Novel Methylation Biomarker for Non-invasive Diagnostics in Lung Cancer

오태정,( Chang Hun Lee2 ),( Min Ki Lee ),( Yeul Hong Kim ),( Sang Yull Lee ),( Hyo Sung Jeon ),( Shin Yup Lee ),( Seung Soo Yoo ),( Jae Yong Park ),( Sung Whan An ) 대한결핵 및 호흡기학회 2012 대한결핵 및 호흡기학회 추계학술대회 초록집 Vol.114 No.-

To identify aberrantly hypermethylated DNA in lung cancer cells we established a genome-wide analysis for hypermethylation sites, namely Methyl DNA Isolation and Amplification (MeDIA) coupled-CpG microarray analysis. In the comprehensive methyaltion profiling analysis between human lung cancer, A549 cells and normal NHBE cells, we observed that several clusters of genes show a significant level of aberrancy in CpG island methylation pattern in cancer cells compared to normal cells. We further identified PCDHGA12 gene as a new marker of non-invasive diagnostics for lung cancer based on followings. 1) Transcription of PCDHGA12 gene is reactivated after treatment of A549 cells with demethylating agent. 2) Bisulfide clonal-sequencing reveals that CpG island of PCDHGA12 shows a distinctive differential methylation between two cell lines. 3) Pyrosequencing-based quantitative methylation assay for such region in tumor and non-tumorous tissues from lung cancer patients shows aberrant hypermethylation in 37 (92%) of the 40 tumor tissues. In clinical validation by pyrosequencingin induced-sputum of lung cancer patients (n=87) and healthy controls (n=51), we observed aberrant hypermethylation incident at significantly elevated level in samples derived from lung cancer patients. According to the optimal threshold calculated by ROC curve analysis, sensitivity and specificity of PCDHGA12 was 86.2% and 82.4%, respectively. PCDHGA12 methylation status could be a potential methylation biomarker alone or combined with others for the screen and the detection of relapse of lung cancer.

Polymorphisms in cancer-related pathway genes and lung cancer

Lee, Shin Yup,Kang, Hyo-Gyoung,Choi, Jin Eun,Jung, Deuk Kju,Lee, Won Kee,Lee, Hyun Chul,Lee, So Yeon,Yoo, Seung Soo,Lee, Jaehee,Seok, Yangki,Lee, Eung Bae,Cha, Seung Ick,Cho, Sukki,Kim, Chang Ho,Lee, European Respiratory Society 2016 The European respiratory journal Vol.48 No.4

<P>We evaluated the associations between potentially functional variants in a comprehensive list of cancer-related genes and lung cancer in a Korean population.</P><P>A total of 1969 potentially functional single nucleotide polymorphisms (SNPs) of 1151 genes involved in carcinogenesis were evaluated using an Affymetrix custom-made GeneChip in 610 nonsmall cell lung cancer patients and 610 healthy controls. A replication study was conducted in an independent set of 490 cases and 486 controls. 68 SNPs were significantly associated with lung cancer in the discovery set and tested for replication.</P><P>Among the 68 SNPs, three SNPs (corepressor interacting with RBPJ 1 (<I>CIR1</I>) rs13009079T>C, ribonucleotide reductase M1 (<I>RRM1</I>) rs1465952T>C and solute carrier family 38, member 4 (<I>SLC38A4</I>) rs2429467C>T) consistantly showed significant associations with lung cancer in the replication study. In combined analysis, adjusted odds ratio for <I>CIR1</I> rs13009079T>C, <I>RRM1</I> rs1465952T>C and <I>SLC38A4</I> rs2429467C>T were 0.69, 0.71 and 0.73, respectively (p=4×10<SUP>−5</SUP>, 0.01 and 0.001, respectively) under the dominant model. The relative mRNA expression level of <I>CIR1</I> was significantly associated with rs13009079T>C genotypes in normal lung tissues (ptrend=0.03).</P><P>These results suggest that the three SNPs, particularly <I>CIR1</I> rs13009079T>C, may play a role in the pathogenesis of lung cancer.</P>

Thematic Poster : TP-57 ; Association of Polymorphisms in the MicroRNA Target Sites and Survival of Patients in Early-Stage Non-Small-Cell Lung Cancer

