Bone marrow chimerism and tolerance induced by single-dose cyclophosphamide

BACKGROUND: Establishment of hematopoietic chimerism is the most stable strategy for donor-specific tolerance. Safer pretreatment regimens are needed for clinical application. We evaluated the efficacy of a simple protocol using cyclophosphamide (CYP) on induction of chimerism and organ transplant tolerance across major histocompatibility complex (MHC) barriers in the rat. MATERIALS AND METHODS: Bone marrow cells from BN (RT1(n)) donors were infused to LEW (RT1(l)) recipients on day 0 after a single injection of CYP at various doses on day -1. Donor-derived hematopoietic chimerism was evaluated by flowcytometry. The recipients received BN or third party (BUF) heart allografts on day 100. RESULTS: While pretreatment with 200 mg/kg of CYP induced high levels of hematopoietic chimerism, six of eight recipients died of severe graft-versus-host-disease (GVHD). CYP at dose of 150 mg/kg induced 36.5 +/- 24.1% of donor-derived chimerism on day 10, and sustained macrochimerism was seen until day 100 without GVHD. Pretreatment with 100 mg/kg of CYP resulted in only transient chimerism (4.8 +/- 5.2%) which disappeared by day 20. In the recipients with 50 mg/kg of CYP, donor bone marrow cells were rapidly rejected and no chimerism was observed. The recipients with 150 mg/kg of CYP accepted BN heart allografts (>100 days x 5), while rejecting BUF allografts by day 12 (n = 4). BN heart allografts were rejected in the recipients with 100 (MST: 57 days, n = 5) and 50 mg/kg (MST: 7 days, n = 5) of CYP. CONCLUSIONS: A single dose of CYP can induce hematopoietic chimerism across MHC-barriers. The dose of 150 mg/kg seems to be optimal to induce organ transplant tolerance without developing GVHD.