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Biomedical subjects

R J Scheper

Publications and source records attributed to R J Scheper.

At least 235 records · Page 13Linked to original sources
PubMed1989

Effects of cyclosporin A and verapamil on the intracellular daunorubicin accumulation in Chinese hamster ovary cells with increasing levels of drug-resistance.

Multidrug-resistance (MDR) is characterized by the presence of a 170 kDa glycoprotein (P-glycoprotein) in the plasma membrane. P-glycoprotein is thought to act as an efflux pump, leading to reduced drug accumulation in MDR cells. This defect in drug accumulation can be overcome by membrane transport modulating agents (MTMAs). We determined the concentration of MTMA needed for maximal restoration of daunorubicin content in 4 Chinese hamster ovary cell lines with increasing levels of drug-resistance using flow cytometry. Stimulation of daunorubicin accumulation occurred in a dose-dependent manner. The required level of MTMA needed for maximal drug accumulation increased with the level of drug-resistance. CHrA3 cells, which have a level of resistance comparable to clinical samples, needed relatively low concentrations of MTMA for maximal restoration of drug accumulation. This indicates that, in trial combining drugs and MTMAs, low dosages of MTMAs could be sufficient for optimal potentiation of cytotoxicity.

Animals↗
PubMed1989

Localization and imaging of radiolabelled monoclonal antibody against squamous-cell carcinoma of the head and neck in tumor-bearing nude mice.

Nude mice carrying human squamous-cell carcinoma xenografts were given i.v. injections of radiolabelled monoclonal antibodies (MAbs). MAb E 48, which reacts with squamous-cell carcinomas, was labelled with 131I, while a second control MAb of similar immunoglobulin subclass was labelled with 125I. Both antibodies were injected simultaneously, then the mice were scanned with a gamma camera or their tissues were removed and antibody uptake was calculated as a percentage of the injected dose. Uptake of E 48 reached a peak value of 16%/g on day 3, while uptake of the control antibody was less than 1.8%/g. By 24 hr after injection tumor could be visualized without subtraction techniques. At days 3 and 7, only xenografts were visible on imaging. These findings suggest that E 48 is capable of high specificity in targeting isotopes to squamous-cell carcinomas in an experimental setting.

Animals↗
PubMed1989

Resistance mechanisms in three human small cell lung cancer cell lines established from one patient during clinical follow-up.

Mechanisms for resistance were studied in three classic type, human small cell lung cancer cell lines, GLC14, GLC16, and GLC19, that were established from one patient during clinical follow-up. Clinically the tumor changed from sensitive (GLC14) to completely resistant to (chemo)therapy (GLC19) during this period. The stain with JSB-1 antibody, detecting the Mr 170,000 multidrug resistance associated glycoprotein, was most pronounced in GLC16 and absent in GLC19. Intracellular Adriamycin (Adr) concentrations were decreased in GLC16 and GLC19 versus GLC14. Glutathione levels were 12.9, 15.5, and 16.6 micrograms/mg protein; total sulfhydryl groups were 36.5, 45.7, and 48.8 micrograms/mg protein; and glutathione S-transferase activity was 13, 29, and 43 nmol I-chloro-2,4-dinitrobenzene/min/mg protein for GLC14, GLC16, and GLC19, respectively. Incubation with DL-buthionine-S,R-sulfoximine increased Adr and cisplatin induced cytotoxicity, whereas X-ray induced cytotoxicity remained the same. Catalase activity increased from 0.88 to 1.73 to 3.83 mumol H2O2/min/mg protein in, respectively, GLC14, GLC16, and GLC19. Compared to GLC14 and GLC16, Adr induced a higher amount of DNA strand breaks in GLC19. In none of the three cell lines could Adr induced DNA strand breaks be repaired. X-ray induced a comparable amount of DNA strand breaks in all three cell lines but all cell lines were capable of repairing the X-ray induced DNA strand breaks within 90 min. It is concluded that a number of different mechanisms are operative and that some but not all of the observed changes in mechanisms for drug resistance in these lines correlate with the clinical data.

