HES6 Interacts with Notch Signaling in Prostate Cancer Progression
Filipe Carvalho, Ashley E Ross, Luigi Marchionni, Edward M Schaeffer, David M Berman. Johns Hopkins University School of Medicine, Baltimore, MD
Background: Although a leading cause of cancer death, more than 80% of men with prostate cancer die of unrelated causes. To avoid overtreatment and more effectively treat when needed, differences between indolent and lethal prostate cancer need to be clarified. The Gleason grading system best predicts the lethal potential of prostate cancers, but its molecular basis is not understood. We recently published a meta-analysis indicating that members of the Notch signaling pathway, particularly the Hairy and Enhancer Split 6 (HES6) transcription factor, distinguish prostate cancers with high Gleason grade. HES6 is known to facilitate invasion and migration in brain tumors and to modulate levels of Notch signaling during embryonic neurogenesis. Here we use prostate cancer cell lines to explore the function of HES6 in tumor growth, tumor cell migration, and Notch signaling.
Design: Using a custom TaqMan real time reverse transcription–polymerase chain reaction (qRT-PCR) assay, we compared expression of all Notch pathway components in benign prostate epithelial (PRE) cells to those in PCa cell lines - 22Rv1, LNCaP, DU145, PC3. We further confirmed differential expression using additional qRT-PCR assays. We up- or down-regulated HES6 using plasmid-mediated gene transfer or small interfering RNA (siRNA). We measured expression of HES1, a well-known readout of Notch pathway activity. We performed immunohistochemical staining for HES6 in benign, low-grade and high-grade tumors.
Results: HES6 transcripts were 4.5-fold higher in cancer cells compared with benign cells. Engineered HES6 overexpression conferred a modest proliferative advantage to both benign and cancer cells (p=0.03), whereas siRNA-mediated HES6 siliencing decreased proliferation, most notably in cancer cells (p=0.01). Compared to control cells, PC3 cells overexpressing HES6 migrated dramatically faster. In most PCa lines, HES6 induced HES1 expression, indicating that HES6 activates the Notch pathway. Immunohistochemistry confirmed an increase in strong nuclear HES6 expression with increasing Gleason grade.
Conclusions: HES6 expression confers growth and migration advantages to prostate cells, supporting a function for this protein in prostate carcinogenesis. This function may result from a direct action of HES6 on transcriptional targets or from cross-talk with other members of the Notch pathway. Since HES6 is overexpressed in high-grade tumors, Notch signaling, and particularly HES6, may be useful in distinguishing indolent from lethal prostate cancers and may be useful targets in those cases that require treatment.
Wednesday, March 21, 2012 9:30 AM
Poster Session V # 269, Wednesday Morning