Transcription Factor–Mediated Reprogramming of Cancer-Associated Fibroblasts Reveals Targetable Vulnerabilities in Solid Tumors

Cancer-associated fibroblasts (CAFs) contribute to immune exclusion and therapy resistance in solid tumors, limiting the efficacy of chimeric antigen receptor (CAR) T cell and immune cell therapy. To overcome this, we developed a transcription factor (TF)-based strategy to reprogram prostate-derived CAFs (pCAFs) into normal fibroblast-like cells (NFs). We prioritized TFs enriched in quiescent stellate cells—Vitamin D receptor (VDR), Peroxisome Proliferator-Activated Receptor gamma (PPARγ), and p53—and selected VDR for proof-of-concept studies. Lentiviral VDR expression in pCAFs produced VDR-reprogrammed NFs (VDR-rpNFs) with reduced CAF markers, increased ATP, and suppressed TGF-β and IL6, indicating phenotypic and metabolic reversion. In both in vitro 3D co-cultures and in vivo , VDR-rpNFs disrupted tumor architecture, enhanced CAR T cell infiltration, and reduced necrosis. PPARγ- and p53-rpNFs showed similar reprogramming effects. These results suggest TF-guided fibroblast reprogramming as a viable strategy to remodel the tumor microenvironment and improve CAR T cell efficacy in solid tumors.