China’s Bitter Lesson From Pirating Patented Properties

A rift between United States and China this week can only be understood by the two of them. United States is accusing China of not being transparent. China is also saying it is the United States who should be transparent and admitting for the first time that the virus is not theirs, but the U.S’s. How did it find its way to China?

A patent document below shows that the virus was first patented by United States in 2006 and again in 2018. The patent is still pending till 2035. It belongs to the U.S military. How did it get to Wuhan laboratories?

Normally, if it is established that someone has stolen a patented intellectual property of another, the victim can charge the thief for monetary damages. But if it were you, would you come forward to claim damages, considering the tremendous damages the stolen property is causing around the globe, leaving huge humanitarian and economic devastations along its trails?

The Modified Coronavirus, Covid-19, as its affectionately called, is a deadly biological arsenal that has been developed by the U.S military. It was manufactured to destroy the enemies's front, and not for the civilian population. It is a weapon of mass destruction that cannot be easily controlled. Until today, March 13, it has killed 5,123 civilians, 140,214 are infected, of which 70,733 have recovered. Economies of the world are melting down, several global and local companies have lost their incomes and millions of workers and their families are on the threat to lose their daily bread.

China’s economy and its citizens have suffered the most, the economy of the communist nation has been reduced to its humiliating lowest. Everything has to start again. And it’s going to take several years to come back.

It is not every item that should be stolen. There are items, if you steal, you will end up killing your household. This is exactly what has happened to China. If they come out of the problem, I am not sure they would steal intellectual properties again.

This will also serve as a warning to the nations, companies and individuals who are pirating others’ patent properties, especially dangerous biological and chemical substances without considering the consequences of spillage.

China took a great offence at comments made by U.S. officials accusing it of being slow to react to the virus, first detected in Wuhan late last year, and of not being transparent enough. On Wednesday, March 11, the U.S. National Security Adviser Robert O'Brien said the speed of China's reaction to the emergence of the coronavirus had probably cost the world two months when it could have been preparing for the outbreak.

In a strongly worded tweet, written in English on his verified Twitter account, Chinese Foreign Ministry spokesman Zhao Lijian said it was the United States that lacked transparency. “When did patient zero begin in US? How many people are infected? What are the names of the hospitals? It might be US army who brought the epidemic to Wuhan. Be transparent! Make public your data! US owe us an explanation!" Zhao wrote.

Who is saying the truth in these tweets of events?
Copyright infringes are common occurrences round the globe. The most frequent copyright violations are related to computer and digital technology, mainly software piracy. Software piracy includes file sharing, illegal uploading, burning of CDs and DVDs, etc,. But pirating of biological substances are not common, though they occur.

Countries and nations invest in strong data protection solutions, which involve large expenses. However, the workers, who have access to the national or company data become the source of data thefts and pilferages.

China is known globally as one of the top pirates of intellectual properties. Intellectual property theft or piracy involves robbing people or companies and nations of their ideas, inventions, and creative expressions—known as “intellectual property”—which can include everything from trade secrets and proprietary products and parts to movies, music, and software.

Intellectual property (IP) theft occurs when someone uses one’s intellectual property for any reason without the owner’s permission. Laws protect intellectual property rights, including trademarks, copyrights, and patents. If one has the proper protections, the one can sue for money damages.

To prove that a work, being reproduced, displayed, or distributed by someone was derived from one’s exclusively copyrighted work, and no consent had been granted by the intellectual property owner for its use, the date and patent number must be provided. If a patented product or idea is found to be stolen, it amounts to fraud.

If one feels that he has been a victim of an intellectual property crime, there are national coordinating centers one can make a report. In the United States, the National Intellectual Property Registry or Coordination Center (IPR Center) is responsible for the offense. The victim can fill an online form or call 1-866-IPR-2060 , or contact a field office of the FBI.

