Salt and solid forms of a hpk1 inhibitor

Abstract

Various salt forms and free base of Compound (I) represented by the following formula are disclosed.

Description

[0001] SALT AND SOLID FORMS OF A HPK1 INHIBITOR

[0002] RELATED APPLICATIONS

[0003] This application claims priority to International Application No. PCT / CN2023 / 139882, filed on December 19, 2023. The entire contents of the foregoing application are expressly incorporated herein by reference.

[0004] BACKGROUND

[0005] Hematopoietic Progenitor Kinase 1 (HPK1), also known as Mitogen-Activated Protein Kinase Kinase Kinase Kinase 1 (MAP4K1), is a protein kinase that acts upstream of the classic 3 -tiered MAPK pathway that includes MAP3K (MAP Kinase Kinase Kinase), which activates MAP2K (MAP Kinase Kinase), which in turn activates the dual Thr and Tyr MAPK family member JNK (c-Jun N-terminal kinase). Originally cloned in hematopoietic progenitor cells, HPK1 / MAP4K1 is predominantly expressed in lymphoid organs / tissues, including the bone marrow, fetal liver, lymph node, placenta, spleen, and thymus (Hu et al., Gene & Dev. 10(18): 2251-2264, 1996; Kiefer et al., The EMB O J. 15(24): 7013-7025, 1996). At the cellular level, HPK1 is expressed in all cell types in the hematopoietic compartment, including hematopoietic progenitor cells, T cells, B cells, macrophages, dendritic cells, neutrophils, and mast cells (Hu, supra; Kiefer, supra).

[0006] HPK1 / MAP4K1 is one of the six MAP4Ks that include HPK1 (MAP4K1), GCK (MAP4K2), GLK (MAP4K3), HGK / NIK (MAP4K4), KHS / GCKR (MAP4K5), and MINK (MAP4K6). Together, these MAP4Ks are members of about 26 mammalian Ste20-like serine / threonine kinases identified so far, which are homologs of the yeast sterile20 protein (Ste20p), a putative MAP4K that activates a MAP3K in the yeast pheromone signaling pathway. These mammalian Ste20-like kinases are divided into two subfamilies based on the domain structures: the p21 -activated kinases (PAKs) and the germinal center kinases (GCKs). Among the GCK subfamily, several of them can activate the MAP3K kinase cascade, leading to JNK activation.

[0007] The MAP4Ks are highly similar structurally, with an N-terminal kinase domain (KD), followed by 2-4 proline-rich motifs, and a C-terminal citron-homology domain (CNH).

[0008] The ATP -binding site of the kinase domain of HPK1 includes Lys-46. Mutation of this residue to Met (HPK1-M46) abolishes catalytic activation of HPK1 (Hu, supra). There are multiple conserved Ser / Thr phosphorylation sites within the kinase domain of HPK1, and a conserved Tyr phosphorylation site between its first two proline-rich motifs. Phosphorylation of Tyr379 (in mouse, or Tyr381 in human) by LCK / ZAP70 appears to be required for HPK1 activation, since the deficiency of LCK or ZAP70 abolishes Tyr-379 phosphorylation and kinase activity of HPK1 in Jurkat T cells upon anti-CD3 stimulation (Ling et al., JBC 276(22): 18908-18914, 2001; Liou et al., Immunity 12(4):399-408, 2000; Sauer et al., JBC 276(48):45207, 45216, 2001). On the other hand, Thr-355 autophosphorylation regulates ubiquitination and degradation of HPK1. Thr-355 is a PP4- targeted dephosphorylation site; this dephosphorylation prevents CUL7 / Fbxw8-mediated ubiquitination and proteasomal degradation of activated HPK1 (Wang et al., Cancer Res. 69(3): 1063-1070, 2009), thus HPK1 is also stabilized and activated by protein phosphatase 4 (PP4) (Zhou et al., JBC 279(47):49551-49561, 2004).

[0009] The Tyr phosphorylation site is also adjacent to a caspase cleavage site (DDVD). It has been shown that the full-length HPK1 can be cleaved by caspase-3 at this site in apoptotic cells, resulting in an enhanced catalytic activity of the N-terminal HPK1 fragment (Chen et al., Oncogene 18:7370-7377, 1999).

[0010] The four Pro-rich motifs of HPK1 mediate the interaction of HPK1 with many SH3 domain-containing proteins (Boomer & Tan, JCB 95(1): 34-44, 2005).

[0011] The CNH domain of HPK1 may be involved in HPKl-mediated lymphocyte adhesion because the citron homology domain in another Ste20-like kinase TNIK binds to Rap2 and regulates actin cytoskeleton (Taira et al., JBC 279(47):49488-49496, 2004).

[0012] The MAP4Ks play important roles in the immune system, particularly in lymphocytes, through regulating cellular signaling, immune cell activation, cell transformation, and cell migration. HPK1 knock-out (KO) mice show enhanced T-cell activation, increased cytokine production, and increased antibody production after KLH immunization. HPK1 KO mice are also more susceptible to EAE induction. HPK1 KO T cells and B cells show enhanced cell activation and antigen receptor signaling. HPK1 KO dendritic cells show higher levels of co-stimulating molecules and pro-inflammatory cytokines (Alzabin et al., J. Immunol. 182(10):6187-6194, 2009; Shui et al., Nat. Immunol. 8(1):84-91, 2007).

[0013] Overexpression in cell lines (e.g., HEK293 and COS-1 cells, and hematopoietic Jurkat T cells and leukemia HL-60 cells) demonstrated that HPK1 can activate the MAPK JNK (but not p38 or ERK MAP kinases) through multiple MAP3Ks (including TAK1, MEKK1, and MLK3), which all activate MAP2Ks MKK4 and MKK7, which in turn activate JNK.

[0014] Interestingly, the regulatory functions of MAP4Ks in immune cells appear to be largely mediated by JNK -independent mechanisms. It has been demonstrated that HPK1 kinase activation is required for IKK-NF-KB activation, and it is believed that HPK1 does so via regulating CARMA1. CARMA1 is an adaptor protein in the so-called CBM (CARMA1 / BCL10 / MALT1) complex that facilitates IKKP activation in Jurkat T cells upon anti-CD3 stimulation. Activated IKK cleaves IKB and releases the associated NF-KB nuclear transcriptional factor. In particular, HPK1 is inducibly associated with CARMA1, and directly phosphorylates CARMA1 at Ser-551 that is required for NF-KB activation (Brenner et al., PNAS USA, 196(34): 14508-14513, 2009).

[0015] In T cells, upon TCR stimulation, lymphocyte protein tyrosine kinase (Lek) phosphorylates the immunoreceptor tyrosine-based activation motifs (IT AMs) on the cytosolic side of the TCR / CD3 complex. Zap-70 is then recruited to the TCR / CD3 complex, where it becomes phosphorylated and activated. Activated ZAP-70 phosphorylates an adaptor protein called SLP-76, which translocates to the plasma membrane, and promotes the formation of a multi-protein signalosome complex by binding to a number of proteins including HPK1. These proteins collectively transmit TCR signaling to different effector molecules, leading to activation, survival, and proliferation of T-Lymphocytes.

[0016] During this process, HPK1 directly binds to the SH2 domain of SLP-76, and primarily serves as a negative regulator of TCR signaling. For example, TCR signaling is enhanced in HPK1 KO primary T cells, in that they show hyperproliferation and IL-2 production upon TCR ligation in vitro (Shui, supra). It is believed that HPK1 can down-regulate TCR signaling through a negative feedback mechanism, by phosphorylating the SLP-76 adaptor protein at Ser-376. Upon Ser-376 phosphorylation by HPK1, SLP-76 binds to 14-3-3 through the phosphorylated Ser-376 residue, leading to ubiquitination at Lys-30 (K30) residue of SLP-76 which is subsequently targeted for proteasome degradation. HPK1 also down-regulates TCR signaling by a similar mechanism in other adaptor proteins, including GADS (e.g., by phosphorylating Thr-254 of GADS and promote 14-3-3 interaction).

[0017] Thus it appears that HPK1 plays dual and opposite roles in JNK activation and TCR signaling. While HPK1 has been demonstrated to directly activate the JNK pathway in different overexpression systems through the MAP3K-MAP2K-MAPK pathway, HPK1- mediated inhibition of SLP-76 activation would also lead to the inhibition of JNK activity in TCR signaling. This is consistent with the observation that HPK1 knockout primary T cells show unaffected JNK activity (Shui, supra). Similarly, HPK1 seems to regulate IKK activation in two different and contrasting mechanisms - on the one hand, HPK1 activates IKK by directly phosphorylating CARMA1; on the other hand, HPK1 also negatively regulates IKK activation by inhibiting SLP-76 activation. This seemingly contrasting dual roles played by HPK1 is best understood that HPK1 facilitates JNK and IKK activation in the initial phase of TCR signaling but plays a critical role in dampening TCR signaling in the late phase.

[0018] HPK1 also plays a similar negative regulatory role in BCR-induced cell activation and proliferation in B cells. B cells use SLP-76-like adaptor protein called BLNK to transduce BCR signaling, including JNK and IKK activation. In B cells, Tyr kinases Syk and Lyn promote Tyr phosphorylation and activation of HPK1, and the resulting pY379 of HPK1 mediates HPK1-BLNK binding. The negative feedback by HPK1 of BLNK is through Thr- 152 of BLNK. pT152 binding by 14-3-3 leads to BLNK ubiquitination at multiple Lys residues, and subsequent proteasome degradation of BLNK (thus dampening BCR signaling).

[0019] Interestingly, HPK1 appears to be a positive regulator of suppressive functions of regulatory T cells (Treg) (Sawasdikosol et al., J Immunol. 188(supp. 1): 163, 2012). HPK1 deficient mouse Foxp3+ Tregs were defective in suppressing TCR-induced effector T cell proliferation, and paradoxically gained the ability to produce IL-2 following TCR engagement (Sawasdikosol, supra). Thus, HPK1 is an important regulator of Treg functions and peripheral self- tolerance.

[0020] HPK1 is also involved in PGE2 -mediated inhibition of CD4+ T cell activation (Ikegami et al., J Immunol. 166(7):4689-4696, 2001). US2007 / 0087988 shows that HPK1 kinase activity was increased by exposure to physiological concentrations of PGE2 in CD4+ T cells through PGE2-induced PKA activation. The proliferation of HPK1 deficient T cells was resistant to the suppressive effects of PGE2 (US 2007 / 0087988). Therefore, PGE2- mediated activation of HPK1 may represent a novel regulatory pathway of modulating immune response.

[0021] Other than TCR and BCR, HPK1 also transduces signals downstream of the TGF-R (transforming growth factor receptor) (Wang et al., JBC 272(36):22771-22775, 1997), or Gs- coupled PGE2 receptors (EP2 and EP4) (Ikegami et al., J Immunol. 166(7):4689-4696, 2001). HPK1 negatively regulates immune cell adhesion. In T cells, TCR activation also induces integrin activation, resulting in T-cell adhesion and immunological synapse formation. This is achieved by SLP-76 binding of the degranulation-promoting adaptor protein (ADAP), which is required for TCR-induced integrin activation (Wang et a!.. J. Exp. Med. 200(8): 1063-1074, 2004), though its constitutively associated SKAP55 protein that targets the activated small GTPase Rapl to the plasma membrane, leading to integrin activation (Kliche et al., MCB 26(19):7130-7144, 2006). In other words, the SLP- 76 / ADAP / SKAP55 ternary complex relays the TCR signaling to adhesion molecules of the integrin family, thereby promoting T-cell adhesion. HPK1 negatively regulates this pathway, not only through down-regulating SLP-76 (supra) but also through competing with ADAP for the same SH2 binding site on SLP-76, which in turn dampens the activity of ADAP downstream effector Rapl (Patzak et al., Eur. J. Immunol. 40(11):3220-3225, 2010).

[0022] HPK1 similarly negatively regulates integrin activation and cell adhesion in B cells. There, HPK1 is associated with a SKAP55 homologue called SKAP-HOM (Konigsberger et al., PloS One 5(9). pii: el2468, 2010), which is required for B-cell adhesion (Togni et al., MCB 25(18):8052-8063, 2005). HPK1 is believed to induce a negative phosphorylation site on SKAP-HOM, which in turn suppresses Rapl activation.

[0023] In neutrophils, however, HPK1 positively regulates their adhesion. Neutrophil trafficking, including slow rolling, tight binding, cell spreading, and diapedesis, is controlled by the outside-in signaling of P2-Integrin activation, which induces the interaction between actin and HIP-55 (HPK1 -interacting protein of 55 kDa). This reinforces the high-affinity conformation of p2-integrin, contributing to neutrophil adhesion (Hepper et al., J. Immunol. 188(9):4590-4601, 2012; Schymeinsky et al., Blood 114(19):4209-4220, 2009). HPK1 colocalizes with HIP-55 and actin at the lamellipodium of neutrophils upon p2-integrin- mediated adhesion (Jakob et al., Blood 121(20):4184-4194, 2013). CXCL1 -mediated neutrophil adhesion is abolished by either HPK1 deficiency or HIP-55 deficiency in vitro and in vivo (Jakob, supra; Schymeinsky, supra).

[0024] Consistent with its role in down-regulating TCR and BCR function, HPK1 negatively regulates adaptive immune responses, and loss of HPK1 -mediated regulation of T-cell activation and immune responses may be a crucial mechanism for autoimmune pathogenesis. In HPK1 KO mice, although the development of T and B cells appeared unaffected (Shui, supra), T cells from these animals showed dramatically increased activation of TCR proximal signaling and downstream ERK, leading to hyperproliferation of these cells in vitro upon anti-CD3 stimulation (Shui, supra). T cells from immunized HPK1 -deficient mice are hyper- responsive upon antigenic specific stimulation, and produce significantly higher levels of inflammatory cytokines such as IL-2, IFN-y, and IL-4. Such mice also produce much higher levels of IgM and IgG isoforms, suggesting enhanced functioning of HPK1 knockout B cells (Shui, supra).

[0025] HPK1 also negatively controls autoimmunity in mice, since HPK1 KO mice are more sensitive to the induction of experimental autoimmune encephalomyelitis (EAE) (Shui, supra). HPK1 attenuation also contributes to the abnormal T- and B-cell activation and to autoimmunity in human patients. HPK1 is down-regulated in peripheral blood mononuclear cells of psoriatic arthritis patients, or T cells of systemic lupus erythematosus (SLE) patients.

[0026] The physiological function of HPK1 is not limited to lymphocytes, for HPK1 also negatively regulates dendritic cell (DC) maturation and activation through an unknown mechanism (Alzabin, supra). In the HPK1 KO mice, the bone marrow-derived dendritic cells (BMDCs) display enhanced levels of co-stimulatory molecules CD80 / CD86, and increased production of proinflammatory cytokines (Alzabin, supra). Consequently, antigen presentation activity of dendritic cells is more efficient in HPK1 KO mice (Alzabin, supra). More importantly, tumor eradication by HPK1 KO BMDC-mediated CTL response is more effective than that by wild-type BMDCs (Alzabin, supra). Furthermore, HPK1 can also control antitumor immunity via T- and B- lymphocyte-dependent mechanisms. It has been shown that adoptive transfer of HPK1 deficient T cells was more effective in controlling tumor growth and metastasis than wild-type T cells (Alzabin et al., Cancer Immunol Immunother 59(3):419-429, 2010). Similarly, BMDCs from HPK1 knockout mice were more efficient to mount a T cell response to eradicate Lewis lung carcinoma as compared to wild-type BMDCs (Alzabin et al., J Immunol. 182(10):6187-6194, 2009).

[0027] Thus, there is a need for HPK1 inhibitory compounds for treating diseases or disorders through modulating HPK1 activity.

[0028] International Publication No. W02022 / 002237, the entire teachings of which are incorporated herein by references discloses highly potent, highly selective inhibitors of HPK1. The structure of one of the inhibitors disclosed in International Publication No. W02022 / 002237, referred to herein as “Compound (I)” is shown below: Compound (I)

[0029] Compound (I) is a potent, selective, small molecule inhibitor of HPK1. Its potency is demonstrated in vitro in cellular (half-maximal inhibitory concentration, [IC50] = < 1 nM) for HPK1 enzymes. Further, Compound (I) exhibits excellent efficacy in inducing IL-2 production in Human Pan T Cells.

[0030] The successful development of pharmaceutically active agents, such as Compound (I), typically requires the identification of a solid form with properties that enable ready isolation and purification following synthesis, that are amendable to large scale manufacture, that can be stored for extended periods of time with minimal absorption of water, decomposition or transformation into other solid forms, that are suitable for formulation and that can be readily absorbed following administration to the subject (e.g., are soluble in water and in gastric fluids).

[0031] SUMMARY

[0032] Described herein are polymorph forms of the free base of Compound (I) and salt / hydrate forms of Compound (I).

[0033] It has now been found that Compound (I) can form a salt upon the addition of a variety of acids. In salt screening, total 5 salts forms were identified. Therein, phosphate hydrate Form A and maleate Form A demonstrated favorable solid state properties and were selected as lead salts for solubility and stability studies. Phosphate hydrate Form A had higher solubility in water than maleate Form A. Phosphate hydrate Form A and maleate Form A showed similar solubility in SGF (-300 pg / mL) and in FeSSIF (-100 pg / mL). In FaSSIF, the solubility of maleate Form A was -5.4 folds that of phosphate hydrate Form A. Phosphate hydrate Form A and maleate Form A were physically stable under both high temperature (60 °C) and high humidity conditions (25 °C / 92.5% RH). However, phosphate Form A could be easily prepared in a variety of solvents, thus providing synthetic advantages towards a manufacturing process. Additionally, the free base of Compound (I) can be crystallized under well-defined conditions to afford a single crystalline form. Form A was less soluble than the salt forms in SGF, FeSSIF, and water, but showed comparable solubility in FaSSIF to the salt forms of Compound (I). Form A also demonstrated proved stable at 60 °C and 25 °C / 92.5% RH after 11 days.

