Preconditioning of the tumor microenvironment by means of low dose chemotherapies for an effective immunotherapy of breast cancer

Breast cancer mortality is mainly due to distant recurrence of the disease arising from dormant tumor cells established by cancer therapies. Patients who initially respond to cancer therapies often succumb to distant recurrence of the disease. It is not clear why people with the same type of breast cancer respond to treatments differently; some escape from dormancy and relapse earlier than others. In addition, some tumor clones respond to immunotherapy while others do not. We investigated how autophagy plays a role in accelerating or delaying recurrence of neu overexpressing mouse mammary carcinoma (MMC) following adriamycin (ADR) treatment, and in affecting response to immunotherapy. We explored two strategies: 1) transient blockade of autophagy with chloroquine (CQ), which blocks fusion of autophagosomes and lysosomes during ADR treatment, and 2) permanent inhibition of autophagy by a stable knockdown of ATG5 (ATG5KD), which inhibits the formation of autophagosomes in MMC during and after ADR treatment. We found that while CQ prolonged tumor dormancy, but that stable knockdown of autophagy resulted in early escape from dormancy and recurrence. Interestingly, ATG5KD MMC contained an increased frequency of ADR-induced polyploid-like cells and rendered MMC resistant to immunotherapy. On the other hand, a transient blockade of autophagy did not affect the sensitivity of MMC to immunotherapy. Our observations suggest that while chemotherapy-induced autophagy may facilitate tumor relapse, cell-intrinsic autophagy delays tumor relapse, in part, by inhibiting the formation of polyploid-like tumor dormancy. Although immunotherapy of breast cancer by means of anti-HER2 antibodies prolongs survival of breast cancer patients, disease recurrence remains a major challenge. On the other hand administration of human vaccines against infectious disease in a preventive setting or during latency/dormancy has been successful in offering a cure. Here, we sought to use adoptive immunotherapy (AIT) at the time of tumor dormancy in order to prevent progression of breast cancer. We used a low dose immunogenic chemotherapy by means of 5-FU, Adriamycin, and Cyclophosphamide (FAC) in order to stabilize tumor progression prior to AIT using autologous tumor-reactive lymphocytes. Low dose FAC established local tumor dormancy, inhibited distant tumor dormancy occurring long before distant metastasis, and induced predominate a Ki67- quiescent type of tumor dormancy, which is less susceptible to tumor immunoediting. Dormant tumor cells expressed the cell survival pathways, including the endothelin receptor/ligand (ETRA, ETRB and ET-1) and PD-L1, thereby protecting them from elimination by AIT. In addition, tumor-reactive CD8+ T cells also produced ET-1 as a survival ligand for ETRA positive tumor cells. A combination of AIT with the blockade of tumor cell survival pathways resulted in a significant improvement of AIT against tumor dormancy. We also showed that the inhibition Bcl-xL downstream of the tumor cell survival pathways is specifically effective against dormant tumor cells, suggesting a combination of AIT with small molecules inhibitors of Bcl-xL. Altogether, we showed that distant tumor dormancy is established long before distant recurrence of breast cancer, and that the expression of several tumor cell survival pathways in dormant cells protects them from immunotherapy. Our results suggest that immunotherapeutic targeting of tumor dormancy combined with the blockade of a common downstream cell survival pathway could prevent tumor progression and recurrence of the disease