Information needs of oncologists, general practitioners and other professionals caring for patients with cancer.

Oncology is a rapidly developing field with a growing number of publications every year. The main goal of this survey was to learn more about the information needs of oncologists and general practitioners. Data were collected using a standardised questionnaire developed in collaboration with the German Cancer Society (Deutsche Krebsgesellschaft) and the German Association of General Practitioners (Deutscher Hausärzteverband). A total of 495 questionnaires could be evaluated. Medical congresses were the preferred source of information for all participants. General practitioners preferred textbooks, while oncologists preferred journals and the Internet (all p < .001). Reasons for a lack of confidence during patient consultation were lack of time (60% of participants), lack of knowledge (61% of general practitioners and 26% of oncologists) and lack of data (>50%). Oncologists felt more confident in searching scientific databases than general practitioners did. Both groups required rapid access to transparent information. For general practitioners, reviews and comments by experts helped to put new information in the context of cancer treatment. Oncologists and general practitioners showed significantly different information needs and different ways to access specific information. In order to better integrate general practitioners while simultaneously serving the needs of oncologists, a database that is up to date, rapidly accessible and does not incur high costs would be helpful.

Cellular receptor levels and glucocorticoid responsiveness of lymphoma cells.

A series of mouse lymphoma cell lines of independent origin was investigated with respect to glucocorticoid sensitivity, cellular receptor levels, and properties of receptors. The concentrations of the glucocorticoid dexamethasone required to produce comparable growth-inhibitory effects varied considerably amongst these cell lines. Also a wide range in the number of receptors per cell was found. When the receptor-steroid complexes were compared with respect to nuclear binding properties and affinities for DNA, no differences were seen. For 7 out of 10 cell lines studied we obtained a direct correlation between hormonal sensitivity and the number of cellular receptor sites divided by the dissociation constants KD for the receptor-dexamethasone complexes. This suggests that the receptor is a major quantitative determinant for steroid responsiveness. The limitations of receptor measurements for glucocorticoid therapy of lymphoid neoplastic disease are discussed.

Subunit dissociation and activation of wild-type and mutant glucocorticoid receptors.

Apparent molecular weights of wild-type and nti ('increased nuclear transfer') mutant glucocorticoid receptors were obtained from Stokes radii and sedimentation coefficients. At low salt concentrations molecular forms of Mr 328,000 and 298,000 of the wild-type and mutant, respectively, were predominant. Increasing ionic strength resulted in receptor dissociation. Dissociated forms of Mr 130,000 and 63,000 of the wild-type and mutant, respectively, were obtained at 300 mM KCl and above. Some metal oxi-anions prevented dissociation. Receptor activation to allow DNA binding produced the dissociated forms which could be separated from non-activated receptors by filtration through DNA-cellulose or by DEAE-cellulose chromatography. Non-activated wild-type and nti receptors eluted from DEAE-cellulose under identical conditions while activated wild-type and nti receptors eluted differently. Partially proteolyzed wild-type receptors behaved identically to nti receptors. We conclude that the large forms of wild-type and nti receptors are heteromeric and contain only one hormone-building polypeptide per complex.

Immortalization of a fetal rat brain cell line that expresses corticotropin-releasing factor mRNA.

By introducing the SV40 T antigen under the control of promoter sequences from the gene encoding the rat corticotropin-releasing factor (CRF) into primary cultures from fetal rat brain, a stable cell line was established that expresses the SV40 T antigen and transcribes its endogenous CRF gene. The cell line exhibits neuronal properties, as demonstrated by positive immunostaining with antibodies against neuronal markers, and it provides a useful system for the detailed analysis of the transcriptional regulation of the neuropeptide gene at an early stage of development. Promoter activities of the CRF 5'-flanking region were tested in transient expression assays after transfection of the cell line with different fusion constructs with CRF promoter regions linked to the bacterial chloramphenicol acetyltransferase (CAT) gene. Basal activity of the promoter was determined by DNA sequences between positions -269 and -222 upstream of the transcriptional start site and showed weak induction upon treatment with forskolin.

Primary structure and functional expression of a glutaminyl cyclase.

