Tumor-infiltrating lymphocyte therapy

Abstract

The present disclosure generally relates to methods of treating cancer by administering autologous tumor infiltrating lymphocytes (TILs) isolated from a subject who has previously been treated with an anti-cancer therapy that includes an anti-clusterin antibody or an antigen-binding fragment thereof. The disclosed methods include administering the anti-cancer therapy to the subject, isolating the TILs, and re-infusing the TILs into the subject. The disclosure also relates to the use of anti-clusterin antibodies or antigen-binding fragments thereof in in vitro or ex vivo methods of generating tumor infiltrating lymphocytes (TILs).

Description

[Technical field]

[0001] The present disclosure generally relates to methods of treating cancer by administering autologous tumor infiltrating lymphocytes (TILs) isolated from a subject who has previously been treated with an anti-cancer therapy comprising an anti-clusterin antibody or an antigen-binding fragment thereof. The disclosed methods include administering the anti-cancer therapy to the subject, isolating the TILs, and re-infusing the TILs into the subject. The disclosure also relates to the use of anti-clusterin antibodies or antigen-binding fragments thereof in in vitro or ex vivo methods of generating tumor infiltrating lymphocytes (TILs). [Background technology]

[0002] Immune cell therapy of solid tumors consists of two different approaches: the adoptive transfer of naturally occurring tumor-specific T cells isolated from tumor-infiltrating lymphocytes (TILs) or the transfer of genetically modified T lymphocytes expressing transgenic T cell receptors (tg-TCRs) specific for tumor antigens or chimeric antigen receptors (CARs) composed of a single-chain variable region of a monoclonal antibody fused to the endodomain of a T cell signaling molecule.

[0003] TIL therapy has a long history of development with multiple clinical trials at centers around the world and has consistently demonstrated long-lasting clinical response rates (approximately 50%) in advanced melanoma and, more recently, cervical cancer.

[0004] The distinct advantages of TIL treatment are the broad nature of T cell recognition of both defined and undefined tumor antigens, the context of all possible MHC molecules rather than a single specificity of tg-TCR or CAR-transduced T cells, and the limited MHC coverage of tg-TCR T cells. On-target / off-tumor toxicity is relatively rare with TIL therapy but is a major issue faced by genetically modified T cell therapy.

[0005] Thus, the treatment of refractory cancers using adoptive transfer of TILs represents a powerful approach to the treatment of patients with poor prognosis. Gattinoni et al., Nat. Rev. Immunol. 2006, 6, 383-393. The "rapid expansion process" (REP) following IL-2-based TIL expansion has become the preferred method of TIL expansion due to its speed and effectiveness (Dudley et al., Science 2002, 298, 850-54; Dudley et al., J. Clin. Oncol. 2005, 23, 2346-57; Dudley et al., J. Clin. Oncol. 2008, 26, 5233-39; Riddell et al., Science 1992, 257, 238-41; Dudley et al., J. Immunother. 2003, 26, 332-42). REP can result in a 1,000-fold expansion of TILs over a 14-day period, but requires an excess (e.g., 200-fold) of irradiated allogeneic peripheral blood mononuclear cells (PBMCs, also known as mononuclear cells (MNCs)) as feeder cells, often from multiple donors, as well as anti-CD3 antibodies (OKT3) and high doses of interleukin 2 (IL-2) (Dudley et al., J. Immunother. 2003, 26, 332-42). TILs that have undergone the REP procedure have been successfully used for adoptive cell therapy following host immunosuppression in patients with melanoma. Current infusion tolerance parameters depend on the composition of the TILs (e.g., CD28, CD8, or CD4 positive) and the readout of fold expansion and viability of the REP product.

[0006] Methods of preparing and expanding TILs are described, for example and without limitation, in Jin J. et al., J Immunother. 35(3):283-292, 2012; Dudley, ME et al., J Immunother. 26(4):332-342, 2003; International Patent Application No. PCT / US2018 / 01633, filed January 5, 2018, published October 4, 2018 as WO 2018 / 182817; and International Patent Application No. PCT / US2019 / 052681, filed September 24, 2019, published April 2, 2020 as WO 2020 / 068816; No. PCT / US2015 / 025313, filed on October 15, 2015 and published as WO 2015 / 157636, and International Patent Application No. PCT / US2019 / 052681, filed on September 24, 2019 and published on April 2, 2020 as WO 2020 / 068816, the entire contents of which are incorporated herein by reference.

[0007] Unfortunately, some patients have tumors that are poorly infiltrated by immune cells and therefore adoptive cell therapy is expected to be of limited benefit.

[0008] There remains a need to increase the presence of TILs in tumors for modulation of in vivo antitumor immune responses and adoptive cell therapy. [Prior art documents] [Patent documents]

[0009] [Patent Document 1] International Patent Application No. PCT / US2018 / 01633 [Patent Document 2] International Publication No. 2018 / 182817 [Patent Document 3] International Patent Application No. PCT / US2019 / 052681 [Patent Document 4] International Publication No. 2020 / 068816 [Patent Document 5] International Patent Application No. PCT / US2015 / 025313 [Patent Document 6] International Publication No. 2015 / 157636 [Patent Document 7] International Publication No. 2020 / 068816 [Patent Document 8] International Patent Application No. PCT / CA2021 / 050572 [Patent Document 9] International Patent Application No. PCT / CA2022 / 050632 [Patent Document 10] International Patent Application No. PCT / CA2006 / 001505 [Patent Document 11] International Publication No. 2007 / 030930 [Patent Document 12] International Patent Application No. PCT / CA2010 / 0001882 [Patent Document 13] International Publication No. 2011 / 063523 [Patent Document 14] US Patent Application Publication No. 2018 / 0282694 [Non-patent literature]

[0010] [Non-Patent Document 1] Gattinoni et al., Nat. Rev. Immunol. 2006, 6, 383-393 [Non-Patent Document 2] Dudley et al., Science 2002, 298, 850-54 [Non-Patent Document 3] Dudley et al., J. Clin. Oncol. 2005, 23, 2346-57 [Non-Patent Document 4] Dudley et al., J. Clin. Oncol. 2008, 26, 5233-39 [Non-Patent Document 5] Riddell et al., Science 1992, 257, 238-41 [Non-Patent Document 6] Dudley et al., J. Immunother. 2003, 26, 332-42 [Non-Patent Document 7] Jin J. et al., J Immunother. 35(3):283-292, 2012 [Non-Patent Document 8] Dudley, M.E. et al., J Immunother. 26(4): 332-342, 2003 [Non-Patent Document 9] Ojima et al., Expert Opin Ther Pat. 2016: 26(1): 1-20 [Non-Patent Document 10] Gacerez, AT et al., J Cell Physiol. 231(12):2590–2598 (2016) [Non-Patent Document 11] Sadelain, M. et al., Cancer Discovery, 3(4):388-98 (2013) [Non-Patent Document 12] Zhang, C. et al., Biomarker Research, 5:22 (2017) [Non-Patent Document 13] Antibody Engineering Vol. 2, Chapter 3 by Andrew CR Martin Summary of the Invention

[0011] The applicant has surprisingly discovered that treatment with an anti-clusterin antibody or an antigen-binding fragment thereof results in increased intratumoral immune infiltration (see International Patent Application No. PCT / CA2021 / 050572, filed April 27, 2021, and International Patent Application No. PCT / CA2022 / 050632, filed April 26, 2022, the entire contents of each of which are incorporated by reference into this specification).

[0012] Preliminary data from a Phase II clinical trial (NCT04364620) aimed at evaluating a combination treatment including an anti-clusterin antibody (AB-16B5, also known as humanized 16B5) and docetaxel in subjects with metastatic non-small cell lung cancer show similar intratumoral immune cell infiltration (see International Patent Application No. PCT / CA2022 / 050632, filed April 26, 2022).

[0013] Thus, an anti-cancer therapy comprising an anti-clusterin antibody or an antigen-binding fragment thereof can be administered to a subject with cancer to promote infiltration of immune cells in the tumor microenvironment. Thus, a preparation of tumor-infiltrating lymphocytes is generated from the tumor of a treated subject for use in adoptive cell therapy.

[0014] The present disclosure provides a method of treating a subject having cancer, the method comprising administering to the subject an anti-cancer therapy comprising an anti-clusterin antibody or an antigen-binding fragment thereof, isolating and expanding tumor-infiltrating lymphocytes (TILs) from the subject's tumor, and re-injecting the preparation of TILs into the subject.

[0015] In some embodiments, the method involves administering a preparation of TILs disclosed herein. In some embodiments, the preparation of TILs comprises one or more TIL cultures. In some embodiments, the preparation of TILs is a TIL culture.

[0016] In accordance with the present disclosure, the anti-cancer treatment consists of an anti-clusterin antibody or antigen-binding fragment thereof provided as the sole anti-cancer agent.

[0017] In accordance with the present disclosure, the anti-cancer treatment comprises an anti-clusterin antibody or antigen-binding fragment thereof and another anti-cancer agent. Thus, the anti-cancer treatment can be a combination therapy.

[0018] In some embodiments, the combination therapy comprises an anti-clusterin antibody, or an antigen-binding fragment thereof, and radiation therapy.

[0019] In other embodiments, the combination therapy comprises an anti-clusterin antibody or antigen-binding fragment thereof and chemotherapy.

[0020] The present disclosure also provides a method of treating cancer with tumor-infiltrating lymphocytes (TILs) isolated and expanded from a tumor isolated from a subject treated with an anti-cancer therapy comprising an anti-clusterin antibody or an antigen-binding fragment thereof.

[0021] In some embodiments, subjects undergo lymphodepletion conditioning prior to TIL infusion.

[0022] In some embodiments, the TILs are isolated and expanded by in vitro or ex vivo tumor infiltrating lymphocyte generation methods to generate a preparation of TILs. In some embodiments, the methods involve culturing the TILs.

[0023] In some embodiments, the methods may include removing tumor cells from the TIL culture.

[0024] In some embodiments, the method further comprises: + A step of selecting the cells may be included.

[0025] In some embodiments, the method comprises obtaining CD3 + A step of selecting the cells may be included.

[0026] In some embodiments, the method further comprises: + A step of selecting the cells may be included.

[0027] In some embodiments, the method comprises obtaining CD8 + A step of selecting the cells may be included.

[0028] In some embodiments, the methods may include selecting cells with intermediate to high levels of INFγ secretion.

[0029] In some embodiments, the TILs are selected for their anti-tumor activity in vitro.

[0030] Typically, TILs with anti-tumor activity are selected for use in autologous adoptive cell therapy.

[0031] In some embodiments, the TILs are isolated from a subject who is being treated or will be treated with an anti-cancer therapy that includes an anti-clusterin antibody, or an antigen-binding fragment thereof, as a single agent.

[0032] In some embodiments, the TILs are isolated from a subject who has been or will be treated with a combination therapy comprising an anti-clusterin antibody, or antigen-binding fragment thereof, and a chemotherapeutic agent.

[0033] Exemplary embodiments of chemotherapeutic agents include alkylating agents, antimetabolites, alkaloids, antitumor antibiotics, or combinations thereof.

[0034] In some cases, the alkylating agent may be selected from altretamine, busulfan, carboplatin, carmustine, cisplatin, cyclophosphamide, dacarbazine, ifosfamide, lomustine, melphalan, temozolomide, trabectedin, or a derivative or analog thereof.

[0035] In some cases, the antimetabolite may be selected from, for example, 5-fluorouracil, 6-mercaptopurine, azacitidine, capecitabine, clofarabine, cytarabine, floxuridine, fludarabine, gemcitabine, methotrexate, pemetrexed, pentostatin, pralatrexate, trifluridine, tipiracil, or a derivative or analog thereof.

[0036] In some cases, the alkaloid can be selected from, for example, vincristine, vinblastine, vinorelbine, taxanes, etoposide, teniposide, irinotecan, topotecan, or derivatives or analogs thereof.

[0037] Exemplary embodiments of taxanes include docetaxel, paclitaxel, and derivatives or analogs thereof, such as, but not limited to, Abraxane®, cabazitaxel, larotaxel, mirataxel, ortataxel, tesetaxel, and others described in Ojima et al., Expert Opin Ther Pat. 2016: 26(1): pp. 1-20, the entire contents of which are incorporated herein by reference.

[0038] In some instances, the antitumor antibiotic may be selected from, for example, daunorubicin, doxorubicin, liposomal doxorubicin, epirubicin, idarubicin, valrubicin, derivatives or analogs thereof.

[0039] In some embodiments, the chemotherapeutic agent is docetaxel.

[0040] In some embodiments, the chemotherapeutic agent is paclitaxel.

[0041] In some embodiments, the tumor is resectable.

[0042] In some embodiments, the subject has a functional immune system.

[0043] In some embodiments, the TILs are obtained from a tumor or tumor fragment isolated by biopsy.

[0044] In some embodiments, the TILs are obtained by a method that includes an initial culture phase and an expansion phase.

[0045] In accordance with the present disclosure, a method for generating tumor infiltrating lymphocytes in vitro or ex vivo may include contacting a tumor fragment with an anti-clusterin antibody or an antigen-binding fragment thereof.

[0046] In accordance with the present disclosure, anti-clusterin antibodies or antigen-binding fragments thereof may be present and / or maintained during the initial culture phase of the tumor infiltrating lymphocyte generation method.

[0047] Alternatively, in accordance with the present disclosure, anti-clusterin antibodies or antigen-binding fragments thereof may be present and / or maintained during the expansion phase of the tumor infiltrating lymphocyte generation method.

[0048] In some embodiments, the methods of the disclosure include administering TILs that are not genetically modified. However, TILs can be genetically modified to cause them to express or overexpress a protein or peptide.

[0049] In some embodiments, the preparation of TILs is not genetically modified.

[0050] In some embodiments, the preparation of TILs comprises genetically modified TILs.

[0051] In some embodiments, the preparation of TILs comprises TILs that express a chimeric antigen receptor.

[0052] In some embodiments, the preparation of TILs comprises TILs that express a transgenic T cell receptor.

[0053] In some embodiments, the preparation of TILs comprises TILs isolated from a primary tumor.

[0054] In some embodiments, the preparation of TILs comprises TILs isolated from a metastatic lesion.

[0055] In accordance with the present disclosure, TILs can be isolated from a subject who has undergone previous treatment with an anti-cancer therapy described herein.

[0056] In an exemplary embodiment, the subject may have undergone prior treatment with an anti-clusterin antibody, or antigen-binding fragment thereof, and a taxane, such as docetaxel or paclitaxel.

[0057] In accordance with the present disclosure, anti-clusterin antibodies or antigen-binding fragments thereof may be administered at a dose and / or dosing interval and / or treatment duration sufficient to result in infiltration of immune cells into the tumor microenvironment.

[0058] In accordance with the present disclosure, docetaxel may be administered at a dose and / or dosing interval and / or treatment duration sufficient to allow chemotherapy-induced immunogenic modulation of the tumor.

[0059] In accordance with the present disclosure, the method includes administering an anti-clusterin antibody or an antigen-binding fragment thereof comprising a light chain variable region comprising the complementarity determining region (CDR) of the light chain variable region set forth in SEQ ID NO: 9, and a heavy chain variable region comprising the CDR of the heavy chain variable region set forth in SEQ ID NO: 10.

[0060] In accordance with the present disclosure, the method includes administering an anti-clusterin antibody or an antigen-binding fragment thereof comprising a light chain variable region having an amino acid sequence that has at least 80% identity to the amino acid sequence set forth in SEQ ID NO:9, and a heavy chain variable region having an amino acid sequence that has at least 80% identity to the amino acid sequence set forth in SEQ ID NO:10.

[0061] In accordance with the present disclosure, the method includes administering an anti-clusterin antibody or antigen-binding fragment thereof comprising a light chain having an amino acid sequence that has at least 80% identity to the amino acid sequence set forth in SEQ ID NO:11, and a heavy chain having an amino acid sequence that has at least 80% identity to the amino acid sequence set forth in SEQ ID NO:12.

[0062] In accordance with the present disclosure, the method includes administering an anti-clusterin antibody or antigen-binding fragment thereof comprising a light chain variable region having the amino acid sequence set forth in SEQ ID NO:9 and a heavy chain variable region having the amino acid sequence set forth in SEQ ID NO:10 to bind to clusterin.

[0063] In some embodiments, the anti-clusterin antibody or antigen-binding fragment thereof is administered prior to isolation of the TILs. In some embodiments, the anti-clusterin antibody or antigen-binding fragment thereof and the chemotherapeutic agent are administered prior to isolation of the TILs. In some embodiments, one or more treatment cycles are administered prior to isolation of the TILs.

[0064] In some embodiments, the anti-cancer treatments described herein may also be administered after the adoptive cell therapy.

[0065] In some embodiments, the anti-clusterin antibody or antigen-binding fragment thereof is administered after infusion of the preparation of TILs. In some embodiments, the anti-clusterin antibody or antigen-binding fragment thereof and the chemotherapeutic agent are administered after infusion of the TILs. In some embodiments, one or more treatment cycles are administered after infusion of the TILs.

[0066] In some embodiments, the anti-clusterin antibody or antigen-binding fragment thereof is administered at a dose of between about 3 mg / kg and about 20 mg / kg prior to isolation of the TILs or after infusion of the TILs.

[0067] In some embodiments, the anti-clusterin antibody or antigen-binding fragment thereof is administered at a dose of approximately 6 mg / kg.