Copyright infringement is generally a civil matter, which the copyright owner must pursue in federal court. Under certain circumstances, the infringement may also constitute a criminal misdemeanor or felony, which in United States, could be prosecuted by the U.S. Department of Justice.

The Country Which Owns The Patent For Coronavirus
The Coronavirus was first patented by United States of America. The patent number was US2006257852

Patent Submission Date was 2006-11-16
Patent Title: Severe acute respiratory syndrome coronavirus

Structure:
Status: Live
The Virus was modified eleven years later on 2017-01-24.

Available since 2011-12-03
Deposit on 2011-12-03
Patent Abstract
An outbreak of a virulent respiratory virus, now known as Severe Acute Respiratory Syndrome (SARS), was identified in Hong Kong, China and a growing number of countries around the world in 2003. The invention relates to nucleic acids and proteins from the SARS coronavirus. These nucleic acids and proteins can be used in the preparation and manufacture of vaccine formulations, diagnostic reagents, kits, etc. The invention also provides methods for treating SARS by administering small molecule antiviral compounds, as well as methods of identifying potent small molecules for the treatment of SARS.

Names of Inventors (country of citizenry)

1. CHIEN DAVID [US]
2. DONNELLY JOHN [US]
3. GREGERSEN JENS P [DE]
4. HAN JANG [US]
5. HOUGHTON MICHAEL [US]
6. KLENK HANS D [DE]
7. MASIGNANI VEGA [IT]
8. POLO JOHN M [US]
9. RAPPUOLI RINO [IT]
10. SEO MI-YOUNG [KR]
11. SONG HYUN C [US]
12. STADLER KONRAD [AU]
13. VALIANTE NICHOLAS [US]
14. WEINER AMY [US]

The Chemistry of the Virus
Compound CID: 6

• Name: 1-Chloro-2,4-dinitrobenzene
• Molecular Formula: C6H3ClN2O4
• Molecular Weight, g/mol: 202.55

Compound CID: 107

• Name: 3-Phenylpropionic acid
• Molecular Formula: C9H10O2
• Molecular Weight, g/mol: 150.17

Compound CID: 135

• Name: 4-Hydroxybenzoic acid
• Molecular Formula: C7H6O3
• Molecular Weight, g/mol: 138.12

Compound CID: 196

• Name: Adipic acid
• Molecular Formula: C6H10O4
• Molecular Weight, g/mol: 146.14

Compound CID: 227

• Name: Anthranilic acid
• Molecular Formula: C7H7NO2
• Molecular Weight, g/mol: 137.14
• Molecular Structures
• Substance SID: 127354536

Compound CID: 11984

• Substance Synonyms:
• Data Source: IBM
• Substance SID: 127362351

Compound CID: 0

• Substance Synonyms:
• Data Source: IBM
• Substance SID: 127365630

Compound CID: 0

• Substance Synonyms:
• Data Source: IBM
• Substance SID: 127373080

Compound CID: 11979

• Substance Synonyms:
• Data Source: IBM
• Substance SID: 127384203

Compound CID: 65220

• Substance Synonyms:
• Data Source: IBM

The Modified Virus was registered a year later. The patent of the modified virus wax expected to expire in 2035.

• Patent. Nr.: US10130701B2 United States
• Inventor: Erica Bickerton, Sarah Keep, Paul Britton
• Current Assignee: PIRBRIGHT INSTITUTE
• Application granted: 2018-11-20
• Anticipated expiration: 2035-07-23

Abstract
The present invention provides a live, attenuated coronavirus comprising a variant replicase gene encoding polyproteins comprising a mutation in one or more of non-structural protein(s) (nsp)-10, nsp-14, nsp-15 or nsp-16. The coronavirus may be used as a vaccine for treating and/or preventing a disease, such as infectious bronchitis, in a subject.

C07K14/005 Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from viruses.

