Peptide conjugates of maytansinoids and uses thereof

Abstract

The present application relates to conjugates of maytansinoids comprising a sortilin- targeting peptide and a linker, and salts thereof. These conjugates were shown to exhibit more potent antitumor effects than corresponding unconjugated maytansinoids in mouse models of Sortilin-expressing colorectal and breast cancers. The present application also relates to pharmaceutical compositions comprising the conjugates or salts thereof, as well as to methods and uses of the conjugates, salts thereof or pharmaceutical compositions for the treatment of Sortilin-expressing cancers.

Description

[0001] PEPTIDE CONJUGATES OF MAYTANSINOIDS AND USES THEREOF

[0002] CROSS REFERENCE TO RELATED APPLICATIONS

[0003] The present application claims the benefit of U.S. provisional Patent application No. 63 / 648,310, filed on May 16, 2024. The entire content of this application is incorporated herein by reference.

[0004] SEQUENCE LISTING

[0005] A sequence listing is submitted herewith as an XML file named G11718-00494-SSS_Seq listing.xml, that was created on May 15, 2025, and having a size of ~21 ,826 bytes. The content of the aforementioned file is hereby incorporated by reference in its entirety.

[0006] TECHNICAL FIELD

[0007] The present invention generally relates to the field of oncology, and more particularly to the treatment of cancers using microtubulin polymerization inhibitors.

[0008] BACKGROUND ART

[0009] Mitotic inhibitors (also called microtubule inhibitors or tubulin inhibitors) are drugs that inhibit mitosis, or cell division, and are used in the treatment of various diseases including cancer. Mitotic inhibitors prevent tumor cells from undergoing mitosis by disrupting microtubule polymerization, thus preventing cancerous growth.

[0010] Maytansine and its derivatives (generally referred to as maytansinoids) are members of the ansamycins superfamily and contains a 19-member macrocyclic lactam attached to a chlorinated benzene. Maytansine is originally isolated from an Ethiopian shrub Maytenus ovatus and exerts extremely high anti-mitotic potency. Maytansinoids are microtubule-targeting agents that share the same binding site with vinca and function by depolymerizing microtubules and arresting cells in the mitosis stage. Maytansinoids exhibit over 100-fold elevated cytotoxicity in cells as compared to vinca alkaloids. However, in human clinical trials, maytansine showed a small therapeutic window due to its neurotoxicity and harmful effects on the gastrointestinal tract.

[0011] Common side effects associated with maytansinoid-containing therapies may include bone marrow suppression, peripheral neuropathy, liver toxicity, gastrointestinal issues (nausea, vomiting, diarrhea, and constipation), fatigue, hair loss (alopecia), skin reactions and mucositis.

[0012] Colorectal cancer (CRC) is one of the leading causes of mortality and morbidity in the world. The management of colorectal cancer involves a combination of therapies that may include surgery, chemotherapy, radiation therapy, targeted therapy, and immunotherapy. The choice of treatment largely depends on the stage of the cancer, its location, the patient’s overall health, and specific genetic markers of the tumor. Surgery is commonly the first-line treatment for localized colorectal cancer and may involve the removal of the tumor and surrounding tissue.

[0013] In advanced stages, chemotherapy is often employed to shrink tumors, control disease progression, and alleviate symptoms. Historically, the treatment of metastatic CRC (mCRC) has been based on the combination of cytotoxic agents, such as irinotecan or oxaliplatin, associated with 5-FU and leucovorin or capecitabine (FOLFIRI / FOLFOX / FOLFOXIRI or CAPIRI / CAPOX regimens), which resulted in an average survival of 18 months.

[0014] Targeted therapies using antibodies designed to interfere with specific molecular targets associated with cancer growth and progression are also used. Drugs like bevacizumab (Avastin®) that target vascular endothelial growth factor (VEGF), and cetuximab (Erbitux®) or panitumumab (Vectibix®) that target epidermal growth factor receptor (EGFR), respectively, and are used in certain metastatic cases. Immunotherapy, which harnesses the patient's immune system to attack cancer cells, has emerged as a treatment option for some patients with advanced colorectal cancer, particularly those with high microsatellite instability (MSI-H) or mismatch repair deficiency (dMMR).

[0015] Mutations in the BRAF gene are examples of such oncogenic events and are found in about 10% of CRC patients. Mutations in BRAF is associated with poor prognosis and resistance to standard therapies, notably anti-EGFR therapies, with a median overall survival (mOS) of approximately 12 months. Similarly, KRAS-mutant CRCs, which correspond to about 40% of CRCs, are insensitive to clinically-used EGFR-targeting antibodies, with an mOS of about 17 months.

[0016] There is thus a need for the development of novel approaches for the treatment of cancers, and more particularly cancers that are resistant to conventional therapies such as EGFR- and / or VEGF-targeting antibodies.

[0017] There is thus a need for novel approaches for the use of maytansinoids in the treatment of cancers, including cancers that are resistant to conventional therapies such as EGFR- and / or VEGF-targeting antibodies.

[0018] The present description refers to a number of documents, the content of which is herein incorporated by reference in their entirety.

[0019] SUMMARY

[0020] The present disclosure relates to products, such as conjugates or salts thereof, and compositions thereof, and methods and uses thereof, such as for the treatment of cancer.

[0021] In various aspects and embodiments, the present disclosure provides the following items 1 to 66:

[0022] 1. A conjugate comprising the following structure I, or a pharmaceutically acceptable salt thereof: A — L — B (i); wherein

[0023] A is a maytansinoid;

[0024] B is a peptide binding to a Sortilin receptor, wherein the peptide comprises an amino acid sequence having at least 60% identity with the amino acid sequence set forth in SEQ ID NO: 1 or 2:

[0025] X1GVRAKAGVRN(Nle)FKSESYX2(SEQ ID NO:1)

[0026] X1YKSLRRKAPRWDAPLRDPALRQLLX2(SEQ ID NO:2); wherein X1is cysteine (C) or is absent, and X2is cysteine (C) or is absent;

[0027] L is a linker, wherein a first end of the linker is attached to a side chain of one or more of the amino acid or modified side chain residues of the peptide, and / or to the N- and / or C- terminal end of the peptide, and a second end of the linker is attached to the maytansinoid.

[0028] 2. The conjugate of item 1 , wherein L is a cleavable linker, wherein a first end of the linker is attached to a side chain of one or more of the lysine (K) and / or cysteine (C) residues of the peptide, and / or to the N- and / or C-terminal end of the peptide, and a second end of the linker is attached to a sulfur atom of the maytansinoid.

[0029] 3. The conjugate or pharmaceutically acceptable salt thereof of item 1 or 2, wherein the conjugate or salt thereof is of one of the following formulas la, lb, Ic or Id:

[0030] A1 — L1 — B (la);

[0031] A1 — L1 — B — L2 — A2 (lb); wherein L1 , L2, L3 and L4 are linkers which may be the same or different, and A1 , A2, A3 and A4 are maytansinoid molecules which may be the same or different. 4. The conjugate or pharmaceutically acceptable salt thereof of any one of items 1 to 3, wherein the maytansinoid is DM1 (N2-deacetyl-N2-(3-mercapto-1-oxopropyl)-maytansine, mertansine) and / or DM4 (N2-deacetyl-N2-(4-mercapto-4-methyl-1 -oxopentyl) maytansine, ravtansine).

[0032] 5. The conjugate or pharmaceutically acceptable salt thereof of item 4, wherein the maytansinoid is DM1.

[0033] 6. The conjugate or pharmaceutically acceptable salt thereof of item 4, wherein the maytansinoid is DM4.

[0034] 7. The conjugate or pharmaceutically acceptable salt thereof of any one of items 1 to 6, wherein the peptide comprises an amino acid sequence having at least 80% identity with the amino acid sequence set forth in SEQ ID NO: 1 or 2.

[0035] 8. The conjugate or pharmaceutically acceptable salt thereof of item 7, wherein the peptide comprises an amino acid sequence having at least 90% identity with the amino acid sequence set forth in SEQ ID NO: 1 or 2.

[0036] 9. The conjugate or pharmaceutically acceptable salt thereof of item 8, wherein the peptide comprises the amino acid sequence set forth in SEQ ID NO: 1 or 2.

[0037] 10. The conjugate or pharmaceutically acceptable salt thereof of item 8, wherein the peptide consists of the amino acid sequence set forth in SEQ ID NO: 1 or 2.

[0038] 11 . The conjugate or pharmaceutically acceptable salt thereof of any one of items 1 to 10, wherein X1and / or X2is C, and wherein the first end of the linker is attached to the side chain of X1and / or X2.

[0039] 12. The conjugate or pharmaceutically acceptable salt thereof of item 11 , wherein X1is absent and X2is C, and wherein the first end of the linker is attached to the side chain of X2.

[0040] 13. The conjugate or pharmaceutically acceptable salt thereof of any one of items 1 to 11 , wherein X1and X2are absent.

[0041] 14. The conjugate or pharmaceutically acceptable salt thereof of any one of items 1 to 13, wherein the linker is a cleavable linker.

[0042] 15. The conjugate or pharmaceutically acceptable salt thereof of any one of items 1 to 13, wherein the cleavable linker is a protease-cleavable linker.

[0043] 16. The conjugate or pharmaceutically acceptable salt thereof of item 15, wherein the protease is a cysteine protease.

[0044] 17. The conjugate or pharmaceutically acceptable salt thereof of item 16, wherein the cysteine protease is cathepsin B.

[0045] 18. The conjugate or pharmaceutically acceptable salt thereof of any one of items 1 to 17, wherein the linker is of one of the following structures:

[0046] • S-X3-CO, wherein X3is a linear or branched Ci-C8alkyl.

[0047] 19. The conjugate or pharmaceutically acceptable salt thereof of item 18, wherein the linker is one of the following structures: S-CH(CH3)-CH2-CH2-CO or S-CH2-CH2-CO. 20. The conjugate or pharmaceutically acceptable salt thereof of item 1 , wherein the conjugate has one of the following structures:

[0048] The conjugate or pharmaceutically acceptable salt thereof of item 20, wherein the conjugate has the following structure: The conjugate or pharmaceutically acceptable salt thereof of item 20, wherein the conjugate has the following structure:

[0049] 23. A pharmaceutical composition comprising the conjugate or salt thereof of any one of items 1 to 22, and a pharmaceutically acceptable excipient.

[0050] 24. A method for treating a Sortilin-expressing cancer in a subject in need thereof comprising administering to the subject an effective amount of the conjugate or salt thereof of any one of items 1 to 22, or the pharmaceutical composition of item 23.

[0051] 25. The method of item 24, wherein the Sortilin-expressing cancer is a hematological cancer, ovarian cancer, endometrial cancer, cervix cancer, skin cancer, brain cancer, breast cancer, colorectal cancer, small intestine cancer, liver cancer, lung cancer, eye cancer, prostate cancer, head and neck cancer, stomach cancer, bone cancer, thyroid cancer, testis cancer, bladder cancer, kidney cancer, or pancreatic cancer.

[0052] 26. The method of item 24 or 25, wherein the Sortilin-expressing cancer is a poor prognosis cancer.

[0053] 27. The method of any one of items 24 to 26, wherein the Sortilin-expressing cancer is an immunologically cold cancer.

[0054] 28. The method of any one of items 24 to 27, wherein the conjugate, salt thereof or composition is administered in combination with one or more additional active agents or therapies for cancer.

[0055] 29. The method of item 28, wherein the one or more additional active agents or therapies for cancer comprise an immunotherapy.

