Genetic aberrations detected by comparative genomic hybridization in biliary tract cancers.

In order to elucidate cytogenetic changes characteristic of biliary tract cancer, we examined the genetic imbalances in 18 biliary tract cancers using comparative genomic hybridization (CGH). The most common sites of increases in copy number, in order of frequency, were 17q (33% of the cases), 5p (28%), 3q (22%), 7p (22%), 8q (22%), and 12p (22%), whereas copy number decreases of 6q (28%), 18q (28%), 4q (22%), 5q (22%), and 9p (22%) were frequent. The average number of chromosomal aberrations was significantly greater in stage IV than in stage III tumors (7.9 vs. 2.2/tumor, p < 0.05). The frequent aberrations detected in this study may be related to the development and/or progression of biliary tract cancers. This is the first report on CGH of biliary tract cancers.     

Genetic aberrations detected by comparative genomic hybridisation in vulvar cancers

Squamous cell carcinoma of the vulva is a disease of significant clinical importance, which arises in the presence or absence of human papillomavirus. We used comparative genomic hybridisation to document non-random chromosomal gains and losses within human papillomavirus positive and negative vulvar cancers. Gain of 3q was significantly more common in human papillomavirus-positive cancers compared to human papillomavirus-negative cancers. The smallest area of gain was 3q22-25, a chromosome region which is frequently gained in other human papillomavirus-related cancers. Chromosome 8q was more commonly gained in human papillomavirus-negative compared to human papillomavirus-positive cancers. 8q21 was the smallest region of gain, which has been identified in other, non-human papillomavirus-related cancers. Chromosome arms 3p and 11q were lost in both categories of vulvar cancer. This study has demonstrated chromosome locations important in the development of vulvar squamous cell carcinoma. Additionally, taken together with previous studies of human papillomavirus-positive cancers of other anogenital sites, the data indicate that one or more oncogenes important in the development and progression of human papillomavirus-induced carcinomas are located on 3q. The different genetic changes seen in human papillomavirus-positive and negative vulvar squamous cell carcinomas support the clinicopathological data indicating that these are different cancer types.     

Additional chromosomal abnormalities detected by array comparative genomic hybridization in AML

Improved outcome of acute myeloid leukemia (AML) depends on the better differentiation of subtypes to predict treatment response and the identification of new target for treatment. In this study, array comparative genomic hybridization (aCGH) was used to distinguish eight cases of AML cases. Validation was performed by FISH and quantitative genomic PCR. The aCGH revealed new large and small recurrent genomic imbalances, such as gains of 1p36, 10q26, 11p15, 20q13, 22q23, harboring many proto-oncogenes. These results better define genetically the studied cases and could be used to understand the multiple phenomena involved in leukemogenesis.     

Genetic aberrations detected by comparative genomic hybridization in ovarian clear cell adenocarcinomas

Genetic abnormalities were detected by comparative genomic hybridization (CGH) in 12 ovarian clear cell adenocarcinomas. DNA sequence copy number abnormalities (CNAs) occurring in more than 20% of the cancers included increased copy numbers of 8q11-q13, 8q21-q22, 8q23, 8q24-qter, 17q25-qter, 20q13-qter and 21q22-qter and reduced copy numbers of 19p. Increases in copy numbers of 8q11-q13, 8q21-q22, 8q23 and 8q24-qter occurred more frequently in disease-free patients than in recurrent/non-surviving patients (p < 0.05). However, increases in copy numbers of 17q25-qter and 20q13-qter occurred more frequently in recurrent/non-surviving patients than in disease-free patients (p < 0.05). Furthermore, increases in copy numbers of 17q25-qter and 20q13-qter occurred together (p < 0.05). Additionally, there were negative correlations between increases in copy numbers of 8q21-q22 and 17q25-qter, and between 8q21-q22 and 20q13-qter (p < 0.05). It appears that ovarian clear cell adenocarcinomas can be classified into two subtypes, one being cancer with an increase in copy numbers of 8q and the other being cancer with increases in copy numbers of 17q25-qter and 20q13-qter.     

