be used to reduce flushing side effects of chemotherapeutic agents, such as cyclophosphamide and tamoxifen.
  • sirtuin-modulating compounds that increase the level and/or activity of a sirtuin protein may be used to reduce flushing side effects of calcium channel blockers, such as amlodipine.
  • sirtuin-modulating compounds that increase the level and/or activity of a sirtuin protein may be used to reduce flushing side effects of antibiotics.
  • sirtuin-modulating compounds that increase the level and/or activity of a sirtuin protein can be used in combination with levofloxacin.
  • One aspect of the present invention is a method for inhibiting, reducing or otherwise treating vision impairment by administering to a patient a therapeutic dosage of sirtuin modulator selected from a compound disclosed herein, or a pharmaceutically acceptable salt, prodrug or a metabolic derivative thereof.
  • the vision impairment is caused by damage to the optic nerve or central nervous system.
  • optic nerve damage is caused by high intraocular pressure, such as that created by glaucoma.
  • optic nerve damage is caused by swelling of the nerve, which is often associated with an infection or an immune (e.g., autoimmune) response such as in optic neuritis.
  • the vision impairment is caused by retinal damage.
  • retinal damage is caused by disturbances in blood flow to the eye (e.g., arteriosclerosis, vasculitis).
  • retinal damage is caused by disruption of the macula (e.g., exudative or non- exudative macular degeneration).
  • Exemplary retinal diseases include Exudative Age Related Macular Degeneration, Nonexudative Age Related Macular Degeneration, Retinal Electronic Prosthesis and RPE Transplantation Age Related Macular Degeneration, Acute Multifocal Placoid Pigment Epitheliopathy, Acute Retinal Necrosis, Best Disease, Branch Retinal Artery Occlusion, Branch Retinal Vein Occlusion, Cancer Associated and Related Autoimmune Retinopathies, Central Retinal Artery Occlusion, Central Retinal Vein Occlusion, Central Serous Chorioretinopathy, Eales Disease, Epimacular Membrane, Lattice Degeneration, Macroaneurysm, Diabetic Macular Edema, Irvine-Gass Macular Edema, Macular Hole, Subretinal Neovascular Membranes, Diffuse Unilateral Subacute Neuroretinitis, Nonpseudophakic Cystoid Macular Edema, Presumed Ocular Histoplasmosis Syndrome,
  • exemplary diseases include ocular bacterial infections (e.g. conjunctivitis, keratitis, tuberculosis, syphilis, gonorrhea), viral infections (e.g. Ocular Herpes Simplex Virus, Varicella Zoster Virus, Cytomegalovirus retinitis,
  • ocular bacterial infections e.g. conjunctivitis, keratitis, tuberculosis, syphilis, gonorrhea
  • viral infections e.g. Ocular Herpes Simplex Virus, Varicella Zoster Virus, Cytomegalovirus retinitis,
  • ocular diseases include fungal infections (e.g. Candida choroiditis, histoplasmosis), protozoal infections (e.g. toxoplasmosis) and others such as ocular toxocariasis and sarcoidosis.
  • fungal infections e.g. Candida choroiditis, histoplasmosis
  • protozoal infections e.g. toxoplasmosis
  • others such as ocular toxocariasis and sarcoidosis.
  • One aspect of the invention is a method for inhibiting, reducing or treating vision impairment in a subject undergoing treatment with a chemotherapeutic drug (e.g., a neurotoxic drug, a drug that raises intraocular pressure such as a steroid), by administering to the subject in need of such treatment a therapeutic dosage of a sirtuin modulator disclosed herein.
  • a chemotherapeutic drug e.g., a neurotoxic drug, a drug that raises intraocular pressure such as a steroid
  • Another aspect of the invention is a method for inhibiting, reducing or treating vision impairment in a subject undergoing surgery, including ocular or other surgeries performed in the prone position such as spinal cord surgery, by administering to the subject in need of such treatment a therapeutic dosage of a sirtuin modulator disclosed herein.
  • Ocular surgeries include cataract, iridotomy and lens replacements.
  • Another aspect of the invention is the treatment, including inhibition and prophylactic treatment, of age related ocular diseases include cataracts, dry eye, age- related macular degeneration (AMD), retinal damage and the like, by administering to the subject in need of such treatment a therapeutic dosage of a sirtuin modulator disclosed herein.
  • Another aspect of the invention is the prevention or treatment of damage to the eye caused by stress, chemical insult or radiation, by administering to the subject in need of such treatment a therapeutic dosage of a sirtuin modulator disclosed herein.
  • Radiation or electromagnetic damage to the eye can include that caused by CRT's or exposure to sunlight or UV.
  • a combination drug regimen may include drugs or compounds for the treatment or prevention of ocular disorders or secondary conditions associated with these conditions.
  • a combination drug regimen may include one or more sirtuin activators and one or more therapeutic agents for the treatment of an ocular disorder.
  • a sirtuin modulator can be administered in conjunction with a therapy for reducing intraocular pressure. In another embodiment, a sirtuin modulator can be administered in conjunction with a therapy for treating and/or preventing glaucoma. In yet another embodiment, a sirtuin modulator can be administered in conjunction with a therapy for treating and/or preventing optic neuritis. In one embodiment, a sirtuin modulator can be administered in conjunction with a therapy for treating and/or preventing CMV Retinopathy. In another embodiment, a sirtuin modulator can be administered in conjunction with a therapy for treating and/or preventing multiple sclerosis. Mitochondrial-Associated Diseases and Disorders
  • the invention provides methods for treating diseases or disorders that would benefit from increased mitochondrial activity.
  • the methods involve administering to a subject in need thereof a therapeutically effective amount of a sirtuin activating compound.
  • Increased mitochondrial activity refers to increasing activity of the mitochondria while maintaining the overall numbers of mitochondria (e.g., mitochondrial mass), increasing the numbers of mitochondria thereby increasing mitochondrial activity (e.g., by stimulating mitochondrial biogenesis), or combinations thereof.
  • diseases and disorders that would benefit from increased mitochondrial activity include diseases or disorders associated with mitochondrial dysfunction.
  • methods for treating diseases or disorders that would benefit from increased mitochondrial activity may comprise identifying a subject suffering from a mitochondrial dysfunction.
  • Methods for diagnosing a mitochondrial dysfunction may involve molecular genetic, pathologic and/or biochemical analyses.
  • Diseases and disorders associated with mitochondrial dysfunction include diseases and disorders in which deficits in mitochondrial respiratory chain activity contribute to the development of pathophysiology of such diseases or disorders in a mammal.
  • Diseases or disorders that would benefit from increased mitochondrial activity generally include for example, diseases in which free radical mediated oxidative injury leads to tissue degeneration, diseases in which cells inappropriately undergo apoptosis, and diseases in which cells fail to undergo apoptosis.
