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

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.

Excellent Quality of Lives for Previously Treated Adult T-Cell Leukemia/Lymphoma Patients After Autologous Dendritic Cell Vaccine Therapy Targeting Htlv-1 Tax Antigens

ABSTRACT Aim: Adult T-cell leukemia/lymphoma (ATL) is a human T-cell leukemia virus type-1 (HTLV-1)-infected T-cell malignancy with extremely poor prognosis due to frequent relapse. Among HTLV-1 antigens, the viral regulatory protein Tax is widely recognized as a target of cytotoxic T lymphocytes (CTLs). We investigated the safety and feasibility of Tax peptide-pulsed dendritic cell (Tax-DC) vaccine therapy administered to augment Tax-specific CTL responses in ATL patients. Methods: Three previously treated acute-type ATL patients possessing HLA-A*0201, A*2402 and/or A*1101 in stable condition were enrolled in the study. Autologous monocyte-derived dendritic cells (DCs) that exhibited the mature phenotype were pulsed with Tax peptides corresponding to the major epitopes of Tax-specific CTL previously identified from ATL patients who received stem cell transplantation. The vaccine protocol consists of three subcutaneous injections of Tax peptide-pulsed DC at 2-week intervals in a total 8-week observation period to assess adverse events. The primary endpoint of the study was to evaluate safety and feasibility of Tax-DC vaccine. Results: DC vaccine-related toxicities were grade 1 or 2 and were considered relatively mild, and non-hematological toxicity was acceptable. Two patients achieved partial remission and one obtained stable disease. The performance status improved after vaccination in all patients and two of them are currently alive without any additional chemotherapy for 17 and 12 months respectively, while the third patient developed slowly progressive disease, although additional therapy was not required until 14 months after vaccination. In all three patients, the level of HTLV-1 proviral load in the peripheral blood mostly revealed below 100 copies per 1,000 PBMCs during the observation period, and Tax-specific CTL responses were observed with a peak at 16-20 weeks. Conclusions: The good quality of lives and long-term treatment-free survival after vaccination observed in this first pilot study indicates that the Tax-DC vaccine is a safe and promising long-lasting maintenance therapy for ATL. Disclosure: All authors have declared no conflicts of interest.

A Therapeutic Vaccine In Adult T-Cell Leukemia Patients With Autologous Dendritic Cells Pulsed With Peptides Corresponding To Tax-Specific Cytotoxic T Lymphocyte Epitopes

Adult T-cell leukemia (ATL) is an aggressive peripheral T-cell neoplasm with extremely poor prognosis. It typically develops due to long-term infection with human T-cell leukemia virus type 1 (HTLV-1). Among HTLV-1 antigens, the viral regulatory protein Tax is widely recognized as a target of cytotoxic T lymphocytes (CTLs), and Tax-specific CTLs are often activated in ATL patients following hematopoietic stem cell transplantation (HSCT). This strongly suggests the presence of Tax expression in vivo and potential contributions of CTL to GVL effects. Based on these findings, we developed an anti-ATL therapeutic vaccine consisting of autologous dendritic cells (DC) pulsed with Tax peptides corresponding to the major epitopes of Tax-specific CTL that were previously identified from human leukocyte antigen (HLA)-A2, A24 or A11-possessing ATL patients post-HSCT (Cancer Res. 2004, J Virol. 2005). In preliminary experiments, the DC induced with a conventional method showed matured phenotype and produced interleukin (IL)-12 in two of three ATL patients tested.After approval by the institutional ethics committees, we conducted a phase I clinical trial of anti-ATL immunotherapy for pretreated ATL patients in stable condition in order to evaluate whether HTLV-1 Tax targeting immunotherapy is safe and feasible. The vaccine protocol consists of three subcutaneous injections of peptide-pulsed DC at 2-week intervals in a total 8-week observation period to assess adverse events. Three acute-type patients enrolled and completed the course of the study. Modest clinical symptoms, such as injection site reaction, fever, and skin damage, were observed in all cases. DC vaccine-related toxicities were grade 1 or 2 and were considered relatively mild, and non-hematological toxicity was acceptable. In addition, significant reduction in sIL-2R levels were observed at least in one patient, and the HTLV-1 proviral loads were maintained below 60 copies /1000 PBMCs during the monitored period after administration of DC in the all patients. Enhanced proliferative responses of Tax-specific CTLs were also observed in vitro after therapy. Clinical effects, as evaluated by RECIST, were partial remission in two patients and stable disease in one patient. Durable remissions and improvements in performance status were observed in all three patients without the requirement for new treatment for 4-10 months after treatment with Tax-targeted DC vaccination. These results indicate that Tax-specific DC vaccination is safe and well tolerated, and may be promising as a new therapeutic strategy for ATL patients. Disclosures:No relevant conflicts of interest to declare.

