Remodeling of Myeloid as Well as Lymphoid Compartments Following Checkpoint Blockade Immunotherapy
Gubin Matthew M1,Ward JP2, Esaulova E1, Malkova ON1, Runci D1, Wong P1, Noguchi T1, Arthur CD1, Meng W1, Schreiber RD1 and Artyomov MN1 1Department of Pathology & Immunology, 2Divison of Oncology, Department of Medicine Washington University School of Medicine, St. Louis, Missouri, USA
Although immune checkpoint blockade (ICB) therapy can induce durable clinical responses in a subset of cancer patients, the molecular and cellular changes associated with successful ICB therapy remain incompletely defined. We used two complementary forms of high dimensional profiling to define and compare differences in intratumoral CD45+ cells isolated from either progressively growing tumors in mice treated with control monoclonal antibody (mAb) or tumors undergoing complete clinical responses in mice treated with anti-PD-1 and/or anti-CTLA-4. Unbiased assessment of the transcriptional profile of tumor infiltrating immune cells by single cell RNA sequencing (scRNAseq) identified common and distinct alterations induced by the different ICB treatments in both lymphoid and myeloid cell populations. Specifically, we identified multiple subpopulations of effector CD4+ and CD8+ T cells, Tregs and NK cells and, strikingly, five distinct macrophage subpopulations. The latter were distinguishable by the combinatorial presence or absence of mrc1 (CD206), CX3CR1, CD1d1 and/or Nos2 (iNOS). The macrophage subpopulations represented a wide spectrum of activation states and changed dynamically during ICB therapy. These findings were confirmed and extended at the protein level using Time of Flight Mass Cytometry (CyTOF), which confirmed significant alterations in the make-up of the macrophage clusters infiltrating tumors upon ICB therapy, with a loss of cells expressing CX3CR1 and an accumulation of iNOS+ cells. Distinct subpopulations of CD103+ CD1d+ CD83+ and plasmacytoid dendritic cells are also apparent in the tumor environment. Unsupervised clustering analysis of lymphoid populations by CyTOF identified three separate populations of Foxp3+ Tregs, which were differentially affected by either anti-PD-1 or -CTLA-4 therapies. Metal-labeled peptide-MHC tetramers captured alterations in populations of neoantigen-specific CD8+ T cells upon ICB therapy, with an accumulation of antigen-specific cells expressing KLRG1. This study thus reveals that both myeloid and lymphoid compartments in a tumor undergo massive transcriptional and functional remodeling following successful ICB therapy.
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