Improved system for producing adenoviral vectors

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First generation non-replicative adenoviral vectors show promise as anti-cancer agents and as prophylactic vaccines. Current methods for manufacturing these vectors use either the 293 cell line or the PER.C6 cell line. These producer cells provide complementary E1a and E1b viral function in the cell for disabled adenoviral vectors. The 293 cell line has been very useful and widely used for making relatively small batches of vector for early stage clinical trials. However the frequency with which it generates replication competent adenovirus, which disqualifies from 10 to 50% of batches depending on the vector, means that, as the manufacturing processes are scaled up in later stage clinical trials, or for marketing, it becomes less and less acceptable. As scale increases, increasing numbers of batches exceed the FDA mandated allowable contamination level of 1RCA/3x10e10 viral particles, and such a manufacturing process cannot be claimed to be ?controlled? by FDA standards if the failure rate is greater than ~ 10%. PER.C6 cells use a specifically designed matched vector to eliminate sequence overlap between producer line and vector. The system is not compatible with most current clinical vectors and is not available to academic researchers or small companies. A modest 10 fold reduction in RCA frequency over that seen in 293 cells would provide a robust production system if the vector productivity were equivalent. We have constructed a cell line named c24 based on the human tumor line A549, which is as productive as 293 cells, is stable, is free of mycoplasma and carries 1-2 copies of an E1 expression plasmid with very limited overlap with conventional adenoviral vectors. We propose to rigorously test the RCA frequency in c24 compared to 293 cells, and to adapt it to serum free growth. If the RCA generation can be reduced by >10-fold, and in a serum free environment, then c24 can provide the basis for a reliable vector production system with a further improved safety profile that can be widely used to manufacture adenoviral vector products, including Advantagene?s own late stage pipeline.