[0056] 30. The method of item 29, wherein the immunotherapy comprises an immune checkpoint inhibitor.

[0057] 31 . The method of item 30, wherein the immune checkpoint inhibitor is a PD1 or PD-L1 inhibitor.

[0058] 32. The method of item 31 , wherein the PD1 or PD-L1 inhibitor is an anti-PD1 or anti-PD-L1 antibody. 33. The method of item 28, wherein the one or more additional active agents or therapies for cancer comprise an anti-vascular endothelial growth factor (VEGF) therapy.

[0059] 34. The method of any one of items 24 to 33, wherein the cancer harbors mutations in a Ras protein or Ras pathway protein.

[0060] 35. The method of item 34, wherein the Ras protein is KRAS.

[0061] 36. The method of item 34, wherein the Ras pathway protein is BRAF.

[0062] 37. The method of any one of items 24 to 36, wherein the cancer is resistant to monotherapy with VEGF inhibitors.

[0063] 38. Use of the conjugate or salt thereof of any one of items 1 to 22, or the pharmaceutical composition of item 23, for treating a Sortilin-expressing cancer in a subject.

[0064] 39. Use of the conjugate or salt thereof of any one of items 1 to 22, or the pharmaceutical composition of item 23, for the manufacture of a medicament for treating a Sortilin-expressing cancer in a subject.

[0065] 40. The use of item 38 or 39, wherein the Sortilin-expressing cancer is a hematological cancer, ovarian cancer, endometrial cancer, cervix cancer, skin cancer, brain cancer, breast cancer, colorectal cancer, small intestine cancer, liver cancer, lung cancer, eye cancer, prostate cancer, head and neck cancer, stomach cancer, bone cancer, thyroid cancer, testis cancer, bladder cancer, kidney cancer, or pancreatic cancer.

[0066] 41. The use of any one of items 38 to 40, wherein the Sortilin-expressing cancer is a poor prognosis cancer.

[0067] 42. The use of any one of items 38 to 41 , wherein the Sortilin-expressing cancer is an immunologically cold cancer.

[0068] 43. The use of any one of items 38 to 42, wherein the conjugate, salt thereof, composition or medicament is for administration in combination with one or more additional active agents or therapies for cancer.

[0069] 44. The use of item 43, wherein the one or more additional active agents or therapies for cancer comprise an immunotherapy.

[0070] 45. The use of item 44, wherein the immunotherapy comprises an immune checkpoint inhibitor.

[0071] 46. The use of item 45, wherein the immune checkpoint inhibitor is a PD1 or PD-L1 inhibitor.

[0072] 47. The use of item 46, wherein the PD1 or PD-L1 inhibitor is an anti-PD1 or anti-PD-L1 antibody.

[0073] 48. The use of item 43, wherein the one or more additional active agents or therapies for cancer comprise an anti-vascular endothelial growth factor (VEGF) therapy.

[0074] 49. The use of any one of items 38 to 48, wherein the cancer harbors mutations in a Ras protein or Ras pathway protein.

[0075] 50. The use of item 49, wherein the Ras protein is KRAS.

[0076] 51 . The use of item 49, wherein the Ras pathway protein is BRAF. 52. The use of any one of items 38 to 51 , wherein the cancer is resistant to monotherapy with VEGF inhibitors.

[0077] 53. The conjugate or salt thereof of any one of items 1 to 22, or the pharmaceutical composition of item 23, for use in the treatment of a Sortilin-expressing cancer in a subject.

[0078] 54. The conjugate, salt thereof or pharmaceutical composition for use according to item 53, wherein the Sortilin-expressing cancer is a hematological cancer, ovarian cancer, endometrial cancer, cervix cancer, skin cancer, brain cancer, breast cancer, colorectal cancer, small intestine cancer, liver cancer, lung cancer, eye cancer, prostate cancer, head and neck cancer, stomach cancer, bone cancer, thyroid cancer, testis cancer, bladder cancer, kidney cancer, or pancreatic cancer.

[0079] 55. The conjugate, salt thereof or pharmaceutical composition for use according to item 54, wherein the Sortilin-expressing cancer is a poor prognosis cancer.

[0080] 56. The conjugate, salt thereof or pharmaceutical composition for use according to item 54 or 55, wherein the Sortilin-expressing cancer is an immunologically cold cancer.

[0081] 57. The conjugate, salt thereof or pharmaceutical composition for use according to any one of items 54 to 56, wherein the conjugate, salt thereof or composition is for administration in combination with one or more additional active agents or therapies for cancer.

[0082] 58. The conjugate, salt thereof or pharmaceutical composition for use according to item 57, wherein the one or more additional active agents or therapies for cancer comprise an immunotherapy.

[0083] 59. The conjugate, salt thereof or pharmaceutical composition for use according to item 58, wherein the immunotherapy comprises an immune checkpoint inhibitor.

[0084] 60. The conjugate, salt thereof or pharmaceutical composition for use according to item 59, wherein the immune checkpoint inhibitor is a PD1 or PD-L1 inhibitor.

[0085] 61. The conjugate, salt thereof or pharmaceutical composition for use according to item 60, wherein the PD1 or PD-L1 inhibitor is an anti-PD1 or anti-PD-L1 antibody.

[0086] 62. The conjugate, salt thereof or pharmaceutical composition for use according to item 57, wherein the one or more additional active agents or therapies for cancer comprise an anti- vascular endothelial growth factor (VEGF) therapy.

[0087] 63. The conjugate, salt thereof or pharmaceutical composition for use according to any one of items 54 to 62, wherein the cancer harbors mutations in a Ras protein or Ras pathway protein.

[0088] 64. The conjugate, salt thereof or pharmaceutical composition for use according to item 63, wherein the Ras protein is KRAS.

[0089] 65. The conjugate, salt thereof or pharmaceutical composition for use according to item 64, wherein the Ras pathway protein is BRAF.

[0090] 66. The conjugate, salt thereof or pharmaceutical composition for use according to of any one of items 54 to 65, wherein the cancer is resistant to monotherapy with VEGF inhibitors. Other objects, advantages and features of the present disclosure 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.

[0091] BRIEF DESCRIPTION OF DRAWINGS

[0092] In the appended drawings:

[0093] FIG. 1A shows the synthetic scheme for the TH19P01-Lys5 13-SPDP-DM1 conjugate (TH2307).

[0094] FIG. 1B shows the synthetic scheme for the TH19P01-Lys5 13-SPDP-DM4 conjugate (TH2311).

[0095] FIG. 1C shows the synthetic scheme for the TH19P01-Lys5 13-SMCC-DM1 conjugate (TH2401).

[0096] FIG. 1D shows the synthetic scheme for the TH19P01-Lys5 13-SPP-DM1 conjugate (TH2402).

[0097] FIGs. 2A-G show the chemical structures of 1 : 1 and 2:1 maytansinoid conjugates according to embodiments disclosed herein. FIG. 2A: non-cleavable 1 :1 TH19P01-Lys5-SMCC-DM1 conjugate. FIG. 2B: Non-cleavable 1 :1 TH19P01-Lys13-SMCC-DM1 conjugate. FIG. 2C: non- cleavable 2:1 TH19P01-Lys5 13-SMCC-DM1 (TH2401) conjugate. FIG. 2D: cleavable 2:1 TH19P01-Lys5 13-SPDP-DM1 (TH2307) conjugate. FIG. 2E: cleavable 2:1 TH19P01-Lys5 13- SPDP-DM4 (TH2311) conjugate. FIG. 2F: cleavable 2:1 TH19P01-SPP5 13-DM1 (TH2402) conjugate. FIG. 2G: cleavable 2:1 TH19P01-SPDP5 13-DM4 conjugate.

[0098] FIGs. 3A-C depict the dose-response anti-cancer activity of TH2307 on HT-29 CRC xenograft model and a comparison with T-DM1 (trastuzumab emtansine, Kadcyla®). HT-29 cells were subcutaneously implanted in the flanks of mice. The animals were treated weekly with either vehicle, DM1 or TH2307 as single agents at the indicated doses. Treatments were initiated when tumor reached about 100-150 mm3. FIG. 3A: Tumor volume was measured using a caliper as described in Example 2. FIG. 3B: Mouse body weight was also monitored for the different groups and remained within the range established for the endpoint limits. FIG. 3C: Published anti-cancer activity of T-DM1 on HT-29 CRC xenograft model (excerpt from FIG. 3A of Chung et al., Eur J Clin Invest. 2020 Apr 29:e13255. doi: 10.1111 / eci.13255).

[0099] FIGs. 4A-B depict the anti-cancer activity of TH2307 (DM1) and TH2311 (DM4) on HCT- 116 CRC xenograft model. HCT-116 cells were subcutaneously implanted in the flanks of mice. The animals were treated weekly with either vehicle, DM1 , TH2307 and TH2311 as single agents at the indicated doses. Treatments were initiated when tumor reached about 100-150 mm3. FIG. 4A: Tumor volume was measured using a caliper as described in Example 2. FIG. 4B: Mouse body weight was also monitored for the different groups and remained within the range established for the endpoint limits.

[0100] FIGs. 5A-B depict the anti-cancer activity of TH2307 (DM1) and TH2311 (DM4) on MDA- MB-231 TNBC xenograft model. MDA-MB-231 cells were subcutaneously implanted in the flanks of mice. The animals were treated weekly with either vehicle, DM1 , TH2307 and TH2311 as single agents at the indicated doses. Treatments were initiated when tumor reached about 100-150 mm3. FIG. 5A: Tumor volume was measured using a caliper as described in Example 2. FIG. 5B: Mouse body weight was also monitored for the different groups and remained within the range established for the endpoint limits.

[0101] FIGs. 6A-B depict the anti-cancer activity of TH2307 (DM1) and TH2311 (DM4) on SKOV3 ovarian cancer xenograft model. SKOV3 cells were subcutaneously implanted in the flanks of mice. The animals were treated weekly with either vehicle, DM1 , TH2307 and TH2311 as single agents at the indicated doses. Treatments were initiated when tumor reached about 100-150 mm3. FIG. 6A: Tumor volume was measured using a caliper as described in Example 2. FIG. 6B: Mouse body weight was also monitored for the different groups and remained within the range established for the endpoint limits.

[0102] FIGs. 7A-B show that the uptake of the TH2307 conjugate is sortilin-dependent in HT-29 cancer cells lines. Uptake of TH2307 was measured in HT-29 cancer cells transfected with either control scrambled siRNA (siScr) or siRNA against sortlin (siSortl). Following cells staining with anti-TH19P01 primary antibody and Alexa-Fluor488-conjugated secondary antibody, the fluorescence signal was acquired by confocal microscopy as described in the Materials and Methods section. FIG. 7A: Obtained images showed that the uptake of TH2307 was reduced in cells transfected with siSortl when compared to the uptake measured in cells transfected with siScr. FIG. 7B: Quantification of the fluorescence signal level indicated that the uptake of TH2307 in cells transfected with siSortl was drastically reduced.

[0103] FIG. 8 depicts the colocalization of anti-TH19P01 and anti-DM1 Abs in TH2307 treated HT- 29 cells. HT-29 cancer cells were treated with TH2307 then incubated with anti-TH19P01 and anti-DM1 antibodies. Confocal microscopy was used to acquired fluorescence signals as described in the Materials and Methods section. Obtained images showed a clear and strong colocalization between anti-TH19P01 and anti-DM1 antibodies following 15 minutes uptake time.

