A phase II study of G17DT in gastric carcinoma.

PURPOSE: G17DT is a gastrin immunogen, raising antibodies that blockade gastrin-stimulated growth. The aim of the study was to characterise antibody response and assess safety and tolerability of G17DT given to patients with gastric cancer. EXPERIMENTAL DESIGN: G17DT was administered to 52 patients with gastric adenocarcinoma at weeks 0, 2 and 6 by intramuscular injection at doses of 10, 100 and 250 microg. Antibody levels were measured by an ELISA assay. A radioligand displacement assay determined the ability of G17DT-immunised patients' sera to inhibit binding of 125IG17 to cholecystokinin (CCK)-2 receptors. RESULTS: By week 12 of the study, 6/12 evaluable stage I-III patients achieved an antibody response in the 10 microg group, 7/11 in the 100 microg group, and 11/12 in the 250 microg group. Stage IV patients dosed at 250 microg achieved a similar response rate to stage I-III patients dosed at 10 or 100 microg. G17DT was well tolerated in 47/52 patients. Two patients suffered significant adverse reactions including injection site pain and abscess. G17DT antibodies displaced iodinated gastrin from CCK-2 receptors, with the level of displacement correlating with antibody titre. CONCLUSIONS: G17DT immunisation is a well-tolerated method of raising functional antibodies to 17 amino acid gastrin forms in patients with gastric carcinomas.

A vaccination strategy for the long-term suppression of androgens in advanced prostate cancer.

OBJECTIVES: We have previously reported the ability of D17DT (formerly GnRH-DT) vaccination to produce castrate levels of androgens in men with advanced prostate cancer. This study examines the efficacy and tolerability of 3 and 15 micrograms of D17DT in 12 patients with advanced prostate cancer to establish a dose-response relationship. METHODS: 12 patients received either 3 or 15 micrograms of D17DT as 3 deep intramuscular injections over 6 weeks. Outcome was assessed in terms of physical and biochemical evaluations of clinical progression and antibody titres. RESULTS: Significant titres of anti-GnRH antibodies were detected in 2 out of 6 subjects who received 15 micrograms of D17DT; suppression of testosterone to castrate levels accompanied by a significant and prolonged reduction in PSA was also demonstrated. No responses were seen following treatment with 3 micrograms of D17DT. CONCLUSION: The induction of anti-GnRH antibodies through vaccination with 15 micrograms D17DT can produce and sustain castrate levels of testosterone in men with advanced prostate cancer.

Rational use of in vitro P-glycoprotein assays in drug discovery.

P-glycoprotein (Pgp) affects the absorption, distribution, and clearance of a variety of compounds. Thus, identification of compounds that are Pgp substrates can aid drug candidate selection and optimization. Our goal was to evaluate three assays used to determine whether compounds are Pgp substrates. Sixty-six compounds were tested in monolayer efflux, ATPase, and calcein-AM assays. Assay results yielded two categories of compounds. Category I (n = 35) exhibited concordance across the assays. Category II (n = 31) revealed differences among the assays that related to the apparent permeability (P(app)) of the compounds. Within category II, two groups were discerned based on the absence (group IIA, n = 10, nontransported substrates) or presence (group IIB, n = 21, transported substrates) of monolayer efflux. Detection of efflux (group IIB) was associated with compounds having low/moderate P(app) values (mean = 16.6 nm/s), whereas inability to detect efflux (group IIA) was associated with compounds having high P(app) values (mean = 535 nm/s). The calcein-AM and ATPase assays revealed Pgp interactions for highly permeable group IIA compounds but were less responsive than monolayer efflux for low/moderate P(app) compounds of group IIB. All assays detected substrates across a broad range of P(app), but the efflux assay was more prone to fail at high P(app), whereas the calcein-AM and ATPase assays were more prone to fail at low P(app). When P(app) is low, efflux is a greater factor in the disposition of Pgp substrates. The efflux assay is more reliable at low/moderate P(app) and is the method of choice for evaluating drug candidates despite low throughput and reliance on liquid chromatography with tandem mass spectrometry.

Anti-GnRH antibodies can induce castrate levels of testosterone in patients with advanced prostate cancer.

D17DT consists of the GnRH decapeptide linked to diphtheria toxoid. The aim of this pilot study was to assess the tolerance of D17DT and the production of anti-GnRH antibodies from two doses, 30 and 100 microg, in patients with locally advanced prostate cancer. Twelve patients with histologically proven prostate cancer in whom hormonal therapy was indicated were recruited. Patients received either 30 or 100 microg given intramuscularly on three separate occasions over six weeks. Patients were followed up and blood was taken for estimation of serum testosterone, PSA and anti-GnRH antibody titre. Overall the drug was well tolerated. In 5 patients a significant reduction in serum testosterone and PSA was seen. Castrate levels of testosterone were achieved in 4 and maintained for up to 9 months. Patients with the highest antibody titre had the best response in terms of testosterone suppression. This study shows that it is possible to immunize a patient with prostate cancer against GnRH to induce castrate levels of testosterone. This state appears to be reversible. This novel form of immunotherapy may have advantages over conventional forms of hormonal therapy and further studies are warranted in order to try and increase the proportion of responders.

