Drug conjugate compounds for stimulating the antitumor immune response

Abstract

Despite significant progresses made in the treatment of cancers with the development of immune checkpoint inhibitors (ICI), several cancers exhibit resistance to these agents. Certain tumors are considered immunologically cold tumors usually respond poorly to cancer immunotherapies such as immune checkpoint blockade (ICB) therapy, and thus there is a need to develop novel approaches for the treatment of such cancers. The present application discloses methods and uses for enhancing the anti-tumor immune response in a subject suffering from a Sortilin-expressing cancer and / or treating a subject suffering from a Sortilin-expressing cancer, such as a cancer resistant to an immunotherapy, through the administration of conjugate compounds comprising a Sortilin-targeting agent conjugated to an anticancer agent. Combination therapies comprising the conjugate compounds and an immunotherapy such as ICB are also disclosed.

Description

[0001] TITLE

[0002] DRUG CONJUGATE COMPOUNDS FOR STIMULATING THE ANTITUMOR IMMUNE RESPONSE

[0003] CROSS REFERENCE TO RELATED APPLICATIONS

[0004] The present application claims the benefit of U.S. provisional Patent application No. 63 / 383,561 , filed on November 14, 2022. The entire content of this application is incorporated herein by reference.

[0005] SEQUENCE LISTING

[0006] A sequence listing is submitted herewith as an XML file named G 11718-00471 -SSS_Seq listing.xml, that was created on November 14, 2023, and having a size of ~100 kilobytes. The content of the aforementioned file is hereby incorporated by reference in its entirety.

[0007] TECHNICAL FIELD

[0008] The present invention generally relates to the field of oncology, and more specifically to the treatment of cancers.

[0009] BACKGROUND ART

[0010] The prevalence of cancer in human and animal populations and its role in mortality means there is a continuing need for new drugs which are effective against tumors. Elimination of a tumour or a reduction in its size or reducing the number of cancer cells circulating in the blood or lymph node systems may be beneficial in a variety of ways; reducing pain or discomfort, preventing metastasis, facilitating operative intervention, and more importantly prolonging life.

[0011] Various attempts have been made to help the immune system to fight tumors. One early approach, in the late 19thcentury, involved a general stimulation of the immune system, e.g., through the administration of bacteria (live or killed) to elicit a general immune response which would also be directed against the tumor.

[0012] Recent approaches aimed at helping the immune system specifically to recognize tumorspecific antigens (TSAs) (or tumor associated antigens, TAAs) involve administration of tumorspecific antigens, typically combined with an adjuvant to the subject. However, a lack of a powerful immune response to TAAs is often observed in cancer. One of the factors responsible for the weak response to TAAs is the induction of inhibitory pathways / signals that suppress the immune response (often referred to as “immune checkpoints”). Whereas such inhibitory signals are important for maintenance of self-tolerance and to protect tissues from damage when the immune system is responding to pathogenic infection, they may also reduce what could otherwise be a helpful response by the body to the development of tumors. A novel therapeutic era has come of age with immune checkpoint inhibitors or blockers (ICB) targeting inhibitory T-cell receptors such as CTLA-4, PD-L1 and PD-1 (Marabelle, Oncolmmunology 2016). This burgeoning field has even been awarded by the 2018 Nobel Prize in Medicine. These immunotherapeutic agents provide good clinical results in several advanced cancers including lung (Reck, NEJM 2016), melanoma (Robert, NEJM 2011), genitourinary (Motzer, NEJM 2018) as well as head and neck (Ferris, NEJM 2016). However, primary resistance rates range from 35-44% in patients with non-small cell lung cancer (NSCLC) while secondary resistance rates approach 100% (Reck, NEJM 2016).

[0013] There are several types of tumors that are considered immunologically “cold” tumors, i.e., tumors that are not likely to trigger a strong immune response and that usually respond poorly to cancer immunotherapies such as ICB therapy (see, e.g., Bonaventura et al., “Cold Tumors: A Therapeutic Challenge for Immunotherapy.” Frontiers in immunology vol. 10 168 (2019). These tumors are notably characterized by the absence of infiltration of tumor antigen-specific immune cells such as tumor-infiltrating lymphocytes (TILs) in the tumor.

[0014] Therefore, there is a need for the development of novel approaches to induce or stimulate the immune response against tumors, notably immunologically cold tumors and / or tumors resistant to immunotherapies.

[0015] The present description refers to a number of documents, the content of which is herein incorporated by reference in their entirety.

[0016] SUMMARY

[0017] In various aspects and embodiments, the present disclosure provides the following items

[0018] 1 to 78:

[0019] 1 . A method for (i) enhancing the anti-tumor immune response in a subject suffering from a Sortilin-expressing cancer and / or (ii) treating a subject suffering from a Sortilin-expressing cancer resistant to an immunotherapy, the method comprising administering to the subject an effective amount of a conjugate compound or a pharmaceutically acceptable salt thereof, wherein the conjugate compound has the formula of A-(B)n, wherein

[0020] A is a peptide compound of 30 residues or less comprising an amino acid sequence having at least 60% sequence identity to the amino acid sequence of any one of SEQ ID NOs: 1- 13:

[0021] X1X2X8X4X5GVX6AKAGVX7NX8FKSESY (SEQ ID NO: 1)

[0022] (X9)nGVX10AKAGVX11NX12FKSESY (SEQ ID NO: 2)

[0023] YKX13LRRX14APRWDX15PLRDPALRX16X17L (SEQ ID NO: 3)

[0024] YKX18LRR(X19)NPLRDPALRX20X21L (SEQ ID NO: 4)

[0025] IKLSGGVQAKAGVINMDKSESM (SEQ ID NO: 5)

[0026] IKLSGGVQAKAGVINMFKSESY (SEQ ID NO: 6)

[0027] I KLSGG VQAKAG I NM FKSESYK (SEQ ID NO: 7) GVQAKAGVINMFKSESY (SEQ ID NO: 8)

[0028] GVRAKAGVRNMFKSESY (SEQ ID NO: 9)

[0029] GVRAKAGVRN(Nle)FKSESY (SEQ ID NO: 10)

[0030] YKSLRRKAPRWDAPLRDPALRQLL (SEQ ID NO: 1 1)

[0031] YKSLRRKAPRWDAYLRDPALRQLL (SEQ ID NO: 12)

[0032] YKSLRRKAPRWDAYLRDPALRPLL (SEQ ID NO: 13) wherein

[0033] Xi , X2, X3, X4, X5, Xe, X7, Xs, X9, X10, X11 , X12, X13, X14, Xis, Xis and X19 are independently chosen from any amino acid;

[0034] X16, X17, X20 and X21 are independently chosen from Q, P, Y, I and L; n is 0, 1 , 2, 3, 4 or 5; when X9 is present more than once, each of said X9is independently chosen from any amino acid; when X19 is present more than once, each of said X9is independently chosen from any amino acid, optionally the peptide compound is cyclic,

[0035] B is at least one therapeutic agent, wherein B is connected to A directly or via a linker.

[0036] 2. The method of item 1 , wherein the peptide compound comprises or consists of an amino acid sequence having at least 90% sequence identity to the amino acid sequence of any one of SEQ ID NOs: 1-13.

[0037] 3. The method of item 1 or 2, wherein the peptide compound comprises or consists of the amino acid sequence of any one of SEQ ID NOs: 1-13, and further comprises 1 to 3 additional amino acids at its amino- and / or carboxy-terminal end.

[0038] 4. The method of item 3, wherein the peptide compound comprises a cysteine residue at its amino- and / or carboxy-terminal end.

[0039] 5. The method of item 4, wherein the peptide compound comprises or consists of one of the following amino acid sequences:

[0040] Z1X1X2X3X4X5G X6AKAG X7NX8FKSESYZ2 (SEQ ID NO:34)

[0041] ZT (X9)nGVX10AKAGVX11 NX12FKSESYZ2(SEQ ID NO:35)

[0042] Z1YkX13LRRX14APRWDX15PLRDPALRX16X17LZ2 (SEQ ID NO:36)

[0043] Z1YKX18LRR(X19)NPLRDPALRX2OX2ILZ2 (SEQ ID NO:37)

[0044] ZT I KLSG GVQAKAGVI NM DKSESMZ2 (SEQ ID NO:38)

[0045] ZT I KLSG GVQAKAG I N M FKSESYZ2 (SEQ ID NO:39)

[0046] ZT I KLSG GVQAKAGVI N M FKSESYKZ2 (SEQ ID NQ:40)

[0047] Z! GVQAKAGVI NM FKSESYZ2 (SEQ ID NO:41)

[0048] Z1GVRAKAGVRNMFKSESYZ2(SEQ ID NO:42)

[0049] Z! G VRAKAG VRN (N le) F KSESYZ2 (SEQ ID NO:43) Z1YKSLRRKAPRWDAPLRDPALRQLLZ2(SEQ ID NO:44)

[0050] Z1YKSLRRKAPRWDAYLRDPALRQLLZ2(SEQ ID NO:45)

[0051] Z1YKSLRRKAPRWDAYLRDPALRPLL z2(SEQ ID NO:46), wherein X1-X21 are as defined in item 1 ; Z1 is a cysteine residue or is absent; Z2is a cysteine residue or is absent, and at least one of Zi and Z2is present.

[0052] 6. The method of item 1 or 5, wherein the peptide compound comprises or consists of the amino acid sequence of SEQ ID NO: 1 or 2.

[0053] 7. The method of item 1 or 5, wherein the peptide compound comprises or consists of the amino acid sequence of SEQ ID NO: 3 or 4.

[0054] 8. The method of item 1 or 5, wherein the peptide compound comprises or consists of the amino acid sequence of SEQ ID NO: 5.

[0055] 9. The method of item 1 or 5, wherein the peptide compound comprises or consists of the amino acid sequence of SEQ ID NO: 6.

[0056] 10. The method of item 1 or 5, wherein the peptide compound comprises or consists of the amino acid sequence of SEQ ID NO: 7.

[0057] 11. The method of item 1 or 5, wherein the peptide compound comprises or consists of the amino acid sequence of SEQ ID NO: 8.

[0058] 12. The method of item 1 or 5, wherein the peptide compound comprises or consists of the amino acid sequence of SEQ ID NO: 9.

[0059] 13. The method of item 1 or 5, wherein the peptide compound comprises or consists of the amino acid sequence of SEQ ID NO: 10.

[0060] 14. The method of item 13, wherein the peptide compound comprises or consists of the amino acid sequence GVRAKAGVRN(Nle)FKSESYC (SEQ ID NO:47).

[0061] 15. The method of item 1 or 5, wherein the peptide compound comprises or consists of the amino acid sequence of SEQ ID NO: 11 .

[0062] 16. The method of item 1 or 5, wherein the peptide compound comprises or consists of the amino acid sequence of SEQ ID NO: 12.

[0063] 17. The method of item 1 or 5, wherein the peptide compound comprises or consists of the amino acid sequence of SEQ ID NO: 13.

[0064] 18. The method of any one of items 1 to 17, wherein the peptide compound comprises at least one modifying group at its amino- and / or carboxy-terminal end.

[0065] 19. The method of item 17, wherein the at least one modifying group is acetyl or succinyl.

[0066] 20. The method of item 1 , wherein the peptide compound is represented by SEQ ID NO:14, SEQ ID NO:15, SEQ ID NO:16, SEQ ID NO:17, SEQ ID NO:18, or SEQ ID NO:48:

[0067] Acetyl-GVRAKAGVRNMFKSESY (SEQ ID NO: 14)

[0068] Acetyl-GVRAKAGVRN(Nle)FKSESY (SEQ ID NO: 15)

[0069] Acetyl-YKSLRRKAPRWDAPLRDPALRQLL (SEQ ID NO: 16) Acetyl-YKSLRRKAPRWDAYLRDPALRQLL (SEQ ID NO: 17) Acetyl- YKSLRRKAPRWDAYLRDPALRPLL (SEQ ID NO: 18) Acetyl-GVRAKAGVRN(Nle)FKSESYC (SEQ ID NO: 48).

[0070] 21 . The method of any one of items 1 to 20, wherein B is connected to A at a free amine of said peptide compound, at an N-terminal position of said peptide compound, at a free -SH of said peptide compound, and / or at a free carboxyl of said peptide compound.

[0071] 22. The method of any one of items 1 to 21 , wherein B is connected to A via a linker.

[0072] 23. The method of any one of items 1 to 22, wherein the conjugate is represented by formula (bill) or (LIV):

[0073] GVRAK(J1)AGVRN(Nle)FK(J2)SESY (bill) (SEQ ID NO:22);

[0074] Acetyl-GVRAK(J1)AGVRN(Nle)FK(J2)SESY (LIV) (SEQ ID NO:23); wherein J1and J2are each independently a therapeutic agent attached to a lysine (K) residue.

[0075] 24. The method of any one of items 1 to 23, wherein the therapeutic agent is an antitumor agent, for example a radionuclide or a chemotherapeutic agent.

[0076] 25. The method of item 24, wherein the chemotherapeutic agent is a taxane.

[0077] 26. The method of item 25, wherein the chemotherapeutic agent is docetaxel.

[0078] 27. The method of any one of items 1 to 26, further comprising treating the subject with an immunotherapy.

[0079] 28. The method of any one of items 1 to 27, wherein the immunotherapy comprises immune checkpoint inhibitor (ICI) therapy.

[0080] 29. The method of item 28, wherein the ICI is a Programmed cell death-1 (PD-1) inhibitor, a cytotoxic T-lymphocyte-associated antigen 4 (CTLA-4) inhibitor, or a Programmed death-ligand 1 (PD-L1) inhibitor.

[0081] 30. The method of item 28 or 29, wherein the ICI is a blocking antibody.

[0082] 31 . The method of item 28 or 29, wherein the ICI is a PD-L1 inhibitor.

[0083] 32. The method of any one of items 1 to 31 , wherein the cancer is an immunologically cold cancer.

[0084] 33. The method of any one of items 1 to 32, wherein the cancer is brain cancer, pancreatic cancer, ovarian cancer, breast cancer, endometrial cancer, thyroid cancer, lung cancer, prostate cancer or melanoma.

[0085] 34. Use of the conjugate compound or pharmaceutically acceptable salt thereof defined in any one of items 1 to 26 for (i) enhancing the anti-tumor immune response in a subject suffering from a Sortilin-expressing cancer and / or (ii) treating a subject suffering from a Sortilin-expressing cancer resistant to an immunotherapy.

[0086] 35. Use of the conjugate compound or pharmaceutically acceptable salt thereof defined in any one of items 1 to 26 for the manufacture of a medicament for (i) enhancing the anti-tumor immune response in a subject suffering from a Sortilin-expressing cancer and / or (ii) treating a subject suffering from a Sortilin-expressing cancer resistant to an immunotherapy.

[0087] 36. The use of item 34 or 35, wherein the conjugate compound, pharmaceutically acceptable salt thereof, or medicament is for use in combination with an immunotherapy.

[0088] 37. The use of any one of items 34 to 36, wherein the immunotherapy comprises immune checkpoint inhibitor (ICI) therapy.

[0089] 38. The use of item 37, wherein the ICI is a Programmed cell death-1 (PD-1) inhibitor, a cytotoxic T-lymphocyte-associated antigen 4 (CTLA-4) inhibitor, or a Programmed death-ligand 1 (PD-L1) inhibitor.

[0090] 39. The use of item 37 or 38, wherein the ICI is a blocking antibody.

[0091] 40. The use of item 38 or 39, wherein the ICI is a PD-L1 inhibitor.

[0092] 41 . The use of any one of items 34 to 40, wherein the cancer is an immunologically cold cancer.

[0093] 42. The use of any one of items 34 to 41 , wherein the cancer is brain cancer, pancreatic cancer, ovarian cancer, breast cancer, endometrial cancer, thyroid cancer, lung cancer, prostate cancer or melanoma.

[0094] 43. The conjugate compound or pharmaceutically acceptable salt thereof defined in any one of items 1 to 26 for use in (i) enhancing the anti-tumor immune response in a subject suffering from a Sortilin-expressing cancer and / or (ii) treating a subject suffering from a Sortilin-expressing cancer resistant to an immunotherapy.

[0095] 44. The conjugate compound or pharmaceutically acceptable salt thereof for use according to item 43, wherein the peptide compound is for use in combination with an immunotherapy.

[0096] 45. The conjugate compound or pharmaceutically acceptable salt thereof for use according to item 43 or 44, wherein the immunotherapy comprises immune checkpoint inhibitor (ICI) therapy.

[0097] 46. The conjugate compound or pharmaceutically acceptable salt thereof for use according to item 45, wherein the ICI is a Programmed cell death-1 (PD-1) inhibitor, a cytotoxic T-lymphocyte- associated antigen 4 (CTLA-4) inhibitor, or a Programmed death-ligand 1 (PD-L1) inhibitor.

[0098] 47. The conjugate compound or pharmaceutically acceptable salt thereof for use according to item 45 or 46, wherein the ICI is a blocking antibody.

[0099] 48. The conjugate compound or pharmaceutically acceptable salt thereof for use according to item 46 or 47, wherein the ICI is a PD-L1 inhibitor.

[0100] 49. The conjugate compound or pharmaceutically acceptable salt thereof for use according to any one of items 43 to 48, wherein the cancer is an immunologically cold cancer.

[0101] 50. The conjugate compound or pharmaceutically acceptable salt thereof for use according to any one of items 43 to 49, wherein the cancer is brain cancer, pancreatic cancer, ovarian cancer, breast cancer, endometrial cancer, thyroid cancer, lung cancer, prostate cancer or melanoma. 51 . A method for treating a Sortilin-expressing cancer in a subject comprising administering to the subject a therapeutically effective amount of the conjugate compound or pharmaceutically acceptable salt thereof defined in any one of items 1 to 26 in combination with an immunotherapy.

[0102] 52. The method of item 51 , wherein the immunotherapy comprises immune checkpoint inhibitor (ICI) therapy.

[0103] 53. The method of item 52, wherein the ICI is a Programmed cell death-1 (PD-1) inhibitor, a cytotoxic T-lymphocyte-associated antigen 4 (CTLA-4) inhibitor, or a Programmed death-ligand 1 (PD-L1) inhibitor.

[0104] 54. The method of item 52 or 53, wherein the ICI is a blocking antibody.

[0105] 55. The method of item 53 or 54, wherein the ICI is a PD-L1 inhibitor.

[0106] 56. The method of any one of items 51 to 55, wherein the cancer is an immunologically cold cancer.

[0107] 57. The method of any one of item 51 to 56, wherein the cancer is brain cancer, pancreatic cancer, ovarian cancer, breast cancer, endometrial cancer, thyroid cancer, lung cancer, prostate cancer or melanoma.

[0108] 58. The method of any one of item 51 to 57, wherein the conjugate compound or pharmaceutically acceptable salt thereof and the immunotherapy are in distinct compositions.

[0109] 59. The method of any one of item 51 to 57, wherein the conjugate compound or pharmaceutically acceptable salt thereof and the immunotherapy are in the same composition.

[0110] 60. Use of the conjugate compound or pharmaceutically acceptable salt thereof defined in any one of items 1 to 26 in combination with an immunotherapy for the treatment of a Sortilin- expressing cancer.

[0111] 61 . Use of the conjugate compound or pharmaceutically acceptable salt thereof defined in any one of items 1 to 26 in combination with an immunotherapy for the manufacture of a medicament for the treatment of a Sortilin-expressing cancer.

[0112] 62. The use of item 60 or 61 , wherein the immunotherapy comprises immune checkpoint inhibitor (ICI) therapy.

[0113] 63. The use of item 62, wherein the ICI is a Programmed cell death-1 (PD-1) inhibitor, a cytotoxic T-lymphocyte-associated antigen 4 (CTLA-4) inhibitor, or a Programmed death-ligand 1 (PD-L1) inhibitor.

[0114] 64. The use of item 62 or 63, wherein the ICI is a blocking antibody.

[0115] 65. The use of item 63 or 64, wherein the ICI is a PD-L1 inhibitor.

[0116] 66. The use of any one of items 60 to 65, wherein the cancer is an immunologically cold cancer.

[0117] 67. The use of any one of items 61 to 66, wherein the cancer is brain cancer, pancreatic cancer, ovarian cancer, breast cancer, endometrial cancer, thyroid cancer, lung cancer, prostate cancer or melanoma. 68. The use of any one of items 61 to 67, wherein the conjugate compound or pharmaceutically acceptable salt thereof and the immunotherapy are in distinct compositions.

[0118] 69. The use of any one of items 61 to 67, wherein the conjugate compound or pharmaceutically acceptable salt thereof and the immunotherapy are in the same composition.

[0119] 70. A combination therapy comprising the conjugate compound or pharmaceutically acceptable salt thereof defined in any one of items 1 to 26 and an immunotherapy for use in the treatment of a Sortilin-expressing cancer.

[0120] 71. The combination therapy for use according to item 70, wherein the immunotherapy comprises immune checkpoint inhibitor (ICI) therapy.

[0121] 72. The combination therapy for use according to item 71 , wherein the ICI is a Programmed cell death-1 (PD-1) inhibitor, a cytotoxic T-lymphocyte-associated antigen 4 (CTLA-4) inhibitor, or a Programmed death-ligand 1 (PD-L1) inhibitor.

[0122] 73. The combination therapy for use according to item 71 or 72, wherein the ICI is a blocking antibody.

[0123] 74. The combination therapy for use according to item 72 or 73, wherein the ICI is a PD-L1 inhibitor.

[0124] 75. The combination therapy for use according to any one of items 71 to 74, wherein the cancer is an immunologically cold cancer.

[0125] 76. The combination therapy for use according to any one of item 70 to 75, wherein the cancer is brain cancer, pancreatic cancer, ovarian cancer, breast cancer, endometrial cancer, thyroid cancer, lung cancer, prostate cancer or melanoma.

[0126] 77. The combination therapy for use according to any one of item 70 to 76, wherein the conjugate compound or pharmaceutically acceptable salt thereof and the immunotherapy are in distinct compositions.

