NZ729411B2 - INHIBITORS OF a-AMINO-ß-CARBOXYMUCONIC ACID SEMIALDEHYDE DECARBOXYLASE - Google Patents

INHIBITORS OF a-AMINO-ß-CARBOXYMUCONIC ACID SEMIALDEHYDE DECARBOXYLASE Download PDF

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NZ729411B2
NZ729411B2 NZ729411A NZ72941115A NZ729411B2 NZ 729411 B2 NZ729411 B2 NZ 729411B2 NZ 729411 A NZ729411 A NZ 729411A NZ 72941115 A NZ72941115 A NZ 72941115A NZ 729411 B2 NZ729411 B2 NZ 729411B2
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formula
compound
disease
disorder
pharmaceutically acceptable
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NZ729411A
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NZ729411A (en
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Johan Auwerx
Roberto Pellicciari
Nadia Raffaelli
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Tes Pharma Srl
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Priority claimed from PCT/EP2015/069808 external-priority patent/WO2016030534A1/en
Publication of NZ729411A publication Critical patent/NZ729411A/en
Publication of NZ729411B2 publication Critical patent/NZ729411B2/en

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    • A61K31/495—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with two or more nitrogen atoms as the only ring heteroatoms, e.g. piperazine or tetrazines
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    • A61K31/505—Pyrimidines; Hydrogenated pyrimidines, e.g. trimethoprim
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Abstract

The present disclosure discloses compounds capable of modulating the activity of a-amino-ß-carboxymuconic acid semialdehyde decarboxylase (ACMSD), which are useful for the prevention and/or the treatment of diseases and disorders associated with defects in NAD+ biosynthesis, e.g., metabolic disorders, neurodegenerative diseases, chronic inflammatory diseases, kidney diseases, and diseases associated with ageing. The present application also discloses pharmaceutical compositions comprising said compounds and the use of such compounds as a medicament.

Description

[Annotation] KEB INHIBITORS OF OL—AMINO—B—CARBOXYMUCONIC ACID SEMIALDEHYDE DECARBOXYLASE RELATED APPLICATIONS This application claims the bene?t of and priority to US. provisional application No. ,853, filed August 29, 2014, the entire contents of which are incorporated herein by reference in its entirety.
FIELD OF THE DISCLOSURE The present disclosure s to compounds capable of modulating the activity of Ot— amino—B-carboxymuconic acid semialdehyde decarboxylase (ACMSD). The compounds of the disclosure may be used in methods for the prevention and/or the treatment of diseases and ers associated with s in NAD+ biosynthesis, e.g., metabolic ers, neurodegenerative diseases, chronic in?ammatory diseases, kidney diseases, and diseases associated with ageing.
BACKGROUND OF THE DISCLOSURE ACMSD is a critical enzyme for tryptophan metabolism, and regulates NAD+ biosynthesis from tryptophan. ACMSD is a zinc-dependent amidohydrolase that participates in picolinic acid (PA), quinolinic acid (QA) and NAD homeostasis. ACMSD stands at a branch point of the NAD+ biosynthetic pathway from tryptophan and determines the ?nal fate of the amino acid, i.e., ormation into PA, complete ion through the citric acid cycle, or conversion into NAD+ through QA synthesis.
ACMSD has been puri?ed from liver, kidney, and brain human tissues. There are two isoforms ACMSD] and ACMSD2 derived from a differential ng ofACMSD gene transcription but only ACMSDl is endowed with enzymatic activity. ACMSDl directs ACMS (OL—amino-w—carboxymuconic acid semialdehyde) to the acetyl-CoA y, and when ACMSDl is ted, ACMS is zymatically converted to quinolinic acid (QA) leading to the formation ofNAD+ and an increase in the intracellular level of NADI.
Increased levels ofNAD+ have been shown to protect against neuronal degeneration, improve muscle function and oxidative metabolism in mice, and enhance lifespan in worms.
Whilst reduced levels ofNAD+ have been associated with a range of pathophysiological [Annotation] KEB states including type 2 diabetes (T2D), hyperlipidemia (elevated cholesterol and TAGS), mitochondrial diseases, neutropenia, cancers, and kidney disorders.
The inhibition of ACMSD thus represents a novel approach to increase NAD+ levels and modify disease pathophysiologies ated with defects in NAD+ biosynthesis.
Y OF THE DISCLOSURE It is an object of embodiments of the disclosure to provide novel series of compounds capable of modulating the ty of oc-amino-B-carboxymuconic acid semialdehyde decarboxylase (ACMSD), which compounds are useful for the prevention and/or the treatment of diseases and disorders associated with defects in NAD+ biosynthesis, e.g., metabolic disorders, neurodegenerative diseases, c in?ammatory diseases, kidney diseases, and diseases associated with ageing.
Compounds of a (I), as d herein, may be used in the treatment of a e or disorder in which ACMSD plays a role. The disclosure es methods of ng a e or disorder associated with ACMSD ction or with abnormalities in NAD+ biosynthesis by administering to subjects suffering from or susceptible to ping a disease or disorder associated with ACMSD dysfunction a therapeutically effective amount of one or more compounds that increases intracellular NAD+ by ACMSDI inhibition, in an amount sufficient to activate sirtuins (SIRTs) and the downstream targets of SIRTs, such as PGC—loc, FoxOl and/or superoxide dismutase (SOD). The methods of the present disclosure can be used in the treatment ofACMSD dependent diseases by inhibiting ACMSD.
Inhibition of ACMSD may provide a novel approach to the prevention and treatment of metabolic disorders, neurodegenerative diseases, chronic in?ammatory diseases, kidney diseases, diseases associated with ageing and other ACMSD dependent diseases, or diseases characterized by defective NAD+ synthesis.
Accordingly, a ?rst aspect of the present disclosure relates to a compound ented by Formula (I): or a pharmaceutically acceptable salt or tautomer thereof, ation] KEB wherein: X is O, OH, or Cl; L is mCH2CH2-, _(CH2)mY(CH2)p'2 "(CH2)mC(O)(CH2)p'a —(CH2)mC(O)O(CH2)p—, mC(O)NR2(CH2)p-, or mNR2C(O)(CH2)p; Y is O, N or S(O)q; R1 is C6-C10 aryl or heteroaryl, wherein the aryl and heteroaryl are substituted with Ra and Rb, and optionally substituted with one or more Re; R2 is H or C1-C6 alkyl; one of R3 and Rb is hydrogen and the other is —(CH2)rCOZRX, 02RX, -(CH2)rtetrazole, —(CH2)roxadiazolone, -(CH2)rtetrazolone, -(CH2)rthiadiazolol, —(CH2)r isoxazol—3 —ol, —(CH2)IP(O)(OH)ORX, —(CH2)rS(O)ZOH, —(CH2)IC(O)NHCN, or —(CH2)rC(O)NHS(O)2alkyl; Rc is H, C1-C6 alkyl, C1—C6 haloalkyl, halogen, —CN, —ORx, -COsz, or N02; Rd is methyl, optionally tuted 5— to lO-membered aryl, optionally substituted 5— or 6-membered heteroaryl, or optionally substituted 5- or 6-membered carbocycle; each RX is independently at each occurrence hydrogen or C1—C6 alkyl; each R6 is independently C1-C6 alkyl, C2—C6 alkenyl, C2-C6 alkynyl, halogen, -ORy, C1—C6 haloalkyl, -NHRZ, —OH, or —CN; Rf is H or absent; each Ry and RZ is independently hydrogen, C1-C6 alkyl, or C1-C6 kyl; each m and p independently is 0, l or 2, wherein m + p < 3; q is 0, 1, or 2; r is 0 or 1; and the dotted line is an optional double bond; with the proviso that Rc is not hydrogen or —CN when X is O, L is —SCH2- and Rd is optionally substituted phenyl, Rc is not C1-C6 alkyl when X is O, L is —SCH2— and Rd is methyl, and that Rc is not —CN when X is O, L is —SCH2— and Rd is 2-furyl.
A second aspect of the present disclosure relates to pharmaceutical compositions comprising a nd of Formula (I), or a pharmaceutically acceptable salt thereof, and at least one of a pharmaceutically acceptable carrier, diluent, or excipient.
A third aspect of the disclosure relates to a method of treating a disease or disorder associated with oc—amino—B—carboxymuconate-s—semialdehyde decarboxylase (ACMSD) dysfunction sing administering to the subject suffering from or susceptible to [Annotation] KEB developing a disease or disorder ated with ACMSD dysfunction a therapeutically effective amount of one or more compounds of Formula (I).
A fourth aspect of the disclosure relates to a method of preventing a disease or disorder associated with 0t—amino—B—carboxymuconate—a—semialdehyde decarboxylase (ACMSD) dysfunction comprising administering to the subject suffering from or tible to developing a disease or disorder associated with ACMSD dysfunction a therapeutically effective amount of one or more compounds of Formula (I).
A fifth aspect of the disclosure relates to a method of reducing the risk of a disease or disorder ated with oc—amino—B—carboxymuconate—8—semialdehyde decarboxylase (ACMSD) dysfunction comprising administering to the subject suffering from or susceptible to developing a disease or disorder associated with ACMSD dysfunction a eutically effective amount of one or more compounds of Formula (I).
A sixth aspect of the disclosure relates to a method of treating a disease or disorder associated with reduced nicotinamide adenine dinucleotide (NAD+) levels comprising administering to the subject suffering from or susceptible to developing a disease or disorder associated with reduced NAD+ levels a therapeutically effective amount of one or more compounds of Formula (I).
A seventh aspect of the disclosure relates to a method of preventing a disease or disorder ated with reduced nicotinamide e dinucleotide (NADl) levels comprising administering to the subject suffering from or susceptible to developing a disease or disorder associated with reduced NAD+ levels a therapeutically effective amount of one or more compounds of Formula (I).
An eighth aspect of the disclosure relates to a method of reducing the risk of a disease or disorder ated with reduced nicotinamide adenine dinucleotide (NAD+) levels comprising administering to the subject suffering from or susceptible to developing a disease or disorder associated with reduced NAD+ levels a therapeutically effective amount of one or more compounds of Formula (I).
An ninth aspect of the disclosure s to a method of treating a disorder associated with mitochondrial dysfunction comprising stering to the t ing from or tible to ping a metabolic disorder a therapeutically effective amount of one or more compounds of Formula (I) that increases intracellular nicotinamide adenine dinucleotide (NADl).
[Annotation] KEB A tenth aspect of the disclosure relates to a method of promoting oxidative metabolism comprising administering to the subject suffering from or susceptible to developing a metabolic disorder a therapeutically effective amount of one or more compounds of Formula (I) that increases ellular nicotinamide adenine eotide (NAD+).
An eleventh aspect of the disclosure relates to a method for the manufacture of a medicament for ng a disease or condition mediated by ACMSD, wherein the medicament comprises a compound of Formula (I), or a pharmaceutically acceptable salt thereof.
A twelfth aspect of the disclosure relates to a pharmaceutical composition for use in a method for treating a disease or condition mediated by ACMSD, wherein the medicament comprises a compound of Formula (I), or a ceutically acceptable salt thereof.
A thirteenth aspect of the disclosure s to a method of treating a disease or disorder ated with a—amino—B—carboxymuconate—s—semialdehyde decarboxylase ) dysfunction, comprising administering to a subject in need thereof, a therapeutically effective amount of compound having one of the following Formulae: ation] KEB [Annotation] KEB or a pharmaceutically acceptable salt thereof.
A fourteenth aspect of the disclosure relates to the use of a compound of a (I), or a pharmaceutically acceptable salt thereof in the manufacture of a medicament for treating, preventing or reducing the risk of a disease or disorder ated with OL-amino-B- carboxymuconate—s—semialdehyde decarboxylase ) ction.
A ?fteenth aspect of the disclosure relates to the use of a compound of Formula (I), or a pharmaceutically acceptable salt thereof in the manufacture of a medicament for treating, preventing or reducing the risk of a e or disorder associated with reduced nicotinamide adenine dinucleotide (NAD+) levels.
A sixteenth aspect of the disclosure relates to the use of a compound of Formula (I), or a pharmaceutically acceptable salt thereof in the manufacture of a medicament for treating, preventing or reducing the risk of a disorder associated with mitochondrial dysfunction.
A eenth aspect of the disclosure s to the use of a compound of Formula (I), or a pharmaceutically acceptable salt thereof in the manufacture of a medicament for promoting oxidative metabolism.
A eighteenth aspect of the disclosure relates to the use of a compound of Formula (I), a pharmaceutically acceptable salt thereof in the manufacture of a medicament for treating, preventing or reducing the risk of a disease or disorder associated with Ot-al’l’liHO-B- carboxymuconate—s—semialdehyde decarboxylase (ACMSD) dysfunction.
A nineteenth aspect of the disclosure relates to a compound of Formula (I), or a pharmaceutically acceptable salt thereof for use as a medicament for treating, preventing or reducing the risk of a e or er associated with reduced nicotinamide adenine dinucleotide (NADl) levels.
A twentieth aspect of the disclosure relates to a compound of Formula (I), or a pharmaceutically acceptable salt thereof for use as a medicament for treating, ting or reducing the risk of a disorder ated with mitochondrial dysfunction.
A twenty ?rst aspect of the disclosure relates to a compound of Formula (I), or a pharmaceutically acceptable salt thereof for use in treating, preventing or reducing the risk of a disease or disorder associated with reduced namide adenine dinucleotide (NAD+) A twenty second aspect of the disclosure relates to a compound of Formula (I), or a ceutically acceptable salt thereof for use in for treating, preventing or reducing the risk of a disorder associated with mitochondrial dysfunction.
[Annotation] KEB A twenty third aspect of the disclosure relates to a compound of Formula (I), or a ceutically acceptable salt thereof for use in promoting oxidative metabolism.
A twenty fourth aspect of the disclosure relates to a compound having the one of the following Formula: [Annotation] KEB or a pharmaceutically acceptable salt thereof in the manufacture of a ment for treating a disease or disorder associated With oc— amino—B—carboxymuconate—s—semialdehyde decarboxylase (ACMSD) dysfunction.
A twenty ?fth aspect of the disclosure relates to a compound having the one of the following Formula: ation] KEB or a pharmaceutically acceptable salt thereof [Annotation] KEB for use as a medicament for treating a disease or disorder associated with oc-amino-B- carboxymuconate—s—semia1dehyde decarboxylase (ACMSD) dysfunction.
A twenty sixth aspect of the disclosure relates to a compound having the one of the ing Formula: [Annotation] KEB or a pharmaceutically acceptable salt f for use in treating a disease or disorder associated with oc-amino—B—carboxymuconate—a— semialdehyde decarboxylase (ACMSD) ction.
In certain s, the ACMSD modulating compounds may be administered alone or in combination with other compounds, including other ACMSD modulating compounds, or other eutic agents.
Unless ise de?ned, all technical and scientific terms used herein have the same meaning as commonly understood by one of ry skill in the art to which this disclosure belongs. In the specification, the singular forms also include the plural unless the context clearly dictates otherwise. Although methods and materials similar to or equivalent to those described herein can be used in the practice and testing of the disclosure, suitable methods and materials are described below. All publications, patent applications, patents, and other references mentioned herein are incorporated by reference. The references cited herein are not admitted to be prior art to the claimed disclosure. In the case of con?ict, the t speci?cation, including de?nitions, will control. In addition, the materials, methods, and examples are illustrative only and not intended to be limiting.
Other es and advantages of the disclosure will be apparent from the following [Annotation] KEB detailed description and .
BRIEF DESCRIPTION OF THE DRAWINGS is a graph of the measured NAD+ levels in human y hepatocytes treated with Compound 4, detected by LC—MS/MS. is a graph of the measured NAD+ levels in murine primary hepatocytes treated with different concentrations of Compound 17 for 24 hours and detected by MS. The data indicate an se in NAD+ levels in murine primary cytes treated with Compound 17. is a graph of the measured NAD+ content in human primary hepatocytes treated with various concentrations of Compound 1 and 2-ethylhexyl)phthalate (MEHP), as a control. is a graph of gene expression of Acmsd, Sod—1, and Sod—2 as determined by RT-qPCR in AML-12 cells treated with Compound 1 for 24 hours. is a graph of gene expression of Sod—1 and Sod—2 as determined by RT-qPCR in Hepa—1.6 cells treated with Compound 1 for 24 hours. is a graph of gene expression of Acmsd, Sod—l, Sod— 2, and chla as determined by RT—qPCR in primary mouse hepatocytes treated with Compound 17 for 24 hours. Bar graphs represent mean i SEM, ***p 5 0.005 is a graph showing the modulation of SOD2 activity in AML-12 cells treated for 24 hours with Compound 1. shows a graph of the modulation of SOD2 activity in AML—12 cells treated for 24 hours with Compound 17. shows a graph of the modulation of SOD2 activity in primary murine hepatocytes cells treated for 24 hours with Compound 17. is a gel showing the effect of Compound 1 on the FoxOl phosphorylation . is a gel showing the effect of Compound 17 on the FoxOl phosphorylation levels. is a graph of changes in acsmd-I and sod-3 expression at mRNA levels measured in N2 wild type worms at day 2 of ood by acmsd-l RNAi silencing in habditis elegans (C. elegans). is a graph of the induction of sod-3 expression at protein levels in N2 worms at day 3 of adulthood, quanti?ed by using SOD-3 gfp reporter strain, after acmsd—J RNAi silencing in C. elegans. is a graph showing the survival of worms upon downregulation of acmsd—J by feeding a speci?c RNAi in C. elegans. FIG.
[Annotation] KEB 7C shows that downregulation of acmsd—I improves the survival ofworms in a SIR-2.1 and DAF—l6 dependent manner. is a graph showing that downregulation of acmsd-I improves the stress—resistance of worms when they are exposed to paraquat—induced oxidative stress. is a graph showing the mobility of worms over time fed with acmsd—J RNAi under paraquat-induced oxidative stress condition. As FIGs. 7C—7E show, reduced acmsd—I expression improves the survival and fitness of worms under paraquat-induced oxidative stress. is a graph that shows the survival of worms under paraquat—induced stress conditions when exposed to acmsd—I RNAi during different stages of development. illustrates that the ement of the survival of worms under paraquat ions is independent of the developmental stage at which the worms were exposed to the acmsd-I RNAi. is a graph showing worm survival under paraquat—induced stress ions upon gulation of acmsd—J combined with daf-16 downregulation by feeding a specific RNAi in C. elegans. shows that the improved survival of worms with gulated acmsd—J is daf-I6 dependent under paraquat—induced oxidative stress conditions. is a graph of changes in caspase3/7 activity induced by tin in MDCK cells when treated with different trations of Compound 18 in combination with cisplatin. is a graph of changes in e3/7 activity induced by cisplatin in MDCK cells when treated with ent concentrations of nd 18 one hour prior to the addition of cisplatin.
DETAILED DESCRIPTION OF THE DISCLOSURE Compounds ofFormula (I) The present disclosure relates to compounds of Formula (I): or a pharmaceutically acceptable salt or tautomer thereof, wherein: X is O, OH, or Cl; L is ‘(CH2)mCH2CH2-, —(CH2)mY(CH2)p—, "(CH2)mC(O)(CH2)p'a [Annotation] KEB —(CH2)mC(O)O(CH2)p—, —(CH2)mC(O)NR2(CH2)p—, or —(CHz)mNR2C(O)(CH2)p; Y is O, N or S(O)q; R1 is C6-C10 aryl or heteroaryl, wherein the aryl and aryl are substituted with Ra and Rb, and optionally substituted with one or more Re; R2 is H or C1-C6 alkyl; one of R3 and Rb is hydrogen and the other is -(CH2)rC02Rx, -OCH2C02RX, -(CH2)rtetrazole, -(CH2)roxadiazolone, -(CH2)rtetrazolone, rthiadiazolol, r isoxazol—3 —ol, —(CH2)rP(O)(OH)ORX, rS(O)20H, —(CH2)rC(O)NHCN, or —(CH2)rC(O)NHS(O)2alkyl; Rc is H, C1-C6 alkyl, C1-C6 haloalkyl, halogen, —CN, —ORX, —COZRX, or N02; Rd is methyl, optionally substituted 5— to 10-membered aryl, optionally substituted 5— or 6—membered heteroaryl, or optionally tuted 5— or 6—membered ycle; each Rx is independently at each occurrence hydrogen or C1-C6 alkyl; each R6 is independently C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, halogen, -ORy, C1-C6 haloalkyl, -NHRZ, -OH, or -CN; Rf is H or absent; each Ry and RZ is ndently hydrogen, C1-C6 alkyl, or C1—C6 haloalkyl, each m and p independently is 0, l or 2, wherein m + p < 3; q is 0, l, or 2; r is 0 or 1; and the dotted line is an optional double bond; with the o that Rc is not hydrogen or —CN when X is O, L is —SCH2- and RC1 is optionally substituted phenyl, Rc is not C1-C6 alkyl when X is O, L is —SCH2- and Rd is methyl, and that RC is not —CN when L is —SCH2— and Rd is 2-furyl.
In some embodiments of Formula (I), X is O, OH, or Cl. In other embodiments, X is O. In other embodiments, X is OH. In other embodiments, X is Cl.
In some embodiments of Formula (I), L is —(CH2)mCH2CH2-, —(CH2)mY(CH2)p—, —(CH2)mC(O)(CH2)p—, —(CH2)mC(O)O(CH2)p—, —(CH2)mC(O)NR2(CH2)p—, or -(CH2)mNR2C(O) (CH2)p. In other embodiments, L is —CH2CH2—, —CH2CH2CH2—,—SCH2—, —SCH2CH2—, —Cst—, —CHZSCH2-, —CH2CHZS—, —S(O)CH2—, —S(O)CH2CH2—, —CHZS(O)—, —CHZS(O)CH2-, 2$(O)—, —S(O)2CH2-, —S(O)2CH2CH2-, —Cst(O)2—, —CH28(O)2CH2—, —CH2CHZS(O)2—, —OCH2—, —OCH2CH2—, —CH20-, —CHzOCH2—, —CH2CH20—,—NR2CH2—, — CH2NR2-, -CH2NR2CH2-, — CH2CH2NR2-, —NR2CH2CH2-, -C(O)CH2-, -C(O)CH2CH2-, [Annotation] KEB -C(O)O-, —C(O)OCH2—, —CH2C(O)O—, —C(O)NR2—, —C(O)NR2CHz-, —NR2C(O), —NR2C(O)CH2, or —CH2NR2C(O). In other embodiments, L is —CH2CH2—, —CH2CH2CH2—, —SCH2—, —SCH2CH2—, —S(O)CH2—, —S(O)CHZCH2—,—S(O)2CH2—, —S(O)2CH2CH2—, —OCH2—, H2—, —NR2CH2-, 2CH2-, —C(O)CH2—, —C(O)CH2CH2—, —C(O)O-, —C(O)OCH2— ,—CH2C(O)O—, —C(O)NR2—, —C(O)NR2CH2—, — NR2C(O), or — NR2C(O)CH2. In other embodiments, L is -CH2CH2-, -CH2C(O)-, -C(O)CH2-, -NR2CH2-, -CH2NR2-, , -CH20-, —SCH2—, —CH2s—, H2—, —CHZS(O)—, O)2—, or CH2—.
In some ments of Formula (I), R1 is C5-C10 aryl or heteroaryl, wherein the aryl and heteroaryl are substituted with R2‘ and Rb, and optionally substituted with one or more Re.
In other embodiments, R1 is C6-C10 aryl substituted with Ra and Rb, and optionally substituted with one or more Re. In other embodiments, R1 is heteroaryl substituted with R21 and Rb, and optionally substituted with one or more Re. In further embodiments, R1 is phenyl substituted with R21 and Rb, and optionally substituted with one or more Re.
In some embodiments of Formula (I), Ra is —(CH2)rCOZRX, —OCH2C02RX, -(CH2)rtetrazole, roxadiazolone, -(CH2)rtetrazolone, -(CH2)rthiadiazolol, —(CH2)r ol—3 —ol, —(CH2)rP(O)(OH)ORX, —(CH2)rS(O)ZOH, —(CH2)rC(O)NHCN, or —(CH2)rC(O)NHS(O)2alky1. In other embodiments, Ra is -(CH2)rC02Rx, OZRX, tetrazole, -(CH2)tetrazole, oxadiazolone, -(CH2)oxadiazolone, tetrazolone, -(CH2)tetrazolone, thiadiazolol, -(CH2)thiadiazolol, isoxazolol, -(CH2) isoxazolol, —P(O)(OH)ORX, —(CH2)P(O)(OH)ORX, —S(O)20H , —(CH2)S(O)ZOH, —C(O)NHCN —(CH2)C(O)NHCN, —C(O)NHS(O)zalkyl, or -(CH2)C(O)NHS(O)zalkyl. In other embodiments, R3 is hydrogen, COZR", CHZCOZRX, tetrazole, or zolone. In further embodiments, Ra is hydrogen, COZH, CHZCOZH, tetrazole, or 1,2,4—oxadiazol—5(4H)-one.
In some embodiments of Formula (I), Rb is —(CH2)rC02RX, —OCH2C02RX, -(CH2)rtetrazole, —(CH2)roxadiazolone, -(CH2)rtetrazolone, -(CH2)rthiadiazolol, —(CH2)r isoxazol—3 —ol, —(CH2)IP(O)(OH)ORX, —(CH2)rS(O)ZOH, —(CH2)IC(O)NHCN, or —(CH2)rC(O)NHS(O)2alkyl. In other embodiments, Rb is -(CH2)rCOZRX, —OCH2COZRX, tetrazole, tetrazole, oxadiazolone, —(CH2)oxadiazolone, tetrazolone, -(CH2)tetrazolone, thiadiazolol, -(CH2)thiadiazolol, isoxazolol, -(CH2) isoxazolol, —P(O)(OH)ORX, P(O)(OH)ORX, —S(O)20H , —(CH2)S(O)20H, —C(O)NHCN —(CH2)C(O)NHCN, —C(O)NHS(O)2alkyl, or —(CH2)C(O)NHS(O)2alkyl. In other embodiments, Rb is hydrogen, COZR", RX, tetrazole, or oxadiazolone. In further [Annotation] KEB ments, Rb is en, COZH, CHZCOZH, tetrazole, or l,2,4—oxadiazol—5(4H)—one. In further embodiments, Rb is hydrogen.
In some embodiments of Formula (I), R0 is H, C1—C6 alkyl, C1-C6 haloalkyl, halogen, —CN, —ORX, —C02Rx, or N02. In other embodiments, RC is C1—C6 alkyl, C1—C6 haloalkyl, halogen, —CN, —ORX, , or N02. In other embodiments, Rc is n, —CN, —OR", or C1-C6 alkyl. In other embodiments, Rc is halogen, —CN, —ORX, or C1-C3 alkyl. In other embodiments, Rc is H, —CN, or n. In other embodiments, Rc is —CN or halogen.
In some embodiments of Formula (I), Rd is methyl, optionally tuted 5- to 10- membered aryl, optionally substituted 5— or ered heteroaryl, or optionally substituted - or 6-membered carbocycle. In other embodiments, Rd is methyl, optionally cyclohexyl, ally substituted pyridinyl, ally substituted thiazolyl, optionally substituted phenyl, or optionally substituted thienyl. In other embodiments, Rd is methyl, cyclohexyl, pyridinyl, thiazolyl, phenyl, or thienyl. In other embodiments, Rd is cyclohexyl, pyridinyl, thiazolyl, phenyl, or thienyl, n each is optionally substituted with one or more substituents independently selected from halogen, C1—C6 alkyl, C1—C6 hydroxyalkyl, C1—C6 alkoxy, C1-C6 haloalkyl, C1-C6 haloalkoxy, -OH, CN, and amino. In other embodiments, Rd is cyclohexyl, pyridinyl, thiazolyl, phenyl, or thienyl, wherein each is optionally substituted with one or more substituents independently selected from halogen, C1—C6 alkyl, C1—C6 hydroxyalkyl, C1-C6 alkoxy, C1-C6 haloalkyl, and C1-C6 haloalkoxy. In other embodiments, Rd is cyclohexyl, pyridinyl,thiazoly1, phenyl, or l, wherein each is optionally substituted with one or more halogen. In yet other embodiments, Rd is exyl, pyridinyl, thiazolyl, phenyl, or l. In other embodiments, Rd is cyclohexyl, pyridinyl, thiazolyl, phenyl, 4-chlorophenyl, 4—methylphenyl, or thienyl.
In some embodiments of Formula (I), each R6 is independently C1-C6 alkyl, C2—C6 alkenyl, C2-C6 alkynyl, halogen, -ORy, C1-C6 haloalkyl, —NHRZ, -OH, or —CN. In other embodiments, C1-C4 alkyl, C2-C4 alkenyl, C2—C4 alkynyl, halogen, —ORy, C1—C4 haloalkyl, - NHRZ, —OH, or —CN.
In some embodiments of Formula (I), Rf is H or absent. In other embodiments, Rf is H. In other embodiments, Rf is absent, when N to which it is attached participates in a double bond.
[Annotation] KEB In some embodiments of Formula (I), RX is hydrogen or C1—C6 alkyl. In other ments, Rx is hydrogen or C1—C3 alkyl, In further embodiments, Rx is en, , ethyl, n-propyl, or iso—propyl.
In some embodiments of Formula (I), Ry is independently hydrogen, C1-C6 alkyl, or C1-C6 haloalkyl. In other embodiments, Ry is hydrogen, C1-C3 alkyl, or C1-C3 haloalkyl.
In some embodiments of a (I), each RZ is independently hydrogen, C1-C6 alkyl, or C1-C6 haloalkyl. In other embodiments, RZ is hydrogen, C1-C3 alkyl, or C1—C3 haloalkyl.
In some embodiments of Formula (I), m is 0, l or 2. In other embodiments, m is 0.
In other embodiments, m is 1. In yet other embodiments, m is 2.
In some embodiments of Formula (I), p is 0, l or 2. In other ments, p is 0. In other embodiments, p is 1. In yet other embodiments, p is 2.
In some embodiments of Formula (I), m + p < 3; In some embodiments of Formula (I), q is 0, l, or 2. In other embodiments, q is 0. In other embodiments, q is 1. In other ments, q is 2.
In some embodiments of Formula (I), r is 0 or 1. In other embodiments, r is 0. In other embodiments, r is 1.
In some embodiments of Formula (I), the dotted line is a single bond. In other ments, the dotted line is a double bond.
In some embodiments of Formula (I), one of Ra and Rb is hydrogen and the other is COzRX, CH2C02RX, tetrazole, or oxadiazolone. In other embodiments, Rb is hydrogen and R8 is CHZCOZH, tetrazole, or (l,2,4-oxadiazol-5(4H)-one).
In some embodiments of Formula (I), Rb is hydrogen, RC is —CN, Rd is thienyl, and R3 is H, tetrazole, or (l,2,4-oxadiazol-5(4H)-one).
In some embodiments of Formula (I), Rc is halogen, R8 is —COZH, and Rb is H. In other embodiments, Rc is -Br, Ra is —COZH, and Rb is H. In further embodiments, Rc is -Cl, Ra is —C02H, and Rb is H.
In some embodiments of Formula (I), R0 is halogen, Ra is tetrazole, and Rb is H. In other embodiments, Rc is —Br, Ra is tetrazole, and Rb is H. In further embodiments, RC is -Cl, Ra is tetrazole, and Rb is H.
[Annotation] KEB In some embodiments of Formula (I), R0 is halogen, Ra is —CH2COZH, and Rb is H. In other embodiments, RC is —Br, R3 is ZH, and Rb is H. In further embodiments, Rc is - c1, Ra is 2H, and Rb is H.
In some embodiments of a (I), R6 is halogen, R3 is (l,2,4-oxadiazol-5(4H)— one), and Rb is H. In other embodiments, Rc is -Br, Ra is (l,2,4-oxadiazol-5(4H)—one), and Rb is H. In other embodiments, Rc is —Cl, Ra is (l,2,4-oxadiazol-5(4H)—one), and Rb is H.
In some embodiments of Formula (I), R0 is -CN, Ra is —C02H, and R1) is H. In other embodiments, RC is —CN, R21 is —CH2C02H, and Rb is H. In other embodiments, Rc is —CN, R3 is tetrazole, and Rb is H. In yet other embodiments, Rc is -CN, Ra is (l,2,4-oxadiazol-5(4H)— one), and Rb is H.
In some embodiments of Formula (I), R0 is not hydrogen or —CN and X is O, L is — SCHz- and Rd is optionally substituted phenyl. In other embodiments, Rc is not C1-C6 alkyl and X is O, L is —SCH2- and Rd is methyl. In other embodiments, Rc is not —CN and X is O, L is —SCH2- and Rd is l.
In some embodiments of Formula (I), Rc is not hydrogen or —CN when X is O, L is — SCHZ— and Rd is optionally substituted phenyl.
In some embodiments of Formula (I), Rc is not C1-C6 alkyl when X is O, L is —SCH2- and Rd is methyl.
In some embodiments of Formula (I), Rc is not —CN when X is O, L is —SCH2- and Rd is 2-furyl.
In one embodiment, the compound of Formula (I) is represented by Formula (Ia): | 1 Rd NAL/R (Ia) or a pharmaceutically acceptable salt, or tautomer thereof, L is ‘(CH2)mCH2CH2-, ‘(CH2)mY(CH2)p'a '(CH2)mC(O)(CH2)p'a —(CH2)mC(O)O(CH2)p—, mC(O)NR2(CH2)p—, or —(CH2)mNR2C(O)(CH2)p; Y is O, N or S(O)q; [Annotation] KEB R1 is C6-C10 aryl or heteroaryl, wherein the aryl and heteroaryl are substituted with Ra and Rb, and optionally substituted with one or more Re; R2 is H or C1-C6 alkyl; one of R3 and Rb is hydrogen and the other is —(CH2)rCOZRx, -OCH2C02RX, -(CH2)rtetrazole, -(CH2)roxadiazolone, -(CH2)rtetrazolone, -(CH2)rthiadiazolol, -(CH2)r isoxazol-3 -ol, -(CH2)rP(O)(OH)ORX, -(CH2)rS(O)20H, -(CH2)rC(O)NHCN, or —(CH2)rC(O)NHS(O)2alkyl; Rc is H, C1—C6 alkyl, C1-C6 kyl, halogen, —CN, —ORX, —COzRX, or N02; Rd is methyl, optionally substituted 5— to 10-membered aryl, optionally substituted 5— or 6—membered heteroaryl, or optionally substituted 5— or 6—membered carbocycle; each Rx is independently at each ence hydrogen or C1-C6 alkyl; each R6 is independently C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, halogen, -ORy, C1—C6 haloalkyl, -NHRZ, —OH, or —CN; each Ry and RZ is independently en, C1-C6 alkyl, or C1-C6 haloalkyl, each m and p independently is 0, l or 2, wherein m + p < 3; q is 0,1, or 2; and r is 0 or 1; with the proviso that RC is not en or —CN when L is —SCH2- and Rd is optionally substituted phenyl, Rc is not C1-C6 alkyl when L is —SCH2- and Rd is methyl, and that RC is not —CN when L is —SCH2— and Rd is 2-furyl.
In some embodiments of Formula (Ia), L is -CH2CH2-, -CH2C(O)-, -C(O)CH2-, -NR2CH2-, -CH2NR2-, -OCH2-, —CH20—, —SCH2—, , —S(O)CH2—, —CHZS(O)—, —CHZS(O)2—, or —S(O)2CH2—; Y is O, N or S(O)q; R1 is C6—C10 aryl or heteroaryl, wherein the aryl and heteroaryl are substituted with Ra and Rb, and optionally substituted with one or more Re; R2 is H or C1-C6 alkyl; one of Ra and Rb is hydrogen and the other is —(CH2)rCOZRX, —OCH2C02RX, -(CH2)rtetrazole, -(CH2)roxadiazolone, -(CH2)rtetrazolone, -(CH2)rthiadiazolol, —(CH2)r ol—3 —ol, rP(O)(OH)ORX, —(CH2)rS(O)ZOH, rC(O)NHCN, or r S(O)2alkyl; Rc is C1-C6 alkyl, C1-C6 haloalkyl, halogen, —CN, —ORX, —COZRX, or N02; Rd is methyl, optionally substituted 5- to 10-membered aryl, optionally substituted 5— or 6—membered heteroaryl, or optionally substituted 5— or 6—membered carbocycle; [Annotation] KEB each RX is independently at each occurrence hydrogen or C1—C6 alkyl; each R6 is independently C1—C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, halogen, -ORy, C1—C6 haloalkyl, -NHRZ, -OH, or —CN; each Ry and R2 is ndently hydrogen, C1-C6 alkyl, or C1-C6 haloalkyl; each m and p independently is 0, l or 2, n m + p < 3; q is 0,1, or 2; and r is 0 or 1; with the proviso that R0 is not —CN when L is —SCH2- and Rd is optionally substituted phenyl, RC is not C1-C6 alkyl when L is —SCH2— and Rd is methyl, and that R6 is not —CN when L is —SCH2- and Rd is 2-furyl.
In some embodiments of Formula (Ia), L is —(CH2)mCH2CH2-, —(CH2)mY(CH2)p-, -(CH2)mC(O)(CHz)p-, —(CH2)mC(O)O(CH2)p—, mC(O)NR2(CHz)p-, or -(CH2)mNR2C(O) (CH2)p. In other embodiments, L is —CH2CH2—, —CH2CH2CH2—,—SCH2—, —SCH2CH2—, —Cst—, —CHZSCH2-, —CH2CHZS-, —S(O)CH2—, —S(O)CH2CH2—, —CHZS(O)—, —CHZS(O)CH2-, —CH2CH2S(O)-, —S(O)2CH2-, —S(O)2CH2CH2-, —CHZS(O)2-, —CHZS(O)2CH2—, ZS(O)z-, —OCH2—, —OCH2CH2—, —CH20—, —CH20CH2-, —CH2CHzO—,—NR2CH2—, — CHZNR2-, -CH2NR2CH2-, — CHZCHgNR2-, —NR2CH2CH2-, H2-, -C(O)CH2CH2-, -, CH2—, —CH2C(O)O—, —C(O)NR2—, —C(O)NR2CH2-, —NR2C(O), —NR2C(O)CH2, or —CH2NR2C(O). In other embodiments, L is —CH2CH2—, —CH2CH2CH2—, —SCH2—, —SCH2CH2—, —S(O)CH2—, —S(O)CH2CH2-,—S(O)2CH2-, —S(O)2CH2CH2-, —OCH2—, —OCH2CH2-, —NR2CH2—, —NR2CHZCH2—, —C(O)CH2—, —C(O)CH2CHz-, -C(O)O-, CH2— ,—CH2C(O)O—, —C(O)NR2—, -C(O)NR2CH2—, — NR2C(O), or — NR2C(O)CH2. In other embodiments, L is —CH2CH2—, —CH2C(O)—, —C(O)CH2-, —NR2CH2—, —CH2NR2—, —OCH2—, -CHzO-, -SCH2-, -Cst-, -S(O)CH2-, O)-, -CH2$(O)2-, or -S(O)2CH2-.
In some embodiments of Formula (Ia), R1 is C6-C10 aryl or heteroaryl, wherein the aryl and heteroaryl are substituted with R8 and Rb, and optionally substituted with one or more Re. In other embodiments, R1 is C6-C10 aryl substituted with R8 and Rb, and optionally substituted with one or more Re. In other embodiments, R1 is heteroaryl substituted with Ra and Rb, and optionally substituted with one or more Re. In further embodiments, R1 is phenyl substituted with Ra and Rb, and optionally substituted with one or more Re.
