The aim of this research proposal is to understand how transcriptional activity and its modification can be used to direct HIV-derived lentiviral vector integration in hematopoietic stem cells. Recent awareness of known lentiviral insertion site preferences for transcriptionally active genomic sites and reports of insertional mutagenesis after Moloney murine leukemia virus vector transduction of stem cells have highlighted the need to more fully investigate the biology of proviral integration. I herein propose to develop in vitro and in vivo model systems to explore strategies that should ultimately minimize insertion related risks of HIV-lentiviral gene replacement therapy. We hypothesize that systematic modifications in the transcriptional profile of hematopoietic target cells may serve to """"""""attract"""""""" insertion events to specific genomic locations while minimizing mutagenic potential and transcriptional interference at others, such as oncogenes and tumor suppressor loci. Critically, the proposed in vivo studies will aim to reconcile the competing requirements for transcriptional profiling and efficient lentiviral gene transfer with the retention of stem cell properties, including target cell engraftment and long-term repopulation. These studies will provide mechanistic insight into how transcription, be it a surrogate, or a causally related event, affects proviral insertion, and will inform future studies for genomic targeting. The proposed mentored training program will help me acquire additional skills and gain understanding through the mentorship by Dr. David Kabat and Dr. Markus Grompe, and their scientific expertise in virology and stem cell biology, respectively. The proposal is further augmented by input and guidance from advisors/collaborators in the areas of hematopoiesis, DNA array technology, and biostatistics, as well as course work. A career advisory committee will help direct my progress to independence and career advancement. The comprehensive training environment at Oregon Health & Science University (OHSU) and the substantial institutional commitment to my career will allow me to accomplish the goals of this proposal.

Agency
National Institute of Health (NIH)
Institute
National Heart, Lung, and Blood Institute (NHLBI)
Type
Clinical Investigator Award (CIA) (K08)
Project #
5K08HL077231-02
Application #
7089976
Study Section
Special Emphasis Panel (ZHL1-CSR-B (M2))
Program Officer
Mondoro, Traci
Project Start
2005-07-04
Project End
2010-06-30
Budget Start
2006-07-01
Budget End
2007-06-30
Support Year
2
Fiscal Year
2006
Total Cost
$129,060
Indirect Cost
Name
Oregon Health and Science University
Department
Pediatrics
Type
Schools of Medicine
DUNS #
096997515
City
Portland
State
OR
Country
United States
Zip Code
97239
Su, Shaoyong; Zhu, Haidong; Xu, Xiaojing et al. (2014) DNA methylation of the LY86 gene is associated with obesity, insulin resistance, and inflammation. Twin Res Hum Genet 17:183-91
O'Neill, Lee S; Skinner, Amy M; Woodward, Josha A et al. (2010) Entry kinetics and cell-cell transmission of surface-bound retroviral vector particles. J Gene Med 12:463-76
Skinner, Amy M; O'Neill, S Lee; Kurre, Peter (2009) Cellular microvesicle pathways can be targeted to transfer genetic information between non-immune cells. PLoS One 4:e6219
Pan, Yung-Wei; Kurre, Peter (2009) Avoiding lentiviral transduction culture induced MSC senescence. J Cell Mol Med 13:1186-7
Skinner, Amy M; O'Neill, S Lee; Grompe, Markus et al. (2008) CXCR4 induction in hematopoietic progenitor cells from Fanca(-/-), -c(-/-), and -d2(-/-) mice. Exp Hematol 36:273-82
Harkey, Michael A; Kaul, Rajinder; Jacobs, Michael A et al. (2007) Multiarm high-throughput integration site detection: limitations of LAM-PCR technology and optimization for clonal analysis. Stem Cells Dev 16:381-92
Pan, Yung-Wei; Scarlett, Jarrad M; Luoh, Tammy T et al. (2007) Prolonged adherence of human immunodeficiency virus-derived vector particles to hematopoietic target cells leads to secondary transduction in vitro and in vivo. J Virol 81:639-49