Whole body clonality analysis in an aggressive STLV-1 associated leukemia (ATLL) reveals an unexpected clonal complexity.

HTLV-1 causes Adult T cell Leukemia/Lymphoma (ATLL) in humans. We describe an ATL-like disease in a 9 year-old female baboon naturally infected with STLV-1 (the simian counterpart of HTLV-1), with a lymphocyte count over 1010/L, lymphocytes with abnormal nuclear morphology, and pulmonary and skin lesions. The animal was treated with a combination of AZT and alpha interferon. Proviral load (PVL) was measured every week. Because the disease continued to progress, the animal was euthanized. Abnormal infiltrates of CD3+CD25+ lymphocytes and Tax-positive cells were found by histological analyses in both lymphoid and non-lymphoid organs. PVL was measured and clonal diversity was assessed by LM-PCR (Ligation-Mediated Polymerase Chain Reaction) and high throughput sequencing, in blood during treatment and in 14 different organs. The highest PVL was found in lymph nodes, spleen and lungs. One major clone and a number of intermediate abundance clones were present in blood throughout the course of treatment, and in organs. These results represent the first multi-organ clonality study in ATLL. We demonstrate a previously undescribed clonal complexity in ATLL. Our data reinforce the usefulness of natural STLV-1 infection as a model of ATLL.

Enhancement of anti-STLV-1/HTLV-1 immune responses through multimodal effects of anti-CCR4 antibody.

Human T-cell leukemia virus type 1 (HTLV-1) causes adult T-cell leukemia and inflammatory diseases. Because anti-HTLV-1 immune responses are critical for suppressing infected cells, enhancing cellular immunity is beneficial for the treatment of HTLV-1-associated diseases. Using simian T-cell leukemia virus type 1 (STLV-1) infected Japanese macaques, we analyzed the immune responses to viral antigens and the dynamics of virus-infected cells. The chemokine receptor CCR4 is expressed on STLV-1 infected cells, and administration of humanized monoclonal antibody to CCR4, mogamulizumab, dramatically decreased the number of STLV-1-infected cells in vivo. Concurrently, mogamulizumab treatment enhanced STLV-1 specific CD4(+) and CD8(+) T cell responses by simultaneously targeting CCR4(+) effector regulatory T (Treg) cells and infected cells. Mogamulizumab promoted the phagocytosis of CCR4(+) infected cells by macrophages, which likely enhanced antigen presentation. Vaccination with recombinant vaccinia virus (rVV) expressing viral antigens suppressed the proviral load and the number of Tax-expressing cells. Enhanced T-cell responses were also observed in some ATL patients who were treated with mogamulizumab. This study shows that mogamulizumab works not only by killing CCR4(+) infected cells directly, but also by enhancing T cell responses by increasing the phagocytosis of infected cells by antigen-presenting cells and suppressing CCR4(+) effector Treg cells.

Thymosin α 1 potentiates the release by CD8(+) cells of soluble factors able to inhibit HIV-1 and human T lymphotropic virus 1 infection in vitro.

BACKGROUND: Thymosin α-1 (Tα1) exploits a specific action on lymphoid cells and is able to induce in peripheral blood mononuclear cells (PBMCs) a strong transcriptional response. CD8 antiviral factor activity plays a role in the control or prevention of HIV-1 infection by a non-cytolytic mechanism. The ability of Tα1 to modulate the release of antiretroviral soluble factors by CD8(+) cells was investigated. METHODS: Supernatants from lipopolysaccharide (LPS) stimulated CD8(+)-isolated cells treated with Tα1 were screened on in vitro infection of human monocyte-derived macrophages (MDMs) and PBMCs with HIV-1, and of PBMCs with human T lymphotropic virus 1 (HTLV-1). In CD8(+) cells, as well as in PBMCs of healthy donors as from HIV(+) individuals, a microarray analysis to assess the transcriptional response after treatment was performed. RESULTS: Tα1 potentiates the release, in LPS-stimulated CD8(+) cells, of soluble factors able to inhibit both in vitro HIV-1 infection of MDMs and PBMCs and in vitro HTLV-1 infection of PBMCs. A distinctive transcriptional profile was induced by Tα1 in PBMCs from HIV(+) donors. CONCLUSIONS: These findings suggest that Tα1 would represent a re-evaluated approach to antiretroviral therapy in combination with innovative treatments and with vaccine administration.

Highly active antiretroviral treatment against STLV-1 infection combining reverse transcriptase and HDAC inhibitors.

