Inhibition of triclabendazole metabolism in vitro by ketoconazole increases disruption to the tegument of a triclabendazole-resistant isolate of Fasciola hepatica

A study has been carried out to investigate whether the action of triclabendazole (TCBZ) against Fasciola hepatica is altered by inhibition of drug metabolism. The cytochrome P450 (CYP 450) enzyme pathway was inhibited using ketoconazole (KTZ) to see whether a TCBZ-resistant isolate could be made more sensitive to TCBZ action. The Oberon TCBZ-resistant and Cullompton TCBZ-susceptible isolates were used for these experiments. The CYP 450 system was inhibited by a 2-h pre-incubation in ketoconazole (40 mu M), then incubated for a further 22 h in NCTC medium containing either KTZ, KTZ+nicotinamide adenine dinucleotide phosphate (NADPH) (1 nM), KTZ+NADPH+TCBZ (15 mu g/ml), or KTZ+NADPH+triclabendazole sulphoxide (TCBZ. SO; 15 mu g/ml). Changes to fluke ultrastructure following drug treatment and metabolic inhibition were assessed using transmission electron microscopy. After treatment with either TCBZ or TCBZ. SO on their own, there was greater disruption to the TCBZ-susceptible than TCBZ-resistant isolate. However, co-incubation with KTZ+TCBZ, but more particularly KTZ+TCBZ. SO, led to more severe changes to the TCBZ-resistant isolate than with each drug on its own: in the syncytium, for example, there was severe swelling of the basal infolds and their associated mucopolysaccharide masses, accompanied by an accumulation of secretory bodies just below the apex. Golgi complexes were greatly reduced or absent in the tegumental cells and the synthesis, production, and transport of secretory bodies were badly disrupted. With the TCBZ-susceptible Cullompton isolate, there was limited potentiation of drug action. The results support the concept of altered drug metabolism in TCBZ-resistant flukes and this process may play a role in the development of drug resistance.

Enhancement of triclabendazole action in vivo against a triclabendazole-resistant isolate of Fasciola hepatica by co-treatment with ketoconazole

An in vivo study in the laboratory rat model was carried out to monitor morphological changes in adult Fasciola hepatica over a 4-day period resulting from combination treatment of triclabendazole (TCBZ) and the metabolic inhibitor, ketoconazole (KTZ). Rats were infected with the TCBZ-resistant Oberon isolate of F. hepatica and divided into 3 groups at 12 weeks post-infection. The first group was dosed orally with TCBZ at a dosage of 10 mg/kg and KTZ at a dosage of 10 mg/kg. Flukes were recovered at 24, 48, 72 and 96h post-treatment (p.t.). A second group of rats was treated with TCBZ alone (10 mg/kg) and sacrificed at 96 h p.t. The third group acted as untreated controls. Surface changes were monitored by scanning electron microscopy (SEM). In flukes from the TCBZ+ KTZ-treated group, the results showed a progressive and time-dependent increase in the level of disruption to the tegumental syncytium. Swelling, furrowing, blebbing and sloughing of the syncytium increased with time p.t. Another feature seen was a thick layer of tegumental shedding in some fluke samples at different times p.t. By comparison, flukes treated with TCBZ alone remained unaffected. The results demonstrated that the Oberon isolate is only sensitive to drug action in the presence of ketoconazole, indicating that combining triclabendazole with a metabolic inhibitor could be used to preserve the effectiveness of the drug against TCBZ-resistant populations of F. hepatica. (C) 2010 Elsevier B.V. All rights reserved.

Potentiation of triclabendazole action in vivo against a triclabendazole-resistant isolate of Fasciola hepatica following its co-administration with the metabolic inhibitor, ketoconazole

