Publications by authors named "Miralem Mrkonjic"

10 Publications

  • Page 1 of 1

MLH1 epimutation is a rare mechanism for Lynch syndrome: A case report and review of the literature.

Genes Chromosomes Cancer 2021 May 2. Epub 2021 May 2.

Department of Pathology and Laboratory Medicine, Sinai Health System and University of Toronto, Toronto, Ontario, Canada.

Endometrial carcinoma is one of the prototypical malignancies associated with Lynch syndrome, an inherited cancer syndrome most commonly caused by germline mutations in DNA mismatch repair (MMR) genes, although rare alternative mechanisms also exist. In this report, we describe a patient first diagnosed with colorectal cancer at age 33, then vulvar squamous cell carcinoma, facial sebaceous adenoma/sebaceoma, and finally endometrial carcinoma at age 52. All tumors were MLH1/PMS2-deficient by immunohistochemistry, and MLH1 promoter methylation was identified in the endometrial cancer. Germline MLH1 testing was negative for pathogenic variants, but she was subsequently diagnosed with Lynch syndrome secondary to a germline monoallelic constitutional epimutation of the MLH1 promoter. Identification of patients displaying a Lynch syndrome phenotype but lacking germline MMR mutations is important to avoid delays in the diagnosis of Lynch syndrome as well as the initiation of appropriate cancer screening and genetic counseling.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1002/gcc.22957DOI Listing
May 2021

Review of the current state of digital image analysis in breast pathology.

Breast J 2020 06 27;26(6):1208-1212. Epub 2020 Apr 27.

Sinai Health System, Toronto, ON, Canada.

Advances in digital image analysis have the potential to transform the practice of breast pathology. In the near future, a move to a digital workflow offers improvements in efficiency. Coupled with artificial intelligence (AI), digital pathology can assist pathologist interpretation, automate time-consuming tasks, and discover novel morphologic patterns. Opportunities for digital enhancements abound in breast pathology, from increasing reproducibility in grading and biomarker interpretation, to discovering features that correlate with patient outcome and treatment. Our objective is to review the most recent developments in digital pathology with clear impact to breast pathology practice. Although breast pathologists currently undertake limited adoption of digital methods, the field is rapidly evolving. Care is needed to validate emerging technologies for effective patient care.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1111/tbj.13858DOI Listing
June 2020

Breast specimen handling and reporting in the post-neoadjuvant setting: challenges and advances.

J Clin Pathol 2019 Feb;72(2):120-132

Laboratory Medicine Program, University Health Network, Toronto, Ontario, Canada

Neoadjuvant systemic therapy is becoming more commonly used in patients with earlier stages of breast cancer. To assess tumour response to neoadjuvant chemotherapy, pathological evaluation is the gold standard. Depending on the treatment response, the pathological examination of these specimens can be quite challenging. However, a uniform approach to evaluate post-neoadjuvant-treated breast specimens has been lacking. Furthermore, there is no single universally accepted or endorsed classification system for assessing treatment response in this setting. Recent initiatives have attempted to create a standardised protocol for evaluation of post-neoadjuvant breast specimens. This review outlines the necessary information that should be collected prior to macroscopic examination of these specimens, the recommended and most pragmatic approach to tissue sampling for microscopic examination, describes the macroscopic and microscopic features of post-therapy breast specimens, summarises two commonly used systems for classifying treatment response and outlines the critical variables that should be included in the final pathology report.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1136/jclinpath-2018-205598DOI Listing
February 2019

The dynamic DNA methylation landscape of the shore is altered by -93G>A polymorphism in normal tissues and colorectal cancer.

Clin Epigenetics 2017 9;9:26. Epub 2017 Mar 9.

Lunenfeld-Tanenbaum Research Institute, Sinai Health System, 60 Murray St., Toronto, Ontario M5T 3L9 Canada.

Background: Colorectal cancers (CRCs) undergo distinct genetic and epigenetic alterations. Expression of (), a mismatch repair gene that corrects DNA replication errors, is lost in up to 15% of sporadic tumours due to mutation or, more commonly, due to DNA methylation of its promoter CpG island. A single nucleotide polymorphism (SNP) in the CpG island of (-93G>A or rs1800734) is associated with CpG island hypermethylation and decreased expression in CRC tumours. Further, in peripheral blood mononuclear cell (PBMC) DNA of both CRC cases and non-cancer controls, the variant allele of rs1800734 is associated with hypomethylation at the shore, a region upstream of its CpG island that is less dense in CpG sites

