HTLV-I is a member of the Deltaretroviruses, which include HTLV-II, bovine leukemia virus and simian T-cell leukemia virus (STLV). The latter two viruses also cause lymphoid malignancies in the host, similar to the case with HTLV-I. HTLV and STLV are thought to originate from common ancestors, and share molecular, virological and epidemiological features. Therefore, they have been designated primate T-cell leukemia viruses (PTLVs). Phylogenetical analyses have revealed that HTLV-Ic first diverged from simian leukemia virus around 50,000 ± 10,000 years ago, while the spread of PTLV-I in Africa is estimated to have occurred at least 27,300 ± 8,200 years ago. Subsequently, HTLV-Ia, which is the most common subtype in Japan, diverged from the African strain 12,300 ± 4,900 years ago 9. Thus, these viruses have had a long history with humans after the interspecies transmission. In contrast, human immunodeficiency virus type 1 (HIV-1) is thought to originate from simian immunodeficiency virus in chimpanzees (SIV_CPZ) 10, and the interspecies transmission to humans is estimated to have occurred recently.

There are approximately 10–20 million HTLV-I carriers in the world 11. In particular, HTLV-I is endemic in Japan, parts of central Africa, the Caribbean basin and South America. In addition, epidemiological studies of HTLV-I have revealed high seroprevalence rates in Melanesia, Papua New Guinea and the Solomon islands, as well as among Australian aborigines 12. In Japan, approximately 1.2 million individuals are estimated to be infected by HTLV-I, and more than 800 cases of ATL are diagnosed each year 13. Moreover, this virus also causes the neurodegenerative disease, HTLV-I-associated myelopathy/tropical spastic paraparesis (HAM/TSP) 1415. The cumulative risks of ATL among HTLV-I carriers in Japan are estimated to be about 6.6% for men and 2.1% for women, indicating that most HTLV-I carriers remain asymptomatic throughout their lives 16.

The HTLV-I provirus has a similar structure to other retroviruses: a long terminal repeat (LTR) at both ends and internal sequences such as the gag, pol and env genes. A characteristic of HTLV-I is the presence of the pX region, which exists between env and the 3'-LTR. This region encodes several accessory genes, which include the tax, rex, p12, p21, p30, p13 and HBZ genes. Among these, the tax gene plays central roles in viral gene transcription, viral replication and the proliferation of HTLV-I-infected cells. Tax enhances viral gene transcription from the 5'-LTR via interaction with cyclic AMP responsive element binding protein (CREB). Tax also interacts with cellular factors and activates transcriptional pathways, such as NF-κB, AP-1 and SRF 817181920. For example, activation of NF-κB induces the transcription of various cytokines and their receptor genes, as well as anti-apoptotic genes such as bcl-xL and survivin 212223. The activation of NF-κB has been demonstrated to be critical for tumorigenesis both in vitro and in vivo 2425. On the other hand, Tax variant without activation of NF-κB has also been reported to immortalize primary T-lymphocytes in vitro 26, suggesting that mechanisms of immortalization are complex. In addition to NF-κB, activation of other transcriptional pathways such as CREB by Tax should be implicated in the immortalization and leukemogenesis.

Tax also interferes with the functions of p53, p16 and MAD1 27282930. These interactions enable HTLV-I-infected cells to escape from apoptosis, and also induce genetic instability. Although inactivation of p53 function by Tax is reported to be mediated by p300/CBP 272831 or NF-κB activation 32, Tax can still repress p53's activity in spite of loss of p300/CBP binding or in cells lacking NF-κB activation 33, indicating the mechanism of p53 inactivation by Tax needs further investigation.

