Introduction

Coronaviruses, a name coined in 1968 that derives from the crown morphology of its structure, are a group of viruses that affect the respiratory tract. Coronavirus COVID-19 spreads predominantly through the respiratory tract with a high degree of infectivity. They belong to the subfamily Coronavirinae which together with Torovirinae form the family Coronaviridae in the order Nidovirales and are known to infect mammals and birds. Genotipically, coronaviruses can be divided into three groups. Group III viruses are found exclusively in birds, while Group I and II viruses have mammals as their host (1).

Coronaviruses belong to the subfamily Coronavirinae which together with Torovirinae form the family Coronaviridae in the order Nidovirales (2) created in 1996.

Infection and Mortality

Influenza epidemics lead to increased mortality due both to influenza itself and pneumonia, but also to complications and aggravation of other chronic illnesses due to influenza. In the absence of laboratory tests, it is not easy to determine whether the causes of deaths can be attributed to influenza as the primary cause.

Between 250,000 and 500,000 influenza-related deaths occur each year, according to WHO estimates (3).

The pandemic influenza pandemic of 1918/1919 so-called Spanish (H1N1 strain), developed with a three-wave model and caused the deaths of about 50 million people (4) with a worldwide mortality rate of 2-3%. Although criticized, one study (5) believed that there were phylogenetic similarities between Spanish and coronavirus. Most infected age groups: Young adults

The first coronavirus was discovered in 1930 (6).

The second coronavirus, in the years 1957-1958 ( HCoV-229E and HCoV-OC43) or so-called Asian influenza (strain H2N2), of presumably avian origin, developed in February in Guizhou province in southern China, spread in March in Hunan province, in April in Singapore and Hong Kong, in a two-wave model caused the death of about 1/4 million people worldwide with a mortality rate of 0.2%. Most infected age groups: all age groups (7).

The third coronavirus in 1968 or Hong Kong flu (H3N2 strain), presumably of avian origin, caused the deaths of about 1 million people with a mortality rate similar to 1957 of 0.2%. Most infected age groups: all age groups (6).

In November 2002 in Foshan, Guangdong Province, mainland China, SARS-CoV developed in Foshan, which had a new coronavirus, the fourth causative agent. The origin appears to come from two animals: civet (Paguma larvata) and raccoon (Nyctereutes procynonoides), sold in markets as culinary delicacies. SARS-CoV infected more than 8000 people and caused 774 deaths in 26 countries on five continents. Most infected age groups: all age groups (8).

In June 2009, a coronavirus, the fifth, resulting from a combination of genetic segments of avian, porcine and human influenza viruses originating in Mexico (H1N1pdm09) caused the deaths of about 200-400,000 people worldwide with a mortality rate of 0.02%. Most infected age groups: children and adults. The pandemic was declared officially over by the WHO in August 2010. This influenza showed an anomalous figure compared to similar seasonal epidemics: about 80% of deaths affected people under 65 years of age, while an estimated 80-90% of deaths caused by previous seasonal epidemics in the older population aged 65 and over (9).

In April 2020 COVID-19 (developed in Wuhan, China) infected nearly 3 million people and caused the death of more than 200,000 people with a mortality rate of 6.7%.

Coronavirus COVID-19 developed in China from two strains of different animal origin: bat and pangolin (10) although other studies believe COVID-19 is closely related to coronaviruses derived from wild animals, including Paguma larvata, Paradoxurus hermaphroditus, Civet, Aselliscus stoliczkanus and Rhinolophus sinicus, located in the same branch of the phylogenetic tree. However, the genome and ORF1a homology show that the virus is not the same coronavirus as that derived from these animals, while the virus has the highest homology with the bat coronavirus isolated RaTG13. On pangolin the genome affinity is not yet clear (11).

Some evidence, however, indicates that it could come from laboratories. In fact, an episode of Coronavirus SARS occurred in September 2003 in Singapore, China. The patient, with confirmed symptoms of SARS, was working in a laboratory and reported working on West Nile virus, but the laboratory was doing live SARS work at the time (12).

