lead generation

active

A compound showing an effect in a biological screen, comprising of an assay designed to detect compounds that interact with the drug target.

hit

A compound showing an effect in a biological screen by the desired mechanism.

lead

A compound identified as a hit, which can be optimised by chemical modification to produce a drug candidate.

Bioavailability

the proportion of administered drug that reaches the circulation and can have its pharmacological effect

What are the criteria for a hit compound to reach the status of lead?

• show required biological activity

• be amenable to structural modification

• not be extremely polar or lipophilic

• no toxic groups

• interact reversibly with biological target

Does the lead compound need to be potent or selective?

• no

• chemical modification should be able to improve both properties

What are the problems associated with polar or lipophilic compounds?

• prevent uptake to the body and cause problems with bioavailability

• polarity is not easily reduced

What is polarity and lipid solubility measured by?

chemical assay

Chemical considerations for testing of lead properties

• Ease of synthesis.

• Amenability of the molecule to chemical modification.

• Solubility

• Polarity/ lipid solubility

• Structural elements likely to cause toxicity

• similarity to competitor compounds - ability to patent

which structural agents are likely to cause toxicity?

alkylating agents, ketone groups, aromatic amines, planar 3-ring aromatic systems

drug likeness

• if a compound can reach its site of action when administered via intended route

• defined by lipinskis rule of 5

Lipinsk’is rule of 5?

• Contain < 5 hydrogen bond donors (NH and OH bonds).

• Contain <10 hydrogen bond acceptors (N and O atoms).

• Have molecular mass < 500 Daltons.

• Have logP < 5.

What is the lipinskis rule of 5 for acetlsalyicylic acid and codeine?

• acetylsalicylic acid - pass

• codeine - pass

where do leads come from

• company compound collections

• academic compound collections

• natural products

• new synthesis

• antibodies and other biologics

company compound libraries

• reflect history of company - identified targets

• limited in structural diversity

• many will be lacking in drug likeness

company compound collaboration

• with academic and commerical

• may reduce limitations with diversity

which early drugs were derived from plants and are known as botanicals?

aspirin and digoxin

Examples of microbials

• penicillin

• botox

Muscarine

• selective agonist to muscarinic receptors

• found in some mushrooms

Which animals are a rich source of potential leads?

• venomous

• toxins from snake, spider, scorpion, amphibian venom and a range of marine animals

Why is venom a rich source of potential leads?

• proteins that produce poisonous effect have very specific interactions with drug targets

Explain what is meant by combinatorial chemistry and how it is used in lead generation

• mechanism for producing large numbers of new compounds

• uses isolated enzymes and cells with structurally related compounds

• identification of new hits and optimisation of leads

• mix and split synthesis, multiple parallel synthesis

mix and split synthesis

• if hit identified need to see which compound it was

• A components are mixed, B is added to each split to mix and generate a pool

  • small combinations identified

  • assay used

  • then can be tested individually and identified

multiple parallel synthesis

• used for lead optimisation

• dependent on automated methods, robots, due to large scale

• rectangle plate often used

• each well has a unique combination of A and B molecules

Which products have more diversity? Give an example

• natural products

• unexpected and predictable structures

• eg protein based glue secreted by australian frog is stronger than available medical glues

What are the cons of natural products?

• lack of consistency in same species - seasonal, environmental

• rarity of origin of species

• hard to scale up to industrial

natural products for industrial production

• hard to scale up to industrial quantities

• aim is to identify the structure of the active molecule and generate it industrially using synthetic chemistry or biotechnology

approaches to lead generation

• rational design

• empirical screening

• high thoroughput screening

rational design relies on what

• relies on available knowledge of target structure

  • endogenous ligand for the target or synthetic compound which interacts with the target

in silico rational design example

• in silico studies may be able to suggest structural components likely to interact with the target

• ACE inhibitor captopril was the first drug discovered as a result of knowledge of the target structure

modification of natural substrate example for rational design

• target based assay for analogues of ligands

• bethanechol - synthetic analogue of acetylcholine

• used to treat bladder dysfunction

• muscarinic agonist resistant to metabolism from acetylcholinesterase

fast follower approach

modify a known compound with activity of interest

fast follower approach example

1. nifedipine - calcium antagonist for treating hypertension

2. template to generate analogues for screening new calcium antagonists such as amlodipine with a longer duration of action

When is rational design chosen? What information is required?

One or more of:

• Structural information.

• Low molecular weight starting molecule(s).

• Mechanistic information.

• Target class analogies.

What are the general steps for rational design?

• information from computational chemistry and structural biology is required for molecular modelling in silico

• chemical ideas will already exist if ligand knowledge is known

• may be synthesised, screened and then reviewed

  • if there is a lead compound found, optimisation

  • if not producing hits, compounds may be further modified and tested until lead is identified

when is empirical screening the only option available?

• if there is no information about the structure of target and there are no known ligand structures

advantage of using empirical screening when there is structural information available

• unpredicted structures can produce hits

• information on novel structure activity relationships

• increase chances of patentable discovery

What does empirical screening involve?

• screening any libraries of synthetic or natural compounds

• no assumptions about what type of structure might have activity @ the target

What is the main benefit of empirical screening?

hits can be identified from low molecular weight compounds even when nothing is known about the structure of the target

What does success of empirical screening depend on?

success depends on good compound libraries

How is empirical screening performed for no structural activity ?

• high thoroughput screening, over 1 million tested

• initial hits

• synthesis, testing and review until lead is ideintified for lead optimisation

high thoroughput screening

• co-ordinated series of processes to test large numbers of compounds quickly and efficiently against a drug target

  • robots for most tests

What are the key requirements for thoroughput screening?

• Large numbers of diverse compounds to test.

• Efficient methods to deliver compounds to the screen.

• Robust, reliable, and cost-effective assay technologies.

• Effective automation.

• Efficient and effective data processing and information management systems.

Steps for designing screening cascade

1. Decide what assay or screening strategy to use, e.g. rational or empirical design

2. Set up small scale assays, test and validate. Demonstrate that the assay detects the appropriate biological activity. Establish proof that the assay measures what is required, it is reproducible, reliable, and robust.

3. Develop and optimise the final method. This involves scaling up the assay and optimising it for automated use in a high throughput system.

4. Validate the method as 'fit for purpose'.

For high thoroughput screen to be validated the following criteria must be met:

1. activity measured must be reproducible between wells within a multi-well plate and between plates.

2. activity detected by an assay must be reproducible when the assay is repeated on different days,

3. Reproducibility must be maintained at the level of throughput that screening will employ

4. There must be correlation between test runs.

5. The long-term reliability of the screen

6. The screen must be be robust as indicated by both the variation in the measurements between assay runs, and the dynamic range of the signal measured.

dynamic range

• ratio of signal to the variation in the background signal above which the signal is measured

• the ability of screening assays and biophysical methods to accurately detect and quantify compound activity across a broad range of potencies