Compounds for mastl inhibition

Abstract

The present disclosure provides compounds of Formula (I). The compounds of Formula (I) may be used as MASTL inhibitors. Also provided are pharmaceutical compositions comprising the compounds of Formula (I) and compounds of Formula (I) for use in a method of treatment.

Description

[0001] 008863987

[0002] 1

[0003] Compounds forMASTL Inhibition

[0004] This application claims priority from GB 2500231.2 filed 09 January 2025, the contents and elements of which are herein incorporated by reference for all purposes.

[0005] Field of the Invention

[0006] The present invention relates to compounds which act as MASTL inhibitors and use of the compounds in a method of treatment.

[0007] Background

[0008] The present invention relates to novel compounds which can be used as microtubule-associated serine / threonine kinase-like (MASTL) inhibitors. The invention also relates to compositions containing the compounds and uses of the compounds for the treatment of MASTL related diseases such as cancer.

[0009] MASTL, also known as Greatwall kinase, is a key mitotic kinase that regulates mitotic progression and maintains mitotic integrity. MASTL plays a unique role in the control of cell cycle progression through the regulation of specific Protein Phosphatase 2A (PP2A) complexes critical for execution of mitosis via control of the mitotic CDK phosphor-proteome during entry and exit from mitosis.

[0010] Notably, MASTL is highly expressed in multiple types of cancers, including breast, head and neck, gastric, thyroid and colorectal cancers. MASTL targeting has reduced tumour growth in vitro and in vivo tumour models.

[0011] Recent studies have suggested that MASTL acts as an oncogenic kinase by inactivating tumour suppressive PP2A-B55 complex in breast cancer cells, thereby regulating various oncogenic properties, such as cellular transformation, chromosome instability, and metastasis.

[0012] MASTL is a promising target for selective anticancer treatment. Good biological properties such as metabolic stability are desirable for an inhibitor intended for medical use.

[0013] WO 2024 / 003773 A1 describes several 2,7-naphthyridine compounds which show activity as MASTL inhibitors.

[0014] The development of alternative MASTL inhibitors with anti-tumour efficacy is needed, particularly with significant anti-tumour activity.

[0015] Additionally, there have been a limited number of MASTL inhibitors reported with significant anti-tumour activity in combination with good biological properties such as metabolic stability. MASTL inhibitors with favourable biological properties, such as good metabolic stability, in combination with anti-tumour efficacy are particularly desirable.

[0016] The present invention has been devised in light of the above considerations.008863987

[0017] Summary of the Invention

[0018] In a general aspect, the invention provides novel compounds which are MASTL inhibitors. In a first aspect of the invention, there is provided a compound of Formula (I) and pharmaceutically acceptable salts, solvates, protected forms and prodrug forms thereof.

[0019] The compound of Formula (I) refer to compounds having the following structure:

[0020]

[0021] Formula (I)

[0022] wherein:

[0023] R1is an optionally substituted heteroaryl;

[0024] X, Y and Z are independently selected from N, CH, and CR5wherein at least one of X, Y or Z is N and wherein R5is selected from Ci-Ce alkyl and Ci-Ce alkoxy;

[0025] V and Q are selected from N, CH, and CR5, wherein at least one of V or Q is N and wherein R5is selected from Ci-Ce alkyl and Ci-Ce alkoxy;

[0026] and wherein:

[0027] R2is hydrogen or C1-C4 alkyl; R3is fluoro-substituted C3-C10 cycloalkyl, or R3together with the carbon atom to which it is attached and a carbon atom of the pyridine ring forms a fluoro-substituted 5- to 8-membered ring that is fused to the pyridine ring; or R2and R3together with the carbon atom to which they are attached form a fluoro- substituted C3-C10 cycloalkyl group; and R4is hydrogen; or

[0028] or

[0029] R2is hydrogen or C1-C4 alkyl; and R3and R4together with the carbon and nitrogen atoms to which they are attached form a fluoro-substituted 5- to 10-membered heterocycloalkyl; or

[0030] or

[0031] R2is hydrogen or C1-C4 alkyl; R3is unsubstituted C1-C4 alkyl; and R4together with the nitrogen atom to which it is attached and a carbon atom of the pyridine ring forms a fluoro-substituted 5- to 8-membered ring that is fused to the pyridine ring.008863987

[0032] 3

[0033] In the compounds of Formula (1) the groups R2, R3, and R4must together include at least one fluoro-substituted ring system.

[0034] The compounds of the invention differ from known MASTL inhibitors such as those of

[0035] WO 2024 / 003773 in that they comprise a fluorinated cycloalkyl or heterocycloalkyl group. The compounds of the invention have been shown to have good MASTL inhibition in combination with unexpectedly good metabolic stability. In particular, the compounds of the invention have improved metabolic stability compared to the compounds of WO 2024 / 003773 while maintaining MASTL inhibition activity.

[0036] The compounds of the invention are also predicted to have improved MASTL binding affinity compared to the compounds of WO 2024 / 003773 while maintaining good selectivity for MASTL. In a further aspect of the invention, there is provided a pharmaceutical composition comprising a compound of formula (I) and a pharmaceutically acceptable excipient.

[0037] In a further aspect, there is provided a compound of Formula (I), or a pharmaceutical composition comprising a compound of Formula (I), for use in a method of treatment or prophylaxis.

[0038] The present invention also provides a method of treatment, the method comprising the step of administering a compound of formula (I), or a pharmaceutical composition comprising a compound of formula (I), to a subject in need thereof.

[0039] The method may involve the inhibition of MASTL and the method may be for the treatment of cancer.

[0040] The invention includes the combination of the aspects and preferred features described except where such a combination is clearly impermissible or expressly avoided.

[0041] Detailed Description of the Invention

[0042] Aspects and embodiments of the present invention will now be discussed with reference to the accompanying figures. Further aspects and embodiments will be apparent to those skilled in the art. All documents mentioned in this text are incorporated herein by reference.

[0043] Compound of Formula (I)

[0044] The present invention relates to compounds of Formula (I)008863987

[0045]

[0046] Formula (I)

[0047] and pharmaceutically acceptable salts, solvates, protected forms and prodrug forms thereof. In some embodiments the compound of Formula (I) is provided as a pharmaceutically acceptable salt or solvate.

[0048] In some embodiments, the compound of Formula (I) is provided in free base form. That is, the amines are in their neutral form and not in the form of a salt.

[0049] R2, R3and R4

[0050] The groups R2, R3, and R4must together include at least one fluorine atom attached to a cyclic ring system. Preferably groups R2, R3, and R4together include one or two fluoro substituents. Where there are two fluoro substituents, these are preferably attached to the same carbon atom.

[0051] R2may be hydrogen or C1-C4 alkyl or together with R3and the carbon atom to which they are attached form a fluoro-substituted C3-C10 cycloalkyl group. In some embodiments, R2is hydrogen or C1-C4 alkyl, such as methyl or ethyl. Preferably, R2is hydrogen or methyl. Most preferably, R2is hydrogen.

[0052] R3may be fluoro-substituted C3-C10 cycloalkyl; or R3together with the carbon atom to which it is attached and a carbon atom of the pyridine ring forms a fluoro-substituted 5- to 8-membered ring that is fused to the pyridine ring; or together with R2and the carbon atom to which they are attached form a fluoro-substituted C3-C10 cycloalkyl group; or together with R4and the carbon and nitrogen atoms to which they are attached form a fluoro-substituted 5- to 10-membered heterocycloalkyl. Preferably, R3may be fluoro-substituted C3-C10 cycloalkyl.

[0053] R4may be hydrogen; or together with R3and the carbon and nitrogen atoms to which they are attached form a fluoro-substituted 5- to 10-membered heterocycloalkyl; or R4together with the nitrogen atom to which it is attached and a carbon atom of the pyridine ring forms a fluoro-substituted 5- to 8-membered ring that is fused to the pyridine ring. Preferably, R4is hydrogen. In some embodiments, R2and R3together with the carbon atom to which they are attached form a fluoro-substituted C3-C10 cycloalkyl group, such as C3-C8 cycloalkyl or C3-C6 cycloalkyl. In these embodiments, R4is hydrogen. The cycloalkyl group formed by R2and R3may include008863987

[0054] 5

[0055] one or more ring structures, including fused and bridged bicyclic structures. In such embodiments, preferably, R2and R3together form a monocyclic C3-C6 cycloalkyl group, such as cyclopropyl, cyclobutyl, cyclopentyl or cyclohexyl. Preferably, the cycloalkyl has one or two fluoro substituents.

[0056] In some embodiments, R3together with the carbon atom to which it is attached and a carbon atom of the pyridine ring forms a fluoro-substituted 5- to 8-membered ring that is fused to the pyridine ring. In these embodiments, R2is hydrogen or C1-C4 alkyl and R4is hydrogen. In these embodiments, R3connects to a carbon atom in the 3- or 5-position of the neighbouring pyridine ring to form a second ring structure. The second ring is preferably a 5- or 6-membered and is preferably non-conjugated. The second ring structure may have one or two fluoro substituents. Preferably, the second ring structure has two fluoro substituents. In some such embodiments, R2may be C1-C4 alkyl, such as methyl or ethyl, such as ethyl.

[0057] In some embodiments R3and R4together with the carbon and nitrogen atoms to which they are attached form a fluoro-substituted 3- to 10-membered heterocycloalkyl. In these embodiments R2is hydrogen or C1-C4 alkyl. The ring system formed by R3and R4is non-conjugated, and may include one or more ring structures, including fused and bridged structures. Preferably, R3and R4together form a 5- or 6-membered monocyclic ring. In these embodiments, R2may be C1-C4 alkyl, such as methyl or ethyl.

[0058] In some embodiments, R4together with the nitrogen atom to which it is attached and a carbon atom of the pyridine ring forms a fluoro-substituted 5- to 8-membered ring that is fused to the pyridine ring. In these embodiments, R2is hydrogen or C1-C4 alkyl and R3is C1-C4 alkyl. In these embodiments, R4connects to a carbon atom in the 3- or 5-position of the neighbouring pyridine ring to form a second ring structure. The second ring is preferably a 5- or 6-membered ring, and is preferably non-conjugated. The second ring structure may have one or two fluoro substituents. Preferably, the second ring structure has two fluoro substituents. In these embodiments, R2and R3may each independently be C1-C4 alkyl, such as methyl or ethyl, such as ethyl.

[0059] In preferred embodiments, R3is fluoro-substituted C3-C10 cycloalkyl, such as C3-C8 or C3-C6 cycloalkyl. In these embodiments, R2is hydrogen or C1-C4 alkyl; and R4is hydrogen.

