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1.
Bioorg Med Chem Lett ; 110: 129887, 2024 Sep 15.
Artigo em Inglês | MEDLINE | ID: mdl-39002936

RESUMO

Human cathepsin K (CatK) stands out as a promising target for the treatment of osteoporosis, considering its role in degrading the bone matrix. Given the small and shallow S2 subsite of CatK and considering its preference for proline or hydroxyproline, we now propose the rigidification of the leucine fragment found at the P2 position in a dipeptidyl-based inhibitor, generating rigid proline-based analogs. Accordingly, with these new proline-based peptidomimetics inhibitors, we selectively inhibited CatK against other human cathepsins (B, L and S). Among these new ligands, the most active one exhibited a high affinity (pKi = 7.3 - 50.1 nM) for CatK and no inhibition over the other cathepsins. This specific inhibitor harbors two novel substituents never employed in other CatK inhibitors: the trifluoromethylpyrazole and the 4-methylproline at P3 and P2 positions. These results broaden and advance the path toward new potent and selective inhibitors for CatK.


Assuntos
Catepsina K , Peptidomiméticos , Prolina , Catepsina K/antagonistas & inibidores , Catepsina K/metabolismo , Peptidomiméticos/farmacologia , Peptidomiméticos/química , Peptidomiméticos/síntese química , Prolina/química , Prolina/farmacologia , Humanos , Relação Estrutura-Atividade , Estrutura Molecular , Relação Dose-Resposta a Droga
2.
Invest New Drugs ; 39(4): 1139-1149, 2021 08.
Artigo em Inglês | MEDLINE | ID: mdl-33475938

RESUMO

Despite the great advances in the understanding of the molecular basis of acute leukemia, very little of this knowledge has been translated into new therapies. Stathmin 1 (STMN1), a phosphoprotein that regulates microtubules dynamics, is highly expressed in acute leukemia cells and promotes cell cycle progression and proliferation. GDP366 has been described as a STMN1 and survivin inhibitor in solid tumors. This study identified structural GDP366 analogs and the cellular and molecular mechanisms underlying their suppressive effects on acute leukemia cellular models. STMN1 mRNA levels were higher in AML and ALL patients, independent of risk stratification (all p < 0.001). Cheminformatics analysis identified three structural GDP366 analogs, with AD80 more potent and effective than GSK2606414 and GW768505A. In acute leukemia cells, GDP366 and AD80 reduced cell viability and autonomous clonal growth in a dose- and/or time-dependent manner (p < 0.05) and induced apoptosis and cell cycle arrest (p < 0.05). At the molecular level, GDP366 and AD80 reduced Ki-67 (a proliferation marker) expression and S6 ribosomal protein (a PI3K/AKT/mTOR effector) phosphorylation, and induced PARP1 (an apoptosis marker) cleavage and γH2AX (a DNA damage marker) expression. GDP366 induced STMN1 phosphorylation and survivin expression, while AD80 reduced survivin and STMN1 expression. GDP366 and AD80 modulated 18 of the 84 cytoskeleton regulators-related genes. These results indicated that GDP366 and AD80 reduced the PI3K/STMN1 axis and had cytotoxic effects in acute leukemia cellular models. Our findings further highlight STMN1-mediated signaling as a putative anticancer target for acute leukemia.


Assuntos
Antineoplásicos/farmacologia , Compostos Heterocíclicos de 4 ou mais Anéis/farmacologia , Leucemia Mieloide Aguda/tratamento farmacológico , Leucemia-Linfoma Linfoblástico de Células Precursoras/tratamento farmacológico , Adenina/administração & dosagem , Adenina/análogos & derivados , Adenina/farmacologia , Antineoplásicos/administração & dosagem , Compostos Bicíclicos Heterocíclicos com Pontes/administração & dosagem , Compostos Bicíclicos Heterocíclicos com Pontes/farmacologia , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Relação Dose-Resposta a Droga , Compostos Heterocíclicos de 4 ou mais Anéis/administração & dosagem , Humanos , Indóis/administração & dosagem , Indóis/farmacologia , Células Jurkat , Leucemia Mieloide Aguda/patologia , Compostos de Fenilureia/administração & dosagem , Compostos de Fenilureia/farmacologia , Fosfatidilinositol 3-Quinase/metabolismo , Leucemia-Linfoma Linfoblástico de Células Precursoras/patologia , Inibidores de Proteínas Quinases/administração & dosagem , Inibidores de Proteínas Quinases/farmacologia , RNA Mensageiro/metabolismo , Estatmina/genética , Fatores de Tempo , Células U937
3.
J Chem Inf Model ; 61(9): 4733-4744, 2021 09 27.
Artigo em Inglês | MEDLINE | ID: mdl-34460252

