ADME in NAFLD: Genes and Big Jeans Increase the Risk of Adverse Drug Reactions. Drug-Induced Liver Injury

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1 ADME in NAFLD: Genes and Big Jeans Increase the Risk of Adverse Drug Reactions Nathan J. Cherrington, Ph.D. Drug-Induced Liver Injury Minimal Toxicity Toxicity Exposure 1

2 Obesity: An Increasing Health Crisis Dec 11 th, 23 The Economist Obesity Trends Among U.S. Adults BRFSS, 1985 (BMI 3, or ~ 3 lbs. overweight for 5 4 person) No Data <1% 1% 14% 2

3 Obesity Trends Among U.S. Adults BRFSS, 1986 (BMI 3, or ~ 3 lbs. overweight for 5 4 person) No Data <1% 1% 14% Obesity Trends Among U.S. Adults BRFSS, 1987 (BMI 3, or ~ 3 lbs. overweight for 5 4 person) No Data <1% 1% 14% 3

4 Obesity Trends Among U.S. Adults BRFSS, 1988 (BMI 3, or ~ 3 lbs. overweight for 5 4 person) No Data <1% 1% 14% Obesity Trends Among U.S. Adults BRFSS, 1989 (BMI 3, or ~ 3 lbs. overweight for 5 4 person) No Data <1% 1% 14% 4

5 Obesity Trends Among U.S. Adults BRFSS, 199 (BMI 3, or ~ 3 lbs. overweight for 5 4 person) No Data <1% 1% 14% Obesity Trends Among U.S. Adults BRFSS, 1991 (BMI 3, or ~ 3 lbs. overweight for 5 4 person) No Data <1% 1% 14% 15% 19% 5

6 Obesity Trends Among U.S. Adults BRFSS, 1992 (BMI 3, or ~ 3 lbs. overweight for 5 4 person) No Data <1% 1% 14% 15% 19% Obesity Trends Among U.S. Adults BRFSS, 1993 (BMI 3, or ~ 3 lbs. overweight for 5 4 person) No Data <1% 1% 14% 15% 19% 6

7 Obesity Trends Among U.S. Adults BRFSS, 1994 (BMI 3, or ~ 3 lbs. overweight for 5 4 person) No Data <1% 1% 14% 15% 19% Obesity Trends Among U.S. Adults BRFSS, 1995 (BMI 3, or ~ 3 lbs. overweight for 5 4 person) No Data <1% 1% 14% 15% 19% 7

8 Obesity Trends Among U.S. Adults BRFSS, 1996 (BMI 3, or ~ 3 lbs. overweight for 5 4 person) No Data <1% 1% 14% 15% 19% Obesity Trends Among U.S. Adults BRFSS, 1997 (BMI 3, or ~ 3 lbs. overweight for 5 4 person) No Data <1% 1% 14% 15% 19% 2% 8

9 Obesity Trends Among U.S. Adults BRFSS, 1998 (BMI 3, or ~ 3 lbs. overweight for 5 4 person) No Data <1% 1% 14% 15% 19% 2% Obesity Trends Among U.S. Adults BRFSS, 1999 (BMI 3, or ~ 3 lbs. overweight for 5 4 person) No Data <1% 1% 14% 15% 19% 2% 9

10 Obesity Trends Among U.S. Adults BRFSS, 2 (BMI 3, or ~ 3 lbs. overweight for 5 4 person) No Data <1% 1% 14% 15% 19% 2% Obesity Trends Among U.S. Adults BRFSS, 21 (BMI 3, or ~ 3 lbs. overweight for 5 4 person) No Data <1% 1% 14% 15% 19% 2% 24% 25% 1

11 Obesity Trends Among U.S. Adults BRFSS, 22 (BMI 3, or ~ 3 lbs. overweight for 5 4 person) No Data <1% 1% 14% 15% 19% 2% 24% 25% Obesity Trends Among U.S. Adults BRFSS, 23 (BMI 3, or ~ 3 lbs. overweight for 5 4 person) No Data <1% 1% 14% 15% 19% 2% 24% 25% 11

