title: Flt3-L augments the engraftment of donor-derived bone marrow cells when combined with sublethal irradiation and costimulatory (CD28/B7 and CD40/CD40L) blockade.
abstract: T-cell costimulatory blockade as a constituent for recipient conditioning prior to bone marrow transplantation has led to the development of less toxic protocols for the establishment of donor cell chimerism. We therefore hypothesized that the addition of the hematopoietic growth factor, Flt3-ligand (Flt3-L), to the perioperative inhibition of the CD28/B7 and CD40/CD40 ligand costimulatory pathways would enhance the engraftment of allogeneic bone marrow. Recipient BALB/c ByJ (H-2(d), Mls(c), Vbeta6+/Vbeta8+ TCR) received a single sublethal dose of total body irradiation (300 rad) 6 h prior to transplantation IV with unfractionated donor CBA/J (H-2(k), Mls(d), Vbeta6-/Vbeta8+ TCR) bone marrow cells. CTLA4-Ig and/or MRI were administered at 500 microg IP on days 0, 2, 4, and 6 posttransplantation. Flt3-L was administered at 10 microg IP on days 0-6. Donor cell chimerism was determined on days 30-90 by flow cytometric analysis. Donor-specific tolerance was assessed by skin grafting. In vitro TCR cross-linking assays and flow cytometry were utilized to explore the deletion of donor-reactive T cells. Recipients receiving CTLA4-Ig and MRI engrafted allogeneic bone marrow cells in the peripheral blood (3/6; 50%) with chimerism being detected at 2-31%. Addition of Flt3-L to this preconditioning regimen enhanced the incidence of engraftment of donor bone marrow cells (10/13; 3-70%). Long-term survival of donor but not third-party-specific skin grafts demonstrated that donor-specific tolerance had been achieved in the chimeric recipients. Deletion of the donor-reactive T cells within the chimeric recipients was also observed. The addition of hematopoietic growth factors and cytokines to the nonmyeloablative regimen of sublethal irradiation and T-cell costimulatory blockade provides a novel strategy for the establishment of donor cell chimerism and for the induction of stable and robust donor-specific tolerance. The deletion of donor-reactive T cells using this protocol suggests the reliability and feasibility of this protocol for clinical transplantation.
authors.affiliation: Thomas E. Starzl Transplantation Institute, Department of Surgery, University of Pittsburgh Medical Center, PA 15261, USA. woodwardje@msx.upmc.edu
authors.forename: Jennifer E
authors.initials: JE
authors.lastname: Woodward
authors.forename: Abdus
authors.initials: A
authors.lastname: Salam
authors.forename: Alison J
authors.initials: AJ
authors.lastname: Logar
authors.forename: Adam T
authors.initials: AT
authors.lastname: Schaefer
authors.forename: Abdul S
authors.initials: AS
authors.lastname: Rao
mesh_terms: D000069594:Abatacept; D000818:Animals; D015703:Antigens, CD; D000943:Antigens, Differentiation / Q000494:pharmacology; D001854:Bone Marrow Cells / Q000166:cytology / Q000187:drug effects* / Q000528:radiation effects; D016026:Bone Marrow Transplantation / Q000379:methods*; D018106:CD28 Antigens / Q000187:drug effects / Q000378:metabolism; D015704:CD4 Antigens / Q000276:immunology; D019013:CD40 Antigens / Q000187:drug effects / Q000378:metabolism; D060908:CTLA-4 Antigen; D002455:Cell Division / Q000187:drug effects / Q000502:physiology; D004305:Dose-Response Relationship, Drug; D006085:Graft Survival / Q000187:drug effects* / Q000528:radiation effects; D006086:Graft vs Host Disease / Q000188:drug therapy / Q000503:physiopathology / Q000517:prevention & control; D015395:Histocompatibility Antigens Class I / Q000494:pharmacology; D018796:Immunoconjugates; D008297:Male; D008565:Membrane Proteins / Q000494:pharmacology*; D051379:Mice; D008807:Mice, Inbred BALB C; D008810:Mice, Inbred C57BL; D008808:Mice, Inbred CBA; D015778:Minor Histocompatibility Antigens; D011827:Radiation; D016693:Receptors, Antigen, T-Cell, alpha-beta / Q000276:immunology; D018183:Transplantation Chimera / Q000502:physiology*; D014184:Transplantation, Homologous
chemical_list: D015703:Antigens, CD; D000943:Antigens, Differentiation; D018106:CD28 Antigens; D015704:CD4 Antigens; D019013:CD40 Antigens; D060908:CTLA-4 Antigen; C556706:CTLA4 protein, human; C556707:Ctla4 protein, mouse; D015395:Histocompatibility Antigens Class I; D018796:Immunoconjugates; D008565:Membrane Proteins; D015778:Minor Histocompatibility Antigens; C490628:Mr1 protein, mouse; D016693:Receptors, Antigen, T-Cell, alpha-beta; C081278:T cell receptor Vbeta6.7a, human; C084881:flt3 ligand protein; D000069594:Abatacept