Methods of administering bryozoin for inducing tumor-associated antigens

Abstract

The bryozoxin-1 stimulates the expression of tumor-associated antigens in blood system tumors and other tumors, thereby enabling these tumors to be recognized by immunotherapies and increasing clinical responses to various immunotherapies. Accordingly, provided herein are methods of treating cancer in a subject by administering bryozoin-1 or a functional analog based on this example.

Description

[0001] CROSS-REFERENCE TO RELATED APPLICATIONS

[0002] This application claims the benefit of priority under 35 USC §119(e) to U.S. Provisional Application No. 63 / 305,198, filed on January 31, 2022. The disclosure of the prior application is considered part of the disclosure of the present application and is incorporated by reference in its entirety into the disclosure of the present application. Technical Field

[0003] The present invention relates generally to cancer, and more specifically to the induction of tumor-associated antigens to improve immune surveillance of abnormal cells in a subject and provide an enhanced number of targets for immuno-oncology drugs primarily in the setting of hematological malignancies. Background Art

[0004] Cancer is a disease in which cells escape normal regulatory processes, transform and grow rapidly. These rapidly growing cells may appear in the blood (hematological malignancies) or may appear in various organs as solid tumors. Abnormal, rapidly growing cells are distinguished by the expression of unique antigens, which are recognized by the host immune system and are controlled by various immune mechanisms. When the transformed cells escape normal immune regulatory mechanisms, the rapidly growing cells can become malignant tumors, whether in the blood or as solid tumors.

[0005] Such tumors express various antigens called tumor-associated antigens (TAAs): cellular proteins constitutively expressed by tumor cells. These antigens provide stimulation for host immune effector functions and additionally serve as targets for immunotherapy, in particular monoclonal antibodies or fragments thereof, antibody-drug conjugates (ADCs) and chimeric antigen receptor (CAR)-T cells, and are examples of such immuno-oncology approaches for hematological malignancies.

[0006] An important aspect of these immune-mediated therapies is the adequate expression of tumor antigens so that immunotherapy can identify abnormal cells for destruction. In B-cell malignancies, TAAs may be specific to the tumor type or may be shared between tumors because they are primarily derived from B cells. CD19 and CD20, as well as other TAAs, are associated with non-Hodgkin's lymphoma and are targets for several approved tumor agents. Similarly, for example, other TAAs are associated with chronic lymphocytic leukemia or acute lymphoblastic leukemia (CD22), acute myeloid leukemia (CD33 and CD123), and multiple myeloma (CD38 and B cell maturation antigen (BCMA)). Several studies of these hematological malignancies have shown that the quantitative expression or density of TAAs expressed by tumor cells is an important factor in the efficacy of immunotherapy. Although less common, T-cell malignancies (lymphomas and leukemias, such as T-cell acute lymphoblastic leukemia) also express a variety of tumor-associated antigens.

[0007] Bryostatin-1 and its analogs are modulators of protein kinase C (PKC) that stimulate PKC activity at low concentrations and cause PKC downregulation at high concentrations. Stimulation of PKC induces the expression of certain genes, including those for tumor-associated antigens, and interacts with various intracellular signaling mechanisms. For example, bryostatin-1 has been shown to increase cell surface expression of CD22, a tumor-associated antigen in B-cell tumors such as acute lymphoblastic leukemia and chronic lymphocytic leukemia (ALL and CLL) (Varterasian et al., 2000; Biberacher et al., 2012; Ramakrishna et al., 2019).

[0008] In earlier studies, bryostatin-1 has been studied as a single-agent for the treatment of cancer. These studies used various dosing regimens; in particular, long continuous infusion times of up to 3 weeks. The reason for extended administration is to stimulate cancer cell differentiation and / or stop growth. In the present invention, short-term administration provides the 'trigger' required to stimulate tumor antigen expression. This shorter 'trigger administration' method will increase the therapeutic index of bryostatin-1 by minimizing the drug exposure required to stimulate antigen expression to a brief pulse or trigger.

[0009] The present invention relates to a method for administering bryostatin-1 or a functional analogue thereof to a subject, wherein the method is used to treat B-cell or T-cell malignancies in a subject in need thereof or to induce, enhance or prolong an anti-tumor response in the subject, thereby enhancing the therapeutic effect on cancer by inducing, enhancing or prolonging the subject's anti-tumor response to an anti-cancer agent, particularly an immunotherapeutic agent.