[0034] The present disclosure provides a method of inhibiting HPK1 activity in a subject in need thereof, the method comprising administering to the subject a therapeutically effective amount of the salt or the free base (including both amorphous form and crystalline form) of Compound (I) disclosed herein, or the corresponding pharmaceutical composition thereof.

[0035] The present disclosure further provides a method of treating a subject with a disease or condition as described herein, such as cancer (such as breast cancer, colorectal cancer, lung cancer, ovarian cancer, and pancreatic cancer), the method comprising administering to the subject a therapeutically effective amount of the salt or the free base (including both amorphous form and crystalline forms) of Compound (I) disclosed herein, or the corresponding pharmaceutical composition thereof.

[0036] The present disclosure also provides a use of the salt or freebase of Compound (I) of the disclosure or a pharmaceutical composition thereof comprising the same for the treatment of any of the disease recited in the previous paragraph. In one embodiment, provided is the salt or freebase of the disclosure or a pharmaceutical composition thereof comprising the same for use in any of the method of the disclosure described herein. In another embodiment, provided is use of the salt of the disclosure or freebase or a pharmaceutical composition thereof comprising the same for the manufacture of a medicament for any of the method of the disclosure described.

[0037] BRIEF DESCRIPTION OF THE DRAWINGS

[0038] Figure 1 A shows the X-ray Powder Diffraction (XRPD) pattern of crystalline Form A of the phosphate hydrate salt of Compound (I).

[0039] Figure IB shows the Thermogravimetric Analysis (TGA) and Differential Scanning Calorimetry Analysis (DSC) thermograms of crystalline Form A of the phosphate hydrate salt of Compound (I).

[0040] Figure 2 A shows the X-ray Powder Diffraction (XRPD) pattern of crystalline Form A of the maleate salt of Compound (I). Figure 2B shows the Thermogravimetric Analysis (TGA) and Differential Scanning Calorimetry Analysis (DSC) thermograms of crystalline Form A of the maleate salt of Compound (I).

[0041] Figure 3 A shows the X-ray Powder Diffraction (XRPD) pattern of crystalline Form A of the hydrochloride salt of Compound (I).

[0042] Figure 3B shows the Thermogravimetric Analysis (TGA) and Differential Scanning Calorimetry Analysis (DSC) thermograms of crystalline Form A of the hydrochloride salt of Compound (I).

[0043] Figure 4A shows the X-ray Powder Diffraction (XRPD) pattern of crystalline Form B of the hydrochloride salt of Compound (I).

[0044] Figure 4B shows the Thermogravimetric Analysis (TGA) and Differential Scanning Calorimetry Analysis (DSC) thermograms of crystalline Form B of the hydrochloride salt Compound (I).

[0045] Figure 5 A shows the X-ray Powder Diffraction (XRPD) pattern of crystalline Form A of the sulfate salt of Compound (I).

[0046] Figure 5B shows the Thermogravimetric Analysis (TGA) and Differential Scanning Calorimetry Analysis (DSC) thermograms of crystalline Form A of the sulfate salt of Compound (I).

[0047] Figure 6A shows the X-ray Powder Diffraction (XRPD) pattern of crystalline Form A of the calcium salt of Compound (I).

[0048] Figure 6B shows the Thermogravimetric Analysis (TGA) and Differential Scanning Calorimetry Analysis (DSC) thermograms of crystalline Form A of the calcium salt of Compound (I).

[0049] Figure 7A shows the X-ray Powder Diffraction (XRPD) pattern of crystalline Form A of the free base of Compound (I).

[0050] Figure 7B shows the Thermogravimetric Analysis (TGA) and Differential Scanning Calorimetry Analysis (DSC) thermograms of crystalline Form A of the free base of Compound (I).

[0051] Figure 8 shows the single crystal structure of the crystalline Form A of the phosphate hydrate salt of Compound (I).

[0052] Figure 9 shows the X-ray Powder Diffraction (XRPD) pattern of the amorphous form of the free base of Compound (I). DETAILED DESCRIPTION

[0053] The present disclosure is directed to: i) novel crystalline forms of the of Compound (I), including unsolvated forms, solvated forms, amorphous forms, and crystalline forms ii) novel solid forms of pharmaceutically acceptable salts of Compound (I), including unsolvated forms, solvated forms, amorphous forms, and crystalline forms (hereinafter collectively referred to as “salt forms”); and iii) methods of use and preparation of the crystalline forms and salt forms of Compound (I).

[0054] In one aspect, the present disclosure provides crystalline forms of Compound (I). The form of Compound (I) can be crystalline and can exist as one or more polymorph forms. These polymorphic forms can be solvated or unsolvated forms. These polymorphic or crystalline forms differ with respect to their X-ray powder diffractions (XRPD) patterns, spectroscopic, physicochemical, and pharmacokinetic properties, as well as their thermodynamic stability.

[0055] In another aspect, the present disclosure provides crystalline forms of Compound (I) as salt forms. Many different salt forms can exist depending on the salt used. Each salt forms can exist as one or more polymorph salt forms. The polymorph salt form can be solvated or unsolvated. These polymorphic or crystalline salt forms differ with respect to their X-ray powder diffractions (XRPD) patterns, spectroscopic, physicochemical, and pharmacokinetic properties, as well as their thermodynamic stability.

[0056] It is desirable to have access to different polymorphic forms of Compound (I) for several reasons. Distinct polymorph forms may exhibit different physical properties such as melting point, hygroscopicity, solubility, flow properties or thermodynamic stability, and therefore, distinct polymorph forms allow the choice of the most suitable form for a given use or aspect, for example, in distinct administration forms such as capsules, or in the manufacture of a drug form having optimum pharmacokinetic properties.

[0057] The present invention provides a polymorphic forms of (S)-2-(l-(5-chloro-2-((6- fluoro-2-methyl-l,2,3,4-tetrahydroisoquinolin-7-yl)amino)pyrimidin-4-yl)-3-methylindolin- 3-yl)acetic acid, also referred to as “Compound (I)”, originally described in International Publication No. W02022 / 002237, Synthetic Example 5 & 6. The contents of W02022 / 002237 is incorporated herewith by reference, in particular the disclosure related to the synthesis, Synthetic Examples 5 and 6. As shown in Figure 9, Compound (I) was obtained as an amorphous solid. It has now been surprisingly found that under certain conditions new solid forms of Compound (I), can be provided which are described hereinafter as Form A, HC1 salt Form A, HC1 salt Form B, maleate salt Form A, phosphate hydrate salt Form A, sulfate Form A, and calcium Form A, which have advantageous utilities and properties. In particular, phosphate hydrate Form A of the Compound (I) shows excellent stability properties when subject to stress conditions. Phosphate hydrate Form A further demonstrated high crystallinity and high physical and chemical stability. This high degree of stability of phosphate hydrate Form A provides advantageous properties and benefits in terms of its suitability for use in a pharmaceutical composition, for example, in terms of its shelf-life and ease of manufacture.

[0058] Salt Forms o f Compound (I)

[0059] Phosphate Hydrate Salt and Crystalline Forms

[0060] In one embodiment, the present disclosure provides a phosphate hydrate salt of Compound (I). In one specific embodiment, the molar ratio of Compound (I) to phosphoric acid is 1 to 1.

[0061] In another embodiment, the present disclosure provides a phosphate hydrate salt of Compound (I), wherein the molar ratio of Compound (I) to phosphoric acid to water is 1 to 1 to 0.5.

[0062] In another embodiment, the present disclosure provides a phosphate hydrate salt of Compound (I), wherein the phosphate hydrate salt is crystalline.

[0063] In another embodiment, the present disclosure provides a phosphate hydrate salt of Compound (I), wherein the phosphate hydrate salt is crystalline Form A characterized by an X-ray powder diffraction pattern which comprises peaks at 13.8°, 14.8°, 20.6°, and 24.1° ± 0.2 in 20.

[0064] In another embodiment, the present disclosure provides a phosphate hydrate salt of Compound (I), wherein the phosphate hydrate salt is crystalline Form A characterized by an X-ray powder diffraction pattern which comprises at least five, six, seven or eight peaks chosen from 8.1°, 12.6°, 13.8°, 14.8°, 17.1°, 20.6°, 24.1°, and 24.5° ± 0.2 in 20.

[0065] In another embodiment, the present disclosure provides a phosphate hydrate salt of Compound (I), wherein the phosphate hydrate salt is crystalline Form A characterized by an X-ray powder diffraction pattern which comprises peaks at 6.2°, 8.1°, 12.6°, 13.8°, 14.8°, 17.1°, 18.7°, 20.6°, 23.4°, 24.1°, 24.5°, and 29.6° ± 0.2 in 20. In another embodiment, the present disclosure provides a phosphate hydrate salt of Compound (I), wherein the phosphate hydrate salt is crystalline Form A characterized by a differential scanning calorimeter (DSC) peak phase transition temperature of 228.8 ± 2 °C.

[0066] Representative XRPD peaks are tabulated Table 1. The XRPD patterns and peaks are shown in Figure 1 A. The Thermogravimetric Analysis (TGA) and Differential Scanning

[0067] Calorimetry Analysis (DSC) thermograms are shown in Figure IB.

[0068] Table 1. Representative XRPD peaks of crystalline Form A of phosphate hydrate salt of Compound (I)

[0069] Maleate Salt and Its Crystalline Form

[0070] In one embodiment, the present disclosure provides a maleate salt of Compound (I), the molar ratio of Compound (I) to maleic acid is 1 to 1.

[0071] In another embodiment, the present disclosure provides crystalline Form A of maleate salt of Compound (I), wherein the molar ratio between Compound (I) and maleic acid is 1 : 1, which is characterized by an X-ray powder diffraction pattern which comprises peaks at 8.5°, 11.3°, 17.2° and 17.8° ± 0.2 in 26. In another embodiment, the present disclosure provides a crystalline Form A of maleate salt of Compound (I), wherein the molar ratio between Compound (I) and maleic acid is 1 : 1, which is characterized by an X-ray powder diffraction pattern which comprises at least five, six, or seven peaks chosen from 8.5°, 8.8°, 11.3°, 17.2°, 17.8°, 22.0°, and 27.2° ± 0.2 in 26.

[0072] In another embodiment, the present disclosure provides a crystalline Form A of maleate salt of Compound (I), wherein the molar ratio between Compound (I) and maleic acid is 1 : 1, which is characterized by an X-ray powder diffraction pattern which comprises peaks at 8.5°, 8.8°, 11.3°, 15.7°, 17.2°, 17.8°, 22.0°, 23.0°, 26.0°, and 27.2° ± 0.2 in 29.

[0073] In another embodiment, the present disclosure provides a crystalline Form A of maleate salt of Compound (I), wherein the molar ratio between Compound (I) and maleic acid is 1 : 1, which is characterized by a differential scanning calorimeter (DSC) peak phase transition temperature of 156.9 ± 2 °C.

[0074] In one embodiment, crystalline Form A of maleate salt of Compound (I) is an anhydrate.

[0075] Representative XRPD peaks are tabulated in Table 2. The XRPD patterns and peaks are shown in Figure 2A. The Thermogravimetric Analysis (TGA) and Differential Scanning Calorimetry Analysis (DSC) thermograms are shown in Figure 2B.

[0076] Table 2. Representative XRPD peaks of crystalline Form A of Maleate Salt of Compound (I)

[0077] Hydrochloride Salt and Its Crystalline Forms

[0078] In one embodiment, the present disclosure provides a hydrochloride salt of Compound (I), the molar ratio of Compound (I) to hydrochloric acid is 1 to 1.

[0079] Hydrochloride Salt Form A

[0080] In another embodiment, the present disclosure provides crystalline Form A of hydrochloride salt of Compound (I), wherein the molar ratio between Compound (I) and hydrochloric acid is 1 : 1, which is characterized by an X-ray powder diffraction pattern which comprises peaks at 6.5°, 13.3°, 19.7°, and 23.8° ± 0.2 in 29.

[0081] In another embodiment, the present disclosure provides crystalline Form A of hydrochloride salt of Compound (I), wherein the molar ratio between Compound (I) and hydrochloric acid is 1 : 1, which is characterized by an X-ray powder diffraction pattern which comprises at least five, six, seven, or eight peaks chosen from 6.5°, 13.3°, 15.7°, 19.7°, 22.2°, 23.8°, 24.2°, and 28.6° ± 0.2 in 29.

[0082] In another embodiment, the present disclosure provides crystalline Form A of hydrochloride salt of Compound (I), wherein the molar ratio between Compound (I) and hydrochloric acid is 1 : 1, which is characterized by an X-ray powder diffraction pattern which comprises peaks at 6.5°, 7.8°, 13.3°, 14.0°, 15.7°, 16.8°, 19.7°, 22.2°, 23.5°, 23.8°, 24.2°, and 28.6° ± 0.2 in 29.

[0083] In another embodiment, the present disclosure provides a crystalline Form A of hydrochloride of Compound (I), wherein the molar ratio between Compound (I) and hydrochloric acid is 1 : 1, which is characterized by a differential scanning calorimeter (DSC) peak phase transition temperature of 120.2 ± 2 °C, 236.4 ± 2 °C, and 268.4 ± 2 °C.

[0084] In one embodiment, crystalline Form A of hydrochloride salt of Compound (I) is a hydrate.

[0085] Representative XRPD peaks are tabulated in Table 3. The XRPD patterns and peaks are shown in Figure 3 A. The Thermogravimetric Analysis (TGA) and Differential Scanning Calorimetry Analysis (DSC) thermograms are shown in Figure 3B.

[0086] Table 3. Representative XRPD peaks of crystalline Form A of Hydrochloride Salt of Compound (I)

[0087] Hydrochloride Salt Form B

[0088] In another embodiment, the present disclosure provides crystalline Form B of hydrochloride salt of Compound (I), wherein the molar ratio between Compound (I) and hydrochloric acid is 1 : 1, which is characterized by an X-ray powder diffraction pattern which comprises peaks at 5.7°, 6.6°, 6.9°, and 17.9° ± 0.2 in 29.

[0089] In another embodiment, the present disclosure provides crystalline Form B of hydrochloride salt of Compound (I), wherein the molar ratio between Compound (I) and hydrochloric acid is 1 : 1, which is characterized by an X-ray powder diffraction pattern which comprises at least five, six, seven, or eight peaks chosen from 5.7°, 6.6°, 6.9°, 7.2°, 13.6°, 14.4°, 17.9°, and 25.8° ± 0.2 in 29.

[0090] In another embodiment, the present disclosure provides crystalline Form B of hydrochloride salt of Compound (I), wherein the molar ratio between Compound (I) and hydrochloric acid is 1 : 1, which is characterized by an X-ray powder diffraction pattern which comprises peaks at 5.7°, 6.6°, 6.9°, 7.2°, 13.6°, 14.4°, 15.7°, 16.5°, 17.9°, 24.3°, 25.1°, and 25.8° ± 0.2 in 29.

[0091] In another embodiment, the present disclosure provides a crystalline Form B of hydrochloride salt of Compound (I), wherein the molar ratio between Compound (I) and hydrochloric acid is 1 : 1, which is characterized by a differential scanning calorimeter (DSC) peak phase transition temperature of 201.6 ± 2 °C, and 215.8 ± 2 °C.

[0092] In one embodiment, crystalline Form B of hydrochloride salt of Compound (I) is a solvate.

[0093] Representative XRPD peaks are tabulated in Table 4. The XRPD patterns and peaks are shown in Figure 4A. The Thermogravimetric Analysis (TGA) and Differential Scanning Calorimetry Analysis (DSC) thermograms are shown in Figure 4B. Table 4. Representative XRPD peaks of crystalline Form B of Hydrochloride Salt of Compound (I)

[0094] Sulfate Salt Form A

[0095] In another embodiment, the present disclosure provides crystalline Form A of sulfate salt of Compound (I), wherein the molar ratio between Compound (I) and sulfuric acid is 1 : 1, which is characterized by an X-ray powder diffraction pattern which comprises peaks at 6.1°, 13.6°, 18.9°, 23.5°, and 23.8° ± 0.2 in 20.

[0096] In another embodiment, the present disclosure provides crystalline Form A of sulfate salt of Compound (I), wherein the molar ratio between Compound (I) and sulfuric acid is 1 : 1, which is characterized by an X-ray powder diffraction pattern which comprises at least five, six, seven, or eight peaks chosen from 6.1°, 9.0°, 13.6°, 18.9°, 20.1°, 20.7°, 23.5°, 23.8° and 25.1° ± 0.2 in 20.

[0097] In another embodiment, the present disclosure provides crystalline Form A of sulfate salt of Compound (I), wherein the molar ratio between Compound (I) and sulfuric acid is 1 : 1, which is characterized by an X-ray powder diffraction pattern which comprises peaks at 6.1°, 9.0°, 13.2°, 13.6°, 14.2°, 14.8°, 16.3°, 17.9°, 18.9°, 20.1°, 20.7°, 23.5°, 23.8°, and 25.1° ± 0.2 in 20.

[0098] In another embodiment, the present disclosure provides a crystalline Form A of sulfate salt of Compound (I), wherein the molar ratio between Compound (I) and sulfuric acid is 1 : 1, which is characterized by a differential scanning calorimeter (DSC) peak phase transition temperature of 61.2 ± 2 °C and 207.3 ± 2 °C.

[0099] In one embodiment, crystalline Form A of sulfate salt of Compound (I) is a solvate. Representative XRPD peaks are tabulated in Table 5. The XRPD patterns and peaks are shown in Figure 5A. The Thermogravimetric Analysis (TGA) and Differential Scanning Calorimetry Analysis (DSC) thermograms are shown in Figure 5B.

