Risk factors for CAR-T cell manufacturing failure among DLBCL patients: A nationwide survey in Japan.

For successful chimeric antigen receptor T (CAR-T) cell therapy, CAR-T cells must be manufactured without failure caused by suboptimal expansion. In order to determine risk factors for CAR-T cell manufacturing failure, we performed a nationwide cohort study in Japan and analysed patients with diffuse large B-cell lymphoma (DLBCL) who underwent tisagenlecleucel production. We compared clinical factors between 30 cases that failed (7.4%) with those that succeeded (n = 378). Among the failures, the proportion of patients previously treated with bendamustine (43.3% vs. 14.8%; p < 0.001) was significantly higher, and their platelet counts (12.0 vs. 17.0 × 104 /µL; p = 0.01) and CD4/CD8 T-cell ratio (0.30 vs. 0.56; p < 0.01) in peripheral blood at apheresis were significantly lower than in the successful group. Multivariate analysis revealed that repeated bendamustine use with short washout periods prior to apheresis (odds ratio [OR], 5.52; p = 0.013 for ≥6 cycles with washout period of 3-24 months; OR, 57.09; p = 0.005 for ≥3 cycles with washout period of <3 months), low platelet counts (OR, 0.495 per 105 /µL; p = 0.022) or low CD4/CD8 ratios (

[Manufacturing results of tisagenlecleucel for acute lymphoblastic leukemia: a survey by the CAR-T cell therapy taskforce of the Japan Society of Transfusion Medicine and Cell Therapy].

The frequency of the manufacturing failure of chimeric antigen receptor (CAR)-T cell therapy in clinical practice is unknown. To clarify the current state of how likely CAR-T cell production is to succeed or fail for B-cell acute lymphoblastic leukemia (B-ALL), we analyzed cases in which the production of tisagenlecleucel was performed for patients with B-ALL at 15 facilities in Japan from October 2019 to March 2022. Total 81 patients (47 males and 34 females) were analyzed. The median age at apheresis was 13 years (1-25) with a median number of prior treatments of 4 (1-9). The numbers of patients with histories of allogeneic transplantation, inotuzumab ozogamicin, or blinatumomab treatments were 51 (63.0%), 26 (32.1%), and 37 (45.7%), respectively. The median blast percentage and CD3+ cell counts in peripheral blood were 0% (0-91.5), and 611/µl (35-4,210) at apheresis, and the median number of CD3+ cells shipped was 2.2×109 (0.5-8.3). While cases with a history of heavy prior treatment before apheresis were included, no manufacturing failures were observed. Continuing to monitor the status of manufacturing failures is necessary as the number of B-ALL cases treated with CAR-T cell therapy increases.

Comparison of cryoprotectants in hematopoietic cell infusion-related adverse events.

BACKGROUND: The standard cryoprotectant for human cellular products is dimethyl sulfoxide (DMSO), which is associated with hematopoietic cell infusion-related adverse events (HCI-AEs) in hematopoietic stem cell transplantation including peripheral blood stem cell (PBSC) transplantation (PBSCT). DMSO is often used with hydroxyethyl starch (HES), which reduces DMSO concentration while maintaining the postthaw cell recovery. The cryoprotectant medium CP-1 (Kyokuto Pharmaceutical Industrial) is widely used in Japan. After mixture of a product with CP-1, DMSO and HES concentrations are 5% and 6%, respectively. However, the safety profile of CP-1 in association with HCI-AEs has not been investigated. STUDY DESIGN AND METHODS: To compare CP-1 with other cryoprotectants, we conducted a subgroup analysis of PBSCT recipients in a prospective surveillance study for HCI-AEs. Moreover, we validated the toxicity of CP-1 in 90 rats following various dose administration. RESULTS: The PBSC products cryopreserved with CP-1 (CP-1 group) and those with other cryoprotectants, mainly 10% DMSO (non-CP-1 group), were infused into 418 and 58 recipients, respectively. The rate of ≥grade 2 HCI-AEs was higher in the CP-1 group, but that of overall or ≥grade 3 HCI-AEs was not significantly different, compared to the non-CP-1 group. Similarly, after propensity score matching, ≥grade 2 HCI-AEs were more frequent in the CP-1 group, but the ≥grade 3 HCI-AE rate did not differ significantly between the groups. No significant toxicity was detected regardless of the CP-1 dose in the 90 rats. CONCLUSIONS: Infusion of a CP-1-containing PBSC product is feasible with the respect of HCI-AEs.

Simultaneous quantification of dasatinib, nilotinib, bosutinib, and ponatinib using high-performance liquid chromatography-Photodiode array detection.

