VEGF receptor inhibitors block the ability of metronomically dosed cyclophosphamide to activate innate immunity-induced tumor regression

In metronomic chemotherapy, frequent drug administration at lower than maximally tolerated doses can improve activity while reducing the dose-limiting toxicity of conventional dosing schedules. Although the antitumor activity produced by metronomic chemotherapy is attributed widely to antiangiogenesis, the significance of this mechanism remains somewhat unclear. In this study, we show that a 6-day repeating metronomic schedule of cyclophosphamide administration activates a potent antitumor immune response associated with brain tumor recruitment of natural killer (NK) cells, macrophages, and dendritic cells that leads to marked tumor regression. Tumor regression was blocked in nonobese diabetic/severe combined immunodeficient (NOD/SCID-γ) mice, which are deficient or dysfunctional in all these immune cell types. Furthermore, regression was blunted by NK cell depletion in immunocompetent syngeneic mice or in perforin-deficient mice, which are compromised for NK, NKT, and T-cell cytolytic functions. Unexpectedly, we found that VEGF receptor inhibitors blocked both innate immune cell recruitment and the associated tumor regression response. Cyclophosphamide administered at a maximum tolerated dose activated a transient, weak innate immune response, arguing that persistent drug-induced cytotoxic damage or associated cytokine and chemokine responses are required for effective innate immunity-based tumor regression. Together, our results reveal an innate immunity-based mechanism of tumor regression that can be activated by a traditional cytotoxic chemotherapy administered on a metronomic schedule. These findings suggest the need to carefully evaluate the clinical effects of combination chemotherapies that incorporate antiangiogenesis drugs targeting VEGF receptor.