Although Tax promotes the proliferation of infected cells, it is also the major target of cytotoxic T-lymphocytes (CTLs) in vivo. Moreover, excess expression of Tax protein is considered to be harmful to infected cells. Therefore, HTLV-I has redundant mechanisms to suppress Tax expression. Rex binds to Rex-responsive element (RxRE) in the U3 and R regions of the 3'-LTR, and enhances the transport of the unspliced gag/pol and the singly spliced env transcripts. By this mechanism, double-spliced tax/rex mRNA decreases, resulting in suppressed expression of Tax 34. On the other hand, p30 binds to tax/rex transcripts, and retains them in the nucleus 35. The HBZ gene is encoded by the complementary strand of HTLV-I, and contains a leucine zipper domain. HBZ directly interacts with c-Jun or JunB 36, or enhances their degradation 37, resulting in the suppression of Tax-mediated viral transcription from the LTR.

Transforming growth factor-β (TGF-β) is an inhibitory cytokine that plays important roles in development, the immune system and oncogenesis. Since TGF-β generally suppresses the growth of tumor cells, most tumor cells acquire escape mechanisms that inhibit TGF-β signaling, including mutations in its receptor and in the Smad molecules that transduce the signal from the receptor. Tax has also been reported to inhibit TGF-β signaling by binding to Smad2, 3 and 4 or CBP/p300 3839. Inhibition of TGF-β signaling enables HTLV-I-infected cells to escape TGF-β-mediated growth inhibition.

ATL cells have been reported to show remarkable chromosomal abnormalities 40, which should be implicated in the disease progression. Tax has been reported to interact with the checkpoint protein MAD1, which forms a complex with MAD2 and controls the mitotic checkpoint. This functional hindrance of MAD1 by Tax protein causes chromosomal instability, suggesting the involvement of this mechanism in oncogenesis 30. Recently, Tax has been reported to interact with Cdc20 and activate Cdc20-associated anaphase-promoting complex, an E3 ubiquitin ligase that controls the metaphase-to-anaphase transition, thereby resulting in mitotic abnormalities 41.

In contrast to HTLV-I, HTLV-II promotes the proliferation of CD8-positive T-lymphocytes in vivo. Although it was first discovered in a patient with variant hairy cell leukemia, HTLV-II is less likely to have oncogenic properties since there is no obvious association between HTLV-II infections and cancers. Regardless of the homology of their tax sequences, the oncogenic potential of Tax1 (HTLV-I Tax) is more prominent than that of Tax2 (HTLV-II Tax). The most striking difference is that Tax2 lacks the binding motif at C-terminal end to PDZ domain proteins, while Tax 1 retains it 42. When the PDZ domain of Tax1 is added to Tax2, the latter acquires oncogenic properties in the rat fibroblast cell line Rat-1, indicating that this domain is responsible for the transforming activity of HTLV-I 43.

To understand the pleiotropic actions of Tax protein more clearly, transcriptome analyses are essential. The transcriptional changes induced by Tax expression have been studied using DNA microarrays, which revealed that Tax upregulated the expression of the mixed-lineage kinase MLK3. MLK3 is involved in NF-κB activation by Tax as well as NIK and MEKK1 44. In addition to transcriptional changes, Tax is also well known to interact with cellular proteins and impair or alter their functions. For example, proteomic analyses of Tax-associated complexes showed that Tax could interact with cellular proteins, including the active forms of small GTPases, such as Cdc42, RhoA and Rac1, which should be implicated in the migration, invasion and adhesion of T-cells, as well as in the activation of the JNK pathway 45.

As described, some ATL cells can proliferate without functional Tax protein, suggesting that somatic (genetic and epigenetic) alterations cause transcriptional or functional changes to the host genes. The p53 gene is frequently mutated in various cancers, and these mutations are associated with disease progression and a poor prognosis. The mutation rate of the p53 gene in ATL cells has been reported to be 36% (4/11) and 30% (3/10) 697071. The p16 gene is an inhibitor of cyclin-dependent kinase 4/6, and blocks the cell cycle. Genetic changes in this gene (deletion in most cases) have been described in many types of cancer cells. Deletion of the p16 gene has also been reported in ATL cells 72. Moreover, DNA methylation of the promoter region of the p16 gene is also implicated in the suppression of p16 73. In addition, genetic changes in the p27^KIP1, RB1/p105 and RB2/p130 genes have been reported in ATL, although they are relatively rare: 2/42 (4.8%) for the p27^KIP1 gene; 2/40 (5%) for the RB1/p105 gene; and 1/41 (2.4%) for the RB2/p130 gene) 74. The fact that higher frequencies of genetic changes in these tumor suppressor genes are observed among aggressive forms of ATL suggests that such genetic changes are implicated in disease progression.

