17. New approaches to viral vaccines

Question 1

How are the main vaccine platforms classified?

Answer 1

1. 1st generation
2. 2nd generation
3. 3rd generation

Question 2

What are 1st generation vaccine platforms?

Answer 2

1. Whole virus vaccines
2. These are either inactivated or attenuated
3. This is the oldest vaccine platform.

Question 3

What are 2nd generation vaccine platforms?

Answer 3

1. Protein based vaccines
2. Subunit vaccines
3. Replicating and non-replicating viral vector vaccines.
4. These use only a piece of the pathogen and use vectors to deliver genetic material.

Question 4

What are 3rd generation vaccine platforms?

Answer 4

1. Nucleic acid vaccines either DNA or mRNA.
2. Nanoparticle based vaccines

Question 5

How do whole virus vaccines work?

Answer 5

1. You take the whole pathogen and alter it so it cannot cause disease.
2. You can inactivate the pathogen chemically or using heat.
3. You can attenuate the pathogen in the lab by introducing deletions or modifications in essential genes.
4. These altered whole virus particles are then inject into patients and trigger an immune response.

Question 6

What vaccine platform was the first point of call for traditional vaccine design?

Answer 6

1. Whole virus vaccines
2. It is intuitive and produces really good immune responses.

Question 7

What are the problems with live attenuated vaccines?

Answer 7

1. Attenuated vaccines don’t replicate well in the host but in immunocompromised patients the vaccines could replicate well enough to cause disease.
2. The whole pathogen has good immunogenic epitopes but it also has virulence factors or epitopes that are poorly immunogenic. You want to ensure the desired epitopes are being recognised.

Question 8

How do viral vector vaccines work?

Answer 8

1. They use a modified virus to deliver genetic material from other pathogen.
2. This triggers the immune response.
3. VSV or adenovirus are common vectors.

Question 9

What is an example of an inactivated whole pathogen vaccine?

Answer 9

Poliovirus vaccine

Question 10

What is an example of an attenuated whole pathogen vaccine?

Answer 10

MMR vaccine

Question 11

What are replicating viral vector vaccines?

Answer 11

1. These are bioengineered viruses.
2. They can reproduce and express target viral proteins.
3. They mimic a natural infection to enhance the immune response.
4. As they are replicating they could cause disease in immunocompromised patients.

Question 12

What are non-replicating viral vector vaccines?

Answer 12

1. These cannot reproduce.
2. They transfer the genetic material of the pathogen and stimulate the immune response.
3. There is no further viral replication.
4. For example the ChadOx covid vaccine.

Question 13

How do protein based vaccines work?

Answer 13

1. They contain protein subunit from a specific virus.
2. These proteins are injected directly into the host to stimulate an immune response.
3. Often rely on adjuvants

Question 14

How do virus like particle vaccines work?

Answer 14

1. These mimic the structure of the virus but lack the genetic material.
2. They rely on the outer shell protein of the virus self assembling into a virus like shape so only possible for some viruses.
3. This happens spontaneously without the genetic material but as the viral proteins accumulate.
4. The immune system sees it as an actual virus.
5. Vaccines against the outer shell of the virus are good as it stops viral entry.

Question 15

What are recombinant protein vaccines?

Answer 15

1. These are protein based vaccines.
2. They use viral proteins produced using recombinant DNA technology.
3. This best example of this is the HPV vaccine.

Question 16

How do nucleic acid vaccines work?

Answer 16

1. They use DNA or RNA to get pathogen proteins made and expressed in cells for presentation.
2. They use the cellular machinery to make the proteins.
3. Nucleic acids are fragile so you put them in a lipid nanoparticle to get them into cells.
4. DNA vaccines need to reach the nucleus for transcription which is tricky.
5. RNA vaccines only need to reach the cytoplasm to work so they are more flexible then DNA vaccines.

Question 17

Why are lipid nanoparticles used in nucleic acid vaccines?

Answer 17

1. Cell membrane are made of lipids
2. Using lipid particles means it can interact and fuse with the cell membrane and help with vaccine entry.

Question 18

Are vaccine platforms specific to viruses?

Answer 18

1. No
2. All vaccine platforms are non specific and can be altered.
3. But the majority of vaccines are for viruses.

Question 19

What are most vaccines against?

