Identification of Radioresponsive Genes in Esophageal Cancer from Longitudinal and Single Cell Exome Sequencing

The majority (70%) of the esophageal squamous cell carcinoma (ESCC) cases in the world occur in China, where radiotherapy is the most common treatment, yet the majority of ESCC patients still relapse. To better understand the genetic basis of radiotherapy resistance for ESCC, we performed longitudinal, whole-exome sequencing (WES) throughout radiotherapy on 42 patient tumor samples, including single cell whole-exome sequencing (scWES) for 147 cells for two patients. Significant allelic changes were observed during clinical irradiation, with 42 recurrent radioresponsive genes (sensitive and resistant) identified in multiple patients, including NOTCH1, MAML3, CDKN2A, NFE2L2, GAS2L2, OBSCN and TP53, with the last three genes implicated as radioresponsive in both bulk and scWES. Most (37/42) radioresponse genes showed regional variegation of both radioresistant and radiosensitive mutations, with a paucity of resistant-only mutations (2.5%). A subset of sensitive mutations in 10 genes and resistant mutations in 18 genes defined a significantly improved prognosis and the shortest time for the local-region recurrence (LRR) status of the tumors, respectively, indicating possible clinical utility. We also confirmed these significant mutational signatures in orthogonal TCGA ESCC cohorts. Overall, our results quantify the allelic shifts underlying radioresponse in bulk and single-cell ESCC exomes for the first time, provide a temporal resolution to such mutational dynamics, and offer new therapeutic target genes and loci for esophageal and potentially other cancers.