MOLECULAR DRUG TARGETS

MOLECULAR DRUG TARGETS

LEARNING OUTCOMES At the end of this session student shall be able to: List different types of druggable targets Describe forces involved in drug-receptor interactions Describe theories of drug receptor interaction Discuss the methods used to evaluate drugreceptor interaction Discuss the process of drug signal transduction

MOLECULAR DRUG TARGETS Molecular drug targets are cellular proteins that selectively bind the drug and initiate the drug s effect A drug target has two characteristics: Recognition capacity Amplification capacity: ability to initiate a response Drug targets include: Enzymes, Receptors, Ion channels and Transporters There are about 8,000 targets in human proteins Existing drugs interact only with 324 targets (266 human and 58 pathogens)

TERMINOLOGIES Ligand: a molecule (drug) that binds to a target. Pharmacophore: a fragment in the drug that enables it to bind to the receptor Binding domain: a region on a target where a ligand binds Signal transduction: the mechanism by which a message carried by the ligand is translated into a tissue response

TYPES OF LIGANDS Target Types of ligands Description Enzyme Ion channel Transporter Receptor activator competitive inhibitors non-competitive inhibitors openers inhibitors (blockers) inhibitors agonist antagonist inverse agonists bind to enzymes and increase their enzymatic activity Bind at the active site and inhibit enzyme activity Bind at allosteric site and inhibit enzyme activity bind to ion channels and allosterically open the ion channel bind to ion channels and physically block the pore or cause an allosteric change that closes the pore bind to transporters and cause allosteric changes that prevent proper functioning of the transporters attach to binding site and give a similar response to that of the endogenous ligand prevent the binding of the endogenous ligand and thus abolish the response prevent the binding of agonists, elicit a response inverse to that of agonists

DRUG-RECEPTOR INTERACTIONS Equilibrium between a drug, a receptor and a drug-receptor complex is: The biological activity of a drug depends on the stability of the D-R complex i.e. its affinity to the receptor The D-R complex can be stable if: the interaction between the drug and receptor is stronger than interaction between drug and receptor with solvent molecules. the enthalpy of interaction can compensate the entropic loss for both receptor and the drug. Thus, the Driving force for drug-receptor interaction is low energy state of drug-receptor complex The stability is measured as: K d = D [R] [D R complex] the smaller the larger the more stable the cmplex

FORCES INVOLVED IN D-R COMPLEX Bonding between a drug and a receptor occurs if there is total decrease in free energy (i.e. G is negative) G = -RTlnK eq K eq = binding equilibrium constant Small change in energy have a major effect on drug-receptor equilibrium E.g. decrease in G of ~ 5.5 kcal/mol changes binding equilibrium from 1% in drug-receptor complex to 99% in drug-receptor complex

FORCES INVOLVED IN D-R COMPLEX Interactions in drug-receptor complex may involve covalent bond or non-covalent forces such as : Hydrophobic interactions Ion-dipole and dipole-dipole Hydrogen bonding Van-der-waals interactions Charge-transfer Ionic bonds

Covalent interactions It is an irreversible link between the drug and the receptor It is the strongest bond G from -40 to -110 kcal/mol but is seldom found in drug action except with enzyme and DNA

Ionic interactions Ionic bonding: are usually reversible and their strength depends on the distance btw the two ions ( G -5 kcal/mol) Arg, Lys and His can provide cations Asp and Glu can provide anions O N H pivagabine O O H 2 N NH 2 NH

Ion-dipole and dipole-dipole interactions ion dipole interaction: a dipole in a drug is attracted by an ion in the receptor dipole dipole interaction: a dipole in a drug is attracted by a dipole in the receptor N C N HO G -1 to -7 kcal/mol zaleplon O N N dipole-dipole ion-dipole H 3 C O O - N CH 3

Hydrogen bonds Are a type of dipole-dipole interaction between H on X-H and O, N or F (X is an electronegative atom) G -3 to -5 kcal/mol intramolecular O H O O H :O H intermolecular

Charge transfer complexes Are forces btw an electron donor and an electron acceptor groups donors are usually π-electron-rich species Acceptors contain electron deficient π-orbitals G -1 to -7 kcal/mol CN Chlorothalonil fungicide Cl Cl Cl Cl + CN acceptor OH donor

Hydrophobic interactions It occurs when non-polar sections of molecules are forced together by a lack of water solubility increased entropy of water decrease the free energy and stabilize the drug-receptor complex. G -0.7 kcal/mol (per CH 2 /CH 2 interaction)

Hydrophobic Interaction O NH 2 O butamben butamben - topical anesthetic

van der Waals forces Occurs due to the formation of transient dipoles within a structure G -0.5 kcal/mol per CH 2 /CH 2 interaction

Drug receptor interaction - example charge transfer or hy drogen bond :N CH 3 CH 2 CH 2 CH 2 O hy drophobic hydrogen bond H ionic or ion-dipole N H + CH 2 CH 2 N CH 2 CH 3 O CH 2 CH 3 hy drophobic hy drophobic dipole-dipole or hy drogen bond dibucaine hy drophobic Dibucaine - local anesthetic

