Objective: Human papillomavirus (HPV) infection is a significant risk factor for head and neck squamous-cell carcinoma (HNSCC). This study aims to identify specific signaling pathways, biomarkers, and potential targeted drugs for HPV-positive HNSCC through competitive endogenous RNA (ceRNA) network analysis.
Methods: High-throughput transcriptomic sequencing was performed on UPCI:SCC154 (HPV-positive HNSCC), Cal27 (HPV-negative HNSCC), and normal human oral keratinocytes to identify differentially expressed mRNAs and miRNAs associated with HPV-positive HNSCC. The interactions of competitive endogenous RNAs were predicted using TargetScan and miRanda, and protein–protein interactions were predicted using the String database to construct a ceRNA network. Network topology analysis was employed to identify hub genes in HPV-positive HNSCC. Further, upstream miRNAs of the hub genes were screened by integrating multiple databases, including encori, mircode, miRDB, miRTarBase, miRWalk, and picTAR, to construct miRNA-mRNA signaling axes. Drug repositioning analysis was conducted using the Connectivity Map database to predict small-molecule drugs targeting the hub genes. Molecular docking and dynamics simulations were used to evaluate the binding affinity and stability of the small molecules with the target proteins. The efficacy of the drugs was validated through cellular functional assays.
Results: A total of 3,253 differentially expressed mRNAs and 391 miRNAs were identified in HPV-positive HNSCC, and RPAP2 was identified as a hub gene. Integrated multi-database analysis revealed that hsa-miR-181b-3p potentially targets and regulates RPAP2. Drug repositioning screening identified five potential drugs, among which molecular docking and dynamics simulations indicated that Enzastaurin exhibited excellent binding affinity and stability with RPAP2. Cellular experiments demonstrated that, compared to cisplatin, Enzastaurin significantly reduced the viability of HPV-positive HNSCC cells and suppressed the expression levels of RPAP2 and hsa-miR-181b-3p.
Conclusion: The miR-181b-3p/RPAP2 axis plays a key regulatory role in HPV-positive HNSCC. Enzastaurin demonstrates promising therapeutic potential, offering a foundation for targeted drug development.