US10130701B2
United States
Classifications and Vaccines

• C07K14/005 Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from viruses
• A61K39/215 Coronaviridae, e.g. avian infectious bronchitis virus
• C12N7/00 Viruses; Bacteriophages; Compositions thereof; Preparation or purification thereof
• C12N9/127 RNA-directed RNA polymerase (2.7.7.48), i.e. RNA replicase
• C12Y207/07048 RNA-directed RNA polymerase (2.7.7.48), i.e. RNA replicase
• A61K2039/5254 Virus avirulent or attenuated
• A61K2039/54 Medicinal preparations containing antigens or antibodies characterised by the route of administration
• A61K2039/70 Multivalent vaccine
• C12N2770/20021 Viruses as such, e.g. new isolates, mutants or their genomic sequences
• C12N2770/20022 New viral proteins or individual genes, new structural or functional aspects of known viral proteins or genes
• C12N2770/20034 Use of virus or viral component as vaccine, e.g. live-attenuated or inactivated virus, VLP, viral protein
• C12N2770/20051 Methods of production or purification of viral material
• C12N2770/20062 Methods of inactivation or attenuation by genetic engineering
• C12N2770/20071 Demonstrated in vivo effect

Hide more classifications
Description and Some Advices of Vaccines
FIELD OF THE INVENTION
The present invention relates to an attenuated coronavirus comprising a variant replicase gene, which causes the virus to have reduced pathogenicity. The present invention also relates to the use of such a coronavirus in a vaccine to prevent and/or treat a disease.

BACKGROUND TO THE INVENTION
Avian infectious bronchitis virus (IBV), the aetiological agent of infectious bronchitis (IB), is a highly infectious and contagious pathogen of domestic fowl that replicates primarily in the respiratory tract but also in epithelial cells of the gut, kidney and oviduct. IBV is a member of the Order Nidovirales, Family Coronaviridae, Subfamily Corona virinae and Genus Gammacoronavirus; genetically very similar coronaviruses cause disease in turkeys, guinea fowl and pheasants.

Clinical signs of IB include sneezing, tracheal rales, nasal discharge and wheezing. Meat-type birds have reduced weight gain, whilst egg-laying birds lay fewer eggs and produce poor quality eggs. The respiratory infection predisposes chickens to secondary bacterial infections which can be fatal in chicks. The virus can also cause permanent damage to the oviduct, especially in chicks, leading to reduced egg production and quality; and kidney, sometimes leading to kidney disease which can be fatal.

IBV has been reported to be responsible for more economic loss to the poultry industry than any other infectious disease. Although live attenuated vaccines and inactivated vaccines are universally used in the control of IBV, the protection gained by use of vaccination can be lost either due to vaccine breakdown or the introduction of a new IBV serotype that is not related to the vaccine used, posing a risk to the poultry industry.

Further, there is a need in the industry to develop vaccines which are suitable for use in ovo, in order to improve the efficiency and cost-effectiveness of vaccination programmes. A major challenge associated with in ovo vaccination is that the virus must be capable of replicating in the presence of maternally-derived antibodies against the virus, without being pathogenic to the embryo.

Current IBV vaccines are derived following multiple passage in embryonated eggs, this results in viruses with reduced pathogenicity for chickens, so that they can be used as live attenuated vaccines. However such viruses almost always show an increased virulence to embryos and therefore cannot be used for in ova vaccination as they cause reduced hatchability. A 70% reduction in hatchability is seen in some cases.

Attenuation following multiple passage in embryonated eggs also suffers from other disadvantages.

It is an empirical method, as attenuation of the viruses is random and will differ every time the virus is passaged, so passage of the same virus through a different series of eggs for attenuation purposes will lead to a different set of mutations leading to attenuation. There are also efficacy problems associated with the process: some mutations will affect the replication of the virus and some of the mutations may make the virus too attenuated. Mutations can also occur in the S gene which may also affect immunogenicity so that the desired immune response is affected and the potential vaccine may not protect against the required serotype. In addition there are problems associated with reversion to virulence and stability of vaccines.

