RESUMO
Malaria remains a global health concern as drug resistance threatens treatment programs. We identified a piperidine carboxamide (SW042) with anti-malarial activity by phenotypic screening. Selection of SW042-resistant Plasmodium falciparum (Pf) parasites revealed point mutations in the Pf_proteasome ß5 active-site (Pfß5). A potent analog (SW584) showed efficacy in a mouse model of human malaria after oral dosing. SW584 had a low propensity to generate resistance (minimum inoculum for resistance [MIR] >109) and was synergistic with dihydroartemisinin. Pf_proteasome purification was facilitated by His8-tag introduction onto ß7. Inhibition of Pfß5 correlated with parasite killing, without inhibiting human proteasome isoforms or showing cytotoxicity. The Pf_proteasome_SW584 cryoelectron microscopy (cryo-EM) structure showed that SW584 bound non-covalently distal from the catalytic threonine, in an unexplored pocket at the ß5/ß6/ß3 subunit interface that has species differences between Pf and human proteasomes. Identification of a reversible, species selective, orally active series with low resistance propensity provides a path for drugging this essential target.
Assuntos
Antimaláricos , Piperidinas , Plasmodium falciparum , Inibidores de Proteassoma , Piperidinas/química , Piperidinas/farmacologia , Plasmodium falciparum/efeitos dos fármacos , Plasmodium falciparum/enzimologia , Animais , Antimaláricos/farmacologia , Antimaláricos/química , Humanos , Camundongos , Inibidores de Proteassoma/farmacologia , Inibidores de Proteassoma/química , Inibidores de Proteassoma/síntese química , Administração Oral , Complexo de Endopeptidases do Proteassoma/metabolismo , Malária/tratamento farmacológico , Malária/parasitologia , Amidas/química , Amidas/farmacologia , Amidas/síntese química , Malária Falciparum/tratamento farmacológico , Feminino , Estrutura MolecularRESUMO
In addition to vector control, long-lasting insecticidal nets and case management, the prevention of infection through vaccination and/or chemoprevention are playing an increasing role in the drive to eradicate malaria. These preventative approaches represent opportunities for improvement: new drugs may be discovered that target the early infectious stages of the Plasmodium parasite in the liver (rather than the symptomatic, abundant blood stage), and new, exciting vaccination technologies have recently been validated (using mRNA or novel adjuvants). Exploiting these possibilities requires the availability of humanized mouse models that support P. falciparum infection yet avoid the hazardous use of infectious mosquitoes. Here, we show that commercially available P. falciparum sporozoites and FRG mice carrying human hepatocytes and red blood cells faithfully recapitulate the early human malaria disease process, presenting an opportunity to use this model for the evaluation of prophylactic treatments with a novel mode of action.
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The Plasmodium falciparum aspartic protease plasmepsin X (PMX) is essential for the egress of invasive merozoite forms of the parasite. PMX has therefore emerged as a new potential antimalarial target. Building on peptidic amino alcohols originating from a phenotypic screening hit, we have here developed a series of macrocyclic analogues as PMX inhibitors. Incorporation of an extended linker between the S1 phenyl group and S3 amide led to a lead compound that displayed a 10-fold improved PMX inhibitory potency and a 3-fold improved half-life in microsomal stability assays compared to the acyclic analogue. The lead compound was also the most potent of the new macrocyclic compounds in in vitro parasite growth inhibition. Inhibitor 7k cleared blood-stage P. falciparum in a dose-dependent manner when administered orally to infected humanized mice. Consequently, lead compound 7k represents a promising orally bioavailable molecule for further development as a PMX-targeting antimalarial drug.