Results: To determine whether this genotype-epigenotype association is present in other tissue types, including colorectal tumours, we assessed DNA methylation in matched normal colorectal tissue, tumour, and PBMC DNA from 349 population-based CRC cases recruited from the Ontario Familial Colorectal Cancer Registry. Using the semi-quantitative real-time PCR-based MethyLight assay, shore methylation was significantly higher in tumour tissue than normal colon or PBMCs ( < 0.01). When shore methylation levels were stratified by SNP genotype, normal colorectal DNA and PBMC DNA were significantly hypomethylated in association with variant SNP genotype ( < 0.05). However, this association was lost in tumour DNA. Among distinct stages of CRC, metastatic stage IV CRC tumours incurred significant hypomethylation compared to stage I-III cases, irrespective of genotype status. Shore methylation of was not associated with MSI status or promoter CpG island hypermethylation, regardless of genotype. To confirm these results, bisulfite sequencing was performed in matched tumour and normal colorectal specimens from six CRC cases, including two cases per genotype (wildtype, heterozygous, and homozygous variant). Bisulfite sequencing results corroborated the methylation patterns found by MethyLight, with significant hypomethylation in normal colorectal tissue of variant SNP allele carriers.

Conclusions: These results indicate that the normal tissue types tested (colorectum and PBMC) experience dynamic genotype-associated epigenetic alterations at the shore, whereas tumour DNA incurs aberrant hypermethylation compared to normal DNA.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1186/s13148-017-0326-6DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5345264PMC
December 2017

MLH1 region polymorphisms show a significant association with CpG island shore methylation in a large cohort of healthy individuals.

PLoS One 2012 11;7(12):e51531. Epub 2012 Dec 11.

Department of Laboratory Medicine and Pathobiology, University of Toronto, Toronto, Ontario, Canada.

Single nucleotide polymorphisms (SNPs) are the most common form of genetic variation. We previously demonstrated that SNPs (rs1800734, rs749072, and rs13098279) in the MLH1 gene region are associated with MLH1 promoter island methylation, loss of MLH1 protein expression, and microsatellite instability (MSI) in colorectal cancer (CRC) patients. Recent studies have identified less CpG-dense "shore" regions flanking many CpG islands. These shores often exhibit distinct methylation profiles between different tissues and matched normal versus tumor cells of patients. To date, most epigenetic studies have focused on somatic methylation events occurring within solid tumors; less is known of the contributions of peripheral blood cell (PBC) methylation to processes such as aging and tumorigenesis. To address whether MLH1 methylation in PBCs is correlated with tumorigenesis we utilized the Illumina 450 K microarrays to measure methylation in PBC DNA of 846 healthy controls and 252 CRC patients from Ontario, Canada. Analysis of a region of chromosome 3p21 spanning the MLH1 locus in healthy controls revealed that a CpG island shore 1 kb upstream of the MLH1 gene exhibits different methylation profiles when stratified by SNP genotypes (rs1800734, rs749072, and rs13098279). Individuals with wild-type genotypes incur significantly higher PBC shore methylation than heterozygous or homozygous variant carriers (p<1.1×10(-6); ANOVA). This trend is also seen in CRC cases (p<0.096; ANOVA). Shore methylation also decreases significantly with increasing age in cases and controls. This is the first study of its kind to integrate PBC methylation at a CpG island shore with SNP genotype status in CRC cases and controls. These results indicate that CpG island shore methylation in PBCs may be influenced by genotype as well as the normal aging process.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0051531PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3519863PMC
May 2013

Promoter methylation of Wnt antagonists DKK1 and SFRP1 is associated with opposing tumor subtypes in two large populations of colorectal cancer patients.

Carcinogenesis 2011 May 8;32(5):741-7. Epub 2011 Feb 8.

Department of Laboratory Medicine and Pathobiology, University of Toronto, Toronto, Ontario, Canada.

