Neoantigens heterogeneity as a prognostic marker in patients with resected intrahepatic cholangiocarcinoma

Background: The prognostic significance of neoantigens and the understanding of neoantigen-associated heterogeneity in intrahepatic cholangiocarcinoma (iCCA) is still unclear. This study aims to investigate the prognostic role of neoantigen s and their intratumor heterogeneity (ITH) in postoperative prognoses of patients with resected iCCA. Methods: A cohort enrolled 68 iCCA patients underwent surgical resections was performed whole-exome sequencing to analyze the impact of neoantigens on postoperative survival. Immunohistochemistry for CD4 and CD8 was performed and T cells in tumor were quantified by visually counted. RNA sequencing and TCR sequencing were performed to demonstrate tumor immunogenicity and immune recognition associated with neoantigen heterogeneity. Results: The neoantigen intratumor heterogeneity (NeoITH) metric was an independent prognostic factor, with NeoITH-low iCCA associated with improved prognosis in the discovery and the validation cohort. Immunogenetic analyses revealed that NeoITH-low iCCA harbored an increased CD8 + T cell infiltration and higher immunogenicity compared to NeoITH-high iCCA. Characterization on the TCR repertoire identified 36 iCCA-related TCR clusters which were enriched in NeoITH-low tumors. Patients identified as the high number of iCCA-related TCR clusters and NeoITH-low iCCAs had the best postoperative prognosis, and vice versa. A retrospective immunotherapy cohort confirmed that patients with NeoITH-low iCCAs had significant therapeutic advantages then patients with NeoITH-high iCCAs. Conclusions: This study provides a novel insight into neoantigen heterogeneity for predicting postoperative survival and immunotherapeutic prognosis in iCCA patients.