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Spagnol LW, Polettini J, Silveira DA, Wegner GRM, Paiva DFF. P16 gene promoter methylation is associated with oncogenesis and progression of gastric carcinomas: A systematic review and meta-analysis. Crit Rev Oncol Hematol 2022; 180:103843. [DOI: 10.1016/j.critrevonc.2022.103843] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/30/2022] [Revised: 10/02/2022] [Accepted: 10/11/2022] [Indexed: 11/06/2022] Open
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Zhao H, Cheng Y, Kalra A, Ma K, Zheng Y, Ziman B, Tressler C, Glunde K, Shin EJ, Ngamruengphong S, Khashab M, Singh V, Anders RA, Jit S, Wyhs N, Chen W, Li X, Lin DC, Meltzer SJ. Generation and multiomic profiling of a TP53/CDKN2A double-knockout gastroesophageal junction organoid model. Sci Transl Med 2022; 14:eabq6146. [PMID: 36449602 PMCID: PMC10026384 DOI: 10.1126/scitranslmed.abq6146] [Citation(s) in RCA: 16] [Impact Index Per Article: 5.3] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/02/2022]
Abstract
Inactivation of the tumor suppressor genes tumor protein p53 (TP53) and cyclin-dependent kinase inhibitor 2A (CDKN2A) occurs early during gastroesophageal junction (GEJ) tumorigenesis. However, because of a paucity of GEJ-specific disease models, cancer-promoting consequences of TP53 and CDKN2A inactivation at the GEJ have not been characterized. Here, we report the development of a wild-type primary human GEJ organoid model and a CRISPR-edited transformed GEJ organoid model. CRISPR-Cas9-mediated TP53 and CDKN2A knockout (TP53/CDKN2AKO) in GEJ organoids induced morphologic dysplasia and proneoplastic features in vitro and tumor formation in vivo. Lipidomic profiling identified several platelet-activating factors (PTAFs) among the most up-regulated lipids in CRISPR-edited organoids. PTAF/PTAF receptor (PTAFR) abrogation by siRNA knockdown or a pharmacologic inhibitor (WEB2086) reduced proliferation and other proneoplastic features of TP53/CDKN2AKO GEJ organoids in vitro and tumor formation in vivo. In addition, murine xenografts of Eso26, an established human esophageal adenocarcinoma cell line, were suppressed by WEB2086. Mechanistically, TP53/CDKN2A dual inactivation disrupted both the transcriptome and the DNA methylome, likely mediated by key transcription factors, particularly forkhead box M1 (FOXM1). FOXM1 activated PTAFR transcription by binding to the PTAFR promoter, further amplifying the PTAF-PTAFR pathway. Together, these studies established a robust model system for investigating early GEJ neoplastic events, identified crucial metabolic and epigenomic changes occurring during GEJ model tumorigenesis, and revealed a potential cancer therapeutic strategy. This work provides insights into proneoplastic mechanisms associated with TP53/CDKN2A inactivation in early GEJ neoplasia, which may facilitate early diagnosis and prevention of GEJ neoplasms.
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Affiliation(s)
- Hua Zhao
- Center for Craniofacial Molecular Biology, Herman Ostrow School of Dentistry, and Norris Comprehensive Cancer Center, University of Southern California, Los Angeles, CA 90033, USA
- Division of Gastroenterology and Hepatology, Department of Medicine and Oncology, Sidney Kimmel Comprehensive Cancer Center, Johns Hopkins University School of Medicine, Baltimore, MD 21287, USA
- Clinical Laboratory, The First Affiliated Hospital of Xi’an Jiaotong University, No. 277 Yanta West Road, Xi’an 710061, Shaanxi, China
| | - Yulan Cheng
- Division of Gastroenterology and Hepatology, Department of Medicine and Oncology, Sidney Kimmel Comprehensive Cancer Center, Johns Hopkins University School of Medicine, Baltimore, MD 21287, USA
| | - Andrew Kalra
- Division of Gastroenterology and Hepatology, Department of Medicine and Oncology, Sidney Kimmel Comprehensive Cancer Center, Johns Hopkins University School of Medicine, Baltimore, MD 21287, USA
| | - Ke Ma
- Division of Gastroenterology and Hepatology, Department of Medicine and Oncology, Sidney Kimmel Comprehensive Cancer Center, Johns Hopkins University School of Medicine, Baltimore, MD 21287, USA
- Einstein Healthcare Network, Philadelphia, PA 19136, USA
| | - Yueyuan Zheng
- Department of Medicine, Cedars-Sinai Medical Center, Los Angeles, CA 90048, USA
| | - Benjamin Ziman
- Center for Craniofacial Molecular Biology, Herman Ostrow School of Dentistry, and Norris Comprehensive Cancer Center, University of Southern California, Los Angeles, CA 90033, USA
