# Patent

## Title
Methyl/fluoro-pyridinyl-methoxy substituted pyridinone-pyridinyl compounds and fluoro-pyrimidinyl-methoxy substituted pyridinone-pyridinyl compounds

## Abstract

The present disclosure provides methyl/fluoro-pyridinyl-methoxy substituted pyridinone-pyridinyl compounds and fluoro-pyrimidinyl-methoxy substituted pyridinone-pyridinyl compounds useful in the treatment of p38 kinase mediated diseases, such as lymphoma and auto-inflammatory disease, including rheumatoid arthritis, having the structure of Formula (I):




wherein R1, R2, R3, R4, R5 and X are as defined in the detailed description; pharmaceutical compositions comprising at least one of the compounds; and methods for treating p38 kinase mediated diseases using the compound.

## Background
CROSS-REFERENCE TO RELATED APPLICATIONS
This application is a non-provisional application which claims the benefit of and priority to Provisional Application No. 61/832,736, filed 7 Jun. 2013, the entire disclosure of which is incorporated herein by reference.


FIELD
The present disclosure generally relates to a compound having enzyme inhibitory activity, pharmaceutical compositions comprising the compound, and methods useful for treating diseases. More specifically, the present disclosure relates to a class of pyridinone-pyridinyl compounds, pharmaceutical compositions comprising the compound, and methods useful for treating p38 kinase mediated diseases.
BACKGROUND
Mitogen-activated protein kinases (MAPK) are a conserved family of enzymes that relay and propagate external stimuli, using phosphorylation cascades to generate a coordinated cellular response to the environment. The MAPK are proline-directed serine/threonine-specific protein kinases that regulate cellular activities, such as gene expression, mitosis, differentiation, and cell survival/apoptosis. To date, four distinct classes of mammalian MAPK have been identified: the extracellular signaling kinases (ERK1 and 2), the c-jun N-terminal kinase-1 (JNK1-3), the p38 MAPK (p38α, β, γ, and δ), and ERK5. The MAPK are activated by the dual phosphorylation of Thr and Tyr residues within a TXY activation motif by coordinated dual-specificity MAPKK, where X is Glu, Pro, and Gly in ERK, JNK, and p38 MAPK, respectively. MAPK are 60-70% identical to each other, yet differ in their activation loop sequences and sizes. The activation loop is adjacent to the enzyme-active site, and its phosphorylation allows the enzyme to reposition active-site residues into the optimal orientation for substrate binding and catalysis. Downstream substrates of MAPK include mitogen-activated protein-kinase-activated protein (MAPKAP) kinases and transcription factors, the phosphorylation of which, either directly or indirectly, regulates gene expression at several points, including transcription, nuclear export, and mRNA stability and translation. The cellular consequences of MAPK activation include inflammation, apoptosis, differentiation, and proliferation.
Distinct genes encode four p38 MAPKinases in humans: p38α, β, γ, and δ. Significant amino acid sequence homology is observed among the 4 isoforms, with 60%-75% overall sequence identity and >90% identity within the kinase domains. Tissue-selective expression is observed, with p38γ found predominantly in skeletal muscle, p38δ in the testes, pancreas, and small intestine. In contrast, p38α and β are more ubiquitously expressed.
An understanding of the broad biologic and pathophysiological roles of p38 MAPK family members has grown significantly over the past decade, as has the complexity of the signaling network leading to their activation. Scientific exploration of this pathway from biological, cellular, and in vivo perspectives was largely enabled by the availability of well-behaved, selective, small-molecule inhibitors of p38 MAPK that target the α and, to a lesser extent, β isoforms. p38α MAPK is the major isoform involved in the immune and inflammatory response. As such its function is critical for the production and activity of multiple pro-inflammatory cytokines, including TNFα, IL-1, IL-6, and IL-8, in cells such as macrophages, monocytes, synovial cells, and endothelial cells. p38 MAPK is also responsible for the induction of key inflammatory enzymes such as COX2 and iNOS, the major sources of eicosanoids and nitric oxide at sites of inflammation, respectively. Additionally, the p38 MAPK pathway regulates the expression of matrix metalloproteinases (MMP), including MMP2, MMP9, and MMP13.
The use of selective and potent inhibitors has facilitated the discovery of several families of p38 MAPK substrates, including transcription factors, MAPKAP kinases, and other enzymes. p38 MAPK can directly phosphorylate several transcription factors, such as myocyte-specific enhancer binding factor 2C (MEF2C), CHOP, peroxisome proliferator-activated receptor (PPAR) α, PPAR γ co-activator 1 and p53. These transcription factors are involved in cellular functions such as apoptosis, gluconeogenesis, and synthesis of enzymes involved in fatty acid oxidation. p38 MAPK is also involved in the direct or indirect phosphorylation of enzyme substrates, such as cytosolic phospholipase A2, and the Cdc25 phosphatases, which are involved in the activation of cyclin-dependent protein kinase activity and cell-cycle regulation. Therefore in addition to its role in the inflammatory response, p38 MAPK has other functions associated with normal and abnormal cell growth and survival as well as cellular function and homeostasis.
The MAPKAP kinases—MK2, MK-3, and PRAK—are selectively phosphorylated by p38 MAPK, while the phosphorylation of MSK1/2, MNK1/2, and RSKb is catalyzed by both p38 MAPK and ERK. Activation of RSKb is thought to play a role in cell survival, although the identification of substrates has been difficult, due to the lack of specific inhibitors. MNK is involved in the phosphorylation of eukaryotic initiation factor-4E, which binds to the ‘cap’ structure of mRNA and enhances protein translation. MNK phosphorylates the mRNA binding protein hnRNP-A0, a protein that regulates mRNA stability of transcripts encoding inflammatory proteins. MSK1/2 is involved in the phosphorylation of the transcription factors CREB and ATF-1, which regulate AP-1 binding proteins. In addition, MSK1/2 can phosphorylate Histone H3, which is involved in chromatin remodeling. While evidence suggests that MSK and MNK play a role in the mediation of pro-inflammatory cytokines, in vivo data with selective inhibitors and/or knockout mice are lacking.
MK-2, MK-3, and PRAK, once phosphorylated and activated by p38 MAPK, share similar substrate specificities. All of these kinases can phosphorylate the small heat-shock protein Hsp27. Studies have shown that the PRAK- and MK3-deficient mice do not display any resistance to endotoxic shock or a decrease in lipopolysaccharide-(LPS)-induced cytokine production. In contrast, MK-2-deficient mice show a resistance to endotoxic shock and an impaired inflammatory response, as well as a significantly decreased production of cytokines such as TNFα, IFNγ and IL-6. Thus, the p38/MK2 axis specifically is necessary and sufficient for mediating pro-inflammatory responses.
Recently, Davidson et al (2004) Discovery and characterization of a substrate selective p38alpha inhibitor, Biochemistry 43:11658-71 described a novel approach for increasing selectivity of a p38 MAPK inhibitors. In these studies, a high throughput screen was carried out using an assay that measured the p38-dependent phosphorylation and activation of MK2. The p38:MK2 complex is very stable with a Kd of 6 nM. The binding affinity of p38 for MK2 is driven by the C-terminal domain of MK2 containing several positively charged amino acid residues. Crystallographic studies of the p38:MK2 complex demonstrated that the C-terminal region of MK2 wraps around p38α and binds to the negatively charged ED binding site. The tight binding of p38 to MK2 may give rise to conformational changes providing additional binding pockets for inhibitors that would specifically be dependent upon the p38:MK2 interaction.
Taking advantage of the p38:MK2 interaction and using MK2 as the p38 substrate, a novel inhibitor of p38α was discovered exhibiting interesting properties (Davidson et al). This inhibitor demonstrated substrate selectivity by preventing the p38α dependent phosphorylation of MK2 (Ki app 300 nM) while sparing the p38α dependent phosphorylation of ATF2 (Ki app>20 uM). This novel inhibitor is functionally unique compared with traditional p38 ATP competitive inhibitors that block the p38-dependent phosphorylation of all p38 substrates. A second independent study also describes p38 inhibitors with unique mechanistic properties. This work demonstrates a novel mechanism for the selective inhibition of the p38 dependent phosphorylation of MK2. Unlike the previous study of Davidson et al., these mechanistically unique compounds are competitive with ATP and stabilize the p38/MK2 complex. Taken together, these two studies clearly prove the concept that selective p38/MK2 axis blockade is achievable with small molecule inhibitors. In comparison to traditional p38 MAPK inhibitors these p38/MK2 inhibitors should retain or enhance potency and exhibit improved safety features in animal models of disease or in human clinical settings.
The p38/MK2 role in the regulation of inflammatory cytokines (TNFα, IL-1β, IL-6) and enzymes responsible for inflammation (COX-2, iNOS, and MMPs) makes it an attractive drug target. Several classical p38 MAPK inhibitors have progressed to testing in clinical trials. Some of these candidates have failed, for safety or other reasons, but several have reported clinical data in diseases such as rheumatoid arthritis, pain, Crohn's disease, acute coronary syndrome, multiple myeloma and chronic obstructive pulmonary disease. In addition to these diseases several IL-1β mediated diseases could be impacted by a p38 inhibitor based upon the key role for the p38 MAPK pathway in the biosynthesis and activity of this cytokine. These diseases include the family of cryopyrin associated periodic disorders (CAPS), chronic gout, diabetes, Still's disease, Familial Mediterranean Fever among others.
Rheumatoid arthritis (RA) is a systemic, autoimmune, chronic inflammatory disorder characterized by joint synovial inflammation leading to cartilage and bone destruction. Current treatment for RA includes oral disease modifying anti-rheumatic drugs (DMARDs) (methotrexate, leflunomide, sulfasalazine), and parenterally administered biologic agents specifically directed against IL-1 (Ankinra®) or TNFα (Enbrel®, Remicade®, and Humira®), two key proinflammatory cytokines implicated in RA pathogenesis. The superior efficacy of these latter agents is somewhat offset by potential shortcomings, including requirement for parenteral administration, difficult dose titration, poor reversibility due to prolonged plasma half-lives, induction of host neutralizing antibody responses and high cost of treatment. Based on a p38 inhibitor's potential to inhibit a broad range of pro-inflammatory mediators purported to play a central role in RA pathogenesis (including TNFα, IL-1β and IL-6) it is expected that a p38 inhibitor will have clinical efficacy equivalent or superior to biologies restricted to single cytokine modulation (e.g., TNFα). An orally administered DMARD with improved efficacy offers multiple advantages to both the patient and physician with respect to convenience and compliance of administration, lack of injection site/allergic reactions, superior dose titratability, and favorable cost of goods. A safe and effective p38 inhibitor thus potentially fulfills an evident unmet medical need and promises high potential to generate significant value for patients and physicians that deal with RA.
Several classical p38 MAPK inhibitors have progressed to testing in clinical trials. Some of these candidates have failed, for safety or other reasons, but several have reported clinical data in diseases such as rheumatoid arthritis, pain, Crohn's disease, acute coronary syndrome, multiple myeloma and chronic obstructive pulmonary disease. In addition to these diseases several IL-ip mediated diseases could be impacted by a p38 inhibitor based upon the key role for the p38 MAPK pathway in the biosynthesis and activity of this cytokine. These diseases include the family of cryopyrin associated periodic disorders (CAPS), chronic gout, diabetes, Still's disease, Familial Mediterranean Fever among others.
In addition to human inflammatory pathways, p38 MAPK has been linked to canine B cell growth and survival. The role of p38 MAPK in B cell growth suggests that inhibition of this enzyme may be therapeutically beneficial for the treatment of canine B cell lymphoma. Canine lymphoma is one of the most common malignancies diagnosed in companion animals representing 10-25% of canine neoplasms and >80% of the hematopoietic tumors. An orally available, selective B cell growth inhibitor would meet a significant unmet medical need.
Compounds useful for treating diseases and conditions caused or exacerbated by unregulated p38 MAP Kinase and/or TNF activity are described in WO 2000/017175 published 30 Mar. 2000. The compounds described therein include a class of substituted urea compounds.
Compounds useful for treating diseases and conditions caused or exacerbated by unregulated p38 MAP Kinase and/or TNF activity are described in WO 2000/071535 published 30 Nov. 2000. The compounds described therein include a class of indole-type compounds.
Compounds useful for treating diseases and conditions caused or exacerbated by unregulated p38 MAP Kinase and/or TNF activity are described in WO 2002/042292 published 30 May 2002. The compounds described therein include a class of coupled indole-type derivatives.
Compounds useful for prophylaxis or treatment of circulatory diseases, metabolic diseases and/or central nervous system diseases are described in WO 2008/062905 published 29 May 2008. The compounds described therein include an alkyl-pyrimidinone-phenyl compounds wherein the phenyl fragment is substituted with a cyclopropyl radical, e.g., 6-butyl-3-(3-cyclopropylphenyl)-2-methyl-5-{[2′-(5-oxo-4,5-dihydro-1,2,4-oxadizol-3-yl)biphenyl-4-yl]methyl}pyrimidin-4(3H)-one.
Various potential inhibitors or modulators of p38 kinase and the p38 kinase pathway are described in WO 2005/018557 published 3 Mar. 2005. The compounds described therein include di-fluorophenyl-methoxy-pyridinone-pyridyl compounds wherein the pyridyl fragment is substituted with various radicals including alkyl, alkenyl, hydroxyalkyl, halo, cyano, amino, carboxy, carbamoyl, methoxycarbonyl and hydroxyalkenylimino radicals.
Compounds useful for treating diseases and conditions caused or exacerbated by unregulated p38 MAP Kinase and/or TNF activity are described in US 2007/0167621 published 19 Jul. 2007. The compounds described therein include di-fluorophenyl-methoxy-pyrimidinone-phenyl compounds wherein the phenyl fragment is substituted with methyl amido radical.
Compounds useful for treating diseases and conditions caused or exacerbated by unregulated p38 MAP Kinase and/or TNF activity are described in WO 2004/087677 published 14 Oct. 2004. The compounds described therein include di-fluorophenyl-methoxy-pyrimidinone-phenyl compounds wherein the phenyl fragment is substituted with piperazinyl or a morpholinyl radical through a carbonyl bridge.
Pyrimidinone derivatives (as inhibitors of protein kinases and useful in treating disorders related to abnormal protein kinase activities such as inflammatory diseases and certain types of cancer), are described in WO 2007/081901 published 19 Jul. 2008. The compounds described therein include di-fluorophenyl-methoxy-pyrimidinone-phenyl compounds wherein the phenyl fragment is substituted with a cyclopropanyl or a morpholinyl radical through an amidoalkylamido bridge.
Pyrimidinone derivatives (as inhibitors of protein kinases and useful in treating disorders related to abnormal protein kinase activities such as inflammatory diseases and certain types of cancer) are described in WO 2008/153942 published 18 Dec. 2008. The compounds described therein include di-fluorophenyl-methoxy-pyrimidinone-phenyl compounds where the phenyl radical is substituted with cyclopentyl or a cyclohexyl radical through an amido bridge.
Compounds useful for treating diseases and conditions caused or exacerbated by unregulated p38 MAP Kinase and/or TNF activity are described in U.S. Pat. No. 7,067,540 published 27 Jun. 2007. The compounds described therein include di-fluorophenyl-methoxy-pyridinone-phenyl compounds wherein the phenyl radical is substituted with a five-membered heteroaryl radical (e.g., pyrazolyl or imidazolyl).
Substituted pyridinone-pyridinyl compounds useful in the treatment of p38 kinase mediated diseases, such as lymphoma and auto-inflammatory disease, are described in U.S. 2012/0142709 published on 7 Jun. 2012. The compounds described therein have a pyridinone-pyridinyl centroid with halo and alkyl substituents.
A counterpart of U.S. 2012/0142709 was published as WO 2012/078684 on 14 Jun. 2012.
SUMMARY
In one embodiment, there is provided a compound of Formula (I):




or a pharmaceutically acceptable salt thereof; wherein R1, R2, R3, R4, R5 and X are as defined in the detailed description.

