LAUSR

Marburg virus (MARV) belongs to the Filoviridae family, along with the related Ebola virus (EBOV). Although MARV is less renowned than EBOV, it causes an equally devastating disease, with clinical symptoms similar to EBOV infection and a remarkably high mortality rate. To date, about 600 MARV cases have been reported globally, with an average mortality rate of around 80% (Brauburger et al. 2012). Due to the severity of disease caused by MARV, as well as its threat to global public health, MARV is classified as a risk group 4 pathogen by WHO, necessitating containment measures equivalent to biosafety level 4 (BSL-4). Moreover, MARV is considered a potential agent of bioterrorism (Leffel and Reed 2004) and, since 2015, has been listed as one of ten priority pathogens on WHO’s Blueprint of Priority. However, drug development specific for MARV is seldomly reported, although some studies for drugs against EBOV have identified compounds that may also be effective against MARV (Madrid et al. 2013; Bixler et al. 2017; Cross et al. 2018). MARV is a single-stranded negative-sense RNA virus. Its genome encodes seven proteins, of which the glycoprotein (GP) is the only membrane protein and is solely responsible for mediating virus adhesion and entry into host cells (Brauburger et al. 2012). Because GP is one of the key elements determining virus infection, it is therefore the main target of MARV drug, antibody and vaccine research. Importantly, despite the fact that MARV and EBOV belong to the same virus family, the amino acid homology between their glycoproteins is only * 30% (Flyak et al. 2015; Flyak et al. 2016). Thus, drugs against EBOV may not be useful against MARV, and the importance of searching for MARV-specific drugs cannot be neglected. The requirement for BSL-4 facilities greatly restricts the progress of research into MARV. To address this restriction, we previously developed a replication-defective, HIV-based pseudovirus expressing MARV GP, and we demonstrated the utility of this pseudovirus in both highthroughput screening in vitro and in a bioluminescent imaging (BLI) mouse model (Zhang et al. 2017). Importantly, this pseudovirus system can be safely used in BSL-2 laboratories. In the present study, we have leveraged our pseudovirus system to screen a large panel of clinicallyapproved drugs against MARV. The compound library, which was obtained from the National Institutes for Food and Drug Control in China, contains 767 approved drugs that have each passed strict clinical trials and for which the pharmacokinetics, toxicity and side effects have all been clearly defined. The experimental design of the in vitro screen is illustrated in supplementary Fig. S1A. The details of the experiment were shown in Supplementary Materials and Methods. The experiment was optimized and the quality of the screen was confirmed before the formal testing by assessing a set of commonly used parameters (Wang et al. 2014): signal-to-noise ratio (S/B), Z’ factor, and coefficient of variation (CV). The high S/B value (769), a Z’ factor value of over 0.5 (0.62), and a very low CV (0.5%) indicate the reliability of this screen (supplementary Fig. S1B). Following assay validation, compounds were screened as outlined in supplementary Fig. S1C. Initially, a primary screen of all 767 compounds was performed using a single dose of 200 lmol/L. This screen Li Zhang and Shan Lei have contributed equally to this work.