| | - Caitlin Tressler
- Russell H. Morgan Department of Radiology and Radiological Science, Division of Cancer Imaging Research, Johns Hopkins University School of Medicine, Baltimore, MD 21287, USA
| | - Kristine Glunde
- Russell H. Morgan Department of Radiology and Radiological Science, Division of Cancer Imaging Research, Johns Hopkins University School of Medicine, Baltimore, MD 21287, USA
- The Sidney Kimmel Comprehensive Cancer Center, Johns Hopkins University School of Medicine, Baltimore, MD 21287, USA
- Department of Biological Chemistry, Johns Hopkins University School of Medicine, Baltimore, MD 21287, USA
| | - Eun Ji Shin
- Division of Gastroenterology and Hepatology, Department of Medicine and Oncology, Sidney Kimmel Comprehensive Cancer Center, Johns Hopkins University School of Medicine, Baltimore, MD 21287, USA
| | - Saowanee Ngamruengphong
- Division of Gastroenterology and Hepatology, Department of Medicine and Oncology, Sidney Kimmel Comprehensive Cancer Center, Johns Hopkins University School of Medicine, Baltimore, MD 21287, USA
| | - Mouen Khashab
- Division of Gastroenterology and Hepatology, Department of Medicine and Oncology, Sidney Kimmel Comprehensive Cancer Center, Johns Hopkins University School of Medicine, Baltimore, MD 21287, USA
| | - Vikesh Singh
- Division of Gastroenterology and Hepatology, Department of Medicine and Oncology, Sidney Kimmel Comprehensive Cancer Center, Johns Hopkins University School of Medicine, Baltimore, MD 21287, USA
| | - Robert A. Anders
- Department of Pathology, Johns Hopkins University School of Medicine, Baltimore, MD 21287, USA
| | - Simran Jit
- Division of Gastroenterology and Hepatology, Department of Medicine and Oncology, Sidney Kimmel Comprehensive Cancer Center, Johns Hopkins University School of Medicine, Baltimore, MD 21287, USA
| | - Nicolas Wyhs
- The Sidney Kimmel Comprehensive Cancer Center, Johns Hopkins University School of Medicine, Baltimore, MD 21287, USA
| | - Wei Chen
- Clinical Laboratory, The First Affiliated Hospital of Xi’an Jiaotong University, No. 277 Yanta West Road, Xi’an 710061, Shaanxi, China
| | - Xu Li
- Center for Translational Medicine, The First Affiliated Hospital of Xi’an Jiaotong University, Xi’an 710061, Shaanxi, China
| | - De-Chen Lin
- Center for Craniofacial Molecular Biology, Herman Ostrow School of Dentistry, and Norris Comprehensive Cancer Center, University of Southern California, Los Angeles, CA 90033, USA
| | - Stephen J. Meltzer
- Division of Gastroenterology and Hepatology, Department of Medicine and Oncology, Sidney Kimmel Comprehensive Cancer Center, Johns Hopkins University School of Medicine, Baltimore, MD 21287, USA
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Panarese I, De Vita F, Ronchi A, Romano M, Alfano R, Di Martino N, Zito Marino F, Ferraraccio F, Franco R. Predictive biomarkers along gastric cancer pathogenetic pathways. Expert Rev Anticancer Ther 2017; 17:417-425. [PMID: 28277834 DOI: 10.1080/14737140.2017.1301207] [Citation(s) in RCA: 21] [Impact Index Per Article: 2.6] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/23/2022]
Abstract
INTRODUCTION Gastric cancer is the second leading cause of cancer all over the world. Unfortunately, several gastric cancers are diagnosed in an advanced stage and chemotherapy and/or target therapies remain the only options to treat patients. Areas covered: Herein we evaluate the new molecular proposal of gastric cancer classification, offering the possibility to recognize different pathogenetic mechanisms and molecular biomarkers potentially useful for target therapies. Expert commentary: The possibility of introducing new specific tests for identification of molecular biomarkers critical for targeted therapies response represents the new frontier in the selection of gastric cancer patients to improve their survival. Besides HER2, already used in clinical settings as a target biomarker for biological therapy in gastric cancer patients with tissue cancer cells overexpressing HER2, other promising target biomarkers which are deregulated in gastric cancer, such as MET and FGFR, could be identified in tissue and then used for therapeutic purposes. In addition immunotherapy represents the most promising possibility of advanced gastric cancer treatment. In particular, as in other solid tumors, PD-1/PDL1 pathway has emerged in several clinical trials as an interesting therapeutic target.