  • the invention provides methods for treating a disease or disorder that would benefit from increased mitochondrial activity that involves administering to a subject in need thereof one or more sirtuin activating compounds in combination with another therapeutic agent such as, for example, an agent useful for treating mitochondrial dysfunction or an agent useful for reducing a symptom associated with a disease or disorder involving mitochondrial dysfunction.
  • the invention provides methods for treating diseases or disorders that would benefit from increased mitochondrial activity by administering to a subject a therapeutically effective amount of a sirtuin activating compound.
  • diseases or disorders include, for example, neuromuscular disorders (e.g., Friedreich's Ataxia, muscular dystrophy, multiple sclerosis, etc.), disorders of neuronal instability (e.g., seizure disorders, migraine, etc.), developmental delay, neurodegenerative disorders (e.g., Alzheimer's Disease, Parkinson's Disease, amyotrophic lateral sclerosis, etc.), ischemia, renal tubular acidosis, age-related neurodegeneration and cognitive decline, chemotherapy fatigue, age-related or chemotherapy-induced menopause or irregularities of menstrual cycling or ovulation, mitochondrial myopathies, mitochondrial damage (e.g., calcium accumulation, excitotoxicity, nitric oxide exposure, hypoxia, etc.), and mitochondrial deregulation.
  • mitochondrial myopathies e.g., calcium accumulation, excitotoxicity,
  • Muscular dystrophy refers to a family of diseases involving deterioration of neuromuscular structure and function, often resulting in atrophy of skeletal muscle and myocardial dysfunction, such as Duchenne muscular dystrophy.
  • sirtuin activating compounds may be used for reducing the rate of decline in muscular functional capacities and for improving muscular functional status in patients with muscular dystrophy.
  • sirtuin modulating compounds may be useful for treatment mitochondrial myopathies.
  • Mitochondrial myopathies range from mild, slowly progressive weakness of the extraocular muscles to severe, fatal infantile myopathies and multisystem encephalomyopathies. Some syndromes have been defined, with some overlap between them.
  • Established syndromes affecting muscle include progressive external ophthalmoplegia, the Kearns-Sayre syndrome (with ophthalmoplegia, pigmentary retinopathy, cardiac conduction defects, cerebellar ataxia, and sensorineural deafness), the MELAS syndrome (mitochondrial encephalomyopathy, lactic acidosis, and stroke-like episodes), the MERFF syndrome (myoclonic epilepsy and ragged red fibers), limb-girdle distribution weakness, and infantile myopathy (benign or severe and fatal).
  • sirtuin activating compounds may be useful for treating patients suffering from toxic damage to mitochondria, such as, toxic damage due to calcium accumulation, excitotoxicity, nitric oxide exposure, drug induced toxic damage, or hypoxia.
  • sirtuin activating compounds may be useful for treating diseases or disorders associated with mitochondrial deregulation.
  • the invention provides methods for enhancing muscle performance by administering a therapeutically effective amount of a sirtuin activating compound.
  • sirtuin activating compounds may be useful for improving physical endurance (e.g., ability to perform a physical task such as exercise, physical labor, sports activities, etc.), inhibiting or retarding physical fatigues, enhancing blood oxygen levels, enhancing energy in healthy individuals, enhance working capacity and endurance, reducing muscle fatigue, reducing stress, enhancing cardiac and cardiovascular function, improving sexual ability, increasing muscle ATP levels, and/or reducing lactic acid in blood.
  • the methods involve administering an amount of a sirtuin activating compound that increase mitochondrial activity, increase mitochondrial biogenesis, and/or increase mitochondrial mass.
  • Sports performance refers to the ability of the athlete's muscles to perform when participating in sports activities. Enhanced sports performance, strength, speed and endurance are measured by an increase in muscular contraction strength, increase in amplitude of muscle contraction, shortening of muscle reaction time between stimulation and contraction. Athlete refers to an individual who participates in sports at any level and who seeks to achieve an improved level of strength, speed and endurance in their performance, such as, for example, body builders, bicyclists, long distance runners, short distance runners, etc. Enhanced sports performance in manifested by the ability to overcome muscle fatigue, ability to maintain activity for longer periods of time, and have a more effective workout.
  • the methods of the present invention will also be effective in the treatment of muscle related pathological conditions, including acute sarcopenia, for example, muscle atrophy and/or cachexia associated with burns, bed rest, limb immobilization, or major thoracic, abdominal, and/or orthopedic surgery.
  • the invention provides novel dietary compositions comprising sirtuin modulators, a method for their preparation, and a method of using the compositions for improvement of sports performance. Accordingly, provided are therapeutic compositions, foods and beverages that have actions of improving physical endurance and/or inhibiting physical fatigues for those people involved in broadly-defined exercises including sports requiring endurance and labors requiring repeated muscle exertions.
  • Such dietary compositions may additional comprise electrolytes, caffeine, vitamins, carbohydrates, etc.
  • Sirtuin-modulating compounds that increase the level and/or activity of a sirtuin protein may be used for treating or preventing viral infections (such as infections by influenza, herpes or papilloma virus) or as antifungal agents.
  • sirtuin-modulating compounds that increase the level and/or activity of a sirtuin protein may be administered as part of a combination drug therapy with another therapeutic agent for the treatment of viral diseases.
  • sirtuin-modulating compounds that increase the level and/or activity of a sirtuin protein may be administered as part of a combination drug therapy with another anti-fungal agent.
  • Subjects that may be treated as described herein include eukaryotes, such as mammals, e.g., humans, ovines, bovines, equines, porcines, canines, felines, non- human primate, mice, and rats.
  • Cells that may be treated include eukaryotic cells, e.g., from a subject described above, or plant cells, yeast cells and prokaryotic cells, e.g., bacterial cells.
  • modulating compounds may be administered to farm animals to improve their ability to withstand farming conditions longer.
  • Sirtuin-modulating compounds that increase the level and/or activity of a sirtuin protein may also be used to increase lifespan, stress resistance, and resistance to apoptosis in plants.
  • a compound is applied to plants, e.g., on a periodic basis, or to fungi.
  • plants are genetically modified to produce a compound.
  • plants and fruits are treated with a compound prior to picking and shipping to increase resistance to damage during shipping. Plant seeds may also be contacted with compounds described herein, e.g., to preserve them.
  • sirtuin-modulating compounds that increase the level and/or activity of a sirtuin protein may be used for modulating lifespan in yeast cells.
  • yeast cells include any process in which yeast is used, e.g., the making of beer, yogurt, and bakery items, e.g., bread.
  • Use of yeast having an extended lifespan can result in using less yeast or in having the yeast be active for longer periods of time.
  • Yeast or other mammalian cells used for recombinantly producing proteins may also be treated as described herein.
  • Sirtuin-modulating compounds that increase the level and/or activity of a sirtuin protein may also be used to increase lifespan, stress resistance and resistance to apoptosis in insects. In this embodiment, compounds would be applied to useful insects, e.g., bees and other insects that are involved in pollination of plants.
  • a compound would be applied to bees involved in the production of honey.
