In vitro and in vivo effect of levopraziquantel, dextropraziquantel versus racemic praziquantel on different developmental stages of Schistosoma japonicum.

AIM: To compare the antischistosomal effect of racemic praziquantel (Pra) and its enantiomers, levopraziquantel (L-Pra) and dextropraziquantel (D-Pra), on different developmental stages of Schistosoma japonicum. METHODS: The in vitro effects of the drugs were determined in different stages of schistosomes maintained in RPMI 1640 supplemented with 20% calf serum. In vivo study mice infected with schistosome cercariae were treated intragastrically (ig) with Pra, L-Pra or D-Pra at different intervals after infection. The efficacy of the drugs was evaluated by residual mean worm number. RESULTS: Based on the degree of tegument damage induced by L-Pra, d28 and d35 schistosomes were most susceptible to L-Pra, while d14 schistosomules being least susceptible. At comparable concentrations of 0.1-1 g/ml, L-Pra was more active than Pra even when the concentration of L-Pra was reduced to one-half of the minimum effective concentration of Pra. At above-mentioned concentrations D-Pra exhibited no apparent in vitro effect on different stages of schistosomes. When infected mice were treated ig with L-Pra, Pra or D-Pra at a single dose of 300 mg/kg or 500 mg/kg, only the former two drugs showed apparent effect on d0, d21, d28 and d35 schistosomes and less or much less effect on d3, d7 and d14 schistosomules. D-Pra only exhibited a negligible effect on d35 adult schistosomes as compared with L-Pra and Pra. When mice infected with d35 adult schistosmes were treated ig with L-Pra 150 mg/kg, the efficacy was similar to that of mice treated with Pra 300 mg/kg. CONCLUSION: L-Pra is the principal active component against schistosomes in racemic Pra.

Tegument changes of Schistosoma japonicum and Schistosoma mansoni in mice treated with artemether.

AIM: To study the effect of artemether (Art) on the tegument of schistosomes. METHODS: Mice infected with S japonicum cercariae for 7 and 35 d, or with S mansoni cercariae for 49 d were treated intragastrically with Art 200-300 mg.kg-1.d-1 for 2 d. Schistosomes were collected in groups of 2 mice at various intervals after medication for scanning electron microscopic observation. RESULTS: The tegumental changes induced by Art appeared to be similar in S japonicum and S mansoni: swelling and fusion of tegumental surfaces, vesicle formation and collapse of discoid-like sensory structures. In S japonicum the emergence of tegumental alterations was earlier in 7-d-old schistosomulae than that in 35-d-old adult worms. CONCLUSION: Art injured the teguments of S japonicum and S mansoni.

Concentration time course of praziquantel in Filipinos with mild Schistosoma japonicum infection.

Despite extensive use of praziquantel, the current drug of choice for the treatment of schistosomiasis and other helminthic infections, little information is available about its pharmacokinetics in individuals living in geographic areas where such infections are endemic. We investigated the pharmacokinetics of praziquantel by determining its serum concentration-time course in four selected Filipino volunteers with mild Schistosoma japonicum infection who lived in an endemic area in the Southern Philippines. At specified intervals during a 24-hour time period after a single oral dose of praziquantel (25 mg/kg BW), intravenous samples of blood were drawn, processed and analyzed for praziquantel using reverse phase high pressure liquid chromatography. The same study was repeated one week later to assess pharmacokinetic reproducibility. A third study, simulating current field practice, consisted of dosing the patient four hours apart and analyzing for praziquantel in serial blood samples drawn at specified time intervals after the first and second dose. The following results were obtained: 1) Serum concentration-time course of praziquantel was reproducible for each patient but varied from patient to patient. 2) Praziquantel was rapidly absorbed in the gastrointestinal tract as measurable amounts appeared in the blood as early as 15 minutes after dosing. Time to peak serum concentration ranged from 1.50 to 6.00 hours with almost complete elimination from blood by 24 hours whether it was administered as a single dose (1 x 25 mg/kg BW) or as a twice a day dose (2 x 25 mg/kg BW) 4 hours apart. Half-life values ranged from 1.00 to 2.50 hours.(ABSTRACT TRUNCATED AT 250 WORDS)

Uptake and effect of praziquantel and the major human oxidative metabolite, 4-hydroxypraziquantel, by Schistosoma japonicum.

