Microsatellite Instability Testing in Colorectal Carcinoma: Choice of Markers Affects Sensitivity of Detection of Mismatch Repair-Deficient Tumors (original) (raw)
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Bulletin du Cancer, 2019
Microsatellite instability (MSI), which is caused by deficiency of the DNA MisMatch repair (MMR) system, is the molecular abnormality observed in tumors associated with Lynch syndrome (LS). LS represents one of the most frequent conditions of cancer predisposition in human, thus requiring specific care and genetic counseling. Moreover, research has recently increasingly focused on MMR deficiency due to its positive predictive value for the efficacy of immune checkpoints inhibitors (ICKi) in metastatic tumors, regardless of their primary origin. MSI has also been demonstrated to constitute an independent prognostic factor in several tumor types, being also associated with alternative response to chemotherapy. These observations have led many professional medical organizations to recommend universal screening of all newly diagnosed colorectal cancers (CRC) for dMMR/MSI status and increasing evidence support the evaluation of MSI in all human tumors regardless of the cancer tissue of origin. Currently, two standard reference methods, namely immunohistochemistry and PCR, are recommended for the detection of dMMR/MSI status. These methods are equally valid as the initial screening test for dMMR/MSI in CRC. To date, there is no recommendation for the detection of dMMR/MSI in other primary tumors. In this review, we will present a comprehensive overview of the methods used for evaluation of tumor dMMR/MSI status in CRC, as well as in other tumor sites. We will see that the evaluation of this status remains challenging in some clinical settings, with the need to improve the above methods in these specific contexts. RESUME L'instabilité des microsatellites (MSI), due à la déficience du système MMR (pour MisMatch Repair), est l'anomalie moléculaire observée dans les tumeurs du syndrome de Lynch. Le syndrome de Lynch représente le plus fréquent des syndromes de prédisposition aux cancers, ce qui nécessite une prise en charge spécifique et un avis du conseil génétique. Par ailleurs, la recherche s'est récemment beaucoup intéressée au statut MSI en raison de sa valeur prédictive quant à l'efficacité des inhibiteurs des immune checkpoints (ICKi) dans les tumeurs à un stade métastatique, quelle que soit leur origine. Le statut MSI constitue également un facteur pronostique indépendant dans plusieurs types tumoraux, du fait de son association à des réponses alternatives à la chimiothérapie. Ces données ont conduit un grand nombre de sociétés savantes à recommander un screening universel pour la détermination du statut dMMR/MSI de tout cancer colorectal (CCR), de même que de nombreux autres types tumoraux. Deux méthodes de référence, qui sont l'immunohistochimie et la biologie moléculaire, sont recommandées pour la détermination du statut dMMR/MSI. Ces deux méthodes sont équivalentes pour le dépistage de ce statut dans le CCR. En revanche, il n'y a pas de recommandation dans d'autres sites tumoraux. Nous ferons un état des lieux des méthodes à disposition pour l'évaluation du statut dMMR/MSI, à la fois dans le CCR, mais également dans d'autres sites tumoraux. Nous verrons que l'évaluation de ce statut reste difficile dans certains contextes cliniques, avec la nécessité d'améliorer en conséquence les méthodes de détection dans ces contextes particuliers.
