Identification of an Epithelial Disease Signature for Recurrent Respiratory Papillomatosis Enables Drug Repurposing Studies

Introduction: Recurrent Respiratory Papillomatosis (RRP) causes significant morbidity and healthcare burden, costing $40-$120 million per year in the US. Historically treatment has relied heavily on surgical management, yet surgeries are non-curative and patients can face up to hundreds in their lifetime. Transcriptomic techniques were applied to papilloma vs non-diseased primary epithelial cells to better understand differentially expressed pathways and enable identification of candidate therapeutics for a drug repurposing screen.

Methods: Bulk RNA sequencing was performed on matched 2D primary cells derived from papilloma versus non-diseased patient tissues. Raw fastq sequences were aligned to the hg19 genome, transcript abundances calculated, and differential expression analysis performed in R using established pipelines. Enrichr was used for functional enrichment analysis of differentially expressed genes. Candidate therapeutics inversely matching the disease signature were identified using CLUE v1.0, EMUDRA, and iLINCS tools and ranked. Top candidates were tested for efficacy using patient-derived 2D cells and 3D organotypic epithelial rafts, measured by viability of non-diseased vs papilloma tissues.

Results: A shared transcriptome signature across patients was defined using RNA-sequencing on matched 2D primary cells derived from papilloma vs non-diseased patient tissues. Pathway analysis highlighted significance for adhesion and epithelial to mesenchymal transition (EMT) upregulation. The top six FDA-approved therapeutic candidates were identified for testing using a novel pipeline. Differences in viability of patient-derived papilloma vs non-diseased specimens were found to be highly variable in cultured 3D organotypic rafts and were not observed in 2D primary cells.

Conclusions: Altogether, these studies reveal insight into pathways upregulated in epithelial cells harboring RRP disease that may be targeted with drug repurposing. Initial drug screenings highlight the importance of model selection in studies of HPV-driven disease.