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Results for “amplification”

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PubMed2002

[HER-2 and TOP 2A gene amplifications in urinary bladder carcinoma].

The HER-2/neu gene is frequently amplified in bladder cancer. Topoisomerase 2 Alpha (TOP2A) which is located nearby the HER-2/neu gene is an important molecular target for several anti cancer drugs. The frequency of TOP2A amplification in urinary bladder cancer is unknown. It was the aim of this study to determine the frequency of HER-2 and TOP2A amplification in urinary bladder cancer and to evaluate the association of these amplifications with tumor phenotype. For this purpose a tissue microarray containing 768 pTa, 425 pT1 and 571 pT2-4 carcinomas was analyzed by fluorescence in situ hybridization (FISH). Amplifications of both genes were significantly associated with advanced tumor stage and high grade. HER-2 amplification was found in 1.6% of pTa, 7.2% of pT1 and 13.8% of pT2-4 carcinomas (p < 0.0001). HER-2 amplification was present in only 1.1% of grade 1 and 0.8% of grade 2 tumors but in 14.2% of grade 3 tumors (p < 0.0001). TOP2A amplification was present in 0.7% pTa, 1.8% pT1 and 3.4% pT2-4 carcinomas (p < 0.0001). TOP2A was found in none of the grade 1, in 0.2% of grade 2 and 3.8% of grade 3 tumors (p < 0.0001). 1% of all analyzed tumors had simultaneously high level amplification of TOP2A and HER-2. Amplification of both genes were significantly associated with tumor specific survival if all tumors were analyzed together. Given the high frequency of HER-2 amplification in urinary bladder cancer, some of these tumors may respond favorable to Herceptin therapy. The TOP2A amplification status may influence response to anthracyclin treatment.

Antigens, Neoplasm↗

PubMed1995

In situ amplification of measles virus RNA by the self-sustained sequence replication reaction.

BACKGROUND: The self-sustained sequence replication (3SR) reaction is an isothermal method for nucleic acid amplification that has several features that make it an attractive alternative to PCR. We have studied the feasibility of the in situ 3SR reaction in cells using a measles virus-infected cell line as a model. EXPERIMENTAL DESIGN: The study was carried out in four steps. First, using RNA extracted from a measles-infected Vero Green monkey kidney cell line, conditions for the in vitro amplification of a segment of the nucleocapsid portion of the RNA viral genome were optimized for 420- and 119-bp 3SR products, and the results were compared. Second, 3SR was performed on intact infected cells in suspension, and the amount of RNA product was compared with infected cells without 3SR. Then, the 3SR reaction was conducted on cytospin preparation slides, followed by in situ hybridization for detection of the amplification product. Finally, 3SR was carried out on sections of formalin-fixed, paraffin-fixed, paraffin-embedded cells, and the degree of amplification as detected by ISH was quantified and compared between infected cells with and without 3SR reaction. RESULTS: Specific amplification of measles was observed in each of these types of preparations with an 8.5-fold rate of amplification in paraffin sections of formalin-fixed cells (a mean of 272.5 +/- 65.3 grains/cell after 3SR amplification in comparison to 31.97 +/- 4.2 grains/cell without amplification). CONCLUSIONS: A significant amount of amplification of RNA is possible with in situ 3SR (IS-3SR) and, in combination with ISH, offers several advantages compared with in situ PCR (IS-PCR), such as ease of use, lack of conditions that lead to cell damage, and a specificity for RNA amplification. This is the first report of specific amplification of RNA within cells using the IS-3SR procedure, a technique that has a wide range of potential applications in pathology and molecular biology.

Animals↗

PubMed1997

Androgen receptor gene amplification: a possible molecular mechanism for androgen deprivation therapy failure in prostate cancer.