[0060] Preferably, R2is hydrogen or methyl. Most preferably, R2is hydrogen.

[0061] In these embodiments, the fluoro-substituted C3-C10 cycloalkyl may include one or more ring structures, and may include bridged structures and fused ring structures. Preferably, the cycloalkyl group is monocyclic or bicyclic, such as bicyclic. Most preferably, the cycloalkyl group is a Cs-Ce bicyclic group. In these embodiments, R3may be selected from the group consisting of:

[0062]

[0063] and008863987

[0064] Preferably, R3is:

[0065]

[0066] R1is an optionally substituted heteroaryl, such as C5-10 heteroaryl. Preferably, R1is attached to the rest of the compound via a carbon atom. The optional substituents on the heteroaryl group may be selected from C1-C4 alkyl or halo. Preferably the optional substituents are selected from methyl or fluoro.

[0067] In some embodiments, R1comprises an optionally substituted azole, such as pyrazole, imidazole, triazole or tetrazole. Preferably, R1comprises an imidazole or 1,2,3-triazole ring. The azole ring may be substituted on a nitrogen or a carbon atom. Preferably, the azole ring is substituted with one or two C1-C4 alkyl or fluoroalkyl groups.

[0068] In some embodiments, R1comprises a 1,2,3-triazole substituted with C1-C4 alkyl, such as methyl. Preferably, the substitution is on a nitrogen atom.

[0069] In some embodiments, R1comprises imidazole fused to a second ring. Preferably, the second ring is 5- to 6-membered and is unconjugated. The second ring may have one or more fluorosubstituents, such as one or two fluoro-substituents. Where the second ring has two fluorosubstituents, these are preferably on the same carbon atom.

[0070] R1may be selected from the group consisting of:

[0071]

[0072] Preferably R1is selected from the group consisting of:

[0073]

[0074] 008863987

[0075] More preferably, R1is:

[0076]

[0077] X, Y and Z

[0078] X, Y and Z are independently selected from N, CH, and CR5. At least one of X, Y and Z is nitrogen and R5is selected from Ci-Ce alkyl and Ci-Ce alkoxy. Preferably, X is nitrogen.

[0079] In some preferred embodiments, X is nitrogen and Y and Z are both CH.

[0080] In some embodiments, X, is nitrogen and one of either Y or Z is also nitrogen.

[0081] In some embodiments, X is nitrogen, Y is nitrogen and Z is CH.

[0082] V and Q

[0083] V and Q are selected from N, CH, and CR5. At least one of V or Q is nitrogen and R5is selected from Ci-Ce alkyl and Ci-Ce alkoxy. Preferably, Q is nitrogen.

[0084] In some preferred embodiments, Q is nitrogen and V is CH or CR5. Preferably, V is CH.

[0085] In some embodiments, Q is nitrogen and V is also nitrogen. In these embodiments, Y and Z are preferably CH.

[0086] In some preferred embodiments, X is nitrogen, Y and Z are both CH, Q is nitrogen and V is CH.

[0087] R5

[0088] When present, R5is selected from Ci-Ce alkyl and Ci-Ce alkoxy. In some embodiments, R5is Ci-Ce alkoxy, such as C1-C4 alkoxy, such as C1-C3 alkoxy. In some embodiments, R5is Ci-Ce alkyl, such as C1-C4 alkyl, such as C1-C3 alkyl. Preferably, R5is methyl or ethyl. More preferably, R5is methyl.

[0089] Embodiments

[0090] In preferred embodiments, R2is hydrogen or methyl, R4is hydrogen, R3is selected from the group consisting of:008863987

[0091] 8

[0092] >

[0093]

[0094] X is nitrogen, Y and Z are both CH, Q is nitrogen and V is CH.

[0095] The compound of the invention may be selected from any of the following compounds, including all isomeric forms (e.g. enantiomeric forms) and mixtures (e.g. racemic mixtures) thereof:

[0096]

[0097] 008863987

[0098] 9

[0099] >

[0100] >

[0101] ""

[0102]

[0103] 008863987

[0104]

[0105] 008863987

[0106] 11

[0107] <

[0108] <

[0109] <

[0110] < < "

[0111]

[0112] 008863987

[0113]

[0114] 008863987

[0115] 13

[0116] "

[0117] <

[0118] <

[0119] "

[0120] >

[0121]

[0122] 008863987

[0123] 14

[0124] ><

[0125] "

[0126] <<

[0127] "

[0128] >

[0129] "

[0130]

[0131] 008863987

[0132] 15

[0133] <

[0134]

[0135] and salts, solvates, protected forms and prodrug forms thereof.

[0136] <

[0137] Preferably, the co mpound of the present invention is:

[0138] F

[0139]

[0140] or

[0141] or a salt, solvate, protected form or prodrug form thereof. The compound may be in the form of a racemic mixture or may be an optically enriched (i.e. enantiomerically enriched) form.

[0142] Definitions

[0143] As used herein, “compound of the invention” may refer to any compound of Formula (I) as defined in the present invention.

[0144] As used herein, “fluoro-substituted” refers to a group in which one or more hydrogen atoms are replaced with a fluorine atom. There may be one, two or three fluoro substituents on one atom.008863987

[0145] 16

[0146] “alkyl” refers to a saturated linear or branched monovalent carbon chain. Where the prefix C1-6 is used this refers to the number carbon atoms in the group including those in branched off the main chain. Example include but are not limited to methyl, ethyl, n-propyl, iso-propyl, sec-butyl. “alkoxy” refers to a group -O-alkyl wherein the oxygen is monovalent and joined to the main structure at the relevant position. Where the prefix C1-6 is used this refers to the number carbon atoms in the alkyl group including those in branched off the main chain. Examples include but are not limited to methoxy, ethoxy, n-propoxy, iso-propoxy, sec-butoxy.

[0147] “Cycloalkyl” refers to a non-aromatic (i.e. saturated or partially saturated) hydrocarbon ring system that has the specified number of carbon atoms. This includes monocyclic, bicyclic or polycyclic ring systems. The bicyclic or polycyclic ring systems can be bridged or fused bicyclic systems.

[0148] “Heteroaryl” refers to an aromatic heterocyclic group. This includes monocyclic, bicyclic and polycyclic ring systems containing at least one non-carbon atom, such as nitrogen wherein at least the ring containing the at least one non-carbon atom is aromatic. Examples include, but are not limited to, azoles, such as imidazole and triazole, pyridine and indole. A heteroaryl group may be substituted or unsubstituted. Where the prefix C5-10 is used this refers to the number of ring atoms of any type including any heteroatoms. For example, in the case of pyridine there are 5 carbon and 1 nitrogen atoms in the “ring system” and this would be considered a “Ce-heteroaryl” group as there are 6 atoms in total in the ring system. In the case of fused heteroaryl groups the numbering refers to all ring atoms in all rings including any rings that are not aromatic themselves.

[0149] “Heterocycloalkyl” to a non-aromatic (i.e. saturated or partially saturated) ring system containing the specified number of ring atoms and containing at least one heteroatom, such as N, as a ring member. Heterocycloalkyl rings include monocyclic, bicyclic or polycyclic ring systems. The bicyclic or polycyclic ring systems can be bridged or fused bicyclic systems.

[0150] Heterocycloalkyl rings may be substituted or unsubstituted. Substitution may occur on a carbon atom or on a nitrogen.

[0151] “Azole” refers to any aromatic 5-membered ring containing a nitrogen atom and at least one other non-carbon atom. Azoles include, but are not limited to, imidazole, pyrazole, 1 ,2,3-triazole, 1,2,4-triazole and tetrazole. Azoles may be substituted or unsubstituted. Substitution may occur on a carbon or a nitrogen atom.

[0152] As used herein, “wedged bonds” in chemical structures (either dashed or complete) are intended to indicate absolute configuration of the respective groups around a potential chiral centre.

[0153] As used herein, “line bonds” are intended to indicate connectivity of atoms but not to define any particular arrangement of groups e.g. around a chiral centre. In the case of a single chiral centre, a structure provided with line bonds only encompasses racemic mixtures as well as enantiomerically enriched material of either the (R)- or (S)- enantiomers.008863987

[0154] 17

[0155] In the definitions of the R3and R4groups in the present application, one possibility for these groups refers to connected to the “neighbouring pyridine ring” or the “pyridine ring”. The “pyridine ring” refers to the pyridine ring in the dashed circle below for completeness:

[0156]

[0157] Protected Forms

[0158] Compounds of the invention may be provided in a protected form. Here, reactive functionality, such as amino functionality, may be masked in order to prevent its reaction during a synthesis step. A protecting group is provided to mask the reactive functionality, and this protecting groups may be removed at a later stage of the synthesis to reveal the original reactive functionality.

[0159] In one embodiment, the protected form is a compound where the amino group of the compound is protected. Protecting groups, such as those for amino acid residues, are well known and well described in the art.

[0160] Where a protecting group is used is it is removable under conditions that do not substantially disrupt the structure of the polymyxin core, for example conditions that do not alter the stereochemistry of the amino acid residues.

[0161] In one embodiment, the protecting groups are acid-labile, base labile, or are removable under reducing conditions.

[0162] Example protecting groups for amino functionality include Boc (tert-butoxycabonyl), Bn (benzyl, Bzl), CbZ (benzyloxycarbonyl, Z), 2-CL-Z (2-chloro), ivDde (1-[4,4-dimethyl-2,6-dioxocylcohex-1-ylidene]-3-methylbutyl), Fmoc (fluorenylmethyloxycarbonyl), HSO3-Fmoc (sulfonylated Fmoc, such as 2-sulfo-Fmoc, as described in e.g. Schechter et al, J. Med Chem 2002, 45 (19) 4264), Dde (1-[4,4-dimethyl-2,6-dioxocylcohex-1-ylidene]ethyl), Mmt (4-methoxytrityl), Mtt (4 methyltrityl), Nvoc (6 nitroveratroyloxycarbonyl), Tfa (trifl uroacetyl), and Alloc (allyloxycarbonyl). Example protecting groups for aromatic nitrogen functionality includes Boc, Mtt, Trt and Dnp (dinitrophenyl).

[0163] Salts, Solvates and Other Forms

[0164] Examples of salts of compound of Formula (I) include all pharmaceutically acceptable salts, such as, without limitation, acid addition salts of strong mineral acids such as HCI and HBr salts008863987

[0165] 18

[0166] and addition salts of strong organic acids such as a methanesulfonic acid salt. Further examples of salts include sulfates and acetates such as acetate itself, trifluoroacetate or trichloroacetate. Examples of suitable salt forms can be found in e.g. Bharate, S.S., Drug Discovery Today (2020) “Recent developments in pharmaceutical salts: FDA approvals from 2015 to 2019’’, and all references therein.