In some embodiments of Formula (Ia), R21 is -(CH2)rCOZRX, —OCH2C02RX, -(CH2)rtetrazole, —(CH2)roxadiazolone, rtetrazolone, rthiadiazolol, —(CH2)r isoxazol—3 —ol, —(CH2)rP(O)(OH)ORX, —(CH2)rS(O)ZOH, —(CH2)rC(O)NHCN, or [Annotation] KEB —(CH2)rC(O)NHS(O)2alky1. In other embodiments, Ra is —(CH2)rCOZRX, -OCH2COZRX, tetrazole, —(CH2)tetrazole, oxadiazolone, oxadiazolone, tetrazolone, -(CH2)tetrazolone, thiadiazolol, -(CHz)thiadiazolol, isoxazolol, —(CH2) isoxazolol, —P(O)(OH)ORX, —(CH2)P(O)(OH)ORX, -S(O)20H , —(CH2)S(O)ZOH, HCN —(CH2)C(O)NHCN, —C(O)NHS(O)2alkyl, or —(CH2)C(O)NHS(O)2alky1. In other embodiments, R3 is hydrogen, COZR", CHZCOZRX, tetrazole, or oxadiazolone. In further embodiments, R2‘ is hydrogen, COZH, CHZCOZH, tetrazole, or 1,2,4-oxadiazol-5(4H)-one.
In some embodiments of Formula (Ia), Rb is -(CH2)rCOZRx, 'OCHZCOQRX, -(CH2)rtetrazole, -(CH2)roxadiazolone, -(CH2)rtetrazolone, -(CH2)rthiadiazolol, r isoxazol—3 —ol, —(CH2)rP(O)(OH)ORX, -(CH2)rS(O)zOH, —(CH2)rC(O)NHCN, or —(CH2)rC(O)NHS(O)2alky1. In other embodiments, Rb is -(CH2)rC02RX, OZRX, tetrazole, -(CH2)tetrazole, oxadiazolone, -(CH2)oxadiazolone, tetrazolone, -(CH2)tetrazolone, thiadiazolol, thiadiazolol, isoxazolol, —(CH2) isoxazolol, —P(O)(OH)ORX, —(CH2)P(O)(OH)ORX, 0H , S(O)ZOH, —C(O)NHCN —(CH2)C(O)NHCN, HS(O)2alkyl, or —(CH2)C(O)NHS(O)2alkyl. In other embodiments, Rb is hydrogen, COZR", CHZCOZRX, tetrazole, or oxadiazolone. In r embodiments, Rb is hydrogen, COZH, CHZCOZH, tetrazole, or 1,2,4-oxadiazol-5(4H)-one. In further embodiments, Rb is en.
In some embodiments of Formula (Ia), RC is H, C1-C6 alkyl, C1-C5 haloalkyl, halogen, —CN, —ORX, —C02RX, or N02. In other embodiments, Rc is C1—C6 alkyl, C1—C6 haloalkyl, halogen, —CN, —ORX, -C02RX, or N02. In other embodiments, Rc is halogen, —CN, —ORX, or C1-C6 alkyl. In other embodiments, Rc is n, —CN, —ORX, or C1-C3 alkyl. In other embodiments, Rc is H, —CN, or halogen. In other embodiments, Rc is —CN or halogen.
In some embodiments of Formula (Ia), Rd is methyl, optionally substituted 5- to 10- membered aryl, optionally substituted 5— or 6-membered heteroaryl, or optionally substituted — or 6-membered carbocycle. In other embodiments, Rd is methyl, optionally cyclohexyl, ally tuted pyridinyl, optionally substituted thiazolyl, optionally substituted phenyl, or optionally substituted thienyl. In other embodiments, Rd is cyclohexyl, pyridinyl, thiazolyl, phenyl, or thienyl, wherein each is optionally substituted With one or more substituents independently selected from halogen, C1-C6 alkyl, C1-C6 yalkyl, C1—C6 alkoxy, C1-C6 haloalkyl, C1-C6 haloalkoxy, -OH, CN, and amino. In other ments, Rd is cyclohexyl, pyridinyl, thiazolyl, phenyl, or thienyl, wherein each is optionally substituted with one or more substituents independently selected from halogen, C1—C6 alkyl, C1—C6 [Annotation] KEB hydroxyalkyl, C1—C6 alkoxy, C1-C6 haloalkyl, and C1-C6 haloalkoxy. In other embodiments, Rd is cyclohexyl, pyridinyl, thiazolyl, phenyl, or thienyl, n each is optionally substituted With one or more halogen. In other embodiments, Rd is methyl, cyclohexyl, pyridinyl, thiazolyl, phenyl, or thienyl. In yet other ments, Rd is cyclohexyl, pyridinyl, thiazolyl, phenyl, or thienyl. In other embodiments, Rd is exyl, pyridinyl, thiazolyl, phenyl, 4-chlorophenyl, ylphenyl, or thienyl.
In some embodiments of a (Ia), each R6 is independently C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, halogen, -ORy, C1-C6 kyl, -NHRZ, -OH, or —CN. In other embodiments, C1-C4 alkyl, C2-C4 alkenyl, C2-C4 alkynyl, halogen, -ORy, C1-C4 haloalkyl, - NHRZ, —OH, or —CN.
In some embodiments of Formula (Ia), Rx is hydrogen or C1-C6 alkyl. In other embodiments, Rx is hydrogen or C1-C3 alkyl. In further embodiments, RX is en, methyl, ethyl, n—propyl, or iso—propyl.
In some embodiments of Formula (Ia), Ry is independently hydrogen, C1—C5 alkyl, or C1-C6 kyl. In other embodiments, Ry is hydrogen, C1-C3 alkyl, or C1-C3 haloalkyl.
In some embodiments of Formula (Ia), each RZ is independently en, C1-C6 alkyl, or C1—C6 haloalkyl. In other embodiments, RZ is hydrogen, C1-C3 alkyl, or C1—C3 haloalkyl.
In some embodiments of Formula (Ia), m is 0, l or 2. In other embodiments, m is 0.
In other embodiments, m is 1. In yet other embodiments, m is 2.
In some embodiments of a (Ia), p is 0, 1 or 2. In other embodiments, p is 0. In other embodiments, p is 1. In yet other embodiments, p is 2.
In some embodiments of Formula (Ia), q is 0, l, or 2. In other embodiments, q is 0. In other embodiments, q is 1. In other embodiments, q is 2.
In some embodiments of Formula (Ia), r is 0 or 1. In other embodiments, r is 0. In other embodiments, r is 1.
In some embodiments of Formula (Ia), one of R8 and Rb is hydrogen and the other is COZRX, CHzCOZRX, tetrazole, or oxadiazolone. In other embodiments, Rb is hydrogen and Ra is CHZCOZH, tetrazole, or (1,2,4-oxadiazol-5(4H)-one).
In some embodiments of a (Ia), Rb is hydrogen, Rc is —CN, Rd is thienyl, and Ra is CHZCOZH, tetrazole, or (1,2,4-oxadiazol-5(4H)-one).
[Annotation] KEB In some embodiments of Formula (Ia), Rc is halogen, Ra is —C02H, and Rb is H. In other embodiments, RC is —Br, R3 is —COzH, and Rb is H. In further embodiments, RC is -Cl, Ra is —COzH, and Rb is H.
In some ments of Formula (Ia), Rc is halogen, R3 is tetrazole, and Rb is H. In other embodiments, Rc is -Br, Ra is tetrazole, and Rb is H. In further ments, Rc is -Cl, R3 is tetrazole, and Rb is H.
In some embodiments of Formula (Ia), Rc is halogen, Ra is —CH2C02H, and Rb is H.
In other embodiments, RC is -Br, R8 is —CH2C02H, and Rb is H. In further embodiments, Rc is -Cl, Ra is —CH2C02H, and Rb is H.
In some embodiments of Formula (Ia), Rc is halogen, R3 is (l,2,4-oxadiazol-5(4H)- one), and Rb is H. In other embodiments, Rc is -Br, Ra is (l,2,4-oxadiazol-5(4H)—one), and Rb is H. In other embodiments, Rc is -C1, R3 is (l,2,4-oxadiazol-5(4H)-one), and Rb is H.
In some embodiments of Formula (Ia), Rc is -CN, Ra is —COZH, and Rb is H. In other embodiments, RC is -CN, R21 is —CH2C02H, and Rb is H. In other embodiments, Rc is -CN, Ra is tetrazole, and Rb is H. In yet other embodiments, Rc is -CN, R3 is (l,2,4-oxadiazol-5(4H)— one), and Rb is H.
In some embodiments of a (Ia), Rc is not hydrogen or —CN and L is —SCH2- and Rd is optionally tuted phenyl. In other embodiments, Rc is not C1-C6 alkyl and L is —SCH2- and Rd is methyl. In other embodiments, Rc is not —CN and L is —SCH2- and Rd is 2- furyl.
In some embodiments of Formula (Ia), Rc is not hydrogen or —CN when L is —SCH2- and Rd is optionally substituted phenyl.
In some embodiments of Formula (Ia), Rc is not C1-C6 alkyl when L is —SCH2- and Rd is methyl.
In some embodiments of Formula (Ia), Rc is not —CN when L is —SCH2- and Rd is 2- furyl.
In another ment, the compound of Formula (I) is represented by Formula (Ib): [Annotation] KEB or a pharmaceutically acceptable salt thereof wherein: Ra and Rb is en and the other is —(CH2)rC02RX, —OCH2COZRX, rtetrazole, -(CH2)roxadiazolone, -(CH2)rtetrazolone, -(CH2)rthiadiazolol, —(CH2)r isoxazol—3 —ol, —(CH2)rP(O)(OH)ORX, —(CH2)rS(O)ZOH, —(CH2)rC(O)NHCN, or —(CH2)rC(O)NHS(O)2alky1; Rc is H, C1-C6 alkyl, C1-C6 haloalkyl, halogen, —CN, —ORX, —COzRX, or N02; Rd is methyl, optionally substituted 5- to 10-membered aryl, optionally substituted 5- or 6—membered heteroaryl, or optionally substituted 5— or 6—membered carbocycle; each RX is ndently at each occurrence hydrogen or C1—C6 alkyl; each R6 is independently C1—C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, halogen, -ORy, C1-C6 haloalkyl, —NHRZ, -OH, or -CN; each Ry and RZ is independently hydrogen, C1-C6 alkyl, or C1-C6 haloalkyl, and n is 0,1, 2, or 3; with the proviso that Rc is not hydrogen or —CN when and Rd is optionally substituted phenyl, Rc is not C1-C6 alkyl when Rd is methyl, and that R0 is not —CN when Rd is 2-furyl.
In some ments of Formula (Ib), one of Ra and Rb is en and the other is —(CH2)rCOZRX, —OCH2C02RX, -(CH2)rtetrazole, —(CH2)roxadiazolone, —(CH2)rtetrazolone, —(CH2)rthiadiazolol, r isoxazol-3—ol, —(CH2)rP(O)(OH)ORX, rS(O)2OH, —(CH2)rC(O)NHCN, or —(CH2)r C(O)NHS(O)2a1kyl; Rc is C1—C6 alkyl, C1-C6 haloalkyl, halogen, —CN, —ORX, —C02RX, or N02; Rd is methyl, optionally substituted 5— to 10-membered aryl, optionally substituted 5— or 6—membered heteroaryl, or optionally tuted 5— or 6—membered carbocycle; each RX is independently at each occurrence hydrogen or C1—C6 alkyl; [Annotation] KEB each R6 is independently C1-C6 alkyl, C2—C6 alkenyl, C2-C6 alkynyl, halogen, -ORy, C1—C6 haloalkyl, -NHRZ, —OH, or —CN; with the proviso that Rc is not hydrogen or —CN when Rd is optionally substituted , Rc is not C1-C6 alkyl when Rd is methyl, and that Rc is not —CN when Rd is 2—furyl.
In some embodiments of formula (Ib), one of R3 and Rb is hydrogen and the other is COZRX, CHZCOZRX, tetrazole, or oxadiazolone; RC is n, —CN, —OR", or C1-C5 alkyl; Rd is methyl, optionally substituted 5— to lO-membered aryl, optionally substituted 5— or 6-membered heteroaryl, or optionally substituted 5— or 6—membered carbocycle; and RK is hydrogen or C1-C6 alkyl; each R6 is independently C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, halogen, -ORy, C1—C6 haloalkyl, -NHRZ, —OH, or —CN; each Ry and RZ is independently hydrogen, C1-C6 alkyl, or C1-C6 haloalkyl; and n is 0,1, 2, or 3; with the proviso that R0 is not —CN when Rd is optionally substituted phenyl, Rc is not cl—c6 alkyl when Rd is methyl, and that Rc is not —CN when Rd is 2-furyl.
In some ments of Formula (Ib), Ra is —(CH2)rCOZRX, —OCH2C02RX, -(CH2)rtetrazole, —(CH2)roxadiazolone, -(CHz)rtetrazolone, rthiadiazolol, r isoxazol—3 —ol, —(CH2)rP(O)(OH)ORX, —(CH2)rS(O)20H, —(CH2)rC(O)NHCN, or —(CH2)rC(O)NHS(O)2alkyl. In other embodiments, Ra is -(CH2)rCOZRX, -OCH2COZRX, tetrazole, —(CH2)tetrazole, oxadiazolone, oxadiazolone, tetrazolone, -(CH2)tetrazolone, thiadiazolol, -(CH2)thiadiazolol, isoxazolol, —(CH2) isoxazolol, -P(O)(OH)ORX, -(CH2)P(O)(OH)ORX, -S(O)2OH , S(O)2OH, -C(O)NHCN —(CH2)C(O)NHCN, —C(O)NHS(O)2alkyl, or —(CH2)C(O)NHS(O)2alkyl. In other embodiments, Ra is hydrogen, COZR", CHZCOZRX, tetrazole, or oxadiazolone. In further embodiments, R3 is hydrogen, COZH, CHZCOZH, tetrazole, or l,2,4-oxadiazol-5(4H)-one.
In some embodiments of Formula (1b), Rb is —(CH2)rCOZRx, -OCH2C02RX, rtetrazole, -(CH2)roxadiazolone, -(CH2)rtetrazolone, -(CH2)rthiadiazolol, —(CH2)r ol—3 —ol, —(CH2)rP(O)(OH)ORX, IS(O)ZOH, rC(O)NHCN, or —(CH2)rC(O)NHS(O)2alky1. In other embodiments, Rb is -(CH2)rC02RX, -OCH2COZRX, tetrazole, -(CH2)tetrazole, oxadiazolone, -(CH2)oxadiazolone, tetrazolone, -(CH2)tetrazolone, thiadiazolol, -(CH2)thiadiazolol, olol, —(CH2) isoxazolol, [Annotation] KEB —P(O)(OH)ORX, —(CH2)P(O)(OH)OR", -S(O)zOH, -(CH2)S(O)20H, -C(O)NHCN -(CH2)C(O)NHCN, -C(O)NHS(O)2alkyl, or -(CH2)C(O)NHS(O)2alkyl. In other embodiments, Rb is en, COZR", CHZCOZRX, tetrazole, or zolone. In further embodiments, Rb is hydrogen, COZH, CHZCOZH, tetrazole, or 1,2,4—oxadiazol-5(4H)—one. In further embodiments, Rb is hydrogen.
In some embodiments of Formula (Ib), Rc is H, C1-C6 alkyl, C1-C6 haloalkyl, halogen, —CN, —ORX, —COZRX, or N02. In other embodiments, Rc is C1—C6 alkyl, C1—C6 haloalkyl, halogen, —CN, —OR", -C02RX, or N02. In other embodiments, Rc is halogen, —CN, —ORX, or C1-C6 alkyl. In other embodiments, Rc is halogen, —CN, —ORX, or C1-C3 alkyl. In other embodiments, RC is H, —CN, or halogen. In other embodiments, Rc is —CN or n.
In some embodiments of Formula (Ib), Rd is methyl, optionally substituted 5- to 10- membered aryl, optionally substituted 5— or 6-membered heteroaryl, or optionally substituted — or ered carbocycle. In other embodiments, Rd is methyl, optionally cyclohexyl, optionally tuted pyridinyl, optionally substituted thiazolyl, optionally substituted phenyl, or optionally tuted thienyl. In other embodiments, Rd is cyclohexyl, pyridinyl, thiazolyl, phenyl, or thienyl, wherein each is ally substituted with one or more tuents independently selected from halogen, C1—C6 alkyl, C1—C6 hydroxyalkyl, C1—C6 alkoxy, C1-C6 haloalkyl, C1-C6 haloalkoxy, -OH, CN, and amino. In other embodiments, Rd is cyclohexyl, pyridinyl, thiazolyl, phenyl, or thienyl, wherein each is optionally substituted with one or more substituents independently selected from halogen, C1—C6 alkyl, C1—C6 hydroxyalkyl, C1—C6 alkoxy, C1-C6 kyl, and C1-C6 haloalkoxy. In other embodiments, Rd is cyclohexyl, pyridinyl, thiazolyl, phenyl, or thienyl, wherein each is optionally substituted with one or more n. In other embodiments, Rd is methyl, exyl, pyridinyl, thiazolyl, phenyl, or thienyl. In yet other embodiments, Rd is cyclohexyl, pyridinyl, thiazolyl, , or thienyl. In other embodiments, Rd is cyclohexyl, pyridinyl, thiazolyl, phenyl, 4-chlorophenyl, ylphenyl, or thienyl.
In some embodiments of Formula (Ib), each R6 is independently C1-C6 alkyl, C2—C6 alkenyl, C2—C6 alkynyl, halogen, —ORy, C1—C6 haloalkyl, —NHRZ, —OH, or —CN. In other embodiments, C1-C4 alkyl, C2-C4 alkenyl, C2-C4 alkynyl, halogen, -ORy, C1-C4 haloalkyl, - NHRZ, —OH, or —CN.
[Annotation] KEB In some embodiments of Formula (Ib), Rx is en or C1—C6 alkyl. In other embodiments, Rx is hydrogen or C1—C3 alkyl, In further embodiments, Rx is hydrogen, methyl, ethyl, n-propyl, or iso—propyl.
In some embodiments of Formula (Ib), Ry is independently hydrogen, C1-C6 alkyl, or C1-C6 haloalkyl. In other embodiments, Ry is hydrogen, C1-C3 alkyl, or C1-C3 haloalkyl.
In some embodiments of Formula (Ib), each R2 is independently hydrogen, C1—C6 alkyl, or C1—C6 haloalkyl. In other embodiments, RZ is hydrogen, C1—C3 alkyl, or C1—C3 haloalkyl.
In some embodiments of Formula (Ib), n is 0, l, 2, or 3. In other embodiments, n is 0 or 1. In further embodiments, n is 0.
In some embodiments of a (Ib), one of Ra and Rb is hydrogen and the other is COzRX, CH2C02RX, ole, or oxadiazolone. In other embodiments, Rb is hydrogen and R3 is CHZCOZH, tetrazole, or (l,2,4-oxadiazol-5(4H)-one).
In some embodiments of Formula (Ib), Rb is en, RC is —CN, Rd is thienyl, and Ra is CH2C02H, tetrazole, or -oxadiazol-5(4H)-one).
In some embodiments of Formula (Ib), Rc is halogen, R3 is —C02H, and Rb is H. In other embodiments, Rc is -Br, Ra is —COzH, and Rb is H. In further embodiments, Rc is -Cl, Ra is —C02H, and Rb is H.
In some embodiments of a (Ib), Rc is n, Ra is tetrazole, and Rb is H. In other embodiments, Rc is —Br, R3 is tetrazole, and Rb is H. In further embodiments, RC is -C1, Ra is tetrazole, and Rb is H.
In some embodiments of Formula (Ib), Rc is halogen, R3 is —CH2COzH, and Rb is H.
In other embodiments, Rc is -Br, R3 is —CH2C02H, and Rh is H. In r embodiments, Rc is —C1, Ra is —CH2COZH, and RI) is H.
In some embodiments of Formula (Ib), Rc is halogen, Ra is (l,2,4-oxadiazol-5(4H)- one), and Rb is H. In other ments, Rc is —Br, Ra is (1,2,4-oxadiazol-5(4H)—one), and Rb is H. In other embodiments, Rc is -Cl, Ra is (l,2,4-oxadiazol-5(4H)—one), and Rb is H.
In some embodiments of Formula (Ib), Rc is -CN, R3 is —C02H, and Rb is H. In other embodiments, Rc is —CN, R21 is —CH2C02H, and Rb is H. In other embodiments, Rc is —CN, Ra is tetrazole, and Rb is H. In yet other embodiments, Rc is —CN, Ra is (l,2,4-oxadiazol-5(4H)— one), and Rb is H.
[Annotation] KEB In some embodiments of Formula (Ib), Rc is not hydrogen or —CN and Rd is ally substituted phenyl. In other embodiments, RC is not C1—C6 alkyl and Rd is methyl.
In other embodiments, Rc is not —CN and Rd is 2-furyl.
In some embodiments of Formula (Ib), Rc is not hydrogen or —CN when and Rd is optionally substituted phenyl.
In some embodiments of Formula (Ib), Rc is not C1-C6 alkyl when Rd is methyl.
In some embodiments of Formula (Ib), Rc is not —CN when Rd is 2-furyl.
In another embodiment, the compound of Formula (I) is represented by Formula (II): A COZH or a ceutically acceptable salt thereof, wherein: Rc is halogen, —CN, —ORX, or C1-C6 alkyl; Rd is methyl, optionally substituted 5— to 10-membered aryl, optionally substituted 5— or 6—membered aryl, or optionally tuted 5— or 6—membered carbocycle; and RK is hydrogen or C1-C6 alkyl with the proviso that R0 is not —CN when and Rd is optionally substituted phenyl, Rc is not C1—C6 alkyl when Rd is methyl, and that R0 is not —CN when Rd is 2-furyl.
In some embodiments of Formula (II), Rc is halogen, —CN, —ORX, or C1-C6 alkyl; Rd is methyl, optionally substituted 5— to lO-membered aryl, ally substituted 5— or 6— membered heteroaryl, or optionally tuted 5— or 6—membered carbocycle; and RK is hydrogen or C1-C6 alkyl with the proviso that R0 is not C1-C6 alkyl when Rd is methyl, and that RC is not —CN when Rd is 2—fury1.
In some embodiments of Formula (II), Rc is n, —CN, —ORX, or C1-C6 alkyl. In other embodiments, Rc is halogen, —CN, —OR", or C1-C3 alkyl. In further embodiments, Rc is —CN or halogen.
In some ments of Formula (II), Rd is methyl, optionally substituted 5- to 10- membered aryl, optionally substituted 5— or 6—membered aryl, or optionally substituted [Annotation] KEB — or 6—membered carbocycle. In other embodiments, Rd is cyclohexyl, pyridinyl, thiazolyl, phenyl, or thienyl, wherein each is optionally substituted with one or more substituents independently selected from n, C1-C6 alkyl, C1-C6 yalkyl, C1-C6 alkoxy, C1-C6 haloalkyl, C1-C6 haloalkoxy, -OH, CN, and amino. In other embodiments, Rd is cyclohexyl, pyridinyl, lyl, phenyl, or thienyl, wherein each is optionally substituted with one or more substituents independently ed from halogen, C1-C5 alkyl, C1-C5 hydroxyalkyl, C1- C6 alkoxy, C1-C6 haloalkyl, and C1-C6 haloalkoxy. In other embodiments, Rd is cyclohexyl, nyl, thiazolyl, phenyl, or thienyl, wherein each is optionally substituted with one or more halogen. In further embodiments, Rd is methyl, cyclohexyl, pyridinyl, thiazolyl, phenyl, or thienyl.
In some embodiments of Formula (II), Rb is en, COZR", CHZCOZRX, tetrazole, or oxadiazolone. In other ments, Rb is hydrogen, COZH, CH2C02H, tetrazole, or 1,2,4- oxadiazol—5(4H)—one. In further embodiments, Rb is hydrogen.
In some embodiments of Formula (II), R3 is hydrogen, COZRX, CHZCOZRX, tetrazole, or oxadiazolone. In further embodiments, Ra is hydrogen, COZH, CH2C02H, ole, or l,2,4—oxadiazol—5(4H)—one.
In some embodiments of Formula (II), each R6 is independently C1-C6 alkyl, C2—C6 alkenyl, C2-C6 alkynyl, halogen, -ORy, C1-C6 haloalkyl, —NHRZ, —OH, or —CN.
In some embodiments of Formula (II), each Ry is independently hydrogen, C1-C6 alkyl, or C1-C6 haloalkyl.
In some embodiments of Formula (II), each RZ is independently hydrogen, C1-C6 alkyl, or C1-C6 haloalkyl.
In some embodiments of Formula (II), n is 0, l, 2, or 3. In other ments, n is 0 or 1. In further embodiments, n is 0.
In some embodiments of Formula (II), one of R3 and Rb is hydrogen and the other is COzR", CHZCOZRX, tetrazole, or oxadiazolone. In other embodiments, Rb is hydrogen and Ra is CHZCOZH, tetrazole, or (l,2,4-oxadiazol-5(4H)-one).
In some embodiments of Formula (II), Rb is en, Rc is —CN, Rd is thienyl, and Ra is CHZCOZH, tetrazole, or -oxadiazol-5(4H)-one).
In some embodiments of Formula (II), R0 is halogen, Ra is —COZH, and Rb is H. In other embodiments, Rc is -Br, R3 is —C02H, and Rb is H. In further embodiments, Rc is -C1, Ra is —COZH, and Rb is H.
[Annotation] KEB In some ments of Formula (II), Rc is halogen, Ra is tetrazole, and Rb is H. In other embodiments, RC is —Br, R3 is tetrazole, and Rb is H. In further embodiments, RC is -Cl, Ra is tetrazole, and Rb is H.
In some embodiments of a (II), Rc is halogen, R3 is —CH2C02H, and Rb is H.
In other embodiments, Rc is -Br, R3 is —CH2COZH, and Rb is H. In further embodiments, Rc is —c1, R3 is —CH2C02H, and Rb is H.
In some embodiments of Formula (II), RC is halogen, Ra is (l,2,4-oxadiazol-5(4H)- one), and Rb is H. In other embodiments, Rc is —Br, Ra is (l,2,4-oxadiazol-5(4H)—one), and Rb is H. In other embodiments, Rc is -Cl, R81 is (l,2,4-oxadiazol-5(4H)—one), and Rb is H.
In some embodiments of Formula (II), Rc is -CN, Ra is —COzH, and Rb is H. In other embodiments, Rc is —CN, R21 is —CH2C02H, and Rb is H. In other embodiments, RC is —CN, R3 is tetrazole, and Rb is H. In yet other embodiments, Rc is -CN, Ra is (l,2,4-oxadiazol-5(4H)— one), and Rb is H.
In another embodiment, the compound of Formula (I) is represented by Formula (III): | x (Ron / /\\ Ra or a pharmaceutically able salt thereof wherein: Ra and Rb is hydrogen and the other is —(CH2)rC02RX, —OCH2COZRX, -(CH2)rtetrazole, -(CH2)roxadiazolone, -(CH2)rtetrazolone, -(CH2)rthiadiazolol, -(CH2)r isoxazol—3—ol, —(CH2)rP(O)(OH)ORX, -(CH2)rS(O)20H, -(CHz)rC(O)NHCN, or -(CH2)rC(O)NHS(O)2alky1; Rc is H, C1-C6 alkyl, C1-C6 haloalkyl, halogen, —CN, —ORX, , or N02; Rd is methyl, ally substituted 5— to bered aryl, optionally tuted 5— or 6—membered heteroaryl, or optionally substituted 5— or 6—membered ycle; each Rx is independently at each occurrence hydrogen or C1-C6 alkyl; each R6 is independently C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, halogen, -ORy, C1-C6 haloalkyl, —NHRZ, -OH, or -CN; each Ry and RZ is independently hydrogen, C1-C6 alkyl, or C1-C6 haloalkyl, and [Annotation] KEB n is 0,1, 2, or 3.
In some embodiments of Formula (III), one of Ra and Rb is hydrogen and the other is —(CH2)rCOZRX, —OCH2C02RX, -(CH2)rtetrazole, -(CH2)roxadiazolone, -(CH2)rtetrazolone, -(CH2)rthiadiazolol, —(CH2)r isoxazol—3 —ol, —(CH2)rP(O)(OH)ORX, —(CH2)rS(O)ZOH, —(CH2)rC(O)NHCN, or -(CH2)r C(O)NHS(O)2alkyl; Rc is C1-C6 alkyl, C1-C6 haloalkyl, halogen, —CN, —ORX, —C02RX, or N02; Rd is , optionally substituted 5- to lO-membered aryl, optionally substituted 5— or 6—membered heteroaryl, or ally substituted 5— or 6—membered carbocycle; each RX is independently at each occurrence en or C1—C6 alkyl; each R6 is independently C1—C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, halogen, —ORy, C1-C6 kyl, -NHRZ, —OH, or —CN.
In some ments of formula (III), one of Ra and Rb is hydrogen and the other is COZRX, CHZCOZRX, tetrazole, or oxadiazolone; Rc is halogen, —CN, —ORX, or C1-C6 alkyl; Rd is methyl, optionally substituted 5- to bered aryl, optionally substituted 5— or 6—membered heteroaryl, or ally substituted 5— or 6—membered carbocycle; and RX is hydrogen or C1-C6 alkyl; each R6 is independently C1—C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, halogen, —ORy, C1—C6 haloalkyl, -NHRZ, -OH, or —CN; each Ry and R2 is independently en, C1-C6 alkyl, or C1-C6 haloalkyl; and n is 0,1, 2, or 3; with the o that R0 is not hydrogen or —CN when Rd is optionally substituted phenyl and that Rc is not —CN when Rd is 2-furyl.
In some embodiments of Formula (III), Ra is rCOZRX, —OCH2COZRX, -(CH2)rtetrazole, -(CH2)roxadiazolone, -(CH2)rtetrazolone, -(CH2)rthiadiazolol, —(CH2)r isoxazol—3 —ol, —(CH2)rP(O)(OH)ORX, —(CH2)rS(O)zOH, —(CH2)rC(O)NHCN, or —(CH2)rC(O)NHS(O)2alky1. In other embodiments, R8 is -(CH2)rC02RX, -OCH2COZRX, tetrazole, -(CH2)tetrazole, oxadiazolone, -(CH2)oxadiazolone, tetrazolone, -(CH2)tetrazolone, thiadiazolol, -(CH2)thiadiazolol, isoxazolol, -(CH2) isoxazolol, —P(O)(OH)ORX, —(CH2)P(O)(OH)ORX, —S(O)20H , —(CH2)S(O)ZOH, —C(O)NHCN [Annotation] KEB -(CH2)C(O)NHCN, -C(O)NHS(O)zalkyl, or -(CH2)C(O)NHS(O)2alkyl. In other embodiments, Ra is hydrogen, COZR", CH2C02RX, tetrazole, or oxadiazolone. In further embodiments, Ra is en, COZH, CHZCOZH, tetrazole, or 1,2,4—oxadiazol—5(4H)—one.
In some embodiments of Formula (III), Rb is -(CH2)rCOZRX, -OCH2C02RX, rtetrazole, -(CH2)roxadiazolone, -(CH2)rtetrazolone, -(CH2)rthiadiazolol, —(CH2)r isoxazol—3 -ol, —(CH2)rP(O)(OH)ORx, —(CH2)rS(O)2OH, —(CH2)rC(O)NHCN, or —(CH2)rC(O)NHS(O)2alkyl. In other embodiments, Rb is -(CH2)rCOZRX, -OCH2C02R", tetrazole, tetrazole, oxadiazolone, -(CH2)oxadiazolone, tetrazolone, -(CH2)tetrazolone, thiadiazolol, -(CH2)thiadiazolol, olol, —(CH2) isoxazolol, —P(O)(OH)ORX, —(CH2)P(O)(OH)ORX, —S(O)zOH , —(CH2)S(O)20H, —C(O)NHCN —(CH2)C(O)NHCN, —C(O)NHS(O)2alkyl, or —(CH2)C(O)NHS(O)2alkyl. In other embodiments, Rb is hydrogen, COZR", CHZCOZRX, tetrazole, or zolone. In further embodiments, Rb is hydrogen, COZH, CHZCOZH, ole, or 1,2,4-oxadiazol-5(4H)—one. In r embodiments, Rb is hydrogen.
In some embodiments of Formula (III), Rc is H, C1-C6 alkyl, C1-C6 haloalkyl, halogen, —CN, —ORX, —COzRX, or N02. In other embodiments, Rc is C1—C6 alkyl, C1—C6 haloalkyl, halogen, —CN, —ORX, —C02RX, or N02. In other embodiments, RC is halogen, —CN, —OR", or C1-C6 alkyl. In other embodiments, RC is halogen, —CN, —ORX, or C1-C3 alkyl. In other embodiments, RC is H, —CN, or n. In other embodiments, Rc is —CN or n.
In some embodiments of Formula (III), Rd is methyl, optionally substituted 5- to 10- membered aryl, optionally substituted 5- or 6-membered heteroaryl, or optionally substituted — or 6—membered carbocycle. In other embodiments, Rd is methyl, optionally cyclohexyl, optionally substituted pyridinyl, optionally substituted thiazolyl, optionally substituted phenyl, or optionally substituted thienyl. In other ments, Rd is cyclohexyl, pyridinyl, thiazolyl, phenyl, or thienyl, n each is optionally tuted with one or more substituents independently ed from halogen, C1—C6 alkyl, C1—C6 hydroxyalkyl, C1—C6 alkoxy, C1-C6 haloalkyl, C1-C6 haloalkoxy, -OH, CN, and amino. In other embodiments, Rd is cyclohexyl, pyridinyl, thiazolyl, phenyl, or thienyl, wherein each is optionally substituted with one or more substituents independently selected from halogen, C1—C6 alkyl, C1—C6 hydroxyalkyl, C1—C6 alkoxy, C1—C6 haloalkyl, and C1—C6 haloalkoxy. In other embodiments, Rd is cyclohexyl, pyridinyl, thiazolyl, phenyl, or thienyl, wherein each is optionally substituted with one or more n. In other embodiments, Rd is methyl, cyclohexyl, nyl, thiazolyl, phenyl, or l. In yet other embodiments, Rd is cyclohexyl, [Annotation] KEB nyl, thiazolyl, phenyl, or thienyl. In other embodiments, Rd is cyclohexyl, pyridinyl, lyl, phenyl, 4-chlorophenyl, 4—methylphenyl, or thienyl.
In some embodiments of Formula (III), each R6 is ndently C1-C6 alkyl, C2—C6 alkenyl, C2-C6 alkynyl, halogen, -ORy, C1-C6 haloalkyl, —NHRZ, -OH, or —CN. In other embodiments, C1-C4 alkyl, C2-C4 alkenyl, C2-C4 alkynyl, halogen, —ORy, C1-C4 haloalkyl, - NHRZ, -OH, or —CN.
In some embodiments of Formula (III), RX is en or C1-C6 alkyl. In other embodiments, Rx is hydrogen or C1—C3 alkyl. In further embodiments, Rx is hydrogen, methyl, ethyl, n-propyl, or iso—propyl.
In some embodiments of Formula (III), Ry is independently hydrogen, C1-C6 alkyl, or C1-C6 haloalkyl. In other embodiments, Ry is hydrogen, C1-C3 alkyl, or C1-C3 haloalkyl.
In some embodiments of Formula (III), each R2 is independently hydrogen, C1—C6 alkyl, or C1—C6 kyl. In other embodiments, RZ is hydrogen, C1—C3 alkyl, or C1—C3 haloalkyl.
In some embodiments of Formula (III), n is 0, l, 2, or 3. In other embodiments, n is 0 or 1. In further embodiments, n is 0.
In some embodiments of Formula (III), one of Ra and Rb is hydrogen and the other is (:0sz, CH2C02RX, tetrazole, or oxadiazolone. In other embodiments, Rb is hydrogen and R8 is CH2C02H, tetrazole, or (l,2,4-oxadiazol-5(4H)-one).
In some embodiments of Formula (111), Rb is hydrogen, RC is —CN, Rd is thienyl, and Ra is CHZCOZH, tetrazole, or (l,2,4-oxadiazol-5(4H)-one).
In some embodiments of Formula (III), Rc is halogen, Ra is —COzH, and Rb is H. In other ments, Rc is -Br, Ra is —COzH, and Rb is H. In further embodiments, Rc is -Cl, Ra is —C02H, and Rb is H.
In some embodiments of Formula (III), R0 is halogen, Ra is tetrazole, and Rb is H. In other embodiments, Rc is —Br, Ra is tetrazole, and Rb is H. In further ments, RC is —Cl, Ra is tetrazole, and Rb is H.
In some embodiments of Formula (111), RC is halogen, Ra is —CH2COZH, and R13 is H.
In other embodiments, Rc is -Br, R3 is —CH2C02H, and Rb is H. In further embodiments, Rc is -Cl, Ra is —CH2C02H, and Rb is H.
[Annotation] KEB In some embodiments of Formula (III), R0 is halogen, Ra is (l,2,4-oxadiazol-5(4H)- one), and Rb is H. In other embodiments, RC is -Br, R3 is (l,2,4-oxadiazol-5(4H)—one), and Rb is H. In other embodiments, Rc is -Cl, Ra is (l,2,4-oxadiazol-5(4H)—one), and Rb is H.
In some embodiments of a (III), Rc is -CN, R8 is —C02H, and Rb is H. In other embodiments, Rc is —CN, Ra is —CH2C02H, and Rb is H. In other embodiments, Rc is —CN, Ra is tetrazole, and Rb is H. In yet other ments, RC is -CN, R3 is (l,2,4—oxadiazol—5(4H)— one), and Rb is H.
In some embodiments of Formula (III), Rc is not hydrogen or —CN when Rd is optionally substituted phenyl, Rc is not C1-C6 alkyl when Rd is methyl, and that R0 is not —CN when Rd is 2-furyl.
In some embodiments of Formula (I), (la), (Ib), (II) and (III), one of Ra or Rb is a carboxylic acid or a carboxylic acid bioisostere.
In some embodiments of Formula (I), (la), (Ib), (II) and (III), Ra is -COzH, —(CH2)C02H, or —OCH2C02H. In other embodiments, Ra is —COzCH3, —COzCH2CH3, —COzCH2CH2CH3, (CH3)2, —(CH2)C02CH3, —(CH2)COzCH2CH3, —(CH2)COZCH2CH2CH3, or —(CH2)COZCH(CH3)2.
In some embodiments of a (I), (Ia), (Ib), (II) and (III), R8 is -P(O)(OH)OH, —(CH2)P(O)(OH)OH, —P(O)(OH)OCH3, —P(O)(OH)OCH2CH3, —P(O)(OH)OCH2CH2CH3, —P(O)(OH)OCH(CH3)2, —(CH2) P(O)(OH)OCH3, -(CH2)P(O)(OH)OCH2CH3, —(CH2)P(O)(OH)OCH2CH2CH3, or —(CH2)P(O)(OH)OCH(CH3)2.
In some embodiments of Formula (I), (la), (Ib), (II) and (III), R8 is -S(O)20H, —(CH2)S(O)ZOH, —C(O)NHCN, or C(O)NHCN.
In some embodiments of Formula (I), (la), (Ib), (II) and (III), Ra is —C(O)NHS(O)2CH3, —C(O)NHS(O)2CH2CH3, HS(O)2CH2CH2CH3, —C(O)NHS(O)2CH(CH3)2, C(O)NHS(O)2CH3, —(CH2)C(O)NHS(O)2CH2CH3, —(CH2)C(O)NHS(O)2CH2CH2CH3, or -(CH2)C(O)NHS(O)2CH(CH3)2.
In some embodiments of Formula (I), (la), (Ib), (II) and (III), Ra is OH OH 0H x OH OH 0H \ x"((4 g):N \ I , '?‘N‘NH ,N ‘s N | ‘N 3" \ \ | N J5» \ o | N \ , ‘5; ‘6 5:5; , , | N | N o, N 3 0 0 8’ § o, 5 5 5 9 3 [Annotation] KEB N OH iH\O/\EN\s\ I \J \ N E:«\NH f/[gqs’ ,§\E\(I N ‘ f 0 3’ , ,or , .
