Section 6

Question 1

What do candidate drug target profiles (CDTPs) consist of?

Answer 1

Values which a molecule will need to reach in order to progress onto the next stage. These include:

Biology (IC50, target selectivity, hERG receptor affinity)

DMPK (clearance, oral bioavailability, plasma protein binding, P450 inhibition IC50, dose to man prediction)

Physicochemistry (molecular weight, cLogP)

Question 2

What is an in vivo oral gavage?

Answer 2

Investigation to determine the ADME of a drug. Drug is fed directly into the stomach of a mouse via gastric tube attached to a syringe and effects are investigated using turnover rate of substrate to product.

Question 3

What do selectivity wheel/plots show?

Answer 3

The level of inhibition (%) of a compound at various receptors/enzymes. Easily shows how effective the drug is at the drug target and its selectivity based on activity at off-target receptors.

Question 4

What is the ideal logP of drugs for oral bioavailability?

Answer 4

2-4

Question 5

What is the ideal logP of drugs for BBB penetration?

Answer 5

3-5

Question 6

What are the pros and cons of a high logP?

Answer 6

Pros:
Increased binding to enzyme/receptor
Increased absorption/permeability through membrane.

Cons:
Decreased aqueous solubility.
Increased binding to P450 metabolising enzymes.
Increased binding to blood/tissue proteins so less free drug to act.
Increased binding to hERG heart ion channel (cardiotoxicity).
More compartmentilisation into lipid membranes.

Question 7

Why do drugs need to be soluble?

Answer 7

Requirement to pass through numerous aqueous compartments means acceptable solubility in this medium is critical.

Question 8

Why is the balance between a drug’s solubility and lipophilicity important?

Answer 8

To reach the desired molecular target a drug molecule needs to be lipophilic enough to permeate a cell membrane but still hydrophilic enough to be soluble.

Question 9

Describe the parallel artificial membrane permeability assay (PAMPA).

Answer 9

3 well plates are prepared:
A donor well plate filled with buffer solution and test compound of known concentration with artificial membranes at the bottom.
An acceptor well plate underneath containing buffer solution. A lid covering the other plates.
The test compound diffuses down its concentratoin gradient from the donor to the acceptor plate. Regular samples are taken from the acceptor plate and the concentration is measured to calculate the rate of diffusion.

Question 10

Describe the Caco-2 assay.

Answer 10

Measures the rate of transit of a drug through a monolayer of cells.
An apical chamber with a permeable membrane and cell monolayer base is filled with buffer solution to represent the GIT. This is placed inside a larger basolateral chamber which is filled with buffer solution to represent the blood.

The drug can be added to either side to measure apical–>basolateral or basolateral–>apical transport.

The measurements can be used to calculate the apparent permeability coefficient (Papp, cm/s)

Question 11

Describe an in vitro assay which can be used to predict the in vivo metabolic stability of a drug.

Answer 11

Obtain liver microsimes (vesicles obtained from hepatocyte endoplasmuc reticulum which contain membrane phase I and II enzymes) from the liver using a series of 3 homogenisation and centrifugation steps to collect a pellet of microsomes.

Incubate the compound with the microsimes and measure the turnover rate.

Alternatively use whole hepatocytes.

Question 12

How is oral bioavailability measured?

Answer 12

(AUC(po)/AUC(iv)) x 100

basically difference of drug concentration in blood after IV and oral dose over time.

Question 13

What is ligand efficiency (LE)?

Answer 13

LE = -deltaG/HAC
(HAC = heavy atom count).

It is the free energy of binding of a ligand, averaged for each non-hydrogen (or heavy atom) in the molecule.
It represents the ratio of molecular weight to affinity.

A higher LE means higher efficiency.

Question 14

What are the 2 calculations for Gibbs free energy?

Answer 14

ΔG= -RTlnKd

ΔG=change in Gibbs Free Energy
R= gas constant = 8.314
T= temperature in Kelvin
lnKd = natural log of equilibrium dissociation constant.

ΔG=ΔH - TΔS

ΔG=change in Gibbs Free Energy
ΔH=change in enthalpy
T = temperature in Kelvin
ΔS = change in entropy

Question 15

What is lipophilic ligand efficiency?

Answer 15

LLE = pIC50 - LogP

Indicates how the lipophilicty of a compound contributes to its potency.

The higher the LLP, the better. Usually 5-7 is deemed good.
If a compound’s LLE is lower than this it is too lipophilic.

Question 16

How was the reverse transcriptase inhibitor Lersivirine developed?

Answer 16

The initial lead had an LLE of 1.9 (too low = too lipophilic) so the lipophilicty was reduced by:
Swapping lipophilic Cl groups for cyano groups (CN)
Replacing the benzylic methylene for an oxygen atom to create an ether linkage between the phenyl ring and the pyrazole.

This structure was Lersivirine, which had a LLE of 4.9.

Question 17

What is a rotatable bond?

Answer 17

Any single non-ring bond, attached to a non-terminal, non-hydrogen atom.
(Not including C-N bonds as resonance prevents rotation)

Question 18

What is the relationship between the number of rotatable bonds and bioavailability? Why?

Answer 18

Higher number of rotatable bonds = lower bioaviailability.

This is because when a molecule passes through a lipid membrane from the aqueous extracellular environment, it partitions into the densley packed lipid envelope which restricts its freedom of movement, thus forcing it to adopt a fixed conformation.
This reduces entropy, so if the molecule is more rigid to start with (less rotatable bonds) there is a less significant decrease in entropy (ΔS). Since ΔG = ΔH – TΔS, this means ΔG is less positive and the impact on ΔG is less (so process is more favourable).