Development of peripheral T-cell lymphoma not otherwise specified in an HTLV-1 carrier

Human T-cell leukemia virus type 1 (HTLV-1) causes adult T-cell leukemia (ATL) after a long latency period of about 60 years. As the mature T-cell neoplasms that emerge in patients infected with HTLV-1 are often ATL, T-cell neoplasms developing in such patients tend to be diagnosed simply as ATL without further investigation. However, not all T-cell neoplasms that develop in HTLV-1-infected cases are ATL. Mature T-cell malignancies other than ATL should be carefully excluded in patients infected with HTLV-1, as these sometimes closely resemble ATL in their clinical, morphological, and histological features. Here, we present a case of peripheral T-cell lymphoma not otherwise specified (PTCL-NOS) in an HTLV-1 carrier. Confirmation of monoclonal integration of the virus with Southern blotting leads to a definite diagnosis of ATL. Although we did not detect the monoclonal integration band of HTLV-1 in this case, the high HTLV-1 proviral load complicated the diagnosis. Multicolor flow cytometric analysis clearly showed that HTLV-1 was not integrated in the tumor cells, and facilitated discrimination of PTCL-NOS from ATL.

HTLV-1-associated inflammatory myopathies: Low proviral load and moderate inflammation in 13 patients from West Indies and West Africa

BackgroundThe Human T-cell Leukemia Virus type 1 (HTLV-1) is the causative agent of several inflammatory diseases, including HTLV-1-associated inflammatory myopathies (HAIM). Little is known about the virological and immunological characteristics of this viral disease. ObjectivesTo characterize the histological and virological features of HAIM patients, in order to better understand the pathogenetic mechanisms and unravel new biological markers of this disease. Study designWe conducted a retrospective study on 13 patients with HAIM, based on blood and muscle samples. We included blood samples from HTLV-1-infected individuals without myopathy as controls. Muscle biopsies were used for a broad immunohistological evaluation of tissue damage and inflammation, as well as identification of infected cells through in situ hybridization. DNA extracted from patients’ PBMC was used to identify the virus genotype by sequencing and to assess the proviral load by quantitative PCR. Anti-viral antibodies in plasma samples were titrated by indirect immunofluorescence. ResultsPatients originate from HTLV-1 endemic areas, the West Indies and West Africa. Histological alterations and inflammation in patients muscles were mostly moderate, with classical features of idiopathic myositis and rare HTLV-1-infected infiltrating cells. In all patients, HTLV-1 belonged to the A subtype, transcontinental subgroup. Anti-HTLV-1 antibodies titers were high, but the proviral load was not elevated compared to asymptomatic HTLV-1 carriers. ConclusionWe show here that muscle inflammation is moderate in HAIM, and accompanied by a low HTLV-1 proviral load, suggesting that the pathogenetic events do not exactly mirror those of other HTLV-1-associated inflammatory diseases.

Lack of evidence to support the association of a single IL28B genotype SNP rs12979860 with the HTLV-1 clinical outcomes and proviral load

BackgroundThe Interleukin 28B (IL28B) rs12979860 polymorphisms was recently reported to be associated with the human T-cell leukemia virus type 1 (HTLV-1) proviral load (PvL) and the development of the HTLV-1-associated myelopathy/tropical spastic paraparesis (HAM/TSP).MethodsIn an attempt to examine this hypothesis, we assessed the association of the rs12979860 genotypes with HTLV-1 PvL levels and clinical status in 112 unrelated Brazilian subjects (81 HTLV-1 asymptomatic carriers, 24 individuals with HAM/TSP and 7 with Adult T cell Leukemia/Lymphoma (ATLL)).ResultsAll 112 samples were successfully genotyped and their PvLs compared. Neither the homozygote TT nor the heterozygote CT mutations nor the combination genotypes (TT/CT) were associated with a greater PvL. We also observed no significant difference in allele distribution between asymptomatic carriers and patients with HTLV-1 associated HAM/TSP.ConclusionsOur study failed to support the previously reported positive association between the IL28B rs12979860 polymorphisms and an increased risk of developing HAM/TSP in the Brazilian population.

