The carcinogenetic role of human papillomavirus (HPV) in mobile tongue cancer remains unclear because of conflicting results reported in the literature. This disparity is likely to be due to variations in the samples and methods used. Furthermore, despite a tendency for increased prevalence of mobile tongue cancer in young adults, only a few reports specifically in young patients have been published. In the present study on 32 patients, including six in their 20s, we genotyped the prevalence of HPV using a highly sensitive detection tool in fresh-frozen samples from surgical specimens and a novel detection device with electrochemical DNA chip and loop-mediated isothermal amplification. In addition, we confirmed HPV prevalence by in situ hybridization and immunohistochemistry for the p16INK4a protein, regarded as a biomarker of HPV-associated cancers. The frequency of 13 genotypes of highrisk HPV was 0 ⁄ 32 (0%), which was further confirmed by in situ hybridization. Overexpression of p16INK4a protein was observed in six of the 32 patients (19%), with four (67%) also overexpressing p53. Because there is usually a lack of p53 overexpression in HPVassociated cancer, it is unlikely that p16INK4a protein overexpression is correlated with HPV infection. Consequently, it is unlikely that HPV infection plays an important role in mobile tongue carcinogenesis, in particular in young adults. In addition, our data suggest that the overexpression of p16INK4a protein is not an appropriate biomarker for HPV association in mobile tongue carcinogenesis. (Cancer Sci 2012; 103: 161-168) H uman papillomavirus (HPV) is a small DNA virus and more than 130 different types of HPV have been identified based on DNA sequence variations.(1,2) Human papillomavirus is divided into low-risk and high-risk types, depending on the carcinogenic power.(3) It has been conclusively established that more than 99% of uterine cervical cancers are associated with HPV infection.(2,4) Because HPV-associated cancers may be prevented with vaccination, (5)(6)(7) it is important to determine whether cancer of extracervical organs is associated with HPV infection. (8,9) Among the head and neck cancers, oropharyngeal cancer, particularly tonsillar cancer, and basal tongue cancer are known to be strongly associated with HPV infection.(3) However, the role of HPV in mobile tongue cancer (MTC) remains unclear because of the disparity of results published so far, which have reported HPV infection rates ranging from 0% to 100%. (10)(11)(12)(13)(14)(15)(16)(17)(18)(19)(20) This disparity may be due to differences in the types of samples used and the methods of detection. A recent study has reported an increasing incidence of MTC among young adults.(21) Thus far, alcohol consumption and heavy cigarette smoking have been suggested as causal factors for MTC. (11,13) However, it is difficult to explain the high incidence of MTC in young adults simply as a result of accumulated exposure to alcohol and smoking. (11,21,22) To date, there have been only a few reports on re...