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Biomedical subjects

R A Hilger

Publications and source records attributed to R A Hilger.

33 records · Page 2Linked to original sources
PubMed2000

Clinical phase I dose escalation and pharmacokinetic study of high-dose chemotherapy with treosulfan and autologous peripheral blood stem cell transplantation in patients with advanced malignancies.

A Phase I dose escalation and pharmacokinetic study of the alkylating cytotoxic agent treosulfan was conducted to evaluate the maximum tolerated dose and the dose-limiting toxicities in patients with advanced malignancies rescued by autologous peripheral blood stem cell transplantation. Twenty-two patients (15 ovarian and 7 other carcinomas/lymphomas) with a median age of 48 years were treated with 28 high-dose courses. Treosulfan was infused over 2 h at escalating doses from 20 to 56 g/m2, and pharmacokinetic parameters were analyzed. At 56 g/m2, three of six patients experienced dose-limiting toxicities: diarrhea grade III/IV in three patients; mucositis/stomatitis grade III in one patient; toxic epidermal necrolysis in one patient; and grade III acidosis in one patient. Other low-grade side effects, including erythema, pain, fatigue, and nausea/vomiting, were recorded. Two patients died within 4 weeks after treatment because of rapid tumor progression and fungal infection, respectively. Plasma half-life, distribution volume, and renal elimination of treosulfan were independent of dose, whereas the increase in area under the curve was linear up to 56 g/m2 treosulfan. The maximum tolerated dose of high-dose treosulfan is 47 g/m2. A split-dose or continuous infusion regimen is recommended for future high-dose trials. In consideration of antineoplastic activity and limited organ toxicity, inclusion of high-dose treosulfan in combination protocols with autologous peripheral blood stem cell transplantation seems worthwhile.

Adult↗
PubMed1999

Reversal of MDR1-associated resistance to topotecan by PAK-200S, a new dihydropyridine analogue, in human cancer cell lines.

Recent data suggest that expression of the membrane P170-glycoprotein (P-gp) may confer resistance to the topoisomerase-I-interactive agent topotecan. The present study describes the cellular effects of a new dihydropyridine analogue, PAK-200S, on P-gp-mediated resistance to topotecan in human breast and ovarian tumour cells. PAK-200S at a non-cytotoxic concentration of 2.0 microM completely reversed resistance to topotecan in P-gp-expressing MCF-7/adr (breast) and A2780/Dx5 (ovarian) tumour cells, respectively, with no effects on parental cells. Cellular pharmacokinetic studies by reversed-phase high-performance liquid chromatography analysis showed significantly lower cellular drug concentrations of the pharmacologically active closed-ring lactone of topotecan in multidrug-resistant cells than in parental cells. PAK-200S was effective in restoring the cellular lactone concentrations of topotecan in resistant MCF-7/adr cells to levels comparable to those obtained in parental cells. Furthermore, exposure of MCF-7/adr cells to topotecan in the presence of PAK-200S significantly increased the induction of protein-linked DNA breaks. PAK-200S did not alter nuclear topoisomerase I-mediated ex vivo pBR322 DNA plasmid unwinding activity and topoisomerase-I protein expression. These results suggest that reversal of P-gp-mediated resistance to topotecan by PAK-200S was related to the restoration of cellular drug concentrations of the active lactone form of topotecan rather than a direct effect on topoisomerase-I function.

ATP Binding Cassette Transporter, Subfamily B, Mem↗
PubMed1998

Clinical pharmacokinetics of intravenous treosulfan in patients with advanced solid tumors.