RESUMO

Covalent inhibitors are assuming central importance in drug discovery projects, especially in this pandemic scenario. Many research groups have focused their attention on inhibiting viral proteases or human proteases such as cathepsin L (hCatL). The inhibition of these critical enzymes may impair viral replication. However, molecular modeling of covalent ligands is challenging since covalent and noncovalent ligand-bound states must be considered in the binding process. In this work, we evaluated the suitability of free energy perturbation (FEP) calculations as a tool for predicting the binding affinity of reversible covalent inhibitors of hCatL. Our strategy relies on the relative free energy calculated for both covalent and noncovalent complexes and the free energy changes have been compared with experimental data for eight nitrile-based inhibitors, including three new inhibitors of hCatL. Our results demonstrate that the covalent complex can be employed to properly rank the inhibitors. Nevertheless, a comparison of the free energy changes in both noncovalent and covalent states is valuable to interpret the effect triggered by the formation of the covalent bond on the interactions played by functional groups distant from the warhead. Overall, FEP can be employed as a powerful predictor tool in developing and understanding the activity of reversible covalent inhibitors.


Assuntos
Descoberta de Drogas , Entropia , Humanos , Ligantes , Modelos Moleculares , Termodinâmica
4.
Bioorg Med Chem ; 41: 116211, 2021 07 01.
Artigo em Inglês | MEDLINE | ID: mdl-33991733

RESUMO

Peptidomimetics of the class of dipeptidyl nitrile analog peptoids were synthesized as inhibitors of mammalian cysteine proteases of the papain superfamily. The dipeptidyl nitrile side chains were attached to the peptide backbone's nitrogen atom, not to the α-carbons. Synthesized nitrile-based peptoid analogs that lack the hydrogen amide at P2-P3 are responsible for many of the secondary structure elements in peptides and proteins, making them resistant to proteolysis. The designed peptoids would lose a hydrogen bond with cruzain Asp161 decreasing the affinity toward the enzyme. A structure-activity relationship and matched molecular pair-based analysis between the dipeptidyl nitrile Neq0409 and its peptoid 4a yielded the following cruzain affinities: pKiNeq0409 = 6.5 and pKi4a = 5.2. respectively. A retrosynthetic matched molecular pair cliff (RMMP-cliff) analysis with a ΔpKiNeq0409-4a of 1.3 log is found for this transformation. These novel peptoids were then optimized, leading to compound 4i, with high cruzain inhibition (pKi = 6.8). Cross-class cathepsin activity was observed for some of these novel compounds against cathepsins K, L and S, while other compounds presented a selective inhibition of cathepsin K (4b, 4c, 4k) over ten times higher than the other enzymes. The putative mode of binding was determined by using covalent docking, which also aided to describe the structure-activity relationship (SAR). Interestingly, none of the peptoids inhibited CatB to any appreciable extent. These results provide guidance to identify novel bioactive nitrile-based peptoids.


Assuntos
Inibidores de Cisteína Proteinase/farmacologia , Nitrilas/química , Peptídeos/farmacologia , Inibidores de Cisteína Proteinase/química , Modelos Moleculares , Simulação de Acoplamento Molecular , Estrutura Molecular , Peptídeos/química
5.
Bioorg Med Chem ; 29: 115827, 2021 01 01.
Artigo em Inglês | MEDLINE | ID: mdl-33254069

RESUMO

Human cathepsin B (CatB) is an important biological target in cancer therapy. In this work, we performed a knowledge-based design approach and the synthesis of a new set of 19 peptide-like nitrile-based cathepsin inhibitors. Reported compounds were assayed against a panel of human cysteine proteases: CatB, CatL, CatK, and CatS. Three compounds (7h, 7i, and 7j) displayed nanomolar inhibition of CatB and selectivity over CatK and CatL. The selectivity was achieved by using the combination of a para biphenyl ring at P3, halogenated phenylalanine in P2 and Thr-O-Bz group at P1. Likewise, compounds 7i and 7j showed selective CatB inhibition among the panel of enzymes studied. We have also described a successful example of bioisosteric replacement of the amide bond for a sulfonamide one [7e â†’ 6b], where we observed an increase in affinity and selectivity for CatB while lowering the compound lipophilicity (ilogP). Our knowledge-based design approach and the respective structure-activity relationships provide insights into the specific ligand-target interactions for therapeutically relevant cathepsins.


