Human T-cell Leukemia Virus Type 1 Proviral Load Research Articles

Multicenter, randomized, double-blind, placebo-controlled phase 3 study of mogamulizumab with open-label extension study in a minimum number of patients with human T-cell leukemia virus type-1-associated myelopathy.

Human T-cell leukemia virus type 1 (HTLV-1)-associated myelopathy/tropical spastic paraparesis (HAM/TSP) is a chronic neurodegenerative disease. This multicenter, randomized phase 3 study evaluated the efficacy and safety of 0.3mg/kg intravenous mogamulizumab, a monoclonal antibody targeting-CC chemokine receptor 4, every 12weeks in HAM/TSP patients. This study comprised a 24-week double-blind, placebo-controlled period, 24-week open-label period, and extension treatment period. The primary endpoint was the proportion of patients with a ≥ 1-grade improvement in the Osame motor disability score (OMDS). Secondary endpoints were changes in HTLV-1 proviral load, 10-m timed walk, cerebrospinal fluid (CSF) neopterin levels, and safety. The exploratory endpoint was CSF chemokine C-X-C motif ligand 10 (CXCL10) levels. Thirty-four and 33 patients were randomized to mogamulizumab and placebo arms, respectively. At the end of the double-blind period, no significant difference was found in the OMDS improvement rate or other secondary efficacy endpoints assessing motor activities. However, the mogamulizumab arm showed a significant decrease in HTLV-1 proviral load (- 59.39 ± 29.91% vs. placebo 2.32 ± 36.31%) and CSF neopterin (p < 0.001)/CXCL10 levels (p = 0.004). The baseline OMDS pattern and the 60-80% HTLV-1 proviral load reduction were sustained through the open-label and extension treatment periods. Although a higher incidence of rash (69.2%) was reported, the safety profile was similar compared with a previous phase 1/2a study. We found no significant difference in clinical benefit; however, mogamulizumab may provide long-term clinical benefit by preventing disease progression, as CSF neopterin/CXCL10 levels are associated with long-term prognosis in HAM/TSP.Clinical Trial Registration Number: NCT03191526 (registered date: 6-June-2017).

EZH1/2 dual inhibitors suppress HTLV-1-infected cell proliferation and hyperimmune response in HTLV-1-associated myelopathy.

Human T-cell leukemia virus type 1 (HTLV-1) causes HTLV-1-associated myelopathy (HAM), adult T-cell leukemia/lymphoma (ATL), HTLV-1-associated uveitis, and pulmonary diseases. Although both HAM and ATL show proliferation of infected cells, their pathogeneses are quite different. In particular, the pathogenesis of HAM is characterized by hyperimmune responses to HTLV-1-infected cells. Recently, we demonstrated the overexpression of histone methyltransferase EZH2 in ATL cells and the cytotoxic effects of EZH2 inhibitors and EZH1/2 dual inhibitors on these cells. However, these phenomena have never been studied in HAM. Furthermore, what effect these agents have on the hyperimmune response seen in HAM is completely unknown. In this study, we investigated histone methyltransferase expression levels in infected cell populations (CD4+ and CD4+CCR4+ cells) from patients with HAM using microarray and RT-qPCR analyses. Next, using an assay system that utilizes the spontaneous proliferation characteristic of peripheral blood mononuclear cells derived from patients with HAM (HAM-PBMCs), we investigated the effects of EZH2 selective inhibitors (GSK126 and tazemetostat) and EZH1/2 dual inhibitors (OR-S1 and valemetostat, also known as DS-3201), particularly on cell proliferation rate, cytokine production, and HTLV-1 proviral load. We also examined the effect of EZH1/2 inhibitors on the proliferation of HTLV-1-infected cell lines (HCT-4 and HCT-5) derived from patients with HAM. We found elevated expression of EZH2 in CD4+ and CD4+CCR4+ cells from patients with HAM. EZH2 selective inhibitors and EZH1/2 inhibitors significantly inhibited spontaneous proliferation of HAM-PBMC in a concentration-dependent manner. The effect was greater with EZH1/2 inhibitors. EZH1/2 inhibitors also reduced the frequencies of Ki67+ CD4+ T cells and Ki67+ CD8+ T cells. Furthermore, they reduced HTLV-1 proviral loads and increased IL-10 levels in culture supernatants but did not alter IFN-γ and TNF-α levels. These agents also caused a concentration-dependent inhibition of the proliferation of HTLV-1-infected cell lines derived from patients with HAM and increased annexin-V(+)7-aminoactinomycin D(-) early apoptotic cells. This study showed that EZH1/2 inhibitors suppress HTLV-1-infected cell proliferation through apoptosis and the hyperimmune response in HAM. This indicates that EZH1/2 inhibitors may be effective in treating HAM.

Long-term safety and efficacy of mogamulizumab (anti-CCR4) for treating virus-associated myelopathy.

