Abstract 4044: The immune phenotype in serial biopsies from metastatic TNBC undergoing chemo-immunotherapy
Authors: Deng Jiehui , Shah Suhagi , Thennavan Aatish , Krogsgaard Michelle , Perou Charles M. , Wong Kwok-Kin , Adams Sylvia 
Published: August 2019

Abstract
Triple negative breast cancer (TNBC) is an aggressive subtype of breast cancer with poor prognosis when metastatic. Immunotherapy, especially in combination with taxane chemotherapy has recently been shown to improve outcomes with PD-L1 as predictive biomarker for survival. To understand the mechanisms of response/resistance to chemo-immunotherapy for TNBC, we obtained serial core biopsies of TNBC patients (n=30) undergoing nab-paclitaxel and pembrolizumab treatment in a prospective trial at three time points, including baseline (before treatment), after 1 cycle (nab-paclitaxel treatment without immunotherapy), and after 2 cycles (nab-paclitaxel plus pembrolizumab treatment). fluorescence-activated cell sorter (FACS). We performed 10x genomics single-cell RNA sequencing (scRNA-seq) of immune cells isolated from these fresh, serial tumor specimens from two patients, a patient with subsequent RECIST response and a patient with rapid disease progression. Results were further validated by FACS and immunohistochemistry analyses. ScRNA-seq showed significant baseline heterogeneity as well as perturbations of the immune cell infiltrates with chemotherapy and chemo-immunotherapy. In the responder patient there was a significant expansion of T cells, including both CD4+ and CD8+ T cells. Before the pembrolizumab treatment, there are 1.84% of CD4+ T cells and 6.45% of CD8+ cells among total tumor infiltrating leukocytes (TILs). After the treatment, the percentage of CD4+ and CD8+ cells increased to 33.32% and 23.42% among TILs, respectively, as determined by FACS analysis. Cytotoxic T lymphocytes (CTLs) showed enrichments for transcripts of tissue-resident memory T cells (Trm). In contrast, tumors from a patient with rapid disease progression showed persistence of a myeloid compartment that suppress anti-tumor immunity. Although T cells were initially present, they mainly consisted of a naïve T cell phenotype and did not expand even after treatment. No Trm cells were detected within TILs by scRNAseq from this patient after pembrolizumab treatment. When comparing baseline samples from these two patients, we found there is a distinct tumor cell population in the non-responder with differentially expressed genes related to oxygen transport and proliferation. The association between this population and the response to chemo-immunotherapy remains to be explored. Our study provides proof of concept that single cell analyses of serial biopsies are a feasible tool to understand or even predict responses to chemo-immunotherapy for TNBC patients.
Citation Format: Jiehui Deng, Suhagi Shah, Aatish Thennavan, Michelle Krogsgaard, Charles M. Perou, Kwok-Kin Wong, Sylvia Adams. The immune phenotype in serial biopsies from metastatic TNBC undergoing chemo-immunotherapy [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 4044.