WO2022056572A1 - Treatment of solid tumours - Google Patents

Abstract

The present invention relates to a method for the treatment of cancer, in particular colorectal cancer (colon cancer), comprising administering an effective amount of dasatinib to the subject; and administering an effective amount of an immune checkpoint inhibitor to the subject. Preferably said immune checkpoint inhibitor is selected from the group consisting of a PD-1 inhibitor, a PD-L1 inhibitor and a CTLA-4 inhibitor, such as pembrolizumab, nivolumab, cemiplimab, atezolizumab, avelumab, durvalumab and ipilimumab.

Description

TREATMENT OF SOLID TUMOURS

All documents cited or referenced herein, and all documents cited or referenced in herein cited documents, together with any manufacturer’s instructions, descriptions, product specifications, and product sheets for any products mentioned herein or in any document incorporated by reference herein, are hereby incorporated herein by reference in their entirety.

FIELD

The present disclosure relates to methods of treating cancers, in particular solid cancers such as colorectal cancer.

BACKGROUND

Colorectal cancer (colon cancer) is the second most common cause of cancer worldwide. In Australia, colorectal cancer is estimated to be the fourth most commonly diagnosed cancer in 2020 with an estimated 15,494 patients in a population of about 26 million (Cancer Australia Government statistics). Unfortunately, 30-50% of patients have occult or overt metastases at presentation and once tumours have metastasised, prognosis is poor with a five year survival rate dropping to around 14%.

In most cases, surgery is required to remove the tumour and depending on the stage of the cancer, further treatment by radiotherapy and/or chemotherapy may be required. However, these methods are invasive and can result in unwanted side effects if radiotherapy or chemotherapy is also involved.

In the past decade, the field of immunotherapy has witnessed remarkable advances for the treatment of cancer patients. The success of these therapies relies on the capacity of a patient’s immune system to single out cancer cells and kill them through T-cell mediated mechanisms. In the majority of cases, this requires administration of antibodies that block inhibitory immune checkpoints such as PD1/PDL1 and CTLA4. However, effective responses to immunotherapy are limited to select patients, particularly those with immunogenically “hot” tumours that are densely populated by cytotoxic T-cells. However, most cancer patients harbor immune effector cell-excluded “cold” tumours characterised by abundant immune suppressive regulatory T cells (Tregs), myeloid-derived suppressor cells (MDSC), as well as alternatively-activated macrophages (AAMs). To this end, strategies to convert the immunosuppressive tumour microenvironment by restricting the recruitment and polarization of AAMs have shown beneficial effects when combined with immune checkpoint blockade, suggesting the benefit of co-targeting these cells to augment anti-tumour immunity (De Henau O et al., (2016) 539(7629):443-447).

Elevated expression of the myeloid-specific SRC family kinase Hematopoietic Cell Kinase (HCK), and its associated kinase activity occurs in a majority of solid malignancies and correlates with poor patient survival (Poh AR et al., (2015) Oncotarget 6(18): 15752-15771).

The inventors have previously shown that genetic reduction of HCK expression favours the repolarisation of tumour associated macrophages (TAMs) from a tumour promoting AAM endotype to a tumouricidal conventionally activated endotype (Poh AR et al., (2017) Cancer Cell 31 (4):563-575.e5).

There is a need in the art for methods to treat colorectal cancer that may avoid the need for surgery or chemotherapy.

SUMMARY OF THE DISCLOSURE

The present disclosure is based on the finding that a pan-tyrosine kinase inhibitor dasatinib improved the anti-tumour activity conferred by anti-PD1.

Accordingly, in a first aspect, there is provided a method of treating a cancer in a subject, comprising: administering an effective amount of dasatinib to the subject; and administering an effective amount of an immune checkpoint inhibitor to the subject.

In some examples, the immune checkpoint inhibitor is selected from the group consisting of a PD-1 inhibitor, a PD-L1 inhibitor and a CTLA-4 inhibitor. In some examples, the immune checkpoint inhibitor is a PD-1 inhibitor. In some examples, the immune checkpoint inhibitor is selected from the group consisting of pembrolizumab, nivolumab, cemiplimab, atezolizumab, avelumab, durvalumab and ipilimumab. In some examples, the immune checkpoint inhibitor is a PD-1 inhibitor selected from the group consisting of pembrolizumab, nivolumab and cemiplimab.

In some examples, the cancer is a solid cancer. In some examples, the cancer is selected from the group consisting of colorectal cancer, gastric cancer, esophageal cancer, lung cancer, breast cancer, melanoma, head and neck squamous cell cancer, cutaneous squamous cell carcinoma, Merkel cell carcinoma, renal cell carcinoma, urothelial carcinoma, cervical cancer, hepatocellular carcinoma, endometrial carcinoma and tumour mutational burden-high cancer. In some examples, the cancer is colorectal cancer.

In some examples, the cancer is a blood cancer. In some examples, the blood cancer is selected from the group consisting of lymphoma and leukemia. In some examples, the blood cancer is selected from the group consisting of chronic myeloid leukemia, acute lymphoblastic leukemia, classical Hodgkin lymphoma and primary mediastinal large B-cell lymphoma.

In some examples, the subject is human.

In some examples, dasatinib and the immune checkpoint inhibitor are administered separately, sequentially or simultaneously.

In some examples, dasatinib is administered once per day in an amount of up to 150mg. In some examples, dasatinib is administered once per day in an amount of about 100mg, or about 140mg. In some examples, dasatinib is administered once per day in an amount of up to 80mg.

In some examples, dasatinib is administered orally.

In some examples, the immune checkpoint inhibitor is pembrolizumab, which is administered in an amount of about 200mg per 3 weeks, or about 400mg per 6 weeks. In some examples, the immune checkpoint inhibitor is pembrolizumab, which is administered in an amount of up to 150mg per 2 weeks, or in an amount of up to 300mg per 6 weeks.

In some examples, the immune checkpoint inhibitor is nivolumab, which is administered in an amount of up to 3mg/kg per 2 weeks. In some examples, the immune checkpoint inhibitor is nivolumab, which is administered in an amount of up to 2mg/kg per 2 weeks.

In some examples, the immune checkpoint inhibitor is cemiplimab, which is administered in an amount of about 350mg per 3 weeks. In some examples, the immune checkpoint inhibitor is cemiplimab, which is administered in an amount of up to 300mg per 3 weeks.

In some examples, the immune checkpoint inhibitor is ipilimumab, which is administered in an amount of up to 3mg/kg per week. In some examples, the immune checkpoint inhibitor is ipilimumab, which is administered in an amount of up to 2mg/kg per week. In some examples, the immune checkpoint inhibitor is administered intravenously by infusion or injection.

There is also provided dasatinib for use in treating a cancer, wherein dasatinib is administered in combination with an immune checkpoint inhibitor.

There is also provided an immune checkpoint inhibitor for use in treating a cancer, wherein the immune checkpoint inhibitor is administered in combination with dasatinib.

There is also provided use of dasatinib for the manufacture of a medicament for the treatment of a cancer, wherein the medicament is administered in combination with an immune checkpoint inhibitor.

There is also provided use of an immune checkpoint inhibitor for the manufacture of a medicament for the treatment of a cancer, wherein the medicament is administered in combination with dasatinib.

There is also provided a method of treating a cancer in a subject, comprising: administering an effective amount of a pharmaceutical composition comprising dasatinib and a pharmaceutically acceptable excipient to the subject; and administering an effective amount of a pharmaceutical composition comprising an immune checkpoint inhibitor and a pharmaceutically acceptable excipient to the subject.

There is also provided a pharmaceutical composition comprising dasatinib and a pharmaceutically acceptable excipient, for use in treating a cancer, wherein the pharmaceutical composition is administered in combination with an immune checkpoint inhibitor.

