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HOME > Cancer Res Treat > Volume 58(1); 2026 > Article
Review Article Diagnostic Challenges and Clinical Implications of Microsatellite Instability/Mismatch Repair Deficiency in Solid Tumors
Yoonjin Kwak1orcid, Jeong Mo Bae1,2orcid, Hye Seung Lee1,2orcid
Cancer Research and Treatment : Official Journal of Korean Cancer Association 2026;58(1):1-14.
DOI: https://doi.org/10.4143/crt.2025.1161
Published online: December 26, 2025

1Department of Pathology, Seoul National University Hospital, Seoul National University College of Medicine, Seoul, Korea

2Cancer Research Institute, Seoul National University College of Medicine, Seoul, Korea

Correspondence: Hye Seung Lee, Department of Pathology, Seoul National University Hospital, Seoul National University College of Medicine, 103 Daehak-ro, Jongno-gu, Seoul 03080, Korea
Tel: 82-2-740-8269 E-mail: hye2@snu.ac.kr
*Yoonjin Kwak and Jeong Mo Bae contributed equally to this work.
• Received: October 23, 2025   • Accepted: December 26, 2025

Copyright © 2026 by the Korean Cancer Association

This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/4.0/) which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited.

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  • The mismatch repair (MMR) system plays a crucial role in correcting replication errors; when the MMR system is deficient (dMMR), replication errors (particularly within microsatellites) accumulate throughout the genome, leading to microsatellite instability (MSI). The key MMR genes include MLH1, MSH2, MSH6, and PMS2. Germline mutations in these genes are associated with Lynch syndrome, whereas in sporadic solid tumors, dMMR is often caused by hypermethylation of the MLH1 promoter. As the clinical use of dMMR/MSI expands, the importance of reliable testing for dMMR or MSI in companion diagnostics continues to increase. dMMR/MSI is diagnosed using immunohistochemistry (IHC) for MMR proteins, polymerase chain reaction with fragmentation analysis, or next-generation sequencing. Although IHC has technical limitations, it requires less tissue, has a short processing time, and is cost-effective. Experienced or specialized pathologists and educational efforts are helpful for reliable diagnosis, in addition to the technical aspects. Solid tumors with dMMR/MSI exhibit distinct clinicopathological features, including prognostic significance and a predictive role in adjuvant cytotoxic chemotherapy. Solid tumors with dMMR/MSI are also characterized by a higher tumor mutational burden and abundant immune cell infiltration, making them promising candidates for immune checkpoint inhibitor therapy. However, the oncogenic processes and immune microenvironment are not identical across the organs of origin, between patients, and even within the same patient, which should be considered in future studies. This review provides an overview of the practical aspects of dMMR/MSI testing, along with the molecular mechanisms and immune microenvironments associated with dMMR/MSI solid tumors.
  • Key words: Microsatellite instability, DNA mismatch repair, Companion diagnostics, Immune microenvironment
Microsatellites are short tandemly repeated nucleotide sequences that are widely distributed throughout the genome. They typically consist of 5-50 times repeats of 1 to 6-10 nucleotides [1]. As an example of mononucleotide repeats, BAT26 is a poly (A)26 tract (26 repeats of A) localized in the fifth intron of MSH2, and BAT25 is a poly (A)25 tract localized in intron 16 of c-kit. There are 50,000-100,000 dinucleotide repeats in the human genome. For example, D5S346 is a repeat of CA ([CA]n). Microsatellites have been found predominantly in noncoding DNA within introns or intergenic regions of the genome, which is related to that exome accounts for only 1%-2% of the entire genome [2,3].
Spontaneous mutations are frequently found during DNA replication, and mismatch repair (MMR) systems correct the replication errors that occur in microsatellites [4]. Uncorrected DNA replication errors lead to changes in the microsatellite length. The four key MMR genes are MLH1, PMS2, MSH2, and MSH6, and the minor MMR genes include MLH3, MSH3, and PMS1. MLH1 and PMS2 form a functional heterodimer, as do MSH2 and MSH6. Germline mutations in MMR genes or epithelial cell adhesion molecule (EPCAM) deletion cause Lynch syndrome, and MLH1 hypermethylation is associated with sporadic deficient MMR (dMMR) or microsatellite instability–high (MSI-H) cancers.
This review aims to provide an overview of the practical aspects of MMR/microsatellite instability (MSI) testing, along with the molecular mechanisms and immune microenvironments associated with dMMR/MSI-H solid tumors.
dMMR results in alterations in the length of microsatellite repeats, which induce MSI-H. In addition to noncoding DNA, frameshift mutations in the coding regions of tumor suppressor genes (TSGs) occur in association with dMMR/MSI-H, and functional inactivation of TSGs is related to tumorigenesis. Several coding-region targets containing microsatellites, including TGFβRII, BAX, TCF4, MSH3, ACVR2, PTHL3, HT001, AC1, SLC23A1 MACS, NUDFC2, and TAF1B, have been reported. TGFβRII is one of the most frequently altered genes, and in our previous study, its frameshift mutation was observed in approximately 90% of MSI-H gastric cancer (GC) [5]. The genes most frequently targeted by MSI vary across tumor types. For example, TGFβRII, BAX, and ACVR2 are specific to colorectal cancer (CRC) and GC, whereas JAK1, TFAM, PDS5B, and ESRP1 are specific to endometrial cancer (EC) [6-8].
TGFβRII gene has poly (A)10 tract. Loss of functional transforming growth factor beta receptor II (TGFβRII) protein caused by the frameshift mutation of TGFβRII inactivates the canonical transforming growth factor beta (TGFβ) signaling pathway. In the canonical pathway, TGFβ binds to TGFβRII, activates TGFβRI, and then phosphorylates Smad proteins [9]. Smad proteins are involved in the regulation of gene expression. In early carcinogenesis, through the canonical pathway, TGFβ signaling inhibits cell growth and induces apoptosis, suggesting its roles as a tumor suppressor. However, as cancer progresses, TGFβ signaling switches to promote tumor progression, invasion, metastasis, fibrosis, and immune evasion [10], which is related to noncanonical TGFβ signaling pathways with the activation of the MEK/extracellular signal-regulated kinase (ERK), phosphoinositide 3-kinase (PI3K)/Akt, or Rho-like GTPase pathways. These opposing roles of TGFβ signaling are important to understand the biology of dMMR/MSI-H tumors [11].
RNF43 is another frequently mutated gene in dMMR/MSI-H tumors. The G659fs frameshift at a mononucleotide tract (c.1970_1976 G7) in the C-terminal region is the most common in MSI-H CRCs (approximately 80%), and approximately half of the MSI-H GC and EC harbor RNF43 mutations [12-14]. RNF43 is a transmembrane E3 ubiquitin ligase that negatively regulates the Wnt signaling pathway by ubiquitinating Frizzled (FZD) receptors, leading to endocytosis and degradation [15,16]. It functions as a tumor suppressor, and its loss promotes epithelial stemness and proliferation. Although these alterations are not considered classical driver mutations, functional studies using CRISPR/Cas9 have demonstrated that C-terminal truncations attenuate RNF43 signaling and reduce surface FZD expression, resulting in the loss of regulatory control over Wnt activity [17,18]. Clinically, RNF43 mutations have been associated with improved outcomes and responses to programmed death-1 (PD-1)/programmed death-ligand 1 (PD-L1) blockade in several retrospective studies [19-21]. However, whether RNF43 adds predictive value beyond MSI/MMR status itself requires more sophisticated prospective validation.
Earlier studies have aimed to reveal the mechanism of action and functional roles of dMMR/MSI-H in carcinogenesis and cancer progression. To screen for Lynch syndrome and predict drug responses, MMR/MSI tests have recently been used as in vitro diagnostics (IVD) or companion diagnostics (CDx) in daily practice, especially for patients with CRC, GC, and EC (Fig. 1). CDx are medical devices, often IVD, that provide essential information for the safe and effective use of a corresponding drug or biological product [22]. Therefore, CDx should be validated both analytically and clinically. The MSI/MMR status can be examined by polymerase chain reaction (PCR) and fragmentation analysis, MMR protein immunohistochemistry (IHC), or next-generation sequencing (NGS). The U.S. Food and Drug Administration (U.S. FDA) has approved FoundationOne CDx (Foundation Medicine) and MI Cancer Seek (MCS) (Caris Life Sciences) for MSI-H and the Ventana MMR RxDx Panel (Ventana Medical Systems) for MLH1, MSH2, MSH6, and PMS2 as CDx [12].
1. IHC for four MMR proteins
dMMR can be diagnosed with IHC of four MMR proteins, including MLH1, MSH2, MSH6, and PMS2. MMR proteins are expressed in the nuclei of normal cells and are lost owing to biallelic genetic inactivation. Concurrent loss of MLH1 and PMS2 is observed in MLH1-mutated or MLH1-methylated tumors. PMS2 forms a functional heterodimer with MLH1, and PMS2 is dependent on MLH1 for stability. Similarly, in MSH2-mutated tumors, concurrent loss of MSH2 and MSH6 expression is observed, although isolated loss of MSH6 or PMS2 expression can occur in MSH6- or PMS2-mutated tumors. Considering these functional heterodimers, two antibody testing algorithms (MSH6 and PMS2) have been proposed for Lynch syndrome screening [23]. Loss of expression is rarely observed in additional MMR proteins beyond the expected heterodimer partner. For example, loss of MSH6 expression is rarely observed in MLH1/PMS2 or PMS2-deficient cancers, which is due to a secondary mutation in the coding mononucleotide tract or an independent somatic mutation in MSH6 [24-26]. Accurate interpretation in routine practice requires a review by specialized pathologists and must be accompanied by dedicated training sessions because atypical expression patterns and interpretive pitfalls are common [27].
MMR IHC should begin by confirming the nuclear staining of adjacent normal cells as an internal positive control. With that control established, tumors are defined as proficient MMR (pMMR) if the expression of all four MMR proteins is retained, whereas dMMR is defined as the absence of expression of one or more MMR proteins. Earlier studies have reported indeterminate MMR IHC results, most often when the internal positive control shows equivocal or faint nuclear staining [28]. Therefore, panels designed to ensure strong internal control staining have been adopted in clinical practice. Many pathology laboratories now use the U.S. FDA–approved Ventana MMR RxDx Panel, which typically shows strong staining intensity with robust positivity in internal controls. A known issue with the RxDx test is that, in some MLH1-deficient cases, abnormal punctate nucleolar staining may appear on MLH1 IHC (Fig. 2) [29,30]. In such cases, performing IHC for PMS2 or using an alternative MLH1 antibody can help clarify MMR status. In addition, missense or in-frame mutations in MMR genes may produce inactive MMR proteins that can bind to primary antibodies and cause false-positive staining [3].
Although MMR IHC is subject to certain technical limitations, it remains advantageous owing to its minimal tissue requirements, cost-effectiveness, rapid turnaround time, applicability in specimens with low tumor purity, and the ability to directly correlate with histopathological findings on hematoxylin and eosin slides [31].
2. PCR and fragmentation analysis for MSI testing
Traditional MSI testing involves PCR and fragmentation analysis using several microsatellite markers [32]. As several markers have been reported to well indicate the MSI status, the National Cancer Institute/Bethesda panel selected five markers with two mononucleotide markers (BAT25 and BAT26) and three dinucleotide markers (D5S346, D2S123, and D17S250) [33]. To overcome the relatively low sensitivity of dinucleotide markers, Suraweera et al. [34] proposed and validated a pentaplex panel of quasimonomorphic mononucleotide markers (NR-27, NR-21, NR-24, BAT-25, and BAT-26). The multiplex fluorescent MSI analysis system of Promega has been developed, using the pentaplex panel with the two additional pentanucleotide repeats of Penta C and D for quality control [35]. MSI testing requires paired tumor and normal DNA. If the tumor DNA test results show additional bands compared with the normal DNA results, they are considered unstable. MSI-H is defined as two or more markers being unstable, MSI-low (MSI-L) as one marker being unstable, and microsatellite-stable (MSS) as all five markers being stable.
MSI testing using the above PCR method evaluates five microsatellite markers; the procedures are relatively simple, and MSI results can be obtained within a few days [36]. In addition, multiplex PCR and fragmentation analysis with 2-5 markers are possible using either Bethesda or pentaplex panels. However, MSI testing based on PCR and fragmentation analysis has several disadvantages. It requires fragmentation analysis equipment, > 20% tumor cell purity through macroscopic dissection, and large volumes of tissue. DNA extracted from old paraffin blocks can exhibit multiple noise bands, and inexperienced interpreters can overlook subtle shifts, resulting in false-negative results.
These conventional MSI markers have been developed and validated for CRC and Lynch syndrome, and it might be difficult to interpret MSI status because of subtle shifts, especially in non-CRC [37]. The shift in the length of each microsatellite marker for MSI detection is significantly shorter in EC than in CRC and GC [38,39]. Long mononucleotide repeat, with 52-60 repeats of A ([A]52-60), has a higher sensitivity for MSI testing than Bethesda or pentaplex panels [38]. In addition, several PCR-based MSI testing techniques have been reported [3]. Drop-off droplet digital PCR (ddPCR) assays targeting BAT-26, ACVR2A, and DEFB105A/B microsatellite markers have been suggested, particularly for liquid biopsy [40]. High-resolution melting and denaturing high-performance liquid chromatography have been applied for post-amplification microsatellite amplicon analysis during MSI testing, and real-time PCR techniques using peptide nucleic acid probes have been investigated [3]. However, the diagnostic sensitivity and reliability of these methods have not been confirmed.
3. Next-generation sequencing
NGS-based comprehensive genomic profiling (CGP) has been rapidly integrated into clinical practice over the past decade [41,42]. CGP testing enables the simultaneous analysis of hundreds of genetic alterations in a cost-effective manner and is highly effective in identifying actionable targets for precision therapies. An additional advantage of using CGP for MSI analysis is its ability to evaluate both somatic mutations and MSI status simultaneously, allowing MSI assessment even in tumor types where MSI testing is not routinely performed, beyond CRC and EC [43].
Bioinformatic algorithms for analyzing MSI using NGS data can be categorized into two groups: microsatellite locus- and mutation signature-based methods. Locus-based methods, such as mSINGS, MSI sensors, and MANTIS, identify microsatellite loci from NGS data and evaluate changes in repeat length relative to normal references [44-46]. Although this approach achieves the highest accuracy when matched normal NGS data are available, it can also be applied to tumor-only samples by comparison with the baseline reference datasets. Mutation signature-based methods, such as MSIseq, analyze the mutational patterns induced by MSI, including mutation counts, indel burden, ratio of indels:substitutions, indels in homopolymers or repeat tracts, and trinucleotide context/substitution spectrum (SBS signatures), to train predictive classification models [47,48].
NGS-based MSI testing has been reported to accurately determine MSI status. The overall percent agreement values of NGS-based MSI testing in FoundationOne CDx are 97.7% and 97.8% for PCR-based MSI testing and IHC, respectively [49]. However, the diagnostic performance of NGS-based MSI testing varies according to tumor type [43]. In CRC and EC, NGS-based MSI detection demonstrated 100% sensitivity and 99.3% specificity compared with conventional PCR-based MSI assays, whereas in other cancer types, the sensitivity and specificity were 96.6% and 100%, respectively.
However, the performance of NGS-based MSI testing can be influenced by multiple technological and biological factors, including tumor purity, platform-specific error profiles, and analytical algorithms. The accuracy of NGS-based MSI testing can be decreased when tumor purity falls below 10%-20% [50]. Platform-specific error signatures introduce additional complexity. Semiconductor-based sequencers generate higher indel error rates in homopolymers, potentially inflating instability estimates, whereas sequencing-by-synthesis platforms may under-detect small indels in long mononucleotide runs unless specialized callers are applied [51,52]. Benchmark evaluations of various NGS-based MSI detection algorithms have demonstrated that their analytical performance can differ substantially across methods [53].
The College of American Pathologists (CAP) recommends the use of NGS-based MSI analysis algorithms that have been analytically validated through comparison with PCR-based MSI tests or IHC [54]. For CRC, the CAP further supports NGS-based MSI testing as an alternative to conventional PCR or IHC methods. The U.S. FDA has approved the FoundationOne CDx assay as a CDx test to identify patients with MSI-H tumors who may benefit from pembrolizumab treatment [22].
4. Circulating tumor DNA
Liquid biopsy is a minimally invasive approach that detects genetic alterations in circulating tumor DNA (ctDNA) in the plasma [55]. It provides valuable molecular information in clinical situations in which tissue biopsy is not feasible, radiologically measurable residual disease is not present, or intratumoral heterogeneity limits the representativeness of tissue specimens. Recent guidelines from the European Society for Medical Oncology (ESMO) recommend performing MSI testing using ctDNA when tumor tissue is unavailable [56].
