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R Apfel

Publications and source records attributed to R Apfel.

16 recordsLinked to original sources
PubMed1995

The orphan receptor TAK1 acts as a repressor of RAR-, RXR- and T3R-mediated signaling pathways.

Recently, we reported the cloning and characterization of the novel orphan receptor TAK1. In this study, we analyze the interaction of TAK1 with a variety of response elements (RE's) and demonstrate that TAK1 binds effectively to RE's composed of the core motif PuGGTCA configured in direct repeats spaced by one or more nucleotides. TAK1 bound poorly to palindromic or inverted palindromic motifs and was unable to bind to a single core motif, suggesting that a dimeric site is required for binding. Transfection experiments with CV-1 cells revealed that TAK1 is able to repress retinoid- and thyroid-hormone-induced transactivation through a subset of retinoid and thyroid hormone RE's. Our studies indicate that the antagonism of RAR-mediated transactivation does not involve the formation of heterodimers between TAK1 and RAR or RXR but is due to the competition of TAK1 homodimers with RAR-RXR heterodimers and RXR homodimers for binding to RARE and RXRE, respectively. Our results suggest that the orphan receptor TAK1 can be a negative modulator of the regulation of gene expression mediated by retinoid and thyroid hormone signaling pathways.

Animals↗
PubMed1994

Cloning of a novel malignant melanoma-derived growth-regulatory protein, MIA.

Growth and progression of malignant melanoma cells is influenced by a complex network of growth-stimulating and -inhibiting factors produced by both the tumor cells and the local environment. Here we report the purification and molecular cloning of a novel growth regulating protein, designated melanoma inhibitory activity (MIA) and provide a preliminary functional characterization. MIA is translated as a 131-amino acid precursor and processed into a mature 107-amino acid protein after cleavage of a putative secretion signal. A murine complementary DNA was isolated that encoded a MIA-protein with 88% amino acid identity. MIA is secreted into the culture supernatant by several malignant melanoma cell lines as an M(r) 11,000 autocrine growth factor and acts as a potent tumor cell growth inhibitor for malignant melanoma cells and some other neuroectodermal tumors, including gliomas. MIA has no homology to any other known protein and, therefore, represents a novel type of growth-regulatory factor. Furthermore, we describe a molecular approach to express functionally active MIA in Escherichia coli, which might be attractive as a future antitumor therapeutical substance.

Amino Acid Sequence↗
PubMed1994

A novel orphan receptor specific for a subset of thyroid hormone-responsive elements and its interaction with the retinoid/thyroid hormone receptor subfamily.

The steroid/hormone nuclear receptor superfamily comprises several subfamilies of receptors that interact with overlapping DNA sequences and/or related ligands. The thyroid/retinoid hormone receptor subfamily has recently attracted much interest because of the complex network of its receptor interactions. The retinoid X receptors (RXRs), for instance, play a very central role in this subfamily, forming heterodimers with several receptors. Here we describe a novel member of this subfamily that interacts with RXR. Using a v-erbA probe, we obtained a cDNA which encodes a novel 445-amino-acid protein, RLD-1, that contains the characteristic domains of nuclear receptors. Northern (RNA) blot analysis showed that in mature rats, the receptor is highly expressed in spleen, pituitary, lung, liver, and fat. In addition, weaker expression is observed in several other tissues. Amino acid sequence alignment and DNA-binding data revealed that the DNA-binding domain of the new receptor is related to that of the thyroid/retinoid subgroup of nuclear receptors. RLD-1 preferentially binds as a heterodimer with RXR to a direct repeat of the half-site sequence 5'-G/AGGTCA-3', separated by four nucleotides (DR-4). Surprisingly, this binding is dependent to a high degree on the nature of the spacing nucleotides. None of the known nuclear receptor ligands activated RLD-1. In contrast, a DR-4-dependent constitutive transcriptional activation of a chloramphenicol acetyltransferase reporter gene by the RLD-1/RXR alpha heterodimer was observed. Our data suggest a highly specific role for this novel receptor within the network of gene regulation by the thyroid/retinoid receptor subfamily.

Amino Acid Sequence↗
PubMed1993

Autoinduction of platelet derived growth factor (PDGF) A-chain mRNA expression in a human malignant melanoma cell line and growth inhibitory effects of PDGF-A-chain mRNA-specific antisense molecules.

