T cell neoepitope discovery in colorectal cancer by high throughput profiling of somatic mutations in expressed genes

D Mennonna, C Maccalli, MC Romano, Claudio Garavaglia, F Capocefalo, R Bordoni, M Severgnini, Gianluca De Bellis, J Sidney, Alessandro Sette, A Gori, Renato Longhi, M Braga, Luca Ghirardelli, L Baldari, E Orsenigo, L Albarello, E Zino, K Fleischhauer, G MazzolaN Ferrero, Antonio Amoroso, G Casorati, G Parmiani, P Dellabona

Research output: Contribution to journalArticlepeer-review


OBJECTIVE: Patient-specific (unique) tumour antigens, encoded by somatically mutated cancer genes, generate neoepitopes that are implicated in the induction of tumour-controlling T cell responses. Recent advancements in massive DNA sequencing combined with robust T cell epitope predictions have allowed their systematic identification in several malignancies. DESIGN: We undertook the identification of unique neoepitopes in colorectal cancers (CRCs) by using high-throughput sequencing of cDNAs expressed by standard cancer cell cultures, and by related cancer stem/initiating cells (CSCs) cultures, coupled with a reverse immunology approach not requiring human leukocyte antigen (HLA) allele-specific epitope predictions. RESULTS: Several unique mutated antigens of CRC, shared by standard cancer and related CSC cultures, were identified by this strategy. CD8+ and CD4+ T cells, either autologous to the patient or derived from HLA-matched healthy donors, were readily expanded in vitro by peptides spanning different cancer mutations and specifically recognised differentiated cancer cells and CSC cultures, expressing the mutations. Neoepitope-specific CD8+ T cell frequency was also increased in a patient, compared with healthy donors, supporting the occurrence of clonal expansion in vivo. CONCLUSIONS: These results provide a proof-of-concept approach for the identification of unique neoepitopes that are immunogenic in patients with CRC and can also target T cells against the most aggressive CSC component. Published by the BMJ Publishing Group Limited. For permission to use (where not already granted under a licence) please go to http://www.bmj.com/company/products-services/rights-and-licensing/
Original languageEnglish
Pages (from-to)454-463
Number of pages10
Issue number3
Publication statusPublished - 2017


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