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E Weiss

Publications and source records attributed to E Weiss.

At least 37 records · Page 2Linked to original sources
PubMed2000

HPV oncoprotein E6 is a structure-dependent DNA-binding protein that recognizes four-way junctions.

E6 is an oncoprotein implicated in cervical cancers, produced by "high-risk" human papillomaviruses. E6 is thought to promote tumorigenesis by stimulating cellular degradation of the tumour suppressor p53, but it might display other activities. Sequence similarity was recently detected between E6 and endonuclease VII, a protein of phage T4 that recognizes and cleaves four-way DNA junctions. Here, we purified recombinant E6 proteins and demonstrated that high-risk E6 s bind selectively to four-way junctions in a structure-dependent manner. Several residues in the C-terminal zinc-binding domain, the region of E6 similar to endonuclease VII, are necessary for the junction-binding activity. E6 binds to the junction as a monomer. Comparative electrophoresis shows that E6-bound junctions migrate in an extended square conformation. Magnesium inhibits the electrophoretic migration of the complexes but does not seem to influence their formation at equilibrium. This work is the first demonstration of specific binding of purified active E6 to a well-characterized DNA ligand, and suggests new modes of action of E6 in oncogenesis.

Amino Acid Sequence↗
PubMed2000

Production of fluorescent single-chain antibody fragments in Escherichia coli.

We describe a novel vector-host system suitable for the efficient preparation of fluorescent single-chain antibody Fv fragments (scFv) in Escherichia coli. The previously described pscFv1F4 vector used for the bacterial expression of functional scFv to the E6 protein of human papillomavirus type 16 was modified by appending to its C-terminus the green fluorescent protein (GFP). The expression of the scFv1F4-GFP fusion proteins was monitored by analyzing of the typical GFP fluorescence of the transformed cells under UV illumination. The brightest signal was obtained when scFv1F4 was linked to the cycle 3 GFP variant (GFPuv) and expressed in the cytoplasm of AD494(DE3) bacteria under control of the arabinose promoter. Although the scFv1F4 expressed under these conditions did not contain disulfide bridges, about 1% of the molecules were able to bind antigen. Fluorescence analysis of antigen-coated agarose beads incubated with the cytoplasmic scFv-GFP complexes showed that a similar proportion of fusions retained both E6-binding and green-light-emitting activities. The scFv1F4-GFPuv molecules were purified by affinity chromatography and successfully used to detect viral E6 protein in transfected COS cells by fluorescence microscopy. When an anti-beta-galactosidase scFv, which had previously been adapted to cytoplasmic expression at high levels, was used in this system, it was possible to produce large amounts of functional fluorescent antibody fragments. This indicates that these labeled scFvs may have many applications in fluorescence-based single-step immunoassays.

Animals↗
PubMed2000

Rhabdoid tumors of the central nervous system.

Rhabdoid tumors of the central nervous system are rare malignancies with a still almost uniformly fatal outcome. There is still no proven curative therapy available. We report our experience with nine patients with central nervous system rhabdoid tumors. Gross complete surgical removal of the tumor was achieved in six patients. Seven patients received intensive chemotherapy. Four of these were treated in addition with both neuroaxis radiotherapy and a local boost directed to the tumor region, while two patients received local radiotherapy only. The therapy was reasonably well tolerated in most cases. Despite the aggressive therapy, eight of the nine patients died from progressive tumor disease, and one patient died from hemorrhagic brain stem lesions of unknown etiology. The mean survival time was 10 months after diagnosis. Conventional treatment, although aggressive, cannot change the fatal prognosis of central nervous system rhabdoid tumors. As these neoplasms are so rare, a coordinated register would probably be a good idea, offering a means of learning more about the tumor's biology and possible strategies of treatment.

Adolescent↗
PubMed2000

Do prior abdominal operations alter the outcome of laparoscopically assisted right hemicolectomy?

