Characterization of alternative splicing events in HPV-negative head and neck squamous cell carcinoma identifies an oncogenic DOCK5 variant

Purpose: Head and neck squamous cell carcinoma (HNSCC) is one of the most common cancers worldwide, and alternative splicing is considered to play important roles in tumor progression. Our study is designed to identify alternative splicing events (ASEs) in human papillomavirus (HPV)-negative HNSCC. Experimental Design: RNA sequencing data of 407 HPVnegative HNSCC and 38 normal samples were obtained from The Cancer Genome Atlas (TCGA), and splice junctions were discovered using MapSplice. Outlier analysis was used to identify significant splicing junctions between HPV-negative HNSCC and normal samples. To explore the functional role of the identified DOCK5 variant, we checked its expression with qRT-PCR in a separate primary tumor validation set and performed proliferation, migration, and invasion assays. Results: A total of 580 significant splicing events were identified in HPV-negative HNSCC, and the most common type of splicing events was an alternative start site (33.3%). The prevalence of a given individual ASE among the tumor cohort ranged from 9.8% and 64.4%. Within the 407 HPV-negative HNSCC samples in TCGA, the number of significant ASEs differentially expressed in each tumor ranged from 17 to 290. We identified a novel candidate oncogenic DOCK5 variant confirmed using qRT-PCR in a separate primary tumor validation set. Lossand gain-of-function experiments indicated that DOCK5 variant promoted proliferation, migration, and invasion of HPV-negative HNSCC cells, and patients with higher expression of DOCK5 variant showed decreased overall survival. Conclusions: Analysis of ASEs in HPV-negative HNSCC identifies multiple alterations likely related to carcinogenesis, including an oncogenic DOCK5 variant.