COVID-19 – Viral Vector Versus mRNA Vaccines

In January this year, we published a blog that covered the development of the first two COVID-19 vaccines that were given emergency use authorization (EUA) (The Promise and Hope of RNA-based vaccines for COVID-19), and which are both RNA-based vaccines. Since then, other vaccines have been granted emergency use authorization in the U.S. and elsewhere. Those new vaccines are based on a different technology, that uses alternative engineered viral vectors. This post will cover the viral vector vaccine technology.

A quick recap on the various vaccine types

The vaccines in development cover a broad design range and fall into four general categories (Figure 1 provides an overview of the different vaccine types):

1. Attenuated and inactivated vaccines which rely on wild-type or natural versions of SARS-CoV-2 that have been chemically or physically treated to make the virus either completely inactive or very weakened with regard to its infectious potential. This is an older but tried-and-true strategy. An example for this type of vaccine is the inactivated polio or Salk vaccine.
   1. Examples for this type include:
      1. Several inactivated SARS-CoV-2 vaccines are being developed in China including Coronavac, a product of Sinovac Biotech in China, which currently is in phase III trial.
      1. Codagenix has partnered with the Serum Institute of India to develop a live-attenuated SARS-CoV-2 vaccine.
2. Nucleic acid vaccines which are based on engineered DNA or RNA molecules that code for one or more of the various SARS-CoV-2 proteins or parts thereof. This is generally a newer and less well established strategy. Stand-out examples for this type of vaccine are the Pfizer-BioNTech B162B2 and Moderna mRNA-1273 vaccines, both of which are currently in use.
   1. Another example:
      1. Inovio Pharmaceuticals is developing INO-4800 a plasmid DNA vaccine for COVID-19 for electroporation delivery.
3. Subunit vaccines where the immunogenic element(s) is a protein or protein fragment of SARS-CoV-2. One of the better known examples of a subunit vaccine already in use is Recombivax HB, based on a viral envelope protein and used for prevention of Hepatitis B infection.
   1. Examples include:
      1. Novavax is developing NVX-CoV2373 which contains SARS-CoV-2 spike proteins produced in a baculovirus expression system and carried in a synthetic lipid nanoparticle.
      1. Another interesting approach is that of ZF2001 from Anhui Zhifei Longcom Biopharmaceutical which uses a dimeric form of the receptor-binding domain (RBD) of the SARS-CoV-2 spike protein as the antigen.
4. Viral vector vaccines which carry as the immunizing agent some part of the SARS-CoV-2 coding sequence inserted into the modified backbone of another virus, for instance an Adenovirus.A prototype example for this technology is the recombinant vesicular stomatitis virus (rVSV)-based vaccine against Zaire Ebola virus (rVSV-ZEBOV) licensed in the EU and US for combatting Ebola infection.
   1. Examples of this type include the four vaccines we will cover in this blog: Ad26.COV2.S (Johnson & Johnson), AZD1222 (Oxford and AstraZeneca), Sputnik V (Gamaleya), and Ad5-nCoV (CanSino).

Adenovirus-based vaccines

The vaccines we will discuss in this post are all created by modification of human or primate adenovirus strains (Figure 2), and summarized in the Forni et al. paper (2021). In each case, the native virus has been engineered to be replication incompetent. Each technology choice has benefits and potential pitfalls. Human adenoviruses are good candidates as they can be manufactured at scale, have well understood safety profiles, and are highly immunogenic (Ewer et al., 2021). However, pre-existing immunity to adenovirus vectors can potentially hamper use as was outlined by Dong and team (2020). Primate adenovirus offers some of the potential benefits while hopefully limiting pre-existing immunity issues. Adenoviral vector vaccines are also generally less heat-labile so supply-chain cold transport challenges should be a less significant issue compared to mRNA-based vaccines.

Figure 1: An overview of the various vaccines types. Image credits: A systematic review of SARS-CoV-2 vaccine candidates.

Figure 2: Adenovirus-based vaccines. Images credits: COVID-19 vaccines: where we stand and challenges ahead.