[0068] In some embodiments, the anti-clusterin antibody or antigen-binding fragment thereof is administered at a dose of approximately 9 mg / kg.

[0069] In some embodiments, the anti-clusterin antibody or antigen-binding fragment thereof is administered at a dose of approximately 12 mg / kg.

[0070] In some embodiments, the anti-clusterin antibody, or antigen-binding fragment thereof, is administered weekly.

[0071] In some embodiments, docetaxel is administered at approximately 60 mg / m prior to isolation of the TILs or after infusion of the TILs. 2 ~ approx. 100 mg / m 2It is administered in doses between 100 and 200 mg / kg.

[0072] In some embodiments, docetaxel is administered once every three weeks.

[0073] In some embodiments, docetaxel is approximately 60 mg / m 2 is administered at a dose of

[0074] In some embodiments, docetaxel is approximately 75 mg / m 2 is administered at a dose of

[0075] In some embodiments, the subject is receiving a weekly dose of approximately 12 mg / kg of an anti-clusterin antibody or antigen-binding fragment thereof and approximately 75 mg / m 2 once every three weeks.

[0076] In some embodiments, the subject is receiving a weekly dose of approximately 12 mg / kg of an anti-clusterin antibody or antigen-binding fragment thereof and approximately 60 mg / m 2 once every three weeks.

[0077] In some embodiments, the subject is receiving a weekly dose of approximately 9 mg / kg of an anti-clusterin antibody or antigen-binding fragment thereof and approximately 75 mg / m 2 once every three weeks.

[0078] In some embodiments, the subject is receiving a weekly dose of approximately 9 mg / kg of an anti-clusterin antibody or antigen-binding fragment thereof and approximately 60 mg / m 2 once every three weeks.

[0079] In some embodiments, the subject is receiving a weekly dose of approximately 6 mg / kg of an anti-clusterin antibody or antigen-binding fragment thereof and approximately 75 mg / m 2 once every three weeks.

[0080] In some embodiments, the subject is receiving a weekly dose of approximately 6 mg / kg of an anti-clusterin antibody or antigen-binding fragment thereof and approximately 60 mg / m 2 once every three weeks.

[0081] In some embodiments, the subject is receiving a weekly dose of about 3 mg / kg of an anti-clusterin antibody or antigen-binding fragment thereof and about 75 mg / m 2 once every three weeks.

[0082] In some embodiments, the subject is receiving a weekly dose of about 3 mg / kg of an anti-clusterin antibody or antigen-binding fragment thereof and about 60 mg / m 2 once every three weeks.

[0083] In some embodiments, the anti-clusterin antibody, or antigen-binding fragment thereof, and docetaxel are administered on the same day.

[0084] In some embodiments, the anti-clusterin antibody or antigen-binding fragment thereof and / or docetaxel are administered by infusion over a time frame of approximately one hour.

[0085] In some embodiments, the methods of the disclosure are for treating a subject as described herein.

[0086] In some embodiments, the subject has carcinoma.

[0087] In some embodiments, the subject has metastatic cancer.

[0088] In some embodiments, the subject has endometrial cancer, breast cancer, liver cancer, prostate cancer, kidney cancer, bladder cancer, cervical cancer, ovarian cancer, colorectal cancer, pancreatic cancer, lung cancer, gastric cancer, head and neck cancer, thyroid cancer, cholangiocarcinoma, mesothelioma, or melanoma.

[0089] In some embodiments, the subject has metastatic endometrial cancer, metastatic breast cancer, metastatic liver cancer, metastatic prostate cancer, metastatic kidney cancer, metastatic bladder cancer, metastatic cervical cancer, metastatic ovarian cancer, metastatic colorectal cancer, metastatic pancreatic cancer, metastatic lung cancer, metastatic gastric cancer, metastatic head and neck cancer, metastatic thyroid cancer, metastatic cholangiocarcinoma, metastatic mesothelioma, or metastatic melanoma.

[0090] In some embodiments, the subject is not immunosuppressed or has not received an immunosuppressant within 14, 13, 12, 11, 10, 9, 8, 7, 6, 5, 4, 3, 2 days, or 1 day prior to treatment with the anti-clusterin antibody or antigen-binding fragment thereof or treatment with the anti-clusterin antibody or antigen-binding fragment thereof and docetaxel combination therapy.

[0091] In some embodiments, the subject is a human subject.

[0092] The methods of the present disclosure include + This may result in the preparation of TILs or TIL cultures that include T cells.

[0093] The methods of the present disclosure include + This may result in the preparation of TILs or TIL cultures that include T cells.

[0094] The methods of the disclosure may result in the preparation of TILs or TIL cultures that include B cells.

[0095] The methods of the present disclosure may result in the preparation of TILs or TIL cultures that include NK cells.

[0096] The methods of the disclosure may result in the preparation of TILs or TIL cultures that contain NK T cells.

[0097] The methods of the disclosure may result in the preparation of TILs or TIL cultures with anti-tumor activity.

[0098] Accordingly, the present disclosure provides a preparation of tumor infiltrating lymphocytes (TILs) obtained by the methods described herein.

[0099] Accordingly, the present disclosure provides tumor infiltrating lymphocyte (TIL) cultures obtained by the methods described herein.

[0100] Accordingly, the present disclosure also provides a preparation of tumor infiltrating lymphocytes (TILs) obtained by a method of treating a subject with cancer with an anti-cancer therapy comprising an anti-clusterin antibody, or an antigen-binding fragment thereof.

[0101] In some embodiments, the preparation of TILs is a preparation of expanded TILs.

[0102] The present disclosure also provides a TIL culture obtained by the method of treating a subject with cancer with an anti-cancer therapy comprising an anti-clusterin antibody or an antigen-binding fragment thereof.

[0103] In some embodiments, the TIL preparation or TIL culture is obtained from a subject who has been or will be treated with an anti-clusterin antibody or antigen-binding fragment thereof, either as a single agent or as combination therapy with a chemotherapeutic agent.

[0104] In some embodiments, the TILs are not genetically modified.

[0105] In other embodiments, the TILs are genetically modified.

[0106] In some embodiments, the preparation of TILs includes TILs that are genetically modified.

[0107] In some embodiments, the preparation of TILs comprises TILs that express a chimeric antigen receptor.

[0108] In some embodiments, the preparation of TILs comprises TILs that express a transgenic T cell receptor.

[0109] In some embodiments, the preparation of TILs is provided in an infusion bag.

[0110] In some embodiments, the preparation of tumor infiltrating lymphocytes (TILs) comprises a majority of CD45 + Contains cells.

[0111] In some embodiments, the preparation of tumor infiltrating lymphocytes (TILs) comprises a majority of CD3 + Contains cells.

[0112] In some embodiments, the preparation of tumor infiltrating lymphocytes (TILs) comprises a majority of CD4 + Contains cells.

[0113] In some embodiments, the preparation of tumor infiltrating lymphocytes (TILs) comprises a majority of CD8 + Contains cells.

[0114] In some embodiments, a preparation of tumor infiltrating lymphocytes (TILs) comprises a majority of CD4 + or CD8 + Includes cells that are cells.

[0115] In some examples, the preparation of tumor infiltrating lymphocytes may contain at least 50% CD8+ lymphocytes. In other examples, the preparation of tumor infiltrating lymphocytes may contain at least 60% CD8+ lymphocytes. In still other examples, the preparation of tumor infiltrating lymphocytes may contain at least 70% CD8+ lymphocytes. In additional examples, the preparation of tumor infiltrating lymphocytes may contain at least 75% CD8+ lymphocytes. In additional examples, the preparation of tumor infiltrating lymphocytes may contain greater than 75% CD8+ lymphocytes. The preparation of tumor infiltrating lymphocytes may secrete intermediate to high levels of INFγ.

[0116] In an exemplary embodiment, the preparation of tumor infiltrating lymphocytes may consist of tumor infiltrating lymphocyte cultures each comprising at least 50% CD8+ lymphocytes. In another exemplary embodiment, the preparation of tumor infiltrating lymphocytes may consist of tumor infiltrating lymphocyte cultures each comprising at least 50% CD8+ lymphocytes and secreting intermediate to high levels of INFγ. In another exemplary embodiment, the preparation of tumor infiltrating lymphocytes consists of tumor infiltrating lymphocyte cultures each comprising at least 60% CD8+ lymphocytes and secreting high levels of INFγ. In an additional exemplary embodiment, the preparation of tumor infiltrating lymphocytes consists of tumor infiltrating lymphocyte cultures each comprising at least 70% CD8+ lymphocytes and secreting high levels of INFγ. In yet an additional exemplary embodiment, the preparation of tumor infiltrating lymphocytes consists of tumor infiltrating lymphocyte cultures each comprising at least 75% CD8+ lymphocytes and secreting high levels of IFNγ. In yet a further exemplary embodiment, the preparation of tumor infiltrating lymphocytes is comprised of tumor infiltrating lymphocyte cultures each of which contains greater than 75% CD8+ lymphocytes and secretes high levels of IFNγ.

[0117] In some embodiments, each of the tumor infiltrating lymphocyte cultures may be obtained from the same tumor. In other embodiments, each of the tumor infiltrating lymphocyte cultures may be obtained from a different tumor.

[0118] In other examples, the preparation of tumor infiltrating lymphocytes may contain less than 10% CD4+ lymphocytes. In yet other examples, the preparation of tumor infiltrating lymphocytes may contain less than 7.5% CD4+ lymphocytes. In other examples, the preparation of tumor infiltrating lymphocytes may contain less than 5% CD4+ lymphocytes. In other examples, the preparation of tumor infiltrating lymphocytes may contain 2% CD4+ lymphocytes or less.

[0119] The preparation of tumor infiltrating lymphocytes may be provided as an article of manufacture, exemplary embodiments of which include vials, flasks, syringes, infusion bags, and the like. [Brief description of the drawings]

[0120] [Figure 1] 4T1 lung metastases are immunologically "cold" and prevent immune lymphocytic infiltration. CD3+ and CD8+ T cells are present at the periphery of 4T1 lung metastases due to the creation of a restrictive tumor microenvironment as a result of epithelial-mesenchymal transition, which prevents lymphocytic infiltration. [Figure 2A] Inhibition of EMT by 16B5 antibody sCLU mAb results in B (B220) and T (CD3, CD4, CD8) lymphocyte infiltration in 4T1 lung metastases. [Figure 2B] Photographs of human tumor biopsies from patients treated with AB-16B5 as a single agent. [Figure 2C] Photographs of human tumor biopsies from patients treated with AB-16B5 as a single agent. [Figure 2D] Photographs of human tumor biopsies from patients treated with AB-16B5 as a single agent. [Diagram 3] 1 is a graph of the number of metastatic lung nodules in 4T1-implanted animals treated with AB-16B5 as monotherapy or in combination with docetaxel. [Figure 4A] FIG. 4T1 lung metastases from animals treated with AB-16B5 as monotherapy or in combination with docetaxel are infiltrated with B and T lymphocytes. 4T1 lung metastases were dissected 36 days after implantation and processed with collagenase and hyaluronidase for immunophenotyping by flow cytometry. [Figure 4B] FIG. 4T1 lung metastases from animals treated with AB-16B5 as monotherapy or in combination with docetaxel are infiltrated with B and T lymphocytes. 4T1 lung metastases were dissected 36 days after implantation and processed with collagenase and hyaluronidase for immunophenotyping by flow cytometry. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0121] Further scope, applicability, and advantages of the present disclosure will become apparent from the following non-limiting detailed description, which, however, should be understood to be given by way of example only, although it refers to the accompanying drawings and illustrates exemplary embodiments of the present disclosure.

[0122] definition Unless otherwise indicated, the amino acid numbering shown for the dimerization domains is according to the EU numbering system.

[0123] In the context of describing embodiments (particularly in the context of the claims), the use of the terms "a," "an," and "the," and similar references should be construed to include both the singular and the plural, unless otherwise indicated herein or clearly contradicted by context.

[0124] Unless specifically stated otherwise or clear from the context, as used herein, the term "or" is understood to be inclusive and includes both "or" and "and."

[0125] The term "and / or", when used herein, should be considered as a specific disclosure of each of the specified features or components in the presence or absence of the other.

[0126] The terms "comprising," "having," "including," and "containing" should be construed as open-ended (i.e., meaning "including but not limited to"), unless otherwise noted. The term "consisting of" should be construed as exclusive.

[0127] For the purposes of this disclosure, the term "treatment" refers to both therapeutic and prophylactic treatment measures. Subjects in need of treatment include those already with the disorder as well as those prone to have the disorder or those in which the disorder is to be prevented.

[0128] The term "EMT signature," as used herein, refers to changes indicative of a loss of an epithelial phenotype and / or a gain of a mesenchymal phenotype, which are observable at the cellular level and / or observable or measurable at the gene or protein level.

[0129] The term "about" or "approximately" with respect to a given value means that variation of the value is contemplated. In some embodiments, the term "about" or "approximately" generally refers to a range within + / - 20 percent, within + / - 10 percent, within + / - 5 percent, within + / - 4 percent, within + / - 3 percent, within + / - 2 percent, or within + / - 1 percent of a given value or range.

[0130] The term "functional immune system" with respect to a subject means that the subject's immune system is essentially unaffected by cancer or medication, or that the subject is not immunosuppressed.

[0131] Methods and Uses Administration of an anti-cancer therapy comprising an anti-clusterin antibody or an antigen-binding fragment thereof promotes the infiltration of tumor cells in the tumor microenvironment. Tumor-infiltrating lymphocytes are isolated from primary tumors or tumor metastases and expanded in vitro. Preparations of tumor-infiltrating lymphocytes can be used for adoptive cell therapy.

[0132] Thus, the present disclosure provides a method of treating a subject having cancer, comprising administering to the subject an anti-cancer therapy comprising an anti-clusterin antibody or an antigen-binding fragment thereof, isolating and expanding tumor-infiltrating lymphocytes (TILs) from the subject's tumor, and re-injecting the preparation of TILs into the subject.

[0133] The present disclosure also provides a method of treating cancer with tumor-infiltrating lymphocytes (TILs) isolated and expanded from a tumor isolated from a subject treated with an anti-cancer therapy comprising an anti-clusterin antibody or an antigen-binding fragment thereof.

[0134] Thus, TILs can be isolated from a subject who has undergone previous treatment with at least one anti-clusterin antibody or antigen-binding fragment thereof.

[0135] In some examples, the anti-cancer treatment is administered at least 2 weeks prior to TIL isolation. In other examples, the anti-cancer treatment is administered at least 3 weeks prior to TIL isolation. In yet other examples, the anti-cancer treatment is administered at least 4 weeks prior to TIL isolation. In further examples, the anti-cancer treatment is administered at least 5 weeks prior to TIL isolation. In yet further examples, the anti-cancer treatment is administered at least 6 weeks prior to TIL isolation.

[0136] In some embodiments, the anti-cancer treatment is a combination therapy comprising an anti-clusterin antibody or antigen-binding fragment thereof and docetaxel. The anti-cancer treatment may be administered as a treatment cycle consisting of administering the anti-clusterin antibody or antigen-binding fragment thereof once a week and docetaxel once every three weeks.

[0137] In an exemplary embodiment, the anti-cancer therapy is administered for at least one cycle of treatment. In another exemplary embodiment, the anti-cancer therapy is administered for at least two cycles of treatment. In yet another exemplary embodiment, the anti-cancer therapy is administered for more than two cycles of treatment.

[0138] The methods of the disclosure may also include administering an anti-cancer treatment comprising an anti-clusterin antibody or antigen-binding fragment thereof following the adoptive cell therapy.

[0139] In some embodiments, the anti-cancer treatment may be administered at least one week after the adoptive cell therapy. In another embodiment, the anti-cancer treatment may be administered at least two weeks after the adoptive cell therapy. In yet another embodiment, the anti-cancer treatment may be administered at least three weeks after the adoptive cell therapy. In a further embodiment, the anti-cancer treatment may be administered at least four weeks after the adoptive cell therapy.

[0140] In some embodiments, the subsequent anti-cancer therapy is a combination therapy comprising an anti-clusterin antibody or antigen-binding fragment thereof and docetaxel. The subsequent anti-cancer therapy may be administered as a treatment cycle consisting of administering the anti-clusterin antibody or antigen-binding fragment thereof once a week and docetaxel once every three weeks.

[0141] In an exemplary embodiment, the subsequent anti-cancer therapy is administered for at least one cycle of treatment. In another exemplary embodiment, the subsequent anti-cancer therapy is administered for at least two cycles of treatment. In yet another exemplary embodiment, the subsequent anti-cancer therapy is administered for more than two cycles of treatment.

[0142] In some embodiments, TILs can be obtained by methods known to those of skill in the art.

[0143] In some embodiments, TILs are isolated and expanded by in vitro or ex vivo tumor infiltrating lymphocyte generation methods.

[0144] TILs are usually treated by a method that includes an initial culture phase and an expansion phase.

[0145] The initial culture phase may be performed by culturing tumor digests and / or tumor fragments, typically in 24-well plates. During the initial culture phase, the TILs may become suspended in the cell culture medium and the tumor cells may become attached to the cell culture plate.

[0146] Tumor fragments can originate from primary tumors or tumor metastases obtained from subjects treated with the anti-cancer therapies disclosed herein.

[0147] The initial culture phase involves culturing the TILs in the presence of tumor cells. In some embodiments, each fragment is cultured separately to obtain a separate TIL culture.

