Zika virus (ZIKV) is one of the mosquito-borne flaviviruses that exhibits a unique tropism to nervous systems and is associated with Guillain-Barre syndrome and congenital Zika syndrome (CZS). Dengue virus (DENV) and yellow fever virus (YFV), the other two mosquito-borne flaviviruses, have also been circulating for a long time and cause severe diseases, such as dengue hemorrhagic fever and yellow fever, respectively. However, there are no safe and effective antiviral drugs approved for the treatment of infections or coinfections of these flaviviruses. Here, we found that zafirlukast, a pregnancy-safe leukotriene receptor antagonist, exhibited potent antiviral activity against infections of ZIKV strains from different lineages in different cell lines, as well as against infections of DENV-2 and YFV 17D. Mechanistic studies demonstrated that zafirlukast directly and irreversibly inactivated these flaviviruses by disrupting the integrity of the virions, leading to the loss of viral infectivity, hence inhibiting the entry step of virus infection. Considering its efficacy against flaviviruses, its safety for pregnant women, and its neuroprotective effect, zafirlukast is a promising candidate for prophylaxis and treatment of infections or coinfections of ZIKV, DENV, and YFV, even in pregnant women.

This study aimed to evaluate diagnostic accuracy of SARS-CoV-2 RNA detection in saliva samples treated with a guanidine-based or guanidine-free inactivator, using nasopharyngeal swab samples (NPS) as referents. Based on the NPS reverse transcription-polymerase chain reaction (RT-PCR) results, participants were classified as with or without COVID-19. Fifty sets of samples comprising NPS, self-collected raw saliva, and saliva with a guanidine-based, and guanidine-free inactivator were collected from each group. In patients with COVID-19, the sensitivity of direct RT-PCR using raw saliva and saliva treated with a guanidine-based and guanidine-free inactivator was 100.0%, 65.9%, and 82.9%, respectively, with corresponding concordance rates of 94.3% (κ=88.5), 82.8% (κ=64.8), and 92.0% (κ=83.7). Among patients with a PCR Ct value of <30 in the NPS sample, the positive predictive value for the three samples was 100.0%, 80.0%, and 96.0%, respectively. The sensitivity of SARS-CoV-2 RNA detection was lower in inactivated saliva than in raw saliva and lower in samples treated with a guanidine-based than with a guanidine-free inactivator. However, in individuals contributing to infection spread, inactivated saliva showed adequate accuracy regardless of the inactivator used. Inactivators can be added to saliva samples collected for RT-PCR to reduce viral transmission risk while maintaining adequate diagnostic accuracy.

Time-dependent inactivation (TDI) of cytochrome P450 (CYP) enzymes may result in clinical drug-drug interactions (DDIs). Therefore, designing out of CYP TDI prior to advancing a compound to clinical development is highly desirable. As TDI of CYP3A is a common occurrence in small molecule drug discovery, high-throughput methods are sought to help identify the mechanism of inactivation and enable design strategies to mitigate CYP3A TDI. CYP inactivation via modification or destruction of the prosthetic heme group results in loss of the ability of the enzyme to bind carbon monoxide. Additionally, formation of a tight binding complex with the heme iron, referred to as a metabolic intermediate (MI) complex, also results in enzyme inactivation. The methods described herein provide a high-throughput means of identifying and comparing compounds for their ability to inactivate via destruction/modification of the heme via loss of the ability to bind carbon monooxide, as well as via formation of an MI complex.

Dynamic regulation of integrin activation and inactivation is critical for precisely controlled cell adhesion and migration in physiological and pathological processes. The molecular basis for integrin activation has been intensively studied; however, the understanding of integrin inactivation is still limited. Here, we identify LRP12 as an endogenous transmembrane inhibitor for α4 integrin activation. The LRP12 cytoplasmic domain directly binds to the integrin α4 cytoplasmic tail and inhibits talin binding to the β subunit, thus keeping integrin inactive. In migrating cells, LRP12-α4 interaction induces nascent adhesion (NA) turnover at the leading-edge protrusion. Knockdown of LRP12 leads to increased NAs and enhanced cell migration. Consistently, LRP12-deficient T cells show an enhanced homing capability in mice and lead to aggravated chronic colitis in a T cell-transfer colitis model. Altogether, LRP12 is a transmembrane inactivator for integrins that inhibits α4 integrin activation and controls cell migration by maintaining balanced NA dynamics.

In addition to antibacterial effects, macrolide antibiotics exhibit other extensive pharmacological effects, such as anti-inflammatory and antiviral activities. Erythromycin estolate, one of the macrolide antibiotics, was previously investigated to effectively inhibit infections of various flaviviruses including Zika virus, dengue virus, and yellow fever virus, but its antiviral effect against human coronavirus remains unknown. Thus, the current study was designed to evaluate the antiviral efficacy of erythromycin estolate against human coronavirus strain OC43 (HCoV-OC43) and to illustrate the underlying mechanisms. Erythromycin estolate effectively inhibited HCoV-OC43 infection in different cell types and significantly reduced virus titers at safe concentration without cell cytotoxicity. Furthermore, erythromycin estolate was identified to inhibit HCoV-OC43 infection at the early stage and to irreversibly inactivate virus by disrupting the integrity of the viral membrane whose lipid component might be the target of action. Together, it was demonstrated that erythromycin estolate could be a potential therapeutic drug for HCoV-OC43 infection.

