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J Styles

Publications and source records attributed to J Styles.

At least 19 recordsLinked to original sources
PubMed2005

Influence of oestradiol and tamoxifen on oestrogen receptors-alpha and -beta protein degradation and non-genomic signalling pathways in uterine and breast carcinoma cells.

Tamoxifen acts as an oestrogen antagonist in the breast reducing cell proliferation, but in the uterus as an oestrogen agonist resulting in increased cell proliferation. Tamoxifen exerts its tissue-specific effects through the oestrogen receptors (ERalpha or ERbeta). The levels and functions of the two ERs affect the response of the target tissue to oestrogen and tamoxifen. We examined the control of ER stability in breast and uterine cell lines using western blotting and RT-PCR. In MCF-7 breast-derived cells, ERalpha and ERbeta proteins were rapidly degraded via the proteasome pathway in response to oestradiol; conversely tamoxifen stabilised both receptors. In Ishikawa uterine-derived cells, oestradiol and tamoxifen stabilised ERalpha but led to degradation of ERbeta by the proteasome pathway. Further investigations showed that oestradiol induced activation of the non-genomic ERalpha/Akt signalling pathway in MCF-7 cells. We have demonstrated that the alternative Erk signalling pathway is activated in Ishikawa cells following oestradiol treatment in the absence of an active proteasome pathway and therefore increased levels of ERbeta. In conclusion, our data have demonstrated tamoxifen or oestradiol control of ER subtype stability and that non-genomic activation of transcription pathways is cell specific.

Breast Neoplasms↗
PubMed2001

Short-term dosing of alpha-hydroxytamoxifen results in DNA damage but does not lead to liver tumours in female Wistar/Han rats.

It is now generally accepted that activation of tamoxifen occurs as a result of metabolism to alpha-hydroxytamoxifen. In this study, alpha-hydroxytamoxifen was given to female Wistar/Han rats (0.103 or 0.0103 mmol/kg, intraperitoneally, daily for 5 days). This resulted in liver DNA damage, determined by (32)P-post-labelling, of 3333 +/- 795 or 343 +/- 68 adducts/10(8) nucleotides, respectively (mean +/- SD, n = 4). Following HPLC separation, the retention times of the major alpha-hydroxytamoxifen DNA adducts were similar to those seen following the administration of tamoxifen. However, after rats were treated with alpha-hydroxytamoxifen (0.103 mmol/kg) for 5 days and the animals kept for up to 13 months, no liver tumours developed (0/7 rats), even with phenobarbital promotion (0/5 rats). GST-P foci were detected in the liver, but only after 13 months was their number or area significantly increased over the corresponding controls. When alpha-hydroxytamoxifen was given to female lambda/lacI transgenic rats (0.103 mmol/kg orally for 10 days) and the animals killed 46 days later, there was an approximate 1.8-fold increase in mutation frequency but no significant increase in G:C to T:A transversions as described after tamoxifen treatment. It is concluded that DNA damage alone, resulting from the short-term administration of alpha-hydroxytamoxifen, is not sufficient to initiate liver tumours even with phenobarbital promotion. As with tamoxifen, long-term exposure may be required to allow promotion and progression of transformed cells.

Animals↗
PubMed2000

Association of tamoxifen biliary excretion rate with prior tamoxifen exposure and increased mdr1b expression.

ATPase transporter proteins are commonly found in the hepatocyte canalicular membrane. Some of these, in particular the multidrug resistance (mdr1b) gene, have been previously demonstrated to be inducible genes. In this study, we found that tamoxifen induced expression of the mdr1b gene in the liver up to 40-fold after 14 days' exposure to tamoxifen in the diet at a concentration of 420 ppm. As tamoxifen and its metabolites are primarily excreted into the bile, we investigated if the increased expression of mdr1b in the liver following tamoxifen exposure had any effect on its excretion in rats. We found that the excretion of tamoxifen and its metabolites into bile was increased from 8 +/- 1% to 51 +/- 18% (mean +/- SD) of an administered dose of 180 nmol/kg over a collection period of 3 hr in rats that had received tamoxifen (35 mg/kg) orally for 12 days (plus a 3-day rest) prior to the experiment. These data suggest that prolonged treatment with tamoxifen may result in lower serum and tumour concentrations, due to a self-mediated enhancement of excretion via mdr1b gene-encoded P-glycoprotein. This may have implications for other drugs sharing the same route of excretion and co-administered with tamoxifen.

ATP Binding Cassette Transporter, Subfamily B, Mem↗
PubMed2000

Chemoprevention of breast cancer by tamoxifen: risks and opportunities.