An in vivo study in the laboratory rat model has been carried out to monitor morphological changes in adult Fasciola hepatica over a 4-day period resulting from co-treatment with triclabendazole (TCBZ) and ketoconazole (KTZ), a cytochrome P450 inhibitor. Rats were infected with the triclabendazole-resistant Oberon isolate of F. hepatica, dosed orally with triclabendazole at a dosage of 10mg/kg live weight and ketoconazole at a dosage of 10mg/kg live weight. Flukes were recovered at 24, 48, 72 and 96 h post-treatment (p.t.) and changes to fluke ultrastructure were assessed using transmission electron microscopy (TEM). Results showed an increase in the severity of changes to the fluke ultrastructure with time p.t. Swelling of the basal infolds and the associated mucopolysaccharide masses became more severe with time. Golgi complexes, if present, were greatly reduced in size and number by 96 h p.t., and sub-tegumental flooding was seen from the 72 h time-period onwards. Some sloughing of the tegumental covering over the spines was observed at 96 h p.t. The results demonstrated that the Oberon isolate is more sensitive to TCBZ action in the presence of KTZ than to TCBZ alone, reinforcing the idea that altered drug metabolism is involved in the resistance mechanism. Moreover, they support the concept that TCBZ+inhibitor combinations (aimed at altering drug pharmacokinetics and potentiating the action of TCBZ) could be used in the treatment of TCBZ-R populations of F. hepatica.

Fabrication of Potentiometric Sensors for the Selective Determination of Ketoconazole

The fabrication and analytical applications of two types of potentiometric sensors for the determination of ketoconazole (KET) are described. The sensors are based on the use of KET-molybdophosphoric acid (MPA) ion pair as electroactive material. The fabricated sensors include both polymer membrane and carbon paste electrodes. Both sensors showed a linear, stable and near Nernstian slope of 57.8mV=decade and 55.2mV=decade for PVC membrane and carbon paste sensors respectively over a relatively wide range of KET concentration (1×10-2-5×10-5 and 1×10-2-1×10-6). The sensors showed a fast response time of <30 sec and <45 sec. A useful pH range of 3–6 was obtained for both types of sensors. A detection limit of 2.96 10 5M was obtained for PVC membrane sensor and 6.91 10 6M was obtained for carbon paste sensor. The proposed sensors proved to have a good selectivity for KET with respect to a large number of ions. The proposed sensors were successfully applied for the determination of KET in pharmaceutical formulations. The results obtained are in good agreement with the values obtained by the standard method.

The effect of Ketoconazole on experimental paracoccidioidomycosis in the Syrian hamster: immunological and histopathological study

The effect of Ketoconazole (KTZ) on the hamster experimental intratesticular paracoccidioidomycosis was studied employing different treatment schedules. KTZ long course treatment beginning at an early stage of the infection was effective in preventing fungal proliferation, dissemination to lymph nodes, spleen and kidneys, and in maintaining low levels of humoral and cellular specific immune responses. KTZ short course treatment starting at an advanced stage of disease resulted in a more severe histopathological picture without significant changes in the immunological profile. The drug prolonged the life span of hamsters infected with Paracoccidioides brasiliensis, but did not prevent mortality. Toxic necrosis of the bone marrow occurred in normal animals receiving 120 mg/kg/day of KTZ but with lower doses no morphologic alterations were observed in heart, lungs, kidneys, adrenals, spleen, liver, intestine or bone marrow. © 1984 Dr W. Junk Publishers.

Paracoccidioidomycosis: A comparative study of the evolutionary serologic, clinical and radiologic results for patients treated with ketoconazole or amphotericin B plus sulfonamides

A comparative study of two groups of patients with paracoccidioidomycosis was carried out with the objective of comparing the evolutionary serologic, clinical and radiologic results after 6, 12, 15 and 18 months of treatment with ketoconazole (22 patients) or amphotericin B plus sulfonamides (32 patients). The serologic data analyzed as a whole showed a tendency to sharper drops in antibody titers in the patients treated with ketoconazole. Clinically patients treated with ketoconazole fared better but the differences were not statistically significant. No statistical difference was detected between groups in terms of the results of radiologic evolution. © 1985 Martinus Nijhoff/Dr W. Junk Publishers.

Ketoconazole in the treatment of experimental murine paracoccidioidomycosis

In a murine model of chronic disseminated paracoccidioidomycosis (strain 18; intravenous route), Ketoconazole (200 mg/kg in 0.2% agar) was given daily by gavage in three different schedules. Continuous treatment from an early stage of infection (day 3) up to week 20 was the most effective protocol, leading to remission of histopathological lesions and of both humoral and cellular anti-P. brasiliensis immune response, and clearance of the fungus in lungs; only 1 treated animal at week 20 showed pulmonary granulomas, although less extensive than control mice. Continuous treatment from early stage up to week 8, followed by a 16 week-period of drug discontinuity, caused remission of lesions in all but 3 treated mice which showed active pulmonary paracoccidioidomycosis similar to controls (14.2% of unresponsiveness to treatment). The continuous Ketoconazole protocol since a late stage of infection (week 4) up to week 20 produced a slower remission of lesions and immune response when compared with the first drug schedule. In this model of paracoccidioidomycosis, Ketoconazole showed no detectable side-effects and was a very effective drug especially in a prolonged administration protocol from an early stage of infection.