Although both ddPCR and NGS have been reported to robustly detect MSI in ctDNA, NGS-based ctDNA testing is generally preferred in clinical practice because it allows the simultaneous detection of multiple genomic alterations [40,57,58]. The accuracy of MSI detection in NGS-based ctDNA assays is affected by both the amount of input DNA and the fraction of tumor-derived DNA within the sample. For the Guardant360 assay, the limit of detection was 0.1% with 30 ng of input DNA, whereas it increased to 0.4% when only 5 ng was used. The sensitivity of MSI detection was 93.1% when the maximum variant allele frequency (maxVAF) was ≥ 1% but decreased to 86% at a maxVAF threshold of ≥ 0.2% [59]. In the case of FoundationOne Liquid CDx, the sensitivity for MSI detection was 71.4% when the tumor fraction was ≥ 1%, but markedly decreased to 6% when the tumor fraction was < 1% [60].
Nearly all clinical trials seeking to demonstrate the efficacy of immune checkpoint inhibitors (ICIs) in MSI-H tumors have assessed MSI status using tumor tissue [61-63]. In addition, to date, no ctDNA-based MSI assay has been approved as CDx by the U.S. FDA. Consequently, the predictive value of ctDNA test-based MSI assays for ICI treatment should be evaluated on the basis of real-world evidence. A retrospective study reported an overall response rate of 77% among nine patients with pancreatic cancer characterized as plasma MSI-H [64]. In a pan-cancer retrospective study, among patients identified as MSI-H by the Guardant360 assay, treatment with immuno-oncology (IO) agents was associated with a significantly longer real-world time to treatment discontinuation than non-IO treatment across all tumor types, except non–small cell lung cancer [65].
5. Practical consideration of MMR/MSI testing
The CAP, Association of Molecular Pathology, American Society of Clinical Oncology, and Fight Colorectal Cancer report recommendations for dMMR/MSI-H detection in patients with solid tumor malignancies being considered for ICI therapy [54]. MMR IHC and/or MSI by PCR is preferred for patients with CRC, and validated NGS assay can also be used. They also recommended MMR IHC and/or MSI by PCR testing for patients with GC rather than NGS. For patients with EC, MMR IHC is recommended over MSI using PCR or NGS. They also recommended that tumor mutational burden (TMB) should not be used as a surrogate marker for dMMR detection. If TMB-high is identified, IHC and/or MSI by PCR should be performed.
With recent advances in cancer treatment, the number of essential biomarkers has increased. Beyond MMR/MSI testing, CRC also requires the assessment of RAS mutations, BRAF mutation, and human epidermal growth factor receptor 2 (HER2) status, whereas GC requires HER2 and CLDN18.2, and PD-L1 should be evaluated with multiple clones (22C3, 28-8, and SP263). As shown in Table 1, when surgically resected tissue is available, the amount of tissue is generally not a limiting factor. However, for many patients who initially present with stage IV CRC or GC, only small endoscopic biopsy specimens are available. For endoscopic biopsy specimen, MMR IHC typically requires four unstained slides with a 4-μm thickness. In contrast, MSI test requires 8-10 unstained slides with a thickness of 10 μm, and NGS often requires 10-20 unstained slides. Therefore, it is important to plan the number of tests required for biomarker testing, tissues that can be used, and methods that have been validated or approved. In addition to diagnostic reliability and validity, the amount and quality of available tissue should also be considered in daily practice.
The concordance rates between dMMR and MSI-H status are approximately 98% in CRC [66] and GC [28] and approximately 94% in EC [67]. The overall prevalence of dMMR or MSI-H is approximately 5%-25% in CRC, GC, and EC (Table 2) [5,50,68-83]; thus, the rate of discordance between dMMR and MSI-H can be 4-20 times higher among cases exhibiting dMMR or MSI-H. For example, in our previous study, MSI-H or dMMR was observed in 9.7% (549 of 5,676) of GC cases, and 47 indeterminate or discordant cases (0.8% of the total, but 8.6% of MSI-H or dMMR cases) were subsequently confirmed as MSI-H or dMMR upon reevaluation [28]. When IHC was repeated or the slides were reviewed by experienced pathologists, some discordant cases were reclassified as concordant [28,66]. In addition to the technical limitations of testing methods and potential misinterpretation by pathologists, factors such as the subclonal loss of MMR proteins, lower tumor purity, and missense mutations may also contribute to the discordance between dMMR and MSIH results.
MSI is important for four key aspects: screening for Lynch syndrome, as a distinct molecular subtype, as a poor predictive biomarker for adjuvant chemotherapy, and as a good candidate for immunotherapy.
1. Screening Lynch syndrome
Hereditary nonpolyposis CRC is diagnosed by clinical criteria, and the diagnosis of Lynch syndrome needs presence of pathogenic germline mutations in MMR genes or deletion of 3′ exons of the EPCAM gene. In patients with Lynch syndrome, 10%-80% are at risk for CRC, 15%-71% for EC, 1%-13% for GC, 4%-20% for ovarian cancer, and 1%-12% for small bowel cancer. The estimated average age of cancer presentation in Lynch syndrome is lower than that of sporadic cancer: 44 years for CRC, 49 years for EC, 52 years for GC, and 47 years for small bowel cancer. Therefore, MMR IHC or MSI testing is required for patients with tumors in the Lynch syndrome spectrum, especially those with a family history of Lynch syndrome or tumors that develop at a younger age.
2. Molecular subtype with distinctive characteristics
Table 2 summarizes the prevalence and clinical significance of dMMR/MSI-H tumors in the individual organs. The dMMR/MSI-H subtype is more common in Lynch syndrome-related cancers and has been reported in approximately 10%-20% of CRCs, 16%-26% of ECs, 8%-22% of GCs, and 10%-26% of small bowel cancer [68-76]. However, it is rarely observed in other tumors, such as prostate adenocarcinoma (1%-3%), non–small cell lung cancer (< 1%), breast ductal carcinoma (< 1%), and glioblastoma (< 1%) [50,79,81-83]. In addition, dMMR/MSI-H tumors are one of the major molecular subtypes of some cancers according to The Cancer Genome Atlas, including CRC [84], GC [14], and EC [85].
In CRC, dMMR/MSI-H defines a distinct clinicopathological subtype with right-sided predominance, poor or medullary differentiation, mucin production, a marked immune reaction with tumor-infiltrating lymphocytes, and a Crohn-like reaction [68]. This subtype aligns with the consensus molecular subtype, CMS1 (“MSI immune”), characterized by hypermutation and strong immune activation [86]. Sporadic MSI-H cases most commonly arise via MLH1 promoter hypermethylation and frequently harbor BRAF V600E, whereas Lynch-associated MSI-H tumors occur at a younger age and rarely carry BRAF mutations [87]. Biologically, the hypermutated immunogenic phenotype is associated with fewer positive lymph nodes at presentation and a favorable stage-adjusted prognosis in nonmetastatic disease [88].
MSI-H subtype also exhibits the distinctive features in GC. They are associated with older age, distal tumor location, elevated or localized gross type, intestinal type by the Lauren classification, expansile growth by the Ming classification, and the absence of venous or perineural invasion [5]. In early-stage GC, MSI-H tumors tend to show a relatively high proportion of lymphatic invasion and occult nodal metastasis, as demonstrated by cytokeratin IHC in patients initially staged as node-negative. By contrast, in advanced GC, MSI-H tumors display fewer lymph node metastases than other molecular subtypes, and this feature is associated with a more favorable prognosis [74].
MSI-H EC is predominantly endometrioid and shows a high somatic mutation burden with a conspicuous tumor-infiltrating lymphocyte response [85]. A low histologic grade is less common in this subtype than in MSS tumors [85]. Most MSI-H ECs are sporadic and arise through MLH1 promoter hypermethylation, whereas a smaller proportion are associated with Lynch syndrome [72]. At the molecular level, MSI-H EC shows frequent alterations in the PI3K pathway, with PTEN mutations most common, followed by PIK3CA and PIK3R1. PIK3CA and PIK3R1 alterations are generally mutually exclusive, yet either one often co-occurs with a PTEN mutation [7,72]. Dedifferentiated and undifferentiated ECs are often dMMR or MSI-H and commonly show abnormalities in the SWI/SNF complex [89].
3. Predictive biomarker for adjuvant therapy
The dMMR/MSI-H status is a crucial biomarker for guiding adjuvant treatment, particularly in stage II CRC. Pooled analyses of randomized clinical trials have indicated that patients with dMMR/MSI-H stage II CRC have minimal to no benefit from fluorouracil-based monotherapy, whereas those with pMMR/MSS tumors demonstrate a favorable response [90-92]. In contrast, recent studies have shown that fluoropyrimidine monotherapy provides little or no benefit in stage III disease, whereas the addition of oxaliplatin improves disease-free outcomes [93]. Similarly, in stage II-III GC, retrospective cohorts and post hoc analyses of CLASSIC trials have indicated that adjuvant chemotherapy improves outcomes in MSS disease after D2 resection, whereas patients with MSI-H tumors do not show a clear benefit from conventional fluoropyrimidine-based or capecitabine plus oxaliplatin regimens [94,95]. However, another study reported opposite findings, showing that MSI-H patients receiving adjuvant chemotherapy demonstrated improved disease-free survival and overall survival relative to patients with MSS tumors [96]. Based on these studies, the most recent ESMO guidelines recommend considering adjuvant chemotherapy carefully in resected MSI-H GC [97]. In EC, molecularly stratified results from the PORTEC 3 trial showed that the dMMR subgroup did not gain a relevant benefit from combined chemoradiation compared with radiotherapy alone, in contrast to the p53 abnormal subgroup, where the benefit was concentrated [98]. The recent European Society of Gynaecological Oncology (ESGO)–European Society for Radiotherapy and Oncology (ESTRO)–European Society of Pathology (ESP) guidelines recommend administering adjuvant treatment based on prognostic risk group stratification incorporating molecular classification rather than applying uniform treatment according to stage alone [99].
4. Predictive biomarker for immunotherapy
In May 2017, the U.S. FDA issued the first site-agnostic approval, granting accelerated approval to pembrolizumab for adult and pediatric patients with unresectable or metastatic dMMR/MSI-H solid tumors that had progressed on prior therapy, specifically as a second-line or later treatment option [100]. In March 2023, this indication was converted to full approval based on enlarged datasets from KEYNOTE-158, KEYNOTE-164, and pediatric cases of KEYNOTE-051, confirming durable activity across diverse solid tumors.
In addition to pan-tumor indications, MSI or MMR status serves as a tumor-specific biomarker that guides immunotherapy in several malignancies. In CRC, pembrolizumab was approved as the first-line therapy for metastatic dMMR/MSI-H tumors in June 2020, based on KEYNOTE-177 [101], whereas nivolumab, alone or in combination with ipilimumab, was approved in April 2025 and showed durable benefits in previously untreated disease [102]. In GC and gastroesophageal junction cancers, although there is no disease-specific MSI-H label beyond the site agnostic indication, dMMR/MSI-H consistently predicted high response rates to pembrolizumab across multiple KEYNOTE trials, supporting its use, although no tumor-specific label beyond site agnostic approval exists [103]. Taken together, these examples highlight that the MSI/MMR status not only defines a pan-tumor biomarker but also functions as a clinically actionable predictor within individual cancer types.
Beyond these approved indications, MSI/MMR status is also being investigated as a predictive biomarker in resectable disease. Recent phase II neoadjuvant trials in locally advanced dMMR/MSI-H CRC have demonstrated substantial clinical and pathological responses, and have even explored organ-preserving, non-operative strategies [104-106]. Although these approaches remain investigational and have not led to specific regulatory approvals, they suggest that the role of MSI/MMR status as a biomarker may further expand from guiding treatment in advanced disease to informing curative-intent immunotherapy in localized settings.
5. Clinical significance of MSI-L
MSI-L can be detected when tumors show minor microsatellite slippage, but current evidence suggests it does not represent a distinct biological or histological category separate from MSS tumors [107]. Although many non–MSI-H tumors display some degree of instability, no specific clinicopathological or molecular features consistently distinguish MSI-L from MSS tumors [108,109]. With respect to prognostic significance, prior studies have yielded conflicting results. Some investigations demonstrated that MSI-L was associated with poorer overall or disease-free survival, whereas others reported no difference or even better outcomes [109-112]. However, it should be noted that the diagnosis of MSI-L is affected by the number of microsatellite markers employed, the types of loci evaluated, and the criteria used to define MSI-L [109]. In addition, MSI-L cannot be determined on the basis of MMR IHC alone.
1. Immune microenvironment in dMMR/MSI-H tumors
dMMR/MSI-H tumors exhibit pronounced hypermutations that produce numerous nonsynonymous frameshift mutations and a high burden of nonfunctional truncated proteins across pathways, including DNA repair, epigenetic regulation, apoptosis, miRNA processing, and cell signaling [6,113]. These aberrant proteins generate neoantigens that provoke immune responses, resulting in a tumor microenvironment enriched with tumor-infiltrating lymphocytes and inflammatory signaling accompanied by the upregulation of multiple immune checkpoints [114,115].
This leads to a common immunogenic phenotype in different malignancies. In CRC, MSI-H tumors mostly display dense intratumoral lymphocytes and a Crohn-like lymphoid reaction at the invasive edge, which are well-established favorable prognostic factors [64]. Transcriptomic and tissue-level studies further showed a microenvironment oriented toward type 1 helper T cell and cytotoxic T lymphocyte activity, together with the inducible expression of inhibitory checkpoint molecules [116]. Similarly, MSI-H GCs frequently exhibit PD-L1 positivity and a high density of CD8-positive lymphocytes [117] and carry a greater mutational load than MSS tumors [118]. Similarly, in EC, dMMR tumors consistently harbor abundant lymphocytes with increased CD8-positive cells and enriched PD-1/PD-L1 pathway relative to MSS tumors [119,120].
In general, dMMR/MSI-H tumors, which carry abundant neoantigens, are immunologically defined by chemokine-driven T-cell infiltration and adaptive upregulation of immune checkpoints. Consequently, dMMR/MSI-H solid tumors are promising candidates for immunotherapy.
2. Resistance to immunotherapy in dMMR/MSI-H tumors
Although dMMR/MSI-H tumors are generally characterized by high immunogenicity, their antitumor immune function is not uniform. Several mechanisms contribute to this variability in treatment response. One key mechanism is defective antigen presentation, which allows immune escape and promotes resistance to immune checkpoint blockade [121]. Loss of β2-microglobulin or broader loss of human leukocyte antigen class I expression can disrupt antigen display. However, in MSI-H CRC, these alterations do not consistently predict a lack of benefit from PD-1 blockade [122,123], underscoring the role of tumor- and disease-specific context and the need for further validation of these associations.
Even with a high mutation burden, dMMR/MSI-H tumors may develop an immunosuppressive tumor microenvironment that limits immune activity. Activation of the Wnt/β-catenin pathway impairs T-cell infiltration, while a TGFβ-rich stroma restricts immune cell trafficking and inhibits Th1 and cytotoxic T-cell functions [124]. In CRC models, inhibition of TGFβ signaling restores immune infiltration and responsiveness [125]. Furthermore, strong immune activation in dMMR/MSI-H tumors can induce compensatory expression of alternative immune checkpoints—such as lymphocyte activation gene-3 (LAG-3), T cell immunoreceptor with Ig and ITIM domains (TIGIT), and T cell immunoglobulin and mucin-domain containing-3 (TIM-3)—attenuating the efficacy of PD-1 blockade [116].
Heterogeneity in MSI/MMR status itself also represents a potential mechanism of resistance to immunotherapy in dMMR/MSI-H tumors. Spatial and temporal variations in MMR protein expression and MSI status have been reported across several cancer types [126]. In GC, heterogeneous MMR staining and discordant MSI results have been observed, including cases of MSS primary tumors with MSI-H nodal metastases [127]. In CRC, heterogeneous MMR expression was reported in 11.5% of stage II-III resection cases [128], and a metastatic CRC cohort showed discordant MMR status between primary and matched metastatic sites in 11.9% of patients [129].
Collectively, these findings indicate that even among dMMR/MSI-H solid tumors, the immune microenvironment is not homogeneous, and resistance mechanisms such as defective antigen presentation, immune suppression, and MSI/MMR heterogeneity can influence responsiveness to immunotherapy. These insights emphasize the need to consider the tumor-specific immune context when interpreting MMR/MSI status and optimizing immunotherapeutic strategies.
dMMR generates MSI-H, a hypermutable state that drives frameshift mutations in coding mononucleotide tracts. These events disable key growth suppressors and promote tumor initiation and progression, with a set of recurrent targets differing according to the tumor type. Clinically, dMMR/MSI status is used for Lynch syndrome screening, defining a distinct molecular subtype, predicting limited benefit from adjuvant chemotherapy, and guiding the use of ICIs as CDx. Because treatment decisions now depend on MSI-CDx results, an accurate and reliable diagnosis of MMR/MSI status using CDx assays is essential and should be guided by a clear understanding of the characteristics and technical limitations of each assay. To meet this standard in routine practice, interpretation should be performed by experienced or specialized pathologists and supported by structured educational sessions. Finally, a thorough understanding of MSI biology is required to guide immunotherapy selection, as dMMR/MSI-H tumors are heterogeneous and differences within a tumor and across lesions, including the functional immune microenvironment, can influence response.

