RESUMEN
There is an urgent need for potent and selective antivirals against SARS-CoV-2. Pfizer developed PF-07321332 (PF-332), a potent inhibitor of the viral main protease (Mpro, 3CLpro) that can be dosed orally and that is in clinical development. We here report that PF-332 exerts equipotent in vitro activity against the four SARS-CoV-2 variants of concerns (VoC) and that it can completely arrest replication of the alpha variant in primary human airway epithelial cells grown at the air-liquid interface. Treatment of Syrian Golden hamsters with PF-332 (250 mg/kg, twice daily) completely protected the animals against intranasal infection with the beta (B.1.351) and delta (B.1.617.2) SARS-CoV-2 variants. Moreover, treatment of SARS-CoV-2 (B.1.617.2) infected animals with PF-332 completely prevented transmission to untreated co-housed sentinels.
Asunto(s)
Tratamiento Farmacológico de COVID-19 , Modelos Animales de Enfermedad , Lactamas/administración & dosificación , Leucina/administración & dosificación , Nitrilos/administración & dosificación , Prolina/administración & dosificación , SARS-CoV-2/efectos de los fármacos , Inhibidores de Proteasa Viral/administración & dosificación , Células A549 , Administración Oral , Animales , COVID-19/prevención & control , COVID-19/transmisión , COVID-19/virología , Chlorocebus aethiops , Proteasas 3C de Coronavirus/antagonistas & inhibidores , Cricetinae , Humanos , Lactamas/farmacocinética , Leucina/farmacocinética , Mesocricetus , Nitrilos/farmacocinética , Prolina/farmacocinética , Mucosa Respiratoria/efectos de los fármacos , Mucosa Respiratoria/virología , SARS-CoV-2/enzimología , SARS-CoV-2/fisiología , Células Vero , Inhibidores de Proteasa Viral/farmacocinética , Replicación Viral/efectos de los fármacosAsunto(s)
Tratamiento Farmacológico de COVID-19 , Lactamas/uso terapéutico , Leucina/uso terapéutico , Nitrilos/uso terapéutico , Prolina/uso terapéutico , Ritonavir/efectos adversos , Ritonavir/uso terapéutico , Inhibidores de Proteasa Viral , Interacciones Farmacológicas , Quimioterapia Combinada , Humanos , Ritonavir/farmacocinética , Ritonavir/farmacología , Inhibidores de Proteasa Viral/efectos adversos , Inhibidores de Proteasa Viral/farmacocinética , Inhibidores de Proteasa Viral/uso terapéuticoRESUMEN
The worldwide outbreak of COVID-19 caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has become a global pandemic. Alongside vaccines, antiviral therapeutics are an important part of the healthcare response to countering the ongoing threat presented by COVID-19. Here, we report the discovery and characterization of PF-07321332, an orally bioavailable SARS-CoV-2 main protease inhibitor with in vitro pan-human coronavirus antiviral activity and excellent off-target selectivity and in vivo safety profiles. PF-07321332 has demonstrated oral activity in a mouse-adapted SARS-CoV-2 model and has achieved oral plasma concentrations exceeding the in vitro antiviral cell potency in a phase 1 clinical trial in healthy human participants.
