Role of Endothelial Progenitors in Hematopoietic Stem Cell Expansion

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Human umbilical cord blood has been demonstrated to be an effective source of hematopoietic stem cells (HSC) for clinical transplantation. A major limitation to the more widespread use of this material for adult patients is that the number of HSC in a cord blood sample may be insufficient to engraft in adult patients, particularly unrelated and mismatched recipients. Determination of a method to expand HSC in cord blood would be a major advance in the field. We have recently identified a hierarchy of endothelial progenitor cells (EPC) that circulate in the blood, but also comprise a portion of the endothelial cell intima in human and rhesus macaque blood vessels. Similar to the hematopoietic system, EPC are organized in a hierarchy in which the most proliferative progenitors give rise to numerous progeny with less proliferative potential but greater evidence of maturation with the most mature endothelial cells displaying no proliferative potential. In preliminary studies, human umbilical cord blood was enriched in EPC compared to adult blood. We hypothesize that the frequency and distribution of EPC changes during ontogeny such that the number of circulating cells and EPC residing in vessels decline with age and there is loss of proliferative potential that correlates with telomerase activity. In other preliminary data, we have determined that cord blood EPC co- cultured with human marrow stem cells expands HSC >260 fold compared to freshly isolated marrow HSC. We now hypothesize that EPCwith the highest proliferative potential will better promote HSC expansion than EPC with lower proliferative potential. To test these hypotheses, we proposed the following aims: Specific Aim 1. Determine the role of human cord blood EPC in expanding human cord blood HSC ex vivo. Human cord blood will be co-cultured with EPC ¿ growth factors and the repopulating ability compared following injection into rhesus fetal hosts in utero. Specific Aim 2. Determine the frequency and proliferative potential of EPC derived from blood and vascular endothelium throughout rhesus ontogeny. Aortic endothelial cells and peripheral blood of rhesus subjects from fetal to aged adults will be examined for EPC content, proliferative potential, and telomerase activity. We hypothesize that EPC proliferative potential declines with age and the rhesus model permits us an opportunity to sample vessels and blood throughout the entire species ontogeny. Specific Aim 3. Determine the role of EPC derived from fetal versus adult blood vessels in the in vitro expansion of fetal rhesus cord blood and adult marrow HSC. We hypothesize that the fetal EPC will more optimally expand fetal than adult HSC but adult HSC will be better expanded on fetal than adult EPC. Indiana University, Indianapolis, IN PHS 398 (Rev. 09/04) Page 98 Form Page 2 Principal Investigator/Program Director (Last, First, Middle): Tarantal, Alice F (Project 1) KEY PERSONNEL. See instructions. Use continuation pages as needed to provide the required information in the format shown below. Start with Principal Investigator. List all other key personnel in alphabetical order, last name first. Name eRA Commons User Name Organization Role on Project Yoder, Men/inC. Indiana Unversity Project Leader Ingram, David A. Indiana University Co-Investigator OTHER SIGNIFICANT CONTRIBUTORS Name Organization Role on Project Human Embryonic Stem Cells [3 No Q Yes If the proposed project Involves human embryonic stem cells, list below the registration number of the specific cell line(s) from the following list: http://stemcells.nih.aov/reqistrv/index.asp. Usecontinuation pages as needed. If a specificline cannot be referenced at this time, include a statement that one from the Registry will be used. Cell Line Disclosure Permission Statement. Applicable toSBIR/STTR Only. See instructions. I I Yes I I No PHS 398 (Rev. 09/04) Page 99 Form Page 2-continued Number the followingpages consecutively throughout the application. Do not use suffixes such as 4a, 4b. Principal Investigator/Program Director (Last, First, Middle): Tarantal, Alice F. (Project 1) DETAILED BUDGET FOR INITIAL BUDGET PERIOD FROM THROUGH DIRECT COSTS ONLY 09/01/05 08/31/06 PERSONNEL (Applicant organization only) % DOLLAR AMOUNT REQUESTED (omit cents) TYPE EFFORT INST. ROLE ON APPT. ON BASE SALARY FRINGE NAME PROJECT (months) PROJ. SALARY REQUESTED BENEFITS TOTAL Principal Mervin C. Yoder Investigator 12 10.0 180,000 18,000 3 227 21,227 David Ingram Co-l 12 5.0 0 0 0 Jennifer Stewart Res Tech 12 100.0 39,140 39,140 12 263 51,403 SUBTOTALS 57,140 15,490 72,6301 ^ CONSULTANT COSTS None 0 EQUIPMENT (Itemize} None 0 SUPPLIES (Itemize by category) ILJ Laboratory supplies 37,500 TRAVEL None PATIENT CARE COSTS INPATIENT OUTPATIENT ALTERATIONS AND RENOVATIONS (Itemize by category) OTHER EXPENSES (Itemize by category) Wells Center Research Core Use Fee $3,576 3,576 CONSORTIUM/CONTRACTUAL COSTS DIRECT COSTS SUBTOTAL DIRECT COSTS FOR INITIAL BUDGET PERIOD (Item 7a, Face Page) $ 113,706 CONSORTIUM/CONTRACTUAL COSTS FACILITIES AND ADMINISTRATIVE COSTS 58,559 TOTAL DIRECT COSTS FOR INITIAL BUDGET PERIOD $ 172,265 SBIR/STTR Only: FEE REQUESTED PHS 398 (Rev. 09/04) Page 100 Form Page 4