Answer 19

viruses

Question 20

What does modern vaccine design consist of?

Answer 20

1. High through put analysis
2. Structural vaccinology
3. Synthetic biology/vaccines
4. Systems vaccinology and immunology

Question 21

Overview of high throughput analysis in vaccine design

Answer 21

1. Enables us to track development of different variants.
2. Helps us understand the sequences of the viral epitopes we want to target.

Question 22

Overview of structural vaccinology in vaccine design

Answer 22

1. Epitope recognition happens in 3D.
2. So inducing effective recognition and immune responses requires knowledge of structures in high detail.
3. This can make the difference between a successful and unsuccessful vaccine.

Question 23

Overview of synthetic vaccines in vaccine design

Answer 23

1. This means making all the vaccine components in the lab
2. Including nucleic acids or proteins.
3. Traditionally they were isolated from patients.

Question 24

Overview of systems vaccinology in vaccine design

Answer 24

This aims to understand why different people react in different ways to the same vaccine.

Question 25

What is high through put viral sequencing?

Answer 25

1. It uses a variety of sequencing approaches. 2. It is used to track variants of a virus and update vaccines accordingly. 3. It is essential for viruses that mutate quickly or viruses that spread quickly.

Question 26

What sequencing techniques can high throughput viral sequencing use?

Answer 26

1. PCR based methods 2. Next generation sequencing methods. 3. Metagenomics

Question 27

What is metagenomics?

Answer 27

1. Looking at the genetic material of a community of organisms. 2. So you would sequence everything in a sample not just the pathogen of interest

Question 28

Why is using the right sequencing technique in high through put viral sequencing important?

Answer 28

1. Different techniques have different efficiency and sensitivities. 2. If you have a low viral load you need to select the right technology otherwise what you get from a sample is limited. 3. Different sequencing technologies give different information about the sequence and have different abilities to get high quality sequences. 4. These are the sequences used to make vaccines so it is important they are right.

Question 29

What is the importance of sequence surveillance?

Answer 29

1. It is important for tracking the epidemiology of a viral disease. 2. You need to know how the sequences of the virus is evolving so you can update the vaccine accordingly. 3. You can also use it to know the circulating subtypes and use this information to design seasonal vaccines like for influenza.

Question 30

How does sequence inform vaccine design using SARS-CoV-2 as an example?

Answer 30

1. The 1st role out of the vaccines induced high levels of neutralising antibody titres. 2. But as the virus mutates the neutralising antibodies decrease and the risk of increase of severe infection. 3. This is random. 4. Protection lessens if you don’t update the vaccine 5. Sequencing of the new variants provide the information to update the vaccines. 6. Nucleic acid vaccines are more adaptable so this is easy.

Question 31

How does sequencing inform seasonal influenza vaccine design?

Answer 31

1. It starts with collecting patient specimens. 2. Then full genetic sequencing of lots of virus strains. 3. This is only possible due to high through put sequencing. 4. You use this predict the immunogenicity and make lots of vaccines to be prepared. 5. This takes lots of time and money.

Question 32

Why is knowing the structure of a proteins, not just the sequence, important for vaccine design?

Answer 32

1. Immune interactions happen in 3D. 2. The sequence doesn’t allow you to infer the structure or these interactions. 3. Different state of the same protein can have different potential to serve as a neutralising target for antibodies.

Question 33

How did the structure of the RSV fusion protein effect vaccine design?

Answer 33

1. The Pre fusion state of the protein is unstable and a potent immunogen. 2. The post fusion state of the protein is more stable but poorly immunogenic. 3. The pre fusion state generates good neutralising antibodies. 4. Effective vaccines to RSV could only be developed once the structure of the fusion protein was known as targeting the pre fusion form was key.

Question 34

What is structure-based immunogen design?

Answer 34

1. It is used to enhance neutralising antibody responses by focusing on specific epitopes and conformations. 2. It aims to present the epitopes that produce a good immune response. 3. It aims to hide the epitopes that prevent the immune response.

Question 35

What does structure based immunogen design consist of?

Answer 35

1. Conformational stabilisation 2. Silencing of non-neutralising sites. 3. Germline targeting 4. Epitope scaffolding 5. It can use one or an combination of these.

Question 36

What is conformational stabilisation?