Droplet transmission is the main route (13) and travel was the main source of transmission of COVID-19 cases during the early stages of the current epidemic in Italy (14) and China. This study found that 779 cases would have been exported from China by 15 February 2020 due to the lack of border restrictions or travel restrictions and that the travel blocking measures applied by the Chinese government subsequently avoided 70.5 % of contagion (15). In particular, the coincidence of the Spring Festival in China increased travel volumes from Wuhan, Hubei Province, epicentre of COVID-19 spreading the epidemic (16).

The defenses

Since no vaccine or drug has been approved to date, the population remains defenceless from COVID-19 entering the human body mainly due to exposure to clouds of respiratory droplets expanding about 5 meters or more, caused by sneezing or coughing from infected people. These clouds can travel in the air and at least 20% can stay alive from 3 hours to 6 days (17).

Currently, the only defence available is to maintain the distance between individuals and respiratory protection devices that are used by those who have to defend themselves against radioactive, biological, chemical materials.

Half face respirators are easy to wear and lightweight. There are currently several types of these respirators on the market which are classified in Europe (EN 149:2001) as FFP1 (minimum filtration efficiency 80%), FFP2 (minimum filtration efficiency 94%) and FFP3 (minimum filtration efficiency 99%), while the US National Institute for Occupational Safety and Health (NIOSH) classifies them as N95 (minimum filtration efficiency 95%), N99 (minimum filtration efficiency 99%), N100 (minimum filtration efficiency 99.97%) (18). Surgical masks commonly used by doctors in the operating room are also available on the market.

Some studies have verified the effective protection of these respirators.

Influenza viruses and coronavirus are particles between 0.04 and 0.2 um.

This 2008 study concludes that the N95 respirators tested provided about 8-12 times better protection than surgical masks. However, approximately 29% of the N95 respirators tested had a PF of less than 10, indicating that the PF 10 value established by the US Occupational Safety and Health Administration (OSHA) overestimates the effective protection provided by N95 respirators against bacteria and viruses. The N95 filter mask respirators with valves showed almost the same protection against bacterial and viral particles as those without valves (19).

In a more recent study in 2016, laboratory tests showed that 10% of FFP2 respirators and 28.2% of FFP3 respirators had lower protection factors than those assigned by the European standard EN 149:2001 (20).

The search for therapy

No vaccine or drug has been approved to date 

Monoclonal antibodies represent the main class of biotherapy for passive immunotherapy to combat viral infections (21). However, they have contraindications that should be evaluated on a case-by-case basis. Tocilizumab (distributed free of charge by Roche while stocks last) can help the patient to breathe independently and leave the intensive care unit.

A temporary suggestion, probably resistant to new coronavirus mutations, is the use of angiotensin type 1 receptor blockers as a therapy to reduce aggressiveness and mortality from SARS-CoV-2 infections (22).

Nucleoside analogs such as favipiravir (T-705 or 6-fluoro-3-oxo-3,4-dihydropyrazine-2-carboxamide) approved for new strains that do not respond to current antivirals, marketed and approved in Japan under the name Avigan®, as well as ribavirin (Tribavirin or Virazole) and experimental nucleoside analogues such as remdesivir (a nucleotide-like formula currently in clinical trials for the treatment of Ebola virus infections) and galidesivir, may have potential on 2019-nCoV (23) but remdesivir appeared superior in efficacy to lopinavir/ritonavir and interferon beta against MERS-CoV in a model of humanized transgenic mice (24).  Regarding efficacy, favipiravir demonstrated an IC50 (concentration required to inhibit 50% of the target) of 601 μM versus 3.9 μM attributed to ribavirin (25), but its use is approved with limitations as it has a risk of teratogenicity and embotoxicity (26).

This study describes the first generation of MERS-CoV fusion inhibitors with potencies in the low micromolar range (27).

Chloroquine phosphate, an already effective drug for the treatment of malaria, has been shown to have apparent efficacy and acceptable safety against COVID-19 associated pneumonia in multicentre clinical trials conducted in China (28) in 500mg tablets twice daily for 10 days in patients diagnosed as mild cases, moderate and severe new coronavirus-related pneumonia without contraindications to chloroquine (29) and already proposed 20 years ago by the authors of this study as effective in vitro against a wide range of viruses (30), while another study considers hydroxychloroquine more potent and more tolerable (31).