Treosulfan (L-threitol- 1,4-bis-methanesulfonate, Ovastat) is a prodrug of a bifunctional alkylating agent with activity in ovarian carcinoma and other solid tumors. For a clinical and pharmacology study, patients with advanced, refractory, or resistant solid tumors were treated with a single-dose intravenous 30-min infusion of 8 or 10 g/m2 treosulfan. A sensitive method for the determination of treosulfan in plasma and urine by reverse-phase high-performance liquid chromatography was developed. A total of 14 plasma and urine treosulfan pharmacokinetics determinations were analyzed in the 8-g/m2 group and 7 were analyzed in the 10-g/m2 group, the maximum tolerated dose for this group of pretreated patients. The terminal half-life of treosulfan was in the range of 1.8 h. AUC and Cmax values were significantly (P < 0.01) higher in the 10-g/m2 group (AUC 708+/-168 versus 977+/-182 microg ml(-1) h, Cmax 465+/-98 versus 597+/-94 microg/ml). The mean urinary excretion of the parent compound was about 25% of the total dose delivered over 48 h (range 5-49%), and about 20% was excreted during the first 6 h after administration. Currently, a clinical phase I pharmacokinetics and dose-escalation trial with autologous blood stem-cell support has been started at 20 g/m2 treosulfan using a 2-h infusion protocol.

Adult↗
PubMed1997

Cellular determinants of resistance to indolocarbazole analogue 6-N-formylamino-12,13-dihydro-1,11-dihydroxy-13(beta-D-glucopyranosyl)- 5H-indolo[2,3-alpha]pyrrolo[3,4-c]carbazole-5,7(6H)-dione (NB-506), a novel potent topoisomerase I inhibitor, in multidrug-resistant human tumor cells.

Membrane protein-associated alterations in cellular drug accumulation have been recently implicated in resistance to topoisomerase I (TOP-I)-interactive drugs. The present study investigated the cellular determinants of resistance to the indolocarbazole compound NB-506 [6-N-formylamino-12,13-dihydro-1,11-dihydroxy-13(beta-D-glucopyranosyl)- 5H-indolo[2,3-alpha]pyrrolo[3,4-c]carbazole-5,7(6H)-dione], a structurally novel TOP-I-interactive drug, in parental and multidrug-resistant tumor cells expressing either the P-170 glycoprotein (Pgp170) or multidrug resistance protein (MRP). MRP-expressing 250-fold doxorubicin-resistant human fibrosarcoma HT1080/DR4 tumor cells were drug sensitive to NB-506 and camptothecin (CPT) (resistance factor: 0.7 and 0.8, respectively) with no alterations of TOP-I parameters including DNA relaxation, expression of TOP-I protein and mRNA. In contrast, doxorubicin-resistant human ovarian A2780/Dx5 tumor cells [pgp170 phenotype] were 6.2-fold resistant to NB-506, whereas resistance to CPT was 2.6-fold. HPLC analysis of cellular NB-506 accumulation showed no significant differences between A2780 and A2780/Dx5 cells (peak intracellular concentrations after 120-min exposure to 10 microM NB-506: 400+/-85.0 and 352+/-95.1 nmol NB-506/mg protein, respectively). However, resistant A2780/Dx5 cells expressed a lower amount of TOP-I mRNA and 29% protein levels of TOP-I compared to parental A2780 cells, resulting in decreased TOP-I catalytic activity (3.17+/-0.02 vs. 1.16+/-0.15 rel.U/microg nuclear protein) and reduced induction of NB-506-mediated cleavable complex formation in A2780/Dx5 cells. Furthermore, the lower induction of NB-506-induced protein-linked DNA breaks (PLDB) in A2780/Dx5 cells correlated with significantly decreased DNA 12.2-440 kb size fragmentation in these cells. The present study demonstrates that expression of MRP and Pgp170 does not confer resistance to NB-506. Resistance to indolocarbazole substance NB-506 in A2780/Dx5 cells was only related to downregulation of TOP-I associated with lower induction of cleavable complex formation and DNA fragmentation. The data reported herein may indicate that the new indolocarbazole compound NB-506 has potent antitumor efficacy in membrane-associated multidrug resistance.

ATP Binding Cassette Transporter, Subfamily B, Mem↗
PubMed1996

Inhibition of leukotriene formation and IL-8 release by the paf-receptor antagonist SM-12502.

