Decreased PU.1 expression in mature B cells induces lymphomagenesis.

Diffuse large B-cell lymphoma (DLBCL) is the most common subtype of lymphoma, accounting for 30% of non-Hodgkin lymphomas. Although comprehensive analysis of genetic abnormalities has led to the classification of lymphomas, the exact mechanism of lymphomagenesis remains elusive. The Ets family transcription factor, PU.1, encoded by Spi1, is essential for the development of myeloid and lymphoid cells. Our previous research illustrated the tumor suppressor function of PU.1 in classical Hodgkin lymphoma and myeloma cells. In the current study, we found that patients with DLBCL exhibited notably reduced PU.1 expression in their lymphoma cells, particularly in the non-germinal center B-cell-like (GCB) subtype. This observation suggests that downregulation of PU.1 may be implicated in DLBCL tumor growth. To further assess PU.1's role in mature B cells in vivo, we generated conditional Spi1 knockout mice using Cγ1-Cre mice. Remarkably, 13 of the 23 knockout mice (56%) showed splenomegaly, lymphadenopathy, or masses, with some having histologically confirmed B-cell lymphomas. In contrast, no wild-type mice developed B-cell lymphoma. In addition, RNA-seq analysis of lymphoma cells from Cγ1-Cre Spi1F/F mice showed high frequency of each monoclonal CDR3 sequence, indicating that these lymphoma cells were monoclonal tumor cells. When these B lymphoma cells were transplanted into immunodeficient recipient mice, all mice died within 3 weeks. Lentiviral-transduced Spi1 rescued 60% of the recipient mice, suggesting that PU.1 has a tumor suppressor function in vivo. Collectively, PU.1 is a tumor suppressor in mature B cells, and decreased PU.1 results in mature B-cell lymphoma development.

Extranodal NK/T-cell lymphoma with localized relapse in bone marrow of lower leg detected using PET-CT.

Extranodal natural killer (NK)/T-cell lymphoma (ENKTL) is a rare subtype of non-Hodgkin lymphoma (NHL) with poor prognosis, particularly in relapsed or refractory patients. Thus, timely detection of relapse and appropriate disease management are crucial. We present two patients with ENKTL, wherein positron emission tomography-computed tomography (PET-CT) with total-body coverage after induction therapy, detected newly relapsed regions in the bone marrow of the lower leg prior to progression. Case 1: A 47-year-old woman with nasal obstruction, showing 18F-fluoro-deoxyglucose (FDG) uptake in the nasal cavity (Lugano stage IE). After induction therapy (RT-2/3 DeVIC), PET-CT revealed abnormal uptake only in the right fibula. Case 2: A 68-year-old man with a skin nodule/ulcer and an enlarged right inguinal lymph node was diagnosed with advanced ENKTL. A PET-CT scan revealed abnormal uptake in the subcutaneous mass of the right medial thigh, lymph nodes, and descending colon (Lugano stage IV). After induction therapy, PET-CT revealed new abnormal uptake only in the left tibia. In both patients, CT-guided biopsy confirmed ENKTL recurrence. Moreover, PET-CT with whole-body coverage was useful for the timely assessment of relapse and detection of asymptomatic bone involvement. This approach allowed for modifications to treatment strategies in certain patients.

Waldenstrom's macroglobulinemia-like B cell lymphoma with MYD88 L265P mutation and t(14;18)(q32;q21) involving IGH-MALT1.

A 65-year-old woman was referred to the hospital for further investigation of weight loss, hyperproteinemia, and anemia. Serum immunofixation electrophoresis revealed IgM-κ M protein. Bone marrow examination revealed an increase in the number of B -cells with immunoglobulin kappa light-chain restriction. Although the MYD88 L265P mutation was identified in bone marrow mononuclear cells, which suggested the diagnosis of Waldenstrom's macroglobulinemia (WM), a fusion signal of IgH-MALT1, which is commonly observed in extranodal marginal zone lymphoma of mucosa-associated lymphoid tissue (MALT) lymphoma, was also identified. Here, we describe a rare case of low-grade B-cell lymphoma with MYD88 L265P mutations accompanying IgH-MALT1.

