A Comprehensive Study of the Immunophenotype and its Clinicopathologic Significance in Adult T-Cell Leukemia/Lymphoma.

Adult T-cell leukemia/lymphoma (ATLL) is a mature T-cell tumor caused by human T-lymphotropic virus type 1 (HTLV-1). The typical ATLL immunophenotypes are described in the 2017 World Health Organization Classification of Tumours of Haematopoietic and Lymphoid Tissues (positive: CD2, CD3, CD5, CD4, and CD25; negative: CD7, CD8, and cytotoxic markers; and partially positive: CD30, CCR4, and FOXP3). However, limited studies are available on the expression of these markers, and their mutual relationship remains unknown. Furthermore, the expression status of novel markers associated with T-cell lymphomas, including Th1 markers (T-bet and CXCR3), Th2 markers (GATA3 and CCR4), T follicular helper markers (BCL6, PD1, and ICOS), and T-cell receptor (TCR) markers, and their clinicopathologic significance is unclear. In this study, we performed >20 immunohistochemical stains in 117 ATLL cases to determine the comprehensive immunophenotypic profile of ATLL, which were compared on the basis of clinicopathologic factors, including morphologic variants (pleomorphic vs anaplastic), biopsy locations, treatments, Shimoyama classification-based clinical subtype, and overall survival. CD3+/CD4+/CD25+/CCR4+ was considered a typical immunophenotype of ATLL, but approximately 20% of cases did not conform to this pattern. Simultaneously, the following new findings were obtained: (1) most cases were negative for TCR-ß and TCR-δ (104 cases, 88.9%), indicating the usefulness of negative conversion of TCR expression to provide differentiation from other T-cell tumors; (2) the positivity of CD30 and CD15 and the negativity of FOXP3 and CD3 were significantly associated with anaplastic morphology; and (3) atypical cases, such as T follicular helper marker-positive (12 cases, 10.3%) and cytotoxic molecule-positive cases (3 cases, 2.6%), were identified. No single markers could predict the overall survival among patients with acute/lymphoma subtypes of ATLL. The results of this study illustrate the diversity of ATLL phenotypes. In T-cell tumors occurring in HTLV-1 carriers, the possibility of ATLL should not be eliminated even when the tumor exhibits an atypical phenotype, and the confirmation of HTLV-1 in the tissue is recommended.

Genetic Alterations in Adult T-Cell Leukemia/Lymphoma: Novel Discoveries with Clinical and Biological Significance.

Adult T-cell leukemia/lymphoma (ATLL) is a refractory T-cell neoplasm that develops in human T-cell leukemia virus type-I (HTLV-1) carriers. Large-scale comprehensive genomic analyses have uncovered the landscape of genomic alterations of ATLL and have identified several altered genes related to prognosis. The genetic alterations in ATLL are extremely enriched in the T-cell receptor/nuclear factor-κB pathway, suggesting a pivotal role of deregulation in this pathway in the transformation of HTLV-1-infected cells. Recent studies have revealed the process of transformation of HTLV-1-infected cells by analyzing longitudinal samples from HTLV-1 carriers and patients with overt ATLL, an endeavor that might enable earlier ATLL diagnosis. The latest whole-genome sequencing study discovered 11 novel alterations, including CIC long isoform, which had been overlooked in previous studies employing exome sequencing. Our study group performed the targeted sequencing of ATLL in Okinawa, the southernmost island in Japan and an endemic area of HTLV-1, where the comprehensive genetic alterations had never been analyzed. We found associations of genetic alterations with HTLV-1 strains phylogenetically classified based on the tax gene, an etiological virus factor in ATLL. This review summarizes the genetic alterations in ATLL, with a focus on their clinical significance, geographical heterogeneity, and association with HTLV-1 strains.

Spindle cell tumor with histiocytic and myogenic marker expression in the lymph node of a human T-cell leukemia virus type 1 carrier.

