* Your assessment is very important for improving the workof artificial intelligence, which forms the content of this project
Download Understanding Oral Bioavailability Challenges Associated With
Discovery and development of non-nucleoside reverse-transcriptase inhibitors wikipedia , lookup
Discovery and development of cephalosporins wikipedia , lookup
Psychopharmacology wikipedia , lookup
Drug design wikipedia , lookup
Prescription costs wikipedia , lookup
Zoopharmacognosy wikipedia , lookup
Neuropsychopharmacology wikipedia , lookup
Pharmaceutical industry wikipedia , lookup
Neuropharmacology wikipedia , lookup
Plateau principle wikipedia , lookup
Discovery and development of tubulin inhibitors wikipedia , lookup
Pharmacognosy wikipedia , lookup
Magnesium transporter wikipedia , lookup
Gastrointestinal tract wikipedia , lookup
Pharmacogenomics wikipedia , lookup
Drug discovery wikipedia , lookup
AAPS Chicagoland Pharmaceutical Discussion Group Understanding Oral Bioavailability Challenges Associated With Poorly Soluble Compounds That Are Also Effluxed or Metabolized Ming Hu, Ph.D. College of Pharmacy University of Houston Houston, TX 77030 [email protected] 832--842832 842-8320 OUTLINE ¾ Overview of Gut and Liver Biology and Physiology ¾ Barriers to Oral Bioavailability ¾ Challenges Associated with Poorly Soluble Compounds ¾ Concluding Remark OUTLINE ¾ Overview of Gut and Liver Biology and Physiology ¾ Barriers to Oral Bioavailability ¾ Challenges Associated with Poorly Soluble Compounds ¾ Concluding Remarks OVERVIEW ¾ Intestinal and Gut Biology and Physiology ¾ Overview of Intestinal and Hepatic Enzymes and Transporters ¾ Models Useful for Study of Intestinal and Hepatic Enzymes and Transporters Physiology: The Digestive System Human Physiology, 1998 Intestinal Barrier: I t ti l Epithelium Intestinal E ith li (LUMEN) Intestinal Epithelium apical basolateral (LUMEN) Intestinal Epithelium apical basolateral (LUMEN) The Tight Junction (LUMEN) Summary of GI Tract Features From Physiological Pharmaceutics: Davis, 2002 Liver Biology and Physiology From Drug Metabolism by L. Gan Liver Lobule 9/20/2013 Bar = 250 nm 12 Hepatocytes ¾ Hepatocytes are exceptionally active in synthesis and transport. An ultra structural examination of hepatocytes reveals bountiful quantities of both rough and smooth endoplasmic reticulum. reticulum. In contrast to most glandular epithelial g p cells that contain a single g Golgi g organelle, hepatocytes typically contain many stacks of Golgi membranes. The ultra structural appearance of hepatocytes reflects their function as metabolic superstars.. superstars - Robert Bowen 9/20/2013 13 Blood Vessels Vs Vs. Cannicular 9/20/2013 14 Bile Cannicular 9/20/2013 15 Transport p Pathways y and Mechanisms Luminal pH pHd = 6.0 Transport Paracellular Intracellular ppH pHi=7.4 Active Diffusion Active F ili Facilitated d Parent Diffusion Facilitated Metabolite Serosal pH pHr = 7.4 Uptake and Efflux Transporters FDA DDI Guidance G id 2012 Draft D f Uptake Transporters Ê Ê Ê Ê Ê Ê Amino acids Small peptides (oligopeptides) Nucleotides OATPs OCT Others Peptide Transporter (PepT1) Expression Large quantities in duodenum, jejunum, ileum, nearly absent (<10% of small intestine) from colon in humans. (1. Drug Metab Dispos. 2007 35(8):1333--40. 2. Drug Metab Dispos. 2007 35(8):1333 35(4):590--4 ) 35(4):590 High expression level in duodenum, jejunum and ileum (used whole ileum) ileum), but low expression in colon. (Peptides. 