Recent studies have found that fapiravir also has an inhibitory effect on the novel coronavirus (EC50 = 61.88 uM), and does not have such high requirements for the tissue distribution of the drug. Therefore, fapiravir is also a potential anti-CoV drug. On February 15, the State Food and Drug Administration approved the use of fapiravir (also known as favaravir) produced by Haizheng Pharmaceutical to treat new or re-exchangeable influenza in adults (only when other anti- influenza virus drugs are ineffective or poorly effective). At the same time, the clinical trial of the new coronavirus was approved on the same day and has been launched at the First Affiliated Hospital of Zhejiang University School of Medicine.
Written by | Enron Medicine
The current outbreak of the novel coronavirus (COVID-19) in China has become a major black swan event this year. Scientists from all over the world are working hard to solve more problems such as the structure, nature and relationship between the new coronavirus. The most important topic at present is to find special drugs against the novel coronavirus. targets the new coronavirus. The current mainstream drug research and development strategies include new use of old drugs and structure-based redesign.
Old medicines new use
Novel coronavirus belongs to the family Coronavirus family, and the coronaviruses we are familiar with also include SARS-CoV and MERS-CoV. Their genomes are all sense single-stranded RNA, with a total length of 26-32 Kb, decoding multiple proteins, including structural proteins (envelope glycoprotein spike), envelope proteins (E), membrane proteins (M), nucleocapsid proteins (N), etc.), non-structural proteins (helicase, protease (3CLpro, PLpro), RNA-dependent RNA polymerase and other proteins) and auxiliary proteins (Orf 3a, etc.).
Previous studies on SARS-CoV have found that auxiliary proteins are not necessary during the replication of viruses. Their function may mainly lie in interacting with proteins in host cells, thereby affecting the virus's infection process. The Spike protein in structural proteins acts with the receptor on the host cell (COVID-19 and SARS - CoV: ACE2; MERS - CoV: DPP4) to induce endocytosis of the virus into the cells, while the helicase, protease and RNA-dependent RNA polymerase in non-structural proteins play an important role in the replication process of viruses. Therefore, in order to develop direct antiviral drugs, previous drug development strategies focused on non-structural proteins (especially helicase, 3CLpro, PLpro and RNA-dependent RNA polymerase) and structural proteins (mainly Spike proteins).
Recent studies have found that the overall viral RNA sequence of SARS-CoV and COVID-19 are nearly 80% identical (this is why the World Health Organization named the novel coronavirus SARS-CoV-2), 96% identical on the amino acid sequence of the RNA-dependent RNA polymerase and the protease 3CLpro, 83% identical on the amino acid sequence of the protease PLpro, and 76% identical on the amino acid sequence of the Spike protein.
Therefore, one of the main strategies for drug discovery against novel coronavirus is to use previously discovered drugs that are effective against SARS-CoV against novel coronaviruses. Although 17 years have passed since the SARS epidemic that broke out in 2003, due to factors such as the rapid outbreak and short duration, no drugs that have been clinically proven to be effective for SARS-CoV. However, many antiviral drugs have been shown to inhibit the replication of SARS-CoV in vitro.
Table 1. Potential anti-COVID-19 old drugs (unit: uM)
Drugs targeting RNA-dependent RNA polymerases include remdesivir, ribavirin and fapiravir.