[Follicular T-cell lymphoma successfully treated with long-term corticosteroid].

Follicular T-cell lymphoma (FTCL) is a rare disease, recently defined in the revised WHO classification Tumours of Haematopoietic and lymphoid tissues (4th edition). Although angioimmunoblastic T-cell lymphoma (AITL) and FTCL share similar T follicular helper (TFH) cell immunophenotypes and gene mutations, the clinical course of FTCL is not well characterized. Herein, we report the case of a 91-year-old woman with FTCL, who was successfully treated with corticosteroid. The patient, who had systemic lymphadenopathy and splenomegaly, was first diagnosed with necrotizing lymphadenitis. Re-biopsy was performed because of her persistent lymphadenopathy, which revealed FTCL. She was treated with corticosteroid because of her advanced age, poor performance, edema, and pleural effusion. After administering 100 mg prednisone, her condition improved and was discharged with prednisone tapering. Six-month positron emission tomography-computed tomography (PET-CT) scan showed complete metabolic remission. With a low dose of prednisone (6-10 mg), she remained disease-free for >3 years. Thus, these findings suggest that corticosteroid treatment is effective in some patients with peripheral T-cell lymphoma of TFH origin, including FTCL.

Predictive impact of soluble interleukin-2 receptor and number of extranodal sites for identification of patients at very high risk of CNS relapse in diffuse large B-cell lymphoma.

There remains an unmet clinical need to identify which patients with diffuse large B-cell lymphoma (DLBCL) would benefit from central nervous system (CNS) prophylaxis, due to the low positive predictive value (PPV; 10%-15%) of the currently available predictive models. To stratify patients at high risk of developing CNS relapse, we retrospectively analyzed 182 patients with DLBCL initially treated with rituximab, cyclophosphamide, doxorubicin, vincristine, and prednisone (R-CHOP), or a R-CHOP-like regimen. Among them, 17 patients relapsed with CNS involvement, and the 2-year rate of CNS relapse was 7.9%. Upon carrying out multivariate analysis, ≥3 extranodal sites and elevated soluble interleukin-2 receptor (sIL-2R) levels at diagnosis were identified as independent risk factors for CNS relapse. The 2-year and 3.5-year rates of CNS relapse were 57.1% and 78.6%, respectively, in patients with both elevated sIL-2R and ≥3 extranodal sites. Furthermore, combined use of these risk factors of both elevated sIL-2R and ≥3 extranodal sites resulted in a high PPV (71.4%), negative predictive value (93.1%), and overall accuracy (92.3%) for undergoing CNS relapse. In conclusion, we propose a simple and valuable tool to predict patients with DLBCL at very high risk of CNS relapse.

Beneficial impact of first-line mogamulizumab-containing chemotherapy in adult T-cell leukaemia-lymphoma.

Chemotherapy in combination with mogamulizumab (Mog) was approved in Japan in 2014 for untreated aggressive adult T-cell leukaemia-lymphoma (ATL), but the survival benefit remains unclear. Therefore, we retrospectively analysed clinical outcomes in 39 transplant-ineligible patients with untreated aggressive ATL at Kumamoto University Hospital between 2010 and 2021. The probability of four-year overall survival was 46.3% in the first-line Mog-containing treatment group compared to 20.6% in the chemotherapy-alone group (p = 0.033). Furthermore, this survival benefit was observed even in the elderly. In conclusion, first-line Mog-containing treatment can be a promising strategy for transplant-ineligible patients with ATL, especially in the elderly.

Two-year outcomes of tirabrutinib monotherapy in Waldenström's macroglobulinemia.

