Gene Therapy for Urea Cycle Disorders

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DESCRIPTION (Provided by Applicant): This application requests two years of revised support for Project I of a program project entitled "Gene Therapy for Urea Cycle Disorders" which is in its second year of funding. All three projects of the P0I have been productive in advancing the aims of the application and remain on schedule according to the original timelines. Specific Aim 3 of Project I was to evaluate the role of inflammation in the performance of AAV vectors. The hypothesis was that a critical step in rendering AAV-transduced hepatocytes susceptible to killing by cytotoxic T lymphocytes (CTLs) was up-regulation of MHC class I and improved presentation of antigen. A series of adoptive transfer studies did confirm this hypothesis. Transfer of lacZ-specific CTLs into mice whose livers were transduced with AAV-LacZ failed to clear transgene expression unless the animals were treated with ligands for Toll-like receptors (TLRs) at the time of the adoptive transfer; under these conditions there was complete loss of transgene expression and clearance of the vector genome concurrent with up-regulation of MHC class I. The investigators recently extended these studies beyond the scope of the program to evaluate the possibility that innate immunity may be sufficient to extinguish transgene expression independent of capsid or transgene T cells. In fact, they showed that injection of low doses of an adenoviral vector expressing an irrelevant transgene with TLR ligands two weeks subsequent to AAV gene transfer was sufficient to completely abolish AAV-encoded transgene expression; vector genomes were diminished but not eliminated, suggesting transcriptional shut off may play a role as well as partial destabilization of the vector genome. The revision aims to further evaluate the role of inflammation in inhibiting transgene expression from AAV vectors in liver. The revised Specific Aim 1 will evaluate a number of important questions regarding the investigators' recent observation, including: Does the low dose Ad contribute through the generation of CTLs or via direct activation of APCs? Do the TLR ligands contribute through engagement of their cognate receptors? Are pro-inflammatory cytokines necessary and/or sufficient? And finally, what is the relative role of signaling via the hepatocyte versus resident APCs? The revised Specific Aim 2 will utilize a mouse model to evaluate the impact of chronic hepatitis infection on AAV vector performance when hepatitis is present at the time of gene transfer or subsequent to gene transfer. RELEVANCE: Adeno-associated viruses hold promise for liver-directed gene therapy in the treatment of a variety of inherited and acquired disorders. This application evaluates the role of liver inflammation in the efficacy and safety of liver-directed gene transfer. The results will be important in the selection of candidate diseases and design of clinical trials.