Phase I and pharmacologic study of the combination of paclitaxel, cisplatin, and topotecan administered intravenously every 21 days as first-line therapy in patients with advanced ovarian cancer.

PURPOSE: To evaluate the feasibility of administering topotecan in combination with paclitaxel and cisplatin without and with granulocyte colony-stimulating factor (G-CSF) support as first-line chemotherapy in women with incompletely resected stage III and stage IV ovarian carcinoma. PATIENTS AND METHODS: Starting doses were paclitaxel 110 mg/m2 administered over 24 hours (day 1), followed by cisplatin 50 mg/m2 over 3 hours (day 2) and topotecan 0.3 mg/m2/d over 30 minutes for 5 consecutive days (days 2 to 6). Treatment was repeated every 3 weeks. After encountering dose-limiting toxicities (DLTs) without G-CSF support, the maximum-tolerated dose was defined as 5 microg/kg of G-CSF subcutaneously starting on day 6. RESULTS: Twenty-one patients received a total of 116 courses at four different dose levels. The DLT was neutropenia. At the first dose level, all six patients experienced grade 4 myelosuppression. G-CSF support permitted further dose escalation of cisplatin and topotecan. Nonhematologic toxicities, primarily fatigue, nausea/vomiting, and neurosensory neuropathy, were observed but were generally mild. Of 15 patients assessable for response, nine had a complete response, four achieved a partial response, and two had stable disease. CONCLUSION: Neutropenia was the DLT of this combination of paclitaxel, cisplatin, and topotecan. The recommended phase II dose is paclitaxel 110 mg/m2 (day 1), followed by cisplatin 75 mg/m2 (day 2) and topotecan 0.3 mg/m2/d (days 2 to 6) with G-CSF support repeated every 3 weeks.

Melanin-related metabolites as markers of the skin pigmentary system.

Three different groups of chemical intermediates are known to be formed during the synthesis of melanin in melanocytes: phenolic compounds, phenolic thio-conjugates, and indolic compounds. All these substances and their metabolites can be detected in urine. We measured the urinary excretion of 3,4-dihydroxyphenylalanine (dopa), 5-S-cysteinyldopa (5-S-CD), and 2 indolic compounds, namely 5-hydroxy-6-methoxyindole (5H6MI) and 5-hydroxy-6-methoxyindole-2-carboxylic acid (5H6MI2C) in urine samples of 4 groups of people with different contents of cutaneous melanin: Asian group, white group, and 2 groups of whites 1 with vitiligo and 1 with tyrosinase-negative oculocutaneous albinism. Dopa and 5-S-CD were determined with the method using high-performance liquid chromatography with an electrochemical detection. Indolic substances were measured by mass fragmentography with deuterium-labeled internal standards. Comparison of the melanin-related metabolites excreted in urine of people with different capacities for melanin biosynthesis indicates that, of all measured substances, 5H6MI2C is the best urinary marker of melanin formation in the skin pigmentary system.

Cyclic-AMP level and eicosanoid release from alveolar macrophages are differentially affected by high and low dose of platelet activating factor.

Antigen challenged alveolar macrophages (ac-AM) showed much higher basal prostaglandin E2 (PGE2) release (4,4-fold) and cAMP content (2,4-fold) than naive alveolar macrophages (AM). In naive AM 1 fM platelet activating factor (PAF) enhanced PGE2 release from 115 to 157 ng/5 x 10(6) cells but was inactive at 1 nM or 1 microM. In ac-AC 1 fM PAF enhanced PGE2 release from 510 to 670 ng/5 x 10(6) cells and inhibited leukotriene B4 (LTB4) release (from 6.0 to 4.8 ng/5 x 10(6) cells). At a 10(6)-fold higher concentration PAF inhibited PGE2 release (from 510 to 400 ng/5 x 10(6) cells) and stimulated LTB4 release (from 6.0 to 8.2 ng/5 x 10(6) cells). PAF-induced increase or decrease in PGE2 release was paralleled by changes in cellular cAMP (+35 and -17%, respectively). The specific PAF-antagonist BN 52021 completely reversed all PAF-induced effects while indomethacin inhibited only PAF-induced increase in PGE2 release and cAMP leaving LTB4 release unaffected. Similarly, the lipoxygenase inhibitor AA-861 inhibited PAF-induced rise in LTB4 release leaving the enhancement in PGE2 release and cAMP content unaffected. Present data show that PAF dose-dependently affects eicosanoid production and cAMP level in alveolar macrophages.

