Enhancing clinical genomic accuracy with panelGC: a novel metric and tool for quantifying and monitoring GC biases in hybridization capture panel sequencing.

Accurate assessment of fragment abundance within a genome is crucial in clinical genomics applications such as the analysis of copy number variation (CNV). However, this task is often hindered by biased coverage in regions with varying guanine-cytosine (GC) content. These biases are particularly exacerbated in hybridization capture sequencing due to GC effects on probe hybridization and polymerase chain reaction (PCR) amplification efficiency.

Characterizing the G12C mutation in metastatic colorectal cancer: a population-based cohort and assessment of expression differences in The Cancer Genome Atlas.

Introduction: In metastatic colorectal cancer (mCRC), mutations impart inferior survival and resistance to anti-epidermal growth factor receptor (EGFR) antibodies. G12C inhibitors have been developed and we evaluated how G12C differs from other RAS mutations.

Patients And Methods: This retrospective review evaluated patients in British Columbia, Canada with mCRC and testing performed between 1 January 2016 and 31 December 2018.

Effect of preexamination conditions in a centralized-testing model of non-invasive prenatal screening.

Objectives: Non-invasive prenatal testing requires the presence of fetal DNA in maternal plasma. Understanding how preexamination conditions affect the integrity of cell-free DNA (cfDNA) and fetal fraction (FF) are a prerequisite for test implementation. Therefore, we examined the adjusted effect that EDTA and Streck tubes have on the cfDNA quantity and FF.

Use of Treatment-Focused Tumor Sequencing to Screen for Germline Cancer Predisposition.

Next-generation sequencing assays are capable of identifying cancer patients eligible for targeted therapies and can also detect germline variants associated with increased cancer susceptibility. However, these capabilities have yet to be routinely harmonized in a single assay because of challenges with accurately identifying germline variants from tumor-only data. We have developed the Oncology and Hereditary Cancer Program targeted capture panel, which uses tumor tissue to simultaneously screen for both clinically actionable solid tumor variants and germline variants across 45 genes.

A clinical transcriptome approach to patient stratification and therapy selection in acute myeloid leukemia.

As more clinically-relevant genomic features of myeloid malignancies are revealed, it has become clear that targeted clinical genetic testing is inadequate for risk stratification. Here, we develop and validate a clinical transcriptome-based assay for stratification of acute myeloid leukemia (AML). Comparison of ribonucleic acid sequencing (RNA-Seq) to whole genome and exome sequencing reveals that a standalone RNA-Seq assay offers the greatest diagnostic return, enabling identification of expressed gene fusions, single nucleotide and short insertion/deletion variants, and whole-transcriptome expression information.

Assessing Limit of Detection in Clinical Sequencing.

Clinical reporting of solid tumor sequencing requires reliable assessment of the accuracy and reproducibility of each assay. Somatic mutation variant allele fractions may be below 10% in many samples due to sample heterogeneity, tumor clonality, and/or sample degradation in fixatives such as formalin. The toolkits available to the clinical sequencing community for correlating assay design parameters with assay sensitivity remain limited, and large-scale empirical assessments are often relied upon due to the lack of clear theoretical grounding.

Population-based Screening for in Metastatic Colorectal Cancer Reveals Increased Prevalence and Poor Prognosis.

Purpose: mutations portend poor prognosis in metastatic colorectal cancer (mCRC); however, the true prevalence and prognosis are unknown, as unwell patients may not undergo sequencing.

Experimental Design: We reviewed a population-based cohort of 1,898 patients with colorectal cancer that underwent reflexive IHC mismatch repair (MMR) and testing. Outcomes among IHC-detected mCRC ( ) were compared with patients with next-generation sequencing (NGS)-identified -mutated mCRC from two institutions ( ) with patients spanning from 2004 to 2018.

Sample Tracking Using Unique Sequence Controls.

Sample tracking and identity are essential when processing multiple samples in parallel. Sequencing applications often involve high sample numbers, and the data are frequently used in a clinical setting. As such, a simple and accurate intrinsic sample tracking process through a sequencing pipeline is essential.

Fixation Effects on Variant Calling in a Clinical Resequencing Panel.

Formalin fixation is the standard method for the preservation of tissue for diagnostic purposes, including pathologic review and molecular assays. However, this method is known to cause artifacts that can affect the accuracy of molecular genetic test results. We assessed the applicability of alternative fixatives to determine whether these perform significantly better on next-generation sequencing assays, and whether adequate morphology is retained for primary diagnosis, in a prospective study using a clinical-grade, laboratory-developed targeted resequencing assay.

Barnacle: detecting and characterizing tandem duplications and fusions in transcriptome assemblies.

Background: Chimeric transcripts, including partial and internal tandem duplications (PTDs, ITDs) and gene fusions, are important in the detection, prognosis, and treatment of human cancers.

Results: We describe Barnacle, a production-grade analysis tool that detects such chimeras in de novo assemblies of RNA-seq data, and supports prioritizing them for review and validation by reporting the relative coverage of co-occurring chimeric and wild-type transcripts. We demonstrate applications in large-scale disease studies, by identifying PTDs in MLL, ITDs in FLT3, and reciprocal fusions between PML and RARA, in two deeply sequenced acute myeloid leukemia (AML) RNA-seq datasets.

Genomic and epigenomic landscapes of adult de novo acute myeloid leukemia.

Background: Many mutations that contribute to the pathogenesis of acute myeloid leukemia (AML) are undefined. The relationships between patterns of mutations and epigenetic phenotypes are not yet clear.

Methods: We analyzed the genomes of 200 clinically annotated adult cases of de novo AML, using either whole-genome sequencing (50 cases) or whole-exome sequencing (150 cases), along with RNA and microRNA sequencing and DNA-methylation analysis.

Dissect: detection and characterization of novel structural alterations in transcribed sequences.

Motivation: Computational identification of genomic structural variants via high-throughput sequencing is an important problem for which a number of highly sophisticated solutions have been recently developed. With the advent of high-throughput transcriptome sequencing (RNA-Seq), the problem of identifying structural alterations in the transcriptome is now attracting significant attention. In this article, we introduce two novel algorithmic formulations for identifying transcriptomic structural variants through aligning transcripts to the reference genome under the consideration of such variation.