Human T-cell lymphotropic virus type 1 (HTLV-1)-associated myelopathy/tropical spastic paraparesis (HAM/TSP) develops in 1–5% of HTLV-1-infected individuals. Previous studies by us and others have shown that the expression of negative immune checkpoint receptors (NCRs) is significantly increased on CD8 T cells in various chronic viral infections and are associated with poor anti-viral immunity. We have previously identified the differential expression of NCRs on CD8 T cells in blood from patients with HAM/TSP and in central nervous system (CNS) tissues of HTLV-1 infected humanized mice and defined the association with neurological complications. In this study, we determined the co-expression patterns of several key NCRs (PD-1, TIGIT, TIM-3, and LAG-3) and their cognate ligands in HTLV-1 infection and assessed how combination strategies targeting these pathways would impact HTLV-1-specific CD8 T-cell effector functions as an approach to reduce CNS disease outcomes. We found that global CD8 T cells from HAM/TSP patients co-express multiple NCRs at significantly higher frequencies than asymptomatic carriers (AC). Moreover, NCR ligands (PVR and PD-LI) on both plasmacytoid and myeloid dendritic cells were also expressed at higher frequencies in HAM/TSP compared to AC. In both AC and HAM/TSP subjects, combination dual PD-L1/TIGIT or triple PD-L1/TIGIT/TIM-3 blockade with monoclonal antibodies resulted in increases in intracellular cytokine expression in CD8 T cells after virus stimulation, particularly CD107a, a marker of degranulation, and TNF-α, a key cytokine that can directly inhibit viral replication. Interestingly, almost all blockade combinations resulted in reduced IL-2+ HTLV-1-specific CD8 T cell frequencies in HAM/TSP subjects, but not in AC. These results define a novel combinatorial NCR immunotherapeutic blockade strategy to reduce HAM/TSP disease burden.

Targeting inhibitory immune checkpoint receptor pathways has shown remarkable success in improving anticancer T cell responses for the elimination of tumors. Such immunotherapeutic strategies are being pursued for HIV remission. Metformin has shown favorable clinical outcomes in enhancing the efficacy of programmed cell death-1 (PD-1) blockade and restoring antitumor T cell immunity. Furthermore, monocytes are known to be a strong predictor of progression-free survival in response to anti-PD-1 immunotherapy. In a single-arm clinical trial, we evaluated the immunological effects over an 8-week course of metformin therapy in seven euglycemic, virally suppressed HIV-infected participants on combination antiretroviral therapy (cART). We assessed changes in peripheral HIV-Gag-specific T cell responses to immune checkpoint blockade (ICB) with anti-PD-L1 and anti-T cell immunoreceptor with immunoglobulin and ITIM domain (TIGIT) monoclonal antibodies (mAbs) and changes in CD8 T cell and monocyte subsets using flow cytometry. Study participants were all male, 71% (5/7) Caucasian, with a median age of 61 years, CD4 count of 739 cells/lL, and plasma HIV RNA of <50 copies/mL on stable cART for >1 year. Ex vivo polyfunctional HIV-Gag-specific CD8 T cell responses to anti-PD-L1 mAb significantly improved (p < .05) over the 8-week course of metformin therapy. Moreover, frequencies of both intermediate (CD14 + CD16 + ; r = 0.89, p = .01) and nonclassical (CD14 low CD16 + ; r = 0.92, p = .01) monocytes at entry were predictive of the magnitude of the anti-HIV CD8 T cell responses to PD-L1 blockade. Collectively, these findings highlight that 8-week course of metformin increases the polyfunctionality of CD8 T cells and that baseline monocyte subset frequencies may be a potential determinant of PD-L1 blockade efficacy. These data provide valuable information for HIV remission trials that utilize ICB strategies to enhance anti-HIV CD8 T cell immunity.

Human T-cell leukemia virus type 1 (HTLV-1) has infected approximately 20 million people worldwide. While 90% are asymptomatic, 5% develop severe diseases including adult T-cell leukemia/lymphoka (ATLL) and HTLV-1-associated myelopathy/tropical spastic paraparesis (HAM/TSP). No vaccine against HTLV-1 exists, and screening programs are not universal. However, patients with chronic HTLV-1 infection have high frequencies of HTLV-1-activated CD8+ T cells, and the two main HLA alleles (A2, A24) are present in 88% of infected individuals. We thus utilized an immunoproteomics approach to characterize MHC-I restricted epitopes presented by HLA-A2+, A24+ MT-2 and SLB-1 cell lines. Unlike traditional motif prediction algorithms, this approach identifies epitopes associated with cytotoxic T-cell responses in their naturally processed forms, minimizing differences in antigen processing and protein expression levels. Out of nine identified peptides, we confirmed six novel MHC-I restricted epitopes that were capable of binding HLA-A2 and HLA-A24 alleles and used in vitro and in vivo methods to generate CD8+ T cells specific for each of these peptides. MagPix MILLIPLEX data showed that in vitro generated epitope-specific CD8+ T cells secreted IFN-γ, granzyme B, MIP-1α, TNF-α, perforin and IL-10 when cultured in the presence of MT-2 cell line. Degranulation assay confirmed cytotoxic response through surface expression of CD107 on CD8+ T cells when cultured with MT-2 cells. A CD8+ T-cell killing assay indicated significant antiviral activity of CD8+ T cells specific against all identified peptides. In vivo generated CD8+ T cells similarly demonstrated immunogenicity on ELISpot, CD107 degranulation assay, and MagPix MILLIPLEX analysis. These epitopes are thus candidates for a therapeutic peptide-based vaccine against HTLV-1, and our results provide preclinical data for the advancement of such a vaccine.

Objectives: Chemokine receptor CCR5 is the principal co-receptor for entry of M-tropic HIV virus into immune cells. It is expressed in the central nervous system and may contribute to neuro-inflammation. The CCR5 antagonist maraviroc (MVC) has been suggested to improve HIV-associated neurocognitive impairment (NCI). Design: A double-blind, placebo-controlled, 48-week, randomized study of MVC vs placebo in people living with HIV (PLWH) on stable antiretroviral therapy (ART) > 1 year in Hawaii and Puerto Rico with plasma HIV RNA <50 copies/mL and at least mild NCI defined as an overall or domain-specific neuropsychological (NP) Z score < -0.5. Methods: Study participants were randomized 2:1 to intensification of ART with MVC vs placebo. The primary endpoint was change in global and domain-specific NP Z scores (NPZ) modeled from study entry to week 48. Covariate adjusted treatment comparisons of average changes in cognitive outcome were performed using winsorized NPZ data. Monocyte subset frequencies and chemokine expression as well as plasma biomarker levels were assessed. Results: Forty-nine participants were enrolled with 32 individuals randomized to MVC intensification and 17 to placebo. At baseline, worse NPZ scores were seen in the MVC arm. Comparison of 48-week NPZ change by arm revealed no differences except for a modest improvement in the Learning and Memory domain in the MVC arm which did not survive multiplicity correction. No significant changes between arms were seen in immunologic parameters. Conclusions: This randomized controlled study found no definitive evidence in favor of MCV intensification among PLWH with mild cognitive difficulties.