Application of PAXgene Blood ccfDNA Tubes* as a standardized approach for therapy monitoring in late-stage lung cancer

Schmidt1,2, E. Arslan2, D.Reinicke1 , T. Voss3, A. Ullius3, S. Eisenmann1 , M. Fleischhacker2

1Universitätsklinikum Halle/Saale, Med. Klinik I, Schwerpunkt Pneumologie, 2DRK Kliniken Berlin, Klinik für Innere Medizin – Schwerpunkt Pneumologie, 3QIAGEN, Hilden, Germany



In June 2016, the FDA approved the first blood-based genetic test to detect gene mutations of epidermal growth factor receptor (EGFR) in non-small cell lung cancer (NSCLC). With this move, the genetic characterization of cell-free DNA became a routine application for the care of NSCLC patients. Nevertheless, there are many unresolved preanalytical issues.

Particularly important is defining the optimal method for blood draw and sample handling before plasma preparation. The aim of this research study is to answer two questions:

– Would the PAXgene Blood ccfDNA Tube* (PAXgene Tubes) be a viable alternative to EDTA blood tubes as the current gold standard?

– Could a longitudinal analysis of methylated cell-free plasma DNA allow predicting response to NSCLC treatment?

Experimental procedures

To date, 45 advanced-stage lung cancer patients were enrolled in this ongoing research study. Blood samples from 17 were used for the first study part to compare the quantity of plasma mPTGER4 and mSHOX2 DNA in both tubes types. At diagnosis, blood from these therapy-naive subjects was drawn into EDTA and PAXgene tubes. The EDTA tubes were processed immediately (within 4 h after blood draw) and spun twice to separate the plasma. The PAXgene tubes were stored for up to 7 days at room temperature (15–25°C) before processing to mimic transport between clinical and laboratory sites. All plasma samples were stored at -–80°C before DNA isolation and quantification of the two methylated sequences by real-time PCR. For the longitudinal study, blood was drawn from 28 subjects into PAXgene tubes only in approximately one week intervals, starting before therapy until conventional re-staging. The quantity of mPTGER4 and mSHOX2 plasma DNA was measured as in the comparative research study.

Summary of data

The amount of plasma mPTGER4 and mSHOX2 DNA in EDTA and PAXgene tubes was completely congruent for both markers in all 17 samples of the first study part.

The analysis of the two methylation markers in the second study part allowed predicting response/non-response to the given therapy in 25/28 subjects (88%), when compared to standard computer tomography-based re-staging. PCR results show that an assessment of therapy success could already be possible within two to three weeks after therapy start.


PAXgene tubes stabilize blood cells and thus prevent a dilution of cell-free DNA by genomic DNA from white blood cells. No change in the amount of tumor-derived cell-free DNA was seen. Therefore, PAXgene Blood ccfDNA Tubes* could be a good alternative to EDTA tubes. Therapy monitoring by longitudinal measurements of mPTGER4 and mSHOX2 plasma DNA could be several weeks faster than re-staging by the currently used digital imaging method.

*The PAXgene Blood ccfDNA Tube is for Research Use Only in the US.

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