It is important that new and safer vaccines are developed for the control of IBV. Thus there is a need for IBV vaccines which are not associated with these issues, in particular vaccines which may be used for in ovo vaccination.

SUMMARY OF ASPECTS OF THE INVENTION
The present inventors have used a reverse genetics approach in order to rationally attenuate IBV. This approach is much more controllable than random attenuation following multiple passages in embryonated eggs because the position of each mutation is known and its effect on the virus, i.e. the reason for attenuation, can be derived.

Using their reverse genetics approach, the present inventors have identified various mutations which cause the virus to have reduced levels of pathogenicity. The levels of pathogenicity may be reduced such that when the virus is administered to an embryonated egg, it is capable of replicating without being pathogenic to the embryo. Such viruses may be suitable for in ovo vaccination, which is a significant advantage and has improvement over attenuated IBV vaccines produced following multiple passage in embryonated eggs.

Thus in a first aspect, the present invention provides a live, attenuated coronavirus comprising a variant replicase gene encoding polyproteins comprising a mutation in one or more of non-structural protein(s) (nsp)-10, nsp-14, nsp-15 or nsp-16.

The coronavirus according to the first aspect of the invention has reduced pathogenicity compared to a coronavirus expressing a corresponding wild-type replicase, such that when the virus is administered to an embryonated egg, it is capable of replicating without being pathogenic to the embryo.

In a second aspect, the present invention provides a variant replicase gene as defined in connection with the first aspect of the invention.

In a third aspect, the present invention provides a protein encoded by a variant coronavirus replicase gene according to the second aspect of the invention.

In a fourth aspect, the present invention provides a plasmid comprising a replicase gene according to the second aspect of the invention.

In a fifth aspect, the present invention provides a method for making the coronavirus according to the first aspect of the invention which comprises the following steps:

• (i) transfecting a plasmid according to the fourth aspect of the invention into a host cell;
• (ii) infecting the host cell with a recombining virus comprising the genome of a coronavirus strain with a replicase gene;
• (iii) allowing homologous recombination to occur between the replicase gene sequences in the plasmid and the corresponding sequences in the recombining virus genome to produce a modified replicase gene; and
• (iv) selecting for recombining virus comprising the modified replicase gene.

The recombining virus may be a vaccinia virus.
The method may also include the step:

• (v) recovering recombinant coronavirus comprising the modified replicase gene from the DNA from the recombining virus from step (iv).

In a sixth aspect, the present invention provides a cell capable of producing a coronavirus according to the first aspect of the invention.

In a seventh aspect, the present invention provides a vaccine comprising a coronavirus according to the first aspect of the invention and a pharmaceutically acceptable carrier.

In an eighth aspect, the present invention provides a method for treating and/or preventing a disease in a subject which comprises the step of administering a vaccine according to the seventh aspect of the invention to the subject.

Further aspects of the invention provide:

• the vaccine according to the seventh aspect of the invention for use in treating and/or preventing a disease in a subject.
• use of a coronavirus according to the first aspect of the invention in the manufacture of a vaccine for treating and/or preventing a disease in a subject.

The disease may be infectious bronchitis (IB).

The method of administration of the vaccine may be selected from the group consisting of; eye drop administration, intranasal administration, drinking water administration, post-hatch injection and in ovo injection.

Vaccination may be by in ova vaccination.
The present invention also provides a method for producing a vaccine according to the seventh aspect of the invention, which comprises the step of infecting a cell according to the sixth aspect of the invention with a coronavirus according to the first aspect of the invention.

Coronavirus
Gammacoronavirus is a genus of animal virus belonging to the family Coronaviridae. Coronaviruses are enveloped viruses with a positive-sense single-stranded RNA genome and a helical symmetry.

The genomic size of coronaviruses ranges from approximately 27 to 32 kilobases, which is the longest size for any known RNA virus.

