An Overview on Target-Based Drug Design against Kinetoplastid Protozoan Infections: Human African Trypanosomiasis, Chagas Disease and Leishmaniases.

The protozoan diseases Human African Trypanosomiasis (HAT), Chagas disease (CD), and leishmaniases span worldwide and therefore their impact is a universal concern. The present regimen against kinetoplastid protozoan infections is poor and insufficient. Target-based design expands the horizon of drug design and development and offers novel chemical entities and potential drug candidates to the therapeutic arsenal against the aforementioned neglected diseases. In this review, we report the most promising targets of the main kinetoplastid parasites, as well as their corresponding inhibitors. This overview is part of the Special Issue, entitled "Advances of Medicinal Chemistry against Kinetoplastid Protozoa (Trypanosoma brucei, Trypanosoma cruzi and Leishmania spp.) Infections: Drug Design, Synthesis and Pharmacology".

A systematic review of anti-Entamoeba histolytica activity of medicinal plants published in the last 20 years.

Amoebiasis has emerged as a major health problem worldwide. It is endemic in the present scenario is different and sub-tropical regions especially in Asia, Latin America and also in Africa. Causative of amoebiasis is a protozoan known as Entamoeba histolytica. We screened all the databases such as PubMed, Science Direct, Medline and Google Scholar by using the keywords 'anti-Entamoeba histolytica activity of medicinal plants, anti-Entamoeba histolytica activity of herbal drugs, the anti-amoebic activity of natural drugs'. In the present study, we found 7861 articles, where all articles were screened for bias analysis and included 32 full-matching articles in total reporting the use of medicinal plants as a remedy for amoebiasis. Through these articles, we found 42 herbs having anti-amoebic activity. In bias analysis, we also found four articles under high bias risk. In our study, seven medicinal plants were concluded to possess the most potent anti-amoebic activity based on their IC50 value, which was less than 1 µg mL−1. On bias analysis, we found four articles with high bias risk, hence these studies can be repeated for better results.

Defining Stage-Specific Activity of Potent New Inhibitors of Cryptosporidium parvum Growth In Vitro.

Cryptosporidium parvum and Cryptosporidium hominis have emerged as major enteric pathogens of infants in the developing world, in addition to their known importance in immunocompromised adults. Although there has been recent progress in identifying new small molecules that inhibit Cryptosporidium sp. growth in vitro or in animal models, we lack information about their mechanism of action, potency across the life cycle, and cidal versus static activities. Here, we explored four potent classes of compounds that include inhibitors that likely target phosphatidylinositol 4 kinase (PI4K), phenylalanine-tRNA synthetase (PheRS), and several potent inhibitors with unknown mechanisms of action. We utilized monoclonal antibodies and gene expression probes for staging life cycle development to define the timing of when inhibitors were active during the life cycle of Cryptosporidium parvum grown in vitro These different classes of inhibitors targeted different stages of the life cycle, including compounds that blocked replication (PheRS inhibitors), prevented the segmentation of daughter cells and thus blocked egress (PI4K inhibitors), or affected sexual-stage development (a piperazine compound of unknown mechanism). Long-term cultivation of C. parvum in epithelial cell monolayers derived from intestinal stem cells was used to distinguish between cidal and static activities based on the ability of parasites to recover from treatment. Collectively, these approaches should aid in identifying mechanisms of action and for designing in vivo efficacy studies based on time-dependent concentrations needed to achieve cidal activity.IMPORTANCE Currently, nitazoxanide is the only FDA-approved treatment for cryptosporidiosis; unfortunately, it is ineffective in immunocompromised patients, has varied efficacy in immunocompetent individuals, and is not approved in infants under 1 year of age. Identifying new inhibitors for the treatment of cryptosporidiosis requires standardized and quantifiable in vitro assays for assessing potency, selectivity, timing of activity, and reversibility. Here, we provide new protocols for defining which stages of the life cycle are susceptible to four highly active compound classes that likely inhibit different targets in the parasite. We also utilize a newly developed long-term culture system to define assays for monitoring reversibility as a means of defining cidal activity as a function of concentration and time of treatment. These assays should provide valuable in vitro parameters to establish conditions for efficacious in vivo treatment.