[0127] 78. The combination therapy for use according to any one of item 70 to 76, wherein the conjugate compound or pharmaceutically acceptable salt thereof and the immunotherapy are in the same composition.

[0128] Other objects, advantages and features of the present invention will become more apparent upon reading of the following non-restrictive description of specific embodiments thereof, given by way of example only with reference to the accompanying drawings.

[0129] BRIEF DESCRIPTION OF DRAWINGS

[0130] In the appended drawings:

[0131] FIGs. 1A-I show the sustained and prolonged antitumor activity of TH 1902 in an immunosuppressed MDA-MB-231 TNBC-derived xenograft model. An in vivo MDA-MB-231 TNBC xenograft model was generated as previously described [5] in immunocompromised nude mice. Mice treatments were performed with intravenous injections of either docetaxel at the MTD of 15 mg / kg / wk, or with TH 1902 at 35 mg / kg / wk and halted four days after either (i) three cycles, or for TH 1902 after (ii) six cycles. An additional group was treated with TH 1902 for (Hi) six cycles on followed by three cycles off. FIG. 1A: Tumor growth was monitored at the indicated days as described in the Examples. Data are represented as mean ± SEM (three mice / group). Representative sections of the tumors were H&E stained (FIG. 1B), or stained for SORT1 , Ki67, STING, and CD45 (FIG. 1C) as described in Example 1. FIG. 1D: Vasculogenic mimicry was assessed by monitoring CD317PAS+ staining, whereas normal vasculature was accounted for by CD31+ / PAS+staining. FIG. 1E: Mice weights were monitored as described in Example 1. Mice weights are expressed as percentage of initial weight upon initiation of treatment (day 0). Data are represented as mean ± SEM (three mice / group). FIGs. 1F and G: Differential effects of TH1902 and docetaxel on p21 and p53 expression in MDA-MB-231 cells. MDA-MB-231 cells were treated with vehicle (DMSO), or 50 nM docetaxel or TH 1902 for 5 minutes, then followed by 96 hours of incubation in fresh complete medium. Cell lysates were harvested as described in Example 1 , and representative immunoblotting was performed with anti-p21 , anti-p53, and anti- GAPDH antibodies (FIG. 1F), and protein expression was quantified using densitometry (FIG. 1F). FIGs. 1H and I: PD-L1 quantification in MDA-MB-231 / Luc tumors in vivo measured with QuPath software. FIG. 1H: % of PD-L1 positive area; FIG. 11: % of PD-L1 positive cells.

[0132] FIGs. 2A-C show SORT 1 expression in melanoma tissues and cell line models. FIG. 2A: Tissue microarrays of healthy tissues and of clinically annotated melanomas from stages ll-IV were assessed for SORT1 expression by immunohistochemistry. FIG. 2B: IHS scoring was performed as described in Example 1 (Normal n=2; Stage II n=2; Stage III n=3; Stage IV n=5). FIG. 2C: Cell lysates (20 pg) from different cancer cell lines were assessed for their SORT1 expression levels by Western blot analysis. B16-F10, murine B16-F10 melanoma cells; SKMEL, human SK-MEL-28 melanoma cells; A375, human A375 melanoma cells; MDA, human TNBC- derived MDA-MB-231 cells.

[0133] FIGs. 3A-G show that TH1902 exerts in vitro anti-proliferative and apoptotic activities and induces senescence in SORT1 -positive melanoma cells. Cell proliferation in response to docetaxel or TH1902 was assessed in SK-MEL-28 (FIG. 3A) and B16-F10 (FIG. 3B) melanoma cells as described in Example 1 , and IC5o values extracted for each test article (FIG. 3C). FIG. 3D: Cell apoptosis was assessed in B16-F10 melanoma cells following docetaxel (black bars) and TH 1902 (grey bars) treatments at the indicated concentrations. FIG. 3E: Fluorescence microscopy was used to assess cellular senescence in B16-F10 melanoma cells following docetaxel, TH1902, or etoposide (as a positive control for senescence induction) treatment and representative pictures are shown. FIG. 3F: Senescence-associated p-galactosidase activity was quantified as described in Example 1 (n=3). FIG. 3G: Changes in cell morphology were assessed using crystal violet staining of cells upon vehicle (DMSO), 100 nM docetaxel, or 50 nM TH1902 treatment (equivalent docetaxel content). FIGs. 4A-E show the infiltration of leukocytes within B16-F 10 tumors treated with vehicle, docetaxel, or TH1902. FIG. 4A: Tumor growth in syngeneic mice treated with vehicle, 15 mg / kg / wk docetaxel (MTD), or 35 mg / kg / wk TH1902 (equivalent docetaxel content). Data are represented as mean ± SEM (9 mice / group for vehicle and docetaxel, 10 mice / group for TH 1902). FIG. 4B: B16-F10 melanoma tumors were excised, and pictures taken. FIG. 4C: Excised tumors were fixed with formalin and processed for immunohistochemistry analysis. The top layer of images was stained with haematoxylin and eosin; the bottom layer was assessed immunohistochemically using a monoclonal antibody against CD45 (pan-immune cells). Representative images of whole and magnified tumors are shown for both stainings (black scale bar = 2 mm, white scale bar = 100 pm). FIG. 4D: The quantities of CD45 stained area were compared for the three groups of mice by one-way ANOVA, followed by Tukey’s multiple comparisons test. The area staining positive for leukocytes was significantly greater in TH 1902- treated animals than in animals treated with vehicle or docetaxel. Data are represented as mean ± SEM (* p <0.05, ** p <0.01 , n = 4 tumors analyzed per group). FIG. 4E: Mice weights in syngeneic mice treated with vehicle, 15 mg / kg / wk docetaxel (MTD), or 35 mg / kg / wk TH1902 (equivalent docetaxel content). Mice weights are expressed as the percent of initial weight upon initiation of treatment (day 0). Data are represented as mean ± SEM (9 mice / group for vehicle and docetaxel, 10 mice / group for TH1902).

[0134] FIGs. 5A-D show the effects of docetaxel and TH1902 on the levels of tumor-infiltrating lymphocytes and macrophages within syngeneic tumors. FIG. 5A: Each row of photomicrographs depicts cells from B16-F10 tumors in animals treated with either vehicle, docetaxel, or TH1902 (same samples as FIGs. 4A-E). Each column includes representative IHC images of immune cell markers CD3 (T-cells), CD8 (cytotoxic T-cells), CD4 (helper T-cells), FoxP3 (regulatory T-cells), and CD161c (NK cells) performed on paraffin sections from primary tumors (white scale bar = 100 pm). FIG. 5B: Quantification of IHC stainings from FIG. 5A. Expression was compared using one-way ANOVA with Tukey’s multiple comparisons test. Data are represented as mean ± SEM (* p <0.05, ** p <0.01 , *** p <0.001 , n = 4 tumors analyzed per group). FIGs. 5C-D are graphs showing the quantification of immune cell infiltration in docetaxel- and TH1902-treated B16-F10 xenograft tumors. FIG. 5C: Natural killer (NK) cells (CD161c+) and FIG.5D: Tumor-Associated Macrophages (TAMs) including M1 (CD68+) and M2 (CD206+) macrophages.

[0135] FIGs. 6A-B show the effects of docetaxel and TH 1902 on expression of tumor markers for immune-stimulated apoptosis. FIG. 6A: Each row of photomicrographs depicts cells from B16- F10 tumors in animals treated with either vehicle, docetaxel, or TH1902 (same samples as Figure 4). Each column includes representative IHC images of markers involved in immune-stimulated apoptosis cleaved caspase-3, perforin, and granzyme B (white scale bar = 100 pm). FIG. 6B: Quantification of IHC stainings from FIG. 6A. Expression was compared using one-way ANOVA with Tukey’s multiple comparisons test. Data are represented as mean ± SEM (* p <0.05, ** p <0.01 , n = 4 tumors analysed per group).

[0136] FIGs. 7A-D show the effects on tumor growth and mice survival of TH 1902 in combination with a checkpoint inhibitor on tumor growth and mice survival. B16-F10 cells were subcutaneously implanted in immunocompetent C57BL / 6 mice. FIG. 7A: Effect on B16-F10 tumor growth of docetaxel, TH1902, and anti-PD-L1 alone or in combination. Mice were treated weekly via intravenous administration of either vehicle, docetaxel (7.5 mg / kg), or TH1902 (17.5 mg / kg; equivalent dose of docetaxel) or bi-weekly via intraperitoneal administration of either anti-PD-L1 (9 mg / kg) and control isotype (9 mg / kg) alone or in combination (docetaxel / anti-PD-L1 or TH1902 / anti-PD-L1) for two cycles of treatment. Data are represented as mean ± SEM (n=8 mice / group). FIG. 7B and C: Effect on survival (FIG. 7B) and tumor growth (FIG. 7C) of increased doses of TH1902 and anti-PD-L1 alone or in combination in B16-F10 tumor-bearing mice. Mice were treated weekly via intravenous administration with either vehicle and TH1902 (4.37, 8.75, 17.5 mg / kg) or bi-weekly via intraperitoneal administration of either anti-PD-L1 (9 mg / kg) and control isotype (9 mg / kg) alone or in combination for continuous cycles of treatment until one of the defined endpoints was reached as described in Example 1 . Kaplan-Meier curves were plotted to estimate mice survival and presented as probability of survival in percentage (upper graph) whereas tumor growth curves were plotted until one individual within a given group reached the tumor size endpoint (tumor >2,000 mm3; lower graph). Data are represented as mean ± SEM (n=6 mice / group). FIG. 7D: Mice weights are expressed as the percent of initial weight upon initiation of treatment (day 0). Data are represented as mean ± SEM (n=8 mice / group).

[0137] FIG. 8 depicts graphs showing the effect of various checkpoint inhibitors on B16-F10 melanoma tumor growth (corresponding to Figure 1A from Ueha et al., Cancer Immunol Res (2015) 3 (6): 631-640). Mice bearing B16-F10 melanoma tumors were injected intraperitoneally (i.p.) with anti-CD4 mAb (200 mg / mouse) on day 5 and 9, and with anti-immune checkpoint mAbs (anti-PD1 , anti-PD-L1 , anti-PD-L2, anti-CTLA-4, anti-OX40, anti-LAG-3, anti-TIM-3, anti-BTLA or anti-GITR) on day 4, 8, 14, and 18 after tumor inoculation.

[0138] FIGs. 9A-F show that TH1902 induces downstream effectors of the STING pathway and the cell surface expression of PD-L1 and MHC-I in B16-F10 melanoma cells. Cells were treated with increasing docetaxel or TH 1902 concentrations for 5 minutes, followed by 96 hours of incubation in fresh complete medium. Cell lysates were harvested as described in Example 1 and immunoblotting was performed with anti-STING and anti-GAPDH antibodies (FIG. 9A), and protein expression was quantified using densitometry (FIG. 9B). FIG. 9C: Lysates were further processed for the indicated expression of STING downstream effectors at 100 nM docetaxel or TH1902, and densitometric quantification was performed (FIG. 9D). FIG. 9E: Total RNA was extracted and gene expression of IL-6 and TNFa assessed using RT-qPCR in cells treated or not with 100 nM docetaxel or TH 1902 for 5 minutes, followed by 24 hours of incubation in fresh complete medium. FIG. 9F: Immunophenotyping of MHC-I and PD-L1 cell surface expression was performed by flow cytometry as described in Example 1 .

[0139] FIGs. 10A-B show that TNFa, but not IL-6, triggers PD-L1 and MHC-1 gene expression in B16-F10 melanoma cells. Cells were treated with the indicated TNFa or IL-6 concentrations for 96 hours. Levels of PD-L1 and MHC-I expression were estimated by flow cytometry as described in the Methods section. PD-L1 and MHC-I cell surface expression upon TNFa treatment (FIG. 10A), or upon IL-6 treatment (FIG. 10B).

[0140] FIGs. 11 A-B depict graphs showing the effect of treatment with the checkpoint inhibitors anti-PD1 (FIG. 11 A) and anti-PD-L1 (FIG. 11B) in the LL / 2 (Lewis Lung) lung carcinoma model (from https: / / drugdevelopment.labcorp.com / industry-solutions / oncology / preclinical / tumor- spotlights / ll-2-an-immunosuppressive-murine-tumor-model.html, Figure 5).

[0141] FIG. 11C shows the effects of TH1902 and Docetaxel on LL / 2 lung cancer xenografts. LL / 2 cells were subcutaneously implanted in the dorsal area of C57BL / 6 mice and treated 3 days after. Tumor volume was measured following weekly administrations of vehicle, docetaxel (15 mg / kg) or TH1902 (35 mg / kg). Data are represented as mean ± SEM (n = 7 mice / group. Exponential curve analysis (extra sum-of squares F test) show that all curves are significantly different for one another.

[0142] FIG. 12 depicts the amino acid sequence of human Sortilin-1 (SORT1 , UniProtKB accession No. Q99523).

[0143] DETAILED DISCLOSURE

[0144] The use of the terms "a" and "an" and "the" and similar referents in the context of describing the invention (especially in the context of the following claims) are to be construed to cover both the singular and the plural, unless otherwise indicated herein or clearly contradicted by context.

[0145] The terms "comprising", "having", "including", and "containing" are to be construed as open-ended terms (i.e., meaning "including, but not limited to") unless otherwise noted.

[0146] Recitation of ranges of values herein are merely intended to serve as a shorthand method of referring individually to each separate value falling within the range, unless otherwise indicated herein, and each separate value is incorporated into the specification as if it were individually recited herein. All subsets of values within the ranges are also incorporated into the specification as if they were individually recited herein.

[0147] All methods described herein can be performed in any suitable order unless otherwise indicated herein or otherwise clearly contradicted by context.

[0148] The use of any and all examples, or exemplary language (“e.g.”, "such as", etc.) provided herein, is intended merely to better illustrate the invention and does not pose a limitation on the scope of the invention unless otherwise claimed. No language in the specification should be construed as indicating any non-claimed element as essential to the practice of the invention.

[0149] Herein, the term "about" has its ordinary meaning. The term “about” is used to indicate that a value includes an inherent variation of error for the device or the method being employed to determine the value, or encompass values close to the recited values, for example within 10% of the recited values (or range of values).

[0150] Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention belongs.

[0151] The scope of the claims should not be limited by the preferred embodiments set forth in the examples, but should be given the broadest interpretation consistent with the description as a whole.

[0152] Any and all combinations and subcombinations of the embodiments and features disclosed herein are encompassed by the present invention.

[0153] In the studies described herein, the present inventors have demonstrated in a murine model of an immunologically cold tumor that a sortilin-targeting peptide conjugated to a chemotherapeutic agent (docetaxel) was able to induce antitumor immune cell infiltration in the tumor and inhibit tumor growth, and to enhance the antitumor response of an immune checkpoint inhibitor.

[0154] Accordingly, in a first aspect, the present disclosure provides a method for (i) enhancing the anti-tumor immune response in a subject suffering from cancer and / or (ii) treating a subject suffering from a cancer resistant to an immunotherapy (e.g., immune checkpoint inhibitor therapy), the method comprising administering to the subject an effective amount of a conjugate compound defined herein or a pharmaceutically acceptable salt thereof. The present disclosure also provides the use of a conjugate compound defined herein or a pharmaceutically acceptable salt thereof for (i) enhancing the anti-tumor immune response in a subject suffering from cancer and / or (ii) treating a subject suffering from a cancer resistant to an immunotherapy (e.g., immune checkpoint inhibitor therapy). The present disclosure also provides the use of a conjugate compound defined herein or a pharmaceutically acceptable salt thereof for the manufacture of a medicament for (i) enhancing the anti-tumor immune response in a subject suffering from cancer and / or (ii) treating a subject suffering from a cancer resistant to an immunotherapy (e.g., immune checkpoint inhibitor therapy). The present disclosure also provides the conjugate compound defined herein or a pharmaceutically acceptable salt thereof for use in (i) enhancing the anti-tumor immune response in a subject suffering from cancer and / or (ii) treating a subject suffering from a cancer resistant to an immunotherapy (e.g., immune checkpoint inhibitor therapy).

[0155] In an embodiment, the method or use is for enhancing the anti-tumor immune response in a subject suffering from cancer. In another embodiment, the method or use is for treating a subject suffering from a cancer resistant to an immunotherapy (e.g., immune checkpoint inhibitor therapy). In another embodiment, the method or use is for enhancing the anti-tumor immune response in a subject suffering from a cancer resistant to an immunotherapy (e.g., immune checkpoint inhibitor therapy).

[0156] The conjugates suitable for the methods and uses disclosed herein are conjugates comprising therapeutic agents such as antitumor agents (e.g., chemotherapeutic agents) conjugated to agents capable of binding to Sortilin and being internalized by tumor cells such as peptides so as to deliver the antitumor agent to the tumor cells. In an embodiment, the conjugate (or conjugate compound) comprises an anti-sortilin antibody or an antigen-binding fragment thereof conjugated to an antitumor agent (i.e., an antibody-drug conjugate, ADC). Examples of anti-sortilin antibodies and antigen-binding fragments thereof include anti-sortilin monoclonal antibody clone 2D8-E3 (Ghaemimanesh F. et al., Monoclon Antib Immunodiagn Immunother 2015;34(6):390-5), the antibodies and antigen-binding fragments thereof disclosed in PCT publications Nos. WO 2016 / 164637, WO 2017 / 009327, WO 2019 / 016247, WO 2020 / 014617, WO 2020 / 252066, WO 2021 / 116290, WO 2021 / 263279, WO 2022 / 261648, and WO 2023 / 122766, the content of which is incorporated by reference herein.

[0157] Peptide compounds that bind to sortilin are disclosed in PCT publications Nos. WO 2017 / 088058, WO 2018 / 213928, WO 2020 / 037434, WO 2021 / 078833, WO 2021 / 028404, and WO 2023 / 165476, the content of which is incorporated by reference herein.

[0158] In an embodiment, the conjugate (or conjugate compound) is an antitumor agent-peptide compound conjugate as described in PCT publications Nos. WO 2017 / 088058, WO 2018 / 213928 and WO 2020 / 037434.

[0159] The term “Sortilin” or “Sortilin receptor” as used herein refers to a neuronal type-1 membrane glycoprotein, encoded by the SORT 1 gene, belonging to the Vacuolar Protein Sorting 10 protein (Vps10) family of receptors. Sortilin (also known as the neurotensin receptor 3; UniProtKB Accession number Q99523) is expressed or overexpressed in a number of cancers including for example ovarian, breast, colon and prostate cancer. The encoded preproprotein (residues 34-831 , residues 1-33 corresponding to the signal peptide) is proteolytically processed after amino acid 77 by furin (or other homologous proteases) to generate the mature receptor with a molecular weight of about 100-110 kDa (residues 78-831). Amino acid residues of sortilin referenced herein correspond to positions in the full-length form (j.e., UniProtKB Accession number Q99523, FIG. 12).

[0160] In an embodiment, the conjugate compound is of the formula A-(B)n, wherein

[0161] A is a peptide compound comprising an amino acid sequence having at least 60% sequence identity to one of the sequences of formulas (l)-(XIII):

[0162] X1X2X3X4X5GVX6AKAGVX7NX8FKSESY (I) (SEQ ID NO: 1)

[0163] (X9)nGVXioAKAGVXiiNXi2FKSESY (II) (SEQ ID NO: 2) YkXisLRRX PRWDXisPLRDPALRXieXlyL (III) (SEQ ID NO: 3)

[0164] YKX18LRR(X19)NPLRDPALRX20X21L (IV) (SEQ ID NO: 4) IKLSGGVQAKAGVINMDKSESM (V) (SEQ ID NO: 5) IKLSGGVQAKAGVINMFKSESY (VI) (SEQ ID NO: 6) IKLSGGVQAKAGVINMFKSESYK (VII) (SEQ ID NO: 7)

[0165] G VQ AKAG VI N M F KSESY (VIII) (SEQ ID NO: 8) GVRAKAGVRNMFKSESY (IX) (SEQ ID NO: 9)

[0166] GVRAKAGVRN(Nle)FKSESY (X) (SEQ ID NO: 10) YKSLRRKAPRWDAPLRDPALRQLL (XI) (SEQ ID NO: 1 1) YKSLRRKAPRWDAYLRDPALRQLL (XII) (SEQ ID NO: 12) YKSLRRKAPRWDAYLRDPALRPLL (XIII) (SEQ ID NO: 13) wherein

[0167] Xi , X2, X3, X4, X5, Xe, X7, X8, X9, X10, X11 , Xi2, X13, X14, Xis, X-i8and X19 are independently chosen from any amino acid; Xi6, X17, X20and X2iare independently chosen from Q, P, Y, I and L; n is an integer from 1 to 10, for example 1 , 2, 3, 4 or 5; when X9is present more than once, each of said X9is independently chosen from any amino acid; when X19 is present more than once, each of said X9is independently chosen from any amino acid, optionally the peptide compound is cyclic, and wherein the peptide compound binds to Sortilin.

[0168] B is at least one therapeutic agent such as an antitumor agent, wherein B is connected to A either directly or via a linker, optionally at a free amine of said peptide compound, at an N- terminal position of said peptide compound, at a free -SH of said peptide compound, or at a free carboxyl of said peptide compound, or a pharmaceutically acceptable salt thereof.

[0169] The term “amino acid” refers to the common natural (genetically encoded) or synthetic amino acids and common derivatives thereof, known to those skilled in the art. When applied to amino acids, “standard” or “proteinogenic” refers to the genetically encoded 20 amino acids in their natural configuration. Similarly, when applied to amino acids, “non-standard,” “unnatural” or “unusual” refers to the wide selection of non-natural, rare or synthetic amino acids such as those described by Hunt, S. in Chemistry and Biochemistry of the Amino Acids, Barrett, G.C., ed., Chapman and Hall: New York, 1985. Some examples of non-standard amino acids include nonalpha amino acids and D-amino acids. In an embodiment, the peptide compound comprises only natural amino acids. In another embodiment, the peptide compound comprises one or more nonnatural or synthetic amino acids, such as D-amino acids.

