Abstract PD2-02: Longitudinal local and peripheral immunologic changes associated with checkpoint inhibition response in murine models of breast cancer
Authors: Hanna Ann , Sun Xiaopeng , Sheng Quanhu , Sanders Melinda , Balko Justin 
Published: March 2023

Abstract
Immune checkpoint inhibitors (ICI) have significantly enhanced patient survival in some cancer types but yield limited success in breast cancer. ICIs activate anti-tumor immunity by overriding the inhibition of tumor infiltrating lymphocytes (TILs). Clinical trials in triple negative breast cancer (TNBC) patients, who are more likely to harbor TILs within tumor stroma, have demonstrated increased progression-free survival (IMpassion130) and pathologic complete response (KEYNOTE-522) to ICI. Consequently, combinations of ICI and chemotherapy have been FDA-approved for metastatic TNBC. However, the therapeutic benefit of ICIs is highly heterogeneous among breast cancer patients; as such, we sought to model ICI response in vivo to evaluate therapeutic resistance and response heterogeneity, as well as ascertain predictive biomarkers for favorable outcomes to ICI in breast cancer. We used an immunocompetent EMT6 orthotopic mammary tumor model to investigate the efficacy of ICI (anti-PD-L1). Analysis of the primary tumor immune landscape was performed by flow cytometry and single-cell RNA sequencing. Matched longitudinal samples of the tumor microenvironment (collected by fine-needle aspiration) and peripheral blood (PBMC) from mice were profiled by bulk RNA and T-cell receptor (TCR) sequencing to identify systemic genomic alterations and T-cell expansion, respectively. Single-agent ICI robustly suppressed primary tumor growth (p =0.0046) and extended survival (p&lt; 0.0001) beyond the control group in the EMT6 model. The addition of chemotherapy (paclitaxel and/or doxorubicin) demonstrated moderate therapeutic efficacy but failed to enhance ICI benefit. Phenotypic profiling of the tumor microenvironment (TME) revealed increased T cells, dendritic cells, and NK cells in anti-PD-L1 only and chemotherapy combination groups. Despite using a genetically identical tumor model and murine host, we found that PD-L1 blockade induced heterogeneous responses, similar to clinical outcomes in breast cancer patients, ranging from complete response to intrinsic resistance. Analysis of the primary tumor microenvironment showed upregulated signatures of cytotoxic T cell response and activation, specifically inflammatory interferon signaling (both prior to and post ICI administration) that corresponded to favorable response to anti-PD-L1 in individual mice. Longitudinal analysis of the peripheral blood identified modest changes among mice at baseline that progressively deviated by response type (non responders-vs-responder mice). Moreover, mice harbored enriched myeloid signatures and clonal T cell expansion during therapy corresponding to ICI resistance and response, respectively. Further investigations of matched peripheral blood and the primary tumor microenvironment signatures may identify systemic biomarkers and tumor antigen-specific T cell clones to accurately predict ICI response in patients and uncover mechanisms for sensitizing tumors refractory to ICI. In conclusion, we identify a heterogeneously ICI-responsive in vivo model that emulates TNBC patient response to combinatorial ICI approaches. We describe host-specific signatures, specifically myeloid cell responses, that correlate with differential responses to immunotherapy, which may serve as a basis for tracking immunotherapy response in peripheral blood from breast cancer patients.
Citation Format: Ann Hanna, Xiaopeng Sun, Quanhu Sheng, Melinda Sanders, Justin Balko. Longitudinal local and peripheral immunologic changes associated with checkpoint inhibition response in murine models of breast cancer [abstract]. In: Proceedings of the 2022 San Antonio Breast Cancer Symposium; 2022 Dec 6-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2023;83(5 Suppl):Abstract nr PD2-02.