Increasing evidence has demonstrated the critical roles of
mRNA modification regulators on multiple types of
cancers. However, it is still poorly known about the prognostic and therapeutic value of
mRNA modification regulators in
HNSCC.
METHODS: The gene expression profile of 36
mRNA modification regulators and their corresponding clinical data were obtained from The
Cancer Genome Atlas (TCGA) and Gene Expression Omnibus (GEO). Stepwise regression in R with both directions was used to construct a model for the prognosis of
HNSCC. Univariate Cox regression survival analysis was performed to identify the most significant risk gene. Gene set enrichment analysis (GSEA) was applied to determine the
cancer-associated pathways with NAT10. Immunohistochemistry (IHC) staining was performed to evaluate the expression of NAT10 in
formalin fixed
paraffin-embedded (FFPE) samples of
HNSCC. Univariate and multivariate Cox regression survival analysis performed to identify the independent risk factors associated with the OS of patients with
HNSCC.
HNSCC cell lines (Cal-27, FaDu, and Detroit-562) were transfected with
short interfering RNA (
siRNA) targeting NAT10 or treated with
Remodelin, a small-molecule inhibitor of NAT10. Knockdown efficiency of
siRNA was assessed by quantitative real-time PCR (qRT-PCR) and western blotting. In addition,
CCK-8 assay, scratch assay and transwell assay were used to examine the proliferation, migration, and invasion abilities of the three
HNSCC cell lines after NAT10 was inhibited genetically and pharmaceutically. Cell cycle and cell apoptosis assays were performed by flow cytometry. Finally, the therapeutic value of
Remodelin in
HNSCC was evaluated via a patient-derived xenograft (PDX) model. The statistical analysis was performed with SPSS 23.0.
RESULTS: A risk prediction model containing 10
mRNA modification regulators was constructed and showed prognostic value in
HNSCC. NAT10 was further identified as a key risk gene and independent prognostic factor in TCGA
HNSCC dataset. The GSEA analysis suggested that high NAT10 expression was associated with MYC, E2F, G2M checkpoint,
mTORC1, DNA repair and oxidative phosphorylation pathways. NAT10
protein expression was significantly up-regulated in tumour cells compared to normal epithelial cells in FFPE samples and increased NAT10
protein expression was correlated with poor overall survival of 267
HNSCC patients. Genetic depletion of NAT10 using
siRNA or chemical inhibition of NAT10 using
Remodelin resulted in reduced cell proliferation, migration and invasion abilities in Cal-27, FaDu and Detroit-562 cells. Knockdown of NAT10 using
siRNA significantly increased cell cycle arrest in S/G2-phase.
Remodelin significantly inhibited tumour growth and tumour cell proliferation in the PDX model of
HNSCC.
CONCLUSIONS: