Ropeginterferon alfa-2b, a monopegylated interferon α-2b, is a cytoreductive treatment for polycythemia vera (PV). In Japan, the current regimen involves titration in 50-µg increments every 2 weeks until reaching the maximum dose (500 µg), which requires considerable time. This phase 3b, open-label, single-arm, multicenter study (October 2023–July 2024) assessed the efficacy and safety of a three-step dose escalation regimen (day 1: 250 µg, week 2: 350 µg, week 4: 500 µg) of ropeginterferon alfa-2b in Japanese patients with PV (NCT06002490). Twenty-one patients were included (mean age: 57 years); most (95.2

Early achievement of deep remission improves patients' outcome in chronic myeloid leukemia (CML) treatment, highlighting the need for predictive indicators before therapy initiation. This study aimed to investigate whether baseline hematologic profiles are associated with molecular response in CML. Using hierarchical clustering of complete blood count (CBC) data at diagnosis, patients were stratified into two clusters. Patients in Cluster 1 had higher BCR::ABL1IS mRNA levels at 3 and 6 months post-treatment and lower rates of major molecular response compared with cluster 2. Cluster 1 also showed increased granulocyte and immature white blood cell counts and decreased erythroid parameters. Flow cytometric analysis of bone marrow mononuclear cells revealed that cluster 1 had a significant increase in hematopoietic stem cell fractions and a higher ratio of granulocyte-macrophage progenitors to megakaryocyte-erythroid progenitors compared with cluster 2. These findings suggest that differences in bone marrow progenitor cell differentiation affect peripheral blood profiles. Artificial intelligence-driven ghost cytometry (GC) comprehensively captured these differences, distinguishing poor responders, with diagnostic GC scores correlating with BCR::ABL1IS mRNA levels. The study indicates that multivariate analysis of CBC and GC captures biological features associated with early molecular response in CML.

Early achievement of deep remission improves patients' outcome in chronic myeloid leukemia (CML) treatment, highlighting the need for predictive indicators before therapy initiation. This study aimed to develop a tool to predict CML treatment responses to guide optimal therapy selection. Using hierarchical clustering of complete blood count (CBC) data at diagnosis, patients were stratified into two clusters. Patients in Cluster 1 had higher BCR::ABL1 IS mRNA levels at 3 and 6 months post-treatment and lower rates of major molecular response compared to cluster 2. Cluster 1 also showed increased granulocyte and immature white blood cell counts and decreased erythroid parameters. Flow cytometric analysis of bone marrow mononuclear cells revealed that cluster 1 had a significant increase in hematopoietic stem cell fractions and a higher ratio of granulocyte-macrophage progenitors to megakaryocyte-erythroid progenitors compared to cluster 2. These findings suggest that differences in bone marrow progenitor cell differentiation affect peripheral blood profiles. Artificial intelligence-driven ghost cytometry (GC) was evaluated for its ability to comprehensively capture these changes and successfully distinguished patients with poorer treatment responses, with GC scores at diagnosis strongly correlating with BCR::ABL1 IS mRNA levels at 3 and 6 months post-treatment initiation. The study indicates that multivariate analysis of CBC or GC analysis may enable simple, early prediction of CML treatment efficacy, potentially contributing to effective and individualized CML therapy. ### Competing Interest Statement This work was funded by Sysmex Corporation to T.T. as a collaborative research project. K.S. and T.T. have filed patent applications related to the label-free CML cell detection method. K.S. and K.Y. are employees of Sysmex Corporation. Other authors declare no competing financial interests regarding this study. ### Funding Statement This work was funded by Sysmex Corporation to T.T. as a collaborative research project. ### Author Declarations I confirm all relevant ethical guidelines have been followed, and any necessary IRB and/or ethics committee approvals have been obtained. Yes The details of the IRB/oversight body that provided approval or exemption for the research described are given below: This study was approved by the Ethics Committee of Juntendo University School of Medicine and Sysmex Corporation and adhered to the Declaration of Helsinki. Patients provided informed consent by accessing the study information published on the website and were free to withdraw their consent at any point during the study, as per the Ethical Guidelines for Medical and Health Research Involving Human Subjects. The ethics committee waived the requirement for written informed consent. I confirm that all necessary patient/participant consent has been obtained and the appropriate institutional forms have been archived, and that any patient/participant/sample identifiers included were not known to anyone (e.g., hospital staff, patients or participants themselves) outside the research group so cannot be used to identify individuals. Yes I understand that all clinical trials and any other prospective interventional studies must be registered with an ICMJE-approved registry, such as ClinicalTrials.gov. I confirm that any such study reported in the manuscript has been registered and the trial registration ID is provided (note: if posting a prospective study registered retrospectively, please provide a statement in the trial ID field explaining why the study was not registered in advance). Yes I have followed all appropriate research reporting guidelines, such as any relevant EQUATOR Network research reporting checklist(s) and other pertinent material, if applicable. Yes The datasets generated and/or analyzed during the current study are available from the corresponding author upon reasonable request.