[0170] The expression "sequence identity" as used herein refers to the percentage of sequence identity between two polypeptide sequences or two nucleic acid sequences. To determine the percent identity of two amino acid sequences or of two nucleic acid sequences, the sequences are aligned for optimal comparison purposes (e.g., gaps can be introduced in the sequence of a first amino acid or nucleic acid sequence for optimal alignment with a second amino acid or nucleic acid sequence). The amino acid residues or nucleotides at corresponding amino acid positions or nucleotide positions are then compared. When a position in the first sequence is occupied by the same amino acid residue or nucleotide as the corresponding position in the second sequence, then the molecules are identical at that position. The percent identity between the two sequences is a function of the number of identical positions shared by the sequences ( / .e., % identity=number of identical overlapping positions / total number of positions x 100%). In one embodiment, the two sequences are the same length. The determination of percent identity between two sequences can also be accomplished using a mathematical algorithm. BLAST protein searches can be performed with the XBLAST program parameters set, e.g., to score-50, wordlength=3 to obtain amino acid sequences homologous to a protein molecule of the present disclosure. To obtain gapped alignments for comparison purposes, Gapped BLAST can be utilized. Alternatively, PSI- BLAST can be used to perform an iterated search which detects distant relationships between molecules (Id.). When utilizing BLAST, Gapped BLAST, and PSI-Blast programs, the default parameters of the respective programs (e.g., of XBLAST and NBLAST) can be used (see, e.g., the NCBI website). Another preferred, non-limiting example of a mathematical algorithm utilized for the comparison of sequences is the algorithm of Myers and Miller, 1988, CABIOS 4:11-17. Such an algorithm is incorporated in the ALIGN program (version 2.0) which is part of the GCG sequence alignment software package. When utilizing the ALIGN program for comparing amino acid sequences, a PAM 120 weight residue table, a gap length penalty of 12, and a gap penalty of 4 can be used. The percent identity between two sequences can be determined using techniques similar to those described above, with or without allowing gaps. In calculating percent identity, typically only exact matches are counted.

[0171] The expression “pharmaceutically acceptable” means compatible with the treatment of subjects such as animals or humans. Also provided herein is a pharmaceutically acceptable salt of a conjugate compound described herein. The expression “pharmaceutically acceptable salt” means an acid addition salt or basic addition salt which is suitable for or compatible with the treatment of subjects such as animals or humans. The expression “pharmaceutically acceptable acid addition salt” as used herein means any non-toxic organic or inorganic salt of any compound of the present disclosure, or any of its intermediates. Illustrative inorganic acids which form suitable salts include hydrochloric, hydrobromic, sulfuric and phosphoric acids, as well as metal salts such as sodium monohydrogen orthophosphate and potassium hydrogen sulfate. Illustrative organic acids that form suitable salts include mono-, di-, and tricarboxylic acids such as glycolic, lactic, pyruvic, malonic, succinic, glutaric, fumaric, malic, tartaric, citric, ascorbic, maleic, benzoic, phenylacetic, cinnamic and salicylic acids, as well as sulfonic acids such as p-toluenesulfonic and methanesulfonic acids. Either the mono or di-acid salts can be formed, and such salts may exist in either a hydrated, solvated or substantially anhydrous form. In general, the acid addition salts of the compounds of the present disclosure are more soluble in water and various hydrophilic organic solvents, and generally demonstrate higher melting points in comparison to their free base forms. The selection of the appropriate salt will be known to one skilled in the art. Other non- pharmaceutically acceptable salts, e.g., oxalates, may be used, for example, in the isolation of the compounds of the present disclosure, for laboratory use, or for subsequent conversion to a pharmaceutically acceptable acid addition salt. The expression “pharmaceutically acceptable basic addition salt” as used herein means any non-toxic organic or inorganic base addition salt of any acid compound of the disclosure, or any of its intermediates. Acidic compounds of the disclosure that may form a basic addition salt include, for example, where CO2H is a functional group. Illustrative inorganic bases which form suitable salts include lithium, sodium, potassium, calcium, magnesium or barium hydroxide. Illustrative organic bases which form suitable salts include aliphatic, alicyclic or aromatic organic amines such as methylamine, trimethylamine and picoline or ammonia. The selection of the appropriate salt will be known to a person skilled in the art. Other non-pharmaceutically acceptable basic addition salts may be used, for example, in the isolation of the compounds or conjugate compounds of the disclosure, for laboratory use, or for subsequent conversion to a pharmaceutically acceptable acid addition salt.

[0172] In an embodiment, the peptide compound comprises or consists of the sequences of any one of formulas (l)-(XIII). In an embodiment, the peptide compound comprises or consists of the amino acid sequence of any one of SEQ ID NOs: 1-13. In embodiments, the peptide compound comprises 50, 45, 40, 35, 30, 25, or 20 amino acids or less.

[0173] In embodiments, the peptide compound comprises or consists of an amino acid sequence having at least 60%, at least 61%, at least 62%, at least 63%, at least 64%, at least 65%, at least 66%, at least 67%, at least 68%, at least 69%, at least 70%, at least 71%, at least

[0174] 72%, at least 73%, at least 74%, at least 75%, at least 76%, at least 77%, at least 78%, at least

[0175] 79%, at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least

[0176] 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least

[0177] 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% sequence identity to a peptide compound represented by formula (I) or SEQ ID NO: 1 , wherein the peptide compound binds to Sortilin.

[0178] In embodiments, the peptide compound comprises or consists of an amino acid sequence having at least 60%, at least 61%, at least 62%, at least 63%, at least 64%, at least 65%, at least 66%, at least 67%, at least 68%, at least 69%, at least 70%, at least 71%, at least

[0179] 72%, at least 73%, at least 74%, at least 75%, at least 76%, at least 77%, at least 78%, at least

[0180] 79%, at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least

[0181] 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least

[0182] 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% sequence identity to a peptide compound represented by formula (II) or SEQ ID NO: 2, wherein the peptide compound binds to Sortilin.

[0183] In embodiments, the peptide compound comprises or consists of an amino acid sequence having at least 60%, at least 61%, at least 62%, at least 63%, at least 64%, at least 65%, at least 66%, at least 67%, at least 68%, at least 69%, at least 70%, at least 71%, at least

[0184] 72%, at least 73%, at least 74%, at least 75%, at least 76%, at least 77%, at least 78%, at least

[0185] 79%, at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least

[0186] 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least

[0187] 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% sequence identity to a peptide compound represented by formula (III) or SEQ ID NO: 3, wherein the peptide compound binds to Sortilin.

[0188] In embodiments, the peptide compound comprises or consists of an amino acid sequence having at least 60%, at least 61%, at least 62%, at least 63%, at least 64%, at least 65%, at least 66%, at least 67%, at least 68%, at least 69%, at least 70%, at least 71%, at least

[0189] 72%, at least 73%, at least 74%, at least 75%, at least 76%, at least 77%, at least 78%, at least

[0190] 79%, at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least

[0191] 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least

[0192] 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% sequence identity to a peptide compound represented by formula (IV) or SEQ ID NO: 4, wherein the peptide compound binds to Sortilin.

[0193] In embodiments, the peptide compound comprises or consists of an amino acid sequence having at least 60%, at least 61%, at least 62%, at least 63%, at least 64%, at least 65%, at least 66%, at least 67%, at least 68%, at least 69%, at least 70%, at least 71%, at least

[0194] 72%, at least 73%, at least 74%, at least 75%, at least 76%, at least 77%, at least 78%, at least

[0195] 79%, at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least

[0196] 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least

[0197] 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% sequence identity to a peptide compound represented by formula (V) or SEQ ID NO: 5, wherein the peptide compound binds to Sortilin.

[0198] In embodiments, the peptide compound comprises or consists of an amino acid sequence having at least 60%, at least 61%, at least 62%, at least 63%, at least 64%, at least 65%, at least 66%, at least 67%, at least 68%, at least 69%, at least 70%, at least 71%, at least

[0199] 72%, at least 73%, at least 74%, at least 75%, at least 76%, at least 77%, at least 78%, at least

[0200] 79%, at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least

[0201] 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least

[0202] 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% sequence identity to a peptide compound represented by formula (VI) or SEQ ID NO: 6, wherein the peptide compound binds to Sortilin.

[0203] In embodiments, the peptide compound comprises or consists of an amino acid sequence having at least 60%, at least 61%, at least 62%, at least 63%, at least 64%, at least 65%, at least 66%, at least 67%, at least 68%, at least 69%, at least 70%, at least 71%, at least

[0204] 72%, at least 73%, at least 74%, at least 75%, at least 76%, at least 77%, at least 78%, at least

[0205] 79%, at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least

[0206] 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least

[0207] 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% sequence identity to a peptide compound represented by formula (VII) or SEQ ID NO: 7, wherein the peptide compound binds to Sortilin.

[0208] In embodiments, the peptide compound comprises or consists of an amino acid sequence having at least 60%, at least 61%, at least 62%, at least 63%, at least 64%, at least 65%, at least 66%, at least 67%, at least 68%, at least 69%, at least 70%, at least 71%, at least

[0209] 72%, at least 73%, at least 74%, at least 75%, at least 76%, at least 77%, at least 78%, at least

[0210] 79%, at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least

[0211] 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least

[0212] 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% sequence identity to a peptide compound represented by formula (VIII) or SEQ ID NO: 8, wherein the peptide compound binds to Sortilin.

[0213] In embodiments, the peptide compound comprises or consists of an amino acid sequence having at least 60%, at least 61%, at least 62%, at least 63%, at least 64%, at least 65%, at least 66%, at least 67%, at least 68%, at least 69%, at least 70%, at least 71%, at least

[0214] 72%, at least 73%, at least 74%, at least 75%, at least 76%, at least 77%, at least 78%, at least

[0215] 79%, at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least

[0216] 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least

[0217] 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% sequence identity to a peptide compound represented by formula (IX) or SEQ ID NO: 9, wherein the peptide compound binds to Sortilin.

[0218] In embodiments, the peptide compound comprises or consists of an amino acid sequence having at least 60%, at least 61%, at least 62%, at least 63%, at least 64%, at least 65%, at least 66%, at least 67%, at least 68%, at least 69%, at least 70%, at least 71%, at least

[0219] 72%, at least 73%, at least 74%, at least 75%, at least 76%, at least 77%, at least 78%, at least

[0220] 79%, at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least

[0221] 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least

[0222] 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% sequence identity to a peptide compound represented by formula (X) or SEQ ID NO: 10, wherein the peptide compound binds to Sortilin.

[0223] In embodiments, the peptide compound comprises or consists of an amino acid sequence having at least 60%, at least 61%, at least 62%, at least 63%, at least 64%, at least 65%, at least 66%, at least 67%, at least 68%, at least 69%, at least 70%, at least 71%, at least

[0224] 72%, at least 73%, at least 74%, at least 75%, at least 76%, at least 77%, at least 78%, at least

[0225] 79%, at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least

[0226] 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least

[0227] 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% sequence identity to a peptide compound represented by formula (XI) or SEQ ID NO: 11 , wherein the peptide compound binds to Sortilin.

[0228] In embodiments, the peptide compound comprises or consists of an amino acid sequence having at least 60%, at least 61%, at least 62%, at least 63%, at least 64%, at least 65%, at least 66%, at least 67%, at least 68%, at least 69%, at least 70%, at least 71%, at least

[0229] 72%, at least 73%, at least 74%, at least 75%, at least 76%, at least 77%, at least 78%, at least

[0230] 79%, at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least

[0231] 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least

[0232] 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% sequence identity to a peptide compound represented by formula (XII) or SEQ ID NO: 12, wherein the peptide compound binds to Sortilin.

[0233] In embodiments, the peptide compound comprises or consists of an amino acid sequence having at least 60%, at least 61%, at least 62%, at least 63%, at least 64%, at least 65%, at least 66%, at least 67%, at least 68%, at least 69%, at least 70%, at least 71%, at least

[0234] 72%, at least 73%, at least 74%, at least 75%, at least 76%, at least 77%, at least 78%, at least

[0235] 79%, at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least

[0236] 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least

[0237] 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% sequence identity to a peptide compound represented by formula (XIII) or SEQ ID NO:13, wherein the peptide compound binds to Sortilin.

[0238] In an embodiment, the peptide compound comprises 30, 25 or 20 residues or less and comprises the sequence GVRAKAGVRN(Nle)FKSESY (SEQ ID NO: 10). In another embodiment, the peptide compound comprises 30, 25 or 20 residues or less and comprises the sequence GVRAKAGVRN(Nle)FKSESYC (SEQ ID NO:31).

[0239] In an embodiment, at least one modifying group is connected to said peptide compound at an N- and / or C-terminal end. In an embodiment, the peptide compound comprises a modifying group at the N-terminal end. In an embodiment, the peptide compound comprises a modifying group at the C-terminal end. Such modifying groups may be useful for protecting the peptide compound from modification or degradation (e.g., protease degradation). In an embodiment, the amino-terminal modifying group is a C1-C16 or C8-Ci6 acyl group (linear or branched, saturated or unsaturated), in a further embodiment, a saturated Ci-C6acyl group (linear or branched) or an unsaturated C8-C6 acyl group (linear or branched). In a further embodiment, the amino-terminal modifying group is an acetyl group (CH8-CO-, Ac) or a succinyl group (CO-CH2-CH2-CO-). The carboxy-terminal modifying group may be, e.g., a hydroxylamine group (NHOH) attached to the carboxyl group (-C(=O)-NHOH), or an amine attached to the carboxyl group (-C(=O)-NRR’), the amine being a primary, secondary or tertiary amine, and preferably the amine is an aliphatic amine preferably of one to ten carbons, such as methyl amine, iso-butylamine, iso-valerylamine or cyclohexylamine, an aromatic amine or an arylalkyl amine, such as aniline, napthylamine, benzylamine, cinnamylamine, or phenylethylamine, a preferred amine being -NH2.

[0240] In an embodiment, a succinyl group is connected to the peptide compound. For example, the peptide compound has the sequence of Succinyl-IKLSGGVQAKAGVINMFKSESY, corresponding to SEQ ID NO: 6 and having a succinyl group attached thereto at the N-terminal end.

[0241] In an embodiment, an acetyl group is connected to the peptide compound. For example, the peptide compound has the sequence of Acetyl-GVRAKAGVRNMFKSESY (SEQ ID NO: 14). For example, the peptide compound has the sequence of Acetyl-GVRAKAGVRN(Nle)FKSESY (SEQ ID NO: 15). For example, the peptide compound has the sequence of Acetyl- YKSLRRKAPRWDAPLRDPALRQLL (SEQ ID NO:16). For example, the peptide compound has the sequence of Acetyl-YKSLRRKAPRWDAYLRDPALRQLL (SEQ ID NO:17). For example, the peptide compound has the sequence of Acetyl-YKSLRRKAPRWDAYLRDPALRPLL (SEQ ID NO:18).

[0242] In an embodiment, the peptide compounds can be modified at the C- and / or N-terminal by the addition of one or more (e.g., 1 to 5 or 1 to 3) amino acid residues in order to obtain or increase preferential binding sites at the peptide terminal end. For example, the amino acid can be cysteine. For example, the amino acid can be lysine. For example, the amino acid can be cysteine added at the N- and / or C-terminal of a peptide. In an embodiment, the peptide compound is modified by the addition of cysteine at the C-terminal.

[0243] Thus, in embodiments, the peptide compound comprises or consists of one the following amino acid sequences:

[0244] Z1X1X2X3X4X5GVX6AKAGVX7NX8FKSESYZ2 (SEQ ID NO:34)

[0245] ZT (X9)nGVXioAKAGVXi 1 NX12FKSESYZ2(SEQ ID NO:35)

[0246] Z1YkX13LRRX14APRWDX15PLRDPALRX16X17LZ2(SEQ ID NO:36)

[0247] Z1YKX18LRR(X19)NPLRDPALRX20X21LZ2(SEQ ID NO:37)

[0248] ZT I KLSGGVQAKAG I NM DKSESMZ2(SEQ ID NO:38)

[0249] ZT I KLSG G VQAKAG VI N M FKSESYZ2(SEQ ID NO:39) ZT I KLSG G VQAKAG VI N M FKSESYKZ2(SEQ ID NO:40)

[0250] Z! G QAKAG I NM FKSESYZ2(SEQ ID N0:41)

[0251] Z1G RAKAG RNMFKSESYZ2(SEQ ID NO:42)

[0252] Z! G VRAKAG VRN (N le) F KSESYZ2(SEQ ID NO:43)

[0253] Z1YKSLRRKAPRWDAPLRDPALRQLLZ2(SEQ ID NO:44)

[0254] Z1YKSLRRKAPRWDAYLRDPALRQLLZ2(SEQ ID NO:45)

[0255] Z1YKSLRRKAPRWDAYLRDPALRPLLZ2(SEQ ID NO:46) wherein X1-X21 are as previously defined; Z1 is a cysteine residue or is absent; and Z2is a cysteine residue or is absent. In an embodiment, Z1 is absent and Z2is a cysteine residue. In embodiments, at least one of Z1 and Z2is present. In an embodiment, Z2is absent and Z1 is a cysteine residue. In an embodiment, Z1 is a cysteine residue and Z2is a cysteine residue.

[0256] In a specific embodiment, the peptide compound has the sequence of GVRAKAGVRN(Nle)FKSESYC (SEQ ID NO:47) or Acetyl-GVRAKAGVRN(Nle)FKSESYC (SEQ ID NO:48) corresponding to SEQ ID NO: 10 and SEQ ID NO: 15, respectively, modified by the addition of a cysteine residue at the C-terminal.

[0257] The conjugate compound may comprise, for example, from 1 to 10 or from 1 to 5 (e.g., 1 , 2, 3 or 4) molecules of a therapeutic agent (e.g., antitumor agent) connected thereto. These molecules of therapeutic agent can be the same or different, e.g., 2, 3, 4 or more different therapeutic agents could be connected to the peptide compounds. The therapeutic agent(s) are connected to the peptide compound via at least one covalent bond, at least one atom or at least one linker. In an embodiment, at least 2 molecules of a therapeutic agent are attached to A. In an embodiment, the at least two molecules are molecules of the same therapeutic agent, e.g., chemotherapeutic agent.

[0258] The antitumor agent may be any compound that has the ability to inhibit the growth and / or kill tumor cells and includes, for example, small molecules, peptides, proteins, oligonucleotides (e.g., siRNA, shRNA), radionuclide agents, antibodies, as well as drug delivery systems including nanoparticles, liposomes, nanotubes, graphene particles loaded with a therapeutic antitumor agent.

[0259] In an embodiment, the antitumor agent is a chemotherapeutic agent. The term “chemotherapeutic agent” refers to agents that kill tumor cells and / or inhibit their proliferation / growth. Examples in chemotherapeutic agents include alkylating agents (e.g., Cyclophosphamide, Ifosfamide, Mechlorethamine, Chlorambucil, Melphalan, Dacarbazine, Nitrosoureas, Temozolomide, Carmustine, Lomustine, Streptozocin, Busulfan, Procarbazine), anthracyclines (e.g., Daunorubicin, Doxorubicin, Epirubicin, Idarubicin, Mitoxantrone, Valrubicin), cytoskeletal disruptors (e.g., taxanes such as Paclitaxel, Docetaxel, Abraxane, Taxotere, cabazitaxel), histone deacetylase inhibitors (e.g., Vorinostat, Romidepsin), topoisomerase I inhibitors (e.g., Irinotecan, Topotecan), topoisomerase II inhibitors (e.g., Etoposide, Teniposide, Tafluposide), kinase inhibitors (e.g., Bortezomib, Erlotinib, Gefitinib, Imatinib, Vemurafenib, Vismodegib, Dasatinib, Nilotinib, Osimertinib, Crizotinib, Dabrafenib, Vemurafenib, Trametinib, Ibrutinib), nucleotide analogs and precursor analogs (e.g., Azacitidine, Azathioprine, Capecitabine, Cytarabine, Doxifluridine, Fluorouracil (5-FU), Gemcitabine, Hydroxyurea, Mercaptopurine, Methotrexate, Tioguanine (Thioguanine)), peptide antibiotics (e.g., Bleomycin, Actinomycin), platinum-based agents (e.g., Carboplatin, Cisplatin, Oxaliplatin), retinoids (Tretinoin, Alitretinoin, Bexarotene), mitotic inhibitors such as vinca alkaloids and derivative (e.g., Vinblastine, Vincristine, Vindesine, Vinorelbine), toxins such as Maytansinoids, Auristatins, Calicheamicins, Amatoxin or Amanitin, as well as natural phytochemicals having anti-tumor properties such as curcumin, Alkaloids (e.g., Chlorogenic acid, Theobromine, Theophylline), Anthocyanins (e.g., Cyanidin, Malvidin, Carotenoids (Beta-Carotene, Lutein, Lycopene), Coumestans, Flavan-3-Ols, Flavonoids (e.g., Epicatechin, Hesperidin, Isorhamnetin, Kaempferol, Myricetin, Naringin, Nobiletin, Proanthocyanidins, Quercetin, Rutin, Tangeretin), Hydroxycinnamic Acids (e.g., Chicoric acid, Coumarin, Ferulic acid, Scopoletin), Isoflavones (e.g., Daidzein, Genistein), Lignans (e.g., Silymarin), Monoterpenes (e.g., Geraniol, Limonene), Organosulfides (e.g., Allicin, Glutathione, lndole-3-Carbinol, Isothiocyanates, Sulforaphane), Damnacanthal, Digoxin, Phytic acid, Phenolic Acids (e.g., Capsaicin, Ellagic Acid, Gallic acid, Rosmarinic acid, Tannic Acid), Phytosterols (e.g., Beta-Sitosterol), Saponins, Stylbenes (e.g., Pterostilbene, Resveratrol), Triterpenoids (e.g., Ursolic acid), Xanthophylls (e.g., Astaxanthin, Beta-Cryptoxanthin), and Monophenols (e.g., Hydroxytyrosol).