Identification of Phenylphthalazinones as a New Class of Leishmania infantum Inhibitors.

Leishmaniasis is a neglected parasitic disease caused by over 20 different Leishmania species. Current treatments often rely on harsh regimes of pentavalent antimonials such as sodium stibogluconate, while more recent drugs suffer other shortcomings such as low stability and rapid emergence of treatment failure, amongst others. Furthermore, the effectiveness of drugs varies depending on the infecting Leishmania species, thus there is an urgent need for new and effective anti-leishmanial drugs. Screening of an in-house compound library identified the hexahydrophthalazinone NPD-2942 as a low micromolar hit with a pIC50 of 5.8 against L. infantum and a pIC50 of 4.6 for cytotoxicity against human MRC-5 fibroblasts. To derive structure-activity relationships, we modified the cyclohexyl ring of the hexahydrophthalazinone scaffold and 1,2,3-triazoles were attempted as replacement for the pyrazole ring, amongst others. Ultimately, the 2,3-pyrazole-substituted hexahydrophthalazinone NPD-1289 was identified as the most potent analogue in this series with a pIC50 of 6.3, although some cytotoxicity toward MRC-5 cells (pIC50 =5.1) was recorded as well. Replacement of the unsubstituted 2,3-pyrazole with 1,2,3-triazoles led to compounds with lower anti-leishmanial activity. The current scaffold is a valuable new starting point for optimization toward novel anti-leishmanial drugs.

Biopharmaceutical optimization in neglected diseases for paediatric patients by applying the provisional paediatric biopharmaceutical classification system.

AIMS: Unavailability and lack of appropriate, effective and safe formulations are common problems in paediatric therapeutics. Key factors such as swallowing abilities, organoleptic preferences and dosage requirements determine the need for optimization of formulations. The provisional Biopharmaceutics Classification System (BCS) can be used in paediatric formulation design as a risk analysis and optimization tool. The objective of this study was to classify six neglected tropical disease drugs following a provisional paediatric BCS (pBCS) classification adapted to three paediatric subpopulations (neonates, infants and children). METHODS: Albendazole, benznidazole, ivermectin, nifurtimox, praziquantel and proguanil were selected from the 5th edition of the Model List of Essential Medicines for Children from the World Health Organization. Paediatric drug solubility classification was based on dose number calculation. Provisional permeability classification was based on log P comparison versus metoprolol log P value, assuming passive diffusion absorption mechanisms and no changes in passive membrane permeability between paediatric patients and adults. pBCS classes were estimated for each drug, according to different doses and volumes adapted for each age stage and were compared to the adult classification. RESULTS: All six drugs were classified into provisional pBCS in the three paediatric subpopulations. Three drugs maintained the same classification as for adults, ivermectin and benznidazole changed solubility class from low to high in neonates and proguanil changed from low to high solubility in all age stages. CONCLUSION: Provisional pBCS classification of these six drugs shows potential changes in the limiting factors in oral absorption in paediatrics, depending on age stage, compared to the adult population. This valuable information will aid the optimization of paediatric dosing and formulations and can identify bioinequivalence risks when comparing different formulations and paediatric populations.

The in vitro leishmanicidal activity of hexadecylphosphocholine (miltefosine) against four medically relevant Leishmania species of Brazil.

The in vitro leishmanicidal activity of miltefosine® (Zentaris GmbH) was assessed against four medically relevant Leishmania species of Brazil: Leishmania (Leishmania) amazonensis, Leishmania (Viannia) braziliensis, Leishmania (Viannia) guyanensis and Leishmania (Leishmania) chagasi. The activity of miltefosine against these New World species was compared to its activity against the Old World strain, Leishmania (Leishmania) donovani, which is known to be sensitive to the effects of miltefosine. The IC50 and IC90 results suggested the New World species harboured similar in vitro susceptibilities to miltefosine; however, miltefosine was approximately 20 times more active against the Old World L. (L.) donovani than against the New World L. (L.) chagasi species. The selectivity index varied from 17.2-28.9 for the New World Leishmania species and up to 420.0 for L. (L.) donovani. The differences in susceptibility to miltefosine suggest that future clinical trials with this drug should include a laboratory pre-evaluation and a dose-defining step.

