Development of adenovirus serotype 35 as a gene transfer vector.

While 51 human adenoviral serotypes have been identified to date, the vast majority of adenoviral vectors designed for gene transfer have been generated in the adenovirus serotype 5 (Ad5) backbone. Viral infections caused by Ad5 are endemic in most human populations and the majority of humans carry preexisting humoral and/or cellular immunity to Ad5 which may severely limit the use of Ad5-based vectors for gene therapy applications. To circumvent this preexisting Ad5 immunity, we have identified Ad35 as an alternative adenoviral serotype to which the majority of humans do not have neutralizing antibodies.

An oncolytic adenovirus selective for retinoblastoma tumor suppressor protein pathway-defective tumors: dependence on E1A, the E2F-1 promoter, and viral replication for selectivity and efficacy.

The use of oncolytic adenoviruses as a cancer therapeutic is dependent on the lytic properties of the viral life cycle, and the molecular differences between tumor cells and nontumor cells. One strategy for achieving safe and efficacious adenoviral therapies is to control expression of viral early gene(s) required for replication with tumor-selective promoter(s), particularly those active in a broad range of cancer cells. The retinoblastoma tumor suppressor protein (Rb) pathway is dysregulated in a majority of human cancers.

Sustained human factor VIII expression in hemophilia A mice following systemic delivery of a gutless adenoviral vector.

Gutless adenoviral vectors are devoid of all viral coding regions and display reduced cytotoxicity, diminished immunogenicity, and an increased coding capacity compared with early generation vectors. Using hemophilia A, a deficiency in clotting factor VIII (FVIII), as a model disease, we generated and evaluated a gutless vector encoding human FVIII. The FVIII gutless vector grew to high titer and was reproducibly scaled-up from vector seed lots.

Breeding of retroviruses by DNA shuffling for improved stability and processing yields.

Manufacturing of retroviral vectors for gene therapy is complicated by the sensitivity of these viruses to stress forces during purification and concentration. To isolate viruses that are resistant to these manufacturing processes, we performed breeding of six ecotropic murine leukemia virus (MLV) strains by DNA shuffling. The envelope regions were shuffled to generate a recombinant library of 5 x 106 replication-competent retroviruses.

In vivo evaluation of an adenoviral vector encoding canine factor VIII: high-level, sustained expression in hemophiliac mice.

Hemophilia A is the most common severe hereditary coagulation disorder and is caused by a deficiency in blood clotting factor VIII (FVIII). Canine hemophilia A represents an excellent large animal model that closely mimicks the human disease. In previous studies, treatment of hemophiliac dogs with an adenoviral vector encoding human FVIII resulted in complete correction of the coagulation defect and high-level FVIII expression [Connelly et al.