Barry M. Forman, MD, PhDAssociate Professor

Beckman Research InstituteCity of Hope National Medical Center

Duarte, California

PolypharmacyPolypharmacy & OTCs:& OTCs:Impact of Dietary Supplements on Gene Networks Impact of Dietary Supplements on Gene Networks

that Regulate Drug Clearancethat Regulate Drug Clearance

Dietary Supplement Use in the ElderlyDietary Supplement Use in the ElderlyNational Institutes of Health

Nuclear Hormone ReceptorsCoactivator Recruitment

HREHRE

RXR Rec

TATA

Rec

CoactivatorsCoactivators

Activated TranscriptionActivated Transcription

LigandLigand

Orphan Nuclear ReceptorsA New Endocrinology

LigandsN - DNA - CNovel Ligands?

SteroidThyroidRetinoid

Vitamin D

Hormonal Ligands

SteroidsThyroidRetinoidsVitamin D

Classical Receptors

MetabolitesLipidsFatty acidsSterols Bile Acids

Xenobiotics

Orphan Receptors

SXR / PXRA steroid & xenobiotic receptor

Expression: Liver & IntestineExpression: Liver & Intestine

Target Genes: Cyp3A4Target Genes: Cyp3A4

PXRE

RXRRXR PXRPXR Ligands: PhenobarbitalRifampicinClotrimazole

Ligands: PhenobarbitalRifampicinClotrimazole

Function: Protection from Xenotoxins

Enhancement of Drug MetabolismMediator of Drug-Drug Interactions

CyclosporinHIV Protease InhibitorsBirth Control “Pill”

Function: Protection from Xenotoxins

Enhancement of Drug MetabolismMediator of Drug-Drug Interactions

CyclosporinHIV Protease InhibitorsBirth Control “Pill”

TaxolChemotherapeutic agent

Efficacy: Breast & Ovarian CancersActivity: Microtubule StabilizationMetabolism: Cyp3A4 & Cyp2C8

OAc

OHO

OAcOO

O

OOH

O

N

O

OH

H

Taxol is a Specific Activator of hPXRPXR vs other receptors

Taxol10 µM

1

5

9

13

17

21

25

29

hPXR

mPX

RER

αVD

RTR

βR

ARα

LXRα

PPAR

αPP

ARγ

PPA

R δC

ARβ

Fold

Act

ivat

ion

FXR

PXR & Taxol DegradationCyp3A4 & Cyp2C8

Primary Human Hepatocytes

Taxo

l

Rifa

mpi

-

n

Cyp 3A4

Cyp 2C8

GAPDH

Hea

rt

Brai

n

Plac

enta

Lung

Live

r

Skel

etal

Mus

cle

Kidn

ey

Panc

reas

Thym

us

Pros

tate

Test

is

Ova

ry

Smal

l int

estin

e

Col

on

Leuk

ocyt

es

Sple

en

hPXR ExpressionLiver & Intestine

Adapted from Blumberg et al. , Lehmann et al., & Bertilsson et al.

PXR Function in the Intestine?Regulation of MDR1

Taxo

l

Rifa

mpi

-

n

MDR1

Human Intestinal CellsLS180

PXRA master regulator of drug clearance

Metabolism

Multi-Drug Resistance?

Excretion

Taxol & TaxotereRelated chemotherapeutic agents

Efficacy: Breast & Ovarian CancersActivity: Microtubule Stabilization

OAc

OHO

OAcOO

O

OOH

O

N

O

OH

H

Taxol

OH

OHO

OAcOO

O

OOH

O

NO

O

OH

H

Taxotere

Taxol vs TaxotereTaxotere does NOT activate hPXR

Taxol

GAL-hPXRFo

ld A

ctiv

atio

n

Taxotere

50

40

30

20

10

0

0 5 10 15 20 25 30

Taxane(mM)

Taxol vs TaxotereTaxotere does NOT induce drug clearance

MetabolismMetabolism

0

1

2

3

4

Taxol Taxotere-

P-G

lygo

prot

ein

Activ

ity

EffluxEfflux

Intestine

Taxol Taxotere-0

25

50

75

100

125

Cyp

3A4

Activ

ity3’

p-O

H T

axol

Pro

duct

ion

Hepatocytes

Taxo

l Ef

flux

Rat

e

HIV Protease Inhibitors & PXRRitonavir is a PXR ligand

TransfectionTransfectionhPXR

0

200

400

600

800

1000

1200

1400

|

UnlabeledProtease Inhibitor

hPXR+

[3 H]-SR12813

Hyp

erfo

rinR

itona

vir

Saqu

inav

ir

Nel

finav

irIn

dina

vir

Ligand BindingLigand BindinghPXRhPXR

Spec

ific

Bind

ing

(cpm

)

