RESUMO
Iron is an essential growth component in all living organisms and plays a central role in numerous biochemical processes due to its redox potential and high affinity for oxygen. The use of iron chelators has been suggested as a novel therapeutic approach towards parasitic infections, such as malaria, sleeping sickness and leishmaniasis. Known iron chelating agents such as Deferoxamine and the 3-hydroxypyridin-4-one (HPO) Deferiprone possess anti-parasitic activity but suffer from mammalian toxicity, relatively modest potency, and/or poor oral availability. In this study, we have developed novel derivatives of Deferiprone with increased anti-parasitic activity and reduced cytotoxicity against human cell lines. Of particular interest are several new derivatives in which the HPO scaffold has been conjugated, via a linker, to the 4-aminoquinoline ring system present in the known anti-malaria drug Chloroquine. We report the inhibitory activity of these novel analogues against four parasitic protozoa, Trypanosoma brucei, Trypanosoma cruzi, Leishmania infantum and Plasmodium falciparum, and, for direct comparison, against human cells lines. We also present data, which support the hypothesis that iron starvation is the major cause of growth inhibition of these new Deferiprone-Chloroquine conjugates in T. brucei.
Assuntos
Aminoquinolinas/química , Antiprotozoários/farmacologia , Quelantes de Ferro/farmacologia , Piridonas/farmacologia , Antiprotozoários/síntese química , Antiprotozoários/química , Deferiprona , Relação Dose-Resposta a Droga , Quelantes de Ferro/síntese química , Quelantes de Ferro/química , Leishmania infantum/efeitos dos fármacos , Estrutura Molecular , Testes de Sensibilidade Parasitária , Plasmodium falciparum/efeitos dos fármacos , Piridonas/síntese química , Piridonas/química , Relação Estrutura-Atividade , Trypanosoma brucei brucei/efeitos dos fármacos , Trypanosoma cruzi/efeitos dos fármacosRESUMO
The membrane bound drug efflux pump P-glycoprotein (P-gp) transports a wide variety of functionally and structurally diverse cytotoxic drugs out of tumour cells. Overexpression of P-glycoprotein is one of the predominant mechanisms responsible for development of multiple drug resistance in tumour therapy. Thus, inhibition of P-gp represents a promising approach for treatment of multidrug resistant tumours. This review highlights concepts for identification and optimization of new inhibitors of P-glycoprotein.
Assuntos
Membro 1 da Subfamília B de Cassetes de Ligação de ATP/antagonistas & inibidores , Membro 1 da Subfamília B de Cassetes de Ligação de ATP/química , Membro 1 da Subfamília B de Cassetes de Ligação de ATP/classificação , Membro 1 da Subfamília B de Cassetes de Ligação de ATP/metabolismo , Animais , Desenho de Fármacos , Resistência a Múltiplos Medicamentos , Resistencia a Medicamentos Antineoplásicos , Humanos , Modelos Químicos , Relação Quantitativa Estrutura-AtividadeRESUMO
In line with our studies on propafenone-type inhibitors of P-glycoprotein (P-gp), we applied several methods to approach virtual screening tools for identification of new P-gp inhibitors on one hand and the molecular basis of ligand-protein interaction on the other hand. For virtual screening, a combination of autocorrelation vectors and selforganising artificial neural networks proved extremely valuable in identifying P-gp inhibitors with structurally new scaffolds. For a closer view on the binding region for propafenone-type ligands we applied a combination of pharmacophore-driven photoaffinity labeling and protein homology modeling. On LmrA, a bacterial homologue of P-gp, we were able to identify distinct regions on transmembrane helices 3, 5 and 6 which show significant changes in the labeling pattern during different steps of the catalytic cycle.