The phase II study of tirabrutinib monotherapy at a daily dose of 480 mg under fasting conditions for treatment-naïve and relapsed/refractory Waldenström's macroglobulinemia (ONO-4059-05 study) demonstrated a promising efficacy and tolerable safety profile. We conducted an unplanned analysis with a median follow-up of 24.8 months to update the efficacy and safety results and to report patient-reported quality of life. Of 27 enrolled patients, 22 patients continued receiving the study drug. The major response assessed by an independent review committee was observed in 25 patients (93%), including one and five patients who newly achieved complete response and very good partial response, respectively, after the primary analysis. The progression-free and overall survival rates at 24 months were 92.6% and 100%, respectively. Serum IgM levels in all patients except one declined and were maintained at low levels, although transient increases occurred after temporal interruption of the study drug. The disease-related symptoms including recurrent fever and hyperviscosity mostly disappeared. Health-related quality of life, assessed by cancer-specific questionnaires, was mostly maintained. Grade 3-4 neutropenia, lymphopenia, and leukopenia were newly recognized in three, two, and one patient, respectively. Grade 3 treatment-related hypertriglyceridemia was also recognized. Nine patients experienced grade 1-2 bleeding events (33%), one patient experienced grade 2 treatment-related atrial fibrillation, and one patient experienced grade 1 treatment-related hypertension. Treatment-related skin adverse events were observed in 14 patients (52%). Taken together, tirabrutinib has durable efficacy with an acceptable safety profile for treatment-naïve and refractory/relapsed Waldenström's macroglobulinemia.

[Adult T-cell leukemia/lymphoma diagnosed by RNA in situ hybridization for HTLV-1 bZIP factor].

A 62-year-old man visited the Department of Otorhinolaryngology at our hospital with a chief complaint of a pharyngeal mass. He was admitted to our department with a diagnosis of T-cell lymphoma based on a biopsy of a mesopharyngeal tumor. Although clonality analysis was not performed due to the lack of an appropriate sample, we considered the possibility of lymphoma-type (Lugano classification stage II) adult T-cell leukemia-lymphoma (ATL), as the anti-HTLV-1 antibody was positive. During the course of the disease, the peripheral blood smear revealed atypical lymphocytes with cleaved nuclei, and inverse PCR was performed with DNA extracted from those cells; however, the result showed that the pattern of HTLV-1 proviral DNA integration sites was polyclonal. Further, we performed RNA in situ hybridization targeting HTLV-1 bZIP factor (HBZ-ISH) using the formalin-fixed paraffin-embedded (FFPE) tissue samples of the mesopharyngeal tumor, and a high expression of HBZ was found in the tumor cells, leading to the diagnosis of ATL. These findings suggest the effectiveness of the novel diagnostic method using FFPE tissue samples for ATL.

Lymphoma during pregnancy in Japan: a multicenter retrospective cohort study.

OBJECTIVE: This study was conducted to characterize lymphoma occurring during pregnancy and to investigate the outcomes of the patients and the fetuses. METHODS: Clinical data were gathered retrospectively from 29 patients at 13 participating institutions, and data from 28 eligible patients were analyzed. RESULTS: Six (21%) patients had Hodgkin lymphoma (HL) and 22 (79%) patients had non-Hodgkin lymphoma (NHL). All patients with HL presented with lymphadenopathy, but 15 (68%) of the 22 patients with NHL presented with extranodal sites only. At the median follow-up period of 1325 (range 6-4461) days, the 5-year overall survival rate was 63% for patients with NHL and 100% for patients with HL. Three of the 13 patients who received chemotherapy during pregnancy (23%) developed Pneumocystis jiroveci pneumonia (PCP). There was 1 intrauterine fetal death, 1 spontaneous abortion in the first trimester, and 15 (54%) preterm births. CONCLUSION: This study showed a higher proportion of NHL than HL during pregnancy in Japan, which was inconsistent with the proportions observed in Western countries. The high incidence of maternal PCP and preterm birth suggested the need for improvements in our management of lymphoma during pregnancy.

Clinical significance of the immunoglobulin G heavy-chain repertoire in peripheral blood mononuclear cells of adult T-cell leukaemia-lymphoma patients receiving mogamulizumab.