Marine sponges: potential sources of new antimicrobial drugs.

Sponges (phylum Porifera) are sessile marine filter feeders that have developed efficient defense mechanisms against foreign attackers such as viruses, bacteria, or eukaryotic organisms. Marine sponges are among the richest sources of pharmacologically-active chemicals from marine organisms. It is suggested that (at least) some of the bioactive secondary metabolites isolated from sponges are produced by functional enzyme clusters, which originated from the sponges and their associated microorganisms. More than 5,300 different products are known from sponges and their associated microorganisms, and more than 200 new metabolites from sponges are reported each year. As infectious microorganisms evolve and develop resistance to existing pharmaceuticals, the marine sponge provides novel leads against bacterial, viral, fungal and parasitic diseases. Many marine natural products have successfully advanced to the late stages of clinical trials, as for example ara-A (vidarabine), an anti-viral drug used against the herpes simplex encephalitis virus. This substance is in clinical use for many years. Moreover, a growing number of candidates have been selected as promising leads for extended preclinical assessment, including manzamine A (activity against malaria, tuberculosis, HIV, and others), lasonolides (antifungal activity) and psammaplin A (antibacterial activity). In this review we have surveyed the discoveries of products derived from marine sponges and associated bacteria that have shown in vivo efficacy or potent in vitro activity against infectious and parasitic diseases, including bacterial, viral, fungal and protozoan infections. Our objective was to highlight the substances that have the greatest potential to lead to clinically useful treatments.

Synthesis and biological evaluation of novel 4-substituted 1-{[4-(10,15,20-triphenylporphyrin-5-yl)phenyl]methylidene}thiosemicarbazides as new class of potential antiprotozoal agents.

A novel series of 4-substituted 1-{[4-(10,15,20-triphenylporphyrin-5-yl)phenyl]methylidene}thiosemicarbazide, 4a-4n, was synthesized in 9-21% yield by the condensation of 4-(10,15,20-triphenylporphyrin-5-yl)benzaldehyde (3) with various substituted thiosemicarbazides in presence of catalytic amount of AcOH. These compounds were assayed for in vitro antiamoebic activity, and the results showed that out of 14 compounds 9 were found with IC(50) values lower than metronidazole corresponding to 1.05- to 4.7-fold increase in activity. MTT Assay showed that all the compounds are nontoxic to human kidney epithelial cell line. 4-(m-Toluidinyl)-1-{[4-(10,15,20-triphenylporphyrin-5-yl)phenyl]methylidene}thiosemicarbazide (4h) showed the highest antiamoebic activity with least cytotoxicity. Some of the compounds were screened for their antimalarial activities and ability to inhibit beta-haematin formation, but none of them showed an activity better than chloroquine and quinine. Only one compound out of six showed an activity comparable to standard drug.

Dihydropyrido[2,3-d]pyrimidines as a new class of antileishmanial agents.

A series of dihydropyrido[2,3-d]pyrimidines have been synthesized and screened for its in vitro antileishmanial activity profile in promastigote and amastigote models. Compounds 2a-2l have shown 83-100% inhibition against promastigotes and 79-100% inhibition against amastigotes at a concentration of 50 microg/mL.

[Canine leishmaniasis: evolution of the chemotherapeutic protocols]. / Evoluzione dei protocolli terapeutici in corso di leishmaniosi canina.