Some carriers of human T-cell leukemia virus type 1 (HTLV-1), a retrovirus that primarily infects CD4+ T cells and causes lifelong infection, develop HTLV-1-associated myelopathy/tropical spastic paraparesis (HAM/TSP). Current treatments for HAM/TSP are insufficient with problematic long-term side effects. This study evaluated the long-term safety and efficacy of the anti-CCR4 antibody mogamulizumab in patients with HAM/TSP over a 4-year period. We conducted an open-label, extended long-term study (UMIN trial number: UMIN000019942) of a Phase 1-2a trial with mogamulizumab for HAM/TSP (UMIN000012655). The study participants were patients with corticosteroid-resistant HAM/TSP who could walk 10 m with or without assistive tools. Mogamulizumab was administered at 0.01, 0.03, 0.1, or 0.3 mg/kg at intervals of ≥8 weeks (0.01 and 0.03 mg/kg) or ≥12 weeks (0.1 and 0.3 mg/kg). HTLV-1 proviral load, cerebrospinal fluid inflammatory markers, and clinical symptoms were summarized by descriptive statistics. Missing observations were imputed using the last-observation-carried-forward method. As a post-hoc analysis, we evaluated the therapeutic effect of mogamulizumab on gait function by comparing it with contemporary control data from a HAM/TSP patient registry. Of the 21 participants in the Phase 1-2a, 18 (86%) enrolled in the long-term study and 15 (71%) continued repeated doses of mogamulizumab for 4 years. The median dose was 0.1 mg/kg after 4 years. Seventeen of 21 participants (81%) experienced grade 1-2 skin-related adverse events. Observed grade 3 drug-related adverse effects included three cases of lymphopenia and one case each of microscopic polyangiitis, elevated levels of aspartate aminotransferase, and neutropenia. Four of 21 participants (19%) developed neutralizing antibodies. After 4 years, the peripheral blood proviral load and the number of infected cells in CSF decreased by 60.7% and 66.3%, respectively. Neopterin and CXCL10 CSF concentrations decreased by 37.0% and 31.0%, respectively. Among the 18 participants, spasticity and Osame Motor Disability Score (OMDS) improved in 17 (94%) and 4 (22%), respectively. However, 10 m walking time worsened by 7.3% on average. Comparison with the contemporary control group demonstrated that mogamulizumab inhibited OMDS progression (p = 0.02). The results of the study suggest that mogamulizumab has long-term safety and inhibitory effect on lower limb motor disability progression in corticosteroid-treated patients with HAM/TSP. This will provide a basis for the application of mogamulizumab in HAM/TSP treatment.

HAS-Flow May Be an Adequate Method for Evaluating Human T-Cell Leukemia Virus Type 1 Infected Cells in Human T-Cell Leukemia Virus Type 1-Positive Rheumatoid Arthritis Patients Receiving Antirheumatic Therapies: A Retrospective Cross-Sectional Observation Study.

The study aims to assess the usefulness of human T-cell leukemia virus type 1 (HTLV-1)-infected cell analysis using flow cytometry (HAS-Flow) as a monitoring method for adult T-cell leukemia (ATL) development in HTLV-1-positive patients with rheumatoid arthritis (RA) under treatment with antirheumatic therapies. A total of 13 HTLV-1-negative and 57 HTLV-1-positive RA patients participated in this study, which was used to collect clinical and laboratory data, including HAS-Flow and HTLV-1 proviral load (PVL), which were then compared between the two groups. CADM1 expression on CD4+ cells in peripheral blood (PB) was used to identify HTLV-1-infected cells. The population of CADM1+ CD4+ cells was significantly higher in HTLV-1-positive RA patients compared to HTLV-1-negative RA patients. The population of CADM1+ CD4+ cells was correlated with HTLV-1 PVL values. There were no antirheumatic therapies affecting both the expression of CADM1 on CD4+ cells and PVLs. Six HTLV-1-positive RA patients who indicated both high HTLV-1 PVL and a predominant pattern of CADM1+ CD7neg CD4+ cells in HAS-Flow can be classified as high-risk for ATL progression. HAS-Flow could be a useful method for monitoring high-risk HTLV-1-positive RA patients who are at risk of developing ATL during antirheumatic therapies.

Elevation of the Plasma Levels of TNF Receptor 2 in Association with Those of CD25, OX40, and IL-10 and HTLV-1 Proviral Load in Acute Adult T-Cell Leukemia.

Adult T-cell leukemia/lymphoma (ATL) cells express TNF receptor type-2 (TNFR2) on their surface and shed its soluble form (sTNFR2). We previously reported that sTNFR2 levels were highly elevated in the plasma of patients with acute ATL. To investigate whether its quantitation would be helpful for the diagnosis or prediction of the onset of acute ATL, we examined the plasma levels of sTNFR2 in a large number of specimens obtained from a cohort of ATL patients and asymptomatic human T-cell leukemia virus type 1 (HTLV-1) carriers (ACs) and compared them to those of other candidate ATL biomarkers (sCD25, sOX40, and IL-10) by enzyme-linked immunosorbent assays (ELISA) and HTLV-1 proviral loads. We observed that sTNFR2 levels were significantly elevated in acute ATL patients compared to ACs and patients with other types of ATL (chronic, smoldering, and lymphoma). Importantly, sTNFR2 levels were significantly correlated with those of sCD25, sOX40, and IL-10, as well as proviral loads. Thus, the present study confirmed that an increase in plasma sTNFR2 levels is a biomarker for the diagnosis of acute ATL. Examination of plasma sTNFR2 alone or in combination with other ATL biomarkers may be helpful for the diagnosis of acute ATL.