Procalcitonin, a prohormone precursor of calcitonin, a diagnostic biomarker approved by the FDA in 2005, for patients with more severe symptoms of Coronavirus 2019 (COVID-19)(32) in intensive care.

In in vitro experiments have been found to be effective against pangolin-borne coronavirus, cepharanthin alkaloid, selamectin, and mefloquine hydrochloride, another known remedy against malaria (33).

During the severe COVID-19 epidemic, many hospitals administered Tocilizumab to intubated patients who were able to breathe independently thanks to this monoclonal antibody.

People in old age are more likely to contract acute respiratory distress syndrome (ARDS) and this study suggests treatment with methylprednisolone (34), synthetic glucocorticoid and potent anti-inflammatory drug.

A highly standardized blend of active compounds derived from the action of Lentinula Edodes Mycelia (AHCC) that can promote a protective response to a wide range of viral infections, and the current absence of effective vaccines could support its use in the prevention of diseases caused by human pathogenic coronaviruses, including COVID-19 (35).

It is important to understand the defence dynamics of coronavirus to try to inhibit its replication, a very complex process aimed at preserving its large RNA genome. Replication and maturation of the virus have essential strengths in several proteases including Mpro or 3CL and papain-like protease (PLpro) capable of generating developed proteins. 

In previous MERS-CoV and SARS-CoV disulfiram (Tetraethylthiuram disulfide, Antabuse™) had been shown to inhibit papain protease and act as an allosteric inhibitor for MERS-CoV and as a competitive (or mixed) SARS-CoV inhibitor (36). However, it should not be co-administered to lopinavir/ritonavir (37).

In fact, PLpro plays a fundamental role in the defense of the virus as it is primarily responsible for shaking off the proteins of host cells that try, with the immune response, to neutralize the virus (38) and also plays a pharmacokinetic interaction on the stimulation of the host interferon through its deubiquitination activity (39).

Disulfiram is a well known remedy against alcoholism, but it has also recently demonstrated its effectiveness against cancer, leukemia and, importantly, low toxicity (40). This study showed that the disulfiram/copper complex significantly induced the arrest of the cell cycle in the G2/M phase in MM.1S and RPMI8226 cells (41).

Between 2005 and 2009 some studies had identified 6-mercaptopurins (an antitumor drug approved by the US Food and Drug Administration (FDA)), 6-thioguanins (a chemotherapeutic drug) and mycophenolic acid as compounds able to inhibit PLpro of MERS-CoV through their synergistic effects and able to constitute a basis for antiviral drugs (42) (43).

In 2018, during the PEDV pig epidemic, 6-thioguanine was suggested as a non-competitive PLpro inhibitor (44).

Another anti-malarial anti-infective drug, mefloquine, structurally related to quinine, which is often combined with artesunate (a semi-synthetic derivative of artemisininin) could provide results in the fight against coronavirus, but has contraindications both for the long treatment period and for potential side effects (45).

References_________________________________________________________________________

(1) van der Hoek L, Pyrc K, Berkhout B. Human coronavirus NL63, a new respiratory virus. FEMS Microbiol Rev. 2006;30(5):760–773. doi:10.1111/j.1574-6976.2006.00032.x

(2)  Belouzard S, Millet JK, Licitra BN, Whittaker GR. Mechanisms of coronavirus cell entry mediated by the viral spike protein. Viruses. 2012;4(6):1011–1033. doi:10.3390/v4061011

(3) Iuliano AD, Roguski KM, Chang HH, et al. Estimates of global seasonal influenza-associated respiratory mortality: a modelling study [published correction appears in Lancet. 2018 Jan 19;:]. Lancet. 2018;391(10127):1285–1300. doi:10.1016/S0140-6736(17)33293-2

(4) Johnson NP, Mueller J.  Updating the accounts: global mortality of the 1918-1920 "Spanish" influenza pandemic. Bull Hist Med. 2002 Spring;76(1):105-15.

