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B Hartley-Asp

Publications and source records attributed to B Hartley-Asp.

At least 37 records · Page 2Linked to original sources
PubMed1991

Estramustine binding protein (EMBP) in rat R3327 Dunning tumors: partial characterization and effect of hormonal withdrawal, hormonal replacement, and cytotoxic treatment on its expression.

Expression of estramustine-binding protein (EMBP) was determined in eight variants of the Dunning R3327 rat prostate adenocarcinoma. This secretory protein was previously isolated from the normal rat prostate and binds estramustine and estromustine, the metabolites exerting anti-mitotic and anti-proliferative activity of the anticancer agent estramustine phosphate (EstracytR). EMBP was found in relatively high amounts only in the androgen-responsive G and H tumors from intact hosts, whereas low (AT-1, HI-F) or nondetectable levels (HI-S, AT-3, MAT-LyLu, MAT-Lu) were obtained in the androgen-independent tumors. Castration decreased EMBP expression in both G and H tumors, as did estradiol and estramustine when given separately to intact rats. Supplementation of castrates with exogenous androgen stimulated EMBP expression above the pre-castration level and was further enhanced when combined with estradiol and in particular estramustine. Partial physicochemical characterization of EMBP in G and H tumors was performed by using ligand-based and immunoblotting techniques. [3H]Estromustine was bound to cytosols and salt extracts from tumors with the same affinity and displayed similar surface-charge distribution, sedimentation behavior, and subunit composition as found for ventral and dorsolateral prostatic tissue from tumor-bearing rats. This study indicates that the synthesis of EMBP is under androgenic control and that its expression may be correlated to androgen responsiveness, metastatic potential, and androgen receptor content but not to growth rate and morphology of the tumors. EMBP may therefore provide a mechanism for concentration of cytotoxic activity at the target site in EMBP-positive tumors and help in the evaluation of antitumor effects obtained when administering estramustine.

Adenocarcinoma↗
PubMed1991

Effect of calcium and calcium antagonists on 45Ca influx and cellular growth of human prostatic tumor cells.

Calcium and calmodulin play significant roles in DNA synthesis and cell proliferation. In this work the effects of verapamil, trifluoperazine, and tamoxifen on 45Ca uptake and cell growth in human prostatic tumor cells (DU 145) and human fibroblast cells (1 BR) were studied. Although the maximum proliferation was achieved at a concentration of around 2 mM CaCl2 in both DU 145 and 1 BR, growth of DU 145 cells was considerably greater than 1 BR at all calcium concentrations (0.1-4 mM). Calcium uptake experiments, using 45Ca, revealed that the unstimulated 45Ca uptake in 1 BR fibroblasts was 4-5 times higher than in DU 145 cancer cells. Depolarization with high extracellular K caused a 2-3-fold increase in 45Ca influx in 1 BR but only 25-55% increase in DU 145 cells. Verapamil caused a significant inhibition of cell growth with an IC50 value of 55 microM. Verapamil paradoxically increased 45Ca uptake in both unstimulated and K-stimulated DU 145 cells. Whereas unstimulated 45Ca uptake could be blocked by very low concentrations of lathanum (10 microM), much higher concentrations (1-10 mM) were required to completely block uptake in K-depolarized cells. Both trifluoperazine and tamoxifen also inhibited cell proliferation with an IC50 concentration of approximately 5 microM. These drugs, had, however, no effect on 45Ca uptake either in unstimulated or depolarized cells. The results suggest that voltage-gated calcium channels exist in both DU 145 cancer cells and fibroblasts. However, verapamil, in contrast to 1 BR, failed to block these channels in DU 145 cells. The mechanism of antiproliferative action of verapamil may be related to the observed, although paradoxical, increase in cellular calcium. The effect of trifluoperazine and tamoxifen does not involve changes in transmembrane calcium movements but could be mediated by their inhibition of calmodulin-mediated reactions within the cell.

Calcium↗
PubMed1990

Characterization of muscarinic cholinergic receptors in membrane preparations from rat prostatic adenocarcinoma.

The binding characteristics of 3H-quinuclidinyl benzilate (QNB) to muscarinic sites in isolated plasma membrane fractions from R-3327 Dunning tumors (H and AT-1 sublines); ventral, dorsolateral prostate; and urinary bladder of the rat were studied. QNB binding to all preparations, except from AT-1 tumors, was specific, saturable, and of high affinity. The AT-1 tumors completely lacked specific QNB binding. The muscarinic receptor density in H tumors was twofold and twentyfold higher than that in the ventral prostate and dorsolateral prostate respectively. The receptor density in the urinary bladder was approximately twofold higher than that in H tumors. The Kd values in H tumors and ventral prostate were very similar and significantly higher than that in dorsolateral prostate or the urinary bladder. QNB binding in H tumors was strongly inhibited by classical muscarinic receptor antagonists atropine and scopolamine, but poorly by the agonists carbacholine and pilocarpine. In contrast to scopolamine or atropine, inhibition by pirenzepine and AF-DX116 was relatively low. These data indicate that the muscarinic receptor in Dunning H tumors is of M3 type.

