Viral vaccines and gene therapy Flashcards
(142 cards)
1
Q
Gene therapy notes.
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Uses of gene therapy
Production mechanism
Desirable qualities
Problems
2
Q
Gene therapy - uses
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Monogenic obvious.
Cancer and other chronic diseases to reverse symptoms.
3
Q
Production mechanism - overview.
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Separately generate structural proteins and vector genome in same cells. These will self-assemble to form viral vectors.
4
Q
Gene therapy - desirable qualities.
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Easy to produce. Safety. Targeted delivery. Transduction and transgene expression. Genome size.
5
Q
Gene therapy - safety
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Needs to be non-toxic and non-immunogenic.
Adeno vs adeno-associated viruses.
Problems - dysregulation of host function.
Uses of immunogenic viruses.
6
Q
Gene therapy - safety. Adeno problems.
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Adeno: most people have been challenged by at least one, so raise an immune response. Most immunogenic of vectors used. Prevents use of multi-dose regimens.
Jesse Gelsinger example.
7
Q
Gene therapy - safety. Adeno getting around problems.
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Adenovirus serotype 5 often used, with hexons expressed from a different serotype since Abs to those from Ad5 are common. Chimera is called rAd vector. BUT cross reactivity is common.
Elimination of all viral genes from genome helps.
Example: rAd-p53.
8
Q
Gene therapy - safety. Adeno example.
A
rAd-p53
Chimera to avoid recognition
Administered directly to tumour for tissue specificity
Encodes p53: leads to massive overexpression in tumour cells with tumour cell death. May have effect on bystander tumour cells, may act synergistically with chemotherapies.
9
Q
Gene therapy safety - adeno
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Problems, solutions, example, Jesse Gelsinger example.
10
Q
Targeted delivery
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CF example - importance.
Limiting delivery
Targeting tissues
11
Q
Targeted delivery - limiting delivery.
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C- type, adeno and AAV.
12
Q
Targeted delivery, targeting tissues.
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Particle and genome expression.
13
Q
Gene therapy - transduction and transgene expression.
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Desired; minimal toxicity, easily detectable, quantifiable and consistent expression.
Persistent expression.
Integration.
14
Q
Gene therapy - transduction and transgene expression. Persistent expression
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Adeno and AAV.
15
Q
Gene therapy - transduction and transgene expression. Integration.
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Could be useful, but issues with safety/progeny. Consider example AAV serotype 2.
16
Q
Gene therapy - genome size
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Herpes
Increasing effective genome size in adeno-associated.
17
Q
Hybrid vectors.
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Wild-type AAV with efficient internalisation and nuclear targeting of adenovirus.
18
Q
Production mechanism - details.
A
Delete coding region of virus, leaving sequences required in cis for packaging. Replace coding regions with cassette of choice. Express packaging proteins in same cell as synthesis of genome occurring in: self-packaging will cause genome with gene of choice to be packaged in viral proteins.
Purification is laborious, and difficult to scale up, but recent technological advances have dealt with this problem on the whole.
19
Q
Gene therapy - desirable qualities.
A
Easy to produce. Safety. Targeted delivery. Transduction and transgene expression. Genome size.
20
Q
Targeted delivery
A
CF example - importance.
Limiting delivery
Targeting tissues
21
Q
Targeted delivery limiting usefulness - C type retroviruses.
A
Can only infect dividing cells – limits usefulness. Recent work lead to nuclear localisation signal engineered into SNV, which made it capable of transducing non-proliferating cells.
22
Q
Targeted delivery limiting usefulness - AAV and adenoviruses.
A
Natural infections of AAV and adenoviruses are limited by transmission route, but this doesn’t occur with therapy.
23
Q
Targeting specific tissues - particles.
A
Adeno-associated. Many available serotypes – varying tissue specificity, so greater understanding is meaning that therapies can be rationally designed using different capsids with more specific/targeted delivery.
24
Q
Targeting specific tissues - transgene expression.
A
- Restricted to particular cell types or switched on and off by promoters, but dissemination of the virus particle itself can have harmful effects.
