WO2023177351A1 - Inhibiteurs de kinase induisant nf қb et méthodes associées - Google Patents

Inhibiteurs de kinase induisant nf қb et méthodes associées Download PDF

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WO2023177351A1
WO2023177351A1 PCT/SG2023/050147 SG2023050147W WO2023177351A1 WO 2023177351 A1 WO2023177351 A1 WO 2023177351A1 SG 2023050147 W SG2023050147 W SG 2023050147W WO 2023177351 A1 WO2023177351 A1 WO 2023177351A1
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optionally substituted
compound
formula
independently
prodrug
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PCT/SG2023/050147
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Venkataramanan RAMADASS
Sook Yee Lee
Vinay TERGAONKAR
Manikandan Lakshmanan
Thamil Selvan VAIYAPURI
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Agency For Science, Technology And Research
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    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P1/00—Drugs for disorders of the alimentary tract or the digestive system
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/33—Heterocyclic compounds
    • A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
    • A61K31/495—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
    • A61K31/505—Pyrimidines; Hydrogenated pyrimidines, e.g. trimethoprim
    • A61K31/519—Pyrimidines; Hydrogenated pyrimidines, e.g. trimethoprim ortho- or peri-condensed with heterocyclic rings
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D487/00—Heterocyclic compounds containing nitrogen atoms as the only ring hetero atoms in the condensed system, not provided for by groups C07D451/00 - C07D477/00
    • C07D487/02—Heterocyclic compounds containing nitrogen atoms as the only ring hetero atoms in the condensed system, not provided for by groups C07D451/00 - C07D477/00 in which the condensed system contains two hetero rings
    • C07D487/04—Ortho-condensed systems

Definitions

  • the present invention relates, in general terms, to NIK inhibitors and their use in treating fibrosis thereof.
  • NF-KB Nuclear factor-KB
  • Dysregulation of NFkB is linked to several human ailments especially those with inflammation as an underlying cause.
  • NFKB activation is governed by two important pathways. The more dominant and pervasive is termed the canonical pathway, which is activated in most cell types by a plethora of stimuli and this pathway involves the master kinases IKKa & IKKg.
  • IKKa & IKKg A more selective & milder activation of NFkB via organogenic cytokines and signals such as Lymphotoxin beta & BAFF is mediated by the kinase NIK.
  • NIK driven non-canonical NFkB signaling is also a key driver of mutant hTERT (telomerase reverse transcriptase) promoter reactivation. Mutations in hTERT promoters are now recognized to the single largest genetic alterations leading to diseases. Reactivation of TERT is the rate-limiting step in 90% of some human diseases irrespective of the other driver mutations.
  • NF-xB-inducing kinase also known as NIK or MAP3K14
  • NIK NF-xB-inducing kinase
  • TNF family receptors such as BAFFR, CD40, RANKL, and TWEAK
  • the receptor-associated TRAF3 degrades and consequently releases NIK from the TRAF3-TRAF2-CIAP1/2 complex.
  • the released NIK is stabilized and activated, which in turn phosphorylates the Serl76 residue of IKKa.
  • Activation of NIK and IKKa results in phosphorylation of the Ser866 and Ser870 residues of plOO, subsequent proteolytic cleavage of plOO, and production of p52.
  • NIK neuropeptide
  • NIK inhibitors There are currently no NIK inhibitors in the market. Inhibiting NIK function in disease is therefore a huge unmet need especially in fibrosis. Accordingly, there is a need to develop compounds for use as NIK inhibitors. There is also a need to identify alternative beneficial therapies which relies on the modulation of NFK;B and to develop suitable compounds.
  • the present invention provides a method of treating fibrosis, comprising a step of administering a compound of Formula (la) or a pharmaceutically acceptable salt, solvate, stereoisomer or prodrug thereof to a subject in need thereof, wherein each Xi, X2, X4 and X5 is independently C or N;
  • Ri and R2 are independently H, or optionally substituted alkyl
  • R3 is H, halo, or cyano
  • R4 is optionally substituted alkyl
  • R is optionally substituted heteroaryl
  • R4 and Rs is linked to give optionally substituted cycloalkyl, or optionally substituted heterocycyl.
  • the present invention provides a use of a compound of Formula (la) or a pharmaceutically acceptable salt, solvate, stereoisomer or prodrug thereof in the manufacture of a medicament for treating fibrosis in a patient in need thereof.
  • the present invention provides a compound of Formula (la) or a pharmaceutically acceptable salt, solvate, stereoisomer or prodrug thereof for use in treating fibrosis in a patient in need thereof.
  • the fibrosis is selected from pulmonary fibrosis, liver fibrosis, renal fibrosis, and long COVID induced pulmonary fibrosis.
  • Rs is optionally substituted 5 membered or 6 membered heteroaryl having at least one heteroatom.
  • 4 and Rs is linked to give optionally substituted cyclobutyl, optionally substituted cyclopentyl, or optionally substituted pyrrolidinonyl.
  • At least one Xi or X2 is N.
  • Xi is C and X2 is N.
  • R4 and Rs are different moieties
  • -C(OH)R4Rs is in the R- configuration.
  • the present invention also provides NIK inhibitors that is suitable for blocking cancers with mutations in promoters of genes like TERT, which are direct targets of NIK.
  • Such inhibitors can target inflammatory diseases such as chronic inflammation, which is a direct underlying cause for most human disorders.
  • the present invention provides a compound of Formula (Ila) or a salt, solvate, stereoisomer or prodrug thereof: wherein
  • X 4 is C and X5 is N, or X4 is N and X5 is C, or X4 and X5 are C;
  • Ri and R2 are independently H, or optionally substituted alkyl
  • R3 is H, halo, or cyano
  • R4 is optionally substituted alkyl
  • Rs is optionally substituted isoxazolyl, optionally substituted pyridinyl, 5- methylthiazolyl; or
  • R4 and Rs is linked to give optionally substituted cycloalkyl, or optionally substituted heterocycyl.
  • the compounds are suitable for treating systemic lupus erythematosus, inflammatory diseases such as IBD & psoriasis, immune inflammatory diseases, fibrosis, multiple myeloma and pancreatic cancer.
  • R4 and Rs are different moieties
  • -C(OH)R4Rs is in the R- configuration.
  • Rs is optionally substituted isoxazolyl.
  • R4 and Rs is linked to give optionally substituted cyclobutyl, optionally substituted cyclopentyl, or optionally substituted pyrrolidonyl.
  • the compound of Formula (Ila) is selected from:
  • the present invention also provides a pharmaceutical composition
  • a pharmaceutical composition comprising an effective amount of a compound of Formula (la) or a pharmaceutically acceptable salt, solvate, stereoisomer or prodrug thereof.
  • the pharmaceutical composition is administrable in combination with another active ingredient.
  • Figure 1 shows expression of fibrotic markers WI-38 human lung fibroblast cells incubated for 24 h with TGF-pi and pretreatment with Compound 12, Compound 21 and Nintedanib.
  • Figure 2 shows the effect of NIK Inhibitors (Compound 12 and Compound 21) and Nintedanib on TGF-gl-induced fibrotic markers protein expression in WI-38 human lung fibroblasts.
  • Figure 3 shows dose response curve of NIK Inhibitors (Compound 12 and Compound 21), ALK5 inhibitor and Nintedanib on FMT assay in WI-38 human lung fibroblasts.
  • Figure 4 shows dose response curve images of NIK Inhibitors (Compound 12 and Compound 21), ALK5 inhibitor and Nintedanib on FMT assay in WI-38 human lung fibroblasts.
  • Figure 5 shows the effect of NIK Inhibitors (Compound 12 and Compound 21), ALK5 inhibitor and Nintedanib on FMT assay at 10 pM concentration in WI-38 human lung fibroblasts.
  • Figure 6 shows digital images of NIK Inhibitors (Compound 12 and Compound 21), ALK5 inhibitor and Nintedanib on FMT assay at 10 M concentration in WI-38 human lung fibroblasts.
  • Figure 7 shows graph of body weight data on Compound 12 and Compound 21 in MTD study respectively.
  • Figure 8 shows line graph showing repeated and high dose PK of Compound 12 and Compound 21 and the respective concentration of compound at different time points are tabulated in the table.
  • Figure 9 shows the effect of NIK Inhibitors (Compound 12 and Compound 21), and Nintedanib on Ashcroft score in BLM induced lung fibrosis model.
  • Figure 10 shows the effect of NIK Inhibitors (Compound 12 and Compound 21), and Nintedanib on lung inflammation in BLM induced lung fibrosis model.
  • Figure 11 shows representative histological images for various groups from H8 ⁇ .E, masson's trichrome and picrosirius red stain. Effect of NIK Inhibitors (Compound 12 and Compound 21), and Nintedanib on lung histology in BLM induced lung fibrosis model.
  • Figure 12 shows effect of NIK siRNA on TWEAK-induced NIK protein expression in U2OS cells.
  • Figure 13 shows effect of NIK siRNA on TGF-pi-induced fibrotic markers protein expression and NIK protein expression in WI-38 human lung fibroblasts.
  • Alkyl refers to monovalent alkyl groups which may be straight chained or branched and preferably have from 1 to 10 carbon atoms or more preferably 1 to 6 carbon atoms. Examples of such alkyl groups include methyl, ethyl, n-propyl, /so-propyl, n-butyl, /so- butyl, n-hexyl, and the like.
  • Halo or halogen refers to fluoro, chloro, bromo and iodo.
  • Cycloalkyl refers to cyclic alkyl groups having a single cyclic ring or multiple condensed rings, preferably incorporating 3 to 11 carbon atoms.
  • Such cycloalkyl groups include, by way of example, single ring structures such as cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cyclooctyl, and the like, or multiple ring structures such as adamantanyl, indanyl, 1,2,3,4-tetrahydronapthalenyl and the like.
  • Heterocyclyl refers to a monovalent saturated or unsaturated group having a single ring or multiple condensed rings, preferably from 1 to 8 carbon atoms and from 1 to 4 hetero atoms selected from nitrogen, sulfur, oxygen, selenium or phosphorous within the ring. The most preferred heteroatom is nitrogen. It will be understood that where, for instance, R2 or R' is an optionally substituted heterocyclyl which has one or more ring heteroatoms, the heterocyclyl group can be connected to the core molecule of the compounds of the present invention, through a C-C or C-heteroatom bond, in particular a C-N bond.