Animals↗
PubMed1990

Estramustine phosphate reversibly inhibits an early stage during adenovirus replication.

Estramustine phosphate, an estradiol-mustard conjugate, was shown to reversibly inhibit a stage during the first hour of productive adenovirus 2 infection of HeLa cells. This drug, employed in the therapy of advanced prostatic cancer, specifically interacts with microtubule-associated proteins (MAPs) of the cytoskeleton. The results obtained under physiological conditions in vivo suggest a MAPs-interference with the microtubule-mediated vectorial migration of the virus inoculum to the nucleus. Virus attachment, uncoating kinetics and the appearance of established uncoating intermediates were not affected.

Adenoviridae Infections↗
PubMed1989

Identification by C-banding of two human prostate tumour cell lines, 1013L and DU 145.

Two human prostate carcinoma cell lines are easily distinguished by C-banding. DU 145 has one metacentric chromosome with a large centromeric band, 1 metacentric and 2 acrocentric chromosomes with interstitial C-bands. The Y chromosome was present in 48% and 42% of the cells at passages 83 and 106, respectively, whereas in 1013L only one metacentric chromosome contained distinct extra C bands--one at the centromere and one interstitially. The Y chromosome was present in 72% of the cells at passage 15 and in 20% at passage 22, but by passage 44 it had disappeared. Karyotype analysis using G-banding revealed that DU 145 had retained several of its original markers, and demonstrated multiple marker chromosomes in 1013L.

Chromosome Aberrations↗
PubMed1989

Differential reaction of secretory and non-secretory proteins in hormone-treated Dunning tumor.

Tumor cells from of the Dunning R-3327 PAP tumor were grown in vivo in the flank region of male Copenhagen rats for 4 months. Untreated control animals, castrated animals, and untreated or castrated animals supplemented with testosterone and estramustine (alone or in combination) bearing tumors were used for immunocytochemical studies of the tumor cells. Antibodies against the following secretory proteins were applied to paraffin sections of formalin-fixed tissue: anti-SVS II, anti-PBP, anti-transglutaminase, anti-acid phosphatase (isoenzymes, isoelectric points [pI] 5.6, 7.1, 8.0), non-secretory proteins comprised antiglucocorticoid-receptor, and antibodies against extracellular matrix proteins and intermediate filaments. A differential expression of marker proteins subsequent to the various treatments was observed immunohistochemically. Castration induced a loss of secretory activity, an increase in keratin-immunoreactive cells, and regressive activity in the secretory cells. Immunoreactivity of the glucocorticoid receptor in the nuclei of the basal cells was also decreased. Testosterone substitution only partially restored secretory activity of tumor cells in castrated animals. In experiments where estramustine had been administered to normal or castrated animals, metaplastic transformation of the epithelium, focal reduction, or increase of secretory and/or regressive activity and persistence of glucocorticoid receptor-like immunoreactivity was observed. The findings indicate that different hormonal situations provide conditions for the development of rather heterogeneous reaction patterns of different tumor cells, allowing partial regression of individual clones of tumor cells along with stimulation of others.

Adenocarcinoma↗
PubMed1989

The effect of estramustine, nor-nitrogen mustard and tauromustine on macromolecular labelling in the human prostatic tumour cell line 1013L.

To further clarify the mode of action of estramustine, the influence on macromolecular synthesis in the human prostatic tumour cell line 1013L was investigated. Cell treatment with estramustine, nor-nitrogen mustard and tauromustine, followed by radioactive nucleotide and leucine incorporations, as a measure of RNA, DNA and protein labelling, were carried out. The initial effect of estramustine clearly differed from that obtained after treatment with nor-nitrogen mustard and tauromustine. No inhibition of DNA synthesis was found whereas an inhibition of overall RNA synthesis was predominant. Adaption of an established RNA separation method was used in an indepth study of RNA labelling after estramustine treatment. An inhibition of 29S, 18S and 4-7S RNA was found after estramustine treatment, indicating disturbances in either RNA processing or RNA transport. The lack of 45S RNA labelling additionally indicates pre-ribosomal inhibition.

