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Purification of mRNA Encoding Chimeric Antigen Receptor Is Critical for Generation of a Robust T-Cell Response.

Authors
  • Foster, Jessica B1, 2
  • Choudhari, Namrata3, 4
  • Perazzelli, Jessica1
  • Storm, Julie1
  • Hofmann, Ted J1
  • Jain, Payal3, 4
  • Storm, Phillip B2, 3, 4, 5
  • Pardi, Norbert6
  • Weissman, Drew6
  • Waanders, Angela J1, 2, 4
  • Grupp, Stephan A1, 2
  • Karikó, Katalin7
  • Resnick, Adam C2, 3, 4, 8
  • Barrett, David M1, 2
  • 1 1 Division of Oncology,, The Children's Hospital of Philadelphia, Philadelphia, Pennsylvania.
  • 2 2 Center for Childhood Cancer Research, The Children's Hospital of Philadelphia, Philadelphia, Pennsylvania.
  • 3 3 Division of Neurosurgery, The Children's Hospital of Philadelphia, Philadelphia, Pennsylvania.
  • 4 4 Center for Data-Driven Discovery in Biomedicine, The Children's Hospital of Philadelphia, Philadelphia, Pennsylvania.
  • 5 5 Department of Neurosurgery, Perelman School of Medicine at the University of Pennsylvania.
  • 6 6 Division of Infectious Diseases, Perelman School of Medicine at the University of Pennsylvania, Philadelphia.
  • 7 7 BioNTech RNA Pharmaceuticals, Mainz, Germany. , (Germany)
  • 8 8 Department of Biomedical and Health Informatics, The Children's Hospital of Philadelphia, Philadelphia, Pennsylvania.
Type
Published Article
Journal
Human Gene Therapy
Publisher
Mary Ann Liebert
Publication Date
Feb 01, 2019
Volume
30
Issue
2
Pages
168–178
Identifiers
DOI: 10.1089/hum.2018.145
PMID: 30024272
Source
Medline
Keywords
Language
English
License
Unknown

Abstract

T cells made with messenger RNA (mRNA) encoding chimeric antigen receptor (CAR) offer a safe alternative to those transduced with viral CARs by mitigating the side effects of constitutively active T cells. Previous studies have shown that mRNA CAR T cells are transiently effective but lack persistence and potency across tumor types. It was hypothesized that the efficacy of mRNA CARs could be improved by utilizing recent advancements in RNA technology, such as incorporating a modified nucleoside, 1-methylpseudouridine, into the mRNA and applying a novel purification method using RNase III to eliminate dsRNA contaminants. T cells electroporated with nucleoside-modified and purified mRNA encoding CD19 CAR showed an initial twofold increase in CAR surface expression, as well as a twofold improvement in cytotoxic killing of leukemia cells that persisted up to 5 days. T cells generated with nucleoside-modified and purified CAR mRNA also showed reduced expression of checkpoint regulators and a differential pattern of genetic activation compared to those made with conventional mRNA. In vivo studies using a leukemia mouse model revealed that the most robust 100-fold suppression of leukemic burden was achieved using T cells electroporated with purified mRNAs, regardless of their nucleoside modification. The results provide a novel approach to generate mRNA for clinical trials, and poise mRNA CAR T cells for increased efficacy during testing as new CAR targets emerge.

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