Author Contributions

Conceived and designed the analysis: Lee HS.

Collected the data: Kwak Y, Bae JM, Lee HS.

Contributed data or analysis tools: Kwak Y, Bae JM, Lee HS.

Performed the analysis: Kwak Y, Bae JM, Lee HS.

Wrote the paper: Kwak Y, Bae JM, Lee HS.

Conflicts of Interest

Conflict of interest relevant to this article was not reported.

Funding

This study was supported by the Korea-US Collaborative Cancer R&D Program, funded by the Ministry of Health and Welfare, Republic of Korea (RS-2024-00442017).

Fig. 1.
Diagnostic approaches for deficient mismatch repair (dMMR)/microsatellite instability–high (MSI-H). This schematic summarizes four laboratory methods used in practice. Immunohistochemistry (IHC) for MLH1, PMS2, MSH2, and MSH6 (clinical use) evaluates nuclear staining in tumor cells with adjacent normal elements as the internal positive control, and classifies tumors as proficient MMR or dMMR. Polymerase chain reaction–based MSI testing with Bethesda or pentaplex panels (clinical use) examines length shifts in predefined microsatellite markers to call MSI-H or microsatellite-stable (MSS). Tissue next-generation sequencing with MSI calling (clinical use for selected panels; others for research use) assesses MSI status within a broader genomic assay and reports MSI-H or MSS together with relevant alterations. Circulating tumor DNA (ctDNA)–based MSI testing (research use) applies similar principles to plasma-derived cell-free DNA and may be informative when tissue is limited. ddPCR, droplet digital PCR; NGS, next-generation sequencing.
crt-2025-1161f1.jpg
Fig. 2.
Example of abnormal punctate nucleolar staining on MLH1 immunohistochemistry (×200). Four immunohistochemistry panels from the same tumor show loss of MLH1 and PMS2 in tumor nuclei, with retention of MSH2 and MSH6; MLH1 displays punctate nucleolar staining despite intact internal positive controls. This represents deficient mismatch repair with MLH1/PMS2 loss, and the nucleolar puncta should not be interpreted as true nuclear retention.
crt-2025-1161f2.jpg
Table 1.
Requirement of tissue amount for MMR or MSI testing
Tests Thickness of slides (µm) No. of slides
Endoscopic biopsy MSI 10 ×8-10
NGS 10 ×10-20
MMR IHC 4 ×4
Surgical resection (1×1 cm) MSI 10 ×2
NGS 10 ×2
MMR IHC 4 ×4

IHC, immunohistochemistry; MMR, mismatch repair; MSI, microsatellite instability; NGS, next-generation sequencing.

Table 2.
Prevalence and clinical features of dMMR/MSI-H tumors
Tumor type Overall prevalence (%) Clinical feature Reference
Colorectal adenocarcinoma 10-20 Better prognosis, less benefit from adjuvant therapy, lower stage, right colon, poorly differentiated or mucinous histology, expansile growth, high TIL infiltration, high PD-L1 (stage II, 20%; stage III, 10%-15%; stage, IV 4%-5%) (right colon, 13.5%-27%; left colon, 2.0%-2.2%; rectum, 2.2%-9.2%) [68-70]
Endometrial carcinoma (uterine) 16-26 Intermediate prognosis, controversial benefit from adjuvant therapy, endometrioid subtype; grade 3, high TIL infiltration, high PD-L1 [69,71-73]
Gastric adenocarcinoma 8-22 Better prognosis, controversial benefit from adjuvant therapy, lower stage, antral location, localized gross type, intestinal type, expansile growth, high TIL infiltration, high PD-L1 [5,69,71,74]
Small bowel adenocarcinoma 10-26 Lower stage; less common in metastatic disease [69,75,76]
Ovarian carcinoma 5-15 Common in endometrioid and clear cell carcinoma [69,77]
AOV carcinoma ~18 - [78]
Urothelial carcinoma 5-10 - [69]
Prostate adenocarcinoma 1-3 - [50,79]
Biliary tract adenocarcinoma 0-5 - [69,70]
Pancreatic ductal adenocarcinoma 0-2 Medullary or colloid histology, high tumor mutational burden [78,80]
Non–small cell lung cancer 0.4-0.6 Smoking, high tumor mutational burden, MLH1 inactivation [71,81]
Breast ductal carcinoma 0-0.2 - [79,82]
Glioblastoma 0-1 - [79,83]

AOV, ampulla of Vater; dMMR, deficient mismatch repair; MSI-H, microsatellite instability–high; PD-L1, programmed death-ligand 1; TIL, tumor-infiltrating lymphocytes.

