%A Rosie Woodruff
%A Farhaan Parekh
%A Katarina Lamb
%A Leila Mekkaoui
%A Christopher Allen
%A Katerina Smetanova
%A Jasmine Huang
%A Alex Williams
%A Gerardo Santiago Toledo
%A Koki Lilova
%A Claire Roddie
%A James Sillibourne
%A Martin Pule
%T Large-scale manufacturing of base-edited chimeric antigen receptor T cells
%J Molecular Therapy: Methods & Clinical Development
%K base editing, BE4max, chimeric antigen receptor, circular RNAPD-1TIM3
%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.
%L discovery10180184
%I Elsevier BV
%C United States
%V 31
%O © 2023 The Authors. Published by Elsevier under a Creative Commons license (http://creativecommons.org/licenses/by-nc-nd/4.0/).
%D 2023