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Cited by 21CRISPR and Genetic EngineeringCAR-T cell therapy researchImmunotherapy and Immune Responses

Reprogramming MHC specificity by CRISPR-Cas9-assisted cassette exchange

Open access
Kelton, William, Waindok, Ann Cathrin, Pesch, Theresa, Pogson, Mark, Ford, Kyle, Parola, Cristina, Reddy, Sai T.·2017·Scientific Reports
DOI 10.1038/srep45775PMC PMC5379551
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Abstract

The development of programmable nucleases has enabled the application of new genome engineering strategies for cellular immunotherapy. While targeted nucleases have mostly been used to knock-out or knock-in genes in immune cells, the scarless exchange of entire immunogenomic alleles would be of great interest. In particular, reprogramming the polymorphic MHC locus could enable the creation of matched donors for allogeneic cellular transplantation. Here we show a proof-of-concept for reprogramming MHC-specificity by performing CRISPR-Cas9-assisted cassette exchange. Using murine antigen presenting cell lines (RAW264.7 macrophages), we demonstrate that the generation of Cas9-induced double-stranded breaks flanking the native MHC-I H2-Kd locus led to exchange of an orthogonal H2-Kb allele. MHC surface expression allowed for easy selection of reprogrammed cells by flow cytometry, thus obviating the need for additional selection markers. MHC-reprogrammed cells were fully functional as they could present H2-Kd-restricted peptide and activate cognate T cells. Finally, we investigated the role of various donor template formats on exchange efficiency, discovering that templates that underwent in situ linearization resulted in the highest MHC-reprogramming efficiency. These findings highlight a potential new approach for the correcting of MHC mismatches in cellular transplantation.

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  • Reprogramming the polymorphic MHC locus removes the immune barrier and increases transplant acceptance.

    “as shown by removing the immune barrier by reprogramming the polymorphic MHC locus to increase the transplant acceptance (310)”

    From Blood-Brain Barrier: From Physiology to Disease and Back (2019) · cites this paper as [310]

    The Abstract describes MHC reprogramming as a potential way to create matched donors, and the Discussion reports successful allele exchange in cell lines. It does not show that the immune barrier was removed or transplant acceptance increased; the Discussion says in vivo studies are still needed to demonstrate tolerance of the engineered cells. (LLM, conf=0.90)

    Experimental — accuracy not yet measured

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Cited by

Sorted by citations

Authors
Kelton, W., Waindok, A. C., Pesch, T., Pogson, M., Ford, K., Parola, C., & Reddy, S. T.
Affiliations
ETH Zurich (CH)
Corresponding author
Reddy, Sai T.
ORCID
Venue
Scientific Reports
Publisher
Springer Science and Business Media LLC
Published
2017-04-04
Volume
7
Issue
1
Pages
45775-45775
ISSN
2045-2322 (ISSN-L)
DOI
10.1038/srep45775
PMC
PMC5379551
OpenAlex
W2604715983
Cited by
21
References
6
Citations by year
2026: 1 · 2025: 2 · 2024: 4 · 2023: 3 · 2022: 1 · 2021: 2 · 2019: 4 · 2018: 3 · 2017: 1
FWCI
0.89
Type
journal article
Language
en
Open access
Licence
Locations
  • Scientific Reports (publishedVersion, cc-by)
  • PubMed Central (publishedVersion, cc-by)
  • PubMed (publishedVersion)
  • Repository for Publications and Research Data (ETH Zurich) (submittedVersion, cc-by)
  • ETH Zürich Research Collection (cc-by)
Funders
  • National Science Foundation
  • Schweizerischer Nationalfonds zur Förderung der Wissenschaftlichen Forschung
  • Eidgenössische Technische Hochschule Zürich
Topics
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › CRISPR and Genetic Engineering
  • Health Sciences › Medicine › Oncology › CAR-T cell therapy research
  • Life Sciences › Immunology and Microbiology › Immunology › Immunotherapy and Immune Responses
Keywords
CRISPR/Cas9, H-2Kb, MHC class I, allogeneic transplantation, MHC mismatch, antigen-presenting cells, macrophages, T cell activation, genome editing
MeSH terms
Animals, Cell Line, Fibroblasts, Flow Cytometry, H-2 Antigens, Macrophages, Major Histocompatibility Complex, Mice, Inbred BALB C, Cell Line, Tumor, Mice, DNA Breaks, Double-Stranded, CRISPR-Cas Systems

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