Graphene-based materials as nanoplatforms for antiviral therapy and prophylaxis.

INTRODUCTION: The dramatic effects caused by viral diseases have prompted the search for effective therapeutic and preventive agents. In this context, 2D graphene-based nanomaterials (GBNs) have shown great potential for antiviral therapy, enabling the functionalization and/or decoration with biomolecules, metals and polymers, able to improve their interaction with viral nanoparticles. AREAS COVERED: This review summarizes the most recent advances of the antiviral research related to 2D GBNs, based on their antiviral mechanism of action. Their ability to inactivate viruses by inhibiting the entry inside cells, or through drug/gene delivery, or by stimulating the host immune response are here discussed. As reported, biological studies performed in vitro and/or in vivo allowed to demonstrate the antiviral activity of the developed GBNs, at different stages of the virus life cycle and the evaluation of their long-term toxicity. Other mechanisms closely related to the physicochemical properties of GBNs are also reported, demonstrating the potential of these materials for antiviral prophylaxis. EXPERT OPINION: GBNs represent valuable tools to fight emerging or reemerging viral infections. However, their translation into the clinic requires standardized scale-up procedures leading to the reliable and reproducible synthesis of these nanomaterials with suitable physicochemical properties, as well as more in-depth pharmacological and toxicological investigations. We believe that multidisciplinary approaches will give valuable solutions to overcome the encountered limitations in the application of GBNs in biomedical and clinical field.

Design, synthesis, and structure-activity relationships of a novel class of quinazoline derivatives as coronavirus inhibitors.

There remain great unmet needs to treat coronavirus infections in clinic, and the development of novel antiviral agents is highly demanded. In this work, a phenotypic screening against our in-house compound library identified several cajanine derivatives with moderate antiviral activity against HCoV-OC43. Based on the scaffold of cajanine, a series of quinazoline derivatives were designed employing a scaffold-hopping strategy. After an iterative structural optimization campaign, several quinazoline derivatives with potent antiviral efficacy (EC50: ∼0.1 µM) and high selectivity (SI > 1000) were successfully identified. The preliminary mechanism of action study indicated that such quinazoline derivatives functioned at the early stage of infection. In aggregate, this work delivered a new chemical type of coronavirus inhibitors, which could be employed not only for further development of antiviral drugs but also as important chemical tools to delineate the target of action.

In silico screening of FDA approved drugs against ACE2 receptor: potential therapeutics to inhibit the entry of SARS-CoV-2 to human cells.

An unknown coronavirus that emerged sometime at the end of 2019 in China, the novel SARS-CoV-2, now called COVID-19, has spread all over the world. Several efforts have been made to prevent or treat this disease, though not with success. The initiation of COVID-19 viral infection involves specific binding of SARS-CoV-2 to the host surface of the receptor, ACE2. The ACE2- SARS-CoV-2 complex then gets transferred into the endosomes where the endosomal acidic proteases cleave the S protein present in SARS-CoV-2, activating its fusion and release of the viral genome. We have carried out detailed and thorough in silico studies to repurpose FDA approved compounds to inhibit human ACE2 receptor so as to prevent the viral entry. Our study reveals that five compounds show good binding to the ACE2 receptor and hence are potential candidates to interact with ACE2 and prevent it's recognition by the virus, SARS-CoV-2. Communicated by Ramaswamy H. Sarma.

Identification of SARS-CoV-2 entry inhibitors among already approved drugs.

To discover effective drugs for COVID-19 treatment amongst already clinically approved drugs, we developed a high throughput screening assay for SARS-CoV-2 virus entry inhibitors using SARS2-S pseudotyped virus. An approved drug library of 1800 small molecular drugs was screened for SARS2 entry inhibitors and 15 active drugs were identified as specific SARS2-S pseudovirus entry inhibitors. Antiviral tests using native SARS-CoV-2 virus in Vero E6 cells confirmed that 7 of these drugs (clemastine, amiodarone, trimeprazine, bosutinib, toremifene, flupenthixol, and azelastine) significantly inhibited SARS2 replication, reducing supernatant viral RNA load with a promising level of activity. Three of the drugs were classified as histamine receptor antagonists with clemastine showing the strongest anti-SARS2 activity (EC50 = 0.95 ± 0.83 µM). Our work suggests that these 7 drugs could enter into further in vivo studies and clinical investigations for COVID-19 treatment.

An Oleanolic Acid Derivative Inhibits Hemagglutinin-Mediated Entry of Influenza A Virus.