There is also provided a pharmaceutical composition comprising an immune checkpoint inhibitor and a pharmaceutically acceptable excipient, for use in treating a cancer, wherein the pharmaceutical composition is administered in combination with dasatinib.

There is also provided a method of treating a cancer in a subject, comprising: administering an effective amount of RK-20449 to the subject; and administering an effective amount of an immune checkpoint inhibitor to the subject.

In some examples, the immune checkpoint inhibitor is selected from the group consisting of a PD-1 inhibitor, a PD-L1 inhibitor and a CTLA-4 inhibitor. In some examples, the immune checkpoint inhibitor is a PD-1 inhibitor. In some examples, the immune checkpoint inhibitor is selected from the group consisting of pembrolizumab, nivolumab, cemiplimab, atezolizumab, avelumab, durvalumab and ipilimumab. In some examples, the immune checkpoint inhibitor is a PD-1 inhibitor selected from the group consisting of pembrolizumab, nivolumab and cemiplimab.

In some examples, the cancer is a solid cancer. In some examples, the cancer is selected from the group consisting of colorectal cancer, gastric cancer, esophageal cancer, lung cancer, breast cancer, melanoma, head and neck squamous cell cancer, cutaneous squamous cell carcinoma, Merkel cell carcinoma, renal cell carcinoma, urothelial carcinoma, cervical cancer, hepatocellular carcinoma, endometrial carcinoma and tumour mutational burden-high cancer. In some examples, the cancer is colorectal cancer.

In some examples, the cancer is a blood cancer. In some examples, the blood cancer is selected from the group consisting of lymphoma and leukemia. In some examples, the blood cancer is selected from the group consisting of chronic myeloid leukemia, acute lymphoblastic leukemia, classical Hodgkin lymphoma and primary mediastinal large B-cell lymphoma.

In some examples, the subject is human.

In some examples, RK-20449 and the immune checkpoint inhibitor are administered separately, sequentially or simultaneously.

In some examples, the immune checkpoint inhibitor is pembrolizumab, which is administered in an amount of about 200mg per 3 weeks, or about 400mg per 6 weeks. In some examples, the immune checkpoint inhibitor is pembrolizumab, which is administered in an amount of up to 150mg per 2 weeks, or in an amount of up to 300mg per 6 weeks.

In some examples, the immune checkpoint inhibitor is nivolumab, which is administered in an amount of up to 3mg/kg per 2 weeks. In some examples, the immune checkpoint inhibitor is nivolumab, which is administered in an amount of up to 2mg/kg per 2 weeks.

In some examples, the immune checkpoint inhibitor is cemiplimab, which is administered in an amount of about 350mg per 3 weeks. In some examples, the immune checkpoint inhibitor is cemiplimab, which is administered in an amount of up to 300mg per 3 weeks.

In some examples, the immune checkpoint inhibitor is ipilimumab, which is administered in an amount of up to 3mg/kg per week. In some examples, the immune checkpoint inhibitor is ipilimumab, which is administered in an amount of up to 2mg/kg per week.

In some examples, the immune checkpoint inhibitor is administered intravenously by infusion or injection.

There is also provided RK-20449 for use in treating a cancer, wherein dasatinib is administered in combination with an immune checkpoint inhibitor.

There is also provided an immune checkpoint inhibitor for use in treating a cancer, wherein the immune checkpoint inhibitor is administered in combination with RK-20449.

There is also provided use of RK-20449 for the manufacture of a medicament for the treatment of a cancer, wherein the medicament is administered in combination with an immune checkpoint inhibitor.

There is also provided use of an immune checkpoint inhibitor for the manufacture of a medicament for the treatment of a cancer, wherein the medicament is administered in combination with RK-20449.

There is also provided a method of treating a cancer in a subject, comprising: administering an effective amount of a pharmaceutical composition comprising RK-20449 and a pharmaceutically acceptable excipient to the subject; and administering an effective amount of a pharmaceutical composition comprising an immune checkpoint inhibitor and a pharmaceutically acceptable excipient to the subject.

There is also provided a pharmaceutical composition comprising RK-20449 and a pharmaceutically acceptable excipient, for use in treating a cancer, wherein the pharmaceutical composition is administered in combination with an immune checkpoint inhibitor.

There is also provided a pharmaceutical composition comprising an immune checkpoint inhibitor and a pharmaceutically acceptable excipient, for use in treating a cancer, wherein the pharmaceutical composition is administered in combination with RK-20449.

DESCRIPTION OF THE FIGURES

Figure 1A and B shows tumour volume of WT and Hck^KO mice following subcutaneous inoculation of MC38 cells and treatment with RK20449 (30 mg/kg, twice daily), Dasatinib (30mg/kg, twice daily) and/or anti-PD1 (200pg, once every 3 days). Treatment continued until tumour volume reached approx.. 600mm³. N> 10 mice per group.

Figure 2 shows the results of a C57BL/6 colon cancer model in which the effect of antibody-mediated stimulation of CD40 was assessed for HCK KO mice and mice treated with an HCK inhibitor.

DETAILED DESCRIPTION

General Techniques and Selected Definitions

The term “and/or”, e.g., “X and/or Y” shall be understood to mean either “X and Y” or “X or Y” and shall be taken to provide explicit support for both meanings or for either meaning.

Reference to the singular forms “a”, “an” and “the” is also understood to imply the inclusion of plural forms unless the context dictates otherwise.

Any discussion of documents, acts, materials, devices, articles or the like which has been included in the present specification is not to be taken as an admission that any or all of these matters form part of the prior art base or were common general knowledge in the field relevant to the present disclosure as it existed before the priority date of each claim of this application.

Throughout this specification, unless specifically stated otherwise or the context requires otherwise, reference to a single step, composition of matter, group of steps or group of compositions of matter shall be taken to encompass one and a plurality (i.e. one or more) of those steps, compositions of matter, groups of steps or group of compositions of matter.

Each example described herein is to be applied mutatis mutandis to each and every other example of the disclosure unless specifically stated otherwise.

Those skilled in the art will appreciate that the disclosure is susceptible to variations and modifications other than those specifically described. It is to be understood that the disclosure includes all such variations and modifications. The disclosure also includes all of the steps, features, compositions and compounds referred to or indicated in this specification, individually or collectively, and any and all combinations or any two or more of said steps or features.

The present disclosure is not to be limited in scope by the specific examples described herein, which are intended for the purpose of exemplification only. Functionally-equivalent products, compositions and methods are clearly within the scope of the disclosure.

The present invention as described herein can be performed using, unless otherwise indicated, conventional techniques of molecular biology and cellular biology. Such procedures are described, for example, in Sambrook, Fritsch & Maniatis, Molecular Cloning: A Laboratory Manual, Cold Spring Harbor Laboratories, New York, Second Edition (1989), whole of vols I, II, and III; DNA Cloning: A Practical Approach, Vols. I and II (D. N. Glover, ed., 1985), IRL Press, Oxford, whole of text; Oligonucleotide Synthesis: A Practical Approach (M. J. Gait, ed, 1984) IRL Press, Oxford, whole of text, and particularly the papers therein by Gait, ppl-22; Atkinson et al, pp35-81 ; Sproat et al, pp 83-115; and Wu et al, pp 135-151 ; 4. Nucleic Acid Hybridization: A Practical Approach (B. D. Hames & S. J. Higgins, eds., 1985) IRL Press, Oxford, whole of text; Immobilized Cells and Enzymes: A Practical Approach (1986) IRL Press, Oxford, whole of text; Perbal, B., A Practical Guide to Molecular Cloning (1984); Methods In Enzymology (S. CoIowick and N. Kaplan, eds., Academic Press, Inc.), whole of series, Sakakibara, D., Teichman, J., Lien, E. Land Fenichel, R.L. (1976). Biochem. Biophys. Res. Commun. 73 336-342; Merrifield, R.B. (1963). J. Am. Chem. Soc. 85, 2149-2154; Barany, G. and Merrifield, R.B. (1979) in The Peptides (Gross, E. and Meienhofer, J. eds.), vol. 2, pp. 1-284, Academic Press, New York. 12. Wunsch, E., ed. (1974) Synthese von Peptiden in Houben-Weyls Metoden der Organischen Chemie (Muler, E., ed.), vol. 15, 4th edn., Parts 1 and 2, Thieme, Stuttgart; Bodanszky, M. (1984) Principles of Peptide Synthesis, Springer-Verlag, Heidelberg; Bodanszky, M. & Bodanszky, A. (1984) The Practice of Peptide Synthesis, Springer-Verlag, Heidelberg; Bodanszky, M. (1985) Int. J. Peptide Protein Res. 25, 449-474; Handbook of Experimental Immunology, Vols. I-IV (D. M. Weir and C. C. Blackwell, eds., 1986, Blackwell Scientific Publications); and Animal Cell Culture: Practical Approach, Third Edition (John R. W. Masters, ed., 2000), ISBN 0199637970, whole of text.

