Vaccines, nasal drug delivery and Gene delivery Flashcards
what kinds of vaccines already exist and when were they invented?
Live and virulent <1800
Live and attenuated 1870<
Recombinant after 1982
What are the two types of immune responses
Adaptive Immune response
Innate immune response
What is the innate response
It is conducted by myeloid cells
It is the first line of defence
Cellular rapids
Cells involved in innate immunity include:
Dendritic cells
Macrophages, mast cells. natural killer cells. basophils, complement protein, eosinophils, Neutrophils, T-cells and Natural killer T-cells.
What is the Adaptive immune response and what cells are involved
Specialised immune, immuneological memory, humoral and is slow.
B cells, antibodies, CD4 T-cell and CD8 T-cells
Describe as a summary the immune response
The pathogen antigen binds to APC’s recognition receptors.
The APC will digest it, break it down and presents fragments of it on to T-cells and B-cells which activates the Adaptive immune system
What are the current convectional vaccines
Live attenuated : live microbial agent that is mutated to reduce the ability to grow in human cells
- Not safe for immunocompromised patient
- Bacillus Calmette-Guerin, measles, mumps, yellow fever and rubella (MMR)
Inactivated: Microorganism or viruses treated with heat or chemical (CH2O).
- Inactivated vaccine are less infective but are safer
- Polio, Hep A, cholera, influenzas
Toxoids: Inactivated toxins that cause disease (Tetnus and diphtheria).
-Can be as vaccines or to improve immunogenicity of other vaccines (haemophilus influenza)
List some more conventional vaccines
Subunit vaccines: uses small part of the organism - gene from genome.
Recombinant DNA tech helps the development of these vaccines.
Characteristic of subunit vaccines
Unable to revert to pathogenic form Decreased toxicity Reproducible production Improved antigen specificity Immune response is though short lived Several booster doses are needed
What are the types of covid-19 vaccines available
Live attenuates, inactivated, Replicating viral vector, non-replicating viral vector, DNA vaccines, RNA Vaccine and subunit vaccines
How can subunit vaccine efficacy be increased?
use of adjuvants: used in combo with specific antigen to produce robust immune response
Aluminium salts eg Aluminium hydroxide, phosphate, potassium phosphate and aluminium.
How does adjuvants aid immunisation?
Induces a polarised immune response (Th-2 biased immunity- strongly humoral).
High levels IgG1 antibodies and cytokines such as IL-4
Which adjuvants are used for which vaccines?
Aluminium salts: Various diseases
MF59: Influenzas
AS03: Influenzas
AS04: HPV, HBV
virosome: Influenza and HAV
What are the types of Adjuvant mediated immunity
Danger model Signal 0 Recombinant signal 2 Emulsion Depot effect
What is the danger model for an adjuvant
Parenteral delivery -> cells rupture releasing intracellular contents, mitochondria, uric acid and heat shock proteins Alarmins.
Describe the Signal 0
Binding of Alarmins and PAMP to pathogen binding receptors on cells such as TLR, antibody Fc receptors.
Leads to activation of the cells and causes proinflammtory cytokines, chemokines and co-stimulatory (CD80/CD86) molecules to activate T-cells
Describe recombinant signal 2
Adjuvant activate macrophage and B-cells to induce CD80 AND CD86- upregulate the expression on the dendritic cells.