BACKGROUND: Dasatinib, nilotinib, and bosutinib, second-generation tyrosine kinase inhibitors (TKIs), and ponatinib, a third-generation TKI, are approved pharmaceuticals used in the treatment of chronic myeloid leukemia (CML). Although liquid chromatography-tandem mass spectrometry assays for simultaneous quantification of the four TKIs in human serum have been reported in the literature, a high-performance liquid chromatography (HPLC) assay that simultaneously quantifies these compounds has not yet been developed. This study aims to establish and validate an efficient HPLC analytical method using a photodiode array (PDA) detector for the simultaneous quantification of the four TKIs. METHODS: Calibration standards were prepared by serial dilution of serum samples containing the four TKIs, followed by solid-phase extraction. The four TKIs were eluted in order within 10 min using a binary HPLC gradient system. RESULTS: The calibration ranges were 2-500 ng/ml for dasatinib, 100-5000 ng/ml for nilotinib, and 10-500 ng/ml for bosutinib and ponatinib. Intra-day and inter-day precision and accuracy values were found to be in accordance with the U.S. Food and Drug Administration guidelines. The recovery rates were 92.9%-96.0%, 80.7%-86.1%, 91.6%-99.0%, and 86.4%-92.6% for dasatinib, nilotinib, bosutinib, and ponatinib, respectively. CONCLUSION: To the best of our knowledge, this is the first report of an HPLC-PDA analytical method that allows efficient simultaneous quantification of the four TKIs in the serum of patients with CML. We believe that the method developed herein can improve the efficiency of therapeutic drug monitoring in patients with CML in clinical practice.

Toxoplasmosis after allogeneic hematopoietic stem cell transplantation: Impact of serostatus-based management.

Toxoplasmosis caused by Toxoplasma gondii (T. gondii) is a serious infectious complication after allogeneic hematopoietic stem cell transplantation (HSCT). The incidence of toxoplasmosis varies widely because of the variabilities of seroprevalence among patient populations. The incidence and the optimal management of toxoplasmosis after allogeneic HSCT in a patient population with a low seroprevalence have not been fully evaluated. We conducted a single-center retrospective study evaluating toxoplasmosis in Japanese patients who underwent allogeneic HSCT. Of the 728 evaluable patients, only 5 developed toxoplasmosis with a median onset of day 60 post-transplant (range, day 55-393). The cumulative incidence was 0.7% (95% CI: 0.3%-1.5%) at day 500 post-transplant. Four of the five patients succumbed due to toxoplasmosis. The more recently treated 220 patients (not the earlier 508 patients) were screened for the T. gondii serostatus, and prophylactic treatment with trimethoprim/sulfamethoxazole was applied. All five patients with toxoplasmosis were in the unscreened group, and there was no case of toxoplasmosis after the introduction of the screening and prophylactic treatment. Our results suggest that toxoplasmosis after allogeneic HST is rare but can develop as a life-threatening complication even in the populations with low seroprevalence, and that prophylactic treatment for seropositive patients could effectively prevent toxoplasmosis.

Clinical Results of a Massive Blood Transfusion Protocol for Postpartum Hemorrhage in a University Hospital in Japan: A Retrospective Study.

Background and objectives: Massive postpartum hemorrhage (PPH) is the most common cause of maternal death worldwide. A massive transfusion protocol (MTP) may be used to provide significant benefits in the management of PPH; however, only a limited number of hospitals use MTP protocol to manage massive obstetric hemorrhages, especially in Japan. This study aimed to assess the clinical outcomes in patients in whom MTP was activated in our hospital. Materials and Methods: We retrospectively reviewed the etiology of PPH, transfusion outcomes, and laboratory findings among the patients treated with MTP after delivery in our hospital. Results: MTP was applied in 24 cases (0.7% of deliveries). Among them, MTP was activated within 2 h of delivery in 15 patients (62.5%). The median estimated blood loss was 5017 mL. Additional procedures to control bleeding were performed in 19 cases, including transarterial embolization (18 cases, 75%) and hysterectomy (1 case, 4.2%). The mean number of units of red blood cells, fresh frozen plasma, and platelets were 17.9, 20.2, and 20.4 units, respectively. The correlation coefficients of any two items among red blood cells, fresh frozen plasma, platelets, blood loss, and obstetrical disseminated intravascular coagulation score ranged from 0.757 to 0.892, indicating high levels of correlation coefficients. Although prothrombin time and activated partial thromboplastin time levels were significantly higher in the <150 mg/dL fibrinogen group than in the ≥150 mg/dL fibrinogen group at the onset of PPH, the amount of blood loss and blood transfusion were comparable between the two groups. Conclusions: Our MTP provides early access to blood products for patients experiencing severe PPH and could contribute to improving maternal outcomes after resuscitation in our hospital. Our study suggests the implementation of a hospital-specific MTP protocol to improve the supply and utilization of blood products to physicians managing major obstetric hemorrhage.