Early diagnosis and treatment initiation of chronic myeloid leukemia (CML) are considered to increase the rate of deep molecular response. However, the early diagnosis of CML is challenging due to the absence of clinical symptoms and peripheral blood test anomaly, especially at the timing of peripheral white blood cell count is within a normal range. This study explored the possibility of artificial intelligence (AI)-based quantitative detection of CML cells using ghost cytometry (GC) technology. We created pre-trained AI models, using the morphological information data of the peripheral blood leukocytes obtained from patients newly diagnosed with CML and healthy individuals. We applied these models to peripheral blood samples from CML patients after initiating tyrosine kinase inhibitor treatment at various time points, which contains smaller amounts of tumor cells. The AI model accurately detected CML cells and a strong correlation between AI-detected CML cells and actual BCR::ABL1IS mRNA levels was observed. We concluded that the multidimensional morphological information of single cells obtained using GC, combined with machine learning algorithms, enables the quantitative detection of label-free CML cells. Our finding may enable the development of a screening test that can identify early-stage patients with CML before numerical abnormalities appear in blood tests.

Polycythemia vera (PV) is a Philadelphia chromosome-negative myeloproliferative neoplasm driven by JAK2 mutations, leading to the overproduction of blood cells. Ropeginterferon-α-2b (RopegIFN) has emerged as a promising therapy, capable of lowering the JAK2V617F allele burden and maintaining a complete hematologic response (CHR). Here, we report the cases of four patients with PV who discontinued RopegIFN after achieving CHR. One patient had sustained long-term remission post-discontinuation, with the JAK2V617F allele burden reduced below 10

Ropeginterferon alfa-2b (ropegIFN), a new-generation interferon-based agent, has been approved in Japan for patients with polycythemia vera (PV) who are ineligible for or respond inadequately to conventional treatment. However, long-term outcomes with ropegIFN in Japanese patients have not been reported. This extension of a phase 2 study of ropegIFN in Japanese patients with PV aimed to determine its long-term safety/efficacy, and changes over time in JAK2 V617F allele burden. Here, we report data from the phase 2 study and subsequent extension over a period of 36 months. The primary endpoint was the complete hematologic response (CHR) maintenance rate without phlebotomy (hematocrit value < 45