Assuntos
Membro 1 da Subfamília B de Cassetes de Ligação de ATP/antagonistas & inibidores , Desenho de Fármacos , Marcadores de Fotoafinidade/metabolismo , Propafenona/farmacologia , Membro 1 da Subfamília B de Cassetes de Ligação de ATP/química , Antiarrítmicos/farmacologia , Proteínas de Bactérias/antagonistas & inibidores , Proteínas de Bactérias/metabolismo , Catálise , Ligantes , Proteínas Associadas à Resistência a Múltiplos Medicamentos/antagonistas & inibidores , Proteínas Associadas à Resistência a Múltiplos Medicamentos/metabolismo , Redes Neurais de Computação , Marcadores de Fotoafinidade/química , Ligação Proteica , Conformação ProteicaRESUMO
The drug efflux pump P-glycoprotein (P-gp) has been shown to promote multidrug resistance (MDR) in tumors as well as to influence ADME properties of drug candidates. Here we synthesized and tested a series of benzophenone derivatives structurally analogous to propafenone-type inhibitors of P-gp. Some of the compounds showed ligand efficiency and lipophilic efficiency (LipE) values in the range of compounds which entered clinical trials as MDR modulators. Interestingly, although lipophilicity plays a dominant role for P-gp inhibitors, all compounds investigated showed LipE values below the threshold for promising drug candidates. Docking studies of selected analogues into a homology model of P-glycoprotein suggest that benzophenones show an interaction pattern similar to that previously identified for propafenone-type inhibitors.
Assuntos
Membro 1 da Subfamília B de Cassetes de Ligação de ATP/antagonistas & inibidores , Benzofenonas/síntese química , Resistência a Múltiplos Medicamentos/efeitos dos fármacos , Membro 1 da Subfamília B de Cassetes de Ligação de ATP/metabolismo , Benzofenonas/química , Benzofenonas/farmacologia , Sítios de Ligação , Linhagem Celular Tumoral , Cristalografia por Raios X , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Ensaios de Seleção de Medicamentos Antitumorais , Canal de Potássio ERG1 , Canais de Potássio Éter-A-Go-Go/antagonistas & inibidores , Canais de Potássio Éter-A-Go-Go/metabolismo , Humanos , Ligantes , Modelos Moleculares , Conformação Molecular , Propafenona/análogos & derivados , Propafenona/síntese química , Propafenona/química , Propafenona/farmacologia , Relação Quantitativa Estrutura-Atividade , Proteínas da Membrana Plasmática de Transporte de Serotonina/metabolismo , Relação Estrutura-AtividadeRESUMO
P-glycoprotein (P-gp) is an energy-dependent multidrug efflux pump conferring resistance to cancer chemotherapy. Characterization of the mechanism of drug transport at a molecular level represents an important prerequisite for the design of pump inhibitors, which resensitize cancer cells to standard chemotherapy. In addition, P-glycoprotein plays an important role for early absorption, distribution, metabolism, excretion, and toxicity profiling in drug development. A set of propafenonetype substrate photoaffinity ligands has been used in this study in conjunction with matrix-assisted laser desorption/ionization time-of-flight mass spectrometry to define the substrate binding domain(s) of P-gp in more detail. The highest labeling was observed in transmembrane segments 3, 5, 8, and 11. A homology model for P-gp was generated on the basis of the dimeric crystal structure of Vibrio cholerae MsbA, an essential lipid transporter. Thereafter, the labeling pattern was projected onto the 3D atomic-detail model of P-gp to allow a visualization of the binding domain(s). Labeling is predicted by the model to occur at the two transmembrane domain/transmembrane domain interfaces formed between the amino- and carboxyl-terminal half of P-gp. These interfaces are formed by transmembrane (TM) segments 3 and 11 on one hand and TM segments 5 and 8 on the other hand. Available data on LmrA and AcrB, two bacterial multidrug efflux pumps, suggest that binding at domain interfaces may be a general feature of polyspecific drug efflux pumps.
