A newly identified gene Ahed plays essential roles in murine haematopoiesis.

The development of haematopoiesis involves the coordinated action of numerous genes, some of which are implicated in haematological malignancies. However, the biological function of many genes remains elusive and unknown functional genes are likely to remain to be uncovered. Here, we report a previously uncharacterised gene in haematopoiesis, identified by screening mutant embryonic stem cells. The gene, 'attenuated haematopoietic development (Ahed)', encodes a nuclear protein. Conditional knockout (cKO) of Ahed results in anaemia from embryonic day 14.5 onward, leading to prenatal demise. Transplantation experiments demonstrate the incapacity of Ahed-deficient haematopoietic cells to reconstitute haematopoiesis in vivo. Employing a tamoxifen-inducible cKO model, we further reveal that Ahed deletion impairs the intrinsic capacity of haematopoietic cells in adult mice. Ahed deletion affects various pathways, and published databases present cancer patients with somatic mutations in Ahed. Collectively, our findings underscore the fundamental roles of Ahed in lifelong haematopoiesis, implicating its association with malignancies.

Asciminib in Patients With CML-CP Previously Treated With ≥ 2 Tyrosine Kinase Inhibitors: 96-Week Results From the Japanese Subgroup Analysis of the ASCEMBL Study.

Asciminib is a first-in-class BCR::ABL1 inhibitor that Specifically Targets the ABL1 Myristoyl Pocket (STAMP). It is approved worldwide and in Japan for chronic myeloid leukemia in chronic phase (CML-CP) with resistance or intolerance to previous tyrosine kinase inhibitor (TKI) therapy. In the Phase 3 ASCEMBL study, patients with CML-CP who received ≥ 2 prior ATP-competitive TKIs were randomized (2:1) to asciminib 40 mg twice-daily or bosutinib 500 mg once-daily. Here, we report the 96-week results of the subgroup analysis of Japanese patients (asciminib, n = 13; bosutinib, n = 3) in the ASCEMBL study. The MMR rate at Week 96 was 46.2% in asciminib-treated patients, increasing from Weeks 24 and 48. Patients who achieved MMR at Week 24 remained in MMR up to the Week 96 cutoff. While a high proportion of patients treated with asciminib remained on treatment at cutoff, none randomized to bosutinib were on treatment at Week 96. Despite the longer duration of exposure to asciminib, its safety and tolerability continued to be favorable with no new or worsening safety findings. Overall, the efficacy and safety outcomes in the Japanese subgroup were comparable with the ASCEMBL global study population, which supports the use of asciminib in Japanese patients with previously treated CML-CP.

Differentiating Between Epstein-Barr Virus-positive Lymphoid Neoplasm Relapse and Post-transplant Lymphoproliferative Disorder After Sex-mismatched Hematopoietic Stem Cell Transplantation.

After allogeneic hematopoietic stem cell transplantation (HSCT), accurate differentiation between donor-derived post-transplant lymphoproliferative disorder (PTLD) and relapse of recipient-derived lymphoproliferative disorder (LPD) is crucial for determining treatment. Conventional diagnostic approaches for PTLD include histopathological examination, flow cytometry, and chimerism analysis of bulk tumor tissue. However, these methods are inconclusive in cases in which the primary disease is an Epstein-Barr virus (EBV)-positive LPD and is of the same lineage as that of the post-HSCT LPD tumor cells. Particularly, in cases where the number of tumor cells in the tissue is low, it is difficult to determine the origin of tumor cells. In this study, we developed a new method to simultaneously detect signals using sex chromosome fluorescence in situ hybridization, immunofluorescence staining, and EBV-encoded small RNA in situ hybridization on a single section of formalin-fixed paraffin-embedded histopathological specimen. The utility of the method was validated using specimens from 6 cases of EBV-positive LPD after sex-mismatched HSCT that were previously difficult to diagnose, including Hodgkin lymphoma-like PTLD that developed after HSCT for Hodgkin lymphoma and recurrence of chronic active EBV infection. This method successfully preserved the histologic structure after staining and allowed accurate determination of tumor cell origin and lineage at the single-cell level, providing a definitive diagnosis in all cases. This method provides a powerful tool for the diagnosis of LPDs after sex-mismatched HSCT.