The antiestrogen tamoxifen is widely used in the adjuvant therapy of breast cancers in women and helps to prevent the occurrence of breast tumors in healthy women. However, epidemiological studies have shown tamoxifen treatment to be associated with a 2- to 5-fold increased risk of endometrial cancer. In rats but not in mice, long-term administration of tamoxifen results in an increase in hepatocellular carcinomas. Mechanistically, this occurs through metabolic activation of the drug, mainly by the CYP3A family, to an electrophilic species, that causes DNA damage in target tissues, and subsequently leads to gene mutations. It is controversial whether low levels of DNA damage occur in human uterine tissues, and there is no evidence that this can be causally related to the mechanisms of carcinogenesis. In healthy women, the risk:benefits for the use of tamoxifen is in part related to the risk of developing breast cancer. The results from the carcinogenicity studies in rats do not predict the likelihood that women will develop liver cancer or indeed cancers in other organs. The mechanism of endometrial cancer in women remains unresolved, but the experience with tamoxifen has highlighted the potential problems that need to be addressed in the assessment of future generations of selective estrogen receptor modulators.

Animals↗
PubMed1996

Phase I trial and tumour localisation of the anti-EGFR monoclonal antibody ICR62 in head and neck or lung cancer.

The purpose of this study was to determine the effect of the first rat monoclonal antibody (MAb ICR62) to the epidermal growth factor receptor (EGFR) in a phase I clinical trial in patients with unresectable squamous cell carcinomas. This antibody effectively blocks the binding of EGF, transforming growth factor (TGF)-alpha and HB-EGF to the EGFR, inhibits the growth in vitro of tumour cell lines which overexpress the EGFR and eradicates such tumours when grown as xenografts in athymic mice. Eleven patients with squamous cell carcinoma of the head and neck and nine patients with squamous cell carcinoma of the lung, whose tumours expressed EGFR, were recruited. Groups of three patients were treated with 2.5 mg, 10 mg, 20 mg or 40 mg of ICR62 and a further eight patients received 100 mg. All patients were evaluated for toxicity using WHO criteria. Patients' sera were tested for the clearance of MAb ICR62 and the development of human anti-rat antibodies (HARA). No serious (WHO Grade III-IV) toxicity was observed in patients treated with up to 100 mg of antibody ICR62. Antibody ICR62 could be detected at 4 h and 24 h in the sera of patients treated with 40 mg or 100 mg of ICR62. Only 4/20 patients showed HARA responses (one at 20 mg, one at 40 mg and two at 100 mg doses) and of these only the former two were anti-idiotypic responses. In four patients receiving doses of ICR62 at 40 mg or greater, biopsies were obtained from metastatic lesions 24 h later and examined for the localisation of ICR62 using anti-rat antibody reagent. In these patients we showed the localisation of MAb ICR62 to the membranes of tumour cells; this appeared to be more prominent at the higher dose of 100 mg. On the basis of these data we conclude that MAb ICR62 can be administered safely to patients with squamous cell carcinomas and that it can localise efficiently to metastases even at relatively low doses.

Adult↗
PubMed1996

Phosphoinositide-dependent in vitro phosphorylation of profilin by protein kinase C. Phospholipid specificity and localization of the phosphorylation site.

Phosphoinositides bind to profilin and regulate actin-based cytoskeletal protein assembly. We report here that profilin is phosphorylated in vitro by protein kinase C (PKC) in the presence of phosphoinositides and micromolar concentrations of calcium. PKC-mediated phosphorylation of profilin was observed only in the presence of phosphoinositides; phosphatidylserine and diacylglycerol (known activators of PKC) and other lipids, including phosphatidic acid and phosphatidylglycerol phosphate, did not activate the phosphorylation. The activation of PKC-mediated phosphorylation of profilin by phosphoinositides was as follows: phosphatidylinositol (PI) 4-phosphate (K(m) = 18 microM) > PI 4,5-bisphosphate (K(m) = 30 microM) > PI (no activation). About 0.5 mol phosphate was incorporated per mol of profilin. Phosphorylation of profilin by PKC was not affected by the presence of various concentrations of actin. Phospho-amino acid analysis showed serine to be the only amino acid phosphorylated. The amino acid sequence of a phosphopeptide from CNBr-digested profilin corresponded to the COOH-terminal peptide of profilin (Ala-Ser-His-Leu-Arg-Ser-Gln-Tyr). Further digestion of this phosphopeptide by trypsin generated two phosphopeptides (Arg-Ser-Gln-Tyr and Ser-Gln-Tyr), thereby confirming that the phosphorylation site was the antepenultimate Ser (Ala-Ser-His-Leu-Arg-Arg-Ser(P)-Gln-Tyr).

