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Assessment of Oxidative Damage to DNA - Cancer, Metals, and Cardiovascular Disease

Posted By David Quig, Phd - VP Scientific Support, Doctor's Data, Inc., Thursday, November 17, 2016
Reactive oxygen species (ROS), both radical and non-radical, are formed continuously in all cells and they are indeed part of physiological and metabolic processes. Under normal physiological conditions there is a balance between the generation of ROS and the body’s ability to appropriately quench them.  When the capacity to quench is exceeded, varying degrees of oxidative stress can ensue.  When sustained major oxidative stress occurs, radical oxygen species such as the hydroxyl radical can cause oxidative damage to important macromolecules such as DNA.  The hydroxyl radical (.OH) readily oxidizes guanosine nucleobases and most of the resultant 8-OH-2’-deoxyguanosine (8-OH-dG) is excised and excreted in urine. 8-OH-dG is the most heavily studied biomarker of excessive intracellular oxidative stress (over 6,200 articles). The level of 8-OH-dG in urine is an important clinical biomarker of oxidative damage to DNA, degenerative diseases, accelerated aging processes, excessive exposure to metals, and cancer. 

We are all subjected to at least low level exposure to environmental and food derived toxicants, and the Centers for Disease Control and Prevention have stated that the epidemic of epidemics of CVD, neurological diseases, and immunological diseases is likely associated with environmental toxicants. Based upon published biomedical literature it is likely that we can add cancers to that list as well.  A primary common effect of environment toxicants, and their partially metabolized toxins, is that they induce oxidative stress in the form of electrophiles and ROS. ROS are formed continuously in all cells and are absolutely essential for life as they participate in cell signaling pathways, and both radical and non-radical ROS are deployed by the immune system to kill invading microorganisms (systemically and in the gastrointestinal tract).  Physiological levels of non-radical oxygen species such as hydrogen peroxide can be readily neutralized by glutathione peroxidase and catalase enzymes.  Likewise even the radical superoxide anion can be neutralized by innate superoxide dismutases. However, when hydrogen peroxide and/or the superoxide radical are produced in excess of the body’s capacity to neutralize the ROS, the .OH is generated from hydrogen peroxide in the Fenton (Fe+) and Haber-Weis (superoxide anion) reactions, respectively. Unlike hydrogen peroxide and the superoxide radical, the extremely reactive .OH has a half-life of only 10-9 seconds and cannot be eliminated by an enzymatic reaction. There aren’t clinically applicable ways to directly measure the .OH, but the damage that it causes to proteins, lipids and DNA can be assessed in various biological matrices.

Oxidative damage to DNA can result in base and sugar modifications, covalent crosslinks, and single- and double-stranded breaks. However with respect to cancer, nucleobase oxidation has been subject of the most research. The C8 position of the 2-deoxyguanosine nucleoside base is extremely vulnerable to oxidation by the .OH resulting in 8-OH-dG. The 8-OH-dG is a radical in and of it-self and is involved in the initiation and promotion of carcinogenesis when not quantitatively removed by DNA repair mechanisms (e.g. the OGG1 protein). 8-OH-dG has a major role in spontaneous mutagenesis. It induces C- to- T transversions which are among the most frequent somatic mutations found in human cancers. Elevated levels of 8-OH-dG in urine have been associated with prostate, bladder, and lung cancer. In recent years numerous studies have identified the importance of urinary levels of 8-OH-dG in human lung cancer, and this association has been linked with exposure to tobacco smoke, diesel exhaust particles (metals and polycyclic hydrocarbons), oil fly ash (vanadium, manganese, nickel and lead), and urban air pollution.  Other research has focused on the effects of occupational exposures to known carcinogens such as benzene, styrene, and inorganic arsenic and urine 8-OH-dG. Therein a dose response relationship has consistently been found.

Exposures to inorganic arsenic, chromium, and mercury have also been found to be associated with elevated levels of urine 8-OH-dG.  Evaluation of the urine levels of arsenic, chromium, and 8-OH-dG for children (n=142, 10-12 years old) in multiple schools in China were determined. One school was adjacent to and downwind of 8 coal-fired power plants (effluent smoke and dust) and the other two schools were upwind in a more suburban area. Urine 8-OH-dG levels were correlated with urine chromium and arsenic levels, and the highest levels of 8-OH-dG were exhibited in the children with high levels of both chromium and arsenic.  The data were analyzed by co-variate analysis of variance and smoking in the home was not a significant co-variate.  In a more recent study, low level exposure to inorganic arsenic in utero and during early childhood was associated with higher levels of urine 8-OH-dG than for non-exposed children.  As methylation reactions are essential for innate detoxification and elimination of inorganic arsenic it is not surprising that those with higher levels of urine arsenic and 8-OH-dG had lower methylation indices (e.g. low plasma SAM : SAH ratios).  Consistent with the mechanism for the appearance of 8-OH-dG in urine, arsenic exposed children that had lower expression of the DNA base excision repair protein (OGG1) also had lower levels of 8-OH-dG in urine.