BACKGROUND: Adhesions can increase the difficulty of both laparoscopic surgery and laparotomy. The aim of this study was to compare the results of laparoscopically assisted right hemicolectomy in patients after prior abdominal operations (PAOs) with the results in patients without prior abdominal operations (NPAOs). METHODS: Between August 1991 and September 1998, 85 patients underwent laparoscopically assisted right hemicolectomy. The Mann-Whitney test or Fisher's exact test was used for statistical analysis. RESULTS: In this study, 36 patients (21 women and 15 men), with an average age of 57.5 years (range, 15-87 years) had undergone a mean of 1.25 (range, 1 to 3) PAOs, whereas 49 patients (20 women and 29 men), with an average age of 60.0 years (range, 16 to 87 years) (p = 0.44) had undergone NPAOs. Overall in the PAO and NPAO groups, respectively, there were no significant differences in the incidence of intraoperative complications (3 versus 4; p = 1.0). The mean operative time was 151 min (range, 90 to 260 min) versus 148 min (range, 70 to 270 min) (p = 0.66), and the mean length of stay was 6. 8 days (range, 3 to 18 days) versus 7.6 days (range, 3 to 19 days) (p = 0.13). The procedure was converted to laparotomy (p = 0.754) for six patients in the PAO group (5 because of adhesions) and 6 patients in the NPAO-group (1 because of adhesions; p = 0.078). In the PAO group 17 patients (47%) had 22 postoperative complications: 11 general medical and 4 wound-related complications. Seven patients (19%) had prolonged postoperative ileus. In the NPAO-group 18 patients (38%) had a total of 22 complications: 7 general medical problems, 6 wound-related complications, and 8 prolonged postoperative ileus, none of which were statistically significant. One patient in the NPAO group had an anastomotic leak. During a mean follow-up period of 41 months (range, 3 to 89 months), three patients in the PAO group and four in the NPAO group developed incisional hernias. CONCLUSIONS: Although there is a trend toward more conversions because of adhesions in patients with a history of prior abdominal operations (p = 0.078), no increase in morbidity resulted. Therefore, laparoscopically assisted right hemicolectomy can be offered to patients with PAO, whose rate of adhesions can be expected to equal that of patients with NPAO.

Abdomen↗
PubMed2000

The use of rectal ultrasound for the correct diagnosis and treatment of rectal villous tumors.

BACKGROUND: One of the difficulties associated with surgery for rectal villous tumors (RVT) is the finding of invasive adenocarcinoma after transanal excision (TAE) and the possible need for more radical procedures or adjuvant therapy. Improved preoperative evaluation may eliminate this dilemma. The aim of our study was to evaluate the role of transrectal ultrasound (TRUS) in establishing the correct diagnosis of RVT. METHODS: All patients with biopsy proven RVT, who were referred for TAE, underwent preoperative TRUS in addition to the routine evaluation. If invasion beyond the submucosa was suspected by TRUS, multiple biopsies were taken before any surgical intervention in order to exclude invasive cancer. If no invasion was noted, biopsies were avoided and a TAE was performed. The final pathology results were compared with both the preoperative diagnosis and TRUS results. RESULTS: Thirty-five patients (19 female, 16 male; mean age 67.5 years, range 36 to 88) were studied. The mean distance of the distal extent of the lesion above the anal verge was 5.8 cm (1.5 to 6). In 27 patients, the tumor was limited to the submucosa (uT0, uT1) on TRUS and, therefore, TAE was performed. In 26 of 27 patients (96%), pathology examination confirmed the presence of RVT without evidence of malignancy. One patient was found to have invasion of the muscularis propria and required postoperative radiation therapy. In 8 patients (23%), TRUS showed extension beyond the submucosa; 3 of these patients had uT2 lesions, 4 had uT3 tumors, and 1 had perirectal nodes. These 8 patients underwent repeated biopsies with the finding of invasive adenocarcinoma in 7. Two patients underwent abdominoperineal resection, 3 had a low anterior resection, and 3 had a TAE. Final pathology confirmed the preoperative diagnosis of invasive adenocarcinoma in 7 patients. In the 1 patient with a uT2 lesion and negative biopsies, the final diagnosis was RVT with no evidence of malignancy. CONCLUSIONS: Preoperative TRUS provides an accurate diagnosis of RVT. In conjunction with TRUS-directed biopsies, directed management of these tumors could be achieved.

Adenoma, Villous↗
PubMed2000

Synthesis and antimalarial activities of base-catalyzed adducts of 11-azaartemisinin.

A series of N-substituted 11-azaartemisinins were prepared in high yield employing base-catalyzed additions to an amide nitrogen of olefins and terminal acetylenes conjugated with electron withdrawing groups (EWGs). When the terminal acetylene was conjugated with carbomethoxy, N,N-dimethyl amide or carbonyl groups, the E-adducts resulted. A mixture of E- and Z-adducts were obtained when the EWG was a nitrile. In vitro antimalarial activities of each compound were determined against two drug-resistant strains of Plasmodium falciparum. Many of the compounds prepared were several times more active than artemisinin.