Annals of Surgical Oncology, 2010
Background In colorectal cancer (CRC), microsatellite instability (MSI) is a valuable marker of defective DNA mismatch repair that identifies cancers with distinct phenotypic properties, including favorable survival. However, the optimal assay for MSI status is unknown. We have evaluated a simplified 3-marker assay for MSI and compared it with the 5-marker (NCI) assay to see if technical variations in MSI testing are important. Materials and Methods DNA samples from 357 CRCs were evaluated for MSI using the 5 microsatellite markers recommended for the NCI assay (BAT 25, BAT26, D2S123, D5S346, and D17S250). Results were compared with a simplified 3-marker assay (BAT25, BAT26, and D2S123). CRCs identified as MSI were evaluated for their clinical, pathological, and genetic characteristics. Results The 5-marker assay identified 96 cancers as MSI. Only 56 of these were MSI by the 3-marker assay (3-marker+ group), leaving 40 cases identified as MSI only by NCI criteria (3-marker− group). The remaining 261 cancers were microsatellite stable (MSS). The 3-marker+ MSI tumors had features characteristic of MSI tumors: more proximal, poorly differentiated, associated with hereditary nonpolyposis colorectal cancer (HNPCC), more BRAF mutations, fewer KRAS mutations, better 5-year disease-specific survival, more frequent mismatch repair (MMR) protein loss, and less likely to be metastatic on presentation (P < .05). Chromosomal arm loss was observed only in 3-marker− MSI and MSS cancers (P < .05). Conclusion The 3-marker MSI assay outperforms the traditional 5-marker assay for identifying patients with favorable prognosis and homogeneous clinical and genetic features. More accurate MSI testing should improve prognostic and predictive scoring systems for colorectal cancer.
Oncology Letters, 2013
Microsatellite instability (MSI) is a mutator phenotype that results from a defective mismatch repair (MMR) pathway. The present study examined the incidence of MSI and loss of heterozygosity (LOH) according to five markers from the panel of the National Cancer Institute (NCI) in 38 colorectal cancer (CRC) patients from the United Arab Emirates (UAE). MSI and LOH were analyzed using fragment analyses in a multiplex PCR setting on a capillary array electrophoresis platform. The expression of the MMR proteins, hMLH1 and hMSH2, was analyzed using immunohistochemistry. The cohort consisted of 17 females (44.7%) and 21 males (55.3%) with mean ages of 59.9 and 63.3 years, respectively. The overall MSI incidence was 31.3% (95% CI, 16.1-50.0), and included three patients with high MSI (MSI-H; 9.4%; 95% CI, 2.0-25.0) and seven patients with low MSI (MSI-L; 21.9%; 95% CI, 10.7-39). LOH was detected in three patients, while the remaining 25 patients (65.8%) showed no instability and were therefore classified as microsatellite stable (MSS). MSI was detected in the following screened markers: Bat25 in seven patients, Bat26 in three patients, adenomatous polyposis coli (APC; D5S346) in five patients, AFM093xh3 (D2S123) in two patients and Mfd15 (D17S250) in three patients. Of the five MSI-positive patients, four (80%) were evidently younger, aged 38, 48, 49 and 59 years, respectively. The MSI-H incidence (9.4%) was lower compared with that of other ethnic groups. In terms of the MMR proteins, hMLH1 expression was deficient in seven patients, of whom three were MSI-H patients, and hMSH2 was deficient in three patients. Fisher's exact test showed significant associations between hMLH1 and MSI when classified as MSS, MSI-L or MSI-H (P=0.0003). No such association was observed with abnormal MMR protein expression, age, cancer stage or gender.
ESMO open, 2021
Background: DNA mismatch repair system deficiency (dMMR) is found in 15% of colorectal cancers (CRCs). Two methods are used to determine dMMR, immunohistochemistry (IHC) of MMR proteins and molecular testing of microsatellite instability (MSI). Only studies with a low number of patients have reported rates of discordance between these two methods, ranging from 1% to 10%. Materials and methods: Overall, 3228 consecutive patients with CRCs from two centers were included. Molecular testing was carried out using the Pentaplex panel and IHC evaluated four (MLH1, MSH2, MSH6, and PMS2; cohort 1; n ¼ 1085) or two MMR proteins (MLH1 and MSH2; cohort 2; n ¼ 2143). The primary endpoint was the rate of discordance between MSI and MMR IHC tests. Results: Fifty-one discordant cases (1.6%) were initially observed. Twenty-nine out of 51 discordant cases were related to IHC misclassifications. In cohort 1, after re-reading IHC and/or carrying out new IHC, 16 discordant cases were reclassified as nondiscordant. In cohort 2, after the addition of MSH6/PMS2 IHC and re-examination, 13 were reclassified as nondiscordant. In addition, 10 misclassifications of molecular tests were identified. Finally, only 12 discordant cases (0.4%) remained: 5 were proficient MMR/MSI and 7 were dMMR/microsatellite stable. Conclusions: Our study confirmed the high degree of concordance between MSI and MMR IHC tests. Discordant cases must be reviewed, and if needed, tests must be repeated and analyzed by an expert team.