Question 19

What is a solvation shell?

Answer 19

A shell of water around a drug molecule, with H bonds joining the molecule to the water and the water to each other.
The water molecules in this solvation shell have a lower entropy than the free water around it.

Question 20

What energy changes occur when a dissolved drug moves from the aqueous extracellular environment into a membrane.

Answer 20

1. 2 water molecules move away from solvation layer so the drug can access the phospholipid head groups. In doing so, this breaks the H-bonds between the drug and themselves, which increases entropy (disorder).
2. Drug molecule diffuses towards lipid membrane. The 2 previous water molecules form H bonds with other molecules in the solvation shell, which decreases entropy.
3. Interactions between drug and membrane increase in strength, forcing additional bonds between the drug and water molecules in the shell to break, increasing entropy.
4. Drug partitions into membrane and engages in hydrophobic/Van der Waals interactions with the lipid interior (more negative enthalpy), overcoming any remaining bonds between the drug and solvation shell, increasing entropy.

ΔG = ΔH – TΔS
So a lower H and a higher S mean more negative Gibbs free energy, which is favourable.

Question 21

How does LogP contribute to the Gibbs free energy for partitioning and bioavailability?

Answer 21

Low logP = high HBAs and HBDs, low lipophilicty, and high water solubility. This means bioavailability is low as large +ve ΔH required to break H-bonds meaning ΔG for partitioning between aqueous and lipid compartments is more positive which is not favourable.

High logP = low HBAs and HBDs, high lipophilicty, and low water solubility.
This means bioavailability is low as less H-bonds between drug and water mean that less are broken so change on entropy (disorder) is minimal. Therefore Gibbs free energy is less negative and less favourable,

Question 22

What are the 2 key factors affecting aqueous solubility of a drug molecule?

Answer 22

Polarity of the drug molecule.

Crystal lattice energy of the solid crystal form.

Question 23

What are the 2 main ways to increase solubility of a molecule?

Answer 23

Introduce new polar functional groups such as alcohols, amines, acids to areas of the molecule which aren’t required for target binding.

Disrupt crystal packing.
Crystal packing is the ordered arrangement of the same molecules held together by covalent or non-covalent bonds. They make dissolution difficult as it requires the energy required to break a molecule free from the crystal lattice to be less than the energy released from the new interactions between the water and drug molecules. To make dissolution easier, the formation of a crystal lattice can be prevented by removing or replacing atoms necessary for the intermolecular bonds.

Question 24

What are some ways to reduce rapid metabolism?

Answer 24

Reduce lipophilicity of molecule or just area susceptible to metabolism.

Invert stereochemistry if possible.

Modify steric environment around area susceptible to metabolism.

Alter electronic characteristics.

Introduce conformational constraints.

Question 25

How was the neurokinase-1 antagonist Aprepitant developed?

Answer 25

Lead compound was found to possess a benzyl ring linkage which was prone to CYP450 oxidation to an ester and subsequent hydrolysis, as well as a benzene which was prone to oxidation to phenol and Phase II metabolism.

To reduce this, a methyl group was added to the linkage and a fluorine was added to the para position of the benzene. The resulting molecule was Aprepitant.

Question 26

How was Tetrabenazine modified to reduce metabolism?

Answer 26

Usually, O-dealkylation occurred at the two methoxy (OCH3) groups.
The Hs on the methyl were swapped for hydrogen deuteriums (hydrogen isotopes with 1 proton, 1 neutron and 1 electron, instead of just 1 proton and 1 electron). This new molecule, deutrabenazine, had reduced the O-dealkylation.

Question 27

What are bioisosteres?

Answer 27

Compounds or groups that possess near-equal molecular shapes and volumes, approximately the same distribution of electrons and which exhibit similar physical properties.

Question 28

What are bioisosteres used for?

Answer 28

To replace functional groups in order to change a molecule’s physicochemical properties but maintain activity.

Question 29

What is the difference between classical and nonclassical bioisosteres?

Answer 29

Classical bioisosteres are atoms, ions, and molecules which possess the same number of bonds and/or peripheral electron layers.

Non-classical bioisosteres do not necessarily possess the same number of atoms or the same number of valance electrons, yet still result in similar pharmacological activity.

Question 30

What are privileged structures?

Answer 30

Structural motifs that are capable of binding to a variety of pharmacological targets.

Question 31

Given an example of a privileged structure?

Answer 31

Benzodiazepine core structure also appears in that of Devazepide (an appetite suppressant) and has been found to bind to several GPCRs and ion channels.

Question 32

Why effects do fluorine have on drug properties?

Answer 32

Very strongly electronegative (influences pKa).

Hydrogen bond acceptor (can improve affinity as can make additional interactions with binding pocket without changing steric environment).

Exerts electronic effects on neighbouring acid/base moieties which tune and optimise their acidity/basicity.

Increased lipophilicity (improves permeability and brain penetratoin).

Improves metabolic stability (swapping H for F can prevent metabolism).

Deactivation of oxidative metabolism of aromatic rings (due to electron-withdrawing nature).

Can easily be convert a molecule into an imaging agent for PET if fluorine is replaced by the radioactive isotope fluorine 18.

Question 33

What are Veber’s rules for oral bioavailability?

Answer 33

A compound should have no more than 10 rotatable bonds.

The hydrogen-bond count (donors + acceptors) should be no more than 12, or polar surface area (PSA) should be no more than 140A.