Studies on CYP2C8-mediated drug interactions

Useiden lääkkeiden yhtäaikainen käyttö on nykyään hyvin yleistä, mikä lisää lääkeaineiden haitallisten yhteisvaikutusten riskiä. Lääkeaineiden poistumisessa elimistöstä ovat tärkeässä osassa niitä hajottavat (metaboloivat) maksan sytokromi P450 (CYP) entsyymit. Vasta aivan viime vuosina on havaittu, että CYP2C8-entsyymillä voi olla tärkeä merkitys mm. lääkeaineyhteisvaikutuksissa. Eräät lääkeaineet voivat estää (inhiboida) CYP2C8-entsyymin kautta tapahtuvaa metaboliaa. Tässä työssä selvitettiin CYP2C8-entsyymiä estävien lääkkeiden vaikutusta sellaisten lääkeaineiden pitoisuuksiin, joiden aikaisemman tiedon perusteella arveltiin metaboloituvan CYP2C8-välitteisesti. Näiden lääkeaineiden metaboliaa tutkittiin myös koeputkiolosuhteissa (in vitro -menetelmillä). Lisäksi CYP2C8-entsyymiä estävän lipidilääke gemfibrotsiilin yhteisvaikutusmekanismia tutkittiin selvittämällä interaktion säilymistä koehenkilöillä gemfibrotsiilin annostelun lopettamisen jälkeen. Yhteisvaikutuksia tutkittiin terveillä vapaaehtoisilla koehenkilöillä käyttäen vaihtovuoroista koeasetelmaa. Koehenkilöille annettiin CYP2C8-entsyymiä estävää lääkitystä muutaman päivän ajan ja tämän jälkeen kerta-annos tutkimuslääkettä. Koehenkilöiltä otettiin useita verinäytteitä, joista määritettiin lääkepitoisuudet nestekromatografisilla tai massaspektrometrisillä menetelmillä. Gemfibrotsiili nosti ripulilääke loperamidin pitoisuudet keskimäärin kaksinkertaiseksi. Gemfibrotsiili lisäsi, mutta vain hieman, kipulääke ibuprofeenin pitoisuuksia, eikä sillä ollut mitään vaikutusta unilääke tsopiklonin pitoisuuksiin toisin kuin aiemman kirjallisuuden perusteella oli odotettavissa. Toinen CYP2C8-estäjä, mikrobilääke trimetopriimi, nosti diabeteslääke pioglitatsonin pitoisuuksia keskimäärin noin 40 %. Gemfibrotsiili nosti diabeteslääke repaglinidin pitoisuudet 7-kertaiseksi ja tämä yhteisvaikutus säilyi lähes yhtä voimakkaana vielä 12 tunnin päähän viimeisestä gemfibrotsiiliannoksesta. Tehdyt havainnot ovat käytännön lääkehoidon kannalta merkittäviä ja ne selvittävät CYP2C8-entsyymin merkitystä useiden lääkkeiden metaboliassa. Gemfibrotsiilin tai muiden CYP2C8-entsyymiä estävien lääkkeiden yhteiskäyttö loperamidin kanssa voi lisätä loperamidin tehoa tai haittavaikutuksia. Toisaalta CYP2C8-entsyymin osuus tsopiklonin ja ibuprofeenin metaboliassa näyttää olevan pieni. Trimetopriimi nosti kohtalaisesti pioglitatsonin pitoisuuksia, ja kyseisten lääkkeiden yhteiskäyttö voi lisätä pioglitatsonin annosriippuvaisia haittavaikutuksia. Gemfibrotsiili-repaglinidi-yhteisvaikutuksen päämekanismi in vivo näyttää olevan CYP2C8-entsyymin palautumaton esto. Tämän vuoksi gemfibrotsiilin estovaikutus ja yhteisvaikutusriski säilyvät pitkään gemfibrotsiilin annostelun lopettamisen jälkeen, mikä tulee ottaa huomioon käytettäessä sitä CYP2C8-välitteisesti metaboloituvien lääkkeiden kanssa.