[0104] FIG. 9 shows that DM1 is released from TH2307 in vitro. Western blots were performed with DM1 , TH2307 and TH19P01 samples prepared in non-reducing condition (left side), reducing conditions without boiling (middle side), or in reducing conditions with boiling (right side).

[0105] FIGs. 10A-C depict TH2307 accumulation in HT-29 treated cells. HT-29 cells were incubated for different times (from 15 minutes up to 72 hours) with TH2307 conjugate (3 pM) in serum-free medium at 37°C, washed before cell lysis and Western blots were performed on the cell lysates using anti-TH19P01 and anti-DM1 antibodies. As standards (loading controls), DM1 and TH2307 were loaded on the same gels. FIG. 10A: Detection with anti-TH19P01 antibody. FIG. 10B: Detection with anti-DM1 antibody in non-reducing conditions. FIG. 10C: Detection with anti-DM1 antibody in reducing conditions.

[0106] FIGs. 11A-B show the synergistic anti-cancer activity when combining TH2307 with anti- VEGF antibody on HCT-116 CRC xenograft model. HCT-116 cells were subcutaneously implanted in the flanks of mice. The animals were treated weekly with either vehicle, DM1 and TH2307, or biweekly with anti-VEGF as single agents or in combination with TH2307 at the indicated doses. Treatments were initiated when tumor reached about 100-150 mm3. FIG. 11A: Tumor volume was measured using a caliper as described in Example 2. FIG. 11B: Mouse body weight was also monitored for the different groups and remained within the range established for the endpoint limits.

[0107] FIGs. 12A-B show the synergistic anti-cancer activity when combining TH2307 with anti- VEGF antibody in the CRC PDX model. CRC PDX tumors were subcutaneously implanted in the flanks of mice. The animals were treated weekly with either vehicle, DM 1 and TH2307 or biweekly with anti-VEGF as single agents or in combination with TH2307 and anti-VEGF at the indicated doses. Treatments were initiated when tumor reached about 100-200 mm3. FIG. 12A: Tumor volume was measured using a caliper as described in Example 1. FIG. 12B: Mouse body weight was also monitored for the different groups and remained within the range established for the endpoint limits.

[0108] FIG. 13 shows the heavy and light chain amino acid sequences of the anti-VEGF antibody bevacizumab (Avastin®; KEGG drug entry D0649).

[0109] DETAILED DISCLOSURE

[0110] The use of the terms "a" and "an" and "the" and similar referents in the context of describing the technology (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.

[0111] 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.

[0112] All methods described herein can be performed in any suitable order unless otherwise indicated herein or otherwise clearly contradicted by context.

[0113] The use of any and all examples, or exemplary language (“e.g.”, "such as") provided herein, is intended merely to better illustrate embodiments of the claimed technology and does not pose a limitation on the scope unless otherwise claimed.

[0114] No language in the specification should be construed as indicating any non-claimed element as essential to the practice of embodiments of the claimed technology.

[0115] 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).

[0116] 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.

[0117] Where features or aspects of the disclosure are described in terms of Markush groups or list of alternatives, those skilled in the art will recognize that the disclosure is also thereby described in terms of any individual member, or subgroup of members, of the Markush group or list of alternatives.

[0118] Unless specifically defined otherwise, all technical and scientific terms used herein shall be taken to have the same meaning as commonly understood by one of ordinary skill in the art (e.g., in stem cell biology, cell culture, molecular genetics, immunology, immunohistochemistry, protein chemistry, and biochemistry).

[0119] Unless otherwise indicated, the recombinant protein, cell culture, and immunological techniques utilized in the present disclosure are standard procedures, well known to those skilled in the art. Such techniques are described and explained throughout the literature in sources such as, J. Perbal, A Practical Guide to Molecular Cloning, John Wiley and Sons (1984), J. Sambrook et al., Molecular Cloning: A Laboratory Manual, Cold Spring Harbour Laboratory Press (1989), T. A. Brown (editor), Essential Molecular Biology: A Practical Approach, Volumes 1 and 2, IRL Press (1991), D. M. Glover and B. D. Hames (editors), DNA Cloning: A Practical Approach, Volumes 1- 4, IRL Press (1995 and 1996), and F. M. Ausubel et al. (editors), Current Protocols in Molecular Biology, Greene Pub. Associates and Wiley-lnterscience (1988, including all updates until present), Ed Harlow and David Lane (editors) Antibodies: A Laboratory Manual, Cold Spring Harbour Laboratory, (1988), and J. E. Coligan et al. (editors) Current Protocols in Immunology, John Wiley & Sons (including all updates until present).

[0120] The present disclosure provides a conjugate comprising the following structure I, or a pharmaceutically acceptable salt thereof:

[0121] A — L — B (I); wherein

[0122] A is a maytansinoid;

[0123] B is a peptide binding to a Sortilin receptor, wherein the peptide comprises an amino acid sequence having at least 60% identity with the amino acid sequence set forth in SEQ ID NO: 1 or 2:

[0124] X1GVRAKAGVRN(Nle)FKSESYX2(SEQ ID NO:1)

[0125] X1YKSLRRKAPRWDAPLRDPALRQLLX2(SEQ ID NO:2); wherein X1is cysteine (C) or is absent, and X2is cysteine (C) or is absent;

[0126] L is a linker, wherein a first end of the linker is attached to a side chain of one or more of the amino acid or modified side chain residues of the peptide (such as, azido, double bond, triple bond and others), and / or to the N- and / or C-terminal end of the peptide, and a second end of the linker is attached to a hydroxy, carboxy or nitrogen atom of the maytansinoid.

[0127] 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 ( / .e., UniProtKB Accession number Q99523).

[0128] 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.

[0129] 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 PAM120 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.

[0130] In embodiments, the peptide comprises or consists of an amino acid sequence having 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 72%, at least 73%, at least 74%, at least 75%, at least 76%, at least 77%, at least 78%, at least 79%, at least 80%, at least 81%, at least 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% sequence identity to the sequence set forth in SEQ ID NO: 1 , wherein the peptide binds to Sortilin.

[0131] In an embodiment, the peptide comprises 25, 24, 23, 22, 21 , 20, or 19 residues or less and comprises the sequence GVRAKAGVRN(Nle)FKSESY (SEQ ID NO:3). In another embodiment, the peptide compound comprises 25, 24, 23, 22, 21 , 20, or 19 residues or less and comprises the sequence GVRAKAGVRN(Nle)FKSESYC (SEQ ID NO:4).

[0132] In embodiments, the peptide has 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

[0133] 68%, at least 69%, at least 70%, at least 71%, at least 72%, at least 73%, at least 74%, at least

[0134] 75%, at least 76%, at least 77%, at least 78%, at least 79%, at least 80%, at least 81%, at least

[0135] 82%, at least 83%, at least 84%, at least 85%, at least 86%, at least 87%, at least 88%, at least

[0136] 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least

[0137] 96%, at least 97%, at least 98% or at least 99% sequence identity to the sequence set forth in SEQ ID NO: 2, wherein the peptide binds to Sortilin.

[0138] In an embodiment, the peptide comprises 30, 29, 28, 27, or 26 residues or less and comprises the sequence YKSLRRKAPRWDAPLRDPALRQLL (SEQ ID NO:5). In another embodiment, the peptide compound comprises 30, 29, 28, 27, or 26 residues or less and comprises the sequence YKSLRRKAPRWDAPLRDPALRQLLC (SEQ ID NO:6).

[0139] The peptide may comprise one or more covalent modifications. Covalent modifications include reacting targeted amino acid residues of the peptides with an organic derivatizing agent that is capable of reacting with selected side chains or the N- or C- terminal residues of the peptides. 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 peptides 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 peptide.

[0140] In an embodiment, the peptide comprises one or more modifications at the N- and / or C- terminal end, for example to protect the peptide from proteolytic degradation. In an embodiment, the amino-terminal modifying group is a C1-C16 or C3-C16 acyl group (linear or branched, saturated or unsaturated), in a further embodiment, a saturated Ci-C6acyl group (linear or branched) or an unsaturated C3-C6acyl group (linear or branched). In a further embodiment, the amino-terminal modifying group is an acetyl group (CH3-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 ortertiary 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.

[0141] In an embodiment, the peptide comprises or consists of the sequence *GVRAKAGVRN(Nle)FKSESY, wherein * is an acetyl group (SEQ ID NO:7). In an embodiment, the peptide comprises or consists of the sequence *GVRAKAGVRN(Nle)FKSESYC, wherein * is an acetyl group (SEQ ID NO:8). In an embodiment, the peptide comprises or consists of the sequence of ‘YKSLRRKAPRWDAPLRDPALRQLL, wherein * is an acetyl group (SEQ ID NO:9). In an embodiment, the peptide comprises or consists of the sequence of ‘YKSLRRKAPRWDAPLRDPALRQLLC, wherein * is an acetyl group (SEQ ID NO: 10).

[0142] The term “maytansinoid” as used herein refers to compounds having a structure similar to that of maytansine (notably containing a 19-membered macrocyclic lactam ring backbone) and having the ability to inhibit microtubule assembly. Examples of maytansinoids include DM1 (N2- deacetyl-N2-(3-mercapto-1-oxopropyl)-maytansine, mertansine), DM3, and DM4 (N2-deacetyl- N2-(4-mercapto-4-methyl-1 -oxopentyl) maytansine, ravtansine).

[0143] Ri = CH2CH2SH (DM1)

[0144] R1 = CH2CH2CH(CH3)SH (DM3)

[0145] R1 = CH2CH2C(CH3)2SH (DM4)

[0146] In an embodiment, the maytansinoid is DM1. In another embodiment, the maytansinoid is DM4.

[0147] The term “linker” as used herein means a chemical structure connecting a peptide compound herein disclosed to the maytansinoid. The linker can be connected to the peptide compound at different functional groups on the peptide compounds. 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. The linker may also be attached to suitable groups / moieties on modified side chain residues of the peptide (such as azido, double bond, and triple bond).

[0148] In an embodiment, L is a cleavable linker, wherein a first end of the linker is attached to a side chain of one or more of the lysine (K) and / or cysteine (C) residues of the peptide, and / or to the N- and / or C-terminal end of the peptide, and a second end of the linker is attached to an oxygen, nitrogen, or sulfur atom of the maytansinoid.

[0149] Through the use of cleavable linkers, the maytansinoid can be cleaved from the peptide at the surface of a Sort il i n receptor-expressing cell, inside the cell, or inside a cell organelle, e.g. , by an enzyme (e.g., a protease), thereby releasing the maytansinoid, e.g., in the vicinity of the cell or inside the cell or cell organelle). The linkers described herein are used to attach the maytansinoid to the peptides that binds to a cancer cell (e.g., binds to cancer cells expressing a Sortilin receptor) and gets internalized into the cell (e.g., through the endosome and / or lysosomal compartment(s)). Once internalized, the linkers are cleaved or degraded to release the maytansinoid.