Sensitization enhances the adenylyl cyclase responsiveness in alveolar macrophages. Changes induced at post-receptor level.

Using membrane fractions (MF) from guinea pig alveolar macrophages (AM), we investigated the effects of sensitization and antigen challenge on the stepwise activation of adenylyl cyclase considering receptor binding, G-protein coupling and direct stimulation of the enzyme. Receptor binding studies, using [125I]ICYP as the beta-adrenoceptor specific ligand, show that neither receptor number (Bmax) nor receptor affinity constants (Kd values) were affected by sensitization or antigen challenge. Using forskolin as a direct stimulant of AC, alterations in the enzymatic activity of AC could be excluded. Pretreatment of the different MF with cholera toxin (CT, a toxin which eliminates GTPase activity) and subsequent stimulation of AC with GTP, shows an increased responsiveness in MF from sensitized and antigen challenged AM. In addition, pretreatment of MF from naive AM with increasing doses of CT results in a maximal AC response at the higher concentrations used (50-100 micrograms/mL), an effect not observed in MF from sensitized and antigen challenged AM. In these MF, the AC response still increases after pretreatment with such doses of CT. These data suggest that the enhanced AC responsiveness in AM, induced by sensitization and antigen challenge, results from alterations in alpha s-subunits.

Antigen challenge modifies the cyclic AMP response of inflammatory mediators and beta-adrenergic drugs in alveolar macrophages.

Adenylate cyclase activity was determined in alveolar macrophages (AMs) obtained from bronchoalveolar lavage (BAL) fluids of naive and antigen-challenged guinea pigs. After the anaphylactic reaction in ovalbumin-sensitized guinea pigs, the basal levels of cyclic AMP in AMs were significantly increased compared to the levels in naive AMS (1.87 +/- 0.22 versus 5.26 +/- 0.45 pmol cyclic AMP/5.10(6) cells). Prostaglandin E2 (PGE2), prostacyclin (DC-PGI2), histamine, isoprenaline and salbutamol stimulated adenylate cyclase activity more effectively in AMs obtained from sensitized guinea pigs after the booster injection compared to AMs obtained from non-treated animals. Moreover, DC-PGI2 and histamine, which were hardly able to induce a rise in cyclic AMP levels in naive AMs, become effective activators in AMs obtained after antigen challenge (100 and 60% increase in the response, respectively). Using selective receptor ligands, we have shown that beta 2-adrenoceptors and H2-subtype histamine receptors are functionally coupled to macrophage adenylate cyclase activity. The present data indicate that sensitization does not affect the configuration of the receptor on the outer membrane (no change in affinity constants), but affects other parts of the transmembrane signal system leading to the intracellular production of cyclic AMP (e.g. regulatory binding proteins or increases in the number of receptors).

Stimulation of cyclic AMP production in human alveolar macrophages induced by inflammatory mediators and beta-sympathicomimetics.

We have investigated the effects of inflammatory mediators and beta-adrenoceptor agonists on the adenylyl cyclase responsiveness in alveolar macrophages from control subjects, patients suffering from chronic obstructive pulmonary disease (COPD) and asthmatics. Basal cyclic AMP (cAMP) levels in alveolar macrophages from COPD patients were significantly elevated (plus 42%) as compared to controls. In addition, the adenylyl cyclase responsiveness to prostaglandin E2, histamine and the beta-adrenoceptor agonist salbutamol was significantly impaired in alveolar macrophages from COPD patients and asthmatics. The lipid mediator platelet activating factor showed no effect on cAMP production in all three alveolar macrophage populations. Furthermore, the cAMP-enhancing effects of isoprenaline, salbutamol and histamine appeared to be mediated via beta 2-adrenoceptors and histamine H2-receptor subtypes respectively. Taken together, these data suggest an intrinsic desensitization phenomenon in alveolar macrophages from COPD patients and asthmatics.