- Tumour specific transcriptional targeting: homologous recombination brought promoter in conjunction with reporter gene. This only occurs in tumour cells, as this is the only place that conditionally replicating adenovirus vector replicates.
- Transductional targeting by redirecting to specific cellular receptors.
25
Targeting specific tissues - CF problems.
Problem with CF; rapid turnover of lung epithelium means need to be delivered to basolateral surface – difficult.
26
Gene therapy - transduction and transgene expression. Persistent expression, adeno.
Adeno: Persist in nucleus as episomes, but not passed on to daughter cells. – persistent transgene expression in non-proliferating cells, but not stable genetic alteration for dividing cells.
27
Gene therapy - transduction and transgene expression. Persistent expression, AAV
Adeno-associated: persistent, can infect dividing or quiescent. – so could be used to transduce haematopoietic stem cells ex vivo.
28
Gene therapy - transduction and transgene expression. Adeno associated virus.
Adeno-associated viruses. Integrates into host chromosome 19 if helper virus not present. Older AAV therapies didn’t have this ability.
• Required: cis-active sequence ITR for replication and integration. So is Rep78, Rep68 and integration efficiency element.
• Required cellular factors unknown.
• Concerns about effect of Rep on cells means not included, so AAV genome maintained as an episome, not integrated.
29
Gene therapy - transduction and transgene expression. AAV example.
AAV serotype 2
• Integrates in host cell genome reliably into 19q13.4
• Recombinant viruses can take advantage of AAV integration by homologous recombination.
o Efficient gene targeting of cells. 0.1 - %1
o Safety: can elicit host immune response.
30
Increasing effective genome size of adeno-associated virus vector.
Despite relatively small genome size, formation of recombinant concatemers, and trans-splicing means that a gene can be split between two vectors. If both are delivered to the cell, it can be recombined to give a normal gene product. Allows deliver of genes up to 9 kb in size.
31
Genome size - herpes.
Herpes simplex in contrast. Large amount of genomic space for fragments of foreign DNA. Replication defective forms can carry up to 40 kB of foreign DNA.
32
Jesse Gelsinger example - illness and treatment.
Ornithine transcarbamylase deficiency treatment with intra-hepatic delivery of adenovirus vector capable of replicating in vivo.
33
Jesse Gelsinger cause of death.
• Cytokine storm lead to death.
o Lapses in reporting of toxicity
o Recombinant DNA advisory committee in disarray when approved
o Dissemination from liver unexpected.
o Possibly...
concurrent undetected viral infection – parvovirus B19.
Unstable phase of OTC that day.
34
Potential uses of immunogenic vectors with short-lived transgene expression.
o Some toxicity/immunogenicity could enhance anti-tumour effects in cancer therapies?
o In some cardiovascular therapies, transient expression is what you want.
35
Adeno-associated viruses, safety.
Immunogenicty and example (scAAV2)
36
Adeno-associated viruses, safety. Immunogenicity.
o Many available serotypes
o Little immune response in most of the clinical trials tried so far but CTL response can be concerning. Example: scAAV2.
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AAV example for safety - surprising toxicity.
scAAV2 for haemophilia B. Gene transfer and successful expression of human coagulation factor IX in haemophilia B patients. Several ongoing trials. Unexpected liver toxicity due to T cell activation by capsid.
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Dysregulation of host function due to insertion.
Example: SCID-X1 trial report
• Carries gene for y-c chain cytokine receptor: cures X-SCID without bone marrow transplant.
• Safety: insertion on or near oncogene LMO2 leukaemia.
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Virus vaccines
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History
Passive vs active
Live attenuated or dead.
Modern development
Therapeutic
Challenges.
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History of vaccination.
Variolation (Lady Wortley Montague).
Jenner
Earliest vaccination - vaccines that do not cause such bad disease.
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Passive vs active.
Antibodies vs antigens
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Live attenuated or dead
Comparisons:
Multiple immunisations, adjuvant.
Expense, cold chain.
Reversion, contamination.
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Virus vaccines
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History
Passive vs active
Live attenuated or dead.