In some embodiments of a (I), (Ia), (Ib), (II), and (III), R8 is OH OH (KEN HO N’ HO N’ \ \ N ,{NKNH ?N \J:N:3 a \yN‘NH o f O f N or O 7 7 7 7 7 ' In some embodiments of Formula (I), (la), (Ib), (II) and (III), Rb is -COZH, COZH, or —OCH2C02H. In other embodiments, R1) is —COzCH3, 2CH3, —C02CH2CH2CH3, —COzCH(CH3)2, —(CH2)C02CH3, —(CH2)COZCH2CH3, —(CH2)COZCH2CH2CH3, or —(CH2)COZCH(CH3)2.
In some embodiments of Formula (I), (Ia), (Ib), (II) and (III), Rb is —P(O)(OH)OH, —(CH2)P(O)(OH)OH, —P(O)(OH)OCH3, —P(O)(OH)OCH2CH3, —P(O)(OH)OCH2CH2CH3, —P(O)(OH)OCH(CH3)2, —(CH2) P(O)(OH)OCH3, -(CH2)P(O)(OH)OCH2CH3, —(CH2)P(O)(OH)OCH2CH2CH3, or —(CH2)P(O)(OH)OCH(CH3)2.
In some embodiments of Formula (I), (la), (Ib), (II) and (III), Rb is -S(O)20H, —(CH2)S(O)ZOH, —C(O)NHCN, or —(CH2)C(O)NHCN.
In some ments of Formula (I), (la), (Ib), (II) and (III), Rb is —C(O)NHS(O)2CH3, —C(O)NHS(O)2CH2CH3, —C(O)NHS(O)2CH2CH2CH3, —C(O)NHS(O)2CH(CH3)2, C(O)NHS(O)2CH3, —(CH2)C(O)NHS(O)2CH2CH3, —(CH2)C(O)NHS(O)2CH2CH2CH3, or -(CH2)C(O)NHS(O)2CH(CH3)2.
In some embodiments of Formula (I), (la), (Ib), (II) and (III), Rb is OH 0" OH ([‘(N ‘féOH OH OH HO N;N‘ ’N‘s NKNH | ‘N :3 \ | ‘N ’2 | \ ' N \ , N w , N g N 0’ a | | 3 s, o 0’ 7 7 7 7A 7 7 :N OH ’N‘ E \NH I \N ’g \ :95; \le \g/ X \ S, I IN 0 or S , , , .
In some embodiments of Formula (I), (la), (Ib), (II), and (III), Rb is ation] KEB OH CH HO =N\ =N\ IN 3 NH | \N ~s I \ I Y ff»:I «’1;"0):": O 1H. N O ‘ N 3/3‘6"" O or 0 7 7 7 , , .
In some embodiments, the compound of Formula (I) is a compound having any one of the following Formulae: [Annotation] KEB or a pharmaceutically acceptable salt f.
The above de?nition of the compounds of Formula (1) is referred to herein by the expressions "compound of Formula (1)" as de?ned herein, or simply "compounds of Formula (1)", etc. The above de?nition of the nds of Formula (Ia) is ed to herein by the expressions "compound of a (1a)" as de?ned herein, or simply unds of Formula (1a)", etc. The above de?nition of the compounds of Formula (1b) is referred to herein by the expressions "compound of Formula (1b)" as de?ned herein, or simply unds of Formula (Ib)", etc. The above de?nition of the compounds of Formula (11) is referred to herein by the expressions "compound of Formula (11)" as de?ned herein, or simply "compounds of a (11)", etc. The above de?nition of the compounds of Formula (111) is referred to herein by the expressions und of Formula (111)" as de?ned herein, or simply "compounds of a (111)", etc. It should be tood, that such references are intended to encompass not only the above general formula, but also each and every of the embodiments, etc. discussed in the following. It should also be understood, that unless stated to the opposite, such references also encompass isomers, mixtures of isomers, pharmaceutically acceptable salts, solvates and prodrugs of the compounds of Formula (1), Formula (1a), Formula (1b), a (11), and Formula (111).
De?nitions The term "alkyl" as used herein refers to a saturated, straight or branched hydrocarbon chain. The hydrocarbon chain preferably contains from one to eight carbon atoms (CH;— alkyl), more preferred from one to six carbon atoms (Cm—alkyl), in particular from one to four carbon atoms (CM-alkyl), including methyl, ethyl, propyl, isopropyl, butyl, isobutyl, secondary butyl, tertiary butyl, pentyl, isopentyl, neopentyl, tertiary pentyl, hexyl, isohexyl, heptyl and octyl. In a preferred embodiment "alkyl" represents a C1_4-alkyl group, which may in particular include methyl, ethyl, propyl, isopropyl, butyl, yl, secondary butyl, and tertiary butyl. Correspondingly, the term "alkylene" means the corresponding biradical (—alkyl—).
The term "cycloalkyl" or "carbocycle" as used herein refers to a cyclic alkyl group, preferably containing from three to ten carbon atoms (C3_10-cycloalkyl or C3_10—carbocycle), [Annotation] KEB such as from three to eight carbon atoms (CH-cycloalkyl or C3_10-carbocycle), preferably from three to six carbon atoms (CM—cycloalkyl or carbocycle), including ropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl and cyclooctyl. Furthermore, the term "cycloalkyl" as used herein may also include clic groups such as for example bicyclo[2.2.2]octyl, bicyclo[2.2. anyl, decalinyl and adamantyl. Correspondingly, the term "cycloalkylene" means the corresponding biradical (-cycloalkyl-). Alkyl and cycloalkyl groups may be optionally substituted with 1—4 substituents. Examples of substituents on alkyl groups include, but are not limited to, alkyl, l, alkynyl, halogen, haloalkyl, alkoxy, heteroaryl, aryl, carbocyclyl, hydroxyl, carbamoyl, oxo, and —CN.
The term "alkenyl" as used herein refers to a straight or branched hydrocarbon chain or cyclic hydrocarbons containing one or more double bonds, including di—enes, tri—enes and poly-enes. Typically, the l group comprises from two to eight carbon atoms (C24;— alkenyl), such as from two to six carbon atoms (CM—alkenyl), in particular from two to four carbon atoms (CM-alkenyl), including at least one double bond. Examples of alkenyl groups e ethenyl; l— or 2-propenyl; 1-, 2— or 3-butenyl, or 1,3—but-dienyl; 1-, 2—, 3—, 4- or 5— l, or l,3-hex-dienyl, or hex-trienyl; l-, 2-, 3—, 4-, 5-, 6—, or 7-octenyl, or l,3— enyl, or 1,3,5-octatrienyl, or l,3,5,7-octatetraenyl, or cyclohexenyl. Correspondingly, the term "alkenylene" means the corresponding biradical (-alkenyl—). Alkenyl groups may be optionally substituted with 1-4 substituents. Examples of substituents on alkenyl groups include, but are not limited to, alkyl, alkenyl, l, halogen, haloalkyl, alkoxy, heteroaryl, aryl, carbocyclyl, hydroxyl, carbamoyl, oxo, and —CN.
The term "alkynyl" as used herein refers to a straight or branched hydrocarbon chain containing one or more triple bonds, including di-ynes, tri-ynes and nes. Typically, the alkynyl group comprises of from two to eight carbon atoms (Cm-alkynyl), such as from two to six carbon atoms (CM-alkynyl), in particular from two to four carbon atoms (CH-alkynyl), ing at least one triple bond. Examples of preferred alkynyl groups include ethynyl; l- or 2-propynyl; 1—, 2- or 3—butynyl, or l,3-but-diynyl; 1-, 2-, 3-, 4- or 5-hexynyl, or l,3—hex- diynyl, or 1,3,5-hex—triynyl; 1—, 2—, 3—, 4-, 5—, 6—, or 7—octynyl, or l,3—oct—diynyl, or 1,3,5—oct— triynyl, or l,3,5,7-oct-tetraynyl. Correspondingly, the term ylene" means the corresponding biradical (-alkynyl-). Alkynyl groups may be optionally substituted with 1-4 substituents. Examples of substituents on alkynyl groups include, but are not limited to,, alkyl, alkenyl, alkynyl, halogen, haloalkyl, alkoxy, heteroaryl, aryl, carbocyclyl, hydroxyl, carbamoyl, oxo, and —CN.
[Annotation] KEB The terms "halo" and "halogen" as used herein refer to ?uoro, chloro, bromo or iodo.
Thus a trihalomethyl group represents, 6.3., a tri?uoromethyl group, or a trichloromethyl group. Preferably, the terms "halo" and "halogen" designate ?uoro or Chloro.
The term "haloalkyl" as used herein refers to an alkyl group, as de?ned herein, which is substituted one or more times with one or more n. Examples of haloalkyl groups include, but are not limited to, tri?uoromethyl, di?uoromethyl, penta?uoroethyl, trichloromethyl, etc.
The term "alkoxy" as used herein refers to an "alkyl-O-" group, wherein alkyl is as de?ned above.
The term "hydroxyalkyl" as used herein refers to an alkyl group (as defined hereinabove), which alkyl group is substituted one or more times with hydroxy. Examples of hydroxyalkyl groups e HO—CH2-, -CHZ- and CH3—CH(OH)—.
The term "oxy" as used herein refers to an "—0—" group.
The term "oxo" as used herein refers to an "=0" group.
The term "amine" as used herein refers to primary (R-NHz, R :2 H), ary ((R)2- NH, (R)2 :2 H) and tertiary ((R)3-N, R 72 H) amines. A substituted amine is intended to mean an amine where at least one of the hydrogen atoms has been replaced by the tuent.
The term "carbamoyl" as used herein refers to a "H2N(C=O)—" group.
The term "aryl", as used , unless otherwise indicated, includes carbocyclic aromatic ring systems derived from an aromatic hydrocarbon by removal of a hydrogen atom.
Aryl furthermore es bi-, tri- and clic ring systems. Examples of preferred aryl moieties include phenyl, naphthyl, indenyl, indanyl, ?uorenyl, biphenyl, indenyl, naphthyl, anthracenyl, phenanthrenyl, pentalenyl, azulenyl, and biphenylenyl. Preferred "aryl" is phenyl, naphthyl or indanyl, in particular phenyl, unless otherwise stated. Any aryl used may be optionally substituted. Correspondingly, the term "arylene" means the corresponding cal (-aryl-). Aryl groups may be optionally substituted with 1—4 substituents. Examples of substituents on aryl groups include, but are not limited to, alkyl, alkenyl, alkynyl, halogen, kyl, alkoxy, heteroaryl, aryl, carbocyclyl, hydroxyl, and —CN.
The term "heteroaryl", as used herein, refers to aromatic groups containing one or more heteroatoms selected from O, S, and N, ably from one to four heteroatoms, and more preferably from one to three heteroatoms. Heteroaryl rmore includes bi—, tri— and polycyclic groups, wherein at least one ring of the group is aromatic, and at least one of the rings contains a heteroatom selected from O, S, and N. Heteroaryl also include ring systems [Annotation] KEB substituted with one or more oxo moieties. Examples of preferred heteroaryl moieties include N—hydroxytetrazolyl, oxytriazolyl, N—hydroxyimidazolyl, furanyl, triazolyl, pyranyl, thiadiazinyl, benzothiophenyl, dihydro-benzo[b]thiophenyl, xanthenyl, isoindanyl, acridinyl, benzisoxazolyl, quinolinyl, isoquinolinyl, phteridinyl, azepinyl, diazepinyl, imidazolyl, thiazolyl, carbazolyl, nyl, pyridazinyl, pyrimidinyl, pyrazolyl, pyrazinyl, tetrazolyl, fury], thienyl, olyl, oxazolyl, isothiazolyl, pyrrolyl, indolyl, benzimidazolyl, benzofuranyl, cinnolinyl, indazolyl, indolizinyl, phthalazinyl, triazinyl, isoindolyl, purinyl, zolyl, thiadiazolyl, furazanyl, benzofurazanyl, benzothiophenyl, benzotriazolyl, benzothiazolyl, benzoxazolyl, quinazolinyl, quinoxalinyl, naphthyridinyl, dihydroquinolyl, tetrahydroquinolyl, dihydroisoquinolyl, tetrahydroisoquinolyl, benzofuryl, furopyridinyl, pyrolopyrimidinyl, azaindolyl, pyrazolinyl, l,2,4-oxadiazol-5(4H)—one, and pyrazolidinyl.
Non-limiting examples of partially hydrogenated derivatives are 1,2,3,4-tetrahydronaphthyl, hydronaphthyl, and l-octalin. Correspondingly, the term "heteroarylene" means the corresponding biradical (—heteroaryl-). Heteroaryl groups may be optionally substituted with 1—4 tuents. es of substituents on heteroaryl groups include, but are not limited to, alkyl, alkenyl, alkynyl, halogen, haloalkyl, alkoxy, heteroaryl, aryl, carbocyclyl, hydroxyl, and -CN.
The term "heterocyclyl" as used herein, refers to cyclic omatic groups containing one or more heteroatoms selected from O, S, and N, preferably from one to four heteroatoms, and more preferably from one to three heteroatoms. Heterocyclyl furthermore includes bi-, tri- and polycyclic non-aromatic groups, and at least one of the rings ns a atom selected from O, S, and N. Heterocyclyl also include ring systems substituted With one or more oxo moieties. es of heterocyclic groups are e, pyrrolidinyl, pyrrolyl, 3H—pyrrolyl, oxolanyl, furanyl, thiolanyl, thiophenyl, pyrazolyl, lidinyl, imidazolyl, imidazolidinyl, azolyl, 1,2-oxazolyl, 1,3-oxazolyl, 1,2-thiazolyl, 1,3— thiazolyl, oxadiazolyl, piperidinyl, pyridinyl, oxanyl, 2-H-pyranyl, 4-H—pyranyl, thianyl, 2H-thiopyranyl, pyridazinyl, l,2-diazinanyl, pyrimidinyl, 1,3-diazinanyl, pyrazinyl, piperazinyl, l,4-dioxinyl, 1,4-dioxanyl, l,3-diazinanyl, 1,4-oxazinyl, morpholinyl, thiomorpholinyl, 1,4-oxathianyl, benzofuranyl, isobenzofuranyl, indazolyl, benzimidazolyl, quinolinyl, isoquinolinyl, chromayl, isochromanyl, 4H-chromenyl, lH-isochromenyl, cinnolinyl, quinazolinyl, quinoxalinyl, phthalazinyl, purinyl, yridinyl, pteridinyl, indolizinyl, lH-pyrrolizinyl, 4H-quinolizinyl and bicyclo[3.2.l]octane.
Correspondingly, the term "heterocyclylene" means the corresponding biradical (—heterocyclyl—). Heterocyclyl groups may be optionally tuted with 1—4 substituents.
[Annotation] KEB Examples of substituents on heterocyclyl groups include, but are not limited, to alkyl, alkenyl, alkynyl, halogen, haloalkyl, , heteroaryl, aryl, carbocyclyl, hydroxyl, and —CN.
The term "N-heterocyclic ring" as used herein, refers to a heterocyclyl or a heteroaryl, as de?ned hereinabove, having at least one nitrogen atom, and being bound via a nitrogen atom. es of such N—heterocyclic rings are pyrrolidinyl, pyrrolyl, 3H-pyrrolyl, pyrazolyl, pyrazolidinyl, imidazolyl, imidazolidinyl, 3H-pyrazolyl, 1,2-oxazolyl, 1,2- thiazolyl, 1,3-thiazolyl, piperidinyl, pyridinyl, pyridazinyl, pyrazinyl, piperazinyl, morpholinyl, pyridinyl, pyridazinyl, pyrimidinyl, lyl, pyrazinyl, tetrazolyl, etc.
Isomers In the present speci?cation, the structural formula of the nd represents a certain isomer for convenience in some cases, but the present disclosure includes all isomers, such as geometrical isomers, optical isomers based on an asymmetrical carbon, stereoisomers, tautomers, and the like. Accordingly, it should be understood that the definition of compounds of Formulae (I), (la), (lb), (11) and (111) include each and every dual isomer corresponding to the Formula: Formulae (I), (la), (1b), (11) and (111), ing cis-trans isomers, stereoisomers and tautomers, as well as racemic mixtures of these and ceutically acceptable salts thereof. Hence, the de?nition of compounds of Formulae (I), (la), (1b), (11) and (III) are also intended to encompass all R- and S-isomers of a chemical structure in any ratio, e. g., with enrichment (i. 6., enantiomeric excess or diastereomeric excess) of one of the possible isomers and ponding smaller ratios of other isomers. In addition, a crystal polymorphism may be present for the nds represented by Formulae (I), (la), (lb), (11) and (111). It is noted that any crystal form, l form mixture, or anhydride or hydrate thereof is included in the scope of the present sure. Furthermore, so-called metabolite which is produced by degradation of the present compound in vivo is included in the scope of the present disclosure.
"Isomerism" means compounds that have identical molecular formulae but differ in the sequence of bonding of their atoms or in the arrangement of their atoms in space. Isomers that differ in the arrangement of their atoms in space are termed oisomers".
Stereoisomers that are not mirror images of one another are termed ereoisomers", and stereoisomers that are perimposable mirror images of each other are termed "enantiomers" or sometimes optical isomers. A mixture containing equal amounts of individual enantiomeric forms of opposite ity is termed a ic mixture".
A carbon atom bonded to four non-identical substituents is termed a "chiral center".
[Annotation] KEB Chiral isomer" means a compound with at least one chiral center. Compounds with more than one chiral center may exist either as an individual diastereomer or as a mixture of diastereomers, termed "diastereomeric mixture". When one chiral center is present, a stereoisomer may be characterized by the absolute con?guration (R or S) of that chiral center.
Absolute con?guration refers to the arrangement in space of the substituents attached to the chiral center. The substituents attached to the chiral center under eration are ranked in accordance with the Sequence Rule of Cahn, Ingold and Prelog. (Cahn et al., Angew. Chem.
Inter. Edit. 1966, 5, 385; errata 511; Cahn er al., Angew. Chem. 1966, 78, 413; Cahn and Ingold, J. Chem. Soc. 1951 (London), 612; Cahn et (11., Experientia 1956, 12, 81; Cahn, J.
Chem. Educ. 1964, 41, 116).
Diastereoisomers, i.e., perimposable stereochemical isomers, can be separated by conventional means such as chromatography, distillation, crystallization or sublimation.
The optical isomers can be obtained by resolution of the racemic mixtures according to conventional processes, for example by formation of diastereoisomeric salts by treatment with an optically active acid or base. Examples of appropriate acids e, without limitation, tartaric, diacetyltartaric, dibenzoyltartaric, ditoluoyltartaric and camphorsulfonic acid. The mixture of diastereomers can be separated by crystallization followed by liberation of the lly active bases from these salts. An alternative process for separation of optical isomers includes the use of a chiral chromatography column optimally chosen to maximize the separation of the omers. Still another ble method involves synthesis of covalent diastereoisomeric molecules by reacting compounds of Formula (I), (la), (lb), (11) or (III) with an optically pure acid in an activated form or an optically pure isocyanate. The synthesized diastereoisomers can be separated by tional means such as chromatography, distillation, crystallization or sublimation, and then hydrolyzed to obtain the enantiomerically pure compound. The optically active compounds of Formulae (I), (la), (lb), (11) and (111) can likewise be obtained by utilizing lly active starting materials and/or by utilizing a chiral catalyst. These isomers may be in the form of a free acid, a free base, an ester or a salt. Examples of chiral separation techniques are given in Chiral Separation ques, A Practical Approach, 2nd ed. by G. Subramanian, Wiley—VCH, 2001.
"Geometric " means the reomers that owe their existence to ed on about double bonds. These con?gurations are differentiated in their names by the pre?xes cis and trans, or Z and E, which indicate that the groups are on the same or opposite side of the double bond in the molecule according to the Cahn—Ingold—Prelog rules.
[Annotation] KEB Furthermore, the structures and other compounds discussed in this disclosure include all atropic isomers thereof. "Atropic isomers" are a type of stereoisomer in which the atoms of two isomers are arranged differently in space. Atropic isomers owe their existence to a restricted rotation caused by hindrance of rotation of large groups about a central bond. Such atropic isomers typically exist as a mixture, however as a result of recent advances in chromatography techniques; it has been le to separate mixtures of two atropic isomers in select cases.
"Tautomer" is one of two or more structural isomers that exist in equilibrium and is readily converted from one isomeric form to r. This conversion results in the formal migration of a hydrogen atom accompanied by a switch of adjacent conjugated double bonds.
Tautomers exist as a mixture of a tautomeric set in solution. In solid form, usually one tautomer predominates. In solutions where tautomerization is le, a chemical equilibrium of the tautomers will be reached. The exact ratio of the ers depends on several factors, including temperature, t and pH. The concept of tautomers that are interconvertible by tautomerizations is called tautomerism.
Of the various types of tautomerism that are possible, two are commonly observed. In keto-enol tautomerism a simultaneous shift of electrons and a hydrogen atom occurs. Ring— chain tautomerism arises as a result of the aldehyde group (-CHO) in a sugar chain molecule reacting with one of the hydroxy groups (-OH) in the same molecule to give it a cyclic (ringshaped ) form as exhibited by glucose.
Common eric pairs are: -enol, amide-nitrile, lactam-lactim, amide— imidic acid tautomerism in heterocyclic rings (e.g., in nucleobases such as guanine, thymine and cytosine), amine-enamine and enamine-enamine. It is to be understood that the compounds of the present disclosure may be depicted as different ers. It should also be understood that when nds have tautomeric forms, all tautomeric forms are intended to be included in the scope of the t disclosure, and the naming of the compounds does not exclude any tautomer form.
The term al polymorphs", orphs" or "crystal forms" means crystal structures in which a compound (or a salt or solvate thereof) can crystallize in ent crystal packing arrangements, all of which have the same elemental composition. Different crystal forms usually have different X-ray diffraction patterns, infrared spectral, melting points, y hardness, l shape, optical and electrical properties, stability and solubility. Recrystallization solvent, rate of crystallization, storage temperature, and other [Annotation] KEB factors may cause one crystal form to te. Crystal polymorphs of the compounds can be ed by crystallization under different conditions.
Additionally, the compounds of the present disclosure, for example, the salts of the compounds, can exist in either hydrated or unhydrated (the anhydrous) form or as solvates with other solvent molecules. Nonlimiting examples of hydrates include monohydrates, dihydrates, etc. Nonlimiting examples of solvates include ethanol solvates, acetone solvates, "Solvate" means solvent addition forms that contain either iometric or non— stoichiometric amounts of t. Some compounds have a tendency to trap a ?xed molar ratio of solvent molecules in the crystalline solid state, thus forming a solvate. If the solvent is water the solvate formed is a hydrate; and if the solvent is alcohol, the solvate formed is an alcoholate. es are formed by the ation of one or more molecules of water with one molecule of the substance in which the water retains its molecular state as H20.
The present disclosure is intended to include all isotopes of atoms occurring in the present compounds. Isotopes include those atoms having the same atomic number but different mass numbers. By way of general example and without limitation, isotopes of en include tritium and ium, and isotopes of carbon include C—13 and C—14. hout the description and claims of this speci?cation, the words "comprise" and "contain" and variations of the words, for example "comprising" and "comprises", mean "including but not limited to" and do not exclude other moieties, additives, components, rs or steps. Throughout the description and claims of this specification, the singular encompasses the plural unless the t otherwise es. In particular, where the inde?nite article is used, the specification is to be understood as contemplating plurality as well as singularity, unless the context requires otherwise.
All references, including any patent or patent application, cited in this specification are hereby incorporated by reference. No admission is made that any reference constitutes prior art. r, no ion is made that any of the prior art constitutes part of the common general knowledge in the art.
Method of treatment In another aspect, the present disclosure relates to a method of treating a disease or er in which oc—amino-B—carboxymuconate-e—semialdehyde decarboxylase (ACMSD) plays a role comprising administering to the subject in need thereof a therapeutically effective [Annotation] KEB amount of one or more compounds of Formula (I), Formula (Ia), Formula (Ib), Formula (II), or Formula (III), or a ceutically acceptable salt thereof.
Another aspect of the t disclosure relates to a method of treating a disease or disorder associated with 0t—amino—B—carboxymuconate—a—semialdehyde decarboxylase (ACMSD) dysfunction comprising stering to the subject suffering from or susceptible to ping a disease or disorder associated with ACMSD dysfunction a therapeutically effective amount of one or more compounds of Formula (I), Formula (Ia), Formula (Ib), Formula (II), or Formula (III), or a pharmaceutically acceptable salt thereof.
In another aspect, the present disclosure relates to a method of ting a disease or er associated with a-amino-B-carboxymuconate—s-semialdehyde decarboxylase (ACMSD) dysfunction comprising administering to the subject suffering from or susceptible to developing a disease or disorder associated with ACMSD dysfunction a therapeutically ive amount of one or more nds of Formula (I), Formula (Ia), Formula (Ib), Formula (II), or Formula (111), or a pharmaceutically acceptable salt f In yet another aspect, the present disclosure relates to a method of reducing the risk of a disease or er associated with oc—amino-B—carboxymuconate—8—semialdehyde decarboxylase (ACMSD) dysfunction comprising administering to the subject suffering from or susceptible to developing a disease or disorder associated with ACMSD dysfunction a therapeutically effective amount of one or more compounds of Formula (I), Formula (Ia), a (Ib), Formula (II), or Formula (III), or a pharmaceutically acceptable salt thereof.
Another aspect of the present sure relates to a method of ameliorating the risk of a disease or disorder ated with OL—amino-B—carboxymuconate—s—semialdehyde oxylase (ACMSD) dysfunction comprising administering to the subject suffering from or susceptible to developing a e or disorder associated with ACMSD dysfunction a therapeutically effective amount of one or more compounds of a (I), Formula (Ia), Formula (Ib), Formula (II), or Formula (III), or a pharmaceutically acceptable salt thereof.
In another aspect, the present disclosure relates to a method of treating a disease or disorder in which nicotinamide adenine dinucleotide (NAD+) modulation plays a role comprising administering to the subject in need thereof a therapeutically effective amount of one or more compounds of Formula (1), Formula (Ia), Formula (1b), a (II), or Formula (III), or a pharmaceutically acceptable salt thereof.
[Annotation] KEB Another aspect of the present disclosure relates to a method of preventing a disease or disorder in which nicotinamide adenine dinucleotide (NAD+) modulation plays a role comprising administering to the subject in need thereof a therapeutically effective amount of one or more compounds of Formula (1), Formula (Ia), Formula (lb), Formula (II), or Formula (111), or a pharmaceutically acceptable salt thereof.
In another aspect, the t disclosure relates to a method of reducing the risk of a disease or disorder in which nicotinamide adenine dinucleotide (NAD+) tion plays a role comprising administering to the subject in need thereof a therapeutically effective amount of one or more nds of Formula (1), Formula (Ia), a (lb), Formula (II), or Formula (111), or a pharmaceutically acceptable salt thereof Another aspect of the present disclosure relates to a method of ameliorating a disease or disorder in which nicotinamide adenine dinucleotide (NAD+) modulation plays a role comprising administering to the subject in need thereof a therapeutically effective amount of one or more compounds of Formula (1), Formula (Ia), Formula (1b), Formula (II), or Formula (111), or a pharmaceutically acceptable salt thereof.
In r aspect, the present sure relates to a method of treating a disease or disorder ated with reduced namide adenine dinucleotide (NAD+) levels comprising administering to the subject suffering from or susceptible to developing a e or disorder ated with reduced NAD+ levels a therapeutically effective amount of one or more nds of Formula (1), Formula (Ia), Formula (1b), a (II), or Formula (111), or a pharmaceutically acceptable salt thereof.
The present disclosure also relates to a method of preventing a disease or disorder associated with reduced nicotinamide adenine dinucleotide (NAD+) levels comprising administering to the t suffering from or susceptible to developing a disease or disorder associated with reduced NAD+ levels a eutically effective amount of one or more compounds of Formula (1), Formula (Ia), Formula (1b), Formula (II), or Formula (111), or a pharmaceutically able salt thereof.
Another aspect of the present disclosure relates to a method of reducing the risk of a disease or disorder associated with reduced nicotinamide adenine dinucleotide (NAD+) levels comprising administering to the subject ing from or susceptible to developing a disease or disorder associated with reduced NAD+ levels a therapeutically effective amount of one or ation] KEB more compounds of Formula (I), Formula (Ia), Formula (Ib), Formula (II), or Formula (III), or a pharmaceutically acceptable salt thereof.
In another aspect, the present disclosure relates to a method of ameliorating a e or disorder associated with reduced nicotinamide adenine dinucleotide (NAD+) levels comprising stering to the subject suffering from or tible to developing a disease or disorder associated with reduced NADI levels a eutically effective amount of one or more compounds of Formula (I), Formula (Ia), Formula (Ib), Formula (II), or Formula (III), or a pharmaceutically acceptable salt thereof Another aspect of the present disclosure relates to a method of treating a disorder associated with mitochondrial dysfunction comprising stering to the subject suffering from or susceptible to developing a lic disorder a therapeutically effective amount of one or more compounds of Formula (I), Formula (Ia), Formula (Ib), Formula (II), or Formula (III), or a pharmaceutically acceptable salt thereof. In one embodiment, the er associated with mitochondrial dysfunction is an inherited mitochondrial disease, a common metabolic disorder, a neurodegenerative disease, an aging related disorder, a kidney disorder, or a chronic in?ammatory disease. In a preferred embodiment, the disorder associated With mitochondrial dysfunction is a common metabolic disorder such as obesity or type II diabetes.
In another aspect, the present disclosure relates to a method of ting a disorder associated with mitochondrial dysfunction comprising administering to the subject suffering from or susceptible to developing a metabolic er a therapeutically effective amount of one or more compounds of Formula (I), Formula (Ia), Formula (Ib), Formula (II), or Formula (III), or a ceutically acceptable salt thereof. In one embodiment, the disorder associated with mitochondrial dysfunction is an inherited mitochondrial disease, a common metabolic disorder, a neurodegenerative disease, an aging related disorder, a kidney er, or a chronic in?ammatory disease. In a preferred embodiment, the er associated with mitochondrial ction is a common lic disorder such as obesity or type II diabetes.
Another aspect of the present disclosure relates to a method of reducing the risk of a disorder associated with mitochondrial dysfunction comprising administering to the subject suffering from or susceptible to developing a metabolic disorder a therapeutically ive amount of one or more compounds of Formula (I), Formula (Ia), Formula (Ib), Formula (II), or Formula (III), or a pharmaceutically acceptable salt thereof. In one embodiment, the disorder associated with mitochondrial dysfunction is an inherited mitochondrial disease, a [Annotation] KEB common metabolic disorder, a neurodegenerative disease, an aging related disorder, a kidney disorder, or a chronic in?ammatory disease. In a preferred embodiment, the disorder associated with mitochondrial dysfunction is a common metabolic disorder such as obesity or type II diabetes.
Another aspect of the t disclosure relates to a method of ameliorating a disorder associated with mitochondrial dysfunction comprising administering to the subject suffering from or susceptible to ping a metabolic disorder a therapeutically effective amount of one or more compounds of Formula (1), a (Ia), Formula (Ib), Formula (II), or Formula (III), or a pharmaceutically acceptable salt thereof. In one ment, the disorder associated with mitochondrial dysfunction is an inherited mitochondrial disease, a common metabolic disorder, a neurodegenerative disease, an aging related disorder, a kidney disorder, or a chronic in?ammatory disease. In a preferred embodiment, the disorder associated with mitochondrial dysfunction is a common metabolic disorder such as obesity or type II diabetes.
In another aspect, the t disclosure relates to a method of ing ive metabolism comprising administering to the subject suffering from or susceptible to developing a metabolic disorder a therapeutically effective amount of one or more compounds of Formula (I), Formula (Ia), Formula (Ib), Formula (II), or a (III), or a pharmaceutically acceptable salt thereof, that increases intracellular nicotinamide e dinucleotide (NAD+).
In yet another aspect, the t disclosure relates to a method for the manufacture of a medicament for treating a disease or condition mediated by ACMSD, n the medicament ses a compound of Formula (I), Formula (Ia), Formula (Ib), Formula (II), or Formula (III), or a pharmaceutically acceptable salt thereof. r aspect of the present disclosure relates to a method for the manufacture of a medicament for preventing a disease or condition mediated by ACMSD, wherein the medicament comprises a compound of Formula (I), Formula (Ia), Formula (Ib), Formula (II), or a (III), or a pharmaceutically acceptable salt thereof.
In r aspect, the present disclosure relates to a method for the manufacture of a medicament for ng the risk of a disease or condition mediated by ACMSD, n the ment comprises a compound of Formula (1), Formula (Ia), Formula (Ib), Formula (II), or Formula (III), or a pharmaceutically acceptable salt thereof.
In yet another aspect, the present disclosure relates to a method for the manufacture of a medicament for ameliorating a disease or condition mediated by ACMSD, wherein the [Annotation] KEB medicament ses a compound of Formula (I), Formula (Ia), Formula (lb), Formula (II), or Formula (III), or a pharmaceutically acceptable salt thereof.
In another aspect, the present disclosure relates to a pharmaceutical composition for use in a method for treating a disease or condition mediated by ACMSD, n the medicament ses a compound of Formula (I), Formula (Ia), Formula (lb), a (II), or a (III), or a pharmaceutically acceptable salt thereof.
In another aspect, the present disclosure relates to a pharmaceutical composition for use in a method for preventing a disease or condition mediated by ACMSD, wherein the medicament comprises a compound of Formula (I), Formula (Ia), Formula (lb), Formula (II), or Formula (III), or a pharmaceutically able salt thereof.
In another aspect, the present disclosure relates to a pharmaceutical composition for use in a method for ng the risk of a disease or condition mediated by ACMSD, wherein the medicament comprises a compound of Formula (I), Formula (Ia), Formula (lb), a (II), or Formula (III), or a pharmaceutically acceptable salt thereof.
Another aspect of the present disclosure relates to a pharmaceutical composition for use in a method for rating a e or condition mediated by ACMSD, wherein the medicament comprises a compound of Formula (I), Formula (Ia), Formula (lb), Formula (II), or Formula (III), or a pharmaceutically acceptable salt thereof In yet another aspect, the present disclosure relates to a compound for use in a method for treating a disease or condition mediated by ACMSD, wherein the nd comprises a compound of Formula (I), Formula (Ia), Formula (lb), Formula (II), or Formula (III), or a pharmaceutically acceptable salt f.
Another aspect of the present disclosure relates to a compound for use in a method for preventing a disease or condition mediated by ACMSD, wherein the compound comprises a compound of Formula (I), Formula (Ia), a (lb), Formula (II), or Formula (III), or a pharmaceutically acceptable salt thereof.
In another aspect, the t disclosure relates to a compound for use in a method for reducing the risk of a disease or ion mediated by ACMSD, wherein the compound ses a compound of Formula (1), a (Ia), Formula (lb), Formula (II), or Formula (III), or a pharmaceutically acceptable salt thereof.
Another aspect of the present disclosure relates to a compound for use in a method for ameliorating a disease or condition mediated by ACMSD, wherein the compound comprises a compound of Formula (1), Formula (Ia), Formula (lb), a (II), or Formula (III), or a pharmaceutically acceptable salt thereof.
[Annotation] KEB Another aspect of the present disclosure relates to the use of a nd of Formula (1), a (la), Formula (lb), Formula (II), or Formula (III), or a pharmaceutically acceptable salt thereof in the manufacture of a medicament for treating, preventing or reducing the risk of a disease or disorder associated with no—[3-carboxymuconate—e- semialdehyde decarboxylase (ACMSD) dysfunction.
In another aspect, the present sure relates to the use of a compound of Formula (1), Formula (la), Formula (lb), Formula (II), or Formula (III), or a pharmaceutically acceptable salt thereof in the manufacture of a medicament for treating the risk of a disease or disorder ated with oc—amino—B—carboxymuconate—8—semialdehyde decarboxylase (ACMSD) dysfunction.
Another aspect of the present disclosure relates to the use of a compound of Formula (1), Formula (la), Formula (lb), Formula (II), or Formula (III), or a pharmaceutically acceptable salt thereof in the manufacture of a ment for preventing a disease or disorder associated with no—B—carboxymuconate—8—semialdehyde oxylase (ACMSD) dysfunction.
In another aspect, the present disclosure relates to the use of a compound of Formula (I), Formula (Ia), Formula (Ib), Formula (II), or Formula (III), or a pharmaceutically acceptable salt thereof in the cture of a medicament for reducing the risk of a disease or disorder associated with oc—amino—B—carboxymuconate—8—semialdehyde decarboxylase (ACMSD) dysfunction.
Another aspect of the present disclosure relates to the use of a compound of Formula (1), Formula (la), Formula (lb), Formula (II), or Formula (III), or a ceutically able salt thereof in the cture of a medicament for ameliorating a disease or disorder associated with a—amino—[3—carboxymuconate—e—semialdehyde decarboxylase (ACMSD) dysfunction.
In another , the present disclosure relates to the use of a compound of a compound of Formula (1), Formula (Ia), Formula (lb), Formula (II), or Formula (III), or a pharmaceutically acceptable salt thereof in the manufacture of a medicament for treating, ting or reducing the risk of a disease or disorder associated with reduced nicotinamide adenine dinucleotide (NAD+) .
Another aspect of the present disclosure relates to the use of a compound of a compound of Formula (1), Formula (la), Formula (lb), Formula (II), or Formula (III), or a pharmaceutically acceptable salt thereof in the manufacture of a medicament for treating a ation] KEB disease or disorder ated with d namide adenine dinucleotide (NADl) levels.
In another aspect, the present disclosure relates to the use of a compound of a compound of Formula (I), Formula (Ia), Formula (lb), Formula (II), or a (III), or a pharmaceutically acceptable salt thereof in the manufacture of a medicament for preventing a disease or disorder associated with reduced nicotinamide adenine dinucleotide (NADl) levels.
Another aspect of the present disclosure relates to the use of a compound of a compound of Formula (I), Formula (Ia), Formula (lb), Formula (II), or Formula (III), or a pharmaceutically acceptable salt thereof in the manufacture of a medicament ng the risk of a disease or disorder associated with reduced nicotinamide adenine dinucleotide (NADl) levels.
In another aspect, the present disclosure relates to the use of a compound of a compound of a (I), Formula (Ia), Formula (lb), Formula (II), or a (III), or a ceutically acceptable salt thereof in the manufacture of a medicament for ameliorating a disease or disorder associated with reduced nicotinamide adenine dinucleotide (NAD+) levels.
Another aspect of the present sure relates to the use of a compound of Formula (I), Formula (Ia), Formula (lb), Formula (II), or Formula (III), or a pharmaceutically acceptable salt thereof in the manufacture of a medicament for treating, preventing or reducing the risk of a disorder associated with mitochondrial ction.
In another aspect, the t disclosure relates to the use of a compound of Formula (I), Formula (Ia), Formula (Ib), Formula (II), or Formula (III), or a pharmaceutically acceptable salt thereof in the manufacture of a medicament for treating a disorder associated with mitochondrial dysfunction.
Another aspect of the present disclosure relates to the use of a compound of Formula (I), Formula (Ia), Formula (Ib), Formula (II), or Formula (III), or a pharmaceutically acceptable salt thereof in the manufacture of a medicament for preventing a disorder associated with mitochondrial ction.
In another aspect, the t disclosure s to the use of a compound of Formula (I), a (Ia), Formula (lb), Formula (II), or Formula (III), or a pharmaceutically acceptable salt thereof in the manufacture of a medicament for reducing the risk of a disorder associated with mitochondrial dysfunction.
[Annotation] KEB Another aspect of the present disclosure relates to the use of a compound of Formula (I), Formula (Ia), Formula (Ib), Formula (II), or Formula (III), or a pharmaceutically acceptable salt thereof in the cture of a medicament for ameliorating a disorder associated with mitochondrial dysfunction.