Cyclic AMP enhancing drugs modulate eicosanoid release from human alveolar macrophages.

The effect of the phosphodiesterase inhibitor isobutyl-methylxanthine (IBMX), salbutamol and sodium nitroprusside was evaluated regarding PGE2 and LTB4 release and cAMP and cGMP level in human alveolar macrophages obtained from controls and COPD patients. Basal levels per five million control- respectively COPD alveolar macrophages: cAMP 1.2 and 1.0 pmole; cGMP 8.4 and 9.1 fmole; PGE2 120 and 63 pg and LTB4 19.2 and 14.8 pg. In both populations IBMX increased cAMP level by 55-93% and salbutamol+IBMX by 285-252%. Except for the 61% rise in LTB4 release by salbutamol+IBMX the drugs hardly affected PGE2 and LTB4 release from control macrophages. In COPD alveolar macrophages, however, IBMX and IBMX+salbutamol largely reduced PGE2 release (63 vs 11 pg per 10(6) cells) but less efficiently increased LTB4. In both macrophage populations sodium nitroprusside (SNP) substantially increased (3-4 fold) cGMP level but did not affect eicosanoid production. Present results indicate that drugs which enhance cAMP level decrease PGE2 release from COPD macrophages and stimulate the release of LTB4 a chemotactic mediator involved in bronchial inflammatory reactions.

Identification of beta 2-adrenoceptors on guinea pig alveolar macrophages using (-)-3-[125I]iodocyanopindolol.

The beta-adrenoceptor antagonist (-)-3-[125I]iodocyanopindolol ([125I]ICYP) binds with high affinity and in saturable way to membranes of guinea pig alveolar macrophages. The equilibrium dissociation constant for [125I]ICYP is 24.3 +/- 1.2 pM, and the number of binding sites is 166.3 +/- 13.7 fmol/mg protein (N = 4, +/- SEM). Displacement studies with selective antagonists showed that [125I]ICYP labels beta 2-adrenoceptors on guinea pig alveolar macrophages.

Pulmonary granulocyte influx and impaired alveolar macrophage adenylyl cyclase responsiveness in developing respiratory distress.

Alveolar macrophages have recently been postulated as being involved in the aetiology of adult respiratory distress syndrome (ARDS). To evaluate their role, basal cyclic AMP levels and responsiveness of adenylyl cyclase alveolar macrophages were determined at four intermediate stages of developing respiratory distress in piglets using a protocol with repeated lung lavage. Examination of alveolar cells recovered from the subsequent lavages reveals an influx of granulocytes (neutrophils and eosinophils) within 1 h of two intensive lung lavages. During the developing respiratory distress the basal cyclic AMPlevel of alveolar macrophages increases and adenylyl cyclase responsiveness to prostaglandin E(2) (PGE(2)) and isoprelanaline diminishes. The previously observed impairment of macrophage activity can then be explained at a subcellular level.

Involvement of eicosanoids in platelet-activating factor-induced modulation of adenylyl cyclase activity in alveolar macrophages.

Platelet-activating factor (PAF) induces a dose-dependent biphasic response in adenylyl cyclase activity in antigen-challenged alveolar macrophages (AM), but not in naive AM. Intracellular cyclic AMP levels are enhanced by very low concentrations of PAF (10(-13)-10(-10) M) and decreased by higher PAF concentrations (10(-8)-10(-5) M). The PAF response of adenylyl cyclase could be completely blocked by pretreatment with the PAF receptor antagonist BN 52021. The adenylyl cyclase stimulatory and inhibitory phases are reversed by indometacin (inhibiting cyclo-oxygenase) and AA 861 (inhibiting lipoxygenase). These results show that the PAF-induced response of adenylyl cyclase activity in antigen-challenged AM is achieved by its modulation of intracellular arachidonic acid metabolism.