[0010] The present invention further relates to compounds that induce tumor antigen expression, and a method of administering ('triggered administration') said compounds to stimulate or induce tumor associated antigen expression. This allows the compound to be administered over a short period of time ('trigger'), thereby improving the therapeutic index or safety. In theory, short-term administration provides the 'trigger' required to stimulate tumor antigen expression. This shorter 'triggered administration' method will further improve the therapeutic index of bryostatin-1 by minimizing the drug exposure required to stimulate antigen expression to a brief pulse or trigger.

[0011] Specific methods of administering bryostatin-1 or a functional analogue described and claimed herein involve 'triggered dosing', which is a method of administration in which a short pulse of bryostatin-1 or a functional analogue causes tumor antigen expression on the cell surface for a much longer period of time after the blood level of bryostatin-1 or a functional analogue is no longer measurable. Specific methods of administration involve administering bryostatin-1 or a functional analogue thereof by intravenous, intraperitoneal, intramuscular, subcutaneous or oral administration. These methods of administration are intended to produce a short pulse or 'trigger' to obtain the desired effect.

[0012] It is further known that bryostatin-1 and functional analogs can induce T cell activation, a beneficial effect in the treatment of cancer.The specific 'triggered administration' approach described herein is also effective in triggering T cell activation. Summary of the invention

[0013] The present invention is not limited to the specific compositions, methods and experimental conditions described, as such compositions, methods and conditions may vary. In addition, the terminology used herein is for the purpose of describing specific embodiments only and is not intended to be limiting, as the scope of the present invention will be limited only by the appended claims.

[0014] All publications, patents, and patent applications mentioned in this specification are herein incorporated by reference to the same extent as if each individual publication, patent, or patent application was specifically and individually indicated to be incorporated by reference.

[0015] Unless otherwise defined, all technical and scientific terms used herein have the same meaning as those commonly understood by those of ordinary skill in the art to which the invention belongs. Although any methods and materials similar or equivalent to those described herein may be used in the practice or testing of the present invention, it should be understood that modifications and variations are encompassed within the spirit and scope of the present disclosure.

[0016] In one embodiment, the technology described herein relates to a method of treating cancer in a subject, the method comprising administering to the subject a bryostatin compound at a dose effective to induce upregulation of a tumor antigen in the subject for a period of time, thereby treating the cancer in the subject.

[0017] In some embodiments of the methods described herein, the bryostatin compound is bryostatin-1 or a bryostatin analog. In some embodiments, the administration time is about 0.5 to 24 hours. In addition, in some embodiments, the administration time is about 30 minutes, about 45 minutes, about 1 hour, about 2 hours, or about 3 hours.

[0018] Embodiments herein also include a method further comprising administering an immunotherapeutic agent. In some embodiments, the immunotherapeutic agent is administered prior to, concurrently with, and / or after the administration of a bryostatin compound.

[0019] In various embodiments of the methods described herein, the cancer is selected from the group comprising a hematological neoplasia comprising B-cell lymphoma, T-cell malignancies, multiple myeloma, acute myeloid leukemia, chronic myeloid leukemia, myelodysplastic syndrome, acute lymphoblastic leukemia, chronic lymphocytic leukemia, and hairy cell leukemia.

[0020] In some embodiments of the methods described herein, the immunotherapeutic agent is selected from a monoclonal antibody or a fragment thereof, a bispecific antibody, an antibody-drug conjugate, a chimeric antigen receptor (CAR)-T cell, and an anti-tumor vaccine product or a combination thereof. In various embodiments, the immunotherapeutic agent binds to a protein selected from the group consisting of: CD22, CD19, CD20, CD33, CD37, CD38, CD123, and BCMA.

[0021] In addition, in some embodiments of the methods described herein, the method further comprises administering to the subject an agent selected from the following: antibodies against tumor-associated antigens (TAA), chemotherapeutic agents, immunotoxic agents, vaccines, immunomodulatory drugs, immunometabolism modifying drugs, targeted therapies, radiation, anti-angiogenic agents, CAR-T cell therapy, bispecific antibodies, T cell engagers, agents that reduce immunosuppression, or a combination thereof. BRIEF DESCRIPTION OF THE DRAWINGS

[0022] Figure 1 is a graph showing the increase in CD19 expression in human peripheral blood mononuclear cells (PBMCs) in response to bryostatin-1 (1 nM) administration for 45 minutes, 3 hours, or 24 hours, demonstrating that a 45 minute incubation produced the same increase as longer incubations.

