Cytogenetic and molecular aspects of gastric cancer: clinical implications

Gastric cancer is of major importance world-wide being the second most common cause of cancer-related death in the world. According to Lauren's histological classification gastric cancer is divided in two groups, the better differentiated intestinal carcinomas and the poorly differentiated diffuse-type cancers. The genetic changes underlying the initiation and progression of gastric cancer are not well defined. Gastric carcinogenesis is a multistep process involving a number of genetic and epigenetic factors. Although it has been proposed that different genetic pathways exist for differentiated and undifferentiated carcinomas, the two histological subtypes of gastric cancer share some common genetic alterations. Currently, tumor histology and pathologic stage are the major prognostic variables used in the clinical practice for gastric cancer patients. However, it is known that tumors with similar morphology may differ in biological aggressiveness, prognosis and response to treatment. Molecular genetic analysis of gastric cancer revealed a number of associations of certain genetic changes with pathological features, tumor biological behavior and prognosis of gastric cancer patients, suggesting that these genetic abnormalities might play an important role in gastric tumorigenesis. Increasing evidence suggests that the molecular genetic changes could be helpful in the clinical setting, contributing to prognosis and management of patients. Regarding epigenetic events in gastric tumorigenesis, a number of methylating markers have been proposed for risk assessment, prognostic evaluation and as therapeutic targets. However, further research is required in order to systematically investigate the genetic changes in gastric cancer estimating also their usefulness in the clinical practice. A good understanding of the genetic changes underlying gastric carcinogenesis may provide new perspectives for prognosis and screening of high risk individuals. Some of the genetic alterations could definitely improve tumor classification and management of gastric cancer patients. Also, based on molecular data identified in gastric cancer novel therapeutics might help to improve the treatment of this disease.     

Search for new biomarkers of gastric cancer through serial analysis of gene expression and its clinical implications

Gastric cancer is one of the most common human cancers and is the second most frequent cause of cancer-related death in the world. Serial analysis of gene expression (SAGE) is a powerful technique to allow genome-wide analysis of gene expression in a quantitative manner without prior knowledge of the gene sequences. SAGE on 5 samples of gastric cancer with different histology and clinical stages have created large SAGE libraries of gastric cancer that enable us to identify new cancer biomarkers. Commonly up-regulated genes in gastric cancer in comparison with normal gastric epithelia included CEACAM6, APOC1 and YF13H12. By comparing gene expression profiles of gastric cancers at early and advanced stages, several genes differentially expressed by tumor stage were also identified, including FUS, CDH17, COL1A1 and COL1A2, which should be novel genetic markers for high-grade malignancy. Regenerating gene type IV (REGIV) is one of the most up-regulated genes in a SAGE library of a scirrhous-type gastric cancer. In vitro studies using RegIV-transfected cells revealed that RegIV is secreted by cancer cells and inhibits apoptosis, suggesting that RegIV may serve as a novel biomarker and therapeutic target for gastric cancer. Production of RNA aptamers could be a useful approach to establish a detection system in blood. A custom-made array, named Ex-STOMACHIP, consisting of 395 genes, including highly differentially expressed genes identified by our SAGE and other known genes related to carcinogenesis and chemosensitivity, is useful to study the molecular pathogenesis of gastric cancer and to obtain information about biological behavior and sensitivity to therapy in the clinical setting. Combined analyses of gene expression profile, genetic polymorphism and genetic instability will aid not only cancer detection, but also characterization of individual cancers and patients, leading to personalized medicine and cancer prevention.     

Molecular-pathological diagnosis of gastric cancer

Gastric cancer is one of the most common human cancers and is the second most frequent cause of cancer-related death in the world. In the course of multistep carcinogenesis of the stomach, various alterations of oncogenes, tumor suppressor genes, DNA repair genes, growth factors/receptors, cell cycle regulators and cell adhesion molecules are accumulated. These can be biological markers of malignancy for molecular-pathological diagnosis. The gene expression profile and scoring system, based on microarray analysis, have great potential for differentiating the character of gene expression in individual cancers and predicting biological behavior and chemosensitivity. Genetic polymorphism is an important determinant of the endogenous cause of cancer and can be used as a marker for gastric cancer risk that will directly connect with personalized cancer prevention. Genetic polymorphisms have been also associated with therapeutic efficacy and toxicity of anti-cancer drugs. Serial analysis of gene expression (SAGE) is a powerful technique to allow genome-wide analysis of gene expression in a quantitative manner. By comparing SAGE libraries of gastric cancer with those of various normal tissues, novel genes specifically expressed in gastric cancer can be selected. If the gene is not expressed in normal crucial organs, the gene can be not only a cancer-biomarker but also a therapeutic target with minimal adverse effects. A custom-made array, based on SAGE libraries, is useful to study the molecular pathogenesis of gastric cancer and obtain information on biological behavior. Combined analyses of gene expression profile and genetic polymorphism will enable the characterization of the individual cancer and person, that is directly connected with personalized medicine and cancer prevention.     