Assuntos
Antimaláricos , Peptidomiméticos , Camundongos , Animais , Antimaláricos/farmacologia , Antimaláricos/metabolismo , Peptidomiméticos/farmacologia , Peptidomiméticos/metabolismo , Inibidores de Proteases/farmacologia , Inibidores de Proteases/metabolismo , Ácido Aspártico Endopeptidases , Plasmodium falciparum/metabolismo , Proteínas de ProtozoáriosRESUMO
The development of new combinations of antimalarial drugs is urgently needed to prevent the spread of parasites resistant to drugs in clinical use and contribute to the control and eradication of malaria. In this work, we evaluated a standardized humanized mouse model of erythrocyte asexual stages of Plasmodium falciparum (PfalcHuMouse) for the selection of optimal drug combinations. First, we showed that the replication of P. falciparum was robust and highly reproducible in the PfalcHuMouse model by retrospective analysis of historical data. Second, we compared the relative value of parasite clearance from blood, parasite regrowth after suboptimal treatment (recrudescence), and cure as variables of therapeutic response to measure the contributions of partner drugs to combinations in vivo. To address the comparison, we first formalized and validated the day of recrudescence (DoR) as a new variable and found that there was a log-linear relationship with the number of viable parasites per mouse. Then, using historical data on monotherapy and two small cohorts of PfalcHuMice evaluated with ferroquine plus artefenomel or piperaquine plus artefenomel, we found that only measurements of parasite killing (i.e., cure of mice) as a function of drug exposure in blood allowed direct estimation of the individual drug contribution to efficacy by using multivariate statistical modeling and intuitive graphic displays. Overall, the analysis of parasite killing in the PfalcHuMouse model is a unique and robust experimental in vivo tool to inform the selection of optimal combinations by pharmacometric pharmacokinetic and pharmacodynamic (PK/PD) modeling.
Assuntos
Antimaláricos , Malária Falciparum , Animais , Camundongos , Antimaláricos/farmacologia , Antimaláricos/uso terapêutico , Plasmodium falciparum , Estudos Retrospectivos , Peróxidos , Malária Falciparum/tratamento farmacológico , Malária Falciparum/parasitologia , Combinação de MedicamentosRESUMO
Innovative cross-over study designs were explored in non-human primate (NHP) studies to determine the value of this approach for the evaluation of drug efficacy against tuberculosis (TB). Firstly, the pharmacokinetics (PK) of each of the drugs Isoniazid (H), Rifampicin (R), Pyrazinamide (Z) and Ethambutol (E), that are standardly used for the treatment of tuberculosis, was established in the blood of macaques after oral dosing as a monotherapy or in combination. Two studies were conducted to evaluate the pharmacokinetics and pharmacodynamics of different drug combinations using cross-over designs. The first employed a balanced, three-period Pigeon design with an extra period; this ensured that treatment by period interactions and carry-over could be detected comparing the treatments HR, HZ and HRZ using H37Rv as the challenge strain of Mycobacterium tuberculosis (M. tb). Although the design accounted for considerable variability between animals, the three regimens evaluated could not be distinguished using any of the alternative endpoints assessed. However, the degree of pathology achieved using H37Rv in the model during this study was less than expected. Based on these findings, a second experiment using a classical AB/BA design comparing HE with HRZ was conducted using the M. tb Erdman strain. More extensive pathology was observed, and differences in computerized tomography (CT) scores and bacteriology counts in the lungs were detected, although due to the small group sizes, clearer differences were not distinguished. Type 1 T helper (Th1) cell response profiles were characterized using the IFN-γ ELISPOT assay and revealed differences between drug treatments that corresponded to decreases in disease burden. Therefore, the studies performed support the utility of the NHP model for the determination of PK/PD of TB drugs, although further work is required to optimize the use of cross-over study designs.
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Plasmepsin X (PMX) is an essential aspartyl protease controlling malaria parasite egress and invasion of erythrocytes, development of functional liver merozoites (prophylactic activity), and blocking transmission to mosquitoes, making it a potential multistage drug target. We report the optimization of an aspartyl protease binding scaffold and the discovery of potent, orally active PMX inhibitors with in vivo antimalarial efficacy. Incorporation of safety evaluation early in the characterization of PMX inhibitors precluded compounds with a long human half-life (t1/2) to be developed. Optimization focused on improving the off-target safety profile led to the identification of UCB7362 that had an improved in vitro and in vivo safety profile but a shorter predicted human t1/2. UCB7362 is estimated to achieve 9â¯logâ¯10 unit reduction in asexual blood-stage parasites with once-daily dosing of 50 mg for 7 days. This work demonstrates the potential to deliver PMX inhibitors with in vivo efficacy to treat malaria.