[0100] Table 5. Representative XRPD peaks of crystalline Form A of Sulfate Salt of Compound (I)

[0101] Calcium Salt Form A

[0102] In another embodiment, the present disclosure provides crystalline Form A of calcium salt of Compound (I), wherein the molar ratio between Compound (I) and calcium hydroxide is 1 : 1, which is characterized by an X-ray powder diffraction pattern which comprises peaks at 5.9°, 18.9°, 26.7°, and 29.3° ± 0.2 in 26.

[0103] In another embodiment, the present disclosure provides crystalline Form A of calcium salt of Compound (I), wherein the molar ratio between Compound (I) and calcium hydroxide is 1 : 1, which is characterized by an X-ray powder diffraction pattern which comprises at least five, six, seven, or eight peaks chosen from 5.9°, 17.4°, 18.9°, 20.7°, 26.7°, 29.3°, 38.6°, and 39.3° ± 0.2 in 29.

[0104] In another embodiment, the present disclosure provides crystalline Form A of calcium salt of Compound (I), wherein the molar ratio between Compound (I) and calcium hydroxide is 1 : 1, which is characterized by an X-ray powder diffraction pattern which comprises peaks at 5.9°, 9.9°, 17.4°, 18.9°, 20.7°, 22.1°, 23.0°, 26.7°, 28.0°, 29.3°, 38.6°, and 39.3° ± 0.2 in 29.

[0105] In another embodiment, the present disclosure provides a crystalline Form A of calcium salt of Compound (I), wherein the molar ratio between Compound (I) and calcium hydroxide is 1 : 1, which is characterized by differential scanning calorimeter (DSC) peak phase transition temperature of 141.7 ± 2 °C, and 255.1 ± 2 °C. In one embodiment, crystalline Form A of calcium salt of Compound (I) is a hydrate.

[0106] Representative XRPD peaks are tabulated in Table 6. The XRPD patterns and peaks are shown in Figure 6A. The Thermogravimetric Analysis (TGA) and Differential Scanning Calorimetry Analysis (DSC) thermograms are shown in Figure 6B.

[0107] Table 6. Representative XRPD peaks of crystalline Form A of Calcium Salt of Compound (I)

[0108] Crystalline Forms o f Compound (I) Free Base

[0109] Form A

[0110] In one embodiment, the present disclosure provides crystalline Form A of Compound (I), which is characterized by an X-ray powder diffraction pattern which comprises peaks at 10.8°, 15.4°, 18.1°, and 25.1° ± 0.2 in 26.

[0111] In another embodiment, the present disclosure provides a crystalline Form A of Compound (I), which is characterized by an X-ray powder diffraction pattern which comprises at least five, six, seven, eight, nine, or ten peaks chosen from 10.8°, 13.2°, 15.4°, 18.1°, 18.6°, 19.9°, 21.8°, 22.8°, 24.0°, and 25.1° ± 0.2 in 29.

[0112] In another embodiment, the present disclosure provides a crystalline Form A of Compound (I), which is characterized by an X-ray powder diffraction pattern which comprises peaks at 10.8°, 11.8°, 13.2°, 15.4°, 18.1°, 18.6°, 19.9°, 20.4°, 21.8°, 22.8°, 24.0°, and 25.1° ± 0.2 in 29.

[0113] In another embodiment, the present disclosure provides a crystalline Form A of Compound (I), which is characterized by differential scanning calorimeter (DSC) peak phase transition temperature of 213.1 ± 2 °C.

[0114] Representative XRPD peaks are tabulated in Table 7. The XRPD patterns and peaks are shown in Figure 7A. The Thermogravimetric Analysis (TGA) and Differential Scanning Calorimetry Analysis (DSC) thermograms are shown in Figure 7B. Table 7. Representative XRPD peaks of crystalline Form A of Compound (I)

[0115] Methods of Treatment

[0116] Treatable Diseases

[0117] The HPK1 inhibitors, pharmaceutically acceptable salts thereof, pharmaceutical compositions thereof, may be used in methods of modulating ( / .< ., inhibiting) HPK1 activity, said method comprising administering to a patient / subject in need thereof an HPK1 inhibitor compound of the invention, or a pharmaceutically acceptable salt thereof, as described herein.

[0118] In particular, the present invention provides the use of the compounds of the invention, or stereoisomers, tautomers, N-oxides, hydrates, solvates, and salts thereof, particularly pharmaceutically acceptable salts thereof, or mixtures of same, for use in the treatment or prophylaxis of diseases, in particular cancer (particularly hematopoietic and solid tumors) or conditions with dysregulated immune responses or other disorders associated with aberrant MAP4K1 signaling. The pharmaceutical activity of the compounds according to the invention can at least be partially explained by their activity as MAP4K1 inhibitors.

[0119] In certain embodiments, the compounds of the invention, or pharmaceutically acceptable salts thereof, are useful for therapeutic administration to a subject in need thereof to treat a disease or indication including, but not limited to, benign hyperplasia, atherosclerotic disorder, sepsis, autoimmune disorder, vascular disorder, viral infection, neurodegenerative disorder, in inflammatory disorder, and male fertility control disorder.

[0120] In certain embodiments, the compounds of the invention, or pharmaceutically acceptable salts thereof, are useful for therapeutic administration to enhance, stimulate and / or increase immunity in treating cancer.

[0121] The HPK1 inhibitor compounds of the invention can be used alone, or in combination with other agents or therapies, or as an adjuvant or neoadjuvant for the treatment of diseases or disorders, including cancers.

[0122] In certain embodiments, the methods of the invention can be used to treat cancers that include, but are not limited to, bone cancer, pancreatic cancer, skin cancer, cancer of the head or neck, cutaneous or intraocular malignant melanoma, uterine cancer, ovarian cancer, rectal cancer, cancer of the anal region, stomach cancer, testicular cancer, uterine cancer, carcinoma of the fallopian tubes, carcinoma of the endometrium, endometrial cancer, carcinoma of the cervix, carcinoma of the vagina, carcinoma of the vulva, Hodgkin’s Disease, non-Hodgkin’s lymphoma, cancer of the esophagus, cancer of the small intestine, cancer of the endocrine system, cancer of the thyroid gland, cancer of the parathyroid gland, cancer of the adrenal gland, sarcoma of soft tissue, cancer of the urethra, cancer of the penis, chronic or acute leukemias including acute myeloid leukemia, chronic myeloid leukemia, acute lymphoblastic leukemia, chronic lymphocytic leukemia, solid tumors of childhood, lymphocytic lymphoma, cancer of the bladder, cancer of the kidney or urethra, carcinoma of the renal pelvis, neoplasm of the central nervous system (CNS), primary CNS lymphoma, tumor angiogenesis, spinal axis tumor, brain stem glioma, pituitary adenoma, Kaposi’s sarcoma, epidermoid cancer, squamous cell cancer, T-cell lymphoma, environmentally induced cancers including those induced by asbestos, and combinations of the above cancers.

[0123] In some embodiments, cancers treatable with compounds of the invention include melanoma (e.g., metastatic malignant melanoma), renal cancer (e.g, clear cell carcinoma), prostate cancer (e.g, hormone refractory prostate adenocarcinoma), breast cancer, triplenegative breast cancer, colon cancer and lung cancer (e.g., non-small cell lung cancer and small cell lung cancer). Additionally, refractory or recurrent malignancies whose growth may be inhibited using the compounds of the invention are also treatable.

[0124] In some embodiments, cancers that are treatable using the compounds of the invention include, but are not limited to, solid tumors (e.g., prostate cancer, colon cancer, esophageal cancer, endometrial cancer, ovarian cancer, uterine cancer, renal cancer, hepatic cancer, pancreatic cancer, gastric cancer, breast cancer, lung cancer, respiratory tract, brain cancer, eye cancer, thyroid and parathyoid cancer, skin cancer, cancers of the head and neck, cancer of the reproductive organs, cancer of the digestive tract, cancer of the urinary tract, glioblastoma, sarcoma, bladder cancer, etc.), hematological cancers (e.g., lymphoma, leukemia such as acute lymphoblastic leukemia (ALL), acute myelogenous leukemia (AML), chronic lymphocytic leukemia (CLL), chronic myelogenous leukemia (CML), DLBCL, mantle cell lymphoma, Non-Hodgkin lymphoma (including relapsed or refractory NHL and recurrent follicular), Hodgkin lymphoma or multiple myeloma), sarcoma, and distant metastasis thereof.

[0125] In some embodiments, diseases and indications that are treatable using the compounds of the invention include, but are not limited to, hematological cancers, sarcomas, lung cancers, gastrointestinal cancers, genitourinary tract cancers, liver cancers, bone cancers, nervous system cancers, gynecological cancers, and skin cancers.

[0126] Exemplary hematological cancers include lymphomas and leukemias, such as acute lymphoblastic leukemia (ALL), acute myelogenous leukemia (AML), acute promyelocyte leukemia (APL), chronic lymphocytic leukemia (CLL), chronic myelogenous leukemia (CML), diffuse large B-cell lymphoma (DLBCL), mantle cell lymphoma, Non-Hodgkin lymphoma (including relapsed or refractory NHL and recurrent follicular), Hodgkin lymphoma, myeloproliferative diseases (e.g., primary myelofibrosis (PMF), polycythemia vera (PV), essential thrombocytosis (ET), myelodysplasia syndrome (MDS), T-cell acute lymphoblastic lymphoma (T-ALL), multiple myeloma, cutaneous T-cell lymphoma, Waldenstrom’s Macroglubulinemia, hairy cell lymphoma, chronic myelogenic lymphoma, and Burkitt’s lymphoma.

[0127] Exemplary sarcomas include chondrosarcoma, Ewing’s sarcoma, osteosarcoma, rhabdomyosarcoma, angiosarcoma, fibrosarcoma, liposarcoma, myxoma, rhabdomyoma, rhabdosarcoma, fibroma, lipoma, harmatoma, and teratoma.

[0128] Exemplary lung cancers include non-small cell lung cancer (NSCLC), small cell lung cancer, bronchogenic carcinoma (squamous cell, undifferentiated small cell, undifferentiated large cell, adenocarcinoma), alveolar (bronchiolar) carcinoma, bronchial adenoma, chondromatous hamartoma, and mesothelioma.

[0129] Exemplary gastrointestinal cancers include cancers of the esophagus (squamous cell carcinoma, adenocarcinoma, leiomyosarcoma, lymphoma), stomach (carcinoma, lymphoma, leiomyosarcoma), pancreas (ductal adenocarcinoma, insulinoma, glucagonoma, gastrinoma, carcinoid tumors, vipoma), small bowel (adenocarcinoma, lymphoma, carcinoid tumors, Kaposi’s sarcoma, leiomyoma, hemangioma, lipoma, neurofibroma, fibroma), large bowel (adenocarcinoma, tubular adenoma, villous adenoma, hamartoma, leiomyoma), and colorectal cancer. Exemplary genitourinary tract cancers include cancers of the kidney (adenocarcinoma, Wilm’s tumor [nephroblastoma]), bladder and urethra (squamous cell carcinoma, transitional cell carcinoma, adenocarcinoma), prostate (adenocarcinoma, sarcoma), and testis (seminoma, teratoma, embryonal carcinoma, teratocarcinoma, choriocarcinoma, sarcoma, interstitial cell carcinoma, fibroma, fibroadenoma, adenomatoid tumors, lipoma).

[0130] Exemplary liver cancers include hepatoma (hepatocellular carcinoma), cholangiocarcinoma, hepatoblastoma, angiosarcoma, hepatocellular adenoma, and hemangioma.

[0131] Exemplary bone cancers include, for example, osteogenic sarcoma (osteosarcoma), fibrosarcoma, malignant fibrous histiocytoma, chondrosarcoma, Ewing’s sarcoma, malignant lymphoma (reticulum cell sarcoma), multiple myeloma, malignant giant cell tumor chordoma, osteochronfroma (osteocartilaginous exostoses), benign chondroma, chondroblastoma, chondromyxofibroma, osteoid osteoma, and giant cell tumors.

[0132] Exemplary nervous system cancers include cancers of the skull (osteoma, hemangioma, granuloma, xanthoma, osteitis deformans), meninges (meningioma, meningiosarcoma, gliomatosis), brain (astrocytoma, meduoblastoma, glioma, ependymoma, germinoma (pinealoma), glioblastoma, glioblastoma multiform, oligodendroglioma, schwannoma, retinoblastoma, congenital tumors), and spinal cord (neurofibroma, meningioma, glioma, sarcoma), as well as neuroblastoma and Lhermitte-Duclos disease.

[0133] Exemplary gynecological cancers include cancers of the uterus (endometrial carcinoma), cervix (cervical carcinoma, pre -tumor cervical dysplasia), ovaries (ovarian carcinoma (serous cystadenocarcinoma, mucinous cystadenocarcinoma, unclassified carcinoma), granulosa-thecal cell tumors, Sertoli-Leydig cell tumors, dysgerminoma, malignant teratoma), vulva (squamous cell carcinoma, intraepithelial carcinoma, adenocarcinoma, fibrosarcoma, melanoma), vagina (clear cell carcinoma, squamous cell carcinoma, botryoid sarcoma (embryonal rhabdomyosarcoma), and fallopian tubes (carcinoma).

[0134] Exemplary skin cancers include melanoma, basal cell carcinoma, squamous cell carcinoma, Kaposi’s sarcoma, Merkel cell skin cancer, moles dysplastic nevi, lipoma, angioma, dermatofibroma, and keloids. In some embodiments, diseases and indications that are treatable using the compounds of the invention include, but are not limited to, sickle cell disease (e.g., sickle cell anemia), triple-negative breast cancer (TNBC), myelodysplastic syndromes, testicular cancer, bile duct cancer, esophageal cancer, and urothelial carcinoma.

[0135] Exemplary head and neck cancers include glioblastoma, melanoma, rhabdosarcoma, lymphosarcoma, osteosarcoma, squamous cell carcinomas, adenocarcinomas, oral cancer, laryngeal cancer, nasopharyngeal cancer, nasal and paranasal cancers, and thyroid and parathyroid cancers.

[0136] In some embodiments, the subject HPK1 inhibitors may be used to treat tumors producing PGE2 (e.g., Cox-2 overexpressing tumors) and / or adenosine (CD73 and CD39 over-expressing tumors). Overexpression of Cox-2 has been detected in a number of tumors, such as colorectal, breast, pancreatic and lung cancers, where it correlates with a poor prognosis. Overexpression of COX-2 has been reported in hematological cancer models such as RAJI (Burkitt’s lymphoma) and U937 (acute promonocyte leukemia) as well as in patient’s blast cells. CD73 is up-regulated in various human carcinomas, including those of colon, lung, pancreas and ovary. Higher expression levels of CD73 have been associated with tumor neovascularization, invasiveness, and metastasis, and with shorter patient survival time in breast cancer.

[0137] Examples of treatable breast cancers include, but are not limited to, triple negative breast cancer, invasive ductal carcinoma, invasive lobular carcinoma, ductal carcinoma in situ, and lobular carcinoma in situ.

[0138] Examples of cancers of the respiratory tract include, but are not limited to, small-cell and non-small-cell lung carcinoma, as well as bronchial adenoma and pleuropulmonary blastoma.

[0139] Examples of treatable brain cancers include, but are not limited to, brain stem and hypophtalmic glioma, cerebellar and cerebral astrocytoma, glioblastoma, medulloblastoma, ependymoma, as well as neuroectodermal and pineal tumor.

[0140] Treatable tumors of the male reproductive organs include, but are not limited to, prostate and testicular cancer.

[0141] Treatable tumors of the female reproductive organs include, but are not limited to, endometrial, cervical, ovarian, vaginal, and vulvar cancer, as well as sarcoma of the uterus.

[0142] Treatable ovarian cancer include, but are not limited to serous tumor, endometrioid tumor, mucinous cystadenocarcinoma, granulosa cell tumor, Sertoli-Leydig cell tumor and arrhenoblastoma.

[0143] Treatable cervical cancer include, but are not limited to squamous cell carcinoma, adenocarcinoma, adenosquamous carcinoma, small cell carcinoma, neuroendocrine tumor, glassy cell carcinoma and villoglandular adenocarcinoma.

[0144] Treatable tumors of the digestive tract include, but are not limited to, anal, colon, colorectal, esophageal, gallbladder, gastric, pancreatic, rectal, small-intestine, and salivary gland cancers.

[0145] Treatable esophageal cancer include, but are not limited to esophageal cell carcinomas and adenocarcinomas, as well as squamous cell carcinomas, leiomyosarcoma, malignant melanoma, rhabdomyosarcoma and lymphoma.

[0146] Treatable gastric cancer include, but are not limited to intestinal type and diffuse type gastric adenocarcinoma.

[0147] Treatable pancreatic cancer include, but are not limited to ductal adenocarcinoma, adenosquamous carcinomas and pancreatic endocrine tumors.

[0148] Treatable tumors of the urinary tract include, but are not limited to, bladder, penile, kidney, renal pelvis, ureter, urethral and human papillary renal cancers.

[0149] Treatable kidney cancer include, but are not limited to renal cell carcinoma, urothelial cell carcinoma, juxtaglomerular cell tumor (reninoma), angiomyolipoma, renal oncocytoma, Bellini duct carcinoma, clear-cell sarcoma of the kidney, mesoblastic nephroma and Wilms' tumor.

[0150] Treatable bladder cancer include, but are not limited to transitional cell carcinoma, squamous cell carcinoma, adenocarcinoma, sarcoma and small cell carcinoma.

[0151] Treatable eye cancers include, but are not limited to, intraocular melanoma and retinoblastoma.

[0152] Treatable liver cancers include, but are not limited to, hepatocellular carcinoma (liver cell carcinomas with or without fibrolamellar variant), cholangiocarcinoma (intrahepatic bile duct carcinoma), and mixed hepatocellular cholangiocarcinoma.

[0153] Treatable skin cancers include, but are not limited to, squamous cell carcinoma, Kaposi's sarcoma, malignant melanoma, Merkel cell skin cancer, and non-melanoma skin cancer.