  • the methods described herein may be applied to any organism, e.g., eukaryote, which may have commercial importance. For example, they can be applied to fish (aquaculture) and birds (e.g., chicken and fowl).
  • Higher doses of sirtuin-modulating compounds that increase the level and/or activity of a sirtuin protein may also be used as a pesticide by interfering with the regulation of silenced genes and the regulation of apoptosis during development.
  • a compound may be applied to plants using a method known in the art that ensures the compound is bio-available to insect larvae, and not to plants. At least in view of the link between reproduction and longevity, sirtuin- modulating compounds that increase the level and/or activity of a sirtuin protein can be applied to affect the reproduction of organisms such as insects, animals and microorganisms.
  • an agent may be a nucleic acid, such as an aptamer.
  • Assays may be conducted in a cell based or cell free format.
  • an assay may comprise incubating (or contacting) a sirtuin with a test agent under conditions in which a sirtuin can be modulated by an agent known to modulate the sirtuin, and monitoring or determining the level of modulation of the sirtuin in the presence of the test agent relative to the absence of the test agent.
  • the level of modulation of a sirtuin can be determined by determining its ability to deacetylate a substrate.
  • Exemplary substrates are acetylated peptides which can be obtained from BIOMOL (Plymouth Meeting, PA).
  • Preferred substrates include peptides of p53, such as those comprising an acetylated K382.
  • a particularly preferred substrate is the Fluor de Lys-SIRTl (BIOMOL), i.e., the acetylated peptide Arg-His-Lys-Lys.
  • Other substrates are peptides from human histones H3 and H4 or an acetylated amino acid. Substrates may be fluorogenic.
  • the sirtuin may be SIRTl, Sir2, SIRT3, or a portion thereof. For example, recombinant SIRTl can be obtained from BIOMOL.
  • the reaction may be conducted for about 30 minutes and stopped, e.g., with nicotinamide.
  • the HDAC fluorescent activity assay/drug discovery kit (AK-500, BIOMOL Research Laboratories) may be used to determine the level of acetylation. Similar assays are described in Bitterman et al. (2002) J. Biol. Chem. 277:45099.
  • the level of modulation of the sirtuin in an assay may be compared to the level of modulation of the sirtuin in the presence of one or more (separately or simultaneously) compounds described herein, which may serve as positive or negative controls.
  • Sirtuins for use in the assays may be full length sirtuin proteins or portions thereof.
  • proteins for use in the assays include N- terminal portions of sirtuins, e.g., about amino acids 1-176 or 1-255 of SIRTl; about amino acids 1-174 or 1-252 of Sir2.
  • a screening assay comprises (i) contacting a sirtuin with a test agent and an acetylated substrate under conditions appropriate for the sirtuin to deacetylate the substrate in the absence of the test agent ; and (ii) determining the level of acetylation of the substrate, wherein a lower level of acetylation of the substrate in the presence of the test agent relative to the absence of the test agent indicates that the test agent stimulates deacetylation by the sirtuin, whereas a higher level of acetylation of the substrate in the presence of the test agent relative to the absence of the test agent indicates that the test agent inhibits deacetylation by the sirtuin.
  • Methods for identifying an agent that modulates, e.g., stimulates, sirtuins in vivo may comprise (i) contacting a cell with a test agent and a substrate that is capable of entering a cell in the presence of an inhibitor of class I and class II HDACs under conditions appropriate for the sirtuin to deacetylate the substrate in the absence of the test agent ; and (ii) determining the level of acetylation of the substrate, wherein a lower level of acetylation of the substrate in the presence of the test agent relative to the absence of the test agent indicates that the test agent stimulates deacetylation by the sirtuin, whereas a higher level of acetylation of the substrate in the presence of the test agent relative to the absence of the test agent indicates that the test agent inhibits deacetylation by the sirtuin.
  • a preferred substrate is an acetylated peptide, which is also preferably fluorogenic, as further described herein.
  • the method may further comprise lysing the cells to determine the level of acetylation of the substrate.
  • Substrates may be added to cells at a concentration ranging from about l ⁇ M to about 1OmM, preferably from about lO ⁇ M to ImM, even more preferably from about lOO ⁇ M to ImM, such as about 200 ⁇ M.
  • a preferred substrate is an acetylated lysine, e.g., ⁇ -acetyl lysine (Fluor de Lys, FdL) or Fluor de Lys-SIRTl.
  • a preferred inhibitor of class I and class II HDACs is trichostatin A (TSA), which may be used at concentrations ranging from about 0.01 to lOO ⁇ M, preferably from about 0.1 to lO ⁇ M, such as l ⁇ M.
  • TSA trichostatin A
  • Incubation of cells with the test compound and the substrate may be conducted for about 10 minutes to 5 hours, preferably for about 1-3 hours. Since TSA inhibits all class I and class II HDACs, and that certain substrates, e.g., Fluor de Lys, is a poor substrate for SIRT2 and even less a substrate for SIRT3-7, such an assay may be used to identify modulators of SIRTl in vivo. 5.
  • sirtuin-modulating compounds described herein may be formulated in a conventional manner using one or more physiologically or pharmaceutically acceptable carriers or excipients.
  • sirtuin-modulating compounds and their pharmaceutically acceptable salts and solvates may be formulated for administration by, for example, injection (e.g. SubQ, EVI, IP), inhalation or insufflation (either through the mouth or the nose) or oral, buccal, sublingual, transdermal, nasal, parenteral or rectal administration.
  • a sirtuin- modulating compound may be administered locally, at the site where the target cells are present, i.e., in a specific tissue, organ, or fluid (e.g., blood, cerebrospinal fluid, etc.).
  • Sirtuin-modulating compounds can be formulated for a variety of modes of administration, including systemic and topical or localized administration. Techniques and formulations generally may be found in Remington's
  • the compounds can be formulated in liquid solutions, preferably in physiologically compatible buffers such as Hank's solution or Ringer's solution.
  • physiologically compatible buffers such as Hank's solution or Ringer's solution.
  • the compounds may be formulated in solid form and redissolved or suspended immediately prior to use. Lyophilized forms are also included.
  • the pharmaceutical compositions may take the form of, for example, tablets, lozenges, or capsules prepared by conventional means with pharmaceutically acceptable excipients such as binding agents (e.g., pregelatinized maize starch, polyvinylpyrrolidone or hydroxypropyl methylcellulose); fillers (e.g., lactose, microcrystalline cellulose or calcium hydrogen phosphate); lubricants (e.g., magnesium stearate, talc or silica); disintegrants (e.g., potato starch or sodium starch glycolate); or wetting agents (e.g., sodium lauryl sulphate).
  • binding agents e.g., pregelatinized maize starch, polyvinylpyrrolidone or hydroxypropyl methylcellulose
  • fillers e.g., lactose, microcrystalline cellulose or calcium hydrogen phosphate
  • lubricants e.g., magnesium stearate, talc or silica
  • disintegrants e.g
  • Liquid preparations for oral administration may take the form of, for example, solutions, syrups or suspensions, or they may be presented as a dry product for constitution with water or other suitable vehicle before use.
