Distinguished University Professor Paul A. Bunn; Jr. Endowed Chair in Cancer Research Professor of Surgery and Pharmacology Director, University of Colorado Cancer CenterView Slides
RalA and RalB are small GTPases that support malignant development and progression in experimental models of lung, pancreatic, colon, prostate, and bladder cancer. However, demonstration of their clinical relevance in human tumors remained lacking. Hence, we developed tools to evaluate Ral protein expression, activation, and transcriptional output and evaluated their association with clinicopathologic parameters in common human tumor types. To determine the relevance of Ral activation and transcriptional output, we identified and evaluated a transcriptional signature of genes that correlates with depletion of RalA and RalB in vivo. The Ral transcriptional signature score, but not protein expression as evaluated by immunohistochemistry, predicted disease stage, progression to muscle invasion, and survival in human bladder cancers and metastatic and stem cell phenotypes in bladder cancer models. In prostate cancer, the Ral transcriptional signature score was associated with seminal vesicle invasion, androgen-independent progression, and reduced survival.
Together, Dan’s team’s findings demonstrated for the first time the clinical relevance of Ral in several human cancer types and provided a rationale for the development of Ral-directed therapies. Therefore, they used protein structure analysis and virtual screening to identify drug-like molecules that bind a site on the GDP-form (inactive) of Ral. Compounds RBC6, RBC8, and RBC10 inhibited Ral binding to its effector RalBP1, Ral-mediated cell spreading in murine fibroblasts and anchorage-independent growth of human cancer cell lines. Binding of RBC8 derivative BQU57 to RalB was confirmed by isothermal titration calorimetry, surface plasma resonance and 15N-HSQC NMR. RBC8 and BQU57 show selectivity for Ral relative to Ras or Rho and inhibit xenograft tumor growth similar to depletion of Ral by siRNA. These results show the utility of structure-based discovery for development of therapeutics for Ral-dependent cancers.