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Advanced Medicinal Chemistry Barrie Martin AstraZeneca R&D Charnwood Lecture 5: Drug Metabolism and Pharmokinetics - 2.

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Presentation on theme: "Advanced Medicinal Chemistry Barrie Martin AstraZeneca R&D Charnwood Lecture 5: Drug Metabolism and Pharmokinetics - 2."— Presentation transcript:

1 Advanced Medicinal Chemistry Barrie Martin AstraZeneca R&D Charnwood Lecture 5: Drug Metabolism and Pharmokinetics - 2

2 Quantitative DMPK Quantitative DMPK involves the measurement of a number of pharmacokinetic parameters which describe the fate of compounds in the body. These can be used to compare compounds, to highlight deficiencies in compounds (e.g. high metabolism) and to predict how the potential drug will behave in man – generate dose predictions.

3 The One Compartment Model The simplest model to describe the fate of a compound in the body is the ‘one compartment model’, which is analogous to the metabolism of the compound in a beaker containing an enzyme solution. Although simplistic, many of the basic parameters of pharmacokinetics (half-life, clearance and volume of distribution) are well illustrated using this model. BLOOD Injection

4 Half-Life (T 1/2 ) and the Elimination Rate Constant (k el ) Plasma conc Time x x x x x x C = C o e -k el t Time ln c ln c = ln c o - k el t At,t1t1 / 2 c coco = 1 2, ln (0.5) = - k el t1t1 / 2 t1t1 / k el = x x x x x x Following an iv injection, we might expect the plasma concentration of a compound to vary over time as shown below: Under first order kinetics, rate of metabolism is proportional to the compound concentration, so the rate (and gradient) decreases over time. By plotting ln c vs. time, we can determine the elimination rate constant (k el ) from the slope of the line and, by extrapolation back to t = 0, the initial plasma concentration c 0. The main use of k el is to determine the half-life (t 1/2 ) of the compound, defined as: The time taken for the concentration of drug in the blood or plasma to decline to half of its original value. - k el = slope

5 Volume of Distribution (V D ) In biological systems, compounds can distribute out of the plasma into tissues. The volume of distribution (V D ) is therefore: The theoretical volume (L) that all the drug in the body would have to occupy if it were present at the same concentration as that found in plasma. It is a measure of how readily drug diffuses out of the plasma into the tissues and can affect t 1/2. –Low V D : drug confined to plasma (vulnerable to the liver and metabolism) –High V D : drug equilibrates with tissues Generally: Acids - high PPB, low V D Neutrals - ready equilibration, but not necessarily retained in the tissue - higher V D Bases - high affinity for phospholipids in membranes (negatively charged) - highest V D When the volume doubles (red line), the initial concentration halves and the half life doubles Time ln c Consider now the case where a compound is dissolved in double the volume, what would happen to t 1/2 ?

6 Clearance (Cl) Clearance (Cl) is a measure of how readily compounds are eliminated (i.e. metabolised or excreted) It is defined as: The volume of plasma (or blood) from which all drug is removed per unit time. (n.b. units of Cl are units of flow ml/min) Cl is a constant, characteristic of a drug in a particular species. It is a scaling factor that relates the plasma concentration of a compound to the rate of elimination Rate of elimination (ng/min) = Clearance x concentration (ng/ml) dD dt ( ) Clearance (ml/min) = Rate of elimination (ng/min) concentration (ng/ml)

7 Plasma Conc Time dt Conc Clearance (Cl) Over a small interval of time, dt: Amount eliminated during interval dt = Cl x Conc x dt Integrating this over the whole concentration-time profile gives: Total amount eliminated = Dose = Cl x AUC Cl = Dose/AUC k el = Conc. x Cl Clearance is related to t 1/2 and V D. If Cl halves, then the half life doubles (because the rate of metabolism halves) and, as we have seen, doubling the volume doubles the half-life (because the concentration at the metabolising enzyme has halved). The precise equation is: t 1/2 = x V D /Cl

8 Half-life is not predictable from clearance alone - clearance characterises elimination of drug from plasma/blood - half-life also depends on distribution of drug outside the plasma (systemic circulation not a closed system) For a rapid (bolus) iv dose; where D is the amount of drug in the body at time t; Clearance, Volume and Half- Life Rate ofh elimination, dt dD = k el D = Clearance x concentration But D = V D x concentration, so k el x V D = Cl Rem: = k el = x V D Cl, NB: t ½ is NOT a measure of how rapidly the drug is metabolised or k el = VDVD Cl t 1/2

9 For the three main elimination processes, metabolism, renal, and biliary, clearances are additive, i.e. Cl T = Cl M + Cl R + Cl B Can relate clearance to liver blood flow (Q): Cl = E x Q (E = extraction ratio) If Cl ~ Q (i.e. E~1), then 1 st pass metabolism is a likely problem. Q: Rat ~ 70 ml/min/kg Dog~ 40 ml/min/kg Man~ 20 ml/min/kg e.g. If Cl T = 20ml/min/Kg and %dose as parent in urine = 20% Then Cl R = 4ml/min/Kg Q = flow (ml/min) C in = conc. entering liver C out = conc. leaving liver E = (C in –C out ) / C in Fraction absorbed (f a ) can now be worked out if know %F Clearance, Extraction and Absorption

