Biomedical subjects

Susanne M Gollin

Publications and source records attributed to Susanne M Gollin.

At least 19 recordsLinked to original sources

PubMed2007

Relationship between FRA11F and 11q13 gene amplification in oral cancer.

Common fragile sites (CFS) are nonstaining gaps or breaks in chromosomes that are expressed under conditions inducing replicative stress. CFS have been suggested to play a role in epithelial cancers by their association with loss of heterozygosity, loss of gene expression, and/or gene amplification in the form of homogeneously staining regions (hsrs). In oral squamous-cell carcinomas (OSCC), amplification of chromosomal band 11q13 occurs in the form of an hsr. We suggested previously that CFS flanking 11q13 may be susceptible to breakage induced by tobacco or other carcinogens and/or human papillomavirus, promoting formation of the 11q13 amplicon. Examination of OSCC cell lines with 11q13 amplification using fluorescence in situ hybridization showed loss of FRA11F sequences, whereas cell lines without 11q13 amplification displayed an intact FRA11F site. Cell lines with more complex 11q rearrangements expressed FRA11F in the form of an inverted duplication, characteristic of breakage-fusion-bridge cycles. Our findings suggest that gene amplification involving chromosomal band 11q13 in OSCC may be initiated by breakage at FRA11F.

Carcinoma, Squamous Cell↗

PubMed2006

The influence of clinical and demographic risk factors on the establishment of head and neck squamous cell carcinoma cell lines.

The purpose of this study was to generate stable cell cultures from head and neck squamous cell carcinomas (HNSCC), and retrospectively analyze the factors associated with successful cell line establishment. Fifty-two HNSCC cell lines were isolated from a series of 199 tumors collected between 1992 and 1997 at the University of Pittsburgh Medical Center. Cell lines were characterized at the molecular and cellular level to determine the features associated with cell line formation. Successful cell line formation was dependent on multiple factors, including gene amplification involving chromosomal band 11q13, local and/or regional involvement of lymph nodes, and alcohol usage. The establishment of HNSCC cell lines enriches the resources available for cancer research. Our findings indicate that generation of stable cell lines from HNSCC is biased towards tumors with a poor prognosis. Our 52 stable lines comprise one of the largest series of HNSCC cell lines in the literature, with complete demographic, histopathologic, clinical, and survival data.

Adult↗

PubMed2006

Mechanisms leading to nonrandom, nonhomologous chromosomal translocations in leukemia.

Nonrandom, reciprocal translocations between nonhomologous chromosomes are critical cellular events that lead to malignant transformation. Therefore, understanding the mechanisms involved in these chromosomal rearrangements is essential for understanding the process of carcinogenesis. There has been substantial discussion in the literature over the past 10 years about mechanisms involved in constitutional chromosomal rearrangements, including deletions, duplications, and translocations. Yet our understanding of the mechanisms of chromosomal rearrangements in cancer is still developing. This review presents what is known about the mechanisms involved in selected nonrandom chromosomal translocations in leukemia.

Cell Nucleus↗

PubMed2006

Overexpression of glycosylphosphatidylinositol (GPI) transamidase subunits phosphatidylinositol glycan class T and/or GPI anchor attachment 1 induces tumorigenesis and contributes to invasion in human breast cancer.

Based on the oncogenic role of phosphatidylinositol glycan (PIG) class U in human tumors, we explored the role of two additional subunits of the glycosylphosphatidylinositol (GPI) transamidase complex in human breast cancer. We found that PIG class T (PIG-T) and GPI anchor attachment 1 (GPAA1) were overexpressed in breast cancer cell lines and primary tumors. Forced expression of PIG-T and GPAA1 transformed NIH3T3 cells in vitro and increased tumorigenicity and invasion of these cells in vivo. Suppression of PIG-T expression in breast cancer cell lines led to inhibition of anchorage-independent growth. Moreover, we found that PIG-T and GPAA1 expression levels positively correlated with paxillin phosphorylation in invasive breast cancer cell lines. Furthermore, suppression of PIG-T and GPAA1 expression led to a decrease in paxillin phosphorylation with a concomitant decrease in invasion ability. These results suggest that the GPI transamidase complex is composed of a group of proto-oncogenes that individually or as a group contribute to breast cancer growth. This aberrant growth is mediated, at least partially, by phosphorylation of paxillin, contributing to invasion and progression of breast cancer.

Acyltransferases↗

PubMed2006

Overexpression of Cdc20 leads to impairment of the spindle assembly checkpoint and aneuploidization in oral cancer.