[0167] A compound of Formula (I) can also be formulated as a prodrug. Prodrugs can include a MASTL inhibiting compound herein described in which one or more amino groups are protected with a group which can be cleaved in vivo, to liberate the biologically active compound. In one embodiment the prodrug is an “amine prodrug”. Examples of amine prodrugs include sulphomethyl, as described in e.g., Bergen et al, Antimicrob. Agents and Chemotherapy, 2006, 50, 1953 or HSO3-FMOC, as described in e.g. Schechter et al, J. Med Chem 2002, 45(19) 4264, and salts thereof. Further examples of amine prodrugs are given by Krise and Oliyai in Biotechnology: Pharmaceutical Aspects, 2007, 5(2), 101-131.

[0168] A reference to a compound of Formula (I) or any other compound described herein, is also a reference to a solvate of that compound. Examples of solvates include hydrates.

[0169] A compound of Formula (I) or any other compound described herein, includes a compound where an atom is replaced by a naturally occurring or non-naturally occurring isotope. In one embodiment the isotope is a stable isotope. Thus a compound described here includes, for example deuterium containing compounds and the like. For example, H may be in any isotopic form, including 1 H, 2H (D), and 3H (T); C may be in any isotopic form, including 12C, 13C, and 14C.

[0170] Chirality

[0171] Certain compounds of Formula (I) may exist in one or more isomers particular geometric, optical, enantiomeric, diasteriomeric, epimeric, atropic, stereoisomeric, tautomeric, conformational, or anomeric forms, including but not limited to, cis- and trans-forms; E- and Z-forms; c-, t-, and r- forms; endo- and exo-forms; R-, S-, and meso-forms; D- and L-forms; d- and l-forms; (+) and (-) forms; keto-, enol-, and enolate-forms; syn- and anti-forms; synclinal- and anticlinal-forms; a- and p-forms; axial and equatorial forms; boat-, chair-, twist-, envelope-, and halfchair-forms; and combinations thereof, hereinafter collectively referred to as “isomers” (or “isomeric forms”).

[0172] In particular, compounds of Formula (I) may have a chiral centre on the carbon bonded to R2and R3. Formula (I) may therefore have two enantiomeric forms (as shown below). These forms may be stereoisomers if there are further chiral centres present in the molecule.008863987

[0173]

[0174] Both enantiomers have high biological activity.

[0175] In one embodiment, the compound of Formula (I) is provided as a racemic mixture. That is, the compound of Formula (I) contains a 50:50 mixture of the R:S forms.

[0176] In one embodiment, the compound of formula (I) is provided in a form that is at least 60% optically enriched (i.e. , 60% of the compound, on a molar basis, is the one of the R or S stereoisomer or enantiomer, and the remaining 40% is the S or R stereoisomer or enantiomer respectively), e.g., at least 70% optically pure, e.g., at least 80% optically pure, e.g., at least 90% optically pure, e.g., at least 95% optically pure, e.g., at least 97% optically pure, e.g., at least 98% optically pure, e.g., at least 99% optically pure.

[0177] In one embodiment, the compound of formula (I) is provided in a form that is at least 60% optically pure R stereoisomer or enantiomer (i.e., 60% of the compound, on a molar basis, is the R stereoisomer or enantiomer, and 40% is the S stereoisomer or enantiomer), e.g., at least 70% optically pure, e.g., at least 80% optically pure, e.g., at least 90% optically pure, e.g., at least 95% optically pure, e.g., at least 97% optically pure, e.g., at least 98% optically pure, e.g., at least 99% optically pure.

[0178] In one embodiment, the compound of formula (I) is provided in a form that is at least 60% optically pure S stereoisomer or enantiomer (i.e., 60% of the compound, on a molar basis, is the S stereoisomer or enantiomer, and 40% is the R stereoisomer or enantiomer), e.g., at least 70% optically pure, e.g., at least 80% optically pure, e.g., at least 90% optically pure, e.g., at least 95% optically pure, e.g., at least 97% optically pure, e.g., at least 98% optically pure, e.g., at least 99% optically pure.

[0179] Enantiomeric purity may be measured by methods such as chiral HPLC.

[0180] Unless otherwise specified, a reference to a particular compound includes all such isomeric forms, including mixtures (e.g., racemic mixtures) thereof. Methods for the preparation (e.g., asymmetric synthesis) and separation (e.g., fractional crystallisation and chromatographic means) of such isomeric forms are either known in the art or are readily obtained by adapting the methods taught herein, or known methods, in a known manner.

[0181] For completeness, the term “isomer(s)” as used herein does not include structural (or constitutional) isomers (i.e., isomers which differ in the connections between atoms rather than merely by the position of atoms in space). For example, a reference to a methoxy008863987

[0182] 20

[0183] group, -OCH3, is not to be construed as a reference to its structural isomer, a hydroxymethyl group, -CH2OH. A reference to isomeric forms of the compounds of the invention requires the connections between atoms as required by the formulae (and terminology) herein. However, a reference to a class of structures may well include structurally isomeric forms falling within that class (e.g., C1-6 alkyl includes n-propyl and iso-propyl; butyl includes n-, iso-, sec-, and tert-butyl; methoxyphenyl includes ortho-, meta-, and para-methoxyphenyl).

[0184] Formulations

[0185] In one aspect, the present invention provides a pharmaceutical composition comprising a compound of formula (I) together with a pharmaceutically acceptable carrier. The pharmaceutical composition may additionally comprise a second active agent. In an alternative embodiment, where a second active agent is provided for use in therapy, the second active agent may be separately formulated from the compound of formula (I). The comments below made in relation to the compound of formula (I) may therefore also apply to the second active agent, as separately formulated.

[0186] While it is possible for the compound of formula (I) to be administered alone or together with the second active agent, it is desirable to present it as a pharmaceutical formulation (e.g., composition, preparation, medicament) comprising at least one compound of formula (I) as described herein, together with one or more other pharmaceutically acceptable ingredients well known to those skilled in the art, including, but not limited to, pharmaceutically acceptable carriers, diluents, excipients, adjuvants, fillers, buffers, preservatives, anti-oxidants, lubricants, stabilisers, solubilisers, surfactants (e.g., wetting agents), masking agents, colouring agents, flavouring agents, and sweetening agents. The formulation may further comprise other active agents, for example, other therapeutic or prophylactic agents.

[0187] Thus, the present invention further provides pharmaceutical compositions, as defined above, and methods of making a pharmaceutical composition comprising admixing at least one compound of formula (I) as described herein, together with one or more other pharmaceutically acceptable ingredients well known to those skilled in the art, e.g., carriers, diluents, excipients, etc. If formulated as discrete units (e.g., tablets, etc.), each unit contains a predetermined amount (dosage) of the compound. The composition optionally further comprises the second active agent in a predetermined amount.

[0188] The term “pharmaceutically acceptable,” as used herein, pertains to compounds, ingredients, materials, compositions, dosage forms, etc., which are, within the scope of sound medical judgment, suitable for use in contact with the tissues of the subject in question (e.g., human) without excessive toxicity, irritation, allergic response, or other problem or complication, commensurate with a reasonable benefit / risk ratio. Each carrier, diluent, excipient, etc. must also be “acceptable” in the sense of being compatible with the other ingredients of the formulation.

[0189] Suitable carriers, diluents, excipients, etc. can be found in standard pharmaceutical texts, for example, Remington's Pharmaceutical Sciences, 18th edition, Mack Publishing Company, Easton, Pa., 1990; and Handbook of Pharmaceutical Excipients, 5th edition, 2005.008863987

[0190] 21

[0191] The formulations may be prepared by any methods well known in the art of pharmacy. Such methods include the step of bringing into association the compound of formula (I) with a carrier which constitutes one or more accessory ingredients. In general, the formulations are prepared by uniformly and intimately bringing into association the compound with carriers (e.g., liquid carriers, finely divided solid carrier, etc.), and then shaping the product, if necessary.

[0192] The formulation may be prepared to provide for rapid or slow release; immediate, delayed, timed, or sustained release; or a combination thereof.

[0193] Formulations may suitably be in the form of liquids, solutions (e.g., aqueous, non-aqueous), suspensions (e.g., aqueous, non-aqueous), emulsions (e.g., oil-in-water, water-in-oil), elixirs, syrups, electuaries, mouthwashes, drops, tablets (including, e.g., coated tablets), granules, powders, losenges, pastilles, capsules (including, e.g., hard and soft gelatin capsules), cachets, pills, ampoules, boluses, suppositories, pessaries, tinctures, gels, pastes, ointments, creams, lotions, oils, foams, sprays, mists, or aerosols.

[0194] The formulation may be suitable for methods of administration including but not limited to: intratumoral administration, oral administration, parenteral administration (e.g. intravenous or subcutaneous injection), pulmonary administration and transdermal administration.

[0195] Medical Uses

[0196] In some embodiments, the compounds of formula (I) may be for use in a method of treatment or prophylaxis of the human or animal body by therapy.

[0197] The compounds of formula (I) may be for use in a method of treating cancer. In some cases, the cancer may be a solid tumour. In some cases, the cancer may be breast cancer, colon cancer, colorectal cancer, head and neck squamous cell carcinoma, non-small cell lung cancer (NSCLC) gastric cancer, pancreatic cancer, prostate cancer or thyroid cancer. In particular, the cancer may be breast cancer, colorectal cancer, non-small cell lung cancer (NSCLC) or pancreatic cancer.

[0198] The compounds of formula (I) or pharmaceutical formulations containing these compounds, are suitable for use in methods of treatment and prophylaxis. The compounds may be administered to a subject in need thereof. In some aspects of the invention, a compound of formula (I) may be administered to a subject, such as a mammalian subject (e.g. a human), in order to treat a cancer.

[0199] Another aspect of the present invention pertains to use of a compound of formula (I) in the manufacture of a medicament for use in treatment. In one embodiment, the medicament comprises a compound of formula (I). In one embodiment, the medicament is for use in the treatment of a cancer.

[0200] The term “treatment,” as used herein in the context of treating a condition, pertains generally to treatment and therapy, whether of a human or an animal (e.g., in veterinary applications), in which some desired therapeutic effect is achieved, for example, the inhibition of the progress of the condition, and includes a reduction in the rate of progress, a halt in the rate of progress,008863987

[0201] 22

[0202] alleviation of symptoms of the condition, amelioration of the condition, and cure of the condition. Treatment as a prophylactic measure (i.e., prophylaxis) is also included. For example, use with patients who have not yet developed the condition, but who are at risk of developing the condition, is encompassed by the term “treatment.”

