key: cord-0961861-2aaz3zo1 authors: Zarn, Katherine E.; Jaramillo, Sierra A.; Zapata, Anthony R.; Stone, Nathan E.; Jones, Ashley; Nunnally, Haley; Settles, Erik W.; Ng, Ken; Keim, Paul; Knudsen, Steen; Nuijten, Patricia; Tijsma, Aloys; French, Christopher T. title: Stenoparib, an inhibitor of cellular poly (ADP-ribose) polymerases (PARPs), blocks in vitro replication of SARS-CoV-2 variants date: 2021-11-05 journal: bioRxiv DOI: 10.1101/2021.11.03.467186 sha: a97395d8e6b0ca557307e663e9ffcd09dafbd018 doc_id: 961861 cord_uid: 2aaz3zo1 We recently published a preliminary assessment of the activity of a poly (ADP-ribose) polymerase (PARP) inhibitor, stenoparib, also known as 2X-121, which inhibits viral replication by affecting pathways of the host. Stenoparib is an inhibitor of mammalian poly (ADP-ribose) polymerases (PARPs). Here we show that stenoparib effectively inhibits additional SARS-CoV-2 variants, including an additional wild-type strain (Germany/BavPat1/2020), and the variants alpha (B.1.1.7), beta (B.1.351) and gamma (P.1) in vitro, with 50% effective concentration (EC50) estimates of 4.1 μM, 8.5 μM, 24.2 μM and 13.6 μM, respectively. A second study focusing on a combination of 10 μM stenoparib and 0.5 μM remdesivir resulted in over 90% inhibition of the alpha (B.1.1.7) variant, which is substantially greater than what was achieved with stenoparib or remdesivir alone at these concentrations. Importance The coronavirus disease (COVID-19) pandemic, caused by SARS-CoV-2, has caused over 247 million infections and over 5 million deaths (1). Although protective vaccines are available, the pandemic continues and both old and new SARS-CoV-2 variants may exhibit degrees of resistance to vaccination. To date, only two antiviral drugs, remdesivir and molnupiravir, or treatment with monoclonal antibodies, have been approved by the United States Food and Drug Administration as COVID-19 therapies in certain situations. Additional effective therapeutics are urgently needed. Here we describe the activity of a small molecule, stenoparib, that effectively inhibits replication of SARS-CoV-2 wild-type and variant strains in vitro. Stenoparib is an inhibitor of mammalian poly (ADP-ribose) polymerases (PARPs). A host-targeting therapeutic like stenoparib could be a significant benefit for COVID-19 patients as a standalone therapy, or especially as part of a combinatorial COVID-19 treatment strategy with an antiviral drug such as remdesivir or molnupiravir. The 2 controls included on all assay plates were 1) infected and untreated cells and 2) 73 uninfected and untreated cells. The experiment was conducted in quadruplicate and the results 74 were calculated as the average fraction of wells that showed positive staining for the virus using 75 ViroSpot assay (10). Inhibition of virus replication by stenoparib was dose-dependent, with EC50 76 ranging between 4.1 μM to 24.2 μM across the 4 variants tested (Fig. 1A) . Interestingly, the 77 EC50 for the beta variant (24.2 μM) was 1.8-to 5.8-fold higher than for the wt, alpha, and 78 gamma variants (4.1, 8.5, and 13.6 μM, respectively). A similar phenomenon was noted for 79 remdesivir (Fig. 1B) , where the EC50 for the beta variant (9.9 μM) was 2.0-to 2.6-fold higher 80 than for the wt, alpha, and gamma variants (3.8, 5.0, and 4.3 μM, respectively). According to a 81 recent report, the beta variant appears to exhibit a significantly reduced eclipse period and more 82 rapid replication in vitro (11), which would likely affect its dose response toward stenoparib and (13). For the analysis shown in Fig. 2A and 2B, we 99 combined lower doses of stenoparib (2.5, 5.0, and 10 μM) with the previously reported 50% 100 effective concentration (EC50) of remdesivir (0.5 μM) (6). In Fig. 2A Prevention USCfDCa. 2020. 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Alpha: 221 USA/CA_CDC_5574/2020, BEI Resources, Cat# NR-54011 Gamma: hCoV-19/Japan/TY7-503/2021 (Brazil 223 P.1), BEI Resources