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As chimeric antigen receptor (CAR) T cell therapy targeting CD19 has shown favorable outcomes in patients with relapsed or refractory (r/r) mature B cell lymphomas and B cell acute lymphoblastic leukemia (B-ALL), an increasing number of patients are waiting to receive these treatments. Optimized protocols for T cell collection by lymphapheresis for chimeric antigen receptor (CAR) T cell therapy are urgently needed to provide CAR T cell therapy for patients with refractory and progressive disease and/or a low number of lymphocytes owing to prior chemotherapy. The predicted efficiency of CD³⁺ cell collection in apheresis can guide protocols for apheresis, but a clinically applicable model to produce reliable estimates has not yet been established. In this study, we prospectively analyzed 108 lymphapheresis procedures for tisagenlecleucel therapy at 2 centers. The apheresis procedures included 20 procedures in patients with B cell acute lymphoblastic leukemia and 88 procedures in patients with diffuse large B cell lymphoma, with a median age at apheresis of 58 years (range, 1 to 71 years). After lymphapheresis with a median processing blood volume of 10 L (range, 3 to 16 L), a median of 3.2 × 10⁹ CD³⁺ cells (range, .1 to 15.0 × 10⁹ cells) were harvested. Collection efficiency 2 (CE2) for CD³⁺ cells was highly variable (median, 59.3%; range, 11.0% to 199.8%). Multivariate analyses revealed that lower hemoglobin levels, higher circulating CD3+ cell counts, and higher platelet counts before apheresis significantly decreased apheresis CE2. Based on multivariate analyses, we developed a novel formula that estimates CE2 from precollection parameters with high accuracy (r = .56; P < .01), which also suggests the necessary processing blood volume. Our strategy for lymphapheresis should help reduce collection failure, as well as achieve efficient utilization of medical resources in clinical practice, thereby allowing delivery of CAR T cell therapy to more patients in a timely manner., キメラ抗原受容体T細胞療法におけるリンパ球採取効率化の取り組み --最適な治療戦略策定への貢献に期待--. 京都大学プレスリリース. 2022-06-20.

Background & Aim Fetal bovine serum (FBS) is a common component of culture media and usually used for cellular research, as well as recent cell-based medical products. However, due to the high risk of contaminations and the variation from batch to batch, FBS might influence the outcome of research or cellular manufacturing. FBS also contains moral concerns because it harvested from bovine fetuses taken from pregnant cows. In addition, FBS is most expensive part of cell culture. To overcome several problems regarding FBS, we developed a new serum, adult bovine platelet lysate-derived cell culture supplement (NeoSERA), and investigate its usefulness as an alternative to FBS. Methods, Results & Conclusion NeoSERA is produced in Japan, considered to be a negligible bovine spongiform encephalopathy (BSE) risk country. We obtain adult donor bovine blood from animal less than 36 months old according to the regulation of EMA for BSE (EMA/410/01 rev.3). Using apheresis medical devices with closed disposable kits, sterile platelet-rich plasma (PRP) is collected from healthy donor bovine receiving a regular veterinary check. After stimulation and removal of coagulated fibrin by centrifugation, NeoSERA is collected in a completely closed system. To meet the scope of directives that apply to produce medicinal products from the European Agency for the Evaluation of Medicinal Products (EMEA/CVMP/743/00) and the United States Department of Agriculture (9CFR§113.450), NeoSERA is finally g-irradiated at a dose of over 30 kGy. NeoSERA completely meets the standard for biological ingredients in Japan (Ministry of Health, Labour and Welfare Notification No. 375 2014) and also obtained the certificate of eligibility for the raw material of regenerative medicine from Pharmaceuticals and Medical Devices Agency (PMDA, No. 0417002, 2017. 4.17). To test whether NeoSERA is useful for the expansion of mesenchymal stem cells (MSCs), a cell culture experiment was performed. 3 to 5 days after NeoSERA treatment, the proliferation of human bone marrow-, adipose tissue-, umbilical cord- and amnion-derived MSCs was significantly increased (p Our results confirm that safe, less ethical and economical bovine NeoSERA profoundly enhances the proliferation of MSCs for regenerative medicine and several cell lines for vaccine production.

Acute myeloid leukemia may manifest as myeloid sarcoma in a variety of extramedullary (EM) tissues at diagnosis or at relapse. Although EM relapse after allogeneic hematopoietic stem cell transplantation (alloSCT) has been considered to be rare, recent studies have suggested that it occurs in 5% to 12% of patients who receive alloSCT, accounting for 7% to 46% of total relapses. The incidence of EM relapse after immunomodulation (eg, donor lymphocyte infusion) or a second SCT is even higher. Moreover, patients with EM relapse are more likely to have had preceding acute graft-versus-host disease or chronic graft-versus-host disease relative to those with bone marrow relapse. Collectively, these observations suggest that the preferential occurrence of the graft-versus-leukemia effect underlies the pathogenesis of EM relapse. Establishing an early diagnosis of EM relapse has been challenging because of the immense diversity in the relapse sites; however, recent studies have suggested the usefulness of (18)F-fluorodeoxyglucose positron emission tomography scans in the detection of EM relapse. As a treatment for EM relapse, a combination of local and systemic therapy should be considered, because local therapy alone often results in subsequent systemic relapse. The prognosis for patients who develop EM relapse after SCT remains poor but is slightly better than that after bone marrow relapse. In addition to an early diagnosis with new modalities, clinical studies using new agents that may offer systemic activity while preserving the graft-versus-leukemia effect are warranted as part of an effort to improve the clinical outcome.