[0148] For the initial culture phase, TIL cultures may be provided with cytokines, exemplary embodiments of which include IL-2 (recombinant human IL-2), IL-7 (recombinant human IL-7), IL-15 (recombinant human IL-15), and combinations thereof.

[0149] The initial culture phase is typically carried out for a period ranging from 2 to 5 weeks. In some instances, the initial culture phase is carried out for at least 2 weeks. In other instances, the initial culture phase is carried out for at least 3 weeks. In yet other instances, the initial culture phase is carried out for at least 4 weeks. In further instances, the initial culture phase is carried out for greater than at least 4 weeks.

[0150] Each TIL culture may be tested during or at the end of the initial culture phase to identify the culture with the desired anti-tumor activity. Alternatively, each TIL culture may be tested during or at the end of the initial culture phase to identify the culture with the highest percentage of lymphocytes. In some cases, T lymphocytes may be identified by cytometry using markers such as, but not limited to, CD3, CD45, or a combination thereof. In some cases, the TIL culture with the highest percentage of cytotoxic lymphocytes may be selected.

[0151] The antitumor activity of a given TIL culture can be evaluated, for example, by the INFγ level secreted in the presence of tumor cells. More specifically, an increase in INFγ secretion in the presence of tumor cells compared to baseline INFγ secretion can indicate the potential antitumor activity of a given TIL culture. In yet another example, the antitumor activity of a given TIL culture can be determined by the expression of an activation marker. An exemplary embodiment of an activation marker is CD37. The expression of an activation marker can be determined, for example, by cytometry. Other methods of testing antitumor activity can also be used.

[0152] TILs that exhibit anti-tumor activity are specifically contemplated for administration to a subject.

[0153] In some embodiments, TIL cultures that show evidence of INFγ secretion or increased INFγ secretion when co-cultured with tumor cells compared to baseline may be selected for the expansion phase.

[0154] In an exemplary embodiment, IFNγ secretion levels equal to or greater than 100 pg / ml are specifically contemplated for selection into the expansion phase.

[0155] In other exemplary embodiments, IFNγ secretion levels equal to or greater than 300 pg / ml (mid-level) are specifically contemplated for selection into the expansion phase.

[0156] In yet another exemplary embodiment, IFNγ secretion levels equal to or greater than 500 pg / ml (high level) are specifically contemplated for selection into the expansion phase.

[0157] In some embodiments, INFγ secretion is determined after at least 2 weeks of culture. In other embodiments, INFγ secretion is determined after at least 3 weeks of culture. In other embodiments, INFγ secretion is determined after at least 4 weeks of culture.

[0158] In some embodiments, the TIL culture with the highest percentage of cytotoxic T cells may be selected for the expansion phase or for administration to a subject. For example, the TIL culture with the highest percentage of CD8 + TIL cultures having T lymphocytes are selected. In another example, at least 50% of the TILs have a CD8 + TIL cultures with T lymphocytes are selected.

[0159] TIL cultures with desirable characteristics may be pooled before or after the expansion phase, or alternatively, individual TIL cultures may be expanded.

[0160] In some embodiments, the expansion phase may involve removing tumor cells from the TIL culture or isolating immune cells from the culture. In some instances, CD8+ T cells may be specifically selected from the culture for subsequent transfer into the subject.

[0161] For the expansion phase, TIL cultures may also be fed with cytokines, if desired. Exemplary embodiments of cytokines include IL-2 (recombinant human IL-2), IL-7 (recombinant human IL-7), IL-15 (recombinant human IL-15), and combinations thereof. If desired, one or more cytokines may be omitted from the expansion phase.

[0162] The augmentation phase is typically carried out for a period ranging from 1 to 5 weeks. In some examples, the augmentation phase may be carried out for at least 1 week. In other examples, the augmentation phase may be carried out for at least 2 weeks. In yet other examples, the augmentation phase may be carried out for at least 3 weeks. In further examples, the augmentation phase may be carried out for at least 4 weeks.

[0163] The TIL preparations may be further tested for anti-tumor activity.

[0164] The methods of the disclosure can involve treating TIL cultures or preparations to improve their characteristics. Treating can occur at one or more time points throughout the initial culture phase or throughout the expansion phase.

[0165] For example, TIL cultures or preparations may be treated to remove components that may negatively affect anti-tumor activity. In another example, TIL cultures or preparations may be treated to remove components that may interfere with the growth or activity of cytotoxic lymphocytes.

[0166] In some exemplary embodiments, TIL cultures or preparations may be treated to remove TRegs.

[0167] In another exemplary embodiment, TIL cultures or preparations may be treated to deplete NKT cells.

[0168] In some embodiments, the methods may include removing tumor cells from the TIL culture or TIL preparation.

[0169] In some embodiments, the method comprises: detecting CD45 from a TIL culture or TIL preparation. + A step of selecting the cells may be included.

[0170] In some embodiments, the method comprises: + A step of selecting the cells may be included.

[0171] In some embodiments, the method comprises: + A step of selecting the cells may be included.

[0172] In some embodiments, the methods may include selecting tumor infiltrating lymphocyte cultures or TIL preparations that secrete INFγ at levels equal to or greater than 100 pg / ml.

[0173] In some embodiments, the method may include selecting tumor infiltrating lymphocyte cultures or TIL preparations that secrete INFγ at levels equal to or greater than 300 pg / ml (mid-level).

[0174] In some embodiments, the method may include selecting tumor infiltrating lymphocyte cultures or TIL preparations that secrete INFγ at levels equal to or greater than 500 pg / ml (high level).

[0175] In some embodiments, the method may include selecting a tumor infiltrating lymphocyte culture or TIL preparation comprising at least 50% CD8+ lymphocytes. In other embodiments, the method may include selecting a tumor infiltrating lymphocyte culture or TIL preparation comprising at least 60% CD8+ lymphocytes. In yet other embodiments, the method may include selecting a tumor infiltrating lymphocyte culture or TIL preparation comprising at least 70% CD8+ lymphocytes. In further embodiments, the method may include selecting a tumor infiltrating lymphocyte culture or TIL preparation comprising at least 75% CD8+ lymphocytes. In additional embodiments, the method may include selecting a tumor infiltrating lymphocyte culture or TIL preparation comprising more than 75% CD8+ lymphocytes.

[0176] In a further embodiment, the method may include selecting a tumor infiltrating lymphocyte culture or TIL preparation that contains CD8+ lymphocytes and secretes intermediate to high levels of INFγ.

[0177] Thus, in some examples, the method may include selecting a tumor infiltrating lymphocyte culture or TIL preparation that includes at least 50% CD8+ lymphocytes and secretes intermediate to high levels of INFγ. In other examples, the method may include selecting a tumor infiltrating lymphocyte culture or TIL preparation that includes at least 60% CD8+ lymphocytes and secretes intermediate to high levels of INFγ. In yet other examples, the method may include selecting a tumor infiltrating lymphocyte culture or TIL preparation that includes at least 70% CD8+ lymphocytes and secretes intermediate to high levels of IFNγ. In additional examples, the method may include selecting a tumor infiltrating lymphocyte culture or TIL preparation that includes at least 75% CD8+ lymphocytes and secretes intermediate to high levels of IFNγ. In additional examples, the method may include selecting a tumor infiltrating lymphocyte culture or TIL preparation that includes more than 75% CD8+ lymphocytes and secretes intermediate to high levels of IFNγ.

[0178] In some embodiments, the methods may include pooling tumor infiltrating lymphocyte cultures or TIL preparations that contain CD8+ lymphocytes and secrete intermediate to high levels of INFγ.

[0179] Thus, in some embodiments, the method may include pooling tumor infiltrating lymphocyte cultures each of which contains at least 50% CD8+ lymphocytes and secretes intermediate to high levels of IFNγ. In other exemplary embodiments, the method may include pooling tumor infiltrating lymphocyte cultures each of which contains at least 60% CD8+ lymphocytes and secretes intermediate to high levels of IFNγ. In additional exemplary embodiments, the method may include pooling tumor infiltrating lymphocyte cultures each of which contains at least 70% CD8+ lymphocytes and secretes intermediate to high levels of IFNγ. In yet additional exemplary embodiments, the method may include pooling tumor infiltrating lymphocyte cultures each of which contains at least 75% CD8+ lymphocytes and secretes intermediate to high levels of IFNγ. In other exemplary embodiments, the method may include pooling tumor infiltrating lymphocyte cultures each of which contains more than 75% CD8+ lymphocytes and secretes intermediate to high levels of IFNγ.

[0180] In an exemplary embodiment, the method may include selecting and / or pooling tumor infiltrating lymphocyte cultures that secrete intermediate levels of INFγ.

[0181] In an exemplary embodiment, the method may include selecting and / or pooling tumor infiltrating lymphocyte cultures that secrete high levels of INFγ.

[0182] Similarly, preparations of TILs with diverse characteristics may be pooled.

[0183] In some embodiments, the preparation of TILs is obtained from a subject described herein.

[0184] In some embodiments, the preparation of TILs is obtained from a subject who has been or will be treated with an anti-clusterin antibody, or antigen-binding fragment thereof, as a single agent.

[0185] In some embodiments, the preparation of TILs is obtained from a subject who has been or will be treated with a combination therapy comprising an anti-clusterin antibody, or antigen-binding fragment thereof, and a chemotherapeutic agent.

[0186] In some embodiments, the chemotherapeutic agent is docetaxel.

[0187] In some embodiments, the tumor is resectable.

[0188] In some embodiments, the subject has a functional immune system.

[0189] In some embodiments, the TILs are obtained from a tumor or tumor fragment isolated by biopsy.

[0190] In accordance with the present disclosure, a method for generating tumor infiltrating lymphocytes in vitro or ex vivo comprises contacting a tumor fragment with an anti-clusterin antibody or an antigen-binding fragment thereof.

[0191] In accordance with the present disclosure, anti-clusterin antibodies or antigen-binding fragments thereof may be present and / or maintained during the initial culture phase of the tumor infiltrating lymphocyte generation method.

[0192] In accordance with the present disclosure, anti-clusterin antibodies or antigen-binding fragments thereof may be present and / or maintained during the expansion phase of the tumor infiltrating lymphocyte generation method.

[0193] TILs may or may not be genetically modified. For example, TILs may express chimeric antigen receptors. The basic structure of chimeric antigen receptors has been described in the literature (e.g., Gacerez, AT et al., J Cell Physiol. 231(12):2590-2598 (2016); Sadelain, M. et al., Cancer Discovery, 3(4):388-98 (2013); Zhang, C. et al., Biomarker Research, 5:22 (2017)). Chimeric antigen receptors usually include an extracellular antigen-binding domain, typically in the form of a single-chain Fv, a transmembrane domain, a costimulatory domain, and an intracellular signaling domain.

[0194] In other examples, the TILs may express a transgenic T cell receptor.

[0195] TILs can be isolated from primary tumors or tumor metastases.

[0196] In accordance with the present disclosure, anti-clusterin antibodies or antigen-binding fragments thereof may be administered at a dose and / or dosing interval and / or treatment duration sufficient to result in infiltration of immune cells in the tumor microenvironment.

[0197] In accordance with the present disclosure, docetaxel may be administered at a dose and / or dosing interval and / or treatment duration sufficient to allow chemotherapy-induced immunogenic modulation of the tumor.

[0198] In some embodiments, the anti-clusterin antibody or antigen-binding fragment thereof is as disclosed herein. For example, in some embodiments, the anti-clusterin antibody or antigen-binding fragment thereof is humanized 16B5.

[0199] In some embodiments, the anti-clusterin antibody or antigen-binding fragment thereof is administered prior to isolation of the TILs. In some embodiments, the anti-clusterin antibody or antigen-binding fragment thereof and the chemotherapeutic agent are administered prior to isolation of the TILs. In some embodiments, one or more treatment cycles are administered prior to isolation of the TILs.

[0200] In some embodiments, the anti-clusterin antibody or antigen-binding fragment thereof is administered after infusion of the TILs. In some embodiments, the anti-clusterin antibody or antigen-binding fragment thereof and the chemotherapeutic agent are administered after infusion of the TILs. In some embodiments, one or more treatment cycles are administered after infusion of the TILs.

[0201] In some embodiments, the preparation of TILs is CD3 + Includes T cells.

[0202] In some embodiments, the preparation of TILs is CD4 + Includes T cells.

[0203] In some embodiments, the preparation of TILs is CD8 + Includes T cells.

[0204] In some embodiments, the preparation of TILs comprises B cells.

[0205] In some embodiments, the preparation of TILs comprises NK cells.

[0206] In some embodiments, the preparation of TILs comprises NK T cells.

[0207] In some embodiments, the preparation of TILs is selected for tumor antigen recognition.

[0208] In accordance with the present disclosure, anti-clusterin antibodies or antigen-binding fragments thereof may be administered in the dosages, regimens, and / or schedules disclosed herein.

[0209] In accordance with the present disclosure, docetaxel may be administered in the dosages, regimens, and / or schedules disclosed herein.

[0210] In accordance with the present disclosure, the combination of an anti-clusterin antibody or antigen-binding fragment thereof and docetaxel may be administered in the dosages, regimens, and / or schedules disclosed herein.

[0211] In accordance with the present disclosure, the subject can have a carcinoma, for example a metastatic carcinoma.

[0212] In yet another aspect, the present disclosure provides a method of treating a subject having cancer, comprising administering, as a single agent or as a combination therapy with chemotherapy, tumor-infiltrating lymphocytes (TILs) obtained by an in vitro or in vivo method comprising contacting a tumor fragment with an anti-clusterin antibody or an antigen-binding fragment thereof.

[0213] In some embodiments, the subject may have been previously treated with an anti-clusterin antibody or antigen-binding fragment thereof, or a combination therapy.

[0214] In some embodiments, the subject has not been previously treated with an anti-clusterin antibody or antigen-binding fragment thereof or a combination therapy.

[0215] According to the present disclosure, TIL is re-infused into the subject.TIL infusion protocol is described in the literature.In some embodiments, the subject undergoes lymphodepletion conditioning before infusion of TIL.In some embodiments, the subject receives IL-2.

[0216] For example, prior to infusion of the TIL product, patients were treated with cyclophosphamide (60 mg / kg / day intravenously for 2 days) and fludarabine (25 mg / m 2 / day intravenously for 5 days). Intravenous adoptive transfer of TILs may be followed by intravenous IL-2 (Proleukin) (600,000 IU / kg / dose every 8 hours until tolerated or up to a maximum of 15 doses).

[0217] Preparation of TILs and TIL cultures The present disclosure also provides a preparation of tumor infiltrating lymphocytes (TILs) obtained by the methods described herein.

[0218] The present disclosure also provides TIL cultures obtained by the methods described herein.

[0219] The terms "preparation of TILs" and "TIL preparation" are used interchangeably.

[0220] Generally, the term "preparation of TILs" is used to refer to a composition for administration in adoptive cell therapy. The term "TIL culture" generally refers to a composition that has been isolated, expanded, or is in the process of being isolated and / or expanded. A "TIL culture" may originate from a single cell clone or a mixed cell population. In some embodiments, a preparation of TILs may consist of a single TIL culture or several TIL cultures.

[0221] It should be understood that a "preparation of TILs" or a "TIL culture" may have similar or identical characteristics. In some embodiments, the preparation of TILs is a TIL culture.

[0222] In some embodiments, the preparation of TILs or TIL cultures are obtained from a subject described herein.

[0223] In some embodiments, the preparation of TILs is a preparation of expanded TILs.

[0224] Accordingly, the present disclosure also provides a preparation of expanded tumor infiltrating lymphocytes (TILs) or TIL cultures obtained by a method of treating a subject having cancer with an anti-clusterin antibody or an antigen-binding fragment thereof, and isolating and expanding tumor infiltrating lymphocytes (TILs) from the subject's tumor.

[0225] In some embodiments, preparations of TILs or TIL cultures are obtained from subjects who have been or will be treated with anti-clusterin antibodies or antigen-binding fragments thereof, either as a single agent or as combination therapy with a chemotherapeutic agent.

[0226] In some embodiments, the TILs are not genetically modified.

[0227] In some embodiments, the TILs are genetically modified.

[0228] In some embodiments, the TILs express a chimeric antigen receptor.

[0229] In some embodiments, the TILs express a transgenic T cell receptor.

[0230] In some embodiments, the TILs are provided in an infusion bag.

[0231] Preparations of tumor-infiltrating lymphocytes or TIL cultures can secrete intermediate to high levels of INFγ.

[0232] In an exemplary embodiment, preparations of tumor infiltrating lymphocytes or TIL cultures secrete INFγ at levels equal to or greater than 100 pg / ml.

[0233] In another exemplary embodiment, preparations of tumor infiltrating lymphocytes or TIL cultures secrete INFγ at levels equal to or greater than 300 pg / ml (mid-level).

[0234] In yet other exemplary embodiments, preparations of tumor infiltrating lymphocytes or TIL cultures secrete INFγ at levels equal to or greater than 500 pg / ml (high level).

[0235] In some embodiments, the preparation of tumor infiltrating lymphocytes (TIL) or TIL cultures contains a majority of CD45 + For example, in some embodiments, the TIL preparation or TIL culture comprises at least 80% CD45 + For example, in other embodiments, the TIL preparation or TIL culture may comprise at least 90% CD45 + In other embodiments, the preparation of TILs or TIL cultures may comprise at least 95% CD45 +In yet other embodiments, the preparation of TILs or TIL cultures may comprise at least 99% CD45 + In another embodiment, the preparation of TILs or TIL cultures may comprise CD45 + It may contain only cells.

