Engineered AAV vectors for combinatorial treatment of rare genetic brain diseases

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Project Summary AAV-based gene therapy requires the development of safe, efficient, and target-specific vectors. AAV-mediated gene therapy for peripheral tissues (blood and skeletal muscle) has made great strides, focused mainly on gene replacement for loss of function diseases. However, there has been little research on dominantly inherited CNS disorders that are caused by toxic gene products. We have assembled a world class, multi-disciplinary academic research team supported by our industrial partners to develop innovative AAV-based gene knockdown and replacement treatments for rare genetic diseases including Spinocerebellar Ataxia Type 7 (SCA7) and valosin- containing protein (VCP) multisystem proteinopathy. We respond to the RFA Project Objectives and propose three Specific Aims. In Aim 1, we will design and manufacture new AAV vectors with improved critical quality attributes (safety, efficacy, target specificity) for gene therapy. We propose innovative neuron specific gene delivery, temporal control of gene expression and reduced immune responses in the CNS. We have constructed AAV vectors that express EGFP and mRNA barcodes for improved screening to support our proposed AAV treatments. In Aim 2, we will develop advanced quantitative analytics by combining next-generation sequencing and bar-coded AAVs for efficient assessment of in vivo gene delivery in the mouse model. We will screen and compare AAV-mediated gene expression with different capsid variants using different promoters / enhancers. Single-cell RNAseq also will be used to assess immunological responses of selected AAV vectors by different administration routes. In Aim 3, to effectively treat the SCA7 and VCP diseases, we will develop novel AAV vectors that simultaneously knockdown toxic gene products while replacing normal gene products that are codon optimized to be unaffected by knockdown. This proposed combinatorial treatment will establish a proof-of- concept for many other dominant inherited diseases, where loss of normal allele expression due to non-specific silencing causes its own problems. We have well developed models of the SCA7 and VCP diseases for Aim 3 studies. Our team has great expertise in developing, manufacturing and applying AAV vectors in basic research and preclinical application. Our published work and preliminary data establish the feasibility and key methodologies for the proposed research. Critically, we have an established viral production facility and distribution platform operating out of our UCI Center for Neural Circuit Mapping that supports viral reagent design, validation, and manufacturing.