Answer 36

Locking viral proteins in the prefusion state to boost antibody induction

Question 37

What is silencing of non-neutralising sites?

Answer 37

1. Clearing the way for the antibody to get to the neutralising target. 2. Usually getting rid of glycosylation

Question 38

What is germline targeting?

Answer 38

1. Using modified viral proteins to prime antibody precursors. 2. Good potential for HIV vaccines. 3. You target a B cell population and push them to produce broadly neutralising antibodies. 4. It is tricky but feasible for highly mutable pathogens

Question 39

What is epitope scaffolding?

Answer 39

Neutralising epitopes are transplanted onto synthetic scaffolds to focus immune responses.

Question 40

Why did a whole pathogen vaccine not work for SARS-CoV-2?

Answer 40

1. The immune system was probably recognising the post fusion spike protein. 2. This is not very immunogenic. 3. Some trialled whole pathogen vaccines lost effectiveness after 1/2 months. 4. You want to target the pre fusion spike but it is unstable by itself

Question 41

How do we stabilise the desired immunogen structure for vaccination?

Answer 41

1. You introduce some targeted mutations into the protein structure. 2. This increase the structural rigidity of the protein and can lock the protein in the pre fusion state. 3. This was used to stabilise the pre fusion SARS-CoV-2 spike.

Question 42

What mutations were used to stabilise the SARS-CoV-2 spike protein vaccines?

Answer 42

1. The main method was the the S-2P mutations. 2. These introduced 2 proline substitutions to lock the spike protein in its prefusion conformation and prevent premature structural changes. 3. K986P and V987P 4. This improved immune recognition, the neutralising antibody responses and led to higher vaccine efficacy. 5. This was used in the Pfizer, Moderna, Novavax and Johnson&Johnson vaccines.

Question 43

What needs to be balanced when using structure stabilisation?

Answer 43

1. Additional substitutions for make a more stable structure sounds good. 2. But you can over do it. 3. You need to find the balance between locking the structure and retaining the important epitopes for recognition. 4. Too many changes mean the immune system might not recognise it as the same protein.

Question 44

What is epitope mapping?

Answer 44

1. It is the mapping of sites on antigens that are more effective at generating neutralising antibodies. 2. This helps improve the immunogenicity. 3. You know where the antibodies are binding to the target antigen. 4. You can use this to generate more effective neutralising antibodies from a vaccine.

Question 45

What do antibodies to HIV normally target?

Answer 45

1. The HIV envelope but it is very changeable. 2. Most epitopes recognised by infected patients are not very neutralising especially once HIV mutates within the patient.

Question 46

What are broadly neutralising antibodies to HIV?

Answer 46

1. They recognise a distinct epitopes on a range of HIV strains. 2. They are generated by a limited group of HIV patients. 3. Mapping the sites these antibodies bind on the HIV envelope can help to design a vaccine that elicits a similar broadly neutralising response and reduce immune escape.

Question 47

What is the problem with making a HIV broadly neutralising antibody response?

Answer 47

1. They are not easily produced in patients that don’t normally produce them. 2. They were originally isolated from a very limited group of patients. 3. It is hard to program the immune response to elicit broadly neutralising antibodies with 1 method across a population. This is needed for a vaccine.

Question 48

How could you design a broadly neutralising antibody HIV vaccine?

Answer 48

1. Need to design an immunogen that binds to a BCR on a B cells that has the potential to produce a broadly neutralising antibody response. 2. Once activated the antibody will undergo somatic hypermutation and maturation which is random. 3. To ensure a broadly neutralising antibody is generated you need use selective boosting and push the antibody to what you want it to be. 4. This is very hard. 5. You need to know the detailed structure of the immunogen, the boosters and what mutations we want to induce. 6. But once successful this germline targeting could be used against a range of highly mutable pathogens.

Question 49

Is 1 broadly neutralising antibody enough?

Answer 49

1. No 2. there are multiple circulating strains of HIV that wouldn’t be covered by 1 broadly neutralising antibody. 3. You need to produce loads to cover multiple epitopes and strains. 4. You would use a multi-epitope vaccines to produce a very broad response.

Question 50

How do highly mutable pathogens use glycosylation?

Answer 50

1. HIV and influenza use it to cover key epitopes. 2. It doesn’t want the immune system to recognise these are they are the conserved antigens. 3. This ensures only high mutable epitope are exposed to the immune system.

