Diagnostic value of circulating cell-free mtDNA in patients with suspected thyroid cancer: ND4/ND1 ratio as a new potential plasma marker

Thyroid cancer is the most common endocrine malignancy, and its incidence continues to rise. For clinicians with cancer patients, choosing and interpreting diagnostic laboratory studies has become increasingly important. Previously, changes in plasma free mitochondrial DNA levels have been found in colorectal, breast, lung, and urinary cancers, and have demonstrated diagnostic value. In this study, we investigated whether the occurrence and development of thyroid cancer might be predicted using mtDNA copy number (ND1), mtDNA integrity (ND4/ND1) and levels of cell-free nDNA (GAPDH). We analyzed ND1, ND4, and GAPDH levels in plasma and blood cells from 75 patients with thyroid cancer, 40 patients with nodular goiter, and 107 normal controls using real-time PCR. Although both the thyroid nodule and thyroid cancer patients had significantly increased ND1 levels, the ND4/ND1 ratio in the thyroid cancer group was higher than the thyroid nodule group (P < 0.05), and significantly higher than the normal control group (P < 0.01). Plasma levels of nuclear DNA (GAPDH) in the thyroid cancer group were also higher compared to normal (P < 0.05). These results indicate that increased intactness of plasma free mtDNA is associated with increased levels of plasma cell-free nDNA, and that the ND4/ ND1 ratio has the potential to be a new detection indicator in thyroid cancer. Furthermore, we classified thyroid cancer patients according to clinical data including age, tumor size, and metastasis. We found significantly higher levels of GAPDH in malignant tissues. Because ND4/ND1 correlated with plasma GAPDH in the plasma studies, this also suggests a potential relationship between ND4 intactness and thyroid tumor tissue size. Taken together, our findings suggest a tumor-specific process involving increased release of intact mtDNA, detectable in the plasma, which differentiates normal patients from patients with thyroid cancer.