  • heterocyclyl groups include, but are not limited to, aziridine, oxirane, thiirane, azetidine, 2,3-dihydroazete, 1,3-diazetidine, oxetane, 2H-oxete, thietane, 2H- thiete, azetidine-2-one, pyrrolidine, pyrroline, pyrrolidinone, pyrrole, pyrazolidine, imidazolidine, pyrazoline, imidazoline, tetra hydrofuran, 1,3-dioxolane, tetrahydrothiophene, oxathiolane, sulfolane, 2,4-thiazolidinedione, succinimide, 2- oxazolidone, hydantoin, piperidine, piperazine, tetra hydropyran, 2H-pyran, 4H-pyran, 1,4-dioxane,
  • Aryl refers to an unsaturated aromatic carbocyclic group having a single ring (eg. phenyl) or multiple condensed rings (eg. naphthyl or anthryl), preferably having from 6 to 14 carbon atoms.
  • aryl groups include phenyl, naphthyl and the like.
  • Heteroaryl refers to a monovalent aromatic heterocyclic group which fulfils the Huckel criteria for aromaticity (ie. contains 4n + 2 n electrons) and preferably has from 2 to 10 carbon atoms and 1 to 4 heteroatoms selected from oxygen, nitrogen, selenium, and sulfur within the ring (and includes oxides of sulfur, selenium and nitrogen).
  • Such heteroaryl groups can have a single ring (eg. pyridyl, pyrrolyl or N- oxides thereof or furyl) or multiple condensed rings (eg. indolizinyl, benzoimidazolyl, coumarinyl, quinolinyl, isoquinolinyl or benzothienyl).
  • heteroaryl groups include, but are not limited to, oxazole, pyrrole, imidazole, pyrazole, pyridine, pyrazine, pyrimidine, pyridazine, indolizine, isoindole, indole, indazole, purine, isoquinoline, quinoline, phthalazine, naphthylpyridine, quinoxaline, quinazoline, cinnoline, pteridine, carbazole, carboline, phenanthridine, acridine, phenanthroline, isothiazole, phenazine, isoxazole, isothiazole, phenoxazine, phenothiazine, thiazole, thiadiazoles, oxadiazole, oxatriazole, tetrazole, thiophene, benzo[b]thiophene, imidazopyridine and the like.
  • a group may or may not be further substituted or fused (so as to form a condensed polycyclic group) with one or more groups selected from hydroxyl, acyl, alkyl, alkoxy, alkenyl, alkenyloxy, alkynyl, alkynyloxy, amino, aminoacyl, thio, arylalkyl, arylalkoxy, aryl, aryloxy, carboxyl, acylamino, cyano, halogen, nitro, phosphono, sulfo, phosphorylamino, phosphinyl, heteroaryl, heteroarylalkyl, heteroaryloxy, heterocyclyl, heterocyclylalkyl, heterocyclyloxy, oxyacyl, oxime, oxime ether, hydrazone, oxyacylamino, oxysulfonylamino, aminoacyloxy, trihalomethyl, trialkyl, alkoxy, alkenyl, alken
  • “Isomer” includes especially optical isomers (for example essentially pure enantiomers, essentially pure diastereomers, and mixtures thereof) as well as conformation isomers (i.e. isomers that differ only in their angles of at least one chemical bond), position isomers (particularly tautomers), and geometric isomers (e.g. cis-trans isomers).
  • optical isomers for example essentially pure enantiomers, essentially pure diastereomers, and mixtures thereof
  • conformation isomers i.e. isomers that differ only in their angles of at least one chemical bond
  • position isomers particularly tautomers
  • geometric isomers e.g. cis-trans isomers
  • Compounds described herein can comprise one or more asymmetric centers, and thus can exist in various isomeric forms, e.g., enantiomers and/or diastereomers.
  • the compounds described herein can be in the form of an individual enantiomer, diastereomer or geometric isomer, or can be in the form of a mixture of stereoisomers, including racemic mixtures and mixtures enriched in one or more stereoisomer.
  • Isomers can be isolated from mixtures by methods known to those skilled in the art, including chiral high pressure liquid chromatography (HPLC) and the formation and crystallization of chiral salts; or preferred isomers can be prepared by asymmetric syntheses.
  • HPLC high pressure liquid chromatography
  • the invention additionally encompasses compounds described herein as individual isomers substantially free of other isomers, and alternatively, as mixtures of various isomers.
  • "Optically-enriched,” as used herein, means that the compound is made up of a significantly greater proportion of one enantiomer.
  • the compound of the present invention is made up of at least about 90% by weight of a preferred enantiomer.
  • the compound is made up of at least about 95%, 98%, or 99% by weight of a preferred enantiomer.
  • Preferred enantiomers may be isolated from racemic mixtures by any method known to those skilled in the art, including chiral high pressure liquid chromatography (HPLC) and the formation and crystallization of chiral salts or prepared by asymmetric syntheses.
  • HPLC high pressure liquid chromatography
  • Jacques et al. Enantiomers, Racemates and Resolutions (Wiley Interscience, New York, 1981); Wilen et al., Tetrahedron 33:2725 (1977); Eliel, Stereochemistry of Carbon Compounds (McGraw-Hill, NY, 1962); and Wilen, Tables of Resolving Agents and Optical Resolutions p. 268 (E.L. Eliel, Ed., Univ, of Notre Dame Press, Notre Dame, IN 1972).
  • an "inhibitor” is a molecule that binds to a substrate and decreases its activity.
  • a substrate may be an enzyme, protein or small molecule. Blocking a substrate's activity can kill a pathogen or correct a metabolic imbalance.
  • the binding of an inhibitor can stop another molecule (biomolecule) from entering the substrate's active site and/or hinder the substrate from catalyzing its reaction.
  • Inhibitor binding is either reversible or irreversible. Irreversible inhibitors usually react with the substrate and change it chemically (e.g. via covalent bond formation). These inhibitors modify key amino acid residues needed for enzymatic activity.
  • reversible inhibitors bind non-covalently and different types of inhibition are produced depending on the binding and complexation. For example the inhibition may be competitive, uncompetitive, non-competitive or mixed.
  • inhibitor refers to an act of decreasing a substrate's activity as described above. This action may be performed by a molecule which may be an inhibitor.
  • NIK is a key player in non-canonical NF-kB pathway which in turn is implicated in fibrosis and immune- inflammatory disease condition. Inhibition of NIK can help resolve such conditions. In particular, inhibition of overexpression of NIK can help ameliorate fibrosis and immune- inflammatory disease condition.
  • Nintedanib is considered as a comparator for fibrosis treatment, Nintedanib competitively inhibits both non-receptor tyrosine kinases (nRTKs) and receptor tyrosine kinases (RTKs).
  • NRTK targets of Nintedanib include Lek, Lyn, and Src.
  • Nintedanib inhibits platelet-derived growth factor receptor (PDGFR), vascular endothelial growth factor receptor (VEGFR) and fibroblast growth factor receptor (FGFR).
  • PDGFR platelet-derived growth factor receptor
  • VEGFR vascular endothelial growth factor receptor
  • FGFR fibroblast growth factor receptor
  • Pirfenidone is another antifibrotic compound, which is also a non-selective growth factor inhibitors.
  • the present compounds are clean and more selectively inhibit NIK.
  • the compounds show better cellular activity (target engagement) as compared to a comparator compound.
  • the compounds are shown to be useful as NIK inhibitors suitable for treating fibrosis. This is supported and substantiated with various assays such as in-vitro cell-based assay (fibrotic gene expression, fibrotic protein expression), golden standard FMT assay, high content image analysis in TGF-pi induced collagen 1A. Preclinical trials was substantiated with the golden standard preclinical animal fibrosis model called BLM induced lung fibrosis model in C57 mice. In both in vitro and in vivo models, the compounds showed superior activity compared to clinical standard Nintedanib.
  • the present invention provides a compound of Formula (I) or a salt, solvate, stereoisomer or prodrug thereof: wherein each of Xi, X2, X3, X4 and X5 is independently C or N;
  • Ri and R2 are independently H, or optionally substituted alkyl
  • R3 is H, halo, or cyano
  • R4 is optionally substituted alkyl
  • R is optionally substituted heteroaryl
  • R4 and Rs is linked to give optionally substituted cycloalkyl, or optionally substituted heterocyclyl.
  • the compounds are useful for treating fibrosis, inflammatory diseases such as asthma and chronic obstructive pulmonary disease (COPD) and immune inflammatory diseases such as rheumatoid arthritis, psoriasis, systemic lupus erythematosus (SLE), crohn disease (CD), and ulcerative collitis (UC).
  • COPD chronic obstructive pulmonary disease
  • immune inflammatory diseases such as rheumatoid arthritis, psoriasis, systemic lupus erythematosus (SLE), crohn disease (CD), and ulcerative collitis (UC).
  • SLE systemic lupus erythematosus
  • CD crohn disease
  • ulcerative collitis ulcerative collitis
  • the compounds are also useful for treating cancer, proliferative diseases and/or solid tumors, such as multiple myeloma and pancreatic cancer.
  • Ri is H or optionally substituted alkyl. In other embodiments, Ri is H. In other embodiments, Ri is optionally substituted C1-C5 alkyl. In other embodiments, Ri is optionally substituted methyl, optionally substituted ethyl, optionally substituted n-propyl, optionally substituted iso-propyl, optionally substituted n-butyl, optionally substituted sec-butyl, optionally substituted iso-butyl, or optionally substituted tert-butyl.
  • R2 is H or optionally substituted alkyl. In other embodiments, R2 is H. In other embodiments, R2 is optionally substituted C1-C5 alkyl. In other embodiments, R2 is optionally substituted methyl, optionally substituted ethyl, optionally substituted n-propyl, optionally substituted iso-propyl, optionally substituted n-butyl, optionally substituted sec-butyl, optionally substituted iso-butyl, or optionally substituted tert-butyl.
  • R3 is halo, or cyano. In other embodiments, R3 is fluoro, chloro, bromo or cyano.
  • R4 is optionally substituted alkyl. In other embodiments, R4 is optionally substituted C1-C5 alkyl. In other embodiments, R4 is optionally substituted methyl, optionally substituted ethyl, optionally substituted n-propyl, optionally substituted iso-propyl, optionally substituted n-butyl, optionally substituted sec-butyl, optionally substituted iso-butyl, or optionally substituted tert-butyl.
  • Rs is optionally substituted heteroaryl. In other embodiments, Rs is optionally substituted 5 membered or 6 membered heteroaryl having at least one heteroatom. In other embodiments, Rs is optionally substituted 5 membered or 6 membered heteroaryl having at least one N or 0.