Trial in Progress: A Phase II Trial of Belinostat As Consolidation Therapy with Zidovudine for Adult T-Cell Leukemia-Lymphoma (ATLL)

Background: Adult T-cell leukemia-lymphoma (ATLL), a lymphoid malignancy associated with human T-cell leukemia virus type 1 (HTLV-1), is endemic in regions such as southwestern Japan, the Caribbean basin, and parts of South America, and central Africa where HTLV-1 is prevalent. Generally, ATLL cannot be cured with chemotherapy alone, and has dismal long-term outcomes. Previous clinical trials have only yielded modest results.Zidovudine (ZDV) is a synthetic nucleoside analog that inhibits reverse transcriptase and DNA polymerase resulting in inhibition of DNA replication and cell proliferation. Studies have demonstrated ZDV plus interferon (IFNα) therapy, as induction, followed by maintenance therapy, can be an efficacious first line treatment in leukemic forms of ATLL. A meta-analysis showed ZDV/IFNα therapy compared better to conventional chemotherapy for acute and chronic leukemic type ATLL in terms of overall survival. NCCN guidelines include ZDV/IFNα as a first-line therapy option for ATLL. While ZDV-IFNα can be effective in some ATLL patients, as with chemotherapy, molecular response rates are suboptimal, and patients usually relapse and succumb to their disease. Therefore, new therapeutic approaches and strategies are urgently needed to treat ATLL.Belinostat, a pan-HDAC inhibitor, obtained accelerated approval in 2015 for treatment of relapsed/refractory PTCL based on efficacy and duration of response. We hypothesize that belinostat will exert anti-neoplastic effects in ATLL cells through a variety of molecular mechanisms including activation of silenced key cellular genes and suppression of HBZ (only HTLV-1 protein consistently expressed in all ATLL tumors) and through HDAC inhibition, belinostat will reactivate HTLV-1 provirus in ATLL cells and infected T-cell reservoirs, thus eliciting an immune response against virus infected cells.Here, we describe a trial designed to evaluate safety and response of belinostat in combination with ZDV as consolidation therapy for treatment of ATLL. Correlative studies include measuring HTLV-1 reactivation in peripheral blood T-cells, assessing cytotoxic T-cell response in vivo, and investigating molecular effects of belinostat in ATLL cells in vivo.Study Design: This phase 2, single arm, open-label trial will evaluate the combination of belinostat and ZDV as consolidation therapy followed by standard ZDV-based maintenance therapy with optional IFNα-2b or pegylated interferon-alfa-2b (PEG-IFN-α-2b). The study will include up to 20 adult participants with histologically or cytologically documented ATLL, HTLV-1 infection, who have achieved and maintained at least a partial hematologic response to prior ZDV/IFNα therapy or chemotherapy. Residual ATLL in peripheral blood is required prior to enrollment. Patients must have measurable, clinically evaluable or molecular disease, with adequate end organ and bone marrow function (unless due to lymphomatous infiltration) and KPS ≥50% or ECOG performance status ≤3.The active study period includes a treatment period of up to 8 cycles (21 days each = ~6-months). Additional assessments will occur at the end of Cycle 8, Month 9, Month 12 and an end-of-treatment visit (≤ 30-days after the last dose of study treatment). Patients who achieve/maintain CR and complete 1 year of therapy will undergo follow-up assessments every 3 months for 1 year, and survival assessment every 6 months up to Year 5.ZDV will be administered in the outpatient setting as 300mg tablets orally, three times daily, starting at least 24-hours prior to the first dose of belinostat. Belinostat will be administered as 1,000 mg/m 2 IV infusion over 30 minutes on Days 1-5, every 21 days, on cycles 1- 8, followed by continuation of ZDV (+/- IFN-α) as maintenance therapy up to the end of Month 12. ZDV may be administered for a minimum 12 months from the beginning of the study.Primary objectives are to evaluate safety and determine complete molecular response rate after addition of belinostat as consolidation therapy for ATLL during ZDV-based maintenance treatment. Secondary outcomes will include evaluation of clinical response rates, investigation of whether belinostat disrupts HTLV-1 latency load in vivo, investigation of whether belinostat provokes immune or cytotoxic T-cell response in vivo and determine the impact of belinostat/ZDV (+/- IFNα) on HTLV-1 proviral load (measure of HTLV-1 infected reservoirs), in vivo. [Display omitted] DisclosuresNo relevant conflicts of interest to declare.