( Shin Yup Lee ),( Jin Eun Choi ),( Hyo Sung Jeon ),( Mi Jung Hong ),( Yi Young Choi ),( Won Kee Lee ),( Seung Soo Yoo ),( Jae Hee Lee ),( Eung Bae Lee ),( Seung Ick Cha ),( Chang Ho Kim ),( Young Tae 대한결핵 및 호흡기학회 2014 대한결핵 및 호흡기학회 추계학술대회 초록집 Vol.118 No.0

Background: MicroRNAs (miRNAs) have a key role in carcinogenesis through negative regulation of their target genes. Therefore, genetic variations in miRNAs or their target sites may affect miRNA-mRNA interactions, thereby result in altered expression of target genes. This study was conducted to investigate the associations between single nucleotide polymorphisms (SNP) located in the miRNA target sites (poly-miRTSs) and survival of patients with early-stage non-small cell lung cancer (NSCLC). Methods: Using public SNP database and miRNA target sites prediction program, 354 poly-miRTSs were selected for genotyping. Among these, 154 SNPs applicable to Sequenom’s MassARRAY platform were investigated in 357 patients. A replication study was performed on an independent patient population (n = 479). Renilla luciferase assay and reverse transcription- PCR were conducted to examine functional relevance of potentially functional poly-miRTSs. Results: Of the 154 SNPs analyzed in a discovery set, 14 SNPs were significantly associated with survival outcomes. Among these, KRT81 rs3660G>C was found to be associated with survival outcomes in the validation cohort. In combined analysis, patients with the rs3660 GC+CC genotype had a significantly better overall survival (OS) compared with those with GG genotype (adjusted hazard ratio [aHR] for OS, 0.65; 95% confidence interval [CI] 0.50-0.85; P= 0.001). An increased expression of the reporter gene for the C allele of rs3660 compared with the G allele was observed by luciferase assay. Consistently, the C allele was associated with higher relative expression level of KRT81 in tumor tissues. Conclusions: The rs3660G>C affects KRT81 expression and thus influences survival in early-stage NSCLC. The analysis of the rs3660G>C polymorphism may be useful to identify patients at high risk of a poor disease outcome.

TSC2 genetic variant and prognosis in non‐small cell lung cancer after curative surgery

Lee, Yong Hoon,Do, Sook Kyung,Lee, Shin Yup,Kang, Hyo‐,Gyoung,Choi, Jin Eun,Hong, Mi Jeong,Lee, Jang Hyuck,Lee, Eung Bae,Jeong, Ji Yun,Shin, Kyung Min,Lee, Won Kee,Seok, Yangki,Cho, Sukki,Yoo, S Wiley-Blackwells 2019 Thoracic Cancer Vol.10 No.2

<P>This study was conducted to investigate the associations between polymorphisms of genes involved in the LKB1 pathway and the prognosis of patients with non‐small cell lung cancer (NSCLC) after surgical resection. Twenty‐three single nucleotide polymorphisms (SNPs) in the LKB1 pathway were investigated in 782 patients with NSCLC who underwent curative surgery. The association of SNPs with overall survival (OS) and disease‐free survival (DFS) were analyzed. Among the 23 SNPs investigated, <I>TSC2</I> rs30259G > A was associated with significantly worse OS and DFS (adjusted hazard ratio for OS 1.88, 95% confidence interval 1.21–2.91, <I>P</I> = 0.005; adjusted hazard ratio for DFS 1.65, 95% confidence interval 1.15–2.38, <I>P</I> = 0.01, under codominant models, respectively). Subgroup analysis showed that SNPs were significantly associated with survival outcomes in squamous cell carcinoma, ever‐smokers, and stage I, but not in adenocarcinoma, never‐smokers, and stage II–IIIA. The results suggest that <I>TSC2</I> rs30259G > A may be useful to predict prognosis in patients with NSCLC, especially squamous cell carcinoma, after curative surgery.</P>

Regulatory variants in cancer-related pathway genes predict survival of patients with surgically resected non-small cell lung cancer

