T-cell and Soluble Co-inhibitory Receptor Expression in Patients With Visceral Leishmaniasis Are Markers of Treatment Response and Clinical Outcome

Background: Co-inhibitory receptors (immune checkpoints) regulate activated immune cells. Their expression on T cells can limit host defense. We hypothesized that chronic Leishmania donovani infection in patients with visceral leishmaniasis (VL) leads to expression of co-inhibitory receptors that could be markers of treatment response and clinical outcome. Method: A prospective cohort of 21 subjects with VL (7 with HIV coinfection) and 10 controls was established to measure T-cell expression of co-inhibitory receptors (PD-1, Tim-3, LAG-3, CTLA-4, and TIGIT) by flow cytometry in discarded remnants of diagnostic splenic or bone marrow aspirates and peripheral blood collected before and after treatment. Plasma levels of soluble co-inhibitory proteins (sPD-1, sTim-3, sLAG-3, and sCTLA-4) and selected cytokines were determined by immunoassay. Results: Expression of co-inhibitory receptors in peripheral blood T cells generally reflected findings in spleen and bone marrow aspirates. PD-1 and Tim-3 were upregulated in CD4+ T cells in HIV-negative and HIV-positive subjects with VL compared to controls. CD8+ T cells from HIV-negative subjects with VL displayed a similar pattern. Plasma levels of sPD-1 and sTim-3 were also greater in VL patients than controls. CD8+ and CD4+ T cells coexpressing PD-1 and Tim-3 showed considerable decline with treatment. Mortality in HIV-negative VL patients was associated with increased CD8+ T cells coexpressing Tim-3 and PD-1, triple-positive CD4+ and CD8+ T cells (PD-1⁺Tim-3⁺LAG-3⁺), and elevated sLAG3. Conclusions: Tim-3 and PD-1 expression on CD4+ and CD8+ T cells, and increased plasma sLAG-3, were markers of treatment response and clinical outcome in patients with VL.