ribavirin
ribavirin is mainly used in combination with interferon for the treatment of chronic hepatitis C (HCV) and respiratory fusion virus (RSV) infection. In vitro antiviral activity experiments demonstrated that ribavirin inhibits type 1b HCV, SARS-CoV and COVID-19 at the same level (86 for EC50; 82-328 and 109.5 uM, respectively). Therefore, the effectiveness of ribavirin combined with interferon on COVID-19 can be expected. The National Health Commission also recommended the use of ribavirin interferon for antiviral treatment in the "Diagnosis and Treatment Plan for Novel Coronavirus Pneumonia (Trial Fifth Edition, Revised Edition)" released on February 8, 2020.Fapiravir is mainly used to treat influenza, and in vitro studies have shown that it has certain inhibitory activity against Ebola virus (EBOV) and Zika virus (Zika virus) (EC50 = 10.5 and 3.5 - 3.8 uM). EBOV, Zika and influenza viruses are also single-stranded RNA viruses, and the participation of RNA-dependent RNA polymerase is required during the virus replication process. Fapiravir
. A clinical trial (JIKI trial) conducted in Guinea that assessed the effectiveness of favipiravir (1200 mg, bid) against EBOV was found to be very safe, but did not show obvious effectiveness. The failure of this clinical trial may be because the blood concentration at this dosage does not reach EC50 in vitro anti-EBOV. Recent studies have found that fapiravir also has an inhibitory effect on the novel coronavirus (EC50 = 61.88 uM), which is on the same order of magnitude as the anti-EBOV activity. In addition, EBOV infects almost all organs and tissues. The novel coronavirus is currently found to mainly cause lung infections, so there is no high requirement for the tissue distribution of drugs. Therefore, fapiravir is also a potential anti-CoV drug. On February 16, the State Food and Drug Administration approved the use of fapiravir (also known as favaravir) produced by Haizheng Pharmaceutical to treat new or re-expat influenza in adults (only when other anti-influenza virus drugs are ineffective or have poor results).
Remdesivir
At present, the most anticipated drug is Gilead's remdesivir. Although its inhibitory activity against the new coronavirus in vitro is nearly 10 times different from SARS-CoV and MERS-CoV, it is still the most publicly disclosed drug with the strongest in vitro anti-COVID-19 activity (EC50 = 0.77 uM), and it has shown clinical effectiveness in some patients. Several phase 3 clinical trials are currently being carried out. We look forward to its good results.
lopinavir and ritonavir
drugs that target HIV aspartate proteases have played a certain role in the SARS epidemic. However, the proteases of the coronavirus belong to the cysteine protease family, so the activity of lopinavir on SARS-CoV and MERS-CoV is about 1,000 times lower than that of HIV-1, but it still shows a certain inhibitory activity (EC50 is 17.1 and 8.0 uM, respectively). Therefore, in the "Diagnosis and Treatment Plan for Novel Coronavirus Pneumonia (Trial Fifth Edition, Revised Edition)", it is also recommended to use rilopinavir and ritonavir (according to the recommended dose for treating HIV) to treat anti-novel coronavirus.
chloroquine
lysosomal reagent chloroquine is an antimalarial drug. It was later found that it has a broad-spectrum antiviral activity and showed certain inhibitory activities against viruses such as HIV-1, HBV and human coronavirus (HCoV-229E). Chloroquine will aggregate and capture protons in the lysosome, thereby increasing the pH of the lysosome, thereby blocking the lysosome's cleavage of virus envelope and reducing the virus's ability to release nucleic acids. In addition, it can interact with many proteins in the host cell to regulate the immune response of the cell. Studies have shown that chloroquine has inhibitory activities on SARS-CoV and COVID-19 (8.8 and 1.33 uM EC50, respectively).
At the press conference of the State Council’s Joint Prevention and Control Mechanism on February 15, 2020, Zhang Xinmin, director of the Biological Center of the Ministry of Science and Technology, said that chloroquine phosphate is currently conducting clinical research in more than ten hospitals including Beijing and Guangzhou, and has accumulated more than 100 patients. The clinical results have initially shown that it has certain therapeutic effects on COVID-19. Nitazonid (or its active metabolite tezonit), which targets the human immune system, is also a broad-spectrum antiviral drug. Studies have shown that tezonit can activate PKR and eIF2-a to inhibit the replication of HCV virus, or inhibit the replication of EBOV by enhancing the expression of proteins such as RIG-like receptors, mitochondrial antiviral signaling proteins and interferon regulator 3. In vitro studies have shown that nitazonid (or tezonidide) has moderate inhibitory activity against HBV and COVID-19 (0.2 and 2.12 uM EC50, respectively). However, the effectiveness of nitazonid in anti-HBV, HCV and HIV has not been clinically proven, so the effectiveness of nitazonid in the treatment of COVID-19 also requires clinical verification.