After exposure to praziquantel in vitro at a concentration of 1 microgram/ml for 0.5-2 hr, amounts of praziquantel in Schistosoma japonicum varied from 2.1 +/- 1.2 to 3.7 +/- 1.6 ng/male worm and 1.3 +/- 1.2 to 2.2 +/- 1.5 ng/female worm during the time studied. At 30 micrograms/ml, praziquantel amounts were 11-33-fold higher. However, within 2 hr after removal from a medium containing 30 micrograms/ml praziquantel, 95% of the drug was released from the parasites. When S. japonicum worm pairs were incubated in vitro with 1, 10, and 30 micrograms/ml of 4-hydroxypraziquantel, the major human oxidative metabolite of praziquantel, 0.2 +/- 0.2, 3.8 +/- 1.3, and 7.4 +/- 1.3 ng/worm pair, respectively, were found after a 2-hr incubation. 15-30-fold lower than corresponding worm pair amounts of praziquantel. In vivo, when 4- or 5-wk S. japonicum-infected mice were treated orally with praziquantel (300 mg/kg), peak concentrations of praziquantel in plasma determined by high pressure liquid chromatography were 14.7 +/- 1.5 micrograms/ml (4-wk infection) and 16.7 +/- 2.8 micrograms/ml (5-wk infection) 15 min after treatment. Corresponding in vivo worm praziquantel amounts were 1.8 +/- 0.4 ng/male worm and 2.4 +/- 1.1 ng/female worm, respectively, in the 4-wk infection and 4.6 +/- 1.6 ng/male worm and 5.6 +/- 1.2 ng/female worm in the 5-wk infection. Peak plasma concentrations of 4-hydroxypraziquantel were similar but corresponding in vivo worm amounts were 1-20-fold lower, depending on the time after drug administration.(ABSTRACT TRUNCATED AT 250 WORDS)

Formation of N-(5-nitro-2-thiazolyl)-N'-carboxymethylurea from 5-hydroxyniridazole. Role of aldehyde dehydrogenase in the oxidative metabolism of niridazole.

N-(5-nitro-2-thiazolyl)-N'-carboxymethylurea (NTCU) has been identified as a urinary metabolite of the antischistosomal drug niridazole [1-(5-nitro-2-thiazolyl)-2-imidazolidinone]. When DBA/2J mice were treated with [14C]niridazole, a metabolite comprising 12-14% of the total radioactivity in 24-hr urine samples was resolved by HPLC. The compound was subsequently isolated from pooled urine of niridazole-treated patients. It was identified as NTCU by mass spectrometry, and the deduced structure was confirmed by chemical synthesis. NTCU is unique among known niridazole metabolites, because it lacks an intact imidazolidinone ring. Its structure allows for a ketoenol tautomerism in which the enolate is stabilized by conjugation with the nitrothiazole ring, as evidenced by a pH-dependent 80-nm red shift in the absorption spectrum. We hypothesized that NTCU arises via oxidation of an acyclic aldehyde tautomer of 5-hydroxyniridazole, one of two proximate oxidative niridazole metabolites. Indirect evidence for the aldehyde tautomer included the fact that 5-hydroxyniridazole displayed the same pH-dependent spectral shift as NTCU with a single isobestic point at 388 nm. The proposed precursor-product relationship was confirmed when we found that NTCU formation from 5-hydroxyniridazole was catalyzed by NAD(+)-dependent aldehyde dehydrogenase (EC 1.2.1.3). The activity copurified with benzaldehyde dehydrogenase activity from mouse liver cytosol. Furthermore, benzaldehyde was a competitive inhibitor of 5-hydroxyniridazole dehydrogenase activity. These results demonstrate that 5hydroxyniridazole is not an end product of niridazole metabolism. Because biotransformation of niridazole to its 4- and 5-hydroxy derivatives has been implicated in the drug's carcinogenicity and central nervous system toxicity, NTCU formation appears to represent a detoxication pathway in mammals.

