Allogeneic Stem Cell Transplantation in CML With Partial T Cell Depletion and Preemptive Donor Lymphocyte Infusion.

Patients were conditioned with oral busulfan 12mg/kg (days -6 to -4), cyclophosphamide 120mg/kg (days -3,-2), rabbit antithymocytic globulin, (Fresenius, Bad Hamburg, Germany) 25mg/kg (days -5 to -1) and fludarabine 200 mg/kg (days -7 to-3). Final busulfan dose was individually determined based on measurements of serum busulfan levels with a target dose of 850-1400 microM x minute.

Transplants were performed in reverse isolation rooms equipped with high-efficiency particulate air filtration systems (HEPA). No post-transplant GvHD prophylaxis was given. Post-transplant infection prophylaxis consisted of acyclovir, itraconazole, trimethoprim-sulfamethoxazole and penicillin VK. Cytomegalovirus (CMV) status was determined weekly using PCR for CMV-DNA and pp65 antigenemia in blood leukocytes, followed by preemptive ganciclovir administration when positive.

Donors Donors were human leukocyte antigen (HLA) A,B,C serologically matched and DR and DQ molecularly matched siblings. Donor stem cells were collected following mobilization with 10 µg/kg/day G-CSF, given subcutaneously for 5 consecutive days. CD34 cells were positively selected using anti-CD34 antibody conjugated to iron-dextran microbeads using CliniMACS device (Miltenyi Biotech, Bergisch Gladbach, Germany) with an aim to collect > 5.0 x 106 CD34 cells/kg.

Disease monitoring Following transplant, all patients were under close surveillance for the presence of minimal residual disease (MRD) using cytogenetic analysis and PCR for the detection of BCR/ABL transcripts. Bone marrow and peripheral blood samples were examined every 3 months in the first year post transplant and every 3-6 months in the subsequent years.

PCR method: RQ-PCR was performed according to the Europe Against Cancer (EAC) protocol.19 The BCR-ABL and ABL copy numbers were calculated by comparing with the standard curve generated using IPSOGEN FusionQuant Standards. The results of quantifying BCR-ABL transcripts were expressed as percentage ratios relative to total ABL transcripts.