[0150] Various types of cleavable linkers may be used in the conjugates described herein. For example, the linker may be an acid-cleavable linker that exploits the natural acidity of endosomes and lysosomes (pH 4.5-6.2) which contrasts with the neutral pH (7.4) of the plasma. The most commonly used acid-cleavable linkers are hydrazones and carbonates. The cleavable linker may also be a reducible or disulfide linker. Such linkers are stable at physiological pH but susceptible to nucleophilic attacks from thiols. Glutathione (GSH) that contains exposed and highly reactive thiol groups is present in the cytosol of the cells, especially in cells under oxidative stress such as tumor cells. Other types of cleavable linkers include phosphatase cleavable linkers (cleaved by phosphatase and pyrophosphates in lysosomes) and sulfatase cleavable linkers (cleaved by sulfatases in lysosomes). The cleavable linker may also be an enzyme-cleavable linker, i.e., a linker that is cleaved by the action of intracellular and / or extracellular enzymes expressed by the target cells. Examples of enzyme-cleavable linkers include glycosidase-cleavable linkers (e.g., |3- glucuronidase or p-galactosidase cleavable linker) as well as protease cleavable linkers.

[0151] In an embodiment, the cleavable linker is a protease-cleavable linker. The protease- cleavable linker can release the maytansinoid under the action of proteases such as cathepsin (e.g., cathepsin A. cathepsin B), trypsin or other proteases expressed by the cell (e.g., tripeptidyl- peptidase I), for example expressed in the endosomal and / or lysosomal compartment(s) of the cell.

[0152] In an embodiment, the cleavable linker comprises one or more amino acid residues, e.g., natural and / or or non-natural amino acid residues. In an embodiment, the cleavable linker comprises from 2 to 4 amino acid residues. In an embodiment, the cleavable linker comprises one of the following dipeptides: Val-Lys, Val-Cit, Phe-Lys, Trp-Lys, Asp-Lys, Val-Arg, or Val-Ala. In other embodiments, the cleavable linker comprises one of the following tetrapeptides: Val-Phe- Gly-Sar, Val-Cit-Gly-Sar, Val-Lys-Gly-Sar, Val-Ala-Gly-Sar, Val-Phe-Gly-Pro (SEQ ID NO:11), Val-Cit-Gly-Pro, Val-Lys-Gly-Pro (SEQ ID NO: 12), or Val-Ala-Gly-Pro (SEQ ID NO: 13).

[0153] The linkers may be prepared from linker precursors that contain reactive groups at one or both ends of the molecule. The reactive groups can be selected to allow conjugation to a maytansinoid at one end, and also facilitate conjugation to the peptide at the other end. It is desirable for the maytansinoid to contain an amine, a hydroxyl, hydrazone, hydrazide or a sulfhydryl group in order to facilitate conjugation to the linker. In an embodiment, the linker is conjugated to a sulfhydryl group of the maytansinoid. In an embodiment, the linker is of the following structure: S-X3-CO, wherein X3is a Ci-C8alkyl (linear or branched), for example a Ci-C6alkyl or a C2-C4alkyl (linear or branched). In a further embodiment, the linker is of the following structure: S-CH(CH3)-CH2-CH2-CO. In another embodiment, the linker is of the following structure: S-CH2-CH2-CO.

[0154] The linker may further comprise additional groups, e.g., spacer groups, to increase the distance between the peptide and the maytansinoid. Suitable spacer groups may comprise an alkylene, alkenylene, alkynylene, heteroalkylene (e.g., polyethylene glycol, PEG), carbocyclyl, heterocyclyl, aryl, heteroaryl, or a combination thereof. For example, the spacer group may comprise a PEG group, an alkylene group, or a combination thereof. The spacer group may be substituted or unsubstituted, e.g., the spacer group may comprise a substituted alkylene, substituted heteroalkylene, or a combination thereof. For example, the spacer group may comprise a PEG group (or PEG spacer), an alkylene group (or alkylene spacer), one or more heteroatoms, and / or one or more or cyclic groups. In an embodiment, the spacer group comprises 1 to 20 PEG groups. In further embodiments, the spacer group comprises 1 to 10, 1 to 8, 1 to 6, 1 to 5, or 2 to 4 PEG groups. In a further embodiment, the spacer group comprises 3 PEG groups.

[0155] In an embodiment, the linker is of one of the following structures:

[0156] • CO(CH2)aCONH(CH2)bCO, wherein a and b are independently 1 , 2, 3, or 4;

[0157] • CO(CH2)d(OCH2CH2)eCONH(CH2)fCO, wherein d and f are independently 1 , 2, 3, or 4, and e is an integer from 2 to 8; or

[0158] It is to be understood that several maytansinoid molecules A (which may be the same or different) may be attached to a single peptide molecule B through several linkers L (which may be the same or different). In an embodiment, 1 , 2, 3 or 4 maytansinoid molecules are attached to a single peptide molecule through linkers.

[0159] Thus, in an embodiment, the conjugate or salt thereof is of one of the following formulas la, lb, Ic or Id:

[0160] A1 — L1 — B (la);

[0161] A1 — L1 — B — L2 — A2 (lb); wherein L1 , L2, L3 and L4 are linkers (same or different), and A1 , A2, A3 and A4 are maytansinoid molecules (same or different). In an embodiment, A1 , A2, A3 and A4 are the same maytansinoid molecules. In an embodiment, L1 , L2, L3 and L4 are the same cleavable linkers.

[0162] In an embodiment, X1and / or X2in SEQ ID NO:1 or 2 is C, and wherein the first end of one of the linkers is attached to the side chain of X1and / or X2. In a further embodiment, X2is C, and wherein the first end of one of the linkers is attached to the side chain of X2.

[0163] In an embodiment, the first end of one or more of the linkers is attached to the side chain of one or more lysine (K) residues of the peptide B. In a further embodiment, the first ends of two of the linkers are attached to the side chain of two lysine (K) residues of the peptide B.

[0164] In an embodiment, the conjugate has one of the following structures:

[0165]

[0166] 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.

[0167] 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 agents, 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.

[0168] In another aspect, the present disclosure provides a method for treating a Sortilin- expressing cancer in a subject in need thereof comprising administering to the subject an effective amount of the conjugate compound, salt thereof or composition disclosed herein. The present disclosure also provides the use of the conjugate compound, salt thereof or composition disclosed herein for treating a Sortilin-expressing cancer, or for the manufacture of a medicament for treating a Sortilin-expressing cancer, in a subject. The present disclosure also provides the conjugate compound, salt thereof or composition disclosed herein for use in treating a Sortilin- expressing cancer in a subject.

[0169] The Sortilin-expressing cancer may be any type of cancer, including a primary (or original) cancer, a relapsing cancer or a metastatic cancer, in which at least a subset of the tumor cells expresses Sortilin. 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. In an embodiment, the Sortilin-expressing cancer is a hematological cancer (e.g., AML), ovarian cancer, endometrial cancer, cervix cancer, skin cancer (e.g., melanoma, squamous cell skin cancer), brain cancer (e.g., glioma, glioblastoma), breast cancer (e.g., triplenegative breast cancer), colorectal cancer, small intestine cancer, liver cancer, lung cancer (e.g., small cell lung carcinoma, non small cell lung carcinoma), eye cancer (e.g., ocular melanoma), prostate cancer, head and neck cancer, stomach cancer, bone cancer, thyroid cancer, testis cancer, bladder cancer, kidney cancer, or pancreatic cancer. In a further embodiment, the cancer is colon cancer, breast cancer, ovarian cancer or pancreatic cancer.

[0170] In an embodiment, the cancer is a poor prognosis cancer. The term “poor prognosis cancer” as used herein refers to a subtype of a given cancer that is associated with lower survival rate (e.g., 5-year or 10-year survival rate) relative to other subtype(s) of the same cancer. Poor prognosis cancer is generally associated with specific characteristics of the cancer subtype, for example the presence of certain mutations, chromosomal abnormalities, etc., that renders them more resistant to treatment. Poor prognosis is also associated with cancers diagnosed at a later stage (e.g., with distant metastasis). For example, for breast cancer, triple-negative breast cancer (TNBC) is considered a poor prognosis breast cancer as it is associated with a lower 5-year relative survival rate relative to other breast cancer subtypes. For ovarian cancer, invasive epithelial ovarian cancer and fallopian tube cancer are generally associated with a lower 5-year relative survival rate relative to ovarian stromal tumors and germ cell tumors. The 5-year overall survival rate of pancreatic cancer is very low (about 3%), which is partly because more than half of the patients are diagnosed at an advanced stage. Diagnosis of pancreatic cancer at stage II l / IV (with distant metastasis) is associated with very poor prognosis. Similarly, for prostate cancer, diagnosis at stage IV (with distant metastasis) is associated with poor prognosis (5-year relative survival rate of less than 30% compared to at least 80-85% for diagnosis at stages l-lll). For lung cancer, small cell lung cancer is associated with particularly poor prognosis, especially when diagnosed at a later stage (e.g., with regional or distant metastasis). Non-small cell lung cancer diagnosed at a later stage (e.g., with distant metastasis) is also associated with poor prognosis. In colorectal cancer, mucinous adenocarcinomas (characterized by the presence of abundant extracellular mucin) have been associated with reduced response to chemotherapy and poor prognosis. Peritoneal involvement and BRAF mutations also constitute poor prognosis markers for colorectal cancer. For kidney cancer, clear cell RCC is associated with worse outcomes (e.g., lower 5-year relative survival rate) than papillary RCC. In skin cancer, thicker tumors, nodal involvement and diagnosis at a later stage (e.g., with regional or distant metastasis) are associated with lower survival in melanoma.

[0171] In an embodiment, the poor prognosis cancer is a stage lll / IV cancer. In an embodiment, the poor prognosis cancer is a stage III cancer. In an embodiment, the poor prognosis cancer is a stage IV cancer.

[0172] In an embodiment, the poor prognosis cancer is a cancer having a 5-year relative survival rate of less than 60%. In an embodiment, the poor prognosis cancer is a cancer having a 5-year relative survival rate of less than 50%. In an embodiment, the poor prognosis cancer is a cancer having a 5-year relative survival rate of less than 40%. In an embodiment, the poor prognosis cancer is a cancer having a 5-year relative survival rate of less than 30%. In an embodiment, the poor prognosis cancer is a cancer having a 5-year relative survival rate of less than 20%. In an embodiment, the poor prognosis cancer is a cancer having a 5-year relative survival rate of less than 10%. In an embodiment, the poor prognosis cancer is a cancer having a 5-year relative survival rate of less than 5%.

[0173] In an embodiment, the cancer is an immunologically cold (or ignorant) cancer. The term “immunologically cold cancer” or “cold cancer” refers to a cancer that 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). 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.

[0174] 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.

[0175] 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), VEGF / VEGFR (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).

[0176] The combination of active agents and / or compositions comprising same may be administered or co-administered (e.g., consecutively, simultaneously, at different times) in any conventional dosage form. Co-administration 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.

[0177] 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).

[0178] 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.

[0179] 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.

[0180] 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.

[0181] In an embodiment, the cancer is resistant to monotherapy with EGFR inhibitors or VEGF inhibitors. Resistance to EGFR inhibitors may be caused by different mechanisms including mutations in EGFR, mutations in Ras proteins or Ras pathway proteins, activation of alternative signaling pathways, the aberrance of downstream pathways, and impairment of the EGFR- mediated apoptosis pathway. Resistance to VEGF inhibitors may be caused by different mechanisms including increase in HIF-1a expression, increased local invasiveness and distant metastasis, redundancy in angiogenic signaling pathways, mutations in Ras proteins or Ras pathway proteins, increase in Ang-2 expression, increase in bombina variegate peptide 8 (Bv8) expression, increase in FGF expression, increase in PDGF expression, increase in TGF-p expression, and increase in expression of matrix metalloproteinases (MMPs).