[0236] In some embodiments, the preparation of tumor infiltrating lymphocytes (TILs) comprises a majority of CD4 + For example, in some embodiments, the TIL preparation or TIL culture comprises greater than 50% CD4 + In other embodiments, the TIL preparation or TIL culture may comprise at least 60% CD4 + In yet other embodiments, the TIL preparation or TIL culture may comprise at least 70% CD4 + In some embodiments, the TIL preparation or TIL culture may comprise at least 80% CD4 + In a further embodiment, the preparation of TILs or TIL cultures may comprise at least 90% CD4 + In other embodiments, the TIL preparation or TIL culture may comprise at least 95% CD4 + In yet other embodiments, the preparation of TILs or TIL cultures may comprise at least 99% CD4 + In another embodiment, the preparation of TILs or TIL cultures may comprise CD4 + It may contain only cells.

[0237] In some embodiments, preparations of tumor infiltrating lymphocytes (TILs) or TIL cultures contain a majority of CD8 + In some examples, the preparation of tumor infiltrating lymphocytes or the TIL culture may contain at least 50% CD8+ lymphocytes. For example, in some embodiments, the preparation of tumor infiltrating lymphocytes or the TIL culture may contain more than 50% CD8 + In other embodiments, the preparation of tumor infiltrating lymphocytes or the TIL culture may comprise at least 60% CD8 +In yet other embodiments, the preparation of tumor infiltrating lymphocytes or TIL cultures may comprise at least 70% CD8 + In yet other embodiments, the preparation of tumor infiltrating lymphocytes or TIL cultures may comprise at least 75% CD8 + In some embodiments, the preparation of tumor infiltrating lymphocytes or the TIL culture may comprise at least 80% CD8 + In a further embodiment, the preparation of tumor infiltrating lymphocytes or the TIL culture may comprise at least 90% CD8 + In other embodiments, the preparation of tumor infiltrating lymphocytes or the TIL culture may comprise at least 95% CD8 + In yet other embodiments, the preparation of tumor infiltrating lymphocytes or TIL cultures may comprise at least 99% CD8 + In another embodiment, the preparation of tumor infiltrating lymphocytes or the TIL culture may comprise CD8 + In some examples, the CD8+ cells are CD8+ T lymphocytes.

[0238] In some embodiments, preparations of tumor infiltrating lymphocytes or TIL cultures may contain CD8+ lymphocytes and may secrete intermediate to high levels of INFγ.

[0239] In some embodiments, a preparation of tumor infiltrating lymphocytes may consist of tumor infiltrating lymphocyte cultures, each of which contains CD8+ lymphocytes and secretes intermediate to high levels of IFNγ.

[0240] In an exemplary embodiment, the preparation of tumor infiltrating lymphocytes may consist of tumor infiltrating lymphocyte cultures each comprising at least 50% CD8+ lymphocytes. In another exemplary embodiment, the preparation of tumor infiltrating lymphocytes may consist of tumor infiltrating lymphocyte cultures each comprising at least 50% CD8+ lymphocytes and secreting intermediate to high levels of IFNγ. In another exemplary embodiment, the preparation of tumor infiltrating lymphocytes may consist of tumor infiltrating lymphocyte cultures each comprising at least 60% CD8+ lymphocytes and secreting intermediate to high levels of IFNγ. In an additional exemplary embodiment, the preparation of tumor infiltrating lymphocytes may consist of tumor infiltrating lymphocyte cultures each comprising at least 70% CD8+ lymphocytes and secreting intermediate to high levels of IFNγ. In yet an additional exemplary embodiment, the preparation of tumor infiltrating lymphocytes may consist of tumor infiltrating lymphocyte cultures each comprising at least 75% CD8+ lymphocytes and secreting intermediate to high levels of IFNγ. In yet further exemplary embodiments, the preparation of tumor infiltrating lymphocytes is comprised of tumor infiltrating lymphocyte cultures each of which comprises at least 80% CD8+ lymphocytes and secretes intermediate to high levels of IFNγ. In yet further exemplary embodiments, the preparation of tumor infiltrating lymphocytes is comprised of tumor infiltrating lymphocyte cultures each of which comprises at least 85% CD8+ lymphocytes and secretes intermediate to high levels of IFNγ. In yet further exemplary embodiments, the preparation of tumor infiltrating lymphocytes is comprised of tumor infiltrating lymphocyte cultures each of which comprises at least 90% CD8+ lymphocytes and secretes intermediate to high levels of IFNγ. In yet further exemplary embodiments, the preparation of tumor infiltrating lymphocytes is comprised of tumor infiltrating lymphocyte cultures each of which comprises at least 95% CD8+ lymphocytes and secretes intermediate to high levels of IFNγ.

[0241] In some embodiments, the preparation of tumor infiltrating lymphocytes (TIL) or TIL cultures contains a majority of CD4 + or CD8 + For example, in some embodiments, the preparation of TILs or TIL cultures include cells that are CD4 + or CD8 +In another embodiment, the TIL preparation or TIL culture may comprise more than 50% cells that are CD4 + or CD8 + In yet other embodiments, the TIL preparation or TIL culture may comprise at least 60% cells that are CD4 + or CD8 + In some embodiments, the TIL preparation or TIL culture may comprise at least 70% cells that are CD4 + or CD8 + In a further embodiment, the TIL preparation or TIL culture may comprise at least 80% cells that are CD4 + or CD8 + In another embodiment, the TIL preparation or TIL culture may comprise at least 90% cells that are CD4 + or CD8 + In another embodiment, the TIL preparation or TIL culture may comprise at least 95% cells that are CD4 + or CD8 + In another embodiment, the TIL preparation or TIL culture may comprise at least 99% cells that are CD4 + or CD8 + The antibody may contain only cells that are

[0242] In other examples, the tumor infiltrating lymphocyte preparation or TIL culture may contain less than 10% CD4+ lymphocytes. In yet other examples, the tumor infiltrating lymphocyte preparation or TIL culture may contain less than 7.5% CD4+ lymphocytes. In other examples, the tumor infiltrating lymphocyte preparation or TIL culture may contain less than 5% CD4+ lymphocytes. In other examples, the tumor infiltrating lymphocyte preparation or TIL culture may contain 2% CD4+ lymphocytes or fewer.

[0243] In an exemplary embodiment, the preparation of TILs or TIL cultures are characterized by an INFγ secretion level equal to or greater than 100 pg / ml.

[0244] In another exemplary embodiment, the preparation of TILs or TIL cultures are characterized by an INFγ secretion level equal to or higher than 300 pg / ml (mid-level).

[0245] In yet another exemplary embodiment, the preparation of TILs or TIL cultures are characterized by an INFγ secretion level equal to or greater than 500 pg / ml (high level).

[0246] In some embodiments, a preparation of TILs disclosed herein is administered to a subject in need thereof. The preparation of TILs is autologous to the subject from which it was originally isolated.

[0247] In the methods of the present disclosure, a preparation of TILs is infused into a subject. Typically, the cells are 8 ~10 11 The individual is used to treat the subject. The subject may undergo lymphodepletion treatment prior to adoptive cell therapy.

[0248] Subjects may also receive high doses of IL-2. Exemplary embodiments of high dose IL-2 include 600,000 IU / kg or 720,000 IU / kg. High dose IL-2 may be provided by IV infusion every 8 hours. High dose IL-2 may be provided in up to 15 consecutive doses. The consecutive doses may be provided, for example, over a period of 5 days.

[0249] Anti-clusterin antibody or antigen-binding fragment thereof In some embodiments, the anti-clusterin antibodies, or antigen-binding fragments thereof, of the present disclosure are capable of inhibiting epithelial-mesenchymal transition.

[0250] In some embodiments, the anti-clusterin antibody or antigen-binding fragment thereof of the present disclosure is capable of binding to amino acids 421 and 443 of the C-terminal portion of the beta subunit of human clusterin (SEQ ID NO: 41, see International Patent Application No. PCT / CA2006 / 001505, published as WO 2007 / 030930, and International Patent Application No. PCT / CA2010 / 0001882, published as WO 2011 / 063523, the entire contents of which are incorporated by reference herein).

[0251] In some embodiments, the anti-clusterin antibodies or antigen-binding fragments thereof of the present disclosure are capable of binding to an epitope contained within amino acids 421 and 443 of the C-terminal portion of the beta subunit of human clusterin (SEQ ID NO: 41; see PCT / CA2006 / 001505, published as WO 2007 / 030930, and PCT / CA2010 / 0001882, published as WO 2011 / 063523, the entire contents of which are incorporated by reference herein).

[0252] In some embodiments, the anti-clusterin antibody, or antigen-binding fragment thereof, comprises the CDRs of an anti-clusterin antibody, or antigen-binding fragment thereof, of the present disclosure.

[0253] In some embodiments, the anti-clusterin antibody or antigen-binding fragment thereof is an antibody or antigen-binding fragment thereof that is capable of competing with the anti-clusterin antibody or antigen-binding fragment thereof of the present disclosure for binding to clusterin (e.g., secreted clusterin (sCLU) or tumor-associated sCLU (TA-sCLU)), or for binding to a polypeptide comprising the amino acid sequence set forth in SEQ ID NO: 41.

[0254] In some embodiments, CDRs are identified using methods known to those of skill in the art, which are reviewed in Antibody Engineering Vol. 2, Chapter 3 by Andrew CR Martin, the entire contents of which are incorporated herein by reference.

[0255] In certain embodiments, all CDRs are identified using the Kabat definition, which is the most commonly used definition (Wu and Kabat, 1970).

[0256] In certain embodiments, all CDRs are identified using the contact definition (MacCallum et al., 1996), which is believed to be most useful for those wishing to perform mutagenesis to alter the affinity of the antibody since these are the residues involved in interactions with the antigen.

[0257] In certain embodiments, the anti-clusterin antibody or antigen-binding fragment thereof comprises a light chain variable region comprising the complementarity determining regions (CDRs) of the light chain variable region set forth in SEQ ID NO: 9, and a heavy chain variable region comprising the CDRs of the heavy chain variable region set forth in SEQ ID NO: 10.

[0258] In some exemplary embodiments, the anti-clusterin antibody or antigen-binding fragment thereof comprises a light chain variable region comprising CDRL1 having the amino acid sequence set forth in SEQ ID NO:1, CDRL2 having the amino acid sequence set forth in SEQ ID NO:2, and CDRL3 having the amino acid sequence set forth in SEQ ID NO:3.

[0259] In some exemplary embodiments, the anti-clusterin antibody or antigen-binding fragment thereof comprises a heavy chain variable region comprising a CDRH1 having the amino acid sequence set forth in SEQ ID NO:4, a CDRH2 having the amino acid sequence set forth in SEQ ID NO:5, and a CDRH3 having the amino acid sequence set forth in SEQ ID NO:6.

[0260] In some exemplary embodiments, the anti-clusterin antibody or antigen-binding fragment thereof comprises a heavy chain variable region comprising a CDRH1 having the amino acid sequence set forth in SEQ ID NO:35, a CDRH2 having the amino acid sequence set forth in SEQ ID NO:36, and a CDRH3 having the amino acid sequence set forth in SEQ ID NO:37.

[0261] In some embodiments, the anti-clusterin antibody or antigen-binding fragment thereof comprises a light chain variable region comprising CDRL1 having the amino acid sequence set forth in SEQ ID NO:1, CDRL2 having the amino acid sequence set forth in SEQ ID NO:2, and CDRL3 having the amino acid sequence set forth in SEQ ID NO:3, and a heavy chain variable region comprising CDRH1 having the amino acid sequence set forth in SEQ ID NO:4, CDRH2 having the amino acid sequence set forth in SEQ ID NO:5, and CDRH3 having the amino acid sequence set forth in SEQ ID NO:6.

[0262] In some embodiments, the anti-clusterin antibody or antigen-binding fragment thereof comprises a light chain variable region comprising CDRL1 having the amino acid sequence set forth in SEQ ID NO:1, CDRL2 having the amino acid sequence set forth in SEQ ID NO:2, and CDRL3 having the amino acid sequence set forth in SEQ ID NO:3, and a heavy chain variable region comprising CDRH1 having the amino acid sequence set forth in SEQ ID NO:35, CDRH2 having the amino acid sequence set forth in SEQ ID NO:36, and CDRH3 having the amino acid sequence set forth in SEQ ID NO:37.

[0263] In some embodiments, the anti-clusterin antibody or antigen-binding fragment thereof comprises a light chain variable region having an amino acid sequence that is at least 80% identical to the amino acid sequence set forth in SEQ ID NO:7, and a heavy chain variable region having an amino acid sequence that is at least 80% identical to the amino acid sequence set forth in SEQ ID NO:8.

[0264] In some embodiments, the anti-clusterin antibody or antigen-binding fragment thereof comprises a light chain variable region having an amino acid sequence that is at least 90% identical to the amino acid sequence set forth in SEQ ID NO:7, and a heavy chain variable region having an amino acid sequence that is at least 90% identical to the amino acid sequence set forth in SEQ ID NO:8.

[0265] In some embodiments, the anti-clusterin antibody or antigen-binding fragment thereof comprises a light chain variable region having an amino acid sequence identical to the amino acid sequence set forth in SEQ ID NO:7, and a heavy chain variable region having an amino acid sequence identical to the amino acid sequence set forth in SEQ ID NO:8.

[0266] In some embodiments, the anti-clusterin antibody or antigen-binding fragment thereof is capable of competing with an antibody comprising a light chain variable region having the amino acid sequence set forth in SEQ ID NO:7 and a heavy chain variable region having the amino acid sequence set forth in SEQ ID NO:8 for binding to clusterin (e.g., secreted clusterin (sCLU) or tumor-associated sCLU (TA-sCLU)), or for binding to a polypeptide comprising the amino acid sequence set forth in SEQ ID NO:41.

[0267] In some embodiments, the anti-clusterin antibody or antigen-binding fragment thereof comprises a light chain variable region having an amino acid sequence that is at least 80% identical to the amino acid sequence set forth in SEQ ID NO:9, and a heavy chain variable region having an amino acid sequence that is at least 80% identical to the amino acid sequence set forth in SEQ ID NO:10.

[0268] In some embodiments, the anti-clusterin antibody or antigen-binding fragment thereof comprises a light chain variable region having an amino acid sequence that is at least 90% identical to the amino acid sequence set forth in SEQ ID NO:9, and a heavy chain variable region having an amino acid sequence that is at least 90% identical to the amino acid sequence set forth in SEQ ID NO:10.

[0269] In some embodiments, the anti-clusterin antibody or antigen-binding fragment thereof comprises a light chain variable region having an amino acid sequence identical to the amino acid sequence set forth in SEQ ID NO:9, and a heavy chain variable region having an amino acid sequence identical to the amino acid sequence set forth in SEQ ID NO:10.

[0270] In some embodiments, the anti-clusterin antibody or antigen-binding fragment thereof is capable of competing with an antibody comprising a light chain variable region having the amino acid sequence set forth in SEQ ID NO:9 and a heavy chain variable region having the amino acid sequence set forth in SEQ ID NO:10 for binding to clusterin (e.g., secreted clusterin (sCLU) or tumor-associated sCLU (TA-sCLU)), or for binding to a polypeptide comprising the amino acid sequence set forth in SEQ ID NO:41.

[0271] In some embodiments, the anti-clusterin antibody or antigen-binding fragment thereof comprises a light chain having an amino acid sequence at least 80% identical to the amino acid sequence set forth in SEQ ID NO:11, and a heavy chain having an amino acid sequence at least 80% identical to the amino acid sequence set forth in SEQ ID NO:12.

[0272] In some embodiments, the anti-clusterin antibody or antigen-binding fragment thereof comprises a light chain having an amino acid sequence at least 90% identical to the amino acid sequence set forth in SEQ ID NO:11 and a heavy chain having an amino acid sequence at least 90% identical to the amino acid sequence set forth in SEQ ID NO:12.

[0273] In some embodiments, the anti-clusterin antibody or antigen-binding fragment thereof comprises a light chain having an amino acid sequence identical to the amino acid sequence set forth in SEQ ID NO:11, and a heavy chain having an amino acid sequence identical to the amino acid sequence set forth in SEQ ID NO:12.

[0274] In some embodiments, the anti-clusterin antibody or antigen-binding fragment thereof is capable of competing with an antibody comprising a light chain having the amino acid sequence set forth in SEQ ID NO:11 and a heavy chain having the amino acid sequence set forth in SEQ ID NO:12 for binding to clusterin (e.g., secreted clusterin (sCLU) or tumor-associated sCLU (TA-sCLU)), or for binding to a polypeptide comprising the amino acid sequence set forth in SEQ ID NO:41.

[0275] In another specific embodiment, the anti-clusterin antibody or antigen-binding fragment thereof comprises a light chain variable region comprising CDRL1 having the amino acid sequence set forth in SEQ ID NO: 15, CDRL2 having the amino acid sequence set forth in SEQ ID NO: 16, and CDRL3 having the amino acid sequence set forth in SEQ ID NO: 17.

[0276] In some exemplary embodiments, the anti-clusterin antibody or antigen-binding fragment thereof comprises a heavy chain variable region comprising CDRH1 having the amino acid sequence set forth in SEQ ID NO: 18, CDRH2 having the amino acid sequence set forth in SEQ ID NO: 19, and CDRH3 having the amino acid sequence set forth in SEQ ID NO: 20.

