Relationship between the loss of the retinoblastoma tumor suppressor and radiosensitivity.
34 Background: Prostate cancer is the most frequently diagnosed malignancy and the second leading cause of cancer death in U.S. men. The retinoblastoma tumor suppressor protein, RB, plays a critical role in cell cycle regulation. Loss of RB has been observed in 25–30% of prostate cancers and is correlated with increasing tumor stage and grade. The clinical consequences of RB loss are unknown. We have previously shown that RB loss results in a castrate resistant phenotype. We hypothesized that RB loss would downregulate the G1-S cell cycle arrest normally induced by irradiation, inhibit DNA repair, and subsequently sensitize cells to mitotic catastrophe. Methods: Experimental work was performed with multiple isogenic prostate cancer cell lines (hormone sensitive: LNCaP and LAP-C4 cells and hormone resistant C42 cells; stable knockdown of RB using shRNA). Gamma H2AX assays were used to quantitate DNA damage and PARP cleavage and Caspase 3 were used to quantitate apoptosis. FACS analysis with BrdU was used to assess the cell cycle. Cell survival was measured using the clonogenic cell survival assay. In vivo work was performed in nude mice with tumor xenografts. Results: We observed that loss of RB increased radioresponsiveness in both transient and clonogenic cell survival assays in all cell lines (p<0.05). Cell death was not mediated through increased apoptosis, however, there was increased cell cycling despite the presence of DNA damage in the RB knockdown cells. In vivo xenografts of the RB deficient tumors exhibited diminished tumor mass, lower PSA kinetics and decreased tumor growth after treatment with single fraction of ionizing radiation in comparison to RB intact tumors (p<0.05). Conclusions: Loss of RB results in a differential response to ionizing radiation. Isogenic cells with RB knockdown are more sensitive to DNA damage and result in reduced cell survival. RB status is integral to determining which therapeutic modality should be employed in the management of prostate cancer. No significant financial relationships to disclose.