Targeting Src SH3 domain-mediated glycolysis of hepatic stellate cells suppresses transcriptome, myofibroblastic activation, and colorectal liver metastasis

Background and Aims: TGF beta 1 induces hepatic stellate cell (HSC) activation into metastasis-promoting cancer-associated fibroblasts (CAFs), but how the process is fueled remains incompletely understood. We studied metabolic reprograming induced by TGF beta 1 in HSCs. Approaches and Results: Activation of cultured primary human HSCs was assessed by expression of myofibroblast markers. Glucose transporter 1 (Glut1) of murine HSC was disrupted by Cre/LoxP recombination. Plasma membrane (PM) Glut1 and glycolysis were studied by biotinylation assay and the Angilent Seahorse XFe96 Analyzer. Subcutaneous HSC/tumor co-implantation and portal vein injection of MC38 colorectal cancer cells into HSC-specific Glut1 knockout mice were performed to determine in vivo relevance. Transcriptome was obtained by RNA sequencing of HSCs and spatialomics with MC38 liver metastases. TGF beta 1-induced CAF activation of HSCs was accompanied by elevation of PM Glut1, glucose uptake, and glycolysis. Targeting Glut1 or Src by shRNA, pharmacologic inhibition, or a Src SH3 domain deletion mutant abrogated TGF beta 1-stimulated PM accumulation of Glut1, glycolysis, and CAF activation. Mechanistically, binding of Src SH3 domain to SH3BP5 led to a Src/SH3BP5/Rab11/Glut1 complex that activated Rab11-dependent Glut1 PM transport under TGF beta 1 stimulation. Deleting the Src SH3 domain or targeting Glut1 of HSCs by shRNA or Cre/LoxP recombination suppressed CAF activation in mice and MC38 colorectal liver metastasis. Multi-omics revealed that Glut1 deficiency in HSCs/CAFs suppressed HSC expression of tumor-promoting factors and altered MC38 transcriptome, contributing to reduced MC38 liver metastases. Conclusion: The Src SH3 domain-facilitated metabolic reprogramming induced by TGF beta 1 represents a target to inhibit CAF activation and the pro-metastatic liver microenvironment.