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Related post: linked on 4ql3-q21 have only two functional receptors encoded by genes on 2q34-q35, Finally, we have cloned the human A2 adenosine receptor cDNA and gene. PHa eiMO (Hev. b«2) 7-9 DEPARTMENT OF HEALTH AND HUMAN SERVICES - PUBLIC HEALTH SERVICE NOTICE OF INTRAMURAL RESEARCH PROJECT PROJECT NUMBER ZOl AI 00644-01 PERIOD COVERED October 1, 1991 to September 30, 1992 TITLE OF PROJECT (80 characters or less. TiUe must fit on one line between the borders.) Biology of Peripheral Blood Progenitors: a Target for Gene Transfer. PRINCIPAL INVESTIGATOR (Lisl other prolessional personnel below the Principal Investigator) (Name, title, laboratory, and institute alliliation) PI: Harry L. Malech, M.D. Assistant Laboratory Chief LHD/NIAID Others; Sudhir Sekhsaria, M.D. John I. Gallin, M.D. Clinical Associate Laboratory Chief LHD/NIAID LHD/NIAID COOPERATING UNITS (11 any) LAB«RANCH Laboratory of Host Defenses INSTITUTE AND LOCATION NJAID, NIH, Bethesda, MD 2089: TOTAL STAFF YEARS: 1.5 PROFESSIONAL: 1.0 OTHER: 0.5 CHECK Dexamethasone 0.1 Eye Drops APPROPRIATE BOX(ES) Kl (a) Human subjects E (a1) Minors D (a2) Interviews D (b) Human tissues D (c) Neither SUMMARY OF WORK (Use standard unreduced type. Do not exceed the space provided.) This project studies peripheral blood progenitors (PEP) as mediated gene therapy of inherited diseases affecting a target for retrovirus the function of human which phagocytic cells , including neutrophils and monocytes . PBPs which may include totipotent stem cells are similar or identical to the blood precursors in bone marrow , but circulate in small numbers in peripheral blood. PBP harvested and concentrated from peripheral blood by centrif ugation apheresis have been used successfully by a number of investigators as an autologous transplant in lieu of or combined with progenitors harvested from bone marrow to reconstitute the bone marrow of patients with cancer treated with cytotoxic chemotherapy. Our hypothesis is that PBP harvested from patients with inherited immune diseases such as chronic granulomatous disease or leukocyte adhesion deficiency are a readily available target for functional correction by gene transfer. Using either whole peripheral blood or apheresis derived leukocytes enriched for PBP, we used monoclonal antibodies to enrich PBP several thousand fold yielding a population of cells where 40-90% of cells expressed the surface antigen CD34 characteristic of early blood progenitors. Using both liquid and soft agarose cultures we determined optimum conditions for growth and differentiation of PBP into colonies of mature neutrophils demonstrating a cloning efficiency of up to 10%. We showed that maximum proliferation occurred when the following growth factors were added to the culture: multilineage growth factor ( MGF ) (also called stem cell factor [ SCF ] ) , FGF-P, IL6, IL3, GM-CSF, and G-CSF . SCF acted as a co-factor to increase both the number and size of colonies. We also showed that the number of colony forming PBP in circulating blood was remarkably constant between individuals at 12 colony forming PBP/ml blood. This number increased from 2 to 4 fold in volunteers administered 3 ng/kg endotoxin. In related studies using the retroviral construct LXSN to transfer neomycin resistance to PBP we showed that under ideal growth conditions up to 30% of colony forming PBP cells could be transduced to acquire neomycin resistance. Thus, we have established methodology to harvest and purify PBP, to grow PBP in culture, and to transfer and express new genes in PBP. These represent important steps in the development of techniques for gene therapy of immune diseases affecting blood phagocytic cells. PHl> 6040 (Hev. 5/92) 7-10 DEPARTMENT OF HEALTH AND HUMAN SERVICES - PUBLIC HEALTH SERVICE NOTICE OF Dexamethasone Acetate Tablets INTRAMURAL RESEARCH PROJECT PROJECT NUMBER ZOl AI 00645-01 PERIOD COVERED October 1, 1991 to September 30, 1992 TITLE OF PROJECT (80 characlers or less. TiOe must fil on one line between me borders.) Gene Therapy for Immune Deficiencies. PRINCIPAL INVESTIGATOR (List other protassional personnel Pms Dexamethasone 4 Mg below the Principal Investigator.) (Name, title, laboratory, and institute alliliation) PI: Harry L. Malech, M.D. CoPI: John I. Gallin, M.D. Others: Sudhir Sekhsaria, M.D. Julie Katkin, M.D. Susan Dorman Assistant Laboratory Chief LHD/NIAID Laboratory Chief LHD/NIAID Clinical Associate LHD/NIAID Special Volunteer LHD/NIAID Guest Reseacher LHD/NIAID COOPERATING UNITS (il any)
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