Vaccines based on modern RNA technology

Two Covid-19 vaccines have been approved for emergency use by the FDA in the USA and one by the Medicines and Healthcare products Regulatory Agency (MHRA) in the U.K. Both of them are based on mRNA technology (BNT162b2 by Pfizer/BioNTech and mRNA-1273 by Moderna/Lonza). They contain a self-replicating mRNA that encodes the entire spike (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 body¹. Both require a booster shot, 30 days after the first one. The mRNA in both vaccines has a modified nucleic acid (1-methyl-pseudouridine) that increases the translation of mRNA into the S protein and helps avoid harmful overactivation of the innate immune system. Both vaccines are encapsulated with lipid nanoparticles and were over 90% effective in Phase 2/3 clinical trials. The Moderna vaccine can be stored at -20 oC 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 oC. Both vaccines are already being mass-produced. Hopefully, as many as 40 million doses (enough for 20 million people) will be ready for distribution by the end of this month. Priority is being given to the most vulnerable people (elderly and healthcare workers). In addition, vaccine candidates that are based on different technologies are being shown to be safe and effective – even in people over 70. Fortunately, there have been no cases of antibody-dependent enhancement (ADE) of viral infectivity or severity of symptoms in any of these clinical trials. Also, ADE was not seen in a recently approved monoclonal antibody, bamlanivimab². It was approved by the FDA for treating Covid-19.

Other important vaccines use an attenuated cold virus (adenovirus) to produce the S protein that activates the desired immune response. The Chinese have one (CoronaVac) that has been given to members of the Chinese military and others for months^3,4. It was shown to be safe and effective in a recent clinical trial⁴. Another (Sputnik V) is being given in Russia to some of its military and the most exposed groups (doctors, healthcare workers, teachers, and social workers^5,6. Millions of doses are now available. Adenovirus vectors are pre-eminent in inducing a strong T-cell response. They have been used to produce three different Ebola vaccines that were given to over 60,000 people, as well as two anticancer drugs for over 30,000 people. The Sputnik V vaccine is called Gam-Covid-Vac Lyo in ongoing clinical trials in the USA⁷. Two trials are being conducted. In one of them, people will be given either placebo or one dose. In the other trial, they will be given either placebo or two doses.

Two other vaccine candidates using adenovirus vectors were developed by Oxford and AstraZeneca (ChAdOx1 nCoV-19 and AZD1222)⁸ and Johnson & Johnson (JNJ-78436735)⁹. Moreover, validated procedures already exist for manufacturing billions of doses of this vaccine safely and inexpensively¹⁰. Oxford’s partner, AstraZeneca, is ready to produce 3 billion doses of their vaccine in 2021. The AZD1222 vaccine candidate was found to be 90% effective when given as a half dose, followed by a full dose at least six months later¹¹. It was 62% effective when given as two full doses. It appears to be better tolerated in older adults than in younger adults and has similar immunogenicity in all age groups (18-55 years, 56-69 years, and ≥70 years) after the second dose. One important limitation is that nobody in the study had substantial chronic illnesses or frailities¹².

So, several types of vaccines against Covid-19 are becoming widely available. They may be much more effective than the seasonal flu vaccines¹³. The first vaccines that are becoming widely available in the USA use modern mRNA technology. This technology has much potential not just as vaccines, but as cures for other diseases. Even though vaccines are usually thought of as preventions, they can also cure diseases. A vaccine is something that stimulates a person’s immune system to produce immunity to a specific disease, protecting the person from that disease. For example, tumors produce specific antigens, much like viruses and pathogenic organisms. Many potential cancer vaccines would stimulate a person’s immune system, causing it to target the tumor antigen (TA). TAs are important in tumor initiation, progression, and metastasis¹⁴. They can include tumor-associated antigens (TAAs) and tumor-specific antigens (TSAs). So, mRNA vaccines would deliver genetic information encoding TAs. Moreover, mRNA collected from a patients’ tumor samples can be used to find and target patient-specific TAs¹⁵. The goal is to identify every unique somatic mutation in an individual patient’s tumor sample¹⁶. Then neoepitope cancer vaccines can be designed rationally. Therapeutic cancer vaccines should stimulate cell-mediated immune responses that are capable of clearing or reducing the size and metastasis of tumors. So, mRNA vaccines could potentially be used on infectious diseases and cancer. Unlike vaccines based on genetically engineered viruses, mRNA is not infectious. There is no risk of it becoming mutated. Antiviral immunity is not a problem, so it can be given repeatedly. Finally, millions of mRNA vaccines can be produced rapidly and inexpensively. So, researchers are developing mRNA vaccines that may cure many types of cancer. They can be inserted into dendritic cells (DCs) that have been isolated from a patient’s blood. That is, DCs are key activators of a proper immune response. They link the innate and acquired immune systems, causing responses against TAs. There are several clinical trials of mRNA vaccines against a variety of tumors¹⁶.