Assuntos
Antimaláricos , Antagonistas do Ácido Fólico , Malária , Animais , Humanos , Antimaláricos/farmacologia , Antimaláricos/uso terapêutico , Plasmodium falciparum/metabolismo , Ácido Aspártico Endopeptidases , Malária/tratamento farmacológicoRESUMO
Human and animal malaria parasites increase their host erythrocyte permeability to a broad range of solutes as mediated by parasite-associated ion channels. Molecular and pharmacological studies have implicated an essential role in parasite nutrient acquisition, but inhibitors suitable for development of antimalarial drugs are missing. Here, we generated a potent and specific drug lead using Plasmodium falciparum, a virulent human pathogen, and derivatives of MBX-2366, a nanomolar affinity pyridazinone inhibitor from a high-throughput screen. As this screening hit lacks the bioavailability and stability needed for in vivo efficacy, we synthesized 315 derivatives to optimize drug-like properties, establish target specificity, and retain potent activity against the parasite-induced permeability. Using a robust, iterative pipeline, we generated MBX-4055, a derivative active against divergent human parasite strains. MBX-4055 has improved oral absorption with acceptable in vivo tolerability and pharmacokinetics. It also has no activity against a battery of 35 human channels and receptors and is refractory to acquired resistance during extended in vitro selection. Single-molecule and single-cell patch-clamp indicate direct action on the plasmodial surface anion channel, a channel linked to parasite-encoded RhopH proteins. These studies identify pyridazinones as novel and tractable antimalarial scaffolds with a defined mechanism of action. SIGNIFICANCE STATEMENT: Because antimalarial drugs are prone to evolving resistance in the virulent human P. falciparum pathogen, new therapies are needed. This study has now developed a novel drug-like series of pyridazinones that target an unexploited parasite anion channel on the host cell surface, display excellent in vitro and in vivo ADME properties, are refractory to acquired resistance, and demonstrate a well defined mechanism of action.
Assuntos
Antimaláricos , Antagonistas do Ácido Fólico , Animais , Ânions/química , Ânions/metabolismo , Antimaláricos/farmacologia , Eritrócitos/metabolismo , Humanos , Nutrientes , Plasmodium falciparum/metabolismoRESUMO
Drug resistance and a dire lack of transmission-blocking antimalarials hamper malaria elimination. Here, we present the pantothenamide MMV693183 as a first-in-class acetyl-CoA synthetase (AcAS) inhibitor to enter preclinical development. Our studies demonstrate attractive drug-like properties and in vivo efficacy in a humanized mouse model of Plasmodium falciparum infection. The compound shows single digit nanomolar in vitro activity against P. falciparum and P. vivax clinical isolates, and potently blocks P. falciparum transmission to Anopheles mosquitoes. Genetic and biochemical studies identify AcAS as the target of the MMV693183-derived antimetabolite, CoA-MMV693183. Pharmacokinetic-pharmacodynamic modelling predict that a single 30 mg oral dose is sufficient to cure a malaria infection in humans. Toxicology studies in rats indicate a > 30-fold safety margin in relation to the predicted human efficacious exposure. In conclusion, MMV693183 represents a promising candidate for further (pre)clinical development with a novel mode of action for treatment of malaria and blocking transmission.