Interglandular cytogenetic heterogeneity detected by comparative genomic hybridization in pancreatic cancer

The aim of this study is to explore the mechanisms of intratumoral cytogenetic heterogeneity (ICH) in pancreatic cancer. Using comparative genomic hybridization (CGH) analysis, we investigated interglandular variation in 20 primary invasive ductal adenocarcinomas of the pancreas. Three or four adjacent neoplastic glands were individually microdissected from a tumor specimen. Extracted DNA from each gland was amplified by degenerate oligonucleotide primed-PCR, followed by CGH. In addition, DNA index (DI) was measured by laser scanning cytometry in each case. CGH profiles displayed a wide variety of differences between glands within the same tumor in all cases, i.e., interglandular cytogenetic heterogeneity was distinct in pancreatic cancers. In this study, genetic changes detected in all regions of a tumor were classified as "region-independent" alterations, whereas changes seen in at least one, but not all regions were designated as "region-dependent" alterations, which resulted in ICH. The degree of ICH, which was manifested as the ratio of these two types of alterations, correlated closely with DI (Spearman rho = 0.842; P = 0.0002). Therefore, DI might be a surrogate marker for ICH. These results suggest that with tumor progression, ICH and DNA aneuploidy result from the successive appearance of region-dependent alterations attributable to chromosomal instability in tumor cells. Our data support a concept of individual cell heterogeneity in pancreatic cancer.     

Progressive genetic aberrations detected by comparative genomic hybridization in squamous cell cervical cancer

Genetic changes orchestrated by human papillomaviruses are the most important known factors in carcinogenesis of the uterine cervix. However, it is clear that additional genetic events are necessary for tumour progression. We have used comparative genomic hybridization to document non-random chromosomal gains and losses within a subset of 37 cervical carcinomas matched for clinical stage Ib, but with different lymph node status. There were significantly more chromosomal changes in the primary tumours when the lymph nodes were positive for metastases. The most frequent copy number alterations were loss of 3p, 11q, 6q and 10q and gain of 3q. The smallest areas of loss and gain on chromosome 3 were 3p14-22 and 3q24-26. The study identifies progressive DNA copy number changes associated with early-stage invasive cervical cancers with and without lymph node metastases, a factor of potential prognostic and therapeutic value.     

Chromosomal aberrations in colorectal cancers and liver metastases analyzed by comparative genomic hybridization.

Comparative genomic hybridization (CGH) was used to screen for changes in the number of DNA sequence copies in 30 primary colorectal cancers and 16 liver metastases, to identify regions that contain genes important for the development and progression of colorectal cancer. In primary colorectal cancer, we found frequent gains at 7p21 (36.7%), 7q31-36 (30%), 8q23-24 (43.0%), 12p (30%), 14q24-32 (33.3%), 16p (40.0%), 20p (33.3%), 20q (63.3%) and 21q (36.3%), while loss was often noted at 18q12-23 (36.7%). In metastatic tumors, there were significantly more gains and losses of DNA sequences than in primary tumors, with gains at 8q23-24 (found in 62.5% of recurrences vs. 43.0% of primary tumors), 15q21-26 (37.5% vs. 20.0%), 19p (43.8% vs. 20.0%) and 20q (81.3% vs. 63.3%) and losses at 18q12-23 (50.0% vs. 36.7%). The pattern of genetic changes seen in metastatic tumors, with frequent gains at 8q23-24 and 20q and loss at 18q12-23, suggests the progression of colorectal cancer. We investigated a clinical follow-up study for all patients examined by CGH and directed our attention to the genetic changes consisting of gains at 8q and 20q. The incidence of liver metastases was higher in patients with primary colorectal cancer with these genetic changes. Gains at 8q and 20q might be useful to identify patients at high risk for developing liver metastases.     

Chromosomal aberrations detected by comparative genomic hybridization (CGH) in human astrocytic tumors

We investigated chromosomal aberrations in 16 patients with astrocytic tumors of various histologic malignancies by comparative genomic hybridization (CGH). The degree of chromosomal loss was shown to be negatively correlated with histologic malignancy. Losses of portions of chromosomes 1p, 19q and 22q were the three chromosomal aberrations observed most frequently. Alterations in multiple chromosomes were observed more frequently in glioblastomas than in astrocytomas or anaplastic astrocytomas (P < 0.001). Primary glioblastomas showed a high frequency of genomic DNA gains (5/7), whereas recurrent glioblastomas from anaplastic astrocytomas did not (0/3). We found CGH to be a powerful tool for surveying DNA alterations in tumors and characterizing the biology of tumors of astrocytic lineage.     

