FL/GCSF/AMD3100-mobilized Hematopoietic Stem Cells Induce Mixed Chimerism With Nonmyeloablative Conditioning and Transplantation Tolerance

Abstract

Background. Mobilization of hematopoietic stem cells (HSCs) has become the preferred approach for HSC transplantation. AMD3100, a competitive inhibitor of C-X-C motif chemokine receptor-4, has been found to be a rapid mobilizing agent. The present study evaluated approaches to optimize the product collected. Methods. Mobilized peripheral blood mononuclear cells (mPBMCs) from B6 mice were transplanted to recipient BALB/c mice conditioned with ablative or nonmyeloablative approaches. Results. The optimal dose of AMD3100 was found to be 5.0 mg/kg. Optimal HSC mobilization was observed when AMD3100 (day 10) was coadministered with Flt3 ligand (FL) (days 1–10) and granulocyte colony-stimulating factor (GCSF) (days 4–10). There was a 228.8-fold increase of HSC with FL/GCSF/AMD3100 compared with AMD3100 treatment alone. When unmodified mPBMCs were transplanted into ablated allogeneic recipients, all recipients expired by day 40 from severe acute graft versus host disease (GVHD). When T cells were depleted from mPBMC, long-term survival and engraftment were achieved in majority of the recipients. When PBMC mobilized by FL/GCSF/AMD3100 were transplanted into recipients conditioned nonmyeloablatively with anti-CD154/rapamycin plus 100, 200, and 300 cGy of total body irradiation, 42.9%, 85.7%, and 100% of mice engrafted, respectively. Donor chimerism was durable, multilineage, and stable. Lymphocytes from mixed chimeras showed no response to host or donor antigens, suggesting functional bidirection T-cell tolerance in vitro. Most importantly, none of the engrafted mice exhibited clinical features of GVHD. Conclusions. FL/GCSF/AMD3100 is an efficient treatment to maximally mobilize HSC. Durable engraftment and donor-specific tolerance can be achieved with mPBMC in nonmyeloablative conditioning without GVHD.