The short arm of chromosome 3 (3p) contains several critical regions that have increased frequencies of allelic deletions and harbor a set of tumor suppressor genes. In particular, the range of functions performed by RASSF1A (LUCA region, 3p21.31) includes those potentially associated with carcinogenesis. Among 3p genes, RASSF1A has the highest methylation frequency in epithelial tumors of various locations. For the first time, two different methods (methylation-specific PCR and methylation-sensitive restriction analysis) independently showed that the methylation level of the CpG island in the RASSF1A promoter region significantly correlated with grade and clinical stage of clear cell renal cell carcinoma (RCC). An analysis of 23 3p polymorphic markers in a representative set of 80 RCC cases characterized clinically and histologically revealed that RCC progression significantly correlated with the frequency of allelic imbalances in some critical regions of 3p (LUCA and AP20), but not in 3p as a whole. These data suggest that RCC progression is associated with the methylation of the RASSF1A promoter and, possibly, with structural and functional alterations in other 3p genes. In addition, significant correlation between RASSF1A methylation and allelic losses at the nearby polymorphic marker locus suggests the “two hit” model for the inactivation of this tumor suppressor gene in RCC.