Chromosomal aberrations detected by comparative genomic hybridization predict outcome in patients with colorectal carcinoma

To obtain comprehensive information regarding the correlation between genomic changes and clinicopathological parameters such as disease stage, metastases, and survival, we investigated genomic changes by comparative genomic hybridization (CGH) in 73 patients with colorectal cancer (CRC), and assessed the associations of such charges with clinicopathological parameters. Gains of 8q21-22, 13q21-31 and 20q12-qter and loss of 17p12-pter were detected in >50% of stage I tumors. Gain of 8q23-qter and losses of 8p12-pter and 18q12-qter were observed more frequently in stage III/IV tumors than in stage I tumors (all P<0.05). Loss of 8p12-pter and gain of 8q23-qter were linked to nodal metastasis (all P<0.05). Loss of 18q12-qter and gain of 8q23-qter were associated with distant organ metastasis at diagnosis and/or recurrence after surgery (all P<0.05). Moreover, losses of 8p12-pter and 18q12-qter and gains of 8q23 and 8q24-qter were associated significantly with unfavorable prognosis (all P<0.05). Furthermore, combined examination of the above four changes can provide a more accurate assessment for patient's prognosis. Specifically, 11 of 19 patients with these four changes died, but only 1 of 21 cases without these four changes died during the follow-up period (P<0.0001). Multivariate analysis revealed that loss of 18q12-qter is an independent prognostic marker (P=0.031). Our findings indicate that genetic aberrations detected by CGH may predict outcome in patients with CRC.     

Detection of genetic alterations in pancreatic cancers by comparative genomic hybridization coupled with tissue microdissection and degenerate oligonucleotide primed polymerase chain reaction

The aim of this study was to elucidate cytogenetic changes in pancreatic cancers (PCs) and to examine their clinical implications. We screened for genetic alterations in 32 primary PCs including 4 cases with distant organ metastasis using comparative genomic hybridization coupled with tissue microdissection and degenerate oligonucleotide primed polymerase chain reaction (DOP-PCR). The present study revealed frequent gains of chromosomes 13q and 15q and a loss of Xq in addition to a high prevalence of chromosomal imbalances. The average number of total genetic alterations and gains tended to be higher in N1 tumors (TNM classification) than in N0 tumors. The average number of amplifications was significantly higher in M1 tumors than in M0 tumors (p = 0.024). Gain/amplification of 20q was more frequently observed in M1 tumors than in M0 tumors (p = 0.016), and this change was also detected in all of 4 distant metastatic lesions. Losses of 6q, 8p, 9p, 17p, and 18q were recurrent in N0 and M0 tumors, and these alterations were also retained in N1 and M1 tumors. These observations suggest that these genetic losses contribute to the development of PCs and that increases in the DNA copy number confer an aggressive character on cancer cells. Especially, gain/amplification of 20q was associated with the potential of distant organ metastasis of tumor cells.     

采用对比基因杂交技术建立嗅神经母细胞瘤基因畸变模型

目的建立嗅神经母细胞瘤(ENB)的基因畸变模型并分析其特征。方法采用对比基因杂交(CGH)技术对12例原发、3例复发、7例转移ENB标本进行基因检测,用特殊软件系统对所采集照片的荧光信号进行量化分析,确定肿瘤DNA与正常对照DNA之间的基因差别。结果ENB常见DNA丢失主要见于1P、2q、3p／q、4p／q、5p／q、6q、8p／q、9p、10p／q、11P、12q、13q、18q和21q,DNA过度表达见于1p、7q、9q、1lq、14q、16p／q、17p／q、19p／q、20p／q和22p／q。ENB的1p21-p31DNA丢失与该类肿瘤患者预后差有明显相关性。死于ENB的患者均具有以下共性：1p21-p31DNA丢失、临床分期为C或D期、分化程度低(Ⅲ或Ⅳ级)。对4例发生转移、复发的患者,将检测结果与其原发病灶基因畸变情况进行对比分析表明,转移、复发病灶与其原发病灶间具有高度的克隆一致性。结论采用CGH能建立ENB的典型基因畸变模型,该模型有助于解释此肿瘤的生物学特性并对其预后进行评估,并与神经母细胞瘤、小细胞肺癌以及头颈部鳞癌进行鉴别。     