Defects in the spindle assembly checkpoint are thought to be responsible for an increased rate of aneuploidization during tumorigenesis. Despite a plethora of information on the correlation between BUB-MAD gene expression levels and defects in the spindle checkpoint, very little is known about alteration of another important spindle checkpoint protein, Cdc20, in human cancer and its role in tumor aneuploidy. We observed overexpression of CDC20 in several oral squamous cell carcinoma (OSCC) cell lines and primary head and neck tumors and provide evidence that such overexpression of CDC20 is associated with premature anaphase promotion, resulting in mitotic abnormalities in OSCC cell lines. We also reconstituted the chromosomal instability phenotype in a chromosomally stable OSCC cell line by overexpressing CDC20. Thus, abnormalities in the cellular level of Cdc20 may deregulate the timing of anaphase promoting complex (APC/C) in promoting premature anaphase, which often results in aneuploidy in the tumor cells.

Anaphase-Promoting Complex-Cyclosome↗

PubMed2006

Fatty acid synthase gene overexpression and copy number gain in prostate adenocarcinoma.

Cancer cells frequently exhibit a significant increase in overexpression and activity of fatty acid synthase (FASN). Elevated FASN pathway activity also occurs in prostate cancer, the second leading cause of cancer-related death in men in the United States. Studies show that genes associated with an increase in protein expression, such as HER2/neu in breast cancer, are associated with an increase in gene copy number as well as an increase in transcription. In the present study, we evaluated whether FASN follows a similar paradigm in prostate cancer. To date, elevated FASN expression in prostate cancer has not been correlated with gene copy number alterations. Using immunohistochemistry and fluorescence in situ hybridization analysis in paraffin-embedded tissue microarrays, we observed gene copy gain in 24% of all prostate adenocarcinoma specimens examined with concurrent increased FASN protein expression. Immunohistochemistry alone showed 59% of prostate cancer specimens in the same tissue microarray with high FASN expression. Increased FASN gene was observed in 53% of all prostate tissues expressing elevated FASN protein levels and in 2 of 5 prostate tumor cell lines tested. These findings suggest that FASN gene copy number increases may be involved in the resultant increase in FASN protein expression observed in prostatic disease.

Adenocarcinoma↗

PubMed2006

Loss of DNA polymerase zeta causes chromosomal instability in mammalian cells.

Rev3L encodes the catalytic subunit of DNA polymerase zeta (pol zeta) in mammalian cells. In yeast, pol zeta helps cells bypass sites of DNA damage that can block replication enzymes. Targeted disruption of the mouse Rev3L gene causes lethality midway through embryonic gestation, and Rev3L-/- mouse embryonic fibroblasts (MEFs) remain in a quiescent state in culture. This suggests that pol zeta may be necessary for tolerance of endogenous DNA damage during normal cell growth. We report the generation of mitotically active Rev3L-/- MEFs on a p53-/- genetic background. Rev3L null MEFs exhibited striking chromosomal instability, with a large increase in translocation frequency. Many complex genetic aberrations were found only in Rev3L null cells. Rev3L null cells had increased chromosome numbers, most commonly near pentaploid, and double minute chromosomes were frequently found. This chromosomal instability associated with loss of a DNA polymerase activity in mammalian cells is similar to the instability associated with loss of homologous recombination capacity. Rev3L null MEFs were also moderately sensitive to mitomycin C, methyl methanesulfonate, and UV and gamma-radiation, indicating that mammalian pol zeta helps cells tolerate diverse types of DNA damage. The increased occurrence of chromosomal translocations in Rev3L-/- MEFs suggests that loss of Rev3L expression could contribute to genome instability during neoplastic transformation and progression.

Animals↗

PubMed2006

Comprehensive genome and transcriptome analysis of the 11q13 amplicon in human oral cancer and synteny to the 7F5 amplicon in murine oral carcinoma.

11q13 amplification occurs in a wide variety of tumors, including almost half of oral squamous cell carcinomas (OSCC) where it has been correlated with a poor outcome. In this study, we compiled 3.6 Mb of DNA sequence in the 11q13 amplicon core and refined the physical map of the amplicon. In the process, we determined the genomic structure and normal tissue expression patterns of two recently identified genes, TAOS2/TMEM16A and MRGF, which reside in the amplicon core. We then quantified DNA copy number and mRNA expression of all genes in the 11q13 amplicon in cell lines and primary tumors from OSCC. With the exception of FGF3, FGF4, FGF19, and MRGF, all genes were overexpressed in most tumors with genomic amplification. Furthermore, we found that the expression of genes in the amplicon appeared to be highly coordinated, making it difficult to determine which gene or genes are driving amplification. However, in nonamplified primary tumors, three genes, TAOS2/TMEM16A, OCIM, and TPCN2, are frequently overexpressed, whereas CCND1 and EMS1 are not. These results suggest that in addition to CCND1 and EMS1, other important genes also may be target genes driving 11q13 amplification. We hypothesize that 11q13 amplification may be driven by a cassette of genes that provide growth or metastatic advantage to cancer cells. This is supported by the finding that the human 11q13 amplicon core is syntenic to mouse chromosomal band 7F5, which is frequently amplified in chemically induced murine OSCC. This article contains Supplementary Material available at http://www.interscience.wiley.com/jpages/1045-2257/suppmat

Animals↗

PubMed2006

Splenic small B-cell lymphoma with IGH/BCL3 translocation.