In another embodiment, there is provided a pharmaceutical composition comprising a compound of Formula (I) or a pharmaceutically acceptable salt thereof, and a pharmaceutically-acceptable carrier.
In various embodiments, the pharmaceutical composition further comprises one or more additional pharmaceutically active compounds.
In yet another embodiment, there is provided a method for treating a condition comprising administering to a subject a therapeutically effective amount of a compound of Formula (I), wherein the condition to be treated includes, but is not limited to, autoimmune disorders, chronic inflammatory disorders, acute inflammatory disorders, auto-inflammatory disorders, pain, atherosclerosis, diabetes, fibrotic diseases, metabolic disorders, cancer, neoplasia, leukemia, lymphoma, and rheumatoid arthritis.
In various embodiments, the method comprises administering a combination of a compound of Formula (I) and at least one additional pharmaceutically active compound.
In yet another embodiment, there is provided a method for preparing a compound of Formula (I) or a pharmaceutically acceptable salt thereof.
In yet another embodiment, there is provided an intermediate useful in making a compound of Formula (I) or a pharmaceutically acceptable salt thereof.
The compounds above are more fully described in the detailed description that follows.


DETAILED DESCRIPTION
The following description is merely exemplary in nature and is not intended to limit the present disclosure, application, or uses.
A. DEFINITIONS
The use of generic terms in the description of the compounds are herein defined for clarity.
This specification uses the terms “substituent”, “radical”, “group”, “moiety”, and “fragment” interchangeably.
The term “hydrido” denotes a single —H atom (H) and may be used interchangeably with the symbol “H” or the term “hydrogen”.
If a substituent is described as being “optionally substituted,” the substituent may be either (1) not substituted or (2) substituted. If a substitutable position is not substituted, the default substituent is a hydrido radical.
As used herein, the singular forms “a” and “an” may include plural reference unless the context clearly dictates otherwise.
The term “alkyl”, either alone or within other terms such as “haloalkyl” and “alkylaryl”, refers to an acyclic alkyl radical containing 1, 2, 3, 4, 5, 6, 7, 8, 9 or 10 carbon atoms. In some embodiments, alkyl is a C1-C10 alkyl group or a C1-C6 alkyl group. Examples of alkyl groups include, but are not limited to, methyl, ethyl, propyl, isopropyl, butyl, isobutyl, t-butyl, pentyl, hexyl, heptyl, octyl, nonyl and decyl.
The term “haloalkyl” refers to an alkyl moiety substituted with one or more halo groups. Examples of haloalkyl groups include —CF3 and —CHF2.
The term “alkoxy” is RO— where R is alkyl as defined herein. Non-limiting examples of alkoxy groups include methoxy, ethoxy and propoxy. The terms alkyloxy and alkoxy may be used interchangeably.
The term “cyano” denotes a carbon radical having 3 of 4 covalent bonds shared by a nitrogen atom.
The term “aryl” refers to any monocyclic, bicyclic or tricyclic carbon ring of up to 6 atoms in each ring, wherein at least one ring is aromatic, or an aromatic ring system of 5 to 14 carbons atoms which includes a carbocyclic aromatic group fused with a 5- or 6-membered cycloalkyl group. Examples of aryl groups include, but are not limited to, phenyl, naphthyl, tetrahydronaphthyl and indanyl.
The term “arylalkyl” embraces an aryl-substituted alkyl radical and may be used interchangeably with the term “aralkyl”. Examples include benzyl, diphenylmethyl, triphenylmethyl, phenylethyl and diphenylethyl. The terms benzyl and phenylmethyl are interchangeable.
The term “aryloxy” is RO—, where R is aryl. “Arylthio” is RS—, where R is aryl.
The term “heteroaryl” refers to a monocyclic, bicyclic or tricyclic ring having up to 6 atoms in each ring, wherein at least one ring is aromatic and contains from 1 to 4 heteroatoms in the ring selected from the group consisting of N, O and S, Non-limiting examples of heteroaryl include pyridyl, thienyl, furanyl, pyrimidyl, imidazolyl, pyranyl, pyrazolyl, thiazolyl, thiadiazolyl, isothiazolyl, oxazolyl, isoxazoyl, pyrrolyl, pyridazinyl, pyrazinyl, quinolinyl, isoquinolinyl, benzofuranyl, dibenzofuranyl, dibenzothiophenyl, benzothienyl, indolyl, benzothiazolyl, benzooxazolyl, benzimidazolyl, isoindolyl, benzotriazolyl, purinyl, thianaphthenyl and pyrazinyl. Attachment of heteroaryl can occur via an aromatic ring, or, if heteroaryl is bicyclic or tricyclic and one of the rings is not aromatic or contains no heteroatoms, through a non-aromatic ring or a ring containing no heteroatoms. “Heteroaryl” is also understood to include the N-oxide derivative of any nitrogen containing heteroaryl.
The term “hydroxyalkyl” refers to a linear or branched monovalent C1-C10 hydrocarbon group substituted with at least one hydroxy group and examples of hydroxyalkyl groups include, but are not limited to, hydroxymethyl, hydroxyethyl, hydroxypropyl and hydroxybutyl.
The number of carbon atoms in a hydrocarbyl substituent can be indicated by the prefix “CX—CY” where X is the minimum and Y is the maximum number of carbon atoms in the substituent.
The term “pharmaceutically-acceptable” means suitable for use in pharmaceutical preparations, generally considered as safe for such use, officially approved by a regulatory agency of a national or state government for such use, or being listed in the U.S. Pharmacopoeia or other generally recognized pharmacopoeia for use in animals, and more particularly in humans.
The term “pharmaceutically-acceptable salt” refers to a salt which may enhance desired pharmacological activity. Examples of pharmaceutically-acceptable salts include acid addition salts formed with inorganic or organic acids, metal salts and amine salts. Examples of acid addition salts formed with inorganic acids include salts with hydrochloric acid, hydrobromic acid, sulfuric acid, nitric acid and phosphoric acid. Examples of acid addition salts formed with organic acids such as acetic acid, propionic acid, hexanoic acid, heptanoic acid, cyclopentanepropionic acid, glycolic acid, pyruvic acid, lactic acid, malonic acid, succinic acid, malic acid, maleic acid, fumaric acid, tartaric acid, citric acid, benzoic acid, o-(4-hydroxy-benzoyl)-benzoic acid, cinnamic acid, mandelic acid, methanesulfonic acid, ethanesulfonic acid, 1,2-ethanedisulfonic acid, 2-hydroxyethane-sulfonic acid, benzenesulfonic acid, p-chlorobenzenesulfonic acid, 2-naphthalenesulfonic acid, p-toluenesulfonic acid, camphorsulfonic acid, 4-methyl-bicyclo[2.2.2]oct-2-ene1-carboxylic acid, gluco-heptonic acid, 4,4′-methylenebis(3-hydroxy-2-naphthoic) acid, 3-phenylpropionic acid, trimethyl-acetic acid, tertiary butylacetic acid, lauryl sulfuric acid, gluconic acid, glutamic acid, hydroxy-naphthoic acids, salicylic acid, stearic acid and muconic acid. Examples of metal salts include salts with sodium, potassium, calcium, magnesium, aluminum, iron, and zinc ions. Examples of amine salts include salts with ammonia and organic nitrogenous bases strong enough to form salts with carboxylic acids.
The term “therapeutically-effective amount” refers to an amount of a compound that, when administered to a subject for treating a disease, is sufficient to effect treatment for the disease. “Therapeutically effective amount” can vary depending on the compound, the disease and its severity, the age, the weight, etc. of the subject to be treated.
Compounds of the present invention can exist in tautomeric, geometric or stereoisomeric forms. The compounds' corresponding esters, metabolites, oximes, prodrugs, oniums and N-oxides are also embraced by the invention. The present invention contemplates all such compounds, including cis- and trans-geometric isomers, E- and Z-geometric isomers, R- and S-enantiomers, diastereomers, atropisomers, d-isomers, 1-isomers, mixtures of isomers and racemates thereof, as falling within the scope of the invention.
The terms “cis” and “trans” denote a form of geometric isomerism in which two carbon atoms connected by a double bond will each have a radical atom on the same side of the double bond (“cis”) or on opposite sides of the double bond (“trans”).
Some of the compounds described contain one or more stereocenters and are meant to include R, S and mixtures of R and S forms for each stereocenter present.
The term “atropisomerism” refers to a type of isomerism resulting from hindered rotation around a single bond due to steric strain of the substituents. This phenomenon creates stereoisomers which display axial chirality.
The following scheme illustrates “atropisomerism” with reference to specific pyridinone-pyridine compounds of the invention:




The bond between the B and C rings of the title compounds is hindered and does not allow for facile rotation. The steric strain barrier to rotation is sufficiently high such that individual conformers can be isolated. The compounds of the invention may also exist as atropisomers, i.e., chiral rotational isomers. The invention encompasses racemates, resolved atropisomers, and mixtures thereof. Atropisomers may be separated via supercritical fluid chromatography using a mobile phase of carbon dioxide and ethanol/methanol.
Atropisomers are generally stable but can often be equilibrated thermally. Atropisomers will have the same but opposite optical rotation. Like chiral compounds each atropisomers may have different properties when bound to an enzyme or receptor with one isomer often being more potent than the other. Atropisomers are frequently used as pharmaceutical agents. Known examples include Vancomycin and derivatives.
The term “3,5-difluoropyridin-2-yl” refers to a moiety of structure:




The term “3-fluoropyridin-2-yl” refers to a moiety of structure




The term “5-fluoro-3-methylpyridin-2-yl” refers to a moiety of structure:




The term “6-fluoropyridin-2-yl” refers to a moiety of structure:




The term “6-fluoro-4-methylpyridin-2-yl” refers to a moiety of structure:




The term “3-fluoro-5-methylpyridin-2-yl” refers to a moiety of structure:




The term “5-fluoropyridin-2-yl” refers to a moiety of structure:




The term “4-fluoropyridin-3-yl” refers to a moiety of structure:




The term “5-fluoropyrimidin-4-yl” refers to a moiety of structure:




LIST OF ABBREVIATIONS
ACN acetonitrile
Boc tert-butyloxycarbonyl
Bu butyl
Bpy 2,2′-bipyridine
DCA dichloroacetic acid
DCI dicyclohexylcarbodiimide
DCM dichloromethane or methylenechloride
DIPEA diisopropylethylamine
DMA dimethylacetamide
DMAP 4-dimethylaminopyridine or N,N-dimethylaminopyridine
DME 1,2-dimethoxyethane
DMF N,N-dimethylformamide
DMSO dimethylsulfoxide
CuBr2 copper(II)bromide
EDAC N-(3-dimethylaminopropyl)-N′-ethylcarbodiimide hydrochloride
eq. equivalents
Et ethyl
EtOAC ethyl acetate
EtOH ethanol
HPLC high pressure liquid chromatography
h hour(s)
IPA isopropyl alcohol
K2CO3 potassium carbonate
KOtBu potassium tert-butoxide
LAH lithium aluminum hydride
LC/MS liquid chromatography mass spectrometry
LC/MS/MS liquid chromatography tandem mass spectrometry
mCPBA m-chloroperbenzoic acid
Me methyl
MeCN acetonitrile
MeOH methanol
MgSO4 magnesium sulfate
mL milliliter
mmol millimole
NaH sodium hydride
NaN(TMS)2 sodium bis(trimethylsilyl)amide
NCS n-chloro succinimide
NMR nuclear magnetic resonance
Pd/C palladium on carbon
Ph phenyl
PPA polyphosphoric acid
TEA triethylamine
TFA trifluoroacetic acid
THF tetrahydrofuran
TOSMIC toluenesulfonylmethyl isocyanide
TSA p-toluenesulfonic acid.
B. COMPOUNDS
The present disclosure provides a compound having the structure of Formula (I):




or a pharmaceutically acceptable salt thereof, wherein:

    
    
        X is CH or N;
        R1 is selected from the group consisting of H, C1-C6 alkyl, fluoro, chloro, bromo, cyano, or —CF3;
        R2 is selected from the group consisting of H, methyl, cyano, or fluoro;
        R3 is selected from the group consisting of:
    
    






    
    
        R4 is selected from the group consisting of H, methyl, OH, and —O—CH3;
        R5 is H or C1-C3 alkyl;
        m is 1 or 2;
        n is 0 or 1;
        p is 1; and
        q is 0 or 1.
    
    


In another embodiment, there is provided a compound having the structure of Formula (II):




or a pharmaceutically acceptable salt thereof, wherein:

    
    
        X is CH or N;
        R1 is selected from the group consisting of H, C1-C6 alkyl, fluoro, chloro, bromo, cyano, or —CF3;
        R2 is selected from the group consisting of H, methyl, cyano, or fluoro;
        R4 is selected from the group consisting of H, methyl, OH, and —OCH3;
        R5 is H or C1-C3 alkyl;
        m is 1 or 2; and
        n is 0 or 1.
    