Assuntos
Amidas/farmacologia , Aminas/farmacologia , Catepsina B/antagonistas & inibidores , Catepsina L/antagonistas & inibidores , Desenho de Fármacos , Inibidores Enzimáticos/farmacologia , Amidas/síntese química , Amidas/química , Aminas/síntese química , Aminas/química , Catepsina B/metabolismo , Catepsina L/metabolismo , Relação Dose-Resposta a Droga , Inibidores Enzimáticos/síntese química , Inibidores Enzimáticos/química , Humanos , Simulação de Dinâmica Molecular , Estrutura Molecular , Relação Estrutura-Atividade
6.
Bioorg Med Chem Lett ; 30(18): 127439, 2020 09 15.
Artigo em Inglês | MEDLINE | ID: mdl-32717373

RESUMO

Cysteine protease B (CPB) can be targeted by reversible covalent inhibitors that could serve as antileishmanial compounds. Here, sixteen dipeptidyl nitrile derivatives were synthesized, tested against CPB, and analyzed using matched molecular pairs to determine the effects of stereochemistry and p-phenyl substitution on enzyme inhibition. The compound (S)-2-(((S)-1-(4-bromophenyl)-2,2,2-trifluoroethyl)amino)-N-(1-cyanocyclopropyl)-3-phenylpropanamide (5) was the most potent CPB inhibitor (pKi = 6.82), which was also selective for human cathepsin B (pKi < 5). The inversion of the stereochemistry from S to R was more detrimental to potency when placed at the P2 position than at P3. The p-Br derivatives were more potent than the p-CH3 and p-OCH3 derivatives, probably due to intermolecular interactions with the S3 subsite.


Assuntos
Cisteína Endopeptidases/metabolismo , Inibidores de Cisteína Proteinase/síntese química , Inibidores de Cisteína Proteinase/metabolismo , Cisteína/química , Nitrilas/síntese química , Sítios de Ligação , Catepsina B/metabolismo , Avaliação Pré-Clínica de Medicamentos , Humanos , Modelos Moleculares , Ligação Proteica , Estereoisomerismo , Relação Estrutura-Atividade
7.
J Chem Inf Model ; 60(2): 880-889, 2020 02 24.
Artigo em Inglês | MEDLINE | ID: mdl-31944110

RESUMO

One tactic for cysteine protease inhibition is to form a covalent bond between an electrophilic atom of the inhibitor and the thiol of the catalytic cysteine. In this study, we evaluate the reaction free energy obtained from a hybrid quantum mechanical/molecular mechanical (QM/MM) free energy profile as a predictor of affinity for reversible, covalent inhibitors of rhodesain. We demonstrate that the reaction free energy calculated with the PM6/MM potential is in agreement with the experimental data and suggest that the free energy profile for covalent bond formation in a protein environment may be a useful tool for the inhibitor design.


Assuntos
Cisteína Proteases/metabolismo , Inibidores de Cisteína Proteinase/metabolismo , Teoria Quântica , Cisteína Proteases/química , Ligantes , Modelos Moleculares , Conformação Proteica , Termodinâmica
8.
J Chem Inf Model ; 60(3): 1666-1677, 2020 03 23.
Artigo em Inglês | MEDLINE | ID: mdl-32126170