Isolated chromosomal translocations are important defining features of many non-Hodgkin lymphomas, especially of B-cell type. In contrast to some other translocations, the significance of IGH/BCL3 translocations is not well defined. Although often considered a feature of the ill-defined entity atypical chronic lymphocytic leukemia, very few cases are reported in which involvement of BCL3 and the precise B-cell neoplasm are both well documented. For this reason, we report a splenic-based CD5(-), CD10(-), CD43(-), CD23(-), CD103(-), FMC7(+), CD25(+) small B-cell lymphoma associated with epithelioid histiocyte clusters and a t(14;19)(q32;q13) representing an IGH/BCL3 translocation based on classical cytogenetic studies, chromosomal painting, and fluorescence in situ hybridization studies. The previously reported neoplasms with t(14;19)(q32;q13) or IGH/BCL3 translocations are also reviewed. The present case did not fall into any of the classic B-cell lymphoma categories and clearly did not represent chronic lymphocytic leukemia/small lymphocytic lymphoma. This case suggests that the IGH/BCL3 translocation may help to define a new clinicopathologic entity.

Aged↗

PubMed2005

Deletion 6q is not a characteristic marker of nodal lymphoplasmacytic lymphoma.

Lymphoplasmacytic lymphoma (LPL) is a small B-cell neoplasm with plasmacytic differentiation that does not fulfill the criteria for any other type of B-cell leukemia or lymphoma. In many cases, LPL is associated with Waldenström macroglobulinemia (WM), although WM may also be associated with other types of lymphoma. Recent studies have demonstrated that del(6q) is the most common structural abnormality in patients with bone marrow-based LPL. It is unknown whether del(6q) might also be associated with nodal LPL. We, therefore, examined 10 well-characterized LPL involving lymph nodes or other extramedullary tissues for del(6q) using paraffin section interphase fluorescence in situ hybridization (FISH). Dual-color FISH was performed using a chromosome 6 centromere probe (CEP6) and a probe for 6q21 (RP11-91C23). The latter probe has previously been reported as deleted in up to 63% of cases of bone marrow-based LPL. In contrast, no nuclei containing a del(6q) pattern were identified in any case of extramedullary LPL examined in this study, and 89-98.5% of nuclei contained a normal signal pattern. These results indicate that del(6q) is at least uncommon in nodal LPL, and cannot be employed as a diagnostic marker to identify this type of lymphoma. Furthermore, these findings suggest that nodal LPL and bone marrow-based cases of LPL may be associated with different cytogenetic findings.

Biomarkers, Tumor↗

PubMed2005

Somatic mutation and gain of copy number of PIK3CA in human breast cancer.

INTRODUCTION: Phosphatidylinositol 3-kinases (PI3Ks) are a group of lipid kinases that regulate signaling pathways involved in cell proliferation, adhesion, survival, and motility. Even though PIK3CA amplification and somatic mutation have been reported previously in various kinds of human cancers, the genetic change in PIK3CA in human breast cancer has not been clearly identified. METHODS: Fifteen breast cancer cell lines and 92 primary breast tumors (33 with matched normal tissue) were used to check somatic mutation and gene copy number of PIK3CA. For the somatic mutation study, we specifically checked exons 1, 9, and 20, which have been reported to be hot spots in colon cancer. For the analysis of the gene copy number, we used quantitative real-time PCR and fluorescence in situ hybridization. We also treated several breast cancer cells with the PIK3CA inhibitor LY294002 and compared the apoptosis status in cells with and without PIK3CA mutation. RESULTS: We identified a 20.6% (19 of 92) and 33.3% (5 of 15) PIK3CA somatic mutation frequency in primary breast tumors and cell lines, respectively. We also found that 8.7% (8 of 92) of the tumors harbored a gain of PIK3CA gene copy number. Only four cases in this study contained both an increase in the gene copy number and a somatic mutation. In addition, mutation of PIK3CA correlated with the status of Akt phosphorylation in some breast cancer cells and inhibition of PIK3CA-induced increased apoptosis in breast cancer cells with PIK3CA mutation. CONCLUSION: Somatic mutation rather than a gain of gene copy number of PIK3CA is the frequent genetic alteration that contributes to human breast cancer progression. The frequent and clustered mutations within PIK3CA make it an attractive molecular marker for early detection and a promising therapeutic target in breast cancer.

