Characterization of Dysregulated VHL-Mediated pRb Regulation in Clear Cell Renal Cell Carcinoma

Abstract

The von Hippel-Lindau (VHL) tumor suppressor plays a pivotal role in maintaining cellular homeostasis as a key component of E3 ubiquitin ligase complexes, which direct proteins for proteasomal degradation. Loss of VHL function, through mutation or transcriptional silencing, is a defining feature of most sporadic cases of clear cell renal cell carcinoma (ccRCC). VHL loss leads to the constitutive stabilization of its E3 ligase substrates, most notably hypoxia-inducible factor α (HIF-α), which drives ccRCC tumorigenesis through the activation of hypoxia-responsive genes. While HIF-α is well established as a driver of ccRCC, the HIF-independent functions of VHL in oncogenesis remain poorly understood. In this study, we sought to identify novel VHL targets and evaluate their therapeutic potential in ccRCC. Using proximity labeling to identify proteasome-sensitive VHL interactors, we discovered that retinoblastoma protein (pRb) is a novel substrate of VHL. Immunoprecipitation experiments revealed that VHL binds nuclear pRb under normoxic conditions in a phosphorylation-independent manner, facilitating its ubiquitin-mediated proteasomal degradation. In VHL-deficient ccRCC cells, pRb becomes hyperstabilized, leading to suppressed apoptosis and widespread transcriptional reprogramming that promotes tumorigenesis. Proteomic analysis of VHL-pRb interactors revealed an enrichment of proteins involved in genome-wide transcriptional remodeling, further implicating this pathway in ccRCC progression. To explore therapeutic strategies, we investigated pharmacological targeting of the VHL-pRb axis. Our findings revealed a pRb-dependent interaction between the CDK4/6 inhibitor Abemaciclib and the E2F1 activator β-lapachone, resulting in decreased viability across various ccRCC cell lines. While these results suggest a promising therapeutic approach, in vivo validation remains necessary. Overall, my work uncovers a previously unrecognized VHL-pRb pathway, providing new mechanistic insights into ccRCC pathogenesis and identifying a potential therapeutic strategy to mitigate tumor progression.