Journal
MOLECULAR THERAPY-METHODS & CLINICAL DEVELOPMENT
Volume 20, Issue -, Pages 30-38Publisher
CELL PRESS
DOI: 10.1016/j.omtm.2020.10.020
Keywords
-
Categories
Funding
- National Institutes of Health [DP1 DA039543, RO1 HD103571]
Ask authors/readers for more resources
The TOP vector is an optimized tool for delivering guide RNAs and transgenes into primary T cells with high transduction efficiency, making it a promising tool for functional screening and immunotherapy applications.
Efficient delivery of nucleic acids for the engineering of primary T cells is central to the study of the basic biology of these key immune effector cells and has clinical implications. To date, lentiviral vectors delivering guide RNAs for CRISPRCas9 editing are not optimal for use in primary cells. Herein, we describe the T cell optimized for packaging (TOP) vector for delivering guide RNAs and transgenes into primary T cells. The TOP vector produces high-titer virus compared to a routinely used guide RNA vector, resulting in a similar to 10-fold increase in transduction in T cells. Moreover, a TOP vector expressing a chimeric antigen receptor and a guide RNA targeting the T cell receptor showed an similar to 5- to 9-fold increased transduction efficiency with similar to 2- to 3-fold higher expression compared to the commonly used epHIV7 vector and was simultaneously able to mediate efficient knockout of the endogenous T cell receptor in >71% of transduced cells upon Cas9 electroporation. The increased packaging of the TOP vector genome into viral particles appears to contribute to its higher transduction efficiency. The TOP vector represents an optimal tool for tandem delivery of transgenes and guide RNAs to primary T cells for use in functional screens and immunotherapy applications.
Authors
I am an author on this paper
Click your name to claim this paper and add it to your profile.
Reviews
Recommended
No Data Available