Synthesis, antimicrobial and cytotoxic activities of sulfone linked bis heterocycles

A new class of sulfone linked bis heterocycles viz., pyrrolyl/pyrazolyl arylaminosulfonylmethyl 1,3,4-oxadiazoles, 1,3,4-thiadiazoles, and 1,2,4-triazoles were prepared and tested for antimicrobial activity and cytotoxicity. The chloro-substituted compounds 5c, 8c and 14c showed comparable antibacterial activity to chloramphenicol against Pseudomonasaeruginosa and compound 5c exhibited comparable antifungal activity to ketoconazole against Penicilliumchrysogenum. One of the compounds, vinylsulfonyl oxadiazole showed appreciably cytotoxic activity on A549 lung carcinoma cells with an IC50 at a concentration of 31.7 mu M. (C) 2012 Elsevier Masson SAS. All rights reserved.

High-throughput synergy screening identifies microbial metabolites as combination agents for the treatment of fungal infections

The high mortality rate of immunocompromised patients with fungal infections and the limited availability of highly efficacious and safe agents demand the development of new antifungal therapeutics. To rapidly discover such agents, we developed a high-throughput synergy screening (HTSS) strategy for novel microbial natural products. Specifically, a microbial natural product library was screened for hits that synergize the effect of a low dosage of ketoconazole (KTC) that alone shows little detectable fungicidal activity. Through screening of approximate to 20,000 microbial extracts, 12 hits were identified with broadspectrum antifungal activity. Seven of them showed little cytotoxicity against human hepatoma cells. Fractionation of the active extracts revealed beauvericin (BEA) as the most potent component, because it dramatically synergized KTC activity against diverse fungal pathogens by a checkerboard assay. Significantly, in our immunocompromised mouse model, combinations of BEA (0.5 mg/kg) and KTC (0.5 mg/kg) prolonged survival of the host infected with Candida parapsilosis and reduced fungal colony counts in animal organs including kidneys, lungs, and brains. Such an effect was not achieved even with the high dose of 50 mg/kg KTC. These data support synergism between BEA and KTC and thereby a prospective strategy for antifungal therapy.

Development of Electrochemical Sensors for the Determination of Certain Pharmaceuticals

The quality of a drug is determined after establishing its authenticity by testing its purity and quality of the pure substance in the drug and its formulations.Ion selective sensors have become one of the most effective and powerful means for analytical scientists in the determination of drug substances and are playing an increasing role in pharmaceutical analysis.ISEs are cost effective,easy to prepare and can be rapidly manipulated . Potentiometric ion selective sensors have been developed for six drugs namely trimethoprim, ketoconazole, lamivudine, domperidon, nimesulide and Lomefloxacin. The sensors fabricated include both PVC membrane sensor as well as carbon paste sensor. A total of sixteen different sensors have been developed. The response parameters of all the sensors have been studied and the sensors were applied to the determination of the drugs in pharmaceutical formulations and also in real samples like urine.

Micoses sistémicas

Este trabalho teve como objetivo a análise das micoses sistémicas, no que respeita à sua identificação, morfologia dos agentes patogénicos que lhes dão origem, onde são endémicas, os sintomas que apresentam, os métodos de diagnóstico e as terapêuticas disponíveis para cada patologia. As micoses sistémicas são infeções causadas por fungos patogénicos primários que têm o trato respiratório como porta de entrada, e a partir daí podem disseminar-se por todo o organismo. Os agentes antifúngicos são usados no tratamento destas infeções, em que, no caso das infeções sistémicas, predominam o uso de cetoconazol (polieno), fluconazol e itraconazol (azóis). Existem ainda estudos que demonstram que as vacinas atenuadas podem ser usadas na profilaxia destas infeções. Com este trabalho conclui-se que as infeções fúngicas estão longe de ser extintas, uma vez que se verifica cada vez mais a existência de resistências por parte dos fungos aos fármacos, sendo o diagnóstico e a terapêutica usada dois parâmetros fundamentais para um tratamento eficaz.