Animals↗
PubMed2000

Bypassing hybridoma technology: HLA-C reactive human single-chain antibody fragments (scFv) derived from a synthetic phage display library (HuCAL) and their potential to discriminate HLA class I specificities.

The generation of discriminative, monospecific anti-HLA antibodies used to be a difficult endeavor. Phage display technology, using single-chain antibody fragments (scFv) offers a powerful alternative obtaining target-specific, genetically stable reagents. Most of scFv obtained to date have been enriched by panning phage libraries to solid-phase coupled antigens. In the present study, HLA-C-specific scFv were isolated using a synthetic phage library in combination with a Cw*0602 overexpressing cell line. ScFv from this procedure precipitated HLA-Cw*0602 heavy chains from whole cell lysates. Flow cytometry analysis revealed that scFv stained HLA-Cw*0602-positive cells, but not cells expressing HLA alleles Cw*0302, Cw*0802, A*0201, B*2705, or Gm1*01011, indicating the specificity of scFv. Similarly they showed an ability to discriminate Cw*0602-positive from Cw*0602-negative peripheral blood lymphocytes (PBL). The results of our study demonstrate the feasibility to genetically engineer single-chain HLA-class I-specific antibodies, by phage display technology. This approach might be a valuable tool to develop a broad range of novel monospecific antibodies against HLA-class I specificities.

Alleles↗
PubMed2000

Off-label use of antipsychotic drugs.

Despite the fact that most antipsychotics have only been formally evaluated for the treatment of schizophreniform disorder, schizophrenia, mania, and schizoaffective disorder (defined as "classical indications"), antipsychotics are widely used for the treatment of a broad range of symptoms and disorders. In this study, 173 patients who were having their prescriptions for antipsychotics filled at local pharmacies were interviewed. In 115 patients (66.5%), an antipsychotic was prescribed for off-label indications. Patients most often stated that they took antipsychotics as a tranquilizer or an anxiolytic. Neither gender, education, duration of treatment, nor efficacy of treatment showed an influence on the prescription practices for antipsychotics. In contrast, family status and side effects showed a significant influence. A classical indication was more often found in married and widowed patients than in unmarried or divorced ones. Patients in whom antipsychotics were prescribed for the treatment of schizophrenia, schizophreniform disorder, mania, or schizoaffective disorder experienced side effects more often than others. Age was also important for the indication of antipsychotics. Classical indications of antipsychotics were most often found in patients aged 30 to 49 years. In older patients (49-70 years), antipsychotics were almost exclusively used for off-label indications. In classical indications, clozapine was used more frequently (50%) than other antipsychotics. Melperone was primarily prescribed for off-label use.

Adolescent↗
PubMed2000

Effect of various mixing ratios on antibacterial properties and hardness of endodontic sealers.

Most chemically cured two-component dental materials, including endodontic sealers, are marketed with mixing instructions but with no strict mixing ratios. The present study evaluated the antibacterial properties and hardness of three endodontic sealers: Roth's cement (RC), CRCS, and AH26, mixed to four controlled consistencies within the range of the manufacturer recommendations. Using Enterococcus faecalis as the test microorganism, antibacterial activity was evaluated by agar diffusion and direct contact test. Surface hardness of sealers with the same consistency was evaluated on week-old specimens by the Knoop Hardness Number tester. In the agar diffusion test, light consistency of RC showed larger zones of inhibition than heavier consistency, whereas no significant differences were found with AH26 or CRCS. In the direct contact test, RC and CRCS exhibited complete inhibition of bacterial growth at all consistencies, whereas AH26 with the heavier consistencies did not inhibit bacterial growth at 24 h samples. The hardness of AH26 and CRCS was significantly lower with lighter consistencies. It is concluded that endodontic sealers possess different antibacterial and physical properties according to their mixing consistencies.

Anti-Infective Agents, Local↗
PubMed1999

In vivo biotinylated recombinant antibodies: high efficiency of labelling and application to the cloning of active anti-human IgG1 Fab fragments.