In another aspect, the present disclosure relates to the use of a compound of a compound of Formula (I), Formula (Ia), a (Ib), Formula (II), or Formula (III), or a pharmaceutically acceptable salt f in the manufacture of a medicament for ing oxidative metabolism. r aspect of the present sure relates to a compound of Formula (I), Formula (Ia), Formula (Ib), Formula (II), or Formula (III), or a pharmaceutically acceptable salt thereof for use in the manufacture of a medicament for treating, preventing or ng the risk of a disease or er associated with oc—amino—B—carboxymuconate—s—semialdehyde decarboxylase (ACMSD) dysfunction.
In another , the present disclosure relates to a compound of Formula (I), Formula (Ia), Formula (Ib), Formula (II), or Formula (III), or a pharmaceutically acceptable salt f for use in the manufacture of a medicament for treating a disease or disorder associated with oe—amino-B—carboxymuconate-s—semialdehyde decarboxylase (ACMSD) dysfunction.
Another aspect of the t disclosure relates to a compound of Formula (I), Formula (Ia), Formula (Ib), Formula (II), or Formula (III), or a pharmaceutically acceptable salt thereof for use in the manufacture of a medicament for preventing a e or disorder associated with oe-amino—B-carboxymuconate—e—semialdehyde decarboxylase (ACMSD) dysfunction.
In another aspect, the present disclosure relates to a compound of Formula (I), Formula (Ia), Formula (Ib), a (II), or Formula (III), or a pharmaceutically acceptable salt f for use in the manufacture of a medicament for reducing the risk of a disease or disorder associated with oc—amino—B—carboxymuconate—8—semialdehyde decarboxylase (ACMSD) ction.
Another aspect of the present disclosure relates to a compound of Formula (I), Formula (Ia), Formula (Ib), Formula (II), or Formula (III), or a pharmaceutically acceptable salt thereof for use in the cture of a medicament for ameliorating a disease or disorder associated with Ot—amino-B—carboxymuconate—e—semialdehyde decarboxylase (ACMSD) dysfunction.
[Annotation] KEB In another aspect, the present disclosure relates to a compound of Formula (I), Formula (Ia), Formula (Ib), a (II), or Formula (III), or a pharmaceutically acceptable salt thereof for use as a ment for treating, preventing or reducing the risk of a disease or disorder associated with reduced nicotinamide adenine dinucleotide (NAD+) levels.
Another aspect of the present disclosure relates to a compound of a (I), Formula (Ia), a (Ib), a (II), or Formula (III), or a pharmaceutically acceptable salt thereof for use as a medicament for treating a disease or disorder associated with reduced nicotinamide adenine dinucleotide (NAD+) levels.
In another aspect, the present disclosure s to a compound of Formula (I), Formula (Ia), Formula (Ib), Formula (II), or Formula (III), or a pharmaceutically acceptable salt thereof for use as a medicament for preventing a disease or disorder associated with reduced nicotinamide adenine dinucleotide (NADI) .
Another aspect of the present disclosure relates to a compound of Formula (1), Formula (Ia), Formula (Ib), Formula (II), or Formula (III), or a pharmaceutically acceptable salt thereof for use as a medicament for reducing the risk of a disease or er associated with reduced nicotinamide adenine dinucleotide (NADI) levels.
In another aspect, the present disclosure s to a compound of Formula (I), Formula (Ia), Formula (Ib), a (II), or Formula (III), or a pharmaceutically able salt thereof for use as a medicament for ameliorating a disease or disorder associated with reduced nicotinamide adenine dinucleotide (NAD+) levels.
Another aspect of the present disclosure relates to a compound of Formula (I), Formula (Ia), Formula (Ib), Formula (II), or Formula (III), or a pharmaceutically acceptable salt thereof for use as a medicament for treating, ting or reducing the risk of a disorder ated with mitochondrial dysfunction.
In another aspect, the present disclosure relates to a compound of a (I), a (Ia), Formula (Ib), Formula (II), or Formula (III), or a pharmaceutically acceptable salt thereof for use as a ment for treating a disorder associated with mitochondrial dysfunction.
Another aspect of the t disclosure relates to a nd of Formula (I), Formula (Ia), Formula (Ib), Formula (II), or Formula (III), or a pharmaceutically acceptable salt thereof for use as a medicament for preventing a disorder associated with mitochondrial dysfunction.
In another aspect, the present disclosure relates to a compound of Formula (I), Formula (Ia), Formula (Ib), Formula (II), or Formula (III), or a pharmaceutically acceptable [Annotation] KEB salt thereof for use as a medicament for reducing the risk of a disorder associated with mitochondrial dysfunction. r aspect of the present disclosure relates to a compound of Formula (I), Formula (la), Formula (lb), Formula (II), or Formula (III), or a pharmaceutically acceptable salt thereof for use as a medicament for ameliorating a disorder associated with mitochondrial dysfunction.
In another aspect, the present disclosure relates to a compound of Formula (I), Formula (Ia), Formula (lb), Formula (II), or Formula (111), or a pharmaceutically acceptable salt thereof for use as a medicament for promoting oxidative metabolism.
Another aspect of the present disclosure relates to a compound of Formula (I), Formula (la), Formula (lb), Formula (II), or Formula (III), or a pharmaceutically acceptable salt thereof for use in treating, preventing or reducing the risk of a disease or disorder associated with reduced nicotinamide adenine dinucleotide (NAD+) levels.
In another aspect, the present disclosure relates to a compound of a (I), Formula (Ia), Formula (lb), Formula (II), or Formula (111), or a pharmaceutically acceptable salt thereof for use in treating a disease or disorder associated with reduced namide adenine dinucleotide (NADl) levels.
Another aspect of the present disclosure relates to a compound of Formula (I), Formula (Ia), Formula (lb), Formula (II), or Formula (III), or a pharmaceutically able salt thereof for use in preventing a disease or er associated with reduced nicotinamide adenine dinucleotide (NADl) levels.
In another aspect, the t disclosure s to a compound of Formula (1), Formula (Ia), Formula (lb), Formula (II), or Formula (III), or a pharmaceutically acceptable salt thereof for use in reducing the risk of a disease or disorder associated with d nicotinamide adenine dinucleotide (NADl) levels.
Another aspect of the present disclosure relates to a compound of Formula (I), Formula (Ia), Formula (lb), Formula (II), or Formula (III), or a pharmaceutically able salt thereof for use in ameliorating a disease or disorder associated With reduced namide adenine dinucleotide (NAD+) levels.
In another aspect, the t disclosure relates to a nd of a (I), a (Ia), Formula (lb), Formula (II), or Formula (III), or a ceutically acceptable salt thereof for use in for treating, preventing or reducing the risk of a er associated with mitochondrial dysfunction.
[Annotation] KEB Another aspect of the present sure relates to a compound of a (I), Formula (Ia), Formula (lb), Formula (II), or Formula (III), or a pharmaceutically acceptable salt thereof for use in for treating a disorder associated with mitochondrial dysfunction.
In another , the present disclosure relates to a compound of Formula (I), Formula (Ia), Formula (lb), a (II), or Formula (III), or a pharmaceutically acceptable salt f for use in for preventing a disorder associated with ondrial dysfunction.
Another aspect of the present disclosure relates to a compound of a (I), Formula (Ia), Formula (lb), a (II), or Formula (III), or a pharmaceutically acceptable salt thereof for use in for reducing the risk of a disorder associated with mitochondrial dysfunction.
In another aspect, the present disclosure relates to a compound of Formula (I), Formula (Ia), Formula (lb), Formula (II), or Formula (III), or a pharmaceutically able salt thereof for use in for ameliorating a disorder associated with mitochondrial dysfunction.
Another aspect of the t disclosure relates to a compound of Formula (I), Formula (Ia), Formula (lb), Formula (II), or Formula (111), or a pharmaceutically acceptable salt thereof for use in promoting oxidative metabolism.
In another aspect, the present disclosure relates to a method of treating, preventing, ameliorating or reducing the risk of a disease or disorder associated with OL—amino—B— carboxymuconate—s—semialdehyde decarboxylase (ACMSD) dysfunction, comprising administering to a subject in need thereof, a therapeutically effective amount of compound having the following Formulae: O 0 NC NC NH NH I I ation] KEB [Annotation] KEB 0 O Nf/U 2 N’)\/\© N/J\)K©/COZH 9 H 7 I"1%me COH2 ["le NC W/UCOZHN NC NH NH :JE‘IAGACOZHO I OJN\H N N/\©H NC:J\,N\©/COZH::NC LHON NH O / I:/J\|N'7J\©/\C02H NH O NH 0 NH NC 0w NC NH NH 0r :Ao/U 2 7 or a pharmaceutically acceptable salt thereof.
In a further aspect, the present disclosure relates to a method of treating, preventing, ameliorating or reducing the risk of a disease or disorder associated with OL—amino—B— carboxymuconate-s—semialdehyde decarboxylase (ACMSD) ction, comprising administering to a subject in need f, a therapeutically effective amount of compound having the following Formulae: O 0 NC NC N/NHS/Ucoszo NH lNASUcozH ation] KEB [Annotation] KEB or a pharmaceutically acceptable salt thereof.
Hence, the disclosure also relates to a compound of Formula (1), Formula (Ia), Formula (1b), Formula (II), or a (111), or a pharmaceutically acceptable salt thereof, as de?ned herein, for use as a medicament.
Another aspect of the present disclosure relates to the use of a nd having the one of the following Formula: 0 O NH NH ation] KEB [Annotation] KEB o o 0 NC NC NC NH 0 NH NH I | I / / COH / N N N o o 7 7 7 :AN/UCOZH o 0 "° NC "" ZH I N/ N/\@1" NCN’J\/N\©/C°2HN: MO NH 0 I H II:J\"J\©/\C02H IN/AEJKO:02HNNH 0 NH 0 NH NA’O/UCOZH I 1" CI N/ N/ 0/01" o/\©/Cozl'I 0r 7 7 or a pharmaceutically acceptable salt thereof in the manufacture of a medicament for treating, preventing, ameliorating or ng the risk of a disease or er associated with oc—amino—B—carboxymuconate—s—semialdehyde decarboxylase (ACMSD) dysfunction.
Another aspect of the present disclosure relates to the use of a compound having the one of the following Formula: 0 0 NC NC NH NH I I ation] KEB ation] KEB Cl 0 N\\ NC | \1 H / co2H S /A N or a pharmaceutically acceptable salt thereof in the manufacture of a medicament for treating, preventing, rating or reducing the risk of a disease or disorder associated with oc—amino—B—carboxymuconate—s—semialdehyde decarboxylase (ACMSD) dysfunction.
In another aspect, the present disclosure relates to a compound having the one of the following Formula: ation] KEB ation] KEB 0 0 NC W/UCOZHN NC NH NH NH :unx\©/\COZHO I NAM/\© m:’J\/N\©/002H: Marl NH 0 I H EWJKQACOZH NH O NH O NH NC GAO/UcozH NC NH NH or :Ao/\© , 7 or a pharmaceutically acceptable salt thereof, for use as a medicament for treating, preventing, ameliorating or reducing the risk of a e or disorder associated with a-amino-B-carboxymuconate-e-semialdehyde decarboxylase (ACMSD) dysfunction.
In another aspect, the present disclosure relates to a compound having the one of the following Formula: NC NC [Annotation] KEB or a ceutically acceptable salt thereof, [Annotation] KEB for use as a medicament for treating, preventing, ameliorating or reducing the risk of a disease or disorder ated with a-amino—B—carboxymuconate—a-semialdehyde decarboxylase (ACMSD) dysfunction.
Another aspect of the t disclosure relates to a compound having the one of the following Formula: NC NC ation] KEB ation] KEB o o (SILNH NC NAG/UCOZH I or NAG/D , 7 or a pharmaceutically acceptable salt thereof for use in treating, preventing, rating or reducing the risk of a disease or disorder associated with oe-amino—B-carboxymuconate—e-semialdehyde decarboxylase (ACMSD) dysfunction.
In another aspect, the present disclosure relates to a compound having the one of the following Formula: NC NC [Annotation] KEB or a pharmaceutically acceptable salt thereof for use in treating, preventing, ameliorating or reducing the risk of a e or disorder associated with a—amino-B—carboxymuconate—s—semialdehyde decarboxylase (ACMSD) dysfunction.
As used , "treating" or "treat" describes the management and care of a t for the purpose of reversing, inhibiting, or combating a e, ion, or disorder and includes the administration of a compound of the present disclosure (i.e., a compound of Formula (1), Formula (1a), Formula (1b), Formula (11), or Formula (111)), or a pharmaceutically acceptable salt, prodrug, metabolite, polymorph or solvate thereof, to e the disease, condition, or disorder, eliminate the disease, condition, or disorder, or inhibit the process of the e, condition, or disorder.
A compound of the present disclosure (i.e., a compound of Formula (1), Formula (1a), Formula (1b), Formula (11), or Formula (111)), or a pharmaceutically acceptable salt, prodrug, [Annotation] KEB metabolite, polymorph or solvate thereof, can also be used to prevent a disease, condition, or disorder or one or more symptoms of such disease, condition, or disorder. As used herein, "preventing" or "prevent" describes reducing or eliminating the onset of the symptoms or complications of the disease, condition, or disorder.
A compound of the present disclosure (i.e., a compound of Formula (1), Formula (1a), Formula (1b), Formula (11), or Formula (111)), or a pharmaceutically acceptable salt, prodrug, metabolite, polymorph or solvate thereof, can also be used to alleviate one or more symptoms of such disease, ion, or disorder. As used herein, the term "alleviate" is meant to describe a process by which the ty of a sign or symptom of a disorder is sed.
Importantly, a sign or symptom can be alleviated without being eliminated. Preferably treatment is curative or ameliorating.
Methods for the re aration of com ounds of ae 1 1a 1b 11 and III The compounds of the present disclosure (e.g., compounds of Formula (1), Formula (1a), Formula (1b), Formula (11), and Formula (11)) can be prepared in a number of ways well known to those skilled in the art of organic synthesis. By way of e, compounds of the present sure can be synthesized using the methods described below, together with synthetic methods known in the art of synthetic organic try, or variations thereon as appreciated by those skilled in the art. red methods include but are not limited to those methods described below. The ?nal products of the reactions described herein may be isolated by conventional techniques, e. g., by extraction, crystallisation, distillation, chromatography, etc.
Compounds of the present disclosure can be synthesized by following the steps outlined in General Scheme A to E which comprise different ces of assembling intermediates Ia-Ih and Ij-Io. Starting materials are either commercially available or made by known ures in the reported literature or as illustrated. Useful steps that may be used in the preparation steps of the compounds will be known to the skilled person. The method below is given as a non-limiting example on how the nds may be prepared.
General Scheme A O 0 RC Re I NH 1 +x—L—R1 ~ I i Rd H S Rd N/ s/ \R1 Ia Ib (I) [Annotation] KEB wherein R1, RC, Rd, and L are de?ned as in Formula (I).
The general way of preparing compounds of Formula (I) by using intermediates Ia, and lb is outlined in General Scheme A. Coupling of la with Ib using a base, i.e., potassium carbonate (K2C03), in a solvent, i.e., acetonitrile (CH3CN), optionally at elevated temperature provides the desired produce of Formula (I). Bases that can be used include, but are not limited to, sodium ate 3), potassium carbonate (K2C03), N,N— diisopropylethylamine (DIPEA) and triethylamine. Solvents used in the coupling reaction can be polar or non-polar solvents. For example, the t can be acetonitrile (CH3CN), acetone, or dimethylsulfoxide (DMSO).
General Scheme B o o REL R" N 2 NH I I i + R‘NA 1 fL Rd NJ\X H 9R Rd N’J‘N’W1 H H p Ic Id (1) wherein X is a good leaving group, i.e., Cl, Br, -SCH3, or S(O)2CH3, and R1, R2, RC, Rd, and p are de?ned as in Formula (I).
Alternatively, compounds of Formula (I) can be prepared using intermediates Ic and Id as outlined in General Scheme B. Amination of Intermediate Ic with Ie using a base, i.e., sodium hydroxide , potassium hydroxide (KOH), etc., in a solvent, i. e., methanol , ethanol (EtOH), water (H20), etc., es compounds of Formula (I).
General Scheme C o o RYL no N NH Rd nJ‘x p Ra MAO/Hm Ie If (I) wherein X is a good leaving group, i.e., Cl, Br, —SCH3, or S(O)2CH3, and R1, R2, RC, Rd, and p are de?ned as in a (1). nds of Formula (I) can also be prepared using intermediates Ie and If as outlined in General Scheme C. Amination of Intermediate Ie with If using a base, i.e., ation] KEB sodium hydroxide (NaOH), potassium hydroxide (KOH), etc., in a solvent, i. e., methanol , ethanol (EtOH), water (H20), etc., provides compounds of Formula (I).
General Scheme D R1’§o \ CN —> R1’\/ —> R1’\/CN —> Ih lj NH 0 R° HCI NH —> | Ik Im I wherein and R1, RC, and Rd are de?ned as in Formula (1).
Alternatively, compounds of Formula (I) can also be prepared using intermediates Ig, Ih, Ij, Ik, and Im as ed in General Scheme D. Ole?nation of intermediate Ig using a base i.e., potassium carbonate (K2C03) and diethyl (cyanomethyl)phosphonate in a solvent, i.e., tetrahydrofuran (THF), water (H20), optionally at an elevated temperature provides Intermediate Ih. Hydrogenation of Ih using a metal catalyst, i.e., palladium on carbon (Pd/C), um dioxide (PtOz), etc, and hydrogen (H2) gas in a solvent, i.e., ethanol (EtOH) and/ or tetrahydrofuran (THF), provides Intermediate Ij. Intermediate Ik is obtained by treating Intermediate Ij with an acid, i.e., hydrochloric acid (HCI) in a solvent, i.e., ethanol (EtOH), dichloromethane (CHzClz), etc., and then subsequent treatment with a base, i.e., ammonia (NH3). ation of Intermediate Ik and Im using a base, i.e., sodium hydroxide (NaOH), potassium hydroxide (KOH), etc., in a t, i.e., dimethylacetamide (DMA), optionally at elevated temperature provides compounds of Formula (I).
General Scheme E 0 O R‘YL 0 RC NH NH 0 I NANHZ —> + LCI R 1|)L Rd Rd NAEJLRaI In 10 (I) n and R1, R6, and Rd are de?ned as in Formula (I).
[Annotation] KEB Alternatively, compounds of Formula (I) can be prepared using intermediates In and lo as outlined in General Scheme D. Acylation of Intermediate In with Io using a base, i. e., sodium hydroxide (NaOH), potassium hydroxide (KOH), etc., in a solvent, i. e., methanol (MeOH), ethanol , water (H20), etc., provides compounds of a (I).
A mixture of enantiomers, diastereomers, cis/trans s resulting from the process described above can be ted into their single components by chiral salt que, chromatography using normal phase, reverse phase or chiral column, ing on the nature of the separation.
It should be tood that in the description and formula shown above, the various groups R1, R2, X, L, Y, Ra, Rb, RC, Rd, Re, Rf, RX, Ry, RZ, m, n, p, q, r and other variables are as de?ned herein above, except where otherwise indicated. Furthermore, for synthetic purposes, the compounds of General Schemes 1 and 2 are mere representative with d radicals to illustrate the general synthetic methodology of the compounds of Formula (I) as defined herein.
Biological Assays and Animals Studies Method ofScreening ACMSD] Inhibition The activity of compounds as inhibitors ofACMSDl is determined in a spectrophotometrical in vitro assay. The pre-assay mixture is incubated and a compound of Formula (1), Formula (Ia), Formula (Ib), Formula (II), or Formula (111), or a pharmaceutically acceptable salt thereof, and ACMSDl solution is then added. The effect of ACMS tration on the enzyme activity is investigated by varying 3-hydroxyanthranilic acid (3 OH—HA) concentration in the pre-assay mixture. Kinetic parameters are calculated from the initial velocity data using a Lineweaver-Burk plot.
Celluiar Assay Methods The mouse hepatocytes cell lines are grown and plated. The cells are maintained in culture at 37 oC and once the cells are attached, different concentrations of a compound of a (1), Formula (Ia), Formula (lb), Formula (II), or Formula (111), or a pharmaceutically able salt thereof, or DMSO are added. Primary cytes are harvested about 24 hrs later.
[Annotation] KEB Determination ofACMSD-I Modulation in HEK293T Cells.
HEK293T cells are seeded and transfected to transiently express ACMSD. The cells are then stimulated with different concentrations of Compound 1, and then lysed to measure the ACMSD activity in a spectrophotometrical in Vitro assay. The amount of the whole protein content in cell lysates is detected by Bradford analysis and used to get the speci?city activity of the enzyme normalized in all samples. ination ofNAD+ content in Human Primary Hepatocytes Primary hepatocytes are d with different concentrations of a compound of Formula (1), Formula (Ia), Formula (1b), a (II), or Formula (111), or a pharmaceutically acceptable salt thereof, or MEHP (control) after seeding. The compound is replaced every 24 hours, and then cells are directly harvested and lysed to detect NAD+ content through LC MS/MS (liquid chromatography mass spectrometry/mass spectroscopy).
Modulation ofSOD2 activity in AMLIZ cells and Marine Primary Hepatocytes Primary hepatocytes or AML—l2 cells are lysed and total protein tration is determined using the Bradford assay. SOD2 activity is determined at indicated times after treatment with a compound of Formula (I), Formula (Ia), Formula (Ib), Formula (II), or Formula (111), or a pharmaceutically acceptable salt thereof, using a SOD Assay Kit.
Absorbance is determined and results are expressed in U/ml/mg of protein according to the standard curve and measured protein concentration.
Determination ofNAD+ content in Marine Primary Hepatocytes NAD+ is ted using acidic extraction method and samples are collected and homogenized. After insoluble protein parts are ed, the samples are separated by high- mance liquid chromatography (HPLC) and analyzed by mass-spectrometry. The proteins in the pellet are quanti?ed by Bradford assay and are used for normalization.
RNA preparation and RT—qPCR is ofACMSD and SIRTI-regulated Genes in Cells, Cells 2, .6, HEK—293, y human and murine hepatocytes) are treated with different concentrations of a compound of Formula (1), Formula (Ia), a (1b), a (II), or Formula (111), or a pharmaceutically acceptable salt thereof and the gene expression of ACMSD, ch1 a, Sod], and Sod2 (MnSOD) is determined using RT-qPCR.
Total RNA is extracted from cells and the extracted RNA is treated With DNase and used for reverse transcription (RT).
[Annotation] KEB Modulation of Caspase 3/7 Activity in MDCK Cells MDCK cells are cultured in base medium to a ?nal concentration of 10%. Cells are plated into 96 wells and 24 hours after cell plating the medium is changed with fresh medium supplemented with 1% FBS. Cisplatin is then used to induce cell injury. Different concentrations of Formula (1), Formula (Ia), Formula (1b), Formula (II), or Formula (111), or a pharmaceutically acceptable salt thereof (in DMSO) are added in ation with Cisplatin or prior to adding Cisplatin. Caspase 3/7 activity (Promega) is determined according to standard procedures using a luminescent signal readout on a plate reader. Each experiment/condition is performed in triplicate. Caspase activity is analyzed as percentage effect ized to the Cisplatin alone and vehicle treated cells.
Cytotoxicity and hERG screening HePGZ and AML-l2 cells are seeded and a dose-response of the nd is performed at various concentrations. Cells are stimulated and the supernatant is used to perform LDH release as a e of necrosis while the cells are lysed to detect ATP levels for determining cell viability.
The Predictor hERG assay kit is stably transfected with hERG potassium channel and a high—af?nity red ?uorescent hERG channel ligand and is used for the ination of hERG channel y binding of compounds of Formula (1), Formula (Ia), Formula (Ib), Formula (II), or Formula (111), or a pharmaceutically able salt thereof. Compounds that bind to the hERG l protein (competitors) are identi?ed by their ability to displace the tracer which results in a lower ?uorescence polarization.
Celegans experiments - ACMSD] ing, lifespan assays, mobility assessment and GFP quantification ACMSDI silencing: Bacterial feeding RNAi experiments to determine the s of downregulation or silencing of acmsd—I on gene expression and survival are carried out in the nematode Caenorhabditis elegans (C. elegans). The clones used for the ial feeding experiments are acmsd—J, SIR-2.1 and DAF-16. Total RNA is extracted from cells and the extracted RNA is treated with DNase, and used for reverse transcription (RT).
Worms are grown on NGM agar plates additionally containing Carbenicillin and IPTG and seeded with bacterial cultures. After RNAi ent, worms are transferred to plates containing paraquat and seeded with RNAi bacteria. Control animals are grown on RNAi ia containing an empty vector (control) and then erred to plates containing [Annotation] KEB paraquat and seeded with RNAi bacteria. Quanti?cation of gene expression of sod-3 at mRNA levels and protein levels using RT-qPCR and survival analyses are med. The movement of worms is recorded at days 1, 3, and 5 of adulthood.
Anti-diabetic E?ects studies in C57Bl/6J and KK-Ay mice Mice are fed with regular chow or a high fat diet (HFD). A compound of Formula (1), Formula (Ia), Formula (Ib), Formula (II), or Formula (111), or a ceutically acceptable salt thereof, is dosed daily and blood and tissues are harvested for RNA ion, lipid ements and histology. Oxygen consumption is measured and histological analysis and transmission electron microscopy are performed. An oral glucose tolerance test and an intraperitoneal insulin tolerance test are also performed to quantify glucose and to measure plasma insulin concentrations.
Anti-diabetic and Anti-obesity studies in db/db Mice with LepR Mutation Animals are fed a high-fat diet (HFD). For subchronic intervention, the animals are treated once/day with a compound of Formula (I), Formula (Ia), a (Ib), a (II), or Formula (III), or a pharmaceutically able salt thereof, for 14 days. Blood samples are collected and glucose concentrations of each blood sample are determined. For acute intervention, l blood samples are collected and then nds of Formula (1), Formula (Ia), a (Ib), Formula (II), or Formula (III), or a pharmaceutically acceptable salt thereof, are administered. Diet—access is then restricted, and a second blood sample is collected. The mice are subjected to an oral glucose tolerance test and blood glucose concentrations are determined.
For the euglycemic—hyperinsulinemic clamps assay, the animals receive a - continuous [3 —3H] glucose infusion and a blood sample is then collected to determine plasma insulin, e and [3—3H]glucose concentrations and to calculate basal endogenous e appearance rates. The mice then receive vehicle or a compound of Formula (I), Formula (Ia), Formula (Ib), Formula (II), or Formula (III), or a pharmaceutically acceptable salt thereof, via gavage. Subsequently, the animals receive a [3—3H] glucose infusion containing insulin causing a moderate net—increase in plasma insulin concentrations. Blood glucose concentrations are measured and target glycemia is established by adjusting the rate of glucose on. 2—deoxy—D—[l—14C] glucose is then given intravenously and blood s are collected. The mice are then sacri?ced. Gastrocnemius muscle and epididymal adipose [Annotation] KEB tissue are collected and plasma [3H]— and [14C]—radioactivity is determined in deproteinized Body weights are assessed and brown adipose tissue (BAT) and gonadal white adipose tissue (WAT) are ted and weighed. Volume oxygen (V02) and volume carbon dioxide tion (VCOz) are measured and are reported as average V02 per hour normalized to body weight (mL/h/kg). Activity counts by infrared beam interruptions and food intake are simultaneously measured.
Non-alcoholic fatty liver disease (NAFLD) and non-alcoholic steatohepatitis (NASH) s in Male C57BU6J mice Mice are fed a ‘Western’ HF—HSD (high fat-high sucrose diet) or normal chow diet (NCD) as control. The animals are then treated with a compound of Formula (1), Formula (Ia), Formula (Ib), Formula (II), or Formula (III), or a pharmaceutically acceptable salt thereof, for 4, 12 or 20 weeks, and then sacri?ced. Body weight and food intake are monitored weekly and total fat mass is analysed. An intraperitoneal glucose tolerance test (IPGTT) is also performed and tail vein glucose levels are measured after glucose administration. Insulin resistance is calculated using the tasis Model of Insulin Resistance. The mice are then ced by blood sampling via cardiac puncture. Plasma is obtained and tissues were collected together with the plasma for further biochemical and molecular analyses or for histological analysis.
Non-alcoholic fatty liver disease (NAFLD) and non-alcoholic steatohepatitis (NASH) Studies in in nine and Choline De?cient mice Mice weighing 25 g are either fed a methionine- and choline-de?cient diet (MCD to induce NASH) or chow diet (as a control). Animal experiments and evaluation D and NASH are conducted as described above in for C57BL/6J mice fed the high fat and high sucrose diet.
Atherosclerosis Studies in High Cholesterol Fed LDL-R Knockout mice LDL-R knockout (KO) mice are sacri?ced about 12 weeks after the initiation of the genic diet, after which the heart and aorta are perfused with PBS and subsequently ?xed. Atherosclerosis and biochemistry parameters are measured with the riate commercially available kits. For the in vivo lipopolysaccharide (LPS) study, mice are [Annotation] KEB intraperitoneally injected with LPS, and blood is taken from the tail vein. TNFoc levels are ?ed with a Mouse TNFoc ELISA assay. Blood cell counts are determined.
Inherited Mitochondrial Disease Studies in ScoZKO/KI mice Compounds of Formula (1), Formula (Ia), Formula (1b), Formula (II), or Formula (111) are dissolved in water and added to a standard powder diet at the appropriate concentration.
The diet supply is changed every three days and administered ad m for one month.
Tissues are collected for histological analysis. For the muscle quadriceps samples, the spectrophotometric activity of cI, cII, c111, and CW, as well as CS, is measured. NAD+ is extracted from tissues using acidic and alkaline extraction methods, respectively, and analysed with mass spectrometry.
Inherited Mitochondrial Disease Studies in Deletor mice Deletor and WT male mice are administered either chow diet (CD) or a nd of Formula (1), Formula (Ia), Formula (Ib), Formula (II), or Formula (111) admixed with the CD.
The mice are regularly monitored for weight, food ption, and physical endurance and their exercise capability is measured. Oxygen consumption and carbon dioxide production, as well as spontaneous moving and g activities, are recorded. Tissue ns are ted and prepared from the quadriceps, liver, and BAT. Frozen sections from quadriceps are assayed for in situ histochemical COX and succinate dehydrogenase (SDH) activities, crista content in both BAT and muscle is determined from on micrographs and skeletal muscle samples are analysed for citrate synthase activity.
Kidney Disease Studies C57BL/6J WT mice are fed a standard commercial diet and divided into four groups: control; cisplatin; a compound of Formula (1), Formula (Ia), Formula (1b), Formula (II), or a (111), or a pharmaceutically acceptable salt thereof, and cisplatin; and a compound of Formula (1), Formula (Ia), Formula (lb), Formula (II), or Formula (111), or a pharmaceutically acceptable salt thereof, alone. The mice are sacri?ced and tissue samples and serum are collected. Serum creatinine and BUN levels are measured and the lammatory cytokines TNF-a, IL-lb, and IL-6 from serum or homogenates from kidney tissue are quanti?ed. Mouse kidneys are ted and stained for analysis. Tubular damage is examined and scored based on the percentage of cortical tubular necrosis. Neutrophil in?ltration is quantitatively assessed on stained tissue by counting the number of neutrophils per high—power ?eld.
[Annotation] KEB atively, C57BL/6J WT mice are numbered and kept in acclimatization for a period and then ized into different treatment groups based on their body weight.
Different groups are maintained on a speci?ed diet for a period of time. Body weight measurements are taken and food consumption is evaluated. Blood is collected by retro— orbital puncture under mild anesthesia and used for analysis of basal blood urea nitrogen levels (BUN).
Mice are anesthetized and placed on a surgical platform. Both kidneys are exposed through incisions and renal pedicles are occluded using vascular . The clamp is then removed and the surgical site is sutured. The sham—operated group is subjected to similar al procedures, except that the occluding clamp is not applied. Animals are monitored until recovery from esia and returned to their home cage. Animals are observed every day for general clinical signs and symptoms and mortality.
One day prior to termination, animals are individually housed in metabolic cages and urine is collected for estimation of urea, creatinine, sodium and potassium. Blood is also collected by retro orbital puncture under mild anesthesia and plasma is used for analysis of blood urea nitrogen levels (BUN) and serum nine. Animals are then euthanized and organs are collected. One kidney is ?xed and the other is ?ash frozen and used for the estimation of lipid peroxidation, GSH, MPO and SOD levels.
Ischemia/Repeiy‘usion-induced Acute Kidney Injury Studies CD-l (ICR) mice are treated with a compound of Formula (1), Formula (Ia), Formula (lb), a (II), or Formula (III), or a pharmaceutically able salt f, by oral gavage once per day. CD-1 mice are divided into four : (1) young mice with sham injury; (2) young mice with ischemic/ reperfusion (I/R) injury; (3) adult mice with sham injury; and (4) adult mice with I/R injury. An additional 27 adult mice are ized into two groups: mice receiving a compound of Formula (1), Formula (Ia), Formula (1b), Formula (H), or Formula (111), or a pharmaceutically acceptable salt f, and mice receiving the vehicle as a control. The serum creatinine level is measured and BUN ements are recorded. Renal tissue is then evaluated and tubular injury is scored.
Determination of the E?ects 0n F0x0] Phosphorylation levels AML-12 cells are treated with different concentrations of a compound of Formula (1), Formula (Ia), Formula (lb), Formula (II), or Formula (111), or a pharmaceutically acceptable salt thereof. Cells are then lysed, and analyzed by SDS—PAGE/western blot. Blocking and [Annotation] KEB antibody incubations are then done and each n present is detected with its speci?c antibody.
Inhibitory e?ect The t disclosure also relates to a compound of Formula (I), Formula (Ia), Formula (Ib), Formula (II), or Formula (III), or a pharmaceutically able salt f, as de?ned herein, in a method for inhibiting the ty of ACMSD. The method includes contacting a cell with a compound of Formula (I), Formula (Ia), Formula (Ib), Formula (II), or a (III), or a pharmaceutically acceptable salt thereof. In a d embodiment, the method further provides that the compound is present in an amount effective to produce a concentration sufficient to selectively inhibit ACMSD in the cell.
Thus, preferably in an assay for ACMSD inhibition (i.e., an ACMSD assay described herein, e.g., Example 29, or an ACMSD assays known in the literature), the preferred compounds of Formula (I), Formula (Ia), Formula (Ib), Formula (II), or Formula (III), or a pharmaceutically acceptable salt thereof, are compounds capable of ng or preferably inhibiting ACMSD and increasing NAD+ levels and/or activating SIRTs and the downstream targets of SIRTs, such as PGC—la, FoxOl and/or SOD. Preferably, said inhibition is determined as the IC50 of said compound of Formula (I), Formula (Ia), Formula (Ib), Formula (II), or Formula (III) with respect to said ACMSD inhibition assay. Preferred compounds of Formula (I), Formula (Ia), Formula (Ib), Formula (II), or Formula (III), or a pharmaceutically acceptable salt thereof, have an IC50 at or below 1 uM, more preferably less than 300 nM, for example less than 100 nM, such as less than 50 nM with respect to inhibition of ACMSD.
Pharmaceutically acceptable salts The nd of Formula (1), a (Ia), Formula (Ib), Formula (II), or a (III) may be ed in any form suitable for the intended administration, in particular including pharmaceutically acceptable salts, solvates and prodrugs of the compound of Formula (I), Formula (Ia), Formula (Ib), Formula (II), or Formula (III).
Pharmaceutically acceptable salts refer to salts of the compounds of Formula (I), Formula (Ia), Formula (Ib), Formula (II), or Formula (111) which are considered to be acceptable for clinical and/or nary use. Typical pharmaceutically acceptable salts include those salts prepared by reaction of the compounds of Formula (1), Formula (Ia), a (Ib), Formula (II), or Formula (III) and a mineral or organic acid or an organic or inorganic base. Such salts are known as acid addition salts and base addition salts, respectively. It will be recognized that the particular r-ion forming a part of any salt is [Annotation] KEB not of a critical nature, so long as the salt as a whole is pharmaceutically acceptable and as long as the counter—ion does not contribute undesired qualities to the salt as a whole. These salts may be prepared by methods known to the skilled person. Pharmaceutically acceptable salts are, e. g., those described and discussed in Remington's Pharmaceutical Sciences, 17. Ed. o R. Gennaro (Ed.), Mack Publishing Company, Easton, PA, U.S.A., 1985 and more recent editions and in opedia of Pharmaceutical Technology.
Examples of pharmaceutically acceptable addition salts include acid addition salts formed with inorganic acids, e. g., hydrochloric, hydrobromic, sulfuric, nitric, hydroiodic, metaphosphoric, or oric acid; and organic acids e.g., succinic, maleic, , fumaric, , tartaric, benzoic, tri?uoroacetic, malic, lactic, formic, propionic, glycolic, gluconic, camphorsulfuric, isothionic, mucic, gentisic, isonicotinic, saccharic, glucuronic, , glutamic, ic, anthranilic, salicylic, phenylacetic, mandelic, c (pamoic), ethanesulfonic, pantothenic, stearic, sul?nilic, alginic and galacturonic acid; and arylsulfonic, for example benzenesulfonic, p-toluenesulfonic, methanesulfonic or naphthalenesulfonic acid; and base addition salts formed with alkali metals and alkaline earth metals and organic bases such as N,N—dibenzylethylenediamine, chloroprocaine, choline, diethanolamine, ethylenediamine, ine (N-methylglucamine), lysine and ne; and ally formed salts. It should be tood that all references to pharmaceutically able salts include solvent addition forms (solvates) or l forms (polymorphs) as de?ned herein, of the same salt.
The compound of Formula (1), Formula (Ia), Formula (lb), Formula (II), or Formula (III), or a pharmaceutically acceptable salt thereof, may be provided in dissoluble or indissoluble forms together with a pharmaceutically acceptable solvent such as water, ethanol, and the like. Dissoluble forms may also include hydrated forms such as the mono— hydrate, the dihydrate, the drate, the trihydrate, the tetrahydrate, and the like.
The compound of Formula (1), Formula (Ia), Formula (Ib), Formula (II), or Formula (III), or a pharmaceutically acceptable salt thereof, may be provided as a prodrug. The term "prodrug" used herein is intended to mean a compound which — upon exposure to certain physiological conditions — will liberate the compound of Formula (I), Formula (Ia), a (Ib), Formula (II), or Formula (III), or a pharmaceutically acceptable salt thereof, which then will be able to exhibit the desired biological action. A typical example is a labile carbamate of an amine.
Since prodrugs are known to enhance numerous desirable qualities of pharmaceuticals (e.g., solubility, bioavailability, cturing, etc.), the compounds of the present disclosure [Annotation] KEB can be delivered in prodrug form. Thus, the present sure is intended to cover prodrugs of the presently claimed compounds, methods of delivering the same and compositions containing the same. "Prodrugs" are intended to include any covalently bonded carriers that release an active parent drug of the t disclosure in vivo when such prodrug is administered to a t. Prodrugs in the present disclosure are ed by modifying functional groups present in the compound in such a way that the ations are cleaved, either in routine manipulation or in vivo, to the parent compound. Prodrugs include compounds of the present disclosure wherein a hydroxy, amino, sulfhydryl, carboxy or carbonyl group is bonded to any group that may be cleaved in vivo to form a free yl, free amino, free sulfhydryl, free carboxy or free carbonyl group, respectively.
Examples of prodrugs include, but are not limited to, esters (e. g., acetate, dialkylaminoacetates, formates, phosphates, sulfates and benzoate derivatives) and carbamates (e. g., N,N—dimethylaminocarbonyl) of hydroxy functional groups, esters (e.g.,C1_6 alkyl esters, e. g., methyl , ethyl esters, 2-propyl esters, phenyl , 2-aminoethyl esters, morpholinoethanol esters, etc.) of carboxyl functional groups, N—acyl derivatives (e. g., N—acetyl) N—Mannich bases, Schiff bases and enaminones of amino functional groups, , acetals, ketals and enol esters of ketone and aldehyde functional groups in compounds of the disclosure, and the like. See Bundegaard, H., Design ofProdrugs, p1-92, Elesevier, New York—Oxford .