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Affiliation(s)
- Iacopo Panarese
- a Pathology Unit, Department of Mental and Physical Health and Preventive Medicine , Università della Campania 'Luigi Vanvitelli,' , Naples , Italy
| | - Ferdinando De Vita
- b Division of Medical Oncology, Department of Internal and Experimental Medicine 'F. Magrassi' , Università della Campania 'Luigi Vanvitelli,' , Naples , Italy
| | - Andrea Ronchi
- a Pathology Unit, Department of Mental and Physical Health and Preventive Medicine , Università della Campania 'Luigi Vanvitelli,' , Naples , Italy
| | - Marco Romano
- c Division of Hepatogastroenterology, Department of Clinical and Experimental Medicine , Università della Campania 'Luigi Vanvitelli,' , Naples , Italy
| | - Roberto Alfano
- d General Surgery Unit, Department of Anesthesiology , Surgery and Emergency, Università della Campania 'Luigi Vanvitelli,' , Naples , Italy
| | - Natale Di Martino
- e Department of Internal Medicine , Surgical, Neurological Metabolic Disease and Geriatric Medicine, Università della Campania 'Luigi Vanvitelli,' , Naples , Italy
| | - Federica Zito Marino
- a Pathology Unit, Department of Mental and Physical Health and Preventive Medicine , Università della Campania 'Luigi Vanvitelli,' , Naples , Italy.,f Pathology Unit, Istituto dei tumori 'Fondazione G. Pascale'
| | - Francesca Ferraraccio
- a Pathology Unit, Department of Mental and Physical Health and Preventive Medicine , Università della Campania 'Luigi Vanvitelli,' , Naples , Italy
| | - Renato Franco
- a Pathology Unit, Department of Mental and Physical Health and Preventive Medicine , Università della Campania 'Luigi Vanvitelli,' , Naples , Italy
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Mahajan A. Practical issues in the application of p16 immunohistochemistry in diagnostic pathology. Hum Pathol 2016; 51:64-74. [DOI: 10.1016/j.humpath.2015.12.021] [Citation(s) in RCA: 33] [Impact Index Per Article: 3.7] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 06/19/2015] [Revised: 12/21/2015] [Accepted: 12/23/2015] [Indexed: 11/25/2022]
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Zhang X, Li J, He Z, Duan H, Gao W, Wang H, Yu S, Chen W, Zheng Y. Associations between DNA methylation in DNA damage response-related genes and cytokinesis-block micronucleus cytome index in diesel engine exhaust-exposed workers. Arch Toxicol 2015; 90:1997-2008. [DOI: 10.1007/s00204-015-1598-2] [Citation(s) in RCA: 22] [Impact Index Per Article: 2.2] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/22/2015] [Accepted: 09/10/2015] [Indexed: 01/07/2023]
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Nakamura J, Tanaka T, Kitajima Y, Noshiro H, Miyazaki K. Methylation-mediated gene silencing as biomarkers of gastric cancer: A review. World J Gastroenterol 2014; 20:11991-12006. [PMID: 25232236 PMCID: PMC4161787 DOI: 10.3748/wjg.v20.i34.11991] [Citation(s) in RCA: 39] [Impact Index Per Article: 3.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 10/26/2013] [Revised: 01/29/2014] [Accepted: 04/09/2014] [Indexed: 02/06/2023] Open
Abstract
Despite a decline in the overall incidence of gastric cancer (GC), the disease remains the second most common cause of cancer-related death worldwide and is thus a significant global health problem. The best means of improving the survival of GC patients is to screen for and treat early lesions. However, GC is often diagnosed at an advanced stage and is associated with a poor prognosis. Current diagnostic and therapeutic strategies have not been successful in decreasing the global burden of the disease; therefore, the identification of reliable biomarkers for an early diagnosis, predictive markers of recurrence and survival and markers of drug sensitivity and/or resistance is urgently needed. The initiation and progression of GC depends not only on genetic alterations but also epigenetic changes, such as DNA methylation and histone modification. Aberrant DNA methylation is the most well-defined epigenetic change in human cancers and is associated with inappropriate gene silencing. Therefore, an increasing number of genes methylated at the promoter region have been targeted as possible biomarkers for different purposes, including early detection, classification, the assessment of the tumor prognosis, the development of therapeutic strategies and patient follow-up. This review article summarizes the current understanding and recent evidence regarding DNA methylation markers in GC with a focus on the clinical potential of these markers.