Immunopathogenesis of Human T-Cell Leukemia Virus Type-1-Associated Myelopathy/Tropical Spastic Paraparesis: Recent Perspectives

Human T-cell leukemia virus type-1 (HTLV-1) is a replication-competent human retrovirus associated with two distinct types of disease only in a minority of infected individuals: the malignancy known as adult T-cell leukemia (ATL) and a chronic inflammatory central nervous system disease HTLV-1-associated myelopathy/tropical spastic paraparesis (HAM/TSP). HAM/TSP is a chronic progressive myelopathy characterized by spastic paraparesis, sphincter dysfunction, and mild sensory disturbance in the lower extremities. Although the factors that cause these different manifestations of HTLV-1 infection are not fully understood, accumulating evidence from host population genetics, viral genetics, DNA expression microarrays, and assays of lymphocyte function suggests that complex virus-host interactions and the host immune response play an important role in the pathogenesis of HAM/TSP. Especially, the efficiency of an individual's cytotoxic T-cell (CTL) response to HTLV-1 limits the HTLV-1 proviral load and the risk of HAM/TSP. This paper focuses on the recent advances in HAM/TSP research with the aim to identify the precise mechanisms of disease, in order to develop effective treatment and prevention.

HTLV-1 proviral load levels in adult T-cell leukemia/lymphoma (ATL) in Bahia, Brazil

In human T-cell leukemia virus type 1 (HTLV-1) infection, high levels of proviral load (PVL) have been associated with the development of HTLV-1-associated myelopathy/tropical spastic paraparesis(HAM/TSP) and infective dermatitis associated with HTLV-1 (IDH). Few data are available on PVL in the different clinical types of adult T-cell leukemia/lymphoma (ATL), a malignancy associated with HTLV-1. PVL assessment may constitute a useful tool for classifying types of ATL. The objective of this study was to assess the PVL levels of patients with different clinical types of ATL. The study included 39 samples from patients with the following clinical subtypes of ATL: acute (n = 11), chronic (n = 11), smoldering (n = 10), lymphoma (n = 5) and primary cutaneous tumoral (n = 2). PVL was quantified using quantitative real-time PCR, taking the tax region of provirus into consideration and using β-globulin as the reference gene. PVL levels were highest in the chronic and acute subtypes, but there was no statistically significant difference between these two subtypes (p = 0.6936). In smoldering ATL, PVL levels were lower compared to the levels found in the acute (p = 0.0124) and chronic (p = 0.0067) subtypes. In the primary cutaneous tumoral and lymphoma subtypes, PVL levels were intermediate but there were no statistically significant differences between these and the other clinical subtypes. The present results suggest that PVL levels in ATL are related to the lymphocytosis observed in the acute and chronic subtypes, but appear to be unrelated to the aggressiveness of the disease.

Double control systems for human T‐cell leukemia virus type 1 by innate and acquired immunity

Human T-cell leukemia virus type 1 (HTLV-1) is the causative retrovirus of adult T-cell leukemia (ATL) and HTLV-1-associated myelopathy/tropical spastic paraparesis (HAM/TSP). HTLV-1-specific T-cell responses elicit antitumor and antiviral effects in experimental models, and are considered to be one of the most important determinants of the disease manifestation, since they are activated in HAM/TSP but not in ATL patients. The combination of low T-cell responses and elevated HTLV-1 proviral loads are features of ATL, and are also observed in a subpopulation of HTLV-1 carriers at the asymptomatic stage, suggesting that these features may be underlying risk factors. These risks may potentially be reduced by vaccination to activate HTLV-1-specific T-cell responses. HAM/TSP and ATL patients also differ in their levels of HTLV-1 mRNA expression, which are generally low in vivo but slightly higher in HAM/TSP patients. Our recent study indicated that viral expression in HTLV-1-infected T-cells is suppressed by stromal cells in culture through type-I IFNs. The suppression was reversible after isolation from the stromal cells, mimicking a long-standing puzzling phenomenon in HTLV-1 infection where the viral expression is very low in vivo and rapidly induced in vitro. Collectively, HTLV-1 is controlled by both acquired and innate immunity in vivo: HTLV-1-specific T-cells survey infected cells, and IFNs suppress viral expression. Both effects would contribute to a reduction in viral pathogenesis, although they may potentially influence or conflict with one another. The presence of double control systems for HTLV-1 infection provides a new concept for understanding the pathogenesis of HTLV-1-mediated malignant and inflammatory diseases.