Affinity Labels↗
PubMed1988

Distribution and metabolism of estramustine in HeLa cells and the human prostatic tumour cell line 1013L.

The metabolism of estramustine [estradiol-3-N-bis(2-chloroethyl) carbamate] was investigated in the human prostatic tumour cell line 1013L and the human cervix tumour cell line HeLa S3. Uptake studies revealed that estramustine (EM), and its 17-ketoanalogue estromustine (EoM), differed in their nuclear binding pattern in 1013L cells but not in HeLa cells. Most of the nuclear radioactivity from both EM and EoM was found in the fraction containing the majority of the phospholipids. HPLC studies on EM-treated 1013L cells showed the presence of the oxidized metabolite EoM, in the medium, an enrichment of estradiol and estrone in whole cells and EM and EoM bound to the nuclear protein matrix. Similar studies on the HeLa cell line showed a completely different pattern, no metabolites other than EoM were found in the cell medium and whole cells but several very lipophilic metabolites were found bound to the nuclear protein matrix. On investigation of other tumour cell lines these metabolites were found to be unique to HeLa cells. The results extend our knowledge concerning EM and demonstrate that the cell line 1013L is a relevant model system for studying drugs active against human prostatic tumours.

Cell Line↗
PubMed1988

Antiinvasive activity of estramustine on malignant MO4 mouse cells and on DU-145 human prostate carcinoma cells in vitro.

Estramustine (EM) is a conjugate of estradiol and nor-nitrogen mustard (nor-HN2), which is effective in the treatment of prostate cancer. We have compared the effect of EM with that of the known microtubule inhibitor vinblastine (VLB) on the following functions of malignant MO4 mouse cells and of DU-145 human prostate cancer cells in vitro: directional migration, invasion; and the organization and the assembly/disassembly equilibrium of microtubule complexes. The circular area covered by cells migrating from an aggregate explanted on a solid substrate was taken as an index of directional migration. Invasion was studied through confrontation of MO4 or DU-145 cells with fragments of embryonic chick heart in organ culture. Microtubules were investigated immunocytochemically and through immunodetection on protein blots. VLB and EM inhibited directional migration and invasion of MO4 and DU-145 cells in a dose-dependent manner; equimolar combinations of estradiol plus nor-nitrogen mustard did not mimic these effects. At anti-invasive concentrations VLB led to partial disassembly of microtubule complexes, whereas EM resulted in an abnormal pattern of microtubule complexes without alteration of the overall assembly/disassembly equilibrium. Combined treatment with VLB and EM resulted in an enhanced VLB effect, namely complete disassembly. In all tests DU-145 cells were more sensitive to both VLB and EM than were MO4 cells, and the effects were less reversible. The present experiments showed that EM shares an anti-invasive activity with other microtubule inhibitors.

Animals↗
PubMed1988

Anti-tumour, toxicological and pharmacokinetic properties of a novel taurine-based nitrosourea (TCNU).

A novel nitrosourea, 1-(2-chloroethyl)-3-[2-(dimethylaminosulfonyl) ethyl]-1-nitrosourea (TCNU) tauromustine, has been investigated in a broad anti-tumour screen and, in depth toxicology and initial pharmacokinetics carried out. TCNU and its two metabolites were found to exhibit equal or better oral efficacy than that of BCNU, CCNU, MeCCNU or chorozotocin against L1210 leukemia, Walker mammary carcinoma, Lewis Lung, Harding Passey melanoma and colon carcinoma C26. The toxicological profile of TCNU after acute and 3 months treatment was similar in mice and rats to that of CCNU, with the exception that, TCNU did not cause the chronic liver disturbances found for CCNU. In dogs treated for 6 weeks with TCNU leucopenia and thrombocytopenia were the major side effects. Parent TCNU was found in all dogs. The absorption was fast, the maximum level being reach after 25 mins and the mean absorption time was 22 mins. The mean half life was 16.1 mins after intravenous and 17.4 after oral administration. The combination of these factors make TCNU an interesting clinical candidate.

Animals↗
PubMed1988

Growth inhibiting effect of estramustine on two prostatic carcinoma cell lines, LNCaP and LNCaP-r.

