Although allogeneic hematopoietic stem cell transplantation has long been the only curative therapy for nonmalignant blood diseases, recent clinical successes have demonstrated the transformative potential of autologous gene therapies as definitive treatments for these disorders. By providing durable life-changing clinical benefit while reducing the need for suitable allogeneic donors and avoiding immunologic complications like graft rejection and graft-versus-host disease, autologous transplantation with genetically modified hematopoietic stem cells provides a potentially safer and, in some respects, more accessible pathway to cure. However, several formidable barriers still prevent broader use of autologous gene therapy, including the risk of adverse acute and chronic effects from the busulfan-based conditioning regimens used to deplete host hematopoietic stem cells before transplant. In this article, we define the role transplant conditioning regimens play in autologous gene therapy, including contrasting the goals of care in this setting with those for stem cell transplantation for malignancies. We also discuss the conditioning strategies that are used by approved and select investigational autologous gene therapies to date, while highlighting exciting preclinical and clinical advances aimed at making transplant conditioning safer for myriad nonmalignant and malignant disease applications.

Adult T-cell leukemia/lymphoma (ATL) is associated with a dismal prognosis in aggressive subtypes, with a median overall survival (OS) of less than one year. In Japan, first line chemotherapy followed by allogeneic stem cell transplantation (allo-SCT) is recommended for eligible patients, yet this strategy is scarcely reported elsewhere, especially among ethnic minorities. We prospectively evaluated the feasibility and outcome of allo-SCT in this setting. Between 2016 and 2024, 80 ATL patients were enrolled; 74 (median age 50 years) received frontline chemotherapy (CHO(E)P regimen +/-Brentuximab Vedotin in CD30 positive cases). Of these, 39 subsequently proceeded to allo-SCT using a Thiotepa-Busulfan-Fludarabine conditioning regimen after at least one line of chemotherapy. Two-year OS from first line therapy was 45 %. Time-dependent analysis demonstrated a significant survival benefit for allo-SCT (hazard ratio, 0.32). Uncontrolled disease was the primary reason for transplant ineligibility. At transplantation, 27 patients were in complete remission (CR,) 5 in partial response (PR) and 7 had stable/progressive disease. Donor sources included haploidentical (56%), matched sibling (26%), and unrelated donors (18%). Three-year OS and progression-free survival were both 52%, while one-year non-relapse mortality and cumulative relapse incidence were 15% and 35%, respectively. Absence of CR/PR at allo-SCT was the main risk factor for poor outcomes. Tailored HTLV-1/ATL molecular and flow-cytometric monitoring of assessable patients in remission, revealed that tumor clones became undetectable after allo-SCT, suggesting a potent graft-versus-ATL effect. Overall, our findings support the benefit of allo-SCT in eligible ATL patients. Clinical Trial Identifier: NCT05237245.

Aldehyde dehydrogenase 2 (ALDH2) is a key mitochondrial enzyme that detoxifies reactive aldehydes including formaldehyde, thereby mitigating their genotoxic effects. While ALDH2 deficiency exacerbates bone marrow (BM) failure in Fanconi anemia models, its role under exogenous genotoxic stress is less clear. In this study, we used Aldh2[-/-] mice to investigate how ALDH2 deficiency impacts hematopoiesis following treatment with busulfan (BSF), sublethal total body irradiation (TBI), or a combined (BSF+TBI) regimen. Under BSF or TBI monotherapy, Aldh2[-/-] mice exhibited moderate increases in DNA damage, measured by γH2AX expression in hematopoietic stem and progenitor cells (HSPCs), with a mild decline in BM cell engraftment, compared to Aldh2[+/+] controls. However, BSF+TBI treatment triggered more severe HSPC DNA damage confirmed by concordant γH2AX and 53BP1 elevation, and a sharper decline in mature hematopoietic cells. Notably, this dual stress significantly upregulated DNA repair and metabolic adaptation genes, including Ccno, Pold4, Taf1c, Npr2, and Aldh18a1. These results suggest that BSF+TBI treatment drives advanced DNA damage and compensatory gene expression shifts in the absence of ALDH2. We conclude that the impact of ALDH2 deficiency on hematopoiesis is dependent on the severity of exogenous stress. TEASER ABSTRACT: The ALDH2 rs671 loss-of-function polymorphism is carried by 30-40% of East Asian individuals, yet its hematopoietic consequences under clinical genotoxic stress remain poorly understood. Using Aldh2[-/-] mice, we demonstrate that ALDH2 deficiency increases DNA damage in hematopoietic stem and progenitor cells (HSPCs) following busulfan (BSF) or total body irradiation (TBI) monotherapy, without altering mature hematopoietic cell populations or long-term repopulating capacity. However, combined BSF+TBI treatment unmasks a clear, stress-severity-dependent phenotype: Aldh2[-/-] mice exhibit significantly greater HSPC apoptosis and DNA damage confirmed by concordant γH2AX and 53BP1 elevation, and more severe peripheral blood cytopenias. Transcriptomic analysis reveals compensatory upregulation of DNA repair pathways and Aldh18a1, suggesting a metabolic adaptation to mitigate genotoxic burden. These findings establish ALDH2 deficiency as a latent hematopoietic risk factor with direct implications for personalizing conditioning regimens in ALDH2 rs671 carriers undergoing hematopoietic stem cell transplantation.

