The Insulin Receptor Substrate 1 (Irs1) in Intestinal Epithelial Differentiation and in Colorectal Cancer

Diana L Esposito,Rossano Lattanzio,Lavinia Vittoria Lotti,Antonio Moschetta,Reza Malekzadeh,Federica Aru,Annalisa Morgano,Michela Abbondanza,Antonio Russo,Rosa Valanzano,Faraz Bishehsari,Renato Mariani-Costantini,Mauro Piantelli,Venugopalan Cheriyath

doi:10.1371/journal.pone.0036190

Abstract

Colorectal cancer (CRC) is associated with lifestyle factors that affect insulin/IGF signaling, of which the insulin receptor substrate 1 (IRS1) is a key transducer. We investigated expression, localization and pathologic correlations of IRS1 in cancer-uninvolved colonic epithelium, primary CRCs with paired liver metastases and in vitro polarizing Caco2 and HT29 cells. IRS1 mRNA and protein resulted higher, relative to paired mucosa, in adenomas of familial adenomatous polyposis patients and in CRCs that overexpressed c-MYC, ß-catenin, InsRß, and IGF1R. Analysis of IRS1 immunostaining in 24 cases of primary CRC with paired colonic epithelium and hepatic metastasis showed that staining intensity was significantly higher in metastases relative to both primary CRC (P<0.01) and colonic epithelium (P<0.01). Primary and metastatic CRCs, compared to colonic epithelium, contained significantly higher numbers of IRS1-positive cells (P = 0.013 and P = 0.014, respectively). Pathologic correlations in 163 primary CRCs revealed that diffuse IRS1 staining was associated with tumors combining differentiated phenotype and aggressive markers (high Ki67, p53, and ß-catenin). In Caco 2 IRS1 and InsR were maximally expressed after polarization, while IGF1R was highest in pre-polarized cells. No nuclear IRS1 was detected, while, with polarization, phosphorylated IRS1 (pIRS1) shifted from the lateral to the apical plasma membrane and was expressed in surface cells only. In HT29, that carry mutations constitutively activating survival signaling, IRS1 and IGF1R decreased with polarization, while pIRS1 localized in nuclear spots throughout the course. Overall, these data provide evidence that IRS1 is modulated according to CRC differentiation, and support a role of IRS1 in CRC progression and liver metastatization.

Highlights

Colorectal cancer (CRC) has been linked to lifestyle risk factors, most notably diets based on energy-dense foods and low physical activity [1,2,3]
To explore the modulation of insulin receptor substrate 1 (IRS1) and of other insulin/insulin-like growth factors (IGFs) pathway components, we assessed by western blot the protein levels of IRS1, InsRß, IGF1Rß and ß-catenin in 7 of the 8 abovereported CRC cases, and in paired colonic mucosa and adenoma samples from two unrelated familial adenomatous polyposis (FAP) patients [33]
In the two FAP cases, IRS1 markedly increased in the adenoma relative to mucosa, together with InsRß, IGF1Rß and ß-catenin, and IHC showed diffuse IRS1 in adenomas (Figure S1)

Summary

Introduction

Colorectal cancer (CRC) has been linked to lifestyle risk factors, most notably diets based on energy-dense foods and low physical activity [1,2,3]. Epidemiological and experimental evidences indicate that the hormone insulin and the insulin-like growth factors (IGFs) 1 and 2 play key role(s) in mediating the complex effect(s) of diet and exercise on CRC risk [4,5,6,7,8,9,10]. Over-expression of the insulin receptor (InsR) and of the closely related IGF1 receptor (IGF1R) is critical for insulin/IGF system over-activation in cancer [4,7,9]. APC-mutated cells show high cytoplasmic and nuclear ß-catenin; the latter, after binding to TCF/LEF transcription factors, forms complexes that, by switching on several cancer-related genes, impose a proliferative crypt progenitor phenotype APC-mutated cells show high cytoplasmic and nuclear ß-catenin; the latter, after binding to TCF/LEF transcription factors, forms complexes that, by switching on several cancer-related genes, impose a proliferative crypt progenitor phenotype (reviewed at http://www.stanford. edu/,rnusse/wntwindow.html) [17]

Methods

Results

Conclusion