Phase II trial with Brequinar (DUP-785, NSC 368390) in patients with metastatic colorectal cancer: a study of the Early Clinical Trials Group of the EORTC.
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Adenocarcinoma↗
Biomedical subjects
T Skovsgaard
Publications and source records attributed to T Skovsgaard.
Epirubicin or epirubicin and vindesine in advanced breast cancer. A phase III study.
One hundred thirty-three evaluable patients with advanced breast cancer entered a randomized trial comparing epirubicin 60 mg/m2 with a combination of epirubicin 45 mg/m2 and vindesine 3 mg/m2 day 1 and 8 every 4 weeks. In all 10 premenopausal women an oophorectomy was performed. Seventy-five patients had previously received cyclophosphamide, methotrexate, and 5-fluorouracil (CMF) for advanced disease and 68 had received adjuvant chemotherapy (cyclophosphamide or CMF). Among evaluable patients (72 in the epirubicin group and 61 in the epirubicin + vindesine group) response rates were as follows: complete response--seven versus six; partial response--31 versus 22; no change--16 versus 17 (p greater than 0.40). Median time to disease progression was 6 months in both groups and median survival times were identical (12 months). Thrombocytopenia was less frequent in the epirubicin + vindesine group (p less than 0.01). In the epirubicin + vindesine group, mild to moderate peripheral neuropathy was observed in 40% of the patients. Congestive heart failure developed in one patient with a cumulative dose of epirubicin less than 1000 mg/m2 and in 7 of 15 patients who had greater than 1000 mg/m2. Four died of this cause. In conclusion, epirubicin is effective as a single agent for advanced breast cancer. The combination with vindesine does not increase its efficacy.
Adult↗
Daunorubicin and vincristine binding to plasma membrane vesicles from daunorubicin-resistant and wild type Ehrlich ascites tumor cells.
Tumor cell resistance to anthracyclines, epipodophyllotoxins and vinca alkaloids, called multi-drug resistance (MDR) is intimately linked to changes in the plasma membrane which facilitate an increased energy dependent drug extrusion in the resistant cell compared to the wild type cell. Isolated plasma membrane vesicles from wild type Ehrlich ascites tumor cells (EHR2) and the daunorubicin (DNR) resistant subline EHR2/DNR+ were utilised to study binding and possible transport of DNR and vincristine (VCR). A significant ATP enhanced increase in VCR binding to vesicles from EHR2/DNR+ compared to EHR2 was demonstrated. Furthermore, an increase in ATP enhanced VCR binding in proportion to content of the MDR associated P-glycoprotein was seen in plasma membrane vesicles prepared from various benign human endocrine tumors. VCR binding to EHR2/DNR+ vesicles was inhibited by other vinca alkaloids greater than actinomycin D greater than colchicine greater than anthracyclines, with 35-75 microM concentrations of anthracyclines needed for 50% inhibition. VCR binding to EHR2/DNR+ vesicles was pH and temperature dependent with an activation energy of -30 kJ/mol and was decreased by replacement of Na+ with K+ and by addition of Ca2+. Preincubation of vesicles with monoclonal antibody against the C terminal of P-glycoprotein had no effect on VCR binding and osmolality tests failed to show genuine transmembranal transport of VCR. DNR binding was similar in plasma membrane vesicles from both cell lines, and showed none of the characteristics mentioned for VCR. Furthermore, a radiolabeled N-hydroxysuccinimide ester derivative of doxorubicin, which inhibited VCR binding to EHR2/DNR+ membranes to an even greater extent than doxorubicin, labeled plasma membrane proteins from EHR2 and EHR2/DNR+ identically and did not demonstrate any binding to P-glycoprotein. Therefore, even though the study confirms the close link between vinca alkaloid binding and P-glycoprotein, it could not detect a similar association between anthracyclines and P-glycoprotein thus attesting to the complexity of the MDR phenotype.
ATP Binding Cassette Transporter, Subfamily B, Mem↗
Freeze-fracture study of plasma membranes in wild type and daunorubicin-resistant Ehrlich ascites tumor and P388 leukemia cells.