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Role of p16 gene promoter methylation in gastric carcinogenesis: a meta-analysis. Mol Biol Rep 2014; 41:4481-92. [PMID: 24610350 DOI: 10.1007/s11033-014-3319-y] [Citation(s) in RCA: 5] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/27/2013] [Accepted: 02/24/2014] [Indexed: 01/30/2023]
Abstract
This meta-analysis was performed to evaluate the relationships between promoter DNA methylation in tumor suppressor gene p16 and gastric carcinogenesis. The PubMed, CISCOM, CINAHL, Web of Science, Google Scholar, EBSCO, Cochrane Library and CBM databases were searched for relevant articles published before November 1st, 2013 without any language restrictions. Meta-analysis was conducted using the STATA 12.0 software. Crude odds ratios (ORs) with 95% confidence intervals (95% CI) were calculated. Forty-seven clinical cohort studies that met all inclusion criteria were included in this meta-analysis. A total of 2,813 gastric cancer (GC) patients were assessed. Our meta-analysis results revealed that the frequencies of p16 promoter methylation in the GC tissues were higher than those of normal and adjacent tissues (Normal: OR = 23.04, 95% CI = 13.55-39.15, P < 0.001; Adjacent: OR = 4.42, 95% CI = 1.66-11.76, P = 0.003; respectively). Furthermore, we observed significant associations of p16 promoter methylation with TNM stage, histologic grade, invasive grade, lymph node metastasis of GC (TNM stage: OR = 3.60, 95% CI: 2.17-5.98, P < 0.001; Histologic grade: OR = 2.63, 95% CI: 1.55-4.45, P < 0.001; Invasive grade: OR = 3.44, 95% CI: 1.68-7.06, P = 0.001; Lymph node metastasis: OR = 2.68, 95% CI: 1.66-4.32, P < 0.001; respectively). However, there were no correlations of p16 promoter methylation with the TNM stage and Helicobacter pylori (HP) infection of GC (Tumor size: OR = 0.76, 95% CI: 0.14-4.07, P = 0.746; HP infection: OR = 1.31, 95% CI: 0.75-2.27, P = 0.342; respectively). Our findings provide empirical evidence that p16 promoter methylation may play an important role in gastric carcinogenesis. Thus, p16 promoter methylation may be a promising potential biomarker for the early diagnosis of GC.
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Lee S, Kim H, Kim H, Kim C, Kim I. The Utility of p16INK4a and Ki-67 as a Conjunctive Tool in Uterine Cervical Lesions. KOREAN JOURNAL OF PATHOLOGY 2012; 46:253-60. [PMID: 23110011 PMCID: PMC3479762 DOI: 10.4132/koreanjpathol.2012.46.3.253] [Citation(s) in RCA: 5] [Impact Index Per Article: 0.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Subscribe] [Scholar Register] [Received: 10/17/2011] [Revised: 04/13/2012] [Accepted: 05/29/2012] [Indexed: 11/29/2022]
Abstract
Background Immunohistochemical staining for p16INK4a and Ki-67 has been used to improve the accuracy in making a diagnosis of the uterine cervix cancer on biopsy. This study was conducted to examine the usefulness of these markers in the pathological diagnosis based on cervical biopsy. Methods We selected a consecutive series of 111 colposcopically directed cervical punch biopsies. Using these biopsy samples, we performed an immunohistochemical staining for p16INK4a and Ki-67 to establish a diagnosis. The slides were circulated among four pathologists in a sequential order: the hematoxylin and eosin (H&E) slide, H&E slide and p16INK4a-stained slide, and H&E slide, p16INK4a- and Ki-67-stained slides. Results The overall rates of the concordance in the first, the second, and the third diagnoses were 77.5%, 82.0%, and 82.0%, respectively. The rate of the concordance in the diagnosis of cervical intraepithelial neoplasm (CIN) 2/3 was increased from 62.2% to 73.0%. But there was a variability in the rate of the revision of the diagnosis between the pathologists. With the application of criteria for interpreting the expressions of p16INK4a and Ki-67, benign and CIN 1 lesions showed a p16INK4a expression score of 0 or 1. But CIN 2 and CIN 3 lesions showed a p16INK4a expression score of 2 and 3, respectively. Conclusions The immunostain for p16INK4a and Ki-67 might be useful in reducing an inter-observer variability. But criteria for interpreting both markers should be strictly applied.
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Affiliation(s)
- Sangho Lee
- Department of Pathology, Gachon University Gil Hospital, Gachon University of Medicine and Science, Incheon, Korea
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p16 Expression in Squamous and Trophoblastic Lesions of the Upper Female Genital Tract. Int J Gynecol Pathol 2010; 29:513-22. [DOI: 10.1097/pgp.0b013e3181e2fe70] [Citation(s) in RCA: 17] [Impact Index Per Article: 1.1] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/25/2022]
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Neves Filho EHC, Alves MKS, Lima VP, Rabenhorst SHB. MTHFR C677T polymorphism and differential methylation status in gastric cancer: an association with Helicobacter pylori infection. Virchows Arch 2010; 457:627-33. [DOI: 10.1007/s00428-010-0996-3] [Citation(s) in RCA: 8] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/25/2010] [Revised: 10/01/2010] [Accepted: 10/05/2010] [Indexed: 12/11/2022]
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Abbaszadegan MR, Moaven O, Sima HR, Ghafarzadegan K, A'rabi A, Forghani MN, Raziee HR, Mashhadinejad A, Jafarzadeh M, Esmaili-Shandiz E, Dadkhah E. p16 promoter hypermethylation: a useful serum marker for early detection of gastric cancer. World J Gastroenterol 2008; 14:2055-2060. [PMID: 18395906 PMCID: PMC2701527 DOI: 10.3748/wjg.14.2055] [Citation(s) in RCA: 69] [Impact Index Per Article: 4.1] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 12/21/2007] [Revised: 02/10/2008] [Indexed: 02/06/2023] Open
Abstract
AIM To determine p16 promoter hypermethylation in gastric tumoral tissue and serum samples, its impact on p16-protein expression, and correlation with clinical and histological features. METHODS Samples were obtained from 52 histologically confirmed cases of gastric adenocarcinoma. Gastric tissue and serum of 50 age- and sex-matched individuals with normal gastroscopy and biopsy were obtained as control samples. Methylation-specific polymerase chain reaction (MSP) was used to evaluate methylation status of p16 promoter. p16-protein expression was analyzed by immunohistochemical staining on paraffin-embedded sections. RESULTS Methylation was detected in 44.2% (23/52) of tumoral tissues. 60.9% of them were also methylated in serum, i.e., 26.9% of all patients (14/52). Methylation was not detected in tissue and sera of control samples. p16-protein expression was decreased in 61.5% of cases (32/52), and was significantly associated with promoter hypermethylation (P < 0.001). Methylation was significantly more frequent in higher pathological grades (P < 0.05). Methylation was not associated with other clinicopathological features and environmental factors including H pylori infection and smoking. CONCLUSION p16 promoter hypermethylation is an important event in gastric carcinogenesis. It is the principle mechanism of p16 gene silencing. It is related to malignant tumor behavior. Detection of DNA methylation in serum may be a biomarker for early detection of gastric cancer.