RESUMO

Reversible and irreversible covalent ligands are advanced cysteine protease inhibitors in the drug development pipeline. K777 is an irreversible inhibitor of cruzain, a necessary enzyme for the survival of the Trypanosoma cruzi (T. cruzi) parasite, the causative agent of Chagas disease. Despite their importance, irreversible covalent inhibitors are still often avoided due to the risk of adverse effects. Herein, we replaced the K777 vinyl sulfone group with a nitrile moiety to obtain a reversible covalent inhibitor (Neq0682) of cysteine protease. Then, we used advanced experimental and computational techniques to explore details of the inhibition mechanism of cruzain by reversible and irreversible inhibitors. The isothermal titration calorimetry (ITC) analysis shows that inhibition of cruzain by an irreversible inhibitor is thermodynamically more favorable than by a reversible one. The hybrid Quantum Mechanics/Molecular Mechanics (QM/MM) and Molecular Dynamics (MD) simulations were used to explore the mechanism of the reaction inhibition of cruzain by K777 and Neq0682. The calculated free energy profiles show that the Cys25 nucleophilic attack and His162 proton transfer occur in a single step for a reversible inhibitor and two steps for an irreversible covalent inhibitor. The hybrid QM/MM calculated free energies for the inhibition reaction correspond to -26.7 and -5.9 kcal mol-1 for K777 and Neq0682 at the MP2/MM level, respectively. These results indicate that the ΔG of the reaction is very negative for the process involving K777, consequently, the covalent adduct cannot revert to a noncovalent protein-ligand complex, and its binding tends to be irreversible. Overall, the present study provides insights into a covalent inhibition mechanism of cysteine proteases.


Assuntos
Cisteína Proteases , Trypanosoma cruzi , Cisteína Endopeptidases , Inibidores de Cisteína Proteinase/farmacologia , Proteínas de Protozoários
9.
Bioorg Med Chem ; 28(22): 115743, 2020 11 15.
Artigo em Inglês | MEDLINE | ID: mdl-33038787

RESUMO

Leishmania mexicana is an obligate intracellular protozoan parasite that causes the cutaneous form of leishmaniasis affecting South America and Mexico. The cysteine protease LmCPB is essential for the virulence of the parasite and therefore, it is an appealing target for antiparasitic therapy. A library of nitrile-based cysteine protease inhibitors was screened against LmCPB to develop a treatment of cutaneous leishmaniasis. Several compounds are sufficiently high-affinity LmCPB inhibitors to serve both as starting points for drug discovery projects and as probes for target validation. A 1.4 Å X ray crystal structure, the first to be reported for LmCPB, was determined for the complex of this enzyme covalently bound to an azadipeptide nitrile ligand. Mapping the structure-activity relationships for LmCPB inhibition revealed superadditive effects for two pairs of structural transformations. Therefore, this work advances our understanding of azadipeptidyl and dipeptidyl nitrile structure-activity relationships for LmCPB structure-based inhibitor design. We also tested the same series of inhibitors on related cysteine proteases cathepsin L and Trypanosoma cruzi cruzain. The modulation of these mammalian and protozoan proteases represents a new framework for targeting papain-like cysteine proteases.


Assuntos
Compostos Aza/farmacologia , Catepsina B/antagonistas & inibidores , Inibidores de Cisteína Proteinase/farmacologia , Leishmania mexicana/efeitos dos fármacos , Tripanossomicidas/farmacologia , Compostos Aza/síntese química , Compostos Aza/química , Catepsina B/metabolismo , Cristalografia por Raios X , Cisteína Endopeptidases/metabolismo , Inibidores de Cisteína Proteinase/síntese química , Inibidores de Cisteína Proteinase/química , Dipeptídeos/síntese química , Dipeptídeos/química , Dipeptídeos/farmacologia , Relação Dose-Resposta a Droga , Leishmania mexicana/enzimologia , Simulação de Dinâmica Molecular , Estrutura Molecular , Nitrilas/síntese química , Nitrilas/química , Nitrilas/farmacologia , Proteínas de Protozoários/antagonistas & inibidores , Proteínas de Protozoários/metabolismo , Relação Estrutura-Atividade , Tripanossomicidas/síntese química , Tripanossomicidas/química
10.
Bioorg Chem ; 101: 104039, 2020 08.
Artigo em Inglês | MEDLINE | ID: mdl-32629285

RESUMO

Cysteine proteases (CPs) are involved in a myriad of actions that include not only protein degradation, but also play an essential biological role in infectious and systemic diseases such as cancer. CPs also act as biomarkers and can be reached by active-based probes for diagnostic and mechanistic purposes that are critical in health and disease. In this paper, we present the modulation of a CP panel of parasites and mammals (Trypanosoma cruzi cruzain, LmCPB, CatK, CatL and CatS), whose inhibition by nitrile peptidomimetics allowed the identification of specificity and selectivity for a given CP. The activity cliffs identified at the CP inhibition level are useful for retrieving trends through multiple structure-activity relationships. For two of the cruzain inhibitors (10g and 4e), both enthalpy and entropy are favourable to Gibbs binding energy, thus overcoming enthalpy-entropy compensation (EEC). Group contribution of individual molecular modification through changes in enthalpy and entropy results in a separate partition on the relative differences of Gibbs binding energy (ΔΔG). Overall, this study highlights the role of CPs in polypharmacology and multi-target screening, which represents an imperative trend in the actual drug discovery effort.


