BACKGROUND

Cancer, also known as malignant neoplasm, is characterized by an abnormal growth of cells that display uncontrolled cell division, invasion and destruction of adjacent tissues, and sometimes metastasis to other locations in the body. There are more than 100 types of cancer, including breast cancer, skin cancer, lung cancer, colon cancer, prostate cancer, and lymphoma. Cancer is the second leading cause of death in America and it causes about 13% of all deaths. Cancer may affect people at all ages, even fetuses, but the risk for most types of cancer increases with age.

Tumor immunoevasion is an advanced phase of cancer immunosurveillance in which tumor cells acquire the ability to circumvent host immune systems and exploit protumorigenic inflammation. The interaction between tumor cells and host immune cells plays an important role in multiple stages of tumorigenesis, and evidence suggests that host immune responses are a factor in the clinical outcome of cancer patients. Manipulation of the endogenous immune system has emerged as an effective anticancer therapy in patients with advanced cancer. (Mellman et al. (2011) Nature 480: 480-489; Rosenberg (2012) Sci Transl Med 4: 127ps8).

The tumor microenvironment has a significant impact on the functional properties of immunoregulatory components that regulate whether host responses promote or antagonize tumor growth. Tumor cells and tumor-infiltrating lymphocytes adopt strategies to evade antitumor processes and may enhance the metastatic potential through the activation of chronic inflammatory signals. The innate and adaptive immune systems play a critical role in detecting and eliminating transformed cells by activating multiple sets of myeloid cells and lymphocytes. Tumor-infiltrating immune cells also contribute to tumor progression by triggering tumor angiogenesis and immune suppression.

Tumor immunoevasion has multiple impacts on tumorigenesis by triggering tolerogenic responses to tumor cells and by exploiting pro-tumor inflammation. Phosphatidylserine (PS) is a constitutive anionic plasma membrane phospholipid selectively exposed on the surface of apoptotic or stressed cells. Its presence is widely believed to be an “eat me signal”, through which the innate immune system, including NK cells, T-cells and monocytes/macrophages recognize and remove target cells. However it has also been observed that contact with PS expressing tumor cells strongly activates immunosuppressive regulatory T-cells, and thus may provide a mechanism by which tumor cells evade recognition by the immune system.

Biological therapies hold tremendous potential for the treatment of cancers, yet there still exists a need for a therapy that provides for effective killing of tumor cells through endogenous biological systems. The present invention addresses this need.

SUMMARY OF THE INVENTION

Methods are provided for treatment of cancer by administering an effective dose of an agent that blocking the binding sites of phosphatidylserine (PS) on tumor cells. In some embodiments the blocking agent is a protein that binds to PS, thereby competitively blocking the available binding sites. Polypeptides of interest include, without limitation, annexin V protein and PS binding fragments derived therefrom, and include wild-type and mutant (e.g. annexin V-128) sequences. Annexin V protein is of particular interest.

In some embodiments of the invention, an effective dose of annexin V protein is administered, e.g. by parenteral administration, locally or systemically to an individual with cancer, where cancers include, without limitation carcinomas, sarcomas, lymphomas, leukemias, gliomas, melanomas, etc. In some embodiments the administration is performed by continuous systemic infusion. In some embodiments such continuous systemic infusion comprises intraperitoneal infusion. In some embodiments, an osmotic pump is deployed. In some embodiments the cancer is a solid tumor, including without limitation carcinomas.

Without being bound by any theory, the subject methods indicate that exogenously administered annexin V has an anti-tumor effect through the binding of PS on tumor cells, thereby blocking T-reg immunosuppression of the immune system. Local immunosuppression via PS expression is a feature of many tumors including aggressive forms of breast cancer (triple negative), neuroblastoma, and head and neck cancer. The methods of the invention provide a well-tolerated blockade of PS, that enhances host immune response to tumor.

In some embodiments of the invention, an effective dose of annexin V protein is administered parenterally to an individual diagnosed with cancer, where the administration may be intra-tumoral, i.v., i.p., or the like, in particular intraperitoneal continuous infusion. The effective dose in a human may be up to about 50 μg/kg, up to about 100 μg/kg, up to about 250 μg/kg, up to about 500 μg/kg, up to about 750 μg/kg, up to about 1 mg/kg, up to about 1.5 mg/kg, up to about 2 mg/kg, up to about 5 mg/kg, up to about 7.5 mg/kg, up to about 10 mg/kg, up to about 20 mg/kg.

The effective dose of annexin V may be combined with other treatment modalities, including without limitation chemotherapeutic drugs, radiation therapy, anti-cancer biologic agents such as monoclonal antibodies directed to tumor antigens, VEGF, etc.; and the like. In some embodiments a synergistic effect is observed when the annexin V therapy is combined with an anti-cancer biologic agent. In some embodiments, the biologic agent is an antibody that specifically binds to a “checkpoint inhibitor”. In other embodiments, the treatment modality is radiation, which may be locally administered.

In some embodiments, the Annexin V protein is administered in a manner that provides for prolonged blood clearance of the protein, for example where the half-life of the protein in circulation is at least about 30 minutes, at least about 1 hour, at least about 1.5 hours, at least about 2 hours, at least about 2.5 hours, at least about 3 hours or more. In some embodiments the manner of administration is intra-peritoneal injection, or osmotic pump. In other embodiments the route of administration is intra-venous injection over an extended period of time, for example where a daily dosage as described above is delivered over a period of up to 30 minutes, up to one hour, up to 2 hours, up to 4 hours, up to 6 hours, up to 8 hours, up to 12 hours, up to 16 hours, up to 24 hours.

In some embodiments of the invention, the presence of excess PS on cancer cells is imaged with an annexin V reagent, where an individual with a tumor positive for expression of surface PS is then selected for treatment with the methods of the invention. In some embodiments of the invention, the presence of cancer stem cells expressing surface PS is determined prior to treatment. The methods of the invention also find use in blocking immunosuppression induced by cancer stem cells. Stem cells also express levels of PS to avoid immune-recognition and therefore can also be treated with annexin V.

Another aspect of the present invention relates to the use of an annexin V agent in the manufacture of a medicament to stabilize, prevent or reduce tumor growth, including methods of reducing the growth of cancer stem cells. The medicament may be administered to an individual diagnosed with PS positive cancer cells.

Still another aspect of the present invention provides a kit to stabilize, prevent or reduce tumor growth. The kit includes a therapeutic annexin V agent, which blocks PS on the surface of cancer cells, including cancer stem cells, in an amount sufficient to stabilize, prevent or reduce disease. The kit may also include reagents for phenotyping cancers for cell surface expression of PS. The kit may also instructions for use, reagents for monitoring cancer, and the like.