The study investigated the inhibitory activity of protocetraric and salazinic acids against SARS-CoV-2 3CLpro. The kinetic parameters were determined by microtiter plate-reading fluorimeter using a fluorogenic substrate. The cytotoxic activity was tested on murine Sertoli TM4 cells. In silico analysis was performed to ascertain the nature of the binding with the 3CLpro. The compounds are slow-binding inactivators of 3CLpro with a Ki of 3.95 μM and 3.77 μM for protocetraric and salazinic acid, respectively, and inhibitory efficiency kinact/Ki at about 3 × 10-5 s-1µM-1. The mechanism of inhibition shows that both compounds act as competitive inhibitors with the formation of a stable covalent adduct. The viability assay on epithelial cells revealed that none of them shows cytotoxicity up to 80 μM, which is well below the Ki values. By molecular modelling, we predicted that the catalytic Cys145 makes a nucleophilic attack on the carbonyl carbon of the cyclic ester common to both inhibitors, forming a stably acyl-enzyme complex. The computational and kinetic analyses confirm the formation of a stable acyl-enzyme complex with 3CLpro. The results obtained enrich the knowledge of the already numerous biological activities exhibited by lichen secondary metabolites, paving the way for developing promising scaffolds for the design of cysteine enzyme inhibitors.

In recent years, infectious diseases caused by viral infections have seriously endangered human health, especially COVID-19, caused by SARS-CoV-2, which continues to spread worldwide. The development of broad-spectrum antiviral inhibitors is urgently needed. Here, we report a series of small-molecule compounds that proved effective against human coronaviruses (HCoV), such as SARS-CoV-2 and its variants of concern (VOCs), including Alpha (B.1.1.7), Beta (B.1.351), Gamma (P.1), Delta (B.1.617.2), and Omicron (B.1.1.529), SARS-CoV, MERS-CoV, HCoV-OC43, and other viruses with class I viral fusion proteins, such as influenza virus, Ebola virus (EBOV), Nipah virus (NiV), and Lassa fever virus (LASV). They are also effective against class II enveloped viruses represented by ZIKV and class III enveloped viruses represented by vesicular stomatitis virus (VSV). Further studies have shown that these compounds may exert antiviral effects through a variety of mechanisms, including inhibiting the formation of the six-helix bundle, which is a typical feature of enveloped virus fusion with cell membranes, and/or targeting viral membrane to inactivate cell-free virions. These compounds are expected to become drug candidates against SARS-CoV-2 and other enveloped viruses.

Virus inactivator can inactivate cell-free virions without relying on their replication cycle, potentially reducing the impact of viral infection on cells. Previously, we successfully constructed a HIV-1 protein inactivator, 2DLT, by conjugating the D1D2 region of CD4 to the fusion inhibitor T1144 via a 35-amino acid linker. Therefore, it targets both the CD4 binding site in gp120 and NHR region in gp41. Considering that small-molecule agents have the advantages of fast production, low cost, good stability, and oral availability, we herein report the design of a new small-molecule HIV-1 inactivator, FD028, by conjugating FD016 (an analog of NBD-556, a gp120-CD4 binding inhibitor) with FD017 (an analog of 11d, an HIV-1 fusion inhibitor). The results showed that FD028 inactivated cell-free virions at a moderate nanomolar concentration by targeting both HIV-1 gp120 and gp41. Moreover, FD028 has broad-spectrum inhibition and inactivation activity against HIV-1 resistant strains and primary isolates of different subtypes without significant cytotoxicity. Therefore, FD028 has potential for further development as an HIV-1 inactivator-based therapeutic.

Bovine coronavirus (BCoV), a major causative pathogen of bovine enteric and respiratory diseases and a zoonotic pathogen transmissible between animals and humans, has led to severe economic losses in numerous countries. BCoV belongs to the genus Betacoronavirus, which is a model of a pathogen that is threatening human health and includes severe acute respiratory syndrome coronavirus (SARS-CoV), SARS-CoV-2, and Middle East respiratory syndrome coronavirus. This study aimed to determine whether photocatalytic material effectively reduces CoVs in the environment. Using the film adhesion method of photocatalytic materials, we assessed its antiviral activity and the effect of visible light irradiation according to methods defined by the International Organization for Standardization. Consequently, photocatalytic material was found to have antiviral activity, reducing the viral loads by 2.7 log TCID50 (tissue culture infective dose 50)/0.1 mL (500 lux), 2.8 log TCID50/0.1 mL (1000 lux), and 2.4 log TCID50/0.1 mL (3000 lux). Hence, this photocatalytic material might be applicable not only to reducing CoVs in the cattle breeding environment but also perhaps in other indoor spaces, such as offices and hospital rooms. To our knowledge, this study is the first to evaluate the antiviral activity of a photocatalytic material against CoV.

Recently, Zika virus (ZIKV) has attracted much attention in consideration of its association with severe neurological complications including fetal microcephaly. However, there are currently no prophylactic vaccines or therapeutic drugs approved for clinical treatments of ZIKV infection. To determine the potential anti-ZIKV inhibitors, we screened a library of clinical drugs with good safety profiles. Erythromycin estolate (Ery-Est), one of the macrolide antibiotics, was found to effectively inhibit ZIKV infection in different cell types and significantly protect A129 mice from ZIKV-associated neurological signs and mortality. Through further investigation, Ery-Est was verified to inhibit ZIKV entry by disrupting the integrity of the viral membrane which resulted in the loss of ZIKV infectivity. Furthermore, Ery-Est also showed inhibitory activity against dengue virus (DENV) and yellow fever virus (YFV). Thus, Ery-Est may be a promising drug for patients with ZIKV infection, particularly pregnant women.