key: cord-0802048-9tls06d2 authors: Lee, Jihye; Lee, JinAh; Kim, Hyeon Ju; Ko, Meehyun; Jee, Youngmee; Kim, Seungtaek title: TMPRSS2 and RNA-dependent RNA polymerase are effective targets of therapeutic intervention for treatment of COVID-19 caused by SARS-CoV-2 variants (B.1.1.7 and B.1.351) date: 2021-04-08 journal: bioRxiv DOI: 10.1101/2021.04.06.438540 sha: bbee94858d3d3f2948c3baae194e1f3df91f6c6c doc_id: 802048 cord_uid: 9tls06d2 SARS-CoV-2 is a causative agent of COVID-19 pandemic and the development of therapeutic interventions is urgently needed. So far, monoclonal antibodies and drug repositioning are the main methods for drug development and this effort was partially successful. Since the beginning of COVID-19 pandemic, the emergence of SARS-CoV-2 variants has been reported in many parts of the world and the main concern is whether the current vaccines and therapeutics are still effective against these variant viruses. The viral entry and viral RNA-dependent RNA polymerase (RdRp) are the main targets of current drug development, thus the inhibitory effects of TMPRSS2 and RdRp inhibitors were compared among the early SARS-CoV-2 isolate (lineage A) and the two recent variants (lineage B.1.1.7 and lineage B.1.351) identified in the UK and South Africa, respectively. Our in vitro analysis of viral replication showed that the drugs targeting TMPRSS2 and RdRp are equally effective against the two variants of concern. COVID-19 is an emerging infectious disease caused by a novel coronavirus, severe acute 43 respiratory syndrome coronavirus 2 (SARS-CoV-2) (1) and it was declared as a pandemic by 44 the World Health Organization (WHO) on March 11, 2020. In order to address this 45 unprecedented global challenge, intensive investigations have been simultaneously conducted 46 by global scientific communities and industries to develop diagnostic tools, vaccines, and 47 therapeutics. Remarkably, within ten months after release of the SARS-CoV-2 genome 48 sequence, a couple of vaccines were successfully developed and are now being used for 49 vaccination of people after emergency use authorization (EUA). Drug development was also 50 partially successful, especially in the development of monoclonal antibodies (2)(3). Notably, 51 the vaccines and monoclonal antibodies currently being used are heavily dependent on the 52 structure and sequence of viral Spike protein, which is a surface glycoprotein responsible for 53 virus entry by interacting with the host receptor, angiotensin-converting enzyme 2 (ACE2). 54 Thus, if there is any mutation in this protein, it is likely to affect the efficacy of both vaccines 55 and antibodies. Since the beginning of COVID-19 pandemic, variants of SARS-CoV-2 have been reported in 57 many parts of the world and the recent variants identified in the UK (lineage B.1.1.7), South 58 Africa (lineage B.1.351), and Brazil (lineage P.1) are of particular concern due to multiple 59 mutations in the Spike gene ( Figure 1 ) (4)(5). Indeed, several results are being published, 60 which demonstrated reduced neutralization capacity of convalescent plasma, vaccine sera, 61 and monoclonal antibodies against these variants (6)(7)(8)(9). In addition to monoclonal antibodies, small molecule inhibitors are also being developed as 63 potential antiviral agents. Targets of such small molecule inhibitors are often transmembrane 64 serine protease 2 (TMPRSS2) (10)(11)(12)(13) and viral RNA-dependent RNA polymerase 65 (RdRp) (14) (15). TMPRSS2 is known to possess serine protease activity, which primes the 66 viral Spike protein for fusion between the viral membrane and the host cell membrane prior 67 to the release of viral genome into the cytoplasm. Camostat and nafamostat are representative 68 drug candidates as TMPRSS2 inhibitors and currently being tested in several phase 2 and 3 69 clinical trials in many countries. On the other hand, RdRp is a target of remdesivir, which is 70 the first approved drug for treatment of COVID-19 patients (16). In this study, we investigated whether the antiviral drug candidates targeting TMPRSS2 and 72 RdRp are still effective against the recent SARS-CoV-2 variants of concern by assessing in 73 vitro viral replication capacity after drug treatment. The amino acid sequence of NSP12 was also well conserved among the three lineages of 102 SARS-CoV-2 ( Figure 1 ) and we did not find any substantial differences among them with 103 regard to drug efficacy of the two representative RdRp inhibitors (remdesivir and 104 molnupiravir) (Figures 2 and 3) . Both remdesivir and molnupiravir are nucleoside analogs 105 but the two drugs differ from each other in that remdesivir works as a chain terminator but In summary, we analyzed efficacy of potential drug candidates (i.e., TMPRSS2 inhibitors, 127 RdRp inhibitors and others) against the recent SARS-CoV-2 variants of concern and we 128 found that all of them were equally effective in suppressing replication of B.1.1.7 and A pneumonia outbreak associated with a new coronavirus 199 of probable bat origin SARS-CoV-2 Neutralizing Antibody LY-CoV555 in Outpatients with Covid-19 REGN-COV2, a Neutralizing Antibody Cocktail, 210 in Outpatients with Covid-19 SARS-CoV-2 212 variants and ending the COVID-19 pandemic SARS-CoV-2 Viral Variants-Tackling a 214 Moving Target Comprehensive mapping of mutations in the SARS-CoV-2 receptor-binding 217 domain that affect recognition by polyclonal human plasma antibodies Identification of SARS-CoV-2 spike mutations that 222 attenuate monoclonal and serum antibody neutralization Circulating SARS-CoV-2 spike N439K variants maintain fitness while evading 234 antibody-mediated immunity 2021. mRNA vaccine-elicited antibodies to SARS-CoV-2 and 241 circulating variants SARS-CoV-2 Cell Entry Depends on ACE2 and TMPRSS2 and Is Blocked by a 245 Clinically Proven Protease Inhibitor Nafamostat Mesylate Blocks Activation of SARS-CoV-2: New Treatment 248 Option for COVID-19 The Anticoagulant Nafamostat Potently Inhibits SARS-CoV-2 S 252 Fusion in a Cell Fusion Assay System and Viral Infection In Vitro in 253 a Cell-Type-Dependent Manner Comparative analysis of antiviral efficacy of FDA-approved drugs against SARS-CoV-2 in human lung cells Broad-spectrum antiviral GS-5734 inhibits both epidemic and 261 zoonotic coronaviruses An orally bioavailable broad-spectrum antiviral inhibits 267 SARS-CoV-2 in human airway epithelial cell cultures and multiple coronaviruses in 268 mice Remdesivir for the Treatment of Covid-19 -Final Report TMPRSS2: A potential target 278 for treatment of influenza virus and coronavirus infections Identification of 280 antiviral drug candidates against SARS-CoV-2 from FDA-approved drugs tropism, entry, replication, and propagation: Considerations for drug discovery and 285 11 development Identification of a New Ribonucleoside Inhibitor of Ebola Virus Replication 2018. β-291 d-N4-Hydroxycytidine Is a Potent Anti-alphavirus Compound That Induces a High 292 Level of Mutations in the Viral Genome Characterization of orally efficacious 296 influenza drug with high resistance barrier in ferrets and human airway epithelia Hydroxycytidine Inhibits a Proofreading-Intact Coronavirus with a High Genetic 302 Barrier to Resistance