    


Non-limiting examples of Formula (II) compounds include the following compounds and pharmaceutically acceptable salts thereof:












No.
Structure
Compound Name

















1




3-chloro-4-((5-fluoropyridin-2-yl)methoxy)-2′-(2- (2-hydroxypropan-2-yl)pyrimidin-4-yl)-5′,6- dimethyl-2H-[1,4′-bipyridin]-2-one


 


2




3-chloro-4-((3-fluoropyridin-2-yl)methoxy)-2′-(2- (2-hydroxypropan-2-yl)pyrimidin-4-yl)-5′,6- dimethyl-2H-[1,4′-bipyridin]-2-one


 


3




3-chloro-4-((5-fluoro-3-methylpyridin-2- yl)methoxy)-2′-(2-(2-hydroxypropan-2- yl)pyrimidin-4-yl)-5′,6-dimethyl-2H-[1,4′- bipyridin]-2-one


 


4




3-chloro-4-((6-fluoropyridin-2-yl)methoxy)-2′-(2- (2-hydroxypropan-2-yl)pyrimidin-4-yl)-5′,6- dimethyl-2H-[1,4′-bipyridin]-2-one


 


5




3-chloro-4-((6-fluoro-4-methylpyridin-2- yl)methoxy)-2′-(2-(2-hydroxypropan-2- yl)pyrimidin-4-yl)-5′,6-dimethyl-2H-[1,4′- bipyridin]-2-one


 


6




3-chloro-4-((3-fluoro-5-methylpyridin-2- yl)methoxy)-2′-(2-(2-hydroxypropan-2- yl)pyrimidin-4-yl)-5′,6-dimethyl-2H-[1,4′- bipyridin]-2-one


 


7




3-bromo-4-((5-fluoropyridin-2-yl)methoxy)-2′-(2- (2-hydroxypropan-2-yl)pyrimidin-4-yl)-5′,6- dimethyl-2H-[1,4′-bipyridin]-2-one


 


8




3-chloro-4-((5-fluoropyridin-2-yl)methoxy)-6′′- (2-hydroxypropan-2-yl)-5′,6-dimethyl-2H- [1,4′:2′,2′′-terpyridin]-2-one


 


9




3-bromo-4-((5-fluoropyridin-2-yl)methoxy)-6′′- (2-hydroxypropan-2-yl)-5′,6-dimethyl-2H- [1,4′:2′,2′′-terpyridin]-2-one


 


10




3-chloro-4-((5-fluoropyridin-2-yl)methoxy)-2′-(2- (2-hydroxypropan-2-yl)-5-methylpyrimidin-4-yl)- 5′,6-dimethyl-2H-[1,4′-bipyridin]-2-one


 


11




3-bromo-4-((5-fluoropyridin-2-yl)methoxy)-2′-(2- (2-hydroxypropan-2-yl)-5-methylpyrimidin-4-yl)- 5′,6-dimethyl-2H-[1,4′-bipyridin]-2-one


 


12




3-chloro-4-((5-fluoropyridin-2-yl)methoxy)-6′′- (2-hydroxypropan-2-yl)-3′′,5′,6-trimethyl-2H- [1,4′:2′,2′′-terpyridin]-2-one


 


13




3-bromo-4-((5-fluoropyridin-2-yl)methoxy)-6′′- (2-hydroxypropan-2-yl)-3′′,5′,6-trimethyl-2H- [1,4′:2′,2′′-terpyridin]-2-one


 


14




3-bromo-4-((3-fluoropyridin-2-yl)methoxy)-2′-(2- (2-hydroxypropan-2-yl)pyrimidin-4-yl)-5′,6- dimethyl-2H-[1,4′-bipyridin]-2-one


 


15




3-chloro-4-((3-fluoropyridin-2-yl)methoxy)-6′′- (2-hydroxypropan-2-yl)-5′,6-dimethyl-2H- [1,4′:2′,2′′-terpyridin]-2-one


 


16




3-bromo-4-((3-fluoropyridin-2-yl)methoxy)-6′′- (2-hydroxypropan-2-yl)-5′,6-dimethyl-2H- [1,4′:2′,2′′-terpyridin]-2-one


 


17




3-chloro-4-((3-fluoropyridin-2-yl)methoxy)-2′-(2- (2-hydroxypropan-2-yl)-5-methylpyrimidin-4-yl)- 5′,6-dimethyl-2H-[1,4′-bipyridin]-2-one


 


18




3-bromo-4-((3-fluoropyridin-2-yl)methoxy)-2′-(2- (2-hydroxypropan-2-yl)-5-methylpyrimidin-4-yl)- 5′,6-dimethyl-2H-[1,4′-bipyridin]-2-one


 


19




3-chloro-4-((3-fluoropyridin-2-yl)methoxy)-6′′- (2-hydroxypropan-2-yl)-3′′,5′,6-trimethyl-2H- [1,4′:2′,2′′-terpyridin]-2-one


 


20




3-bromo-4-((3-fluoropyridin-2-yl)methoxy)-6′′- (2-hydroxypropan-2-yl)-3′′,5′,6-trimethyl-2H- [1,4′:2′,2′′-terpyridin]-2-one


 


21




3-bromo-4-((5-fluoro-3-methylpyridin-2- yl)methoxy)-2′-(2-(2-hydroxypropan-2- yl)pyrimidin-4-yl)-5′,6-dimethyl-2H-[1,4′- bipyridin]-2-one


 


22




3-chloro-4-((5-fluoro-3-methylpyridin-2- yl)methoxy)-6′′-(2-hydroxypropan-2-yl)-5′,6- dimethyl-2H-[1,4′:2′,2′′-terpyridin]-2-one


 


23




3-bromo-4-((5-fluoro-3-methylpyridin-2- yl)methoxy)-6′′-(2-hydroxypropan-2-yl)-5′,6- dimethyl-2H-[1,4′:2′,2′′-terpyridin]-2-one


 


24




3-chloro-4-((5-fluoro-3-methylpyridin-2- yl)methoxy)-2′-(2-(2-hydroxypropan-2-yl)-5- methylpyrimidin-4-yl)-5′,6-dimethyl-2H-[1,4′- bipyridin]-2-one


 


25




3-bromo-4-((5-fluoro-3-methylpyridin-2- yl)methoxy)-2′-(2-(2-hydroxypropan-2-yl)-5- methylpyrimidin-4-yl)-5′,6-dimethyl-2H-[1,4′- bipyridin]-2-one


 


26




3-chloro-4-((5-fluoro-3-methylpyridin-2- yl)methoxy)-6′′-(2-hydroxypropan-2-yl)-3′′,5′,6- trimethyl-2H-[1,4′:2′,2′′-terpyridin]-2-one


 


27




3-bromo-4-((5-fluoro-3-methylpyridin-2- yl)methoxy)-6′′-(2-hydroxypropan-2-yl)-3′′,5′,6- trimethyl-2H-[1,4′:2′,2′′-terpyridin]-2-one


 


28




3-bromo-4-((6-fluoropyridin-2-yl)methoxy)-2′-(2- (2-hydroxypropan-2-yl)pyrimidin-4-yl)-5′,6- dimethyl-2H-[1,4′-bipyridin]-2-one


 


29




3-chloro-4-((6-fluoropyridin-2-yl)methoxy)-6′′- (2-hydroxypropan-2-yl)-5′,6-dimethyl-2H- [1,4′:2′,2′′-terpyridin]-2-one


 


30




3-bromo-4-((6-fluoropyridin-2-yl)methoxy)-6′′- (2-hydroxypropan-2-yl)-5′,6-dimethyl-2H- [1,4′:2′,2′′-terpyridin]-2-one


 


31




3-chloro-4-((6-fluoropyridin-2-yl)methoxy)-2′-(2- (2-hydroxypropan-2-yl)-5-methylpyrimidin-4-yl)- 5′,6-dimethyl-2H-[1,4′-bipyridin]-2-one


 


32




3-bromo-4-((6-fluoropyridin-2-yl)methoxy)-2′-(2- (2-hydroxypropan-2-yl)-5-methylpyrimidin-4-yl)- 5′,6-dimethyl-2H-[1,4′-bipyridin]-2-one


 


33




3-chloro-4-((6-fluoropyridin-2-yl)methoxy)-6′′- (2-hydroxypropan-2-yl)-3′′,5′,6-trimethyl-2H- [1,4′:2′,2′′-terpyridin]-2-one


 


34




3-bromo-4-((6-fluoropyridin-2-yl)methoxy)-6′′- (2-hydroxypropan-2-yl)-3′′,5′,6-trimethyl-2H- [1,4′:2′,2′′-terpyridin]-2-one


 


35




3-bromo-4-((6-fluoro-4-methylpyridin-2- yl)methoxy)-2′-(2-(2-hydroxypropan-2- yl)pyrimidin-4-yl)-5′,6-dimethyl-2H-[1,4′- bipyridin]-2-one


 


36




3-chloro-4-((6-fluoro-4-methylpyridin-2- yl)methoxy)-6′′-(2-hydroxypropan-2-yl)-5′,6- dimethyl-2H-[1,4′:2′,2′′-terpyridin]-2-one


 


37




3-bromo-4-((6-fluoro-4-methylpyridin-2- yl)methoxy)-6′′-(2-hydroxypropan-2-yl)-5′,6- dimethyl-2H-[1,4′:2′,2′′-terpyridin]-2-one


 


38




3-chloro-4-((6-fluoro-4-methylpyridin-2- yl)methoxy)-2′-(2-(2-hydroxypropan-2-yl)-5- methylpyrimidin-4-yl)-5′,6-dimethyl-2H-[1,4′- bipyridin]-2-one


 


39




3-bromo-4-((6-fluoro-4-methylpyridin-2- yl)methoxy)-2′-(2-(2-hydroxypropan-2-yl)-5- methylpyrimidin-4-yl)-5′,6-dimethyl-2H-[1,4′- bipyridin]-2-one


 


40




3-chloro-4-((6-fluoro-4-methylpyridin-2- yl)methoxy)-6′′-(2-hydroxypropan-2-yl)-3′′,5′,6- trimethyl-2H-[1,4′:2′,2′′-terpyridin]-2-one


 


41




3-bromo-4-((6-fluoro-4-methylpyridin-2- yl)methoxy)-6′′-(2-hydroxypropan-2-yl)-3′′,5′,6- trimethyl-2H-[1,4′:2′,2′′-terpyridin]-2-one


 


42




3-bromo-4-((3-fluoro-5-methylpyridin-2- yl)methoxy)-2′-(2-(2-hydroxypropan-2- yl)pyrimidin-4-yl)-5′,6-dimethyl-2H-[1,4′- bipyridin]-2-one


 


43




3-chloro-4-((3-fluoro-5-methylpyridin-2- yl)methoxy)-6′′-(2-hydroxypropan-2-yl)-5′,6- dimethyl-2H-[1,4′:2′,2′′-terpyridin]-2-one


 


44




3-bromo-4-((3-fluoro-5-methylpyridin-2- yl)methoxy)-6′′-(2-hydroxypropan-2-yl)-5′,6- dimethyl-2H-[1,4′:2′,2′′-terpyridin]-2-one


 


45




3-chloro-4-((3-fluoro-5-methylpyridin-2- yl)methoxy)-2′-(2-(2-hydroxypropan-2-yl)-5- methylpyrimidin-4-yl)-5′,6-dimethyl-2H-[1,4′- bipyridin]-2-one


 


46




3-bromo-4-((3-fluoro-5-methylpyridin-2- yl)methoxy)-2′-(2-(2-hydroxypropan-2-yl)-5- methylpyrimidin-4-yl)-5′,6-dimethyl-2H-[1,4′- bipyridin]-2-one


 


47




3-chloro-4-((3-fluoro-5-methylpyridin-2- yl)methoxy)-6′′-(2-hydroxypropan-2-yl)-3′′,5′,6- trimethyl-2H-[1,4′:2′,2′′-terpyridin]-2-one


 


48




3-bromo-4-((3-fluoro-5-methylpyridin-2- yl)methoxy)-6′′-(2-hydroxypropan-2-yl)-3′′,5′,6- trimethyl-2H-[1,4′:2′,2′′-terpyridin]-2-one









In another embodiment, there is provided a compound having the structure of Formula (III):




or a pharmaceutically acceptable salt thereof, wherein:

    
    
        X is CH or N;
        R1 is chloro or bromo; and
        R2 is —H or methyl.
    
    


Non-limiting examples of Formula (III) compounds include the following compounds and pharmaceutically acceptable salts thereof:












No.
Structure
Compound Name







49




3-chloro-4-((3,5-difluoropyridin-2-yl)methoxy)- 2′-(2-(2-hydroxypropan-2-yl)pyrimidin-4-yl)- 5′,6-dimethyl-2H-[1,4′-bipyridin]-2-one


 


50




3-bromo-4-((3,5-difluoropyridin-2-yl)methoxy)- 2′-(2-(2-hydroxypropan-2-yl)pyrimidin-4-yl)- 5′,6-dimethyl-2H-[1,4′-bipyridin]-2-one


 


51




3-chloro-4-((3,5-difluoropyridin-2-yl)methoxy)- 6′′-(2-hydroxypropan-2-yl)-5′,6-dimethyl-2H- [1,4′:2′,2′′-terpyridin]-2-one


 


52




3-bromo-4-((3,5-difluoropyridin-2-yl)methoxy)- 6′′-(2-hydroxypropan-2-yl)-5′,6-dimethyl-2H- [1,4′:2′,2′′-terpyridin]-2-one


 


53




3-chloro-4-((3,5-difluoropyridin-2-yl)methoxy)- 2′-(2-(2-hydroxypropan-2-yl)-5- methylpyrimidin-4-yl)-5′,6-dimethyl-2H-[1,4′- bipyridin]-2-one


 


54




3-bromo-4-((3,5-difluoropyridin-2-yl)methoxy)- 2′-(2-(2-hydroxypropan-2-yl)-5- methylpyrimidin-4-yl)-5′,6-dimethyl-2H-[1,4′- bipyridin]-2-one


 


55




3-chloro-4-((3,5-difluoropyridin-2-yl)methoxy)- 6′′-(2-hydroxypropan-2-yl)-3′′,5′,6-trimethyl-2H- [1,4′:2′,2′′-terpyridin]-2-one


 


56




3-bromo-4-((3,5-difluoropyridin-2-yl)methoxy)- 6′′-(2-hydroxypropan-2-yl)-3′′,5′,6-trimethyl-2H- [1,4′:2′,2′′-terpyridin]-2-one









In another embodiment, there is provided a compound having the structure of Formula (IV):




or a pharmaceutically acceptable salt thereof, wherein:

    
    
        X is CH or N;
        R1 is selected from the group consisting of H, C1-C6 alkyl, fluoro, chloro, bromo, cyano, or —CF3;
        R2 is selected from the group consisting of H, methyl, cyano, or fluoro;
        R4 is selected from the group consisting of H, methyl, OH, and —OCH3;
        R5 is H or C1-C3 alkyl;
        m is 1 or 2; and
        n is 0 or 1.
    
    


In another embodiment, there is provided a compound having the structure of Formula (V):




or a pharmaceutically acceptable salt thereof, wherein:

    
    
        X is CH or N;
        R1 is chloro or bromo; and
        R2 is —H or methyl.
    
    


Non-limiting examples of Formula (V) compounds include the following compounds and pharmaceutically acceptable salts thereof:












No.
Structure
Compound Name







57




3-chloro-4-((4-fluoropyridin-3-yl)methoxy)-2′- (2-(2-hydroxypropan-2-yl)pyrimidin-4-yl)-5′,6- dimethyl-2H-[1,4′-bipyridin]-2-one


 


58




3-bromo-4-((4-fluoropyridin-3-yl)methoxy)-2′- (2-(2-hydroxypropan-2-yl)pyrimidin-4-yl)-5′,6- dimethyl-2H-[1,4′-bipyridin]-2-one


 


59




3-chloro-4-((4-fluoropyridin-3-yl)methoxy)-6′′- (2-hydroxypropan-2-yl)-5′,6-dimethyl-2H- [1,4′:2′,2′′-terpyridin]-2-one


 


60




3-bromo-4-((4-fluoropyridin-3-yl)methoxy)-6′′- (2-hydroxypropan-2-yl)-5′,6-dimethyl-2H- [1,4′:2′,2′′-terpyridin]-2-one


 


61




3-chloro-4-((4-fluoropyridin-3-yl)methoxy)-2′- (2-(2-hydroxypropan-2-yl)-5-methylpyrimidin-4- yl)-5′,6-dimethyl-2H-[1,4′-bipyridin]-2-one


 


62




3-bromo-4-((4-fluoropyridin-3-yl)methoxy)-2′- (2-(2-hydroxypropan-2-yl)-5-methylpyrimidin-4- yl)-5′,6-dimethyl-2H-[1,4′-bipyridin]-2-one


 


63




3-chloro-4-((4-fluoropyridin-3-yl)methoxy)-6′′- (2-hydroxypropan-2-yl)-3′′,5′,6-trimethyl-2H- [1,4′:2′,2′′-terpyridin]-2-one


 


64




3-bromo-4-((4-fluoropyridin-3-yl)methoxy)-6′′- (2-hydroxypropan-2-yl)-3′′,5′,6-trimethyl-2H- [1,4′:2′,2′′-terpyridin]-2-one









In another embodiment, there is provided a compound having the structure of Formula (VI):




or a pharmaceutically acceptable salt thereof, wherein:

    
    
        X is CH or N;
        R1 is selected from the group consisting of H, C1-C6 alkyl, fluoro, chloro, bromo, cyano, or —CF3;
        R2 is selected from the group consisting of H, methyl, cyano, or fluoro;
        R4 is selected from the group consisting of H, methyl, OH, and —OCH3;
        R5 is H or C1-C3 alkyl;
        p is 1; and
        q is 0 or 1.
    
