Mutational Landscape of Esophageal Squamous Cell Carcinoma in an Indian Cohort

Abstract

Esophageal squamous cell carcinoma (ESCC) is the most common histological subtype of esophageal cancer in India. Cigarette smoking and chewing tobacco are known risk factors associated with ESCC. However, genomic alterations associated with ESCC in India are not well-characterized. In this study, we carried out exome sequencing to characterize the mutational landscape of ESCC tumors from subjects with a varied history of tobacco usage. Whole exome sequence analysis of ESCC from an Indian cohort revealed several genes that were mutated or had copy number changes. ESCC from tobacco chewers had a higher frequency of C:G > A:T transversions and 2-fold enrichment for mutation signature 4 compared to smokers and non-users of tobacco. Genes, such as TP53, CSMD3, SYNE1, PIK3CA, and NOTCH1 were found to be frequently mutated in Indian cohort. Mutually exclusive mutation patterns were observed in PIK3CA-NOTCH1, DNAH5-ZFHX4, MUC16-FAT1, and ZFHX4-NOTCH1 gene pairs. Recurrent amplifications were observed in 3q22-3q29, 11q13.3-q13.4, 7q22.1-q31.1, and 8q24 regions. Approximately 53% of tumors had genomic alterations in PIK3CA making this pathway a promising candidate for targeted therapy. In conclusion, we observe enrichment of mutation signature 4 in ESCC tumors from patients with a history of tobacco chewing. This is likely due to direct exposure of esophagus to tobacco carcinogens when it is chewed and swallowed. Genomic alterations were frequently observed in PIK3CA-AKT pathway members independent of the history of tobacco usage. PIK3CA pathway can be potentially targeted in ESCC which currently has no effective targeted therapeutic options.

Keywords: esophageal cancer; mutation signatures; squamous cell carcinoma; tobacco; whole exome sequencing.

Figure 1

Mutational landscape of esophageal squamous cell carcinoma. Top panel depicts mutation load per Mb, middle panel depicts sample details including tumor grade, gender, age, and tobacco consumption history. Each column represents a sample and each row depicts a gene. Somatic mutations are colored based on mutation type and left panel depicts percentage of samples that harbor mutations.

Figure 2

Examples of mutated genes that showed mutual exclusivity. Schematic representation of coding variant distribution on proteins (A) PIK3CA, (B) NOTCH1, (C) ZFHX4, (D) FAT1, (E) DNAH5, and (F) MUC16. Green indicates missense mutation, black indicates splice site and purple indicates silent mutation. Total number of different type of mutations for each gene is given within the circle on right side. X-axis represents length of the protein (amino acid) and y-axis represents number of samples. Orange flame indicates COSMIC hotspot mutations. Instances of gene pairs with mutually exclusive mutation pattern in ESCC (G–J) PIK3CA and NOTCH1, MUC16 and FAT1, DNAH5 and ZFHX4, ZFHX4 and NOTCH1. Column represents samples and row represents genes. Dark shaded circles indicate the mutant allele and the gray circle indicates wild type allele.

Figure 3

Mutation load observed in tumors from tobacco users and non-users in this study and publicly available datasets of esophageal squamous cell carcinoma (ESCC), oral squamous cell carcinoma (OSCC), lung squamous cell carcinoma (LUSC), and lung adenocarcinoma (LUAD). Black dots represents outliers identified using 97.5 percentile interval. Mutation burden for each sample in ESCC, OSCC, LUSC, and LUAD datasets is provided in Supplementary Table 5 and plotted as per tobacco consumption history.

Figure 4

Analysis of mutation signatures enriched in ESCC from smokers, chewers and tobacco non-users. (A) Mutation signatures associated with ESCC from smokers, chewers and tobacco non-users. Height of the bar represents contribution of the base substitution across different trinucleotide contexts. (B) Mutation load of C > A transversions and C > T transitions in smokers, chewers and tobacco non-users. (C) Results of mutation signature decomposition in ESCC samples from chewers, smokers and tobacco non-users. (Red) Chewers, (Gray) Non-users, and (Orange) Smokers.

Figure 5

Schematic depicting recurrent genomic amplification in cytoband 3q22-3q29 in esophageal squamous cell carcinoma samples.

Figure 6

Schematic representation of PIK3CA/AKT pathway depicting genes altered in ESCC. The numbers beside rectangles indicate percentage of samples that carry mutations (pink) and/or copy number variations (white) in respective genes. Green color represents genes with both mutations and CNVs, yellow color represents genes with only CNVs, and blue color are genes with only mutations.