CD16 and Siglec expression refine the phenotypic heterogeneity of steady-state myeloid-derived suppressor cells

Polymorphonuclear myeloid-derived suppressor cells (PMN-MDSCs) mediate cancer immune suppression by promoting an immunosuppressive microenvironment that inhibits effective anti-tumor immunity. However, they are still a poorly understood, heterogeneous mix of neutrophil subsets. This study aims to determine the Siglec expression profile on several neutrophil subsets and assess their immunosuppressive ability.

We identified CD16^high and CD16^low neutrophil subsets from the low-density fractions of human peripheral blood and compared them to high-density neutrophils. We profiled the expression of the entire family of Siglecs on these three key neutrophil populations under steady-state conditions in healthy subjects as well as cancer patients. Moreover, the ability of these populations, isolated from healthy subjects, to suppress T cell proliferation was assessed.

Two distinct subpopulations were investigated within the low-density fraction of human peripheral blood (CD15⁺CD66b⁺CD16^low and CD15⁺CD66b⁺CD16^high) and compared to high-density neutrophils (CD15⁺CD66b⁺CD16^high). We found that in addition to CD33 (Siglec-3), Siglec-5/-14, -7, and -9, are differentially expressed on the CD16^low and CD16^high low-density subsets in both healthy, steady-state subjects, and cancer patients. Upregulated expression of CD33 on the CD16^low cells led to the initial speculation that they are MDSCs. As the differential expression of Siglec-9 between these two populations was striking, we used CD16 and Siglec-9 double staining to quantify these populations, which demonstrated that the CD16^lowSiglec-9^low population is greatly upregulated in cancer patients. The CD16^high low-density and high-density neutrophils, but not the CD16^low low-density neutrophils from healthy subjects, inhibited T cell proliferation, indicating that the CD16^lowSiglec-9^low population are not MDSCs.

These results demonstrate that Siglecs are differentially expressed on neutrophil subsets, and along with CD16, may be used to help further define what is a PMN-MDSC. Consistent with current observations by others, PMN-MDSCs may encompass an array of neutrophil subtypes, including low-density neutrophils, and point to the need for more work to precisely define the genetic signatures of PMN-MDSCs.