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

Xu, Hong MD1; Zhu, Ziqiang MD1; Huang, Yiming MD1; Ildstad, Suzanne T. MD1

doi: 10.1097/TP.0000000000002657
Original Basic Science—General
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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.

1 Institute for Cellular Therapeutics, University of Louisville, Louisville, KY.

Received 16 October 2018. Revision received 14 December 2018.

Accepted 30 December 2018.

The authors declare no funding to disclose.

Dr S.T.I. is the Chief Scientific Officer and Founding Scientist of Regenerex, Inc., a biotechnology company that has been formed to develop and commercialize a bone marrow product to treat numerous diseases. All other authors have no conflicts of interest to disclose.

H.X. designed experiments, performed experiments, analyzed data, and prepared manuscript for publication. Z.Z. performed experiments, analyzed data, and prepared manuscript. Y.H. edited manuscript. S.T.I. designed experiments, interpreted data, manuscript preparation, and reviewed all data and manuscript.

Correspondence: Suzanne T. Ildstad, MD, Institute for Cellular Therapeutics, University of Louisville, CFO, Regenerex, Inc., 570 S. Preston St, Suite 404, Louisville, KY 40202. (suzanne.ildstad@louisville.edu).

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