Assuntos
Cisteína Proteases/química , Animais , Mamíferos , Simulação de Dinâmica Molecular , Estrutura Molecular , Relação Estrutura-Atividade
11.
Exp Parasitol ; 219: 108032, 2020 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-33137308

RESUMO

Chagas disease affects several countries around the world with health and sanitation problems. Cysteine proteases are essential for the virulence and replication of the Trypanosoma cruzi, being modulated by dipeptidyl nitriles and derivatives. Here, four dipeptidyl nitrile derivatives were assayed in three T. cruzi morphologies and two strains (Tulahuen and Y) using a set of assays: (i) analysis of the inhibitory activity against cysteine proteases; (ii) determination of the cytotoxic activity and selectivity index; (iii) verification of the inhibition of the trypomastigote invasion in the host cell. These compounds could inhibit the activity of cysteine proteases using the selective substrate Z-FR-MCA for the trypomastigote lysate and extracellular amastigotes. Interestingly, these compounds did not present relevant enzymatic inhibition for the epimastigote lysate. Most of the substances were also cytotoxic and selective against the trypomastigotes and intracellular amastigotes. The best compound of the series (Neq0662) could reduce the enzymatic activity of the cysteine proteases for the trypomastigotes and amastigotes. It was equipotent to the benznidazole drug in the cytotoxic studies using these two parasite forms. Neq0662 was also selective for the parasite, and it inhibited the invasion of the mammalian host cell in all conditions tested at 10 µM. The stereochemistry of the trifluoromethyl group was an important factor for the bioactivity when the two diastereomers (Neq0662 and Neq0663) were compared. All-in-all, these results indicate that these compounds could move further in the drug development stage because of its promising bioactive profile.


Assuntos
Inibidores de Cisteína Proteinase/farmacologia , Nitrilas/farmacologia , Trypanosoma cruzi/efeitos dos fármacos , Análise de Variância , Animais , Antiparasitários/química , Antiparasitários/farmacologia , Área Sob a Curva , Linhagem Celular , Sobrevivência Celular , Inibidores de Cisteína Proteinase/química , Inibidores de Cisteína Proteinase/toxicidade , Haplorrinos , Rim/citologia , Nitrilas/química , Proteólise , Estereoisomerismo , Sais de Tetrazólio , Tiazóis , Trypanosoma cruzi/enzimologia , Trypanosoma cruzi/crescimento & desenvolvimento , Trypanosoma cruzi/metabolismo
12.
J Enzyme Inhib Med Chem ; 35(1): 639-649, 2020 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-32048531

RESUMO

Leishmaniasis is a neglected disease caused by the protozoa Leishmania ssp. Environmental differences found by the parasites in the vector and the host are translated into cellular stress, leading to the production of heat shock proteins (Hsp). These are molecular chaperones involved in the folding of nascent proteins as well as in the regulation of gene expression, signalling events and proteostasis. Since Leishmania spp. use Hsp90 to trigger important transitions between their different stages of the life cycle, this protein family becomes a profitable target in anti-parasite drug discovery. In this work, we implemented a multidisciplinary strategy coupling molecular modelling with in vitro assays to identify small molecules able to inhibit Hsp90 from L. braziliensis (LbHsp90). Overall, we identified some compounds able to kill the promastigote form of the L. braziliensis, and to inhibit LbHsp90 ATPase activity.


Assuntos
Antiprotozoários/farmacologia , Inibidores Enzimáticos/farmacologia , Proteínas de Choque Térmico HSP90/antagonistas & inibidores , Leishmania braziliensis/efeitos dos fármacos , Chaperonas Moleculares/farmacologia , Bibliotecas de Moléculas Pequenas/farmacologia , Adenosina Trifosfatases/antagonistas & inibidores , Adenosina Trifosfatases/metabolismo , Antiprotozoários/síntese química , Antiprotozoários/química , Relação Dose-Resposta a Droga , Descoberta de Drogas , Inibidores Enzimáticos/síntese química , Inibidores Enzimáticos/química , Proteínas de Choque Térmico HSP90/metabolismo , Leishmania braziliensis/química , Modelos Moleculares , Chaperonas Moleculares/síntese química , Chaperonas Moleculares/química , Estrutura Molecular , Testes de Sensibilidade Parasitária , Bibliotecas de Moléculas Pequenas/síntese química , Bibliotecas de Moléculas Pequenas/química , Relação Estrutura-Atividade
13.
Bioorg Med Chem ; 27(22): 115083, 2019 11 15.
Artigo em Inglês | MEDLINE | ID: mdl-31561938

