Abstract
Abstract
The Rb pathway functions as a cell cycle checkpoint and deregulation of its components, commonly found in malignancies, causes progression from G1 to S phase, promoting cellular proliferation. Because of Rb's central role in checkpoint regulation, abrogation of the pathway can occur through multiple non-redundant mechanisms including Rb loss, hypermethylation or mutation, and CCND1 or CCNE amplification. Emerging evidence shows that tumour types can often be distinguished by particular alterations in one member of the pathway suggesting that different mechanisms of Rb abrogation may regulate tumour behaviour. We hypothesize that Rb pathway deregulation is frequent in HGSC, the most common and most aggressive histotype of ovarian cancer, and that the mechanism of Rb pathway deregulation identifies clinically distinct subgroups of HGSC.
Micro-dissected epithelium from HGSC and normal FTE samples were analyzed for differential gene expression using the Affymetrix U133 Plus 2.0 gene-chips, and expression values for p53, p21, p27, p16, CCND1, CCNE and Rb genes determined. Protein expression was assessed by immunohistochemistry (IHC) on tissue microarrays composed of ovarian/tubal carcinomas inclusive of the major histotypes. Digitized stained slides were quantified using automated image analysis and correlated with clinico-pathologic variables including outcome. Rb loss of heterozygosity (LOH) was tested by an Rb diagnostics protocol involving D13S153 and RB1.2 polymorphic marker analyses using PCR amplification, followed by comparisons of the tumour and its corresponding normal sample by MicroGene Clipper sequencers.
Gene expression analysis showed statistically significant over-expression of p53, CCNE E2F1/3 and p16 and down-regulation of p21 and CCND1 in HGSC compared to normal fallopian tube epithelium (p<0.001). Protein expression determined by IHC analysis of HGSC revealed a similar pattern of expression when compared to normal fallopian tube, the site of origin of this carcinoma. There were important differences in the expression of these proteins between HGSC subgroups, where up-regulation was observed for p16, CCNE, CCND1 and BIRC5 in 58.3%, 57%, 33.3%, and 58.3% respectively. Rb however, showed no statistically significant differences at the RNA level, but 40% of all HGSC profiled had a significant decrease in protein expression. Interestingly, LOH analysis revealed 76% of HGSC had Rb inactivation at the gene level. Furthermore, we observed statistically significant correlations (p=0.029) between p16 over-expression and Rb protein loss, using Fisher's Exact test.
HGSC is characterized by both genetic and protein abrogation in the Rb pathway. Additionally, we observed differences in the mechanism of this G1/S checkpoint inactivation amongst HGSC patient samples which may represent important biological/clinical differences amongst sub-groups of serous cancer patients.
Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 333. doi:10.1158/1538-7445.AM2011-333