[0154] Treatable head-and-neck cancers include, but are not limited to, squamous cell cancer of the head and neck, laryngeal, hypopharyngeal, nasopharyngeal, oropharyngeal cancer, salivary gland cancer, lip and oral cavity cancer and squamous cell.

[0155] Treatable lymphomas include, but are not limited to, AIDS-related lymphoma, non- Hodgkin's lymphoma, cutaneous T-cell lymphoma, Burkitt lymphoma, Hodgkin's disease, and lymphoma of the central nervous system.

[0156] Treatable sarcomas include, but are not limited to, sarcoma of the soft tissue, osteosarcoma, malignant fibrous histiocytoma, lymphosarcoma, and rhabdomyosarcoma.

[0157] Treatable leukemias include, but are not limited to, acute myeloid leukemia, acute ymphoblastic leukemia, chronic lymphocytic leukemia, chronic myelogenous leukemia, and hairy cell leukemia.

[0158] In certain embodiments, the compounds of the invention can be used to treat a variety of other disorders in which MAP4K1 is involved, such as cardiovascular and lung diseases.

[0159] In certain embodiments, the compounds of the invention can be used in medicaments for the treatment and / or prophylaxis of cardiovascular, inflammatory and fibrotic disorders, renal disorders, in particular of acute and chronic renal insufficiency, and also of acute and chronic renal failure.

[0160] Here, the term “renal insufficiency” comprises both acute and chronic manifestations of renal insufficiency, and also underlying or related renal disorders such as diabetic and nondiabetic nephropathies, hypertensive nephropathies, ischaemic renal disorders, renal hypoperfusion, intradialytic hypotension, obstructive uropathy, renal stenoses, glomerulopathies, glomerulonephritis (such as, for example, primary glomerulonephritides; minimal change glomerulonephritis (lipoidnephrosis); membranous glomerulonephritis; focal segmental glomerulosclerosis (FSGS); membrane-proliferative glomerulonephritis; crescentic glomerulonephritis; mesangioproliferative glomerulonephritis (IgA nephritis, Berger's disease); post-infectious glomerulonephritis; secondary glomerulonephritides), diabetes mellitus, lupus erythematosus, amyloidosis, Goodpasture syndrome, Wegener granulomatosis, Henoch- Schonlein purpura, microscopic polyangiitis, acute glomerulonephritis, pyelonephritis (for example as a result of: urolithiasis, benign prostate hyperplasia, diabetes, malformations, abuse of analgesics, Crohn's disease), glomerulosclerosis, arteriolonecrose of the kidney, tubulointerstitial diseases, nephropathic disorders such as primary and congenital or aquired renal disorder, Alport syndrome, nephritis, immunological kidney disorders such as kidney transplant rejection and immunocomplex -induced renal disorders, nephropathy induced by toxic substances, nephropathy induced by contrast agents, diabetic and non-diabetic nephropathy, renal cysts, nephrosclerosis, hypertensive nephrosclerosis and nephrotic syndrome which can be characterized diagnostically, for example by abnormally reduced creatinine and / or water excretion, abnormally elevated blood concentrations of urea, nitrogen, potassium and / or creatinine, altered activity of renal enzymes, for example glutamyl synthetase, altered urine osmolarity or urine volume, elevated microalbuminuria, macroalbuminuria, lesions on glomerulae and arterioles, tubular dilatation, hyperphosphataemia and / or the need for dialysis.

[0161] In certain embodiments, the compounds of the invention can be used for the treatment and / or prophylaxis of sequelae of renal insufficiency, for example pulmonary oedema, heart failure, uremia, anemia, electrolyte disturbances (for example hypercalemia, hyponatremia) and disturbances in bone and carbohydrate metabolism.

[0162] In certain embodiments, the compounds of the invention can be used for the treatment and / or prevention of sequelae of renal insufficiency, for example pulmonary oedema, heart failure, uraemia, anaemia, electrolyte disturbances (for example hyperkalaemia, hyponatraemia) and disturbances in bone and carbohydrate metabolism.

[0163] In certain embodiments, the compounds of the invention are further suitable for the treatment and / or prevention of polycystic kidney disease (PCKD) and of the syndrome of inappropriate ADH secretion (SIADH).

[0164] In certain embodiments, the compounds of the invention are also suitable for the treatment and / or prophylaxis of metabolic syndrome, hypertension, resistant hypertension, acute and chronic heart failure, coronary heart disease, stable and unstable angina pectoris, peripheral and cardiac vascular disorders, arrhythmias, atrial and ventricular arrhythmias and impaired conduction, for example atrioventricular blocks degrees 1-111 (AB block 1-111), supraventricular tachyarrhythmia, atrial fibrillation, atrial flutter, ventricular fibrillation, ventricular flutter, ventricular tachyarrhythmia, Torsade de pointes tachycardia, atrial and ventricular extrasystoles, AV-junctional extrasystoles, sick sinus syndrome, syncopes, AV- nodal re-entry tachycardia, Wolff-Parkinson-White syndrome, of acute coronary syndrome (ACS), autoimmune cardiac disorders (pericarditis, endocarditis, valvolitis, aortitis, cardiomyopathies), shock such as cardiogenic shock, septic shock and anaphylactic shock, aneurysms, boxer cardiomyopathy (premature ventricular contraction (PVC)), for treatment and / or prophylaxis of thromboembolic disorders and ischaemias such as myocardial ischaemia, myocardial infarction, stroke, cardiac hypertrophy, transient and ischaemic attacks, preeclampsia, inflammatory cardiovascular disorders, spasms of the coronary arteries and peripheral arteries, oedema formation, for example pulmonary oedema, cerebral oedema, renal oedema or oedema caused by heart failure, peripheral circulatory disturbances, reperfusion damage, arterial and venous thromboses, myocardial insufficiency, endothelial dysfunction, to prevent restenoses, for example after thrombolysis therapies, percutaneous transluminal angioplasties (PTA), transluminal coronary angioplasties (PTCA), heart transplants and bypass operations, and also micro- and macrovascular damage (vasculitis), increased levels of fibrinogen and of low-density lipoprotein (LDL) and increased concentrations of plasminogen activator inhibitor 1 (PAI-1), and also for treatment and / or prophylaxis of erectile dysfunction and female sexual dysfunction.

[0165] In certain embodiments, the compounds of the invention are also suitable for treatment and / or prophylaxis of asthmatic disorders, pulmonary arterial hypertension (PAH) and other forms of pulmonary hypertension (PH) including left-heart disease, HIV, sickle cell anaemia, thromboembolisms (CTEPH), sarcoidosis, COPD or pulmonary fibrosis-associated pulmonary hypertension, chronic-obstructive pulmonary disease (COPD), acute respiratory distress syndrome (ARDS), acute lung injury (ALI), alpha- 1 -antitrypsin deficiency (AATD), pulmonary fibrosis, pulmonary emphysema (for example pulmonary emphysema induced by cigarette smoke) and cystic fibrosis (CF).

[0166] In certain embodiments, the compounds of the invention are also effective for the control of central nervous system disorders characterized by disturbances of the NO / cGMP system. They are suitable in particular for improving perception, concentration, learning or memory after cognitive impairments like those occurring in particular in association with situations / diseases / syndromes, such as mild cognitive impairment, age-associated learning and memory impairments, age-associated memory losses, vascular dementia, craniocerebral trauma, stroke, dementia occurring after strokes (post stroke dementia), post-traumatic craniocerebral trauma, general concentration impairments, concentration impairments in children with learning and memory problems, Alzheimer's disease, Lewy body dementia, dementia with degeneration of the frontal lobes including Pick's syndrome, Parkinson's disease, progressive dementia with corticobasal degeneration, amyolateral sclerosis (ALS), Huntington's disease, demyelinization, multiple sclerosis, thalamic degeneration, Creutzfeld- Jacob dementia, HIV dementia, schizophrenia with dementia or Korsakoff s psychosis.

[0167] In certain embodiments, the compounds of the invention are also suitable for treatment and / or prophylaxis of central nervous system disorders such as states of anxiety, tension and depression, CNS-related sexual dysfunctions and sleep disturbances, and for controlling pathological disturbances of the intake of food, stimulants and addictive substances. In certain embodiments, the compounds of the invention are furthermore also suitable for controlling cerebral blood flow and thus represent effective agents for controlling migraines.

[0168] In certain embodiments, the compounds of the invention are also suitable for the prophylaxis and control of sequelae of cerebral infarction (cerebral apoplexy) such as stroke, cerebral ischaemia and craniocerebral trauma. The compounds according to the invention can likewise be used for controlling states of pain and tinnitus.

[0169] In certain embodiments, the compounds of the invention have anti-inflammatory action and can therefore be used as anti-inflammatory agents for treatment and / or prophylaxis of sepsis (SIRS), multiple organ failure (MODS, MOF), inflammatory disorders of the kidney, chronic intestinal inflammations (IBD, Crohn's disease, UC), pancreatitis, peritonitis, rheumatoid disorders, inflammatory skin disorders and inflammatory eye disorders.

[0170] In certain embodiments, the compounds of the invention can also be used for treatment and / or prophylaxis of autoimmune diseases.

[0171] In certain embodiments, the compounds of the invention are also suitable for treatment and / or prophylaxis of fibrotic disorders of the internal organs, for example the lung, the heart, the kidney, the bone marrow and in particular the liver, and also dermatological fibroses and fibrotic eye disorders.

[0172] As used herein, the term “fibrotic disorders” includes in particular the following: hepatic fibrosis, cirrhosis of the liver, pulmonary fibrosis, endomyocardial fibrosis, nephropathy, glomerulonephritis, interstitial renal fibrosis, fibrotic damage resulting from diabetes, bone marrow fibrosis and similar fibrotic disorders, scleroderma, morphea, keloids, hypertrophic scarring (also following surgical procedures), naevi, diabetic retinopathy, proliferative vitroretinopathy and disorders of the connective tissue (for example sarcoidosis).

[0173] In certain embodiments, the compounds of the invention are also suitable for controlling postoperative scarring, for example as a result of glaucoma operations.

[0174] In certain embodiments, the compounds of the invention can also be used cosmetically for ageing and keratinized skin.

[0175] In certain embodiments, the compounds of the invention are suitable for treatment and / or prophylaxis of hepatitis, neoplasms, osteoporosis, glaucoma and gastroparesis.

[0176] In certain embodiments, the compounds of the invention are suitable for treatment and / or prophylaxis of viral infections (e.g., HIV and Kaposi's sarcoma); inflammatory and autoimmune diseases (e.g., colitis, arthritis, Alzheimer's disease, glomerulonephritis and wound healing); bacterial, fungal and / or parasitic infections; skin diseases (e.g., psoriasis); diseases based on hyperplasia which are characterised by an increase in the number of cells (e.g., fibroblasts, hepatocytes, bones and bone marrow cells, cartilage or smooth muscle cells or epithelial cells (e.g., endometrial hyperplasia)); bone diseases and cardiovascular diseases (e.g., restenosis and hypertrophy).

[0177] In another embodiment, the inventive compounds can also be used to treat or to prevent uterine fibroids (uterine leiomyoma or uterine myoma) in women. Uterine fibroids are benign tumors of the myometrium, the smooth muscle layer of the uterus. Uterine fibroids grow slowly during a women's life, and their growth is dependent on the female sexual hormones estradiol and progesterone. Therefore, the highest prevalence of uterine fibroids with approx. 70% and >80% in white and afro-american women, respectively, is found from 35 years of age onwards to menopause, when they shrink due to reduced hormone levels. Approx. 30% and 45% of white and afro-american women, respectively, do show clinically relevant symptoms due to their fibroids, which are heavy menstrual bleeding and pain, which is related to the menstrual cycle (David el al., Eur J Ob stet Gynecol Reprod Biol. 199: 137-140, 2016). Heavy menstrual bleeding in this respect is defined by a blood loss of more than 80 mL in a menstrual bleeding period. Submucosal position of the uterine fibroids, e.g., those located directly below the endometrium, seems to have an even more severe effect on uterine bleeding, which may result in anemia in affected women. Furthermore, uterine fibroids, due to their symptoms, do severly affect the quality of life of affected women.

[0178] In certain embodiments, the compounds of the invention are useful for the treatment and / or prophylaxis of chronic renal disorders, acute and chronic renal insufficiency, diabetic, inflammatory or hypertensive nephropaties, fibrotic disorders, cardiac insufficiency, angina pectoris, hypertension, pulmonary hypertension, ischemias, vascular disorders, thromboembolic disorders, arteriosclerosis, sickle cell anemia, erectile dysfunction, benign prostate hyperplasia, dysuria associated with benign prostate hyperplasia, Huntington, dementia, Alzheimer and Creutzfeld-Jakob.

[0179] The present invention provides a method for the treatment and / or prophylaxis of chronic renal disorders, acute and chronic renal insufficiency, diabetic, inflammatory or hypertensive nephropathies, fibrotic disorders, cardiac insufficiency, angina pectoris, hypertension, pulmonary hypertension, ischemias, vascular disorders, thromboembolic disorders, arteriosclerosis, sickle cell anemia, erectile dysfunction, benign prostate hyperplasia, dysuria associated with benign prostate hyperplasia, Huntington, dementia, Alzheimer and Creutzfeld- Jakob.

[0180] The present invention further provides the use of the compounds according to the invention for treatment and / or prophylaxis of disorders, especially the disorders mentioned above.

[0181] The present invention further provides a method for treatment and / or prophylaxis of disorders, in particular the disorders mentioned above, using an effective amount of at least one of the compounds according to the invention.

[0182] Thus, compounds of the present invention can be utilized to inhibit, block, reduce or decrease MAP4K1 activation by exogenous and / or endogenous ligands for the reduction of tumor growth and the modulation of dysregulated immune responses, e.g., to block immunosuppression and increase immune cell activation and infiltration in the context of cancer and cancer immunotherapy. This method comprises administering to a mammal in need thereof, including a human, an amount of a compound of this invention, or a pharmaceutically acceptable salt, isomer, polymorph, metabolite, hydrate, solvate or ester thereof; which is effective to treat the disorder.

[0183] The present invention also provides methods of treating a variety of other disorders wherein MAP4K1 is involved such as, but not limited to, disorders with dysregulated immune responses, inflammation, vaccination for infection & cancer, viral infections, obesity and diet- induced obesity, adiposity, metabolic disorders, hepatic steatosis and uterine fibroids. These disorders have been well characterized in humans, but also exist with a similar etiology in other mammals, and can be treated by administering pharmaceutical compositions of the present invention.

[0184] Combination Therapy

[0185] The compounds of the invention may be used in combination therapy with one or more additional / secondary therapeutic agents suitable for treating a disease or indication treatable by the subject compounds.

[0186] Thus in certain embodiments, for example, methods of the invention using compounds of the invention may comprise administering to the subject in need thereof a further therapeutic agent. The further therapeutic agent may be: (i) an immunomodulatory agent which blocks or inhibits an immune system checkpoint, which checkpoint may or may not be a component of the NF-KB pathway; and / or (ii) an agent which directly stimulates an immune effector response, such as a cytokine, or a tumor specific adoptively transferred T cell population, or an antibody specific for a protein expressed by a tumor cell; and / or (iii) a composition comprising a tumor antigen or immunogenic fragment thereof; and / or (iv) a chemotherapeutic agent.

[0187] In certain embodiments, the second therapeutic agent comprises an inhibitor of the PI3K-AKT-mT0R pathway, an inhibitor of the Raf-MAPK pathway, an inhibitors of the JAK-STAT pathway, an inhibitor of the beta catenin pathway, an inhibitor of notch pathway, an inhibitor of the hedgehog pathway, an inhibitor of the Pirn kinases, and / or an inhibitor of protein chaperones and cell cycle progression. In certain embodiments, combination therapy of the invention reduces the likelihood of drug-resistance arising in a cell population, and / or reduces the toxicity of treatment.

[0188] In certain embodiments, the HPK1 inhibitor compounds of the invention can be combined with one or more inhibitors of the following kinases for the treatment of cancer: Aktl, Akt2, Akt3, TGF-pPv, PKA, PKG, PKC, CaM-kinase, phosphorylase kinase, MEKK, ERK, MAPK, mTOR, EGFR, HER2, HER3, HER4, INS-R, IGF-1R, IR-R, PDGFaR, PDGFPR, CSFIR, KIT, FLK-II, KDR / FLK-1, FLK-4, flt-1, FGFR1, FGFR2, FGFR3, FGFR4, c-Met, Ron, Sea, TRKA, TRKB, TRKC, FLT3, VEGFR / Flt2, Flt4, EphAl, EphA2, EphA3, EphB2, EphB4, Tie2, Src, Fyn, Lek, Fgr, Btk, Fak, SYK, FRK, JAK, ABL, ALK, and B-Raf.

[0189] In certain embodiments, the HPK1 inhibitor compounds of the invention can be combined with one or more of the following inhibitors for the treatment of cancer, including an FGFR inhibitor (FGFR1, FGFR2, FGFR3 or FGFR4, e.g, AZD4547, BAY 1187982, ARQ087, BGJ398, BIBF1120, TKI258, lucitanib, dovitinib, TAS-120, J J-42756493, Debiol347, INCB54828, INCB62079, and INCB63904), a JAK inhibitor (JAK1 and / or JAK2, e.g., ruxolitinib, baricitinib, or itacitinib (INCB39110)), an IDO inhibitor (e.g., epacadostat and NLG919), an LSD1 inhibitor (e.g., GSK2979552, INCB59872 and INCB60003), a TDO inhibitor, a PI3K-delta inhibitor (e.g., INCB50797 and INCB50465), a PI3K-gamma inhibitor such as a PI3K-gamma selective inhibitor, a CSFIR inhibitor (e.g., PLX3397 and LY3022855), a TAM receptor tyrosine kinases (Tyro-3, Axl, and Mer), an aryl hydrocarbon receptor (AhR) modulator (such as laquinimod, aminoflavone, CB7993113, CH223191, 6, 2’, 4’ -trimethoxyflavone (TMF), GNF351 (N-(2-(lH-indol-3-yl)ethyl)-9- isopropyl-2-(5-methylpyridin-3-yl)-9H-purin-6-amine), aminoflavone, NKI150460, indole-3- carbinol, P -naphthoflavone and dimer thereof, diindolylmethane (DIM), 4- Hydroxytamoxifen, leflunomide, raloxifene, tranilast, flutamide, mexiletine, nimodiphine, omeprazole, sulindac, tranilast, and TCDD (2,3,7,8-tetrachlorodibenzo-p-dioxin)), an angiogenesis inhibitor, an interleukin receptor inhibitor, bromo and extra terminal family members inhibitors (for example, bromodomain inhibitors or BET inhibitors such as OTX015, CPI-0610, INCB54329, and INCB57643), and an adenosine receptor antagonist or combinations thereof.

