467. Consistency of Research Grade Lentiviral Vector Manufacturing Over 60 Batches

Lentiviral vectors (LV) have shown to be efficient gene transfer vehicles in a growing list of gene therapy applications and are currently being evaluated in clinical trials. bluebird bio has developed a GMP manufacturing process for large-scale production of third generation VSV-G pseudotyped HIV-1 based lentiviral vectors to support such trials. To cope with the increased demand for researchgrade LV material for preclinical testing, a mirror small-scale manufacturing model is used at bluebird bio to generate LV batches at scales ranging between 2 and 10L. The manufacturing process is based on the transient transfection of adherent HEK 293T cells with packaging and envelope plasmids along with a transfer plasmid coding for a gene of interest. Harvested supernatants are subsequently concentrated and purified using a combination of membrane filtration and chromatography steps. Analytical data obtained over 60 batches of LV manufactured using this process indicates that it consistently delivers high quality vector preparations with an infectious titer u003e 1E+08 TU/mL and a particle/infectious ratio between 100-500, emphasizing the robustness of the process design. Process yields typically range between 30 and 60 %, according to both infectious particle determinations using the HOS cell titer assay and total particle determinations as assessed by p24 ELISA. A linear correlation between harvested supernatant LV titers and final product LV titers is observed. This process is used to generate LV preparations to support ALD, β-Thalassemia, Sickle Cell Disease and CAR-T programs.