Progression of prostate cancer during endocrine therapy is a major clinical problem, the molecular mechanisms of which remain poorly understood. Amplification of the androgen receptor (AR) gene was recently described in recurrent prostate carcinomas from patients who had failed androgen deprivation therapy. To evaluate the hypothesis that amplification of the AR gene is a cause for the failure of androgen deprivation therapy in prostate cancer, we studied whether AR amplification leads to gene overexpression, whether the amplified AR gene is structurally intact, and whether tumors with AR amplification have distinct biological and clinical characteristics. Tumor specimens were collected from 54 prostate cancer patients at the time of a local recurrence following therapy failure. In 26 cases, paired primary tumor specimens from the same patients prior to therapy were also available. Fifteen (28%) of the recurrent therapy-resistant tumors, but none of the untreated primary tumors, contained AR gene amplification as determined by fluorescence in situ hybridization. According to single-stranded conformation polymorphism analysis, the AR gene was wild type in all but one of the 13 AR amplified cases studied. In one tumor, a presumed mutation in the hormone-binding domain at codon 674 leading to a Gly --> Ala substitution was found, but functional studies indicated that this mutation did not change the transactivational properties of the receptor. AR amplification was associated with a substantially increased level of mRNA expression of the gene by in situ hybridization. Clinicopathological correlations indicated that AR amplification was most likely to occur in tumors that had initially responded well to endocrine therapy and whose response duration was more than 12 months. Tumors that recurred earlier or those that showed no initial therapy response did not contain AR amplification. The median survival time after recurrence was two times longer for patients with AR amplification in comparison to those with no amplification (P = 0.03, Willcoxon-Breslow test). In conclusion, failure of conventional androgen deprivation therapy in prostate cancer may be caused by a clonal expansion of tumor cells that are able to continue androgen-dependent growth despite of the low concentrations of serum androgens. Amplification and the increased expression of a wild-type AR gene may play a key role in this process.

Aged↗

PubMed1996

Cyclin D1 amplification as a new predictive classification for squamous cell carcinoma of the esophagus, adding gene information.

The cyclin D1, referred to as PRAD-1, has been mapped to the 11q13 region, and its expression has been detected in squamous cell lines and several primary esophageal carcinomas. We assessed cyclin D1 amplification in 122 squamous cell carcinomas of the esophagus. Samples for DNA extraction were obtained from formalin-fixed paraffin-embedded specimens, and 10 microgram of each DNA sample were subjected to slot blot analysis. The presence of more than three gene copies was considered evidence of gene amplification. Amplification of cyclin D1 was detected in 28 (23%) of 122 cases of squamous cell carcinoma of the esophagus. There were no significant differences between the clinicopathological background factors in groups positive and negative for cyclin D1 amplification, but the survival rate of patients exhibiting amplification was significantly lower (P < 0.001). The groups were stratified according to the pN (pathological N category) factor and pT (pathological T category) factor in the TNM classification, and the cumulative survival rates in the amplification groups were always significantly lower. Amplification of cyclin D1 was correlated with distant organ metastasis after curative operations, but there was no significant difference in lymph node recurrence rates of patients with or without amplification. Cyclin D1 amplification had the second highest partial regression coefficient in the multivariate analysis, after the pN factor. Amplification of cyclin D1 was independent of the TNM classification as a prognostic factor, and was a useful marker for predicting outcome and distant organ metastasis in patients with squamous cell carcinoma of the esophagus. It appears that appropriate treatment can be selected by evaluating both TNM factors and cyclin D1 amplification.

Adult↗

PubMed1995

Strategies to respond to polymerase chain reaction deoxyribonucleic acid amplification failure in a preimplantation genetic diagnosis program.

OBJECTIVES: Our purpose was to identify and evaluate practical methods within a preimplantation genetic diagnosis program that will increase the percentage of embryos for which a genetic diagnosis can be obtained, including clinical responses after failure of deoxyribonucleic acid amplification has occurred. STUDY DESIGN: Known human lymphoblast cell lines and human embryo blastomeres were evaluated in a single-cell, nested primer polymerase chain reaction system with primer sequences for the specific locus surrounding the four base pair insertion mutation on exon 11 of beta-hexosaminidase A-Tay-Sachs disease, the delta F508 mutation of cystic fibrosis, and the sex-determining region on the Y chromosome. Reamplification polymerase chain reaction with standard polymerase chain reaction and primer extension preamplification was performed in deoxyribonucleic acid preparations after previous polymerase chain reaction amplification attempts had resulted in failure of amplification. RESULTS: The amplification efficiency of Tay-Sachs disease, 51% (97/187), was significantly lower than that for cystic fibrosis, 85% (87/107), and for the sex-determining region on the Y chromosome, 85% (77/90). Tay-Sachs disease polymerase chain reaction amplification occurred in 51% of one-cell lymphoblasts, 89% of two-cell lymphoblasts, and 94% of samples when more than two cells were processed together. When previous amplification failure had occurred, standard Tay-Sachs disease polymerase chain reaction resulted in an amplification efficiency of 16% (three of 19), whereas primer extension preamplification polymerase chain reaction for Tay-Sachs disease resulted in amplification of 52% (31/59) lymphoblasts and 54% (13/24) of polyspermic human blastomeres. Four of six human blastomeres in which amplification failure occurred in a Tay-Sachs disease preimplantation genetic diagnosis cycle amplified by primer extension preamplification polymerase chain reaction, which increased the diagnostic information obtained from four to six of the seven embryos on which biopsy was performed. CONCLUSIONS: We suggest that practical approaches for consideration within a clinical preimplantation genetic diagnosis program to limit the net effect of amplification failure (i.e., reduced embryo transfer number) include increasing the deoxyribonucleic acid content in the polymerase chain reaction tube by using more than one blastomere and by using primer extension preamplification when the initial attempt at amplification fails.