Carriers of oncogenic human T-cell leukemia virus type 1 (HTLV-1) can develop adult T-cell leukemia/lymphoma (ATLL). While an increasing number of animal models of HTLV-1 infection have revealed that malignant tumors with a histiocytic phenotype can arise, they have not been reported in humans. Here, we present a 79-year-old female HTLV-1 carrier who presented with a swollen lymph node. Histological examination revealed that the lymph node was replaced with a malignant spindle cell tumor, but not ATLL. Immunohistochemical analysis indicated that the tumor was positive for histiocytic (CD68 and CD163) and myogenic (α-smooth muscle actin, desmin, and caldesmon) markers, suggesting some differential diagnoses. We could not reach a definitive diagnosis under the current notion of the disease entity. In addition, we could not provide an exact causal relationship between HTLV-1 infection and the development of the current tumor. Nevertheless, this tumor may be a human counterpart of murine HTLV-1-related histiocytic tumors. Curiously, the tumor showed a good response to chemotherapy with the combination of cyclophosphamide, vincristine, and prednisone, a standard approach for ATLL. This case might represent a novel entity of an HTLV-1-related malignant tumor. Further accumulation of case reports will certainly contribute to our understanding of human HTLV-1-related disease and the mechanism of viral oncogenesis.

Acute type adult T-cell leukemia cells proliferate in the lymph nodes rather than in peripheral blood.

A massive increase in the number of mature CD4+ T-cells in peripheral blood (PB) is a defining characteristic of acute type of adult T-cell leukemia (ATL). To date, the site of proliferation of ATL cells in the body has been unclear. In an attempt to address this question, we examined the expression of the proliferation marker, Ki-67, in freshly isolated ATL cells from PB and lymph nodes (LNs) of patients with various types of ATL. Our findings reveal that LN-ATL cells display higher expression of the Ki-67 antigen than PB-ATL cells in acute type patients. The gene expression of T-cell quiescence regulators such as Krüppel-like factor 2/6 and forkhead box protein 1 was substantially high in acute type PB-ATL cells. The expression of human telomerase reverse transcriptase, which is involved in T-cell expansion, was significantly low in PB-ATL cells from acute type patients, similar to that in normal resting T-cells. These findings suggest that ATL cells proliferate in the LNs rather than in PB.

The Positivity of Phosphorylated STAT3 Is a Novel Marker for Favorable Prognosis in Germinal Center B-Cell Type of Diffuse Large B-Cell Lymphoma.

On the basis of immunohistochemistry, diffuse large B-cell lymphoma (DLBCL) is categorized as a germinal center B-cell (GCB) or non-GCB subtype. Recent integrated genomic analyses have highlighted the importance of the JAK-STAT3 pathway in the molecular pathogenesis of DLBCL. However, its relevance to clinical outcomes remains controversial. Therefore, we evaluated the extent of the nuclear expression of phosphorylated STAT3 (pSTAT3), a surrogate marker of signal transducer and activator of transcription 3 (STAT3) activation, by immunohistochemistry. We also analyzed the potential relationship between pSTAT3 positivity (defined as ≥40% positive neoplastic cells) and clinicopathologic characteristics in 294 patients with DLBCL. pSTAT3 was detected in 122 patients (42%), with a higher rate in the non-GCB subtype than in the GCB subtype (57% vs. 28%, P<0.001). Factors potentially activating STAT3, MYD88L265P, and Epstein-Barr virus-encoded small RNA were identified in the pSTAT3-positive non-GCB subtype, whereas the pSTAT3-positive GCB subtype often showed STAT3 mutations and lacked EZH2 mutations and the rearrangements of BCL2 and MYC. Multivariate analyses revealed that the pSTAT3-positive GCB subtype showed a favorable prognosis (HR: 0.17; 95% confidence interval, 0.04-0.7; P=0.014). These findings suggest that pSTAT3 positivity may have a unique impact on the clinicopathologic characteristics of DLBCL, making it a promising novel marker for the favorable prognosis of patients with the GCB subtype.

Clinicopathological features of adult T-cell leukemia/lymphoma with HTLV-1-infected Hodgkin and Reed-Sternberg-like cells.

