Aberrant expression of miR-10b has been described in many cancers but remains unexplored in nasopharyngeal carcinoma (NPC). Therefore, we aimed to study the miR-10b expression level in 43 NPC biopsies collected from Tunisian patients and three NPC xenografts. Then, we investigated the correlation between miR-10b expression and its upstream regulators LMP1/Twist1 as well as its adjacent gene HoxD4. We showed that miR-10b was significantly up-regulated in NPC biopsies compared to non-tumor nasopharyngeal tissues (fold change 153; p = 0.004) and associated with advanced clinical stage and young age at diagnosis (p = 0.005 and p = 0.011, respectively). In addition, over-expression of miR-10b was positively associated with the transcription factor Twist1 as well as the EBV oncoprotein LMP1 (fold change 6.32; p = 0.014, fold change 6.58; p = 0.01 respectively). Furthermore, higher level of miR-10b was observed in tumors with simultaneous expression of LMP1 and Twist1, compared to those expressing only Twist1 (fold change 2.49; p = 0.033). Meanwhile, the analysis of the link between miR-10b and its neighbor gene HoxD4 did not show any significant correlation (Fisher test p = 0.205; Mann-Whitney test p = 0.676). This study reports the first evidence of miR-10b over-expression in NPC patients. Furthermore, our findings can support hsa-miR-10b gene regulation through LMP1/Twist1 in NPC malignancy.

To further explore the epigenetic changes in nasopharyngeal carcinoma (NPC), methylation-sensitive arbitrarily primed PCR was performed on NPC biopsies and nontumor nasopharyngeal samples. We have shown mainly two DNA fragments that appeared to be differentially methylated in NPCs versus nontumors. The first, defined as hypermethylated, corresponds to a CpG island at the 5′-end of the tetratricopeptide repeat domain 40 (TTC40) gene, whereas the second, defined as hypo-methylated, is located on repetitive sequences at chromosomes 16p11.1 and 13.1. Thereafter, the epigenetic alteration on the 5′-TTC40 gene was confirmed by methylation-specific PCR, showing a significant aberrant methylation in NPCs, compared to nontumors. In addition, the bisulfite sequencing analysis has shown a very high density of methylated cytosines in C15, C17, and X666 NPC xenografts. To assess whether TTC40 gene is silenced by aberrant methylation, we examined the gene expression by reverse transcription-PCR. Our analysis showed that the mRNA expression was significantly lower in tumors than in nontumors, which is associated with 5′-TTC40 gene hypermethylation. In conclusion, we found that the 5′-TTC40 gene is frequently methylated and is associated with the loss of mRNA expression in NPCs. Hypermethylation of 5′-TTC40 gene might play a role in NPC development; nevertheless, other studies are needed.