Prevention of insulin receptor isoform A formation inhibits MASH-driven hepatocellular carcinoma

Introduction: Metabolic dysfunction-associated steatohepatitis (MASH) has emerged as a leading driver of hepatocellular carcinoma (HCC). Our previous work identified that the loss of the RNA splicing factor SRSF3 is an early event in chronic liver disease. This loss leads to the aberrant skipping of exon 11 in the insulin receptor ( INSR ) gene, shifting the isoform expression from the normal hepatic IR-B to the fetal IR-A isoform. Unlike IR-B, IR-A binds insulin-like growth factor 2 (IGF2) with high affinity, potentially establishing a mitogenic autocrine loop. We hypothesized that this specific splicing switch is a critical driver of hepatocarcinogenesis and that preventing it could inhibit tumor formation. Methods: To test this hypothesis, we generated a novel "Exon 11+" mouse model in which Insr exon 11 inclusion is genetically enforced, thereby preventing the production of the IR-A isoform. We subjected Exon 11+ mice and wild-type (WT) littermates to a chemical-dietary carcinogenesis protocol. Mice received a single injection of diethylnitrosamine (DEN) at 2 weeks of age, followed by chronic administration of thioacetamide (TAA) and a Western diet (MASH conditions) for 24 weeks to induce HCC. Tumor burden was assessed by surface counting and measuring. Liver tissues were analyzed via histology (H&E, Sirius Red, Reticulin), immunohistochemistry (Ki67, CD45), and qPCR. Results: WT mice developed extensive liver tumors accompanied by severe fibrosis and inflammation. In striking contrast, Exon 11+ mice were significantly protected against tumorigenesis, exhibiting a profound reduction in both the number (decrease >80%, p<0.001) and size (p<0.05) of surface tumors. Histological analysis confirmed that Exon 11+ mice had significantly reduced liver fibrosis (Sirius Red, p<0.001) and immune cell infiltration (CD45, p<0.01) compared to WT controls. Notably, Reticulin staining revealed that Exon 11+ livers maintained normal hepatic plate architecture (1-2 cell thickness), whereas WT livers displayed disorganization and thickening characteristic of HCC. Furthermore, enforcing IR-B expression significantly suppressed hepatocyte proliferation (Ki67, p<0.0001) and downregulated the expression of key proinflammatory (Tnf, Il6, Ccl2) and profibrogenic (Col1a1, Col3a1) genes (all p<0.05). Conclusions: Our data provide the first in vivo genetic evidence that the alternative splicing switch of the insulin receptor from IR-B to IR-A is a necessary event for MASH-driven HCC progression. Blocking this specific splicing error is sufficient to suppress inflammation, fibrosis, and tumor development in a harsh carcinogenic environment. These findings suggest that targeting the IGF2/IR-A signaling axis or correcting INSR splicing represents a potent therapeutic strategy for preventing MASH-associated liver cancer.