Modern development
Challenges.
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Modern vaccine development
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Subunit.
Rational attenuation.
Viral vectors.
Nucleic acid vaccination
Adjuvants
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Challenges
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No effective immune correlate for sterilising immunity.
Mutability
Multiple clades
Incomplete understanding of immunity.
Animal models
HIV
Danger.
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Modern vaccine development
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Subunit.
Rational attenuation.
Viral vectors.
Nucleic acid vaccination
Adjuvants
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Challenges
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No effective immune correlate for sterilising immunity.
Mutability
Multiple clades
Incomplete understanding of immunity.
Animal models
HIV
Danger
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48
Live attenuated comparison, adjuvant.
* Enhances uptake of antigen
* Stimulates immune response
* Localises immune response to one area (depot effect)
* Targets antigens to particular pathways. Some adjuvants appear to be able to target subunits and dead viruses to MHC II pathways, inducing cytotoxic response.
49
Modern vaccine development, subunit vaccines.
Sometimes certain subunits are more immunogenic than others – in fact, some others may dampen immune response. Empty capsids are also non-infectious, but immunogenic
• HepB vaccine
• HPV vaccine – yeast expression systems.
50
Modern vaccine development, rational attenuation.
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DIS
Replication fidelity
Codon deoptimisation
miRNA control elements.
Zn finger nuclease control of virus production.
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51
Modern vaccine development, viral vectors
Mechanism
Previous exposure
Example
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Modern vaccine development, nucleic acid vaccination.
Injection of mice with influenza NP protein lead to immune response sufficient to protect the mouse from infection.
Stable, manipulatable, authentic post-translational modification, possible to make many different antigens (broad spectrum).
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Modern vaccine development, adjuvants.
Controlled release
Depot delivery
Immunostimulatory
Tolerogenic
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Modern vaccine development, adjuvants, immunostimulatory.
* TLR agonists
| * Flu HA-flagellin chimeras
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Modern vaccine development, tolerogenic
SIV study showing induction of CD8+ Tregs due to bacterial adjuvants lead to decreased activation of CD4+ cells and hence decreased early viral infection load (good for long term prognosis).
56
Challenges to vaccine development - mutability.
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especially RNA viruses.
• Error prone
• Intracellular recombination
• Rapid replication cycles
• Formation of escape mutants (passed on, become common in population)
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57
Challenges to vaccine development. Multiple clades.
* HIV: even among HIV-1 M subtypes, subtypes B and C can have 30-40% sequence diversity in places.
* HCV
* Dengue – careful. Immunity to a single serotype can increase severity of infections by other serotypes (Dengue haemorrhagic fever and Dengue shock syndrome). Antibody dependent enhancement of replication in Fc-receptor bearing cells. Therefore any vaccine must be tetravalent.
58
Challenges to HIV vaccine development.
Mutability, multiple clades. Stimulation provides targets.
Poor antibodies in natural infection.
Memory T cells take time to activate, so would already have infected T cells.
Timing - early stages crucial but difficult to access.
Translating genetic insights into intervention.
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Challenges - incomplete understanding.
HIV vaccines: some aimed at generating CD8+ response actually lead to increased infection: this was with adenovirus vectors expressing Gag, pol or Nef. STEP trial.
60
Challenges to vaccine development - animal models.
* HCV, HIV; highly species specific
* No validated animal model for Dengue
* Humanised small animal models: expensive, limited in number.
61
Challenges to vaccine development - animal models, HIV
In SIV macaque model, rCMV vector expressing Gag, Pol, Tat, Rev, Nef, Env and Pol lead to decreased viraemia and apparent clearance in 50% of monkeys, due to induction of peripheral effector memory T cells. But
This is a different disease, different host, different vectors from those tried in humans.
62
Challenges to vaccine development - animal models, dengue.
• No validated animal model for Dengue
o Difficult to do initial studies
o Lack of knowledge about dengue immunology
difficult to assess protective mechanisms of vaccines.
63
Challenges to vaccine development - animal models, humanised small animal models.
o For HCV, potential to use humanised mice, with human hepatocytes
o For Dengue
64
Challenges to vaccine development, HIV - genetic insights.
