Clinical Application Of Next-generation Sequencing Research Articles

Abstract IA11: Clinical application of next-generation sequencing as a guide to treatment selection

Abstract Profound and durable responses are often observed in early stage clinical trials of novel cancer agents in only a small minority of patients. It has long been postulated that these responses have a definable genetic basis but until now it was not feasible to perform a comprehensive genomic analysis of such patients. Rather, at best a few candidate genes were examined. Technical feasibility thus ensured that oncology trials were designed to identify agents that have a statistically significant benefit in a genetically unselected population, a paradigm that has led to the development of many agents that have modest or no benefit in the vast majority of patients. Agents with profound activity in only a small number of patients were on the other had deemed inactive and abandoned. Our preliminary experience shows that next generation sequencing methodology can now be feasibly applied in the clinical setting to identify previously unrecognized biomarkers of response for agents that show profound responses in only a minority of patients. As an example, we explored the molecular basis of an outlier phenotype, an apparent disease cure of a 51-year old woman with recurrent metastatic small cell carcinoma. The patient was treated on a phase 1 clinical trial that combined topoisomerase I inhibition with an ATP-competitive inhibitor of checkpoint kinase 1 (Chk1). Whole genome sequencing revealed a complex but highly clonal tumor genome with a somatic, missense mutation in the Mre11 complex gene RAD50, which functions to initiate double stranded break (DSB) repair by homologous recombination or non- homologous end joining. Through modeling in yeast, we confirmed that this heterozygous RAD50 mutation, which affects a highly conserved residue and whose wild type copy was focally deleted, conferred sensitivity to topoisomerase I inhibition. Drug sensitivity was markedly enhanced upon genetic ablation of the DNA damage checkpoint pathway, suggesting a synthetic lethal interaction between checkpoint kinase inhibition and clastogenic chemotherapy. These results and additional examples to be presented demonstrate the feasibility of using whole-genome sequencing in the clinical setting to identify previously occult biomarkers of drug sensitivity that can aid in the identification of patients most likely to respond to targeted anticancer drugs. The use of novel clinical trial designs to confirm genotype-phenotype associations will also be discussed. Citation Format: David B. Solit. Clinical application of next-generation sequencing as a guide to treatment selection. [abstract]. In: Proceedings of the AACR Precision Medicine Series: Integrating Clinical Genomics and Cancer Therapy; Jun 13-16, 2015; Salt Lake City, UT. Philadelphia (PA): AACR; Clin Cancer Res 2016;22(1_Suppl):Abstract nr IA11.

Capture-based high-coverage NGS: a powerful tool to uncover a wide spectrum of mutation types.

Next-generation sequencing (NGS) has been widely applied to clinical diagnosis. Target-gene capture followed by deep sequencing provides unbiased enrichment of the target sequences, which not only accurately detects single-nucleotide variations (SNVs) and small insertion/deletions (indels) but also provides the opportunity for the identification of exonic copy-number variants (CNVs) and large genomic rearrangements. Capture NGS has the ability to easily detect SNVs and small indels. However, genomic changes involving exonic deletions/duplications and chromosomal rearrangements require more careful analysis of captured NGS data. Misaligned raw sequence reads may be more than just bad data. Some mutations that are difficult to detect are filtered by the preset analytical parameters. "Loose" filtering and alignment conditions were used for thorough analysis of the misaligned NGS reads. Additionally, using an in-house algorithm, NGS coverage depth was thoroughly analyzed to detect CNVs. Using real examples, this report underscores the importance of the accessibility to raw sequence data and manual review of suspicious sequence regions to avoid false-negative results in the clinical application of NGS. Assessment of the NGS raw data generated by the use of loose filtering parameters identified several sequence aberrations, including large indels and genomic rearrangements. Furthermore, NGS coverage depth analysis identified homozygous and heterozygous deletions involving single or multiple exons. Our results demonstrate the power of deep NGS in the simultaneous detection of point mutations and intragenic exonic deletion in one comprehensive step.Genet Med 18 5, 513-521.

Clinical application of next-generation sequencing for Mendelian diseases

Over the past decade, next-generation sequencing (NGS) has led to an exponential increase in our understanding of the genetic basis of Mendelian diseases. NGS allows for the analysis of multiple regions of the genome in one single reaction and has been shown to be a cost-effective and efficient tool in investigating patients with Mendelian diseases. More recently, NGS has been successfully deployed in the clinics, with a reported diagnostic yield of ~25 %. However, recommendations on clinical implementation of NGS are still evolving with numerous key challenges that impede the widespread use of genetics in everyday medicine. These challenges include when to order, on whom to order, what type of test to order, and how to interpret and communicate the results, including incidental findings, to the patient and family. In this review, we discuss these challenges and suggest guidelines on implementing NGS in the routine clinical workflow.

Genome Medicine in Cancer: What's in a Name?