Background: Acute graft-versus-host disease (GVHD) is a major complication of allogeneic hematopoietic stem cell transplantation (allo-HSCT) with significant morbidity and mortality, and efficacy of currently available therapeutics are limited. Acute and chronic GVHD are similar in that both are initiated by antigen presenting cells and activation of alloreactive B-cells and T-cells, subsequently leading to inflammation, tissue damage, and organ failure. One difference is that acute GVHD is mostly attributed to T-cell activation and cytokine release, whereas B-cells are the key players in chronic GVHD. Ibrutinib is an irreversible inhibitor of the Bruton's tyrosine kinase (BTK), which is part of B-cell receptor signaling. Ibrutinib is currently used for treating chronic GVHD, but its efficacy towards acute GVHD is unknown. Besides BTK, ibrutinib also inhibits interleukin-2 inducible T-cell kinase (ITK), which is predominantly expressed in T-cells and a crucial enzyme for activating the downstream pathway of TCR signaling. ITK activates PLCγ2 and facilitates signaling through NF-κB, NFAT, and MAPK, leading to activation and proliferation of T-cells and enhanced cytokine production. Therefore, the TCR signaling pathway is indispensable for development of acute GVHD, and ITK inhibition by ibrutinib would be a rational therapeutic approach. Case presentation: A 56-year-old male acute myeloid leukemia patient with Myeloid neoplasms with germline DEAD-box RNA helicase 41 (DDX41) mutation underwent cord blood transplantation and developed severe gastrointestinal (GI) acute GVHD which was refractory to steroids and mesenchymal stem cell therapy. While acute GVHD accommodated by multiple life-threatening GI bleeding events persisted, chronic cutaneous GVHD developed, and ibrutinib 420 mg/day was initiated from day 147 of transplant. Although ibrutinib was commenced targeting the chronic GVHD, unexpected and abrupt remission of acute GVHD along with remission of chronic GVHD was observed. Conclusion: Ibrutinib is a promising therapeutic for treating acute GVHD, and further studies are warranted.

Persistent COVID-19 is a well recognized issue of concern in patients with hematological malignancies. Such patients are not only at risk of mortality due to the infection itself, but are also at risk of suboptimal malignancy-related outcomes because of delays and terminations of chemotherapy. We report two lymphoma patients with heavily pretreated persistent COVID-19 in which ensitrelvir brought about radical changes in the clinical course leading to rapid remissions. Patient 1 was on ibrutinib treatment for mantle cell lymphoma when he developed COVID-19 pneumonia which was severe and ongoing for 2 months despite therapy with molnupiravir, multiple courses of remdesivir, one course of sotrovimab, tocilizumab, and steroids. Patient 2 was administered R-CHOP therapy for diffuse large B-cell lymphoma when he developed COVID-19 which was ongoing for a month despite treatment with multiple courses of remdesivir and one course of sotrovimab. A 5-day administration of ensitrelvir promptly resolved the persistent COVID-19 accommodated by negative conversions of RT-qPCR tests in both patients within days. Ensitrelvir is a novel COVID-19 therapeutic that accelerates viral clearance through inhibition of the main protease of SARS-CoV-2, 3-chymotrypsin-like protease, which is vital for viral replication. Ensitrelvir is a promising treatment approach for immunocompromised lymphoma patients suffering from persisting and severe COVID-19.