High incidence of unbalanced chromosomal changes in mantle cell lymphoma detected by comparative genomic hybridization

Comparative genomic hybridization (CGH) was carried out in 30 mantle cell lymphoma (MCL) patients at the time of diagnosis. CGH results were supported by conventional cytogenetics (CC), FISH, molecular genetic PCR methods and 2 patients were examined by array CGH. Using all cytogenetic, molecular cytogenetic and PCR methods, chromosomal changes were detected in 28 (93%) patients. Using CGH, unbalanced chromosomal changes were detected in 24 (80%) cases. The most frequent aberrations were losses of 1p (8 cases), 8p (10 cases), 9q (6 cases), 11q (11 cases), 13q (10 cases) and 17p (9 cases), and gains of chromosome 3 and 3q (12 cases) and 8q (7 cases). Total number of 60 gains and 116 losses were detected. The primary chromosomal change t(11;14) was detected using FISH and/or PCR in 20 (66.6%) patients, and in 9 of them, the breakpoint was determined using PCR in the major translocation cluster (MTC). The evaluation of the frequencies of CGH changes in groups of patients with and without t(11;14) revealed the differences only in losses 6q and 9q, which were only found in patient with t(11;14). An important result was obtained using array CGH method. In a patient without the primary t(11;14), the gain of CCND1 gene was found. Our results show high heterogeneity of the additional chromosomal changes in MCL cases, which involved specific chromosomal subregions. We did not confirm the importance of subdividing of MCL cases with and without t(11;14). Also, statistical significance in survival rates between both subgroups was not confirmed.     

Detection of chromosomal aberrations by comparative genomic hybridization during transformation of human breast epithelial cells in vitro

Breast cancer is the most frequent malignancy in women. It is well recognized that tumorigenesis is a multistep process resulting from the accumulation of sequential genetic alterations. In breast cancers LOH has been described on one or both arms of multiple chromosomes. Comparative genomic hybridization (CGH) analysis was performed to identify chromosomal imbalances in the breast epithelial cells (HBEC). We have used a human in vitro-in vivo system in which the environmental carcinogen benz(a)pyrene (BP) and the c-Ha-ras oncogene were utilized for inducing in vitro transformation of HBEC. Immortal MCF-10F cells were treated with BP which resulted in the transformed cell line BP-1 that was further enhanced by transfection with the c-Ha-ras to generate the cell line BP-1-Tras. This cell line is tumorigenic when injected in severe combined immunodeficient (SCID) mice, generating the tumor cell line BP-1-Tras T J#4. Our comparative genomic hybridization analysis indicates that the most overrepresented segment after cell transformation and in the BP-1, BP-1-Tras and in the tumor cell line were 1p (80%), 5q21-ter (80%), 8q24.1 (90%) and Xq27-28 (60%). DNA sequence amplification at 10p14-15 was observed in BP-1-Tras T J#4 cells. Allelic losses of chromosome 4, 8p11-21 and 15q11-12, occur after cell transformation and are maintained consistently during tumorigenesis.     

Distinct genomic profiles in hereditary breast tumors identified by array-based comparative genomic hybridization

Mutations in BRCA1 and BRCA2 account for a significant proportion of hereditary breast cancers. Earlier studies have shown that inherited and sporadic tumors progress along different somatic genetic pathways and that global gene expression profiles distinguish between these groups. To determine whether genomic profiles similarly discriminate among BRCA1, BRCA2, and sporadic tumors, we established DNA copy number profiles using comparative genomic hybridization to BAC-clone microarrays providing <1 Mb resolution. Tumor DNA was obtained from BRCA1 (n = 14) and BRCA2 (n = 12) mutation carriers, as well as sporadic cases (n = 26). Overall, BRCA1 tumors had a higher frequency of copy number alterations than sporadic breast cancers (P = 0.00078). In particular, frequent losses on 4p, 4q, and 5q in BRCA1 tumors and frequent gains on 7p and 17q24 in BRCA2 tumors distinguish these from sporadic tumors. Distinct amplicons at 3q27.1-q27.3 were identified in BRCA1 tumors and at 17q23.3-q24.2 in BRCA2 tumors. A homozygous deletion on 5q12.1 was found in a BRCA1 tumor. Using a set of 169 BAC clones that detect significantly (P < 0.001) different frequencies of copy number changes in inherited and sporadic tumors, these could be discriminated into separate groups using hierarchical clustering. By comparing DNA copy number and RNA expression for genes in these regions, several candidate genes affected by up- or down-regulation were identified. Moreover, using support vector machines, we correctly classified BRCA1 and BRCA2 tumors (P < 0.0000004 and 0.00005, respectively). Further validation may prove this tumor classifier to be useful for selecting familial breast cancer cases for further mutation screening, particularly, as these data can be obtained using archival tissue.     