[0260] In another embodiment, the antitumor agent is an antibody or antigen-binding fragment thereof that recognizes an antigen expressed by tumor cells.

[0261] In an embodiment, B is connected to A at a free amine of a lysine residue of said peptide compound, optionally via a linker, or at an N-terminal position of said peptide compound, optionally via a linker. In an embodiment, B is connected to A (optionally via a linker) at a cysteine residue added to an end of the peptide compound, for example the C-terminal end.

[0262] In an embodiment, B is connected to A via a linker, optionally a cleavable linker.

[0263] The term “linker” as used herein means a chemical structure connecting a peptide compound disclosed herein to at least one therapeutic agent. The linker can be connected to the peptide compound at different functional groups on the peptide compound. For example, the linker can be connected to the peptide compound at the primary amines (amines (-NH2): this group exists at the N-terminus of each polypeptide chain (called the alpha-amine) and in the side chain of lysine (Lys, K) residues (called the epsilon-amine). For example, the linker can be connected to the peptide compound at the carboxyls (-COOH): this group exists at the C-terminus of each polypeptide chain and in the side chains of aspartic acid (Asp, D) and glutamic acid (Glu, E). For example, the linker can be connected to the peptide compound at the Sulfhydryls (-SH): This group exists in the side chain of cysteine (Cys, C). Often, as part of a protein's secondary or tertiary structure, cysteines are joined together between their side chains via disulfide bonds (- S-S-). These must be reduced to sulfhydryls to make them available for crosslinking by most types of reactive groups. For example, the linker can be connected to the peptide compound at the Carbonyls (-CHO): Ketone or aldehyde groups can be created in glycoproteins by oxidizing the polysaccharide post-translational modifications (glycosylation) with sodium meta-periodate.

[0264] The following table summarizes the reactivity class and the chemical group of some of the principal linkers for standard chemical conjugation:

[0265] For example, homobifunctional and heterobifunctional crosslinkers can be used. For example, Disuccinimidyl suberate (DSS) is a homobifunctional crosslinker that has identical amine-reactive NHS-ester groups at either end of a short spacer arm. For example, Sulfosuccinimidyl 4- (N-maleimidomethyl)cyclohexane-l -carboxylate (Sulfo-SMCC) is a heterobifunctional crosslinker that has an amine-reactive sulfo-NHS-ester group at one end and a sulfhydryl reactive maleimide group at the opposite end of a cyclohexane spacer arm. This allows for sequential, two-step conjugation procedures. Among the commercially available homobifunctional cross-linkers are: BSOCOES (Bis(2-[Succinimidooxycarbonyloxy]ethyl) sulfone; DPDPB (1 ,4-Di-(3’-[2pyridyldithio]-propionamido) butane; DSS (disuccinimidyl suberate); DST (disuccinimidyl tartrate); Sulfo DST (sulfodisuccinimidyl tartrate); DSP (dithiobis(succinimidyl propionate); DTSSP (3,3’-Dithiobis(sulfosuccinimidyl propionate); EGS (ethylene glycol bis(succinimidyl succinate)); and BASED (Bis(|3-[4-azidosalicylamido]-ethyl)disulfide iodinatable).

[0266] The peptide compounds may be conjugated through a variety of linkers, e.g., sulfhydryl groups, amino groups (amines), or any appropriate reactive group. The linker can be a covalent bond. The linker group may comprise a flexible arm, e.g., 2, 3, 4, 5, 6, 7, 8, 9, 10, 11 , 12, 13, 14, or 15 carbon atoms.

[0267] Exemplary linkers include, without limitation, pyridinedisulfide, thiosulfonate, vinylsulfonate, isocyanate, imidoester, diazine, hydrazine, thiol, carboxylic acid, multi-peptide linkers, and acetylene. Alternatively other linkers that can be used include BS3[Bis(sulfosuccinimidyl)suberate] (which is a homobifunctional N-hydroxysuccinimide ester that targets accessible primary amines), NHS / EDC (N-hydroxysuccinimide and 1-ethyl-3-(3- dimethylaminopropyl)carbodiimide (NHS / EDC allows for the conjugation of primary amine groups with carboxyl groups), sulfo-EMCS ([N-s-maleimidocaproic acid]hydrazide (sulfo-EMCS are heterobifunctional reactive groups that are reactive toward sulfhydryl and amino groups), hydrazide (most proteins contain exposed carbohydrates and hydrazide is a useful reagent for linking carboxyl groups to primary amines).

[0268] To form covalent bonds, one can use as a chemically reactive group a wide variety of active carboxyl groups (e.g., esters) where the hydroxyl moiety is physiologically acceptable at the levels required to modify the peptide compound. Particular agents include for example N- hydroxysuccinimide (NHS), N-hydroxy-sulfosuccinimide (sulfo-NHS), maleimide-benzoyl- succinimide (MBS), gamma-maleimido-butyryloxy succinimide ester (GMBS), maleimido propionic acid (MPA), maleimido hexanoic acid (MHA), and maleimido undecanoic acid (MUA).

[0269] Primary amines are the principal targets for NHS esters; NHS esters react with primary amines to form covalent amide bonds. Accessible a-amine groups present on the N-termini of proteins and the s-amine of lysine react with NHS esters. Thus, conjugated compounds herein disclosed can include a linker having an NHS ester conjugated to an N-terminal amino of a peptide compound, or to an s-amine of lysine. An amide bond is formed when the NHS ester reacts with primary amines releasing N-hydroxysuccinimide. Succinimide containing reactive groups may be referred to more simply as succinimidyl groups. In some embodiments, the functional group on the peptide compound will be a thiol group and the chemically reactive group will be a maleimido- containing group such as gamma-maleimide-butylamide (GMBA or MPA). Such maleimide- containing groups may be referred to herein as maleido groups.

[0270] Amine-to-amine linkers include NHS esters, imidoesters, and others, examples of which are listed below. _

[0271] The linker may also be a sulfhydryl-to-sulfhydryl linker, such as the maleimides and pyridyldithiols listed below. The linker may be an amine-to-sulfhydryl linker, which includes NHS ester / maleimide compounds. Examples of these compounds are provided below.

[0272] The linker can react with an amino group and a non-selective entity. Such linkers include

[0273] NHS ester / aryl azide and NHS ester / diazirine linkers, examples of which are listed below.

[0274] TMEA and TSAT reach through their maleimide groups with sulfhydryl groups. The hydroxyl groups and carboxy group of THPP can react with primary or secondary amines. Other useful linkers conform to the formula Y=C=N-Q-A-C(O)-Z, where Q is a homoaromatic or heteroaromatic ring system; A is a single bond or an unsubstituted or substituted divalent Ci -30 bridging group, Y is O or S; and Z is Cl, Br, I, N3, N-succinimidyloxy, imidazolyl, 1- benzotriazolyloxy, OAr where Ar is an electron-deficient activating aryl group, or OC(O)R where R is -A-Q-N=C=Y or C4-20 tertiary-alkyl (see U.S. Patent No. 4,680,338). O S or another chemical structure that is able to delocalize the lone pair electrons of the adjacent nitrogen and R4is a pendant reactive group capable of linking R3to a peptide compound (see for example U.S. Patent No. 5,306,809).

[0275] The linker may include at least one amino acid residue and can be a peptide of at least or about 2, 3, 4, 5, 6, 7, 10, 15, 20, 25, 30, 40, or 50 amino acid residues. Where the linker is a single amino acid residue it can be any naturally or non-naturally occurring amino acid (e.g., Gly or Cys). Where the linker is a short peptide, it can be a glycine-rich peptide (which tend to be flexible) such as a peptide having the sequence [Gly-Gly-Gly-Gly-Ser]nwhere n is an integer from 1 to 6, inclusive (see U.S. Patent No. 7,271 ,149) or a serine-rich peptide linker (see U.S. Patent No. 5,525,491). Serine rich peptide linkers include those of the formula [X-X-X-X-Gly]ywhere up to two of the X are Thr, the remaining X are Ser, and y is an integer greater than 1 , for example from 1 to 5, inclusive (e.g., [Ser-Ser-Ser-Ser-Gly]y(SEQ ID NO: 19), where y is an integer equal to or greater than 1 , for example from 1 to 5). Other linkers include rigid linkers (e.g., PAPAP (SEQ ID NO:20) and (PT)nP, where n is 2, 3, 4, 5, 6, or 7) and a-helical linkers (e.g., A(EAAAK)nA (SEQ ID NO:21), where n is 1 , 2, 3, 4, or 5).

[0276] The linker can be an aliphatic linker (e.g., with an amide bond to the polypeptide and an ester bond to the therapeutic agent). Where an aliphatic linker is used, it may vary with regard to length (e.g., C1-C20, C1-C12, Ci-C6) and the chemical moieties it includes (e.g., an amino group or carbamate).

[0277] Examples of suitable amino acid linkers are succinic acid, Lys, Glu, and Asp, or a dipeptide such as Gly-Lys. When the linker is succinic acid, one carboxyl group thereof may form an amide bond with an amino group of the amino acid residue, and the other carboxyl group thereof may, for example, form an amide bond with an amino group of the peptide or substituent. When the linker is Lys, Glu, or Asp, the carboxyl group thereof may form an amide bond with an amino group of the amino acid residue, and the amino group thereof may, for example, form an amide bond with a carboxyl group of the substituent. When Lys is used as the linker, a further linker may be inserted between the E-amino group of Lys and the substituent. The further linker may be succinic acid, which can form an amide bond with the E- amino group of Lys and with an amino group present in the substituent. In one embodiment, the further linker is Glu or Asp (e.g., which forms an amide bond with the E-amino group of Lys and another amide bond with a carboxyl group present in the substituent), that is, the substituent is a N£-acylated lysine residue.

[0278] The linker can also be a branched polypeptide. Exemplary branched peptide linkers are described in U.S. Patent No. 6,759,509.

[0279] The linker can provide a cleavable linkage (e.g., a thioester linkage) or a non-cleavable linkage (e.g., a maleimide linkage). For example, a cytotoxic protein can be bound to a linker that reacts with modified free amines, which are present at lysine residues within the polypeptide and at the amino-terminus of the polypeptide. Thus, linkers useful in the present conjugate compounds can comprise a group that is reactive with a primary amine on the polypeptide or modified polypeptide to which the therapeutic agent moiety is conjugated. More specifically, the linker can be selected from monofluoro cyclooctyne (MFCO), bicyclo[6.1.0]nonyne (BCN), N-succinimidyl- S-acetylthioacetate (SATA), N-succinimidyl-S-acetylthiopropionate (SATP), maleimido and dibenzocyclooctyne ester (a DBCO ester). Useful cyclooctynes, within a given linker, include OCT, ALO, MOFO, DIFO, DIBO, BARAC, DIBAC, and DIMAC.

[0280] The linker may comprise a flexible arm, such as for example, a short arm (<2 carbon chain), a medium-size arm (from 2-5 carbon chain), or a long arm (3-6 carbon chain).

[0281] Click chemistry can also be used for conjugation on a peptide (DBCO, TCO, tetrazine, azide and alkyne linkers). These families of linkers can be reactive toward amine, carboxyl and sulfhydryl groups. In addition, these linkers can also be biotinylated, pegylated, modified with a fluorescent imaging dye, or phosphoramidited for incorporation onto an oligonucleotide sequence.

[0282] In an embodiment, the antitumor agent-peptide compound conjugate is represented by formula (LIII) or (LIV):

[0283] GVRAK(J1)AGVRN(Nle)FK(J2)SESY (LIII) (SEQ ID NO:22);

[0284] Acetyl-GVRAK(J1)AGVRN(Nle)FK(J2)SESY (LIV) (SEQ ID NO:23); wherein J1and J2are each independently an antitumor (e.g., chemotherapeutic) agent attached to the lysine (K) residues.

[0285] In an embodiment, the conjugate compound is GVRAK(curcumin)AGVRN(Nle)FK(curcumin)SESY - Formula (XIV) (SEQ ID NO:24) that comprises the peptide compound having SEQ ID NO: 10 wherein each lysine residue has a curcumin molecule connected thereto; or

[0286] YK(curcumin)SLRRK(curcumin)APRWDAPLRDPALRQLL - Formula (XV) (SEQ ID NO:25) that comprises the peptide compound having SEQ ID NO:11 wherein each lysine residue has a curcumin molecule connected thereto.

[0287] In an embodiment, the conjugate compound is Acetyl- GVRAK(curcumin)AGVRN(Nle)FK(curcumin)SESY - Formula (XVI) (SEQ ID NO:26) that comprises the peptide compound having SEQ ID NO: 15 wherein each lysine residue has a curcumin molecule connected thereto, or Acetyl-

[0288] YK(curcumin)SLRRK(curcumin)APRWDAPLRDPALRQLL - Formula (XVII) (SEQ ID NO:27) that comprises the peptide compound having SEQ ID NO: 16 wherein each lysine residue has a curcumin molecule connected thereto.

[0289] In an embodiment, the conjugate compound is GVRAK(docetaxel)AGVRN(Nle)FK(docetaxel)SESY - Formula (XIX) (SEQ ID NO:28) that comprises the peptide compound having SEQ ID NO: 10 wherein each lysine residue has a docetaxel molecule connected thereto.

[0290] In another embodiment, the conjugate compound is Acetyl- GVRAK(docetaxel)AGVRN(Nle)FK(docetaxel)SESY - Formula (XXIII) (SEQ ID NO:29) that comprises the peptide compound having SEQ ID NO: 15 wherein each lysine residue has a docetaxel molecule connected thereto.

[0291] In an embodiment, the conjugate compound is GVRAK(doxorubicin)AGVRN(Nle)FK(doxorubicin)SESY - Formula (XXVI) (SEQ ID NO:30) that comprises the peptide compound having SEQ ID NO: 10 wherein each lysine residue has a doxorubicin molecule connected thereto.

[0292] In another embodiment, the conjugate compound is Acetyl-GVRAK(doxorubicin)AGVRN(Nle)FK(doxorubicin)SESY - Formula (XXVIII) (SEQ ID NO:31) that comprises the peptide compound having SEQ ID NO:15 wherein each lysine residue has a doxorubicin molecule connected thereto.

[0293] In an embodiment, the conjugate compound is GVRAKAGVRN(Nle)FKSESYC(aldoxorubicin) - Formula (LI) (SEQ ID NO:32) that comprises the peptide compound having SEQ ID NO:47 wherein cysteine residue has an aldoxorubicin molecule connected thereto, or that comprises the peptide compound having SEQ ID NO: 10 wherein a cysteine residue is added to C-terminal of said peptide compound, and wherein the cysteine residue has an aldoxorubicin molecule connected thereto.

[0294] In an embodiment, the conjugate compound is Acetyl- GVRAKAGVRN(Nle)FKSESYC(aldoxorubicin) - Formula (Lil) (SEQ ID NO:33) that comprises the peptide compound having SEQ ID NO:48 wherein cysteine residue has an aldoxorubicin molecule connected thereto, or that comprises the peptide compound having SEQ ID NO: 15 wherein a cysteine residue is added to C-terminal of said peptide compound, and wherein the cysteine residue has an aldoxorubicin molecule connected thereto.

[0295] In an embodiment, the conjugate is administered in the form of a prodrug. The term “prodrug” as used herein refers to a derivative of an active form of a known compound or composition which derivative, when administered to a subject, is gradually converted to the active form to produce a better therapeutic response and / or a reduced toxicity level. In general, prodrugs will be functional derivatives of the compounds disclosed herein which are readily convertible in vivo into the compound from which it is notionally derived. Prodrugs include, without limitation, acyl esters, carbonates, phosphates, and urethanes. These groups are exemplary and not exhaustive, and one skilled in the art could prepare other known varieties of prodrugs. Prodrugs may be, for example, formed with available hydroxy, thiol, amino or carboxyl groups. For example, the available OH and / or NH2in the conjugates of the disclosure may be acylated using an activated acid in the presence of a base, and optionally, in inert solvent (e.g. an acid chloride in pyridine). Some common esters which have been utilized as prodrugs are phenyl esters, aliphatic (C1-C24) esters, acyloxymethyl esters, carbamates and amino acid esters. In certain instances, the prodrugs of the compounds of the disclosure are those in which the hydroxy and / or amino groups in the compounds is masked as groups which can be converted to hydroxy and / or amino groups in vivo. Conventional procedures for the selection and preparation of suitable prodrugs are described, for example, in “Design of Prodrugs” ed. H. Bundgaard, Elsevier, 1985.

[0296] Covalent modifications of the conjugate are included within the scope of this disclosure. Covalent modifications include reacting targeted amino acid residues of the conjugate with an organic derivatizing agent that is capable of reacting with selected side chains or the N- or C- terminal residues of the conjugate. Other modifications include deamidation of glutaminyl and asparaginyl residues to the corresponding glutamyl and aspartyl residues, respectively, hydroxylation of proline and lysine, phosphorylation of hydroxyl groups of seryl or threonyl residues, methylation of the a-amino groups of lysine, arginine, and histidine side chains (T.E. Creighton, Proteins: Structure and Molecular Properties, W.H. Freeman & Co., San Francisco, pp. 79-86 (1983)). Other types of covalent modification of the conjugate included within the scope of this disclosure include linking the conjugate to proteins (e.g., albumin) or to nonproteinaceous polymers, e.g., polyethylene glycol (PEG), polypropylene glycol, or polyoxyalkylenes, which may for example increase the in vivo half-life of the conjugate.

[0297] In an embodiment, the conjugate compound or pharmaceutically acceptable salt thereof disclosed herein is formulated into a pharmaceutical composition. In an embodiment, the pharmaceutical composition further comprises a pharmaceutically acceptable carrier or excipient. Such compositions may be prepared in a manner well known in the pharmaceutical art by mixing the conjugate compound having a suitable degree of purity with one or more optional pharmaceutically acceptable carriers or excipients (see Remington: The Science and Practice of Pharmacy, by Loyd Allen, Jr, 2012, 22ndedition, Pharmaceutical Press; Handbook of Pharmaceutical Excipients, by Rowe et al., 2012, 7thedition, Pharmaceutical Press). The carrier / excipient can be suitable for administration of the conjugate compound by any conventional administration route, for example, for oral, intravenous, parenteral, subcutaneous, intramuscular, intracranial, intraorbital, ophthalmic, intraventricular, intracapsular, intraspinal, intrathecal, epidural, intracisternal, intraperitoneal, intranasal or pulmonary (e.g., aerosol) administration. In an embodiment, the carrier / excipient is adapted for administration of the conjugate compound or salt thereof by the intravenous or subcutaneous route. In an embodiment, the carriers / excipients are adapted for administration of the conjugate compound by the intravenous route. In another embodiment, the carriers / excipients are adapted for administration of the conjugate compound or salt thereof by the subcutaneous route. In another embodiment, the carriers / excipients are adapted for administration of the conjugate compound or salt thereof by the oral route.

[0298] An "excipient" as used herein has its normal meaning in the art and is any ingredient that is not an active ingredient (drug) itself. Excipients include for example binders, lubricants, diluents, fillers, thickening agents, disintegrants, plasticizers, coatings, barrier layer formulations, lubricants, stabilizing agent, release-delaying agents and other components. "Pharmaceutically acceptable excipient" as used herein refers to any excipient that does not interfere with effectiveness of the biological activity of the active ingredients and that is not toxic to the subject, i.e., is a type of excipient and / or is for use in an amount which is not toxic to the subject. Excipients are well known in the art, and the present system is not limited in these respects. In certain embodiments, the composition may include excipients such as one or more binders (binding agents), thickening agents, surfactants, diluents, release-delaying agents, colorants, flavoring agents, fillers, disintegrants / dissolution promoting agents, lubricants, plasticizers, silica flow conditioners, glidants, anti-caking agents, anti-tacking agents, stabilizing agents, anti-static agents, swelling agents and any combinations thereof. As those of skill would recognize, a single excipient can fulfill more than two functions at once, e.g., can act as both a binding agent and a thickening agent. As those of skill will also recognize, these terms are not necessarily mutually exclusive. Examples of commonly used excipients for injectable formulations include water, saline, phosphate buffered saline, dextrose, glycerol, ethanol, and the like, as well as combinations thereof. In many cases, it will be preferable to include isotonic agents, for example, sugars, polyalcohols, such as mannitol, sorbitol, or sodium chloride in the composition. Additional examples of pharmaceutically acceptable substances are wetting agents or auxiliary substances, such as emulsifying agents, preservatives, or buffers, which increase the shelf life or effectiveness.

[0299] The exact amount / dosage of conjugate to be administered will vary according to factors such as the specific cancer cell involved, and the specific cancer disease; the degree of or involvement or the severity of the cancer disease; the size, age, and general health of the cancer patient; the response of the individual patient; the particular compound administered; the bioavailability characteristics of the preparation administered; the dose regimen selected; whether the conjugate is administered alone or in combination with other agents; pharmacodynamic characteristics of the conjugate and their mode and route of administration; and other relevant characteristics that the physician or as one skilled in the art, will readily determine by the use of known techniques and by observing results obtained under analogous circumstances. The conjugate / composition is suitably administered to the patient at one time or over a series of treatments. Preferably, it is desirable to determine the dose-response curve in vitro, and then in useful animal models prior to testing in humans. The present disclosure provides dosages for the conjugates and compositions comprising same. For example, depending on the type and severity of the disease, about 1 pg / kg to 1000 mg per kg (mg / kg) of body weight per day. Further, the effective dose may be 0.5 mg / kg, 1 mg / kg, 5 mg / kg, 10 mg / kg, 15 mg / kg, 20 mg / kg / 25 mg / kg, 30 mg / kg, 35 mg / kg, 40 mg / kg, 45 mg / kg, 50 mg / kg, 55 mg / kg, 60 mg / kg, 70 mg / kg, 75 mg / kg, 80 mg / kg, 90 mg / kg, 100 mg / kg, 125 mg / kg, 150 mg / kg, 175 mg / kg, 200 mg / kg, and may increase by 25 mg / kg increments up to 1000 mg / kg, or may range between any two of the foregoing values. A typical daily dosage might range from about 1 pg / kg to 100 mg / kg or more, depending on the factors mentioned above. For repeated administrations over several days or longer, depending on the condition, the treatment is sustained until a desired suppression of disease symptoms occurs. However, other dosage regimens may be useful. The progress of this therapy is easily monitored by conventional techniques and assays.

