Abstract A44: Suppression of insulin-induced fatty acid synthase gene expression and colon cancer cell proliferation by members of the Krüppel-like family of transcription factors
Authors: Brown Adam R. , Simmen Rosalia CM , Simmen Frank A. 
Published: June 2012

Obesity and the associated hyperinsulinemia are major risk factors for colorectal cancer (CRC). The Fatty Acid Synthase (FASN) gene encodes the protein responsible for de novo synthesis of fatty acids. FASN mRNA and protein are insulin-induced in the colonic epithelium, over-expressed early in CRC, and positively associated with tumor cell proliferation, cell survival and chemo-resistance. FASN and its transcriptional regulators may therefore provide mechanistic links between obesity and CRC predisposition and may be exploitable for CRC prevention. Recent attention has focused on several members of the Krüppel-like family (KLF) of transcription factors in pathogenesis of CRC. Published studies have demonstrated that Krüppel-like Factors 4 and 6 (KLF4, KLF6) are important suppressors of CRC development. Recent work from our laboratory and others has implicated KLF9 as a third potential KLF protein that may be a tumor suppressor in colon and rectum. In the present study, we evaluated the possible actions of KLF9 and KLF4 to oppose the inductions by insulin of colon cancer cell FASN expression and proliferation. Addition of insulin to cell medium to mimic hyperinsulinemia (5nM-25nM, compared to vehicle) led to an increase in proliferation (13–25%, P&lt;0.05, evaluated by MTT assay) of the human colorectal cancer cell line HT-29. An increased insulin level also led to induction of FASN mRNA abundance (P=0.018). Gene expression and cell proliferation were examined after transfection of HT-29 cells with siRNAs targeting KLF4, KLF9, and KLF13 (the latter is highly similar in primary sequence to KLF9). Knockdown of KLF9 caused induction of FASN at both the mRNA (P&lt;0.001) and protein (P=0.01) levels. FASN mRNA levels also increased with siRNA knockdown of KLF13 (P=0.001) and tended (P=0.075) to increase with KLF4 knockdown. Hyperinsulinemia elicited a greater increase in proliferation of cells transfected with KLF9 siRNA than of cells transfected with control (scrambled) siRNA. While transfection of KLF9 expression vector produced a trend of reduced proliferation (P=0.082; compared to empty vector), hyperinsulinemia rescued cells from this effect. A hyperinsulinemic environment coupled with siRNA knockdown of KLF9 augmented FASN mRNA expression, compared to KLF9 knockdown alone. Increased insulin concentrations caused a marked induction of KLF4 but not KLF9 or KLF13, mRNA expression. As KLF4 has been shown to inhibit proliferation of colorectal cancer cells, it is possible that induction of KLF4 by insulin may constitute a counter-regulatory mechanism to control their proliferation. In conclusion, our findings implicate KLF family members KLF9, KLF13 and KLF4 in transcriptional repression of proto-oncogenic FASN in CRC cells. They also highlight the potential cancer-suppressive roles of KLF proteins in normo- and hyperinsulinemic conditions, which clearly warrant further investigation. Supported by NIH RO1 CA136493 and grants from the Arkansas Biosciences Institute and Sturgis Foundation.

Citation Information: Cancer Prev Res 2011;4(10 Suppl):A44.