Potential of FAK/PYK2 inhibition in overcoming immune checkpoint inhibitor resistance

Summary: Immune checkpoint inhibitors (ICIs) have transformed cancer therapy, yet many patients ultimately develop acquired resistance. IFNγ-mediated induction of PD-L1 and STAT1 has been implicated in this resistance, but the underlying signaling pathways remain incompletely understood. Phosphoproteomic analyses of IFNγ-stimulated melanoma cells provided significant enrichment of phosphorylation events associated with the cytoskeleton and cell adhesion—processes in which FAK and PYK2 play central regulatory roles. To determine whether these kinases contribute to IFNγ-driven responses, we stimulated cells with IFNγ in the presence of the FAK/PYK2 inhibitor VS6063. Under these conditions, IFNγ-induced PD-L1 and STAT1 expression was suppressed by more than 90%, pinpointing FAK as a central regulator of IFNγ-driven signaling. In 470 TCGA melanoma samples, gene set enrichment analysis comparing samples with high and low FAK expression revealed significant enrichment of ICI resistance-related terms in the high FAK expression group. Furthermore, a re-analysis of existing RNA sequencing data from 56 human melanoma specimens showed that FAK levels were significantly elevated in ICI non-responders compared to responders (p<0.05). In co-culture assays with human CD8⁺ T cells, FAK/PYK2 inhibition significantly enhanced tumor cell killing (n=4, p<0.05). Network analysis identified paxillin (PXN) as a key IFNγ-responsive signaling hub, suggesting that PXN-related pathways further support immune suppression. These findings demonstrate that dual blockade of FAK and PYK2 effectively suppresses IFNγ-driven PD-L1 and STAT1 expression, offering a promising strategy to overcome ICI resistance in melanoma. Importantly, similar results were observed in breast cancer and glioblastoma models, suggesting that this approach could have broader applications in cancers with immune checkpoint resistance. Yuto Mizuno¹, Masanari Umemura², Momoko Nagai³, Jo Nishino³, Mamoru Kato³, Kazuo Horikawa², Tomoko Akiyama², Yayoi Kimura², Yukie Yamaguchi², Yoshihiro Ishikawa¹ 1. Yokohama Shiritsu Daigaku, Yokohama, Kanagawa Prefecture, Japan. 2. Yokohama Shiritsu Daigaku Igakubu Daigakuin Igaku Kenkyuka, Yokohama, Kanagawa Prefecture, Japan. 3. Department of Bioinformatics, Research Institute National Cancer Center, Tokyo, Japan. Pigmentation, Melanoma, and Melanoma Immune Surveillance