BTE722 Vaccine Technology PDF

BTE722 Vaccine technology History of vaccine development- Importance of vaccines. Immunological response to vaccines Different types of vaccines: Inactivated toxins, Inactivated whole bacteria or viruses, Live attenuated bacteria or viruses; Subunit vaccines, Polysaccharide...

BTE722 Vaccine technology History of vaccine development- Importance of vaccines. Immunological response to vaccines Different types of vaccines: Inactivated toxins, Inactivated whole bacteria or viruses, Live attenuated bacteria or viruses; Subunit vaccines, Polysaccharide vaccines, Conjugated vaccines ; Recombinant DNA vaccines, Edible vaccines, Virus like particles; Next-generation vaccines: Human Immunome project; Human antibodies as vaccines. Vaccine design and development: Epitope identification; Vaccine efficacy, Adjuvants ; Delivery methods: microspheres, nanoparticles; ISCOMS and immunomodulators. Production techniques used for vaccines ; Storage and preservation of vaccines. Regulatory issues in vaccine production: OIE guidelines for production and seed lot management; Manufacturing recommendation; Final product release tests. Vaccine safety-the debate.  There are so many types of vaccines. Polio is being given in live attenuated and inactivated….Why ? Different vaccines can cause different adverse reactions? Contraindications in vaccination – additional risks? Immunizing pregnant women or immunocompromised people? Do we really need two different vaccines for SARS-CoV-2 ?  Generations and vaccines A fourth generation vaccine ? First generation :  LIVE ATTENUATED VACCINES (LAV) Examples: BCG; OPV; Measles; Rotavirus; Yellow fever LAVs Vaccine Safety Basics- WHO; https://vaccine-safety-training.org/ Adverse reactions of LAVs  Tuberculosis (BCG): Fatal dissemination of BCG infection; (0.19–1.56 per million) BCG osteitis BCG is cross reactive against some forms of leprosy and Buruli ulcer Image : Terreri, M., & Yamada, A.F. (2007). Osteitis caused by BCG vaccination. Pediatric Radiology, 38, 481. Oral polio vaccine: Vaccine-associated paralytic poliomyelitis (VAPP) (1 in 2.7 million) Vaccine derived poliovirus (VDPV) Circulating Vaccine derived poliovirus (cVDPV) (750 cases till date) Image: https://www.who.int/immunization/diseases/poliomyelitis Measles: Febrile seizures (mainly restricted to children under 5; not fatal in most cases; 0.3%) Image: https://www.stjohnvic.com.au/news/what-is-febrile-seizure/ Thrombocytopenic purpura (0.03%) Anaphylaxis (mostly due to vaccine Components like sorbitol) INACTIVATED WHOLE-CELL (KILLED ANTIGEN) VACCINES Examples: IPV; Whole-cell pertussis Vaccine Safety Basics- WHO; https://vaccine-safety-training.org/ Adverse reactions: The pertussis vaccine has been shown to lead to some cases of Hypotonic, hyporesponsive episodes (HHE) (approx 1 in 4,762) National Pulse Polio and India’s turnover story- India constituted over 60% of all global polio cases …. In 2014 it was declared polio free (27th March) 1970 – Sabin vaccine was developed in India “India committed to the resolution passed by World Health Assembly for global polio eradication in 1988. The country introduced polio vaccine under Expanded Programme on Immunization (EPI, 1978), and subsequently in Universal Immunization Programme (UIP, 1985), but started carrying out special polio campaigns from 1995.” Response to Polio Vaccination-IPV versus OPV 18  Subunit (purified) vaccines Subunit vaccines, like inactivated whole-cell vaccines, do not contain live components of the pathogen. They differ from inactivated whole-cell vaccines, by containing only the antigenic parts of the pathogen. These parts are necessary to elicit a protective immune response.  Immunologically relevant part ? Bacteria- LPS (gram negative) Virus- Capsid proteins; Parasite- polysaccharide, proteins Which is better ? Why ? Protein based subunit vaccines Present an antigen to the immune system, using a specific, isolated protein of the pathogen (expensive; immunological memory uncertain (?) , boosters required) Example: Hepatitits (hepatitis B surface antigen (HBsAg)), Acellcular pertussis Polysaccharide vaccines Polysacharide – any limitation ? Not be effective in infants and young children (under 18–24 months) Induce only short-term immunity (slow immune response, slow rise of antibody levels, no immune memory). Example: Pneumococcal vaccine; Meningococcal vaccine Conjugate vaccines Polysaccharide to a carrier protein Example: Pneumococcal vaccine (PCV 7); Haemophilus influenzae b Costly ? Stable ? Vaccine Safety Basics- WHO; https://vaccine-safety-training.org/  