The association of polymorphisms (rs2227981 and rs10204525) of PDCD1 gene with susceptibility to human T-cell leukemia virus type 1

Human T-cell leukemia virus type 1 (HTLV-1) is the first retrovirus as the causative agent of two serious diseases in human is known.Programmed cell death-1 (PD-1) is a regulatory protein that has an important role in immune system response and created exhaustion phenotype in T cells at chronic infections; therefore, it can impair anti-viral responses. Since the single nucleotide polymorphisms (SNP) in PD-1gene may influence infection with HTLV-1virus, so in this research, association between SNPs in exon 5 of PD-1 gene with susceptibility to HTLV-1 infection and proviral load (PVL) in Iran's population studied. In this case-control study, PD-1 rs2227981 and rs10204525 polymorphisms were evaluated in 81 HTLV-1 asymptomatic carriers (ACs) and 162 healthy individuals (control groups). These polymorphisms were genotyped by Polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP). Moreover, PVL was detected by quantitative real-time PCR (Q-RT-PCR). The results indicated that frequency of GA and AA genotypes in rs10204525 polymorphism was higher in ACs group (82.7%) than control group (26.5%) significantly; and GA + AA genotypes were significantly associated with HTLV-1 infection (OR = 13.244, 95%CI = 6.755–25.968, p = 0.000); but CT + TT genotypes in rs2227981 polymorphism, were as a protective factor against HTLV-1 infection (OR = 0.473, 95%CI = 0.279–0.813, P = 0.009). However, there was no significant difference between these polymorphisms and HTLV-1 PVL.

Tocilizumab has no direct effect on cell lines infected with human T-cell leukemia virus type 1.

ObjectiveIt remains unclear whether human T-cell leukemia virus type 1 (HTLV-1) infection influences therapeutic responses in patients with rheumatic diseases and whether immunosuppressive treatments increase the risk of HTLV-1-related complications in HTLV-1 carriers with rheumatic diseases. We examined the effects of tocilizumab (TCZ), an interleukin (IL)-6 receptor antagonist, on two HTLV-1-infected T-cell lines (HCT-5 and MT-2) in vitro.MethodsWe evaluated production of cytokines and chemokines, expression of HTLV-I associated genes, HTLV-1 proviral load (PVL), expression of HTLV-1 structural proteins, and apoptosis.ResultsThere were no significant differences in cytokine and chemokine levels in the culture supernatants of HCT-5 and MT-2 cells treated with phosphate-buffered saline (PBS) or TCZ. No significant differences were detected in mRNA abundance of Tax or HBZ, PVL, expression of the HTLV-1 structural protein GAG, or apoptosis among HCT-5 and MT-2 cells treated with PBS or TCZ.ConclusionsTCZ had no effect the cytokine profiles, HTLV-1 gene and protein expression, PVL, or apoptosis in HTLV-1-infected T-cell lines. Thus, TCZ treatment has no effect on HTLV-1 infection in vitro.

In vitro Evaluation of the Safety of Adalimumab for the Eye Under HTLV-1 Infection Status: A Preliminary Study.

Adalimumab (ADA), a fully human monoclonal tumor necrosis factor (TNF)-α antibody, is one of the most widely used biologics in the treatment of inflammatory diseases. However, ADA can exacerbate infectious conditions, induce paradoxical reactions such as inflammation, and cause neoplasia. Human T-cell leukemia virus type 1 (HTLV-1) is an infectious agent that induces inflammation and neoplastic infiltration in the eye. To date, numerous HTLV-1 carriers have been treated with adalimumab to suppress inflammation out of necessity, when standard anti-inflammatory drugs such as steroids and immunosuppressive agents have proven inadequate to control the inflammation. Here, we clarify the safety of adalimumab for the eye under HTLV-1 infectious conditions in vitro. We used the adult retinal pigment epithelial cell line (ARPE)-19 cell line as ocular resident cells, and used MT2 and TL-Om1 as HTLV-1-infected cells. ARPE-19 and MT2/TL-Om1 were co-cultured, and then adalimumab was administered. Production of cytokines and chemokines, TNF-α receptor (TNF-R), HTLV-1 proviral load (PVL), and apoptosis were measured to assess the effects of adalimumab. Contact between ARPE-19 and MT2/TL-Om1 produced inflammatory cytokines such as TNF, interleukin (IL)-6, IL-8 and IL-10, and transduced chemokines such as interferon-inducible protein-10 (IP-10), monocyte chemotactic protein-1 (MCP-1), monokine induced by interferon-γ (MIG), and regulated on activation, normal T cell expressed and secreted (RANTES). No inflammatory cytokines and chemokines were exacerbated by adalimumab. Expression of TNF-R on ARPE-19 and MT2/TL-Om1 cells, HTLV-1 PVLs of MT2/TL-Om1 cells, and cell growth rate and apoptotic rate of ARPE-19 were unaffected by adalimumab. In conclusion, adalimumab does not appear to exacerbate HTLV-1-associated inflammatory conditions in the eye or increase PVL in HTLV-1-infected T cells. These data suggest that adalimumab could be used safely for the eye under HTLV-1 infectious conditions from the perspective of in vitro assessment.

Pulmonary Disease Is Associated With Human T-Cell Leukemia Virus Type 1c Infection: A Cross-sectional Survey in Remote Aboriginal Communities.