[0203] The term “therapeutically-effective amount,” as used herein, pertains to that amount of a compound, or a material, composition or dosage form comprising a compound, which is effective for producing some desired therapeutic effect, commensurate with a reasonable benefit / risk ratio, when administered in accordance with a desired treatment regimen.

[0204] The term “treatment” includes combination treatments and therapies, in which two or more treatments or therapies are combined, for example, sequentially or simultaneously.

[0205] *****

[0206] The features disclosed in the foregoing description, or in the following claims, or in the accompanying drawings, expressed in their specific forms or in terms of a means for performing the disclosed function, or a method or process for obtaining the disclosed results, as appropriate, may, separately, or in any combination of such features, be utilised for realising the invention in diverse forms thereof.

[0207] While the invention has been described in conjunction with the exemplary embodiments described above, many equivalent modifications and variations will be apparent to those skilled in the art when given this disclosure. Accordingly, the exemplary embodiments of the invention set forth above are considered to be illustrative and not limiting. Various changes to the described embodiments may be made without departing from the spirit and scope of the invention.

[0208] For the avoidance of any doubt, any theoretical explanations provided herein are provided for the purposes of improving the understanding of a reader. The inventors do not wish to be bound by any of these theoretical explanations.

[0209] Any section headings used herein are for organizational purposes only and are not to be construed as limiting the subject matter described.

[0210] Throughout this specification, including the claims which follow, unless the context requires otherwise, the word “comprise” and “include”, and variations such as “comprises”, “comprising”, and “including” will be understood to imply the inclusion of a stated integer or step or group of integers or steps but not the exclusion of any other integer or step or group of integers or steps. It must be noted that, as used in the specification and the appended claims, the singular forms “a,” “an,” and “the” include plural referents unless the context clearly dictates otherwise. Ranges may be expressed herein as from “about” one particular value, and / or to “about” another particular value. When such a range is expressed, another embodiment includes from the one particular value and / or to the other particular value. Similarly, when values are expressed as approximations, by the use of the antecedent “about,” it will be understood that the particular value forms another embodiment. The term “about” in relation to a numerical value is optional and means for example + / - 10%.008863987

[0211] Examples

[0212] Synthesis Examples

[0213] Compound 1

[0214]

[0215] To a stirred mixture of bicyclo[1.1.1]pentane-1,3-dicarboxylic acid (36.6 g, 234 mmol, 1.0

[0216] equiv) in H2O (1.0 L) were added 4-(chloromethyl)-1-fluoro-1,4-diazabicyclo[2.2.2]octane-1,4-diium; bis(tetrafluoroboranuide) (207.6 g, 586 mmol, 2.5 equiv) and AgNOa (2.99 g, 17.6 mmol, 0.075 equiv) at room temperature under nitrogen atmosphere. The resulting mixture was stirred at 67 °C for 4 h under nitrogen atmosphere. The mixture was allowed to cool down to room temperature. The resulting mixture was filtered, the filter cake was washed with H2O. The

[0217] filtrate was extracted with tert-Butyl Methyl ether (3 x 1000 mL). The combined organic layers

[0218] were washed with brine, dried over anhydrous Na2SO4. After filtration, the filtrate was

[0219] concentrated under reduced pressure. The resulting mixture was concentrated under

[0220] vacuum. This resulted in 3-fluorobicyclo[1.1.1]pentane-1-carboxylic acid (19.0 g, crude) as

[0221] a brown solid.

[0222] LCMS (ESI, m / z): 129.05 [M-H]’008863987

[0223] 24

[0224] Step (ii)

[0225]

[0226] To a stirred solution of 3-fluorobicyclo[1.1.1]pentane-1 -carboxylic acid (10.4 g, 71.0 mmol, 1.0 equiv) in THF (100 mL) were added UAIH4 (5.39 g, 142.0 mmol, 2.0 equiv) in portions at 0 °C. The resulting mixture was stirred at room temperature for 2 h. The reaction was quenched with 1 N HCI (aq.) at 0 °C. The resulting mixture was extracted with EtOAc (3 x 15 mL). The combined organic layers were washed with brine, dried over anhydrous Na2SO4. After filtration, the filtrate was concentrated under reduced pressure. This resulted in {3-fluorobicyclo[1.1.1]pentan-1-yl}methanol (7.30 g, crude) as a yellow oil.

[0227] Step (Hi)

[0228]

[0229] A solution of (COCI)2 (16.39 g, 129 mmol, 1.5 equiv) in DCM (528 mL) was treated with DMSO (20.18 g, 258 mmol, 3.0 equiv) at -78 °C for 15 min under nitrogen atmosphere followed by the addition of {3-fluorobicyclo[1.1.1]pentan-1-yl}methanol (10.0 g, 86.107 mmol, 1.0 equiv) in DCM (128 mL) dropwise at -78 °C. The resulting mixture was stirred at -78 °C for 50 min under nitrogen atmosphere. To the above mixture was added Et3N (88 mL) dropwise over 5 min at -78 °C. The resulting mixture was stirred at -78 °C for additional 15 min. The mixture was warmed up to room temperature. The reaction was quenched with water at room temperature. The resulting mixture was extracted with DCM (3 x 500 mL). The combined organic layers were washed with brine, dried over anhydrous Na2SO4. After filtration, the filtrate was concentrated under reduced pressure. This resulted in 3-fluorobicyclo[1.1.1]pentane-1-carbaldehyde (9.60 g, crude) as a yellow oil.

[0230] Step (iv)

[0231]

[0232] Step 4

[0233] To a stirred mixture of 3-fluorobicyclo[1.1.1]pentane-1-carbaldehyde (9.60 g, 84.1 mmol, 1.0 equiv) and 2-methylpropane-2-sulfinamide (48.02 g, 396 mmol, 4.71 equiv) in THF (50 mL) were added tetrakis(propan-2-yloxy)titamum (23.91 g, 84.1 mmol, 1.0 equiv) at room008863987

[0234] 25

[0235] temperature under nitrogen atmosphere. The resulting mixture was stirred at 65 °C for 3 h under air atmosphere. The reaction was quenched with water at room temperature. The resulting mixture was filtered, the filter cake was washed with ethyl acetate (3 x 100 mL). The filtrate was extracted with EtOAc (3 x 200 mL). The combined organic layers were washed with brine, dried over anhydrous Na2SO4. After filtration, the filtrate was concentrated under reduced pressure. The residue was purified by silica gel column chromatography, eluted with PE I EA (3:1) to afford / V-[(1E)-{3-fluorobicyclo[1.1.1]pentan-1-yl}methylidene]-2-methylpropane-2-sulfinamide (8.6 g, 47.05%) as a yellow solid.

[0236] LCMS (ESI, m / z): 218.15[M+H]+

[0237]

[0238] A solution of 2,4-dibromopyridine (18.75 g, 79.2 mmol, 2.0 equiv) in Diethyl ether (200 mL) was treated with n-BuLi (5.07 g, 79.2 mmol, 2.0 equiv) at -78 °C for 15 min under nitrogen atmosphere followed by the addition of / V-[(1E)-{3-fluorobicyclo[1.1.1]pentan-1-yl}methylidene]-2-methylpropane-2-sulfinamide (8.6 g, 39.6 mmol, 1.0 equiv) in Diethyl ether (86 mL) dropwise at -78 °C. The resulting mixture was stirred at -78 °C for 1 h under nitrogen atmosphere. The reaction was quenched with sat. NH4CI (aq.) at 10 °C. The resulting mixture was extracted with EtOAc (3 x 300 mL). The combined organic layers were washed with brine, dried over anhydrous Na2SO4. After filtration, the filtrate was concentrated under reduced pressure. The residue was purified by silica gel column chromatography, eluted with PE I EA (1:1.2) to afford / V-[(2-bromopyridin-4-yl)({3-fluorobicyclo[1.1.1]pentan-1-yl})methyl]-2-methylpropane-2-sulfinamide (4.0 g, 26.93%) as a yellow solid.

[0239] LCMS (ESI, m / z): 375.05 [M+H]+

[0240]

[0241] Step 6

[0242] To a stirred mixture of / V-[(2-bromopyridin-4-yl)({3-fluorobicyclo[1.1.1]pentan-1-yl})methyl]-2-methylpropane-2-sulfinamide (3.6 g, 9.59 mmol, 1 equiv) and diphenylmethanimine (2.09 g, 11.51 mmol, 1.2 equiv) in toluene (120 mL) were added CS2CO3 (9.38 g, 28.78 mmol, 3.0 equiv), BINAP (597.3 mg, 0.96 mmol, 0.1 equiv) and Pd(OAc)2 (215.4 mg, 0.96 mmol, 0.1008863987

[0243] 26

[0244] equiv) at room temperature. The resulting mixture was stirred at 120 °C for 3 h

[0245] under nitrogen atmosphere. The reaction was quenched with water at room temperature. The resulting mixture was concentrated under reduced pressure. The residue was purified by reversed-phase flash chromatography [with the following conditions: column, C18 silica gel; mobile phase, MeCN in Water (0.05% NH3 H2O), 70% to 100% gradient in 25 min; detector, UV 254 nm]. This resulted in / V-({2-[(diphenylmethylidene)amino]pyridin-4-yl}({3-fluorobicyclo[1.1.1]pentan-1-yl})methyl)-2-methylpropane-2-sulfinamide (1.0 g, 21.92%) as a yellow solid.

[0246] LCMS (ESI, m / z): 476.15 [M+H]+

[0247] Step (vii)

[0248]

[0249] To a stirred mixture of / V-({2-[(diphenylmethylidene)amino]pyridin-4-yl}({3-fluorobicyclo[1.1.1]pentan-1-yl})methyl)-2-methylpropane-2-sulfinamide (1.0 g, 2.10 mmol, 1 equiv) in THF (5 mL) and H2O (5 mL) were added HCI (1 M, 5 mL) dropwise at 0 °C. The resulting mixture was stirred at room temperature for 1 h. The mixture was basified to pH 8 with saturated NaHCCh (aq.). The residue was purified by reversed-phase flash chromatography [with the following conditions: column, C18 silica gel; mobile phase, MeCN in Water (0.05% NH3 H2O), 0% to 40% gradient in 10 min; detector, UV 254 nm]. This resulted in / \ / -[(2-aminopyridin-4-yl)({3-fluorobicyclo[1.1.1]pentan-1-yl})methyl]-2-methylpropane-2-sulfinamide (380 mg, 58.04%) as a yellow solid.