The compounds, or pharmaceutically acceptable salts, esters or prodrugs thereof, are administered orally, nasally, ermally, ary, inhalationally, buccally, sublingually, intraperintoneally, subcutaneously, uscularly, intravenously, rectally, intrapleurally, intrathecally and parenterally. In one embodiment, the compound is administered orally.
One skilled in the art will recognize the advantages of n routes of administration.
The dosage regimen utilizing the compounds is selected in accordance with a variety of factors including type, species, age, weight, sex and medical condition of the patient; the ty of the condition to be treated; the route of administration; the renal and hepatic function of the patient; and the ular compound or salt thereof employed. An ordinarily skilled ian or veterinarian can readily determine and prescribe the effective amount of the drug required to prevent, counter or arrest the progress of the condition.
Techniques for formulation and administration of the disclosed compounds of the disclosure can be found in Remington: the Science and Practice ofPharmacy, 19th edition, Mack Publishing Co., Easton, PA (1995). In an embodiment, the compounds described herein, and the pharmaceutically acceptable salts thereof, are used in pharmaceutical [Annotation] KEB preparations in combination with a pharmaceutically acceptable carrier or diluent. Suitable pharmaceutically able carriers include inert solid ?llers or diluents and sterile aqueous or organic ons. The compounds will be present in such pharmaceutical compositions in s suf?cient to provide the desired dosage amount in the range described .
In one aspect of this disclosure, there is provided a pharmaceutical composition comprising at, as an active ingredient, at least one compound of Formula (I), Formula (Ia), Formula (Ib), Formula (II), or a (III), or a pharmaceutically acceptable salt thereof, as defined herein, and optionally one or more pharmaceutically acceptable excipients, diluents and/or carriers. The compounds of Formula (1), Formula (Ia), Formula (Ib), Formula (II), or a (III), or a pharmaceutically acceptable salt thereof, may be administered alone or in combination with pharmaceutically acceptable carriers, diluents or ents, in either single or multiple doses. Suitable pharmaceutically acceptable carriers, diluents and excipients include inert solid diluents or ?llers, sterile aqueous solutions and various organic solvents.
A "pharmaceutical composition" is a formulation containing the nds of the present disclosure in a form suitable for administration to a subject. The pharmaceutical compositions may be formulated with pharmaceutically acceptable carriers or diluents as well as any other known adjuvants and excipients in ance with conventional techniques such as those disclosed in Remington: The e and Practice of Pharmacy, 21st Edition, 2000, Lippincott Williams & Wilkins.
As used , the phrase "pharmaceutically acceptable" refers to those nds, materials, compositions, rs, and/or dosage forms which are, within the scope of sound medical nt, suitable for use in contact with the tissues of human beings and animals without excessive toxicity, irritation, allergic se, or other problem or cation, commensurate with a reasonable benefit/risk ratio.
"Pharmaceutically acceptable excipient" means an excipient that is useful in preparing a pharmaceutical composition that is generally safe, non—toxic and neither biologically nor otherwise undesirable, and includes ent that is acceptable for veterinary use as well as human pharmaceutical use. A "pharmaceutically acceptable excipient" as used in the speci?cation and claims includes both one and more than one such excipient.
The pharmaceutical compositions formed by combining a compound of Formula (I), Formula (Ia), Formula (Ib), Formula (II), or Formula (III), or a pharmaceutically acceptable salt thereof, as defined herein, with pharmaceutically acceptable rs, diluents or excipients can be readily administered in a variety of dosage forms such as tablets, powders, lozenges, syrups, itories, injectable solutions and the like. In powders, the carrier is a ation] KEB ?nely divided solid such as talc or starch which is in a mixture with the ?nely d active component. In tablets, the active component is mixed with the carrier having the necessary binding properties in suitable proportions and compacted in the shape and size desired.
The pharmaceutical compositions may be speci?cally prepared for administration by any le route such as the oral and parenteral (including subcutaneous, uscular, intrathecal, intravenous and intradermal) route. It will be appreciated that the preferred route will depend on the general condition and age of the subject to be treated, the nature of the condition to be treated and the active ingredient chosen.
Pharmaceutical compositions for oral administration include solid dosage forms such as capsules, tablets, dragees, pills, es, powders, and granules. Where appropriate, they can be ed with coatings such as enteric coatings or they can be prepared so as to e controlled release of the active ingredient such as sustained or prolonged release according to methods well known in the art.
For oral administration in the form of a tablet or capsule, a compound of Formula (I), a (Ia), Formula (Ib), Formula (II), or Formula (III), or a pharmaceutically acceptable salt thereof, as de?ned herein, may suitably be combined with an oral, non-toxic, pharmaceutically acceptable carrier such as ethanol, glycerol, water, or the like. Furthermore, suitable s, lubricants, disintegrating agents, ?avouring agents, and colourants may be added to the mixture, as appropriate. Suitable binders include, e.g., lactose, glucose, starch, gelatin, acacia gum, anth gum, sodium alginate, ymethylcellulose, polyethylene glycol, waxes, or the like. Lubricants include, e. g., sodium oleate, sodium te, magnesium stearate, sodium benzoate, sodium acetate, sodium chloride, or the like.
Disintegrating agents include, e.g., starch, methyl cellulose, agar, ite, xanthan gum, sodium starch glycolate, crospovidone, croscarmellose sodium, or the like. Additional excipients for capsules include macrogels or lipids.
For the preparation of solid compositions such as tablets, the active compound of Formula (I), Formula (Ia), Formula (Ib), Formula (II), or a (III), or a pharmaceutically acceptable salt thereof, is mixed with one or more ents, such as the ones bed above, and other pharmaceutical diluents such as water to make a solid pre—formulation composition containing a homogenous mixture of a compound of Formula (I), Formula (Ia), Formula (Ib), Formula (II), or a (III), or a pharmaceutically acceptable salt thereof.
The term "homogenous" is understood to mean that the compound of Formula (I), Formula (Ia), Formula (Ib), Formula (II), or Formula (III), or a pharmaceutically acceptable salt [Annotation] KEB thereof, is dispersed evenly throughout the composition so that the composition may readily be subdivided into equally ive unit dosage forms such as tablets or capsules.
Liquid compositions for either oral or parenteral administration of the compound of Formula (1), a (Ia), Formula (Ib), Formula (II), or Formula (111), or a pharmaceutically acceptable salt thereof, include, e.g., aqueous solutions, syrups, elixirs, aqueous or oil suspensions and emulsion with edible oils such as cottonseed oil, sesame oil, coconut oil, or peanut oil. Suitable dispersing or suspending agents for aqueous suspensions include synthetic or natural gums such as tragacanth, alginate, acacia, n, sodium carboxymethylcellulose, gelatin, cellulose, or polyvinylpyrrolidone.
Pharmaceutical compositions for parenteral administration include sterile aqueous and non—aqueous able ons, dispersions, suspensions or ons as well as sterile powders to be tituted in sterile inj ectable solutions or dispersions prior to use.
For intravenous administration, suitable carriers e physiological saline, bacteriostatic water, Cremophor ELTM (BASF, Parsippany, NJ.) or phosphate buffered saline (PBS). In all cases, the composition must be sterile and should be ?uid to the extent that easy syringeability exists. It must be stable under the conditions of manufacture and e and must be preserved against the contaminating action of microorganisms such as bacteria and fungi. The carrier can be a solvent or dispersion medium containing, for example, water, ethanol, polyol (for example, glycerol, propylene glycol, and liquid polyethylene , and the like), and suitable mixtures f. The proper ?uidity can be maintained, for example, by the use of a coating such as lecithin, by the maintenance of the ed particle size in the case of dispersion and by the use of surfactants. Prevention of the action of microorganisms can be achieved by various cterial and ngal agents, for example, parabens, chlorobutanol, phenol, ascorbic acid, thimerosal, and the like. In many cases, it will be preferable to include isotonic agents, for example, sugars, cohols such as manitol, sorbitol, and sodium chloride in the composition. Prolonged absorption of the inj ectable compositions can be brought about by including in the composition an agent which delays absorption, for example, aluminum monostearate and gelatin.
The preparation of all these solutions under sterile conditions is readily accomplished by standard pharmaceutical techniques well known to those skilled in the art.
For example, sterile injectable solutions can be ed by incorporating the active compound in the required amount in an appropriate t with one or a combination of ingredients enumerated above, as required, followed by ?ltered sterilization. Generally, ation] KEB dispersions are prepared by incorporating the active compound into a sterile vehicle that contains a basic dispersion medium and the required other ients from those enumerated above. In the case of sterile powders for the preparation of sterile injectable solutions, methods of preparation are vacuum drying and freeze-drying that yields a powder of the active ingredient plus any additional desired ingredient from a previously sterile-filtered solution thereof. Depot injectable compositions are also contemplated as being within the scope of the present sure.
For parenteral administration, solutions containing a compound of Formula (I), Formula (Ia), Formula (Ib), Formula (II), or Formula (III), or a pharmaceutically acceptable salt thereof, in sesame or peanut oil, aqueous propylene glycol, or in sterile aqueous solution may be employed. Such aqueous ons should be suitably buffered if necessary and the liquid diluent first rendered isotonic with sufficient saline or glucose. These particular aqueous solutions are especially suitable for intravenous, intramuscular, subcutaneous and intraperitoneal administration. The oily solutions are suitable for intra-articular, intramuscular and subcutaneous ion purposes.
In addition to the aforementioned ingredients, the compositions of a compound of a (I), Formula (Ia), Formula (1b), Formula (II), or Formula (III), or a pharmaceutically able salt thereof, may include one or more additional ingredients such as diluents, buffers, ?avouring agents, colourant, surface active agents, thickeners, preservatives, e.g., methyl hydroxybenzoate (including xidants), emulsifying agents and the like.
The term "therapeutically effective amount", as used herein, refers to an amount of a pharmaceutical agent to treat, ameliorate, or prevent an identi?ed disease, disorder, or condition, or to exhibit a detectable therapeutic or inhibitory effect. The effect can be detected by any assay method known in the art. The e effective amount for a subject will depend upon the subject’s body weight, size, and health; the nature and extent of the condition; and the therapeutic or combination of therapeutics selected for stration.
Therapeutically effective amounts for a given situation can be ined by routine experimentation that is within the skill and nt of the clinician. In a preferred , the disease or disorder to be d is a disease or disorder associated with a-amino-B— carboxymuconate—s—semialdehyde decarboxylase (ACMSD) dysfunction.
For any compound, the therapeutically effective amount can be estimated lly either in cell culture , e.g., in cells, or in animal , usually rats, mice, rabbits, dogs, or pigs. The animal model may also be used to determine the appropriate concentration [Annotation] KEB range and route of administration. Such information can then be used to determine useful doses and routes for administration in humans. Therapeutic/ prophylactic ef?cacy and toxicity may be determined by standard pharmaceutical procedures in cell cultures or experimental animals, e.g., ED50 (the dose therapeutically effective in 50% of the tion) and LD50 (the dose lethal to 50% of the population). The dose ratio between toxic and therapeutic effects is the therapeutic index, and it can be expressed as the ratio, LDso/EDso.
Pharmaceutical compositions that exhibit large therapeutic indices are preferred. The dosage may vary within this range depending upon the dosage form employed, sensitivity of the patient, and the route of administration.
Dosage and administration are ed to provide sufficient levels of the active agent(s) or to maintain the desired effect. Factors which may be taken into account include the severity of the disease state, l health of the subject, age, weight, and gender of the subject, diet, time and ncy of administration, drug combination(s), reaction sensitivities, and tolerance/response to therapy. Long-acting pharmaceutical compositions may be administered every 3 to 4 days, every week, or once every two weeks depending on ife and clearance rate of the particular formulation.
A suitable dosage of the nd of Formula (1), Formula (Ia), Formula (Ib), Formula (II), or Formula (111), or a pharmaceutically acceptable salt thereof, will depend on the age and condition of the t, the severity of the disease to be treated and other factors well known to the practicing ian. The compound may be stered for example either orally, parenterally or topically according to different dosing schedules, e.g., daily or with intervals, such as weekly intervals. In general a single dose will be in the range from 0.01 to 500 mg/kg body weight, preferably from about 0.05 to 100 mg/kg body weight, more preferably between 0.1 to 50 mg/kg body weight, and most preferably between 0.1 to 25 mg/kg body weight. The compound may be administered as a bolus (i.e., the entire daily dose is administered at once) or in divided doses two or more times a day. Variations based on the aforementioned dosage ranges may be made by a physician of ordinary skill taking into account known considerations such as weight, age, and condition of the person being treated, the severity of the af?iction, and the particular route of administration.
As used , a "subject" or "subject in need thereof’ is a t having a disease or disorder associated with a—amino-B-carboxymuconate-a-semialdehyde decarboxylase ) dysfunction. A "subject" includes a mammal. The mammal can be e.g., any [Annotation] KEB mammal, e.g., a human, primate, bird, mouse, rat, fowl, dog, cat, cow, horse, goat, camel, sheep or a pig. Preferably, the mammal is a human.
The compounds of Formula (1), Formula (Ia), Formula (1b), Formula (II), or Formula (111), or a pharmaceutically acceptable salt thereof, may also be prepared in a pharmaceutical composition comprising one or more further active substances alone, or in ation with pharmaceutically able carriers, diluents, or excipients in either single or multiple doses.
The suitable pharmaceutically acceptable carriers, diluents and excipients are as bed herein above, and the one or more further active substances may be any active substances, or preferably an active substance as described in the section "combination treatment" herein below.
Clinical conditions and other uses ofcompounds The compounds according to Formula (1), Formula (Ia), Formula (1b), Formula (II), or Formula (111), or a pharmaceutically acceptable form thereof, itions, medicaments, and compounds for use, as defined , are useful for treatment of a disease or disorder in which 0t-amino—B-carboxymuconate—a-semialdehyde decarboxylase (ACMSD) modulation plays a role. The compounds may be used either in human or in veterinary medicine and the patient may be any mammal, but especially a human. The treatment may include administering to any mammal, but especially a human, suffering from a disease or disorder in which 0t-amino-B-carboxymuconate-s-semialdehyde decarboxylase (ACMSD) modulation plays a role, a therapeutically ive amount of a nd according to Formula (1), Formula (Ia), Formula (Ib), Formula (II), or Formula (111), or a pharmaceutically acceptable salt thereof, as de?ned herein.
The present disclosure also relates to a compound of Formula (1), Formula (Ia), a (lb), Formula (II), or Formula (111), or a ceutically acceptable salt thereof, as defined herein, for use in a e or disorder ated with oc-amino—B—carboxymuconate- s—semialdehyde decarboxylase (ACMSD) dysfunction, such as obesity, type II es and its complications (e.g., diabetic retinopathy and pathy), non-alcoholic fatty liver disease (NAFLD), non—alcoholic steatohepatitis (NASH), or c kidney disease.
By the term "disease or disorder associated with oc—amino—B—carboxymuconate—e— semialdehyde decarboxylase (ACMSD) dysfunction" is meant any disease characterized by reduced nicotinamide adenine dinucleotide (NADT) expression and/or activity in at least in some instances of the e, or a disease which is ameliorated by elevation of the levels of [Annotation] KEB The s, medicaments and compounds for use of the present disclosure are useful to treat, alleviate the ms of, or delay the onset of a disorder associated with aberrant mitochondrial function. Disorders associated with aberrant mitochondrial function e, for example, metabolic disorders, neurodegenerative disorders, aging related disorders, and chronic in?ammatory disorders. Mitochondrial disorders also include diseases with inherited and/or acquired mitochondrial dysfunction (i.e., Charcot-Marie-Tooth e, Type 2A2, Mitochondrial Encephalopathy Lactic Acidosis and Stroke (MELAS), Leigh syndrome, Barth syndrome, and Leber’s optic neuropathy), fatty acid oxidation disorders, inherited forms of deafness and blindness, and metabolic alities induced by exposure to toxic chemicals and/or drugs (e.g., cisplatin induced deafness).
Metabolic disorders include, for example, type II diabetes, obesity, hyperglycemia, e intolerance, n resistance (i.e., hyperinsulinemia, metabolic syndrome, syndrome X), hypercholesterolemia, hypertension, hyperlipoproteinemia, hyperlipidemia (e.3., dyslipidemia), hypertriglylceridemia, cardiovascular disease, atherosclerosis, peripheral ar disease, kidney disease, ketoacidosis, thrombotic disorders, nephropathy, diabetic neuropathy, diabetic retinopathy, sexual dysfunction, dermatopathy, dyspepsia, hypoglycemia, cancer, and edema.
Neurodegenerative disorders e diseases such as photoreceptor ration (i.e., retinitis pigmentosa), Dementia, Alzheimer’s disease, Parkinson’s disease, and Huntington’s disease.
Chronic in?ammatory diseases include diseases such as celiac disease, itis, lupus, chronic obstructive pulmonary disease (COPD), irritable bowel disease, atherosclerosis, arthritis, and psoriasis.
Aging related disorders include diseases such as cancer, dementia, vascular disease (i.e., arteriosclerosis), ension, diabetes mellitus (type I or type II), arthritis, cataracts, Alzheimer's e, macular degeneration, and osteoporosis.
The subject can be suffering from or susceptible to developing a metabolic disorder.
Subjects ing from or at risk of developing a metabolic disorder are identi?ed by methods known in the art. For example, es can be diagnosed by measuring fasting blood glucose levels or insulin or by glucose tolerance test. Normal adult glucose levels are n about 60-126 mg/dl. Normal insulin levels are about 7 mU/mL : 3mU.
Hypertension can be diagnosed by a blood pressure reading consistently at or above about . Cardiovascular disease can be diagnosed by measuring cholesterol levels. For example, LDL cholesterol above about 137 or total cholesterol above about 200 is indicative [Annotation] KEB of vascular disease. Hyperglycemia can be diagnosed by a blood glucose level higher than about 10 mmol/l (180 mg/dl). Glucose intolerance can be diagnosed by glucose levels of 140 to 199 mg per dL (7.8 to 11.0 mmol) after ting a 75 g oral two—hour glucose tolerance test. Insulin resistance can be diagnosed by a fasting serum insulin level of greater than approximately 60 pmol/L. Hypoglycemia can be diagnosed by a blood e level lower than about 2.8 to 3.0 mmol/L (50 to 54 mg/dl). Obesity can be diagnosed, for example, by body mass index. Body mass index (BMI) is measured in kg/m2 (or lb/in2 X 704.5).
Alternatively, waist circumference (estimates fat bution), waist—to-hip ratio (estimates fat distribution), skinfold ess (if ed at several sites, tes fat bution), or bioimpedance (based on principle that lean mass conducts current better than fat mass (i.e., fat mass impedes current), estimates % fat) can be measured. The parameters for normal, overweight, or obese individuals are as follows: Underweight: BMI < 18.5; Normal: BMI about 18.5 to about 24.9; Overweight: BMI = about 25 to about 29.9. Overweight individuals are terized as having a waist ference of > 94 cm for men or > 80 cm for women and waist to hip ratios of3 0.95 in men and 3 0.80 in women. Obese individuals are characterized as having a BMI of 30 to 34.9, being greater than 20% above "normal" weight for height, having a body fat percentage > 30% for women and 25% for men, and having a waist circumference >102 cm (40 inches) for men or 88 cm (35 inches) for women.
Individuals with severe or morbid obesity are characterized as having a BMI of 3 35.
The methods described herein may lead to a reduction in the severity or the alleviation of one or more ms of a metabolic disorder. For example, symptoms of diabetes include elevated fasting blood glucose levels, blood pressure at or above 140/90 mm/Hg; abnormal blood fat levels, such as high-density lipoproteins (HDL) less than or equal to 35 mg/dL, or triglycerides greater than or equal to 250 mg/dL (mg/dL = milligrams of glucose per deciliter of blood). Ef?cacy of treatment is determined in association with any known method for diagnosing the metabolic disorder. Alleviation of one or more symptoms of the metabolic disorder indicates that the compound confers a al benefit.
The methods of the present disclosure are useful to treat, alleviate the symptoms of, or delay the onset of a kidney disorder. Kidney disorders include acute kidney injury (AKI) and chronic kidney disease (CKD).
The subject can be suffering from or susceptible to developing acute kidney injury (AKI). The acute kidney injury can be characterized by one or more al criteria or conditions (i.e., an abrupt decrease in the ability of the kidneys to excrete enous waste products from the blood, resulting in azotemia). Subjects suffering from or at risk of [Annotation] KEB developing acute kidney injury (AKI) are identi?ed by s known in the art. For example, the acute kidney injury can be characterized by an increase in serum creatinine by at least 50% over baseline, an absolute increase in serum creatinine of at least 0.3 mg/dL over baseline, a reduction in glomerular ?ltration rate of at least 25% compared to baseline, a decrease in urine output to 0.5 ml per kilogram ofbody weight or less per hour ting for at least 6 hours, or any combination thereof. An acute kidney injury may be caused by ischemia, drugs or toxic agents (116., radiocontrast media, a non—steroidal anti—inflammatory drug (NSAID), alcohol, or a chemotherapy agent), viruses, and obstruction.
The subject can be suffering from or susceptible to developing chronic kidney disease (CKD). Chronic kidney disease (CKD) is de?ned as either (1) having kidney damage as de?ned by structural or functional abnormalities of the kidney for 3 months or longer with or without a decreased glomerular ?ltration rate (GFR) or (2) having a GFR of less than 60 mL/min/1.73 m2 for 3 months or longer with or t kidney damage. Subjects suffering from or at risk of developing a chronic kidney disease (CKD) are identi?ed by methods known in the art. Structural or functional abnormalities are manifested by symptoms such as either pathologic alities or markers of kidney damage, including abnormalities identi?ed in imaging studies or the composition of blood or urine.
For example, CKD can be diagnosed by testing for speci?c . For example, markers of kidney damage include a plasma creatinine concentration of above about 1.6 mg/dL and a blood urea nitrogen (BUN) tration of above about 20 mg/dL. Typically, both of these markers are elevated in individuals with CKD. Additional markers of kidney damage can include hematuria (i.e., any detectable amount of blood in the urine), proteinuria (i.e., protein concentrations in urine above about 100 , albuminuria (i.e., albumin concentrations in urine above about 100 mg/dL), an intact parathyroid e (PTH) concentration in the blood above about 150 pg/mL, or blood phosphate levels of above about 4.5 mg/dL. One speci?c marker of kidney disease is a GFR rate above normal (17.6., a GFR above about 90 mL/min/l.73 n12), however a below normal GFR also indicates CKD.
The methods of the present sure are useful to treat, alleviate the symptoms of, or delay the onset of non-alcoholic fatty liver disease WAFLD) and/or non-alcoholic steatohepatitis (NASH). The subject can be suffering from or susceptible to developing non— alcoholic fatty liver disease (NAFLD) and/or non-alcoholic steatohepatitis (NASH). Subjects suffering from or at risk of developing a non-alcoholic fatty liver disease (NAFLD) and/or non-alcoholic hepatitis (NASH) are ?ed by methods known in the art. For e, NAFLD and/ or NASH can be diagnosed by liver biopsy.
[Annotation] KEB Non—alcoholic fatty liver disease (NAFLD), as de?ned herein, is a disease with fat deposition in the liver, which occurs in ts whose alcohol ingestion history is not long enough to cause liver . Non-alcoholic fatty liver disease (NAFLD) can be r ?ed into simple fatty liver, steatohepatitis and cirrhosis. Nonalcoholic steatohepatitis (NASH) refers to a pathology associated with inflammation, liver cell necrosis, ballooning and ?brosis. The onset of nonalcoholic simple fatty liver is induced by fat deposition in liver cells, and this fat accumulation is defined by the e n increasing factors (in?ux and synthesis of fats in liver cells) and decreasing factors (catabolism of fats and their release from liver cells). Once damage of liver cells , in addition to this fat deposition, nonalcoholic simple fatty liver will progress to nonalcoholic steatohepatitis. Nonalcoholic steatohepatitis is progressive and may ?nally ss to cirrhosis and hepatocellular carcinoma.
Combination treatment A nd, compositions, medicaments and compounds for use of Formula (1), Formula (Ia), Formula (Ib), Formula (II), or Formula (111), or a pharmaceutically acceptable salt thereof, may also be used to advantage in combination with one or more other therapeutic agents. Such therapeutic agents include, but are not limited to other ACMSD inhibitors; anti-diabetic agents such as PPARy agonists, PPARoc/y dual agonists, PPARS agonists, biguanides, protein tyrosine phosphatase-1B (PTP-lB), dipeptidyl peptidase IV (DPP-IV) inhibitors, sulfonylureas, meglitinides, alpha glucoside hydrolase inhibitors, alpha-amylase inhibitors, n secreatagogues, A2 antagonists, insulin or n mimetics, glycogen orylase inhibitors, GLP-l agonists, non—thiazolidinediones, glycokinase, and 11 [3 HSD-l inhibitor; anti-obesity agents such as uncoupling Protein (UCP-l, UCP—2, and UCP— 3) activators, B3 adrenergic receptor ([33), thyroid hormone [3 agonists, fatty acid synthase (PAS) inhibitors, phosphodieterase (PDE) inhibitors, lipase inhibitors, serotonin reuptake inhibitors, monoamine reuptake inhibitors, Mc4r agonists, 5HT2c agonists, growth hormone secretagogue (GHS) agonists, CNTF tives, ciliary neurotrophic factors , cholecystokinin-A ) agonists, opioid antagonists, orexin antagonists, acyl-estrogens, leptin, NPY 5 antagonists, neuropeptide Y5 (NPYS) antagonists, neuropeptide Y2 (NPY2) agonists, melanin—concentrating hormone receptor (MCHLR) antagonists and melanin— concentrating hormone 2 receptor (MCHZR), MCHlR antagonists, neuropeptide Yl, ghrelin antagonists, inoid receptor 1 (CB-l), serotonin (5HT) transport inhibitors, CCK—A agonists and histamine 3 (H3) antagonist/inverse agonists; cholesterol lower agents such as 3— [Annotation] KEB hydroxy—3 ~methylglutaryl—coenzyme A (HMG CoA) reductase tors, HMG—CoA synthase inhibitors, squalene epoxidase inhibitors, ?bric acids, bile acid-binding resins probucol and niacin (nicotinic acid) ; compounds that boost NADi levels such as NAD"L precursors (17.6., nicotinamide ribose (NA), nicotinamide mononucleotide WMN), nicotinic acid (NA) and namide); and compounds that inhibit NAD’i ption such as PARP inhibitors and CD38 inhibitors.
PPARy ts useful in the present disclosure include, but are not limited to, glitazones (e. g., balaglitazone, ciglitazone, darglitazone, englitazone, isaglitazone (MCC— 555), pioglitazone, itazone, tazone, CLX-0921, 5-BTZD, and the like); GW—0207, LG-100641, LY—3005 12, LY-5198 18, R483 ), T131 (Tularik), and compounds disclosed in W097/27857, 97/28115, 97/28137 and 97/27847; and pharmaceutically acceptable salts or esters thereof. PPAROt/y dual agonists useful in the present disclosure, include, but are not limited to, CLX—0940, GW-153 6, GW1929, GW-2433, KRP—297, L- 796449, LR—90, MK-0767, SB 219994, and muraglitazar, and pharmaceutically acceptable salts or esters thereof. KRP-297 is 5-[(2,4- 5-thiazolidinyl)methyl]methoxy-N-[[ 4- (tri?uoromethyl) phenyl] methyl]benzamide, and pharmaceutically able salts or esters f. PPARS agonists useful in the present sure include, but are not d to, GW 501516, GW 590735, and compounds disclosed in JP 10237049, and W0 02/14291; and pharmaceutically able salts or esters thereof.
Biguanides useful in the present disclosure include, but are not limited to, buformin, metformin, and phenformin, and pharmaceutically acceptable salts or esters thereof.
Metformin (Glucophage®) is indicated for patients with non-insulin dependent diabetes mellitus, particularly those with refractory obesity. Physician's Desk Reference® page 1080— 1086, (56th ed. 2002).
Protein tyrosine phosphatase—1B (PTP-lB) inhibitors useful in the present disclosure include, but are not limited to, A—401,674, KR 61639, OC—060062, OC—83839, OC—297962, , MC52453, and the compounds disclosed in WO 02/26707, WO 02/26743, JP 2002114768, and pharmaceutically acceptable salts or esters thereof.
Dipeptidyl peptidase IV (DPP-IV) inhibitors, such as cine thiazolidide; NVP— DPP728; P32/98; and LAP 237, P 3298, TSL 225, valine pyrrolidide, TMC—2A/2B/2C, CD- 26 inhibitors, FE 999011, P9310/K364, VIP 0177, DPP4, SDZ 274A444; and the compounds disclosed in WO 03/00449; WO 03/004496; EP 1 258 476; WO 02/083128; WO 021062764; [Annotation] KEB WO 03/000250; WO 530; WO 03/002531; WO 03/002553; WO 03/002593; WO 03/000180; and WO 03/000181.
Sulfonylureas useful in the present disclosure include, but are not limited to, acetohexamide, chlorpropamide, ese, glibenclamide, glipizide, glyburide, glimepiride, gliclazide, tide, gliquidone, glisolamide, tolazamide, and amide, pharmaceutically acceptable salts or esters thereof. Meglitinides useful in the present disclosure include, but are not limited to, repaglinide and inide, and pharmaceutically acceptable salts or esters thereof.
Alpha glucoside hydrolase inhibitors (or glucoside inhibitors) useful in the present sure include, but are not limited to, acarbose, adiposine, camiglibose, emiglitate, miglitol, voglibose, pradimicin-Q, salbostatin, CKD-711, MDL-25,637, MDL—73,945, and MOR 14, and pharmaceutically acceptable salts or esters thereof, and the nds disclosed in US. Pat. Nos. 4,062,950, 4,174,439, 4,254,256, 4,701,559, 4,639,436, 772, 4,634,765, 5,157,116, 5,504,078, 5,091,418, 5,217,877, and 5,091,524. Alpha— amylase inhibitors useful in the present disclosure include, but are not limited to, tendamistat, trestatin, and A1-3688, and pharmaceutically acceptable salts and esters f, and the compounds disclosed in US. Pat. Nos. 4,451,455, 4,623,714, and 4,273,765.
Insulin tagogues useful in the present disclosure include, but are not limited to, linogliride and A—4166, and pharmaceutically acceptable salts and esters thereof.
Fatty acid oxidation inhibitors useful in the present disclosure e, but are not limited to, clomoxir, and ir, and pharmaceutically acceptable salts and esters thereof.
A2 antagonists useful in the present disclosure include, but are not 'limited to, midaglizole, isaglidole, deriglidole, an, earoxan, ?uparoxan, and pharmaceutically acceptable salts and esters thereof. Insulin or insulin mimetics useful in the present disclosure include, but are not limited to, biota, LP-100, novarapid, insulin detemir, insulin lispro, insulin glargine, insulin zinc suspension (lente and ultralente), Lys-Pro insulin, GLP—l (73-7) (insulintropin), and GLP-1 (7-36)—NH2), and pharmaceutically acceptable salts or esters thereof. en phosphorylase inhibitors useful in the present disclosure include, but are not limited to, CP—368, 296, CP-316,819, BAYR3401, and nds disclosed in WO 01/94300, and WO 02/20530, and pharmaceutically acceptable salts or esters thereof. GLP—l agonists useful in the present disclosure include, but are not d to, exendin-3 and exendin—4, and compounds disclosed in US 2003087821 and NZ 504256, and pharmaceutically acceptable salts or esters thereof.
[Annotation] KEB Non-thiazolidinediones useful in the present disclosure include, but are not limited to, JT-501, and farglitazar (GW-2570/GI-262579), and ceutically able salts or esters thereof. Glycokinase tors useful in this disclosure, include, but are not limited to, fused heteroaromatic compounds such as those disclosed in US 2002103199, and isoindolin— 1—one—substituted propionamide compounds such as those disclosed in WO 02/48106.
Serotonin (5HT) transport inhibitors useful in this disclosure include, but are not limited to, paroxetine, ?uoxetine, fen?uramine, mine, sertraline, and imipramine.
Norepinephrine (NE) transport tors useful in this disclosure include, but are not limited to, GW 320659, despiramine, ram, and nomifensine. Cannabinoid receptor 1 (CB-1) antagonist/inverse ts useful in the t sure include: US. Pat. Nos. 5,532,237, 4,973,587, 5,013,837, 122, 5,112,820, 5,292,736, 5,624,941 and US. Pat. No. 084, and PCT Application Nos. WO 96/33159, WO 98/33765, W098/43636, W098/43635, WO 01/09120, WO 98/31227, WO 98/41519, WO 98/37061, WO 00/10967, WO 00/10968, WO 97/29079, WO 99/02499, WO 01/58869, WO 02/076949, WO 01/64632, WO 01/64633, WO 01/64634, and WO 03/007887, and EPO Application No. EP—658546.
Speci?c CB—1 antagonists/inverse agonists useful in the present disclosure include, but are not limited to, rimonabant(Sanof1 Synthelabo), SR-147778 (Sanofi Synthelabo), BAY 65— 2520 (Bayer), and SLY 319 (Solvay). CCK—A agonists useful in the present disclosure include G1 181771, and SR 146,131. Ghrelin antagonists useful in the present disclosure, include: PCT Application Nos. WO 01/87335, and WO 02/08250. Histamine 3 (H3) antagonist/inverse agonists useful in the present disclosure include: PCT Application No. W0 02/ 15905, and O-[3-(1H-imidazol4-yl)propanol]carbamates (Kiec-Kononowicz, K. et al., Pharmazie, 55:349-55 (2000)), piperidine-containing histamine H3 -receptor nists (Lazewska, D. et al., Pharmazie, 56:927-32 (2001), benzophenone derivatives and related compounds (Sasse, A. et al. Arch. Pharm.(Weinheim) 334:45—52 (2001)), substituted N — phenyl carbamates (Reidemeister, S. et al., zie, 55:83-6 (2000)), and proxifan derivatives (Sasse, A. et al., J. Med. Chem. 43:3335—43 ). Speci?c H3 antagonists/inverse agonists useful in the present disclosure include, but are not limited to, thioperamide, 3—(1H-imidazol4-yl)propy1N—4—pentenyl)carbamate, clobenpropit, iodophenpropit, xifan, GT2394 (Gliatech), and A331440.
Melanin-concentrating hormone receptor (MCHLR) nists and melanin- concentrating e 2 receptor (MCH2R) agonist/antagonists useful in the present disclosure include PCT Patent Application Nos. WO 01/82925, WO 01/87834, WO 02/06245, WO 02/04433, and WO 02/51809, and se Patent Application No. JP ation] KEB 13226269. Speci?c MCHlR antagonists useful in the present disclosure include, but are not limited to, T—226296 (Takeda), SB 568849, and SNAP 7941. Neuropeptide Y1 (NPYl) antagonists useful in the present disclosure, include: US. Pat. No. 6,001,836, and PCT Application Nos. WO 96/14307, WO 01/233 87, WO 99/51600, WO 01/85690, WO 98, WO 01/85173, and WO 01/89528. Speci?c examples ofNPY1 antagonists useful in the t disclosure include, but are not limited to, BIBP3226, J-115814, BIBO 3304, LY—357897, CP—671906, and GI—264879A. Neuropeptide Y2 (NPY2) agonists useful in the present disclosure, include, but are not limited to, peptide YY (PYY), and PYY3 _3 6, peptide YY analogs, PYY agonists, and the compounds disclosed in WO 591, WO 03/057235, and WO 03/027637. Neuropeptide Y5 (NPY5) antagonists useful in the present disclosure, e, but are not limited to, the nds described in: US. Pat. Nos. 6,140,354, 6,191,160, 6,258,837, 6,313,298, 6,337,332, 6,329,395, and 6,340,683, US. Pat. Nos. 6,326,375, 6,329,395, 6,337,332, 6,335,345, European Patent Nos. EP—01010691, and EP 01044970, and PCT-International Patent Publication Nos. W0 97/ 19682, WO 97/20820, WO 97/20821, WO 97/20822, WO 97/20823, WO 98/27063, WO 00/107409, 85714, WO 00/185730, WO 00/64880, WO 97, WO 49, wo 01/09120, wo 01/85714, WO 01/85730, WO 01/07409, WO 01/02379, WO 01/02379, WO 01/23388, WO 01/23389, WO 01/44201, WO 01/62737, WO 3 8, WO 20, WO 02/20488, WO 02/22592, WO 02/48152, WO 02/49648, and W0 01/14376. Speci?c NPY5 antagonists useful in the combinations of the present disclosure, include, but are not limited to GW—569180A, GW— 594884A, 081X, GW-548118X, FR 235,208, FR226928, FR 240662, FR252384, 1229U91, GI-264879A, CGP71683A, LY-377897, 77, PD-160170, SR-120562A, SR—120819A, 4, and H409/22. Additional speci?c NPY5 antagonists useful in the combinations of the present disclosure, include, but are not limited to the compounds bed in Norman et a1., J. Med. Chem. 43:42884312 (2000). Leptin includes, but is not limited to, recombinant human leptin (PEG-OB, Hoffman La Roche) and recombinant methionyl human leptin (Amgen). Leptin derivatives (e.g., ted forms o?eptin) useful in the present disclosure include: Pat. Nos. 5,552,524, 5,552,523, 5,552,522, 5,521,283, and PCT International Publication Nos. WO 96/23513, WO 96/23514, WO 96/23515, WO 96/23516, WO 96/23517, WO 96/23518, WO 19, and WO 96/23520.
Opioid nists useful in the present disclosure include: PCT Application No. WO 00/21509. Speci?c opioid antagonists useful in the present disclosure include, but are not limited to, nalmefene (Revex®), 3-methoxynaltrexone, naloxone, and naltrexone. Orexin antagonists useful in the present disclosure include: PCT Patent Application Nos. W0 [Annotation] KEB 01/96302, WO 01/68609, WO 02/51232, WO 02/5183 8, and WO 03/023561. Speci?c orexin nists useful in the present disclosure include, but are not limited to, SB—334867—A.
Acyl—estrogens useful in the present disclosure e oleoyl—estrone (del Mar—Grasa, M. et al., Obesity Research, 9:202-9 (2001)). Cholecystokinin-A (CCK—A) agonists useful in the present disclosure include US. Pat. No. 5,739,106. Speci?c CCK—A agonists include, but are not d to, AR-R 15849, GIl8177l,JMV-l80, A-71378, A-71623 and SR146131. Speci?c ciliary neurotrophic factors (CNTh) useful in the present disclosure include, but are not limited to, GI-18l771 (Glaxo-SmithKline), SR146131 (Sano? Synthelabo), butabindide, PD170,292, PD 149164 (P?zer). CNTF derivatives useful in the present disclosure include, but are not limited to, axokine (Regeneron), and PCT ation Nos. WO 94/09134, WO 98/22128, and WO 13. Growth hormone secretagogue (GHS) agonists useful in the present disclosure include: US. Pat. No. 6,358, 951, and US. Patent Application Nos. 2002/049196 and 2002/02263 7, and PCT Application Nos. WO 01/56592, and WO 02/32888. c GHS ts include, but are not d to, NN703, hexarelin, MK— 0677, SM—130686, CP424 391, L-692,429 and L—163,255. 5HT2c agonists useful in the present disclosure include: US. Pat. No. 250, and PCT Application Nos. WO 02/36596, WO 02/48124, WO 69, WO 01/66548, WO 02/44152, WO 02/51844, WO 02/40456, and WO 02/40457. Speci?c 5HT2c agonists useful in this disclosure e, but are not limited to, BVT93 3, DPCA37215, 1K264, PNU 223 94, 503, R—1065, and YM 348.