We analyzed the effect of the PAF receptor antagonist (+)-cis-3,5-dimethyl-2-(3-pyridyl)thiazolidin-4-one hydrochloride (SM-12502) on the release of leukotriene B4 and IL-8 from human leukocytes. Peripheral blood from healthy donors was separated in two different fractions: polymorphonuclear leukocytes (PMN) and a lymphocyte, monocyte and basophil granulocyte cell fraction (LMB). After incubation of the cell population with different concentrations of SM-12502 the cells were subsequently stimulated with either the Ca ionophore A23187, the bacterial derived peptide fMLP, or with an activator of heterotrimetric G-proteins, the sodium fluoride (NaF, in the presence of Al3+). The PAF receptor antagonist led to a concentration and time dependent inhibition of LTB4 formation and IL-8 release from PMN and LMB. Our data clearly indicate an inhibitory effect of the PAF receptor antagonist SM-12502 on the formation of mediators of the lipoxygenase pathway and on the release of IL-8.

Aluminum↗
PubMed1996

Effect of the PAF-receptor antagonist SM-12502 on human platelets.

We analyzed the effect of the PAF receptor antagonist (+)-cis-3,5-dimethyl-2-(3-pyridyl)thiazolidin-4-one hydrochloride (SM-12502) on human platelet aggregation as well as mediator release. After incubation of human platelet with different concentrations of SM-12502 the cells were subsequently stimulated with either the Ca ionophore A23187, with human thrombin, or with an activator of heterotrimeric G-proteins, sodium fluoride (NaF, in the presence of Al3+). Preincubation of platelets with the PAF receptor antagonist led to an inhibition of 12-lipoxygenase derived 12(S)-HETE and cyclooxygenase derived 12(S)-HHT. Pretreatment of platelets with the PAF receptor antagonist SM-12502 prior to activation with the Ca ionophore A23187 or PAF also inhibited platelet aggregation. Our data clearly indicate an inhibitory effect of the new PAF receptor antagonist SM-12502 on the formation of platelet derived inflammatory mediators of the lipoxygenase pathway as well as of the cyclooxygenase pathway, and furtherone, treatment with the PAF receptor antagonist diminished platelet aggregation after subsequent specific and unspecific activation.

12-Hydroxy-5,8,10,14-eicosatetraenoic Acid↗
PubMed1996

Cetirizine exerts anti-inflammatory effects on human neutrophils.

Leukotrienes are potent lipid mediators involved in acute and chronic inflammatory processes and allergic inflammation. Cetirizine is an H1-receptor antagonist used in the treatment of allergic symptoms. We analyzed the effect of cetirizine on the formation of leukotriene B4 (LTB4) after stimulation of human peripheral blood neutrophils. The inflammatory mediators were analyzed after cellular activation with different stimuli: the Ca ionophore A23187, which bypasses membranous signal transduction elements; the bacterial peptide formyl-methionine-leucyl-phenylalanine (fMLP), which activates cells by binding to a GTP-protein (G-protein)-coupled receptor, and with sodium fluoride (NaF), which directly activates G-proteins. After cellular preincubation with cetirizine, the amounts of LTB4 generated from neutrophil granulocytes decreased significantly when the cells were subsequently stimulated with either fMLP or NaF, in contrast to stimulations with the Ca ionophore. The data provide evidence that cetirizine exerts anti-inflammatory effects apart from H1 antagonism.

Anti-Inflammatory Agents↗
PubMed1996

Dual effect of ammonium persulfate on the generation of leukotrienes from human neutrophil granulocytes.

Occupationally induced hypersensitive reactions towards chemical compounds are increasingly observed. In this regard ammonium persulfate (APS) is reported as an inducer of erythema, urticaria, eczema, rhinitis and bronchial asthma. On the basis of these reactions, persulfates obviously show some characteristics of skin-sensitizing as well as respiratory allergens. We measured the effect of APS on the release of inflammatory mediators, i.e. leukotriene B4 (LTB4), from human polymorphonuclear neutrophils (PMN). The effect of the persulfate was analyzed in comparison of the corresponding sulfate. After different cellular activation either with the Ca ionophore A23187, the tripeptide formyl-methionyl-leucyl-phenylalanine (fMLP) or sodium fluoride (NaF) coincubation of PMN with APS led to a diminished generation of LTB4. In addition it was demonstrated that APS decreased the stability of leukotrienes in cell-free systems. However, cells which were preactivated with APS and subsequently washed showed an increase in leukotriene formation after stimulation with fMLP or NaF but not with the Ca ionophore. Thus, the concentration of APS at local tissue sites as well as the occurrence and nature of a secondary cell-activating stimulus finally determine to what extent persulfates will interfere with cellular functions, e.g. mediator suppression or induction, which are then responsible for clinical disease processes.

