Abstract
A subset of cancer-associated fibroblasts (FAP+/CAF-S1) mediates immunosuppression in breast cancers, but its heterogeneity and its impact on immunotherapy response remain unknown. Here, we identify 8 CAF-S1 clusters by analyzing more than 19,000 single CAF-S1 fibroblasts from breast cancer. We validate the five most abundant clusters by flow cytometry and in silico analyses in other cancer types, highlighting their relevance. Myofibroblasts from clusters 0 and 3, characterized by extracellular matrix proteins and TGFβ signaling, respectively, are indicative of primary resistance to immunotherapies. Cluster 0/ecm-myCAF upregulates PD-1 and CTLA4 protein levels in regulatory T lymphocytes (Tregs), which, in turn, increases CAF-S1 cluster 3/TGFβ-myCAF cellular content. Thus, our study highlights a positive feedback loop between specific CAF-S1 clusters and Tregs and uncovers their role in immunotherapy resistance. SIGNIFICANCE: Our work provides a significant advance in characterizing and understanding FAP+ CAF in cancer. We reached a high resolution at single-cell level, which enabled us to identify specific clusters associated with immunosuppression and immunotherapy resistance. Identification of cluster-specific signatures paves the way for therapeutic options in combination with immunotherapies.This article is highlighted in the In This Issue feature, p. 1241.
Highlights
Cancer-Associated Fibroblasts (CAF) represent one of the most abundant components in adenocarcinomas and play key pro-tumorigenic functions [1,2,3,4,5,6]
We considered EPCAM- CD45- CD31- CD235a- as the fraction of cells enriched in fibroblasts and performed Fibroblast Activation Protein (FAP) and CD29 staining (Fig. S1A), which enabled us to distinguish CAF-S1 (FAPHigh CD29Med-High) from the other CAF subpopulations (CAF-S2: FAPNeg CD29Low; CAF-S3: FAPNeg CD29Med; CAF-S4: FAPNeg CD29High) (Fig. S1A), as previously established in [8,10]
By performing scRNA-seq on more than 19 000 CAF-S1 fibroblasts from breast cancers (BC) patients, we identified 8 cellular clusters within the CAF-S1 immunosuppressive subset in human BC
Summary
Cancer-Associated Fibroblasts (CAF) represent one of the most abundant components in adenocarcinomas and play key pro-tumorigenic functions [1,2,3,4,5,6]. CAF-S1 fibroblasts stimulate immunosuppression by increasing attraction, survival and overall content of CD4+ CD25+ FOXP3+ regulatory T cells (Tregs) in tumor micro-environment [8,10]. In line with these observations, FAPHigh CAF have been suspected to contribute to primary resistance to immunotherapies [7,9,25]. To our knowledge, the direct role of FAPHigh CAF in immunotherapy resistance has not yet been addressed in human cancer This observation, coupled with the recent finding that FAPHigh CAF-S1 fibroblasts exert their immunosuppressive function through a multi-step mechanism [8,10], prompted us to study their heterogeneity and specific roles in primary resistance to immunotherapy in cancer patients
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