Fas antigen was the first identified death receptor. It transduces the death signal by binding of its ligand, Fas ligand (FasL). ATL cells highly express Fas antigen on their cell surface 75, and are highly susceptible to death signals mediated by agonistic antibodies to Fas antigen, such as CH-11. Genetic changes of Fas gene in ATL cells, which confer resistance to the Fas-mediated signal, have been reported 7677. Normal activated T-lymphocytes express FasL as well as Fas antigen. Apoptosis induced by autocrine mechanisms is designated activation-induced cell death (AICD) and this controls the immune response 78. Although ATL cells express Fas antigen, they do not produce FasL, thereby enabling ATL cells to escape from AICD. Attempts to isolate hypermethylated genes from ATL cells identified the EGR3 gene as a hypermethylated gene compared to PBMCs from carriers 79. EGR3 is a transcriptional factor with a zinc finger domain, that is essential for transcription of the FasL gene 80. The finding that EGR3 gene transcription is silenced in ATL cells could account for the loss of FasL expression, and the escape of ATL cells from AICD. Thus, alterations of the Fas (genetic) and EGR3 (epigenetic) genes are examples of ATL cell evolution in vivo.

Disordered DNA methylation has been identified in the genome of ATL cells compared with that of PBMCs from carriers: hypomethylation is associated with aberrant expression of the MEL1S gene 81, while hypermethylation silences transcription of the p16 73, EGR3 and KLF4 genes as well as many others 79. It is reasonable to consider that other currently unidentified genes are involved in such alterations of the genome in ATL cells, and play roles in leukemogenesis.

Transcriptome analyses using DNA microarrays have revealed transcriptional changes that are specific to ATL cells. Among 192 up-regulated genes, the expressions of the tumor suppressor in lung cancer 1 (TSLC1), caveolin 1 and prostaglandin D2 synthase genes were increased more than 30-fold in fresh ATL cells compared with normal CD4+ and CD4+, CD45RO+ T-cells 82. TSLC1 is a cell adhesion molecule that acts as a tumor suppressor in lung cancer. Although TSLC1 is not expressed on normal T-lymphocytes, all acute ATL cells show ectopic TSLC1 expression. Enforced expression of TSLC1 enhances both the self-aggregation and adhesion abilities to vascular endothelial cells in ATL cells. Thus, TSLC1 expression is implicated in the adhesion or infiltration of ATL cells. By screening a retrovirus cDNA library from ATL cells, a gene with oncogenic potency was identified in NIH3T3 cells, and designated the Tgat gene 83. Ectopic expression of the Tgat gene is observed in aggressive forms of ATL, and in vitro experiments showed that its expression is associated with an invasive phenotype.

The host immune system, especially the cellular response, against HTLV-I exerts critical control over virus replication and the proliferation of infected cells 84. CTLs against the virus have been extensively studied, and Tax protein was found to be the dominant antigen recognized by CTLs in vivo 63. HTLV-I-specific CD8-positive CTLs are abundant and chronically activated. The paradox is that the frequency of Tax-specific CTLs is much higher in HAM/TSP patients than in carriers. Since the provirus load is higher in HAM/TSP patients, this finding suggests that the CTLs in HAM/TSP cannot control the number of infected cells. One explanation for this is that the CTLs in HAM/TSP patients show less efficient cytolytic activity toward infected cells, whereas CTLs in carriers can suppress the proliferation of infected cells 85. Hence, the gene expression profiles of circulating CD4+ and CD8+ lymphocytes were compared between carriers with high and low provirus loads. The results revealed that CD8+ lymphocytes from individuals with a low HTLV-1 provirus load show higher expressions of genes associated with cytolytic activities or antigen recognition than those from carriers with a high provirus load 86. Thus, CD8+ T-lymphocytes in individuals with a low provirus load successfully control the number of HTLV-I-infected cells due to their higher CTL activities. Thus, the major determinant of the provirus load is thought to be the CTL response to HTLV-I.

