Performance characteristics of a polymerase chain reaction-based assay for the detection of EGFR mutations in plasma cell-free DNA from patients with non-small cell lung cancer using cell-free DNA collection tubes.

Survival rates in non-small cell lung cancer (NSCLC) are low. Detection of circulating tumor DNA in liquid biopsy (plasma) is increasingly used to identify targeted therapies for clinically actionable mutations, including EGFR mutations in NSCLC. The cobas® EGFR Mutation Test v2 (cobas EGFR test) is FDA-approved for EGFR mutation detection in tissue or liquid biopsy from NSCLC. Standard K2EDTA tubes require plasma separation from blood within 4 to 8 hours; however, Roche Cell-Free DNA (cfDNA) Collection Tubes (Roche cfDNA tube) enable whole blood stability for up to 7 days prior to plasma separation. This analysis assessed performance of Roche cfDNA tubes with the cobas EGFR test for the detection of EGFR mutations in plasma from healthy donors or patients with NSCLC. Overall, test performance was equally robust with either blood collection tube, eg, regarding limit of detection, linearity, and reproducibility, making Roche cfDNA tubes suitable for routine clinical laboratory use in this setting. Importantly, the Roche cfDNA tubes provided more flexibility for specimen handling versus K2EDTA tubes, eg, in terms of tube mixing, plasma separation, and sample stability, and do not require processing of blood within 8 hours thereby increasing the reach of plasma biopsies in NSCLC.

Eating more sardines instead of fish oil supplementation: Beyond omega-3 polyunsaturated fatty acids, a matrix of nutrients with cardiovascular benefits.

Omega-3 polyunsaturated fatty acids (n-3 PUFA) play a significant role in the prevention and management of cardiometabolic diseases associated with a mild chronic pro-inflammatory background, including type 2 diabetes, hypertension, hypertriglyceridaemia, and fatty liver disease. The effects of n-3 PUFA supplements specifically, remain controversial regarding reducing risks of cardiovascular events. n-3 PUFA supplements come at a cost for the consumer and can result in polypharmacy for patients on pharmacotherapy. Sardines are a well-known, inexpensive source of n-3 PUFA and their consumption could reduce the need for n-3 PUFA supplementation. Moreover, sardines contain other cardioprotective nutrients, although further insights are crucial to translate a recommendation for sardine consumption into clinical practice. The present review discusses the matrix of nutrients contained in sardines which confer health benefits for cardiometabolism, beyond n-3 PUFA. Sardines contain calcium, potassium, magnesium, zinc, iron, taurine, arginine and other nutrients which together modulate mild inflammation and exacerbated oxidative stress observed in cardiovascular disease and in haemodynamic dysfunction. In a common serving of sardines, calcium, potassium, and magnesium are the minerals at higher amounts to elicit clinical benefits, whilst other nutrients are present in lower but valuable amounts. A pragmatic approach towards the consumption of such nutrients in the clinical scenario should be adopted to consider the dose-response relationship effects on physiological interactions. As most recommendations currently available are based on an indirect rationale of the physiological actions of the nutrients found in sardines, randomised clinical trials are warranted to expand the evidence on the benefits of sardine consumption.

Innovative Tumor Tissue Dissection Tool for Molecular Oncology Diagnostics.

Formalin-fixed, paraffin-embedded (FFPE) tissue is the most commonly used material for tumor molecular profiling, therapy selection, and prognostication. Tumor tissue enrichment by tissue dissection is highly recommended to generate quality data reproducibly for use in downstream assays, such as real-time PCR and next-generation sequencing. The aim of this study was to evaluate the performance of the automated tissue dissection tool AVENIO Millisect System compared with a manual dissection method using 18 FFPE tissue specimens. The study assessed performance of these two methods with paraffinized and deparaffinized sections at 5- and 10-µm thickness as well as at low (5% to 10%) and high (>50%) tumor content. In addition, compatibility with various nucleic acid and protein extraction methods was assessed. Overall, dissection by Millisect resulted in statistically significantly higher yields of nucleic acids and protein compared with manual dissection (P = 0.00524). In downstream analysis on a statistically nonpowered sample set, EGFR mutation testing by PCR led to highly concordant results, and next-generation sequencing testing yielded significantly higher allelic frequencies when tissue was dissected by Millisect compared with manual scraping, demonstrating noninferiority of the automated method. In summary, the AVENIO Millisect System may replace manual labor and support automation of FFPE tumor tissue workflows in clinical molecular laboratories with high testing volumes with adequate validation.