[0300] The conjugate compound or salt thereof or composition comprising same described herein may be used in combination with one or more additional active agents or therapies (radiotherapy, surgery, vaccines, etc.) for the treatment the targeted disease / condition or for the management of one or more symptoms of the targeted disease / condition (e.g., pain killers, anti- nausea agents, etc.). In an embodiment, the conjugate compound described herein is used in combination with one or more chemotherapeutic agents, immunotherapies, checkpoint inhibitors, cell-based therapies, etc. Examples of chemotherapeutic agents suitable for use in combination with the conjugate described herein include, but are not limited to, vinca alkaloids, agents that disrupt microtubule formation (such as colchicines and its derivatives), anti-angiogenic agents, therapeutic antibodies, EGFR targeting agents, tyrosine kinase targeting agent (such as tyrosine kinase inhibitors), transitional metal complexes, proteasome inhibitors, antimetabolites (such as nucleoside analogs), alkylating agents, platinum-based agents, anthracycline antibiotics, topoisomerase inhibitors, macrolides, retinoids (such as all-trans retinoic acids or a derivatives thereof); geldanamycin or a derivative thereof (such as 17-AAG), and other cancer therapeutic agents recognized in the art. In some embodiments, chemotherapeutic agents for use in combination with the conjugate described herein comprise one or more of adriamycin, colchicine, cyclophosphamide, actinomycin, bleomycin, duanorubicin, doxorubicin, epirubicin, mitomycin, methotrexate, mitoxantrone, fluorouracil, carboplatin, carmustine (BCNU), methyl-CCNU, cisplatin, etoposide, interferons, camptothecin and derivatives thereof, phenesterine, taxanes and derivatives thereof (e.g., taxol, paclitaxel and derivatives thereof, taxotere and derivatives thereof, and the like), topetecan, vinblastine, vincristine, tamoxifen, piposulfan, nab-5404, nab-5800, nab- 5801 , Irinotecan, HKP, Ortataxel, gemcitabine, Oxaliplatin, Herceptin®, vinorelbine, Doxil®, capecitabine, Alimta®, Avastin®, Velcade®, Tarceva®, Neulasta®, lapatinib, sorafenib, erlotinib, erbitux, derivatives thereof, and the like. In an embodiment, the conjugate compound or composition comprising same described herein is used in combination with an EGFR or tyrosine kinase targeting agent, for example an EGFR inhibitor (RTK inhibitor). The conjugate compound or salt thereof or composition comprising same described herein may also be used in combination with one or more therapeutic antibodies or antibody fragments, e.g., therapeutic antibodies or antibody fragments used for the treatment of tumors. Examples of antibodies used for the treatment of cancers include antibodies targeting CD52 (e.g., Alemtuzumab), VEGFA / EGFR (e.g., Bevacizumab, Ramucirumab), EGFR (e.g., Cetuximab, Necitumumab, Panitumumab), CD38 (e.g., Daratumumab, Isatuximab), RANKL (e.g., Denosumab), GD2 (e.g., Dinutuximab, Naxitamab-gqgk), SLAMF7 (e.g., Elotuzumab), HER2 (e.g., Margetuximab-cmkb, Pertuzumab), CCR4 (e.g., Mogamulizumab), CD20 (Obinutuzumab, Ofatumumab, Rituximab), BCMA (e.g., Teclistamab), CD19 (e.g., Tafasitamab), CTLA-4 (e.g., Tremelimumab), LAG-3 (e.g., Relatlimab), PD-1 (e.g., Tislelizumab, Penpulimab, Sintilimab, Toripalimab, Retifanlimab, Dostarlimab), PD- L1 (e.g., Durvalumab, Avelumab, Atezolizumab), EpCAM (e.g., Oportuzumab, Edrecolomab), Nectin-4 (e.g., Enfortumab), CD79b (e.g., Polatuzumab).

[0301] In an embodiment, the conjugate compound or salt thereof defined herein is used in combination with an immunotherapy (e.g., immunotherapeutic agent). Thus, in another aspect, the present disclosure provides a method for treating cancer in a subject comprising administering to the subject a therapeutically effective amount of a peptide compound or conjugate defined herein in combination with an immunotherapy (e.g., immune checkpoint inhibitor therapy). The present disclosure also provides the use of a peptide compound or conjugate defined herein in combination with an immunotherapy (e.g., immune checkpoint inhibitor therapy) for the treatment of cancer, or for the manufacture of a medicament for the treatment of cancer. The present disclosure also provides a combination therapy comprising a peptide compound or conjugate defined herein and an immunotherapy (e.g., immune checkpoint inhibitor therapy) for use in the treatment of cancer.

[0302] In an embodiment, the peptide compound or conjugate and immunotherapy exhibit a synergistic effect (e.g., on inhibition of tumor growth, survival, etc.) in the subject.

[0303] The term immunotherapy as used herein refers to an anti-tumor treatment that enhances or boosts the immune response against the tumor cells. Immunotherapies include cell-based immunotherapies, for example administration of immune cells that are able to recognize tumor cells, such as chimeric antigen receptor (CAR) T cells and NK cells, or T cells having a TCR specific for a tumor antigen, or antigen-presenting cells (APCs such as dendritic cells) capable of expressing tumor antigens at their surface. Immunotherapies also include the administration of specific antibodies that recognize antigens expressed by tumor cells and target them for destruction by the immune system, or the administration of cytokines (interferons, interleukins) that stimulates the immune response. Another type of immunotherapy comprises administration of an immune checkpoint inhibitor (ICI). A combination of different types of immunotherapies may be used, for example administration of immune cells (CAR T or NK cells) in combination with an immune checkpoint inhibitor.

[0304] The term “immune checkpoint inhibitor” (ICI) or “immune checkpoint blocker” (ICB) as used herein refers to an agent that block or inhibit the activity of a negative regulator of the immune response. In an embodiment, the ICI blocks or inhibits the activity of T cells (e.g., CTLs and / or CD4 helper T cells) and / or of NK cells. Examples of such negative regulators of the immune response (i.e., immune checkpoint) include Adenosine A2A receptor (A2AR), B7-H3 (CD276), B7-H4 (VTCN1), B and T Lymphocyte Attenuator (BTLA or CD272), Cytotoxic T- Lymphocyte-Associated protein 4 (CTLA-4, CD152), CD47 / SIRPa, Indoleamine 2,3-dioxygenase (IDO), Killer-cell Immunoglobulin-like Receptor (KIR), Lymphocyte Activation Gene-3 (LAG3), nicotinamide adenine dinucleotide phosphate NADPH oxidase isoform 2 (NOX2), poliovirus receptor-related immunoglobulin (PVRIG), carcinoembryonic antigen-related cell adhesion molecule 1 (CEACAM1), Programmed Death 1 (PD-1) receptor, PD-L1 , PD-L2, T-cell Immunoglobulin domain and Mucin domain 3 (TIM-3), V-domain Ig suppressor of T cell activation (VISTA), and Sialic acid-binding immunoglobulin-type lectin 7 (SIGLEC7 or CD328) and SIGLEC9 (CD329). In an embodiment, the immune checkpoint inhibitor is an inhibitor of CTLA-4, PD-1 or PD-L1. Examples of immune checkpoint inhibitors includes anti-PD-1 antibodies / blockers (e.g., Tislelizumab, Penpulimab, Pidilizumab, Sintilimab, Toripalimab, Retifanlimab, Dostarlimab, Nivolumab, Cemiplimab, Pembrolizumab, Spartalizumab, Camrelizumab, JTX-4014, INCMGA00012 (MGA012), AMP-224, AMP-514), anti-PD-L1 antibodies / blockers (e.g., Durvalumab, Avelumab, Atezolizumab, KN035, CK-301 , AUNP12, CA-170, BMS-986189), anti- CTLA-4 antibodies (e.g., Tremelimumab, Ipilimumab), anti-LAG-3 antibodies (e.g., Relatlimab, LAG525 (IMP701), REGN3767 (R3767), Bl 754,091 , tebotelimab (MGD013), eftilagimod alpha (IMP321), FS118), anti-TIM-3 antibodies (MBG453, Sym023, TSR-022), anti-B7-H3 / H4 antibodies (e.g., MGC018, FPA150), adenosine signaling pathway inhibitors including A2AR inhibitors (e.g., Inupadenant (EGS100850), Etrumadenant (AB928), Imaradenant (AZD4635), Ciforadenant, NIR178, CS3005, PBF-999, INCB106385, CPI-444), CD73 antagonists / anti-CD73 antibodies (e.g., Mupadolimab (CPI-006), Oleclumab (MEDI9447), Uliledlimab, AB680, BMS- 986179, NZV930, AK119, SYM024, INCA00186, ORIC-533, IPH5301 , PSB-1248937), and CD39 antagonists (TTX-030, IPH5201 , SRF617), anti-NKG2A antibodies (Monalizumab), anti-PVRIG (e.g., COM701), anti-CEACAM1 antibodies (e.g., CM24), and CD47 blockers / inhibitors (Evorpacept (ALX148), Hu5F9-G4 (5F9), TTI-662, RRx-001) (see, e.g., Marin-Acevedo et al., Next generation of immune checkpoint inhibitors and beyond, Journal of Hematology & Oncology, volume 14, Article number: 45 (2021); Xia et al., CD39 / CD73 / A2AR pathway and cancer immunotherapy, Molecular Cancer, volume 22, Article number: 44 (2023)). The chemical structures and sequences of the above-noted immune checkpoint inhibitors are incorporated herein by reference.

[0305] In an embodiment, the immune checkpoint inhibitor is an inhibitor of PD-1 , such as an anti-PD-1 antibody. In an embodiment, the immune checkpoint inhibitor is an inhibitor of PD-L1 , such as an anti-PD-L1 antibody. In an embodiment, the immune checkpoint inhibitor is an inhibitor of CTLA-4, such as an anti-CTLA-4 antibody. In an embodiment, a combination of ICIs is used, for example a combination of a CTLA-4 inhibitor and a PD-1 inhibitor (e.g., Ipilimumab with nivolumab), or a combination of a CTLA-4 inhibitor and a PD-L1 inhibitor (e.g., Durvalumab with tremelimumab).

[0306] The combination of active agents (e.g., conjugate compound + immunotherapeutic agent) and / or compositions comprising same may be administered or co-administered (e.g., consecutively, simultaneously, at different times) in any conventional dosage form. Coadministration in the context of the present invention refers to the administration of more than one therapeutic in the course of a coordinated treatment to achieve an improved clinical outcome. Such co-administration may also be coextensive, that is, occurring during overlapping periods of time. For example, a first agent (e.g., the conjugate compound described herein) may be administered to a patient before, concomitantly, before and after, or after a second active agent (e.g., a chemotherapeutic agent or an immunotherapy) is administered. The agents may in an embodiment be combined / formulated in a single composition and thus administered at the same time.

[0307] In an embodiment, enhancing the anti-tumor immune response comprises (a) increasing the number of tumor-infiltrating lymphocytes (TILs), tumor-associated macrophages (TAMs) and / or Natural Killer (NK) cells in the tumor; and / or (b) decreasing the levels of immunoregulatory cells in the tumor. In an embodiment, the TILs comprise activated and / or memory CD4+and / or CD8+T cells, such as cytotoxic CD8+T cells. In an embodiment, the TAMs comprise type 1 macrophages (M1). In an embodiment, the NK cells comprise cytotoxic NK cells. In an embodiment, the immunoregulatory cells comprises CD4+regulatory T cells (Tregs), type 2 macrophages (M2), and / or NK regulatory cells (NKreg).

[0308] The cancer may be any type of cancer, including a primary (or original) cancer, a relapsing cancer or a metastatic cancer. Examples of cancers include heart sarcoma, lung cancer, small cell lung cancer (SCLC), non-small cell lung cancer (NSCLC), bronchogenic carcinoma (squamous cell, undifferentiated small cell, undifferentiated large cell, adenocarcinoma), alveolar (bronchiolar) carcinoma, bronchial adenoma, sarcoma (e.g., Ewing’s sarcoma, Karposi's sarcoma), lymphoma, chondromatous hamartoma, mesothelioma; cancer of the gastrointestinal system, for example, esophagus (squamous cell carcinoma, adenocarcinoma, leiomyosarcoma, lymphoma), stomach (carcinoma, lymphoma, leiomyosarcoma), gastric, pancreas (ductal adenocarcinoma, insulinoma, glucagonoma, gastrinoma, carcinoid tumors, vipoma), small bowel (adenocarcinoma, lymphoma, carcinoid tumors, Karposi's sarcoma, leiomyoma, hemangioma, lipoma, neurofibroma, fibroma), large bowel (adenocarcinoma, tubular adenoma, villous adenoma, hamartoma, leiomyoma); cancer of the genitourinary tract, for example, kidney cancer (adenocarcinoma, Wilm's tumor [nephroblastoma], lymphoma, leukemia), bladder and / or urethra cancer (squamous cell carcinoma, transitional cell carcinoma, adenocarcinoma), prostate cancer (adenocarcinoma, sarcoma), testis cancer (seminoma, teratoma, embryonal carcinoma, teratocarcinoma, choriocarcinoma, sarcoma, interstitial cell carcinoma, fibroma, fibroadenoma, adenomatoid tumors, lipoma); liver cancer, for example, hepatoma (hepatocellular carcinoma, HCC), cholangiocarcinoma, hepatoblastoma, angiosarcoma, hepatocellular adenoma, hemangioma, pancreatic endocrine tumors (such as pheochromocytoma, insulinoma, vasoactive intestinal peptide tumor, islet cell tumor and glucagonoma); bone cancer, for example, osteogenic sarcoma (osteosarcoma), fibrosarcoma, malignant fibrous histiocytoma, chondrosarcoma, 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; cancer of the nervous system, for example, neoplasms of the central nervous system (CNS), primary CNS lymphoma, skull cancer (osteoma, hemangioma, granuloma, xanthoma, osteitis deformans), meninges (meningioma, meningiosarcoma, gliomatosis), brain cancer (astrocytoma, medulloblastoma, glioma, ependymoma, germinoma [pinealoma], glioblastoma multiform, oligodendroglioma, schwannoma, retinoblastoma, congenital tumors), spinal cord neurofibroma, meningioma, glioma, sarcoma); cancer of the reproductive system, for example, gynecological cancer, uterine cancer (endometrial carcinoma), cervical cancer (cervical carcinoma, pre-tumor cervical dysplasia), ovarian cancer (ovarian carcinoma [serous cystadenocarcinoma, mucinous cystadenocarcinoma, unclassified carcinoma], granulosa-thecal cell tumors, Sertoli-Leydig cell tumors, dysgerminoma, malignant teratoma), vulvar cancer (squamous cell carcinoma, intraepithelial carcinoma, adenocarcinoma, fibrosarcoma, melanoma), vaginal cancer (clear cell carcinoma, squamous cell carcinoma, botryoid sarcoma (embryonal rhabdomyosarcoma), fallopian tube cancer (carcinoma); placenta cancer, penile cancer, prostate cancer, testicular cancer; cancer of the hematologic system, for example, blood cancer (acute myeloid leukemia (AML), chronic myeloid leukemia (CML), acute lymphoblastic leukemia (ALL), chronic lymphocytic leukemia (CLL), myeloproliferative diseases, multiple myeloma, myelodysplastic syndrome), Hodgkin's disease, non-Hodgkin's lymphoma [malignant lymphoma]; cancer of the oral cavity, for example, lip cancer, tongue cancer, gum cancer, palate cancer, oropharynx cancer, nasopharynx cancer, sinus cancer; skin cancer, for example, malignant melanoma, cutaneous melanoma, basal cell carcinoma, squamous cell carcinoma, Karposi's sarcoma, moles dysplastic nevi, lipoma, angioma, dermatofibroma, and keloids; adrenal gland cancer: neuroblastoma; and cancers of other tissues including connective and soft tissue, retroperitoneum and peritoneum, eye cancer, intraocular melanoma, and adnexa, breast cancer (e.g., ductal breast cancer), head or / and neck cancer (head and neck squamous cell carcinoma), anal cancer, thyroid cancer, parathyroid cancer; secondary and unspecified malignant neoplasm of lymph nodes, secondary malignant neoplasm of respiratory and digestive systems and secondary malignant neoplasm of other sites.

[0309] In an embodiment, the cancer is an immunologically cold (or ignorant) cancer. The term “immunologically cold cancer” or “cold cancer” refers to a cancerthat does not trigger an antitumor immune response in the patient and / or that does not respond to cancer immunotherapies such as ICI therapy (see, e.g., Bonaventura et al., “Cold Tumors: A Therapeutic Challenge for Immunotherapy.” Frontiers in immunology vol. 10 168 (2019). Immunologically cold cancers are characterized by the absence of infiltration of antitumor immune cells such as TILs, TAMs and / or NK cells in the tumor and / or the presence of high levels of immunoregulatory cells (e.g., Tregs). Immunologically cold tumors can be subdivided into so-called immune deserts, in which immune effector cells such as T cells are absent from the tumor and its periphery, and immune-excluded tumors, in which immune effector cells such as T cells accumulate near the tumor but do not efficiently infiltrate it. In an embodiment, the immunologically cold cancer is an immune desert. In another embodiment, the immunologically cold cancer is an immune-excluded cancer. A number of breast cancers, ovarian cancers, prostate cancers, pancreatic cancers, and glioblastomas are considered immunologically cold cancers. Subtypes of several cancers are also considered immunologically cold cancers including subtypes of lung cancers such as non-small- cell lung cancers (NSCLC) (Cascone et al., Tumor Immunology and Immunotherapy of Non- Small-Cell Lung Cancer, Cold Spring Harb Perspect Med. 2022 May 27;12(5):a037895), renal cancers such as chromophobe renal cell carcinoma (non-clear cell renal cell carcinoma, nccRCC) (Zarrabi et al., Immune Checkpoint Inhibition in Advanced Non-Clear Cell Renal Cell Carcinoma: Leveraging Success from Clear Cell Histology into New Opportunities, Cancers vol. 13,15 3652. 21 Jul. 2021), high tumor mutational burden RCC (Yakirevich etal., Tumor mutational burden and immune signatures interplay in renal cell carcinoma. Ann Transl Med. 2020;8(6):269), colorectal cancers (CRC) such as consensus molecular subtype (CMS) 2 and CMS3 CRC (Roelands et al., Immunogenomic Classification of Colorectal Cancer and Therapeutic Implications, Int J Mol Sci. 2017;18(10):2229), head and neck squamous cell carcinoma (Ribbat-ldel et al., Immunologic “Cold” Squamous Cell Carcinomas of the Head and Neck Are Associated With an Unfavorable Prognosis, Frontiers in medicine, 8, 622330), oesophageal cancer (Puhr et al., Immunotherapy for Esophageal Cancers: What Is Practice Changing in 2021?, Cancers vol. 13,18 4632), liver cancer such as stage II hepatocellular carcinoma (Nguyen etal., Nature Communications volume 13, Article number: 1441 (2022)). Homozygous deletion of 9p21.3, one of the most frequent genomic defects occurring in ~13% of all cancers including melanoma (SKCM), bladder (BLCA), pancreatic cancer (pancreatic adenocarcinoma), gastric cancer (stomach adenocarcinoma), lung adeno- (LUAD) and squamous-cell carcinoma (LUSC)], has been shown to be associated with an immunologically cold phenotype.

[0310] In an embodiment, the methods and uses described herein further comprises a step of determining whether the patient suffers (or identifying a patent suffering) from an immunologically cold cancer. The determination of whether a patient suffers from an immunologically cold cancer may be made by various methods known in the art. For example, this determination may be made by assessing the presence of immune cells such as TILs (cytotoxic CD8+T cells), TAMs (M1) and / or NK cells (cytotoxic NK cells) in the tumor microenvironment (TME), wherein the absence or a low number of immune cells in the TME is indicative that the patient suffers from an immunologically cold cancer, and / or by assessing the presence of immunoregulatory cells (e.g., Tregs) in the TME, wherein the presence of immunoregulatory cells (e.g., high number of immunoregulatory cells) (e.g., Tregs) in the TME is indicative that the patient suffers from an immunologically cold cancer. “Low number of immune cells” means a number of immune cells that is significantly lower relative to the mean or average number of immune cells detected in corresponding tumors of the same type, and “high number of immunoregulatory cells” means a number of immunoregulatory cells (e.g., Tregs) that is significantly higher relative to the mean or average number of immunoregulatory cells (e.g., Tregs) detected in corresponding tumors of the same type.

[0311] In an embodiment, the cancer is resistant to an immunotherapy, i.e., a cancer in which the immunotherapy does not lead to inhibition of tumor growth in the patient. A cancer resistant to immunotherapy may be a cancer that has never responded to immunotherapy (primary resistance) or that has developed resistance to immunotherapy treatment after a period of (responsive) treatment (acquired resistance).

[0312] In a further embodiment, the cancer is resistant to PD-1 or PD-L1 inhibitor-based therapy (anti-PD-1 / PD-L1 therapy). In a further embodiment, the cancer is resistant to PD-1 or PD-L1 inhibitor-based therapy is melanoma, lung cancer, renal cell carcinoma, Hodgkin lymphoma, head and neck cancer, colon cancer, liver cancer, stomach cancer, squamous cell skin cancer or myeloma.

