Creation of Chimerism in a Thalassemic Mouse Model

Abstract 882 Hematology-Oncology I Platform, Saturday, 5/1

Treatments for genetic erythrocytic diseases such as sickle cell anemia and Cooley's anemia have been limited to transfusion and treatment of symptoms of either the disease or the iron build-up associated with transfusions. Recently, BMT has been shown to provide a cure for certain patients. However, concerns over toxicity of conditioning regimens, inability to predict clinical severity, identification of HLA-matched donors, and consequences of GVHD have raised concerns and limited this approach. Current protocols, based on those developed for the treatment of malignant marrow disease, assume that complete ablation is necessary. In a few scattered cases, chimeras resulted with the survival of both host and graft precursor cells. In these cases, clinical benefits have been consistently noted. Recently, significant success was reported in development of transplantation protocols in mice which do not require complete ablation and which are less toxic. Stable mixed chimeras have been formed even with incompatible donor marrow. We have investigated the use of low dose radiation host treatment (100 cGy) and high levels of stem cells (40 million male BALB/c cells) for the nontoxic creation of allochimerism. We utilized an H2 mismatched murine transplantation model, B6.SJL to BALB/c, for these studies. The concepts involved are 1)low dose (100 cGy) host treatment avoids the "cytokine storm" which probably plays a major role in GVHD and graft acceptance, while nontoxically but dramatically reducing the number of host stem cells; 2)high levels of donor stem cells (40 × 10⁶) enhance donor:host stem cell ratios and thus increase the final % donor phenotype and may act to "tolerize." We applied this approach utilizing the Hbbth¹/Hbbth¹ thalassemic mouse, which is on a B6 background. We transplanted 3 sets of mice: 5 female and 3 male Hbbth¹/Hbbth¹ with sex-matched B6.SJL marrow following donor cells with the CD45.1 MAB and cell sorter MoFlo analysis. We treated all Hbbth¹/Hbbth¹ mice with 100 cGy whole body irradiation. In 3 male Hbbth¹/Hbbth¹ and 2 female Hbbth¹/Hbbth¹ we infused 40 million B6.SJL and in 3 female Hbbth¹/Hbbth¹, we infused 100 million marrow cells. Chimerism was detected in the marrow and ranged from 18% to 44%. We have now mixed chimerism using Ektacytometry and cell density profiles of peripheral blood. RBC survival studies demonstrate normal survival of the red cells of donor origin in contrast to shortened survival of thalassemic cells. Complete hematologic correction was noted in all chimeric animals. Our results indicate that engraftment can occur in this model, although it is quite variable, and that we can create long-term, high level chimerism with a 40 million cell dose. Furthermore, at this level of chimerism the anemia and other phenotypic manifestations of thalassemia were corrected. We believe application of this approach to humans is warranted.