Inactivation of the retinoblastoma gene in human bladder and renal cell carcinomas

The retinoblastoma (RB) gene was the first tumor suppressor gene isolated and its inactivation is associated with the pathogenesis of several types of human cancer. In this study, we investigated the involvement of the RB gene in bladder and renal cell carcinomas by determining the loss of heterozygosity (LOH) at the RB locus and by DNA, RNA, and protein analysis of the RB gene. Whenever possible, the latter included Western blotting and immunohistochemical staining of the RB protein. In bladder carcinoma, 2 of the 8 cell lines we studied had an inactivated RB gene; one cell line lacked RB expression without a gross RB deletion, whereas the other cell line expressed only the underphosphorylated form of the RB protein. None of 16 low-grade noninvasive bladder carcinomas showed an alteration in RB protein by direct Western blot analysis, whereas 2 of 14 high-grade, invasive tumors had no RB protein as measured by both Western blotting and immunohistochemical staining. This suggests that the loss of RB function may be more important in the progression of bladder cancer than in its initiation, although more extensive studies are required. LOH within the RB locus was observed in 5 of 27 informative cases of primary bladder, ureter, or renal pelvis carcinoma. However, none of the 5 cases with LOH at the RB locus had a functional loss of RB protein expression. In renal cell carcinoma, one of the 12 cell lines had a gross homozygous deletion of the RB gene, and 2 of 32 primary tumors were negative for RB protein expression. LOH at the RB locus also was found in only 2 of 30 informative cases, one of which lacked RB expression. These results are the first to demonstrate the involvement of RB inactivation in the development of advanced primary bladder carcinoma and suggest that RB loss could have a role in certain renal cell carcinomas. Our data, however, show no correlation between LOH at the RB locus in bladder cancer and actual inactivation of the RB gene at the protein level. This may suggest that there is a second tumor suppressor or recessive cancer gene on chromosome 13 in bladder cancer and/or that the mechanism of RB inactivation in bladder cancer frequently involves independent mutations of each RB allele.