Journal article
Engineering improved T cell receptors using an alanine-scan guided T cell display selection system
Journal of immunological methods, Vol.392(1-2), pp.1-11
06/28/2013
DOI: 10.1016/j.jim.2013.02.018
PMCID: PMC3668434
PMID: 23500145
Abstract
T cell receptors (TCRs) on T cells recognize peptide-major histocompatibility complex (pMHC) molecules on the surface of antigen presenting cells and this interaction determines the T cell immune response. Due to negative selection, naturally occurring TCRs bind self (tumor) peptides with low affinity and have a much higher affinity for foreign antigens. This complicates isolation of naturally occurring, high affinity TCRs that mediate more effective tumor rejection for therapeutic purposes. An attractive approach to resolve this issue is to engineer high affinity TCRs in vitro using phage, yeast or mammalian TCR display systems. A caveat of these systems is that they rely on a large library by random mutagenesis due to the lack of knowledge regarding the specific interactions between the TCR and pMHC. We have focused on the mammalian retroviral display system because it uniquely allows for direct comparison of TCR–pMHC-binding properties with T-cell activation outcomes. Through an alanine-scanning approach, we are able to quickly map the key amino acid residues directly involved in TCR–pMHC interactions thereby significantly reducing the library size. Using this method, we demonstrate that for a self-antigen-specific human TCR (R6C12) the key residues for pMHC binding are located in the CDR3β region. This information was used as a basis for designing an efficacious TCR CDR3α library that allowed for selection of TCRs with higher avidity than the wild-type as evaluated through binding and activation experiments. This is a direct approach to target specific TCR residues in TCR library design to efficiently engineer high avidity TCRs that may potentially be used to enhance adoptive immunotherapy treatments.
•Alanine scanning of TCR CDR3 regions for optimal design of TCR display library•Conduct analysis and selected T-cell clones based on functional T-cell potency•Generate T cell clones with increased binding affinity and increased functionality•Use mammalian display system to generate specific and cross-reactive T cell clones•TCR binding avidity and off-rate determine functionality for specific T cell clones.
Details
- Title: Subtitle
- Engineering improved T cell receptors using an alanine-scan guided T cell display selection system
- Creators
- Karolina Malecek - New York UniversityShi Zhong - NYU Cancer institute, New York University School of Medicine, USAKatelyn McGary - New York UniversityConnie Yu - New York UniversityKevin Huang - New York UniversityLaura A. Johnson - National Cancer InstituteSteven A. Rosenberg - National Institutes of HealthMichelle Krogsgaard - New York University
- Resource Type
- Journal article
- Publication Details
- Journal of immunological methods, Vol.392(1-2), pp.1-11
- Publisher
- Elsevier B.V
- DOI
- 10.1016/j.jim.2013.02.018
- PMID
- 23500145
- PMCID
- PMC3668434
- ISSN
- 0022-1759
- eISSN
- 1872-7905
- Grant note
- name: National Institute of Health, award: U01CA137070, R01GM085586; name: American Cancer Society Research Scholar, award: RSG-09-070-01-LIB; name: Cancer Research Investigator grant; name: Pew Scholar in Biomedical Sciences; name: Pew Trust; name: American Heart Association Fellowship; name: NIH T32 Molecular Biophysics Training Grant
- Language
- English
- Date published
- 06/28/2013
- Academic Unit
- Surgery
- Record Identifier
- 9984502460202771
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