Approximately 3% of all human T-lymphotropic virus type 1 (HTLV-1)-infected persons will develop a disabling inflammatory disease of the central nervous system known as HTLV-1-associated myelopathy/tropical spastic paraparesis, against which there is currently no efficient treatment. As correlation exists between the proviral load (PVL) and the clinical status of the carrier, it is thought that diminishing the PVL could prevent later occurrence of the disease. We have conducted a study combining valproate, an inhibitor of histone deacetylases, and azidothymidine, an inhibitor of reverse transcriptase, in a series of baboons naturally infected with simian T-lymphotropic virus type 1 (STLV-1), whose PVL was equivalent to that of HTLV-1 asymptomatic carriers. We show that the combination of drugs caused a strong decrease in the PVL and prevented the transient rise in PVL that is seen after treatment with histone deacetylases alone. We then demonstrate that the PVL decline was associated with an increase in the STLV-1-specific cytotoxic T-cell population. We conclude that combined treatment with valproate to induce viral expression and azidothymidine to prevent viral propagation is a safe and effective means to decrease PVL in vivo. Such treatments may be useful to reduce the risk of HAM/TSP in asymptomatic carriers with a high PVL.

Neuromyelitis optica associated with subacute human T-lymphotropic virus type 1 infection.

Although human T-lymphotropic virus type 1 (HTLV-1)-associated myelopathy, or tropical spastic paraparesis (HAM/TSP), is usually considered as a progressive myelopathy, a subacute variant has been described. It is unusual for optic neuritis (ON) to be associated with an HTLV-1 infection. Neuromyelitis optica (NMO) is characterised by severe attacks of acute transverse myelitis and ON of unknown aetiology. We report a 61-year-old Afro-Caribbean male patient with subacute HAM/TSP associated with bilateral ON that occurred 5years previously. To our knowledge this is the first report of recurrent NMO syndrome associated with HTLV-1 infection.

Discovery and significance of new human T-lymphotropic viruses: HTLV-3 and HTLV-4.

Human T-lymphotropic virus type 1 (HTLV-1) and type 2 (HTLV-2) were discovered approximately 30 years ago and they are associated with various lymphoproliferative and neurological diseases. The estimated number of infected people is 10-20 million worldwide. In 2005, two new HTLV-1/HTLV-2-related viruses were detected, HTLV-3 and HTLV-4, from the same geographical area of Africa. In the last 4 years, their complete genomic sequences were determined and some of their characteristic features were studied in detail. These newly discovered retroviruses alongside their human (HTLV-1 and -2) and animal relatives (simian T-lymphotropic virus type 1-3) are reviewed. The potential risks associated with these viruses and the potential antiretroviral therapies are also discussed.

Histone deacetylase inhibitors induce growth arrest and apoptosis of HTLV-1-infected T-cells via blockade of signaling by nuclear factor kappaB.

Adult T-cell leukemia/lymphoma (ATL) is a highly aggressive disease with a poor prognosis in which nuclear factor kappa B (NF-kappaB) is thought to play a role. This study explored the effects of histone deacetylase inhibitors (HDACIs) MS-275, suberoylanilide hydroxamic acid (SAHA), and LBH589 on both human T-cell lymphotropic virus type I (HTLV-1)-infected T cells (MT-1, -2, -4, and HUT102) and freshly isolated ATL cells harvested from patients. HDACIs effectively inhibited the proliferation of these cells. For example, MS-275, SAHA, and LBH589 effectively inhibited the proliferation of MT-1 cells with ED(50s) of 6microM, 2.5microM, and 100nM, respectively, as measured by 3-(4,5-dimethylithiazol-2-yl)-2,5-diphenyl tetrazolium bromide assay on day 2 of culture. In addition, HDACIs induced cell cycle arrest at the G2/M phase and apoptosis of HTLV-1-infected T-cells in conjunction with regulation of apoptosis-related proteins. Electrophoretic mobility shift assay showed that exposure of HTLV-1-infected T-cells to HDACIs for 48h inhibited formation of the NF-kappaB/DNA binding complex. Moreover, we found that HDACIs accumulated NF-kappaB and inhibitory subunit of NF-kappaB in the cytoplasm in conjunction with the down-regulation of NF-kappaB in the nucleus, suggesting that HDACIs blocked nuclear translocation of NF-kappaB. Based on these findings, we believe HDACIs can be useful for treating patients with ATL or other types of cancer in which NF-kappaB plays a role.

12th International Conference on Human Retrovirology: HTLV and Related Retroviruses.