[0313] Immune checkpoint inhibitors have been approved or are currently being tested in phase III and IV clinical trials for several cancers including lung cancer (e.g., non-small cell lung cancer (NSCLC) and small cell lung cancer, squamous cell lung carcinoma), head and neck cancer (e.g., head and neck squamous cell carcinoma, renal cell carcinoma, gastric adenocarcinoma, nasopharyngeal neoplasms, urothelial carcinoma, colorectal cancer, mesothelioma (e.g., pleural mesothelioma), breast cancer (e.g., triple-negative breast cancer, TNBC), esophageal neoplasms, multiple myeloma, gastric and gastroesophageal junction cancer, gastric adenocarcinoma, melanoma, Merkel-cell carcinoma (MCC), lymphoma (e.g., Hodgkin and nonHodgkin lymphoma, diffuse Large B-cell lymphoma), liver cancer (e.g., hepatocellular carcinoma), melanoma, ovarian cancer, fallopian tube cancer, peritoneal neoplasms, bladder cancer, transitional cell carcinoma, prostatic neoplasms and biliary tract neoplasms (see, e.g., Darvin et al., Experimental & Molecular Medicine volume 50, Article number: 165 (2018)). Thus, in an embodiment, the cancer is one of the above-noted cancer for which immune checkpoint inhibitors have been approved or are currently being tested in phase III and IV clinical trials.

[0314] Currently approved immune checkpoint inhibitors include the anti-CTLA-4 Ipilimumab (melanoma and lung cancer), the anti-PD-1 Nivolumab (melanoma, lung cancer, renal cell carcinoma, Hodgkin lymphoma, head and neck cancer, colon cancer, and liver cancer), Pembrolizumab (melanoma, lung cancer, head and neck cancer, Hodgkin lymphoma, renal cell carcinoma and stomach cancer), and Cemiplimab (squamous cell skin cancer, myeloma, and lung cancer), and the anti-PD-L1 atezolizumab (NSCLC, small cell lung cancer, TNBC), Avelumab (NSCLC, MCC) and Durvalumab (urothelial carcinoma, lung cancer). Thus, in an embodiment, the cancer is one of the above-noted cancer for which immune checkpoint inhibitors have been approved.

[0315] In an embodiment, the doses of the conjugate compound described herein and / or of the immunotherapy that are used / administered in the methods, uses, compositions, combination therapy of the disclosure is a suboptimal dose. “Suboptimal dose” as used herein refers to a dose of one of the compound(s) of the combination described herein (the peptide compound described herein and / or of the immunotherapy), which, when used in the absence of the other compound of the combination, results in a biological effect of 50% or less, in an embodiment of 40% or less, in a further embodiment of 30% or less, in a further embodiment of 20% or less, in a further embodiment of 10% or less. As such, use of a combination of the compounds described herein, where one or more compounds in the combination is used at a suboptimal dose, may achieve increased efficacy / biological effect relative to using the compound(s) in the absence of the other(s), at a comparable suboptimal dose.

[0316] As used herein, a synergistic effect is achieved when the effect of the combined compounds is greater than the theoretical sum of the effect of each agent in the absence of the other. One potential advantage of combination therapy with a synergistic effect is that lower dosages (e.g., a suboptimal dose) of one or both of the drugs or therapies may be used in order to achieve high therapeutic activity with low toxicity. In an embodiment, the combination therapy (the peptide compound described herein and / or of the immunotherapy) results in at least a 5% increase in the effect relative to the predicted theoretical additive effect of the agents. In a further embodiment, the combination therapy results in at least a 10% increase in the effect relative to the predicted theoretical additive effect of the agents. In a further embodiment, the combination therapy results in at least a 20% increase in the effect relative to the predicted theoretical additive effect of the agents. In a further embodiment, the combination therapy results in at least a 30% increase in the effect relative to the predicted theoretical additive effect of the agents. In a further embodiment, the combination therapy results in at least a 50% increase in the effect relative to the predicted theoretical additive effect of the agents. A further advantage of using the drugs in combination is that efficacy may be achieved in situations where either drug alone would not have an effect, for example for a cancer or tumor resistant to the ICI. Resistance means that the administration of the ICI alone does not lead to a significant therapeutic effect, e.g., a significant reduction in tumor volume or tumor cell number, or an increase in survival time. Examples of cancers for which resistance to ICI has been reported in patients and / or animal models include lung cancer (e.g., NSCLC), pancreatic cancer, prostate cancer, melanoma, ovarian cancer, urothelial cancer, renal cell carcinoma (see, e.g., Fares et al., American Society of Clinical Oncology Educational Book 39, 147-164, 2019; Pandey et al., Cancer Drug Resist 2019; 2:178- 188). In an embodiment, the synergistic effect is on inhibition or reduction of tumor growth. In an embodiment, the synergistic effect is on the increase of survival time.

[0317] Thus, in other aspects, the present disclosure also provides a combination therapy comprising the peptide compound described herein and an immunotherapy, such as an immune checkpoint inhibitor (ICI) therapy. The present disclosure also provides the use of a combination therapy comprising the peptide compound described herein and an immunotherapy (i.e., immunotherapeutic agent), such as an ICI, for treating a subject suffering from a cancer (e.g., a cancer resistant to immunotherapy such as ICI monotherapy). The present disclosure also provides the use of a combination therapy comprising the peptide compound described herein and an immunotherapy, such as an immune checkpoint inhibitor, for the manufacture of a medicament for treating a subject suffering from a cancer (e.g., a cancer resistant to immunotherapy such as ICI monotherapy). The present disclosure also provides a method for treating a subject suffering from a cancer (e.g., a cancer resistant to immunotherapy such as ICI monotherapy) comprising administering to the subject an effective amount of a combination therapy comprising the peptide compound described herein and an immunotherapy such as an ICI.

[0318] As used herein, the term "subject" or “patient” denotes a mammal, such as a rodent, a feline, a canine, and a primate. Preferably a subject or patient according to the disclosure is a human.

[0319] EXAMPLES

[0320] The present invention is illustrated in further details by the following non-limiting examples.

[0321] Example 1 : Materials and Methods

[0322] Cells and reagents.

[0323] Human SK-MEL-28 melanoma cells with stage I and II melanosomes were obtained from American Type Culture Collection (ATCC, Manassas, VA; #HTB-72) [17, 18] and cultured in Eagle’s Minimum Essential Medium (EMEM; Wisent, #217-010-XK) containing 1 mM Na pyruvate and 10% fetal bovine serum (FBS; Hyclone, #SH30396.03) at 37°C in a humidified atmosphere (5% CO2). Human A375 melanoma cells were also obtained from ATCC (#CRL-1619) and cultured in Dulbecco’s Modified Eagle Medium (DMEM; #319-005-CL) containing 10% FBS. Human TNBC-derived MDA-MB-231 / Luc cells were obtained from Cell Biolabs, Inc. (San Diego, CA; #AKR-231) and cultured in DMEM containing 10% FBS. Murine B16-F10 melanoma cells (ATCC; #CRL-6475), a stage lll / IV melanosome model [19, 20], were cultured in DMEM with 10% FBS at 37°C in a humidified atmosphere (5% CO2) and used in the in vivo syngeneic studies. Cell counts and cell viability were assessed with a BioRad TC20 automated cell counter. The polyclonal anti-SORT1 antibody directed against amino acids 800 to the C-terminus of the intracellular domain of SORT 1 was obtained from Abeam (Cambridge, MA; ab16640). The APC anti-mouse CD274 (B7-H1 , PD-L1) antibody (#124312) and its APC rat immunoglobulin G (IgG) 2b, K isotype control antibody (#400612) were obtained from BioLegend (San Diego, CA). The APC major histocompatibility complex class I (MHC-1 ; H-2Db) monoclonal antibody (mAb; 28-14- 8; #17-5999-82) and its APC mouse lgG2a K isotype control (eBM2a) (#17-4724-81) were obtained from Thermo Fisher Scientific (Agawam, MA). All other reagents were from Sigma- Aldrich (Oakville, ON).

[0324] Tissue microarray probing and analysis.

[0325] SORT1 expression was evaluated using high-density tissue microarrays (TMAs) of human melanoma cancers as well as of healthy tissues, at the Institute for Research in Immunology and Cancer (IRIC; Montreal, QC). The TMAs were provided with subtypes information allowing the assessment of SORT1 expression in different grades of melanoma samples. Immunostaining was performed on 4 pm sections of formalin-fixed, paraffin-embedded material. Briefly, SORT1 antigen retrieval was performed by heat-induced epitope retrieval techniques with ER1 solution (Leica) for 30 minutes at 100°C. Mouse anti-SORT1 (clone F11) primary antibody was incubated at 20 pg / mL for 30 minutes at room temperature. Target antigen was detected using the Bond™ Polymer Refine Detection (Leica Biosystems, Buffalo Grove, IL) with diaminobenzidine chromogen for visualization according to the manufacturer's instructions. Sections were then counterstained with Leica proprietary hematoxylin and mounted for analysis. Analysis of the images from the immunohistochemistry was performed by an expert pathologist. SORT1 labeling was scored by the IHS method on a scale ranging from 0 to 3 as follows: 0, negative staining; 1 , weak staining; 2, moderate staining; 3, strong staining. The raw data were converted to the IHS by multiplying the quantity and staining intensity scores. Therefore, the IHS score ranged from 0 to 12.

[0326] Western blotting

[0327] Cells were homogenized in 1% sodium dodecyl sulfate (SDS) lysis buffer supplemented with a complete protease inhibitor cocktail from Calbiochem (San Diego, CA). Cells were incubated for 30 minutes at room temperature (RT) with vortexing every five minutes, sonicated and centrifuged at 15,000g for 10 minutes at RT. Equal amounts of protein (20 pg) were separated by SDS-polyacrylamide gel electrophoresis (PAGE). Proteins were then electrotransferred to a polyvinylidene fluoride (PVDF) membrane and blocked for one hour at RT using 5% non-fat dry milk in Tris-buffered saline (150 mM NaCI, 20 mM Tris-HCI, pH 7.5) containing 0.1% Tween™- 20 (TBST). Membranes were washed in TBST and incubated overnight with primary antibodies against SORT1 (1 / 1 ,000 dilution) or glyceraldehyde 3-phosphate dehydrogenase (GAPDH; #MAb6C5, 1 / 40,000 dilution) diluted in TBST containing 3% bovine serum albumin (BSA) and 0.05% NaN3. STING (#50494, 1 / 1 ,000 dilution), p21 (#ab109199, 1 / 1 ,000 dilution), TANK-binding kinase 1 (TBK1 ; #3504, 1 / 1 ,000 dilution), phosphoTBKI (#5483, 1 / 1 ,000 dilution), P65 (#8242, 1 / 1 ,000 dilution), phosphoP65 (#3033, 1 / 1 ,000 dilution), and p53 (#ab131442, 1 / 5,000 dilution). Membranes were washed in TBST and incubated for one hour at RT with horseradish peroxidase- conjugated anti-mouse or anti-rabbit IgG (1 / 5,000 dilution), in TBST containing 5% non-fat dry milk. Membranes were washed again in TBST, and signals were detected using chemiluminescence (Bio-Rad, Saint-Laurent, QC).

[0328] Animals

[0329] Female CD-1 nude mice (Crl:NU-Foxn7™, 4-6 weeks old) were used for xenograft tumor models and female immunocompetent C57BL / 6 mice (C57BL / 6NCrl, 4-6 weeks old) were used for syngeneic tumor models. All mice were obtained from Charles River Laboratories, Inc. (St- Constant, QC). Animals were allowed to acclimate for 5 days before experiments. All mice were maintained in a pathogen-free environment and handled in accordance with the Guidelines of the Canadian Council on Animal Care (CCAC) for care and use of experimental animals.

[0330] Preparation of test articles for injection

[0331] TH 1902 (Acetyl-GVRAK(docetaxel)AGVRN(Nle)FK(docetaxel)SESY) was synthesized as previously described (PCT publication No. WO 2017 / 088058). Stock solution (10 mg / ml) of formulated TH1902 was prepared as sterile aliquots and frozen. On the day of animal dosing, frozen aliquots were thawed then diluted with sterile 5% Dextrose Injection USP (D5W) to the desired concentration for injection. Vehicle frozen stock solution was processed exactly as per the dilutions of TH 1902 stock solution to match the quantities of excipients present in the TH 1902 animal groups. Docetaxel (Wonda Science Inc, Lexington, MA) was prepared the same day as animal dosing and to match the docetaxel content of the highest administered TH1902 dose for comparison. Docetaxel was solubilized to 50 mg / ml with injectable EtOH, diluted to 25 mg / ml with polysorbate-80, then diluted with sterile 5% D5W to the desired concentration for injection (i.e., 2.5 mg / ml). All diluted solutions were filtered before animal administrations (Millex-GP 0.22 pm syringe filter, PES membrane, Millipore). Note that, in terms of docetaxel content, the 15 mg / kg docetaxel dose is equivalent to 35 mg / kg TH 1902. Similarly, the 7.5 mg / kg docetaxel dose is equivalent to 17.5 mg / kg TH1902 (the conversion factor of 2.3 considers molecular weights of the two docetaxel moieties within each TH1902 molecule). Therefore, docetaxel makes 44% of each TH1902 molecule. The anti-PD-L1 mAb (clone 10F.9G2; BP0101) and isotype control (clone LTF- 2; BP0090) were purchased from Bio X Cell (Lebanon, NH) and diluted according to the manufacturer’s instructions (IP0065 and IP0070, respectively) to the desired concentration for injection. All diluted solutions were filtered before animal administrations (Millex-GP 0.22 pm syringe filter, polyethersulfone (PES) membrane, MilliporeSigma, Burlington, MA).

[0332] In vivo therapeutic efficacy assessment of docetaxel and TH 1902 in MDA-MB-231 immunocompromised xenograft model, and in a B16-F10 melanoma syngeneic immunocompetent model. The MDA-MB-231 xenograft model was described previously [4, 5], Generation of the B16-F10 melanoma syngeneic model was performed as follows: B16-F10 cells were resuspended in 100 pl of implantation medium (MilliporeSigma; HBSS, #H6648) and Matrigel (Corning Inc., Corning, NY; #356231), in a 1 :1 proportion to inject 1x105cells in 100 pl (1x106cells / ml). Tumors were established by subcutaneous inoculation of B16-F10 cells in the dorsal region of immunocompetent C57BL / 6 mice under light isoflurane anesthesia. The injection schedules, doses, number of treatment cycles, and administration method of injected substances are detailed in the figures. Briefly, mice were treated weekly with either vehicle, docetaxel (15 mg / kg), or TH1902 (35 mg / kg) via intravenous (IV) tail vein injection. Treatments were initiated when MDA-MB-231 tumors reached an average size of about 100 mm3or three days following B16-F10 cell implantation. MDA-MB-231 -tumors were collected ( / ) four days following three cycles of treatment of either vehicle, docetaxel, or TH1902; ( / / ) four days following six cycles of treatment with TH 1902; or (Hi) four days following six cycles of treatment with TH 1902, then three cycles off-treatment, for prolonged tumor growth observation. B16-F10-tumors were collected four days following two cycles of treatment of either vehicle, docetaxel, or TH 1902. Tumors were collected at their respective timepoints then fixed in 10% buffered formalin and processed for immunohistochemistry (IHC) analysis. For all in vivo studies, tumor growth was monitored by two- dimensional measurements taken with an electronic caliper, and tumor volume was calculated according to the following formula: tumor volume (mm3) = TT / 6 X length x width2. Animal weights were measured with a precision of ±10 mg.

[0333] In vivo assessment of combined therapeutic efficacies of docetaxel or TH 1902 with anti-PD-L1 antibody in the B16-F10 syngeneic model.

[0334] Generation of the B16-F10 melanoma syngeneic model was performed as described above. The first study investigated the effect on tumor growth of docetaxel, TH 1902, or anti-PD- L1 alone or in combination (docetaxel / anti-PD-L1 or TH1902 / anti-PD-L1) in primary tumors three days post-implantation of B16-F10 cells into mice, which were treated for two cycles. The treatment doses and schedules for each cycle (indicated in the figure legend) were as follows: (i) control group treated with appropriate vehicles and isotype control for anti-PD-L1 (9 mg / kg, i.p., biweekly); (ii) docetaxel at half its maximum tolerated dose (MTD; 7.5 mg / kg, IV, weekly); (Hi) TH1902 at an equivalent dose of docetaxel (17.5 mg / kg, IV, weekly); (iv) combination of docetaxel with anti-PD-L1 ; (v) combination of TH1902 with anti-PD-L1. The second study investigated the effect on mice survival of escalating doses of TH1902 (4.37, 8.75, and 17.5 mg / kg, IV, weekly) or anti-PD-L1 (9 mg / kg, intraperitoneally [IP], biweekly) alone or in combination. Cycles of treatment were continued until one of the study endpoints was reached (tumor size >2,000 mm3, weight loss of >20% of initial weight, ulcerated tumor, death). In both studies, tumor growth and mice weights were monitored as described above. In vivo therapeutic efficacy assessment of docetaxel and TH1902 using the syngeneic Lewis lung carcinoma (LL / 2) xenograft model.

[0335] Tumors were established by subcutaneous inoculation of 1x106cells in 100 l (LL / 2) HBSS / Matrigel (50:50). All cells were injected into the dorsal area of immunocompetent (C57BL / 6) mice under light isoflurane anesthesia. For LL / 2 tumors, mice were treated weekly, 3 days post-implantation, with either vehicle, docetaxel (15 mg / kg) or TH1902 at equivalent docetaxel amounts (35 mg / kg) via intravenous (IV) tail vein injection through 11 days, encompassing 2 treatments (on Days 0 and 7 post start of treatment). For all indicated studies, tumor growth was monitored by two-dimensional measurements taken with an electronic caliper and tumor volume was calculated according to the following formula: tumor volume (mm3) = TT / 6 x length x width2. Animal weights were measured with a precision of ±10 mg. Tumors were collected when mice in the vehicle group reached approximately 600-1000 mm3in size.

[0336] Immunohistochemistry staining of tissue microarray and in vivo tumors.

[0337] SORT1 expression was evaluated using high-density tissue microarrays (TMAs) of human melanomas (IMH-366; Novus Biologicals, Centennial, CO) as well as of healthy tissues (IMH-373; Novus Biologicals) at the Institute for Research in Immunology and Cancer (IRIC; Montreal, QC). The TMAs were supplied along with subtype information, allowing assessment of SORT1 expression in different grades of melanoma samples. Immunostaining of TMAs was performed as previously described [5], An expert pathologist analyzed the IHC images. SORT1 labeling was scored by the IHS method on the following 0-3 scale: 0, negative staining; 1 , weak staining; 2, moderate staining; 3, strong staining. The raw data were converted to IHS scores by multiplying the quantity and staining intensity scores. Consequently, the IHS scores ranged from O to 12.

[0338] Primary MDA-MB-231 and B16-F10 tumors were isolated from mice upon sacrifice, fixed in 10% buffered formalin, then embedded in paraffin. Hematoxylin and Eosin (H&E; 23-314631 and 23-245657, Fisher Scientific) staining was done to evaluate general morphology. IHC was performed on consecutive slides using the Leica Bond Max automated platform (Leica Biosystems, Nussloch GmbH) with the bond polymer refine detection kit (DS9800, Leica Biosystems). Briefly, 4-micron sections from tissue blocks were taken on coated slides. These were then deparaffinized, hydrated, and blocked with hydrogen peroxide. Heat-induced or enzymatic antigen retrieval was achieved using appropriate buffer for each antibody. Slides were incubated with primary antibody against SORT1 , Ki-67, CD31 , STING, CD45, CD3, CD8, CD4, FoxP3, CD161c, F4 / 80, CD68, CD206, Perforin, Granzyme B, or cleaved Caspase-3 followed by secondary antibody. Staining was completed with diaminobenzidine (DAB) chromogen and hematoxylin was used as a counterstain. The quantification of immune cells infiltrating the tumor cores was achieved using the National Institutes of Health (NIH) Imaged Version 1 .4.21 software. Mean color intensity using identical threshold settings per staining were divided by tumor region of interest surface area and expressed as positive area in percentage. For each section, analysis was performed on the entire tumor surface but with the outer edge, invasive margins, and necrotic regions excluded. Details on all primary antibodies, commercial supplier, dilution, antigen retrieval, and treatment conditions are provided in Table 1.

[0339] Table 1 : List of antibodies and staining conditions used in the immunohistochemistry experiments described herein

[0340] 1Primary antibody dilution with or without blocking solution (Bk; Leica Biosystems, no. PV6122).2H1 : HIER 1 , Citrate buffer pH 6, Leica Biosystems, no. AR9961 ; HIER 2, EDTA buffer pH 9,

[0341] Leica Biosystems, no. AR9640; ENZ: enzymatic digestion, Leica Biosystems, no. AR9551.

[0342] 3Incubation primary antibody / secondary antibody / polymer.

[0343] The detection of healthy blood vessels and VM-associated structures was performed as previously described [4], Briefly, formalin-fixed, paraffin-embedded tumor tissue sections were stained with primary antibodies against mouse CD31. Then, the sections were treated with 1% periodic acid-Schiff (PAS) solution for 10 minutes. After rinsing with distilled water for two minutes, tumor tissue sections were placed into Schiff solution for 30 minutes in a dark chamber and rinsed with distilled water three times. Finally, the slides were counterstained with Leica proprietary hematoxylin and mounted for analysis. Healthy vessels are CD31-positive / PAS-positive whereas VM-associated structures are CD31-negative / PAS-positive. Images were captured using Nanozoomer slide scanner (Hamamatsu Photonics K.K., Hamamatsu, Japan) and analyzed with Aperio ImageScope (Leica Biosystems, version 12.4.3.5008, Buffalo Grove, IL).

[0344] Cell proliferation assay.

[0345] To assess the effects of docetaxel and TH1902 on SK-MEL-28 and B16-F10 melanoma cell proliferation, cells were seeded in 96-well plates (PerkinElmer, Waltham, MA) and treated with various concentrations of drugs in complete cell culture medium. After 120 hours of incubation for SK-ML-28 and 72 hours for B16-F10, cell proliferation was measured using the 3- (4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay in accordance with the protocol described by Mosmann

[0021] with the following modifications. The cells were incubated with MTT (0.5 mg / ml) at 37°C under a humidified atmosphere containing 5% CO2for four hours. After incubation, 100 pL of DMSO (solubilizing reagent) was added to each well and mixed thoroughly for five minutes to dissolve the dark blue crystals. The presence of viable cells was visualized by development of a purple color due to formation of formazan crystals. The plates were read on a SpectraMax™ Plus reader (Molecular Devices, San Jose, CA) using a test wavelength of 570 nm and reference wavelength of 650 nm. Analyseswere made in quadruplicate for each condition.