Some cancer vaccines use mRNAs that encode a patient’s own specific TAs to activate their immature DCs¹⁷. The DCs are then put back in the patient, thus initiating protective immune responses. With the help of computer-aided design, new, more potent TAs will be discovered and optimum mRNAs will be prepared. That is, the mRNA will not only encode a TA, but also attach untranslated regions (UTRs) to the 5’ and 3’ ends. This maximizes the translation of the mRNA into the desired TA.

One TA is becoming the major target for the design and development of cancer vaccines. It's aberrantly glycosylated mucin-1 (MUC-1)¹⁸. It is a transmembrane glycoprotein that is on the surface of almost all epithelial cells. It’s on the apical surface of most glandular epithelial cells, including those of the mammary gland, lung, pancreas, kidney, female reproductive tract, and stomach. Aberrantly glycosylated MUC1 is associated with human cancers. Properly glycosylated MUC1 helps form a physical barrier that lubricates and protects normal epithelial tissues and mediates intercellular communication. However, aberrant expression of MUC1 occurs in esophageal, gastric, breast, ovarian and bladder cancer. It is especially prominent in breast cancer cells. Aberrantly glycosylated MUC1 is a well known TSA on epithelial cell tumors¹⁸. It is a target in adoptive immunotherapy for treating pancreatic cancer¹⁹.

Between 2007 and 2012, 42 patients with unresectable or recurrent pancreatic cancer were treated at the Department of Digestive Surgery and Surgical Oncology (Department of Surgery II) of the Yamaguchi University Graduate School of Medicine¹⁹. This therapy was not a clinical trial, but a medical treatment approved as advanced health care by the Japanese Ministry of Health, Labor and Welfare. Pancreatic adenocarcinoma is the fourth leading cause of cancer death worldwide and has an overall 5-year survival rate of only 6%. No adequate therapy for pancreatic cancer has yet been found, and most patients die within a year of diagnosis. Immunotherapy has an advantage over radiation and chemotherapies. It acts specifically against the tumor without damaging normal tissue. Mucin 1 (MUC1) is overexpressed in an incompletely glycosylated form in pancreatic cancer. Cytotoxic T lymphocytes (CTLs) recognize MUC1 molecules. They can be used for all cancer patients that express the MUC1 antigen. DCs are potent antigen-presenting cells for inducing immune responses. Gemcitabine (GEM), which is a standard chemotherapeutic agent for pancreatic cancer may enhance responses to some vaccines. Treatment with GEM sensitizes human pancreatic carcinoma cell lines against CTL-mediated destruction. To create a more effective therapy for pancreatic cancer, mature DCs were transfected with MUC1-mRNA (MUC1-CTLs). These CTLs were induced by co-culture with a human pancreatic cancer cell line, and then with interleukin-2. Patients were treated with GEM, while MUC1-DCs were injected intradermally, and MUC1-CTLs were administered intravenously. The median survival time was 13.9 months. The 1-year survival rate was 51.1%. Of 42 patients, one patient had complete response (2.4%), three patients had partial response (7.1%) and 22 patients had stable disease (52.4%)¹⁹.

Progress is being made in treating lung cancer, too. CureVac AG, in Germany, tested its mRNA, called CV9201²⁰. It is based on RNActive® technology. Its mRNA encodes five antigens that are specific to non-small cell lung cancer (NSCLC). The tumor antigens that were selected were based on their roles in causing NSCLC, their emergence in malignant cells and their ability to induce the production of a proper immune response, the production of CTLs and/or antigen-specific antibodies. In a phase I/IIa clinical trial, patients with NSCLC received five intradermal injections of CV9201 (400–1600 μg of mRNA). It was safe and helped some of the patients live longer than expected. The 2- and 3-year survival rates were 26.7% and 20.7%, respectively. The results should lead to further clinical investigation²⁰.

There is another exciting antiviral strategy that is based on RNA that is produced using CRISPR technology. CRISPR is a naturally-occurring defense mechanism that bacteria use to keep them from being infected by viruses²¹. To do this, bacteria use the parts of their genomes that contain base sequences that are repeated many times, with unique sequences in between the repeats. They are called “clustered regularly interspaced short palindromic repeats” or CRISPR. They keep pieces of viral genomes in the bacterial DNA so they can recognize viruses and defend themselves against future infections. Scientists and engineers have learned to use the CRISPR to edit genes (DNA) and make new types of RNA, called CRISPR RNA, or crRNA. CRISPR is being used to improve livestock and seafood production, create better animal models of diseases, help in the development of improved vaccines and new prescription drugs, and possibly eventually eradicate malaria. It can also be used in the field of synthetic biology to make entirely new organisms and may be able to bring back extinct species, such as the wooly mammoth. CRISPR has been predicted to become one of the key technologies that will be part of the fourth industrial revolution²¹.