A single-institution pre-post comparison of subcutaneous immunoglobulin replacement therapy in allogeneic haematopoietic cell transplantation recipients.

Immunoglobulin replacement therapy (IgRT) reduces the risk of infection in hypogammaglobulinaemia secondary to chronic lymphocytic leukaemia and multiple myeloma. However, the benefit of IgRT, especially subcutaneous IgRT (ScIgRT), has not been assessed in hypogammaglobulinaemia after allogeneic haematopoietic cell transplantation (allo-HCT). We performed a pre-post comparison of the clinical impact of ScIgRT after allo-HCT in a retrospective analysis of 209 patients who underwent allogeneic HCT at our institution from 2011 to 2019. Since ScIgRT became available at our institution in April 2017, we categorized patients treated from January 2011 to March 2017 as the Pre-ScIgRT group (n = 118) and those treated from April 2017 to December 2019 as the Post-ScIgRT group (n = 91). The 2-year overall survival rate was 65% in the Pre-ScIgRT group and 81% in the Post-ScIgRT group (p = 0.02). The cumulative incidence (CI) of non-relapse mortality at 2 years was 18% and 7% (p = 0.02). There were 78 infectious events in 44 patients in the Pre-ScIgRT group and 28 such events in 19 patients in the Post-ScIgRT group. The CI of the documented infection during the observation period was between 38% and 21% (p = 0.01). Our study suggests that ScIgRT may reduce infection rates and improve prognosis after allo-HCT.

Gilteritinib in peritransplant period for relapsed or refractory FLT3-mutated acute myeloid leukemia: A case report of three patients.

Patients with relapsed or refractory acute myeloid leukemia (RR-AML) with mutations of FMS-like tyrosine kinase 3 (FLT3) have a poor prognosis even after allogeneic hematopoietic cell transplantation (allo-HCT). Multiple FLT3 inhibitors, including gilteritinib, have been developed and serve as treatment options for RR-AML. Here, we describe three cases of FLT3 mutated RR-AML that were successfully treated with gilteritinib administration before and after allo-HCT. Gilteritinib treatment before HCT was helpful in achieving remission. However, HCT often resulted in mild liver damage, and careful introduction of gilteritinib after HCT at a lower dose may be helpful for its safe usage. The three cases discussed had a successful clinical outcome in terms of disease control as well as the management of side effects associated with gilteritinib treatment.

Relationship between muscle thickness measured by ultrasound and physical functions: A 2-year follow-up study of allogeneic hematopoietic stem cell transplantation recipients.

BACKGROUND: Allogeneic hematopoietic stem cell transplantation (allo-HSCT) is a potentially curative treatment for hematological malignancies. Several complications following allo-HSCT, such as graft-versus-host disease, infection, and malnutrition, often cause physical dysfunction, and the assessment of physical function and evaluation of muscle mass are incompletely performed. Use of ultrasound (US) allows muscle mass measurement in patients with poor general conditions. In allo-HSCT recipients, the correlation between physical function and muscle thickness, as measured by US, remains unclear. OBJECTIVE: To clarify whether muscle thickness measured by US correlated with physical function in allo-HSCT recipients. DESIGN: A single-center prospective cohort study. SETTING: Hospital. PATIENTS: Ninety-two patients underwent allo-HSCT at our hospital from April 2017 to March 2019. INTERVENTIONS: Not applicable. MAIN OUTCOME MEASURE(S): Biceps and quadriceps muscle thickness measured by US, grip strength, isometric muscle strength (elbow flexion and knee extension), and 6-minute walking test (6MWT) before allo-HSCT and on days 30, 90, 180, 1 year, and 2 years after allo-HSCT. The implementation rates of these assessments were also investigated. RESULTS: Correlations were observed between biceps thickness and elbow flexion muscle strength/grip strength before allo-HSCT, on days 30, 90, 180, 1 year, and 2 years after allo-HSCT (r = 0.71/0.74, 0.73/0.72, 0.70/0.79, 0.67/0.75, 0.72/0.75, and 0.85/0.79, respectively, all p < .001). At the same time points, quadriceps thickness moderately correlated with knee extensor strength (r = 0.49, 0.50, 0.45, 0.64, 0.61, and 0.58, all p < .001). However, biceps and quadriceps thicknesses did not correlate with the 6MWT. The percentages of patients measured with US and 6MWT were 93.4% and 82.4% (p = .01) on day 30 and 97.5% and 87.8% (p = .02) on day 90, respectively. CONCLUSIONS: US assessment may be a useful alternative method for estimating muscle strength in fragile allo-HSCT recipients, particularly when physical function assessment is difficult to quantify.

