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B A Gusterson

Publications and source records attributed to B A Gusterson.

At least 37 records · Page 2Linked to original sources
PubMed1998

Experimental pathology and breast cancer genetics: new technologies.

The goal is to understand the critical events in tumour development and to apply this understanding to new approaches to diagnosis, prevention and treatment. It is clear that breast cancer is a heterogeneous disease at the molecular level, raising the possibility of a future functional classification based on mechanisms rather than morphology. These molecular phenotypes will also confer predictive value on the potential of the tumour to invade, metastasise and respond to or resist new therapeutic strategies. Studies of the genome in individuals are predicted also to enable the identification of polymorphisms that are associated with increased susceptibility to environmental factors, in addition to possibly explaining de novo variations in responses to drugs and radiation. The difficulty is how to identify which, of the approximately 30,000 genes expressed by a typical cancer cell alone or in combination, are the ones involved in these processes. The majority of breast cancers have such a multitude of molecular changes that it is difficult to distinguish between those that are critical to tumour progression and those that are epiphenomena of genetic instability and abnormalities in DNA repair. The identification of the earliest events in carcinogenesis must be the best hope, as it will then be possible to target the events that predispose to other secondary changes before they occur. Genomics and proteomics is the current hope to take us forward. This involves the application of a number of new technologies to facilitate the profiling of individual tumours, including laser-guided microdissection of microscopic lesions, comparative genomic hybridisation and loss of heterozygosity analysis of DNA using microarray technology to study DNA and expressed RNAs and protein profiling using 2D gel mass spectroscopy. With over 100,000 mRNAs and proteins to examine in complex tissues and in various combinations, there is obviously going to be a requirement for a large investment in computing power (bioinformatics) to facilitate the analysis of these data in relation to the clinical characteristics of the individual tumour and the patient.

Bacteriophages↗
PubMed1998

Differential expression of type XIV collagen/undulin by human mammary gland intralobular and interlobular fibroblasts.

Immunolocalisation of type XIV collagen/undulin in the human mammary gland revealed greater deposition in the interlobular stroma than in the intralobular stroma. The interlobular stroma is located between the breast lobules and their associated intralobular stroma. Fibroblasts isolated from the interlobular stroma synthesised 3- to 5-fold more type XIV collagen/undulin than intralobular fibroblasts, but synthesised type I and type IV collagens in similar amounts. The differential expression of type XIV collagen/undulin was maintained with passage in culture. The results suggest a role for type XIV collagen/undulin in stabilising dense collagen fibrils. The maintenance of two types of structurally distinct stromas may be important during developmental processes in the mammary gland.

Breast↗
PubMed1998

Prediction of local recurrence of ductal carcinoma in situ of the breast using five histological classifications: a comparative study with long follow-up.

The increased detection of ductal carcinoma in situ (DCIS) by mammographic screening and the more widespread use of breast-conserving surgery have led to a search for histological features associated with the risk of recurrence. In a case control study of 141 patients with long follow-up, we compared the ability of five morphological classifications to predict recurrence after local excision. A significant correlation was not found between recurrence and growth pattern when a traditional classification based on architecture was used nor with necrosis when a scheme based principally on this feature was employed. A correlation was, however, found between recurrence and "differentiation" as defined by nuclear features and cell polarization in a classification recently formulated by the European Pathologists Working Group (EPWG), but this failed to reach statistical significance at the 5% level. A stronger and statistically significant correlation was found between nuclear grade as defined by the EPWG and recurrence when cell polarization was disregarded, using the classification currently employed by the UK National Health Service and European Commission-funded Breast Screening Programmes. This was attributable to a small number of recurring cases being downgraded as a consequence of exhibiting polarized cells. A significant correlation between histology and recurrence was also observed using the Van Nuys classification, which is based on nuclear grade and necrosis. Whether the tumor recurred as in situ or invasive carcinoma was unrelated to histological classification, as was the time course over which it occurred. These findings strongly support the use of nuclear grade to identify cases of DCIS at high risk of recurrence after local excision, but further work is necessary to determine whether nuclear grade or necrosis is more appropriate to subdivide the non-high-grade cases.

