DNA methylation profiling of ovarian carcinomas and their in vitro models identifies HOXA9, HOXB5, SCGB3A1, and CRABP1 as novel targets

Abstract

Background: The epigenetics of ovarian carcinogenesis remains poorly described. We have in the present study investigated the promoter methylation status of 13 genes in primary ovarian carcinomas (n = 52) and theirin vitromodels (n = 4; ES-2, OV-90, OVCAR-3, and SKOV-3) by methylation-specific polymerase chain reaction (MSP). Direct bisulphite sequencing analysis was used to confirm the methylation status of individual genes. The MSP results were compared with clinico- pathological features.Results: Eight out of the 13 genes were hypermethylated among the ovarian carcinomas, and altogether 40 of 52 tumours were methylated in one or more genes. Promoter hypermethylation ofHOXA9,RASSF1A,APC,CDH13,HOXB5,SCGB3A1 (HIN-1),CRABP1, andMLH1was found in 51% (26/51), 49% (23/47), 24% (12/51), 20% (10/51), 12% (6/52), 10% (5/52), 4% (2/48), and 2% (1/51) of the carcinomas, respectively, whereasADAMTS1,MGMT,NR3C1,p14^ARF, andp16^INK4awere unmethylated in all samples. The methylation frequencies ofHOXA9andSCGB3A1were higher among relatively early-stage carcinomas (FIGO I-II) than among carcinomas of later stages (FIGO III-IV;P= 0.002,P= 0.020, respectively). The majority of the early-stage carcinomas were of the endometrioid histotype. Additionally,HOXA9hypermethylation was more common in tumours from patients older than 60 years of age (15/21) than among those of younger age (11/30;P= 0.023). Finally, there was a significant difference inHOXA9methylation frequency among the histological types (P= 0.007).Conclusion: DNA hypermethylation of tumour suppressor genes seems to play an important role in ovarian carcinogenesis andHOXA9,HOXB5,SCGB3A1, andCRABP1are identified as novel hypermethylated target genes in this tumour type.