  • Rs is optionally substituted thiazolyl, optionally substituted pyridinyl, optionally substituted isoxazolyl, optionally substituted imidazolyl, optionally substituted pyrazolyl, optionally substituted triazolyl, optionally substituted tetrazolyl, optionally substituted oxazolyl, optionally substituted isothiazolyl, optionally substituted oxadiazolyl, optionally substituted thiadiazolyl, optionally substituted thiadiazolyl, optionally substituted pyridazinyl, optionally substituted pyrimidinyl, optionally substituted pyrazinyl, or optionally substituted triazinyl.
  • Rs is optionally substituted thiazolyl, pyridinyl, isoxazolyl, or imidazolyl.
  • R4 and Rs is linked to give optionally substituted cycloalkyl, or optionally substituted heterocyclyl.
  • R4 and Rs is linked to give optionally substituted fused cycloalkyl, optionally substituted bridged cycloalkyl, optionally substituted spiro cycloalkyl, optionally substituted fused heterocyclyl, optionally substituted bridged heterocyclyl or optionally substituted spiro heterocyclyl.
  • the optionally substituted cycloalkyl is an optionally substituted C3-C8 cycloalkyl.
  • the optionally substituted cycloalkyl is an optionally substituted cyclopropanyl, optionally substituted cyclobutyl, optionally substituted cyclopentyl, optionally substituted cyclohexyl, optionally substituted cycloheptyl, or optionally substituted cyclooctanyl.
  • the optionally substituted cycloalkyl is an optionally substituted cyclopropanyl, optionally substituted cyclobutyl, optionally substituted cyclopentyl, optionally substituted cycloheptyl, or optionally substituted cyclooctanyl.
  • the optionally substituted cycloalkyl is an optionally substituted cyclobutyl, or optionally substituted cyclopentyl.
  • the optionally substituted heterocyclyl is optionally substituted 5 membered or 6 membered heterocyclyl having at least one heteroatom.
  • the optionally substituted heterocyclyl is optionally substituted pyrrolidinonyl, optionally substituted pyrrolidinedionyl, optionally substituted thiazolidinedionyl, optionally substituted oxazolidonyl, optionally substituted pyrrolidinyl, optionally substituted pyrrolinyl, optionally substituted pyrrolyl, optionally substituted pyrazolidinyl, optionally substituted imidazolidinyl, optionally substituted pyrazolinyl, optionally substituted imidazolinyl, optionally substituted tetrahydrofuranyl, optionally substituted 1,3-dioxolanyl, optionally substituted tetrahydrothiophenyl, optionally substituted oxathiolanyl, optionally substituted sulfolanyl, optionally substituted 2,4-thiazolidinedionyl, optionally substituted hydantoinyl, optionally substituted
  • the optional substitutent is halo or alkyl. In some embodiments, the optional substitutent is halo or C1-C5 alkyl.
  • At least one Xi or X2 is N. In some embodiments, Xi is C and X2 is N.
  • each X3, X4 and X5 is independently C or N. In some embodiments, at least one of X3, X4 and X5 is N.
  • Stereoisomers are isomers that have the same chemical composition but differ in the orientation of those parts in space.
  • enantiomers are stereoisomers that are mirror images of each other.
  • the compounds of Formula (I) or sub-formulae thereof bears a chiral carbon, which is denoted by below; i.e. -C(OH)R4Rs when R4 and Rs are different moieties.
  • stereochemistry is in the R-configuration. In certain other embodiments the stereochemistry is in the S-configuration. Preferably, the stereochemistry is in the R-configuration.
  • the compound of Formula (I) is a compound of Formula (I') or a salt, solvate or prodrug thereof: wherein each of Xi, X2, X3, X4 and X5 is independently C or N;
  • Ri and R2 are independently H, or optionally substituted alkyl
  • R3 is H, halo, or cyano
  • R4 is optionally substituted alkyl
  • Rs is optionally substituted heteroaryl.
  • the compound of Formula (I) is a compound of Formula (I") or a salt, solvate or prodrug thereof:
  • each of Xi, X2, X3, X4 and X5 is independently C or N;
  • Ri and R2 are independently H, or optionally substituted alkyl
  • R3 is H, halo, or cyano
  • R4 and Rs is linked to give optionally substituted cycloalkyl, or optionally substituted heterocyclyl.
  • the compound of Formula (I) is a compound of Formula (la): wherein Xi, X2, X4, Xs, Ri, R2, R3, R4 and Rs are as disclosed herein.
  • the compound of Formula (I) is a compound of Formula (la') or a salt, solvate or prodrug thereof: da') wherein each of Xi, X2, X4 and Xs is independently C or N;
  • Ri and R2 are independently H, or optionally substituted alkyl
  • R3 is H, halo, or cyano
  • R4 is optionally substituted alkyl
  • Rs is optionally substituted heteroaryl.
  • the compound of Formula (I) is a compound of Formula (la") or a salt, solvate or prodrug thereof: da") wherein each of Xi, X2, X4 and X5 is independently C or N;
  • Ri and R2 are independently H, or optionally substituted alkyl
  • R3 is H, halo, or cyano
  • R4 and Rs is linked to give optionally substituted cycloalkyl, or optionally substituted heterocyclyl.
  • the compound of Formula (I) is a compound of Formula (II): wherein X3, X4, Xs, Ri, R2, R3, R4 and Rs are as disclosed herein.
  • X3, X4, Xs are independently C or N;
  • Ri and R2 are independently H, or optionally substituted alkyl
  • R3 is H, halo, or cyano
  • R4 is optionally substituted alkyl
  • Rs is optionally substituted isoxazolyl, optionally substituted pyridinyl, 5- methylthiazolyl; or
  • R4 and Rs is linked to give optionally substituted cycloalkyl, or optionally substituted heterocycyl.
  • X3, X4 are C and X5 is N, or X3, X4 are N and X5 is C, or X3, X4 and X5 is C.
  • the compounds of Formula (II) or sub-formulae thereof bears a chiral carbon, which is denoted by below; i.e. -C(OH)R4Rs when R4 and Rs are different moieties.
  • stereochemistry is in the R-configuration. In certain other embodiments the stereochemistry is in the S-configuration. Preferably, the stereochemistry is in the R-configuration.
  • the compound of Formula (I) is a compound of Formula (II'): wherein X3, X4, Xs, Ri, R2, R3 are as disclosed herein;
  • R4 is optionally substituted alkyl
  • Rs is optionally substituted isoxazolyl, optionally substituted pyridinyl, 5- methylthiazolyl.
  • the compound of Formula (I) is a compound of Formula (II"):
  • R4 and Rs is linked to give optionally substituted cycloalkyl, or optionally substituted heterocycyl.
  • the compound of Formula (I) is a compound of Formula (Ila): wherein
  • X4, X5 are independently C or N;
  • Ri and R2 are independently H, or optionally substituted alkyl
  • R3 is H, halo, or cyano
  • R4 is optionally substituted alkyl
  • Rs is optionally substituted isoxazolyl, optionally substituted pyridinyl, 5- methylthiazolyl; or
  • R4 and Rs is linked to give optionally substituted cycloalkyl, or optionally substituted heterocycyl.
  • X4 is C and X5 is N, or X4 is N and X5 is C, or X4 and X5 are C.
  • the compound of Formula (I) is a compound of Formula (Ila'):
  • X 4 , X5 are independently C or N;
  • Ri and R2 are independently H, or optionally substituted alkyl
  • R3 is H, halo, or cyano
  • R4 is optionally substituted alkyl
  • Rs is optionally substituted isoxazolyl, optionally substituted pyridinyl, 5- methylthiazolyl.
  • the compound of Formula (I) is a compound of Formula (Ila"): wherein
  • X 4 , X5 are independently C or N;
  • Ri and R2 are independently H, or optionally substituted alkyl
  • R3 is H, halo, or cyano
  • R4 and Rs is linked to give optionally substituted cycloalkyl, or optionally substituted heterocycyl.
  • the compound of Formula (I) is a compound of Formula (lib) or a salt, solvate, stereoisomer or prodrug thereof:
  • Ri and R2 are independently H, or optionally substituted alkyl
  • R3 is H, halo, or cyano
  • R4 is optionally substituted alkyl; Rs is optionally substituted isoxazolyl, optionally substituted pyridinyl; or R4 and Rs is linked to give optionally substituted pyrrolidonyl.
  • the compound of Formula (I) is a compound of Formula (lib) or a salt, solvate, stereoisomer or prodrug thereof: wherein Ri and R2 are independently H, or optionally substituted alkyl;
  • R3 is halo, or cyano
  • R4 is optionally substituted alkyl
  • R4 and Rs is linked to give optionally substituted pyrrolidonyl.
  • the compound of Formula (I) is a compound of Formula (lib') or a salt, solvate or prodrug thereof: wherein Ri, R2 and R3 are as disclosed herein;
  • R4 is optionally substituted alkyl
  • Rs is optionally substituted isoxazolyl, optionally substituted pyridinyl.
  • the compound of Formula (I) is a compound of Formula (lib") or a salt, solvate or prodrug thereof: wherein Ri, R2 and R3 are as disclosed herein; and
  • R4 and Rs is linked to give optionally substituted pyrrolidonyl.
  • the compound of Formula (I) is a compound of Formula (lie) or a salt, solvate, stereoisomer or prodrug thereof: wherein Ri and R2 are independently H, or optionally substituted alkyl;
  • R3 is H, halo, or cyano
  • R4 is optionally substituted alkyl
  • R is optionally substituted heteroaryl
  • R4 and Rs is linked to give optionally substituted cycloalkyl, or optionally substituted heterocycyl.
  • the compound of Formula (I) is a compound of Formula (lie) or a salt, solvate, stereoisomer or prodrug thereof: wherein Ri and R2 are independently H, or optionally substituted alkyl;
  • R3 is halo, or cyano
  • R4 is optionally substituted alkyl
  • R is optionally substituted heteroaryl
  • R4 and Rs is linked to give optionally substituted cycloalkyl, or optionally substituted heterocycyl.
  • the compound of Formula (I) is a compound of Formula (lie) or a salt, solvate, stereoisomer or prodrug thereof: wherein Ri and R2 are independently H, or optionally substituted alkyl;
  • R3 is H, halo, or cyano
  • R4 is optionally substituted alkyl
  • Rs is optionally substituted isoxazolyl, optionally substituted pyridinyl; or
  • R4 and Rs is linked to give optionally substituted cyclobutyl, optionally substituted cyclopentyl, optionally substituted cyclohexyl, or optionally substituted pyrrolidonyl.
