Avian adenoassociated virus and uses thereof
Inventors
Bossis, Ioannis • Chiorini, John A.
Assignees
General Hospital Corp • National Institutes of Health NIH • US Department of Health and Human Services
Publication Number
US-8927269-B2
Publication Date
2015-01-06
Expiration Date
2024-05-18
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Abstract
The present invention provides an Avian adeno-associated virus (AAAV) virus and vectors and particles derived therefrom. In addition, the present invention provides methods of delivering a nucleic acid to a cell using the AAAV vectors and particles. Methods of isolating the AAAV are provided.
Core Innovation
The present invention provides an avian adeno-associated virus (AAAV), vectors, and particles derived therefrom, along with methods for delivering nucleic acids to cells using AAAV vectors and particles. It also discloses methods for isolating AAAV. The genome of AAAV is cloned and sequenced, revealing a distinct genetic and physicochemical profile compared to known primate AAVs. AAAV possesses inverted terminal repeats (ITRs) that form characteristic T-shaped hairpin structures and can serve as a universal origin of replication, recognized by Rep proteins from different AAV serotypes, including AAV2 and AAV5.
The problem addressed is the need for new and efficient gene delivery vectors with different tissue tropism and immunological properties to overcome limitations in existing AAV vectors. Existing AAVs are mostly from primate origins, limiting their efficiency and utility in certain applications, especially in avian species or in patients with pre-existing immunity to primate AAV serotypes. The invention solves this by providing AAAV, which is serologically distinct and exhibits a unique tropism favoring avian cells, expanding the versatility of AAV vectors in gene therapy, gene transfer, transgenic animal development, vaccination, and recombinant protein production.
Further, AAAV vectors incorporate novel features such as unique capsid proteins (VP1, VP2, VP3) differing from primate AAVs, which enable altered tissue tropism. The AAAV genome organization parallels other AAVs but maintains distinct sequences especially in the capsid proteins and Rep ORFs, providing improved transduction efficiency in avian cells and with potential effectiveness in subjects having neutralizing antibodies against primate AAVs. These genetic and structural differences establish AAAV as a valuable novel vector system for nucleic acid delivery.
Claims Coverage
The claims disclose multiple inventive features centered on nucleic acid vectors and recombinant virions comprising AAAV inverted terminal repeats (ITRs), capsid proteins, and heterologous nucleic acids, highlighting the novel sequences and their use in gene delivery applications.
Nucleic acid vector with AAAV-specific inverted terminal repeats
A nucleic acid vector comprising a pair of inverted terminal repeats (ITRs) capable of forming T-shaped hairpin structures, where at least one ITR comprises specific nucleotide sequences (SEQ ID NO:20 and SEQ ID NO:21), serving as an origin of replication, and containing a heterologous nucleic acid sequence between the ITRs.
Recombinant AAAV virion containing a vector with AAAV ITRs and heterologous nucleic acid
A recombinant AAAV virion comprising a pair of inverted terminal repeats as defined above and a heterologous nucleic acid between the ITRs, where the ITRs serve as a replication origin.
Vectors encoding AAAV capsid proteins
Vectors, including plasmids, yeast artificial chromosomes, or non-AAV viral vectors, encoding AAAV capsid proteins VP1, VP2, or VP3 (SEQ ID NOS:11, 13, 15) useful for producing infectious AAAV particles.
Vector system for producing infectious AAAV particles
A vector system comprising at least one nucleic acid molecule with AAAV ITRs and heterologous nucleic acid, and/or recombinant nucleic acid encoding AAAV capsid proteins, designed to produce infectious AAAV particles.
Chimeric and hybrid constructs combining AAAV components
Nucleic acid vectors comprising AAAV inverted terminal repeats, capsid encoding sequences, and rep encoding sequences (SEQ ID NOS:3, 5, 7, 9) enabling assembly and production of functional AAAV particles.
Overall, the claims cover inventive nucleic acid vectors and recombinant virions featuring distinctive AAAV inverted terminal repeats capable of replication, heterologous nucleic acid inserts, and AAAV capsid proteins. These allow for the production and use of infectious AAAV particles with unique genetic components distinct from known AAV serotypes for nucleic acid delivery.
Stated Advantages
AAAV vectors have unique tissue tropism distinct from primate AAVs, enabling efficient transduction of avian cells and cell types not well transduced by primate AAVs.
AAAV is serologically distinct from primate AAVs, allowing gene delivery in subjects with neutralizing antibodies against other AAV serotypes.
The AAAV ITR functions as a universal origin of replication compatible with Rep proteins from AAV2 and AAV5, enhancing vector versatility and packaging efficiency.
Recombinant AAAV vectors can be used to develop transgenic animals, facilitate vaccination in avian species, and produce recombinant proteins in eggs.
Documented Applications
Gene therapy and gene transfer applications involving delivery of nucleic acids to cells or subjects using AAAV vectors and particles.
Development of transgenic animals via gene transfer using AAAV vectors.
Vaccination of eggs or avian species with AAAV vectors encoding vaccine antigens.
Production of recombinant proteins in avian cells or embryonated chicken eggs by administering AAAV particles carrying nucleic acids encoding the proteins.
Screening cells for permissiveness to AAAV infection based on binding to N-linked terminal lactose.
Use of AAAV vectors for subjects possessing neutralizing antibodies to primate AAV serotypes, facilitating gene delivery.
Blocking AAAV infection by using lactose conjugates or lectins targeting N-linked terminal lactose as a receptor.
Modification of AAAV capsid proteins to alter tissue tropism, antigen presentation, or immunogenicity for therapeutic or vaccine purposes.
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