Single cell RNA sequencing reveals distinct gene expression signatures of myeloidderived suppressor cells in breast cancer

Abstract Myeloid-derived suppressor cells (MDSCs) are a heterogeneous population of immature myeloid cells with potent immune suppressive activity. MDSCs regulate anti-tumor immunity by suppressing T cell proliferation. MDSCs can be further classified into granulocytic MDSCs (G-MDSCs) and monocytic MDSC (M-MDSCs). However, there is a lack of MDSC-specific markers since MDSCs, normal neutrophil granulocytes, and monocytes are defined by the same flow cytometry markers (CD11b+Gr1+). We used a breast cancer mouse model (MMTV-PyMT) to define cellular and molecular properties of MDSCs in single cell resolution. To test the capacity of MDSCs to inhibit immune responses, CD11b+Gr1+ cells from PyMT and wild type (WT) mice were sorted by fluorescence-activated cell sorting (FACS) from bone marrow, lung, and spleen, and then subjected to a T cell activation assay in co-culture with T cells. We found that predominantly spleen-derived CD11b+Gr1+ cells from PyMT significantly suppressed CD4 and CD8 T cell proliferation, while CD11b+Gr1+cells from bone marrow and lung showed no effect on T cell proliferation. Thus, we focused on spleen-derived CD11b+Gr1+ cells and utilized single cell RNA sequencing (scRNAseq) to compare breast cancer derived MDSCs to the respective cell populations harvested from normal, and non-tumor bearing hosts. Our computational analysis of 14,646 single cell transcriptomes revealed a separate G-MDSCs cluster from normal neutrophils and a M-MDSCs cluster from normal monocytes and a distinct gene expression signature for the entire MDSCs population. Our studies provide crucial insights into the biology of MDSCs, which may ultimately form the basis for novel marker and therapeutic avenues to improve cancer immunotherapy.