    


In another embodiment, there is provided a compound having the structure of Formula (VII):




or a pharmaceutically acceptable salt thereof, wherein:

    
    
        X is CH or N;
        R1 is chloro or bromo; and
        R2 is —H or methyl.
    
    


Non-limiting examples of Formula (VII) compounds include the following compounds and pharmaceutically acceptable salts thereof:












No.
Structure
Compound Name







65




3-chloro-4-((5-fluoropyrinindin-4-yl)methoxy)-2′- (2-(2-hydroxypropan-2-yl)pyrimidin-4-yl)-5′,6- dimethyl-2H-[1,4′-bipyridin]-2-one


 


66




3-bromo-4-((5-fluoropyrimidin-4-yl)methoxy)- 2′-(2-(2-hydroxypropan-2-yl)pyrimidin-4-yl)- 5′,6-dimethyl-2H-[1,4′-bipyridin]-2-one


 


67




3-chloro-4-((5-fluoropyrimidin-4-yl)methoxy)- 6′′-(2-hydroxypropan-2-yl)-5′,6-dimethyl-2H- [1,4′:2′,2′′-terpyridin]-2-one


 


68




3-bromo-4-((5-fluoropyrimidin-4-yl)methoxy)- 6′′-(2-hydroxypropan-2-yl)-5′,6-dimethyl-2H- [1,4′:2′,2′′-terpyridin]-2-one


 


69




3-chloro-4-((5-fluoropyrimidin-4-yl)methoxy)-2′- (2-(2-hydroxypropan-2-yl)-5-methylpyrimidin-4- yl)-5′,6-dimethyl-2H-[1,4′-bipyridin]-2-one


 


70




3-bromo-4-((5-fluoropyrimidin-4-yl)methoxy)- 2′-(2-(2-hydroxypropan-2-yl)-5- methylpyrimidin-4-yl)-5′,6-dimethyl-2H-[1,4′- bipyridin]-2-one


 


71




3-chloro-4-((5-fluoropyrimidin-4-yl)methoxy)- 6′′-(2-hydroxypropan-2-yl)-3′′,5′,6-trimethyl-2H- [1,4′:2′,2′′-terpyridin]-2-one


 


72




3-bromo-4-((5-fluoropyrimidin-4-yl)methoxy)- 6′′-(2-hydroxypropan-2-yl)-3′′,5′,6-trimethyl-2H- [1,4′:2′,2′′-terpyridin]-2-one









In another embodiment, there is provided a method to make a compound of Formula W comprising:

    
    
        contacting a compound of Formula Y with Compound Z in a polar solvent to form a mixture;
        heating the mixture at a temperature and for a time sufficient; and
        in the presence of an acid having a pKa less than about 2, forming a reaction product having the structure of Formula W:
    
    






wherein R10 is either bromo or chloro. In another embodiment, the polar solvent is dioxane. In another embodiment, the temperature is in a range from about 70° C. to about 105° C.
In another embodiment, the time sufficient to form a reaction product is in the range of two to about six hours. In another embodiment, the acid is sulfuric acid. In another embodiment, R10 is chloro. In another embodiment, R10 is bromo.
In another embodiment, there is provided a pharmaceutical composition comprising a therapeutically effective amount of a compound of Formula I or a pharmaceutically acceptable salt thereof, and a pharmaceutically acceptable carrier.
In another embodiment, there is provided a pharmaceutical composition comprising a therapeutically effective amount of a compound of Formula I or a pharmaceutically acceptable salt thereof, a pharmaceutically acceptable carrier, and a therapeutically effective amount of an active pharmaceutical ingredient selected from the group consisting of anti-inflammatory drugs, anti-atherosclerotic drugs, immunosuppressive drugs, immunomodulatory drugs, cytostatic drugs, angiogenesis inhibitors, kinase inhibitors, cytokine blockers and inhibitors of cell adhesion molecules.
In another embodiment, there is provided a method for treating a condition comprising administering to a subject in need thereof a therapeutically effective amount of a compound of Formula I or pharmaceutically acceptable salt thereof, wherein the condition is selected from the group consisting of autoimmune disorders, chronic inflammatory disorders, acute inflammatory disorders, auto-inflammatory disorders, atherosclerosis, diabetes, fibrotic diseases, metabolic disorders, cancer, neoplasia, leukemia and lymphoma. In another embodiment, the subject is a mammal selected from a canine and a human. In another embodiment, the condition is lymphoma. In another embodiment, the condition is rheumatoid arthritis.
C. GENERAL SYNTHETIC SCHEMES
The compounds of the present invention can be prepared using the methods illustrated in the general synthetic schemes and experimental procedures detailed below. These general synthetic schemes and experimental procedures are presented for purposes of illustration and are not intended to be limiting. The starting materials used to prepare the compounds of the present invention are commercially available or can be prepared using routine methods known in the art.
Representative procedures for the preparation of compounds of invention are outlined in Schemes 1 and 2. The substituted pyridine starting material can be purchased or prepared using methods known in the art with a representative procedure provided as an intermediate. Scheme 1 highlights the synthesis of the fully elaborated 1,4′-bipyridin-2-ones. The synthesis of pyridinone 1c can be accomplished by reaction of acetal 1a and pyridine 1b in a solvent such as dioxane. Alkylation of the phenol of 1c with the desired heteroaryl substituent (R3) gives alkylated 1d. Pyridinone 1d may be converted to the title compound via one of three routes depending on the R2 and X-substituents. For instance if R2 is methyl, reaction of 1d with a vinyl tin reagent in the presence of a palladium catalyst provides methyl ketone 1i. Halogenation of 1i using N-chlorosuccinimide (or N-bromosuccinimide if the corresponding bromo is desired) in a solvent such as isopropanol provides 1j. In situ enamine formation by reaction of 1j with N,N-dimethylformamide dimethyl acetal provides an intermediate, which is then reacted with 2-hydroxy-2-methylpropionamidine in a solvent such as DMF to give pyridinone 1g. Alternatively, if X is N 1d may be carboxylated by treating the halide with carbon monoxide in the presence of a palladium catalyst in ethanol to give ester 1e. Hydrolysis of the ester of 1e with lithium hydroxide in water followed by treating the intermediate carboxylic acid with CDI, and subsequently with methoxymethylamine and an amine base such as diisopropylethylamine under Weinreb conditions gives 1 h. Reaction of the Weinreb amide 1 h with the desired R2 Grinard reagent in a solvent such as THF provides 1i. Ketone 1i is then coverted to 1j and then the final compound 1g, as indicated above. Another option to set the pyridine or pyrimidine D-ring is to react 1d with the desired boronic acid under Suzuki conditions using an appropriate palladium catalyst to give the coupled intermediate which is then halogenated using NCS or NBS to provide 1f. Addition of methyl magnesium bromide to 1f in a solvent such as THF provides the title compound 1g.




The synthesis of the desired compounds wherein the benzyl substituent R3 is added in the last step is shown in Scheme 2. Pyridinone 2c can be accomplished by reaction of acetal 2a and pyridine 2b in a solvent such as dioxane as described in Scheme 1. Protection of the phenol of 2c with para-methoxybenzyl bromide gives benzylated 2d. Reaction of 2d with a vinyl tin reagent in the presence of a palladium catalyst provides methyl ketone 2e. Halogenation of 1e using N-chlorosuccinimide (or N-bromosuccinimide if the corresponding bromo is desired) in a solvent such as isopropanol provides 2f. In situ enamine formation by reaction of 2f with N,N-dimethylformamide dimethyl acetal provides an intermediate, which is then reacted with 2-hydroxy-2-methylpropionamidine in a solvent such as DMF to give pyrimidinone 2g. Deprotection of the benzyl group by treating 2g with an acid such as TFA or HCl provides 2h. Alkylation of phenol 2h with the desired benzyl halide substituent (R3CH2Br or R3CH2Cl) provides the desired pyridinones 2i.







D. SPECIES COMPOUNDS PREPARATIONS
Intermediates
Intermediate I: Preparation of 2-chloromethyl-3,5-difluoro-pyridine




Step A: Preparation of 3,5-difluoro-pyridine-2-carboxylic acid ethyl ester




To a suspension of 3,5-difluoropyridine-2-carboxylic acid (2.0 g, 12.6 mmol) in ethanol (5 mL), cooled using an ice water bath, was added thionyl chloride (2 mL) in a dropwise manner. The solution was heated at 60° C. for 3 h. The reaction was returned to ambient temperature and was concentrated in vacuo to provide the ethyl ester, hydrochloride salt as a yellow oil (2.5 g).
Step B: Preparation of (3,5-difluoro-pyridin-2-yl)-methanol




To a solution of 3,5-difluoro-pyridine-2-carboxylic acid ethyl ester of part A (2.5 g, 12.6 mmol) in ethanol (10 mL), cooled using an ice water bath, was added sodium borohydride (1.43 g, 37.8 mmol) in a portion wise manner. The solution was stirred at 0° C. for thirty minutes and at ambient temperature for 2 h. The reaction was returned to 0° C. and saturated ammonium chloride was added dropwise. The solvent was removed in vacuo and the resulting residue was partitioned between ethyl acetate and water. The organic layer was washed with saturated ammonium chloride, water and brine, and dried over magnesium sulfate. The slurry was filtered and concentrated to provide the alcohol as a yellow oil (1.8 g): MS (ES) m/e 146 (M+H).
Step C: Preparation of 2-chloromethyl-3,5-difluoro-pyridine




To a solution of (3,5-difluoro-pyridin-2-yl)-methanol from part B (1.8 g, 12.3 mmol) in dichloromethane (20 mL) was added three drops of N,N-dimethylformamide and cooled using an ice water bath. Thionyl chloride (2 mL) was added dropwise and the solution was stirred at ambient temperature for one hour. The solution was concentrated in vacuo to provide the chloro compound as a light brown liquid (1.75 g).
Compound No. 49, Example A
Preparation of 3-chloro-4-((3,5-difluoropyridin-2-yl)methoxy)-2′-(2-(2-hydroxypropan-2-yl)pyrimidin-4-yl)-5′,6-dimethyl-2H-[1,4′-bipyridin]-2-one




Step A: Preparation of 2′-chloro-4-hydroxy-6,5′-dimethyl-[1,4′]bipyridinyl-2-one




To a screw top vial with rubber septa inset was added 2,2-dimethyl-6-(2-oxo-propyl)-[1,3]dioxin-4-one, prepared as described in Organic Letters, 11(21), 4910-4913; 2009, (500 mg, 2.7 mmol) and 2-chloro-5-methyl-pyridin-4-ylamine (575 mg, 4 mmol, 1.5 eq). The mixture was dissolved in anhydrous 1,4-dioxane (10 mL). Once the mixture was homogeneous the vial was placed on a stirrer/hot plate preset to 90° C. The reaction vessel was heated at this temperature for 3.5 h. The reaction vial was removed from heat and analyzed by HPLC which showed that the reaction was >95% complete. The vial was placed back on the hot plate. To the heated mixture was added H2SO4 (250 μL) and the reaction was heated for 1 h. The reaction vial was removed from the heat and after cooling to ambient temperature, the dioxane was removed by passing a stream of air over the top of the open vial to give a brown residue. Water ( ̃4 mL) was added to the vial, and the mixture was stirred for 30 min. The resulting tan solid was filtered off with washing from additional water and the diethyl ether to give the desired product (531 mg, 57% based on being the sulfate salt) as a tan solid which by HPLC was  ̃95% pure: MS (ES) m/e 250 (M+H).
Step B: Preparation of 2′-chloro-4-((4-methoxybenzyl)oxy)-5′,6-dimethyl-2H-[1,4′-bipyridin]-2-one




To a solution of 2′-chloro-4-hydroxy-6,5′-dimethyl-[1,4′]bipyridinyl-2-one of part A (6.0 g, 20.1 mmol) in N,N-dimethylformamide (20 mL) was added 4-methoxybenzylchloride (2.73 mL, 20.1 mmol), potassium carbonate (6.93 g, 50.2 mmol) and 18-crown-6 (100 mg). The slurry was heated at 60° C. for 3 h and was stirred at ambient temperature for 18 h. The reaction mixture was partitioned between ethyl acetate and water. The organic layer was washed with water and brine and dried over magnesium sulfate. The solution was concentrated in vacuo to provide a brown oil. Normal phase chromatography (ethyl acetate/heptane) provided the alkylated product as a light yellow solid (4.6 g): MS (ES) m/e 371 (M+H).
Step C: Preparation of 2′-acetyl-4-((4-methoxybenzyl)oxy)-5′,6-dimethyl-2H-[1,4′-bipyridin]-2-one




A solution of 2′-chloro-4-((4-methoxybenzyl)oxy)-5′,6-dimethyl-2H-[1,4′-bipyridin]-2-one of part B (4.6 g, 12.4 mmol), tributyl(1-ethoxyvinyl)tin (4.6 mL, 13.6 mmol) and PdCl2(PPh3)2 (87 mg, 0.12 mmol) in 1,4-dioxane (30 mL) was irradiated using a CEM ExplorerTM microwave at 130° C. for 2 h. The resulting dark solution was filtered through Celite, rinsing with ethyl acetate. The filtrate was concentrated and the residue was dissolved into tetrahydrofuran (5 mL) and treated with concentrated HCl until hydrolysis was complete. The solution was concentrated in vacuo and purified using normal phase chromatography (ethyl acetate/heptane) to provide the acetyl compound as a yellow oil (3.3 g): MS (ES) m/e 379 (M+H).
Step D: Preparation of 2′-acetyl-3-chloro-4-hydroxy-5′-methyl-[1,4]bipyridinyl-2-one




To a solution of 2′-acetyl-4-((4-methoxybenzyl)oxy)-5′,6-dimethyl-2H-[1,4′-bipyridin]-2-one of part C (3.3 g, 8.7 mmol) in 2-propanol (100 mL) was added N-chlorosuccinimide (1.27 g, 9.6 mmol) and 10 drops of dichloroacetic acid. The slurry was heated at 60° C. for 3 h. The resulting slurry was concentrated and the residue was partitioned between ethyl acetate and water. The organic layer was washed with water and brine and dried over magnesium sulfate. The solution was concentrated in vacuo. The residue was suspended into dichloromethane and the resulting white solid was collected by vacuum filtration to provide the chlorinated deprotected product (1.16 g): MS (ES) m/e 293 (M+H).
Step E: Preparation of 2′-acetyl-3-chloro-4-(3,5-difluoro-pyridin-2-ylmethoxy)-5′-methyl-[1,4]bipyridinyl-2-one




To a solution of 2′-acetyl-3-chloro-4-hydroxy-5′-methyl-[1,4′]bipyridinyl-2-one of part D (500 mg, 1.7 mmol) in N,N-dimethylformamide (3 mL) was added 2-chloromethyl-3,5-difluoro-pyridine (277 mg, 1.7 mmol), potassium carbonate (590 mg, 4.28 mmol) and 18-crown-6 (10 mg) and the reaction was stirred at 60° C. for 4 h. After cooling the solution was partitioned between ethyl acetate and water. The organic layer was washed with water and brine and dried over magnesium sulfate. The solution was filtered and concentrated in vacuo. The crude material was purified using normal phase chromatography (ethyl acetate/heptane) to provide alkylated product as a yellow solid (397 mg): MS (ES) m/e 420 (M+H).
Step F: Preparation of 3-chloro-4-((3,5-difluoropyridin-2-yl)methoxy)-2′-(2-(2-hydroxypropan-2-yl)pyrimidin-4-yl)-5′,6-dimethyl-2H-[1,4′-bipyridin]-2-one




