ABSTRACT:Hypodiploid and BCR::ABL1+ B-cell precursor acute lymphoblastic leukemia (BCP-ALL) confers a high risk of disease relapse. We investigated post-hematopoietic stem cell transplantation (HSCT) outcomes within the prospective FORUM trial, comparing outcomes among these genetic subgroups with those of patients without these lesions. The use of pre- and post-HSCT add-on treatments, including tyrosine kinase inhibitors (TKI) and immunotherapies, was also assessed. Multivariate analysis evaluated associations with overall survival (OS), event-free survival (EFS), and cumulative incidence of relapse (CIR). The FORUM trial enrolled 741 patients aged ≥4 years with BCP-ALL who underwent HSCT from HLA-matched donors (2013-2023). The 3-year OS and EFS did not differ significantly between patients with BCR::ABL1 fusion, hypodiploidy, and neither of these 2 genetic lesions. However, patients with hypodiploid BCP-ALL in second complete remission (CR2) showed inferior OS and EFS, driven by higher nonrelapse mortality (NRM), which occurred exclusively in near-diploid cases. No NRM occurred in severe hypodiploid cases conditioned with total body irradiation. Minimal residual disease (MRD) positivity at transplant predicted worse OS, EFS, and CIR in all genetic groups. Patients with hypodiploid BCP-ALL were difficult to salvage after relapse, even with chimeric antigen receptor T-cell therapy. By contrast, BCR::ABL1+ patients had favorable outcomes, even when MRD positive before HSCT. Prophylactic TKI use after HSCT improved EFS and reduced CIR. BCR::ABL1+ patients who received a transplant in CR2 had a 3-year OS of 96%. In conclusion, the standardized FORUM protocol yielded comparable outcomes across genetic subgroups. Posttransplant TKI maintenance improved outcomes in BCR::ABL1+ BCP-ALL. This trial was registered at www.clinicaltrials.gov as #NCT01949129 and at www.clinicaltrialsregister.eu as #EudraCT2012-0032-22.

Introduction: Kaposiform hemangioendothelioma (KHE) is a rare vascular tumor frequently complicated by Kasabach-Merritt phenomenon (KMP). Case Presentation: A 21-month-old boy presented with an initial spontaneous scalp hematoma, recurrent hemorrhagic episodes, and progressive consumptive coagulopathy. Imaging revealed an extensive peri- and paravertebral infiltrative mass involving the thoracic and lumbosacral spine, consistent with KHE. Biopsy was deferred because of high bleeding risk. Treatment with sirolimus led to rapid clinical and hematologic remission. At 3.5 years of follow-up, the patient remains free of recurrent KMP with sustained disease control and mild radiological regression. Conclusion: This case highlights the diagnostic challenges of deep-seated KHE and the pivotal role of imaging when biopsy is unsafe.

In the randomized cohort of the international phase-III FORUM trial, which showed the superiority of total-body irradiation (TBI) over chemotherapy-based conditioning prior to hematopoietic stem cell transplantation (HSCT) for pediatric acute lymphoblastic leukemia (ALL), type of conditioning and remission phase, but not pre-HSCT minimal residual disease (MRD), were associated with outcome. We report the impact of MRD within the extended FORUM cohort. Patients (n=1014), 4–21 years old, transplanted from a matched donor who had ≥1 MRD measurement prior to and/or 100 days and/or 1 year after HSCT were eligible. A threshold of 0.01% defined MRD positivity versus negativity. Prior to HSCT, 21% of patients were MRDpos. Three-year event-free survival (EFS) was 0.73 and 0.59 (p<0.001), and 3-year cumulative incidence of relapse (CIR) was 0.20 and 0.33 (p<0.001) in MRDneg and MRDpos patients, respectively. The level of MRD positivity pre-HSCT (<0.1% versus ≥0.1%), did not significantly affect outcome. Pre-HSCT MRDneg and TBI/etoposide conditioning were associated with a 2-fold lower risk of relapse, whereas MRDpos had a 2-fold higher risk of any failure and/or death. No detrimental effect of MRDpos pre-HSCT could be demonstrated in patients with T-cell ALL. MRDpos versus MRDneg patients at day 100 had an EFS of 0.47 versus 0.77 (p<0.001) and a CIR of 0.51 versus 0.17 (p<0.001), respectively, but post-HSCT MRDpos did not necessarily imply relapse. In conclusion, the MRD status pre-HSCT and at day 100 post-HSCT was a strong prognostic factor for children transplanted for ALL in the extended FORUM cohort.