[0250] LCMS (ESI, m / z): 312.00 [M+H]+

[0251] Step (viii)

[0252]

[0253] p

[0254] A solution of 6-chloro-4-methylpyridine-3-carbonitrile (2.0 g, 13.11 mmol, 1 equiv) in THF (50 mL) was treated with LiHMDS (1.0 M in THF) (2.5 mL, 3.81 equiv) at -78 °C for 30 min under nitrogen atmosphere followed by the addition of methyl 1-methyl-1 / 7-1,2,3-triazole-5-carboxylate008863987

[0255] 27

[0256] (1.85 g, 13.11 mmol, 1.0 equiv) dropwise at -78 °C. The resulting mixture was stirred at room temperature for 2 h under nitrogen atmosphere. To the above mixture was added NH4OAC (10.31 g, 133.70 mmol, 10.2 equiv) and acetic acid (74.0 mL) dropwise over 5 min at 0 °C. The resulting mixture was stirred at 70 °C for additional 3 h. The resulting mixture was concentrated under reduced pressure. The resulting mixture was diluted with water (5 mL). The residue was basified to pH 8 with saturated K2CO3 (aq.). The precipitated solids were collected by filtration and washed with H2O, PE and DCM. The resulting solid was dried under vacuum. This resulted in 6-chloro-3-(3-methyl-1,2,3-triazol-4-yl)-2,7-naphthyridin-1-amine (2.68 g, 78.43%) as a brown solid.

[0257] LCMS (ESI, m / z): 261.10 [M+H]+

[0258] Step (ix)

[0259]

[0260] To a stirred mixture of / V-[(2-aminopyridin-4-yl)({3-fluorobicyclo[1.1.1]pentan-1-yl})methyl]-2-methylpropane-2-sulfinamide (380 mg, 1.22 mmol, 1 equiv) and 6-chloro-3-(3-methyl-1,2,3-triazol-4-yl)-2,7-naphthyridin-1-amine (477.2 mg, 1.83 mmol, 1.5 equiv) in Dioxane (5 mL) were added CS2CO3 (1.19 g, 3.66 mmol, 3.0 equiv) and BrettPhos Pd G3 (331.8 mg, 0.37 mmol, 0.3 equiv) at room temperature under nitrogen atmosphere. The resulting mixture was stirred at 100 °C for 2 h under nitrogen atmosphere. The resulting mixture was concentrated under reduced pressure. The residue was purified by reversed-phase flash chromatography [with the following conditions: column, C18 silica gel; mobile phase, MeCN in Water (0.05% NH3 H2O), 10% to 50% gradient in 10 min; detector, UV 254 nm]. This resulted in / \ / -[(2-{[8-amino-6-(3-methyl-1 ,2,3-triazol-4-yl)-2,7-naphthyridin-3-yl]amino}pyridin-4-yl)({3-fluorobicyclo[1.1.1 ]pentan-1-yl})methyl]-2-methylpropane-2-sulfinamide (140 mg, 21.42%) as a light yellow solid.

[0261] LCMS (ESI, m / z): 536.20 [M+H]008863987

[0262]

[0263] Into a 50 mL round-bottom flask were added HCI in 1,4-dioxane (4.0 M, 2 mL) at room temperature. The resulting mixture was stirred at room temperature for 1 h under nitrogen atmosphere. The mixture was basified to pH 8 with saturated NaHCCh (aq.). The residue was purified by reversed-phase flash chromatography [with the following conditions: column, C18 silica gel; mobile phase, MeCN in Water (0.05% NH3 H2O), 0% to 40% gradient in 10 min; detector, UV 254 nm]. This resulted in N6-{4-[amino({3-fluorobicyclo[1.1.1]pentan-1-yl}) methyl] pyridin-2-yl}-3-(3-methyl- 1 ,2,3-triazol-4-yl)-2,7-naphthyridine-1 ,6-diamine (72.9 mg, 64.64%) as a light yellow solid.

[0264] LCMS (ESI, m / z): 432.10 [M+H]+

[0265] 1H N MR (400 MHz, DMSO-cfe) 69.89 (s, 1H), 9.31 (s, 1H), 8.29 (s, 1H), 8.25 - 8.18 (m, 2H), 7.35 (s, 2H), 7.29 - 7.21 (m, 2H), 6.85 (dd, J = 5.2, 1.4 Hz, 1 H), 4.39 (s, 3H), 4.09 (s, 1 H), 1.94 - 1.70 (m, 6H).

[0266] Separation of Enantiomers of Compound 1

[0267]

[0268] The R- and S- enantiomers of Compound 1 were separated by chiral HPLC using a CHIRALPAK IK-3 column in 50:50 Hex(0.1%DEA):ETOH. The flow rate was 1.67ml / min and the analysis was performed at room temperature. Detection wavelength was 254 nm. Retention times were:

[0269] Compound 1A (enantiomer 1): 1.360 min

[0270] Compound 1B (enantiomer 2): 1.971 min

[0271] The absolute configuration of enantiomer 1 and enantiomer 2 is not known.008863987

[0272] 29

[0273] Compound 2

[0274]

[0275] To a mixture of methyl 6,6-difluorobicyclo[3.1.0]hexane-3-carboxylate (19.0 g, 107 mmol, 1.00 equiv) in THF (200 mL) was added UAIH4 (8.19 g, 215 mmol, 2.00 equiv) dropwise at 0 °C. The resulting mixture was stirred at room temperature for 2 h. The reaction was quenched with HCI

[0276] (1 M) at 0 °C. The resulting mixture was extracted with EtOAc (3 x 200 mL), dried over

[0277] anhydrous Na2SC>4. After filtration, the filtrate was concentrated under reduced pressure. This resulted in {6,6-difluorobicyclo [3.1.0] hexan-3-yl} methanol as a yellow oil (15.0 g, crude).

[0278] Step (77)

[0279]

[0280] crude008863987

[0281] 30

[0282] To a mixture of {6,6-difluorobicyclo[3.1.0] hexan-3-yl}methanol (15.0 g, 101 mmol, 1.00 equiv) in DCM (200 mL) was added Dess-Martin (85.8 g, 202 mmol, 2.00 equiv) in portions at 0 °C. The resulting mixture was stirred at room temperature for 3 h. The reaction was quenched with water at 0 °C. The resulting mixture was extracted with CH2CI2 (3 x 200 mL), dried over anhydrous Na2SO4. After filtration, the filtrate was concentrated under reduced pressure. The residue was purified by silica gel column chromatography, eluted with PE I EA (5:1) to afford 6,6-difluorobicyclo [3.1.0] hexane-3-carbaldehyde as a yellow oil (13.0 g crude).

[0283] LCMS (ESI, m / z): No mass

[0284] Step (Hi)

[0285]

[0286] 45.0%

[0287] To a mixture of 6,6-difluorobicyclo [3.1.0] hexane-3-carbaldehyde (13.0 g, 88.9 mmol, 1.00 equiv) and tert-butanesulfinamide (12.9 g, 106 mmol, 1.20 equiv) in THF (300 mL) was added Ti(0Et)4 (40.5 g, 177 mmol, 2.00 equiv) at room temperature. The resulting mixture was stirred at 65 °C for 3 h. The reaction was quenched with water at room temperature. The resulting mixture was filtered, the filter cake was washed with water (3 x 100 mL). The filtrate was concentrated under reduced pressure. The resulting mixture was extracted with EtOAc (3 x 300 mL), dried over anhydrous Na2SC>4. After filtration, the filtrate was concentrated under reduced pressure. The residue was purified by silica gel column chromatography, eluted with PE I EA (5:1) to afford N-[(1E)-{6,6-difluorobicyclo [3.1.0] hexan-3-yl}methylidene]-2-methylpropane-2-sulfinamide as a yellow oil (10.0 g, 45.0%).

[0288] LCMS (ESI, m / z): 250.10 [M+H]+

[0289]

[0290] A solution of 2,4-dibromopyridine (19.0 g, 80.2 mmol, 2.00 equiv) in 2-(propan-2-yloxy)propane (150 mL) was treated with n-BuLi (2.5 M, 32.1 mL, 80.2 mmol, 2.00 equiv) at -78 °C for 30 min under nitrogen atmosphere followed by the addition of N-[(1E)-{6,6-difluorobicyclo[3.1.0]hexan-3-yl}methylidene]-2-methylpropane-2-sulfinamide (10.0 g, 40.1 mmol, 1.00 equiv) in Et20 dropwise at -78 °C. The resulting mixture was stirred at -78 °C for 1 h under nitrogen atmosphere. The reaction was quenched with sat. NH4CI (aq.) at 0 °C. The008863987

[0291] 31

[0292] resulting mixture was extracted with EtOAc (3 x 150 mL), dried over anhydrous Na2SO4. After filtration, the filtrate was concentrated under reduced pressure. The residue was purified by silica gel column chromatography, eluted with PE I EA (1:1) to afford N-[(2-bromopyridin-4-yl) ({6,6-difluorobicyclo [3.1.0] hexan-3-yl}) methyl]-2-methylpropane-2-sulfinamide as a yellow oil (5.00 g, 30.6%).

[0293] LCMS (ESI, m / z): 407.10 [M+H]+

[0294] Step (v)

[0295]

[0296] p

[0297] 40.1%

[0298] To a mixture of N-[(2-bromopyridin-4-yl)({6,6-difluorobicyclo[3.1.0]hexan-3-yl})methyl]-2-methylpropane-2-sulfinamide (5.00 g, 12.2 mmol, 1.00 equiv) and a-phenyl-benzenemethanimine (2.67 g, 14.7 mmol, 1.20 equiv) in toluene (150 mL) were added CS2CO3 (12.0 g, 36.8 mmol, 3.00 equiv), BINAP (764 mg, 1.22 mmol, 0.10 equiv) and Pd(OAc)2 (275 mg, 1.22 mmol, 0.10 equiv) at room temperature under nitrogen atmosphere. The resulting mixture was stirred at 120 °C for 3 h under nitrogen atmosphere. The reaction was quenched with water at room temperature. The resulting mixture was concentrated under reduced pressure. The resulting mixture was extracted with EtOAc (3 x 100 mL), dried over anhydrous Na2SC>4. After filtration, the filtrate was concentrated under reduced pressure. The residue was purified by silica gel column chromatography, eluted with PE / EA (1:1) to afford N-({6,6-difluorobicyclo [3.1.0] hexan-3-yl} ({2-[(diphenylmethylidene)amino] pyridin-4-yl}) methyl)-2-methylpropane-2-sulfinamide as a yellow oil (2.50 g, 40.1%).