Ammonium Sulfate↗
PubMed1992

Differential increase in 12-HETE release and CD29/CD49f expression of platelets from normal donors and from patients with atopic dermatitis by Staphylococcus aureus.

The generation of the arachidonic acid-derived inflammatory mediator 12-hydroeicosatetraenoic acid (HETE) and the expression of CD29 as well as CD49f from unstimulated and stimulated platelets has been studied in patients with atopic dermatitis (AD) as well as in healthy volunteers. Heat-killed clinical isolates of Staphylococcus aureus served as stimuli. Unstimulated platelets from patients with AD produced higher amounts of 12-HETE compared to platelets from normal donors. The absolute 12-HETE release from platelets of patients with AD was significantly higher compared to the control group after stimulation with heat-killed S. aureus, whereas the relative increase remained. The expression of CD29 and CD49f on unstimulated platelets of patients with AD was markedly enhanced compared to platelets from normal donors. Stimulation with S. aureus led to similar results as to the CD29 expression on normal and atopic platelets or to a markedly higher expression of CD49f on platelets from normal donors. As compared to platelets from normal donors the CD49f expression on atopic platelets was slightly enhanced by S. aureus. Our data emphasize that platelets may play an important role in the pathogenesis of AD by an increased preactivation and by an enhanced responsiveness to S. aureus which colonizes permanently the skin of patients with AD.

12-Hydroxy-5,8,10,14-eicosatetraenoic Acid↗
PubMed1992

Leukotriene A4 modulates generation of leukotriene B4 and sulphidopeptide leukotrienes by human neutrophils.

We investigated the influence of exogenous leukotriene A4 (LTA4) on the reactivity of polymorphonuclear leucocytes (PMN). PMN were either prestimulated with LTA4 or incubated simultaneously with LTA4 and the Ca ionophore A23187 or sodium fluoride (NaF). The Ca ionophore A23187 and NaF induced generation of LTB4 from PMN was significantly diminished in the presence of LTA4 while the formation of LTC4 was enhanced. In contrast, preincubation of cells with LTA4 followed by subsequent stimulation with NaF synergistically increased the LTB4 generation from PMN. LTA4, either alone or in combination with the calcium ionophore A23187 or NaF, decreases GTPase activity in human PMN. This decrease was abolished when LTA4 pretreated cells were subsequently stimulated with NaF, but not with calcium ionophore A23187, suggesting a regulatory role of LTA4 on G-proteins. The results demonstrate dual functions of LTA4: it serves as a substrate for the generation of leukotrienes and also regulates the susceptibility of human PMN for subsequent response.

Calcimycin↗
PubMed1991

Analysis of leukotriene B4 metabolism in human promyelocytic HL-60 cells.

We previously reported that human alveolar macrophages rapidly metabolize the chemotactic active lipid mediator leukotriene B4 (LTB4) into the dihydro-LTB4 by reduction of one of the conjugated double bonds. We herein report that human HL-60 cells (a myeloid precursor which can be differentiated into granulocyte- as well as monocyte-like cells by dimethyl sulphoxide or phorbol myristate acetate) express a highly active LTB4 reductase in the undifferentiated state. Differentiation by dimethyl sulphoxide (1.3%) along the granulocyte lineage, as confirmed by light microscopy, conversion of NitroBlue Tetrazolium into formazan, failed to induce a substantial capacity for omega-oxidation of LTB4; this reaction is exclusively found in mature granulocytes. Studies with the cell homogenate of undifferentiated HL-60 cells indicated that the activity of the enzyme depends on the presence of NADPH, Ca2+ and Mg2+, with a pH optimum of 7.5 at 37 degrees C. The enzyme was not released into the supernatant after stimulation of HL-60 cells with phorbol myristate acetate (100 ng) or Ca2+ ionophore (7.5 microM). Subcellular fractionation revealed evidence that the LTB4 reductase is located within the membrane fraction. Purification of the enzyme by gel filtration and gel electrophoresis suggests an apparent molecular mass of 40 kDa.

