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J C Barrett

Publications and source records attributed to J C Barrett.

At least 163 records · Page 9Linked to original sources
PubMed1996

A defined chromosome 6q fragment (at D6S310) harbors a putative tumor suppressor gene for breast cancer.

Recent evidence obtained by cytogenetic and molecular studies indicates that in breast cancer chromosome 6q is often affected by genetic changes suggesting the existence of putative tumor suppressor genes (TSGs). However the function of gene(s) on this chromosome in breast cancer suppression is not understood. To substantiate further the presence of breast cancer related TSGs at 6q and to define their location, we first performed microcell-mediated transfer of chromosome 6 to CAL51 breast cancer cells for studying possible suppression of malignant phenotype and secondly, we analysed DNAs from 46 primary breast cancers for loss of constitutive heterozygosity (LOH) using 24 poly-morphic microsatellite markers. The chromosome transfer resulted in loss of tumorigenicity and reversion of other neoplastic properties of the microcell hybrids. Polymorphism analysis of single hybrids revealed that they harbored only a small donor chromosome fragment defined by the marker D6S310 (6q23.3-q25) and flanked by D6S292 and D6S311. The LOH data suggest that four tumor suppressor gene loci mapped to the central and distal portion of 6q may be independently deleted in breast cancer. One of these regions corresponds to the region identified by chromosome transfer.

Breast Neoplasms↗
PubMed1996

Molecular modeling of the amino-terminal zinc ring domain of BRCA1.

The equine herpes virus zinc ring domain nuclear magnetic resonance structure was used for homology-based modeling of the amino-terminal zinc ring domain of the BRCA1 breast and ovarian cancer susceptibility gene. The zinc ring domain of BRCA1 is of particular interest because it is the location of significant and frequently occurring missense (Cys(61)Gly, Cys(64)Gly, and Cys(64)Tyr) and frameshift (185delAG) mutations observed in several high-risk kindreds. The BRCA1 zinc ring domain possesses 54% sequence similarity with the equine herpes virus zinc ring domain. The model structure undergoes little conformational variance after 140 ps of solvated molecular dynamics. This model proposes BRCA1 zinc ring domain residues that may play a role in DNA binding and/or protein-protein interactions. These predictions provide a point of departure for the design of mutants to probe BRCA1 zinc ring domain functionality.

Amino Acid Sequence↗
PubMed1996

Molecular genetic evidence of the occurrence of breast cancer as an integral tumor in patients with the hereditary nonpolyposis colorectal carcinoma syndrome.

BACKGROUND: The hereditary nonpolyposis colorectal carcinoma (HNPCC) syndrome is an autosomal dominant genetic disorder caused by the inheritance of a mutation in one of a family of genes encoding DNA mismatch repair (MMR) proteins. HNPCC manifests as genetic instability in linked tumors. Clinically, the syndrome is characterized by early onset malignancies, primarily of the colon and endometrium, with an increased incidence of tumors at other gastrointestinal sites, upper urologic tract, ovary, and pancreas as well. However, the inclusion of breast cancer as an integral tumor of this syndrome is controversial. METHODS: Mutation screening of MMR genes was carried out by single strand conformation polymorphism (SSCP) and sequencing analyses of genomic DNA prepared from normal lymphocytes. Expression analysis was performed by SSCP. Sequence analyses of cDNA was prepared from breast tumor tissue and normal lymphocytes. Genetic instability was assessed by comparing the electrophoretic mobility of polymerase chain reaction (PCR) products using multiple microsatellite markers. RESULTS: A 4-bp frameshift mutation in the hMLH1 gene was found to segregate with disease in the germlines of the affected members of a large kindred HNPCC. Expression of only the mutant allele was observed in the breast cancer tissue of one family member, however both alleles were observed in her normal tissue. This breast cancer exhibited widespread microsatellite instability, as did breast cancers obtained from several other HNPCC kindreds. CONCLUSIONS: These data indicate that breast cancer may result from the inheritance of a mutant MMR gene, and that breast cancer may occur as an integral tumor in the HNPCC syndrome.