2006 27(4):85027(4):850-7) Drugs Transported by PEPT1 Pflugers Arch - Eur J Physiol (2004) 447:610–618 Efflux Transporters P-Glycoprotein, MDR1 (other MDRs in other species) p ) Ê BCRP Ê MRPs (1 (1, 22, 33, 44, 55, 66, 7) Ê BSEP Ê MATEs Ê OATs Ê P-glycoprotein Ê Ê Ê Victor Ling, 1976 Design and development of inhibitors to enhance cancer therapy in vivo has been the dreams of newly four generations of scientists. If a drug can be made, Dr. Ling will receive a Nobel price for medicine. Ê Ê Ê Ê Ê First crystal structure of binding subunit, published in 1998 First bacteria crystal structure of a model protein, protein 2002 First mammalian crystal structure published in 2009 Countless papers of high quality Billions of dollars and tens of thousands of hours of intellectual pursuit. S G Aller et al. Science 2009;323:1718-1722 Binding of novel cyclic peptide P-gp inhibitors. Published by AAAS Fig. 4. Model of substrate transport by P-gp. S G Aller et al. Science 2009;323:1718-1722 Published by AAAS Metabolic Enzymes Phase I Enzymes CYPs FMO AO Hydrolases (esterases) Phase II Enzymes (Transferases) UGTs SULTs GTs Characteristics of Intestinal E Enzymes andd Transporters T Ê Region Region--Dependent Ê Concentration C Concentrationi -Dependent D d Rat Intestinal Efflux Transporter Expression Drug Metab Dispos. 2008 36(7):124936(7):1249-54. LC-MS LC MS Based Assay Fig.10. Amount of Bcrp in various rat tissues BCRP protein ( (pmol/mg 25 2.5 2 15 1.5 MRM 1 MRM 3 1 05 0.5 0 Liver WT Liver KO Duo WT Duo KO Jej WT Jej KO iLe WT iLe KO Lung WT Lung KO Amount of M A Metabolite E Excreted (nm mol/30min) Region-Dependent: Region Dependent: MRP2 14 12 10 8 6 4 2 0 Intestinal Excretion of Genistein Conjugates Wistar Duodenum Jejunum Ileum TR TR- Colon Region-Dependent Expression Panel C: Microsomal Rates Rattes of Glucuronidation (nmol/m min/mg of pro otein) 4.0 * 3.0 20 2.0 * * * 1.0 0.0 Duodenum Jejunum Ileum Colon Am mount of Conjugates Excreted (nmol/30min n) Panel B: Excretion 20 * * Duodenum Jejunum * 15 10 5 0 Ileum Colon Wang et al, DMD, 34(11):1837-48, 2006 Summary Features of Enzymes and T Transporters i Intestine in I i ¾ Transporter: ¾ ¾ ¾ Enzymes: ¾ ¾ ¾ Region-specific (or dependent) expression Region((“absorption absorption window”) window ) Polarized (apical vs. basolateral) Region R Regioni -Specific S ifi Expression E i Membrane--bound vs. Cytosolic Membrane Excretion: c et o : ¾ ¾ Region-Specific RegionApical vs. Basolateral Summary Features of Enzymes and T Transporters i Liver in Li ¾ Transporter: ¾ ¾ Enzymes: ¾ ¾ Polarized (apical vs. basolateral) Membrane--bound vs. Cytosolic Membrane Excretion: ¾ Apical vs. Basolateral Models and Experimental Systems Physicochemical: log P, log D, solubility, pKa, PAMPA and etc. Ê Biologically based models: Ê Fidelity Cost ¾ ¾ ¾ ¾ ¾ ¾ Human PK Animal PK In situ Excised tissue Cell culture M b Membrane vesicles/Microsomes i l /Mi Resolution Speed Epithelial Cell Culture Model AP Side (Lumen) BL Side (Serosal) B to A A to B Apical to Basolateral Transport: Absorptive Transport Basolateral to Apical Transport: Efflux Transport The Perfused Rat Intestinal Model 0.1-0.4 ml/min Heated to 37oC Perfusion Pump Perfusion Solution Perfusate D J I C Control Four site simultaneous perfusion model of rat intestine Four-site Flow rate may y be changed g to increase absorption/metabolism p during single pass perfusion. OUTLINE ¾ Overview of Gut and Liver Biology and Physiology ¾ Barriers to Oral Bioavailability ¾ Challenges Associated with Poorly Soluble Compounds ¾ Concluding Remarks Barriers to Oral Bioavailability Solubility and Dissolution ¾ Absorption ¾ Excretion ¾ Metabolism ¾ Transport p to Specific p Targets g ¾ Bioavailability Barriers Intestine Liver Target 1 Systemic Portal vein 2 4 Blood ? 3 ? 