Quantitation of cell-free DNA and RNA in plasma during tumor progression in rats.

BACKGROUND: To clarify the implications of cell-free nucleic acids (cfNA) in the plasma in neoplastic disease, it is necessary to determine the kinetics of their release into the circulation. METHODS: To quantify non-tumor and tumor DNA and RNA in the plasma of tumor-bearing rats and to correlate such levels with tumor progression, we injected DHD/K12-PROb colon cancer cells subcutaneously into syngenic BD-IX rats. Rats were sacrificed and their plasma was analyzed from the first to the eleventh week after inoculation. RESULTS: The release of large amounts of non-tumor DNA into plasma was related to tumor development from its early stages. Tumor-specific DNA was detected in 33% of tumor-bearing rats, starting from the first week after inoculation and at an increasing frequency thereafter. Animals that were positive for tumor DNA in the plasma had larger tumors than those that were negative (p = 0.0006). However, the appearance of both mutated and non-mutated DNA fluctuated with time and levels of both were scattered among individuals in each group. The release of non-tumor mRNA was unaffected by tumor progression and we did not detect mutated RNA sequences in any animals. CONCLUSIONS: The release of normal and tumor cfDNA into plasma appeared to be related to individual-specific factors. The contribution of tumor DNA to the elevated levels of plasma DNA was intermittent. The release of RNA into plasma during cancer progression appeared to be an even more selective and elusive phenomenon than that of DNA.

Release of cell-free DNA into the bloodstream leads to high levels of non-tumor plasma DNA during tumor progression in rats.

To examine the implications of cell-free DNA in the plasma in neoplastic disease, it is necessary to clarify various features of this DNA, such as the contribution of DNA from the host's normal cells and the kinetics of the release of this latter DNA. To quantify non-tumor DNA in the plasma of tumor-bearing rats and to correlate levels of this DNA with tumor progression, we injected DHD/K12-PROb colon cancer cells subcutaneously into BD-IX rats and recorded tumor diameters weekly. After euthanasia, we collected plasma from each rat and quantified non-mutated and mutated DNA in the plasma. Overall, levels of non-mutated (non-tumor) DNA in plasma of tumor-bearing animals were significantly higher than those in healthy animals measured by real-time PCR (p=0.001). However, 5 weeks after inoculation, levels were similar to those in healthy animals. As a whole, levels of non-mutated DNA were not significantly related to tumor size or to metastasis. However, when we excluded animals that were analyzed earliest, we found a positive and statistically significant correlation between levels of non-mutated DNA and tumor diameter (p=0.002). Release of cell-free DNA into the plasma during tumor progression appears to follow a predictable time course in a homogeneous population. In addition, large amounts of non-tumor DNA are released during tumor progression and, in particular, at early stages. Our findings support the hypothesis that interactions between tumor cells and host cells result in release of cell-free DNA.