D. PROTEIN FORMULATIONS 2 Flashcards
properties of protein and peptide drugs
- large, hydrophilic molecules 0.5-150 kDa
- not well transported across biological membranes
- typically <2% bioavailability even for small peptides
- injected polypeptides have a small half-life
- major disadvantages compared with small MW drugs
- physical, chemical, biological instabilities
physical stability of proteins and peptide drugs
- easily lose their native 3D structure by denaturation
- due to hydrophobic conditions, surfactants, pH, solvent, temperature, dehydration, lyophilisation
- can happen in the bottle or during manufacturing
aggregation/precipitation of drugs
- denatured, unfolded proteins may interact
- aggregation becomes precipitation and we can see large particles
- loss of function
adsoprtion of drugs
- polar and non-polar residues are adsorbed at interfaces
air-water: foaming
air solid: Insulin adsorbs to many surfaces like plastic tubing or containers so we use delivery pumps, glass or plastic syringes - to decrease the dose lost
chemical stability of proteins and peptide drugs
- deamidation: asparagine and glutamine residues
hydrolysed to form a carboxylic acid - oxidation: methionine, cysteine by oxygen in air
mechanism via oxygen radical
catalysed by transition metals (e.g. Fe and Cu)
mediated by antioxidants (e.g. Ascorbate)
PEG can result in peroxide formation - photo-oxidation
biological stability of protein and peptide drugs
- hydrolysed to amino acids and small peptides in the GIT
- Pepsin (preferred aa: phe, tyr, trp)
- Trypsin, chymotrypsin and aminopeptidases [small intestine]
- Carboxypeptidases - produced in pancreas, act in small intestine
- denatured by gastric acid
- Very few peptides are stable to biodegradation: Cyclosporin (11 residues, cyclic - therefore very small) and TRH (very small)
- Colon: Fewer digestive enzymes but substantial microbial enzymes
what is the order for pre-formulation
- selection of candidate drug (protein)
- determination of properties eg: MW, 3D structure
- a) chemical/biological alteration
b) select pH, salts, solvents, surfactants
c) liquid or dried formulation - stable, active formulation
what conditions need to be decided
- physical state, pH , ionic strength, temperature: needs to be active and stable at 37 degrees Celsius in body
- stability enhanced when dry and frozen (i.e. freeze-dried) as water is removed and left with pure protein
how do additives affect proteins
- salts decrease denaturation via binding to the protein (also metal ions - calcium)
- polyalcohols (glycerol) stabilise by selective solvation of functional groups on the surface
- surfactants prevent adsorption of proteins at surfaces and aggregation
*additives need to be non-toxic
what chemical modifications can proteins have
- synthetic polymers (PEG) which makes proteins more stable in aq environments
- lipids covalently bound to proteins to make proteins more stable in non-aq environments
how does primary sequence alteration affect proteins
- specific amino acids changed which doesn’t alter final function of protein
- improved physical and chemical stability
what problems are there with parenteral route for proteins
- repeated administration
- patient compliance
- stability of dosage form (liquid is not stable - fridge)
what is the most preferable route of administration
oral as easy to administer
what is the problem with oral administration
low absorption except for a few cases:
- cyclosporin, TRH, captopril
advantages of parenteral route
- controlled drug release
- site-specific delivery
what advantages does site-specific delivery have
- Pharmacokinetic advantages
- Improvement of therapeutic index
- Protection from unwanted drug disposition
- Extravascular access (cancer-drugs: don’t want it escaping target area due to side effects)
what are the methods of site-specific delivery
- particulate systems: non-soluble, protein delivered to site
disadvantages: problems with RES as particles can’t cross vascular endothelium - soluble carriers
disadvantages: problems with stability/transport
how are implantables delivered
- IM or SC route
advantage of implantables
drug release for periods up to 1 year
types of implantable system
- Polymer gel matrix (biodegradable)
- Polymer fibre system
- Osmotic mini-pumps
- Tablet-type implants