  • the compound of Formula (I) is a compound of Formula (lie) or a salt, solvate, stereoisomer or prodrug thereof:
  • Ri and R2 are independently H, or optionally substituted alkyl
  • R3 is halo, or cyano
  • R4 is optionally substituted alkyl
  • Rs is optionally substituted isoxazolyl, optionally substituted pyridinyl; or
  • R4 and Rs is linked to give optionally substituted cyclobutyl, optionally substituted cyclopentyl, optionally substituted cyclohexyl, or optionally substituted pyrrolidonyl.
  • the compound of Formula (I) is a compound of Formula (lie') or a salt, solvate or prodrug thereof: wherein Ri, R2 and R3 are as disclosed herein;
  • R4 is optionally substituted alkyl
  • Rs is optionally substituted heteroaryl.
  • R4 is optionally substituted alkyl
  • Rs is optionally substituted isoxazolyl, optionally substituted pyridinyl.
  • the compound of Formula (I) is a compound of Formula (lie”) or a salt, solvate or prodrug thereof: die”) wherein Ri, R2 and R3 are as disclosed herein; and
  • R4 and Rs is linked to give optionally substituted cycloalkyl, or optionally substituted heterocycyl.
  • R4 and Rs is linked to give optionally substituted cyclobutyl, optionally substituted cyclopentyl, optionally substituted cyclohexyl, or optionally substituted pyrrolidonyl.
  • the compound of Formula (I) is a compound of Formula (lid) or a salt, solvate, stereoisomer or prodrug thereof: wherein Ri and R2 are independently H, or optionally substituted alkyl;
  • R3 is H, halo, or cyano
  • R4 is optionally substituted alkyl
  • R is optionally substituted heteroaryl
  • R4 and Rs is linked to give optionally substituted cycloalkyl, or optionally substituted heterocycyl.
  • the compound of Formula (I) is a compound of Formula (lid) or a salt, solvate, stereoisomer or prodrug thereof:
  • Ri and R2 are independently H, or optionally substituted alkyl
  • R3 is halo, or cyano
  • R4 is optionally substituted alkyl
  • R is optionally substituted heteroaryl
  • R4 and Rs is linked to give optionally substituted cycloalkyl, or optionally substituted heterocycyl.
  • the compound of Formula (I) is a compound of Formula (lid) or a salt, solvate, stereoisomer or prodrug thereof: wherein Ri and R2 are independently H, or optionally substituted alkyl;
  • R3 is halo, or cyano
  • R4 is optionally substituted alkyl
  • Rs is optionally substituted isoxazolyl, optionally substituted pyridinyl; or
  • R4 and Rs is linked to give optionally substituted cyclobutyl, optionally substituted cyclopentyl, optionally substituted cyclohexyl, or optionally substituted pyrrolidonyl.
  • the compound of Formula (I) is a compound of Formula (lid') or a salt, solvate or prodrug thereof:
  • R4 is optionally substituted alkyl; and Rs is optionally substituted heteroaryl.
  • R4 is optionally substituted alkyl
  • Rs is optionally substituted isoxazolyl, optionally substituted pyridinyl.
  • the compound of Formula (I) is a compound of Formula (lid") or a salt, solvate or prodrug thereof: did") wherein Ri, R2 and R3 are as disclosed herein; and
  • R4 and Rs is linked to give optionally substituted cycloalkyl, or optionally substituted heterocycyl.
  • R4 and Rs is linked to give optionally substituted cyclobutyl, optionally substituted cyclopentyl, optionally substituted cyclohexyl, or optionally substituted pyrrolidonyl.
  • the compound of Formula (I) is a compound of Formula (He):
  • the compound of Formula (I) is a compound of Formula (He): wherein Ri and R2 are independently H, or optionally substituted alkyl;
  • R3 is H, halo, or cyano
  • R4 is optionally substituted alkyl
  • R is optionally substituted heteroaryl
  • R4 and Rs is linked to give optionally substituted cycloalkyl, or optionally substituted heterocycyl.
  • R3 is a halo or cyano
  • the compounds are particularly selective for NIK.
  • the compound of Formula (I) is a compound of Formula (He) or a salt, solvate, stereoisomer or prodrug thereof: wherein Ri and R2 are independently H, or optionally substituted alkyl;
  • R3 is halo, or cyano
  • R4 is optionally substituted alkyl
  • Rs is optionally substituted isoxazolyl, optionally substituted pyridinyl, 5- methylthiazolyl; or
  • R4 and Rs is linked to give optionally substituted cycloalkyl, or optionally substituted heterocycyl.
  • the compound of Formula (I) is a compound of Formula (He) or a salt, solvate or prodrug thereof: wherein Ri and R2 are independently H, or optionally substituted alkyl;
  • R3 is halo, or cyano
  • R4 is optionally substituted alkyl
  • R4 and Rs is linked to give optionally substituted cycloalkyl, or optionally substituted heterocycyl.
  • the compound of Formula (I) is a compound of Formula (He) or a salt, solvate or prodrug thereof: wherein Ri and R2 are independently H, or optionally substituted alkyl;
  • R3 is halo, or cyano
  • R4 is optionally substituted alkyl
  • Rs is optionally substituted isoxazolyl, optionally substituted pyridinyl, 5- methylthiazolyl; or
  • FU and Rs is linked to give optionally substituted cyclobutyl, optionally substituted cyclopentyl, optionally substituted cyclohexyl, or optionally substituted pyrrolidonyl.
  • the compound of Formula (I) is a compound of Formula (He) or a salt, solvate or prodrug thereof: wherein Ri and R2 are independently H, or optionally substituted alkyl;
  • R3 is halo, or cyano
  • R4 is optionally substituted alkyl
  • R4 and Rs is linked to give optionally substituted cyclobutyl, optionally substituted cyclopentyl, or optionally substituted pyrrolidonyl.
  • the compound of Formula (I) is a compound of Formula (He) or a salt, solvate or prodrug thereof: wherein Ri and R2 are independently H, or optionally substituted alkyl;
  • R3 is halo, or cyano
  • R4 is optionally substituted alkyl
  • R4 and Rs is linked to give optionally substituted pyrrolidonyl.
  • the compound of Formula (I) is a compound of Formula (He'): wherein Ri, R2 and R3 are as disclosed herein;
  • R4 is optionally substituted alkyl
  • Rs is optionally substituted heteroaryl.
  • R4 is optionally substituted alkyl
  • Rs is optionally substituted isoxazolyl, optionally substituted pyridinyl, 5- methylthiazolyl.
  • R4 is optionally substituted alkyl and Rs is optionally substituted isoxazolyl.
  • the compound of Formula (I) is a compound of Formula (He"): wherein Ri, R2 and R3 are as disclosed herein; and
  • R4 and Rs is linked to give optionally substituted cycloalkyl, or optionally substituted heterocycyl.
  • R4 and Rs is linked to give optionally substituted cyclobutyl, optionally substituted cyclopentyl, optionally substituted cyclohexyl, or optionally substituted pyrrolidonyl. In some embodiments, R4 and Rs is linked to give optionally substituted pyrrolidonyl.
  • the compound of Formula (I), (II), or sub-formulae thereof is selected from:
  • the compound of Formula (I), (II), or sub-formulae thereof is selected from:
  • the compounds are characterised by a-SMA fiber inhibition EC50 of about 0.1 pM to about 10 pM.
  • the EC50 is about 0.1 pM to about 9 pM, about 0.1 pM to about 8 pM, about 0.1 pM to about 7 pM, about 0.1 pM to about 6 pM, about 0.1 pM to about 5 pM, about 0.1 pM to about 4 pM, about 0.1 pM to about 3 pM, about 0.1 pM to about 2 pM, or about 0.1 pM to about 1 pM.
  • the present invention also provides a pharmaceutical composition
  • a pharmaceutical composition comprising an effective amount of a compound of Formula (I), sub-formulae or a pharmaceutically acceptable salt, solvate, stereoisomer or prodrug thereof.
  • the present invention also provides a pharmaceutical composition
  • a pharmaceutical composition comprising an effective amount of a compound of Formula (II), sub-formulae or a pharmaceutically acceptable salt, solvate, stereoisomer or prodrug thereof.
  • compositions according to the invention may be prepared in a manner known in the art and are those suitable for enteral, such as oral or rectal, and parenteral administration to mammals (warm-blooded animals), particularly humans, comprising a therapeutically effective amount of at least one pharmacologically active combination partner alone, e.g., as indicated above, or in combination with one or more pharmaceutically acceptable carriers or diluents, especially suitable for enteral or parenteral application.
  • Suitable pharmaceutical compositions contain, e.g., from about 0.1% to about 99.9%, preferably from about 1 % to about 60%, of the active ingredient(s).
  • Suitable pharmaceutically acceptable salts include, but are not limited to salts of pharmaceutically acceptable inorganic acids such as hydrochloric, sulphuric, phosphoric, nitric, carbonic, boric, sulfamic, and hydrobromic acids, or salts of pharmaceutically acceptable organic acids such as acetic, propionic, butyric, tartaric, maleic, hydroxymaleic, fumaric, maleic, citric, lactic, mucic, gluconic, benzoic, succinic, oxalic, phenylacetic, methanesulphonic, toluenesulphonic, benezenesulphonic, salicyclic sulphanilic, aspartic, glutamic, edetic, stearic, palmitic, oleic, lauric, pantothenic, tannic, ascorbic and valeric acids.
  • pharmaceutically acceptable inorganic acids such as hydrochloric, sulphuric, phosphoric, nitric
  • Base salts include, but are not limited to, those formed with pharmaceutically acceptable cations, such as sodium, potassium, lithium, calcium, magnesium, ammonium and alkylammonium.
  • the present invention includes within its scope cationic salts eg sodium or potassium salts, or alkyl esters (eg methyl, ethyl) of the phosphate group.
  • Basic nitrogen-containing groups may be quarternised with such agents as lower alkyl halide, such as methyl, ethyl, propyl, and butyl chlorides, bromides and iodides; dialkyl sulfates like dimethyl and diethyl sulfate; and others.
  • lower alkyl halide such as methyl, ethyl, propyl, and butyl chlorides, bromides and iodides
  • dialkyl sulfates like dimethyl and diethyl sulfate; and others.