[0299] LCMS (ESI, m / z): 508.10 [M+H]+

[0300] Step (vi)

[0301]

[0302] To a mixture of N-({6,6-difluorobicyclo [3.1.0] hexan-3-yl} ({2-[(diphenylmethylidene)amino] pyridin-4-yl}) methyl)-2-methylpropane-2-sulfinamide (2.00 g, 3.94 mmol, 1.00 equiv) in H2O (20.0 mL) and THF (20.0 mL) was added HCI (1 M, 20.0 mL) dropwise at 0 °C. The resulting mixture was stirred at room temperature for 1 h. The mixture was neutralized to pH

[0303] 7 with NaOH. The resulting mixture was concentrated under reduced pressure. The residue was008863987

[0304] 32

[0305] purified by reversed-phase flash chromatography [with the following conditions: column, C18 silica gel; mobile phase, ACN in water (10 mmol / L NH4HCO3), 0% to 100% gradient in 40 min; detector, UV 254 nm]. This resulted in N-[(2-aminopyridin-4-yl) ({6,6-difluorobicyclo [3.1.0] hexan-3-yl}) methyl]-2-methylpropane-2-sulfinamide as a yellow solid (800 mg, 59.1%).

[0306] LCMS (ESI, m / z): 344.10 [M+H]+

[0307] Step (vii)

[0308]

[0309] To a mixture of N-[(2-aminopyridin-4-yl) ({6,6-difluorobicyclo [3.1.0] hexan-3-yl})methyl]-2-methylpropane-2-sulfinamide (750 mg, 2.18 mmol, 1.00 equiv) and 6-chloro-3-(3-methyl-1,2,3-triazol-4-yl)-2,7-naphthyridin-1 -amine (853 mg, 3.27 mmol, 1.50 equiv) in 1,4-dioxane (24.0 mL) were added CS2CO3 (2.13 g, 6.55 mmol, 3.00 equiv) and BrettPhos Pd G3 (39.5 mg, 0.04 mmol, 0.02 equiv) at room temperature under nitrogen atmosphere. The resulting mixture was stirred at 100 °C for 2 h under nitrogen atmosphere. The reaction was quenched

[0310] with water at room temperature. The resulting mixture was concentrated under reduced pressure. The residue was purified by reversed-phase flash chromatography [with the following conditions: column, C18 silica gel; mobile phase, MeCN in Water (10 mmol / L NH4HCO3), 0% to 100% gradient in 40 min; detector, UV 254 nm]. This resulted in N-[(2-{[8-amino-6-(3-methyl-1,2,3-triazol-4-yl)-2,7-naphthyridin-3-yl] amino} pyridin-4-yl) ({6,6-difluorobicyclo [3.1.0] hexan-3-yl}) methyl]-2-methylpropane-2-sulfinamide as a yellow solid (400 mg, 32.2%).

[0311] LCMS (ESI, m / z): 368.10 [M+H]+008863987

[0312] Step (viii)

[0313]

[0314] Compound 2

[0315] Into an 8 mL round-bottom flask were added N-[(2-{[8-amino-6-(3-methyl-1,2,3-triazol-4-yl)-2,7-naphthyridin-3-yl] amino} pyridin-4-yl) ({6,6-difluorobicyclo [3.1.0] hexan-3-yl}) methyl]-2-methylpropane-2-sulfinamide (400 mg, 0.71 mmol, 1.00 equiv) and HCI in 1,4-dioxane (4.0 M, 4.0 mL) at room temperature. The resulting mixture was stirred at room temperature for 1 h. The resulting mixture was concentrated under reduced pressure. The crude product was purified by reverse phase flash with the following conditions [Column: X-Bridge Prep OBD C18 Column 30 *150 mm, 5 pm; Mobile Phase A: Water (10 mmol / L NH4HCC>3+0.05% NH3.H2O), Mobile Phase B: ACN; Flow rate: 60 mL / min; Gradient (B%): 16% B to 46% B in 10 min; Wave Length:

[0316] 254nm / 220 nm; RT1 (min): 9.37] to afford N6-{4-[amino({6,6-difluorobicyclo[3.1.0]hexan-3-yl})methyl]pyridin-2-yl}-3-(3-methyl-1 ,2,3-triazol-4-yl)-2,7-naphthyridine-1 ,6-diamine as a light yellow solid (134.8 mg, 41.2%).

[0317] LCMS (ESI, m / z): 464.15 [M+H]+

[0318] 1H NMR (400 MHz, DMSO-cfe) 69.87 (s, 1H), 9.30 (s, 1H), 8.33 - 8.17 (m, 3H), 7.38 - 7.19 (m, 4H), 6.86 - 6.81 (m, 1 H), 4.39 (s, 3H), 3.53 - 3.44 (m, 1 H), 2.27 - 2.05 (m, 4H), 1.93-1.87 (m, 1 H), 1.73-1.67 (m, 1 H), 1.62 - 1.52 (m, 1 H).

[0319] Separation of Enantiomers of Compound 2

[0320]

[0321] The R- and S- enantiomers of Compound 2 were separated by chiral HPLC using a CHIRALPAK IK-3 column with 50:50 Hex(0.1%DEA):(EtOH:CAN=5:1). Flow rate was 1.67 ml / min and the analysis was performed at room temperature. The detection wavelength was254 nm. Retention times were:

[0322] Compound 2A (enantiomer 1): 0.866 min

[0323] Compound 2B (enantiomer 2): 1.113 min008863987

[0324] 34

[0325] The absolute configuration of enantiomer 1 and enantiomer 2 is not known.

[0326] Comparative Example 1

[0327] &

[0328]

[0329] Step 1 crude

[0330] A solution of 6-chloro-4-methylpyridine-3-carbonitrile (1.00 g, 6.55 mmol, 1.00 equiv) in THF

[0331] (40.0 mL) was treated with LiHMDS (25.0 mL, 149 mmol, 22.8 equiv) at -65 °C for 30 min under nitrogen atmosphere followed by the addition of methyl 3-methyl-1,2,3-triazole-4-carboxylate

[0332] (926 mg, 6.56 mmol, 1.00 equiv) dropwise at -65 °C. The resulting mixture was stirred at room temperature for 2 h under nitrogen atmosphere. To the above mixture was added NH4OAC (5.20 g, 67.4 mmol, 10.2 equiv) in portions over 5 min at 0 °C. The resulting mixture was stirred at008863987

[0333] 35

[0334] 70 °C for additional 3 h. The resulting mixture was concentrated under reduced pressure. The mixture neutralized to pH 8 with K2CC>3(aq.). The precipitated solids were collected by filtration and washed with water (3 x 20 mL). This resulted in 6-chloro-3-(3-methyl-1,2,3-triazol-4-yl)-2,7-naphthyridin-1-amine as a brown solid (1.60 g, crude).

[0335] LCMS (ESI, m / z): 261.15 [M+H]+

[0336] Step (ii)

[0337]

[0338] ,

[0339] Step 2

[0340] crude

[0341] To a mixture of 3-pentanone (20.0 g, 232 mmol, 1.00 equiv) and tetraethoxytitanium (105 g, 464 mmol, 2.00 equiv) in THF (150 mL) was added terf-butanesulfinamide (28.1 g, 232 mmol, 1.00 equiv) at room temperature. The resulting mixture was stirred at 65 °C for overnight. The reaction was quenched with water at room temperature. The resulting mixture was extracted with EtOAc (3 x 200 mL). The combined organic layers were washed with brine (3 x 200 mL), dried over anhydrous Na2SO4. After filtration, the filtrate was concentrated under reduced pressure. This resulted in 2-methyl- / V-(pentan-3-ylidene)propane-2-sulfinamide as a yellow oil (26.0 g, crude).

[0342] LCMS (ESI, m / z): 190.10 [M+H]+

[0343] Step (Hi)

[0344]

[0345] A solution of 2,4-dibromopyridine (20.0 g, 84.4 mmol, 2.00 equiv) in Diethyl ether (300 mL) was treated with n-BuLi (34.0 mL, 530 mmol, 12.5 equiv) at -78 °C for 30 min under nitrogen atmosphere followed by the addition of 2-methyl- / V-(pentan-3-ylidene)propane-2-sulfinamide (8.00 g, 42.2 mmol, 1.00 equiv) dropwise at -78 °C. The resulting mixture was stirred at -78 °C for 1 h under nitrogen atmosphere. The reaction was quenched with sat. NH4CI (aq.) at 0 °C. The resulting mixture was extracted with EtOAc (4 x 100 mL), dried over

[0346] anhydrous Na2SO4. After filtration, the filtrate was concentrated under reduced pressure. The residue was purified by silica gel column chromatography, eluted with PE / EA (1:1) to afford N-008863987

[0347] 36

[0348] [3-(2-bromopyridin-4-yl)pentan-3-yl]-2-methylpropane-2-sulfinamide as a yellow oil (5.00 g, 34.0%).

[0349] LCMS (ESI, m / z): 347.10 [M+H]+

[0350] Step (iv) and (v)

[0351]

[0352] 2. 1M HCI Step 4 & 5

[0353] crude

[0354] To a mixture of / V-[3-(2-bromopyridin-4-yl)pentan-3-yl]-2-methylpropane-2-sulfinamide (5.00 g, 14.3 mmol, 1.00 equiv) and diphenylmethanimine (3.13 g, 17.2 mmol, 1.20 equiv) in toluene (75 mL) was added CS2CO3 (14.0 g, 43.1 mmol, 3.00 equiv), BINAP (900 mg, 1.44 mmol, 0.10 equiv) and Pd(OAc)2 (320 mg, 1.44 mmol, 0.10 equiv) at room temperature. The resulting mixture was stirred at 110 °C for overnight under nitrogen atmosphere. The reaction was quenched with water at room temperature. The resulting mixture was filtered, the filter cake was washed with toluene (3 x 50 mL). The filtrate was concentrated under reduced pressure. To the above mixture was added THF (20 mL), H2O (20 mL) and HCI (20 mL, 1M) in portions over 2 min at 0 °C. The resulting mixture was stirred at room temperature for additional 2 h. The residue was purified by reversed-phase flash chromatography [with the following conditions: column, C18 silica gel; mobile phase, MeCN in Water (0.1% FA), 0% to 100% gradient in 30 min; detector, UV 254 nm]. This resulted in / V-[3-(2-aminopyridin-4-yl)pentan-3-yl]-2-methylpropane-2-sulfinamide as a yellow oil (2.20 g, crude).