Shin, Kyung Min,Hong, Mi Jeong,Lee, Shin Yup,Jin, Cheng Cheng,Baek, Sun Ah,Lee, Jang Hyuck,Choi, Jin Eun,Kang, Hyo-Gyoung,Lee, Won Kee,Seok, Yangki,Lee, Eung Bae,Jeong, Ji Yun,Yoo, Seung Soo,Lee, Jaeh Elsevier BV 2018 Gene Vol.646 No.-

<P><B>Abstract</B></P> <P><B>Background</B></P> <P>We conducted this study to identify genetic variants in cancer-related pathway genes which can predict prognosis of NSCLC patients after surgery, using a comprehensive list of regulatory single nucleotide polymorphisms (SNPs) prioritized by RegulomeDB.</P> <P><B>Method</B></P> <P>A total of 509 potentially functional SNPs in cancer-related pathway genes selected from RegulomeDB were evaluated. These SNPs were analyzed in a discovery set (n=354), and a replication study was performed in an independent set (n=772). The association of the SNPs with overall survival (OS) and disease-free survival (DFS) were analyzed.</P> <P><B>Results</B></P> <P>In the discovery set, 76 SNPs were significantly associated with OS or DFS. Among the 76 SNPs, the association was consistently observed for 5 SNPs (<I>ERCC1</I> rs2298881C>A, <I>BRCA2</I> rs3092989G>A, <I>NELFE</I> rs440454C>T, <I>PPP2R4</I> rs2541164G>A, and <I>LTBP4</I> rs3786527G>A) in the validation set. In combined analysis, <I>ERCC1</I> rs2298881C>A, <I>BRCA2</I> rs3092989, <I>NELFE</I> rs440454C>T, and <I>PPP2R4</I> rs2541164G>A were significantly associated with OS and DFS (adjusted HR ·aHR· for OS=1.46, 0.62, 078, and 0.76, respectively; <I>P</I> =0.003, 0.002, 0.007, and 0.003 respectively; and aHR for DFS=1.27, 0.69, 0.86, and 0.82, respectively; <I>P</I> =0.02, 0.002, 0.03, and 0.008, respectively). The <I>LTBP4</I> rs3786527G>A was significantly associated with better OS (aHR=0.75; <I>P</I> =0.003).</P> <P><B>Conclusion</B></P> <P>Our results suggest that five SNPs in the cancer-related pathway genes may be useful for the prediction of the prognosis in patients with surgically resected NSCLC.</P> <P><B>Highlights</B></P> <P> <UL> <LI> Five among 509 SNPs from RegulomeDB were validated in two independent cohorts. </LI> <LI> Five SNPs in the cancer-related pathway genes were independent prognostic factors. </LI> <LI> RegulomeDB is a practical tool for selecting functional SNPs in regulatory regions. </LI> </UL> </P>

Poster Session : PS 1597 ; Lung Cancer : Association of Polymorphisms in the MicroRNA Target Sites and Survival of Patients in Early-Stage Non- Small-Cell Lung Cancer

( Shin Yup Lee ),( Jin Eun Choi ),( Hyo Sung Jeon ),( Mi Jung Hong ),( Yi Young Choi ),( Won Kee Lee ),( Seung Soo Yoo ),( Jae Hee Lee ),( Eung Bae Lee ),( Seung Ick Cha ),( Chang Ho Kim ),( Y Oung Ta 대한내과학회 2014 대한내과학회 추계학술대회 Vol.2014 No.1

Background: MicroRNAs (miRNAs) have a key role in carcinogenesis through negative regulation of their target genes. Therefore, genetic variations in miRNAs or their target sites may affect miRNA-mRNA interactions, thereby result in altered expression of target genes. This study was conducted to investigate the associations between single nucleotide polymorphisms (SNP) located in the miRNA target sites (poly-miRTSs) and survival of patients with early-stage non-small cell lung cancer (NSCLC). Methods: Using public SNP database and miRNA target sites prediction program, 354 poly-miRTSs were selected for genotyping. Among these, 154 SNPs applicable to Sequenom`s MassARRAY platform were investigated in 357 patients. A replication study was performed on an independent patient population (n = 479). Renilla luciferase assay and reverse transcription-PCR were conducted to examine functional relevance of potentially functional poly-miRTSs. Results: Of the 154 SNPs analyzed in a discovery set, 14 SNPs were significantly associated with survival outcomes. Among these, KRT81 rs3660G>C was found to be associated with survival outcomes in the validation cohort. In combined analysis,patients with the rs3660 GC+CC genotype had a signifi cantly better overall survival (OS) compared with those with GG genotype (adjusted hazard ratio [aHR] for OS, 0.65; 95% confi dence interval [CI] 0.50-0.85; P= 0.001). An increased expression of the reporter gene for the C allele of rs3660 compared with the G allele was observed by luciferase assay. Consistently, the C allele was associated with higher relative expressionlevel of KRT81 in tumor tissues. Conclusions: The rs3660G>C affects KRT81 expression and thus infi uences survival in early-stage NSCLC. The analysis of the rs3660G>C polymorphism may be useful to identify patients at high risk of a poor disease outcome.