Base Sequence↗

PubMed1996

Epidermal growth factor receptor gene and c-erbB-2 gene amplification in ovarian cyst fluid.

OBJECTIVE: To test for a significant difference between functional and neoplastic ovarian cyst with respect to epidermal growth factor (EGF) receptor and c-erbB-2 proto-oncogene amplification rates. METHODS: We determined amplification of EGF-receptor and c-erbB-2 genes by differential polymerase chain reaction (PCR) on 138 ovarian-cyst aspirates. The semiquantitative differential PCR is based on simultaneous co-amplification of a target gene and a reference gene. Amplification rates were detected by densitometry of silver-stained polyacrylamide gels and were scored as single-, low-, or high-copy numbers. Wilcoxon ranked sum test, Pearson correlation coefficient, and multiple logistic regression were used to evaluate the differences in oncogene amplification and to predict histology. RESULTS: There were 71 (51.5%) women with functional cysts, whereas 67 (48.5%) had benign (n = 59) or malignant (n = 8) tumors. Low-copy (two- to fourfold copy numbers) EGF-receptor gene amplification was found in 22 of 67 (33%) women with neoplastic cysts, but in only eight (11%) of those with functional cysts. Neoplastic histology differed significantly from functional histology in correlation to EGF-receptor low-copy gene amplification (r = .279, P < .001). There was no significant difference in c-erbB-2 gene amplification with respect to functional and neoplastic histology (r = .083, P = .32). CONCLUSIONS: Low-copy EGF-receptor gene amplification seems to be a market for neoplastic histology. Epidermal growth factor receptor gene amplification may be involved in proliferation and growth in an early stage of tumorigenesis. However, further studies are required to investigate this gene structure abnormality on a gene function level. Amplification of c-erbB-2 proto-oncogene does not appear to be a common factor in the development of ovarian tumors.

Adolescent↗

PubMed1995

Allelic drop-out and preferential amplification in single cells and human blastomeres: implications for preimplantation diagnosis of sex and cystic fibrosis.

Previously the diagnosis of sex and cystic fibrosis status has been studied on single cells using the polymerase chain reaction (PCR). It has been suggested that allelic drop-out (PCR failure of one allele) and/or preferential amplification (hypo-amplification of one allele) may contribute to poor reliability and misdiagnosis, although this remains controversial as some reports suggest that allelic drop-out does not occur. We investigated an improved method of diagnosing sex and cystic fibrosis in single cells using a new technology (fluorescent PCR) to determine the base level of PCR artefacts (allelic drop-out and preferential amplification) which, in combination with improved sensitivity, should improve PCR reliability and accuracy. Fluorescent PCR gives high reliability (approximately 97%) and accuracy rates (approximately 97%) in somatic cells for both sex and cystic fibrosis diagnosis and its lower detection threshold allows allelic drop-out and preferential amplification to be easily distinguished. We also achieved high reliability and accuracy in diagnosing cystic fibrosis in human blastomeres. This study confirms earlier reports of both allelic drop-out and preferential amplification in single cell analysis. We demonstrate that both allelic drop-out and preferential amplification occur in somatic cells and suggest these are separate phenomena. Preferential amplification appeared common in single cell PCR while allelic drop-out apparently occurred at random in each allele. Preferential amplification was mainly amplification of the larger allele. We suggest that some inaccuracy/misdiagnosis may be due to both preferential amplification as well as allelic drop-out. Other findings were variability in drop-out between PCR and that amplification of signals from human blastomeres may be linked to embryo quality. We suggest that allelic drop-out is dependent on the number of cells within the sample.

