Showing papers by "Ie Ming Shih published in 2006"
TL;DR: Notch3 is required for proliferation and survival of Notch3-amplified tumors and inactivation of notch3 can be a potential therapeutic approach for ovarian carcinomas, according to results.
Abstract: Gene amplification is one of the common mechanisms that activate oncogenes. In this study, we used single nucleotide polymorphism array to analyze genome-wide DNA copy number alterations in 31 high-grade ovarian serous carcinomas, the most lethal gynecologic neoplastic disease in women. We identified an amplicon at 19p13.12 in 6 of 31 (19.5%) ovarian high-grade serous carcinomas. This amplification was validated by digital karyotyping, quantitative real-time PCR, and dual-color fluorescence in situ hybridization (FISH) analysis. Comprehensive mRNA expression analysis of all 34 genes within the minimal amplicon identified Notch3 as the gene that showed most significant overexpression in amplified tumors compared with nonamplified tumors. Furthermore, Notch3 DNA copy number is positively correlated with Notch3 protein expression based on parallel immunohistochemistry and FISH studies in 111 high-grade tumors. Inactivation of Notch3 by both gamma-secretase inhibitor and Notch3-specific small interfering RNA suppressed cell proliferation and induced apoptosis in the cell lines that overexpressed Notch3 but not in those with minimal amount of Notch3 expression. These results indicate that Notch3 is required for proliferation and survival of Notch3-amplified tumors and inactivation of Notch3 can be a potential therapeutic approach for ovarian carcinomas.
283 citations
TL;DR: Analysis of sequence mutations in PIK3CA and AKT2 genes using purified tumor cells that were isolated from high-grade ovarian serous carcinomas and serous borderline tumors and a dual-color FISH on tissue microarrays provides unambiguous evidence that somatic sequence mutations of PIK2CA and ATK2 are rare in ovarianSerous tumors but amplification of both genes may play an important role in the development of high- grade ovarian serious carcinoma.
Abstract: Sequence mutations and gene amplifications lead to activation of the PIK3CA-AKT2 signaling pathway and have been reported in several types of neoplasms including ovarian cancer. Analysis of such genetic alterations, however, is usually complicated by contamination of normal cell DNA, artifacts associated with formalin-fixed tissues and the sensitivity of the techniques employed. In this study, we analyzed the sequence mutations in PIK3CA and AKT2 genes using purified tumor cells that were isolated from high-grade ovarian serous carcinomas and serous borderline tumors (SBTs) and assessed gene amplification using a dual-color FISH on tissue microarrays. Somatic sequence mutations in the kinase domain of AKT2 were not detected in any of the 65 ovarian tumors analyzed. Mutations of PIK3CA were rare, occurring only in one (2.3%) of 44 high-grade serous carcinomas and in only one (4.8%) of 21 SBTs. Dual-color FISH demonstrated that PIK3CA and AKT2 were not amplified in SBTs but amplified in 13.3% and 18.2% high...
186 citations
TL;DR: It is found that ovarian carcinoma cell lines exhibit greater sensitivity to apoptosis in response to proteasome inhibitors than immortalized ovarian surface epithelial cells, which correlates with increased cellular proliferation rate and UPS stress rather than absolute proteasomes levels.
Abstract: The ubiquitin-proteasome system (UPS) mediates targeted protein degradation. Notably, the UPS determines levels of key checkpoint proteins controlling apoptosis and proliferation by controlling protein half-life. Herein, we show that ovarian carcinoma manifests an overstressed UPS by comparison with normal tissues by accumulation of ubiquitinated proteins despite elevated proteasome levels. Elevated levels of total ubiquitinated proteins and 19S and 20S proteasome subunits are evident in both low-grade and high-grade ovarian carcinoma tissues relative to benign ovarian tumors and in ovarian carcinoma cell lines relative to immortalized surface epithelium. We find that ovarian carcinoma cell lines exhibit greater sensitivity to apoptosis in response to proteasome inhibitors than immortalized ovarian surface epithelial cells. This sensitivity correlates with increased cellular proliferation rate and UPS stress rather than absolute proteasome levels. Proteasomal inhibition in vitro induces cell cycle arrest and the accumulation of p21 and p27 and triggers apoptosis via activation of caspase-3. Furthermore, treatment with the licensed proteasome inhibitor PS-341 slows the growth of ES-2 ovarian carcinoma xenograft in immunodeficient mice. In sum, elevated proliferation and metabolic rate resulting from malignant transformation of the epithelium stresses the UPS and renders ovarian carcinoma more sensitive to apoptosis in response to proteasomal inhibition.
