Changes of Total and Tumor Specific Cell Free Dna During an Exhaustive Cycling Exercise Test

Abstract

PURPOSE: Circulating, cell free DNA (cfDNA) is known to directly accumulate in a load-dependent fashion in various aerobic and anaerobic exercise sessions within minutes. Furthermore, increases of cfDNA at baseline have been reported for different patient groups with cancer to depend on malignancy and tumor progression. Typically, such analysis did not differentiate between cfDNA and circulating tumor derived levels of DNA (ctDNA). Here we investigated for the first time, whether exercise to voluntary exhaustion induces increases in cfDNA and ctDNA in patients with advanced stage cancer. METHODS: In a pilot study, individuals with solid tumors of diverse entities and an age matched control group (n = 5) were subjected to a step-wise incremental cycling spiroergometry until exhaustion. Blood samples were taken before, directly after, and 90 minutes after the end of the test. Total cfDNA was quantified directly from the blood plasma using a qPCR targeting repetitive Line-1 elements and ctDNA was measured following silica-based total DNA purification and enrichment from plasma and subsequent specific quantification using a nested qPCR assay targeting the seven most frequent oncogenic KRAS sequences in comparison to wild-type sequence. Additionally, DNase activity reduction was measured with a commercial ELISA kit. RESULTS: We quantified total cfDNA in all blood samples and detected and quantified ctDNA concentrations in all three samples of one cancer patient with metastatic colon cancer. Total cfDNA concentrations increased 1.7-fold (95% CI 1.28-2.22-fold; p<0.01) in all tumor patients during exercise, but to a significantly lesser extent (p=0.003) than in the healthy control group (3.7-fold; CI 2.23-6.15-fold, p<0.01). The decrease of cfDNA concentrations during the recovery period was significantly lower in tumor patients than in the healthy control (1.8-fold vs 5.5-fold decrease, p=0.001), independent of the DNase activity. Tumor DNA increased only slightly, causing a relative decrease in the ctDNA during exercise. CONCLUSIONS: cfDNA kinetics seem to be less dynamic in tumor patients, especially the depletion of cfDNA from the plasma seems to be impaired. This effect does not seem to be a result of circulating DNA originating from the Tumor.