  • the compounds of Formula (I), (II), sub-formulae or a salt, solvate, stereoisomer or prodrug thereof may be in crystalline form either as the free compound or as a solvate (e.g. hydrate) and it is intended that both forms are within the scope of the present invention.
  • Methods of solvation are generally known within the art.
  • prodrug any compound that is a prodrug of the compound of Formula (I), (II), sub-formulae is also within the scope and spirit of the invention.
  • the compound of the invention can be administered to a subject in the form of a pharmaceutically acceptable pro-drug.
  • pro-drug is used in its broadest sense and encompasses those derivatives that are converted in vivo to the compound of the invention. Such derivatives would readily occur to those skilled in the art.
  • Other texts which generally describe prodrugs (and the preparation thereof) include: Design of Prodrugs, 1985, H. Bundgaard (Elsevier); The Practice of Medicinal Chemistry, 1996, Camille G.
  • amine moieties can be quarternised to improve its solubility.
  • a therapeutically effective amount is intended to include at least partially attaining the desired effect, or delaying the onset of, or inhibiting the progression of, or halting or reversing altogether the onset or progression of macular degeneration.
  • the term "effective amount" relates to an amount of compound which, when administered according to a desired dosing regimen, provides the desired therapeutic activity. Dosing may occur at intervals of minutes, hours, days, weeks, months or years or continuously over any one of these periods. Suitable dosages may lie within the range of about 0.1 ng per kg of body weight to 1 g per kg of body weight per dosage, such as is in the range of 1 mg to 1 g per kg of body weight per dosage. In one embodiment, the dosage may be in the range of 1 mg to 500 mg per kg of body weight per dosage. In another embodiment, the dosage may be in the range of 1 mg to 250 mg per kg of body weight per dosage. In yet another embodiment, the dosage may be in the range of 1 mg to 100 mg per kg of body weight per dosage, such as up to 50 mg per body weight per dosage.
  • Suitable dosage amounts and dosing regimens can be determined by the attending physician and may depend on the severity of the condition as well as the general age, health and weight of the patient to be treated.
  • the compound of the invention may be administered in a single dose or a series of doses. While it is possible for the active ingredient to be administered alone, it is preferable to present it as a composition, preferably as a pharmaceutical composition.
  • the formulation of such compositions is well known to those skilled in the art.
  • the composition may contain any suitable carriers, diluents or excipients. These include all conventional solvents, dispersion media, fillers, solid carriers, coatings, antifungal and antibacterial agents, dermal penetration agents, surfactants, isotonic and absorption agents and the like. It will be understood that the compositions of the invention may also include other supplementary physiologically active agents.
  • compositions may conveniently be presented in unit dosage form and may be prepared by any methods well known in the art of pharmacy. Such methods include the step of bringing into association the active ingredient with the carrier which constitutes one or more accessory ingredients. In general, the compositions are prepared by uniformly and intimately bringing into association the active ingredient with liquid carriers or finely divided solid carriers or both, and then if necessary shaping the product.
  • the compound may be injected directly to the eye, and in particular the vitreous of the eye.
  • the compound, composition or combination of the invention can be administered to the vitreous of the eye using any intravitreal or transscleral administration technique.
  • the compound, composition or combination can be administered to the vitreous of the eye by intravitreal injection.
  • Intravitreal injection typically involves administering a compound of the invention or a pharmaceutically acceptable salt, solvate or prodrug in a total amount between 0.1 ng to 10 mg per dose.
  • Injectables for such use can be prepared in conventional forms, either as a liquid solution or suspension or in a solid form suitable for preparation as a solution or suspension in a liquid prior to injection, or as an emulsion.
  • Carriers can include, for example, water, saline (e.g., normal saline (NS), phosphate-buffered saline (PBS), balanced saline solution (BSS)), sodium lactate Ringer's solution, dextrose, glycerol, ethanol, and the like; and if desired, minor amounts of auxiliary substances, such as wetting or emulsifying agents, buffers, and the like can be added.
  • saline e.g., normal saline (NS), phosphate-buffered saline (PBS), balanced saline solution (BSS)
  • NS normal saline
  • PBS phosphate-buffered saline
  • BSS balanced saline solution
  • Proper fluidity can be maintained, for example, by using a coating such as lecithin, by maintaining the required particle size in the case of dispersion and by using surfactants.
  • the compound, composition or combination can be dissolved in a pharmaceutically effective carrier and be injected into the vitreous of the eye with a fine gauge hollow bore needle (e.g., 30 gauge, 1/2 or 3/8 inch needle) using a temporal approach (e.g., about 3 to about 4 mm posterior to the limbus for human eye to avoid damaging the lens).
  • a person skilled in the art will appreciate that other means for injecting and/or administering the compound, composition or combinations to the vitreous of the eye can also be used.
  • These other means can include, for example, intravitreal medical delivery devices.
  • These devices and methods can include, for example, intravitreal medicine delivery devices, and biodegradable polymer delivery members that are inserted in the eye for long term delivery of medicaments.
  • These devices and methods can further include transscleral delivery devices.
  • solutions or suspensions of the compound, composition or combinations of the invention may be formulated as eye drops, or as a membranous ocular patch, which is applied directly to the surface of the eye.
  • Topical application typically involves administering the compound of the invention in an amount between 0.1 ng and 10 mg.
  • the compound, composition or combinations of the invention may also be suitable for intravenous administration.
  • a compound of Formula (I), (I'), (II), (IT) or a pharmaceutically acceptable salt, solvate or prodrug thereof may be administered intravenously at a dose of up to 16 mg/m 2 .
  • the compound, composition or combinations of the invention may also be suitable for oral administration and may be presented as discrete units such as capsules, sachets or tablets each containing a predetermined amount of the active ingredient; as a powder or granules; as a solution or a suspension in an aqueous or non-aqueous liquid; or as an oil-in-water liquid emulsion or a water-in-oil liquid emulsion.
  • the active ingredient may also be presented as a bolus, electuary or paste.
  • the compound of Formula (I), (II), sub-formulae or a pharmaceutically acceptable salt, solvate or prodrug is orally administerable.
  • a tablet may be made by compression or moulding, optionally with one or more accessory ingredients.
  • Compressed tablets may be prepared by compressing in a suitable machine the active ingredient in a free-flowing form such as a powder or granules, optionally mixed with a binder (e.g inert diluent, preservative disintegrant (e.g. sodium starch glycolate, cross-linked polyvinyl pyrrolidone, cross-linked sodium carboxymethyl cellulose) surface-active or dispersing agent.
  • a binder e.g inert diluent, preservative disintegrant (e.g. sodium starch glycolate, cross-linked polyvinyl pyrrolidone, cross-linked sodium carboxymethyl cellulose) surface-active or dispersing agent.
  • Moulded tablets may be made by moulding in a suitable machine a mixture of the powdered compound moistened with an inert liquid diluent.
  • the tablets may optionally be coated or scored and may be formulated so as to provide slow or controlled release of the active ingredient therein using, for example, hydroxypropylmethyl cellulose in varying proportions to provide the desired release profile. Tablets may optionally be provided with an enteric coating, to provide release in parts of the gut other than the stomach.
  • the compound, composition or combinations of the invention may be suitable for topical administration in the mouth including lozenges comprising the active ingredient in a flavoured base, usually sucrose and acacia or tragacanth gum; pastilles comprising the active ingredient in an inert basis such as gelatine and glycerin, or sucrose and acacia gum; and mouthwashes comprising the active ingredient in a suitable liquid carrier.
  • lozenges comprising the active ingredient in a flavoured base, usually sucrose and acacia or tragacanth gum
  • pastilles comprising the active ingredient in an inert basis such as gelatine and glycerin, or sucrose and acacia gum
  • mouthwashes comprising the active ingredient in a suitable liquid carrier.
  • the compound, composition or combinations of the invention may be suitable for topical administration to the skin may comprise the compounds dissolved or suspended in any suitable carrier or base and may be in the form of lotions, gel, creams, pastes, ointments and the like.
  • suitable carriers include mineral oil, propylene glycol, polyoxyethylene, polyoxypropylene, emulsifying wax, sorbitan monostearate, polysorbate 60, cetyl esters wax, cetearyl alcohol, 2-octyldodecanol, benzyl alcohol and water.
  • Transdermal patches may also be used to administer the compounds of the invention.
  • the compound, composition or combination of the invention may be suitable for parenteral administration include aqueous and non-aqueous isotonic sterile injection solutions which may contain anti-oxidants, buffers, bactericides and solutes which render the compound, composition or combination isotonic with the blood of the intended recipient; and aqueous and non-aqueous sterile suspensions which may include suspending agents and thickening agents.
  • the compound, composition or combination may be presented in unit-dose or multi-dose sealed containers, for example, ampoules and vials, and may be stored in a freeze-dried (lyophilised) condition requiring only the addition of the sterile liquid carrier, for example water for injections, immediately prior to use.
  • Extemporaneous injection solutions and suspensions may be prepared from sterile powders, granules and tablets of the kind previously described.
  • Preferred unit dosage composition or combinations are those containing a daily dose or unit, daily sub-dose, as herein above described, or an appropriate fraction thereof, of the active ingredient.
  • composition or combination of this invention may include other agents conventional in the art having regard to the type of composition or combination in question, for example, those suitable for oral administration may include such further agents as binders, sweeteners, thickeners, flavouring agents disintegrating agents, coating agents, preservatives, lubricants and/or time delay agents.
  • suitable sweeteners include sucrose, lactose, glucose, aspartame or saccharine.
  • Suitable disintegrating agents include cornstarch, methylcellulose, polyvinylpyrrolidone, xanthan gum, bentonite, alginic acid or agar.
  • Suitable flavouring agents include peppermint oil, oil of Wintergreen, cherry, orange or raspberry flavouring.
  • Suitable coating agents include polymers or copolymers of acrylic acid and/or methacrylic acid and/or their esters, waxes, fatty alcohols, zein, shellac or gluten.
  • Suitable preservatives include sodium benzoate, vitamin E, alpha-tocopherol, ascorbic acid, methyl paraben, propyl paraben or sodium bisulphite.
  • Suitable lubricants include magnesium stearate, stearic acid, sodium oleate, sodium chloride or talc.
  • Suitable time delay agents include glyceryl monostearate or glyceryl distearate.
  • the compound of Formula (I), (II), sub-formulae can be administered in combination with another drug.