Emulsion or particles in the adjuvant mediated immunity
Vesicular structures carry antigen either by entrapment or adsorption. Vesicles are naturally occurring and appropriately sized to be endocytosed by cells (composition and size critical)
Depot effect in Adjuvant Mediated Immunity
Localisation of antigen (with or without adjuvant) at the injection site and not in the lymphoid organs (although increased presentation of antigen in the lymphoid organs may be a direct consequence of the depot-effect and is often the desired effect) [route of injection, viscosity, particle size]
what are the characteristic of an ideal adjuvant
> Must contain sufficient PAMPs to alert immune system without hyper-stimulation
Cause some tissue damage to release Alarmins
Stays associated with antigen until uptake
Free antigens, peptides and genes rapidly degraded or cleared
Typical adjuvants forming a depot
- Chemical \precipitates (Aluminium salts) (? Uric acid/danger signal)
- Oil-in-Water / Water-in-Oil (IFA, Montanide MF59) [MF59: 0.5% Tween 80, 0.5% Span 85, 4.3% Squalene, Water for injection, 10nM Sodium citrate] – direct effect on Cytokine levels
- Protein precipitates (IC31)
Typical adjuvants activating DCs
> CpGs, IC31 (TLR9), MPL (TLR4), TLR-2, 3,4,5,6,8 ligands
Peptidoglycan (nods), QS21, Saponin
Heat-shock proteins
Typical adjuvants targeting DCs
> Liposome
Virosome and virosome like particles
Microparticles (polymers)
ISCOMs (Th1 & Th2) /Iscomatrix (Th2) [Immune stimulating complexes, 40nm, saponins (Quil A), lipids, cholesterol and antigen; chol binds to saponin forming 12 nm rings that are fixed together by lipids to form spherical NPs. Hydrophobic and amphipathic antigens preferred. Increased antigen presentation to B cells and uptake by APC inducing a potent humoural and cellular response]
IC31 [11-mer antibacterial peptide (KLKL(5)KLK) and a synthetic oligodeoxynucleotide (ODN1a) which is a Toll-like receptor 9 agonist without containing cytosine phosphate guanine (CpG) motifs –humoural response]
what are Nano-enabled Vaccine Formulations
Liposomes, niosomes and other vesicular adjuvants
Enhancing protection and APC uptake of antigen
Can include immunomodulating molecules in liposomes prepared with dimethyldioctadecylammonium
Factor to consider concerning parenteral vaccination
> Needle-phobia especially in children making vaccination unnecessarily stressful
Needle stick injuries (300,000 injuries annually in US)
Re-use of needles (developing countries-cost)
Need for sterility
Need for cold transport
Need for trained healthcare professionals
IgG and IgM produced circulate in the blood
-Thus no mucosal antigen produced
-Most infections start at a mucosal surface
Factor to take into consideration for mucosal vaccines
> Improved patient compliance
No need for sterility
No need for cold transport
No need for trained healthcare professionals
MALT (mucosal associated lymphoid tissue)
IgA secreted at mucosal surfaces and IgM produced circulate in the blood
-Most infections start at a mucosal surface
Types of mucosal vaccinations
> Oral: Antibody response in SI (proximal segment), ascending colon, mammary and salivary glands
Rectal: Antibody response –small in SI and proximal colon
Nasal or tonsilar: Antibody response in upper airway mucosa, regional secretions such saliva, nasal secretions (no gut response)
Vaginal but also nasal: strong IgG antibody response in human cervicovaginal mucosa
-Menstrual status can affect intensity of immune response in genital secretions
What the examples of oral mucosal vaccines
-Fleas from cows infected with cowpox
-Sabin oral polio vaccines
-Typhoid fever
Cholera
How does oral mucosal vaccines work
Recognised by microfold (M-cells) in the peyer’s patch of the intestines and by DC
What is the formulation ingredients for Pfizers covid-vaccine
Active: ModRNA encoding viral pike glycoprotein
Lipids: Ionisable Lipid: ALC-0315: (4-hydroxybutyl) azanediyl)bis (hexane-6,1-diyl)bis(2-hexyldecanoate)
Peg Lipid: ALC-0159: 2-[(polyethylene glycol)-2000]-N,N-ditetradecylacetamide
DSPC: 1,2-Distearoyl-snglycero-3-phosphocholine
Cholesterol
Salts: Potassium chloride, monobasic potassium phosphate,
Sodium chloride, potassium dihydrogen phosphate
Others: Dihydrate sucrose, water for injection
What are the ingredient for Az/oxford vaccine formulation
Active: Non-replicating chimpanzee AdV5 expressing spike protein
Excipients:
>Polysorbate 80 (0.1mM) [Non-ionic surfactant/emulsifier]
>Sodium Chloride (35mM)
>Magnesium chloride (1mM) [Stabiliser heat, Immune response booster]
>Histidine (10mM) (pKa 6.1)
>Sucrose (7.5% w/v)
>Ethanol (0.002 mg/0.5mL dose) [Not enough to cause noticeable effects – Deemed negligible by Muslim scholars (comparable amount of ethanol in natural foods or bread)]
