Desenvolvimento de estratégias antivirais frente a vírus emergentes

Abstract

Emerging viruses pose growing threats to public health, particularly in Brazil, where environmental and sociodemographic factors favour epidemic outbreaks. This work aimed to identify and characterise antiviral compounds against the emerging viruses Orthobunyavirus oropoucheense (OROV) and Betacoronavirus pandemicum (SARS-CoV-2) through medium-throughput screening of the Pandemic Response Box (PRB) and Global Health Priority Box (GHPB) libraries from the Medicines for Malaria Venture (MMV). OROV is the causative agent of Oropouche fever, which caused over 13,000 cases in Brazil in 2024, with unprecedented geographical expansion, vertical transmission, neurological complications and confirmed fatalities, with no approved antiviral treatment available. SARS-CoV-2, responsible for the COVID-19 pandemic with over 779 million cases recorded through 2026, remains a therapeutic challenge, particularly against the Delta and Omicron variants of concern. The experimental approach employed fluorescent reporter viruses, BUNV-eGFP (Bunyamwera virus with an enhanced green fluorescent protein reporter) as a prototypic model for OROV, and SARS-CoV-2-mCherry (SARS-CoV-2 with a monomeric red fluorescent protein reporter) for direct screening, in A549 and A549-AT cells, with automated readout using the IncuCyte S3 system. Of the 640 compounds tested, active hits were identified and validated for both viral targets. Against OROV, nine compounds from the PRB inhibited BUNV replication by more than 90%. Following validation against wild-type OROV (epidemic strain AM0088), three hits were confirmed: Trimetrexate, MMV1634385, and GSK-983, being GSK the compound with greatest inhibition (EC50 = 0.5 μM against OROV; DHODH inhibitor; SI > 20), reported here for the first time as an anti-orthobunyavirus compound and identified as the most promising candidate. Against SARS-CoV-2, two hits were mechanistically characterised. Revaprazan (EC50 = 1.3 μM; SI > 7.7), a clinically approved potassium-competitive acid blocker (P-CAB), inhibited viral entry while retaining activity against Delta and Omicron BA.2, representing the first report of antiviral activity for a P-CAB-class compound. Milbemectin (EC50 = 1.6 μM; SI > 6), a milbemycin-class macrolide antiparasitic structurally related to ivermectin, acted predominantly at post-entry stages, interfering with virion assembly and egress. Together, these findings establish GSK-983, Revaprazan and Milbemectin as priority candidates for preclinical investigation, expand the antiviral spectrum of compounds with established safety profiles, and validate the fluorescent reporter-based screening platform as a robust strategy for antiviral discovery against emerging viral threats.