COMBINATION VACCINES Combination vaccines consist of two or more antigens in the same preparation. This approach has been used for over 50 years in many vaccines such as DTwP and MMR. Testing ? Too many antigens overwhelming the immune system? Adjuvants, delivery system, other components compatibility ? ✔Reducing the cost of stocking and administering separate vaccines, ✔Reducing the cost of extra health care visits, ✔Improving timeliness of vaccination (some parents and health-care providers object to administering more than two or three injectable vaccines during a single visit because of a child's fear of needles and pain, and because of concerns regarding safety), ✔Facilitating the addition of new vaccines into immunization programmes Vaccine Safety Basics- WHO; https://vaccine-safety-training.org/ MMRV ?  Toxoids (inactivated toxins) Examples: Tetanus; Diptheria The protein-based toxin is rendered harmless (toxoid) Inactivated or killed toxin (poison) used in vaccine production Vaccine Safety Basics- WHO; https://vaccine-safety-training.org/  Adverse effects Very less- known adverse effects of Tetanus is Anaphylaxis Sometimes swelling due to improper injection module What generation does it belong to ? How do you make them ? Inactivated: formalin; β-propiolactone; heat; radiation Attenuation: pathogenic organism different host; cold stress; salt, bile etc Toxoid: formalin Subunit : blood derived; heat killed The story of the 1st hepatitis vaccine Baruch Bloomberg and the Australian antigen Second generation vaccines :  How do you make Recombinant subunit vaccines ? Recombinant vaccines are products of genetic engineering, where typically a bacterial or yeast cells are programmed to produce antigens of harmful pathogens. A small piece of DNA is taken from the virus or bacterium against which we want to protect. This is inserted into other cells to make them produce large quantities of active ingredient for the vaccine (usually just a single protein or sugar). HPV vaccine; MenB vaccine.  Hepatitis : Ref to : Bitter, Ph.D., Grant A. & Egan, Kevin & Burnette, W. Neal & Samal, Babru & Fieschko, John & Peterson, Darrel & Downing, Michael & Wypych, Jette & Langley, Keith. (1988). Hepatitis B vaccine produced in yeast. Journal of medical virology. 25. 123-40. 10.1002/jmv.1890250202. Alternative host : Hansenula polymorpha and Pichia pastoris  When using peptides: Use discrete portion (domain) of a surface protein as Vaccine These domains are ‘epitopes’- antigenic determinants are recognized by antibodies Often accompanied by carriers Limited success? Image: https://www.biologydiscussion.com/biotechnology/vaccines/types-of-recombinant-vaccines-3-t ypes/10080  Herpes Simplex Virus: Image: https://www.biologydiscussion.com/biotechnology/vaccines/types-of-recombinant-vaccines- 3-types/10080  Human Immunodeficiency virus: Image: https://www.biologydiscussion.com/biotechnology/vaccines/types-of-recombinant-vacci nes-3-types/10080 Current status HIV vaccine- where are we now ? Non-clinical vaccines- in different stages of work  Edible vaccines Edible vaccines (Charles Arnetz) refers to any foods; typically plants, that produce vitamins, proteins or other nourishment that act as a vaccine against a certain disease. Edible vaccines are currently being developed for measles, cholera, foot and mouth disease, Hepatitis B and Hepatitis C Edible Vaccine involves introduction of selected desired genes into plant and then inducing these altered plants to manufacture the altered protein. 39 Why the need for edible vaccines? How do you prepare it ? WILLIAM H. R. LANGRIDGE, Edible Vaccine, Copyright 2000 Scientific American, Inc Some different methods Birtukan Girma, Dereje Shegu, Ayelech Muluneh and Fanos Tadesse woldemariyam. (2019). Vaccine Production in Transgenic Plants for Animal and Human Diseases. Archives of Veterinary and Animal Sciences 1(1).  Other questions What are the factors considered before selecting a gene? Is there a selective advantage of one plant versus other ? Selection of promoter? Gene expression and integration system? Ethical issues and regulations? Other systems Algae, insects ?  