Adult↗
PubMed1998

Predicted anti-oestrogen resistance in BRCA-associated familial breast cancers.

There is controversy concerning the prognosis of breast cancers arising in women carrying loss of function mutations in the breast cancer susceptibility genes BRCA1 and BRCA2. This study was carried out to assess the likely hormone dependence of this group of tumours in comparison with an age and grade matched group of control sporadic tumours. We used quantitative immunohistochemical analysis for the oestrogen receptor (ER), progesterone receptor (PgR), cyclin D1 and pS2 on sections of primary tumours and ductal carcinoma in situ (DCIS). Expression of PgR (P < 0.05) and cyclin D1 (P < 0.01) was low in the BRCA1- and BRCA2-associated cancers compared with sporadic cases. The low frequency of expression of ER (9/40), PgR (2/40) cyclin D1 (5/36) and pS2 (5/36) in the familial tumours indicates that the majority of such tumours will be oestrogen insensitive and unlikely to respond to hormonal manipulation even at the in situ stage in their evolution. The low level of PgR (2/40 cases) suggests that there may be some abnormality of transactivating function of the ER in these tumours.

Adult↗
PubMed1998

Breast development gives insights into breast disease.

AIMS: Studies of developing human breasts are essential for understanding the organogenesis as well as molecular pathogenesis of benign and malignant breast diseases. In this study we have examined the distribution of TGF-alpha, TGF-beta 1, tenascin-C and collagen type IV with the aim of starting to build a picture of the profile of molecules that may be involved in the development of the human breast. METHODS AND RESULTS: Ten fetal breasts (16 to 23 weeks of gestation) and 45 infant breasts, ranging in age from newborn to 2 years, were used in this study. Paraffin sections from these samples were immunostained with antibodies for these proteins and for Ki67 to elucidate the level of proliferative activity in different stages of breast development. TGF-alpha immunoreactivity was observed both in the stromal and the epithelial cells within fetal and infant breasts up to 25 days. TGF-beta 1 immunoreactivity was localized in the extracellular matrix. Tenascin-C was found around the neck of the developing breast bud and in the extracellular matrix of the infant with peaks in the newborn at 6-12 weeks. The immunoreactivity for type IV collagen was more intense in the region of the breast bud neck in the fetal breasts and reduced around the tips of lobular and terminal-end buds within the infant breasts. CONCLUSIONS: The distribution of the growth factors and extracellular matrix proteins within the developing human breast indicates that they play a significant role in different cellular compartments during morphogenesis and provides insights into breast disease.

Breast↗
PubMed1998

Interindividual variation in the metabolic activation of heterocyclic amines and their N-hydroxy derivatives in primary cultures of human mammary epithelial cells.

The heterocyclic amines, 2-amino-3-methylimidazo[4,5-f]quinoline (IQ), 2-amino-3,4-dimethylimidazo[4,5-f]quinoline (MeIQ) and 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP) are pyrolysis products formed when meat is cooked and are rodent mammary carcinogens. They are thought to be metabolically activated by N-hydroxylation, catalysed by cytochrome P450 (CYP), followed by O-acetylation catalysed by N-acetyltransferases. Primary cultures of human mammary epithelial cells (HMECs) prepared from up to 26 individuals for each compound, were treated with IQ, MeIQ, or PhIP (500 microM) or with N-hydroxy-2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (N-OH-PhIP) or N-hydroxy-2-amino-3-methylimidazo[4,5-f]quinoline (N-OH-IQ) (20 microM) and the levels of adduct formation in their DNA analysed by 32P-post-labelling. In order to investigate whether pharmacogenetic polymorphisms influence DNA adduct formation, the NAT2 genotype of each individual was determined by a polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) method that distinguishes between the wild-type and four variant alleles. Presence of two variant alleles designates a slow NAT2 acetylator, whereas individuals with one or two wild-type alleles are designated fast NAT2 acetylators. Interindividual variations in total DNA adduct levels ranged for IQ from 0.64-63.1 DNA adducts per 10(8) nucleotides (mean 7.80), for MeIQ from 1.99-17.8 (mean 6.63), for PhIP from 0.13-4.0 (mean 0.96), for N-OH-PhIP from 6.32-497 (mean 176) and for N-OH-IQ from 0.92-30.6 (mean 9.24). The higher adduct levels observed in cells treated with the N-OH metabolites suggests that N-hydroxylation is the rate-limiting step in HMECs and this may be due to low CYP levels. In contrast, the Phase II reaction catalysed by N-acetyltransferases is probably the major step in the metabolic activation of heterocyclic amines that occurs in the breast. Higher mean levels of heterocyclic amine-DNA adduct formation were detected in the cells of NAT2 fast acetylators compared with slow acetylators, with mean adduct levels per 10(8) nucleotides following IQ treatment, of 12.74 and 3.57 respectively, following PhIP treatment, of 1.20 and 0.74, respectively, following MeIQ treatment, of 7.90 and 5.08, respectively and following N-OH-PhIP-treatment, of 243.1 and 130.0, respectively. However, due to the large variations in adduct levels, these differences in mean values were not statistically significant with the limited number of individuals studied. This appears to be the first pilot study to demonstrate interindividual variations in the metabolic activation of heterocyclic amines and their metabolic intermediates in primary cultures of HMECs in vitro.

