Expression profiling of primary and metastatic oral squamous cell carcinoma identifies progression-associated transcriptome changes and therapeutic vulnerabilities

Oral squamous cell carcinoma (OSCC) constitutes a subset of head and neck squamous cell carcinoma primarily caused by the excessive consumption of alcohol, tobacco, and areca nut. According to the latest estimates of the global burden of cancer (GLOBOCAN 2022), cancers of the lip and oral cavity account for 389,485 new cancer cases and 188,230 deaths annually. Despite significant progress in OSCC diagnosis and therapy only about 65% of patients survive five years or longer. This poor prognosis is mainly due to the fact that about half of OSCC patients are diagnosed with advanced disease. Hence, a better understanding of the molecular mechanisms responsible for the development and metastatic progression of OSCC is needed in order to identify novel biomarkers and therapeutic targets. We isolated RNA from 87 primary tumours and 21 lymph node metastases from a total of 72 OSCC patients. The RNA was subjected to next-generation sequencing (~62 Mio. reads per sample) upon depletion of ribosomal RNA. Comprehensive transcriptome-wide expression and correlation analyses were performed in order to identify expression subtypes, disease-associated genes and isoforms. The functional and therapeutic relevance of selected candidates was further investigated in vitro using gain- and loss-of-function analyses. Unsupervised clustering of primary tumours identified three clusters (C1-C3) that show different overall survival (OS) probabilities, especially for C3. Furthermore, gene-level analyses identified SHMT2 as a prognostic biomarker associated with poor OS. Importantly, targeting of SHMT2 using siRNAs or a small molecule inhibitor (SHIN1/RZ-2994) decreased cell proliferation and induced apoptosis in a panel of OSCC cell lines and establish SHMT2 as a promising therapeutic target. Comparison of primary tumours and their matched lymph node metastases identified additional druggable metastasis-associated genes as well as several isoform switches. For example, the canonical full-length isoform of WNT5A was higher expressed in metastases and its overexpression in SAS enhanced cell invasion capacity. Our profiling study provides a valuable resource for the community that will foster functional analyses to gain insights into the molecular mechanisms of OSCC progression, and it will help to identify novel biomarkers and therapeutic targets in the future.