Poster abstracts
Poster number 18 submitted by Safiya Khurshid
Modulation of insulin receptor alternative splicing to develop cancer therapeutics
Safiya Khurshid (Nationwide Childrens Hospital)
Abstract:
Insulin receptor (IN-R) undergoes alternative splicing to produce two isoforms: the full-length IN-RB and exon 11 skipped IN-RA isoform. The IN-RA 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 tumor-cell hallmarks like proliferation & angiogenesis. Although this splicing event is profound and could hamper the therapeutic efficiency of drugs, very little is known about what causes it and whether we can reinstate the normal splicing pattern in the cancer cells. To determine how this change is brought about we surveyed different kinds of stress including hypoxia, heat-shock and DNA-damage. Our data shows that hypoxia causes a significant increase in the expression of IN-RA and this change is Hif1a dependent. Furthermore, we found that there is a significantly increased expression of IN-RA levels in a cohort of Rhabdomyosarcoma (RMS) patients as well as in multiple cell lines derived from these tumors as compared to the control samples. To allow the possibility for therapeutic intervention, we utilized the antisense oligonucleotide technology (ASO) to modulate the levels of IN-RA in RMS cell lines. Based on published data we designed the ASO, which binds to the cugbp binding site in the intron 10 and found that the application of ASO shifts the IN-R splicing towards IN-RB and significantly reduces proliferation, migration and angiogenesis in multiple RMS cell lines. In order to delineate the downstream consequences of this splicing change, we subjected the control and ASO treated cells to Mass spectrometric analysis. Preliminary analysis of this data reveals that proliferation markers such as Ki67, PI3-kinase pathway components, protein clusters such as cell-cell adhesion & cell division among others are significantly down regulated in cells treated with ASO where the splicing has been restored to the IN-RB isoform.
Our data opens a new paradigm of how alternative splicing regulates cell signaling to mediate cancer-causing changes and our ASO compounds show promising insight into how we can reinstate the splicing pattern and impede tumorigenesis.
Keywords: Alternative Splicing , Insulin receptor, Antisense Oligonucleotides