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Tamoxifen And CYP2D6: Using Pharmacogenetics to discover a new drug

Tamoxifen And CYP2D6: Using Pharmacogenetics to discover a new drug

Matthew P. Goetz, MDAssociate Professor of Oncology

Mayo Clinic

CP1229323-1

Ohio State University Center: Conference on Personalized Health Care

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Current Applications of Pharmacogenomics in Breast Cancer

• Current state of adjuvant hormonal therapy

• Pharmacogenetics: CYP2D6 and tamoxifen

• Future directions: endoxifen as a novel hormonal therapy for breast cancer

• Current state of adjuvant hormonal therapy

• Pharmacogenetics: CYP2D6 and tamoxifen

• Future directions: endoxifen as a novel hormonal therapy for breast cancer

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Breast Cancer2008 (USA)

• Invasive breast cancer: 178,480new cases

• Ductal carcinoma in situ: 62,030 cases

• 2/3 are estrogen positive — candidates for hormonal therapy

American Cancer Society: Breast Cancer Facts & Figures, 2007-2008

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0 5 10 0 5 10

Oxford Overview: 5 Years of Tamoxifen vs NotER Positive vs ER Negative

Oxford Overview: 5 Years of Tamoxifen vs NotER Positive vs ER Negative

5 Years Tamoxifen vsNot Recurrence

ER+/ER UnknownEntry age 50

5 Years Tamoxifen vsNot Recurrence

ER+/ER UnknownEntry age 50

5 Years Tamoxifen vs NotRecurrence

ER – Poor PR – Poor

5 Years Tamoxifen vs NotRecurrence

ER – Poor PR – Poor

Re

cu

rren

ce

Re

cu

rren

ce

YearsYears

50

40

30

20

10

0

Control40.3%

Control40.3%

5-yr TAM23.5%

5-yr TAM23.5%

10-yr gain 15.0% (SE 1.1)Logrank 2P<0.0000110-yr gain 15.0% (SE 1.1)Logrank 2P<0.00001

28.228.2

15.115.1

5-yr TAM29.0%

5-yr TAM29.0%

21.321.3

20.820.8

YearsYears

Loss 1.9% (SE 1.5)Logrank 2P>0.1; NSLoss 1.9% (SE 1.5)Logrank 2P>0.1; NS

Control27.1%

Control27.1%

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Cohort 1

Direct Comparison of Tamoxifen and an Aromatase

Inhibitor (9,856 Patients)

Ingle et al. SABC 2008

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Cohort 1

5 years of AI vs

tamoxifen

ER+

10

0

20

30

40

50

% SE

% SE

0 5 8Years

15.3%AI

12.6%

9.6%

5-yr gain 2.9% (SE 0.7)8-yr gain 3.9% (SE 1.0)

Logrank 2P<0.00001

Years 0-1 2-4 ≥5

AI 1.69 (163/9647) 2.31 (261/11297) 2.33 (160/6879)

Tamoxifen 2.46 (234/9510) 2.81 (307/10938) 2.78 (180/6478)

Rate ratio, 0.67 SE 0.08 0.81 SE 0.08 0.83 SE 0.10 from (O-E)/V -38.4/96.6 -29.5/137.9 -15.7/83.0

Tamoxifen19.2%

Recurrence

Ingle et al. SABC 2008

3013063-7Jin Y et al: J Natl Cancer Inst 97:30, 2005

Tamoxifen Metabolic Pathway (Humans)

CP1230355-21

400-600 nM

5-10 nM

20-180 nM

200-300 nM

Binding affinity relative to estradiol

1.00 for metabolites 4-OH TAM and endoxifen 0.01 for tamoxifen and N-desmethyl tamoxifen

chromosome 22 CYP2D8 (pseudogene)

CYP2D7 (pseudogene)

CYP2D6

functional alleles*1, *2, *35

nonfunctional (null, *0)

*3, *4, *5, *6, *7, *8, *12,*13, *14, *15, *16, *18, *19, *20, *21, *38 ....