(5) Gibbs, M. J., Armstrong, J. S. & Gibbs, A. J. 2001 Recombination in the hemagglutinin gene of the 1918 ‘Spanish flu’.

Science 293, 1842–1845.

(6)https://www.cdc.gov/flu/pandemic-resources/1968-pandemic.html

(7) https://www.who.int/influenza/preparedness/pandemic/PIRM_withCoverPage_201710_FINAL.pdf?ua=1

(8) Joseph S.M. Peiris, M.D., D.Phil., Kwok Y. Yuen, M.D., Albert D.M.E. Osterhaus, Ph.D., and Klaus Stöhr, Ph.D.  The Severe Acute Respiratory Syndrome  December 18, 2003 N Engl J Med 2003; 349:2431-2441 DOI: 10.1056/NEJMra032498

(9) Dawood FS, Iuliano AD, Reed C, et al. Estimated global mortality associated with the first 12 months of 2009 pandemic influenza A H1N1 virus circulation: a modelling study [published correction appears in Lancet Infect Dis. 2012 Sep;12(9):655]. Lancet Infect Dis. 2012;12(9):687–695. doi:10.1016/S1473-3099(12)70121-4

(10) Fan HH, Wang LQ, Liu WL, An XP, Liu ZD, He XQ, Song LH, Tong YG. Repurposing of clinically approved drugs for treatment of coronavirus disease 2019 in a 2019-novel coronavirus (2019-nCoV) related coronavirus model.   Chin Med J (Engl). 2020 Mar 6. doi: 10.1097/CM9.0000000000000797.

(11) Li C, Yang Y, Ren L. Genetic evolution analysis of 2019 novel coronavirus and coronavirus from other species [published online ahead of print, 2020 Mar 10]. Infect Genet Evol. 2020;104285. doi:10.1016/j.meegid.2020.104285

(12) https://www.nas.gov.sg/archivesonline/data/pdfdoc/20030923-MOH.pdf

(13)  Han Q, Lin Q, Ni Z, You L.Uncertainties about the transmission routes of 2019 novel coronavirus.   Influenza Other Respir Viruses. 2020 Mar 4. doi: 10.1111/irv.12735.

(14) Porcheddu R, Serra C, Kelvin D, Kelvin N, Rubino S. Similarity in Case Fatality Rates (CFR) of COVID-19/SARS-COV-2 in Italy and China.    J Infect Dev Ctries. 2020 Feb 29;14(2):125-128. doi: 10.3855/jidc.12600.

(15) Wells CR, Sah P, Moghadas SM, et al. Impact of international travel and border control measures on the global spread of the novel 2019 coronavirus outbreak [published online ahead of print, 2020 Mar 13]. Proc Natl Acad Sci U S A. 2020;202002616. doi:10.1073/pnas.2002616117

(16) Zhong P, Guo S, Chen T. Correlation between travellers departing from Wuhan before the Spring Festival and subsequent spread of COVID-19 to all provinces in China [published online ahead of print, 2020 Mar 17]. J Travel Med. 2020;taaa036. doi:10.1093/jtm/taaa036

(17) Ijaz MK, Brunner AH, Sattar SA, Nair RC, Johnson-Lussenburg CM. Survival characteristics of airborne human coronavirus 229E. J Gen Virol. 1985;66 ( Pt 12):2743–2748. doi:10.1099/0022-1317-66-12-2743

(18) National Institute for Occupational Safety and Hygiene (NIOSH) NIOSH Guide to the Selection and Use of Particulate Respirators Certified Under 42 CFR 84. Cincinnati, Ohio, USA: National Institute for Occupational Safety and Hygiene (NIOSH); 1996

(19) Lee SA, Grinshpun SA, Reponen T. Respiratory performance offered by N95 respirators and surgical masks: human subject evaluation with NaCl aerosol representing bacterial and viral particle size range. Ann Occup Hyg. 2008;52(3):177–185. doi:10.1093/annhyg/men005

(20) Lee SA, Hwang DC, Li HY, Tsai CF, Chen CW, Chen JK. Particle Size-Selective Assessment of Protection of European Standard FFP Respirators and Surgical Masks against Particles-Tested with Human Subjects. J Healthc Eng. 2016;2016:8572493. doi:10.1155/2016/8572493

(21) Shanmugaraj B, Siriwattananon K, Wangkanont K, Phoolcharoen W.Perspectives on monoclonal antibody therapy as potential therapeutic intervention for Coronavirus disease-19 (COVID-19).    Asian Pac J Allergy Immunol. 2020 Mar 4. doi: 10.12932/AP-200220-0773.

