The insulin receptor (IN-R) alternatively spliced variant A, encoding a receptor lacking exon 11, is the predominant isoform expressed in both pediatric sarcoma cells and vascular endothelial cells. Exon 11 of the IN-R gene is composed of 36 nucleotides that encode 12-amino acids that reside at the carboxyl terminus of the alpha subunit. Though only a small change in the protein composition, deletion of these 12 amino acids results in a receptor that has increased binding affinity for the IGF-2 growth factor and is exquisitely capable of responding to autocrine and paracrine signaling. Cells expressing both the IN-R A variant and the IGF-2 growth factor are therefore programmed for growth and angiogenesis, two essential steps in tumorigenesis. Furthermore, the IGF-2 can maintain angiogenesis through IN-R A when the IGF-1 receptor is blocked. We also demonstrate that hypoxia induces IN-R alternative splicing. This finding is particularly intriguing since recent studies have shown that adaptation to hypoxic environments is a hallmark of the neoplastic phenotype. We therefore hypothesize that the expression of splicing factors is altered by hypoxia leading to the generation of the IN-R A isoform, which contributes to cancer progression beyond the micrometastatic stage.
To characterize sequence elements and RNA binding proteins in the regulation of IN-R alternative splicing, we have designed a hypoxia-induced splicing system that recapitulates the IN-R splicing seen in tumors. We have shown that sequence elements flanking the regulated exon are important for the hypoxia-induced splicing changes. Furthermore, have performed an antisense oligonucleotide (ASO) walk to target these splicing enhancer and repressor sequences to interfere with IN-R splicing. We have successfully targeted the CUGBP1 negative regulator of splicing that resides in intron 10 to increase the IN-R B full-length isoform. We have shown that an ASO used to drive production of the IN-R B isoform in a rhabdomyosarcoma (RMS) cell line, which normally expresses the IN-R A isoform, affects cancer cell behavior to induce reduced motility and cell migration. We therefore postulate that modulation of the splicing of the IN-R, alone or in conjunction with IGF-1R blocking antibodies, could be used as a therapeutic intervention in pediatric RMS cells.
Citation Format: Dawn S. Chandler, Chelsea Brown, Hemant Bid, Thomas Bebee, Daniel F. Comiskey, Frank Rigo, Peter J. Houghton. Targeting insulin receptor splicing as a therapeutic intervention for pediatric rhabdomyosarcoma. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr LB-014.