  • 1. Dumbovic G, Forcales SV, Perucho M. Emerging roles of macrosatellite repeats in genome organization and disease development. Epigenetics. 2017;12:515–26. ArticlePubMedPMCPDF
  • 2. Bagshaw AT. Functional mechanisms of microsatellite DNA in eukaryotic genomes. Genome Biol Evol. 2017;9:2428–43. ArticlePubMedPMC
  • 3. Gasparello J, Piva VM, Angerilli V, Ceccon C, Sabbadin M, Luchini C, et al. Lights and shadows of microsatellite status characterization in gastrointestinal cancers in the era of cancer precision therapy. Pathologica. 2025;117:204–19. ArticlePubMedPMC
  • 4. Schlotterer C. Genome evolution: are microsatellites really simple sequences? Curr Biol. 1998;8:R132–4. ArticlePubMed
  • 5. Lee HS, Choi SI, Lee HK, Kim HS, Yang HK, Kang GH, et al. Distinct clinical features and outcomes of gastric cancers with microsatellite instability. Mod Pathol. 2002;15:632–40. ArticlePubMedPDF
  • 6. Cortes-Ciriano I, Lee S, Park WY, Kim TM, Park PJ. A molecular portrait of microsatellite instability across multiple cancers. Nat Commun. 2017;8:15180.ArticlePubMedPMCPDF
  • 7. Kim TM, Laird PW, Park PJ. The landscape of microsatellite instability in colorectal and endometrial cancer genomes. Cell. 2013;155:858–68. ArticlePubMedPMC
  • 8. Shia J. The diversity of tumours with microsatellite instability: molecular mechanisms and impact upon microsatellite instability testing and mismatch repair protein immunohistochemistry. Histopathology. 2021;78:485–97. ArticlePubMedPDF
  • 9. Garg P, Pareek S, Kulkarni P, Horne D, Salgia R, Singhal SS. Exploring the potential of TGFβ as a diagnostic marker and therapeutic target against cancer. Biochem Pharmacol. 2025;231:116646.ArticlePubMed
  • 10. Zhang YE. Non-Smad pathways in TGF-beta signaling. Cell Res. 2009;19:128–39. ArticlePubMedPMCPDF
  • 11. Principe DR, Doll JA, Bauer J, Jung B, Munshi HG, Bartholin L, et al. TGF-β: duality of function between tumor prevention and carcinogenesis. J Natl Cancer Inst. 2014;106:djt369.ArticlePubMedPMC
  • 12. Giannakis M, Hodis E, Jasmine Mu X, Yamauchi M, Rosenbluh J, Cibulskis K, et al. Rnf43 is frequently mutated in colorectal and endometrial cancers. Nat Genet. 2014;46:1264–6. ArticlePubMedPMCPDF
  • 13. Wang K, Yuen ST, Xu J, Lee SP, Yan HH, Shi ST, et al. Whole-genome sequencing and comprehensive molecular profiling identify new driver mutations in gastric cancer. Nat Genet. 2014;46:573–82. ArticlePubMedPDF
  • 14. Cancer Genome Atlas Research Network. Comprehensive molecular characterization of gastric adenocarcinoma. Nature. 2014;513:202–9. ArticlePubMedPMCPDF
  • 15. Koo BK, Spit M, Jordens I, Low TY, Stange DE, van de Wetering M, et al. Tumour suppressor RNF43 is a stem-cell E3 ligase that induces endocytosis of Wnt receptors. Nature. 2012;488:665–9. ArticlePubMedPDF
  • 16. Tsukiyama T, Fukui A, Terai S, Fujioka Y, Shinada K, Takahashi H, et al. Molecular role of RNF43 in canonical and noncanonical Wnt signaling. Mol Cell Biol. 2015;35:2007–23. ArticlePubMedPMCPDF
  • 17. Yan HH, Lai JCW, Ho SL, Leung WK, Law WL, Lee JF, et al. Rnf43 germline and somatic mutation in serrated neoplasia pathway and its association with BRAF mutation. Gut. 2017;66:1645–56. ArticlePubMed
  • 18. Yu J, Yusoff PA, Woutersen DT, Goh P, Harmston N, Smits R, et al. The functional landscape of patient-derived RNF43 mutations predicts sensitivity to Wnt inhibition. Cancer Res. 2020;80:5619–32. ArticlePubMedPDF
  • 19. Samstein RM, Lee CH, Shoushtari AN, Hellmann MD, Shen R, Janjigian YY, et al. Tumor mutational load predicts survival after immunotherapy across multiple cancer types. Nat Genet. 2019;51:202–6. PubMedPMC
  • 20. Principe DR. Patients deriving long-term benefit from immune checkpoint inhibitors demonstrate conserved patterns of site-specific mutations. Sci Rep. 2022;12:11490.ArticlePubMedPMCPDF
  • 21. Huang K, Ding S, Chen K, Guo C, Wu M, Zhang R, et al. Rnf43 mutation as a predictor of immunotherapeutic efficacy in colorectal cancer. Am J Cancer Res. 2023;13:5549–58. PubMedPMC
  • 22. Companion diagnostics [Internet]. Food and Drug Administration; 2023 [cited 2025 Oct 1]. Available from: https://www.fda.gov/medical-devices/in-vitro-diagnostics/companion-diagnostics
  • 23. Shia J, Tang LH, Vakiani E, Guillem JG, Stadler ZK, Soslow RA, et al. Immunohistochemistry as first-line screening for detecting colorectal cancer patients at risk for hereditary nonpolyposis colorectal cancer syndrome: a 2-antibody panel may be as predictive as a 4-antibody panel. Am J Surg Pathol. 2009;33:1639–45. PubMed
  • 24. Shia J, Zhang L, Shike M, Guo M, Stadler Z, Xiong X, et al. Secondary mutation in a coding mononucleotide tract in MSH6 causes loss of immunoexpression of MSH6 in colorectal carcinomas with MLH1/PMS2 deficiency. Mod Pathol. 2013;26:131–8. ArticlePubMedPMCPDF
  • 25. Reitsam NG, Markl B, Dintner S, Waidhauser J, Vlasenko D, Grosser B. Concurrent loss of MLH1, PMS2 and MSH6 immunoexpression in digestive system cancers indicating a widespread dysregulation in DNA repair processes. Front Oncol. 2022;12:1019798.ArticlePubMedPMC
  • 26. Moreno E, Rosa-Rosa JM, Caniego-Casas T, Ruz-Caracuel I, Perna C, Guillen C, et al. Next generation sequencing to decipher concurrent loss of PMS2 and MSH6 in colorectal cancer. Diagn Pathol. 2020;15:84.ArticlePubMedPMCPDF
  • 27. Aiyer KT, Doeleman T, Ryan NA, Nielsen M, Crosbie EJ, Smit VT, et al. Validity of a two-antibody testing algorithm for mismatch repair deficiency testing in cancer: a systematic literature review and meta-analysis. Mod Pathol. 2022;35:1775–83. ArticlePubMedPMCPDF
  • 28. Park Y, Nam SK, Seo SH, Park KU, Oh HJ, Park YS, et al. Comprehensive study of microsatellite instability testing and its comparison with immunohistochemistry in gastric cancers. J Gastric Cancer. 2023;23:264–74. ArticlePubMedPMCPDF
  • 29. Niu BT, Hammond RF, Leen SL, Faruqi AZ, Trevisan G, Gilks CB, et al. Artefactual punctate MLH1 staining can lead to erroneous reporting of isolated PMS2 loss. Histopathology. 2018;73:703–5. ArticlePubMedPDF
  • 30. Zhang Q, Young GQ, Yang Z. Pure discrete punctate nuclear staining pattern for MLH1 protein does not represent intact nuclear expression. Int J Surg Pathol. 2020;28:146–52. ArticlePubMedPDF
  • 31. Wang C, Zhang L, Vakiani E, Shia J. Detecting mismatch repair deficiency in solid neoplasms: immunohistochemistry, microsatellite instability, or both? Mod Pathol. 2022;35:1515–28. ArticlePubMedPDF
  • 32. Kim HS, Woo DK, Bae SI, Kim YI, Kim WH. Microsatellite instability in the adenoma-carcinoma sequence of the stomach. Lab Invest. 2000;80:57–64. ArticlePubMedPDF
  • 33. Boland CR, Thibodeau SN, Hamilton SR, Sidransky D, Eshleman JR, Burt RW, et al. A national cancer institute workshop on microsatellite instability for cancer detection and familial predisposition: development of international criteria for the determination of microsatellite instability in colorectal cancer. Cancer Res. 1998;58:5248–57. PubMed
  • 34. Suraweera N, Duval A, Reperant M, Vaury C, Furlan D, Leroy K, et al. Evaluation of tumor microsatellite instability using five quasimonomorphic mononucleotide repeats and pentaplex PCR. Gastroenterology. 2002;123:1804–11. ArticlePubMed
  • 35. Murphy KM, Zhang S, Geiger T, Hafez MJ, Bacher J, Berg KD, et al. Comparison of the microsatellite instability analysis system and the Bethesda panel for the determination of microsatellite instability in colorectal cancers. J Mol Diagn. 2006;8:305–11. ArticlePubMedPMC
  • 36. Bonneville R, Krook MA, Chen HZ, Smith A, Samorodnitsky E, Wing MR, et al. Detection of microsatellite instability biomarkers via next-generation sequencing. Methods Mol Biol. 2020;2055:119–32. ArticlePubMedPMC
  • 37. Wu X, Snir O, Rottmann D, Wong S, Buza N, Hui P. Minimal microsatellite shift in microsatellite instability high endometrial cancer: a significant pitfall in diagnostic interpretation. Mod Pathol. 2019;32:650–8. ArticlePubMedPDF
  • 38. Chung Y, Nam SK, Chang HE, Lee C, Kang GH, Lee HS, et al. Evaluation of an eight marker-panel including long mononucleotide repeat markers to detect microsatellite instability in colorectal, gastric, and endometrial cancers. BMC Cancer. 2023;23:1100.ArticlePubMedPMCPDF
  • 39. Wang Y, Shi C, Eisenberg R, Vnencak-Jones CL. Differences in microsatellite instability profiles between endometrioid and colorectal cancers: a potential cause for false-negative results? J Mol Diagn. 2017;19:57–64. PubMedPMC
  • 40. Silveira AB, Bidard FC, Kasperek A, Melaabi S, Tanguy ML, Rodrigues M, et al. High-accuracy determination of microsatellite instability compatible with liquid biopsies. Clin Chem. 2020;66:606–13. ArticlePubMedPDF
  • 41. Frampton GM, Fichtenholtz A, Otto GA, Wang K, Downing SR, He J, et al. Development and validation of a clinical cancer genomic profiling test based on massively parallel DNA sequencing. Nat Biotechnol. 2013;31:1023–31. ArticlePubMedPMCPDF
  • 42. Gibbs SN, Peneva D, Cuyun Carter G, Palomares MR, Thakkar S, Hall DW, et al. Comprehensive review on the clinical impact of next-generation sequencing tests for the management of advanced cancer. JCO Precis Oncol. 2023;7:e2200715ArticlePubMedPMC
  • 43. Middha S, Zhang L, Nafa K, Jayakumaran G, Wong D, Kim HR, et al. Reliable pan-cancer microsatellite instability assessment by using targeted next-generation sequencing data. JCO Precis Oncol. 2017;2017:PO.17.00084.ArticlePubMedPMC
  • 44. Niu B, Ye K, Zhang Q, Lu C, Xie M, McLellan MD, et al. MSIsensor: microsatellite instability detection using paired tumor-normal sequence data. Bioinformatics. 2014;30:1015–6. ArticlePubMedPMCPDF
