Combination therapy targeting hypoxia inducible factor-α (HIF-1 α) and Tissue Factor (TF) by the mTOR inhibitor Rapamycin and the histone deacetylase inhibitor LBH589

1033

Tumor hypoxia plays an important role in the onset of growth factor production and subsequent new vessel formation by tumor released Vascular Endothelial Growth Factor (VEGF). VEGF production and signaling is partly dependent on mTOR induced expression of the transcription factor hypoxia inducible factor-1α (HIF-1α). The mTOR inhibitor Rapamycin was initially developed as an immunosuppressant and is now in clinical development as an anticancer agent and HIF-1α inhibitor. We have previously reported that HDAC inhibitors suppress tumor induced angiogenesis by reducing HIF-1α protein level. Based on these data, we hypothesized that the combination of Rapamycin and the HDAC inhibitor LBH589 has greater antiangiogenic and antitumor activity than single agents. Combination treatment of both endothelial cells as well as a VHL-deficient renal cell carcinoma (RCC) cell line UMRC2 (C2 cells) resulted in a significantly reduced HIF-1α expression compared to either agent alone. In addition, in an orthotopic model with C2 cells the combination of Rapamycin and LBH589 produced significant antitumor and antiangiogenic activity as measured by tumor weight and microvessel density, respectively. One of the proposed antitumor mechanisms of Rapamycin is also induction of tumor specific thrombosis by upregulation of Tissue Factor (TF), the main regulator of coagulation. TF is abundantly expressed in tumor tissue, including the tumor vasculature, and is known to promote tumor angiogenesis. We could not detect Rapamycin induced intratumoral thrombosis nor increased TF expression by immunohistochemical staining in the C2 orthotopic tumor model. Thus, we studied the effect of LBH589 on Rapamycin induced TF activity on endothelial cells in vitro and found that LBH689 completely abolished Rapamycin induced TF-activity of VEGF stimulated human umbilical vein endothelial cells. In addition, TF activity and expression in C2 cells was significantly inhibited by Rapamycin, LBH589 and combination treatment by approximately 50%. To our knowledge we report for the first time that an HDAC inhibitor prevents VEGF and Rapamycin induced TF activity. In conclusion, these results suggest that the novel combination of mTOR and HDAC inhibitors represents a rationale therapeutic strategy targeting HIF-1α and TF that warrants clinical testing in patients with advanced cancer.