Pri-let-7a-2 rs1143770C>T Associated with Prognosis of Surgically Resected Non-Small Cell Lung Cancer

( Yong Dae Lee ),( Kyung Min Shin ),( Deuk Kju Jung ),( Won Kee Lee ),( Hyewon Seo ),( So Yeon Lee ),( Seung Soo Yoo ),( Shin Yup Lee ),( Seung Ick Cha ),( Jaehee Lee ),( Chang Ho Kim ),( Yangki Seok 대한결핵 및 호흡기학회 2015 대한결핵 및 호흡기학회 추계학술대회 초록집 Vol.120 No.0

Background: Evidence indicates that the let-7 microRNA (miRNA) may be a prognostic factor in lung cancer. Genetic variation in miRNA precursors could influence the processing and expression of miRNAs, which could affect the prognosis of lung cancer. We aimed to investigate the impact of single nucleotide polymorphisms (SNPs) of pri-let-7 on the prognosis of non-small cell lung cancer (NSCLC). Methods: Seven hundred sixty-one patients with surgically resected NSCLC were included. The four SNPs (pri-let-7a-2 rs1143770 and rs629367, pri-let-7a-1 rs10739971, and pri-let-7f-2 rs17276588) were genotyped using a polymerase chain reaction-restriction fragment length polymorphism assay. The genotype associations with overall survival (OS) and disease-free survival (DFS) were evaluated. Results: Of the four SNPs analyzed, the rs1143770C>T was identified to be significantly associated with OS and DFS. The rs1143770 CT or TT genotype exhibited a significantly better OS and DFS compared with the rs1143770 CC genotype (adjusted hazard ratio for OS = 0.67, 95% confidence interval = 0.49―0.91, P = 0.01 and adjusted hazard ratio for DFS = 0.74, 95% confidence interval = 0.58―0.95, P = 0.02). Conclusions: This observation indicates that the pri-let-7a-2 rs1143770C>T may have a prognostic impact on surgically resected NSCLC.

Clinical Utility of CT-Based Bronchial Aspirate TB-PCR for the Rapid Diagnosis of Pleural Tuberculosis

Lee, Jaehee,Lee, So Yeon,Choi, Keum Ju,Lim, Jae Kwang,Yoo, Seung Soo,Lee, Shin Yup,Cha, Seung Ick,Park, Jae Yong,Kim, Chang Ho The Korean Academy of Tuberculosis and Respiratory 2013 Tuberculosis and Respiratory Diseases Vol.75 No.4

Background: Thoracoscopic pleural biopsy is often required for rapid and confirmative diagnosis in patients with suspected pleural tuberculosis (PL-TB). However, this method is more invasive and costly than its alternatives. Therefore, we evaluated the clinical utility of the chest computed tomography (CT)-based bronchial aspirate (BA) TB-polymerase chain reaction (PCR) test in such patients. Methods: Bronchoscopic evaluation was performed in 54 patients with presumptive PL-TB through diagnostic thoracentesis but without a positive result of sputum acid-fast bacilli (AFB) smear, pleural fluid AFB smear, or pleural fluid TB-PCR test. Diagnostic yields of BA were evaluated according to the characteristics of parenchymal lesions on chest CT. Results: Chest radiograph and CT revealed parenchymal lesions in 25 (46%) and 40 (74%) of 54 patients, respectively. In cases with an absence of parenchymal lesions on chest CT, the bronchoscopic approach had no diagnostic benefit. BA TB-PCR test was positive in 21 out of 22 (95%) patients with early-positive results. Among BA results from 20 (37%) patients with patchy consolidative CT findings, eight (40%) were AFB smear-positive, 18 (90%) were TB-PCR-positive, and 19 (95%) were culture-positive. Conclusion: The BA TB-PCR test seems to be a satisfactory diagnostic modality in patients with suspected PL-TB and patchy consolidative CT findings. For rapid and confirmative diagnosis in these patients, the bronchoscopic approach with TB-PCR may be preferable to the thoracoscopy.