Throughout this specification, unless the context requires otherwise, the word "comprise", or variations such as "comprises" or "comprising", will be understood to imply the inclusion of a stated step or element or integer or group of steps or elements or integers but not the exclusion of any other step or element or integer or group of elements or integers. The term “about”, as used herein when referring to a range is meant to encompass variations of ±20% or ±10%, more preferably ±5%, even more preferably ±1 % from the specified amount.

As used herein, the term “treat” or “treatment” or “treating” shall be understood to refer to the medical management of a patient with the intent to cure, ameliorate, stabilize, or prevent a disease, pathological condition or disorder. This term includes active treatment, i.e. treatment directed specifically toward the improvement of a disease, pathological condition, or disorder. In addition, this term includes palliative treatment, i.e. treatment designed for the relief of symptoms rather than curing the disease, pathological condition or disorder; and supportive treatment, i.e. treatment employed to supplement another specific therapy directed towards the improvement of the associated disease, pathological condition or disorder.

As used herein, a “PD-1 inhibitor” is any pharmacologic or biologic agent or medicinal product that reduces the activity or expression of PD-1 and/or modulates PD- 1 interactions with its ligands and/or other molecules and/or inhibits PD-1 signalling and/or pathway activity.

The term “subject” as used herein refers to a mammal including human and nonhuman animals. More particularly, the mammal is a human. Terms such as “subject”, “patient” or “individual” are terms that can, in context, be used interchangeably in the present disclosure.

As used herein "an increase in expression or phosphorylation" refers to an amount of gene expression, protein expression or protein phosphorylation that is at least about 0.05 fold more (for example 0.1 , 0.2, 0.3, 0.4, 0.5, 1 , 5, 10, 25, 50, 100, 1000, 10,000-fold or more) than the amount of gene expression, protein expression or protein phosphorylation in a subject not undergoing PD-1 inhibition or in a subject prior to undergoing PD-1 inhibition according to the methods described herein. "Increased" as it refers to gene expression, protein expression or protein phosphorylation also means at least about 5% more (for example 5, 6, 7, 8, 9, 10, 15, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95, 99 or 100%) than the amount of gene expression, protein expression or protein phosphorylation in a subject not undergoing PD-1 inhibition or in a subject prior to undergoing PD-1 inhibition according to the methods described herein. Amounts can be measured according to methods known in the art for determining amounts of gene expression, protein expression or protein phosphorylation. Ranges provided herein are understood to be shorthand for all of the values within the range. For example, a range of 1 to 50 is understood to include any number, combination of numbers, or sub-range from the group consisting 1 , 2, 3, 4, 5, 6, 7, 8, 9, 10, 11 , 12, 13, 14, 15, 16, 17, 18, 19, 20, 21 , 22, 23, 24, 25, 26, 27, 28, 29, 30, 31 , 32, 33, 34, 35, 36, 37, 38, 39, 40, 41 , 42, 43, 44, 45, 46, 47, 48, 49, or 50 (as well as fractions thereof unless the context clearly dictates otherwise).

Dasatinib

The present disclosure relates to therapeutic methods and uses involving dasatinib. Dasatinib is a kinase inhibitor having the chemical name: N-(2-chloro-6- methylphenyl)-2-[[6-[4-(2-hydroxyethyl)-1-piperazinyl]-2-methyl-4-pyrimidinyl]amino]-5- thiazolecarboxamide. It has the chemical structure:

Dasatinib is the active ingredient present in the product SPRYCEL.

Dasatinib, its preparation, and uses of the compound are disclosed in, for example, US6,596,746, US7,125,875, US7, 153,856, US7,491 ,725 and US8,680,103, the contents of each of which are incorporated herein by reference.

The present disclosure encompasses all forms of dasatinib, including the free base and salt forms, and all physical forms, solvates and polymorphs.

Dasatinib is most commonly used in the form of the free base. In some examples, dasatinib is used in the form of the free base. In some other examples, a salt form of dasatinib is used. Suitable salts include those formed with organic or inorganic acids. Typically, a pharmaceutically acceptable salt is used. Exemplary acid addition salts include, but are not limited to, sulfate, citrate, acetate, oxalate, chloride, bromide, iodide, nitrate, bisulfate, phosphate, acid phosphate, isonicotinate, lactate, salicylate, acid citrate, tartrate, oleate, tannate, pantothenate, bitartrate, ascorbate, succinate, maleate, gentisinate, fumarate, gluconate, glucuronate, saccharate, formate, benzoate, glutamate, methanesulfonate, ethanesulfonate, benzenesulfonate, p-toluenesulfonate, and pamoate (i.e., 1 ,1 '-methylene-bis-(2-hydroxy-3-naphthoate)) salts. Dasatinib is most commonly used in the form of a monohydrate. Those skilled in the art of medicinal chemistry will appreciate that many organic compounds can form complexes with solvents in which they are reacted or from which they are precipitated or crystallized. These complexes are known as "solvates". For example, a complex with water is known as a "hydrate". As used herein, the phrase “pharmaceutically acceptable solvate” or “solvate” refer to an association of one or more solvent molecules and a compound of the present disclosure. Examples of solvents that form pharmaceutically acceptable solvates include, but are not limited to, water, isopropanol, ethanol, methanol, DMSO, ethyl acetate, acetic acid, and ethanolamine. It will be understood that the present disclosure encompasses solvated forms, including hydrates, of dasatinib, as well as unsolvated dasatinib. In some examples, dasatinib is used in the form of a monohydrate.

RK-20449

The present disclosure also relates to therapeutic methods and uses involving RK-20449. RK-20449 is an inhibitor of Src family kinases, including HCK. It is a pyrrolo- pyrimidine derivative having the structure below:

RK-20449 is also known by the name A-419259, and was described in, for example, Wilson et ai, Oncogene, 2002, 21(53) p8075-8088. It is available in salt forms, including as a hydrochloride salt.

The present disclosure encompasses all forms of RK-20449, including the free base and salt forms, and all physical forms, solvates and polymorphs. Suitable salts include those formed with organic or inorganic acids. Typically, a pharmaceutically acceptable salt is used. Exemplary acid addition salts include, but are not limited to, sulfate, citrate, acetate, oxalate, chloride, bromide, iodide, nitrate, bisulfate, phosphate, acid phosphate, isonicotinate, lactate, salicylate, acid citrate, tartrate, oleate, tannate, pantothenate, bitartrate, ascorbate, succinate, maleate, gentisinate, fumarate, gluconate, glucuronate, saccharate, formate, benzoate, glutamate, methanesulfonate, ethanesulfonate, benzenesulfonate, p-toluenesulfonate, and pamoate (i.e., 1 ,1 '- methylene-bis-(2-hydroxy-3-naphthoate)) salts.

