Abstract
BackgroundAlthough the anti-PD-1+LAG-3 and the anti-PD-1+CTLA-4 combinations are effective in advanced melanoma, it remains unclear whether their mechanisms of action overlap.MethodsWe used single cell (sc) RNA-seq, flow cytometry and IHC analysis of responding SM1, D4M-UV2 and B16 melanoma flank tumors and SM1 brain metastases to explore the mechanism of action of the anti-PD-1+LAG-3 and the anti-PD-1+CTLA-4 combination. CD4+ and CD8+ T cell depletion, tetramer binding assays and ELISPOT assays were used to demonstrate the unique role of CD4+T cell help in the antitumor effects of the anti-PD-1+LAG-3 combination.ResultsThe anti-PD-1+CTLA-4 combination was associated with the infiltration of FOXP3+regulatory CD4+ cells (Tregs), fewer activated CD4+T cells and the accumulation of a subset of IFNγ secreting cytotoxic CD8+T cells, whereas the anti-PD-1+LAG-3 combination led to the accumulation of CD4+T helper cells that expressed CXCR4, TNFSF8, IL21R and a subset of CD8+T cells with reduced expression of cytotoxic markers. T cell depletion studies showed a requirement for CD4+T cells for the anti-PD-1+LAG-3 combination, but not the PD-1-CTLA-4 combination at both flank and brain tumor sites. In anti-PD-1+LAG-3 treated tumors, CD4+T cell depletion was associated with fewer activated (CD69+) CD8+T cells and impaired IFNγ release but, conversely, increased numbers of activated CD8+T cells and IFNγ release in anti-PD-1+CTLA-4 treated tumors.ConclusionsTogether these studies suggest that these two clinically relevant immune checkpoint inhibitor (ICI) combinations have differential effects on CD4+T cell polarization, which in turn, impacted cytotoxic CD8+T cell function. Further insights into the mechanisms of action/resistance of these clinically-relevant ICI combinations will allow therapy to be further personalized.
Funder
Melanoma Research Foundation
National Institutes of Health
National Cancer Institute
Florida Department of Health
The Department of Defense
Subject
Cancer Research,Pharmacology,Oncology,Molecular Medicine,Immunology,Immunology and Allergy
Cited by
2 articles.
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