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Circadian Rhythm of Methylated Septin 9, Cell-Free DNA Amount and Tumor Markers in Colorectal Cancer Patients

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Pathology & Oncology Research

Abstract

To determine the level of cell-free DNA (cfDNA), Septin 9 (SEPT9) and tumor markers (CEA, AFP, CA19–9, TPA, CA72–4). Plasma samples were collected four times a day (06:00, 12:00, 18:00, 24:00) from 9 patients with CRC (5 stage I-II, 4 stage III-IV), from one with colorectal adenoma and from one healthy control. CfDNA was isolated, quantified and bisulfite-converted. CfDNA and methylated SEPT9 were determined by RT-PCR. Plasma levels of conventional tumor markers were also measured. The lowest cfDNA concentrations were observed at 24:00 and 18:00 in stage I-III patients. In stage IV samples low cfDNA level (mean 48.2 ng/ml) were observed at several time points (6:00, 12:00, 18:00). The highest cfDNA levels were measured at 6:00 and 12:00 in CRC I-III stages and at 24:00 in stage IV samples (78.65 ng/ml). Higher in-day differences were found in stage II (43–48%) than in stage I samples (22%). Interestingly, the highest SEPT9 methylation level was found at 24:00 in most CRC cases, in contrast to the cfDNA levels. At 24:00, all cancer and adenoma cases were positive for SEPT9 methylation. At other time points (6:00, 12:00, 18:00) only 77.7% of CRC samples showed SEPT9 positivity. Stage I samples were SEPT9 positive only at 24:00. CEA and CA19–9 levels displayed correlation with the amount of cfDNA in case of late stage cases. Daytime activity can influence SEPT9 positivity in cases with low concentration of cfDNA. Thus, it may improve screening sensitivity by collecting samples earlier in the morning.

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Acknowledgements

We thank both the Endoscopy Unit of the 2nd Department of Internal Medicine, Semmelweis University, and the nurses at the 2nd Department of Internal Medicine for their technical assistance. We also thank Bernadett Tóth for blood sample and data collection and plasma preparation. We thank Prof. Barna Vásárhelyi, Tünde Holczer and all the colleagues at the Department of Laboratory Medicine for the technical assistance. Finally, we thank all of the included patients for the blood collection.

The authors thank Theo deVos the help, scientific comments and support our review paper.

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Authors and Affiliations

  1. 2nd Department of Internal Medicine, Semmelweis University, Szentkirályi utca 46, Budapest, 1088, Hungary

    Kinga Tóth, Árpád V. Patai, Alexandra Kalmár, Barbara Kinga Barták, Zsófia Brigitta Nagy, Zsolt Tulassay & Béla Molnár

  2. Molecular Medicine Research Unit, Hungarian Academy of Sciences, Budapest, Hungary

    Alexandra Kalmár, Orsolya Galamb, Barnabás Wichmann, Zsolt Tulassay & Béla Molnár

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  1. Kinga Tóth
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Correspondence to Kinga Tóth.

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Tóth, K., Patai, Á.V., Kalmár, A. et al. Circadian Rhythm of Methylated Septin 9, Cell-Free DNA Amount and Tumor Markers in Colorectal Cancer Patients. Pathol. Oncol. Res. 23, 699–706 (2017). https://doi.org/10.1007/s12253-016-0174-2

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