RESUMO

The capsid (CA) subunit of the HIV-1 Gag polyprotein is involved in several steps of the viral cycle, from the assembly of new viral particles to the protection of the viral genome until it enters into the nucleus of newly infected cells. As such, it represents an interesting therapeutic target to tackle HIV infection. In this study, we screened hundreds of compounds with a low cost of synthesis for their ability to interfere with Gag assembly in vitro. Representatives of the most promising families of compounds were then tested for their ability to inhibit HIV-1 replication in cellulo. From these molecules, a hit compound from the benzimidazole family with high metabolic stability and low toxicity, 2-(4-N,N-dimethylaminophenyl)-5-methyl-1-phenethyl-1H-benzimidazole (696), appeared to block HIV-1 replication with an IC50 of 3 µM. Quantitative PCR experiments demonstrated that 696 does not block HIV-1 infection before the end of reverse transcription, and molecular docking confirmed that 696 is likely to bind at the interface between two monomers of CA and interfere with capsid oligomerization. Altogether, 696 represents a promising lead molecule for the development of a new series of HIV-1 inhibitors.

Assuntos

Infecções por HIV , HIV-1 , Benzimidazóis/farmacologia , Proteínas do Capsídeo , Humanos , Simulação de Acoplamento Molecular , Replicação Viral

RESUMO

Trematode infections such as schistosomiasis and fascioliasis cause significant morbidity in an estimated 250 million people worldwide and the associated agricultural losses are estimated at more than US$ 6 billion per year. Current chemotherapy is limited. Triosephosphate isomerase (TIM), an enzyme of the glycolytic pathway, has emerged as a useful drug target in many parasites, including Fasciola hepatica TIM (FhTIM). We identified 21 novel compounds that selectively inhibit this enzyme. Using microscale thermophoresis we explored the interaction between target and compounds and identified a potent interaction between the sulfonyl-1,2,4-thiadiazole (compound 187) and FhTIM, which showed an IC50 of 5 µM and a Kd of 66 nM. In only 4 hours, this compound killed the juvenile form of F. hepatica with an IC50 of 3 µM, better than the reference drug triclabendazole (TCZ). Interestingly, we discovered in vitro inhibition of FhTIM by TCZ, with an IC50 of 7 µM suggesting a previously uncharacterized role of FhTIM in the mechanism of action of this drug. Compound 187 was also active against various developmental stages of Schistosoma mansoni. The low toxicity in vitro in different cell types and lack of acute toxicity in mice was demonstrated for this compound, as was demonstrated the efficacy of 187 in vivo in F. hepatica infected mice. Finally, we obtained the first crystal structure of FhTIM at 1.9 Å resolution which allows us using docking to suggest a mechanism of interaction between compound 187 and TIM. In conclusion, we describe a promising drug candidate to control neglected trematode infections in human and animal health.

Assuntos

Anti-Helmínticos/química , Anti-Helmínticos/farmacologia , Trematódeos/efeitos dos fármacos , Trematódeos/enzimologia , Infecções por Trematódeos/tratamento farmacológico , Triose-Fosfato Isomerase/antagonistas & inibidores , Animais , Anti-Helmínticos/uso terapêutico , Cristalografia por Raios X , Descoberta de Drogas , Inibidores Enzimáticos/química , Inibidores Enzimáticos/farmacologia , Inibidores Enzimáticos/uso terapêutico , Fasciola hepatica/efeitos dos fármacos , Fasciola hepatica/enzimologia , Fasciolíase/tratamento farmacológico , Fasciolíase/parasitologia , Feminino , Masculino , Mesocricetus , Camundongos , Camundongos Endogâmicos BALB C , Modelos Moleculares , Schistosoma mansoni/efeitos dos fármacos , Schistosoma mansoni/enzimologia , Esquistossomose mansoni/tratamento farmacológico , Esquistossomose mansoni/parasitologia , Infecções por Trematódeos/parasitologia , Triose-Fosfato Isomerase/metabolismo

RESUMO

Feline immunodeficiency virus (FIV) is a member of the retroviridae family of viruses. It causes acquired immunodeficiency syndrome (AIDS) in worldwide domestic and non-domestic cats and is a cause of an important veterinary issue. The genome organization of FIV and the clinical characteristics of the disease caused by FIV are similar to human immunodeficiency virus (HIV). Both viruses infect T lymphocytes, monocytes, and macrophages, with a similar replication cycle in infected cells. Thus, the infection of cats with FIV is also a useful tool for the study and development of novel drugs and vaccines against HIV. Anti-retroviral drugs studied extensively with regards to HIV infection have targeted different steps of the virus replication cycle: (1) disruption of the interaction with host cell surface receptors and co-receptors; (2) inhibition of fusion of the virus and cell membranes; (3) blocking of the reverse transcription of viral genomic RNA; (4) interruption of nuclear translocation and integration of viral DNA into host genomes; (5) prevention of viral transcript processing and nuclear export; and (6) inhibition of virion assembly and maturation. Despite the great success of anti-retroviral therapy in slowing HIV progression in humans, a similar therapy has not been thoroughly investigated for FIV infection in cats, mostly because of the little structural information available for FIV proteins. The FIV capsid protein (CA) drives the assembly of the viral particle, which is a critical step in the viral replication cycle. During this step, the CA protein oligomerizes to form a protective coat that surrounds the viral genome. In this work, we perform a large-scale screening of four hundred molecules from our in-house library using an in vitro assembly assay of p24, combined with microscale thermophoresis, to estimate binding affinity. This screening led to the discovery of around four novel hits that inhibited capsid assembly in vitro. These may provide new antiviral drugs against FIV.