[0023] Figure 2is a graph showing the increase in CD22 expression in human peripheral blood mononuclear cells (PBMCs) in response to bryostatin-1 (1 nM) administration for 45 minutes, 3 hours, or 24 hours, demonstrating that a 45 minute incubation produced the same increase as longer incubations.

[0024] Figure 3A and 3B is a graph showing that the increase in antigen expression remains elevated over 7 days (Ramakrishna, S. et al. (2019) Modulation of target density improves CAR T-cell functionality and persistence. Clinical Cancer Research, 25(17), 5329-5341). DETAILED DESCRIPTION

[0025] As used in this specification and the appended claims, the singular forms "a," "an," and "the" include plural referents unless the context clearly dictates otherwise. Thus, for example, reference to "the method" includes one or more methods and / or steps of the type described herein that will become apparent to those skilled in the art after reading this disclosure.

[0026] As used herein, the terms "subject," "patient," or "subjects" refer to humans or other mammals, including ungulates, rodents, or primates; for example, humans, horses, cows, sheep, pigs, goats, llamas, camels, dogs, cats, birds, ferrets, rabbits, squirrels, mice, opossums, lemurs, or rats. In some embodiments, the subject is a human subject.

[0027] The term "effective amount" refers to an amount of a compound, composition or formulation sufficient to treat a condition or disease, produce a desired effect or result, or alleviate, relieve or stop the symptoms of a condition or disease. "Effective amount" can be used interchangeably with the term "therapeutically effective amount".

[0028] As used herein, the term "tumor-associated antigen" (TAA) is an antigen present in a subject's tumor cells or tissues. TAAs are proteins expressed on the surface of tumor cells that can be recognized by the immune system and immunotherapeutic agents. Such proteins can be glycoproteins, phosphoproteins, glycophosphoproteins, phosphoglycoproteins, etc. The quantitative expression of tumor-associated antigens (also referred to as tumor antigen density) is a known correlate of clinical response. These antigens may also be expressed in normal cells, reflecting their constitutive expression and normal biological functions.

[0029] Bryostatin-1 or its functional analogs (also referred to as "bryologs" or bryostatin compounds) refer to a group of macrolide lactones known to be potent modulators of protein kinase C (PKC). Bryostatin-1 is derived from the marine bryozoan Bugula neritina and is a potent modulator of the protein kinase C (PKC) enzyme system, a family of proteins involved in cell signaling, cell proliferation, and cell death. Therefore, bryostatin-1 has applications in a variety of therapeutic indications. To date, 20 different bryostatin compounds have been isolated. Examples of bryostatin analogs include compounds such as Picolog, Acetal 7c, and Merle 23. Bryostatin-1 is the best characterized analog and is the only member of the bryostatin family that has been evaluated in human clinical trials. At low concentrations, it is a potent PKC activator and causes activation of cell signaling, induction of transcription, and cellular expression of membrane-associated tumor and / or human immunodeficiency virus (HIV) antigens. Thus, bryostatin-1 renders these cells more responsive to the host immune system and exogenously administered immunotherapeutics targeting these antigens. See, e.g., U.S. Pat. No. 8,816,122 to Wender et al., which is incorporated herein by reference.

[0030] As used herein, the term "treatment" refers to a method or regimen intended to improve or alleviate the symptoms of a disease, illness, or infirmity. Treatment may reduce pain, improve quality of life, or reduce the size, number, or distribution of a tumor, cancer, or cancer cell. Treatment of cancer also occurs when symptoms or tests of cancer or cancer cells improve, or when symptoms or tests of cancer or cancer cells do not worsen or stabilize.

[0031] As used herein, the term "bryostatin compound" refers to a compound having a potential bryostatin pharmacophore based on the bryostatin natural product and in some cases having a scaffold characterized by a macrocyclic lactone-containing ring comprising three embedded six-membered rings (e.g., tetrahydropyran rings designated as A, B, and C rings), and an arrangement of carbon atoms numbered C1 to C26, as exemplified in the structure of bryostatin 1 shown below. The lactone of the scaffold is defined by a bond between the C1 carbonyl and the oxygen of the C25 hydroxyl.

