Hepatocellular Carcinoma (HCC) is predicted to be the sixth most commonly diagnosed cancer and the fourth leading cause of cancer death worldwide. Rates of both incidence and mortality are 2 to 3 times higher among men and thus liver cancer ranks second in terms of deaths for males. In the United States alone, an estimated 42,810 adults (31,762 men and 11,048 women) will be diagnosed with primary liver cancer in 2020. These statistics, combined with the fact that the death rate of liver cancer has increased by 43% in the last decade, necessitates unconventional treatment approaches. Genomic studies have established the landscape of molecular changes in HCC, however, only ~25% of tumors harbor known targetable drivers. On the other hand, recent advances in high throughput sequencing technologies have uncovered a surprising number of alternatively spliced variants associated with tumorigenesis, implicating de-regulated splicing in the tumor phenotype. Hence, we have turned our attention to the alternatively or aberrantly spliced transcripts in the HCC ?spliceome? to identify new therapeutic targets. Insulin receptor has uniquely evolved to undergo alternative splicing to produce two isoforms: the full- length INSR-B and exon 11 skipped INSR-A isoform. Data from TCGA liver cancer cohorts as well as our own multiple in-house patient cohorts show that normal liver tissue primarily expresses the insulin receptor B isoform, whereas human HCC patient samples express more INSR-A. INSR-A, in addition to binding to insulin, has abnormally high affinity for IGF2 and accelerates the onset of tumor-cell hallmarks like proliferation and angiogenesis. Our data further show that this conversion of INSR-B to INSR-A takes place in the presence of stress conditions such as hypoxia. These observations are particularly relevant to HCC because 1) Hif1a has been shown to be significantly elevated and associated with worse progression in HCC and 2) IGF2 has been referred to as an epigenetic onco-driver of HCC. We therefore hypothesize that altering the splice pattern of INSR in liver cancer will abrogate the proliferative signaling downstream and impede the tumorigenic process. To achieve therapeutic intervention, we propose to use splice-switching oligonucleotide (SSO) technology to restore the normal INSR splicing pattern in liver cells. In this proposal, we aim to generate a clinically relevant mouse model of HCC that faithfully recapitulates the INSR splicing changes seen in the human condition. The current HCC mouse models do not express INSR alternatively spliced isoforms and thus do not predict responsiveness to therapies targeting the IGF pathway. There is therefore a critical need for new mouse models of HCC that will allow accurate testing of therapeutic modalities.
The focus of this proposal is to generate novel mouse models for hepatocellular carcinoma (HCC) that express the alternatively spliced variants of the insulin receptor gene (INSR). Because INSR splice variants modify the HCC disease severity and treatment response in patients, these mouse models will be used to answer numerous questions pertaining to the therapeutic possibilities of targeting the insulin receptor for the treatment of HCC.