[0190] In certain embodiments, the HPK1 inhibitor compounds of the invention can be combined with inhibitors of HD AC, such as panobinostat and vorinostat.

[0191] In certain embodiments, the HPK1 inhibitor compounds of the invention can be combined with inhibitors of c-Met, such as onartumzumab, tivantnib, and capmatinib (INC- 280).

[0192] In certain embodiments, the HPK1 inhibitor compounds of the invention can be combined with inhibitors of BTK, such as ibrutinib.

[0193] In certain embodiments, the HPK1 inhibitor compounds of the invention can be combined with inhibitors of mTOR, e.g, rapamycin, sirolimus, temsirolimus, and everolimus.

[0194] In certain embodiments, the HPK1 inhibitor compounds of the invention can be combined with inhibitors of MEK, such as trametinib, selumetinib and GDC-0973.

[0195] In certain embodiments, the HPK1 inhibitor compounds of the invention can be combined with inhibitors of Hsp90 (e.g., tanespimycin), cyclin dependent kinases (e.g., palbociclib), PARP (e.g., olaparib) and Pirn kinases ( LGH447, INCB053914, and SGI- 1776).

[0196] In certain embodiments, the HPK1 inhibitor compounds of the invention can be combined with an agonist of DNA sensor (c-GAS) and / or its downstream adaptor protein STING.

[0197] The cGAS (cyclic GMP-AMP Synthase)-STING (Stimulator of Interferon Genes) pathway is a component of the innate immune system that functions to detect the presence of cytosolic DNA and, in response, trigger expression of inflammatory genes that can lead to senescence or to the activation of defense mechanisms. Localization of DNA from the usual nuclear localization to the cytosol is associated with tumorigenesis or viral infection. cGAS is found in the cytosol and, upon direct binding to cytosolic DNA, cGAS forms dimers to catalyze production of 2’3’-cGAMP from ATP and GTP. The resulting cGAMP then acts a second messenger to bind STING and to trigger activation of the transcription factor IRF3. Activated IRF3 leads to transcription of type-1 IFN-P, and a number of downstream target genes to initiate a diverse array of biological responses, such as viral response, tumor surveillance, autoimmunity, and cellular senescence. In many tumor cells, constitutively active DNA damage response leads to the accumulation of cytoplasmic DNA and activation of the cGAS / STING pathway. It has been shown in lymphoma cells that the NKG2D ligand, Rael, was upregulated in a STING / IRF3 dependent manner, so as to aid in NK-mediated tumor clearance. The activation of c-GAS-STING pathway in antigen-presenting cells, such as dendritic cells, has been shown to enhance their function and boost anti-tumor immunity.

[0198] In certain embodiments, the HPK1 inhibitor compounds of the invention can be combined with one or more immune checkpoint inhibitors.

[0199] Effector T cell activation is normally triggered by the TCR recognizing antigenic peptide presented by the MHC complex. The type and level of activation achieved is then determined by the balance between signals which stimulate and signals which inhibit the effector T cell response. “Immune system checkpoint” is used herein to refer to any molecular interaction which alters the balance in favor of inhibition of the effector T cell response. That is, a molecular interaction which, when it occurs, negatively regulates the activation of an effector T cell. Such an interaction might be direct, such as the interaction between a ligand and a cell surface receptor which transmits an inhibitory signal into an effector T cell. Or it might be indirect, such as the blocking or inhibition of an interaction between a ligand and a cell surface receptor which would otherwise transmit an activatory signal into the effector T cell, or an interaction which promotes the upregulation of an inhibitory molecule or cell, or the depletion by an enzyme of a metabolite required by the effector T cell, or any combination thereof.

[0200] Examples of immune system checkpoints include: a) The interaction between indoleamine 2,3 -dioxygenase (IDO1) and its substrate; b) The interaction between PD1 and PD-L1 and / or PD1 and PD-L2; c) The interaction between CTLA-4 and CD86 and / or CTLA- 4 and CD80; d) The interaction between B7-H3 and / or B7-H4 and their respective ligands; e) The interaction between HVEM and BTLA; f) The interaction between GAL9 and TIM3; g) The interaction between MHC class I or II and LAG 3; and h) The interaction between MHC class I or II and KIR; i) The interaction between 0X40 (CD134) and OX40L (CD252); j) The interaction between CD40 and CD40L (CD 154); k) The interaction between 4-1 BB (CD 137) and ligands including 4-1 BBL; 1) The interaction between GITR and ligands including GITRL.

[0201] Thus exemplary immune checkpoint inhibitors include inhibitors against immune checkpoint molecules such as CD20, CD27, CD28, CD39, CD40, CD 122, CD96, CD73, CD47, 0X40, GITR, CSF1R, JAK, PI3K delta, PI3K gamma, TAM, arginase, CD137 (also known as 4-1BB), ICOS, A2AR, B7-H3, B7-H4, BTLA, CTLA-4, LAG3, TIM3, VISTA, PD-1, PD-L1, and PD-L2.

[0202] A representative checkpoint for the purposes of the present invention is checkpoint (b), namely the interaction between PD1 and either of its ligands PD-L1 and PD-L2. PD1 is expressed on effector T cells. Engagement with either ligand results in a signal which downregulates activation. The ligands are expressed by some tumors. PD-L1 in particular is expressed by many solid tumors, including melanoma. These tumors may therefore down regulate immune mediated anti -turn or effects through activation of the inhibitory PD-1 receptors on T cells. By blocking the interaction between PD1 and one or both of its ligands, a checkpoint of the immune response may be removed, leading to augmented anti-tumor T cell responses. Therefore, PD1 and its ligands are examples of components of an immune system checkpoint which may be targeted in the method of the invention.

[0203] Another checkpoint for the purposes of the present invention is checkpoint (c), namely the interaction between the T cell receptor CTLA-4 and its ligands, the B7 proteins (B7-1 and B7-2). CTLA-4 is ordinarily upregulated on the T cell surface following initial activation, and ligand binding results in a signal which inhibits further / continued activation. CTLA-4 competes for binding to the B7 proteins with the receptor CD28, which is also expressed on the T cell surface but which upregulates activation. Thus, by blocking the CTLA-4 interaction with the B7 proteins, but not the CD28 interaction with the B7 proteins, one of the normal check points of the immune response may be removed, leading to augmented anti-tumor T cell responses. Therefore, CTLA-4 and its ligands are examples of components of an immune system checkpoint which may be targeted in the method of the invention.

[0204] In some embodiments, the immune checkpoint molecule is a stimulatory checkpoint molecule selected from CD27, CD28, CD40, ICOS, 0X40, GITR, and CD137.

[0205] In some embodiments, the immune checkpoint molecule is an inhibitory checkpoint molecule selected from A2AR, B7-H3, B7-H4, BTLA, CTLA-4, IDO, KIR, LAG3, N0X2, PD-1, TIM3, SIGLEC7, SIGLEC9, and VISTA, and binding partners thereof (such as PD-L1 and PD-L2). In some embodiments, the compounds provided herein can be used in combination with one or more agents selected from KIR inhibitors, TIGIT inhibitors, LAIR1 inhibitors, CD 160 inhibitors, 2B4 inhibitors and TGFR beta inhibitors.

[0206] In some embodiments, the compounds provided herein can be used in combination with immune checkpoint inhibitors that are small molecule inhibitors (SMI), which are typically small organic molecules. For example, in certain embodiments, inhibitors of IDO 1 include Epacadostat (INCB24360), Indoximod, GDC-0919 (NLG919) and F001287. Other inhibitors of IDO 1 include 1 -methyltryptophan (1 MT).

[0207] In some embodiments, the inhibitor of an immune checkpoint molecule is also known as an “immunomodulatory agent,” which includes any agent which, when administered to a subject, blocks or inhibits the action of an immune system checkpoint, resulting in the upregulation of an immune effector response in the subject, typically a T cell effector response, which may comprise an anti-tumor T cell effector response.

[0208] The immunomodulatory agent used in the method of the present invention may block or inhibit any of the immune system checkpoints described above. The agent may be an antibody or any other suitable agent which results in said blocking or inhibition. The agent may thus be referred to generally as an inhibitor of a said checkpoint.

[0209] An “antibody” as used herein includes whole antibodies and any antigen binding fragment ( / .< ., “antigen-binding portion”) or single chains thereof. An antibody may be a polyclonal antibody or a monoclonal antibody, and may be produced by any suitable method. Examples of binding fragments encompassed within the term “antigen-binding portion” of an antibody include a Fab fragment, a F(ab’)2 fragment, a Fab’ fragment, a Fa fragment, a Fv fragment, a dAb fragment, and an isolated complementarity determining region (CDR). Single chain antibodies such as scFv and heavy chain antibodies such as VHH and camel antibodies are also intended to be encompassed within the term “antigen-binding portion” of an antibody.

[0210] In certain embodiments, the immunomodulatory agent used with the HPK1 inhibitor of the invention is anti-PDl antibody, anti-PD-Ll antibody, anti-PD-L2 antibody, or anti- CTLA-4 antibody.

[0211] In some embodiments, the inhibitor of an immune checkpoint molecule is an inhibitor of PD-1, e.g., an anti-PD-1 monoclonal antibody. In some embodiments, the anti-PD-1 monoclonal antibody is nivolumab (MDX-1106), pembrolizumab (Merck 3475 or Lambrolizumab), pidilizumab (CT-011), Tislelizumab (BGB-A317), Camrelizumab (SHR- 1210), spartalizumab (PDR001), or AMP-514 (MEDI0680). In some embodiments, the anti- PD-1 monoclonal antibody is nivolumab or pembrolizumab. In some embodiments, the anti- PD1 antibody is pembrolizumab. In some embodiments, the anti PD-1 antibody is Camrelizumab (SHR-1210). In certain embodiments, the inhibitor of PD-1 is AMP -224 (PD- L2 Fcfusion protein that binds PD-1) or AUNP-12 (anti-PD-1 peptide).

[0212] In some embodiments, the inhibitor of an immune checkpoint molecule is an inhibitor of PD-L1, e.g., an anti-PD-Ll monoclonal antibody. In some embodiments, the anti-PD-Ll monoclonal antibody is BMS-935559, BMS-936559 (MDX-1105), MEDI-4736 (durvalumab), MPDL3280A (also known as RG7446), YW243.55.S70 (HPAB-0381-WJ), or MSB0010718C. In some embodiments, the anti-PD-Ll monoclonal antibody is MPDL3280A or MEDI-4736. In certain embodiments, anti-PD-Ll antibodies include atezolizumab, avelumab, durvalumab or MEDI-4736, and MPDL3280A.

[0213] In some embodiments, the inhibitor of an immune checkpoint molecule is an inhibitor of CTLA-4, e.g., an anti-CTLA-4 antibody. In some embodiments, the anti-CTLA-4 antibody is ipilimumab, tremelimumab, or any of the antibodies disclosed in W02014 / 207063 (incorporated herein by reference). Other molecules include polypeptides, or soluble mutant CD86 polypeptides. In certain embodiments, the antibody is Ipilumumab.

[0214] In certain embodiments, the inhibitor of an immune checkpoint molecule is a combination of two or more of the modulators described herein, such as a combination that targets two or more different targets (e.g., PD-1, PD-L1 and PD-L2). Exemplary combinations include: a-PD-1 and a-PD-Ll; a-CTLA-4, a-PD-Ll, and a-CD20; etc..

[0215] In some embodiments, the inhibitor of an immune checkpoint molecule is an antibody which blocks or inhibits the interaction between 4-1 BB and its ligand, including utomilumab.

[0216] In some embodiments, the inhibitor of an immune checkpoint molecule is an inhibitor of CSFIR, e.g., an anti-CSFIR antibody. In some embodiments, the anti-CSFIR antibody is IMC-CS4 or RG7155.

[0217] In some embodiments, the inhibitor of an immune checkpoint molecule is an inhibitor of LAG3, e.g., an anti-LAG3 antibody. In some embodiments, the anti-LAG3 antibody is BMS-986016, LAG525, IMP321 or GSK2831781.

[0218] In some embodiments, the inhibitor of an immune checkpoint molecule is an inhibitor of GITR, e.g., an anti-GITR antibody. In some embodiments, the anti-GITR antibody is TRX518, MK-4166, MK1248, BMS-986156, MEDI1873, or GWN323.

[0219] In some embodiments, the inhibitor of an immune checkpoint molecule is an inhibitor of 0X40, e.g., an anti-OX40 antibody or OX40L fusion protein. In some embodiments, the anti-OX40 antibody is MEDI0562, MEDI6469, MOXR0916, PF-04518600, or GSK3174998. In some embodiments, the OX40L fusion protein is MED 16383.

[0220] In some embodiments, the inhibitor of an immune checkpoint molecule is an inhibitor of TIM3, e.g., an anti-TIM3 antibody. In some embodiments, the anti-TIM3 antibody is MBG-453.

[0221] In some embodiments, the inhibitor of an immune checkpoint molecule is an inhibitor of CD20, e.g., an anti-CD20 antibody. In some embodiments, the anti-CD20 antibody is obinutuzumab or rituximab.

[0222] In some embodiments, the compounds of the invention can be used in combination with one or more metabolic enzyme inhibitors. In some embodiments, the metabolic enzyme inhibitor is an inhibitor of IDO 1, TDO, or arginase. Examples of IDO 1 inhibitors include epacadostat and NGL919. An example of an arginase inhibitor is CB-1158.

[0223] In some embodiments, the compounds of the invention can be used in combination with bispecific antibodies. In some embodiments, one of the domains of the bispecific antibody targets PD-1, PD-L1, CTLA-4, GITR, 0X40, TIM3, LAG3, CD137, ICOS, CD3 or TGFp receptor.

[0224] In some embodiments, the compounds of the invention can be used in combination with one or more agents for the treatment of diseases such as cancer. In some embodiments, the agent is an alkylating agent, a proteasome inhibitor, a corticosteroid, or an immunomodulatory agent. Examples of an alkylating agent include bendamustine, nitrogen mustards, ethylenimine derivatives, alkyl sulfonates, nitrosoureas and triazenes, uracil mustard, chlormethine, cyclophosphamide (Cytoxan™), ifosfamide, melphalan, chlorambucil, pipobroman, triethylene-melamine, triethylenethiophosphoramine, busulfan, carmustine, lomustine, streptozocin, dacarbazine, and temozolomide. In some embodiments, the proteasome inhibitor is carfilzomib. In some embodiments, the corticosteroid is dexamethasone (DEX). In some embodiments, the immunomodulatory agent is lenalidomide (LEN) or pomalidomide (POM).

[0225] The compounds of the present disclosure can further be used in combination with other methods of treating cancers, for example by chemotherapy, irradiation therapy, tumor- targeted therapy, adjuvant therapy, immunotherapy or surgery. Examples of immunotherapy include cytokine treatment (e.g., interferons, GM-CSF, G-CSF, IL-2), CRS-207 immunotherapy, cancer vaccine, monoclonal antibody, adoptive T cell transfer, oncolytic virotherapy and immunomodulating small molecules, including thalidomide or JAK1 / 2 inhibitor and the like.

[0226] The compounds of the invention can be administered in combination with one or more anti-cancer drugs, such as a chemotherapeutics. Example chemotherapeutics include any of: abarelix, abiraterone, afatinib, aflibercept, aldesleukin, alemtuzumab, alitretinoin, allopurinol, altretamine, anastrozole, arsenic trioxide, asparaginase, axitinib, azacitidine, bevacizumab, bexarotene, baricitinib, bicalutamide, bleomycin, bortezombi, bortezomib, brivanib, buparlisib, busulfan intravenous, busulfan oral, calusterone, capecitabine, carboplatin, carmustine, cediranib, cetuximab, chlorambucil, cisplatin, cladribine, clofarabine, crizotinib, cyclophosphamide, cytarabine, dacarbazine, dacomitinib, dactinomycin, dalteparin sodium, dasatinib, dactinomycin, daunorubicin, decitabine, degarelix, denileukin, denileukin diftitox, deoxycoformycin, dexrazoxane, docetaxel, doxorubicin, droloxafine, dromostanolone propionate, eculizumab, enzalutamide, epidophyllotoxin, epirubicin, erlotinib, estramustine, etoposide phosphate, etoposide, exemestane, fentanyl citrate, filgrastim, floxuridine, fludarabine, fluorouracil, flutamide, fulvestrant, gefitinib, gemcitabine, gemtuzumab ozogamicin, goserelin acetate, histrelin acetate, ibritumomab tiuxetan, idarubicin, idelalisib, ifosfamide, imatinib mesylate, interferon alfa 2a, irinotecan, lapatinib ditosylate, lenalidomide, letrozole, leucovorin, leuprolide acetate, levamisole, lomustine, meclorethamine, megestrol acetate, melphalan, mercaptopurine, methotrexate, methoxsalen, mithramycin, mitomycin C, mitotane, mitoxantrone, nandrolone phenpropionate, navelbene, necitumumab, nelarabine, neratinib, nilotinib, nilutamide, nofetumomab, oserelin, oxaliplatin, paclitaxel, pamidronate, panitumumab, pazopanib, pegaspargase, pegfilgrastim, pemetrexed disodium, pentostatin, pilaralisib, pipobroman, plicamycin, ponatinib, prednisone, procarbazine, quinacrine, rasburicase, regorafenib, reloxafine, rituximab, ruxolitinib, sorafenib, streptozocin, sunitinib, sunitinib maleate, tamoxifen, tegafur, temozolomide, teniposide, testolactone, thalidomide, thioguanine, thiotepa, topotecan, toremifene, tositumomab, trastuzumab, tretinoin, triptorelin, uracil mustard, valrubicin, vandetanib, vinblastine, vincristine, vinorelbine, vorinostat, and zoledronate. Other anti-cancer agent(s) include antibody therapeutics such as trastuzumab (Herceptin), antibodies to costimulatory molecules such as CTLA-4 (e.g., ipilimumab or tremelimumab), 4- IBB, antibodies to PD-1 and PD-L1, or antibodies to cytokines (IL- 10, TGF-P, efc.). Examples of antibodies to PD-1 and / or PD-L1 that can be combined with compounds of the present disclosure for the treatment of cancer or infections such as viral, bacteria, fungus, and parasite infections include, but are not limited to, nivolumab, pembrolizumab, MPDL3280A, MEDI-4736, and SHR-1210.