Mc4r agonists useful in the present disclosure include: PCT Application Nos. WO 99/64002, WO 00/74679, WO 01/991752, WO 01/74844, WO 01/70708, WO 01/70337, WO 01/91752, WO 02/059095, WO 02/059107, WO 02/059108, WO 02/059117, wo 02/12166, WO 02111715, WO 02/12178, WO 02/15909, WO 3 87, W0 388, WO 02/067869, WO 03/007949, and WO 03/009847. Speci?c Mc4r agonists useful in the present disclosure include CIR86036 (Chiron), 42, and ME—10145 (Melacure).
Monoamine reuptake tors useful in the present sure include: PCT Application Nos. WO 01/27068, and WO 01/62341. Speci?c monoamine reuptake inhibitors useful in the present disclosure include, but are not limited to, sibutramine (Meridia O /Reductil®) disclosed in US. Pat. Nos. 4,746,680, 4,806,570, and 5,436,272, and US. Patent Publication No. 2002/0006964.
Serotonin reuptake inhibitors, and releasers, useful in the present disclosure include: dexfen?uramine, ?uoxetine, and other serotonin reuptake inhibitors, including, but not [Annotation] KEB limited to, those in US. Pat. No. 6,365,633, and PCT Patent ation Nos. WO 01/27060, and WO 01/162341. 11 [3 HSD-l inhibitor useful in the present disclosure include, but are not limited to, BVT 3498, BVT 2733, and those compounds disclosed in WO 91, WO 01/90090, WO 01/90092. Uncoupling n (UCP—l, UCP—2, and UCP—3) tors useful in the present disclosure include: PCT Patent Application No. WO 99/00123. Speci?c uncoupling protein (UCP-l, UCP-2, and UCP—3) activators useful in the present disclosure include, but are not limited to, phytanic acid, 4—[ (E)( 5 ,6, 7,8-tetrahydro-5,5 ,8,8-tetramethyl—2—napthalenyl)- 1-propeny1]benzoic acid (TTNPB), and retinoic acid. [33 adrenergic receptor ([33) agonists useful in the present disclosure include: US. Pat.
No. 5,705,515 and US. Pat. No. 5,451,677 and PCT Patent Application Nos. WO 01/74782, and WO 02/32897. Speci?c [3 agonists useful in the present sure include, but are not d to, AD9677/TAK677 (Dainippon/Takeda), CL—316,243, SB 418790, BRL-37344, L- 796568, BMS—196085, BRL— 35135A, CGP12177A, BTA—243, GW 427353, Trecadrine, Zeneca D7114, and SR 59119A.
Thyroid hormone [3 agonists useful in the present disclosure include: PCT Application No. W0 02/15845 and Japanese Patent ation No. JP 2000256190. Speci?c thyroid e [3 agonists useful in the present disclosure include, but are not limited to, KB—2611 (KaroBioBMS). Speci?c fatty acid synthase (PAS) tors useful in the present disclosure, include, but are not limited to, Cerulenin and C75. Speci?c phosphodieterase (PDE) inhibitors useful in the present sure, include, but are not limited to, ylline, pentoxifY11ine, zaprinast, sildena?l, arnrinone, milrinone, cilostamide, rolipram, and cilomilast.
Lipase inhibitors useful in the present disclosure include, but are not d to, those sed in PCT Application No. WO 01/77094, and US. Pat. Nos. 4,598,089, 4,452,813, ,512,565, 5,391,571, 5,602,151, 4,405,644, 438, and 4,242,453. Speci?c lipase inhibitors useful in the present disclosure include, but are not limited to, tetrahydrolipstatin (orlistat/Xenical®), Triton WR1339, RHC80267, lipstatin, teasaponin, and lumbelliferyl phosphate, , WAY—121898, Bay-N—3176, valilactone, esteracin, ebelactone A, tone B, and RHC 80267.
Examples of HMG—CoA reductase inhibitors include, but are not limited to, lovastatin, simvastatin, pravastatin and ?uvastatin. Examples of HMG-CoA synthase inhibitors are the beta-lactone derivatives disclosed in US. Patents 4,806,564, 4,816,477, [Annotation] KEB 4,847,271, and 475123 7; the beta—lactam derivatives disclosed in US. 4,983,597 and U.S.S.N. 07/540,992 ?led June 20, 1990; and the substituted oxacyclopropane analogues disclosed in European Patent Publication EP 0 411 703. Examples of ne epoxidase inhibitors are disclosed in European Patent Publication EP 0 318 860 and in Japanese Patent Publication 102 169—571A. Examples of LDL—receptor gene inducer molecules are disclosed in US. Patent 5,182,298 ?led March 18, 1991. Other cholesterol lowering agents that may be administered include niacin, ol, ?bric acids (116., clo?brate and gem?brozil), and LDL—receptor gene inducers.
Examples of PARP inhibitors include, but are not limited to, iodonitocoumarin, 5— iodo—6-nitrocoumarin, 3,4—dihydro-S—methyl-isoquinolinone, o—l,8—naphthalimide, 3- methoxybenzamide, 8-hydroxymethyl hydro-quinazolin0ne, 2— {3-[4—(4- ?uorophenyl)—3,6-dihydro—l(2h)-pyridinyl]propyl}methyl-4(3h)— quinazolinone, S-?uoro- 1-[4-(4—phenyl-3,6-dihydropyridin-1(buty1]quinazoline-2,4(lh,3h)-dione, 3-(4-ch10rophenyl) quinoxaline-S-carboxamide, 2-(3‘-methoxyphenyl)benzimidazolecarboxam, benzamide, 3- aminobenzamide, 3-aminophtalhydrazide, and l,5-dihydroxyisoquinoline.
The above—mentioned compounds, which can be used in combination with a nd of Formula (1), Formula (Ia), Formula (1b), a (II), or Formula (111), or a pharmaceutically acceptable salt thereof, can be prepared and administered as described in the art such as in the documents cited above.
The above compounds are only illustrative of the ACMSD inhibitors, anti-diabetic , anti-obesity agents, cholesterol lower agent, compounds that boost NAD+ levels, compounds that inhibit NAD+ consumption that can be used in the compositions of the present disclosure. As this listing of compounds is not meant to be comprehensive, the methods of the present disclosure may employ any anti-obesity agent and any anti-diabetic agent, and are not limited to any particular structural class of compounds.
As used herein, "combination therapy" includes the stration of a nd of the t disclosure, or a pharmaceutically acceptable salt, prodrug, metabolite, rph or solvate thereof, and at least a second agent as part of a speci?c treatment regimen intended to provide the bene?cial effect from the co-action of these therapeutic agents. The bene?cial effect of the combination includes, but is not limited to, a cooperative, e.g., synergistic, effect and/or a pharmacokinetic or pharmacodynamic co-action, or any combination thereof, ing from the combination of eutic agents. Administration of these therapeutic agents in combination typically is carried out over a de?ned time period (usually minutes, hours, days or weeks depending upon the combination selected). nation therapy" [Annotation] KEB may be, but generally is not, intended to encompass the administration of two or more of these therapeutic agents as part of separate monotherapy regimens that ntally and arbitrarily result in the combinations of the present disclosure.
"Combination therapy" is intended to embrace administration of these therapeutic agents in a sequential manner, wherein each therapeutic agent is administered at a different time and in any order, or in alternation and in any order, as well as stration of these therapeutic agents, or at least two of the therapeutic agents, in a substantially simultaneous manner. Substantially simultaneous administration can be accomplished, for example, by administering to the t a single capsule having a ?xed ratio of each therapeutic agent or in le, single capsules for each of the therapeutic agents. Sequential or substantially simultaneous administration of each therapeutic agent can be effected by any appropriate route including, but not limited to, oral routes, enous routes, intramuscular routes, and direct absorption through mucous membrane tissues. The therapeutic agents can be administered by the same route or by different routes. For example, a first therapeutic agent of the combination selected may be administered by intravenous ion while the other therapeutic agents of the combination may be administered orally. Alternatively, for example, all therapeutic agents may be administered orally or all therapeutic agents may be administered by intravenous injection. The sequence in which the eutic agents are stered is not narrowly critical.
All percentages and ratios used herein, unless otherwise indicated, are by weight.
Other features and advantages of the present disclosure will become apparent from the different examples. The provided examples illustrate different components and ology useful in practicing the t disclosure. lly speaking, the disclosure extends to any novel one, or any novel combination, of the features disclosed in this specification (including the accompanying claims and drawings). The examples do not limit the claimed disclosure.
Thus, features, integers, teristics, compounds or chemical moieties described in conjunction with a ular aspect, embodiment or example of the disclosure are to be understood to be able to any other aspect, embodiment or example described herein, unless incompatible therewith. Based on the present disclosure the skilled artisan can identify and employ other components and methodology useful for practicing the present disclosure. Moreover, unless stated otherwise, any feature disclosed herein may be replaced by an alternative e serving the same or a r purpose.
The Disclosure will now be described by way of example only with reference to the Examples below: [Annotation] KEB EXEMPLIFICATION I. ND PREPARATION General Methods and Materials All chemicals were purchased from Sigma-Aldrich, Alfa Aesar. 1H NMR spectra were recorded at 200 and 400 MHz and C NMR spectra were recorded at 100.6 and 50.3 MHz by using deuterated solvents indicated below. TLC were performed on aluminium backed silica plates (silica gel 60 F254). All the reactions were med under nitrogen atmosphere using distilled solvents. All tested compounds were found to have > 95% purity determined by HPLC analysis. HPLC-grade water was obtained from a tandem Milli- Ro/Milli-Q apparatus. The ical HPLC measurements were made on a Shimadzu LC- 20AProminence equipped with a CBM-20A communication bus module, two LC—20AD dual piston pumps, a 0A photodiode array detector and a ne 7725i injector with a uL stainless steel loop.
Scheme 1: ation of Intermediate 1.4 0 NC S NH NC COzEt + JP KCO EtOH + H 2 3’ ’ I ,g H2N NH2 —> N s Reflux 1.1 1.2 1.3 1.4 Exam le 1.‘ Pre aration 0 Intermediate 1.4 To a stirred solution of compound 1.1 (0.52 ml, 4.9 mmol), 1.2 (3 72 mg, 4.9 mmol) and 1.3 (0.5 mL, 0.83 mL, 4.9 mmol) in ethanol (25 mL) was added K2C03 (812 mg, 5.88 mmol). Stirring was continued at re?ux overnight. The pale yellow solid was collected after cooling, taken up with boiling water and d again. The aqueous phase was acidi?ed to pH 5 with AcOH (15 drops), the precipitate was filtered and dried under vacuum. The title compound 1.4 was obtained as pale yellow solid (500 g, 2.18 mmol). Yield 44%.
[Annotation] KEB Scheme 2: Preparation of Intermediate 2.2 0 NC s NH Et + JL + K2C03, EtOH, \ H I A H2N NH2 \ _. \ N s Reflux \ H 1.1 1.2 2.1 22 Exam le 2.‘ Pre n 0 Intermediate 2.2 To a stirred solution of compound 1.1 (0.96 g, 8.8 mmol), 1.2 (672 mg, 8.8 mmol) and 2.1 (lg, 0.83 mL) in ethanol (55 mL) was added K2CO3 (1.57 g, 11.44 mmol). Stirring was ued at re?ux overnight. The yellowish solid was collected after cooling, taken up with hot water and ?ltered again. The aqueous phase was acidi?ed to pH 1, the precipitate was ?ltered and dried under vacuum. The title compound 2.2 was obtained as yellowish solid (1 g, 4.25 mmol). Yield 49%.1H NMR (200 MHz, DMSO) 5 7.22 (m, 1H), 7.68 (m, 1H), 7.85 (d, J: 4.8 Hz, 1H), 8.05 (s, 1H).
Scheme 3: Preparation of Intermediate 3.2 0 NC S NH /\ KCO3’ EtOH NC C02Et + JL 2 + / H ’ | —> /J§ H2N NH2 I / N s S I H Reflux S 1.1 1.2 3.1 3.2 Exam le 3: Pre aration 0 Intermediate 3.2 To a stirred solution of compound 1.1(0.96 mL, 8.8 mmol), 1.2 (672 mg, 8.8 mmol) and 3.1 (lg, 1.29 mL) in l (55 mL) was added K2C03 (1.57 g, 11.44 mmol). Stirring was continued at re?ux overnight. The yellowish solid was collected after cooling, taken up with hot water and ?ltered again. The aqueous phase was acidi?ed to pH 1, the precipitate was ?ltered and dried under vacuum. The title compound 3.2 was obtained as yellowish solid (1 g, 4.25 mmol). Yield 49%.
[Annotation] KEB Scheme 4: Preparation of Intermediate 4.2 /\ iperiine,t NC CO Et + + H , Reflux 1.1 1.2 4.1 4.2 Exam le 4.‘ Pre aration 0 Intermediate 4.2 To a stirred solution of compound 1.1 (1.42 mL, 13.37 mmol), 1.2 (1.01 g, 13.3 mmol) and 4.1 (1.62 mL, 13.3 mL) in l (50 mL) was added piperidine (2.64 mL, 26.7 mmol). Stirring was continued at re?ux overnight. The solid was collected after cooling, taken up with hot water and ?ltered again. The aqueous phase was acidi?ed to le and extracted with EtOAc (3 x 25 mL). The organic phase was washed with brine and dried over NazSO4.The crude of reaction was puri?ed by ?ash chromatography (CHClg/MeOH as gradient, from 0 to 2% for product), affording the title compound 4.2 (930 mg, 3.95 mmol) as white solid. Yield 30%.
Scheme 5: Preparation of Intermediate 5.2 0 NC s NH Et + JL K2003,EtOH, I & + |\ H H2N NH2 —> u S N / I Reflux N / 1.1 1.2 5.1 5-2 Exam le 5.‘ Pre aration 0 Intermediate 5.2 To a stirred solution of compound 1.1 (0.49 mL, 4.67 mmol), 1.2 (355 mg, 4.67 mmol) and 5.1 (0.44 mL, 4.67 mmol) in ethanol (25 mL) was added K2CO3 (773 mg, 5.6 mmol). Stirring was continued at re?ux overnight. The white solid was ted after cooling, dried under vacuum and used for the next step without r puri?cation. The title compound 5.2 was obtained as white solid (300 mg, 1.3 mmol). Yield 29%.1H NMR (400 MHz, DMSO) 8 7.64 (d, J = 4.7 Hz, 2H), 8.78 (d, J= 4.7 Hz, 2H), 12.98 (s, 1H).
[Annotation] KEB Scheme 6: Preparation of Intermediate 6.2 NC COzEt j: NC I EtO'Na", EtOH NH + H2N NH2 N/KS OEt reflux H 6.1 1.2 6.2 Exam le 6: Pre arati0n 0 ediate 6.2 To a stirred solution NaOEt (1.02 mL, 2.73 mmol) in EtOH abs (20 mL) was added compound 6.1 (500 mg, 2.73 mmol) and 1.2 (207 mg, 2.73 mmol). Stirring was continued at re?ux 4 h. The volatiles were removed under vacuum. The crude of reaction was taken up with water and acidi?ed with AcOH. The precipitate was collected dissolved in water, washed with a mixture of CHC13 and MeOH. The aqueous phase was extracted with EtOAc (3 x 20 mL). The collected c phase was washed with brine, dried over NazSO4. The title compound 6.2 was obtained as white solid (250 mg, 1.49 mmol). Yield 55%.
Scheme 7: Preparation of Intermediate 7.3 C80 EtOH NC 0Et NC 8 \ Piperidine K2003, EtOH.
NCACO2Et + ESI / / + JL l O H2N NH2 N\ N/j:S'K+ N Reflux &8 1.17.1 7.2 1.2 7.3 Exam le 7a: Pre aration 0 Intermediate 7.2 To a stirred solution of nd 1.1 (0.14 mL, 1.3 mmol) and 7.1 (150 mg, 1.3 mmol) in EtOH (5 mL) was added piperidine (1 drop). Stirring was continued at room temperature overnight. The solvent was removed under vacuum. The crude of reaction was puri?ed by ?ash chromatography affording the title nd 7.2 (160 mg, 0.77 mmol) as yellowish solid. Yield 58%.
Exam le 71): Pre aration 0 ediate 7.3 To a stirred suspension of compound 7.2 (150 mg, 0.72 mmol) and nd 1.2 (55 mg, 0.72 mmol) in EtOH (5 mL) was added K2C03 (99 mg, 0.72 mmol). Stirring was continued at re?ux overnight. The white precipitate was collected and used as well for the [Annotation] KEB next step without further ation. The title nd 7.3 (150 mg, 0.48 mmol) was obtained as yellowish solid as di—potassium salt. Yield 67%.
Scheme 8: Preparation of Intermediate 8.5 NH NH WMNchos, NH20H*HCI, \©)LNDH C'OCOOEL M6200, H ULWOYOHH NaOH, H20 o H20, EtOH, 80°C; 8.1 8.2 8.3 - N—O NaOH hl :>=o \©/‘\H NBS, AIBN, CCI4, re?ux; Br/U‘LN>=OH 8.4 8",, Exam le 8a: Pre aration 0 Intermediate 8.2 To a stirred on of NH20H*HCl and NaHCO3 in water (7 mL) was gradually added a solution of m-tolunitrile (8.1) (2 mL, 17.0 mmol) in EtOH (13.3 mL). Stirring was continued at 80°C for 4 h. The volatiles were removed under vacuum. The crude of reaction was taken up with water, extracted with EtOAc (3 x 25 mL). The organic phase were collected, washed with brine and dried over Na2SO4 affording the title compound 8.2 (1.5 g, 9 mmol) as white solid. Yield 59%.
Exam le 81?: Pre n 0 Intermediate 8.3 To a solution of compound 8.2 (lg, 6 mmol) in dry acetone (5 mL), was added dropwise at 0 OC EtOCOCl (0.63 mL, 6.6 mmol). Stirring was continued at this temperature for 1h. Then a 5% NaOH solution was added to the mixture. Stirring was continued for additional 1h. The solvent ws removed under vacuum. The crude of reaction was poured in water, extracted with EtOAc (3 x 50 mL). The ted organic phase was washed with brine, dried over Na2S04. The title compound 8.3 (600 mg, 2.7 mmol) was ed as white solid. Yield 45%.
Exam 1e 86: Pre aration 0 Intermediate 8.4 To a solution of compound 8.3 (300 mg, 1.35 mmol) in EtOH abs (5 mL) was added sodium (50 mg) portion wise. Stirring was continued at room temperature for additional 4h.
The reaction was quenched by the addition of MeOH. The t was removed under reduced pressure and the crude was d by ?ash chromatography. The title compound 8.4 (150 mg, 0.85 mmol) was obtained as white solid. Yield 63%.
[Annotation] KEB Exam le 8d: Pre aration 0 Intermediate 8.5 To a suspension of compound 8.4 (326 mg, 1.85 mmol) in CCl4 (10 mL) was added AIBN (60.7 mg, 0.37 mmol) and NBS (493 mg, 2.77 mmol). Stirring was ued at re?ux overnight. The solvent was removed under reduced re. The reaction was taken up with water, extracted with EtOAc (3 x 20 mL) washed with brine and dried over NaZSO4.The crude was puri?ed by ?ash chromatography, eluting with Petroleum ether (Pet. Ether) /EtOAc (30% for product) ing the title compound 8.5 (280 mg, 1.09 mmol) was obtained as white solid. Yield 59%.
Scheme 9: Preparation of Intermediate 9.2 UCOZH NBS, AIBN, Br COZH CCI4, reflux; 9.1 9.2 Examgle 9: Pregaration 0f Intermediate 9.2 To a suspension of nd 9.1 (750 mg, 5 mmol) in CC14 (15 mL) was added AIBN (41 mg, 0.25 mmol) and NBS (933.7 mg, 5.24 mmol). Stirring was continued at re?ux overnight. The solvent was removed under reduced pressure. The reaction was taken up with water, extracted with EtOAc (3 x 20 mL) washed with brine and dried over .The crude was puri?ed by ?ash chromatography, eluting with CHZCIZ/MeOH (3% for product) affording the title compound 9.2 (800 mg, 3.49 mmol) as white solid. Yield 70%.
Scheme 10: Preparation of Intermediate 10.
N’N. N411.
\©///N l + ,N l ,N Et3NH C|-, NaN3 N NBS, AIBN, Br N H —> H toluene, 110°C CH3CN, reflux; .1 10.2 10-3 Exam le 10a: Pre aration 0 Intermediate 10.2 A mixture of compound 10.1 (1.02 mL, 8.54 mmol), NaN3 (832 mg, 12.8 mmol) and Et3N*HCl (1.76 g, 12.8 mmol) was heated at re?ux 4 h. The solvent was d under vacuum. The crude was poured in water, acidi?ed to pH 1 with 3N HCl and extracted with EtOAc (3 x 20 mL). The organic phase was washed with brine, dried over Na2S04 and [Annotation] KEB concentrated under reduced pressure. The title compound 10.2 (1.22 g, 7.6 mmol) was obtained as white solid. Yield 89%.
Exam le 1019: Pre n 0 ediate 10.3 To a suspension of compound 10.2 (300 mg, 1.87 mmol) in CH3CN (15 mL) was added AIBN (31 mg, 0.18 mmol) and NBS (333 mg, 1.87 mmol). Stirring was continued at re?ux overnight. The solvent was removed under reduced re. The reaction was taken up with water, extracted with EtOAc (3 x 20 mL) washed with brine and dried over NazSO4.The crude was purified by ?ash chromatography, eluting with CHZCIZ/MeOH (7% for product) affording the title compound 10.3 (150 mg, 0.62 mmol) as light yellow solid.
Yield 34%.
Scheme 11: Preparation of Intermediate 11.3 OH OH :3 0 CH CI EtN H NH2 2 2’ 3 N + am w 11.1 11.2 11.3 Exam le 1].'Pre aration 0 Intermediate 11.3 To a solution of compound 11.1 (2.5 g, 23 mmol) in CH2C12 (25 mL) was added pyridine (1.63 mL, 20.3 mmol) and compound 11.2 (1.68 mL, 20.3 mmol). Stirring was continued at room ature overnight. The solvent was removed under reduced pressure.
The reaction was taken up with water, extracted with CH2C12 (3 x 30 mL) washed with brine and dried over NazSO4. The crude was puri?ed by ?ash chromatography, eluting with Pet.
Ether/EtOAc (25% for product) affording the title nd 11.3 (735 mg, 3.19 mmol) as brownish solid. Yield 14%.
Scheme 12: Preparation of Intermediate 12.2 o o H 0 EtO‘Na+ EtOH NH OH + Jk I & H2N NH2 N S reflux H 12.1 1.2 12.2 [Annotation] KEB Exam le 12: Pre aration 0 Intermediate 12.2 To a solution of compound 12.1 (2 g, 10.41 mmol) in EtOH (15 mL) was added EtONa (7 mL, 18.7 mmol) and compound 1.2 (1.18 g, 15.61 mmol). Stirring was ued at re?ux ght. The solvent was removed under reduced re. The reaction was taken up with water, acidi?ed to pH 3, extracted with EtOAc (3 x 20 mL) washed with brine and dried over NaZSO4.The crude was puri?ed by ?ash chromatography, eluting with CHzClz/MeOH (2.5% for product) affording the title compound 12.2 (500 mg, 2.44 mmol) as white solid. Yield 24%.
Scheme 13: Preparation of Intermediate 13.2 o o o LDA, THF, EtOAc JOL EtO‘Na’", EtOH \ Cl \ OEt \ \ H N2 NH2 S ‘78 DC S reflux 13.1 13.2 1.2 13-3 Exam le 13a: Pre aration 0 Intermediate 13.2 To a stirred on of DIPA (7.6 mL, 54 mmol) in THF (53 mL) was added n-BuLi (21.6 mL) at 0 OC. Stirring was ued at this temperature 10 minutes. The mixture was then cooled to —78 °C and EtOAc (2.4 mL, 27 mmol) was added se. Stirring was continued at this temperature 30 minutes. After that, a solution of compound 13.1 (3 mL, 27 mmol) in THF (20 mL) was added dropwise. The reaction was allowed to warm to room temperature and was stirred overnight. The crude of reaction was poured in water and extracted with EtOAc (3 x 30 mL). The collected organic phase were washed with brine, dried over NaZSO4 and concentrated under vacuum. The title nd 13.2 was obtained as brownish oil (4.8 g, 24.3 mmol). Yield 90%.
Exam le 13b: Pre aration 0 Intermediate 13.3 To a solution of intermediate 13.2 (2 g, 10 mmol) in EtOH (15 mL) was added EtONa (21% wt/wt in EtOH) (7.5 mL, 20 mmol) and compound 1.2 (1.15 g, 15.1 mmol). Stirring was continued at re?ux overnight. The solvent was removed under reduced pressure. The reaction was taken up with water. At pH 10 was recovered unreacted starting material. The e was then acidi?ed to pH 5, extracted with EtOAc (3 x 20 mL) washed with brine and dried over Na2S04. The crude was puri?ed by ?ash chromatography, eluting with [Annotation] KEB CHzClz/MeOH (7% for product) ing the title compound 13.3 (435 mg, 2.06 mmol) as yellowish solid. Yield 21%.
Scheme 14: Preparation of Compound 1 NC K2003 NH NH I I COZH CH3CN / + Cl/\©/ C02H N SAG N 8 reflux 1.4 3-(Chloromethy|)benzoic acid 1 Exam [6 14: Pre arationo Com ound 1 To a stirred suspension of intermediate 1.4 (1.6 g, 6.98 mmol) and K2C03 (2.88 g, .9 mmol) in CH3CN (80 mL) was added 3—(chloromethyl)benzoic acid (1.19 g, 6.98 mmol). Stirring was continued overnight at re?ux. The volatiles were removed under vacuo.
The crude was taken up with water, acidified to pH 5 and washed with EtOAc to remove impurities. Then the pH was adjusted to 3/4 and the mixture was extracted with EtOAc (3x 50 mL). Titration with hot e afforded compound 1 (936 mg, 2.78 mmol) as yellowish solid. Yield 40%. 1H NMR (400 MHz, DMSO) 5 4.58 (s, 2H), 7.44 (t, J= 7.5 Hz, 1H), 7.54- 7.61 (m, 3H), 7.67 (d, J= 7.1 Hz, 1H), 7.83 (d, J= 7.5 Hz, 1H), 7.91 (d, J= 7.27 Hz, 2H), 8.04 (s, 1H), 13 (s, 1H); 13C NMR (100 MHz, DMSO) 5 33.5, 93.2, 115.6, 128.2, 128.4, 128.4, 128.5, 128.5, 128.6, 129.7, 130.8, 131.5,133.3, 135.1, 137.4, 165.4, 166.8, 167.3.
HPLC: 96.3% Scheme 15: Preparation of nd 4 NC ch03 l co H2 CH30N + CI/U \ N S reflux \ H 2.2 3-Chloromethylbenzoic acid 4 Examgle 15: Pregaration of Compound 4 To a stirred sion of intermediate 2.2 (250 mg, 1.06 mmol) and K2C03 (440 mg, 3.18 mmol) in CH3CN (15 mL) was added 3—(chloromethyl)benzoic acid (180 mg, 1.06 mmol). Stirring was continued overnight at re?ux. The volatiles were removed under vacuo.
The crude was taken up with water, washed with EtOAc, acidified to pH 1 and extracted with EtOAc (3x 50 mL). ion with hot acetone afforded compound 4 (45 mg, 0.12 mmol) as [Annotation] KEB yellowish solid. Yield 12%. 1H NMR (400 MHz, DMSO) 5 4.62 (s, 2H), 7.33 (t, J = 4.3 Hz, 1H), 7.44 (t, J = 7.6 Hz, 1H), 7.72 (d, J = 7.5 Hz, 1H), 7.82 (d, J = 7.5 Hz, 1H), 8.05 (m, 2H), 8.26 (d, J= 3.8 Hz, 1H), 12.99 (s, 1H); 13C NMR (100 MHz, DMSO) 5 33.9, 88.7, 116.5, 128.8, 1293,1299, 130.2, 131.5, 132.1, 1337,1354, 1379, 1397,1590, 1612,1653, 167.4. HPLC: 97.2% Scheme 16: ation of Compound 3 NC K2003 I cozH CH3CN + Cl l N S re?ux 3.2 3—(Chloromethyl)benzoic acid 3 Exam le 16: Pre arationo Com oand 3 To a stirred suspension of intermediate 3.2 (250 mg, 1.06 mmol) and K2CO3 (440 mg, 3.18 mmol) in CH3CN (15 mL) was added 3—(chloromethyl)benzoic acid (180 mg, 1.06 mmol). Stirring was continued ght at re?ux. The volatiles were removed under vacuo.
The crude was taken up with water, washed with EtOAc, acidi?ed to pH 1 and extracted with EtOAc (3x 50 mL). Titration with a mixture of EtZO/acetone ed compound 3 (260 mg, 0.7 mmol) as yellowish solid. Yield 70%. 1H NMR (400 MHz, DMSO) 5 4.63 (s, 2H), 7.44 (t, J= 7.6 Hz, 1H), 7.69 (d, J= 7.7 Hz, 1H), 7.74 (dd, J= 5 Hz, J = 2.9 Hz, 1H), 7.84 (m, 2H), 8.05 (s, 1H), 8.58 (m, 1H), 13.0 (s, 1H); 13C NMR (100 MHz, DMSO) 5 35.3, 90.1, 118.0,130.2,130.7,131.3,131.6,132.9,133.5,135.1,136.8,139.3,141.1,160.4,162.7, 166.7, 168.8. HPLC: 95.0% Scheme 17: Preparation of Compound 6 NC KZCOS NH NH I | A CH3CN + N/st COZH N S reflux H el/UcozH U 4.2 3-(Chloromethy|)benzoic acid 6 Example 17: Preparation of Compound 6 To a d suspension of intermediate 4.2 (250 mg, 1.18 mmol) and K2C03 (495 mg, 3.56 mmol) in CH3CN (15 mL) was added 3—(chloromethyl)benzoic acid (202 mg, 1.18 mmol). Stirring was continued overnight at re?ux. The volatiles were removed under vacuo.
[Annotation] KEB The crude was taken up with water, washed with EtOAc, acidi?ed to pH 1 and extracted with EtOAc (3 x 50 mL). Titration with EtZO afforded compound 6 (90 mg, 0.24 mmol) as white solid. Yield 21%. 1H NMR (400 MHz, DMSO) 5 1.24 (m, 3H), 1.60 (m, 7H), 2.74 (m, 1H), 4.52 (s, 2H), 7.45 (t, J = 7.18 Hz, 1H), 7.67 (d, J = 6.83 Hz, 1H), 7.82 (d, J = 7.17 Hz, 1H), 8.04 (s, 1H), 13.0 (s, 1H).13C NMR (100 MHz, DMSO) 5 25.4, 25.7, 25.7, 30.3, 30.3, 33.8, 44.9, 94.1,115.3,128.6,129.2,130.l, 131.3,133.6, 138.5, 161.1,166.2, 167.4, 177.9. HPLC: 98.1% Scheme 18: Preparation of Compound 7 NC DIPEA NH | A CO H2 DMSO I + Cl \ N/st C02H I H s NI / U .2 3-(Chloromethyl)benzoic acid 7 Example 18: Pregaration of Compound 7 To a stirred suspension of intermediate 5.2 (220 mg, 0.95 mmol) and DIPEA (0.18 mL, 1.05 mmol) in DMSO (5 mL) was added 3—(chloromethyl)benzoic acid (178 mg, 1.05 mmol). Stirring was continued overnight at room temperature. The light yellow solid was collected, washed with crushed ice and water, and dried under vacuum. Trituration with hot EtOAc afforded nd 7 (180 mg, 0.49 mmol) as light yellow solid. Yield 53%. 1 NMR (400 MHz, DMSO) 5 4.55 (s, 2H), 7.44 (m, 1H), 7.66 (d, J= 6 Hz, 1H), 7.81 (m, 3H), 8.02 (s, 1H), 8.80 (s, 2H), 13.1 (s, 1H);13C NMR (100 MHZ, DMSO) 5 34.1, 94.8, 115.8, 122.8, 122.8, 128.7, 129.2, 130.4,131.3, 133.9, 138.1,143.1, 150.5, 150.5, 161.8, 165.7, 167.4, 167.4. HPLC: 95.1% Scheme 19: Preparation of nd 11 K2003 H Br2 9002 Br W/UCOZH?CHSCN AQ/ —’ACOH I CO H2 3: CO H2 N SAG reflux 12.2 3—(Chloromethyl)benzoic [Annotation] KEB Exam le 19a: Pre n 0 Com ound 14 To a stirred suspension of intermediate 12.2 (100 mg, 0.43 mmol) and K2CO3 (178 mg, 1.29 mmol) in CH3CN (15 mL) was added oromethyl)benzoic acid (74 mg, 0.43 mmol). Stirring was continued overnight at re?ux. The volatiles were removed under vacuo.
The crude was taken up with water, washed with EtOAc, acidi?ed to pH 3 and ted with EtOAc (3 x 50 mL). Titration with a mixture of EtZO/Acetone afforded compound 14 (30 mg, 0.088 mmol) as white solid. Yield 21%. 1H NMR (400 MHz, DMSO) 5 4.59 (s, 2H), 6.69 (s, 1H), 7.41 (m, 1H), 7.46 (m, 3H), 7.71 (d, J= 7.5 Hz, 1H), 7.81 (d, J= 7.74 Hz, 1H), 8.06 (m, 3H), 12.85 (s, 2H). 13C NMR (100 MHz, DMSO) 6 33.8, 127.3, 127.3, 128.5, 129.1, 129.2, 130.1, 131.0, 13l.3,131.5, 133.6, 136.3,138.8, 167.5.
Exam [6 19b: Pre arationo Com ound I] To a stirred solution of compound 14 (100 mg, 0.29 mmol) in acetic acid (5 mL) was added lead dioxide (77.2 mg, 0.32 mmol) and bromine (0.02 mL, 0.32 mmol). Stirring was continued for 6 hrs at room temperature. The mixture was poured in a solution ofNazszos and was extracted with EtOAc (3 x 20 mL). The collected organic phases were washed with water and brine, and then they were dried over NaZSO4. Titration with a e of EtZO/Acetone afforded compound 11 (40 mg, 0.09 mmol) as white solid. Yield 33%.1H NMR (400 MHz, DMSO) 5 4.44 (s, 2H), 7.42 (t, J= 7.6 Hz, 1H), 7.48 (m, 3H), 7.61-7.65 (m, 3H), 7.82 (d, J = 7.5 Hz, 1H), 8.0 (s, 1H), 13.1 (m, 2H). 13c NMR (100 MHz, DMSO) 5 33.9, 128.4,128.6,129.14,129.3, 129.3, 129.3,130.1, 130.2,131.3,133.9, 138.4, 167.5.
HPLC: 94.2% Scheme 20: Preparation of Compound 12 0 O 0 K2C03 H Brz PbOz COzH CH3CN AcOH COZH —> COZH \\S +c'/\©/ reflux 13.3 3-(Chloromethyl)benzoic acid N/N:S/\©/ "(NE/U Exam le 200: Pre aration 0 Com ound 15 To a stirred suspension of intermediate 13.3 (235 mg, 1.11 mmol) and K2C03 (460 mg, 3.33 mmol) in CH3CN (15 mL) was added 3—(chloromethyl)benzoic acid (190 mg, 1.11 mmol). Stirring was continued overnight at re?ux. The volatiles were removed under vacuo.
The crude was taken up with water, washed with EtOAc, acidi?ed to pH 3 and extracted with [Annotation] KEB EtOAc (3 x 50 mL). Titration with a mixture of EtZO/Acetone afforded nd 15 (150 mg, 0.44 mmol) as white solid. Yield 39%. 1H NMR (400 MHz, DMSO) 5 4.55 (s, 2H), 6.64 (s, 1H), 7.19 (dd, J= 4.9 Hz, J= 3.8 Hz, 1H), 7.43 (t, J: 7.7 Hz, 1H), 7.74 (d, J= 7.6 Hz, 1H), 7.77 (d, J = 4.9 Hz, 1H), 7.81 (d, J= 7.8 Hz, 1H), 7.90 (d, J = 3.6 Hz, 1H), 8.08 (s, 1H), 12.80 (s, C NMR (100 MHz, DMSO) 5 33.5, 101.6, 128.1, 128.6, 129.1, 129.1, 130.1, 131.1, l31.4,133.7, 138.9,141.7,167.4.
Exam [6 20b: Pre arationo Com ound 12 To a stirred solution of compound 15 (134 mg, 0.39 mmol) in acetic acid (5 mL) was added lead dioxide (102 mg, 0.42 mmol) and bromine (0.022 mL, 0.42 mmol). Stirring was continued for 6 hrs at room temperature. The mixture was poured in a solution ofNazszos and was extracted with EtOAc (3 x 20 mL). The ted organic phases were washed with water and brine, and then they were dried over NazSO4. The crude of reaction was subjected to flash chromatography purification eluting with CHZClg/MeOH (10% for product).
Compound 12 (45 mg, 0.11 mmol) was ed as white solid. Yield 27%. 1H NMR (400 MHz, DMSO) 5 4.56 (s, 2H), 7.27 (t, J= 3.5 Hz, 1H), 7.44 (t, J = 7.6 Hz, 1H), 7.72 (d, J= 7.5 Hz, 1H), 7.82 (d, J = 7.7 Hz, 1H), 7.92 (d, J = 4.5 Hz), 8.07 (s, 1H), 8.33 (d, J= 2.9 Hz, 1H), 13.1 (m, 2H). 13c NMR (100 MHz, DMSO) 5 33.8, 128.6, 128.7, 128.7, 129.2, 130.1, 131.4,132.3,132.8, 133.6,138.3, 141.1, 1521,1585, 1596,1674. HPLC: 95.2% Scheme 21: Preparation of Compound 13 Exam [6 21.'Pre arationo Com our/16113 To a stirred on of compound 14 (100 mg, 0.29 mmol) in acetic acid (5 mL) was added lead dioxide (55.8 mg, 0.35 mmol) and N—chlorosuccinimmide (47 mg, 0.35 mmol).
Stirring was continued for 6 hrs at room temperature. The mixture was poured in water and was extracted with EtOAc (3 x 20 mL). The collected organic phases were washed with water and brine, and then they were dried over NazSO4. ion with a mixture of EtZO/acetone afforded compound 13 (40 mg, 0.1 mmol) as white solid. Yield 37%.1H NMR (400 MHz, DMSO) 5 4.47 (s, 2H), 7.43 (t, J = 7.7 Hz, 1H), 7.49 (m, 3H), 7.64 (d, J = 7.2 Hz, 1H), 7.71 [Annotation] KEB (m, 2H), 7.83 (d, J= 7.5 Hz, 1H), 8.02 (s, 1H), 13.1 (s, 1H), 13.25 (s, 1H); 13C NMR (100 MHz, DMSO) 8 33.9, 1284,1284, 128.6, 1291,1294, 130.2, 131.3, 131.3, 1338,1365, 138.3, 167.5. HPLC: 95.3% Scheme 22: Preparation of Compound 22 0 CI NC NC POCI3, 70 °C CO H \J:/ 2 CO H2 N S/\©/ N s/\©/ 1 22 Exam [6 22.‘ Pre arationo Com ound 22 A stirred suspension of compound 1 (160 mg, 0.44 mmol) and POC13 (3 mL) was heated at 70 °C for 6 h. The white suspension turned red. The excess of POCl3 was carefully destroyed with d ice and then water. The mixture was extracted with EtOAc (3 x 20 mL). The collected organic phase were washed with brine, dried over Na2SO4 and evaporated. Flash chromatography purification (gradient /MeOH) afforded the title compound 22 (60 mg, 0.16 mmol) as white solid. Yield 36%. 1H NMR (400 MHz, DMSO) 6 4.58 (s, 2H), 7.44 (t, J= 7.7 Hz, 1H), 7.58—7.62 (m, 2H), 7.66 (d, J= 7.17 Hz, 1H), 7.70 (d, J = 7.7 Hz, 1H), 7.82 (d, J: 7.7 Hz, 1H), 7.94 (d, J= 7.1 Hz, 2H), 8.08 (s, 1H), 12.98 (s, 1H); 13C NMR(100 MHz, DMSO) 5 34.9, 102.5, 115.2, 128.7, 129.2, 129.2, 129.6, 129.6, 130.4, 131.4, 132.7, 1339, 1347,1380, 162.9, 167.5,169.0, 174.3. HPLC: 98.8% Scheme 23: Preparation of Compound 10 O 0 NCIL): K2003 NC + Cl/\©/ COZH CH3CN | —> /J1" CO H2 H 3 reflux N 3/0 6.2 3- 10 (Chloromethyl)benzoic Exam [6 23: Pre arationo Com ound 10 To a stirred sion of intermediate 6.2 (145 mg, 0.86 mmol) and K2CO3 (599 mg, 4.33 mmol) in CH3CN (15 mL) was added 3—(chlor0methyl)benzoic acid (148 mg, 0.86 mmol). Stirring was continued overnight at re?ux. The les were removed under vacuo.