A case of severe oral mucosal GVHD induced by heterologous SARS-CoV-2 vaccination after cord blood transplantation.

Patients who have undergone hematopoietic cell transplantation (HCT) are at a higher risk of severe acute respiratory syndrome coronavirus 2 (SARS-Cov-2) infection than the general population. Therefore, early vaccination is recommended for post-transplant patients. Although exacerbation of chronic graft-versus-host disease (cGVHD) after the initial vaccination has been reported, it is unknown whether severe cGVHD occurs when different RNA vaccines are combined. We treated a patient who developed severe oral mucosal cGVHD after receiving two different RNA vaccines. Visual inspection showed that the patient presented with typical mucocutaneous cGVHD, and cGVHD in this case responded well to low-dose steroids compared to common oral GVHD exacerbations. Histopathological findings revealed T cell, B cell, and conspicuous neutrophil infiltration. Multiple doses of SARS-Cov2 vaccination are required in post-transplant recipients. In conclusion, it is essential to obtain the vaccination history of allo-HSCT recipients with cGVHD exacerbation. Furthermore, reviewing the pathological findings may help treat patients with lower doses of steroids.

Successful Bridging to Allogeneic Hematopoietic Stem Cell Transplantation by Azacitidine and Venetoclax in a Case of Acute Myeloid Leukemia With t(3;3)(q21.3;q26.2) Developed Early After Orthotopic Heart Transplantation.

A 48-year-old male patient developed acute myeloid leukemia (AML) with t(3;3)(q21.3;q26.2) chromosomal mutation 8 months after orthotopic heart transplantation from a human leukocyte antigen-unmatched brain-dead donor for cardiac sarcoidosis. He had sequelae of stroke and chronic renal failure at the time of AML diagnosis. He received 3 cycles of azacitidine and venetoclax induction therapy and achieved complete hematological remission with incomplete count recovery without causing severe complications, including infection. He sequentially underwent allogeneic peripheral blood stem cell transplantation from a HLA-8/8 matched, ABO-blood matched, unrelated female donor and successfully achieved donor cell engraftment. His transplanted heart was viable, and the coronary vessels were not damaged even after allogeneic peripheral blood stem cell transplantation. Although AML relapsed afterward, azacytidine/venetoclax was a tolerable bridging therapy even for early-onset AML after heart transplantation.

[Acquired hemophilia A following BNT162b2 mRNA COVID-19 vaccination].

Acquired hemophilia A (AHA) is a rare disease characteized by bleeding symptoms caused by decreased factor VIII activity due to the appearance of inhibitors to factor VIII triggered by malignancy or collagen disease. An 86-year-old woman developed purpura on her extremities after the first dose of the BNT162b2 mRNA COVID-19 vaccine. This symptom subsided after a few days. After the second dose of the BNT162b2 mRNA COVID-19 vaccine, purpura appeared again, and the patient was referred to our hospital Her APTT was remarkably prolonged to 110 seconds, and a cross-mixing test revealed an inhibitor pattern. Since FVIII activity was <1% and FVIII inhibitor was 51.6 BU, she was diagnosed with AHA. Prednisolone therapy was started, and coagulative complete remission was achieved. Because acquired hemophilia can develop after mRNA COVID-19 vaccination, as in this case, it is critical to monitor the appearance of bleeding symptom.

Prolonged gut microbial alterations in post-transplant survivors of allogeneic haematopoietic stem cell transplantation.