Alleles↗

PubMed1992

Preferential PCR amplification of alleles: mechanisms and solutions.

The preferential PCR amplification of one allele relative to another in a heterozygous sample could result in an incorrect or ambiguous genetic typing of that sample. There are several mechanisms that could potentially lead to such preferential PCR amplification. First, preferential amplification can result from significant GC% differences between alleles if the conditions of the reaction (denaturation temperature (Tden), duration at the Tden' salt and co-solvent concentrations, etc.) allow the denaturation of one allele but not the other (differential denaturation). For example, the DQa1.1, -1.2, and -1.3 alleles of the HLA-DQa locus do not amplify at a Tden < 89 degrees C; these same conditions still allow amplification of the DQa2, -3, and -4 alleles. However, no differences in amplification efficiency were found between the different HLA-DQa alleles when the Tden was set at the recommended Tden of 94 degrees C, even after as many as 102 cycles of amplification. Second, for PCR-based genetic typing systems in which the PCR products from different alleles differ in length, preferential amplification of the shorter allelic product can occur. Experiments in which the variable number tandem repeat (VNTR) marker D17S5 (YNZ22) was amplified under various conditions suggest that the smaller allelic products are amplified preferentially when Taq polymerase is limiting. Preferential amplification of VNTR alleles can also occur if the target DNA is sufficiently degraded. Third, when the initial number of genomes sampled is very small, stochastic fluctuation in the number of copies of each allele can result in what appears to be preferential amplification. Finally, less efficient priming of DNA synthesis of one allele versus another can occur because of mismatches between the primer and the specific allelic template, resulting in preferential amplification of the other allele. General strategies to avoid preferential amplification are discussed.

Alleles↗

PubMed1995

Prevalence and significance of HER-2/neu amplification in epithelial ovarian cancer.

Amplification of the HER-2/neu oncogene was assessed in 80 cases of epithelial ovarian tumors using differential polymerase chain reaction. HER-2/neu gene was amplified in 22 of 46 invasive cancers (48%) and in 5 of 34 borderline cancers (15%), but none of the 20 specimens of normal ovaries showed amplification. THis difference is statistically significant (p = 0.00004). The incidence of HER-2/neu amplification in late stage (III-IV, 77%) was significantly higher than that in early stage (I-II, 21%) in invasive epithelial carcinoma (p = 0.0004). There was no correlation between HER-2/neu amplification and cell type or grade of tumor. In cases of ovarian tumors of borderline malignant potential, the amplification of HER-2/neu was not correlated with clinicopathologic features. Follow-up with a mean of 22 months (6-50 months) was available for 39 cases of invasive ovarian cancers and all 34 borderline ovarian cancers. The incidence of HER-2/neu amplification in the invasive cancer and borderline cancer patients who were alive with disease was 50 and 50%, and is not statistically different from that in the patients who were alive with no evidence of disease (p = 0.662 and 0.345, respectively). The incidence of amplification in the invasive cancers of patients who died of the disease (86%) was higher than that in the patients who were still alive (44%), but the difference is not statistically significant (p = 0.175). This study supports the association of HER-2/neu amplification with progression of invasive ovarian cancer. It also suggests that HER-2/neu amplification may be an adjunctive prognostic factor of invasive epithelial ovarian cancer, shown to be associated with an unfavorable clinical course. In addition, HER-2/neu amplification occurs relatively infrequently in early invasive and borderline ovarian cancers, making it unlikely that such amplification is a general early event in ovarian carcinogenesis.

Base Sequence↗

PubMed2006

c-myc amplification is frequent in esophageal adenocarcinoma and correlated with the upregulation of VEGF-A expression.

