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Synthetic combinatorial aav capsid library for targeted gene therapy

Inactive Publication Date: 2016-12-22
UNIV OF FLORIDA RES FOUNDATION INC
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Benefits of technology

The present invention provides a better way to create viruses called adeno-associated virus (AAV) that can target specific tissues and organs in the body. By using a new method, researchers can modify the virus's surface to improve its ability to target specific cells. This method involves creating new libraries of viral DNA and using them to generate new variants of the virus. This new approach makes it more likely to find useful viral variants that can be tried out in humans.

Problems solved by technology

In addition, gene synthesis allows virtual recombination between naturally-occurring sequences at the nucleotide level, producing unique combinations that cannot be obtained through other methods.

Method used

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  • Synthetic combinatorial aav capsid library for targeted gene therapy
  • Synthetic combinatorial aav capsid library for targeted gene therapy
  • Synthetic combinatorial aav capsid library for targeted gene therapy

Examples

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example 1

AAV Vector Library Construction

[0149]Methodologies to improve existing AAV vectors for gene therapy include either rational approaches or directed evolution to derive capsid variants characterized by superior transduction efficiencies in targeted tissues. In the present invention, both approaches were integrated in one unified design strategy of “virtual family shuffling” to derive a combinatorial capsid library whereby only variable regions on the surface of the capsid are modified. Individual sub-libraries were first assembled in order to pre-select compatible amino acid residues within restricted surface-exposed regions to minimize the generation of dead-end variants. Subsequently, the successful families were interbred to derive a combined library of about 1×108 complexity. Next-Gen sequencing of the packaged viral DNA revealed capsid surface areas susceptible to directed evolution thus providing guidance for future designs. The utility of the library in gene therapy application...

example 2

Integrating Combinatorial and Rational Approaches to Derive Novel Aav Variants

[0154]AAV is a single-stranded DNA virus belonging to the Parvoviridae family (Muzyczka and Berns, 2001). AAV-derived vectors are promising tools for human gene therapy applications because of their absence of pathogenicity, episomal localization and stable transgene expression (Wu et al., 2006). However, significant limitations to the clinical use of AAV are its promiscuity and its susceptibility to neutralization by human antibodies (Louis-Jeune et al., 2013). Both of these limitations are determined by the nature of amino acid residues exposed at the surface of the capsid. Two main strategies are generally employed to improve AAV vectors: 1) mutagenizing capsid residues to facilitate binding, entry, and / or intracellular trafficking through a rational approach based on the knowledge of virus biology (Wu et al., 2000; Zhong et al., 2008; Lochrie et al., 2005; Aslanidi et al., 2012; Li et al., 2012; Gabrie...

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PUM

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Abstract

Disclosed are compositions and methods for producing modified adeno-associated virus (AAV) cap genes and combinatorial libraries of chimeric AAV vectors and virions; selecting for virions displaying cell-specific tropisms; and, in certain embodiments, producing helper vectors containing one or more modified AAV cap genes. The synthetic combinatorial AAV capsid libraries of the invention are useful in introducing into selected target host cells one or more nucleic acid molecules. The viral vectors and genetic constructs disclosed herein are also useful in a variety of diagnostic and / or therapeutic gene-therapy regimens.

Description

CROSS-REFERENCE TO RELATED APPLICATIONS[0001]The present application claims priority to U.S. Provisional Patent Appl. No. 61 / 883,063, filed Sep. 26, 2013 (pending; Atty. Dkt. No. 36689.341). The content of which is specifically incorporated herein in its entirety by express reference thereto.STATEMENT REGARDING FEDERALLY SPONSORED RESEARCH OR DEVELOPMENT[0002]This invention was made with government support under Grant Nos. HL-097088 and GM-082946 awarded by the National Institutes of Health. The government has certain rights in the invention.NAMES OF THE PARTIES TO A JOINT RESEARCH AGREEMENT[0003]Not Applicable.BACKGROUND OF THE INVENTION[0004]Field of the Invention[0005]The present invention relates to the fields of molecular biology, medicine, and virology. More particularly, the invention relates to compositions and methods for producing modified adeno-associated virus (AAV) cap genes and combinatorial libraries of chimeric AAV vectors and virions; and in particular embodiments, ...

Claims

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Application Information

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IPC IPC(8): C12N15/86C12N7/00C07K14/005
CPCC12N15/86C07K14/005C12N2750/14143C12N2750/14122A61K48/00C12N7/00C12N2750/14145
Inventor MARSIC, DAMIENZOLOTUKHIN, SERGEIAGBANDJE-MCKENNA, MAVIS
Owner UNIV OF FLORIDA RES FOUNDATION INC
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