Frequent concomitant inactivation of miR-34a and miR-34b/c by CpG methylation in colorectal, pancreatic, mammary, ovarian, urothelial, and renal cell carcinomas and soft tissue sarcomas

The microRNA encoding genes miR-34a and miR-34b/c represent direct p53 target genes and possess tumor suppressive properties as they mediate apoptosis, cell cycle arrest, and senescence. We previously reported that the miR-34a gene is subject to epigenetic inactivation by CpG methylation of its promoter region in primary prostate cancer and melanomas, and in 110 different cancer cell lines of diverse origin. Here we analyzed the methylation status of miR-34a and miR-34b/c in additional primary tumors of divergent sites. We found methylation of miR-34a or miR-34b/c in formalin-fixed, paraffin-embedded (FFPE) tumor samples from 178 patients with the following frequencies: colorectal cancer (74% miR-34a, 99% miR-34b/c; n = 114), pancreatic cancer (64%, 100%; n = 11), mammary cancer (60%, 90%; n = 10), ovarian cancer (62%, 69%; n = 13), urothelial cancer (71%, 57%; n = 7), and renal cell cancer (58%, 100%; n = 12). Furthermore, soft tissue sarcomas showed methylation of miR-34 gene promoters in FFPE samples (64%, 45%; n = 11), in explanted, cultured cells (53%, 40%; n = 40), and in frozen tissue samples (75%, 75%, n = 8). In the colorectal cancer samples a statistically significant correlation of miR-34a methylation and the absence of p53 mutation was detected. With the exception of sarcoma cell lines, the inactivation of miR-34a and miR-34b/c was concomitant in most cases. These results show that miR-34 inactivation is a common event in tumor formation, and suggest that CpG methylation of miR-34a and miR-34-b/c may have diagnostic value. The mutual exclusiveness of miR-34a methylation and p53 mutation indicates that miR-34a inactivation may substitute for loss of p53 function in cancer.