[0182] In an embodiment, the cancer harbors one or more mutations in a Ras protein or Ras pathway protein. In an embodiment, the cancer harbors one or more mutations in a Ras protein. In an embodiment, the mutation is in K-Ras, in a further embodiment the mutation is at residues 12, 13 and / or 61 of K-Ras. In another embodiment, the mutation is in N-Ras, in a further embodiment the mutation is at residues 12, 13 and / or 61 of N-Ras. In another embodiment, the mutation is in H-Ras, in a further embodiment the mutation is at residues 12, 13 and / or 61 of H- Ras. In an embodiment, the mutation at residue 12 of K-Ras, N-Ras and / or H-Ras is G12A, G12C, G12D, G12R, G12S, or G12V. In an embodiment, the mutation at residue 13 of K-Ras, N-Ras and / or H-Ras is G13A, G13C, G13D, G13R, G13S, or G13V. In an embodiment, the mutation at residue 61 of K-Ras, N-Ras and / or H-Ras is Q61 E, Q61 H, Q61 K, Q61 L, Q61 P or Q61 R. In an embodiment, the mutation is at residue 12 of K-Ras, more particularly G12D.

[0183] In an embodiment, the cancer harbors one or more mutations in a Ras pathway protein. In a further embodiment, the Ras pathway protein is BRAF. In a further embodiment, the mutation is at residue 464 (for example G464V, G464E), 466 (for example G466V, G466E, G466A), 467 (for example S467L), 469 (for example G469A, G469V, G469R, G469E), 581 (for example N581S), 594 (for example D594A, D594E, D594G, D594H, or D594N), 595 (for example F595L), 596 (for example G596R, G596D), 597 (for example L597Q, L597V), 600 (for example V600E, V600M, V600K, V600R, or 600D), and / or 601 (for example K601 E, K601 N, or K601T), of BRAF. In a further embodiment, the mutation is at residue 600 of BRAF, more particularly V600E.

[0184] 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 a cancer resistant to monotherapy with EGFR inhibitors or EGF inhibitors.

[0185] 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 a cancer harboring one or more of the above-noted mutations in a Ras protein or Ras pathway protein.

[0186] Mutations in BRAF have been reported in several cancers include colorectal cancers, melanoma, multiple myeloma, non-small cell lung cancers, bladder urothelial cancers, chronic lymphocytic leukemia, brain glioblastoma multiforme, head and neck squamous cell cancers, kidney cancers, liver cancers, thyroid cancers, prostate cancers and gastric cancers.

[0187] Mutations in Ras proteins or Ras pathway proteins have been reported in several cancers including pancreatic ductal adenocarcinoma, colorectal carcinoma, non-small cell lung carcinoma, melanoma, urinary bladder carcinoma, thyroid carcinoma, head and neck squamous cell cancers, brain glioblastoma multiforme, lower grade glioma, breast cancers, ovarian cancers, bladder urothelial cancers, as well as hematopoietic malignancies.

[0188] In an embodiment, the cancer harboring one or more of the above-noted mutations in a Ras protein or Ras pathway protein is a gastrointestinal cancer, such as colorectal cancer. In an embodiment, the cancer is colon cancer. In another embodiment, the cancer is rectal cancer. In an embodiment, the colorectal cancer is characterized by microsatellite instability (MSI). In an embodiment, the colorectal cancer is characterized by microsatellite stability (MSS). In an embodiment, the colorectal cancer is characterized by RAS (e.g., KRAS) and / or RAF mutations. In another embodiment, the colorectal cancer is resistant / refractory to chemotherapy. In another embodiment, the colorectal cancer is resistant / refractory to EGFR inhibitors.

[0189] 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.

[0190] EXAMPLES

[0191] The present disclosure is illustrated in further details by the following non-limiting examples.

[0192] Example 1: Synthesis of peptide-drug (Maytansinoids) conjugates

[0193] General Analytical Methods and Methodology for purification and Characterization of

[0194] As used herein the term RT (min) refers to the LCMS retention time, in minutes, associated with the compound. Unless otherwise indicated, the instruments and columns used to obtain the desired compounds. Instrument and column: Waters Acquity UPLC BEH C18 1.7 mm, 2.1 x 50 mm. Agilent 1290 Infinity II LUNA C185 mm, 4.6 x 150 mm. AKTA pure protein purification system using 30 RPC resins.

[0195] The following abbreviations may be used as follows:

[0196] Aq. = aqueous cone = concentrate

[0197] DCM = Dichloromethane

[0198] DIPEA = Diisopropylethylamine

[0199] DMF = dimethylformamide

[0200] DMSO = Dimethylsulfoxide

[0201] EtOAc = Ethyl acetate

[0202] Mol = molar

[0203] MeOH = Methanol

[0204] RT = room temperature

[0205] TEA = Triethylamine

[0206] THF = Tetra hydrofuran

[0207] Synthesis of TH19P01-Lys5 13-SPDP-DM1 (TH2307) (FIG. 1A)

[0208] Step 1 :

[0209] To a solution of DM1 (1) (250 mg, 0.339 mmol) in DCM (10 ml), 3-(2-Pyridyldithio)propionic acid N-hydroxysuccinimide ester (SPDP) (159 mg, 0.508 mmol) and N,N-Diisopropylethylamine (88.5 pL, 0.508 mmol) were added. The reaction mixture was stirred under nitrogen atmosphere at room temperature for 3-4 hours and monitored by UPLC. Once starting material was consumed, the solution was concentrated and purified by column chromatography using EtOAc in DCM as a gradient and pure fractions were evaporated to afford (14S,16S,32S,33S,2R,4S,10E,12E,14R)- 86-chloro- 14- hydroxy-85, 14-dimethoxy-33,2,7, 10-tetram ethyl- 12,6-dioxo-7-aza-1 (6,4)- oxazinana-3(2,3)-oxirana-8(1 ,3)-benzenacyclotetradecaphane-10,12-dien-4-yl N-(3-((3-((2,5- dioxopyrrolidin-1-yl)oxy)-3-oxopropyl)disulfaneyl)propanoyl)-N-methyl-L-alaninate (2) (275 mg, 86%) as off white solid. The LC-MS exhibited more than 99% purity of product. The product obtained was used as such in the next step.

[0210] Step 2:

[0211] To a solution of (14S,16S,32S,33S,2R,4S,10E,12E,14R)-86-chloro-14-hydroxy-85,14- dimethoxy-33,2,7,10-tetramethyl-12,6-dioxo-7-aza-1 (6,4)-oxazinana-3(2,3)-oxirana-8(1 ,3)- benzenacyclotetradecaphane-10,12-dien-4-yl N-(3-((3-((2,5-dioxopyrrolidin-1-yl)oxy)-3- oxopropyl)disulfaneyl)propanoyl)-N-methyl-L-alaninate (2) (271 mg, 0.288 mmol) in DMSO (3.29 ml), TH19P01 (250 mg, 0.130 mmol) in DMSO (1.1 ml_) and DI PEA (0.865 mmol, 151 pL) were added. The reaction was stirred at room temperature and monitored with UPLC. After the completion, the reaction mixture was injected directly to the AKTA and purified using 10-80% ACN (0.1% FA) in H2O (0.1%FA). The pure fractions were pooled and freeze dried to afford TH2307 (374 mg, 81%) as white solid. MH+3 1191.76 is observed at 1.75 minutes with > 99% purity.

[0212] Synthesis of TH19P01-Lys5 13-SPDP-DM4 (TH2311) (FIG. 1B)

[0213] Step 1 :

[0214] To a solution of DM4 (1) (260 mg, 0.339 mmol) in DCM, 3-(2-Pyridyldithio)propionic acid N- hydroxysuccinimide ester (117 mg, 0.373 mmol) and N,N-Diisopropylethylamine; DIEA; Hunig's base (65.0 pL, 0.373 mmol) were added. Reaction was allowed to stir for 3-4 hours and once all the starting material was consumed, DCM was evaporated by blowing nitrogen gas to afford (14S, 16S,32S,33S,2R,4S, 10E, 12E, 14R)-86-chloro- 14-hydroxy-85, 14-dimethoxy-33,2,7, 10- tetramethyl-12,6-dioxo-7-aza-1 (6,4)-oxazinana-3(2,3)-oxirana-8(1 ,3)- benzenacyclotetradecaphane-10,12-dien-4-yl N-(3-((3-((2,5-dioxopyrrolidin-1-yl)oxy)-3- oxopropyl)disulfaneyl)-3-methylbutanoyl)-N-methyl-L-alaninate (2). The crude was used as such in the next step. Step 2:

[0215] 2,5-dioxopyrrolidin- 1 -yl 4-((4-(((S)-1 -(((14S, 16S,32S,33S,2R,4S, 10E, 12E, 14R)-86-chloro- 14-hydroxy-85,14-dimethoxy-33,2,7,10-tetramethyl-12,6-dioxo-7-aza-1 (6,4)-oxazinana-3(2,3)- oxirana-8(1 ,3)-benzenacyclotetradecaphane-10,12-dien-4-yl)oxy)-1-oxopropan-2- yl)(methyl)amino)-2-methyl-4-oxobutan-2-yl)disulfaneyl)-4-oxobutanoate (2) (147 mg, 0.119 mmol) was taken in DMSO (1.80 mL) and to that, Hunig’s base (62.1 pL, 0.356 mmol) and TH19P01 (103 mg, 0.0534 mmol) were added. The reaction was monitored by LCMS. Once all the starting material was consumed and there was no more progress of the reaction, it was injected directly to the AKTA and purified using acetonitrile in water (0-40%, 0.1 % formic acid) as a gradient. The pure fractions were collected and lyophilized to afford TH2311 (85.0 mg, 44 %). MH+3 (1219.6) is observed at 1.89 minutes on 4 min run with > 97% purity on UPLC.

[0216] Synthesis of TH19P01- Lys5 13-SMCC-DM1 (TH2401) (FIG. 1C)

[0217] To a solution of DM1 (1) (25.0 mg, 0.0339 mmol) in AON (1.00 ml) and phosphate buffer (2.00 ml), 4-(N-Maleimidomethyl)cyclohexanecarboxylic acid N-hydroxysuccinimide ester, SMCC (11 .3 mg, 0.0339 mmol) was added. The reaction mixture was stirred at room temperature for 30 minutes and monitored with the help of UPLC. Once starting material was consumed, to the reaction mixture, TH19P01 (29.3 mg, 0.0152 mmol) was added. To the solution, 10 pL of the triethyl amine was added and product formation was observed. The reaction mixture was quenched by addition of water and filtered. The filter cake was dried. Because of solubility problem, the solid obtained was dissolved in 200 ml of 40% mixture of acetonitrile:water and diluted to 10% of acetonitrile in water. The solution was pumped directly to the AKTA and purified using acetonitrile in water (0-40%, 0.1% formic acid) and pure fractions were lyophilized to afford TH2401. MH+2 (1920.42) is observed on QTOF.