Genome-wide analysis of gene expression in intestinal-type gastric cancers using a complementary DNA microarray representing 23,040 genes

To shed light on mechanisms that underlie development and/or progression of intestinal-type gastric cancer, we compared expression profiles of cancer cells obtained by laser-capture microdissection of 20 intestinal-type gastric tumors with expression of genes in corresponding noncancerous mucosae, by a cDNA microarray consisting of 23,040 genes. We identified 61 genes that were commonly up-regulated and 63 that were commonly down-regulated in the cancer tissues. Altered expression of 12 of those genes was associated with lymph node metastasis. A "predictive score," based on expression profiles of five of the genes that were able to distinguish tumors with metastasis from node-negative tumors in our panel, correctly diagnosed the lymph node status of nine additional gastric cancers. This genome-wide information contributes to an improved understanding of molecular changes during the development of intestinal-type gastric cancers. It may help clinicians predict metastasis to lymph nodes and assist researchers in identifying novel therapeutic targets for this type of cancer.     

S100A11 gene identified by in-house cDNA microarray as an accurate predictor of lymph node metastases of gastric cancer

Gastric cancer is one of the most common malignancies in the world, and in Asian countries its incidence and mortality rates are very high. Worldwide, Japan ranks first in the incidence of this type of cancer for both sexes. To shed light on the mechanisms underlying the development and/or progression of gastric cancer, we compared the expression profiles in gastric cancer cells obtained from surgical dissection of 20 gastric adenocarcinoma specimens with those in the corresponding non-cancerous mucosa, by cDNA microarray analysis. In total, 8,000 cDNA clones were randomly picked up and their 5'-end nucleotide sequences were determined. On the basis of sequence information, 4,608 independent clones were selected and used to produce the cDNA microarray. We identified 26 genes that were commonly up-regulated and 44 genes that were commonly down-regulated in cancerous tissues. To validate the cDNA microarray analysis, real-time PCR was performed. We found that gene S100A11 expression was associated with the development of lymph node metastases. S100A11 gene expression was clearly up-regulated in specimens from patients with lymph node metastases relative to those from patients without lymph node metastases. S100A11 gene expression status was useful to distinguish gastric cancers with lymph node metastases from those without lymph node metastasis. This genome-wide information contributes to an improved understanding of molecular changes during the development of gastric cancers. It may also help clinicians predict the development of lymph node metastases and assist researchers in identifying novel therapeutic targets for patients with gastric cancer.     

Molecular biology of gastric cancer

Abstract Despite its decreasing incidence overall, gastric cancer is still a challenging disease. Therapy is based mainly upon surgical resection when the tumour remains localised in the stomach. Conventional chemotherapy may play a role in treating micrometastatic disease and is effective as palliative therapy for recurrent or advanced disease. However, the knowledge of molecular pathways implicated in gastric cancer pathogenesis is still in its infancy and the contribution of molecular biology to the development of new targeted therapies in gastric cancer is far behind other more common cancers such as breast, colon or lung. This review will focus first on the difference of two well defined types of gastric cancer: intestinal and diffuse. A discussion of the cell of origin of gastric cancer with some intriguing data implicating bone marrow derived cells will follow, and a comprehensive review of different genetic alterations detected in gastric cancer, underlining those that may have clinical, therapeutic or prognostic implications. *Supported by an unrestricted educational grant from Sanofi-Aventis.     

The aberrant p53 protein (review)

According to the current concept of carcinogenesis, neoplastic transformation consists of multistep accumulations of adverse genetic and epigenetic events. Recent advances in molecular genetics have demonstrated aberrations of oncogenes and tumor-suppressor genes in a variety of human cancers. The loss of wild-type p53 gene expression has exceptionally been implicated in the development of a wide variety of human cancers and it is generally accepted that p53 is a component in biochemical pathways central to human carcinogenesis. Although the role of the p53 gene in cancer genesis and development has fueled as many questions, study of p53 has come to the forefront of cancer research and detection of its abnormalities during the development of tumors may have diagnostic, prognostic and therapeutic implications. To be of value in clinical practice, immunohistochemical assessment of p53 protein should provide clinically relevant information. The degree of concordance between p53 gene mutation and the accumulation of p53 protein cannot be perfect, however, the immunohistochemical assay using anti-p53 antibodies is the most widely applicable approach for detection of tumors in routine investigations, particularly with regard to diagnosis or prognosis.     

