Nearly 100 million people have received at least one dose of a Covid-19 vaccine, either as part of clinical trials or after their government’s regulatory agency approved the vaccines. They are based on different technologies that have advantages and disadvantages1. The clinical trials showed that all the approved vaccines were safe and effective. At the beginning of the first trials, the original SARS-CoV-2 virus that originated in Wuhan was widespread. By March, a variant with a mutation in amino acid number 614 became more widespread. The original virus had an aspartic acid (abbreviated as D) at this position. This has been changed to a glycine (G) in variants that have become dominant, due to its higher transmissibility (higher viral load, wider distribution of viral replication)2. The mutation is abbreviated as D614G. Amino acid 614 is in the spike (S) protein that binds to the human ACE2 receptor that is found on the outer portion of cell membranes and is crucial for heart and lung health. The presence of a G instead of D made this variant more stable and transmissible. The density of S proteins on the surface of the virus increased four to five times over the original form. As a result, each mutant virion particle can infect human host cells more efficiently. This could help explain why the SARS-CoV-2 virus has spread so rapidly throughout most of the Americas. The G614 genotype was not detected in February (among 33 sequences) and existed at low frequency in March (26%), but increased rapidly by April (65%) and May (70%), indicating a transmission advantage over viruses with the D614 genotype3. Others have estimated that it was almost 100% prevalent by the end of April4. Fortunately, both genotypes (D614 and G614) are similarly susceptible to neutralizing antisera, indicating that antibody-mediated control of viruses carrying either genotype would be similar.
So, clinical trials were effective against these two forms of the virus. Recently, a potentially more transmissible variant has emerged in England5. It’s been named VUI 202012/01 (Variant Under Investigation, year 2020, month 12, variant 01). It’s also referred to by its lineage, B.1.1.7. However, mutations are naturally expected for viruses and are most often simply neutral regional markers useful for contact tracing. The mutations seen previously rarely have affected viral fitness and almost never affect clinical outcome, but the detailed effects of these mutations remain to be determined fully. The UK NERVTAG group of experts concluded, 'In summary, NERVTAG has moderate confidence that VUI-202012/01 demonstrates a substantial increase in transmissibility compared to other variants'6. Still, there is some concern that one or more of the vaccines being used may not be very effective against either VUI-202012/01 or future variants that are bound to emerge. So, the international scientific community is monitoring the evolution of this virus. We are using every resource available (including artificial intelligence) to predict the effects of mutations on viral transmissibility. This effort is being coordinated, in large part by the Global Initiative on Sharing All Influenza Data (GISAID)3. They have been doing the same thing for the influenza virus since 2008, enabling us to change the flu vaccine each year so it will be as effective as possible against the most widespread variants. At this time, we don’t know how long immunity will last with the various Covid-19 vaccines or the frequency with which adverse side effects will occur, but we are collecting and analyzing many data. This is called a Phase 4 trial, or post-market surveillance. So, the aims of this month’s article are to describe the progress being made in vaccinating millions of people and provide more information about mutations in the SARS-CoV-2 virus.