Coronaviruses primarily infect the upper respiratory or gastrointestinal tract of mammals and birds. Five to six different currently known strains of coronaviruses infect humans. The most publicized human coronavirus, SARS-CoV which causes severe acute respiratory syndrome (SARS), has a unique pathogenesis because it causes both upper and lower respiratory tract infections and can also cause gastroenteritis. Middle East respiratory syndrome coronavirus (MERS-CoV) also causes a lower respiratory tract infection in humans. Coronaviruses are believed to cause a significant percentage of all common colds in human adults.

Coronaviruses also cause a range of diseases in livestock animals and domesticated pets, some of which can be serious and are a threat to the farming industry. Economically significant coronaviruses of livestock animals include infectious bronchitis virus (IBV) which mainly causes respiratory disease in chickens and seriously affects the poultry industry worldwide; porcine coronavirus (transmissible gastroenteritis, TGE) and bovine coronavirus, which both result in diarrhoea in young animals. Feline coronavirus has two forms, feline enteric coronavirus is a pathogen of minor clinical significance, but spontaneous mutation of this virus can result in feline infectious peritonitis (FIP), a disease associated with high mortality.

There are also two types of canine coronavirus (CCoV), one that causes mild gastrointestinal disease and one that has been found to cause respiratory disease. Mouse hepatitis virus (MHV) is a coronavirus that causes an epidemic murine illness with high mortality, especially among colonies of laboratory mice.

Coronaviruses are divided into four groups, as shown below:

Alpha

• Canine coronavirus (CCoV)
• Feline coronavirus (FeCoV)
• Human coronavirus 229E (HCoV-229E)
• Porcine epidemic diarrhoea virus (PEDV)
• Transmissible gastroenteritis virus (TGEV)
• Human Coronavirus NL63 (NL or New Haven)

Beta

• Bovine coronavirus (BCoV)
• Canine respiratory coronavirus (CRCoV)—Common in SE Asia and Micronesia
• Human coronavirus OC43 (HCoV-OC43)
• Mouse hepatitis virus (MHV)
• Porcine haemagglutinating encephalomyelitis virus (HEV)
• Rat coronavirus (Roy). Rat Coronavirus is quite prevalent in Eastern Australia where, as of March/April 2008, it has been found among native and feral rodent colonies.
• (No common name as of yet) (HCoV-HKU1)
•  Severe acute respiratory syndrome coronavirus (SARS-CoV)
• Middle East respiratory syndrome coronavirus (MERS-CoV)

Gamma

• Infectious bronchitis virus (IBV)
• Turkey coronavirus (Bluecomb disease virus)
• Pheasant coronavirus
• Guinea fowl coronavirus

Delta

• Bulbul coronavirus (BuCoV)
• Thrush coronavirus (ThCoV)
• Munia coronavirus (MuCoV)
• Porcine coronavirus (PorCov) HKU15

The variant replicase gene of the coronavirus of the present invention may be derived from an alphacoronavirus such as TGEV; a betacoronavirus such as MHV; or a gammacoronavirus such as IBV.

As used herein the term “derived from” means that the replicase gene comprises substantially the same nucleotide sequence as the wild-type replicase gene of the relevant coronavirus. For example, the variant replicase gene of the present invention may have up to 80%, 85%, 90%, 95%, 98% or 99% identity with the wild type replicase sequence. The variant coronavirus replicase gene encodes a protein comprising a mutation in one or more of non-structural protein (nsp)-10, nsp-14, nsp-15 or nsp-16 when compared to the wild-type sequence of the non-structural protein.

IBV
Avian infectious bronchitis (IB) is an acute and highly contagious respiratory disease of chickens which causes significant economic losses. The disease is characterized by respiratory signs including gasping, coughing, sneezing, tracheal rales, and nasal discharge. In young chickens, severe respiratory distress may occur. In layers, respiratory distress, nephritis, decrease in egg production, and loss of internal egg quality and egg shell quality are common.