Hypodiploid (Hypo) and BCR::ABL1-positive (BCR-ABL+) B-cell precursor acute lymphoblastic leukemia (BCP-ALL) confers a high risk of disease relapse. We investigated post–hematopoietic stem cell transplantation (HSCT) outcomes within the prospective FORUM trial, comparing these genetic subgroups to patients without these lesions. The use of pre- and post-HSCT add-on treatments, including tyrosine-kinase inhibitors (TKIs) and immunotherapies, was also assessed. Multivariate analysis evaluated associations with overall survival (OS), event-free survival (EFS), cumulative incidence of relapse (CIR). The FORUM trial enrolled 741 patients ≥4 years of age with BCP-ALL who underwent HSCT from HLA-matched donors (2013–2023). The 3-year OS (0.86 [95% CI, 0.76–0.92], 0.79 [0.65–0.87], and 0.79 [0.76–0.82]) and EFS (0.71 [0.59–0.80], 0.73 [0.59–0.83], and 0.67 [0.63–0.71]) did not differ significantly between BCR-ABL+, Hypo, and Neither patients, respectively. However, Hypo patients in second complete remission (CR2) showed inferior OS and EFS, driven by higher non-relapse mortality (NRM), which occurred exclusively in near-diploid cases. No NRM occurred in severe hypodiploid cases conditioned with TBI. MRD positivity at transplant predicted worse OS, EFS, and CIR in all genetic groups. Hypo patients were difficult to salvage post-relapse, even with CAR-T therapy. By contrast, BCR-ABL+ patients had favorable outcomes, even when MRD-positive prior HSCT. Prophylactic TKI use post-HSCT improved EFS and reduced CIR. BCR::ABL+ patients transplanted in CR2 had a 3-year OS of 96%. In conclusion, the standardized FORUM protocol yielded comparable outcomes across genetic subgroups. Posttransplant TKI maintenance improved outcomes in BCR-ABL+ BCP-ALL. EudraCT: 2012-0032-22; ClinicalTrials.gov: NCT01949129

Risk-based stratification approaches using measurable residual disease (MRD) successfully help to identify T-acute lymphoblastic leukemia (T-ALL) patients at risk of relapse, whose treatment outcomes are very poor. Because of T-ALL heterogeneity and rarity, a reliable and standardized approach for flow cytometry (FC)-based MRD measurement and analysis is often missing. Within the international AIEOP-BFM-ALL-FLOW study group we made a consensus on markers and a standard operating procedure for common 8- and 12-color T-ALL MRD panels. Custom manufactured tubes with dried backbone antibodies were tested in parallel to local FC standards. Altogether, 66 diagnostic and 67 day 15 samples were analyzed. We designed two guided MRD gating strategies to identify blast cells in parallel to expert-based evaluation. We proved that the optimized tubes allowed the correct identification of blast cells in all diagnostic samples. Both, expert and guided analysis of day 15 samples correlated to local standard (Spearman R=0.98 and R=0.94, respectively). Only in 2 (3 %) and 4 (6 %) patients expert gating and guided analysis results were substantially discordant from local standard, respectively. The cases that require an individualized approach may be partially identified at diagnosis through a rare immunophenotype or mixed phenotype acute leukemia status. Our work shows that standardized operating procedures together with guided analysis are applicable in a great majority of T-ALL cases. Further improvement of MRD detection is needed, as in some cases an individualized analytical approach is still required due to the challenging nature of the T-ALL phenotype.

Abstract The superiority of total body irradiation (TBI)–based vs chemotherapy conditioning for allogeneic hematopoietic stem cell transplantation (allo-HSCT) in children with acute lymphoblastic leukemia (ALL) has been established in the international, prospective phase-3 FORUM study, randomizing 417 patients aged 4-18 years in complete remission (CR), who received allo-HSCT from HLA-matched sibling or unrelated donors. Because of the unavailability of TBI in some regions and to accommodate individual contraindications, this study reports the prespecified comparison of outcomes of patients receiving busulfan (BU)– or treosulfan (TREO)–based regimens from 2013 to 2018. Overall, 180 and 128 patients received BU/thiotepa (THIO)/fludarabine (FLU) or TREO/THIO/FLU, respectively. Data were analyzed as of February 2023, with a median follow-up of 4.2 years (range, 0.3-9.1). 3-year overall survival was 0.71 (BU, 95% confidence interval [0.64-0.77]) and 0.72 (TREO, [0.63-0.79]) and 3-year event-free survival was 0.60 (BU, [0.53-0.67]) and 0.55 (TREO, [0.46-0.63]). The 3-year cumulative incidence of relapse (BU, 0.31 [0.25-0.38]; TREO, 0.36 [0.27-0.44]); and nonrelapse mortality (BU, 0.08 [0.05-0.13]; TREO, 0.09 [0.05-0.15]) were comparable. One case of fatal veno-occlusive disease occurred in each group. No significant differences in acute and chronic graft-versus-host disease (GVHD) or 3-year GVHD-free and relapse-free survival (BU, 0.48 [0.41-0.55]; TREO, 0.45 [0.37-0.54]) were recorded. Outcomes for patients in first and second CR were similar irrespective of the regimen. In conclusion, BU/THIO/FLU or TREO/THIO/FLU regimens can be an alternative to TBI for patients with ALL aged >4 years with contraindications or lack of access to TBI. This trial was registered at www.ClinicalTrials.gov as #NCT01949129.

