Phase III Randomized Trial Comparing Clofarabine, Fludarabine, and Busulfan vs. Busulfan, Cyclophosphamide, and Melphalan in Pediatric Acute Myeloid Leukemia Transplant Conditioning

The primary objective of this study is to evaluate whether a conditioning regimen consisting of one alkylator (**Busulfan**) combined with two antimetabolites (**Clofarabine** and **Fludarabine**) results in superior 2-year acute grade III to IV-free, chronic non-limited **Graft-versus-Host Disease (GvHD)**-free, relapse-free survival (GRFS) compared to a regimen combining three alkylating agents (**Busulfan, Cyclophosphamide, and Melphalan**) in pediatric patients with **Acute Myeloid Leukemia** undergoing allogeneic stem cell transplantation. This is clinically relevant as it aims to improve long-term outcomes and reduce treatment-related complications in this vulnerable population.

Secondary objectives include comparing the following outcomes between the two trial arms: - neutrophil and platelet engraftment, - rate of primary and secondary graft failure, - cumulative incidence of disease relapse, - cumulative incidence of transplant-related mortality, - disease-free and overall survival, - incidence of grade II-IV and III-IV acute GvHD, - incidence of chronic GvHD, - rates of Grade ≥ 3 toxicity according to the Common Terminology Criteria for Adverse Events (CTCAE) Version 5.0, - incidence of infections, - immunological recovery, - quality of life, - late effects, - nutritional status. Additionally, the study aims to analyze the association between pre-hematopoietic cell transplantation (HCT) Minimal Residual Disease and incidence of relapse, disease-free survival, and overall survival, as well as to estimate and analyze outcomes and associations as described above.

The clinical trial involves a total of **28 participants** diagnosed with **Acute Myeloid Leukemia**. The study population includes both male and female subjects, with an age range of up to 21 years at the time of transplantation. Participants were selected based on specific inclusion criteria, such as being in hematological remission with no evidence of extramedullary disease and having a related or unrelated donor meeting certain HLA matching criteria. The trial does not focus on a vulnerable population. Lifestyle considerations such as diet and physical activity are not specified. The selection process ensures that participants are in remission and have a suitable donor, which is crucial for the study's objective of comparing different conditioning regimens for transplantation.

The clinical trial is a **randomized**, multi-center Phase III study designed to compare two conditioning regimens in pediatric patients with **Acute Myeloid Leukemia** (AML) undergoing allogeneic stem cell transplantation. The trial aims to evaluate whether a regimen containing one alkylator (Busulfan) combined with two antimetabolites (Clofarabine and Fludarabine) results in superior 2-year acute grade III to IV-free, chronic non-limited Graft-versus-Host Disease (GvHD)-free, relapse-free survival compared to a regimen combining three alkylating agents (Busulfan, Cyclophosphamide, and Melphalan). The study is **double-blind** and controlled, ensuring unbiased results.

The trial is expected to run from January 2022 to December 2029, with participant involvement lasting up to four years, depending on the treatment regimen. Participants will undergo a series of study visits, beginning with an inclusion (screening) visit to confirm eligibility based on criteria such as age, remission status, and donor compatibility. Follow-up visits will be scheduled to monitor the primary endpoint, which is the 2-year acute grade III to IV-free, chronic non-limited GvHD-free, relapse-free survival, as well as secondary endpoints like disease-free survival, overall survival, and transplant-related mortality. The end-of-study visit will conclude the participant's involvement, assessing long-term outcomes and any late effects of the treatment.

Participants may be withdrawn from the study early if they experience significant adverse events, fail to comply with the study protocol, or withdraw consent. The trial's methodology ensures rigorous monitoring and data collection to achieve its objectives, contributing valuable insights into the optimal conditioning regimen for pediatric AML patients undergoing transplantation.

The clinical trial involves the administration of several experimental medications, each with specific pharmaceutical forms, dosages, and administration routes. **Sendoxan**, containing the active substance **cyclophosphamide**, is provided as a powder for solution for injection. It is administered via infusion with a maximum daily dose of 60 mg/kg and a total dose of 120 mg/kg over a treatment period of up to 2 days. The pharmaceutical form is a solution for injection, and the product is manufactured by Baxter Medical AB.

**Grafalon**, with the active substance **anti-T lymphocyte immunoglobulin for human use, rabbit**, is a concentrate for solution for infusion. It is administered through IV infusion with a maximum daily dose of 15 mg/kg and a total dose of 45 mg/kg over a maximum treatment period of 3 days. This product is manufactured by Neovii Biotech GmbH.

**Busulfan Fresenius Kabi** is a concentrate for solution for infusion containing the active substance **busulfan**. It is administered via IV infusion with a maximum daily dose of 5.2 mg/kg and a total dose of 90 mg/kg over a treatment period of up to 4 days. The product is manufactured by Fresenius Kabi Deutschland GmbH.

**Evoltra**, containing **clofarabine**, is a concentrate for solution for infusion. It is administered through IV infusion with a maximum daily dose of 30 mg/m² and a total dose of 120 mg/m² over a treatment period of up to 4 days. The product is manufactured by Sanofi B.V.

**Fludarabin Actavis** is a concentrate for solution for injection/infusion containing **fludarabine phosphate**. It is administered via IV infusion with a maximum daily dose of 10 mg/kg and a total dose of 40 mg/kg over a treatment period of up to 4 days. The product is manufactured by Actavis Group PTC EHF.

**Thymoglobuline**, with the active substance **antithymocyte immunoglobulin**, is a powder for solution for infusion. It is administered through IV infusion with a maximum daily dose of 2.5 mg/kg and a total dose of 10 mg/kg over a treatment period of up to 4 days. The product is manufactured by Genzyme Europe B.V.

**Alkeran**, containing **melphalan**, is a solution for injection. It is administered via IV infusion with a maximum daily and total dose of 140 mg/m² over a treatment period of 1 day. The product is manufactured by Aspen Pharma Trading Limited.

Participant compliance with the dosing schedules is monitored throughout the trial to ensure adherence to the treatment protocols. No non-experimental treatments, such as standard-of-care therapy, placebo, or comparator treatments, are specified in this trial. The trial aims to evaluate the efficacy of these medications in children with Acute Myeloid Leukemia undergoing allogeneic stem cell transplantation.

The efficacy of the clinical trial will be assessed using a combination of primary and secondary endpoints. The primary endpoint is the 2-year acute grade III to IV-free, chronic non-limited **Graft-versus-Host Disease (GvHD)**-free, relapse-free survival. This will be evaluated by monitoring the absence of acute GvHD grades III to IV, chronic non-limited GvHD, relapse, or death, up to the last follow-up. Secondary endpoints include Disease-Free Survival (DFS), Overall Survival (OS), Cumulative Incidence of Relapse (CIR), Transplant-related Mortality (TRM), Hematologic Recovery, Graft Failure (GF), Immune Reconstitution, Chronic GvHD, Infections, Toxicity, Transplant-associated hormonal and gonadal late effects, and Nutritional status.

The trial will compare two conditioning regimens in children with Acute Myeloid Leukemia undergoing allogeneic stem cell transplantation. The efficacy parameters will be collected and analyzed at specified intervals throughout the study duration, with the final analysis occurring at the end of the trial. The trial is designed to determine if a conditioning regimen containing one alkylator (Busulfan) combined with two antimetabolites (Clofarabine and Fludarabine) results in superior outcomes compared to a regimen combining three alkylating agents (Busulfan, Cyclophosphamide, and Melphalan).