Determining Microsatellite Instability Status Using Multiplex PCR-Based Methods

Determining Microsatellite Instability Status Using Multiplex PCR-Based Methods

Sponsored Content by Promega Corporation Jan 24 2020 insights from industry Shaun Peterson Global Strategic Manager, Clinical Diagnostics Business Promega Corporation An interview with Shaun Peterson, Global Strategic Manager, Clinical Diagnostics Business, Promega Corporation Please give an overview of microsatellite instability (MSI) status and its applications within oncology. Historically Microsatellite Instability (MSI) has been used to screen for Lynch Syndrome, a dominant hereditary cancer propensity. Recently, MSI status has been rediscovered as a biomarker for prolonged and durable response to immune checkpoint inhibitors, making MSI status an increasingly relevant tool in genetic- and immuno-oncology. Visit the MSI Educational Resources MSI testing functionally measures the genomic accumulation of insertion or deletion (INDEL) errors caused by cells deficient mismatch-repair system (dMMR) that occurs in certain types of solid tumors, and this screening may be used to better characterize tumors and guide therapeutic choices for MSI-High cancer types. Read the Full Article from Promega's Lead R&D Scientist To learn more about both the classical and new utility of microsatellite instability please visit our educational resource. Also, to learn more about this history of MSI please read this article from our lead R&D scientist: What recent developments have been made that make the MSI status of tumor tissue important? May 23, 2017, the U.S. Food and Drug Administration granted accelerated approval to pembrolizumab for adult and pediatric patients with unresectable or metastatic, microsatellite instability-high (MSI-H) or mismatch repair deficient (dMMR) solid tumors. This is the first time the agency has granted approval for a cancer therapeutic agnostic to tissue/site of origin indication. Additionally, on July 31, 2017, the U.S. FDA granted accelerated approval to nivolumab for the treatment of patients 12 years and older with mismatch repair deficient (dMMR) and microsatellite instability high (MSI-H) metastatic colorectal cancer. Since then the agency has made additional approvals for MSI-H tumors. Tumors with MSI-High status have been shown to respond to immune checkpoint inhibitor (ICI) therapies. This outcome may be explained by MSI-driven tumor expression of mutation-associated neoantigens (MANA) that are believed to cause immune cell infiltration into the tumor microenvironment. Tumor induced inhibition of immune cell activity can be overcome with ICI therapies, allowing for tumor cell destruction by the immune cells. What techniques can be used to establish MSI status? What are the advantages and limitations of these methods? The current Promega Research Use Only MSI assay, is a PCR and Capillary Electrophoresis-based test that is designed to match the robustness needed when working with fragmented DNA extracted from FFPE samples. Image credit: Promega The assay is easy to use and has been available and used in the market as part of Lab-Developed Tests since 2004. This patent-protected technology is considered the gold standard molecular assay for detecting DNA mismatch-repair deficiency. Promega intends to seek US FDA approval and CE-IVD marking for its MSI assay to help oncologists and pathologists make treatment decisions for colorectal cancer patients. The MSI Assay is reimbursable, has a fast turnaround time and most importantly there is a large body of evidence supporting use of MSI in colorectal cancer decisions. Immunohistochemistry (IHC) analysis of the presence or absence of mismatch repair proteins (MMR) is often considered a surrogate and equal analysis method, but peer reviewed literature demonstrates IHC-MMR is not an equal comparison to MSI by PCR. The presence of MMR protein expression is not necessarily a conclusive measure of MMR function. There can be a loss in the function of these proteins without a corresponding loss of the protein in the cell. There can be a loss in the function of these proteins without a corresponding loss of the protein in the cell. It has been estimated that 5-12% of MSI-H tumors are not recognized by an IHC test in part because of the expression of non-functional MMR proteins. MSI by PCR directly measures changes in DNA caused by loss of MMR protein function as opposed to measuring the proteins themselves as in IHC. This makes MSI by PCR a functional measure of mismatch repair deficiency that detects loss in MMR repair function, regardless of the origin. While there are large NGS cancer gene panels used by specialty service providers that will provide an estimate of the MSI status of a tumor and other signatures of genomic instability, such as tumor mutation burden. However, each laboratory attempts to analyze different loci for MSI and the MSI determination is made using different bioinformatic algorithms. Thus, one should be careful in selecting the laboratory. These types of assays are often quite costly and the long turnaround time argues against using them for MSI determination unless there are other reasons to wait for the results with regard to other cancer markers covered by the panel. What does Promega offer to determine the MSI status of tissues? What is the workflow of this technique? The MSI Analysis System, Version 1.2 (RUO), is a fluorescent multiplex PCR-based method for detecting microsatellite instability (MSI). Microsatellites are loci or regions of DNA where one or a few bases are tandemly repeated many times. MSI is a form of genomic instability caused by the insertion or deletion of repeat elements at these loci during DNA replication and the failure of the mismatch repair system (MMR) to correct these errors. This system has been the gold standard test available for clinical research since 2004. Following DNA isolation, specific mononucleotide sequences within the samples are amplified using polymerase chain reaction performed on a thermocycler. Microsatellite markers can be amplified in multiple, parallel reactions or multiplexed in a single reaction. When multiplexed primers for individual markers are fluorescently labeled, many different fragments of similar sizes can be detected in the same reaction. Related Stories



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