In broilers, coughing and rattling are common clinical signs, rapidly spreading in all the birds of the premises. Morbidity is 100% in non-vaccinated flocks. Mortality varies depending on age, virus strain, and secondary infections but may be up to 60% in non-vaccinated flocks.

The first IBV serotype to be identified was Massachusetts, but in the United States several serotypes, including Arkansas and Delaware, are currently circulating, in addition to the originally identified Massachusetts type.

The IBV strain Beaudette was derived following at least 150 passages in chick embryos. IBV Beaudette is no longer pathogenic for hatched chickens but rapidly kills embryos.

H120 is a commercial live attenuated IBV Massachusetts serotype vaccine strain, attenuated by approximately 120 passages in embryonated chicken eggs. H52 is another Massachusetts vaccine, and represents an earlier and slightly more pathogenic passage virus (passage 52) during the development of H120. Vaccines based on H120 are commonly used.

IB QX is a virulent field isolate of IBV. It is sometimes known as “Chinese QX” as it was originally isolated following outbreaks of disease in the Qingdao region in China in the mid 1990s. Since that time the virus has crept towards Europe. From 2004, severe egg production issues have been identified with a very similar virus in parts of Western Europe, predominantly in the Netherlands, but also reported from Germany, France, Belgium, Denmark and in the UK.

The virus isolated from the Dutch cases was identified by the Dutch Research Institute at Deventer as a new strain that they called D388. The Chinese connection came from further tests which showed that the virus was 99% similar to the Chinese QX viruses. A live attenuated QX-like IBV vaccine strain has now been developed.

IBV is an enveloped virus that replicates in the cell cytoplasm and contains an non-segmented, single-stranded, positive sense RNA genome. IBV has a 27.6 kb RNA genome and like all coronaviruses contains the four structural proteins; spike glycoprotein (S), small membrane protein (E), integral membrane protein (M) and nucleocapsid protein (N) which interacts with the genomic RNA.

The genome is organised in the following manner: 5′UTR—polymerase (replicase) gene—structural protein genes (S-E-M-N)—UTR 3′; where the UTR are untranslated regions (each ˜500 nucleotides in IBV).

The lipid envelope contains three membrane proteins: S, M and E. The IBV S protein is a type I glycoprotein which oligomerizes in the endoplasmic reticulum and is assembled into homotrimer inserted in the virion membrane via the transmembrane domain and is associated through non-covalent interactions with the M protein. Following incorporation into coronavirus particles, the S protein is responsible for binding to the target cell receptor and fusion of the viral and cellular membranes. The S glycoprotein consists of four domains: a signal sequence that is cleaved during synthesis; the ectodomain, which is present on the outside of the virion particle; the transmembrane region responsible for anchoring the S protein into the lipid bilayer of the virion particle; and the cytoplasmic tail.

All coronaviruses also encode a set of accessory protein genes of unknown function that are not required for replication in vitro, but may play a role in pathogenesis. IBV encodes two accessory genes, genes 3 and 5, which both express two accessory proteins 3a, 3b and 5a, 5b, respectively.

The variant replicase gene of the coronavirus of the present invention may be derived from an IBV. For example the IBV may be IBV Beaudette, H120, H52, IB QX, D388 or M41.

The IBV may be IBV M41. M41 is a prototypic Massachusetts serotype that was isolated in the USA in 1941. It is an isolate used in many labs throughout the world as a pathogenic lab stain and can be obtained from ATCC (VR-21™). Attenuated variants are also used by several vaccine producers as IBV vaccines against Massachusetts serotypes causing problems in the field. The present inventors chose to use this strain as they had worked for many years on this virus, and because the sequence of the complete virus genome is available.

The M41 isolate, M41-CK, used by the present inventors was adapted to grow in primary chick kidney (CK) cells and was therefore deemed amenable for recovery as an infectious virus from a cDNA of the complete genome. It is representative of a pathogenic IBV and therefore can be analysed for mutations that cause either loss or reduction in pathogenicity.