Hematopoietic stem cell transplantation (HSCT) represents a curative therapeutic modality for children, adolescents, and young-adult (CAYA) patients with various life-threatening hematological malignancies and non- malignant haematological disorders. Conditioning regimens, a cornerstone of HSCT, typically involve total body irradiation (TBI) or busulfan (Bu)-based myeloablation. However, the optimal conditioning strategy for CAYA populations remains controversial, and high-quality evidence regarding long-term complications is lacking. This systematic review and meta-analysis aimed to comprehensively compare the efficacy and safety profiles of TBI and Bu-based conditioning regimens in CAYA-HSCT recipients.

Busulfan (BUS), a chemotherapeutic agent used to treat chronic myeloid leukemia, is associated with hepatotoxicity and nephrotoxicity. This study used transmission electron microscope (TEM) to characterize BUS-induced ultrastructural alterations in the liver and kidney of rats, and to evaluate the protective effects of quercetin (QCT), bradykinin potentiating peptide (BPP) and their combination. To our knowledge, these ultrastructural changes have not been previously reported. Rats received two BUS doses (10 mg/kg b.w.) at a 21-day interval. BUS caused marked hepatic alterations, including mitochondrial swelling with loss of cristae, rough endoplasmic reticulum (RER) dilatation, sinusoidal endothelial cell damage and increased collagen synthesis. Renal changes included thrombotic microangiopathy affecting the glomerular capillary, thickened glomerular basement membranes, reactive changes of podocyte and parietal epithelial cells, mesangial cell proliferation, Bowman's space dilatation, and increased periglomerular collagen deposition. Concomitant treatment with QCT markedly reduced most hepatic alterations, whereas the BUS+BPP group exhibited comparatively limited protection. The combined QCT+BPP treatment produced the greatest hepatic protection. In the kidney, co-treatment with QCT partially ameliorated BUS-induced damage, while BUS+BPP and BUS+QCT+BPP restored ultrastructural features to near normal. In conclusions, these findings demonstrate that BUS induces marked ultrastructural injury in the liver and kidney, QCT, BPP, particularly in combination, provide substantial protection against these alterations.

Autologous haematopoietic stem cell gene therapy (HSC-GT) is an emerging therapy for treatment-refractory neurodegenerative lysosomal storage disorders (LSD). Mucopolysaccharidosis IIIA (MPSIIIA) is a devastating LSD where pathogenic variants in the SGSH gene lead to toxic heparan sulfate accumulation, inflammation and cell death. Children present with developmental delay, neurodegeneration, significant behavioural disturbance, and die in late teens. We report 80-month outcomes for a patient with severe MPSIIIA following treatment with investigational autologous HSC-GT (OTL-201), at 30-months of age. Haematopoietic stem and progenitor cells (HSPC) were collected from peripheral blood following mobilisation with G-CSF and Plerixafor. HSPC were transduced with a lentiviral vector containing the SGSH gene and the CD11b promoter. The product was infused following myeloablative conditioning with busulfan. Product engraftment was rapid, with production of sustained supra-physiological SGSH enzyme measured in leukocytes, plasma and cerebrospinal fluid (CSF), and reductions in substrate heparan sulfate to within the normal range in urine, plasma and CSF. Neurocognitive outcomes on the Bayley Scale of Infant Development (BSID-III) and parental-reported outcomes showed improvements compared to natural history. This demonstrates sustained long-term efficacy and safety of HSC-GT in a patient with severe MPSIIIA, setting the scene for a phase I/II clinical trial in younger patients.