So, a new CRISPR-based antiviral strategy called PAC-MAN (Prophylactic Antiviral CRISPR in huMAN cells) targets the mRNA of the SARS-CoV-2 virus²². It was used to make CRISPR RNAs (crRNAs) that targeted either the H1N1 influenza virus or over 90% of 1087 recently sequenced coronaviruses²². PAC-MAN technology was combined with bioinformatics to prepare a possible treatment for Covid-19²³. First, they identified conserved regions in SARS-CoV-2 RNA. Then, they designed several crRNAs that targeted the different coronaviruses that have been identified and sequenced to date. One key advantage of this technology is that it can be adapted to treat multiple coronaviruses. It could use a single cocktail containing different crRNAs that target conserved regions in coronaviruses. Their computational analysis predicted that just three crRNAs could be enough to target the coronaviruses that cause SARS, MERS, and Covid-19. By using several crRNAs, mutants that emerge could be easily targeted. PAC-MAN technology may even be applied to other viruses that infect animals like bats that pose a future threat. PAC-MAN could be used to prepare potential treatments before a pandemic can occur²².

One key question about any vaccine for Covid-19 is how long it will be effective. If it’s like the flu (influenza) vaccine, it will only be effective for a year. Immunity decreases over time. The same thing could happen with Covid-19. Even though it seems to be rare, some people are getting Covid-19 twice²⁴. Even though there were reports of suspected re-infections as early as April, the first confirmed case was reported on 24 August. Since then, at least 24 other re-infections have been confirmed, but that is an underestimate. Still, that is a very small fraction of the nearly 70 million cases that have occurred worldwide. For a re-infection to be confirmed, a patient must have tested positive for the SARS-CoV-2 virus twice with at least one symptom-free month in between. However, a second test can also be positive simply because the patient has a residue of non-replicating viral RNA from their original infection. That is, the virus can linger in the gut, sometimes causing diarrhea, but not appearing in the nasal cavity during the second test. So, it’s important to emphasize that asymptomatic people can have the virus and spread it. In fact, asymptomatic people are major spreaders of Covid-19. So, most scientific journals want to see two full virus sequences (from the first and second illnesses) that are sufficiently different. So far, no evidence exists that mutations have emerged that would make the virus more pathogenic or that might help the virus evade immunity. SARS-CoV-2 can persist for months inside the gut. Persistent infections may help explain the extraordinarily long-lasting symptoms that afflict some Covid-19 survivors²⁴.

Political, economic and social factors

Vaccines alone will not end the Covid-19 pandemic in the USA, even if they are safe, effective and widely available. Covid-19 is not a simple infectious disease. It moves through our society easily^25,26. Covid-19 clusters with pre-existing conditions, such as hypertension, diabetes, cardiovascular disease, respiratory disorders and obesity. This is driven by systemic racism, mistrust in science lies from our leadership and a fragmented healthcare system. Essential, but underpaid workers include many African-Americans, Asians, immigrants and minority ethnic communities. Some of them are deemed essential because they work in slaughterhouses and meatpacking plants. At the same time, an economic crisis has emerged. We must reverse profound disparities in economic opportunities, access to healthy food and a good education. In contrast, the political leadership and communal responsibility of the people of New Zealand have been exemplary. Covid-19 does not have to be such a large problem, as it is in most rich countries^25,26.

Glossary of Terms

ACE-2 receptor: angiotensin-converting enzyme-2 that reduces blood pressure. It’s a receptor that’s bound to human cell membranes. The SARS-CoV-2 virus that causes Covid-19 infects human cells by binding to ACE-2 receptors.
Antibodies: proteins made by immune cells that bind to antigens from a pathogen (like the spike protein in the SARS-CoV-2 virus).
Antigens: parts of a pathogen (like the spike protein in the SARS-CoV-2 virus) that induces the production of antibodies in a host cell.
Epitope: the part of an antigen that binds to an antibody, thus activating the immune system. It’s also known as an antigenic determinant.
mRNA: messenger RNA, which is translated into proteins.
Neoepitope: A new epitope that appears on the surface of a cancer cell.
RNA: ribonucleic acid (many types exist, such as mRNA, tRNA, rRNA, lncRNA, microRNA, piRNA, pri-RNA, pre-miRNA, miRNA, shRNA, siRNA, snoRNA).
Polymerase chain reaction (PCR): A way to amplify (make many copies) of DNA, using a chain reaction that’s catalyzed by a DNA polymerase.
T-cells: immune cells made in the thymus gland. In the process of developing immunity, the body retains a few T-cells that are called memory cells. When the same person is exposed to the same infectious agent again later in life, the memory T-cells identify it and stimulate B-cells to produce antibodies to attack and eliminate the virus or other infectious agent.
Transcription: the process of transcribing a piece of DNA into a molecule of mRNA.