[Spontaneous remission of relapsed skin lesions of ALK-positive anaplastic large cell lymphoma after autologous stem cell transplantation].

A 44-year-old woman was diagnosed with anaplastic lymphoma kinase (ALK)-positive anaplastic large cell lymphoma (ALCL) with clinical stage IVA (nodal and bladder involvement). Complete response (CR) was achieved after the CHOP chemotherapy; however, 12 months after the last course of chemotherapy, ALCL relapsed in the form of skin lesions without nodal involvement. After achieving a second CR with chemotherapy, autologous stem cell transplantation was performed. Two months after transplantation, the disease again relapsed as multiple skin lesions. Electron beam irradiation was performed; however, other skin lesions appeared thereafter and spontaneously disappeared. At present, 3.4 years after the transplantation, the patient is free from disease. ALK-positive ALCL relapsing as skin lesions may behave differently from the nodal relapse. An accumulation of cases is required to elucidate ALCL characteristics relapsing as skin lesions.

Seasonal changes in indoor airborne fungal concentration in a hematology ward.

Invasive fungal disease (IFD) is an important infectious complication of hematological disorders, especially in hematopoietic stem cell transplantation recipients. Evidences suggest seasonal and/or geographical variations in the airborne fungal counts and a relationship between those counts and the incidence of IFD. We evaluated the concentrations of indoor airborne fungi quantitated over the course of one year in a hematology ward in Japan. In January, April, July, and October, fixed volumes of air samples were obtained by an air sampler in a hematology ward not equipped with a high-efficiency particulate air filter and incubated in fugal cultures. Samples were also obtained from a protective environment in the same ward and were evaluated. The number of fungal colonies per 50 L of sampled air was highest in October (median 2.25 (range, 0.2-7.0)), which was significantly higher than those in the other three months (0.1 (range, 0-1.0) in January; 0 (0-0) in April; 0.55 (0-2.5) in July; P < 0.01)). Commonly identified pathogens included Penicillium and Cladosrporium species, but Aspergillus species was detected only in July and October samples. These results suggest a seasonal variation in indoor airborne fungal concentrations in Japan, which could affect the epidemiology of IFD.

[Two serial events of delayed hemolytic transfusion reactions due to different primary immune response-induced irregular antibodies].

A 74-year-old woman with a history of pregnancies, but without previous transfusions, received a red blood cell transfusion for aplstic anemia. She lost consciousness due to severe anemia two weeks later and was transported by ambulance to our hospital. Delayed hemolytic transfusion reaction (DHTR) was diagnosed based on the detection of anti-E antibody and positive E antigen of the previously transfused product. A transfusion of E antigen-negative red cell products was performed. However, DHTR due to anti-c antibody developed 16 d after the transfusion of a c antigen-positive product. Based on the onset of ≥14 d after the transfusions and the detection of a causative IgM-type antibody, DHTR due to a primary immune response was diagnosed. Because the incidence of DHTR is low, physicians rarely experience it in clinical practice. However, in our case, DHTR due to a primary immune response, which is even rarer in DHTR cases, developed twice within a short period. A history of transfusion and pregnancy as well as preexisting irregular antibodies have been identified as risk factors for DHTR. Thus, more attention should be paid to the risk of DHTR redevelopment by repeated transfusions.

Human Herpesvirus 6 Reactivation Evaluated by Digital Polymerase Chain Reaction and Its Association With Dynamics of CD134-Positive T Cells After Allogeneic Hematopoietic Stem Cell Transplantation.

BACKGROUND: Human herpesvirus 6 (HHV-6) causes life-threatening central nervous system disorders after allogeneic hematopoietic stem cell transplantation (allo-HSCT). Recent studies implicated CD134 as a specific receptor of HHV-6B and demonstrated that its expression levels in CD4-positive T cells after allo-HSCT could be related to the reactivation of HHV-6. We prospectively evaluated the relationship between HHV-6 reactivation and CD134+ T cells in the recipients of allo-HSCT. METHODS: HHV-6 viral load in plasma was quantitatively measured weekly after allo-HSCT by digital polymerase chain reaction in 34 patients. The ratio of CD134 in CD4+ T cells (CD134/CD4 ratio) was serially measured by flow cytometry before and after transplantation. RESULTS: HHV-6 reactivation was detected in 23 patients (68%). The CD134/CD4 ratio before conditioning was significantly higher in patients with HHV-6 reactivation than in those without (median, 3.8% vs 1.5%, P < .01). In multivariate analysis, a higher CD134/CD4 ratio before conditioning was significantly associated with the incidence of HHV-6 reactivation (odds ratio, 10.5 [95% confidence interval, 1.3-85.1], P = .03). CONCLUSIONS: A higher CD134/CD4 ratio before conditioning was associated with a higher risk of HHV-6 reactivation, suggesting that the rate may be a promising marker for predicting HHV-6 reactivation after allo-HSCT.

