Figure 1: “Reverse Immunology” approach to identify immunogenic cancer neoantigens for designing neoepitope-based immunotherapy. The left flowchart shows the process of identifying cancer neoantigens for target of immunotherapy, consisting of three steps: screening, selection, and validation of the candidate immunogen. The right hierarchy chart* shows the process of narrowing candidate antigens through each step. First, the whole genome/exome sequence profile is comprehensively screened to identify tumor-specific somatic mutations (cancer neoantigens) by massive parallel sequencing of tumor and normal tissues, respectively. Second, computational algorithms are used for predicting the affinity of the mutation-derived peptides with the patient’s own HLA and/or TCR. Third, synthetic mutated peptides and wild-type peptides are used to validate the immunogenicity and specificity of the identified antigens by in vitro T-cell assay or in vivo immunization. Validated antigens could be applied to immunotherapy as either biomarkers of responses to immunotherapy, or as targets of cancer immunotherapy, including cancer vaccines and adoptive T-cell therapy. *Each area does not necessarily illustrate a precise proportion.