Assuntos
Antimaláricos , Antagonistas do Ácido Fólico , Malária Falciparum , Malária Vivax , Malária , Animais , Antimaláricos/farmacologia , Antimaláricos/uso terapêutico , Malária/tratamento farmacológico , Malária Falciparum/tratamento farmacológico , Malária Vivax/tratamento farmacológico , Camundongos , Ácido Pantotênico/análogos & derivados , Plasmodium falciparum/genética , RatosRESUMO
A series of 5-aryl-2-amino-imidazothiadiazole (ITD) derivatives were identified by a phenotype-based high-throughput screening using a blood stage Plasmodium falciparum (Pf) growth inhibition assay. A lead optimization program focused on improving antiplasmodium potency, selectivity against human kinases, and absorption, distribution, metabolism, excretion, and toxicity properties and extended pharmacological profiles culminated in the identification of INE963 (1), which demonstrates potent cellular activity against Pf 3D7 (EC50 = 0.006 µM) and achieves "artemisinin-like" kill kinetics in vitro with a parasite clearance time of <24 h. A single dose of 30 mg/kg is fully curative in the Pf-humanized severe combined immunodeficient mouse model. INE963 (1) also exhibits a high barrier to resistance in drug selection studies and a long half-life (T1/2) across species. These properties suggest the significant potential for INE963 (1) to provide a curative therapy for uncomplicated malaria with short dosing regimens. For these reasons, INE963 (1) was progressed through GLP toxicology studies and is now undergoing Ph1 clinical trials.
Assuntos
Antimaláricos , Antagonistas do Ácido Fólico , Malária Falciparum , Malária , Animais , Antimaláricos/farmacologia , Antimaláricos/uso terapêutico , Antagonistas do Ácido Fólico/uso terapêutico , Malária/tratamento farmacológico , Malária Falciparum/tratamento farmacológico , Camundongos , Camundongos SCID , Plasmodium falciparumRESUMO
The production of reactive oxygen species (ROS) increases considerably in situations of cellular stress, inducing lipid peroxidation and multiple alterations in proteins and nucleic acids. However, sensitivity to oxidative damage varies between organs and tissues depending on the triggering process. Certain drugs used in the treatment of diverse diseases such as malaria have side effects similar to those produced by oxidative damage, although no specific study has been conducted. For this purpose, cell membrane microarrays were developed and the superoxide production evoked by the mitochondrial activity was assayed in the presence of specific inhibitors: rotenone, antimycin A and azide. Once the protocol was set up on cell membrane isolated from rat brain areas, the effect of six antimalarial drugs (atovaquone, quinidine, doxycycline, mefloquine, artemisinin, and tafenoquine) and two essential oils (Rosmarinus officinalis and Origanum majoricum) were evaluated in multiple human samples. The basal activity was different depending on the type of tissue, the liver, jejunum and adrenal gland being the ones with the highest amount of superoxide. The antimalarial drugs studied showed specific behavior according to the type of human tissue analyzed, with atovaquone and quinidine producing the highest percentage of superoxide formation, and doxycycline the lowest. In conclusion, the analysis of superoxide production evaluated in cell membranes of a collection of human tissues allowed for the characterization of the safety profile of these antimalarial drugs against toxicity mediated by oxidative stress.
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In recent years, enzymes have risen as promising therapeutic tools for different pathologies, from metabolic deficiencies, such as fibrosis conditions, ocular pathologies or joint problems, to cancer or cardiovascular diseases. Treatments based on the catalytic activity of enzymes are able to convert a wide range of target molecules to restore the correct physiological metabolism. These treatments present several advantages compared to established therapeutic approaches thanks to their affinity and specificity properties. However, enzymes present some challenges, such as short in vivo half-life, lack of targeted action and, in particular, patient immune system reaction against the enzyme. For this reason, it is important to monitor serum immune response during treatment. This can be achieved by conventional techniques (ELISA) but also by new promising tools such as microarrays. These assays have gained popularity due to their high-throughput analysis capacity, their simplicity, and their potential to monitor the immune response of patients during enzyme therapies. In this growing field, research is still ongoing to solve current health problems such as COVID-19. Currently, promising therapeutic alternatives using the angiotensin-converting enzyme 2 (ACE2) are being studied to treat COVID-19.