Progress in vaccinations
In July, China started by vaccinating some of its military personnel with the vaccine from CanSino called Ad5-nCoV7,8. It was safe and induced significant immune responses in the majority of recipients after a single immunization according to a publication in The Lancet8. Also, Sinovac and Sinopharm have vaccinated many with their Coronavac vaccine9. It can be stored and transported in normal refrigerators, making it especially useful in regions that lack cold storage capacities. China approved the vaccine for general public use while promising free vaccinations. They aim to vaccinate 50 million people ahead of New Year celebrations in February. That’s in addition to the 1.5 million doses already given to high-risk groups. The Sinovac vaccine has also been approved in Singapore, Brazil, the United Arab Emirates (UAE), Turkey, Indonesia and Bahrain, where vaccinations have begun. Then, Russia started vaccinating some of its military personnel in August with its Sputnik V vaccine10,11. Sputnik V was approved in Algeria, Argentina, Belarus, Bolivia, Guinea, Palestine, Paraguay, Palestine, Russia, Serbia, and Venezuela. Mexico, India, Nepal, Egypt, Brazil, Uzbekistan and Nepal also plan to buy the vaccine. On 30 Dec., the UK Medicines and Healthcare products Regulatory Agency (MHRA) approved AstraZeneca-Oxford’s AZD1222, also known as COVISHIELD12, followed by India on 3 Jan., 2021. Bangladesh, Argentina, the Dominican Republic, El Salvador, Mexico and Nepal have also approved it. These vaccines use an attenuated adenovirus to produce the S protein that activates and trains the immune system1. India also approved Bharat Biotech’s COVAXIN (also known as BBV152)13. It is based on an inactivated SARS-CoV-2 virus. The original, active virus was isolated from tourists who arrived in New Delhi and is the most prevalent strain in India. Its genome was deposited in the GISAID database as EPIISL420545. It’s 99.97% identical to the original strain isolated in Wuhan, China. Sinophram in China has another vaccine, BBIBP-CorV, which is based on an inactivated SARS-CoV-2 virus. It’s been approved in China, Egypt, Jordan, the UAE and Bahrain. Bharat Biotech also entered into a licensing agreement with the Washington University School of Medicine in St. Louis, Missouri (USA) for a novel “chimp-adenovirus” (adenovirus that infects chimpanzees), single-dose intranasal vaccine for Covid-19.
Meanwhile, Johnson & Johnson’s Janssen Pharmaceutical Companies has a vaccine candidate (JNJ-78436735) that also uses an attenuated adenovirus, but it only requires one dose14,15. It is in Phase 3 clinical trial (called ENSEMBLE) that is being conducted in Argentina, Brazil, Chile, Colombia, Mexico, Peru, South Africa and the U.S. It also expects to include people with and without comorbidities linked to increased risk for progression to severe disease. In the U.S. the trial includes significant representation of Black, Hispanic/Latinx, American Indian and Alaskan Native volunteers.
Another vaccine candidate from Novavax (NVX-CoV2372) has begun a Phase 3 clinical trial16. It’s based on a recombinant SARS-CoV glycoprotein that is encapsulated within lipid nanoparticles. It elicited immunity in baboons and immunoprotection in mice. Purified NVX-CoV2373 S proteins from 27.2 nm nanoparticles that are thermostable and bind with high affinity to the human ACE2 receptor17. The vaccine also contains a Matrix-MTM adjuvant that contains saponins (triterpene glycosides found in many plants)18. It stimulates the entry of antigen-presenting cells into the injection site and enhances antigen presentation in local lymph nodes, thus boosting immune response. Novavax plans to deliver 10 million doses of NVX CoV2373 that could be used in Phase 2/3 trials, or under an Emergency Use Authorization if approved by the FDA. They can increase that to 100 million, if needed. They have a recent history of success - quadrivalent influenza (flu) vaccine. It met all primary objectives in its pivotal Phase 3 clinical trial in older adults18.
