cell-therapy-public-health-congress-2019-posters

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April 15-16, 2019 | Milan, Italy

OF EXCELLENCE

IN INTERNATIONAL

MEETINGS

YEARS

Joint Event on

PUBLIC HEALTH,

EPIDEMIOLOGY AND NUTRITION

World Congress on

CELL AND GENE THERAPY

International Conference on

Cell and Gene Therapy 2019 & Public Health Congress 2019

Archives of General Internal Medicine | ISSN: 2591-7951 | Volume 3

TOWARDS A 3

GENERATION AAV MANUFACTURING PLATFORM AND IN-PROCESS

CONTROLS

Ales Strancar

BIA Separations, Slovenia

deno associated virus (AAV) is the leading vector in the field of gene therapy because of its low toxicity,

good overall safety profile and ability to maintain stable expression for long periods of time. It is therefore

crucial to develop a robust and high efficiency platform for its manufacturing. One of the key challenges in

manufacturing viral vectors is managing the interface between upstream and downstream processing. Exper-

imental observations indicate that, in addition to losses due to shear, processing methods such as precipita-

tion, freeze/thaw and Tangential Flow Filtration (TFF) promote formation of stable associations between virus,

DNA and proteins. These complexes lower virus recovery and process capacity, depress robustness, and inflate

contamination at each processing step. They accordingly have strong potential to influence long term clinical

safety, especially with respect to residual DNA. This paper will present a 2

generation AAV manufacturing

platform engineered specifically to address these issues. After removal of cell debris by filtration, AAV is cap-

tured and fractionated by hydrophobic interaction chromatography (HIC). The AAV fraction from HIC is diluted

and loaded onto a cation exchanger under conditions to dissociate AAV from virus-protein-DNA complexes

and strongly bind remaining protein-DNA complexes. The cation exchange fraction is then applied to an anion

exchanger as a final DNA dissociation-polishing step and to separate empty and full capsids. This orthogonal

process achieves very high recoveries (>70%) with AAVs that are secreted from the host cells. It is fully scalable,

very robust, and has proven effective for all AAV serotypes evaluated to date. In addition, the 3-step chromato-

graphic process without preliminary TFF is very efficient and improves overall reduction of all contaminating

viruses. A 3rd generation process is under development to fully extend these benefits to AAVs processed by cell

lysis and to reach extra lowDNA impurity profile. Rapid sensitive analytics usingmultiple in-line HPLC detectors

to provide insightful process development and manufacturing documentation will also be presented.

Arch Gen Intern Med 2019, Volume 3 | DOI: 10.4066/2591-7951-C2-027