3 4 4 ? Bypass Hepatocytes 4 Bile 5 Importance of Solubility Ê Vabs =PSC Ê Mabs = P S C TR C: Mass Driving Force Ê Ê F passive For i diffusion, diff i P is i dependent d d on partition ii coefficients, diffusion coefficients, and unstirred water layer y thickness. For carriercarrier-mediated transport, P is dependent on Jmax and Km of the transporter, and concentration. P decreases with C when C approaches Km. Transport p Pathways y and Mechanisms Luminal pH pHd = 6.0 Transport Paracellular Intracellular ppH pHi=7.4 Active Diffusion Active F ili Facilitated d Parent Diffusion Facilitated Metabolite Serosal pH pHr = 7.4 Oral Absorption Pathways: Nutrient Carriers Variety: amino acids, glucose, peptides peptides,, nucleotides,, nucleobases,, vitamins,, etc. ¾ Variable capacity: Km ranging from several μM to several mM ¾ Structure: typically hydrophilic molecules with nutrient like structures ¾ Stringent: critical structure motif needed ¾ Other Drug Carriers Organic Anion Transporters Ê Organic Anion Transporting Peptides Ê Organic Cation Transporters Ê Monocarboxylic M b li Acid A id T Transporters Ê Excretion ¾ Efflux transporters were first recognized as a normal intestinal functions about 2 decades ago. Many efflux transporters recognize small hydrophobic molecules, especially those with planar structures. ¾ Efflux transporters were found to be necessary for cellular production of phase II conjugates was recognized i d about b 1 decade d d ago. Intestinal Excretion ¾ ¾ Counter the Absorption of Lipophilic Substances Efflux of Xenobiotics and Their Metabolites ¾ ¾ ¾ ¾ P-Glycoprotein (MDRs) BCRP MRP and MRPs d Oth Other ABC T Transporters t Other Classes of Transporters Efflux of Parent L i l Luminal Paracellular Efflux Intracellular pH ppHi=7.4 Enzy yme Efflux S Serosal l Hepatic Excretion ¾ ¾ Eliminate Lipophilic Substances from Blood Efflux of Xenobiotics and Their Metabolites ¾ ¾ ¾ ¾ P-Glycoprotein (MDRs) BCRP MRP and MRPs d Oth Other ABC T Transporters t Other Classes of Transporters Metabolism First Pass Metabolism Ê Phase I Ê Phase II Ê Interplay I l Ê Interplay ¾ Consequences of Interplay: ¾ ¾ ¾ Efficient production of phase II conjugates Multiple or repeated exposure to the same enzymes Recycling of xenobiotics (e.g., phenolics) Interplay: Excretion of Phase II Metabolites L i l Luminal Excretion Efflux Enzy yme Intracellular pH ppHi=7.4 S Serosal l Interplay: E Excretion ti off Parent P t and d Metabolite M t b lit L i l Luminal Metabolism and Efflux Efflux Enzy yme Intracellular pH ppHi=7.4 S Serosal l Enteric and Enterohepatic p Recycling y g LUMEN Glucoside Microflora Microflora Aglycone LPH MRP Conjugates BCRP SGLT1 MRP2 OAT β-glucosidase WALL Glucoside Aglycone Glucuronic Acid Sulfate OAT MRP ? bile MESSENTARY BLOOD Portal Vein LIVER MRP2 Heart Saturation of Protein Mediated Di Disposition iti (Metabolism (M t b li or Efflux) Effl ) Ê Vmet met/ex /ex = Vmax C/( C/(K Km+C +C)) Ê Mmet C/(K Km+C +C) C ) TR met/ex /ex = Vmax C/( Ê Ê Ê Ê C: Saturation means higher concentration will not increase rate of metabolism but may decrease it (substrate inhibition). Saturation of efflux will result in favorable mass action. C Compound d with ith fast f t permeability bilit andd poor protein t i binding bi di may escape metabolism or fair much better than predicted using microsomes. Inward concentration gradient and outward protein concentration gradient across the intestinal epithelium p favor uptake/absorption. p p Recycling of parent compound in the gut due to efflux may increase the extent of metabolism, OUTLINE ¾ Overview of Gut and Liver Biology and Physiology ¾ Barriers to Oral Bioavailability ¾ Challenges Associated with Poorly Soluble Compounds ¾ Concluding Remarks Biopharmaceutical Cl ifi i System Classification S Class I Class II High Dissolution/Solubility High g Permeabilityy Class III Low Dissolution/Solubility High g Permeabilityy Class IV High i h Dissolution/Solubility L Permeability Low P bilit Low Dissolution/Solubility L Permeability Low P bilit Class II