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CD19-directed chimeric antigen receptor (CAR) T cells have evolved as new standard-of-care (SOC) treatment in patients with relapsed/refractory large B-cell lymphoma (LBCL). Here, we report the first German real-world data on SOC CAR-T cell therapies with the aim to explore risk factors associated with outcome.

Differences in major and minor histocompatibility antigens between donor and recipient trigger powerful graft-versus-host reactions after allogeneic hematopoietic stem cell transplantation (HSCT). The clinical effects of alloreactivity present a Janus-face: detrimental graft-versus-host disease increases non-relapse mortality, beneficial graft-versus-malignancy may cure the recipient. The ultimate consequences on long-term outcome remain a matter of debate. We hypothesized that increasing donor-recipient antigen matching would decrease the negative effects, while preserving antitumor alloreactivity. We analyzed retrospectively a predefined cohort of 32 838 such patients and compared it to 59 692 patients with autologous HSCT as reference group. We found a significant and systematic decrease in non-relapse mortality with decreasing phenotypic and genotypic antigen disparity, paralleled by a stepwise increase in overall and relapse-free survival (Spearman correlation coefficients of cumulative excess event rates at 5 years 0.964; P<0.00; respectively 0.976; P<0.00). We observed this systematic stepwise effect in all main disease and disease-stage categories. The results suggest that detrimental effects of alloreactivity are additive with each step of mismatching; the beneficial effects remain preserved. Hence, if there is a choice, the best match should be donor of choice. The data support an intensified search for predictive genomic and environmental factors of ‘no-graft-versus-host disease’.

Background: Determining the onset of peripheral blood stem cell apheresis is known to be associated with several advantages. The method applied most commonly so far is the immunological analysis of cells expressing the CD34 antigen by flow cytometry. In this study a new parameter for monitoring was tested: the measurement of the so-called human progenitor cell (HPC) parameter by an automated hematology analyzer. Materials and Methods: Eleven multiple myeloma patients were included in this study. Following the white blood cell nadir, monitoring of CD34+ cells by flow cytometry and of IMI-total (immature myeloid information) and HPC counts by a hematology analyzer (Sysmex SE-9000 TM ) were performed daily. The quality of the harvest product was determined by flow-cytometric analysis. Results: Monitoring was performed on 46 days (total). Comparing the IMI-total results with the immunological CD34+ cell analyses in peripheral blood, a correlation of r = 0.609 (p < 0.05) was obtained. When comparing the CD34+ cell analysis with the HPC count, a correlation of r = 0.477 (p = 0.05) resulted. A total of 16 PBSC aphereses was performed. When comparing the preapheresis peripheral blood measurements to the CD34+ cell content of the harvest product, a correlation of r = 0.383 (p = 0.14) and of r = 0.254 (p = 0.34) was calculated for the IMI-total counts, and the HPC counts, respectively. The CD34+ pre-apheresis results showed an excellent correlation with the quality of the peripheral blood stem cell apheresis product (r = 0.937; p < 0.05). Conclusion: The gold standard for determining the timing of PBSC apheresis remains the analysis of CD34+ stem and progenitor cells by flow cytometry. The determination of the timing of an apheresis is not possible with the IMI-total or HPC results. In addition, a predictive impression of the graft quality to be expected is only possible with flow cytometry. However, the HPC counts may be applicable for the determination of the onset of CD34+ cell flow-cytometric analyses. Further tests will have to be performed to prove this finding.

Background: Long-term plateletpheresis has been suspected to cause depression in cell-mediated immunity. To prove this hypothesis, we analyzed blood samples by flow-cytometric immunophenotyping in plateletpheresis donors over a period of 26 months. Methods: Donors were included beginning with their first donation (n = 20). A non-donor control group was matched for age and sex (n = 20). Samples were taken before each apheresis and at 3-month intervals (control group: ± 2 days), and analyzed by flow cytometry. A minimum of 6 procedures per year (range 6–14) was the precondition for each donor to be included in the study. Results: No significant differences were seen within the two groups regarding the absolute white blood cell count, total lymphocyte count, relative and absolute counts of CD3+ (p = 0.82), CD4+ (p = 0.91), CD8+ (p = 0.74), CD4:CD8 ratio (p = 0.65), CD16+56+ (p = 0.54), CD25+ (p = 0.82) and CD3+/HLA-DR+ cells (p = 0.66). Conclusions: Long-term plateletpheresis does not seem to alter the absolute white blood cell count, total lymphocyte count or lymphocyte subsets in donors. Regarding the cells and subsets analyzed, plateletpheresis is a safe procedure and does not seem to have a detrimental effect on the cellular immune competence.

