To identify genes that could potentially serve as molecular therapeutic markers for human head and neck cancer (HNC), we employed differential display analysis to compare the gene expression profiles between HNC and histopathologically normal epithelial tissues. Using reverse transcription-polymerase chain reaction and Western blot analysis, desmoglein 3 (DSG3) was identified as being differentially expressed at both the RNA and protein levels. Of 56 patients assayed, 34 (61%) had overexpression of DSG3, which correlated statistically with T stage (P=0.009), N stage (P=0.047), overall stage (P=0.011), tumor depth (P=0.009) and extracapsular spread in lymph nodes (P=0.044), suggesting that DSG3 participates in carcinogenesis of HNC. Consistent with the clinical findings, inhibition of DSG3 by RNA interference (RNAi) significantly reduced cell growth and colony formation to 57-21% in three HNC cell lines. Use of an in vitro wound healing and Matrigel invasion assays, we found that cell migration and invasive ability were also inhibited to 30-48% in three cell lines tested. An in vivo xenograft study showed that administration of DSG3-RNAi plasmid significantly inhibited tumor growth for 2 months in BALB/C nude mice. In conclusion, DSG3 is identified overexpressed in HNC, with the degree of overexpression associated with clinicopathologic features of the tumor. Inhibition of DSG3 significantly suppresses carcinogenic potential in cellular and in vivo animal studies. These findings suggest that DSG3 is a potential molecular target in the development of adjuvant therapy for HNC.