In this study, we found an overall upregulation of EIF5A2 mRNA and the transcripts of several key enzyme genes involved in polyamine metabolism, e.g., ODC1, SRM, and SMOX, in HPV− HNSCC, and blocking DHPS/eIF5Ahpu consistently inhibited OSCC cell growth and the expression of genes involved in regulation of epithelial–mesenchymal transition (EMT) and cancer stem cell (CSC) properties both in vitro and in vivo. This evidence concerns the gene EIF5A2 and head and neck squamous cell carcinoma.