As described in last month’s article, two vaccines based on mRNA technology were approved in December 2020 (BNT162b2 by Pfizer/BioNTech and mRNA-1273 by Moderna/Lonza)19. They are now being administered to many people. The Pfizer/BioNTech vaccine has been approved in the U.S., Argentina, Chile, Colombia, Ecuador, Iraq, Jordan, Oman, Panama, the Philippines, Costa Rica, the United Kingdom (U.K.), the European Union (EU), Switzerland, Canada, Israel, Bahrain, Kuwait, Qatar, Saudia Arabia, Serbia, Slovakia and the World Health Organization (WHO). The Moderna vaccine has been approved in the U.S., Israel, the EU and Switzerland. They contain a self-replicating mRNA that encodes the entire S protein that the SARS-CoV-2 virus uses to bind to ACE-2 receptors on the membranes of host cells in the nose, lungs, heart and many other parts of the body. The mRNA is packed into lipid nanoparticles. When these particles are injected into the human body, they fuse to the cell membrane of human cells, allowing the mRNA to gain access to the host cell’s enzymes that are needed to translate it into the many copies of the S protein. Once the viral proteins are released into the bloodstream, the immune system produces antibodies against them. Both vaccines require a booster shot, about 21 (Pfizer/BioNTech) or 28 days (Moderna/Lonza) after the first one. Both vaccines were over 90% effective in Phase 2/3 clinical trials. It might be wise to get a good night’s sleep, drink no alcohol and don’t take ibuprofen or acetaminophen the day before being vaccinated. However, both ibuprofen and acetaminophen are very useful in reducing pain after being vaccinated20. The Moderna vaccine can be stored at -20 °C in freezers that are widely available in doctors’ offices and pharmacies, like currently used vaccines such as chickenpox. It can also be stored in a refrigerator for up to 30 days. Pfizer’s vaccine must be stored at -75 °C. The difference in stability is due to differences in the contents of the lipid nanoparticles.
The lipid nanoparticles used in Moderna’s mRNA-1273 vaccine contain four different lipids21,22:
- SM-102 — a proprietary phospholipid that makes up the basic structure of the nanoparticle wall.
- DSPC — another commercially available phospholipid that will aid in the nanoparticle wall structure.
- Cholesterol — native to all of our cell membranes, cholesterol helps to add fluidity or stiffness to a cell membrane depending on temperature. The addition of cholesterol will aid in the stability of the nanoparticle.
- PEG-2000 DMG — a lipid fused to polyethylene glycol which aids in the formation of the nanoparticle.
- mRNA – is translated into the viral S protein.
The lipid nanoparticles used in Pfizer’s BNT162b2 vaccine contain four different lipids22,23:
- (4-hydroxybutyl)azanediyl)bis(hexane-6,1-diyl)bis(2-hexyldecanoate), ALC-0315 - Proprietary phospholipid that makes up the basic structure of the nanoparticle wall. These molecules are modeled after native phospholipids found in living cells.
- 2-[(polyethylene glycol)-2000]-N,N-ditetradecylacetamide, ALC-0159 — Another commercially available phospholipid linked to polyethylene glycol.
- 1,2-distearoyl-sn-glycero-3-phosphocholine — A commercially available glycophospholipid used as a component of the cell wall.
- Cholesterol — A native molecule ubiquitous in the human body. Cholesterol aids in the fluidity or rigidity of a lipid membrane depending on temperature.
There were some allergic reactions to the Pfizer/BioNTech vaccine in a few people who had a history of suffering anaphylactic shock24,25. It may have been due to the components that contain polyethylene glycol (PEG)24. PEG is a polymer that is used in many medicines, foods and cosmetics. It contains ethylene glycol repeating units, -(CH2CH2O)n -. Also, Polysorbate 80 contains these units and may also cause allergic reactions. Skin prick and intradermal tests are available to test for allergies to PEG and Polysorbate 801. According to the WHO, “Any person with a history of anaphylaxis to a vaccine, medicine or food should not receive the Pfizer BioNTech vaccine”25.
New variants of the virus
A key question has emerged. Will these and other vaccines still be effective against new variants of the SARS-CoV-2 virus that are formed by mutations? Unfortunately, another variant (501Y.V2) has emerged26. This variant emerged in South Africa after the first epidemic wave in the worst affected metropolitan area within the Eastern Cape Province. This was followed by the rapid spread. Now, it is the dominant variant in Eastern and Western Cape Provinces. It has eight mutations in the receptor-binding domain of the S protein. This includes K417N, E484K and N501Y) at amino acids 417, 484 and 501. That is, lysine (K) at amine acid 417 was changed to asparagine (N), glutamic acid (E) at amino acid 484 was mutated to lysine (K) and asparagine (N) at amino acid 501 was mutated into tyrosine (Y). These mutations may make this variant more transmissible. The K417N mutation appeared independently in the U.S. and several European countries. This mutation is associated with escape from neutralization by monoclonal and polyclonal antibodies in blood serum27. This could make some vaccines less effective in some people.