  • 45. Salipante SJ, Scroggins SM, Hampel HL, Turner EH, Pritchard CC. Microsatellite instability detection by next generation sequencing. Clin Chem. 2014;60:1192–9. ArticlePubMedPDF
  • 46. Kautto EA, Bonneville R, Miya J, Yu L, Krook MA, Reeser JW, et al. Performance evaluation for rapid detection of pan-cancer microsatellite instability with MANTIS. Oncotarget. 2017;8:7452–63. ArticlePubMedPMC
  • 47. Huang MN, McPherson JR, Cutcutache I, Teh BT, Tan P, Rozen SG. MsiSeq: software for assessing microsatellite instability from catalogs of somatic mutations. Sci Rep. 2015;5:13321.PubMedPMC
  • 48. Nowak JA, Yurgelun MB, Bruce JL, Rojas-Rudilla V, Hall DL, Shivdasani P, et al. Detection of mismatch repair deficiency and microsatellite instability in colorectal adenocarcinoma by targeted next-generation sequencing. J Mol Diagn. 2017;19:84–91. ArticlePubMedPMC
  • 49. Lin DI, Quintanilha JCF, Danziger N, Lang L, Levitan D, Hayne C, et al. Pan-tumor validation of a NGS fraction-based MSI analysis as a predictor of response to Pembrolizumab. NPJ Precis Oncol. 2024;8:204.ArticlePubMedPMCPDF
  • 50. Abida W, Cheng ML, Armenia J, Middha S, Autio KA, Vargas HA, et al. Analysis of the prevalence of microsatellite instaYoonjin bility in prostate cancer and response to immune checkpoint blockade. JAMA Oncol. 2019;5:471–8. ArticlePubMedPMC
  • 51. Walker CJ, Miranda MA, O’Hern MJ, Blachly JS, Moyer CL, Ivanovich J, et al. MonoSeq variant caller reveals novel mononucleotide run indel mutations in tumors with defective DNA mismatch repair. Hum Mutat. 2016;37:1004–12. ArticlePubMedPMC
  • 52. Loman NJ, Misra RV, Dallman TJ, Constantinidou C, Gharbia SE, Wain J, et al. Performance comparison of benchtop high-throughput sequencing platforms. Nat Biotechnol. 2012;30:434–9. ArticlePubMedPDF
  • 53. Anthony H, Seoighe C. Performance assessment of computational tools to detect microsatellite instability. Brief Bioinform. 2024;25:bbae390.ArticlePubMedPMCPDF
  • 54. Bartley AN, Mills AM, Konnick E, Overman M, Ventura CB, Souter L, et al. Mismatch repair and microsatellite instability testing for immune checkpoint inhibitor therapy: guideline from the College of American Pathologists in collaboration with the Association for Molecular Pathology and Fight Colorectal Cancer. Arch Pathol Lab Med. 2022;146:1194–210. ArticlePubMedPDF
  • 55. Ignatiadis M, Sledge GW, Jeffrey SS. Liquid biopsy enters the clinic: implementation issues and future challenges. Nat Rev Clin Oncol. 2021;18:297–312. ArticlePubMedPDF
  • 56. Pascual J, Attard G, Bidard FC, Curigliano G, De Mattos-Arruda L, Diehn M, et al. ESMO recommendations on the use of circulating tumour DNA assays for patients with cancer: a report from the ESMO Precision Medicine Working Group. Ann Oncol. 2022;33:750–68. ArticlePubMed
  • 57. Vidula N, Lipman A, Kato S, Weipert C, Hesler K, Azzi G, et al. Detection of microsatellite instability high (MSI-H) status by targeted plasma-based genotyping in metastatic breast cancer. NPJ Breast Cancer. 2022;8:117.ArticlePubMedPMCPDF
  • 58. Nakamura Y, Okamoto W, Denda T, Nishina T, Komatsu Y, Yuki S, et al. Clinical validity of plasma-based genotyping for microsatellite instability assessment in advanced GI cancers: SCRUM-Japan GOZILA substudy. JCO Precis Oncol. 2022;6:e2100383ArticlePubMedPMC
  • 59. Willis J, Lefterova MI, Artyomenko A, Kasi PM, Nakamura Y, Mody K, et al. Validation of microsatellite instability detection using a comprehensive plasma-based genotyping panel. Clin Cancer Res. 2019;25:7035–45. ArticlePubMedPDF
  • 60. Mishima S, Nakamura Y, Tukachinsky H, Taniguchi H, Kadowaki S, Kato K, et al. Validity and utility of blood tumor mutational burden (bTMB) is dependent on circulating tumor DNA (ctDNA) shed: SCRUM-Japan MONSTAR-SCREEN. J Liq Biopsy. 2023;1:100003.ArticlePubMedPMC
  • 61. Le DT, Uram JN, Wang H, Bartlett BR, Kemberling H, Eyring AD, et al. PD-1 blockade in tumors with mismatch-repair deficiency. N Engl J Med. 2015;372:2509–20. PubMedPMC
  • 62. Andre T, Shiu KK, Kim TW, Jensen BV, Jensen LH, Punt C, et al. Pembrolizumab in microsatellite-instability-high advanced colorectal cancer. N Engl J Med. 2020;383:2207–18. PubMed
  • 63. Marabelle A, Le DT, Ascierto PA, Di Giacomo AM, De Jesus-Acosta A, Delord JP, et al. Efficacy of pembrolizumab in patients with noncolorectal high microsatellite instability/mismatch repair-deficient cancer: results from the phase II KEYNOTE-158 study. J Clin Oncol. 2020;38:1–10. PubMed
  • 64. Chakrabarti S, Bucheit L, Starr JS, Innis-Shelton R, Shergill A, Dada H, et al. Detection of microsatellite instability-high (MSI-H) by liquid biopsy predicts robust and durable response to immunotherapy in patients with pancreatic cancer. J Immunother Cancer. 2022;10:e004485ArticlePubMedPMC
  • 65. Kasi PM, Bucheit LA, Liao J, Starr J, Barata P, Klempner SJ. Pan-cancer prevalence of microsatellite instability-high (MSI-H) identified by circulating tumor DNA and associated real-world clinical outcomes. JCO Precis Oncol. 2023;7:e2300118ArticlePubMed
  • 66. Guyot D’Asnieres De Salins A, Tachon G, Cohen R, Karayan-Tapon L, Junca A, Frouin E, et al. Discordance between immunochemistry of mismatch repair proteins and molecular testing of microsatellite instability in colorectal cancer. ESMO Open. 2021;6:100120.ArticlePubMedPMC
  • 67. Stelloo E, Jansen AM, Osse EM, Nout RA, Creutzberg CL, Ruano D, et al. Practical guidance for mismatch repair-deficiency testing in endometrial cancer. Ann Oncol. 2017;28:96–102. ArticlePubMed
  • 68. Fan WX, Su F, Zhang Y, Zhang XL, Du YY, Gao YJ, et al. Oncological characteristics, treatments and prognostic outcomes in MMR-deficient colorectal cancer. Biomark Res. 2024;12:89.ArticlePubMedPMCPDF
  • 69. Ponti G, Gelsomino F, Tomasi A, Ozben T, Losi L, Manfredini M. Hereditary micro‑satellite instable cancers associated with Lynch Syndrome: predictive biomarkers and novel immuno-therapeutic approaches. Crit Rev Clin Lab Sci. 2025;62:477–90. ArticlePubMed
  • 70. Eso Y, Shimizu T, Takeda H, Takai A, Marusawa H. Microsatellite instability and immune checkpoint inhibitors: toward precision medicine against gastrointestinal and hepatobiliary cancers. J Gastroenterol. 2020;55:15–26. ArticlePubMedPMCPDF
  • 71. Vanderwalde A, Spetzler D, Xiao N, Gatalica Z, Marshall J. Microsatellite instability status determined by next-generation sequencing and compared with PD-L1 and tumor mutational burden in 11,348 patients. Cancer Med. 2018;7:746–56. PubMedPMC
  • 72. Espinosa I, D’Angelo E, Prat J. Endometrial carcinoma: 10 years of TCGA (the cancer genome atlas): a critical reappraisal with comments on FIGO 2023 staging. Gynecol Oncol. 2024;186:94–103. ArticlePubMed
  • 73. Stelloo E, Bosse T, Nout RA, MacKay HJ, Church DN, Nijman HW, et al. Refining prognosis and identifying targetable pathways for high-risk endometrial cancer: a TransPORTEC initiative. Mod Pathol. 2015;28:836–44. ArticlePubMedPDF
  • 74. Choi J, Nam SK, Park DJ, Kim HW, Kim HH, Kim WH, et al. Correlation between microsatellite instability-high phenotype and occult lymph node metastasis in gastric carcinoma. APMIS. 2015;123:215–22. ArticlePubMed
  • 75. Aparicio T, Zaanan A, Svrcek M, Laurent-Puig P, Carrere N, Manfredi S, et al. Small bowel adenocarcinoma: epidemiology, risk factors, diagnosis and treatment. Dig Liver Dis. 2014;46:97–104. ArticlePubMed
  • 76. Latham A, Shia J, Patel Z, Reidy-Lagunes DL, Segal NH, Yaeger R, et al. Characterization and clinical outcomes of DNA mismatch repair-deficient small bowel adenocarcinoma. Clin Cancer Res. 2021;27:1429–37. ArticlePubMedPMCPDF
  • 77. Xiao X, Melton DW, Gourley C. Mismatch repair deficiency in ovarian cancer: molecular characteristics and clinical implications. Gynecol Oncol. 2014;132:506–12. ArticlePubMed
  • 78. Luchini C, Scarpa A. Microsatellite instability in pancreatic and ampullary carcinomas: histology, molecular pathology, and clinical implications. Hum Pathol. 2023;132:176–82. ArticlePubMed
  • 79. Hause RJ, Pritchard CC, Shendure J, Salipante SJ. Classification and characterization of microsatellite instability across 18 cancer types. Nat Med. 2016;22:1342–50. ArticlePubMedPDF
  • 80. Hu ZI, Shia J, Stadler ZK, Varghese AM, Capanu M, Salo-Mullen E, et al. Evaluating mismatch repair deficiency in pancreatic adenocarcinoma: challenges and recommendations. Clin Cancer Res. 2018;24:1326–36. ArticlePubMedPMCPDF
  • 81. Yang SR, Gedvilaite E, Ptashkin R, Chang J, Ziegler J, Mata DA, et al. Microsatellite instability and mismatch repair deficiency define a distinct subset of lung cancers characterized by smoking exposure, high tumor mutational burden, and recurrent somatic MLH1 inactivation. J Thorac Oncol. 2024;19:409–24. ArticlePubMedPMC
  • 82. Ren XY, Song Y, Wang J, Chen LY, Pang JY, Zhou LR, et al. Mismatch repair deficiency and microsatellite instability in triple-negative breast cancer: a retrospective study of 440 patients. Front Oncol. 2021;11:570623.ArticlePubMedPMC
  • 83. Touat M, Li YY, Boynton AN, Spurr LF, Iorgulescu JB, Bohrson CL, et al. Mechanisms and therapeutic implications of hypermutation in gliomas. Nature. 2020;580:517–23. PubMedPMC
  • 84. Cancer Genome Atlas Network. Comprehensive molecular characterization of human colon and rectal cancer. Nature. 2012;487:330–7. ArticlePubMedPMCPDF
  • 85. Cancer Genome Atlas Research Network; Kandoth C, Schultz N, Cherniack AD, Akbani R, Liu Y, et al. Integrated genomic characterization of endometrial carcinoma. Nature. 2013;497:67–73. ArticlePubMedPMCPDF
  • 86. Guinney J, Dienstmann R, Wang X, de Reynies A, Schlicker A, Soneson C, et al. The consensus molecular subtypes of colorectal cancer. Nat Med. 2015;21:1350–6. PubMedPMC
