Extramedullary disease (EMD) is a rare presentation in Acute Myeloid Leukaemia (AML). On behalf of PDWP/EBMT we studied the effect of EMD compared to isolated BM involvement in the outcomes following non-TBI conditioning hematopoietic stem cell transplantation (HCT) for AML in children. Patients were grouped into three categories: BM involvement only (Group A), BM + CNS involvement only (Group B) and BM+ other EMD+/- CNS (Group C). A total of 958 patients were included, 685 (71.5%) in Group A, 135 (14.1%) in Group B and 138 (14.4%) in Group C. The median post HCT follow-up was 5.4 years. Five years LFS, OS, RI and NRM were 62%, 68.4%, 26.4% and 11.6%, respectively. Multivariate analysis showed higher RI in Group C compared to Group A (HR = 1.45 (1.01-2.06) p = 0.04). In the multivariate analysis patients with EMD had no significant difference in LFS, OS and NRM. Our findings suggest that children with AML and BM with EMD+/- CNS involvement (group C) have a higher incidence of relapse after HCT compared to those with BM only or BM + CNS only disease. However, the presence of EMD did not have an impact on overall, leukaemia-free survival or non-relapse mortality.

ABSTRACT:Chronic granulomatous disease (CGD) is an inborn error of immunity characterized by defective NAD phosphate oxidase function, leading to impaired microbial killing, recurrent infections, and granulomatous inflammation. Allogeneic hematopoietic stem cell transplantation (HSCT) is a curative treatment for CGD, particularly effective when a fully HLA-matched donor is available. However, the place of HLA-haploidentical HSCT remains less established. This retrospective multicenter study analyzed outcomes of 64 patients with CGD (53 males; 46 with X-linked CGD) who underwent a first HSCT with HLA-haploidentical family donors, with either in vitro T-cell receptor (TCR)αβ/CD19 depletion or in vivo depletion using posttransplant cyclophosphamide (PTCY). The mean age at transplant was 5.8 years (range, 0-33). Patients exhibited a high disease burden before HSCT, with 45% experiencing infections in the 6 months before HSCT and 67% exhibiting inflammation. Outcomes in the entire cohort showed a 3-year overall survival, event-free survival (EFS), and grade 3 to 4 graft-versus-host disease (GVHD)-free EFS of 75.9%, 70.2%, and 56.1%, respectively, and were not affected by the type of depletion or age. The cumulative incidence (CI) of primary graft failure (PGF) was 20.6%. The CI of grade 2 to 4 acute GVHD was higher in the PTCY group (P = .04), whereas the CI of grade 3 to 4 GVHD was not. These results indicate that HLA-haploidentical HSCT is a feasible transplant option for patients with CGD lacking HLA-matched donors. Further refinement of transplant protocols is necessary to mitigate graft failure and acute GVHD, ultimately improving access and outcomes for this life-saving therapy.

Evolution of acute myeloid leukaemia (AML) treatments and transplantation procedures may affect outcomes after second haematopoietic stem cell transplantation (HSCT2) for relapsed paediatric AML. We analysed 345 paediatric patients reported to the European Society for Bone Marrow Transplantation (EBMT) registry for HSCT2 performed for AML relapse post-HSCT between 2000 and 2022. Multivariable analyses were adjusted for sex, age, transplant period, donor, disease status pre-HSCT2, cytogenetics, conditioning, total body irradiation (TBI) and post-first haematopoietic stem cell transplantation (HSCT1) remission duration. At three years leukaemia-free survival (LFS), overall survival (OS), non-relapse mortality (NRM), relapse incidence (RI) and graft-versus-host disease (GVHD)/relapse-free survival (GRFS) were 30.2%, 37.5%, 19.1%, 50.7% and 20.7% respectively. Compared with the 2000-2013 period, HSCT2 performed in 2014-2022 had better LFS (hazard ration [HR]: 0.66, 95% confidence interval [95% CI]: 0.48-0.90; 3-year: 34.3% vs. 26.3%), OS (HR: 0.60, 95% CI: 0.42-0.84; 3-year: 42.9% vs. 32.8%), RI (HR: 0.66, 95% CI: 0.46-0.98; 3-year: 46.0% vs. 54.7%) and GRFS (HR: 0.65, 95% CI: 0.48-0.90; 3-year: 25.3% vs. 16.1%) while NRM and GVHD incidence were stable. Relapse >6 months post-HSCT1 and remission pre-HSCT2 were associated with better LFS, OS and RI. Conditioning and cytogenetics did not influence outcomes. Mismatched unrelated donor negatively affected OS. These results highlight the improving survival after HSCT2 and support it in selected patients, particularly those relapsing later and in remission at HSCT2.