Question 51

How can glycoengineering be used to optimise vaccine epitopes?

Answer 51

1. Glycoengineering can modify viral immunogen glycosylation. 2. This selectively removes the sugars to expose the epitopes you want the immune system to recognise. 3. Not yet tested in trials but could be good.

Question 52

How is epitope scaffolding used in vaccine design?

Answer 52

1. It stabilises and presents key epitopes on a structured protein framework. 2. It enhances the immune response and antibody recognition. 3. Used for HCV vaccine

Question 53

What are synthetic viral vaccines?

Answer 53

1. Synthetic vaccines are rationally designed immunogens. 2. They are created using genetic, structural and computational approaches to optimise antigen presentation and immune activation. 3. These include mRNA, self-amplifying RNA, DNA, protein subunit, virus-like particles and nanoparticle vaccines. 4. Uses insights from high-throughput sequencing and structural vaccinology.

Question 54

What was the mRNA vaccine for SARS-CoV-2?

Answer 54

1. Synthetic mRNA with single ORF encoding the spike protein. 2. Mixed in with lipids to form a synthetic lipid nanoparticle. 3. This protect the mRNA from degradation and enables it to cross the cell membrane and be translated. 4. The spike proteins appears on the surface of transfected cells and the individual makes a immune response to it.

Question 55

How are mRNA vaccines produced?

Answer 55

1. The Vaccine is designed based on the sequence. 2. The mRNA is made using an in vitro transcription system using DNA. 3. The mRMA is purified. 4. It is mixed with the lipids and precipitated. 5. finally it is filtered and you have the pure vaccine.

Question 56

What are the advantages of mRNA vaccines?

Answer 56

1. Rapid development - can be less then a year. 2. induces a strong immune response. 3. Scalable and flexible manufacturing.

Question 57

What are the current limitations of mRNA vaccines?

Answer 57

1. Cold storage chain requirement to prevent degradation. 2. RNA is fragile 3. Immunity can be less persistent then other platforms but we are not sure why

Question 58

What are self amplifying mRNA vaccines?

Answer 58

1. They are the same as normal mRNA vaccines but have an additional replicase gene from the harmless alpha virus. 2. This makes more copies of the RNA from the template. 3. More RNA = More proteins = more antigen presentation.

Question 59

Why are self-amplifying mRNA vaccines thought to be better than mRNA vaccines?

Answer 59

1. It is assumed the fading immunity seen in mRNA vaccines is based on having limited numbers of mRNA in the cell. 2. So a self-amplifying mRNA vaccine would solve this problem. 3. However this is just an assumption and it could be partly or wholly due to something else.

Question 60

What is the main vaccine that uses Virus-Like particles?

Answer 60

the HPV vaccine

Question 61

What is the HPV vaccine?

Answer 61

1. It protects against HPV which causes cervical, anal, oropharyngeal and other cancers 2. It also causes genital warts 3. It is a synthetic vaccine that mimics HPV structure. 4. It induces a strong immune response 5. There are 4 main types of vaccine that differ on the number of strains it protects against.

Question 62

What are the 4 different HPV vaccines?

Answer 62

1. Gardasil 4 or 9 depending on the number of types it protects against. 2. Cervarix and Cecolin are bivalent vaccines that protect against HPV types 16 and 18. 3. Gelcolin is a quadrivalent HPV vaccine that targets HPV 6, 11, 16, 18 and is approved in china

Question 63

How does the HPV VLP vaccine work?

Answer 63

1. It uses the L1 structural proteins from HPV. 2. L1 spontaneously assembles into a virus like particle. 3. This is presented to the immune system.

Question 64

How is the HPV VLP vaccine made?

Answer 64

1. The L1 proteins is transcribed and translated. 2. This is done in a prokaryotic or eukaryotic expression system. 3. The eukaryotic expression system will include the post translational modifications. 4. Prokaryotic systems are newer and cheaper and work well.

Question 65

How are recombinant protein vaccines made for HBV?

Answer 65

1. The HBV surface antigens can self assemble into a virus like particle. 2. Using recombinant DNA technology to insert the gene into an expression system to express the protein. 3. The protein self assembles into the virus. 4. It is a good vaccine and offer >90% protection.