Assuntos
Membro 1 da Subfamília B de Cassetes de Ligação de ATP/química , Membro 1 da Subfamília B de Cassetes de Ligação de ATP/metabolismo , Modelos Moleculares , Fragmentos de Peptídeos/química , Fragmentos de Peptídeos/metabolismo , Marcadores de Fotoafinidade/metabolismo , Homologia Estrutural de Proteína , Animais , Sítios de Ligação , Linhagem Celular , Membrana Celular/química , Membrana Celular/metabolismo , Insetos , Marcadores de Fotoafinidade/química , Propafenona/química , Propafenona/metabolismo , Ligação Proteica , Estrutura Terciária de Proteína , Especificidade por Substrato , Propriedades de SuperfícieRESUMO
LmrA is an ATP dependent multidrug transporter from Lactococcus lactis conferring antibiotic resistance to 17 out of 21 most frequently administered antibiotics. Starting from the dimeric crystal structure of Vc-MsbA, we built two homology models, with NBD:NBD interfaces reflecting the nonenergized and energized state, respectively. The TMD:TMD topology of the dimer is consistent with the previously obtained substrate photoaffinity labeling pattern suggesting binding of substrates at the TMD:TMD interface involving helix 3 of one monomer and helices 5 and 6 of the other monomer.
Assuntos
Proteínas de Bactérias/química , Resistência a Múltiplos Medicamentos , Proteínas Associadas à Resistência a Múltiplos Medicamentos/química , Homologia Estrutural de ProteínaRESUMO
Lipophilicity is one of the major determining physicochemical descriptors for P-glycoprotein (P-gp) inhibitory activity. Recently, Pajeva and Wiese showed that in case of P-gp interaction, lipophilicity may be regarded as space-directed property. In the present study, a series of propafenone-type P-gp inhibitors with systematically varying hydrophobicity distribution within the molecules were synthesised and pharmacologically tested. QSAR studies on the basis of multiple linear regression analysis showed that with increasing lipophilicity of the substituents on the amine moiety, the statistical significance of the indicator variables, denoting the substitution pattern on the central aromatic ring system, also increases. This indicates that the distribution of hydrophobicity within the molecules influences the mode of interaction with P-gp.
Assuntos
Antineoplásicos/química , Resistência a Múltiplos Medicamentos/efeitos dos fármacos , Propafenona/análogos & derivados , Propafenona/química , Membro 1 da Subfamília B de Cassetes de Ligação de ATP/antagonistas & inibidores , Membro 1 da Subfamília B de Cassetes de Ligação de ATP/química , Antineoplásicos/síntese química , Antineoplásicos/farmacologia , Linhagem Celular Tumoral , Desenho de Fármacos , Resistencia a Medicamentos Antineoplásicos , Humanos , Interações Hidrofóbicas e Hidrofílicas , Modelos Lineares , Propafenona/síntese química , Propafenona/farmacologia , Relação Quantitativa Estrutura-AtividadeRESUMO
The very late antigen-4 (VLA-4), also known as integrin alpha4beta1, is expressed on monocytes, T- and B-lympohocytes, basophils, and eosinophils and is involved in the massive recruitment of granulocytes in different pathological conditions such as multiple sclerosis and asthma. VLA-4 interacts with its endogenous ligand VCAM-1 during chronic inflammation, and blockade of VLA-4 /VCAM-1 interaction is a potential target for immunosuppression. Two classes of VLA-4 antagonists have so far been reported: beta-amino acid derivatives containing a diaryl urea moiety (BIO-1211) and phenylalanine derivatives (TR-14035). With the aim of clarifying the structural basis responsible for VLA-4 recognition by phenylalanine derivatives, we developed a combined computational study on a set of 128 antagonists available through the literature. Our computational approach is composed of three parts. (i) A VCAM-1 based pharmacophore was constructed with a restricted number of phenylalanine derivatives to identify the region of the protein that resembles synthetic antagonists. The pharmacophore was instrumental in constructing an alignment of a set of 128 compounds. This alignment was exploited to build a pseudoreceptor model with the RECEPTOR program. (ii) 3D-QSAR analysis was carried out on the computed electrostatic and steric interaction energies with the pseudoreceptor surface. The 3D-QSAR analysis yielded a predictive model able to explain much of the variance of the 128 antagonists. (iii) A homology modeling study of the headpiece of VLA-4 based on the crystal structure of alphavbeta3 was performed. Docking experiments of TR-14035 into the binding site of VLA-4 aided the interpretation of the 3D-QSAR model. The obtained results will be fruitful for the design of new potent and selective antagonists of VLA-4.