The human T-cell leukemia virus type 1 (HTLV-1) subtype c is endemic to central Australia. We report the first large-scale, community-based, health survey of HTLV-1 and its disease associations in this setting. Aboriginal community residents aged >2 years in 7 remote communities were invited to do a health survey that included a questionnaire, spirometry, and clinical examination by a physician blinded to HTLV-1 status, clinical records, and spirometry results. Blood was drawn for HTLV-1 serology and proviral load (PVL). Pulmonary disease was assessed clinically and spirometrically and, where records were available, radiologically after the clinical assessment. Associations between specific diseases and HTLV-1 status were determined using logistic regression, adjusting for available confounders. Overall, 579 residents (164 children aged 3-17 years; 415 adults) were examined (37.7% of the estimated resident population). HTLV-1 prevalences for children and adults were 6.1% and 39.3%, respectively. No associations were found between HTLV-1 and any assessed clinical condition among children. Chronic pulmonary disease and gait abnormalities were more common among adults with HTLV-1 infection. Adjusted odds ratios among participants with PVL ≥1000 per 105 peripheral blood leukocytes were 7.08 (95% confidence interval [CI], 2.67-18.74; P < .001), 9.81 (95% CI, 3.52-27.35; P < .001), and 14.4 (95% CI, 4.99-41.69; P < .001) for clinically defined chronic pulmonary disease, moderate-severe expiratory airflow limitation, and radiologically determined bronchiectasis/bronchiolitis, respectively, and 5.21 (95% CI, 1.50-18.07; P = .009) for gait abnormalities. In the first study of HTLV-1 disease associations based on community recruitment and blinded assessment, HTLV-1 infection was strongly associated with pulmonary disease and gait abnormalities.

Inhibition of ABL1 tyrosine kinase reduces HTLV-1 proviral loads in peripheral blood mononuclear cells from patients with HTLV-1-associated myelopathy/tropical spastic paraparesis.

Human T-cell leukemia virus type 1 (HTLV-1) causes incurable adult T-cell leukemia and HTLV-1-associated myelopathy/tropical spastic paraparesis (HAM/TSP). Patients with HAM/TSP have increased levels of HTLV-1-infected cells compared with asymptomatic HTLV-1 carriers. However, the roles of cellular genes in HTLV-1-infected CD4+ T cells await discovery. We performed microarray analysis of CD4+ T cells from HAM/TSP patients and found that the ABL1 is an important gene in HAM/TSP. ABL1 is a known survival factor for T- and B-lymphocytes and is part of the fused gene (BCR-ABL) known to be responsible for chronic myelogenous leukemia (CML). ABL1 tyrosine kinase inhibitors (TKIs), including imatinib, nilotinib, and dasatinib, are used clinically for treating CML. To evaluate whether ABL1 is indeed important for HAM/TSP, we investigated the effect of TKIs on HTLV-1-infected cells. We developed a propidium monoazide-HTLV-1 viability quantitative PCR assay, which distinguishes DNA from live cells and dead cells. Using this method, we were able to measure the HTLV-1 proviral load (PVL) in live cells alone when peripheral blood mononuclear cells (PBMCs) from HAM/TSP cases were treated with TKIs. Treating the PBMCs with nilotinib or dasatinib induced significant reductions in PVL (21.0% and 17.5%, respectively) in live cells. Furthermore, ABL1 siRNA transfection reduced cell viability in HTLV-1-infected cell lines, but not in uninfected cell lines. A retrospective survey based on our clinical records found a rare case of HAM/TSP who also suffered from CML. The patient showed an 84.2% PVL reduction after CML treatment with imatinib. We conclude that inhibiting the ABL1 tyrosine kinase specifically reduced the PVL in PBMCs from patients with HAM/TSP, suggesting that ABL1 is an important gene for the survival of HTLV-1-infected cells and that TKIs may be potential therapeutic agents for HAM/TSP.

Safety of Infliximab for the Eye Under Human T-Cell Leukemia Virus Type 1 Infectious Conditions in vitro.

Use of biologics has been widely advocated for inflammatory diseases recently. Anti-tumor necrosis factor (TNF)-α antibody therapy is reportedly effective against ocular inflammation. However, side effects of TNF-α inhibition have been reported, particularly in the form of exacerbation of infections such as tuberculosis. Paradoxical reactions such as exacerbated inflammation are also well known. Around 20 million humans are infected with human T-cell leukemia virus type 1 (HTLV-1) globally, and this virus can cause adult T-cell leukemia, HTLV-1-associated myelopathy and HTLV-1 uveitis. As for ophthalmic concerns, it has not been identified whether anti-TNF-α antibody stimulates HTLV-1-infected cells and ocular cells to induce HTLV-1 uveitis in HTLV-1 carriers. Here we investigated the effects of anti-TNF-α antibody on ocular status under HTLV-1 infectious conditions using ocular cells and HTLV-1-infected cells in vitro. We used the ARPE-19 human retinal pigment epithelial cell line as ocular cells considered to play an important role in the blood-ocular barrier, and the MT2 HTLV-1-infected cell line. Jurkat cells were used as controls. Infliximab (IFX) was used as an anti-TNF-α antibody to achieve TNF-α inhibition. We evaluated the production of inflammatory cytokines and intercellular adhesion molecule (ICAM)-1, proliferation of ARPE-19, expression of TNF-α receptor (TNF-R) and HTLV-1 proviral DNA, and the percentage of apoptotic ARPE-19. Inflammatory cytokines such as interleukin (IL)-6, IL-8, TNF, and ICAM-1 were significantly elevated through contact between ARPE-19 and MT2. Treatment with IFX tented to inhibit TNF production, although the level of production was low, but changes in IL-6, IL-8, and ICAM-1 remained unaffected. Expression of TNFR was unaltered by IFX treatment. HTLV-1 proviral DNA was not significantly changed with treatment. No change in cell growth rate or apoptotic rate of ARPE-19 was seen with the addition of IFX. In conclusion, IFX did not exacerbate production of inflammatory cytokines, and did not affect expression of TNFR, proliferation of ARPE-19, HTLV-1 proviral load, or apoptosis of ARPE-19. These results suggest that IFX does not exacerbate HTLV-1-related inflammation in the eye and represents an acceptable treatment option under HTLV-1 infectious conditions.