The variant with a mutation in amino acid number 614 (D614G) and higher transmissibility emerged in March 2020 and quickly became the predominant version of the virus3. Many other mutations have appeared since then, and scientists have deposited their genomes into the GISAID database. In December, the United Kingdom reported the emergence of the VUI 202012/01 variant that has even higher transmissibility. It was defined by a total of 17 mutations, with 10 of them in the part of the mRNA that codes for the spike (S) protein (deletion 69-70, deletion 145, N501Y, A570D, D614G, P681H, T716I, S982A, D1118H, where D, P, H, T and I are aspartic acid, proline, histidine, threonine and isoleucine, respectively). An increasing fraction in Southern England (all from clade GR) share several of these mutations and a handful have been seen through imports in other countries. Based on evaluation of the effect on virus structure and function, the most relevant might be N501Y (asparagine, N, in the previous variants, mutated into tyrosine, Y)3. Still, scientists are urging calm when communicating with most people28. They echo the advice of the WHO when they say that viral mutations are quite common and to be expected. However, the first rule of communication is “know your audience”. So, this is good advice for the educated, international audience that read the Wall Street International. However, in my opinion, it is not very good advice for most of the 70 million people in the USA who voted for Donald Trump, who chose not to wear masks in public and then gathered in large groups (like the rioters in the Capitol on 6 January). They should be advised not to be so calm about the pandemic, but take it seriously and wear masks. Still, by not wearing masks, many of the rioters were quickly identified by police and arrested.
The first case of re-infection from genetically distinct SARS-CoV-2 lineage in Brazil was reported on 6 January 202129. It had the E484K mutation in the S protein, which is associated with escape from neutralizing antibodies. The new variant found in Brazil belongs to the B.1.1.248 lineage of the coronavirus and has 12 mutations, including N501Y and E484K. It has become widespread in several Brazilian states. This variant (or lineage) was also described previously in the U.K., the U.S., Australia and Portugal.
On 13 January 2021, researchers at the Ohio State University announced the emergence of the Columbus strain, named COH.20G/501Y30. It has three mutations not seen before. It appears to have become the most widespread form of Covid in Columbus, Ohio.
In the meantime, it appears that repeat infections by the most widespread lineages (variants) of the SARS-CoV-2 virus are rare31. That is, almost all people who have tested positive for the virus in the past will not become re-infected for several months. Still, that could change, as escape mutants emerge.
It is very interesting to compare the results of Chinese and American efforts. In the USA there have been over 400,000 deaths, 24 million cases and continuing problems with sufficient and accurate testing in some areas. China has had only 4635 deaths and 88,118 cases (as of 16 January 2021). They have tested over 160 million people. Moreover, China recently locked down Shijiazhuang, a city of 11 million, after only 117 people were found to have the SARS-CoV-2 virus. The majority (67) were asymptomatic.
1 Smith, R.E. Developing vaccines and treatments for Covid-19. Progress report, 24 June, 2020. Wall Street International, 2020.
2 Zhang, L. et al. The D614G mutation in the SARS-CoV-2 spike protein reduces S1 shedding and increases infectivity. Scripps Research, 2020.
3 Zhang, L. et al. The D614G Mutation in the SARS-CoV-2 Spike Protein Reduces S1 Shedding and Increases Infectivity. Scripps Research, 2020.
4 Cortey, M. et al. SARS-CoV-2 Amino Acid Substitutions Widely Spread in the Human Population Are Mainly Located in Highly Conserved Segments of the Structural Proteins. BioRxiv preprint, 17 May, 2020.