To a solution of 2′-acetyl-3-chloro-4-(3,5-difluoro-pyridin-2-ylmethoxy)-5′-methyl-[1,4′]bipyridinyl-2-one from step E (397 mg, 0.95 mmol) in N,N-dimethylformamide (3 mL) was added N,N-dimethylformamide dimethyl acetal (0.18 mL, 1.42 mmol) and the solution was heated to 55° C. for 18 h. The solution was concentrated to half volume and 2-hydroxy-2-methylpropionamidine HCl (195 mg, 1.42 mmol) and potassium carbonate (393 mg, 2.85 mmol) were added. The slurry was heated at 75° C. for 18 h. The slurry was returned to ambient temperature and partitioned between ethyl acetate and water. The organic layer was washed with brine and dried over magnesium sulfate. The solution was concentrated and purified using normal phase chromatography (ethyl acetate/heptane) to provide the title compound as a light yellow solid (255 mg, 46%): MS (ES) m/e 514 (M+H).
Chiral resolution of 3-chloro-4-((3,5-difluoropyridin-2-yl)methoxy)-2′-(2-(2-hydroxypropan-2-yl)pyrimidin-4-yl)-5′,6-dimethyl-2H-[1,4′-bipyridin]-2-one
Racemic 3-chloro-4-((3,5-difluoropyridin-2-yl)methoxy)-2′-(2-(2-hydroxypropan-2-yl)pyrimidin-4-yl)-5′,6-dimethyl-2H-[1,4′-bipyridin]-2-one (250 mg, 0.49 mmol) was separated using supercritical fluid chromatography (Thar 80, preparative SFC, ChiralCel OD-H, 250×30 mmID column) with a mobile phase of carbon dioxide and ethanol. The separation method used an isocratic method of 40% ethanol with a flow rate of 50 mL/min and a cycle time of 10 min. Optical rotation was determined using a WZZ-2S polarimeter.
The faster isomer eluted at 1.77 minutes yielded 115 mg of atropisomer 1: [α]D20−60.7° (CH3OH); 1H NMR (400 MHz, DMSO-d6) δ ppm 8.97 (d, J=5.09 Hz, 1H), 8.86 (s, 1H), 8.69 (s, 1H), 8.61 (s, 1H), 8.24 (d, J=5.08 Hz, 1H), 8.10 (t, 1H), 6.85 (s, 1H), 5.50 (s, 2H), 5.26 (s, 1H), 2.11 (s, 3H), 1.98 (s, 3H), 1.54 (s, 3H), 1.52 (s, 3H); MS (ES) m/e 514 (M+H).
The slower isomer eluted at 3.68 minutes yielded 112 mg of atropisomer 2: [α]D20+61.9° (CH3OH); 1H NMR (400 MHz, DMSO-d6) δ ppm 8.97 (d, J=5.09 Hz, 1H), 8.86 (s, 1H), 8.69 (s, 1H), 8.61 (s, 1H), 8.24 (d, J=5.08 Hz, 1H), 8.10 (t, 1H), 6.85 (s, 1H), 5.50 (s, 2H), 5.26 (s, 1H), 2.11 (s, 3H), 1.98 (s, 3H), 1.54 (s, 3H), 1.52 (s, 3H); MS (ES) m/e 514 (M+H).
Compound No. 65, Example B
Preparation of 3-chloro-4-((5-fluoropyrimidin-4-yl)methoxy)-2′-(2-(2-hydroxypropan-2-yl)pyrimidin-4-yl)-5′,6-dimethyl-2H-[1,4′-bipyridin]-2-one




The title compound was prepared following Compound 49, Example A, up to Step E, but alkylating 3-chloro-4-hydroxy-2′-(2-(2-hydroxypropan-2-yl)pyrimidin-4-yl)-5′,6-dimethyl-2H-[1,4′-bipyridin]-2-one with 4-(chloromethyl)-5-fluoropyrimidine instead of 2-(chloromethyl)-3,5-difluoropyridine to give the desired benzyl ether. The title compound was prepared following the general procedure of Compound 49, Example A, Step F.
E. METHODS OF TREATMENT
The present disclosure further provides methods for treating a condition in a subject having or susceptible to having such a condition, by administering to the subject a therapeutically-effective amount of one or more compounds as described above. In one embodiment, the treatment is preventative treatment. In another embodiment, the treatment is palliative treatment. In another embodiment, the treatment is restorative treatment.
1. Conditions
The conditions that can be treated in accordance with the present invention include, but are not limited to, autoimmune disorders, chronic inflammatory disorders, acute inflammatory disorders, auto-inflammatory disorders, pain, atherosclerosis, diabetes, fibrotic diseases, metabolic disorders, cancer, neoplasia, leukemia, lymphoma and the like.
In some embodiments the methods described herein are used to treat patients with disorders arising from dysregulated cytokine, enzymes and/or inflammatory mediator production, stability, secretion, posttranslational processing. Examples of cytokines that may be dysregulated include interleukins 1, 2, 6, 8, 10, 12, 17, 22 and 23 along with tumor necrosis factor alpha and interferons alpha, beta and gamma. Examples of inflammatory mediators that may be dysregulated include nitric oxide, prostaglandins and leukotrienes. Examples of enzymes include cyclo-oxygenase, nitric oxide synthase and matrixmetalloprotease.
In some embodiments the methods described herein are used to treat patients with dysregulated p38 activity, activation, biosynthesis or pathway function.
In some embodiments, the methods described herein are used to treat a patient in need thereof suffering from an autoimmune disorder, chronic and/or acute inflammatory disorder and/or auto-inflammatory disorder. Examples of disorders include, but are not limited to colitis, multiple sclerosis, arthritis, rheumatoid arthritis, osteoarthritis, juvenile arthritis, psoriatic arthritis, cryopyrin associated periodic syndromes, Muckle-Wells Syndrome, Familial Cold Auto-inflammatory Syndrome, neonatal-onset multisystem inflammatory disease, TNF receptor associated periodic syndrome, acute pancreatitis, chronic pancreatitis, atherosclerosis, inflammatory bowel disease, Crohn's disease, ulcerative colitis, Diabetes mellitus type 1, Diabetes mellitus type 2, diabetic retinopathy, Still's disease, multiple sclerosis, vasculitis, sarcoidosis, pulmonary inflammation, acute respiratory distress syndrome, wet and dry age-related macular degeneration, autoimmune hemolytic syndromes, autoimmune hepatitis, autoimmune neuropathy, autoimmune ovarian failure, autoimmune orchitis, autoimmune thrombocytopenia, reactive arthritis, ankylosing spondylitis, silicone implant associated autoimmune disease, Sjogren's syndrome, Familial Mediterranean Fever, systemic lupus erythematosus, vasculitis syndromes (such as, for example, giant cell arteritis, Behcet's disease & Wegener's granulomatosis), Vitiligo, secondary hematologic manifestation of autoimmune diseases (such as, for example, anemias), drug-induced autoimmunity, Hashimoto's thyroiditis, hypophysitis, idiopathic thrombocytic pupura, metal-induced autoimmunity, myasthenia gravis, pemphigus, autoimmune deafness (including, for example, Meniere's disease), Goodpasture's syndrome, Graves' disease, HW-related autoimmune syndromes and Gullain-Barre disease; Examples of inflammatory conditions include, but are not limited to sepsis, septic shock, endotoxic shock, exotoxin-induced toxic shock, gram negative sepsis, toxic shock syndrome, glomerulonephritis, peritonitis, interstitial cystitis, psoriasis, atopic dermatitis, hyperoxia-induced inflammations, asthma, chronic obstructive pulmonary disease (COPD), vasculitis, graft vs. host reaction (i.e., graft vs. host disease), allograft rejections (e.g., acute allograft rejection, and chronic allograft rejection), early transplantation rejection (e.g., acute allograft rejection), reperfusion injury, acute pain, chronic pain, neuropathic pain, Fibromyalgia, pancreatitis, chronic infections, meningitis, encephalitis, myocarditis, gingivitis, post-surgical trauma, tissue injury, traumatic brain injury, hepatitis, enterocolitis, sinusitis, uveitis, ocular inflammation, optic neuritis, gastric ulcers, esophagitis, peritonitis, periodontitis, dermatomyositis, gastritis, myositis, polymyalgia, pneumonia and bronchitis. Fibrotic diseases; Metabolic disorders, including but not limited obesity, steroid-resistance, glucose intolerance, metabolic syndrome. In some embodiments, the methods described herein can be used to treat a patient in need thereof and suffering from neoplasia. Examples of these conditions include but not limited to angiogenesis, multiple myeloma, leukemia, B cell lymphoma, T cell lymphoma, mast cell tumors, lymphoma, Hodgkin's disease, cancer of the bone, mouth/pharynx, oesophagus, larynx, stomach, intestine, colon, rectum, lung, liver, pancreas, nerve, brain, head and neck, throat, ovary, uterus, prostate, testis, bladder, kidney, breast non-small cell lung carcinoma, melanoma, skin cancer, teratoma, rhabdomyosarcoma, glioma, metastatic and bone disorders. In some embodiments, the disease associated with dysregulated p38 include Cardiovascular and Cerebrovascular diseases, including but not limited to atherosclerosis, restenosis of an atherosclerotic coronary artery, Acute coronary syndrome, myocardial infarction, cardiac-allograft vasculopathy and stroke; central nervous system disorders with an inflammatory or apoptotic component, Alzheimer's disease, Parkinson's disease, Huntington's disease, amyotrophic lateral sclerosis, spinal cord injury, neuronal ischemia and peripheral neuropathy. The term patient refers to both humans and nonhuman animals with the abovementioned conditions. Nonhuman animals could be companion animals such as, but not limited to canine and feline species.
2. Subjects
Suitable subjects to be treated according to the present invention include mammalian subjects. Mammals according to the present invention include, but are not limited to, human, canine, feline, bovine, caprine, equine, ovine, porcine, rodents, lagomorphs, primates, and the like, and encompass mammals in utero. Subjects may be of either gender and at any stage of development.
3. Administration and Dosing
The compounds of the present invention are generally administered in a therapeutically effective amount.
The compounds of the present invention can be administered by any suitable route in the form of a pharmaceutical composition adapted to such a route, and in a dose effective for the treatment intended. An effective dosage is typically in the range of about 0.001 to about 100 mg per kg body weight per day, preferably about 0.01 to about 30 mg/kg/day, in single or divided doses. Depending on age, species and condition being treated, dosage levels below the lower limit of this range may be suitable. In other cases, still larger doses may be used without harmful side effects. Larger doses may also be divided into several smaller doses, for administration throughout the day.
F. PHARMACEUTICAL COMPOSITIONS
For the treatment of the conditions referred to above, the compounds of described herein can be administered as follows:
Oral Administration
The compounds of the present invention may be administered orally, including by swallowing, so that the compound enters the gastrointestinal tract, or absorbed into the blood stream directly from the mouth (e.g., buccal or sublingual administration).
Suitable compositions for oral administration include solid formulations such as tablets, lozenges and capsules, which can contain liquids, gels, or powders.
Compositions for oral administration may be formulated as immediate or modified release, including delayed or sustained release, optionally with enteric coating.
Liquid formulations can include solutions, syrups and suspensions, which can be used in soft or hard capsules. Such formulations may include a pharmaceutically acceptable carrier, for example, water, ethanol, polyethylene glycol, cellulose, or an oil. The formulation may also include one or more emulsifying agents and/or suspending agents.
In a tablet dosage form the amount of drug present may be from about 0.05% to about 95% by weight, more typically from about 2% to about 50% by weight of the dosage form. In addition, tablets may contain a disintegrant, comprising from about 0.5% to about 35% by weight, more typically from about 2% to about 25% of the dosage form. Examples of disintegrants include methyl cellulose, sodium or calcium carboxymethyl cellulose, croscarmellose sodium, polyvinylpyrrolidone, hydroxypropyl cellulose, starch and the like.
Suitable lubricants, for use in a tablet, may be present in amounts from about 0.1% to about 5% by weight, and include calcium, zinc or magnesium stearate, sodium stearyl fumarate and the like.
Suitable binders, for use in a tablet, include gelatin, polyethylene glycol, sugars, gums, starch, hydroxypropyl cellulose and the like. Suitable diluents, for use in a tablet, include mannitol, xylitol, lactose, dextrose, sucrose, sorbitol and starch.
Suitable surface active agents and glidants, for use in a tablet, may be present in amounts from about 0.1% to about 3% by weight, and include polysorbate 80, sodium dodecyl sulfate, talc and silicon dioxide.
Parenteral Administration
Compounds of the present invention may be administered directly into the blood stream, muscle, or internal organs. Suitable means for parenteral administration include intravenous, intra-muscular, subcutaneous intraarterial, intraperitoneal, intrathecal, intracranial, and the like. Suitable devices for parenteral administration include injectors (including needle and needle-free injectors) and infusion methods.
Compositions for parenteral administration may be formulated as immediate or modified release, including delayed or sustained release.
Most parenteral formulations are aqueous solutions containing excipients, including salts, buffering agents and carbohydrates.
Parenteral formulations may also be prepared in a dehydrated form (e.g., by lyophilization) or as sterile non-aqueous solutions. These formulations can be used with a suitable vehicle, such as sterile water. Solubility-enhancing agents may also be used in preparation of parenteral solutions.
Topical Administration
Compounds of the present invention may be administered topically to the skin or transdermally. Formulations for this topical administration can include lotions, solutions, creams, gels, hydrogels, ointments, foams, implants, patches and the like. Pharmaceutically acceptable carriers for topical administration formulations can include water, alcohol, mineral oil, glycerin, polyethylene glycol and the like. Topical administration can also be performed by electroporation, iontophoresis, phonophoresis and the like.
Compositions for topical administration may be formulated as immediate or modified release, including delayed or sustained release.
G. COMBINATIONS AND COMBINATION THERAPY
The compounds of the present invention can be used, alone or in combination with other pharmaceutically active compounds, to treat conditions such as those previously described above. The compound(s) of the present invention and other pharmaceutically active compound(s) can be administered simultaneously (either in the same dosage form or in separate dosage forms) or sequentially. Accordingly, in one embodiment, the present invention comprises methods for treating a condition by administering to the subject a therapeutically-effective amount of one or more compounds of the present invention and one or more additional pharmaceutically active compounds.
In another embodiment, there is provided a pharmaceutical composition comprising one or more compounds of the present invention, one or more additional pharmaceutically active compounds, and a pharmaceutically acceptable carrier.
In another embodiment, the one or more additional pharmaceutically active compounds is selected from the group consisting of anti-inflammatory drugs, anti-atherosclerotic drugs, immunosuppressive drugs, immunomodulatory drugs, cytostatic drugs, anti-proliferative agents, angiogenesis inhibitors, kinase inhibitors, cytokine blockers and inhibitors of cell adhesion molecules.
p38 inhibitor compositions described herein are also optionally used in combination with other therapeutic reagents that are selected for their therapeutic value for the condition to be treated In general, the compositions described herein and, in embodiments where combinational therapy is employed, other agents do not have to be administered in the same pharmaceutical composition, and, because of different physical and chemical characteristics, are optionally administered by different routes. The initial administration is generally made according to established protocols, and then, based upon the observed effects, the dosage, modes of administration and times of administration subsequently modified. In certain instances, it is appropriate to administer a p38 inhibitor composition as described herein in combination with another therapeutic agent. By way of example only, if one of the side effects experienced by a patient upon receiving a p38 inhibitor composition as described herein is rash, then it is appropriate to administer an anti-histamine agent in combination with the initial therapeutic agent. Or, by way of example only, the therapeutic effectiveness of a p38 inhibitor is enhanced by administration of another therapeutic agent (which also includes a therapeutic regimen) that also has therapeutic benefit. In any case, regardless of the disease, disorder or condition being treated, the overall benefit experienced by the patient is either simply additive of the two therapeutic agents or the patient experiences a synergistic benefit.
Therapeutically effective dosages vary when the drugs are used in treatment combinations. Methods for experimentally determining therapeutically effective dosages of drugs and other agents for use in combination treatment regimens are documented methodologies. Combination treatment further includes periodic treatments that start and stop at various times to assist with the clinical management of the patient. In any case, the multiple therapeutic agents (one of which is a p38 inhibitor as described herein) are administered in any order, or even simultaneously. If simultaneously, the multiple therapeutic agents are optionally provided in a single, unified form, or in multiple forms (by way of example only, either as a single pill or as two separate pills).
In some embodiments, one of the therapeutic agents is given in multiple doses, or both are given as multiple doses. If not simultaneous, the timing between the multiple doses optionally varies from more than zero weeks to less than twelve weeks.
In addition, the combination methods, compositions and formulations are not to be limited to the use of only two agents, the use of multiple therapeutic combinations are also envisioned. It is understood that the dosage regimen to treat, prevent, or ameliorate the condition(s) for which relief is sought, is optionally modified in accordance with a variety of factors. These factors include the disorder from which the subject suffers, as well as the age, weight, sex, diet, and medical condition of the subject. Thus, the dosage regimen actually employed varies widely, in some embodiments, and therefore deviates from the dosage regimens set forth herein.
The pharmaceutical agents which make up the combination therapy disclosed herein are optionally a combined dosage form or in separate dosage forms intended for substantially simultaneous administration. The pharmaceutical agents that make up the combination therapy are optionally also administered sequentially, with either agent being administered by a regimen calling for two-step administration. The two-step administration regimen optionally calls for sequential administration of the active agents or spaced-apart administration of the separate active agents. The time period between the multiple administration steps ranges from, a few minutes to several hours, depending upon the properties of each pharmaceutical agent, such as potency, solubility, bioavailability, plasma half-life and kinetic profile of the pharmaceutical agent. Circadian variation of the target molecule concentration is optionally used to determine the optimal dose interval.
In another embodiment, a p38 inhibitor is optionally used in combination with procedures that provide additional or synergistic benefit to the patient. A p38 inhibitor and the additional therapy(ies) are optionally administered before, during or after the occurrence of a disease or condition, and the timing of administering the composition containing a p38 inhibitor varies in some embodiments. Thus, for example, a p38 inhibitor is used as a prophylactic and is administered continuously to subjects with a propensity to develop conditions or diseases in order to prevent the occurrence of the disease or condition. A p38 inhibitor and compositions are optionally administered to a subject during or as soon as possible after the onset of the symptoms. While embodiments of the present invention have been shown and described herein, it will be obvious to those skilled in the art that such embodiments are provided by way of example only. Numerous variations, changes, and substitutions will now occur to those skilled in the art without departing from the invention. It should be understood that in some embodiments of the invention various alternatives to the embodiments described herein are employed in practicing the invention.
A p38 inhibitor can be used in combination with drugs from the following classes: NSAIDs, immunosuppressive drugs, immunomodulatory drugs, cytostatic drugs, anti-proliferative agents, angiogenesis inhibitors, biological agents, steroids, vitamin D3 analogs, retinoids, other kinase inhibitors, cytokine blockers, corticosteroids and inhibitors of cell adhesion molecules. Where a subject is suffering from or at risk of suffering from atherosclerosis or a condition that is associated with atherosclerosis, a p38 inhibitor composition described herein is optionally used together with one or more agents or methods for treating atherosclerosis or a condition that is associated with atherosclerosis in any combination. Examples of therapeutic agents/treatments for treating atherosclerosis or a condition that is associated with atherosclerosis include, but are not limited to any of the following: torcetrapib, aspirin, niacin, HMG CoA reductase inhibitors (e.g., atorvastatin, fluvastatin, lovastatin, pravastatin, rosuvastatin and simvastatin), colesevelam, cholestyramine, colestipol, gemfibrozil, probucol and clofibrate.)
Where a subject is suffering from or at risk of suffering from an inflammatory condition, a p38 inhibitor composition described herein is optionally used together with one or more agents or methods for treating an inflammatory condition in any combination. Examples of therapeutic agents/treatments for treating an autoimmune and/or inflammatory condition include, but are not limited to any of the following: corticosteroids, nonsteroidal antiinflammatory drugs (NSAID) (e.g. ibuprofen, naproxen, acetaminophen, aspirin, Fenoprofen (NALFON), Flurbiprofen (ANSAID), Ketoprofen, Oxaprozin (DAYPRO), Diclofenac sodium (VOLTAREN), Diclofenac potassium (CATAFLAM), Etodolac (LODINE), Indomethacin (INDOCIN), Ketorolac (TORADOL), Sulindac (CLINORIL), Tolmetin (TOLECTIN), Meclofenamate (MECLOMEN), Mefenamic acid (PONSTEL), Nabumetone (RELAFEN), Piroxicam (FELDENE), COX-2 inhibitors (e.g., celecoxib (CELEBREX)), immunosuppressants (e.g., methotrexate (RHEUMATREX), leflunomide (ARAVA), azathioprine (IMURAN), cyclosporine (NEORAL, SANDIMMUNE), tacrolimus and cyclophosphamide (CYTOXAN), CD20 blockers (RITUXIMAB), Tumor Necrosis Factor (TNF) blockers (e.g., etanercept (ENBREL), infliximab (REMICADE) and adalimumab (HUMIRA), Abatacept (CTLA4-Ig) and interleukin-1 receptor antagonists (e.g. Anakinra (KINERET)), interleukin 6 inhibitors (e.g., ACTEMRA), interleukin 17 inhibitors (e.g., AIN457), Janus kinase inhibitors (e.g., TASOCITINIB), Syk inhibitors (e.g. R788), chloroquine and its derivatives.
For use in cancer and neoplastic diseases a p38 inhibitor is optimally used together with one or more of the following classes of drugs: wherein the anti-cancer agent is an EGFR kinase inhibitor, MEK inhibitor, VEGFR inhibitor, anti-VEGFR2 antibody, KDR antibody, AKT inhibitor, PDK-1 inhibitor, PI3K inhibitor, c-kit/Kdr tyrosine kinase inhibitor, Bcr-Abl tyrosine kinase inhibitor, VEGFR2 inhibitor, PDGFR-beta inhibitor, KIT inhibitor, Flt3 tyrosine kinase inhibitor, PDGF receptor family inhibitor, Flt3 tyrosine kinase inhibitor, RET tyrosine kinase receptor family inhibitor, VEGF-3 receptor antagonist, Raf protein kinase family inhibitor, angiogenesis inhibitor, Erb2 inhibitor, mTOR inhibitor, IGF-1R antibody, NFkB inhibitor, proteosome inhibitor, chemotherapy agent, or glucose reduction agent.
H. BIOLOGICAL EVALUATIONS
List of Biological Evaluation Abbreviations