Effect of microsatellite instability on histopathological parameters and prognosis in colon cancers
Cukurova Medical Journal, 2023
The aim of this study was to compare the clinicopathological features, while evaluating the frequency of MSI, and the survival rates in these patients in our patient group with MSI and microsatellite stabil (MSS) colorectal carcinomas (CRCs). Materials and Methods: We retrospectively enrolled 146 patients who underwent colon resection between the years of 2014-2022. The expression of MSI status was evaluated by immunohistochemistry. The association of MSI status, presense of tumor-infiltrating lymphocytes (TILs), and tumor budding score with a patient's survival was assessed by the Kaplan-Meier method and Cox regression analysis. Results: There were 104 (71.2%) MSS cases and, 42 (28.8%) MSI cases. 15 (10.3%) MSI tumors were found to be MSI-low, and 27 (18.5%) to be MSI-high. MSI tumors were significantly associated with younger patients (<50), earlier stage (T1-T2), right localization, lower rate of lymph node metastasis, presence of mucinous component and TILs response. The Cox-regression model revealed TILs, tumor budding score, and MSI are variables that significantly affect survival. The presence of TILs exhibited a protective effect (Hazard ratio (HR)=0.446), which decreased the mortality risk by 2.24 times for the follow-up period, while the presence of high TBS increased the risk of mortality by HR=3.22. Conclusion: This study revealed that patients with MSI CRCs may show unique clinicopathological features and should be evaluated using some guiding parameters that will improve survival. Amaç: Bu çalışmada, MSİ ile mikrosatellit stabil (MSS) kolorektal karsinomlu (KRK) hasta grubumuzda, klinikopatolojik özellikleri karşılaştırmayı, MSİ sıklığını ve bu hastalarda sağkalım oranlarını değerlendirmeyi amaçladık Gereç ve Yöntem: Çalışmaya retrospektif olarak 2014-2022 yılları arasında kolon rezeksiyonu yapılan 146 hasta dahil edildi. MSI durumu, immünohistokimyal belirteçlerle değerlendirildi. MSI durumu, tümörü infiltre eden lenfositlerin (TIL'ler) varlığı ve tümör tomurcuklanma skorunun sağkalım ile olan ilişkisi, Kaplan-Meier yöntemi ve Cox regresyon analizi ile değerlendirildi. Bulgular: Olgularımızın 104'ü (%71,2) MSS, 42'si (%28,8) MSİ'dir. MSI tümörlerin 15'i (%10,3) MSI-düşük, 27'si (%18,5) MSİ-yüksek olarak bulundu. MSİ tümörler, genç yaş (<50 yaş), erken evre (T1-T2), sağ kolon lokalizasyonu, düşük lenf nodu metastazı oranı, müsinöz komponent ve TİL varlığı ile istatistiksel olarak anlamlı şekilde ilişkiliydi. Cox-regresyon modeli, TIL'lerin, tümör tomurcuklanma skorunun ve MSİ'nin sağkalımı önemli ölçüde etkileyen değişkenler olduğunu ortaya çıkardı. TIL'lerin varlığı, takip süresince ölüm riskini 2,24 kat azaltan koruyucu bir etki (Tehlike oranı (HR)=0,446) sergilerken, yüksek tümör tomurcuklanma skorunun ölüm riskini (HR=3,22) artırdığı bulundu. Sonuç: Çalışmamızda, MSİ KRK'li hastaların, kendilerine özgü klinikopatolojik özellikler gösterebileceği ve sağkalımda yol gösterici bazı parametrelerle birlikte değerlendirilmesi gerektiği vurgulanmaktadır.