Influence of passive permeability on apparent P-glycoprotein kinetics.

PURPOSE: The objectives of this work were to evaluate the importance of moderate passive permeability on apparent P-glycoprotein (P-gp) kinetics, and demonstrate that inspection of basolateral to apical and apical to basolateral (BL-AP/AP-BL) permeability ratios may result in a compound being overlooked as a P-gp substrate and inhibitor of another drug's transport via P-gp inhibition. METHODS: The permeability ratios of nicardipine, vinblastine, cimetidine, and ranitidine were determined across Caco-2 monolayers that express P-gp, in the presence and absence of the specific P-gp inhibitor, GF120918. In addition, the permeability ratio of vinblastine was studied after pretreatment of Caco-2 monolayers with nicardipine, ranitidine, or cimetidine. Similar studies were repeated with hMDRI-MDCK monolayers. RESULTS: The permeability ratios for cimetidine and vinblastine were >2. The permeability ratios for nicardipine and ranitidine were close to unity, and were not affected by the addition of GF120918. Based solely on ratios, only compounds with moderate transcellular permeability (vinblastine and cimetidine) would be identified as P-gp substrates. Although the permeability ratios appeared to be unity for nicardipine and ranitidine, both compounds affected the permeability of vinblastine, and were identified as substrates and inhibitors of P-gp. Studies performed in hMDR1-MDCK cells confirmed these experimental results. Data were explained in the context of a kinetic model, where passive permeability and P-gp efflux contribute to overall drug transport. CONCLUSIONS: Moderate passive permeability was necessary for P-gp to reduce the AP-BL drug permeability. Inspection of the permeability ratio after directional transport studies did not effectively identify P-gp substrates that affected the P-gp kinetics of vinblastine. Because of the role of passive permeability, drug interaction studies with known P-gp substrates, rather than directional permeability studies, are needed to elucidate a more complete understanding of P-gp kinetics.

P-glycoprotein-mediated transport of morphine in brain capillary endothelial cells.

Cell accumulation, transendothelial permeability, and efflux studies were conducted in bovine brain capillary endothelial cells (BBCECs) to assess the role of P-glycoprotein (P-gp) in the blood-brain barrier (BBB) transport of morphine in the presence and absence of P-gp inhibitors. Cellular accumulation of morphine and rhodamine 123 was enhanced by the addition of the P-gp inhibitors N-{4-[2-(1,2,3,4-tetrahydro-6,7dimethoxy-2-isoquinolinyl)-ethyl]-phenyl}-9,10-dihydro-5-methoxy-9- carboxamide (GF120918), verapamil, and cyclosporin A. Positive (rhodamine 123) and negative (sucrose and propranolol) controls for P-gp transport also were assessed. Morphine glucuronidation was not detected, and no alterations in the accumulation of propranolol or sucrose were observed. Transendothelial permeability studies of morphine and rhodamine 123 demonstrated vectorial transport. The basolateral to apical (B:A) fluxes of morphine (50 microM) and rhodamine (1 microM) were approximately 50 and 100% higher than the fluxes from the apical to the basolateral direction (A:B), respectively. Decreasing the extracellular concentration of morphine to 0.1 microM resulted in a 120% difference between the B:A and A:B permeabilities. The addition of GF120918 abolished any significant directionality in transport rates across the endothelial cells. Efflux studies showed that the loss of morphine from BBCECs was temperature- and energy-dependent and was reduced in the presence of P-gp inhibitors. These observations indicate that morphine is transported by P-gp out of the brain capillary endothelium and that the BBB permeability of morphine may be altered in the presence of P-gp inhibitors.

Role of P-glycoprotein on the CNS disposition of amprenavir (141W94), an HIV protease inhibitor.