As mentioned above, the provirus load is considered to be controlled by host factors. Considering that the cellular immune responses are critically implicated in the control of HTLV-I infection, human leukocyte antigen (HLA) should be a candidate for such a host genetic factor. From analyses of HAM/TSP patients and asymptomatic carriers, HLA-A02, and Cw08 are independently associated with a lower provirus load and a lower risk of HAM/TSP. In addition, polymorphisms of other genes (TNF-α, SDF-1, HLA-B54, HLA-DRB-10101 and IL-15) are also associated with the provirus load, although their associations are not as significant compared with HLA-A02, and Cw08 8788. Regarding the onset of ATL, only a polymorphism of TNF-α gene was reported to show an association 89. However, familial clustering of ATL cases is a well-known phenomenon, strongly suggesting that genetic factors are implicated in the onset of ATL 909192.

Spontaneous remission is more frequently observed in patients with ATL than those with other hematological malignancies 9093. Usually, this phenomenon is associated with infectious diseases, suggesting that immune activation of the host enhances the immune response against ATL cells. If the immune response against HTLV-I is implicated in spontaneous remission, this suggests the possibility of immunotherapy for ATL patients by the induction of an immune response to HTLV-I 94, for example via antigen-stimulated dendritic cells.

Immunodeficiency in ATL patients is pronounced, and results in frequent opportunistic infections by various pathogens, including Pneumocystis carinii, cytomegalovirus, fungus, Strongyloides and bacteria, due to the inevitable impairment of the T-cell functions 95. To a lesser extent, impaired cell-mediated immunity has also been demonstrated in HTLV-I carriers 96. Such immunodeficiency in the carrier state may be associated with the leukemogenesis of ATL by allowing the proliferation of HTLV-I-infected cells. A prospective study of HTLV-I-infected individuals found that carriers who later develop ATL have a higher anti-HTLV-I antibody and a low anti-Tax antibody level for up to 10 years preceding their diagnosis. This finding indicates that HTLV-I carriers with a higher anti-HTLV-I titer, which is roughly correlated with the HTLV-I provirus load, and a lower anti-Tax reactivity may be at the greatest risk of developing ATL 97. The anti-HTLV-I antibody and soluble IL-2 receptor (sIL-2R) levels are correlated with the HTLV-I provirus load 53, and a high antibody titer and high sIL-2R level are risk factors for developing ATL among carriers 98. Taken together, these findings suggest that a higher proliferation of HTLV-I-infected cells and a low immune response against Tax may be associated with the onset of ATL. Given these findings, potentiation of CTLs against Tax via a vaccine strategy may be useful for preventing the onset of ATL 99.

EBV-associated lymphomas frequently develop in individuals with an immunodeficient state associated with transplantation or AIDS. This has also been reported in an ATL patient 100. Does such an immunodeficient state influence the onset of ATL? Among 24 patients with post-transplantation lymphoproliferative disorders (PT-LPDs) after renal transplantation in Japan, 5 cases of ATL have been reported. Considering that most PT-LPDs are of B-cell origin in Western countries, this frequency of ATL in Japan is quite high. Although the high HTLV-I seroprevalence is due to blood transfusion during hemodialysis, the immunodeficient state during renal transplantation apparently promotes the onset of ATL 101. In addition, when experimental allogeneic transplantation was performed to 12 rhesus monkeys and immunosuppressive agents (cyclosporine, prednisolone or lymphocyte-specific monoclonal antibodies) were administered to prevent rejection, 4 of the 7 monkeys that died during the experiment showed PT-LPDs. Importantly, the STLV provirus was detected in all PT-LPD samples 102. These observations emphasize that transplantation into HTLV-I-infected individuals or from HTLV-I positive donors require special attention.