Mode of action: Antigen in transgenic plant - Ingestion -Delivered by bioencapsulation -Taken up by Mcell - Pass on to the Macrophage - IgG, IgE responses - Local IgA response & Memorycells - Neutralize the attack by the real infectious agent. WILLIAM H. R. LANGRIDGE, Edible Vaccine, Copyright 2000 Scientific American, Inc  Pros ✔ Cost effective. ✔ Easy to administer. ✔ Easy to store. ✔ Acceptable to poor/developing country. ✔ Fail safe ✔ Activate both mucosal and systemic immunity. ✔ Heat stable ? ✔ Do not required cold chain maintenance. ✔ No fear of contamination. ✔ Easy to propagate Cons ▪ Transgenic contamination can occur. ▪ Antibiotic resistance marker genes can spread from GM food to pathogenic Bacteria. ▪ Difficulty in dose maintenance. ▪ Denaturation during cooking ? ▪ Difference in glycosylation ▪ Effect of stomach 46 Where are we now? Transgenic bananas with anti-hemagglutination specific antibodies to combat measles. Transgenic tomatoes to treat Alzheimer's,plague, ( Read: Youm, J. W., Jeon, J. H., Kim, H., Kim, Y. H., Ko, K., Joung, H., & Kim, H. (2008). Transgenic tomatoes expressing human beta-amyloid for use as a vaccine against Alzheimer's disease. Biotechnology letters, 30(10), 1839–1845. https://doi.org/10.1007/s10529-008-9759-5) https://doi.org/10.1186/s43088-023-00453-x Relevance of Edible vaccines  Virus-like particle vaccines Virus-like particles (VLPs) are multiprotein structures that mimic the organization and conformation of authentic native viruses but lack the viral genome, potentially yielding safer and cheaper vaccine candidates. This consist of viral proteins derived from the structural proteins of a virus. These proteins can self-assemble into particles that resemble the virus from which they were derived but lack viral nucleic acid, meaning that they are not infectious. Because of their highly repetitive, multivalent structure, virus-like particles are typically more immunogenic than subunit vaccines. The human papillomavirus (HPV) and Hepatitis C virus vaccines are two virus-like particle-based vaccines currently in clinical use. They are non-infectious because they contain no viral genetic material. They can be naturally occurring or synthesized through the individual expression of viral structural proteins, which can then self assemble into the virus-like structure 50  Negative staining electron microscopy Image: https://thenativeantigencompany.com/products/mayaro-virus-vlp/ Image : (2008). 1. Human papillomavirus (HPV) infection 1.1 Structure and molecular biology of human papilomaviruses. Schiller JT, Müller M. Next generation prophylactic human papillomavirus vaccines. Lancet Oncol. 2015 May;16(5):e217-25. doi: 10.1016/S1470-2045(14)71179-9. PMID: 25943066. https://www.slideshare.net/gauravg/cervical-cancer-vaccine-do-we-need-it-in-india https://www.paho.org/spanish/ad/fch/im/31_Andrus_HPVvaccines_Feb2008.pdf  Expression systems for the production of VLPs and these include: (1) various mammalian cell lines, either transiently or stably transfected or transduced with viral expression vectors; (2) the baculovirus/insect cell system (convenience, speed, high yield of expressed proteins; problem : simpler N-glycosylation pattern) (3) various species of yeast including Saccharomyces cerevisiae and Pichia pastoris( rapid cell growth, high yield of expression proteins, scalability, cost-effective production, and providing a degree of PTM processes ) (4) Escherichia coli and other bacteria. (lack of PTM system, incomplete disulfide bond formation and protein solubility problems,-not suitable platforms for producing enveloped VLPs) (5) Plants Great at producing neutralizing antibodies Caldeira, Jerri & Perrine, Michael & Pericle, Federica & Cavallo, Federica. (2020). Virus-Like Particles as an Immunogenic Platform for Cancer Vaccines. Viruses. 12. 488. Prophylactic HPV L1 VLP vaccines Delivered intramuscularly rapid access of VLPs to blood vessels and local lymph nodes potent activators of APC induce good T helper responses for B cells Efficacy >90% for persistent infection 100% for disease (5 years post vaccination) in subjects naïve for vaccine HPV types Immunogenic high antibody concentrations up to 1000x > than in natural HPV infection Duration of vaccine induced antibody levels protection maintained over 5 years Safe no vaccine related serious adverse events identified in the trials to date (70,000 women) Antibody response Mosquirix – the VLP The mosquito life cycle and malaria Viruses Different viruses, different VLPs Relevance of VLP What factors affect self aggregation? Ph, temperature, molar ratio, glycosylation ?, lysis, shear force.. As a Delivery tool Virus-Like Particle; Progress in Molecular Biology and Translational Science  Bacterial ghost Empty cell envelopes of Gram negative bacteria Tested in animals (for veterinary) Vaccines for V.cholerae on animal testing ✔Simple production, packaging ✔No cold chain ✔Long storage- freeze drying possible ✔Both cellular and humoral immunity ✔Natural adjuvant ✔Excellent