In vitro and in vivo studies of the effect of artemether on Schistosoma mansoni.

To determine whether artemether, a derivative of the antimalarial agent qinghaosu, is therapeutically active against Schistosoma mansoni, we determined the in vitro, in vivo, and histopathologic effects of the drug on S. mansoni worms. In vitro, toxic effects of artemether on S. mansoni were not seen at concentrations of less than 100 micrograms/ml. However, in vivo, 30 and 50% reductions in the lengths of male and female worms, respectively, were observed 14 days after treatment. By 56 days worm dimensions had returned to control values. Similar reversible effects on male testes and female ovaries were seen. In vivo, a single oral dose of artemether (300 mg/kg) induced a shift of worms towards the liver within 8 h after treatment. By 3 and 14 days after treatment, 99 and 76%, respectively, of worms were still in the liver. In vivo, the therapeutic effect of artemether on adult S. mansoni treated on day 56 after infection was modest. Doses as high as 1,200 mg (200 mg/kg per day, six doses) resulted in a worm reduction rate of only 39%. However, in infected mice treated on day 14 or 21 after infection, worm reduction rates of 83 to 98% were obtained. Thus, artemether exhibited modest in vitro and in vivo activities against adult S. mansoni but was twofold more active against 2- to 3-week-old liver-stage parasites.

Clinical use of praziquantel in China.

Praziquantel, a broad-spectrum anthelmintic developed by E. Merck and Bayer AG in Germany in the early 1970s, was synthesized in the People's Republic of China in 1977 and given the Chinese name Pyquiton. After a series of pharmacological and toxicological studies in China, praziquantel was released for clinical use in 1978. The drug is now available for treatment of human schistosome and other trematode infections as well as the treatment of cestode infections including cysticercosis. However, much of the clinical work has been published in Chinese journals that may not be universally familiar. This article summarizes some of the key aspects of these reports, dealing with the clinical use of praziquantel against schistosomiasis japonica and other helminth infections in China.

Streptococcus mitis. A cause of serious infection in adults.

Twenty strains of Streptococcus mitis were isolated from blood or body fluids at the Cleveland Veterans Administration Medical Center from Jan 1, 1981, to April 30, 1984. Fifteen (75%) isolates were considered contaminants. Five (25%) were clinically important and associated with a serious infection of the oropharynx or gastrointestinal tract (three of five), endovascular system (one of five), or a prosthetic hip. Four of five patients required surgical intervention for treatment. Two of five died; one death was directly attributable to S mitis infection. Eighteen strains were available for detailed bacteriologic study. Three strains had a minimum inhibitory concentration of greater than 0.1 micrograms/mL of penicillin and six other strains were tolerant to penicillin. This review suggests that S mitis can be an important pathogen in adults and may cause infections other than endocarditis.

Effects of praziquantel on different developmental stages of Schistosoma mansoni in vitro and in vivo.

To discern whether stage-specific resistance of Schistosoma mansoni to praziquantel occurs in vitro, we determined minimal effective concentrations (MECs) of drug needed to increase motor activity, produce contraction and/or paralysis, and cause tegumental vesiculation of developmental stages from day 0 to day 42 of S. mansoni. Recovery of these stages from exposure to praziquantel in vitro was also evaluated. MECs of praziquantel inducing increased motor activity and muscular contraction or paralysis or both were 0.005-0.01 micrograms/ml, irrespective of the stage examined. However, day-3 lung forms were more resistant than other stages when either drug-induced tegumental vesiculation (MEC, 1 microgram/ml) or recovery from drug exposure was tested. Three-day infections with S. mansoni in CF1 mice were also less responsive to praziquantel treatment than were infections of shorter or longer duration. The concentrations of praziquantel and periods of drug exposure causing gross tegumental damage to S. mansoni in vitro correlated with the peak serum levels and time course of unchanged praziquantel associated with reduction of worm burden in vivo. Thus, stage-specific resistance of S. mansoni to praziquantel does occur in vitro and correlates better with the tegumental than the muscular action of the drug.