* Determinant of progress of HIV infection: HLA class I alleles in T cell antigen. HLA-C affects cell surface expression. How to translate these into therapies is difficult, as resulting differences in cell physiology subtle, species specific, difficult to reproduce in vitro and etc.
* Reversing virally mediated downregulation of genes very difficult e.g. HLA-A and B to prevent CTL, but not C and E.
65
Modern vaccine development, viral vectors, mechanism
Attenuated (replication defective) virus has gene of different virus inserted. MVA (well understood, highly attenuated, induces IFN, no soluble IFNR, expreses IL-B receptor) or DISC herpes virus Challenge with this: things do not always happen as we expect (mouse pox immunisation)
66
Modern vaccine development, viral vectors, previous exposure.
If the patient has already been exposed to the vector, it could just cause a more vigorous response to the vector while dampening response to vaccinating antigens.
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Modern vaccine development, viral vectors, example
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Chimeric vaccines (e.g. Dengue and Yellow Fever)
• Yellow fever backbone with Dengue preM and E genes inserted. Safe, neutralising Ab titres moderate to high, but only 30% efficacy.
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68
Modern vaccine development, rational attenuation, codon deoptimisation.
Early days. Mutate viral genomes to alter codon pair frequencies decreasing translation efficiency. Increased CpG and UpA dinucleotide frequencies also attenuate replication.
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Modern vaccine development, rational attenuation, replication fidelity
Mutate polymerase, e.g. Gly64Ser, to decrease replication fidelity enough to cripple the virus (most viral fidelities are somewhere like 1/no. Of bp in genome)
70
Modern vaccine development, rational attenuation, DISC
disabled attenuated single cycle – because viral particles can infect, but progeny cannot, the immune response is accurately balanced and strong, but little chance of reversion.
• E.g. gH gene in cellular genome, deleted from viral genome. Viral genome packaged, buds out acquiring gH, but progeny have no source of gH.
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Modern vaccine development, rational attenuation, miRNA control elements.
* Change cis-acting factors to alter tropism.
| * Species specific attenuation of Influenza A virus.
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Rate of new infections
Rate of new infections =β x I/N x S where β = probability of transmission on meeting, N = population size, I = infective individuals and S = susceptible individuals.
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Epidemic turnover
When rate of new infections begins to decline, that is when the epidemic turns over. This occurs when the number of infectives >> susceptible, and no. of susceptible is a limiting factor.
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R0
Basic reproductive ratio. Summary of potential for spread, but only really accurate at start of epidemic, as later herd immunity leads to a falling effective R0 (e.g. R). Only in completely susceptible population, so R is more helpful in real-life scenarios.
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Calculating R0
R_0=βT_1, where T_1 is duration of infectiousness. β encompasses both the probability of transmission per contact, and the number of contacts per time.
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Calculating R (effective R0)
R=R_0 x S/N
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S/N is also called
s. Proportion of population which is susceptible. If we vaccinate p, then s = 1 - p.
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When R=1, s =
1/R_0. No epidemic occurs.
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p (proportion vaccinated) =
1 - s.
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p required to eradicate disease.
To eradicate disease s = 1/R.
| Since p = 1 - s, then to eradicate disease, p = 1 - 1/R.
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Problems in vaccination uptake.
decrease dangerous even if previous coverage. Especially as often clustered e.g. affluent boroughs in London. Some countries do not have infrastructure to allow necessary levels to be achieved. Measles: Democratic Republic of Congo reports 100% coverage, but this is unlikely given infrastructure and political turmoil. Also, marginalized communities.
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Structuring of populations.
People only contact a few regularly: not a well mixed population. Trace individual contacts and offer vaccination or prophylactic treatment.
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Things affecting efficiency of contact tracing.
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Infectious period before recognised (helpful if short)
Latent period (helpful if long)
Low economic impact of treatment (or people will refuse).
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84
Ideal diseases for contact tracing.