Association of high levels of plasma OX40 with acute adult T-cell leukemia.

OX40, a member of the tumor necrosis factor receptor (TNFR) superfamily, co-stimulates activated T cells following interaction with its own ligand OX40L. Human T-cell leukemia virus type-1 (HTLV-1) is an etiological agent of adult T-cell leukemia (ATL). ATL cells are known to express cell surface OX40; however, the level of soluble OX40 (sOX40) in blood samples from ATL patients is unknown. Quantitative enzyme-linked immune-sorbent assay (ELISA) showed that sOX40 levels were significantly higher in plasma from acute ATL patients than those from asymptomatic HTLV-1 carriers and healthy donors, and correlated with sCD25 levels and HTLV-1 proviral loads in peripheral blood mononuclear cells (PBMCs). Fresh PBMCs from acute ATL patients showed a higher percentage of OX40-positive cells compared with those from carriers, and shed sOX40 into culture supernatants. Shedding of sOX40 was partially inhibited by a matrix metalloproteinase (MMP) inhibitor, GM6001. A fraction of sOX40 was capable of binding to OX40L. These results suggest that high levels of sOX40 are shed into blood from a large number of ATL cells in acute ATL patients. Thus, abnormally elevated plasma sOX40 levels may be useful as an additional diagnostic marker of acute ATL.

The time-sequential changes of risk factors for adult T-cell leukemia development in human T-cell leukemia virus-positive patients with rheumatoid arthritis: a retrospective cohort study

Objective: This study aimed to investigate the time-sequential changes of risk factors for adult T-cell leukemia (ATL) development in human T-cell leukemia virus type 1 (HTLV-1)-positive rheumatoid arthritis (RA) patients.Methods: HTLV-1 infection was screened using particle agglutination assay and confirmed via western blotting in 365 RA patients. Twenty-three HTLV-1-positive RA patients were included in the study cohort. Blood samples were obtained from these patients at each observation time point. The values of HTLV-1 proviral load (PVL) and serum soluble IL-2 receptor (sIL2-R), which are risk factors for ATL development, were measured using real-time PCR and enzyme immunoassay, respectively.Results: The study cohort comprised 79 person-years. The median HTLV-1 PVL and sIL2-R values of the HTLV-1-positive RA patients were 0.44 copies per 100 white blood cells (WBCs) and 406 U/mL, respectively. Three HTLV-1-positive RA patients showed a high PVL value. No remarkable changes were observed in the PVL and sIL2-R values during the observation period. However, one elderly HTLV-1-positive RA patient who had a high PVL value developed ATL during treatment with methotrexate and infliximab.Conclusion: A thorough clinical assessment of the risk factors for ATL development may be necessary in daily clinical practice for RA patients in HTLV-1-endemic areas in Japan.

HTLV-1 proviral load in cerebrospinal fluid may not be a good marker to differentiate asymptomatic carriers with high proviral load in blood from HAM/TSP patients.

An elevated human T cell leukemia virus type 1 (HTLV-1) proviral load (PVL) is an important risk factor for HTLV-1-associated myelopathy/tropical spastic paraparesis (HAM/TSP), although there is a considerable frequency of asymptomatic carriers (AC) with high PVL in blood. Our objective was to evaluate whether PVL quantified in cerebrospinal fluid (CSF) is helpful to distinguish AC from HAM when AC have high PVL in blood (ACH). ACH (n = 7) were characterized to have high PVL in blood by quantification of samples collected over time (mean 7years). HAM patients (n = 14) also had analyzed blood samples collected at different times (mean 9years). Comparing paired CSF and blood samples of each individual, CSF PVL mean was 4.7-fold higher than blood PVL in the ACH group and 10.8-fold in the HAM group. CSF PVL was significantly greater than blood PVL in the HAM group (p = 0.004), but not in the ACH group. Important to highlight, CSF PVL was not significantly different between the ACH and the HAM groups. These results suggested that significantly higher PVL in CSF than in blood is a hallmark of HAM/TSP patients, but this is also true for asymptomatic carriers with high PVL in blood, thus reducing its usefulness as a marker for HAM/TSP. A greater number of ACH should be analyzed, but whether they will eventually develop HAM/TSP or why they have not developed the disease are still questions to be clarified. Longitudinal studies are necessary to answer these questions.