'Monitoring of immune responses following mogamulizumab-containing treatment in patients with adult T-cell leukaemia-lymphoma (ATL)' (MIMOGA) is a multicentre prospective clinical study (UMIN000008696). In the MIMOGA study, we found that a lower percentage of CD2- CD19+ B cells in peripheral blood mononuclear cells (PBMC) was a significant unfavourable prognostic factor for overall survival (OS). Accordingly, we then analysed the immunoglobulin G (IgG) heavy-chain repertoire in PBMC by high-throughput sequencing. Of the 101 patients enrolled in the MIMOGA study, for 81 a sufficient amount of PBMC RNA was available for repertoire sequencing analysis. Peripheral IgG B cells in patients with ATL had a restricted repertoire relative to those in healthy individuals. There was a significant positive correlation between the Shannon-Weaver diversity index (SWDI) for the IgG repertoire and proportions of B cells in the PBMC of the patients. Multivariate analysis identified two variables significantly affecting OS: a higher serum soluble interleukin-2 receptor level, and a lower SWDI for the IgG repertoire [hazard ratio, 2·124; 95% confidence interval, 1·114-4·049; n = 44]. The present study documents the importance of humoral immune responses in patients receiving mogamulizumab-containing treatment. Further investigation of strategies to enhance humoral immune responses in patients with ATL is warranted.

Mogamulizumab for adult T-cell leukemia-lymphoma: a multicenter prospective observational study.

Monitoring of Immune Responses Following Mogamulizumab-Containing Treatment in Patients with Adult T-Cell Leukemia-Lymphoma (ATL) (MIMOGA) is a multicenter prospective observational study to establish the most effective and safe treatment strategy using mogamulizumab for ATL patients (UMIN000008696). Mogamulizumab-naive patients were enrolled (n = 102), of whom 101 received mogamulizumab-containing treatment (68 acute, 18 lymphoma, 12 chronic, and 3 smoldering subtypes). At enrollment, there was a significant inverse correlation between serum soluble interleukin-2 receptor (sIL-2R) levels and percentages of Tax-specific cytotoxic T lymphocytes (Tax-CTLs) in the entire lymphocyte population or in the CD8+ T cell subset, but there was not a correlation with cytomegalovirus pp65-specific cytotoxic T lymphocytes (CMV-CTLs). The overall response rate was 65%, and median progression-free survival and overall survival (OS) were 7.4 and 16.0 months, respectively. A higher percentage of Tax-CTLs, but not CMV-CTLs, within the entire lymphocyte population or in the CD8+ T cell subset was significantly associated with longer survival. Multivariate analysis identified the clinical subtype (acute or lymphoma type), a higher sIL-2R level, and a lower percentage of CD2-CD19+ B cells in peripheral blood mononuclear cells as significant independent unfavorable prognostic factors for OS. This indicates that a higher percentage of B cells might reflect some aspect of a favorable immune status leading to a good outcome with mogamulizumab treatment. In conclusion, the MIMOGA study has demonstrated that mogamulizumab exerts clinically meaningful antitumor activity in ATL. The patient's immunological status before mogamulizumab was significantly associated with treatment outcome. Further time series immunological analyses, in addition to comprehensive genomic analyses, are warranted.

A multicenter, open-label, phase II study of tirabrutinib (ONO/GS-4059) in patients with Waldenström's macroglobulinemia.

Tirabrutinib is a second-generation Bruton's tyrosine kinase inhibitor with greater selectivity than ibrutinib. Here, we conducted a multicenter, phase II study of tirabrutinib in patients with treatment-naïve (Cohort A) or with relapsed/refractory (Cohort B) Waldenström's macroglobulinemia (WM). Patients were treated with tirabrutinib 480 mg once daily. The primary endpoint was major response rate (MRR; ≥ partial response). Secondary endpoints included overall response rate (ORR; ≥ minor response), time to major response (TTMR), progression-free survival (PFS), overall survival (OS), and safety. In total, 27 patients (18 in Cohort A; 9 in Cohort B) were enrolled. The median age was 71 y, and the median serum immunoglobulin M level was 3600 mg/dL. Among the patients, 96.2% had the MYD88L265P mutation. MRR and ORR were 88.9% and 96.3%, respectively (Cohort A: MRR, 88.9%; ORR, 94.4%; Cohort B: MRR, 88.9%; ORR, 100%). Median TTMR was 1.87 mo. PFS and OS were not reached with a median follow-up of 6.5 and 8.3 mo for Cohorts A and B, respectively. The most common adverse events (AEs) were rash (44.4%), neutropenia (25.9%), and leukopenia (22.2%), with most AEs classified as grade 1 or 2. Grade ≥ 3 AEs included neutropenia (11.1%), lymphopenia (11.1%), and leukopenia (7.4%). No grade 5 AEs were noted. All bleeding events were grade 1; none were associated with drug-related atrial fibrillation or hypertension. Although the follow-up duration was relatively short, the study met the primary endpoint. Therefore, tirabrutinib monotherapy is considered to be highly effective for both untreated and relapsed/refractory WM with a manageable safety profile. (JapicCTI-173646).