Those skilled in the art of medicinal chemistry will appreciate that many organic compounds can form complexes with solvents in which they are reacted or from which they are precipitated or crystallized. These complexes are known as "solvates". For example, a complex with water is known as a "hydrate". As used herein, the phrase “pharmaceutically acceptable solvate” or “solvate” refer to an association of one or more solvent molecules and a compound of the present disclosure. Examples of solvents that form pharmaceutically acceptable solvates include, but are not limited to, water, isopropanol, ethanol, methanol, DMSO, ethyl acetate, acetic acid, and ethanolamine. It will be understood that the present disclosure encompasses solvated forms, including hydrates, of RK-20449, as well as unsolvated RK-20449.

Immune checkpoint inhibitors

It is known to those skilled in the art that the immune system provides an inhibitory signal for its components to balance the immune response. Known immune checkpoint proteins include CTLA-4, PD1 and its ligands PD-L1 and PD-L2, and additionally LAG-3, BTLA, B7H3, B7H4, TIM3, KIR. It is recognized in the art that pathways involving LAG3, BTLA, B7H3, B7H4, TIM3 and KIR constitute immune checkpoint pathways similar to those dependent on CTLA-4 and PD-1 (Pardoll, (2012) Nature Rev Cancer 12: 252-264; Mellman et al., (2011) Nature 480: 480-490). In one example, the immune checkpoint protein is a human immune checkpoint protein.

In particular examples, the present disclosure is directed to the use of an immune checkpoint protein inhibitor, or “immune checkpoint inhibitor”. The term “immune checkpoint inhibitor” as used herein refers to any compound that inhibits the function of an immune checkpoint protein. The inhibition includes the spectrum from reduced function to complete blockade. In a particular example, the immune checkpoint protein inhibitor is an inhibitor of human immune checkpoint protein.

Immune checkpoint proteins have been reported in the art (Pardoll, (2012) Nature Rev. cancer 12:252-264). The term immune checkpoint includes experimental demonstration of stimulation of T lymphocyte responses caused by antigen receptors by inhibition of immune checkpoint protein in vitro or in vivo, e.g., mice lacking expression of immune checkpoint protein show signs of enhanced antigen-specific T lymphocyte response or autoimmunity (disclosed in Waterhouse et al., (1995) Science 270: 985-988; Nishimura et al., (1999) Immunity 11 : 141-151). It may also include demonstration of inhibition of the CD4+ or CD8+ T cell response caused by antigen receptors due to the intentional stimulation of immune checkpoint proteins in vitro or in vivo (Zhu et al., (2005) Nature Immunol 6:1245-1252).

Preferred immune checkpoint protein inhibitors are antibodies that specifically recognize immune checkpoint proteins. Examples include CTLA-4, PD1 , PDL-1 , PD- L2, LAG-3, BTLA, B7H3, B7H4, TIM3 and KIR inhibitors.

Ipilimumab is a fully human CTLA-4 blocking antibody currently sold under the name Yervoy® (Bristol-Myers Squibb). A further CTLA-4 inhibitor is tremelimumab (referenced in Ribas et al., (2013) J. Clin. Oncol. 31 : 616-22).

Examples of PD-1 inhibitors include humanized antibodies that block human PD- 1 , such as lambrolizumab (e.g, WO 2008/156712; Hamid et al., (2013) N. Engl. J. Med. 369:134-144). HPD109A and its humanized derivatives h409A11 , h409A16 and h409A17) or pidilizumab (disclosed in Rosenblatt et al., (2011) J Immunother. 34: 409- 18), and nivolumab (formerly known as MDX-1106 or BMS-936558, Topalian et al., (2012) N. Eng. J. Med. 366: 2443-3454). Other PD-1 inhibitors include, but are not limited to, the PD-L2 Fc fusion protein also known as B7-DC-lg or AMP-244 (Mkrticyan M, et al. (2012) J Immunol. 189: 2338-47). Further, immune checkpoint inhibitors include humanized or fully human antibodies that block PD-L1 , such as MEDI-4736 (WO2011106389A1), MPDL3280A (US8217149B2) and MIH1 (US8217149B2). Affymetrix (16.59883.82) available through eBioscience and pembrolizumab (sold under the trade name KEYTRUDA) as well as other PD-L1 inhibitors currently under investigation.

In one example, the immune checkpoint inhibitor is selected from the group consisting of a CTLA-4 inhibitor, PD-1 inhibitor, and PD-L1 inhibitor. In one example, the inhibitor is selected from the group consisting of ipilimumab, tremelimumab, labrolizumab, nivolumab, pidilizumab, pembrolizumab, spartalizumab, AMP-244, MEDI- 4736, MPDL3280A, and MIH1. Lambrolizumab is also known by alternative names MK- 3475 and pembrolizumab including analogs, in particular chimerized forms, humanized forms or humanized antibodies.

In some examples, the inhibitor is one that directly or indirectly stimulates or enhances antigen-specific T lymphocytes. Such inhibitors include, but are not limited to, agents that target immune checkpoint proteins and pathways involving PD-L2, LAG3, BTLA, B7H4 and TIM3. For example, human PD-L2 inhibitors known in the art include MIH18 (Pfistershammer et al., (2006) Eur J Immunol. 36: 1104-13). Another example of LAG3 inhibitors known in the art include soluble LAG3 (IMP321 or LAG3-lg as disclosed in W020090443273A2 and Brignon et al. (2009) Clin. Cancer Res. 15: 6225-6231) and human LAG3. Mouse antibodies or humanized antibodies (disclosed in W0200832601A1 , derived from IMP 701) or fully human antibodies (disclosed in EP 2320940A2) that inhibit human LAG3. Another example is provided by the use of blocking agents against BTLA, including but not limited to antibodies that block the interaction between human BTLA and its ligands (eg, 4C7 disclosed in WO2011014438). Yet another example includes, but is not limited to, an antibody against human B7H4 (disclosed in WO2013025779A1 and WO2013067492A1) or an antibody against soluble recombinant B7H4 (disclosed in US20120176745A1 , or antihuman B7H4 provided by the use of an agent that neutralizes B7H4, including clone H74: eBioscience # 14-5948). Yet another example is provided by agents that neutralize B7-H3, including but not limited to antibodies that neutralize human B7-H3 (e.g, MGA271 disclosed as BRCA84D and a derivative in US2012029496A1). An example includes an antibody that targets human TIM3 (such as those disclosed in WG2013006490A2, or Jones et al., J Exp Med. (2008) Nov 24; 205 (12): Provided by an agent that targets TIM3, including the anti-human TIM3 blocking antibody F38-2E2) disclosed by 2763-79.

Cancer Treatment

Cancers of various etiologies frequently contain PD-1 receptor-expressing cancer cell subpopulations. Tumour cell-expressed PD-1 modulates downstream pathways, signalling mediators of which can serve as biomarkers for predicting and monitoring response to therapeutic anti-PD-1 antibodies.

The methods as described herein are useful for treating cancer in a subject. In a particular example, the cancer is one that expresses PD-1. Expression of PD-1 by a cancer cell can be determined by various means including immunofluorescence, immunohistochemistry, flow cytometry, immunoblot, and in situ hybridization.

In one example, the cancer is a solid tumour. A solid tumour is defined herein as a mass of tissue that usually does not contain cysts or liquid areas. In another example, the solid tumour is a sarcoma, carcinoma or lymphoma. The solid tumour may be a tumour selected from the group consisting of colorectal cancer, gastric cancer, esophageal cancer, lung cancer, breast cancer, melanoma, head and neck squamous cell cancer, cutaneous squamous cell carcinoma, Merkel cell carcinoma, renal cell carcinoma, urothelial carcinoma, cervical cancer, prostate cancer, ovarian cancer, bladder cancer, hepatocellular carcinoma, endometrial carcinoma and tumour mutational burden-high cancer. PD-1 expression, signally or activity can also be indicated by p-S6 expression.