Hodgkin and Reed-Sternberg (HRS) cells, a hallmark of classic Hodgkin lymphoma (CHL), are occasionally detected in non-Hodgkin lymphomas, including adult T-cell leukemia/lymphoma (ATLL), a lymphoid neoplasm caused by human T-cell leukemia virus type 1 (HTLV-1). HRS-like cells associated with ATLL have been described to be of B-cell lineage and infected with Epstein-Barr virus (EBV), not HTLV-1. We herein describe clinicopathological findings in 8 cases (4 males and 4 females; median age, 73 years [range, 55-81 years]) of ATLL with HTLV-1-infected HRS-like cells identified by ultrasensitive RNA in situ hybridization for HTLV-1 basic leucine zipper factor (HBZ-ISH), a specific viral transcript of HTLV-1. All patients showed nodal or mediastinal lesions, and 5 of the 8 patients were at an advanced disease stage. HRS-like cells were positive for CD30, CD15, MUM1, CD25, and HBZ-ISH and negative for B-cell markers, including PAX5, pan-T-cell antigens, and EBV in all cases. Five cases were positive for CD4, and 6 cases were positive for fascin. HBZ was identified in both HRS-like cells and surrounding lymphoid cells in 1 case with an aggressive clinical course and only HRS-like cells in 7 cases, most of whom showed a clinical response regardless of the chemotherapeutic regimen. Even though the definitive lineage typing of the HTLV-1-infected HRS cells is one of the limitations of this study in the absence of single-cell microdissection for polymerase chain reaction analysis, the combination of diffuse HBZ-ISH positivity and negativity for PAX5 and EBV deemed these cases distinct from CHL arising in HTLV-1 carriers.

Genetic profile of adult T-cell leukemia/lymphoma in Okinawa: Association with prognosis, ethnicity, and HTLV-1 strains.

Genetic alterations in adult T-cell leukemia/lymphoma (ATLL), a T-cell malignancy associated with HTLV-1, and their clinical impacts, especially from the perspective of viral strains, are not fully elucidated. We employed targeted next-generation sequencing and single nucleotide polymorphism array for 89 patients with ATLL in Okinawa, the southernmost islands in Japan, where the frequency of HTLV-1 tax subgroup-A (HTLV-1-taxA) is notably higher than that in mainland Japan, where most ATLL cases have HTLV-1-taxB, and compared the results with previously reported genomic landscapes of ATLL in mainland Japan and the USA. Okinawan patients exhibited similar mutation profiles to mainland Japanese patients, with frequent alterations in TCR/NF-ĸB (eg, PRKCB, PLCG1, and CARD11) and T-cell trafficking pathways (CCR4 and CCR7), in contrast with North American patients who exhibited a predominance of epigenome-associated gene mutations. Some mutations, especially GATA3 and RHOA, were detected more frequently in Okinawan patients than in mainland Japanese patients. Compared to HTLV-1-taxB, HTLV-1-taxA was significantly dominant in Okinawan patients with these mutations (GATA3, 34.1% vs 14.6%, P = .044; RHOA, 24.4% vs 6.3%, P = .032), suggesting the contribution of viral strains to these mutation frequencies. From a clinical viewpoint, we identified a significant negative impact of biallelic inactivation of PRDM1 (P = .027) in addition to the previously reported PRKCB mutations, indicating the importance of integrated genetic analysis. This study suggests that heterogeneous genetic abnormalities in ATLL depend on the viral strain as well as on the ethnic background. This warrants the need to develop therapeutic interventions considering regional characteristics.

A new diagnostic algorithm using biopsy specimens in adult T-cell leukemia/lymphoma: combination of RNA in situ hybridization and quantitative PCR for HTLV-1.

