Saul J. Karpen, M.D., Ph.D.

Separating the metabolic benefits from lipoprotein abnormalities

associated with FXR agonists--gut vs. liver effects

Raymond F. Schinazi Distinguished Biomedical Chair

Professor of Pediatrics

Paris July 1, 2016

No Financial Disclosures

OCA/plac. x 72 w.

ALT

FXR AGONIST • Chol

• HDL

• LDL

• ~ TG

-0,25

-0,15

-0,05

0,05

0,15

0,25

OCA (126)

Plac (131)

Chol HDL LDL TG

D L

ipid

(m

M)

P= 0.0009

P= 0.01

P< 0.0001 P= 0.88

Gastro 2015 PMID:25500425

0

50

100

150

200

250

300

Chol LDL HDL TG

Plac

OCA (10 mg)

OCA (25 mg)

* *

* *

EO

T [

Day 8

5]

Lip

id L

evels

(m

g/d

l)

FXR AGONIST • Chol #

• HDL

• ~ LDL #

• ~ TG

# : Different from NASH

17 D

28 D

Pencek, Diabetes, Obesity & Metabolism 2016, PMID:27109453

OCA in 68 Healthy Volunteers: Lipid Analyses

Transporters & FXR: Ileal & Hepatic Components of the EHC

Ileum

Liver

Schaap, Nat Rev GI Hep 2014

LXR FXR

MAPK

PI3K

PKC

TGR5

AP1 FAS

TRAIL

PGC1

CAR

VDR

PXR

Cell

Signaling

Apoptosis

Nuclear

Receptors

JNK

p38MAPK

ERK1/2

SHP

FGF19

S1PR2

Multiple Molecular Roles for Bile Acids

Cholic Acid Chenodeoxycholic acid

Lithocholic Acid Z-Guggulsterone

FXR Agonists

FXR Antagonists

GW4064

EC50: 4-10 M 37-80 nM 20 M

IC50:

Deoxycholic acid

19-50 M

6--ethyl-CDCA

(Obeticholic acid) 99 nM

10 M

Makishima Science 1999

Parks Science 1999

Wang Mol Cell 1999

Urizar Science 2002

Yu JBC 2002

Pellicciari J Med Chem 2002

Hawkins JCI 2002

Dussault JBC 2003

Downes Mol Cell 2003

Carter Ped Res 2007 10-30 M

Stigmasterol

10 M

Fexaramine

25 nM

Fexarine

38 nM

Fexarene

36 nM

AGN31

2 M (also RXR)

2016, PMID: 26812075 OCA x 24 h: FXR targets

Caveats:

• “The hPCLS used for this study were obtained from patients with a

high BMI (35–43 kg/m2).”

• Dedifferentiated human cells in culture—CYP7A1 & CYP8B1 were not

downregulated.

SHP BSEP OST MDR3 FGF19

Protein

RNA:

J Clin Invest. 2015;125:386–402.

Intestinal FXR Antagonism

improves NASH in mice

Intestinal FXR Agonism

improves NASH in mice

Essential, but seemingly contradictory effects of FXR & BA signaling in NAFLD

Intestinal FXR ko protects against

HFD-induced hepatic steatosis

Nat Med. 2015;21:159–165.

Fexaramine (Intestinal FXR agonist) improves

HFD-induced hepatic steatosis

Glyco-Muricholic Acid:

• Intestinal FXR antag.

• Brown Fat activity

• Intest. Ceramides

NAFLD & NASH:

FXR Agonism or FXR Antagonism

Both work Why?

Intact Enterohepatic

BA Recirculation

ASBT

Ileal ASBT

Inhibition

Interrupted Enterohepatic

BA Recirculation

ASBT

Ileal ASBT inhibition will

improve the hepatic and

whole body response to

HFD in mice

Hypothesis:

Colon

BA’s

Microbial BA metab.

TGR5 signaling

BA Pool size

Liver

BA Synthesis

Cholesterol

Ileum

BA Uptake

FXR-FGF15/19 signaling

ALIOS (45% fat; 0.2% cholesterol), + Added Sugars in the Drinking Water

Chow

HFD

Asbti [SC-435] x 16wk

HFD Asbti

HFD:

Asbti: 0.006% SC-435, 10 mg/kg/day

Mice: Male, C57BL6J, 4-6 weeks, n=7-16/group

Study Design & Endpoints

4 8 12 16 . . . . . 0

Weeks • Weekly Body Weights

• Weekly Caloric & fluid intake

• Week 15 GTT, ITT

• Week 16

• Serum Liver Indices

• Feces Bile Acids

• Ileum RNA

• Colon RNA

• Liver Histology

• Lipids & Bile Acids

• RNA-Seq

• RNA & Protein

• Hydroxyproline

• Statistics: Mean ± SD

• ANOVA

Tetri LH. Am J Physiol GI 2008 Nov;295(5):G987–95.