BACKGROUND: Deregulation of c-myc plays a major role in the carcinogenesis of human malignancies. We investigated the amplification of the c-myc gene in a surgical series of Barrett cancers. METHODS: Primary resected esophageal (Barrett) adenocarcinomas (n = 84) were investigated for c-myc amplification using chromogene in situ hybridization. Tumor samples were assembled in a tissue microarray. c-myc gene dosage was correlated with clinicopathologic parameters, including the survival and gene expression of cyclooxygenases (COX-1 and COX-2) and proangiogenic growth factors (VEGF-A and VEGF-C). RESULTS: The majority (70 of 84; 83.3%) exhibited amplification of the c-myc gene. There were low-level amplifications in 63 (75.0%) cases and high-level amplifications in 7 (8.3%) cases. No amplification was found in 14 (16.7%) cases. Tumors without c-myc amplification had lower VEGF-A, VEGF-C, and COX-2 expression levels than tumors with low-level and high-level c-myc amplification (statistically significant for VEGF-A; P = .0348). c-myc amplification was not correlated with clinicopathological parameters or survival. Only diffuse and mixed-type tumors, according to Lauren classification, exhibited c-myc amplifications more frequently (P = .0466). CONCLUSIONS: Amplifications of the c-myc gene are frequent in Barrett cancer. c-myc may be involved in the regulation of angiogenesis.

Adenocarcinoma↗

PubMed1999

Concomitant gastrin and ERBB2 gene amplifications at 17q12-q21 in the intestinal type of gastric cancer.

Our recent studies using comparative genomic hybridization showed that gain or amplification at the 17q12-q21 region is very common in the intestinal type of gastric cancer. Here, we describe a fluorescence in situ hybridization study with gastrin (GAS)-specific and ERBB2-specific probes on ten specimens of gastric carcinoma that, by using comparative genomic hybridization, showed 1) DNA copy number gain or amplification at 17q12-q21, a region known to harbor the GAS and ERBB2 genes (four cases); 2) gain of the entire chromosome 17 (three cases); or 3) normal copy number of chromosome 17 (three cases). GAS and ERBB2 protein expression was studied by Western immunoblotting from gastric cancer cell lines with or without gain at 17q12-q21 as well as a breast cancer cell line with ERBB2 amplification. Our results showed that simultaneous amplification of both GAS and ERBB2 was four- to ninefold in the tumors with the 17q12-q21 amplification. Both genes were amplified in the same nuclei, and the hybridization signals were localized to the same region of the nucleus. Overexpression of GAS and ERBB2 was observed by Western immunoblotting only in the gastric cancer cell line with gain at 17q12-q21. The ERBB2 amplification is also a recurrent change in breast cancer. To investigate whether the GAS amplification is unique in gastric cancer, fluorescence in situ hybridization analysis was performed on 40 breast cancer cell lines. The ERBB2 amplification was observed in 11 cell lines, but none of the lines showed the GAS amplification. This indicates that the formation of an amplicon, in which both the GAS and the ERBB2 genes are amplified, might be unique in gastric cancer, especially in its intestinal type, and that simultaneous amplification of both genes is important to the tumorigenesis of intestinal gastric cancer. We demonstrate here for the first time that a gene of a physiological hormone is amplified in tumors that originate from cells that normally secrete the hormone.

Breast Neoplasms↗

PubMed2001

c-erbB-2 (HER-2/neu) gene amplification is a better indicator of poor prognosis than protein over-expression in operable breast-cancer patients.

Our aim was to compare the prognostic value of c-erbB-2 gene amplification analyzed by Southern blot with that of protein (p185) over-expression measured by immunohistochemistry in 172 patients with operable breast cancer (BC). Amplification and p185 over-expression were found in 31 (18%) and 51 (30%) BCs, respectively. All but 1 of the tumors showed both amplification and over-expression, while 21 (12%) tumors displayed over-expression without amplification. The risk of death associated with c-erbB-2 gene amplification and p185 over-expression was evaluated by multivariate analysis, taking into account tumor size, histoprognostic grade, hormone receptors and axillary node status. During a mean follow-up of 9.5 (+/-2) years, node involvement (p < 0.001), c-erbB-2 gene amplification (p = 0.02) and negative hormone receptors (p = 0.02) were found to be independent prognostic indicators of the risk of death. Over-expression of p185 with no amplification was not correlated with this risk. When the risk of death associated with c-erbB-2 amplification was studied according to chemo- and hormone therapy, no significant difference was observed between subgroups of subjects. Amplification was also associated (p = 0.02) with the risk of multifocal distant metastases (i.e., metastases detected concomitantly in at least 2 sites) and, thus, with BC aggressiveness. These data show the importance of c-erbB-2 gene amplification in predicting the long-term outcome of patients and in selecting eligible patients for c-erbB-2-targeted therapies.

Biomarkers, Tumor↗

PubMed2002

Prognostic significance of c-myc and AIB1 amplification in hepatocellular carcinoma. A broad survey using high-throughput tissue microarray.

