Microsatellite Instability (MSI)

In this article, learn more about microsatellite instability (MSI) and its association with various cancers including colon, gastric, and ovarian cancers. Discover how next-generation sequencing (NGS) can be used to measure this form of tumor mutation load and its advantages over other detection methods.

Microsatellites are tandem repeats of DNA, where a sequence motif of one to six nucleotides is repeated multiple times. Also called short tandem repeats (STRs), these repetitive sequences can occur in thousands of regions and have a higher mutation rate than other areas of the human genome (1). Thus, these regions have higher genetic diversity and STR polymorphisms have been used for human identification and population genetics.

As the name suggests, DNA mismatch repair (MMR) is the process whereby errors, such as insertions and deletions, that occur during DNA replication are recognized and fixed. Microsatellite instability (MSI) occurs due to MMR not functioning properly, with the target microsatellite gaining or losing repeat units, resulting in a somatic change in length.

In the 1990s, MSI was discovered to be a biomarker for hereditary nonpolyposis colorectal cancer (HNPCC), also known as Lynch syndrome (2). The widespread instability associated with deficient MMR indicates a rapid accumulation of somatic mutations that could inactivate genes in key regulatory processes and cause tumorigenesis. Current research has identified MMR deficiency in many forms of cancer, more often in early-stage disease (3) (Figure 1).

Analyzing MSI is becoming an increasingly important tool in cancer research and immuno-oncology. Given the prevalence of MSI in colorectal cancer and the need to better understand both clinical and histological manifestations, recommended MSI testing criteria were established by key leaders in the field (4). Known as the revised Bethesda guidelines, these recommendations established microsatellite targets in the testing panel (e.g., BAT25, BAT26) and defined how one would classify a cancer tumor as microsatellite stable (MSS), MSI-low (MSI-L), and MSI-high (MSI-H) based on the number of biomarkers found with a mutation. The Bethesda guidelines and MSI testing have helped with patient prognosis, guiding therapy and serving as a screen for HNPCC.

Studies have shown that colorectal cancers with MMR deficiency (MSI-H) are sensitive to immune checkpoint blockade with antibodies to programmed death receptor-1 (PD-1) (5,6). MMR-deficient colorectal cancer is hypothesized to have a larger proportion of neoantigens that would make it more sensitive to immunotherapy. The latest research data supports this hypothesis, and MMR deficient cancers are sensitive to immune checkpoint inhibitors, regardless of cancer type (3).

As a result of MSI studies in cancer research, the U.S. Food and Drug Administration (FDA) has approved a PD-1 inhibitor for the treatment of adult and pediatric patients with MSI-H or MMR solid tumors, regardless of tissue type. The FDA also approved the companion MSI biomarker assay, which defines the group of patients that can be treated (7,8).

If you would like to learn more about immuno-oncology and the impact of NGS in this field, explore our immuno-oncology learning center articles.

The aforementioned approval of a PD-1 inhibitor for patients with MSI-H or MMR solid tumors is the first tissue-agnostic drug approval by the FDA and the first approved companion assay for any cancer therapy (8). Established MSI tests following Bethesda guidelines typically use capillary electrophoresis and analysis with paired normal-tumor samples to perform a qualitative assessment of MSI for each biomarker.

NGS and the ability to sequence millions of DNA molecules rapidly in a parallel fashion has transformed molecular genetics. It has also enabled precision medicine and offers many advantages for MSI analysis in both the research and healthcare setting. The high sequencing throughput of NGS means that high multiplexing of samples is now possible, as well as the capability to significantly increase the number of microsatellites in a single assay.

Only five microsatellite markers are recommended by the Bethesda guidelines. Given this limitation, greater stringency is required to ensure that even one marker does not fail, which would impact panel performance. By significantly increasing the number of microsatellites analyzed in a panel, NGS can provide greater reliability and reproducibility with MSI analysis. A more accurate assessment of MSI-H and MSS can also be achieved with NGS. MSI-H refers to changes in two or more of the five recommended markers in the guidelines. Low MSI is defined by have only one change out of the five markers. MSI analysis with NGS can provide a more quantitative and granular assessment of what MSI-H or MSI-L is. For example, a scoring system can be implemented to indicate the degree of MSI-H.

Many MSI markers are now reported in different publications. By including these markers, NGS can achieve greater sensitivity and can better capture information on MMR deficiency across multiple cancer types.  Increasing the number of molecular MSI targets would provide a tumor assay that is truly agnostic to tissue type. Researchers and clinicians could improve patient stratification based on MSI scoring, enabling improved clinical research, as well as potentially improving patient outcome.

1. Brinkmann B, et al. Am J Hum Genet 62:1408 (1998)
2. De la Chapelle A. New Engl J Med 349:290 (2003)
3. Le DT, et al. Science 357:409 (2017)
4. Umar A, et al. J Natl Cancer Inst 18:261 (2004)
5. Le DT, et al. New Engl J Med 372:25090 (2015)
6. Overman MJ, et al. J Clin Onc 35:519 (2017)
7. Lemery S, et al. New Engl J Med 377:1409 (2017)
8. Chan TA, et al. Ann Oncol 30:44 (2019)

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