Abstract 2458: Targeting the CXCR3 axis in HER2+ breast cancer to enhance anti-tumor immunity

Abstract Breast cancer (BC) is the most frequently diagnosed cancer in women worldwide with more than 2 million new cases in 2020. The efficacy of immunotherapies for BC is limited, with response rates of <10% for anti-PD-1/PDL1 monotherapy in pre-treated triple-negative breast cancer (TNBC) patients and even lower in subtypes that express human epidermal growth factor receptor 2 (HER2+) and/or hormone receptors. Hence, there is an urgent need to improve immune based strategies for BC. One possible approach to enhance response is the application of intratumoral (IT) immunotherapy utilizing IL-12; a potent pro-inflammatory cytokine that is critical in producing an anti-tumor response. However, systemic delivery of IL-12 has substantial toxicity and localized delivery of protein has insufficient residence time. To circumvent these issues, we have previously reported the efficacy of intratumorally injected plasmid IL-12 (pIL-12-EP) followed by electroporation in a TNBC mouse model and in clinical trials. Following pIL-12-EP, treated tumor expressed genes that were associated with increased CD8+ T cell infiltration, activation, trafficking, antigen presentation, and exhaustion. Single-cell RNA sequencing (scRNAseq) of tumor infiltrating immune cells from treated tumors revealed a striking enrichment of the CXCL9/10/11/CXCR3 family. This CXCR3 axis is upregulated in response to IFNγ production and has been studied extensively in melanoma and NSCLC patients as a potential predictive biomarker for responses to immune checkpoint inhibitors but has not been well characterized in BC. We hypothesized that this axis may be the critical component downstream of pIL-12-EP treatment that is responsible for anti-tumor responses and that targeting it will be effective in other molecular subtypes of BC beyond TNBC. We first evaluated the expression of CXCR3 by tumor infiltrating lymphocytes (TILs) in biopsies of human HER2+ BC. Using a single cell resolution spatial transcriptomic Xenium instrument we find not only expression of CXCR3 by TILs, but can also track the distance and gradients of the chemokine ligands CXCL9 and CXCL10 in this tumor microenvironment. Having confirmed the activity of this axis in human HER2+ BC, we used our well-established spontaneous HER2-driven mouse model to compare treatment of HER2+ tumors with pIL-12-EP or pCXCL9-EP intratumorally. We observed tumor growth inhibition in both treatment groups as compared to control treated tumors. Interestingly, the abscopal effect previously seen with pIL-12-EP was not evident for distant tumors in mice treated with pCXCL9-EP. These data suggest that IT CXCL9 therapies may benefit from combination with a systemic immune activator like a vaccine or immunogenic chemotherapy. Ongoing studies will continue to evaluate ways to better target this axis therapeutically and define clinically active combination therapies. Citation Format: Anchit Bhagat, Jesuchristopher Joseph, Joshua D. Ginzel, Christopher Rabiola, Amy C. Hobeika, Joshua C. Snyder, Herbert K. Lyerly, Erika J. Crosby. Targeting the CXCR3 axis in HER2+ breast cancer to enhance anti-tumor immunity [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 2458.