  • Such liquid preparations may be prepared by conventional means with pharmaceutically acceptable additives such as suspending agents (e.g., sorbitol syrup, cellulose derivatives or hydrogenated edible fats); emulsifying agents (e.g., lecithin or acacia); non-aqueous vehicles (e.g., almond oil, oily esters, ethyl alcohol or fractionated vegetable oils); and preservatives (e.g., methyl or propyl-p- hydroxybenzoates or sorbic acid).
  • suspending agents e.g., sorbitol syrup, cellulose derivatives or hydrogenated edible fats
  • emulsifying agents e.g., lecithin or acacia
  • non-aqueous vehicles e.g., almond oil, oily esters, eth
  • the preparations may also contain buffer salts, flavoring, coloring and sweetening agents as appropriate.
  • Preparations for oral administration may be suitably formulated to give controlled release of the active compound.
  • sirtuin- modulating compounds may be conveniently delivered in the form of an aerosol spray presentation from pressurized packs or a nebuliser, with the use of a suitable propellant, e.g., dichlorodifluoromethane, trichlorofluoromethane, dichlorotetrafluoroethane, carbon dioxide or other suitable gas.
  • a suitable propellant e.g., dichlorodifluoromethane, trichlorofluoromethane, dichlorotetrafluoroethane, carbon dioxide or other suitable gas.
  • the dosage unit may be determined by providing a valve to deliver a metered amount.
  • Capsules and cartridges of e.g., gelatin, for use in an inhaler or insufflator may be formulated
  • Sirtuin-modulating compounds may be formulated for parenteral administration by injection, e.g., by bolus injection or continuous infusion.
  • Formulations for injection may be presented in unit dosage form, e.g., in ampoules or in multi-dose containers, with an added preservative.
  • the compositions may take such forms as suspensions, solutions or emulsions in oily or aqueous vehicles, and may contain formulatory agents such as suspending, stabilizing and/or dispersing agents.
  • the active ingredient may be in powder form for constitution with a suitable vehicle, e.g., sterile pyrogen-free water, before use.
  • Sirtuin-modulating compounds may also be formulated in rectal compositions such as suppositories or retention enemas, e.g., containing conventional suppository bases such as cocoa butter or other glycerides.
  • sirtuin-modulating compounds may also be formulated as a depot preparation. Such long acting formulations may be administered by implantation (for example subcutaneously or intramuscularly) or by intramuscular injection.
  • sirtuin- modulating compounds may be formulated with suitable polymeric or hydrophobic materials (for example as an emulsion in an acceptable oil) or ion exchange resins, or as sparingly soluble derivatives, for example, as a sparingly soluble salt.
  • Controlled release formula also includes patches.
  • the compounds described herein can be formulated for delivery to the central nervous system (CNS) (reviewed in Begley, Pharmacology & Therapeutics 104: 29-45 (2004)).
  • CNS central nervous system
  • Conventional approaches for drug delivery to the CNS include: neurosurgical strategies (e.g., intracerebral injection or intracerebroventricular infusion); molecular manipulation of the agent (e.g., production of a chimeric fusion protein that comprises a transport peptide that has an affinity for an endothelial cell surface molecule in combination with an agent that is itself incapable of crossing the BBB) in an attempt to exploit one of the endogenous transport pathways of the BBB; pharmacological strategies designed to increase the lipid solubility of an agent (e.g., conjugation of water-soluble agents to lipid or cholesterol carriers); and the transitory disruption of the integrity of the BBB by hyperosmotic disruption (resulting from the infusion of a mannitol solution into the carotid artery or the use of a biologically
  • Liposomes are a further drug delivery system which is easily injectable. Accordingly, in the method of invention the active compounds can also be administered in the form of a liposome delivery system.
  • Liposomes are well-known by a person skilled in the art. Liposomes can be formed from a variety of phospholipids, such as cholesterol, stearylamine of phosphatidylcholines. Liposomes being usable for the method of invention encompass all types of liposomes including, but not limited to, small unilamellar vesicles, large unilamellar vesicles and multilamellar vesicles.
  • cyclodextrin is meant OC-, ⁇ -, or ⁇ -cyclodextrin.
  • Cyclodextrins are described in detail in Pitha et al., U.S. Pat. No. 4,727,064, which is incorporated herein by reference. Cyclodextrins are cyclic oligomers of glucose; these compounds form inclusion complexes with any drug whose molecule can fit into the lipophile- seeking cavities of the cyclodextrin molecule.
  • Rapidly disintegrating or dissolving dosage forms are useful for the rapid absorption, particularly buccal and sublingual absorption, of pharmaceutically active agents.
  • Fast melt dosage forms are beneficial to patients, such as aged and pediatric patients, who have difficulty in swallowing typical solid dosage forms, such as caplets and tablets. Additionally, fast melt dosage forms circumvent drawbacks associated with, for example, chewable dosage forms, wherein the length of time an active agent remains in a patient's mouth plays an important role in determining the amount of taste masking and the extent to which a patient may object to throat grittiness of the active agent.
  • compositions may comprise from about 0.00001 to 100% such as from 0.001 to 10% or from 0.1% to 5% by weight of one or more sirtuin-modulating compounds described herein.
  • the pharmaceutical composition comprises: (i) 0.05 to 1000 mg of the compounds of the invention, or a pharmaceutically acceptable salt thereof, and (ii) 0.1 to 2 grams of one or more pharmaceutically acceptable excipients.
  • a sirtuin-modulating compound described herein is incorporated into a topical formulation containing a topical carrier that is generally suited to topical drug administration and comprising any such material known in the art.
  • the topical carrier may be selected so as to provide the composition in the desired form, e.g., as an ointment, lotion, cream, microemulsion, gel, oil, solution, or the like, and may be comprised of a material of either naturally occurring or synthetic origin. It is preferable that the selected carrier not adversely affect the active agent or other components of the topical formulation.
  • suitable topical carriers for use herein include water, alcohols and other nontoxic organic solvents, glycerin, mineral oil, silicone, petroleum jelly, lanolin, fatty acids, vegetable oils, parabens, waxes, and the like.
  • Formulations may be colorless, odorless ointments, lotions, creams, microemulsions and gels.
  • Sirtuin-modulating compounds may be incorporated into ointments, which generally are semisolid preparations which are typically based on petrolatum or other petroleum derivatives.
  • ointments which generally are semisolid preparations which are typically based on petrolatum or other petroleum derivatives.
  • the specific ointment base to be used is one that will provide for optimum drug delivery, and, preferably, will provide for other desired characteristics as well, e.g., emolliency or the like.
  • an ointment base should be inert, stable, nonirritating and nonsensitizing.
  • Sirtuin-modulating compounds may be incorporated into lotions, which generally are preparations to be applied to the skin surface without friction, and are typically liquid or semiliquid preparations in which solid particles, including the active agent, are present in a water or alcohol base.
