Novel heteroplasmic frameshift and missense somatic mitochondrial DNA mutations in oral cancer of betel quid chewers

Abstract

Mitochondrial DNA (mtDNA) has been proposed to be involved in carcinogenesis because of its high susceptibility to oxidative DNA damage and limited repair mechanisms. For investigation of the potential role of somatic mtDNA mutations in the tumorigenesis of oral cancer, we screened the occurrence of mtDNA mutations by the temporal temperature gradient gel electrophoresis method. We amplified the entire mitochondrial genome by use of 32 pairs of overlapping primers, and to identify the mutations, we sequenced DNA fragments showing different banding patterns between normal and tumor mtDNA. Fourteen of eighteen (77.8%) oral carcinomas displayed somatic mtDNA mutations, with a total of 26 mutations. Among them, six were in the mRNA coding region. Three were missense mutations (C14F, H186R, T173P) in NADH dehydrogenase subunit 2, and one was a frameshift mutation, 9485delC, in cytochrome c oxidase subunit III. Eight (44%) tumors had insertion or deletion mutations in the nucleotide position 303-309 poly C region of the D-loop. Multiple large deletions were also observed. Our results demonstrate that somatic mtDNA mutations occur in oral cancer. Some missense and frameshift mutations may play an important role in the tumorigenesis of this carcinoma. More extensive biochemical and molecular studies will be necessary for determining the pathologic effect of these somatic mutations.

Clinicopathological significance of mitochondrial D-Loop mutations in head and neck carcinoma

“…They found all mtDNA mutations in a part of the D-Loop and a mtDNA microsatellite instability, defined as insertions or deletions in the D310 repeat, in 42% of the tumours. Another study showed 66% of D-Loop mutations in 18 oral cancers of betel quid chewers (Tan et al, 2003), which however cannot be really assimilated to HNSCC of tobacco and alcohol consumers. The vast majority of these mutations were located between nucleotides 204 and 489 and 44% of the tumours harboured mutations in the D310 repeat.…”

“…Betel quid contains tender areca nuts and lime that have been shown to generate ROS and induce oxidative DNA damage (Nair et al, 1987;Stich and Anders, 1989), which can initiate or promote oral carcinogenesis. These betel quid compounds could therefore preferentially target mtDNA, which may explain the high proportion of oral tumours with mtDNA mutation observed in betel quid chewers (Tan et al, 2003), compared to that observed in cigarette smokers.…”

“…Its role in apoptosis supports this hypothesis (Zamzami and Kroemer, 2001) and it was shown that mtDNA mutations may lead to a dysregulation of oxidative phosphorylation that can enhance production of the carcinogenic ROS. Over last years, somatic mtDNA mutations have been reported in many human tumours (Polyak et al, 1998;Maximo et al, 2000;Richard et al, 2000;Yeh et al, 2000;Hibi et al, 2001b;Sanchez-Cespedes et al, 2001;Kirches et al, 2001;Liu et al, 2001;Lievre et al, 2005), including head and neck squamous cell carcinoma (HNSCC) that were found mutated in small series in 37 -77% of the cases (Fliss et al, 2000;Sanchez-Cespedes et al, 2001;Ha et al, 2002;Tan et al, 2003;Poetsch et al, 2004). Although mutations may occur throughout the mitochondrial genome, the vast majority of them have been described in the noncoding region of the D-Loop and particularly in a mononucleotide repeat named D310 (C-tract, nucleotide position: 303 -315) that has emerged as a mutational hotspot in HNSCC (Fliss et al, 2000;Sanchez-Cespedes et al, 2001;Ha et al, 2002;Tan et al, 2003;Poetsch et al, 2004).…”

Comprehensive One-Step Molecular Analyses of Mitochondrial Genome by Massively Parallel Sequencing

“…A female carrying a pathogenic mtDNA mutation is at risk of passing the mutation to her offspring, even if she is asymptomatic with low-level heteroplasmy in somatic cells. In addition, there is growing evidence that somatic mtDNA alterations are associated with aging and a variety of common diseases, including diabetes and cancers (5)(6)(7)(8)(9)(10).…”

“…Somatic mtDNA alterations have been reported in many types of cancers and may play a role in tumorigenesis (5,(32)(33)(34)(35). However, relatively few cancers harbor detectable deleterious somatic mtDNA mutations (6,36,37 ). This hypothesis may be tested by using the deep sequencing approach which, in theory, reaches a level of detection equivalent to singlemolecule analysis.…”