In some embodiments, the cancer is not a fibrotic cancer. In some embodiments, the cancer is not pancreatic cancer. In some embodiments, the methods described herein are not for preventing, reducing or inhibiting metastasis of a primary tumour.

In one example, the cancer is a blood cancer. In another example, the blood cancer is a cancer selected from the group consisting of chronic myeloid leukemia, acute lymphoblastic leukemia, classical Hodgkin lymphoma and primary mediastinal large IB- cell lymphoma.

In some examples, the cancer is one that has previously been identified as not responding or poorly responding to PD-1. The clinician will be able to determine whether a cancer is not responding to PD-1 therapy. Examples include a cancer that does not decrease in size or severity.

Responsiveness to therapy can be assessed by measuring the size of the tumour following therapy or severity/spread of the tumour following therapy (e.g. whether the tumour has increased in stage or metastasised). In another example, response can in some cases be assessed by a decrease in one or more of the phosphorylation status of auto-phosphorylation sites on SRC-family kinases, including SRC, HCK, FGR, ribosomal protein S6 (pS6), AKT, SRC homology 2-containing protein-tyrosine- phosphatase, phosphoinositide 3-kinase, extracellular signal regulated kinase, Translation initiation factor 4E binding protein 1 , Eukaryotic translation initiation factor 4B, Eukaryotic translation initiation factor 4G, mammalian target of rapamycin, PRAS40, or S6 kinase 1.

In one example, the present disclosure provides a method of treating cancer in a subject, comprising: administering an effective amount of a pharmaceutical composition comprising dasatinib and a pharmaceutically acceptable excipient to the subject; and administering an effective amount of a pharmaceutical composition comprising an immune checkpoint inhibitor and a pharmaceutically acceptable excipient to the subject. In one example, the subject to be treated according to a method described herein is one which has failed first line treatment with chemotherapy. In another example, the subject is one that has previously not responded or poorly responded to PD-1 or another immune checkpoint inhibitor.

In a further example, the present disclosure provides a pharmaceutical composition comprising dasatinib and a pharmaceutically acceptable excipient, for use in treating a cancer, wherein the pharmaceutical composition is administered in combination with an immune checkpoint inhibitor.

In a further example, the disclosure provides a pharmaceutical composition comprising an immune checkpoint inhibitor and a pharmaceutically acceptable excipient, for use in treating a cancer, wherein the pharmaceutical composition is administered in combination with dasatinib.

In some examples, dasatinib and the immune checkpoint inhibitor can be used or combined with one or more additional therapeutic agents or therapies (including, but not limited to, radiation therapy or chemotherapy). The additional therapeutic agent may be administered concurrently or sequentially with dasatinib and the immune checkpoint inhibitor.

Pharmaceutical compositions

The PD-1 inhibitor may be administered to the subject in the format in which it is approved by a regulatory agency (e.g. FDA or TGA). Similarly, dasatinib may be administered to the subject in its approved form according to the product information (PI). RK-20449 may also be administered in any suitable composition.

The pharmaceutical compositions provided herein, may contain pharmaceutical carriers or diluents. The term "composition" as used herein is intended to cover products containing the specified ingredients in the specified amounts, as well as any products produced directly or indirectly from a combination of the specified ingredients in the specified amounts. "Pharmaceutically acceptable" means that the carrier, diluent or excipient must be compatible with the other ingredients of the formulation and not harmful to its recipient. A biological product, such as an antibody of the disclosure, may be composed of a pharmaceutical composition containing one or more antibodies or fragments thereof and a pharmaceutically acceptable carrier. As used herein, a "pharmaceutically acceptable carrier" includes any and all solvents, dispersion media, coatings, antibacterial and antifungal agents, isotonic and absorption delaying agents, and physiologically compatible similar. Preferably, the carrier is suitable for intravenous, intramuscular, subcutaneous, parenteral, spinal or epidermal administration (e.g., by injection or infusion).

The pharmaceutical composition of the present invention may include one or more pharmaceutically acceptable salts, antioxidants, aqueous and non-aqueous carriers and/or adjuvants such as preservatives, wetting agents, emulsifying agents and dispersing agents. Pharmaceutical compositions for administration of the compounds and agents of the present disclosure are suitably presented in unit dosage form and can be prepared by any of the methods well known in pharmacology and drug delivery technology.

In pharmaceutical compositions, the active target compound is included in an amount sufficient to effect the desired process or condition of the disease. Pharmaceutical compositions containing active ingredients may be in a form suitable for oral use (e.g. tablet). Compositions intended for oral use may be prepared according to any method known in the art for making pharmaceutical compositions, and such compositions may contain one or more agents selected from the group consisting of sweeteners, flavourings, agents, colorants, antioxidants and preservatives in order to provide a pharmaceutical delicate and palatable formulation. Formulations for oral use can also be presented in the form of hard gelatin capsules in which the active ingredient is mixed with an inert solid diluent such as calcium carbonate, calcium phosphate or kaolin; or in the form of soft gelatin capsules in which the active ingredient is in water or oil (e.g, peanut oil, liquid paraffin, or olive oil).

The pharmaceutical composition of the present invention may also be in the form of an oil-in-water emulsion. The oily phase may be a vegetable oil, such as olive oil or flower oil; or a mineral oil, such as liquid paraffin, or a mixture of these oils. Suitable emulsifiers may be naturally occurring gums, such as acacia or tragacanth; naturally occurring phospholipids, such as soybean, lecithin; and esters or partial esters derived from fatty acids and hexitol anhydrides, such as sorbitan sugar alcohol monooleate; and condensation products of these partial esters with ethylene oxide, such as polyoxyethylene sorbitan monooleate. The emulsion may also contain sweeteners and flavouring agents. Syrups and elixirs can be formulated with sweeteners such as glycerol, propylene glycol, glucose or sucrose. Such formulations may also contain a demulcent, a preservative, a flavouring agent, and a colouring agent. Oral solutions can be prepared in combination with, for example, cyclodextrin, PEG and surfactants. The compounds of the present disclosure can be formulated for placement in medical devices, which can include a variety of conventional grafts, stents (including stent grafts), catheters, expanders, baskets, or implantable or permanently implanted body cavities. For example, compounds can be delivered to a tumour or the microenvironment surrounding the tumour.

In some examples, dasatinib is administered in the form of a pharmaceutical composition which is a tablet, e.g. a coated tablet.

A tablet may be made for example by compression or moulding, optionally with one or more accessory ingredients. Compressed tablets may for example be prepared by compressing in a suitable machine the active ingredient in a free-flowing form such as a powder or granules, optionally mixed with a binder, lubricant, inert diluent, lubricating, surface active, or dispersing agent. Moulded tablets may for example be made by moulding in a suitable machine a mixture of the powdered compound moistened with an inert liquid diluent. Tablets may be optionally coated or scored, and may be formulated so as to provide slow or controlled release of the active compound.

The active can, for example, be administered in a form suitable for immediate release or extended release. Immediate release or extended release can be achieved by the use of suitable pharmaceutical compositions comprising the active or, particularly in the case of extended release, by the use of devices such as subcutaneous implants or osmotic pumps. An active may also be administered liposomally.

Exemplary compositions may contain, for example, excipients such as fillers, binders, extenders, disintegrants, diluents, and/or lubricants such as those known in the art. Suitable binders include starch, gelatin, natural sugars such as glucose or betalactose, corn sweeteners, natural and synthetic gums such as acacia, tragacanth or sodium alginate, carboxymethylcellulose, polyethylene glycol, waxes, and the like.

Disintegrators include without limitation, starch, methylcellulose, agar, bentonite, xanthan gum, and the like.

Lubricants, glidants, flavours, colouring agents, and stabilisers may also be added for ease of fabrication and use. Lubricants used in these dosage forms include sodium oleate, sodium stearate, magnesium stearate, sodium benzoate, sodium acetate, sodium chloride, and the like.