Human T cell leukemia virus type 1 infection drives spontaneous proliferation of natural killer cells

Most human T cell leukemia virus type 1 (HTLV-1) infected subjects remain asymptomatic throughout their lives, with a few individuals developing HTLV-1 associated myelopathy/tropical spastic paraparesis (HAM/TSP) or adult T cell leukemia. Lymphocytes from about half of HTLV-1 infected subjects spontaneously proliferate in vitro, and how this phenomenon relates to symptomatic disease outcome and viral burden is poorly understood. Spontaneous proliferation was measured in lymphocyte subsets, and these findings were correlated with HTLV-1 proviral load and Tax expression in PBMCs. We found that in addition to previously described vigorous CD8+ T cell spontaneous proliferation, natural killer (NK) cells spontaneously proliferated to a similar high level, resulting in expansion of CD56-expressing NK cells. Spontaneous NK cell proliferation positively correlated with HTLV-1 proviral load but not with Tax expression or the presence of HAM/TSP. The strongest correlate with clinical outcome in this cohort was the ability of cells to express Tax, while HTLV-1 proviral load was more closely related to spontaneous NK cell proliferation. These results demonstrate that spontaneous proliferation, Tax expression, and proviral load are inter-related but not equivalent, and that spontaneous lymphocyte proliferation is not restricted to T cells, the targets of HTLV-1 infection.

Neuraminidase enhances the initial steps of human T-cell leukemia virus type 1 replication

Human T-cell leukemia virus type 1 (HTLV-1) infection is involved in the development of adult T-cell leukemia and HTLV-1-associated myelopathy/tropical spastic paraparesis. A high HTLV-1 proviral load in circulating lymphocytes of HTLV-1 carriers is a risk factor for HTLV-1-related diseases. The virus–cell interaction is linked to viral tropism and pathogenesis. Characterization of the factors that affect HTLV-1 infection is important for preventing HTLV-1 infection. HTLV-1 virions are believed to be weakly infectious under cell culture conditions; however, we found that the treatment of HTLV-1 virions with microbial neuraminidase, an enzyme catalyzing the removal of sialic acid residues from various glycoconjugates, enhanced the number of proviral DNAs in infected cells in a dose-dependent manner. Neuraminidase treatment of virions, but not target cells, enhanced viral binding and entry into cells and viral infectivity; treatment of target cells prior to infection had no effect. Moreover, the number of HTLV-1-mediated syncytia was higher in the presence of neuraminidase. Our results suggest a possible contribution of microbial agents carrying neuraminidase activity to HTLV-1 pathogenesis.

Immunogenetics of human T-cell leukemia virus type 1 (HTLV-1)-associated myelopathy/tropical spastic paraparesis (HAM/TSP)

Human T-cell leukemia virus type 1 (HTLV-1) is a replication-competent human retrovirus associated with two distinct types of disease: the malignancy known as adult T-cell leukemia (ATL) and a chronic inflammatory central nervous system disease HTLV-1-associated myelopathy/tropical spastic paraparesis (HAM/TSP), whereas the vast majority of infected individuals remain asymptomatic carriers of the virus in lifetime. It is not yet fully understood why do certain individuals develop ATL or HAM/TSP, and how does HTLV-1 persist in spite of host immune response. This review focuses on the complex virus-host interactions and the cellular immune responses to HTLV-1 infection seen in HAM/TSP patients, which are important factors in determining HTLV-1 proviral load and the risk of developing disease.

Successful treatment of adult T‐cell leukemia with unrelated cord blood transplantation

This study reports the first well-documented case of adult T-cell leukemia (ATL) successfully treated with unrelated cord blood transplantation (UCBT). A 49-year-old woman was diagnosed with acute-type of ATL. Chemotherapy induced complete remission, but the human T-cell leukemia virus type 1 (HTLV-1) proviral load was detected in mononuclear cells of her peripheral blood. The patient received UCBT with a conditioning regimen consisting of total body irradiation, cytarabine, and cyclophosphamide. She remains in remission 30 months after UCBT and the HTLV-1 proviral load has fallen to undetectable levels. This result suggests that UCBT should be a therapeutic option for ATL patients who do not have suitable donors and those who urgently require treatment.

Allogeneic Stem Cell Transplantation for Adult T-Cell Leukemia/Lymphoma

Adult T-cell leukemia/lymphoma (ATLL) develops in elderly individuals who have been infected with human T-cell leukemia virus type 1 (HTLV-1), and the prognosis for patients with ATLL has been extremely poor. Retrospective studies of allogeneic stem cell transplantation (alloSCT) for selected populations of patients have achieved several encouraging results; however, the reported incidence of transplantation-related mortality (TRM) have been high, even though more than 80% of patients received stem cells from related donors and the patients were relatively young for ATLL. This report documents a prospective feasibility study of alloSCT with reduced-intensity conditioning (RIST) for elderly ATLL patients (>50 years). Regimen-related toxicities and nonhematologic toxicities were acceptable. Fourteen of 15 evaluable patients achieved complete donor chimerism within 90 days, and 1 patient had early TRM after RIST. The HTLV-1 proviral load became undetectable in 8 of 15 patients, suggesting that RIST has potential as an antiviral treatment. The results of alloSCT are promising, and 30% to 40% of patients who achieve remission and have suitable donors can now become long-term survivors with either conventional alloSCT or RIST. It is clear that a graft-versus-ATLL effect is present after alloSCT, regardless of the conditioning regimen or the stem cell source.