SNS

NN N N N O

O

O OH

O

Ritonavir

StructureStructurehPXR

1

4

7

10

13

16

19GAL-hPXR

Fold

Act

ivat

ion

Rifa

mpi

ci

Rito

navi

Saqu

inav

ir

Nel

finav

irIn

dina

vir

Protease Inhibitor

rn

Steroid & Xenobiotic Receptor (PXR/SXR)PXR ligands are Cyp3A4 substrates

Function: Master regulator of drug & xenobiotic clearance

Function: Master regulator of drug & xenobiotic clearance

Expression: Liver & IntestineExpression: Liver & Intestine

Target Genes: MDR1, MRP2, OATP2 Cyp3A4, Cyp2C8, Cyp2BGSTa1, UGT1A, CARAminolevulinate synthase (ALAS), Por

Target Genes: MDR1, MRP2, OATP2 Cyp3A4, Cyp2C8, Cyp2BGSTa1, UGT1A, CARAminolevulinate synthase (ALAS), Por

PXRE

RXRRXR PXRPXR

Ligands: Common Drugs & XenobioticsEndogenous Ligands?Dietary Supplements?

Ligands: Common Drugs & XenobioticsEndogenous Ligands?Dietary Supplements?

Triol: a Potent PXR Agonist Cyp3A4 & PXR utilize common sterol substrates

HO OH

OH

NoneNone PCNPCN0

1

2

3

4

5

Rep

orte

r Act

ivity

Transfection AssayMouse PXR

Transfection AssayMouse PXR

6

TriolTriol

EC503 µM

5β-cholestane-3α,7α,12α-Triol55ββ--cholestanecholestane--33αα,7,7αα,12,12αα--TriolTriol

Cyp3A4Cyp3A4

HO OH

OHOHOH

55ββ--cholestanecholestane--33αα,7,7αα,12,12αα,25,25--TetrolTetrol

Cyp27 KO: Triol Accumulation & PXR Activation

OH OH

OH

Cyp27Cyp27TriolTriol Cholic AcidCholic Acid

0

25

50

75

Min

utes

Tribromoethanol Sleep Time

Tribromoethanol Tribromoethanol Sleep TimeSleep Time

Cyp27Cyp27--//--

WTWT

Cyp2CCyp2C

OATP2OATP2

GAPDHGAPDH

Cyp3A11Cyp3A11

+/+ -/-Cyp27:

Hepatic Target GenesHepatic Target Genes

PXRSpecies-specific responses

Human PXRHuman PXR Mouse PXRMouse PXR

29

1

5

9

13

17

21

25

29

Fold

Act

ivat

ion

PCN

PCN

PCN

PCN

Rifa

mpi

cin

Rifa

mpi

cin

Rifa

mpi

cin

Rifa

mpi

cin

33

25

1

5

9

13

17

d Ac

tivat

ion 21

Fol

Widespread Use of Dietary Supplements

Widespread Use of Dietary Supplements

Widespread Use of Dietary Supplements

Future Research Needs & ResourcesPredict and prevent herbal-drug interactions

Require comprehensive study of herbal meds for effects on:Require comprehensive study of herbal meds for effects on:• PXR activation• Endogenous PXR ligands• Expression of genes involved in drug clearance• Activity of P450 enzymes and transporters• Drug clearance in humanized mice• Human trials using non-invasive measures of drug clearance• Similar studies on related receptor CAR

Teach clinicians & patientsTeach clinicians & patients• Natural product ≠ Safety • Medical histories should document use of drug supplements • Create online database• Modify supplement use is counterindicated• FDA-like regulation? Standardize potency, efficacy, safety?

Explanation of Explanation of interindividualinterindividual differences in drug clearance?differences in drug clearance?• Polymoprphisms in PXR• Polymorphisms in CAR

Ex- Lab MembersJenny ChenHaibo Wang

Hye-Dong Yoo

Research AssociatesMing Fan

Michael HoKevin Hollister

Min LinIrina Ianculescu

Karol RostamianiEric Wang

Ex- Lab MembersJenny ChenHaibo Wang

Hye-Dong Yoo

Research AssociatesMing Fan

Michael HoKevin Hollister

Min LinIrina Ianculescu

Karol RostamianiEric Wang

CoH CollaborationsTom Balon

Larry SowersTim Synold

Mark ShermanJohn Termini

Outside CollaborationsR. Chandraratna (Allergan)

S. Erickson (UCSF)G. Salen (UMDNJ)V. LaMorte (UCI)

M. Stemerman (UCR)J. Shyy (UCR)

F. Sladek (UCR)

CoH CollaborationsTom Balon

Larry SowersTim Synold

Mark ShermanJohn Termini

Outside CollaborationsR. Chandraratna (Allergan)

S. Erickson (UCSF)G. Salen (UMDNJ)V. LaMorte (UCI)

M. Stemerman (UCR)J. Shyy (UCR)

F. Sladek (UCR)

City of HopeCity of Hope

FellowsGregg Van CittersIsabelle Dussault

Lily LaiShyamali MallickKohei ShimadaCorinne Solier

Xiaolei XuDonna Yu

Xiaohui Yuan

FellowsGregg Van CittersIsabelle Dussault

Lily LaiShyamali MallickKohei ShimadaCorinne Solier

Xiaolei XuDonna Yu

Xiaohui Yuan

National Center for Complementary and Alternative MedicineNational Institutes of Health

National Center for Complementary and Alternative MedicineNational Institutes of Health