Electric Field-Induced Release and Measurement Liquid Biopsy for Noninvasive Early Lung Cancer Assessment.

Previously, we detected circulating tumor DNA that contained two EGFR mutations (p.L858R and exon19 del) in plasma of patients with late-stage non-small-cell lung carcinoma (NSCLC) using the electric field-induced release and measurement (EFIRM) platform. Our aim was to determine whether EFIRM technology can detect these mutations in patients with early-stage NSCLC. Prospectively, 248 patients with radiographically determined pulmonary nodules were recruited. Plasma was collected before biopsy and histologic examination of the nodule. Inclusion criteria were histologic diagnosis of benign nodule (control) and stage I or II adenocarcinoma harboring either p.L858R or exon19 delEGFR mutations. Plasma samples were available from 44 patients: 23 with biopsy-proven benign pulmonary nodules and 21 with stage I or II adenocarcinoma (12 p.L858R and 9 exon19 delEGFR variants). Samples were analyzed for the EGFR mutations using the EFIRM platform. Assay sensitivity was 92% for p.L858R (11 of 12 samples positive) and 77% for exon19 del (7 of 9 samples positive). Specificity was 91% with two false-positive results in 23 patients with EGFR-positive nodules and 95% for the entire 44-patient series. Concordance was 100% with identical mutations discovered in plasma and nodule biopsy. The EFIRM platform is able to noninvasively detect two EGFR mutations in individuals with early-stage NSCLC.

Assessing the necessity of confirmatory testing for exome-sequencing results in a clinical molecular diagnostic laboratory.

PURPOSE: Sanger sequencing is currently considered the gold standard methodology for clinical molecular diagnostic testing. However, next-generation sequencing has already emerged as a much more efficient means to identify genetic variants within gene panels, the exome, or the genome. We sought to assess the accuracy of next-generation sequencing variant identification in our clinical genomics laboratory with the goal of establishing a quality score threshold for confirmatory Sanger-based testing. METHODS: Confirmation data for reported results from 144 sequential clinical exome-sequencing cases (94 unique variants) and an additional set of 16 variants from comparable research samples were analyzed. RESULTS: Of the 110 total single-nucleotide variants analyzed, 103 variants had a quality score ≥Q500, 103 (100%) of which were confirmed by Sanger sequencing. Of the remaining seven variants with quality scores

Current landscape and new paradigms of proficiency testing and external quality assessment for molecular genetics.

CONTEXT: Participation in proficiency testing (PT) or external quality assessment (EQA) programs allows the assessment and comparison of test performance among different clinical laboratories and technologies. In addition to the approximately 2300 tests for individual genetic disorders, recent advances in technology have enabled the development of clinical tests that quickly and economically analyze the entire human genome. New PT/EQA approaches are needed to ensure the continued quality of these complex tests. OBJECTIVES: To review the availability and scope of PT/EQA for molecular genetic testing for inherited conditions in Europe, Australasia, and the United States; to evaluate the successes and demonstrated value of available PT/EQA programs; and to examine the challenges to the provision of comprehensive PT/EQA posed by new laboratory practices and methodologies. DATA SOURCES: The available literature on this topic was reviewed and supplemented with personal experiences of several PT/EQA providers. CONCLUSIONS: Proficiency testing/EQA schemes are available for common genetic disorders tested in many clinical laboratories but are not available for most genetic tests offered by only one or a few laboratories. Provision of broad, method-based PT schemes, such as DNA sequencing, would allow assessment of many tests for which formal PT is not currently available. Participation in PT/EQA improves the quality of testing by identifying inaccuracies that laboratories can trace to errors in their testing processes. Areas of research and development to ensure that PT/EQA programs can meet the needs of new and evolving genetic tests and technologies are identified and discussed.

Opportunities and challenges associated with clinical diagnostic genome sequencing: a report of the Association for Molecular Pathology.

This report of the Whole Genome Analysis group of the Association for Molecular Pathology illuminates the opportunities and challenges associated with clinical diagnostic genome sequencing. With the reality of clinical application of next-generation sequencing, technical aspects of molecular testing can be accomplished at greater speed and with higher volume, while much information is obtained. Although this testing is a next logical step for molecular pathology laboratories, the potential impact on the diagnostic process and clinical correlations is extraordinary and clinical interpretation will be challenging. We review the rapidly evolving technologies; provide application examples; discuss aspects of clinical utility, ethics, and consent; and address the analytic, postanalytic, and professional implications.

Should we be applying warfarin pharmacogenetics to clinical practice? No, not now.

The U.S. Food and Drug Administration modified warfarin labeling in 2007 to suggest, but not mandate, pharmacogenetic testing. Genetic analysis is now commercially available. However, results predict only one third of all dosing variation, the value of testing in reducing bleeding and thrombosis rates remains unproved, and cost-effectiveness is not established. Careful consideration of clinical factors that influence dosing, conscientious prothrombin time monitoring, and sage dosage adjustment remain paramount in warfarin management. Further study is required before routine warfarin pharmacogenetic testing can be recommended.

Keeping up with the next generation: massively parallel sequencing in clinical diagnostics.

The speed, accuracy, efficiency, and cost-effectiveness of DNA sequencing have been improving continuously since the initial derivation of the technique in the mid-1970s. With the advent of massively parallel sequencing technologies, DNA sequencing costs have been dramatically reduced. No longer is it unthinkable to sequence hundreds or even thousands of genes in a single individual with a suspected genetic disease or complex disease predisposition. Along with the benefits offered by these technologies come a number of challenges that must be addressed before wide-scale sequencing becomes accepted medical practice. Molecular diagnosticians will need to become comfortable with, and gain confidence in, these new platforms, which are based on radically different technologies compared to the standard DNA sequencers in routine use today. Experience will determine whether these instruments are best applied to sequencing versus resequencing. Perhaps most importantly, along with increasing read lengths inevitably comes increased ascertainment of novel sequence variants of uncertain clinical significance, the postanalytical aspects of which could bog down the entire field. But despite these obstacles, and as a direct result of the promises these sequencing advances present, it will likely not be long before next-generation sequencing begins to make an impact in molecular medicine. In this review, technical issues are discussed, in addition to the practical considerations that will need to be addressed as advances push toward personal genome sequencing.