Background/Objectives: Busulfan is a cornerstone conditioning agent for pediatric hematopoietic stem cell transplantation (HSCT), yet its narrow therapeutic index and substantial pharmacokinetic (PK) variability complicate dosing. This review synthesizes current evidence on pediatric busulfan PK, exposure-guided dosing, and the transition from conventional therapeutic drug monitoring (TDM) to model-informed precision dosing (MIPD). Methods: We conducted a focused narrative search of PubMed, Web of Science, and ScienceDirect from database inception to August 2026 and synthesized relevant literature on pediatric busulfan PK, exposure-response relationships, TDM, population PK modeling, Bayesian methods, and emerging quantitative approaches. Results: Body size is the primary determinant of busulfan clearance in children, while maturation, underlying disease, concomitant medications, and pharmacogenetic variation may contribute additional variability, although their effects are inconsistent across populations. Busulfan exposure is associated with HSCT outcomes and toxicity; however, reported targets differ by dosing regimen, conditioning intensity, disease category, and exposure metric. Reliable exposure estimation requires accurate sampling records and validated bioanalysis, while limited sampling strategies may improve clinical feasibility. Integrating population PK models with Bayesian estimation enables individualized dose adjustment, although routine implementation requires external validation and compatibility with local workflows. Emerging approaches, including physiologically based PK, PK/PD, and machine learning models, show promise but lack sufficient clinical validation for routine use. Conclusions: TDM remains the foundation of pediatric busulfan precision dosing, while MIPD provides a framework for integrating patient characteristics, measured concentrations, and validated models. Standardized workflows, population-appropriate exposure targets, prospective validation, and evidence of improved clinical outcomes are essential for broader implementation.

BACKGROUND: Chemoimmunotherapy (ChIt) has been proven to be an effective treatment for patients with refractory high-risk neuroblastoma (HR-NBL). The objective was to assess the appearance of veno-occlusive disease/sinusoidal obstruction syndrome (VOD/SOS) after high-dose chemotherapy (HDC) and autologous stem cell transplant (ASCT) in patients previously treated with ChIt for refractory HR-NBL. METHODS: Nineteen patients were treated with ChIt based on dinutuximab beta for refractory NBL and underwent ASCT. Six patients underwent HDC with ASCT according to the SIOPEN protocol (Busulfan and Melphalan (BuMel)). Treosulfan and Melphalan (TreoMel) were used in 13 patients. The occurrence of VOD/SOS was analysed. The Mann-Whitney U test was used to analyse the correlation between VOD/SOS and potential risk factors. RESULTS: VOD/SOS developed in five out of six patients when BuMel was used after ChIt (3 pts, grade 4; 2 pts, grade 3), resulting in two deaths. No VOD/SOS was observed in patients receiving TreoMel. Other analysed factors had no influence of VOD/SOS occurrence and the only identified risk factor was BuMel administered after ChIt. CONCLUSIONS: Patients with refractory HR-NBL treated with ChIt and DB did not present life-threatening toxicities or severe liver toxicities during the ChIt cycles and continued to HDC with ASCT according to SIOPEN guidelines. The incidence of severe VOD/SOS after ChIt and subsequent BuMel was higher than expected, with severe course resulting in two deaths. Taking into account the possible increase in the risk of VOD/SOS after BuMel used after chemotherapy combined with DB, careful observation with early VOD/SOS treatment in patients after ChIt is recommended and TreoMel regimen may be considered as less toxic.

Allogeneic hematopoietic stem cell transplantation (HCT) is a curative treatment for acquired aplastic anemia (AA). More than 1000 cases of transplants for AA are performed annually in China, and transplant protocols have been continuously refined over recent years. To characterize contemporary transplant practices in China and provide useful insights for global professionals, this study analyzed the 2024 Chinese National Registry data. In 2024, 1919 transplants were reported among 1893 patients by 158 centers. Haploidentical donor (HID) was the leading donor source (57.22%). Matched sibling donor (MSD) and unrelated donor (URD) were used in 19.23% and 17.87% of cases, respectively. The G-CSF/ATG-based platform (84.42%) was the predominant transplant modality, followed by the ATG/ATLG + PTCY platform (14.17%). Low-dose busulfan was frequently used in conditioning regimens for both platforms. Due to the dominant use of HID and peripheral blood stem cell grafts, novel graft-versus-host disease (GVHD) prophylactic strategies have been explored. Based on a representative contemporary cohort of 226 patients, we report a favorable 1-year survival rate of 92.0% and a GVHD-/failure-free survival rate of 77.8%, which are comparable to reports from the EBMT and CIBMTR. Moreover, modern HCT practice supports broader consideration of transplantation for older patients, especially those aged 40-50 years. In conclusion, our report offers a comprehensive overview of the current HCT landscape in China and serves as a valuable resource for global practice.