    p38 Class of mitogen-activated protein kinases that are responsive to stress stimuli
    MAP Mitogen activated protein kinase
    MK2 Also known as MAPKAPK2. Refers to MAP kinase-activated protein kinase 2
    PRAK p38 regulated/activated kinase
    GST Glutathione S-transferase
    Hsp27 Heat-shock protein 27
    BSA Bovine serum albumin
    DTT Dithiothreitol
    ATP Adenosine triphosphate
    IC50 Amount of a drug that's needed to inhibit a process by half
    EC50 concentration of a drug which induces a response halfway between the baseline and maximum after a specified exposure time
    TNF Tumor necrosis factor
    IL Interleukin
    JNK c-Jun N-terminal kinase
    RPMI Roswell Park Memorial Institute medium. A medium for cell and tissue culture
    HWB Human whole blood
    DMEM Dulbecco's modified Eagle's medium. A vitamin and nutrient-enriched cell culture.
    FBS Fetal bovine serum
    RASF Rheumatoid arthritis synovial fibroblasts


Example C
p38 Inhibitory Potency and p38/MK2 Substrate Selectivity
This study evaluated the invention compound potency in inhibiting the p38 pathway. p38 activates MK2 and PRAK via phosphorylation, which both then interact with Hsp27, leading to increased inflammation and decreased ability to manage shock. The study measured the amount of the invention compound necessary to inhibit activation of MK2 and PRAK by half. This is a measurement of how effective the invention compound is in helping to lower inflammatory response, which helps treat many diseases, including autoimmune conditions, lymphoma, and rheumatoid arthritis. The novel, MK2 substrate-selective inhibitory mechanism of compounds is evaluated in enzyme assays comparing inhibitor potency in blocking p38/MK2 versus p38/PRAK induced phosphorylation of an HSP-27 derived peptide substrate. The ability of compounds to inhibit activated phospho-p38α is evaluated using a p38α/MK2 and a p38α/PRAK cascade assay format. The kinase activity of p38α is determined by its ability to phosphorylate GST-MK2 or GST-PRAK. Activation of MK2 or PRAK by p38α is quantitated by measuring the phosphorylation of a fluorescently-labeled, MK2/PRAK specific peptide substrate, Hsp27 peptide (FITC-KKKALSRQLSVAA). The phosphorylation of the Hsp27 peptide is quantified using IMAP technology (Molecular Devices, Sunnyvale Calif.). Kinase reactions are carried out in a 384-well plate (Greiner, 781280) in 20 mM HEPES pH 7.5, 10 mM MgCl2, 0.01% Triton X-100, 0.01% BSA, 1 mM DTT, and 2% DMSO. The inhibitor concentration is varied between 0.02-30,000 nM, while the Hsp27 peptide substrate and MgATP are held constant at 1 μM and 10 μM, respectively. Activated p38α is added to a final concentration of 30 μM for reactions with nonphosphorylated 1 nM GST-MK2 in the cascade reaction. For the p38α/PRAK cascade, unactivated GST-PRAK is held constant at 10 nM while p38α is added in to a final concentration of 200 pM. Kinase reactions are incubated at room temperature and quenched after 120 minutes by the addition of IMAP Binding Solution. Under these conditions, approximately 20% of the substrate Hsp27 peptide is phosphorylated. Reactions are initiated by the addition of activated p38α except for preincubation experiments, where reactions are initiated by the addition of Hsp27 peptide and MgATP. Preincubation of p38α with inhibitor or p38α with unactivated GST-MK2 or unactivated GST-PRAK and inhibitor are performed at 2× final assay concentrations at room temperature 240 minutes prior to adding ATP and Hsp27 peptide to initiate catalysis. The p38α compound inhibitory potency is quantitated from dose-response IC50 values or Ki values from p38α/MK2 cascade assays while the substrate selectivity is calculated as a ratio of p38α/PRAK:p38α/MK2 IC50 values. Species compounds of Formula (I), described hereinabove, evaluated in this assay, are expected to provide a therapeutic benefit in the treatment of p38 kinase mediated diseases, such as autoimmune diseases and lymphoma.
Compounds were tested in accordance with the above described assay, yielding IC50 values described below:














Compound

p38/MK2 IC50
p38/PRAK
Selectivity


Number
Structure
(μM)
IC50 (μM)
Ratio







49




0.021
8.1
385x









Example D
Cytokine Regulation in Human Monocytes
The p38 pathway has been shown to be critical for the biosynthesis of a number of proinflammatory cytokines including TNFα, IL-1β and IL-6. Therefore, inhibition of the p38 MAPK pathway will lower the inflammatory response by decreasing biosynthesis of proinflammatory cytokines. This study shows the amount of the invention compound necessary to inhibit biosynthesis of TNFα, IL-6, and IL-1β (proinflammatory cytokines) by half. This is a reflection of the invention compound's effectiveness in helping to lower inflammation, an effect which helps treat many diseases, including autoimmune conditions, lymphoma, and rheumatoid arthritis. Evaluation of the potency and efficacy of p38 inhibitors to block cytokine production is carried out using the human U937 cell line. The U937 human pre-monocytic cell line is obtained from the American Type Culture Collection (Rockville, Md.). These cells are differentiated to a monocytic/macrophage phenotype as described by Burnette (Burnette et al, (2009). SD0006: a potent, selective and orally available inhibitor of p38 kinase, Pharmacology 84(1):42-60). Differentiated U937 cells are seeded into 96-well tissue culture plates (200,000 cells/well) in complete media. After 24 hours, the cells are pretreated for 60 minutes in the presence or absence of compound and then stimulated with LPS (0.1 μg/mL) for 4 hours. Culture media are then collected for determination of TNFα, IL-6 or IL-1β levels by ELISA. Cytokine concentrations are extrapolated from recombinant protein standard curves using a four-parameter logistic model and solving for IC50 after iterating to the best least-squares fit. Species compounds of Formula (I), described hereinabove, evaluated in this assay, are expected to provide a therapeutic benefit in the treatment of p38 kinase mediated diseases, such as lymphoma or inflammation.
Compounds were tested in accordance with the above described assay, yielding IC50 values described below:


















U937,
A549
HWB
HWB
HWB




TNFα
IL-6
TNFα
IL-1β
IL-6


Compound

IC50
IC50
IC50
IC50
IC50


Number
Structure
(μM)
(μM)
(μM)
(μM)
(μM)







