Targeting AEBP1 to mitigate pro-tumor activity of cancer-associated fibroblasts and increase therapeutic efficacy to anti-PD-1

Cancer-associated fibroblasts (CAFs) are one of the components in tumor microenvironment (TME) that can facilitate tumor progression, invasion, and metastasis directly through tumor-CAF interactions, or indirectly by crosstalking with tumor infiltrating immune cells and inducing immunosuppressive TME. Indeed, high stromal signature has been associated with reduced therapeutic efficacy and resistance to Immune Checkpoint Blockade. AEBP1 is highly expressed in myofibroblast cells whose expression is further upregulated in activated fibroblasts and the extracellular matrix (ECM)-producing CAFs. AEBP1 has two alternatively spliced isoforms that the extracellular isoform binds to collagen and promotes collagen remodeling, and the intracellular isoform modulates transcription and signaling. We observed expression of both isoforms of AEBP1 in primary human CAFs. Our data showed that combined knock out (KO) of both isoforms of AEBP1 by gene editing technology decreased CAF proliferation, collagen gel contractility and CAF-mediated tumor cell proliferation in vitro. Pan AEBP1 KO mouse fibroblasts also showed reduced activity in vitro and in vivo in the co-implantation mouse model. We found that knocking out both isoforms of AEBP1 downregulated the collagen biosynthesis and ECM organization related pathways in both mouse fibroblasts and primary human CAFs through RNA sequencing studies. In addition, loss of AEBP1 in fibroblasts impacted tumor cell phenotypes resulting in significant decrease of tumor cells with,with epithelial-mesenchymal transition signature in the co-implantation mouse model. Furthermore, AEBP1 KO in CAFs enhanced the anti-PD-1 induced effector T cell activity and the anti-PD-1 efficacy in this co-implantation model. Altogether, our results demonstrate that AEBP1 is important in regulating CAF function in the TME. Inhibiting AEBP1, therefore, may be a novel strategy in targeting CAF tumor-supporting activity and overcoming cancer resistance to anti-PD-1 immunotherapy. RNAseq profiling of wild-type NIH3T3 cell line and three clonal derivatives with AEBP1 knocked out.