carrier ✔Mucosal immunity ✔Mostly used as delivery tools How is bacterial ghost made ? How to make bacterial ghosts ? What generation ? Bacterial ghosts as delivery systems.. https://microbenotes.com/vaccines-introduction-and-types/  Third generation vaccines “For over a hundred years vaccination has been effected by one of two approaches: either introducing specific antigens against which the immune system reacts directly; or introducing live attenuated infectious agents that replicate within the host without causing disease synthesize the antigens that subsequently prime the immune system” Nucleic acids as vaccines who.int Nucleic acid vaccines DNA Vaccine- West Nile Virus Coronavirus  Types DNA RNA Vector based Viral – Replicating ; Non replicating Non-vector based DNA Vaccines It involves the direct introduction into appropriate tissues of a plasmid containing the DNA sequence encoding the antigen(s) against which an immune response is sought, and relies on the in situ production of the target antigen. Alternatively, the DNA may be encapsulated in protein to facilitate cell entry. Instead of the DNA, the cDNA may be used What should the plasmid contain ??? The plasmid carrying DNA vaccine normally contains a promoter site, cloning site for the DNA vaccine gene, origin of replication, a selectable marker sequence (e.g. a gene for ampicillin resistance) and a terminator sequence (a poly—A tail) Different modes of delivery can be used – injection (intramascular, intravenous, intradermal); gene gun; mucosal delivery and topical administration (direct or liposome mediated)Interestingly, skin and mucus membranes possibly best sites … ??? Immune response in DNA vaccine  Current status of DNA Vaccine Data from preclinical studies great – protective immunity induced. The initial development of DNA vaccines in larger animals and human studies showed that DNA is well tolerated and has an excellent safety record. Clinical studies of first-generation vaccines, primarily consisting of naked DNA, showed that this platform induces only low levels of immunity. Recent studies have generated new leads from basic research on insert design, RNA structure, variation in codon usage, and leader-sequence optimizations — all of which improve the immune potency of DNA vaccines. New formulations, including lipids and polymers, and new delivery devices. Strong molecular adjuvants that are included in plasmid formulations seem to be important Multiple vaccines in preparation Cancer vaccine- DNA Status of DNA vaccine in SARS-CoV-2 In non-infectious model Advantages: Antigen presentation by both Class I and Class II- stimulation of both B- and T-cell responses improved vaccine stability, the absence of any infectious agent – no risk of infection the relative ease of large-scale manufacture. Stability of storage and shipping No need of costly production of proteins hence Cheap In-vivo expression Disadvantages: Limited to protein immunogens (not useful for non-protein based antigens such as bacterial polysaccharides) Possibility of inducing antibody production against DNA Possibility of tolerance to the antigen (protein) produced Potential for atypical processing of bacterial and parasite proteins Potential when using nasal spray administration of plasmid DNA nanoparticles to transfect non-target cells Important considerations for a plasmid vector: Highly active expression vectors Strong viral promoter Including Intron A sequence Strong polyadenylation/transcriptional termination signal Polycistronic vectors for simultaneous expression of two proteins Codon usage optimization Preferential stimulation of B/T cells High in GC Avoid chi sites  RNA Vaccines Use RNA containing vector The RNA sequence codes for antigens/proteins that are identical or resembling those of the pathogen Delivery can be naked particles; lipid nanoparticles as well as ex-vivo Ex-vivo: Dendritic cells are extracted from the patient’s blood, transfected with the RNA vaccine, then given back to the patient to stimulate an immune reaction. Conventional mRNA vaccines include in their simplest an ORF for the target antigen, flanked by untranslated regions (UTRs) and with a terminal poly(A) tail. Working of a mRNA Vaccine Immune response of a mRNA Vaccine Suitability of mRNA Vaccines Strategies for optimizing mRNA Vaccine  Are RNA Vaccines better than the DNA counterpart? ✔ No risk of host genome integration- cytosol ✔ Modified nucleosides can be incorporated to avoid immune response ✔ The ORF and UTR of mRNA can be optimized to amplify/increase translation Unintended immune reaction. Some