  • Lotions are usually suspensions of solids, and may comprise a liquid oily emulsion of the oil-in-water type.
  • Sirtuin-modulating compounds may be incorporated into creams, which generally are viscous liquid or semisolid emulsions, either oil-in-water or water- in- oil.
  • Cream bases are water- washable, and contain an oil phase, an emulsifier and an aqueous phase.
  • the oil phase is generally comprised of petrolatum and a fatty alcohol such as cetyl or stearyl alcohol; the aqueous phase usually, although not necessarily, exceeds the oil phase in volume, and generally contains a humectant.
  • the emulsifier in a cream formulation as explained in Remington 's, supra, is generally a nonionic, anionic, cationic or amphoteric surfactant.
  • Sirtuin-modulating compounds may be incorporated into microemulsions, which generally are thermodynamic ally stable, isotropically clear dispersions of two immiscible liquids, such as oil and water, stabilized by an interfacial film of surfactant molecules (Encyclopedia of Pharmaceutical Technology (New York: Marcel Dekker, 1992), volume 9).
  • Sirtuin-modulating compounds may be incorporated into gel formulations, which generally are semisolid systems consisting of either suspensions made up of small inorganic particles (two-phase systems) or large organic molecules distributed substantially uniformly throughout a carrier liquid (single phase gels). Although gels commonly employ aqueous carrier liquid, alcohols and oils can be used as the carrier liquid as well.
  • sunscreen formulations e.g., other antiinflammatory agents, analgesics, antimicrobial agents, antifungal agents, antibiotics, vitamins, antioxidants, and sunblock agents commonly found in sunscreen formulations including, but not limited to, anthranilates, benzophenones (particularly benzophenone-3), camphor derivatives, cinnamates (e.g., octyl methoxycinnamate), dibenzoyl methanes (e.g., butyl methoxydibenzoyl methane), p-aminobenzoic acid (PABA) and derivatives thereof, and salicylates (e.g., octyl salicylate).
  • sunscreen formulations including, but not limited to, anthranilates, benzophenones (particularly benzophenone-3), camphor derivatives, cinnamates (e.g., octyl methoxycinnamate), dibenzoyl methanes (e.g., but
  • the active agent is present in an amount in the range of approximately 0.25 wt. % to 75 wt. % of the formulation, preferably in the range of approximately 0.25 wt. % to 30 wt. % of the formulation, more preferably in the range of approximately 0.5 wt. % to 15 wt. % of the formulation, and most preferably in the range of approximately 1.0 wt. % to 10 wt. % of the formulation.
  • Conditions of the eye can be treated or prevented by, e.g., systemic, topical, intraocular injection of a sirtuin-modulating compound, or by insertion of a sustained release device that releases a sirtuin-modulating compound.
  • a sirtuin- modulating compound that increases the level and/or activity of a sirtuin protein may be delivered in a pharmaceutically acceptable ophthalmic vehicle, such that the compound is maintained in contact with the ocular surface for a sufficient time period to allow the compound to penetrate the corneal and internal regions of the eye, as for example the anterior chamber, posterior chamber, vitreous body, aqueous humor, vitreous humor, cornea, iris/ciliary, lens, choroid/retina and sclera.
  • the pharmaceutically-acceptable ophthalmic vehicle may, for example, be an ointment, vegetable oil or an encapsulating material.
  • the compounds of the invention may be injected directly into the vitreous and aqueous humour.
  • the compounds may be administered systemically, such as by intravenous infusion or injection, for treatment of the eye.
  • Sirtuin-modulating compounds described herein may be stored in oxygen free environment.
  • resveratrol or analog thereof can be prepared in an airtight capsule for oral administration, such as Capsugel from Pfizer, Inc.
  • Cells e.g., treated ex vivo with a sirtuin-modulating compound
  • an immunosuppressant drug e.g., cyclosporin A.
  • the reader is referred to Cell Therapy: Stem Cell Transplantation, Gene Therapy, and Cellular Immunotherapy, by G. Morstyn & W. Sheridan eds, Cambridge University Press, 1996; and Hematopoietic Stem Cell Therapy, E. D. Ball, J. Lister & P. Law, Churchill Livingstone, 2000.
  • Toxicity and therapeutic efficacy of sirtuin-modulating compounds can be determined by standard pharmaceutical procedures in cell cultures or experimental animals.
  • the LDso is the dose lethal to 50% of the population.
  • the EDso is the dose therapeutically effective in 50% of the population.
  • the dose ratio between toxic and therapeutic effects (LDso/EDso) is the therapeutic index.
  • Sirtuin-modulating compounds that exhibit large therapeutic indexes are preferred. While sirtuin- modulating compounds that exhibit toxic side effects may be used, care should be taken to design a delivery system that targets such compounds to the site of affected tissue in order to minimize potential damage to uninfected cells and, thereby, reduce side effects.
  • the data obtained from the cell culture assays and animal studies can be used in formulating a range of dosage for use in humans.
  • the dosage of such compounds may lie within a range of circulating concentrations that include the EDso with little or no toxicity.
  • the dosage may vary within this range depending upon the dosage form employed and the route of administration utilized.
  • the therapeutically effective dose can be estimated initially from cell culture assays.
  • a dose may be formulated in animal models to achieve a circulating plasma concentration range that includes the ICso (i.e., the concentration of the test compound that achieves a half-maximal inhibition of symptoms) as determined in cell culture.
  • ICso i.e., the concentration of the test compound that achieves a half-maximal inhibition of symptoms
  • levels in plasma may be measured, for example, by high performance liquid chromatography. 6. Kits
  • kits e.g., kits for therapeutic purposes or kits for modulating the lifespan of cells or modulating apoptosis.
  • a kit may comprise one or more sirtuin-modulating compounds, e.g., in premeasured doses.
  • a kit may optionally comprise devices for contacting cells with the compounds and instructions for use. Devices include syringes, stents and other devices for introducing a sirtuin-modulating compound into a subject (e.g., the blood vessel of a subject) or applying it to the skin of a subject.
  • the invention provides a composition of matter comprising a sirtuin modulator of this invention and another therapeutic agent (the same ones used in combination therapies and combination compositions) in separate dosage forms, but associated with one another.
  • a sirtuin modulator of this invention and another therapeutic agent (the same ones used in combination therapies and combination compositions) in separate dosage forms, but associated with one another.
  • the term "associated with one another" as used herein means that the separate dosage forms are packaged together or otherwise attached to one another such that it is readily apparent that the separate dosage forms are intended to be sold and administered as part of the same regimen.
  • the agent and the sirtuin modulator are preferably packaged together in a blister pack or other multi-chamber package, or as connected, separately sealed containers (such as foil pouches or the like) that can be separated by the user (e.g., by tearing on score lines between the two containers).