122 citations
TL;DR: NAC-1 is a tumor recurrence-associated gene with oncogenic potential, and the interaction between BTB/POZ domains of Nac-1 proteins is critical to form the discrete NAC- 1 nuclear bodies and essential for tumor cell proliferation and survival.
Abstract: Recent studies have suggested an oncogenic role of the BTB/POZ-domain genes in hematopoietic malignancy. The aim of this study is to identify and characterize BTB/POZ-domain genes in the development of human epithelial cancers, i.e., carcinomas. In this study, we focused on ovarian carcinoma and analyzed gene expression levels using the serial analysis of gene expression (SAGE) data in all 130 deduced BTB/POZ genes. Our analysis reveals that NAC-1 is significantly overexpressed in ovarian serous carcinomas and several other types of carcinomas. Immunohistochemistry studies in ovarian serous carcinomas demonstrate that NAC-1 is localized in discrete nuclear bodies (tentatively named NAC-1 bodies), and the levels of NAC-1 expression correlate with tumor recurrence. Furthermore, intense NAC-1 immunoreactivity in primary tumors predicts early recurrence in ovarian cancer. Both coimmunoprecipitation and double immunofluorescence staining demonstrate that NAC-1 molecules homooligomerize through the BTB/POZ domain. Induced expression of the NAC-1 mutant containing only the BTB/POZ domain disrupts NAC-1 bodies, prevents tumor formation, and promotes tumor cell apoptosis in established tumors in a mouse xenograft model. Overexpression of full-length NAC-1 enhanced tumorigenicity of ovarian surface epithelial cells and NIH 3T3 cells in athymic nu/nu mice. In summary, NAC-1 is a tumor recurrence-associated gene with oncogenic potential, and the interaction between BTB/POZ domains of NAC-1 proteins is critical to form the discrete NAC-1 nuclear bodies and essential for tumor cell proliferation and survival.
117 citations
TL;DR: The molecular signatures unique to OC/PPC and DMPM should provide a molecular basis to study both tumors and new potential markers for facilitating their differential diagnosis.
Abstract: Purpose: Ovarian/primary peritoneal serous carcinoma (OC/PPC) and diffuse peritoneal malignant mesothelioma (DMPM) are highly aggressive tumors that are closely related morphologically and histogenetically. It remains unclear whether both tumors are molecularly distinct neoplasms. The current study compared global gene expression patterns in OC/PPC and DMPM. Experimental Design: Ten OC/PPC and five DMPM effusions were analyzed for gene expression profiles using the Affymetrix U133 Plus 2 arrays and the dCHIP analysis program. Differentially expressed candidate genes were validated using quantitative real-time PCR and immunohistochemistry. Results: Unsupervised hierarchical clustering using all 54,675 genes in the array classified the samples into two groups: DMPM specimens versus OC/PPC specimens. A total of 189 genes that were differentially expressed in these two groups were selected based on statistical significance. Genes overexpressed in DMPM ( n = 68) included calretinin, vitronectin, claudin 15, α 4 laminin, hyaluronan synthase 1, cadherin 11, RAB7, v-maf , and the epidermal growth factor–containing fibulin-like extracellular matrix protein 1 . Genes overexpressed in OC/PPC ( n = 121) included insulin-like growth factor II ( IGF-II ); IGF-II binding protein 3 ; cyclin E1 ; folate receptors 1 and 3 ; RAB25 ; MUC4 ; endothelin-1 ; CD24 ; kallikreins 6, 7 , and 8 ; claudins 3, 4 , and 6 ; Notch3 ; and MMP-7 . Quantitative real-time PCR validated the differential expression of 13 genes, and immunohistochemistry confirmed the differences for four gene products. Conclusions: Expression profiling separates OC/PPC from DMPM and identifies a number of genes that are differentially expressed in these tumors. The molecular signatures unique to OC/PPC and DMPM should provide a molecular basis to study both tumors and new potential markers for facilitating their differential diagnosis.