Question 66

What is systems immunology in the context of vaccines?

Answer 66

1. Systems immunology integrates multi-omics data to provide a comprehensive analysis of vaccine induced immune responses. 2. Also uses clinical data and loots at different cohorts and populations. 3. It helps address key questions such as why some people develop strong protective immunity while other fail to response effectively to the same vaccine/immunogen.

Question 67

How was system immunology used to compare the efficacy of 2 different SARS-CoV-2 vaccines?

Answer 67

1. Compared ChAdOx1 and mRNA vaccines. 2. Showed they were both very immumogenic. 3. The mRNA vaccine induced slightly strong immune responses. 4. ChAdOX caused some thrombotic events due to increased expression of thrombosis proteins. 5. Systems immunology found the link between the immune signature of ChAdOx and blood clots.

Question 68

How was systems immunology used to examine non-responders to the HBV vaccine?

Answer 68

1. It is rare to not respond to HBV vaccine but some patients don’t produce the protective antibody titres. 2. Systems immunology found these non-responders had less IL-10+ B cells. 3. Il-10 promotes B cell differentiation and antibody secretion. 4. So these patients had an immune systems that was less primed for defence then responders.

Question 69

How was systems immunology used to observe the impact of age?

Answer 69

1. Younger people have a functional thymus and strong naive T cell response leading to better vaccine responses. 2. As you age the thymus reduces in function and causes T cell senescence. 3. This reduces immune surveillance and make older people less responsive to vaccination. 4. This shows that early vaccination is important and age specific vaccine strategies are needed.

Question 70

How was systems immunology used to observe the impact of sex?

Answer 70

1. Broad population studies are the only way to study this making systems immunology important. 2. Women tend to have more autoimmune disease. 3. But this is not stable as trends in inflammation change as you age. 4. The ratio between immune cells and the types of recognition change through sexes and as you age.

Question 71

How can systems immunology be used to interpret adverse events to vaccines?

Answer 71

1. When you vaccinate large cohorts they adverse events will not be equally distributed between the sexes. 2. Using systems immunology you can see the trends and what populations are more at risk. 3. For the covid vaccine boosters both age and sex were the main drivers of adverse events. 4. Younger females were more likely to have an adverse events but as age increased males became more likely to have an adverse event.

Question 72

What do synthetic vaccine provide?

Answer 72

1. Rationally designed immunogens that enhance antigen stability, 2, Immune activation 3. Vaccine durability 4. Reduction in risks compared to whole pathogen vaccines.

Question 73

What is overcoming immune evasion essential for?

Answer 73

designing next generation vaccines that provide broader and longer-lasting immunity

Question 74

Why is a HIV vaccine needed?

Answer 74

Due to rising resistance to anti retroviral drugs

Question 75

How do HIV broadly neutralising antibodies naturally develop in patients?

Answer 75

1. Broadly neutralising antibodies develop in a small subset of patients. 2. It only occurs after months/years of infection with HIV then infrequent germline B cell priming with extensive antibody somatic hypermutation. 3. In patients broadly neutralising antibodies are normally only of a single clonal lineage and contribute a small part to the patient's HIV antibody response.

Question 76

What are the traits of a HIV broadly neutralising antibody?

Answer 76

1. Under gone extensive somatic hypermutation 2. Selection of improbable antibody mutations that are necessary for broadly neutralising activity. 3. Presence of long heavy chain complementarity determining region 3

Question 77

How could HIV vaccine induce broadly neutralising antibodies?

Answer 77

1. Induce CD4+ Tfh differentiation which help induce broadly neutralising antibodies. 2. Minimise Treg and NK cell induction 3. Induce somatic hypermutation in B cells and reduce time for maturation of broadly neutralising antibodies.

Question 78

What was the first RSV vaccine and why did it fail?

Answer 78

1. A formalin inactivate RSV vaccine. 2. The immunity elicited by it primed for more severe disease after natural infection. 3. 80% of infected and immunised children were hospitalised and 2 died.

Question 79

Why is RSV a larger problem in low and middle income countries?

Answer 79

1. They have a high disease burden but low awareness of disease. 2. Limited availability and cost of diagnostic testing leads to a lack of understanding around the RSV disease burden and benefits of immunisation. 3. Cost of good and transport of a vaccine.