Mells JE J Nutr Biochem. 2015 Mar;26(3):285–92. AASLD 2015

SC-435 Inhibits Ileal ASBT function

Liver

0

1

2

3

4

Chow HFD HFD Asbti

a

b

a

Cyp7a1

0

1

2

3

4Cyp8b1

a

b

a

Chow HFD HFD Asbti

Rela

tive

gen

e e

xp

res

sio

n

Colon

0

5

1 0

1 5

2 0 Ilbp

a a

b

Chow HFD HFD Asbti

Rela

tive

gen

e e

xp

res

sio

n

ASBT

Fec

al B

ile

Ac

ids

(

M/2

4 h

r)

HFD Asbti

Feces

HFD

a

b

0

1

2

3

Fgf15

a

b

c

Chow HFD HFD Asbti

Ileum

Rela

tive

gen

e e

xp

res

sio

n

Asbt

HFD Asbti

Chow HFD

Shp

a

b

a

AASLD 2015

Glu

co

se (

mg

/dL

)

Time (mins)

ASBTi Improves Glucose Tolerance

Chow HFD HFD Asbti

* *

*

Chow

HFD + Asbti

HFD

GT

T A

UC

a

b

a

*: Significantly different from HFD + Asbti

GTT

AASLD 2015

Triglycerides Cholesteryl Ester

(g

/mg

liv

er)

(g

/mg

liv

er)

a

b

a

a

b

a

Chow HFD HFD Asbti

Chow HFD HFD Asbti

ASBTi Reduces Hepatic Lipids, But Not Total Bile Acids

Chow HFD HFD Asbti

(pm

ol/

mg

liv

er

tiss

ue

)

Total Bile Acids

AASLD 2015

a

b b

Chow HFD HFD Asbti

Liver Wt/Body Wt

ASBTi Improves Hepatic NAS & Steatosis Scores

Chow HFD HFD + Asbti

NAS Score

Chow HFD HFD Asbti

a

b

c

Steatosis Score

Chow HFD HFD Asbti

a

b

c

AASLD 2015

FXR Antagonist FXR Agonist

HFD

58% 42%

HFD + Asbti

83%

17%

Bile

Aci

d (

pm

ol/

mg

tiss

ue

) ASBTi Markedly Alters Hepatic BA Composition

* * * * * -TMCA b-TMCA w-TMCA THDCA TUDCA -MCA b-MCA

FXR Antagonist

HFD

HFD + Asbti

TCA TCDCA TDCA TLCA CA

*

*

*

FXR Agonist

*

AASLD 2015

HFD

Insulin resistance

Hyperglycemia Hyperinsulinemia

ChREBP

Lipogenic genes TG

SREBP-1c LXR Hepatic

cholesterol

FXR Hepatic

BA Composition

ASBTi

ASBTi

ASBTi

Hepatic

Steatosis

ASBT

Hypothesized Mechanisms of Action of ASBTi in Liver

FXR Antagonist

FXR Agonist

HFD

58% 42%

HFD + Asbti

83%

17%

Ileal ASBT inhibition Markedly Alters Hepatic BA Composition

TMCA’s THDCA TUDCA

TCA TCDCA TDCA

AASLD 2015

March 2016, PMID: 26708144

Nature. 2014 Mar 26.

HFD, 11w of VSG in mice KO = FXR-/-

Intact BA signaling, through its receptor, FXR,

mediates the response to Bariatric Surgery

Tilg NEJM June 23, 2016

Interaction of Diet (PC), microbes, BAs, Genes (FMO3) CV Disease

Bile acid based therapeutic trials (~ 200 in clinicaltrials.gov)

FXR agonists: Obeticholic Acid

TGR5 agonists:

ASBT inhibitors:

NorUDCA:

NASH

PBC

PSC PSC

Satiety

Constipation

Pruritus in cholestasis (ALGS, PFIC’s)

IBS-C

PSC

Glycocholic Acid: BA Synthesis Defect

BA Sequestrant: Colesevelam Diabetes

NASH

Obesity

BA diarrhea

Alcohol

Fibrosis

Summary: FXR & the Lipids in NASH

• Bile acid (BA) biology: Opportunities for discovering new

linked components of the Gut-Liver-Microbe-Gene Axis

– Differential effects of FXR & BAs in Ileum, Colon, Liver, Fat, …

– Individual BA’s have distinct functional properties

• FXR Agonists in NAFLD & NASH: Lipid Issues

– Reduces CYP7A1 & BA synthesis

– Total Cholesterol

– LDL

– HDL

• Further evidence that we will need to attack NASH

from multiple therapeutic angles – FXR Agonism & Antagonism both improve NASH in mice

– ASBT inhibition improves NASH in mice

• Reduces Hepatic TG & Cholesterol

Saul Karpen, MD, PhD

Paul Dawson, PhD

Astrid Kosters, PhD

Anuradha Rao, PhD

Angelica Amanso, PhD

JP Berauer, MD

Gina Ramirez

Funding (NIH) • R01 DK056239

• R01 DK047987

• Philanthropies:

• Alpard Foundation

• Spain Fund

• Moss Fund

Anya Mezina,MD MSCR

Courtney Ferrebee

Jamie Mells, PhD

Kim Pachura

Jianing Li, PhD

Grace Wynn

Prabhu Shankar, MD

Emory University (Saul-Paul Lab)

Hong Yin, MD (Pathology)

Dean Jones, PhD (Metabolomics)

Sophia Banton

Shuzhao Li

Hao Wu, PhD (School of Public Health)

Emory University

Brad Keller, PhD (Lumena/Shire)

Ken Setchell, PhD

Wujuan Zhang, PhD

Cincinnati Children’s

HDL Pathway targets LDL Pathway targets

Genes by OCA:

• Abca1

• Tgm2

• Fgl1

• Npc1l1

• Angptl4

• Hif1

• Ghr