BACKGROUND: Amplifications of 1q21, c-myc at 8q24.1, and AIB1 at 20q12 are genetic alterations that are detected frequently in hepatocellular carcinoma (HCC). The authors evaluated the association of these amplifications with the prognosis of patients with HCC. METHODS: In the current study, amplification of 1q21, c-myc, and AIB1 was analyzed in 560 specimens from 400 patients with HCC and 20 patients with benign liver lesions using fluorescence in situ hybridization with high-throughput tissue microarray. Differences of amplification patterns were compared between small and large HCC, single nodular and multiple nodular HCC, primary and metastatic HCC, and primary and recurrent HCC. RESULTS: Significant differences between single nodular and multiple nodular HCC were detected in c-myc amplification (12% vs. 38%; P < 0.01) and AIB1 amplification (16% vs. 30%; P < 0.05). More frequent c-myc amplification was detected in metastatic HCC (45%) compared with primary HCC (29%) and in recurrent HCC (60%) compared with primary HCC (38%). Similarly, more frequent AIB1 amplification was observed in metastatic HCC (41%) compared with primary HCC (23%) and in recurrent HCC (60%) compared with primary HCC (29%). However, no significant differences in 1q21 amplification were observed. CONCLUSIONS: The current results strongly suggest that amplifications of the c-myc and AIB1 oncogenes are late genetic alterations in the progression of HCC and are correlated with a poor prognosis.

Carcinoma, Hepatocellular↗

PubMed2007

Nonrandom distribution of oncogene amplifications in bilateral breast carcinomas: Possible role of host factors and survival bias.

Amplification of HER2, C-MYC and CCND1 oncogenes is a hallmark of breast cancer (BC); however, its involvement in the bilateral form of this disease has not been investigated yet. In this study, 50 bilateral BC (biBC) pairs (100 tumors) and 72 control unilateral BC were examined using real-time PCR analysis of microdissected archival tissues. In biBC, the frequency of >3-fold oncogene amplification was 6/100 (6%) for HER2, 6/100 (6%) for C-MYC and 7/100 (7%) for CCND1. Altogether, 18/100 (18%) biBC tumors had increased gene dosage of at least one oncogene. Tumors forming synchronous biBC pairs had amplification in 11/46 cases (24%). In 3 of 8 patients with amplification-positive carcinomas, the amplification was detected in both neoplasms: 2 biBC had concordant activation of the same oncogene (HER2 and CCND1, respectively), and in the remaining case distinct oncogenes were affected (HER2 and C-MYC). In contrast, amplifications in metachronous biBC were strongly discordant: none of 27 first carcinomas carried this abnormality, while the frequency of amplification in second tumors (7/27; 26%) was similar to the one observed in unilateral BC (20/72; 28%). The trend toward concordance of oncogene amplification status in synchronous but not in metachronous biBC pairs can be explained by the nearly identical natural history of the disease in simultaneously arising tumors. The skewed pattern of amplifications in metachronous biBC might be attributed to their association with adverse BC prognosis; it appears that only patients with amplification-negative first BC have sufficient chances to survive until the development of the contralateral carcinoma.

Adult↗

PubMed1989

The significance of oncogene amplification in primary breast cancer.

Alterations in the gene copy numbers of the proto-oncogenes HER2/neu and c-myc in primary human breast cancer investigated in 73 patients. We detected amplification of HER2/neu in 17 patient samples and amplification of c-myc in 11, while amplification of both was seen in 6 samples. There was no correlation of age, hormone receptor positivity or tumour size with amplification of either proto-oncogene. Amplification of HER2/neu was significantly correlated with the stage of the disease. HER2/neu amplification was observed in 18.5% and 38% of node-negative and node-positive patients, respectively; the association between HER2/neu amplification and advanced stage of the disease was statistically significant (p = 0.05). Since this is a prospective study, the clinical significance of oncogene amplification is not known. The relatively high frequency of HER2/neu amplification points to a functional role in human breast cancer, particularly in the progression of the disease. The method used in our study allows oncogene amplification to be studied in conjunction with hormone receptor determination and thus may be of value in large clinical trials to determine the significance of oncogene abnormalities in breast cancer.

Breast Neoplasms↗

PubMed1994

Analysis of gene amplification and overexpression in human esophageal-carcinoma cell lines.