Concentration-time course of niridazole and six metabolites in the serum of four Filipinos with Schistosoma japonicum infection.

Niridazole and six of its metabolites have been quantitated by high-pressure liquid chromatography in sera of four male Filipino patients with mild Schistosoma japonicum infections given single oral doses of niridazole (15 mg/kg) on two occasions 10 days apart. Of the five oxidative metabolites measured, 4-hydroxyniridazole and 4-ketoniridazole achieved the highest concentrations, reaching peak values of 0.9 +/- 0.3 microgram/ml of serum (mean +/- S.D., n = 4) and 0.7 +/- 0.1 microgram/ml of serum within 1 to 4 hr. 4-Ketoniridazole achieved peak serum levels 1 hr after the other oxidative metabolites in three of four patients and was the predominant metabolite in the serum of all patients 6 to 10 hr after dosing. By 24 hr, both 4-ketoniridazole and 4-hydroxyniridazole had largely disappeared from the serum. Niridazole and three other oxidative metabolites, 4,5-dihydroxyniridazole, 5-hydroxyniridazole and 4,5-dehydroniridazole, appeared within 1 hr in serum but failed to exceed 0.4 microgram/ml; none of these compounds were detected in the 24-hr serum samples. The pharmacokinetic pattern of niridazole and the oxidative metabolites showed marked interindividual variation but was quite reproducible in the same individual studied 10 days later. 1-Thiocarbamoyl-2-imidazolidinone was analyzed in serum samples by a different high-pressure liquid chromatographic procedure. This reductive metabolite attained maximal levels of 50 to 150 ng/ml of serum 6 to 12 hr after drug administration and remained at 40% or more of its peak concentration even after 24 hr.(ABSTRACT TRUNCATED AT 250 WORDS)

4-Keto niridazole: a major niridazole metabolite with central nervous system toxicity different than niridazole.

4-Keto niridazole, isolated by high-pressure liquid chromatography, was identified by high resolution electron impact mass spectral analysis as a major drug metabolite of niridazole in the serum or plasma of rats and mice treated orally or i.p. with niridazole. This metabolite has a pKa of 5.8 and is approximately 40% bound at physiologic pH to serum proteins of mice receiving therapeutic doses of niridazole. After i.p. injection of niridazole (160 mg/kg), peak serum levels of 4-keto niridazole (10.4 micrograms/ml) were reached within 6 hr in DBA/2J mice. The acute LD50 for 4-keto niridazole i.p. was 55 mg/kg in DBA/2J mice and 51 mg/kg in C57BL/6J mice; the comparable value for niridazole was 220 mg/kg in DBA/2J mice. Signs of acute 4-keto niridazole toxicity were different from those of niridazole toxicity and consisted of profound sedation and labored, irregular breathing terminating in respiratory arrest. Daily i.p. injection of 30 mg/kg of 4-keto niridazole for 5 days into DBA/2J mice resulted in no evidence of cumulative toxicity. The serum and brain concentrations of 4-keto niridazole after a 70-mg/kg i.p. LD90 dose of this compound were 93 micrograms/ml and 7.5 micrograms/g just before death. If an LD90 dose of niridazole (285 mg/kg) was injected into DBA/2J mice, the serum and brain concentrations of 4-keto niridazole just before death were 15 and 5%, respectively, of those found after an LD90 dose of 4-keto niridazole. Thus, 4-keto niridazole does not appear to account for the central nervous system toxicity of niridazole.

1-thiocarbamoyl-2-imidazolidinone, a metabolite of niridazole in Schistosoma mansoni.