  • the compound can be used in combination with other drugs as a therapeutic intervention for NIK mediated disease.
  • the term "combination” relates to the co-administration of another active ingredient to a single patient, and are intended to include treatment regimens in which the agents are not necessarily administered by the same route of administration or at the same time.
  • the therapeutic compounds or treatments used in such combination therapies may be administered together with a compound of Formula (I), (II), sub-formulae or a pharmaceutically acceptable salt, solvate or prodrug, one after the other, separately in one combined unit dosage or in separate unit dosage forms.
  • the present invention provides a method of treating NIK mediated diseases.
  • the method of treating NIK mediated diseases is via NIK inhibition.
  • the present invention provides a method of treating proliferative diseases, comprising a step of administering a compound of Formula (I) or a pharmaceutically acceptable salt, solvate, stereoisomer or prodrug thereof to a subject in need thereof, wherein each of Xi, X2, X3, X4 and X5 is independently C or N; Ri and R2 are independently H, or optionally substituted alkyl;
  • R3 is H, halo, or cyano
  • R4 is optionally substituted alkyl
  • R is optionally substituted heteroaryl
  • R4 and Rs is linked to give optionally substituted cycloalkyl, or optionally substituted heterocycyl.
  • the present invention provides a method of treating proliferative diseases, comprising a step of administering a compound of Formula (la) or a pharmaceutically acceptable salt, solvate, stereoisomer or prodrug thereof to a subject in need thereof, wherein Xi, X2, X4, X5, Ri, R2, R3, R4 and Rs are as disclosed herein.
  • proliferative disease refers to a disease which cells are growing and increasing in number rapidly. A unifying concept is that excessive proliferation of cells and turnover of cellular matrix contribute significantly to the pathogenesis of several diseases.
  • the terms broadly encompasses any neoplastic disease including those which are potentially malignant (pre-cancerous) or malignant (cancerous). The term therefore encompasses the treatment of tumours.
  • proliferative disease also encompasses atherosclerosis, rheumatoid arthritis, psoriasis, fibrosis, idiopathic pulmonary fibrosis, scleroderma, cirrhosis of the liver, stem cell and diabetic retinopathy.
  • tumor is used generally to define any malignant cancerous or pre-cancerous cell growth, and may include leukemias and carcinomas such as melanomas, colon, lung, ovarian, skin, breast, pancreas, pharynx, brain prostate, CNS, and renal cancers, as well as other cancers.
  • the compound may be used in the treatment of a cancer.
  • cancer may be but not limited to, cervical, colon, breast, bladder, head and neck, gastric, lung, ovarian, prostate, thyroid, non-small-cell lung, chronic lymphocytic leukemia, mesothelioma, melanoma, pancreatic adenocarcinoma, basal cell carcinoma, osteosarcoma, hepatocellular carcinoma, Wilm's tumor, medulloblastoma or chola ng io-carci noma.
  • the present invention provides a use of a compound of Formula (I), sub-formulae or a pharmaceutically acceptable salt, solvate, stereoisomer or prodrug thereof in the manufacture of a medicament for treating proliferative diseases in a patient in need thereof.
  • the present invention provides a compound of Formula (I), sub-formulae or a pharmaceutically acceptable salt, solvate, stereoisomer or prodrug thereof for use in treating proliferative diseases in a patient in need thereof.
  • the present invention provides a method of treating proliferative diseases, comprising a step of administering a compound of Formula (II) or a pharmaceutically acceptable salt, solvate, stereoisomer or prodrug thereof to a subject in need thereof,
  • X3, X4 are C and X5 is N, or X3, X4 are N and X5 is C, or X3, X4 and X5 is C;
  • Ri and R2 are independently H, or optionally substituted alkyl
  • R3 is H, halo, or cyano
  • R4 is optionally substituted alkyl
  • Rs is optionally substituted isoxazolyl, optionally substituted pyridinyl, 5- methylthiazolyl; or
  • R4 and Rs is linked to give optionally substituted cycloalkyl, or optionally substituted heterocycyl.
  • the present invention provides a method of treating proliferative diseases, comprising a step of administering a compound of Formula (Ila) or a pharmaceutically acceptable salt, solvate, stereoisomer or prodrug thereof to a subject in need thereof, wherein X4, X5, Ri, R2, R3, R4 and Rs are as disclosed herein.
  • the present invention provides a use of a compound of Formula (II), sub-formulae or a pharmaceutically acceptable salt, solvate, stereoisomer or prodrug thereof in the manufacture of a medicament for treating proliferative diseases in a patient in need thereof.
  • the present invention provides a compound of Formula (II), sub-formulae or a pharmaceutically acceptable salt, solvate, stereoisomer or prodrug thereof for use in treating proliferative diseases in a patient in need thereof.
  • the present invention provides a method of treating fibrosis, comprising a step of administering a compound of Formula (I) or a pharmaceutically acceptable salt, solvate, stereoisomer or prodrug thereof to a subject in need thereof, wherein each of Xi, X2, X3, X4 and X5 is independently C or N; Ri and R2 are independently H, or optionally substituted alkyl;
  • R3 is H, halo, or cyano
  • R4 is optionally substituted alkyl
  • R is optionally substituted heteroaryl
  • R4 and Rs is linked to give optionally substituted cycloalkyl, or optionally substituted heterocycyl.
  • the present invention provides a method of treating fibrosis, comprising a step of administering a compound of Formula (la) or a pharmaceutically acceptable salt, solvate, stereoisomer or prodrug thereof to a subject in need thereof, wherein Xi, X2, X4, X5, Ri, R2, R3, R4 and Rs are as disclosed herein.
  • the compounds can be used in the treatment of fibrosis.
  • fibrosis can be but not limited to, pulmonary fibrosis, liver fibrosis or renal fibrosis.
  • Fibrosis describes the development of fibrous connective tissue as a reparative response to injury or damage. Fibrosis may refer to the connective tissue deposition that occurs as part of normal healing or to the excess tissue deposition that occurs as a pathological process. Fibrosis involves stimulated fibroblasts laying down connective tissue, including collagen and glycosaminoglycans. The process is initiated when immune cells such as macrophages release soluble factors that stimulate fibroblasts. For example, pro-fibrotic mediator, TGF-gl, is released by macrophages as well as any damaged tissue between surfaces called interstitium.
  • TGF-gl pro-fibrotic mediator
  • CTGF connective tissue growth factor
  • PDGF platelet-derived growth factor
  • IL-10 interleukin 10
  • the compounds can be used to treat COVID related fibrosis such as long and/or post COVID induced pulmonary fibrosis.
  • Long COVID is a term to describe the effects of Covid-19 that continue for weeks or months beyond the initial illness. It is believed that fibrosis resulting from COVID is (or is mechanistically similar to) a NIK mediated disease, and/or which can be modelled using a bleomycin (BLM) induced lung fibrosis model.
  • BLM model is a model of the fibrotic response following acute lung injury, and acute lung injury and acute respiratory distress syndrome (ARDS) are the major cause of mortality in COVID-19. Therefore, compounds developed for treating fibrosis using BLM models can be beneficial in COVID-19, both in the acute phase of the illness and in preventing long-term complications.
  • COVID-induced pulmonary fibrosis caused by COVID can be estimated based on a 15-year observational study of lung pathology after SARS. Most SARS patients with fibrotic lung damage recovered within the first year and then remained healthy; however, in 20% of the cases, significant fibrosis progression was found in 5- 10 years. Based on these data, the incidence rate of post-COVID lung fibrosis can be estimated at 2-6% after moderate illness. The persistent respiratory complications may cause substantial population morbidity, long-term disability, and even death due to the lung fibrosis progression.
  • the present invention provides a use of a compound of Formula (I), sub-formulae or a pharmaceutically acceptable salt, solvate, stereoisomer or prodrug thereof in the manufacture of a medicament for treating fibrosis in a patient in need thereof.
  • the present invention provides a compound of Formula (I), sub-formulae or a pharmaceutically acceptable salt, solvate, stereoisomer or prodrug thereof for use in treating fibrosis in a patient in need thereof.
  • the present invention also relates to a method of treating fibrosis, comprising a step of administering a compound of Formula (II) or a pharmaceutically acceptable salt, solvate, stereoisomer or prodrug thereof, wherein
  • X3, X4 are C and X5 is N, or X3, X4 are N and X5 is C, or X3, X4 and X5 is C;
  • Ri and R2 are independently H, or optionally substituted alkyl
  • R3 is H, halo, or cyano
  • R4 is optionally substituted alkyl
  • Rs is optionally substituted isoxazolyl, optionally substituted pyridinyl, 5- methylthiazolyl; or
  • R4 and Rs is linked to give optionally substituted cycloalkyl, or optionally substituted heterocycyl.
  • the present invention also relates to a method of treating fibrosis, comprising a step of administering a compound of Formula (Ila) or a pharmaceutically acceptable salt, solvate, stereoisomer or prodrug thereof,
  • the present invention also relates to a use of a compound of Formula (II), sub-formulae or a pharmaceutically acceptable salt, solvate, stereoisomer or prodrug thereof in the manufacture of a medicament for treating fibrosis in a patient in need thereof.
  • the present invention also relates to a compound of Formula (II), sub-formulae or a pharmaceutically acceptable salt, solvate, stereoisomer or prodrug thereof for use in treating fibrosis in a patient in need thereof.
  • the present invention provides a method of treating inflammatory diseases and/or immune inflammatory diseases, comprising a step of administering a compound of Formula (I) or a pharmaceutically acceptable salt, solvate, stereoisomer or prodrug thereof to a subject in need thereof, wherein each of Xi, X2, X3, X4 and X5 is independently C or N; Ri and R2 are independently H, or optionally substituted alkyl;
  • R3 is H, halo, or cyano
  • R4 is optionally substituted alkyl
  • R is optionally substituted heteroaryl
  • R4 and Rs is linked to give optionally substituted cycloalkyl, or optionally substituted heterocycyl.
  • the present invention provides a method of treating inflammatory diseases and/or immune inflammatory diseases, comprising a step of administering a compound of
  • Inflammation disease is when the immune system attacks the body's own tissues, resulting in inflammation. Inflammation is the body's normal response to illness and infection. But sometimes inflammation can be misdirected. The immune system attacks healthy tissue instead. Asthma is an example of an inflammatory disease.
  • An immune-mediated inflammatory disease is any of a group of conditions or diseases that lack a definitive etiology, but which are characterized by common inflammatory pathways leading to inflammation, and which may result from, or be triggered by, a dysregulation of the normal immune response.