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Milne ANA, Sitarz R, Carvalho R, Polak MM, Ligtenberg M, Pauwels P, Offerhaus GJA, Weterman MAJ. Molecular analysis of primary gastric cancer, corresponding xenografts, and 2 novel gastric carcinoma cell lines reveals novel alterations in gastric carcinogenesis. Hum Pathol 2007; 38:903-913. [PMID: 17376510 DOI: 10.1016/j.humpath.2006.12.010] [Citation(s) in RCA: 23] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 07/11/2006] [Revised: 12/06/2006] [Accepted: 12/08/2006] [Indexed: 02/05/2023]
Abstract
We report the molecular characterization of 8 primary gastric carcinomas, corresponding xenografts, and 2 novel gastric carcinoma cell lines. We compared the tumors and cell lines, with respect to histology, immunohistochemistry, copy number, and hypermethylation of up to 38 genes using methylation-specific multiplex ligation-dependent probe amplification, and TP53 and CDH1 mutation analysis where relevant. The primary tumors and xenografts were histologically comparable and shared expression of 11 of 14 immunohistochemical markers (E-cadherin, beta-catenin, COX-2, p53, p16, TFF1, cyclin E, MLH1, SMAD4, p27, KLK3, CASR, CHFR, and DAPK1). Gains of CASR, DAPK1, and KLK3--not yet described in gastric cancer--were present in the primary tumors, xenografts, and cell lines. The most prominent losses occurred at CDKN2A (p16), CDKN2B (p15), CDKN1B (p27/KIP1), and ATM. Except for ATM, these losses were found only in the cell line or xenograft, suggesting an association with tumor progression. However, examination of p16 and p27 in 174 gastric cancers using tissue microarrays revealed no significant correlation with tumor stage or lymph node status. Further losses and hypermethylation were detected for MLH1, CHFR, RASSF1, and ESR, and were also seen in primary tumors. Loss of CHFR expression correlated significantly with the diffuse phenotype. Interestingly, we found the highest rate of methylation in primary tumors which gave rise to cell lines. In addition, both cell lines harbored mutations in CDH1, encoding E-cadherin. Xenografts and gastric cancer cell lines remain an invaluable research tool in the uncovering of the multistep progression of cancer. The frequent gains, losses, and hypermethylation reported in this study indicate that the involved genes or chromosomal regions may be relevant to gastric carcinogenesis.
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Affiliation(s)
- Anya N A Milne
- Department of Pathology, University Medical Centre, 3508 GA Utrecht, The Netherlands.