Arylamine N-Acetyltransferase↗
PubMed1998

Determination of the enzymes responsible for activation of the heterocyclic amine 2-amino-3-methylimidazo[4,5-f]quinoline in the human breast.

The heterocyclic amine 2-amino-3-methylimidazo[4,5-f]quinoline (IQ) is a potent mutagen and is a mammary carcinogen in rodents. In man, hepatic activation is carried out by cytochrome P450 (CYP) 1A2 and the ultimate DNA-reactive species is thought to be a nitrenium ion formed via an acetoxy ester of an exocyclic amino group. Because most human breast tumours are ductal in origin, we investigated the ability of cell types present in the mammary gland (breast epithelial cells and neutrophils present in milk) to activate IQ to DNA-binding species using 32P-postlabelling. Phorbol myristate acetate-stimulated neutrophils produced a similar pattern of IQ-DNA adducts to that produced by human mammary epithelial cells. Adduct formation in stimulated neutrophils was inhibited 80% by the myeloperoxidase inhibitor sodium azide (1 mM) but was not affected by proadifen (100 microM), indomethacin (100 microM), or eicosatetraynoic acid (100 microM), inhibitors of cytochrome P450, prostaglandin synthetase, and lipoxygenase, respectively. Similar experiments in human mammary epithelial cells showed no azide inhibition of IQ-DNA adduct formation. Analysis of gene expression by reverse transcription-polymerase chain reaction showed that CYP1A1 and CYP1B1, but not CYP1A2, were expressed at detectable levels in untreated mammary epithelial cells, whereas in neutrophils cytochrome P450 expression was confined to low levels of CYP1A1. In cultured epithelial cells, IQ-DNA adduct formation and CYP1A1, but not CYP1B1 expression were induced threefold by benz[a]anthracene treatment; IQ-DNA adduct formation was inhibited by alpha-naphthoflavone. Our results indicate possible mechanisms for the metabolic activation of dietary carcinogens in the human breast.

5,8,11,14-Eicosatetraynoic Acid↗
PubMed1997

Multistep carcinogenesis of breast cancer and tumour heterogeneity.

Breast cancer emerges by a multistep process which can be broadly equated to transformation of normal cells via the steps of hyperplasia, premalignant change and in situ carcinoma. The elucidation of molecular interdependencies, which lead to development of primary breast cancer, its progression, and its formation of metastases is the main focus for new strategies targetted at prevention and treatment. Cytogenetic and molecular genetic analysis of breast cancer samples demonstrates that tumour development involves the accumulation of various genetic alterations including amplification of oncogenes and mutation or loss of tumour suppressor genes. Amplification of certain oncogenes with concomitant overexpression of the oncoprotein seems to be specific for certain histological types. Loss of normal tumour suppressor protein function can occur through sequential gene mutation events (somatic alteration) or through a single mutational event of a remaining normal copy, when a germline mutation is present. The second event is usually chromosome loss, mitotic recombination, or partial chromosome deletion. Chromosome loci 16q and 17p harbour tumour suppressor genes, which seem to be pathognomonic for the development or progression of a specific histological subtype. There are an overwhelming number of abnormalities that have been identified at the molecular level which fit the model of multistep carcinogenesis of breast cancer. When the functions of all of these genes are known and how they participate in malignant progression, we will have the tools for a more rational approach to diagnosis, prevention and treatment. This review deals only with the factors that are involved in the conversion of a normal breast cell into a malignant cell rather than those required for invasion and metastases. A key critical long-term step in the molecular analysis of breast cancer will be to link the specific molecular damage with the effects of environmental carcinogens.

