L2: Lead optimisation

Question 1

what is the goal of lead optimisation? [4]

Answer 1

improve properties of lead compounds by:
- improving efficacy
- improving potency
- reduce adverse effects and toxicity profile
- is the NCR suitable for the intended route of administration? @stability, absorption an distribution
- improving ADME: onset and duration of action

Question 2

key criteria in creating a LEAD series [7]

Answer 2

1. binding/functional potency in primary assay: IC50< 100nM
2. potency in secondary assay [cell proliferation assay]: GI50<500nM [has higher threshold for secondary assay]
3. fulfills Lipinski’s rule of 5
4. in vitro ADME liabilities [half likfe>60mins]
5. synthesis in less than 10 steps
6. multiple points of modification to allow for improving and creating LEAD series
7. patentable

Question 3

steps to LEAD optimisation

Answer 3

1. chemical modifications: alter/remove functional groups using chemical synthesis and test the activity of analog [altered molecule] –> then infer role of the groups in binding and function
2. bioisosteric replacement: substitute atoms or groups of atoms in parent molecule to produce compounds w broadly similar biological properties to the parent but have structural diversity. eg. have diff size and structure to stick into diff parts of pocket, have diff charges to interact w target

Question 4

give an example of bioisosteric replacement

Answer 4

resveratrol. its a compound found in red food, but need to consume large amts to see beneficial effects observed in mice –> optimise it by replacing linker with phenol ring substitution–> formed compound 13G that became the new lead molecule

Question 5

how can we improve drug absorption? [4]

Answer 5

1. increasing H20 solubility: impt for intravenous administration and dissolution in GI tract
2. acid base properties: affects solubility in GI tract. physiological range is pKa 1.5-8
3. lipophilicity: influences ability of small molecules to pass through lipid bilayer
4. molecular size: smaller -> BETTER! shld be<500Daltons

Question 6

formula for absorption:

Answer 6

solubility x permeability

Question 7

which is the most convenient and cost-effective drug administration method?

Answer 7

oral administration. patient can consume at their own convenience.

Question 8

does form of drug affect rate of dissolution?

Answer 8

yes. tablet, capsule, suspension and solution etc. solution is fastest.

Question 9

what limits drug absorption?

Answer 9

1. Drug: lipophilicity and extent of ionisation of drug
2. External factors: what food is ingested with drug
3. Clearance: may pass through portal vein and enter liver –> metabolised

Question 10

how to ensure stability of drug?

Answer 10

observed at different pH and temperatures

Question 11

state an example in which stability and solubility plays a part in drug absorption.

Answer 11

gemcitabine. active drug impairs DNA replication and induces apoptosis. but we create a prodrug version that is not soluble in low pH [in stomach]. prodrug promotes oral-mediated absorption of gemcitabine w less toxicity. at higher pH [in intestine], lower hydrolysis of drug and increased solubility of drug

Question 12

describe what area under curve represents

Answer 12

the total amt of drug in circulation. plot drug conc in plasma against time

Question 13

absorption affects bioavailability. T/F?

Answer 13

True

Question 14

formula for bioavailabity

Answer 14

[AUC of test x dose of i.v] / [AUC of i/v x dose of test] x 100%

Question 15

how are drugs distributed in the body?

Answer 15

via blood stream. diff drug conc are attained in diff tissues/organs. a drug may be preferentially distributed to target organ or not at all.

Question 16

recap: how are drug cleared from the body?

Answer 16

via metabolism in liver [for more lipophilic drugs to be converted in polar metabolites] or excretion in kidneys [drug remains unchanged]

Question 17

drug is cleared only through liver and kidneys. T/F?

Answer 17

False. can be by perspiration etc

Question 18

recap: describe the process of metabolism.

Answer 18

Phase 1: functionalisation by CYP450 enzymes –> these enzyme changes the functional grps through monooxygenase and drug loses activity eg. Paclitaxel converted to inactive metabolite

Phase 2: conjugation
- addition of highly polar conjugates to allow for rapid excretion
eg. SN-38 [active metabolite of irinotecan] inactivated by UGTs which adds glucuronic acid

Question 19

how do we observe/know ADME properties of HIT/LEADS?

Answer 19

conduct in vitro ADME assays
1. eg. using microsome metabolism
process: add animal/human liver microsome and lead drug candidate into plate and incubate –> analyse % of parent drug remaining at diff timepoints using LC/MS, compare with control [addition of a substance that we know wont metabolise as quickly]

2. PAMPA assay [parallel artificial memb permeability assay]
   Process: drug put in donor well and have to pass through lipid memb into receiver well. measures amt of drug in donor/receiver well after certain timepoint

Question 20

after a round of lead optimisation, what shld be done?

Answer 20

perform secondary assays in vitro studies on cells isolated from disease tissue grown in monoculture and in vivo.
-we want to do as many expts to find the BEST drug possible
- types of assays: eg. gene expression, proliferation, motility for anti-cancer drug–> assay depends on function of drug
- good to conduct expts in 3D models aka organoid spheroids

Question 21

why do we have to do in vivo expts in animal based models?

Answer 21

• required to evaluate efficacy and toxicity of drug
• can validate in vivo biomarkers for drug efficacy
• allow for in vivo evaluation of PK/PD in normal and disease animal models
• can analyse histology, tissue sample [RNA/DNA/protein]+ in vivo imaging of disease progression

Question 22

human chimera mice alws have lower metabolism than human chimera mice. T/F?

Answer 22

False, may not alws be the case

Question 23

why is lead optimisation an iterative process?

Answer 23

starts with synthesis/chemical modifications –> test in vivo and in vitro [check for PK/PD, metabolism, potency etc] –> restarts cycle if not optimal