Alterations of the p16-pRb pathway and the chromosome locus 9p21-22 in non-small-cell lung carcinomas - Relationship with p53 and MDM2 protein expression

The p16-pRb and p53-MDM2 pathways represent vital cell cycle checkpoints. R ecent studies provide evidence that these pathways are directly linked via MDM2-pRb interaction and p53 suppression of the RBI gene. In the present st udy we investigated the alterations of this G1 phase protein network using immunohistochemical and molecular methods in a series of 68 non-small-cell lung carcinomas (NSCLCs) and correlated the findings with clinicopathologic al features and prognosis of the patients. Aberrant expression (Ab) of p16 and pRb was observed in 33 (49%) and 27 (40%) of the carcinomas, respective ly. Analysis of the region that encodes for p16 by deletion mapping, a poly merase chain reaction (PCR) based methylation assay and PCR single-strand c onformation polymorphism (SSCP) analysis revealed that deletions and transc riptional silencing by methylation might represent the main mechanisms of C DKN2/p16(ink4a) inactivation in NSCLCs, The results of deletion mapping als o suggest that other tumor suppressor genes may reside at the 9p21-22 regio n, which encodes for CDKN2/MTS1/p16(ink4a), r14(ARF), and MTS2/p15(ink4b), In addition, microsatellite instability was observed with a frequency of 16 % in the 9p21-22 chromosome area. Overexpression (P) of p53 and MDM2 protei ns was found in 39 (58%) and 47 (70%) of the cases, respectively, A highly significant association was observed between p53 overexpression and p53 mut ations (P = 0.006), Statistical analysis of the expression patterns of the biologically relevant molecules (p16/pRb, p53/MDM2, MDM2/pRb, and p53/pRb) showed coincident overexpression of p53 and MDM2 (P = 0.04) and that abnorm al pRb was correlated with elevated levels of MDM2 (P = 0.013) and p53 (P = 0.01), respectively, We suggest that deregulated expression of these molec ules may act synergistically. An important finding of the study was that mu ltiple impairments (three and four molecules affected) of the p16/pRb/p53/M DM2 network occurred in a large proportion (43%) of the carcinomas, This fi nding in addition to the absence of correlation with clinical stage of the tumors suggests that multiple hits of this network may be a relatively earl y event in the development of a subset of NSCLCs, The relationship between the factors examined in the present study, clinicopathological features, an d survival of the patients did not reveal any significant correlations with the exception of smoking, which was associated with microsatellite alterat ions (loss of heterozygosity and microsatellite instability) at the 9p21-22 locus (P = 0.04) and the immunophenotypes p53(P)/MDM2(P) (P = 0.04) and p1 6(Ab)/pRb(Ab)/p53(P)/MDM2(P) (P = 0.03), respectively. We suggest that in a subset of NSCLCs, simultaneous deregulation of the members of this network may represent one way of initiating the oncogenic procedure whereas in oth er NSCLC subgroups alternative pathways may play this role.