A glutaminyl cyclase (QC) that is probably involved in the biosynthesis of pyroglutamyl peptides such as gonadotropin-releasing hormone and thyrotropin-releasing hormone has been purified to homogeneity from bovine anterior pituitary. On the basis of N-terminal sequence analysis, a 2088-base-pair cDNA clone was isolated from a bovine anterior pituitary library. From the nucleotide sequence of this clone, the primary structure of a 330-residue protein and a preceding 31-residue prepropeptide sequence was deduced. By transfection of COS-7 monkey cells with a QC cDNA/pCDM8 vector construct, QC activity was expressed. Hybridization with mRNAs of various bovine tissues revealed expression of QC mainly in brain tissue.

Active domains in wild-type and mutant glucocorticoid receptors.

[3H]Triamcinolone acetonide was used to tag covalently specific glucocorticoid receptors by photoaffinity labelling at lambda greater than or equal to 320 nm. Receptors of wild-type mouse lymphoma cells and two glucocorticoid resistant mutants of "nuclear transfer deficient" (nt-) and "increased nuclear transfer" (nti) phenotypes, respectively, were used. Wild-type and nt- receptors yielded radiolabelled polypeptide bands of mol. wt. 98 000 as revealed by gel electrophoresis under denaturing conditions and fluorography. In contrast, the nti receptor had a mol. wt. of 42 000. Partial proteolysis of the wild-type receptor with alpha-chymotrypsin resulted in a fragment of mol. wt. 39 000 which still contained the steroid binding site but had increased affinity for DNA indistinguishable from that of the nti receptor. Chymotrypsin thus removed a domain from the wild-type receptor polypeptide which is involved in modulating DNA binding. The same domain is missing from the nti receptor.

Immunochemical characterization of wild-type and variant glucocorticoid receptors by monoclonal antibodies.

Monoclonal antibodies raised against the rat liver glucocorticoid receptor were used to investigate receptors of wild-type and glucocorticoid-resistant variants of mouse lymphoma cells. Two of the variant types contained receptors of 'nuclear transfer deficient' (nt-) and 'increased nuclear transfer' (nti) phenotypes, respectively, while the third was of the 'receptorless' (r-) phenotype with negligible hormone binding activity. Three monoclonal antibodies of the IgM class and one of the IgG class reacted with both wild-type and nt- receptors but not with the steroid binding form of nti receptors. Some of the antibodies bound the wild-type and nt- receptors more efficiently after activation at 20 degrees C. By use of an immuno-competition assay we were able to detect cross-reacting material in considerable amounts in extracts of nti and r- cell variants. This material was further characterized by gel filtration and immunoblotting. The immunoreactive material of wild-type, nti and r- cells gave a major band of mol. wt. 94 000 upon SDS-gel electrophoresis while the steroid-binding polypeptides of wild-type and nti receptors have mol. wts. of 94 000 and 40 000, respectively. The data show that in S49.1 mouse lymphoma cells the products of two receptor alleles can be distinguished.

Partial overlapping of binding sequences for steroid hormone receptors and DNaseI hypersensitive sites in the rabbit uteroglobin gene region.

Four DNaseI hypersensitive (HS) chromatin regions were found in the uteroglobin locus located at -3.7, -2.4, -0.1 and +4.1 kb with respect to the transcription start site of the gene. The three sites upstream of the gene are only detected in the hormonally stimulated endometrium and disappear after hormone withdrawal, whereas the site at +4.1 is also found in tissues that do not express uteroglobin. In the -2.4 HS region, which is strictly dependent on progesterone treatment, three DNaseI sites are clustered within a 240 bp DNA segment that contains 20 imperfect repeats of an octanucleotide motif. Upstream of the uteroglobin gene there are three regions containing binding sites for the glucocorticoid and the progesterone receptors, located at -3.7, -2.6/-2.7 and -2.4. The -2.4 region contains two binding sites for the hormone receptors flanking the central HS site. In footprinting experiments with naked DNA binding of the receptor also renders this site more susceptible towards digestion with DNaseI. The -2.6/-2.7 region contains three binding sites for the hormone receptors located 140 bp upstream of the HS -2.4. While the -3.7 HS is also located within a receptor binding fragment, there is no binding of the hormone receptors to the promoter region. Thus, interaction of the receptor with DNA sequences far upstream from the promoter alters the chromatin conformation of neighbouring sequences and results in transcriptional activation.