Chromosomal aberrations in human hepatocellular carcinomas associated with hepatitis C virus infection detected by comparative genomic hybridization.

Thirty-five hepatocellular carcinomas (HCCs) associated with hepatitis C virus (HCV) were analysed by comparative genomic hybridization (CGH), to screen for changes in copy-number of DNA sequences. Chromosomal losses were noted in 1p34-36 (37%), 4q12-21 (48%), 5q13-21 (35%), 6q13-16 (23%), 8p21-23 (28%), 13q (20%), 16q (33%) and 17p13 (37%). Gains were noted in 1q (46%), 6p (20%), 8q21-24 (31%) and 17q (43%). High level gains indicative of gene amplifications were found in 7q31 (3%), 11q13 (3%), 14q12 (6%) and 17q12 (3%); amplification at 14q12 may be characteristic for HCCs. No significant difference in chromosomal aberrations was noted between carcinomas associated with HCV-infection in our study and those reported earlier in HCCs infected with hepatitis B virus (HBV), indicating that both HBV- and HCV-related carcinomas may progress through a similar cascade of molecular events.     

Identification of chromosome aberrations in sporadic microsatellite stable and unstable colorectal cancers using array comparative genomic hybridization

Colorectal cancer (CRC) is one of the most common cancers in Denmark and in the western world in general, and the prognosis is generally poor. According to the traditional molecular classification of sporadic colorectal cancer, microsatellite stable (MSS)/chromosome unstable (CIN) colorectal cancers constitute approximately 85% of sporadic cases, whereas microsatellite unstable (MSI) cases constitute the remaining 15%. In this study, we used array comparative genomic hybridization (aCGH) to identify genomic hotspot regions that harbor recurrent copy number changes. The study material comprised fresh samples from 40 MSS tumors and 20 MSI tumors obtained from 60 Danish CRC patients. We identified five small genomic regions (<15 megabases) exhibiting recurrent copy number loss, which, to our knowledge, have not been reported in previously published aCGH studies of CRC: 3p25.3, 3p21.2-p21.31, 5q13.2, 12q24.23-q24.31, and 12q24.23-q24.31. These regions contain several potentially important tumor suppressor genes that may play a role in a significant proportion of both sporadic MSS CRC and MSI CRC. Furthermore, the generated aCGH data are in support of the recently proposed classification of sporadic CRC into MSS CIN+, MSI CIN-, MSI CIN+, and MSS CIN- cancers.     

DNA copy number aberrations associated with the clinicopathological features of colorectal cancers: Identification of genomic biomarkers by array-based comparative genomic hybridization

The aim of the present study was to investigate the chromosomal aberrations that are linked with the crucial clinicopathological features of colorectal cancer (CRC) and its prognosis by array-based comparative genomic hybridization (CGH). Fresh-frozen tumor tissues of 94 cases of CRC were analyzed by using bacterial artificial chromosome (BAC) CGH slides spotted with 4030 human BAC clones, which covered the whole range of the human genome at an average interval of 0.83 mega base pairs. DNA copy number aberrations (DCNAs) were identified in association with clinicopathological features: a gain of 8q24.3 and losses of 9q33.1 and 20p12.2 were associated with lymph node metastasis, gain of 8q24.3 and loss of 9q33.1 with disease stage, gain of 8q21.11 and loss of 10q21.3 with lymphovascular invasion and losses of 3p25.1, 10p15.3, 12q15 and 17p13.1 for venous invasion. These aberrations can be regarded as genomic biomarkers to predict the clinical outcome of patients with CRC, and are expected to serve to individualize the treatment of CRC patients.     