The 12th International Conference on Human Retrovirology: HTLV and Related Retroviruses, was held at the Half Moon Hotel in Montego Bay, Jamaica, from June 22nd to June 25th 2005. The scientific conference, sponsored by the International Retrovirology Association, is held biennially at rotating international venues around the world. The meeting brings together basic scientists, epidemiologists and clinical researchers to discuss findings to prevent HTLV infection or develop new therapies against HTLV-mediated diseases. The Association fosters the education and training of young scientists to bring new approaches to the complex problems of HTLV research, such as translational research to bring findings from the laboratory into clinical trials that benefit HTLV-infected patients. The breadth and quality of research presentations and workshops at the 12th International Conference indicate that these goals are being accomplished. As HTLV research enters its third decade a new generation of scientists face many challenges. However, HTLV scientists and clinicians displayed exciting new approaches and discoveries during plenary talks and poster sessions. The conference encouraged research in HTLV infections and disease, fostered collaborations, and stimulated new partnerships between clinicians and scientists to encourage clinical trials and novel therapeutic interventions.

Dichotomy of all-trans retinoic acid inducing signals for adult T-cell leukemia.

We previously reported that all-trans retinoic acid (ATRA) inhibits growth in human T-cell leukemia virus type 1 (HTLV-1)-positive T-cell lines and fresh cells from patients with adult T-cell leukemia. However, the mechanism of this inhibition is not clear. In the present study, we observed that NF-kappaB transcriptional activity as well as cell growth decreased significantly in HTLV-1-positive T-cell lines in the presence of ATRA. Furthermore, we observed that ATRA reduced HTLV-1 proviral DNA, HTLV-1 genes (gag, tax, or pol mRNA) using the real-time quantitative polymerase chain reaction. SIL-2R was reduced by ATRA in both protein level (culture supernantant) and mRNA level in HTLV-1-positive T-cell lines. Interestingly, ATRA significantly inhibited RT activity similar to azidothimidine (AZT) in HTLV-1-positive T-cell lines. Moreover, AZT inhibited proviral DNA but not NF-kappaB transcriptional activity, and sIL-2R on HTLV-1; however, ATRA inhibited of NF-kappaB, proviral DNA and sIL-2R on HTLV-1. These results suggested that the decrease in sIL-2R induced by ATRA may be caused by the actions of a NF-kappaB inhibitor acting on the NF-kappaB/sIL-2R signal pathway. These results suggested that ATRA could have two roles, as a NF-kappaB inhibitor and as an RT inhibitor.

International Retrovirology Association brings together scientists and clinicians to bridge discoveries about human T-lymphotropic viruses from the laboratory to clinical trials.

Human T-lymphotropic virus type 1 (HTLV-1) and HTLV-2 were among the first human retroviruses discovered in the early 1980's. The International Retrovirology Association is an organized effort that fostered the efforts of scientists and clinicians to form interdisciplinary groups to study this group of retroviruses and their related diseases. The Association promotes excellent science, patient education, and fosters the training of young scientists to promote "bench-to-bedside" research. The International Conference on Human Retrovirology: HTLV and Related Viruses sponsored by the Association supports clinicians and researchers in the exchange of research findings and stimulation of new research directions. This years conference will be held from June 22 to 25, in Montego Bay, Jamaica http://www.htlvconference.org.jm/. Since its inception in 1988, these conferences have provided a highly interactive forum for the global community of HTLV scientists. This is of particular importance as HTLV research enters its third decade and a new generation of scientists takes over this important work. Many of the scientists attending the meeting will be from developing countries where HTLV is endemic, consistent with the history of international collaborations that have characterized HTLV research. The International Conference on Human Retrovirology provides a unique opportunity for researchers of all disciplines interested in HTLV infections to meet their peers and to address the questions facing clinicians and scientists who study retroviruses, like HTLV.

Clinical similarities and differences between human T-cell lymphotropic virus type 1-associated bronchiolitis and diffuse panbronchiolitis.