PURPOSE: To determine the role of P-glycoprotein (Pgp) on the CNS penetration of the HIV protease inhibitor (PI) amprenavir (141W94) and to test the hypothesis that co-administration of a second HIV PI (ritonavir) could enhance amprenavir's brain penetration in vivo. METHODS: Pgp-mediated efflux was investigated in vitro with Caco-2 cells and in vivo by whole-body autoradiography (WBA). "Genetic" mdr1a/1b double knockout mice, "chemical" Pgp knockout mice generated by administration of the Pgp inhibitor GF120918, and mice pretreated with ritonavir were used in WBA studies to investigate the effects of Pgp modulation on the CNS penetration of amprenavir. RESULTS: Amprenavir, indinavir, ritonavir, and saquinavir had 2- to 23-fold higher transport rates from the basolateral to apical direction than from the apical to basolateral direction across Caco-2 monolayers. Incubation with GF120918 negated this difference, suggesting that the efflux was Pgp-mediated. WBA studies demonstrated a 13- and 27-fold increase in the brain and a 3.3-fold increase in the CSF concentrations of amprenavir in mice pretreated with GF120918 and in mdr1a/1b double knockout mice. In contrast, pretreatment with ritonavir did not alter the CNS exposure of amprenavir. CONCLUSIONS: These results provide evidence that amprenavir and other HIV PIs are Pgp substrates and that co-administration of a specific Pgp inhibitor will enhance amprenavir's CNS penetration in vivo. These results will have an important therapeutic impact in the treatment of AIDS dementia.

Comparison of antihypertensive and lipid actions of terazosin and atenolol in essential hypertension.

Terazosin is a selective alpha 1-adrenoceptor antagonist; its actions on the serum lipoprotein profile were compared with those of the cardioselective beta-adrenoceptor antagonist atenolol in 40 patients with mild to moderate hypertension. Atenolol and terazosin were titrated over six weeks until blood pressure control (diastolic blood pressure less than 90 mmHg or greater than 10 mmHg fall in blood pressure) or a maximum dose of atenolol 100 mg or terazosin 10 mg had been achieved. Patients not controlled were then prescribed additional diuretic therapy (cyclopenthiazide 0.5 mg and potassium 1200 mg). At each visit blood pressure and adverse events were recorded; plasma lipids were measured at baseline, six and 12 weeks. During titration there was a linear decrease in systolic (-29 mmHg on atenolol and -24 mmHg on terazosin) and diastolic blood pressure (-17 and -12 mmHg) in both groups without subsequent change over the next six weeks; atenolol reduced heart rate (-11 bpm) without change on terazosin. During the initial six weeks the total cholesterol fell in both groups; however, there were significant between-treatment differences in triglyceride responses with a fall on terazosin and a rise on atenolol. Comparing atenolol and terazosin over the total 12 week study (irrespective of thiazide treatment) increased triglyceride levels and reduced cholesterol ratios and HDL were demonstrated on atenolol, contrasting with reduced triglyceride levels and elevated HDL and cholesterol ratios of terazosin.(ABSTRACT TRUNCATED AT 250 WORDS)

A relative cost-effectiveness analysis of different methods of screening for diabetic retinopathy.

The relative cost and cost-effectiveness of different methods of screening diabetic patients for sight-threatening retinopathy are assessed. The resource costs per screening visit, both to the health service and to patients, of ophthalmoscopic examination by primary screeners including general practitioners, hospital physicians, and ophthalmic opticians are estimated together with those of a similar screening test by ophthalmological clinical assistants. The total resource cost per screen of screening using non-mydriatic photography is also estimated. Using estimates of sensitivity, specificity, and prevalence generated in the screening of 3318 diabetic patients in three UK centres, the relative cost-effectiveness of screening methods is estimated in terms of their cost per true positive case detected. On the assumption that a patient makes a special trip for eye screening, the cost per true positive case detected for primary screeners ranges from 633 pounds for a GP-screened group in one centre to 1079 pounds for another GP-screened group in a second centre; the cost per true positive case detected of photography ranges from 497 pounds for a camera that is taken to general practices in one centre to 1546 pounds for a hospital-based camera. Relative cost-effectiveness changes if, in some contexts, the screening can take place without requiring an additional patient visit, and is strongly related to the relative sensitivity of the screening methods and to the prior probability (prevalence or incidence) of retinopathy in the diabetic population.

Screening for treatable diabetic retinopathy: a comparison of different methods.

The results of the screening of 3318 diabetic patients for sight-threatening diabetic retinopathy in three UK centres are reported. The aims of the study were to determine the extent of diabetic retinopathy in the screened population and to assess the relative effectiveness of different screening methods in appropriately referring cases from a diabetic population, in a context very close to a routine clinical service. Patients were assessed by ophthalmoscopic examination by an ophthalmological clinical assistant. The clinical assistants' referral grades formed the reference standard against which to assess the effectiveness of other screening methods including ophthalmoscopy by primary screeners who were general practitioners (GPs), ophthalmic opticians and hospital physicians, and the assessment by consultant ophthalmologists of non-mydriatic Polaroid fundus photography. The performance of primary screeners based on ophthalmoscopy ranged from a sensitivity of 0.41, with a specificity of 0.89, for one of the GP groups, to a sensitivity of 0.67, with a specificity of 0.96, for the hospital physician group. The performance of the non-mydriatic camera ranged from a sensitivity of 0.35, with a specificity of 0.95, to a sensitivity of 0.67, with a specificity of 0.98.