This is an exciting time to be in cancer medicine. New technologies, such as next-generation sequencing (NGS), have increased our understanding of the molecular aberrations that define cancer. This, in turn, has led to the identification of cancer-specific molecular targets and potential drugs to confront these targets. As these new technologies move toward clinical application, a new vocabulary of "genome medicine" has been introduced to the field of oncology. Unfortunately, unclear or incorrect use of the new terminology has led to semantic misunderstandings that impair communication between the basic research and clinical practice arenas. These misunderstandings have led to assumptions regarding the clinical application of NGS and other technologies that may or may not be true. For example, some organizations that perform NGS testing on clinical samples have endorsed use of the results of such tests to direct specific therapies based on laboratory hypotheses, but without clinical testing of the hypotheses to show utility for these potential predictive claims. Here, we review some simple, and hopefully universally acceptable, definitions, concepts, and trial designs so that laboratory researchers and clinicians can move closer toward speaking the same language.

Real-time clinical application of next-generation sequencing (NGS): Results from a multicenter program.

11016 Background: NGS techniques enable the identification of actionable mutations in clinical tumor samples. The objective of this study is to assess feasibility and explore the impact of real-time targeted NGS on therapeutic decision-making. Methods: Patients (pts) with advanced solid tumors underwent a biopsy of a metastatic lesion. The first phase was performed with Sequenom MassARRAY somatic genotyping and Pacific Biosciences RS-targeted NGS. The second phase broadened genomic coverage in both Sequenom and Illumina MiSeq. All pts had a molecular profiling (mp) report issued after identified actionable mutations were verified by Sanger sequencing in a CLIA-lab and reviewed by an expert panel. “Actionability” was defined as having prognostic, predictive or diagnostic implications on patient management. Details of clinical outcomes and subsequent matched therapy, if applicable, were captured. Referring physicians were surveyed on the impact of mutation results on their treatment recommendations. Results: These are summarized in the Table. Conclusions: Broader mp platforms resulted in more identified actionable mutations which required a longer time for verification prior to reporting, but may yield a greater impact on clinical decision-making. However, the matching of pts to drugs based on their molecular profiles depends highly on drug access. For mp to be clinically relevant, it must be coupled with access to approved drugs or to investigational agents on clinical trials. Clinical trial information: NCT01345513. [Table: see text]

Integrating next‐generation sequencing into the diagnostic testing of inherited cancer predisposition

The clinical application of next-generation sequencing (NGS) as a diagnostic tool has become increasingly evident. The coupling of NGS technologies with new genomic sequence enrichment methods has made the sequencing of panels of target genes technically feasible, at the same time as making such an approach cost-effective for diagnostic applications. In this article, we discuss recent studies that have applied NGS in the diagnostic setting in relation to hereditary cancer.

Deploying next-generation sequencing in a hospital setting

Personalized medicine is the ability to tailor healthcare decisions based on an individual's unique characteristics (genetics, demographic information, healthcare experience, environment and social factors) to more accurately diagnose the individual's disease, predict its outcomes, and select treatments that increase the chances of a successful outcome and reduce possible adverse reactions. Moreover, it is the ability to predict an individual's susceptibility to diseases with the goal of taking measures to prevent or mitigate the extent to which an individual will experience a disease. The promise of personalized medicine in healthcare delivery includes: • avoiding the cost of a drug or therapy with little chance of success for each patient • avoiding exposing patients to side-effects from the wrong treatment • avoiding increased disease burden from delays in finding the right treatment • being able to act preventatively in cases where we can predict disease susceptibility, to delay onset and/or progression. These changes in healthcare delivery promise improved patient outcomes as well as reduced healthcare and societal costs of managing disease. A key component in personalized medicine is the emergence of whole-genome-scale sequencing as a platform to identify gene variants. Many challenges exist, however, to the use of large-scale sequencing in disease gene discovery and the efficacious application of such knowledge to health benefit. Leveraging upon the Ontario Government's CAD$7 million committed investment to build a clinical genomics operational framework for personalized healthcare at Mount Sinai Hospital, our project is developing a portfolio of computational tools and methods to enable acquisition, analysis, interpretation and reporting of high-throughput sequence data and the ethical, economic, psychosocial and knowledge synthesis support needed to move deep sequencing datasets and their findings into clinical practice. The challenges to moving next-generation sequencing (NGS) into the hospital setting are both technical and regulatory. Since introduced into the research community in 2007, broad use of NGS has been greatly enabled by development of sophisticated informatics and analytic tools. While such tools have reduced many of the barriers to clinical application of NGS, the investment needed to bring NGS into medical practice remains significant, the scale of IT required being unprecedented at most hospitals. In this context, hospitals are, to some extent, at a similar place to that of research institutes when NGS first emerged. Some of the IT barriers to clinical use of NGS have been bypassed by the expertise in NGS data management developed at genome centers and across the research community. However, even this wealth of experience does not fully address the road blocks inherent to integrating NGS information into existing health workflows and IT systems, and the added challenges posed by the regulatory controls in place across the health system. This talk will describe strategies our group is using to overcome these hurdles and thereby expedite clinical integration of NGS capability and its translation to delivery of personalized healthcare.

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.