The influence of linker chain length on the sequence specificity of DNA damage by n-bromoalkylphenanthridinium bromides in plasmid DNA and in intact human cells |
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| Affiliation: | 1. School of Biochemistry and Molecular Genetics, University of New South Wales, Sydney, NSW 2052, Australia;2. School of Chemistry, University of Melbourne, Parkville, VIC 3052, Australia;3. Department of Chemistry, University of Wollongong, Wollongong, NSW 2522, Australia;1. School of Public Health, University of California, Berkeley, 94720 Berkeley, CA, USA;2. Institute for Global Health Sciences, University of California, San Francisco, 94158 San Francisco, CA, USA;3. Centro de Estudios en Salud, Universidad del Valle de Guatemala, Guatemala City, Guatemala;4. School of Nursing, University of California, San Francisco, 94158 San Francisco, CA, USA;5. Epidemiology and Biostatistics Department, University of Georgia, 30606 Athens, GA, USA;6. Climate, Air Quality Research Unit, School of Public Health and Preventive Medicine, Monash University, Melbourne, VIC, Australia;7. Nell Hodgson Woodruff School of Nursing, Emory University, 30322 Atlanta, GA, USA;8. Department of Medicine, University of California, San Francisco, San Francisco, CA, USA;1. Department of Epidemiology, University of Groningen, University Medical Center Groningen, 9713 GZ, Groningen, the Netherlands;2. Laboratory of Environmental Medicine and Developmental Toxicology, Shantou University Medical College, 515041, Guangdong, China;3. Department of Endocrinology, University of Groningen, University Medical Center Groningen, 9713 GZ, Groningen, the Netherlands;4. Department of Bioinformatics, Isfahan University of Medical Sciences, Isfahan 81746-7346, Iran;5. Department of Laboratory Medicine, University of Groningen, University Medical Center Groningen, 9713 GZ, Groningen, the Netherlands;6. Laboratory of Environmental Medicine and Developmental Toxicology, Guangdong Key Laboratory of Environmental Pollution and Health, School of Environment, Jinan University, 510632, Guangdong, China;7. Department of Genetics, University of Groningen, University Medical Center Groningen, 9713 GZ, Groningen, the Netherlands;1. Departments of Genetic Toxicology and Environmental Genomics, The Key Laboratory of Modern Toxicology of Ministry of Education, Center for Global Health, Jiangsu Key Laboratory of Cancer Biomarkers, Prevention and Treatment, Collaborative Innovation Center for Cancer Personalized Medicine, School of Public Health, Nanjing Medical University, Nanjing, China;2. The Affiliated Taizhou People''s Hospital of Nanjing Medical University, Taizhou, China;3. Key Laboratory of Environmental Medicine Engineering, Ministry of Education of China, School of Public Health, Southeast University, Nanjing, China;4. Department of Respiratory Medicine, Children''s Hospital of Nanjing Medical University, Nanjing, China;5. Department of Biostatistics, Center for Global Health, School of Public Health, Nanjing Medical University, Nanjing, China;6. The Affiliated Suzhou Hospital of Nanjing Medical University, Suzhou Municipal Hospital, Gusu School, Nanjing Medical University, Suzhou, China |
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| Abstract: | The sequence specificity of DNA damage of n-bromoalkylphenanthridinium bromides, with linker chain lengths (n) of 4,6,8 and 10 methylene groups, was investigated in the plasmid pUC8 and in intact human cells. A linear amplification assay was used to elucidate the DNA sequence specificity of the alkylating agents. In this assay Taq DNA polymerase extends from an oligonucleotide primer up to the damage site and the products run on a DNA sequencing gel to reveal the precise sites of DNA damage. For both the plasmid and cellular experiments, the compound that caused the most damage to DNA was the n = 6 compound, followed by (in decreasing order) the n = 4, n = 8, and n = 10 compounds. There were significant differences in the sequence specificity of DNA damage between n-bromoalkylphenanthridinium bromides of different linker chain length: (1) the main sites of damage were at guanines for the n = 4,6 and 8 compounds but at guanines and adenines for the n = 10 compound; (2) a consensus sequence of 5′-c(a/t)Ggg-3′ was obtained for the n = 4,6 and 8 compounds but 5′-c(a/c)(G/A)(g/a)-3′ for the n = 10 compound; (3) runs of consecutive Gs were the major site of damage for the n = 4,6 and 8 compounds, but consecutive Gs or consecutive As for the n = 10 compound; (4) for damage at single isolated guanines, the most damaged sequences were at 5′-Ga-3′ for the n = 4 compound but at 5′-Gt-3′ for the n = 6,8 and 10 compounds. The tandemly repeated alpha RI DNA sequence was the DNA target in intact human K562 cells. In intact human cells, the compounds produced damage with similar DNA sequence selectivity to that found in plasmid DNA. The n = 4 and 6 compounds possess marginal anti-tumour activity and these compounds produced the most damage in intact human cells. The n = 8 and 10 compounds do not demonstrate significant anti-tumour activity and these compounds resulted in the least damage in cells. |
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