RESUMO

The structure-activity relationship for nitrile-based cruzain inhibitors incorporating a P2 amide replacement based on trifluoroethylamine was explored by deconstruction of a published series of inhibitors. It was demonstrated that the P3 biphenyl substituent present in the published inhibitor structures could be truncated to phenyl with only a small loss of affinity. The effects of inverting the configuration of the P2 amide replacement and linking a benzyl substituent at P1 were observed to be strongly nonadditive. We show that plotting affinity against molecular size provides a means to visualize both the molecular size efficiency of structural transformations and the nonadditivity in the structure-activity relationship. We also show how the relationship between affinity and lipophilicity, measured by high-performance liquid chromatography with an immobilized artificial membrane stationary phase, may be used to normalize affinity with respect to lipophilicity.


Assuntos
Amidas/química , Cisteína Endopeptidases/síntese química , Proteínas de Protozoários/antagonistas & inibidores , Proteínas de Protozoários/síntese química , Estrutura Molecular , Relação Estrutura-Atividade
14.
Phys Chem Chem Phys ; 21(44): 24723-24730, 2019 Nov 28.
Artigo em Inglês | MEDLINE | ID: mdl-31680132

RESUMO

Nitrile reversible covalent inhibitors of human cathepsin L (hCatL) bind covalently to the side chain of the catalytic Cys25 residue in the S1 pocket to form thioimidates. Predicting the binding of reversible covalent inhibitors is essential for their practical application in drug design. In this report, five nitrile-based inhibitors coded Neq0570, Neq0710, Neq0802, Neq0803 and Neq0804 had their hCatL inhibition constants, Ki, determined. These analogs of the prototypical Neq0570 are halogenated reversible covalent inhibitors of hCatL, which bear a halogen atom in the meta position of the P3 benzyl ring that can form a halogen bond with the Gly61 of the hCatL. To describe halogen bonding interaction in an inhibitor-hCatL complex, we applied an extra point (EP) of charge to represent the anisotropic distribution of charge on the iodine, bromine and chlorine atoms. Besides, we have used alchemical free energy calculations for evaluating the overall relative binding free energies of these inhibitors using a two-state binding model: noncovalent and covalent bond states. Our results show that free energy perturbation (FEP) can predict the hCatL binding affinities of halogenated reversible covalent inhibitors in close agreement with experiments.

15.
Exp Parasitol ; 200: 84-91, 2019 May.
Artigo em Inglês | MEDLINE | ID: mdl-30954455

RESUMO

Cysteine proteases are involved in critical cell processes to the protozoa from Leishmania genus, and their inhibition is a therapeutic alternative to treat the disease. In this work, derivatives of dipeptidyl nitriles acting as reversible covalent inhibitors of cysteine proteases were studied as cytostatic agents. The proteolytic activity inside the living and lysed parasite cells was quantified using a selective substrate for cysteine proteases (Z-FR-MCA) from Leishmania amazonensis and L. infantum. The overall proteolytic activity of intact cells and even cell extracts was only marginally affected at high concentrations, with the observation of cytostatic activity and cell cycle arrest of promastigotes. However, the cytotoxic effects were only observed for infected J774 macrophages, which impaired further analysis of the amastigote infection. Therefore, the proteolytic inhibition in intact L. amazonensis and L. infantum promastigotes had no relationship to the cytostatic activity, which emphasizes that these dipeptidyl nitriles act through another mechanism of action.


