Single-cell RNA sequencing reveals tumor heterogeneity and potential mechanisms of response/resistance in breast cancer treated with neoadjuvant therapy

BackgroundNeoadjuvant therapy (NAT) with chemotherapy (CT) backbone remains the mainstay for most early-stage aggressive breast cancer (BC) subtypes such as triple-negative (TNBC), HER2-positive, and luminal B-like BC. However, patients without a pathological complete response (pCR) will have a high risk of recurrence. This study aims to provide insights into cellular and molecular heterogeneity and potential resistance mechanisms to NAT using single-cell RNA sequencing (scRNA-seq).MethodsWe obtained high-quality scRNA-seq data from prospectively collected samples of 37 patients: 29 treated with CT alone (including luminal B, HER2-positive, and TNBC) and 8 with TNBC treated with NAT chemo-immunotherapy. Cells were processed using the 10x Genomics Chromium Next GEM Single Cell 5’ platform and analyzed for gene expression and T-cell and B-cell repertoire diversity.ResultsA total of 286,346 cells were annotated from baseline biopsies and the tumor bed and normal adjacent tissues at surgery, with significant differences in cell type composition between the tumor and normal adjacent tissues. TNBC tumor cells expressed markers suggestive of a basal/progenitor origin, while luminal and HER2-positive tumor cells displayed a luminal hormone-responsive origin. Sub-clonal analysis revealed multiple tumor clones within the same patient exhibiting distinct transcriptional programs, which may contribute to treatment resistance. Of interest, we identified clonotype variations in T-cell receptor repertoires and found CD8+ CXCL13+ enrichment with clonotype expansion in baseline samples of responders to CT. An analysis of the cell cycle phases showed a significant association of the cell cycle G1 phase in tumor and T cells with non-pCR (p<0.05) and of G2/M phases in CD8+ T-cells (p<0.01), NK and Treg cells (p<0.05) with pCR. We also described a population of ITM2C+ T-cells with opposite trends of enrichment in pCR versus non-pCR among patients treated with CT alone versus CT-immunotherapy.ConclusionsOur findings underscore the complexity of tumor heterogeneity and provide insights into the cellular dynamics, with the potential to unveil predictive biomarkers of response to NAT in BC.Editorial acknowledgementThis research project was supported by ESMO. Any views, opinions, findings, conclusions, or recommendations expressed in this material are those solely of the author(s) and do not necessarily reflect those of ESMO.Legal entity responsible for the studyThe authors.FundingInstitut Jules Bordet.DisclosureAll authors have declared no conflicts of interest.