Objectives: The development of tyrosine kinase inhibitors (TKIs) has significantly improved the prognosis of chronic myeloid leukemia (CML) patients, making their life expectancy nearly equal to that of healthy individuals. However, lower rates of deep molecular response (DMR) and treatment-free remission (TFR) in Sokal or ELTS high-risk patients remain concerns. These scores are calculated based on age, peripheral blood blast count, platelet count, and splenomegaly, which is thought to worsen over time. Therefore, we hypothesize that early diagnosis and treatment intervention could improve the DMR achievement rate and potentially contribute to an increased rate of TFR. Recently, mitochondrial dynamics have drawn attention due to their reported involvement in many diseases; however, there are few reports on that in CML. In this study, we explored the mitochondrial alterations in CML cells and their potential as a novel biomarker for the early diagnosis. Methods: The BCR::ABL1 oncogene was transduced into UT-7/EPO cells. Mitochondrial observations were performed using transmission electron microscopy and confocal laser microscopy. Protein expressions were analyzed by Western blotting. For the detection of CML cells, we introduced a novel technology called ghost cytometry (GC), which is a combination of flow cytometry and artificial intelligence (AI). GC enables the acquisition of signals containing subtle morphological information of the cells in a label-free manner, which are then identified using AI (Ota S et al., Science 2018). Results: Through observing leukocytes from patients with CML and BCR::ABL1-transduced UT-7/EPO, we found that mitochondria were excessively fragmented in these cells compared to leukocytes from healthy individuals and gene-untransduced UT-7/EPO. We discovered that the phosphorylation of dynamin related protein (DRP) 1 and extracellular signal-regulated kinase (ERK) 1/2 in BCR::ABL1-transduced UT-7/EPO were enhanced, while inhibiting DRP1 suppressed excessive mitochondrial fragmentation. Moreover, inhibition of MEK1/2 reduced ERK1/2 and DRP1 phosphorylation, indicating that mitogen-activated protein kinase (MAPK) pathway was involved in BCR::ABL1-induced mitochondrial fragmentation. GC distinguished between BCR::ABL1-positive and negative cells with area under the receiver operating characteristic curve of >0.8, both in cell line and real-world clinical specimens. Subsequently, we created an AI learning model using the peripheral blood leukocytes of CML patients at diagnosis and healthy individuals. We then tested peripheral blood samples from CML patients after TKI initiation, which had white blood cell counts and BCR::ABL1 (IS) mRNA values of 5,960 (1,680 - 11,690) /uL and 48.8 (8.8 - 71.4) %, respectively. The percentages of AI-detected CML cells in these samples showed a high correlation (r = 0.87) with the actual IS values. In addition, we found that AI-based discrimination scores at diagnosis were significantly higher in patients whose IS values were higher than 1% at three months of treatment compared to those lower than 1% (p < 0.05). Discussion: In this study, we revealed that BCR::ABL1 induces excessive mitochondrial fragmentation by activating DRP1 through the MAPK pathway. These morphological alterations could be identified by AI-driven flow cytometry, enabling label-free detection of CML cells before numerical abnormalities in white blood cell counts appear. Moreover, AI-detectable morphological changes might occur between patient groups with different treatment responses. Since clinical symptoms are rarely observed in chronic phase CML, early diagnosis is challenging, especially in patients before the abnormal increase in white blood cell count. By employing this automated AI-driven label-free method as a screening test, GC holds potential for early diagnosis of CML, which may contribute to improving the rate of DMR. Previous studies have shown that patients are more likely to have a prolonged duration of DMR when the IS values are lower than 1.5% at three months of treatment (Sasaki K et al., Cancer 2018). Based on our findings, GC-derived morphological information may predict early treatment response to TKI therapy and subsequent DMR achievement. In conclusion, BCR::ABL1-induced mitochondrial fragmentation could serve as a novel biomarker for the early diagnosis and prediction of treatment efficacy in CML.

Introduction: Splenomegaly is frequently encountered in patients with myeloproliferative neoplasms. Splenomegaly is associated with an increased incidence of engraftment failure during allogeneic hematopoietic stem cell transplantation (allo-HSCT), and some centers perform prior low-dose splenic irradiation (LDSI) which has been reported to be both safe and effective. However, we report conflicting results by presenting three allo-HSCT patients undergoing LDSI that subsequently developed sinusoidal obstruction syndrome (SOS). Case Presentation: The underlying diseases of the presented cases were atypical chronic myeloid leukemia, secondary myelofibrosis following essential thrombocythemia, and acute myeloid leukemia transforming from myeloproliferative neoplasm, unclassifiable. Endothelial Activation and Stress Index (EASIX) scores of the 3 patients were 0.40, 3.82, and 4.40, respectively. Conclusion: SOS is a potentially fatal complication of allo-HSCT, and the mortality rates of severe cases are reported to be above 80%. LDSI has not been recognized as a risk factor of SOS to date. Alternative management methods of splenomegaly during allo-HSCT such as ruxolitinib administration may be safer compared to LDSI.

Objectives:Hemorrhagic events are a rare but potentially fatal complication in patients with polycythemia vera (PV).Methods:We analyzed the characteristics of hemorrhagic events in 267 patients with PV.Results:A median follow-up of 4.8 years revealed that 23 (8.6%) hemorrhagic events occurred. Significantly more hemorrhagic events occurred in younger patients aged below 60 years (n = 72) than in older patients aged 60 years or above (n = 191) (n = 12 [16.7%] vs. n = 11 [5.8%], respectively, P = 0.012). In univariate analysis among the younger patients, white blood cell (WBC) count >= 15 x 109/L (hazard ratio [HR] = 7.746, 95% confidence interval [CI] 2.082-28.830, P = 0.002), palpable splenomegaly (HR = 5.629, 95% CI 1.193-26.550, P = 0.029), and JAK2V617F allele burden >= 80% (HR = 22.850, 95% CI 2.885-181.00, P = 0.003) were associated with an increased risk of hemorrhagic events. In multivariate analysis, JAK2V617F allele burden >= 80% (HR = 9.394, 95% CI 1.046-84.380, P = 0.046) was a significant risk factor.Conclusions:There is an increased risk of hemorrhagic events after diagnosis in younger PV patients with a high JAK2V617F allele burden, high WBC count or palpable splenomegaly. It is important to consider treatment options that aim to avoid hemorrhagic events by reducing the JAK2V617F allele burden in younger PV patients.