BACKGROUND: AML SCT-BFM 2007 was a prospective, on site monitored, and standardized hematopoietic stem cell transplantation trial for patients with pediatric acute myeloid leukemia (AML). In this trial the graft was the investigational product and therefore needed approval by the Paul-Ehrlich-Institute (PEI). Here we report the outcomes of transplants with matched unrelated donors (MUDs) in comparison to matched sibling donors (MSDs). METHODS: Bone marrow as graft source was utilized in 90, peripheral blood stem cells in 50 patients. Children and adolescents with high-risk de novo AML or relapsed AML were eligible for enrolment. Between May 2010 and February 2016, 140 children (age < 12 years, n=91) and adolescents (age > 12, years, n=49) were recruited in Germany, Austria and the Czech Republic. A matched donor was either a MSD or an at least 9/10 HLA-matched MUD. Subjects with AML in CR1 (n=45) or CR2 (n=47) were conditioned with a myeloablative regiment consisting of Busulfan (age adjusted i.v. dosing: 3.2 - 4,8 mg/kg BW on days -7 through -4), Cyclophosphamide (60mg/kg i.v. on days -3 and -2), and Melphalan (140 mg/m2 i.v. on day -1) (BuCyMel). Graft-versus-Host disease (GvHD) prophylaxis was cyclosporine (CSA) and short-term methotrexate. Patients with MUD received anti-T-lymphocyte globuline at a dose of 20 mg/kg/day on days -3 through -1) in addition. Patients with poor response to AML induction therapy (n=48) were stratified to receive a reduced-intensity regimen consisting of a cytoreductive block with Fludarabine (30mg/m2/d i.v.), Amsacrine (100 mg/m2/d i.v.), and Cytarabine (2g/m2/d i.v.) (FLAMSA) on days -12 through -9, immediately followed by 4 Gy TBI on day -5 and Cyclophosphamide (60 (unrelated)/40 (related) mg/kg/day i.v.) on days -4 through -3. After early taper of MMF and CSA, three prophylactic donor lymphocyte infusions were scheduled on days 120, 150, and 180. ATLG was added for MUDs (20 mg/kg x d) and MSDs (10 mg/kg x d) on days -4 through -2. RESULTS: Of the 140 patients (68 female, 72 male), 99 received a graft from a MUD (10/10 n=70, 9/10 n=29), 32 from a MSD. Data on ethnicity were not collected. Nine patients without a matched donor were eligible for Haplo-HCT and excluded from this analysis. Overall survival (OS) at 4 years was 65% (SE 5%) for patients transplanted from MUD and not different from that for MDS transplants with 62% (SE 9%). Similarly, there was not difference for MUD and MSD transplants in the 4-years cumulative incidence rates (CI) of aGvHD II-IV with 33% (SE 5%) versus 28% (SE 8%), cGVHD 12% (SE 3%) versus 7% (SE 7%), relapse 28% (SE 5%) versus 34% (SE 9%), and TRM 14% (SE 4%) versus 16% (SE 6%), respectively. Patients in CR1 or CR2 conditioned with BuCyMel had a similar OS after MUD and MDS transplants of 76% (SE 5%) versus 62% (SE 10%), and similar CI rates of aGVHD II-IV 33% (SE 16%) versus 30% (SE 6%), relapse 21% (SE 5%) versus 29% (SE 10%) and TRM 12% (SE 5%) versus 17% (SE 5%). OS after FLAMSA-RIC was 33% (SE 10%) versus 63% (SE 17%) (p=0.17), and CIs of aGvHD II-IV 31% (SE 9%) versus 25% (SE 17%), relapse 46% (SE 10%) versus 50% (SE 20%), and TRM 20% (SE 8%) versus 0% (SE 0%) (p=0.18) in MUD and MSD transplants, respectively. CONCLUSION: With improving HLA-typing technology, OS and CI of chronic and acute GvHD II-IV are identical after MUD- and MSD-transplantation for pediatric AML, if ATLG is added for MUDs at a dose of 20 mg/kg x day on days -4 through -2. This result may have major ethical implications for the current use of minor sibling donors.