Antineoplastic Agents↗
PubMed1989

Immunohistochemical detection of P-glycoprotein in human tumor cells with a low degree of drug resistance.

We report the immunohistochemical detection of the 170-180 kDa multi-drug-resistance-related P-glycoprotein in human tumor cells with a low level of resistance. A series of human squamous lung cancer cell lines with increasing levels of resistance to doxorubicin (DOX) was developed and stained for P-glycoprotein, using the JSB-IMAb. Subline SW1573/50A with a 4- to 6-fold cross-resistance to daunorubicin (DNR) and vincristine (VCR) showed rather uniform positive staining for P-glycoprotein apparently at cytoplasmic sites. Only in cells with higher degrees of resistance (greater than 10-fold) could plasma-membrane-associated P-glycoprotein be made visible. DNR efflux was increased in SW1573/50A as compared to the parent line SW1573 (52 and 70% DNR were retained during 3 min efflux respectively). Verapamil partially reversed DNR and VCR resistance in SW1573/50A. Cells obtained from a metastasized renal cell carcinoma and cultured in vitro stained in a similar way to SW1573/50A and showed some sensitivity to verapamil modulation of VCR cytotoxicity. Our results suggest that weakly resistant cancer cells obtained from patients can be routinely detected with JSB-I on cytospins, and implicate that in such weakly resistant cells P-glycoprotein may be present, while plasma membrane expression is not yet readily detectable.

ATP Binding Cassette Transporter, Subfamily B, Mem↗
PubMed1989

Immunohistochemical detection and quantitation of P-glycoprotein in multiple drug-resistant human myeloma cells: association with level of drug resistance and drug accumulation.

Using several multiple drug-resistant human myeloma cell lines as standards, we developed an immunohistochemical staining technique and means of quantitating P-glycoprotein in individual myeloma cells. The level of staining intensity for P-glycoprotein in individual myeloma cells was quantitated by measuring the average optical density of each cell with a microscopic computerized cell analysis system. Using this system, we observed that the level of P-glycoprotein for individual cells within a cell population of known drug sensitivity was very homogeneous (coefficient of variation less than or equal to 13%). Analysis of cell lines with gradually increasing levels of multidrug resistance (8226/S, 8226/Dox6 and 8226/Dox40) demonstrated a close association between the level of resistance to doxorubicin, defined by the mean lethal dose (D0) and the amount of P-glycoprotein on individual cells determined by the optical density (r = 0.82, P less than 0.0005). Intracellular doxorubicin (DOX) accumulation in the individual cell lines was inversely related to the level of drug resistance expressed as D0. P-glycoprotein was also detected in the marrow-derived myeloma cells of patients with drug refractory disease using immunohistochemical staining. The amount of P-glycoprotein in the cells of one patient was directly compared to the amount found in the simultaneously stained standard cell lines (8226/Dox6 and 8226/Dox40) by comparing the optical densities for individual cells. Using this immunohistochemical technique to detect and quantitate P-glycoprotein in patient myeloma cells and comparing it to standard multidrug resistant myeloma cell lines may be of value in determining the contribution of P-glycoprotein to clinical drug resistance in patients with multiple myeloma.

ATP Binding Cassette Transporter, Subfamily B, Mem↗
PubMed1989

Cell-mediated immunity is enhanced by cytostatic drugs continuously released at the site of antigenic stimulation.

Immunopotentiation by cytostatic drugs continuously released from osmotic minipumps, was investigated in a guinea-pig contact-sensitivity model. These pumps are designed to release their content within a period of 7 days. Vepeside (VP-16) and 5-fluorouracil were released into oxazolone-stimulated lymph nodes by subcutaneous implantation of pumps containing either of these drugs. The pumps were implanted at the intended sensitization site, 2 days before sensitization. Strong potentiation of T-cell-mediated immunity, evaluated by delayed-type hypersensitivity measurements, was observed with both drugs tested. Daily injections with VP-16 also caused an enhancement of the immune response. However, daily injections with 5-fluorouracil, a drug assumed to be cell-cycle-specific in its action, failed to potentiate delayed hypersensitivity to oxazolone. Intralesional administration of cytostatic drugs has been put forward as an effective treatment modality in various types of cancer. Therapeutic effects may depend on both local tumor cytotoxic and immunopotentiating activities. Our present results suggest that osmotic minipumps can be applied to broaden the applicability and effectiveness of local chemotherapy.