[Annotation] KEB The crude was taken up with water, washed with EtOAc, acidi?ed to pH 3 and extracted with EtOAc (3 X 50 mL). The crude of reaction was puri?ed by ?ash chromatography, eluting with (CHzClz/ MeOH + ACOH 3%) affording the title compound 10 (60 mg, 0.2 mmol) as white solid. Yield 23%. 1H NMR (400 MHz, DMSO) 8 2.44 (s, 3H), 4.49 (s, 2H), 7.45 (t, J = 7.6 Hz, 1H), 7.68 (d, J: 7.4 Hz, 1H), 7.82 (d, J: 7.6 Hz, 1H), 8.05 (m, 1H), 13.1 (s, 1H); 13C NMR (100 MHZ, DMSO) 5 23.3, 33.9, 95.3, 115.6, 128.7, 129.1, 130.6, 131.2, 134.1, 138.0, 161.1, 165.7, 167.4,170.9.HPLC 96.5% Scheme 24: Preparation of Compound 5 \ ch03 l A COZH CH3CN + Cl <\: N S'K+ reflux 7.3 3—(Chloromethy|)benzoic acid 5 Exam le 24: Pre arationo Com owl/1615 To a stirred suspension of intermediate 7.3 (414 mg, 1.27 mmol) and K2C03 (526 mg, 3.81 mmol) in CH3CN (20 mL) was added 3-(chloromethyl)benzoic acid (217 mg, 1.27 mmol). Stirring was ued overnight at re?ux. The volatiles were removed under vacuo.
The crude was taken up with water, washed with EtOAc, acidi?ed to pH 3 and extracted with EtOAc (3 x 50 mL). The title compound 5 has been obtained (260 mg, 0.7 mmol) as pure light yellow solid after titration with a mixture of EtZO/Acetone. Yield 55%. 1H NMR (400 MHz, DMSO) 5 4.62 (s, 2H), 7.45 (m, 1H), 7.74 (d, J= 5.9 Hz, 1H), 7.82 (d, J = 6.1 Hz, 1H), 8.08 (s, 1H), 8.21 (d, J = 7.2 Hz, 2H), 12.9 (s, 1H); 13C NMR (100 MHz, DMSO) 8 34.0, 90.5,114.9, 128.8, 129.3, 131.4, 133.7,138.1,146.2, 156.7,161.9, 163.8, 166.5, 167.4. HPLC 96.5% Scheme 25: Preparation of Compound 19 NC HN40 DMSO, \ ,0 DIPEA & + Br N —> \ N S \ H 22 8.5 19 [Annotation] KEB Example 25: Preparation of Compound 19 To a stirred suspension of intermediate 2.2 (100 mg, 0.42 mmol) and DIPEA (0.07 mL, 0.47 mmol) in DMSO (5 mL) was added intermediate 8.5 (120 mg, 0.47 mmol). Stirring was continued overnight at room temperature. The crude was poured in water, washed with EtOAc then acidi?ed to pH 3 and extracted with EtOAc (3 x 50 mL). The title compound 19 has been obtained (65 mg, 0.15 mmol) as pure orange solid after ?ash chromatography puri?cation eluting with CH2C12/MeOH (10% for product). Yield 38%. 1H NMR (400 MHZ, DMSO) 5 4.37 (s, 2H), 7.20 (t, J = 4 Hz, 1H), 7.45 (t, J: 7.6 Hz, 1H), 7.63 (t, J: 7.2 Hz, 2H), 7.75 (d, J = 4.3 Hz, 1H), 7.88 (s, 1H), 8.07 (d, J = 3 Hz, 1H); 13c NMR (100 MHz, DMSO) 5 33.7, 85.7, 120.4, 124.9, 125.4, 126.7, 128.7, 128.8, 129.5, 131.1, 132.3, 140.6, 142.2, 159.1, 159.9, 163.3,1704, 171.3. HPLC 94.1%.
Scheme 26: ation of Compound 18 I k 811% + Br c02H , \ u S 2.2 9.2 18 Exam [6 26: Pre arationo Com ound18 To a stirred sion of intermediate 2.2 (500 mg, 0. mmol) and DIPEA (0.4 mL, 2.12 mmol) in DMSO (5 mL) was added intermediate 9.2 (487 mg, 2.12 mmol). Stirring was continued ght at room temperature. The crude was poured in water, washed with EtOAc then acidi?ed to pH 3 and extracted with EtOAc (3 x 50 mL). The title compound 18 has been obtained (200 mg, 0.52 mmol) as pure yellowish solid after ?ash chromatography puri?cation eluting with CH2C12/MeOH (10% for t) and titration with a mixture of EtZO/Acetone. Yield 25%. 1H NMR (400 MHz, DMSO) 5 3.49 (s, 2H), 4.53 (s, 2H), 7.16 (d, J: 6.8 Hz, 1H), 7.26 (t, J: 7.2 Hz, 1H), 7.36 (m, 3H), 8.05 (d, J: 4.4 Hz, 1H), 8.27 (s, 1H), 12.13 (s, 1H); 13C NMR (100 MHz, DMSO) 5 34.3, 40.9, 88.5, 116.8, 127.6, 128.9, 129.1, 129.8, 130.4, 131.9, 1352,1358, 137.0, 139.9,159.1, 161.6, 165.7, 172.9. HPLC 95.8%.
[Annotation] KEB Scheme 27: ation of Compound 17 NC N1. KCO NH ' ,N | n cigcrii + Br N _.
\ N S H reflux \ H 2.2 10.3 17 Exam le 27: Pre arationo Com ound17 To a stirred suspension of intermediate 2.2 (160 mg, 0.66 mmol) and DIPEA (0.09 mL, 0.55 mmol) in DMSO (3 mL) was added intermediate 10.3 (171 mg, 0.55 mmol).
Stirring was continued overnight at room temperature. The crude was poured in water, washed with EtOAc then acidi?ed to pH 3 and extracted with EtOAc (3 x 50 mL). The title nd 17 has been obtained (90 mg, 0.22 mmol) as pure orange solid after ?ash chromatography puri?cation eluting with CHzClz/MeOH (5% for product) and prior titration with a mixture of EtZO/Acetone. Yield 23%. 1H NMR (400 MHZ, DMSO) 5 4.59 (s, 2H), 7.29 (t, J= 4.6 Hz, 1H), 7.54 (t, J: 7.6 Hz, 1H), 7.68 (t, J= 7.8 Hz, 1H), 7.91 (d, J = 7.7 Hz, 1H), 7.96 (d, J = 4.9 Hz, 1H), 8.16 (s, 1H), 8.22 (d, J = 3.8 Hz, 1H); 130 NMR (100 MHz, DMSO) 5 33.9, 88.9, 117.6, 125.0, 126.2, 127.9, 129.6, 129.9, 131.2, 131.9, 134.4, 139.3, 140.4, 155.8, 159.1, 163.8, 167.0. HPLC 96.2%.
Scheme 28: Preparation of Compound 23 NC 0" H KCO NH 2 3’ NH 0H l k N Acetone + Br \ N s MAS/\n/NH \ H o 2.2 11.2 23 Exam le 28: Pre arationo Com out/16123 To a stirred suspension of intermediate 2.2 (150 mg, 0.63 mmol) and K2CO3 (96.6 mg, 0.70 mmol) in acetone (10 mL) was added intermediate 11.2 (173 mg, 0.72 mmol). ng was continued overnight at room temperature. The volatiles were removed under vacuo. The crude was taken up with water, acidi?ed to pH 3 and extracted with EtOAc (3 x 50 mL). The title nd 23 has been obtained (150 mg, 0.39 mmol) as pure orange solid [Annotation] KEB after ?ash chromatography puri?cation eluting with CHzClz/MeOH (5% for t) and prior titration with hot EtOAc. Yield 62%. 1H NMR (400 MHz, DMSO) 5 3.92 (s, 2H), 6.72 (t, J: 7.7 Hz, 1H), 6.79 (d, J = 7.6 Hz, 1H), 6.88 (t, J = 7.3 Hz, 1H), 7.20 (t, J = 4.26 Hz, 1H), 7.77 (d, J = 4.7 Hz, 1H); 7.92 (d, J: 7.8 Hz, 1H), 8.0 (d, J= 3.5 Hz, 1H); 9.59 (s, 1H), 9.80 (s, 1H); 13C NMR (100 MHz, DMSO) 5 35.12, 86.2, 115.5, 119.2, 120.1, 121.0, 124.4, 127.0, 128.7, 129.2, 131.4, 141.8, 147.3, 159.2, 167.9, 169.1, 170.5. HPLC 97.7%. 11. BIOLOGICAL ACTIVITY Example 29: Determination otACMSD] inhibition The activity of compounds 1—19 and 21—23 as inhibitors ofACMSDl was determined by measuring the sion of thrani1ic Acid into product (i. e., ACMS) in a spectrophotometrical in Vitro assay.
The pre—assay mixture consisting of oxyanthranilic acid (3 OH—HA), 3- hydroxyanthranilic acid, 3,4—diOxygenase (HAO), and a dialyzed crude extract of E. coii BL21 (DE3) cells expressing the recombinant enzyme, was incubated at 25 °C with monitoring of the increase in absorbance at 360 nm due to the formation ofACMS from 3OH-HA. After the reaction was completed within ~ 2 mins, an aliquot of ACMSDI solution (prepared and purified from Pichia Pastoris overexpressing the inant enzyme) was added, and the decrease in absorbance at 360 nm was followed at 15 second intervals. The effect ofACMS concentration on the enzyme activity was investigated by varying 3OH-HA tration from 2 to 20 uM. Kinetic parameters were calculated from the initial velocity data by using the aver—Burk plot.
The rate of the decrease in absorbance caused by ACMSDl was calculated by subtracting that of the control reaction mixture without ACMSD from that described above.
One unit ofACMSD activity was indicated as the amount of enzyme that converts 1 mmol of ACMS per minute at 25 °C. The absence or a ion of ACMSDI activity (e. g., by using ACMSD inhibitors) results in a slow pontaneous degradation (i.e., cyclization to form quinolic acid).
The enzymatic activity was determined at a HAA concentration of 10uM in the presence of the compounds in Table 1 below. The compounds were tested at the concentration of about 5 "M and 10uM and the IC50 was calculated for compounds showing tory activity higher than 50%. The results are shown in Table 1. ation] KEB TABLE 1: Act1v1ty. .
Compound N0. Structure hACMSD ICSO 1 I 0.050 [Annotation] KEB Act1v1ty. . nd N0. Structure hACMSD ICSO 12 0.088 [Annotation] KEB Activity nd N0. Structure hACMSD IC50 [Annotation] KEB Activity Compound No. Structure hACMSD IC50 Exam le 30: Determination of ACMSD-l modulation in HEK293T cells HEK293T cells (ATCC) were seeded in six—well plates and transfected using Fugene HD to express transiently ACMSD. 24 hrs post transfection, the cells were stimulated for 48 hrs to 72 hrs with different concentrations of nd 1 and then lysed to measure the ACMSD activity, by measuring the conversion of 3OH—Anthranilic Acid into product (Le. Ot- amino—beta-carboxymuconate-a-semialdehyde, ACMS) in a spectrophotometrical in vitro assay. The amount of the whole protein content in cell lysates was detected by Bradford is. This value was used to get the specificity activity of the enzyme normalized in all samples (mU/ml or AE/At/mg of total protein). l enzyme is known to be expressed in liver, kidney and brain; available cell lines for these cell types were therefore tested to determine the expression levels of ACMSD.
We determined that ACMSD-l is not expressed in transformed cell lines from liver and [Annotation] KEB , such as HepG2, HEK293T, Hep3B etc. ection of ACMSD was performed to express the enzyme in different cellular backgrounds such as COS—7, T, and HepG2.
The HEK293T ar background proved to be the best system, with the highest protein production allowing robust measurement ACMSDl enzyme activity. This is ly due to the better transfection efficacy observed in HEK293T.
Having determined the optimum stimulation time and transfection protocol cells were stimulated with different trations of Compound 1 (about 50 nM to about 5 uM).
Compound 1 inhibited ACMSD-1 activity, in a dose dependent manner, in this over- expression cell—based assay.
Example 31: Determination ofNAD+ content in Human Primam Hepatocytes Treated with Compound 4 The NAD+ concentration or content was determined in human primary hepatocytes d with Compound 4. Vehicle (NT) was used as a control.
At least three experiments were run treating primary hepatocytes with different concentrations of Compound 4 (0.5 uM and 5 uM) after 48 hrs from seeding. nd 4 was replaced every 24 hrs, and then cells were directly harvested and lysed with ACN/HZO (ratio 5:1). LCMS/MS was used to detect and measure NAD+ tration/content.
Screening data showed that Compound 4 inhibits ACMSD-l enzyme at concentrations as low as 0.5uM and SuM. ( Example 32: Determination ofNAD+ content in Human Primam Hepatocytes Treated with Compound 1 The NAD+ concentration or content was determined in human primary hepatocytes treated with Compound 1 and MEHP, a known ACMSD inhibitor. MEHP was used as a control.
At least three experiments were run treating primary hepatocytes with different trations of Compound 1 (0.5 uM, 5 uM, and 50 uM) after 48 hrs from seeding.
Compound 1 was replaced every 24 hrs, and then cells were directly harvested and lysed with ACN/H20 (ratio 5:1). LCMS/MS was used to detect and e NAD+ concentration/content. Screening data showed that SOOuM of MEHP inhibits 70% of purified ACMSD-1 enzyme, and that 0.5uM of Compound 1 has similar inhibition activity as 250uM of MEHP. ( [Annotation] KEB Example 33: Modulation of SOD2 activity in AML-12 cells and Murine Primam Hepatocytes The modulation of SOD-2 activity in AML-12 cells and murine primary hepatocytes treated with either Compound 1 or 17 was measured.
The mouse hepatocytes cell line AML—l2 (alpha mouse liver 12) was obtained from ATCC and grown at 37 0C in a humidi?ed here of 5% % air in Dulbecco’s Modi?ed Eagle Medium / Nutrient Mixture F—12 (DMEM / F-l2) supplemented with 0.005 mg/ml insulin, 0.005 mg/ml transferrin, 5 ng/ml um, 40 ng/ml dexamethasone and 1% ycin. ACMSD inhibitor was initially diluted from powder in DMSO to a stock concentration of 1 mM. This stock was further diluted with water to a concentration of 100 uM which was used for the cell treatments.
Primary hepatocytes were prepared from eek—old C57BL/6J mice by collagenase perfusion method. Mouse livers were perfused with Hank's balanced salt solution (HBSS, KCl, 5.4 mM; KH2P04, 0.45 mM; NaCl, 138 mM; NaHCO3, 4.2 mM; NaZHPO4, 0.34 mM; glucose, 5.5 mM; HEPES, 1 M; EGTA, 50 mM; CaClz, 50 mM; pH 7.4). Livers were then washed at a rate of 5 ml/min h the portal vein. After washing, livers were perfused with collagenase (0.025%) solution. Cell viability was assessed by the trypan blue method. ed primary hepatocytes were plated with DMEM medium (Gibco) ing % FCS, 10 units per m1 penicillin and HEPES for buffering. The cells were maintained in culture at 37 0C in a humidi?ed atmosphere of 5% cog/95% air. After 6-8 hrs of attachment, this medium was replaced with media containing different concentrations of an ACMSD inhibitor (i.e., Compound 1 or Compound 17) or with the corresponding concentration of DMSO (as a control). Primary hepatocytes were harvested about 24 hrs later if not indicated differently.
Primary hepatocytes or AML-12 cells were then lysed in a 20 mM HEPES buffer (Gibco), pH 7.2, containing 1 mM EGTA (Sigma), 210 mM mannitol (Sigma), and 70 mM e (AMRESCO). Total protein concentration was determined using the Bradford assay (BioRad). SOD—2 activity was determined at ted times after ACMSD inhibitor ent by the SOD Assay Kit (Cayman Chemical) according to the manufacturer’s ctions. In order to speci?cally detect the SOD2 activity 2 mM potassium cyanide was added to the assay, which inhibited both Cu/Zn—SOD and extracellular SOD, resulting in the detection of only Mn—SOD (SOD—2) activity. Absorbance was determined with a Victor X4 multi-label plate reader (Perkin-Elmer) at 450 nm. Results are expressed in U/ml/mg of protein according to the standard curve and measured protein concentration.
[Annotation] KEB The oxidative stress resistance pathway, which seemed to be induced upon ACSMD inhibition, was explored by measuring the activity of SOD2. The results showed that SOD2 was induced in a dose-dependent manner in both AML-12 and primary murine cytes by both Compound 17 and Compound 1. In primary hepatocytes, which express ACMSD at signi?cantly higher levels than AML-12, effects were observed at a dose of about 5 nM and reached a maximum at dose of about 50 nM. Both Compound 17 and Compound 1 were able to induce the activity of SOD2 in a dose-dependent manner. ( and ) Example 34: Determination ofNAD+ Content in Murine Primag Hepatocytes NAD+ levels were determined in human primary hepatocytes treated with Compound NAD+ was extracted using acidic extraction method. Samples were collected and nized in 70% ice—cold perchloric acid ). After insoluble protein parts were pelleted by adding ium carbonate (K2C03), the samples were separated by high— performance liquid chromatography (HPLC) and analyzed by mass—spectrometry. The ns in the pellet were quanti?ed by Bradford assay and were used for ization.
The exposure of primary hepatocytes to 5 nM, 10 nM and 50 nM of the ACMSD inhibitor Compound 17 for 24 hours induced a signi?cant and dose-dependent se in intra—cellular NAD+ levels. A icant effect on NAD+ levels was observed at concentrations as low as 5 nM concentration. ( Example 35: RT-gPCR analysis of SIRTl-regplated genes in AML-12 cells, Hepa—l.6 cells and Primam Murine cytes treated with Compound 1 or 17 Gene expression ofACMSD and genes known to be regulated by SIRTl, (an enzyme that is strictly NAD+ dependent) such as PgCJa, Sodl, Sod2 (MnSOD), were analysed in AML-l2 cells, Hepa-l.6 cells and primary murine hepatocytes treated with Compound 1 or Cells (AML-12, Hepa-l .6, HEK-293, primary human and murine hepatocytes) were treated with different concentrations of Compound 1 or Compound 17. Total RNA was extracted from cells using TRIzol (Invitrogen) according to the manufacturer’s ctions.
The RNA was treated with DNase, and 2 ug of RNA was used for reverse transcription (RT). 50X diluted cDNA was used for RT—quantitative PCR (RT-qPCR) reactions. The RT—qPCR ons were performed using the Light-Cycler system (Roche Applied Science) and a [Annotation] KEB qPCR Supermix (QIAGEN) with the indicated primers. The average of at least three technical repeats was used for each biological data point.
A dose—dependent se in mRNA expression levels of genes known to be regulated by SIRTl, (an enzyme that is ly NAD+ dependent) such as chla, Sod2 (MnSOD), but not Sod] (Cu—Zn SOD), was observed in primary mouse hepatocytes treated for 24 hrs with Compound 17 (5 nM - 500 nM range). The observed increase in the gene sion was dose—dependent, which is in line with the dose—dependent increase in SOD2 enzymatic activity observed in Example 32 (. S002 mRNA levels were also increased in a dose-dependent manner in the AML-12 cells and Hepa-1.6 hepatic cell lines after 24 hrs of treatment with Compound 1. These changes in mRNA expression are compatible with the activation of SIRTl, subsequent to the induction in NAD+ levels by inhibition of ACMSDl ty by Compound 17. (and Example 36: Modulation of Caspase 3/7 Activity in MDCK Cells An in vitro study was performed to determine the effects of compounds of Formula (1), Formula (Ia), Formula (Ib), Formula (II), or Formula (111), or a pharmaceutically acceptable salt thereof, on Acute Kidney Injury in MDCK cells.
MDCK cells (MDCK (NBL—2) ATCC® CCL—34TM) were cultured in base medium ATCC—formulated Eagle's Minimum Essential Medium, g No. 30-2003 with fetal bovine serum (FBS) to a ?nal tration of 10%. 10,000 cells were plated into 96 wells and 24 hours after cell g the medium was changed with fresh medium supplemented with 1% FBS. Cisplatin (SOuM for 16 hrs) was then used to induce cell injury. Different concentrations of Compound 18 (in 1% DMSO) were added in combination with cisplatin ( or 1 hour prior adding cisplatin (.
Caspase 3/7 ty (Promega) was determined ing to standard ures using a luminescent signal readout on a Victor V plate reader (PerkinElmer). Each experiment/condition was performed in triplicate.
Caspase activity was analyzed as percentage effect ized to the cisplatin alone (100%) and vehicle treated cells as 0% of caspase activity. Data were analyzed by GraphPad Software. One-way analysis of variance (Dunnett’s Multiple Comparison test) was used for statistical analyses.
As shown in MDCK cells were treated with different concentrations of Compound 18 M to 100uM) in combination with cisplatin (cisp). The EC50 value was [Annotation] KEB ated as equal to 70uM. Cells treated with Compound 18 decreased e activity in signi?cant manner at a concentration of 100uM compared to cells treated with cisplatin alone (p<0.001).
MDCK cells were also treated with different concentrations (1 HM to 125uM) of Compound 18 one hour prior to the addition of cisplatin (cisp). As shown in cells treated with Compound 18 decreased caspase activity in signi?cant manner at a tration of about 30 uM to about 125uM compared to cells treated with tin alone ( p<0.001).
The EC50 value was calculated as equal to 30uM.
Data show that Compound 18 ses, in signi?cant , the activity of caspase 3/7 induced by cisplatin (Fig. 1) the protective effect is particularly noteworthy if Compound 18 is added before insult with the injury agent (cisplatin) as shown in Fig.2.
Example 37: Cytotoxicity and hERG screening Cytotoxicity: 20000 HePG2 and AML-l2 cells were seeded in 96 well plate (Viewplate PerkinElmer). Dose—response of the compound in Table 2 was med using HP D300 digital dispenser, ranging from 10 nM to 300 uM with constant DMSO 1% in . Cells were stimulated for 4 hrs at 37 °C; the atant was used to perform LDH release (Cytotox—one, Promega) as a measure of is while the cells were lysed to detect ATP level for determining cell viability (Celltiter—glo, Promega) ing to manufacturer’s instructions.
The Predictor hERG assay kit (Invitrogen), containing membrane preparations from Chinese hamster ovary cells stably transfected with hERG potassium channel and a high- af?nity red ?uorescent hERG channel ligand (tracer), was used for the determination of hERG channel af?nity binding of the test compounds in Table 2. Compounds that bind to the hERG channel protein (competitors) were identi?ed by their ability to displace the tracer, resulting in a lower ?uorescence polarization. The ?nal concentration of DMSO in each well was maintained at 1%. The assays were performed according to the manufacturer’s protocol (lnvitrogen).
The results are shown in Table 2.
[Annotation] KEB TABLE 2: Cytotoxicity Compound Structure Compound No Activity Toxic No Activity Toxic No Activity No ty No Activity [Annotation] KEB Cytotoxicity Compound Structure Compound No Activity No Activity Exam le 38: Cele ans ex eriments -ACMSDl silencin lifes an assa s t assessment and GFP ?uorescence uanti?cation C. elegans (Caenorhabditis elegans) strains were provided by the Caenorhabditis Genetics Center (University of Minnesota). Worms were maintained on Nematode Growth Medium (NGM) agar plates seeded with Ecoli OP50 bacteria at 20°C, unless stated otherwise. The s used for the experiments were the following: Bristol N2, NL2099 (rrf— 3(pk1426)II), KN259 (huIs33[sod—3:2GFP+pRF4(rol—6(sul006))]). ial feeding RNAi experiments were carried out as follows: worms were grown on NGM agar plates containing Carbenicillin and IPTG at ?nal trations of 25 ug/ml and lmM tively and seeded with bacterial cultures taken from Ahringer library.
Clones used were acmsd—l (Y7lD11A.3), sir—2.1 (R11A8.4), and daf—l6 (R13H8. l). Clones were purchased from GeneSerVice and their identity was con?rmed by sequencing. For the double RNAi experiments bacterial cultures were mixed before seeding on NGM plates. The control RNAi in this kind of experiments was 50% diluted with control empty vector RNAi bacteria.
The nematode Caenorhabditis elegans was used as a model system to con?rm the tion of the oxidative stress defence that we have observed in cells at the level of an intact organism. The effects of acmsd-I RNAi were assessed in C. elegans by RT—qPCR.
[Annotation] KEB The total RNA was extracted from cells using TRIzol (Invitrogen) ing to the cturer’s instructions. RNA was treated with DNase, and 2 pg of RNA was used for reverse transcription (RT). 50X diluted cDNA was used for ntitative PCR (RT- qPCR) reactions. The RT—qPCR ons were performed using the Light—Cycler system (Roche Applied Science) and a qPCR Supermix (QIAGEN) with the indicated primers. The average of at least three technical repeats was used for each biological data point.
C. elegans lifespan assays were carried at 20 °C as follows. Animals were exposed to NAC (N-acetyl cysteine) at a final concentration of 5 mM from a 0.5 M aqueous stock from the young adult stage. Sodium pyruvate was added at a ?nal concentration of 2.5 mM to NGM plates containing carbenicillin (100 lg mL)l) and seeded with UV-killed OP50. After days of RNAi ent, worms were transferred to plates containing paraquat and seeded with acmsd-l RNAi bacteria. Control animals were grown during the first 5 days of adulthood on RNAi bacteria containing the empty vector and then transferred to plates containing paraquat and seeded with acmsd—l RNAi bacteria. Survival analyses were performed using the Kaplan—Meier method, and the significance of differences between survival curves was calculated using the log rank test. The statistical software used was XLSTAT 2007 T, Brooklyn, NY, USA), and all P—values < 0.05 were considered signi?cant. 100 worms were used per condition and scored every 2 days. The reasons for censoring were the ded vulva» phenotype or worms that crawled off the plate. Where indicated, at was added on top of the agar plates at the indicated tration. Once the paraquat on was completely dried, L4 worms were transferred to these agar plates and monitored for 5-6 days every day. By day 6 all the paraquat tests were stopped because a small percentage in worm population could start to die lly and rather than dying due to the paraquat effects.
The movement ofworms was recorded for 45 seconds at days 1, 3, and 5 of adulthood using a Nikon DS—L2 / DS—Fil camera and controller setup, ed to both a computer and a standard bright field microscope. For each condition five plates of worms, with 10 worms per plate were used. The movement of worms during aging was calculated by taking an integral of the speed value which was ed by following the worm centroids with a modified version of the freely-available for the Parallel Worm Tracker for MATLAB.
Fluorescence intensity in worm strains expressing GFP-reporter proteins was quanti?ed using Victor X4 plate reader (Perkin Elmer). The animals were prepared in the following way: eighty worms per condition (at the corresponding ages) were picked (20 [Annotation] KEB worms per well of a walled 96—well plate) and placed into the M9 medium. Each experiment was repeated at least twice.
The expression level of acmsd—J mRNA was signi?cantly reduced con?rming the ef?cacy of RNAi mediated gene knock-down (). The worm ortholog of MnSOD, SOD—3, was d at its mRNA level with concomitant downregulation of the acmsd—I gene with RNAi. A signi?cant increase at the protein level of SOD-3 was also observed at Day 3 of adulthood. () ACMSD downregulation improved worm lifespan and this improvement was SIR—2. 1— and DAF—16—dependent. () Moreover, worms exposed to acmsd—I RNAi lived longer and showed improved performance in mobility assays when treated with paraquat, a well—known ROS inducer that is widely used to mimic oxidative stress in C. elegans. (FIGs. 7D and 7E) The better survival at paraquat conditions was independent on the developmental stage at which worms were d to acmsd—I RNAi. () This increase in lifespan under oxidative stress conditions was no longer observed when , the worm og of FoxOl, was downregulated, meaning that better ive stress resistance was DAF—16 dependent. () Example 39: Study of the Anti-diabetic Effects of Compounds of Formula (1), Formula (Ia), Formula (Ib), Formula (II), or Formula (III) in C57BL/6J and KK-Ay Mice A glucose tolerance test is performed on male C57BL/6J and KK-Ay mice to determine the effects of compounds of Formula (I), a (Ia), Formula (Ib), Formula (II), or Formula (III) on glucose and insulin levels.
Male C57BL/6J and KK—Ay mice, 6—7 weeks of age, are obtained, e.g., from Charles River Laboratories France and CLEA Japan, respectively. Mice are fed from the age of 8 weeks onwards with regular chow (CD—Harlan 2018), a high fat diet (HFD—Harlan 06414).
A compound of Formula (1), Formula (Ia), Formula (Ib), Formula (II), or Formula (III), or a pharmaceutically acceptable salt thereof, is mixed with the HFD at 180 mg kg‘1 of food. On the basis of their daily food intake, this results in a daily dose of about 15 mg kg'1 body weight. The mice are fasted for 4 hrs before blood and tissues are ted for RNA isolation, lipid ements and histology. Oxygen consumption is ed with the Oxymax apparatus (Columbus Instruments). Histological analysis and ission electron microscopy are performed.
[Annotation] KEB An oral e tolerance test is performed in the s that are fasted overnight.
Glucose is administered by gavage at a dose of 2 g/kg. An intraperitoneal insulin tolerance test is performed in animals fasted for 4 hrs. Insulin is injected at a dose of 0.75 U/kg body weight. Glucose is quanti?ed with the Maxi Kit Glucometer 4 (Bayer Diagnostic) or Glucose RTU (bioMerieux Inc.) and plasma insulin concentrations are ed by ELISA (Cristal Chem Inc.). Statistical differences are determined by either ANOVA or Student’s .
Example 40: Study of the Anti-diabetic and Obesity Effects of Compounds of a {I}, Formula (Ia), a (Ib), Formula gII), or Formula (III! in db/db Mice with LepR Mutation A study of the anti-diabetic effects of the compounds of Formula (I), Formula (Ia), Formula (Ib), Formula (II), or Formula (III), or a pharmaceutically acceptable salt thereof, is conducted in genetically obese Leprdb /J (db/db) mice.
Animals are bred and housed in a temperature- and humidity—controlled environment in compliance with FELASA-protocols. From an age of three weeks, mice are fed a high-fat diet (HFD) (Harlan 06414). Most pharmacological studies are started in diabetic eight—week— old db/db and wild type (wt) references.
Subchronic intervention db/db mice are treated once/day with a compound of Formula (I), Formula (Ia), Formula (Ib), Formula (II), or Formula (III), or a pharmaceutically able salt thereof, for 14 days between 5—6 PM before dark-phase onset (6 PM). Blood samples are collected after 4 hrs of fasting the mice prior to the ?rst dose and at 18 i 2 hrs after the last dose. e concentrations of each blood sample are determined.
Acute intervention Glucose l blood samples are collected in random—fed db/db mice between 6—8 AM after light-phase-onset (6 AM), then compounds of Formula (1), Formula (Ia), Formula (Ib), Formula (II), or a (III), or a pharmaceutically acceptable salt thereof, are administered, diet—access is cted, and the second blood sample is collected 4 hrs post-treatment.
Thereafter, mice are subjected to an oral glucose tolerance test (OGTTl: 1 g glucose/kg body mass) and blood glucose concentrations are determined at 0.5, 1, 2, 3, and 4 hrs after each glucose challenge.
[Annotation] KEB Euglycemic-hyperinsulinemic Clamps Assay db/db mice receive a permanent jugular vein catheter under ketamine/xylazine esia. For six to seven days, later (after 6 AM) food—access is restricted. Conscious mice are placed in oversized rat-restrainers and warmed by warming pads. Catheter-ends are then connected to syringes in CMA402—pumps (Axel Semrau, Sprockhoevel, Germany).
After 110 minutes of primed-continuous [3 —3H]glucose on (1.85 kBq/min), a blood sample is collected to ine plasma insulin, glucose and [3—3 H]glucose concentrations and to ate basal endogenous glucose appearance rates. The mice then receive vehicle or a compound of a (1), Formula (Ia), Formula (1b), Formula (II), or Formula (111), or a pharmaceutically acceptable salt thereof, via gavage.
Subsequently, glucose—1 clamps are d with a [3—3H] glucose infusion (3.7 kBq/min) containing insulin (36 pmol/kg*min'l; HumulinR, Lilly, USA) causing a moderate net—increase in plasma insulin concentrations. Blood e concentrations are measured every 10 minutes and target glycemia is established by adjusting the rate of a 20% glucose infusion (GIR). At minute 120, y-D—[1—14 C] glucose (3 70 kBq) is given intravenously.
Blood samples are collected at minute 30, 60, 90, 100, 110, 120, 122, 125, 130, and 140. The mice are then sacrificed (126., through an intravenous ketamine/xylazine-overdose). cnemius muscle and ymal adipose tissue are collected, ately snap—frozen in liquid en, and stored at —80 OC. 2—[14C]deoxyglucose—6—phosphate is extracted from the tissue and glucose uptake rates (Rg) are calculated .
Plasma [3H]- and [14C]-radioactivity is determined in deproteinized plasma after [3HZO] evaporation. Glucose ?uxes under basal conditions and between glucose clamp minute 60 to 90 and 90 to 120 are estimated as follows: whole—body glucose disappearance rate (Rd) = [3—3H]GIR (dpm/min)/plasma g1ucose speci?c activity (dpm/min*mol); basal Endo Ra=[3 -3H]GIR (dpm/min)/plasma [3 -3H]glucose specific activity (dpm/min*mol); glucose—clamp Endo Ra = GIR—Rd. Ultima-Gold scintillation-cocktail, radioisotopes, and a Tri-Carb2910TR are obtained from Perkin Elmer (Germany).
Assaysfrom blood, plasma, urine Blood samples are collected from l tail veins. Blood glucose is measured With a glucometer (Contour, Bayer Vital, Germany), urine and plasma glucose with a colorimetric Glucose LabAssay (Wako, Germany), and HbAlc with AlcNow+ (Bayer Vital) or Clover Analyzer (Inopia, South Korea).
[Annotation] KEB Analyses ofdisease onset and survival Disease onset is de?ned as the last day of individual peak body weight before gradual loss occurs. The stages of disease are de?ned as follows: the early stage of disease is de?ned as the duration of time between peak body weight until loss of 10% of peak body weight. The late stage of disease is de?ned as the duration of time between 10% loss of peak body weight until the end stage of disease. The end stage of disease is de?ned as the day when an animal could no longer right itself within 30 s for three consecutive trials when placed on its side.
Animals are euthanized at the end stage of disease.
Body ition measurements Body weights are assessed weekly for at least 13 weeks. Brown adipose tissue (BAT) and gonadal white adipose tissue (WAT) are dissected and weighed at the indicated age.
Total lean mass, % ofWAT and BMD (bone mineral density) are determined by DEXA (PlXImus DEXA; GE).
Indirect calorimetry, food intake and activity Animals are initially weighed and ated to the test cage. Volume oxygen (V02) and volume carbon dioxide production (VCOz) are measured every 20 min using the Oxymax Comprehensive Laboratory Animal Monitoring System (CLAMS) (Columbus Instruments) and are reported as average V02 per hour ized to body weight kg). Using the CLAMS machine, activity counts by infrared beam uptions and food intake are simultaneously measured. More speci?cally, food intake is measured by deducting the weight of powderized food pellets at the end of experimentation from the starting weight at the beginning of experimentation. To complement this experiment and to l for a novel environment that may affect feeding behaviour, we also perform a more ‘manual’ experiment, wherein a set weight of food pellets is placed at the same time each day into a clean home cage, which holds a mouse. The next day the weight of the remaining pellets is ed and deducted from the ng weight. This ment is performed for 14 days straight. The body weight of each mouse is also recorded daily. Results for each genotype are similar to that acquired from the CLAMS.
Statistical analyses.
Considering a l-B larger than 0.9 statistically powerful, we estimate appropriate group s from pilot studies a priori. One- or two—way Analyses of Variance rroni post—tests) or t—tests are performed.
[Annotation] KEB Example 41: Study of the Effects of Compounds of Formula (1), Formula (Ia), Formula 1b a II or Formula 111 on Non-alcoholic Fat Liver Disease NAFLD and Non-alcoholic Steatohepatitis g SASH) in Mice A study is performed to determine the effects of compounds of Formula (1), Formula (Ia), Formula (1b), Formula (II), or a (111), or a pharmaceutically acceptable salt thereof, on non—alcoholic fatty liver disease (NAFLD) and non-alcoholic steatohepatitis (NASH) in male C57BL/6J fed a high fat and high sucrose diet.
Male C57BL/6J mice (The Jackson Laboratory, Bar Harbor, Maine, USA) are housed under a 14 hrs light—10 hrs dark cycle at 21—23°C and have ad libitum access to water during the entire experiment. From the age of 6 weeks, mice are fed a ‘Western’ HF—HSD with 44.6% of kcal derived from fat (of which 61% saturated fatty acids) and 40.6% of kcal d from ydrates (primarily sucrose 340 g/kg diet) (TD.08811, 45% kcal Fat Diet, Harlan Laboratories Inc., Madison, sin, USA) or normal chow diet (NCD) as control (V1534—000 ssniff lUM-H, ssniff Spezialdiaten GmbH, Soest, Germany). The animals are then treated with a compound of Formula (1), Formula (Ia), Formula (Ib), Formula (II), or Formula (111), or a pharmaceutically acceptable salt thereof, or a control for 4, 12 or 20 weeks (n = 8 per group for every time point), after which they are sacrificed.
Body weight and food intake are monitored weekly on the same day. After sedation with sodium pentobarbital peritoneal injection, 50 mg/kg body weight), total fat mass is analysed by nergy X-ray absorptiometry (DEXA) (PIXImus densitometer, Lunar Corp., Madison, Wisconsin, USA). eritoneal glucose tolerance test (IPGTT) is performed in 6 hrs fasted mice. Tail vein glucose levels are measured with a Bayer r glucometer ately before (time point 0 min) and 15, 30, 60, 90 and 150 min after glucose administration (1 g glucose/kg body weight). Insulin resistance is calculated using the Homeostasis Model of Insulin ance (HOMA-IR) index: (fasting insulin (ng/mL) >< fasting glucose (mg/dL))/405.
Sacrifice After a 6 hrs fasting period, mice are anaesthetised with sodium pentobarbital (intraperitoneal injection, 50 mg/kg body ) and sacrificed by blood sampling via cardiac puncture. Plasma is obtained by centrifugation of blood (6000 rpm for 5 min at 4 °C) that is collected in heparinised syringes. Tissues are either snap frozen in liquid nitrogen or stored at —80°C together with the plasma until further biochemical and molecular analyses or preserved for ogical analysis.