Although the mechanism of immunodeficiency remains unknown, some previous reports have provided important clues. One mechanism for immunodeficiency is that HTLV-I infects CD8-positive T-lymphocytes, which may impair their functions 48. Indeed, the immune response against Tax via HTLV-I-infected CD8-positive T-cells renders these cells susceptible to fratricide mediated by autologous HTLV-I-specific CD8-positive T-lymphocytes 103. Fratricide among virus-specific CTLs could impair the immune control of HTLV-I. Another mechanism for immunodeficiency is based on the observation that the number of naive T-cells decreases in individuals infected with HTLV-I via decreased thymopoiesis 48. In addition, CD4+ and CD25+ T-lymphocytes are classified as immunoregulatory T-cells that control the host immune system. Regulatory T-cells suppress the immune reaction via the expression of immunoregulatory molecules on their surfaces. The FOXP3 gene has been identified as a master gene that controls gene expressions specific to regulatory T-cells. FOXP3 gene transcription can be detected in some ATL cases (10/17; 59%) 104. Such ATL cells are thought to suppress the immune response via expression of immunoregulatory molecules on their surfaces, and production of immunosuppressive cytokines.

ATL cells are derived from activated helper T-lymphocytes, which play central roles in the immune system by elaborating cytokines and expressing immunoregulatory molecules. ATL cells are known to retain such features, and this cytokine production or surface molecule expression may modify the pathogenesis.

ATL is well known to infiltrate various organs and tissues, such as the skin, lungs, liver, gastrointestinal tract, central nervous system and bone 95. This infiltrative tendency of leukemic cells is possibly attributable to the expressions of various surface molecules, such as chemokine receptors and adhesion molecules. Skin-homing memory T-cells uniformly express CCR4, and its ligands are thymus and activation-regulated chemokine (TARC) and macrophage-derived chemokine (MDC). CCR4 is expressed on most ATL cells. In addition, TARC and MDC are expressed in skin lesions in ATL patients. Thus, CCR4 expression should be implicated in the skin infiltration 105. On the other hand, CCR7 expression is associated with lymph node involvement 106. OX40 is a member of the tumor necrosis factor family, and was reported to be expressed on ATL cells 107. It was also identified as a gene associated with the adhesion of ATL cells to endothelial cells by a functional cloning system using a monoclonal antibody that inhibited the attachment of ATL cells 108. Thus, OX40 is also implicated in the cell adhesion and infiltration of ATL cells.

Hypercalcemia is frequently complicated in patients with acute ATL (more than 70% during the whole clinical course) 109. In hypercalcemic patients, the number of osteoclasts increases in the bone (Figure 3). RANK ligand, which is expressed on osteoblasts, and M-CSF act synergistically on hematopoietic precursor cells, and induce the differentiation into osteoclasts 110. ATL cells from hypercalcemic ATL patients express RANK ligand, and induced the differentiation of hematopoietic stem cells into osteoclasts when ATL cells were co-cultured with hematopoietic stem cells 111. In addition, the serum level of parathyroid hormone-related peptide (PTH-rP) is also elevated in most of hypercalcemic ATL patients. PTH-rP indirectly increases the number of osteoclasts, as well as activating them 112113, which is also implicated in mechanisms of hypercalcemia.

Regardless of intensive chemotherapies, the prognosis of ATL patients has not so improved. The median survival time of acute or lymphoma-type ATL was reported to be 13 months with the most intensive chemotherapy 114. Such a poor prognosis might be due to: 1) the resistance of ATL cells to anti-cancer drugs; and 2) the immunodeficient state and complicated opportunistic infections as described above. Regarding the resistance to anti-cancer drugs, one mechanism is the activated NF-κB pathway in ATL cells 115, which increases the transcription of anti-apoptotic genes such as bcl-xL and survivin. A proteasome inhibitor, bortezomib, is currently used for the treatment of multiple myeloma. One of its mechanisms is suppression of the NF-κB pathway by inhibiting the proteasomal degradation of IκB protein. Several groups have shown that bortezomib is effective against ATL cells both in vitro and in vivo 116117118119. Since the sensitivity to bortezomib is well correlated with the extent of NF-κB activation, the major mechanism of the anti-ATL effect is speculated to be inhibition of NF-κB. In addition, an NF-κB inhibitor has also been demonstrated to be effective against ATL cells 120.