Influenza A viruses (IAV) have been a major public health threat worldwide, and options for antiviral therapy become increasingly limited with the emergence of drug-resisting virus strains. New and effective anti-IAV drugs, especially for highly pathogenic influenza, with different modes of action, are urgently needed. The influenza virus glycoprotein hemagglutinin (HA) plays critical roles in the early stage of virus infection, including receptor binding and membrane fusion, making it a potential target for the development of anti-influenza drugs. In this study, we show that OA-10, a newly synthesized triterpene out of 11 oleanane-type derivatives, exhibited significant antiviral activity against four different subtypes of IAV (H1N1, H5N1, H9N2 and H3N2) replications in A549 cell cultures with EC50 ranging from 6.7 to 19.6 µM and a negligible cytotoxicity (CC50 > 640 µM). It inhibited acid-induced hemolysis in a dose-dependent manner, with an IC50 of 26 µM, and had a weak inhibition on the adsorption of H5 HA to chicken erythrocytes at higher concentrations (≥40 µM). Surface plasmon resonance (SPR) analysis showed that OA-10 interacted with HA in a dose-dependent manner with the equilibrium dissociation constants (KD) of the interaction of 2.98 × 10-12 M. Computer-aided molecular docking analysis suggested that OA-10 might bind to the cavity in HA stem region which is known to undergo significant rearrangement during membrane fusion. Our results demonstrate that OA-10 inhibits H5N1 IAV replication mainly by blocking the conformational changes of HA2 subunit required for virus fusion with endosomal membrane. These findings suggest that OA-10 could serve as a lead for further development of novel virus entry inhibitors to prevent and treat IAV infections.

Second generation of diazachrysenes: Protection of Ebola virus infected mice and mechanism of action.

Ebola virus (EBOV) causes a deadly hemorrhagic fever in humans and non-human primates. There is currently no FDA-approved vaccine or medication to counter this disease. Here, we report on the design, synthesis and anti-viral activities of two classes of compounds which show high potency against EBOV in both in vitro cell culture assays and in vivo mouse models Ebola viral disease. These compounds incorporate the structural features of cationic amphiphilic drugs (CAD), i.e they possess both a hydrophobic domain and a hydrophilic domain consisting of an ionizable amine functional group. These structural features enable easily diffusion into cells but once inside an acidic compartment their amine groups became protonated, ionized and remain trapped inside the acidic compartments such as late endosomes and lysosomes. These compounds, by virtue of their lysomotrophic functions, blocked EBOV entry. However, unlike other drugs containing a CAD moiety including chloroquine and amodiaquine, compounds reported in this study display faster kinetics of accumulation in the lysosomes, robust expansion of late endosome/lysosomes, relatively more potent suppression of lysosome fusion with other vesicular compartments and inhibition of cathepsins activities, all of which play a vital role in anti-EBOV activity. Furthermore, the diazachrysene 2 (ZSML08) that showed most potent activity against EBOV in in vitro cell culture assays also showed significant survival benefit with 100% protection in mouse models of Ebola virus disease, at a low dose of 10 mg/kg/day. Lastly, toxicity studies in vivo using zebrafish models suggest no developmental defects or toxicity associated with these compounds. Overall, these studies describe two new pharmacophores that by virtue of being potent lysosomotrophs, display potent anti-EBOV activities both in vitro and in vivo animal models of EBOV disease.

Potent influenza A virus entry inhibitors targeting a conserved region of hemagglutinin.

Influenza A viruses (IAVs) induce acute respiratory disease and cause significant morbidity and mortality throughout the world. With the emergence of drug-resistant viral strains, new and effective anti-IAV drugs with different modes of action are urgently needed. In this study, by conjugating cholesterol to the N-terminus of the short peptide KKWK, a lipopeptide named S-KKWK was created. The anti-IAV test indicated that S-KKWK and its derivatives displayed potent antiviral activities against a broad variety of influenza A viral strains including oseltamivir-resistant strains and clinically relevant isolates with IC50 values ranging from 0.7 to 3.0µM. An extensive mechanistic study showed that these peptides functioned as viral "entry blockers" by inhibiting the conformational rearrangements of HA2 subunit, thereby interrupting the fusion of virus-host cell membranes. Significantly, a computer-aided docking simulation and protein sequence alignment identified conserved residues in the stem region of HA2 as the possible binding site of S-KKWK, which may be employed as a potential drug target for designing anti-IAVs with a broad-spectrum of activity. By targeting this region, a potent anti-IAV agent was subsequently created. In addition, the anti-IAV activity of S-KKWK was assessed by experiments with influenza A virus-infected mice, in which S-KKWK reduced the mortality of infected animals and extended survival time significantly. Overall, in addition to providing a strategy for designing broad-spectrum anti-IAV agents, these results indicate that S-KKWK and its derivatives are prospective candidates for potent antivirals.

Defeating enemy outside the border-development of viral entry inhibitors / 复旦学报（医学版）

About 25 years ago,we had pioneered the research fields of developing human immunodeficiency virus (HIV) fusion/entry inhibitors and anti-HIV peptide drugs.Over the past six years,we have gained some promising results in research and development of the HIV,Middle East respiratory syndrome coronavirus (MERS-COV),the Ebola virus (EBOV),and the Zika virus (ZIKV) entry inhibitors.This article provides an overview of the research progress of viral entry inhibitors against the related highly pathogenic viruses.