Adaptor Proteins, Signal Transducing↗
PubMed1996

Molecular genetic analysis of clear cell adenocarcinomas of the vagina and cervix associated and unassociated with diethylstilbestrol exposure in utero.

BACKGROUND: Prenatal exposure to the synthetic estrogen diethylstilbestrol (DES) is associated with the subsequent development of clear cell adenocarcinoma of the lower reproductive tract in young women, and data concerning the molecular genetic alterations involved in the etiology of this tumor type have not previously been reported. Such knowledge would be of potential value by providing insight into the molecular mechanisms of hormonal carcinogenesis in general, as well as by suggesting molecular markers for risk assessment in the estrogen-exposed population. METHODS: A total of 24 samples of clear cell adenocarcinoma of the vagina or cervix, 16 associated with exposure in utero to DES and 8 with no history of DES exposure, were obtained as archival fixed and embedded tissue specimens. DNA was purified from these tissues and used to examine a number of biologically plausible molecular genetic endpoints for tumor specific alterations. RESULTS: No evidence was found for mutations in the K-ras or H-ras protooncogenes, the Wilms' tumor (WT1) tumor suppressor gene, or the estrogen receptor gene. Sporadic overexpression of the p53 tumor suppressor gene was detected in some tumor cell nuclei by immunohistochemistry, but in the absence of detectable p53 gene mutation. Genetic instability as manifested by somatic mutation of microsatellite repeats was widespread in these tumors, with evidence of microsatellite instability in all DES-associated tumors examined, and in 50% of those tumors not associated with DES exposure. CONCLUSIONS: These data are consistent with the hypothesis that the induction of genomic instability may be an important mechanism of DES-induced carcinogenesis.

Adenocarcinoma, Clear Cell↗
PubMed1996

Escape from senescence in hybrid cell clones involves deletions of two regions located on human chromosome 1q.

Human normal cells have been shown to undergo a limited number of cell doublings, a phenomenon termed cellular senescence. Human chromosome 1 has been implicated in this process, and several lines of evidence indicate that there is a senescence-inducing gene or genes on human chromosome 1q. Our approach to analyze the senescence-inducing effect of chromosome 1 includes the use of somatic cell hybrid revertants. We show here that fusion of a hypoxanthine phosphoribosyl transferase-negative mouse cell line (A9) containing a human neo-tagged chromosome 1 with an immortal hamster cell line (10W-2) results in cell hybrids that senesce after a few population doublings. Rare revertants that had escaped senescence were obtained after one large fusion experiment. Thirty-five nonsenescent hybrids were obtained from a total of approximately 1 million hybrids, and 25 of these were subjected to further analysis. The presence of a single copy of human chromosome 1 in the revertant hybrids was confirmed by fluorescence in situ hybridization analysis using a chromosome 1-specific painting probe. No visible translocations or deletions of chromosome 1 were observed in any of the hybrids. Deletion mapping revealed that 11 (56%) of the hybrids analyzed had lost one or more markers on chromosome 1q. Two regions with deletions were detected, one of which has been shown to be implicated in the senescence-inducing effect exerted by chromosome 1 following monochromosome transfer (P. J. Vojta et al., manuscript submitted for publication). The present study suggests that two separate loci on human chromosome 1q may be of importance for the induction of senescence. Moreover, this set of nonsenescent revertants could be useful for future detailed analyses of the senescence-inducing loci.

Animals↗
PubMed1996

Evidence for two senescence loci on human chromosome 1.

