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Insulin receptor substrate 4 (IRS-4) is an adaptor protein for which new evidence suggests plays a role in tumour promotion. We described nuclear IRS-4 in RKO colon cancer cell lines in biopsies of patients with colorectal cancer (CRC) (n = 20) and in matched adjacent normal colorectal (MANC) tissue (n = 20). Treatment with physiological doses of IGF-1 promoted nuclear influx of IRS-4 from cellular cytosol in RKO cells. When exogenous IRS-4 was overexpressed in RKO cells, there was an increase in cyclin D1, cyclin E, E2F1, pRB Ser 809/811 and pRB Ser 705 levels compared with the empty vector-transfected cells. Some of these changes returned to control values after wortmannin treatment. Subcellular fractionation showed an overexpression of IRS-4 in the cytoplasm, membrane, and nuclei of tumour samples, whereas the levels of the protein were barely detectable in the three compartments of normal samples. Immunohistochemical studies showed positive nuclear IRS-4 staining in over 74% of the tumour cells. IRS-4 was strongly overexpressed in tumoural tissues from CRC patients compared to MANC tissues. The up-regulation of IRS-4 in CRC samples correlated significantly with the increase of several G1 checkpoint proteins including cyclin D1 (r = 0.6662), Rb (r = 0.7779), pRb Serine 809/811 (r = 0.6864), pRb serine 705 (r = 0.6261) and E2F1 (r = 0.8702). Taken together, our findings suggest that IRS-4 promotes retinoblastoma–cyclin-dependent kinase activation and it may serve as a pharmacological target since its expression is very low in normal tissue, including colonic epithelium.