How to Control HTLV-1-Associated Diseases: Preventing de Novo Cellular Infection Using Antiviral Therapy.

Five to ten million individuals are infected by Human T-cell Leukemia Virus type 1 (HTLV-1). HTLV-1 is transmitted through prolonged breast-feeding, by sexual contacts and by transmission of infected T lymphocytes through blood transfusion. One to ten percent of infected carriers will develop a severe HTLV-1-associated disease: Adult-T-cell leukemia/lymphoma (ATLL), or a neurological disorder named Tropical Spastic Paraparesis/HTLV-1 Associated Myelopathy (TSP/HAM). In vivo, HTLV-1 is mostly detected in CD4+ T-cells, and to a lesser extent in CD8+ T cells and dendritic cells. There is a strong correlation between HTLV-1 proviral load (PVL) and clinical status of infected individuals. Thus, reducing PVL could be part of a strategy to prevent or treat HTLV-1-associated diseases among carriers. Treatment of ATLL patients using conventional chemotherapy has very limited benefit. Some chronic and acute ATLL patients are, however, efficiently treated with a combination of interferon α and zidovudine (IFN-α/AZT), to which arsenic trioxide is added in some cases. On the other hand, no efficient treatment for TSP/HAM patients has been described yet. It is therefore crucial to develop therapies that could either prevent the occurrence of HTLV-1-associated diseases or at least block the evolution of the disease in the early stages. In vivo, reverse transcriptase (RT) activity is low in infected cells, which is correlated with a clonal mode of viral replication. This renders infected cells resistant to nucleoside RT inhibitors such as AZT. However, histone deacetylase inhibitors (HDACi) associated to AZT efficiently induces viral expression and prevent de novo cellular infection. In asymptomatic STLV-1 infected non-human primates, HDACi/AZT combination allows a strong decrease in the PVL. Unfortunately, rebound in the PVL occurs when the treatment is stopped, highlighting the need for better antiviral compounds. Here, we review previously used strategies targeting HTLV-1 replication. We also tested a series of HIV-1 RT inhibitors in an in vitro anti-HTLV-1 screen, and report that bis-POM-PMEA (adefovir dipivoxil) and bis-POC-PMPA (tenofovir disoproxil) are much more efficient compared to AZT to decrease HTLV-1 cell-to-cell transmission in vitro. Our results suggest that revisiting already established antiviral drugs is an interesting approach to discover new anti-HTLV-1 drugs.

Absolute quantification of HTLV-1 basic leucine zipper factor (HBZ) protein and its plasma antibody in HTLV-1 infected individuals with different clinical status.

BackgroundHuman T cell leukemia virus type 1 (HTLV-1) basic leucine zipper factor (HBZ), which is encoded by a minus strand mRNA, is thought to play important roles in the development of adult T-cell leukemia (ATL) and HTLV-1-associated myelopathy/tropical spastic paraparesis (HAM/TSP). However, a comprehensive analysis of HBZ, including mRNA and protein expression, humoral immunoreactivity against HBZ, and HTLV-1 proviral load (PVL), in HTLV-1-infected individuals with different clinical status has not been reported previously.ResultsIn this study, using novel monoclonal antibody-based in-house enzyme-linked immunosorbent assay systems, we report the absolute quantification of HBZ protein and its plasma antibody in clinical samples from HTLV-1-infected individuals with different clinical status. The data were compared to both HBZ mRNA levels and PVL. The results showed that plasma anti-HBZ antibody was detectable only in 10.4 % (5/48) of asymptomatic carriers (ACs), 10.8 % (13/120) of HAM/TSP patients, and 16.7 % (7/42) of ATL patients. HBZ protein was detected in three out of five patients with acute ATL, but was not detected in patients with HAM/TSP (0/10) or ACs (0/4). Thus, an antibody response to HBZ was not associated with the PVL or the expression of HBZ (both at the mRNA and protein levels) or the clinical status of the infection.ConclusionsThe present results emphasize the extremely low expression and immunogenicity of HBZ in natural HTLV-1 infection. However, there is a possibility that the low but distinct expression of HBZ protein in PBMCs is associated with the survival of HTLV-1-infected cells and the development of ATL.Electronic supplementary materialThe online version of this article (doi:10.1186/s12977-016-0263-z) contains supplementary material, which is available to authorized users.