The effect of estramustine (EM), estradiol-17 beta (E2) or 5 alpha-dihydrotestosterone (DHT) on the growth of two human prostatic carcinoma cell lines, LNCaP and LNCaP-r was investigated. The hormone resistant subline LNCaP-r was derived in our laboratory, from the hormone sensitive LNCaP cell line. E2, 10(-8) or 10(-5) M inhibited the growth of the LNCaP cells, but did not affect the LNCaP-r. DHT, 10(-8) M, had a stabilizing effect at the stationary phase on the growth of the LNCaP cells whereas at higher concentrations, 10(-5) M, the growth rate was decreased. The LNCaP-r cell line was previously reported to be unaffected by DHT. EM inhibited the growth of both cell lines but LNCaP was more sensitive than LNCaP-r. E2 and DHT modulated the effect of EM. When treated with 10(-7) M EM, addition of E2 or DHT (10(-7)-10(-5) M) further inhibited the growth. When EM was used at a higher concentration (10(-5) M), the enhanced effect of growth inhibition by hormone addition was lost. Based on these results it is suggested that the presence of endogenous hormones, or estrogens released from EM on hydrolysis, may play a contributory role in the cytotoxicity of estramustine.

Cell Division↗
PubMed1986

Nuclear protein matrix as a target for estramustine-induced cell death.

The effect of estramustine [estradiol 3-N-bis(2-chloroethyl)carbamate] on the human prostatic tumor cell line 1013L was investigated. Cell proliferation experiments revealed that estramustine cytotoxicity varied during the different phases of cell growth. Maximum cell killing was found in early log phase, but cell death also occurred in the stationary phase. Mitotic arrest was found at cytotoxic concentrations throughout the log phase. Subcellular distribution studies showed that the cellular uptake of estramustine increased throughout the log phase and remained steady during the stationary phase. Nuclear uptake in contrast was similar in all phases, whereas a preferential binding to the nuclear protein matrix was found to increase throughout the log phase and even during the stationary phase of growth. This implicates the nuclear protein matrix as a target for estramustine cytotoxicity.

Cell Line↗
PubMed1986

Cytotoxicity and metabolism of prednimustine, chlorambucil and prednisolone in a Chinese hamster cell line.

Prednimustine and chlorambucil induce dose- and time-dependent cell death in V79 Chinese hamster cells in vitro. Prednimustine was found to be 3-4 times more potent than either chlorambucil or an equimolar mixture of its components chlorambucil and prednisolone after 24 h treatment. Prednimustine was hydrolyzed to prednisolone and chlorambucil in the system, and the concentration of prednimustine was reduced by one half within 15 h. Prednisolone was not further metabolized, but chlorambucil was rapidly inactivated by dechlorination, the half-life being 2.5 h. No dechlorinated prednimustine was formed during the experiments. The higher stability of prednimustine than chlorambucil is probably due to protective binding to different serum proteins from those that bind chlorambucil. Substitution of fetal calf serum by human serum albumin revealed that hydrolysis of prednimustine is catalyzed by esterases present in the serum. In similar substitution experiments cell survival studies indicated that prednimustine itself was not cytotoxic. Rather, cytotoxicity was found to correlate with hydrolysis to chlorambucil. Thus, it appears that the prolonged availability of chlorambucil is responsible for the increased potency of prednimustine in this system.

Animals↗
PubMed1986

The antitumor effect of prednimustine in vitro and in vivo.

Prednimustine is active against a wide variety of experimental tumors both in vivo and in vitro. In many of these tumor systems, prednimustine exhibits distinct advantages over a mixture of its constituents, chlorambucil and prednisolone. In vitro, a higher cell kill is obtained, and in vivo, at doses that are equally effective, prednimustine is less toxic. These differences could be connected with the different pharmacokinetic profiles found between prednimustine and chlorambucil plus prednisolone.

Animals↗
PubMed1985

Effect of hormones on growth and ATP content of a human prostatic carcinoma cell line, LNCaP-r.

The use of ATP content as a measurement for cell growth was evaluated in the LNCaP prostatic cancer cell line. ATP content was found to correlate well with cell counts and was an easy and reliable method for following the effect of substances on cell growth. During cultivation for 9 days no effect on cell counts or ATP content could be seen when testosterone (10(-10) to 10(-6) M), estradiol-17 beta (10(-10) to 10(-5) M), 5 alpha-DHT (10(-9) to 10(-6) M), prolactin, vitamin A, or antiandrogen was added to the cell medium in different combinations. However, a weak positive effect was seen on the mitotic index when 10 or 100 nM 5 alpha-DHT was added to the cells, whereas 1 microM 5 alpha-DHT inhibited cell growth. Thus despite the fact that this LNCaP line contained 16 fmol androgen receptor/mg protein (Kd 0.6 nM), it is unresponsive to hormones and should be designated LNCaP-r (resistant). Chromosome analysis revealed that a shift in the modal chromosome number had occurred from the original LNCaP line, which could account for the lack of hormonal sensitivity.

Adenosine Triphosphate↗