The present invention discloses adeno-associated viral vectors useful in gene therapy methods for the treatment of obesity, insulin resistance, type 2 diabetes, liver cirrhosis and non-alcoholic fatty liver disease (NAFLD)/non-alcoholic steatohepatitis (NASH). The invention also relates to polynucleotides, plasmids, vectors and methods for the production of such adeno-associated viral vectors. The invention also relates to pharmaceutical compositions comprising said vectors.

Methods for separating AAV empty capsids from mixtures of AAV vector particles and AAV empty capsids are described. The methods use column chromatography techniques and provide for commercially viable levels of recombinant AAV virions.

The present disclosure provides methods and compositions for the treatment of diseases and genetic disorders linked to CSTB loss and/or misfunction. The methods and compositions of the present disclosure include rAAV vectors and rAAV viral vectors comprising transgene nucleic acid molecules encoding CSTB polypeptides.

Compositions and methods are provided for preparation of concentrated stock solutions of AAV virions without aggregation. Formulations for AAV preparation and storage are high ionic strength solutions (e.g. μ˜500 mM) that are nonetheless isotonic with the intended target tissue. This combination of high ionic strength and modest osmolarity is achieved using salts of high valency, such as sodium citrate. AAV stock solutions up to 6.4×10.sup.13 vg/mL are possible using the formulations of the invention, with no aggregation being observed even after ten freeze-thaw cycles. The surfactant Pluronic® F68 may be added at 0.001% to prevent losses of virions to surfaces during handling. Virion preparations can also be treated with nucleases to eliminate small nucleic acid strands on virions surfaces that exacerbate aggregation.

Formulations for highly purified viral particles (e.g., adeno-associated virus (AAV) particles) are provided herein. The formulations include purified AAV particles that are substantially free of impurities (e.g., product-related impurities and process-related impurities), and one or more of a buffering agent, a cryoprotectant, a non-ionic surfactant, and optionally a pharmaceutically acceptable salt. In certain aspects, the formulation maintains or enhances stability and/or reduces or prevents aggregation of the purified AAV particles.

Adeno-associated virus rh.20 sequences, vectors containing same, and methods of use are provided.

The invention relates to methods of selectively purifying an adeno-associated virus (AAV) from an aqueous biomass using a flocculent. In embodiments, the flocculent is polydiallyldialkylammonium salt, e.g., polydiallyldimethvlammonium chloride (pDADMAC).

This disclosure relates to the use of size exclusion chromatography and/or size exclusion chromatography with multi-angle light scattering technology to characterize viral particles such as adeno-associated virus and lentivirus particles. The disclosed methods are also useful for estimating the titer of viral particles, determining the integrity of the viral particles and estimating the amount of DNA encapsidated in the viral particle.

Provided herein are improved, cost-effective and environmentally friendly methods of production of recombinant AAV (rAAV) in embryonated avian eggs. Further provided herein is a provides embryonated avian eggs as novel host vehicles for high-yield production of rAAV, including both packaging and propagation. In particular, embryonated chicken eggs provide a novel expression vehicle for AAV of mammalian origin, irrespective of AAV serotype. The disclosed methods may comprise packaging of rAAV in embryonated avian eggs (e.g., chicken eggs) by inoculating an embryonated avian egg with a first nucleic acid vector comprising a transgene and a second nucleic acid vector comprising AAV rep and cap genes, incubating the egg, and isolating rAAV from the egg, wherein the AAV is of non-avian origin. Also provided are methods of purifying and propagating packaged rAAV in embryonated avian eggs or in avian embryonic fibroblasts.

Provided is a method of enriching full adenovirus-associated virus (AAV) capsids from a mixture of full AAV capsids and empty AAV capsids, the method comprising providing a sample comprising a mixture of full AAV capsids and empty AAV capsids, subjecting the sample to anion exchange chromatography comprising an elution buffer comprising an equilibration buffer and a salt-containing buffer in an initial ratio that provides an initial conductivity, and changing the ratio of the equilibration buffer and the salt-containing buffer to provide a step gradient conductivity increase of about 0.5-2.0 mS/cm in each step to elute empty AAV capsids and to provide a fluid enriched with full AAV capsids.