Actins↗
PubMed1995

Perturbation of hepatocyte nuclear populations induced by iron and polychlorinated biphenyls in C57BL/10ScSn mice during carcinogenesis.

The induction of hepatocarcinogenesis by polychlorinated biphenyls (PCBs) in C57BL/10ScSn mice is markedly potentiated by iron. To investigate the effects of iron and PCBs on nuclear populations, C57BL/10ScSn mice received a single dose of iron-dextran (600 mg Fe/kg) and were fed a diet containing 0.01% of the PCBs mixture Aroclor 1254 for up to 6 months. DNA content of isolated nuclei and hepatocytes was estimated by flow cytometry. Cell suspensions and nuclei isolated from Aroclor treated mice after 6 months contained increased diploid (2N) populations compared to controls. In contrast, iron treatment of mice markedly enhanced fractions of octoploid (8N) nuclei by 2 weeks and this effect persisted over the 6 month period. When Aroclor 1254 and iron were administered together there was a synergistic increase in the mononucleated diploid fraction which was significant at 2 weeks and highly significant at 6 months. This became the predominant nuclear effect. At six months, Aroclor 1254 and iron, both alone and in combination, also increased the rate of DNA synthesis in hepatocytes as measured by bromodeoxyuridine (BrdU) incorporation. The chronic polyploidizing effect of iron overload alone was investigated further and shown to be proportional to the dose and was detectable as early as 2 days after 600 mg Fe/kg and 1 week after 150 mg Fe/kg. Polyploidization of nuclei was inhibited by the oral iron chelator CP94. Iron also induced a prolonged reduction in the incidence of binucleated cells. Histologically, nuclear enlargement due to iron was confined to the midzonal region of the liver lobule, whereas iron deposition was greatest in the periportal region. Iron (600 mg/kg) also caused increased nuclear polyploid states in hepatocytes of adult rats and gerbils. Similarly, weanling mice with a dominantly diploid cell population, when treated with iron (300 mg/kg), exhibited a significant shift to a tetraploid (4N) population and a marked increase in proliferation as measured by BrdU incorporation and proliferative cell nuclear antigen (PCNA) detection. These results indicate that Aroclor 1254 and iron induce changes in the mouse hepatocyte population that involve 2N and 8N nuclei respectively. The combination treatment leads to the emergence and proliferation of a mononucleated, diploid population as observed frequently in chemical hepato-carcinogenesis. The reason for the chronic polyploidizing effect of iron is unknown, but may imply both increased DNA synthesis and impairment of nuclear division with implications in human conditions of iron overload.

Animals↗
PubMed1994

Primary structure of the large subunit of trifunctional beta-oxidation complex from pig heart mitochondria.

The amino-terminal and internal sequences of the isolated large subunit of trifunctional beta-oxidation complex from pig heart mitochondria were determined by Edman degradation. The results demonstrated that the sequence of this novel beta-oxidation enzyme is identical with the sequence recently reported for a porcine gastrin binding protein that serves as the gastrin receptor on parietal cell surfaces. Evidence is provided to show that it is unlikely that the porcine gastrin binding protein has such a sequence. The data lead us to conclude that the mature large subunit of porcine trifunctional beta-oxidation complex is composed of 727 amino acid residues with a calculated molecular weight of 79,113, while the precursor of this long-chain fatty acid oxidation enzyme has a mitochondrial presequence consisting of 36 residues and a calculated M(r) of 83,099.

Amino Acid Sequence↗
PubMed1994

Immunotherapy with antibodies to the EGF receptor.

A series of rat monoclonal antibodies (MAbs) has been generated against the extracellular domain of the receptor for EGF which block the binding of EGF and TGF alpha to the receptor and inhibit the growth in vitro of a range of carcinoma cell lines that over-express the receptor for EGF. Some of these antibodies were able also to induce the complete regression of xenografts of EGFR-over-expressing tumours when treatment was started, either at the time of tumour inoculation or later when the tumours were established. The most effective of these antibodies was ICR62, which was also able to activate host immune effector functions. We conclude that antibodies which block growth-factor-ligand interaction can have a profound influence on the proliferative capacity of tumour cells in vivo and may have useful clinical application.

Animals↗
PubMed1994

The expression and processing of human beta-amyloid peptide precursors in Saccharomyces cerevisiae: evidence for a novel endopeptidase in the yeast secretory system.