Plasmatic proinflammatory chemokines levels are tricky markers to monitoring HTLV-1 carriers

The human T-cell leukemia virus type 1 (HTLV-1) is present throughout the world and is associated with HTLV-1-associated myelopathy/tropical spastic paraparesis (HAM/TSP) and other inflammatory conditions. The pathogenesis of HAM/TSP involves a chronic inflammatory response in central nervous system (CNS), with the presence of HTLV-1 infected cells and HTLV-1-specific CD8+ lymphocytes. Chemokines may have a role in the infiltration of these cells into the CNS. In this context, the present study analyzed the level of plasmatic chemokines CCL2 (MCP-1), CCL5 (RANTES), IL8 (CXCL8), CXCL9 (MIG), and CXCL10 (IP-10) and HTLV-1 proviral load from peripheral blood in 162 asymptomatic carriers and 136 HAM/TSP patients to determine the differences that be associated with the clinical status of the HTLV-1 infection. The results showed that patients with HAM/TSP have significantly higher levels of IL8 and CXCL9, and that the level of IL8, CXCL9 and CXCL10 was significantly greater in HTLV-1 infected individuals with high (>1%) than those with low proviral load (<1%). However, the levels of the chemokines tested have not showed high sensitivity to discriminate HAM/TSP patients from asymptomatic carriers. In addition, chemokine profiles in asymptomatic carriers and HAM/TSP groups were similar, with no significant increased frequency of higher producers of chemokines in HAM/TSP individuals. Results indicate that the heterogeneity of the individuals in the groups regarding time of infection, duration of disease, proviral load level and other possible confound factors may impair the use of chemokines levels to monitor HTLV-1 carriers in clinical practice. J. Med. Virol. 88:1438-1447, 2016. © 2016 Wiley Periodicals, Inc.

Triple Therapy with Prednisolone, Pegylated Interferon and Sodium Valproate Improves Clinical Outcome and Reduces Human T-Cell Leukemia Virus Type 1 (HTLV-1) Proviral Load, Tax and HBZ mRNA Expression in Patients with HTLV-1-Associated Myelopathy/Tropical Spastic Paraparesis.

Considering that there is no effective treatment for human T-cell leukemia virus type 1 (HTLV-1)-associated myelopathy/tropical spastic paraparesis, this study aimed to assess the impact of triple combination therapy-interferon-α, valproic acid, and prednisolone-on clinical outcomes, main HTLV-1 viral factors, and host anti-HTLV-1 antibody response. HTLV-1 proviral load (PVL), and HBZ and Tax mRNA expression levels were measured in peripheral blood mononuclear cells of 13 patients with HTLV-1-associated myelopathy/tropical spastic paraparesis before and after treatment with 180 μg pegylated interferon once a week, 10-20 mg/kg/day sodium valproate, and 5 mg/day prednisolone for 25 weeks using a TaqMan real-time polymerase chain reaction assay. Furthermore, anti-HTLV-1 titer, Osame Motor Disability Score, Ashworth spasticity scale, and urinary symptoms (through standard questionnaire and clinical monitoring) were assessed in patients before and after the treatment. HTLV-1 PVL and HBZ expression significantly decreased after the treatment [PVL from 1443 ± 282 to 660 ± 137 copies/10(4) peripheral blood mononuclear cells (p = 0.01); and HBZ from 8.0 ± 1.5 to 3.0 ± 0.66 (p < 0.01)]. Tax mRNA expression decreased after the treatment from 2.26 ± 0.45 to 1.44 ± 0.64, but this reduction was not statistically significant (p = 0.10). Furthermore, anti-HTLV-1 titer reduced dramatically after the treatment, from 3123 ± 395 to 815 ± 239 (p < 0.01). Clinical signs and symptoms, according to Osame Motor Disability Score and Ashworth score, improved significantly (both p < 0.01). Urinary symptoms and sensory disturbances with lower back pain were reduced, though not to a statistically significant degree. Although signs and symptoms of spasticity were improved, frequent urination and urinary incontinence were not significantly affected by the triple therapy. The results provide new insight into the complicated conditions underlying HTLV-1-associated diseases.

Advanced human T-cell leukemia virus type 1 carriers and early-stage indolent adult T-cell leukemia-lymphoma are indistinguishable based on CADM1 positivity in flow cytometry.

We previously reported that the cell adhesion molecule 1 (CADM1) versus CD7 plot in flow cytometry reflects disease progression in human T-cell leukemia virus type 1 (HTLV-1) infection. In CD4+ cells from peripheral blood, CADM1−CD7+ (P), CADM1+CD7dim (D) and CADM1+CD7− (N) subpopulations are observed. The D and N subpopulations increase as asymptomatic HTLV-1 carriers (AC) progress to indolent adult T-cell leukemia-lymphoma (ATL) and the N subpopulation then expands in aggressive ATL. In the present study we examined whether the analysis can estimate the risk of developing ATL in advanced AC. Peripheral blood samples from AC (N = 41) and indolent ATL patients (N = 19) were analyzed by flow cytometry using the CADM1 versus CD7 plot for CD4+ cells and inverse long PCR (clonality analysis) of FACS-sorted subpopulations. Almost all AC with a high HTLV-1 proviral load (>4 copies/100 cells) had a CADM1+ (D + N) frequency of >10%. AC with 25% < CADM1+ ≤ 50% contained expanded clones similar to smoldering-type ATL. In many patients in the 25% < CADM1+ ≤ 50% group, the proportion of abnormal lymphocytes was distributed around the 5% line, which divides AC and smoldering-type ATL in Shimoyama's classification. In conclusion, the CADM1 versus CD7 plot is useful for selection of putative high-risk AC. The characteristics of some AC and smoldering ATL are said to be similar; however, long-term follow up is required and the clinical outcome (e.g. rate of transformation) of these cases should be used to determine whether to include them in the same clinical category.