Aberrant activation of canonical Wnt signaling is a hallmark event in colorectal carcinogenesis. The Dickkopf-1 (DKK1) and Secreted Frizzled Related Protein 1 (SFRP1) genes encode extracellular inhibitors of Wnt signaling that are frequently silenced by promoter hypermethylation in colorectal cancer (CRC). These methylation events have been identified as prognostic markers of patient outcome and tumor subtype in several cancers but similar roles in CRC have not been comprehensively examined. In CRC, the microsatellite instability (MSI) subtype associates with favorable disease outcome but the molecular events that are responsible remain poorly understood. Consequently, we quantified promoter methylation status of the Wnt antagonist genes DKK1 and SFRP1 in a large population-based cohort of CRCs from Ontario (n = 549) and Newfoundland (n = 696) stratified by MSI status. We examined the association between methylation status and clinicopathological features including tumor MSI status and patient survival. DKK1 and SFRP1 were methylated in 13 and 95% of CRCs, respectively. In Ontario, DKK1 methylation was strongly associated with MSI tumors after adjustment for age, sex and tumor location [odds ratio (OR) = 13.7, 95% confidence interval (CI) = 7.8-24.2, P < 0.001]. Conversely, SFRP1 methylation was inversely associated with MSI tumors after these adjustments (OR = 0.3, 95% CI = 0.1-0.9, P = 0.009). Similar results were obtained in Newfoundland. There were no independent associations with recurrence-free survival. This is the first large study to identify associations between Wnt antagonist promoter hypermethylation and CRC MSI subtype. These events provide insight into subtype-specific epigenetic mediation of Wnt signaling in CRC.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1093/carcin/bgr020DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3140140PMC
May 2011

Functional effects of the MLH1-93G>A polymorphism on MLH1/EPM2AIP1 promoter activity.

Oncol Rep 2011 Mar 30;25(3):809-15. Epub 2010 Dec 30.

Department of Laboratory Medicine and Pathobiology, University of Toronto, Samuel Lunenfeld Research Institute, Mount Sinai Hospital, 1 King's College Circle, Toronto, ON, Canada.

Defective mismatch repair leads to the microsatellite instability (MSI) phenotype of colorectal cancer (CRC). We previously showed that the MLH1-93G>A promoter polymorphism is strongly associated with MSI tumours, suggesting a modifier role for this polymorphism in CRC. The MLH1 promoter is bi-directional with the EPM2AIP1 gene located on the antisense strand. In order to evaluate the functional effects of this polymorphism, we transfected a panel of CRC, endometrial cancer and non-tumourigenic cell lines with MLH1 luciferase promoter constructs. We used constructs in reverse orientation to assess the effect of this polymorphism on EPM2AIP1. The luciferase activities were compared using a two-sided Student's t-test. Electrophoretic mobility shift assays (EMSAs) were used to evaluate whether differential protein binding was responsible for the differences in promoter activity. We observed a higher level of activity with the -93G allele in all the cell lines observed; including the CRC cell line, HCT116 (P=0.002), the endometrial cancer cell line, HEC-1-A (P<0.001) and the normal colonic cell line, CCD-841-CoTr (P=0.002). This polymorphism also affected EPM2AIP1 transcription with the -93A allele demonstrating higher promoter activity in the HCT116 (P=0.007) and HEC-1-A (P=0.004) cells. The EMSA results suggest that this polymorphism alters the affinity of nuclear factors that bind to this region. Our findings indicate that the -93G>A polymorphism modifies the efficiency of MLH1/EPM2AIP1 transcription.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.3892/or.2010.1129DOI Listing
March 2011

Specific variants in the MLH1 gene region may drive DNA methylation, loss of protein expression, and MSI-H colorectal cancer.

PLoS One 2010 Oct 13;5(10):e13314. Epub 2010 Oct 13.

Department of Laboratory Medicine and Pathobiology, University of Toronto, Toronto, Ontario, Canada.

Background: We previously identified an association between a mismatch repair gene, MLH1, promoter SNP (rs1800734) and microsatellite unstable (MSI-H) colorectal cancers (CRCs) in two samples. The current study expanded on this finding as we explored the genetic basis of DNA methylation in this region of chromosome 3. We hypothesized that specific polymorphisms in the MLH1 gene region predispose it to DNA methylation, resulting in the loss of MLH1 gene expression, mismatch-repair function, and consequently to genome-wide microsatellite instability.

Methodology/principal Findings: We first tested our hypothesis in one sample from Ontario (901 cases, 1,097 controls) and replicated major findings in two additional samples from Newfoundland and Labrador (479 cases, 336 controls) and from Seattle (591 cases, 629 controls). Logistic regression was used to test for association between SNPs in the region of MLH1 and CRC, MSI-H CRC, MLH1 gene expression in CRC, and DNA methylation in CRC. The association between rs1800734 and MSI-H CRCs, previously reported in Ontario and Newfoundland, was replicated in the Seattle sample. Two additional SNPs, in strong linkage disequilibrium with rs1800734, showed strong associations with MLH1 promoter methylation, loss of MLH1 protein, and MSI-H CRC in all three samples. The logistic regression model of MSI-H CRC that included MLH1-promoter-methylation status and MLH1 immunohistochemistry status fit most parsimoniously in all three samples combined. When rs1800734 was added to this model, its effect was not statistically significant (P-value  = 0.72 vs. 2.3×10(-4) when the SNP was examined alone).