(22) Gurwitz D. Angiotensin receptor blockers as tentative SARS-CoV-2 therapeutics.  Drug Dev Res. 2020 Mar 4. doi: 10.1002/ddr.21656.

(23) Li G, De Clercq E. Therapeutic options for the 2019 novel coronavirus (2019-nCoV).   Nat Rev Drug Discov. 2020 Mar;19(3):149-150. doi: 10.1038/d41573-020-00016-0.               

Dong L, Hu S, Gao J.  Discovering drugs to treat coronavirus disease 2019 (COVID-19).   Drug Discov Ther. 2020;14(1):58-60. doi: 10.5582/ddt.2020.01012.

(24) Martinez MA. Compounds with therapeutic potential against novel respiratory 2019 coronavirus.    Antimicrob Agents Chemother. 2020 Mar 9. pii: AAC.00399-20. doi: 10.1128/AAC.00399-20.

(25) Furuta Y, Gowen BB, Takahashi K, Shiraki K, Smee DF, Barnard DL.  Favipiravir (T-705), a novel viral RNA polymerase inhibitor.  Antiviral Res. 2013 Nov;100(2):446-54. doi: 10.1016/j.antiviral.2013.09.015.

(26) Nagata T, Lefor AK, Hasegawa M, Ishii M. Favipiravir: a new medication for the Ebola virus disease pandemic. Disaster Med Public Health Prep. 2015;9(1):79–81. doi:10.1017/dmp.2014.151 

(27) Kandeel M, Yamamoto M, Al-Taher A, Watanabe A, Oh-Hashi K, Park BK, Kwon HJ, Inoue JI, Al-Nazawi M.  Small Molecule Inhibitors of Middle East Respiratory Syndrome Coronavirus Fusion by Targeting Cavities on Heptad Repeat Trimers.   Biomol Ther (Seoul). 2020 Mar 4. doi: 10.4062/biomolther.2019.202.

(28) Gao J, Tian Z, Yang X.  Breakthrough: Chloroquine phosphate has shown apparent efficacy in treatment of COVID-19 associated pneumonia in clinical studies.   Biosci Trends. 2020 Feb 19. doi: 10.5582/bst.2020.01047.

(29) Zhonghua Jie He He Hu Xi Za Zhi. multicenter collaboration group of Department of Science and Technology of Guangdong Province and Health Commission of Guangdong Province for chloroquine in the treatment of novel coronavirus pneumonia. Expert Consensus on Chloroquine Phosphate for the Treatment of Novel Coronavirus Pneumonia  2020;43(3):185–188. doi:10.3760/cma.j.issn.1001-0939.2020.03.009

(30) Colson P, Rolain JM, Raoult D. Chloroquine for the 2019 novel coronavirus SARS-CoV-2.   Int J Antimicrob Agents. 2020 Feb 15:105923. doi: 10.1016/j.ijantimicag.2020.105923.

(31)  Yao X, Ye F, Zhang M, Cui C, Huang B, Niu P, Liu X, Zhao L, Dong E, Song C, Zhan S, Lu R, Li H, Tan W, Liu D.  In Vitro Antiviral Activity and Projection of Optimized Dosing Design of Hydroxychloroquine for the Treatment of Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2). Clin Infect Dis. 2020 Mar 9. pii: ciaa237. doi: 10.1093/cid/ciaa237.

(32) Lippi G, Plebani M.   Procalcitonin in patients with severe coronavirus disease 2019 (COVID-19): a meta-analysis.   Clin Chim Acta. 2020 Mar 4. pii: S0009-8981(20)30106-6. doi: 10.1016/j.cca.2020.03.004.