[0277] In some exemplary embodiments, the anti-clusterin antibody or antigen-binding fragment thereof comprises a heavy chain variable region comprising CDRH1 having the amino acid sequence set forth in SEQ ID NO:38, CDRH2 having the amino acid sequence set forth in SEQ ID NO:39, and CDRH3 having the amino acid sequence set forth in SEQ ID NO:40.

[0278] In some embodiments, the anti-clusterin antibody or antigen-binding fragment thereof comprises a light chain variable region comprising CDRL1 having the amino acid sequence set forth in SEQ ID NO: 15, CDRL2 having the amino acid sequence set forth in SEQ ID NO: 16, and CDRL3 having the amino acid sequence set forth in SEQ ID NO: 17, and a heavy chain variable region comprising CDRH1 having the amino acid sequence set forth in SEQ ID NO: 18, CDRH2 having the amino acid sequence set forth in SEQ ID NO: 19, and CDRH3 having the amino acid sequence set forth in SEQ ID NO: 20.

[0279] In some embodiments, the anti-clusterin antibody or antigen-binding fragment thereof comprises a light chain variable region comprising CDRL1 having the amino acid sequence set forth in SEQ ID NO: 15, CDRL2 having the amino acid sequence set forth in SEQ ID NO: 16, and CDRL3 having the amino acid sequence set forth in SEQ ID NO: 17, and a heavy chain variable region comprising CDRH1 having the amino acid sequence set forth in SEQ ID NO: 38, CDRH2 having the amino acid sequence set forth in SEQ ID NO: 39, and CDRH3 having the amino acid sequence set forth in SEQ ID NO: 40.

[0280] In some embodiments, the anti-clusterin antibody or antigen-binding fragment thereof comprises a light chain variable region having an amino acid sequence that is at least 80% identical to the amino acid sequence set forth in SEQ ID NO:21, and a heavy chain variable region having an amino acid sequence that is at least 80% identical to the amino acid sequence set forth in SEQ ID NO:22.

[0281] In some embodiments, the anti-clusterin antibody or antigen-binding fragment thereof comprises a light chain variable region having an amino acid sequence at least 90% identical to the amino acid sequence set forth in SEQ ID NO:21, and a heavy chain variable region having an amino acid sequence at least 90% identical to the amino acid sequence set forth in SEQ ID NO:22.

[0282] In some embodiments, the anti-clusterin antibody or antigen-binding fragment thereof comprises a light chain variable region having an amino acid sequence identical to the amino acid sequence set forth in SEQ ID NO:21, and a heavy chain variable region having an amino acid sequence identical to the amino acid sequence set forth in SEQ ID NO:22.

[0283] In some embodiments, the anti-clusterin antibody or antigen-binding fragment thereof is capable of competing with an antibody comprising a light chain variable region having the amino acid sequence set forth in SEQ ID NO:21 and a heavy chain variable region having the amino acid sequence set forth in SEQ ID NO:22 for binding to clusterin (e.g., secreted clusterin (sCLU) or tumor-associated sCLU (TA-sCLU)), or for binding to a polypeptide comprising the amino acid sequence set forth in SEQ ID NO:41.

[0284] In some embodiments, the anti-clusterin antibody or antigen-binding fragment thereof comprises a light chain variable region having an amino acid sequence at least 80% identical to the amino acid sequence set forth in SEQ ID NO:23, and a heavy chain variable region having at least 80% identity to the amino acid sequence set forth in SEQ ID NO:24.

[0285] In some embodiments, the anti-clusterin antibody or antigen-binding fragment thereof comprises a light chain variable region having an amino acid sequence at least 90% identical to the amino acid sequence set forth in SEQ ID NO:23, and a heavy chain variable region having an amino acid sequence at least 90% identical to the amino acid sequence set forth in SEQ ID NO:24.

[0286] In some embodiments, the anti-clusterin antibody or antigen-binding fragment thereof comprises a light chain variable region having an amino acid sequence identical to the amino acid sequence set forth in SEQ ID NO:23, and a heavy chain variable region having an amino acid sequence identical to the amino acid sequence set forth in SEQ ID NO:24.

[0287] In some embodiments, the anti-clusterin antibody or antigen-binding fragment thereof is capable of competing with an antibody comprising a light chain variable region having the amino acid sequence set forth in SEQ ID NO: 23 and a heavy chain variable region having the amino acid sequence set forth in SEQ ID NO: 24 for binding to clusterin (e.g., secreted clusterin (sCLU) or tumor-associated sCLU (TA-sCLU)), or for binding to a polypeptide comprising the amino acid sequence set forth in SEQ ID NO: 41.

[0288] In some embodiments, the anti-clusterin antibody or antigen-binding fragment thereof comprises a light chain having an amino acid sequence at least 80% identical to the amino acid sequence set forth in SEQ ID NO:25, and a heavy chain having at least 80% identity to the amino acid sequence set forth in SEQ ID NO:26.

[0289] The anti-clusterin antibody or antigen-binding fragment thereof comprises a light chain having an amino acid sequence at least 90% identical to the amino acid sequence set forth in SEQ ID NO:25, and a heavy chain having at least 90% identical to the amino acid sequence set forth in SEQ ID NO:26.

[0290] In some embodiments, the anti-clusterin antibody or antigen-binding fragment thereof comprises a light chain having an amino acid sequence identical to the amino acid sequence set forth in SEQ ID NO:25, and a heavy chain having an amino acid sequence identical to the amino acid sequence set forth in SEQ ID NO:26.

[0291] In some embodiments, the anti-clusterin antibody or antigen-binding fragment thereof is capable of competing with an antibody comprising a light chain having the amino acid sequence set forth in SEQ ID NO: 25 and a heavy chain having the amino acid sequence set forth in SEQ ID NO: 26 for binding to clusterin (e.g., secreted clusterin (sCLU) or tumor-associated sCLU (TA-sCLU)), or for binding to a polypeptide comprising the amino acid sequence set forth in SEQ ID NO: 41.

[0292] In yet other specific embodiments, the anti-clusterin antibody or antigen-binding fragment thereof comprises the amino acid sequence of the CDRs, variable regions, or entire chains of the antibodies or antigen-binding fragments thereof set forth in Table 5. The amino acid sequences of the antibodies identified as 16B5, 21B12, 20E11, 11E2, and 16C11 are disclosed in International Patent Application No. PCT / CA2006 / 001505, filed Sep. 13, 2006, and published Mar. 22, 2007 as WO 2007 / 030930, the entire contents of which are incorporated herein by reference. Murine 16B5, humanized 16B5, murine 21B12, and humanized 21B12 are disclosed in International Patent Application No. PCT / CA2010 / 001882, filed November 24, 2010, and published as WO 2011 / 063523 on June 3, 2011, the entire contents of which are incorporated herein by reference.

[0293] In even more specific embodiments, the anti-clusterin antibody or antigen-binding fragment thereof may be capable of competing with one or more of the antibodies or antigen-binding fragments thereof set forth in Table 5.

[0294] subject In some aspects and embodiments of the present disclosure, the subject is a human subject.

[0295] In some aspects and embodiments of the present disclosure, the subject is a subject with cancer.

[0296] In other aspects and embodiments of the present disclosure, the subject has cancer and has a functional immune system.

[0297] In some embodiments, the subject has carcinoma.

[0298] In some embodiments, the subject has endometrial cancer, breast cancer, liver cancer, prostate cancer, kidney cancer, bladder cancer, cervical cancer, ovarian cancer, colorectal cancer, pancreatic cancer, lung cancer, gastric cancer, head and neck cancer, thyroid cancer, cholangiocarcinoma, mesothelioma, or melanoma.

[0299] In some embodiments, the subject has metastatic cancer.

[0300] In some embodiments, the subject has metastatic endometrial cancer, metastatic breast cancer, metastatic liver cancer, metastatic prostate cancer, metastatic kidney cancer, metastatic bladder cancer, metastatic cervical cancer, metastatic ovarian cancer, metastatic colorectal cancer, metastatic pancreatic cancer, metastatic lung cancer, metastatic gastric cancer, metastatic head and neck cancer, metastatic thyroid cancer, metastatic cholangiocarcinoma, metastatic mesothelioma, or metastatic melanoma.

[0301] In some embodiments, the subject has non-small cell lung cancer (NSCLC).

[0302] In some embodiments, the subject has metastatic NSCLC.

[0303] In some embodiments, the subject has stage III-IV NSCLC.

[0304] In some embodiments, the subject has breast cancer.

[0305] In some embodiments, the subject has metastatic breast cancer.

[0306] In some embodiments, the subject has prostate cancer.

[0307] In some embodiments, the subject has metastatic prostate cancer.

[0308] In some embodiments, the subject has gastric cancer.

[0309] In some embodiments, the subject has metastatic gastric cancer.

[0310] In some embodiments, the subject has head and neck cancer.

[0311] In some embodiments, the subject has metastatic head and neck cancer.

[0312] In some embodiments, the subject has thyroid cancer.

[0313] In some embodiments, the subject has metastatic thyroid cancer.

[0314] In some embodiments, the subject has ovarian cancer.

[0315] In some embodiments, the subject has metastatic ovarian cancer.

[0316] In some embodiments, the subject has endometrial cancer.

[0317] In some embodiments, the subject has metastatic endometrial cancer.

[0318] In some embodiments, the subject has liver cancer.

[0319] In some embodiments, the subject has metastatic liver cancer.

[0320] In some embodiments, the subject has colorectal cancer.

[0321] In some embodiments, the subject has metastatic colorectal cancer.

[0322] In some embodiments, the subject has pancreatic cancer.

[0323] In some embodiments, the subject has metastatic pancreatic cancer.

[0324] In some embodiments, the subject has cholangiocarcinoma.

[0325] In some embodiments, the subject has metastatic cholangiocarcinoma.

[0326] In some embodiments, the subject has mesothelioma.

[0327] In some embodiments, the subject has metastatic mesothelioma.

[0328] In some embodiments, the subject has melanoma.

[0329] In some embodiments, the subject has metastatic melanoma.

[0330] In some embodiments, the subject has, or is selected for having, a tumor characterized as immunologically cold.

[0331] In some embodiments, the subject has, or is selected for having, a tumor characterized as immunologically warm or hot that is non-responsive to immunotherapy.

[0332] In some embodiments, the subject has, or is selected for having, a tumor that exhibits signs of an epithelial-mesenchymal transition (EMT) signature.

[0333] As used herein, the term "tumor" refers to a primary tumor or a tumor metastasis or lesion.

[0334] In some embodiments, the subject has, or is selected for having, a carcinoma that has progressed after first-line immune checkpoint therapy.

[0335] In some embodiments, the subject has or is selected for having a carcinoma that has failed previous treatment with immune checkpoint therapy and a platinum-containing doublet treatment.

[0336] In some embodiments, the subject has or is selected for having a carcinoma that has failed previous treatment with concomitantly or sequentially administered immune checkpoint therapy and platinum-containing doublet treatment.

[0337] In some embodiments, the subject has, or is selected for having, a carcinoma that has failed previous treatment with an anti-PD1 or PDL-1 immune checkpoint antibody and a platinum-containing doublet treatment.

[0338] In some embodiments, the subject has, or is selected for having, a carcinoma that has failed previous treatment with ipilimumab, nivolumab, pembrolizumab, cemiplimab, atezolizumab, avelumab, or durvalumab and a platinum-containing doublet treatment.

[0339] In some embodiments, the subject has, or is selected for having, a carcinoma that has failed previous treatment with concurrent or sequential anti-PD1 or PDL-1 immune checkpoint antibody and platinum-containing doublet treatment.

[0340] In some embodiments, the subject is not immunosuppressed.

[0341] In some embodiments, the subject has not received an immunosuppressant medication within 14, 7, 6, 5, 4, 3, 2, or 1 days of treatment, hi some embodiments, the subject may have received a corticosteroid prior to treatment.

[0342] In some embodiments, the subject has not received prior treatment with docetaxel.

[0343] In some embodiments, the subject is treated with at least two cycles of treatment.

[0344] In some embodiments, subjects undergo lymphodepletion conditioning prior to infusion of the TILs.

[0345] Dosage, Treatment Regimens, and Schedules According to one aspect of the present disclosure, a subject is treated with an anti-cancer therapy comprising an anti-clusterin antibody or antigen-binding fragment thereof prior to isolation of tumor-infiltrating lymphocytes.

[0346] Thus, the anti-clusterin antibody or antigen-binding fragment thereof is administered at a dose sufficient to result in infiltration of immune cells in the tumor microenvironment.

[0347] In some embodiments, the dose of the anti-clusterin antibody or antigen-binding fragment thereof is a therapeutically effective and safe dose.

[0348] In accordance with the present disclosure, the anti-clusterin antibody or antigen-binding fragment thereof is administered at intervals sufficient to result in infiltration of immune cells in the tumor microenvironment.

[0349] In accordance with the present disclosure, the anti-clusterin antibody or antigen-binding fragment thereof is administered for a treatment period sufficient to result in infiltration of immune cells in the tumor microenvironment.

[0350] In some embodiments, the anti-clusterin antibody or antigen-binding fragment thereof is administered at a dose, dosing interval, and / or treatment duration sufficient to result in infiltration of immune cells in the tumor microenvironment.

[0351] According to another aspect of the present disclosure, the subject is treated with a combination therapy comprising an anti-clusterin antibody or antigen-binding fragment thereof and docetaxel prior to isolation of tumor-infiltrating lymphocytes.

[0352] In some embodiments, the dose of docetaxel is a therapeutically effective and safe dose.

[0353] In accordance with the present disclosure, docetaxel is administered at a dosing interval sufficient to allow chemotherapy-induced immunogenic modulation of the tumor.

[0354] In accordance with the present disclosure, docetaxel is administered for a treatment period sufficient to allow chemotherapy-induced immunogenic modulation of the tumor.

[0355] In some embodiments, docetaxel is administered at a dose and / or at a dosing interval and / or at a dosing interval sufficient to allow chemotherapy-induced immunogenic modulation of the tumor.

[0356] Thus, the anti-clusterin antibody or antigen-binding fragment thereof and docetaxel are administered at doses sufficient to result in infiltration of immune cells in the tumor microenvironment and / or to enable chemotherapy-induced immunogenic modulation of the tumor.

[0357] According to yet another aspect of the present disclosure, the subject is treated with an anti-cancer therapy comprising an anti-clusterin antibody or antigen-binding fragment thereof after re-infusion of tumor-infiltrating lymphocytes.

[0358] According to a further aspect of the present disclosure, the subject is treated with an anti-cancer therapy comprising an anti-clusterin antibody or antigen-binding fragment thereof and docetaxel after reinfusion of tumor-infiltrating lymphocytes.

[0359] According to an exemplary embodiment of the present disclosure, the anti-clusterin antibody or antigen-binding fragment thereof is administered once a week.

[0360] According to another exemplary embodiment of the present disclosure, the anti-clusterin antibody or antigen-binding fragment thereof is administered twice weekly.

[0361] According to yet another exemplary embodiment of the present disclosure, the anti-clusterin antibody or antigen-binding fragment thereof is administered three times per week.

[0362] According to a further exemplary embodiment of the present disclosure, the anti-clusterin antibody or antigen-binding fragment thereof is administered once every two weeks.

[0363] According to still further exemplary embodiments of the present disclosure, the anti-clusterin antibody or antigen-binding fragment thereof is administered once every three weeks.

[0364] According to an additional exemplary embodiment of the present disclosure, the anti-clusterin antibody or antigen-binding fragment thereof is administered once every four weeks.

[0365] In some embodiments, the anti-clusterin antibody or antigen-binding fragment thereof is administered weekly for a period of at least 2 weeks prior to isolation of the TILs. In other embodiments, the anti-clusterin antibody or antigen-binding fragment thereof is administered weekly for a period of at least 3 weeks prior to isolation of the TILs. In still other embodiments, the anti-clusterin antibody or antigen-binding fragment thereof is administered weekly for a period of at least 4 weeks prior to isolation of the TILs. In further embodiments, the anti-clusterin antibody or antigen-binding fragment thereof is administered weekly for a period of at least 5 weeks prior to isolation of the TILs. In still further embodiments, the anti-clusterin antibody or antigen-binding fragment thereof is administered weekly for a period of at least 6 weeks prior to isolation of the TILs. In accordance with the present disclosure, the anti-clusterin antibody or antigen-binding fragment thereof is administered at a dose between about 3 mg / kg and about 20 mg / kg.

[0366] In some embodiments, the anti-clusterin antibody or antigen-binding fragment thereof is administered at a dose of approximately 3.0 mg / kg.

[0367] In some embodiments, the anti-clusterin antibody or antigen-binding fragment thereof is administered at a dose of approximately 4.0 mg / kg.

[0368] In some embodiments, the anti-clusterin antibody or antigen-binding fragment thereof is administered at a dose of approximately 5.0 mg / kg.

[0369] In some embodiments, the anti-clusterin antibody or antigen-binding fragment thereof is administered at a dose of approximately 6.0 mg / kg.

[0370] In some embodiments, the anti-clusterin antibody or antigen-binding fragment thereof is administered at a dose of approximately 7.0 mg / kg.

[0371] In some embodiments, the anti-clusterin antibody or antigen-binding fragment thereof is administered at a dose of approximately 8.0 mg / kg.

[0372] In some embodiments, the anti-clusterin antibody or antigen-binding fragment thereof is administered at a dose of approximately 9.0 mg / kg.