Gene amplification/overexpression was analyzed in 23 cell lines derived from human esophageal squamous-cell-carcinoma tissues by Southern and Northern hybridizations to c-myc, c-erbB, hst-1 and cyclin-D1 probes. Amplification of the c-myc gene was observed in 5 cell lines derived from well-differentiated carcinomas and all of them were accompanied by co-amplification of other examined oncogenes. The c-erbB gene was amplified in 3 cell lines. Co-amplification of hst-1 and cyclin D1, both of which are located in chromosome 11q13, was found in 9 cell lines. Without exception their amplification was simultaneous and the magnitudes were similar. Their amplification, but not their overexpression, was significantly correlated with poor prognosis in patients from whom the cell lines were established. While hst-1-gene expression was not detected, at least 1 of the genes analyzed was overexpressed in 20 cell lines vs. its expression in normal esophageal mucosal tissues. However, gene amplification was not necessarily accompanied by overexpression of the corresponding genes. Expression of the cyclin D1 gene, which has been assumed to be a target gene for 11q13 amplification, was not detected in one particular cell line with amplification of 11q13. These results suggest that the amplification/overexpression of more than I oncogene is involved in the carcinogenic process of esophageal carcinoma and that c-myc-gene amplification is associated with a well-differentiated subtype. There remains a possibility that key oncogenes other than cyclin D1 are involved in 11q13 amplification.

Animals↗

PubMed2003

High-throughput tissue microarray analysis of 11q13 gene amplification (CCND1, FGF3, FGF4, EMS1) in urinary bladder cancer.

Gene amplification is a common mechanism for oncogene overexpression. High-level amplifications at 11q13 have been repeatedly found in bladder cancer by comparative genomic hybridization (CGH) and other techniques. Putative candidate oncogenes located in this region are CCND1 (PRAD1, bcl-1), EMS1, FGF3 (Int-2), and FGF4 (hst1, hstf1). To evaluate the involvement of these genes in bladder cancer, a tissue microarray (TMA) containing 2317 samples was screened by fluorescence in situ hybridization (FISH). The frequency of gains and amplifications of all genes increased significantly from stage pTa to pT1-4 and from low to high grade. In addition, amplification was associated with patient survival and progression of pT1 tumours. Among 123 tumours with amplifications, 68.3% showed amplification of all four genes; 19.5% amplification of CCND1, FGF4, and FGF3; and 0.8% co-amplification of FGF4, FGF3, and EMS1. Amplification of CCND1 alone was found in 9% of the tumours, while EMS1 alone was amplified in 1.6% and FGF4 in 0.8%. Overall, the amplification frequency decreased with increasing genomic distance from CCND1, suggesting that, among the genes examined, CCND1 is the major target gene in the 11q13 amplicon in bladder cancer.

Adenocarcinoma↗

PubMed1999

Amplification and overexpression of the cyclin D1 and epidermal growth factor receptor genes in non-small-cell lung cancer. Lung Cancer Study Group.

PURPOSE: To study the structure and expression of the cyclin D1 and the epidermal growth factor receptor (EGFR) genes in a cohort of 298 non-small-cell lung cancer (NSCLC) specimens. METHODS: Gene structure was studied by Southern analysis, and gene expression was studied by Northern analysis and immunohistochemical analysis. RESULTS: Amplification of the cyclin D1 locus was found in 14/298 (5%) specimens. All 12/12 specimens with amplification of the cyclin D1 gene for which RNA was available were found to express the cyclin D1 transcript, and 11/12 overexpressed the transcript to levels higher than that of uninvolved lung. The EGFR gene was amplified in 17/286 samples of NSCLC tested, and was overexpressed in 22/169 (13%) cases tested, including 12/13 cases with amplification of the gene for which RNA was available. Cyclin D1 gene amplification was associated with advanced lymph node involvement (P = 0.043), but not with larger tumor size or adverse outcome. Cyclin D1 gene amplification and overexpression occurred independently of retinoblastoma tumor-suppressor gene (RB) inactivation, but tumors with amplification of the cyclin D1 gene were more likely to have EGFR gene amplification (P < 0.005). No correlation of EGFR gene amplification or overexpression with tumor size, lymph node involvement, patient demographic data, or survival was identified. CONCLUSIONS: These data indicate that the cyclin D1 and EGFR genes are amplified and overexpressed in NSCLC, and amplification of the cyclin D1 gene occurs frequently in conjunction with amplification of the EGFR gene.

Adenocarcinoma↗