Fusion of the viral and host cell membranes is a necessary first step for infection by enveloped viruses and is mediated by the envelope glycoprotein. The transmembrane subunits from the structurally defined "class I" glycoproteins adopt an α-helical "trimer-of-hairpins" conformation during the fusion pathway. Here, we present our studies on the envelope glycoprotein transmembrane subunit, GP2, of the CAS virus (CASV). CASV was recently identified from annulated tree boas (Corallus annulatus) with inclusion body disease and is implicated in the disease etiology. We have generated and characterized two protein constructs consisting of the predicted CASV GP2 core domain. The crystal structure of the CASV GP2 post-fusion conformation indicates a trimeric α-helical bundle that is highly similar to those of Ebola virus and Marburg virus GP2 despite CASV genome homology to arenaviruses. Denaturation studies demonstrate that the stability of CASV GP2 is pH dependent with higher stability at lower pH; we propose that this behavior is due to a network of interactions among acidic residues that would destabilize the α-helical bundle under conditions where the side chains are deprotonated. The pH-dependent stability of the post-fusion structure has been observed in Ebola virus and Marburg virus GP2, as well as other viruses that enter via the endosome. Infection experiments with CASV and the related Golden Gate virus support a mechanism of entry that requires endosomal acidification. Our results suggest that, despite being primarily arenavirus like, the transmembrane subunit of CASV is extremely similar to the filoviruses.

Background

The SARS CoV-2 virus has caused one of the deadliest pandemics in recent history, resulting in over 170 million deaths and global economic disruption. There remains an urgent need for clinical trials to test therapies for treatment and prevention.

Design

An online research platform was created to support a registry community of healthcare workers (HCWs) to understand their experiences and conduct clinical studies to address their concerns. The first study, HERO-HCQ, was a double-blind, multicenter, randomized, pragmatic trial to evaluate the superiority of hydroxychloroquine (HCQ) vs placebo for pre-exposure prophylaxis (PrEP) of COVID-19 clinical infection in HCWs. Secondary objectives were to assess the efficacy of HCQ in preventing viral shedding of COVID-19 among HCWs and to assess the safety and tolerability of HCQ.

Methods

HCWs joined the Registry and were pre-screened for trial interest and eligibility. Trial participants were randomized 1:1 to receive HCQ or placebo. On-site baseline assessment included a COVID-19 nasopharyngeal PCR and blood serology test. Weekly follow-up was done via an online portal and included screening for symptoms of COVID-19, self-reported testing, adverse events, and quality of life assessments. The on-site visit was repeated at Day 30.

Discussion

The HERO research platform offers an approach to rapidly engage, screen, invite and enroll into clinical studies using a novel participant-facing online portal interface and remote data collection, enabling limited onsite procedures for conduct of a pragmatic clinical trial. This platform may be an example for future clinical trials of common conditions to enable more rapid evidence generation.