Histopathological distinction between adult T-cell leukemia/lymphoma (ATLL) and other T-cell neoplasms is often challenging. The current gold standard for the accurate diagnosis of ATLL is the Southern blot hybridization (SBH) assay, which detects clonal integration of human T-cell leukemia virus type I (HTLV-1) provirus. However, SBH cannot be performed with small biopsy or formalin-fixed paraffin-embedded (FFPE) tissue samples because this assay requires a large amount of DNA without degradation. Here we developed a new diagnostic algorithm for the accurate diagnosis of ATLL using FFPE samples. This method combines two HTLV-1 detection assays, namely, ultrasensitive RNA in situ hybridization using RNAscope for HTLV-1 bZIP factor (HBZ-RNAscope), and quantitative PCR targeting the tax gene (tax-qPCR). We analyzed 119 FFPE tissue specimens (62 ATLL, and 57 non-ATLL, including 41 HTLV-1 carriers) and compared them with the SBH results using the corresponding fresh-frozen samples. As a result, tax-qPCR had a higher ATLL identification rate than HBZ-RNAscope (88% [52/59], and 63% [39/62], respectively). However, HBZ-RNAscope clearly visualized the localization of HTLV-1-infected tumor cells and its identification rate increased to 94% (17/18) when the analysis was limited to samples up to 2 years old, indicating its usefulness in the daily diagnosis. The diagnostic algorithm combining these two assays successfully evaluated 94% (112/119) of samples and distinguished ATLL from non-ATLL cases including HTLV-1 carriers with 100% sensitivity and specificity. This method is expected to replace SBH and increase the accuracy of the diagnosis of ATLL.

Dermatopathic reaction of lymph nodes in HTLV-1 carriers: a spectrum of reactive and neoplastic lesions.

AIMS: Dermatopathic reaction is a histopathological finding of lymph nodes that usually occurs in patients with inflammatory pruritic cutaneous lesions. However, it is sometimes seen in patients with cutaneous T cell lymphoma. Adult T cell leukaemia/lymphoma (ATLL) is a T cell malignancy caused by infection with human T cell leukaemia virus type I (HTLV-1), which is frequently accompanied by cutaneous lesions. However, the detailed clinicopathological characteristics of the dermatopathic reaction of lymph nodes in ATLL patients and HTLV-1 carriers, addressed in this study, remains to be clarified. METHODS AND RESULTS: We retrospectively analysed 18 nodal lesions with dermatopathic reaction in HTLV-1 carriers. Axillary and inguinal lymph nodes were the primary affected tissues. Three cases with atypical lymphoid cell infiltration were defined as ATLL with dermatopathic reaction (ATLL-D), showing an abnormal T cell immunophenotype and T cell monoclonality. Two of the three ATLL-D patients died 14 and 7 months after diagnosis (the third case had a very short follow-up). The other 15 patients were indistinguishable from reactive lesions and were defined as HTLV-1-associated lymphadenitis with dermatopathic reaction (HAL-D). They showed an indolent clinical course, with only one case eventually transforming to aggressive disease. CONCLUSIONS: Lymph node lesions accompanied by dermatopathic reaction in HTLV1 carriers represent a spectrum that includes reactive and neoplastic conditions. HAL-D should be distinguished from ATLL-D, especially to avoid overtreatment.

Proteomic profiling of HTLV-1 carriers and ATL patients reveals sTNFR2 as a novel diagnostic biomarker for acute ATL.

Adult T-cell leukemia/lymphoma (ATL) is a human T-cell leukemia virus type 1 (HTLV-1)-associated T-cell malignancy with generally poor prognosis. Although only ∼5% of HTLV-1 carriers progress to ATL, early diagnosis is challenging because of the lack of ATL biomarkers. In this study, we analyzed blood plasma profiles of asymptomatic HTLV-1 carriers (ACs); untreated ATL patients, including acute, lymphoma, smoldering, and chronic types; and ATL patients in remission. Through SOMAscan, expression levels of 1305 plasma proteins were analyzed in 85 samples (AC, n = 40; ATL, n = 40; remission, n = 5). Using gene set enrichment analysis and gene ontology, overrepresented pathways in ATL vs AC included angiogenesis, inflammation by cytokines and chemokines, interleukin-6 (IL-6)/JAK/STAT3, and notch signaling. In selecting candidate biomarkers, we focused on soluble tumor necrosis factor 2 (sTNFR2) because of its active role in enriched pathways, extreme significance (Welch's t test P < .00001), high discrimination capacity (area under the curve >0.90), and novelty in ATL research. Quantification of sTNFR2 in 102 plasma samples (AC, n = 30; ATL, n = 68; remission, n = 4) using enzyme-linked immunosorbent assay showed remarkable elevations in acute ATL, at least 10 times those of AC samples, and return of sTNFR2 to AC state levels after achieving remission. Flow cytometry and immunostaining validated the expression of TNFR2 in ATL cells. No correlation between sIL-2 and sTNFR2 levels in acute ATL was found, suggesting the possibility of sTNFR2 as an independent biomarker. Our findings represent the first extensive blood-based proteomic analysis of ATL, suggesting the potential clinical utility of sTNFR2 in diagnosing acute ATL.