Apoptosis↗

PubMed2005

Uncommon mutation, but common amplifications, of the PIK3CA gene in thyroid tumors.

CONTEXT: As in many other human cancers, overactivation of the phosphotidylinositol 3-kinase (PI3K)/Akt signaling pathway occurs frequently in thyroid cancer, but the mechanism is not completely clear. OBJECTIVE: Because activating mutations and genomic amplification of the PIK3CA gene, which encodes the p110a catalytic subunit of PI3K, are common in many cancers, we sought to investigate this phenomenon in thyroid tumors. DESIGN: To search for PIK3CA mutations, we isolated genomic DNA from primary thyroid tumors of various types and performed direct sequencing of the exons of PIK3CA gene that carry the most common mutations in other cancers. We used real-time quantitative PCR to investigate genomic amplification of the PIK3CA gene. RESULTS: We found no PIK3CA gene mutations in 37 benign thyroid adenomas, 52 papillary thyroid cancers, 25 follicular thyroid cancers, 13 anaplastic thyroid cancers, 13 medullary thyroid cancers, and seven thyroid tumor cell lines. We found a C3075T single-nucleotide polymorphism in exon 20 of this gene in two cases. With a copy number of 4 or more defined as amplification, we found PIK3CA gene amplification in four of 34 (12%) benign thyroid adenomas, three of 59 (5%) papillary thyroid cancer, five of 21 (24%) follicular thyroid cancer, none of 14 (0%) medullary thyroid cancer, and five of seven (71%) thyroid tumor cell lines. The PIK3CA gene amplification and consequent Akt activation were confirmed by fluorescence in situ hybridization and Western blotting studies using cell lines, respectively. CONCLUSION: These data suggest that mutation of the PIK3CA gene is not common, but its amplification is relatively common and may be a novel mechanism in activating the PI3K/Akt pathway in some thyroid tumors.

Class I Phosphatidylinositol 3-Kinases↗

PubMed2005

Spindle multipolarity is prevented by centrosomal clustering.

Most tumor cells are characterized by increased genomic instability and chromosome segregational defects, often associated with hyperamplification of the centrosome and the formation of multipolar spindles. However, extra centrosomes do not always lead to multipolarity. Here, we describe a process of centrosomal clustering that prevented the formation of multipolar spindles in noncancer cells. Noncancer cells needed to overcome this clustering mechanism to allow multipolar spindles to form at a high frequency. The microtubule motor cytoplasmic dynein was a critical part of this coalescing machinery, and in some tumor cells overexpression of the spindle protein NuMA interfered with dynein localization, promoting multipolarity.

Antigens, Nuclear↗

PubMed2005

Human papillomavirus-16 associated squamous cell carcinoma of the head and neck (SCCHN): a natural disease model provides insights into viral carcinogenesis.

Uncertainty regarding the causality of human papillomaviruses (HPVs) in squamous cell carcinoma of the head and neck (SCCHN) necessitates better in vitro models. We carried out molecular analyses of a novel, naturally HPV-16-transformed SCCHN cell line (UPCI:SCC090) and show high copy number of HPV-16 DNA, present in a head to tail, tandemly repeated integrated state. Sequence analysis of the HPV-16 long control region (LCR) in UPCI:SCC090 revealed a deletion of 163 bp, removing a portion of the enhancer sequence, including the binding sites for the transcription factors YY1 and NF1. The E6 and E7 oncogenes of HPV-16 are expressed at high levels in this cell lines, as determined by quantitative reverse transcriptase-polymerase chain reaction (RT-PCR). UPCI:SCC090 contains wild-type tumour suppressor TP53 gene, and undetectable p53 protein, except after treatment with cisplatin, specific proteasome inhibitors or by E6 RNA interference, suggesting E6-dependent degradation of p53 in this cell line. The results of our studies are consistent with a causative role of HPV-16 in the pathogenesis of SCCHN.

Blotting, Southern↗

PubMed2005

Mechanisms leading to chromosomal instability.

