Silencing of the Insulin Receptor Isoform A Favors Formation of Type 1 Insulin-Like Growth Factor Receptor (IGF-IR) Homodimers and Enhances Ligand-Induced IGF-IR Activation and Viability of Human Colon Carcinoma Cells

Abstract

Insulin receptor (IR) overexpression is common in cancers, with expression of the A isoform (IR-A, exon 11-) predominating over the B isoform. The IR-A signals a proliferative, antiapoptotic response to IGF-II, which itself can be secreted by tumors to establish an autocrine proliferative loop. Therefore, IGF-II signaling via the IR-A could mediate resistance to type 1 IGF receptor (IGF-IR) inhibitory drugs that are currently in development. This study addressed the role of the IR-A, using a small interfering RNA-based approach in SW480 human colon adenocarcinoma cells that coexpress the IGF-IR. Clonogenic survival was inhibited by depletion of the IGF-IR but not the IR-A, and dual receptor depletion had no greater effect than IGF-IR knockdown alone, suggesting that the IR-A could not compensate for IGF-IR loss. IGF-IR knockdown also resulted in a decrease in viability, whereas IR-A depletion resulted in increased viability. Consistent with this, upon IR-A depletion, we found a concomitant enhancement of IGF-IR activation by IGF-I and IGF-II, reduced formation of IGF-IR:IR-A hybrid receptors and increased IGF-IR homodimer formation. Together, these results suggest that IGF bioactivity is mediated more effectively by the IGF-IR than by the IR-A or receptor hybrids and that signaling via the IGF-IR is dominant to the IR-A in colon cancer cells that express both receptors.

Elevated SRPK1 lessens apoptosis in breast cancer cells through RBM4-regulated splicing events

“…Conventional approaches have revealed a close correlation between a high IR-A:IR-B ratio and malignant features of breast cancer, such as high proliferation and poor prognosis (Costa et al 2008;Mester and Redeuilh 2008;Brierley et al 2010). However, the comprehensive molecular mechanism involved in the isoform shift of IR transcript remains unclear.…”

Insulin receptor isoform switching in intestinal stem cells, progenitors, differentiated lineages and tumors: evidence that IR-B limits proliferation

“…Recently, there has been interest in the potential for IR-A to compensate for IGF1R inhibition in tumor cells (Brierley et al, 2010;Buck et al, 2010;Ulanet et al, 2010). Our finding that IR-B expression can reduce proliferation of aggressively growing SW480 CRC cells, as well as Caco-2 cells, indicate that it will be of considerable interest to test the impact of IR-B overexpression on in vivo tumorigenicity of CRC cells that normally express primarily IR-A in xenograft assays, as well as their responsiveness to IGF1R inhibitors.…”

“…Fragments 210 bp (IR-A) and 246 bp (IR-B) were resolved on 2.5% agarose gels. For human cells, IR-A and IR-B mRNAs were examined using primers 59 and 39 to exon 11 (present in IR-B) and modified from a published protocol (Brierley et al, 2010): denaturation for 5 minutes at 92˚C, 30 cycles of 92˚C for 30 seconds, annealing at 60˚C for 30 seconds, and extension at 72˚C for 30 seconds. Primers were forward in exon 10, 59-GAATGCTGCTCCTGTCCAAA-39 and reverse in exon 12, 59-TCGTGGGCACGCTGGTCGAG-39.…”

Biochemical Characterization of Individual Human Glycosylated pro-Insulin-like Growth Factor (IGF)-II and big-IGF-II Isoforms Associated with Cancer

“…Cell Proliferation Assay-Cellular proliferation assays were conducted essentially as previously described (27) utilizing the CellTiter Glo Luminescent Cell Viability Assay (Promega) according to the manufacturer's instructions.…”