• A high R0 with a low proportion of asymptomatic infectious or proportions of infections prior to symptoms is ideal for contact tracing.
o Smallpox and SARS ideal.
o HIV wouldn’t work
o Influenza doesn’t really work for either atm.
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Equation suggesting that epidemics should tend to an equilibrium.
T=2π√A(T_I+T_E),
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Causes of seasonality of epidemics.
• Animal populations: seasonal births
• Human populations: time outside, Vit D levels, school terms (measles: peaks at start of school terms, decreasing in height through the year), populations movements e.g. agricultural season.
E.g. arrival of new naive population leads to sustained oscillations.
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Effect of reducing birthrate on epidemics.
Reduced birthrates = reduced naïve population = increased time between epidemics. E.g. measles
• Baby boom smaller annual
• Even when rest of country followed London, higher birth rates in Liverpool due to Catholic immigrants annual epidemics.
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Threshold populations and cities.
Need threshold population size to maintain, otherwise goes extinct in between epidemics. But big cities can act as reservoirs, leading to a hierarchy of transmission. Thus London, with a few exceptions, drove epidemics across the UK. Requires travel and large cities (travel – e.g. Norwich).
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Measles in Niger
High birth rate even with 70% vaccination, population susceptibility same as in England and Wales before vaccination.
Surprisingly despite high transmission rates, spatial spread is low, so immunization of nearby districts could be implemented early enough and save lives.
Agricultural season --> high amplitude of seasonal forcing.
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Vaccination and age of acquisition.
Vaccination tends to drive up age of acquisition of diseases since there is less probability that you will contact and infectious person. For many childhood illnesses, later exposure is more dangerous. E.g. Rubella - mild disease except when congenital, yet vaccination increases the mean age of first infection.
Honeymoon period: herd immunity until numbers of unvaccinated have reached a high enough level to cause epidemic (e.g. measles after Wakefield controversy).
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Epidemiology in measles vaccination.
High R0 --> very high vaccine coverage and chance of importations into countries where endemicity has been eliminated.
Wealthy countries have motive to eliminate from poorer.
Epidemics show poor coverage.
Pulse vaccination or routine?
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Pulse or routine vaccination.
Routine vaccination: some children will always be under age. Alternative is pulse vaccination to initially eliminate, then routine vaccination later.
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Importation of measles into previously clear countries.
Increasing travel increases chances of imported measles cases. Eradication of trains of transmission from imported cases is also needed.
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Controlling outbreaks (quarantine and contact tracing) rather than introducing elimination.
* R0>1
* Tg = mean time between infection of individual and infection of individuals that infected individual infects.
* Proportion of transmission before overt clinical symptoms (θ).
* Examples: HIV and SARS.
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θ
Proportion of transmission before overt clinical symptoms .
o Isolating symptomatic individuals can only control outbreaks in which the number of individuals infected by asymptomatic carriers is less than 1. For control through isolation, θ
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Increasing θ.
o Delays in isolation increase θ. Therefore, efficiency in isolation and treatment of infected individuals is key. Rapid isolation after infection is achieved when symptoms occur rapidly, when there is little stigma attached (so people self-diagnose), when symptoms are easily differentiated from clinically similar diseases.
o Symptoms unpleasant autoisolation.
97
Example of isolation effectively limiting isolation.
o SARS: peak infectivity more than a week after onset of symptoms isolation on symptoms can interrupt transmission.
98
Measles dynamics in sub-Saharan Africa.
If outbreaks are highly episodic, and coverage is below threshold level, could lead to build up of naïve population and sudden major epidemics.
High birth rates and high seasonal outbreaks lead to complex multiannual outbreak dynamics.
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High birth rates and high seasonal outbreaks lead to complex multiannual outbreak dynamics. In sub-Saharan Africa.
Occasional large outbreaks followed by years with none: very strong seasonality leads to extremely deep troughs, so CCS is much larger than for Europe, requiring external reintroduction. Major outbreaks will quickly overwhelm health resources: a balance needs to be struck between routine, supplementary and reactive control straties.
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Vaccination for measles in sub-Saharan Africa.
