Different combinations of genetic/epigenetic alterations inactivate the p53 and pRb pathways in invasive human bladder cancers

Inactivation of both the pRb (pRb-cyclin D1/cyclin-dependent kinase 4/6-p16 ) and p53 (p53-p21(WAF1)-p14(ARF)) pathways is thought to be essential for immortalization in vitro and malignant transformation in vivo. We identifie d different combinations of pRb and p53 pathway alterations in 12 invasive transitional cell carcinomas (TCCs) and addressed the functional significan ce of the different combinations observed. Results showed four combinations of alterations including -pRb/-p53 (i.e., pRb inactivated in the pRb pathw ay and p53 inactivated in the p53 pathway; four TCCs), -p16/-p53 (four TCCs ), -p16/-p21(WAF1) (one TCC), and -p16/-p14(ARF) (two TCCs), These groups i nclude two new combinations (i.e., -p16/-p53 and -p16/-p21(WAF1)) not repor ted previously for TCCs, An alteration in the key components of the p53 pat hway was not detected in one invasive TCC that had inactivated p16, Note th at all four TCCs with inactivated pRb had mutant p53; thus, the combination s of -pRb/-p21(WAF1) and -pRb/-p14(ARF) were not observed. Only two of eigh t TCCs with altered p16 had concomitant p14(ARF) loss, demonstrating that s imultaneous inactivation of these two 9p21INK4a tumor suppressor genes is n ot obligatory. To determine the biological phenotypes of TCCs with differen t combinations of pRb and p53 pathway alterations, their downstream respons es to gamma radiation were studied in vitro. As expected, none of eight TCC s with mutant p53 responded to gamma radiation by elevation of p53, p21(WAF 1), or mdm2 or by cell cycle arrest. Only two of four TCCs with wild-type p 53 and wild-type pRb (the combination of -p16/-p14(ARF)) showed normal down stream responses to gamma radiation and underwent cell cycle arrest, Two TC Cs with wild-type pRb and wild-type p53 (the combination of -p16/-p21(WAF1) and one TCC with -p16) failed to show cell cycle arrest in response to rad iation. This was attributed to the absence of p21(WAF1) in one TCC. In summ ary, these data support a model of invasive bladder cancer pathogenesis in which both the pRb and p53 pathways are usually inactivated and the biology of the tumor is impacted by the mechanism of their inactivations.