Background: Relapse after a first allogeneic hematopoietic stem cell transplant (HCT1) remains the leading cause of treatment failure in children with acute myeloid leukaemia (AML). A second allogeneic stem cell transplant (HCT2) is offered to a proportion of patients failing the first allograft. It is potentially curative but carries a high risk of relapse and toxicity. The latest extensive retrospective studies, published around ten years ago, noted a prolonged overall survival rate of around 30%. Recent developments in AML management and transplantation procedures may have impacted the results of HCT2. Methods: Through the EBMT registry, we retrospectively evaluated transplant outcomes of patients under 18 at the time of HCT2 with relapsed AML after HCT1 who then underwent HCT2 between 2000 and 2022. The primary outcome was Leukaemia-Free Survival (LFS). To assess changes in outcomes over time, we categorised patients, according to the median year of HCT2, into two periods: 2000-2013 and 2014-2022. Results: A total of 345 patients were identified, with 168 receiving HCT2 from 2000 to 2013 and 177 since 2014. The median age at HCT2 was 10.4 years (Q1: 5.2; Q3: 14.5). The median time between HCT1 and relapse was 10.8 months (Q1: 6.2; Q3: 17.8). Concerning disease status at HCT2, 265 patients (76.8%) were in complete remission. Donor types included 77 matched related donors (22.4%), 86 mismatched related donors (25%) and 181 unrelated donors (52.6%). At HCT2, 73.7% of patients had a different donor than their HCT1. The stem cell sources were bone marrow (n=112, 32.6%), cord blood (n=57, 16.6%), and peripheral blood (n=175, 50.9%). The conditioning regimen was myeloablative (MAC) in 75.2% of HCT2, total body irradiation was used in 34.4%, Busulfan in 23.1%, and Treosulfan in 18.8% of transplants. The median follow-up after HCT2 was 4.9 years (CI95%: 4.4-6). At 3 years, leukaemia-free survival (LFS), overall survival (OS), non-relapse mortality (NRM), cumulative relapse incidence (RI) and GVHD-free, relapse-free survival (GRFS) were 30.2% (CI95%: 24.9-35.5), 37.5% (CI95%: 31.8-43.2), 19.1 (CI95%: 14.9-23.7), 50.7% (CI95%: 44.9-56.2) and 20.7% (CI95%: 16.1-25.6) respectively. The incidence of grade II-IV acute GVHD (aGVHD) was 34.8% (CI95%: 29.5-40.1) and of grade III-IV 13.2% (CI95%: 9.7-17.2). Three-year extensive chronic GVHD was 11.9% (CI95%: 8.4-16.2). The recent period (2014-2022), compared to the earlier period (2000-2013) for the time of HCT2, was significantly associated with improved LFS (3-year probability: 34.3% vs 26.3%; HR 0.65, p=0.007), OS (3-year probability: 42.9% vs 32.8%; HR 0.60, p=0.003), RI (3-year incidence: 46% vs 54.7%; HR 0.66, p=0.029) and GRFS (3-year probability: 25.2% vs. 16.1%; HR 0.65, p=0.007), NRM (3-year incidence: 19.7% vs 19%; HR 0.59, p=0.095) and GVHD related outcomes did not differ significantly between the two periods. Time from HCT1 to relapse (> 6 months) was a favourable prognostic factor for LFS (HR 0.61, p=0.004), OS (HR 0.66, p=0.024), RI (HR 0.60, p=0.01) and GRFS (HR 0.61, p=0.004). Active disease at HCT2 was a significant risk factor for LFS (HR 1.71, p=0.003), OS (HR 1.63, p=0.01), RI (HR 1.87, p=0.002) and GRFS (HR 1.71, p=0.003). Additionally, poor-risk cytogenetics was a significant risk factor for OS (HR 1.50, p=0.045). Conclusion: For pediatric AML patients with relapse after HCT1, HCT2 is a suitable option able to rescue a proportion of patients, especially if the time elapsing between HCT1 and relapse is longer than 6 months and if a new complete remission was achieved. Still, relapse remains a significant obstacle in this setting. Interestingly, overall survival improved over time, linked to a substantial reduction in relapse incidence.