Outbreak response vaccination increases time between outbreaks, making outbreaks larger. But overall, increasing levels of routine vaccination and ORV lead to decrease in cases and outbreaks.
101
Mechanisms of gene therapy
liposome
naked DNA
Virus vector
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Interest in gene therapy
over 2000 trials
64% cancer
9% monogenic disorders.
Crisis in confidence after Gelsinger disaster.
103
2 approved forms of gene therapy
Gencidine
| Glybera
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Difficulties with gene therapy
Short-lived, immune response.
Complexities of multi-gene disorders.
Breaching Weismann barrier
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Breaching Weismann barrier.
Some therapies may breach the Weismann barrier (between soma and germ-line) protecting the testes, potentially modifying the germline, falling afoul of regulations in countries that prohibit the latter practice.
Insertional mutagenesis. Cost.
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Gene therapy trials: AAV
achromatopsia trial,rAAV gene therapy replaces red cone opsin, resolution for 33 months in canine model
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Gene therapy trials: retroviruses.
September 2010 used lentiviral vector to transfer B globin gene into patients own blood and bone marrow cells : cure
75% people with B thalassaemia dont find a matching donor
Phase I trials approved by FDA
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Gene therapy trials: adenoviruses.
SERCA gene heart trials
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HIV therapeutic vaccines
Immune response sub-optimal. Thes would target immune modulatory mechanisms including PD-1 blockade.
Strategies also in development for inducing reactivation of HIV to allow targeting of latently infected cells.
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Reactivation of latently HIV infected T cells.
Initially - IL-2, IL-7 or global activation with anti-CD3 Abs.
But more recent understanding of latency mechanisms has lead to work on HDAC inhibitors and PKC agonists.
111
RV-144 vaccine
Anti-HIV.
Reached phase III trials in Thailand.
31.2% efficacy, but view with caution, as that was only a difference of 23 people more in the control group.
Abs did not neutralise particles, but directed ADCC against virally infected cells.
112
Vaccine development HIV despite multiple clades.
Recent trial attempted to overcome using prime-boost strategy with HIV DNA clade B gag/pol and nef, and calde A, B and C env genes, followed by rAd5 vaccine expressing some. Reached phase II trials, but stopped due to poor efficacy.
113
When did polio become a problem?
First half of 20th century, with increased mean age of first infection when child no longer protected from viraemia by maternal Abs.
114
Current state of polio elimination.
Eradicated from most countries in world including India. Small epidemic can cause rapid reimportation to previously clear countries. E.g. stopping vaccine in Nigeria in 2002 resulted in spread across Africa, into Middle East and Indonesia.
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Challenges to eradication of polio
* Largely silent disease, so θ is very high. Quarantine therefore is ineffective.
* cVDPV, iVDPV, VAPPs
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Different approaches to vaccination
Routine
Mass
Focused response to outbreaks
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Routine vaccination against polio.
In low season if seasonal. In developing countries, babies would see the virus before immunisation so didn’t work. Preferred route if possible.
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Mass vaccination vs polio.
vaccinate all children under 5 in massive, rapid campaigns. E.g. in India, 120 million were vaccinated in 24 hours. Result: no susceptible hosts in country. Usually 2 or 3 rounds are needs, with mopping up campaigns, and limited routine vaccination as well. Logistically difficult.
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Polio; focused response to disease outbreaks.
Northern Syria
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Different types of polio vaccine.
3 serotypes, so most are trivalent.
Salk (IPV)
Sabin (OPV)
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Polio 3 serotypes.
Immunity to one does not give immunity to others, so most vaccines trivalent (or if a strain not present in a country, bivalent one is used). If a strain has been wiped out in an area, then tends to be better to use bivalent strain as irrelevant serotypes do not interfere with gut infection.
122
Salk vaccine (IPV)
Probably doesn’t prevent infection of the gut, but where hygiene levels are high, respiratory route is more important, so can break transmission that way. A theory, not proven.
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OPV - key themes
Attenuation reversion tunes for growth
OPV will never cause successful eradication.
Withdrawal of OPV will be necessary to eradicate.
