Admission Cell Free DNA Levels Predict 28-Day Mortality in Patients with Severe Sepsis in Intensive Care

et al. (2014) Admission Cell Free DNA Levels Predict 28-Day Mortality in Patients with Severe Sepsis in Intensive Care. PLOS ONE 9(6): e100514. doi:10.1371/journal.pone.0100514 Admission Cell Free DNA Levels Predict 28-Day Mortality in Patients with Severe Sepsis in Intensive Care Avital Avriel Maya Paryente Wiessman Yaniv Almog Yael Perl Victor Novack Ori Galante Moti Klein Michael J. Pencina Amos Douvdevani Olivier Neyrolles, Institut de Pharmacologie et de Biologie Structurale, France Aim: The aim of the current study is to assess the mortality prediction accuracy of circulating cell-free DNA (CFD) level at admission measured by a new simplified method. Materials and Methods: CFD levels were measured by a direct fluorescence assay in severe sepsis patients on intensive care unit (ICU) admission. In-hospital and/or twenty eight day all-cause mortality was the primary outcome. Results: Out of 108 patients with median APACHE II of 20, 32.4% have died in hospital/or at 28-day. CFD levels were higher in decedents: median 3469.0 vs. 1659 ng/ml, p,0.001. In multivariable model APACHE II score and CFD (quartiles) were significantly associated with the mortality: odds ratio of 1.05, p = 0.049 and 2.57, p,0.001 per quartile respectively. Cstatistics for the models was 0.79 for CFD and 0.68 for APACHE II. Integrated discrimination improvement (IDI) analyses showed that CFD and CFD+APACHE II score models had better discriminatory ability than APACHE II score alone. Conclusions: CFD level assessed by a new, simple fluorometric-assay is an accurate predictor of acute mortality among ICU patients with severe sepsis. Comparison of CFD to APACHE II score and Procalcitonin (PCT), suggests that CFD has the potential to improve clinical decision making. - Data Availability: The authors confirm that all data underlying the findings are fully available without restriction. The clinical data of the patients is kept by our Clinical Research Center headed by Prof. Victor Novack (co-author of this manuscript). Funding: This work was supported by the Dr. Montague Robin Fleisher Kidney Transplant Unit Fund and by grant number 3-00000-6087 from the Chief Scientist, Israeli Ministry of Health. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. Competing Interests: Amos Douvdevani submitted a US Patent Application No. 13/659,439 Assay for Detecting Circulating Free Nucleic Acids. All other authors declare that they have no conflict of interest. The authors confirm that this does not alter the authors firm commitment to PLOS ONE standing policies on sharing data and materials. . These authors contributed equally to this work. Decision making process in the setting of the intensive care unit (ICU) is commonly supported by scoring methods such as the Acute Physiology And Chronic Health Evaluation (APACHE) II or the Sequential Organ Failure Assessment (SOFA) score [1,2]. A novel approach, which incorporates specific blood biomarkers has been put forward to improve the early diagnosis and assessment of patients with sepsis [3]. However, today there is no shelf-ready, accessible serological marker available for the routine clinical practice [4]. Circulating cell-free DNA (CFD), a product of cell necrosis, apoptosis and active secretion, is being investigated as a new reliable marker for assessing prognosis in various pathologies, including cancer, trauma and chronic renal failure (CRF) treated by hemodialysis [5,6,7,8]. Several studies evaluated the prognostic accuracy of CFD for ICU general-patients and septic-patients for death prediction and found good correlation with other outcome predicting scores and markers, such as APACHE, SOFA and CRP level [9,10,11,12,13,14]. In a recent publication, Dwivedi et al. found that their CFD assay had a sensitivity of 87.9% and specificity of 93.5% for predicting ICU mortality in patients with severe sepsis- better than multiple organ dysfunction (MOD) and APACHE II scores [14]. Although there is general agreement on the potential value of CFD measurement, to date there is no standard applicable method for routine clinical use. The currently available research methods for CFD measurement require DNA extraction and are labor-intensive. Consequently, CFD measurements are not utilized in the routine patient management. We have developed a novel rapid and direct fluorescent assay for CFD quantification that does not require DNA extraction and PCR amplification, and it has been shown to be inexpensive, accurate and reproducible [15]. Another potentially useful marker for prognostication in patients with severe sepsis is procalcitonin (PCT). PCT was shown to be a relatively reliable method for the outcome prediction in critically ill patients [4,12,16]. The aim of the current study is to assess the mortality prediction accuracy of CFD level at admission, measured by a new simplified method, as compared to the conventional clinical score and procalcitonin. Methods and Materials Population The study was conducted in Soroka University Medical Center, a tertiary 1000 bed hospital. The research protocol was approved by the Local Ethics Committee of the Soroka Medical Center, Beer-Sheva, Israel. Written Informed consent was obtained for patients capable of understanding the study procedures. Institutional review board allowed for an independent physician (not affiliated with the study team) to sign the proxy consent form in a case patient was unconscious (approval number 4636). The study population comprised severe sepsis patients admitted to ICU between March 2009 and 30th of April 2012. We excluded patients less than 18 years of age, pregnant, with CRF, malignancy, and were less than 14 days after surgical procedure (including coronal catheterization) or post trauma. Enrolled patients were assessed during the first 12 hours of ICU stay. CFD levels were measured within 12 hours from ICU admission. PCT levels were measured upon admission in 70 patients. PCT was determined using a chemiluminescent assay (Liaison Brahms PCT; DiaSorin S.P.A., Saluggia, Italy). We recorded clinical data related to the course of hospitalization, including need for invasive mechanical ventilation, treatment with vasopressors, occurrence of system organ failure and clinical outcomes. Severe sepsis and system organ failure was defined based on the ACCP/SCCM consensus document [17]. CFD measurements CFD was detected directly in sera using SYBR Gold Nucleic Acid Gel Stain, (Invitrogen, Paisley, UK) according to the fluorometric method we published [15]. Intra-day coefficient of variation was 16%, 7.9% and 4.8% in the low (383 ng/ml), elevated (1152 ng/ml) and high DNA range (2735 ng/mL), respectively. Day-to-day coefficient of variation was 31%, 6.7% and 8% in the low, elevated and high DNA range, respectively. This method was tested in comparison with the gold standard, QPCR to b-globin and was found to be i (...truncated)