Chromosomal instability is a common feature of cancer cells. Several cellular mechanisms lead to numerical and structural chromosomal instability in cancer cells, including defects in chromosomal segregation, cellular checkpoints that guard against reproduction of abnormal cells, telomere stability, and the DNA damage response. Human papillomavirus interferes with these processes, causing chromosomal instability and tumor formation in some of the epithelial cells which it infects. The rate of discoveries about the mechanisms leading to chromosomal instability in cancer cells is increasing rapidly. Although these mechanisms were thought to be unrelated, they are intimately intertwined, connecting the complex network of cellular pathways. Since chromosomal instability is undoubtedly a major cause of tumor evasion of therapy, understanding the mechanisms leading to chromosomal instability has major translational significance.

Cell Transformation, Neoplastic↗

PubMed2004

Cigarette smoke induces anaphase bridges and genomic imbalances in normal cells.

Exposure to cigarette smoke has long been linked to carcinogenesis, but the emphasis has been placed on mutational changes in the DNA sequence caused by the carcinogens in smoke. Here, we report an additional role for cigarette smoke exposure in contributing to chromosomal aberrations in cells. We have found that cigarette smoke condensate (CSC) induces anaphase bridges in cultured human cells, which in a short time lead to genomic imbalances. The frequency of the induced bridges within the entire population decreases with time, and this decrease is not dependent upon the p53-mediated apoptotic pathway. Additionally, we show that CSC induces DNA double stranded breaks (DSBs) in cultured cells and purified DNA. The reactive oxygen species (ROS) scavenger, 2' deoxyguanosine 5'-monophosphate (dGMP) prevents CSC-induced DSBs, anaphase bridge formation and genomic imbalances. Therefore, we propose that CSC induces bridges and genomic imbalances via DNA DSBs. Furthermore, since the amount of CSC added to the cultures was substantially less than that extracted from a single cigarette, our results show that even low levels of cigarette smoke can cause irreversible changes in the chromosomal constitution of cultured cells.

Antioxidants↗

PubMed2004

Mapping and analysis of HPV16 integration sites in a head and neck cancer cell line.

Human papillomavirus (HPV) is a circular double-stranded DNA virus implicated in at least 90% of cervical and anogenital cancers and has been observed in approximately 20% of squamous cell carcinomas of the head and neck (SCCHN). Transcription of the viral oncogenes E6 and E7 is regulated by expression of the E2 protein. Disruption of the E2 gene sequence due to viral integration results in upregulation of E6 and E7, which promote tumorigenesis by abrogating p53 and pRb functions. HPV integration sites in cervical and anogenital cancers have been mapped primarily to chromosomal fragile sites and in some cases have been shown to integrate within tumor suppressor genes or other cancer-related genes. To study viral integration sites in SCCHN, we examined an HPV16-infected SCCHN cell line cultured from a tongue-base tumor. HPV fluorescence in situ hybridization (FISH) revealed multiple integrated viral DNA copies in blocks throughout the genome. Sequential FISH and spectral karyotyping identified integration sites on chromosomes 3, 6, 9q, 13q and t(1;8)(q;?). Restriction site-polymerase chain reaction (RS-PCR) was performed to identify the viral-cellular junctions. Sequence analyses confirmed integration sites at 9q31.1 and 6p21 and revealed a novel junction at 16p12.3. Subsequent chromosome breakage studies suggested that the observed viral-cellular integration sites may have occurred within common fragile sites. Additional studies using RT-PCR for E6--E7 viral transcripts showed oncoprotein expression from episomal and integrated viral sequences. Our results suggest that viral integration of HPV in SCCHN appears to occur nonrandomly through targeting specific chromosomal sequences prone to breakage.

Adult↗

PubMed2004

DLC1 is unlikely to be a primary target for deletions on chromosome arm 8p22 in head and neck squamous cell carcinoma.

Allelic imbalance on chromosome arm 8p is common in head and neck squamous cell carcinoma (HNSCC). DLC1, a tumour suppressor gene inactivated in liver carcinogenesis and encoding a Rho GTPase activating protein (RhoGAP) maps to one of the deleted regions (8p21.3-22). In order to determine whether inactivation of DLC1 is involved in HNSCC, we have screened tumour cell lines for DLC1 mutations and expression. Pathological mutations were not identified in any of the 17 cell lines tested. Seven polymorphisms were identified; 13 of the 17 of cell lines were homozygous for all seven polymorphisms compared to only 2 of 17 controls suggesting a loss of heterozygosity in a majority of the cell lines. DLC1 expression was observed in all 11 HNSCC cell lines tested, thus excluding the possibility of transcriptional silencing of DLC1 by promoter hypermethylation. Overall, our data suggest that hemizygous deletions of the DLC1 locus are frequent in HNSCCs but this gene is unlikely to be primary target for inactivation on this chromosomal arm.

Carcinoma, Squamous Cell↗