Autoinduction of PDGF-A-chain mRNA's was demonstrated in cells of a human malignant melanoma cell line (HTZ 19), indicating a PDGF-induced amplification of an autocrine growth factor loop mechanism. PDGF-alpha receptors were detected by immunocytochemistry in HTZ 19 melanoma cells. Exogenously administered PDGF-AA exerted an increase in cell proliferation by 70% at 200 ng/ml PDGF in serum-supplemented cell culture medium. Antisense phosphorothioate-oligodeoxynucleotides (S-ODN's), specifically targeted against PDGF-A-chain mRNA, reduced cell proliferation of HTZ 19 melanoma cells by 62% at a concentration of 5.0 microM, also only in serum-enriched medium. These findings strongly suggest that PDGF-AA homodimers may be an autocrine growth factor for HTZ 19 melanoma cells but require synergistic interaction with serum components.

Base Sequence↗
PubMed1993

In vitro studies on interaction of 4-hydroperoxyifosfamide and 2-mercaptoethanesulphonate in malignant gliomas.

Drug interference of ifosfamide and sodium 2-mercaptoethanesulphonate (MESNA) was studied in three malignant glioma cell cultures (HTZ-17, HTZ-209B, and HTZ-243) by a recently developed in vitro method for evaluation of multimodal treatment interactions. Glioma cell cultures were treated in monolayer 96-well tissue-culture plates for 2 h each, with 4-hydroperoxyifosfamide and MESNA combined in both sequences, or alone. Concentrations ranged from 0.01 microM to 50 microM in single-modality exposures, and from 0.01 microM to 10 microM in combination exposures. After five population doubling times, DNA synthesis was determined by a standard [3H]Tdr-incorporation liquid-scintillation-counting protocol. Data points were evaluated for mono- and combined treatment dose effects (adapted with a probit function), and a model-free three-dimensional response surface was created that was compared to the theoretical additive, anticipated response surface. Local additivity was analysed for any ratio of combined treatment. No tumour effects were seen with MESNA in single-drug exposure, whereas ifosfamide resulted in more than 90% inhibition of tumour DNA synthesis. In combination experiments, MESNA could be confirmed to be inert: the anticipated theoretical combination response surfaces formed a three-dimensional extension of the single-drug ifosfamide dose/response curves--the experimental combination response surfaces displayed an identical appearance (P < or = 0.05). In conclusion, these results indicate no drug interference of MESNA and ifosfamide in malignant glioma cells.

Antineoplastic Agents↗
PubMed1993

Effects of diethylstilbestrol (DES) medication during pregnancy: report from a symposium at the 10th international congress of ISPOG.

This article is a report based on presentations at the symposium Effects of Diethylstilbestrol (DES) Medication during Pregnancy at the conference Reproductive Life, 10th International Congress of The International Society of Psychosomatic Obstetrics and Gynaecology (ISPOG) in Stockholm, Sweden June 14-17, 1992. The objective of this symposium, chaired by Eylard van Hall and Ingar Palmlund, was to provide the basis for a discussion of how the risks of DES had been evaluated in different countries. In the general discussion following the presentations everybody present expressed concerns that DES might still be given to pregnant women in many parts of the world, called for measures to alert medical professionals world-wide to the hazards of the use of DES, and requested measures to effect an international ban of DES in medication for pregnant women.

Abnormalities, Drug-Induced↗
PubMed1993

The effect of transforming growth factor-beta 2-specific phosphorothioate-anti-sense oligodeoxynucleotides in reversing cellular immunosuppression in malignant glioma.