The compositions of the present disclosure may also include polymeric excipients/additives or carriers, e.g., polyvinylpyrrolidones, derivatised celluloses such as hydroxymethylcellulose, hydroxyethylcellulose, and hydroxypropylmethylcellulose, Ficolls (a polymeric sugar), hydroxyethylstarch (HES), dextrates (e.g., cyclodextrins, such as 2-hydroxypropyl-p-cyclodextrin and sulfobutylether-p-cyclodextrin), polyethylene glycols, and pectin. The compositions may further include buffers, thickeners, preservatives (including antioxidants), inorganic salts (e.g., sodium chloride), antimicrobial agents (e.g., benzalkonium chloride), antistatic agents, lipids (e.g., phospholipids such as lecithin and other phosphatidylcholines, phosphatidylethanolamines, fatty acids and fatty esters, steroids (e.g., cholesterol)), and/or chelating agents (e.g., EDTA, zinc and other such suitable cations).

For oral administration in liquid form, the oral drug components can be combined with any oral, non-toxic, pharmaceutically acceptable inert carrier such as ethanol, glycerol, water, and the like.

Other pharmaceutical excipients and/or additives suitable for use in the compositions according to the present disclosure are listed in "Remington: The Science & Practice of Pharmacy", 19. sup. th ed., Williams & Williams, (1995), and in the "Physician's Desk Reference", 52. sup. nd ed., Medical Economics, Montvale, N.J. (1998), and in "Handbook of Pharmaceutical Excipients", Third Ed., Ed. A. H. Kibbe, Pharmaceutical Press, 2000.

As discussed above, in the methods of the present disclosure, dasatinib may for example be administered in the form of a coated tablet. In some examples, dasatinib is administered in the form of a coated tablet containing the active, lactose, microcrystalline cellulose, croscarmellose sodium, hydroxypropyl cellulose and magnesium stearate, and having a coating containing hypromellose, titanium dioxide and polyethylene glycol.

In some examples, dasatanib is administered in the form of coated tablets containing 20mg, 50mg, 70mg or 100mg dasatinib.

In some examples, the composition is formulated for parenteral delivery, for example by intravenous infusion or injection. In some examples, the immune checkpoint inhibitor is used in the form of a pharmaceutical composition which is suitable for intravenous infusion or injection.

Compositions for parenteral administration include aqueous and non-aqueous sterile injections, solutions which may contain anti-oxidants, buffers, tonicity modifiers, bacteriostats, and solutes that render the formulation isotonic with the blood of the intended recipient; and aqueous and non-aqueous sterile suspensions which may include suspending agents and thickening agents. The formulations may be presented in unit-dose or multi-dose containers, for example, sealed ampoules and vials, and may be stored in a freeze-dried condition requiring only the addition of a sterile liquid carrier, for example saline or water-for-injection, immediately prior to use. Extemporaneous injection solutions and suspensions may be prepared from sterile powders, granules and tablets of the kind previously described. Exemplary compositions for parenteral administration include injectable solutions or suspensions which can contain, for example, suitable non-toxic, parenterally acceptable diluents or solvents, such as mannitol, 1.3-butanediol, water, Ringer’s solution, an isotonic sodium chloride solution, or other suitable dispersing or wetting and suspending agents, including synthetic mono- or diglycerides, and fatty acids, including oleic acid, or Cremaphor.

For example, the formulation may be a sterile, lyophilized composition that is suitable for reconstitution in an aqueous vehicle prior to injection. In one example, a formulation suitable for parenteral administration conveniently comprises a sterile aqueous preparation of an active ingredient, which may for example be formulated to be isotonic with the blood of the recipient.

In some examples, the immune checkpoint inhibitor is provided in the form of a formulation for intravenous infusion or injection. In some examples, the immune checkpoint inhibitor is provided in the form of a concentrate, for admixing with an aqueous diluent such as saline prior to administration. In some examples, the immune checkpoint inhibitor is provided in the form of a solid composition, for admixing with an aqueous diluent such as saline prior to administration.

In some examples, the immune checkpoint inhibitor is provided in the form of an aqueous composition comprising water for injection and one or more of a surfactant, a buffer, an inorganic salt, and a polyol.

In some examples, the immune checkpoint inhibitor is pembrolizumab, and is provided in the form of an aqueous composition for injection comprising water for injection, L-histidine, polysorbate 80 and sucrose. In some examples the immune checkpoint inhibitor is pembrolizumab and is provided in a unit dosage form containing about 100mg pembrolizumab.

In some examples, the immune checkpoint inhibitor is nivolumab, and is provided in the form of an aqueous composition for injection which comprises water for injection, manitol, pentetic acid, polysorbate 80, sodium chloride and sodium citrate, and which may contain hydrochloric acid and/or sodium hydroxide to adjust the pH to 6. In some examples, the immune checkpoint inhibitor is nivolumab and is provided in a unit dosage form containing about 40mg, about 100mg, or about 240mg nivolumab.

In some examples, the immune checkpoint inhibitor is cemiplimab, and is provided in the form of an aqueous composition for infusion which contains water for injection, L-histidine, L-proline, sucrose and polysorbate 80. In some examples, the immune checkpoint inhibitor is cemiplimab and is provided in a unit dosage form containing about 350mg cemiplimab.

In some examples, the immune checkpoint inhibitor is atezolizumab, and is provided in the form of an aqueous composition for infusion which contains water for injection, histidine, glacial acetic acid, sucrose and polysorbate 20. In some examples, the immune checkpoint inhibitor is atezolizumab and is provided in a unit dosage form containing about 840mg or about 1200mg atezolizumab.

In some examples, the immune checkpoint inhibitor is avelumab, and is provided in the form of an aqueous composition for infusion which contains water for injection, glacial acetic acid, polysorbate 20 and sodium hydroxide. In some examples, the immune checkpoint inhibitor is avelumab and is provided in a unit dosage form containing about 200mg avelumab.

In some examples, the immune checkpoint inhibitor is durvalumab, and is provided in the form of an aqueous composition for infusion which contains water for injection, histidine, trehalose, and polysorbate 80. In some examples, the immune checkpoint inhibitor is durvalumab and is provided in a unit dosage form containing about 120mg or about 500mg durvalumab.

In some examples, the immune checkpoint inhibitor is imipililumab, and is provided in the form of an aqueous composition for infusion which comprises water for injection, diethylene triamine pentaacetic acid (DTPA), mannitol, polysorbate 80, sodium chloride, tris hydrochloride, at pH 7. In some examples, the immune checkpoint inhibitor is imipililumab and is provided in a unit dosage form containing about 50mg, or about 200mg imipililumab.

Dosages

In the methods of the present disclosure, an effective amount of dasatinib is administered and an effective amount of an immune checkpoint inhibitor is administered to the subject. Dasatinib and the immune checkpoint inhibitor may be administered in any appropriate form to the subject, for example they may be administered separately, sequentially or simultaneously. In some examples, dasatinib and the immune checkpoint inhibitor are administered separately to the subject; that means that each active is administered as part of a single treatment regime, but they are administered at different times and different dosage intervals, as will be appropriate for the specific active.

In some examples, the subject is human. In some examples, the subject is an adult human.

As shown by the examples below, administration of a combination of dasatinib and an immune checkpoint inhibitor also provided effects greater than the properties of each agent as monotherapy. Thus, it is anticipated that the combination therapy of the present disclosure may provide improved therapeutic effects, compared for example with administration of each agent as a single therapy. In other words, in the case, where each active agent is dosed at its currently approved therapeutic dose, improved therapeutic effects (e.g. one or more of reduction of tumour growth, reduced rate of tumour growth, increased survival time) may be observed. Alternatively, use of the combination therapy of the present disclosure may enable the dosage amount of one or both actives to be reduced, e.g. compared with their currently approved dosage levels, and may lead to avoidance of or reduction in the level of side effects experienced by the subject.