[0032]

[0033] The bryostatin scaffold may include an olefin between C16 and C17 and an exocyclic olefin at the C13 and C21 positions. The bryostatin scaffold may include, for example, a specific arrangement of stereocenters at C3, C5, C7, C9, C11, C15, C19, C20 and C23, C25 and / or C26. Various substituents and derivative groups (e.g., ester or ether groups) may be included in the subject bryostatin compound (see, for example, Wender et al. WO2018 / 067382). Naturally occurring bryostatins were originally isolated from marine bryozoans and include a family of about 21 known compounds. The term "bryostatin compound" is intended to include naturally occurring bryostatin compounds (e.g., bryostatin 1) and "bryostatin analog compounds", which include non-naturally occurring bryostatin analogs and derivative compounds of interest that retain the functions required for biological activity.

[0034] The term "bryostatin compound" is intended to include compounds having the same potential bryostatin pharmacophore; the pharmacophore is the three hydrogen bond donors and acceptors that provide its binding function and the three-dimensional spatial arrangement of lipid domains that provide its association with the membrane and its derivative functions. The bryostatin pharmacophore or PKC pharmacophore model was introduced by the Wender group in 1986 (see, e.g., Wender, Proceedings of the National Academy of Sciences (PNAS), 1986, 83, 4214-4218), and was extended to bryostatin in 1988 (see, e.g., Wender, Proceedings of the National Academy of Sciences, 1988, 85, 7197-7201), thereby designing the first bryostatin analogs (e.g., Wender, Journal of the American Chemical Society (JACS), 1998, 120, 4534-4535; and Wender, Proceedings of the National Academy of Sciences, 1998, 95, 6624-6629, the disclosures of which are incorporated herein by reference in their entirety). The pharmacophore model is described in U.S. Patent No. 8,735,609, the disclosure of which is incorporated herein by reference in its entirety.

[0035] As used herein, the term "beneficial effect" or "effective" refers to ameliorating symptoms of a disease, reducing adverse symptoms, improving clinical test results, alleviating pain, improving quality of life or other desired results.

[0036] As used herein, the term "administering" refers to any method of administering a treatment to a patient or subject. Administration can be accomplished by topical, intravenous, intramuscular, systemic, oral, or parenteral methods. Administration can be performed at multiple or single time points and can be combined with a variety of treatments.

[0037] As used herein, a molecule that binds to another molecule refers to the ability of a molecule to target another molecule, whether the molecule is a protein, nucleic acid, antibody, or small molecule. For example, proteins such as CD19 and CD22 are common antigens targeted by immunotherapies that include antibodies, antibody fragments, diabodies, or other molecules that bind to cancer-associated antigens.

[0038] As used herein, the term "upregulation" refers to an increase in the number, percentage, concentration or amount of an antigen presented on the surface of a cancer cell. This may be due to an increase in the transcription of the antigen, a decrease in a physiological process that inhibits the production of the antigen by the transcription product, a decrease in a process that removes the antigen from the surface of a cancer cell or tumor, or any other process that results in an increase in the antigen presented on the surface of the cancer cell, thereby allowing the cell to be detected and destroyed by the immune system.

[0039] The present invention is based on the discovery that bryostatin-1 and its analogs can stimulate the expression of TAAs, thereby improving the recognition of abnormal cells (e.g., cancer cells) expressing these antigens and improving the ability of immunotherapy to target and kill these abnormal cells.

[0040] Bryostatin-1 stimulates the expression of tumor-associated antigens in B-cell tumors (hematological malignancies), thereby enabling these tumors to be recognized by immunotherapy and increasing the clinical response to various immunotherapies. Analogs of bryostatin also stimulate the expression of TAAs in hematological malignancies, thereby enabling these tumors to be recognized by the immune system and immunotherapy. Bryostatin compounds can increase the clinical response to various immunotherapies. Bryostatin-1 stimulates the expression of TAAs in leukemias, lymphomas, and myeloma; specifically, myeloid leukemias, non-Hodgkin lymphomas, and multiple myeloma, thereby enabling these tumors to be recognized by immunotherapy to improve the clinical response to various immunotherapies. Analogs of bryostatin also stimulate the expression of tumor-associated antigens in leukemias, lymphomas, and myeloma; specifically, myeloid leukemias, non-Hodgkin lymphomas, and multiple myeloma, thereby enabling these tumors to be recognized by immunotherapy and increasing the clinical response to various immunotherapies.