  • the invention provides a kit comprising in separate vessels, a) a sirtuin modulator of this invention; and b) another
  • Tris(dibenzylideneacetone)dipalladium(0) (18mg, 0.02 mmol) was added, and nitrogen was bubbled through the solution for 5 min. The tube was then sealed and the reaction was heated in the microwave with stirring for 1.5 h at 120 0 C. Water was added (40 mL) and enough 5N HCl was added to bring the pH to 4. The mixture was extracted with ethyl acetate (3 x 25mL) and the organics were washed with brine, dried with sodium sulfate, filtered and concentrated.
  • ⁇ -Chloroquinoline ⁇ -carboxylic acid (1; 100 mg, 0.48 mmol), l-(3-(4,4,5,5- tetramethyl-l,3,2-dioxaborolan-2-yl)phenyl)pyrrolidine (5; 197 mg, 0.72 mmol), dicyclohexyl(2',6'-dimethoxybiphenyl-2-yl)phosphine (16 mg, 0.04 mmol) and K 3 PO 4 (307mg, 1.44 mmol) were suspended in dioxane (2 mL) and water (0.2 mL).
  • Tris(dibenzylideneacetone)dipalladium(0) (18 mg, 0.02 mmol) was added, and nitrogen was bubbled through the solution for 5 min. The tube was then sealed and the reaction was heated in the microwave with stirring for 1.5 h at 120 0 C. Water was added (40 rnL) and enough 5N HCl was added to bring the pH to 4. The mixture was extracted with ethyl acetate (3x25mL) and the organics were washed with brine, dried with sodium sulfate, filtered and concentrated.
  • Tris(dibenzylideneacetone)dipalladium(0) (18 mg, 0.02 mmol) was added, and nitrogen was bubbled through the solution for 5 min. The tube was then sealed and the reaction was heated in the microwave with stirring for 2 h at 120 0 C. Water was added (40 mL) and enough 5N HCl was added to bring the p ⁇ to 4. The mixture was extracted with ethyl acetate (3 x 30 mL) and the organics were washed with brine, dried with sodium sulfate, filtered and concentrated.
  • Methyl- ⁇ -Q ⁇ difluoromethy ⁇ pheny ⁇ quinoline ⁇ -carboxylate 32; 1.36 g, 4.34 mmol
  • LiOH H 2 O 730 mg, 17.36 mmol
  • THF/H 2 O (2:1, 20 ml) was stirred at room temp, for 2 h.
  • the reaction mixture was then concentrated to remove the THF, more water (5 mL) was added and the pH was adjusted to 5.
  • 4-Bromo-2-(pyrrolidin-l-ylmethyl)pyridine 44 was prepared following a similar procedure as that of 2-bromo-5-(pyrrolidin-l-ylmethyl)thiazole 38 above using 4-bromopicolinaldehyde (1.35 g, 30%). MS calcd for Ci 0 Hi 3 BrN2: 240; found: 241 [M+l].
  • Azabenzotriazol-l-yl)- ⁇ W ⁇ f', ⁇ f'-tetramethyluronium hexafluorophosphate 122 mg, 0.32 mmol
  • diisopropylethylamine 62 mg, 0.48 mmol
  • N,N- dimethylformamide 3 mL
  • Tris(dibenzylideneacetone)dipalladium(0) 29 mg, 0.03 mmol was added, and the mixture was sparged with nitrogen another 2 minutes, then sealed and heated in the microwave to 125 0 C for 1.5 h.
  • Water (20 mL) and ethyl acetate (20 mL) were added, and the layers separated.
  • the aqueous layer was adjusted to pH 4 by addition of drops of 5 N HCl, after which time a precipitate formed.
  • the mixture was filtered, and the solid was washed with water, ethyl acetate and ether.
  • This general procedure is used to produce other compounds of the invention containing 8-(3-(trifluoromethyl)phenyl)-l,6-naphthyridine-2-carboxamide by using the appropriate amine component in place of aminopyrazine.
  • Ethyl 2-aminothiazole-4-carboxylate (59; 10.0 g, 58.1 mmol) was taken up in 150 mL of anhydrous THF along with di-tert-butyl carbonate (BoC 2 O, 12.67 g, 58.1 mmol) along with 10 mg of 4-(dimethyl)aminopyridine (DMAP).
  • DMAP 4-(dimethyl)aminopyridine
  • the reaction mixture was stirred at 50 0 C for 4 h and then at room temperature for 18 h. It was then concentrated under reduced pressure to obtain a thick oil. Pentane was added and the resulting crystalline materials were collected by filtration and dried to afford 10.5 g of ethyl 2-(tert-butoxycarbonylamino)thiazole-4-carboxylate 60.
  • This material (60; 10.5 g, 38.5 mmol) was dissolved in 300 mL of anhydrous THF and cooled in Dry Ice-acetonitrile bath. A solution of 1 M Super HydrideTM in THF (85 mL) was then added over a period of 10 min. The resulting reaction mixture was stirred at -45°C for 2 h. Another portion of 1 M Super HydrideTM in THF (35 mL) was then added and the reaction mixture was stirred for an additional 2 h at -45°C. The reaction was quenched at -45°C by the addition of 50 mL of brine. Upon warming to room temperature, the reaction mixture was concentrated under reduced pressure. The resulting mixture was extracted with EtOAc.
  • Step 2) Synthesis of4-(morpholinomethyl)thiazol-2-amine (63): tert-Butyl 4-(hydroxymethyl)thiazol-2-ylcarbamate (61; 2.0 g, 8.7 mmol) was taken up in 25 niL of CH 2 Cl 2 along with Et 3 N (1.82 niL, 13.05 mmol) and cooled to 0 0 C. Methanesulfonyl chloride (0.85 mL, 10.88 mmol) was added and the resulting reaction mixture was stirred at 0 0 C for 60 min. Morpholine (3.0 mL, 35 mmol) was then added and the reaction mixture was stirred at room temperature for 18 h. The reaction mixture was concentrated under reduced pressure.
  • Ethyl 2-aminothiazole-5-carboxylate (71; 10.0 g, 58.1 mmol) was taken up in 150 mL of anhydrous THF along with di-tert-butyl carbonate (12.67 g, 58.1 mmol) along with 10 mg of 4-(dimethyl)aminopyridine. The reaction mixture was stirred at 5O 0 C for 4 h and then at room temperature for 18 h. It was then concentrated under reduced pressure to obtain a thick oil. Pentane was added and the resulting crystalline materials were collected by filtration and dried to afford 10.5 g of ethyl 2- (tert-butoxycarbonylamino)thiazole-5-carboxylate 72.
  • 2-Chloropyridin-4-amine 78 was subjected to the same reaction conditions described above for the preparation of 6-morpholinopyridin-2-amine. Pyrrolidine was used as the amine component instead of morpholine.
