Pathogenic somatic alterations in advanced HPV-negative cell squamous laryngeal carcinoma revealed via targeted next generation sequencing

Abstract

During the last decade, next generation sequencing (NGS) became the major tool for detection of somatic mutations in cancer, but data about mutations in advanced laryngeal squamous carcinomas is still scarce. The aim of this study is to analyze the mutation profile of key oncogenes and tumour suppressor genes in advanced HPV-negative laryngeal squamous cell carcinoma. A total of 57 Bulgarian LSCC patients were included. DNA was isolated from fresh-frozen tissues. Targeted NGS was performed using TruSeq Amplicon Panel on Illumina platform, and data was analysed with VarSeq Software. Results revealed altogether 92 known pathogenic and likely pathogenic variants in 27 tumour-associated genes. Thirteen new variants were predicted to be pathogenic with four or more prediction programs. The most frequently mutated gene was TP53, with mutations in 84.2%, followed by MET in 19.3%, CDKN2A in 15.8%, PIK3CA in 14% and FBXW7 in 8.8% patients. Interestingly in eight of the supraglottic LSCC patients we found two TP53 mutations and in one subglottic LSCC patient - three TP53 mutations. For the first time the mutational spectrum of three LSCC sub-locations was analyzed and the supraglottis LSCC showed more mutated genes (n=20) compared to glottis (n=10) and subglottis (n=12) tumours. In 9% of the samples we found combined high-risk TP53 and RAS mutations, previously associated with poor overall survival and drug resistance. The analysis revealed that NOTCH1 mutations are not common for LSCC in comparison to other HNC loci. In addition, we found three common polymorphisms in TP53 (p.Pro72Arg), KDR (p.Gln472His) and KIT (p.Met541Leu) genes, strongly associated with angiogenesis, poor prognosis and metastasis and drug sensitivity in cancer diseases. In conclusion, NGS targeted sequencing enables discovery of new mutations in key genes relevant to LSCC, some of which might be useful in selection or development of more precise LSCC treatment approaches in the future.