10 The Two Compartment Model The two compartment model more accurately describes observed DMPK data. In this model, the compound is viewed as being able to equilibrate with a second compartment as, in addition to metabolism, drug is distributing into the tissues. Drug accumulates in the tissues because the plasma concentration is initially greater than the tissue concentration and so k 12 >k 21. However, eventually the plasma concentration falls to such an extent that the net drug movement is from tissues back into blood. At this point in time, plasma concentration begins to fall far more slowly as diffusion from tissues back into blood becomes more and more significant. BLOODTISSUES Plasma conc Time x x x x x x ln c x x x x x x k 12 k 21 Injection Distribution phase Elimination phase c p = c 1 e -k1t + c 2 e -k2t

11 Oral Dosing - Bioavailability Time iv oral Plasma conc Upon oral dosing of a drug, there is an initial increase in the systemic concentration of the drug, as it is absorbed from the gut. As absorption is completed and the compound is eliminated from the body, the concentration of drug decreases over time. Absorption phaseElimination phase Oral Bioavailability (F%) is defined as: The fraction of the dose which makes it to the systemic circulation (i.e. survives 1 st pass metabolism). F = AUC after an oral dose AUC after an equivalent iv dose Limiting factors include:Chemical instability, eg acid sensitive compound in the stomach Incomplete absorption - solubility, formulation Gut wall metabolism, 1 st pass metabolism - labile functional groups

12 A number of DMPK parameters may be predicted from in vitro assays to build up understanding of compound properties – predict behaviour in man Absorption – Pampa, Caco-2 Clearance - Microsomes, Hepatocytes Distribution - Plasma protein binding, Cytochrome P450 inhibition (5 major isozymes) Physical parameters – log D, pKa, solubility These assays are used extensively to profile compounds and filter out those that do not possess the required properties to be drugs. Predicting in vivo DMPK using in vitro Measurements

13 Pampa (Parallel Artificial Membrane Permeability Assay) Artificial membrane separates 2 compartments Models transcellular (passive) absorption only No tissue culture Assay 96 cpd (2 days experimental and analysis) Caco-2 Human colon adenocarcinoma Human colon carcinoma cell line which grow as monolayers, similar to small intestine enterocytes All mechanisms modelled - express key transporter proteins (e.g. PGP) Culturing over several days Assay 96 cpds AB or 48 cpd BA Can investigate different directions (A-B) and (B-A) Apparent permeability (P app ) measurements calculated (units are cm/sec x 1E-6) Typically use PAMPA assay as primary assay for absorption followed by oral data in two species Both can be related to human fraction absorbed Permeability in vitro Absorptive Flux (A-B) Basolateral Chamber (blood) drug Apical chamber (gut lumen) Cell monolayer Secretory flux (B-A)drug

14 In vitro Measurement of Metabolism In vivo Cl can also be predicted using in vitro assays: Microsomes (species – rat, dog, human) A subcellular fraction obtained by centrifugation of liver cells. Mainly composed of vesicles containing CYP450 enzymes formed from fragmented endoplasmic reticulum. Perform Phase I reactions. Hepatocytes (species – rat, dog, human) Isolated whole liver cells. Capable of performing both Phase I and II reactions. Rates of metabolism are reported as intrinsic clearance - Cl int (ul/min/10 6 cells) Typically: Rat Hepatocytes Cl int Low ( 20) Human Microsomes Cl int Low ( 30)

15 In vitro – In vivo scaling Cannot measure in vivo human PK until phase I trials. If we can predict consistently how a compound will behave in other species, then we can have greater confidence that it will behave predictably in man. In vitro - in vivo scaling is the prediction of in vivo Cl from Cl int measured in hepatocytes. The in vitro assay gives the maximum possible metabolic rate – but need to factor in drug delivery i.e. liver size, PPB, liver blood flow etc, extrahepatic clearance. Cl int  l/min/10 6 cells 120 x 10 6 hu heps per gram liver  l/min/g liver Cl int *  ml/min/kg g liver per kg body weight  l to ml Liver Species 1Species 2Human Heps low Compound scales Hu heps low In vitro In vivoFTIM DMPK well understood, predictable from hepatocytes, PPB etc.

16 Dose Prediction to Man DMPK measurements enable prediction of human PK parameters. Incorporation of potency and safety data enables Dose to Man (DtM) and safety margin predictions. Predicted human PK appropriate for once a day oral dosing: Therapeutic Dose < 5mg/kg uid Pred. Human DMPKt 1/2 6-12h, F > 30% Plasma conc Time Plasma conc Therapeutic Toxic Ineffective Css min (typically 3 x potency) Css max Safety Margin

17 Summary Definitions and qualitative aspects of absorption, distribution and elimination. Quantitative PK studies allowing the determination of: PermeabilityEffluxAqueous solubility Renal excretion Metabolic stability Biliary excretion Protein binding Tissue binding fafa ClVDVD %Ft 1/2 (poor/med/high)(UID/BID/>3-4x) AbsorptionElimination Distribution Knowledge of these parameters allows identification of where improvements need to be made to end up with a pharmacokinetically optimized drug.


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