[0346] Cell apoptosis assay.

[0347] AnnexinV / PI staining was performed using an Apoptosis Detection Kit according to the manufacturer's instructions (BD Pharmingen, San Diego, CA). Briefly, B16-F10 cells were treated for 15 minutes in serum-free media containing either docetaxel (0.1 or 2 pM) or TH1902 (0.05 or 1 pM). Cells were then washed twice with complete growth medium and incubated for 94 hours in complete growth medium. Cells were harvested, then resuspended in a staining solution of 100 pL of 1X binding buffer containing 5 pL of AnnexinV-FITC and 5 pL of propidium iodide (PI). Cells were finally incubated for 15 minutes at room temperature in the dark; the numbers of apoptotic cells were acquired and analyzed using a BD Accuri C6 flow cytometer (BD Biosciences, Franklin Lakes, NJ).

[0348] Senescence activity assay.

[0349] B16-F10 cells were plated in 8-well chamber slides from ibidi (Grafelfing, Germany) at a density of 20,000 cells / well. After 24 hours, cells were treated for two hours in serum-free media containing 7.5 pM docetaxel or 3.75 pM TH1902. Cells were then washed twice with complete growth medium and incubated for four days in complete growth medium. As a positive control for senescence, cells were treated for 24 hours in serum-free medium with 12.5 pM etoposide, washed, and incubated in complete growth medium for up to four days. CellEvent™ Senescence Green Detection Kit (Thermo Fisher, #010850) was used to detect senescence-associated |3- Galactosidase (SA-p-gal) activity according to manufacturer instructions. Photomicrographs were taken and digitalized at 20x magnification by confocal microscopy (Nikon Al plus, Melville, NY) then analyzed using NIH Imaged Version 1.4.21 software. For cell morphologic assessment, cells were fixed with 10% formalin phosphate, colored with 0.1% crystal violet / 20% methanol, and observed by microscopy.

[0350] Total RNA isolation, cDNA synthesis, and real-time quantitative PCR.

[0351] Total RNA was extracted from cell monolayers using the Qiagen RNeasy™ kit (QIAGEN, Toronto, ON). For copy DNA (cDNA) synthesis, 2 pg of total RNA was reverse-transcribed using a high-capacity cDNA reverse transcription kit (4368814, Applied Biosystems, Foster City, CA). The cDNA was stored at -20°C prior to polymerase chain reaction (PCR). Gene expression was quantified by real-time quantitative PCR using SsoFast EvaGreen Supermix™ (1725201 , BioRad, Hercules, CA). DNA amplification was carried out using an Icycler iQ5 (Bio-Rad) and product detection was performed by measuring the binding of the fluorescent dye SYBR Green I to double-stranded DNA. The following primer sets were from QIAGEN: lnterleukin-6 (IL-6) (QT00098875, Mm_H6_1_SG), Tumor necrosis factor alpha (TNFoc) (QT00104006, Mm_Tnf_1_SG), GAPDH (QT01658692, Mm_Gapdh_3_SG) and Peptidylprolyl Isomerase A (PPIA) (QT00247709, Mm_Ppia_1_SG). The relative quantities of target gene mRNA were normalized against internal housekeeping genes PPIA and GAPDH. The RNA was measured by following a ACTmethod employing an amplification plot (fluorescence signal vs. cycle number). The difference (ACT) between the mean values in the triplicate samples of the target gene and the housekeeping genes was calculated with the CFX Manager Software version 2.1 (Bio-Rad) and the relative quantified value (RQV) was expressed as 2-ACT.

[0352] Flow cytometry.

[0353] Single B16-F10 live cell suspensions were washed with RPMI-1640, supplemented with 8% FBS in PBS and once with FACS buffer. Subsequently, samples were incubated with Fc block mouse (2%) and mouse serum (5%) for 10 minutes, then stained with anti-MHC-l or anti-PD-L1 antibodies for one hour at 4 °C in the dark and finally rinsed twice with PBS containing 0.5% BSA solution. Samples were acquired using the LSR Fortessa (BD Biosciences) and results analyzed with FlowJo software.

[0354] Statistical analysis Data are expressed as means ± standard error of the mean (SEM) or standard deviation (SD) as indicated in the figure legends. Statistical analysis was done using t-test for comparing two samples, whereas analysis by one-way ANOVA followed by Bonferroni’s, Dunnett’s or Tukey’s multiple comparisons was employed for three or more samples. A value of p < 0.05 (*) and p < 0.01 (**) was considered significant and an asterisk (*) identifies the level of such significance in the figures.

[0355] Example 2: Sustained and prolonged antitumor activity of TH1902 in an immunosuppressed MDA-MB-231 TNBC-derived xenograft model correlates with immune cell infiltration and increased STING expression.

[0356] The efficacy of TH 1902 and docetaxel against a TNBC xenograft model was first investigated in vivo. Immunocompromised nude mice were implanted in the right flank with MDA- MB-231 cancer cells, and tumor growth monitored as described in Example 1 . Mice were treated with IV bolus injections of either (i) docetaxel at the MTD of 15 mg / kg / wk (three treatments), or (ii) TH1902 at 35 mg / kg / wk (six treatments). An additional group received TH1902 for (Hi) six treatments and was monitored up to day 60 (FIG. 1A). Treated tumors were collected after cessation of treatment and, in the case of TH 1902, at day 60. In contrast to docetaxel, which at equivalent TH1902 doses only halved tumor growth, TH1902 induced complete tumor regression and maintained this effect for six treatments, up to day 60 (i.e., 25 days after the last treatment). While body weights remained virtually unchanged (FIG. 1E). H&E staining of resected tumors exhibited significantly enlarged SORT1 -positive cancer cell morphology upon six cycles of treatment; this morphology was maintained off-treatment (FIG. 1B). IHC analysis further revealed reduced Ki67 cell proliferation, accompanied by increased STING and CD45 staining, in TH1902- treated tumors and a sustained expression of SORT1 on cancer cells following treatments (FIG. 1C). Increased CD45 immune infiltration staining specifically surrounded the enlarged cancer cells post-treatment (FIG. 1C). TH1902’s anti-vasculogenic mimicry (VM) properties were observed as early as the third cycle of treatment, whereupon CD317PAS+staining was reduced in TH1902-treated tumors (FIG. 1D). In contrast, docetaxel appeared to have a much lesser effect than TH1902 on Ki67, STING, and CD45 staining as well as on VM. Interestingly, unlike docetaxel, TH1902 significantly induced cell senescence-associated expression of both p21 and p53 in vitro in MDA-MB-231 cells (FIGs. 1F-G). Finally, the results depicted in FIGs. 1H-I show that treatment with TH1902 increase PD-L1 expression in MDA-MB-231 tumors in vivo. Collectively, these results provide evidence that TH1902 has the ability to alter the immune tumor microenvironment (iTME) through immune cell infiltration processes, and correlate with the long- lasting effect on tumor regression. Example 3: SORT1 is highly expressed in clinically annotated melanoma tissues and cell line models.

[0357] To assess whether TH 1902 could serve a SORT1 receptor-mediated chemotherapeutic approach in an immunocompetent syngeneic model, SORT 1 expression was assessed in murine B16-F10 melanoma cells along with other cell lines. This screen was also conducted in TMAs of healthy skin tissues and of clinically annotated melanomas from stages I l-IV by IHC (FIG. 2A). IHS scoring showed that SORT1 expression was increased throughout those stages, whereas levels remained low in healthy tissues (FIG. 2B). High SORT1 expression was also validated in lysates from a murine B16-F10 melanoma cell line and from two human melanoma cell lines (A375 and SK-M EL-28), as well as from a positive control (human TNBC-derived MDA-MB-231) (FIG. 2C). High expression of SORT1 in murine B16-F10 cells validated that such a syngeneic model may be used to assess the in vivo therapeutic effects of TH 1902 on the iTME.

[0358] Example 4: TH1902 exerts in vitro anti-proliferative and pro-apoptotic activities and induces cancer cell senescence.

[0359] SORT1 -positive SK-MEL-28 (FIG. 3A) and B16-F10 (FIG. 3B) melanoma cells were selected fortesting the anti-proliferative effect of docetaxel and TH 1902. When TH 1902 biological effects were monitored, the half-maximal inhibitory concentration (IC5o) ofTH1902 was similar to that of docetaxel, averaging 0.38 vs. 0.39 nM, respectively, in human SK-MEL-28 cells, and 2.57 vs. 1.72 nM in murine B16-F10 cells (FIG. 3C), indicating that the anti-proliferative property of docetaxel was unaffected upon its conjugation with a cleavable linker to TH19P01. As the cytotoxicity of docetaxel is associated with its ability to induce cell-cycle arrest and apoptosis, the apoptotic effects of low and high concentrations of docetaxel and TH1902 against B16-F10 cells was investigated. Cells were treated with 0.1 or 2 pM docetaxel (FIG. 3D, black bars), or 0.05 and 1 pM TH 1902 (bearing equimolar conjugated docetaxel concentrations; FIG. 3D, grey bars) for 15 minutes, rinsed, and further incubated in complete medium for up to 96 hours. Apoptosis was then assessed by flow cytometry as described in Example 1. While only elevated docetaxel concentrations induced apoptosis, TH1902 incrementally induced higher apoptosis in a dosedependent manner (FIG. 3D). Given that docetaxel is known to induce senescence in B16-F10 cells

[0022] and to limit proliferative capacity

[0023] , the respective effects of docetaxel and TH1902 on senescence was next tested as described in Example 1 (FIG. 3E). Relative to control conditions, the SA-|3-gal activity-reliant senescence marker was increased in docetaxel-treated cells as well as in those treated with the senescence inducer etoposide (positive control). Relative to docetaxel, TH 1902 further increased senescence as measured by SA-|3-gal activity (FIG. 3F). The increased cellular senescence induction was additionally confirmed morphologically in cells treated with TH 1902, which became larger and flatter relative to the control or docetaxel conditions (FIG. 3G). This data provides evidence that increased apoptotic and senescence effects may support the antitumor effects of TH1902 on B16-F10 melanoma cells.

[0360] Example 5: TH1902 stops tumor growth and triggers leukocyte infiltration in an immunocompetent syngeneic “cold” tumor model.

[0361] B16-F10 melanoma syngeneic tumors were generated (FIG. 4A), with tumor sizes monitored as described in Example 1. Tumors in xenograft-bearing, vehicle-treated mice grew at an exponential rate (FIG. 4A, black circles). Partial inhibition of tumor growth was observed after IV administration of 15 mg / kg / wk docetaxel (FIG. 4A, squares), whereas treatment with a docetaxel-equivalent quantity of TH 1902 (35 mg / kg / wk) induced tumor regression after two treatments over the period measured (FIG. 4A, triangles). Due to rapid tumor growth, only two administrations of the test articles could be performed on a weekly schedule. B16-F10 melanoma tumors from the mice treated with either vehicle, docetaxel, or TH 1902 were then excised, fixed in formalin, and immunohistochemically examined (FIG. 4B). Within the tumors shown, necrotic regions are present and some areas are full of actual cancer cells with significantly enlarged morphology, as previously observed within the TH1902-treated MDA-MB-231 tumors (FIG. 4C). Here again, significant infiltration of immune cells (CD45+ total leukocytes) into the tumors from TH1902-treated animals was detected (FIG. 4D). Tumor parenchyma from animals treated with vehicle showed little immunodetection of CD45 (FIG. 4C, left panels), indicating an absence of leukocytes, while tumors from animals treated with docetaxel exhibited only slightly higher leukocyte infiltration (FIG. 4C, middle panels). In both cases, however, the staining was limited to the outer tumor periphery. This indicates that the tumors are of the immune-excluded phenotype, rather than the related immune-desert phenotype where T-cells are notably absent from either the parenchyma or the stroma of the tumor

[0024] , It is axiomatic that tumors lacking lymphoid cell infiltration are unlikely to respond to checkpoint inhibitors (CPI); this has been established for infiltration of lymphoid cells [25, 26], Animals treated with TH1902, however, showed significantly greater tumor levels of pan-immune cell marker CD45 staining (FIG. 4C, right panels), and quantification confirmed infiltration of most of the leukocytes within the tumor parenchyma (FIG. 4D). Body weights of the mice administered test articles or vehicle were followed as a gross indicator of morbidity. The mice bearing B16-F10 tumors exhibited similar body weights whether they had been administered vehicle, docetaxel, or TH1902 (FIG. 4E), indicating that animal weights remained within the acceptable range.

[0362] Example 6: TH1902 triggers immune cell infiltration within tumors.

[0363] The immune response to tumors is a complex orchestration involving myriad cell types, interacting membrane proteins, and soluble effectors

[0027] , Docetaxel induced moderate lymphocyte cell infiltration of all subclasses by IHC, including cytotoxic T-cells, helper T-cells, regulatory T-cells, and natural killer (NK) cells within the tumor parenchyma (FIG. 5A, horizontal middle panels). However, the increase from TH1902 treatment was systematically and significantly greater than that from docetaxel for all classes of lymphocytes measured (FIG. 5A, horizontal lower panels, and FIG. 5B, grey bars). The CD3 T-cell marker was slightly increased with docetaxel and significantly increased with TH1902 relative to vehicle. IHC data also showed that TH 1902 treatment significantly increased the expression of CD8+ (or cytotoxic) T-cells that are known for their antitumoral immune response. Furthermore, TH1902 increased the infiltration of CD4 T-cells more than did docetaxel, whereas TH1902 induced regulatory T-cell (Treg; FoxP3) infiltration levels. Moreover, CD 161c NK marker expression was increased in TH1902-treated mice relative to vehicle- or docetaxel treated-mice (FIG. 5B).

[0364] Macrophages also represent an important class of immune cells within tumors. In fact, tumor-associated macrophages (TAMs) are generally the most abundant immune cell population within most tumors

[0028] , TAMs can either enhance or antagonize the cytotoxic activity of immune cells, effects often attributed to two distinct subpopulations, M1 and M2, respectively

[0029] , These are not two separate lineages of macrophages but cells whose “polarization” has been switched to M1 or M2 by local humoral factors

[0030] , Much like tumor-infiltrating lymphocytes (TILs), the numbers of tumor-infiltrated TAMs (F4 / 80+), and those of M1 (CD68+) and M2 (CD206+) macrophages, were slightly increased by docetaxel treatment but greatly increased by treatment with TH1902 (FIGs. 5C and 5D). Surprisingly, expression of the CD206+M2 macrophages marker showed the same tendency as CD68+M1 macrophages for TH1902. Thus, the stimulation of leukocyte infiltration of tumors by TH1902 administration applies to multiple classes of immune cells.

[0365] Example 7: TH1902 induces immune-stimulated apoptosis.

[0366] Regardless of the specific lineages of cells that infiltrate the tumor, the effectiveness of a treatment often relies on induction of apoptosis in tumor cells

[0031] , One mechanism by which immune cells (mainly NK cells and cytotoxic T-cells) eliminate other cells is through the granzyme B / perforin apoptotic pathway. The effect of both docetaxel and TH 1902 on this pathway was thus investigated. A small increase for perforin and granzyme B, but not for caspase-3, was found to occur when the animals were treated with docetaxel, relative to the vehicle-treated group (FIG. 6A). However, large, significant increases in perforin, granzyme B, and caspase-3 staining (5-7- fold over docetaxel) occurred following treatment with TH 1902, indicating that TH1902-induced tumor cell apoptosis is at least partially mediated through immune cells (FIG. 6B). Overall, these data support the pronounced effect of TH 1902 on the antitumor response of CD8+ T-cell effectors and NK cells, which lead, following cancer cell death, to complete tumor regression.

[0367] Example 8: Effect of TH1902 / anti-PD-L1 combination against B16-F10 tumors. Treatment of the B16-F10 tumors with TH1902 appears to enable widespread immune cell infiltration of the tumors and to greatly enhance immune cell-induced apoptosis. TH1902 also appears to circumvent the immune-excluded character of these “cold” tumors. Consequently, combining TH1902 with checkpoint inhibitors (CPIs) appears to be feasible. Since anti-PD-L1 immunotherapy is known to be ineffective in the B16-F10 syngeneic model

[0032] , the combination of this antibody with docetaxel orTH1902 was evaluated. To assess synergistic treatment effects, doses of docetaxel and TH1902 were halved. As observed, tumor growth was similar in the vehicle group and in mice treated with the isotype control antibodies (a control for the anti-PD-L1) while administration of anti-PD-L1 itself produced a small but not statistically significant (p = 0.0702) decrease in tumor growth by 35%. These results are similar to previous reports in this tumor model

[0033] , in which docetaxel administration produced a significant but small decrease in tumor growth of 49%, while TH 1902 produced a much larger, significant decrease of 92% (FIG. 7A). While combination treatment with anti-PD-L1 and docetaxel produced a small further decrease in tumor growth (66%), the combination of anti-PD-L1 and TH 1902 produced a significant decrease in tumor size, showing tumor regression at day 11 (regression of 30% relative to initial tumor volume at treatment initiation). The enhanced inhibition of tumor growth by the combination of TH1902 and anti-PD-L1 became more apparent as the experiment continued. While single-drug administrations of docetaxel or anti-PD-L1 showed only partial inhibition of tumor growth, TH1902 effectively reduced this growth after two cycles (FIG. 7A). Interestingly, while the anti-PD-L1 / docetaxel combination was unable to further reduce tumor growth, the anti- PD-L1 / TH1902 combination showed significant and greater tumor growth reduction at day 14 post-treatment relative to that obtained with TH 1902 alone. Docetaxel and TH 1902 both appeared to be well-tolerated, as mouse body weight was almost unaffected when compared to control groups (FIG. 7D). However, anti-PD-L1 , used either alone or in combination with docetaxel or TH1902, led to similar body weight loss, suggesting that the addition ofTH1902 had limited impact on mice body weight (FIG. 7D). Overall, these data support the therapeutic efficiency of TH1902 as monotherapy, which is significantly increased when combined with anti-PD-L1. It also demonstrates that TH1902 reverses the resistance of the tumors to CPIs.

[0368] These results could not be expected since B16-F10 melanoma tumors are generally considered to be immunologically cold with limited immune cell infiltration and poor response to immune checkpoint inhibitors (See https: / / drugdevelopment.labcorp.com / industry- solutions / oncology / preclinical / tumor-spotlights / b16-f10-a-murine-melanoma-model.html and Ueha et al., Cancer Immunol Res (2015) 3 (6): 631-640 [FIG. 8]).

[0369] Example 9: Effects of combining different levels of TH1902 with anti-PD-L1 on tumor growth in survival study. A survival study on mice treated with (or without) anti-PD-L1 along with different dosages of TH1902 was next performed. The results depicted in FIG. 7C show that TH1902 exhibits a strong dose-dependent inhibition of tumor growth. Treatment with the lowest dose of TH1902 or with anti-PD-L1 alone yielded slight but not significant (p = 0.1394 and 0.1787, respectively) tumor growth inhibition (TGI) relative to that in vehicle-treated animals; tumor sizes at 10 days following treatment start were halved similarly to those in vehicle-treated animals, with a median survival of either 14 (TH1902 4.37 mg / kg / wk and anti-PD-L1 , both p = 0.0521) or 11.5 (vehicle) days. However, the tumors in animals treated with either of the higher doses of TH1902 were significantly smaller, with TGI of 95% (8.75 mg / kg / wk) or regression of 54% relative to initial tumor volume at treatment initiation (17.5 mg / kg / wk) and a median survival of 17 and 24 days, respectively. Mice in the survival study showed much better tolerance to the anti-PD-L1 than was seen in the efficacy study. None of the groups of mice in this study displayed significant weight loss, but there did appear to be more weight gain in mice that did not receive the IP injections of control IgG or of the anti-PD-L1 mAb; this may be attributed to the rapid exponential growth of some tumors. Survival curves for each of the nine groups of mice were generated. The two groups receiving 17.5 mg / kg / wk TH1902 (with and without anti-PD-L1) displayed the longest median survival: 24 and 32.5 days, respectively (FIG. 7B). Statistically, animals treated with both of the higher concentrations of TH1902 (8.75 and 17.5 mg / kg / wk) as monotherapy and all three combination TH1902 doses (4.37, 8.75, and 17.5 mg / kg / wk) with anti-PD-L1 exhibited survival curves that were significantly different from that of vehicle-treated animals (Table 2). In addition, the survival curves for all three of those combination TH1902 doses were significantly different (p<0.05) from those of groups treated with TH1902 alone, with increased life spans of 4.5 vs. 2.5, 10 vs. 5.5, and 21 vs. 12.5 days when relative to vehicle, respectively (Table 2). The [TH 1902 17.5 mg / kg / wk I anti-PD-L1] combination significantly and synergistically increased animal survival over either anti-PD-L1 or TH1902 17.5 mg / kg / wk as single agents (21 days increased median survival relative to 2.5 days for anti-PD-L1 alone and 12.5 days for TH1902 17.5 mg / kg / wk) alone).

[0370] Table 2: Results of the survival study in the B16-F10 melanoma xenograft tumor model.

[0371] Groups Dosage Median time survival Increase of life span Significance (p value)

[0372] (mg / kg) (Days) (Days) vs Vehicle group Mono vs Combo

[0373] Vehicle 0 11.5 0 na na

[0374] Control 9 13 1.5 ns (0.1908) na isotype

[0375] Anti-PD-Ll 9 14 2.5 ns (0.0521) na

[0376] TH1902 4.37 14 2.5 ns (0.0521) na

[0377] 8.75 17 5.5 <0.001 na

[0378] 17.5 24 12.5 <0.001 na

[0379] Combination 4.37 + 9 16 4.5 <0.01 <0.05

[0380] TH1902+ 8.75 + 9 21.510<0.001 <0.05

[0381] Anti-PD-Ll 17.5 + 9 32.5 21 <0.001 <0.05

[0382] Example 10: TH1902 induces downstream effectors of the STING pathway and increases the expression of cell surface PD-L1 and MHC-I in B16-F10 melanoma cells.

