An Epigenetic Marker Panel for Detection of Lung Cancer Using Cell-Free Serum DNA

Department of Otolaryngology and Head and Neck Surgery, The Johns Hopkins School of Medicine, Baltimore, Maryland, USA.
Clinical Cancer Research (Impact Factor: 8.72). 05/2011; 17(13):4494-503. DOI: 10.1158/1078-0432.CCR-10-3436
Source: PubMed


We investigated the feasibility of detecting aberrant DNA methylation of some novel and known genes in the serum of lung cancer patients.
To determine the analytic sensitivity, we examined the tumor and the matched serum DNA for aberrant methylation of 15 gene promoters from 10 patients with primary lung tumors by using quantitative methylation-specific PCR. We then tested this 15-gene set to identify the more useful DNA methylation changes in the serum of a limited number of lung cancer patients and controls. In an independent set, we tested the six most promising genes (APC, CDH1, MGMT, DCC, RASSF1A, and AIM1) for further elucidation of the diagnostic application of this panel of markers.
Promoter hypermethylation of at least one of the genes studied was detected in all 10 lung primary tumors. In majority of cases, aberrant methylation in serum DNA was accompanied by methylation in the matched tumor samples. In the independent set, using a single gene that had 100% specificity (DCC), 35.5% (95% CI: 25-47) of the 76 lung cancer patients were correctly identified. For patients without methylated DCC, addition of a logistic regression score that was based on the five remaining genes improved sensitivity from 35.5% to 75% (95% CI: 64-84) but decreased the specificity from 100% to 73% (95% CI: 54-88).
This approach needs to be evaluated in a larger test set to determine the role of this gene set in early detection and surveillance of lung cancer.

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Available from: Andre Carvalho, Jan 27, 2014
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    • "RASSF1A (Ras association domain family 1 isoform A) is a tumor suppressor gene, whose inactivation, mainly achieved by promoter hypermethylation, is involved in the development of many cancers (Donninger et al., 2007, 2015). So far the role of RASSF1A methylation as a biomarker in cfDNA has been investigated, mostly in combination with other parameters, in a variety of tumors such as breast cancer (Papadopoulou et al., 2006; Skvortsova et al., 2006; Agostini et al., 2012), prostate cancer (Papadopoulou et al., 2006), lung cancer (Begum et al., 2011; Ponomaryova et al., 2013), ovarian carcinoma (Bondurant et al., 2011; Liggett et al., 2011; Zhang et al., 2013), gastric cancer (Balgkouranidou et al., 2015), testicular germ cell cancer (Ellinger et al., 2009), nasopharyngeal carcinoma (Wong et al., 2004), renal cell carcinoma (de Martino et al., 2012), bladder cancer (Hauser et al., 2013), and melanoma (Hoon et al., 2004; Koyanagi et al., 2006; Salvianti et al., 2012). In this neoplasia RASSF1A promoter methylation has a frequency of 55% (Spugnardi et al., 2003). "
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    ABSTRACT: Solid tumor release into the circulation cell-free DNA (cfDNA) and circulating tumor cells (CTCs) which represent promising biomarkers for cancer diagnosis. Circulating tumor DNA may be studied in plasma from cancer patients by detecting tumor specific alterations, such as genetic or epigenetic modifications. Ras association domain family 1 isoform A (RASSF1A) is a tumor suppressor gene silenced by promoter hypermethylation in a variety of human cancers including melanoma. The aim of the present study was to assess the diagnostic performance of a tumor-related methylated cfDNA marker in melanoma patients and to compare this parameter with the presence of CTCs. RASSF1A promoter methylation was quantified in cfDNA by qPCR in a consecutive series of 84 melanoma patients and 68 healthy controls. In a subset of 68 cases, the presence of CTCs was assessed by a filtration method (Isolation by Size of Epithelial Tumor Cells, ISET) as well as by an indirect method based on the detection of tyrosinase mRNA by RT-qPCR. The distribution of RASSF1A methylated cfDNA was investigated in cases and controls and the predictive capability of this parameter was assessed by means of the area under the ROC curve (AUC). The percentage of cases with methylated RASSF1A promoter in cfDNA was significantly higher in each class of melanoma patients (in situ, invasive and metastatic) than in healthy subjects (Pearson chi-squared test, p
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    • "Using similar methodology, the influence on meta-regression was determined by omitting one study each time to explore heterogeneity sources. The sample type of tissue or serum would be one of the heterogeneity sources (P < 0.026) when Begum et al. ([12], US) were removed from the meta studies; likewise, the proportion of stage I and aim of the study would become the heterogeneity source when Lin et al. ([17], China), Zhang et al. ([27], China) or Yanagawa et al. ([26], Japan) was removed (P-values were 0.0046, 0.029 and 0.039 respectively). This analysis suggested the above factors should be considered in a future case-control association study. "
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    • "Some of these are as follows: 34 miRNA signatures [6], expression profiles of 11 miRNAs (miR-106a, miR-15b, miR-27b, miR-142-3p, miR-26b, miR-182, miR-126, let7g, let-7i and miR-30e-5p) from serum [7], 7 miRNA signatures [8], overexpression of six snoRNAs [9], and expression of 3 miRs (miR-205, miR-210 and miR-708) in sputum [10]. Additional signatures and markers have also been reported from the plasma proteome [11,12], the salivary proteome [13], the serum epigenome [14], sputum-based genomics [15], and blood-based gene expression studies [16]. However, none of these have progressed sufficiently to provide the necessary specificity and sensitivity required for clinical implementation. "
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