- Automatic feedback system (may need surgery)
examples of implantable - Zoladex
- injectable biodegradable polymer matrix
- SC injection into upper abdominal wall – continuous release over 28 days
- sterile white/cream – coloured cylinder 1 mm in diameter & 5 mm long, preloaded into a special single-use syringe
- indicated for palliative treatment of advanced prostate carcinoma
example of implantable: Levonorgestrel
- polymer fibre system
- > 6 months
- for birth control
what are the 5 ways oral bioavailability can be improved
- peptidase inhibitors
- saturation/bypassing of intestinal peptidases
- penetration enhancers
- reduction of hepatic first pass clearance
- non-passive and paracellular transport
peptidase inhibitors
-improve stability 1
- eg: Amistatin promotes absorption of enkephalins
saturation/bypassing of intestinal peptidases
- increase dosage or load with another peptide
- enteric coating eg: acid-resistant acrylic resin
- use of azo polymers so drug released further down GIT
penetration enhancers
- improvement of passive absorption
- transcellular or paracellular - eg: for insulin
- ionic/non-ionic detergents
- bile salt surfactants
- EDTA and other chelating agents
- problems with irritation/tissue damage
- recently (2020), oral semaglutide (GLP synthetic hormone) for Type 2 diabetes
- sodium N-(8-[2-hydroxybenzoyl] amino) caprylate (SNAC)
- increases local pH, promotes absorption in GI epithelium
reduction of first pass clearance
- saturation of specific enzymes
- promotion of lymphatic uptake
e.g. cyclosporin bioavailability increased from 1% →20-50% when drug in olive-oil based formulation so bypassed hepatic clearance
non–passive and paracellular transport
- carrier mediated for di- and tri- peptides
- endocytic/transcytotic for proteins
- M-cell (Peyer’s patch, small intestine) but only 1000 molecules could be transported which isn’t enough
- role of P-glycoprotein efflux pump
- transport across tight junctions (paracellular)
PKs of protein drugs compared to small drugs
- dose-response relationships non-standard ie: in hormonal systems there is no improved reaction when you get a dose of hormone due to:
- multi-faceted cascade processes
- differences in biological action in males/females:
- How much?
- How often?
- Where to?
- both zero-order or complex delivery systems (non-zero order) may be used depending on situation ie: time of day/season when protein delivered changes actions of protein
limitations of SC insulin injection
- insulin SC fails to mimic endogenous insulin secretion
- lack of acceptance of multiple daily injections by patients
- complications: retinopathy, nephropathy, neuropathy
properties which promote insulin as a candidate for inhalation delivery
- relatively large hydrophilic molecule (5.6 kDa)
- transported by passive paracellular diffusion through tight junction
inhalation delivery for insulin
- 2nd gen dry powder inhalation technology for insulin delivery
- spray-dried insulin powder with stabilisers in blister
- blister loaded at the base of the inhaler and punctured by actuation
- fluidization/deaggregation in aerosolisation chamber by compressed air
- patient inhales the particle cloud through a slow deep breath
PKs of inhaled insulin
- serum concentrations peak earlier and decay more rapidly than after SC injection of regular insulin
- onset of action quicker and duration of action prolonged as compared to rapid-acting insulin analogues
- patients can inhale insulin just 10min before food but they still need a bedtime SC injection
limitations of inhaled insulin
- relative bioavailability of insulin is 15-30% vs SC injection
- fate of unabsorbed dose
non-respiratory effects of inhaled insulin
- hypoglycaemia (frequency and severity similar to SC injections) ie: it wasn’t improved
- increase in insulin antibodies (no clinical effects so far: long-term?)
respiratory effects of inhaled insulin
- cough, increased sputum
- decrease in lung function in some patients (recommended patients undergo lung tests before and periodically thereafter)
why was Exubera and Afrezza inhaled insulin a failure
they were FDA approved then removed from the market
- not good sales due to sizing of device (Exubera)
- cancer-causing warning
- lung-function concerns
what is the only medicinal form of insulin now
solution for injection (can order infusion)