[0218] Synthesis of TH19P01-Lys5 13-SPP-DM1 (TH2402) (FIG. 1D)

[0219] Step 1 :

[0220] To a solution of DM1 (1) (250 mg, 0.339 mmol) in DCM (6.25 mL), 3-(2- Pyridyldithio)propionic acid N-hydroxysuccinimide ester (173 mg, 0.508 mmol), N,N- Diisopropylethylamine; DIEA; Hunig's base (93.6 pL, 0.508 mmol) were added. The reaction mixture was stirred at room temperature for 3h under nitrogen atmosphere. LC-MS showed the completion. The solution was concentrated and purified by normal phase column chromatography using EtOAc in DCM as gradient and pure fractions were evaporated to afford 2,5-dioxopyrrolidin-

[0221] 1 -yl 4-((3-(((S)-1 -(((14S, 16S,32S,33S,2R,4S, 10E, 12E, 14R)-86-chloro-14-hydroxy-85, 14- dimethoxy-33,2,7,10-tetramethyl-12,6-dioxo-7-aza-1 (6,4)-oxazinana-3(2,3)-oxirana-8(1 ,3)- benzenacyclotetradecaphane-10,12-dien-4-yl)oxy)-1-oxopropan-2-yl)(methyl)amino)-3- oxopropyl)disulfaneyl)pentanoate (2) (98.0 mg, 29%) as off white solid.

[0222] Step 2:

[0223] 2,5-dioxopyrrolidin- 1 -yl 4-((3-(((S)-1 -(((14S, 16S,32S,33S,2R,4S, 10E, 12E, 14R)-86-chloro- 14- hydroxy-85, 14-dimethoxy-33,2,7, 10-tetramethyl-12,6-dioxo-7-aza- 1 (6,4)-oxazinana-3(2,3)- oxirana-8(1 ,3)-benzenacyclotetradecaphane-10,12-dien-4-yl)oxy)-1-oxopropan-2- yl)(methyl)amino)-3-oxopropyl)disulfaneyl)pentanoate (98.0 mg, 0.0993 mmol) was taken in DMSO (2.00 mL) followed by addition of TH19P01 (85.9 mg, 0.0447 mmol) dissolved in 1 ml DMSO and Hunig’s base (38.5 mg, 0.298 mmol). The reaction was monitored with UPLC for completion. Once most of the NH-esterwas consumed, reaction mixture was injected to the AKTA using acetonitrile:water (0-80%, containing 0.1 formic acid) as eluent. Pure fractions were pooled together and freeze dried to afford TH2402 (120 mg, 74%) as colorless solid. MH+3 (1210.52) is observed at 1.82 minutes on 4 min run with > 97% purity on UPLC.

[0224] Example 2: Materials and Methods

[0225] Cell lines and cell culture

[0226] Human colorectal cancer cell lines HCT-116 (#CCL-247), LoVo (# CCL-229) and HT-29 (# HTB-38) were purchased from ATCC. Human triple-negative breast cancer (TNBC)-derived MDA-MB-231 / Luc cells was from Cell Biolabs Inc. (San Diego, CA, #AKR-231). Human ovarian cancer SKOV3 (#AKR-232) was purchased from Cell Biolabs Inc. (San Diego, CA) whereas ES-

[0227] 2 (#CRL-1978) cell line was from ATCC. Human pancreatic cancer PANC-1 (#CRL-1469) cell line was from ATCC.

[0228] All cell lines were grown as adherent monolayers at 37°C in a humidified atmosphere (5% CO2) using media described in Table 1.

[0229] Table 1 : Cell Culture Conditions Used for Cell Lines

[0230] For experimental use, cells were detached from the culture flask by a 5-10-minute treatment with trypsin (Wisent), then were 10-fold diluted and neutralized by addition of complete culture media. Cell counts and cell viability were assessed with a TC20 automated cell counter (BioRad) following 0.4% trypan blue (Thermo Fisher Scientific) exclusion staining of cells.

[0231] Western blotting

[0232] For standards preparation, 3 pM of DM1 , TH2307, and TH19P01 were prepared in 2Xtricine sample buffer (BioRad, # 1610739) supplemented or not with 10% p-mercaptoethanol and incubated or not for 5 minutes at 95 °C.

[0233] For TH2307 detection in HT-29 treated cells, 1% SDS supplemented with a complete protease inhibitor cocktail from Calbiochem (San Diego, CA) was used as a lysis buffer. Cells were incubated for 30 min at RT with vortexing every 5 min, sonicated and centrifuged at 15,000g for 10 min at RT. Samples were mixed with 5X sample buffer and incubated or not for 5 minutes at 95°C.

[0234] Standards or equal amounts of protein (10 pg) were resolved on a 16.5% Tris-tricine gel (Bio-Rad). Proteins were then electrotransferred to a nitrocellulose membrane (0.2 pm) and blocked for 1 h at room temperature 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 TH19P01 peptide (1 / 1 ,000 dilution; Covance), DM1 (1 / 100 dilution; Covalab; # mab 0160-P), GAPDH (1 / 50,000 dilution; EMD Millipore; # CB1001) diluted in TBST containing 3% BSA and 0.05% NaN3. Membranes were washed in TBST and incubated for 1 h at room temperature with horseradish peroxidase- conjugated anti-mouse or anti-rabbit IgG (1 / 5000 dilution) in TBST containing 5% non-fat dry milk. Membranes were washed again in TBST and signals were detected using chemiluminescence (Amersham Biosciences, Baie d’Urfe, QC).

[0235] Sortilin RNA interference

[0236] HT-29 cells were transiently transfected for 24 hours with 100 nM of a siRNA scrambled sequence (AllStar Negative Control siRNA, 1027281) or a human siRNA generated against sortilin (Hs_SORT_5 FlexiTube siRNA: SI03115168; Qiagen, Valencia, CA) using Lipofectamine™ 2000 (ThermoFisher Scientific, Burlington, ON). Medium was refreshed for an additional 24h before starting the uptake assay.

[0237] Uptake Assay of TH2307 by confocal microscopy For SORT1 -dependant internalization assay, HT-29 cells were plated for 24 hours on glass coverslips and grown to 40-50% confluency. Cells were then transfected with scrambled siRNA or siSORTI as described above. Cells were washed with HBSS and incubated in HBSS for 15 minutes at 37°C in the presence or absence of 300 nM TH2307. Cells were washed with HBSS, fixed for 10 minutes at RT in 4% paraformaldehyde (PFA), washed again in HBSS before permeabilization for 10 minutes at RT using 0.1% Triton™X-100 in HBSS. Cells were blocked for 1 hour with 1% BSA-HBSS and incubated O / N at 4°C with anti-TH19P01 primary antibody (1 OO; Covance). Cells were washed with PBS and incubated for 1 hour with rabbit Alexa-Fluor™488- conjugated secondary antibody (1 / 1 ,000; Invitrogen).

[0238] For TH19P01 and DM1 co-localization experiment, HT-29 cells were plated for 24 hours on glass coverslips and grown to 70% confluency. Cells were first treated and stained with anti- TH19P01 primary antibody and Alexa-Fluor™488-conjugated secondary antibody as described above. Cells were then washed with HBSS, incubated O / N at 4°C with anti-DM1 primary antibody (1 :100; Covalabs), washed again with PBS before incubation for 1 hour with mouse Alexa- Fluor488-conjugated secondary antibody (1 / 1 ,000; Invitrogen).

[0239] In all these experiments, cells were washed with PBS, stained with DAPI (2 pg / ml, Invitrogen) for 4 min, washed again and mounted onto slides using Prolong™ Gold antifade reagent. Cells were finally digitalized by confocal microscopy (Nikon A1) and analyzed using the NIH Imaged Version 1.4.21 software.

[0240] Cell proliferation assay

[0241] To assess the effects of free (Lys-MHA-DM1 , S-methyl-DM1 , DM1 and DM4) and conjugated drugs on cell proliferation, ES-2 (2,500 cells / well), MDA-MB-231 / luc (3,000 cells / well), PANC-1 (3,000 cells / well), and HT-29 (3,000 cells / well), LoVo (8,000 cells / well), and HCT-116 (1 ,500 cells / well) were first seeded for 24 hours in 96-well plates in complete growth medium. Cells were then treated in complete cell culture medium with various concentrations of drugs. After 72 hours of incubation, viable cells were estimated by MTT, WST-1 or WST-8 (Sigma- Aldrich) assay. GraphPad Prism™ software was used to calculate IC50 values.

[0242] Animals

[0243] Female (HT-29, SKOV3) or male (HCT-116) athymic nude (Crl:NU(NCr)-Foxn1 nu mice and female CD1 nude (Crl:NU-Foxn1 nu; MDA-MB-231) mice aged between six and eight weeks were obtained from Charles River Laboratories, Inc. (St-Constant, QC). Male NOD scid gamma (NSG, NOD-sc / d IL2Rgnu"; PDX) mice aged between six and eight weeks were obtained from The Jackson Laboratories, Inc. (Bar Harbor, ME). 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.

[0244] Establishing xenograft models for in vivo efficacy assessment of peptide drug conjugates drug combinations

[0245] Tumors were established in mice by subcutaneous inoculation of HT-29 (7x106), HCT-116 (3x106), MDA-MB-231 (7x106) or SKOV3 (7x106) cancer cells in 100 pl of HBSS (Sigma #H6648). All cells were injected into the right flank of immunodeficient mice under light isoflurane anesthesia. Mice were treated via intravenous (IV) tail vein or intraperitoneal (IP) injection at doses indicated in the respective figures and preparations of all injectables are described in the section below. 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 reached approximately 1000 mm3in size for each respective group or following prolonged observation upon end of treatments when tumor growth were halted.

[0246] PDX tumors were established in mice by subcutaneous implantation of 1-2 mm3tumors derived from a metastatic colorectal cancer patient. The PDX model (TM00170) from a patient with grade IV metastatic colorectal cancer to the liver was purchased from The Jackson Laboratory. Tumors were implanted into the right flank of immunodeficient mice under light isoflurane anesthesia. Mice were treated via IV tail vein or IP injection at doses indicated in the respective figures and preparations of all injectables are described in the section below. 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 reached approximately 1500 mm3in size for each respective group or following prolonged observation upon end of treatments when tumor growth were halted.

[0247] Preparation of injectables and administration schedule

[0248] TH2307 was solubilized to 16.7 mg / ml with formic acid-acidified DMSO and Solutol™ (1 :2 ratio), then diluted with sterile dextrose 5% in water (D5W) to the desired concentration for injection (i.e., 1.125 mg / ml for 7.5 mg / kg, 0.75 mg / ml for 5 mg / kg or 3.3 mg / kg, or 0.375 mg / ml for 2.5 mg / kg doses). TH2307 was given alone or in combination with anti-VEGF via weekly IV administration as described in the Brief Description of Drawings section.

[0249] TH2311 was solubilized to 16.7 mg / ml with formic acid-acidified DMSO and Solutol™ (1 :2 ratio), then diluted with sterile D5W to the desired concentration for injection (i.e., 0.77 mg / ml for 5.1 mg / kg dose). TH2311 was given alone via weekly IV administration as described in the Brief Description of Drawings section.

[0250] Anti-VEGF antibody (biosimilar to bevacizumab (Avastin®), FIG. 12) was provided by the National Research Council Canada (NRC) as a 3.4 mg / ml stock solution in phosphate buffer saline (PBS) pH 7.4. Stock solution was then diluted with sterile D5W to the desired concentration for injection (i.e., 1 mg / ml for 5 mg / kg dose). Anti-VEGF was given alone or in combination with TH2307 via biweekly IP administration as described in figure legends. The first biweekly dose was given 3 hours following TH2307 administration.

[0251] DM1 was purchased from MedChemExpress LLC (HY-19792) and solubilized to 8.3 mg / ml with formic acid-acidified DMSO and Solutol™ (1 :2 ratio), then diluted with sterile D5W to the desired concentration for injection (i.e., 0.158 mg / ml for 1 mg / kg or 0.079 mg / ml for 0.5 mg / kg doses). DM1 was given alone via weekly IV administration as described in the Brief Description of Drawings section.