[0373] In some embodiments, the anti-clusterin antibody or antigen-binding fragment thereof is administered at a dose of approximately 10.0 mg / kg.

[0374] In some embodiments, the anti-clusterin antibody or antigen-binding fragment thereof is administered at a dose of approximately 11.0 mg / kg.

[0375] In some embodiments, the anti-clusterin antibody or antigen-binding fragment thereof is administered at a dose of approximately 12.0 mg / kg.

[0376] In some embodiments, the anti-clusterin antibody or antigen-binding fragment thereof is administered at a dose of approximately 13.0 mg / kg.

[0377] In some embodiments, the anti-clusterin antibody or antigen-binding fragment thereof is administered at a dose of approximately 14.0 mg / kg.

[0378] In some embodiments, the anti-clusterin antibody or antigen-binding fragment thereof is administered at a dose of approximately 15.0 mg / kg.

[0379] In some embodiments, the anti-clusterin antibody or antigen-binding fragment thereof is administered at a dose of approximately 16.0 mg / kg.

[0380] In some embodiments, the anti-clusterin antibody or antigen-binding fragment thereof is administered at a dose of approximately 17.0 mg / kg.

[0381] In some embodiments, the anti-clusterin antibody or antigen-binding fragment thereof is administered at a dose of approximately 18.0 mg / kg.

[0382] In some embodiments, the anti-clusterin antibody or antigen-binding fragment thereof is administered at a dose of approximately 19.0 mg / kg.

[0383] In some embodiments, the anti-clusterin antibody or antigen-binding fragment thereof is administered at a dose of approximately 20.0 mg / kg.

[0384] In accordance with the present disclosure, the anti-clusterin antibody or antigen-binding fragment thereof is humanized 16B5 and is administered at a dose of between about 3 mg / kg and about 20 mg / kg.

[0385] In accordance with the present disclosure, humanized 16B5 is administered at a dose of between about 4 mg / kg and about 20 mg / kg.

[0386] In accordance with the present disclosure, humanized 16B5 is administered at a dose of between about 5 mg / kg and about 20 mg / kg.

[0387] In accordance with the present disclosure, humanized 16B5 is administered at a dose of between about 6 mg / kg and about 20 mg / kg.

[0388] In accordance with the present disclosure, humanized 16B5 is administered at a dose of between about 6 mg / kg and about 18 mg / kg.

[0389] In accordance with the present disclosure, humanized 16B5 is administered at a dose of between about 6 mg / kg and about 17 mg / kg.

[0390] In accordance with the present disclosure, humanized 16B5 is administered at a dose of between about 6 mg / kg and about 16 mg / kg.

[0391] In accordance with the present disclosure, humanized 16B5 is administered at a dose of between about 6 mg / kg and about 15 mg / kg.

[0392] In accordance with the present disclosure, humanized 16B5 is administered at a dose of between about 6 mg / kg and about 14 mg / kg.

[0393] In accordance with the present disclosure, humanized 16B5 is administered at a dose of between about 6 mg / kg and about 13 mg / kg.

[0394] In accordance with the present disclosure, humanized 16B5 is administered at a dose of between about 6 mg / kg and about 12 mg / kg.

[0395] In accordance with the present disclosure, humanized 16B5 is administered at a dose of between about 7 mg / kg and about 12 mg / kg.

[0396] In accordance with the present disclosure, humanized 16B5 is administered at a dose of between about 8 mg / kg and about 12 mg / kg.

[0397] In accordance with the present disclosure, humanized 16B5 is administered at a dose of between about 9 mg / kg and about 12 mg / kg.

[0398] In some embodiments, the anti-clusterin antibody, or antigen-binding fragment thereof, is humanized 16B5 and is administered at a dose of approximately 3.0 mg / kg.

[0399] In some embodiments, the anti-clusterin antibody, or antigen-binding fragment thereof, is humanized 16B5 and is administered at a dose of approximately 4.0 mg / kg.

[0400] In some embodiments, the anti-clusterin antibody, or antigen-binding fragment thereof, is humanized 16B5 and is administered at a dose of approximately 5.0 mg / kg.

[0401] In some embodiments, the anti-clusterin antibody, or antigen-binding fragment thereof, is humanized 16B5 and is administered at a dose of approximately 6.0 mg / kg.

[0402] In some embodiments, the anti-clusterin antibody, or antigen-binding fragment thereof, is humanized 16B5 and is administered at a dose of approximately 7.0 mg / kg.

[0403] In some embodiments, the anti-clusterin antibody, or antigen-binding fragment thereof, is humanized 16B5 and is administered at a dose of approximately 8.0 mg / kg.

[0404] In some embodiments, the anti-clusterin antibody, or antigen-binding fragment thereof, is humanized 16B5 and is administered at a dose of approximately 9.0 mg / kg.

[0405] In some embodiments, the anti-clusterin antibody, or antigen-binding fragment thereof, is humanized 16B5 and is administered at a dose of approximately 10.0 mg / kg.

[0406] In some embodiments, the anti-clusterin antibody, or antigen-binding fragment thereof, is humanized 16B5 and is administered at a dose of approximately 11.0 mg / kg.

[0407] In some embodiments, the anti-clusterin antibody, or antigen-binding fragment thereof, is humanized 16B5 and is administered at a dose of approximately 12.0 mg / kg.

[0408] In some embodiments, the anti-clusterin antibody, or antigen-binding fragment thereof, is humanized 16B5 and is administered at a dose of approximately 13.0 mg / kg.

[0409] In some embodiments, the anti-clusterin antibody, or antigen-binding fragment thereof, is humanized 16B5 and is administered at a dose of approximately 14.0 mg / kg.

[0410] In some embodiments, the anti-clusterin antibody, or antigen-binding fragment thereof, is humanized 16B5 and is administered at a dose of approximately 15.0 mg / kg.

[0411] In some embodiments, the anti-clusterin antibody, or antigen-binding fragment thereof, is humanized 16B5 and is administered at a dose of approximately 16.0 mg / kg.

[0412] In some embodiments, the anti-clusterin antibody, or antigen-binding fragment thereof, is humanized 16B5 and is administered at a dose of approximately 17.0 mg / kg.

[0413] In some embodiments, the anti-clusterin antibody, or antigen-binding fragment thereof, is humanized 16B5 and is administered at a dose of approximately 18.0 mg / kg.

[0414] In some embodiments, the anti-clusterin antibody, or antigen-binding fragment thereof, is humanized 16B5 and is administered at a dose of approximately 19.0 mg / kg.

[0415] In some embodiments, the anti-clusterin antibody, or antigen-binding fragment thereof, is humanized 16B5 and is administered at a dose of approximately 20.0 mg / kg.

[0416] According to an exemplary embodiment of the present disclosure, docetaxel is administered once per week.

[0417] According to another exemplary embodiment of the present disclosure, docetaxel is administered once every two weeks.

[0418] According to yet another exemplary embodiment of the present disclosure, docetaxel is administered once every three weeks.

[0419] According to a further exemplary embodiment of the present disclosure, docetaxel is administered once every four weeks.

[0420] According to a further exemplary embodiment of the present disclosure, docetaxel is administered once every five weeks.

[0421] According to a further exemplary embodiment of the present disclosure, docetaxel is administered once every six weeks.

[0422] In accordance with the present disclosure, docetaxel is administered at approximately 60 mg / m 2 ~ approx. 100 mg / m 2 is administered at a dose of

[0423] In accordance with the present disclosure, docetaxel is administered at approximately 60 mg / m 2 ~ approx. 95 mg / m 2 It is administered in doses between 100 and 200 mg / kg.

[0424] In accordance with the present disclosure, docetaxel is administered at approximately 60 mg / m 2 ~ approx. 90 mg / m 2 It is administered in doses between 100 and 200 mg / kg.

[0425] In accordance with the present disclosure, docetaxel is administered at approximately 60 mg / m 2 ~ approx. 85 mg / m 2 It is administered in doses between 100 and 200 mg / kg.

[0426] In accordance with the present disclosure, docetaxel is administered at approximately 60 mg / m 2 ~ approx. 80 mg / m 2 It is administered in doses between 100 and 200 mg / kg.

[0427] In accordance with the present disclosure, docetaxel is administered at approximately 60 mg / m 2 ~ approx. 75 mg / m 2 It is administered in doses between 100 and 200 mg / kg.

[0428] In accordance with the present disclosure, docetaxel is administered at approximately 70 mg / m 2 ~ approx. 75 mg / m 2 It is administered in doses between 100 and 200 mg / kg.

[0429] In some embodiments, docetaxel is approximately 60 mg / m 2 is administered at a dose of

[0430] In some embodiments, docetaxel is approximately 65 mg / m 2 is administered at a dose of

[0431] In some embodiments, docetaxel is approximately 70 mg / m 2 is administered at a dose of

[0432] In some embodiments, docetaxel is approximately 75 mg / m 2 is administered at a dose of

[0433] In some embodiments, docetaxel is approximately 80 mg / m 2 is administered at a dose of

[0434] In some embodiments, docetaxel is approximately 85 mg / m 2 is administered at a dose of

[0435] In some embodiments, docetaxel is approximately 90 mg / m 2 is administered at a dose of

[0436] In some embodiments, docetaxel is approximately 95 mg / m 2 is administered at a dose of

[0437] In some embodiments, docetaxel is at approximately 100 mg / m 2 is administered at a dose of

[0438] In some embodiments, the anti-clusterin antibody or antigen-binding fragment thereof is administered once a week at a dose of approximately 12 mg / kg and the docetaxel is administered once a week at a dose of approximately 75 mg / m 2 It is administered once every three weeks at a dose of

[0439] In some embodiments, the anti-clusterin antibody or antigen-binding fragment thereof is administered once a week at a dose of approximately 12 mg / kg and the docetaxel is administered once a week at a dose of approximately 60 mg / m 2 It is administered once every three weeks at a dose of

[0440] In some embodiments, the anti-clusterin antibody or antigen-binding fragment thereof is administered once a week at a dose of approximately 9 mg / kg and the docetaxel is administered once a week at a dose of approximately 75 mg / m 2It is administered once every three weeks at a dose of

[0441] In some embodiments, the anti-clusterin antibody or antigen-binding fragment thereof is administered once a week at a dose of approximately 9 mg / kg and the docetaxel is administered once a week at a dose of approximately 60 mg / m 2 It is administered once every three weeks at a dose of

[0442] In some embodiments, the anti-clusterin antibody or antigen-binding fragment thereof is administered once a week at a dose of approximately 6 mg / kg and the docetaxel is administered once a week at a dose of approximately 75 mg / m 2 It is administered once every three weeks at a dose of

[0443] In some embodiments, the anti-clusterin antibody or antigen-binding fragment thereof is administered once a week at a dose of approximately 6 mg / kg and the docetaxel is administered once a week at a dose of approximately 60 mg / m 2 It is administered once every three weeks at a dose of

[0444] In some embodiments, the anti-clusterin antibody or antigen-binding fragment thereof is administered once a week at a dose of approximately 3 mg / kg and the docetaxel is administered once a week at a dose of approximately 75 mg / m 2 It is administered once every three weeks at a dose of

[0445] In some embodiments, the anti-clusterin antibody or antigen-binding fragment thereof is administered once a week at a dose of approximately 3 mg / kg and the docetaxel is administered once a week at a dose of approximately 60 mg / m 2 It is administered once every three weeks at a dose of

[0446] In some embodiments, the anti-clusterin antibody or antigen-binding fragment thereof is humanized 16B5 and is administered once weekly at a dose of 12 mg / kg, and the docetaxel is administered once weekly at a dose of 75 mg / m 2 It is administered once every three weeks at a dose of

[0447] In some embodiments, the anti-clusterin antibody or antigen-binding fragment thereof is humanized 16B5 and is administered once weekly at a dose of 12 mg / kg, and the docetaxel is administered once weekly at a dose of 60 mg / m 2It is administered once every three weeks at a dose of

[0448] In some embodiments, the anti-clusterin antibody or antigen-binding fragment thereof is humanized 16B5 and is administered once weekly at a dose of 9 mg / kg, and the docetaxel is administered once weekly at a dose of 75 mg / m 2 It is administered once every three weeks at a dose of

[0449] In some embodiments, the anti-clusterin antibody or antigen-binding fragment thereof is humanized 16B5 and is administered once weekly at a dose of 9 mg / kg, and the docetaxel is administered once weekly at a dose of 60 mg / m 2 It is administered once every three weeks at a dose of

[0450] In some embodiments, the anti-clusterin antibody or antigen-binding fragment thereof is humanized 16B5 and is administered once a week at a dose of 6 mg / kg, and the docetaxel is administered once a week at a dose of 75 mg / m 2 It is administered once every three weeks at a dose of

[0451] In some embodiments, the anti-clusterin antibody or antigen-binding fragment thereof is humanized 16B5 and is administered once a week at a dose of 6 mg / kg, and the docetaxel is administered once a week at a dose of 60 mg / m 2 It is administered once every three weeks at a dose of

[0452] In some embodiments, the anti-clusterin antibody or antigen-binding fragment thereof is humanized 16B5 and is administered at a dose of 3 mg / kg once weekly, and the docetaxel is administered at a dose of 75 mg / m 2 It is administered once every three weeks at a dose of

[0453] In some embodiments, the anti-clusterin antibody or antigen-binding fragment thereof is humanized 16B5 and is administered at a dose of 3 mg / kg once weekly, and the docetaxel is administered at a dose of 60 mg / m 2 It is administered once every three weeks at a dose of

[0454] A treatment cycle lasts, for example, 21 days. During one cycle of treatment, a subject may receive, for example, an anti-clusterin antibody or an antigen-binding fragment thereof once a week and docetaxel once every three weeks. A subject may receive two or more consecutive treatment cycles.

[0455] In some embodiments, the treatment cycle will end approximately 7 days after the subject has received both the anti-clusterin antibody, or antigen-binding fragment thereof, and docetaxel.

[0456] For example, if both the anti-clusterin antibody or antigen-binding fragment thereof and docetaxel are administered weekly, a treatment cycle would be considered to be 7 days.

[0457] For example, if the anti-clusterin antibody or antigen-binding fragment thereof is administered weekly and docetaxel is administered every two weeks, the treatment cycle would be considered to be 14 days.

[0458] For example, if the anti-clusterin antibody or antigen-binding fragment thereof is administered weekly and docetaxel is administered every three weeks, the treatment cycle would be considered to be 21 days.

[0459] In some exemplary embodiments, one cycle of treatment is approximately 21 days.

[0460] In some exemplary embodiments, essentially all treatment cycles are approximately 21 days.

[0461] In some exemplary embodiments, each treatment cycle is approximately 21 days.

[0462] Thus, in accordance with the present disclosure, a subject may receive a new treatment cycle every 21 days.

[0463] In accordance with the present disclosure, subjects may undergo at least one cycle of treatment prior to isolation of TILs.

[0464] In accordance with the present disclosure, subjects may receive at least two cycles of treatment prior to isolation of TILs.

[0465] In accordance with the present disclosure, subjects may receive at least three cycles of treatment prior to isolation of TILs.

[0466] In accordance with the present disclosure, subjects may receive at least four cycles of treatment prior to isolation of TILs.

[0467] In accordance with the present disclosure, subjects may receive four or more cycles of treatment prior to isolation of TILs.

[0468] In accordance with the present disclosure, subjects may receive at least five cycles of treatment prior to isolation of TILs.

[0469] In accordance with the present disclosure, subjects may receive at least six cycles of treatment prior to isolation of TILs.

[0470] In accordance with the present disclosure, subjects may receive at least seven cycles of treatment prior to isolation of TILs.

[0471] In accordance with the present disclosure, subjects may receive at least eight cycles of treatment prior to isolation of TILs.

[0472] In accordance with the present disclosure, subjects may receive at least nine cycles of treatment prior to isolation of TILs.

[0473] In accordance with the present disclosure, subjects may undergo at least 10 cycles of treatment prior to isolation of TILs.

[0474] In accordance with the present disclosure, subjects may receive at least 11 cycles of treatment prior to isolation of TILs.

[0475] In accordance with the present disclosure, subjects may receive at least 12 cycles of treatment prior to isolation of TILs.

[0476] In accordance with the present disclosure, subjects may receive at least 13 cycles of treatment prior to isolation of TILs.

[0477] In accordance with the present disclosure, subjects may receive at least 14 cycles of treatment prior to isolation of TILs.

[0478] In accordance with the present disclosure, subjects may receive at least 15 cycles of treatment prior to isolation of TILs.

[0479] In accordance with the present disclosure, subjects may receive at least 16 cycles of treatment prior to isolation of TILs.

[0480] In accordance with the present disclosure, subjects may receive at least 17 cycles of treatment prior to isolation of TILs.

[0481] In accordance with the present disclosure, subjects may receive at least 18 cycles of treatment prior to isolation of TILs.

[0482] In accordance with the present disclosure, subjects may receive at least 19 cycles of treatment prior to isolation of TILs.

[0483] In accordance with the present disclosure, subjects may undergo at least 20 cycles of treatment prior to isolation of TILs.

[0484] In accordance with the present disclosure, a subject may receive more than 20 cycles of treatment prior to isolation of TILs.

[0485] In accordance with the present disclosure, subjects may receive at least one cycle of treatment following infusion of the TILs.

[0486] In accordance with the present disclosure, subjects may receive at least two cycles of treatment following infusion of the TILs.

[0487] In accordance with the present disclosure, subjects may receive at least three cycles of treatment following infusion of the TILs.

