NONMYELOABLATIVE BMT FOR SICKLE CELL DISEASE

Washington University, Saint Louis MO

Investigators

Abstract

DESCRIPTION (adapted from application abstract): Allogeneic bone marrow transplantation (BMT) is the only definitive cure for sickle cell disease. However, the high morbidity and mortality associated with conventional BMT has precluded its use for the overwhelming majority of patients with sickle cell anemia. The long-term goal of this study is to determine whether a less toxic therapeutic approach, non-myeloablative BMT, might be a feasible alternative to conventional BMT for the treatment of sickle cell disease. To achieve this goal, two specific aims are proposed. In Specific Aim 1, a murine model system of non-myeloablative bone marrow transplantation will be developed with the goal of establishing stable red cell chimerism in mice homozygous for a mutation similar to the human beta globin S mutation. The applicant has created a novel mouse model of sickle cell disease by knocking in an alanine- to-isoleucine mutation into codon 6 of the mouse beta globin major gene. This mutation, designated beta-6I (B6I) encodes a beta globin molecule that is predicted to produce a polymerizing form of hemoglobin in homozygous mice. The applicant will use B6I/B6I mice as transplant recipients in non-myeloablative BMT experiments. In both the syngeneic and allogeneic transplant settings, the applicant will investigate factors such as conditioning, stem cell dose, and graft composition in the establishment and maintenance of mixed donor-host hematopoietic chimerism. A real-time PCR assay system will be used to precisely monitor mixed chimerism in peripheral red blood cells, the relevant compartment for the treatment of sickle cell disease. In Specific Aim 2, the applicant will create a unique donor mouse to complement the recipient B6I mouse in non-myeloablative bone marrow transplantation experiments. A Green Fluorescent Protein (GFP) cassette will be inserted into the mouse beta minor locus by gene targeting. Virtually all red cells from this mouse should express GFP, allowing direct quantification by flow cytometry in nonmyeloablated BMT recipients. This system will provide a more clinically realistic cellular view of alloengraftment in comparison to the RNA detection system used initially. Ultimately, the applicant will attempt to correlate levels of red cell chimerism with clinical outcome in the mice. This work will be performed in the laboratory of Dr. Tim Ley, within the Division of Bone Marrow Transplantation and Stem Cell Biology at the Washington University School of Medicine.