Performance Characteristics of a Real-Time Polymerase Chain Reaction Assay for the Detection of Epidermal Growth Factor Receptor (EGFR) Mutations in Plasma Samples of Non-Small Cell Lung Cancer (NSCLC) Patients.

BACKGROUND: Circulating free DNA in plasma is an alternative source of tumor-derived DNA that can be a surrogate for tissue epidermal growth factor receptor (EGFR) testing. OBJECTIVE: We evaluated the analytical performance of the cobas® EGFR Mutation Test v2 (cobas test), a real-time polymerase chain reaction assay designed to detect defined EGFR gene mutations in plasma from patients with advanced non-small cell lung cancer (NSCLC). METHODS: We used K2-ethylenediaminetetraacetic acid plasma samples from NSCLC patients and healthy donors (HDs), along with cell line DNA. Results from a complete technical performance evaluation are described, including a comparison between NSCLC and HD plasma to support the use of surrogate samples and an independent confirmation of the limit of detection (LoD). RESULTS: The cobas test reported an overall percent agreement of approximately 88% for plasma samples when compared with a next-generation sequencing method. The LoD for all EGFR mutations was ≤ 100 copies/mL for plasma samples. An external study confirmed the LoD for exon 19 deletion, L858R, and T790M at ≤ 100 copies/mL using samples derived from NSCLC patient specimens. The cobas test showed linearity between at least 50 and 10,000 copies/mL for plasma samples. An internal repeatability study reported a correct call accuracy of 99.2% for plasma samples. The performance of the cobas test is equivalent when using sheared or intact cell line DNA diluted into either HD plasma or NSCLC patient plasma. CONCLUSIONS: The cobas test is a sensitive, robust, and accurate assay that delivers reproducible results.

Functional Outcomes and Delayed Cerebral Ischemia Following Nonperimesencephalic Angiogram-Negative Subarachnoid Hemorrhage Similar to Aneurysmal Subarachnoid Hemorrhage.

BACKGROUND: The angiogram-negative subarachnoid hemorrhage (SAH) literature includes patients with perimesencephalic hemorrhage, which is recognized to have a much better outcome than aneurysmal SAH. OBJECTIVE: To evaluate the clinical outcomes of Nonperimesencephalic Angiogram-Negative SAH (NPAN-SAH). METHODS: A prospective, spontaneous SAH database of 1311 patients that accrued between April 2006 and December 2014 was screened. All patients with NPAN-SAH and 2 consecutive negative cerebral angiograms were included. RESULTS: We identified 191 (11%) from a total of 1311 patients with spontaneous SAH. Amongst angiogram-negative patients, 83 (4.9%) were adjudicated to have NPAN-SAH. Patient characteristics were similar across the groups, except NPAN-SAH patients were more likely to be men and had higher rates of diabetes. In a multivariable logistic regression model, NPAN-SAH patients were less likely to develop vasospasm, after adjusting for Fisher grade, sex, and diabetes (odds ratio [OR]: 0.197, 95% confidence interval [CI; 0.07-0.55], P = .002). In another adjusted model accounting for Hunt and Hess clinical grade, NPAN-SAH patients were also less likely to develop vasospasm (OR: 0.2, 95% CI [0.07-0.57], P = .002). We found no statistical significance between 2 groups for rebleed, developing hydrocephalus, seizures, or delayed cerebral ischemia. NPAN-SAH patients were equally associated with poor functional outcome (modified Rankin scale ≥3; OR: 1.16, 95% CI [0.615-2.20], P = .6420), and death (OR: 1.22, 95% CI [0.362-4.132], P = .7455) compared to aneurysmal SAH. CONCLUSION: Although the risk of vasospasm may be lower, patients with NPAN-SAH are equally associated with delayed cerebral ischemia, poor outcome, and death as compared to patients with aneurysmal SAH. Furthers studies may be necessary to further clarify these findings.