Conclusions/significance: The observed association of rs1800734 with MSI-H CRC occurs through its effect on the MLH1 promoter methylation, MLH1 IHC deficiency, or both.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0013314PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2954166PMC
October 2010

MSH2 118T>C and MSH6 159C>T promoter polymorphisms and the risk of colorectal cancer.

Carcinogenesis 2007 Dec 17;28(12):2575-80. Epub 2007 Oct 17.

Department of Pathology and Laboratory Medicine, Mount Sinai Hospital, Toronto, Ontario M5T 3L9, Canada.

The most important indicator of colorectal cancer (CRC) risk is the presence of family history of the disease. Inherited genetic changes, such as single nucleotide polymorphisms, in key candidate genes may contribute to CRC risk. We investigated whether promoter polymorphisms in DNA mismatch repair (MMR) genes MSH2 and MSH6 are associated with the risk of CRC. We genotyped 929 CRC patients and 1098 control subjects from Ontario, and 467 patients and 344 controls from Newfoundland and Labrador, for two promoter polymorphisms in the MMR genes MSH2 and MSH6 using the fluorogenic 5' nuclease assay. We used unconditional logistic regression to evaluate the association between each polymorphism and CRC after adjusting for age and sex. The associations between polymorphisms and tumor clinicopathological features were evaluated with a Pearson's chi-squared test or Fisher's exact test. All statistical tests were two sided. We observed strong associations between the MSH2 -118T>C polymorphism and family history of CRC based on the Amsterdam criteria I (P = 0.005) and Amsterdam criteria I and II (P = 0.036) among cases from Ontario. This association was especially evident among female CRC patients in Ontario (for Amsterdam criteria I, and I and II combined, P = 0.003 and P = 0.0001, respectively). The MSH2 -118T>C polymorphism was associated with strong family history of CRC in Ontario patients.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1093/carcin/bgm229DOI Listing
December 2007

MLH1 -93G>A promoter polymorphism and the risk of microsatellite-unstable colorectal cancer.

J Natl Cancer Inst 2007 Mar;99(6):463-74

Department of Pathology, Samuel Lunenfeld Research Institute, Mount Sinai Hospital, Toronto, ON, Canada M5T 3L9.

Background: Although up to 30% of patients with colorectal cancer have a positive family history of colorectal neoplasia, few colorectal cancers can be explained by mutations in high-penetrance genes. We investigated whether polymorphisms in DNA mismatch repair genes are associated with the risk of colorectal cancer.

Methods: We genotyped 929 case patients and 1098 control subjects from Ontario and 430 case patients and 275 control subjects from Newfoundland and Labrador for five polymorphisms in the mismatch repair genes MLH1 and MSH2 with the fluorogenic 5' nuclease assay. Tumor microsatellite instability (MSI) was determined with a polymerase chain reaction-based method; MSI status was assigned as high (MSI-H, > or = 30% unstable markers among all markers tested), low (MSI-L, <30% markers unstable), or stable (MSS, no unstable markers). We used unconditional logistic regression to evaluate the association between each polymorphism and colorectal cancer after adjusting for age and sex. The associations between polymorphisms and tumor clinicopathologic features were evaluated with a Pearson's chi-square or Fisher's exact test. All statistical tests were two-sided.

Results: We observed strong associations between the MLH1 -93G>A polymorphism and MSI-H tumors among case patients from Ontario (P = .001) and Newfoundland (P = .003). When compared with the control populations, homozygosity for the MLH1 -93G>A variant allele was associated with MSI-H tumors among case patients in Ontario (adjusted odds ratio [OR] = 3.23, 95% confidence interval [CI] = 1.65 to 6.30) and in Newfoundland (OR = 8.88, 95% CI = 2.33 to 33.9), as was heterozygosity among case patients in Ontario (OR = 1.84, 95% CI = 1.20 to 2.83) and in Newfoundland (OR = 2.56, 95% CI = 1.14 to 5.75). Genotype frequencies were similar among case patients with MSS and MSI-L tumors and control subjects, and the majority of homozygous variant carriers had MSS tumors. Among case patients from Ontario, an association between the MLH1 -93G>A polymorphism and a strong family history of colorectal cancer (for Amsterdam criteria I and II, P = .004 and P = .02, respectively) was observed.

Conclusion: In two patient populations, the MLH1 -93G>A polymorphism was associated with an increased risk of MSI-H colorectal cancer.
View Article and Find Full Text PDF

Download full-text PDF

Source
http://dx.doi.org/10.1093/jnci/djk095DOI Listing
March 2007