Lec 3- Biotechnology-based pharmaceuticals Flashcards
1
Q
Proteins v peptides
A
- A matter of size generally
- Peptides are compounds consisting of 2 or more amino acids
- Manufactures by either solution or solid phase peptide sysnthesis
- Protein are chains of 50 or more amino acids
- Manufactured by biotech methods
2
Q
Peptides, Proteins and delivery routes
A
- Currently there are only 2 peptides in oral formulations currently approved by the FDA
- Desmopressin
- Cyclosporine
- Look up both of these products, see their clinical use and consider their oral bioavailability
3
Q
Stability of biotech pharmaceuticals
A
- A pharmaceutical protein product should be stabilised against
- Unfolding
- Aggregation and self-association
- Denaturation
- Chemical degradation e.g. oxidation
- As well as
- Changes in the amino acid sequence due to mutation occurring during fermentation
4
Q
Protein structure
A
- Protein structure is not a simple matter
- Unlike traditional, low molecular weight drugs, protein therapeuticsz neeed to be stabilised in 4 levels
5
Q
Protein structure levels
A
- Primary: Refers to the linear amino acid sequence and position of the disulphide links
- Secondary: Defines the spatial relationship of the amino acid residues e.g. a-helix or b-pleated sheets
- Tertiary: This takes into account the extent and nature of stabilising side-chain interactions (H-bonding)
- Quaternary: refers to the spatial relationship between polypeptide subunits of a protein
6
Q
Physicochemical properties of proteins in solution
Hydrodynamic properties
A
- A folded protein is more compact than an unfolded one
- This results in lower solution viscosity and generally faster rate of sedimentation
7
Q
Chemical reactivity
A
- There is a great variety in chemical reactivity of functional groups
- Unreactive: Hydrocarbon side chain
- Reactive: Cysteine groups
8
Q
Water solubility
A
- Charged groups in proteins can increase solubility
- However ion-ion bond can form within the protein structure
- This neutralises the charge and influence solubility (As can electrolytes)
- Unfolding of a protein structure will also influence solubility
- When sufficient water solubility is lost, protein precipitation results (with loss of biological activity)
9
Q
Protein instability
A
- Degradation for proteins can be categorized into 2 distinct classes
- A) Chemical instability (irreversible)- covalent changes
- B) Physical instability (often reversible) - non-covalent changes
10
Q
Mechanisms involved in the degradation of protein pharmaceuticals
A
- Native functional proteins =>
- Paritally or fully unfolded proteins =>
- Non-covalent changes- Aggregation; surface adsorption; precipitation
- Covalent changes- Hydrolysis; deamindation; oxidation; racemization; disulphide exchange
- Paritally or fully unfolded proteins =>
11
Q
Denaturation
A
- Refers to alteration of the global fold of a native molecule
- I.e disruption of the teritary and frequently the secondary structure
- Essentially changes the physical structure of the protein
12
Q
Denaturation
Factors and outcomes
A
- Factors
- Temperature
- pH
- Addition of organic solutes e.g. urea, acetamide and formamide
- Addition of organic solvents e.g. alcohols, acetone
- Outcomes
- Protein becomes insoluble in aq solution (precipitation)
- Loss of biological activity
- Changes in molecular shape
- Susceptibility to enzymatic hydrolysis
13
Q
Stability problems observed with proteins
A
14
Q
Formulation strategies
A
15
Q
Protein stabilisation in the dried solid state
A
- If the solutions are not chemically and physically stable for 1 year or more in solution then lyophilization (freeze drying= using pressure to make product solid without using pressure) can be used
- These techniques dehydrate the protein and inhibit the degradation reactions observed with proteins in solution
16
Q
Formulation strategy: Excipients
A
- Excipients can be essential during lyophilisation to protect the integrity of the protein, during reconstitution and stabilising the protein in solution
