We investigated sirolimus and mycophenolate mofetil (MMF) as graft-versus-host disease (GVHD) prophylaxis in patients with advanced hematologic malignancies receiving myeloablative hematopoietic cell transplantation (HCT) from HLA-identical sibling donors. Based on pre-study stopping rules, the trial was closed to accrual after enrollment of 11 adult patients. Seven of the 11 patients received busulfan-containing preparative regimens. Sirolimus was discontinued in 3 patients due to toxicity-related events of severe sinusoidal obstructive syndrome, portal vein thrombosis, altered mental status and in 1 patient due to risk of poor wound healing. Six of the 11 patients developed grade II-IV acute GVHD (AGVHD) a median of 15.5 days post-HCT. Two of 3 patients with grade IV AGVHD had sirolimus discontinued by 9 days post-HCT. All patients responded to AGVHD therapy without GVHD-related deaths. There were 2 nonrelapse- and 2 relapse-related deaths. At a median follow-up of 38 months (2–47 months), 7 of 11 patients were alive without disease. MMF and sirolimus GVHD prophylaxis did not reduce the risk of acute GVHD, however, there were no GVHD-related deaths. The severe toxicities in the patients receiving the busulfan-containing preparative regimens limited the continued use of sirolimus and MMF for the prevention of AGVHD.