What the the storage requirements for covid vaccines
Pfizer RNA vaccine : -70oC and stable for 5 days at 2-8oC
Moderna RNA vaccine: -20oC and stable for 30 days at 2-8oC
What are the advantages of nasal drug delivery
- Rapid onset of action (e.g. migraine)
- Avoidance of GIT or 1st pass metabolism
- Preferred to parenteral or rectal
- Lower costs
- Chronic disorders
- Delivery of peptides, proteins
- Vaccination esp for resp tract infections as influenza or TB
- CNS access: reach local receptors
- Delivery across the BBB
Describe the nasal cavity
It is around 160cm2 and is made of 3 sections: >Nasal vestibule and atrium >Respiratory region -Inferior turbinate -Middle turbinate -Superior turbinate >Olfactory regions
What is the function of the vestibular region and were is it located
Located at the opening of the nasal cavity and filters airborne particles
Particles filtration:
> 10 µm filtered by vibrissae at nostrils
5
Describe the respiratory regions
Make up 77% of the nasal cavity
Has mucociliary to remove particles (5-10um)
Made of pseudostratified columnar secretory epithelium
What is the olfactory region of the nasal cavity
Make 10% of the nasal cavity
Composed of non-ciliated pseudostratified columnar epithelial with 6-10m olfactory sensory neurons
Used to smell
What are Turinates
They are thin bony structure that line the mucosa and protrude into the nasal airways
Maxillo-turbinate: Cover respiratory epithelium
- Warm, humidify and cleans inhaled air
- Removes H20 from exhaled air
Ethmo-turbinates
- Cover olfactory epithelium
- High SA for neurons
- Associated with cribriform plate
- olfactory receptor nerves synapse with the mitral olfactory nerve cells through the cribriform plate
What is the delivery sites for nasal drug formulations.
-Have large surface area for absorptions Two potential deposition sites 1.anterior nares (nostrils) - longer residence time - low permeability
- posterior turbinates
- high permeability
- shorter residence time
Two delivery routes
- volatile substances via olfactory receptors
- essential oils
- odours - potential non-invasive route for CNS delivery circumventing the BBB
Describe muco-ciliary clearance
Maintain integrity of muco-ciliary system by removing trapped
Ciliary activity is the driving force: 1000 strokes /min
Mucous flow rate of 7mm per min
Types of local nasal delivery formulation
Drops, sprays, powders, washes, semisolids, sticks
Single dose or mutli-dose +/- device
Non-irritating +/- no A/Es on mucosa or cilial function
Consideration when formulating nasal preparations
Critical processes
- Deposition
- Clearance
- Absorption
How does systemic nasal delivery work
Mimic parenteral delivery
Fast onser of action
How does systemic nasal delivery work
Mimic parenteral delivery
Fast onset of action
-Pain relief: morphine, ketamine, enkephalin
-Erectile dysfunction: Apomorhine
Advantages of nasal systemic delivery
Surface area Permeability Avoid 1st pass Low enzymatic activity Self administration Patent extension
Disadvantages of nasal systemic delivery
Limited Dose Size Low absorption of high MW hydrophilic drugs Tissue irritation Low reproducibility Mucociliary clearance
Barriers of drug absorption in nasal delivery
Mucus
Rapid mucociliary clearance of administered formulation
Permeation across epithelial cells
Metabolism (enzymatic degradation in the vestibule and /or crossing epithelium
How the barriers of nasal drug delivery overcome
Inhibit mucociliary clearance/manipulate contact time (viscosity enhancers)
Enhance permeation using absorption/penetration enhancers
E.g. chitosan derivatives able to reversibly open the TJ
Enzyme inhibitors
What are the novel technologies used to overcome
Mucolytics Absorption enhancers Viscocity enhancers Mucoadhesives (sol, dry powder, colloidal) >Reduce MCC >Reduce clearance >Increase local drug concentration >Protect drug from dilution
Absorption enhancers
Insulin + dimethyl-b-cyclodextrin = 100% bioavailability
- More effective when combined with a mucoadhesive
- Bile salts, surfactants, fatty acids (phospholipids and lysophospholipids) improved transcellular transport of drugs but result in membrane damage
Chitosan and positively charged polymers
> Chitosan glutamate (250 KDa, 80% deacetylation)
Sheep: Insulin peak plasma level increased from 34 to 191ml U-1 and a 7-fold increase in AUC
Humans: 9-15% bioavailability of SC Insulin
What are some licenced therapies for nasal delivery
PecFent: Pain relief which contain fentanyl (Higher than plasma level than oral)
>Highly lipophilic
>In a pectin based gel in which the drug diffuses out of. = sustained release