Adjuvants, Immunologic↗
PubMed1989

Mechanisms of human lymphocyte migration and their role in the pathogenesis of disease.

Lymphocyte recirculation is an essential component of the functional immune system, providing a means for constant surveillance of the organism's tissues by immunocompetent cells and, moreover, facilitating interactions between different cell types engaged in the immune response. Adhesive interactions between recirculating lymphocytes and the wall of high endothelial venules (HEV) are thought to play a central role in this process. These interactions are mediated by lymphocyte homing receptors expressed on the lymphocyte cell surface which recognize tissue-specific molecules on the endothelium. Moreover, LFA-1 is also involved in the regulation of lymphocyte traffic. In addition, recent evidence indicating that these mechanisms may also play a role in the pathogenesis of chronic inflammatory diseases and non-Hodgkin's lymphomas is discussed.

Animals↗
PubMed1989

Drug-resistance in multiple myeloma and non-Hodgkin's lymphoma: detection of P-glycoprotein and potential circumvention by addition of verapamil to chemotherapy.

The B-cell neoplasms, multiple myeloma and non-Hodgkin's lymphoma, frequently become drug resistant, despite initial responses to chemotherapeutic drugs. Tumor cells from eight patients with clinically drug-refractory disease were evaluated by immuno-histochemical staining for monoclonal immunoglobulin (Ig) expression, nuclear proliferation antigen, P-glycoprotein (P-gly) expression, and other cellular antigens. P-gly was detected on tumor cells from six of eight patients with drug-resistant disease. Of the six patients with P-gly-positive tumors, five patients had advanced multiple myeloma and one had a drug-refractory non-Hodgkin's lymphoma. Cellular RNA analysis confirmed the over-expression of P-gly. In an effort to overcome drug resistance, a pilot study evaluated possible verapamil enhancement of chemotherapy in these eight patients. All patients had developed progressive disease while receiving a regimen containing vincristine and doxorubicin, and seven of eight patients had previously received continuous infusion vincristine and doxorubicin plus oral dexamethasone (VAD). At the time of progressive disease, continuous infusion verapamil was added to the VAD regimen. Three of the eight patients who were refractory to vincristine and doxorubicin alone responded when verapamil was added to VAD. The three patients who responded had P-gly-positive tumors. Verapamil increased the intracellular accumulation of doxorubicin and vincristine in vitro for both a P-gly-positive myeloma cell line and tumor cells from two patients with end-stage myeloma which over-expressed P-gly. The dose-limiting side effect associated with the addition of verapamil to chemotherapy was temporary impairment of cardiac function, manifest as hypotension and cardiac arrhythmia. We conclude that P-gly expression occurs in drug-refractory B-cell neoplasms and may contribute to the development of clinical drug resistance. However, other factors, such as the proliferative activity of the tumor, may also play a role in determining response to chemotherapy. The administration of verapamil along with VAD chemotherapy may partially circumvent drug resistance in patients whose tumors over-express P-gly.

ATP Binding Cassette Transporter, Subfamily B, Mem↗
PubMed1989

Synergistic effects of locally administered cytostatic drugs and a surfactant on the development of delayed-type hypersensitivity to keyhole limpet haemocyanin in mice.

The immunomodulating effects of two locally administered cytostatic drugs, the active cyclophosphamide-derivative Z 7557 and the plant alkaloid VP-16, were compared with the effects of systemically administered cyclophosphamide and several established adjuvants: Freund's complete adjuvant, dextran sulphate, and dimethyl dioctadecyl ammonium bromide (DDA). All agents tested promoted the development of delayed-type hypersensitivity (DTH) to keyhole limpet haemocyanin (KLH) in mice. Locally administered cytostatic drugs were the most effective immunostimulatory compounds, whereas DDA was the least toxic agent tested. In order to increase the effectiveness and/ or reduce the toxicity of these agents we tested the efficacy of combinations of cytostatic drugs and DDA to enhance DTH. The results show that DDA and suboptimal amounts of locally administered cytostatic drugs act synergistically on DTH.