mRNA-based vaccine platforms induce potent type I interferon responses- inflammation and autoimmunity Image: Zifu, Zhong & Mc Cafferty, Sean & Combes, Francis & Huysmans, Hanne & De Temmerman, Joyca & Gitsels, Arlieke & Vanrompay, Daisy & Portela Catani, João & Sanders, Niek. (2018). mRNA therapeutics deliver a hopeful message.. The pioneers of mRNA vaccine  History of mRNA vaccines Discovered in 1960s 1970s they could be delivered into cells First mice testing 1990 First clinical testing 2013 Katalin Karikó; Michael Buckstein; Houping Ni; Drew Weissman (August 2005). "Suppression of RNA recognition by Toll-like receptors: the impact of nucleoside modification and the evolutionary origin of RNA". Immunity. 23 (2): 165–75. doi:10.1016/J.IMMUNI.2005.06.008. ISSN 1074-7613. PMID 16111635. Wikidata Q24316383. Bart R. Anderson; Hiromi Muramatsu; Subba R Nallagatla; Philip C. Bevilacqua; Lauren H. Sansing; Drew Weissman; Katalin Karikó (10 May 2010). "Incorporation of pseudouridine into mRNA enhances translation by diminishing PKR activation". Nucleic Acids Research. 38 (17): 5884–5892. Clinical trials of mRNA vaccines against infectious diseases  Vector Vaccines: Virus as Antigen Gene Delivery System Antigen Gene A live vector vaccine using viruses is a vaccine that uses a chemically weakened virus to transport pieces of the Virus pathogen in order to stimulate an immune response. Patient Antigen Protein is Made  Usually either replication-competent or replication-defective. Replication-defective vectors are the most common choice in studies because the viruses have had the coding regions for the genes necessary for additional rounds of virion replication and packaging replaced with other genes, or deleted. These virus are capable of infecting their target cells and delivering their viral payload, but then fail to continue the typical lytic pathway that leads to cell lysis and death. Conversely, replication-competent viral vectors contain all necessary genes for virion synthesis, and continue to propagate themselves once infection occurs. Because the viral genome for these vectors is much lengthier, the length of the actual inserted gene of interest is limited compared to the possible length of the insert for replication-defective vectors. Which is better?  History of viral vector vaccines – before COVID 1st vaccination schemes has been investigated using vaccinia virus in 1984. Human clinical trials were conducted for viral vector vaccines against several infectious diseases including Zika virus, influenza viruses, respiratory syncytial virus, HIV, malaria. Two Ebola vaccines that used viral vector technology were used to combat Ebola outbreaks between 2013-2020 ------ The rVSV-ZEBOV vaccine was approved for medical use in the European Union in November. Zabdeno/Mvabea was approved for medical use in the European Union in July 2020.  Pros and Cons ✔ The advantage with vector vaccine is that it stimulates B-lymphocytes (to produce antibodies) and T-lymphocytes (to kill virus infected cells). (Remember subunit vaccines trigger the humoral axis) ✔ 2. Often no adjuvant needed ✔ 3. High efficient gene transduction ✔ 4. Specific delivery ✔ 5. Possibility of vaccinating individuals against different diseases simultaneously. ✔ 6. Possibility of large scale manufacturing Concerns: ?? Safety Toxicity Difficult to ptoduce? Stability Using viruses as delivery vehicle- Features? Step 1: Construction of the vector- Generation Can you make it replication deficient or replication competent ? Helper required ? How large can the construct be ? Step 2: Getting inside Can it infect replicating or non-replicating cells ? How efficient ? Does it integrate ? If so where? Replicated during cell division? In-vivo, ex-vivo ? Step 3: Expression Rate of transgene production? Type of cells? Duration ? Step 4: Immunity: Immunologically relevant? Pre-existing immunity ? Safety ?  