Dysbiosis of the gut microbiota has been reported to increase early complications after allogeneic haematopoietic stem cell transplantation (allo-HSCT). However, it remains unclear whether gut microbial alterations persist during late complications, such as chronic graft-versus-host disease (cGVHD) or secondary cancers. Here, we analysed the gut microbiota of 59 patients who survived for 1-21.7 years (median, 6.4 years) after allo-HSCT. Long-term survivors showed lower gut microbial diversity than the age- and sex-matched healthy controls. This decreased diversity was reflected in the reduced abundance of the butyrate-producing bacteria. Patients with a history of grade 3 acute graft-versus-host disease (aGVHD) exhibited higher Veillonella abundance than patients with a history of grade 1-2 or non-aGVHD cases. The abundance of Faecalibacterium showed no decrease only in limited cGVHD cases. Additionally, the microbial structure in the secondary cancer group was significantly different (p < 0.05) from that in the non-secondary cancer group. This study is the first to show that microbial dysbiosis is present over a 10-year lifetime after discharge following allo-HSCT. Our results suggest that these prolonged gut microbial alterations may be associated with the development and exacerbation of late complications in post-transplant survivors.

Asciminib vs bosutinib in CML patients pretreated with ≥2 tyrosine kinase inhibitors: Results from the Japanese subgroup analysis of ASCEMBL study.

Asciminib, a first-in-class, allosteric inhibitor of BCR-ABL1 that acts by STAMP (Specifically Targeting the ABL Myristoyl Pocket), is a novel therapeutic option for patients with chronic myeloid leukemia (CML). In the global, phase 3, open-label ASCEMBL study in patients with CML in chronic phase (CML-CP) pretreated with ≥2 tyrosine kinase inhibitors (TKIs) (NCT03106779), asciminib (40 mg twice-daily) demonstrated significant superiority over the ATP-competitive TKI bosutinib (500 mg once daily) for the primary endpoint of major molecular response (MMR; BCR::ABL1 transcript levels on the international scale [BCR::ABL1IS ] ≤0.1%) at week 24. Here, we report results from a descriptive subgroup analysis of Japanese patients enrolled in ASCEMBL study (data cut-off: May 25, 2020). Overall, 16 Japanese patients were randomized (asciminib, n = 13; bosutinib, n = 3). At week 24, the MMR rate with asciminib was 30.8% (4/13; 95% confidence interval [CI], 9.09-61.43). BCR::ABL1IS ≤1% and complete cytogenic response (CCyR) at week 24 were 61.5% (8/13 patients) and 50.0% (4/8 patients), respectively. In the bosutinib group, no patient achieved MMR, CCyR, or BCR::ABL1IS ≤1%, but results were limited by the low number of patients. The safety profile of asciminib was comparable to that previously observed in the overall study population. Findings from this Japanese subgroup analysis of the ASCEMBL study support the use of asciminib for the treatment of Japanese patients with CML-CP previously treated with ≥2 TKIs. ClinicalTrials.gov Identifier: NCT03106779.

[Exploring new molecules that regulate hematopoietic stem cells and early stages of lymphoid hematopoiesis: the functional significance of ESAM and SATB1].

Hematopoietic stem cells (HSCs) possess multilineage differentiation capability, which sustains the production of blood and immune cells throughout life. However, the precise mechanisms by which HSCs initiate differentiation toward a particular lineage and the factors that attenuate their lymphopoietic potential with aging are yet to be elucidated. Our group has investigated this issue for over two decades. We initially developed a method for segregating early lymphoid progenitors from HSCs and identified two molecules: endothelial cell-selective adhesion molecule (ESAM), highly expressed in HSCs, and special AT-rich sequence binding protein 1 (SATB1), expressed in early lymphoid progenitors. ESAM marks HSCs across species, including humans. In addition to its significance in stress-induced hematopoiesis, ESAM is also useful in identifying features of human acute myeloid leukemia stem cells. Further, we determined the role of SATB1 in the early HSC differentiation processes toward the lymphoid lineage. Remarkably, SATB1 expression in HSCs significantly decreased with aging, whereas its exogenous induction in aged HSCs rejuvenated their lymphopoietic potential. Furthermore, SATB1-expressing HSCs demonstrated robust lymphopoietic and long-term reconstituting capability, whereas HSCs without SATB1 skewed toward the myeloid lineage. Thus, our continuing research has revealed the significance of ESAM and SATB1 in the fundamental biology of HSCs.