CYP2D gene locus

Molecular Basis of the CYP2D6 Polymorphism

> 100 Genetic Variants duplicated alleles*1x2, *2x2, *35x2

low function alleles*9, *10, *17, *41, *59

q13.1

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0

20

40

60

80

100

120

140

160

180

Wt/Wt Wt/*4 *4/*4

CYP2D6 Genotype and Endoxifen

Jin Y et al: J Natl Cancer Inst 97:30, 2005

CYP2D6*4 (most common genetic variant associated with the CYP2D6 poor metabolizer state)

P<0.001, r2=0.24

Plasmaendoxifen

(nM)

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5 years oftamoxifen(20 mg qd)+ 1 year of

fluoxymesterone(10 mg po bid)

5 years oftamoxifen(20 mg qd)+ 1 year of

fluoxymesterone(10 mg po bid)

5 years of tamoxifen5 years of tamoxifen

NCCTG 89-30-52NCCTG 89-30-52

RRAANNDDOOMMIIZZAATTIIOONN

Postmenopausal women

Postmenopausal women

5 years total therapy5 years total therapy

541 women accrued

541 women accrued

Early ER+

breast cancerEarly ER+

breast cancer

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Time to Recurrence According to CYP2D6 MetabolizerStatus* in Women Receiving Adjuvant Tamoxifen

0

20

40

60

80

100

0 2 4 6 8 10 12 14

%

Years after randomization

P<0.001P<0.001

EM/EM, EM/IMEM/EM, EM/IMEM/PM, IM/IM, PM/IMEM/PM, IM/IM, PM/IMPM/PMPM/PM

(n=108)(n=108)

(n=65)(n=65)

(n=16) (n=16)

PM alleles: *3,*4,*6,

IM alleles: *10, *17, *41

Potent CYP2D6 inhibitor = PM/PM

Goetz et al J Clin Oncol. 2005;23(36):9312-8.Goetz M et al. Breast Cancer Res Treat 101:113-121, 2007Goetz et al J Clin Oncol. 2005;23(36):9312-8.Goetz M et al. Breast Cancer Res Treat 101:113-121, 2007

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ABCSG Trial 8 structure

Primarysurgery

Randomize

Anastrozole (3 years)

Switching period

Sequencing periodJakesz R et al. Lancet 2005

Switch point

Tamoxifen

(2 years)

Tamoxifen(3 years)

Tamoxifen

(2 years)

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Event-free survival following adjuvant therapy switch (n = 2529)

0

75

80

85

90

95

100

0 12 24 36 48 60 72

Time after switch (months)

EFS (%)

Anastrozole (A)Tamoxifen (T)

HR

0.62Events

p-value

0.011

T

69

A

44

Therapy switch(2 years after surgery)

92.8%

88.9%

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ABCSG Trial 8 structure

Primarysurgery

Randomize

Anastrozole (3 years)

Switching period

Sequencing periodJakesz R et al. Lancet 2005

Switch point

Tamoxifen

(2 years)

Tamoxifen(3 years)

Tamoxifen

(2 years)

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Arm A: Tamoxifen for 5 years (n=67 cases)

Risk relative toextensive metabolizers P value

CYP2D6 PM 3.83 (1.27-11.55) 0.017

CYP2D6 IM 0.87 (0.44-1.71) 0.689

CP1229323-18

Arm B: Tamoxifen to anastrozole (n=55 cases)

Risk relative to extensive metabolizers P value

CYP2D6 PM 1.02 (0.21-4.83) 0.985

CYP2D6 IM 0.81 (0.40-1.61) 0.538

CYP2D6 and Relative Risk of Breast Event

Goetz et al. SABC 2008

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Arm A: Tamoxifen Years 3-5 (n=55 cases)

Risk relative toextensive metabolizers P value

CYP2D6 PM 2.81 (0.88-8.97) 0.081

CYP2D6 IM 0.75 (0.36-1.55) 0.431

CP1229323-18

Arm B: Anastrozole Years 3-5 (n=31 cases)