Since MPL mutation is a rare driver gene mutation found in a small number of essential thrombocythemia (ET) patients, the clinical characteristics of patients with MPL mutations and their association with thrombotic events have not yet been elucidated in Japan.We enrolled 579 Japanese ET patients based on the diagnostic criteria of the WHO classification 2017 and compared clinical characteristics of MPL-mutated patients (n = 22; 3.8%) to JAK2V617F-mutated (n = 299; 51.6%), CALR-mutated (n = 144; 24.9%), and triple-negative (TN) (n = 114; 19.7%) patients.Thrombosis during follow up was observed in 4 out of 22 (18.2%) in the MPL-mutated group, which was the highest among all driver gene mutation groups (JAK2V617F-mutated, 8.7%; CALR-mutated, 3.5%; TN,1.8%). The MPL- and JAK2V617F-mutated groups had worse thrombosis-free survival (TFS) than the CALR-mutated (p = 0.043) and TN groups (p = 0.006). Univariable analysis revealed that a history of thrombosis was a possible risk factor for thrombosis among MPL-mutated patients (hazard ratio: 9.572, p = 0.032).MPL-mutated ET patients should require more intensive management to prevent recurrence of thrombosis.

Objectives A proportion of patients with polycythemia vera (PV) and essential thrombocythemia (ET) harbor non-driver mutations associated with poor prognosis. In this study, we analyzed the frequency of non-driver mutations in a large Japanese PV and ET cohort. Furthermore, we studied the relationship of these mutations and prognosis in Japanese patients. Methods We enrolled 843 Japanese patients with PV or ET. Non-driver mutations were analyzed by target resequencing using next-generation sequencing. The association of the mutations with the prognosis was estimated using multivariable logistic regression analysis and log-rank test. Results Non-driver mutations were detected in 31.1% and 24.5% patients with PV and ET, respectively. Among them, ASXL1 mutations were identified as a risk factor for leukemic/myelofibrotic transformation in PV and ET patients (hazard ratio: 4.68, p = .006). The higher-risk groups of the mutation-enhanced international prognostic system (MIPSS)-PV and MIPSS-ET incorporating non-driver mutations exhibited significantly shorter overall survival compared with the low-risk group (p < .001). Conclusions These results implicate the importance of studying non-driver mutations for predicting the prognosis and survival of Japanese PV and ET patients.

Risk-adapted therapy is recommended to prevent major clinical complications, such as thrombo-haemorrhagic events, in patients with essential thrombocythaemia (ET). In this study, we analysed the association between non-driver gene mutations and thrombo-haemorrhagic events in 579 patients with ET. ASXL1 and TP53 mutations were frequently identified in patients with ET complicated by thrombosis (22.7% and 23.1%, respectively), and the DNMT3A mutation was frequently identified in patients who experienced haemorrhage (15.2%). Multivariate analyses of thrombosis-free survival (TFS) revealed that ASXL1 and TP53 mutations are associated with thrombosis (hazard ratio [HR] = 3.140 and 3.752 respectively). Patients harbouring the ASXL1 or TP53 mutation had significantly worse TFS rates than those without mutation (p = 0.002 and p < 0.001 respectively). Furthermore, JAK2V617F-mutated patients with accompanying ASXL1 mutations showed significantly shorter TFS compared with those without ASXL1 mutations (p = 0.003). Multivariate analyses of haemorrhage-free survival (HFS) revealed that the DNMT3A mutation (HR = 2.784) is associated with haemorrhage. DNMT3A-mutated patients showed significantly shorter HFS than those without the mutation (p = 0.026). Non-driver gene mutations should be considered in treatment strategies and may provide important information for personalised treatment approaches.

