Prospective evaluation of DNA mismatch repair protein expression in primary endometrial cancer

doi:10.1016/j.ygyno.2009.05.026

Gynecologic Oncology

Volume 114, Issue 3, September 2009, Pages 486-490

https://doi.org/10.1016/j.ygyno.2009.05.026 Get rights and content

Abstract

Objectives

Immunohistochemical (IHC) stains for mismatch repair (MMR) proteins help screen for Lynch syndrome and identify microsatellite unstable colorectal carcinomas, providing prognostic information. It has been suggested that colorectal and endometrial carcinomas should be screened routinely for a MMR defect, but data are lacking on the practical application of this policy. We report our experience with the prospective evaluation of MMR protein expression in endometrial cancer.

Methods

All cases of primary endometrial cancer at a single institution regardless of age, family history or histologic features were prospectively stained for the MMR proteins MLH1, MSH2, MSH6 and PMS2. Clinical and pathologic correlates were collected from the medical record.

Results

A total of 140 endometrial cancer cases were studied. Over 90% of cases were of endometrioid histology. 119 patients had stage I/II disease, and 21 stage III/IV. Nineteen percent of patients were < age 50. Overall, there was loss of 1 or more MMR proteins in 30 patients (21%), including MLH1 and PMS2 in 24, MSH2 and MSH6 in 4, and MSH6 in 2 patients. None of the patients met clinical criteria for Lynch syndrome. However, using MMR protein expression, age and family history, 11% of patients were referred for genetic counseling. Of these patients, three (20%) scheduled an appointment: one canceled and two tested negative.

Conclusions

Prospective staining for MMR proteins is feasible and allows for primary triage for the evaluation of Lynch syndrome in women with endometrial cancer. However, acceptance of genetic consultation and testing is surprisingly low and deserves further investigation.

Introduction

Lynch syndrome, also known as hereditary non-polyposis colorectal cancer (HNPCC) syndrome, is the most common hereditary colorectal cancer syndrome in Western countries, accounting for 2 to 5% of all colorectal cancers [1], [2]. Lynch syndrome is an autosomal dominant syndrome, caused by inactivating mutations of the DNA mismatch repair (MMR) genes, mostly MSH2, MLH1, and MSH6. MMR genes behave like tumor suppressor genes in that an inherited or acquired mutation in an MMR gene predisposes to cancer.

Patients with Lynch syndrome are at increased risk of developing colorectal, endometrial, gastric, ovarian, small bowel, and urinary tract cancers. A recent study by Hampel et al. [3] showed that approximately 2% of newly diagnosed unselected endometrial cancer patients have Lynch syndrome. Women with Lynch syndrome have a 50 to 85% lifetime risk of developing colorectal cancer and a 40 to 60% lifetime risk of developing endometrial cancer [4], [5].

Screening for Lynch syndrome is traditionally done based on family history using the clinical Amsterdam and Bethesda criteria. However, not all patients with Lynch syndrome meet the established clinical criteria, and not all women meeting the clinical criteria have Lynch syndrome [6], [7]. Thus, alternative or adjunctive methods to screen for Lynch syndrome are imperative. Another screening method is to test tumor tissue for microsatellite instability, which results from defective mismatch repair systems and can be found in > 90% of colon and endometrial cancers associated with Lynch syndrome [8]. Patients with tumors that show microsatellite instability are then referred for further DNA testing. Another way of determining whether the mismatch repair genes are functioning properly is testing the tumor for the presence of mismatch repair proteins by immunohistochemistry [9], [10]. If one or more of the MMR proteins is absent on IHC, then the abnormal protein expression could be caused by an inherited or acquired mutation in one of the MMR genes or by methylation of the genes causing absent protein expression. Lack of protein expression can then be confirmed with genetic testing. Immunohistochemistry is an easy, fast, and inexpensive way of screening that can be done at time of initial pathology evaluation. It has been suggested that CRC and endometrial carcinomas should be screened routinely for a MMR defect, but data are lacking on the practical application of this policy. We report our experience with the prospective evaluation of MMR protein expression in endometrial cancer.

Section snippets

Patients

Beginning April 2007, all patients undergoing hysterectomy as part of their primary surgical management for endometrial adenocarcinoma with tumor in the hysterectomy specimen at The Ohio State University Medical Center and Arthur G. James Cancer Hospital were identified. Endometrial tumors from these patients were prospectively stained for the MMR proteins MLH1, MSH2, MSH6 and PMS2, regardless of age, family history or histologic features.

Immunohistochemistry

Paraffin-embedded tissue was cut at 4 μm and placed on

Results

A total of 140 cases of primary endometrial cancer were studied, and are described in Table 1. The mean age of diagnosis was 60.5 years (range 30 to 91). Twenty-seven patients (19%) were ≤ 50 years at diagnosis, and of those 22% had abnormal protein expression. Ninety percent of cases were endometrioid adenocarcinomas, 62% of these were grade 1, and over 80% were stage I or II.