  • 87. Boland CR, Goel A. Microsatellite instability in colorectal cancer. Gastroenterology. 2010;138:2073–87. ArticlePubMedPMC
  • 88. Popat S, Hubner R, Houlston RS. Systematic review of microsatellite instability and colorectal cancer prognosis. J Clin Oncol. 2005;23:609–18. ArticlePubMed
  • 89. Kobel M, Hoang LN, Tessier-Cloutier B, Meng B, Soslow RA, Stewart CJ, et al. Undifferentiated endometrial carcinomas show frequent loss of core switch/sucrose nonfermentable complex proteins. Am J Surg Pathol. 2018;42:76–83. ArticlePubMedPMC
  • 90. Ribic CM, Sargent DJ, Moore MJ, Thibodeau SN, French AJ, Goldberg RM, et al. Tumor microsatellite-instability status as a predictor of benefit from fluorouracil-based adjuvant chemotherapy for colon cancer. N Engl J Med. 2003;349:247–57. ArticlePubMedPMC
  • 91. Sargent DJ, Marsoni S, Monges G, Thibodeau SN, Labianca R, Hamilton SR, et al. Defective mismatch repair as a predictive marker for lack of efficacy of fluorouracil-based adjuvant therapy in colon cancer. J Clin Oncol. 2010;28:3219–26. PubMedPMC
  • 92. Zeinalian M, Hashemzadeh-Chaleshtori M, Salehi R, Emami MH. Clinical aspects of microsatellite instability testing in colorectal cancer. Adv Biomed Res. 2018;7:28.ArticlePubMedPMC
  • 93. Andre T, de Gramont A, Vernerey D, Chibaudel B, Bonnetain F, Tijeras-Raballand A, et al. Adjuvant fluorouracil, leucovorin, and oxaliplatin in stage II to III colon cancer: updated 10-year survival and outcomes according to BRAF mutation and mismatch repair status of the MOSAIC study. J Clin Oncol. 2015;33:4176–87. ArticlePubMed
  • 94. An JY, Kim H, Cheong JH, Hyung WJ, Kim H, Noh SH. Microsatellite instability in sporadic gastric cancer: its prognostic role and guidance for 5-FU based chemotherapy after R0 resection. Int J Cancer. 2012;131:505–11. ArticlePubMed
  • 95. Choi YY, Kim H, Shin SJ, Kim HY, Lee J, Yang HK, et al. Microsatellite instability and programmed cell death-ligand 1 expression in stage II/III gastric cancer: post hoc analysis of the CLASSIC randomized controlled study. Ann Surg. 2019;270:309–16. PubMed
  • 96. Kim JW, Cho SY, Chae J, Kim JW, Kim TY, Lee KW, et al. Adjuvant chemotherapy in microsatellite instability-high gastric cancer. Cancer Res Treat. 2020;52:1178–87. ArticlePubMedPMC
  • 97. Shitara K, Fleitas T, Kawakami H, Curigliano G, Narita Y, Wang F, et al. Pan-Asian adapted ESMO clinical practice guidelines for the diagnosis, treatment and follow-up of patients with gastric cancer. ESMO Open. 2024;9:102226.ArticlePubMedPMC
  • 98. Leon-Castillo A, de Boer SM, Powell ME, Mileshkin LR, Mackay HJ, Leary A, et al. Molecular classification of the PORTEC-3 trial for high-risk endometrial cancer: impact on prognosis and benefit from adjuvant therapy. J Clin Oncol. 2020;38:3388–97. ArticlePubMedPMC
  • 99. Concin N, Matias-Guiu X, Vergote I, Cibula D, Mirza MR, Marnitz S, et al. ESGO/ESTRO/ESP guidelines for the management of patients with endometrial carcinoma. Int J Gynecol Cancer. 2021;31:12–39. PubMed
  • 100. Marcus L, Lemery SJ, Keegan P, Pazdur R. FDA approval summary: pembrolizumab for the treatment of microsatellite instability-high solid tumors. Clin Cancer Res. 2019;25:3753–8. ArticlePubMedPDF
  • 101. Casak SJ, Marcus L, Fashoyin-Aje L, Mushti SL, Cheng J, Shen YL, et al. FDA approval summary: Pembrolizumab for the first-line treatment of patients with MSI-H/dMMR advanced unresectable or metastatic colorectal carcinoma. Clin Cancer Res. 2021;27:4680–4. ArticlePubMedPMCPDF
  • 102. Andre T, Elez E, Van Cutsem E, Jensen LH, Bennouna J, Mendez G, et al. Nivolumab plus ipilimumab in microsatellite-instability-high metastatic colorectal cancer. N Engl J Med. 2024;391:2014–26. PubMed
  • 103. Yoon J, Kim TY, Oh DY. Recent progress in immunotherapy for gastric cancer. J Gastric Cancer. 2023;23:207–23. ArticlePubMedPMCPDF
  • 104. Cercek A, Lumish M, Sinopoli J, Weiss J, Shia J, Lamendola-Essel M, et al. PD-1 blockade in mismatch repair-deficient, locally advanced rectal cancer. N Engl J Med. 2022;386:2363–76. PubMedPMC
  • 105. Chalabi M, Verschoor YL, Tan PB, Balduzzi S, Van Lent AU, Grootscholten C, et al. Neoadjuvant immunotherapy in locally advanced mismatch repair-deficient colon cancer. N Engl J Med. 2024;390:1949–58. PubMed
  • 106. de Gooyer PG, Verschoor YL, van den Dungen LD, Balduzzi S, Marsman HA, Geukes Foppen MH, et al. Neoadjuvant nivolumab and relatlimab in locally advanced MMR-deficient colon cancer: a phase 2 trial. Nat Med. 2024;30:3284–90. ArticlePubMedPMCPDF
  • 107. Lawes DA, SenGupta S, Boulos PB. The clinical importance and prognostic implications of microsatellite instability in sporadic cancer. Eur J Surg Oncol. 2003;29:201–12. ArticlePubMed
  • 108. Tomlinson I, Halford S, Aaltonen L, Hawkins N, Ward R. Does MSI-low exist? J Pathol. 2002;197:6–13. ArticlePubMed
  • 109. Wright CM, Dent OF, Newland RC, Barker M, Chapuis PH, Bokey EL, et al. Low level microsatellite instability may be associated with reduced cancer specific survival in sporadic stage C colorectal carcinoma. Gut. 2005;54:103–8. ArticlePubMedPMC
  • 110. Lee SY, Kim DW, Lee HS, Ihn MH, Oh HK, Min BS, et al. Low-level microsatellite instability as a potential prognostic factor in sporadic colorectal cancer. Medicine (Baltimore). 2015;94:e2260ArticlePubMedPMC
  • 111. Jiang D, Shu C, Zhang W, Sun L, Zhang M, He Y, et al. Low level of microsatellite instability correlates with short disease-free survival of gastric cancer patients undergoing neoadjuvant chemotherapy. Virchows Arch. 2021;478:231–40. ArticlePubMedPDF
  • 112. Imamura Y, Toihata T, Haraguchi I, Ogata Y, Takamatsu M, Kuchiba A, et al. Immunogenic characteristics of microsatellite instability-low esophagogastric junction adenocarcinoma based on clinicopathological, molecular, immunological and survival analyses. Int J Cancer. 2021;148:1260–75. ArticlePubMedPDF
  • 113. Baretti M, Le DT. DNA mismatch repair in cancer. Pharmacol Ther. 2018;189:45–62. ArticlePubMed
  • 114. Schwitalle Y, Kloor M, Eiermann S, Linnebacher M, Kienle P, Knaebel HP, et al. Immune response against frameshift-induced neopeptides in HNPCC patients and healthy HNPCC mutation carriers. Gastroenterology. 2008;134:988–97. ArticlePubMed
  • 115. Wilbur HC, Le DT, Agarwal P. Immunotherapy of MSI cancer: facts and hopes. Clin Cancer Res. 2024;30:1438–47. ArticlePubMedPDF
  • 116. Llosa NJ, Cruise M, Tam A, Wicks EC, Hechenbleikner EM, Taube JM, et al. The vigorous immune microenvironment of microsatellite instable colon cancer is balanced by multiple counter-inhibitory checkpoints. Cancer Discov. 2015;5:43–51. ArticlePubMedPMCPDF
  • 117. Koh J, Ock CY, Kim JW, Nam SK, Kwak Y, Yun S, et al. Clinicopathologic implications of immune classification by PD-L1 expression and CD8-positive tumor-infiltrating lymphocytes in stage II and III gastric cancer patients. Oncotarget. 2017;8:26356–67. ArticlePubMedPMC
  • 118. Koh J, Nam SK, Roh H, Kim J, Lee BC, Kim JW, et al. Somatic mutational profiles of stage II and III gastric cancer according to tumor microenvironment immune type. Genes Chromosomes Cancer. 2019;58:12–22. ArticlePubMedPDF
  • 119. Willvonseder B, Stogbauer F, Steiger K, Jesinghaus M, Kuhn PH, Brambs C, et al. The immunologic tumor microenvironment in endometrioid endometrial cancer in the morphomolecular context: mutual correlations and prognostic impact depending on molecular alterations. Cancer Immunol Immunother. 2021;70:1679–89. ArticlePubMedPMCPDF
  • 120. Sloan EA, Ring KL, Willis BC, Modesitt SC, Mills AM. PD-L1 expression in mismatch repair-deficient endometrial carcinomas, including Lynch syndrome-associated and MLH1 promoter hypermethylated tumors. Am J Surg Pathol. 2017;41:326–33. ArticlePubMed
  • 121. Zaretsky JM, Garcia-Diaz A, Shin DS, Escuin-Ordinas H, Hugo W, Hu-Lieskovan S, et al. Mutations associated with acquired resistance to PD-1 blockade in melanoma. N Engl J Med. 2016;375:819–29. PubMedPMC
  • 122. Zhang C, Li D, Xiao B, Zhou C, Jiang W, Tang J, et al. B2M and JAK1/2-mutated MSI-H colorectal carcinomas can benefit from anti-PD-1 therapy. J Immunother. 2022;45:187–93. ArticlePubMedPMC
  • 123. Middha S, Yaeger R, Shia J, Stadler ZK, King S, Guercio S, et al. Majority of B2M-mutant and -deficient colorectal carcinomas achieve clinical benefit from immune checkpoint inhibitor therapy and are microsatellite instability-high. JCO Precis Oncol. 2019;3:PO.ArticlePubMedPMC
  • 124. Mariathasan S, Turley SJ, Nickles D, Castiglioni A, Yuen K, Wang Y, et al. TGFβ attenuates tumour response to PD-L1 blockade by contributing to exclusion of T cells. Nature. 2018;554:544–8. ArticlePubMedPMCPDF
  • 125. Tauriello DV, Palomo-Ponce S, Stork D, Berenguer-Llergo A, Badia-Ramentol J, Iglesias M, et al. TGFβ drives immune evasion in genetically reconstituted colon cancer metastasis. Nature. 2018;554:538–43. PubMed
  • 126. Lee HS. Spatial and temporal tumor heterogeneity in gastric cancer: discordance of predictive biomarkers. J Gastric Cancer. 2025;25:192–209. ArticlePubMedPMCPDF
  • 127. Wang X, Jiang K, Hu Y, Zhao X, Yin L, Diao X, et al. An exploration of gastric cancer with heterogeneous mismatch repair status. Virchows Arch. 2023;482:517–23. ArticlePubMedPDF
  • 128. Dencic T, Petrovic A, Jovicic Milentijevic M, Radenkovic G, Jovic M, Zivkovic N, et al. The importance of immunohistochemical heterogeneous expression of MMR protein in patients with colorectal cancer in stage II and III of the disease. Medicina (Kaunas). 2023;59:489.ArticlePubMedPMC
  • 129. Huang Q, Yu T, Li L, Zhang Q, Zhang S, Li B, et al. Intraindividual tumor heterogeneity of mismatch repair status in metastatic colorectal cancer. Appl Immunohistochem Mol Morphol. 2023;31:84–93. ArticlePubMedPMC