  • Example 21 Preparation of N-methyl-./V-(4-(pyrrolidin-l-ylmethyl)phenyl)-8- (3-(trifluoromethyl) phenylquinoline-2-carboxamide (Compound 265): Step 1) Synthesis of l-methyl-l-(4-(N-methyl-8-(3-(trifluoromethyl)phenyl) quinoline-2-carboxamido)benzyl)pyrrolidinium (80):
  • N-(4-(pyrrolidin-l-ylmethyl)phenyl)-8-(3-(trifluoromethyl)phenyl)quinoline- 2-carboxamide (Compound 201; 50 mg, 0.1 mmol), was dissolved in toluene (3.3 mL) and cooled to O 0 C under nitrogen atmosphere. Potassium hexamethyldisilazide (0.5 M, 1.05 mL, 0.53 mmol) was added and the reaction was allowed to warm to room temperature for 15 min, followed by re-cooling to O 0 C. Iodomethane (0.03 niL, 0.53 mmol) was added, and after 5 min. the reaction was again warmed to room temperature.
  • the crude mixture from above (0.1 mmol) was dissolved in neat pyrrolidine (0.8 mL, 10 mmol). The mixture was stirred at 7O 0 C overnight, then sealed and heated in the microwave to HO 0 C for 1 h. Water was added (20 mL), and the mixture was extracted with dichloromethane (5 x 10 mL). The combined organics were washed with brine, dried with sodium sulfate, filtered and concentrated. The crude material was purified by prep. HPLC using 15-85% acetonitrile/water with 0.1% trifluoroacetic acid.
  • solketal (10; 34.4 g, 260 mmol) in THF (150 niL) was added NaH (10.4 g, 260 mmol) at room temperature and the mixture stirred for Ih.
  • 2- chloro-4-aminopyrimidine (82; 15.0 g, 115 mmol) was then added, and the mixture was stirred at 7O 0 C for 48 h.
  • 2-methoxynicotinamide 87 is prepared in a manner similar to the procedures in Bioorg. Med. Chem. Lett. 2007, 17, 2031 and /. Org. Chem. 1975, 40, 3554.
  • 2-methoxynicotinic acid 86; 49 mmol
  • suspended dichloromethane 60 mL
  • DMF 3 drops
  • oxalyl chloride 16.6 mL, 186 mmol
  • the mixture is stirred at room temperature for 1 h, then concentrated and redissolved in petroleum ether (60 mL). This mixture is then added at -35 0 C to 250 mL of anhydrous acetonitrile that has been saturated with ammonia at -30 0 C.
  • Tert-butyl 2-methoxypyridin-3-ylcarbamate 88 is prepared in a manner similar to the procedure in /. Med. Chem. 1988, 31, 2136. To a suspension of 2- methoxynicotinamide (87; 10 mmol) in anhydrous tert-butyl alcohol (25 mL) is added lead tetraacetate (4.44 g, 10 mmol) under nitrogen atmosphere. The reaction mixture is heated to reflux for 2 h, then cooled and filtered through diatomaceous earth. The filtrate is concentrated and the residue dissolved in diethyl ether. The solution is washed with saturated aqueous sodium bicarbonate and brine, then dried with sodium sulfate, filtered, and concentrated to give the desired product 88 which can be purified by recrystallization.
  • Tert-butyl 4-fo ⁇ nyl-2-methoxypyridin-3-ylcarbamate 89 is prepared in a manner similar to the procedure in /. Med. Chem. 1988, 31, 2136.
  • a solution of 2- methoxypyridin-3-ylcarbamate (88; 100 mmol) in dry THF (350 mL) is cooled to - 78 0 C, followed by drop wise addition of tert-butyllithium (120 mL, 2 M in pentane, 240 mmol) at a rate such that the temperature does not exceed -65 0 C.
  • the reaction is stirred at -78 0 C for an additional 15 min., then at -20 0 C for 1.5 h.
  • 8-methoxy-l, 7-naphthyridine-2-carboxylic acid 90 is prepared in a manner similar to the procedure in Tetrahedron Lett. 2000, 41, 8053.
  • the Boc group of yert-butyl 4-formyl-2-methoxypyridin-3-ylcarbamate 89 is removed using trifluoroacetic acid in dichloromethane, followed by an aldol condensation with sodium pyruvate in sodium hydroxide/water to produce the desired product 90.
  • Step 5 Preparation of8-bromo-l,7-naphthyridine-2-carboxylic acid (91): ⁇ -bromo-lj-naphthyridine ⁇ -carboxyric acid 91 is prepared in a manner similar to the procedure in Bioorg. Med. Chem. Lett. 2002, 12, 233. 8-methoxy-l,7- naphthyridine-2-carboxylic acid 90 is dissolved in DMF and treated with PBr 3 at 100 0 C for 30 min to produce the desired product 91.
  • ⁇ -bromo-lj-naphthyridine-l-carboxylic acid 91 is derivatized with the appropriate R 2 group at the 8-position to produce a carboxylic acid intermediate that may be combined with an appropriate amine to produce compounds of the invention containing in a 8-substituted-l,7-naphthyridine-2-carboxamide.
  • Example 25 Biological activity
  • a mass spectrometry based assay was used to identify modulators of SIRTl activity.
  • the mass spectrometry based assay utilizes a peptide having 20 amino acid residues as follows: Ac-EE-K(biotin)-GQSTSSHSK(Ac)NleSTEG-K(5TMR)-EE- NH 2 (SEQ ID NO: 1) wherein K(Ac) is an acetylated lysine residue and NIe is a norleucine.
  • the peptide is labeled with the fluorophore 5TMR (excitation 540 nm/emission 580 nm) at the C-terminus.
  • the sequence of the peptide substrate is based on p53 with several modifications.
  • the mass spectrometry assay is conducted as follows: 0.5 ⁇ M peptide substrate and 120 ⁇ M ⁇ NAD + is incubated with 10 nM SIRTl for 25 minutes at 25°C in a reaction buffer (50 mM Tris-acetate pH 8, 137 mM NaCl, 2.7 mM KCl, 1 mM MgCl 2 , 5 mM DTT, 0.05% BSA). Test compounds may be added to the reaction as described above.
  • the SirTl gene is cloned into a T7 -promoter containing vector and transformed into BL21(DE3). After the 25 minute incubation with
  • SIRTl protein was expressed and purified as follows.
  • the SirTl gene was cloned into a T7 -promoter containing vector and transformed into BL21(DE3).
  • the protein was expressed by induction with 1 mM IPTG as an N- terminal His-tag fusion protein at 18°C overnight and harvested at 30,000 x g.
  • Cells were lysed with lysozyme in lysis buffer (50 mM Tris-HCl, 2 mM Tris[2- carboxyethyl] phosphine (TCEP), 10 ⁇ M ZnCl 2 , 200 mM NaCl) and further treated with sonication for 10 min for complete lysis.