Assuntos
Enzima de Conversão de Angiotensina 2/uso terapêutico , Tratamento Farmacológico da COVID-19 , Terapia Enzimática/métodos , Proteínas Recombinantes/uso terapêutico , Enzima de Conversão de Angiotensina 2/farmacologia , Ensaios Clínicos Fase II como Assunto , Composição de Medicamentos/métodos , Estabilidade Enzimática , Terapia Enzimática/história , Terapia Enzimática/tendências , Meia-Vida , História do Século XX , História do Século XXI , Humanos , Proteínas Recombinantes/farmacologia , SARS-CoV-2/efeitos dos fármacos , SARS-CoV-2/metabolismo , Resultado do Tratamento , Internalização do Vírus/efeitos dos fármacosRESUMO
The emergence and spread of Plasmodium falciparum resistance to first-line antimalarials creates an imperative to identify and develop potent preclinical candidates with distinct modes of action. Here, we report the identification of MMV688533, an acylguanidine that was developed following a whole-cell screen with compounds known to hit high-value targets in human cells. MMV688533 displays fast parasite clearance in vitro and is not cross-resistant with known antimalarials. In a P. falciparum NSG mouse model, MMV688533 displays a long-lasting pharmacokinetic profile and excellent safety. Selection studies reveal a low propensity for resistance, with modest loss of potency mediated by point mutations in PfACG1 and PfEHD. These proteins are implicated in intracellular trafficking, lipid utilization, and endocytosis, suggesting interference with these pathways as a potential mode of action. This preclinical candidate may offer the potential for a single low-dose cure for malaria.
Assuntos
Antimaláricos , Malária Falciparum , Malária , Parasitos , Animais , Antimaláricos/farmacologia , Antimaláricos/uso terapêutico , Endocitose , Malária/tratamento farmacológico , Malária Falciparum/tratamento farmacológico , Plasmodium falciparumRESUMO
Dihydroorotate dehydrogenase (DHODH) has been clinically validated as a target for the development of new antimalarials. Experience with clinical candidate triazolopyrimidine DSM265 (1) suggested that DHODH inhibitors have great potential for use in prophylaxis, which represents an unmet need in the malaria drug discovery portfolio for endemic countries, particularly in areas of high transmission in Africa. We describe a structure-based computationally driven lead optimization program of a pyrrole-based series of DHODH inhibitors, leading to the discovery of two candidates for potential advancement to preclinical development. These compounds have improved physicochemical properties over prior series frontrunners and they show no time-dependent CYP inhibition, characteristic of earlier compounds. Frontrunners have potent antimalarial activity in vitro against blood and liver schizont stages and show good efficacy in Plasmodium falciparum SCID mouse models. They are equally active against P. falciparum and Plasmodium vivax field isolates and are selective for Plasmodium DHODHs versus mammalian enzymes.
