Search

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.

HTLV-1-associated myelopathy/tropical spastic paraparesis (HAM/TSP) is a progressive demyelinating disease of the spinal cord due to chronic inflammation. Hallmarks of disease pathology include dysfunctional anti-viral responses and the infiltration of HTLV-1-infected CD4+ T cells and HTLV-1-specific CD8+ T cells in the central nervous system. HAM/TSP individuals exhibit CD4+ and CD8+ T cells with elevated co-expression of multiple inhibitory immune checkpoint proteins (ICPs), but ICP blockade strategies can only partially restore CD8+ T-cell effector function. Exosomes, small extracellular vesicles, can enhance the spread of viral infections and blunt anti-viral responses. Here, we evaluated the impact of exosomes isolated from HTLV-1-infected cells and HAM/TSP patient sera on dendritic cell (DC) and T-cell phenotypes and function. We observed that exosomes derived from HTLV-infected cell lines (OSP2) elicit proinflammatory cytokine responses in DCs, promote helper CD4+ T-cell polarization, and suppress CD8+ T-cell effector function. Furthermore, exosomes from individuals with HAM/TSP stimulate CD4+ T-cell polarization, marked by increased Th1 and regulatory T-cell differentiation. We conclude that exosomes in the setting of HAM/TSP are detrimental to DC and T-cell function and may contribute to the progression of pathology with HTLV-1 infection. [ABSTRACT FROM AUTHOR]

Human T-cell leukemia virus type 1 (HTLV-1) has infected as many as 10 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-ɣ, 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.

HTLV-I-associatedmyelopathy/tropical spastic paraparesis (HAM/TSP) is a chronic, progressive, neuroinflammatory demyelinating condition of the spinal cord. We have previously shown that aberrant expression and activity of immune checkpoint (ICP) molecules such as PD-1 and PD-L1/PD-L2, negatively associates with the cytolytic potential of T cells in individuals with HAM/TSP. Interestingly, ICPs can exist in a soluble cell-free form and can be carried on extracellular vesicles (EVs) and exosomes (small EVs, <300 nm) while maintaining their immunomodulatory activity. Therefore, we investigated the role of soluble and exosomal ICPs in HTLV-1 associated neuroinflammation. For the very first time, we demonstrate a unique elevated presence of several stimulatory (CD27, CD28, 4-1BB) and inhibitory (BTLA, CTLA-4, LAG-3, PD-1, PD-L2) ICP receptors inHAM/TSP sera, and in purified exosomes from a HAM/TSP-derived HTLV-1-producing (OSP2) cells. These ICPs were found to be co-localized with the endosomal sorting complex required for transport (ESCRT) pathway proteins and exhibited functional binding with their respective ligands. Viral proteins and cytokines (primarily IFNγ) were found to be present in purified exosomes. IFNγ exposure enhanced the release of ICP molecules while antiretroviral drugs (Azidothymidine and Lopinavir) significantly inhibited this process. HTLV-1 b-Zip protein (HBZ) has been linked to factors that enhance EV release and concurrent knockdown here led to the reduced expression of ESCRT associated genes (e.g., Hrs, Vsp4, Alix, Tsg101) as well as abrogated the release of ICP molecules, suggesting HBZ involvement in this process. Moreso, exosomes from OSP2 cells adversely affected CD8 T-cell functions by diminishing levels of cytokines and cytotoxic factors. Collectively, these findings highlight exosomemediated immunomodulation of T-cell functions with HBZ and ESCRT pathways as an underlying mechanism in the context of HTLV-1-induced neuroinflammation. [ABSTRACT FROM AUTHOR]

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. [ABSTRACT FROM AUTHOR]

Infection with human T-cell leukemia/lymphoma virus type 1 (HTLV-1) has been associated with various clinical syndromes including co-infection with Strongyloides stercoralis, which is an intestinal parasitic nematode and the leading cause of strongyloidiasis in humans. Interestingly, HTLV-1 endemic areas coincide with regions citing high prevalence of S. stercoralis infection, making these communities optimal for elucidating the pathogenesis of co-infection and its clinical significance. HTLV-1 co-infection with S. stercoralis has been observed for decades in a number of published patient cases and case series; however, the implications of this co-infection remain elusive. Thus far, data suggest that S. stercoralis increases proviral load in patients co-infected with HTLV-1 compared to HTLV-1 infection alone. Furthermore, co-infection with HTLV-1 has been associated with shifting the immune response from Th2 to Th1, affecting the ability of the immune system to address the helminth infection. Thus, despite this well-known association, further research is required to fully elucidate the impact of each pathogen on disease manifestations in co-infected patients. This review provides an analytical view of studies that have evaluated the variation within HTLV-1 patients in susceptibility to S. stercoralis infection, as well as the effects of strongyloidiasis on HTLV-1 pathogenesis. Further, it provides a compilation of available clinical reports on the epidemiology and pathology of HTLV-1 with parasitic co-infection as well as data from mechanistic studies suggesting possible immunopathogenic mechanisms. Furthermore, specific areas of potential future research have been highlighted to facilitate advancing understanding of the complex interactions between these two pathogens. [ABSTRACT FROM AUTHOR]

Abstract: CCAAT/enhancer-binding protein (C/EBP) basic region/leucine zipper (bZIP) transcription factors have been shown to form heterodimers with cAMP-responsive element binding protein 2 (CREB-2), a transcription factor involved in regulating basal and Tax-mediated transactivation of the human T cell leukemia virus type 1 (HTLV-1) long terminal repeat (LTR). In cells of the monocyte–macrophage lineage (proposed to play a role in HTLV-1 pathogenesis as an accessory target cell), several members of the C/EBP family are expressed at high levels and may have functional impact on both basal and Tax-mediated transactivation of the HTLV-1 LTR. Basal activation of the HTLV-1 LTR was enhanced by overexpression of C/EBPβ, C/EBPδ, or C/EBPε, whereas transactivation of the LTR by Tax was inhibited by overexpression of C/EBPα and C/EBPβ. Inhibition of Tax-mediated transactivation of the HTLV-1 LTR was co-activator-independent, did not require C/EBP binding to the Tax-responsive elements, and may involve heterodimerization with CREB factors. [Copyright &y& Elsevier]