104 citations
TL;DR: Evidence is provided that mesothelin expression is associated with prolonged survival in patients with high-grade ovarian serous carcinoma and with in vitro drug resistance, and differentiation status of tumor cells.
Abstract: Purpose: Mesothelin is an emerging marker for cancer diagnosis and target-based therapy, yet relatively little is known about the clinical significance of mesothelin expression in tumors. In this study, we correlate mesothelin immunoreactivity to clinicopathologic features in ovarian serous carcinoma. Experimental Design: Mesothelin expression levels were compared among 81 publicly available serial analysis of gene expression (SAGE) libraries of various carcinoma and normal tissue types. Immunohistochemistry using a well-characterized mesothelin monoclonal antibody (5B2) was done to evaluate mesothelin expression in 167 high-grade and 31 low-grade ovarian serous carcinomas. Immunohistochemistry staining scores were correlated with patient survival, tumor site, tumor grade, in vitro drug resistance, and differentiation status of tumor cells. Results: SAGE analysis showed that mesothelin was overexpressed in 50% of ovarian and pancreatic carcinomas but rarely in other cancer types, including liver, colon, kidney, prostate, and breast. Mesothelin immunoreactivity (>5% of tumor cells) was present in 55% of ovarian serous carcinomas with no difference in expression between high-grade and low-grade serous tumors ( P = 0.82). Based on Kaplan-Meier analysis, we found that a diffuse mesothelin staining (>50% of tumor cells) in primary high-grade ovarian carcinomas correlated significantly with prolonged survival in patients who had advanced-stage disease and had received optimal debulking surgery followed by chemotherapy ( P = 0.023). Mesothelin expression did not correlate significantly with patient age, tumor site, in vitro drug resistance, or tumor differentiation status ( P > 0.10). Conclusion: Our results provided new evidence that mesothelin expression is associated with prolonged survival in patients with high-grade ovarian serous carcinoma.
89 citations
TL;DR: It is demonstrated that this new genotyping method is capable of detecting zeptomoles of targets and achieve an allele discrimination selectivity factor >105 and analysis of target signals with a multiple-parameter approach to enhance assay sensitivity and specificity.
Abstract: This report describes a new genotyping method capable of detecting low-abundant point mutations in a homogeneous, separation-free format. The method is based on integration of oligonucleotide ligation with a semiconductor quantum dot (QD)-mediated two-color fluorescence coincidence detection scheme. Surface-functionalized QDs are used to capture fluorophore-labeled ligation products, forming QD-oligonucleotide nanoassemblies. The presence of such nanoassemblies and thereby the genotype of the sample is determined by detecting the simultaneous emissions of QDs and fluorophores that occurs whenever a single nanoassembly flows through the femtoliter measurement volume of a confocal fluorescence detection system. The ability of this method to detect single events enables analysis of target signals with a multiple-parameter (intensities and count rates of the digitized target signals) approach to enhance assay sensitivity and specificity. We demonstrate that this new method is capable of detecting zeptomoles of targets and achieve an allele discrimination selectivity factor >10 5 .
80 citations
TL;DR: It is found that cyclin E expression is useful in distinguishing an ETT from a placental site nodule and p16 expression is helpful in distinguishes an ETTs from a cervical squamous carcinoma.