Special AT-Rich Sequence-Binding Protein 1 Supports Survival and Maturation of Naive B Cells Stimulated by B Cell Receptors.

Epigenetic mechanisms underpin the elaborate activities of essential transcription factors in lymphocyte development. Special AT-rich sequence-binding protein 1 (SATB1) is a chromatin remodeler that orchestrates the spatial and temporal actions of transcription factors. Previous studies have revealed the significance of SATB1 in T cell lineage. However, whether and how SATB1 controls B cell lineage development is yet to be clarified. In this study, we show that SATB1 is an important factor during splenic B cell maturation. By analyzing SATB1/Tomato reporter mice, we determined the dynamic fluctuation of SATB1 expression in the B cell lineage. Although SATB1 expression decreased to minimal levels during B cell differentiation in the bone marrow, it resurged markedly in naive B cells in the spleen. The expression was dramatically downregulated upon Ag-induced activation. Splenic naive B cells were subdivided into two categories, namely SATB1high and SATB1-/low, according to their SATB1 expression levels. SATB1high naive B cells were less susceptible to death and greater proliferative than were SATB1-/low cells during incubation with an anti-IgM Ab. Additionally, SATB1high cells tended to induce the expression of MHC class II, CD86, and CD83. Accordingly, naive B cells from B lineage-specific SATB1 conditional knockout mice were more susceptible to apoptosis than that in the control group upon anti-IgM Ab stimulation in vitro. Furthermore, conditional knockout mice were less capable of producing Ag-specific B cells after immunization. Collectively, our findings suggest that SATB1 expression increases in naive B cells and plays an important role in their survival and maturation.

Inotuzumab ozogamicin and blinatumomab sequential therapy for relapsed/refractory Philadelphia chromosome-positive acute lymphoblastic leukemia.

To overcome the unfavorable outcome of refractory/relapsed (R/R) Philadelphia chromosome-positive acute lymphoblastic leukemia (Ph+ALL) and conduct allogeneic stem cell transplantation (allo-SCT) safely, we designed a sequential therapy involving a single cycle of Inotuzumab ozogamicin (InO) and Blinatumomab (Blina). Two heavily treated and aged patients with R/R Ph+ALL were treated with the therapy. Both of them achieved complete molecular remission without cytokine release syndrome and underwent allo-SCT without veno-occlusive disease/sinusoidal obstruction syndrome. Although appropriate central nervous system prophylaxis should be added, the InO-Blina sequential therapy is a promising strategy for treating R/R Ph+ALL as a bridging regimen before allo-SCT.

Impaired B cell terminal differentiation in B cell-specific knockout mice of cell death-defying factor anamorsin.

Anamorsin (AM) is an anti-apoptotic molecule cloned by us as a molecule that confers resistance against apoptosis induced by growth factor deprivation. AM-deficient mice are embryonic lethal, which impedes detailed analyses of the roles of AM in various types of adult cells. To overcome the embryonic lethality, we generated AM conditional knockout (AMflox/flox) mice and cell type-specific genetic modification became possible using the Cre-loxP system. CD19-Cre/AMflox/flox mice with AM deleted specifically in CD19+ B cells exhibited less B220+ B cells in their spleen, peripheral blood, and lymph node compared with control CD19-Cre mice. Using flow cytometry to categorize bone marrow and spleen cells into B cell subsets, we observed significantly less follicular type I cells, which are the most mature follicular B cells, compared with control CD19-Cre mice. These data suggest that AM has an important role in the generation of mature B cells.

Nivolumab-induced systemic lymphadenopathy occurring during treatment of malignant melanoma: a case report.

