ADME of Macromolecules Flashcards
Key differences between biotechnology-
based products and chemical products
Biologics/Biopharmaceutical Products
* Molecular weight: larger, typically in
kilodaltons (kDa)
* Derived from living sources –human &
animal tissues, cells & microorganisms
* Not easily characterized and refined to
high degree of purity.
* Often called by the same name despite
modifications in one or more amino acid
residues.
* Behave more predictably and lesser
side effects.
Traditional Chemical-based drugs
* Molecular weight: typically < 1000 Da
* Can be chemically synthesized and
purified to homogeneity.
* Chemical modification usually leads
to drastic changes in activity and new
drugs for new uses.
* May have off-target effects (side
effects).
what does Biopharmaceuticals consist of?
Recombinant proteins, monoclonal antibodies, nucleic acid-based products, enz, Ab, etc
Challenges of using Biopharmaceuticals
Stability (denaturation and degradation)
immunogenicity if impure and from non-human origin
Poor distribution due to permeability of vasculatures
why does P have poor oral F?
poor Stability, poor permeability
Name 2 transport mechanism for Absorption of protein drugs after SC or IM administration
- Diffusion - Inversely related to MW/size of
proteins → limited for large proteins - Convection - not limited by MW unless the
protein molecules are enormously large and
gets entrapped in the ECM. affected by steric hinderance and charge interactions
For larger proteins (e.g. > 16-20 kDa), movement across capillary membrane
_______; absorption mostly occur via the _______________, drain into ______________
and _____________, then finally into ___________
slow, lymphatic system, lymph nodes, larger lymphatic vessels, circulatory system
For smaller proteins (e.g. < 16-20 kDa), absorption can be via both ___________
and _______________. ____________ (i.e. blood flow throughout tissue) can
influence capillary absorption
circulatory, lymphatic systems, Perfusion
Name 2 rate limiting factors that can cause changes to absorption rates of proteins
drugs after SC/IM administration
- Interstitial fluid transport rate
- Lymphatic transport rate
________________ of protein drugs via ______________ pathway of trans-capillary transportation of proteins accounts mainly
for protein drug distribution.
Passive movement; convective or diffusive
What are the key points of the Two pore model to characterize tissue level protein disposition
Key points of two pore model:
1. Endosomal space represents the porous tissue microvascular endothelium.
2. Two types of pores exist in the endosomal layer: small and large pores.
3. Fluid passes through both small and large pores and can re-circulate.
4. Passive movement of protein molecules from vascular (plasma) space to interstitial space using both small and large pores via diffusion (PS) or fluid phase convection (J). Extent of movement and distribution related to MW/size.
5. Larger proteins: have more limited distribution and display slower distribution into and out of tissues.
6. Mathematical equation utilizing a number of derived parameters to predict PK
profile of proteins of different sizes.
Metabolism of protein drugs is via _________ by _______________ in _________, ___________ and ____________
proteolysis; proteolytic enzymes (e.g. activated proteases); interstitial fluid (extracellular fluid)
present in tissues/organs; On cell surfaces; Intracellularly once protein drugs are
taken up into cells
The ____________ is a ___________________ sharing atructural similarities with _________________
Neonatal Fc receptor (FcRn), IgG Fc receptor; MHC Class I molecule
FcRn binds to _____________
IgG and serum albumin
Role of FcRn
Plays critical role in IgG homeostasis by mediating antibody recycling by
allowing intracellular trafficking of antibodies resulting in antibodies escaping
degradation in lysosomes.