Abidor
Abidor is a Russian anti-influenza virus drug, which mainly binds to the hemagglutinin of the influenza virus, thereby blocking the influenza virus from entering the host cell. At present, the drug has not entered the US and EU markets. In vitro studies have shown that it has certain inhibitory activities on SARS-CoV and EBOV (EC50 is 10 and 2.83 uM, respectively). At present, Abidor Hydrochloride Hydrochloride is conducting several clinical trials against COVID-19 (NCT04260594, NCT04252885 and NCT04261907).
Structure-based drug design
The full sequence gene structure of the novel coronavirus was parsed out, and many strains of the novel coronavirus were isolated. Therefore, it is also a strategy to design drugs based on the structure of the virus. As mentioned earlier, targeting coronaviruses mainly focuses on Spike protein, protease (3CLpro, PLpro) RNA-dependent RNA polymerase and helicase. For small molecule drugs, structural molecular design can be based on protein sequence homology modeling, and a structural model of proteins such as 3CLpro, PLpro and RNA-dependent RNA polymerase of COVID-19 with high homology to SARS-CoV. Then, through virtual screening or further redesign and scoring of previous drugs targeting the above target proteins (such as remdesivir, GPL-001, GRL-0617, etc.) to obtain potentially better compounds.
It should be mentioned here that homologous modeling is not the real protein structure after all, so this type of drug design requires continuous design-synthesis-evaluation-redesign process. Even if the crystal data of the target protein in the future can make the redesign more rational, such a drug discovery process is quite time-consuming, let alone subsequent preclinical evaluation and clinical trials. Therefore, for this epidemic with rapid outbreak and short duration, this solution is not very realistic in solving the current problems. However, if it is to allow us to better face the next similar coronavirus outbreak that may come, such a work is particularly important.
There is another type of therapeutic drug, the development of therapeutic antibodies. Recently, antibodies have been detected in patients with rehabilitation of COVID-19, and patients in Wuhan, Xuzhou and other places have already received such antibodies and showed good clinical effectiveness. However, obtaining such antibodies requires the donation of plasma by recovering people, and its source and effectiveness of the antibodies cannot be guaranteed. Therefore, the best choice is to re-screen based on the protein structure of the virus (especially Spike protein) and artificially purify the preparation of therapeutic antibodies.
regenerator is undoubtedly the leader in this field. This company uses glycoprotein on the surface of the synthetic virus envelope to immunize genes (Veloc Immune mice) containing the variable region genes of the whole human beings, and then screens out antibodies that bind glycoprotein with high activity, strong neutralization ability and high FcγRIIIa receptor ability, and then enrich the corresponding antibodies through CHO cells. Using this screening synthesis system, Regeneron developed an antibody cocktail against EBOV glycoprotein within 10 months: REGN-EB3 (REGN3470, RENG3471 and REGN3479), and successfully developed a therapeutic antibody composition against MERS coronavirus Spike protein (REGN3048 and REGN3051). On February 4, 2020, the U.S. Department of Health and Human Services announced a partnership with Regeneron to develop therapeutic antibodies against COVID-19. Here, we look forward to good news from the regenerative.
The coronavirus is seriously threatening human health, but due to the characteristics of the outbreak and other reasons, people have not developed effective therapeutic drugs for SARS-CoV and MERS-CoV so far. Therefore, in the face of the current new coronavirus, our best strategy is to use existing old drugs for antiviral treatment. However, we don't know when similar coronaviruses will reappear, and we missed the learning opportunities SARS brings, and hopefully this time won't be missed again.
Author Profile: Enron Medicine is a medical person who has just entered the workplace. During his doctorate, he developed the habit of reading literature. He does chemistry in his hands, thinks about signal channels in his mind, and speaks about the footprints of his predecessors. I hope to make good medicines that the people can afford in my lifetime.
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