A compound of Formula (I): pharmaceutically acceptable salts thereof, deuterated analogs thereof, compositions thereof, and methods of treating disease using a compound thereof are disclosed.

FIELD

The present disclosure relates to novel compounds that are inhibitors of the kinase IRAK4. The disclosure also relates to methods for preparing the compounds and to pharmaceutical compositions comprising such compounds.

BACKGROUND

Interleukin-1 receptor-associated kinase-4 (IRAK4) is a serine-threonine kinase which acts as a mediator in interleukin-1/Toll-like receptor (IL-1/TLR) signaling cascades. More particularly, IRAK4 is involved in activation of adaptor protein myeloid differentiation primary response gene 88 (MyD88) signaling cascades and is hypothesized to play a role in inflammatory and fibrotic disorders, such as rheumatoid arthritis (RA), inflammatory bowel disease (IBD), gout, Lyme disease, arthritis, psoriasis, pelvic inflammatory disease, systemic lupus erythematosus (SLE), Sjogren's syndrome, viral myocarditis, acute and chronic tissue injury, non-alcoholic steatohepatitis (NASH), alcoholic hepatitis and kidney disease, including chronic kidney disease and diabetic kidney disease. In addition, IRAK4 plays a role in certain cancers and is hypothesized to play a role in inflammation associated with gastrointestinal infections, includingC. difficile. Signaling through IL-1R/TLR results in the activation of MyD88 which recruits IRAK4 and IRAK1 to form a signaling complex. This complex then interacts with a series of kinases, adaptor proteins, and ligases, ultimately resulting in the activation of nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB), activator protein-I (AP1), cyclic AMP-responsive element-binding protein (CREB) and the interferon-regulatory factors (IRFs), including IRF5 and IRF7, inducing the generation of pro-inflammatory cytokines and type I interferons.

In addition, certain cancers, including lymphomas, may contain one or more mutations in the MYD88 adaptor protein, leading to a constitutively active signaling cascade that may promote survival of tumor cells. (Kelly et al., IRAK4 inhibitors for autoimmunity and lymphoma, J. Exp. Med. 2015 Vol. 212 No. 13 2189-2201)

Therefore, an inhibitor of IRAK4 may be useful in the treatment of cancers, including lymphomas.

There are currently no approved IRAK4 inhibiting pharmaceuticals. Therefore, it would be useful to provide an IRAK4 inhibiting compound with properties suitable for administration as a pharmaceutical agent to a mammal, particularly a human. Considerations for selecting a pharmaceutical compound are multifactorial. Compound characteristics including on-target potency, pharmacokinetics, pKa, solubility, stability (e.g., metabolic stability) and off-target liabilities are frequently profiled.

WO2016210034, WO2016210036, WO2015150995, WO2016127024, and WO2016210037 recite compounds said to be useful as IRAK4 inhibitors.

IRAK1 functions as a cytokine initiated by the binding of ligands to IL-1R and TLRs. Activation of the IL-1 and TLR signaling pathways can be triggered by a variety of stimuli, including recognition of microbial pathogens or products, such as LPS, the presence of reactive oxygen species, recognition of DNA damage, abnormalities in the tissue matrix caused by chronic inflammation, and genetic factors, such as amplification of 1921.3 and overproduction of S100A proteins.

The inflammatory cytokines sIL-17A, sIL-2, and sIL-6 are regulated by IRAK1, and inhibition of IRAK1 by the late-stage clinical compound pacritinib suppresses induced immunglobulin synthesis in normal human lymphocytes. In normal human monocytes, IRAK1 induces inflammatory cytokines upon LPS challenge. (See, for example, Singer, J. et al. INHIBITION OF INTERLEUKIN-1 RECEPTOR-ASSOCIATED KINASE 1 (IRAK1) AS A THERAPEUTIC STRATEGY, Oncotarget, Vol. 9, (No. 70), pp: 33416-33439 (2018)). Therefore, there is strong evidence to support the role of IRAK1 in a variety of inflammatory conditions.

SUMMARY OF THE INVENTION

Provided herein are compounds and pharmaceutical compositions useful as inhibitors of IRAK4 or IRAK1 or both IRAK4 and IRAK1. Some compounds of the disclosure may find use in pharmaceutical compositions, together with at least one pharmaceutically acceptable excipient, for treating a subject in need thereof. Compounds of the present disclosure also have been found to inhibit production of pro-inflammatory cytokines TNFα, IL-6, IL-1β, IL-8, IL-12, IL-23 and type I interferons IFNα and IFNβ, all of which are mediators of inflammation and the immune response. The disclosure also provides compositions, including pharmaceutical compositions, kits that include the compounds, and methods of using and making the compounds.

In one embodiment of the disclosure, there is provided a compound of Formula (I):

In one embodiment, “Het” is selected from:

In one embodiment, R1is C1-10alkyl optionally substituted with Z1.

In still another embodiment, R1is C3-10cycloalkyl substituted with 5-10 membered heteroaryl wherein said 5-10 membered heteroaryl is optionally substituted with Z1a.

In another embodiment, R1is C3-10cycloalkyl substituted with C1-3alkyl and said C1-3alkyl is further substituted with Z1a.

In one embodiment, R2is C1-10alkyl optionally substituted with Z1.

In another embodiment, R2is C1-10alkyl optionally substituted with one or more —F, —OH or combinations thereof.

In another embodiment, R2is a 4-8 membered heterocyclyl optionally substituted with Z1I.

The disclosure also provides a compound of Formula (Ia):

In another embodiment the disclosure provides a pharmaceutical composition comprising a compound of Formula (I) or (Ia) or a pharmaceutically acceptable salt, stereoisomer, mixture of stereoisomers or deuterated analog thereof, together with a pharmaceutically acceptable carrier.

In still other embodiments the disclosure provides a method of treating an inflammatory condition in a patient in need thereof, comprising administering to said patient a compound of Formula (I) or (Ia) or the composition comprising a compound of Formula (I) or (Ia).

In some embodiments the inflammatory condition is selected from IBD, SLE, Psoriasis and Rheumatoid Arthritis.

DETAILED DESCRIPTION OF THE INVENTION

Definitions

A dash (“-”) that is not between two letters or symbols is used to indicate a point of attachment for a substituent. For example, —C(O)NH2is attached through the carbon atom. A dash at the front or end of a chemical group is a matter of convenience; chemical groups may be depicted with or without one or more dashes without losing their ordinary meaning. A wavy line drawn through a line in a structure indicates a point of attachment of a group. Unless chemically or structurally required, no directionality is indicated or implied by the order in which a chemical group is written or named.

The prefix “Cu-v” indicates that the following group has from u to v carbon atoms. For example, “C1-6alkyl” indicates that the alkyl group has from 1 to 6 carbon atoms.

Reference to “about” a value or parameter herein includes (and describes) embodiments that are directed to that value or parameter per se. In certain embodiments, the term “about” includes the indicated amount ±10%. In other embodiments, the term “about” includes the indicated amount ±5%. In certain other embodiments, the term “about” includes the indicated amount ±1%. Also, to the term “about X” includes description of “X”. Also, the singular forms “a” and “the” include plural references unless the context clearly dictates otherwise. Thus, e.g., reference to “the compound” includes a plurality of such compounds and reference to “the assay” includes reference to one or more assays and equivalents thereof known to those skilled in the art.

“Alkenyl” refers to an alkyl group containing at least one carbon-carbon double bond and having from 2 to 20 carbon atoms (i.e., C2-20alkenyl), 2 to 8 carbon atoms (i.e., C2alkenyl), 2 to 6 carbon atoms (i.e., C2-6alkenyl), or 2 to 4 carbon atoms (i.e., C2-4alkenyl). Examples of alkenyl groups include ethenyl, propenyl, butadienyl (including 1,2-butadienyl and 1,3-butadienyl).

“Alkynyl” refers to an alkyl group containing at least one carbon-carbon triple bond and having from 2 to 20 carbon atoms (i.e., C2-20alkynyl), 2 to 8 carbon atoms (i.e., C2-8alkynyl), 2 to 6 carbon atoms (i.e., C2-6alkynyl), or 2 to 4 carbon atoms (i.e., C2-4alkynyl). The term “alkynyl” also includes those groups having one triple bond and one double bond.

“Haloalkoxy” refers to an alkoxy group as defined above, wherein one or more hydrogen atoms are replaced by a halogen.

“Alkylthio” refers to the group “alkyl-S—”.

“Amino” refers to the group —NRyRywherein each Ryis independently selected from the group consisting of hydrogen, alkyl, alkenyl, alkynyl, aryl, heterocyclyl, cycloalkyl or heteroaryl, each of which is optionally substituted, as defined herein.

“Aryl” refers to an aromatic carbocyclic group having a single ring (e.g., monocyclic) or multiple rings (e.g., bicyclic or tricyclic) including fused systems. As used herein, aryl has 6 to 20 ring carbon atoms (i.e., C6-20aryl), 6 to 12 carbon ring atoms (i.e., C6-12aryl), or 6 to 10 carbon ring atoms (i.e., C6-10aryl). Examples of aryl groups include phenyl, naphthyl, fluorenyl, and anthryl. Aryl, however, does not encompass or overlap in any way with heteroaryl defined below. If one or more aryl groups are fused with a heteroaryl, the resulting ring system is heteroaryl. If one or more aryl groups are fused with a heterocyclyl, the resulting ring system is heterocyclyl.

“Cyano” refers to the group —CN.

“Keto” or “oxo” refers to a group ═O.

“Carbamoyl” refers to both an “O-carbamoyl” group which refers to the group —O—C(O)NRyRzand an “N-carbamoyl” group which refers to the group —NRyC(O)ORz, wherein Ryand Rzare independently selected from the group consisting of hydrogen, alkyl, aryl, haloalkyl, or heteroaryl; each of which may be optionally substituted.

“Ester” refers to both —OC(O)R and —C(O)OR, wherein R is a substituent; each of which may be optionally substituted, as defined herein.

“Cycloalkyl” refers to a saturated or partially unsaturated cyclic alkyl group having a single ring or multiple rings including fused, bridged, and spiro ring systems. The term “cycloalkyl” includes cycloalkenyl groups (i.e., the cyclic group having at least one double bond). As used herein, cycloalkyl has from 3 to 20 ring carbon atoms (i.e., C3-20cycloalkyl), 3 to 12 ring carbon atoms (i.e., C3-12cycloalkyl), 3 to 10 ring carbon atoms (i.e., C3-10cycloalkyl), 3 to 8 ring carbon atoms (i.e., C3-8cycloalkyl), or 3 to 6 ring carbon atoms (i.e., C3-6cycloalkyl). Examples of cycloalkyl groups include cyclopropyl, cyclobutyl, cyclopentyl, and cyclohexyl.

“Halogen” or “halo” includes fluoro, chloro, bromo, and iodo. “Haloalkyl” refers to an unbranched or branched alkyl group as defined above, wherein one or more hydrogen atoms are replaced by a halogen. For example, where a residue is substituted with more than one halogen, it may be referred to by using a prefix corresponding to the number of halogen moieties attached. Dihaloalkyl and trihaloalkyl refer to alkyl substituted with two (“di”) or three (“tri”) halo groups, which may be, but are not necessarily, the same halogen. Examples of haloalkyl include difluoromethyl (—CHF2) and trifluoromethyl (—CF3).

“Heteroalkyl” refers to an alkyl group in which one or more of the carbon atoms (and any associated hydrogen atoms) are each independently replaced with the same or different heteroatomic group. The term “heteroalkyl” includes unbranched or branched saturated chain having carbon and heteroatoms. By way of example, 1, 2 or 3 carbon atoms may be independently replaced with the same or different heteroatomic group. Heteroatomic groups include, but are not limited to, —NR—, —O—, —S—, —S(O)—, —S(O)2—, and the like, where R is H, alkyl, aryl, cycloalkyl, heteroalkyl, heteroaryl or heterocyclyl, each of which may be optionally substituted. Examples of heteroalkyl groups include —OCH3, —CH2OCH3, —SCH3, —CH2SCH3, —NRCH3, and —CH2NRCH3, where R is hydrogen, alkyl, aryl, arylalkyl, heteroalkyl, or heteroaryl, each of which may be optionally substituted. As used herein, heteroalkyl include 1 to 10 carbon atoms, 1 to 8 carbon atoms, or 1 to 4 carbon atoms; and 1 to 3 heteroatoms, 1 to 2 heteroatoms, or 1 heteroatom.

“Heteroaryl” refers to an aromatic group having a single ring, multiple rings, or multiple fused rings, with one or more ring heteroatoms independently selected from nitrogen, oxygen, and sulfur. As used herein, heteroaryl includes 1 to 20 ring carbon atoms (i.e., C1-20heteroaryl), 3 to 12 ring carbon atoms (i.e., C3-12heteroaryl), or 3 to 8 carbon ring atoms (i.e., C3-8heteroaryl); and 1 to 5 heteroatoms, 1 to 4 heteroatoms, 1 to 3 ring heteroatoms, 1 to 2 ring heteroatoms, or 1 ring heteroatom independently selected from nitrogen, oxygen, and sulfur. Examples of heteroaryl groups include pyrimidinyl, purinyl, pyridyl, pyridazinyl, benzothiazolyl, and pyrazolyl. Examples of the fused-heteroaryl rings include, but are not limited to, benzo[d]thiazolyl, quinolinyl, isoquinolinyl, benzo[b]thiophenyl, indazolyl, benzo[d]imidazolyl, pyrazolo[1,5-a]pyridinyl, and imidazo[1,5-a]pyridinyl, where the heteroaryl can be bound via either ring of the fused system. Any aromatic ring, having a single or multiple fused rings, containing at least one heteroatom, is considered a heteroaryl regardless of the attachment to the remainder of the molecule (i.e., through any one of the fused rings). Heteroaryl does not encompass or overlap with aryl as defined above.

“Heterocyclyl” refers to a saturated or unsaturated cyclic alkyl group, with one or more ring heteroatoms independently selected from nitrogen, oxygen and sulfur. The term “heterocyclyl” includes heterocycloalkenyl groups (i.e., the heterocyclyl group having at least one double bond), bicyclic heterocyclyl groups, bridged-heterocyclyl groups, fused-heterocyclyl groups, and spiro-heterocyclyl groups. A heterocyclyl may be a single ring or multiple rings wherein the multiple rings may be fused, bridged, or spiro. Any non-aromatic ring containing at least one heteroatom is considered a heterocyclyl, regardless of the attachment (i.e., can be bound through a carbon atom or a heteroatom). Further, the term heterocyclyl is intended to encompass any non-aromatic ring containing at least one heteroatom, which ring may be fused to an aryl or heteroaryl ring, regardless of the attachment to the remainder of the molecule. As used herein, heterocyclyl has 2 to 20 ring atoms (i.e., 4-20 membered heterocyclyl), 2 to ring atoms (i.e., 4-12 membered heterocyclyl), 4 to 10 ring atoms (i.e., 4-10 membered heterocyclyl), 4 to 8 ring atoms (i.e., 4-8 membered heterocyclyl), or 4 to 6 ring carbon atoms (i.e., 4-6 membered heterocyclyl); having 1 to 5 ring heteroatoms, 1 to 4 ring heteroatoms, 1 to 3 ring heteroatoms, 1 to 2 ring heteroatoms, or 1 ring heteroatom independently selected from nitrogen, sulfur or oxygen. A heterocyclyl may contain one or more oxo and/or thioxo groups. Examples of heterocyclyl groups include pyrrolidinyl, piperidinyl, piperazinyl, oxetanyl, dioxolanyl, azetidinyl, azetidinyl, morpholinyl, thiomorpholinyl, 4-7 membered sultam, 4-7 membered cyclic carbamate, 4-7 membered cyclic carbonate, 4-7 membered cyclic sulfide and morpholinyl. As used herein, the term “bridged-heterocyclyl” refers to a four- to ten-membered cyclic moiety connected at two non-adjacent atoms of the heterocyclyl with one or more (e.g., 1 or 2) four- to ten-membered cyclic moiety having at least one heteroatom where each heteroatom is independently selected from nitrogen, oxygen, and sulfur. As used herein, bridged-heterocyclyl includes bicyclic and tricyclic ring systems. Also used herein, the term “spiro-heterocyclyl” refers to a ring system in which a three- to ten-membered heterocyclyl has one or more additional ring, wherein the one or more additional ring is three- to ten-membered cycloalkyl or three- to ten-membered heterocyclyl, where a single atom of the one or more additional ring is also an atom of the three- to ten-membered heterocyclyl. Examples of the spiro-heterocyclyl rings include bicyclic and tricyclic ring systems, such as 2-oxa-7-azaspiro[3.5]nonanyl, 2-oxa-6-azaspiro[3.4]octanyl, and 6-oxa-1-azaspiro[3.3]heptanyl. Examples of the fused-heterocyclyl rings include, but are not limited to, 1,2,3,4-tetrahydroisoquinolinyl, 1-oxo-1,2,3,4-tetrahydroisoquinolinyl, 1-oxo-1,2-dihydroisoquinolinyl, 4,5,6,7-tetrahydrothieno[2,3-c]pyridinyl, indolinyl, and isoindolinyl, where the heterocyclyl can be bound via either ring of the fused system. As used herein, a bicyclic heterocyclyl group is a heterocyclyl group attached at two points to another cyclic group, wherein the other cyclic group may itself be a heterocyclic group, or a carbocyclic group.

As used herein, the term “nitrogen or sulfur containing heterocyclyl” means a heterocyclyl moiety that contains at least one nitrogen atom or at least one sulfur atom, or both a nitrogen atom and a sulfur atom within the ring structure. It is to be understood that other heteroatoms, including oxygen, may be present in addition to the nitrogen, sulfur, or combinations thereof. Examples of nitrogen or sulfur containing heterocyclyls include morpholinyl, thiomorpholinyl, thiazolyl, isothiazolyl, oxazolidinone 1,2 dithiolyl, piperidinyl, piperazinyl, and the like.

“Hydroxy” or “hydroxyl” refers to the group —OH. “Hydroxyalkyl” refers to an unbranched or branched alkyl group as defined above, wherein one or more hydrogen atoms are replaced by a hydroxyl.

“Nitro” refers to the group —NO2.

“Imino” refers to a group ═N—Ry, or ═N—C(O)Ry, wherein Ryis selected from the group consisting of hydrogen, alkyl, aryl, cyano, haloalkyl, or heteroaryl; each of which may be optionally substituted.

“Sulfoximine” or “sulfoximino” refers to a substituted or unsubstituted moiety of the general formula

wherein Ryis selected from the group consisting of hydrogen, alkyl, amino, aryl, cyano, haloalkyl, heterocyclyl, or heteroaryl; V and W are each independently selected from a bond, alkyl, amino, aryl, haloalkyl, heterocyclyl or heteroaryl; each of which may be optionally substituted and wherein Ryand V, Ryand W, and V and W together with the atoms to which they are attached may be joined together to form a ring.

“Sulfonyl” refers to the group —S(O)2R, where R is a substituent, or a defined group.

“Alkylsulfonyl” refers to the group —S(O)2R, where R is a substituent, or a defined group.

“Alkylsulfinyl” refers to the group —S(O)R, where R is a substituent, or a defined group.

“Thiol” refers to the group —SR, where R is a substituent, or a defined group.

“Thioxo” or “thione” refer to the group (═S) or (S).

Certain commonly used alternative chemical names may be used. For example, a divalent group such as a divalent “alkyl” group, a divalent “aryl” group, etc., may also be referred to as an “alkylene” group or an “alkylenyl” group, an “arylene” group or an “arylenyl” group, respectively. Also, unless indicated explicitly otherwise, where combinations of groups are referred to herein as one moiety, e.g., arylalkyl, the last mentioned group contains the atom by which the moiety is attached to the rest of the molecule.

The terms “optional” or “optionally” means that the subsequently described event or circumstance may or may not occur, and that the description includes instances where said event or circumstance occurs and instances in which it does not. Also, the term “optionally substituted” refers to any one or more hydrogen atoms on the designated atom or group may or may not be replaced by a moiety other than hydrogen. “Optionally substituted” may be zero to the maximum number of possible substitutions, and each occurance is independent. When the term “substituted” is used, then that substitution is required to be made at a substitutable hydrogen atom of the indicated substituent. An optional substitution may be the same or different from a (required) substitution.

When a moiety is “optionally substituted,” and reference is made to a general term, such as any “alkyl,” “alkenyl,” “alkynyl,” “haloalkyl,” “cycloalkyl,” “aryl” or “heteroaryl,” then the general term can refer to any antecedent specifically recited term, such as (C1-3alkyl), (C4-6alkyl), —O(C1-4alkyl), (C3-10cycloalkyl), O—(C3-10cycloalkyl) and the like. For example, “any aryl” includes both “aryl” and “—O(aryl) as well as examples of aryl, such as phenyl or naphthyl and the like. Also, the term “any heterocyclyl” includes both the terms “heterocyclyl” and O-(heterocyclyl),” as well as examples of heterocyclyls, such as oxetanyl, tetrahydropyranyl, morpholino, piperidinyl and the like. In the same manner, the term “any heteroaryl” includes the terms “heteroaryl” and “O-(heteroryl),” as well as specific heteroaryls, such as pyridine and the like.

Some of the compounds exist as tautomers. Tautomers are in equilibrium with one another. For example, amide containing compounds may exist in equilibrium with imidic acid tautomers. Regardless of which tautomer is shown, and regardless of the nature of the equilibrium among tautomers, the compounds are understood by one of ordinary skill in the art to comprise both amide and imidic acid tautomers. Thus, the amide containing compounds are understood to include their imidic acid tautomers. Likewise, the imidic acid containing compounds are understood to include their amide tautomers.

Any formula or structure given herein, is also intended to represent unlabeled forms as well as isotopically labeled forms of the compounds. Isotopically labeled compounds have structures depicted by the formulas given herein except that one or more atoms are replaced by an atom having a selected atomic mass or mass number. Examples of isotopes that can be incorporated into compounds of the disclosure include isotopes of hydrogen, carbon, nitrogen, oxygen, phosphorous, fluorine and chlorine, such as, but not limited to2H (deuterium, D),3H (tritium),11C,13C,14C,15N,18F,31P,32P,35S,36Cl and125I. Various isotopically labeled compounds of the present disclosure, for example those into which radioactive isotopes such as13H,13C and14C are incorporated. Such isotopically labelled compounds may be useful in metabolic studies, reaction kinetic studies, detection or imaging techniques, such as positron emission tomography (PET) or single-photon emission computed tomography (SPECT) including drug or substrate tissue distribution assays or in radioactive treatment of patients.

