Mechanistic Study and Target Identification of NNK-Induced Head and Neck Squamous Cell Carcinoma via Systems Bioinformatics, Network Toxicology, and Machine Learning

Background: Head and neck squamous cell carcinoma (HNSCC) is an aggressive malignancy strongly associated with tobacco use. The tobacco-specific nitrosamine NNK is a potent carcinogen, but its molecular mechanisms, particularly its impact on the tumor immune microenvironment and drug resistance in HNSCC, are not fully understood. This study aimed to identify key molecular targets and pathways involved in NNK-induced HNSCC. Methods: We employed an in silico framework integrating systems bioinformatics, network toxicology, and machine learning. Potential targets of NNK and HNSCC-related genes were collected from multiple databases. Core genes were identified from transcriptomic data (GSE107591, GSE138206) through differential expression analysis and machine learning algorithms (XGBoost, Boruta). The stability of the NNK-target interaction was validated using molecular docking and 100 ns molecular dynamics simulations. Immune infiltration and drug sensitivity analyses were also performed. Results: We identified 71 core genes linked to both NNK and HNSCC, from which eight key genes, including monoamine oxidase B (MAOB), were pinpointed as crucial for HNSCC progression. Functional analysis revealed their involvement in the MAPK signaling pathway. MAOB was strongly correlated with an immunosuppressive microenvironment, characterized by increased pro-tumor M2 macrophages and Tregs. Molecular simulations confirmed a stable, high-affinity binding between NNK and MAOB (-7.5 kcal/mol). High MAOB expression was also associated with resistance to MEK inhibitors and Dasatinib. Conclusion: Our computational analysis identifies MAOB as a key mediator in NNK-induced HNSCC, playing a dual role in promoting immune evasion and drug resistance. These findings establish MAOB as a promising therapeutic target and offer new strategies for overcoming treatment resistance in smoking-related HNSCC.