Ad26.COV2.S (Johnson & Johnson)

The next vaccine that received emergency use authorization in the U.S. (February 27, 2021) and EU (March 11, 2021) – following the Moderna and Pfizer/BionTech ‘front-runners’ – was the one developed by Janssen Biotech (Johnson & Johnson). This third vaccine, named Ad26.COV2.S, uses a derivative of adenovirus vectors at its core (Forni et al., 2021), and is contained in a recombinant, replication-incompetent adenovirus serotype 26 (Ad26) vector. Sadoff and team summarized its phase 1-2a clinical trial results in January, 2021. At the time of writing this post, the CDC and FDA have called for pause on the J&J vaccine after blood clot cases.

Ad26.COV2.S highlights

The design and engineering details of the vector and most importantly the immunogen was described in the very interesting Bos et al. paper (2020).

• The S protein gene in the Ad26.COV2.S vaccine is a full-length gene from the SARS-CoV-2 isolate Wuhan-Hu-1 (Genbank accession number MN908947).
• The gene is codon-optimized for expression in human cell lines.
• Critically, the furin cleavage site was mutationally abolished by amino acid changes R682S and R685G.
  • The protein was stabilized in the prefusion conformation by 2 proline substitutions (K986P and V987P) at the apex of the central helix and heptad repeat 1. The stabilization with proline mutations is the same made for the mRNA-1273 (Corbett et al., 2020) and BNT162b2 (Walsh et al., 2020) however the abrogation of the furin site is a major difference.
  • The furin site mutations were found to yield the highest ratio of neutralizing to non-neutralizing antibody in mouse models.

• Although, various single and multi-dose regimens were investigated in early trials (Sadoff et al., 2021) the EUA for Ad26.COV2.S is for a single-shot option.
• Unlike the two mRNA vaccines Ad26.COV2.S does not need to be stored frozen. The vaccine is stable for at least three months at 2-8 °C.
• According to the FDA briefing document the vaccine efficacy against moderate to severe/critical COVID-19 across all trials was 66.9%.
• Full results from the phase III trials are not yet published in a peer-reviewed journal.

AZD1222 (Oxford and AstraZeneca) – marketed as Vaxzevria

AZD1222 is another vaccine that has been granted EUA in the EU (January 29, 2021) and is made via a collaborative effort between Oxford University and AstraZeneca. Unfortunately, the AZD1222 vaccine has also been linked to very rare cases of unusual blood clots. It appears as though EUA for AZD1222 in the US is imminent but an exact date is uncertain.

• AZD1222 also known as ChAdOx1 nCov-19, was developed at Oxford University, and uses a replication-deficient vector that is derived from the simian adenovirus Y25 serotype (van Doremalen et al., 2020). It is thought that use of a non-human adenovirus will limit potential pre-existing anti-vaccine antibody interference.
• AZD1222 expresses a full-length (amino acids-2-1273), codon optimized SARS-CoV-2 spike protein with the leader sequence from tissue-plasminogen activator (tPA).
• Interestingly, the form of the Spike protein produced by AZD1222 does not carry the stabilizing double proline substitutions present in many of the vaccines thus far put into wide use.
  • Though, not tested in the context of the simian adenovirus, Bos et al. (2020) found that using the wild-type signal peptide (SP) rather than tPA leads to more correctly processed and folded mature Spike protein. This in turn leads to more induction of S binding antibodies and higher Neutralizing antibody (NAbs) titer.
  • Similarly, proline stabilizing mutations and abrogation of the furin site leads to highest quality immune response. Whether these difference between AZD1222 and Ad26.COV2.S (J&J vaccine) lead to consequences in real-world large-scale use remains to be seen.
• AZD1222 (ChAdOx1 nCov-19) was tested in mouse and rhesus macaques model organisms early on and found to be capable of generating balanced humoral and cellular immune responses characterized as mainly TH-1, as well as inhibiting disease (van Doremalen et al., 2020).
• Single dose versus multiple dose regimens, safety and immunogenicity of AZD1222 were well characterized in Phase I/II trials recently published in a pair of papers (Barrett et al, 2021, Ewer et al., 2021).
• The first interim phase III results for AZD1222 were reported in December of 2020 and formally published this year (Voysey et al., 2021).
  • Overall, vaccine efficiency across two different two-dose regimens was over 70%.
• AZD1222 is stable when stored at 2-8 °C.
• AZD1222 is based on a modified version of a chimpanzee adenovirus.
• Because of the unique combination of genome surveillance in the UK and the locations of AZD1222 trials there is some practical data on efficacy versus variant strains. Emary et al. (2021) show that AZD1222 is still very effective against the B.1.1.7. variant. An analysis of limited clinical trial results of AZD1222 showed that the vaccine may not be effective against  the B.1.351 viral variant (Madhi et al., 2021).