The disclosure also includes “deuterated analogues” of compounds of Formula I in which from 1 to n hydrogens attached to a carbon atom is/are replaced by deuterium, in which n is the number of hydrogens in the molecule. Such compounds exhibit increased resistance to metabolism and are thus useful for increasing the half-life of any compound of Formula I when administered to a mammal, particularly a human. See, for example, Foster, “Deuterium Isotope Effects in Studies of Drug Metabolism,” Trends Pharmacol. Sci. 5(12):524-527 (1984). Such compounds are synthesized by means well known in the art, for example by employing starting materials in which one or more hydrogens have been replaced by deuterium.

Deuterium labelled or substituted therapeutic compounds of the disclosure may have improved DMPK (drug metabolism and pharmacokinetics) properties, relating to distribution, metabolism and excretion (ADME). Substitution with heavier isotopes such as deuterium may afford certain therapeutic advantages resulting from greater metabolic stability, for example increased in vivo half-life, reduced dosage requirements and/or an improvement in therapeutic index. An18F labeled compound may be useful for PET or SPECT studies. Isotopically labeled compounds of this disclosure and prodrugs thereof can generally be prepared by carrying out the procedures disclosed in the schemes or in the examples and preparations described below by substituting a readily available isotopically labeled reagent for a non-isotopically labeled reagent. It is understood that deuterium in this context is regarded as a substituent in the compound of Formula I.

The concentration of such a heavier isotope, specifically deuterium, may be defined by an isotopic enrichment factor. In the compounds of this disclosure any atom not specifically designated as a particular isotope is meant to represent any stable isotope of that atom. Unless otherwise stated, when a position is designated specifically as “H” or “hydrogen”, the position is understood to have hydrogen at its natural abundance isotopic composition. Accordingly, in the compounds of this disclosure any atom specifically designated as a deuterium (D) is meant to represent deuterium.

In many cases, the compounds of this disclosure are capable of forming acid and/or base salts by virtue of the presence of amino and/or carboxyl groups or groups similar thereto.

Provided are also pharmaceutically acceptable salts, hydrates, solvates, tautomeric forms, polymorphs, and prodrugs of the compounds described herein. “Pharmaceutically acceptable” or “physiologically acceptable” refer to compounds, salts, compositions, dosage forms and other materials which are useful in preparing a pharmaceutical composition that is suitable for veterinary or human pharmaceutical use.

The term “pharmaceutically acceptable salt” of a given compound refers to salts that retain the biological effectiveness and properties of the given compound, and which are not biologically or otherwise undesirable. “Pharmaceutically acceptable salts” or “physiologically acceptable salts” include, for example, salts with inorganic acids and salts with an organic acid. In addition, if the compounds described herein are obtained as an acid addition salt, the free base can be obtained by basifying a solution of the acid salt. Conversely, if the product is a free base, an addition salt, particularly a pharmaceutically acceptable addition salt, may be produced by dissolving the free base in a suitable organic solvent and treating the solution with an acid, in accordance with conventional procedures for preparing acid addition salts from base compounds. Those skilled in the art will recognize various synthetic methodologies that may be used to prepare nontoxic pharmaceutically acceptable addition salts. Pharmaceutically acceptable acid addition salts may be prepared from inorganic and organic acids. Salts derived from inorganic acids include hydrochloric acid, hydrobromic acid, sulfuric acid, nitric acid, phosphoric acid, and the like. Salts derived from organic acids include acetic acid, propionic acid, glycolic acid, pyruvic acid, oxalic acid, malic acid, malonic acid, succinic acid, maleic acid, fumaric acid, tartaric acid, citric acid, benzoic acid, cinnamic acid, mandelic acid, methanesulfonic acid, ethanesulfonic acid, p-toluene-sulfonic acid, salicylic acid, and the like. Likewise, pharmaceutically acceptable base addition salts can be prepared from inorganic and organic bases. Salts derived from inorganic bases include, by way of example only, sodium, potassium, lithium, ammonium, calcium and magnesium salts. Salts derived from organic bases include, but are not limited to, salts of primary, secondary and tertiary amines, such as alkyl amines (i.e., NH2(alkyl)), dialkyl amines (i.e., HN(alkyl)2), trialkyl amines (i.e., N(alkyl)3), substituted alkyl amines (i.e., NH2(substituted alkyl)), di(substituted alkyl) amines (i.e., HN(substituted alkyl)2), tri(substituted alkyl) amines (i.e., N(substituted alkyl)3), alkenyl amines (i.e., NH2(alkenyl)), dialkenyl amines (i.e., HN(alkenyl)2), trialkenyl amines (i.e., N(alkenyl)3), substituted alkenyl amines (i.e., NH2(substituted alkenyl)), di(substituted alkenyl) amines (i.e., HN(substituted alkenyl)2), tri(substituted alkenyl) amines (i.e., N(substituted alkenyl)3, mono-, di- or tri-cycloalkyl amines (i.e., NH2(cycloalkyl), HN(cycloalkyl)2, N(cycloalkyl)3), mono-, di- or tri-arylamines (i.e., NH2(aryl), HN(aryl)2, N(aryl)3), or mixed amines, etc. Specific examples of suitable amines include, by way of example only, isopropylamine, trimethyl amine, diethyl amine, tri(iso-propyl) amine, tri(n-propyl) amine, ethanolamine, 2-dimethylaminoethanol, piperazine, piperidine, morpholine, N-ethylpiperidine, and the like.

The term “substituted” means that any one or more hydrogen atoms on the designated atom or group is replaced with one or more substituents other than hydrogen, provided that the designated atom's normal valence is not exceeded. The one or more substituents include, but are not limited to, alkyl, alkenyl, alkynyl, alkoxy, acyl, amino, amido, amidino, aryl, azido, carbamoyl, carboxyl, carboxyl ester, cyano, guanidino, halo, haloalkyl, haloalkoxy, heteroalkyl, heteroaryl, heterocyclyl, hydroxy, hydrazino, imino, oxo, nitro, alkylsulfinyl, sulfonic acid, alkylsulfonyl, thiocyanate, thiol, thione, or combinations thereof. Polymers or similar indefinite structures arrived at by defining substituents with further substituents appended ad infinitum (e.g., a substituted aryl having a substituted alkyl which is itself substituted with a substituted aryl group, which is further substituted by a substituted heteroalkyl group, etc.) are not intended for inclusion herein. Unless otherwise noted, the maximum number of serial substitutions in compounds described herein is three. For example, serial substitutions of substituted aryl groups with two other substituted aryl groups are limited to ((substituted aryl)substituted aryl) substituted aryl. Similarly, the above definitions are not intended to include impermissible substitution patterns (e.g., methyl substituted with 5 fluorines or heteroaryl groups having two adjacent oxygen ring atoms). Such impermissible substitution patterns are well known to the skilled artisan. When used to modify a chemical group, the term “substituted” may describe other chemical groups defined herein. Unless specified otherwise, where a group is described as optionally substituted, any substituents of the group are themselves unsubstituted. For example, in some embodiments, the term “substituted alkyl” refers to an alkyl group having one or more substituents including hydroxyl, halo, alkoxy, cycloalkyl, heterocyclyl, aryl, and heteroaryl. In other embodiments, the one or more substituents may be further substituted with halo, alkyl, haloalkyl, hydroxyl, alkoxy, cycloalkyl, heterocyclyl, aryl, or heteroaryl, each of which is substituted. In other embodiments, the substituents may be further substituted with halo, alkyl, haloalkyl, alkoxy, hydroxyl, cycloalkyl, heterocyclyl, aryl, or heteroaryl, each of which is unsubstituted One skilled in the art will recognize that substituents and other moieties of the compounds of the generic formula herein should be selected in order to provide a compound which is sufficiently stable to provide a pharmaceutically useful compound which can be formulated into an acceptably stable pharmaceutical composition. Compounds which have such stability are contemplated as falling within the scope of the present invention. It should be understood by one skilled in the art that any combination of the definitions and substituents described above should not result in an inoperable species or compound.

A “solvate” is formed by the interaction of a solvent and a compound. Solvates of salts of the compounds described herein are also provided. Hydrates of the compounds described herein are also provided.

Combinations

Patients being treated by administration of the IRAK4 inhibitors of the disclosure often exhibit diseases or conditions that benefit from treatment with other therapeutic agents. These diseases or conditions can be of an inflammatory nature or can be related to cancer, metabolic disorders, gastrointestinal disorders and the like. Thus, one aspect of the disclosure is a method of treating an inflammation related disease or condition, or a metabolic disorder, gastrointestinal disorder, or cancer and the like comprising administering a compound of the in combination with one or more compounds useful for the treatment of such diseases to a subject, particularly a human subject, in need thereof.

In some embodiments, a compound of the present disclosure is co-formulated with the additional one or more active ingredients. In some embodiments, the other active ingredient is administered at approximately the same time, in a separate dosage form. In some embodiments, the other active ingredient is administered sequentially, and may be administered at different times in relation to a compound of the present disclosure.

Combinations for Inflammatory Diseases and Conditions

Combinations for Metabolic Diseases or Conditions

Examples of metabolic disorders include, without limitation, diabetes, including type I and type II diabetes, metabolic syndrome, dyslipidemia, obesity, glucose intolerance, hypertension, elevated serum cholesterol, and elevated triglycerides. Examples of therapeutic agents used to treat metabolic disorders include antihypertensive agents and lipid lowering agents. Additional therapeutic agents used to treat metabolic disorders include insulin, sulfonylureas peroxisome proliferator activated receptor gamma (PPAR-γ) agonists, such as thiazolidinediones such as pioglitazones, biguanides, alpha-glucosidase inhibitors, Vitamin E and incretin mimetics. Thus, one aspect of the disclosure is a method of treating a metabolic disease comprising administering a compound of the disclosure in combination with one or more compounds useful for the treatment of metabolic diseases to a subject, particularly a human subject, in need thereof.

Pharmaceutical Compositions

While it is possible for the active ingredients to be administered alone it may be preferable to present them as pharmaceutical formulations (compositions). The formulations, both for veterinary and for human use, of the invention comprise at least one active ingredient, as above defined, together with one or more acceptable carriers therefor and optionally other therapeutic ingredients. The carrier(s) must be “acceptable” in the sense of being compatible with the other ingredients of the formulation and physiologically innocuous to the recipient thereof.

In certain embodiments, formulations suitable for oral administration are presented as discrete units such as capsules, cachets or tablets each containing a predetermined amount of the active ingredient.

The amount of active ingredient that is combined with the inactive ingredients to produce a dosage form will vary depending upon the host treated and the particular mode of administration. For example, in some embodiments, a dosage form for oral administration to humans contains approximately 1 to 1000 mg of active material formulated with an appropriate and convenient amount of carrier material (e.g., inactive ingredient or excipient material). In certain embodiments, the carrier material varies from about 5 to about 95% of the total compositions (weight:weight). In some embodiments, the pharmaceutical compositions described herein contain about 1 to 800 mg, 1 to 600 mg, 1 to 400 mg, 1 to 200 mg, 1 to 100 mg or 1 to 50 mg of the compound of Formula I, or a pharmaceutically acceptable salt thereof. In some embodiments, the pharmaceutical compositions described herein contain not more than about 400 mg of the compound of Formula I. In some embodiments, the pharmaceutical compositions described herein contain about 100 mg of the compound of Formula I, or a pharmaceutically acceptable salt thereof.

Veterinary compositions comprising at least one active ingredient as above defined together with a veterinary carrier are further provided.

Veterinary carriers are materials useful for the purpose of administering the composition and may be solid, liquid or gaseous materials which are otherwise inert or acceptable in the veterinary art and are compatible with the active ingredient. These veterinary compositions may be administered orally, parenterally or by any other desired route.

Effective dose of active ingredient depends at least on the nature of the condition being treated, toxicity, whether the compound is being used prophylactically (lower doses), the method of delivery, and the pharmaceutical formulation, and will be determined by the clinician using conventional dose escalation studies.

Routes of Administration

One or more compounds of Formula I (herein referred to as the active ingredients), or a pharmaceutically acceptable salt thereof, are administered by any route appropriate to the condition to be treated. Suitable routes include oral, rectal, nasal, topical (including buccal and sublingual), vaginal and parenteral (including subcutaneous, intramuscular, intravenous, intradermal, intrathecal and epidural), and the like. It will be appreciated that the preferred route may vary with for example the condition of the recipient. An advantage of the compounds of this invention is that they are orally bioavailable and can be dosed orally. Accordingly, in one embodiment, the pharmaceutical compositions described herein are oral dosage forms. In certain embodiments, the pharmaceutical compositions described herein are oral solid dosage forms.

Formulation Example 1

Hard gelatin capsules containing the following ingredients are prepared:

The above ingredients are mixed and filled into hard gelatin capsules.

Formulation Example 2

A tablet Formula is prepared using the ingredients below:

The components are blended and compressed to form tablets.

Formulation Example 3

A dry powder inhaler formulation is prepared containing the following components:

The active ingredient is mixed with the lactose and the mixture is added to a dry powder inhaling appliance.

Formulation Example 4

Tablets, each containing 30 mg of active ingredient, are prepared as follows:

The active ingredient, starch and cellulose are passed through a No. 20 mesh U.S. sieve and mixed thoroughly. The solution of polyvinylpyrrolidone is mixed with the resultant powders, which are then passed through a 16 mesh U.S. sieve. The granules so produced are dried at 50° C. to 60° C. and passed through a 16 mesh U.S. sieve. The sodium carboxymethyl starch, magnesium stearate and talc, previously passed through a No. 30 mesh U.S. sieve, are then added to the granules which, after mixing, are compressed on a tablet machine to yield tablets each weighing 120 mg.

Formulation Example 5

Suppositories, each containing 25 mg of active ingredient are made as follows:

The active ingredient is passed through a No. 60 mesh U.S. sieve and suspended in the saturated fatty acid glycerides previously melted using the minimum heat necessary. The mixture is then poured into a suppository mold of nominal 2.0 g capacity and allowed to cool.

Formulation Example 6

Suspensions, each containing 50 mg of active ingredient per 5.0 mL dose are made as follows:

The active ingredient, sucrose and xanthan gum are blended, passed through a No. 10 mesh U.S. sieve and then mixed with a previously made solution of the microcrystalline cellulose and sodium carboxymethyl cellulose in water. The sodium benzoate, flavor and color are diluted with some of the water and added with stirring. Sufficient water is then added to produce the required volume.

Formulation Example 7

A subcutaneous formulation may be prepared as follows:

Formulation Example 8

An injectable preparation is prepared having the following composition:

Formulation Example 9

A topical preparation is prepared having the following composition:

All of the above ingredients, except water, are combined and heated to 60° C. with stirring.

A sufficient quantity of water at 60° C. is then added with vigorous stirring to emulsify the ingredients and water then added q.s. 100 g.

Formulation Example 10

Sustained Release Composition

Sustained release formulations of this disclosure may be prepared as follows: compound and pH-dependent binder and any optional excipients are intimately mixed(dry-blended). The dry-blended mixture is then granulated in the presence of an aqueous solution of a strong base which is sprayed into the blended powder. The granulate is dried, screened, mixed with optional lubricants (such as talc or magnesium stearate) and compressed into tablets. Preferred aqueous solutions of strong bases are solutions of alkali metal hydroxides, such as sodium or potassium hydroxide, preferably sodium hydroxide, in water (optionally containing up to 25% of water-miscible solvents such as lower alcohols).

The resulting tablets may be coated with an optional film-forming agent, for identification, taste-masking purposes and to improve ease of swallowing. The film forming agent will typically be present in an amount ranging from between 2% and 4% of the tablet weight. Suitable film-forming agents are well known to the art and include hydroxypropyl methylcellulose, cationic methacrylate copolymers (dimethylaminoethyl methacrylate/methyl-butyl methacrylate copolymers—Eudragito® E—Röhm. Pharma) and the like. These film-forming agents may optionally contain colorants, plasticizers and other supplemental ingredients.

The compressed tablets preferably have a hardness sufficient to withstand 8 Kp compression. The tablet size will depend primarily upon the amount of compound in the tablet. The tablets will include from 300 to 1100 mg of compound free base. Preferably, the tablets will include amounts of compound free base ranging from 400-600 mg, 650-850 mg and 900-1100 mg.

In order to influence the dissolution rate, the time during which the compound containing powder is wet mixed is controlled. Preferably the total powder mix time, i.e., the time during which the powder is exposed to sodium hydroxide solution, will range from 1 to 10 minutes and preferably from 2 to 5 minutes. Following granulation, the particles are removed from the granulator and placed in a fluid bed dryer for drying at about 60° C.

Formulation Example 11

A tablet Formula Is prepared using the ingredients below:

The components are blended and compressed to form tablets.

EXAMPLES

List of Abbreviations and Acronyms

EXPERIMENTAL PROCEDURES

General Schemes

The compounds of formula 1.5 may be accessed according to the method outlined in Scheme 1. 1-aminopyrrole 1.1 may be condensed with a suitable coupling partner to produce substituted pyrrolo[1,2-b]pyridazine 1.2 using a suitable catalyst (e.g., HCl, etc.) and suitable solvent (e.g., EtOH, etc.). Halogenation at the position shown using a known halogenating reagent (e.g., NBS, etc.) can form the intermediate 1.3, which can be further substituted either via C—H activation or electrophilic aromatic substitution with a suitable reagent (e.g., selectfluor, etc.) to produce intermediate 1.4. Halogen metal exchange of —X to -M can then be achieved using a suitable reagent (e.g., n-BuLi, etc.) or transition metal coupling using a palladium catalyst and metal source (e.g., B2Pin2, Me6Sn2, etc.) to give intermediate 1.5.

The compounds of the formula 2.3 may be accessed according to the method outlined in Scheme 2. The acid 2.1 can be converted to the corresponding acyl hydrazine using a coupling reagent (e.g., HATU, etc.) in the presence of a base (e.g., DIPEA, etc.). Cyclization of compound 2.2 can be accomplished by heating in the presence of a thionating reagent (e.g., Lawesson's reagent, etc.) to provide compound 2.3.

The compounds of formula 3.6 may be accessed according to the method outlined in Scheme 3. Dihalopyridine 3.1 may be converted to compound 3.2 via displacement of one of the halogen groups (e.g., nucleophilic aromatic substitution, etc.). Further functionalization of compound 3.2 using a metal-containing heterocyclic species (e.g., compound 1.5) with a suitable catalyst, such as a palladium catalyst, can afford compound 3.3. Halogenation at the position shown using a known halogenating reagent (e.g., NBS, etc.) can form the intermediate 3.4 which can be further substituted through a cross-coupling reaction using a suitable catalyst, such as a palladium catalyst, to provide compound 3.5.

Compounds of formula 4.2 may be assembled following Scheme 4. Displacement of the halogen group (e.g., nucleophilic aromatic substitution, etc.) of a halothiadiazole 4.1 with a nucleophile (e.g., an amine, etc.) can provide compound 2.3. Halogenation at the position shown using a known halogenating reagent (e.g., NBS, etc.) can form the intermediate 4.2.

Compounds of formula 3.5 may also be assembled following Scheme 5. Halogen metal exchange of —X to -M can then be achieved using a suitable reagent (e.g., n-BuLi, etc.) or transition metal coupling using a palladium catalyst and metal source (e.g., B2Pin2, Me6Sn2, etc.) to give intermediate 5.1. Functionalization of compound 5.1 can be done utilizing a cross-coupling reaction with compound 4.2 using a suitable catalyst, such as a palladium catalyst, to provide compound 3.5.

An alternative method of access compound 3.5 is shown in Scheme 6. Starting from the nicotinic acid 6.1, the corresponding acyl hydrazine can be prepared using a coupling reagent (e.g., HATU, etc.) in the presence of a base (e.g., DIPEA, etc.). Cyclization of compound 6.3 can be accomplished by heating in the presence of a thionating reagent (e.g., Lawesson's reagent, etc.) to provide compound 6.4. Further functionalization of compound 6.4 using a metal-containing heterocyclic species (e.g., compound 1.5) with a suitable catalyst, such as a palladium catalyst, can afford compound 3.5.

It is also noted that synthetic manipulations of the incorporated R groups are possible following their incorporation. A specific illustrative example of an alteration to the R2group is shown in Scheme 7 wherein the secondary carbamate 7.1 is converted the primary amine 7.2. Other functional groups may also be present in the R2and can be manipulated. These groups and manipulations can include, but are not limited to, oxidation, elimination or displacement using suitable reagents known to those skilled in the art. The order of synthetic manipulations may be carried out in a fashion that is consistent with the methods outlined in Schemes 1-6 and should not be limited to the final step of compound preparation.

Synthesis of Intermediates

Preparation of Intermediate I-1

tert-butyl((1r,4r)-4-(2-formylhydrazine-1-carbonyl)cyclohexyl)carbamate: To a solution of (1r,4r)-4-((tert-butoxycarbonyl)amino)cyclohexane-1-carboxylic acid (250 mg, 1.0 mmol) in DMF (2 mL) was added formic acid hydrazide (80 mg, 1.3 mmol), HATU (469 mg, 1.2 mmol), and finally DIPEA (0.45 mL, 2.6 mmol) and the resulting mixture stirred at room temperature for 15 minutes. Upon completion, the reaction mixture was poured into water (5 mL) and extracted with EtOAc (2×15 mL). The combined organic layers were dried over MgSO4, filtered and concentrated. The resulting crude residue was purified by silica gel chromatography (eluent: EtOAc/hexanes) to give the desired product.

tert-butyl((1r,4r)-4-(1,3,4-thiadiazol-2-yl)cyclohexyl)carbamate: To a solution of tert-butyl((1r,4r)-4-(2-formylhydrazine-1-carbonyl)cyclohexyl)carbamate (193 mg, 0.68 mmol) in dioxane (5 mL) was added Lawesson's Reagent (301 mg, 0.74 mmol) and the resulting reaction mixture heated to 100° C. for 3 hours. Upon completion, the reaction mixture was poured into water (5 mL) and extracted with EtOAc (2×15 mL). The combined organic layers were dried over MgSO4, filtered and concentrated. The resulting crude residue was purified by silica gel chromatography (eluent: EtOAc/hexanes) to give the desired product.

(1r,4r)-4-(1,3,4-thiadiazol-2-yl)cyclohexan-1-amine hydrochloride: tert-butyl((1r,4r)-4-(1,3,4-thiadiazol-2-yl)cyclohexyl)carbamate (59 mg, 0.21 mmol) was then dissolved in HCl (4.0M in dioxane, 4 mL, 16 mmol) and stirred at room temperature for 7 hours after which the reaction mixture was concentrated to dryness directly to give the desired product as an HCl salt which was used without further purification.

