key: cord-0877575-86stuueg authors: Milewska, Aleksandra; Chi, Ying; Szczepanski, Artur; Barreto-Duran, Emilia; Liu, Kevin; Liu, Dan; Guo, Xiling; Ge, Yiyue; Li, Jingxin; Cui, Lunbiao; Ochman, Marek; Urlik, Maciej; Rodziewicz-Motowidlo, Sylwia; Zhu, Fengcai; Szczubialka, Krzysztof; Nowakowska, Maria; Pyrc, Krzysztof title: HTCC as a highly effective polymeric inhibitor of SARS-CoV-2 and MERS-CoV date: 2020-03-31 journal: bioRxiv DOI: 10.1101/2020.03.29.014183 sha: 144fc2c4592474850fc63a415fdd26f745f7a1f3 doc_id: 877575 cord_uid: 86stuueg The beginning of 2020 brought us information about the novel coronavirus emerging in China. Rapid research resulted in the characterization of the pathogen, which appeared to be a member of the SARS-like cluster, commonly seen in bats. Despite the global and local efforts, the virus escaped the healthcare measures and rapidly spread in China and later globally, officially causing a pandemic and global crisis in March 2020. At present, different scenarios are being written to contain the virus, but the development of novel anticoronavirals for all highly pathogenic coronaviruses remains the major challenge. Here, we describe the antiviral activity of previously developed by us HTCC compound (N-(2-hydroxypropyl)-3-trimethylammonium chitosan chloride), which may be used as potential inhibitor of currently circulating highly pathogenic coronaviruses – SARS-CoV-2 and MERS-CoV. pathogenic coronaviruses -SARS-CoV-2 and MERS-CoV. INTRODUCTION Coronaviruses mainly cause respiratory and enteric diseases in humans, other mammals, 50 and birds. However, some species can cause more severe conditions such as hepatitis, 51 peritonitis, or neurological disease. Seven coronaviruses infect humans, four of which (human pangolins were suggested as such 9 . The virus is associated with a respiratory illness that, in a 63 proportion of cases, is severe. The mortality rate varies between locations, but at present, is 64 estimated to reach 3-4% globally. The virus infects primarily ciliated cells and type II 65 pneumocytes in human airways, hijacking the angiotensin-converting enzyme 2 (ACE2) to 66 enter the cell, similarly as SARS-CoV and HCoV-NL63. 67 MERS-CoV is related to SARS-CoV-2, but together with some bat viruses forms 68 a separate Merbecovirus subgenus. Bats are believed to serve as an original reservoir also in 69 this case 10 , but camels were identified as the intermediate host 11 . The virus never fully crossed 70 the species border, as the human-to-human transmission is limited, and almost all the cases are 71 associated with animal-to-human transmission. The entry receptor for MERS-CoV is the 72 dipeptidyl peptidase 4 (DPP4) 12, 13 . In humans, MERS-CoV causes a respiratory illness with severity varying from asymptomatic to potentially fatal acute respiratory distress 14-16 . To date, 74 MERS-CoV infection was confirmed in 27 countries, with over 2,000 cases and a mortality rate 75 of ~35%. 76 Currently, there are no vaccines or drugs with proven efficacy to treat coronavirus 77 infection, and treatment is limited to supportive care. However, a range of therapeutics have Consequently, broad-spectrum antivirals are essential in long-term perspective. we dissected the mechanism of the HTCC antiviral activity. We showed that the polymer 96 interacts with the coronaviral Spike (S) protein and blocks its interaction with the cellular 97 receptor 22-24 . Here, we analyzed the HTCC activity against SARS-CoV-2 and MERS-CoV in 98 vitro using permissive cell lines and ex vivo, using a model of human airway epithelium (HAE). 99 The study showed that the replication of both viruses was efficiently hampered. Overall, our 100 data show that HTCC polymers are potent broad-spectrum anticoronavirals and may be very 101 promising drug candidates for SARS-CoV-2 and MERS-CoV. 105 Previously, we showed that HTCC with different degrees of substitution (DSs) is a Next, the dose-dependence was tested for the HTCCs. The inhibitory activity of selected 134 polymers was verified for three different concentrations, and obtained data are shown in Figure 135 2. Based on the data obtained, the basic parameters were calculated and are presented in Table 1 . The parameters observed for the SARS-CoV-2 appear to be favorable with SI above 12. It is 148 also worth to note that HTCC was previously administered by inhalation in rats, and no adverse 149 reactions were observed 25 . In that study, HTCC was used as a carrier for the active substance, 150 and as such, was reported to be promising for local sustained inhalation therapy of pulmonary 151 diseases. Our previous research showed that the HTCC-mediated inhibition of coronaviral The active compound 257 The HTCC was prepared in the same manner as previously described 22,23,28 . The severe acute respiratory syndrome Middle East respiratory syndrome coronavirus (MERS-CoV): 414 announcement of the Coronavirus Study Group Isolation 417 of a novel coronavirus from a man with pneumonia in Saudi Arabia Identification of a new human coronavirus Croup is associated with the novel coronavirus NL63 A Novel Coronavirus from Patients with Pneumonia in China The proximal origin of SARS-CoV-2 Rooting the phylogenetic tree of middle East respiratory syndrome 429 coronavirus by characterization of a conspecific virus from an African bat Evolutionary Dynamics of MERS-CoV: Potential Recombination Molecular basis of binding between novel human coronavirus MERS-CoV and its 434 receptor CD26 Dipeptidyl peptidase 4 is a functional receptor for the emerging human 436 coronavirus-EMC MERS coronavirus: diagnostics, epidemiology and transmission The emergence of the Middle East respiratory 440 syndrome coronavirus Tropism and replication of Middle East respiratory syndrome coronavirus 442 from dromedary camels in the human respiratory tract: an in-vitro and ex-vivo study Coronavirus Susceptibility to the Antiviral Remdesivir (GS-5734) Is 445 Mediated by the Viral Polymerase and the Proofreading Exoribonuclease A mouse model for Betacoronavirus subgroup 2c using a bat coronavirus 448 strain HKU5 variant A noncovalent class of papain-like protease/deubiquitinase inhibitors blocks 450 SARS virus replication Enhancement of the infectivity of SARS-CoV in BALB/c mice by IMP 453 dehydrogenase inhibitors, including ribavirin Severe acute respiratory syndrome coronavirus 456 as an agent of emerging and reemerging infection HTCC: Broad Range Inhibitor of Coronavirus Entry Novel polymeric inhibitors of HCoV-NL63 THE USE OF CHITOSAN 463 POLYMER IN THE TREATMENT AND PREVENTION OF INFECTIONS CAUSED BY 464 CORONAVIRUSES Cyclosporine A/porous quaternized chitosan microspheres as a novel 466 pulmonary drug delivery system Katarzyna Owczarek 471 , Marek Ochman , Tomasz Stacel View ORCID Profile. Cell Encapsulation Technology 472 and Therapeutics Biopolymeric 474 nano/microspheres for selective and reversible adsorption of coronaviruses Human coronavirus NL63 utilizes heparan sulfate proteoglycans for 477 attachment to target cells This work was supported by the subsidy from the Polish Ministry of Science and Higher 385 Education for the research on the SARS-CoV-2 and a grant from the National Science Center 386 UMO-2017/27/B/NZ6/02488 to KP. The funders had no role in study design, data collection, and analysis, decision to publish, or 388 preparation of the manuscript. The technology is owned by the Jagiellonian University (Krakow, Poland) and protected 390 by a patent no WO2013172725A1 and associated documents.