[0227] Other anti-cancer agents include inhibitors of kinases associated cell proliferative disorder. These kinases include but not limited to Aurora-A, CDK1, CDK2, CDK3, CDK5, CDK7, CDK8, CDK9, ephrin receptor kinases, CHK1, CHK2, SRC, Yes, Fyn, Lek, Fer, Fes, Syk, Itk, Bmx, GSK3, JNK, PAK1, PAK2, PAK3, PAK4, PDK1, PKA, PKC, Rsk, and SGK.

[0228] Other anti-cancer agents also include those that block immune cell migration such as antagonists to chemokine receptors, including CCR2 and CCR4. The compounds of the present disclosure can further be used in combination with one or more anti-inflammatory agents, steroids, immunosuppressants, or therapeutic antibodies.

[0229] In some embodiments, the compounds of the invention can be used in combination with a further therapeutic agent which directly stimulates an immune effector response, such as a cytokine, or a tumor specific adoptively transferred T cell population, or an antibody specific for a protein expressed by a tumor cell.

[0230] As used herein, “an agent which directly stimulates an immune effector response” means any suitable agent, but typically refers to a cytokine or chemokine (or an agent which stimulates production of either), a tumor specific adoptively transferred T cell population, or an antibody specific for a protein expressed by a tumor cell.

[0231] The cytokine may be an interferon selected from IFNa, IPNP, IFNy and IFNA, or an interleukin, such as IL-2. The chemokine may be an inflammatory mediator, for example selected from CXCL9, 10, and 11, which attract T cells expressing CXCR3. The agent which stimulates production of a cytokine or chemokine may be an adjuvant suitable for administration to humans. One example is Bacille Calmette-Guerin (BCG), which is typically administered intravesical (i.e. urethral catheter) for treatment of bladder cancer. A typical dosage regime of BCG for bladder cancer is once per week for six weeks, but given its long safety history it is also administered indefinitely as maintenance. BCG has been shown to stimulate immune responses to bladder cancer. BCG has also been used as an adjuvant in combination with compositions which comprise tumor antigens (i.e. with cancer vaccines), particularly for colon cancer when it is administered typically intradermally. Such uses of BCG are also envisaged in the present invention. The tumor specific adoptively transferred T cell population directly increases the size of the tumor specific T cell population in an individual, and may be generated by any suitable means. However, typically the process involves isolating tumor specific T cells from a tumor sample taken from a patient, and selectively culturing those cells before returning the expanded population of tumorspecific T cells to the patient. Alternatively a tumor specific T cell population may be produced by genetic engineering of the T cell receptor locus, followed by expansion of the altered cell.

[0232] Antibodies specific for proteins expressed by a tumor cell typically stimulate immune activity by binding to the tumor cell and promoting destruction of the cell via antibodydependent cell-mediated cytotoxicity (ADCC). Examples of antibodies of this type include anti-CD20 antibodies such as ofatumumab or rituximab, and anti-CD52 antibodies such as alemtuzumab.

[0233] Thus in certain exemplary embodiments, the compounds of the invention may be used in combination with a calcineurin inhibitor, e.g., cyclosporin A or FK 506; a mTOR inhibitor, e.g., rapamycin, 40-0-(2-hydroxyethyl)-rapamycin, biolimus-7 or biolimus-9; an ascomycin having immunosuppressive properties, e.g, ABT-281, ASM981; a corticosteroid; cyclophosphamide; azathioprene; methotrexate; leflunomide; mizoribine; mycophenolic acid or salt; mycophenolate mofetil; IL-ip inhibitor.

[0234] In another embodiment, compounds of the invention are combined with a co-agent which are PI3 Kinase inhibitors.

[0235] In another embodiment, compounds of the invention are combined with co-agent that influence BTK (Bruton’s tyrosine kinase).

[0236] For the treatment of oncological diseases, compounds of the invention may be used in combination with B-cell modulating agents, e.g, Rituximab, BTK or Syk inhibitors, inhibitors of PKC, PI3 kinases, PDK, PIM, JAK and mTOR and BH3 mimetics.

[0237] In some embodiments, the compounds of the invention, including salts thereof, can be combined with another immunogenic agent, such as cancerous cells, purified tumor antigens (including recombinant proteins, peptides, and carbohydrate molecules), cells, and cells transfected with genes encoding immune stimulating cytokines. Non-limiting examples of tumor vaccines that can be used include peptides of melanoma antigens, such as peptides of gp 100, MAGE antigens, Trp-2, MARTI and / or tyrosinase, or tumor cells transfected to express the cytokine GM-CSF.

[0238] In some embodiments, the compounds of the invention or salts thereof can also be used in combination with a vaccination protocol for the treatment of cancer. In some embodiments, the tumor cells are transduced to express GM-CSF. In some embodiments, tumor vaccines include the proteins from viruses implicated in human cancers such as Human Papilloma Viruses (HPV), Hepatitis Viruses (HBV and HCV) and Kaposi’s Herpes Sarcoma Virus (KHSV). In some embodiments, the compounds of the present disclosure can be used in combination with tumor specific antigen such as heat shock proteins isolated from tumor tissue itself. In some embodiments, the compounds of the invention or salts thereof can be combined with dendritic cells immunization to activate potent anti-tumor responses.

[0239] In some embodiments, the compounds of the invention can be used in combination with bispecific macrocyclic peptides that target Fe a or Fe y receptor-expressing effectors cells to tumor cells. The compounds of the invention can also be combined with macrocyclic peptides that activate host immune responsiveness.

[0240] In some embodiments, the compounds of the invention can be used in combination with bone marrow transplant for the treatment of a variety of tumors of hematopoietic origin.

[0241] Suitable antiviral agents contemplated for use in combination with the compounds of the invention can comprise nucleoside and nucleotide reverse transcriptase inhibitors (NRTIs), non-nucleoside reverse transcriptase inhibitors (NNRTIs), protease inhibitors and other antiviral drugs. Example suitable NRTIs include zidovudine (AZT); didanosine (ddl); zalcitabine (ddC); stavudine (d4T); lamivudine (3TC); abacavir (1592U89); adefovir dipivoxil [bis(POM)-PMEA]; lobucavir (BMS-180194); BCH-10652; emitricitabine [(-)- FTC]; beta-L-FD4 (also called beta-L-D4C and named beta-L-2’,3’-dicleoxy-5-fluoro- cytidene); DAPD ((-)-beta-D-2, 6, -diamino-purine dioxolane); and lodenosine (FddA). Typical suitable NNRTIs include nevirapine (BI-RG-587); delaviradine (BHAP, U-90152); efavirenz (DMP- 266); PNU-142721; AG-1549; MKC-442 (l-(ethoxy-methyl)-5-(l- methylethyl)-6-(phenylmethyl)-(2,4(lH,3H)-pyrimidinedione); and (+)-calanolide A (NSC- 675451) and B. Typical suitable protease inhibitors include saquinavir (Ro 31-8959); ritonavir (ABT-538); indinavir (MK-639); nelfnavir (AG-1343); amprenavir (141W94); lasinavir (BMS-234475); DMP-450; BMS-2322623; ABT-378; and AG-1549. Other antiviral agents include hydroxyurea, ribavirin, IL-2, IL-12, pentafuside and Yissum Project No.11607. It will be appreciated that many of the further therapeutic agents used in the methods of the invention may be biologicals requiring intravenous, intraperitoneal or depot administration. In a further embodiment, the compound of the invention is orally administered and the further therapeutic agent is administered parenterally, for example intravenously, intraperitoneally or as a depot.

[0242] In any of the combination therapies described herein, when more than one pharmaceutical agent is administered to a patient, they can be administered simultaneously, separately, sequentially, or in combination (e.g., for more than two agents).

[0243] In one embodiment, the invention provides a product comprising a compound of the invention, such as a subject compound or any subgroup thereof, and at least one other therapeutic agent as a combined preparation for simultaneous, separate or sequential use in therapy. Products provided as a combined preparation include a composition comprising the compound of the invention or any subgroup thereof and the other therapeutic agent(s) together in the same pharmaceutical composition, or the subject compound or any subgroup thereof and the other therapeutic agent(s) in separate form, e.g, in the form of a kit.

[0244] In one embodiment, the invention provides a kit comprising two or more separate pharmaceutical compositions, at least one of which contains a subject compound, and another contains a second therapeutic agent discussed herein. In one embodiment, the kit comprises means for separately retaining said compositions, such as a container, divided bottle, or divided foil packet. An example of such a kit is a blister pack, as typically used for the packaging of tablets, capsules and the like. The kit of the invention may be used for administering different dosage forms, for example, oral and parenteral, for administering the separate compositions at different dosage intervals, or for titrating the separate compositions against one another. To assist compliance, the kit of the invention typically comprises directions for administration.

[0245] Pharmaceutical Compositions

[0246] The invention provides pharmaceutical compositions which comprise any one of the compounds described herein, or a pharmaceutically acceptable salt thereof, and one or more pharmaceutically acceptable carriers or excipients.

[0247] “Pharmaceutically acceptable excipient” and “pharmaceutically acceptable carrier” refer to a substance that aids the formulation and / or administration of an active agent to and / or absorption by a subject and can be included in the compositions of the present disclosure without causing a significant adverse toxicological effect on the subject. Non-limiting examples of pharmaceutically acceptable carriers and excipients include water, NaCl, normal saline solutions, lactated Ringer’s, normal sucrose, normal glucose, binders, fillers, disintegrants, lubricants, coatings, sweeteners, flavors, salt solutions (such as Ringer’s solution), alcohols, oils, gelatins, carbohydrates such as lactose, amylose or starch, fatty acid esters, hydroxymethycellulose, polyvinyl pyrrolidine, and colors, and the like. Such preparations can be sterilized and, if desired, mixed with auxiliary agents such as lubricants, preservatives, stabilizers, wetting agents, emulsifiers, salts for influencing osmotic pressure, buffers, coloring, and / or aromatic substances and the like that do not deleteriously react with or interfere with the activity of the compounds provided herein. One of ordinary skill in the art will recognize that other pharmaceutical carriers and excipients are suitable for use with disclosed compounds.

[0248] These compositions optionally further comprise one or more additional therapeutic agents. Alternatively, a compound of the invention may be administered to a patient in need thereof in combination with the administration of one or more other therapeutic regimens (e.g. Gleevec or other kinase inhibitors, interferon, bone marrow transplant, farnesyl transferase inhibitors, bisphosphonates, thalidomide, cancer vaccines, hormonal therapy, antibodies, radiation, etc). For example, additional therapeutic agents for conjoint administration or inclusion in a pharmaceutical composition with a compound of this invention may be another one or more anticancer agents.

[0249] As described herein, the compositions of the present invention comprise a compound of the invention together with a pharmaceutically acceptable carrier, which, as used herein, includes any and all solvents, diluents, or other vehicle, dispersion or suspension aids, surface active agents, isotonic agents, thickening or emulsifying agents, preservatives, solid binders, lubricants and the like, as suited to the particular dosage form desired. Remington’s Pharmaceutical Sciences, Fifteenth Edition, E. W. Martin (Mack Publishing Co., Easton, Pa., 1975) discloses various carriers used in formulating pharmaceutical compositions and known techniques for the preparation thereof. Except insofar as any conventional carrier medium is incompatible with the compounds of the invention, such as by producing any undesirable biological effect or otherwise interacting in a deleterious manner with any other component(s) of the pharmaceutical composition, its use is contemplated to be within the scope of this invention. Some examples of materials which can serve as pharmaceutically acceptable carriers include, but are not limited to, sugars such as lactose, glucose and sucrose; starches such as com starch and potato starch; cellulose and its derivatives such as sodium carboxymethyl cellulose, ethyl cellulose and cellulose acetate; powdered tragacanth; malt; gelatin; talc; excipients such as cocoa butter and suppository waxes; oils such as peanut oil, cottonseed oil; safflower oil; sesame oil; olive oil; corn oil and soybean oil; glycols; such a propylene glycol; esters such as ethyl oleate and ethyl laurate; agar; buffering agents such as magnesium hydroxide and aluminum hydroxide; alginic acid; pyrogen-free water; isotonic saline; Ringer’s solution; ethyl alcohol, and phosphate buffer solutions, as well as other nontoxic compatible lubricants such as sodium lauryl sulfate and magnesium stearate, as well as coloring agents, releasing agents, coating agents, sweetening, flavoring and perfuming agents, preservatives and antioxidants can also be present in the composition.

[0250] Formulations

[0251] This invention also encompasses a class of compositions comprising the active compounds of this invention in association with one or more pharmaceutically-acceptable carriers and / or diluents and / or adjuvants (collectively referred to herein as “carrier” materials) and, if desired, other active ingredients.

[0252] In certain embodiments, the invention provides a pharmaceutical formulation for treating cancer, in particular the cancers described herein, comprising a compound of the present invention or a pharmaceutically acceptable salt thereof together with a pharmaceutically acceptable carrier.

[0253] In certain embodiments, the invention provides a pharmaceutical formulation for treating a cancer selected from the group consisting of breast cancer, colorectal cancer, lung cancer, ovarian cancer, and pancreatic cancer, comprising a compound of the present invention or a pharmaceutically acceptable salt thereof together with a pharmaceutically acceptable carrier.

[0254] The compounds of the present invention may be administered by any suitable route, preferably in the form of a pharmaceutical composition adapted to such a route, and in a dose effective for the treatment intended. The compounds and compositions of the present invention may, for example, be administered orally, mucosally, topically, rectally, pulmonarily such as by inhalation spray, or parentally including intravascularly, intravenously, intraperitoneally, subcutaneously, intramuscularly, transdermally, intraorbitally, intrathecally, intraventricularly, intratumorally, intranasally, intrasternally, by implantation, by inhalation, and by infusion techniques, in dosage unit formulations containing conventional pharmaceutically acceptable carriers, adjuvants, and vehicles.

[0255] Typically, the pharmaceutical compositions are tablets or gelatin capsules comprising the active ingredient together with a) diluents, e.g., lactose, dextrose, sucrose, mannitol, sorbitol, cellulose and / or glycine; b) lubricants, e.g., silica, talcum, stearic acid, its magnesium or calcium salt and / or polyethyieneglycol; for tablets also c) binders, e.g., magnesium aluminum silicate, starch paste, gelatin, tragacanth, methylcellulose, sodium carboxymethylcellulose and / or polyvinylpyrrolidone; if desired d) disintegrants, e.g., starches, agar, alginic acid or its sodium salt, or effervescent mixtures; and / or e) absorbents, colorants, flavors and sweeteners. Tablets may be either film coated or enteric coated according to methods known in the art.

[0256] Suitable compositions for oral administration include an effective amount of a compound of the invention in the form of tablets, lozenges, aqueous or oily suspensions, dispersibie powders or granules, emulsion, hard or soft capsules, or syrups or elixirs. Compositions intended for oral use are prepared according to any method known in the art for the manufacture of pharmaceutical compositions and such compositions can contain one or more agents selected from the group consisting of sweetening agents, flavoring agents, coloring agents and preserving agents in order to provide pharmaceutically elegant and palatable preparations. Tablets may contain the active ingredient in admixture with nontoxic pharmaceutically acceptable excipients which are suitable for the manufacture of tablets. These excipients are, for example, inert diluents, such as calcium carbonate, sodium carbonate, lactose, calcium phosphate or sodium phosphate; granulating and disintegrating agents, for example, com starch, or alginic acid; binding agents, for example, starch, gelatin or acacia; and lubricating agents, for example magnesium stearate, stearic acid or talc. The tablets are uncoated or coated by known techniques to delay disintegration and absorption in the gastrointestinal tract and thereby provide a sustained action over a longer period. For example, a time delay material such as glyceryl monostearate or glyceryl distearate can be employed. Formulations for oral use can be presented as hard gelatin capsules wherein the active ingredient is mixed with an inert solid diluent, for example, calcium carbonate, calcium phosphate or kaolin, or as soft gelatin capsules wherein the active ingredient is mixed with water or an oil medium, for example, peanut oil, liquid paraffin or olive oil.

[0257] Certain injectable compositions are aqueous isotonic solutions or suspensions, and suppositories are advantageously prepared from fatty emulsions or suspensions. Said compositions may be sterilized and / or contain adjuvants, such as preserving, stabilizing, wetting or emulsifying agents, solution promoters, salts for regulating the osmotic pressure and / or buffers. In addition, they may also contain other therapeutically valuable substances. Said compositions are prepared according to conventional mixing, granulating or coating methods, respectively, and contain about 0.1-75%, or contain about 1-50%, of the active ingredient. Suitable compositions for transdermal application include an effective amount of a compound of the invention with a suitable carrier. Carriers suitable for transdermal delivery include absorbable pharmacologically acceptable solvents to assist passage through the skin of the host. For example, transdermal devices are in the form of a bandage comprising a backing member, a reservoir containing the compound optionally with carriers, optionally a rate controlling barrier to deliver the compound of the skin of the host at a controlled and predetermined rate over a prolonged period of time, and means to secure the device to the skin.