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        Diagnostic Challenges and Clinical Implications of Microsatellite Instability/Mismatch Repair Deficiency in Solid Tumors
        Cancer Res Treat. 2026;58(1):1-14.   Published online December 26, 2025
        DOI: https://doi.org/10.4143/crt.2025.1161
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      Diagnostic Challenges and Clinical Implications of Microsatellite Instability/Mismatch Repair Deficiency in Solid Tumors
      Image Image
      Fig. 1. Diagnostic approaches for deficient mismatch repair (dMMR)/microsatellite instability–high (MSI-H). This schematic summarizes four laboratory methods used in practice. Immunohistochemistry (IHC) for MLH1, PMS2, MSH2, and MSH6 (clinical use) evaluates nuclear staining in tumor cells with adjacent normal elements as the internal positive control, and classifies tumors as proficient MMR or dMMR. Polymerase chain reaction–based MSI testing with Bethesda or pentaplex panels (clinical use) examines length shifts in predefined microsatellite markers to call MSI-H or microsatellite-stable (MSS). Tissue next-generation sequencing with MSI calling (clinical use for selected panels; others for research use) assesses MSI status within a broader genomic assay and reports MSI-H or MSS together with relevant alterations. Circulating tumor DNA (ctDNA)–based MSI testing (research use) applies similar principles to plasma-derived cell-free DNA and may be informative when tissue is limited. ddPCR, droplet digital PCR; NGS, next-generation sequencing.
      Fig. 2. Example of abnormal punctate nucleolar staining on MLH1 immunohistochemistry (×200). Four immunohistochemistry panels from the same tumor show loss of MLH1 and PMS2 in tumor nuclei, with retention of MSH2 and MSH6; MLH1 displays punctate nucleolar staining despite intact internal positive controls. This represents deficient mismatch repair with MLH1/PMS2 loss, and the nucleolar puncta should not be interpreted as true nuclear retention.