[0252] All injectable solutions were filtered using 0.2 pm syringe filters before animal administrations; anti-VEGF diluted solutions were filtered using PES membrane syringe filters whereas TH2307, TH2311 and DM1 diluted solutions were filtered using DMSO-safe Nylon membrane syringe filters from PALL Corporation.

[0253] Example 3: Effect of maytansinoids and peptide-maytansinoid conjugates on the growth inhibition of various tumor cell lines in vitro

[0254] Table 2 shows the anti-proliferative activity of peptide-maytansinoid conjugates and unconjugated maytansinoids on ES-2 (ovarian), MDA-MB-231 (breast) and PANC-1 (pancreas) cancer cells. Peptide-maytansinoid conjugates were able to inhibit ES-2, MDA-MB-231 and PANC-1 tumor cell growth, confirming that the conjugation of the maytansinoids to the peptides does not inhibit their anti-proliferative activity. The results also demonstrate that conjugates comprising two maytansinoid molecules exhibit a more potent anti-proliferative activity relative to conjugates comprising one maytansinoid molecule, and that the conjugates comprising a cleavable linker have a lower IC50 relative to those comprising a non-cleavable linker.

[0255] Table 2. Evaluation of the anti-proliferation activity of free and conjugated maytansinoids drugs on ES-2, MDA-MB-231 and PANC-1 cancer cell lines. Cancer cells were incubated with increasing concentrations of unconjugated DM1 or peptide-conjugated DM1 as indicated in Example 2. The concentration at which they exert half of their maximal inhibitory effect (IC50) estimated by the WST-1 or WST-8 assay and calculated using the Graph Pad Prism™ software is indicated.

[0256] Additional in vitro experiments were performed on three colorectal tumor cell lines (HT-29, LoVo and HCT-116). As shown in Table 3, the two peptide-maytansinoid conjugates were able to inhibit tumor cell growth in all three cell lines with IC5o similar to that of free DM1 or DM4, confirming that the conjugation of these maytansinoids to the peptides does not inhibit their antiproliferative activity

[0257] Table 3. Evaluation of the anti-proliferation activity of DM1, DM4 and their conjugates on CRC cancer cell lines. HT-29, LoVo and HCT-116 cells were incubated with increasing concentrations of DM1 and TH2307 or DM4 and TH2311 as indicated in Example 2.

[0258] MTT assay was used to determine the IC50 using the Graph Pad prism software.

[0259] Example 4: Effects of maytansinoids and peptide-maytansinoid conjugates in colorectal, ovarian and breast cancer mouse models

[0260] The antitumor effects of unconjugated maytansinoids and peptide-maytansinoid conjugates were tested in vivo in the HT-29 and HCT-116 xenograft colorectal cancer mouse models, the MDA-MB-231 triple-negative breast cancer (TNBC) xenograft model, and the SKOV3 ovarian cancer mouse model. HT-29 cells express a mutated BRAF (BRAF V600E) whereas HCT-116 cells express a mutated K-Ras (G12D), and both cell lines were shown to exhibit resistance to anti-VEGF and anti-EGFR monotherapies (see, e.g., Hein et al., Cancer Cell International, volume 13, Article number: 94 (2013); Troiani et al., Clin Cancer Res (2014) 20 (14): 3775-3786). MDA- MB-231 cells carry the KRAS G13D mutation (Kim et al., Exp Mol Med. 2015 Jan; 47(1): e137).

[0261] The results reported in FIGs. 3A, 4A, 5A and 6A show a weak inhibition of tumor growth in mice treated with the unconjugated maytansinoid DM1. In contrast, administration of DM1 and DM4 as part of a conjugate with the sortilin-targeting peptide of SEQ ID NO:1 (TH2102) led a significantly improved inhibition of tumor growth, with complete tumor eradication at the highest doses in some of the cancer mouse models (FIGs. 3A, 4A, 5A and 6A). Based on results reported in the literature using the HT-29 CRC xenograft model (from Chung et al., EurJ Clin Invest. 2020 Apr 29:e13255. doi: 10.1111 / eci.13255 - FIG. 3A), complete tumor eradication is not achieved with an antibody-drug conjugate comprising an anti-HER2 antibody and DM1 (T-DM1 , Kadcyla) (FIG. 3C). The DM1 conjugate was shown to be more effective than the DM4 conjugate at inhibiting tumor growth in all models (FIGs. 4A, 5A and 6A), which was not expected given that DM4 is considered to be more potent than DM1 in the context of delivery by antibody-drug conjugates (ADCs) (see, e.g., Kalinovsky et al., Curr Issues Mol Biol. 2023 Oct; 45(10): 8112— 8125; Bouchard et al., Bioorganic & Medicinal Chemistry Letters, 24 (2014) 5357-5363).

[0262] Administration of the different conjugates was not associated with significant weight loss in any of the models tested (FIGs. 3B, 4B, 5B and 6B), in contrast to free DM1 that led to an important weight loss in some of the models. Thus, these results demonstrate that the peptide- maytansinoid conjugates are more effective at inhibiting tumor growth and better tolerated ( / .e., less toxic) than the corresponding unconjugated maytansinoids, and are effective against tumors with Ras or Ras pathway mutations.

[0263] Example 5: Internalization of peptide conjugates is sortilin-dependent

[0264] It was next assessed whether down-regulation of the expression of sortilin in HT-29 cancer cells could affect the internalization of the TH2307 conjugate. The results reported in FIGs. 7A- 7B show that that the uptake of TH2307 is reduced in cells transfected with an siRNA specific for Sortilin 1 (siSORTI) relative to the uptake measured in cells transfected with a control siRNA (siScrambled, siScr). These results demonstrate that the TH2307 conjugate specifically binds to sortilin at the surface of the HT-29 cancer cells, and thus that it suitable to deliver maytansinoids to sortilin-expressing cells.

[0265] Example 6: Detection and accumulation of TH2307 and release of DM1 in HT-29 cells

[0266] It was next assessed whether TH2307 may be detected inside HT-29 cells. HT-29 cells were incubated with vehicle or TH2307, followed by incubation with labelled anti-TH19P01 and anti-DM1 antibodies. As shown in FIG. 8, colocalization of the anti-TH19P01 and anti-DM1 antibodies was detected in TH2307-treated HT-29 cells. In the next series of experiments, the release of DM1 from the TH2307 conjugate was measured. As shown in FIG. 9, detection with the anti-TH19P01 antibody showed that TH2307 and TH19P01 bands almost migrated at the same level, and that similar profile was observed between all three tested conditions (non-reducing, reducing + heat, reducing - heat). Anti-DM1 antibody detection showed that DM1 band migrated lower than TH2307 band. It also showed that reducing conditions with heat (boiling) hampers DM1 detection and that reducing conditions without boiling induced a net release of DM1 from TH2307.

[0267] Next, western blots were performed with cell lysates obtained from HT-29 cells treated with TH2307 for 15 minutes up to 72 hours. Detection with the anti-TH19P01 antibody demonstrated the accumulation of TH2307 in cells over time (FIG. 10A) and detection with anti-DM1 antibody in non-reducing conditions confirmed this accumulation and showed that there is no or little release of DM1 from TH2307 conjugate (FIG. 10B). FIG. 10C shows the induction of DM1 release from conjugates by sample preparation under reducing conditions without boiling. Anti-DM1 antibody detection showed an increased DM1 signal over time, further supporting the accumulation of TH2307 in the cells.

[0268] Example 7: Estimation of the maximum tolerated dose for repeated treatments with DM1 and maytansinoid conjugates

[0269] The maximum tolerated dose (MTD) for repeated treatments with DM1 as well as for the TH2307 and TH2311 conjugates was estimated from mouse body weight loss. The results are reported in Table 4. Data are expressed in term of mg / kg / week for these molecules and in terms of payload content (mg of DM1 or DM4 / week). Animals received 7 cycles of both TH2307 and TH2311 conjugates at 5 mg / kg / week for a cumulative dose of 35 mg / kg compared to 3 cycles of DM1 at 0.5 mg / kg / week for a cumulative dose of 1.5 mg / kg for only the payload. Since DM1 content represents about 40% of both conjugate weights, the equivalent DM1 dose administered for both conjugates is 2 mg / kg / week for 7 cycles. Therefore, the total cumulative maytansinoids equivalent dose administered by both conjugates is 14 mg / kg, which is about 9.3-fold higher than the payload DM1.

[0270] Table 4 Example 8: DM1 content in the peptide drug conjugate TH2307 vs. the antibody drug conjugate T-DM1 (Kadcyla)

[0271] DM1 content in both TH2307 and T-DM1 was compared and the results are reported in Table 5. In TH2307, the DM1 content represents about 40% the conjugate mass (0.4 mg of DM1 / mg of TH2307) relative to about 1.7% in T-DM1 (0.017 mg of DM1 / mg of T-DM1). Therefore, the DM1 equivalent doses administered to mice with the usual doses of TH2307 (5 mg / kg) or T- DM1 (15 mg / kg) correspond to 2 mg / kg and 0.25 mg / kg of DM1 , respectively, which means that an 8-fold higher dose of DM1 is administered through the TH2307 conjugate relative to T-DM1.

[0272] Table 5

[0273] Example 9: Combination therapy of a peptide-maytansinoid conjugate with an anti-VEGF therapy in colorectal cancer tumor models

[0274] The effects of combining an anti-VEGF antibody with the maytansinoid conjugate TH2307 was next tested in the HT-116 tumor model. As noted above, HT-116 tumors express a mutated K-Ras (G12D) and were shown to exhibit resistance to anti-VEGF monotherapy. As shown in FIG. 11 A, administration of the anti-VEGF antibody alone led to a weak inhibition of tumor growth, confirming the resistance of the HT-116 cell line to anti-VEGF monotherapy. Administration of the maytansinoid conjugate TH2307 resulted in tumor growth inhibition relative to vehicle, but no tumor regression overthe study period. However, combining the anti-VEGF antibody with TH2307 resulted in potent tumor regression (FIG. 11 A). Administration of the conjugate or combination was not associated with significant weight loss (FIG. 11 B), in contrast to free DM1 that led to an important weight loss. These results provide evidence that the maytansinoid peptide conjugate reverses the resistance of HT-116 tumors to anti-VEGF therapy.

[0275] The combination therapy was next tested in a grade IV metastatic CRC patient-derived xenograft (PDX) model. As shown in FIG. 12A, administration of the anti-VEGF antibody alone led to a weak inhibition of tumor growth, indicating that the metastatic tumor is resistant to anti- VEGF monotherapy. Similarly, a weak inhibition of tumor growth was obtained following administration of DM1. Administration of the maytansinoid conjugate TH2307 resulted in some level of tumor growth inhibition relative to vehicle over the study period. However, combining the anti-VEGF antibody with TH2307 resulted in almost complete tumor growth inhibition (FIG. 12A). Administration of the conjugate or combination was not associated with significant weight loss (FIG. 12B). These results provide evidence that the maytansinoid peptide conjugate reverses the resistance of a patient-derived metastatic CRC to anti-VEGF therapy.

[0276] 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.