During chemotherapy for ATL, chemotherapeutic agents worsen the immunodeficient state of ATL patients. In this regard, antibody therapy against ATL cells has advantages due to its decreased adverse effects. A humanized monoclonal antibody to CD25 has been clinically administered to patients with ATL 121122. In addition, a monoclonal antibody to CD2 is at the preclinical stage 123. As described above, most ATL cells express CCR4 antigen on their surfaces, and a humanized antibody against CCR4 is being developed as an anti-ATL agent 124.

Advances in the treatment of ATL were brought about by allogeneic bone marrow or stem cell transplantation 125126. Absence of graft-versus-host disease (GVHD) was linked with relapse of ATL, suggesting that GVHD or graft-versus-ATL may be implicated in the clinical effects of allogeneic stem cell transplantation 125. Furthermore, 16 patients with ATL, who were over 50 years of age, were treated with allogeneic stem cell transplantation with reduced conditioning intensity (RIST) from HLA-matched sibling donors 127. Among 9 patients in whom ATL relapsed after transplantation, 3 achieved a second complete remission after rapid discontinuation of cyclosporine A. This finding strongly suggests the presence of a graft-versus-ATL effect in these patients. In addition, Tax peptide-recognizing cells were detected by a tetramer assay (HLA-A2/Tax 11–19 or HLA-A24/Tax 301–309) in patients after allogeneic stem cell transplantation 128. In 8 patients, the provirus became undetectable by real-time PCR. Among these, 2 patients who received grafts from HTLV-I-positive donors also became provirus-negative by real-time PCR after RIST. Since the provirus load is relatively constant in HTLV-I-infected individuals 53, this finding indicates an enhanced immune response against HTLV-I after RIST, which suppresses the provirus load. This may account for the effectiveness of allogeneic stem cell transplantation to ATL. However, Tax expression is frequently lost in ATL cells as described above. Many questions arise, such as whether the tax gene status is correlated with the effect of allogeneic stem cell transplantation, and whether the effectiveness of the anti-HTLV-I immune response is against leukemic cells or non-leukemic HTLV-I-infected cells. Nevertheless, these data suggest that potentiation of the immune response against viral proteins such as Tax may be an attractive way to treat ATL patients 94. Such strategies may enable preventive treatment of high-risk HTLV-I carriers, such as those with familial ATL history, predisposing genetic factors to ATL, a higher provirus load, etc.

As described in the first section, HTLV-I has resided in humans for a long time. On the other hand, HIV-1 has only been recently transmitted to humans, probably from chimpanzees. Due to the comparatively small genomic differences between humans and chimpanzees, this virus can quickly adapt to human cells. These two human retroviruses are opposite in many aspects. HIV-1 vigorously replicates in vivo, and the maximum production of HIV-1 virions in the body can reach 10¹⁰ per day. Since reverse transcriptase is an error-prone enzyme due to its lack of proof-reading activity, it produces about one mistake per replication, resulting in tremendous errors in the proviral sequence during replication. Although most of these variations ruin the virus replication due to nonsense mutations or impairment of viral gene functions, some become capable of replicating under different circumstances such as the presence of anti-HIV drugs and activation of the host immune system. This can account for why HIV-1 acquires resistance against anti-HIV drugs, and escape from CTLs. On the other hand, HTLV-I increases its copy number in two ways, namely replication of HTLV-I itself and the proliferation of HTLV-I-infected cells in vivo. Although immune responses (antibodies, CTLs) against viral proteins suggest the presence of active viral replication in vivo, most of increased HTLV-I provirus load (the number of infected cells) is considered to be due to proliferation of infected cells since CTLs efficiently eliminate virus-expressing cells. Therefore, there is much less variation in the HTLV-I provirus sequence compared with HIV-1 129. However, this strategy by which HTLV-I increases the number of infected cells due to clonal expansion generates unfortunate side effects for both the host and the virus, namely oncogenesis of CD4-positive T-lymphocytes and the development of ATL.