Single-cell transcriptomic analysis identified resistant MDSCs and a stress-tolerant gene co-expression network as common MDSC features across multiple disease settings

Tianmeng Chen ^1* Julia Hughes ¹ Alyssa Gregory ¹ Julia Conroy ¹ Patricia Loughran ¹ Jinming Song ² Wei Chen ¹ Timothy Robert Billiar ^1*

• ¹ University of Pittsburgh, Pittsburgh, United States
• ² Moffitt Cancer Center, Tampa, Florida, United States

Background: Myeloid-derived suppressor cells (MDSCs) are a heterogeneous population of immunosuppressive myeloid cells. The identification of a molecular signature common to MDSC regardless of tissue source would aid in the classification of cells as MDSCs. Methods: Single-cell RNA sequencing (scRNA-seq) was performed on GM-CSF+ IL-6 induced human MDSCs to characterize the extent of heterogeneity within monocytic MDSCs (M-MDSCs). Cytokinetreated PBMCs were also cultured in the absence of serum to include an additional element of cell stress. Independent published bulk and single-cell transcriptomic datasets were used for validation. Findings: A cluster of cells with preserved MDSC features was induced by the combination of inflammatory signals and cell stress in the form of serum starvation (resistant MDSCs, rMDSCs). A gene co-expression module (the yellow module) was identified specific to rMDSCs. The genes upregulated in MDSCs can be further classified into stress-tolerant vs. -sensitive features. This yellow module mostly contained stress-tolerant genes and showed excellant separation for distinguishing M-MDSCs from control cells across a range of in vitro and in vivo conditions (ROC AUC=0.954), a feature not found in the stress-sensitive genes. Importantly, rMDSCs were identified in scRNA-seq datasets of immune cells from multiple human cancer types. Tumor C1Q macrophages, which have been associated with immunosuppression, highly expressed the yellow module gene signature. Interpretation: These results demonstrate the importance of the combined roles of inflammation and cellular stress in shaping the features of M-MDSCs, and highlights cellular resilience represented by rMDSCs and the role of stress-tolerant features in defining common MDSC features.