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OPV - attenuation reversion tunes for growth. Mutations.
Thermodynamic stability due to mutation in 5’ NCR, which forms part of the IRES.
At least one attenuating mutation in capsid region.
Mutations that compensate for mutations weakening pentamer assembly.
125
OPV - attenuation reversion tunes for growth. Driving factors.
Attenuation reversion may be driven by hostile gut conditions – so if too hostile, reversion may be driven faster, suggesting that effective attenuation has a narrow phenotypic window.
Some reversion occurs in all vaccine recipients.
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Why will OPV never cause successful eradication?
VAPPs
iVDPVs
cVDPVs
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VAPPs
Reversion leads to poliomyelitis in 1/750000 vaccinees (VAPPs). This may spread to 2 or 3 transmissions, but generally dies out. Type 1 causes fewer of these, because there needs to be more incremental changes to revert to virulence.
128
cVDPV
Reversions in poliovirus from vaccine can lead to circulating vaccine derived poliovirus. E.g. Hispaniola outbreak related to Sabin Type 1, probably been circulating for 2 years before outbreak. Vaccine coverage had fallen to 50%.
Recombinations with enteroviruses also a problem.
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iVDPVs
A few hypogammaglobulinaemic patients become chronic excretors of poliovirus after being given the vaccine: immunodeficiency vaccine derived polioviruses
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Withdrawal of OPV necessary for eradication.
Bring in more general use of IPVs.
| Develop live vaccines that will not revert.
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Bringing in more general use of IPVs.
issues in places with circulating cVDPVs but low levels of hygiene. Also concerns about keeping containment effective while making IPV (which requires growth of large quantities of wild-type virus.)
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Future development of live polio vaccines.
Non-revertant strains.
Strains that only grow in culture.
Stabilise empty capsids.
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Future development of live polio vaccines. NON-REVERTANT STRAINS.
o Codon deoptimisation to make translation less effective. Too many changes revert using natural mutation. High particle: infectivity ratio, but poor growth rates.
o Only mutate capsid region – if recombine with enterovirus, not a poliovirus.
o Codon deoptimisation to make translation less effective. Too many changes revert using natural mutation. High particle: infectivity ratio, but poor growth rates.
o Only mutate capsid region – if recombine with enterovirus, not a poliovirus.
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Future development of live polio vaccines. CULTURE ONLY STRAINS.
Design many mutations that weakens highly structured IRES at 5’ of RNA genome, leading to further mutations preventing replication rather than reversion.
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Future development of live polio vaccines. STABILISED EMPTY CAPSID.
Would need to be genetically engineered.
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Phases of polio disease
Asymptomatic is most common.
Minor disease causes malaise, fever, sore throat.
Major disease can involve aseptic meningitis, spinal/bulbar poliomyelitis, or encephalitis.
137
Non-nucleoside reverse transcriptases
Pyrophosphate analogue for HCMV, foscarnet.
Non-nucleoside reverse transcriptase inhibitors licensed for HIV-1. Efavirenze, Nevirapine (used as monotherapy to prevent transplacental; resistance an issue).
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Protease inhibitors
Used as monotherapies, but now some resistance. HIV treatments such as Lopinivir. Several are boosted by ritonovir, also a protease inhiibtor.
Some are used in HCV e.g. telaprevir.
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Aciclovir resistance
Immunosuppressed HSV patients may cause development of resistant strains because there is prolonged shedding.
Does not work agaist VZV because it is inherently less sensitive.
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Concerns with resistance
o Rapid mutability
o Segmented viruses; shift causing pandemic strain to acquire drug resistance.
o Transmission of drug resistant strain
HSV acyclovir resistance does not appear to transmit, but many others do.
o Latency
o Recombination.
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Dealing with drug resistance in HIV.
o Check before treatment, at failure and after interruption.
o Combination therapies e.g. HAART
o Counseling, education and support to improve compliance.
o Monitor transmission of drug mutation.
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Mechanisms contributing to HIV resistance.
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High mutation rate
Recombination
Latency
Compartmentalisation
Transmitted drug resistance mutations.
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