The main aims of this multiple-PI proposal are: to continue to define the underlying mechanisms of improved hepatocyte transduction by our next generation of recombinant adeno-associated virus (AAV) vectors; to extend these studies to novel AAV2 variants with high liver tropism, enriched from a library, and to AAV3-based vectors, which show high efficacy in human hepatocytes and non-human primate liver; and to evaluate the efficacy and host immune response in gene therapy for hemophilia B. Candidate vectors for clinical trial with high efficacy in small and large animal models will be identified, with particular emphasis on in vivo gene transfer to human/primate hepatocytes and immunobiology. The current application encompassing an interdisciplinary approach should allow us to develop superior next generation vectors for liver-directed human gene therapy, drawing heavily from expertise at the University of Florida in AAV vector biology, immunology of AAV gene transfer, and animal model studies. We will test the following hypotheses: a. Minor changes in the capsid sequence can substantially improve liver gene transfer, in part through postentry mechanisms, allowing us to design superior AAV2- and AAV3-based vectors. b. A combination of molecular evolution and capsid engineering will result in vectors with high transduction efficiency of human/primate hepatocytes and reduced MHC I antigen presentation. c. Capsid-modified AAV vectors will provide therapeutic F.IX expression at low vector doses and with reduced immunogenicity. The following three Specific Aims will be pursued:
Specific Aim 1 : Further optimize and compare in vivo performance of novel AAV2- and AAV3-based vectors, and define the mechanisms responsible for the robust liver gene transfer.
Specific Aim 2 : Identify novel capsid variants for high-efficiency in vivo gene transfer to human hepatocytes and adopt these into an advanced baculovirus production system.
Specific Aim 3 : Test improved vectors in animal models of hemophilia and in non-human primates, and define their immunogenicity. These investigations have a high chance of yielding superior vectors for liver gene transfer in humans.

Public Health Relevance

The main aim of this proposal is to develop the next generation of vectors with which a therapeutic gene can be safely delivered to patients with a bleeding disorder called hemophilia B. These vectors are derived from a virus that causes no known disease, and is therefore, expected to be safer. The development of such a vector for the potential treatment and cure of hemophilia therefore has relevance to public health.

Agency
National Institute of Health (NIH)
Institute
National Heart, Lung, and Blood Institute (NHLBI)
Type
Research Project (R01)
Project #
3R01HL097088-06S1
Application #
9197791
Study Section
Gene and Drug Delivery Systems Study Section (GDD)
Program Officer
Sarkar, Rita
Project Start
2009-07-01
Project End
2019-04-30
Budget Start
2016-05-01
Budget End
2017-04-30
Support Year
6
Fiscal Year
2016
Total Cost
Indirect Cost
Name
University of Florida
Department
Pediatrics
Type
Schools of Medicine
DUNS #
969663814
City
Gainesville
State
FL
Country
United States
Zip Code
32611
Tai, Phillip W L; Xie, Jun; Fong, Kaiyuen et al. (2018) Adeno-associated Virus Genome Population Sequencing Achieves Full Vector Genome Resolution and Reveals Human-Vector Chimeras. Mol Ther Methods Clin Dev 9:130-141
Wang, Dan; Gao, Guangping (2018) Taking a Hint from Structural Biology: To Better Understand AAV Transport across the BBB. Mol Ther 26:336-338
Wang, Dan; Li, Jia; Tran, Karen et al. (2018) Slow Infusion of Recombinant Adeno-Associated Viruses into the Mouse Cerebrospinal Fluid Space. Hum Gene Ther Methods 29:75-85
Biswas, Moanaro; Kumar, Sandeep R P; Terhorst, Cox et al. (2018) Gene Therapy With Regulatory T Cells: A Beneficial Alliance. Front Immunol 9:554
Wang, Dan; Li, Shaoyong; Gessler, Dominic J et al. (2018) A Rationally Engineered Capsid Variant of AAV9 for Systemic CNS-Directed and Peripheral Tissue-Detargeted Gene Delivery in Neonates. Mol Ther Methods Clin Dev 9:234-246
Lu, Yi; Tai, Phillip W L; Ai, Jianzhong et al. (2018) Transcriptome Profiling of Neovascularized Corneas Reveals miR-204 as a Multi-target Biotherapy Deliverable by rAAVs. Mol Ther Nucleic Acids 10:349-360
Rogers, Geoffrey L; Shirley, Jamie L; Zolotukhin, Irene et al. (2017) Plasmacytoid and conventional dendritic cells cooperate in crosspriming AAV capsid-specific CD8+ T cells. Blood 129:3184-3195
Srivastava, Arun; Carter, Barrie J (2017) AAV Infection: Protection from Cancer. Hum Gene Ther 28:323-327
Kumar, Sandeep R P; Hoffman, Brad E; Terhorst, Cox et al. (2017) The Balance between CD8+ T Cell-Mediated Clearance of AAV-Encoded Antigen in the Liver and Tolerance Is Dependent on the Vector Dose. Mol Ther 25:880-891
Kondratov, Oleksandr; Marsic, Damien; Crosson, Sean M et al. (2017) Direct Head-to-Head Evaluation of Recombinant Adeno-associated Viral Vectors Manufactured in Human versus Insect Cells. Mol Ther 25:2661-2675

Showing the most recent 10 out of 59 publications