49




0.0016
0.0018
0.013
0.006
0.035









Example E
Phosphoprotein Analysis in Human Monocytes
This study shows the effectiveness and selectivity of the invention compound in inhibiting the JNK pathway. The JNK pathway leads to increased inflammation by boosting production of inflammatory cytokines. Inhibition of this pathway will lead to less inflammation and therefore will treat many diseases, including autoimmune conditions, lymphoma, and rheumatoid arthritis. Classical p38 inhibitors block the phosphorylation of downstream substrates of p38 while elevating activity of parallel pathways such as JNK. Evaluation of the impact of different classes of p38 inhibitors on p38 and JNK pathway regulation is carried out using phospho-HSP27 and phosphor-JNK for the two pathways, respectively. Evaluation of the potency and efficacy of p38 inhibitors to impact phosphoprotein levels is carried out using the human U937 cell line. The U937 human pre-monocytic cell line is obtained from the American Type Culture Collection (Rockville, Md.). These cells are differentiated to a monocytic/macrophage phenotype as described by Burnette (Burnette et al, (2009). SD0006: a potent, selective and orally available inhibitor of p38 kinase, Pharmacology 84(1):42-60). Suspension cells (approximately 0.5 million per milliliter in T75 cm2 cm tissue culture flasks) are grown in RPMI containing 10% fetal bovine serum (FBS) plus antibiotics. On day one, phorbol 12-myristate 13-acetate (PMA, 20 ng/ml) is added to the culture flask and the cells are incubated overnight at 37° C./5% CO2. The cells are washed on day two by centrifuging and resuspending them in fresh media without PMA. Adherent cells are harvested on day three by scraping, centrifuging and resuspending them in fresh media at a density of 1 million per milliliter. The PMA-differentiated U937 cells are then distributed into each well of a 96-well flat bottom tissue culture plate (100 ml/well) and the 100,000 cells/well are allowed to recover, incubated, overnight. On the day of the assay fresh media (50 ml/well) is added to the plates followed by the addition of compound (25 ml/well, concentration response) for 1 hour. The cells are stimulated with LPS (100 ng/ml) in a final assay volume of 100 ml. After 30 minutes, complete lysis buffer (50 ml/well MSD Tris lysis buffer, supplemented with protease inhibitors and phosphatase inhibitors) is added and the plate is placed on a shaker at 4° C. for 30 minutes before being stored frozen at −20° C. The cellular lysate (25 ml/well) is thawed and transferred from the assay plate to Meso Scale detection plates for determination of phospho-Hsp27/total Hsp27 or phospho-JNK/total JNK.
Compounds were tested in accordance with the above described assay, yielding IC50 and EC50 values described below:
















pHSP27/Total
pJNK/Total



Compound

HSP27
JNK
Selectivity


Number
Structure
IC50 (nM)
EC50 (nM)
Ratio







49




1.15x
117x
102x









Example F
Endotoxin-Induced Cytokine Production from Human Whole Blood
Human whole blood (HWB; 25-45 ml) was collected from an NSAID-free donor into vacutainer collection tubes containing sodium heparin (10 ml, 158 USP units), pooled and rocked gently before being distributed into each well of a 96-well round bottom tissue culture plate (180 ml/well). Compounds (10 ml/well, concentration response) are added and mixed gently for 15-20 seconds using a disposable 96 polypropylene pin tool before the plates are incubated at 37° C./5% CO2 for 1 hour. The HWB is stimulated with LPS (100 ng/ml) in a final assay volume of 200 ml. After 3 hours, the plates are spun at 240×g for 5 minutes to pellet the red cells. The plasma is carefully transferred to another 96-well round bottom plate and diluted 2-fold with assay media (DMEM containing 10% fetal bovine serum (FBS) plus antibiotics). Finally, the diluted plasma (25 ml/well) is transferred to Meso Scale detection plates for determination of IL-1, IL-6 or TNFα.
Example G
Determination of IL-1-Induced IL-6 Production in A549 Cells
Adherent A549 cells (approximately 5 million per T75 cm2 tissue culture flask) are grown in F-12K media containing 10% fetal bovine serum (FBS) plus antibiotics. The cells are trypsinized, washed and resuspended at 0.3 million per milliliter. A549 cells are then distributed into each well of a 96-well flat bottom tissue culture plate (100 ml/well) and the 30,000 cells/well are allowed to recover, incubated, overnight. On the day of the assay fresh media (50 ml/well) is added to the plates followed by the addition of compound (25 ml/well, concentration response) for 1 hour. The cells are stimulated with LPS (100 ng/ml) in a final assay volume of 100 ml. After 3 hours, cultured media (25 ml/well) is transferred from the assay plate to a Meso Scale custom coated detection plate for determination of IL-6 levels. The detection plate is incubated at 4° C. overnight followed by the addition of a sulfo-tagged antibody cocktail (25 ml/well) for 1 hour at room temperature, with vigorous shaking. Read buffer (150 ml/well, MSD 4× read buffer diluted 4-fold with dH2O) is added and the plate is read using the Meso Scale Sector Imager 6000. Upon electrical stimulation of the detection plate, co-reactants in the read buffer enhance an electrochemical reaction resulting in the release of energy in the form of light. This signal is captured by an internal CCD camera and quantitated. Viability of A549 cells is determined using an MTT assay. After the 3 hour incubation of cells with LPS and collection of the cultured media, the assay plates are inverted and gently tapped to remove any remaining liquid. MTT (1 mg/ml solution prepared in assay media) is added (100 ml/well) and the plates are returned to the 37° C./5% CO2 incubator for 3 hours. The plates are again inverted to remove any liquid and allowed to dry overnight. Isopropanol (100 ml/well) is added to solubilize the resulting formazan crystals and the plate is read at 570 nm/650 nm using a Molecular Devices SpectraMax spectrophotometer.
Example H
IL-1β Induced Prostaglandin Production in Rheumatoid Arthritis Synovial Fibroblasts (RASF)
Rheumatoid arthritis synovial fibroblasts (RASF) are derived from the inflamed synovium of a female RA patient who was undergoing total knee replacement. Synovial tissue is teased away from adjacent cartilage and dispersed into single cells with collagenase. Cells are expanded and banked. RASF cells are further cultured as described by Burnette supra. RASF cells are seeded into 96-well tissue culture plates (5×104 cells/well) in complete growth medium. After 24 hours, the medium is replaced with fresh growth medium containing 1% FBS. Cells are treated with serial concentrations (30,000-0.01 nM) of compound or dimethyl-sulfoxide (DMSO) vehicle control for 1 hour then stimulated with 1 ng/mL IL-1β (R&D Systems, Minneapolis, Minn.) for 18-20 hours at 37° C. and conditioned media collected. PGE2 levels the in cultured media are quantitated by ELISA (Cayman Chemical, Ann Arbor, Mich.). Species compounds of Formula (I), described hereinabove, evaluated in this assay, are expected to provide a therapeutic benefit in the treatment of p38 kinase mediated diseases, such as lymphoma or rheumatoid arthritis.
Example J
Substrate Selectivity in HUVEC Cells
When a compound is identified from the biochemical characterization step with selective inhibition of p38/MK2, it is next placed into a cell-based assay to verify enzyme to cell translatability. These assays utilize human umbilical vein endothelial cells (HUVEC) to demonstrate inhibition of Hsp27 phosphorylation (a biomarker of p38/MK2 activation) while sparing production of tissue factor (TF), which is linked to another downstream substrate of p38, MSK. In a 96-well format, adherent HUVEC (at 5 passages or less) are treated for 1 hour with serially-diluted compounds, including a non-selective p38 inhibitor as a reference, or vehicle for controls. For Hsp27 phosphorylation, cells are then stimulated with 500 pg/mL IL-1β for 0.5 hours, media is removed, cells are lysed, and phospho-Hsp27 in the lysate is quantitated by enzyme-linked immunosorbent assay (ELISA)(Life Technologies, Carlsbad, Calif.). The procedure for TF release is a similar ELISA-based assay (American Diagnostica, Stanford, Conn.), except that IL-1β stimulation proceeds for 5 hours. The ratio of TF inhibition IC50:HSP27 phosphorylation inhibition IC50 is defined as the substrate selectivity index in these cells. Species compounds of Formula (I), described hereinabove, evaluated in this assay, are expected to provide a therapeutic benefit in the treatment of p38 kinase mediated diseases, such as lymphoma and auto-inflammatory disease.
Example K
Canine B Cell Growth Regulation
p38 inhibitors have been shown to uniquely inhibit canine B cell proliferation and survival. This selective effect on canine B cells may be exploited in therapeutic treatment for canine B cell lymphoma, a fatal disease that impacts>40,000 companion animals in the United States. Quantitation of impact of p38 inhibitors on B cell growth is a cellular indicator of efficacy in B cell lymphoma. Species compounds of Formula (I), described hereinabove, evaluated in this assay, are expected to provide a therapeutic benefit in the treatment of p38 kinase mediated diseases, such as lymphoma. These assays utilize beagle dog spleens obtained with protocols approved by the Saint Louis University Animal Care and Use Committee in collaboration with Seventh Wave Laboratories. Leukocytes are isolated from splenocytes by centrifugation through Histopaque 1077. To evaluate effect on proliferation, leukocytes are then cultured for 48 hours in 96-well plates in the presence of vehicle or test compounds. Cells are stimulated with LPS for TLR4 stimulation, Staphylococcus aureus B cell mitogen, or concanavalin-A T cell mitogen, then proliferation is quantitated with a BRDU incorporation ELISA (Roche, Mannheim, Germany). For apoptosis experiments, leukocytes are plated on 96-well polypropylene U bottom plates and treated with p38 MAPK inhibitors or staurosporine (as a positive control) for up to 24 hours in the absence or presence of actinomycin D or cycloheximide (if needed to increase apoptosis rate). Apoptosis is determined using Caspase-Glo 3/7 luminescent assay (Promega, Madison, Wis.). In both assays, values generated after incubation with increasing concentrations of the inhibitors are compared to a negative control without inhibitors.
Example L
LPS Induced TNFα Production in Rats
Rats are fasted eighteen hours prior to oral dosing, and allowed free access to water throughout the experiment. Each treatment group consists of five animals. Compounds are prepared as a suspension in a vehicle consisting of 0.5% methylcellulose, (Sigma Aldrich, St. Louis, Mo.), 0.025% Tween 20 (Sigma Aldrich). The compound or vehicle is administered by oral gavage in a volume of 1 mL. Two vehicle groups are used per experiment to control for intra-experiment variability. LPS (E. coli serotype 0111:B4, Sigma Aldrich) is administered four hours after compound intravenous injection at a dose of 1 mg/kg in 0.5 mL sterile saline (Baxter Healthcare, Deerfield, Ill.). Blood is collected in serum separator tubes via cardiac puncture ninety minutes after LPS injection, a time point corresponding to maximal TNFα and IL-1β production. After clotting, serum is withdrawn and stored at −20° C. and IL-1β and TNFα levels quantitated by ELISA (Burnette supra). Species compounds of Formula (I), described hereinabove, evaluated in this assay, are expected to provide a therapeutic benefit in the treatment of p38 kinase mediated diseases, such as lymphoma or inflammation.
All mentioned documents are incorporated by reference as if herein written. When introducing elements of the present invention or the exemplary embodiment(s) thereof, the articles “a,” “an,” “the” and “said” are intended to mean that there are one or more of the elements. The terms “comprising,” “including” and “having” are intended to be inclusive and mean that there may be additional elements other than the listed elements. Although this invention has been described with respect to specific embodiments, the details of these embodiments are not to be construed as limitations.

## Claims

1. A compound of Formula (I):



or a pharmaceutically acceptable salt thereof, wherein:
X is CH or N;
R1 is selected from the group consisting of H, C1-C6 alkyl, fluoro, chloro, bromo, cyano, or —CF3;
R2 is selected from the group consisting of H, methyl, cyano, or fluoro;
R3 is selected from the group consisting of:






R4 is selected from the group consisting of H, methyl, OH, and O—CH3;
R5 is H or C1-C3 alkyl;
m is 1 or 2;
n is 0 or 1;
p is 1; and
q is 0 or 1.

2. The compound of claim 1, wherein the compound has the structure of Formula (II):



or a pharmaceutically acceptable salt thereof, wherein:
X is CH or N;
R1 is selected from the group consisting of H, C1-C6 alkyl, fluoro, chloro, bromo, cyano, or —CF3;
R2 is selected from the group consisting of H, methyl, cyano, or fluoro;
R3 is






R4 is selected from the group consisting of H, methyl, OH, and —OCH3;
R5 is H or C1-C3 alkyl;
m is 1 or 2; and
n is 0 or 1.

3. The compound of claim 2, wherein R3 is selected from the group consisting of 3,5-difluoropyridin-2-yl, 3-fluoropyridin-2-yl, 5-fluoro-3-methylpyridin-2-yl, 6-fluoropyridin-2-yl, 6-fluoro-4-methylpyridin-2-yl, 3-fluoro-5-methylpyridin-2-yl, and 5-fluoropyridin-2-yl.

4. The compound of claim 3, wherein the compound has the structure of Formula (III):



or a pharmaceutically acceptable salt thereof, wherein:
X is CH or N;
R1 is chloro or bromo; and
R2 is —H or methyl.

5. The compound of claim 4, wherein the compound is selected from the group consisting of:
3-bromo-4-((3,5-difluoropyridin-2-yl)methoxy)-6′′-(2-hydroxypropan-2-yl)-3′′,5′,6-trimethyl-2H-[1,4′:2′,2′′-terpyridin]-2-one;
3-chloro-4-((3,5-difluoropyridin-2-yl)methoxy)-6′′-(2-hydroxypropan-2-yl)-3′′,5′,6-trimethyl-2H-[1,4′:2′′,2′′-terpyridin]-2-one;
3-bromo-4-((3,5-difluoropyridin-2-yl)methoxy)-2′-(2-(2-hydroxypropan-2-yl)-5-methylpyrimidin-4-yl)-5′,6-dimethyl-2H-[1,4′-bipyridin]-2-one;
3-chloro-4-((3,5-difluoropyridin-2-yl)methoxy)-2′-(2-(2-hydroxypropan-2-yl)-5-methylpyrimidin-4-yl)-5′,6-dimethyl-2H-[1,4′-bipyridin]-2-one;
3-bromo-4-((3,5-difluoropyridin-2-yl)methoxy)-6′′-(2-hydroxypropan-2-yl)-5′,6-dimethyl-2H-[1,4′:2′,2′′-terpyridin]-2-one;
3-chloro-4-((3,5-difluoropyridin-2-yl)methoxy)-6′′-(2-hydroxypropan-2-yl)-5′,6-dimethyl-2H-[1,4′:2′,2′′-terpyridin]-2-one;
3-bromo-4-((3,5-difluoropyridin-2-yl)methoxy)-2′-(2-(2-hydroxypropan-2-yl)pyrimidin-4-yl)-5′,6-dimethyl-2H-[1,4′-bipyridin]-2-one;
3-chloro-4-((3,5-difluoropyridin-2-yl)methoxy)-2′-(2-(2-hydroxypropan-2-yl)pyrimidin-4-yl)-5′,6-dimethyl-2H-[1,4′-bipyridin]-2-one.