[0041] The main obstacle to the efficacy of cancer immunotherapy is related to the lack of TAA, which has the function of participating in the host immune function and serves as a cellular target for immunotherapeutics, which include monoclonal antibodies or fragments thereof, ADCs and CAR-T agents. Cell membrane presentation of TAA (also known as TAA density) has been described as a predictor of response to immunotherapeutics. In fact, clinical failure or relapse after CD19 CAR-T in acute lymphoblastic leukemia is associated with loss and cellular internalization of target CD19 molecules (e.g., see Fry et al., 2018). In acute lymphoblastic leukemia, a similar phenomenon also occurs after CD22CAR-T, and the phenomenon is associated with loss and cellular internalization or shedding of CD22 target molecules (Fry et al., 2018; Guedan and Delgado, 2019).

[0042] The ability to minimize side effects is an important parameter in therapeutics; this is especially true when drugs are given in combination. Therefore, it is highly desirable to reduce the patient's therapeutic exposure to the minimum dose required to produce an effective result. In most of the early clinical studies on bryostatin-1, long-term infusions were used (see, e.g., Varterasian et al., 2000, 72-hour infusion). For these early studies, the goal was to downregulate protein kinase C and induce tumor cell differentiation, thereby preventing tumor growth.

[0043] In contrast, current approaches are to stimulate antigen expression, making tumor cells visible to cancer immunotherapy, thereby making immunotherapy more effective. Preliminary animal experiments have shown that a single intraperitoneal administration of bryostatin-1 can induce an increase in tumor-associated antigen expression (Ramakrishna et al., 2019).

[0044] There are many immuno-oncology drugs and therapies that can be combined with bryostatin-1 to treat a patient's cancer. Examples of FDA-approved antibodies for treating cancer include, but are not limited to, Avastin (bevacizumab), Bexxar (tositumomab), CDP 870 and CEA-Scan (arcitumomab), denosumab, Erbitux (cetuximab), Herceptin (trastuzumab), Humira (adalimumab), IMC-IIF 8. LeukoScan (sulesomab), Campath (alemtuzumab), MabThera (rituximab), matuzumab, Mylotarg (gemtuzumabozogamicin), natalizumab, panitumamab, Panorex (edrecolomab), Raptiva (efalizumab), Remicade (infliximab), ReoPro (abciximab), Rituximab, Simulect (basiliximab), Synagis (palivizumab), TheraCIM hR3, tocilizumab, Tysabri (natalizumab), Verluma (nofetumomab), Xolair (omalizumab), Zenapax (dacliximab), Zevalin (ibritumomab conjugated to yttrium-90)tiuxetan (IDEC-Y2B8), Gilotrif (afatinib), Lynparza (olaparib), Perjeta (pertuzumab), Opdivo (nivolumab), Bosulif (bosutinib), Cabometyx (cabozantinib), trastuzumab dkst (Ogivri), Sutent (sunitinib malate), Adcetris (brentuximab vedotin), Alecensa (alectinib), Calquence (acalabrutinib), Kymriah (tisagenlecleucel), Yescarta (axicabtagene ciloleucel), Blincyto (blinatumomab), Besponsa (inotuzumab ozogamicin), Lumoxiti (moxetumomab-tdfk), pasudotox-tdfk), LymphoCide (epratuzumab), Verzenio (abemaciclib), Keytruda (pembrolizumab), Aliqopa (copanlisib), Nerlynx (neratinib), Imfinzi (durvalumab), Darzalex (daratumumab), Tecentriq (atezolizumab), Tarceva (erlotinib).

[0045] It has been demonstrated that short-term exposure to bryostatin-1 in vitro is sufficient to increase the expression of tumor-associated antigens. In experiments performed on a number of tumor cell lines and human cells, bryostatin-1 was administered for varying periods of time ranging from 45 minutes to 24 hours, and changes in CD19 and CD22 expression were measured. Short-term exposure (45 minutes) produced the same level of increase in antigen expression as 3-hour or 24-hour exposure ( Figure 1 and 2 ). This increase took time to become evident and reached a maximum at 72 hours. This suggests that short pulses (no longer than 45 minutes) can elicit a response equivalent to much longer incubations, corresponding to short doses rather than long infusions in the clinical setting.

