Next-Generation Sequencing–Based Molecular Profiling of Conjunctival Squamous Cell Carcinoma and Its Potential Application for Therapy

Objective

Targeted next-generation sequencing–based genomic and transcriptomic analyses of conjunctival squamous cell carcinoma (cSCC) samples using a panel of >1700 cancer-related genes.

Design

Prospective case series.

Participants

Twenty patients with invasive cSCC consecutively managed at an academic ocular oncology setting.

Methods

Integrative exome and transcriptome analysis of fresh-frozen tumor and matching normal samples.

Main Outcome Measures

Molecular characterization of invasive cSCCs (for somatic mutations, tumor mutation burden (TMB) and signatures, structural variations, viral transcripts, outlier gene expression) and its potential clinical applications.

Results

Of the 20 invasive cSCCs, only 2 were positive for human papillomavirus (HPV). All 18 HPV-negative tumors had genetic alterations in TP53 and 16 also had alterations in CDKN2A. The 2 HPV-positive tumors showed no alterations in these cell cycle regulators but harbored mutations in PIK3CA. Other frequently altered genes included KMT2C/D (70%), FAT1/3 (65%), and NOTCH1/2/3 (60%), which were implicated in both the HPV-negative and positive tumors. The average TMB was 58.41 mutations per megabase (Mut/Mb). The TMB was >20 Mut/Mb in 13 cases (65%, range: 49.3–160.8 Mut/Mb), of which 11 had ultraviolet (UV) mutational signature, 3 had APOBEC signature, and 2 had microsatellite instability. Most UV-driven tumors (8 of 11) also harbored TERT promoter mutations. The most recurrent large-scale copy number alterations were the deletions affecting chromosomes 3p, 9p, and 14q. Uncommon but potentially drug-targetable copy number gains were also detected affecting several oncogenes. The most frequent gene expression outlier was the aberrantly expressed TP63 found in 19 tumors.

Conclusions

In our invasive cSCC cohort, HPV infection was not a major contributor to tumor etiopathogenesis. Our results confirmed the central role of TP53 genetic alterations and the common presence of UV signature in the HPV-negative cSCCs. Irrespective of HPV status, other commonly observed genetic alterations included those affecting chromatin modifiers followed by Hippo or Notch pathway–related genes. Ultraviolet-driven TERT promoter mutations co-occurred with other driver gene alterations. The commonly observed high TMB makes immunotherapy a good treatment choice for invasive cSCC. Moreover, several cSCC-associated molecular alterations represent potentially actionable targets, while further studies are necessary to understand their roles in cSCC development and invasion.

Financial Disclosure(s)

Proprietary or commercial disclosure may be found in the Footnotes and Disclosures at the end of this article.