Niridazole, a nitro heterocyclic antischistosomal drug, is extensively metabolized to unknown metabolites by Schistosoma mansoni. We report that 1-thiocarbamoyl-2-imidazolidinone was isolated by high pressure liquid chromatography and identified by high resolution electron impact mass spectroscopy as a niridazole metabolite in schistosomes. After a 20-h in vitro incubation in 30 ml of medium containing 10 micrograms ml-1 [14C]niridazole (5.2 Ci mol-1), 100 S. mansoni worm pairs contained approximately 275 ng of 1-thiocarbamoyl-2-imidazolidinone. This amount represented 4% of the total metabolized fraction of niridazole in the parasite. Incubation of schistosomes with 1-thiocarbamoyl-2-[2 14C]imidazolidinone (2.7 Ci mol-1) indicated that this metabolite was not taken up. However, schistosomes released an average of 44 ng ml-1 or 1% of the total 1-thiocarbamoyl-2-imidazolidinone found in the worm back into 1 ml of medium during incubation. No host oxidative metabolites of niridazole were found in the parasites.

Reductive metabolism of niridazole by adult Schistosoma mansoni. Correlation with covalent drug binding to parasite macromolecules.

Niridazole, an antischistosomal nitrothiazole derivative, is metabolized by adult Schistosoma mansoni to one or more reactive intermediates, as evidenced by extensive covalent binding of [14C]niridazole to parasite macromolecules. When worm pairs were incubated for 16 hr in culture medium containing 70 microM [14C]niridazole, 26-34% of the total parasite-associated radioactivity was irreversibly bound to trichloroacetic acid-precipitable material. Drug binding was both time- and [14C]niridazole concentration-dependent. Of the bound drug fraction, 85-90% was associated with parasite proteins, 3-5% with RNA and 4-7% with DNA. When schistosomes were recovered from infected mice, treated with periodic doses of [14C]niridazole, over 40% of the total parasite-associated radioactivity was bound to macromolecules. Niridazole caused up to a 40% decrease in the concentration of total nonprotein thiols in intact schistosomes incubated with the drug over an 8-hr period. Under strictly anaerobic conditions, cell-free schistosome preparations catalyzed a reduced pyridine nucleotide-dependent reduction of niridazole's essential nitro group, as evidenced by disappearance of absorption at 400 nm. Net nitroreduction did not occur under aerobic conditions, although the drug did stimulate oxidation of the pyridine nucleotide cofactor. Covalent binding of [14C]niridazole also took place in this cell-free system, with requirements identical with those needed for enzymatic nitroreduction. Covalent drug binding, but not nitroreduction, was inhibited up to 80-85% by 2 mM L-cysteine, N-acetyl-L-cysteine, or glutathione; S-carboxymethyl-L-cysteine, which has no free sulfhydryl group, was not inhibitory. [14C]4'-Methylniridazole, a nonschistosomicidal analogue of niridazole, was taken up by intact schistosomes in vitro, but was not metabolized and did not bind covalently to parasite macromolecules. Furthermore, 4'-methylniridazole did not affect the concentration of nonprotein thiols in intact parasites and did not serve as a substrate for schistosomal nitroreductase in vitro. These results indicate a positive correlation between proximal metabolic activation of niridazole within these facultative anaerobic organisms and its antiparasitic activity.

Quantitative determination of praziquantel in serum by high-performance liquid chromatography.

A simple method for the quantitation of praziquantel by high-performance liquid chromatography is described. This method has a lower limit of sensitivity of 2.5 ng of drug per ml of human serum and a relative standard deviation of 2.6% at concentrations of 5 ng/ml.

Cercaria-induced histamine release: a factor in the pathogenesis of schistosome dermatitis?

Cercariae, but not schistosomules, of Schistosoma mansoni are capable of activating the release of histamine from rat peritoneal mast cells in vitro. This activation occurs in the absence of serum and does not require adherence to mast cells. The amount of histamine released is proportional to the number of cercariae present. This mast-cell triggering factor from cercariae remains to be characterized biochemically, but may well play a causal role in the "swimmer's itch" associated with primary schistosome exposure.