STUDY OBJECTIVES: Human T-cell lymphotropic virus type 1 (HTLV-1)-associated bronchiolitis and diffuse panbronchiolitis might overlap. We examined whether these conditions can be differentiated by comparing their clinical features and the effect of long-term macrolide treatment. PATIENTS AND METHODS: Fifty-eight Japanese patients, including 15 with HTLV-1-associated bronchiolitis and 43 with diffuse panbronchiolitis. Both conditions were clinically compared using the clinical criteria for diffuse panbronchiolitis, including findings from CT scans and BAL fluid testing. Pulmonary function, blood gas levels, and cold hemagglutinin (CHA) levels were assessed before and after long-term treatment with macrolides. Interleukin-2 receptor (IL-2R) expression in T cells obtained from the BAL fluid of patients with HTLV-1-associated bronchiolitis or diffuse panbronchiolitis was analyzed by flow cytometry. RESULTS: Clinical, laboratory, radiologic, and bacterial features were strikingly similar in both groups, except for the fact that patients with HTLV-1-associated bronchiolitis had a higher ratio of IL-2R-positive cells in the BAL fluid. The histopathologic features were also similar. Long-term treatment with macrolides improved PaO(2), FEV(1), and CHA in patients with HTLV-1-associated bronchiolitis to a lesser extent than in those with diffuse panbronchiolitis, and PaO(2) and FEV(1) in the group of patients with HTLV-1-associated bronchiolitis who had high IL-2R levels did not respond after therapy. CONCLUSIONS: These findings showed that the clinicopathologic features of the two conditions are quite similar, suggesting that diffuse panbronchiolitis is a chronic pulmonary manifestation of HTLV-1 infection. However, HTLV-1-associated bronchiolitis might be associated with conditions that are distinct from those of diffuse panbronchiolitis based on the different responses to macrolide treatment and the difference in the number of activated T cells bearing IL-2R in the lungs.

The effect of antiretroviral therapy on HTLV infection.

No effective treatment for TSP/HAM has been described so far. Interventions with corticosteroids, plasmapheresis, interferon and, more recently, with antiretroviral drugs have been tried with poor results. The main HTLV replication mechanism is thought to be through clonal expansion of HTLV-infected cells, which excludes the involvement of the reverse transcriptase (RT) enzyme. However, a virological and clinical improvement has been noticed in HTLV-I carriers suffering from TSP/HAM receiving zidovudine or lamivudine. Herein, we describe the virological and clinical outcome in two TSP/HAM patients infected with HTLV-I treated with zidovudine plus lamivudine, and in two HTLV-II/HIV-1 co-infected patients receiving triple combinations including lamivudine. While, one TSP/HAM patient experienced a 2 log decrease in HTLV-I proviral load, an increase of 1 log was observed in another patient after several months of treatment with zidovudine plus lamivudine. The two HTLV-II/HIV-1 co-infected patients showed an initial increase in HTLV-II proviral load after beginning HAART followed by a slight decline a few months later. Plasma HIV-1 RNA fell to <50 copies/ml in both patients after beginning therapy. None of the four HTLV positive patients developed genetic changes at the conserved YMDD domain within their respective RT genes, which could be related to lamivudine resistance. No clinical improvement was observed in one TSP/HAM patient after more than 1 year on treatment with nucleoside analogues. The inhibition of the HTLV RT along with the cytostatic effect of some nucleoside analogues, including zidovudine, could reduce HTLV replication, and therefore reduce HTLV proviral load. The clinical consequences of this effect need to be further examined.

Clinical features of human T-lymphotropic virus type 1 uveitis: a long-term follow-up.

To investigate the clinical manifestations of human T-lymphotropic virus type-1 uveitis (HU), 112 HU patients who were followed up periodically for more than one year were retrospectively analyzed with respect to their ophthalmological and systemic complications. The gender ratio (female/male ratio) of the HU patients was 2.0 and the initial complications were foggy vision in 34.5%, ocular floaters in 33.3%, and blurred vision in 15.5%. As for the ocular symptoms, the majority (78.6%) of patients were classified as intermediate uveitis with vitreous inflammation. Recurrence of uveitis episodes was seen in one half of the patients (51.8%); 12 patients had more than six uveitis episodes. The interval of uveitis episodes varied from two weeks to 10 years. Nearly one half of the patients (43.8%) had ocular complications: e.g., cataract in 22 patients, persistent vitreous opacities in 17 patients, and glaucoma in 16 patients. Although the visual prognosis was essentially good, 11 patients had poor visual prognosis (<0.1). The causes of poor vision in these patients were cataract, cystoid macular edema, epiretinal membrane, and optic nerve atrophy. Of the 112 HU patients, two developed HTLV-I-associated myelopathy (TSP/HAM) after the onset of HU, while none developed adult T-cell leukemia. Sixteen HU patients had a previous history of Graves' disease and a past history of methimazole therapy, while Graves' disease was found in another HU patient only after HU onset and methimazole was not administered before the onset of HU. The present data of long-term follow-up indicate that (1) HU causes various ocular complications and its visual prognosis can be poor, (2) TSP/HAM can be induced even after the onset of HU, and (3) methimazole is not a risk factor of HU after Graves' disease.

Toxicity of sulfonamide-reactive metabolites in HIV-infected, HTLV-infected, and noninfected cells.