Congenital Erythropoeitic Porphyria is extremely rare autosomal recessive disorder caused by deficiency of uroporphyrin synthase III deficiency, leading to accumulation of toxic precursors of Heme biosynthesis. Severe Phenotypes present in neonatal period with reddish brown colored urine followed by photosensitivity and erythrodontia. Allogenic hematopoietic stem cell transplant is the only curative option. We retrospectively analyzed 4 patients (2 males , 2 females) with clinically and biochemically confirmed Congenital Erythropoeitic Porphyria who underwent fully matched sibling donor hematopoietic stem cell transplant with myeloablative conditioning Busulphan (Bu) and Cyclophosphamide (Cy) at the Armed Forces Bone Marrow Transplant Centre Pakistan from 2020 to 2025. These four transplants were performed at a centre with over 550 transplants during this period, highlighting the extreme rarity of disease even within a high volume transplant centre. Median Age of onset of symptoms was five days (range 2-15), median age of diagnosis was 27 months (range12-72) and median age of hematopoietic transplant was 100.5 months (range 54-126). All patients received myeloablative conditioning (Bu[16mg/kg], Cy[160-200mg/kg]). Peri-transplant period was complicated by febrile neutropenia and mucositis. Neutrophil engraftment was achieved at a median of Day+16 (range 14-18) and platelet engraftment at median of Day+21 (range 18-21). At a median follow up of 240 days (range 116-2555), all patients were alive, transfusion independent with complete resolution of photosensitivity. Additionally, serum and urine porphyrin levels became normal in all patients. This cohort, though very small supports the efficacy of early myeloablative Hematopoietic stem cell transplant with matched sibling donors to prevent irreversible tissue damage and significantly improve clinical outcomes.

Diamond-Blackfan anemia (DBA) is a congenital bone marrow failure syndrome (CBMFS) primarily managed with corticosteroids and blood transfusions; however, allogeneic hematopoietic stem cell transplantation (allo-HSCT) remains the sole curative option when these therapies fail or relapse occurs. We report the case of a 21-year-old woman diagnosed with DBA caused by an RPS19 mutation shortly after birth. Although she responded well to corticosteroid therapy until 18 years of age, discontinuation led to relapse and subsequent transfusion dependence, necessitating allogeneic transplantation. In the absence of a human leukocyte antigen (HLA)-matched donor, she underwent post-transplantation cyclophosphamide (PT-Cy)-based haploidentical peripheral blood stem cell transplantation (haplo-PBSCT) at 21 years of age, utilizing her father as the donor. Myeloablative conditioning was performed using fludarabine, melphalan, and busulfan. For acute graft-versus-host disease (GVHD) prophylaxis, PT-Cy (50 mg/kg on days 3 and 4), tacrolimus, and mycophenolate mofetil were administered, resulting in successful neutrophil engraftment on day 16 post-transplantation. Although the patient developed acute GVHD (grade II), it resolved with corticosteroid therapy. Post-discharge, she was readmitted due to acute gastrointestinal GVHD (grade II), which improved after methylprednisolone administration. She was discharged on day 92 and achieved complete transfusion independence for the past 5 years, making this the first reported case of long-term survival following PT-Cy-based haplo-PBSCT for this condition. Including our patient, there are only four reported cases of allo-HSCT using PT-Cy for adult patients with DBA; notably, all patients survived without developing severe GVHD or infectious complications. Generally, transplant outcomes for DBA worsen with increasing age due to severe GVHD and organ damage secondary to long-term iron overload. Given that DBA is a non-malignant disease, meticulous attention must be paid to prevent unnecessary GVHD and graft failure. In adult patients with DBA, PT-Cy yielded successful engraftment and safety, with no incidence of severe GVHD, leading to long-term survival and minimal complications. These findings suggest that allo-HSCT utilizing PT-Cy remarkably expands donor options for congenital bone marrow failure syndromes. Consequently, this approach is expected to become a promising therapeutic strategy not only for hematologic malignancies but also for non-malignant disorders.