p53 and DPC4 alterations in the bile of patients with pancreatic carcinoma

INTRODUCTION: Pancreatic cancer is still predominantly diagnosed in advanced stages, and 85%-90% of patients are not eligible for surgery at diagnosis. This is mainly due to the great difficulty in detecting the tumour at an early stage and presently no satisfactory results have been obtained to overcome this problem. Studies on molecular genetic profile of pancreatic cancer may represent an important approach. This study was focused on the mutations of p53 and DPC4 detectable in the bile of patients with histologically proven pancreatic cancers. MATERIALS AND PATIENTS: We analysed specimens of bile collected through percutaneous transhepatic biliary catheters, placed to treat malignant biliary obstruction in 25 patients with pancreatic adenocarcinoma. A percentage of mutation was obtained of 43 % for the microsatellite D17S945 (p53), 54% and 50 % for D18S46 and D18S474 (DPC4), respectively. The percentage of amplification was 67%, 93,6%, and 80%. CONCLUSION: We consider the results encouraging enough to decide to enlarge the number of patients examined. The aim is to determine if a test for DPC4 and p53 mutations is eligible for introduction in clinical routine use. More sets of samples are required to satisfactorily answer this question.     

Abnormalities affecting the APC and MCC tumour suppressor gene loci on chromosome 5q occur frequently in gastric cancer but not in pancreatic cancer

Abnormalities affecting tumour suppressor genes on chromosome 5q21 are increasingly recognised as important in the pathogenesis of a variety of human cancers, particularly of the gastrointestinal tract. We have examined a series of gastric and pancreatic cancers from European patients for loss of heterozygosity (LOH) of markers within and around the APC and MCC genes on chromosome 5q2t using restriction fragment length polymorphism and polymerase chain reaction techniques. We find that LOH of the APC and MCC genes is particularly frequent in gastric cancers of diffuse type, but very infrequent in pancreatic cancers. We have also used single-strand conformational polymorphism to screen for abnormalities of the sequence of the APC and MCC genes in a panel of pancreatic cancer cell lines. Our results suggest that there are distinct differences in the molecular pathogenesis of gastric and pancreatic cancer and that abnormalities of APC and MCC may be involved particularly in the diffuse type of gastric cancer.     

Genetic alterations in gliomas

Conclusion It would seem that distinct genetic changes in different genes, the protein products of which interact in particular growth control mechanism may lead to the same cellular abnormality. This is exemplified in the case of the gliomas by the ways in which p53 and the Rb1 phosphorylation mechanisms may be rendered impotent. It seems likely that many further genetic abnormalities affecting genes coding for proteins, either involved in the cellular mechanisms identified or in new growth control mechanisms, will be found in the future. In total our results indicate both differences and similarities between the molecular genetic alterations in tumors with oligodendroglial and astrocytic differentiation. The loss of genetic information from 19q and 1p as well as the rareness ofTP53 mutations in oligodendroglial tumors suggests that the early events in their oncogenesis are distinct from those associated with astrocytic tumors. However, similarities are indicated by the allelic losses on 9p and 10 in the anaplastic tumors, suggesting the utilization of common pathways of progression.     

Molecular biology in gastric cancer

In gastric cancer, the process of carcinogenesis is thought to occur as a stepwise accumulation of genetic abnormalities. However, the mechanisms of the process of multistage carcinogenesis is still unknown for gastric cancer. Gene abnormalities seen in gastric cancer, including ras, myc, c-erbB-2, met, K-sam and cript are summarized herein. Abnormalities of cancer suppressor genes, including p53, RB and APC are also described. In our studies, the biological malignancy of patients with c-erbB-2 amplification was higher than that of patients without amplification. Moreover, the cases with amplification of c-erbB-2 were found to be highly correlated with distant organ metastasis. However, very little is currently known of the molecular abnormalities leading to gastric cancer. In order to clarify the multiple gene abnormalities in gastric cancer, we used the method of restriction landmark genomic scanning (RLGS). RLGS provides a useful method for genomic analysis of gastric cancer. In the future, new analytical methods that will permit screening of all gene abnormalities at once promise to improve our understanding of the mechanisms of gastric cancer.     