Background and Objectives: In this study we investigated whether platelet activation during apheresis results in the binding of platelets to white blood cells. Material and Methods: Analysis of platelet-leukocyte interaction was performed using multiparameter, three-color flow cytometry. Results: Over the duration of the procedure, there was an increase in the surface expression of CD62p (P-selectin) and CD63 (p&lt;0.05), and also in the binding of platelets to monocytes (p&lt;0.05), neutrophilic granulocytes (p&lt;0.05) and to CD3+ cells (initially to a low degree; p&lt;0.05). Platelet binding to CD19+ cells did not change significantly. Conclusion: This study demonstrates that platelets become activated during apheresis and that following this process, interaction with monocytes and neutrophilic granulocytes occurs.

BACKGROUND:The measurement of CD34+ cells is the most important step in the quality control of peripheral blood progenitor cell apheresis products. For this purpose, flow cytometry is applied. Recently, a new test kit has been introduced for the enumeration of CD34‐expressing cells, in combination with software support for semi‐automation of data acquisition and analysis. STUDY DESIGN AND METHODS:This study evaluated the ProCOUNT kit. Ninety samples obtained from peripheral blood progenitor cell apheresis products from 39 patients with hemato‐oncologic diseases were analyzed. For data acquisition and analysis, ProCOUNT software was used. Data comparison was performed with parallel measurements according to the International Society for Hematotherapy and Graft Engineering (ISHAGE) guidelines and the German reference protocol for analysis of CD34‐expressing cells. RESULTS:Correlation of the German and ISHAGE techniques was excellent (r2= 0.99). The initial correlation coefficient of ProCOUNT analysis with the German protocol was r2= 0.89. In 21 (23.3%) of 90 ProCOUNT analyses, a warning message was encountered from the ProCOUNT software. Following manual reevaluation of these data with CellQUEST software, a correlation of r2= 0.96 with the German protocol and r2= 0.97 with the ISHAGE analyses was obtained. ANOVA testing revealed significant differences between ProCOUNT and ISHAGE techniques (p<0.05) and between ProCOUNT and the German protocol (p<0.05). No statistically significant difference between ISHAGE and German protocol was observed (p = 0.19). CONCLUSION:The ProCOUNT kit and software for semi‐automated data acquisition and analysis represents a further step toward standardization of CD34 cell quantitation in peripheral blood progenitor cell apheresis products. However, the occurrence of software warnings is high, and analysis or data reevaluation by experienced staff is still mandatory. Therefore, currently there is no definite advantage of the kit and software over the existing guidelines for CD34+ analysis in peripheral blood progenitor cell grafts.

Agents capable of reversing P-glycoprotein-associated multidrug resistance have usually failed to enhance chemotherapy activity in patients with solid tumours. Based on its toxicity profile and experimental potency, dexverapamil, the R-enantiomer of verapamil, is considered to be promising for clinical use as a chemosensitizer. The purpose of this early phase II trial was to evaluate the effects of dexverapamil on epirubicin toxicity, activity and pharmacokinetics in patients with metastatic breast cancer. A two-stage design was applied. Patients first received epirubicin alone at 120 mg m(-2) i.v. over 15 min, repeated every 21 days. Patients with refractory disease continued to receive epirubicin at the same dose and schedule but supplemented with oral dexverapamil 300 mg every 6 h x 13 doses. The Gehan design was applied to the dexverapamil/epirubicin cohort of patients. Thirty-nine patients were entered on study, 25 proceeded to receive epirubicin plus dexverapamil. Dexverapamil did not increase epirubicin toxicity. The dose intensity of epirubicin was similar when used alone or with dexverapamil. In nine intrapatient comparisons, the area under the plasma concentration-time curve (AUC) of epirubicin was significantly reduced by dexverapamil (mean 2968 vs 1901 microg ml[-1] h[-1], P= 0.02). The mean trough plasma levels of dexverapamil and its major metabolite nor-dexverapamil were 1.2 and 1.5 microM respectively. The addition of dexverapamil to epirubicin induced partial responses in 4 of 23 patients evaluable for tumour response (17%, CI 5-39%, s.e.P 0.079). The remissions lasted 3, 8, 11 and 11+ months. These data suggest that the concept of enhancing chemotherapy activity by adding chemosensitizers may function not only in haematological malignancies but also in selected solid tumours. An increase in the AUC and toxicity of cytotoxic agents does not seem to be a prerequisite for chemosensitizers to enhance anti-tumour activity.

