Abstract 30: The integrin α6A splice variant regulates colon cancer cell proliferation and the Wnt/β-catenin pathway

Abstract

Abstract Colon cancer is a major cause of death by cancer in North America. Recently, we found that both subunits of the integrin α6β4 are up-regulated in colon primary tumours. The α6 integrin subunit exists as two distinct splice isoforms, α6A and α6B. It is the α6A isoform that was shown to be associated with the proliferative cell population of the normal human colon and up-regulated in colorectal cancer. The aim of this study was to analyze the role of integrin α6Aβ4 in human colon cancer, more precisely its effect on cell proliferation and its mechanism of regulation. To investigate the function of integrin α6Aβ4 in colon cancer, expression of splice variant α6A was specifically abolished, using shRNA technology, in colon adenocarcinoma cell lines (Caco-2/15, T84, SW480, SW620, DLD-1 and HT29). The function of splice variant α6A of the α6β4 integrin was investigated in vitro and in tumour xenografts. In all colon cancer cell lines tested, knockdown of the integrin α6A subunit resulted in a decrease of up to 90% of the α6A variant without affecting the levels of the α6B variant. Furthermore, no compensatory expression of α6Bβ4 integrin was observed following the abolition of the α6A subunit. In the majority of colon cancer cell lines, the knockdown of α6A resulted in a significant decrease in proliferation as show by BrdU incorporation. Moreover, it was accompanied by reduced activity of the Wnt/β-catenin pathway as measured by a decrease of responsive β-catenin/TCF promoter activity and increased β-catenin phosphorylation at Ser37/Thr41 by GSK3β. Preliminary data confirmed that in these cells, the Wnt/β-catenin pathway can be stimulated by pharmacological inhibition of GSK3β (SB216763), as displayed by a decrease of β-catenin phosphorylation at Ser37/Thr41 and an increase in responsive β-catenin/TCF promoter activity. Analysis of splice variants of the α6 integrin subunit in human colorectal cancer revealed that α6A was increased in 80% of tumours as compared to their matched resection margins and a significant correlation was observed between expression of α6A and c-myc in the tumours. Furthermore, to determine the role of the integrin α6A subunit on colon cancer tumorigenesis in vivo, control and α6A knockdown T84, SW620, HT29 and DLD-1 cells were injected into nude mice. Results showed that abolition of the integrin α6A subunit led to a significant reduction in tumour growth for T84, SW620 and HT29 colon cell lines. Moreover, analysis of tumours showed an increase in β-catenin phosphorylation at Ser37/Thr41 in T84 and HT29. Our results show for the first time that splice variant A of the integrin α6 subunit regulates the Wnt/β-catenin pathway to promote cancer cell growth. Since 80% of colon cancers display an up-regulation of integrin α6A expression, targeting this integrin in colon cancers may offer new therapeutic value. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 30. doi:1538-7445.AM2012-30