N-((1r,4r)-4-(1,3,4-thiadiazol-2-yl)cyclohexyl)acetamide (I-1): (1r,4r)-4-(1,3,4-thiadiazol-2-yl)cyclohexan-1-amine hydrochloride (175 mg, 0.8 mmol) was dissolved in CH2Cl2(4 mL) and the reaction mixture was cooled to 0° C. Triethylamine (0.33 mL, 2.39 mmol) was added, followed by acetic anhydride (0.094 mL, 1 mmol). The reaction was stirred at 0° C. for 30 minutes after which the mixture was diluted with CH2Cl2and washed with water. The organic layer was dried over MgSO4, filtered, and concentrated. The crude material was purified by silica gel chromatography (eluent EtOAc/hexanes followed by methanol/EtOAc) to give I-1.

The following intermediates were synthesized as described for I-1 using the appropriate starting carboxylic acid, and appropriate anhydride, carbonyl-chloride, or alkyl triflate:

Preparation of Intermediate I-2

2-bromo-N-isopropylpyridin-4-amine: To a solution of 2-bromo-4-fluoropyridine (1.0 g, 5.68 mmol) in NMP (10 mL) was added isopropylamine (0.8 mL, 12.02 mmol) and N,N-diisopropylethylamine (1.25 mL, 7.18 mmol). The resulting mixture was heated for 30 minutes at 150° C. in a microwave after which the reaction contents were diluted with EtOAc and washed three times with water. The organic layer was dried over MgSO4, filtered and concentrated. The resulting material was purified normal phase SiO2chromatography (eluent: ethyl acetate/hexanes) to provide the desired product.

7-(4-(isopropylamino)pyridin-2-yl)pyrrolo[1,2-b]pyridazine-3-carbonitrile: To a solution of 2-bromo-N-isopropylpyridin-4-amine (1.18 g, 5.14 mmol), (3-cyanopyrrolo[1,2-b]pyridazin-7-yl)boronic acid (2.1 g, 10.34 mmol) (the corresponding pinacol boronic ester is equally competent in this transformation) and Xphos Pd G3 (0.31 g, 0.37 mmol) in DME (12.3 mL) was added aqueous potasium phosphate (2M, 4.9 mL, 9.9 mmol). The resulting solution was degassed with argon for 2 min and heated under microwave conditions for 40 min at 120° C. after which silica gel was added, and the resulting slurry was filtered through celite, rinsing with EtOAc. The material was concentrated, and the resulting crude material was purified by normal phase SiO2chromatography (eluent: ethyl acetate/hexanes) to provide the desired product.

7-(5-bromo-4-(isopropylamino)pyridin-2-yl)pyrrolo[1,2-b]pyridazine-3-carbonitrile (I-2): To a solution of 7-[4-(isopropylamino)-2-pyridyl]pyrrolo[1,2-b]pyridazine-3-carbonitrile (0.75 g, 2.4 mmol) in DCM:MeCN (1:1, 60 mL) at 0° C. was added a solution of N-bromosuccinimide (0.433 g, 2.4 mmol) in 9 mL of 1:1 DCM:MeCN, and the resulting mixture stirred at zero degrees. After 30 minutes, significant precipitate was observed, and the reaction mixture was concentrated under vacuum. The crude material was purified by silica gel chromatography (EtOAc/hexanes) to give I-2.

The following intermediates were prepared as described for I-2, using the appropriate amine for step 1 or boronate ester/boronic acid for step 2

Preparation of Intermediate I-3

The following intermediates were prepared as described for I-3 from the appropriate 3-bromo-pyridines:

Preparation of Intermediate I-4

N-(1-(1,3,4-thiadiazol-2-yl)piperidin-4-yl)acetamide: To a solution of 2-bromo-1,3,4-thiadiazole (100.0 mg, 0.61 mmol) and N-(piperidin-4-yl)acetamide hydrochloride (270.7 mg, 1.5 mmol) in n-butanol (1.2 mL) was added N,N-diisopropylethylamine (0.42 mL, 2.4 mmol). The reaction mixture was heated thermally at 120° C. for 1 hour. The reaction mixture was cooled, concentrated in vacuo to provide the crude product that was used without purification.

N-(1-(5-bromo-1,3,4-thiadiazol-2-yl)piperidin-4-yl)acetamide (1-4): To a solution of crude N-(1-(1,3,4-thiadiazol-2-yl)piperidin-4-yl)acetamide (50.0 mg, 0.22 mmol) in DCM (0.5 mL) and MeCN (0.5 mL) was added N-bromosuccinimide (118.0 mg, 0.66 mmol) in one portion. After stirring at room temperature for 5 minutes, the reaction mixture was concentrated in vacuo and purified by silica gel column chromatography (eluent: MeOH/DCM) to provide I-4.

Preparation of Intermediate I-5

7-(5-bromo-1,3,4-thiadiazol-2-yl)-2-oxa-7-azaspiro[3.5]nonane(I-5):7-(5-bromo-1,3,4-thiadiazol-2-yl)-2-oxa-7-azaspiro[3.5]nonane was prepared as described in Preparation of Intermediate I-4 substituting N-(piperidin-4-yl)acetamide hydrochloride with 2-oxa-7-azaspiro[3.5]nonane.

Preparation of Intermediate I-6

(R)-4-((5-bromo-1,3,4-thiadiazol-2-yl)amino)-3-fluoro-2-methylbutan-2-ol(I-6):(R)-4-((5-bromo-1,3,4-thiadiazol-2-yl)amino)-3-fluoro-2-methylbutan-2-ol was prepared as described in Preparation of Intermediate I-4 substituting N-(piperidin-4-yl)acetamide hydrochloride with (R)-4-amino-3-fluoro-2-methylbutan-2-ol hydrochloride.

Preparation of Intermediate I-7

tert-butyl(2-(1,3,4-thiadiazol-2-yl)ethyl)carbamate (I-7): To a solution of 2-(1,3,4-thiadiazol-2-yl)ethan-1-amine (100.0 mg, 0.77 mmol) and di-tert-butyl dicarbonate (186 mg, 0.85 mmol) in THF at 0° C. was added triethylamine (0.13 mL, 0.93 mmol). The reaction mixture was warmed to room temperature and stirred for 90 minutes. The reaction mixture was concentrated in vacuo and purified by silica gel column chromatography (eluent: MeOH/DCM) to provide I-7.

Preparation of Intermediate I-8

3,3-Diethoxy-2-formylpropionitrile Potassium Salt (I-8C): To a stirred solution of 3,3-diethoxypropane-nitrile (I-8A, 283.80 g, 1.98 moles) and methyl formate (I-8B, 148.80 g, 2.48 moles) in anhydrous THF (1.1 L) at 10° C. was added 1.0 M potassium tert-butoxide in THF (2.2 L, 2.2 moles). The temperature was maintained in the range of 10° C. to 15° C. throughout the 45 minute addition. Following the addition, the resulting slurry was stirred for 2 hours at ambient temperature. Hexane (400 mL) was then added and stirring was continued for another 20 min. The slurry was filtered and the cake washed with 1/1 hexanes/THF and dried overnight at 60° C. in a vacuum oven to provide I-8C.1H-NMR (CD3OD) was consistent with the desired structure.

Pyrrolo[1,2-b]pyridazine-3-carbonitrile (I-8E): A stirred suspension of 3,3-diethoxy-2-formylpropionitrile potassium salt (I-8C, 5.10 g, 24.36 mmol) was cooled to 0° C., and concentrated HCl (7.11 mL, 85.26 mmol) was added dropwise at such a rate that the internal temperature of the reaction did not go above 20° C. After addition was complete, the reaction was stirred at room temperature for 20 minutes. To this reaction mixture was added a solution of 1-aminopyrrole (I-8D, 1.00 g, 12.18 mmol) in methanol (4.0 mL). After addition, the reaction mixture was refluxed at 90° C. for 2 hours. When heating was complete, the reaction was cooled to room temperature and concentrated to about half of the original volume. Saturated aqueous sodium bicarbonate was added carefully to the resulting residue until bubbling stopped. The solution was extracted with two portions of ethyl acetate. The combined organic layers were dried over sodium sulfate, filtered, concentrated in vacuo, and the resulting residue was purified by silica gel chromatography (eluent: EtOAc/hexanes) to provide I-8E.

7-bromopyrrolo[1,2-b]pyridazine-3-carbonitrile (I-8F): To a solution of pyrrolo[1,2-b]pyridazine-3-carbonitrile (I-8E, 840.0 mg, 5.9 mmol) in MeCN (30 mL) at room temperature was added N-bromosuccinimide in one portion. The reaction was stirred at room temperature for 30 minutes then poured into saturated aqueous sodium bicarbonate. The solution was concentrated in vacuo to remove the acetonitrile. The resulting aqueous layer was extracted with three portions of EtOAc. The combined organic layers were dried over sodium sulfate, filtered, concentrated in vacuo, and purified by silica gel chromatography (eluent: EtOAc/hexanes) to provide I-8F.

7-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)pyrrolo[1,2-b]pyridazine-3-carbonitrile (I-8): A microwave vial was charged with 7-bromopyrrolo[1,2-b]pyridazine-3-carbonitrile (I-8F, 416.5 mg, 1.9 mmol), bis(pinacolato)diboron (762.1 mg, 3.0 mmol), potassium acetate (552.3 mg, 5.6 mmol), and bis(triphenylphosphine)palladium(II) dichloride (65.8 mg, 0.094 mmol). Dioxane (8.0 mL) and DMF (4.0 mL) were added, and the reaction mixture was degassed with bubbling argon for 2 minutes. The vial was sealed and the reaction was heated at 120° C. in a microwave reactor for 60 minutes. After cooling, the reaction mixture was filtered and concentrated in vacuo. The resulting residue was partitioned between EtOAc and water. The aqueous layer was extracted with a second portion of EtOAc, and the combined organic layers were dried over sodium sulfate, filtered through a plug of Celite, and concentrated in vacuo. The resulting residue was purified by silica gel chromatography (eluent: EtOAc/hexanes) to provide I-8.

Preparation of Intermediate I-9

tert-butyl((1r,4r)-4-(hydrazinecarbonyl)cyclohexyl)carbamate: To a solution of (1r,4r)-4-((tert-butoxycarbonyl)amino)cyclohexane-1-carboxylic acid (10.0 g, 41.1 mmol) in THF (360 mL) was added 1,1′-carbonyldiimidazole (10.7 g, 65.8 mmol) as a single portion and the resulting mixture stirred for 16 h at room temperature. Hydrazine hydrate (10.0 mL, 206 mmol) was then added as a single portion. After 15 minutes approximately 200 mL THF was removed by rotary evaporation and the resulting slurry filtered rinsing with THF. The solid was dried under vacuum to give tert-butyl((1r,4r)-4-(hydrazinecarbonyl)cyclohexyl)carbamate which was used without further purification.

tert-butyl((1r,4r)-4-(2-(2,2-difluoroacetyl)hydrazine-1-carbonyl)cyclohexyl)carbamate: To a solution of tert-butyl((1r,4r)-4-(hydrazinecarbonyl)cyclohexyl)carbamate (1.50 g, 5.83 mmol) and diisopropylethylamine (2.6 mL, 14.9 mmol) in THF (20 mL) was added difluoroacetic anhydride (0.93 mL, 7.43 mmol) and the reaction mixture allowed to stir at room temperature. After 30 minutes additional difluoroacetic anhydride (0.40 mL, 3.20 mmol) was added and the reaction mixture allowed to stir for 30 minutes. The reaction mixture was then poured into water (20 mL), extracted with EtOAc (2×40 mL), washed with brine (1×15 mL), dried over MgSO4, filtered and concentrated to give crude tert-butyl((1r,4r)-4-(2-(2,2-difluoroacetyl)hydrazine-1-carbonyl)cyclohexyl)carbamate which was used without further purification.

tert-butyl((1r,4r)-4-(5-(difluoromethyl)-1,3,4-oxadiazol-2-yl)cyclohexyl)carbamate: To a solution of tert-butyl((1r,4r)-4-(2-(2,2-difluoroacetyl)hydrazine-1-carbonyl)cyclohexyl)carbamate (1.66 g, 4.96 mmol) in dry acetonitrile (40 mL) was added sequentially triphenylphosphine (3.90 g, 14.9 mmol), hexachloroethane (1.76 g, 7.34 mmol) and diisopropylethylamine (5.2 mL, 29.7 mmol) and the resulting solution allowed to stir for 15 minutes at room temperature. Upon completion the reaction mixture was poured into saturated aqueous NH4Cl (30 mL, and extracted with EtOAc (2×60 mL). The combined organics were washed with brine (1×15 mL), dried over MgSO4, filtered and concentrated to give a crude residue which was further purified using silica gel chromatography (eluent: EtOAc/hexanes) to give the product tert-butyl((1r,4r)-4-(5-(difluoromethyl)-1,3,4-oxadiazol-2-yl)cyclohexyl)carbamate.

(1r,4r)-4-(5-(difluoromethyl)-1,3,4-oxadiazol-2-yl)cyclohexan-1-amine hydrochloride (I-9): Tert-butyl((1r,4r)-4-(5-(difluoromethyl)-1,3,4-oxadiazol-2-yl)cyclohexyl)carbamate (1.26 g, 3.96 mmol) was dissolved in HCl solution (4.0 M in dioxane, 12 mL, 48 mmol) and the resulting mixture was stirred in a preheated 50° C. heating block for 30 minutes. Upon completion the suspension was filtered directly washing with dioxane (1×4 mL) and the solid dried under vacuum to give (1r,4r)-4-(5-(difluoromethyl)-1,3,4-oxadiazol-2-yl)cyclohexan-1-amine hydrochloride (I-9) which was used without further purification.

Preparation of Intermediate I-10

(R)-7-(5-bromo-4-((1-cyanoethyl)amino)pyridin-2-yl)pyrrolo[1,2-b]pyridazine-3-carbonitrile (I-10): A solution of (R)-2-((5-bromo-2-(3-cyanopyrrolo[1,2-b]pyridazin-7-yl)pyridin-4-yl)amino)propanamide (prepared as described for I-2 using the appropriate amine in step 1) (500 mg, 1.30 mmol) in THF (6.5 mL) was cooled to 0° C. To the cooled solution was added pyridine (0.52 mL, 6.49 mmol) followed by trifluoroacetic anhydride (0.27 mL, 1.95 mmol). The reaction mixture was allowed to warm to RT. After 2 hours, trifluoroacetic lanhydride (36 μL, 0.26 mmol) was added. The reaction mixture was stirred at RT for an additional 30 minutes, then concentrated in vacuo and purified by silica gel column chromatography (eluent: MeOH/DCM) to provide I-10.

Preparation of Intermediate I-11

(S)-8-(5-bromo-1,3,4-thiadiazol-2-yl)octahydropyrazino[2,1-c][1,4]oxazine (I-11): To a solution of 2,5-dibromo-1,3,4-thiadiazole (100 mg, 0.41 mmol) and (S)-octahydropyrazino[2,1-c][1,4]oxazine dihydrochloride (106 mg, 0.49 mmol) in 1,4-dioxane (1.0 mL) was added N,N-diisopropylethylamine (0.29 mL, 1.64 mmol). The reaction mixture was heated in a sealed vial for one hour, then concentrated in vacuo and purified by silica gel column chromatography (eluent: MeOH/DCM) to provide I-11.

The following intermediates were synthesized as described for I-11 using the appropriate starting amine:

Preparation of Intermediate I-12

(1R,5S,8r)-3-(5-bromo-1,3,4-thiadiazol-2-yl)-3-azabicyclo[3.2.1]octan-8-amine hydrochloride: To a solution of tert-butyl((1R,5S,8r)-3-(5-bromo-1,3,4-thiadiazol-2-yl)-3-azabicyclo[3.2.1]octan-8-yl)carbamate (synthesized as described for I-11) (40.0 mg, 0.10 mmol) in 1,4-dioxane (0.5 mL) was added hydrochloric acid (4M in dioxane, 0.13 mL, 0.51 mmol). The reaction mixture was stirred at 40° C. for 90 minutes, then concentrated in vacuo and used without additional purification.

The reaction mixture was stirred at RT for 10 minutes, then concentrated in vacuo and purified by silica gel column chromatography (eluent: MeOH/DCM) to provide I-12.

The following intermediates were synthesized as described for I-12 using the appropriate amine in step 1, and the appropriate anhydride, acid chloride, chloroformate, or sulfonyl chloride in step 2

Preparation of Intermediate I-13

Preparation of Intermediate I-14

tert-butyl((1S,2R)-2-((7-bromopyrrolo[2,1-f][1,2,4]triazin-2-yl)amino)cyclohexyl)carbamate: To a solution of 7-bromo-2-chloropyrrolo[2,1-f][1,2,4]triazine (150 mg, 0.65 mmol) and tert-butyl((1S,2R)-2-aminocyclohexyl)carbamate (152 mg, 0.71 mmol) in DMA (2.0 mL) was added N,N-diisopropylethylamine. The reaction mixture was heated to 160° C. in a microwave reactor for one hour. The cooled reaction mixture was diluted with water and extracted 3× with EtOAc. The combined organic layers were washed with saturated aqueous ammonium chloride and brine, then dried over sodium sulfate, isolated by vacuum filtration, and concentrated in vacuo. The resulting oil was purified by silica gel column chromatography (eluent: EtOAc/hexanes) to provide the desired material.

tert-butyl((1S,2R)-2-((7-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)pyrrolo[2,1-f][1,2,4]triazin-2-yl)amino)cyclohexyl)carbamate (I-14): A microwave vial was charged with tert-butyl((1S,2R)-2-((7-bromopyrrolo[2,1-f][1,2,4]triazin-2-yl)amino)cyclohexyl)carbamate (150 mg, 0.37 mmol), bis(pinacolato)diboron (149 mg, 0.59 mmol), trans-dichlorobis(triphenylphosphine)palladium (II) (12.9 mg, 0.018 mmol), and potassium acetate (109 mg, 1.11 mmol). Dioxane (0.82 mL) and DMF (0.4 mL) were added, and the reaction mixture was degassed by bubbling argon through the mixture for 60 seconds. The vial was sealed, and the reaction mixture was heated at 150° C. in a microwave reactor for 20 minutes. The reaction mixture was filtered, and the filtrate was concentrated and purified by silica gel column chromatography (eluent: EtOAc/hexanes) to provide I-14. Mass fragmentation provided the mass of the boronic acid.

Preparation of Intermediates I-15

Preparation of Intermediate I-16

N-(2-azabicyclo[2.2.2]octan-4-yl)acetamide hydrochloride: To a solution of tert-butyl 4-amino-2-azabicyclo[2.2.2]octane-2-carboxylate (200 mg, 0.88 mmol) in DCM (2.0 mL) was added triethylamine (0.25 mL, 1.77 mmol) followed by acetic anhydride (92 μL, 0.97 mmol). The reaction mixture was stirred for 30 minutes, then concentrated in vacuo. To the resulting oil was added HCl (4M in 1,4-dioxane, 2.2 mL, 8.8 mmol). The suspension was stirred at 40° C. for 2 hours, then concentrated and used without additional purification.

N-(2-(5-bromo-1,3,4-thiadiazol-2-yl)-2-azabicyclo[2.2.2]octan-4-yl)acetamide (1-16): To a crude suspension of N-(2-azabicyclo[2.2.2]octan-4-yl)acetamide hydrochloride (132 mg, 0.65 mmol) in dioxane (2 mL) was added 2,5-dibromo-1,3,4-thiadiazole (75.0 mg, 0.31 mmol) followed by N,N-diisopropylethylamine (0.27 mL, 1.5 mmol). The reaction mixture was heated at 120° C. for 1 hour. The cooled reaction mixture was concentrated in vacuo and purified by silica gel column chromatography (eluent: MeOH/DCM) to provide I-16.

The following intermediates were synthesized as described for I-16 using the appropriate starting amine:

Preparation of Intermediate I-17

2-(5-bromo-1,3,4-thiadiazol-2-yl)-7-methyl-2-azaspiro[3.5]nonan-7-ol (I-17): To a solution of crude 2-(5-bromo-1,3,4-thiadiazol-2-yl)-2-azaspiro[3.5]nonan-7-one (155 mg, 0.51 mmol) in THF (1.0 mL) at 0° C. was added methylmagnesium bromide (3M in diethyl ether, 0.26 mL, 0.77 mmol). The reaction mixture was warmed to RT and stirred for one hour. The reaction mixture was quenched with saturated aqueous sodium bicarbonate and extracted with three portions of EtOAc. The combined organic layers were dried over sodium sulfate, isolated by filtration, concentrated in vacuo and purified by silica gel column chromatography (eluent: MeOH/DCM) to provide I-17.

Preparation of Intermediate I-18

2-(3,6-diazabicyclo[3.1.1]heptan-3-yl)-5-bromo-1,3,4-thiadiazole: A solution of tert-butyl 3-(5-bromo-1,3,4-thiadiazol-2-yl)-3,6-diazabicyclo[3.1.1]heptane-6-carboxylate (synthesized following the protocol for I-11 using the appropriate amine) (150 mg, 0.42 mmol) in 1,1,1,3,3,3-hexafluoro-2-propanol (2.2 mL, 20.8 mmol) was heated in a microwave reactor 30 minutes at 150° C. The cooled reaction was concentrated in vacuo to provide the desired product which was used without purification.

1-(3-(5-bromo-1,3,4-thiadiazol-2-yl)-3,6-diazabicyclo[3.1.1]heptan-6-yl)-2-hydroxy-2-methylpropan-1-one (I-18): To a solution of crude 2-(3,6-diazabicyclo[3.1.1]heptan-3-yl)-5-bromo-1,3,4-thiadiazole (108 mg, 0.42 mmol) and 2-hydroxy-2-methylpropanoic acid (56.2 mg, 0.54 mmol) in DMF (0.5 mL) was added N,N-diisopropylethylamine (0.30 mL, 1.66 mmol) followed by a solution of HATU (237 mg, 0.62 mmol) in DMF (0.5 mL). The reaction mixture was stirred at RT for 15 minutes, then concentrated in vacuo and partitioned between water and EtOAc. The aqueous later was extracted two additional times with EtOAc, and the combined organic layers were dried over sodium sulfate, isolated by filtration, concentrated in vacuo, and purified by silica gel column chromatography (eluent: MeOH/DCM) to provide I-18.

