Exploring the feasibility and utility of exome-scale tumour sequencing in a clinical setting.

Background: Technology has progressed from single gene panel to large-scale genomic sequencing. This is raising expectations from clinicians and patients alike. The utility and performance of this technology in a clinical setting needs to be evaluated.

Sensitive NPM1 Mutation Quantitation in Acute Myeloid Leukemia Using Ultradeep Next-Generation Sequencing in the Diagnostic Laboratory.

Context Detection of measurable residual disease after therapy is an important predictor of outcome in acute myeloid leukemia. Objective To investigate the feasibility of using next-generation sequencing (NGS) in the diagnostic laboratory to perform quantitative NPM1 mutation assessment using ultradeep (approximately 300 000×-500 000×) sequencing (NGS-q NPM1) as a method of assessing residual disease burden in patients with acute myeloid leukemia. Design A flexible NGS-based assay for the detection and quantitation of NPM1 mutations was developed by polymerase chain reaction amplification of target DNA sequences, sequencing on an Illumina (San Diego, California) MiSeq, and analyzing data with an in-house-designed bioinformatic pipeline.

Reversion of BRCA1/2 Germline Mutations Detected in Circulating Tumor DNA From Patients With High-Grade Serous Ovarian Cancer.

Purpose Germline BRCA1 or BRCA2 mutations in patients with high-grade serous ovarian cancer (HGSC) are associated with favorable responses to chemotherapy. However, secondary intragenic (reversion) mutations that restore protein function lead to clinically significant rates of acquired resistance. The goal of this study was to determine whether reversion mutations could be found in an unbiased manner in circulating cell-free DNA (cfDNA) to predict treatment response in HGSC.

Clinicopathological differences exist between CALR- and JAK2-mutated myeloproliferative neoplasms despite a similar molecular landscape: data from targeted next-generation sequencing in the diagnostic laboratory.

Mutations in CALR have recently been detected in JAK2-negative myeloproliferative neoplasms (MPNs) and are key pathological drivers in these diseases. CALR-mutated MPNs are shown to have numerous clinicopathological differences to JAK2-mutated MPNs. The basis of these differences is poorly understood.

"Cancer 2015": A Prospective, Population-Based Cancer Cohort-Phase 1: Feasibility of Genomics-Guided Precision Medicine in the Clinic.

"Cancer 2015" is a longitudinal and prospective cohort. It is a phased study whose aim was to pilot recruiting 1000 patients during phase 1 to establish the feasibility of providing a population-based genomics cohort. Newly diagnosed adult patients with solid cancers, with residual tumour material for molecular genomics testing, were recruited into the cohort for the collection of a dataset containing clinical, molecular pathology, health resource use and outcomes data.

Sequence artefacts in a prospective series of formalin-fixed tumours tested for mutations in hotspot regions by massively parallel sequencing.

Background: Clinical specimens undergoing diagnostic molecular pathology testing are fixed in formalin due to the necessity for detailed morphological assessment. However, formalin fixation can cause major issues with molecular testing, as it causes DNA damage such as fragmentation and non-reproducible sequencing artefacts after PCR amplification. In the context of massively parallel sequencing (MPS), distinguishing true low frequency variants from sequencing artefacts remains challenging.

Studying cancer genomics through next-generation DNA sequencing and bioinformatics.

Cancer is a complex disease driven by multiple mutations acquired over the lifetime of the cancer cells. These alterations, termed somatic mutations to distinguish them from inherited germline mutations, can include single-nucleotide substitutions, insertions, deletions, copy number alterations, and structural rearrangements. A patient's cancer can contain a combination of these aberrations, and the ability to generate a comprehensive genetic profile should greatly improve patient diagnosis and treatment.

Bioinformatics pipelines for targeted resequencing and whole-exome sequencing of human and mouse genomes: a virtual appliance approach for instant deployment.

Targeted resequencing by massively parallel sequencing has become an effective and affordable way to survey small to large portions of the genome for genetic variation. Despite the rapid development in open source software for analysis of such data, the practical implementation of these tools through construction of sequencing analysis pipelines still remains a challenging and laborious activity, and a major hurdle for many small research and clinical laboratories. We developed TREVA (Targeted REsequencing Virtual Appliance), making pre-built pipelines immediately available as a virtual appliance.

Massively-parallel sequencing assists the diagnosis and guided treatment of cancers of unknown primary.

The clinical management of patients with cancer of unknown primary (CUP) is hampered by the absence of a definitive site of origin. We explored the utility of massively-parallel (next-generation) sequencing for the diagnosis of a primary site of origin and for the identification of novel treatment options. DNA enrichment by hybridization capture of 701 genes of clinical and/or biological importance, followed by massively-parallel sequencing, was performed on 16 CUP patients who had defied attempts to identify a likely site of origin.