The plasma membrane is considered to play a major role in the development and maintenance of the multidrug resistance (MDR) phenotype, a role which may in part be mediated by an inducible 170 kD transmembrane protein (P-170). The present freeze-fracture study of plasma membranes of daunorubicin-resistant Ehrlich ascites and P388 leukemia cells demonstrated a significant increase in the density of intramembrane particles (IMP) in the P-face, but not the E-face, of resistant sublines compared with wild type cells. Furthermore, a three-dimensional histogram plot of the diameters of P-face IMPs in Ehrlich ascites tumor cells showed the emergence of a subpopulation of 9 X 11 nm IMPs not found in wild type cells. The size of these IMPs would be consistent with a MW of approximately 340 kD, thus indicating that P-170, shown to be present in both resistant cell lines by Western blot analysis and immunohistochemical staining, exists as a dimer in the plasma membrane. Incubation with the calcium channel blocker verapamil, in concentrations known to inhibit daunorubicin efflux in resistant cells, showed evidence of membrane disturbance in the form of IMP clustering in both wild type and resistant Ehrlich ascites tumor cells. However, incubation with daunorubicin itself did not alter the freeze-fracture morphology of the plasma membranes.
Animals↗
Inhibition of vincristine binding to plasma membrane vesicles from daunorubicin-resistant Ehrlich ascites cells by multidrug resistance modulators.
The multidrug resistance (MDR) phenotype is presumed to be mostly dependent on changes in the resistant cell plasma membrane, notably the emergence of a 170 kDa glycoprotein called P-glycoprotein, which facilitate increased drug efflux. We have previously demonstrated that ATP-enhanced binding of vincristine (VCR) to plasma membrane vesicles is much greater in MDR than in wild type cells. The present study has shown that VCR binding to MDR Ehrlich ascites tumour cell plasma membrane vesicles is inhibited 50% most efficiently by quinidine (0.5 microM) followed by verapamil (4.1 microM) and trifluoperazine (23.2 microM). This is the reverse order of the effect on whole cells where a ranking of efficiency in terms of enhancement of VCR accumulation, inhibition of VCR efflux, DNA perturbation and modulation of resistance in a clonogenic assay, was trifluoperazine greater than or equal to verapamil much greater than quinidine. The detergent Tween 80 inhibited VCR binding to plasma membrane vesicles at 0.001% v/v which agreed with the level which modulated resistance and increased VCR accumulation in whole cells. No effect was observed on daunorubicin binding to MDR plasma membrane vesicles after incubation with either Tween 80 (up to 0.1% v/v) or verapamil (up to 25 microM). We conclude that the effect of a modulating drug in reversing resistance to VCR correlates with its ability to raise intracellular VCR levels but not with its capability to inhibit VCR binding to the plasma membrane. Thus, enhancement of VCR accumulation in MDR cells is hardly solely due to competition for a drug binding site on P-glycoprotein. Furthermore, the lack of a demonstrable effect on daunorubicin binding to the plasma membrane by modulators points to transport mechanisms which do not utilise specific drug binding to the plasma membrane.
Animals↗
In vitro evaluation of the potential of aclarubicin in the treatment of small cell carcinoma of the lung (SCCL).
The sensitivity of eight cell lines established from treated and untreated patients with small cell carcinoma of the lung (SCCL) was tested in the clonogenic assay with 1 h and continuous exposure to aclarubicin (ACLA), adriamycin (ADR), daunorubicin (DAU) and mitoxantrone (MITO). The sensitivity to ADR, DAU and MITO covariated, and varied with a factor of five. The sensitivity to ACLA was independent of the sensitivity to ADR and varied only within a factor of two. Only ACLA showed pronounced increased potency with continuous incubation, and ACLA was the most potent drug in the three cell lines least sensitive to ADR. Two resistant cell lines were selected by treating NCI-H69 in vitro with DAU. One cell line (9-fold resistant to DAU) expressed large amounts of P-glycoprotein, the other cell line (4-fold resistant to DAU) had barely detectable glycoprotein. Both lines acquired resistance to ADR, ACLA and MITO. The cross-resistance to ACLA and MITO was only partial and ACLA was still the most potent drug on these lines. The sensitivity to ACLA of the cell lines least sensitive to ADR suggest that ACLA partially circumvents mechanisms of multidrug resistance. Together with the pronounced increase in potency with prolonged exposure, these results suggest that ACLA has a mechanism of action different from the 'classical' anthracyclines. In this context mitoxantrone is more similar to the classical anthracyclines although its structure is more dissimilar.
