OBJECTIVE Graft-versus-host disease (GvHD) is one of the leading causes of morbidity and mortality in patients undergoing allogeneic HSCT, and effective prevention of GvHD is critical for the success of the HSCT procedure. Calcineurin inhibitors (CNI) have been used for decades as the backbone of GvHD prophylaxis. In this study, the efficacy and safety of Cyclosporine A (CsA) and tacrolimus (TCR) were compared in pediatric HSCT for thalassemia. METHODS This is a retrospective analysis of 129 pediatric patients who underwent HSCT with the diagnosis of thalassemia at Medicalpark Göztepe and Antalya Hospitals between January 2017 and December 2020. RESULTS Despite the GvHD prophylaxis, grade II-IV acute GvHD developed in 29 patients. Of these patients, 12 had only gut, 10 had only skin, 6 had combined gut and skin, and one had only liver GvHD. Fifteen of these 29 patients were in the CsA group, and 14 of them were in the TCR group. There was no significant difference between the groups in terms of acute GvHD occurrence, GvHD stage, or involvement sites. In terms of CNI-related toxicity, neurotoxicity in 15 (CsA n = 9, TCR n = 6) and nephrotoxicity in 18 (CsA n = 4, TCR n = 14) patients were observed. While there was no difference between the two groups in terms of neurotoxicity, more nephrotoxicity developed in patients using TCR (p = .013). There was no significant difference between the groups in terms of engraftment syndrome, veno-occlusive disease, CMV reactivation, PRES, or graft rejection. CONCLUSIONS Regarding GvHD, there was no difference in efficacy between TCR and CsA usage. Patients taking TCR experienced noticeably higher nephrotoxicity in terms of adverse effects. This difference should be considered according to the patient's clinical situation while choosing a CNI.