Abstract: Epithelioid trophoblastic tumor (ETT) is a relatively uncommon trophoblastic tumor that can be confused with several trophoblastic and nontrophoblastic lesions, notably the placental site nodule and invasive squamous carcinoma of the cervix In this report, we analyzed the immunoreactivity of two cell cycle-regulated proteins, cyclin E and p16, in ETTs, placental site nodules and cervical squamous carcinomas to determine whether they are useful in their differential diagnosis Other trophoblastic lesions were also evaluated Using an H-score based on both percentage of positively stained cells and immunointensity, we found that ETTs demonstrated a much higher cyclin E staining score than placental site nodules (P 40 Only two placental site nodules had scores above the cutoff and both showed morphologic features that placed them in an intermediate position between a typical placental site nodule and an ETT, so-called "atypical PSN" p16 immunoreactivity, was not detected in any of the ETTs and placental site nodules, whereas it was strongly and diffusely positive in the vast majority of cervical squamous carcinomas examined (83/87 cases) (P<0001) Therefore, cyclin E expression is useful in distinguishing an ETT from a placental site nodule and p16 expression is useful in distinguishing an ETT from a cervical squamous carcinoma The majority of other types of trophoblastic lesions showed diffuse and intense nuclear immunoreactivity for cyclin E whereas none were positive for p16
80 citations
TL;DR: HLA-G is focally expressed in MM and breast carcinoma, while HLA-ABC expression is conserved, which may suggest a possible role in immune response evasion in some tumors.
Abstract: The aim of the present study was to evaluate HLA-G expression in breast carcinoma and malignant mesothelioma (MM). Malignant breast carcinoma effusions (46) and corresponding solid tumors (39) and 104 MM (26 effusions, 78 solid tumors) were analyzed using immunohistochemistry (IHC). HLA-G protein and mRNA expression were further studied using immunoblotting (IB) and RT-PCR. HLA-ABC expression was analyzed using flow cytometry (FCM). IHC showed predominantly focal HLA-G expression in 12 of 46 (26%) breast carcinoma effusions and 16 of 39 (41%) solid lesions. In MM, 20 of 78 (26%) solid lesions and 14 of 26 (54%) effusions were focally HLA-G positive. Expression in MM was higher in effusions (p=0.008). IB showed more frequent HLA-G expression in MM compared with breast carcinoma effusions, while RT-PCR showed HLA-G mRNA expression in both tumors. FCM showed conserved HLA-ABC expression in 15 of 15 effusions. Breast cancer patients with HLA-G-positive tumor cells had shorter disease-free survival (mean 37 vs 85, median 25 vs 31 months), though not significantly (p=0.14). In conclusion, HLA-G is focally expressed in MM and breast carcinoma, while HLA-ABC expression is conserved. However, the up-regulated expression of HLA-G in MM effusions and its possible association with shorter disease-free survival in advanced stage of breast carcinoma suggest a possible role in immune response evasion in some tumors.
78 citations
TL;DR: Digital karyotyping suggests that homozygous deletion or reduced expression of MKK4 may contribute to the development of ovarian serous carcinoma.
Abstract: Analysis of deleted chromosomal regions in tumors has historically led to the identification of tumor suppressor genes. In this study, we used digital karyotyping, a genome-wide, high-resolution technology, to search for chromosomal deletions in ovarian serous carcinoma, the most lethal gynecological malignancy in women. Five purified ovarian serous carcinomas were analyzed by digital karyotyping and small interstitial deletions at chromosome 17p were identified in two tumor samples. Aligning these two deletions identified an overlapping region that spanned 2.4 Mb which harbored a candidate tumor suppressor gene, mitogen-activated protein kinase kinase-4 (MKK4). Dual-color FISH analysis confirmed homozygous deletion of the MKK4 locus in both samples and RT-PCR demonstrated that both carcinomas lacked MKK4 transcript expression. Loss of heterozygosity of 17p occurred in 24 (86%) of 28 high-grade serous carcinomas including both cases with homozygous MKK4 deletion. Additionally, downregulation of MKK4 expression was found in 96 (75%) of 128 ovarian serous carcinomas as compared to benign ovarian tissues. These findings suggest that homozygous deletion or reduced expression of MKK4 may contribute to the development of ovarian serous carcinoma.
52 citations
TL;DR: Evidence is provided that Rsf-1 expression is primarily confined to high-grade serous carcinoma, the most aggressive ovarian cancer, and it is unlikely that Rs f-1 is a critical gene in the development of breast carcinoma.
TL;DR: Findings support a role of Rsf-1 in mediating disease progression and aggressive clinical behavior in this subset of ovarian carcinoma patients and is a novel prognostic marker for patients with post-chemotherapy recurrent disease.
TL;DR: In this paper, the authors describe characteristic chromosomal alterations in a consecutive series of 96 serous ovarian tumors by comparative genomic hybridization and analyze their association with different pathways of progression, histological grade, and clinical outcome.