A Functional Polymorphism in the <i>CHRNA3</i> Gene and Risk of Chronic Obstructive Pulmonary Disease in a Korean Population

Lee, Jae Yeon,Yoo, Seung Soo,Kang, Hyo-Gyoung,Jin, Guang,Bae, Eun Young,Choi, Yi Young,Choi, Jin Eun,Jeon, Hyo-Sung,Lee, Jaehee,Lee, Shin Yup,Cha, Seung-Ick,Kim, Chang Ho,Park, Jae Yong The Korean Academy of Medical Sciences 2012 JOURNAL OF KOREAN MEDICAL SCIENCE Vol.27 No.12

<P>A genome-wide association study has identified the 15q25 region as being associated with the risk of chronic obstructive pulmonary disease (COPD) in Caucasians. This study intended as a confirmatory assessment of this association in a Korean population. The rs6495309C > T polymorphism in the promoter of nicotinic acetylcholine receptor alpha subunit 3 (<I>CHRNA3</I>) gene was investigated in a case-control study that consisted of 406 patients with COPD and 394 healthy control subjects. The rs6495309 CT or TT genotype was associated with a significantly decreased risk of COPD when compared to the rs6495309 CC genotype (adjusted odds ratio = 0.69, 95% confidence interval = 0.50-0.95, <I>P</I> = 0.023). The effect of the rs6495309C > T on the risk of COPD was more evident in moderate to very severe COPD than in mild COPD under a dominant model for the variant T allele (<I>P</I> = 0.024 for homogeneity). The <I>CHRNA3</I> rs6495309C > T polymorphism on chromosome 15q25 is associated with the risk of COPD in a Korean population.</P>

Polymorphisms in the Caspase Genes and the Risk of Lung Cancer

Lee, Shin Yup,Choi, Yi Young,Choi, Jin Eun,Kim, Min Jung,Kim, Jong-Sik,Jung, Deuk Kju,Kang, Hyo-Gyoung,Jeon, Hyo-Sung,Lee, Won Kee,Jin, Guang,Cha, Seung Ick,Kim, Chang Ho,Jung, Tae Hoon,Park, Jae Yong Elsevier 2010 JOURNAL OF THORACIC ONCOLOGY Vol.5 No.8

<P>INTRODUCTION:: Caspases (CASPs) are important regulators and executioners in apoptosis pathway and play a crucial role in the development and progression of cancer. On the basis of the interactions of CASPs in the apoptotic pathway, we evaluated the association between 11 polymorphisms of CASP3, 6, 7, 8, 9, and 10 genes and the risk of lung cancer. METHODS:: The genotypes were determined in 720 patients with lung cancer and 720 healthy controls frequency matched for age and gender. RESULTS:: In individual polymorphism analysis, the CASP7 rs2227310C>G and CASP9 rs4645981C>T were associated with 1.40-fold and 1.28-fold increased risk of lung cancer under recessive and dominant models for the variant allele of each polymorphism, respectively. In the case of the CASP3 77G>A, subjects with the GG genotype were at a 1.19-fold increased risk of lung cancer compared with those with at least one variant allele. When the three polymorphisms were combined, the risk of lung cancer increased in a dose-dependent manner as the number of risk genotypes increased (ptrend <0.001). Subjects with two and three risk genotypes carried a significantly increased risk of lung cancer compared with those with zero risk genotype (adjusted odds ratio = 1.56, 95% confidence interval = 1.14-2.13, p = 0.005 and adjusted odds ratio = 2.54, 95% confidence interval = 1.28-5.02, p = 0.008, respectively). CONCLUSIONS:: These results suggest that a combined analysis of these three CASP gene polymorphisms might better predict the risk of lung cancer than analysis of a single polymorphism.</P>