Microcell-mediated introduction of a neo-tagged human chromosome 1 (HC-1-neo) into several immortal cell lines has previously been shown to induce growth arrest and phenotypic changes indicative of replicative senescence. Somatic cell hybridization studies have localized senescence activity for immortal hamster 10W-2 cells to a cytogenetically defined region between 1q23 and the q terminus. Previous microcell-mediated chromosome transfer experiments showed that a chromosome 1 with an interstitial q-arm deletion (del-1q) lacks senescence inducing activity for several immortal human cell lines that are sensitive to an intact HC-1-neo. In contrast, our studies reveal that the del-1q chromosome retains activity for 10W-2 cells, indicating that there are at least two senescence genes on human chromosome 1. Sequence-tagged site (STS) content analysis revealed that the q arm of the del-1q chromosome has an interstitial deletion of approximately 63 centimorgans (cM), between the proximal STS marker DIS534 and distal marker DIS412, approximately 1q12 to 1q31. This deletion analysis provides a candidate region for one of the senescence genes on 1q. In addition, because this deletion region extends distally beyond 1q23, it localizes the region containing a second senescence gene to approximately 1q31-qter, between DIS422 and the q terminus. STS content analysis of a panel of 11 10W-2 microcell hybrid clones that escaped senescence identified 2 common regions of loss of 1q material below the distal breakpoint of del-1q. One region is flanked by markers DIS459 and ACTN2, and the second lies between markers WI-4683 and DIS1609, indicating that the distal 1q senescence gene(s) localizes within 1q42-43.

Animals↗
PubMed1996

Estrogen-induced cell transformation and DNA adduct formation in cultured Syrian hamster embryo cells.

To study the possible involvement of DNA damage in cell transformation induced by estrogens, we examined whether DNA adduct formation is elicited in cultured Syrian hamster embryo (SHE) cells treated with estrogens and their derivatives by means of the 32P-postlabeling assay. Morphological transformation of the cells was induced by treatment with diethylstilbestrol (DES) at 1-10 micrograms/mL for 24 h but not by treatment with its derivatives trans, trans-dienestrol (alpha-DIES) or cis, cis-dienestrol (beta-DIES) at 1-10 micrograms/mL for 24 h. Similarly, DNA adduct formation was elicited by exposure of SHE cells to DES at 1-10 micrograms/mL for 24 h but not by either alpha-DIES or beta-DIES. Treatment of SHE cells with DES at 1-10 micrograms/mL for 2 h in the presence of exogenous metabolic activation with rat liver post-mitochondrial supernatant enhanced morphological transformation in a dose-dependent manner. Our previous studies have demonstrated that exposure of SHE cells to DES under the same conditions with exogenous metabolic activation induces somatic mutations at the Na+/K+ ATPase locus. Therefore, we examined whether with exogenous metabolic activation DES induced DNA adduct formation in SHE cells. DNA adducts were not detected when SHE cells were treated with DES at 1-10 micrograms/mL for 2 h in the presence of exogenous metabolic activation. Treatment with 17 beta-estradiol (E2), 2-hydroxyestradiol (2-OH E2), or 4-hydroxyestradiol (4-OH E2) at 1 microgram/mL for 24 h induced DNA adduct formation in the cells, in parallel with the induction of cell transformation. The rank order of DNA adduct formation was 4-OH E2 > 2-OH E2 > E2. The results indicate that estrogens induce DNA adduct formation in cultured SHE cells, but the induction may not be the only mechanism relevant to the initiation of cell transformation.

Animals↗
PubMed1996

Mutational analysis of the p21/WAF1/CIP1/SDI1 coding region in human tumor cell lines.

p21/WAF1/CIP1/SDI1 is an important cell-cycle mediator with tumor suppressor gene capabilities, and its inactivation could potentially lead to tumor progression. Because tumor suppressor genes are commonly inactivated by somatic and germline mutations, we analyzed a variety of human tumor cell lines for p21 mutations. We used single-strand conformational analysis and direct sequencing to identify possible mutations in the p21 coding region. Two base-alterations were observed in 41 immortalized human tumor cell lines. A previously reported polymorphism that results in a serine-to-arginine amino-acid substitution at codon 31 was found in 24% (10 of 41) of the tumor cell lines but was also found in 10% (six of 62) of normal parental DNAs tested and 7% (three of 43) of normal DNAs from patients with primary endometrial tumors. Another nucleotide substitution found at codon 80 resulted in the replacement of threonine with methionine. Codon 80 changes were found in 7% (three of 41) of the tumor cell lines (all endometrial) and in 2% (one of 62) of the normal parental DNAs. This change was not found in any of the primary endometrial tumors examined. The biological activity of these base changes was analyzed by using in vitro cyclin-dependent kinase 2-cyclin A kinase assays and calcium phosphate transfections. We observed that wild-type p21 and the p21 variants had similar growth-inhibitory abilities. Thus, our results suggest that mutation of the p21 gene is not prevalent in human tumor cell lines and is not a probable mechanism of inactivation of this gene.