Adenyl cyclase activity in human alveolar macrophages.

The inflammatory mediators PGE2, DC-PGI2 and histamine as well as the beta-adrenergic drugs isoprenaline and salbutamol increase intracellular cyclic AMP-concentrations whereas Platelet Activating Factor does not induce any change in adenyl cyclase activity of normal human alveolar macrophages. Functional H2-histaminergic and beta 2-adrenergic receptor-subtypes are coupled to macrophage adenyl cyclase.

Protection against influenza A virus infection in mice by oral immunization with a polyvalent bacterial lysate.

A study is presented which investigated whether oral immunization with a polyvalent bacterial lysate (Paspat oral) can sufficiently enhance cell-mediated defense mechanisms to protect mice against influenza A virus infection. It was found that oral immunization reduced mortality due to influenza A infection with 15-70%, depending on the quantity of virus administered and and the moment of infection. Cyclosporin A severely reduced the protective effect of oral immunization, suggesting that a major effect of oral immunization in these studies is T-cell activation. The effect of oral immunization on macrophageal activity was evaluated by measuring cyclic-AMP in alveolar macrophages (AMs) obtained by bronchoalveolar lavage. Before infection, basal activity levels of AMs in immunized mice were significantly lower than in controls. Five days after infection, however, basal activity level of AMs in immunized mice was significantly higher than AM activity in controls. Stimulation of AMs with PGE2 significantly reduced cellular activity in both groups, before and after infection. However, cellular activity of AMs from immunized animals was less reduced than cellular activity of control macrophages. Activity of AMs of immunized animals was significantly more reduced by histamine than activity of control macrophages. It is concluded that oral immunization with Paspat oral stimulates T-cell-dependent immune mechanisms, resulting in protection against influenza A virus infection in mice.

Oral topotecan: bioavailablity and effect of food co-administration.

The aims of the study were twofold: (1) to evaluate the effect of food on the relative oral bioavailability of topotecan gelatin capsules in patients with solid tumours, and (2) to determine the absolute bioavailability of oral topotecan with reference to the intravenous (i.v.) formulation. The study had a randomized two-period cross-over design. On day 1 of the first treatment course patients were administered 2.3 mg m(-2) day(-1) of oral topotecan with or without a high-fat breakfast. They crossed over to receive the alternate regimen on day 2. In the second course (3 weeks later) fasted patients received topotecan orally (2.3 mg m(-2) day(-1)) or i.v. (1.5 mg m(-3) day). They crossed over to receive the alternate regimen on day 2. On days 3-5 of both treatment courses patients received oral topotecan. Plasma pharmacokinetics were performed on days 1 and 2 of the first and second course using a high-performance liquid chromatographic assay. Eighteen patients were enrolled in the study. The ratio of the area under the curve to infinity during fasted and high-fat treatment was 0.93+/-0.23 (90% confidence interval (CI) 0.83-1.03). Maximal plasma concentrations of topotecan were similar after ingestion of the capsules with (10.6+/-4.4 ng ml(-1)) or without food (9.2+/-4.1 ng ml(-1)) (P = 0.130). The time needed to reach maximal plasma levels was significantly prolonged after food intake (median 3.1 h, range 2.8-6.1) compared to fasted conditions (2.0 h, range 1.1-8.1) (P = 0.013). The absolute bioavailability of topotecan averaged 42+/-13% (90% CI 37-47%). The apparent terminal half-life was significantly longer after administration of oral topotecan (3.9+/-1.0 h) than after i.v. administration (2.7+/-0.4 h) (P < 0.001). Topotecan demonstrates suitable bioavailability for oral treatment. Co-administration of the topotecan gelatin capsules with a high-fat breakfast leads to a small decrease in absorption rate but does not affect the extent of absorption.