Drugs • • • Low Solubility High Permeability Lipophilic Drug Molecules That Binds Often Selected via HTS. Binds, HTS Dose Number = Dose/(Cs V) Identification of Problems Poor aqueous solubility, less than 5 μM Ê Lower F at a higher dose in preclinical species Ê Complications: Efflux Metabolism Protein Bindingg Class IV Drugs Poor solubility: small versus large dose Ê Poor permeability: efflux vs. vs uptake Ê Poor solubility exasperates the efflux problem and/or first pass metabolism effects. Ê Identification of Problems Ê Ê Ê Low aqueous solubility, less than 5 μM Highly g y variable and poor p F at all doses in preclinical p species High efflux ratio in efflux transporter overexpressed cell culture models. Complications: Efflux Metabolism Food and/or fasting effects Complication: Solubility and Efflux Lower solubility means that higher potential for efflux. Ê Efflux inhibitors, with rare exception, have not been successful in the clinic clinic. Ê Enhancement of solubility will alleviate or even eliminate the problem. problem Ê F will increase with dose. Complication: Solubility and Metabolism Lower solubility means less potential for saturating g intestinal metabolism. Ê Intestinal inhibitors are sometimes successful in the clinic if the enzymes are not present in the liver. Ê Enhancement of solubility will alleviate or even eliminate the problem. Ê F will increase with dose. Complication: Solubility and Microbiome Ê Ê Ê Lower solubility means that intestinal microbiome will be exposed to the unabsorbed compounds Microbiome can mediate highly different metabolic pathway that may not be well conserved across species. i Enhancement of solubility will exasperate the problem bl if absorption b ti remain i largely l l incomplete. i l t Example: Ginsenosides R2O R2O OH OH 20 0 3 HO 6 R1O OR1 PPD type PPT type OH R1O OH R1O R2 Type C R2 Type D 62 Ginsenoside Bioavailability Ê L exposure (oral Low ( l bioavailability) bi il bilit ) off ginsenosides i id Ê Rg3 oral administration in human Ê Ê 0.8 mg/kg: g g below LOQ Q 3.2 mg/kg: Cmax is 15.6 ng/ml In Rat Oral bioavailability (%) Dose (mg/kg) Rh2 4.7 1 Rg3(r) < LOQ 100 CK 3.52 30 Re 0.16-0.28 1 Rb1 4 35 4.35 10 Rd 2.36 10 63 Identification of Problems No phase I metabolism Ê No phase II metabolism Ê No stability problem Ê Poor P solubility, l bili less l than h 5 uM M Ê Perm meability (cm m/s) 3.0E-05 2.5E-05 A to B 2.0E-05 B to A 1.5E-05 1 0E-05 1.0E 05 * 5.0E-06 * * * 0.0E+00 Efflux ratio (Pb-a / Pa-b) 28.5 1.0 1.2 I Intracellular r amounts (nmol/mg) Cell Culture Studies 0.7 0.6 05 0.5 0.4 0.3 0.2 0.1 0 Cell Culture Studies Rh2 Rh2 6.0E-05 B to A 4.0E-05 3 0E 05 3.0E-05 2.0E-05 1.0E-05 ** 0.0E+00 MDCKII MDR1-MDCKII MDR1-MDCKII +20µM CsA Inttracellular Conccentrations (nmol/mgg) 5.0E-05 Permeabillity (cm/s) 08 0.8 A to B ** 0.7 0.6 0.5 0.4 0.3 0.2 0.1 * 0 MDCKII MDR1-MDCKII MDR1-MDCKII +20µM CsA Animal Studies Plasma conccentrations ((µM) P 10 FVB alone Mdr1a/b -// 1 0.1 0.01 0 4 8 12 Time (hr) 16 20 24 Approaches Ê For highly permeable drugs, increase solubility y via nanoparticles, p , solid dispersion, emulsion, excipients excipients,, and other pp Hepatic p clearance formulation approaches. may still be a problem. problem. Ê Cyclosporin A, Griseoflavin Approaches Ê For poorly permeable drugs that are poorly soluble, identify the role of efflux transporters (if any), and then define approaches to the problems. problems Ê Ê Ê If efflux transporters are involved, solubility increase will greatly help. However, this may not solve rapid hepatic clearance problem. bl If extensive metabolism is known, intestinal metabolism may be inhibited but hepatic metabolism will remain a big problem. If no efflux transporters are involved, then prospects are very poor. Prodrugs are