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Luo D, Zhang B, Lv L, Xiang S, Liu Y, Ji J, Deng D. Methylation of CpG islands of p16 associated with progression of primary gastric carcinomas. J Transl Med 2006; 86:591-8. [PMID: 16534497 DOI: 10.1038/labinvest.3700415] [Citation(s) in RCA: 48] [Impact Index Per Article: 2.5] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/31/2022] Open
Abstract
Inactivation of p16 by methylation of CpG islands is a frequent early event in gastric carcinogenesis. The positive relationship between p16 methylation and the clinical characteristics of gastric carcinomas (GC) has not been reported to date. In the present study, a DHPLC assay to quantify p16 methylation was established (detection limit by fluorescence detector: 1:255 (Methlyated vs Unmethylated)). The proportion of methylated p16 in the representative samples was confirmed and standardized by clone sequencing. Then, the DHPLC and two regular methylation-specific PCR (MSP) assays were used to detect p16 methylation in 82 paired, resected GCs and their adjacent normal tissues. Results showed that the average proportion of methylated p16 in GCs was significantly higher than that in their adjacent samples (12.90 vs 0.63%; t-test P=0.005). A much higher proportion of methylated p16 was detected in GCs with metastases (local or distant) than without metastases (14.76 vs 2.61%; t-test P=0.014). A proportional relationship was observed between clinical stages and positive rates of p16 methylation in GCs and/or adjacent tissues: 27.3, 37.5, and 58.8% (by DHPLC) for stage-I, -II, -III-IV of GCs, respectively (two-sided Fisher's exact test P=0.016). To confirm the data obtained by DHPLC, two MSP primer sets (p16-M and p16-M2) were also used to analyze p16 methylation in the same set of samples simultaneously. Data of MSP assay using the primer set p16-M2, but not p16-M, correlated with that of DHPLC. These results imply that the primer set p16-M2 might be more suitable than p16-M to detect p16 methylation in gastric tissues. In conclusion, the present data indicates that p16 methylation correlates with progression of GCs significantly.
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Affiliation(s)
- Daya Luo
- Peking University School of Oncology, Beijing Cancer Hospital and Beijing Institute for Cancer Research, Beijing, China
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Takada H, Imoto I, Tsuda H, Nakanishi Y, Sakakura C, Mitsufuji S, Hirohashi S, Inazawa J. Genomic loss and epigenetic silencing of very-low-density lipoprotein receptor involved in gastric carcinogenesis. Oncogene 2006; 25:6554-62. [PMID: 16715143 DOI: 10.1038/sj.onc.1209657] [Citation(s) in RCA: 24] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/09/2022]
Abstract
Homozygous loss in the genomic sequence, a mechanism for inactivating tumor-suppressor genes (TSGs) in cancer, has been used as a tag for the identification of novel TSGs, and array-based comparative genomic hybridization (array-CGH) has a great potential for high-throughput identification of this change. We identified a homozygous loss of the very-low-density lipoprotein receptor (VLDLR) gene (9p24.2) from genome-wide screening for copy-number alterations in 32 gastric cancer (GC) cell lines using array-CGH. Although previous reports demonstrated mRNA or protein expression of VLDLR in various cancers including GC, the association between genomic losses or epigenetic silencing of this gene and carcinogenesis has never been reported before. Homozygous deletion of VLDLR was also seen in primary GCs, albeit infrequently, and about half of GC cell lines showed lost or reduced VLDLR expression. The VLDLR expression was restored in gene-silenced GC cells after treatment with 5-aza 2'-deoxycytidine. According to methylation analyses, hypermethylation of the VLDLR promoter region, which all of GC lines without its expression showed, occurred in some primary GCs. Restoration of VLDLR type I expression in GC cells reduced colony formation. These results suggest that not only the expression of VLDLR but also genetic or epigenetic silencing of this gene may contribute to tumor formation and be involved in gastric carcinogenesis.
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Affiliation(s)
- H Takada
- Department of Molecular Cytogenetics, Medical Research Institute and Graduate School of Biomedical Science, Tokyo, Japan
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Abstract
The upper gastrointestinal (GI) cancers have various carcinogenic pathways and precursor lesions, such as dysplasia for esophageal squamous cell carcinoma, Barrett esophagus for esophageal adenocarcinoma, and intestinal metaplasia for the intestinal-type of gastric cancer. Recently, many epigenetic events in carcinogenic pathways have been revealed, along with genomic and genetic alterations. This information has provided deeper insight into an understanding of the mechanisms of upper GI carcinogenesis. Moreover, detection methods of aberrant methylation have been applied to clinical fields to stratify high-risk groups, detect early cancer, and to predict clinical outcomes. In this review, a variety of information is summarized regarding gene hypermethylation in esophageal and gastric cancer.
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Affiliation(s)
- Fumiaki Sato
- Department of Pathology, University of Maryland School of Medicine, Baltimore, Maryland 21201, USA.