5 GISAID. UK reports new variant, termed VUI 202012/01. Dec., 2020.
6 NERVTAG meeting minutes. 18 Dec., 2020.
7 Smith, R.E. China starts to vaccinate its military personnel. Developing vaccines and treatments for Covid-19. Progress report. Wall Street International, 24 July, 2020.
8 Zhu F-C et al. Safety, tolerability, and immunogenicity of a recombinant adenovirus type-5 vectored Covid-19 vaccine: a dose-escalation, open-label, non-randomised, first-in-human trial. The Lancet, Volume 395, p. 1845-1854, 24 June, 2020.
9 Biorender. Covid-19 vaccine & therapy tracker: Ad5-nCoV, 2021.
10 Smith, R.E. Russia starts to vaccinate its military personnel. Developing vaccines and treatments for Covid-19. Progress report. Wall Street International, 24 Aug., 2020.
11 Sputnik V. The first registered Covid-19 vaccine, 2021.
12 AstraZeneca. AstraZeneca’s Covid-19 vaccine authorised for emergency supply in the UK. 30 Dec., 2020.
13 Ganneru, B. et al. Evaluation of safety and immunogenicity of an adjuvanted, TH-1 skewed, whole virion inactivated SARS-CoV-2 vaccine – BBV152. bioRx preprint, 12 Sept. 2020.
14 Terry, M. J&J's One-dose Covid-19 vaccine candidate begins phase III trial. 23 Sept., 2020.
15 Johnson & Johnson prepares to resume Phase 3 ENSEMBLE trial of its Janssen Covid-19 vaccine candidate in the U.S. 23 Oct., 2020.
16 NIH. Phase 3 trial of Novavax investigational Covid-19 vaccine opens. 28 Dec., 2020.
17 Tian J-H. et al. SARS-CoV-2 spike glycoprotein vaccine candidate NVX-CoV2373 elicits immunogenicity in baboons and protection in mice. bioRxiv preprint 30 June, 2020.
18 Novavax wins $1.6B in “warp speed” funding; plans first 100M doses of Covid-19 vaccine. Genetic Engineering & Biotechnology News, 7 July, 2020.
19 Smith, R.E. Vaccines based on modern RNA technology. This technology's potential for vaccines and other diseases. 24 Dec., 2020. Wall Street International, 24, 2020.
20 CDC. What to expect after getting a Covid-19 vaccine. Jan., 2021.
21 ModernaTX, Inc. Protocol mRNA-1273-P301, Amendment 3. 20 Aug., 2020.
22 Smith, J. Decoding Pfizer’s Covid-19 vaccine ingredients. Medium coronavirus blog. 10 Dec., 2020.
23 Pfizer and BioNTech. Vaccines and related biological products advisory committee meeting. 10 December, 2020.
24 Cabanillas, B. et al. Allergic reactions to the first Covid-19 vaccine: a potential role for polyethylene glycol. 15 Dec., 2020.
25 World Health Organization. mRNA vaccines against Covid-19: Pfizer-BioNTech Covid-19 vaccine BNT162b2. 22 Dec., 2020.
26 Tegally, H. et al. Emergence and rapid spread of a new severe acute respiratory syndrome-related coronavirus 2 (SARS-CoV-2) lineage with multiple spike mutations in South Africa. medRxiv, preprint, 22 Dec., 2020.
27 Thomson, E.C. et al. The circulating SARS-CoV-2 spike variant N439K maintains fitness while evading antibody-mediated immunity. bioRxiv. 2020.
28 Scientists urge calm about coronavirus mutations, which are not unexpected. New York Times, 21 Dec, 2020.
29 Nonaka, C.K.V. et al. Genomic evidence of a SARS-CoV-2 reinfection case with E484K spike mutation in Brazil. Preprints, 6 Jan., 2021.
30 Leitch, M. Researchers discover new variant of Covid-19 virus in Columbus, Ohio. 13 Jan., 2021.
31 Leford, H. Covid reinfections are unusual — but could still help the virus to spread. Nature News, 14 Jan., 2021.