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Antimaláricos/farmacologia , Desenho de Fármacos , Inibidores Enzimáticos/farmacologia , Oxirredutases atuantes sobre Doadores de Grupo CH-CH/antagonistas & inibidores , Pirróis/farmacologia , Animais , Antimaláricos/química , Di-Hidro-Orotato Desidrogenase , Inibidores Enzimáticos/química , Camundongos , Plasmodium falciparum/efeitos dos fármacos , Pirróis/química , Relação Estrutura-AtividadeRESUMO
BACKGROUND: There is a continued need to develop effective and safe treatments for visceral leishmaniasis (VL). Preclinical studies on pharmacokinetics and pharmacodynamics of anti-infective agents, such as anti-bacterials and anti-fungals, have provided valuable information in the development and dosing of these agents. The aim of this study was to characterise the pharmacokinetic and pharmacodynamic properties of the anti-leishmanial drugs AmBisome and miltefosine in a preclinical disease model of VL. METHODOLOGY / PRINCIPAL FINDINGS: BALB/c mice were infected with L. donovani (MHOM/ET/67/HU3) amastigotes. Groups of mice were treated with miltefosine (orally, multi-dose regimen) or AmBisome (intravenously, single dose regimen) or left untreated as control groups. At set time points groups of mice were killed and plasma, livers and spleens harvested. For pharmacodynamics the hepatic parasite burden was determined microscopically from tissue impression smears. For pharmacokinetics drug concentrations were measured in plasma and whole tissue homogenates by LC-MS. Unbound drug concentrations were determined by rapid equilibrium dialysis. Doses exerting maximum anti-leishmanial effects were 40 mg/kg for AmBisome and 150 mg/kg (cumulatively) for miltefosine. AmBisome displayed a wider therapeutic range than miltefosine. Dose fractionation at a total dose of 2.5 mg/kg pointed towards concentration-dependent anti-leishmanial activity of AmBisome, favouring the administration of large doses infrequently. Protein binding was >99% for miltefosine and amphotericin B in plasma and tissue homogenates. CONCLUSION / SIGNIFICANCE: Using a PK/PD approach we propose optimal dosing strategies for AmBisome. Additionally, we describe pharmacokinetic and pharmacodynamic properties of miltefosine and compare our findings in a preclinical disease model to available knowledge from studies in humans. This approach also presents a strategy for improved use of animal models in the drug development process for VL.
Assuntos
Anfotericina B/farmacocinética , Antiprotozoários/farmacocinética , Leishmaniose Visceral/tratamento farmacológico , Fosforilcolina/análogos & derivados , Anfotericina B/uso terapêutico , Animais , Antiprotozoários/uso terapêutico , Quimioterapia Combinada , Proteínas de Homeodomínio/genética , Humanos , Fígado/parasitologia , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Knockout , Carga Parasitária , Fosforilcolina/farmacocinética , Fosforilcolina/uso terapêutico , Ligação Proteica/fisiologiaRESUMO
P218 is a highly selective dihydrofolate reductase inhibitor with potent in vitro activity against pyrimethamine-resistant Plasmodium falciparum. This single-center, randomized, double-blind, placebo-controlled phase Ib study evaluated P218 safety, pharmacokinetics, and chemoprotective efficacy in a P. falciparum sporozoite (PfSPZ) volunteer infection study (VIS). Consecutive dose safety and tolerability were evaluated (cohort 1), with participants receiving two oral doses of P218 1,000 mg 48 hours apart (n = 6), or placebo (n = 2). P218 chemoprotective efficacy was assessed (cohorts 2 and 3) with direct venous inoculation of 3,200 aseptic, cryopreserved PfSPZ (NF54 strain) followed 2 hours later with two P218 doses of 1,000 mg (cohort 2, n = 9) or 100 mg (cohort 3, n = 9) administered 48 hours apart, or placebo (n = 6). Parasitemia was assessed from day 7 using quantitative PCR targeting the var gene acidic terminal sequence (varATS qPCR). By day 28, all participants in cohort 2 (P218 1,000 mg) and 8/9 in cohort 3 (P218 100 mg) were sterilely protected post-PfSPZ VIS, confirming P218 P. falciparum chemoprotective activity. With placebo, all six participants became parasitemic (geometric mean time to positive parasitemia 10.6 days [90% CI: 9.9-11.4]). P218 pharmacokinetics were similar in participants with or without induced infection. Adverse events of any cause occurred in 45.8% (11/24) of participants who received P218 and 50.0% (4/8) following placebo; all were mild/moderate in severity, transient, and self-limiting. There were no clinically relevant changes in laboratory parameters, vital signs, or electrocardiograms. P218 displayed excellent chemoprotective efficacy against P. falciparum with favorable safety and tolerability.