12 Obesity Trends Among U.S. Adults BRFSS, 24 (BMI 3, or ~ 3 lbs. overweight for 5 4 person) No Data <1% 1% 14% 15% 19% 2% 24% 25% Obesity Trends Among U.S. Adults BRFSS, 25 (BMI 3, or ~ 3 lbs. overweight for 5 4 person) No Data <1% 1% 14% 15% 19% 2% 24% 25% 12

13 Obesity Trends Among U.S. Adults BRFSS, 26 (BMI 3, or ~ 3 lbs. overweight for 5 4 person) No Data <1% 1% 14% 15% 19% 2% 24% 25% 29% 3% Obesity Trends Among U.S. Adults BRFSS, 27 (BMI 3, or ~ 3 lbs. overweight for 5 4 person) No Data <1% 1% 14% 15% 19% 2% 24% 25% 29% 3% 13

14 Obesity Trends Among U.S. Adults BRFSS, 28 (BMI 3, or ~ 3 lbs. overweight for 5 4 person) No Data <1% 1% 14% 15% 19% 2% 24% 25% 29% 3% Obesity Trends Among U.S. Adults BRFSS, 29 (BMI 3, or ~ 3 lbs. overweight for 5 4 person) No Data <1% 1% 14% 15% 19% 2% 24% 25% 29% 3% 14

15 Obesity Trends Among U.S. Adults BRFSS, 21 (BMI 3, or ~ 3 lbs. overweight for 5 4 person) No Data <1% 1% 14% 15% 19% 2% 24% 25% 29% 3% Prevalence of Nonalcoholic Fatty Liver Disease in U.S. 38 million Ref: Ali & Cusi, Ann Med. 29;41(4): McCullough et al. J Clin Gastroenterol. 26 Mar;4 Suppl 1:S

16 Prevalence of Nonalcoholic Fatty Liver Disease in U.S. Healthy million NAFLD 9-12 million 3 4% Metabolic Syndrome in the liver is manifested as Nonalcoholic Fatty Liver Disease Ref: Ali & Cusi, Ann Med. 29;41(4): McCullough et al. J Clin Gastroenterol. 26 Mar;4 Suppl 1:S17-29 Prevalence of Nonalcoholic Fatty Liver Disease in U.S. Healthy million Steatosis 4-15 million 5-17% Steatohepatitis 15-5 million Ref: Ali & Cusi, Ann Med. 29;41(4): McCullough et al. J Clin Gastroenterol. 26 Mar;4 Suppl 1:S

17 Prevalence of Nonalcoholic Fatty Liver Disease in U.S. Healthy million Steatosis 4-15 million 5-17% Diagnosed 3, Steatohepatitis 15-5 million Ref: Ali & Cusi, Ann Med. 29;41(4): McCullough et al. J Clin Gastroenterol. 26 Mar;4 Suppl 1:S17-29 Prevalence of Nonalcoholic Fatty Liver Disease in U.S. Healthy million Steatosis 4-15 million 5-17% Diagnosed 3, Steatohepatitis 15-5 million Ref: Ali & Cusi, Ann Med. 29;41(4): McCullough et al. J Clin Gastroenterol. 26 Mar;4 Suppl 1:S

18 NAFLD Comprises a Spectrum of Pathologic Severity Healthy liver Steatosis NASH w/fat NASH not fatty (Cirrhosis) - benign - reversible - steatosis - irreversible scarring - inflammation - end-stage - progressive fibrosis NAFLD = Nonalcoholic Fatty Liver Disease NASH = Nonalcoholic Steatohepatitis NAFLD Comprises a Spectrum of Pathologic Severity Healthy liver Steatosis NASH w/fat NASH not fatty (Cirrhosis) NAFLD = Nonalcoholic Fatty Liver Disease NASH = Nonalcoholic Steatohepatitis nd hit includes Inflammation and oxidative stress : Global Macrophage Gene and Expression stellate cell activation Analysis 33,252 Cytokine genes- and Affymetrix ROS production Genechip Human 1. ST Array Mitochondrial dysfunction Lipid peroxidation.1 Significance Level 18

