Description:
Abstract Introduction Accurate prediction of T-cell epitopes have remained a bottleneck for developing peptide vaccines for infections and cancer immunotherapy, especially for major histocompatibility complex class II (MHC II). We previously found that cell-surface expression level of the covalently linked peptide-MHC II complex is a good indicator of MHC-peptide interaction (Miyadera et al. 2015 J Clin Invest). We named this assay as “delta MHC (dMHC) assay” and validated its utility in the prediction of CD4+ T-cell epitopes. Method dMHC assay was performed using engineered fibroblast cells that stably express DRA- or DQA1. The cell line was transduced with pMXs-IG (Kitamura et al. 2003 Exp Hematol) that carries DRB1/3/4/5- or DQB1-peptide fusion construct. Cell surface HLA expression was measured by flow cytometry and normalized to the internal control GFP (Miyadera et al. 2015 J Clin Invest). Results and Discussion We conducted systematic binding analysis for HLA class II (27 alleles: HLA-DRB1,3,4,5, and HLA-DQ) and nine peptides derived from Mycobacterium tuberculosis antigens CFP-10 and ESAT-6 that contain potential CD4+ T-cell epitopes (Nagai et al. 2014 J Immunol Res). The cell surface HLA expression level in the presence of the test peptide were normalized to the expression level of the negative control peptide (g9). The normalized expression level (dMHC-g9) showed overall good correlation with the predicted binding affinity (IC50). Moreover, dMHC-g9 outperforms IC50 in predicting potential T-cell epitopes. These data indicate that dMHC assay may be suitable as a platform for large scale analysis of MHC-peptide interaction and data sharing, which should facilitate improved prediction of CD4+ T-cell epitopes.