By Diagnostics World Staff
March 30, 2017 | Last week, the Association for Molecular Pathology (AMP) published consensus recommendations to help clinical laboratory professionals achieve high-quality sequencing results and deliver better care for cancer patients. The report, “Guidelines for Validation of Next Generation Sequencing (NGS)-based Oncology Panels: A Joint Consensus Recommendation of the Association for Molecular Pathology and College of American Pathologists,” was released online in The Journal of Molecular Diagnostics (http://dx.doi.org/10.1016/j.jmoldx.2017.01.011).
The article is extensive, outlining recommendations for sample preparation, library prep, sequencing, data analysis, panel content, choosing a sequencing platform and method, test validation, controls, confirmatory and validation testing, and documentation. It particularly highlights the importance and role of professional pathologists.
In an email interview, Diagnostics World News asked Marina N. Nikiforova, Professor of Pathology at University of Pittsburgh Medical Center, Working Group Member, and 2016 AMP Clinical Practice Committee Chair, about the impetus for the recommendations and how she sees the field progressing.
Diagnostics World News: Why was this paper necessary? The techniques aren’t brand new anymore. What necessitated the extensive outline of the process?
Nikiforova: Although NGS technology itself is not new, targeted NGS panels have relatively recently become a method of choice for detection of somatic variants and are being rapidly adopted by clinical laboratories. The information generated by these panels can inform diagnostic classification, guide therapeutic decisions, and/or provide prognostic insights for a tumor. However, this method remains technically challenging given the variability in next-gen sequencing platforms and chemistries, and difficulty testing oncology samples, which often contain small proportions of neoplastic cells and poorly preserved nucleic acids. Therefore, clinical NGS requires thorough analytical validation to ensure the high quality of sequencing results. AMP believes that these standards are best established through the professional associations’ development of practice guidelines.
Although best practice recommendations for germline NGS testing were developed, the challenges of implementing and providing clinical NGS testing for somatic variant detection are significant enough to warrant their own guideline. AMP convened and led a multidisciplinary subject matter expert working group with liaison representation from CAP to summarize current knowledge, expose challenges, and provide best practice recommendations on how to ensure high-quality sequencing when used for patient care. This joint consensus recommendation represents a significant step forward in this era of precision medicine and the fight against cancer.
AMP Clinical Practice Committee is in the process of providing a series of guidelines designed to improve the entire NGS workflow including analytical, bioinformatics, and reporting aspects. The new report follows the previously published guidelines on interpreting oncology sequence variants. AMP plans to publish the next companion paper in this series focusing on downstream NGS bioinformatics analysis later this year.
Was AMP seeing trends or practices that raised red flags?
Cancer genomics is a rapidly evolving field. We are seeing the adoption of NGS technology by more clinical labs across the country, including small community labs, large academic centers, and commercial companies. These labs are developing and implementing more targeted oncology panels due to the expanding knowledge base of molecular alterations that initiate and drive tumor growth and metastasis. Given these factors, there was a growing community need for the development of a guidance document that will help to standardize the implementation of NGS technology among clinical molecular diagnostics laboratories ensuring best practices and high-quality patient care.
This first version of the Guidelines for Validation of NGS-Based Oncology Panels provides consensus recommendations on validation and ongoing monitoring of targeted NGS panels in the clinical setting and covers a broad spectrum of topics, including NGS platform overview, test design, potential sources of error during NGS assay development process, optimal number of samples for validation, establishing minimal depth of sequencing, implementation and quality control metrics, and others.
How much of the paper is meant as a correction to practices that don’t represent best practices?
AMP has consistently been on the forefront of shepherding new genomic technologies and methodologies into clinical practice through the development of best practice guidelines. This latest manuscript is not intended to correct specific deficiencies but to inform and assist the clinical laboratory community and stakeholders by providing subject matter expert-derived consensus recommendations on validation and ongoing monitoring of clinical NGS testing for detection of somatic variants. This new document summarizes current knowledge about targeted NGS in the field of molecular diagnostics, exposes challenges of this technology, and provides guidance on how to ensure high-quality sequencing when used for patient care. Specifically, the recommendations emphasize the critical role of laboratory director in using an approach that identifies potential sources of errors that may occur throughout the analytical process and addressing these potential errors through test design, method validation, or quality controls so that patients do not suffer as a result of subpar quality testing.
What’s the most “important” section of the paper? In which area are established best practices most needed?
Frankly, it is all quite important. All aspects of NGS testing – from initial panel design to interpretation and reporting of variants – requires not only laboratory technical skills but the experience, judgment, and expertise of the molecular professional who is ultimately responsible for and intimately involved in producing accurate patient results. A rigorous validation process establishes the performance characteristics of the test within the laboratory, enabling laboratory directors to establish quality metrics that ensure the test remains robust and accurate once in clinical use. This paper outlines current best practices for laboratory directors utilizing an error-based approach for patient risk management.
For example, the most common sticking point in evaluations of NGS tests is the number of samples a validation effort must invoke in order to provide confidence in that test and previously available peer-reviewed publications offered no fixed number. This manuscript addresses the volume of samples question specifically by providing calculations depending on how much confidence is needed. No medical test is ever 100% sensitive and 100% specific, but there are several ways to establish confidence about the results of an NGS test, and the goal is always to ensure that any errors will be readily detected and that patients do not suffer as a result of subpar quality testing.
As AMP surveyed all the data sources for the paper, what was the biggest surprise either in lab practice or a lack of knowledge?
It was not surprising to find that there was some variability in practice across the molecular laboratory community as this initially disruptive technology was introduced into clinical practice. This is somewhat expected when a new technology enters any area of medicine. Consensus best practices begin to emerge over time and with additional experience. Given our review of survey data and published literature, along with many informative discussions with colleagues about how they were implementing NGS in their own facilities, AMP’s Clinical Practice Committee believed that providing a set of minimum best practice standards for NGS validation for oncology panels was now possible. Whether laboratory directors are bringing this testing into their laboratory for the first time or have been performing this testing for years, we believe that this manuscript offers information that can help those professionals ensure highly accurate NGS testing continues to be provided to patients.
How often does AMP plan to update the paper?
Cancer genomics is a rapidly evolving field, so our recommendations on how to validate and monitor targeted NGS panels should be reevaluated on an ongoing basis. Many of the best practice principles articulated in this manuscript will stay the same over time, although this set of guidelines addresses specifically the DNA panel testing for platforms and chemistries that are currently available. AMP will continue to monitor the need for an update through existing Clinical Practice Committee oversight processes.