Cancer biomarkers : section A of Disease markers, 2010
Tumour microsatellite instability (MSI) is useful in identifying patients with hereditary non-polyposis colorectal cancer (HNPCC) with defective DNA mismatch repair (MMR) genes. A reference Bethesda panel has limitations resulting from the inclusion of dinucleotide markers, which are less sensitive and specific for detection of tumours with MMR deficiencies. We developed a multiplex PCR assay with additional four mononucleotide markers and one dinucleotide marker (NR-21, NR-24, BAT-40, TGF-BetaR and D18S58) for a rapid and proper classification of MSI-H, MSI-L and MSS colorectal cancers. Two tetranucleotide markers were added to identify sample mix-ups and/or contamination. all the 44 cases test cases were in agreement with previous classification except for three cases: one case MSI-H-Bethesda unstable only for dinucleotides markers shifted to MSI-L category and two cases MSI-L-Bethesda unstable for mononucleotide markers shifted to MSI-H category. Immunohistochemistry analysis rev...
Pathologica, 2020
The American Society for Clinical Pathology (ASCP), College of American Pathologists (CAP), Association for Molecular Pathology (AMP), and the American Society of Clinical Oncology (ASCO) have been recently strongly recommended the evaluation of mismatch repair status (MMS) as molecular biomarkers in colorectal cancer for a better prognostic stratification of patients. This recommendation is emphasized by the recent evidence of Microsatellite Instability (MSI) as a predictive marker for chemotherapy and immunotherapy. In this scenario, the validation of molecular biomarker testing methods seems to be essential to design the most appropriate tailored therapy and the most suitable care strategy, respectively. In this study, we validated an alternative method based on capillary electrophoresis system label-free PCR (Qiaxcel system) to evaluate the MSI Bethesda Panel. We also parallel the results with a standard approach. Our data showed total concordance with the standard approach, with a highly time-efficient and easy procedure combined with high sensitivity for MSI detection. Alternative capillary electrophoresis based on label-free PCR such as the Qiaxel system is a very sensitive and specific method to detect MSI for the management of patients with colorectal cancer. This procedure is adequate and suitable in diagnostic routine for the evaluation of microsatellite repeats compared to standard procedures.
Causes of microsatellite instability in colorectal tumors
Cancer Genetics and Cytogenetics, 2001
Microsatellite instability (MSI) analysis was performed using a "reference panel" of microsatellite markers in 345 unselected primary colorectal cancers (CRC). Thirty-five (10%) tumors were classified as high MSI (MSI-H). We identified 6 (17%) MSI-H tumors with germline mutations in mismatch repair (MMR) genes (tumors from patients with hereditary non-polyposis colorectal cancer (HNPCC) syndrome) and 29 (83%) MSI-H tumors without germline MMR mutations (sporadic MSI-H tumors). Hypermethylation of the hMLH1 promoter was found in 26/29 (90%) sporadic MSI-H tumors but only in 1/6 (17%) HNPCC tumors ( P Ͻ .001). Somatic alterations were identified in both MMR genes in HNPCC tumors but mainly in the hMSH2 gene in sporadic MSI-H tumors. LOH at MMR loci was detected in 3/6 (50%) HNPCC tumors and in 4/26 (15%) informative sporadic MSI-H tumors. These results together indicate different mode of inactivation of MMR genes in sporadic MSI-H tumors versus MSI-H tumors in HNPCC patients. We therefore propose that MSI analysis of newly diagnosed primary CRC followed by methylation analysis of hMLH1 promoter in MSI-H tumors and mutational analysis of MMR genes in MSI-H tumors lacking hMLH1 promoter methylation might be an efficient molecular genetic approach for HNPCC screening.