Analysis of cytogenetic aberrations in sporadic vestibular schwannoma by comparative genomic hybridization

Vestibular schwannomas (VS) are benign tumors of the nervous system that are usually sporadic but also occur in the inherited disorder neurofibromatosis type 2 (NF2). The NF2 gene is a tumor suppressor gene located on chromosome 22. Loss of the NF2 protein product, Merlin, is universal in both sporadic and NF2-related schwannomas and the loss or mutation of the gene is the only established causative event underlying schwannoma formation. Comparative genomic hybridization (CGH) was used to screen 20 sporadic VS to identify additional chromosomal regions that may harbor genes involved in VS-tumorigenesis. The most common change were losses on chromosome 22q. Additionally, losses were observed on chromosome 9p indicating a possible participation of the CDKN2A tumor suppressor gene in the genesis of VS. Gains were observed on 17q, 19p and 19q, which have been reported before in malignant peripheral nerve sheath tumors that are associated with neurofibromatosis type 1. Importantly, high level amplifications have been observed on 16p and 16q as well as on 9q, suggesting the possible involvement of several oncogenes in the tumorigenesis of VS. Our data suggest the involvement of various oncogenes and tumor suppressor genes might play a role in the genesis of the vestibular schwannomas apart from the inactivation of the NF2 gene.     

Chromosomal aberrations in head and neck squamous cell carcinomas in Norwegian and Sudanese populations by array comparative genomic hybridization

We used microarray-based comparative genomic hybridization to explore genome-wide profiles of chromosomal aberrations in 26 samples of head and neck cancers compared to their pair-wise normal controls. The samples were obtained from Sudanese (n=11) and Norwegian (n=15) patients. The findings were correlated with clinicopathological variables. We identified the amplification of 41 common chromosomal regions (harboring 149 candidate genes) and the deletion of 22 (28 candidate genes). Predominant chromosomal alterations that were observed included high-level amplification at 1q21 (harboring the S100A gene family) and 11q22 (including several MMP family members). Regions of copy number increase was also identified at 6p21 (p21), 7p12 (EGFR), 17p13 (p53) and 19p13.2 (p19INK4d), while regions showing deletion included among others 3p25.2 (RAF1) and 9p21 (p15, p16). We found genes from four common biological pathways (MAPK signaling, cytokine-cytokine receptor interaction, ECM-receptor interaction and Jak-STAT signaling) to be predominantly over-represented in areas of gain and loss. The current study provides valuable information on chromosomal aberrations likely to be involved in the pathogenesis of head and neck cancers. An increased copy number of the S100A and MMP gene family members, known to be involved in invasion and metastasis, may play an important role in the development of the tumors. Hierarchical clustering of the chromosomal alterations with clinicopathological parameters showed little correlation, suggesting an occurrence of gains/losses regardless of ethnic differences and clinicopathological status between the patients from the two countries. Our findings indicate the existence of common gene-specific amplifications/deletions in these tumors, regardless of the source of the samples or attributed carcinogenic risk factors.     

采用比较基因组杂交方法分析原发性食管癌染色体异常

背景与目的:有研究表明原发性食管癌常有染色体的改变,包括染色体基因组异常扩增和缺失.比较基因组杂交技术可以显示这些染色体的异常变化.本实验采用比较基因组杂交技术研究和分析原发性食管癌染色体基因组的变化特点及其与预后的关系.方法:采用比较基因组杂交技术检测16例食管癌组织中染色体的异常改变,并分析染色体异常与预后的关系.研究的病例中7例食管癌术后2年内死亡(对照组),9例术后生存3年以上(生存组).结果:食管癌患者中多数染色体基因组发生改变,最常见的染色体基因组高扩增频率发生在1q/p,2q/p,3q,5q/p,8q/p,9q/p,11q/p,17和20q/p染色体区段上,在染色体1q/p,4p,9p,18q和xp中常见染色体基因缺失.染色体7q/p和19扩增频率和染色体4q/p和18q缺失频率,生存组与对照组间存在显著性差异.结论:食管癌患者染色体区段基因易发生异常扩增和缺失,生存组与对照组存在明显差异.