In vivo biotinylation of antibody fragments with a gene fusion approach is a realistic alternative to conventional in vitro chemical labelling. We have previously reported the construction of a vector system suitable for the bacterial expression of the binding fragment of antibody (Fab) genetically linked to the C-terminal domain of Escherichia Coli biotin carboxy carrier protein (BCCP*). A minor fraction of the expressed hybrids was biotinylated in vivo and therefore able to interact with streptavidin. We now show that the large majority of bacterially-expressed Fab-BCCP* fusions are labelled with biotin when plasmid-encoded biotin holoenzyme synthetase (BirA) is co-expressed. The yield of biotinylated Fab is maximal when overexpression of BirA is driven by a second compatible plasmid. We took advantage of this property to develop a novel filter assay for the rapid identification of recombinant Fab reacting with immunoglobulin. Starting with total RNA of two newly established murine hybridoma cell lines producing anti-human IgG1 antibodies, we selected in a single experiment the bacterial clones that expressed in vivo biotinylated anti-IgG1 Fab. Sequence analysis of the isolated Fabs showed that they did not derive from a single B clone. In addition, we found that these recombinant Fabs labelled with biotin in vivo are useful for the specific detection of human IgG1 by a solid-phase immunoassay.

Acetyl-CoA Carboxylase↗
PubMed1999

Mgm101p is a novel component of the mitochondrial nucleoid that binds DNA and is required for the repair of oxidatively damaged mitochondrial DNA.

Maintenance of mitochondrial DNA (mtDNA) during cell division is required for progeny to be respiratory competent. Maintenance involves the replication, repair, assembly, segregation, and partitioning of the mitochondrial nucleoid. MGM101 has been identified as a gene essential for mtDNA maintenance in S. cerevisiae, but its role is unknown. Using liquid chromatography coupled with tandem mass spectrometry, we identified Mgm101p as a component of highly enriched nucleoids, suggesting that it plays a nucleoid-specific role in maintenance. Subcellular fractionation, indirect immunofluorescence and GFP tagging show that Mgm101p is exclusively associated with the mitochondrial nucleoid structure in cells. Furthermore, DNA affinity chromatography of nucleoid extracts indicates that Mgm101p binds to DNA, suggesting that its nucleoid localization is in part due to this activity. Phenotypic analysis of cells containing a temperature sensitive mgm101 allele suggests that Mgm101p is not involved in mtDNA packaging, segregation, partitioning or required for ongoing mtDNA replication. We examined Mgm101p's role in mtDNA repair. As compared with wild-type cells, mgm101 cells were more sensitive to mtDNA damage induced by UV irradiation and were hypersensitive to mtDNA damage induced by gamma rays and H2O2 treatment. Thus, we propose that Mgm101p performs an essential function in the repair of oxidatively damaged mtDNA that is required for the maintenance of the mitochondrial genome.

Alleles↗
PubMed1999

Pathophysiologic basis of contrast enhancement in breast tumors.

While the diagnostic benefits of gadolinium (Gd)-chelate contrast agents are firmly established in magnetic resonance imaging (MRI) of tumors, the pathophysiologic basis of the enhancement observed and its histopathologic correlate remained vague. Tumor angiogenesis is fundamental for growth and metastasis and also of interest in new therapeutic concepts. By correlative analysis of a) histology; b) vascular density (CD31); and c) vascular permeability (vascular permeability factor/vascular endothelial growth factor [VPF/VEGF]), we found a) significantly (P < 0.001) faster exchange rates in malignant compared with benign breast lesions; b) distinct differences in enhancement characteristics between the histologic types (invasive ductal carcinoma, invasive lobular carcinoma, and ductal carcinoma in situ); and c) dependence of enhancement kinetics on the VPF/VEGF expression. The pathophysiologic basis for the differences in contrast enhancement patterns of tumors detectable by MRI is mainly due to vascular permeability, which leads to more characteristic differences than vascular density. MRI is able to subclassify malignant breast tumors due to their different angiogenetic properties.

Adult↗
PubMed1999

Therapeutic outcome and relation of acute and late side effects in the adjuvant radiotherapy of endometrial carcinoma stage I and II.

BACKGROUND AND PURPOSE: Adjuvant radiotherapy is the standard treatment for endometrial carcinoma. However, until now there has been no clear indication that radiotherapy in that setting improves survival. This calls for critical assessment of the relation between the expected benefit and the number and severity of postradiotherapeutic side effects. MATERIAL AND METHODS: In a retrospective study 159 consecutive patients with stage I and II endometrial carcinoma were treated with external radiotherapy and vaginal brachytherapy after hysterectomy and bilateral salpingoophorectomy (no surgical lymph node staging). Recurrence rates, survival and side effects were evaluated, in particular with regard to the relation of acute and late toxicity. RESULTS: Five percent of all patients developed a recurrence either in the abdomen or at distant locations. No pelvic relapses were diagnosed. Five-year relapse-free survival was 92% for stage IA + B, 84% for stage IC, and 79% for stage II disease. Acute toxicity was seen in 65.4% of patients, all grade 1 or 2, late toxicity grade 1 or 2 in 18.8%, grade 3 or 4 complications in 1.8%. The estimated 5-year freedom from late toxicity was 76.8%. Seventy-seven percent of all patients with treatment interruptions or premature end of therapy experienced late injury. In ten of 33 patients with late sequelae there was no free interval between early and late toxicity. No relation of radiation technique/fractionation and the rate of side effects was noticed. CONCLUSIONS: Adjuvant radiotherapy leads to excellent pelvic control, with few serious complications. Major acute toxicity shows significant correlation with the incidence of severe late injury, which we suggest to classify in part as consequential late effects.

