MPhil Molecular Pathology and Genomics, Department of Biological Sciences, Forman Christian College, Lahore, Pakistan.
Consultant Molecular Pathologist, Agha Khan University Hospital Lahore Stat Lab, Lahore, Pakistan.
Asian Pac J Cancer Prev. 2020 Aug 1;21(8):2395-2401. doi: 10.31557/APJCP.2020.21.8.2395.
The purpose of our study was to determine the frequency of BRCA1 promoter hypermethylation and its association with expression changes of BRCA1 and main morphological features in sporadic breast cancer.
A retrospective review of cases was performed to select those with specific morphological features suggestive of breast cancer. BRCA1 promoter hypermethylation and changes in protein expression were evaluated in 30 cancerous and 30 non-cancerous tissue samples. A tissue microarray containing samples from normal and tumor tissue was prepared and stained for BRCA1 protein expression using a commercially available monoclonal antibody against BRCA1 (Ab-1) clone MS110 (mAb). DNA was extracted using modified protocol of Qiagen minikit. DNA was modified using a Bisulfite conversion kit and BRCA1 hypermethylation was detected using a methylation specific PCR.
Promoter hypermethylation was negative in 30 non-cancerous samples with retained BRCA1 protein expression. Methylation was positive in 82.6% (n=19/23) of the sporadic cancer samples that had loss of BRCA1 expression and 50% (n=2/4) of the samples with equivocal protein expression. Methylation was negative in all the sporadic breast cancer samples (n=3/3) with retained protein expression. Chi-square analysis showed significant association of BRCA1 promoter methylation with decreased protein expression (P=0.016) and co-existence of loss of BRCA1 and Her2neu at chromosome 17 (P=0.026) respectively. There was no significant association of BRCA1 methylation with morphological features excluding necrosis (P=0.035). Promoter hypermethylation was found to be most common (68.75%) among Triple Negative Breast Cancer (TNBC) females less than 45 years old.
Our study suggests that BRCA1 promoter hypermethylation has significant contribution in sporadic breast carcinogenesis. This was our preliminary study in Pakistan. Further studies aimed to determine the in-depth mechanisms of BRCA1 epigenetics in TNBC. BRCAness enriched phenotype in TNBC might be used as a biomarker for the exploitation of therapeutic and clinical implications.
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本研究旨在确定 BRCA1 启动子超甲基化的频率及其与散发性乳腺癌中 BRCA1 的表达变化和主要形态特征的关系。
方法：对病例进行回顾性分析，选择具有提示乳腺癌特定形态特征的病例。评估 30 例癌组织和 30 例非癌组织样本中 BRCA1 启动子超甲基化和蛋白表达的变化。制备包含正常和肿瘤组织样本的组织微阵列，并使用针对 BRCA1（Ab-1）克隆 MS110（mAb）的商业可得单克隆抗体对 BRCA1 蛋白表达进行染色。使用 Qiagen minikit 的改良方案提取 DNA。使用亚硫酸氢盐转化试剂盒对 DNA 进行修饰，并使用甲基化特异性 PCR 检测 BRCA1 超甲基化。
结果：保留 BRCA1 蛋白表达的 30 例非癌组织中，启动子超甲基化为阴性。在 23 例 BRCA1 表达缺失的散发性癌样本中，82.6%（n=19/23）的样本呈甲基化阳性，在 4 例蛋白表达不确定的样本中，50%（n=2/4）的样本呈甲基化阳性。所有保留蛋白表达的散发性乳腺癌样本（n=3/3）均呈甲基化阴性。卡方分析显示 BRCA1 启动子甲基化与蛋白表达降低（P=0.016）和 17 号染色体上 BRCA1 和 Her2neu 丢失的共存（P=0.026）显著相关。BRCA1 甲基化与排除坏死的形态特征无显著相关性（P=0.035）。BRCA1 启动子超甲基化在 45 岁以下的三阴性乳腺癌（TNBC）女性中最为常见（68.75%）。
结论：本研究表明 BRCA1 启动子超甲基化在散发性乳腺癌发生中具有重要作用。这是我们在巴基斯坦的初步研究。进一步的研究旨在确定 BRCA1 表观遗传学在 TNBC 中的深入机制。TNBC 中 BRCAness 富集表型可作为探索治疗和临床意义的生物标志物。
