Chronic Chemoimmunotherapy Achieves Cure of Advanced Spontaneous Mammary Tumors via Persistent Blockade of Post-Therapy Counter-Regulation
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Advances in breast cancer treatment and diagnosis have led to enhanced survival and cure of primary tumors, however metastatic disease remains incurable. In many patients, breast cancer has already disseminated in the form of micro-metastases upon diagnosis, emphasizing the need for systemically effective treatments that not only target the primary tumor but also eradicate distant metastases. Cancer immunotherapy remains a promising clinical option as activated lymphocytes have the capacity to specifically kill tumor cells and to circulate throughout the body. Historically, immunotherapy has had limited success at inducing complete tumor regression due to counter-regulatory mechanisms, specifically T-suppressor cell expansion. Like other forms of cancer immunotherapy, intratumoral delivery of cytokines IL-12 and GM-CSF leads to only short-lived tumor regression in a murine mammary tumor model. IL-12/GM-CSF treatment promotes an initial anti-tumor immune response; however T suppressor cells rapidly inhibit tumor-killing effector cells, rendering them nonfunctional. Recently it has been shown that subclinical-dose chemotherapy can enhance immunotherapy treatments by specifically depleting T-suppressor cells. Therefore, in these studies we combined chemotherapy pretreatment with immunotherapy to stimulate tumor-specific immune cells to induce complete tumor regression both locally and systemically, while circumventing regulatory mechanisms. Repeated administration of cyclophosphamide and IL-12/GM-CSF, over a period of six, weeks led to complete and durable regression of spontaneous tumors, while either treatment alone was ineffective at achieving long-term cure. Importantly, whereas contribution of CD8+ T-cells to tumor regression in the immunotherapy alone group was minimal, tumor kill was strictly dependent on CD8+ T-cells in the chemoimmunotherapy group. Further analysis revealed that chemoimmunotherapy enhanced CD8+ T cell proliferation over immunotherapy alone. The addition of chemotherapy to immunotherapy also increased the cytotoxic activity of CD8+ T cells, as assessed by granzyme B and IFNγ expression. Ultimately, persistent suppression of the post-therapy T-suppressor cell rebound led to an elevated activity index (proliferating CD8+ T cell to T-suppressor cell ratio) in the tumor, which correlates with tumor regression. Overall, these data show that combination chemoimmunotherapy achieves enhanced tumor cell death by overcoming homeostatic counter-regulation, a major challenge of immune-based cancer therapies.