Sickle cell disease (SCD) can be cured by bone marrow transplantation (BMT); however because of the morbidity and mortality associated with the procedure, BMT has been used only as a last resort for the most severely affected patients. In order for BMT and/or gene therapy to become a realistic curative therapeutic option for most sickle cell patients, it is imperative to develop less toxic preparative regimens, increase the number of available donors, develop gene therapy/transplantation protocols in which genetically manipulated autologous cells can be used, and determine the proportion of mixed chimerism required to prevent the sequelae of SCD. Umbilical cord blood (CB) has been used as an alternative source of stem cells for BMT and may provide a unique source of stem cells for gene therapy because of its easy and safe availability from individuals prenatally diagnosed with genetic disorders such as SCD. The goal is to provide effective, safe, and minimally toxic BMT and/or gene therapy for SCD patients. The hypothesis to be tested is: CB transplantation can provide stable mixed chimerism in non-ablated or minimally ablated recipients.
The specific aims are to utilize a near-term fetal/newborn mouse blood (FNPB) model of CB transplantation which we and others have developed to: 1) determine the minimum number of FNPB cells which can provide long-term full engraftment or mixed chimerism in non-ablated and minimally ablated recipients; 2) transplant transgenic mice that are models of human SCD with normal FNPB stem/progenitor cells (S/PC) and to determine the proportion of donor:recipient chimerism required to ameliorate or """"""""cure"""""""" sickle cell sequelae in these recipients; 3) use retroviral vectors containing human gamma- and beta-globin genes to transduce FNPB S/PC capable of long term engraftment and to transplant these transduced cells into non-ablated and/or minimally ablated normal and sickle cell mice. These studies are designed to define the characteristics of FNPB S/PC with regard to frequency, stem cell cycle kinetics, """"""""stemness,"""""""" and gene transducibility. BMT, including competitive repopulation assay, short-term and long-term liquid cultures, and methylcellulose cultures will be employed. These """"""""pre-translational"""""""" studies form the basis for developing effective human CB transplantation and autologous CB gene therapy protocols.