Adjuvants, Immunologic↗
PubMed1989

Expression of lymphocyte homing receptor as a mechanism of dissemination in non-Hodgkin's lymphoma.

To investigate whether the lymphocyte homing receptor (LHR), an adhesion molecule believed to play an important role in the control of normal lymphocyte circulation, influences the spread of non-Hodgkin's lymphoma (NHL), expression of LHR was examined in 107 NHL of various histologic and immunophenotypic subclasses. This analysis revealed that whereas NHL with a putative derivation from recirculating mature T and B lymphocytes almost invariably express high levels of LHR, those akin to sessile mature or immature lymphocytes tend to express lower levels of LHR. Furthermore, in a survey among diffuse large-cell lymphomas of the B-lineage, the tumors of 11 of 13 patients with stage III/IV disease expressed moderate to high levels of LHR, whereas only two of 17 patients with stage I/II disease had tumors that did so. These findings suggest that LHR is involved in the dissemination of NHL.

Antibodies, Monoclonal↗
PubMed1988

Characterization of a human small cell lung carcinoma cell line with acquired resistance to cis-diamminedichloroplatinum(II) in vitro.

A 6.4-fold cis-diamminedichloroplatinum(II) (CDDP) resistant human small cell lung carcinoma cell line (GLC4-CDDP) was developed to study acquired CDDP resistance in vitro. Compared to the sensitive cell line (GLC4), the GLC4-CDDP showed an increase in doubling time and a decrease in cloning efficiency, cellular size, double minutes per cell, cellular protein, and nuclear protein content. While a complete cross-resistance for tetraplatin and a partial cross-resistance for doxorubicin, melphalan, cadmium chloride, carboplatin, and cis-dichloro-trans-dihydroxo-cis-bis(isoprolylamine)platinum (IV) (resistance factor, respectively,4.0,5.8,2.1,1.5,2.9) was found, no cross-resistance for vincristine was found. In the GLC4-CDDP line in comparison to the GLC4 line, glutathione and total amount of sulfhydryl compounds was significantly increased, while glutathione S-transferase and glutathione reductase was the same. The platinum content in cells and nuclei was lower in the resistant line, but after correction for cellular protein or volume no difference was found. The amount of platinum bound to DNA was significantly lower in the GLC4-CDDP line. After a 1-h incubation with CDDP, the amount of Pt-GG adducts was the same and the amount of interstrand cross-links was reduced in the GLC4-CDDP line as compared to GLC4. In conclusion, in the GLC4-CDDP line the phenotype and genotype are changed and various mechanisms, such as decreased Pt-DNA binding, elevated glutathione, and reduced interstrand cross-links, play a role in the development of the CDDP resistance.

Amino Acids↗
PubMed1988

Monoclonal antibody JSB-1 detects a highly conserved epitope on the P-glycoprotein associated with multi-drug-resistance.

Resistance to multiple chemotherapeutic agents is a common clinical problem in the treatment of cancer. This resistance may occur before primary therapy or be acquired during treatment. We have generated a monoclonal antibody (MAb) (JSB-I), specific for a conserved epitope on the plasma membrane 170- to 180-kDa glycoprotein, the expression of which is strongly correlated with the degree of multi-drug resistance (MDR). JSB-I strongly binds to both Chinese-hamster-derived MDR cell lines and human MDR cell lines, including cell lines derived from lung and ovary. A drug-sensitive revertant line, and the corresponding drug-sensitive parent lines, showed only weak reactivity or none at all. JSB-I reacts strongly to air-dried or acetone-fixed cells and therefore has potential value for diagnostic detection of MDR cells in human tumor samples.