CYP2D6 and Relative Risk of Breast Event

Risk relative toextensive metabolizers P value

CYP2D6 PM 0.71 (0.06-8.39) 0.782

CYP2D6 IM 0.57 (0.21-1.54) 0.269

Goetz et al. SABC 2008

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Mayo Clinic Rochester: Treatment of Postmenopausal, ER + Breast Cancer

Women should be informed 1) data in support of CYP2D6 as key

enzyme involved in the activation of tamoxifen

2) Discontinuation of potent CYP2D6 inhibitors

3) CYP2D6 genotyping: CYP2D6 PM do not receive tamoxifen

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New identified Mechanism of Action (Endoxifen)

• Does endoxifen differ from Tamoxifen or 4-HT in its interaction with the estrogen receptor?

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Tamoxifen Mechanism of Action

ER

ER

ERER

ER

SRC

D1

Coregulators

Transcription

GeneERE

Tamoxifen

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Endoxifen but not 4HT Results in Increased ERα Protein Turnover

Wu et al: Cancer Res, 2009

3013063-21Jin Y et al: J Natl Cancer Inst 97:30, 2005

Tamoxifen Metabolic Pathway (Humans)

CP1230355-21

400-600 nM

5-10 nM

20-180 nM

200-300 nM

Binding affinity relative to estradiol

1.00 for metabolites 4-OH TAM and endoxifen 0.01 for tamoxifen and N-desmethyl tamoxifen

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High Concentrations of EndoxifenSuppress MCF-7 Cell Proliferation at Clinically

Relevant Conditions

Wu et al: Cancer Res, 2009

0.0

0.5

1.0

1.5

2.0

2.5

3.0

3.5

4.0

Re

lati

ve

fo

ld c

ha

ng

efr

om

ve

hic

le

Vehicle E2 E2+com-

bination(TAM+4HT

+NDT)

E2++20 nM

endoxifen

E2+com-

bination+40 nM

endoxifen

E2+com-

bination+100 nM

endoxifen

E2+com-

bination+1,000 nMendoxifen

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True Personalized Medicine: You need to account for all sources of

variation: germ line, tumor, environmental

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van't Veer LJ et al: J Clin Oncol 23:1631, 2005van't Veer LJ et al: J Clin Oncol 23:1631, 2005

Gene Expression Profiling

Unfixed sample of tumor tissueUnfixed sample of tumor tissue

Surgical removal of tumor tissue

Surgical removal of tumor tissue

Labeled tumor cDNA or cRNALabeled tumor cDNA or cRNA

Tumor RNATumor RNA

Comparative analysis

of gene expression

Molecular signatureMolecular signature

Poor Poor prognosisprognosis

Good Good prognosisprognosis

Labeled control cDNA or cRNA

Labeled control cDNA or cRNA

DNA microarray

DNA microarray

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Sorlie et al: Proc Natl Acad Sci USA 100:8418, 2003Sorlie et al: Proc Natl Acad Sci USA 100:8418, 2003

Gene Expression ProfilingIdentifies Molecularly Distinct Subtypes

Gene Expression ProfilingIdentifies Molecularly Distinct Subtypes

AA BB

CC

DD

EE

FF

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Overexpression of HER-2 in Human Breast Cancer Cells

Human breast cancer cells

Transfect withHER2 gene

DNA synthesisCell growthrateGrowth in soft agar

TumorigenicityMetastaticpotential

Transformed breast cancer cell

Multiple copies of HER2,

high expressor

Single copy of HER2,

low expressor

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Tamoxifen Stimulates the in Vivo Growth ofHER-2 Expressing, ER+ Xenographs

Shou J et al:J Natl Cancer Inst 96:926, 2004

P-MAPKTHr202/Tyr204)

T-MAPK

E2 TAM -E2

1 9030 60

Tumorvolume(mm3)

Days

1,400

1,000

200

0

600

-E2

TAME2

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Tamoxifen, Endoxifen in MCF-7 HER2 expressing Breast Cancer Cells

MCF7-HER2

0

20

40

60

80

100

120

140

160

1 10 100 1000

Drug concentration (nM)

% o

f C

on

tro

l (D

MS

O o

nly

)

TAM

END

Goetz, Reinicke, Ames et al. unpublished

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Tamoxifen versus Endoxifen in HER-2 positive cells

• Does CYP2D6 status (metabolic activation to endoxifen) impact the outcomes of patients with HER-2 positive breast cancer?