[0258] Suitable compositions for topical application, e.g., to the skin and eyes, include aqueous solutions, suspensions, ointments, creams, gels or sprayable formulations, e.g., for delivery by aerosol or the like. Such topical delivery systems will in particular be appropriate for dermal application, e.g., for the treatment of skin cancer, e.g., for prophylactic use in sun creams, lotions, sprays and the like. They are thus particularly suited for use in topical, including cosmetic, formulations well-known in the art. Such may contain solubilizers, stabilizers, tonicity enhancing agents, buffers and preservatives.

[0259] As used herein a topical application may also pertain to an inhalation or to an intranasal application. They may be conveniently delivered in the form of a dry powder (either alone, as a mixture, for example a dry blend with lactose, or a mixed component particle, for example with phospholipids) from a dry powder inhaler or an aerosol spray presentation from a pressurized container, pump, spray, atomizer or nebulizer, with or without the use of a suitable propellant.

[0260] The present invention further provides anhydrous pharmaceutical compositions and dosage forms comprising the compounds of the present invention as active ingredients, since water may facilitate the degradation of certain compounds.

[0261] Anhydrous pharmaceutical compositions and dosage forms of the invention can be prepared using anhydrous or low moisture containing ingredients and low moisture or low humidity conditions. An anhydrous pharmaceutical composition may be prepared and stored such that its anhydrous nature is maintained. Accordingly, anhydrous compositions are packaged using materials known to prevent exposure to water such that they can be included in suitable formulary kits. Examples of suitable packaging include, but are not limited to, hermetically sealed foils, plastics, unit dose containers (e.g., vials), blister packs, and strip packs.

[0262] The invention further provides pharmaceutical compositions and dosage forms that comprise one or more agents that reduce the rate by which the compound of the present invention as an active ingredient will decompose. Such agents, which are referred to herein as “stabilizers,” include, but are not limited to, antioxidants such as ascorbic acid, pH buffers, or salt buffers, etc.

[0263] The pharmaceutically active compounds of this invention can be processed in accordance with conventional methods of pharmacy to produce medicinal agents for administration to patients, including humans and other mammals.

[0264] The amount of compounds which are administered and the dosage regimen for treating a disease condition with the compounds and / or compositions of this invention depends on a variety of factors, including the age, weight, sex and medical condition of the subject, the type of disease, the severity of the disease, the route and frequency of administration, and the particular compound employed. Thus, the dosage regimen may vary widely, but can be determined routinely using standard methods. As mentioned previously, the daily dose can be given in one administration or may be divided between 2, 3, 4 or more administrations.

[0265] For therapeutic purposes, the active compounds of this invention are ordinarily combined with one or more adjuvants, excipients or carriers appropriate to the indicated route of administration. If administered per os, the compounds may be admixed with lactose, sucrose, starch powder, cellulose esters of alkanoic acids, cellulose alkyl esters, talc, stearic acid, magnesium stearate, magnesium oxide, sodium and calcium salts of phosphoric and sulfuric acids, gelatin, acacia gum, sodium alginate, polyvinylpyrrolidone, and / or polyvinyl alcohol, and then tableted or encapsulated for convenient administration. Such capsules or tablets may contain a controlled-release formulation as may be provided in a dispersion of active compound in hydroxypropylmethyl cellulose.

[0266] In the case of skin conditions, it may be preferable to apply a topical preparation of compounds of this invention to the affected area two to four times a day. Formulations suitable for topical administration include liquid or semi-liquid preparations suitable for penetration through the skin (e.g., liniments, lotions, ointments, creams, or pastes) and drops suitable for administration to the eye, ear, or nose. For topical administration, the active ingredient may comprise from 0.001% to 10% w / w, e.g., from 1% to 2% by weight of the formulation, although it may comprise as much as 10% w / w, but preferably not more than 5% w / w, and more preferably from 0.1% to 1% of the formulation.

[0267] The compounds of this invention can also be administered by a transdermal device. Preferably transdermal administration will be accomplished using a patch either of the reservoir and porous membrane type or of a solid matrix variety. In either case, the active agent is delivered - continuously from the reservoir or microcapsules through a membrane into the active agent permeable adhesive, which is in contact with the skin or mucosa of the recipient. If the active agent is absorbed through the skin, a controlled and predetermined flow of the active agent is administered to the recipient. In the case of microcapsules, the encapsulating agent may also function as the membrane. The oily phase of the emulsions of this invention may be constituted from known ingredients in a known manner.

[0268] While the phase may comprise merely an emulsifier, it may comprise a mixture of at least one emulsifier with a fat or an oil or with both a fat and an oil. Preferably, a hydrophilic emulsifier is included together with a lipophilic emulsifier which acts as a stabilizer. It is also preferred to include both an oil and a fat. Together, the emulsifier(s) with or without stabilizer(s) make-up the so-called emulsifying wax, and the wax together with the oil and fat make up the so-called emulsifying ointment base which forms the oily dispersed phase of the cream formulations. Emulsifiers and emulsion stabilizers suitable for use in the formulation of the present invention include Tween 60, Span 80, cetostearyl alcohol, myristyl alcohol, glyceryl monostearate, sodium lauryl sulfate, glyceryl distearate alone or with a wax, or other materials well known in the art.

[0269] The choice of suitable oils or fats for the formulation is based on achieving the desired cosmetic properties, since the solubility of the active compound in most oils likely to be used in pharmaceutical emulsion formulations is very low. Thus, the cream should preferably be a non-greasy, non-staining and washable product with suitable consistency to avoid leakage from tubes or other containers. Straight or branched chain, mono- or dibasic alkyl esters such as di-isoadipate, isocetyl stearate, propylene glycol diester of coconut fatty acids, isopropyl myristate, decyl oleate, isopropyl palmitate, butyl stearate, 2-ethylhexyl palmitate or a blend of branched chain esters may be used. These may be used alone or in combination depending on the properties required.

[0270] Alternatively, high melting point lipids such as white soft paraffin and / or liquid paraffin or other mineral oils can be used. Formulations suitable for topical administration to the eye also include eye drops wherein the active ingredients are dissolved or suspended in suitable carrier, especially an aqueous solvent for the active ingredients.

[0271] The active ingredients are preferably present in such formulations in a concentration of 0.5 to 20%, advantageously 0.5 to 10% and particularly about 1.5% w / w.

[0272] Formulations for parenteral administration may be in the form of aqueous or nonaqueous isotonic sterile injection solutions or suspensions. These solutions and suspensions may be prepared from sterile powders or granules using one or more of the carriers or diluents mentioned for use in the formulations for oral administration or by using other suitable dispersing or wetting agents and suspending agents. The compounds may be dissolved in water, polyethylene glycol, propylene glycol, ethanol, com oil, cottonseed oil, peanut oil, sesame oil, benzyl alcohol, sodium chloride, tragacanth gum, and / or various buffers. Other adjuvants and modes of administration are well and widely known in the pharmaceutical art. The active ingredient may also be administered by injection as a composition with suitable carriers including saline, dextrose, or water, or with cyclodextrin (i.e. Captisol), cosolvent solubilization (i.e. propylene glycol) or micellar solubilization ( / '.< . Tween 80).

[0273] The sterile injectable preparation may also be a sterile injectable solution or suspension in a non-toxic parenterally acceptable diluent or solvent, for example as a solution in 1,3-butanediol. Among the acceptable vehicles and solvents that may be employed are water, Ringer’s solution, and isotonic sodium chloride solution. In addition, sterile, fixed oils are conventionally employed as a solvent or suspending medium. For this purpose any bland fixed oil may be employed, including synthetic mono- or diglycerides. In addition, fatty acids such as oleic acid find use in the preparation of injectables.

[0274] For pulmonary administration, the pharmaceutical composition may be administered in the form of an aerosol or with an inhaler including dry powder aerosol.

[0275] Suppositories for rectal administration of the drug can be prepared by mixing the drug with a suitable nonirritating excipient such as cocoa butter and polyethylene glycols that are solid at ordinary temperatures but liquid at the rectal temperature and will therefore melt in the rectum and release the drug.

[0276] The pharmaceutical compositions may be subjected to conventional pharmaceutical operations such as sterilization and / or may contain conventional adjuvants, such as preservatives, stabilizers, wetting agents, emulsifiers, buffers etc. Tablets and pills can additionally be prepared with enteric coatings. Such compositions may also comprise adjuvants, such as wetting, sweetening, flavoring, and perfuming agents. Pharmaceutical compositions of this invention comprise a compound of the formulas described herein or a pharmaceutically acceptable salt thereof; an additional agent selected from a kinase inhibitory agent (small molecule, polypeptide, antibody, efc.), an immunosuppressant, an anticancer agent, an anti-viral agent, anti-inflammatory agent, antifungal agent, antibiotic, or an anti- vascular hyperproliferation compound; and any pharmaceutically acceptable carrier, adjuvant or vehicle.

[0277] Alternate compositions of this invention comprise a compound of the formulae described herein or a pharmaceutically acceptable salt thereof; and a pharmaceutically acceptable carrier, adjuvant or vehicle. Such compositions may optionally comprise one or more additional therapeutic agents, including, for example, kinase inhibitory agents (small molecule, polypeptide, antibody, efc.), immunosuppressants, anti-cancer agents, anti-viral agents, anti-inflammatory agents, antifungal agents, antibiotics, or anti-vascular hyperproliferation compounds.

[0278] The term “pharmaceutically acceptable carrier or adjuvant” refers to a carrier or adjuvant that may be administered to a patient, together with a compound of this invention, and which does not destroy the pharmacological activity thereof and is nontoxic when administered in doses sufficient to deliver a therapeutic amount of the compound. Pharmaceutically acceptable carriers, adjuvants and vehicles that may be used in the pharmaceutical compositions of this invention include, but are not limited to, ion exchangers, alumina, aluminum stearate, lecithin, self emulsifying drug delivery systems (SEDDS) such as d-atocopherol polyethyleneglycol 1000 succinate, surfactants used in pharmaceutical dosage forms such as Tweens or other similar polymeric delivery matrices, serum proteins, such as human serum albumin, buffer substances such as phosphates, glycine, sorbic acid, potassium sorbate, partial glyceride mixtures of saturated vegetable fatty acids, water, salts or electrolytes, such as protamine sulfate, disodium hydrogen phosphate, potassium hydrogen phosphate, sodium chloride, zinc salts, colloidal silica, magnesium trisilicate, polyvinyl pyrrolidone, cellulose-based substances, polyethylene glycol, sodium carboxymethylcellulose, polyacrylates, waxes, polyethylene-polyoxypropylene-block polymers, polyethylene glycol and wool fat. Cyclodextrins such as u-, P-, and y- cyclodextrin, or chemically modified derivatives such as hydroxyalkylcyclodextrins, including 2 and 3-hydroxypropyl-cyclodextrins, or other solubilized derivatives may also be advantageously used to enhance delivery of compounds of the formulae described herein.

[0279] The pharmaceutical compositions may be orally administered in any orally acceptable dosage form including, but not limited to, capsules, tablets, emulsions and aqueous suspensions, dispersions and solutions. In the case of tablets for oral use, carriers which are commonly used include lactose and corn starch. Lubricating agents, such as magnesium stearate, are also typically added. For oral administration in a capsule form, useful diluents include lactose and dried com starch. When aqueous suspensions and / or emulsions are administered orally, the active ingredient may be suspended or dissolved in an oily phase is combined with emulsifying and / or suspending agents.

[0280] If desired, certain sweetening, flavoring and / or coloring agents may be added. The pharmaceutical compositions may comprise formulations utilizing liposome or microencapsulation techniques, various examples of which are known in the art.

[0281] The pharmaceutical compositions may be administered by nasal aerosol or inhalation. Such compositions are prepared according to techniques well known in the art of pharmaceutical formulation and may be prepared as solutions in saline, employing benzyl alcohol or other suitable preservatives, absorption promoters to enhance bioavailability, fluorocarbons, and / or other solubilizing or dispersing agents, examples of which are also well known in the art.

[0282] Treatment Kits

[0283] One aspect of the present invention relates to a kit for conveniently and effectively carrying out the methods or uses in accordance with the present invention. In general, the pharmaceutical pack or kit comprises one or more containers filled with one or more of the ingredients of the pharmaceutical compositions of the invention. Such kits are especially suited for the delivery of solid oral forms such as tablets or capsules. Such a kit preferably includes a number of unit dosages, and may also include a card having the dosages oriented in the order of their intended use. If desired, a memory aid can be provided, for example in the form of numbers, letters, or other markings or with a calendar insert, designating the days in the treatment schedule in which the dosages can be administered. Optionally associated with such container(s) can be a notice in the form prescribed by a governmental agency regulating the manufacture, use or sale of pharmaceutical products, which notice reflects approval by the agency of manufacture, use or sale for human administration. The following representative examples contain important additional information, exemplification and guidance which can be adapted to the practice of this invention in its various embodiments and the equivalents thereof. These examples are intended to help illustrate the invention, and are not intended to, nor should they be construed to, limit its scope. Indeed, various modifications of the invention, and many further embodiments thereof, in addition to those shown and described herein, will become apparent to those skilled in the art upon review of this document, including the examples which follow and the references to the scientific and patent literature cited herein.

[0284] The contents of the cited references are incorporated herein by reference to help illustrate the state of the art.

[0285] In addition, for purposes of this invention, the chemical elements are identified in accordance with the Periodic Table of the Elements, CAS version, Handbook of Chemistry and Physics, 75thEd., inside cover. Additionally, general principles of organic chemistry, as well as specific functional moieties and reactivity, are described in “Organic Chemistry,” Thomas Sorrell, University Science Books, Sausalito: 1999, and “Organic Chemistry,” Morrison & Boyd (3d Ed), the entire contents of both of which are incorporated herein by reference.

[0286] EXPERIMENTAL

[0287] Abbreviations:

[0288] Abbreviation Solvent Abbreviation Solvent

[0289] A AActive pharmaceutical

[0290] ACN Acetonitrile API / ingredient

[0291] Fasted-state simulated

[0292] DCM Dichloromethane FaSSIF . . . intestinal fluid

[0293] Fed-state simulated

[0294] Tol Toluene FeSSIF . . , , intestinal fluid

[0295] Hept n-Heptane RH Relative humidity

[0296] EA Ethyl Acetate FA Formic Acid

[0297] IPA 2-Propanol RT Room temperature

[0298] EtOH Ethanol SGF Simulated gastric fluid

[0299] MEK 2-Butanone MeOAc Methyl Acetate

[0300] MeOH Methanol DMSO Dimethylsulfoxide

[0301] MTBE tert-Butyl Methyl Ether NMP N-Methyl Pyrrolidone

[0302] / -BuOH tert-butanol TFA Trifluoroacetic Acid

[0303] IPA 2-propanol H2O Water

[0304] IP AC Isopropyl acetate THF Tetrahydrofuran Instruments

[0305] Full Name Abbreviation

[0306] Differential scanning calorimetry DSC

[0307] Dynamic Vapor Sorption DVS

[0308] High Performance Liquid Chromatography HPLC

[0309] Liquid Chromatography-Mass Spectrometry LCMS

[0310] Nuclear Magnetic Resonance NMR

[0311] Polarized Light Microscopy PLM

[0312] X-ray Powder Diffraction XRPD

[0313] Thermogravimetric Analysis TGA

[0314] Infrared Spectroscopy IR

[0315] Units

[0316] Full Name Abbreviation

[0317] Celsius C

[0318] Degrees0

[0319] Equivalents eq.

[0320] Gram g

[0321] Hour H

[0322] Hertz (S'1) Hz

[0323] Milligrams mg

[0324] Milliliters mL

[0325] Minute min

[0326] Relative Humidity RH

[0327] Room temperature RT

[0328] Second sec volume vol.

[0329] Volume ratio v / v

[0330] Weight wt.

[0331] Weight Percentage wt.%

[0332] Analysis Conditions X-ray Powder Diffraction (XRPD)

[0333] XRPD patterns were identified with an X-ray diffractometer (Rigaku Smartlab SE) using parameters as defined in Table 8.

[0334] Thermogravimetric Analysis (TGA) TGA was carried out on a TA, TGA 500 (TA Instruments, US) using parameters as defined in Table 8.

[0335] Differential Scanning Calorimeter (DSC) DSC was performed using a TA, DSC2500 (TA Instruments, US) using parameters as defined in Table 8.

[0336] Polarized Light Microscopy (PLM)

[0337] Light microscopy was performed using a Polarizing Microscope ECLIPSE LVIOOPOL (Nikon, JPN) using parameters as defined in Table 8.

[0338] Dynamic Vapor Sorption (DVS)

[0339] Moisture sorption / desorption data was collected on an Adventure DVS using parameters as defined in Table 8.

[0340] High Pressue Liquid Chromatography (HPLC)

[0341] HPLC was performed using an Agilent 1260 using parameters as defined in Table 8.

[0342] Table 8. HPLC Method for Stability Testing

[0343] Example 1: Experimental Procedures and Preparation of Crystalline Salt Forms of

[0344] Compound (I)

[0345] General Procedure 1: Estimated Solubility Measurement 10 mg of Compound (I) was weighed into a vial. 0.1 mL of the solvent was added and the vial was shaken vigorously for 1 min and placed in a constant temperature device, maintained at a temperature of 25.0 ± 0.5 °C, 50.0 ± 0.5 °C for 15 min. If the substance was not completely dissolved, more solvent was added, then repeated the shaking for 1 min and placed the tube in the constant temperature device for 15 min.

[0346] The results of the estimated solubility measurement are outlined in Table 9.