It has been suggested that the high rates of adverse reactions to sulfonamides among patients with AIDS may be related to an increased sensitivity to reactive drug metabolites among HIV-infected cells. To study this hypothesis, we investigated the toxicity of the hydroxylamine of sulfamethoxazole in HIV-infected and noninfected MOLT-3 cultured human T-lymphoblasts. Toxicity was assessed by trypan blue dye exclusion. The hydroxylamine of sulfamethoxazole produced concentration-dependent toxicity in HIV-infected cells, with marked toxicity seen when HIV-infected cells were incubated with 400 microM of the hydroxylamine (82 +/- 8%); this was significantly greater than the toxicity seen among noninfected cells (p < 0.01). There was no concentration-dependent toxicity seen among noninfected cells or in cells infected with HTLV-I, suggesting that the concentration-dependent toxicity seen was specifically related to HIV infection. HIV-infected cells had significantly lower glutathione concentration than did noninfected cells (p < 0.05). Incubation with the hydroxylamine of sulfamethoxazole produced a concentration-dependent decline in glutathione content that was similar in infected and non-infected cells. Co-incubation with glutathione or N-acetylcysteine significantly reduced the toxicity of hydroxylamine of sulfamethoxazole in HIV-infected cells (p < 0.05). Our data supports the role of reactive sulfonamide metabolites in the pathogenesis of adverse reactions to sulfonamides among patients with AIDS.

Elevated serum levels of soluble Tac peptide in adult T-cell leukemia: correlation with clinical status during chemotherapy.

Adult T-cell leukemia associated with human T-cell leukemia virus type I (HTLV-I) is characterized by a clonal expansion of a CD4-positive subset of T lymphocytes that constitutively express high numbers of interleukin-2 receptors and that frequently infiltrate the skin; osteolytic bone lesions, and hypercalcemia. Using an enzyme-linked immunosorbent assay (ELISA) test, we measured the level of soluble Tac peptide, one chain of the human interleukin-2 receptor, in the serum of 50 patients with adult T-cell leukemia (38 Japanese and 12 American patients), 8 patients with other hematologic malignancies, 8 asymptomatic HTLV-I-antibody-positive carriers, and 17 normal controls. The serum level of soluble Tac peptide (geometric mean U/mL, 95% CI) was elevated at presentation in all patients with adult T-cell leukemia (16,461; 819 to 330,896) when compared with normal controls (238; 112 to 502), patients with other hematologic malignancies (1302; 475 to 3569), and healthy HTLV-I antibody-positive carriers (490; 115 to 2086). The highest levels were seen in patients (n = 33) with acute (32,154; 2587 to 399,598) compared with chronic (5464; 661 to 45,156) disease (n = 14). Serum levels of Tac peptide also tended to be more elevated in patients with adult T-cell leukemia with hypercalcemia (32,072; 2461 to 417,908) compared with normocalcemic patients (13,885; 496 to 388,436). Serial measurements of soluble Tac peptide levels in serum were done in four patients with adult T-cell leukemia during chemotherapy and the levels reflected disease activity. These observations suggest that the measurement of soluble Tac peptide levels in patients with adult T-cell leukemia is useful as a noninvasive measure of tumor burden and will help in the diagnosis of the disease and management of these patients.

Major prognostic factors of adult patients with advanced T-cell lymphoma/leukemia.

Eighty-one adult patients with advanced T-cell lymphoma/leukemia including 54 with adult T-cell leukemia/lymphoma (ATL), who were treated between 1981 and 1983 with vincristine, cyclophosphamide, prednisolone, and doxorubicin (VEPA) or VEPA plus methotrexate (VEPA-M) in randomized fashion, were evaluated for pretreatment characteristics. The overall complete response (CR) and the 4-year survival rates were 39.5% and 19.4%, respectively, and 69% of 32 CR patients had relapses, indicating the need for development of new effective regimens for the disease. In a multiple logistic regression analysis, only three factors, leukemic manifestation, poor performance status (PS), and a high lactate dehydrogenase (LDH) level, were significantly associated with the poor response rate. In a Cox proportional hazards model analysis, shortened survival was again significantly associated with poor PS and a high LDH level, but not with a clinical diagnosis of ATL. The two factors, PS and LDH level, that were found to be significantly associated with both CR and survival rates, were used to construct a model containing six categories of patients at increasing risk for poor response and shortened survival. These categories divided the patients into three groups with respective CR and 4-year survival rates of 75% and 53% for low-risk, 45% and 15% for moderate-risk, and 15% and 0% for high-risk. The results indicate that PS and LDH levels were the most important in predicting the response and survival of an adult patient with advanced T-cell lymphoma/leukemia. The prognosis of patients with usual peripheral T-cell lymphoma, excluding ATL, was comparable with that of advanced B-cell lymphoma. These results have important implications for the design of new prospective therapeutic trials.