Assuntos
Antiprotozoários/farmacologia , Inibidores de Cisteína Proteinase/farmacologia , Citostáticos/farmacologia , Leishmania infantum/efeitos dos fármacos , Leishmania mexicana/efeitos dos fármacos , Nitrilas/farmacologia , Animais , Antiprotozoários/química , Linhagem Celular , Inibidores de Cisteína Proteinase/química , Citostáticos/química , Fibroblastos/efeitos dos fármacos , Citometria de Fluxo , Leishmania infantum/enzimologia , Leishmania mexicana/enzimologia , Macrófagos/efeitos dos fármacos , Camundongos , Nitrilas/química
16.
Phys Chem Chem Phys ; 20(37): 24317-24328, 2018 Sep 26.
Artigo em Inglês | MEDLINE | ID: mdl-30211406

RESUMO

Chagas disease affects millions of people in Latin America. This disease is caused by the protozoan parasite Trypanossoma cruzi. The cysteine protease cruzain is a key enzyme for the survival and propagation of this parasite lifecycle. Nitrile-based inhibitors are efficient inhibitors of cruzain that bind by forming a covalent bond with this enzyme. Here, three nitrile-based inhibitors dubbed Neq0409, Neq0410 and Neq0570 were synthesized, and the thermodynamic profile of the bimolecular interaction with cruzain was determined using isothermal titration calorimetry (ITC). The result suggests the inhibition process is enthalpy driven, with a detrimental contribution of entropy. In addition, we have used hybrid Quantum Mechanical/Molecular Mechanical (QM/MM) and Molecular Dynamics (MD) simulations to investigate the reaction mechanism of reversible covalent modification of cruzain by Neq0409, Neq0410 and Neq0570. The computed free energy profile shows that the nucleophilic attack of Cys25 on the carbon C1 of inhibitiors and the proton transfer from His162 to N1 of the dipeptidyl nitrile inhibitor take place in a single step. The calculated free energy of the inhibiton reaction is in agreement with covalent experimental binding. Altogether, the results reported here suggests that nitrile-based inhibitors are good candidates for the development of reversible covalent inhibitors of cruzain and other cysteine proteases.


Assuntos
Cisteína Endopeptidases/química , Cisteína Proteases/química , Inibidores de Cisteína Proteinase/química , Nitrilas/síntese química , Proteínas de Protozoários/química , Tripanossomicidas/química , Trypanosoma cruzi/enzimologia , Desenho de Fármacos , Simulação de Dinâmica Molecular , Ligação Proteica , Teoria Quântica , Termodinâmica
17.
Bioorg Chem ; 79: 285-292, 2018 09.
Artigo em Inglês | MEDLINE | ID: mdl-29783099

RESUMO

Cruzain is the major cysteine protease of Trypanosoma cruzi, the etiological agent of Chagas disease. Reversible covalent cruzain inhibitors can block the steps of cell differentiation in the parasite and kill the organism. To this end, the description of how inhibitors modified at the P2/P3 positions lead to analogs with greater cruzain affinity to the S2/S3 subsites is of fundamental importance. Albeit many efforts are being employed in the characterization of the interaction processes with S2 subsite, little is known about the cruzain S3 subsite. In this work, we show a brief but consistent study to identify favorable substitutions in P3 of dipeptidyl nitriles that increase cruzain affinity. Using molecular dynamics simulations, we have identified some dipeptidyl nitrile analogs with modifications at P3 position that had higher cruzain inhibition than the original unsubstituted compound. A matched molecular pair analysis shows the importance of including a chlorine atom in the P3-meta position. The modifications implemented in P3 are confirmed when profiling the thermodynamic parameters via isothermal titration calorimetry. The classical enthalpy-entropy compensation phenomenon, in which enthalpy changes are counterbalanced by entropy results in a small modification of ΔG. The inclusion of the chlorine atom in the P3-meta position results in the highest reduction of the detrimental entropic contribution observed in P3.


Assuntos
Inibidores de Cisteína Proteinase/farmacologia , Proteínas de Protozoários/antagonistas & inibidores , Trypanosoma cruzi/enzimologia , Cisteína Endopeptidases/metabolismo , Inibidores de Cisteína Proteinase/síntese química , Inibidores de Cisteína Proteinase/química , Relação Dose-Resposta a Droga , Simulação de Dinâmica Molecular , Estrutura Molecular , Proteínas de Protozoários/metabolismo , Relação Estrutura-Atividade
18.
Bioorg Med Chem Lett ; 27(17): 4001-4006, 2017 09 01.
Artigo em Inglês | MEDLINE | ID: mdl-28774426

RESUMO

Phenotypic assays were performed in prostate cancer cell lines to describe the biological activity of PI3K-AKT-mTOR pathway inhibitors retrieved from the virtual screening initiative. These novel chemicals share in common the aminopyridine scaffold, hitting PC-3 cells in macromolar range, with selectivity index over fibroblast cell lines. Moreover, a preliminary study of the mode of action by flow cytometry assay pointed out that these compounds had a rapamycin-like response for the PI3K-AKT-mTOR pathway modulation.


