Drug screening against F13 protein, the target of tecovirimat, as potential therapies for monkeypox virus

The outbreak of monkeypox virus (MPXV) outside Africa has expanded worldwide since May 2022. To date, there is no approved treatment for MPXV. Tecovirimat (also known as TPOXX or ST-246) is approved for the treatment of the smallpox, another member of the orthopoxvirus family, by targeting VP37 protein. F13, as a viral structural protein of MPXV and the homolog of VP37 protein, plays an important role in the envelopment of the intracellular mature virus with Golgi-derived membrane to form enveloped virus before the release of the virus. Daoqun Li et al. established the 3D structure prediction of F13 protein and analyzed the molecular dynamics simulation of tecovirimat-F13 complex, Lam et al. found hypericin and naldemedine can be repurposable drugs for F13L protein of original european MPXV strain by QVina2.1. And Qiangzhen Yang et al. screened 5903 drugs for 10 proteins (without F13) of MPXV. Tecovirimat was demonstrated to be an effective anti-MPXV medicine clinically. Thus, we think the F13 protein, rather than other untested proteins of MPXV, is a practical and promising drug target for anti-MPXV therapy discovery and development. And we conducted drug screening for MPXV F13.

We utilized AlphaFold2 to predict the protein structure of the latest F13 of MPXV (GenBank ID URK20480.1). The predicted local distance difference test (pLDDT) score of MPXV F13 was 92.22 (Fig. 1b), and most pLDDTs of amino acid residues reached a confidential score, indicating the predicted F13 protein structure is highly accurate. Next, the CurPocket tool, a web-based protein-surface-curvature-based cavity detection approach, of CB-Dock2 was used to detect cavities on the predicted monkeypox virus F13 protein structure. The top 20 pockets were found and prepared as docking configs (pocket 1 shown in Fig. 1c, Table S1).

Then, the docking between tecovirimat and the above-mentioned MPXV F13 model was conducted to verify the accuracy of pocket detecting based on CB-Dock2, a protein-ligand blind docking web server based on AutoDock Vina (version 1.1.2). We found the docking between tecovirimat and MPXV F13 protein gets a better Vina score in pocket 1 (Fig. 1e, f) than in pocket 10 (Fig. 1d), the binding pose previously described by Daoqun Li et al. (−8.5 vs −8.2) and similar to the value Lam et al. reported (−8.4). A 50 ns molecular dynamics simulation of the tecovirimat-F13 complex was used to validate the binding stability of the docking pose by Gromacs (version 2019.3). And the RMSD and RMSF curves suggest the tecovirimat-F13 complex retained as a whole (Fig. 1j, k, green line).

We used DrugRep, an AutoDock Vina-based online virtual screening server, to screen 10,640 compounds (workflow showed in Fig. 1a), containing 3 libraries: FDA-approved drugs (N = 2315), experimental drugs (N = 5935), and traditional Chinese medicine (TCM) monomer compounds (N = 2390). The approved drug library and experimental drug library were accessed from the DrugBank database (version 5.1.7). And TCM library was obtained from Topscience Company, from ∼800 traditional Chinese medicines. 406 compounds were found to have a better or equal binding ability with MPXV F13 protein (Vina score ≤−8.5), including 86 approved drugs, 186 experimental drugs, and 134 monomer compounds from traditional Chinese medicine (top 20 s were listed in Table 1, Table S2). Tucatinib and ubrogepant et al. from the approved drug library, zk-806,450 et al. from the experimental drug library, and protohypericin, hypericin, and bilobetin et al. from the TCM library showed good binding abilities with F13 protein of monkeypox (Table 1). In our result, hypericin and naldemedine, found by Lam et al., were validated (−10.7 and −8.7 respectively).

Furthermore, Among those 406 compounds, some approved antiviral drugs displayed a high affinity with MPXV F13 protein. Glecaprevir, an anti-hepatitis C virus (HCV) drug that inhibits NS3/4A protease of HCV, was dosed as an oral combination therapy with the NS5A inhibitor pibrentasvir. The vina binding score for glecaprevir and F13 protein complex was −10.1 (Fig. 1g), suggesting a more stable binding than the tecovirimat-F13 complex. Dolutegravir, a second-generation integrase strand transfer inhibitor, was originally developed for the treatment of human immunodeficiency virus (HIV). The vina score of the dolutegravir-F13 complex was −9.1, showing good potential as an anti-monkeypox virus medicine (Fig. 1h). Bictegravir, another second-generation integrase strand transfer inhibitor approved for HIV treatment in fixed-dose combination with emtricitabine and tenofovir alafenamide, demonstrated to be a potential candidate for drug repurposing to be an anti-MPXV therapy for its bictegravir-F13 complex vina score was −9.0 (Fig. 1i). To validate the stability of F13 protein binding with glecaprevir, dolutegravir, or bictegravir, a molecular dynamics simulation of the complexes was conducted for 50 ns. And the results showed all 3 protein-drug complexes remained as a whole as the tecovirimat-F13 complex (Fig. 1j, k). Our virtual drug screening work found that many approved drugs can be repurposed for the treatment of monkeypox potentially. Those approved drugs have been studied comprehensively in pre-clinical research and clinical trials for their pharmacokinetics, pharmacodynamics, and toxicology. During the rapid global epidemic of the monkeypox virus, fast track of drug discovery and development can be vital for MPXV patients. Clinical studies for off-label use (unapproved use of an approved drug) of those drugs can be considered after ethical approval for those areas where effective treatment, especially tecovirimat and cidofovir et al., is not accessible.

In this work, we have predicted the structure of the monkeypox F13 protein and detected the potential binding pockets of the predicted protein structure. The docking of tecovirimat and F13 was verified. Then 10,640 compounds of the approved drug library, the experimental library, and the TCM library were virtual screened to dock F13 protein. Approved anti-viral medicines, including glecaprevir, dolutegravir, and bictegravir, other approved medicines, i.e. tucatinib and ubrogepant, and zk-806,450 et al. from experimental drug library, and protohypericin, hypericin, and bilobetin et al. from TCM library be indicated can be potentially repurposed for the treatment of MPXV.