Dogs are the domestic reservoir for Leishmania infantum (syn.: L. chagasi), the parasite causing zoonotic visceral leishmaniasis (ZVL) in both the Old and New Worlds. In foci of canine leishmaniasis (CanL), symptomatic disease occurs in less than 50% of infected dogs, and is characterized by chronic evolution of viscero-cutaneous signs. Among strategies recommended to control ZVL, detection and drug treatment of infected dogs are usually employed in the endemic countries of southern Europe. However, the conventional antileishmanial drugs successfully used in human therapy, such as pentavalent antimonials, amphotericin B, pentamidine or miltefosine, have low efficacy in the treatment of CanL. In dogs, these drugs induce only temporary remission of clinical signs, do not prevent occurrence of relapses, and often cause severe side effects. Leishmaniotic dogs may be classified into 4 groups: 1) Asymptomatic resistant dogs ("contacted dogs"), 2) Asymptomatic dogs (preclinical), 3) Dogs with minimal signs of leishmaniasis (oligosymptomatic dogs? Chronic form of leishmaniasis?), 4) Dogs suffering from different forms of clinical leishmaniasis (symptomatic dogs). The dog's immunological status and the associated clinical signs may influence the efficacy of antileishmanial drugs. Subjects belonging to groups 2, 3 and 4 should be always treated, in order to reduce their parasite load. Parameters that must be considered before starting the antileishmanial treatment are hemogram, renal and hepatic functions, electrophoretic protein pattern, antileishmania antibody titres, and bone marrow and lymph node parasite load. The most common antileishmanial drugs currently used in Italy to treat CanL are pentavalent antimonials (meglumine antimoniate) and allopurinol, alone or in combination. Other used drugs are aminosidine (syn.: paromomycin), pentamidine, metronidazole and spyramicin. Each drug regimen has different duration, from a few weeks (aminosidine), to a few months (meglumine antimoniate) or several months (allopurinol). One of the most recent drug used in human VL is liposomal amphotericin B (AmBisome--L-AMB), a powerful antileishmanial drug in both experimental murine models and in VL patients. In Italy, L-AMB is now considered the drug of choice for the treatment of human cases. However, in HIV co-infected patients high doses of L-AMB are ineffective in obtaining a radical cure. In dogs, L-AMB treatment rapidly leads to clinical recovery but is uneffective to eliminate the parasites. Drugs containing amphotericin B should not be used in veterinary practice in order to avoid selection of parasites resistant to the drug, as it already occurred for the pentavalent antimonials. Currently, there is not a standard protocol for CanL treatment in Italy, as there is an extreme variability of proposed dosages. Clinical studies on immunotherapeutics and new antileishmanial drugs, such as miltefosine and its derivates, are in progress.

Anilino-(2-bromophenyl) acetonitrile: a promising orally effective antileishmanial agent.

Visceral leishmaniasis (VL) or kala-azar is a worldwide disseminated intracellular infection caused by the hemoflagellate protozoan parasites Leishmania donovani. Chemotherapeutic scenario presents a deplorable picture and demands an urgent search for a new and safe anti-VL drugs, preferably active by oral route. In search of new antileishmanial agents, a total of 16 compounds belonging to the anilino-(substituted phenyl)-acetonitrile class were tested in vitro in promastigote/macrophase-amastigote systems and in vivo in L. donvoani/hamster model for their antileishmanial activity. Compound 3, anilino-(2-bromophenyl)-acetonitrile, exhibited most promising activity both in vitro at a concentration of 100 microg/ml (82.33 and 94.36% in promastigote and macrophase-amastigote systems, respectively) and in vivo at a dose of 50 mg/kg for 5 days (82.11 and 80% by i.p. and p.o. routes, respectively), hence this compound was investigated in detail. To maximize its bioavailability, dissolution profile, absorption, the compound was also tested in vivo as its soluble form. But no enhancement in activity was observed. From the results of different parameters for example ED(50) and LD(50) etc. compound 3 appears to be a potent orally effective compound which could further be investigated to establish its potential as a candidate molecule of antileishmanial therapy.

Chemotherapeutic approaches to protozoa: Coccidiae--current level of knowledge and outlook.