BACKGROUND:Auer rods (AuRs) are prominent intracellular structures found almost exclusively in myeloid cell malignancies, such as acute myeloid leukemia (AML), chronic and juvenile myelomonocytic leukemia and myelodysplastic syndrome. Extremely rare AuRs have been reported in patients with acute lymphoblastic leukemia (ALL) or among ambiguous lineage leukemia patients with a dominantly lymphoblastic immunophenotype. PROCEDURE:We report diagnostic and follow-up data of an international cohort of 11 children suffering from leukemias with AuRs and with significant presence of T and myeloid markers, majority of whom categorized as early T-cell precursor (ETP, n = 7); or T-ALL (ETP status unknown, n = 2), ALAL (acute leukemia of ambiguous lineage, n = 1), and AML reclassified from ALAL (n = 1). We described other diagnostic details and treatment types and responses. Moreover, we summarize previously published data. RESULTS:Among the four patients who started and remained on ALL-type therapy, all were in the first complete remission, whereas both patients who started and remained on AML-type therapy relapsed and died. Of the patients who followed either a combined ALL/AML protocol (Interfant 06) or who switched from one of the two types of therapy to the other, one patient died, and the remaining four were in first complete remission at the most recent follow-up. We also searched for similar cases in the literature and found only three additional children with nonmyeloid leukemia and AuRs and 10 adults with this type of leukemia. CONCLUSIONS:Briefly, ALL- or combined ALL/AML-type therapy may be effective for treating AuR-positive leukemia patients with a lymphoid immunophenotype.