[0488] In accordance with the present disclosure, subjects may receive at least four cycles of treatment following infusion of the TILs.

[0489] In accordance with the present disclosure, subjects may receive four or more cycles of treatment following infusion of the TILs.

[0490] In accordance with the present disclosure, subjects may receive at least five cycles of treatment following infusion of the TILs.

[0491] In accordance with the present disclosure, subjects may receive at least six cycles of treatment following infusion of the TILs.

[0492] In accordance with the present disclosure, subjects may receive at least seven cycles of treatment following infusion of the TILs.

[0493] In accordance with the present disclosure, subjects may receive at least eight cycles of treatment following infusion of the TILs.

[0494] In accordance with the present disclosure, subjects may receive at least 9 cycles of treatment following infusion of the TILs.

[0495] In accordance with the present disclosure, subjects may receive at least 10 cycles of treatment following infusion of the TILs.

[0496] According to the present disclosure, subjects may receive at least 11 cycles of treatment following infusion of the TILs.

[0497] In accordance with the present disclosure, subjects may receive at least 12 cycles of treatment following infusion of the TILs.

[0498] According to the present disclosure, subjects may receive at least 13 cycles of treatment following infusion of the TILs.

[0499] In accordance with the present disclosure, subjects may receive at least 14 cycles of treatment following infusion of the TILs.

[0500] In accordance with the present disclosure, subjects may receive at least 15 cycles of treatment following infusion of the TILs.

[0501] In accordance with the present disclosure, subjects may receive at least 16 cycles of treatment following infusion of the TILs.

[0502] In accordance with the present disclosure, subjects may receive at least 17 cycles of treatment following infusion of the TILs.

[0503] In accordance with the present disclosure, subjects may receive at least 18 cycles of treatment following infusion of the TILs.

[0504] In accordance with the present disclosure, subjects may receive at least 19 cycles of treatment following infusion of the TILs.

[0505] In accordance with the present disclosure, subjects may receive at least 20 cycles of treatment following infusion of the TILs.

[0506] In accordance with the present disclosure, subjects may receive more than 20 cycles of treatment following infusion of TILs.

[0507] In some embodiments, the anti-clusterin antibody, or antigen-binding fragment thereof, is administered by infusion over a time frame of approximately one hour.

[0508] In some embodiments, the docetaxel is administered by infusion over a time frame of approximately one hour.

[0509] In accordance with the present disclosure, the anti-clusterin antibody or antigen-binding fragment thereof and docetaxel are administered on the same day.

[0510] The anti-clusterin antibody or antigen-binding fragment thereof and docetaxel may be administered separately.

[0511] The anti-clusterin antibody or antigen-binding fragment thereof and docetaxel may be administered sequentially.

[0512] In some embodiments, the anti-clusterin antibody or antigen-binding fragment thereof is administered by infusion over approximately a one hour time frame, and docetaxel is subsequently administered on the same day by infusion over approximately a one hour time frame.

[0513] In some embodiments, docetaxel is administered by infusion over approximately a one hour time frame, and the anti-clusterin antibody or antigen-binding fragment thereof is subsequently administered on the same day by infusion over approximately a one hour time frame. EXAMPLES

[0514] Effect of AB-16B5 on immune cell infiltration in the tumor microenvironment Balb / c mice, 5 × 10 5 4T1 cells were orthotopically implanted into the fourth mammary fat pad. Animals received saline treatment IP three times a week. Primary tumors were surgically resected 16 days after implantation. Animals were sacrificed on day 36 and lungs were removed. Tissues were fixed in paraformaldehyde and processed for paraffin embedding. Tissue sections were probed with anti-mouse CD3, anti-mouse CD8, and anti-mouse B220 antibodies. Signals were revealed with horseradish peroxidase-specific secondary antibodies and counterstained with hematoxylin and eosin. The results depicted in Figure 1 show that 4T1 lung metastases create an immunologically cold microenvironment that prevents infiltration of B and T lymphocytes in the tumor. The outlined area shows that CD3 and CD8 T lymphocytes are restricted to the tumor periphery as a result of EMT.

[0515] Animals bearing 4T1 tumors were treated with AB-16B5 antibody (mouse 16B5) IP at 10 mg / kg three times a week. Primary tumors were surgically resected 16 days after implantation. Animals were sacrificed on day 36 and lungs were removed. Tissues were fixed in paraformaldehyde and processed for paraffin embedding. Tissue sections were probed with anti-mouse CD3, anti-mouse CD8, and anti-mouse B220 antibodies. Signals were revealed with horseradish peroxidase-specific secondary antibodies and counterstained with hematoxylin and eosin. The results, depicted in FIG. 2, show that lung metastases densely infiltrated with CD3 and CD8 T cells were fewer and significantly smaller. Similarly, evidence of plasma cell penetration in 16B5-treated tumors was also observed.

[0516] Thus, AB-16B5 allows the infiltration of immune cells in the tumor microenvironment in immune-competent mice. AB-16B5 may represent a novel therapeutic avenue for creating a warmer tumor microenvironment to stimulate a strong immune response against tumors.

[0517] In parallel, human tumor biopsies from patients treated with AB-16B5 (humanized 16B5) as a single agent were analyzed (Figures 2B-2E). Needle biopsies obtained from a patient with metastatic thyroid cancer and a patient with inoperable metastatic gastric cancer were sectioned and stained with hematoxylin and eosin. An as-treated biopsy from a patient with thyroid cancer metastasizing to the lung was obtained after a second cycle of treatment with AB-16B5. As shown in Figure 2B, essentially all tumor fragments were necrotic. Lymphoplasmacytic infiltrates were observed on the display along the margins of the fragments. Hemosiderin-depositing macrophages were observed within the necrotic areas, some reflecting erythrocyte extravasation associated with necrosis (not shown). Figure 2C shows a perivascular infiltrate composed of plasma cells along the margins of a tumor fragment from the same patient. Analysis of a pretreatment biopsy from a metastatic gastric cancer case showed several fragments of gastric mucosa infiltrated with diffuse undifferentiated gastric cancer (signet ring cells). The fragment on display showed mainly necrotic foci with acute neutrophilic infiltrates. Figure 2E shows a treatment biopsy obtained after a second cycle of treatment with AB-16B5, consisting of three tumor fragments. The larger fragments consisted of normal superficial gastric mucosa, while the smaller fragments were infiltrated with a mixture of neutrophilic and mononuclear immune cell infiltrates. EXAMPLES

[0518] Effect of combination therapy of AB-16B5 and docetaxel on the infiltration of immune cells in the tumor microenvironment. An immune-competent mouse cancer model was chosen to test the extent of immune response by treatment with AB-16B5 monotherapy or combination therapy of AB-16B5 and docetaxel using mouse 16B5.

[0519] Five groups of 10 female Balb / c mice each were assigned to the study (see Table 1 below). All animals received subcutaneous implantation of 4T1 mouse breast cancer cells into the fourth mammary fat pad. Treatment began on the day of implantation (defined as day 1). Group 1 (Gr. 1) animals received treatment with saline vehicle control IP for the duration of the study. Group 2 (Gr. 2) animals received 10 mg / kg docetaxel IP once a week for 5 weeks. Group 3 (Gr. 3) animals received 10 mg / kg docetaxel once a week for 2 weeks and 10 mg / kg AB-16B5 twice a week for 5 weeks. Group 4 (Gr. 4) animals received 10 mg / kg docetaxel once a week and 5 mg / kg 16B5 twice a week, each for a 5-week treatment course. Group 5 (Gr. 5) animals received AB-16B5 twice a week for 5 weeks. On day 36, primary tumors were excised, and on day 37, animals were sacrificed and the number of metastatic nodules visible to the naked eye on the lung surface was counted.

[0520] [Table 1]

[0521] The results shown in FIG. 3 show that the lungs of animals from groups 4 and 5 contained fewer metastatic lung nodules than saline control-treated mice. Similarly, mice tested in docetaxel monotherapy had as many metastatic lung nodules as the saline control group. Two weeks of treatment with docetaxel in combination with 16B5 resulted in fewer metastatic lung nodules than groups 1 and 2, but the response to treatment was not as extensive as groups 4 and 5. More animals in group 4 had no detectable nodules than any other group. These results suggest that AB-16B5 monotherapy or combination therapy with docetaxel effectively inhibits metastatic invasion in immune-competent mice. These results also suggest that it may be preferable to administer AB-16B5 and docetaxel throughout the course of treatment.

[0522] Primary tumors excised 16 days after implantation were treated with collagenase and hyaluronidase, and immune cells were purified by positive selection using magnetic latex beads coated with anti-CD45 antibodies. The purified cells were transferred to small petri dishes containing culture medium supplemented with IL2 and IL7, and phenotypic analysis was performed. It was found that there were very few CD45+ in the primary tumors taken from animals in groups 1 and 2. In contrast, there were more immune cells in the tumors taken from animals in groups 3, 4, and 5.

[0523] Treatment of mice implanted with 4T1 tumor cells with docetaxel (DTX 5W) was relatively ineffective. 4T1 tumors have an EMT-high signature that causes resistance to many chemotherapeutic agents, including docetaxel. Treatment of mice with docetaxel for 2 weeks and 16B5 for 5 weeks was not as effective as treatment with 16B5 as monotherapy, possibly because transient exposure of tumors to docetaxel resulted in increased tumor resistance. The combination of docetaxel and 16B5 for 5 weeks proved to be the most effective treatment regimen. The combination of increased shed antigens and inhibition of EMT caused by docetaxel resulted in an increased immune response leading to fewer lung metastases in this group compared to 16B5 monotherapy.

[0524] Thus, AB-16B5 as monotherapy and in combination with AB-16B5 and docetaxel enable infiltration of immune cells in the tumor microenvironment in immune-competent mice. EXAMPLES

[0525] Characterization, purification, and generation of tumor-infiltrating lymphocytes Balb / c mice, 5 × 10 54T1 cells were orthotopically implanted into the fourth mammary fat pad. Animals received AB-16B5 (mouse 16B5) 10 mg / kg intraperitoneally (IP) twice weekly (Group 15: animals 1501, 1502, and 1503) combined with docetaxel 10 mg / kg IP once weekly, or AB-16B5 10 mg / kg IP twice weekly (Group 25: animals). Primary tumors were surgically excised 16 days after implantation. Animals were sacrificed on day 36, lungs were removed, and each visible lung metastasis was carefully dissected. Each visible metastatic nodule, if any, was excised and processed for the tumor-infiltrating lymphocyte rapid expansion protocol. Metastatic nodules were minced into 2- to 3-mm cubes and grown individually in 24-well plates containing culture medium supplemented with FBS, IL2, IL7, ITS (1,000 U / mL IL2, 2.0 ng / mL IL7, and 1x insulin-transferrin-selenium cocktail (Gibco 41400-045)).

[0526] After 3 weeks of culture, 100,000 cells were harvested from each of the lymphocyte cultures (6 cultures corresponding to 3 animals from group 15 and 3 animals from group 25) and directly cultured with 100,000 4T1 tumor cells. After overnight co-culture, supernatants were harvested for INFγ quantification by ELISA.

[0527] Results of INFγ secretion from lymphocyte cultures in the presence of 4T1 cells showed that lymphocytes isolated from lung metastatic nodules secreted INFγ at high levels, with the highest mean levels observed in the docetaxel-16B5 group (see Table 2). These results confirm that inhibition of EMT by the antibody sCLU 16B5mAb contributes to the generation of a "warm" tumor microenvironment that allows infiltration of T lymphocytes in the tumor.

[0528] [Table 2]

[0529] Lymphocytes were stimulated with anti-CD3 and anti-CD28 monoclonal antibodies. Lymphocytes from each donor animal were pooled and processed for flow cytometry analysis with antibodies against CD45 (lymphocyte common antigen), CD3, CD4, CD8, and CD19 (B cell biomarker) (Figures 4A and 4B). The resulting single cell preparations were first selected for their size to select cells corresponding to immune cells. They were further gated on an FSC / SSC plot to remove dead cells and debris. Flow cytometry analysis was then performed with antibodies against CD45, CD3, CD19, CD3, CD4, and CD8. CD45 positive immune cells were gated on CD3 and CD19 (P3). CD3+ cells were further gated on CD4 and CD8 (Q1-LR).

[0530] The results showed 80-90% cell viability of CD45+ cells for both groups. CD45+ cells from group 15 (Figure 4A) contained 40.2%-55.0% CD19 cells and 14.0%-21.1% CD3+ cells. CD3+ cells contained 63.7%-66.5% CD4+ T cells and 20.6%-27.0% CD8+ T cells. CD45+ cells from group 25 (Figure 4B) contained 14.0%-35.0% CD19 cells and 21.3%-42.0% CD3+ cells. CD3+ cells contained 47.5%-67.8% CD4+ T cells and 25.9%-41.1% CD8+ T cells. EXAMPLES

[0531] Evaluation of immune reactivity of tumor-infiltrating T lymphocytes from mice treated with AB-16B5 in combination with docetaxel Balb / c mice, 5 × 10 54T1 cells were orthotopically implanted into the fourth mammary fat pad. Animals received AB-16B5 (mouse 16B5) 10 mg / kg intraperitoneally (IP) twice weekly in combination with docetaxel 10 mg / kg IP once weekly. Primary tumors were surgically resected 21 days after implantation. Animals were sacrificed on day 36, lungs removed, and each visible lung metastasis was carefully dissected. Eighteen lymphocyte cultures contained 1-3 small lung metastases in 24-well G-Rex multiwell plates (Wilson-Wolf, # 80192M). TILs were expanded in defined R&D Systems™ ExCellerate Human T Cell Expansion Medium (# CCM030) containing 600 IU / mL IL2. After 3 weeks of culture, 100,000 cells were harvested from each TIL culture, washed in PBS, and cultured with 100,000 4T1 tumor cells. After overnight coculture, the supernatant was removed and INFγ concentrations were assessed by ELISA. The results of INFγ secretion from TIL cultures in the presence of 4T1 cells show that lymphocytes isolated from lung metastatic nodules secrete various levels of INFγ. Based on the current literature, it was established that T cell cultures with IFNγ levels lower than 300 pg / mL are considered weak; cultures containing those between 300 pg / mL and 500 pg / mL are considered intermediate, and cultures above 500 pg / mL are considered high (see Table 3).

[0532] [Table 3]

[0533] As can be seen from Table 3, all TIL cultures had INFγ secretion levels equal to or greater than 100 pg / ml, 14 of these TIL cultures showed INFγ secretion levels equal to or greater than 300 pg / ml, and 11 of 18 TIL cultures showed INFγ secretion levels equal to or greater than 500 pg / ml.

[0534] Lymphocytes were further analyzed by flow cytometry. Lymphocytes were stimulated with anti-CD3 and anti-CD28 monoclonal antibodies. Lymphocytes from each culture were processed for flow cytometry analysis with antibodies against CD45, CD3, CD4, and CD8. The obtained single cell preparations were first selected based on their size to select cells corresponding to immune cells. They were gated on an FSC / SSC plot to remove dead cells and debris. Flow cytometry analysis was then performed with antibodies against CD45, CD3, CD4, and CD8. CD45 positive immune cells were gated on CD3. The results showed that 73%-95% of viable cells were CD3 positive. CD3+ cells were further gated on CD4 and CD8. The results showed that the conditions used to grow immunoreactive TILs were favorable for enriching CD8+ T cells. Interestingly, cultures with low IFNγ production, such as #3 and #5, had a higher content of CD4+ T cells, which may suggest the presence of CD4+ regulatory T cells.

[0535] [Table 4] EXAMPLES

[0536] Rapid growth of detailed TILs The following rapid expansion protocol is derived from Jin J. et al., J Immunother. 35(3):283-292, 2012, the entire contents of which are incorporated herein by reference.

[0537] Initial culture phase TILs were first isolated from tumor fragments (1–8 mm) produced by enzymatic tumor digestion and sharp dissection. 3) were cultured from the tumor. Tumor digests were generated by mechanical dissociation (GentleMACS, Miltenyi Biotec, Auburn, CA) after incubation in enzyme medium (RPMI 1640, 2 mM Glutmax, 10 μg / mL gentamicin, 30 units / mL DNase, and 1.0 mg / mL collagenase). Immediately after placing the tumor in the enzyme medium, the tumor was mechanically dissociated for approximately 1 minute. The solution was then incubated at 37° C., 5% CO 2 After incubation at 4°C for 30 min, the cells were again mechanically dissociated for approximately 1 min. The cells were then incubated again at 37°C, 5% CO 2 After incubation at 4°C for 30 min, the tumors were subjected to a third mechanical dissociation for approximately 1 min. If large tissue fragments were present after the third mechanical dissociation, the tumors were incubated at 37°C, 5% CO 2 One or two additional rounds of mechanical dissociation were applied to the samples with or without an additional 30 min incubation at 50° C. At the end of the final incubation, if the cell suspension contained large numbers of red blood cells or dead cells, a density gradient separation using Ficoll was performed to remove these cells.