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Prall F, Ostwald C, Weirich V, Nizze H. p16(INK4a) promoter methylation and 9p21 allelic loss in colorectal carcinomas: relation with immunohistochemical p16(INK4a) expression and with tumor budding. Hum Pathol 2006; 37:578-85. [PMID: 16647956 DOI: 10.1016/j.humpath.2006.01.005] [Citation(s) in RCA: 11] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 09/10/2005] [Accepted: 01/03/2006] [Indexed: 01/02/2023]
Abstract
In colorectal carcinomas, p16(INK4a) inactivation is known to occur by allelic loss and by promoter methylation, but mutations are rare. p16(INK4a) is up-regulated in tumor buds, and the consequent shutdown of proliferation may be a prerequisite for tumor budding. Fifty-seven colorectal carcinomas from a consecutive series were investigated. Using DNA from tissue homogenates, p16(INK4a) promoter methylation was seen in 17 of 57 tumors by methylation-specific polymerase chain reaction, and this could be confirmed using DNA from laser-capture microdissected material in 16 of these cases. A total loss of immunohistochemical p16(INK4a) expression was seen in 6 of 17 tumors with promoter methylation. Quantification of immunohistochemical p16(INK4a) expression for the remaining 11 cases revealed statistically lower frequencies of expression as compared with cases without p16(INK4a) promoter methylation. 9p21 allelic loss was observed in 9 cases, but p16(INK4a) expression in these carcinomas was not reduced. Attempted linear regression of p16(INK4a) expression in tumor buds on the degree of tumor budding, as counted on pan-cytokeratin immunostains, did not show a correlation. p16(INK4a) promoter methylation can completely abrogate p16(INK4a) expression in colorectal carcinomas. In many cases, however, it has an appreciable but only modulatory influence on p16(INK4a) expression. Possibly, methylations are heterozygous, and/or mosaic in colorectal carcinomas and/or methylations are not totally stable but can be lost between carcinoma cell replication cycles. Up-regulation of p16(INK4a) does not seem to be a strict requirement for tumor budding, hence, the absence of a correlation.
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Affiliation(s)
- Friedrich Prall
- Institute of Pathology, University of Rostock, D-18057 Rostock, Germany
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17
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Abstract
p16 is a cyclin-dependent kinase-4 inhibitor that is expressed in a limited range of normal tissues and tumors. In recent years, immunohistochemistry with p16 antibodies has been used as a diagnostic aid in various scenarios in gynecologic pathology. Diffuse (as opposed to focal) positivity with p16 in the cervix can be regarded as a surrogate marker of the presence of high-risk human papillomavirus (HPV). In cervical squamous lesions, p16 is positive in most high-grade cervical intraepithelial neoplasia (CIN) and in some cases of low-grade CIN, usually those associated with high-risk HPV. p16 may be useful to identify small focal high-grade CIN lesions, to distinguish some cases of CIN involving immature metaplastic squamous epithelium from immature metaplastic squamous epithelium not involved by CIN and to distinguish high-grade CIN from benign mimics. Most cervical carcinomas of squamous, glandular, and small cell type are p16-positive. In cervical glandular lesions, p16 is useful, as part of a panel, in the distinction between adenocarcinoma in situ (diffusely positive) and benign mimics, including tuboendometrial metaplasia and endometriosis, which are usually p16-negative or focally positive. p16 may be used, in combination with other markers, to distinguish between a cervical adenocarcinoma (diffuse positivity) and an endometrioid-type endometrial adenocarcinoma (negative or focally positive). Some uterine serous carcinomas are diffusely positive. In the vulva, p16 is positive in HPV-associated vulval intraepithelial neoplasia (VIN) but negative in VIN not associated with HPV. Similarly, HPV-associated invasive squamous carcinomas are p16-positive, whereas the more common non-HPV-associated neoplasms are largely negative or focally positive. In the uterus, p16 positivity is more common and widespread in leiomyosarcomas than leiomyomas, and this may be a useful aid to diagnosis, although problematic uterine smooth muscle neoplasms have not been extensively studied. Metastatic cervical adenocarcinomas in the ovary are usually diffusely p16-positive, and because these may closely mimic a primary ovarian endometrioid or mucinous adenocarcinoma, this may be a valuable diagnostic aid, although p16 expression in primary ovarian adenocarcinomas of these morphologic subtypes has not been widely investigated. Some ovarian serous carcinomas, similar to their uterine counterparts, are p16-positive.
