Cell Genesys, Inc.

Other

University of Florida Research Foundation, Incorporated

University of Washington

The University of North Carolina at Chapel Hill

Richard O Snyder

Richard Jude Samulski

Xiao Xiao

Mark A Kay

Sergei Zolotukhin

William W Hauswirth

Nicholas Muzyczka

Barry John Byrne

Thomas L Dull

Mitchell H Finer

Edgardo Rodriguez

Stephen F Hardy

Lawrence K Cohen

Ryan McGuinness

Terence R Flotte

Olivier Danos

Brian Donahue

Mark R Potter

Scott Loiler

Vince A Chiodo

Corinna Burger

Kevin R Nash

Thomas J Fraites

Arthur R Thompson

Yoshihisa Sakai

Irine Zolotukhin

Stocks of infectious rAAV are generated using yeast strains, bacterial strains, and bacteriophages engineered to express the required AAV proteins and harboring rAAV vector sequences. Stocks of rAAV virions of all serotypes and pseudotypes can be generated in prokaryotic and eukaryotic cells using the methods described herein.

Production of recombinant AAV virions

The present invention relates to methods and compositions for increasing the production of high titre stocks of recombinant AAV (rAAV) through regulation of expression of the AAV REP and CAP proteins. The methods and compositions of the invention are based on the observation that the low level expression of the AAV REP protein increases the production of AAV viral capsid protein and efficiency of packaging resulting in production of higher titre recombinant viral stocks. The invention encompasses recombinant AAV vectors that direct the expression of AAV REP and CAP proteins and the use of such vectors for the production of novel stable cell lines capable of generating high titre rAAV vectors. The invention provides methods for regulating the expression of the AAV REP gene at the transcriptional and post-translational level. The methods and compositions of the invention can be used to produce high titre stocks of rAAV which can be used in gene therapy for the purpose of transferring genetic information into appropriate host cells for the management and correction of human diseases including inherited and acquired disorders.

Methods for increasing the efficiency of recombinant AAV product

Vectors that encode Adeno-Associated Virus (AAV) Rep and Cap proteins of different serotypes and Adenovirus transcription products that provide helper functions were used to produce pseudotyped recombinant AAV (rAAV) virions. Purification methods generated pseudotyped rAAV virion stocks that were 99% pure with titers of 1&#xd7;10&#x2013;1&#xd7;10vector genomes/ml.

Production of pseudotyped recombinant AAV virions

The instant invention provides methods of expressing polynucleotides in the cells of the liver comprising administering viral particles comprising a recombinant AAV vector into a mammal, preferably a human.

Adeno-associated viral vector-mediated delivery of DNA to cells of the liver

The instant invention provides methods and materials for expressing a polypeptide with factor VIII activity comprising administering an rAAV vector encoding a truncated version of human factor VIII, containing, for example, a 90 kD heavy chain of factor VIII fused to a light chain of factor VIII.

Recombinant AAV vectors for gene therapy of hemophilia A

Suppressor tRNA's are used to regulate expression of transgenes that are toxic, or the expression thereof requires a factor that is toxic, to the host cell.

Use of suppressor tRNA's to regulate cytotoxicity during the production of recombinant gene products

The present invention relates to methods and compositions for increasing the production of high titre stocks of recombinant AAV (rAAV) through regulation of expression of the AAV REP and CAP proteins. The methods and compositions of the invention are based on the observation that the low level expression of the AAV REP 78/68 protein increases the production of AAV viral capsid protein and efficiency of packaging resulting in production of higher titre recombinant viral stocks. The invention encompasses recombinant AAV vectors that direct the expression of AAV REP and CAP proteins and the use of such vectors for the production of novel stable cell lines capable of generating high titre rAAV vectors. The invention provides methods for regulating the expression of the AAV REP 78/68, REP 40/52 and CAP gene at the transcriptional and post-translational level. The methods and compositions of the invention can be used to produce high titre stocks of rAAV which can be used in gene therapy for the purpose of transferring genetic information into appropriate host cells for the management and correction of human diseases including inherited and acquired disorders.

Method for increasing the efficiency of recombinant AAV production

Growing community of inventors

Gainesville, FL, United States of America

Richard O Snyder