Dasatinib has been approved for once daily oral dosing in daily dosage amounts of 100mg and 140mg, depending on the indication.

In some examples of the present disclosure, dasatinib is administered orally. In some examples, dasatinib is administered twice per day or, more preferably, once per day. In some examples dasatinib is administered in an amount of up to 150 mg per day, or up to 140 mg per day, or up to 130mg per day, or up to 120 mg per day, or up to 110 mg per day, or up to 100 mg per day, or up to 90 mg per day, or up to 80 mg per day, or up to 70 mg per day or up to 60 mg per day, or up to 50 mg per day. In some examples, dasatinib is administered in an amount of about 100 mg per day, or in an amount of about 140 mg per day.

In some examples dasatinib is administered once per day in an amount of up to 150 mg per day, or up to 140 mg per day, or up to 130mg per day, or up to 120 mg per day, or up to 110 mg per day, or up to 100 mg per day, or up to 90 mg per day, or up to 80 mg per day, or up to 70 mg per day or up to 60 mg per day, or up to 50 mg per day. In some examples, dasatinib is administered once per day in an amount of about 100 mg per day, or in an amount of about 140 mg per day. In some examples dasatinib is administered orally once per day in an amount of up to 150 mg per day, or up to 140 mg per day, or up to 130mg per day, or up to 120 mg per day, or up to 110 mg per day, or up to 100 mg per day, or up to 90 mg per day, or up to 80 mg per day, or up to 70 mg per day or up to 60 mg per day, or up to 50 mg per day. In some examples, dasatinib is administered orally once per day in an amount of about 100 mg per day, or in an amount of about 140 mg per day.

Many immune checkpoint inhibitors are typically administered intravenously, e.g. by infusion or injection, at specified dosage intervals, e.g. once every 1 , 2 or 3 weeks.

In the case where the immune checkpoint inhibitor is pembrolizumab, it may for example be administered intravenously, e.g. by injection. For example, it may be administered in an amount of about 200mg per 3 weeks, or about 400mg per 6 weeks, or pto 200mg per 3 weeks, or up to 400mg per 3 weeks. In some examples, it may be administered in an amount of up to 150mg per 2 weeks, or in an amount of up to 300mg per 6 weeks.

In the case where the immune checkpoint inhibitor is nivolumab, it may for example be administered intravenously, e.g. by injection. For example, it may be administered in an amount of up to 3mg/kg per 2 weeks, or about 3mg/kg per 2 weeks. In some examples, it may be administered in an amount of up to 2mg/kg per 2 weeks.

In the case where the immune checkpoint inhibitor is cemiplimab, it may for example be administered intravenously, e.g. by infusion. For example, it may be administered in an amount of up to 350mg per 3 weeks, or about 350mg per 3 weeks. In some examples, it may be administered in an amount of up to 300mg per 3 weeks.

In the case where the immune checkpoint inhibitor is ipilimumab, it may for example be administered intravenously, e.g. by injection. For example, it may be administered in an amount of up to 3mg/kg per week, or about 3mg/kg per week. In some examples, it may be administered in an amount of up to 2mg/kg per week.

In the case where the immune checkpoint inhibitor is atezolizumab, it may for example be administered intravenously, e.g. by injection. For example, it may be administered in an amount of up to 840mg per 2 weeks, or up to 1200mg per 3 weeks, or up to 1680 mg per 4 weeks, or about 840mg per 2 weeks, or about 1200 mg per 3 weeks, or about 100mg per 4 weeks. In some examples, it may be administered in an amount of up to 700mg per 2 weeks, or up to 1000mg per 3 weeks, or up to 1400mg per 4 weeks. In the case where the immune checkpoint inhibitor is avelumab, it may for example be administered intravenously, e.g. by infusion. For example, it may be administered in an amount of up to 10mg/kg per 2 weeks, or about 10mg/kg per 2 weeks, or up to 800mg per 2 weeks, or about 800mg per 2 weeks. In some examples, it may be administered in an amount of up to 600mg per 2 weeks, or up to 8mg/kg per 2 weeks.

In the case where the immune checkpoint inhibitor is durvalumab, it may for example be administered intravenously, e.g. by infusion. For example, it may be administered in an amount of up to 10mg/kg per 2 weeks, or about 10mg/kg per 2 weeks. In some examples, it may be administered in an amount of up to 8mg/kg per 2 weeks.

Whilst in some examples, dasatanib and a single immune checkpoint inhibitor may be the only active ingredients administered to the subject, in some other examples, one or more further active ingredients may be administered with dasatinib and the immune checkpoint inhibitor. For example, more than one immune checkpoint inhibitor may be administered in combination wih dasatinib. In some embodiments, a therapeutic regime involving the use of dasatinib, a PD-L1 or PD-1 inhibitor (e.g. nivolumab, pembrolizumab) and a CTLA4 inhibitor (e.g. ipilimumab) is used, for example for the treatment of colon cancer. For DNA mismatch repair (MMR) proficient forms of colon cancer, while these MMR-proficient cancers can remain resistant to single immune checkpoint blockade therapy, it is considered that the overall response rate can be increased by combining a PD-1 or PD-L1 inhibitor plus a CTLA4 inhibitor (Chalabi et al, Nature Medicine, 2020, Vol 26, p566-576). In some embodiments, a combination of nivolumab, ipilimumab and dasatinib is used.

In some embodiments, a combination of dasatinib with a PD-1 or PD-L1 inhibitor and a further active agent which is an immunotherapy which acts via stimulation of CD40 or 0X40 is used.

As a further example, anticancer agents other than dasatanib and an immune checkpoint inhibitor may also be administered as part of the treatment regime.

It will be appreciated by persons skilled in the art that numerous variations and/or modifications may be made to the above-described embodiments, without departing from the broad general scope of the present disclosure. The present embodiments are, therefore, to be considered in all respects as illustrative and not restrictive. EXAMPLES

Materials and Methods

Mice

Age- and sex-matched WT, and Hd^° (Lowell, CA et al. (1994) Genes and Development 8:387-398) mice were bred and maintained in specific pathogen-free facilities at LaTrobe University, and the Austin Hospital, Australia. All animal studies were approved and conduced in accordance with the Animal Ethics Committee of the Olivia Newton John Cancer Research Institute/Austin Hospital.

Cell lines and tumour models

The mouse MC38 colon cell line was maintained in DMEM/F12 supplemented with 10% FCS at 37°C with 10% CO2. Cell lines were tested negative for mycoplasma. Six week old C57BL/6 WT or Hck^⁰ mice were inoculated subcutaneously with 2 x 10⁶ MC38 cells into the right flank. Once palpable tumours formed, mice were randomised into treatment groups. Where indicted, mice were either treated with IgG control, RK20449 (30 mg/kg, diluted in 12% Captisol, twice daily i.p), Dasatinib (30 mg/kg diluted in 12% Captisol, twice daily i.p) and anti-PD1 (clone RMP14, 200 pg once every 3 days i.p). Tumour volume (mm³) was measured using digital calipers using the following formula: (length x width²)/2. T umour growth was measured by an independent assessor who was blinded to the experimental conditions.

Anti-PD1 inhibitor

The anti-PD1 inhibitor used herein was a purified anti-mouse CD279 (PD-1) antibody. The inhibitor can be purchased commercially from a number of sources including BioLegend, Bio X Cell, absolute antibody, and CrownVivo.

Dasatinib

Dasatinib was obtained from Selleck Chemicals.

RK20449

RK20449 is available from, for example, Cayman Chemicals and Reagency.

Western Blot Protein lysates were prepared as previously described and resolved on 10% SDS-polyacrylamide gels (Poh AR et al., (2017) Cancer Immunol Res 8(4):428-435). Following dry transfer, PVDF membranes were blocked for 1 hour in Intercept Blocking Buffer (LI-COR Biosciences) and incubated overnight in primary antibodies at 4°C. The next day, blots were incubated with fluorescent-conjugated secondary antibodies for 1 hour. Signals were detected using the Odyssey Infrared Imaging System (LI-COR Biosciences). Actin was used as a loading control.