Sputnik V (Gamaleya)

A vaccine that has garnered much international attention is the one developed by the Gamaleya Research Institute in Moscow and termed Sputnik V. Perception of Sputnik V has certainly been colored significantly by international politics but there are several peer-reviewed publications that make a convincing case on the scientific front (Logunov et al., 2020, Logunov et al., 2021).

• Sputnik V is unique amongst the current COVID-19 vaccines in that it is based on a two component prime-boost design. The first dose is a recombinant human adenovirus type 26 vector and the second dose is a recombinant adenovirus type 5 vector where the antigen encoded in each case is identical.
  • It is hoped that by switching adenovirus types between the two injections the most robust immune response will be generated while minimizing the effects of potential pre-existing immune responses to the viral vector components (Logunov et al., 2020).
  • Both vector components carry the gene for full-length glycoprotein S but no further information is available on the types of modifications or changes to the spike sequence like those described above for Ad26.COV2.S and AZD1222.
• Interestingly, the vaccine is manufactured in two formulations, one frozen (storage at -18°C) and one lyophilized (storage at 2-8 °C). The lyophilized form could be particularly useful in remote areas where cold storage is difficult.
• In a large phase 3 trial in Russia greater than 16,000 subjects received the full two dose regimen, and vaccine efficacy was determined to be 91.6% (Logunov et al., 2021). Most adverse events were not serious and the overall safety was considered good.

Ad5-nCoV (CanSino)

A fourth adenovirus vector vaccine, Ad5-nCoV, has been developed by the Beijing Institute of Biotechnology in conjunction with CanSino Biologics which is based in China.

• Ad5-nCoV uses a replication defective human adenovirus type 5 vector similar to one of the two components of Sputnik V.
• Ad5-nCoV expresses an optimized full-length spike gene based on Wuhan-Hu-1 with the tPA signal peptide (Zhu et al., 2020). No other description of alterations or additions to the antigen sequence was found.
• The vaccine was phase I trialed at three different dose levels with a single vaccination-regimen.
  • The lowest two doses was advanced to a phase II of which the results suggested that the lowest dose would be used in phase III studies (Zhu et al., 2020b).
  • No phase III trial results have yet been published, and trials are currently proceeding in multiple countries.
• As per a recent February 2021 announcement, the single shot regimen has a 65.3% efficacy in preventing symptomatic COVID-19 and 90% efficacy against severe disease.
• Ad5-nCoV has the advantage of being a single dose regimen like Ad26.COV2.S.
• Ad5-nCoV is also stored at 2-8 °C – potentially reducing supply chain stress.

How things stack up: viral vector vaccines versus mRNA vaccines

All four of the vaccines we have discussed in this post share the essential features of viral vector vaccines. The material used for actual vaccinations consists of mature virus particles grown in the lab and by their external construction very much look like wild-type virus. Thus, the native virus’ ability to infect human cells is preserved and exploited to deliver the DNA encoding the antigen (Forni et al., 2021). In all four cases, the antigen coding sequence is some version of the Spike glycoprotein. Once delivered by the virus, the DNA must transit to the nucleus, be copied into an mRNA and then translated into protein. These proteins produced by the human subjects’ own cells are the source for generation of immune reactions, both humoral and cell-based (Rawat et al., 2021).

The mRNA vaccines we discussed previously (The Promise and Hope of RNA-based vaccines for COVID-19) share much of the characteristics of the viral vector vaccines. They are produced in the lab but in the case of mRNA vaccines the production is enzymatic and completely in vitro, so no requirement of a culture growth step. The mRNA molecules are encapsulated in lipid nanoparticles and therefore share some physical-chemical properties with adenovirus vectors. While the vector vaccines are more directed and designed to infect human cells, the mRNA vaccines depend on non-directed cellular fusion events to reach their target. Once the mRNA is inside the cells and reaches the correct compartment for translation, the process is very much the same as for the viral vector vaccines. Compared to vector vaccines no transcription step is required. mRNA vaccines use specially engineered RNA nucleotides to enhance stability and translation. However, the protein products produced by the mRNA-1273 (Moderna) and BNT-162b2 (Pfizer-BioNTech) are very similar to the ones produced by the four adenovirus vaccines.