19 Diagnosis Sex PCA Cluster by Diagnosis Age The Liver is the Major Organ of Drug Metabolism and Elimination SINUSOIDAL BLOOD Phase I Drug Metabolism - Cytochrome P45s Phase II Drug Conjugation Phase III Transport O OH CO2H BILE OH OH EPATOCYTE OH HEPATOCYTE HEPATOC 19

20 Drug Metabolizing Enzyme Protein Expression in NAFLD Patients Normal Steatosis NASH (fatty liver) NASH (not fatty) Densitometry Statistics CYP1A2 Trend (p=.1) CYP2A6 Trend (p=.2) CYP2C9 CYP2C19 Trend (p=.1) CYP2D6 Trend (p=.68) CYP2E1 Trend (p=.1) CYP3A4 Trend (p=.112) GAPDH Indicates significant trend with progressive disease states (p <.5) Ref: Fisher et al. DMD. 29: 37(1): Drug Metabolizing Enzyme Activity in NAFLD Patients Trend p =.1 Trend p =.4 Trend p =.18 Trend p =.1 Ref: Fisher et al. DMD. 29: 37(1):

21 Nrf2 Activation in All Stages of NAFLD Normal Steatosis Normal Steatosis NASH (fatty) 2X NASH (not fatty) 2X 2X = Nrf2 staining = No Nrf2 staining Ref: Hardwick et al. DMD. 21: 38(12): X The Affect of NAFLD on NQO1 Normal Steatosis NASH (fatty) NASH (not fatty) NQO1 Total ERK Ref: Hardwick et al. DMD. 21: 38(12):

22 Global Gene Expression All genes DMEs Overexpression No change Downregulation Ref: Lake et al. DMD. 211: 39(1): ,252 genes-affymetrix Genechip Human 1. ST Array Global Gene Expression DMEs Ref: Lake et al. DMD. 211: 39(1):

23 Transporters Global Gene Expression Ref: Lake et al. DMD. 211: 39(1): Transporters Global Gene Expression Ref: Lake et al. DMD. 211: 39(1):

24 Uptake Transporters Global Gene Expression Steatosis-NORMAL NASH-Steatosis NASH-NORMAL Ref: Lake et al. DMD. 211: 39(1): Plasma BSP Concentration (mg/1ml) mulative Biliary BSP Excretion (mg/ml) Normal High Fat NASH diet Ref: Fisher et al. 25 Eur J Pharmacol. 29: 613(1-3): Plasma BSP Concentration (mg/1ml) Cumulative Biliary BSP Excretion (mg/ml) Bile Flow ul/min/kg) Disposition in NASH NAFLD Alters BSP Clearance Figure 5 Functional Plasma BSP Disposition in NASH Time (min) Normal High Fat NASH diet Bile Time (min) 24

25 The Liver is the Major Organ of Drug Metabolism and Elimination SINUSOIDAL BLOOD Phase I Drug Metabolism - Cytochrome P45s Phase II Drug Conjugation Phase III Transport O OH CO2H BILE OH OH EPATOCYTE OH HEPATOCYTE HEPATOC Efflux Transporter Protien Expression ABCC1 Normal Steatosis NASH (fatty) NASH (not fatty) ABCC3 ABCC4 ABCC5 ABCC6 ABCB1 ABCG2 Pan-Cadherin Ref: Hardwick et al. DMD. 211: Epub ahead of print 25

