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Quantitation of DNA adducts by stable isotope dilution mass spectrometry
Natalia Tretyakova1, Melissa Goggin, Dewakar Sangaraju
1Department of Medicinal Chemistry and Masonic Cancer Center, University of Minnesota , Minneapolis, Minnesota 55455, USA. trety001@umn.edu
Abstract:
Exposure to endogenous and exogenous chemicals can lead to the formation of structurally modified DNA bases (DNA adducts). If not repaired, these nucleobase lesions can cause polymerase errors during DNA replication, leading to heritable mutations and potentially contributing to the development of cancer. Because of their critical role in cancer initiation, DNA adducts represent mechanism-based biomarkers of carcinogen exposure, and their quantitation is particularly useful for cancer risk assessment. DNA adducts are also valuable in mechanistic studies linking tumorigenic effects of environmental and industrial carcinogens to specific electrophilic species generated from their metabolism. While multiple experimental methodologies have been developed for DNA adduct analysis in biological samples, including immunoassay, HPLC, and ³²P-postlabeling, isotope dilution high performance liquid chromatography-electrospray ionization-tandem mass spectrometry (HPLC-ESI-MS/MS) generally has superior selectivity, sensitivity, accuracy, and reproducibility. As typical DNA adduct concentrations in biological samples are between 0.01-10 adducts per 10⁸ normal nucleotides, ultrasensitive HPLC-ESI-MS/MS methodologies are required for their analysis. Recent developments in analytical separations and biological mass spectrometry, especially nanoflow HPLC, nanospray ionization MS, chip-MS, and high resolution MS, have pushed the limits of analytical HPLC-ESI-MS/MS methodologies for DNA adducts, allowing researchers to accurately measure their concentrations in biological samples from patients treated with DNA alkylating drugs and in populations exposed to carcinogens from urban air, drinking water, cooked food, alcohol, and cigarette smoke.
Insights
DNA adducts, or modified DNA bases, are key biomarkers for carcinogen exposure and cancer risk. Ultrasensitive methods like isotope dilution HPLC-ESI-MS/MS accurately quantify these adducts, aiding mechanistic studies and risk assessment.
Area of Science:
- Environmental Health Sciences
- Molecular Biology
- Analytical Chemistry
Background:
- Chemical exposure can create DNA adducts, which if unrepaired, lead to mutations and cancer.
- DNA adducts serve as crucial biomarkers for carcinogen exposure and are vital for cancer risk assessment.
- Understanding DNA adducts helps link environmental carcinogens to their metabolic electrophilic species.
Purpose of the Study:
- To highlight the importance of DNA adducts in cancer initiation and risk assessment.
- To discuss the role of DNA adducts in mechanistic studies of carcinogenicity.
- To review advanced analytical methodologies for DNA adduct quantitation.
Main Methods:
- Isotope dilution high-performance liquid chromatography-electrospray ionization-tandem mass spectrometry (HPLC-ESI-MS/MS) offers superior selectivity, sensitivity, accuracy, and reproducibility for DNA adduct analysis.
- Ultrasensitive methodologies are required due to low adduct concentrations (0.01–10 adducts per 10⁸ nucleotides).
- Recent advancements include nanoflow HPLC, nanospray ionization MS, chip-MS, and high-resolution MS, enhancing analytical capabilities.
Main Results:
- HPLC-ESI-MS/MS enables precise measurement of DNA adducts.
- Advanced techniques have significantly improved the sensitivity and accuracy of DNA adduct analysis.
- Accurate quantitation allows for assessment of exposure in various populations and patient groups.
Conclusions:
- Accurate quantitation of DNA adducts using advanced HPLC-ESI-MS/MS is essential for cancer risk assessment and mechanistic studies.
- These methods allow for the measurement of DNA adducts in populations exposed to diverse carcinogens.
- The continuous development of analytical techniques further refines the ability to detect and quantify DNA adducts in biological samples.
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