Breast Neoplasms↗
PubMed1997

DNA damage in breast epithelial cells: detection by the single-cell gel (comet) assay and induction by human mammary lipid extracts.

The presence of DNA damage in primary cultures of human mammary epithelial cells (HMECs), and the ability of extracts of human mammary lipid to cause such damage, has been investigated. Lipid extracts, prepared by a solid-phase procedure, and HMECs were obtained from breast tissue removed from healthy women (ages 18-50 years) who were resident in the UK and undergoing elective reduction mammoplasties. DNA single strand breaks (SSBs) were detected using the single-cell gel assay (comet assay) with alkaline electrophoresis (pH 12.3) and quantified by measuring comet tail length (CTL) (microm). Untreated HMECs and HMECs incubated (30 min, 37 degrees C) with a mammary lipid extract, with or without DNA-repair inhibitors hydroxyurea (HU) and cytosine arabinoside (ara-C), were examined. Ionizing radiation was used as a positive control. An active lipid extract gave a linear dose-response over the range 2.0-12.2 g equivalents. When MCL-5 cells, a line of metabolically-competent human lymphoblastoid cells, were used to compare the DNA-damaging properties of lipid extracts from six different donors, significant interindividual variations (median CTLs were 15.0, 53.5, 32.5, <4.0, <4.0 and 77.5 microm respectively) were observed. In eight subjects, the donors' HMECs were examined both before and after treatment with extracts of that donor's own lipid. Pre-existing DNA damage was detected in untreated HMECs from some donors (median CTLs 22.0-37.5 microm) that was not present in others (median CTLs 4.0-11.5 microm), and increases in CTL could be induced by incubation with the matching lipid extract (8 g equivalent) in more than half (five out of eight) the subjects examined (median CTL up to 111.0 microm). There was a tendency for the most active lipid extracts to be those obtained from donors whose HMECs also contained the most pre-existing DNA SSBs. The results of this pilot study may prove to be significant in relation to the initiation of breast cancer.

Adult↗
PubMed1997

Prognostic importance of thymidylate synthase expression in early breast cancer.

PURPOSE: To assess the prognostic importance of thymidylate synthase (TS) expression in breast tumors of patients with early-stage breast cancer, and to determine whether the benefit of chemotherapy (CT) is associated with TS expression. PATIENTS AND METHODS: The level of TS expression was evaluated in 210 node-negative and 278 node-positive patients enrolled onto Trial V of the International Breast Cancer Study Group ([IBCSG] formerly the Ludwig Breast Cancer Study Group) with a median follow-up time of 8.5 years. TS expression was assessed using the immunohistochemical method with the monoclonal antibody TS 106 on paraffin-embedded tissue specimens. RESULTS: High TS expression was associated with a significantly worse prognosis in node-positive but not in node-negative breast cancer patients. Twenty-seven percent of node-positive patients with high TS expression were disease-free at 10 years, compared with 44% of node-positive patients with low TS expression (P = .03). Forty-one percent of patients with node-positive high-TS-expressing tumors were alive after 10 years, compared with 49% of those with low TS expression (P = .06). The association between TS and disease-free survival (DFS) and overall survival (OS) was independent of other prognostic factors such as tumor size, tumor grade, nodal status, vessel invasion, estrogen receptor (ER)/ progestin receptor (PR) status, c-erb B-2, or Ki-67 expression. In node-positive patients, six cycles of standard adjuvant cyclophosphamide, methotrexate, and fluorouracil ([5-FU] CMF) CT improved DFS and OS compared with one cycle of perioperative CMF therapy. The magnitude of this benefit was greatest in patients whose tumors had high TS expression (P < .01 for DFS; P < .01 for OS). Node-negative patients demonstrated no difference in outcome to CT based on TS expression; however, the power to detect differences was limited by the small number of events in this group. CONCLUSION: In early-stage breast cancer, high TS expression is associated with a significantly worse prognosis in node-positive patients. Node-positive patients with high TS levels demonstrate the most significant improvement in DFS and OS when treated with six cycles of conventional adjuvant CMF therapy.