Chanu Rhee, Using Procalcitonin to Guide Antibiotic Therapy, Open Forum Infectious Diseases, Volume 4, Issue 1, Winter 2017, ofw249, https://doi.org/10.1093/ofid/ofw249

(33) Fan HH, Wang LQ, Liu WL, An XP, Liu ZD, He XQ, Song LH, Tong YG. Repurposing of clinically approved drugs for treatment of coronavirus disease 2019 in a 2019-novel coronavirus (2019-nCoV) related coronavirus model.   Chin Med J (Engl). 2020 Mar 6. doi: 10.1097/CM9.0000000000000797.

(34) Wu C, Chen X, Cai Y, et al. Risk Factors Associated With Acute Respiratory Distress Syndrome and Death in Patients With Coronavirus Disease 2019 Pneumonia in Wuhan, China [published online ahead of print, 2020 Mar 13]. JAMA Intern Med. 2020;10.1001/jamainternmed.2020.0994. doi:10.1001/jamainternmed.2020.0994

(35) Di Pierro F, Bertuccioli A, Cavecchia I. Possible therapeutic role of a highly standardized mixture of active compounds derived from cultured Lentinula edodes mycelia (AHCC) in patients infected with 2019 novel coronavirus [published online ahead of print, 2020 Mar 12]. Minerva Gastroenterol Dietol. 2020;10.23736/S1121-421X.20.02697-5. doi:10.23736/S1121-421X.20.02697-5

(36) Lin MH, Moses DC, Hsieh CH, et al. Disulfiram can inhibit MERS and SARS coronavirus papain-like proteases via different modes. Antiviral Res. 2018;150:155–163. doi:10.1016/j.antiviral.2017.12.015

(37) Cvetkovic RS, Goa KL. Lopinavir/ritonavir: a review of its use in the management of HIV infection. Drugs. 2003;63(8):769–802. doi:10.2165/00003495-200363080-00004

(38) Brian DA, Baric RS. Coronavirus genome structure and replication. Curr Top Microbiol Immunol. 2005;287:1-30. 

(39) Báez-Santos YM, St John SE, Mesecar AD. The SARS-coronavirus papain-like protease: structure, function and inhibition by designed antiviral compounds. Antiviral Res. 2015 Mar;115:21-38. doi: 10.1016/j.antiviral.2014.12.015. 

(40) Sheppard JG, Frazier KR, Saralkar P, Hossain MF, Geldenhuys WJ, Long TE. Disulfiram-based disulfides as narrow-spectrum antibacterial agents. Bioorg Med Chem Lett. 2018;28(8):1298–1302. doi:10.1016/j.bmcl.2018.03.023

(41) Xu Y, Zhou Q, Feng X, et al. Disulfiram/copper markedly induced myeloma cell apoptosis through activation of JNK and intrinsic and extrinsic apoptosis pathways [published online ahead of print, 2020 Mar 4]. Biomed Pharmacother. 2020;126:110048. doi:10.1016/j.biopha.2020.110048

(42) Cheng KW, Cheng SC, Chen WY, et al. Thiopurine analogs and mycophenolic acid synergistically inhibit the papain-like protease of Middle East respiratory syndrome coronavirus. Antiviral Res. 2015;115:9–16. doi:10.1016/j.antiviral.2014.12.011

(43) Chou CY, Chien CH, Han YS, et al. Thiopurine analogues inhibit papain-like protease of severe acute respiratory syndrome coronavirus. Biochem Pharmacol 2008;75:1601–1609.

(44) Chu HF, Chen CC, Moses DC, et al. Porcine epidemic diarrhea virus papain-like protease 2 can be noncompetitively inhibited by 6-thioguanine. Antiviral Res. 2018;158:199–205. doi:10.1016/j.antiviral.2018.08.011 

(45) Nevin RL, Byrd AM. Neuropsychiatric Adverse Reactions to Mefloquine: a Systematic Comparison of Prescribing and Patient Safety Guidance in the US, UK, Ireland, Australia, New Zealand, and Canada. Neurol Ther. 2016;5(1):69–83. doi:10.1007/s40120-016-0045-5