Adult↗
PubMed1999

PE-11, a peptide derived from chromogranin B, in the human brain.

This study was performed to investigate the distribution of chromogranin B in the human central nervous system. We used an antiserum raised against a synthetic peptide (PE-11) present in the chromogranin B molecule. PE-11-like immunoreactivity was characterized by molecular size exclusion and reversed-phase high-performance liquid chromatography. Its localization was studied using immunocytochemistry. Only the free peptide and an N-terminally elongated peptide were detected by molecular size exclusion high-performance liquid chromatography, indicating that proteolytic processing of chromogranin B is quite extensive. PE-11-like immunoreactivity was present in differently shaped fibers, varicosities and neurons, but not in glial cells. Its density varied throughout the brain. An especially high density was observed in the bed nucleus of the stria terminalis, the central and cortical nuclei of the amygdala, the hypothalamus, the hippocampus, the raphe complex, the nucleus interpeduncularis, the nucleus of the solitary tract, and laminae I and II of the spinal cord. This study demonstrates a significant processing of chromogranin B and indicates that chromogranin B constitutes a precursor for smaller peptides which are derived by endoproteolytic processing. It provides the neuroanatomical basis to investigate the chromogranin B molecule as a widespread component of large dense-core vesicles in the human central nervous system.

Adult↗
PubMed1999

Detection of soluble HLA-G molecules in plasma and amniotic fluid.

Although the cDNA sequence of HLA-G antigens is compatible with their expression as soluble molecules (sHLA-G), the determination of native sHLA-G levels in body fluids has not yet been described. The lack of this information is likely to reflect the difficulties in developing an assay suitable to measure sHLA-G antigens in the presence of soluble HLA-A, -B and -C (sHLA-I) antigens, since most of the available anti-HLA-G mAb do not detect soluble beta2-m associated HLA-G antigens or crossreact with sHLA-I antigens. Therefore, we have developed a two-step assay which eliminates the interference of classical HLA class I antigens. In the first step, the sample is depleted of sHLA-I antigens and of HLA-E antigens with mAb TP25.99. Then, HLA-G antigens are captured with mAb W6/32 and detected with anti-beta2-m mAb in ELISA. Utilizing this assay, sHLA-G antigen levels were measured in EDTA plasma from 92 controls with known HLA types, 28 women at delivery and the corresponding cord bloods and in 50 amniotic fluids. Mean sHLA-G plasma levels did not differ between males (24.9+/-3.0 SEM ng/ml; n=42) and females (20.1+/-2.1 SEM ng/ml; n = 50). However, sHLA-G levels in HLA-A11 positive probands (mean: 13.0+/-4.4 SEM ng/ml; n=12) were significantly (P<0.05) lower than in HLA-A11 negative ones (mean: 24.5+/-2.0 SEM ng/ml; n=80). sHLA-G levels in women at delivery (mean: 22.9+/-2.2 SEM ng/ml; n=28) were in the range of controls but were significantly (P<0.001) reduced in the corresponding cord bloods (mean: 13.8+/-1.5 SEM ng/ml; n=28). sHLA-G levels in amniotic fluids (mean: 15.5 + 1.0 SEM ng/ml; n=50) were significantly (P<0.001) lower than in plasma. sHLA-G levels were 5 and 11% of those of sHLA-I antigens in plasmas and amniotic fluids, respectively. Individual sHLA-G levels were not correlated with sHLA-I levels. SDS-PAGE analysis of plasma sHLA-G antigens revealed two molecular variants with a 35 kD and a 27 kD MW corresponding to the sizes of sHLA-G1 and -G2 isoforms. In conclusion, our study has shown that the two-step assay we have developed is reliable in measuring sHLA-G antigen levels. This assay will facilitate the analysis of the biological and clinical significance of sHLA-G antigens in plasma.

Adult↗