A comparative study of prostacyclin infusion given before and during cardiopulmonary bypass to assess the first pass effect of the circuit on platelet number and function.

Platelet damage during cardiopulmonary bypass (CPB), although proportional to the duration of bypass, may result in significant dysfunction after the initial contact with an extracorporeal circuit, the so-called 'first pass' phenomenon. The platelet sparing effect of prostacyclin (PGI2) infusion was studied in a double-blind randomized trial on male patients undergoing coronary artery bypass grafts to assess the effect of the 'first pass' through the CPB circuit. Prostacyclin infusion was begun before the onset of CPB or during CPB in two groups which were compared to a placebo control group. A standardized anaesthetic, surgical and perfusion technique were used. Preoperatively and during surgery at pre-set intervals, whole blood platelet aggregation was studied using ADP and collagen agonists. Platelet numbers and function measured by ADP aggregation were conserved in the two PGI2 groups. There was no significant difference between the treated groups. We conclude, therefore, that the initial contact of platelets with the CPB circuit, in the absence of PGI2 did not irreversibly affect platelet function. In addition, the hypotensive action of PGI2 was easier to control once on bypass. It may therefore be preferable to delay PGI2 infusion until CPB has been established.

Comparison of cadmium cytotoxicity in human versus rat nasal epithelial cells in vitro.

Differences in the sensitivity of human and Fischer 344 rat tissues to cadmium sulphate (CdSO(4)) toxicity were investigated in an in vitro model using human and rat nasal turbinate epithelial (NTE) cells. Both rat and human NTE cells were obtained from fresh, normal tissue. Methods were developed for isolating and culturing NTE cells from rat and human tissue using identical procedures, and for measuring the cellular nucleotides by high-performance liquid chromatography. Changes in adenylate energy charge and nucleotide levels were used as toxicity endpoints. Cellular Cd levels were measured by graphite-furnace atomic absorption spectrometry and expressed per unit DNA. Cd uptake was significantly greater in human NTE cells than in rat cells, particularly at the highest exposure concentration (4.8 mm-CdSO(4)). The effects of CdSO(4) on the adenylate energy charge of human and rat NTE cells were similar except at high exposure concentrations and after long exposure times; after a 2-4-hr exposure to 4.8 mm CdSO(4) the adenylate energy charge of human cells was significantly less than that of the rat cells.

Alpha-1 blockers. A new generation of antihypertensive agents.

Alpha-1 blockers have certain disadvantages over conventional and antihypertensive therapies in their haemodynamic profile and metabolic effects. This paper reviews the development of alpha-blockade, the therapeutic efficacy of prazosin, the prototype alpha-1 blocker, and the rationale for the once-daily antihypertensive compounds, terazosin and doxazosin. These drugs offer a useful alternative to first- or second-line therapy in suitable hypertensive patients, particularly with their potentially beneficial effects on serum lipids.

Pharmacokinetics of single and multiple doses of flusoxolol (Ro 31-1411) in healthy subjects.

Flusoxolol (Ro 31-1411) is the pharmacologically active optical isomer of Ro 31-1118, a potent cardioselective beta-adrenoceptor antagonist with partial agonist activity. It was given to 6 healthy volunteers in a single dose, 40 mg, and then in multiple doses, 40 mg daily for 8 days. Plasma concentration data were best described by a linear two-compartment pharmacokinetic model with first order absorption, and the results confirmed linear kinetics. Pharmacokinetic data for flusoxolol were comparable to those for the racemate Ro 31-1118.

Regulatory proteins of the myocardium. Atrial and ventricular tropomyosin and troponin-I in the developing and adult bovine and human heart.

The myofibrillar regulatory proteins, troponin-I and tropomyosin were isolated from human and bovine atria and ventricles and studied in the adult and during foetal development. A number of analytical electrophoretic procedures were employed to detect possible atrial, ventricular or foetal specific forms of these proteins. Biological activity of the regulatory protein complex during development was monitored by measuring the calcium sensitivity of the myofibrillar Mg2+ activated ATPase. No evidence was obtained for unique or foetal specific forms of either troponin I or the alpha and beta subunits of tropomyosin. Although the atria and ventricles possess markedly different contractile properties, no difference was observed between the two chambers at any developmental stage in the relative amounts of alpha and beta tropomyosin present. However, the relative amount of beta tropomyosin increased by 50% in both atria and ventricles during the transition from foetus to adult. A strong inverse correlation existed (r = -0.92) between beta tropomyosin content and heart rate in different species and at different developmental stages in both cardiac chambers. The relative invariance of tropomyosin and troponin I forms in the myocardium was reflected in the high and constant level of calcium sensitivity of myofibrillar Mg2+ ATPase retained in the atria and ventricles throughout development. The implications of these results in relation to control of cardiac contraction are discussed.