3.2 Host-based drug research
3.2.1 Interferon and inducer
After a cell is infected with a virus, it then synthesizes and secretes a substance that interferes with virus replication and enhances the anti-viral ability of neighboring cells, which is called interferon. Human interferons are divided into type I and type II. The host's inherent interferon response is essential for controlling viral replication after infection. The interferon response can be enhanced by recombinant interferon or interferon inducers.
(1) Recombinant interferon α and interferon β can inhibit the replication of SARS-CoV and MERS coronavirus in vitro and animal models. Interferon β1b type has the best antiviral effect on MERS-CoV among various subtypes. Various combinations of interferon alpha or interferon beta with other antiviral drugs (such as ribavirin, lopinavir, and ritonavir) have been used to treat SARS or MERS infected patients.
(2) Polyinosinic acid is a synthetic analogue of dsRNA, which can strongly induce type I interferon, although studies have shown that MERS-CoV 4a protein contains inosine: a double-stranded RNA binding domain that interacts with polycytidylic acid, thereby inhibiting the production of interferon induced by polyinosinic acid: polycytidylic acid or Sendai virus, yet experiments show that polyinosinic acid: polycytic glycoside can still greatly reduce the MERS-CoV load in BALB/c mice.
(3) Nitazoxanide (NTZ) is another effective type I interferon inducer, which is currently used clinically to treat parasitic infections. It is a synthetic thiazolyl-salicylamide derivative. In cell culture assays, nitazoxanide can inhibit the replication of various RNA and DNA viruses, including respiratory syncytial virus, parainfluenza virus, coronavirus, Rotavirus, Norovirus, HBV, HCV, dengue virus, yellow fever, Japanese encephalitis virus, and HIV, which has broad-spectrum antiviral activity.
3.2.2 Cyclophilin inhibitor
Cyclophilin (Cyp) belongs to the family of peptidyl-prolyl isomerases (PPIase) and is used in the replication of RNA viruses (including human immunodeficiency virus 1, hepatitis C virus and influenza A virus). Cyclophilins are currently found in all cells of prokaryotes and eukaryotes. There are seven main types of cyclophilin in the body, namely cyclophilin A (CypA), cyclophilin B (CypB), cyclophilin C (CypC), cyclophilin D (CypD), and cyclophilin E CypE), cyclophilin 40 (Cyp40) and cyclophilin NK (CypNK), which are usually not connected to each other in the genome. Cyclophilin is a potential drug target for coronavirus. Among them, cyclophilin A is very important in human immunodeficiency virus type 1 (HIV-1), hepatitis C virus (HCV), vesicular stomatitis virus (VSV), human papilloma virus and vaccinia virus effect, and is the main cellular target of the immunosuppressive drug cyclosporin A (CsA). Cyclosporin A is a classic immunosuppressive drug that binds to cyclophilin to inhibit calcineurin phosphatase, thereby preventing the transfer of nuclear factors that activate T cells into the nucleus, and inhibiting the transcription of genes encoding cytokines such as interleukin-2. Cyclosporin A can inhibit the replication of almost all species of coronavirus in a dose-dependent manner in vitro, including SARS-CoV, MERS-CoV, human coronavirus 229E, feline coronavirus, avian infectious bronchitis virus, and mouse hepatitis virus, but only in the early stages of replication. As an effective and broad-spectrum coronavirus inhibitor, cyclosporin A and its analogs have good research and development prospects.
3.2.3 Inhibitors of glycosylation
Chloroquine is a widely used antimalarial and autoimmune disease drug, which is recently reported as a potential broad-spectrum antiviral drug. Chloroquine can block viral infection by up-regulating the pH of the endosome required for the fusion of the virus with the cell and inhibiting the glycosylation of the cell receptor, and specifically interacts with sugar-modifying enzymes or glycosyltransferases in human cells. Studies have shown that chloroquine may have an inhibitory effect on quinone reductase 2, which is structurally adjacent to UDP-N-acetylglucosamine 2-epimerase, thereby affecting the biosynthesis of sialic acid. The presence of sialic acid in the coronavirus receptor ACE2 may explain the inhibitory effect of chloroquine on SARS-CoV replication and other functions. Chloroquine is very effective in controlling 2019-nCoV infection in vitro, to play a role in both the entry phase and the post-entry phase of infection in Vero E6 cells. In addition to its antiviral activity, chloroquine also has immunomodulatory activity, which can synergistically enhance its antiviral effect in the body.