[0046] In view of these data, the concept of 'triggered dosing' was introduced, in which short pulses (a few minutes) of exposure to bryostatin-1 or other agents can increase tumor antigen expression for several days. This method of administration reduces the need to expose cancer patients to bryostatin-1 for a long time and eliminates possible interactions between bryostatin-1 and cancer immunotherapy agents. The 'trigger' (bryostatin-1) can be administered once (before or after the cancer immunotherapy agent), or more than once (before and after the cancer immunotherapy agent), and the cancer immunotherapy agent can be administered at another time before or after the trigger agent is administered. Therefore, the administration of the two agents can be separated. This will reduce the possibility of drug interactions and adverse events caused by the combination therapy.

[0047] In some aspects, the disclosure described herein relates to a method of treating cancer in a subject, the method comprising administering to the subject a bryostatin compound at a dose effective to induce upregulation of a tumor antigen in the subject for a period of time, thereby treating the cancer in the subject. In some aspects, the bryostatin compound is bryostatin-1 or a bryostatin analog, whether natural or synthetic.

[0048] In some embodiments, the duration of administration is about 15 minutes to about 24 hours. In addition, in some embodiments, the administration time is about 30 minutes, about 3 hours, or about 24 hours. The administration time can be about 1 minute, about 2 minutes, about 3 minutes, about 4 minutes, about 5 minutes, about 6 minutes, about 7 minutes, about 8 minutes, about 9 minutes, about 10 minutes, about 11 minutes, about 12 minutes, about 13 minutes, about 14 minutes, about 15 minutes, about 16 minutes, about 17 minutes, about 18 minutes, about 19 minutes, about 20 minutes, about 21 minutes, about 22 minutes, about 23 minutes, about 24 minutes, about 25 minutes, about 26 minutes, about 27 minutes, about 28 minutes. , about 29 minutes, about 30 minutes, about 31 minutes, about 32 minutes, about 33 minutes, about 34 minutes, about 35 minutes, about 36 minutes, about 37 minutes, about 38 minutes, about 39 minutes, about 40 minutes, about 41 minutes, about 42 minutes, about 43 minutes, about 44 minutes, about 45 minutes, about 46 minutes, about 47 minutes, about 48 minutes, about 49 minutes, about 50 minutes, about 51 minutes, about 52 minutes, about 53 minutes, about 54 minutes, about 55 minutes, about 56 minutes, about 57 minutes, about 58 minutes, about 59 minutes, about 1 hour, about 1.5 hours, about 2 hours, about 2.5 hours, about 3 hours, about 3.5 hours, about 4 hours, about 4.5 hours, about 5 hours, about 5.5 hours, about 6 hours, about 6.5 hours, about 7 hours, about 7.5 hours, about 8 hours, about 8.5 hours, about 9 hours, about 9.5 hours, about 10 hours, about 10.5 hours, about 11 hours, about 11.5 hours, about 12 hours, about 1 Hours, about 2.5 hours, about 13 hours, about 13.5 hours, about 14 hours, about 14.5 hours, about 15 hours, about 15.5 hours, about 16 hours, about 16.5 hours, about 17 hours, about 17.5 hours, about 18 hours, about 18.5 hours, about 19 hours, about 19.5 hours, about 20 hours, about 20.5 hours, about 21 hours, about 21.5 hours, about 22 hours, about 22.5 hours, about 23 hours, about 23.5 hours or about 24 hours.

[0049] In the methods described herein, the administration of the bryostatin compound or bryostatin analog can be intravenous, intraperitoneal, intramuscular, subcutaneous, oral administration or a combination thereof. The duration of administration of the bolus dose or oral or subcutaneous dose can be different from the duration of administration of the intravenous dose (e.g., its duration is as short as 10 seconds to 3 minutes).