  • lysis buffer 50 mM Tris-HCl, 2 mM Tris[2- carboxyethyl] phosphine (TCEP), 10 ⁇ M ZnCl 2 , 200 mM NaCl
  • the protein was purified over a Ni-NTA column (Amersham) and fractions containing pure protein were pooled, concentrated and run over a sizing column (Sephadex S200 26/60 global). The peak containing soluble protein was collected and run on an Ion-exchange column (MonoQ). Gradient elution (200 mM - 500 mM NaCl) yielded pure protein. This protein was concentrated and dialyzed against dialysis buffer (20 mM Tris-HCl, 2 mM TCEP) overnight. The protein was aliquoted and frozen at -80°C until further use.
  • Sirtuin modulating compounds that activated SIRTl were identified using the assay described above and are shown below in Table 1.
  • the ECi 5 values represent the concentration of test compounds that result in 150% activation of SIRTl .
  • the ECi 5 values for the activating compounds are represented A (ECi 5 ⁇ 1 uM), B (ECi 5 >1 and ⁇ 15 uM), or C (ECi 5 >15 uM).
  • the percent maximum fold activation is represented by A (Fold activation >350%), B (Fold Activation >150% and ⁇ 350%), or C (Fold Activation ⁇ 150%).
  • Example 26 Cell based Assay
  • TNF- ⁇ Tumor necrosis factor
  • SNF- ⁇ Tumor necrosis factor
  • the primary role of TNF- ⁇ is in the regulation of immune cells. TNF- ⁇ is also able to induce apoptotic cell death and to induce inflammation.
  • Dysregulation of TNF- ⁇ production has been implicated in a variety of human diseases, as well as cancer.
  • Sirtuin modulators identified by their ability to activate SIRTl in the in vitro assay described above were tested for their ability to inhibit LPS-induced TNF- ⁇ production in macrophages.
  • Raw Macrophage 264.7 cells were obtained from the American Tissue Culture Collection (ATCC Deposit No. TIB-71). Approximately 4 x 10 4 cells were seeded into microtiter plate wells and incubated for 17-18 hours at 37°C and 5.0% CO 2 prior to each assay.
  • the Raw264.7 Macrophage Growth Culture Media (Assay Media) consists of DMEM Media supplemented with 10% low endotoxin FBS (fetal bovine serum), 100 units/ml penicillin, and 100 ⁇ g/ml streptomycin. Media was stored at 4°C and warmed to 37°C prior to use.
  • IC 50 value i.e., the concentration of compound required to inhibit the LPS-induced production of TNF- ⁇ by 50%.
  • the SIRTl activator was added to the Raw Macrophage 264.7 test cells and incubated for 2 hours at 37°C and 5.0% CO 2 prior to the addition of LPS.
  • a TACE inhibitor compound was added to a control sample at a concentration that completely inhibits TNF- ⁇ secretion (100% inhibition control).
  • the solvent DMSO was added to another control sample (0% inhibition control).
  • LPS Lipopolysaccharide
  • TNF- ⁇ is secreted from mouse RAW 264.7 macrophages (a mouse leukaemic monocyte macrophage cell line) when the cells are exposed to Lipopolysaccharide (LPS).
  • LPS Lipopolysaccharide
  • ELISA Enzyme Linked- Immunosorbent Assay
  • the colorimetric read-out was measured by the Spectramax M5 Plate Reader.
  • the intensity of the produced absorbance is directly proportional to the concentration of MsTNF- ⁇ present in the original specimen.
  • the amount of Ms-TNF- ⁇ secreted in the samples can thus be interpolated and calculated from a standard curve using recombinant mouse TNF- ⁇ , and the percent inhibition is then calculated based on the TACE and DMSO control wells.
  • the MsTNF- ⁇ Assay measures the amount of MsTNF- ⁇ that is secreted by cells, in the presence of LPS and a known SIRTl activator. Using this assay one can assess the ability of a test compound to inhibit MsTNF- ⁇ secretion in a dose- dependent manner. An IC 50 value was determined for each compound tested.
  • any polynucleotide and polypeptide sequences which reference an accession number correlating to an entry in a public database, such as those maintained by The Institute for Genomic Research (TIGR) (www.tigr.org) and/or the National Center for Biotechnology Information (NCBI) (www.ncbi.nlm.nih.gov).
  • TIGR The Institute for Genomic Research
  • NCBI National Center for Biotechnology Information

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Abstract

Cette invention concerne des quinoléines substituées en 8 et analogues associés utilisés comme composés modulant la sirtuine de formule structurelle (I) et leurs procédés d'utilisation. Les modulateurs de sirtuine peuvent être utilisés pour augmenter la durée de vie d'une cellule, et traiter et/ou prévenir un large éventail de maladies et de troubles, notamment les maladies et les troubles associés au vieillissement ou au stress, au diabète, à l'obésité, aux maladies neurodégénératives, aux maladies cardiovasculaires, aux troubles de la coagulation, à l'inflammation, cancer et/ou aux bouffées congestives ainsi qu'aux maladies et aux troubles bénéficiant d'un accroissement de l'activité mitochondriale. L'invention concerne également des compositions contenant un composé modulateur de sirtuine en association avec un autre agent thérapeutique.
PCT/US2010/025963 2009-03-02 2010-03-02 Quinoléines substituées en 8 et analogues associés utilisés comme modulateurs de sirtuine WO2010101949A1 (fr)

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BRPI1011477A BRPI1011477A2 (pt) 2009-03-02 2010-03-02 quinolinas 8-substituidas e analogos relacionados como moduladores de sirtuina
EP10749227A EP2403833A4 (fr) 2009-03-02 2010-03-02 Quinoléines substituées en 8 et analogues associés utilisés comme modulateurs de sirtuine
CN2010800194505A CN102414180A (zh) 2009-03-02 2010-03-02 作为沉默调节蛋白调节剂的8-取代的喹啉及相关的类似物
AU2010221417A AU2010221417A1 (en) 2009-03-02 2010-03-02 8-substituted quinolines and related analogs as sirtuin modulators
CA2754058A CA2754058A1 (fr) 2009-03-02 2010-03-02 Quinoleines substituees en 8 et analogues associes utilises comme modulateurs de sirtuine
JP2011553053A JP2012519211A (ja) 2009-03-02 2010-03-02 サーチュイン調節化合物としての８−置換キノリンおよび関連類似体
MX2011009213A MX2011009213A (es) 2009-03-02 2010-03-02 Quinolinas 8-sustituidas y analogos relacionados como moduladores de sirtuina.
EA201171098A EA201171098A1 (ru) 2009-03-02 2010-03-02 8-замещенные хинолины и родственные аналоги в качестве модуляторов сиртуина
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AU2010221417A1 (en) 2011-09-22
EP2403833A1 (fr) 2012-01-11
BRPI1011477A2 (pt) 2016-03-22
EP2403833A4 (fr) 2012-08-29
IL214943A0 (en) 2011-11-30
JP2012519211A (ja) 2012-08-23
MX2011009213A (es) 2011-12-14
CN102414180A (zh) 2012-04-11
KR20110128908A (ko) 2011-11-30
EA201171098A1 (ru) 2012-04-30

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