Poster abstracts
Poster number 22 submitted by Safiya Khurshid
ASO screen uncovers splicing as a therapeutic vulnerability in the insulin-like growth factor (IGF) signaling pathway
SAFIYA KHURSHID (Nationwide Childrens Hospital, Columbus Ohio), Matias Montes (Nationwide Childrens Hospital, Columbus Ohio), Andy Goodwin (Nationwide Childrens Hospital,Columbus Ohio), Frank Rigo (Ionis Pharmaceuticals, Carlsbad, California, 92010), Peter Houghton (Greehey Childrens Cancer Research Institute, University of Texas Health Science Center, San Antonio, Texas, 78229), Dawn Chandler (Nationwide Childrens Hospital; Department of Pediatrics, The Ohio State University, Columbus, Ohio, 43210)
Abstract:
The insulin receptor (INR) undergoes alternative splicing to produce 2 isoforms: the full-length INRB and exon 11 skipped INRA isoform. The expression of these isoforms is tightly regulated during development, however there is an aberrant increase in INRA expression in cancer. INRA encodes for a receptor which has high affinity for both insulin & IGF2 growth hormones and it exploits the IGF pathway to accelerate the onset of tumors. Rhabdomyosarcoma (RMS) is the most common soft-tissue sarcoma in children and there is inadequacy of less toxic therapies.
We found a significant increase in the expression of INRA levels in a cohort of 40 RMS patients as well as multiple RMS cell lines as compared to the controls. We went on to show that cellular stress such as hypoxia increases alternative splicing to produce more INRA. To get a mechanistic insight into this phenomenon, we utilized this hypoxia-inducible splicing system & performed an antisense-oligonucleotide (ASO) screen to characterize sequence elements & splicing factors involved in the regulation of INR splicing. We found that sequence elements flanking exon 11 are critical to the increased alternative splicing we see under hypoxia. We performed a refined ASO walk to target the regions important for exon inclusion or exclusion and identified a region known to be a binding site for the splicing factor CUGBP1. RMS derived cell lines exclusively express INRA but when treated with our lead ASO compound, they show a decrease in the INRA expression. ASO treated cells exhibit a significant reduction in cell proliferation, migration & angiogenesis.
Our data shows promising insight into how we can impede the IGF2 pathway by causing the splicing shift & reducing INRA expression to consequently mitigate tumor hallmarks like cell-proliferation, migration & angiogenesis. The goal is to use these ASO compounds as therapeutic interventions in conjunction with already established anti IGF1 receptor therapies to treat RMS.
Keywords: Alternative splicing, Insulin receptor, Pediatric cancers