Hierarchy of immunosuppressive strength among myeloid-derived suppressor cell subsets is determined by GM-CSF

CD11b⁺/Gr-1⁺ myeloid-derived suppressor cells (MDSC) contribute to tumor immune evasion by restraining the activity of CD8 ⁺ T-cells. Two major MDSC subsets were recently shown to play an equal role in MDSC-induced immune dysfunctions: monocytic- and granulocytic-like. We isolated three fractions of MDSC, i.e. CD11b ⁺/Gr-1^high, CD11b⁺/ Gr-1^int, and CD11b⁺/Gr-1^low populations that were characterized morphologically, phenotypically and functionally in different tumor models. In vitro assays showed that CD11b⁺/Gr-1^int cell subset, mainly comprising monocytes and myeloid precursors, was always capable to suppress CD8⁺ T-cell activation, while CD11b⁺/Gr-1 ^high cells, mostly granulocytes, exerted appreciable suppression only in some tumor models and when present in high numbers. The CD11b ⁺/Gr-1^int but not CD11b⁺/Gr-1^high cells were also immunosuppressive in vivo following adoptive transfer. CD11b⁺/Gr-1^low cells retained the immunosuppressive potential in most tumor models. Gene silencing experiments indicated that GM-CSF was necessary to induce preferential expansion of both CD11b ⁺/Gr-1^int and CD11b⁺/Gr-1^low subsets in the spleen of tumor-bearing mice and mediate tumor-induced tolerance whereas G-CSF, which preferentially expanded CD11b⁺/Gr-1^high cells, did not create such immunosuppressive environment. GM-CSF also acted on granulocyte-macrophage progenitors in the bone marrow inducing local expansion of CD11b⁺/Gr-1^low cells. These data unveil a hierarchy of immunoregulatory activity among MDSC subsets that is controlled by tumor-released GM-CSF.