Base Sequence↗
PubMed1996

Changes in jun N-terminal kinase activation by stress during aging of cultured normal human fibroblasts.

The molecular changes associated with the aging process include the reduced activity of transcription factors (such as AP-1) and an impaired response to stress, which has been well documented in the case of the heat-shock (HS) response. Using human diploid fibroblasts of early and late passages as an in vitro model for aging, we elucidated changes in the activation of jun N-terminal kinases (JNKs), which play an important role in the mammalian stress response. We found that early-passage cells exhibited a greater degree of JNK activation in response to HS and ultraviolet (UV) C light treatments than did late-passage cells. Decreased JNK activation was dependent on the number of passages but was not affected by varying doses of UV irradiation. Analysis of protein kinase A, mitogen-activated protein kinase, and src-related tyrosine kinases revealed no decreased activities in aged cells, indicating a selective rather than generalized decrease in kinase activities during aging. A further understanding of this impaired activation of JNK may provide insights into the mechanisms of stress response and cellular aging.

Calcium-Calmodulin-Dependent Protein Kinases↗
PubMed1996

Chronic inhalation toxicity and carcinogenicity testing of respirable fibrous particles. Workshop report.

On May 8-10, 1995, a workshop on chronic inhalation toxicity and carcinogenicity testing of respirable fibrous particles was held in Chapel Hill, North Carolina. The workshop was sponsored by the Office of Pollution Prevention and Toxics, U.S. Environmental Protection Agency (EPA), in collaboration with the National Institute of Environmental Health Sciences (NIEHS), the National Institute for Occupational Safety and Health (NIOSH), and the Occupational Safety and Health Administration (OSHA). The goal of the workshop was to obtain input from the scientific community on a number of issues related to fiber testing. Major issues for discussion were: (i) the optimal design and conduct of studies of the health effects of chronic inhalation exposure of animals to fibers; (ii) preliminary studies which would be useful guides in designing the chronic exposure study; (iii) mechanistic studies which would be important adjuncts to the chronic exposure study to enable better interpretation of study results and extrapolation of potential effects in exposed humans; and (iv) available screening tests which can be used to develop a minimum data set for (a) making decisions about the potential health hazard of the fibers and (b) prioritizing the need for further testing in a chronic inhalation study. After extensive discussion and debate of the workshop issues, the general consensus of the expert panel is that chronic inhalation studies of fibers in the rat are the most appropriate tests for predicting inhalation hazard and risk of fibers to humans. A number of guidances specific for the design and conduct of prechronic and chronic inhalation studies of fibers in rodents were recommended. For instance, it was recommended that along with other information (decrease in body weight, systemic toxicity, etc.), data should be obtained on lung burdens and bronchoalveolar lavage fluid analysis to assist in establishing the chronic exposure levels. Lung burden data are also important for quantifying aspects of risk assessment related to dosimetric adjustments before extrapolation. Although mechanistic studies are not recommended as part of the standard chronic inhalation studies, the expert panel stressed the need for obtaining mechanistic information as far as possible during the course of subchronic or chronic inhalation studies. At present, no single assay and battery of short-term assays can predict the outcome of a chronic inhalation bioassay with respect to carcinogenic effects. Meanwhile, several short-term in vitro and in vivo studies that may be useful to assess the relative potential of fibrous substances to cause lung toxicity/carcinogenicity have been identified.

