RESUMO
A new class of chemical inhibitors has been discovered that interferes with the process of herpesvirus DNA replication. To date, the majority of useful herpesvirus antivirals are nucleoside analogues that block herpesvirus DNA replication by targeting the DNA polymerase. The new helicase-primase inhibitors (HPI) target a different enzyme complex that is also essential for herpesvirus DNA replication. This review will place the HPI in the context of previous work on the nucleoside analogues. Several promising highly potent HPI will be described with a particular focus on the identification of drug-resistance mutations. Several HPI have good pharmacological profiles and are now at the outset of phase II clinical trials. Provided there are no safety issues to stop their progress, this new class of compound will be a major advance in the herpesvirus antiviral field. Furthermore, HPI are likely to have a major impact on the therapy and prevention of herpes simplex virus and varicella zoster in both immunocompetent and immunocompromised patients alone or in combination with current nucleoside analogues. The possibility of acquired drug-resistance to HPI will then become an issue of great practical importance.
Assuntos
Antivirais/farmacologia , DNA Helicases/antagonistas & inibidores , DNA Primase/antagonistas & inibidores , Proteínas Virais/antagonistas & inibidores , Animais , Farmacorresistência Viral , Inibidores Enzimáticos/farmacologia , Herpes Genital/tratamento farmacológico , Herpes Genital/enzimologia , Herpes Simples/tratamento farmacológico , Herpes Simples/enzimologia , Herpesvirus Humano 1/efeitos dos fármacos , Herpesvirus Humano 1/enzimologia , Herpesvirus Humano 2/efeitos dos fármacos , Herpesvirus Humano 2/enzimologia , Humanos , Replicação Viral/efeitos dos fármacosRESUMO
Herpes simplex virus infections are the cause of significant morbidity, and currently used therapeutics are largely based on modified nucleoside analogs that inhibit viral DNA polymerase function. To target this disease in a new way, we have identified and optimized selective thiazolylphenyl-containing inhibitors of the herpes simplex virus (HSV) helicase-primase enzyme. The most potent compounds inhibited the helicase, the primase and the DNA-dependent ATPase activities of the enzyme with IC50 (50% inhibitory concentration) values less than 100 nM. Inhibition of the enzymatic activities was through stabilization of the interaction between the helicase-primase and DNA substrates, preventing the progression through helicase or primase catalytic cycles. Helicase-primase inhibitors also prevented viral replication as demonstrated in viral growth assays. One compound, BILS 179 BS, displayed an EC50 (effective concentration inhibiting viral growth by 50%) of 27 nM against viral growth with a selectivity index greater than 2,000. Antiviral activity was also demonstrated for multiple strains of HSV, including strains resistant to nucleoside-based therapies. Most importantly, BILS 179 BS was orally active against HSV infections in murine models of HSV-1 and HSV-2 disease and more effective than acyclovir when the treatment frequency per day was reduced or when initiation of treatment was delayed up to 65 hours after infection. These studies validate the use of helicase-primase inhibitors for the treatment of acute herpesvirus infections and provide new lead compounds for optimization and design of superior anti-HSV agents.
Assuntos
Antivirais/uso terapêutico , DNA Helicases/antagonistas & inibidores , Inibidores Enzimáticos/uso terapêutico , Herpes Simples/tratamento farmacológico , Piridinas/uso terapêutico , Tiazóis/uso terapêutico , Animais , Antivirais/química , DNA Primase , Modelos Animais de Doenças , Desenho de Fármacos , Inibidores Enzimáticos/química , Feminino , Herpes Genital/tratamento farmacológico , Herpes Genital/enzimologia , Herpes Simples/enzimologia , Herpesvirus Humano 1/enzimologia , Herpesvirus Humano 2/enzimologia , Humanos , Técnicas In Vitro , Camundongos , Camundongos Pelados , Proteínas ViraisRESUMO
An amplified enzyme immunoassay (EIA) for herpes simplex virus (Novo Nordisk) was compared with cell culture in 853 genital specimens from a genito-urinary medicine clinic. The sensitivity of the EIA was 86% and its specificity 99.6%. The sensitivity increased to 94% for lesion swabs but decreased to 68% for cervical swabs. Sensitivity for urethral and vulval swabs was 83% and 82%, respectively. It is concluded that the EIA is specific and quick and easy to perform. It will be suitable for testing for genital herpes simplex infections in laboratories without access to local cell culture facilities.