Summary The survival after childhood cancer has improved substantially, therefore the population of childhood cancer survivors is increasing. This growing population of childhood cancer survivors, however, is at risk of a spectrum of adverse health outcomes. Unfortunately, until now, there was a lack of comprehensive follow-up recommendations.The purpuse of this article is to provide information on recently developed harmonized evidence-based guidelines and the structure to provide complex long term follow up for childhood cancer survivors. We pointed out the need for a multidisciplinary pediatric and adult specialist team, who together develop multidisciplinary long-term follow-up clinics.

Background Anti-CD19 chimeric antigen receptor T-cells (CART) have been incorporated into the therapeutic landscape of B-acute lymphoblastic leukemia (B-ALL) and B-non-Hodgkin's lymphoma (B-NHL). The manufacturing process of commercially available autologous CART in patients relapsing after allogeneic hematopoietic cell transplant (allo-HCT) might include T-cells of donor origin. In this setting, there is limited data on graft-versus-host disease (GvHD) as an off-target effect in patients treated with CART after allo-HCT. We hereby report on a large, retrospective, EBMT registry-based study on GvHD in patients treated with CART therapy after allo-HCT. Methods Inclusion criteria were B-ALL and B-NHL adult and pediatric allo-HCT patients, treated with a first anti-CD19 CART (axicabtagene ciloleucel [axi-cel] and tisagenlecleucel [tisa-cel]) from 2018 to August 2022. The primary study endpoints were the cumulative incidences (CI) of new acute GvHD (aGvHD) and chronic GvHD (cGvHD). Secondary endpoints were the 1-year GvHD relapse-free survival (GRFS), non-relapse mortality (NRM), and overall survival (OS). Overall data was analyzed in a descriptive manner. Results A total of 257 allo-HCT patients treated with anti-CD19 CART were included. One hundred seventy-two patients (66.9%) were ≥18 years old. Tisa-cel was the therapy of choice in 184 patients (71.6%), whereas axi-cel was used in 73 patients (28.4%). More than half of the cohort (57.6%) underwent allo-HCT from unrelated donors. Notably, 109 patients (46.6%) and 38 patients (15%) had previously develop aGvHD and cGvHD between allo-HCT and CART infusion. Table 1 describes data on baseline patient, allo-HCT and CART characteristics of the whole cohort and of patients developing aGvHD and cGvHD. In total, 3 patients developed new aGvHD and 6 patients developed new cGvHD after CART therapy. The 100-day CI of new aGvHD was 1.6% (95% CI, 0.4-4.2) and the 12-month CI of new cGvHD was 2.8% (95% CI, 1.1-5.7). No GvHD was observed in the pediatric cohort. The median time from allo-HCT to CART infusion was 15.8 months (range, 3.8-220.3) and the median times from CART to the development of aGvHD and cGvHD were respectively 44 days (range, 8-81) and 144 days (range, 5-182). The 1-year GRFS and NRM were 52.1% (95% CI 45.6-59.4) and 4.7% (95% CI 2.5-8.1) respectively (Figure 1). With a median follow up of 18.8 months (95% CI, 16.2-23.7), the 1-year OS was 76.8% (95% CI 71.5-82.4). The most frequent cause of death was disease-related in 56 patients (74.6%). Conclusion Together, in this large cohort including adult and pediatric allo-HCT patients treated with tisa-cel and axi-cel after allo-HCT, these data show that the incidence of both aGvHD and cGvHD is very low. Of note, GvHD was not observed in the pediatric cohort. Furthermore, no excessive NRM was observed compared to previously published methods of treating relapse post allo-HCT, being disease progression a major hurdle to optimal outcomes in this cohort of patients.

Abstract Langerhans cell histiocytosis (LCH) is a rare neoplasm predominantly affecting children. It occupies a hybrid position between cancers and inflammatory diseases, which makes it an attractive model for studying cancer development. To explore the molecular mechanisms underlying the pathophysiology of LCH and its characteristic clinical heterogeneity, we investigated the transcriptomic and epigenomic diversity in primary LCH lesions. Using single-cell RNA sequencing, we identified multiple recurrent types of LCH cells within these biopsies, including putative LCH progenitor cells and several subsets of differentiated LCH cells. We confirmed the presence of proliferative LCH cells in all analyzed biopsies using IHC, and we defined an epigenomic and gene-regulatory basis of the different LCH-cell subsets by chromatin-accessibility profiling. In summary, our single-cell analysis of LCH uncovered an unexpected degree of cellular, transcriptomic, and epigenomic heterogeneity among LCH cells, indicative of complex developmental hierarchies in LCH lesions. Significance: This study sketches a molecular portrait of LCH lesions by combining single-cell transcriptomics with epigenome profiling. We uncovered extensive cellular heterogeneity, explained in part by an intrinsic developmental hierarchy of LCH cells. Our findings provide new insights and hypotheses for advancing LCH research and a starting point for personalizing therapy. See related commentary by Gruber et al., p. 1343. This article is highlighted in the In This Issue feature, p. 1325