Antineoplastic Combined Chemotherapy Protocols↗
PubMed1996

Brk, a breast tumor-derived non-receptor protein-tyrosine kinase, sensitizes mammary epithelial cells to epidermal growth factor.

brk (breast tumor kinase) shows homology to the src family of non-receptor protein-tyrosine kinases and is expressed in breast carcinomas. In order to investigate the role of brk in breast tumor development, we have examined the growth and transformation properties of human mammary epithelial cells engineered to overexpress Brk. Interestingly, like c-Src, overexpression of Brk leads to sensitization to EGF, and also results in a partially transformed phenotype. Further investigation of the latter activity was attempted by mutational analysis, targeting key residues known to affect tyrosine kinase activity in Src-like kinases. Mutation of amino acid residue Lys-219 to Met, by analogy to Src, abolished both kinase activity and transformation capacity. Mutation of amino acid residue Tyr-447 to Phe, however, resulted in a decrease in transforming potential without affecting kinase activity. These results suggest that while Src and Brk share some functional properties, they act differently during transformation. These differences are discussed in the context of the mechanisms underlying breast cancer development.

3T3 Cells↗
PubMed1996

Regulation of urokinase-type plasminogen activator production by rat mammary myoepithelial cells.

Addition of basic fibroblast growth factor (bFGF) to a rat mammary gland myoepithelial cell line (25.5-G4.2.3) resulted in a six- to eightfold increase in cellular and secreted urokinase-type plasminogen activator (uPA) activity after a lag phase of 5-8 h. bFGF had no effect on the uPA activity of mammary epithelial cells. bFGF was active on myoepithelial cells over a narrow concentration range (0.5-2 ng/ml). The bFGF-induced increase in uPA activity was inhibited in a dose-dependent manner by hydrocortisone and transforming growth factor-beta1 (TGF-beta1). Hydrocortisone also inhibited the basal secretion of uPA, as did interleukin-1beta and phorbol myristate acetate, both of which increase uPA levels in other cell systems. The effects of bFGF could also be inhibited by factors which bind bFGF, e.g., heparin and methylamine a2-macroglobulin. TGF-beta1, but not bFGF, induced the synthesis of plasminogen activator inhibitor-1 in the myoepithelial cell line. Mammary gland myoepithelial cells contribute to the synthesis of and are located next to the basement membrane. Myoepithelial-derived uPA is probably associated with basement membrane turnover. The mammary gland basement membrane undergoes many cycles of remodeling and multiple mechanisms may be required to regulate uPA activity.

Animals↗
PubMed1996

Compartment switching of WNT-2 expression in human breast tumors.

WNT-2 is a secreted polypeptide with mitogenic effects in murine mammary epithelial cells, but its role in human cancer is unknown. Using RNase protection analysis of primary cell preparations and in situ hybridization analysis, we report that WNT-2 is expressed at low levels in normal human breast fibroblasts but not in epithelial cells. WNT-2 was found to be expressed at high levels in both the epithelium and stroma of 5 of 11 infiltrating carcinomas and 2 of 6 fibroadenomas. The high level of WNT-2 expression in tumor epithelium suggests that tumorigenesis may involve the ectopic expression of WNT-2 and the creation of an autocrine Wnt signaling loop.

Breast↗
PubMed1996

Expression of the receptor protein tyrosine kinase myk-1/htk in normal and malignant mammary epithelium.