  • IMID immune-mediated inflammatory disease
  • auto immune diseases such as rheumatoid arthritis, psoriasis, SLE, inflammatory bowel diseases (CD & UC)
  • immune mediated inflammatory pathways play vital role for the disease pathogenesis.
  • Respiratory disorders such as Asthma and COPD are mainly mediated through inflammatory cells.
  • the present invention provides a use of a compound of Formula (I), sub-formulae or a pharmaceutically acceptable salt, solvate, stereoisomer or prodrug thereof in the manufacture of a medicament for treating inflammatory diseases and/or immune inflammatory diseases in a patient in need thereof.
  • the present invention provides a compound of Formula (I), sub-formulae or a pharmaceutically acceptable salt, solvate, stereoisomer or prodrug thereof for use in treating inflammatory diseases and/or immune inflammatory diseases in a patient in need thereof.
  • the present invention provides a method of treating inflammatory diseases and/or immune inflammatory diseases, comprising a step of administering a compound of Formula (II) or a pharmaceutically acceptable salt, solvate, stereoisomer or prodrug thereof to a subject in need thereof, wherein
  • X3, X4 are C and Xs is N, or X3, X4 are N and Xs is C, or X3, X4 and Xs is C;
  • Ri and R2 are independently H, or optionally substituted alkyl
  • R3 is H, halo, or cyano
  • R4 is optionally substituted alkyl
  • Rs is optionally substituted isoxazolyl, optionally substituted pyridinyl, 5- methylthiazolyl; or
  • R4 and Rs is linked to give optionally substituted cycloalkyl, or optionally substituted heterocycyl.
  • the present invention provides a method of treating inflammatory diseases and/or immune inflammatory diseases, comprising a step of administering a compound of Formula (Ila) or a pharmaceutically acceptable salt, solvate, stereoisomer or prodrug thereof to a subject in need thereof, wherein X4, X5, Ri, R2, R3, R4 and Rs are as disclosed herein.
  • the present invention provides a use of a compound of Formula (II), sub-formulae or a pharmaceutically acceptable salt, solvate, stereoisomer or prodrug thereof in the manufacture of a medicament for treating inflammatory diseases and/or immune inflammatory diseases in a patient in need thereof.
  • the present invention provides a compound of Formula (II), sub-formulae or a pharmaceutically acceptable salt, solvate, stereoisomer or prodrug thereof for use in treating inflammatory diseases and/or immune inflammatory diseases in a patient in need thereof.
  • the compound is characterised by a maximum tolerated dose of less than about 200 mg/kg, about 180 mg/kg, about 160 mg/kg, about 140 mg/kg, about 120 mg/kg, or about 100 mg/kg.
  • the dose may be administered twice a day.
  • the present invention also provides a method of inhibiting NIK in a cell, comprising the steps of contacting the compounds as disclosed herein or a pharmaceutically acceptable salt, solvate, stereoisomer or prodrug thereof with the cell.
  • the cell may in vitro or in vivo.
  • Exemplary compounds of the present invention are as follows:
  • the compounds described herein may be prepared by techniques known in the art.
  • the compounds described herein may be prepared by following the reaction sequence as depicted in Scheme-1 to Scheme-10.
  • the following schemes where specific bases, acids, reagents, solvents, coupling agents, etc., are mentioned, it is understood that other bases, acids, reagents, solvents, coupling agents etc., known in the art may also be used and are therefore included within the scope of the present invention.
  • Variations in reaction conditions, for Example, temperature and/or duration of the reaction which may be used as known in the art, are also within the scope of the present invention. All the isomers of the compounds in described in these schemes, unless otherwise specified, are also encompassed within the scope of this invention.
  • Step 1 The 2- Chloropyrollo pyrimidine (1.0 eq), l,3-dibromobenzene( 1.5 eq), K2CO3 (2.0 eq), Cui (0.1 eq) and L-proline (0.2 eq) in DMSO (5 Times) taken in RB flask was stirred at 120°C for 16hrs. On completion of reaction as monitored by TLC, the reaction was quenched in ice-cold water, extracted into ethyl acetate (3 x 25ml) and evaporated to give crude product. The crude mixture was purified using flash column chromatography to obtain the pure compound.
  • Step 2 The product of Step 1 (1.0 eq), NFUOH solution ( 10 Times), NaF (2.0 eq), K2CO3 (2.0 eq) were taken in steel bomb (sealed, pressure vessel)and stirred at 120°C for 16hrs. Evaporated the solvent, redissolved in 20% Methanol in DCM, filtered off the inorganic insoluble compound and evaporated the filtrate to give required product.
  • scheme-5 compound 4 (5.0 g, 14.9 mmol) in DMF (40 mL) was added /V-chlorosuccinimide (2.4 g, 17.9 mmol) at room temperature and stirred for 24 h. Reaction was monitored by TLC. On completion of the reaction, ice-cold water was added and stirred for 30 min. Solid precipitated was filtered through Buchner funnel and dried under vacuum give scheme-5 compound 5 (4.9 g, 89%) as off white solid.
  • reaction mixture was heated at 100°C for 16 h in sealed tube. Reaction mixture was cooled to room temperature, quenched by the addition of water. The reaction mixture was extracted with EtOAc (30 mL X 3). Combined organic layer was washed with brine (50 mL), dried over anhydrous sodium sulphate, filtered and concentrated. The resulting residue was purified by flash column chromatography to afford title scheme-9 compound 3 (480 mg, 36%) as off white solid.
  • Salts of compound of Formula (I) are obtained by dissolving the compound in a suitable solvent and then treated with the desired acid or base as described in Berge S.M. et al. "Pharmaceutical Salts, a review article in Journal of Pharmaceutical sciences volume 66, page 1-19 (1977)" and in “handbook of pharmaceutical salts properties, selection, and use by P.H. Einrich Stahland Camille G. wasmuth, Wiley-VCH (2002)".
  • stereoisomers of the compounds of Formula (I) of the present invention may be prepared by stereospecific syntheses or resolution of the achiral compound using an optically active amine, acid or complex forming agent, and separating the diastereomeric salt/complex by fractional crystallization or by column chromatography. lOOmg of 7-(3-bromophenyl)-pyrrolo[2,3-d]pyrimidin-2-amine, 26mg of Pd(dppf)Cl2.CH2Cl2 and 15mg of copper (I) iodide were added to a dry 10ml round- bottomed flask.
  • reaction mixture was cooled at room temperature and poured in ice water. Filtered off the solid, redissolved in DCM-MeOH, absorbed over silica gel and purified using Hexane: Ethyl acetate. Isolated polar spot as product.
  • Pd(dppf)Cl2.CH2Cl2 and lOmg of copper (I) iodide were added to a dry 10ml round- bottomed flask. 0.1ml of 1-ethynlycyclobutan-l-ol, 0.5ml of DIPEA and 1ml of dry DMF were then added before the mixture was purged with argon. The mixture was stirred at 100°C for 2h under argon. After 2h, the reaction was complete (based on TLC of reaction mixture) and it was slowly added to ice water and stirred vigorously. The resulting mixture was then extracted 3 times with ethyl acetate before the organic layers were combined, dried over magnesium sulphate and the solvent removed to yield the crude product.
  • Compound 4 Compound 8, Compound 9, and Compound 10 can be prepared using similar process as that of Compound 3 using appropriate starting materials.
  • the resulting mixture was then extracted 3 times with ethyl acetate before the organic layers were combined, dried over magnesium sulphate and the solvent removed to yield the crude product.
  • the crude product was then purified using flash column chromatography to yield a somewhat pure product which was precipitated with DCM to obtain the pure product at 14mg (27%).
  • Compound 6, and Compound 14 can be prepared using similar process as that of Compound 5 using appropriate starting materials.
  • Butyn-2-ol (R-isomer), 20mg of Pd(dppf)CI2.CH2CI2 and 5 mg of copper (I) iodide were added to a dry 10ml round-bottomed flask. 0.4ml of DIPEA and 1ml of dry DMF were then added before the mixture was purged with argon. Then the reaction mixture purged with Argon was heated in round bottom flask under Argon at 95°C for 1 hour, 90% completion of reaction observed, reaction stopped. Cooled to room temperature, poured into ice water, filtered off the solid, washed with water and dried to crude product (105 mg). Purified product on 10 g SNAP silica cartridge at 70% ethyl acetate in hexane to give titled product (yield: 30 mg)
  • reaction mixture On completion of about 80% of reaction as confirmed by TLC, reaction mixture was cooled at room temperature and poured in ice water but did not give solid, extracted with ethyl acetate and washed with water and brine, dried over MgSO4 to give crude product (156 mg) absorbed over silica gel and purified using Hexane: Ethyl acetate. Isolated polar spot as product.
  • reaction was complete (based on TLC of reaction mixture) and it was slowly added to ice water and stirred vigorously. The resulting mixture was filtered to yield the crude product. The crude product was then purified using flash column chromatography to yield the pure product at 50mg (92%).
  • Compound 19 can be prepared using similar process as that of Compound 21 using appropriate starting materials.
  • Comparator 1 lOOmg of 7-(3-bromophenyl)-pyrrolo[2,3-d]pyrimidin-2-amine, 20mg of Pd(dppf)Cl2.CH2Cl2 and lOmg of copper (I) iodide were added to a dry 10ml round- bottomed flask. 0.1ml of 2-methyl-3-butyn-2-ol, 0.5ml of DIPEA and 1ml of dry DMF were then added before the mixture was purged with argon. The mixture was stirred at 90-100°C for 20mins under argon.
  • reaction was complete (based on TLC of reaction mixture) and it was slowly added to ice water and stirred vigorously. The resulting mixture was filtered to yield the crude product. The crude product was then purified using flash column chromatography to yield the pure product at 75mg.
  • Comparator 3 can be prepared using similar process as that of Comparator 2 using appropriate starting materials.
  • Comparator 4 is prepared as per Scheme-3, mentioned above starting with appropriate starting materials.
  • Pd(dppf)Cl2.CH2Cl2 and lOmg of copper (I) iodide were added to a dry 10ml round- bottomed flask. 0.1ml of 3,4-dimethyl-l-pentyn-3-ol, 0.5ml of DIPEA and 1ml of dry DMF were then added before the mixture was purged with argon. The mixture was stirred at 100°C for 2h under argon. After 2h, the reaction was complete (based on TLC of reaction mixture) and it was slowly added to ice water and stirred vigorously.