Chromatography, Gel↗
PubMed1991

Conversion of leukotriene A4 by neutrophils and platelets from patients with atopic dermatitis.

The generation of arachidonic acid-derived inflammatory mediators from unstimulated and stimulated neutrophils (PMN) and platelets in the presence of exogenous LTA4 has been studied in patients with atopic dermatitis (AD) as well as in healthy volunteers. PMN were stimulated with the interleukins IL-3, IL-8, C5a, and the Ca-ionophore A23187. In addition, NaF and thrombin were used to stimulate platelets. The release of leukotriene (LT)B4, 20-COOH- and 20-OH-LTB4, cysteinyl-leukotrienes and 12-HETE was measured. The proinflammatory mediator release from PMN and platelets of patients with AD was significantly higher as compared to the control group. The spontaneous conversion of LTA4 by PMN and platelets was markedly enhanced in patients with AD. Different results with receptor-specific and non-specific stimuli (Ca-ionophore A23187) in the presence of exogenous LTA4 were obtained. The results indicate a higher state of activation for enzymes involved in leukotriene formation. Furthermore, the production of 12-HETE by platelets from patients with AD was enhanced in unstimulated and stimulated cells. Our data emphasize that neutrophils and platelets may play an important role in the pathogenesis of AD by an increased responsiveness to receptor-specific stimuli and cell-cell interaction via LTA4.

Adolescent↗
PubMed1991

Interleukin-3, interleukin-8, FMLP and C5a enhance the release of leukotrienes from neutrophils of patients with atopic dermatitis.

The influence of the receptor-specific stimuli interleukin-3 (IL-3), interleukin-8 (IL-8), C5a and formyl-methionyl-leucyl-phenylalanine (FMLP) on the generation of arachidonic acid-derived inflammatory mediators from neutrophils (PMN) has been studied in patients with atopic dermatitis (AD) as well as in healthy, non-atopic volunteers. The release of leukotriene (LT)B4, the omega-oxidation products 20-COOH- and 20-OH-LTB4 and the cysteinyleukotriene LTC4 were measured by reverse-phase HPLC and radioimmunoassay. The incubation of neutrophils with these stimuli led to a significantly higher release of LTB4 and LTC4 in the AD group. The spontaneous leukotriene generation of PMN from patients with AD was on average threefold higher compared to the control group. C5a stimulated the release of LTB4 and its metabolites from atopic cells up to 9 ng in contrast to low amounts from non-atopic cells. Furthermore, FMLP distinctly enhanced the leukotriene release of neutrophils from patients with AD compared to unstimulated cells and to cells of normal donors. IL-3 and IL-8 also significantly stimulated the generation of LTB4 and LTC4 of PMN from atopic patients. Our data emphasize that neutrophils may play an important role in the pathogenesis of AD by an increased responsiveness to receptor-specific stimuli and further suggest that IL-3 and IL-8 influence the acute and chronic inflammatory reactions in patients with AD.

Cells, Cultured↗
PubMedDate not supplied

Immunosuppressive agents enhance the cytokine-induced priming of inflammatory cells.

The influence of the immunosuppressive substances cyclosporin A (CsA), FK506 and rapamycin on inflammatory mediators from cytokine-primed human leukocytes was studied. The cells were primed with granulocyte/macrophage-colony-stimulating factor, granulocyte-stimulating factor (G-CSF) and interleukin-3 (IL-3), and subsequently stimulated with the chemotactic peptide fMLP. The immunosuppressive agents CsA and FK506 enhanced the release of leukotriene B4 (LTB4) from human neutrophil granulocytes dependent on the priming with G-CSF and IL-3. IL-3-primed neutrophils released up to threefold higher amounts of LTB4 after subsequent stimulation with FK506 and fMLP. In contrast, the immunosuppressant rapamycin inhibited the formation of LTB4 after subsequent stimulation with fMLP. In addition to the well-documented inhibitory effects on lymphocyte function, our results indicate an enhancing effect of the immunosuppressive agents on the cellular inflammatory response of human peripheral blood neutrophils.

Cytokines↗