Assuntos
Antimaláricos/administração & dosagem , Antagonistas do Ácido Fólico/administração & dosagem , Malária Falciparum/prevenção & controle , Plasmodium falciparum/efeitos dos fármacos , Esporozoítos/efeitos dos fármacos , Voluntários , Adulto , Animais , Antimaláricos/uso terapêutico , Estudos de Coortes , Método Duplo-Cego , Feminino , Antagonistas do Ácido Fólico/uso terapêutico , Experimentação Humana , Humanos , Malária Falciparum/tratamento farmacológico , Masculino , Camundongos , Camundongos Transgênicos , Pessoa de Meia-Idade , Parasitemia/tratamento farmacológico , Placebos/administração & dosagem , Distribuição AleatóriaRESUMO
BACKGROUND AND PURPOSE: Efficacy of current antimalarial treatments is declining as a result of increasing antimalarial drug resistance, so new and potent antimalarial drugs are urgently needed. Azithromycin, an azalide antibiotic, was found useful in malaria therapy, but its efficacy in humans is low. EXPERIMENTAL APPROACH: Four compounds belonging to structurally different azalide classes were tested and their activities compared to azithromycin and chloroquine. in vitro evaluation included testing against sensitive and resistant Plasmodium falciparum, cytotoxicity against HepG2 cells, accumulation and retention in human erythrocytes, antibacterial activity, and mode of action studies (delayed death phenotype and haem polymerization). in vivo assessment enabled determination of pharmacokinetic profiles in mice, rats, dogs, and monkeys and in vivo efficacy in a humanized mouse model. KEY RESULTS: Novel fast-acting azalides were highly active in vitro against P. falciparum strains exhibiting various resistance patterns, including chloroquine-resistant strains. Excellent antimalarial activity was confirmed in a P. falciparum murine model by strong inhibition of haemozoin-containing trophozoites and quick clearance of parasites from the blood. Pharmacokinetic analysis revealed that compounds are metabolically stable and have moderate oral bioavailability, long half-lives, low clearance, and substantial exposures, with blood cells as the preferred compartment, especially infected erythrocytes. Fast anti-plasmodial action is achieved by the high accumulation into infected erythrocytes and interference with parasite haem polymerization, a mode of action different from slow-acting azithromycin. CONCLUSION AND IMPLICATIONS: The hybrid derivatives described here represent excellent antimalarial drug candidates with the potential for clinical use in malaria therapy.
Assuntos
Antimaláricos , Malária , Animais , Antimaláricos/farmacologia , Antimaláricos/uso terapêutico , Azitromicina/farmacologia , Azitromicina/uso terapêutico , Cloroquina/farmacologia , Cloroquina/uso terapêutico , Cães , Malária/tratamento farmacológico , Camundongos , Plasmodium falciparum , RatosRESUMO
Malaria puts at risk nearly half the world's population and causes high mortality in sub-Saharan Africa, while drug resistance threatens current therapies. The pyrimidine biosynthetic enzyme dihydroorotate dehydrogenase (DHODH) is a validated target for malaria treatment based on our finding that triazolopyrimidine DSM265 (1) showed efficacy in clinical studies. Herein, we describe optimization of a pyrrole-based series identified using a target-based DHODH screen. Compounds with nanomolar potency versus Plasmodium DHODH and Plasmodium parasites were identified with good pharmacological properties. X-ray studies showed that the pyrroles bind an alternative enzyme conformation from 1 leading to improved species selectivity versus mammalian enzymes and equivalent activity on Plasmodium falciparum and Plasmodium vivax DHODH. The best lead DSM502 (37) showed in vivo efficacy at similar levels of blood exposure to 1, although metabolic stability was reduced. Overall, the pyrrole-based DHODH inhibitors provide an attractive alternative scaffold for the development of new antimalarial compounds.