Impact of immunoglobulin G2 subclass level on late-onset bacterial infection after allogeneic hematopoietic stem cell transplantation.

BACKGROUND: Immunoglobulin (Ig) G2 subclass deficiency is known to be associated with recurrent bacterial respiratory infections caused by capsulated bacteria and is found mostly in pediatric patients. However, its impact after allogeneic hematopoietic stem cell transplantation (HSCT) has not been fully assessed. METHODS: We retrospectively evaluated the relationship between IgG2 subclass levels and bacterial pneumonia in 74 adult patients who survived longer than 2 years after allogeneic HSCT. RESULTS: During the evaluation period, nine patients developed bacterial pneumonia. The median IgG2 level was significantly lower in patients with an infectious episode than in those without (143 mg/dL vs 287 mg/dL; P < 0.01). In multivariate analysis, a history of rituximab therapy and cord blood as a stem cell source were significantly associated with decreased levels of both IgG2 and IgG2/IgG ratios (P < 0.05). CONCLUSIONS: Suboptimal serum IgG2 levels could increase susceptibility to late-onset bacterial pneumonia after allogeneic HSCT. IgG2 levels should be considered carefully, especially in patients receiving cord blood transplantation and/or rituximab treatment.

[Bacteriocidal effects of introducing copper products on highly touched areas in hematology ward].

Nosocomial infection via the hospital environment is a serious problem, and highly touched surfaces are the main route of transmission. Copper has been reported to possess bacteriocidal effects, and the introduction of copper-impregnated products is receiving attention as a potential component of hospital infection control. In this study, copper-impregnated door handles as highly touched areas were introduced in a hematology ward, and their bacteriocidal effects were evaluated in comparison with conventional products. All 12 samples obtained from conventional door handles were positive for bacterial cultures, whereas only 5 of 18 samples from copper-impregnated handles were positive (P<0.0001). The mean number of bacterial colonies per milliliter of sample was 300 (range: 40-1.1×106) in samples from conventional handles, but it was significantly lower in samples from copper-impregnated handles (0; range: 0-220, P<0.0001). While various types of bacteria grew on conventional handles, most of the bacteria on copper-impregnated handles were Bacillus subtilis. These results suggest that the introduction of copper-impregnated products would be useful for hospital infection control by reducing the bacterial burden on highly touched areas. However, the efficacy of this approach against spore-forming bacteria should be further investigated.

Recurrent bacterial pneumonia due to immunoglobulin G2 subclass deficiency after allogeneic hematopoietic stem cell transplantation: Efficacy of immunoglobulin replacement.

Immunoglobulin (Ig) G2 subclass deficiency is known to be associated with recurrent bacterial respiratory infections caused by capsulated bacteria. We encountered a case of recurrent pneumonia due to Streptococcus pneumoniae after allogeneic hematopoietic stem cell transplantation (HSCT). IgG2 subclass level was specifically low, and prophylactic Ig replacement successfully prevented subsequent infections. However, the cessation of Ig replacement resulted in subsequent pneumonia. These findings suggested that IgG2 deficiency could be a cause of recurrent pneumococcal infection after allogeneic HSCT.

Single-institutional retrospective analysis of Japanese patients with chronic lymphocytic leukemia.

Unlike in Western countries, chronic lymphocytic leukemia (CLL) is a rare lymphoid malignancy in Japan, and its clinical features remain to be elucidated in the Japanese population. Therefore, we retrospectively analyzed 29 Japanese CLL patients newly diagnosed at our institute. Seventeen (59%) were male, and their median age was 62 years. With a median follow-up period from diagnosis of 69 months (range, 3-170 months), 9 patients received some form of treatment for CLL. Three patients died of disease progression with or without infection (n=2) or skin cancer (n=1). Five-year overall and treatment-free survival rates were 83% (95%CI, 46-96%) and 67% (95%CI, 45-81%), respectively. Two patients received allogeneic hematopoietic stem cell transplantation for refractory disease, and both were alive without disease relapse at 53 and 110 months, respectively, after transplantation. These results suggest the clinical courses of Japanese patients with CLL to be comparable to those in Western countries. However, future studies of larger numbers of patients are needed to further elucidate the features and long-term clinical courses of CLL in the Japanese population.