Genetic detection of free cancer cells in the peritoneal cavity of the patient with gastric cancer: present status and future perspectives

The purpose of this review is to examine the current status and future perspectives of the molecular analysis of peritoneal lavage fluid in patients with gastric cancer. During the past 10 years, the polymerase chain reaction (PCR) has been applied for the molecular detection of free cancer cells in the abdominal cavity of patients with gastric cancer, and its clinical significance in establishing the presence of peritoneal dissemination has been assessed by several groups especially in Japan. The majority of these studies have confirmed the predictive value of the molecular detection of peritoneal metastasis and recurrence using peritoneal lavage fluid. Based on these findings, since April 2006, the genetic diagnosis of body fluids has been included in the Japanese Government public health insurance program for patients with solid tumors. However, there are still many obstacles to overcome before the genetic diagnosis of micrometastasis can be considered a routine laboratory assay. Here we review the importance of the molecular detection of cancer cells in the abdominal cavity, and the molecular techniques used for such diagnosis; we also provide some clinical examples to illustrate the value of molecular diagnosis.     

Prognostic significance of aberrant gene methylation in gastric cancer

Promoter methylation acts as an important alternative to genetic alterations for gene inactivation in gastric carcinogenesis. Although a number of gastric cancer-associated genes have been found to be methylated in gastric cancer, valuable methylation markers for early diagnosis and prognostic evaluation of this cancer remain largely unknown. In the present study, we used methylation-specific PCR (MSP) to analyze promoter methylation of 9 gastric cancer-associated genes, including MLF1, MGMT, p16, RASSF2, hMLH1, HAND1, HRASLS, TM, and FLNc, and their association with clinicopathological characteristics and clinical outcome in a large cohort of gastric cancers. Our data showed that all of these genes were aberrantly methylated in gastric cancer, ranging from 8% to 51%. Moreover, gene methylation was strongly associated with certain clinicopathological characteristics, such as tumor differentiation, lymph node metastasis, and cancer-related death. Of interest, methylation of MGMT, p16, RASSF2, hMLH1, HAND1, and FLNc was closely associated with poor survival in gastric cancer, particularly MGMT, p16, RASSF2 and FLNc. Thus, our findings suggested these epigenetic events may contribute to the initiation and progression of gastric cancer. Importantly, methylation of some genes were closely relevant to poor prognosis in gastric cancer, providing the strong evidences that these hypermethylated genes may be served as valuable biomarkers for prognostic evaluation in this cancer.     

Integrated gene copy number and expression microarray analysis of gastric cancer highlights potential target genes

We performed an integrated array comparative genomic hybridization (aCGH) and expression microarray analysis of 8 normal gastric tissues and 38 primary tumors, including 25 intestinal and 13 diffuse gastric adenocarcinomas to identify genes whose expression is deregulated in association with copy number alteration. Our aim was also to identify molecular genetic alterations that are specific to particular clinicopathological characteristics of gastric cancer. Distinct molecular genetic profiles were identified for intestinal and diffuse gastric cancers and for tumors obtained from 2 different locations of the stomach. Interestingly, the ERBB2 amplification and gains at 20q13.12-q13.33 almost exclusively discriminated intestinal cancers from the diffuse type. In addition, the 17q12-q25 gain was characteristic to cancers located in corpus and the 20q13.12-q13.13 gain was more common in the antrum. Statistical analysis was performed using integrated copy number and expression data to identify genes showing differential expression associated with a copy number alteration. Genes with the highest statistical significance included ERBB2, MUC1, GRB7, PPP1R1B and PPARBP with concomitant changes in copy number and expression. Immunohistochemical analysis of ERBB2 and MUC1 on a tissue microarray containing 78 independent gastric tissues showed statistically significant differences (p < 0.05 and <0.001) in immunopositivity in the intestinal (31 and 70%) and diffuse subtypes (14 and 41%), respectively. In conclusion, our results demonstrate that intestinal and diffuse type gastric cancers as well as cancers located in different sites of the stomach have distinct molecular profiles which may have clinical value.     

Gastroscopy plus cytology for the diagnosis of gastric cancer or precancerous diseases--report of 127 cases

One hundred and twenty-seven patients with gastric tumor or precancerous diseases were examined by gastroscopy plus cytology, and the results were compared with that of histopathology after operation. Out of 127 cases, 70 were diagnosed as gastric cancer (15 cases as early cancer, 55 as advanced cancer). The positive rate for early cancer was 21.4%, it comprised 11.2% of all gastric cancers diagnosed by gastroscopy during the same period. The preoperative positive rate of early gastric cancer was 100% (13/13) as examined by the combination of touch, brush cytology and mucosal biopsy. The combined method may increase the positive rate and the accuracy, not only for advanced but also for early gastric cancers. It is suggested that the combination of these methods be routinely used in the diagnosis of suspicious cancer in the gastric mucosa.     