Assuntos
Antineoplásicos/farmacologia , Inibidores de Fosfoinositídeo-3 Quinase , Neoplasias da Próstata/tratamento farmacológico , Inibidores de Proteínas Quinases/farmacologia , Proteínas Proto-Oncogênicas c-akt/antagonistas & inibidores , Serina-Treonina Quinases TOR/antagonistas & inibidores , Animais , Antineoplásicos/síntese química , Antineoplásicos/química , Células 3T3 BALB , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Relação Dose-Resposta a Droga , Ensaios de Seleção de Medicamentos Antitumorais , Fibroblastos/efeitos dos fármacos , Humanos , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Estrutura Molecular , Fosfatidilinositol 3-Quinases/metabolismo , Neoplasias da Próstata/metabolismo , Neoplasias da Próstata/patologia , Inibidores de Proteínas Quinases/síntese química , Inibidores de Proteínas Quinases/química , Proteínas Proto-Oncogênicas c-akt/metabolismo , Relação Estrutura-Atividade , Serina-Treonina Quinases TOR/metabolismo
19.
Bioorg Med Chem Lett ; 27(3): 546-550, 2017 02 01.
Artigo em Inglês | MEDLINE | ID: mdl-28011219

RESUMO

Prostate cancer is one of the most prevalent types of cancer in male population. It is a hormone driven disease, especially in its initial phase. Hence, androgen deprivation therapy (ADT) is the major chemotherapeutic effort and novel AR inhibitors with improved pharmacological profiles are needed. In this report, a novel bioactive compound was selected and investigated using in silico and cell-based assays. Neq0502 compound was selective for the testosterone stimulated AR-dependent prostate cancer cell (LNCaP, GI50=22.4µM) when compared with unstimulated LNCaP or AR-insensitive (DU145 and PC-3) cell lines. Cell cycle arrest study provided the same profile for Neq0502 and the reference drug enzalutamide. Moreover, this compound is not cytotoxic for fibroblast Balb/C 3T3 clone A31 cells up to 250µM, with a good selectivity ratio (SI>11), which could be used in compound optimization effort to a novel therapeutic alternative.


Assuntos
Acetanilidas/farmacologia , Antineoplásicos/farmacologia , Simulação por Computador , Compostos Heterocíclicos com 3 Anéis/farmacologia , Neoplasias da Próstata/tratamento farmacológico , Receptores Androgênicos/metabolismo , Células 3T3 , Acetanilidas/química , Animais , Antineoplásicos/química , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Relação Dose-Resposta a Droga , Ensaios de Seleção de Medicamentos Antitumorais , Compostos Heterocíclicos com 3 Anéis/química , Humanos , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Estrutura Molecular , Neoplasias da Próstata/metabolismo , Neoplasias da Próstata/patologia , Relação Estrutura-Atividade
20.
Bioorg Med Chem Lett ; 27(22): 5031-5035, 2017 11 15.
Artigo em Inglês | MEDLINE | ID: mdl-29054358

RESUMO

The effects on potency of cruzain inhibition of replacing a nitrile group with alternative warheads were explored. The oxime was almost an order of magnitude more potent than the corresponding nitrile and has the potential to provide access to the prime side of the catalytic site. Dipeptide aldehydes and azadipeptide nitriles were found to be two orders of magnitude more potent cruzain inhibitors than the corresponding dipeptide nitriles although potency differences were modulated by substitution at P1 and P3. Replacement of the α methylene of a dipeptide aldehyde with cyclopropane led to a loss of potency of almost three orders of magnitude. The vinyl esters and amides that were characterized as reversible inhibitors were less potent than the corresponding nitrile by between one and two orders of magnitude.


Assuntos
Cisteína Endopeptidases/metabolismo , Inibidores de Cisteína Proteinase/química , Domínio Catalítico , Catepsina L/química , Catepsina L/metabolismo , Cisteína Endopeptidases/química , Inibidores de Cisteína Proteinase/metabolismo , Dipeptídeos/química , Desenho de Fármacos , Cinética , Nitrilas/química , Relação Estrutura-Atividade
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