[0050] In some aspects of the methods described herein, the dosage of the bryostatin compound or bryostatin analog is about 5-60 μg / m 2 In other aspects of the methods described herein, the dosage of the bryostatin compound or bryostatin analog is about 30-50 μg / m 2In another aspect, the dosage of the bryostatin compound or bryostatin analog is about 45 μg / m2. In various aspects of the methods described herein, the dosage of the bryostatin compound or bryostatin analog is about 5 μg / m2, about 6 μg / m 2 , about 7 μg / m 2 , about 8 μg / m 2 , about 9 μg / m 2 , about 10 μg / m 2 , about 11 μg / m 2 , about 12 μg / m 2 , about 13 μg / m 2 , about 14 μg / m 2 , about 15 μg / m 2 , about 16 μg / m 2 , about 17 μg / m 2 , about 18 μg / m 2 , about 19 μg / m 2 , about 20 μg / m 2 , about μg / m 2 , about 21 μg / m 2 , about 22 μg / m 2 , about 23 μg / m 2 , about 24 μg / m 2 , about 25 μg / m 2 , about 26 μg / m 2 , about 27 μg / m 2 , about 28 μg / m 2 , about 29 μg / m 2 , about 30 μg / m 2 , about 31 μg / m 2 , about 32 μg / m 2 , about 33 μg / m 2 , about 34 μg / m 2 , about 35 μg / m 2 , about 36μg / m 2 , about 37 μg / m 2 , about 38 μg / m 2 , about 39μg / m 2 , about 40 μg / m 2 , about 41 μg / m 2 , about 42 μg / m 2 , about 43 μg / m 2 , about 44 μg / m 2 , about 45 μg / m 2 , about 46μg / m 2 , about 47 μg / m 2 , about 48 μg / m 2 , about 49μg / m2 , about 50 μg / m 2 , about 51 μg / m 2 , about 52 μg / m 2 , about 53 μg / m 2 , about 54 μg / m 2 , about 55 μg / m 2 , about 56μg / m 2 , about 57 μg / m 2 , about 58 μg / m 2 , about 59μg / m 2 or about 60 μg / m 2 .

[0051] The methods described herein further comprise administering an immunotherapeutic agent. In some embodiments, the immunotherapeutic agent is administered prior to, concurrently with, or after administration of the bryostatin compound.

[0052] In various embodiments of the methods described herein, the cancer is selected from the group comprising hematological neoplasia, comprising B-cell lymphoma, T-cell malignancy, multiple myeloma, acute myeloid leukemia, chronic myeloid leukemia, myelodysplastic syndrome, acute lymphoblastic leukemia, chronic lymphocytic leukemia and hairy cell leukemia. The cancer can also be genitourinary cancer, gynecological cancer, lung cancer, gastrointestinal cancer, head and neck cancer, malignant glioblastoma, malignant mesothelioma, non-metastatic or metastatic breast cancer, malignant melanoma, Merkel cell carcinoma or bone and soft tissue sarcoma, hematological neoplasia, non-small cell lung cancer (NSCLC), breast cancer, metastatic colorectal cancer, hormone-sensitive or hormone-refractory prostate cancer, colorectal cancer, ovarian cancer, hepatocellular carcinoma, renal cell carcinoma, pancreatic cancer, gastric cancer, esophageal cancer, hepatocellular carcinoma, cholangiocarcinoma, head and neck squamous cell carcinoma, soft tissue sarcoma or small cell lung cancer.

[0053] In some embodiments of the methods described herein, the immunotherapeutic agent is selected from a monoclonal antibody or a fragment thereof, a bispecific antibody, an antibody-drug conjugate, a chimeric antigen receptor (CAR)-T cell, and an anti-tumor vaccine product or a combination thereof. Immunotherapeutic agents include therapies that are intended to target different types of cancer using components of the immune system. These include antibodies, checkpoint inhibitors, immunomodulators, cytokines, monoclonal antibodies, cancer vaccines, and chimeric antigen receptor T cells approved by the FDA for cancer treatment.

[0054] Checkpoint inhibitors are immune checkpoint antibodies that work by unleashing the activity of tumor antigen-specific T cells by blocking the interaction of their inhibitory receptors with inhibitory ligands (e.g., PD-1:PD-L1 interactions). In principle, this T cell antigen-specific recognition is universal and independent of the disease type, where T cell effector memory is generated against previously encountered tumor antigens. This mechanism is applicable to infectious diseases, where humans have been previously exposed to viral infections (e.g., CMV) and are able to strengthen the recall response if they encounter the antigen again. Checkpoint inhibitors for use in the methods described herein include, but are not limited to, ipilimumab (Yervoy TM ), pembrolizumab (Keytruda TM ), Opdivo TM ), cemiplimab (Libtayo TM ), atezolizumab (Tecentriq TM ), avelumab (Bavencio TM ) and durvalumab (Imfinzi TM ). Cytokines used in the cancer treatment methods described herein include interferon alpha, intron A [2b], Alferon [2a], interleukin-2, and aldesleukin. Cancer vaccines used in the methods described herein include Gardasil, talimogenelaherparepvec (Imlygic TM ) and sipuleucel-T (Provenge TM ). Immunomodulatory drugs used in the methods described herein include, for example, thalidomide, lenalidomide, pomalidomide, and imiquimod.