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5 years oftamoxifen(20 mg qd)+ 1 year of

fluoxymesterone(10 mg po bid)

5 years oftamoxifen(20 mg qd)+ 1 year of

fluoxymesterone(10 mg po bid)

5 years of tamoxifen5 years of tamoxifen

NCCTG 89-30-52NCCTG 89-30-52

RRAANNDDOOMMIIZZAATTIIOONN

Postmenopausal women

Postmenopausal women

5 years total therapy5 years total therapy

541 women accrued

541 women accrued

Early ER+

breast cancerEarly ER+

breast cancer

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HER-2, CYP2D6 and Recurrence Free-Survival

0

20

40

60

80

100

0 2 4 6 8 10 12 14

%%

Years from randomizationYears from randomization

EM/EM, EM/IMEM/PM, IM/IM, PM/IM, PM/PMP=0.006

EM/EM, EM/IMEM/PM, IM/IM, PM/IM, PM/PMP=0.006

n=9n=9

n=10n=10

CP1347559-1

PM alleles: *3, *4, *6IM alleles: *10, *17, *41Potent CYP2D6 inhibitor = PM/PM

PM alleles: *3, *4, *6IM alleles: *10, *17, *41Potent CYP2D6 inhibitor = PM/PM

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Pharmacology 101

• Which tamoxifen molecule is occupying the estrogen receptor – tamoxifen, 4-hydroxy tamoxifen, endoxifen?

• Who cares?

• Pharmacology is boring – if the drug doesn’t work, just use a different drug

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Summary

• The metabolic activation of tamoxifen precludes use for many women

• In HER2 expressing breast cancer, tamoxifen activates but endoxifen potently inhibits tumor growth

• Administration of endoxifen would bypass the genetic and concomitant drug effects on metabolism

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Murine Endoxifen PlasmaConcentrations After PO Adminstration

1

10

100

1,000

10,000

0 240 480 720 960 1,200

En

do

xife

n (

nM

)E

nd

oxi

fen

(n

M)

MinutesMinutes

4 mg/kg4 mg/kg

200 mg/kg200 mg/kg

Reid, JR, Goetz, MP, Ames MM

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• How do we develop Endoxifen?• No intellectual property, thus no

drug company interest• Can NCI facilitate the development

of endoxifen?

Next Steps

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Endoxifen Drug Development Timeline

2009 2010 2010-2011 2012-2015

Precl

inic

al p

harm

acolo

gy

and to

xico

logy

in m

ice

NCI and M

ayo complete preclin

ical

Toxicology; clin

ical gra

de drug availa

ble

Phase II stu

dies begin

Human p

hase I stu

dy begin

s

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Acknowledgments

Matthew Ames LabJoel Reid Katherine ReinickeStephanie SafgrenMary Kuffel Sarah Buhrow

James Ingle Vera J. Suman

Tom Spelsberg labJohn HawseXiaglin WuMalayannan Subramaniam

Richard Weinshilboum labAnn Moyer

OthersRobert JenkinsCarol Reynolds

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• Mayo Clinic Cancer Center

• North Central Cancer Treatment Group

• ABCSG

• Mayo Clinic Cancer Center

• North Central Cancer Treatment Group

• ABCSG

• 1R01CA133049-01 (Goetz)

• Mayo Clinic Breast Cancer SPORE

• K-12 (Paul Calabresi Scholar)

• Pharmacogenetics Research Network (Dick Weinshilboum)