This in vitro study was aimed at restitution of transforming growth factor (TGF)-beta 2-mediated suppression of T-lymphocyte activation within malignant gliomas. In early-passage tumor cell cultures of two glioblastomas (HTZ-153 and HTZ-209) and one malignant astrocytoma classified as World Health Organization Grade III (HTZ-243), autologous peripheral blood mononuclear cells were activated by interleukin-1 alpha and interleukin-2 in vitro (lymphokine-activated killer cells) and tested for cytotoxic and proliferative activity. In expression studies (Western blot and Northern hybridization) of all three tumors, TGF-beta could be detected at the protein and messenger ribonucleic acid (mRNA) levels. A polyclonal anti-TGF-beta neutralizing antibody did not enhance lymphocyte proliferation upon stimulation with tumor targets (3H-thymidine incorporation) and slightly stimulated lymphocyte cytotoxicity against autologous target cells. Preincubation of target cells for 12 hours with TGF-beta 2-specific phosphorothioate-anti-sense oligodeoxynucleotides (S-ODN's) did, however, enhance lymphocyte proliferation up to 2.5-fold and autologous tumor cytotoxicity up to 60%, compared to controls not treated with S-ODN's. Incubation of tumor cells with TGF-beta 2-specific S-ODN's resulted in decreased TGF-beta-specific immunoreactivity in cultured glioma cells, in reduced TGF-beta 2 protein concentration (Western blot), and in a change in the expression pattern of TGF-beta 2 mRNA's. These observations may have implications for in vivo and in vitro activation of a cellular immune response against autologous malignant glioma cells.

Antigens, CD↗
PubMed1992

Purification and analysis of growth regulating proteins secreted by a human melanoma cell line.

Supernatants of a human malignant cell line established from a CNS metastasis, contained several proteins with putative growth regulating functions. BioGel P-10 gel filtration chromatography, reverse phase HPLC purification, and amino-terminal sequencing of purified peptides resulted in characterization of beta 2-microglobulin (beta 2M, 10 kD), ubiquitin (6 kD), and tissue inhibitor of metalloproteinases 2 (TIMP-2, 21 kD). In addition, CNBr cleavage and purification of resulting peptides revealed diazepam binding inhibitor (DBI, 8 kD) and melanoma inhibiting activity (MIA, 11 kD). The secretion of beta 2M as part of the HLA-class I complex may be related to impaired autologous anti-tumour immune function; ubiquitin may play a role in activation or deactivation of extracellular proteins or cell-cell interactions. As HTZ-19 cells respond in a dose-dependent manner to midozolam, DBI may interfere with growth regulation mediated by diazepam receptor sites. In a collagenolytic assay, TIMP-2 interfered with metalloproteinase functions, which are required for degradation of collagen type IV and organotopic metastasis. MIA is clearly associated with a proliferation inhibiting effect on HTZ-19 cells. In conclusion, although this tumour shows a degree of progression, several proteins with putative functions at different cellular levels were identified, related to proliferation as well as to the type of metastasis.

Amino Acid Sequence↗
PubMed1991

In vitro studies on drug interaction of ifosfamide and ACNU in primary and metastatic human brain tumours.

Combination chemotherapy is widely employed in clinical oncology; however, there is no generally accepted model to evaluate individual tumour susceptibility to a given drug combination protocol. We therefore investigated the drug interaction of ifosfamide (4-hydroxyperoxy-ifosfamide) and ACNU in a recently developed in vitro model of paired sequential combination chemotherapy in primary and metastatic malignant brain tumours. A long-term standard [6,3-3H]-thymidine-incorporation assay, employing a liquid scintillation counting protocol, was selected to assess the drug sensitivity of human tumours. In vitro drug exposures were derived from correlating in vivo-(systemic and CNS) and in vitro-pharmacokinetic drug parameters. In combination experiments tumour cells were treated sequentially by two drugs in both sequences: drug exposures were calculated for 2 h with a 1-h drug-free interval in between. "Cut-off" concentrations (maximum in vitro exposure doses) were calculated as 1.74 microM (for primary CNS tumours: 0.58 microM) for ifosfamide and 5.4 microM (for primary CNS tumours: 1.33 microM) for ACNU. Dose/response relations were derived from isotope incorporation rates after cells had grown for approximately five population doubling times. Combination isoboles were plotted after drug doses had been transformed into "equieffective doses", enabling comparison of drug combination effects. In all three glioblastomas (with CNS exposure dose) an additive or supra-additive effect could be observed in either sequence (in one tumour a biphasic additive isobole was found for both sequences). Out of three bronchial carcinomas (small-cell type, brain metastases) in two non-identical sequences a supra-additive effect was observed in two tumours, with antagonistic effects in the third tumour. In all three malignant melanomas and in one renal carcinoma antagonistic effects were observed, whereas in a second renal carcinoma supra-additive effects were demonstrated for both sequences. We conclude that drug combination chemotherapy effects at the cellular level may be extremely heterogeneous.

