BACKGROUND: Somatic mutations of mitochondrial DNA (mtDNA) are increasingly 
being recognized in many human cancers, but automated sequencing of 16.5 kb of 
DNA poses an onerous task. We have recently described an oligonucleotide 
microarray (MitoChip) for rapid and accurate sequencing of the entire 
mitochondrial genome (Zhou et al., J Mol Diagnostics, 8: 9_14, 2006), greatly 
facilitating the analysis of mtDNA mutations in cancer. In this report, we 
perform a comprehensive cataloging of somatic mutations in the mitochondrial 
genome of human pancreatic cancers using our novel array-based approach.
MATERIALS AND METHODS: MitoChip analysis was performed on DNA isolated from 15 
histologically confirmed resection specimens of pancreatic ductal 
adenocarcinomas. In all cases, matched nonneoplastic pancreatic tissue was 
obtained as germline control for mtDNA sequence. DNA was extracted from 
snap-frozen cryostat-embedded specimens and hybridized to the sequencing 
microarray after appropriate polymerase chain reaction amplification and 
labeling steps. The vast majority of somatic mutational analyses of mtDNA in 
human cancers utilize lymphocyte DNA as germline control, without excluding the 
potential for organ-specific polymorphisms. Therefore, we also examined a series 
of 15 paired samples of DNA obtained from nonneoplastic pancreata and 
corresponding EBV-immortalized lymphoblastoid cell lines to determine whether 
lymphocyte DNA provides an accurate surrogate for the mtDNA sequence of 
pancreatic tissue.
RESULTS: We sequenced 497,070 base pairs of mtDNA in the 15 matched samples of 
pancreatic cancer and nonneoplastic pancreatic tissue, and 467,269 base pairs 
(94.0%) were assigned by the automated genotyping software. All 15 pancreatic 
cancers demonstrated at least one somatic mtDNA mutation compared to the control 
germline DNA with a range of 1-14 alterations. Of the 71 somatic mutations 
observed in our series, 18 were nonsynonymous coding region alterations (i.e., 
resulting in an amino acid change), 22 were synonymous coding region 
alterations, and 31 involved noncoding mtDNA segments (including ribosomal and 
transfer RNAs). Overall, somatic mutations in the coding region most commonly 
involved the ND4, COI, and CYTB genes; of note, an A-G transition at nucleotide 
position 841 in the 12sRNA was observed in three independent samples. In the 
paired analysis of nonneoplastic pancreata and lymphoblastoid cell line DNA, 14 
nucleotide discrepancies were observed out of 226,876 nucleotide sequences (a 
concordance rate of 99.99%), with 9 samples demonstrating a perfect match across 
all bases assigned.
CONCLUSIONS: Our findings confirm that somatic mtDNA mutations are common in 
pancreatic cancers, and therefore, have the potential to be a clinically useful 
biomarker for early detection. Further, our studies confirm that lymphocyte DNA 
is an excellent, albeit not perfect, surrogate for nonneoplastic pancreatic 
tissues in terms of being utilized as a germline control. Finally, our report 
confirms the utility of a high-throughput array-based platform for mtDNA 
mutational analyses of human cancers.