[0538] When TIL cultures were initiated in 24-well plates (Costar 24-well cell culture clusters, flat bottom, Corning Incorporated, Corning, NY), each well contained 1 × 10 6 1 tumor digestive cell or approximately 1-8 mm in size 3 One tumor fragment was seeded in 2 mL of complete medium (CM) with IL-2 (6000 IU / mL, Chiron Corp., Emeryville, Calif.). CM consisted of RPMI 1640 with glutamine and supplemented with 10% human AB serum, 25 mM Hepes, and 10 μg / mL gentamicin. Cultures were grown in 40 mL volumes and 10 cm 2 Starting with gas-permeable flasks with gas-permeable silicone bottoms (G-Rex10, Wilson Wolf Manufacturing, New Brighton, MN, USA) (Figure 1), each flask was filled with 10–40 × 10 6Viable tumor digested cells or 5-30 tumor fragments were loaded into 10-40 mL of CM with IL-2 (recombinant human IL-2). Both G-Rex10 and 24-well plates were incubated at 37°C, 5% CO 2 Incubate the cells in a humidified incubator at 4 °C for 5 min. On day 5 after the initiation of culture, remove half of the medium and replace it with fresh CM and IL-2, and after day 5, replace half of the medium every 2-3 days.

[0539] Augmentation Phase The Rapid Expansion Protocol (REP) of TILs is performed using T-175 flasks and gas permeable bags or gas permeable G-Rex® flasks. For TIL REP in T-175 flasks, 1×10 6 TILs were added to each T-175 flask. TILs are cultured at a 1:100 ratio with irradiated (50 Gy) allogeneic peripheral blood mononuclear cells (PBMCs) as "feeder" cells, and cells are cultured in a 1:1 mixture of CM and AIM-V medium (50 / 50 medium) supplemented with 3000 IU / mL IL-2 and 30 ng / mL anti-CD3. The T-175 flasks are incubated at 37° C., 5% CO 2 Incubate at 4°C for 12 h. Half of the medium is replaced on day 5 using 50 / 50 medium with 3000 IU / mL IL-2. On day 7, combine the cells from the two T-175 flasks in a 3 liter bag and 300 mL of AIM V with 5% human AB serum and 3000 IU / mL IL2. Count the number of cells in each bag every day or two and add fresh medium to keep the cell numbers at 0.5-2.0 x 10 6 Maintain between 100 and 150 cells / mL.

[0540] 100cm 2 For TIL REP, 5 × 10 in a 500 mL flask with gas-permeable silicon bottom (G-Rex100, Wilson Wolf) (Figure 1). 6 Pieces or 10 x 10 6TILs are cultured in a 1:100 ratio with irradiated allogeneic PBMCs in 400 mL of 50 / 50 medium supplemented with 5% human AB serum, 3000 IU / mL IL-2 and 30 ng / mL antibody CD3. G-Rex100 flasks are incubated at 37° C., 5% CO 2 On day 5, 250 mL of the supernatant is taken, placed in a centrifuge tube, and centrifuged at 1500 rpm (491×g) for 10 minutes. The TIL pellet is resuspended in 150 mL of fresh medium with 5% human AB serum, 3000 IU / mL IL-2, and returned to the original G-Rex100 flask. The TILs are continuously expanded in the G-Rex100 flasks, and on day 7, the TILs in each G-Rex100 are resuspended in 300 mL of medium present in each flask, and the cell suspension is divided into three 100 mL aliquots that are used to seed three G-Rex100 flasks. Then, 150 mL of AIM-V with 5% human AB serum and 3000 IU / mL IL-2 is added to each flask. The G-Rex100 flasks are incubated at 37° C., 5% CO 2 After 4 days of incubation, 150 mL of AIM V with 3000 IU / mL IL-2 is added to each G-Rex100 flask. Cells are harvested on day 14 of culture.

[0541] Wardell et al. (U.S. Patent Application Publication No. 2018 / 0282694, the entire contents of which are incorporated herein by reference) discloses an improved and shortened process for expanding TILs and for producing therapeutic populations of TILs that are applicable to the present disclosure.

[0542] If desired, anti-CD28 antibody and / or anti-4-1B may be added during the expansion phase.

[0543] Cell counts, viability and flow cytometry Expression of CD3, CD4, CD8, and CD56 is measured by flow cytometry with antibodies from BD Biosciences (BD Biosciences, San Jose, Calif.) using a FACSCanto flow cytometer (BD Biosciences). Cells are manually counted using a disposable c-tip hemocytometer (VWR, Batavia, Ill.) and viability is assessed using trypan blue staining.

[0544] Cytokine release assay TILs are assessed for interferon gamma (IFN-γ) secretion in response to stimulation with OKT3 antibody or co-culture with autologous tumor digest. For OKT3 stimulation, TILs are washed extensively and duplicate wells were plated with 1×10 cells in 0.2 mL of CM in 96-well flat-bottom plates pre-coated with 0.1 or 1.0 μg / mL OKT-3 antibody diluted in PBS. 5 After overnight incubation, the supernatants were harvested and IFN-γ in the supernatants was measured by ELISA (Pierce / Endogen, Woburn, Mass.). For the co-culture assay, TIL cells were placed in 96-well plates with autologous tumor cells. After 24 hours of incubation, the supernatants were harvested and IFN-γ release was measured by ELISA.

[0545] The embodiments and examples described herein are illustrative and are not meant to limit the scope of the claims. Variations of the foregoing embodiments, including alternatives, modifications, and permeants, are intended by the inventors to be encompassed by the scope of the claims. The citations cited in this application are incorporated herein by reference. References Al-Lazikani et al., Standard conformations for the canonical structures of immunoglobulins. J Mol Biol 273:927-948, 1997. Brochet et al. IMGT / V-QUEST: the highly customized and integrated system for IG and TR standardized V-J and V-D-J sequence analysis. Nucl Acids Res 36:W503-W508, 2008. Andrew C.R. Martin, Antibody Engineering Vol. 2, Chapter 3: Protein Sequence and Structure Analysis of Antibody Variable Domains. R. Kontermann and S. Dubel (eds.), DOI 10.1007 / 978-3-642-01147-4_3, # Springer-Verlag Berlin Heidelberg 2010 Shibue, T., Weinberg, R. EMT, CSCs, and drug resistance: the mechanistic link and clinical implications. Nat Rev Clin Oncol 14: 611-629 (2017). Terry, S., Savagner, P., Ortiz-Cuaran, S., Mahjoubi, L., Saintigny, P., Thiery, J.-P. and Chouaib, S., New insights into the role of EMT in tumor immune escape. Mol Oncol, 11: 824-846 (2017). Lenferink, A., Cantin, C., Nantel, A. et al. Transcriptome profiling of a TGF-β-induced epithelial-to-mesenchymal transition reveals extracellular clusterin as a target for therapeutic antibodies. Oncogene 29: 831-844 (2010). New response evaluation criteria in solid tumours: Revised RECIST guideline (version 1.1)" E.A. Eisenhauer, P. Therasse, J. Bogaerts, L.H. Schwartz, D. Sargent, R. Ford, J. Dancey, S. Arbuck, S. Gwyther, M. Mooney, L. Rubinstein, L. Shankar, L. Dodd, R. Kaplan, D. Lacombe, J. Verweij; Eur J Cancer, 45 (2009) 228 -24 . Cristiano Ferrario, Julie Laurin, Leon Van Kempen, Caroline Lambert, Alan Spatz, Oksana Markova, Gerald Batist, Adrian Langleben, Mario Filion, Jacques Jolivet. Phase 1 first-in-human study of anti-clusterin antibody AB-16B5 in patients with advanced solid malignancies [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl): Abstract nr CT098. doi:10.1158 / 1538-7445.AM2017-CT098. Hodge, J.W. Garnett, C.T., Farsaci, B., et al. Chemotherapy-induced immunogenic modulation of tumor cells enhances killing by cytotoxic T lymphocytes and is distinct from immunogenic cell death. Int. J. Cancer. 133: 624-636 (2013). Jiang X, Dudzinski S, Beckermann KE, et al. MRI of tumor T cell infiltration in response to checkpoint inhibitor therapy. Journal for ImmunoTherapy of Cancer 2020;8:e000328. doi:10.1136 / jitc-2019-000328. MacCallum, R. M., Martin, A. C. R. and Thornton, J. T. 'Antibody-antigen interactions: Contact analysis and binding site topography' J. Mol. Biol. 262:732-745, 1996. Wu and Kabat, An analysis of the sequences of the variable regions of Bence Jones proteins and myeloma light chains and their implications for antibody complementarity. J Exp Med 132:211-250, 1993.

[0546]

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Claims

1. A pharmaceutical composition comprising an anti-clusterin antibody or antigen-binding fragment thereof for use in a subject having cancer to obtain a preparation of tumor-infiltrating lymphocytes (TILs) by isolating and expanding TILs from the subject's tumor, said use further comprising treating the subject by re-injecting the TIL preparation into the subject, wherein the anti-clusterin antibody or antigen-binding fragment thereof binds to the C-terminal portion of the β subunit of human clusterin as set forth in SEQ ID NO:

41.

2. A preparation of tumor-infiltrating lymphocytes (TILs) isolated from a tumor in a subject for use in treating cancer in the subject, wherein the subject has previously been treated with a pharmaceutical composition comprising an anti-clusterin antibody or antigen-binding fragment thereof, wherein the anti-clusterin antibody or antigen-binding fragment thereof binds to the C-terminal portion of the beta subunit of human clusterin as set forth in SEQ ID NO:

41.

3. A preparation of tumor-infiltrating lymphocytes (TILs) obtained by a method of treating a subject having cancer with a pharmaceutical composition containing an anti-clusterin antibody or an antigen-binding fragment thereof, and isolating and expanding tumor-infiltrating lymphocytes (TILs) from the tumor of the subject, wherein the anti-clusterin antibody or antigen-binding fragment thereof binds to the C-terminal portion of the β subunit of human clusterin as set forth in SEQ ID NO:

41.

4. A pharmaceutical composition as described in claim 1 or a preparation of TIL as described in claim 2 or a preparation of TIL as described in claim 3, wherein the preparation of TIL is isolated and expanded by an in vitro or ex vivo tumor infiltrating lymphocyte generation method, the generation method optionally including a step of contacting tumor fragments with an anti-clusterin antibody or an antigen-binding fragment thereof, and optionally the anti-clusterin antibody or antigen-binding fragment thereof is present and / or maintained during one or more phases of the method of generating the preparation of tumor infiltrating lymphocytes.

5. A pharmaceutical composition or preparation of TILs described in any one of claims 1 to 3, wherein the anti-clusterin antibody or antigen-binding fragment thereof is for use as a single agent or in combination with a chemotherapeutic agent.

6. 6. The pharmaceutical composition or preparation of TILs according to claim 5, wherein the chemotherapeutic agent is selected from an alkylating agent, an antimetabolite, an alkaloid, an antitumor antibiotic, or a combination thereof.

7. 6. The pharmaceutical composition or preparation of TILs of claim 5, wherein the chemotherapeutic agent is a taxane, optionally docetaxel, paclitaxel, Abraxane®, cabazitaxel, larotaxel, mirataxel, ortataxel, or tesetaxel.

8. A pharmaceutical composition or preparation of TILs according to any one of claims 1 to 3, wherein the tumor is resectable.

9. A pharmaceutical composition or preparation of TILs described in any one of claims 1 to 3, wherein the subject has a functional immune system or the subject is not immunosuppressed or has not received an immunosuppressant within 14, 13, 12, 11, 10, 9, 8, 7, 6, 5, 4, 3, 2, or 1 day prior to treatment with the anti-clusterin antibody or its antigen-binding fragment or treatment with the anti-clusterin antibody or its antigen-binding fragment and docetaxel combination therapy.

10. 4. A pharmaceutical composition or preparation of TILs according to any one of claims 1 to 3, wherein the preparation of TILs comprises non-genetically modified or genetically modified TILs, optionally comprising TILs expressing a chimeric antigen receptor or a transgenic T cell receptor.

11. The anti-clusterin antibody or antigen-binding fragment thereof, a. a light chain variable region comprising the complementarity determining regions (CDRs) of the light chain variable region set forth in SEQ ID NO: 9, and a heavy chain variable region comprising the CDRs of the heavy chain variable region set forth in SEQ ID NO: 10; b. a light chain variable region having an amino acid sequence that has at least 80% identity to the amino acid sequence set forth in SEQ ID NO:9, and a heavy chain variable region having an amino acid sequence that has at least 80% identity to the amino acid sequence set forth in SEQ ID NO:10; or c. a light chain having an amino acid sequence that has at least 80% identity to the amino acid sequence set forth in SEQ ID NO:11, and a heavy chain having an amino acid sequence that has at least 80% identity to the amino acid sequence set forth in SEQ ID NO:

12.

4. A pharmaceutical composition or preparation of TILs according to any one of claims 1 to 3, comprising:

12. A pharmaceutical composition or TIL preparation described in any one of claims 1 to 3, wherein the antibody or its antigen-binding fragment is capable of competing with an antibody comprising a light chain variable region having the amino acid sequence set forth in SEQ ID NO: 9 and a heavy chain variable region having the amino acid sequence set forth in SEQ ID NO: 10 for binding to clusterin.

13. The preparation of TILs is CD4 + A pharmaceutical composition or preparation of TILs according to any one of claims 1 to 3, comprising T cells, CD8 + T cells, B cells, NK cells and / or NK T cells.

14. A pharmaceutical composition or a preparation of TILs described in any one of claims 1 to 3, wherein the preparation of TILs is selected for tumor antigen recognition and / or the preparation of TILs secretes intermediate to high levels of INFγ.

15. A pharmaceutical composition or preparation of TILs according to any one of claims 1 to 3, wherein the anti-clusterin antibody or its antigen-binding fragment is used at a dose of between approximately 3 mg / kg and approximately 20 mg / kg before isolation of TILs or after infusion of TILs, or the anti-clusterin antibody or its antigen-binding fragment is used at a dose of approximately 6 mg / kg, approximately 9 mg / kg, and / or approximately 12 mg / kg before isolation of TILs or after infusion of TILs.

16. Docetaxel was administered at approximately 60 mg / m before isolation of TILs or after infusion of TILs. 2 ~ approx. 100 mg / m 2 or docetaxel is used at a dose of approximately 60 mg / m2 and / or at a dose of approximately 75 mg / m2.

17. a. The anti-clusterin antibody or antigen-binding fragment thereof is used at a dose of approximately 12 mg / kg once a week, and docetaxel is used at approximately 75 mg / m 2 used once every 3 weeks at a dose of; b. the anti-clusterin antibody or antigen-binding fragment thereof is administered once weekly at a dose of approximately 12 mg / kg and docetaxel is administered once every three weeks at a dose of approximately 60 mg / m 2 ; c. the anti-clusterin antibody or antigen-binding fragment thereof is administered once weekly at a dose of approximately 9 mg / kg and docetaxel is administered once every three weeks at a dose of approximately 75 mg / m 2 ; d. the anti-clusterin antibody or antigen-binding fragment thereof is administered once weekly at a dose of approximately 9 mg / kg and docetaxel is administered once every three weeks at a dose of approximately 60 mg / m 2 ; e. the anti-clusterin antibody or antigen-binding fragment thereof is administered once weekly at a dose of approximately 6 mg / kg and docetaxel is administered once every three weeks at a dose of approximately 75 mg / m 2 ; f. the anti-clusterin antibody or antigen-binding fragment thereof is administered once weekly at a dose of approximately 6 mg / kg and docetaxel is administered once every three weeks at a dose of approximately 60 mg / m 2 ; g. the anti-clusterin antibody or antigen-binding fragment thereof is administered once a week at a dose of approximately 3 mg / kg and docetaxel is administered once every three weeks at a dose of approximately 75 mg / m2; or h. the anti-clusterin antibody or antigen-binding fragment thereof is administered once weekly at a dose of approximately 3 mg / kg and docetaxel is administered once every three weeks at a dose of approximately 60 mg / m 2 ; A pharmaceutical composition or preparation of TILs according to claim 7.

18. A pharmaceutical composition or preparation of TILs according to any one of claims 1 to 3, wherein the subject has a carcinoma, optionally wherein the carcinoma is metastatic.

19. 4. The pharmaceutical composition or preparation of TILs of any one of claims 1 to 3, wherein the subject has endometrial cancer, breast cancer, liver cancer, prostate cancer, kidney cancer, bladder cancer, cervical cancer, ovarian cancer, colorectal cancer, pancreatic cancer, lung cancer, gastric cancer, head and neck cancer, thyroid cancer, cholangiocarcinoma, mesothelioma, melanoma, metastatic endometrial cancer, metastatic breast cancer, metastatic liver cancer, metastatic prostate cancer, metastatic kidney cancer, metastatic bladder cancer, metastatic cervical cancer, metastatic ovarian cancer, metastatic colorectal cancer, metastatic pancreatic cancer, metastatic lung cancer, metastatic gastric cancer, metastatic head and neck cancer, metastatic thyroid cancer, metastatic cholangiocarcinoma, metastatic mesothelioma, or metastatic melanoma.

20. A pharmaceutical composition or preparation of TILs according to any one of claims 1 to 3, wherein the subject has undergone lymphodepletion conditioning prior to infusion of the TILs.

21. A preparation of TILs comprising a majority of CD45 + A pharmaceutical composition or preparation of TILs described in any one of claims 1 to 3, comprising cells, a majority of CD3+ cells, a majority of CD4+ cells, a majority of CD8+ cells, a majority of CD4+ or CD8+ cells, or wherein the preparation of TILs comprises TILs that secrete intermediate to high levels of INFγ.

22. A pharmaceutical composition or a preparation of TILs described in any one of claims 1 to 3, wherein the preparation of TILs is for use in adoptive cell therapy.

23. A product comprising a preparation of TILs according to claim 2 or 3, optionally said product being an infusion bag containing said preparation of TILs.

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