Introduction We previously reported an interim analysis of the DADI (dasatinib discontinuation) trial. The results showed that 48% of patients with chronic myeloid leukemia in the chronic phase who maintained a deep molecular response (DMR) for ≥ 1 year could discontinue second- or subsequent-line dasatinib treatment safely at a median follow-up of 20 months. However, the results from longer follow-up periods would be much more useful from a clinical perspective. Patients and Methods The DADI trial was a prospective, multicenter trial conducted in Japan. After confirming a stable DMR for ≥ 1 year, dasatinib treatment subsequent to imatinib or nilotinib was discontinued. After discontinuation, the loss of DMR (even of 1 point) was defined as stringent molecular relapse, thereby triggering therapy resumption. The predictive factors of treatment-free remission (TFR) were analyzed. Results The median follow-up period was 44.0 months (interquartile range, 40.5-48.0 months). The estimated overall TFR rate at 36 months was 44.4% (95% confidence interval, 32.0%-56.2%). Only 2 patients developed a molecular relapse after the 1-year cutoff point. The presence of imatinib resistance was a significant risk factor for molecular relapse. Moreover, high natural killer cell and low γδ+ T-cell and CD4+ regulatory T-cell (CD25+CD127low) counts before discontinuation correlated significantly with successful therapy discontinuation. Conclusion These findings suggest that discontinuation of second- or subsequent-line dasatinib after a sustained DMR of ≥ 1 year is feasible, especially for patients with no history of imatinib resistance. In addition, the natural killer cell count was associated with the TFR.

Objective We investigated whether established graft-vs-host disease (GVHD) could be successfully treated by a second allogeneic bone marrow transplantation (BMT) through elimination of first donor–derived lymphocytes responsible for GVHD. Materials and Methods In a murine GVHD model of BDF1 (H-2 b/d )→B6C3F1(H-2 b/k ), GVHD mice underwent a second BMT using a graft (1 × 10 7 bone marrow and 3 × 10 7 spleen cells) from a major histocompatibility complex (MHC) antigen haploidentically mismatched (to host and also to first donor) mouse strain, B6B10F1(H-2 b/s ), following low-dose total body irradiation (TBI) 2 to 3 weeks after the first BMT. Results Results demonstrated that severe GVHD could be successfully and stably treated by a second allogeneic BMT. For successful treatment of GVHD, rapid achievement of full second–donor T-cell chimerism was required. Furthermore, we showed that mice with GVHD could easily accept MHC haploidentically mismatched second-donor hematopoietic cells even after minimal conditioning (2–4 Gy TBI) because they were in a profoundly immunosuppressed state, and that the mice were relatively resistant to new development of GVHD by second-donor grafts. Furthermore, the timing of the second BMT, the intensity of conditioning treatment (GVHD mice are very sensitive), and donor selection were also found to be important for obtaining successful outcomes. Increased regulatory T cells and reduction of interferon-γ levels may be involved in tolerance induction. Conclusions We demonstrated that established GVHD in a murine GVHD model could be successfully treated by a second BMT from a third-party allogeneic donor.

Objective The incidence of severe graft-vs-host disease (GVHD) in unmanipulated human leukocyte antigen (HLA) 2-3 antigen–mismatched bone marrow transplantation (BMT) using cyclosporine and methotrexate as GVHD prophylaxis is 80% to 90%. We investigated whether pharmacological GVHD prophylaxis consisting of four drugs, including a steroid, effectively suppressed GVHD in this transplantation setting. Materials and Methods Thirty patients who had hematologic malignancies at an advanced stage or with poor prognosis underwent allogeneic BMT using a myeloablative preconditioning regimen consisting of cyclophosphamide (60 mg/kg × 2), total body irradiation (8–10 Gy), and fludarabine (30 mg/m 2 × 4) with or without cytosine arabinoside (2 g/m 2 × 4), and GVHD prophylaxis consisting of a combination of tacrolimus, methotrexate, mycophenolate mofetil, and methylprednisone (2 mg/kg). Early therapeutic intervention for GVH reaction or grade I GVHD was performed, and steroid was slowly tapered. Results All patients achieved donor-type engraftment. Neutrophil (>0.5 × 10 9 /L) and platelet (>20 × 10 9 /L) engraftment was achieved on day 13 and on day 30, respectively. Seventeen patients (56.7%) had no GVHD. Eleven patients (36.7%) developed grade II–III acute GVHD. Seven patients (23.3%) died of transplant-related toxicity, including fungal or viral infections and thrombotic microangiopathy, and four patients died of disease progression. Estimated relapse rate at 3 years was only 20.9%. The probability of survival at 3 years was 49.9%. Conclusions These data suggest that, in unmanipulated HLA-haploidentical allogeneic BMT, this GVHD prophylactic regimen, which includes methylprednisolone 2 mg/kg, and early therapeutic intervention for GVH reaction suppress the incidence of severe GVHD to an acceptable level, while preserving the graft-vs-leukemia effect.

