Cell therapy for transient hematopoietic recovery

先前在犬模型中的研究表明，及时给予单一细胞因子G-CSF可以拯救犬免受400cGy全身照射（TBI），否则这是致命剂量。这种保护作用在500cgy时消失。同样，不能长期移植的mhc错配的骨髓细胞也提供了保护作用，允许450 cGy TBI后的自体重建。在这种情况下，最大耐受辐射剂量尚不清楚，保护机制也不清楚。假设，不匹配的细胞可以提供一个短暂的血细胞群，减轻细胞减少，直到发生自体重建。另外,

Previous studies in the canine model have shown that a single cytokine, G-CSF, given in a timely manner can rescue dogs from 400cGy total body irradiation (TBI), an otherwise lethal dose. This protective effect is lost at 500 cGy. Similarly, MHC-mismatched marrow cells that do not engraft long-term also provide a protective effect that allows for autologous reconstitution following 450 cGy TBI. In this case, the maximum tolerated radiation dose is not known, nor is the protective mechanism. Hypothetically, the mismatched cells could provide a transient population of blood cells that would mitigate the cytopenia until autologous reconstitution occurs. Alternatively, they could, through alloreactivity, trigger a 'cytokine storm' that mimics the injection of G-CSF. If the former proves true, combining growth factors with a non-engrafting cell product might have additive protective effects, increasing the radiation dose threshold from which victims can be rescued. Therefore, the proposed studies are designed to determine the maximum radiation dose at which survival, with autologous hematopoietic reconstitution, can be achieved by infusing a non-matched population of hematopoietic cells, with and without added growth factors. Studies will use the canine irradiation model that has been highly predictive for outcomes in human patients. Three Specific Aims are proposed: Aim 1; given the assumption that the "optimal" cell product consists primarily of committed myeloid progenitors, small-scale methods for ex vivo generation of these cells will be developed. This includes development of in vitro assays for quality control of expanded cell products and optimization of cryopreservation techniques. The optimal ex vivo expansion protocol will then be scaled-up using Good Manufacturing Practice (GMP) guidelines. In Aim 2, the optimal cell product as defined under Aim 1 will be used to determine the maximum radiation dose that allows for autologous recovery when transient hematopoietic support is provided. Donor cells will be distinguished from autologous cells using VNTR analysis. It will then be determined whether immunosuppressive regimens are needed to prevent host-versus-graft reactions that might otherwise neutralize the "rescue effect". In Aim 3, cytokines will be combined with the myeloid progenitor cell product aimed at optimizing the "rescue effect". The cytokines to be tested in addition to G-CSF will be identified through separate support (RO1 AI66498-01). The "window of opportunity" after radiation exposure, during which infusion of the progenitor cell product is effective, will also be determined. The overall goal of this project is to provide 'off-the-shelf' products that can facilitate autologous reconstitution or at least provide a period of support while an appropriate stem cell source (addressed in Projects 5 and 6) is identified.