Intestinal helminth infections in HIV-infected patients in Savannakhet after establishment of an HIV registration network in Lao People's Democratic Republic.

BACKGROUND: In Lao People's Democratic Republic (PDR), which borders China, Vietnam, Cambodia, Thailand, and Myanmar, the number of HIV-infected patients has increased in recent years. HIV-infected patients diagnosed in Lao PDR are enrolled in a registration network and receive antiretroviral therapy (ART) covered by governmental financial support. Based on the registration network, we investigated intestinal helminth infections and coinfection with HTLV-1 in HIV-infected patients treated with an early intervention using ART in Lao PDR. METHODS: This cross-sectional study of all 252 HIV-infected patients at Savannakhet Provincial Hospital, located in the southern part of Lao PDR, was conducted between February and March 2018. Socioepidemiological information and clinical information were collected from a registration network database and by questionnaire administered to participants. Microscopic examination of intestinal helminth infections in stool samples and particle agglutination for anti-HTLV-1 antibody in plasma were performed. RESULTS: The median age of all 252 participants was 39 years old (range, 18-59). Based on the registration network database, there were 156 (61.9%) HIV-infected patients with a CD4-positive cell count ≥ 200 cells/µL and 146 (57.9%) with an HIV viral load < 250 copies/mL. Among 212 stool samples, 75 (35.4%) were found to contain one or more intestinal helminth species, including Opisthorchis viverrini (16.5%), Strongyloides stercoralis (10.8%), hookworm (10.4%), and Taenia saginata (3.3%). This rate of intestinal helminth infections was lower than that of a previous report conducted before the establishment of the registration network for HIV-infected patients in Lao PDR. There was no significant association between intestinal helminth infections and a lower CD4-positive T cell count or higher HIV viral load. HIV-infected patients with anti-HTLV-1 antibody positivity were not found in this cohort. CONCLUSION: The registration network and an early intervention using ART may provide good medical care and improve the clinical course of HIV-infected patients in Lao PDR. However, the incidence of intestinal helminth infections remains high at 35.4%. The development of a specific medical care system for helminth infection for HIV-infected patients is necessary.

Human T-cell leukemia virus type I Tax genotype analysis in Okinawa, the southernmost and remotest islands of Japan: Different distributions compared with mainland Japan and the potential value for the prognosis of aggressive adult T-cell leukemia/lymphoma.

Okinawa, comprising remote islands off the mainland of Japan, is an endemic area of human T-cell leukemia virus type I (HTLV-1), the causative virus of adult T-cell leukemia-lymphoma (ATL) and HTLV-1-associated myelopathy (HAM). We investigated the tax genotype of HTLV-1 among 29 HTLV-1 carriers, 74 ATL patients, and 33 HAM patients in Okinawa. The genotype distribution-60 (44%) taxA cases and 76 (56%) taxB cases-differed from that of a previous report from Kagoshima Prefecture in mainland Japan (taxA, 10%; taxB, 90%). A comparison of the clinical outcomes of 45 patients (taxA, 14; taxB, 31) with aggressive ATL revealed that the overall response and 1-year overall survival rates for taxA (50% and 35%, respectively) were lower than those for taxB (71% and 49%, respectively). In a multivariate analysis of two prognostic indices for aggressive ATL, Japan Clinical Oncology Group-Prognostic Index and Prognostic Index for acute and lymphoma ATL, with respect to age, performance status, corrected calcium, soluble interleukin-2 receptor, and tax genotype, the estimated hazard ratio of taxA compared with taxB was 2.68 (95% confidence interval, 0.87-8.25; P=0.086). Our results suggest that the tax genotype has clinical value as a prognostic factor for aggressive ATL.