What kind of viruses can we use? Non (or low) pathogenic Maybe genetically engineered to reduce/eliminate any remaining pathogenicity Ideal scenario Non toxic; high transduction; stable; safe to handle; high packaging capacity Safety Toxicity Stability Cell type specificity- Tropism Identification  Viruses as backbone Adenovirus: High immunogenicity; High transduction rate, high transgene expression High titter production; can infect both replicating and non-replicating ; large transgene; no integration Pre existing immunity Adeno-associated virus : Integration in nature happens (but specific); non-pathogenic; long term gene expression; can infect non dividing cells ; in dividing cells lost; low immunogenicity; low cytotoxic response very low titer Poxvirus/Vaccinia virus : Licensed: Zabdeno/Mvabea Ebola vaccine Can make both replication competent and deficient strain; High titre production; But preexitsing immunity Retrovirus: can stably integrate their genetic information into the host cell chromosome; target cell specificity; hybrid vectors comprising different vector designs and incorporation of elements from various viruses is feasible; random integration so potential tumorogenic; infects diving cells only Gammaretroviruses; lentiviruses and spumaviruses mostly used Sendai virus: pediatric; high immunogenicity; pre existing immunity Plant viruses Plant viruses can be used to engineer viral vectors, tools commonly used to deliver genetic material into plant cells; they are also sources of biomaterials and nanotechnology devices. Tobacco mosaic virus (TMV) is the first virus to be discovered.. Hybrids Hybrid vectors are vector viruses that are genetically engineered to have qualities of more than one vector. Viruses are altered to avoid the shortcomings of typical viral vectors, which may have limited loading capacity, immunogenicity, genotoxicity, and fail to support long-term adequate transgenic expression A quick comparison Adenovirus as vector vaccine construct Approx 36kb Different generations of adenoviral vectors Methods of preparation of adenoviral vector vaccines https://pubmed.ncbi.nlm.nih.gov/24279313/ Structure of lentivirus  Three major classes of components Gene Products Function gag Capsid Protects the core Matrix Lines envelope Nuclear capsid Protects the genome and forms the core pol Protease Essential for Gag protein cleavage during maturation Reverse transcriptase Reverse transcribes the RNA genome into double stranded DNA Integrase Required for integration of the provirus env Surface glycoprotein Outer envelope glycoprotein; major virus antigen Transmembrane protein Inner component of the mature envelope glycoprotein Lentiviral vectors: Lentiviral vectors are non-replicative, negligibly inflammatory, and not targets of preexisting immunity in human populations. The potential of lentiviral vectors to transduce non-dividing cells is determinant in their strong immunogenicity. Can infect non dividing cells. The viral genome in the form of RNA is reverse-transcribed when the virus enters the cell, which is then inserted into the genome at a random position by viral integrase. The provirus is passed on to the progeny By Peter Znamenskiy - Own work, Public Domain, https://commons.wikimedia.org/w/index.php?curid=912852 Components of a current generation Lentiviral plasmid vector 1. Lentiviral transfer plasmid encoding your insert of interest. The transgene sequence is flanked by long terminal repeat (LTR) sequences, which facilitate integration of the transfer plasmid sequences into the host genome. Typically it is the sequences between and including the LTRs that is integrated into the host genome upon viral transduction. Many lentiviral transfer plasmids are based on the HIV-1 virus. Transfer plasmids are all replication incompetent and may contain an additional deletion in the 3'LTR, rendering the virus “self-inactivating” (SIN) after integration. 2. Packaging plasmid(s) 3. Envelope plasmid Why three or four plasmids ?  Two Viral vector based vaccines  Antibodies as a vaccine ? Monoclonal versus polyclonal ? Advantages and disadvantages 4TH generation? How do antibodies work? Human antibodies versus mouse antibodies Vaccination or Immunization ?  Fourth generation vaccines ? Chiappelli F. 2019-nCoV - Towards a 4th generation vaccine. Bioinformation. 2020;16(2):139-144. Published 2020 Feb 12. doi:10.6026/97320630016139  What if we inject a soluble binary molecule (i.e., ACE2R-ACE2R; [ACE2R] 2) or its quaternary form (i.e. two intertwined ACE2R-ACE2R; [ACE2R] 4 ? https://nat.au.dk/om-fakultetet/nyheder/nyhed/artikel/38-mio-kr-til-nyt-samarbejde-om-bekaempelse-af-covid-19/ Graphics: Daniel Otzen Case study : SARS-CoV-2 and vaccine development https://covid19.trackvaccines.org/ https://www.thno.org/v10p7821.htm https://www.invivogen.com/spotlight-covid-19-vaccine-development Immunologically relevant molecule? Antivirals versus vaccine Case study 2: Mtb and vaccine development  Candidates? 1. Antigen 85 https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7786643/ Other components in vaccine ?  Extra resources https://www.youtube.com/watch?v=XYtV034mQvQ https://www.youtube.com/watch?v=hEXkEuoA3UE https://www.youtube.com/watch?v=hYHbfQe5h-Q https://www.youtube.com/watch?v=GyCO9c2PbtU