Exposure to inorganic mercury with compromised serum redox status has also been associated with elevated urine levels of 8-OH-dG. First morning urine levels of mercury and 8-OH-dG, fasted serum glutathione and total thiols were measured in mercury-exposed (active occupational, n=35) and non-exposed control adults in China.  Serum mercury, urine mercury and urine 8-OH-dG levels were markedly higher for the mercury-exposed group, and the biomarkers of redox status were significantly lower for the mercury-exposed group. It should be noted the serum and urine mercury levels in the exposed subjects were about 40-times greater than the non-exposed subjects.

Urine 8-OH-dG also appears to be an emerging risk factor for cardiac events. Urine 8-OH-dG is derived from both cellular and mitochondrial DNA.  In a 1.8 year prospective study of 186 CVD patients the odds ratio for cardiac events was 4-times higher for patients with elevated urine levels of 8-OH-dG. Elevated levels of 8-OH-dG have also been detected in atherosclerotic plaque from humans. Further, in a very recently published meta-analysis (14 studies) it was found that CVD patients had significantly higher urine and serum levels of 8-OH-dG than control subjects. Other conditions associated with elevated levels of urinary 8-OH-dG include mitochondrial dysfunction, inflammatory conditions (NF-ҡB mediated), and diabetic nephropathy and retinopathy (correlated with HbA1c).

Urine levels of 8-OH-dG have in fact been shown to represent levels of oxidative damage to DNA in cells.  The levels in urine and extracted lymphocytes from subjects were highly correlated when 8-OH-dG was measured by three independent methodologies.  Further, a study of human volunteers that were fed highly oxidized N15-labelled DNA clearly demonstrated that diet does not directly contribute to urinary levels of 8-OH-dG, and studies have indicated that normal cell death does not contribute significant levels of 8-OH-dG.  We and others have validated that levels of 8-OH-dG are highly correlated in 24 hour and first AM urine collections, and levels in urine are inherently higher in young children compared to adolescents/adults; age-appropriate reference ranges should be applied.  Urine 8-OH-dG levels can be conveniently assessed in a first morning urine specimen and provide an excellent, non-invasive indication of excessive intracellular oxidative stress and direct oxidative damage to cellular and mitochondrial DNA.

Select References
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 Accessed 5 September 2016

Valavanidis A et al. 8-hydroxy-2’-deoxyguanosine (8-OHdG): A critical biomarker of oxidative stress and carcinogenesis. J Environ Sci Hlth (2009) 27 Part C:120- 39.  http://www.tandfonline.com/doi/citedby/10.1080/1059050090288568
  Accessed 5 September 2016

Wong R-H et al.  Increased levels of 8-hydroxy-2’-deoxyguanosine attributable to carcinogenic metal exposure among schoolchildren. Env Hlth Perspect (2005)113(10):1386-90.  https://www.ncbi.nlm.nih.gov/pmc/articles/PMC1281285/
 Accessed 5 September

Hinhumpatch P et al. Oxidative DNA damage and repair in children exposed to low levels of arsenic in utero and during early childhood: application of salivary and urinary biomarkers. Toxicol Appl Pharmacol (2013)273(3):569-79.
Doi:10.1016/j.taap.213.10.003  http://www.sciencedirect.com/science/article/pii/S0041008X1300433X
  Accessed 30 October 2016
Chen C et al.  Increased oxidative DNA damage, as assessed by urinary 8-hydroxy-2’-deoxyguanosine concentrations, and serum redox status in persons exposed to mercury. Clin Chem (2005)51(4):759-67.  http://clinchem.aaccjnls.org/content/51/4/759.long
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Roseelo-Lleti E et al.  Impact of cardiovascular risk-factors and inflammatory status on urinary 8-OHdG in essential hypertension. Am J Hypertens (2012)25:236-42.  http://ajh.oxfordjournals.org/content/25/2/236.abstract
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Di Minno A et al. 8-Hydroxy-2-Deoxyguanosine Levels and CardiovascularDisease: A Systematic Review and Meta-Analysis of the Literature. Antioxidants & Redox Signaling (2016)24(10).
DOI: 10.1089/ars.2015.6508  https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4827317/
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Feruson LR et al.  Oxidative DNA damage and repair: Significance and biomarkers. J Nutr (2006)136:2687S-89S. http://jn.nutrition.org/content/136/10/2687S.long
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Jomova K et al.  Metals, oxidative stress and neurodegenerative disorders. Mol Cell Biochem (2010)345:91-104.  http://link.springer.com/article/10.1007/s11010-010-0563-x
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Pilger A and Rudiger HW.  8-hydroxy-2’-deoxyguanosine as a marker of oxidative DNA damage related to occupational and environmental exposures. Int Arch Occup Environ Hlth (2006)80(1):1-15.  http://link.springer.com/article/10.1007%2Fs00420-006-0106-7
 Accessed 5 September 2016

Tags:  cancer  cardiovascular disease  DNA  heavy metals  oxidative medicine 

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