%0 Journal Article
%A Woodruff, Rosie
%A Parekh, Farhaan
%A Lamb, Katarina
%A Mekkaoui, Leila
%A Allen, Christopher
%A Smetanova, Katerina
%A Huang, Jasmine
%A Williams, Alex
%A Toledo, Gerardo Santiago
%A Lilova, Koki
%A Roddie, Claire
%A Sillibourne, James
%A Pule, Martin
%D 2023
%F discovery:10180184
%I Elsevier BV
%J Molecular Therapy: Methods & Clinical Development
%K base editing, BE4max, chimeric antigen receptor, circular RNAPD-1TIM3
%T Large-scale manufacturing of base-edited chimeric antigen receptor T cells
%U https://discovery.ucl.ac.uk/id/eprint/10180184/
%V 31
%X Base editing is a revolutionary gene-editing technique enabling the introduction of point mutations into the genome without generating detrimental DNA double-stranded breaks. Base-editing enzymes are commonly delivered in the form of modified linear messenger RNA (mRNA) that is costly to produce. Here, we address this problem by developing a simple protocol for manufacturing base-edited cells using circular RNA (circRNA), which is less expensive to synthesize. Compared with linear mRNA, higher editing efficiencies were achieved with circRNA, enabling an 8-fold reduction in the amount of RNA required. We used this protocol to manufacture a clinical dose (1 × 108 cells) of base-edited chimeric antigen receptor (CAR) T cells lacking expression of the inhibitory receptor, PD-1. Editing efficiencies of up to 86% were obtained using 0.25 μg circRNA/1 × 106 cells. Increased editing efficiencies with circRNA were attributed to more efficient translation. These results suggest that circRNA, which is less expensive to produce than linear mRNA, is a viable option for reducing the cost of manufacturing base-edited cells at scale.
%Z © 2023 The Authors. Published by Elsevier under a Creative Commons license (http://creativecommons.org/licenses/by-nc-nd/4.0/).