[Annotation] KEB Histological analyses Liver samples are routinely ?xed in buffered formalin (4%) and ed in paraf?n. Serial 4 mm thick sections are stained with H&E and picrosirius red to assess ?brosis. Frozen liver sections are d with Oil Red 0 to assess lipid accumulation. All liver biopsies are analysed by an expert liver pathologist, blinded to the dietary condition or surgical ention. Steatosis, activity and s are semiquantitatively scored according to the NASH-Clinical Research Network criteria. The amount of steatosis (percentage of hepatocytes containing fat droplets) is scored as 0 (<5%), 1 (5—3 3%), 2 (>33—66%) and 3 (>66%). Hepatocyte ballooning is classi?ed as 0 (none), 1 (few) or 2 (many cells/prominent ballooning). Foci of lobular in?ammation are scored as 0 (no foci), 1 (<2 foci per 200>< ?eld), 2 (2—4 foci per 200>< ?eld) and 3 (>4 foci per 200>< ?eld). Fibrosis is scored as stage F0 (no s), stage Fla (mild, zone 3, nusoidal ?brosis), stage Flb (moderate, zone 3, perisinusoidal ?brosis), stage Flc (portal/periportal ?brosis), stage F2 (perisinusoidal and /periportal ?brosis), stage F3 (bridging ?brosis) and stage F4 (cirrhosis). Diagnosis of NASH is based on accepted histological criteria. Severity of the disease is assessed using the NAS (NAFLD activity score) as the unweighted sum of scores of steatosis, cyte ballooning and lobular in?ammation. tage of ?brosis is quantitated by morphometry from digitalised sirius red stained sections using the Aperio system after tuning the threshold of ?brosis detection under visual control. Results are expressed as collagen proportional area.
Example 42: Study of the Effects of Compounds of Formula (I), Formula (Ia), Formula 11b), Formula II or Formula III on Non-alcoholic Fatt Liver Disease AFLD and Non- lic Steatohe atitis ASH in Methionine and Choline De?cient mice A study is performed to determine the effects of compounds of Formula (I), Formula (Ia), Formula (Ib), Formula (II), or Formula (III), or a pharmaceutically acceptable salt thereof, on non—alcoholic fatty liver disease (NAFLD) and non-alcoholic steatohepatitis (NASH) in male pe mice fed a methionine- and e-de?cient diet.
Wildtype mice housed in 12—hour1ight/dark cycles, with free access to food and water are used. At least 5 animals per time point are analysed. All ments are repeated at least three times. For dietary treatment, 8—12 weeks old male mice weighing 25 g are either fed a methionine— and choline-de?cient diet (MCD to induce NASH) or chow diet (as a control). Animal experiments and evaluation ofNAFLD and NASH as described above in Example 40 for mice fed the high fat and high sucrose diet.
[Annotation] KEB Example 43: Study of the Effects of Compounds of Formula (I), Formula (Ia), Formula (Ib), Formula (II), or Formula (III) on Atherosclerosis in High Cholesterol Fed LDL-R Knockout A study is performed to determine the effects of compounds of Formula (I), Formula (Ia), Formula (Ib), Formula (II), or Formula (III), or a pharmaceutically acceptable salt thereof, on atherosclerosis in high cholesterol fed LDL—R knockout mice.
LDL—R knockout (KO) mice are backcrossed for ten generations with the C57BL/6J strain, yielding congenic C57BL/6J animals. The controls that are used are littermates in all experiments. The animals are treated with a compound of a (I), Formula (Ia), Formula (Ib), Formula (II), or a (III), or pharmaceutically acceptable salt thereof, or a l.
Mice are sacrificed 12 weeks after the initiation of the atherogenic diet (TD94059; Harlan), after which the heart and aorta are perfused with PBS and subsequently ?xed on Formal Fixx, Thermo Scienti?c). Atherosclerosis is assessed by an Oil red O staining of the aortic root and quantified with MetaMorph software. Biochemistry ters are measured with the appropriate kits in the COBAS C111 (Roche). For the in viva lipopolysaccharide (LPS) study, mice are eritoneally ed with 100 mg of LPS, and blood is taken from the tail vein. TNFoc levels are quanti?ed with Mouse TNFoc ELISA Ready—SET-Go! (eBioscience) assay. Blood cell counts are determined with Advia2120 (Siemens Healthcare Diagnostics).
The Student’s t test is used to calculate the statistical icance. In case of multiple testing (i.e., the comparison of more than two groups), this test is preceded by the ANOVA test. P < 0.05 is considered statistically cant. Results represent the mean :: SEM.
Example 44: Study of the Effects of Compounds of a 11), a (Ia), Formula (Ib), Formula (II ), or Formula (III) on ted Mitochondrial Disease in ScoZKO/KI mice A study is performed to determine the effects of compounds of Formula (1), Formula (Ia), Formula (Ib), Formula (II), or Formula (III), or a pharmaceutically acceptable salt KO/KI thereof, on inherited mitochondrial disease in Sco2 mice.
Anti-COI, anti-COXSa, anti-Ndufa9, anti-SDH-HA, and anti-Core 2 are from Invitrogen; anti-GAPDH is from Millipore; anti—FoxOl and anti-acetylated-FoxOl are from Cell Signaling and Santa Cruz, respectively. Anti—mouse secondary antibodies are from am. Chemicals are from Sigma. Oligonucleotides are from PRIMM, Italy.
[Annotation] KEB Compounds of Formula (1), a (la), Formula (lb), Formula (II), or Formula (111), or a pharmaceutically acceptable salt f, are dissolved in water and added to a standard powder diet (Mucedola, Italy) at the appropriate tration of 50 mg/Kg/day.
Pellets containing the compounds of Formula (1), Formula (Ia), Formula (Ib), Formula (II), or Formula (111), or a pharmaceutically acceptable salt thereof, or the vehicles are reconstituted by hand and kept frozen at - 20°C until needed. The diet supply is d every three days, and only the amount needed is thawed at each time and administered ad libitum for one month. ScoZKO/KI mice are maintained in a temperature- and humidity-controlled animal—care facility, with a 12 hrs light/dark cycle and free access to water and food. Animals are ced by cervical dislocation.
Morphological Analysis For hemical analysis, tissues are frozen in liquid-nitrogen precooled isopentane.
Series of 8 mm thick sections are stained for COX and SDH.
Biochemical Analysis ofMRC Complexes Muscle quadriceps samples stored in liquid nitrogen are homogenized in 10 mM phosphate buffer (pH 7.4), and the spectrophotometric activity of CI, CH, 0111, and cIV, as well as CS, is measured as bed. Note that in all panels the activity of CH is multiplied by 10 for Visualization clarity.
NAD+ Determination NAD+ is extracted using acidic and alkaline extraction methods, respectively. Tissue NAD+ is analysed with mass spectrometry as usly described.
Example 45: Study of the Effects of Compounds of Formula 11), Formula 11a), Formula 11b), Formula II or Formula 111 on Inherited Mitochondrial Disease in Deletor mice A study is performed to determine the effects of compounds of Formula (1), Formula (Ia), Formula (lb), Formula (II), or Formula (III), or a pharmaceutically acceptable salt thereof, on inherited mitochondrial e in Deletor mice.
The Deletor mouse model is generated in C57BL/6 ic ound and has been previously terized (Tyynismaa et a1, 2005); WT mice are littermates from the same congenic mouse strain C57BL/6J. Deletor and WT male mice are administered either chow diet (CD) or a compound of Formula (1), Formula (Ia), Formula (Ib), Formula (II), or [Annotation] KEB Formula (III), or a pharmaceutically acceptable salt thereof, admixed with the CD at the appropriate concentration. The food pellets are ly prepared by mixing a compound of Formula (I), Formula (Ia), Formula (lb), Formula (II), or Formula (III), or a pharmaceutically acceptable salt thereof, into the powdered food as described for the ScoZKO/KI mice in Example 43 and stored at -20°C. The mice are housed in standard animal facility, under a 12 hrs ight cycle. They have ad libitum access to food and water. The pre-manifestation group consists of 12 Deletors and 12 WT mice, and the post-manifestation group of 24 Deletors and 24 WT mice, receiving either a compound of Formula (1), a (Ia), Formula (Ib), Formula (II), or Formula (III), or a pharmaceutically acceptable salt thereof, or CD diet. During the intervention, the mice are regularly red for weight, food consumption, and al endurance. Their exercise capability is measured twice by treadmill se test (Exer-6M ill, Columbus Instrument) at the start and the end of the diet. The exercise test protocol consists of the initial running speed of 7 m/s which is increased every 2 min by 2 m/s and continued until the animal is unable to run or repeatedly falls from the belt at the stimulus site.
Oxygen ption and carbon dioxide production, as well as spontaneous moving and feeding activities, are recorded by Oxymax Lab Animal Monitoring System (CLAMS; us Instruments, OH, USA). The mice are kept in individual cages inside a CLAMS chamber for 3 days; the first day and night is a nonrecording adjustment period followed by a 24 hrs recording at thermoneutrality (+30 °C). The results of Oz consumption and C02 production are used to calculate respiratory exchange rate and analysed separately from the light (inactive) and dark (active) periods of the day.
Morphologic analysis Tissue sections are prepared from the quadriceps, liver, and BAT. Samples are embedded with OCT Compound Embedding Medium e—Tek) and snap—frozen in 2— methylbutane in liquid nitrogen. Frozen sections (12 lm) from quadriceps are assayed for in situ histochemical COX and succinate dehydrogenase (SDH) activities simultaneously. The activities from the quadriceps sections, the COX—negative and the COX-negative plus SDH ve and normal ?bres are calculated. imately 2000 fibres are counted from each mouse sample. The intensity of COX hemical activity from quadriceps for both oxidative and non—oxidative ?bres is measured with Image J software. Frozen sections (8 pm) from liver and BAT are stained with Oil Red 0. For plastic embedding, quadriceps, liver, and BAT samples are fixed in 2.5% glutaraldehyde, treated with 1% osmium tetroxide, [Annotation] KEB ated in ethanol, and embedded in epoxy resin. Semi-thin (l um) ns are stained with methyl blue (0.5% W/v) and boric acid (1% w/v). The interesting areas for the ultrastructural analyses are selected by inspection of the light microscopic sections. For transmission electron microscopy, ultrathin (60—90 nm) sections are cut on grids and stained with uranyl acetate and lead citrate and viewed with a ission Electron Microscope.
Crista content in both BAT and muscle is determined from electron micrographs, ing a l um "intra-mitochondrial ing stick," placed perpendicular to cristae. Skeletal muscle samples are also analysed for citrate synthase activity.
Example 46: Study of the Effects of Compounds of Formula 11), Formula 11a), Formula 91b), Formula {II}, or Formula (III) on Kidney Disease A study is performed to determine the s of compounds of Formula (1), Formula (Ia), Formula (Ib), Formula (II), or Formula (III), or a pharmaceutically acceptable salt f, on kidney disease in C57BL/6J WT mice. (Wei, Q., et al.. "Mouse model of ischemic acute kidney : technical notes and tricks" an Journal of Physiology—Renal Physiology, 303(11), Fl487—Fl494) C57BL/6J WT mice are purchased from Charles—River. All mice are fed a standard commercial diet While housed at an ambient temperature of 20—22 0C with a relative humidity of 50 i 5% under 12/12 hrs light—dark cycle in a speci?c pathogen—free facility. The experimental mice are 8 weeks old and are divided into four groups: control (n = 5); cisplatin (20 mg/kg; Sigma Chemical, St Louis, MO; n = 5); a compound of Formula (1), Formula (Ia), Formula (Ib), Formula (II), or Formula (III), or a pharmaceutically acceptable salt thereof, and tin (n = 5); and a compound of Formula (1), Formula (Ia), a (Ib), Formula (II), or Formula (III), or a pharmaceutically able salt thereof, alone (40 mg/kg; n = 5).
The dose and time of cisplatin treatment for nephrotoxicity are chosen ing to a published method. A compound of Formula (1), Formula (Ia), Formula (Ib), Formula (II), or Formula (III), or a pharmaceutically acceptable salt thereof, is stered orally once a day for 4 days. Cisplatin is injected once at 12 hrs after the ?rst administration of a compound of Formula (I), Formula (Ia), Formula (Ib), Formula (II), or Formula (III), or a pharmaceutically acceptable salt thereof. The mice are sacrificed at 72 hrs after the single cisplatin injection.
Assaysfor unctional markers andproin?ammatory cytokines For renal function analysis, serum is isolated and stored at —80 0C until use. Serum creatinine and BUN levels are measured using an assay kit according to the manufacturer’s [Annotation] KEB instructions (BioVision, Milpitas, CA). In addition, the proin?ammatory cytokines TNF—OL, lL-lb, and IL-6 from serum or homogenates from kidney tissue are quanti?ed by ELISA ikine Kit; R&D Systems, Minneapolis, MN) according to the manufacturer’s instructions. For measuring cytokines, kidney tissue is homogenized in phosphate buffered saline ning 0.05% Tween-20. Aliquots containing 300 mg of total protein are used. A metabolic cage is used for collecting urine to analyse the level of urinary cytokines. The sample size for each group is five.
Alternative Study ofthe s of Compounds ofFormula (I), Formula (Ia), Formula (lb), Formula (II), or Formula (111) on Kidney Disease Alternatively, C57BL/6J WT mice are numbered and kept in acclimatization for a period of 5—7 days before tion of the experiment. (Wei, Q., et al.. "Mouse model of ic acute kidney injury: technical notes and tricks" American Journal of Physiology— Renal Physiology, 303(11), F1487-F1494) Mice are randomized into different treatment groups based on their body weight. Different groups are maintained on Harlan diet 2916.
Mice are then maintained on the respective diets for 10 days prior to bilateral Ischemic kidney injury. Body weight ement is made once at randomization and once on day 7.
Food consumption is evaluated once on day 7. Blood is collected by retro—orbital puncture under mild Isoflurane anesthesia and used for analysis of basal blood urea nitrogen levels (BUN) on day 9.
Mice are anesthetized with ketamine (80 mg/kg i.p.) and/or Xylazine (10 mg/kg, i.p.) and placed on a surgical platform in a dorsal on. Both kidneys are d through ?ank incisions and renal pedicles are ed using vascular clamps for 25 minutes. The clamp is then removed and the al site is sutured. lml of physiological saline is administered intra—peritoneally after closing the wound to prevent dehydration. The sham- operated group is subjected to similar al procedures, except that the occluding clamp is not applied. Animals are red until recovery from anesthesia and returned to their home cage. Animals are ed every day for general clinical signs and symptoms and mortality.
One day prior to termination, animals are individually housed in metabolic cages for 12h and urine is collected for estimation of urea, creatinine, sodium and potassium.
On days 12, 14, & 16 blood is collected by retro orbital puncture under mild isoflurane anesthesia and plasma is used for analysis of blood urea nitrogen levels (BUN) and serum creatinine. Animals are then euthanized by C02 inhalation and organs are collected.
One kidney is fixed in 10% neutral buffered formalin and the other is ?ash frozen in liquid [Annotation] KEB nitrogen, stored at -80°C and used for the estimation of lipid peroxidation, GSH, MPO and SOD levels.
Histological analysis and neutrophil counting Mouse kidneys are ?xed in 4% dehyde and ed in paraf?n wax. The 5- mm-thick sections are deparaf?nised in xylene and rehydrated h graded concentrations of ethanol. H&E and PAS staining are performed using standard protocols. Images are collected and ed using a light cope (1X71, Olympus, Tokyo, Japan) with DP analyser software BSW, Tokyo, Japan). Tubular damage in PAS-stained kidney sections is examined under a light microscope and scored based on the percentage of cortical tubular necrosis: 0 = normal, 1 = 1—10, 2 = 11—25, 3 = 26—45, 4 = 46—75, and 5 = 76—100%.
Slides are scored in a blinded manner, and results are means i s.d. of 10 representative ?elds/group. Severity criterion for tubular necrosis displaying the loss of the proximal tubular brush border and cast formation are used to fy s. The sample size for each group is 10. Neutrophil in?ltration is quantitatively assessed on PAS stained tissue by a renal pathologist by counting the number of neutrophils per high-power ?eld (x400). At least ?elds are counted in the outer stripe of the outer medulla for each slide.
All values are represented as mean :t s.d. One—way analysis of variance is used to ate the statistical cance of the results of all assays and P—values < 0.05 are considered statistically signi?cant.
Example 47: Study of the Effects of Compounds of Formula 11), Formula 11a), Formula 11b), Formula 111), or Formula 1111) on Ischemia/Reperfusion-induced Acute Kidney Inju? A study is performed to determine the effects of compounds of Formula (I), Formula (Ia), Formula (1b), Formula (II), or Formula (111), or a pharmaceutically acceptable salt thereof, on Ischemia/Reperfusion-induced (UR—induced) Acute Kidney Injury in CD-l (ICR) mice.
CD-l (ICR) mice are purchased from Charles River Laboratory (Wilmington, MA).
Mice are housed in a temperature- and humidity-controlled environment with a 12: 12 hrs light—dark cycle and are allowed freely access to rd rodent chow (TekLad, Madison, WI) and tap water.
Mice are subjected to a midline back incision, and both renal pedicles are clamped for 45 min with microaneurysm clamps (00396-01; Fine Science Tools, Foster City, CA). After [Annotation] KEB removal of the clamp, the kidneys are ted for the restoration of blood flow. The animals are allowed to r, and they are sacri?ced 48 hrs after reperfusion. Mice are treated with 100 mg/kg of a compound of Formula (1), Formula (Ia), Formula (1b), Formula (II), or Formula (III), or a pharmaceutically acceptable salt thereof, by oral gavage once per day. CD—1 mice are divided into four groups: (1) young mice with sham injury (n = 4) (6—7 weeks old); (2) young mice with UK injury (11 = 8); (3) adult mice with sham injury (n = 4) (20—24 weeks old); and (4) adult mice with I/R injury (n = 11). An additional 27 adult mice (20—24 weeks old) are ized into two groups: 13 mice ed a compound of Formula (1), Formula (Ia), Formula (Ib), Formula (II), or Formula (III), or a pharmaceutically acceptable salt thereof, and the other 14 mice received the vehicle as a control.
The serum creatinine level is measured using the QuantiChrom nine Assay Kit (DICT—500, BioAssay Systems, Hayward, CA). BUN measurements are recorded using the In?nity Urea (Nitrogen) Liquid Stable Reagent (TR12421; ThermoTrace, Victoria, AU).
Evaluation ofrenal tissue Kidneys are ?xed in 4% paraformaldehyde, ed in paraf?n, and stained with hematoxylin and eosin (4 mm thick). Tubular injury is scored on a scale of 0—4 on the basis of the percentage of tubules with is, tion, or cell swelling: 0, less than 5%; l, 5— %; 2, 25—50%; 3, 50—75%; and 4, over 75%. All high—power ?elds (x 400) in the cortex and outer medulla are evaluated by a pathologist in a blinded manner.
All values are expressed as mean :: s.e. Statistical analysis is carried out using GraphPad Prism 4.00 (San Diego, CA) with unpaired Student’s t testing for two sets of data and an analysis of variance with a Bonferroni post-test for multiple groups. P < 0.05 was ered signi?cant.
Example 48: Determination of the Effects of Compounds 1 and 17 on FoxOl Phosphorylation levels AML-12 cells were treated with different concentrations of Compound 1 or nd 17 for 24 hours. Cells were then lysed in lysis buffer (50 mM Tris, 150 mM KCl, EDTA lmM, NP40 1%) containing protease and phosphatase inhibitors, and analyzed by SDS—PAGE/Western blot. Blocking and antibody incubations were done in 5% milk. Each protein t was ed with its speci?c antibody. Tubulin antibody was obtained from Sigma Inc, FoxOl and phopho—FoxOl (Ser256) antibodies were obtained from Cell Signaling. Antibody detection reactions were ped by enhanced chemiluminescence (Advansta, CA, USA) using x-ray ?lms.
[Annotation] KEB [Annotation] KEB FoxOl orylation at Ser256 results in its nuclear export and in inhibition of its transcription factor activity. A decrease in FoxOl phosphorylation at Ser256 with increasing dose of Compound 1 and 17 was observed (, indicating an increase in nuclear- translocated FoxOl and therefore increased FoxOl transcriptional activity.
Unless otherwise , all technical and scienti?c terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this disclosure belongs. In the cation, the singular forms also include the plural unless the context clearly es otherwise. Although methods and materials similar or equivalent to those described herein can be used in the practice of testing the present disclosure, suitable methods and materials are described below. All publications, patent applications, patents, and other references mentioned herein are hereby expressly incorporated by reference. The references cited herein are not admitted to be prior art of the claimed disclosure. In the case of con?ict, the present speci?cation, including de?nitions, will control. In on, the materials, methods, and examples are illustrative only and are not intended to be limiting.
Exam 1e 49: S nthesis of Exem li?ed Com ounds 1) diethyl (cyanomethyl) phosphonate, /§o MeOH. HCI Me020\©/§o K2003, THF, H20. 60 c. 1 h; -------> --------------------------------> Reflux 2)H2, 10% Pd/C, EtOH, THF, rt, 2 h MeOzCO/VCN 2a 2b 2c HCI (g) satd in EtOH/CHZCI2 Ho. 1:1, 00, 20 h; Meozc ......................> NH2 ---------------------> 2N NH3.OC, 18 h; "0' NH2 2M NaOH See, Hirose M, et al., "Design and synthesis of novel DFG-out scular endothelial growth factor receptor 2 (VEGFRZ) inhibitors: 3. tion of 5—amino-linked thiazolo[5,4-d]pyrimidine and thiazolo[5,4-b]pyridine tives." Bioorg. Med. Chem. 2012, 15;20(18):5600—15.
[Annotation] KEB H \/© CHZNZV MeOH 002MB HOZCUNHz Benzoylchloroformate, H02C N 0 ‘rr Wolff rearr. N O ........... - ............. - 1r Nchogl H20, Dioxane 78% 0 7b 7c 7d 602Me 2M, NaOH, See, Clift MD, Silverman RB., "Synthesis and evaluation of novel aromatic substrates and competitive inhibitors of GABA aminotransferase," Bioorg. Med. Chem. Lett,. 2008, ;18(10):3122-5.
H2N 002E! N N 002H 0 H N base, DMF | Jk 7e . ---------------r- M X 2M,NaOH, Nc MeOH NH COH | H2" 2 NAN COZH HCI H Commercial X = Cl, Br, SMe, SOZMe A. M. El—Reedy, A. O. Ayyad and A. S. Ali, "Azolopyrimidines and pyrimidoquinazolines from ropyrimidines," J. Het. Chem. 1989, 26, 313-16.
COZMe DMF,COZC|2‘ . DME, 40°C 0 NH 0 Cl COzMe N’J‘N cozH NH base, DMF | A 11d 1 + ..............T- N ""2 O 2M,NaOH.
MeOH NH O 002"" c. NJ\N COZH Commercial See, Iwahashi M, et a1., "Design and synthesis of new prostaglandin D2 receptor antagonists," Bioorg. Med. Chem. 2011, 19(18):5361—71.
Br CO 2Et CaCO 3 , H 2O Dioxane, 6h, 80°C O HO CO 2Et N O CO 2H N base, DMF 13c 13 + O H 2M, NaOH, NC MeOH NH CO 2Me HO CO 2H 7f N O X = Cl, Br, SO 2Me, SMe 13d 13a NH 3-THF, THF, rt, 34h, rt CO 2Me See, U.S. 2008/004,302(A1); and U.S. 8,716,470 (B2).
EQUIVALENTS Those skilled in the art will recognize, or be able to ascertain using no more than routine experimentation, many equivalents to the specific embodiments and methods described herein. Such equivalents are intended to be encompassed by the scope of the t disclosure.
Definitions of specific embodiments of the invention as d herein follow.
According to a first ment of the invention, there is provided compound represented by Formula (I): or a pharmaceutically acceptable salt or tautomer thereof, wherein: X is O, OH, or Cl; L is mY(CH2)p-; Y is O, NH or S(O)q; R1 is C6-C10 aryl or heteroaryl, wherein the aryl and heteroaryl are substituted with Ra and Rb, and optionally substituted with one or more Re; R2 is H or C1-C6 alkyl; one of Ra and Rb is hydrogen and the other is -(CH2)rCO2Rx, -OCH2CO2Rx, -(CH2)rtetrazole, -(CH2)roxadiazolone, -(CH2)rtetrazolone, rthiadiazolol, -(CH2)r isoxazolol, -(CH2)rP(O)(OH)ORx, -(CH2)rS(O)2OH, -(CH2)rC(O)NHCN, or rC(O)NHS(O)2alkyl; Rc is C1-C6 haloalkyl, halogen, –CN, –ORx, -CO2Rx, or NO2; Rd is methyl, 5- to 10-membered aryl, 5- or 6-membered heteroaryl, or 5- or 6- ed carbocycle; each Rx is independently at each occurrence hydrogen or C1-C6 alkyl; each Re is independently C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, halogen, -ORy, C1-C6 haloalkyl, -NHRz, -OH, or -CN; Rf is H or absent; each Ry and Rz is independently hydrogen, C1-C6 alkyl, or C1-C6 haloalkyl; m is 0, 1 or 2; p is 1 or 2, wherein m + p < 3; q is 0, 1, or 2; r is 0 or 1; and the dotted line is an optional double bond; with the proviso that Rc is not –CN when X is O, L is –SCH2- and Rd is phenyl, that Rc is not C1-C6 alkyl when X is O, L is –SCH2- and Rd is methyl, and that Rc is not –CN when X is O, L is –SCH2- and Rd is 2-furyl.
According to a second embodiment of the invention, there is provided a pharmaceutical composition comprising the compound of the first embodiment, or a pharmaceutically able salt thereof, and at least one of a pharmaceutically acceptable carrier, diluent, or excipient.
According to a third ment of the invention, there is provided the use of the compound of the first embodiment or a pharmaceutically acceptable salt thereof in the manufacture of a medicament for treating, preventing, or reducing the risk of a disease or disorder ated with reduced nicotinamide adenine dinucleotide (NAD+) levels, by tion of o-?-carboxymuconate-?-semialdehyde decarboxylase (ACMSD).
According to a fourth embodiment of the invention, there is provided the use of the compound of the first embodiment or a pharmaceutically acceptable salt thereof in the manufacture of a medicament for ng, preventing, or reducing the risk of a disease or disorder associated with reduced namide adenine dinucleotide (NAD+) levels.
According to a fifth embodiment of the invention, there is provided the use of the compound of the first embodiment or a pharmaceutically acceptable salt thereof that increases intracellular nicotinamide adenine dinucleotide (NAD+) in the manufacture of a medicament for treating, preventing, or reducing the risk of a disorder associated with ondrial dysfunction.
According to a sixth embodiment of the invention, there is provided the use of the compound of the first embodiment or a pharmaceutically acceptable salt thereof that increases intracellular namide e dinucleotide (NAD+) in the manufacture of a medicament for promoting oxidative metabolism.
According to a seventh embodiment of the invention, there is provided the use of the compound of the first embodiment or a pharmaceutically acceptable salt thereof in the manufacture of a medicament for treating a disease or condition mediated by ACMSD.
According to an eighth embodiment of the invention, there is ed the use of the compound of the first embodiment or a pharmaceutically acceptable salt f in the cture of a medicament for treating, preventing or reducing the risk of a disease or disorder associated with ?-amino-?-carboxymuconate-?-semialdehyde decarboxylase (ACMSD) dysfunction.
According to a ninth embodiment of the invention, there is provided the use of the compound of the first embodiment or a pharmaceutically acceptable salt thereof in the manufacture of a medicament for treating, preventing or reducing the risk of a er ated with mitochondrial dysfunction. ing to a tenth embodiment of the invention, there is provided the use of the compound of the first embodiment or a pharmaceutically acceptable salt thereof in the manufacture of a medicament for promoting oxidative metabolism.
According to an eleventh ment of the invention, there is provided the use of a compound having one of the following Formula: O O NC NC NH NH CO 2H O CO 2H N S N S , , O O NC NC NH NH CO 2H S CO 2H N S N S S , , O O N NC NH NH N CO 2H CO 2H N S N S S , , NC NH CO 2H N S N , Cl CO 2H, O O NC NC NH NH CO 2Et CO 2H N S N S , , O O Br Br NH NH CO 2H S CO 2H N S N S , , NH H N S CO2H , , O O H NC NH NH S CO2H N S N S , CO2H, O O N N NH N N NH N S N N S CO2H S , S , N O NH N O N S N H N S CO 2H S , , Cl O N NH OH CO 2H S N N S N S , or O , or a pharmaceutically able salt thereof in the manufacture of a medicament for treating a disease or disorder associated with ?- amino-?-carboxymuconate-?-semialdehyde decarboxylase (ACMSD) dysfunction.

Claims (35)

1. A compound represented by a (I): or a pharmaceutically acceptable salt or tautomer thereof, wherein: X is O, OH, or Cl; L is –(CH2)mY(CH2)p-; Y is O, NH or S(O)q; R1 is C6-C10 aryl or heteroaryl, wherein the aryl and heteroaryl are substituted with Ra and Rb, and optionally substituted with one or more Re; R2 is H or C1-C6 alkyl; one of Ra and Rb is hydrogen and the other is rCO2Rx, -OCH2CO2Rx, -(CH2)rtetrazole, -(CH2)roxadiazolone, -(CH2)rtetrazolone, -(CH2)rthiadiazolol, -(CH2)r isoxazolol, -(CH2)rP(O)(OH)ORx, -(CH2)rS(O)2OH, -(CH2)rC(O)NHCN, or -(CH2)rC(O)NHS(O)2alkyl; Rc is C1-C6 haloalkyl, halogen, –CN, –ORx, -CO2Rx, or NO2; Rd is methyl, 5- to 10-membered aryl, 5- or ered heteroaryl, or 5- or 6- membered carbocycle; each Rx is independently at each occurrence hydrogen or C1-C6 alkyl; each Re is independently C1-C6 alkyl, C2-C6 alkenyl, C2-C6 l, halogen, -ORy, C1-C6 haloalkyl, -NHRz, -OH, or -CN; Rf is H or absent; each Ry and Rz is independently hydrogen, C1-C6 alkyl, or C1-C6 haloalkyl; m is 0, 1 or 2; p is 1 or 2, wherein m + p < 3; q is 0, 1, or 2; r is 0 or 1; and the dotted line is an optional double bond; with the o that Rc is not –CN when X is O, L is –SCH2- and Rd is phenyl, that Rc is not C1-C6 alkyl when X is O, L is –SCH2- and Rd is methyl, and that Rc is not –CN when X is O, L is –SCH2- and Rd is 2-furyl.
2. The compound of claim 1, wherein the compound is represented by Formula (Ia) or a pharmaceutically acceptable salt, or tautomer thereof.
3. The compound of claim 1 or 2, wherein the compound is represented by Formula (Ib): or a ceutically acceptable salt thereof, wherein n is 0, 1, 2, or 3;
4. The compound of any one of claims 1-3, wherein: one of Ra and Rb is hydrogen and the other is CO2Rx, Rx, tetrazole, or oxadiazolone; Rc is halogen, –CN, or –ORx; Rd is methyl, 5- to 10-membered aryl, 5- or 6-membered heteroaryl, or 5- or 6- membered carbocycle; and Rx is hydrogen or C1-C6 alkyl; each Re is ndently C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, halogen, -ORy, C1-C6 haloalkyl, -NHRz, -OH, or -CN; each Ry and Rz is independently hydrogen, C1-C6 alkyl, or C1-C6 haloalkyl; and n is 0, 1, 2, or 3; with the proviso that Rc is not –CN when Rd is phenyl and that Rc is not –CN when Rd is 2-furyl.
5. The compound of any one of claims 1-3, wherein the compound is represented by Formula (II): or a pharmaceutically acceptable salt thereof.
6. The compound of claim 5, wherein Rc is halogen, –CN, or –ORx, Rd is methyl, 5- to 10- membered aryl, 5- or 6-membered heteroaryl, or 5- or 6-membered carbocycle, and Rx is hydrogen or C1-C6 alkyl.
7. The compound of any one of claims 1-6, wherein Rc is –CN or halogen.
8. The compound of any one of claims 1-7, wherein Rd is methyl, cyclohexyl, nyl, thiazolyl, phenyl, or thienyl.
9. The compound of any one of claims 1-7, wherein Rd is methyl, cyclohexyl, pyridinyl, thiazolyl, thienyl, or .
10. The compound of any one of claims 1-4, wherein Ra is en, H, tetrazole, or oxadiazolone (1,2,4-oxadiazol-5(4H)-one).
11. The compound of any one of claims 1-4, wherein Rb is hydrogen, CH2CO2H, tetrazole, or zolone (1,2,4-oxadiazol-5(4H)-one).
12. The compound of any one of claims 1-4, wherein n is 0.
13. The nd of claim 1, wherein the compound is represented by Formula (III) (III), or a pharmaceutically acceptable salt thereof.
14. The compound of claim 13, wherein Ra and Rb is en and the other is -(CH2)rCO2Rx, -OCH2CO2Rx, -(CH2)rtetrazole, -(CH2)roxadiazolone, -(CH2)rtetrazolone, -(CH2)rthiadiazolol, -(CH2)r isoxazolol, -(CH2)rP(O)(OH)ORx, -(CH2)rS(O)2OH, -(CH2)rC(O)NHCN, or -(CH2)rC(O)NHS(O)2alkyl; Rc is C1-C6 haloalkyl, halogen, –CN, –ORx, -CO2Rx, or NO2; Rd is methyl, 5- to 10-membered aryl, 5- or 6-membered heteroaryl, or 5- or 6- membered carbocycle; each Rx is independently at each occurrence en or C1-C6 alkyl; each Re is independently C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, halogen, -ORy, C1-C6 haloalkyl, -NHRz, -OH, or -CN; each Ry and Rz is independently hydrogen, C1-C6 alkyl, or C1-C6 haloalkyl; n is 0, 1, 2, or 3.
15. The compound of claim 1, having any one of the following Formula: NH NH CO 2H S CO 2H N S N S S , , N NC NH NH N CO 2H CO 2H N S N S S , , O O NC NC NH NH CO 2H CO 2H N S N S N , , O O Br Br NH NH CO 2H S CO 2H N S N S , , O N NH NH N N CO 2H N S N S N , S , O O N N NH NH N O N S CO 2H N S N S , S , or N S or a pharmaceutically able salt thereof.
16. A pharmaceutical ition comprising the compound of any one of claims 1 to 15, or a pharmaceutically acceptable salt thereof, and at least one of a pharmaceutically acceptable carrier, diluent, or ent.
17. The pharmaceutical composition according to claim 16, which comprises one or more further therapeutic agents.
18. Use of the nd of any one of claims 1 to 15 or a pharmaceutically acceptable salt thereof in the manufacture of a medicament for treating, ting, or reducing the risk of a disease or disorder associated with reduced nicotinamide adenine dinucleotide (NAD+) levels, by inhibition of ?-amino-?-carboxymuconate-?-semialdehyde decarboxylase (ACMSD).
19. Use of the compound of any one of claims 1 to 15 or a pharmaceutically acceptable salt thereof in the manufacture of a medicament for ng, preventing, or reducing the risk of a disease or disorder associated with reduced nicotinamide adenine dinucleotide (NAD+) levels.
20. Use of the compound of any one of claims 1 to 15 or a ceutically acceptable salt thereof that increases intracellular nicotinamide adenine dinucleotide (NAD+) in the manufacture of a medicament for treating, preventing, or reducing the risk of a disorder associated with mitochondrial dysfunction.
21. The use of claim 20, wherein said er associated with mitochondrial dysfunction is an inherited ondrial disease, a common metabolic disorder, a neurodegenerative disease, an aging related disorder, a kidney disorder, or a chronic inflammatory disease.
22. The use of claim 21, wherein the common metabolic disorder is obesity or type II diabetes.
23. Use of the nd of any one of claims 1 to 15 or a pharmaceutically acceptable salt thereof that increases intracellular nicotinamide adenine dinucleotide (NAD+) in the manufacture of a medicament for promoting oxidative metabolism.
24. Use of the nd of any one of claims 1 to 15 or a pharmaceutically acceptable salt thereof in the manufacture of a medicament for ng a disease or ion ed by ACMSD.
25. Use of the compound of any one of claims 1-15 or a pharmaceutically acceptable salt thereof in the manufacture of a medicament for treating, preventing or reducing the risk of a disease or disorder associated with ?-amino-?-carboxymuconate-?-semialdehyde decarboxylase (ACMSD) dysfunction.
26. Use of the compound of any one of claims 1-15 or a pharmaceutically acceptable salt thereof in the manufacture of a medicament for treating, preventing or reducing the risk of a disorder associated with mitochondrial dysfunction.
27. Use of the compound of any one of claims 1-15 or a pharmaceutically acceptable salt thereof in the manufacture of a medicament for promoting oxidative metabolism.
28. Use of a compound having one of the following Formula: O O NC NC NH NH CO 2H O CO 2H N S N S , , O O NC NC NH NH CO2H S CO2H N S N S S , , O O N NC NH NH N CO2H CO2H N S N S S , , NC NH CO 2H N S N , Cl CO 2H, O O NC NC NH NH CO 2Et CO 2H N S N S , , O O Br Br NH NH CO 2H S CO 2H N S N S , , NH H CO 2H N S CO 2H , , O O H NC NH NH S CO2H N S N S , CO2H, O O N N NH N N NH N S N N S CO2H S , S , N O NH N O N S N H N S CO 2H S , , Cl O N NH OH CO 2H S N N S N S , or O , or a pharmaceutically acceptable salt thereof in the manufacture of a medicament for treating a e or disorder associated with ?-amino-?- carboxymuconate-?-semialdehyde decarboxylase (ACMSD) dysfunction.
29. The use of claim 18, wherein said disease or disorder is an inherited mitochondrial e, a common metabolic disorder, a neurodegenerative disease, an aging related disorder, a kidney disorder, a chronic inflammatory disease, non-alcoholic fatty liver disease (NAFLD), or non-alcoholic steatohepatitis (NASH).
30. The use of claim 19, wherein said disease or er is an inherited mitochondrial disease, a common metabolic disorder, a neurodegenerative disease, an aging related disorder, a kidney disorder, a c inflammatory disease, coholic fatty liver disease (NAFLD), or non-alcoholic steatohepatitis .
31. The use of claim 20, wherein said disorder is an inherited mitochondrial disease, a common metabolic disorder, a neurodegenerative disease, an aging related disorder, a kidney disorder, a chronic inflammatory disease, non-alcoholic fatty liver disease (NAFLD), or nonalcoholic steatohepatitis .
32. The use of claim 23, wherein promoting oxidative metabolism is associated with an inherited mitochondrial disease, a common lic disorder, a egenerative disease, an aging related disorder, a kidney disorder, a chronic inflammatory e, non-alcoholic fatty liver disease (NAFLD), or non-alcoholic steatohepatitis (NASH).
33. The use of claim 25, wherein said disease or disorder is an inherited ondrial disease, a common metabolic disorder, a neurodegenerative e, an aging related disorder, a kidney disorder, a c matory disease, coholic fatty liver disease (NAFLD), or non-alcoholic hepatitis (NASH).
34. The use of claim 26, wherein said disorder is an inherited mitochondrial disease, a common metabolic disorder, a neurodegenerative disease, an aging related disorder, a kidney disorder, a chronic inflammatory disease, non-alcoholic fatty liver disease (NAFLD), or nonalcoholic steatohepatitis (NASH).
35. The use of claim 27, wherein promoting oxidative metabolism is associated with an inherited mitochondrial disease, a common metabolic disorder, a neurodegenerative disease, an aging related disorder, a kidney disorder, a chronic inflammatory disease, non-alcoholic fatty liver disease (NAFLD), or non-alcoholic steatohepatitis (NASH). I Compound 4 (0.5 uM) content l nd 4 (5 HM) NT Compound 4 Compound 4 0.5 uM 5 uM No |:l DMSO increase/Protein 5 nM _
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