[0347] Table 9. Estimated Solubility of the Compound (I) (S: mg / mL)

[0348] General Procedure 2: Salt Screening

[0349] 30 mg of Compound (I) and 1 mL of solvent were added into a glass vial and stirred until clear or suspension under RT. 1.1 eq of the the counter-ion was prepared in 0.5 mL of the respective solvent, and then added into the glass vial containing Compound (I) dropwise. The vial was heated to 50°C for 2 h then cooled to RT over 3 h. Any resulting suspension was filted and the solid was collected for XRPD analysis. The remaining filtrate was allowed to evaporate. If a clear solution was obtained, direct evaporation was conducted for collecting a solid for XRPD test. The results of the salt screening are shown in Table 10.

[0350] Table 10. Results of Salt Screening Experiments.

[0351] Characterization of Salt Forms

[0352] The salt forms obtained from salt screening were characterized by differential scanning calorimetry (DSC), thermogravimetric analysis (TGA), and X-ray powder diffraction (XRPD) and polarized light microscopy (PLM). The solid state properties of the salt forms have been shown in Table 11.

[0353] Table 11. Results of Solid State Characterization of Salt Forms

[0354] Example 2: Solubility, Hygroscopicity, and Stability Studies of Phosphate hydrate salt Form A, Maleate salt Form Al and Sulfate salt Form A of Compound (I)

[0355] Solubility Testing

[0356] 20 mg samples of maleate Form A, phosphate hydrate-Form A, and sulfate Form A were accurately weighed and then transferred into 10 mL screw vials. 5 mL of vehicle solutions (water, PH buffer, SGF, FaSSIF, FeSSIF) were separately added into the vials. The vials were sealed and transferred into a constant temperature shaker under 25°C with a shake rate of 150 rpm for 48 h. The vials were checked during the testing and more salt was added if the suspension turned into solution. After 48 hours, about 1 mL of each suspension was filtered and the filtrate was collected for HPLC analysis to test the solubility. The filter cake was analyzed by XRPD. The pH value of the filtrate samples was measured by pH meter. The results of the solubility testing are shown in Table 12.

[0357] Table 12. Results of Solubility Testing of Salt Forms

[0358] Hygroscopicity Studies

[0359] Around 20-50 mg samples of maleate salt Form A, sulfate salt Form A and phosphate hydrate salt Form A were weighed into the sample pan. The sample was subjected to the DVS program as 0%-90%-0% two cycles, the step of P / Po was 10% and dm / dt<0.002.

[0360] Results:

[0361] Maleate salt Form A demonstrated slight moisture absorption, 1.8% at RH=90%. However, the crystalline form did not change during the testing.

[0362] Phosphate hydrate salt Form A demonstrated slight moisture absorption, 3.2% at RH=90%.

[0363] However, the crystalline form did not change during the testing.

[0364] Sulfate salt Form A demosntrated relatively strong moisture absorption, 9% at RH=90%. However, the crystalline formdid not change during the testing. Solid Bulk Stability Testing

[0365] 20 mg samples of maleate Form A, phosphate hydrate-Form A, and sulfate Form A were accurately weighed respectively and then was placed in glass petri dishes. The petri dishes were transferred into the constant temperature and humidity box with different environments: illumination: 5000±500 Lux, high temperature: 60°C (closed), high relative humidity (RH): 25°C, 90 + 5% (open) and tested periodically at 5 days and 11 -days storage. Samples were tested by XRPD, DSC, and assayed for impurities using a high performance liquid chromatography technique (HPLC) method. The results of the solubility testing are shown in Table 13.

[0366] Table 13. Results of Stability Testing of Salt Forms

[0367] Example 3: Stability Studies, Stoichiometry, and Absolute Configuration of Phosphate hydrate salt Form A of Compound (I)

[0368] Suspension Stability Testing HPMC and Tween 80 were weighed in to the 20 mL glass vials respectively for preparation of excipients solution (1 % w / w). The excipients were under ultrasound and stirring until the solution was clear. Phosphate hydrate Form A was weighed into a 50 mL glass beaker, the Tween 80 solution was firstly added into the beaker under stirring with a speed of 500 rpm. The HPMC solution was added into the beaker to form a suspension under stirring.

[0369] The suspension was kept stirring for 2 h at 25°C to ensure the suspension was uniform. The suspension was transferred into a sealed screw-top bottle and then put into a constant temperature which was protected from light. The solution and solid were collected to compare the purity and XRPD patterns of the stressed sample to an unstressed sample. The results of the solubility testing are shown in Table 14.

[0370] Table 14. Results of Suspension Stability Testing of Phosphate Hydrate salt Form A

[0371] Stoichiometry Studies

[0372] The stoichiometry (base / acid) of the phosphate hydrate salt Form A of Compound (I) was quantified using IC. The results are summarized in Table 15.

[0373] Table 15. Results of Stoichiometry Testing of Phosphate Hydrate salt Form A

[0374] The calculated content of phosphate group was 18.7%, which meants that the stoichiometric ratio of salt formation is 1.

[0375] Determination of Absolute Configuration

[0376] The single crystal of phosphate hydrate salt Form A was prepared by vapor diffusion. The solution was 6 mg / mL prepared in ACN:H2O (1 : 1, v / v) and the vapor phase was Acetone.

[0377] The crystal was obtained as a columnar crystal and characterized by Bruker single crystal X- ray diffractometer. The characterization results show that the single crystal belongs to space group Pl and is a phosphate hemihydrate. The absolute configuration of the chiral center of Compound (I) is S configuration. The crystal structure is shown in Figure 8.

[0378] Conclusion

[0379] Salt screening was performed using 17 types of counter-ions in 6 solvent systems. According to preliminary characterization of thermal stability and crystallinity, the maleate, sulfate, phosphate were scaled up and characterized for further studies. The stability studies of the maleate, sulfate, phosphate salts were performed under stress condition at high temperature (60°C), high humidity (92.5%) and illumination over 11 days., The solubility, suspension stability, and hygroscopicity were also tested for the phosphate hydrate salt Form A. Overall, phosphate hydrate salt Form A showed good stability and favorable properties. Procedures and Results

[0380] General Procedure 3: Equilibration with solvent at 25 °C

[0381] About 50 mg of Compound (I) is equilibrated with 1 ml solvent for at least 24 h at 25°C. Then, the solutions were filtered and solid was air-dried to remove residual excess solvent. The solid was analyzed by XRPD. If differences were observed additional investigations would be performed. The results of the equilibration with solvent at 25 °C are summarized in Table 16.

[0382] Table 16. Results of Equilibration with Solvent at 25 °C

[0383] General Procedure 4: Equilibration with solvent at 25 °C

[0384] This study was perfomed following the procedure in general procedure 3. but at 50 °C ± 0.5.

[0385] The hot filtrate was used for determination of the solubility at 50 °C. The results of the equilibration with solvent at 50 °C are summarized in Table 17.

[0386] Table 17. Results of Equilibration with Solvent at 50 °C

[0387] General Procedure 5: Slow Evaporation at 25 °C The filtrate from general procedure 3 was used for evaporation (slow). Samples were slowly evaporated to dryness by nitrogen flow at room temperature. The sample was collected prior to complete dryness and examined by XRPD. The results of the slow evaporation at 25 °C are summarized in Table 18.

[0388] Table 18. Results of Slow Evaporation at 25 °C

[0389] General Procedure 6: Fast Evaporation at 50 °C

[0390] The solutions were prepared at 50 °C and allowed to evaporate at 50 °C (fast evaporation).

[0391] Samples were collected prior to complete dryness and examined by XRPD and other methods as needed. The results of the fast evaporation at 25 °C are summarized in Table 19.

[0392] Table 19. Results of Slow Evaporation at 25 °C General Procedure 7: Precipitation by Addition of Anti-solvent

[0393] Two different solvent combinations were tested. Compound (I) was dissolved in a medium where the solubility was high, and an anti-solvent in which the substance was highly insoluble was added. The results are summarized in Table 20.

[0394] Table 20. Results of Precipitation by Addition of Antisolvent

[0395] General Procedure 8: Behavior under Compression

[0396] 300 mg of Compound (I) were compressed for 5 minutes at 10 MPa with a hydraulic press (diameter of the tablets 13 mm).

[0397] Results:

[0398] This solid obtained from this experiment was analyzed by XRPD and determined to be Form A.

[0399] General Procedure 9: Granulation Simulation Experiments

[0400] Added granulating solvent dropwise to Compound (I) until solid was wetted sufficiently.

[0401] Evaluate solid form and degree of crystallinity by e.g., XRPD and / or DSC if change was detected. The objective was to determine whether polymorphs / solvatemorphs or amorphous phase were readily produced. Results are shown in Table 21.

[0402] Table 21. Results of Anti-solvent Precipitation. Characterization and Definition of Form A

[0403] Form A was the only crystalline form obtained from polymorph studies performed above The XRPD pattern was named as Form A (Figure 7A). The solid state properties are summarized in Table 22.

[0404] Table 22. Solid state Characterization of Form A

[0405] Example 5: Solubiliy, Hygroscopicity, and Stability Studies of Crystalline Form A of Compound (I)

[0406] Solubility Studies

[0407] About 50 mg of crystalline form A of Compound (I) into was weighed into 10 mL vials, then 5 mL of each media (water, buffer solution and simulated physiological solution) was added into the vials respectively. The vials were transferred into the constant temperature and humidity chamber, and shaken for 48 hat 150 rpm under 25°C. The vials were checked during the testing and more crystalline Form A was added if the suspension turned into solution. After 48 hours, the obtained suspension was filtered, and the filtrate was tested by HPLC. The pH of the filtrate was tested using a pH meter and the filter cake was collected and analyzed by XRPD. The results of the solubility testing are shown in Table 23.

[0408] Table 23. Results of Solubility Testing of Salt Forms Hygroscopicity Testing

[0409] Around 20-50 mg samples of crystalline Form A was weighed into the sample pan. The sample was subjected to the DVS program as 0%-90%-0% two cycles, the step of P / Po was 10% and dm / dt<0.002.

[0410] Results:

[0411] Crystalline Form A demonstrated slight moisture absorption, 1.8% at RH=90%. However, the crystalline form did not change during the testing.

[0412] Solid Bulk Stability Testing

[0413] 20 mg samples of crystalline Form A were accurately weighed then placed in glass petri dishes. The petri dishes were transferred into the constant temperature and humidity box with different environments: illumination: 5000±500 Lux, high temperature: 60°C ( closed), high relative humidity (RH): 25°C, 90 + 5% ( open) and tested periodically during 5 / 11- days storage. Samples were tested by XRPD, DSC, and assayed for impurities using a high performance liquid chromatography technique (HPLC) method. The results of the solubility testing are shown in Table 24.

[0414] Table 24. Results of Stability Testing of Salt Forms

[0415] Suspension Stability Testing

[0416] HPMC and Tween 80 were weighed in to the 20 mL glass vials respectively for preparation of excipients solution (1 % w / w). The excipients were under ultrasound and stirred until the solution was clear. Crystalline Form A of Compound (I) was weighed into a 50 mL glass beaker, the Tween 80 solution was firstly added into the beaker under stirring with a speed of 500 rpm. The HPMC solution was added into the beaker to form a suspension under stirring. The suspension was kept stirring for 2 h at 25°C to ensure the suspension was uniform. The suspension was transferred into a sealed screw-top bottle and then put into a constant temperature which was protected from light. The solution and solid were collected to compare the purity and the XRPD pattern of the stressed sample to an unstressed sample. The results of the solubility testing are shown in Table 25.

[0417] Table 25. Results of Suspension Stability Testing of Crystalline Form A Conclusion

[0418] Polymorph screening was performed in 19 solvents using 7 kinds of crystallization methods. Crystalline Form A was the only polymorph form obtained and characterized. The physicochemical stability, solubility, dynamic hygroscopicity of crystalline Form A were invetstigated. Overall, Form A demonstrated good solid state properties.

Claims

CLAIMS1. A phosphate hydrate salt of Compound (I):

2. The phosphate hydrate salt of Compound (I) according to claim 1, wherein the molar ratio of Compound (I) to phosphoric acid to water is 1 to 1 to 0.5.

3. The phosphate hydrate salt of Compound (I) according to claim 1 or 2, wherein the phosphate hydrate salt is crystalline.

4. The phosphate hydrate salt of Compound (I) according to any one of claims 1 to 3, wherein the phosphate hydrate salt is crystalline Form A characterized by an X-ray powder diffraction pattern which comprises peaks at 13.8°, 14.8°, 20.6°, and 24.1° ± 0.2 in 20.

5. The phosphate hydrate salt of Compound (I) according to any one of claims 1 to 3, wherein the phosphate hydrate salt is crystalline Form A characterized by an X-ray powder diffraction pattern which comprises at least five, six, seven or eight peaks chosen from 8.1°, 12.6°, 13.8°, 14.8°, 17.1°, 20.6°, 24.1°, and 24.5° ± 0.2 in 20.

6. The phosphate hydrate salt of Compound (I) according to any one of claims 1 to 3, wherein the phosphate hydrate salt is crystalline Form A characterized by an X-ray powder diffraction pattern which comprises peaks at 6.2°, 8.1°, 12.6°, 13.8°, 14.8°, 17.1°, 18.7°, 20.6°, 23.4°, 24.1°, 24.5°, and 29.6° ± 0.2 in 20.

7. The phosphate hydrate salt of Compound (I) according to any one of claims 1 to 6, characterized by a differential scanning calorimeter (DSC) peak phase transition temperature of 228.8 ± 2 °C.

8. A crystalline Form A of maleate salt of Compound (I):wherein the molar ratio of Compound (I) to maleic acid is 1 to 1, characterized by an X-ray powder diffraction pattern which comprises peaks at 8.5°, 11.3°, 17.2°, and 17.8° ± 0.2 in 26.

9. The crystalline Form A of maleate salt of Compound (I) according to claim 8, characterized by an X-ray powder diffraction pattern which comprises at least five, six, or seven peaks chosen from 8.5°, 8.8°, 11.3°, 17.2°, 17.8°, 22.0°, and 27.2° ± 0.2 in 26.

10. The crystalline Form A of maleate salt of Compound (I) according to claim 8, characterized by an X-ray powder diffraction pattern which comprises peaks at 8.5°, 8.8°, 11.3°, 15.7°, 17.2°, 17.8°, 22.0°, 23.0°, 26.0°, and 27.2° ± 0.2 in 26.

11. The crystalline Form A of maleate salt of Compound (I) according to any one of claims 8 to 10, characterized by a differential scanning calorimeter (DSC) peak phase transition temperature of 156.9 ± 2 °C.

12. A crystalline Form A of hydrochloride salt of Compound (I):wherein the molar ratio of Compound (I) to hydrochloric acid is 1 to 1, characterized by an X-ray powder diffraction pattern which comprises peaks at 6.5°, 13.3°, 19.7°, and 23.8° ± 0.2 in 26.

13. The crystalline Form A of hydrochloride salt of Compound (I) according to claim 12, characterized by an X-ray powder diffraction pattern which comprises at least five, six, seven, or eight peaks chosen from 6.5°, 13.3°, 15.7°, 19.7°, 22.2°, 23.8°, 24.2°, and 28.6° ± 0.2 in 29.

14. The crystalline Form A of hydrochloride salt of Compound (I) according to claim 12, characterized by an X-ray powder diffraction pattern which comprises peaks at 6.5°, 7.8°, 13.3°, 14.0°, 15.7°, 16.8°, 19.7°, 22.2°, 23.5°, 23.8°, 24.2°, and 28.6° ± 0.2 in 29.

15. The crystalline Form A of hydrochloride salt of Compound (I) according to any one of claims 12 to 14, characterized by a differential scanning calorimeter (DSC) peak phase transition temperature of 120.2 ± 2 °C, 236.4 ± 2 °C, and 268.4 ± 2 °C.

16. A crystalline Form A of Compound (I):characterized by an X-ray powder diffraction pattern which comprises peaks at 10.8°, 15.4°, 18.1°, and 25.1° ± 0.2 in 29.

17. The crystalline Form A of Compound (I) according to claim 16, characterized by an X-ray powder diffraction pattern which comprises at least five, six, seven, eight, nine or ten peaks chosen from 10.8°, 13.2°, 15.4°, 18.1°, 18.6°, 19.9°, 21.8°, 22.8°, 24.0°, and 25.1° ± 0.2 in 29.

18. The crystalline Form A of Compound (I) according to claim 16, characterized by an X-ray powder diffraction pattern which comprises peaks at 10.8°, 11.8°, 13.2°, 15.4°, 18.1°, 18.6°, 19.9°, 20.4°, 21.8°, 22.8°, 24.0°, and 25.1° ± 0.2 in 29.

19. The crystalline Form A of Compound (I) according to any one of claims 16 to 18, characterized by a differential scanning calorimeter (DSC) peak phase transition temperature of 213.1 ± 2 °C.

20. A pharmaceutical composition comprising the compound of any one of claims 1 to 19, and a pharmaceutically acceptable carrier.

21. A combination comprising a therapeutically effective amount of the compound of any one of claims 1 to 19, and one or more therapeutically active co-agents.

22. A method for treating a subject with cancer, comprising administering to the subject an effective amount of a compound of any one of claims 1 to 19, or a pharmaceutically acceptable salt thereof, the pharmaceutical composition of claim 20, or the combination of claim 21.

23. The method of claim 22, wherein the cancer is selected from breast cancer, colorectal cancer, lung cancer, ovarian cancer, and pancreatic cancer.

24. A method of inhibiting HPK1 activity in a subject in need thereof, said method comprising administering to the subject an effective amount of the compound of any one of claims 1 to 19, or a pharmaceutically acceptable salt thereof, the pharmaceutical composition of claim 20, or the combination of claim 21.

25. The method of claim 24, wherein the subject has a cancer, and wherein the cancer is treated.

26. The method of claim 25, wherein the cancer is selected from breast cancer, colorectal cancer, lung cancer, ovarian cancer, and pancreatic cancer.

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