Structure-activity relationship of T-cell receptors based on alanine scanning
Shogo Narimatsu1, Yasuo Yoshioka, Tomohiro Morishige
1Laboratory of Biotechnology and Therapeutics, Graduate School of Pharmaceutical Sciences, Osaka University, 1-6, Yamadaoka, Suita, Osaka 565-0871, Japan.
Abstract:
T-cell receptors (TCR) recognize complexes between human leukocyte antigens (HLA) and peptides derived from intracellular proteins. Their therapeutic use for antigen targeting, however, has been hindered by the very low binding affinity of TCRs, typically in the 1- to 100-μM range. Therefore, to construct mutant TCRs with high binding affinity, we need to understand the relationship between the structure and activity of these molecules. Here, we attempted to identify the amino acids of the TCR that are important for binding to the peptide/HLA complex. We used a TCR that recognizes complexes between HLA-A(∗)0201 and the peptide from tyrosinase, antigen overexpressed in melanoma. We changed 16 amino acids in the third complementarity-determining region within the TCR to alanine and examined the effect on binding affinity. Five alanine substitutions decreased the binding affinity to below 10% compared with that of wild-type TCR. In contrast, one alanine substitution caused a faster on-rate and slower off-rate, and increased the binding affinity to three times that of the wild-type TCR. Our results provide fundamental information for constructing mutant TCRs with high binding affinity.
Insights
Researchers engineered T-cell receptors (TCRs) to enhance their binding affinity for cancer antigens. One specific mutation significantly boosted TCR binding, offering a promising strategy for developing advanced immunotherapies.
Area of Science:
- Immunology
- Structural Biology
- Biochemistry
Background:
- T-cell receptors (TCRs) are crucial for adaptive immunity, recognizing peptide-MHC complexes.
- Therapeutic applications of TCRs are limited by their low binding affinity (1-100 μM) to target antigens.
- Understanding TCR structure-activity relationships is key to engineering high-affinity variants.
Purpose of the Study:
- To identify specific amino acids in TCRs that modulate binding affinity to peptide-HLA complexes.
- To engineer enhanced TCRs for potential cancer immunotherapy applications.
Main Methods:
- Systematic alanine scanning mutagenesis of 16 amino acids in the TCR's third complementarity-determining region (CDR3).
- Assessing the binding affinity of mutant TCRs to HLA-A*0201 complexed with a tyrosinase-derived peptide.
Main Results:
- Five alanine substitutions significantly reduced TCR binding affinity (<10% of wild-type).
- One specific alanine substitution enhanced binding affinity threefold by increasing the on-rate and decreasing the off-rate.
- Identified key residues in CDR3 critical for TCR-peptide/HLA interaction.
Conclusions:
- Structure-guided engineering of TCR CDR3 can substantially improve binding affinity.
- This study provides fundamental insights for developing high-affinity TCR-based therapeutics for cancer.
- Optimized TCRs hold promise for enhanced antigen targeting in immunotherapies.
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