Administration, Inhalation↗
PubMed1996

Prostate cancer--biology of metastasis and its clinical implications.

Prostate cancer is one of the most commonly diagnosed cancers and is a major cause of cancer death in men. Although the majority of the diagnosed prostate cancers will remain localized and never produce clinical symptoms during the lifetime of the host, a subset of these cancers will progress to a more malignant state requiring therapeutic intervention. Acquisition of metastatic ability by prostatic cancer cells is the most lethal aspect of prostatic cancer progression. Once this has occurred, definitive therapy is required before the initially localized metastatic cells escape from the prostate. At present, metastatic prostate cancer is incurable. Therefore, there is an urgent need to develop molecular markers that can be used to predict the metastatic potential of prostate cancers. Using somatic cell hybridization, we have demonstrated that acquisition of metastatic ability requires both the loss of metastasis-suppressor function(s) and the activation of oncogenes. In further studies using micro-cell-mediated chromosomal transfer, we located genes on human chromosome, 8, 10cen-q23, 11p11.2-13, and 17pter-q23, which, when introduced into rat prostatic cancer cells, are capable of suppressing their metastatic ability without affecting their tumorigenicity or growth rate in vivo. Initially we focused upon the human chromosome 11p11.2-13 region to clone metastasis-suppressor gene(s) positionally. One such gene, termed KAI-1, encodes a membrane glycoprotein. KAI-1 has been mapped to the p11.2 region of human chromosome 11 by fluorescence in-situ hybridization analysis. Expression of KAI-1 has been detected in all normal human tissues thus far tested, including prostate tissue. When introduced into rat metastatic prostatic cancer cells, KAI-1 significantly suppressed the metastasis without affecting the tumor growth rate. KAI-1 expression is high in human normal prostate and benign prostatic hyperplasia but is dramatically lower in cancer cell lines derived from metastatic prostate tumors.

Animals↗
PubMed1996

Mutation of MSH3 in endometrial cancer and evidence for its functional role in heteroduplex repair.

Many human tumours have length alterations in repetitive sequence elements. Although this microsatellite instability has been attributed to mutations in four DNA mismatch repair genes in hereditary nonpolyposis colorectal cancer (HNPCC) kindreds, many sporadic tumours exhibit instability but no detectable mutations in these genes. It is therefore of interest to identify other genes that contribute to this instability. In yeast, mutations in several genes, including RTH and MSH3, cause microsatellite instability. Thus, we screened 16 endometrial carcinomas with microsatellite instability for alterations in FEN1 (the human homolog of RTH) and in MSH3 (refs 12-14). Although we found no FEN1 mutations, a frameshift mutation in MSH3 was observed in an endometrial carcinoma and in an endometrial carcinoma cell line. Extracts of the cell line were deficient in repair of DNA substrates containing mismatches or extra nucleotides. Introducing chromosome 5, encoding the MSH3 gene, into the mutant cell line increased the stability of some but not all microsatellites. Extracts of these cells repaired certain substrates containing extra nucleotides, but were deficient in repair of those containing mismatches or other extra nucleotides. A subsequent search revealed a second gene mutation in HHUA cells, a missense mutation in the MSH6 gene. Together the data suggest that the MSH3 gene encodes a product that functions in repair of some but not all pre-mutational intermediates, its mutation in tumours can result in genomic instability and, as in yeast, MSH3 and MSH6 are partially redundant for mismatch repair.

Base Sequence↗
PubMed1996

Induction of apoptosis by c-Fos protein.