Novel p97/VCP inhibitor induces endoplasmic reticulum stress and apoptosis in both bortezomib-sensitive and -resistant multiple myeloma cells.

p97/VCP is an endoplasmic reticulum (ER)-associated protein that belongs to the AAA (ATPases associated with diverse cellular activities) ATPase family. It has a variety of cellular functions including ER-associated protein degradation, autophagy, and aggresome formation. Recent studies have shown emerging roles of p97/VCP and its potential as a therapeutic target in several cancer subtypes including multiple myeloma (MM). We conducted a cell-based compound screen to exploit novel small compounds that have cytotoxic activity in myeloma cells. Among approximately 2000 compounds, OSSL_325096 showed relatively strong antiproliferative activity in MM cell lines (IC50 , 100-500 nmol/L). OSSL_325096 induced apoptosis in myeloma cell lines, including a bortezomib-resistant cell line and primary myeloma cells purified from patients. Accumulation of poly-ubiquitinated proteins, PERK, CHOP, and IREα, was observed in MM cell lines treated with OSSL_325096, suggesting that it induces ER stress in MM cells. OSSL_325096 has a similar chemical structure to DBeQ, a known p97/VCP inhibitor. Knockdown of the gene encoding p97/VCP induced apoptosis in myeloma cells, accompanied by accumulation of poly-ubiquitinated protein. IC50 of OSSL_325096 to myeloma cell lines were found to be lower (0.1-0.8 µmol/L) than those of DBeQ (2-5 µmol/L). In silico protein-drug-binding simulation suggested possible binding of OSSL_325096 to the ATP binding site in the D2 domain of p97/VCP. In cell-free ATPase assays, OSSL_325096 showed dose-dependent inhibition of p97/VCP ATPase activity. Finally, OSSL_325096 inhibited the growth of subcutaneous myeloma cell tumors in vivo. The present data suggest that OSSL_325096 exerts anti-myeloma activity, at least in part through p97/VCP inhibition.

Maintenance and enhancement of human peripheral blood mobilized stem/progenitor cell engraftment after ex vivo culture via an HDACi/SALL4 axis (3465).

Currently, there is a growing need for culturing hematopoietic stem/progenitor cells (HSPCs) in vitro for various clinical applications including gene therapy. Compared with cord blood (CB) CD34+ HSPCs, it is more challenging to maintain or expand CD34+ peripheral blood mobilized stem/progenitor cells (PBSCs) ex vivo. To fill this knowledge gap, we have systematically surveyed 466 small-molecule drug compounds for their potential in cytokine-dependent expansion of human CD34+CD90+ HSPCs. We found that epigenetic modifiers, especially histone deacetylase inhibitors (HDACis), could preferentially maintain and expand these cells. In particular, treatment of CD34+ PBSCs with a single dose of HDACi trichostatin A (TSA) at a concentration of 50 nmol/L ex vivo yielded the greatest expansion (11.7-fold) of CD34+CD90+ cells when compared with the control (dimethyl sulfoxide [DMSO] plus cytokines) group. Additionally, TSA-treated PBSC CD34+ cells had a statistically significant higher engraftment rate than the control-treated group in xenotransplantation experiments. Mechanistically, TSA treatment was associated with increased expression of HSPC-related genes such as GATA2 and SALL4. Furthermore, TSA-mediated CD34+CD90+ expansion was reduced by downregulation of SALL4 but not GATA2. Overall, we have developed a robust, short-term (5-day), PBSC ex vivo maintenance/expansion culture technique and found that the HDACi-TSA/SALL4 axis is important for the biological process.