In mammalian cells, the transmembrane beta-amyloid peptide precursor (beta-APP) undergoes a complex series of alternative proteolytic processing steps that result in the secretion of varying proportions of its extra-cellular domain (protease nexin II) and beta-amyloid peptide. The protein is also reinternalized and degraded in the endosomal-lysosomal system. The relative efficiencies of these competing processes determine the yield of beta-amyloid peptide. Several proteases have been implicated in this complex processing pathway, although none has been identified to date. The yeast secretory system contains proteases homologous to mammalian pro-hormone convertases and is susceptible to genetic manipulation. We therefore investigated the expression and processing of the beta-amyloid peptide precursors (beta-APP-695 and beta-APP-751) in Saccharomyces cerevisiae transformed with human beta-APP cDNA's. beta-APP (695 or 751) cDNA either with its authentic signal sequence or the yeast-derived prepro-alpha-factor leader, was inserted into a glucose-regulated expression vector and transfected into a protease-deficient yeast strain. In all instances, expression of beta-APP was about 1% of total protein. Protease protection studies indicated that either the natural human signal sequence or the alpha-factor leader sequence targetted beta-APP to the endoplasmic reticulum and inserted it with the amino-terminal domain in the lumen. All of the beta-APP fused to the alpha-factor leader proceeded to the trans-Golgi, where Kex2 endopeptidase removed the leader and released the normal amino-terminus of beta-APP. About one-half of the beta-APP was also cleaved at the "alpha-secretase" site in the middle of the beta-peptide sequence, 12 residues before the membrane-spanning sequence. A fraction of the alpha-secretase-cleaved beta-APP appeared in the culture medium; however, most of it associated with the exterior of the cells. The carboxyl-terminal fragments formed by cleavage at the alpha-secretase site accumulated in the membranes. Other proteolytic processes generated membrane-associated carboxyl-terminal fragments that also resembled those found in mammalian cells. These results indicate that the secretory system of S. cerevisiae possesses proteases with specificities similar to the mammalian enzymes that process beta-APP.

Amino Acid Sequence↗
PubMed1993

Peptide compositions of the cerebrovascular and senile plaque core amyloid deposits of Alzheimer's disease.

The pathological findings of Alzheimer's disease include amyloid deposition in cerebral blood vessels and in senile plaques. Both deposits are known to include peptides that contain a common sequence. Both forms of amyloid were isolated and their peptide compositions were determined. The peptides were resolved by size-exclusion chromatography in 70% formic acid, and reverse-phase chromatography in 60% formic acid, 0-40% acetonitrile. Senile plaque amyloid cores contain about 25% protein, about 70% of which is composed of peptides containing the beta-amyloid sequence. Amino-terminal sequencing of the core amyloid peptides (CAPs) revealed extensive amino-terminal heterogeneity, with variable amounts of blocked amino termini. Matrix-assisted, laser-desorption-time-of-flight mass spectrometry of the CAP mixture revealed an array of peptides the molecular weights of which corresponded to peptides beginning with each of the first 11 amino acids of the beta-peptide sequence and ending with Ala-42 of that sequence. The carboxyl-terminal residues were identified by tandem mass spectrometry of chymotrypsin digests. CAP possessed a minor degree of carboxyl-terminal heterogeneity. Cerebrovascular amyloid peptides (CVAPs) possessed minor degrees of both amino- and carboxyl-terminal heterogeneity. The major CVAP commenced at Asp-1 and ended at Val-40. Minor components of CAP possessed masses of 8000-9000 Da and the same amino-terminal residues as the major components of CAP. They may be precursors to the smaller CAPs. The differences in amino termini and carboxyl termini of CAPs and CVAPs suggest that the two types of amyloid form by different pathways, on which they encounter different proteases.

Alzheimer Disease↗
PubMed1993

Radioimmunolocalisation in breast cancer using the gene product of c-erbB2 as the target antigen.

Lymph node status is still the single most important prognostic factor in breast cancer. Axillary surgery remains the only reliable means of providing this information. This pilot study evaluates using a highly specific radiolabelled monoclonal antibody to provide equivalent information by a non-invasive technique. After optimisation of labelling conditions, our first antibody, ICR12 (against the gene product of c-erbB-2) was evaluated in a mouse model system. Twenty-four hours post i.v. injection the mice were killed and their organs, blood and tumours harvested for counting. Tumour localisation was four times greater than that into normal tissues, reaching 20% injected dose per gram of tumour. Eight patients have had this Tc99m-ICR12. Patient selection was by immunocytochemical staining of fine needle aspirates from the patient's own breast cancer. After intravenous administration of the immunoconjugate, tomographic images were obtained at 24 h. These results were compared to the subsequent histopathological examinations. Three patients acted as normal controls, one patient was negative due to inappropriate sampling, and two patients had strong membrane staining and provided excellent tumour localisation to both breast primary and regional node metastases. A further two patients only had moderate antigen expression on staining and did not localise well. The good performance of this radiolabelled antibody with patients that strongly stain for the antigen encourages the development of this system as both a method of staging breast cancer and a potential means of immunotherapy in this subgroup of patients.