While survival rates in paediatric acute lymphoblastic leukaemia (ALL) nowadays exceed 90%, systemic ALL relapse, especially after haemopoietic stem cell transplantation (HSCT), is associated with a poor outcome. As there is currently no standardized treatment for this situation, individualized treatment is often pursued. Exemplified by two clinical scenarios, the aim of this article is to highlight the challenge for treating physicians to find a customized treatment strategy integrating the role of conventional chemotherapy, immunotherapeutic approaches and second allogeneic HSCT. Case 1 describes a 2-year-old girl with an early isolated bone marrow relapse of an infant KMT2A-rearranged B-cell precursor ALL after allogeneic HSCT. After bridging chemotherapy and lymphodepleting chemotherapy, chimeric antigen receptor (CAR) T-cells (tisagenlecleucel) were administered for remission induction, followed by a second HSCT from the 9/10 human leukocyte antigen (HLA)-matched mother. Case 2 describes a 16-year-old girl with a late, isolated bone marrow relapse of B-cell precursor ALL after allogeneic HSCT who experienced severe treatment toxicities including stage IV renal insufficiency. After dose-reduced bridging chemotherapy, CAR T-cells (tisagenlecleucel) were administered for remission induction despite a CD19- clone without prior lymphodepletion due to enhanced persisting toxicity. This was followed by a second allogeneic HSCT from the haploidentical mother. While patient 2 relapsed around Day + 180 after the second HSCT, patient 1 is still in complete remission >360 days after the second HSCT. Both cases demonstrate the challenges associated with systemic ALL relapse after first allogeneic HSCT, including chemotherapy-resistant disease and persisting organ damage inflicted by previous therapy. Immunotherapeutic approaches, such as CAR T-cells, can induce remission and enable a second allogeneic HSCT. However, optimal therapy for systemic ALL relapse after first HSCT remains to be defined.

Background Extramedullary disease (EMD), including central nervous system (CNS) and myeloid sarcoma, is a rare presentation in Acute Myeloid Leukemia (AML). We studied the effect of EMD (CNS involvement only and other EMD ± CNS) compared to isolated BM involvement in the outcomes following hematopoietic stem cell transplantation for AML in children. Methods An EBMT registry-based retrospective study to assess the impact of extramedullary disease (EMD) in children (de novo AML, age <18y at transplant, period 2008-2016) with AML who underwent a first allogeneic hematopoietic cell transplant (allo-HCT) with non-TBI conditioning regimen. Outcomes of interest included: leukemia-free survival (LFS), overall survival (OS), relapse incidence (RI) and non-relapse mortality (NRM). Patients were grouped into three categories: BM involvement only (Group A), BM+CNS involvement only (Group B) and BM+ other EMD ± CNS (Group C). Patients with Down syndrome related AML and t(15:17) AML were excluded. Results A total of 958 (529 males) patients met study criteria, including 685 (71.5%) Group A, 135 (14.1%) Group B and 138 (14.4%) Group C patients. Group C patients were transplanted at a younger age (median age at transplant 4.8y vs 9.2y and 6.6y for Group A, B respectively, p <0.001). A higher proportion of Group A patients were in CR1 (70.8%) compared to Group B (63.7%) and C (60.1%) patients. The three groups were comparable regarding the distribution of high and standard risk cytogenetics. Fifty seven percent of the patients received an unrelated donor and 35% a matched sibling/family donor. The median post HCT follow-up was 5.4 years (IQR: 5.2-5.7y). Five years LFS, OS, RI and NRM were 62%, 68.4%, 26.4% and 11.6%, respectively in the entire cohort. Multivariate analysis showed higher RI in Group C compared to Group A (HR= 1.45 (1.01-2.06) p=0.04) and a non-significantly different NRM (HR: 0.61 (0.29-1.29), p=0.2). There was no difference in RI in Group B compared to Group A (Hazard Ratio (HR) : 1.12 (0.76-1.64). In the multivariate analysis patients with EMD (CNS only or EMD+/- CNS) had no significant difference in LFS, OS and NRM. The Hazard ratio of LFS, OS and NRM for Group B and C compared to Group A was: HR=1.13, p= 0.46; HR=0.96, p=0.82; HR=1.09, p=0.76 and HR=1.19, p=0.28; HR=0.96, p = 0.82; HR=0.61, p=0.2, respectively. Further 116 patients underwent hematopoietic stem cell transplantation with active disease. Eighty-nine patients had BM involvement only (group A), 12 patients had BM and CNS disease (Group B) and 15 patients had BM and other sites of EMD +/- CNS (Group C). In this cohort, the 4y-OS was 37% in Group A, 25% in Group B of and 38% in Group C. Four-year LFS was 31%, 25% and 39% in Group A, Groups B and Group C respectively. Conclusions Our findings suggest that children with AML and BM with EMD ± CNS involvement (group C) have a higher incidence of relapse after HCT compared to those with BM only or BM+CNS only disease. However, the presence of EMD with CNS involvement or other extramedullary disease did not have a statistically significant impact on overall survival, leukemia-free survival or non-relapse mortality. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal

The impact of donor (D) and recipient (R) CMV serology on transplant outcomes in children with malignant hematological diseases was analyzed in a large registry-based study. Children below 18 years (y) reported to European Society for Blood and Marrow Transplantation (EBMT) who received an unmanipulated transplant from a matched sibling (MSD) or an unrelated donor (UD) (10/10, 9/10 HLA match) with bone marrow (BM) or peripheral blood (PB) between 2007 and 2020 were included. Patients receiving haploidentical grafts, cord blood and ex vivo T-cell depletion were excluded. Impact of D and R CMV serology on overall survival (OS), progression free survival (PFS), relapse incidence (RI) and non-relapse mortality (NRM) was evaluated in 4 categories (D-/R-, D-/R+, D+/R-, D+/R+), within MSD, UD10/10 and UD9/10 groups. Variables used for adjustment were source of cells, age at transplant, female to male transplant, year of transplant, disease risk index, and total body irradiation use. There were 7033 patients with a median follow-up of 3.5 yrs. In total 3407, 2599 and 1027 patients received MSD, UD10/10 and UD9/10 transplants, respectively (Table I). In multivariable Cox models (Table II), MSD with the combinations D+/R+ and D-/R+ had significantly decreased OS (HR, 1.22; p=0.03 and HR, 1.26; p=0.04, respectively) and increased NRM (HR, 1.63; p=0.008 and HR, 2.01; p=<0.001, respectively) compared to D-/R-. The PFS was significantly decreased in D-/R+ (HR, 1.22; p=0.04) compared to D-/R-. For D+/R-, RI was significantly lower (HR 0.71; p= 0.03) without significant increase of NRM (HR 1.5. p=0.15) and non-significantly different PFS (HR 0.83, p=0.17) but RI was not significantly different for D+/R+ and D-/R+ compared to D-/R- (HR 1.06; p=0.47 and HR 1.08; p=0.5, respectively). UD10/10, D-/R+ had a significantly decreased OS (HR, 1.55; p=<0.001), decreased PFS (HR, 1.37; p=<0.001), increased NRM (HR, 2.03; p=<0.001) compared to D-/R- group. In UD9/10, there was no significant difference in OS between groups. Significantly increased NRM (HR, 1.6; p=0.03) was observed in D-/R+ group. Our data show that CMV serology remains to have a substantial impact on survival in both MSD and UD10/10 transplants in children transplanted for malignant hematological diseases. For MSD, the D-/R+ and D+/R+ combinations have decreased OS and increased NRM compared to D-/R- and donor serology is not of significant impact on OS and PFS in both R+ and R- patients. RI was significantly lower in D+/R- in MSD which may be due to the possible "virus versus leukemia” effect as suggested by previous studies. However, this did not translate into improved PFS. For UD10/10, only D-/R+ (but not D+/R+ or D+/R-) have decreased OS and increased NRM compared to D-/R- transplants. This observation is clearly different from previous registry based studies focusing on adults where D+/R- transplants were also reported to have decreased OS compared to D-/R-. Our study reveals that for UD10/10 transplants selection of serocompatible donor is pivotal in R+ patients for transplants in children with malignant hematological diseases. In conclusion, this large retrospective trial shows that CMV serology represents a substantial determinant of survival in both MSD and UD10/10 transplants in children transplanted for a malignant hematological disease. While selecting donor according to CMV serology has no impact on survival for MSD transplants, selection of a serocompatible donor for a seropositive patient significantly improves overall survival in the unrelated donor setting, which represents a clinically impactful finding. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal

Background: The national Austrian CAR-T network (AT-CAR-T network) consists of 6 CAR-T cell centers and established a stringent nation-wide selection algorithm in order to ensure quality-controlled and regulated access for patients with relapsed/refractory large B-cell lymphomas (LBCL) in line with the EMA-label. Patients were selected according to this algorithm and therapy was approved by local tumor-boards. The algorithm allocates candidates to 3 categories: i) eligible, ii) to be discussed and iii) non-eligible for CAR-T treatment based on 5 major (cardiac and lung function, ECOG performance status, active CNS involvement and current infections) and 7 minor criteria. We here present our first nation-wide outcome analysis assessing the efficacy of the selection process using retrospective real-life data collected over the last three years including efficacy and toxicity data. Patients and methods: Between September 2019 and February 2022, 45 patients were selected strictly based on the selection algorithm including an ECOG <2 and treated with one of the two approved compounds (tisagenlecleucel/ Kymriah® or axicabtagen ciloleucel/ Yescarta®). All patients have signed IC before being treated with these approved products. The analysis included patient characteristics, efficacy and adverse event data. Patients treated with "out of specification" products were also documented. Results: At the time of this analysis, 45 r/r LBCL (n=44 DLBCL-NOS thereof n=13 HGL/ DHL (double hit lymphoma) or THL (triple hit lymphoma), n=1 PMBCL), n=6 pediatric ALL (pALL) patients. IPI was <3 in most patients and >3 in n=6 LBCL patients. Twenty-three patients received tisagenlecleucel and twenty-two patients axicabtagen ciloleucel. Additionally, 6 pALL patients received tisagenlecleucel, but were not selected by the Austrian Selection Algorithm. Therefore, analysis focuses on mainly the LBCL pts. The median number of previous treatment lines was 3 (range 2-7), including autologous transplant (n=15; 33.3%). Bridging therapy was administered in 39 pts (86.7%) consisting of radiotherapy (RT, n=6), combined immunochemotherapy with various regimens mainly Pola-RB (n=9), but also some with R-GeMOX (n=2), R-ICE (n=1), R-GDP (n=3), R-DHAP (n=1), dexamethasone monotherapy (n=3), bispecific antibodies like glofitamab (n=3), small molecules alone or in combination with obinotuzumab, or amongst others ibrutinib (n=2), idelalisib (n=1), lenalidomide (n=3), venetoclax (n=2). PET-CT-scan based response assessment at month 3 after CAR T-infusion was available for n=38 LBCL patients (ORR=62,2%: CR=53,3%, PR=8,9%; SD=2,2%; PD=20%). The median follow-up period for patients censored at last observation was 9.6 months (interquartile range: 5.2-20.2). The 6mo PFS and OS is 74% [95%CI:60-87%] and 87% [95%CI:76-98%] and 12mo PFS and OS is 70% [95%CI:55-84%] and 78% [95%CI:62-93%], respectively. The median PFS and OS calculated from the time of CAR-T administration (d0) was not reached in the overall cohort. Three of six pALL patients were in CR at month 3 after CAR-T infusion, four of the pALL pts relapsed, whereas 5 are still alive being rescued by an allogeneic transplant or a second CAR-T infusion. Side effects were well manageable in all cases. Grade 3 AE like CRS occurred in 2 LBCL patients (axicabtagen ciloleucel), ICANS ≥3 in 4 LBCL cases (1 tisagenlecleucel /3 axicabtagen ciloleucel). We did not observe therapy-related mortality. Long-term neutropenia after day 28 was observed in 9 patients. Summary/Conclusion: Our results support the use of a nationwide consensus for CAR-T patient selection criteria, which serves as basis for local patient allocation. This first analysis of AT-CAR-T patients selected based on a defined algorithm demonstrates favorable outcome and is well comparable to other approaches using centralized selection criteria, e.g. with the French DESCAR-T data. Our data further support that co-morbidities and fitness of CAR-T candidates need to be acknowledged. The limitation of our data is the rather small patient cohort and lack of outcome data for individuals refused for CAR-T cell therapy based on the AT-CAR-T algorithm. Figure 1View largeDownload PPTFigure 1View largeDownload PPT Close modal