Reduction of Human T-Cell Leukemia Virus Type 1 (HTLV-1) Proviral Loads in Rats Orally Infected with HTLV-1 by Reimmunization with HTLV-1-Infected Cells

Human T-cell leukemia virus type 1 (HTLV-1) persistently infects humans, and the proviral loads that persist in vivo vary widely among individuals. Elevation in the proviral load is associated with serious HTLV-1-mediated diseases, such as adult T-cell leukemia and HTLV-1-associated myelopathy/tropical spastic paraparesis. However, it remains controversial whether HTLV-1-specific T-cell immunity can control HTLV-1 in vivo. We previously reported that orally HTLV-1-infected rats showed insufficient HTLV-1-specific T-cell immunity that coincided with elevated levels of the HTLV-1 proviral load. In the present study, we found that individual HTLV-1 proviral loads established in low-responding hosts could be reduced by the restoration of HTLV-1-specific T-cell responses. Despite the T-cell unresponsiveness for HTLV-1 in orally infected rats, an allogeneic mixed lymphocyte reaction in the splenocytes and a contact hypersensitivity response in the skin of these rats were comparable with those of naive rats. HTLV-1-specific T-cell response in orally HTLV-1-infected rats could be restored by subcutaneous reimmunization with mitomycin C (MMC)-treated syngeneic HTLV-1-transformed cells. The reimmunized rats exhibited lower proviral loads than untreated orally infected rats. We also confirmed that the proviral loads in orally infected rats decreased after reimmunization in the same hosts. Similar T-cell immune conversion could be reproduced in orally HTLV-1-infected rats by subcutaneous inoculation with MMC-treated primary T cells from syngeneic orally HTLV-1-infected rats. The present results indicate that, although HTLV-1-specific T-cell unresponsiveness is an underlying risk factor for the propagation of HTLV-1-infected cells in vivo, the risk may potentially be reduced by reimmunization, for which autologous HTLV-1-infected cells are a candidate immunogen.

The genetic background as a determinant of human T-cell leukemia virus type 1 proviral load

Human T-cell leukemia virus type 1 (HTLV-1) is etiologically linked with HTLV-1-associated diseases. HTLV-1 proviral load is higher in persons with adult T-cell leukemia and HTLV-1-associated myelopathy/tropical spastic paraparesis than in asymptomatic carriers. However there are little data available on the factors controlling HTLV-1 proviral load in carriers. To study the effect of genetic background on HTLV-1 proviral load, we employed a mouse model of HTLV-1 infection that we had established. Here we analyzed nine strains of mice and found there is a great variation of proviral load among mouse strains that is not necessarily dependent on major histocompatibility complex. The antibody response is also different among these strains. To our knowledge, this is the first demonstration of the importance of the genetic background other than major histocompatibility complex controlling the HTLV-1 proviral load.

Effect of lamivudine on human T-cell leukemia virus type 1 (HTLV-1) DNA copy number, T-cell phenotype, and anti-tax cytotoxic T-cell frequency in patients with HTLV-1-associated myelopathy.

Patients with human T-cell leukemia virus type 1 (HTLV-1)-associated myelopathy/tropical spastic paraparesis (HAM/TSP) typically have a high HTLV-1 proviral load in peripheral blood mononuclear cells and abundant, activated HTLV-1-specific cytotoxic T lymphocytes (CTLs). No effective treatment for HAM/TSP has been described so far. We report a 10-fold reduction in viral DNA for five patients with HAM/TSP during treatment with the reverse transcriptase inhibitor lamivudine. In one patient with recent-onset HAM/TSP, the reduction in viral DNA was associated with a fall in the frequency of CTLs specific to two peptides in the immunodominant viral antigen Tax. The half-life of peripheral blood mononuclear cell populations was estimated from changes in viral DNA copy number, CTL frequency, reduction in CD25 expression, and the loss of dicentric chromosomes following radiation-induced damage. Each of these four different techniques indicated a cellular half-life of approximately 3 days consistent with continuous lymphocyte replication and destruction. These results indicate that viral replication through reverse transcription significantly contributes to the maintenance of HTLV-1 viral DNA load. The relative contribution of proliferation versus replication may vary between infected people.