Technical Evaluation of Commercial Mutation Analysis Platforms and Reference Materials for Liquid Biopsy Profiling

Molecular profiling from liquid biopsy, in particular cell-free DNA (cfDNA), represents an attractive alternative to tissue biopsies for the detection of actionable targets and tumor monitoring. In addition to PCR-based assays, Next Generation Sequencing (NGS)-based cfDNA assays are now commercially available and are being increasingly adopted in clinical practice. However, the validity of these products as well as the clinical utility of cfDNA in the management of patients with cancer has yet to be proven. Within framework of the Innovative Medicines Initiative (IMI) program CANCER-ID we evaluated the use of commercially available reference materials designed for ctDNA testing and cfDNA derived from Diagnostic Leukaphereses (DLA) for inter-and intra-assay as well as intra-and inter-laboratory comparisons. In three experimental setups, a broad range of assays including ddPCR, MassARRAY and various NGS-based assays were tested. We demonstrate that both reference materials with predetermined VAFs and DLA samples are extremely useful for the performance assessment of mutation analysis platforms. Moreover, our data indicate a substantial variability of NGS assays with respect to sensitivity and specificity

Using Bioinformatics to Uncover Novel Transcription Factor Binding Sites

In response to a changing environment, cells turn off the expression of some genes and turn on the expression of others. In eukaryotes, this is a complex process involving both the binding of one or more transcription factors in the vicinity of the affected genes 1,2,3,4,5 as well as chromatin remodeling 6. Some transcription factors bind immediately upstream of a gene in th

A case-control study on the relationship between HDL2b and non-alcoholic fatty liver disease in Chinese type 2 diabetic patients

Background: It has been shown that low concentrations of high-density lipoprotein cholesterol (HDL-C) are associated with non-alcoholic fatty liver disease (NAFLD) in patients with type 2 diabetes mellitus (T2DM). HDL2b, a major subfraction of high-density lipoprotein (HDL), is more significantly correlated with coronary heart disease (CHD) compared with total HDL. In this study, we analysed HDL2b in a cohort of Chinese T2DM subjects with or without NAFLD. Methods: A highly sensitive and reliable microfluidic chip method was adopted to measure HDL2b. In total, 48 T2DM patients with NAFLD diagnosed by a B-ultrasound were enrolled from our Beijing Community Pre-Diabetes (BCPD) study cohort. A total of 48 age and gender matched diabetic controls without NAFLD were selected from the same population. Results: Clinical characteristics and serum biochemical analyses demonstrated a significantly increased body mass index (BMI), waist circumference, homeostasis model assessment-insulin resistant index (HOMA-IR), total cholesterol (TC), and triglyceride (TG) concentrations in the NAFLD group. However, the concentrations of HDL2b and its ratio to total HDL in NAFLD patients was decreased, compared with controls (p<0.01). Significantly increased concentrations of high sensitive C-reactive protein (hs-CRP) (p<0.01) were also found. Multifactor logistic regression analysis showed that BMI and TG were the predominant risk factors for fatty liver, while HDL2b was a protective factor. Conclusions: T2DM patients with NAFLD have characteristics including obesity, marked insulin resistance, high TG, high hs-CRP, low HDL2b and a low HDL2b ratio to total HDL. These factors may increase the incidence of atherosclerosis as well as the risk of CHD. Clin Chem Lab Med 2009;47:1067–72.Peer Reviewe