possible but often not viable based on business considerations. This may work for second generation of drugs. Phenolic Drugs Many phenolic drugs are rapidly metabolized Ê For highly potent drugs (effective in vivo concentration in nM/pM range), phenolics may not be as bad as feared feared. Ê Futile recycling may result in low bioavailability but long halfhalf-life (many sex hormones and SERMs such as raloxifene). Ê Phase Ph 2 metabolism b li may be b less l variable. i bl Ê Collaboration with Industry Ê Ê Ê Ê Ê Regional Absorption: Should we develop a sustained release formulation ? Solubility Enhancement: Would it really matter? Model System Characterization: Is the model reliable and reproducible? Protein Drug Adsorption: How to minimize the impact in clinical setting? Protein quantitation: Is it ready for prime time in ADME and PK? OUTLINE ¾ Overview of Gut and Liver Biology and Physiology ¾ Barriers to Oral Bioavailability ¾ Challenges Associated with Poorly Soluble Compounds ¾ Concluding Remarks Concluding Remarks Ê Ê Compounds with poor water solubility and high g permeability, p y, as well as slow metabolism or efflux is a prime candidate for elaborate formulation efforts. The caveat is that human disposition should be well established to decrease the chance of erroneous designation of low “metabolism” or “efflux.” Concluding Remarks Ê Compounds with poor solubility and extensive efflux by y intestinal but not hepatic p apical efflux transporters is a also a good candidate for elaborate formulation efforts. Ê Question? Should we even consider the importance of efflux transporters that are only expressed in the kidney? Concluding Remarks Ê Compounds with poor solubility and extensive efflux by y both intestinal and hepatic apical efflux transporters is a g g candidate to develop, p, and mayy challenging not deserve elaborate formulation efforts. Ê Question? Should we really develop poorly soluble compounds that are pp-glycoprotein or efflux transporter inhibitors (e.g., cyclosporin A like)? Concluding Remarks Ê Compounds with poor solubility and extensive metabolism by y both intestinal and hepatic apical enzymes is a challenging p, and should be candidate to develop, abandoned unless it is highly potent and g recycling. y g undergo Ê Question? Could phenolic compounds that are highly potent but extensively metabolized and recycled always succeed? Concluding Remarks Ê Compounds with poor solubility and extensive metabolism by y only y intestinal but not by hepatic apical enzymes is a good candidate to for elaborate formulation efforts to increase its solubility. Ê Question? Do you know many phase II enzymes display substrate inhibition kinetics, meaning lower rates at higher concentration? Concluding Remarks Ê Because intestinal microbiome will be exposed p to unabsorbed compounds p and intestinal microbiome structures appear to correlate with human health,, FDA will begin to mandate studies of a drug’s effect on intestinal microbiome within a decade. The tools should be ready then. Acknowledgement ¾ Postdoctoral Researcher ¾ Ph.D. Students ¾ ¾ ¾ ¾ ¾ ¾ ¾ ¾ ¾ ¾ ¾ Grant Dr. Jun Chen Dr.. Song Gao Dr D B Dr. Beibei ib i Xu X ¾ ¾ ¾ Dr. Stephen Wang D Zh Dr. Zhen Yang Y Dr. Zhu Wei Dr. Kaustubh H Kulkarni, Dr. Dr Rashim Singh Dr. Wen Jiang Dr. Baojian Wu ¾ ¾ ¾ ¾ ¾ Ms. Terry Yin Collaborator ¾ ¾ ¾ Senior Research Associate: ¾ NIH AT 000182 NIH AT 003203 NIH AT 005522 NIH GM 070737 NIH CA 087779 Dr. Ming You, Medical College of Wisconsin Dr. Dr Joseph Petrosino, Petrosino Baylor College of Medicine Dr. Zhongqiu Liu, Guangzhou University of Chi Chinese Medicine M di i Role of MRP2 Amountss Extcretted (nmoll/30 min) 3.0 2.5 Wistar Billary Genistein Conjugates TR- 2.0 1.5 1.0 0.5 0.0 60 90 120 Time (min) ( ) 150