[0355] LCMS (ESI, m / z): 284.10 [M+H]+

[0356] Step (vi)

[0357]

[0358] 29.2%

[0359] To a mixture of / V-[3-(2-aminopyridin-4-yl)pentan-3-yl]-2-methylpropane-2-sulfinamide (2.10 g, 7.40 mmol, 1.00 equiv) and 6-chloro-3-(3-methyl-1,2,3-triazol-4-yl)-2,7-naphthyridin-1-amine008863987

[0360] 37

[0361] (2.32 g, 8.89 mmol, 1.20 equiv) in 1,4-dioxane (21.0 mL) were added CS2CO3 (4.83 g, 14.8 mmol, 2.00 equiv) and BrettPhos Pd G3 (671 mg, 0.74 mmol, 0.100 equiv) at room temperature. The resulting mixture was stirred at 100 °C for 3 h under nitrogen atmosphere. The reaction was quenched with water at room temperature. The resulting mixture was extracted with EtOAc (3 x 50 mL), dried over anhydrous Na2SO4. After filtration, the filtrate was concentrated under reduced pressure. The residue was purified by silica gel column chromatography, eluted with CH2CI21 MeOH (9:1) to afford / \ / -[3-(2-{[8-amino-6-(3-methyl-1,2,3-triazol-4-yl)-2,7-naphthyridin-3-yl]amino}pyridin-4-yl)pentan-3-yl]-2-methylpropane-2-sulfinamideas a yellow solid (1.10 g, 29.2%) .

[0362] LCMS (ESI, m / z): 508.10 [M+H]+

[0363]

[0364]

[0365] Comparative Example 1

[0366] To a mixture of / V-[3-(2-{[8-amino-6-(3-methyl-1,2,3-triazol-4-yl)-2,7-naphthyridin-3-yl]amino}pyridin-4-yl)pentan-3-yl]-2-methylpropane-2-sulfinamide (300 mg, 0.59 mmol, 1.00 equiv) in DCM (3.0 mL) was added HCI in 1,4-dioxane (4.0 M, 1 mL) dropwise at 0 °C. The resulting mixture was stirred at room temperature for 2 h. The resulting mixture was concentrated under reduced pressure. The crude product was purified by reverse phase flash [with the following conditions (Column: XBridge Prep OBD C18 Column 30*150 mm, 5 pm; Mobile Phase A: Water(0.1% FA), Mobile Phase B: ACN; Flow rate: 100 mL / min; Gradient: 2% B to 16% B in 25 min; Wave Length: 254 nm 1220 nm; RT1(min): 22.22] to afford / \ / -6-[4-(3-aminopentan-3-yl)pyridin-2-yl]-3-(3-methyl-1,2,3-triazol-4-yl)-2,7-naphthyridine-1,6-diamine as a yellow green solid (57.3 mg, 24.0%).

[0367] LCMS (ESI, m / z): 404.10 [M+H]+

[0368] 1H NMR (400 MHz, DMSO-cfe) 69.86 (s, 1H), 9.31 (s, 1H), 8.33 (s, 1H), 8.27 - 8.29 (m, 4H), 7.40 -7.35 (m, 3H), 7.27 (s, 1 H), 6.98 (d, J = 5.2 Hz, 1 H), 4.39 (s, 3H), 1.85 - 1.66 (m, 4H), 0.72 (t, J= 7.3 Hz, 6H).008863987

[0369] 38

[0370] Comparative Example 2

[0371]

[0372] To a stirred mixture of 6,8-dichloro-2,7-naphthyridin-3-amine (200 mg, 0.93 mmol, 1.00

[0373] equiv) and bis[(4-methoxyphenyl) methyl] amine (4.81 g, 18.6 mmol, 20.0 equiv) in 1,4-dioxane

[0374] (20.0 mL) was added DIEA (361.9 mg, 2.80 mmol, 3.00 equiv). The resulting mixture was

[0375] stirred at 140 °C for overnight. The resulting mixture was concentrated under educed pressure.

[0376] The residue was purified by reversed-phase flash chromatography [with the following

[0377] conditions: column, C18 silica gel; mobile phase, MeCN in Water (10 mmol / L NH4HCO3), 10%

[0378] to 100% gradient in 30 min; detector, UV 254 nm] to afford 3-chloro- / V1 , A / 1-bis[(4-methoxyphenyl) methyl]-2,7-naphthyridine-1,6-diamine as a yellow solid (380 mg, 93.5%).

[0379] LCMS (ESI, m / z): 435.05 [M+H]+

[0380] Step (ii)

[0381]

[0382]

[0383] p 008863987

[0384] 39

[0385] To a mixture of 3-chloro- / V1 , / V1-bis[(4-methoxyphenyl) methyl]-2,7-naphthyridine-1,6-diamine (350 mg, 0.81 mmol, 1.00 equiv) and oxazolidinone (105 mg, 1.20 mmol, 1.50 equiv) in 1,4-dioxane (4 mL) were added Cs2CC>3(524 mg, 1.61 mmol, 2.00 equiv), XantPhos (93.1 mg, 0.16 mmol, 0.20 equiv) and Pd2(dba)3(92.2mg, 0.081 mmol, 0.10 equiv) at room temperature. The resulting mixture was stirred at 120 °C for 16 h under nitrogen atmosphere. The reaction was quenched with water at room temperature. The resulting mixture was extracted with EtOAc (3 x 15 mL), dried over anhydrous Na2SO4. After filtration, the filtrate was concentrated under reduced pressure. The residue was purified by reversed-phase flash chromatography [with the following conditions: column, C18 silica gel; mobile phase, MeCN in Water (0.1% FA), 0% to 100% gradient in 30 min; detector, UV 254 nm]. This resulted in 3-(6-amino-1-{bis[(4-methoxyphenyl) methyl] amino}-2,7-naphthyridin-3-yl)-1,3-oxazolidin-2-oneas a yellow solid (150 mg, 38.3%).

[0386] LCMS (ESI, m / z): 486.15 [M+H]+

[0387] Step (Hi)

[0388]

[0389]

[0390] Step 3 69.1%

[0391] To a mixture of 3-(6-amino-1-{bis[(4-methoxyphenyl) methyl] amino}-2,7-naphthyridin-3-yl)-1,3-oxazolidin-2-one (140 mg, 0.29 mmol, 1.00 equiv) and / V-[3-(2-bromopyridin-4-yl) pentan-3-yl]-2-methylpropane-2-sulfinamide (130 mg, 0.37 mmol, 1.30 equiv) in 1,4-dioxane (3 mL) were added CS2CO3 (187 mg, 0.58 mmol, 2.00 equiv), xphos (13.8 mg, 0.03 mmol, 0.20

[0392] equiv), and Pd2(dba)s (26.4 mg, 0.03 mmol, 0.10 equiv) at room temperature. The resulting mixture was stirred at 100 °C for 15 h under nitrogen atmosphere. The reaction was quenched with water at room temperature. The residue was purified by reversed-phase flash chromatography [with the following conditions: column, C18 silica gel; mobile

[0393] phase, MeCN in Water (0.1% FA), 0% to 100% gradient in 30 min; detector, UV 254 nm]. This resulted in / \ / -(3-{2-[(8-{bis[(4-methoxyphenyl) methyl] amino}-6-(2-oxo-1,3-oxazolidin-3-yl)-2,7-naphthyridin-3-yl) amino] pyridin-4-yl} pentan-3-yl)-2-methylpropane-2-sulfinamide as a yellow solid (180 mg, 69.1%).

[0394] LCMS (ESI, m / z): 752.35 [M+H]008863987

[0395] Step (iv)

[0396]

[0397] Into an 8 mL round-bottom flask were added / \ / -(3-{2-[(8-{bis[(4-methoxyphenyl) methyl] amino}-6-(2-oxo-1,3-oxazolidin-3-yl)-2,7-naphthyridin-3-yl) amino] pyridin-4-yl} pentan-3-yl)-2-methylpropane-2-sulfinamide (180 mg, 0.24 mmol, 1.00 equiv) and trifluoroacetic acid (3.0 mL) at room temperature. The resulting mixture was stirred at 80 °C for 2 h. The reaction was quenched with water at room temperature. The resulting mixture was concentrated under reduced pressure. The residue product was purified by reverse phase flash [with the following conditions (Column: Xselect CSH Prep C18, 30*150 mm 5 pm; Mobile Phase A: Water (0.1% FA), Mobile Phase B: ACN; Flow rate: 60 mL / min; Gradient: isocratic 2% to 17% B in 12 min; Wave Length: 254 nm / 220 nm; RT1(min): 8.32 min)] to afford 3-(1-amino-6-{[4-(3-aminopentan-3-yl) pyridin-2-yl] amino}-2,7-naphthyridin-3-yl)-1,3-oxazolidin-2-one as a yellow solid (57.0 mg, 58.4%).

[0398] LCMS (ESI, m / z): 408.25 [M+H]+

[0399] 1H NMR (400 MHz, DMSO-cfe) 69.79 (s, 1 H), 9.20 (s, 1 H), 8.37 - 8.14 (m, 4H), 7.33 (d, J= 2.4 Hz, 2H), 7.24 (s, 2H), 6.93 (dd, J= 5.5, 1.7 Hz, 1H), 4.42 (t, J= 8.0 Hz, 2H), 4.18 (t, J= 8.0 Hz, 2H), 1.93 - 1.81 (m, 4H), 0.79 (t, J = 7.2 Hz, 6H).

[0400] Biological Assay Procedure

[0401] Compounds 1 and 2 were tested for MASTL inhibiting activity and metabolic stability (liver microsome assay). The results were compared with Comparative Examples 1 and 2, which are described in WO 2024 / 003773.

[0402] Inhibition of MASTL

[0403] Inhibition of MASTL was determined using an ADP-Glo™ kinase assay (Promega) which measures kinase activity by quantifying the amount of ADP produced during a reaction. Full length human recombinant N-terminal GST tagged MASTL protein, the substrate ARPP19 protein, 5x kinase assay buffer III (200 mM Tris-HCI, pH7.4, 100 mM MgCh and 0.5 mg / ml BSA) and DTT were purchased from SignalChem, SinoBiological. ATP was purchased from

[0404] Promega.008863987

[0405] 41

[0406] Compounds (3 nM- 100 pM, 10 dose 3.16-fold dilutions) were incubated in 384-well plates with MASTL kinase in 1x kinase buffer III and 0.05 mM DTT at25°C for 10 min and 2.5 pL substrate was then added to initiate the reaction. The final reactions, containing 40 mM Tris HCI, pH 7.4, 20 mM MgCI2, 0.1 mg / ml BSA, 0.05 mM DTT, 1% DMSO, 2.2 pM ARPP19 protein, 10 pM ATP and 10 nM MASTL enzyme were incubated for 2 hours, when they were terminated by the addition of ADP Gio reagent, followed by kinase detection reagent (from ADP-Glo™ kinase assay kit (Promega)). After a further hour of incubation, luminescence was read on an Envision plate reader (Revvity). Low controls contain all assay components except MASTL enzyme and compound. High controls contain no compound.