The role of c-Fos in apoptosis was examined in two Syrian hamster embryo cell lines (sup+I and sup-II) and a human colorectal carcinoma cell line (RKO), using the chimeric Fos-estrogen receptor fusion protein c-FosER. As previously reported, contrasting responses were observed when these two cell lines were placed under growth factor deprivation conditions; sup+I cells were highly susceptible to apoptosis, whereas sup-II cells were resistant. In this report, we show that the activated c-FosER protein induces apoptosis in sup-II preneoplastic cells in serum-free medium, indicating that c-Fos protein can induce apoptotic cell death in these cells. c-Fos-induced apoptosis was not blocked by the protein synthesis inhibitor cycloheximide, suggesting that the c-Fos transcriptional activation activity is not involved. This conclusion was further supported by the observation that overexpression of v-Fos, which is highly proficient in transcriptional activation but deficient in the transcriptional repression activity associated with c-Fos, did not induce apoptosis. Constitutively expressed Bcl-2 delayed the onset of low-serum-induced apoptosis in sup+I cells and enhanced survival in sup-II cells. Further, coexpression of Bcl-2 and c-FosER in sup+I or sup-II cells protected the cells from c-FosER-induced apoptosis. The possibility that c-FosER-induced apoptosis requires a p53 function was examined. Colorectal carcinoma RKOp53+/+ cells, which do not normally undergo apoptosis in serum-free medium, showed apoptotic DNA fragmentation upon expression and activation of c-FosER. Further, when the wild-type p53 protein was diminished in the RKO cells by infection with the papillomavirus E6 gene, subsequent c-FosER-induced apoptosis was blocked. The data suggest that c-Fos protein plays a causal role in the activation of apoptosis in a p53-dependent manner. This activity does not require new protein synthesis and is blocked by overexpression of Bcl-2 protein.

Animals↗
PubMed1996

Breast cancer and environmental risk factors: epidemiological and experimental findings.

Breast cancer has long been associated with reproductive hormone exposures. Recently, greater attention has been focused on environmental exposures that may be responsible for some proportion of breast cancer incidence. Several etiologic aspects are discussed. A number of chemicals induce breast cancer in rodents--including solvents, pesticides, and polycyclic aromatic hydrocarbons--and these might serve as leads for studies in humans. In women, strong links have been established between breast cancer risk and ionizing radiation. Evidence for nonionizing radiation (electromagnetic field) exposures and breast cancer is suggestive, albeit limited. Occupational exposures have not been identified as breast cancer risks, but several associations need further study, including solvents and pesticides. Time of life when exposures take place is important, and this claim is strongly supported by data on cigarette smoking and radiation. Also, basic research has demonstrated that mammary tissue is more susceptible to carcinogenesis at certain periods of breast development. Likewise, prenatal, neonatal, and adolescent exposures deserve continuing attention. Research on etiology of breast cancer should measure environmental exposures and take into account the time of life at which these occur. Complex interactions between exogenous and endogenous carcinogenic agents need further focus, as modulated by varying genetically determined individual susceptibilities.

Animals↗
PubMed1996

Metastasis suppressor genes for prostate cancer.

To examine the role of human chromosomes in the development of metastatic prostate cancer, we introduced a copy of human chromosomes into highly metastatic Dunning R-3327 rat prostatic cancer cells by microcell-mediated chromosome transfer. Each microcell hybrid clones containing human chromosomes 8, 10, 11, and 17, respectively, showed decreased ability to metastasize to the lung, without any loss of tumorigenicity. This finding demonstrates that these human chromosomes contain metastasis suppressor genes for prostate cancer. Spontaneous deletion of portions of human chromosomes was observed in human chromosome 10, 11, and 17 studies. In the human chromosome 8 study, irradiated microcell-mediated chromosome transfer was performed to enrich chromosomal arm deletions of human chromosome 8. Relationships between the size of human chromosomes introduced into microcell hybrid clones and the number of lung metastases produced by the clones were analyzed to determine which part of human chromosomes contained metastasis suppressor gene(s) for prostate cancer. Molecular and cytogenetic analyses of microcell hybrid clones demonstrated that metastasis suppressor genes on human chromosomes 8, 10, and 11 were located on 8p23-q12, 10q, 11p13-11.2, respectively. Further analyses are proposed to confirm the potentially useful advantage of this assay system to identify metastasis suppressor gene(s) for prostate cancer.

Animals↗