  • the resulting mixture was then extracted 3 times with ethyl acetate before the organic layers were combined, dried over magnesium sulphate and the solvent removed to yield the crude product.
  • the crude product was then purified using flash column chromatography to yield a somewhat pure product (51mg) which was then further purified using flash column chromatography to yield the pure product at lOmg.
  • Comparator 6 is prepared as per Scheme-8, mentioned above.
  • Comparator 8 l- ⁇ [3-(2-amino-4-methoxy-7H-pyrrolo[2,3-tf]pyi"iniidin-7- yl)phenyl]ethynyl ⁇ cyclobutan-l-ol
  • reaction was complete (based on TLC of reaction mixture) and it was slowly added to ice water and stirred vigorously. The resulting mixture was then extracted 3 times with ethyl acetate before the organic layers were combined, dried over magnesium sulphate and the solvent removed to yield the crude product. The crude product was then purified using flash column chromatography to yield the pure product at 37mg (70%).
  • the compounds demonstrated nano-molar NIK activity.
  • TGF-pi TGF-pi (30 ng/mL) was used to stimulate for the fibrotic markers expression for 12h.
  • Compound 12 (30 pM)
  • Compound 21 (30 pM)
  • Nintedanib (30 pM) were pre-treated before the TGF-pi stimulation.
  • TGF- 81 stimulation fibroblasts were lysed and analysed for mRNA expression of CTGF, Collagen 1A, fibronectin, elastin, o- SMA, and TGF-pi by reverse transcription polymerase chain reaction (RT-PCR).
  • Compound 12 and Compound 21 also significantly reduced most of the fibrotic markers mRNA expression in TGF- pi stimulated normal lung fibroblast ( Figure 1). Nintedanib was used as standard compound.
  • WI-38 cells were stimulated with TGF-pi (30 ng/mL) for 12 h in pretreated with Compound 12 (30 pM), Compound 21 (30 pM) and Nintedanib (30 pM) for lOmin.
  • the mRNA expressions of CTGF, Collagen 1A, fibronectin, elastin, o-SMA, and TGF-pi in WI-38 cells were analyzed by reverse transcription-polymerase chain reaction (RT- PCR) and the results were summarized in Figure 1.
  • Statistical analysis was performed using Graph Pad Prism-Version 8 by one way ANOVA Dunnet's test. *P ⁇ 0.05, **P ⁇ 0.01, ***p ⁇ 0.001, Treated groups vs. TGF-pi stimulated group; # P ⁇ 0.001, TGF- pi stimulated group vs unstimulated group.
  • TGF-pi (20 ng/mL) was used to stimulate the cells for fibrotic markers expression.
  • Compound 12 (10 pM), Compound 21 (10 pM), and Nintedanib (10 pM) were pre-treated 10 min prior to TGF stimulation and post 12 h of TGF stimulation.
  • the cells were harvested and analysed to study the effect of compounds on fibrotic markers such as CTGF, collagen 1A, and fibronectin.
  • Important fibrotic markers such as CTGF, Collagen 1A and Fibronectin were upregulated in TGFp-1 stimulated cells compared to untreated WI-38 cells.
  • FMT assay is one of the important in vitro pharmacological experimental study to screen the anti-fibrotic molecules. This assay was performed in 96 well plate. At day 0, WI-38 cells were plated in the complete medium and on day 1 cells were starved with starvation medium for 24 h. Compound treatment were started on day 3, Nintedanib (clinical standard) and ALK5 inhibitor (TGF-pi receptor antagonist; an assay standard) were kept for as comparators. Compound 12 and Compound 21 were preincubated for one hour prior to TGF-pi stimulation at various concentration from 0.003pM to 10 pM to get a dose dependent curve.
  • MTD maximum tolerated dose
  • Design Female NOD-SCID and Balb/C mice were used in the MTD study for Compound 12 and Compound 21 respectively. Animals were dosed with Compound 12 and Compound 21 (100 mg/kg, PO) twice a day. The animals were watched carefully during the first two hours for any adverse reactions, and monitored morning and evening daily. Compound 12 and Compound 21 were dosed for five consecutive days. On day 6, repeated and high dose PK study was performed on the same animals for both compounds ( Figure 7). Animals were then sacrificed and vital organs were collected for histological observation.
  • V were treated with Compound 12 (50 mg/kg, bid, po), Compound 21(50 mg/kg, bid, po), and Nintedanib (60 mg/kg, qd, po) respectively until the study termination.
  • animals were sacrificed, and bronchoalveolar Lavage (BAL) was performed to collect lung fluid. The whole lung was dissected out, the left lung was taken for histopathological analysis and the right lung was taken for future analysis.
  • BAL bronchoalveolar Lavage
  • the fibrotic score of the lung is an important parameter to understand the lung pathological condition and the disease stage and it is quantified following the score as described by Ashcroft et al (1988), the reference of which is herein incorporated.
  • BLM treatment significantly increased the fibrotic score compared to vehicle control, which was markedly reduced by Compound 12, Compound 21, and Nintedanib treatment ( Figure 9 and 11).
  • FIG 11 shows that BLM caused varying degree of fibrosis in mouse lung parenchyma, accompanied with infiltration of mononuclear cell and polymorphs. It also caused alveolar/bronchial hyperplasia, haemorrhages, and bronchial degeneration. Fibrosis presence is demonstrated with picrosirius red (PSR) and masson's trichrome (MT) staining. Compared to BLM control, the presence of fibrosis is noticed in following groups in descending order: Compound 12 >STD (Nintedanib)> Compound 21. Normal mice group did not show any fibrosis.
  • NIK is thus the master regulator of the noncanonical NF-KB pathway and its inhibition can modulate the associated disease effectively, especially in fibrosis, inhibition of NIK using small molecules approach reduced lung fibrosis progression. It was strongly evidenced by the above-described in vitro and in vivo lung fibrosis models. Upon this, following additional studies were performed. To find the effective role of NIK involvement in lung fibrosis, we have selected three types of NIK siRNA called hs MAP3K14 13.1, 13.5, 13.10, now it is called NIK SiRNA 1, 2, and 3 respectively and the selected NIK siRNAs were evaluated first for their knockdown potential of NIK in U2OS cells, further evaluated for their antifibrotic effect in Wi38 cells.
  • TWEAK Tumor necrosis factor-like weak inducer of apoptosis
  • RelB/p52 heterodimers translocate to the nucleus to act as a transcription factor.
  • TWEAK (20 ng/mL) was used for stimulation and three hours post-stimulation, cells were lysed and then NIK expression and P52 expression levels were analyzed. Before the TWEAK stimulation, cells were transfected with NIK siRNA and incubated for 28 hours. In the study, noncanonical NF-KB markers such as NIK and pP52 expressions were upregulated upon TWEAK stimulation compared to unstimulated U2OS cells. Transfection with NIK siRNA 1, 2, and 3 markedly reduced the NIK expression levels and the results showed that all three NIK siRNA showed NIK inhibition ( Figure 12).
  • NIK siRNA2 was also screened in FMT assay. WI-38 cells were transfected with NIK siRNA2 and incubated for 28 hours before the TGF-pi stimulation. The remaining procedure was followed as described above in the FMT assay and the results were very impressive, the percentage inhibition of NIK siRNA2 on the reduction of a-SMA fiber count was 63.3 ⁇ 8.1%.
  • the NIK inhibitors of the present disclosure were subjected to screening on various in- vitro and in-vivo anti-fibrotic experiments, and results strongly suggested that compound 21 in particular showed marked antifibrotic potential in in-vitro assays by inhibiting TGF-B1 -induced fibrotic markers expression and reducing a-SMA fiber count in FMT assay.
  • the anti-fibrotic effect of compound 21 on in-vitro assay was also translated in an in-vivo mouse BLM-induced lung fibrosis model.
  • compound 21 showed superior in-vivo anti-fibrotic efficacy in mouse BLM-induced lung fibrosis model compared to current standard care Nintedanib, also compound 21 showed good in-vivo PK exposure.
  • NIK knockdown significantly reduced fibrotic marker expressions such as CTGF and Collagen Al upon TGF-pl stimulation, and also reduced o-SMA fiber in FMT assay.

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Abstract

La présente divulgation concerne des composés et des méthodes associées pour traiter des maladies prolifératives, la fibrose, des maladies inflammatoires et/ou des maladies inflammatoires immunitaires. La présente divulgation concerne également des méthodes d'inhibition de la kinase induisant NF-κB dans une cellule.
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WO2018037059A1 (fr) * 2016-08-24 2018-03-01 F. Hoffmann-La Roche Ag Dérivés de 2-azabicyclo [3.1.0] hexan-3-one et procédés d'utilisation
CN109810110A (zh) * 2017-11-22 2019-05-28 中国科学院上海药物研究所 一种具有2-氨基嘧啶结构的化合物，其制备方法和用途

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Publication number Priority date Publication date Assignee Title
WO2018037059A1 (fr) * 2016-08-24 2018-03-01 F. Hoffmann-La Roche Ag Dérivés de 2-azabicyclo [3.1.0] hexan-3-one et procédés d'utilisation
CN109810110A (zh) * 2017-11-22 2019-05-28 中国科学院上海药物研究所 一种具有2-氨基嘧啶结构的化合物，其制备方法和用途

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INAYAMA MAMI, NISHIOKA YASUHIKO, AZUMA MOMOYO, MUTO SUSUMU, AONO YOSHINORI, MAKINO HIDEKI, TANI KENJI, UEHARA HISANORI, IZUMI KEIS: "A Novel IκB Kinase-β Inhibitor Ameliorates Bleomycin-induced Pulmonary Fibrosis in Mice", AMERICAN JOURNAL OF RESPIRATORY AND CRITICAL CARE MEDICINE, AMERICAN THORACIC SOCIETY, US, vol. 173, no. 9, 1 May 2006 (2006-05-01), US , pages 1016 - 1022, XP093093501, ISSN: 1073-449X, DOI: 10.1164/rccm.200506-947OC *
LI, Z. ET AL.: "Discovery of a Potent and Selective NF-KB-Inducing Kinase (NIK) Inhibitor That Has Anti-inflammatory Effects in Vitro and in Vivo", JOURNAL OF MEDICINAL CHEMISTRY, vol. 63, no. 8, 27 March 2020 (2020-03-27), pages 4388 - 4407, XP055884421, [retrieved on 20230510], DOI: 10.1021/ACS.JMEDCHEM.OC00396 *

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