[0055] In various embodiments, the immunotherapeutic agent binds to a protein selected from the group consisting of: CD22, CD19, CD20, CD33, CD37, CD38, CD123, and BCMA.

[0056] In some embodiments of the methods described herein, the method further comprises administering to the subject an agent selected from the following: antibodies against tumor-associated antigens (TAAs), chemotherapeutic agents, immunotoxic agents, vaccines, immunomodulatory drugs, immunometabolism-modifying drugs, targeted therapies, radiation, anti-angiogenic agents, CAR-T cell therapy, bispecific antibodies, T cell engagers, agents that reduce immunosuppression, or a combination thereof.

[0057] Example 1

[0058] The ability of bryostatin-1 to promote antigen expression was evaluated in vitro. Human peripheral blood mononuclear cells (PBMCs) were cultured according to previously published methods (Ramakrishna et al., 2019). Briefly, cells were washed and then treated with bryostatin-1 (1 nM) or DMSO (vehicle control) for various times ranging from 45 minutes to 24 hours as indicated. Antibody binding capacity (ABC) was measured by flow cytometry (NCI Fluidics Laboratory); and plotted directly or as a fold change relative to DMSO / vehicle control ( Figure 1 and 2 ). Figure 1 and 2 The data shown show that a 45 minute incubation produced the same antigenic rise as longer incubations.

[0059] Although the present invention has been described with reference to the above examples, it will be understood that modifications and variations are encompassed within the spirit and scope of the present invention. Accordingly, the present invention is limited only by the following claims.

Claims

1. A method for treating cancer in a subject, the method comprising administering to the subject a bryostatin compound at a dose effective to induce upregulation of a tumor antigen in the subject, thereby treating the cancer in the subject.

2. The method of claim 1, wherein the bryostatin compound is bryostatin-1 or a bryostatin analog.

3. The method of claim 1, wherein the administration time is between about 0.5 hours and 24 hours.

4. The method of claim 3, wherein the administration time is about 30 minutes, about 45 minutes, about 1 hour, about 2 hours or about 3 hours.

5. The method of claim 1, further comprising administering an immunotherapeutic agent to the subject.

6. The method of claim 5, wherein the immunotherapeutic agent is administered prior to, simultaneously with, and / or after administration of the bryostatin compound.

7. The method of claim 1, wherein the cancer is selected from the group consisting of hematologic neoplasia comprising B-cell lymphoma, T-cell malignancies, multiple myeloma, acute myeloid leukemia, chronic myeloid leukemia, myelodysplastic syndrome, acute lymphoblastic leukemia, chronic lymphocytic leukemia, and hairy cell leukemia.

8. The method of claim 5, wherein the immunotherapeutic agent is selected from a monoclonal antibody or a fragment thereof, an antibody-drug conjugate, a bispecific antibody, a chimeric antigen receptor (CAR)-T cell, an anti-tumor vaccine product, or a combination thereof.

9. The method of claim 5, wherein the immunotherapeutic agent binds to a protein selected from the group consisting of: CD22, CD19, CD20, CD33, CD37, CD38, CD123, and BCMA.

10. The method of claim 1, further comprising administering to the subject an agent selected from the following: antibodies against tumor-associated antigens (TAA), chemotherapeutic agents, immunotoxic agents, vaccines, chemotherapeutic agents, immunomodulatory drugs, immunometabolism-modifying drugs, antibody-drug conjugates, targeted therapies, bispecific antibodies, radiation, anti-angiogenic agents, CAR-T cell therapy, T cell engagers, agents that reduce immunosuppression, or a combination thereof.

11. The method of claim 10, wherein the agent is a bispecific antibody, a CAR-T cell therapy, a T cell engager, or a combination thereof.

12. The method of claim 1, wherein the subject is a human.

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