26 ABCC3 Immunohistochemistry Normal Steatosis NASH (fatty) 4X NASH (not fatty) 4X Ref: Hardwick et al. DMD. 211: Epub ahead of print 4X 4X 51 Cellular Localization of ABCC2 Normal Steatosis NASH (fatty) 4X NASH (not fatty) 4X Ref: Hardwick et al. DMD. 211: Epub ahead of print 4X 4X 26

27 Cellular Localization of ABCC2 Normal NASH (not fatty) 1X 1X NASH NASH Normal Steatosis (fatty) (not fatty) ABCC2 2kDa 18kDa Cadherin Ref: Hardwick et al. DMD. 211: Epub ahead of print APAP Elimination in Normal Liver BLOOD LIVER BILE APAP-Gluc APAP-Sulf 27

28 APAP Elimination in Normal Liver BLOOD LIVER BILE APAP-Gluc APAP-Sulf APAP Elimination in NASH BLOOD LIVER BILE APAP-Gluc APAP-Sulf 28

29 APAP Elimination in NASH BLOOD LIVER BILE APAP-Gluc APAP-Sulf APAP Disposition APAP (nmol/ml) APAP-Gluc (nmol/ml) APAP-Sulf (nmol/ml) Rat Plasma Ref: Lickteig et al. DMD. 27: 35(1):197-8 Control SFL NASH Time (min) Rat Bile Control HF MCD Time (min) APAP = Acetaminophen p <.5 significant vs. Control 29

30 APAP Disposition APAP (nmol/ml) Rat Plasma Control SFL NASH APAP-Gluc (nmol/ml) APAP-Sulf (nmol/ml) Ref: Lickteig et al. DMD. 27: 35(1): Time (min) APAP = Acetaminophen p <.5 significant vs. Control APAP Disposition APAP (nmol/ml) Rat Plasma Control SFL NASH Human Plasma Normal Steatosis NASH APAP-Gluc (nmol/ml) APAP-Sulf (nmol/ml) 1 Ref: Merrell et al. unpublished data Time (min) APAP = Acetaminophen p <.5 significant vs. Control 3

31 Ezetemibe Disposition Liver EZE EZE-GLUC Small Intestine Abcc2 and Abcb1 Localization Control MCD 1X 1X Abcb1 Abcc2 Ref: Hardwick et al. unpublished data 1X 1X 31

32 EZE and EZE-Gluc Disposition Plasma Bile Ref: Hardwick et al. unpublished data Drug-Induced Liver Injury Minimal Toxicity Toxicity Exposure 32

33 Summary Drug Metabolizing Enzyme and Transporter expression changes occur during the transition from simple fatty liver to NASH Mechanisms of regulating DME and Transporter function during NASH may be controlled at the transcription, post-translational modification, and cellular localization levels Changes in the expression of the ADME battery of genes results in altered disposition and exposure of numerous drugs and xenobiotics Understanding inter-individual variation in drug metabolism may help lead to personalized medicine Cherrington Lab Andrew Lickteig, Ph.D. Craig Fisher, Ph.D. Lisa Augustine Matthew Merrell, Ph.D. Rhiannon Hardwick Lisa Beilke, Ph.D. Jonathan Jackson, Ph.D. April Lake Mark Canet David Klein Walter Klimecki, Ph.D. Dean Billheimer, Ph.D. Robert P. Erickson, M.D. H. Hesham A-Kader, M.D. Petr Novak, Ph.D. Leif Abrell, Ph.D. Acknowledgments Jose E. Manautou, Ph.D. Sarah Campion, Ph.D. - University of Connecticut Angela Slitt, Ph.D. - University of Rhode Island Lauren Aleksunes, Ph.D. - Rutgers University Michael Goedken, Ph.D. - Merck Steven Ferguson, Ph.D. - Invitrogen Funding HRAA NICHD HD62489 NIDDK DK6839 NIAID AI83927 NIEHS ES11646, ES791, ES