Cell-Free DNA Source of Cell-Free DNA DNA in Urine Other Cancer Indicators Indicator Limitations

Indicator Limitations

Measuring the concentration of DNA in bodily fluids, whether plasma, serum, or urine, results in a very non-specific assay. As we have discussed, many different types of cancer, as well as other conditions such as pregnancy, systemic lupus erythematosus, rheumatoid arthritis, pulmonary embolism and myocardial infarction have been linked to elevated DNA levels. At best, then, this can only be a sign that something may be wrong, and further tests must be conducted to narrow down the possibilities.

Sozzi et al. (2005) reported the degradation of cell-free DNA over long term storage. With a median time of 41 months, they report that DNA levels declined substantially between the two assessments at an average rate of approximately 30% per year. Of greater concern, Herrera et al. (2005) found no difference between esophageal or lung cancer patients and controls when quantitatively measuring circulating DNA concentrations, and suggest that qualitative analysis may be necessary. Wu et al. (2002) reported that although elevated DNA was found in patients with a wide variety of cancer types, only 50% of cancer patients had elevated cell-free DNA. Thus, the potential for false negatives may be quite high.

Apart from the fact that not all cancer types have been correlated with elevated levels of circulating DNA, whether from lack of current research or from lack of positive correlation between elevated levels of circulating DNA, there are other non-cancer disease states that have been correlated to elevated levels of DNA. These disease states may generate false positives while using urinary DNA as a prognostic tool for cancer in the general population.

Authors (year)	Disease State	Results
Lo et al. (2000)	Acute blunt traumatic injury	Patients with adverse outcomes had significantly higherplasma DNA concentrations (11.6- to 12-fold) than those whodid not develop these complications.
Atamaniuk et al. (2004)	Exhaustive exercise	In the resting state, mean cell-free plasma DNA was [18.01pg/µL]. Immediatelyafter the race, plasma DNA was significantly increased [334.4 pg/µL]. Interestingly, by 2 h after the race, cell-freeplasma DNA had returned to baseline [30.44].
Rhodes et al. (2006)	ICU admission	Plasma DNA was higher in ICU patients than in healthy controls and was also higher in patients who developed sepsis. Plasma DNA had a sensitivity of 92% and a specificity of 80% when a concentration higher than 127 ng/ml was taken as a predictor for death on the ICU.
Wijeratne et al. (2004)	ICU admission	ICU patients' median plasma cell-free DNA concentration was 2.6-foldhigher than in healthy subjects. The median cell-free plasma DNA concentration in nonsurvivorswas 2.3-fold greater than that in survivors^.
Chang et al. (2003)	Myocardial infarction	Patients who had suffered MI had average cell-free DNA 10-fold higher than that of normal individuals. (510.6±398 vs. 36.3±23.8 ng/ml)
Leon et al. (1977)	Rheumatoid arthritis	In RA patients, a mean serum DNA concentration of 187 ng/mL was found, while healthy controls had a mean concentration of 13 ng/mL.
Rainer et al. (2003)	Stroke	Medianplasma DNA concentrations taken within 3 h of symptom onsetwere higher in stroke patients who died compared with those who survivedat discharge. Plasma DNA concentrations had a sensitivity of 100%and a specificity of 74.4% for predicting hospital mortalityafter stroke.
Tsai et al. (1994)	Systemic lupus erythematosus	Compared with the controls, plasma DNA was markedly increased in SLE (59.3% in SLE vs 7.4% in controls).
Fournie et al. (1993)	Various acute or chronic illnesses	Plasma DNA was higher in patients suffering from various acute or chronic illnesses than in patients without chronic disease. Plasma DNA was also higher than in a control group of healthy subjects. Increase in plasma DNA concentration was found in various pathological situations associated with cell death phenomena, including infections, cancers with metastasis, hepatitis, irreversible cardiac failure, severe respiratory insufficiency and thrombophlebitis.

Table 4. Other disease states that are positively correlated with elevated circulating DNA.

While the above table does list other indications that may cause an increase in Cell-free DNA levels in the body, none of the above indications would be unknown to the individual prior to the test. For example, prior to testing for Cell-free DNA in urine, the individual would be aware of any recent blunt trauma injury or suffering from an acute chronic illness.

Atamaniuk J, Vidotto C, Tschan H, Bachl N, Stuhlmeier KM, Muller MM. Increased concentrations of cell-free plasma DNA after exhaustive exercise. Clin chem 2004; 50: 1668-1670.

Chang CP, Chia RH, Wu TL, Tsao KC, Sun CF, Wu JT. Elevated cell-free serum DNA detected in patients with myocardial infarction. Clin Chim Acta 2003; 327: 95-101.

Fournie GJ, Martres F, Pourrat JP, Alary C, Rumeau M. Plasma DNA as cell death marker in elderly patients. Gerontology 1993; 39:215-221.

Herrera LJ, Raja S, Gooding WE, El-Hefnawy T, Kelly L, Luketich JD, et al. Quantitative analysis of circulating plasma DNA as a tumor marker in thoracic malignancies. Clin Chem 2005; 51: 113-118.

Leon SA, Ehrlich GE, Shapiro B, Labbate VA. Free DNA in the serum of rheumatoid arthritis patients. J Rheumatol 1977; 4: 139-143.

Leon SA, Shapiro B, Sklaroff DM, Yaros MJ. Free DNA in the serum of cancer patients and the effect of therapy. Cancer Res 1977; 37: 646-650.

Lo YM, Rainer TH, Chan LY, Hjelm NM, Cocks RA. Plasma DNA as a prognostic marker in trauma patients. Clin Chem 2000; 46: 319-323.

Rainer TH, Wong LK, Lam W, Yuen E, Lam NY, Metreweli C, et al. Prognostic use of circulating plasma nucleic acid concentrations in patients with acute stroke. Clin Chem 2003; 49: 562-569.

Rhodes A, Wort SJ, Thomas H, Collinson P, Bennett ED. Plasma DNA concentration as a predictor of mortality and sepsis in critically ill patients. Crit Care 2006; 10: R60.

Sozzi G, Roz L, Conte D, Mariani L, Andriani F, Verderio P, et al. Effects of prolonged storage of whole plasma or isolated plasma DNA on the results of circulating DNA quantification assays. J Natl Cancer Inst 2005; 97: 1848-1850.

Tsai CY, Yu CL, Huang DF, Hsieh SC, Tsai YY, Tsai ST. The elevation of plasma DNA in patients with systemic lupus erythematosus is attributable to increased DNA release and defective DNA binding of mononuclear cells. Zhonghua Yi Xue Za Zhi (Taipei) 1994; 54: 291-299.

Wijeratne S, Butt A, Burns S, Sherwood K, Boyd O, Swaninathan R. Cell-free plasma DNA as a prognostic marker in intensive treatment unit patients. Ann N Y Acad Sci 2004; 1022: 232-238.

Wu TL, Zhang D, Chia JH, Tsao KH, Sun CF, Wu JT. Cell-free DNA: measurement in various carcinomas and establishment of normal reference range. Clin Chim Acta 2002; 321: 77-87.

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