Deficiencia adenozín-deaminázy 2. typu (DADA2) je autoinflamačné ochorenie,

Introduction Anti T lymphocyte globulins (ATG) are effective in preventing both severe acute and chronic Graft-versus-Host Disease (GVHD) and rejection after unrelated donor (UD) hematopoietic stem cell transplantation. The two most commonly used types of ATG are rabbit polyclonal antibodies. Grafalon® (Neovii) is produced by rabbit immunization with the Jurkat T-cell line, while Thymoglobulin® (Sanofi) is made after the animal immunization with human thymocytes. Although both agents are lymphodepleting, the different formulations vary in antibody quantity, antigen recognition, and ability to eliminate or modulate the function of T, B, NK, and dendritic cells. Few studies have directly compared the efficacy and outcomes of Grafalon and Thymoglobulin. Methods Here we report the outcomes of a homogenous cohort of pediatric acute lymphoblastic leukemia (ALL) patients who received over 3 days before the allograft either Grafalon (45 mg/kg total dose (TD)) or Thymoglobulin (7.5 mg/kg, TD) in the international prospective ALL SCTped 2012 FORUM trial (NCT01949129). Patients, aged 4-18 years, were transplanted from a 9/10 or 10/10 HLA matched UD after uniform conditioning with 12 Gy total body irradiation and etoposide (60 mg/kg TD, max 3600 mg). Results From 2012 until 2023, 400 patients from 24 countries were enrolled. 303 received Grafalon (17 countries) and 97 received Thymoglobulin (15 countries). The two cohorts were comparable in terms of gender, age, ALL immunophenotype, remission status, minimal residual disease (MRD) level at transplant, donor/recipient- HLA matching (HLA 10/10 68% vs 58%, and HLA 9/10 32% vs 42%, p=0.13), stem cell source (63% vs 54% bone marrow, p=0.25) and CMV/EBV- donor/recipient IgG results. The 3-yr overall survival (OS) and event-free survival (EFS) rates were similar in both groups with 84% (+/-2) vs 77% (+/-5), (p=0.398), and 73% (+/-3) vs 74% (+/-5) (p=0.708), for Grafalon vs Thymoglobulin, respectively. The 3-yr cumulative incidence of relapse (CIR) was 17% (+/-2) for Grafalon vs 13% (+/-4) for Thymoglobulin (p=0.375). We observed no differences in acute GVHD incidence, with grade III-IV of 6% vs 10%, (p=0.176) for Grafalon and Thymoglobulin. Grafalon patients had a lower 3-year cumulative incidence of any chronic GVHD, estimated at 7% (+/-2) vs 16% (+/-4) (p=0.013). This was not associated with a significant difference in non-relapse mortality (3-yr NRM of 7% (+/-1) for Grafalon vs 13% (+/-4) for Thymoglobulin (p=0.097)). There was a trend towards a higher incidence of mild chronic GVHD in the Thymoglobulin group ((7% +/- 3) vs 3% (+/-1) p=0.05), but no difference was seen in severe chronic GVHD (3% +/-2 vs 2% +/- 1 (p=0.338)). Early grade III-IV toxicity varied between groups, with more frequent disseminated intravascular coagulation (3% vs 0%, p=0.002) in the Thymoglobulin group, but higher incidence of stomatitis, nausea, and elevated liver enzymes after Grafalon treatment. Grafalon patients had higher frequencies of hematological grade III-IV adverse events (cytopenia and hemolysis). Leukocyte engraftment was slightly delayed, with Grafalon patients reaching a WBC count of >1G/l at a median of day 22, vs day 20 for Thymoglobulin (p=0.036), while neutrophil engraftment was similar at day 28. Compared to Grafalon, more patients developed CMV reactivation/disease after Thymoglobulin exposure (14% vs 24% at day 100, p=0.031) and more Thymoglobulin-patients developed EBV reactivation/infection and post-transplant lymphoproliferative disease (PTLD) (Grafalon 17%, Thymoglobulin 38%, p <0.001; PTLD n= 1 vs 5). Subgroup analyses (CR1 and ≥ CR2) showed comparable outcomes (OS, EFS, CIR and NRM). In ≥ CR2 patients, the 3-year cumulative incidence of any chronic GVHD was lower after Grafalon with 6% (+/-2) vs 17% (+/-5) for Thymoglobulin (p=0.032). Conclusion We observed excellent outcome results following serotherapy with both Grafalon and Thymoglobulin in ALL pediatric patients transplanted from 9/10 or 10/10 allelic matched UD conditioned with TBI/etoposide. The incidence of acute GVHD did not differ significantly. Patients treated with Grafalon had lower rates of any chronic GVHD. The frequency of CMV disease after reactivation and EBV reactivation, disease and PTLD was significantly higher after Thymoglobulin. The main outcome measures - OS, EFS, and NRM did not show significant differences between the two treatment groups.

The gut microbiota of paediatric oncology patients undergoing a conditioning regimen before hematopoietic stem cell transplantation is recently considered to play role in febrile neutropenia. Disruption of commensal microbiota and evolution of opportune pathogens community carrying a plethora of antibiotic-resistance genes play crucial role. However, the impact, predictive role and association of patient´s gut resistome in the course of the therapy is still to be elucidated. We analysed gut microbiota composition and resistome of 18 paediatric oncology patients undergoing hematopoietic stem cell transplantation, including 12 patients developing febrile neutropenia, hospitalized at The Bone Marrow Transplantation Unit of the National Institute of Children´s disease in Slovak Republic and healthy individuals (n = 14). Gut microbiome of stool samples obtained in 3 time points, before hematopoietic stem cell transplantation (n = 16), one week after hematopoietic stem cell transplantation (n = 16) and four weeks after hematopoietic stem cell transplantation (n = 14) was investigated using shotgun metagenome sequencing and bioinformatical analysis. We identified significant decrease in alpha-diversity and nine antibiotic-resistance genes msr(C), dfrG, erm(T), VanHAX, erm(B), aac(6)-aph(2), aph(3)-III, ant(6)-Ia and aac(6)-Ii, one week after hematopoietic stem cell transplantation associated with febrile neutropenia. Multidrug-resistant opportune pathogens of ESKAPE, Enterococcus faecium, Staphylococcus aureus, Klebsiella pneumoniae and Escherichia coli found in the gut carried the significant subset of patient’s resistome. Over 50% of patients treated with trimethoprim/sulfamethoxazole, piperacillin/tazobactam and amikacin carried antibiotic-resistance genes to applied treatment. The alpha diversity and the resistome of gut microbiota one week after hematopoietic stem cell transplantation is relevant predictor of febrile neutropenia outcome after hematopoietic stem cell transplantation. Furthermore, the interindividual diversity of multi-drug resistant opportunistic pathogens with variable portfolios of antibiotic-resistance genes indicates necessity of preventive, personalized approach.