[0407] Percent inhibition was determined using the formula: % Inhibition =100-100 x [(Readout - Low control) I (high control - Low control)]. IC50 values were calculated using a four-parameter logistic curve fit in XLfit.

[0408] The results are summarised in Table 1 below.

[0409] Table 1. Results of MASTL inhibition assay

[0410] <

[0411] <

[0412] <

[0413]

[0414] <

[0415] Metabolic Stability Assay

[0416] The metabolic stability of the enantiomerically enriched forms of Compounds 1 and 2 (Compounds 1A and 1B and Compounds 2A and 2B described above) were tested with human liver microsomes as follows.

[0417] The master solution was prepared according to Table 2.

[0418] Table 2. Preparation of Master Solution

[0419]

[0420] Note: The final concentrations of PBS and microsomes are 100 mM and 0.5 mg / mL in the reaction solution, respectively.

[0421] The cofactor solution 10 mM NADPH in ultra-pure H2O was prepared.

[0422] 80 nL of 1 mM test compounds or control compounds was added to the 72 pL of master solution.008863987

[0423] 42

[0424] The reaction was started with the addition of 8 pL of cofactor solution. The final concentrations in the reaction were 1 mM NADPH, 100mM PBS, 0.5mg / mL microsomes, 1 pM test compounds and 0.1% DMSO. The negative control samples were prepared by replacing NADPH solutions with 8 pL of ultra-pure H2O. The negative control is used to exclude the misleading factor that resulted from instability of chemical itself. This study was performed in duplicate. Negative controls were prepared in singlet. The mixture was pre-warmed at 37°C for 10 min.

[0425] Aliquots of 10 pL of 4 test compounds (40 pL in total) were taken from the reaction solution into one well of a new plate containing 120 pL of cold acetonitrile with internal standards (IS, 200 nM labetalol, 100 nM tolbutamide and 100 nM ketoprofen). The same operation was done after 15-, 30-, 45- and 60-minutes incubation at 37°C. Samples were centrifuged at 3, 220 g for 45 minutes. Aliquots of 40 pL of the supernatant were mixed with 40 pL of ultra-pure H2O and then used for LC-MS / MS analysis.

[0426] Data Analysis

[0427] All calculations were carried out using Microsoft Excel. Peak areas were determined from extracted ion chromatograms. The slope value was determined by linear regression of the natural logarithm of the remaining percentage of the parent drug vs. incubation time curve.

[0428] The in vitro half-life (in vitro ti / 2) was determined from the slope value:

[0429] Where: k is the rate constant (k= -slope value)

[0430] Conversion of the in vitro ti / 2 (min) into the in vitro intrinsic clearance (in vitro CLint , in pL / min / mg protein) was done using the following equation (mean of duplicate determinations):

[0431] volume of incubation (pL)

[0432] in vitro CLtat= k x - —;- — — — — - amount of proteins (mg)

[0433] The results are shown in Table 3:

[0434]

[0435] 008863987

[0436] 43

[0437] Computational Data

[0438] Binding affinity prediction model Boltz2 was used to predict MASTL binding affinity (Kj) and selectivity for compounds of the invention and comparative examples.

[0439] Boltz2 is a top-ranked Al program for predicting potency. Details on the model can be found in Passaro et al., ‘Boltz-2: Towards Accurate and Efficient Binding Affinity Prediction’ (2025), which is hereby incorporated by reference.

[0440] For each compound, predictions were run five times and averages and standard deviations were calculated. For compounds having stereoisomers the predictions are based on a racemic mixture. The results are shown in Table 4. Compounds having a Kj standard deviation of greater than 2 nm are excluded from the data provided.

[0441] Selectivity score is a percentage calculated based on the number of predicted off-target hits out of 47, using the formula:

[0442] ((47-n) / 47)*100

[0443] where n is the number of off target hits.

[0444] Table 4. Predicted Ki and Selectivity Score for Example Compounds

[0445]

[0446] 008863987

[0447] 44

[0448] References

[0449] A number of publications are cited above in order to more fully describe and disclose the invention and the state of the art to which the invention pertains. Full citations for these references are provided below. The entirety of each of these references is incorporated herein.

[0450] WO 2024 / 003773

[0451] Bharate, S.S., Drug Discovery Today (2020) “Recent developments in pharmaceutical salts: FDA approvals from 2015 to 2019’’

[0452] doi: https: / / doi.org / 10.1016 / j.drudis.2020.11.016

[0453] For standard molecular biology techniques, see Sambrook, J., Russel, D.W. Molecular Cloning, A Laboratory Manual. 3 ed. 2001, Cold Spring Harbor, New York: Cold Spring Harbor Laboratory Press

[0454] Passaro et al., ‘Boltz-2: Towards Accurate and Efficient Binding Affinity Prediction’ (2025)

Claims

00886398745Claims:

1. A compound of formula (I):Formula (I)or a pharmaceutically acceptable salt, solvate, protected form or prodrug form thereof, wherein:R1is an optionally substituted heteroaryl;X, Y and Z are independently selected from N, CH, and CR5wherein at least one of X, Y or Z is N and wherein R5is selected from Ci-Ce alkyl and Ci-Ce alkoxy;V and Q are selected from N, CH, and CR5, wherein at least one of V or Q is N and wherein R5is selected from Ci-Ce alkyl and Ci-Ce alkoxy;and wherein:R2is hydrogen or C1-C4 alkyl; R3is fluoro-substituted C3-C10 cycloalkyl, or R3together with the carbon atom to which it is attached and a carbon atom of the pyridine ring forms a fluoro-substituted 5- to 8-membered ring that is fused to the pyridine ring; or R2and R3together with the carbon atom to which they are attached form a fluoro- substituted C3-C10 cycloalkyl group; and R4is hydrogen;orR2is hydrogen or C1-C4 alkyl; and R3and R4together with the carbon and nitrogen atoms to which they are attached form a fluoro-substituted 5- to 10-membered heterocycloalkyl;orR2is hydrogen or C1-C4 alkyl; R3is C1-C4 alkyl; and R4together with the nitrogen atom to which it is attached and a carbon atom of the pyridine ring forms a fluoro- substituted 5- to 8-membered ring that is fused to the pyridine ring.

2. The compound according to claim 1, wherein:R1is an optionally substituted heteroaryl;00886398746X, Y and Z are independently selected from N, CH, and CR5wherein at least one of X, Y or Z is N and wherein R5is selected from Ci-Ce alkyl and Ci-Ce alkoxy;V and Q are selected from N, CH, and CR5, wherein at least one of V or Q is N and wherein R5is selected from Ci-Ce alkyl and Ci-Ce alkoxy;and wherein:R2is hydrogen or C1-C4 alkyl; R3is fluoro-substituted C3-C10 cycloalkyl, or R3together with the carbon atom to which it is attached and a carbon atom of the pyridine ring forms a fluoro-substituted 5- to 8-membered ring that is fused to the pyridine ring; or R2and R3together with the carbon atom to which they are attached form a fluoro- substituted C3-C10 cycloalkyl group; and R4is hydrogen.

3. The compound according to claim 2, wherein R2is hydrogen or methyl and R3is fluoro-substituted C3-C10 cycloalkyl, or R2and R3together form a fluoro-substituted C3-C10 cycloalkyl group4. The compound according to claim 3, wherein R2is hydrogen or methyl and R3is fluoro-substituted C3-C10 cycloalkyl such as C3-C8 or C3-C6 cycloalkyl.

5. The compound according to claim 4, wherein R3is a fluoro-substituted bicyclic C5-C10 cycloalkyl group preferably fluoro-substituted C5 or Ce bicyclic cycloalkyl group.

6. The compound according to claim 5, wherein R3is selected from the group consisting of:wherein marks the point of attachment to the rest of the compound.

7. The compound according to any of the preceding claims, wherein X is nitrogen.

8. The compound according to any of the preceding claims wherein Q is nitrogen.

9. The compound according to any of the preceding claims wherein X is nitrogen, Y and Z are both CH, Q is nitrogen and V is CH.

10. The compound according to any of claims 1 to 8 wherein X is nitrogen, Y is nitrogen and Z is CH.00886398711. The compound according to any of the preceding claims, wherein R1comprises an optionally substituted azole ring.

12. The compound according to any of the preceding claims, wherein R1comprises a substituted imidazole or 1,2,3-triazole ring.

13. The compound according to any of the preceding claims, wherein R1is substituted with C1-C4 alkyl, such as methyl.

14. The compound according to claim 11 or 12, wherein the azole ring of R1is fused with a second ring to form a bicyclic ring system.

15. The compound according to claim 14, wherein R1comprises an imidazole ring fused with a second ring, wherein the second ring is an optionally fluoro-substituted 5-membered ring.

16. The compound according to claim 11, wherein R1is selected from the group consisting of:

17. The compound according to claim 16, wherein R1is selected from the group consisting of:

18. The compound according to any of the preceding claims, wherein the compound is provided as a racemic mixture.0088639874819. The compound according to any of claims 1 to 17, wherein the compound is provided in a form that is at least 60% optically enriched with either the R or S enantiomer, such as at least 70% optically enriched, such as at least 80% optically enriched, such as at least 90% optically enriched.

20. The compound according to claim 1, 18 or 19, wherein the compound is selected from:008863987008863987008863987008863987520088639870088639870088639875521. The compound according to claim 1, 18, 19 or 20, wherein the compound is selected from:0088639870088639870088639875822. The compound according to claim 1, 18, 19 or 20 wherein the compound is selected from:

23. A pharmaceutical composition comprising a compound according to any of the preceding claims and a biologically acceptable excipient.

24. The compound according to any of claims 1 to 22 or a pharmaceutical composition according to claim 23 for use in a method of treatment or prophylaxis.

25. The compound according to any of claims 1 to 22 or a pharmaceutical composition according to claim 23 for use in a method of treatment or prophylaxis comprising the inhibition of MASTL.0088639875926. The compound according to any of claims 1 to 22 or a pharmaceutical composition according to claim 23 for use in a method of treating cancer such as breast cancer, colon cancer, colorectal cancer, head and neck squamous cell carcinoma, non-small cell lung cancer (NSCLC) gastric cancer, pancreatic cancer, prostate cancer or thyroid cancer.

27. A method of treatment comprising the step of administering a compound of formula (I) according to any of claims 1 to 22, or a pharmaceutical composition comprising a compound of formula (I) according to claim 23, to a subject in need thereof.

Images