Different criteria have been used to pick predicted peptides. In our case, along with a binding threshold for all mutated peptides, we distinguished two groups relating to the binding capability of the WT peptide. In the primary group , the place WT peptides have been poor or null binders, an necessary proportion of peptides contained the mutation at HLA-anchoring residues (e.g. peptides CAM69-6, CAM73-9). However, within the second group all peptides contained the mutation at non-anchor residues. Indeed, it has been just lately shown the relevance of the position of the mutated amino acid in neoAg selection .

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Library was subjected to paired-end sequencing utilizing the NovaSeq platform (10 Gb 100PE 100Mreads/sample). Alignments had been performed using the STAR technique to human genome build hg38. Duplicates were marked and Sorted utilizing Picard’s MarkDuplicates software. Reads were then split and trimmed using GATK SplitNTrim device. In/Del realignment and base recalibration were carried out utilizing GATK toolbox. Gene counts have been calculated utilizing featureCount , Cufflinks and HTseq .

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