核糖体蛋白S24基因在胃癌及其癌前病变中差异表达的研究

背景与目的:将胃癌组织、正常胃黏膜及胃癌前病变的基因表达进行比较,找出与人胃癌密切相关的基因片段,为进一步探讨胃癌的发生机制、胃癌的早期诊断和治疗方案提供重要的理论根据。材料与方法:应用荧光mRNA差异显示技术(FluorescentmRNAdifferentialdisplay,FDD)分析3例胃癌、3例正常胃粘膜、3例胃癌前病变组织,分析基因差异表达情况。分离差异表达基因片段,进行PCR扩增。将cDNA片段测序,测序结果提交Genbank,经BLAST软件检索与同源性分析。选取其中差异表达的核糖体蛋白S24(RibosomalproteinS24,RPS24)基因,应用Northern杂交进行验证,并进一步对RPS24基因进行基因表达系列分析和组织表达谱分析。结果:mRNA差异显示发现RPS24基因在胃癌组织中的表达明显高于癌前病变和正常胃黏膜组织,并且Northern印迹验证阳性。基因表达系列分析和组织表达谱分析发现RPS24基因在多种肿瘤组织中表达增高以及分布广泛。结论:与非癌组织相比,RPS24基因在胃癌中表达明显增高,RPS24的高表达可能与胃癌的发生、恶性进展有关。     

Molecular biology of gastric cancer

Gastric cancer is one of the leading causes of cancer mortality in the world. Gastric adenocarcinomas account for more than 95% of gastric tumors, whereas gastrointestinal stromal tumors (GISTs) are the most common neoplasms of the rare gastric mesenchymal tumors. Although the incidence of mid-distal gastric adenocarcinomas is decreasing, the incidence of gastroesophageal junctional tumors and Barrett's adenocarcinomas is increasing for unknown reasons. The majority of gastric tumors are sporadic in nature. However, there are rare, inherited gastric cancer predisposition traits, such as germline p53 (Li-Fraumeni syndrome) as well as E-cadherin (CDH1) alterations in familial diffuse gastric cancers. Gastric cancer has been observed to be part of the spectrum of neoplasms associated with germline mismatch repair gene (MMR) alterations that give rise to the hereditary nonpolyposis colorectal cancer (HNPCC) entity. Comparative genomic hybridization analyses have identified several amplifications and losses of DNA copy numbers in gastric cancers. Loss of heterozygosity (LOH) studies have shown several chromosomal loci with significant allelic loss, thus indicating the possibility of harboring a tumor suppressor gene important in gastric tumorigenesis. Microsatellite instability (MIS) and associated alteration of the TGF-bIIR, IGFRII, BAX, E2F-4, hMSH3, and hMSH6 genes are found in a subset of gastric carcinomas. Cell adhesion molecule abnormalities such as those involving CDH1 may play an important role in diffuse-type gastric cancer development. Although, multiple somatic alterations have been described in gastric carcinomas at the molecular level, the significance of these changes in gastric tumorigenesis remains to be established in most instances. The critical molecular alterations in gastric cancers that may lead to advances in our armamentarium to combat this lethal disease remain to be fully characterized.     

Inherited predisposition to glioma

In gliomas, germline gene alterations play a significant role during malignant transformation of progenitor glial cells, at least for families with occurrence of multiple cancers or with specific hereditary cancer syndromes. Scientific evidence during the last few years has revealed several constitutive genetic abnormalities that may influence glioma formation. These germline abnormalities are manifested as either gene polymorphisms or hemizygous mutations of key regulatory genes that are involved either in DNA repair or in apoptosis. Such changes, among others, include hemizygous alterations of the neurofibromatosis 1 (NF1) and p53 genes that are involved in apoptotic pathways, and alterations in multiple DNA repair genes such as mismatch repair (MMR) genes, x-ray cross-complementary genes (XRCC), and O6-methylguanine-DNA methyltransferase (MGMT) genes. Subsequent cellular changes include somatic mutations in cell cycle regulatory genes and genes involved in angiogenesis and invasion, leading eventually to tumor formation in various stages. Future molecular diagnosis may identify new genomic regions that could harbor genes important for glioma predisposition and aid in the early diagnosis of these patients and genetic counseling of their families.