      Fig. 1.

      Fig. 2.

      Diagnostic Challenges and Clinical Implications of Microsatellite Instability/Mismatch Repair Deficiency in Solid Tumors

      Requirement of tissue amount for MMR or MSI testing

      Tests Thickness of slides (µm) No. of slides
      Endoscopic biopsy MSI 10 ×8-10
      NGS 10 ×10-20
      MMR IHC 4 ×4
      Surgical resection (1×1 cm) MSI 10 ×2
      NGS 10 ×2
      MMR IHC 4 ×4

      IHC, immunohistochemistry; MMR, mismatch repair; MSI, microsatellite instability; NGS, next-generation sequencing.

      Prevalence and clinical features of dMMR/MSI-H tumors

      Tumor type Overall prevalence (%) Clinical feature Reference
      Colorectal adenocarcinoma 10-20 Better prognosis, less benefit from adjuvant therapy, lower stage, right colon, poorly differentiated or mucinous histology, expansile growth, high TIL infiltration, high PD-L1 (stage II, 20%; stage III, 10%-15%; stage, IV 4%-5%) (right colon, 13.5%-27%; left colon, 2.0%-2.2%; rectum, 2.2%-9.2%) [68-70]
      Endometrial carcinoma (uterine) 16-26 Intermediate prognosis, controversial benefit from adjuvant therapy, endometrioid subtype; grade 3, high TIL infiltration, high PD-L1 [69,71-73]
      Gastric adenocarcinoma 8-22 Better prognosis, controversial benefit from adjuvant therapy, lower stage, antral location, localized gross type, intestinal type, expansile growth, high TIL infiltration, high PD-L1 [5,69,71,74]
      Small bowel adenocarcinoma 10-26 Lower stage; less common in metastatic disease [69,75,76]
      Ovarian carcinoma 5-15 Common in endometrioid and clear cell carcinoma [69,77]
      AOV carcinoma ~18 - [78]
      Urothelial carcinoma 5-10 - [69]
      Prostate adenocarcinoma 1-3 - [50,79]
      Biliary tract adenocarcinoma 0-5 - [69,70]
      Pancreatic ductal adenocarcinoma 0-2 Medullary or colloid histology, high tumor mutational burden [78,80]
      Non–small cell lung cancer 0.4-0.6 Smoking, high tumor mutational burden, MLH1 inactivation [71,81]
      Breast ductal carcinoma 0-0.2 - [79,82]
      Glioblastoma 0-1 - [79,83]

      AOV, ampulla of Vater; dMMR, deficient mismatch repair; MSI-H, microsatellite instability–high; PD-L1, programmed death-ligand 1; TIL, tumor-infiltrating lymphocytes.
      Table 1. Requirement of tissue amount for MMR or MSI testing

      IHC, immunohistochemistry; MMR, mismatch repair; MSI, microsatellite instability; NGS, next-generation sequencing.

      Table 2. Prevalence and clinical features of dMMR/MSI-H tumors

      AOV, ampulla of Vater; dMMR, deficient mismatch repair; MSI-H, microsatellite instability–high; PD-L1, programmed death-ligand 1; TIL, tumor-infiltrating lymphocytes.

      Table 1.

      Table 2.

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