Antineoplastic Combined Chemotherapy Protocols↗
PubMed1990

Melanoma-inhibiting activity inhibits cell proliferation by prolongation of the S-phase and arrest of cells in the G2 compartment.

Autocrine-secreted melanoma tumor growth-inhibiting activity (MIA, approximately Mr 8000) was isolated from supernatants of a malignant melanoma cell line HTZ-19 dM, established from a central nervous system-melanoma metastasis. Cell cycle kinetic analysis performed with bromodeoxyuridine/Hoechst flow cytometry revealed a MIA-sensitive period at the G0/G1 to S traverse; MIA mediated prolongation of the S-phase and increased arrest of cells in the G2 compartment. Growth inhibition by MIA is cell-density dependent; maximal effect is seen at low densities, and the effect may be partially antagonized by whole serum. MIA may cause growth stimulation at high cell densities and low MIA concentrations. The effect of MIA on different histological neuroectodermal cell types was compared by the same methodology: proliferation of a second malignant melanoma was inhibited, and no effect was observed with an ependymoma; 2 glioblastomas were slightly stimulated. Effects on human fibroblast-like cell strains were inconsistent. The mechanism of MIA is discussed in relation to other endogenous autocrine growth inhibitors.

Cell Count↗
PubMed1989

Autocrine tumor cell growth-inhibiting activities from human malignant melanoma.

Autocrine-secreted tumor cell growth-inhibiting activities were isolated from supernatants of a malignant melanoma cell line, HTZ 19-dM, established from a central nervous system melanoma metastasis. HTZ 19-dM was characterized by cyto- and immunocytochemistry and karyotyping; cells were propagated in defined serum-free tissue culture medium for up to 8 months. Supernatants were ultrafiltrated, dialyzed, lyophilized, and purified by Bio-Gel P-10 gel permeation chromatography, leading to three active fraction pools, MIAI [melanoma-inhibiting activity (MIA), 2 kDa), MIAII (Mr 11,500-17,000) and MIAIII (proteins at the cutoff of Bio-Gel P-10) inhibiting growth of 19-dM cells with 50% inhibitory concentrations of 0.79 microgram/ml (MIAI), 0.13 microgram/ml (MIAII), and 16.7 micrograms/ml (MIAII). MIAII could be further purified by reverse phase high pressure liquid chromatography; the main activity displayed a 50% inhibitory concentration of 0.33 microgram/ml. On sodium dodecyl sulfate-polyacrylamide gel electrophoresis one major band (molecular weight about 14,000) and two minor bands (up to Mr 17,000) were identified. Macromolecular synthesis was inhibited in 19-dM cells up to greater than 99.5%; tumor stem cell colony formation was reduced by 99.89%; the inhibitory effect of MIAII was irreversible, nonsaturable, and partially antagonized by a serum factor (depending on purification stage). MIAII was heat stable (3 min at 100 degrees C) and trypsin labile. The effect of MIAII on allogeneic neuroectodermal tumors was also investigated; proliferation of two of three malignant melanomas and two of four glioblastomas was inhibited up to 85.2%; proliferation of a neuroblastoma cell line could be inhibited to 33.8%, whereas normal fibroblasts and low grade gliomas were not influenced in their proliferation.

Biological Factors↗
PubMed1986

Pregnancy during residency.

To identify possible barriers to women physicians who wish to combine parenting with medical careers, we studied 56 of 64 pregnancies from the past 10 years in 63 of 66 Harvard-affiliated residency programs. Pregnancies during residency were common (one in eight married women in 1983) and were usually planned (77 percent). Most institutions were unprepared for pregnancies among members of the house staff; four fifths of the programs had no maternity-leave policy. No one quit a residency program because of pregnancy, and pregnancy rarely affected achievement of board certification. Whether the pregnant women found pregnancy during residency a "pleasant" experience was determined largely by whether the program director was supportive, whether the issue of pregnancy was openly discussed within the program, and whether the woman was permitted to return to work on a part-time basis for the first weeks after maternity leave. We conclude that pregnancy within residency programs should be expected and planned for and that if proper arrangements are made, it need not be a major problem for either the training program or the pregnant resident.

Attitude of Health Personnel↗
PubMed1978

Treatment of anxiety: a comparison of the usefulness of self-hypnosis and a meditational relaxation technique. An overview.