Preparation of Intermediate I-19

1-(3-(5-bromo-1,3,4-thiadiazol-2-yl)-3,6-diazabicyclo[3.1.1]heptan-6-yl)ethan-1-one (I-19): To a solution of crude 2-(3,6-diazabicyclo[3.1.1]heptan-3-yl)-5-bromo-1,3,4-thiadiazole (41.9 mg, 0.16 mmol) in DCM (3.2 mL) was added triethylamine (92 μL, 0.66 mmol) followed by acetic anhydride (16.7 μL, 0.18 mmol). The reaction mixture was stirred at RT for 30 minutes then concentrated in vacuo and purified by silica gel column chromatography (eluent: MeOH/DCM) to provide I-19.

The following intermediates were synthesized as described for I-19 using the appropriate amine and the appropriate anhydride, acid chloride, chloroformate, or sulfonyl chloride:

Preparation of Intermediate I-20

(±) methyl trans-2-(2-formylhydrazine-1-carbonyl)cyclopropane-1-carboxylate: To a suspension of racemic trans-2-methoxycarbonylcyclopropanecarboxylic acid (2 g, 13.9 mmol) and N-methylmorpholine (1.68 mL, 15.3 mmol) in THF (50 mL) at 0° C. was added isobutyl chloroformate (1.98 mL, 15.3 mmol) dropwise. The suspension was stirred for 15 minutes at 0° C., and then formohydrazide (917 mg, 15.3 mmol) was added in one portion. The suspension was stirred for 10 minutes at 0° C., and was then stirred for 30 minutes at room temperature. Methanol (10 mL) was added to the reaction mixture, and slurry was filtered, rinsing with methanol. The filtrate was concentrated and used without further purification.

(±) methyl trans-2-(1,3,4-thiadiazol-2-yl)cyclopropane-1-carboxylate (t I-20): To a solution of methyl(1S,2S)-2-(2-formylhydrazine-1-carbonyl)cyclopropane-1-carboxylate (2 g, 10.7 mmol) in THF (50 mL) at 65° C. was added Lawesson's Reagent (6.52 g, 16.1 mmol) in one portion. The reaction was stirred at 65° C. for 20 minutes, until conversion of the starting material to desired product was observed by LCMS. The flask was cooled, and diluted with EtOAc (100 mL). The organic layer was washed with water (50 mL). The aqueous layer was back-extracted with EtOAc (2×50 mL), and the combined organic layers were dried over MgSO4, filtered, and concentrated under reduced pressure. The crude material was purified twice by silica gel chromatography (eluent EtOAc/hexanes followed by methanol/EtOAc) to give±I-20 as a clear viscous oil.

Preparation of Intermediates I-21

2-(1-(oxetan-3-yl)piperidin-4-yl)-1,3,4-thiadiazole (I-21): To a solution of 2-(piperidin-4-yl)-1,3,4-thiadiazole hydrochloride (synthesized as described for I-1 using the appropriate carboxylic acid) (200 mg, 0.97 mmol) in CH2Cl2(5 mL) was added N,N-Diisopropylethylamine (0.17 mL, 0.97 mmol). The mixture was cooled to 0° C., and then oxetan-3-one (0.14 g, 1.94 mmol) was added, followed by sodium triacetoxyborohydride (618 mg, 2.92 mmol). The mixture was stirred under nitrogen for 90 minutes, upon which time conversion of the starting material to desired product was observed by LCMS. The reaction was quenched with 5 drops of sat. aq. NaHCO3, and the crude reaction was dry-loaded onto silica. The crude material was purified by silica gel chromatography (eluent EtOAc/hexanes followed by methanol/EtOAc) to give I-21.

Preparation of Intermediate I-22

(1R,5S,8r)-3-benzyl-8-methyl-3-azabicyclo[3.2.1]octan-8-ol(I-22): Toanoven-dried 250 mL round bottom flask was added 3-benzyl-3-azabicyclo[3.2.1]octan-8-one (3 g, 13.9 mmol), and the flask was placed under an N2atmosphere. THF (100 mL) was added, and the solution was cooled to −78° C. MeMgBr (3M in ether, 13.9 mL, 41.8 mmol) was added dropwise, and the reaction was stirred 30 min at −78° C. LCMS aliquot showed conversion to desired product. 10 mL sat. aq. ammonium chloride was added dropwise, and the mixture was allowed to warm to RT. The mixture was diluted with 200 mL EtOAc and 50 mL water, and the layers separated. The aq. layer was extracted twice with 50 mL EtOAc, and the combined organic layers were dried over MgSO4, filtered, and concentrated. The crude material was purified by silica gel chromatography (eluent EtOAc/hexanes) to give I-22.

Preparation of Intermediate I-23

(1R,5S,8r)-8-methyl-3-azabicyclo[3.2.1]octan-8-ol: To a 25 mL round bottom flask was added I-22 (0.75 g, 3.24 mmol) and ethanol (10 mL). Pd on carbon (10% wt, 0.1 g) was added in one portion, and the mixture was degassed with H2before stirring overnight under an H2atmosphere. LCMS showed complete conversion to the desired product, and the mixture was degassed with argon. The mixture was filtered over celite to remove the solids, rinsing with EtOH. The filtrate was concentrated, and used directly for next step.

(1R,5S,8r)-3-(5-bromo-1,3,4-thiadiazol-2-yl)-8-methyl-3-azabicyclo[3.2.1]octan-8-ol (I-23): To a solution of 2,5-dibromo-1,3,4-thiadiazole (777 mg, 3.19 mmol) and (R,5S,8r)-8-methyl-3-azabicyclo[3.2.1]octan-8-ol (450 mg, 3.19 mmol) in DMF (2.0 mL) was added N,N-diisopropylethylamine (1.11 mL, 6.37 mmol). The reaction mixture was stirred at 120° C. in a sealed vial for one hour. The cooled reaction mixture was diluted with EtOAc (50 mL), and washed twice with water (15 mL). The organic layer was dried over MgSO4, filtered, concentrated under reduced pressure, and purified by silica gel column chromatography (eluent: EtOAc/hexanes) to provide 1-23.

Preparation of Intermediate I-24

N-((1R,5S,8s)-3-benzyl-8-methyl-3-azabicyclo[3.2.1]octan-8-yl)acetamide: To a 250 mL round bottomed flask was added 3-benzyl-8-methyl-3-azabicyclo[3.2.1]octan-8-ol (I-22) (2 g, 8.65 mmol) and acetonitrile (15 mL), and the solution was then cooled to 0° C. Concentrated Sulfuric acid (12 mL) was added dropwise, and the reaction was warmed to RT and stirred overnight. The reaction mixture was poured into ice, and the resulting solution was adjusted (carefully) to pH 10 with sat. aq. KOH. Significant precipitates appeared. Mixture extracted with EtOAc (3×200 mL), and organic layers were dried over MgSO4, filtered, and concentrated. The crude material purified by silica gel column chromatography (eluent: EtOAc/hexanes, then MeOH/EtOAc) to provide the product.

N-((1R,5S,8s)-8-methyl-3-azabicyclo[3.2.1]octan-8-yl)acetamide: To a 25 mL round bottom flask was added N-((1R,5S,8s)-3-benzyl-8-methyl-3-azabicyclo[3.2.1]octan-8-yl)acetamide (0.14 g, 0.514 mmol) and ethanol (6 mL). Pd on carbon (10% wt, 55 mg) was added in one portion, and the mixture was degassed with H2 before stirring overnight under an H2atmosphere. LCMS showed complete conversion to the desired product, and the mixture was degassed with argon. The mixture was filtered over celite to remove the solids, rinsing with EtOH. The filtrate was concentrated, and used directly for next step.

N-((1R,5S,8s)-3-(5-bromo-1,3,4-thiadiazol-2-yl)-8-methyl-3-azabicyclo[3.2.1]octan-8-yl)acetamide (I-24): To a solution of 2,5-dibromo-1,3,4-thiadiazole (132 mg, 0.543 mmol) and N-((1R,5S,8s)-8-methyl-3-azabicyclo[3.2.1]octan-8-yl)acetamide (90 mg, 0.494 mmol) in 1,4-dioxane (0.5 mL) was added N,N-diisopropylethylamine (0.22 mL, 1.23 mmol). The reaction mixture was stirred at 120° C. in a sealed vial for one hour. The cooled reaction mixture was concentrated under reduced pressure, and purified by silica gel column chromatography (eluent: EtOAc/hexanes, then MeOH/EtOAc) to provide 1-24.

Preparation of Intermediates I-25 and I-26

Exo-N-((1R,5S,9r)-3-(5-bromo-1,3,4-thiadiazol-2-yl)-3-azabicyclo[3.3.1]nonan-9-yl)acetamide (I-SEA6) and Endo-N-((1R,5S,9s)-3-(5-bromo-1,3,4-thiadiazol-2-yl)-3-azabicyclo[3.3.1]nonan-9-yl)acetamide (I-26): To a solution of 2,5-dibromo-1,3,4-thiadiazole (250 mg, 1.02 mmol) and tert-butyl N-(3-azabicyclo[3.3.1]nonan-9-yl)carbamate (246 mg, 1.02 mmol) in 1,4-dioxane (1 mL) was added N,N-Diisopropylethylamine (0.36 mL, 2.05 mmol). The reaction mixture was stirred at 120° C. in a sealed vial for one hour. The cooled reaction mixture was concentrated under reduced pressure, and purified by silica gel column chromatography (eluent: EtOAc/hexanes) to separate both isomers endo-(I-25) and exo-(I-26). The endo- and exo-isomers were distinguished by 2D NOESY spectroscopy.

Preparation of Intermediate I-27

N-(4-(5-bromo-1,3,4-thiadiazol-2-yl)phenyl)acetamide (I-27): To a suspension of 4-(5-bromo-1,3,4-thiadiazol-2-yl)aniline (200 mg, 1.13 mmol) in CH2Cl2(5 mL) and THF (5 mL) at 0° C. was added triethylamine (0.315 mL, 2.26 mmol) followed by acetic anhydride (0.11 mL, 1.13 mmol). The reaction mixture was stirred at RT for 1 hour, followed by 30 minutes at 40° C. The reaction was diluted with 20 mL EtOAc, and washed with 10 mL water. The aqueous layer was extracted with 3×10 mL EtOac, and the combined organic layers were dried over MgSO4and concentrated under reduced pressure. The residue was purified by silica gel column chromatography (eluent: EtOAc/hexanes, then MeOH/EtOAc) to provide I-27.

Preparation of Intermediates I-28 and I-29

tert-butyl(3R)-3-fluoro-4-hydroxy-4-methylpiperidine-1-carboxylate: To an oven-dried 100 mL round bottom flask was added tert-butyl(3S)-3-fluoro-4-oxo-piperidine-1-carboxylate (1 g, 4.6 mmol), and the flask was placed under an N2atmosphere. THF (40 mL) was added, and the solution was cooled to −78° C. MeMgBr (3M in ether, 3.84 mL, 11.5 mmol) was added dropwise, and the reaction was stirred 5 minutes at −78° C. LCMS aliquot showed conversion to desired product. 5 mL Sat. aq. ammonium chloride was added dropwise, and the mixture was allowed to warm to RT. The mixture was diluted with 100 mL EtOAc and 50 mL water, and the layers separated. The aq. layer was extracted twice with 50 mL EtOAc, and the combined organic layers were dried over MgSO4, filtered, and concentrated. The crude material was purified by silica gel chromatography (eluent EtOAc/hexanes) to give the product as a diastereomeric mixture.

(3R)-3-fluoro-4-methylpiperidin-4-ol hydrochloride: To a solution of tert-butyl(3R)-3-fluoro-4-hydroxy-4-methylpiperidine-1-carboxylate (mixture of diastereomers) (1.03 g, 4.42 mmol) in 1,4-dioxane (8 mL) and methanol (3 mL) was added hydrochloric acid (4M in dioxane, 4.7 mL, 18.7 mmol). The reaction mixture was stirred at RT for 4 hours, then concentrated under reduced pressure and used without additional purification.

(3R,4R)-1-(5-bromo-1,3,4-thiadiazol-2-yl)-3-fluoro-4-methylpiperidin-4-ol (I-28) and (3R,4S)-1-(5-bromo-1,3,4-thiadiazol-2-yl)-3-fluoro-4-methylpiperidin-4-ol (I-29): To a solution of 2,5-dibromo-1,3,4-thiadiazole (800 mg, 3.28 mmol) and (3R)-3-fluoro-4-methylpiperidin-4-ol hydrochloride (700 mg, 4.13 mmol) in DMF (4.0 mL) was added N,N-diisopropylethylamine (1.71 mL, 9.84 mmol). The reaction mixture was stirred at 120° C. in a sealed vial for one hour. The cooled reaction mixture was diluted with EtOAc (50 mL), and washed with sat. aq. NH4C1(20 mL). The aqueous was back-extracted three times with EtOAc (20 mL). The combined organic layers were dried over MgSO4, filtered, concentrated under reduced pressure, and purified by silica gel column chromatography (eluent: EtOAc/hexanes) to provide a mixture of diastereomers, which were further separated by SFC.

The following intermediates were synthesized as described for I-28 using tert-butyl(3S)-3-fluoro-4-hydroxy-4-methylpiperidine-1-carboxylate in step 1

Preparation of Intermediate I-30

Endo-7-benzyl-3-oxa-7-azabicyclo[3.3.1]nonan-9-one oxime: 7-benzyl-3-oxa-7-azabicyclo[3.3.1]nonan-9-one (2 g, 8.65 mmol), hydroxylamine hydrochloride (900 mg, 13 mmol) and pyridine (1.2 mL, 14.9 mmol) in ethanol (20 mL) was heated to 100° C. for 2 hours. The reaction mixture was concentrated under reduced pressure. 2.5 N Aqueous sodium hydroxide solution (10 mL) was added to the residue. The resulting solution was extracted 2× with ethyl acetate (50 mL), washed with water, dried over magnesium sulfate, filtered and concentrated under reduced pressure. The crude product was purified by column chromatography on silica gel (eluent: EtOAc/hexane mixture) to afford the product.

7-benzyl-3-oxa-7-azabicyclo[3.3.1]nonan-9-amine: To a de-oxygentated solution of 7-benzyl-3-oxa-7-azabicyclo[3.3.1]nonan-9-one oxime (1.77 g, 7.19 mmol) in ethanol (50 mL) was added Raney nickel (0.6 g, suspension in water) under an argon atmosphere. The reaction was hydrogenated using a hydrogen balloon overnight. The reaction was filtered through a bed of celite and the filtrate was concentrated to dryness. Care was taken to avoid drying out the filtered nickel catalyst. The crude residue was purified by flash column chromatography on silica gel (eluent: MeOH/CH2Cl2). A mixture of exo- and endo-isomers was obtained.

The reaction mixture was stirred at 0° C. for 15 minutes, then diluted with EtOAc (50 mL) and sat. aq. NH4Cl. The layers were separated, and the aqueous layer was extracted with EtOAc (5×20 mL). the combined organic layers were concentrated under reduced pressure and purified by silica gel column chromatography (eluent: EtOAc/hexanes, then MeOH/EtOAc) to provide the product.

N-(3-oxa-7-azabicyclo[3.3.1]nonan-9-yl)acetamide: To a 100 mL round bottom flask was added N-(7-benzyl-3-oxa-7-azabicyclo[3.3.1]nonan-9-yl)acetamide (1.35 g, 4.92 mmol) and ethanol (20 mL). Pd on carbon (10% wt, 524 mg) was added in one portion, and the mixture was degassed with H2before stirring overnight under an H2atmosphere.

LCMS showed complete conversion to the desired product, and the mixture was degassed with argon. The mixture was filtered over celite to remove the solids, rinsing with EtOH.

The filtrate was concentrated, and used directly for next step.

Endo-N-(7-(5-bromo-1,3,4-thiadiazol-2-yl)-3-oxa-7-azabicyclo[3.3.1]nonan-9-yl)acetamide (I-30): To a solution of 2,5-dibromo-1,3,4-thiadiazole (1.13 g, 4.64 mmol) and N-(3-oxa-7-azabicyclo[3.3.1]nonan-9-yl)acetamide (900 mg, 4.89 mmol) in 1,4-dioxane (6 mL) was added N,N-Diisopropylethylamine (1.7 mL, 9.77 mmol). The reaction mixture was stirred at 120° C. in a sealed vial for one hour. The cooled reaction mixture was concentrated under reduced pressure, and purified by silica gel column chromatography (eluent: EtOAc/hexanes) to yield a mixture of endo- and exo-isomers (approximately 1:4 endo:exo). The mixture was further purified by SFC to isolate the endo-isomer 1-30.

Preparation of Intermediate I-31

methyl 4-((tert-butoxycarbonyl)amino)-1-fluorocyclohexane-1-carboxylate: To a solution of trans-4-aminocyclohexane-1-fluoro-1-carboxylic acid methyl ester hydrochloride (3 g, 14.2 mmol) in THe (50 mL) was added triethylamine (4.35 mL, 31.2 mmol) and di-tert-butyl dicarbonate (3.4 g, 15.6 mmol) at room temperature. The mixture was stirred at room temperature overnight, then poured into saturated aqueous ammonium chloride (50 mL) and extracted with ethyl acetate (3×100 mL). The combined organic phases were washed with water and brine, dried over anhydrous MgSO4and concentrated under reduced pressure to obtain the product. Material was used without further purification.

4-((tert-butoxycarbonyl)amino)-1-fluorocyclohexane-1-carboxylic acid: To a 250 mL round bottom flask with methyl 4-((tert-butoxycarbonyl)amino)-1-fluorocyclohexane-1-carboxylate (3.9 g, 14.2 mmol) was added THF (50 mL), MeOH (10 mL) and water (5 mL). icH (M aq., 42.5 mL, 42.5 mmol) was added, and the mixture was stirred overnight at rt. The mixture was concentrated on the rotovap, and dissolved in EtOAc (100 mL). The mixture was acidified with 50% citric acid, and the layers separated. The aqueous layer was back extracted with 2×50 mL EtOAc. The combined organic layers were dried over MgSO4, filtered, and concentrated under reduced pressure. Material was used without further purification.

tert-butyl(4-fluoro-4-(2-formylhydrazine-1-carbonyl)cyclohexyl)carbamate: To a suspension of 4-((tert-butoxycarbonyl)amino)-1-fluorocyclohexane-1-carboxylic acid (3.6 g, 13.8 mmol) and N-methylmorpholine (1.67 mL, 15.2 mmol) in THF (50 mL) at 0° C. was added isobutyl chloroformate (1.97 mL, 15.2 mmol) dropwise. The suspension was stirred for 20 minutes at 0° C., and then formohydrazide (1.24 mg, 20.7 mmol) was added in one portion. The suspension was stirred for 1 hour at room temperature. Methanol (10 mL) was added to the reaction mixture, and slurry was filtered, rinsing with methanol. The filtrate was concentrated used without further purification.

tert-butyl(4-fluoro-4-(1,3,4-thiadiazol-2-yl)cyclohexyl)carbamate: To a solution of tert-butyl(4-fluoro-4-(2-formylhydrazine-1-carbonyl)cyclohexyl)carbamate (3.79 g, 12.5 mmol) in THF (100 mL) at 65° C. was added Lawesson's Reagent (7.58 g, 18.7 mmol) in one portion. The reaction was stirred at 65° C. for 60 minutes, until conversion of the starting material to desired product was observed by LCMS. The flask was cooled, and diluted with EtOAc (100 mL). The organic layer was washed with water (50 mL). The aqueous layer was back-extracted with EtOAc (2×50 mL), and the combined organic layers were dried over MgSO4, filtered, and concentrated under reduced pressure. The crude material was purified by silica gel chromatography (eluent EtOAc/hexanes followed by methanol/EtOAc) to give the product.

4-fluoro-4-(1,3,4-thiadiazol-2-yl)cyclohexan-1-amine hydrochloride: To a solution of tert-butyl(4-fluoro-4-(1,3,4-thiadiazol-2-yl)cyclohexyl)carbamate (2.5 g, 8.3 mmol) in 1,4-dioxane (20 mL) and methanol (4 mL) was added hydrochloric acid (4M in dioxane, 10 mL, 40 mmol). The reaction mixture was stirred at RT for 1 hour, then overnight at 55° C. The mixture was concentrated under reduced pressure and used without additional purification.

N-(4-fluoro-4-(1,3,4-thiadiazol-2-yl)cyclohexyl)acetamide (I-31): To a suspension of 4-fluoro-4-(1,3,4-thiadiazol-2-yl)cyclohexan-1-amine hydrochloride (2.1 g, 8.83 mmol) in CH2Cl2(50 mL) at 0° C., was added triethylamine (3.70 mL, 26.5 mmol) and then acetic anhydride (0.835 mL, 8.83 mmol). The reaction mixture was stirred at 0° C. for 15 minutes, then diluted with EtOAc (100 mL) and sat. aq. NH4C1(30 mL). The layers were separated, and the aqueous layer was extracted with EtOAc (5×40 mL). The combined organic layers were concentrated under reduced pressure and purified by silica gel column chromatography (eluent: EtOAc/hexanes, then MeOH/EtOAc), then by silica gel column chromatography (eluent: MeOH/EtOAc) to provide the product I-31 as a mixture of diastereomers.

Preparation of Intermediate I-32

methyl trans-3-hydroxy-3-methylcyclobutane-1-carboxylate: To a solution trans-3-hydroxy-3-methylcyclobutane-1-carboxylic acid (850 mg, 6.53 mmol) in methanol (15 mL) was added concentrated sulfuric acid (0.142 mL). The mixture was refluxed overnight and then concentrated. The residue was dissolved in water (10 mL) and the mixture was extracted with ethyl acetate (20 mL×2). The combined organic phases were washed with sat. aq. NaHCO3(100 mL), and dried over MgSO4. Concentration gave the methyl ester, which was carried forward. methyl trans-3-((tert-butyldimethylsilyl)oxy)-3-methylcyclobutane-1-carboxylate: To methyl trans-3-hydroxy-3-methylcyclobutane-1-carboxylate (900 mg, 6.24 mmol) in anhydrous DMF (15 mL), was added imidazole (1.27 g, 18.7 mmol) and tert-Butyldimethylsilyl chloride (2.82 g, 18.7 mmol). The resulting mixture was heated at 80° C. overnight. After cooling to room temperature, water (20 mL) was added and the aqueous layer was extracted with ethyl acetate (40 mL×3). The combined organic layers were washed with brine and dried over anhydrous MgSO4. The solid was filtered off and the filtrate was concentrated in vacuo. The residue was purified by silica gel chromatography, (eluent: EtOAc/hexanes)

trans-3-((tert-butyldimethylsilyl)oxy)-3-methylcyclobutane-1-carboxylic acid: To a 50 mL round bottom flask was added methyl trans-3-((tert-butyldimethylsilyl)oxy)-3-methylcyclobutane-1-carboxylate (1.36 g, 5.26 mmol) and dissolved in MeOH (15 mL) and water (1 mL). Solid LiOH (378 mg, 15.8 mmol) was added, and the mixture was stirred overnight at 50° C. Most of the methanol was evaporated under reduced pressure, and the residue was dissolved in EtOAc (100 mL). The mixture was acidified with 50% aq. citric acid, and the layers were separated. The aq layer was extracted 2×50 mL EtOAc. The combined organic layers were dried over MgSO4, filtered, and concentrated under reduced pressure. The material was carried forward without further purification.

