Computational protocol: Identification of human leukemia antigen A*0201 restricted epitopes derived from epidermal growth factor pathway substrate number 8

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Protocol publication

[…] To identify the potential T-cell epitopes for EPS8, the HLA-A*0201 allele was screened, covering a wide range of populations (). First, the protein sequence was screened for the best binding epitopes using the algorithms SYFPEITHI (http://www.syfpeithi.de/) and Bioinformatics and Molecular Analysis Section (BIMAS; http://www-bimas.cit.nih.gov/). The cut-off score was adjusted to >20 for SYFPEITHI and T1/2>100 for BIMAS, and peptides matching these two criteria and which were shared by the two algorithms were selected. Second, the NetChop algorithm (http://www.cbs.dtu.dk/services/NetChop/) was used to predict whether the selected peptides in the first step would be cleaved at the C-terminus. Third, protein sequences were analyzed with five different algorithms for HLA-A*0201, including immune epitope database (IEDB; http://www.iedb.org/), NetMHC (http://www.cbs.dtu.dk/services/NetMHC/), SVMHC (http://www.sbc.su.se/~pierre/svmhc/), EpiJen (http://www.ddgpharmfac.net/epijen/EpiJen/EpiJen.htm) and Rankpep (http://imed.med.ucm.es/Tools/rankpep.html). Peptides predicted by at least four algorithms were selected. The known HLA-A*0201-restricted carcinoembryonic antigen peptide-1 (CAP-1, YLSGANLNL) was used as a positive control peptide (), while the HBcAg-derived HLA-A*2402-restricted peptide HBc117e125 (EYLVSFGVW) was used as a negative control peptide (), and all peptides were synthesized using standard 9-fluorenylmethyl-oxycarbonyl chemistry by the Chinese Peptide Company (Hangzhou, China). They were of >95% purity as determined by reverse-phase chromatography and their identity was confirmed by mass spectrometry. The lyophilized peptide powder was dissolved according to the manufacturer’s instructions in ultrapure water or in dimeth-ylsulfoxide (BioLegend), diluted in phosphate-buffered saline (PBS; Invitrogen Life Technologies) to a final concentration of 1 mg/ml and stored in aliquots at −80°C until use. […]

Pipeline specifications

Software tools NetChop, NetMHC, SVMHC, EpiJen, RANKPEP
Databases SYFPEITHI IEDB
Organisms Homo sapiens
Diseases Leukemia, Neoplasm Metastasis, Neoplasms, Hematologic Neoplasms, Drug-Related Side Effects and Adverse Reactions
Chemicals Brefeldin A