T cells engineered to express chimeric antigen receptors (CAR-T cells) have shown impressive clinical efficacy in the treatment of B cell malignancies. However, the development of CAR-T cell therapies for solid tumors is hampered by the lack of truly tumor-specific antigens and poor control over T cell activity. Here we present an avidity-controlled CAR (AvidCAR) platform with inducible and logic control functions. The key is the combination of (i) an improved CAR design which enables controlled CAR dimerization and (ii) a significant reduction of antigen-binding affinities to introduce dependence on bivalent interaction, i.e. avidity. The potential and versatility of the AvidCAR platform is exemplified by designing ON-switch CARs, which can be regulated with a clinically applied drug, and AND-gate CARs specifically recognizing combinations of two antigens. Thus, we expect that AvidCARs will be a highly valuable platform for the development of controllable CAR therapies with improved tumor specificity.

Molecular ON-switches in which a chemical compound induces protein–protein interactions can allow cellular function to be controlled with small molecules. ON-switches based on clinically applicable compounds and human proteins would greatly facilitate their therapeutic use. Here, we developed an ON-switch system in which the human retinol binding protein 4 (hRBP4) of the lipocalin family interacts with engineered hRBP4 binders in a small molecule-dependent manner. Two different protein scaffolds were engineered to bind to hRBP4 when loaded with the orally available small molecule A1120. The crystal structure of an assembled ON-switch shows that the engineered binder specifically recognizes the conformational changes induced by A1120 in two loop regions of hRBP4. We demonstrate that this conformation-specific ON-switch is highly dependent on the presence of A1120, as demonstrated by an ∼500-fold increase in affinity upon addition of the small molecule drug. Furthermore, the ON-switch successfully regulated the activity of primary human CAR T cells in vitro. We anticipate that lipocalin-based ON-switches have the potential to be broadly applied for the safe pharmacological control of cellular therapeutics.

Chimeric antigen receptor (CAR) T cells have proven to be a powerful cellular therapy for B cell malignancies. Massive efforts are now being undertaken to reproduce the high efficacy of CAR T cells in the treatment of other malignancies. Here, predictive preclinical model systems are important, and the current gold standard for preclinical evaluation of CAR T cells are mouse xenografts. However, mouse xenograft assays are expensive and slow. Therefore, an additional vertebrate in vivo assay would be beneficial to bridge the gap from in vitro to mouse xenografts. Here, we present a novel assay based on embryonic zebrafish xenografts to investigate CAR T cell-mediated killing of human cancer cells. Using a CD19-specific CAR and Nalm-6 leukemia cells, we show that live observation of killing of Nalm-6 cells by CAR T cells is possible in zebrafish embryos. Furthermore, we applied Fiji macros enabling automated quantification of Nalm-6 cells and CAR T cells over time. In conclusion, we provide a proof-of-principle study that embryonic zebrafish xenografts can be used to investigate CAR T cell-mediated killing of tumor cells. This assay is cost-effective, fast, and offers live imaging possibilities to directly investigate CAR T cell migration, engagement, and killing of effector cells.

Nach jahrelangem Bemühen ist es im Jahr 2014 gelungen, Kinder- und Jugendlichenrehabilitation als gemeinsame Initiative der Sozialversicherungen und der Bundesländer österreichweit vorzusehen. Nach einem 2‑stufigen EU-weiten Vergabeverfahren wurden 2017 und 2018 insgesamt 343 pädiatrische Rehabilitationsbetten in insgesamt 11 Losen an die jeweiligen Bestbieter vergeben. Mittlerweile (Stand Dezember 2019) sind 306 dieser Versorgungseinheiten in Betrieb. Der Zugang zu Rehabilitationsaufenthalten wird niederschwellig gehalten und die Antragstellung ist österreichweit einheitlich geregelt („single point of service“). Mangels entsprechender Vorerfahrung wird die Entwicklung einer entsprechenden Rehabilitationskultur noch ein wenig Zeit in Anspruch nehmen, bei weiterer Kooperation aller Beteiligten sollte dieses wichtige und bisher fehlende Versorgungssegment aber in naher Zukunft optimale Qualität bieten können. Regelmäßige Evaluierungen und ein Monitoring des Patienten- und Elternfeedbacks sollen dazu beitragen.