BACKGROUND: Therapeutic resistance limits durable survival in advanced/metastatic renal cell carcinoma (RCC) treated with first-line tyrosine kinase inhibitor (TKI) plus immune checkpoint inhibitor (ICI). We sought to define key resistance drivers and actionable targets. METHODS: Integrated RNA sequencing of cabozantinib-resistant RCC cells, lipid metabolomics, and PD-L1 correlation analyses identified ENPP2 as a candidate driver. Its role in TKI resistance and survival signaling was validated by apoptosis, CCK-8, and colony formation assays in vitro and by nude-mouse xenograft models in vivo. ELISA, flow cytometry and tumor cell-T-cell co-culture assays were used to dissect ENPP2-dependent CD8+ T-cell dysfunction. The therapeutic benefit of pharmacologic ATX inhibition combined with standard TKI-ICI regimens was tested in RCC patient-derived xenograft models. RESULTS: The ATX-LPA axis conferred TKI resistance via constitutive AKT/mTOR activation and promoted immune evasion by upregulating PD-L1 through TBK1/IRF3 signaling, thereby impairing intratumoral CD8+ T-cell function. ENPP2 enhanced PD-L1 transcription by facilitating IRF3 nuclear translocation and its direct recruitment to the CD274 promoter. ATX inhibition improved the antitumor efficacy of TKI-ICI therapy in preclinical models. CONCLUSIONS: Targeting the ATX-LPA axis represents a promising strategy to overcome resistance to current TKI-ICI combinations.

Epidermal Growth Factor Receptor (EGFR) is often present on the cell surface of a wide variety human malignancies, including non-small-cell lung cancer (NSCLC), glioblastoma (GBM), pancreatic ductal carcinoma (PDC), and castration-resistant prostate cancer (CRPC). Efforts to optimize immunotherapies targeting EGFR are limited by murine intolerance of human EGFR. To overcome this obstacle, we developed C57BL/6 mice in which a truncated variant of human EGFR (hEGFRt), lacking the ligand binding domain and cytoplasmic domain, is expressed from the CAG regulatory elements comprised of the CMV enhancer, β-actin promoter, and β-globin poly-adenylation signals. Cetuximab, a high-affinity, clinically available anti-human EGFR antibody, retains affinity for hEGFRt. The hEGFRt cDNA in the CAG-hEGFRt transgene is flanked by loxP sites to enable its excision thereby reducing interaction of hEGFRt-directed immunotherapies with normal tissues. The CAG-hEGFRt(f/f) transgene is abundantly expressed in hematopoietic lymphoid and myeloid cells, with low-level expression evident also in non-hematopoietic liver, lung, kidney, and brain. Mx1-Cre-mediated transgene excision in adult mice reduces hEGFRt ~ 5-fold in blood mononuclear cells. CAG-hEGFRt(f/f) adult mice are tolerant of syngeneic NSCLC, GBM, prostate, and PDC lines expressing hEGFRt. These mice retain hEGFRt tolerance after transgene deletion. CAG-hEGFRt(f/f) mice provide a new and important tool for the development of immunotherapies targeting hEGFR.

Immune checkpoint inhibitors (ICIs) extend survival in many patients with cancer but are ineffective in patients without pre-existing immunity1–9. Although personalized mRNA cancer vaccines sensitize tumours to ICIs by directing immune attacks against preselected antigens, personalized vaccines are limited by complex and time-intensive manufacturing processes10–14. Here we show that mRNA vaccines targeting SARS-CoV-2 also sensitize tumours to ICIs. In preclinical models, SARS-CoV-2 mRNA vaccines led to a substantial increase in type I interferon, enabling innate immune cells to prime CD8+ T cells that target tumour-associated antigens. Concomitant ICI treatment is required for maximal efficacy in immunologically cold tumours, which respond by increasing PD-L1 expression. Similar correlates of vaccination response are found in humans, including increases in type I interferon, myeloid–lymphoid activation in healthy volunteers and PD-L1 expression on tumours. Moreover, receipt of SARS-CoV-2 mRNA vaccines within 100 days of initiating ICI is associated with significantly improved median and three-year overall survival in multiple large retrospective cohorts. This benefit is similar among patients with immunologically cold tumours. Together, these results demonstrate that clinically available mRNA vaccines targeting non-tumour-related antigens are potent immune modulators capable of sensitizing tumours to ICIs. mRNA vaccines targeting SARS-CoV-2 also sensitize tumours to immune checkpoint inhibitors.