[0383] Cellular senescence, an irreversible cell-cycle arrest, represents a principal barrier against tumorigenesis

[0034] , Docetaxel has been reported to induce senescence in mouse lung and prostate tumor cells [35, 36], in part through persistent DNA damage response

[0037] , Recently, DNA damage conditions were shown to activate the cGAS (cyclic GMP-AMP synthase) / STING (stimulator of interferon genes) pathway in TNBC cells, a process associated with cell proliferation and invasiveness [38, 39], Given the ability of TH1902 to trigger senescence in B16-F10 melanoma cells, and to inhibit cell proliferation in melanoma (as shown herein) and in TNBC cells [5], the involvement of the STING pathway was next addressed. Cell lysates were harvested following treatment with either docetaxel or TH1902; STING protein expression was assessed by immunoblotting (FIG. 9A). Whereas docetaxel induced densitometric quantification of STING expression, TH1902 did so more effectively at concentrations up to 500 nM (FIG. 9B). In an assessment of downstream effectors of the STING pathway (FIG. 9C), TH1902 was found to trigger expression of the p53 transcription factor known to engage the cGAS / STING pathway, as well as cell senescence [40, 41]; TH1902 also triggered expression of p21 , which is known to maintain the viability of DNA damage-induced senescent cells

[0042] (FIG. 9D, grey bars). Consistent with the activation of the STING pathway, greater increases in the phosphorylation status of TBK1 and p65 by TH1902 treatment relative to docetaxel were also observed (FIG. 9D). TBK1 is an important serine / threonine-protein kinase known to mediate NF-KB signaling and to regulate inflammatory cytokine production and the activation of innate immunity

[0043] , TBK1 , independently from NF-KB, also mediates phosphorylation and nuclear translocation of IRF3, which contributes to the induction of type I interferons

[0044] , Cytokine modulation was next assessed in docetaxel- and TH1902-treated cells. Activation of the STING pathway further correlated with increases in interleukin-6 (IL-6) and tumor necrosis factor (TNF)a transcript levels by TH1902 (FIG. 9E). Finally, PD-L1 (FIG. 9F, lower panels) and MHC-I (FIG. 9F, upper panels) were also induced at the cell surface of TH1902- treated B16-F10 melanoma cells, both of which were also induced by interferon gamma (FIG. 9F, right panels). NF-KB-mediated upregulation of MHC-I is believed to enhance T-cell activation

[0045] ; this may increase tumor cell recognition, ultimately making the melanoma cells more susceptible to cytotoxic CD8+T-cell killing. These data indicate increased NF-KB expression through phosphorylation of p65, which is also associated with favorable response to CPI therapy in patients with melanoma [46, 47], TH1902 therefore has the potential to increase antitumor immunity, and the data highlights molecular in vitro evidence suggesting that compensatory upregulation of the CPI ligand PD-L1 may benefit from counteraction by anti-PD-L1 to achieve maximal antitumor immunity in vivo, consistent with the synergistic effect obtained with the combination of TH1902 and anti-PD-L1 in Example 9.

[0384] Example 11 : TNFa, but not IL-6, triggers PD-L1 and MHC-I cell surface expression in B16- F10 melanoma cells.

[0385] In order to decipher the potential crosstalk that may link TNFa / IL-6 inductions triggered by TH1902 to the increased levels of MHC-I and PD-L1 , B16-F10 melanoma cells were treated with the indicated TNFa or IL-6 concentrations for 96 hours. Dose-dependent increases in cell surface expression of PD-L1 and MHC-I were observed upon TNFa treatment (FIG. 10A), whereas cell surface expression of both proteins remained unchanged upon IL-6 treatment (FIG. 10B). These data suggest that TNFa, not IL-6, is the potential main trigger of TH1902-mediated induction of PD-L1 and MHC-I cell surface expression.

[0386] Example 12: Effects of TH1902 in an immunologically cold lung carcinoma model.

[0387] The effect of TH1902 on tumor growth was assessed in another model of immunologically cold tumor expressing SORT1 , the Lewis lung carcinoma (LL / 2) syngeneic model. LL / 2 tumor cells are resistant to immune checkpoint inhibitors (https: / / drugdevelopment.labcorp.com / industry-solutions / oncology / preclinical / tumor-spotlights / ll- 2-an-immunosuppressive-murine-tumor-model.html, FIGs. 11 A and 11 B). The results depicted in FIG. 11C shows that treatment with TH1902 induces an almost complete inhibition of LL / 2 tumor growth.

[0388] The results presented herein provide evidence that a conjugate comprising a sortilin- targeting peptide linked to a chemotherapeutic agent permits to induce immune cell infiltration and cytotoxic cell-mediated tumor killing in otherwise immunologically cold tumors, and that combining such conjugate with an immune checkpoint inhibitor leads to eradication of tumors, including those resistant to treatment with immune checkpoint inhibitor monotherapy.

[0389] Although the present invention has been described hereinabove by way of specific embodiments thereof, it can be modified, without departing from the spirit and nature of the subject invention as defined in the appended claims. In the claims, the word "comprising" is used as an open-ended term, substantially equivalent to the phrase "including, but not limited to". The singular forms "a", "an" and "the" include corresponding plural references unless the context clearly dictates otherwise.

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Claims

WHAT IS CLAIMED IS:1 . A method for (i) enhancing the anti-tumor immune response in a subject suffering from a Sortilin-expressing cancer and / or (ii) treating a subject suffering from a Sortilin-expressing cancer resistant to an immunotherapy, the method comprising administering to the subject an effective amount of a conjugate compound or a pharmaceutically acceptable salt thereof, wherein the conjugate compound has the formula of A-(B)n, whereinA is a peptide compound of 30 residues or less comprising an amino acid sequence having at least 60% sequence identity to the amino acid sequence of any one of SEQ ID NOs: 1- 13:X1X2X3X4X5GVX6AKAGVX NX8FKSESY (SEQ ID NO: 1)(X9)nGVXioAKAGVXn NX12FKSESY (SEQ ID NO: 2)YKX13LRRX14APRWDX15PLRDPALRX16X17L (SEQ ID NO: 3)YKX18LRR(X19)NPLRDPALRX2OX2IL (SEQ ID NO: 4)IKLSGGVQAKAGVINMDKSESM (SEQ ID NO: 5)IKLSGGVQAKAGVINMFKSESY (SEQ ID NO: 6)I KLSGG VQAKAGVI NM FKSESYK (SEQ ID NO: 7)GVQAKAGVINMFKSESY (SEQ ID NO: 8)GVRAKAGVRNMFKSESY (SEQ ID NO: 9)GVRAKAGVRN(Nle)FKSESY (SEQ ID NO: 10)YKSLRRKAPRWDAPLRDPALRQLL (SEQ ID NO: 1 1)YKSLRRKAPRWDAYLRDPALRQLL (SEQ ID NO: 12)YKSLRRKAPRWDAYLRDPALRPLL (SEQ ID NO: 13) whereinXi , X2, X3, X4, X5, Xe, X7, Xs, X9, X10, X11 , X12, X13, X14, Xis, Xis and X19 are independently chosen from any amino acid;X16, X17, X20 and X21 are independently chosen from Q, P, Y, I and L; n is 0, 1 , 2, 3, 4 or 5; when X9 is present more than once, each of said X9is independently chosen from any amino acid; when X19 is present more than once, each of said X9is independently chosen from any amino acid, optionally the peptide compound is cyclic,B is at least one therapeutic agent, wherein B is connected to A directly or via a linker.

2. The method of claim 1 , wherein the peptide compound comprises an amino acid sequence having at least 90% sequence identity to the amino acid sequence of any one of SEQ ID NOs: 1- 13.

3. The method of claim 1 or 2, wherein the peptide compound comprises the amino acid sequence of any one of SEQ ID NOs: 1-13, and further comprises 1 to 3 additional amino acids at its amino- and / or carboxy-terminal end.

4. The method of claim 3, wherein the peptide compound comprises a cysteine residue at its amino- and / or carboxy-terminal end.

5. The method of claim 4, wherein the peptide compound comprises one of the following amino acid sequences:Z1X1X2X3X4X5G X6AKAG X7NX8FKSESYZ2 (SEQ ID NO:34)ZT (X9)nGVXioAKAGVXi 1 NX12FKSESYZ2(SEQ ID NO:35)Z1YkX13LRRX14APRWDX15PLRDPALRX16X17LZ2(SEQ ID NO:36)Z1YKX18LRR(X19)NPLRDPALRX20X21LZ2(SEQ ID NO:37)ZT I KLSGGVQAKAGVI NM DKSESMZ2(SEQ ID NO:38)ZT I KLSG G VQAKAG VI N M FKSESYZ2(SEQ ID NO:39)ZT I KLSG G VQAKAG VI N M FKSESYKZ2(SEQ ID NQ:40)Z! GVQAKAGVI NM FKSESYZ2(SEQ ID NO:41)Z1GVRAKAGVRNMFKSESYZ2(SEQ ID NO:42)Z! G VRAKAG VRN (N le) F KSESYZ2(SEQ ID NO:43)Z1YKSLRRKAPRWDAPLRDPALRQLLZ2(SEQ ID NO:44)Z1YKSLRRKAPRWDAYLRDPALRQLLZ2(SEQ ID NO:45)ZIYKSLRRKAPRWDAYLRDPALRPLL Z2(SEQ ID NO:46), wherein Xi-X2iare as defined in claim 1 ; Zi is a cysteine residue or is absent; Z2is a cysteine residue or is absent, and at least one of Zi and Z2is present.

6. The method of claim 1 or 5, wherein the peptide compound comprises the amino acid sequence of SEQ ID NO: 1 or 2.

7. The method of claim 1 or 5, wherein the peptide compound comprises the amino acid sequence of SEQ ID NO: 3 or 4.

8. The method of claim 1 or 5, wherein the peptide compound comprises the amino acid sequence of SEQ ID NO: 5.

9. The method of claim 1 or 5, wherein the peptide compound comprises the amino acid sequence of SEQ ID NO: 6.The method of claim 1 or 5, wherein the peptide compound comprises the amino acid sequence of SEQ ID NO: 7.

11. The method of claim 1 or 5, wherein the peptide compound comprises the amino acid sequence of SEQ ID NO: 8.

12. The method of claim 1 or 5, wherein the peptide compound comprises the amino acid sequence of SEQ ID NO: 9.

13. The method of claim 1 or 5, wherein the peptide compound comprises the amino acid sequence of SEQ ID NO: 10.

14. The method of claim 13, wherein the peptide compound comprises the amino acid sequence GVRAKAGVRN(Nle)FKSESYC (SEQ ID NO:47).

15. The method of claim 1 or 5, wherein the peptide compound comprises the amino acid sequence of SEQ ID NO: 11 .

16. The method of claim 1 or 5, wherein the peptide compound comprises the amino acid sequence of SEQ ID NO: 12.

17. The method of claim 1 or 5, wherein the peptide compound comprises the amino acid sequence of SEQ ID NO: 13.

18. The method of any one of claims 1 to 17, wherein the peptide compound comprises at least one modifying group at its amino- and / or carboxy-terminal end.

19. The method of claim 17, wherein the at least one modifying group is acetyl or succinyl.

20. The method of claim 1 , wherein the peptide compound is represented by SEQ ID NO:14, SEQ ID NO:15, SEQ ID NO:16, SEQ ID NO:17, SEQ ID NO:18, or SEQ ID NO:47:Acetyl-GVRAKAGVRNMFKSESY (SEQ ID NO: 14)Acetyl-GVRAKAGVRN(Nle)FKSESY (SEQ ID NO: 15)Acetyl-YKSLRRKAPRWDAPLRDPALRQLL (SEQ ID NO: 16)Acetyl-YKSLRRKAPRWDAYLRDPALRQLL (SEQ ID NO: 17)Acetyl-YKSLRRKAPRWDAYLRDPALRPLL (SEQ ID NO: 18)Acetyl-GVRAKAGVRN(Nle)FKSESYC (SEQ ID NO: 48).21 . The method of any one of claims 1 to 20, wherein B is connected to A at a free amine of said peptide compound, at an N-terminal position of said peptide compound, at a free -SH of said peptide compound, and / or at a free carboxyl of said peptide compound.

22. The method of any one of claims 1 to 21 , wherein B is connected to A via a linker.

23. The method of any one of claims 1 to 22, wherein the conjugate is represented by SEQ ID NO:23 or SEQ ID NO:24:GVRAK(J1)AGVRN(Nle)FK(J2)SESY (SEQ ID NO:22);Acetyl-GVRAK(J1)AGVRN(Nle)FK(J2)SESY (SEQ ID NO:23); wherein J1and J2are each independently a therapeutic agent attached to a lysine (K) residue.

24. The method of any one of claims 1 to 23, wherein the therapeutic agent is an antitumor agent, for example a radionuclide or a chemotherapeutic agent.

25. The method of claim 24, wherein the chemotherapeutic agent is a taxane.

26. The method of claim 25, wherein the chemotherapeutic agent is docetaxel.

27. The method of any one of claims 1 to 26, further comprising treating the subject with an immunotherapy.

28. The method of any one of claims 1 to 27, wherein the immunotherapy comprises immune checkpoint inhibitor (ICI) therapy.

29. The method of claim 28, wherein the ICI is a Programmed cell death-1 (PD-1) inhibitor, a cytotoxic T-lymphocyte-associated antigen 4 (CTLA-4) inhibitor, or a Programmed death-ligand 1 (PD-L1) inhibitor.

30. The method of claim 28 or 29, wherein the ICI is a blocking antibody.31 . The method of claim 28 or 29, wherein the ICI is a PD-L1 inhibitor.

32. The method of any one of claims 1 to 31 , wherein the cancer is an immunologically cold cancer.

33. The method of any one of claims 1 to 32, wherein the cancer is brain cancer, pancreatic cancer, ovarian cancer, breast cancer, endometrial cancer, thyroid cancer, lung cancer, prostate cancer or melanoma.

34. Use of the conjugate compound or pharmaceutically acceptable salt thereof defined in any one of claims 1 to 26 for (i) enhancing the anti-tumor immune response in a subject suffering from a Sortilin-expressing cancer and / or (ii) treating a subject suffering from a Sortilin-expressing cancer resistant to an immunotherapy.

35. Use of the conjugate compound or pharmaceutically acceptable salt thereof defined in any one of claims 1 to 26 forthe manufacture of a medicament for (i) enhancing the anti-tumor immune response in a subject suffering from a Sortilin-expressing cancer and / or (ii) treating a subject suffering from a Sortilin-expressing cancer resistant to an immunotherapy.

36. The use of claim 34 or 35, wherein the conjugate compound, pharmaceutically acceptable salt thereof, or medicament is for use in combination with an immunotherapy.

37. The use of any one of claims 34 to 36, wherein the immunotherapy comprises immune checkpoint inhibitor (ICI) therapy.

38. The use of claim 37, wherein the ICI is a Programmed cell death-1 (PD-1) inhibitor, a cytotoxic T-lymphocyte-associated antigen 4 (CTLA-4) inhibitor, or a Programmed death-ligand 1 (PD-L1) inhibitor.

39. The use of claim 37 or 38, wherein the ICI is a blocking antibody.

40. The use of claim 38 or 39, wherein the ICI is a PD-L1 inhibitor.

41. The use of any one of claims 34 to 40, wherein the cancer is an immunologically cold cancer.

42. The use of any one of claims 34 to 41 , wherein the cancer is brain cancer, pancreatic cancer, ovarian cancer, breast cancer, endometrial cancer, thyroid cancer, lung cancer, prostate cancer or melanoma.

43. The conjugate compound or pharmaceutically acceptable salt thereof defined in any one of claims 1 to 26 for use in (i) enhancing the anti-tumor immune response in a subject suffering from a Sortilin-expressing cancer and / or (ii) treating a subject suffering from a Sortilin-expressing cancer resistant to an immunotherapy.

44. The conjugate compound or pharmaceutically acceptable salt thereof for use according to claim 43, wherein the peptide compound is for use in combination with an immunotherapy.

45. The conjugate compound or pharmaceutically acceptable salt thereof for use according to claim 43 or 44, wherein the immunotherapy comprises immune checkpoint inhibitor (ICI) therapy.

46. The conjugate compound or pharmaceutically acceptable salt thereof for use according to claim 45, wherein the ICI is a Programmed cell death-1 (PD-1) inhibitor, a cytotoxic T-lymphocyte- associated antigen 4 (CTLA-4) inhibitor, or a Programmed death-ligand 1 (PD-L1) inhibitor.

47. The conjugate compound or pharmaceutically acceptable salt thereof for use according to claim 45 or 46, wherein the ICI is a blocking antibody.

48. The conjugate compound or pharmaceutically acceptable salt thereof for use according to claim 46 or 47, wherein the ICI is a PD-L1 inhibitor.

49. The conjugate compound or pharmaceutically acceptable salt thereof for use according to any one of claims 43 to 48, wherein the cancer is an immunologically cold cancer.

50. The conjugate compound or pharmaceutically acceptable salt thereof for use according to any one of claims 43 to 49, wherein the cancer is brain cancer, pancreatic cancer, ovarian cancer, breast cancer, endometrial cancer, thyroid cancer, lung cancer, prostate cancer or melanoma.51 . A method for treating a Sortilin-expressing cancer in a subject comprising administering to the subject a therapeutically effective amount of the conjugate compound or pharmaceutically acceptable salt thereof defined in any one of claims 1 to 26 in combination with an immunotherapy.

52. The method of claim 51 , wherein the immunotherapy comprises immune checkpoint inhibitor (ICI) therapy.

53. The method of claim 52, wherein the ICI is a Programmed cell death-1 (PD-1) inhibitor, a cytotoxic T-lymphocyte-associated antigen 4 (CTLA-4) inhibitor, or a Programmed death-ligand 1 (PD-L1) inhibitor.

54. The method of claim 52 or 53, wherein the ICI is a blocking antibody.

55. The method of claim 53 or 54, wherein the ICI is a PD-L1 inhibitor.

56. The method of any one of claims 51 to 55, wherein the cancer is an immunologically cold cancer.

57. The method of any one of claim 51 to 56, wherein the cancer is brain cancer, pancreatic cancer, ovarian cancer, breast cancer, endometrial cancer, thyroid cancer, lung cancer, prostate cancer or melanoma.

58. The method of any one of claim 51 to 57, wherein the conjugate compound or pharmaceutically acceptable salt thereof and the immunotherapy are in distinct compositions.

59. The method of any one of claim 51 to 57, wherein the conjugate compound or pharmaceutically acceptable salt thereof and the immunotherapy are in the same composition.

60. Use of the conjugate compound or pharmaceutically acceptable salt thereof defined in any one of claims 1 to 26 in combination with an immunotherapy for the treatment of a Sortilin- expressing cancer.61 . Use of the conjugate compound or pharmaceutically acceptable salt thereof defined in any one of claims 1 to 26 in combination with an immunotherapy for the manufacture of a medicament for the treatment of a Sortilin-expressing cancer.

62. The use of claim 60 or 61 , wherein the immunotherapy comprises immune checkpoint inhibitor (ICI) therapy.

63. The use of claim 62, wherein the ICI is a Programmed cell death-1 (PD-1) inhibitor, a cytotoxic T-lymphocyte-associated antigen 4 (CTLA-4) inhibitor, or a Programmed death-ligand 1 (PD-L1) inhibitor.

64. The use of claim 62 or 63, wherein the ICI is a blocking antibody.

65. The use of claim 63 or 64, wherein the ICI is a PD-L1 inhibitor.

66. The use of any one of claims 60 to 65, wherein the cancer is an immunologically cold cancer.

67. The use of any one of claims 61 to 66, wherein the cancer is brain cancer, pancreatic cancer, ovarian cancer, breast cancer, endometrial cancer, thyroid cancer, lung cancer, prostate cancer or melanoma.

68. The use of any one of claims 61 to 67, wherein the conjugate compound or pharmaceutically acceptable salt thereof and the immunotherapy are in distinct compositions.

69. The use of any one of claims 61 to 67, wherein the conjugate compound or pharmaceutically acceptable salt thereof and the immunotherapy are in the same composition.

70. A combination therapy comprising the conjugate compound or pharmaceutically acceptable salt thereof defined in any one of claims 1 to 26 and an immunotherapy for use in the treatment of a Sortilin-expressing cancer.

71. The combination therapy for use according to claim 70, wherein the immunotherapy comprises immune checkpoint inhibitor (ICI) therapy.

72. The combination therapy for use according to claim 71 , wherein the ICI is a Programmed cell death-1 (PD-1) inhibitor, a cytotoxic T-lymphocyte-associated antigen 4 (CTLA-4) inhibitor, or a Programmed death-ligand 1 (PD-L1) inhibitor.

73. The combination therapy for use according to claim 71 or 72, wherein the ICI is a blocking antibody.

74. The combination therapy for use according to claim 72 or 73, wherein the ICI is a PD-L1 inhibitor.

75. The combination therapy for use according to any one of claims 71 to 74, wherein the cancer is an immunologically cold cancer.

76. The combination therapy for use according to any one of claim 70 to 75, wherein the cancer is brain cancer, pancreatic cancer, ovarian cancer, breast cancer, endometrial cancer, thyroid cancer, lung cancer, prostate cancer or melanoma.

77. The combination therapy for use according to any one of claim 70 to 76, wherein the conjugate compound or pharmaceutically acceptable salt thereof and the immunotherapy are in distinct compositions.

78. The combination therapy for use according to any one of claim 70 to 76, wherein the conjugate compound or pharmaceutically acceptable salt thereof and the immunotherapy are in the same composition.

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