CpG Island Methylation within the TCF2 Promoter May Enable Epigenetic Modulation of HNF1-beta and Clear Cell Phenotype in Ovarian Clear Cell Carcinoma
J Cuff, S Huang, JP Higgins, TA Longacre, JR Pollack. Stanford University, Stanford, CA
Background: Hepatocyte nuclear factor 1-beta (HNF1-beta), a hepatic and pancreatic specific transcription factor that is encoded by the TCF2 gene, is upregulated in ovarian clear cell carcinoma gene expression arrays. To determine whether HNF1-beta expression is lineage specific or more broadly expressed by other clear cell neoplasms, we developed a lexicon of HNF1-beta protein expression in a series of anatomically and histologically diverse clear cell tumors and evaluated potential epigenetic modulation of HNF1-beta expression.
Design: A monoclonal antibody directed against HNF1-beta (clone C-20, Santa Cruz, titer 1:2000) was evaluated on an anatomic and histologically diverse (n=929) set of tissue microarray and conventional tissue sections enriched for renal and gynecologic primaries with cytoplasmic clearing (n=261). Immunostaining was scored as negative, weak or strongly positive and the significance of expression was judged using the Fisher's exact test. Single-gene methylation analysis was performed to evaluate the CpG island in the promoter region of the TCF2 gene. DNA was extracted from a variety of ovarian carcinomas with a lymphocyte control. Following bisulfite modification, methylation specific PCR with primers to the promoter region of the TCF2 gene (HNF1- beta) was used to amplify a 140 bp fragment which was analyzed by Sanger sequencing.
Results: HNF1-beta was identified in 74% (192/261) of tumors with clear cell change compared to 8.8% (59/668) of tumors without clear cell change (p<0.0001). Differential methylation was identified among subtypes of epithelial ovarian tumors. Serous histology showed a relative increase in CpG island methylation in the promoter of the TCF2 gene in comparison to clear cell, endometrioid and lymphocyte control with an intermediate degree of methylation in a poorly differentiated tumor.
Conclusions: Expression of HNF1-beta is associated with clear cell phenotype across anatomically and histologically diverse tumors. Promoter methylation may serve a permissive mechanism where absent or low level methylation is complicit in transcriptional regulation and potentially, phenotypic differentiation.
Monday, March 9, 2009 9:30 AM
Poster Session I Stowell-Orbison/Autopsy Award # 144, Monday Morning