ATP Binding Cassette Transporter, Subfamily B, Mem↗
Anthracycline resistance.
The major limitation of the usefulness of anthracyclines is the development of drug resistance. Most of the data related to anthracycline resistance has been obtained in experimental animal tumors or in cultured human cell lines. An important characteristic obtained in these models is the common finding of cross-resistance to a series of drugs structurally unrelated to the anthracyclines and with a mechanism of action which apparently differ from that of the anthracyclines. This phenomenon is defined as pleiotropic drug resistance or multidrug resistance (MDR). The cross-resistance demonstrated includes the vinca alkaloids, actinomycin D, colchicin and the epipodophyllotoxins. The best documented mechanism of resistance and mechanism of cross-resistance is decreased cellular drug accumulation. Several findings indicate that the underlying factor of the altered net uptake is an active drug extrusion in the resistant cells. Another important characteristic is the demonstration of a cell surface protein with a molecular weight of about 170,000 (GP 170). Genetic and biochemical evidence indicate that the MDR phenotype in both animal and human cells result from overexpression of the MDR-1 gene which encodes GP 170. Some findings suggest that GP 170 is a transport protein for cytotoxic hydrophobic drugs like the anthracyclines. Other findings suggest that this gene is also associated with intrinsic (natural) anthracycline resistance.
Animals↗
The carboxylic ionophore monensin inhibits active drug efflux and modulates in vitro resistance in daunorubicin resistant Ehrlich ascites tumor cells.
Acquired cellular resistance to anthracycline and vinca alkaloid drugs (pleiotropic resistance) is commonly associated with reduced drug accumulation, a phenomenon which is thought to be partly due to increased energy-dependent drug efflux. We have previously detected increased plasma membrane traffic to, and content of, the acid endosomal compartment in pleiotropic resistant Ehrlich ascites and P388 leukemia cells. This suggested that the endosome could be associated with the pleiotropic resistance phenotype by a mechanism of vesicular drug trapping and transport. The present study was undertaken in order to test the effects of the carboxylic ionophores monensin and nigericin, which are known to both disrupt intracellular vesicular traffic and to raise intravesicular pH, with relation to the pleiotropic resistance phenotype. Both monensin and nigericin increased daunorubicin (DNR) accumulation in daunorubicin resistant Ehrlich ascites tumor cells (EHR2/DNR+) in a dose-dependent manner. Further, monensin inhibited glucose induced DNR efflux from EHR2/DNR+ cells loaded with drug by energy deprivation. On the other hand, monensin had only negligible effect on DNR accumulation and efflux in wild-type Ehrlich ascites tumor cells (EHR2). In a clonogenic assay system, monensin reduced resistance to DNR in EHR2/DNR+, whereas only an additive effect was obtained in EHR2. However, both ionophores proved too toxic in in vivo experiments. These results, showing that drugs known to disrupt endosomal functions also inhibit the pleiotropic resistance phenotype, support the suggested link between the endosome and pleiotropic resistance.
Animals↗
Lectin staining patterns of plasma membranes of daunorubicin and vincristine resistant Ehrlich ascites tumour cells.