Assuntos
Integrina alfa4beta1/química , Fenilalanina/química , Sequência de Aminoácidos , Modelos Moleculares , Dados de Sequência Molecular , Relação Quantitativa Estrutura-Atividade , Homologia de Sequência de Aminoácidos , Molécula 1 de Adesão de Célula Vascular/químicaRESUMO
Resveratrol (3,5,4'-trihydroxy-trans-stilbene) is found in grapes and various medical plants. Among cytotoxic, antifungal, antibacterial cardioprotective activity resveratrol also demonstrates non-selective cyclooxygenase-1 (COX-1) and cyclooxygenase-2 (COX-2) inhibition. In order to find more selective COX-2 inhibitors a series of methoxylated and hydroxylated resveratrol derivatives were synthesized and evaluated for their ability to inhibit both enzymes using in vitro inhibition assays for COX-1 and COX-2 by measuring PGE(2) production. Hydroxylated but not methoxylated resveratrol derivatives showed a high rate of inhibition. The most potent resveratrol compounds were 3,3',4',5-tetra-trans-hydroxystilbene (COX-1: IC(50)=4.713, COX-2: IC(50)=0.0113 microM, selectivity index=417.08) and 3,3',4,4',5,5'-hexa-hydroxy-trans-stilbene (COX-1: IC(50)=0.748, COX-2: IC(50)=0.00104 microM, selectivity index=719.23). Their selectivity index was in part higher than celecoxib, a selective COX-2 inhibitor already established on the market (COX-1: IC(50)=19.026, COX-2: IC(50)=0.03482 microM, selectivity index=546.41). Effect of structural parameters on COX-2 inhibition was evaluated by quantitative structure-activity relationship (QSAR) analysis and a high correlation was found with the topological surface area TPSA (r=0.93). Docking studies on both COX-1 and COX-2 protein structures also revealed that hydroxylated but not methoxylated resveratrol analogues are able to bind to the previously identified binding sites of the enzymes. Hydroxylated resveratrol analogues therefore represent a novel class of highly selective COX-2 inhibitors and promising candidates for in vivo studies.
Assuntos
Inibidores de Ciclo-Oxigenase/síntese química , Prostaglandina-Endoperóxido Sintases/fisiologia , Estilbenos/síntese química , Animais , Ciclo-Oxigenase 2 , Inibidores de Ciclo-Oxigenase 2 , Inibidores de Ciclo-Oxigenase/farmacologia , Humanos , Proteínas de Membrana , Modelos Moleculares , Prostaglandina-Endoperóxido Sintases/química , Estrutura Secundária de Proteína , Resveratrol , Ovinos , Estilbenos/farmacologia , Relação Estrutura-AtividadeRESUMO
Multidrug resistance presents a major obstacle to the treatment of infectious diseases and cancer. LmrA, a bacterial ATP-dependent multidrug transporter, mediates efflux of hydrophobic cationic substrates, including antibiotics. The substrate-binding domain of LmrA was identified by using photo-affinity ligands, proteolytic degradation of LmrA, and identification of ligand-modified peptide fragments with matrix-assisted laser desorption ionization/time of flight mass spectrometry. In the nonenergized state, labeling occurred in the alpha-helical transmembrane segments (TM) 3, 5 and 6 of the membrane-spanning domain. Upon nucleotide binding, the accessibility of TM5 for substrates increased, whereas that of TM6 decreased. Inverse changes were observed upon ATP-hydrolysis. An atomic-detail model of dimeric LmrA was generated based on the template structure of the homologous transporter MsbA from Vibrio cholerae, allowing a three-dimensional visualization of the substrate-binding domain. Labeling of TM3 of one monomer occurred in a predicted area of contact with TM5 or TM6 of the opposite monomer, indicating substrate-binding at the monomer/monomer interface. Inverse changes in the reactivity of TM segments 5 and 6 suggest that substrate binding and release involves a repositioning of these helices during the catalytic cycle.