Myelodysplastic/myeloproliferative neoplasm with ring sideroblasts and thrombocytosis (MDS/MPN-RS-T) is a rare disease, which presents with features of myelodysplastic syndromes with ring sideroblasts and essential thrombocythemia, as well as anemia and marked thrombocytosis. SF3B1 and JAK2 mutations are often found in patients, and are associated with their specific clinical features. This study was a retrospective analysis of 34 Japanese patients with MDS/MPN-RS-T. Median age at diagnosis was 77 (range, 51–88) years, and patients had anemia (median hemoglobin: 9.0 g/dL) and thrombocytosis (median platelet count: 642 × 109/L). Median overall survival was 70 (95

Paroxysmal cold hemoglobinuria (PCH) is an extremely rare subtype of autoimmune hemolytic anemia (AIHA) in adults. PCH is caused by the biphasic Donath-Landsteiner (DL) antibody which fixes complement to red blood cells at low temperatures and dissociates at warmer temperatures, leading to complement-mediated intravascular hemolysis. Autoimmune hematological disorders including AIHA and immune thrombocytopenia have been reported to develop following the mRNA COVID-19 vaccination. However, PCH developing subsequent to mRNA vaccination has never been reported. We report a 59-year-old male who developed PCH approximately a month after his second mRNA COVID-19 vaccination.

Risk-adapted therapy is recommended to prevent thrombosis in essential thrombocythemia (ET) patients. An advanced age, a history of thrombosis, and the presence of the JAK2V617F mutation are well-defined risk factors for thrombosis in ET; however, the impact of cardiovascular risk (CVR) factors on thrombosis in ET remains elusive. Therefore, we herein investigated the impact of CVR factors on thrombosis in 580 ET patients who met the 2017 World Health Organization Classification diagnostic criteria. A univariate analysis identified hypertriglyceridemia and multiple CVR factors as strong risk factors for thrombosis (hazard ratio [HR] 3.530, 95% confidence interval [CI] 1.630–7.643, P = 0.001 and HR 3.368, 95% CI 1.284–8.833, P = 0.014, respectively) and hyper-LDL cholesterolemia as a potential risk factor (HR 2.191, 95% CI 0.966–4.971, P = 0.061). A multivariate analysis revealed that hypertriglyceridemia was an independent risk factor for thrombosis (HR 3.364, 95% CI 1.541–7.346, P = 0.002). Furthermore, poor thrombosis-free survival was observed in patients with a serum triglyceride level ≥ 1.2 mmol/L (HR = 2.592, P = 0.026 vs. < 1.2 mmol/L) or two or more CVR factors (P = 0.011 vs. no CVR factors and P = 0.005 vs. one CVR factor). These results revealed the impact of CVR factors on thrombosis in ET. Since CVR factors are manageable, lifestyle interventions, such as the control of serum triglyceride levels, may effectively prevent thrombosis in ET patients.

TAFRO syndrome is a relatively new disease entity first reported in 2010. We report a case of TAFRO syndrome accommodated by abnormal exacerbation of moderately differentiated gastric adenocarcinoma. The pathophysiology of TAFRO syndrome is largely unknown, but because the disease often responds to immunosuppressive therapy and also because T follicular helper (Tfh) cells are reported to be drastically decreased in TAFRO syndrome, involvement of a dysregulated immune system can be speculated. Growing evidence points toward a pivotal role of Tfh cells in tumor immunity through supporting ectopic lymphoid structures, which are recruitment sites for cells directly engaging in antitumor activity such as CD8(+) T cells, NK cells, and macrophages. In fact, Tfh cells are reported to positively correlate with longer survival in human colorectal and breast cancer. Combined with our observations of hyperprogressive gastric cancer in the presented patient, an impaired tumor immunity is strongly indicated in TAFRO syndrome.