The plasma membrane is considered to play a major role in the development of resistance to anthracycline and vinca alkaloid drugs (pleiotropic resistance). Previous studies have reported an increase in plasma membrane carbohydrates in pleiotropic resistant cells compared with wild-type cells. The present study has utilized a panel of 11 lectins and the streptavidin-biotin histochemical technique in order to compare plasma membrane carbohydrates from wild-type Ehrlich ascites tumour cells with cells from daunorubicin and vincristine resistant sublines. While the lectins ConA, LCA, PSA, PNA after neuraminidase and WGA stained plasma membranes of daunorubicin-resistant cells to a significantly greater degree than those of wild-type cells, no difference was apparent between vincristine-resistant and wild-type cells. PWM and WGA after neuraminidase pretreatment showed similar staining of the wild-type and both resistant sublines, while SBA with and without neuraminidase pretreatment, HPA, DBA, LTA and UEA I demonstrated either very weak or negative reactions with all sublines. We conclude that the observed increase in plasma membrane carbohydrate found in anthracycline-resistant cells is possibly due to drug action during acquisition and maintainance of resistance, and, though conceivably of importance in the development of resistance towards anthracyclines, is without significance for the pleiotropic resistance phenotype itself.
Animals↗
Multidrug resistance: drug extrusion and its counteraction by chemosensitizers.
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Antineoplastic Agents↗
Proopiomelanocortin producing cells of spleen: increase after transplantation with opioid-peptide producing Ehrlich ascites tumor cells.
Proopiomelanocortin (POMC) peptides are produced by many cell systems, including a population of macrophage-like cells in mouse spleen. After transplantation of mice with Ehrlich ascites tumor cells, the number of POMC producing spleen cells increase up to 10-fold by 5 to 6 days. The POMC peptides produced by these cells increase even more, as evidenced by radioimmunoassay. Thus, these data indicate both proliferation of splenic POMC cells and increased production of POMC peptides per cell after tumor challenge. Characterization of the peptides by sequence-specific radioimmunoassays and high performance liquid chromatography documents the presence of both ACTH(1-39) and of ACTH(1-14) in these cells. These peptides have multifacetted effects on immune parameters and may exhibit a general antiinflammatory action, partly mediated through inhibition of interleukin 1-stimulated events. The tumor cells themselves do not produce POMC peptides, but display met- and leu-enkephalin immunoreactivity. Also cultured tumor cells display such immunoreactivity, indicating endogenous production of opioid peptides. The opioid peptides of the tumor cells may both affect host immune defenses and play intratumoral autocrine or paracrine roles.
Animals↗
Effect of verapamil on daunorubicin accumulation in Ehrlich ascites tumor cells.
Previous studies have demonstrated that verapamil may overcome resistance to anthracyclines. In vitro and in vivo experiments were performed on wild-type and resistant Ehrlich ascites tumor cells. Verapamil in concentrations of 25-50 microM enhances the accumulation of daunorubicin (DNR) in resistant cells to the same level as in wild-type cells. No significant effect of verapamil on influx or nuclear binding could be demonstrated, indicating that verapamil enhances DNR uptake by blocking active drug extrusion. Exposure of cells to a high concentration of Ca2+ did not influence the effect of verapamil on DNR accumulation, suggesting a different mode of verapamil action apart from the Ca2+-blocking effect. Attempts to circumvent acquired resistance to DNR in vivo with verapamil showed that the combination of the two drugs was more toxic than DNR given alone. The LD10 of DNR was determined as 3 mg/kg and the LD10 of the combination, as 2.5 mg/kg. The therapeutic effect of verapamil at a dose of 50 mg/kg and DNR of 2.5 mg/kg increased the life span of the mice by 50%. No difference was seen in the wild-type tumor in vivo. These data lead us to conclude that verapamil can reverse DNR resistance completely, but that verapamil at non-toxic dosage only reduces DNR resistance by 50% in vivo.
Animals↗
Increased plasma membrane traffic in daunorubicin resistant P388 leukaemic cells. Effect of daunorubicin and verapamil.