Overall, there was loss of expression of one or more MMR proteins in 30 patients (21%), with 24/30 of these cases

Discussion

Although none of the patients in this study met clinical criteria for Lynch syndrome, approximately 10% of all endometrial cancer patients should be referred for genetic counseling using the screening and triage method described in this study (Fig. 1). Prospective screening for Lynch syndrome using IHC followed by genetic counseling for selective patients with abnormal protein expression is important to detect the estimated 2% of endometrial cancer patients with Lynch syndrome since these

Conflict of interest statement

All authors declare that there are no conflicts of interest, except Heather Hampel who has received an honorarium from Myriad Genetics Laboratories Inc for an advisory board meeting and an honorarium from Falco Biosystems for giving a lecture.

Acknowledgments

We would like to thank Leigha Senter for her help with contacting patients for follow up.

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The prognostic implications of DNA mismatch repair protein (MMRP) have not been determined in endometrial cancer. Therefore, in this study, we aimed to evaluate the clinicopathologic characteristics of DNA MMRP deficiency in endometrial cancer.
We examined the MMRP status of 206 patients with endometrial carcinomas, using immunohistochemistry, and analyzed their clinicopathologic factors and survival outcomes stratified by MMRP status using the Kaplan–Meier method and Cox regression analysis.
Forty-three cases were deficient for at least one MMRP (20.9%). Loss of MLH1 was the most common (13.1%), followed by MSH6 (7.8%). MMRP deficiency was significantly associated with lympho-vascular space invasion, deep myometrial invasion, and adjuvant treatment (P = 0.032, 0.041, and 0.047, respectively). MMRP-deficient patients had a better overall survival (OS), particularly at advanced cancer stages (III/IV) (100% vs. 73.7%, P = 0.170) or if they had received adjuvant treatment (100% vs. 86.7%, P = 0.087).
Although MMRP deficiency was associated with unfavorable prognostic risk factors in endometrial cancer, we found a trend in favor of OS in MMRP-deficient patients. More studies are needed to confirm its prognostic implication.
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Citation Excerpt :
MLH1 promoter methylation testing was conclusive in >98% of MLH1/PMS2-deficient cases (Table 2). The proportion of MMRd endometrial cancers (25.9%) in our study (Table 1) is similar to that in previous reports,9–12,14–17 and MMRd cases were present throughout all grades of endometrioid adenocarcinoma as well as high-grade carcinomas (Table 3). MMRd endometrial carcinoma is 1 of the 4 molecular subgroups of endometrial carcinoma22–24 and has an intermediate recurrence-free survival.24,25
Universal screening of endometrial cancer for underlying Lynch syndrome (LS) using DNA mismatch repair immunohistochemistry (MMR IHC) has been recommended. The objective of this study was to assess the feasibility and outcomes of using office endometrial samplings in a community LS screening program.
A community laboratory adopted Cancer Care Ontario's LS screening recommendations. All new endometrial cancers in women aged <70 years were screened for LS using MMR IHC and MLH1 promoter methylation testing cascade for MLH1/PMS2-deficient cases. This retrospective validation study analyzes the first year's results.
Of 693 new endometrial cancers, 467 (67.4%) were eligible for LS screening. Both MMR IHC and MLH1 promoter methylation testing were conclusive in >98% of cases. MMR deficiency (MMRd), which includes LS screen-positive cases, was identified in 25.9% of patients (121/467). LS screen-positive tumours comprised 5.9% (27/467) of all cases.
Endometrial samplings from community practice are suitable for pre-operative LS screening. This testing can identify MMRd endometrial cancers with significant prognostic implications. Approximately 1 in 20 Ontario women <70 years of age with endometrial cancer screen positive for LS. Pre-operative and/or operative assessment for co-existent colonic neoplasms needs to be considered in this high-risk group. In addition, these women should be referred to genetic counselling.
Le dépistage systématique du cancer de l'endomètre est recommandé pour détecter un syndrome de Lynch (SL) sous-jacent en effectuant l'analyse immunohistochimique des protéines de réparation des mésappariements de l'ADN (IHC des MMR). Cette étude visait à évaluer la faisabilité et les résultats du prélèvement endométrial effectué en cabinet dans un programme communautaire de dépistage du SL.
Un laboratoire communautaire a adopté les recommandations d'Action cancer Ontario pour le dépistage du SL. Tous les nouveaux cas de cancer de l'endomètre chez des femmes de moins de 70 ans ont fait l'objet d'un dépistage du SL par IHC des MMR et de dépistages en cascade pour la méthylation du promoteur de MLH1 dans les cas de perte d'expression de MLH1 et de PMS2. Cette étude de validation rétrospective analyse les résultats de la première année.
Des 693 nouveaux cancers de l'endomètre, 467 (67,4 %) étaient admissibles au dépistage du SL. L'IHC des MMR et les dépistages pour la méthylation du promoteur de MLH1 étaient concluants dans plus de 98 % des cas. Une déficience du système MMR a été observée chez 25,9 % des patientes (121/467), y compris celles ayant obtenu un résultat positif au dépistage du SL. Les tumeurs positives au dépistage du SL représentaient 5,9 % (27/467) de tous les cas.