ATP Binding Cassette Transporter, Subfamily B, Mem↗
PubMed1988

Evidence that leukocyte function-associated antigen-1 is involved in recirculation and homing of human lymphocytes via high endothelial venules.

We report the results of mAb inhibition studies of human lymphocyte-high endothelial venule interaction in vitro. These studies in which T cells from both normal donors and from a LFA-1-deficient patient were used indicate that in addition to a system of organ-specific 90-kDa "homing" receptors on lymphocytes, LFA-1 is also involved in lymphocyte recirculation and homing.

Adult↗
PubMed1988

Suppressive effects of transdermal clonidine administration on contact hypersensitivity reactions in guinea pigs.

In the present study, immunopharmacological effects of clonidine-TTS on allergic contact dermatitis (ACD) to non-related, established contact sensitizers were investigated in guinea pigs. First, to evaluate the hypotensive effect of clonidine-TTS in guinea pigs, intra-arterial blood pressure was recorded. After 4 days of treatment with one (or two) TTS per animal, a reduction of arterial blood pressure from 71 +/- 1 to 51 +/- 2 mm Hg was observed. We subsequently assessed the effects of clonidine-TTS on contact hypersensitivity reactions to 2,4-dinitrochlorobenzene (DNCB) and 4-ethoxymethylene-2-phenyl-oxazolone (Ox). This study indicates that clonidine-TTS suppressed the elicitation of contact hypersensitivity reactions. The observed immunosuppressive effect of clonidine may account for the relatively weak hypersensitivity reactions to this drug in experimental animal studies. Further studies are needed to determine whether such findings are of relevance to the clinical use of clonidine in patient populations.

Administration, Cutaneous↗
PubMed1988

Hapten-specific antibodies in allergic contact dermatitis in the guinea pig.

A guinea pig model was used to investigate the potential role of hapten-specific antibodies in allergic contact dermatitis. Hapten-specific antibodies of both the IgG1, IgG2 and IgM (sub)classes could be readily detected by ELISA in sera obtained after single or repeated exposure to the allergens 2,4-dinitrochlorobenzene (DNCB) or 4-ethoxymethylene-2-phenyloxazolone. Experiments in which animals were strongly boostered with Freund's complete adjuvant support previous reports that hapten-specific antibodies are able to transfer contact skin reactions with a delayed time course into naive recipients. In contrast, epicutaneous sensitization procedures never allowed transfer of such reactivity. Upon transfer of immune sera to previously sensitized animals, existing allergic contact dermatitis was unaffected, except for a distinct suppression of induration, but not erythema, in the DNCB system. In actively sensitized animals, antibody-related suppression of induration was never observed. Hapten-specific hyposensitivity, as induced by repeated epicutaneous exposure, was also not related to circulating antibody levels. Our findings do not support the view that hapten-specific antibodies should be reconsidered as playing a potentially important role in allergic contact dermatitis.

Animals↗
PubMed1987

A rapid and simple hapten conjugation method for monoclonal antibodies to be used in immunoenzyme single and double staining procedures.

A rapid and simple method was developed for the haptenization of monoclonal antibodies (MAbs), to be used in immunoenzyme single and double staining techniques. Using this method minute amounts of MAbs can be haptenized without purification of the antibody or removal of the excess hapten. The haptenized MAb can be ready for use with 2.5 h. The method consists of a direct incubation of the antibody with the haptenizing agent and subsequent addition of an amino acid solution to stop the reaction. Commonly available reagents were tested, of which dinitrofluorobenzene, trinitrobenzene sulfonic acid and an oxazolone derivative gave the best results. The procedure was evaluated by using MAbs directed against lymphoid cell surface membrane antigens in an indirect immunoenzyme staining on frozen sections using peroxidase or alkaline phosphatase-conjugated anti-hapten antibodies as second step antibody. It was found that those MAbs, which show good staining results in conventional indirect immunoenzyme procedures, can also be used successfully after haptenization in single as well as double staining procedures. Combination of haptenized and biotynilated MAbs gave good results when weakly reactive MAbs had to be included in immunoenzyme double staining.

Antibodies, Monoclonal↗