Abstract: In this study, we describe characteristic chromosomal alterations in a consecutive series of 96 serous ovarian tumors by comparative genomic hybridization. We analyze their association with different pathways of progression, histological grade, and clinical outcome. The most striking difference between low-grade and high-grade serous carcinomas was seen in a higher incidence of chromosomal gains at 3q and 20q and losses of 13q in the high-grade carcinomas. In addition, high-level amplifications were significantly more frequent in high-grade carcinomas, specifically involving regions on 3q and 8q. Chromosomal amplifications of 19p and 19q and losses of 4q and 5q were among the most frequent changes found in both low-grade and high-grade carcinomas, distinguishing them from borderline tumors, which had very few recurrent alterations. The most significant impact on survival of patients with invasive carcinomas Stage II-IV was observed for high-level amplifications of regions on 8q (mean overall survival (OS) 69 versus 27 months, P = 0.0006). Interestingly, low-level gains on 8q do not show any impact compared to cases with no alteration. Surprisingly, chromosomal losses on 5q had a protective impact (mean OS 36 versus 76 months, P = 0.0007). Combination of both parameters resulted in two risk groups. Low risk: loss on 5q, no amplification on 8q (mean OS 84 months); high risk: no loss on 5q, amplification on 8q (mean OS 26 months). This difference is even more pronounced, if only cases with residual tumor of less than 2 cm are included, resulting in a 5-year survival of 100% and 0% (P = 0.0005).
TL;DR: During controlled ovarian stimulation with a GnRH antagonist protocol, luteal-phase support with micronized progesterone and 17beta-estradiol seem to increase endometrial L-selectin ligand expression in the luminal endothelium.
Abstract: Context: The impact of different types of luteal phase support on endometrial receptivity after ovarian stimulation has not been investigated. Objective: Our objective was to evaluate the impact of different luteal-phase support protocols on sex steroid levels and on endometrial expression of L-selectin ligand after ovarian hyperstimulation with a GnRH antagonist protocol. Patients and Design: Seventeen oocyte donors who underwent ovarian stimulation with a recombinant FSH/ganirelix acetate protocol were randomized into three groups: group I had no luteal-phase support; group II had luteal support with micronized progesterone; and group III had luteal support with progesterone plus 17β-estradiol. All donors had endometrial biopsies on the day of retrieval, and then 3, 5, and 10 d after retrieval. In addition, they had serum estradiol and progesterone measurements on d 3, 5, and 10. Main Outcome Measures: Endometrial L-selectin ligand expression was detected by immunohistochemical staining in the luminal a...
TL;DR: Controlled ovarian stimulation, using the antagonist protocol in this study, is associated with a reduction of L-selectin ligand expression during the implantation window which may adversely affect the endometrial environment.
TL;DR: RNA in situ hybridization shows that BCOX1 mRNA signal is mainly located in the cytoplasm of breast carcinoma epithelial cells, but not in those of normal epithel cells, stroma cells and lymphocytes, and Computational analyses predict that the putative B COX1 protein is a cy toplasmic protein.
01 Jan 2006
TL;DR: The future application of new advances in molecular genetic technology holds promise for the discovery of novel amplified chemoresistant genes, which may significantly affect the understanding of how tumors become Chemoresistant as well as provide a molecular platform for customized treatment of cancer patients.
Abstract: Resistance to chemotherapeutic agents represents a chief cause of mortality in cancer patients with advanced disease. Gene amplification has been shown to be one of the molecular mechanisms for tumors to escape the effect of chemotherapeutic drugs. The amplification and subsequent overexpression of the chemoresistant gene product are likely the results of tumor cell clonal expansion under the selective pressure of chemotherapeutic agents. In the past few decades, researchers have correlated the amplification of several target genes to drug resistance status in in vitro cell culture models. Although it is possible for gene amplification to be a widespread mechanismof chemore-sistance in cancer patients, only a few well-studied examples are presently available. Therefore, the future application of new advances in molecular genetic technology holds promise for the discovery of novel amplified chemoresistant genes, which may significantly affect our understanding of how tumors become chemoresistant as well as provide a molecular platform for customized treatment of cancer patients.