We have recently reported the molecular characterization of a novel murine receptor PTK (myk-1), belonging to the eph-related family, whose expression was differentially regulated during mammary gland development and elevated in invasive mouse mammary tumours. In this communication we have investigated the cellular origin of myk-1 expression by in situ hybridisation and RNase protection. Murine antisense myk-1 probe specifically recognised ductal and alveolar epithelium in the resting mouse mammary gland. In normal human breast, the expression of htk, the human homologue, was also confined to the secretory luminal epithelial cells. RNase protection analysis of enriched luminal and myoepithelial cells prepared from human reduction mammoplasty tissue confirmed the luminal specificity. Elevated expression of htk was found in several human breast carcinoma cell lines as well as in primary, high grade, infiltrating, ductal carcinomas of the breast. The specific epithelial expression of the myk-1/htk receptor PTK suggest a specialised role of this eph-related PTK in the differentiation and/or maintenance of secretory epithelium in the adult.

Adult↗
PubMed1996

Fusion of the EWS gene to CHN, a member of the steroid/thyroid receptor gene superfamily, in a human myxoid chondrosarcoma.

The specific chromosomal translocation t(9;22)(q22-31;q11-12) has been observed in the myxoid variant of human chondrosarcoma. In agreement with this observation we report that the EWS gene located at chromosome band 22q12 becomes fused to CHN, a member of the steroid/thyroid receptor gene superfamily located at 9q22-31, in a skeletal myxoid chondrosarcoma. CHN appears to be the human homologue of the rat gene NOR1, which was recently identified as a sequence overexpressed in rat brain cells undergoing apoptosis. Our results also indicate that the chimaeric EWS-CHN gene encodes a EWS-CHN fusion protein in which the C-terminal RNA-binding domain of EWS is replaced by the entire CHN protein, comprising a long N-terminal domain, a central DNA binding domain and a C-terminal ligand-binding/dimerisation domain.

Amino Acid Sequence↗
PubMed1996

The molecular pathology of small round-cell tumours--relevance to diagnosis, prognosis, and classification.

Substantial improvements have been made in the treatment and survival of children with SRCT, resulting in an increased emphasis on precise histological diagnosis. Although diagnostic procedures such as electron microscopy and immunocytochemistry contribute in poorly differentiated cases, an accurate diagnosis can remain elusive in a proportion of SRCTs. The cytogenetic and molecular genetic abnormalities characteristic of the different SRCTs can now be consistently and rapidly identified from minimal quantities of tumour material, using the techniques of FISH and PCR. This, coupled with the identification of novel phenotypic characteristics, has had a major impact on SRCT diagnosis. The aim of a tumour classification is to identify disease entities which are biologically distinct and whose recognition is of clinical value. The recent advances described above demonstrate that the SRCTs are genotypically and phenotypically distinct tumour types and that the genetic abnormalities represent key alterations that influence both the morphology and the clinical behaviour of the tumour. This suggests that these advanced phenotypic and genotypic analyses should form an integral and complementary part of the laboratory assessment and clinical management of these forms of paediatric cancer.

Bone Neoplasms↗
PubMed1996

Interphase fluorescence in situ hybridization detection of t(2;13)(q35;q14) in alveolar rhabdomyosarcoma--a diagnostic tool in minimally invasive biopsies.

The identification of t(2;13)(q35;q14) is a useful aid in the accurate diagnosis of rhabdomyosarcoma, distinguishing it from other small round cell tumours and supporting the distinction between alveolar and embryonal forms. Cytogenetic analysis is difficult and with the increased use of minimally invasive biopsy methods and primary chemotherapy, adequate tumour material is not always available. To overcome these difficulties, two-colour interphase fluorescence in situ hybridization (FISH) to detect t(2;13)(q35;q14) was developed and its utility in assessing minimally invasive biopsies was investigated. Two cosmid clones which mapped proximal or distal to the breakpoint region 13q14 and one yeast artificial chromosome clone that mapped distal to the 2q35 breakpoint were identified. In interphase cells containing t(2;13)(q35;q14), the configuration of cosmid and yeast artificial chromosome signals demonstrated the presence of the translocation. Five cases of rhabdomyosarcoma were analysed by interphase FISH. The t(2;13)(q35;q14) was detected in all four alveolar tumours and was confirmed by cytogenetics in two cases, but was absent in one embryonal tumour. This sensitive detection method is applicable to minimal amounts of fresh or frozen tumour.

Adolescent↗