RESUMEN
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.
Asunto(s)
Linfocitos T CD4-Positivos/inmunología , Linfocitos T CD8-positivos/inmunología , Neoplasias Colorrectales/genética , Neoplasias Colorrectales/inmunología , ADN Complementario/análisis , Epítopos de Linfocito T/genética , Proteína de la Poliposis Adenomatosa del Colon/genética , Proteínas de Ciclo Celular/genética , Fosfatidilinositol 3-Quinasa Clase I , Análisis Mutacional de ADN , Epítopos de Linfocito T/inmunología , Proteínas F-Box/genética , Proteína 7 que Contiene Repeticiones F-Box-WD , Expresión Génica , Antígenos HLA/genética , Antígenos HLA/inmunología , Ensayos Analíticos de Alto Rendimiento , Humanos , Células Madre Neoplásicas/inmunología , Fosfatidilinositol 3-Quinasas/genética , Proteínas Proto-Oncogénicas p21(ras)/genética , Proteína Smad4/genética , Proteína Smad4/inmunología , Células Tumorales Cultivadas , Proteína p53 Supresora de Tumor/genética , Ubiquitina-Proteína Ligasas/genéticaRESUMEN
Although cancer immunotherapy shows efficacy with adoptive T cell therapy (ACT) and antibody-based immune checkpoint blockade, efficacious therapeutic vaccination of cancer patients with tumor-associated antigens (TAAs) remains largely unmet. Current cancer vaccines utilize nonmutated shared TAAs that may have suboptimal immunogenicity. Experimental evidence underscores the strong immunogenicity of unique TAAs derived from somatically mutated cancer proteins, whose massive characterization has been precluded until recently by technical limitations. The development of cost-effective, high-throughput DNA sequencing approaches makes now possible the rapid identification of all the somatic mutations contained in a cancer cell genome. This method, combined with robust bioinformatics platforms for T cell epitope prediction and established reverse immunology approaches, provides us with an integrated strategy to identify patient-specific unique TAAs in a relatively short time, compatible with their potential use in the clinic. Hence, it is now for the first time possible to quantitatively define the patient's unique tumor antigenome and exploit it for vaccination, possibly in combination with ACT and/or immune checkpoint blockade to further increase immunotherapy efficacy.