To summarize…

Adenovirus vaccines

• Adenovirus vector vaccine particles are more complicated and difficult to produce than equivalent mRNA vaccines.
• Adenovirus vaccines are potentially subject to an immune reaction due to pre-existing exposure to viral vector component
• Adenovirus vaccines are generally more heat-tolerant than mRNA vaccines.
• Adenovirus vaccines are inherently good at infecting human cells.
• Adenovirus vaccines – both the J&J and AstraZeneca vaccines – have been associated with very rare blood clots that are not fully understood.

mRNA vaccines

• mRNA vaccines are likely to be more quickly re-engineered with altered antigen to meet consequences of viral mutation and variation.
• mRNA vaccines are theoretically incapable of recombination with host DNA so genetic effects and/or viral reversion are prevented.
• mRNA vaccines are produced completely in vitro so pose less risk of biological contamination.

Companies

Johnson & Johnson (J&J) is a very large publicly-traded healthcare company with more than 130K employees and founded in 1886. J&J engages in the research and development, manufacture and sale of a range of products in the healthcare field. J&J is headquartered in New Brunswick, New Jersey, and trades on the NYSE under the symbol JNJ. Janssen is the pharmaceutical arm of J&J and is headquartered in Antwerp, Belgium. Janssen’s focus is on six areas of medicine; oncology, immunology, neuroscience, pulmonary hypertension, infectious disease/vaccines and cardiovascular/Metabolism. Within the infectious disease area, the major efforts are in HIV, respiratory infection, Hepatitis B, pathogens of global concern and bacterial infections.

AstraZeneca is a global pharmaceutical company that discovers, develops, manufactures, and markets prescription medicines. AstraZeneca is publicly-traded company, has more than 10K employees worldwide, and the company stock trades on the NASDAQ under the symbol AZN. AstraZeneca was founded in 1999 and is headquartered in Cambridge, United Kingdom. AstraZeneca has a diverse product development pipeline concentrated generally in oncology, cardiovascular/renal/metabolism and respiratory/immunology. In addition to its COVID-19 vaccine, AZD1222, the company is in clinical trial with AZD7442 for the prevention and treatment of COVID-19. AZD7442 is a combination of two long-acting antibody (LAAB) types derived from convalescent patients after SARS-CoV-2 infection. AstraZeneca is also trialing MEDI3506 a monoclonal antibody (mAb) to IL-33 for treatment of COVID-19.

CanSino Biologics is a small publicly-traded company established in Tianjin, China in 2009 and dedicated to vaccine research, development and commercialization. CanSinoBIO has been listed on the Main Board of Hong Kong Exchange and is dual listed on the Shanghai Stock Exchange. In addition to the COVID-19 vaccine Ad5-nCOV, trade-named Convidecia, CanSinoBIO has a number of other vaccines in clinical trial and one for Ebola (Ad5-EBOV) which is only approved in China.

Moderna Therapeutics was founded in 2010, is headquartered in Cambridge (MA), and trades with the Nasdaq symbol MRNA. Besides mRNA-1273 in distribution, Moderna has mRNA-1647 vaccine in Phase II trial for Cytomegalovirus (CMV) and mRNA-1893 in Phase I trial for Zika virus. Within their oncology pipeline, Moderna has mRNA-4157 personalized cancer vaccine in combination Phase I/II trials for solid tumors and melanoma respectively. In their intratumoral immuno-oncology suite they have mRNA-2416 encoding OX40L for solid tumors/lymphoma and advanced ovarian carcinoma in Phase I. They also have mRNA-2752 encoding OX40L/IL-23/IL-36γ for solid tumors/lymphoma in Phase I. Moderna was also featured in the Immunotherapy and the Basis of Neoantigen Biology: Part I Neoantigens post from April 2019.