Assuntos
Herpes Genital/diagnóstico , Técnicas Imunoenzimáticas/normas , Células Cultivadas , Colo do Útero/microbiologia , Feminino , Herpes Genital/enzimologia , Humanos , Sensibilidade e Especificidade , Simplexvirus/enzimologia , Uretra/microbiologia , Vulva/microbiologiaRESUMO
Mice deficient in RNA-dependent protein kinase (PKR-/-) or deficient in PKR and a functional 2',5'-oligoadenylate synthetase (OAS) pathway (PKR/RL-/-) are more susceptible to genital herpes simplex virus type 2 (HSV-2) infection than wild-type mice or mice that are deficient only in a functional OAS pathway (RL-/-) as measured by survival over 30 days. The increase in susceptibility correlated with an increase in virus titre recovered from vaginal tissue or brainstem of infected mice during acute infection. There was also an increase in CD45+ cells and CD8+ T cells residing in the central nervous system of HSV-2-infected PKR/RL-/- mice in comparison with RL-/- or wild-type control animals. In contrast, there was a reduction in the HSV-specific CD8+ T cells within the draining lymph node of the PKR/RL-/- mice. Collectively, activation of PKR, but not of OAS, contributes significantly to the local control and spread of HSV-2 following genital infection.
Assuntos
Herpes Genital/enzimologia , Herpesvirus Humano 2 , eIF-2 Quinase/genética , 2',5'-Oligoadenilato Sintetase/metabolismo , Animais , Tronco Encefálico/virologia , Complexo CD3/imunologia , Linfócitos T CD8-Positivos/imunologia , Quimiocina CCL3 , Quimiocina CCL4 , Quimiocinas CC/análise , Suscetibilidade a Doenças , Ensaio de Imunoadsorção Enzimática/métodos , Feminino , Citometria de Fluxo/métodos , Herpes Genital/imunologia , Interferon gama/análise , Linfonodos/imunologia , Contagem de Linfócitos , Proteínas Inflamatórias de Macrófagos/análise , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Medula Espinal/virologia , Fator de Necrose Tumoral alfa/análiseRESUMO
Small DNA viruses (adenoviruses, simian virus 40, or human papillomaviruses) induce S-phase progression but prevent cell division to provide precursors for viral DNA replication. Herpes simplex viruses types 1 or 2 (HSV-1 or HSV-2) contain genes which encode DNA-metabolizing enzymes, for example, ribonucleotide reductase, thymidine kinase and dUTPase, suggesting that S-phase factors are not required for an efficient infection. However, several studies indicated that HSV induces some events that occur during cell-cycle progression. To determine if HSV-2 induces S-phase entry, we examined serum-arrested African green monkey kidney cells (CV-1) after infection. Two hours after infection steady-state levels of the S-phase-specific cyclin, cyclin A, increased. S-phase cyclin-dependent kinase activity (CDK2) was stimulated 10-fold 8 h after infection but decreased at 16 or 24 h after infection. Mitotic CDK activity (CDC2) was not activated after infection, in part due to decreases in CDC2 protein levels and inactivation of enzymatic activity resulting from tyrosine phosphorylation of CDC2. Furthermore, CDK4 activity was not dramatically affected by infection. These studies indicate that HSV-2 infection selectively activates CDK2 after infection but cell-cycle progression does not occur. We hypothesize that infection activates certain components of the cell cycle which enhance viral gene expression and DNA replication.