Claims

1. WHAT IS CLAIMED IS:1 . A conjugate comprising the following structure I, or a pharmaceutically acceptable salt thereof:A — L — B (I); whereinA is a maytansinoid;B is a peptide binding to a Sortilin receptor, wherein the peptide comprises an amino acid sequence having at least 60% identity with the amino acid sequence set forth in SEQ ID NO: 1 or 2:X1GVRAKAGVRN(Nle)FKSESYX2(SEQ ID NO:1)X1YKSLRRKAPRWDAPLRDPALRQLLX2(SEQ ID NO:2); wherein X1is cysteine (C) or is absent, and X2is cysteine (C) or is absent;L is a linker, wherein a first end of the linker is attached to a side chain of one or more of the amino acid or modified side chain residues of the peptide, and / or to the N- and / or C- terminal end of the peptide, and a second end of the linker is attached to the maytansinoid.

2. The conjugate of claim 1 , wherein L is a cleavable linker, wherein a first end of the linker is attached to a side chain of one or more of the lysine (K) and / or cysteine (C) residues of the peptide, and / or to the N- and / or C-terminal end of the peptide, and a second end of the linker is attached to a sulfur atom of the maytansinoid.

3. The conjugate or pharmaceutically acceptable salt thereof of claim 1 or 2, wherein the conjugate or salt thereof is of one of the following formulas la, lb, Ic or Id:A1 — L1 — B (la);A1 — L1 — B — L2 — A2 (lb);wherein L1 , L2, L3 and L4 are linkers which may be the same or different, and A1 , A2, A3 and A4 are maytansinoid molecules which may be the same or different.

4. The conjugate or pharmaceutically acceptable salt thereof of any one of claims 1 to 3, wherein the maytansinoid is DM1 (N2-deacetyl-N2-(3-mercapto-1-oxopropyl)-maytansine, mertansine) and / or DM4 (N2-deacetyl-N2-(4-mercapto-4-methyl-1 -oxopentyl) maytansine, ravtansine).

5. The conjugate or pharmaceutically acceptable salt thereof of claim 4, wherein the maytansinoid is DM1.

6. The conjugate or pharmaceutically acceptable salt thereof of claim 4, wherein the maytansinoid is DM4.

7. The conjugate or pharmaceutically acceptable salt thereof of any one of claims 1 to 6, wherein the peptide comprises an amino acid sequence having at least 80% identity with the amino acid sequence set forth in SEQ ID NO: 1 or 2.

8. The conjugate or pharmaceutically acceptable salt thereof of claim 7, wherein the peptide comprises an amino acid sequence having at least 90% identity with the amino acid sequence set forth in SEQ ID NO: 1 or 2.

9. The conjugate or pharmaceutically acceptable salt thereof of claim 8, wherein the peptide comprises the amino acid sequence set forth in SEQ ID NO: 1 or 2.

10. The conjugate or pharmaceutically acceptable salt thereof of claim 8, wherein the peptide consists of the amino acid sequence set forth in SEQ ID NO: 1 or 2.

11. The conjugate or pharmaceutically acceptable salt thereof of any one of claims 1 to 10, wherein X1and / or X2is C, and wherein the first end of the linker is attached to the side chain of X1and / or X2.

12. The conjugate or pharmaceutically acceptable salt thereof of claim 11 , wherein X1is absent and X2is C, and wherein the first end of the linker is attached to the side chain of X2.

13. The conjugate or pharmaceutically acceptable salt thereof of any one of claims 1 to 11 , wherein X1and X2are absent.

14. The conjugate or pharmaceutically acceptable salt thereof of any one of claims 1 to 13, wherein the linker is a cleavable linker.

15. The conjugate or pharmaceutically acceptable salt thereof of any one of claims 1 to 13, wherein the cleavable linker is a protease-cleavable linker.

16. The conjugate or pharmaceutically acceptable salt thereof of claim 15, wherein the protease is a cysteine protease.

17. The conjugate or pharmaceutically acceptable salt thereof of claim 16, wherein the cysteine protease is cathepsin B.

18. The conjugate or pharmaceutically acceptable salt thereof of any one of claims 1 to 17, wherein the linker is of one of the following structures:S-X3-CO, wherein X3is a linear or branched Ci-C8alkyl.

19. The conjugate or pharmaceutically acceptable salt thereof of claim 18, wherein the linker is one of the following structures: S-CH(CH3)-CH2-CH2-CO or S-CH2-CH2-CO.

20. The conjugate or pharmaceutically acceptable salt thereof of claim 1 , wherein the conjugate has one of the following structures:21 . The conjugate or pharmaceutically acceptable salt thereof of claim 20, wherein the conjugate has the following structure:

22. The conjugate or pharmaceutically acceptable salt thereof of claim 20, wherein the conjugate has the following structure:

23. A pharmaceutical composition comprising the conjugate or salt thereof of any one of claims 1 to 22, and a pharmaceutically acceptable excipient.

24. A method for treating a Sortilin-expressing cancer in a subject in need thereof comprising administering to the subject an effective amount of the conjugate or salt thereof of any one of claims 1 to 22, or the pharmaceutical composition of claim 23.

25. The method of claim 24, wherein the Sortilin-expressing cancer is a hematological cancer, ovarian cancer, endometrial cancer, cervix cancer, skin cancer, brain cancer, breastcancer, colorectal cancer, small intestine cancer, liver cancer, lung cancer, eye cancer, prostate cancer, head and neck cancer, stomach cancer, bone cancer, thyroid cancer, testis cancer, bladder cancer, kidney cancer, or pancreatic cancer, more particularly colorectal cancer, breast cancer, ovarian cancer or pancreatic cancer.

26. The method of claim 24 or 25, wherein the Sortilin-expressing cancer is a poor prognosis cancer.

27. The method of any one of claims 24 to 26, wherein the Sortilin-expressing cancer is an immunologically cold cancer.

28. The method of any one of claims 24 to 27, wherein the conjugate, salt thereof or composition is administered in combination with one or more additional active agents or therapies for cancer.

29. The method of claim 28, wherein the one or more additional active agents or therapies for cancer comprise an immunotherapy.

30. The method of claim 29, wherein the immunotherapy comprises an immune checkpoint inhibitor.31 . The method of claim 30, wherein the immune checkpoint inhibitor is a PD1 or PD-L1 inhibitor.

32. The method of claim 31 , wherein the PD1 or PD-L1 inhibitor is an anti-PD1 or anti-PD-L1 antibody.

33. The method of claim 28, wherein the one or more additional active agents or therapies for cancer comprise an anti-vascular endothelial growth factor (VEGF) therapy.

34. The method of any one of claims 24 to 33, wherein the cancer harbors mutations in a Ras protein or Ras pathway protein.

35. The method of claim 34, wherein the Ras protein is KRAS.

36. The method of claim 34, wherein the Ras pathway protein is BRAF.

37. The method of any one of claims 24 to 36, wherein the cancer is resistant to monotherapy with VEGF inhibitors.

38. Use of the conjugate or salt thereof of any one of claims 1 to 22, or the pharmaceutical composition of claim 23, for treating a Sortilin-expressing cancer in a subject.

39. Use of the conjugate or salt thereof of any one of claims 1 to 22, or the pharmaceutical composition of claim 23, for the manufacture of a medicament for treating a Sortilin-expressing cancer in a subject.

40. The use of claim 38 or 39, wherein the Sortilin-expressing cancer is a hematological cancer, ovarian cancer, endometrial cancer, cervix cancer, skin cancer, brain cancer, breast cancer, colorectal cancer, small intestine cancer, liver cancer, lung cancer, eye cancer, prostate cancer, head and neck cancer, stomach cancer, bone cancer, thyroid cancer, testis cancer, bladder cancer, kidney cancer, or pancreatic cancer, more particularly colorectal cancer, breast cancer, ovarian cancer or pancreatic cancer.

41. The use of any one of claims 38 to 40, wherein the Sortilin-expressing cancer is a poor prognosis cancer.

42. The use of any one of claims 38 to 41 , wherein the Sortilin-expressing cancer is an immunologically cold cancer.

43. The use of any one of claims 38 to 42, wherein the conjugate, salt thereof, composition or medicament is for administration in combination with one or more additional active agents or therapies for cancer.

44. The use of claim 43, wherein the one or more additional active agents or therapies for cancer comprise an immunotherapy.

45. The use of claim 44, wherein the immunotherapy comprises an immune checkpoint inhibitor.

46. The use of claim 45, wherein the immune checkpoint inhibitor is a PD1 or PD-L1 inhibitor.

47. The use of claim 46, wherein the PD1 or PD-L1 inhibitor is an anti-PD1 or anti-PD-L1 antibody.

48. The use of claim 43, wherein the one or more additional active agents or therapies for cancer comprise an anti-vascular endothelial growth factor (VEGF) therapy.

49. The use of any one of claims 38 to 48, wherein the cancer harbors mutations in a Ras protein or Ras pathway protein.

50. The use of claim 49, wherein the Ras protein is KRAS.51 . The use of claim 49, wherein the Ras pathway protein is BRAF.

52. The use of any one of claims 38 to 51 , wherein the cancer is resistant to monotherapy with VEGF inhibitors.

53. The conjugate or salt thereof of any one of claims 1 to 22, or the pharmaceutical composition of claim 23, for use in the treatment of a Sortilin-expressing cancer in a subject.

54. The conjugate, salt thereof or pharmaceutical composition for use according to claim 53, wherein the Sortilin-expressing cancer is a hematological cancer, ovarian cancer, endometrial cancer, cervix cancer, skin cancer, brain cancer, breast cancer, colorectalcancer, small intestine cancer, liver cancer, lung cancer, eye cancer, prostate cancer, head and neck cancer, stomach cancer, bone cancer, thyroid cancer, testis cancer, bladder cancer, kidney cancer, or pancreatic cancer, more particularly colorectal cancer, breast cancer, ovarian cancer or pancreatic cancer.

55. The conjugate, salt thereof or pharmaceutical composition for use according to claim 54, wherein the Sortilin-expressing cancer is a poor prognosis cancer.

56. The conjugate, salt thereof or pharmaceutical composition for use according to claim 54 or 55, wherein the Sortilin-expressing cancer is an immunologically cold cancer.

57. The conjugate, salt thereof or pharmaceutical composition for use according to any one of claims 54 to 56, wherein the conjugate, salt thereof or composition is for administration in combination with one or more additional active agents or therapies for cancer.

58. The conjugate, salt thereof or pharmaceutical composition for use according to claim 57, wherein the one or more additional active agents or therapies for cancer comprise an immunotherapy.

59. The conjugate, salt thereof or pharmaceutical composition for use according to claim 58, wherein the immunotherapy comprises an immune checkpoint inhibitor.

60. The conjugate, salt thereof or pharmaceutical composition for use according to claim 59, wherein the immune checkpoint inhibitor is a PD1 or PD-L1 inhibitor.

61. The conjugate, salt thereof or pharmaceutical composition for use according to claim 60, wherein the PD1 or PD-L1 inhibitor is an anti-PD1 or anti-PD-L1 antibody.

62. The conjugate, salt thereof or pharmaceutical composition for use according to claim 57, wherein the one or more additional active agents or therapies for cancer comprise an anti- vascular endothelial growth factor (VEGF) therapy.

63. The conjugate, salt thereof or pharmaceutical composition for use according to any one of claims 54 to 62, wherein the cancer harbors mutations in a Ras protein or Ras pathway protein.

64. The conjugate, salt thereof or pharmaceutical composition for use according to claim 63, wherein the Ras protein is KRAS.

65. The conjugate, salt thereof or pharmaceutical composition for use according to claim 64, wherein the Ras pathway protein is BRAF.

66. The conjugate, salt thereof or pharmaceutical composition for use according to of any one of claims 54 to 65, wherein the cancer is resistant to monotherapy with VEGF inhibitors.

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