The objectives of this project are to study the stem cell compartment of tile bone marrow, its functional organization, and the implications of this organization on the clinical use of cytotoxic agents, cytokines, and radiation. This project is also directed toward improving autologous hematopoietic stem cell transplantation (HSCT) by optimizing donor marrow engraftment, and long-term recipient hematopoiesis and survival.
Specific aim 1 will determine how cytotoxic agents, radiation, and cytokines affect bone marrow stem cell reserve capacity in a murine model. Mechanisms of marrow damage will be explored by using side population (SP) cells. SP cells, isolated by flow cytometric analysis and sorting, constitute a hematopoietic stem cell (HSC) population with very high long-term repopulating ability in mice. Use of SP cells allows direct measurement of damage to progenitors and primitive stem cells, assessment of the repair capacity of stem cells, and the isolation and study of HSC in S-G2M.
Specific aim 2 will determine how cytotoxic agents and cytokines affect bone marrow stem cell reserve capacity through variations in drug dosing (using a high dose cyclophosphamide (CY) model) and through drug-drug interactions (with fludarabine and CY). Autologous HSCT is increasingly successful for treating a variety of malignancies in the clinic. Experimental and clinical data suggest that prior exposure to cytotoxic agents that damage primitive stem cells results in impaired hematopoiesis and an increased the risk of MDS/AML after autologous HSCT.
Specific aim 3 will determine predictive factors in donor HSC most important for impaired hematopoiesis in patients after autologous HSCT. We will determine the frequency and function of SP cells and CD34+KDR+ cells; two recently described human HSC populations, in normal marrow and in mobilized blood. We will assess which marker(s) in donor autologous marrow and mobilized blood predicts for impaired hematopoiesis after autologous HSCT. Transplanted CD34+38- cell, SP cell, and CD34+KDR+ cell content, and the presence of chromosomal abnormalities pre-transplant will be used as markers of intact hematopoiesis. We will correlate these markers with surrogates for HSC reserve capacity (post-HSCT time to platelet engraftment, impaired long-term hematopoiesis, and development of MDS). The successful completion of this specific aim should allow better identification of patients at high risk for marrow failure after autologous HSCT.
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