High Incidence of Afebrile Bloodstream Infection Detected by Surveillance Blood Culture in Patients on Corticosteroid Therapy after Allogeneic Hematopoietic Stem Cell Transplantation.

Bloodstream infections (BSI) are still important complications after allogeneic hematopoietic stem cell transplantation (allo-SCT). Patients who are receiving corticosteroid therapy can develop BSI without fever. The utility of surveillance blood cultures in these situations is controversial. We retrospectively analyzed 74 patients who received a corticosteroid consisting of ≥.5 mg/kg prednisolone or equivalent after allo-SCT. In principle, we performed surveillance blood culture weekly for these patients. Sixteen patients (21.6%) developed definite BSI. In a multivariate analysis, a myeloablative conditioning regimen, high-risk disease status at allo-SCT, and the presence of a central venous catheter at the initiation of corticosteroid therapy were identified as independent significant risk factors for the development of definite BSI. At the first definite BSI episode, 7 patients (46.7%) were afebrile and diagnosed by surveillance blood culture. However, 6 of these 7 afebrile patients showed various signs that could be attributed to infection at the time of positive blood culture. In conclusion, patients receiving corticosteroid therapy after allo-SCT frequently develop afebrile BSI. Although surveillance blood culture might be beneficial in these situations, it also seems important to not miss the signs of BSI, even when patients are afebrile.

Meta-analysis and meta-regression analysis to compare the outcomes of chemotherapy for T- and B-lineage acute lymphoblastic leukemia (ALL): the use of dexamethasone, L-asparaginase, and/or methotrexate may improve the outcome of T-lineage ALL.

The effects of intensive regimens and the roles of drugs used might differ between T- and B-lineage acute lymphoblastic leukemia (ALL). We performed a literature search for clinical studies published from January 1998 to March 2013. Studies were eligible for inclusion in the analyses if they included more than 80 patients with adult ALL who were treated with a uniform regimen and compared T- and B-lineage ALL. Studies that included only adolescent or elderly patients were excluded. We identified 11 clinical studies, which included a total of 381 and 1366 patients with T- and B-lineage ALL, respectively, and performed meta-analyses using the selected studies. Nine studies included patients with Philadelphia chromosome-positive (Ph+) ALL. A meta-analysis using the random-effect model demonstrated superior survival in patients with T-lineage ALL compared to those with B-lineage ALL (hazard ratio 1.78, 95 % confidence interval 1.50-2.11), though the inclusion of patients with Ph+ ALL in B-lineage ALL must have influenced this result strongly. We performed meta-regression analyses, adjusted according to whether or not patients with Ph+ ALL were included in each study. Use of dexamethasone (Dex), higher dose of methotrexate (MTX), and higher dose of L-asparaginase (L-asp) were associated with a significant trend toward a better outcome in T-lineage ALL. A meta-regression analysis including Dex and the dose of L-asp or MTX together as covariates showed that these factors were independently significant. In conclusion, the use of Dex and high-dose L-asp or MTX may improve the outcome of T-lineage ALL. This hypothesis should be tested in a prospective study including only patients with Ph-negative ALL.

Significance of a positive Clostridium difficile toxin test after hematopoietic stem cell transplantation.

Patients with hematological malignancies show a high prevalence of asymptomatic colonization with Clostridium difficile (CD colonization). Therefore, it is difficult to distinguish CD colonization with diarrhea induced by a conditioning regimen from true Clostridium difficile infection (CDI) in hematopoietic stem cell transplantation (HSCT) recipients. We retrospectively analyzed 308 consecutive patients who underwent a CD toxin A/B enzyme immunoassay test for diarrhea within 100 d after HSCT from November 2007 to May 2014. Thirty patients (9.7%) had positive CD toxin results, and 11 of these had positive results in subsequent tests after an initial negative result. Allogeneic HSCT, total body irradiation, stem cell source, acute leukemia, and the duration of neutropenia were significantly correlated with positive CD toxin results. In a logistic regression model, allogeneic HSCT was identified as a significant risk factor (odds ratio 18.6, p < 0.01). In an analysis limited to within 30 d after the conditioning regimen, the duration of neutropenia was the sole risk factor (odds ratio 10.4, p < 0.01). There were no distinctive clinical features for CDI, including the onset or duration of diarrhea. In conclusion, although CDI may be overdiagnosed in HSCT recipients, it is difficult to clinically distinguish between CDI and CD colonization.