Nivolumab is an anti-programmed cell death protein 1 monoclonal antibody that exhibits significant efficacy in treating melanoma and other malignancies. However, various nivolumab-induced immune-related adverse events (irAEs) have been reported, and differentiating irAEs from tumor progression is sometimes difficult. Here, we report a case of reactive lymphadenopathy occurring after treatment with nivolumab. A 56-year-old man with stage IIIC melanoma received adjuvant therapy with nivolumab after wide local excision. He developed systemic lymphadenopathy and autoimmune hemolytic anemia 1 month after receiving seven cycles of nivolumab. Pathological analysis of a cervical lymph node biopsy specimen revealed no metastatic lesion or any other malignancy, including lymphoma. Thus, the patient was diagnosed with nivolumab-induced reactive lymphadenopathy. Systemic corticosteroids were administered to reduce hemolysis, which led to the resolution of lymphadenopathy. When progressive lymphadenopathy is observed in a patient who received immune checkpoint inhibitor therapy, reactive lymphadenopathy should be carefully distinguished from progression to lymphoid metastasis, and biopsy should be performed if needed.

Clinical implications of combination therapy with quizartinib and craniospinal irradiation for refractory acute myeloid leukemia positive for FMS-like tyrosine kinase 3-internal tandem duplication with central nervous system involvement.

FMS-like tyrosine kinase 3-internal tandem duplication (FLT3-ITD) mutation-positive acute myeloid leukemia (AML) has a poor prognosis. We report the first case of successful bridge therapy of novel FLT3 inhibitor, quizartinib, to umbilical cord blood stem cell transplantation for FLT3-ITD-positive AML-primary induction failure patients with central nervous system involvement.

Autoimmune-mediated thrombocytopenia after allogeneic hematopoietic stem cell transplantation: significance of detecting reticulated platelets and glycoprotein-specific platelet autoantibodies.

Autoimmune hematological disorders are rare complications after allogeneic hematopoietic stem cell transplantation (allo-HSCT). Diagnosis of immune thrombocytopenia (ITP) is challenging, especially after allo-HSCT, because various complications such as graft-versus-host disease, disease relapse, viral infection, thrombotic microangiopathy, and drug side effects can also cause thrombocytopenia. Assessment of reticulated platelets (RP) and plasma thrombopoietin (TPO) levels may be useful to distinguish between ITP and hypoplastic thrombocytopenia. ITP is generally characterized by an increased percentage of RP, and a normal or slightly increased plasma TPO level. We now report three cases of thrombocytopenia after allo-HSCT. RP% was elevated in these patients, as it is in primary ITP. However, in contrast to primary ITP, plasma TPO levels were high in two of three patients. Anti-αIIbß3 and anti-GPIb/IX-specific direct IgG antibodies were detected as well, suggesting occurrence of immune-mediated platelet destruction in addition to bone marrow suppression in two patients. All three patients were successfully treated with corticosteroids and/or thrombopoietin receptor agonists (TPO-RAs). These results suggest that increased RP% and detection of glycoprotein-specific platelet autoantibodies are useful for the diagnosis of ITP after HSCT.

Relationship between donor-specific HPA-15 antibodies and poor graft function in HPA-15 mismatched cord blood transplantation.

Poor graft function (PGF) is a fatal complication following hematopoietic stem cell transplantation and is influenced by multiple factors, such as donor-specific anti-HLA antibodies, a poor infused CD34+ cell count, and the donor source. Alloantibodies against human platelet antigen 15 (HPA-15) recognize platelet membrane glycoprotein CD109, which is expressed not only on platelets, but also on megakaryocytes and specific hematopoietic stem cells. HPA-15 antibodies are known to induce platelet transfusion refractoriness and neonatal alloimmune thrombocytopenia, but their effects on graft function following hematopoietic stem cell transplantation remain unknown. We encountered a case of HPA-15 mismatched cord blood transplantation with a high HPA-15b antibody titer. Prolonged PGF and megakaryocyte aplasia with sustained high-titer HPA-15b antibodies were attenuated by rituximab therapy, and rapid recovery of hematopoiesis was achieved. HPA-15-compatible platelet transfusions were highly effective for platelet recovery. Methylcellulose assays and megakaryocyte cultures revealed that patient serum inhibited in vitro hematopoietic development from patient bone marrow cells. These results suggest that HPA-15 antibodies might be a cause of PGF and that reducing the HPA-15 antibody titer might improve graft function in HPA-15 mismatched transplantation.