Investigating the Feasibility of Targeted Next-Generation Sequencing to Guide the Treatment of Head and Neck Squamous Cell Carcinoma

Abstract

Purpose: Head and neck squamous cell carcinoma (HNSCC) is a deadly disease in which precision medicine needs to be incorporated. We aimed to implement next-generation sequencing (NGS) in determining actionable targets to guide appropriate molecular targeted therapy in HNSCC patients.

Materials and methods: Ninety-three tumors and matched blood samples underwent targeted sequencing of 244 genes using the Illumina HiSeq 2500 platform with an average depth of coverage of greater than 1,000×. Clinicopathological data from patients were obtained from 17 centers in Korea, and were analyzed in correlation with NGS data.

Results: Ninety-two of the 93 tumors were amenable to data analysis. TP53 was the most common mutation, occurring in 47 (51%) patients, followed by CDKN2A (n=23, 25%), CCND1 (n=22, 24%), and PIK3CA (n=19, 21%). The total mutational burden was similar between human papillomavirus (HPV)-negative vs. positive tumors, although TP53, CDKN2A and CCND1 gene alterations occurred more frequently in HPV-negative tumors. HPV-positive tumors were significantly associated with immune signature-related genes compared to HPV-negative tumors. Mutations of NOTCH1 (p=0.027), CDKN2A (p < 0.001), and TP53 (p=0.038) were significantly associated with poorer overall survival. FAT1 mutations were highly enriched in cisplatin responders, and potentially targetable alterations such as PIK3CA E545K and CDKN2A R58X were noted in 14 patients (15%).

Conclusion: We found several targetable genetic alterations, and our findings suggest that implementation of precision medicine in HNSCC is feasible. The predictive value of each targetable alteration should be assessed in a future umbrella trial using matched molecular targeted agents.

Keywords: Biomarkers; Clinical trial; Molecular Targeted Therapy; Next-generation sequencing; Squamous cell carcinoma of the head and neck.

Fig. 1.

(A) Mutational spectrum and copy number alterations in head and neck squamous cell carcinomas detected by targeted sequencing. Samples with a greater than 1% incidence of genetic alterations are shown, and are stratified by human papillomavirus (HPV) status and primary tumor anatomic site. Pos, positive; Neg, negative. (B) A heat map of 55 differentially expressed genes with an absolute fold change ≥ 2 and a false discovery rate (FDR) < 0.05. (C) Volcano plot showing the distribution of the fold changes in gene expression. Genes with an absolute fold change ≥ 2 and FDR < 0.05 are indicated in red (high expression in HPV-positive tumors compared to HPV-negative tumors).

Fig. 2.

Kaplan-Meier curves showing the association of single nucleotide variations and overall survival (OS) in patients. (A) Patients with NOTCH1 somatic mutation had poorer overall survival (somatic mutation includes missense, nonsense, splice site mutations, frame shift indels, or in-frame indels). (B) Patients with CDKN2A missense mutations had poorer OS. (C) Patients with TP53 nonsense mutation showed poorer OS.

Fig. 3.

Patients who received cisplatin-based chemotherapy were categorized into responders vs. non-responders and genetic alterations are shown in the order of frequency. Pos, positive; Neg, negative.

Fig. 4.

Gene diagrams for a selection of key mutations in potentially targetable genes PIK3CA (A), CDKN2A (B), and TP53 (C). (D) Signaling pathway deregulation is shown. HPV, human papillomavirus.