* Plays a role in recycling of serum albumin (same intracellular trafficking
mechanism as antibodies) –> increase half-life of IgG and albumin
slide 18/19 ic7
Protein drugs can be eliminated via:
- Proteolytic degradation (extracellular and intracellular)
- Renal (glomerular) filtration (dominates renal excretion of protein
drugs)
Factors affecting renal excretion of proteins:
- Cut-off molecular weight of protein (proteins > ~50 kDa are not renal
eliminated due to them being too big to pass through renal glomerular
barrier). - Charge of protein (positively charged proteins have higher renal
filtration than negatively charged proteins of same size due to
negative charges on glomerular basement membrane). - Shape and rigidity of protein (affect how well proteins undergo
glomerular filtration) - Tubular reabsorption (tubular epithelium have net negative charge →
positively charged proteins get more reabsorbed)
Name 3 Common strategies to improve PK profile of protein therapeutics
- Glycosylation of proteins
- PEGylation of proteins
- Increase in size (MW) by means of fusion proteins
Glycosylation involves addition of _________ (carbohydrates) to specific amino acids in a protein
glycans
One universal strategy: _____________ of proteins increase circulation half-time by increasing size of protein or modifying binding
to glycoprotein receptors
N-linked glycosylation
Polyethylene glycol (PEG) have 2 types :
PEG with free hydroxyl at both ends and methoxylated PEG (mPEG)
with hydroxyl at one or both ends methoxylated
PEG conjugation involves Reactive functional groups of activated PEG/mPEG attached to ________
an amino group on amino acids like lysine, sulfhydryl -SH group in
cysteine or other nucleophilic groups on amino acids
2 types of configurations:
Linear or branched PEG/mPEG polymers conjugated to protein drugs →
give rise to PEGylated proteins
of different extended half-lives
PEGylation increases circulation half-lives of proteins drugs through:
- Increase in the size of conjugated protein
- Glomerular filtration of small proteins is retarded if PEG molecules with
MW 40-50 kDa is conjugated to the proteins. - Decrease elimination by proteolysis
- PEG molecules form a protective layer on surface of protein molecules,
decreasing chance for proteolytic enzymes to interact with and break down
proteins. - Decrease elimination by action of antibodies and activated immune cells
- PEG molecules form a protective layer on surface of protein molecules,
decreasing recognition by antibodies or activated immune cells (e.g.
macrophages, dendritic cells, NK cells, etc). - This is also the property that make PEGylated proteins have reduced
immunogenicity/antigenicity
______________________ fused to a therapeutic protein to utilize FcRn-mediated recycling (so as to increase half-lives of the therapeutic protein) to enhance circulation half-lives.
Fusion proteins with Fc domain of antibody or albumin used to a
therapeutic protein
Name 4 Impact of recombinant DNA (rDNA)
technology on Biopharmaceutical products
cheaper, safer, and abundant supply of
protein-based biopharmaceuticals.
- Overcomes limitations regarding source availability.
Natural sources often rare and expensive. In addition, yield can be low due
to limited amounts present in natural sources. - Allows production of safer biopharmaceuticals. E.g. eliminates transmission
of blood-borne pathogens such as HIV, Hepatitis B virus if product is
directly isolated from infected sources. - Provides an alternative way to obtain protein-based products other than
direct extraction from inappropriate source material (e.g. urine, placenta). - Since gene of interest can be synthetic, offers opportunities for scientists to
design desirable mutations to produce engineered protein-based
biopharmaceutical products that possess advantages (such as greater
clinical efficacy, greater protein stability for longer self-life, as well as
shorter/longer circulation half-life)
Recom
In recombinant protein making,
upstream processes include selection
of the _____ transfected cell that possesses
the best cell growth properties and
highest protein yield to develop a master cell line.
1
ic8 slide 9-11
In pharmaceutical industry, QC is performed to confirm _______________ of final product to pre-determined specifications.
conformance
____________ is a biologic that is almost an identical alternative version of the original biologic (called __________________) that is manufactured by a different
company.
biosimilar; innovator biologic, reference biologic
Can a biosimilar biologic be entirely
identical to the innovator biologic?
Each biological product displays variability even between different
batches of the same product due to the variability of the biological
expression system and manufacturing process. → Process of
manufacturing biopharmaceuticals (upstream and downstream
processing) has crucial influence on the nature of the final biological
product
Mabs are larger proteins with post-translational modifications →
impossible to engineer a biosimilar 100% identical to its innovator
biologic; produce a highly similar biosimilar instead
