<h3>Background</h3> The combination of Clofarabine + Fludarabine + Busulfan (CloFluBu) was found to have synergistic anti-leukemic activity against ALL and AML blasts in vitro (Andersson et al: BBMT 2011). As TBI induces significant late effects in childhood ALL, and AML patients have high relapse rates, we hypothesized that CloFluBu may be a potential alternative to TBI in ALL, and could add anti-leukemic activity in AML. Within the "Dutch COG HCT Working Group" we prospectively studied the outcomes of a CloFluBu-conditioning regimen for lymphoblastic and myeloid malignancies. <h3>Methods</h3> Patients from the 2 pediatric HCT programs (LUMC and UMC Utrecht/Princess Máxima Center for Pediatric Oncology) in the Netherlands with a lymphoblastic or myeloid malignancy receiving their first HCT, between August 2011 and April 2019, were included. Over 4 days, Clofarabine 30mg/m2 was given in 1 hour, followed by Fludarabine 10mg/m2 in 1 hour, followed by a 3-hour infusion of once-daily targeted Busulfan (weight-based dosing + therapeutic drug monitoring to a total Bu-exposure of 90mg*h/L). Thymoglobulin was added in unrelated donors (except in AML receiving cord blood). GvHD-prophylaxis was according to standard protocols. Minimal Residual Disease (MRD) negative was defined as < 10e-4. Primary endpoints were Overall Survival (OS) and Leukemia Free Survival (LFS). Secondary endpoints were Non Relapse Mortality (NRM), Relapse, acute and chronic Graft-versus-Host Disease (GvHD), and VOD/SOS. Cox Proportional Hazard and fine and gray competing risk models were used for data analysis. <h3>Results</h3> 155 children were included; 66 ALL (38 in CR1, 28 ³ CR2), 69 AML (28 in CR1, 40 in CR2, 1 in active disease) and 20 other malignancies (mostly MDS-EB). Median age was 9.7 (0.5-18.6) years. Most donors were unrelated (119 vs 36 related); 79 Bone Marrow (BM), 66 Cord Blood (CB) and 10 Peripheral Blood Stem Cells. Median follow up was 964 (19-2994) days. Overall the 3-yr estimated OS and LFS was 72 ± 4.5% and 65 ± 5% respectively. Estimated 3-yr LFS for MRD-neg ALL, MRD-pos ALL, AML CR1 and AML CR2 was 74 ± 7%, 40 ± 12%, 64 ± 10%, 65 ± 9% respectively; Fig 1). NRM in whole cohort was 10.3 ± 3.0% (with 25 ± 4.3% for AML CR1 (n=28), and 5.8 ± 2.3% in the rest; Fig 2). Other endpoints: only 2 graft-failures were noted, incidence of aGvHD III-IV at 6 months was 11 ± 3%, extensive chronic GvHD at 3-yr was 5.2 ± 2.2%. Relapse at 3-yr was 25 ± 4.3% (MRD-neg ALL 16.2 ± 3.7%, versus 60 ± 4.9% in MRD-pos ALL, in AML 22.5 ± 4.2%) and no VOD/SOS was noted. <h3>Conclusion</h3> CloFluBu in myeloid- and lymphoblastic malignancies, showed very limited toxicity and encouraging LFS in all groups, in particular for MRD negative ALL. More studies, preferably in randomized controlled clinical trials, are needed to draw firm conclusion with regards to the anti-leukemic effect and late effects.