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MESH Headings
- Adenocarcinoma/chemistry
- Adenocarcinoma/diagnosis
- Adenocarcinoma/genetics
- Biomarkers, Tumor/analysis
- Biomarkers, Tumor/genetics
- Carcinoma, Small Cell/chemistry
- Carcinoma, Small Cell/diagnosis
- Carcinoma, Small Cell/genetics
- Cyclin-Dependent Kinase Inhibitor p16/analysis
- Cystadenocarcinoma, Serous/chemistry
- Cystadenocarcinoma, Serous/diagnosis
- Cystadenocarcinoma, Serous/genetics
- Diagnosis, Differential
- Endometrial Neoplasms/chemistry
- Endometrial Neoplasms/diagnosis
- Endometrial Neoplasms/genetics
- Female
- Genes, p16
- Genital Neoplasms, Female/chemistry
- Genital Neoplasms, Female/diagnosis
- Genital Neoplasms, Female/genetics
- Genitalia, Female/chemistry
- Genitalia, Female/physiopathology
- Humans
- Immunohistochemistry
- Ovarian Neoplasms/chemistry
- Ovarian Neoplasms/diagnosis
- Ovarian Neoplasms/genetics
- Tumor Suppressor Proteins/analysis
- Tumor Suppressor Proteins/genetics
- Uterine Cervical Neoplasms/chemistry
- Uterine Cervical Neoplasms/diagnosis
- Uterine Cervical Neoplasms/genetics
- Uterine Neoplasms/chemistry
- Uterine Neoplasms/diagnosis
- Uterine Neoplasms/genetics
- Vulvar Neoplasms/chemistry
- Vulvar Neoplasms/classification
- Vulvar Neoplasms/diagnosis
- Vulvar Neoplasms/genetics
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Affiliation(s)
- Ciaran J O'Neill
- Department of Pathology, Royal Group of Hospitals Trust, Grosvenor Road, Belfast BT12 6BL, Northern Ireland
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18
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Zhu X, Zeisel SH. Gene expression profiling in phosphatidylethanolamine N-methyltransferase knockout mice. ACTA ACUST UNITED AC 2005; 134:239-55. [PMID: 15836921 DOI: 10.1016/j.molbrainres.2004.10.040] [Citation(s) in RCA: 10] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/18/2004] [Revised: 10/08/2004] [Accepted: 10/24/2004] [Indexed: 01/06/2023]
Abstract
Choline is derived from the diet as well as from de novo methylation of phosphatidylethanolamine catalyzed by phosphatidylethanolamine N-methyltransferase (PEMT). Pemt knockout mice have no endogenous synthesis of choline molecules. We previously reported that these mice have excess S-adenosylmethionine and hypermethylated DNA in brain, as well as increased mitosis in neural progenitor cells of the hippocampus in embryonic day 17 (E17) brain. In the present study, E17 fetal brains and adult brains were harvested and total RNA was extracted. In fetal brain, using gene expression profiling and Significance Analysis of Microarrays, we identified 107 significant genes with increased expression and 379 significant genes with decreased expression. In adult brain, we identified 381 significant genes with increased expression and 1037 significant genes with decreased expression. We observed significant changes in expression of genes regulating cell cycle (such as TP53, Fgf4, and Ing1), differentiation and neurogenesis (such as S100A4 and D14Ws), and phospholipid metabolism (such as Pip5k1a, Pitpn, and Pla2g6) as well as in a number of methyltransferase genes (including Gnmt). Some genes with expression known to be regulated by promoter methylation were suppressed in Pemt knockout brain (such as S100a4 and TP53). These findings are consistent with the biochemical changes that we previous reported in fetal brains from Pemt knockout mice. This is the first report of gene profiling in Pemt(-/-) mouse brain.
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Affiliation(s)
- Xiaonan Zhu
- Department of Nutrition, CB #7461, School of Public Health, School of Medicine, University of North Carolina, Chapel Hill, NC 27599, USA
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19
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An C, Choi IS, Yao JC, Worah S, Xie K, Mansfield PF, Ajani JA, Rashid A, Hamilton SR, Wu TT. Prognostic Significance of CpG Island Methylator Phenotype and Microsatellite Instability in Gastric Carcinoma. Clin Cancer Res 2005. [DOI: 10.1158/1078-0432.656.11.2] [Citation(s) in RCA: 38] [Impact Index Per Article: 1.9] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/16/2022]
Abstract
Abstract
Purpose: The influence of molecular characteristics in prognosis of gastric cancer remains unclear. The aim of this study was to evaluate the prognostic value of the CpG island methylator phenotype (CIMP) and microsatellite instability (MSI) in gastric cancer.
Experimental Design: We studied the methylation profiles of tumor suppressor gene p16, DNA mismatch repair gene hMLH1, and four CpG islands (MINT1, MINT2, MINT25, and MINT31) using bisulfite/methylation–specific PCR, and MSI using five microsatellite markers in 83 resected gastric carcinomas. The CIMP and MSI status were compared with clinicopathologic features and overall survival.
Results: Concordant methylation of multiple genes/loci (CIMP-high) was present in 31% of tumors and in 4% of non-neoplastic mucosa, CIMP-low in 55% and 17%, and CIMP-negative in 13% and 79%, respectively (P < 0.001). The prevalence of MSI-high, MSI-low, and MS-stable in tumor was 19%, 17%, and 64%, respectively. MSI status was closely associated with hMLH1 hypermethylation and CIMP status (P = 0.001). In univariate analysis, overall survival was predicted by pathologic stage (P < 0.0001), R0 resection (P = 0.0002), MINT31 methylation (P = 0.04), and CIMP-high status (P = 0.04). MSI status of tumor was not a significant predictor of prognosis. Although CIMP status seemed to be a prognostic predictor of gastric cancer, only pathologic stage remained a significant predictor of prognosis on multivariate analysis (P < 0.001).
Conclusions: Our results indicate that there is an association between CIMP status and MSI status in gastric cancer. Concordant methylation of multiple genes/loci (CIMP-H) is associated with better survival but is not an independent predictor of prognosis in resected gastric cancer.
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Affiliation(s)
| | | | | | | | | | - Paul F. Mansfield
- 3Surgical Oncology, The University of Texas M.D. Anderson Cancer Center, Houston, Texas
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