The landscape of long noncoding RNA during cutaneous squamous cell carcinoma progression

Cutaneous squamous cell carcinoma (cSCC) is a common cancer with a high morbidity rate and poor prognosis for metastatic disease. Disease may progress from premalignant actinic keratosis to invasive and metastatic cSCC, but it is perhaps best characterized as a disease continuum progressing from a differentiated to a progenitor-like state. The critical molecular mediators of this process remain poorly defined. Long noncoding (lnc)RNAs, a relatively unexplored class of RNA molecules > 200 nucleotides long, are likely to have important functional roles in cSCC.

To provide a comprehensive landscape of lncRNA expression during the cSCC continuum and to identify potentially functional lncRNA drivers of disease progression.

We interrogated bulk RNA sequencing (RNAseq) data from 110 patient samples, encompassing healthy sun-exposed skin (n = 26), actinic keratosis (n = 14), primary cSCC (n = 66) and metastases (n = 4), to identify changes in lncRNA expression during disease progression. We developed a bioinformatics pipeline to infer lncRNA function based on co-expression patterns and generated a lncRNA signature score, which we validated in head-and-neck squamous cell carcinoma (HNSC) and pancreatic adenocarcinoma (PAAD). We performed bulk RNAseq on 15 patient-derived cell lines and integrated these data to identify tumour cell-specific lncRNAs and validated our findings in multiple other cSCC gene expression cohorts. Using in vitro knockdown approaches we investigated the functional role of LINC00941.

We found that lncRNA expression alone is sufficient to identify disease states and progression along the cSCC disease continuum. Correlation analysis revealed potentially functionally relevant lncRNAs and the processes they may regulate. We developed a 267 lncRNA signature that correlates with a progenitor-like state and predicts poor prognosis in HNSC and PAAD. Bulk RNAseq of patient-derived cell lines revealed tumour cell-specific lncRNAs, and knockdown of LINC00941 indicated that it is required for cell proliferation and colony formation in vitro.

Our findings provide a comprehensive description of lncRNA transcriptomic changes in cSCC and demonstrate their functional relevance as biomarkers and drivers of disease progression in this and, potentially, other cancers.