Dual loss of VHL and RASSF1A enhances tumorigenicity in clear cell renal cancer.

Introduction: Clear Cell Renal cell carcinoma (ccRCC) is the most common form of kidney cancer. The VHL tumor suppressor is lost or mutated in the majority of ccRCC and hereditary mutations in VHL predispose patients to ccRCC. However, another tumor suppressor, RASSF1A is inactivated at even higher levels in ccRCC. The majority of ccRCC lose function of both proteins. This suggests there may be some functional link between them. We found that the RASSF1A and VHL proteins can form a complex, suggesting they may act in concert to suppress the tumorigenic phenotype. Methods: We generated two matched sets of Renal cancer tumor cell systems where we added back or knocked down both genes. We also made compound transgenic mice with induced, dual inactivation. Results: In the add-back system we found that the proteins cooperated to suppress 3D cell growth and sensitized cells to apoptosis inducing agents. In the knockdown system, we found the double loss cooperated to promote growth in 3D. In cell line xenografts, we found that the loss of VHL alone was insufficient to promote tumorigenesis, but it acted as an accelerant on tumor growth by RASSF1A deficient cells, which showed enhanced vascularization differential deregulation of HIF1α compared to HIF2α. Discussion: This is the first report of an interaction between RASSF1A and VHL and may explain why RASSF1A is so frequently inactivated in Renal cancer. However, further studies in transgenic mice showed that dual inactivation of the tumor suppressors did not result in the formation of tumors or macrocysts. Thus, additional genetic lesions are likely required for the development of ccRCC.