N-[[3-[tert-butyl(dimethyl)silyl]oxy-3-methyl-cyclobutanecarbonyl]amino]formamide: To a suspension of trans-3-((tert-butyldimethylsilyl)oxy)-3-methylcyclobutane-1-carboxylic acid (1.17 g, 4.79 mmol) and N-methylmorpholine (0.579 mL, 5.27 mmol) in 2-MeTHF (15 mL) at 0° C. was added isobutyl chloroformate (0.683 mL, 5.27 mmol) dropwise. The suspension was stirred for 20 minutes at 0° C., and then formohydrazide (575 mg, 9.57 mmol) was added in one portion. The suspension was stirred for 1 hour at room temperature. The resulting slurry was filtered. The filtrate was concentrated used without further purification.

2-(trans-3-((tert-butyldimethylsilyl)oxy)-3-methylcyclobutyl)-1,3,4-thiadiazole: To a solution of N-[[3-[tert-butyl(dimethyl)silyl]oxy-3-methyl-cyclobutanecarbonyl]amino]formamide (1.3 g, 4.54 mmol) in THF (20 mL) at 65° C. was added Lawesson's Reagent (2.75 g, 6.81 mmol) in one portion. The reaction was stirred at 65° C. for 10 minutes, until conversion of the starting material to desired product was observed by LCMS. The flask was cooled, and dry loaded onto silica. The crude material was purified by silica gel chromatography (eluent EtOAc/hexanes followed by methanol/EtOAc) to give the product.

The crude material was purified by silica gel chromatography (eluent EtOAc/hexanes followed by methanol/EtOAc) to give the product I-32.

Preparation of Intermediate I-33

N′-formyl-1,4-dioxaspiro[4.5]decane-8-carbohydrazide: To a suspension of 1,4-dioxaspiro[4.5]decane-8-carboxylic acid (0.8 g, 4.3 mmol) and N-methylmorpholine (0.496 mL, 4.51 mmol) in THF (15 mL) at 0° C. was added isobutyl chloroformate (0.585 mL, 4.51 mmol) dropwise. The suspension was stirred for 30 minutes at 0° C., and then formohydrazide (387 mg, 6.44 mmol) was added in one portion. The suspension was stirred for 30 minutes at room temperature. The resulting slurry was filtered. The filtrate was concentrated used without further purification.

2-(1,4-dioxaspiro[4.5]decan-8-yl)-1,3,4-thiadiazole (I-33): To a solution of N′-formyl-1,4-dioxaspiro[4.5]decane-8-carbohydrazide (0.9 g, 3.94 mmol) in THF (20 mL) at 65° C. was added Lawesson's Reagent (1.91 g, 4.73 mmol) in one portion. The reaction was stirred at 65° C. for 20 minutes, until conversion of the starting material to desired product was observed by LCMS. The flask was cooled, and dry loaded onto silica. The crude material was purified twice by silica gel chromatography (eluent EtOAc/hexanes followed by methanol/EtOAc) to give the product I-33.

Preparation of Intermediate I-34

2-chloro-5-iodo-N-(oxetan-3-yl)pyridin-4-amine: To a solution of 2-chloro-4-fluoro-5-iodo-pyridine (750 mg, 2.91 mmol) and oxetan-3-amine (4.23 g, 68.0 mmol) in NMP (4.0 mL) was added N,N-diisopropylethylamine (319 mg, 4.37 mmol). The reaction mixture was heated in a microwave at 150° C. for 60 minutes, then cooled and diluted with EtOAc (50 mL) and water (10 mL). The resulting mixture was extracted with ethyl acetate (4×20 mL). The combined organic extracts were washed with brine, dried over MgSO4, filtered and concentrated under reduced pressure. The crude material was purified by silica gel chromatography (eluent EtOAc/hexanes) to give the product

4-(5-(6-chloro-4-(oxetan-3-ylamino)pyridin-3-yl)-1,3,4-thiadiazol-2-yl)cyclohexan-1-one: To a vial with 5-(5-(1,4-dioxaspiro[4.5]decan-8-yl)-1,3,4-thiadiazol-2-yl)-2-chloro-N-(oxetan-3-yl)pyridin-4-amine (159 mg, 0.389 mmol) was added acetonitrile (5 mL) and HCl (1N aq, 2 mL, 2 mmol). The reaction was stirred for 4 hours at RT, then diluted with EtOAc (40 mL), and neutralized with sat. aq. NaHCO3. The layers were separated, and the aqueous layer was extracted twice with 20 mL EtOAc. The combined organic layers were dried over MgSO4, filtered, and concentrated under reduced pressure. The crude material was purified by silica gel chromatography (eluent: EtOAc/hexanes, then MeOH/EtOAc) to provide the product.

4-(5-(6-chloro-4-(oxetan-3-ylamino)pyridin-3-yl)-1,3,4-thiadiazol-2-yl)-1-methylcyclohexan-1-ol (I-34): To an oven-dried vial was added 4-(5-(6-chloro-4-(oxetan-3-ylamino)pyridin-3-yl)-1,3,4-thiadiazol-2-yl)cyclohexan-1-one (110 mg, 0.3 mmol), and the flask was placed under an N2atmosphere. THF (5 mL) was added, and the solution was cooled to −78° C. MeMgBr (3M in ether, 0.4 mL, 1.2 mmol) was added dropwise, and the reaction was stirred 15 minutes at −78° C. LCMS aliquot showed conversion to desired product. 5 mL Sat. aq. ammonium chloride was added dropwise, and the mixture was allowed to warm to RT. The mixture was diluted with 30 mL EtOAc and 5 mL water, and the layers separated. The aq. layer was extracted twice with 10 mL EtOAc, and the combined organic layers were dried over MgSO4, filtered, and concentrated. The crude material was purified by silica gel chromatography (eluent EtOAc/hexanes, then MeOH/EtOAc) to afford both separable isomers of the product I-34.

Preparation of Intermediate I-35

N-((1R,5S,8s)-3-(5-bromo-1,3,4-thiadiazol-2-yl)-3-azabicyclo[3.2.1]octan-8-yl)-N-methylacetamide (I-35): To a solution of N-((1R,5S,8s)-3-(5-bromo-1,3,4-thiadiazol-2-yl)-3-azabicyclo[3.2.1]octan-8-yl)acetamide (50 mg, 0.15 mmol) in THF (1.5 mL) was added sodium hydride (5 mg, 0.22 mmol). Gas evolution was observed and the resulting mixture stirred at room temperature for 5 minutes before iodomethane (0.011 mL, 0.18 mmol) was added. The reaction was stirred at room temperature for 3 days before additional sodium hydride (5 mg, 0.22 mmol) and iodomethane (0.011 mL, 0.18 mmol) was added. Upon completion, the reaction mixture was poured into water/NH4Cl and extracted with EtOAc. The combined organic layers were dried over MgSO4, filtered and concentrated. The resulting crude residue was purified by silica gel chromatography (eluent: MeOH/CH2Cl2) to give I-35.

Preparation of Intermediate I-36

2-Chloroethyl((1R,5S,8s)-3-(5-bromo-1,3,4-thiadiazol-2-yl)-3-azabicyclo[3.2.1]octan-8-yl)carbamate: To a solution of (1R,5S,8s)-3-(5-bromo-1,3,4-thiadiazol-2-yl)-3-azabicyclo[3.2.1]octan-8-amine hydrochloride (150 mg, 0.46 mmol) in THF (4.5 mL) was added potassium carbonate (190 mg, 1.37 mmol) and 2-chloroethyl chloroformate (98 mg, 0.68 mmol). The resulting slurry was stirred at room temperature for 2 days. Additional 2-chloroethyl chloroformate (98 mg, 0.68 mmol) was added and stirred for 4.5 hours. The reaction was filtered through celite, washed with EtOAc, and concentrated to dryness. The resulting crude residue was purified by silica gel chromatography (eluent: EtOAc/Hexanes) to give the desired product.

3-((1R,5S,8s)-3-(5-bromo-1,3,4-thiadiazol-2-yl)-3-azabicyclo[3.2.1]octan-8-yl)oxazolidin-2-one (I-36): To a solution of 2-Chloroethyl((1R,5S,8s)-3-(5-bromo-1,3,4-thiadiazol-2-yl)-3-azabicyclo[3.2.1]octan-8-yl)carbamate (24.5 mg, 0.062 mmol) in MeOH (0.75 mL) was added sodium methoxide (0.023 mL, 0.10 mmol). After stirring at room temperature for 3 days, the reaction was concentrated and dissolved in EtOAc. The organic layer was washed with NH4Cl/H2O, dried over MgSO4, and concentrated. The resulting crude residue was purified by silica gel chromatography (eluent: MeOH/CH2Cl2) to give I-36.

Preparation of Intermediate I-37

(1R,5S,8s)-3-(5-bromo-1,3,4-thiadiazol-2-yl)-N-(pyrimidin-2-yl)-3-azabicyclo[3.2.1]octan-8-amine (I-37): To a solution of (1R,5S,8s)-3-(5-bromo-1,3,4-thiadiazol-2-yl)-3-azabicyclo[3.2.1]octan-8-amine hydrochloride (25 mg, 0.077 mmol) in NMP (0.4 mL) was added 2-chloropyridine (19 mg, 0.17 mmol) and N,N-diisopropylethylamine (0.07 mL, 0.40 mmol). The reaction was sealed and heated in a microwave to 150° C. for 30 min. The reaction mixture was purified by RP-HPLC (eluent: water/MeCN*0.1% TFA). The resulting product fractions were combined and concentrated to dryness. The residue was dissolved in CH2Cl2and washed with aqueous bicarbonate. The aqueous layer was back-extracted with CH2Cl2and the combined organic layers were dried over MgSO4and concentrated to give I-37.

Preparation of Intermediate I-38

(1R,5S,8s)-3-(5-bromo-1,3,4-thiadiazol-2-yl)-N-(pyrimidin-2-yl)-3-azabicyclo[3.2.1]octan-8-amine (I-38): To a solution of (1R,5S,8s)-3-(5-bromo-1,3,4-thiadiazol-2-yl)-3-azabicyclo[3.2.1]octan-8-amine hydrochloride (75 mg, 0.23 mmol) in NMP (1.2 mL) was added 2,3-difluoropyridine (82 mg, 0.71 mmol) and N,N-diisopropylethylamine (0.2 mL, 1.15 mmol). The reaction was sealed and heated 120° C. for 4 days. The reaction mixture was purified by RP-HPLC (eluent: water/MeCN*0.1% TFA). The resulting product fractions were combined and concentrated to dryness. The residue was dissolved in CH2Cl2and washed with aqueous bicarbonate. The aqueous layer was back-extracted with CH2Cl2and the combined organic layers were dried over MgSO4and concentrated to give I-38.

Preparation of Intermediate I-39

1-(5-bromo-1,3,4-thiadiazol-2-yl)piperidine-4-carboxylic acid: To a solution of methyl 1-(5-bromo-1,3,4-thiadiazol-2-yl)piperidine-4-carboxylate (22 mg, 0.073 mmol) in MeOH (0.15 mL) and THF (0.3 mL) was added aqueous lithium hydroxide (1M, 0.15 mL, 0.15 mmol). The resulting solution was stirred at room temperature for 3 days. The mixture was concentrated to dryness and used crude in the next step.

1-(5-bromo-1,3,4-thiadiazol-2-yl)-N-methylpiperidine-4-carboxamide (I-39): To a solution of 1-(5-bromo-1,3,4-thiadiazol-2-yl)piperidine-4-carboxylic acid (21 mg, 0.72 mmol) in DMF (0.3 mL) was added a THF solution of methylamine (2M, 0.35 mL, 0.7 mmol), HATU (33 mg, 0.087 mmol), and N,N-diisopropylethylamine (0.1 mL, 0.57 mmol). The reaction was stirred at room temperature for 1.5 h and concentrated to dryness. The resulting crude residue was purified by silica gel chromatography (eluent: MeOH/CH2Cl2) to give I-39.

Preparation of Intermediate I-40

Tert-butyl(4-(5-bromo-1,3,4-thiadiazol-2-yl)cyclohex-3-en-1-yl)carbamate (I-40): To a solution of tert-butyl(4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)cyclohex-3-en-1-yl)carbamate (100 mg, 0.31 mmol) in dioxane (1.5 mL) was added 2,5-dibromo-1,3,4-thiadiazole (150 mg, 0.62 mmol), Tetrakis(triphenylphosphine)palladium (36 mg, 0.031 mmol), and aqueous sodium carbonate (2M, 0.3 mL, 0.6 mmol). The resulting slurry was degassed with argon for 2 min, sealed, and heated in a microwave at 150° C. for 2.5 hours. The resulting mixture was filtered through celite and washed with ethyl acetate. The combine filtrates were concentrated to dryness. The crude residue was dissolved in DMF and purified by RP-HPLC (eluent: water/MeCN*0.1% TFA) to yield I-40 as a trifluoroacetate salt.

Preparation of Intermediate I-41

N-((1s,4s)-4-(1,3,4-thiadiazol-2-yl)cyclohexyl)-2-hydroxy-2-methylpropanamide(I-41): To a slurry of (1s,4s)-4-(1,3,4-thiadiazol-2-yl)cyclohexan-1-amine hydrochloride (100 mg, 0.46 mmol) in DMF (1 mL) was added 2-Hydroxyisobutyric acid (60.0 mg, 0.576 mmol), HATU (190 mg, 0.5 mmol), and N,N-diisopropylethylamine (0.4 mL, 2.3 mmol). The reaction was stirred at room temperature for 16 h and concentrated to dryness. The resulting crude residue was purified by silica gel chromatography (eluent: MeOH/CH2Cl2) to give I-41.

Preparation of Intermediate I-42

(S)-(1-(5-bromo-1,3,4-thiadiazol-2-yl)pyrrolidin-2-yl)methanol (1-42): To a solution of methyl(2S)-1-(5-bromo-1,3,4-thiodiazol-2-yl)pyrrolidine-2-carboxylate (64.74 mg, 0.221 mmol) in tetrahydrofuran (0.55 mL), lithium chloride (28.2 mg, 0.664 mmol) and sodium borohydride (25.1 mg, 0.664 mmol) were added at rt followed by addition of ethanol (1 mL), and the mixture was stirred at the same temperature as above for 24 hours. Water (0.5 mL) was carefully added thereto, and the mixture was concentrated under reduced pressure. Purified on silica using 30%-100% Hexanes/EtOAc, then flushed with 100% MeOH. Compound containing fractions were concentrated to yield 1-42.

Preparation of Intermediate I-43

(S)-1-(5-bromo-1,3,4-thiadiazol-2-yl)-N-methylpyrrolidine-2-carboxamide(1-43): To a solution of methyl(2S)-1-(5-bromo-1,3,4-thiodiazol-2-yl)pyrrolidine-2-carboxylate (38.0 mg, 0.130 mmol) in methanol (0.650 mL), methylamine hydrochloride (13.2 mg, 0.195 mmol) was added at rt. The mixture was stirred at the same temperature as above for 24 hours. Saturated sodium bicarbonate solution (5 mL) was carefully added and the mixture was extracted twice with 10 mL of EtOAc, washed with 5 mL Brine, and dried over MgSO4. The mixture was filtered and concentrated under reduced pressure to provide 1-43 which was used without additional purification.

Preparation of Intermediate I-44

2-hydroxy-2-methyl-N-[4-(1,3,4-thiadiazol-2-yl)-1-bicyclo[2.1.1]hexanyl]propenamide (I-44): To a solution of crude 4-(1,3,4-thiadiazol-2-yl)bicyclo[2.1.1]hexan-1-amine; hydrochloride (45.0 mg, 0.207 mmol) and 2-hydroxy-2-methyl-propanoic acid (23.7 mg, 0.227 mmol) in DMF (0.5 mL) was added N,N-diisopropylethylamine (0.118 mL, 0.661 mmol) followed by the addition of a solution of HATU (82.5 mg, 0.217 mmol) in DMF (0.5 mL). The reaction mixture was stirred at RT overnight then directly purified by RP-HPLC (eluent: water/MeCN*0.1% TFA) to yield I-44 as a trifluoroacetate salt.

Preparation of Intermediate I-45

4-(1-(difluoromethyl)-1H-pyrazol-4-yl)cyclohexan-1-amine hydrochloride: A suspension of tert-butyl(4-(1-(difluoromethyl)-1H-pyrazol-4-yl)cyclohex-3-en-1-yl)carbamate (0.23 g, 0.75 mmol) in EtOH (15 mL) was degassed with argon and vacuum. Pd/C (10%, 91 mg, 0.086 mmol) was added and the mixture was stirred with a balloon of H2 overnight. The reaction was filtered over a Celite plug, rinsed with EtOAc and the filtrate was concentrated to give tert-butyl(4-(1-(difluoromethyl)-1H-pyrazol-4-yl)cyclohexyl)carbamate which was carried forward without further purification assuming quantitative yield. To a solution of tert-butyl(4-(1-(difluoromethyl)-1H-pyrazol-4-yl)cyclohexyl)carbamate (0.24 g, 0.75 mmol) in DCM (6 mL) was added HCl (4.0 M in dioxane, 3 mL, 12 mmol) and the resulting solution stirred at room temperature for 16 h. Upon completion the reaction mixture was concentrated to dryness to give 4-(1-(difluoromethyl)-1H-pyrazol-4-yl)cyclohexan-1-amine hydrochloride (I-9) which was used without further purification.

2-bromo-N-((1r,4r)-4-(1-(difluoromethyl)-1H-pyrazol-4-yl)cyclohexyl)pyridin-4-amine: To a solution of 2-bromo-4-fluoropyridine (0.23 g, 1.29 mmol) in NMP (7 mL) was added 4-(1-(difluoromethyl)-1H-pyrazol-4-yl)cyclohexan-1-amine hydrochloride (0.38 g, 1.51 mmol) and N,N-diisopropylethylamine (0.70 mL, 4.02 mmol). The resulting mixture was heated for 1 h at 160° C. in a microwave after which the reaction contents diluted with EtOAc, washed 3 times with 5% aqueous LiCl, dried and concentrated to give a crude residue which was purified by normal phase SiO2chromatography (eluent: ethyl acetate/hexanes) to give both the cis and trans products. The trans product 2-bromo-N-((1r,4r)-4-(1-(difluoromethyl)-1H-pyrazol-4-yl)cyclohexyl)pyridin-4-amine was isolated and carried forward.

7-(5-bromo-4-(((1r,4r)-4-(1-(difluoromethyl)-1H-pyrazol-4-yl)cyclohexyl)amino)pyridin-2-yl)pyrrolo[1,2-b]pyridazine-3-carbonitrile(I-45):2-bromo-N-((1r,4r)-4-(1-(difluoromethyl)-1H-pyrazol-4-yl)cyclohexyl)pyridin-4-amine was elaborated to the final intermediate 7-(5-bromo-4-(((1r,4r)-4-(1-(difluoromethyl)-1H-pyrazol-4-yl)cyclohexyl)amino)pyridin-2-yl)pyrrolo[1,2-b]pyridazine-3-carbonitrile (I-45) in the same manner as described in steps 2-3 of Preparation of Intermediate I-2.

EXAMPLE PROCEDURES AND COMPOUND EXAMPLES

The following compounds were made with the procedures herein, using the appropriate starting materials and protecting group chemnistry as needed:

(5-(6-chloro-4-(isopropylamino)pyridin-3-yl)-1,3,4-thiadiazol-2-yl)methanol: To a solution of methyl 5-(6-chloro-4-(isopropylamino)pyridin-3-yl)-1,3,4-thiadiazole-2-carboxylate (50.0 mg, 0.16 mmol) in THF (0.5 mL) was added lithium borohydride (2M in THF, 80 μL, 0.16 mmol). The reaction mixture was stirred at room temperature for 15 minutes, then quenched with a few drops of saturated aqueous ammonium chloride. The reaction mixture was concentrated in vacuo and purified by silica gel column chromatography (eluent: MeOH/DCM) to provide (5-(6-chloro-4-(isopropylamino)pyridin-3-yl)-1,3,4-thiadiazol-2-yl)methanol.

The following compounds were made according to the procedures herein, using the appropriate starting materials and protecting group chemistry as needed:

(S)-(5-(6-chloro-4-(isopropylamino)pyridin-3-yl)-1,3,4-thiadiazol-2-yl)(3-hydroxypyrrolidin-1-yl)methanonea: To a solution of methyl 5-(6-chloro-4-(isopropylamino)pyridin-3-yl)-1,3,4-thiadiazole-2-carboxylate (50.0 mg, 0.16 mmol) in MeCOH (0.5 mL) was added (S)-3-hydroxypyrrolidine (13 μL, 0.16 mmol). The reaction mixture was heated at 80° C. for 1 hour, then concentrated in vacuo and purified by silica gel column chromatography (eluent: MeOH/DCM) to provide (S)-(5-(6-chloro-4-(isopropylamino)pyridin-3-yl)-1,3,4-thiadiazol-2-yl)(3-hydroxypyrrolidin-1-yl)methanonea.

The following compounds were made according to the procedures herein, using the appropriate starting materials and protecting group chemistry as needed:

N′-acetyl-6-chloro-4-(isopropylamino)nicotinohydrazide: A solution of 6-chloro-4-(isopropylamino)nicotinic acid (0.2 g, 0.93 mmol), acetohydrazide (138.05 mg, 1.86 mmol), HATU (425.1 mg, 1.12 mmol), and N,N-Diisopropylethylamine (0.41 ml, 2.33 mmol) in DMF (1.8 mL) was stirred at room temperature for 18 hours. The reaction solution was diluted in ethyl acetate and washed three times with aqueous saturated ammonium chloride solution. The organic extract was dried over sodium sulfate, filtered, and concentrated to give N′-acetyl-6-chloro-4-(isopropylamino)nicotinohydrazide

2-chloro-N-isopropyl-5-(5-methyl-1,3,4-thiadiazol-2-yl)pyridin-4-amine: A suspension of N′-acetyl-6-chloro-4-(isopropylamino)pyridine-3-carbohydrazide (0.13 g, 0.46 mmol) and Lawesson's reagent (0.21 g, 0.51 mmol) in 1,4-dioxane (2.3 mL) was heated at 80° C. for 18 hours in an aluminum heating block. The crude reaction mixture was purified by normal phase silica gel chromatography (eluent: EtOAc/hexanes) to give 2-chloro-N-isopropyl-5-(5-methyl-1,3,4-thiadiazol-2-yl)pyridin-4-amine.

