Development of a novel helper plasmid: one step closer to the next generation rAAV vectors

Eric Mauro, Jonathan Havard, Sylvain Julien, Laure Robert, Carine Morel, Patrick Erbacher

To broaden the use of rAAVs to treat a wider spectrum of monogenic diseas for small and larger indications, the aim is to improve specificity, safety and patient affordability. For this, it is key to improve rAAV production yields to decrease manufacturing costs, and improve functional titers to administer lower and safer doses. To this end, our research is focused on developing novel technologies to ensure manufacturing of high yielding rAAV using transient transfection, as well as enhancing features of the rAAV vectors that act on the overall size of packaged material and specificity of delivery.

Here we present our state-of-the art approach to designing new helper plasmids (phelpers) with the aim of improving both the infectiosity (TU/mL) and the quality (full/empty ratio) of the viral particle obtained from suspensions cultures. We took the opportunity to exploit our proprietary DNA assembly method technology, to rationally explore the synergies of multiple genetic features modularly assembled in synthetic plasmids. Comparison of the biological activity of several versions of rationally designed pHelpers led us to identify the optimal configuration able to outperform existing helper plasmids in every tested bioproduction conditions. Our ability to assemble tailor designed DNA plasmids and our expertise in developing scalable transfection solutions for rAAV manufacturing gives us potential to improve both productivity and specificity of gene therapy products.

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