Abstract：
:Lentiviruses are quite effective gene delivery systems for stable production of genetically engineered human cells. However, prior to using lentivirus to deliver genetic materials to cells of interest, the normal course of production of these lentiviruses involves a lengthy collection, purification, preservation, and quantification process. In this report, we demonstrate the ability for producer HEK293T cells to simultaneously produce lentiviral particles and transduce (i.e., infect) target cells through a membrane-based coculture system in a continuous, real-time mode which negates the need for a separate viral collection and quantification process. The coculture system was evaluated for major design features such as variations in HEK293T seeding density, target cell type densities, as well as membrane porosities to identify key relationships between lentiviral particle production rate and infection kinetics for adherent and suspension cell types. As a proof-of-concept for the creation of an engineered cell immunotherapy, we describe the ability to engineer human T cells isolated from PBMCs under the control of this coculture system in under 6 days with a GFP construct. These studies suggest the capability to combine and more closely automate the transfection/transduction process in order to facilitate well-timed and cost-effective transduction of target cell types. These experiments provide novel insight into the forthcoming transition into improved manufacturing systems for viral production and subsequent cell engineering.
journal_name
Cytotechnologyjournal_title
Cytotechnologyauthors
Timmins LM,Patel RS,Teryek MS,Parekkadan Bdoi
10.1007/s10616-019-00343-0subject
Has Abstractpub_date
2019-10-01 00:00:00pages
1019-1031issue
5eissn
0920-9069issn
1573-0778pii
10.1007/s10616-019-00343-0journal_volume
71pub_type
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