BACKGROUND: The toxicity of commonly used myeloablative regimens is the main factor limiting applicability and restricting upper age limit for allogeneic hematopoietic stem cell transplantation (alloHSCT). However, reduced intensity conditioning (RIC) protocols are associated with high risk of relapse. Previous dose-escalation trials revealed high-dose treosulfan-based conditioning an alternative treatment with myeloablative as well as antileukemic potential, accompanied by low non-hematological toxicity. The goal of this multicenter phase II trial was to evaluate safety and efficacy of treosulfan/fludarabine conditioning in AML patients. PATIENTS AND METHODS: Seventy-five patients (median age 45 years, range 19–59) with AML in 1st (80%), 2nd (17%), or 3rd (3%) complete remission were treated with alloHSCT from either matched related (MRD, 40%) or unrelated donors (MUD, 60%). Preparative regimen consisted of treosulfan 14 g/m2/day on days −6 to −4, fludarabine 30 mg/m2/day on days −6 to −2, and in case of MUD-HSCT additionally ATG (Fresenius) 10 mg/kg/day on days −4 to −2. Graft-vs.-host disease (GvHD) prophylaxis consisted of cyclosporine and short-course methotrexate. 25% of the patients had an increased risk of toxicity for standard myeloablative conditioning regimens because of higher age or co-morbidities. RESULTS: All patients engrafted after a period of absolute neutropenia with the median time to ANC >0.5 G/L and PLT >50 G/L of 21 (13–39) days and 19 (11–48) days, respectively. The cumulative incidence of complete donor chimerism was 72% on day +28 and 92% on day +100. Most frequent CTC grade 3 or 4 complications until day 28 after transplantation were febrile neutropenia (28%), infection in neutropenia (16%), and mucositis (5%). None of the patients experienced severe hepatic toxicity, including VOD. After a median follow up of 457 days (98–988), the probabilities of overall (OS) and disease-free survival (DFS) at 2 years equaled 67% resp. 58% (MRD-HSCT) and 58% resp. 45% (MUD-HSCT, p=NS). The 2-year cumulative incidence of relapse was 34% for MRD-HSCT and 37% for MUD-HSCT, whereas non-relapse mortality was 8% and 17%, respectively. Outcome was not significantly influenced by disease status (CR1 vs. ≥CR2) or age. CONCLUSIONS: Conditioning therapy based on treosulfan at a dose of 3x14 g/m2 in combination with fludarabine is very well tolerated, resulting in a low NRM. The regimen demonstrated reliable efficacy in AML patients with confirmed first or subsequent remission. Further randomized studies are warranted to verify advantageous survival results.

BACKGROUND: The toxicity of commonly used myeloablative regimens is the main factor limiting applicability and restricting upper age limit for allogeneic hematopoietic stem cell transplantation (alloHSCT). However, reduced intensity conditioning (RIC) protocols are associated with high risk of relapse. Previous dose-escalation trials revealed high-dose treosulfan-based conditioning an alternative treatment with myeloablative as well as antileukemic potential, accompanied by low non-hematological toxicity. The goal of this multicenter phase II trial was to evaluate safety and efficacy of treosulfan/fludarabine conditioning in AML patients.