VHL L169P Variant Does Not Alter Cellular Hypoxia Tension in Clear Cell Renal Cell Carcinoma.

In the current era of tumor genome sequencing, single amino acid missense variants in the von Hippel-Lindau () tumor suppressor gene are frequently identified in clear cell renal carcinoma (ccRCC). Due to the incomplete knowledge of the structural architecture of protein, the functional significance of many missense mutations cannot be assigned. L169P is one such missense mutation identified in the case of aggressive, metastatic ccRCC. Here, we characterized the biochemical activity, transcriptomic hypoxia signature and biological functions of the L169P variant. Lentiviral vector expressing either wildtype (WT) or L169P were used to transduce two -deficient human ccRCC cell lines, 786-O and RCC4. The stability of the protein and the expression level of , and were analyzed. The impact of restoring L169P or WT on the hypoxia gene expression program in 786-O cells was assessed by mRNA sequencing (RNAseq) and computed hypoxic scores. The impact of restoring expression on the growth of ccRCC models was assessed in cell cultures and in chorioallantoic membrane (CAM) xenografts. In the 786-O cells, the protein stability of L169P was comparable to WT . No obvious difference in the capability of degrading and was observed between WT and L169P in the 786-O or RCC4 cells. The hypoxic scores were not significantly different in the 786-O cells expressing either wildtype or L169P . From the cellular function perspective, both WT and L169P slowed cell proliferation in vitro and in vivo. The L169P variant is comparable to WT in terms of protein stability, ability to degrade factors and ability to regulate hypoxia gene expression, as well as in the suppression of ccRCC tumor cell growth. Taken together, our data indicate that the L169P variant alone is unlikely to drive the oncogenesis of sporadic ccRCC.