6. The compound of claim 3, wherein the compound is selected from the group consisting of:
3-chloro-4-((5-fluoropyridin-2-yl)methoxy)-2′-(2-(2-hydroxypropan-2-yl)pyrimidin-4-yl)-5′,6-dimethyl-2H-[1,4′-bipyridin]-2-one;
3-chloro-4-((3-fluoropyridin-2-yl)methoxy)-2′-(2-(2-hydroxypropan-2-yl)pyrimidin-4-yl)-5′,6-dimethyl-2H-[1,4′-bipyridin]-2-one;
3-chloro-4-((5-fluoro-3-methylpyridin-2-yl)methoxy)-2′-(2-(2-hydroxypropan-2-yl)pyrimidin-4-yl)-5′,6-dimethyl-2H-[1,4′-bipyridin]-2-one;
3-chloro-4-((6-fluoropyridin-2-yl)methoxy)-2′-(2-(2-hydroxypropan-2-yl)pyrimidin-4-yl)-5′,6-dimethyl-2H-[1,4′-bipyridin]-2-one;
3-chloro-4-((6-fluoro-4-methylpyridin-2-yl)methoxy)-2′-(2-(2-hydroxypropan-2-yl)pyrimidin-4-yl)-5′,6-dimethyl-2H-[1,4′-bipyridin]-2-one;
3-chloro-4-((3-fluoro-5-methylpyridin-2-yl)methoxy)-2′-(2-(2-hydroxypropan-2-yl)pyrimidin-4-yl)-5′,6-dimethyl-2H-[1,4′-bipyridin]-2-one;
3-bromo-4-((5-fluoropyridin-2-yl)methoxy)-2′-(2-(2-hydroxypropan-2-yl)pyrimidin-4-yl)-5′,6-dimethyl-2H-[1,4′-bipyridin]-2-one;
3-chloro-4-((5-fluoropyridin-2-yl)methoxy)-6′′-(2-hydroxypropan-2-yl)-5′,6-dimethyl-2H-[1,4′:2′,2′′-terpyridin]-2-one;
3-bromo-4-((5-fluoropyridin-2-yl)methoxy)-6′′-(2-hydroxypropan-2-yl)-5′,6-dimethyl-2H-[1,4′:2′,2′′-terpyridin]-2-one;
3-chloro-4-((5-fluoropyridin-2-yl)methoxy)-2′-(2-(2-hydroxypropan-2-yl)-5-methylpyrimidin-4-yl)-5′,6-dimethyl-2H-[1,4′-bipyridin]-2-one;
3-bromo-4-((5-fluoropyridin-2-yl)methoxy)-2′-(2-(2-hydroxypropan-2-yl)-5-methylpyrimidin-4-yl)-5′,6-dimethyl-2H-[1,4′-bipyridin]-2-one;
3-chloro-4-((5-fluoropyridin-2-yl)methoxy)-6′′-(2-hydroxypropan-2-yl)-3′′,5′,6-trimethyl-2H-[1,4′:2′,2′′-terpyridin]-2-one;
3-bromo-4-((5-fluoropyridin-2-yl)methoxy)-6′′-(2-hydroxypropan-2-yl)-3′′,5′,6-trimethyl-2H-[1,4′:2′,2′′-terpyridin]-2-one;
3-bromo-4-((3-fluoropyridin-2-yl)methoxy)-2′-(2-(2-hydroxypropan-2-yl)pyrimidin-4-yl)-5′,6-dimethyl-2H-[1,4′-bipyridin]-2-one;
3-chloro-4-((3-fluoropyridin-2-yl)methoxy)-6′′-(2-hydroxypropan-2-yl)-5′,6-dimethyl-2H-[1,4′:2′,2′′-terpyridin]-2-one;
3-bromo-4-((3-fluoropyridin-2-yl)methoxy)-6′′-(2-hydroxypropan-2-yl)-5′,6-dimethyl-2H-[1,4′:2′,2′′-terpyridin]-2-one;
3-chloro-4-((3-fluoropyridin-2-yl)methoxy)-2′-(2-(2-hydroxypropan-2-yl)-5-methylpyrimidin-4-yl)-5′,6-dimethyl-2H-[1,4′-bipyridin]-2-one;
3-bromo-4-((3-fluoropyridin-2-yl)methoxy)-2′-(2-(2-hydroxypropan-2-yl)-5-methylpyrimidin-4-yl)-5′,6-dimethyl-2H-[1,4′-bipyridin]-2-one;
3-chloro-4-((3-fluoropyridin-2-yl)methoxy)-6′′-(2-hydroxypropan-2-yl)-3′′,5′,6-trimethyl-2H-[1,4′:2′,2′′-terpyridin]-2-one;
3-bromo-4-((3-fluoropyridin-2-yl)methoxy)-6′′-(2-hydroxypropan-2-yl)-3′′,5′,6-trimethyl-2H-[1,4′:2′,2′′-terpyridin]-2-one;
3-bromo-4-((5-fluoro-3-methylpyridin-2-yl)methoxy)-2′-(2-(2-hydroxypropan-2-yl)pyrimidin-4-yl)-5′,6-dimethyl-2H-[1,4′-bipyridin]-2-one;
3-chloro-4-((5-fluoro-3-methylpyridin-2-yl)methoxy)-6′′-(2-hydroxypropan-2-yl)-5′,6-dimethyl-2H-[1,4′:2′,2′′-terpyridin]-2-one;
3-bromo-4-((5-fluoro-3-methylpyridin-2-yl)methoxy)-6′′-(2-hydroxypropan-2-yl)-5′,6-dimethyl-2H-[1,4′:2′,2′′-terpyridin]-2-one;
3-chloro-4-((5-fluoro-3-methylpyridin-2-yl)methoxy)-2′-(2-(2-hydroxypropan-2-yl)-5-methylpyrimidin-4-yl)-5′,6-dimethyl-2H-[1,4′-bipyridin]-2-one;
3-bromo-4-((5-fluoro-3-methylpyridin-2-yl)methoxy)-2′-(2-(2-hydroxypropan-2-yl)-5-methylpyrimidin-4-yl)-5′,6-dimethyl-2H-[1,4′-bipyridin]-2-one;
3-chloro-4-((5-fluoro-3-methylpyridin-2-yl)methoxy)-6′′-(2-hydroxypropan-2-yl)-3′′,5′,6-trimethyl-2H-[1,4′:2′,2′′-terpyridin]-2-one;
3-bromo-4-((5-fluoro-3-methylpyridin-2-yl)methoxy)-6′′-(2-hydroxypropan-2-yl)-3′′,5′,6-trimethyl-2H-[1,4′:2′,2′′-terpyridin]-2-one;
3-bromo-4-((6-fluoropyridin-2-yl)methoxy)-2′-(2-(2-hydroxypropan-2-yl)pyrimidin-4-yl)-5′,6-dimethyl-2H-[1,4′-bipyridin]-2-one;
3-chloro-4-((6-fluoropyridin-2-yl)methoxy)-6′′-(2-hydroxypropan-2-yl)-5′,6-dimethyl-2H-[1,4′:2′,2′′-terpyridin]-2-one;
3-bromo-4-((6-fluoropyridin-2-yl)methoxy)-6′′-(2-hydroxypropan-2-yl)-5′,6-dimethyl-2H-[1,4′:2′,2′′-terpyridin]-2-one;
3-chloro-4-((6-fluoropyridin-2-yl)methoxy)-2′-(2-(2-hydroxypropan-2-yl)-5-methylpyrimidin-4-yl)-5′,6-dimethyl-2H-[1,4′-bipyridin]-2-one;
3-bromo-4-((6-fluoropyridin-2-yl)methoxy)-2′-(2-(2-hydroxypropan-2-yl)-5-methylpyrimidin-4-yl)-5′,6-dimethyl-2H-[1,4′-bipyridin]-2-one;
3-chloro-4-((6-fluoropyridin-2-yl)methoxy)-6′′-(2-hydroxypropan-2-yl)-3′′,5′,6-trimethyl-2H-[1,4′:2′,2′′-terpyridin]-2-one;
3-bromo-4-((6-fluoropyridin-2-yl)methoxy)-6′′-(2-hydroxypropan-2-yl)-3′′,5′,6-trimethyl-2H-[1,4′:2′,2′′-terpyridin]-2-one;
3-bromo-4-((6-fluoro-4-methylpyridin-2-yl)methoxy)-2′-(2-(2-hydroxypropan-2-yl)pyrimidin-4-yl)-5′,6-dimethyl-2H-[1,4′-bipyridin]-2-one;
3-chloro-4-((6-fluoro-4-methylpyridin-2-yl)methoxy)-6′′-(2-hydroxypropan-2-yl)-5′,6-dimethyl-2H-[1,4′:2′,2′′-terpyridin]-2-one;
3-bromo-4-((6-fluoro-4-methylpyridin-2-yl)methoxy)-6′′-(2-hydroxypropan-2-yl)-5′,6-dimethyl-2H-[1,4′:2′,2′′-terpyridin]-2-one;
3-chloro-4-((6-fluoro-4-methylpyridin-2-yl)methoxy)-2′-(2-(2-hydroxypropan-2-yl)-5-methylpyrimidin-4-yl)-5′,6-dimethyl-2H-[1,4′-bipyridin]-2-one;
3-bromo-4-((6-fluoro-4-methylpyridin-2-yl)methoxy)-2′-(2-(2-hydroxypropan-2-yl)-5-methylpyrimidin-4-yl)-5′,6-dimethyl-2H-[1,4′-bipyridin]-2-one;
3-chloro-4-((6-fluoro-4-methylpyridin-2-yl)methoxy)-6′′-(2-hydroxypropan-2-yl)-3′′,5′,6-trimethyl-2H-[1,4′:2′,2′′-terpyridin]-2-one;
3-bromo-4-((6-fluoro-4-methylpyridin-2-yl)methoxy)-6′′-(2-hydroxypropan-2-yl)-3′′,5′,6-trimethyl-2H-[1,4′:2′,2′′-terpyridin]-2-one;
3-bromo-4-((3-fluoro-5-methylpyridin-2-yl)methoxy)-2′-(2-(2-hydroxypropan-2-yl)pyrimidin-4-yl)-5′,6-dimethyl-2H-[1,4′-bipyridin]-2-one;
3-chloro-4-((3-fluoro-5-methylpyridin-2-yl)methoxy)-6′′-(2-hydroxypropan-2-yl)-5′,6-dimethyl-2H-[1,4′:2′,2′′-terpyridin]-2-one;
3-bromo-4-((3-fluoro-5-methylpyridin-2-yl)methoxy)-6′′-(2-hydroxypropan-2-yl)-5′,6-dimethyl-2H-[1,4′:2′,2′′-terpyridin]-2-one;
3-chloro-4-((3-fluoro-5-methylpyridin-2-yl)methoxy)-2′-(2-(2-hydroxypropan-2-yl)-5-methylpyrimidin-4-yl)-5′,6-dimethyl-2H-[1,4′-bipyridin]-2-one;
3-bromo-4-((3-fluoro-5-methylpyridin-2-yl)methoxy)-2′-(2-(2-hydroxypropan-2-yl)-5-methylpyrimidin-4-yl)-5′,6-dimethyl-2H-[1,4′-bipyridin]-2-one;
3-chloro-4-((3-fluoro-5-methylpyridin-2-yl)methoxy)-6′′-(2-hydroxypropan-2-yl)-3′′,5′,6-trimethyl-2H-[1,4′:2′,2′′-terpyridin]-2-one; and
3-bromo-4-((3-fluoro-5-methylpyridin-2-yl)methoxy)-6′′-(2-hydroxypropan-2-yl)-3′′,5′,6-trimethyl-2H-[1,4′:2′,2′′-terpyridin]-2-one.

7. The compound of claim 1, wherein the compound has the structure of Formula (IV):



or a pharmaceutically acceptable salt thereof, wherein:
X is CH or N;
R1 is selected from the group consisting of H, C1-C6 alkyl, fluoro, chloro, bromo, cyano, or —CF3;
R2 is selected from the group consisting of H, methyl, cyano, or fluoro;
R4 is selected from the group consisting of H, methyl, OH, and —OCH3;
R5 is H or C1-C3 alkyl;
m is 1 or 2; and
n is 0 or 1.

8. The compound of claim 7, wherein the compound has the structure of Formula (V):



or a pharmaceutically acceptable salt thereof, wherein:
X is CH or N;
R1 is chloro or bromo; and
R2 is —H or methyl.

9. The compound of claim 8, wherein the compound is selected from the group consisting of:
3-chloro-4-((4-fluoropyridin-3-yl)methoxy)-2′-(2-(2-hydroxypropan-2-yl)pyrimidin-4-yl)-5′,6-dimethyl-2H-[1,4′-bipyridin]-2-one;
3-bromo-4-((4-fluoropyridin-3-yl)methoxy)-2′-(2-(2-hydroxypropan-2-yl)pyrimidin-4-yl)-5′,6-dimethyl-2H-[1,4′-bipyridin]-2-one;
3-chloro-4-((4-fluoropyridin-3-yl)methoxy)-6′′-(2-hydroxypropan-2-yl)-5′,6-dimethyl-2H-[1,4′:2′,2′′-terpyridin]-2-one;
3-bromo-4-((4-fluoropyridin-3-yl)methoxy)-6′′-(2-hydroxypropan-2-yl)-5′,6-dimethyl-2H-[1,4′:2′,2′′-terpyridin]-2-one;
3-chloro-4-((4-fluoropyridin-3-yl)methoxy)-2′-(2-(2-hydroxypropan-2-yl)-5-methylpyrimidin-4-yl)-5′,6-dimethyl-2H-[1,4′-bipyridin]-2-one;
3-bromo-4-((4-fluoropyridin-3-yl)methoxy)-2′-(2-(2-hydroxypropan-2-yl)-5-methylpyrimidin-4-yl)-5′,6-dimethyl-2H-[1,4′-bipyridin]-2-one;
3-chloro-4-((4-fluoropyridin-3-yl)methoxy)-6′′-(2-hydroxypropan-2-yl)-3′′,5′,6-trimethyl-2H-[1,4′:2′,2′′-terpyridin]-2-one; and
3-bromo-4-((4-fluoropyridin-3-yl)methoxy)-6′′-(2-hydroxypropan-2-yl)-3′′,5′,6-trimethyl-2H-[1,4′:2′,2′′-terpyridin]-2-one.

10. The compound of claim 1, wherein the compound has the structure of Formula (VI):



or a pharmaceutically acceptable salt thereof, wherein:
X is CH or N;
R1 is selected from the group consisting of H, C1-C6 alkyl, fluoro, chloro, bromo, cyano, or —CF3;
R2 is selected from the group consisting of H, methyl, cyano, or fluoro;
R4 is selected from the group consisting of H, methyl, OH, and —OCH3;
R5 is H or C1-C3 alkyl;
p is 1; and
q is 0 or 1.

11. The compound of claim 10, wherein the compound has the structure of Formula (VII):



or a pharmaceutically acceptable salt thereof, wherein:
X is CH or N;
R1 is chloro or bromo; and
R2 is —H or methyl.

12. The compound of claim 11, wherein the compound is selected from the group consisting of:
3-chloro-4-((5-fluoropyrimidin-4-yl)methoxy)-2′-(2-(2-hydroxypropan-2-yl)pyrimidin-4-yl)-5′,6-dimethyl-2H-[1,4′-bipyridin]-2-one;
3-bromo-4-((5-fluoropyrimidin-4-yl)methoxy)-2′-(2-(2-hydroxypropan-2-yl)pyrimidin-4-yl)-5′,6-dimethyl-2H-[1,4′-bipyridin]-2-one;
3-chloro-4-((5-fluoropyrimidin-4-yl)methoxy)-6′′-(2-hydroxypropan-2-yl)-5′,6-dimethyl-2H-[1,4′:2′,2′′-terpyridin]-2-one;
3-bromo-4-((5-fluoropyrimidin-4-yl)methoxy)-6′′-(2-hydroxypropan-2-yl)-5′,6-dimethyl-2H-[1,4′:2′,2′′-terpyridin]-2-one;
3-chloro-4-((5-fluoropyrimidin-4-yl)methoxy)-2′-(2-(2-hydroxypropan-2-yl)-5-methylpyrimidin-4-yl)-5′,6-dimethyl-2H-[1,4′-bipyridin]-2-one;
3-bromo-4-((5-fluoropyrimidin-4-yl)methoxy)-2′-(2-(2-hydroxypropan-2-yl)-5-methylpyrimidin-4-yl)-5′,6-dimethyl-2H-[1,4′-bipyridin]-2-one;
3-chloro-4-((5-fluoropyrimidin-4-yl)methoxy)-6′′-(2-hydroxypropan-2-yl)-3′′,5′,6-trimethyl-2H-[1,4′:2′,2′′-terpyridin]-2-one; and
3-bromo-4-((5-fluoropyrimidin-4-yl)methoxy)-6′′-(2-hydroxypropan-2-yl)-3′′,5′,6-trimethyl-2H-[1,4′:2′,2′′-terpyridin]-2-one.

13. A pharmaceutical composition comprising a therapeutically effective amount of a compound of claim 1 or a pharmaceutically acceptable salt thereof, and a pharmaceutically acceptable carrier.

14. The pharmaceutical composition of claim 13, further comprising a therapeutically effective amount of an active pharmaceutical ingredient selected from the group consisting of anti-inflammatory drugs, anti-atherosclerotic drugs, immunosuppressive drugs, immunomodulatory drugs, cytostatic drugs, angiogenesis inhibitors, kinase inhibitors, cytokine blockers and inhibitors of cell adhesion molecules.