Le prélèvement endométrial effectué en pratique communautaire convient au dépistage préopératoire du SL. Ce dépistage peut détecter les cancers de l'endomètre avec déficience du système MMT, ce qui a d'importantes conséquences sur le pronostic. En Ontario, chez les femmes de moins de 70 ans atteintes d'un cancer de l'endomètre, environ 1 cas sur 20 se révèle positif au dépistage du SL. Il faut envisager l’évaluation préopératoire et/ou peropératoire pour détecter les tumeurs coliques concomitantes chez ce groupe à risque élevé. De plus, il y a lieu d'orienter ces femmes vers une consultation génétique.
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Findings on impact of mismatch repair deficiency (MMRd) on patient outcomes in endometrial cancer (EC) have been inconsistent to date. The objective of this study was to compare the oncologic outcomes and recurrence patterns between MMRd and MMR-intact (MMRi) endometrioid EC (EEC).
Between 2015 and 2018, we prospectively recruited 492 EEC cases from three cancer centers in Ontario, Canada. Tumors were reflexively assessed for MMR protein expression by immunohistochemistry (IHC). Clinicopathological, survival and recurrence patterns were compared between MMRd and MMRi cases.
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During the study, there were 2045 endometrial cancers: 1399 in the physician-order group and 646 in the universal testing group. In the physician-order group: among women < age 60, 34% underwent IHC; 9.6% were abnormal, and 3% were possible LS after methylation testing; among women ≥60, 11% underwent IHC, 3% were abnormal and <1% were possible LS. In the universal group, 87% of women age <60 had IHC, 19.4% were abnormal, and 6% were possible LS; Among women age ≥60, 82% underwent IHC, 26% were abnormal, and 2% were possible LS. There were no differences in LS cases between the physician-order group and the universal group in either age strata (<60: 3% vs. 3.6%, p=0.62; ≥60: <1% vs. 1%, p=0.63) Factors associated with LS were younger age (odds ratio (OR) 0.11, 95% confidence interval (CI) 0.04–0.29) and lower body mass index (BMI), (OR 0.38 95% CI 0.18–0.80).
Universal IHC screening did not result in increased LS detection in EC.
MMR deficiency in urothelial carcinoma of the bladder presents with temporal and spatial homogeneity throughout the tumor mass
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Microsatellite instability (MSI), a hypermutator phenotype described in many cancers, has emerged as a predictive biomarker for immune checkpoint inhibitor therapy. Cancer heterogeneity represents a potential obstacle for the analysis of predicitive biomarkers. MSI has been reported in bladder cancer, but data on the possible extent of intratumoral heterogeneity are lacking.
To study MSI heterogeneity in bladder cancer, a tissue microarray (TMA) comprising 598 muscle-invasive urothelial carcinomas of the bladder was utilized to screen for MSI by immunhistochemistry with antibodies for MLH1, PMS2, MSH2, and MSH6.
In 9 cases suspicious for MSI, MMR status was further evaluated by large section examination and polymerase chain reaction (PCR)-based analysis of microsatellites (“Bethesda panel”) resulting in the identification of 5 validated MSI cases from 448 interpretable cancers (prevalence 1.1%). MMR deficiency always involved PMS2 loss, in 3 cases with additional loss or reduction of MLH1 expression. Four cancers were MSI-high and 1 was MSI-low in the PCR analysis. Parallel sequencing revealed an inactivating MLH1 mutation in 1 tumor but no further known pathogenic MMR gene mutations were found. Immunostaining of all available 72 cancer-containing tissue blocks of the 5 confirmed bladder cancer with MSI including prior and subsequent biopsies showed complete homogeneity of the MMR protein defects and the status of the 4 MMR proteins did not markedly change in sequential resections. In all 4 cases with noninvasive precursor lesions, MSI was also detectable.
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^☆: This study was presented in abstract and poster form at the Society of Gynecologic Oncology Annual Meeting in Tampa Florida in March 2008.

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Prospective evaluation of DNA mismatch repair protein expression in primary endometrial cancer☆

Abstract

Objectives

Methods

Results

Conclusions

Introduction

Section snippets

Patients

Immunohistochemistry

Results

Discussion

Conflict of interest statement

Acknowledgments

Gastroenterology

Gastroenterology

Gastroenterology

J. Mol. Diagn.

Genetic susceptibility for non-polyposis colorectal cancer

J. Med. Genet.

Screening for the Lynch syndrome (hereditary nonpolyposis colorectal cancer

N. Engl. J. Med.

Screening for Lynch syndrome (hereditary nonpolyposis colorectal cancer) among endometrial cancer patients

Cancer Res.

Cancer risk in mutation carriers of DNA-mismatch-repair genes

Int. J. Cancer

Clinical findings with implications for genetic testing in families with clustering of colorectal cancer

N. Engl. J. Med.

Sensitivity and specificity of clinical criteria for hereditary nonpolyposis colorectal cancer associated mutations in MSH2 and MLH1

J. Med. Genet.