Antibodies

The antibodies used herein as summarised in Table 1 below.

Table 1 Antibodies

Statistics

All experiments were performed at least twice with a minimum of three age- and sex-matched mice per group. The specific n (number of animals) used per cohort is indicated in the respective figure legends. Unless otherwise stated, comparisons between mean values were performed with a 2-tailed Student’s t-test as appropriate using Prism 8 software (GraphPad). A P value of less than 0.05 was considered statistically significant. Example 1 Pharmacologic inhibition of HCK enhances therapeutic efficacy of immune checkpoint blockade

Wild-type (WT) mice were treated with the pan-tyrosine kinase inhibitor Dasatinib or the HCK-specific pyrrolo-pyrimidine derivative RK20449. Consistent with the characterised activity of Dasatinib on HCK (Montero, J.C et al. (2011) Clin Cancer Res 17(17):5546-52), and the associated capacity to suppress the appearance of the autophosphorylated kinase isoform (Figure 1A), the inventors observed that both tyrosine kinase inhibitors improved the anti-tumour activity conferred by anti-PD1. This is demonstrated in Figure 1 B.

The contribution of Dasatinib-mediated HCK inhibition was functionally assessed. This was done by comparing MC38 allograft (murine colon adenocarcinoma cells) growth between Dasatinib- treated WT and Hck knock-out Hck^° hosts. As shown in Figure 1 B and C, Dasatinib treatment improved anti-PD1 mediated anti-tumour immune responses primarily through inhibition of HCK signalling.

Example 2: Colon Cancer Model and Effect of CD40 Stimulation

Six-week-old C57BL/6 WT or Hck^° mice were inoculated subcutaneously with 2x10⁶ MC38 into the right flank. Once palpable tumours formed, mice were randomized into treatment groups. Where indicated, mice were either treated with vehicle (12% Captisol), IgG control, RK20449 (30mg/kg, diluted in 12% Captisol, twice daily i.p) or aCD40 (Clone FGK45, 100pg once every 3 days i.p) for 10 days. The results are shown in Figure 2. Antibody-mediated stimulation of CD40 was found to reduce tumour burden more in Hck KO hosts than in WT hosts; or in RK20449 (Hck-inhibitor) treated WT mice.

Claims

28 CLAIMS:

1. A method of treating a cancer in a subject, comprising: administering an effective amount of dasatinib to the subject; and administering an effective amount of an immune checkpoint inhibitor to the subject.

2. A method as claimed in claim 1 , wherein the immune checkpoint inhibitor is selected from the group consisting of a PD-1 inhibitor, a PD-L1 inhibitor and a CTLA-4 inhibitor.

3. A method as claimed in claim 2, wherein the immune checkpoint inhibitor is a PD-1 inhibitor.

4. A method as claimed in claim 1 or claim 2, wherein the immune checkpoint inhibitor is selected from the group consisting of pembrolizumab, nivolumab, cemiplimab, atezolizumab, avelumab, durvalumab and ipilimumab.

5. A method as claimed in claim 2 or 4, wherein the immune checkpoint inhibitor is a PD-1 inhibitor selected from the group consisting of pembrolizumab, nivolumab and cemiplimab.

6. A method as claimed in any of claims 1 to 5, wherein the cancer is a solid cancer.

7. A method as claimed in claim 6, wherein the cancer is selected from the group consisting of colorectal cancer, gastric cancer, esophageal cancer, lung cancer, breast cancer, melanoma, head and neck squamous cell cancer, cutaneous squamous cell carcinoma, Merkel cell carcinoma, renal cell carcinoma, urothelial carcinoma, cervical cancer, hepatocellular carcinoma, endometrial carcinoma and tumour mutational burden-high cancer.

8. A method as claimed in claim 7, wherein the cancer is colorectal cancer.

9. A method as claimed in any of claims 1 to 5, wherein the cancer is a blood cancer.

10. A method as claimed in claim 9, wherein the blood cancer is selected from the group consisting of lymphoma and leukemia.

11. A method as claimed in claim 10, wherein the blood cancer is selected from the group consisting of chronic myeloid leukemia, acute lymphoblastic leukemia, classical Hodgkin lymphoma and primary mediastinal large B-cell lymphoma.

12. A method as claimed in any of claims 1 to 11 , wherein the subject is human.

13. A method as claimed in any of claims 1 to 12, wherein dasatinib and the immune checkpoint inhibitor are administered separately, sequentially or simultaneously.

14. A method as claimed in any of claims 1 to 13, wherein dasatinib is administered once per day in an amount of up to 150mg.

15. A method as claimed in claim 14, wherein dasatinib is administered once per day in an amount of about 100mg, or about 140mg.

16. A method as claimed in claim 14, wherein dasatinib is administered once per day in an amount of up to 80mg.

17. A method as claimed in any of claims 1 to 16, wherein dasatinib is administered orally.

18. A method as claimed in any of claims 1 to 17, wherein the immune checkpoint inhibitor is pembrolizumab, which is administered in an amount of about 200mg per 3 weeks, or about 400mg per 6 weeks.

19. A method as claimed in any of claims 1 to 17, wherein the immune checkpoint inhibitor is pembrolizumab, which is administered in an amount of up to 150mg per 2 weeks, or in an amount of up to 300mg per 6 weeks.

20. A method as claimed in any of claims 1 to 17, wherein the immune checkpoint inhibitor is nivolumab, which is administered in an amount of up to 3mg/kg per 2 weeks.

21. A method as claimed in any of claims 1 to 17, wherein the immune checkpoint inhibitor is nivolumab, which is administered in an amount of up to 2mg/kg per 2 weeks.

22. A method as claimed in any of claims 1 to 17, wherein the immune checkpoint inhibitor is cemiplimab, which is administered in an amount of about 350mg per 3 weeks.

23. A method as claimed in any of claims 1 to 17, wherein the immune checkpoint inhibitor is cemiplimab, which is administered in an amount of up to 300mg per 3 weeks.

24. A method as claimed in any of claims 1 to 17, wherein the immune checkpoint inhibitor is ipilimumab, which is administered in an amount of up to 3mg/kg per week.

25. A method as claimed in any of claims 1 to 17, wherein the immune checkpoint inhibitor is ipilimumab, which is administered in an amount of up to 2mg/kg per week.

26. A method as claimed in any of claims 1 to 25, wherein the immune checkpoint inhibitor is administered intravenously by infusion or injection.

27. Dasatinib for use in treating a cancer, wherein dasatinib is administered in combination with an immune checkpoint inhibitor.

28. An immune checkpoint inhibitor for use in treating a cancer, wherein the immune checkpoint inhibitor is administered in combination with dasatinib.

29. Use of dasatinib for the manufacture of a medicament for the treatment of a cancer, wherein the medicament is administered in combination with an immune checkpoint inhibitor.

30. Use of an immune checkpoint inhibitor for the manufacture of a medicament for the treatment of a cancer, wherein the medicament is administered in combination with dasatinib.

31. A method of treating a cancer in a subject, comprising: administering an effective amount of a pharmaceutical composition comprising dasatinib and a pharmaceutically acceptable excipient to the subject; and administering an effective amount of a pharmaceutical composition comprising an immune checkpoint inhibitor and a pharmaceutically acceptable excipient to the subject.

32. A pharmaceutical composition comprising dasatinib and a pharmaceutically acceptable excipient, for use in treating a cancer, wherein the pharmaceutical composition is administered in combination with an immune checkpoint inhibitor.

33. A pharmaceutical composition comprising an immune checkpoint inhibitor and a pharmaceutically acceptable excipient, for use in treating a cancer, wherein the pharmaceutical composition is administered in combination with dasatinib.