Asunto(s)
Tratamiento Farmacológico de COVID-19 , Lactamas/farmacología , Lactamas/uso terapéutico , Leucina/farmacología , Leucina/uso terapéutico , Nitrilos/farmacología , Nitrilos/uso terapéutico , Prolina/farmacología , Prolina/uso terapéutico , SARS-CoV-2/efectos de los fármacos , Inhibidores de Proteasa Viral/farmacología , Inhibidores de Proteasa Viral/uso terapéutico , Administración Oral , Animales , COVID-19/virología , Ensayos Clínicos Fase I como Asunto , Coronavirus/efectos de los fármacos , Modelos Animales de Enfermedad , Quimioterapia Combinada , Humanos , Lactamas/administración & dosificación , Lactamas/farmacocinética , Leucina/administración & dosificación , Leucina/farmacocinética , Ratones , Ratones Endogámicos BALB C , Pruebas de Sensibilidad Microbiana , Nitrilos/administración & dosificación , Nitrilos/farmacocinética , Prolina/administración & dosificación , Prolina/farmacocinética , Ensayos Clínicos Controlados Aleatorios como Asunto , Ritonavir/administración & dosificación , Ritonavir/uso terapéutico , SARS-CoV-2/fisiología , Inhibidores de Proteasa Viral/administración & dosificación , Inhibidores de Proteasa Viral/farmacocinética , Replicación Viral/efectos de los fármacosRESUMEN
Human noroviruses (HuNoVs) are the most common cause of viral gastroenteritis resulting annually in ~219,000 deaths and a societal cost of ~USD 60 billion, and no antivirals or vaccines are available. Here, we assess the anti-norovirus activity of new peptidomimetic aldehydes related to the protease inhibitor rupintrivir. The early hit compound 4 inhibited the replication of murine norovirus (MNV) and the HuNoV GI.1 replicon in vitro (EC50 ~1 µM) and swiftly cleared the HuNoV GI.1 replicon from the cells. Compound 4 still inhibits the proteolytic activity. We selected a resistant GI.1 replicon, with a mutation (I109V) in a highly conserved region of the viral protease, conferring a low yield of resistance against compound 4 and rupintrivir. After testing new derivatives, compound 10d was the most potent (EC50 nanomolar range). Molecular docking indicated that the aldehyde group of compounds 4 and 10d bind with Cys139 in the HuNoV 3CL protease by a covalent linkage. Finally, compound 10d inhibited the replication of HuNoV GII.4 in infected zebrafish larvae, and PK studies in mice showed an adequate profile.
Asunto(s)
Infecciones por Caliciviridae/tratamiento farmacológico , Norovirus/efectos de los fármacos , Inhibidores de Proteasa Viral/farmacología , Animales , Infecciones por Caliciviridae/virología , Línea Celular , Efecto Citopatogénico Viral/efectos de los fármacos , Farmacorresistencia Viral , Isoxazoles/farmacología , Pruebas de Sensibilidad Microbiana , Simulación del Acoplamiento Molecular , Mutación , Norovirus/enzimología , Norovirus/genética , Norovirus/fisiología , Péptido Hidrolasas/genética , Péptido Hidrolasas/metabolismo , Fenilalanina/análogos & derivados , Fenilalanina/farmacología , Pirrolidinonas/farmacología , ARN Viral/metabolismo , Replicón , Bibliotecas de Moléculas Pequeñas , Valina/análogos & derivados , Valina/farmacología , Inhibidores de Proteasa Viral/química , Inhibidores de Proteasa Viral/farmacocinética , Replicación Viral/efectos de los fármacos , Pez Cebra/virologíaRESUMEN
The main protease (Mpro) of the SARS-CoV-2 virus is one focus of drug development efforts for COVID-19. Here, we show that interactive molecular dynamics in virtual reality (iMD-VR) is a useful and effective tool for creating Mpro complexes. We make these tools and models freely available. iMD-VR provides an immersive environment in which users can interact with MD simulations and so build protein complexes in a physically rigorous and flexible way. Recently, we have demonstrated that iMD-VR is an effective method for interactive, flexible docking of small molecule drugs into their protein targets (Deeks et al. PLoS One 2020, 15, e0228461). Here, we apply this approach to both an Mpro inhibitor and an oligopeptide substrate, using experimentally determined crystal structures. For the oligopeptide, we test against a crystallographic structure of the original SARS Mpro. Docking with iMD-VR gives models in agreement with experimentally observed (crystal) structures. The docked structures are also tested in MD simulations and found to be stable. Different protocols for iMD-VR docking are explored, e.g., with and without restraints on protein backbone, and we provide recommendations for its use. We find that it is important for the user to focus on forming binding interactions, such as hydrogen bonds, and not to rely on using simple metrics (such as RMSD), in order to create realistic, stable complexes. We also test the use of apo (uncomplexed) crystal structures for docking and find that they can give good results. This is because of the flexibility and dynamic response allowed by the physically rigorous, atomically detailed simulation approach of iMD-VR. We make our models (and interactive simulations) freely available. The software framework that we use, Narupa, is open source, and uses commodity VR hardware, so these tools are readily accessible to the wider research community working on Mpro (and other COVID-19 targets). These should be widely useful in drug development, in education applications, e.g., on viral enzyme structure and function, and in scientific communication more generally.