Pulmonary dysfunction is a significant complication following allogeneic hematopoietic stem cell transplantation (HSCT), and is associated with significant morbidity and mortality. Effective antimicrobial prophylaxis and treatment strategies have increased the incidence of noninfectious lung injury, which can occur in the early posttransplant period or in the months and years that follow. Late-onset noninfectious pulmonary complications are frequently encountered, but diagnostic criteria and terminology for these disorders can be confusing and therapeutic approaches are suboptimal. As a consequence, inaccurate diagnosis of these conditions may hamper the appropriate data collection, enrollment into clinical trials, and appropriate patient care. The purpose of this review is to clarify the pathogenesis and diagnostic criteria of representative conditions, such as bronchiolitis obliterans syndrome and bronchiolitis obliterans organizing pneumonia, and to discuss the appropriate diagnostic strategies and treatment options.

The major problems in human leukocyte antigen (HLA)-mismatched stem cell transplantation (SCT) are graft failure and graft-versus-host disease (GVHD). Less-intensive regimens should be associated with a lower release of inflammatory cytokines and possibly less GVHD. The objective of this study was to investigate whether HLA-haploidentical SCT can be performed using nonmyeloablative conditioning and pharmacologic GVHD prophylaxis, including glucocorticoids. Using conditioning consisting of fludarabine, busulfan, and anti–T-lymphocyte globulin and GVHD prophylaxis consisting of tacrolimus and methylprednisolone (1 mg/kg/day), 26 patients who had hematologic malignancies in an advanced stage or with a poor prognosis underwent transplantation using peripheral blood stem cells from an HLA-haploidentical donor (2–3 antigen mismatches in the graft-versus-host [GVH] direction) without T-cell depletion. All patients except for 1 achieved donor-type engraftment. Rapid hematologic engraftment was achieved (neutrophils > 0.5 × 109/L on day 12 and platelets > 20 × 109/L on day 12), with full donor chimerism achieved by day 14. Fifteen patients did not develop acute GVHD clinically, and only 5 patients developed grade II GVHD. The recovery of CD4+ T cells was delayed compared with that of CD8+ T cells. Sixteen of the 26 patients are alive in complete remission. Four died of transplantation-related causes, and 6 died of progressive disease. These data suggest that nonmyeloablative conditioning, GVHD prophylaxis consisting of tacrolimus and methylprednisolone, and early therapeutic intervention for the GVH reaction allow stable engraftment and effectively suppress GVHD in HLA 2–3 antigen-mismatched SCT.

Although the mobilization of peripheral blood stem cells from normal donors using granulocyte colony-stimulating factor is widely used, the ideal method for the administration of filgrastim has not been determined. Therefore, we compared the efficacy of peripheral blood stem cell mobilization on day 4 of filgrastim between once-daily (group O) and twice-daily (group T) administration of filgrastim at 400 microg/m(2)/d. In all, 38 and 34 donors were randomly assigned to groups O and T, respectively. The number of CD34(+) cells collected on day 4 was not significantly different (1.74 x 10(6) cells/kg in group O and 2.08 x 10(6) cells/kg in group T, P = .37). The incidence and severity of adverse events were similar in the two groups. The baseline white blood cell count was the strongest predictor of poor mobilization. Donor age, sex, and serum concentrations of several cytokines did not significantly affect the CD34(+) cell yield. In conclusion, once-daily administration of filgrastim at 400 microg/m(2)/d appeared to be appropriate for the mobilization of CD34(+) cells in normal donors when apheresis is planned on day 4 of filgrastim. Selection of a donor with a steady-state white blood cell count of 5.0 x 10(9)/L or more may lead to a lower incidence of poor mobilization.

The leaching behavior of heat-treated waste ash was studied to verify the possibility of the thermal treatment of waste incineration ash in existing incinerators and boiler combustion chambers. The influence of temperature, oxygen concentration and treatment time on the leaching behaviors of harmful heavy metals, especially lead (Pb) and chromium(VI) (Cr (VI)) were studied to clarify effective treatment conditions to suppress leaching. By examining the leaching behavior of Pb and Cr from ash heat-treated under various conditions, it was found that leaching can be suppressed by heat-treating the ash under conventional combustion conditions of around 900–1000 °C at 5–10% oxygen concentration. The leaching behaviors of Pb and Cr (VI) from real ash with different particle sizes and from model samples were also investigated in detail to find an effective method to suppress Pb leaching. It was found that the formation and growth of gehlenite (SiO 2 ·2CaO·Al 2 O 3 ) in the ashes led to the decrease in the amount of Pb leaching. Therefore it was considered that the addition of an inorganic matrix with a high silica content that can promote gehlenite growth in the ash, for example, coal ash or waste glass is effective to suppress of Pb leaching.