7-(5-(5-(2-aminoethyl)-1,3,4-thiadiazol-2-yl)-4-(isopropylamino)pyridin-2-yl)pyrrolo[1,2-b]pyridazine-3-carbonitrile (Example 26): To a solution of tert-butyl(2-(5-(6-(3-cyanopyrrolo[1,2-b]pyridazin-7-yl)-4-(isopropylamino)pyridin-1-3-yl)-1,3,4-thiadiazol-2-yl)ethyl)carbamate (Example 31) (obtained as described in Procedure 1) (4.6 mg, 0.0091 mmol) in 1,4-dioxane (0.25 mL) was added HCl (4M in 1,4-dioxane, 11.0 μL, 0.046 mmol). The reaction mixture was heated to 30° C. for 30 minutes, then concentrated in vacuo and purified by reverse phase high pressure liquid chromatography (eluent: water/MeCN*0.1% TFA) to provide the final compound Example 26.

The following compounds were made according to the procedures herein, using the appropriate starting materials and protecting group chemistry as needed:

2-chloro-N-isopropyl-5-(1,3,4-thiadiazol-2-yl)pyridin-4-amine: To a solution of methyl 5-(6-chloro-4-(isopropylamino)pyridin-3-yl)-1,3,4-thiadiazole-2-carboxylate (50.0 mg, 0.16 mmol) in 1,4-dioxane (2.0 mL) was added potassium carbonate (2.4M in water, 0.20 mL, 0.48 mmol). The reaction mixture was heated at 110° C. for 2 hours. The reaction was concentrated and partitioned between water and EtOAc. The organic layer was isolated, and the aqueous layer was extracted with two additional portions of EtOAc. The organic layers were combined, dried over sodium sulfate, filtered, and concentrated to provide 2-chloro-N-isopropyl-5-(1,3,4-thiadiazol-2-yl)pyridin-4-amine which was used without additional purification.

Methyl 4-(2-(6-chloro-4-(isopropylamino)nicotinoyl)hydrazinyl)-4-oxobutanoate: To a solution of known compound 6-chloro-4-(isopropylamino)nicotinohydrazide (2 g, 8.75 mmol) in dichoromethane (100 mL) was added triethylamine (3.66 mL, 26.2 mmol), and the reaction was cooled to 0° C. To the reaction was added methyl 4-chloro-4-oxobutanoate (1.72 mL, 14 mmol) dropwise, and the resulting mixture stirred at room temperature for 5 hours. Afterward, saturated aqueous NaHCO3was added, and layers were separated. The organic layer was extracted with dichloromethane. The combined organic layers were dried over MgSO4, filtered and concentrated. The resulting crude residue was purified by silica gel chromatography (eluent: EtOAc/hexanes, then MeOH/EtOAc) to give the desired product.

Methyl 3-(5-(6-chloro-4-(isopropylamino)pyridin-3-yl)-1,3,4-thiadiazol-2-yl)propanoate: To a solution of methyl 4-(2-(6-chloro-4-(isopropylamino)nicotinoyl)hydrazinyl)-4-oxobutanoate (710 mg, 2.07 mmol) in THF (50 mL) was added Lawesson's reagent (1.68 g, 4.14 mmol) in one portion, and the reaction was stirred at reflux for 90 minutes. Afterward, the reaction was cooled, and the mixture was diluted with EtOAc, and the organics were washed twice with 50% (by volume) aqueous NaHCO3. The organic layer was dried over MgSO4, filtered and concentrated. The resulting crude residue was purified by silica gel chromatography (eluent: EtOAc/hexanes) to give the desired product.

4-(5-(6-chloro-4-(isopropylamino)pyridin-3-yl)-1,3,4-thiadiazol-2-yl)-2-methylbutan. 2-ol: To a flask with methyl 3-(5-(6-chloro-4-(isopropylamino)pyridin-3-yl)-1,3,4-thiadiazol-2-yl)propanoate (216 mg, 0.63 mmol) under N2was added THF (7 mL), and the reaction was cooled to 0° C. To the reaction was added methylmagnesium bromide (3M solution in Et2O, 1.48 mL, 4.44 mmol) dropwise, and the reaction was stirred for 30 min at 0° C. Afterward, the reaction was quenched by dropwise addition of saturated aqueous NH4Cl, and the mixture was diluted with EtOAc and water. The layers were separated, and the aqueous layer was washed once with EtOAc. The combined organic layers were dried over MgSO4, filtered and concentrated. The resulting crude residue was purified by silica gel chromatography (eluent: EtOAc/hexanes) to give the desired product.

7-(5-(5-(3-hydroxy-3-methylbutyl)-1,3,4-thiadiazol-2-yl)-4-(isopropylamino)pyridin-2-yl)pyrrolo[1,2-b]pyridazine-3-carbonitrile (Example 31): To a microwave vial was added 4-(5-(6-chloro-4-(isopropylamino)pyridin-3-yl)-1,3,4-thiadiazol-2-yl)-2-methylbutan-2-ol (50 mg, 0.15 mmol), 7-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)pyrrolo[1,2-b]pyridazine-3-carbonitrile (I-8) (55 mg, 0.21 mmol), [1,1′-Bis(diphenylphosphino)ferrocene]dichloropalladium(II) (21.8 mg, 0.029 mmol). To the vial was added DME (1.0 mL) and sodium carbonate (2M solution in water, 0.15 mL, 0.29 mmol). The mixture was degassed with argon for 1 min, sealed, and heated under microwave conditions for min at 120° C. Afterward, the vial was cooled and the mixture was concentrated under vacuum. The crude material was diluted with DMF (1 mL). This mixture was filtered, and purified by RP-HPLC (eluent: water/MeCN*0.1% TFA) to yield the product Example 31 as a trifluoroacetate salt.

The following compounds were made according to the procedures set forth previously, using the appropriate starting materials and protecting group chemistry as needed:

The following compounds were made according to the previous procedures, using the appropriate starting materials and protecting group chemistry as needed:

N-((1R,5S,8s)-3-(5-(6-chloro-4-fluoropyridin-3-yl)-1,3,4-thiadiazol-2-yl)-3-azabicyclo[3.2.1]octan-8-yl)acetamide: A microwave vial was charged with N-((1R,5S,8s)-3-(5-bromo-1,3,4-thiadiazol-2-yl)-3-azabicyclo[3.2.1]octan-8-yl)acetamide (200 mg, 0.60 mmol), 2-chloro-4-fluoro-5-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)pyridine (171 mg, 0.66 mmol), palladium(II) acetate (20.3 mg, 0.091 mmol), Xantphos (105 mg, 0.18 mmol), and cesium carbonate (590 mg, 0.18 mmol). Dioxane (3.0 mL) was added, and the reaction mixture was degassed by bubbling argon for 60 seconds. The vial was sealed and heated with stirring at 120° C. for 20 minutes in a microwave reactor. The cooled reaction mixture was filtered through a pad of Celite with EtOAc, concentrated in vacuo, and used without additional purification.

(R)-2-((5-(5-((1R,5S,8S)-8-acetamido-3-azabicyclo[3.2.1]octan-3-yl)-1,3,4-thiadiazol-2-yl)-2-chloropyridin-4-yl)amino)propenamide: To a solution of crude N-((1R,5S,8s)-3-(5-(6-chloro-4-fluoropyridin-3-yl)-1,3,4-thiadiazol-2-yl)-3-azabicyclo[3.2.1]octan-8-yl)acetamide (231 mg, 0.60 mmol) in butyronitrile (3.0 mL) was added D-alaninamide hydrochloride (90.3 mg, 0.73 mmol) and N,N-diisopropylethylamine (0.47 mL, 2.7 mmol). The reaction was heated at 130° C. for 45 minutes. The cooled reaction was concentrated in vacuo and used without additional purification.

N-((1R,5S,8S)-3-(5-(6-chloro-4-(((R)-1-cyanoethyl)amino)pyridin-3-yl)-1,3,4-thiadiazol-2-yl)-3-azabicyclo[3.2.1]octan-8-yl)acetamide: A solution of crude (R)-2-((5-(5-((1R,5S,8S)-8-acetamido-3-azabicyclo[3.2.1]octan-3-yl)-1,3,4-thiadiazol-2-yl)-2-chloropyridin-4-yl)amino)propanamide (272 mg, 0.60 mmol) in THF (3.0 mL) was cooled to 0° C. To the solution was added pyridine (0.24 mL, 3.0 mmol) and trifluoroacetic anhydride (0.13 mL, 0.91 mmol). The reaction was allowed to warm to room temperature while stirred for 25 minutes. The reaction mixture was concentrated in vacuo and purified by silica gel column chromatography (eluent: MeOH/DCM) to provide the desired product.

N-((1R,5S,8S)-3-(5-(4-(((R)-1-cyanoethyl)amino)-6-(3-cyanopyrrolo[1,2-b]pyridazin-7-yl)pyridin-3-yl)-1,3,4-thiadiazol-2-yl)-3-azabicyclo[3.2.1]octan-8-yl)acetamide (Example 53): A microwave vial was charged with N-((1R,5S,8S)-3-(5-(6-chloro-4-(((R)-1-cyanoethyl)amino)pyridin-3-yl)-1,3,4-thiadiazol-2-yl)-3-azabicyclo[3.2.1]octan-8-yl)acetamide (84.0 mg, 0.19 mmol), 7-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)pyrrolo[1,2-b]pyridazine-3-carbonitrile (Intermediate I-8, 78.5 mg, 0.29 mmol), and [1,1′-Bis(diphenylphosphino)ferrocene]dichloropalladium(II) (28.9 mg, 0.039 mmol). To the flask was added DME (4.0 mL) and a 2M aqueous solution of sodium carbonate (0.39 mL, 0.78 mmol). The reaction mixture was degassed by bubbling argon for 60 seconds. The vial was sealed and heated with stirring at 120° C. for 15 minutes in a microwave reactor. The cooled reaction mixture was concentrated and purified by RP-HPLC (eluent: water/MeCN*0.1% TFA) to yield the product Example 53 as a trifluoroacetate salt.

The following compounds were made according to the previous procedures, using the appropriate starting materials and protecting group chemistry as needed:

N-((1R,5S,8S)-3-(5-(6-(2-(((1R,2S)-2-aminocyclohexyl)amino)pyrrolo[2,1-f][1,2,4]triazin-7-yl)-4-(((R)-1-cyanoethyl)amino)pyridin-3-yl)-1,3,4-thiadiazol-2-yl)-3-azabicyclo[3.2.1]octan-8-yl)acetamide (Example 61): To a solution of tert-butyl((1S,2R)-2-((7-(5-(5-((1R,5S,8S)-8-acetamido-3-azabicyclo[3.2.1]octan-3-yl)-1,3,4-thiadiazol-2-yl)-4-(((R)-1-cyanoethyl)amino)pyridin-2-yl)pyrrolo[2,1-f][1,2,4]triazin-2-yl)amino)cyclohexyl)carbamate (synthesized as described in Example P-GJB2 using the appropriate boronate ester in place of I-8) (25.2 mg, 0.035 mmol) in DCM (1.0 mL) was added trifluoroacetic acid (0.5 mL, 7.0 mmol). The reaction mixture was stirred at RT for 20 minutes, then concentrated in vacuo and purified by RP-HPLC (eluent: water/MeCN*0.1% TFA) to yield the product Example 61 as a trifluoroacetate salt.

The following compounds were made according to the previous procedures, using the appropriate starting materials and protecting group chemistry as needed:

(R)-2-((2-chloro-5-iodopyridin-4-yl)amino)propenamide: To a solution of 2-chloro-4-fluoro-5-iodo-pyridine (3.50 g, 13.6 mmol) and D-alaninamide hydrochloride (4.23 g, 68.0 mmol) in NMP (30.0 mL) was added N,N-diisopropylethylamine (11.8 mL, 68.0 mmol). The reaction mixture was heated to 150° C. overnight, then cooled and diluted with water (125 mL). The solution was stirred at RT for 2 hours, then the resulting solid was isolated by vacuum filtration, washed with water, and dried by high vacuum. The crude solids were used without additional purification.

(R)-2-((5-(5-(4-acetamidobicyclo[2.2.2]octan-1-yl)-1,3,4-thiadiazol-2-yl)-2-chloropyridin-4-yl)amino)propenamide: (R)-2-((5-(5-(4-acetamidobicyclo[2.2.2]octan-1-yl)-1,3,4-thiadiazol-2-yl)-2-chloropyridin-4-yl)amino)propenamide was prepared following the protocol in Procedure 1, using the appropriate coupling partners.

(R)—N-(4-(5-(6-chloro-4-((1-cyanoethyl)amino)pyridin-3-yl)-1,3,4-thiadiazol-2-yl)bicyclo[2.2.2]octan-1-yl)acetamide: (R)—N-(4-(5-(6-chloro-4-((1-cyanoethyl)amino)pyridin-3-yl)-1,3,4-thiadiazol-2-yl)bicyclo[2.2.2]octan-1-yl)acetamide was prepared following the protocol in step 3 of Procedure 11.

(R)—N-(4-(5-(4-((1-cyanoethyl)amino)-6-(3-cyanopyrrolo[1,2-b]pyridazin-7-yl)pyridin-3-yl)-1,3,4-thiadiazol-2-yl)bicyclo[2.2.2]octan-1-yl)acetamide (Example 72): Example 72 was prepared following the protocol in step 4 of Procedure 11 using the appropriate coupling partners to provide Example 72 as a trifluoroacetate salt.

The following compounds were made according to the previous procedures, using the appropriate starting materials and protecting group chemistry as needed:

tert-butyl(1-(2-(6-chloro-4-(isopropylamino)nicotinoyl)hydrazine-1-carbonyl)-2-oxabicyclo[2.2.2]octan-4-yl)carbamate: To a solution of 6-chloro-4-(isopropylamino)nicotinohydrazide (123 mg, 0.54 mmol), 4-((tert-butoxycarbonyl)amino)-2-oxabicyclo[2.2.2]octane-1-carboxylic acid (175 mg, 0.65 mmol), and HATU (215 mg, 0.57 mmol) in DMF (0.5 mL) was added N,N-diisopropylethylamine (0.31 mL, 1.72 mmol). The reaction mixture was stirred at RT for 15 minutes, then diluted with water (10 mL) and extracted with EtOAc (2×10 mL). The combined organic layers were dried over sodium sulfate, isolated by vacuum filtration, concentrated in vacuo, and purified by silica gel column chromatography (eluent: MeOH/DCM) to provide the desired product.

tert-butyl(1-(5-(6-chloro-4-(isopropylamino)pyridin-3-yl)-1,3,4-thiadiazol-2-yl)-2-oxabicyclo[2.2.2]octan-4-yl)carbamate: To a solution of tert-butyl(1-(2-(6-chloro-4-(isopropylamino)nicotinoyl)hydrazine-1-carbonyl)-2-oxabicyclo[2.2.2]octan-4-yl)carbamate (240 mg, 0.50 mmol) in THF (6.0 mL) at 65° C. was added Lawesson's Reagent (302 mg, 0.75 mmol). The RM was stirred at 65° C. for 15 minutes, then concentrated in vacuo and purified by silica gel column chromatography (eluent: MeOH/DCM) to provide the desired product.

N-(1-(5-(6-chloro-4-(isopropylamino)pyridin-3-yl)-1,3,4-thiadiazol-2-yl)-2-oxabicyclo[2.2.2]octan-4-yl)acetamide: Tert-butyl(1-(5-(6-chloro-4-(isopropylamino)pyridin-3-yl)-1,3,4-thiadiazol-2-yl)-2-oxabicyclo[2.2.2]octan-4-yl)carbamate (152 mg, 0.32 mmol) was suspended in HCl (4M in dioxane, 0.8 mL, 3.17 mmol) and heated with stirring at 40° C. for 2 hours. The reaction mixture was cooled and concentrated in vacuo. The resulting residue was suspended in DCM (2.0 mL). To the suspension was added triethylamine (60 μL, 0.43 mmol) and acetic anhydride (11.2 μL, 0.12 mmol). The reaction mixture was stirred at RT for 15 minutes, then concentrated in vacuo and purified by silica gel column chromatography (eluent: MeOH/DCM) to provide the desired product.

N-(1-(5-(6-(3-cyanopyrrolo[1,2-b]pyridazin-7-yl)-4-(isopropylamino)pyridin-3-yl)-1,3,4-thiadiazol-2-yl)-2-oxabicyclo[2.2.2]octan-4-yl)acetamide (Example 83): Example 83 was prepared following the protocol in step 4 of Procedure 11 using the appropriate coupling partners to provide Example 83 as a trifluoroacetate salt.

The following compounds were made according to the previous procedures, using the appropriate starting materials and protecting group chemistry as needed:

2-((1,2-trans)-2-(5-(6-chloro-4-(isopropylamino)pyridin-3-yl)-1,3,4-thiadiazol-2-yl)cyclobutyl)propan-2-ol: To a solution of methyl(1,2-trans)-2-(5-(6-chloro-4-(isopropylamino)pyridin-3-yl)-1,3,4-thiadiazol-2-yl)cyclobutane-1-carboxylate (prepared following the protocol in steps 1 and 2 of Procedure 14 using the appropriate carboxylic acid) (100 mg, 0.24 mmol) in THF (0.5 mL) at 0° C. was added methylmagnesium bromide (3M in diethyl ether, 0.12 mL, 0.37 mmol). The reaction mixture was allowed to warm to RT with stirring over 30 minutes, then quenched with saturated aqueous ammonium chloride. The reaction was partitioned between water and EtOAc, and the aqueous layer was extracted two additional times with EtOAc. The combined organic layers were dried over sodium sulfate, isolated by vacuum filtration, concentrated in vacuo, and purified by silica gel column chromatography (eluent: MeOH/DCM) to provide the desired product.

7-(5-(5-((1,2-trans)-2-(2-hydroxypropan-2-yl)cyclobutyl)-1,3,4-thiadiazol-2-yl)-4-(isopropylamino)pyridin-2-yl)pyrrolo[1,2-b]pyridazine-3-carbonitrile (Example 88): Example 88 was prepared following the protocol in step 4 of Procedure 11 using the appropriate coupling partners to provide Example 88 as a trifluoroacetate salt and a mixture of trans-stereoisomers.

The following compound was made according to Procedure 9, using the appropriate starting materials and protecting group chemistry as needed:

tert-butyl((1R,5S,8s)-3-(5-(6-(3-cyanopyrrolo[1,2-b]pyridazin-7-yl)-4-(isopropylamino)pyridin-3-yl)-1,3,4-thiadiazol-2-yl)-3-azabicyclo[3.2.1]octan-8-yl)carbamate: Tert-butyl((1R,5S,8s)-3-(5-(6-(3-cyanopyrrolo[1,2-b]pyridazin-7-yl)-4-(isopropylamino)pyridin-3-yl)-1,3,4-thiadiazol-2-yl)-3-azabicyclo[3.2.1]octan-8-yl)carbamate was prepared from tert-butyl((1R,5S,8s)-3-(5-(6-chloro-4-(isopropylamino)pyridin-3-yl)-1,3,4-thiadiazol-2-yl)-3-azabicyclo[3.2.1]octan-8-yl)carbamate (prepared following the protocol in steps 1 and 2 of Procedure 11 using the appropriate 2-bromo-thiadiazole in step 1 and the appropriate amine in step 2) following the protocol in step 4 of Procedure 11.

(R)—N-((1R,5S,8s)-3-(5-(6-(3-cyanopyrrolo[1,2-b]pyridazin-7-yl)-4-(isopropylamino)pyridin-3-yl)-1,3,4-thiadiazol-2-yl)-3-azabicyclo[3.2.1]octan-8-yl)-2-hydroxypropanamide (Example 90): To a solution of 7-(5-(5-((1R,5S,8s)-8-amino-3-azabicyclo[3.2.1]octan-3-yl)-1,3,4-thiadiazol-2-yl)-4-(isopropylamino)pyridin-2-yl)pyrrolo[1,2-b]pyridazine-3-carbonitrile hydrochloride (12.0 mg, 0.023 mmol) and (R)-2-hydroxypropanoic acid (2.7 mg, 0.03 mmol) in DMF (0.5 mL) was added N,N-diisopropylethylamine (20.5 μL, 0.12 mmol). To the solution was added HATU (13.1 mg, 0.035 mmol) in DMF (0.5 mL). The reaction mixture was stirred at RT for 15 minutes then purified by RP-HPLC (eluent: water/MeCN*0.1% TFA) to yield the product Example 90 as a trifluoroacetate salt.

The following compounds were made according to Procedure 16, using the appropriate starting materials and protecting group chemistry as needed:

N-((1R,5S,8s)-3-(5-(6-(3-cyanopyrrolo[1,2-b]pyridazin-7-yl)-4-methylpiperazine-1-carboxamide (Example 96): To a suspension of 7-(5-(5-((1R,5S,8s)-8-amino-3-azabicyclo[3.2.1]octan-3-yl)-1,3,4-thiadiazol-2-yl)-4-(isopropylamio)pyridin-2-yl)pyrrolo[1,2-b]pyridazine-3-carbonitrile hydrochloride (synthesized following the protocol in Procedure 16) (12.0 mg, 0.023 mmol) in DMF (0.5 mL) was added N,N-diisopropylethylamine (20.5 μL, 0.12 mmol). To the resulting solution was added a solution of 4-methylpiperazine-1-carbonyl chloride hydrochloride (6.0 mg, 0.030 mmol) and N,N-diisopropylethylamine (8.2 μL, 0.046 mmol) in DMF (0.5 mL). The reaction mixture was stirred at RT for 3 hours then directly purified by RP-HPLC (eluent: water/MeCN*0.1% TFA) to yield the product Example 96 as a trifluoroacetate salt.

