Drug design: finding a lead

Question 1

What is the lead candidate?

Answer 1

A compound demonstrating a property likely to be therapeutically useful
• Used as the starting point for drug design and development

Question 2

How is the lead candidate found?

Answer 2

• Found by design (molecular modelling, X-ray crystallography or NMR) or by screening existing compounds (natural or synthetic)
• Need to design a suitable test in order to identify a lead compound

Question 3

What is teprotide?

Answer 3

• Teprotide is a peptide which was isolated from the venom of the Brazilian viper and was one of the lead compounds for the development of the antihypertensive agent captopril
• Teprotide – o linked peptide, drug lead to develop captopril

Question 4

What is Atracurium?

Answer 4

• Atracurium is used during surgical procedure or mechanical ventilation
• Used by native Americans for hunting
• Some structural similarity – tertiary amines, aromatic rings

Question 5

What are endogenous compounds?

Answer 5

• Natural ligands for receptors e.g. adrenaline - drug lead to give idea how to produce propranolol and histamine - triggers inflammatory response in body

Question 6

What are peptides?

Answer 6

protease inhibitors (natural substrates for enzymes)

Question 7

What is saquinavir?

Answer 7

An antiretroviral drug used in HIV therapy

Question 8

Where can the knowledge about the structure of enzyme or receptor binding/active site be obtained?

Answer 8

 X-ray crystallography (preferably with the ligand bound to this binding site)
 NMR spectroscopy (allows to determine the protein structures in solutions)
 A computer-generated model of the target proteins based on the known structure of an analogous protein
• Structure can be found experimentally

Question 9

What is the strategy for carrying out drug design?

Answer 9

• Carry out drug design based on the identifies interactions between the lead compound and the target binding site

Question 10

What is step 1 of structure-based drug design?

Answer 10

Step 1 (X-ray based)
•Crystallise target protein with bound ligand
•Acquire structure by X-ray crystallography

Step 1 (NMR based)
•Alternatively, prepare an NMR sample of protein with bound ligand
•Record high-resolution 2D NMR spectra
•Identify NMR signals and any inter-nuclear interactions to reconstruct the 3D structure

Question 11

What is step 2 of structure-based drug design?

Answer 11

• Download the structure to computer for molecular modelling studies
• Identify where the ligand is, and thus identify the binding site
• Identify the binding interactions between ligand and target by measuring the distances between the ligand and neighbouring atoms in the binding site
• If Ligand has strong interactions = strong ability to inhibit enzyme

Question 12

What is step 3 of structure-based drug design?

Answer 12

• Identify vacant regions for extra binding interactions
• Remove the ligand from the binding site in silico
• Using receptor-based Virtual Screening, ‘fit’ analogues into the binding site in silico to test their binding capabilities
• Alternative approach involves rational de novo design of lead molecules
• Removing ligand from active centre

Question 13

What is de novo?

Answer 13

Designing own ligand from scratch

Question 14

What is receptor-based virtual screening?

Answer 14

• Involves docking of ligands from a large structural database into the active (or allosteric) site of the target provided that the 3D structure of the target molecule is known either from X-ray crystallography or NMR analysis

Question 15

What does virtual screening help to asses?

Answer 15

Helps to assess whether known compounds from a large database are likely to be lead candidates for a particular target
If you know 3D structure - then you can dock a huge number of different small molecule ligand compounds onto active centre and programme calculates binding energy etc and therefore can decide which one has strongest interactions

Question 16

Illustration of de novo design?

Answer 16

Active centre with functional groups present – crucial for binding with a ligand/drug lead
Link them with flexible linkers so ligand can be accommodative and to make sure reactions are optimal

Question 17

What is step 4 of structure-based drug design?

Answer 17

• Identify the most promising analogues
• Synthesise and test for activity
• Crystallise a promising analogue with the target protein
• Alternatively, prepare a sample of a selected ligand with the target protein and record high-resolution 2D NMR spectra
• REPEAT THE WHOLE PROCESS

Question 18

What is the strategy for designing lead compounds using NMR spectrometry?

Answer 18

• Based on identification of small molecules (fragments), which will bind to specific but different regions of a protein binding site
• The separate fragment structures bind to only one part of the binding site
• Thus, they have no activity themselves against the target
• However, linking the two separate fragments with an appropriate spacer may lead to a larger molecule that binds to the whole binding site
• This may considerably increase the binding affinity (and thus activity) of the resulting lead compound towards the protein active site

Question 19

What is the procedure for designing lead compounds using NRM?

Answer 19

• A range of low-molecular weight compounds is screened against the target
• Binding can be detected by observing a shift in the 15N or 13C NMR signals
• This shift can be induced only by the interaction of the fragment with the target
• This will also reveal which part of the binding site is occupied by the fragment
• Once two fragments have been identified, the structure of each can be further optimised to enhance binding affinity
• Then the new ligand can be designed where the two fragments are linked together
• This approach offers a high level of structural diversity for drug leads
• Optimise by synthesising
• Very cost efficient

Question 20

NMR spectra

Answer 20

• If fragment binds – some signals shift because they’re interacting
• Record spectrum FIRST then bind the ligand and record spectra again to see change in chemical shift