Adult↗
PubMed1993

Immunotherapy of human tumour xenografts overexpressing the EGF receptor with rat antibodies that block growth factor-receptor interaction.

Athymic mice bearing xenografts of human tumours that overexpress the receptor (EGFR) for EGF and TGF alpha have been used to evaluate the therapeutic potential of three new rat monoclonal antibodies (mAbs) directed against two distinct epitopes on the extracellular domain of the human EGFR. The antibodies, ICR16 (IgG2a), ICR62 (IgG2b) and ICR64 (IgG1), have been shown (Modjtahedi et al., 1993) to be potent inhibitors of the growth in vitro of a number of human squamous cell carcinomas because they block receptor-ligand interaction. When given i.p. at 200 micrograms dose, the three antibodies were found to induce complete regression of xenografts of the HN5 tumour if treatment with antibody commenced at the time of tumour implantation (total doses: ICR16, 3.0 mg; ICR62, 1.2 mg; ICR64, 2.2 mg). More importantly when treatment was delayed until the tumours were established (mean diam. 0.5 cm) both ICR16 and ICR62 induced complete or almost complete regression of the tumours. Furthermore, treatment with a total dose of only 0.44 mg of ICR62 was found to induce complete remission of xenografts of the breast carcinoma MDA-MB 468, but ICR16 was less effective at this dose of antibody and only 4/8 tumours regressed completely. ICR16 and ICR62 were poor inhibitors of the growth in vitro of the vulval carcinoma A431, but both induced a substantial delay in the growth of xenografts of this tumour and 4/8 tumours regressed completely in the mice treated with ICR62 (total dose 2.2 mg). Although ICR16 and ICR64 were more effective than ICR62 as growth inhibitors in vitro, ICR62 was found to be substantially better at inducing regression of the tumour xenografts due perhaps to additional activation of host immune effector functions by the IgG2b antibody. We conclude that these antibodies may be useful therapeutic agents that can be used alone without conjugation to other cytotoxic moieties.

Animals↗
PubMed1992

Preoperative imaging of colorectal cancers. Targeting the epithelial membrane antigen with a radiation-labeled monoclonal antibody.

The epithelial membrane antigen (EMA) is expressed by the majority of colorectal cancers but has not previously been investigated as a target for radiation-labeled monoclonal antibodies (MoAb) in the imaging of patients with colorectal cancer. A rat IgG2a MoAb that recognizes EMA, ICR2, was labeled with Indium-111 (100 megabecquerel per milligram [MBq/mg]MoAb) using the bicyclic anhydride of the chelating agent diethylene triamine pentacetic acid (ccDTPA) and was administered intravenously to 22 patients known to have or thought to have colorectal cancer. Daily gamma camera imaging was performed for 3 days during the time between the administration of the radiation-labeled antibody and surgical procedure. At operation, the biopsies were done of the tumors and the normal colon, and the uptake of radiation-labeled MoAb was measured in a gamma well-counter. Immunocytochemistry for EMA expression also was done on resected tumors. Independent unblinded and blinded reporting was done on all scans. The sensitivity of 111In-ICR2 for detecting cancers preoperatively was 80% and 60%, respectively, on unblinded and blinded reporting, and the corresponding specificity 20% and 60%. The low unblinded specificity was attributable to a false-positive localization in severely dysplastic benign tumors (n = 2) and inflammatory tissue (n = 2). Liver metastases present in three patients were cold relative to normal liver. Lymph node metastases were localized in 1 of 6 patients preoperatively. The mean absolute uptake of 111In-ICR2 in tumor tissue was 7.75 +/- 3.77 x 10(-3) percent of injected dose per gram, and the ratio to normal colon was 2.10 +/- 0.92:1. On immunohistochemistry, EMA was expressed by 16 of the 17 primary cancers, both dysplastic adenomas, and all nodal metastatic deposits. EMA-negative tumors (1 cancer + 1 colonic lipoma) had negative antibody scans, and patients whose tumor was negative or only focally positive for EMA expression had lower tumor/normal colon ratios of radioactivity (1.30 +/- 0.26 versus 2.45 +/- 0.65, P = 0.005) on gamma well-counting of excised specimens. These results suggest a possible role for 111In-ICR2 in the detection of colorectal cancer and metastases but not its liver deposits.

Aged↗