Drug Targeting and Resistance

Question 1

What is drug targeting?

Answer 1

Is ensuring that your inhibitor has a very high affinity for the target (enzyme)

Question 2

What is drug resistance?

Answer 2

Is the result of mutations in the target (enzyme)

Question 3

What are some factors that need to be considered when considering if your drug will reach the target cell?

Answer 3

Oral or parenteral administration
Metabolism by liver, plasma enzymes etc. (is it stable?)
Excretion in the kidneys
Segregation in specific tissues, binding to plasma proteins
Blood-brain barrier; placenta; blood-testes barrier etc.

Question 4

What are some blockages that prevent the drug entering the target cell?

Answer 4

Lipophillic or hydrophilic
Transporters, channels
Multidrug resistance proteins (ABC transporters)

Question 5

What are the ways to cross the plasma membrane?

Answer 5

Diffusion
Facilitated diffusion
Active transport

Question 6

What molecules can get across by diffusion?

Answer 6

Highly lipophilic molecules only (not water soluble)
Not specific: Enters any cell
Wide distribution may mean higher dosage and toxicity
High lipophilicity also means increased penetration across blood brain barrier and similar
Absorption through skill may be possible

Question 7

What is an example of molecules that can cross through diffusion?

Answer 7

Example: steroids

Question 8

What is facilitated diffusion mediated by?

Answer 8

Transport proteins

Question 9

What compounds usually pass through in facilitated diffusion?

Answer 9

Hydrophobic compounds

Question 10

What is specific targeting?

Answer 10

Enters only cells expressing appropriate

Question 11

What is equilibrative?

Answer 11

Allows the passage across the plasma membrane until there is equilibrium between intracellular and extracellular concentrations

Question 12

What can intracellular concentration never be higher then in facilitated diffusion?

Answer 12

Plasma concentration

Question 13

What is active transport mediated by?

Answer 13

Transport proteins

Question 14

What is active transport depend on?

Answer 14

Energy dependent

Question 15

What is concentrative?

Answer 15

Continues to ‘pump’ the substrate into the cell even against high concentration gradient

Question 16

How can intracellular concentration differ to plasma concentration in active transport?

Answer 16

Can grossly exceed plasma concentration

Question 17

What does the increased intracellular concentration increase?

Answer 17

Specifity and keep required plasma concentrations low

Question 18

What are transporters characteristics?

Answer 18

Highly specific
Concentrative if energy dependent
Equilibrative if not energy dependent
Cause of resistance if lost

Question 19

What do transporters define?

Answer 19

What molecules reach the intracellular target
What concentrations are achievable within the parasite/target cell
How fast the drug is taken up
Drug resistance and cross-resistance

Question 20

What is active transport?

Answer 20

The combination of high affinity and high rates of uptake requires energy

Question 21

What direction is active transport?

Answer 21

Uni-directional and accumulates substrates in the cell above the gradient

Question 22

What is an example of active transport?

Answer 22

Pentamidine given at 1 nanometer accumulates to mM inside trypanosomes

Question 23

What are the 3 types of active transport?

Answer 23

ATP dependent
Using sodium gradient
Using proton-motive force (H+ gradient)

Question 24

What is ATP dependent active transport dependent on?

Answer 24

Ion pumps
P-glycoproteins
Multi-drug resistance (mdr) proteins

Question 25

What is sodium gradient active transport dependent on?

Answer 25

High affinity mammalian transporters | Usually expressed in specialised tissues

Question 26

What is proton-motive force active transport dependent on?

Answer 26

Most protozoan transporters

Question 27

What does the BBB consist of?

Answer 27

BBB consists of brain capillary endothelial cells, connected by tight junctions, plus astrocytes

Question 28

What does the composition of the BBB do?

Answer 28

Keeps many unwanted substance and pathogens out but essential nutrients still need to be taken into the brain: specific transporters

Question 29

What are transporters in the BBB?

Answer 29

Mostly active and uni-directional

Question 30

What do efflux pumps do in the BBB?

Answer 30

Severely limit the ingress of drugs

Question 31

What are the most potential causes of drug resistance?

Answer 31

``` Mutation of the target protein Loss of the target Overexpression of the target Overexpression of pathway bypassing target Loss of enzyme activating prodrug Detoxification with elevated levels of glutathione etc. Expression of ABC transporters Loss of transporters ```

Question 32

What does expression of ABC transporters rely on?

Answer 32

MDR | P-glycopoprotein

Question 33

How can loss of transporters occur?

Answer 33

Mutations Gene deletions Loss of expression

Question 34

What is an example of mutation of target?

Answer 34

Benzimidazoles as anthelminthics Most widely used anthelmintics Thiabendazole (1961). Later analogues reduced toxicity Widely used in human and veterinary medicine Effective against most intestinal and systemic nematodes (roundworms) and cestodes (tapeworms) and some trematodes (flukes)

Question 35

How does this mutation occur?

Answer 35

High affinity, but irreversible binding to beta-tubulin This cases ultra structural changes to intestinal cells of nematodes and to the tegument of cestodes Disruption of cytoskeleton Inhibition of microtubule-mediated vesicular transport

Question 36

What are microtubules?

Answer 36

13 proto-filament strands

Question 37

What are proto-filaments?

Answer 37

Made up of strings of alpha-beta hetero-dimers

Question 38

What does binding to beta-tubulin prevent?

Answer 38

Tubule elongation

Question 39

What is normal tubule elongation?

Answer 39

alphabeta: alphabeta:alphabeta etc

Question 40

What does benzimidazole treatment lead to?

Answer 40

Tubule capping and degredation

Question 41

What do macrolides inhibit?

Answer 41

Including erythrocmycin | Inhibit protein synthesis by binding to 23 S ribosomal subunit

Question 42

How is resistance associated by alterations to target?

Answer 42

Methylation of rRNA | Plasmid-encoded methyl transferase

Question 43

How is resistance by reduced cellular concentration?

Answer 43

Plasmid-encoded efflux pump

Question 44

What is arsenical resistance in trypanosomes?

Answer 44

Tyrpanosoma brucei spp cause sleeping sickness Are transmitted by the tsetse fly Sleeping sickness is treated with the organo-arsenic compound melarsoprol

Question 45

What can melarsoprol resistance be the result of?

Answer 45

Detoxificarion plus-extrusion Loss of uptake into parasite Combination of both

Question 46

What does Melarsoprol (MelB) enter the parasite through?

Answer 46

P2 adenosine transporter

Question 47

What does MelB form?

Answer 47

Adduct with trypanothione (MelT)

Question 48

What is the adduct exported by?

Answer 48

TbMRPA (ABC transporter)

Question 49

What is the most common mechanism?

Answer 49

ABC-transporter-mediated efflux

Question 50

What is non-expression of (uptake) transporters?

Answer 50

Unlikely to be result of loss-of-function mutations Heterologous population followed by selection → survival of non-expressors → relapse with resistant cancer Individual differences between people