3.2.4 Inhibitors of endocytosis
Chlorpromazine is an antipsychotic drug used to treat schizophrenia. After attaching to the host surface receptors, most viruses use cellular endocytosis mechanisms (clathrin-dependent and independent pathways) to enter cells. SARS-CoV utilizes clathrin-dependent mechanisms to enter host cells. Chlorpromazine, an inhibitor of clathrin-dependent endocytosis, can significantly inhibit the replication of SARS-CoV. The study found that chlorpromazine is a broad-spectrum virus inhibitor that can inhibit HCV, alpha virus, and a variety of coronaviruses including human coronavirus 229E, SARS-CoV and MERS-CoV in vitro.
3.2.5 Kinase inhibitors
Imatinib (Gleevec) is a small molecule inhibitor targeting Abl2 kinase (Abelson tyrosine-protein kinase 2), a non-receptor tyrosine kinase present in the nucleus and mitochondria, mediated from the embryo morphogenesis to multiple cellular processes of viral infection. Abl2 kinase is required for SARS-CoV and MERS-CoV replication in vitro. Experiments showed that as an in vitro inhibitor of SARS-CoV and MERS-CoV, imatinib can significantly reduce the SARS-CoV and MERS-CoV virus titers, and inhibit the SARS-CoV and MERS-CoV pseudotype virions process. Imatinib's anti-coronavirus activity is mainly reflected in the early stages of infection, mainly by inhibiting the fusion of viral particles on the endosomal membrane.
3.3 Vaccines and antibodies
3.3.1 Monoclonal antibody
Monoclonal antibodies (mAbs) have been successfully applied to treat various diseases. Passive immunotherapy using neutralizing monoclonal antibodies (mAbs) is an effective preventive and therapeutic agent against emerging viruses. SARS-CoV and MERS-CoV have always been global public health threats, and so far, there are no approved vaccines or specific therapies.
Neutralizing antibodies are the main components of protective immunity against human viral infections. Antibody reactions in the body mobilize a dynamic mixture of mAbs. These antibodies work in concert to target various antigens on the viral envelope glycoprotein. Neutralizing the mAb can be achieved by a variety of techniques, such as hybridoma technology, humanized mice, phage or yeast, and single B cell isolation. More and more mAbs have been developed so far, which have shown higher potency against emerging viruses both in vitro and in animal models of infection. The main problem with neutralizing mAb therapy is the escape of mutants under selective pressure, which can be solved by combining neutralizing mAbs targeting different epitopes.
Spike glycoprotein plays a key role in mediating virus entry and has the ability to induce protective antibody responses in infected individuals. Consistent with this, the spike protein is the main target for neutralizing antibodies. At present, a number of mAbs targeting SARS-CoV and MERS-CoV are undergoing a phase I clinical trial, which has the prospect of developing new viral diseases. Based on the above research, neutralizing antibodies against the spike protein on the surface of SARS-CoV-2 may be the first therapy considered by the researchers.
3.3.2 Serum of recovered patients
Immunotherapy based on blood sources of recovered patients can be used to treat infections caused by measles virus, Lhasa virus, SARS coronavirus and influenza A H5N1 virus. This method is also applicable to the treatment of 2019-nCoV. Rehabilitation patients with COVID-19 will produce high-titer polyclonal antibody immune responses to different antigens of SARS-CoV-2. Some of these polyclonal antibodies may neutralize the virus and prevent a new round of infection.
Although a number of drugs with good inhibitory effects on 2019-nCoV have emerged in in vitro tests, the experimental environment in vitro is relatively simple, and these drugs may not achieve similar therapeutic effects in vitro as in vivo. In addition, safety and other issues need to be considered, and data still need to be waited for further tests for verification. At present, the most effective method for combating viruses is still a vaccine. Compared with the long period required for drug research and development, the vaccine requires a relatively short time and a strong protective effect.
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