A comprehensive investigation of D-Loop mutations in the mitochondrial DNA as potential diagnostic molecular markers for colorectal cancer.

SR Fradley, BSc Genetics

University of Wales Swansea, Swansea, West Glamorgan United Kingdom

AIM The human mitochondrial genome (mtDNA) contains a short hypervariable, non-coding control region known as the D-Loop. Recent investigations have revealed that a proportion of colorectal adenocarcinomas harbour mtDNA mutations, not present in normal surrounding mucosa, classified as tumour-specific. Tumour specific mtDNA mutations have also been observed in other tumours including lung and bladder. It has been suggested that these mutations may serve as diagnostic markers for cancer as the D-Loop may be rapidly and cost-effectively scanned for mutations. Previous studies searching for tumour-specific mtDNA mutations in colorectal cancer have relied on small sample sizes or have failed to reveal D-Loop mutations. METHODS This comprehensive survey of the D-Loop in adenocarcinoma and normal mucosa of twenty patients uses PCR-SSCP and DNA sequencing to (i) demonstrate the usefulness of the D-Loop in providing tumour-specific markers for colorectal cancer, (ii) reveal the types and distribution of mutations within the D-Loop, (iii) estimate mutation frequency within this region in adenocarcinomas. RESULTS Of the colorectal adenocarcinomas, 25% showed tumour specific mutations. Sequencing revealed the mutations as 1-basepair C:G deletion and 1-basepair C:G insertion at nucleotide position 309, two C:G/T:A transitions at positions 61 and 52 and one T:A/A:T transversion at nucleotide 55. This study is currently ongoing, we aim to increase the sample size and analyse adenomas and hyperplastic polyps. CONCLUSIONS Our results will allow predictions to be made concerning the causative factors of mtDNA mutations in colorectal cancer and give insight into whether these mutations precede neoplasia or are a consequence of tumorigenesis.

KEY WORDS: tumour-specific markers, colorectal cancer, mitochondrial DNA, .

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Paper presented at the International Symposium on Predictive Oncology and Intervention Strategies; Paris, France; February 9 - 12, 2002; in the section on Prognostic Markers.