Assuntos
Antimaláricos/uso terapêutico , Inibidores Enzimáticos/uso terapêutico , Malária Falciparum/tratamento farmacológico , Oxirredutases atuantes sobre Doadores de Grupo CH-CH/antagonistas & inibidores , Pirróis/uso terapêutico , Animais , Antimaláricos/síntese química , Antimaláricos/metabolismo , Antimaláricos/farmacocinética , Linhagem Celular Tumoral , Cristalografia por Raios X , Di-Hidro-Orotato Desidrogenase , Cães , Inibidores Enzimáticos/síntese química , Inibidores Enzimáticos/metabolismo , Inibidores Enzimáticos/farmacocinética , Feminino , Humanos , Masculino , Camundongos SCID , Microssomos Hepáticos/metabolismo , Estrutura Molecular , Oxirredutases atuantes sobre Doadores de Grupo CH-CH/metabolismo , Testes de Sensibilidade Parasitária , Plasmodium falciparum/efeitos dos fármacos , Plasmodium falciparum/enzimologia , Plasmodium vivax/efeitos dos fármacos , Plasmodium vivax/enzimologia , Ligação Proteica , Pirróis/síntese química , Pirróis/metabolismo , Pirróis/farmacocinética , Ratos , Relação Estrutura-AtividadeRESUMO
Malaria eradication is critically dependent on new therapeutics that target resistant Plasmodium parasites and block transmission of the disease. Here, we report that pantothenamide bioisosteres were active against blood-stage Plasmodium falciparum parasites and also blocked transmission of sexual stages to the mosquito vector. These compounds were resistant to degradation by serum pantetheinases, showed favorable pharmacokinetic properties, and cleared parasites in a humanized mouse model of P. falciparum infection. Metabolomics revealed that coenzyme A biosynthetic enzymes converted pantothenamides into coenzyme A analogs that interfered with parasite acetyl-coenzyme A anabolism. Resistant parasites generated in vitro showed mutations in acetyl-coenzyme A synthetase and acyl-coenzyme A synthetase 11. Introduction and reversion of these mutations in P. falciparum using CRISPR-Cas9 gene editing confirmed the roles of these enzymes in the sensitivity of the malaria parasites to pantothenamides. These pantothenamide compounds with a new mode of action may have potential as drugs against malaria parasites.
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Acetilcoenzima A/biossíntese , Antimaláricos/farmacologia , Vias Biossintéticas/efeitos dos fármacos , Ácido Pantotênico/análogos & derivados , Ácido Pantotênico/farmacologia , Plasmodium falciparum/metabolismo , Animais , Antimaláricos/química , Antimaláricos/farmacocinética , Modelos Animais de Doenças , Resistência a Medicamentos/efeitos dos fármacos , Humanos , Malária Falciparum/parasitologia , Malária Falciparum/transmissão , Masculino , Camundongos Endogâmicos BALB C , Mutação/genética , Ácido Pantotênico/química , Parasitemia/tratamento farmacológico , Parasitos/efeitos dos fármacos , Parasitos/metabolismo , Proteínas de Protozoários/genética , Reprodução Assexuada/efeitos dos fármacos , Resultado do Tratamento , Trofozoítos/efeitos dos fármacos , Trofozoítos/metabolismoRESUMO
There is a pressing need for safe and highly effective Plasmodium falciparum (Pf) malaria vaccines. The circumsporozoite protein (CS), expressed on sporozoites and during early hepatic stages, is a leading target vaccine candidate, but clinical efficacy has been modest so far. Conversely, whole-sporozoite (WSp) vaccines have consistently shown high levels of sterilizing immunity and constitute a promising approach to effective immunization against malaria. Here, we describe a novel WSp malaria vaccine that employs transgenic sporozoites of rodent P. berghei (Pb) parasites as cross-species immunizing agents and as platforms for expression and delivery of PfCS (PbVac). We show that both wild-type Pb and PbVac sporozoites unabatedly infect and develop in human hepatocytes while unable to establish an infection in human red blood cells. In a rabbit model, similarly susceptible to Pb hepatic but not blood infection, we show that PbVac elicits cross-species cellular immune responses, as well as PfCS-specific antibodies that efficiently inhibit Pf sporozoite liver invasion in human hepatocytes and in mice with humanized livers. Thus, PbVac is safe and induces functional immune responses in preclinical studies, warranting clinical testing and development.