The following compounds were made according to the previous procedures, using the appropriate starting materials and protecting group chemistry as needed:

N-((1R,5S,8s)-3-(5-(6-(3-cyanopyrrolo[1,2-b]pyridazin-7-yl)-4-(isopropylamino)pyridin-3-yl)-1,3,4-thiadiazol-2-yl)-3-azabicyclo[3.2.1]octan-8-yl)piperazine-1-carboxamide hydrochloride: Tert-butyl 4-(((1R,5S,8s)-3-(5-(6-(3-cyanopyrrolo[1,2-b]pyridazin-7-yl)-4-(isopropylamino)pyridin-3-yl)-1,3,4-thiadiazol-2-yl)-3-azabicyclo[3.2.1]octan-8-yl)carbamoyl)piperazine-1-carboxylate (synthesized following the protocol in Procedure 2 using the appropriate coupling partners) (130 mg, 0.19 mmol) was stirred in a solution of HCl (4M in 1,4-dioxane, 0.93 mL, 3.73 mmol) at 40° C. for 1 hour. The reaction mixture was concentrated and the resulting solid used without purification.

4-acetyl-N-((1R,5S,8s)-3-(5-(6-(3-cyanopyrrolo[1,2-b]pyridazin-7-yl)-4-(isopropylamino)pyridin-3-yl)-1,3,4-thiadiazol-2-yl)-3-azabicyclo[3.2.1]octan-8-yl)piperazine-1-carboxamide (Example 104): To a suspension of crude N-((1R,5S,8s)-3-(5-(6-(3-cyanopyrrolo[1,2-b]pyridazin-7-yl)-4-(isopropylamino)pyridin-3-yl)-1,3,4-thiadiazol-2-yl)-3-azabicyclo[3.2.1]octan-8-yl)piperazine-1-carboxamide hydrochloride (10.0 mg, 0.016 mmol) in DMF was added triethylamine (8.8 μL, 0.063 mmol) followed by acetic anhydride (1.8 μL, 0.019 mmol). The reaction mixture was stirred at RT for 15 minutes, then directly purified by RP-HPLC (eluent: water/MeCN*0.1% TFA) to yield the product Example 104 as a trifluoroacetate salt.

The following compounds were made according to Procedure 18, using the appropriate starting materials and protecting group chemistry as needed:

7-(5-(5-(3,6-diazabicyclo[3.1.1]heptan-3-yl)-1,3,4-thiadiazol-2-yl)-4-(isopropylamino)pyridin-2-yl)pyrrolo[1,2-b]pyridazine-3-carbonitrile: A solution of tert-butyl 3-(5-(6-(3-cyanopyrrolo[1,2-b]pyridazin-7-yl)-4-(isopropylamino)pyridin-3-yl)-1,3,4-thiadiazol-2-yl)-3,6-diazabicyclo[3.1.1]heptane-6-carboxylate (synthesized following the protocol in Procedure 2 using the appropriate coupling partners) (153 mg, 0.28 mmol) in 1,1,1,3,3,3-hexafluoro-2-propanol (1.9 mL, 17.9 mmol) was heated in a microwave reactor 30 minutes at 150° C. The cooled reaction was concentrated in vacuo to provide the desired product which was used without purification.

7-(5-(5-(6-(2-((dimethyl(oxo)-λ6-sulfanylidene)amino)acetyl)-3,6-diazabicyclo[3.1.1]heptan-3-yl)-1,3,4-thiadiazol-2-yl)-4-(isopropylamino)pyridin-2-yl)pyrrolo[1,2-b]pyridazine-3-carbonitrile (Example 107): To a solution of crude 7-(5-(5-(3,6-diazabicyclo[3.1.1]heptan-3-yl)-1,3,4-thiadiazol-2-yl)-4-(isopropylamino)pyridin-2-yl)pyrrolo[1,2-b]pyridazine-3-carbonitrile (15.0 mg, 0.033 mmol) and 2-((dimethyl(oxo)-λ6-sulfanylidene)amino)acetic acid (6.4 mg, 0.043 mmol) in DMF (0.5 mL) was added N,N-diisopropylethylamine (23 μL, 0.13 mmol) followed by the addition of a solution of HATU (18.7 mg, 0.049 mmol) in DMF (0.5 mL). The reaction mixture was stirred at RT for 15 minutes then directly purified by RP-HPLC (eluent: water/MeCN*0.1% TFA) to yield the product Example 107 as a trifluoroacetate salt.

The following compounds were made according to the previous procedures, using the appropriate starting materials and protecting group chemistry as needed:

The following compounds were made according to Procedure 19, using the appropriate starting materials and protecting group chemistry as needed:

tert-butyl(4-(5-(6-(3-cyanopyrrolo[1,2-b]pyridazin-7-yl)-4-(isopropylamino)pyridin-3-yl)-1,3,4-thiadiazol-2-yl)bicyclo[2.2.2]octan-1-yl)carbamate:Tert-butyl(4-(5-(6-(3-cyanopyrrolo[1,2-b]pyridazin-7-yl)-4-(isopropylamino)pyridin-3-yl)-1,3,4-thiadiazol-2-yl)bicyclo[2.2.2]octan-1-yl)carbamate was synthesized from tert-butyl(4-(5-(6-chloro-4-(isopropylamino)pyridin-3-yl)-1,3,4-thiadiazol-2-yl)bicyclo[2.2.2]octan-1-yl)carbamate (synthesized following the protocol in steps 1 and 2 of Procedure 14 using the appropriate carboxylic acid) following the protocol in step 4 of Procedure 14.

7-(5-(5-(4-aminobicyclo[2.2.2]octan-1-yl)-1,3,4-thiadiazol-2-yl)-4-(isopropylamino)pyridin-2-yl)pyrrolo[1,2-b]pyridazine-3-carbonitrile: A solution of tert-butyl(4-(5-(6-(3-cyanopyrrolo[1,2-b]pyridazin-7-yl)-4-(isopropylamino)pyridin-3-yl)-1,3,4-thiadiazol-2-yl)bicyclo[2.2.2]octan-1-yl)carbamate in 1,1,1,3,3,3-hexafluoro-2-propanol was heated at 150° C. in a microwave reactor for 90 minutes. The cooled reaction mixture was concentrated to provide the desired product which was used without purification.

N-(4-(5-(6-(3-cyanopyrrolo[1,2-b]pyridazin-7-yl)-4-(isopropylamino)pyridin-3-yl)-1,3,4-thiadiazol-2-yl)bicyclo[2.2.2]octan-1-yl)-2-hydroxy-2-methylpropanamide (Example 120): To a solution of crude 7-(5-(5-(4-aminobicyclo[2.2.2]octan-1-yl)-1,3,4-thiadiazol-2-yl)-4-(isopropylamino)pyridin-2-yl)pyrrolo[1,2-b]pyridazine-3-carbonitrile (10.0 mg, 0.021 mmol) and 2-hydroxy-2-methylpropanoic acid (2.8 mg, 0.027 mmol) in DMF (0.5 mL) was added N,N-diisopropylethylamine (14.7 μL, 0.083 mmol) followed by a solution of HATU (11.8 mg, 0.031 mmol) in DMF (0.5 mL). The reaction mixture was stirred at RT for 15 minutes then directly purified by RP-HPLC (eluent: water/MeCN*0.1% TFA) to yield the product Example 120 as a trifluoroacetate salt.

The following compounds were made according to Procedure 22, using the appropriate starting materials and protecting group chemistry as needed:

7-(5-(5-((1R,5S,8s)-8-amino-3-azabicyclo[3.2.1]octan-3-yl)-1,3,4-thiadiazol-2-yl)-4-(isopropylamino)pyridin-2-yl)pyrrolo[1,2-b]pyridazine-3-carbonitrile hydrochloride: Tert-butyl((1R,5S,8s)-3-(5-(6-(3-cyanopyrrolo[1,2-b]pyridazin-7-yl)-4-(isopropylamino)pyridin-3-yl)-1,3,4-thiadiazol-2-yl)-3-azabicyclo[3.2.1]octan-8-yl)carbamate (synthesized following the protocol in Procedure 2 using the appropriate coupling partners) (451 mg, 0.77 mmol) was stirred with HCl (4M in 1,4-dioxane, 3.9 mL, 15.4 mmol) at 40° C. for 1 hour. The cooled reaction was concentrated in vacuo to provide the desired product which was used without purification.

tert-butyl 4-(((1R,5S,8s)-3-(5-(6-(3-cyanopyrrolo[1,2-b]pyridazin-7-yl)-4-(isopropylamino)pyridin-3-yl)-1,3,4-thiadiazol-2-yl)-3-azabicyclo[3.2.1]octan-8-yl)carbamoyl)piperidine-1-carboxylate: To a solution of crude 7-(5-(5-((1R,5S,8s)-8-amino-3-azabicyclo[3.2.1]octan-3-yl)-1,3,4-thiadiazol-2-yl)-4-(isopropylamino)pyridin-2-yl)pyrrolo[1,2-b]pyridazine-3-carbonitrile hydrochloride (50.0 mg, 0.096 mmol) and 1-(tert-butoxycarbonyl)piperidine-4-carboxylic acid (65.9 mg, 0.29 mmol) in DMF (0.5 mL) was added N,N-diisopropylamine (0.21 mL) followed by a solution of HATU (142 mg, 0.37 mmol) in DMF (0.5 mL). The reaction mixture was stirred at RT for 15 minutes then partitioned between water and EtOAc. The aqueous layer was extracted with two additional portions of EtOAc. The combined organic layers were dried over sodium sulfate, isolated by vacuum filtration, and purified by silica gel column chromatography (eluent: EtOAc/hexanes) to provide the desired product.

N-((1R,5S,8s)-3-(5-(6-(3-cyanopyrrolo[1,2-b]pyridazin-7-yl)-4-(isopropylamino)pyridin-3-yl)-1,3,4-thiadiazol-2-yl)-3-azabicyclo[3.2.1]octan-8-yl)piperidine-4-carboxamide: A solution of tert-butyl 4-(((1R,5S,8s)-3-(5-(6-(3-cyanopyrrolo[1,2-b]pyridazin-7-yl)-4-(isopropylamino)pyridin-3-yl)-1,3,4-thiadiazol-2-yl)-3-azabicyclo[3.2.1]octan-8-yl)carbamoyl)piperidine-1-carboxylate (135 mg, 0.19 mmol) in 1,1,1,3,3,3-hexafluoro-2-propanol (2.0 mL, 19.4 mmol) was heated in a microwave reactor for 75 minutes at 140° C. The cooled reaction was concentrated to provide the desire product which was used without purification.

1-acetyl-N-((1R,5S,8s)-3-(5-(6-(3-cyanopyrrolo[1,2-b]pyridazin-7-yl)-4-(isopropylamino)pyridin-3-yl)-1,3,4-thiadiazol-2-yl)-3-azabicyclo[3.2.1]octan-8-yl)piperidine-4-carboxamide (Example 123): To a solution of crude N-((1R,5S,8s)-3-(5-(6-(3-cyanopyrrolo[1,2-b]pyridazin-7-yl)-4-(isopropylamino)pyridin-3-yl)-1,3,4-thiadiazol-2-yl)-3-azabicyclo[3.2.1]octan-8-yl)piperidine-4-carboxamide (10.0 mg, 0.017 mmol) in DMF (1.0 mL) was added triethylamine (7.0 μL, 0.050 mmol) followed by acetic anhydride (2.0 μL, 0.022 mmol). The reaction mixture was stirred at RT for 15 minutes then directly purified by RP-HPLC (eluent: water/MeCN*0.1% TFA) to yield the product Example 123 as a trifluoroacetate salt.

The following compounds were made according to Procedure 1, using the appropriate starting materials and protecting group chemistry as needed:

7-(4-(isopropylamino)-5-(5-(piperidin-4-yl)-1,3,4-thiadiazol-2-yl)pyridin-2-yl)pyrrolo[1,2-b]pyridazine-3-carbonitrile (Example 129): To an RBF with tert-butyl 4-(5-(6-(3-cyanopyrrolo[1,2-b]pyridazin-7-yl)-4-(isopropylamino)pyridin-3-yl)-1,3,4-thiadiazol-2-yl)piperidine-1-carboxylate (prepared via Procedure 1) (40 mg, 0.061 mmol) was added 1,4-dioxane (7 mL) and methanol (1.5 mL). HCl (4.0 M in dioxane, 1 mL) was added, and the reaction was stirred at 50° C. for 2 hours. The reaction was then concentrated under reduced pressure, and the crude material was dissolved in DMF, then purified by RP-HPLC (eluent: water/MeCN*0.1% TFA) to yield the product Example 129 as a trifluoroacetate salt.

The following compounds were made according to Procedure 2, using the appropriate starting materials and protecting group chemistry as needed:

±methyl trans-2-(5-(6-(3-cyanopyrrolo[1,2-b]pyridazin-7-yl)-4-(isopropylamino)pyridin-3-yl)-1,3,4-thiadiazol-2-yl)cyclopropane-1-carboxylate: To a vial was added 7-(5-bromo-4-(isopropylamino)pyridin-2-yl)pyrrolo[1,2-b]pyridazine-3-carbonitrile (I-2) (60 mg, 0.168 mmol), (±) methyl trans-2-(1,3,4-thiadiazol-2-yl)cyclopropane-1-carboxylate (I-20) (47 mg, 0.253 mmol), Pd(OAc)2(9.5 mg, 0.042 mmol), Xantphos (49 mg, 0.084 mmol), copper(I) iodide (16 mg, 0.084 mmol), and cesium carbonate (165 mg, 0.5 mmol). 1,4-Dioxane (1 mL), was added, and the mixture was degassed with argon for 1 minute. The vial was sealed and stirred 2 hours at 105° C. Afterward, the vial was cooled and the crude material was diluted with DMF (0.5 mL). This mixture was filtered, and purified by RP-HPLC (eluent: water/MeCN*0.1% TFA) to yield the product as a trifluoroacetate salt. The purified material was dissolved in EtOAc and neutralized with sat. aq. NaHCO3. The layers were separated, and the organic layer was dried over MgSO4, filtered, and concentrated under reduced pressure.

±7-(5-(5-(trans-2-(2-hydroxypropan-2-yl)cyclopropyl)-1,3,4-thiadiazol-2-yl)-4-(isopropylamino)pyridin-2-yl)pyrrolo[1,2-b]pyridazine-3-carbonitrile (Example 132): To a solution of ±methyl trans-2-(5-(6-(3-cyanopyrrolo[1,2-b]pyridazin-7-yl)-4-(isopropylamino)pyridin-3-yl)-1,3,4-thiadiazol-2-yl)cyclopropane-1-carboxylate (40 mg, 0.087 mmol) in THF (2 mL) at −78° C. was added methyllithium (1.6M in ether, 0.16 mL, 0.26 mmol) dropwise. The reaction was quenched at −78° C. with dropwise addition of saturated aqueous ammonium chloride, then slowly allowed to warm to room temperature. The reaction was partitioned between water and EtOAc, and the aqueous layer was extracted two additional times with EtOAc. The combined organic layers were dried over magnesium sulfate, filtered, concentrated under reduced pressure, and then purified by RP-HPLC (eluent: water/MeCN*0.1% TFA) to yield the product Example 132 as a trifluoroacetate salt.

The following compounds were made according to the previous procedures, using the appropriate starting materials and protecting group chemistry as needed:

methyl trans-2-(5-(6-chloro-4-(oxetan-3-ylamino)pyridin-3-yl)-1,3,4-thiadiazol-2-yl)cyclopropane-1-carboxylate: To a vial was added 2-chloro-5-iodo-N-(oxetan-3-yl)pyridin-4-amine (synthesized in step 1 of I-34) (60 mg, 0.0.19 mmol), (±) methyl trans-2-(1,3,4-thiadiazol-2-yl)cyclopropane-1-carboxylate (1-20) (40 mg, 0.213 mmol), Pd(OAc)2(8.7 mg, 0.039 mmol), Xantphos (45 mg, 0.077 mmol), copper(I) iodide (15 mg, 0.077 mmol), and cesium carbonate (189 mg, 0.58 mmol). 1,4-Dioxane (1 mL), was added, and the mixture was degassed with argon for 1 minute. The vial was sealed and stirred 90 minutes at 105° C. Afterward, the vial was cooled and the crude material was diluted with DMF (0.5 mL). This mixture was filtered, and purified by RP-HPLC (eluent: water/MeCN*0.1% TFA) to yield the product as a trifluoroacetate salt. The purified material was dissolved in EtOAc and neutralized with sat. aq. NaHCO3. The layers were separated, and the organic layer was dried over MgSO4, filtered, and concentrated under reduced pressure.

2-(trans-2-(5-(6-chloro-4-(oxetan-3-ylamino)pyridin-3-yl)-1,3,4-thiadiazol-2-yl)cyclopropyl)propan-2-ol: To a solution of methyl trans-2-(5-(6-chloro-4-(oxetan-3-ylamino)pyridin-3-yl)-1,3,4-thiadiazol-2-yl)cyclopropane-1-carboxylate (80 mg, 0.218 mmol) in THF (10 mL) at −78° C. was added methyllithium (1.6M in ether, 0.2 mL, 0.33 mmol) dropwise, and the reaction was stirred 5 minutes at −78° C. The reaction was quenched at −78° C. with dropwise addition of 1 mL saturated aqueous ammonium chloride, then slowly allowed to warm to room temperature. The reaction was partitioned between water and EtOAc, and the aqueous layer was extracted two additional times with EtOAc. The combined organic layers were dried over magnesium sulfate, filtered, concentrated under reduced pressure, and then purified by silica gel chromatography (eluent: EtOAc/hexanes, then MeOH/EtOAc) to yield the product.

7-(5-(5-(trans-2-(2-hydroxypropan-2-yl)cyclopropyl)-1,3,4-thiadiazol-2-yl)-4-(oxetan-3-ylamino)pyridin-2-yl)pyrrolo[1,2-b]pyridazine-3-carbonitrile (±P-SEA6): A microwave vial was charged with 2-(trans-2-(5-(6-chloro-4-(oxetan-3-ylamino)pyridin-3-yl)-1,3,4-thiadiazol-2-yl)cyclopropyl)propan-2-ol (50 mg, 0.136 mmol), 7-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)pyrrolo[1,2-b]pyridazine-3-carbonitrile (I-8, 51.3 mg, 0.191 mmol), and [1,1′-Bis(diphenylphosphino)ferrocene]dichloropalladium(II) (20 mg, 0.027 mmol). To the flask was added 1,4-dioxane (1.0 mL) and a 2M aqueous solution of sodium carbonate (0.136 mL, 0.27 mmol). The reaction mixture was degassed by bubbling argon for 30 seconds. The vial was sealed and heated with stirring at 120° C. for 20 minutes in a microwave reactor. The cooled reaction mixture was concentrated and purified by RP-HPLC (eluent: water/MeCN*0.1% TFA) to yield the product Example 136 as a trifluoroacetate salt.

Single isomers were isolated by chiral supercritical fluid chromatography separation.

The following compounds were made according to the previous procedures, using the appropriate starting materials and protecting group chemistry as needed:

N-((1R,5S,8S)-3-(5-(4-(((R)-1-cyanoethyl)amino)-6-(3-cyanopyrrolo[1,2-b]pyridazin-7-yl)pyridin-3-yl)-1,3,4-thiadiazol-2-yl)-3-azabicyclo[3.2.1]octan-8-yl)cyclopropanecarboxamide (Example 148): To a vial containing (R)-(4-((1-cyanoethyl)amino)-6-(3-cyanopyrrolo[1,2-b]pyridazin-7-yl)pyridin-3-yl)boronic acid as a trifluoroacetate salt (25 mg, 0.056 mmol), N—((R,5S,8s)-3-(5-bromo-1,3,4-thiadiazol-2-yl)-3-azabicyclo[3.2.1]octan-8-yl)cyclopropanecarboxamide (20 mg, 0.056 mmol), Pd(dppf)C12(1.8 mg, 0.014 mmol), was added 1,4-dioxane (1 mL) and sodium carbonate (2M Aqueous, 0.085 mL, 0.17 mmol). The mixture was degassed with argon for 30 seconds, sealed, and heated for 1 hour at 120° C. Afterward, the vial was cooled and the crude material was diluted with DMF (0.5 mL). This mixture was filtered, and purified by RP-HPLC (eluent: water/MeCN*0.1% TFA) to yield the product Example 148 as a trifluoroacetate salt.

The following compounds were made according to the previous procedures, using the appropriate starting materials and protecting group chemistry as needed:

The following compounds were made according to the previous procedures, using the appropriate starting materials and protecting group chemistry as needed:

7-(5-(5-(4-hydroxy-4-methylcyclohexyl)-1,3,4-thiadiazol-2-yl)-4-(oxetan-3-ylamino)pyridin-2-yl)pyrrolo[1,2-b]pyridazine-3-carbonitrile (Example 206 from Isomer 1): A microwave vial was charged with 4-(5-(6-chloro-4-(oxetan-3-ylamino)pyridin-3-yl)-1,3,4-thiadiazol-2-yl)-1-methylcyclohexan-1-ol (I-34 Isomer 1) (21.1 mg, 0.055 mmol), 7-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)pyrrolo[1,2-b]pyridazine-3-carbonitrile (I-8, 20.9 mg, 0.078 mmol), and [,1′-Bis(diphenylphosphino)ferrocene]dichloropalladium(II) (8.2 mg, 0.011 mmol). To the flask was added DME (1.0 mL) and a 2M aqueous solution of sodium carbonate (0.06 mL, 0.11 mmol). The reaction mixture was degassed by bubbling argon for 30 seconds. The vial was sealed and heated with stirring at 120° C. for 20 minutes in a microwave reactor. The cooled reaction mixture was concentrated and purified by RP-HPLC (eluent: water/MeCN*0.1% TFA) to yield the product Example 206 from Isomer 1 as a trifluoroacetate salt.

The following compounds were made according to the previous procedures, using the appropriate starting materials and protecting group chemistry as needed:

tert-butyl (S)-((1-(5-(6-chloro-4-(isopropylamino)pyridin-3-yl)-1,3,4-thiadiazole-2-carbonyl)pyrrolidin-2-yl)methyl)carbamate: To a solution of methyl 5-(6-chloro-4-(isopropylamino)pyridin-3-yl)-1,3,4-thiadiazole-2-carboxylate (50.0 mg, 0.16 mmol) in MeOH (0.5 mL) was added tert-butyl (S)-(pyrrolidin-2-ylmethyl)carbamate (38.4 mg, 0.19 mmol). The reaction mixture was heated at 80° C. for 1 hour, then concentrated in vacuo and purified by silica gel column chromatography (eluent: MeOH/DCM) to provide tert-butyl (S)-((1-(5-(6-chloro-4-(isopropylamino)pyridin-3-yl)-1,3,4-thiadiazole-2-carbonyl)pyrrolidin-2-yl)methyl)carbamate.

