Jeronimo Carmen, Monteiro Paula, Henrique Rui, Dinis-Ribeiro Mário, Costa Isabel, Costa Vera L, Filipe Luísa, Carvalho André L, Hoque Mohammad O, Pais Irene, Leal Conceição, Teixeira Manuel R, Sidransky David
Department of Genetics, Portuguese Oncology Institute, Rua Dr. Antonio Bernardino Almeida, Porto, Portugal.
Breast Cancer Res Treat. 2008 May;109(1):27-34. doi: 10.1007/s10549-007-9620-x. Epub 2007 Jun 5.
We hypothesized that comprehensive breast cancer methylation profiling might provide biomarkers for diagnostic assessment of suspicious breast lesions using fine needle aspiration biopsy (FNA).
Twenty-three gene promoters were surveyed by quantitative methylation-specific PCR in bisulfite-modified DNA from 66 breast carcinomas (BCa), 31 fibroadenomas (FB) and 12 normal breast (NT) samples to define a set of genes differentially methylated in malignant and non-malignant tissues. This set was tested in 78 FNA washings obtained pre-operatively (66 malignant, 12 benign), with histopathological diagnosis. Receiver operator characteristic (ROC) curve analysis identified a gene panel which might distinguish cancer from non-cancerous lesions. Finally, this panel was validated in an independent series of FNA washings (45 cases) in which cytomorphology did not reach definitive diagnosis.
In tissue samples, 14-3-3-sigma, DAPK, CCND2, RASSF1A, CALCA, APC, HIN1, RARbeta2, TIG1, and GSTP1 methylation levels differed significantly among BCa, FB, and NT. ROC curve analysis identified a panel of four gene loci (CCND2, RASSF1A, APC, and HIN1) that discriminated BCa from benign lesions in a set of 78 FNA washings from histologically characterized breast lesions. When this panel was tested in the validation dataset of 45 FNA washings, breast cancer was identified with perfect specificity (100%) when 3 of 4 gene loci tested positive, providing estimated added information of 91% over cytomorphologic evaluation alone.
Our data provide evidence that multigene methylation analysis augments diagnostic accuracy of cytological assessment of suspicious breast lesions, and might be a valuable ancillary tool for breast cancer diagnosis.
我们假设全面的乳腺癌甲基化谱分析可能为使用细针穿刺活检（FNA）对可疑乳腺病变进行诊断评估提供生物标志物。
通过定量甲基化特异性PCR对66例乳腺癌（BCa）、31例纤维腺瘤（FB）和12例正常乳腺（NT）样本经亚硫酸氢盐修饰的DNA中的23个基因启动子进行检测，以确定一组在恶性和非恶性组织中差异甲基化的基因。该组基因在78份术前获得的FNA冲洗液（66例恶性，12例良性）中进行检测，并进行组织病理学诊断。受试者操作特征（ROC）曲线分析确定了一个可能区分癌症与非癌性病变的基因panel。最后，该panel在一个独立的FNA冲洗液系列（45例）中进行验证，这些病例的细胞形态学未达到明确诊断。
在组织样本中，14-3-3-西格玛、DAPK、CCND2、RASSF1A、CALCA、APC、HIN1、RARβ2、TIG1和GSTP1的甲基化水平在BCa、FB和NT之间存在显著差异。ROC曲线分析确定了一个由四个基因位点（CCND2、RASSF1A、APC和HIN1）组成的panel，该panel在一组78份来自组织学特征明确的乳腺病变的FNA冲洗液中区分了BCa与良性病变。当该panel在45份FNA冲洗液的验证数据集中进行检测时，当检测的4个基因位点中有3个呈阳性时，乳腺癌的诊断特异性为100%，与单独的细胞形态学评估相比，估计增加了91%的信息。
我们的数据提供了证据，表明多基因甲基化分析提高了可疑乳腺病变细胞学评估的诊断准确性，可能是乳腺癌诊断的一种有价值的辅助工具。