BioNTech was founded in 2008, is based in Mainz (Germany), and after IPO on Oct 19, 2019 trades on the NASDAQ under the symbol BNTX. Its founders are clinical scientists Prof. Dr. Ugur Sahin and Prof. Dr. Christoph Huber who are affiliated with the Gutenberg University. As well as BNT162 now being distributed widely, BioNTech has an mRNA-based Individualized Neoantigen Specific Immunotherapy (iNeST) termed BNT122 in Phase II trial for treatment of metastatic melanoma. The company also has a number of RNA vaccines to Fixed combination of shared cancer antigens (FixVac) in Phase I for various cancer indications. BioNTech was also featured in Immunotherapy and the Basis of Neoantigen Biology: Part II Cancer Immunotherapies May 2019.

References

Barrett et al., Phase 1/2 trial of SARS-CoV-2 vaccine ChAdOx1 nCoV-19 with a booster dose induces multifunctional antibody responses. (2021) Nat Med., Feb;27(2):279-288.

Bos et al., Ad26 vector-based COVID-19 vaccine encoding a prefusion-stabilized SARS-CoV-2 Spike immunogen induces potent humoral and cellular immune responses. (2020) NPJ Vaccines, Sep 28;5:91.

Corbett et al., SARS-CoV-2 mRNA vaccine design enabled by prototype pathogen preparedness. (2020) Nature, Oct;586(7830):567-571.

Dong et al., A systematic review of SARS-CoV-2 vaccine candidates. (2020) Signal Transduct Targer Ther., Oct 13;5(1):237.

Emary et al., Efficacy of ChAdOx1 nCoV-19 (AZD1222) vaccine against SARS-CoV-2 variant of concern 202012/01 (B.1.1.7): an exploratory analysis of a randomized controlled trial. (2021) Lancet, Apr 10;397(10282):1351-1362.

Ewer et al., T cell and antibody responses induced by a single dose of ChAdOx1 nCoV-19 (AZD1222) vaccine in a phase 1/2 clinical trial. (2021) Nat Med., Feb;27(2):270-278.

Forni et al., COVID-19 vaccines: where we stand and challenges ahead. (2021) Cell Death and Differentiaion, Feb;28(2):626-639.

Logunov et al., Safety and immunogenicity of an rAd26 and rAd5 vector-based heterologous prime-boost COVID-19 vaccine in two formulations: two open, non-randomised phase 1/2 studies from Russia. (2020) Lancet, Sep 26;396(10255):887-897.

Logunov et al., Safety and efficacy of an rAd26 and rAd5 vector-based heterologous prime-boost COVID-19 vaccine: an interim analysis of a randomised controlled phase 3 trial in Russia. (2021) Lancet, Feb 20;397(10275):671-681.

Madhi et al., Efficacy of the ChAdOx1 nCoV-19 Covid-19 Vaccine against the B.1.351 Variant. (2021) N Engl J Med., Mar 16. doi: 10.1056/NEJMoa2102214.

Rawat et al., COVID-19 vaccine: A recent update in pipeline vaccines, their design and development strategies. (2021) Eur J Pharmacol., Feb 5;892:173751.

Sadoff et al., Interim Results of a Phase 1-2a Trial of Ad26.COV2.S Covid-19 Vaccine. (2021) N Engl J Med., Oct 13;5(1):237.

van Doremalen et al., ChAdOx1 nCoV-19 vaccine prevents SARS-CoV-2 pneumonia in rhesus macaques. (2020) Nature, Oct;586(7830):578-582.

Voysey et al., Safety and efficacy of the ChAdOx1 nCoV-19 vaccine (AZD1222) against SARS-CoV-2: an interim analysis of four randomised controlled trials in Brazil, South Africa, and the UK. (2021) Lancet, Jan 9;397(10269):99-111.

Walsh et al., Safety and Immunogenicity of Two RNA-Based Covid-19 Vaccine Candidates. (2020) N Engl J Med., Oct 14;NEJMoa2027906.

Zhu 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. (2020) Lancet, Jun 13;395(10240):1845-1854.

Zhu et al., Immunogenicity and safety of a recombinant adenovirus type-5-vectored COVID-19 vaccine in healthy adults aged 18 years or older: a randomized, double-blind, placebo-controlled, phase 2 trial. (2020b) Lancet, Aug 15;396(10249):479-488.