(S)-7-(5-(5-(2-(aminomethyl)pyrrolidine-1-carbonyl)-1,3,4-thiadiazol-2-yl)-4-(isopropylamino)pyridin-2-yl)pyrrolo[1,2-b]pyridazine-3-carbonitrile (Example 229): To a solution of tert-butyl (S)-((1-(5-(6-(3-cyanopyrrolo[1,2-b]pyridazin-7-yl)-4-(isopropylamino)pyridin-3-yl)-1,3,4-thiadiazole-2-carbonyl)pyrrolidin-2-yl)methyl)carbamate (19 mg, 0.032 mmol) in dioxane (0.25 mL) was added hydrochloric acid (4N in dioxane, 121.2 μL, 0.48 mmol) and stirred for 3 hours. The reaction mixture was concentrated, filtered and purified by reverse phase high pressure liquid chromatography (eluent: water/MeCN*0.1% TFA) to provide the final compound Example 229.

The following compounds were made according to the previous procedures, using the appropriate starting materials and protecting group chemistry as needed:

7-(5-(5-((1r,3r)-3-aminocyclobutyl)-1,3,4-thiadiazol-2-yl)-4-(isopropylamino)pyridin-2-yl)pyrrolo[1,2-b]pyridazine-3-carbonitrile hydrochloride: tert-butyl((1r,3r)-3-(5-(6-(3-cyanopyrrolo[1,2-b]pyridazin-7-yl)-4-(isopropylamino)pyridin-3-yl)-1,3,4-thiadiazol-2-yl)cyclobutyl)carbamate (150 mg, 0.28 mmol) was then dissolved in HCl (4.0 M in dioxane, 2 mL, 8 mmol) and stirred at room temperature for 1 hour after which the reaction mixture was concentrated to dryness directly to give the desired product as an HCl salt which was used without further purification.

Example 282 was made according to Procedure 14, using the appropriate starting materials and protecting group chemistry as needed:

7-[4-(isopropylamino)-5-[5-[rac-(2R)-2-(aminomethyl)pyrrolidine-1-carbonyl]-1,3,4-thiadiazol-2-yl]-2-pyridyl]pyrrolo[1,2-b]pyridazine-3-carbonitrile (Example 283): To a solution of 7-(5-(5-((1r,3r)-3-aminocyclobutyl)-1,3,4-thiadiazol-2-yl)-4-(isopropylamino)pyridin-2-yl)pyrrolo[1,2-b]pyridazine-3-carbonitrile hydrochloride (30 mg, 0.064 mmol), oxetan-3-one (32.4 mg, 0.45 mmol) in 1 mL of DCE and 1 mL of acetic acid, sodium triacetoxy borohydride (95.3 mg, 0.45 mmol) was added to the suspension. It was stirred for overnight. Oxetan-3-one (32.4 mg, 0.45 mmol) and sodium triacetoxy borohydride (95.3 mg, 0.45 mmol) was added to the mixture. After 5 hours, diluted with EtOAc (20 mL) and washed with 10 mL of saturated NaHCO3. The organic layer was dried and concentrated. This mixture was filtered, and purified by RP-HPLC (eluent: water/MeCN*0.1% TFA) to yield the product Example 283 as a trifluoroacetate salt. ES/MS: 487.6 [M+H+].

The following compounds were made according to the previous procedures, using the appropriate starting materials and protecting group chemistry as needed:

Tert-butyl((1r,4r)-4-(5-(6-(3-cyanopyrrolo[1,2-b]pyridazin-7-yl)-4-(isopropylamino)pyridin-3-yl)-1,3,4-thiadiazol-2-yl)cyclohexyl)carbamate: To a slurry of tert-butyl((1r,4r)-4-(5-(6-chloro-4-(isopropylamino)pyridin-3-yl)-1,3,4-thiadiazol-2-yl)cyclohexyl)carbamate (prepared following the protocol in steps 1 and 2 of Procedure 14 using the appropriate carboxylic acid) (89 mg, 0.20 mmol) in DME (3 mL) was added 7-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)pyrrolo[1,2-b]pyridazine-3-carbonitrile (85 mg, 0.32 mmol), XPhos Pd G3 (12 mg, 0.15 mmol) and aqueous potassium phosphate, tribasic (2M, 0.2 mL, 0.4 mmol). The resulting mixture was degassed with argon for 2 min, sealed, and heated in a microwave at 120° C. for 20 min. The resulting slurry was diluted with Et2O, filtered, and washed with Et20. The resulting solid was purified by silica gel chromatography (eluent: MeOH/CH2Cl2) to give the desired product.

7-(5-(5-((1R,4R)-4-aminocyclohexyl)-1,3,4-thiadiazol-2-yl)-4-(isopropylamino)pyridin-2-yl)pyrrolo[1,2-b]pyridazine-3-carbonitrile: To a solution of tert-butyl((1r,4r)-4-(5-(6-(3-cyanopyrrolo[1,2-b]pyridazin-7-yl)-4-(isopropylamino)pyridin-3-yl)-1,3,4-thiadiazol-2-yl)cyclohexyl)carbamate (75 mg, 0.13 mmol) in CH2Cl2(2.5 mL) was added solution of hydrochloric acid in dioxane (4M, 0.5 mL, 2 mmol). The resulting slurry was stirred at 50° C. for 1 hour. Additional hydrochloric acid in dioxane (4M, 0.5 mL, 2 mmol) and methanol (0.5 mL) was added and the solution was stirred at 50° C. for 2 hours, followed by stirring at room temperature for 18 hours. The reaction mixture was concentrated and the crude product was used directly in the next step.

N-((1r,4r)-4-(5-(6-(3-cyanopyrrolo[1,2-b]pyridazin-7-yl)-4-(isopropylamino)pyridin-3-yl)-1,3,4-thiadiazol-2-yl)cyclohexyl)-3-methyloxetane-3-carboxamide (Example 314): To a slurry of 7-(5-(5-((1R,4R)-4-aminocyclohexyl)-1,3,4-thiadiazol-2-yl)-4-(isopropylamino)pyridin-2-yl)pyrrolo[1,2-b]pyridazine-3-carbonitrile in DMF (0.8 mL) and MeOH (0.5 mL) was added N,N-diisopropylethylamine (0.06 mL, 0.34 mmol), 3-[2-[2-[2-(tert-butoxycarbonylamino)ethoxy]ethoxy]ethoxy]propanoic acid (18 mg, 0.056 mmol), and HATU (20 mg, 0.053 mmol). The solution was stirred at room temperature for 16 hours and concentrated. The crude material was dissolved in DMF and purified by RP-HPLC (eluent: water/MeCN*0.1% TFA) to yield the product Example 314 as a trifluoroacetate salt.

Example 315 was made according to Procedure 34, using the appropriate starting materials and protecting group chemistry as needed:

(R)-2-((2-chloro-5-iodopyridin-4-yl)amino)propanenitrile: To a solution of (R)-2-((2-chloro-5-iodopyridin-4-yl)amino)propanamide (10.1 g, 30 mmol) in methyl-THF (165 mL) was added trimethylamine (21 mL, 154 mmol). To the slurry was slowly added trifluoroacetic anhydride (1.15 mL, 8.27 mmol) in methyl-THF (10 mL) over 15 min. The resulting solution is stirred at room temperature for 45 min before dilution with H2O and washing with aqueous NH4Cl. The aqueous layers are back-extracted with ethyl acetate and the combine organic layers are dried over MgSO4and concentrated to dryness. The crude material was dissolved in hot ethyl acetate, to which hexanes was added until precipitation initiated. The solution was allowed to cool to room temperature for 18 hours and the resulting precipitate was filtered and washed with hexanes to provide the desired product.

4-(5-(6-chloro-4-(isopropylamino)pyridin-3-yl)-1,3,4-thiadiazol-2-yl)-2-(methyl-d3)butan-1,1,1-d3-2-ol: To a flask with methyl 3-(5-(6-chloro-4-(isopropylamino)pyridin-3-yl)-1,3,4-thiadiazol-2-yl)propanoate (100 mg, 0.29 mmol) under N2was added THF (2 mL), and the reaction was cooled to 0° C. To the reaction was added Methyl-d3-lithium, as complex with lithium iodide solution (117 mg, 0.73 mmol) dropwise, and the reaction was stirred for 30 min at 0° C. Afterward, the reaction was quenched by dropwise addition of saturated aqueous NH4Cl, and the mixture was diluted with EtOAc and water. The layers were separated, and the aqueous layer was washed once with EtOAc. The combined organic layers were dried over MgSO4, filtered and concentrated. The resulting crude residue was purified by silica gel chromatography (eluent: EtOAc/hexanes) to give the desired product.

7-(5-(5-(3-hydroxy-3-(methyl-d3)butyl-4,4,4-d3)-1,3,4-thiadiazol-2-yl)-4-(isopropylamino)pyridin-2-yl)pyrrolo[1,2-b]pyridazine-3-carbonitrile (Example 317): Coupling performed as in Procedure 9 to yield the product Example 317 as a trifluoroacetate salt.

Example 318 was made according to Procedure 9, using the appropriate starting materials and protecting group chemistry as needed:

2-(5-(6-(3-cyanopyrrolo[1,2-b]pyridazin-7-yl)-4-(isopropylamino)pyridin-3-yl)-1,3,4-thiadiazol-2-yl)acetic acid (Example 319): To ethyl 2-(5-(6-(3-cyanopyrrolo[1,2-b]pyridazin-7-yl)-4-(isopropylamino)pyridin-3-yl)-1,3,4-thiadiazol-2-yl)acetate (10 mg, 0.022 mmol) in THF (0.5 mL) was added lithium hydroxide (3 mg, 0.125 mmol). The reaction mixture was stirred at RT for 6 hours then directly purified by RP-HPLC (eluent: water/MeCN*0.1% TFA) to yield the product Example 319 as a trifluoroacetate salt.

1-(5-(6-chloro-4-(isopropylamino)pyridin-3-yl)-1,3,4-thiadiazol-2-yl)-2-methylpropan-2-ol Ethyl (60 mg, 0.22 mmol) in THF (1 mL) was added lithium diisopropylamide (2.0 M THF, 0.56 mmol). The temperature was maintained in the range of −70° C. to −78° C. throughout the 15 minute addition. Following the addition, the resulting slurry was stirred for 2 hours at ambient temperature then directly purified by RP-HPLC (eluent: water/MeCN*0.1% TFA) to yield the product 7-(5-(5-(2-hydroxy-2-methylpropyl)-1,3,4-thiadiazol-2-yl)-4-(isopropylamino)pyridin-2-yl)pyrrolo[1,2-b]pyridazine-3-carbonitrile (Example 320): Coupling performed as in Procedure 9 to yield the product Example 320 as a trifluoroacetate salt.

The following compounds were made according to the previous procedures, using the appropriate starting materials and protecting group chemistry as needed:

methyl 6-chloro-4-((tetrahydro-2H-pyran-4-yl)amino)nicotinate: To a solution of methyl 4,6-dichloropyridine-3-carboxylate (4.00 g, 19.4 mmol) and tetrahydropyran-4-amine hydrochloride (4.01 g, 29.1 mmol) in THF (20 mL) was added N,N-diisopropylethylamine (10.1 mL, 58.2 mmol). The reaction mixture was heated to 120° C. overnight. The reaction was cooled to rt then partitioned between water and EtOAc, and the aqueous layer was extracted two additional times with EtOAc. The combined organic layers were washed with brine and dried over magnesium sulfate, isolated by vacuum filtration, concentrated in vacuo, and purified by silica gel column chromatography (eluent: EtOAc/Hexanes) to provide the desired product.

6-chloro-4-((tetrahydro-2H-pyran-4-yl)amino)nicotinohydrazide: To a solution of methyl 6-chloro-4-((tetrahydro-2H-pyran-4-yl)amino)nicotinate (3.03 g, 11.2 mmol) in EtOH (18 mL) was added hydrazine hydrate (4.42 mL, 90.9 mmol). The solution was refluxed at 80° C. for 3 hours. The solution was cooled, concentrated and carried forward without further purification.

tert-butyl(4-(2-(6-chloro-4-((tetrahydro-2H-pyran-4-yl)amino)nicotinoyl)hydrazine-1-carbonyl)bicyclo[2.2.2]octan-1-yl)carbamate: To a solution of crude 6-chloro-4-((tetrahydro-2H-pyran-4-yl)amino)nicotinohydrazide (2.7 g, 9.97 mmol) and 4-(tert-butoxycarbonylamino)bicyclo[2.2.2]octane-1-carboxylic acid (2.8 g, 10.5 mmol) in DMF (50 mL) was added N,N-diisopropylethylamine (5.70 mL, 31.9 mmol) followed by a solution of HATU (4.55 g, 12.0 mmol). The reaction mixture was stirred at RT for 15 minutes, then concentrated in vacuo, and purified by silica gel column chromatography (eluent: MeOH/DCM).

tert-butyl(4-(5-(6-chloro-4-((tetrahydro-2H-pyran-4-yl)amino)pyridin-3-yl)-1,3,4-thiadiazol-2-yl)bicyclo[2.2.2]octan-1-yl)carbamate: To a solution of tert-butyl(4-(2-(6-chloro-4-((tetrahydro-2H-pyran-4-yl)amino)nicotinoyl)hydrazine-1-carbonyl)bicyclo[2.2.2]octan-1-yl)carbamate (5.00 g, 9.58 mmol) in 2-methyltetrahydrofuran (47.9 mL), Lawesson's Reagent (4.26 g, 10.5 mmol) was added, and the resulting reaction mixture heated to 50° C. overnight. Upon completion, the reaction mixture concentrated and purified by silica gel chromatography (eluent: EtOAc/hexanes). Product containing fractions were combined and stirred with 5 g of 10% Palladium on carbon and filtered. The solution was concentrated and purified by silica gel chromatography (eluent: MeOH/DCM).

tert-butyl(4-(5-(6-(3-cyanopyrrolo[1,2-b]pyridazin-7-yl)-4-((tetrahydro-2H-pyran-4-yl)amino)pyridin-3-yl)-1,3,4-thiadiazol-2-yl)bicyclo[2.2.2]octan-1-yl)carbamate: A microwave vial was charged with tert-butyl(4-(5-(6-chloro-4-((tetrahydro-2H-pyran-4-yl)amino)pyridin-3-yl)-1,3,4-thiadiazol-2-yl)bicyclo[2.2.2]octan-1-yl)carbamate (86.0 mg, 0.136 mmol), 2,7-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)pyrrolo[1,2-b]pyridazine-3-carbonitrile (I-8, 58.7 mg, 0.218 mmol), XPhos Pd G3 (8.63 mg, 0.010 mmol), 2M aqueous potassium phosphate tribasic (0.138 mL, 0.276 mmol). DMF (2.0 mL) was added, and the reaction mixture was degassed by bubbling argon for 60 seconds. The vial was sealed and heated with stirring at 120° C. for 30 minutes in a microwave reactor. The cooled reaction mixture was diluted with THF and filtered through a syringe filer, concentrated in vacuo, and purified by RP-HPLC (eluent: water/MeCN*0.1% TFA) to yield the product as a trifluoroacetate salt.

7-(5-(5-(4-aminobicyclo[2.2.2]octan-1-yl)-1,3,4-thiadiazol-2-yl)-4-((tetrahydro-2H-pyran-4-yl)amino)pyridin-2-yl)pyrrolo[1,2-b]pyridazine-3-carbonitrile (Example 336): To a solution of tert-butyl(4-(5-(6-(3-cyanopyrrolo[1,2-b]pyridazin-7-yl)-4-((tetrahydro-2H-pyran-4-yl)amino)pyridin-3-yl)-1,3,4-thiadiazol-2-yl)bicyclo[2.2.2]octan-1-yl)carbamate (24.8 mg, 0.039 mmol) in 0.04 mL 1,2-dichloroethane was added 4M HCl in dioxane (0.0964 mL, 0.039 mmol). The solution was stirred for 1 h at rt during which time a precipitate formed. The reaction mixture was concentrated in vacuo, and purified by RP-HPLC (eluent: water/MeCN*0.1% TFA) to yield the product Example 336 as a trifluoroacetate salt.

N-(4-(5-(6-(3-cyanopyrrolo[1,2-b]pyridazin-7-yl)-4-((tetrahydro-2H-pyran-4-yl)amino)pyridin-3-yl)-1,3,4-thiadiazol-2-yl)bicyclo[2.2.2]octan-1-yl)-2-hydroxy-2-methylpropanamide (Example 337): To a solution of crude 7-(5-(5-(4-aminobicyclo[2.2.2]octan-1-yl)-1,3,4-thiadiazol-2-yl)-4-((tetrahydro-2H-pyran-4-yl)amino)pyridin-2-yl)pyrrolo[1,2-b]pyridazine-3-carbonitrile hydrochloride (12.0 mg, 0.018 mmol) and 2-hydroxy-2-methyl-propanoic acid (23.7 mg, 0.227 mmol) in DMF (0.5 mL) was added N,N-diisopropylethylamine (0.011 mL, 0.059 mmol) followed by the addition of a solution of HATU (7.48 mg, 0.019 mmol) in DMF (0.1 mL). The reaction mixture was stirred at RT overnight then directly purified by RP-HPLC (eluent: water/MeCN*0.1% TFA) to yield the product Example 337 as a trifluoroacetate salt.

Single isomers were isolated by chiral supercritical fluid chromatography separation.

The following compounds were made according to the previous procedures, using the appropriate starting materials and protecting group chemistry as needed:

Proton NMR data for each compound exemplified is shown in Table 1.

Compounds comprising the following components or combined with other components exemplified herein might be prepared according to the Examples and Procedures described herein using the appropriate starting materials and protecting group chemistry as needed.

R1and Het pairings to form the below components present in these compounds would be generated from intermediates I-2 and I-3. These would be paired with a thiadiazole-R2component.

The above components would be combined with intermediates and components attached to the below R2groups and/or to exemplified R2groups by the methods described herein.

Also, any of the exemplified compounds may be made, using, as components, the following “Het” groups:

Biological Assays

Biological assays were conducted to measure activity against TNFα and IRAK4. As summarized in Table 2, the test compounds are inhibitors of IRAK4.

Cryopreserved human monocytes (Stem Cell Technologies) were thawed, diluted in RPMI with GlutaMAX™ (Gibco® 200 mM L-alanyl-L-glutamine) (10 mM HEPES, 1× Pen-Strep, 55 μM ß-mercaptoethanol, 1 mM Sodium pyruvate) media containing 10% FBS to 0.125×106cells/ml and recovered at 37° C. for 2 hours. The cell suspension was then plated at a density of 5,000 cells/well onto black 384 well Greiner clear bottom plates. Plates were pre-spotted with test compounds and serially diluted in DMSO where 40 nL/well were delivered using the Echo 550 acoustic liquid dispenser (Labcyte®) for a final DMSO concentration of 0.1%. Plated cells were treated with compound for 1 hour at 37° C. Cells were then stimulated with 50 pg/ml of LPS (Sigma) excluding outside columns of plate used for unstimulated cell control wells. Cells were incubated for an additional 4 hours at 37° C. Cells were then spun out of the media and 5 μl of sample were taken and analyzed for total TNFα content using the TR-FRET Human TNFα detection system (CisBio). This system utilizes two labeled antibodies (cryptate and XL665) that bind to two different epitopes of the TNFα molecule and produce FRET signal proportional to the concentration of TNFα in the sample. Detection antibodies are mixed 50:50 and 5 μL were dispensed into each well. Plates were covered with clear seals and incubated at room temp overnight. The following morning plates were read using an Envision 2103 Multilabeled reader (PerkinElmer) with excitation/emission/FRET emission at 340 nm/615 nm/665 nm, respectively. Fluorescence intensities at 615 nm and 665 nm emission wavelengths were expressed as a ratio (665 nm/615 nm). Percent of control was calculated as follows:
% Control=100×(Ratiosample−Ratio0% stimulation)/(Ratio100% Stimulation−Ratio0% Stimulation)
where unstimulated cells (0% stimulation) were the negative control and stimulated cells (100% stimulation) were used as the positive control.
IRAK4 Biochemical Assay Procedure:

IRAK4 enzyme (Carna Biosciences, Chuo-ku, Kobe, Japan) activity was measured by detecting phosphorylated peptide substrate formation using an antibody against the phosphorylated peptide substrate. This is a time-resolved fluorescence resonance energy transfer (TR-FRET) immunoassay, based on the STK1 KinEASE Assay (Cisbio, Bedford, Mass.). The assay was designed as a simple two-step, endpoint assay (a 5 μl enzyme reaction followed by 5 μl stop and detect Solution) performed in ProxiPlate-384 Plus plates (Perkin Elmer, Waltham, Mass.). Staurosporine, a non-selective kinase inhibitor was used as a positive control. Compounds diluted in DMSO were spotted into 384 well plates using a Labcyte® Echo 550 Liquid Handling System prior to addition of IRAK4 enzyme and peptide substrate. Reaction solutions were delivered using a Multi-Flo (Bio-Tek Instruments). The enzyme and peptide solution was incubated with compound for 15 minutes at room temp before the reaction was initiated by the addition of ATP. The standard 5 μl reaction mixture contained 500 μM ATP, 2 M peptide (STK1 Peptide), 0.75 nM of IRAK4 in reaction buffer (50 mM HEPES, pH 7.0, 0.02% NaN3, 0.01% BSA, 0.1 mM Orthovanadate, 5 mM MgC2, 0.025% NP-40, 1 mM DTT). After 120 min of incubation at room temperature, 5 μl of Stop and Detect Solution (1:100 Cryptate labeled anti-phosphorylated peptide antibody solution and 125 nM Tracer in a 50 mM HEPES pH 7.0 detection buffer containing sufficient EDTA) was added. The plate was then further incubated for 60 minutes at room temperature and read on Envision 2103 Multilabeled reader (PerkinElmer) with excitation/emission/FRET emission at 340 nm/615 nm665 nm, respectively. Fluorescence intensities at 615 nm and 665 nm emission wavelengths were expressed as aratio (665 nm/615 nm). Percentage of inhibition was calculated as below:
% Inhibition=100×(Ratiosample−Ratio0% Inhibition)/(Ratio100% Inhibition−Ratio0% Inhibition)

The 0% inhibition value comes from control wells lacking inhibitor. The 100% inhibition value comes from control wells containing a saturating amount of known inhibitor staurosporine.