Hit to Lead Activities 1

what does lead optimisation produce?

produce a pre-clinical candidate where it is evaluated in a complex biological system for its toxicity and drug like properties to assess if its safe to advance for clinical development

what does a drug need to have a good balance of?

• potency

• stability

• PK properties

• safety

• side effects

need to compromise to produce best balance

what are the different approaches to medicinal chemistry in drug discovery programme?

2 approaches:

• we don’t know how good a compound is until you make it

• we can utilise resources to predict enough data to ensure we make better compounds and succeed sooner

what is target validation?

verifies that the interactions with the target will impact the disease → identify biomarkers

what are the 3 ways in which a drug can be excreted?

• faeces → if drug is not absorbed into blood then excreted here

• urine → main route if drug enters blood

• exhalation

what are the key properties to consider in hit to lead activities? (8)

• aqueous solubility

• lipophilicity (log P/D)

• microsomal stability

• P450 inhibition

• cytotoxicity

• Hep G2 hepatotoxicity

• Caco-2 permeability (P_app)

• MDR1-MDCK permeability (P_app)

why is aqueous solubility and lipophilicity important to consider in hit to lead activities?

• too soluble: stays in aqueous phase

• too lipophilic: stays in fatty tissue

why is microsomal stability important to consider in hit to lead activities?

organs will try to protect against foreign exogenous substance → mainly liver that does this :. detoxifies what you’re taking

• liver = rich in microsomal enzymes → drug first exposed to liver :. defines what fraction of drug enters the systemic circulation

• assay measures compound clearance and can give you an idea of how fast it will be cleared out in vivo

why is P450 inhibition important to consider in hit to lead activities?

CYP450 = main enzyme in liver → create hooks = functional groups (phase 1 metabolism) which increase polarity :. dissolves in water :. excreted in urine

• drugs that exhibit CYP450 inhibition slows phase 1 metabolism of itself and other drugs metabolised using this enzyme

why is caco-2 permeability (P_app) important to consider in hit to lead activities?

caco 2 = intestinal epithelial cell line that forms a monolayer → used as a model to understand how well the drug crosses intestinal membrane :. assesses how well drug is absorbed from gut

why is MDR1-MDCK permeability (P_app) important to consider in hit to lead activities?

MDCK (man, die calvin klein) cells contain MDR1 gene which encodes p-glycoprotein efflux pump

• mimics ejection of drugs from cells

• can be used to predict intestinal and brain permeability

why is Hep G2 hepatotoxicity important to consider in hit to lead activities?

most drugs pass liver for metabolism → if drug is toxic to liver then this is reduced

• hep G2 = liver cells

what is the target value for aqueous solubility?

less than 100 micromolar (microM)

what is the target value for lipophilicity (logD)

0-3 → for BBB penetration then 0-2

(>3 too lipophilic)

what is the problem with lipophilic drugs?

bind to many receptors :. many off target side effects

what is the target value for microsomal stability?

less than 30 microlitres/min/mg of protein

what is the target value for CYP450 inhibition?

more than 10 micromolar (microM)

what is the target value for caco-2 permeability

more than 1 × 10^-6 cm^-1

what is the target value for hep G2 hepatoxicity?

no effect at 50 x IC₅₀ or EC₅₀

what is the target value for cytotoxicity in suitable cell line?

no effect at 50 x IC₅₀ or EC₅₀

what do hit compounds look like?

• functional groups arranged around a central core of scaffold

• small

• core will usually be flat, rigid, aromatic

why are hit compounds usually small?

during optimisation, adding substitutions will increase MW but aim is to stay beneath 500 Da :. if big, you cannot add freely

why are hit compounds usually aromatic?

• allows points for modification :. rapid generation of many analogues

• allows fine tuning of properties

• can do template hopping = when screening compounds you find a hit which is already patented, you can make modifications whilst maintaining pharmacophore :. novel intellectual property for patent

what is the criteria for hits?

• reproducible in vitro affinity/efficacy

• favourable properties (MW, pKa, solubility etc)

• chemical tractability = able to synthesise compound in sufficient quantities

• structure-activity relationship established

• patentable or strategy in place to enhance chemical novelty

• no significant toxicity alerts from compounds or known metabolites

how was imatinib developed?

imatinib = tyrosine kinase BCR-ABL inhibitor

• known protein kinase C inhibitor was taken → large number of analogues of this was prepared

• these analogues maintained the core scaffold

• x ray crystallography confirmed binding of imatinib to target :. imatinib found here

define bioavailability

fraction of unchanged dose reaching systemic circulation

• measured on a scale of 0-1

what is absorption?

entry of drugs into the bloodstream from the site of administration.

comment on the importance of free drug in plasma

• only free drug exerts pharmacological activity

• free drug is in equilibrium across the system

• drug is reversibly bound to tissues and in plasma

• only free drug will be cleared

comment on the importance of plasma protein binding in hit to lead activities. what is its target value?

high binding affects dose, half-life and may have safety implications

PPB is expressed as f_u = fraction unbound → value greater than 90% preferred

• too high limits diffusion to the site of action

what is the criteria from hit to lead phase?

• improved affinity/efficacy

• selective over related targets (greater than 100 fold)

• acceptable plasma protein binding, CYP inhibition/induction and hERG profiles

• selectivity over non-related targets

• efficacy in animal dose → dose-response relationship

• no toxicity or mutagenicity at efficacious dose

• define patient strategy

why is a balance during lead optimisation important?

• drug experiences different environment before it reaches systemic circulation

• optimising molecular properties are important to allow the drug to reach the target (but not getting renally or hepatically cleared from plasma)