Zapnometinib and IAV H3N2/Fukui infection remodel HLA-I ligandome in human lung cells, modulating antiviral responses

Current antiviral therapies for influenza A virus (IAV) face challenges like drug resistance and limited efficacy, necessitating novel approaches. Host-directed drugs, such as MEK inhibitors, represent a promising alternative by targeting host cellular pathways essential for viral replication or immune response. Zapnometinib, a clinical-stage MEK inhibitor, has demonstrated both antiviral and immunomodulatory properties. However, its specific impact on antigen presentation, particularly at the level of the HLA class I (HLA-I) ligandome, which is crucial for T-cell mediated immunity against viral infections, remained uncharacterized.

Researchers investigated the effects of zapnometinib treatment and Influenza A virus (IAV) H3N2/Fukui infection on HLA-I-presented peptides in human lung adenocarcinoma Calu-3 cells. Cells were infected with IAV and treated with zapnometinib. Surface HLA-I expression was quantified using flow cytometry. Immunopeptidomic analyses, employing label-free LC-MS/MS, were then applied to assess alterations in the ligandome and identify functionally enriched cellular pathways. This approach allowed for a detailed characterization of how the drug and virus independently and synergistically influence antigen presentation.

Neither zapnometinib treatment nor IAV infection significantly altered HLA-I surface expression, with observed fold changes 1.01-1.13 and p > 0.05. However, immunopeptidomics revealed significant, allotype-specific changes in the relative abundance of HLA-I-presented peptides, showing approximately 3-12% change per allotype. Statistically significant modulation was observed in defined ligand subsets, affecting about 3-14% of ligands per condition (with log2 fold change ≥ 2 and adjusted p < 0.05). Functional annotation analysis demonstrated condition-specific enrichment in distinct cellular pathways. Interferon-induced pathways, including IFI16 upregulation during infection, were selectively modulated, with IFI16 downregulation upon MEK inhibition. Additionally, cell cycle and RNA-processing pathways were affected by zapnometinib, and adhesion-related pathways by combined treatment. > Ligandome remodeling after IAV H3N2/Fukui infection strongly affected antiviral hubs (IFIH1, DHX58, IFI16), whereas zapnometinib caused no consistent or significant reduction in these specific hubs.