Progress in the treatment and prophylaxis of cyst-forming coccidial infections (Neospora, Sarcocystis Toxoplasma) and Cryptosporidium infections has been limited (Table 1: Haberkorn 1996: Croft 1997: Wang 1997). However, new possibilities have been opened up in the treatment of Eimeria-induced coccidiosis in poultry and mammals. due to improvements in treatment and, or metaphylaxis. A new polyether antibiotic. semduramycin, has recently been added to the range of effective prophylactic preparations. The development of resistance to anticoccidial agents is now posing similar problems to those encountered with malaria, coccidiosis in poultry being particularly affected. Because no new active ingredient from a new family of chemical substances has been developed for more than 10 years, the following approaches are being adopted to get round this problem: the use of older preparations which have not been used for a long time, the introduction of combinations such as narasin nicarbazin or methyl benzoquate clopidol and the alternating use of anticoccidial agents in rotation and shuttle programmes. The goal of a real alternative, i.e. vaccination, has been achieved to a certain extent in the form of live vaccines for laying hens and broiler breeders and is being practiced in some countries.

Chemotherapeutic approaches to protozoa: Giardia, Trichomonas and Entamoeba--current level of knowledge and outlook.

The situation regarding the treatment of human Giardia and Trichomonas infections and the intestinal and tissue stages of Entamoeba histolytica with metronidazole and other 5-nitroimidazoles is currently satisfactory (Table 1; Mehlhorn 2000). Following correct and rapid diagnosis, the parasites are eliminated reliably and completely. The situation in cases of infection with Acanthamoeba (often involving the eyes) or with Naegleria (often involving the brain) is serious, however. In both cases, there is no drug of choice available. Treatment consists of relieving the symptoms and/or preventing local degeneration.

Molecular search of new active drugs against Toxoplasma gondii.

Molecular connectivity has been applied to the search of new compounds with activity against the protozoan Toxoplasma gondii, using a stepwise linear discriminant analysis (SLDA) which is able to classify a compound according its activity either as active or as inactive. Among the selected compounds, andrographolide and dibenzotiophene sulfone stand out, both with IC50 values lower than 1 microgram/ml, which are comparable to these of drugs such as sulfamethoxazole, pyrimethamine and trimethoprim, with IC50 values equal to 1.1, 0.04 and 2.31 micrograms/ml, respectively. These results confirm the usefulness of our topological approach for the selection and design of new-lead drugs active against Toxoplasma gondii.

[Treatment of infantile visceral leishmaniasis]. / Traitement de la leishmaniose viscérale infantile.

Visceral leishmaniasis is an endemic disease in the Mediterranean Basin. Children are one of the targets of the infection. Treatment usually requires parenteral injections of pentavalent antimony (Glucantime or Pentostam), but the high frequency of adverse events and the occurrence of primary or secondary resistance cases limit the use of these medications. Diamidines (Pentacarinat) or amphotericin B derivatives are alternatives to antimony. Unfortunately, pharmacokinetics and optimal dosage of diamidines are not well-known, and numerous adverse events are described. Liposomal preparations of amphotericin B enhance its efficiency and tolerance, and the duration of treatment may be reduced to 5 days. Moreover, primary resistance to amphotericin B is not described in immunocompetent children. Allopurinol associated with antimony seems no more efficient than antimony alone. Aminosidine is not evaluated.

Tratamiento de las enteroparasitosis / Treatment of the intestinal parasites

La parasitosis intestinal es un grave problema de salud mundial que afecta principalmente a los países subdesarrollados carentes de recursos económicos, sanitarios y educacionales. El Perú no escapa a esta situación por lo que es necesario que los profesionales de la salud tengan conocimientos clínicos, epidemiológicos y terapéuticos básicos para ser aplicados eficientemente en beneficio de la comunidad. Se presenta una revisión actualizada del tratamiento de las enteroparasitosis en un esquema terapéutico y adecuado a nuestra realidad

[Multiparameter analysis approach of antiparasitic structure-activity relationships in function of the parasite taxonomy]. / Approche par analyse multiparamétrique de relations structure-activité antiparasitaire en fonction de la taxonomie des parasites.

In this paper the authors have studied the structure-parasitic activity relationships through the Chemical Abstracts data base which groups together the works published in this field for the last twenty years. With mathematical methods of multidimensional data reduction using a computer it has been possible to validate the classification of human parasites on the base of their sensitivity to chemical families. The specificity of the pharmacochemical relationships was described through two factorial maps. This work shows that it is possible to find new chemical series with potential parasitic activity.