We have investigated prospectively the efficacy of two nonpharmacologic relaxation techniques in the therapy of anxiety. A simple, meditational relaxation technique (MT) that elicits the changes of decreased sympathetic nervous system activity was compared to a self-hypnosis technique (HT) in which relaxation, with or without altered perceptions, was suggested. 32 patients with anxiety neurosis were divided into 2 groups on the basis of their responsivity to hypnosis: moderate-high and low responsivity. The MT or HT was then randomly assigned separately to each member of the two responsivity groups. Thus, 4 treatment groups were studied: moderate-high responsivity MT; low responsivity MT; moderate-high responsivity HT; and low responsivity HT. The low responsivity HT group, by definition largely incapable of achieving the altered perceptions essential to hypnosis, was designed as the control group. Patients were instructed to practice the assigned technique daily for 8 weeks. Change in anxiety was determined by three types of evaluation: psychiatric assessment; physiologic testing; and self-assessment. There was essentially no difference between the two techniques in therapeutic efficacy according to these evaluations. Psychiatric assessment revealed overall improvement in 34% of the patients and the self-rating assessment indicated improvement in 63% of the population. Patients who had moderate-high hypnotic responsivity, independent of the technique used, significantly improved on psychiatric assessment (p = 0.05) and decreased average systolic blood pressure from 126.1 to 122.5 mm Hg over the 8-week period (p = 0.048). The responsivity scores at the higher end of the hypnotic responsivity spectrum were proportionately correlated to greater decreases in systolic blood pressure (p = 0.075) and to improvement by psychiatric assessment (p = 0.003). There was, however, no consistent relation between hypnotic responsivity and the other assessments made, such as diastolic blood pressure, oxygen consumption, heart rate and the self-rating questionnaires. The meditational and self-hypnosis techniques employed in this investigation are simple to use and effective in the therapy of anxiety.

Anxiety↗
PubMedDate not supplied

MIA, a novel serum marker for progression of malignant melanoma.

MIA was isolated previously as a small soluble protein secreted from malignant melanoma cell lines in vitro. Highly restricted expression patterns in melanocytic tumors were identified in vivo. We therefore quantitated serum levels of MIA protein by means of a non-radioactive ELISA and investigated whether MIA provides clinically relevant parameters in patients with malignant melanomas. Here we report enhanced MIA serum levels in 13% and 23% of patients with stage I and II disease, respectively, and in 100% with stage III or IV disease. Response to therapy in stage IV disease correlated with changes in MIA serum levels and surgical removal of metastases led to normalization of serum values. Repeated measuring of sera from 350 patients with a history of stage I or II melanoma during follow-up, we detected 32 patients developing positive MIA values. At the time of serum analysis 15 of them had developed metastases and one presented with metastatic disease 6 months later. In conclusion, MIA represents a novel serum marker for systemic malignant melanoma revealing the highest sensitivity and specificity among currently available markers.

Adult↗
PubMedDate not supplied

In vitro studies on interaction of 4-hydroperoxy-ifosfamide and radiotherapy in malignant gliomas.

Drug-radiation multimodal chemotherapy for malignant brain tumor cells was stimulated in vitro by a recently developed computer program, based on evaluation of 3D-response surfaces and interaction isoboles. Three malignant glioma tumor cell cultures (HTZ17, 146 and 209 B) were sequentially treated in vitro by increasing doses of 4-hydroperoxy-ifosfamide, according to in vivo/in vitro pharmacokinetic correlation (0.01 to 10 microM), and increasing single doses of Gamma-radiation (clinical60Co-radiotherapy unit, 0.26 to 4 Gy or 1 to 3.38 Gy, respectively). After approximately five population doubling times with standard tissue culture conditions, 3H-Tdr-incorporation was determined by a liquid scintillation counting protocol. Data points were evaluated for mono- and combined treatment dose-effects. A model-free 3D-response surface was created and compared to the theoretical additive response surface. Local additivity was analysed for any desired ratio of combined treatment as well as for isoboles. No significant sub- or supraadditive effects were observed, indicating additive effects in all 3 tumors. No sequence dependence of effects could be demonstrated. In case radiotherapy and ifosfamide-chemotherapy are active treatment modalities and additive effects are found, we conclude that the combination of ifosfamide and radiotherapy might be attractive for the treatment of malignant brain tumors and should be further studied.

Astrocytoma↗