Numerous studies have indicated that the plasma membrane plays an important role in the development of resistance to anthracycline and vinca alkaloid drugs (pleiotropic resistance). We have previously shown that pleiotropically resistant Ehrlich ascites tumour cells, which are of epithelial origin, have a significantly increased plasma membrane traffic (endo/exocytosis) to the endosomal compartment compared to sensitive cells. The present study, using the same ultrastructural morphometric technique, has demonstrated a similar significant difference in plasma membrane traffic between daunorubicin resistant and sensitive P388 cell lines (which are of lymphoid origin). Furthermore, we have shown that this difference between the P388 sublines is accompanied by an approximately 4 fold increase in the plasma membrane area participating in recycling together with an increased endosomal volume, number and membrane area in resistant cells. Plasma membrane traffic in resistant cells was significantly inhibited by the calcium channel blocker verapamil, a well known modulator of anthracycline resistance, but unaffected by daunorubicin itself. The confirmation of this phenotype in an additional pleiotropically resistant cell type with a different histogenesis further supports a hypothesis of endosomal drug trapping and vesicular extrusion as a possible resistance mechanism.
Animals↗
Increase in nonspecific adsorptive endocytosis in anthracycline- and vinca alkaloid-resistant Ehrlich ascites tumor cell lines.
Numerous biochemical and drug transport studies have focused on the role of the plasma membrane or membrane-associated processes in anthracycline and vinca alkaloid resistance. Cationized ferritin, which binds electrostatically to anionic sites on the plasma membrane, was used as a membrane marker in a morphometrical ultrastructural study. The nonspecific adsorptive endocytosis was significantly increased in Ehrlich ascites tumor cell lines resistant to daunorubicin (DNR), doxorubicin, vincristine, and vinblastine compared to that in sensitive cells. Thus cells resistant to DNR internalized 2.04% of their plasma membrane per minute, in contrast to 1.17% in sensitive cells. Because the measured cell surface areas in sensitive and resistant cells were unchanged during the experiments, increased membrane traffic (recycling) from intracytoplasmic compartments to the plasma membrane must have occurred in resistant cells. Possible implications of these findings include a resistance-linked phenotype without significance for resistance per se, an increase in membrane repair, and a connection between increased membrane traffic and increased active drug extrusion in resistant cells.
Adsorption↗
Pharmacodynamic aspects of aclarubicin with special reference to daunorubicin and doxorubicin.
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Aclarubicin↗
Interaction among two subpopulations of Ehrlich ascites tumor in vivo: evidence of a contact mediated immune response.
Clonal interaction among two subpopulations of Ehrlich ascites carcinoma was studied during in vivo growth in immune competent N/D mice in which the cell lines had been propagated for several years as ascites tumors. A growth inhibitory interaction by a subcutaneously slow growing subpopulation (E1.15) on a fast growing subpopulation (E1.95) was demonstrated only when the cells had contact during solid tumor growth. The effect was dependent on the relative proportion of the suppressing cell line. An identical effect was exerted by radiation killed inhibitor cells. The inhibition was only transient. If the tumor cell lines were grown intraperitoneally as ascites tumors without cellular contact, no interaction was found using flow cytometric DNA analysis to determine alterations in the relative proportions of the cell lines. Ascites from the inhibitor cell line E1.15 had no inhibitory effect on E1.95. Pre-immunization with radiation killed E1.15 cells or simultaneous growth of E1.15 in the opposite flank did not affect the growth of E1.95 significantly. A mononuclear cell infiltrate was found to surround the subcutaneously growing E1.15 tumors in immune competent N/D mice. This was not the case in T-lymphocyte deficient athymic nude mice in which E1.15 grew without delay subcutaneously. It is suggested that the E1.15 cell line was able to elicit a T-lymphocyte immune response only when grown subcutaneously, and that the close contact between E1.15 and E1.95 in mixed tumors would induce a non-specific growth inhibition of E1.95 cells which themselves were not able to induce a T-lymphocyte response.
Animals↗
Treatment of small cell lung cancer with 6-mercaptopurine: a phase II study.
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Adult↗
Chemosensitizers counteracting acquired resistance to anthracyclines and vinca alkaloids in vivo. A new treatment principle.
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Animals↗