Means of measuring drug-receptor interactions Drug receptor interaction is measured by comparing a dose-response curve of endogenous ligand and the drug molecule. If a molecule produce the same maximal response as the ligand is called full agonist If it show no response to the receptor but block the effect of natural ligand depending on concentration of the ligand is a competitive antagonist If the effect of the antagonist is independent of the concentration of the ligand is called non-competitive antagonist. If it produced less than the maximal response is called partial agonist If it showed opposite response is an inverse agonist

% Muscle Contraction Y showed less than maximal response is a partial agonist Dose-response curve of Ach W showed same response as Ach is agonist X showed no response X blocked the response depending on conc. of Ach is a competitive antagonist X blocks the response independent on conc. of Ach is a non-competitive antagonist Z showed full response but opposite to Ach is an inverse agonist Z showed less maximal response opposite to Ach is a partial inverse agonist

Stages in drug-receptor interactions Drug receptor interaction involves two stages 1. Affinity: capacity of drug to bind to the receptor 2. Efficacy ( ): ability of drug to initiate a biological effect Affinity and efficacy are uncoupled: a compound can have great affinity but poor efficacy (and vice versa). Agonist and antagonist depends on the biological system. A compound can be an agonist for one receptor and an antagonist or inverse agonist for another receptor Equal affinities Different efficacies Equal efficacies Different affinities

Drug-Receptor Theories Occupancy theory Rate theory Induced-Fit theory Activation-Aggregation theory Multistate model

Occupancy Theory In 1926 Intensity of pharmacological effect is directly proportional to number of receptors occupied Does not rationalize why two drugs that can occupy the same receptor can act differently. (as agonists, antagonist or inverse agonists)

Rate Theory (1961) In 1961 Activation of receptors is proportional to the total number of encounters of a drug with its receptor per unit time. Does not rationalize why different types of compounds exhibit the characteristics they do.

Induced Fit Theory In 1958 Agonist induces conformational change response Antagonist does not induce conformational change - no response Partial agonist induces partial conformational change - partial response

Activation-Aggregation Theory In 1965-1967 Receptor is always in a state of dynamic equilibrium between activated form (R o ) and inactive form (T o ). R o T o biological response no biological response Agonists shift equilibrium to R o Antagonists shift equilibrium to T o Partial agonists bind to both R o and T o Binding sites in R o and T o may be different, accounting for structural differences in agonists vs. antagonists

Two-state (Multi-state) Receptor Model R and R* are in equilibrium (equilibrium constant L), which defines the basal activity of the receptor. Full agonists bind only to R* Full inverse agonists bind only to R Partial agonists bind preferentially to R* Partial inverse agonists bind preferentially to R Antagonists have equal affinities for both R and R* (no effect on basal activity) In the multi-state model there is more than one R state to account for variable agonist and inverse agonist behavior for the same receptor type.

G-PROTEIN-COUPLED RECEPTORS (GPCRs) Are integral plasma membrane proteins that transduce signals from extracellular ligands to signals in intracellular G-proteins (GTP binding proteins) GPCRs has seven transmembrane helices in a single chain of 350 1,200 residues, The amino-terminal contains N-linked glycosylation sites. Interaction with G-protein is through the third loop, and the C-terminal

Types of GPCRs Receptors About 860 genes of the human genome encode GPCRs. More than 50% of GPCRs are activated by sensory stimuli (8 by light, 22 by taste compounds and 388 to 460 by odorant stimuli). The endogenous ligands of GPCRs are small neurotransmitters, neuropeptides, peptide hormones, inflammatory mediators, lipids and ions

Types of GPCRs Receptors Endogenous ligand Example Biogenic amines Acetylcholine, Adrenaline, Dopamine, Histamine, Serotonin Peptides/Proteins Adrenocorticotrophin (ACTH, Adrenomedullin, Amylin, Angiotensin II, Bradykinin, chemokines, Gastric inhibitory peptide, Gastrin, Neuropeptide Y/W/FF, Opioids Amino acids Glutamate, GABA Lipids Leukotriene, Lysophosphatidylcholine, Platelet-activating factor, Prostacyclin, Prostaglandin, Thromboxane A Nucleotides/Nucleosides Adenosine, ADP, ATP, UDP, UTP Proteases Thrombin, Trypsin Ions Calcium

G-protein consist is complex of three units GαGβγ. At rest Gα bound to GDP When a receptor is activated: 1. GDP is converted to GTP. 2. complex dissociates to active Gα-GTP and Gβγ 3. Hydrolysis of GTP leads to reassociation

G-protein signaling pathways G-protein Gα s Gα i Gα q Gα 12/13 Gα transducin Gβγ Pathway activates adenylyl cyclases, increase camp, stimulates PKA inhibit most adenylyl cyclases activate PI3K increase IP 3 and DAG that release Ca +2 and activate PKC, respectively enhance Rho kinase Release cgmp actiavte vision and taste systems inhibit opening of Ca v channels stimulate PLC β and (PI3K). Note Signaling may take up to tens of seconds to be completed. in a few cases, such as vision using rhodopsin and transducin, the responses take only tens of milliseconds.

G-protein signaling pathways