SNPs and Cancer

SNPs and Drug Interactions

Transporter

Drug

Absorption in the breast

Drug in breast tissue

Metabolism in the liver

Excretion in the kidney

Drug becomes inactive or toxic

Transportation in the blood

Drug in bloodstream

SNPs A SNPs B

SNPs C SNPs D

SNPs May Be the Solution

KANSER FARMAKOGENTİĞİ

Kemoterapi Rejimi vücut yüzey alanına göre

Neden kanser tedavileri başarısız olabiliyor?

Tümör hücrelerinin ilaca azalmış hassasiyeti, ilaca karşı direnç gelişimi

- doğal veyaprimer rezistans

- kazanılmış veya sekonder rezistans (kemoterapiden sonra da gelişebilir)

Tedavi (Eş zamanlı ilaç kullanımı)

Genetic polimorfizm

- İlaç metabolize eden enzimler

- ilaç hedef molekülleri (enzimler, reseptörler)

Genotyping tests in clinical oncology

I- Gene polymorphisms for toxicity of chemotherapeutics

5-FU,irinotecan, thiopurin

DPD, UDP glucuronyltransferase, thiopurin S methyl

transferase, metabolysing enzymes for purin-pyrimidin

analogs, folat metab. enzymes, CYP enz (CYP2B6, CYP2C8,

CYP3A), transferases, transport proteins, receptors

2- Tests for response to molecular therapeutics

Imatinib- BCR- ABL gene mutations

Rituximab- FCGR3A and FCGR2A gene mutations

Gefitinib- EGFR gene mutations

Trastuzumab- HER2 expression

Cisplatin- ERCC1 ve BRCA1 mutation

Biomarker

Label Context

Representative Label Test Drug

Her2/neu Over-expression

Overexpresion of Her2/neu necessary for selection of patients appropriate for drug therapy 1 Trastuzumab (Herceptin)

UGT1A1 Variants

UGT1A1 mutation in patients, exposure to drug and hence their susceptibility to toxicity 2

Irinotecan

(Campto)

EGFR expression Epidermal Growth Factor Receptor presence or absence- 3

Cetuximab

Gefitinib(Iressa)

Erlotinib (Tarceva)

Examples of Valid Genomic Biomarkers

Kanser ilaçlarının metabolizması ile ilişkili enzimler ve genler

Tiopurin metil transferaz (TMPT)

Dihidropirimidin dehidrogenaz (DPD)

Sitokrom p 450 (CYP)

5,10 metiltetrahidrofolat redüktaz (MTHFR)

N-asetil transferaz (NAT)

UDP glukoroniltransferaz (UGT1 ve UGT2)

Multi-drug direnç geni (MDR1)

DNA hasar onarım geni(ERCC1)

Sulfotransferaz SULT1A1)

Onkolojide kullanılan farmakogenetik testler

GEN İLAÇ SONUÇ TMPT

(Tiopurin S-metil Transferaz)

6-MP

Lösemide etkinlik ve yan etki

DHD

(DihidropirimidinDehidrogenaz)

MTHFR

(5’-10’ metilen tetrahidrofolatreduktaz

TSER(TS)

(Timilat sentaz)

5-FU

5-FU toksisitesi

UGT1A1

(UDP-Glukuronozil Transferaz)

irinotekan

Metabolizma ve toksisite

ERCC1

(Nükleotid eksizyon tamir genleri)

GST

(Glutatyon S-transferazlar)

platinum

Tümör cevabı ve sağkalımda azalma

CYP2D6

(Sitokrom p450 enzimleri)

TMX

(Tamoksifen)

Adjuvan sonuçları etkiler

MTHFR

(5’-10’ metilen tetrahidrofolatreduktaz)

MTX

(Metotreksat)

5-FU

Toksisite riski artar

Kemoterapötiklerin Toksisitesinin Belirlenmesinde Kullanılan Testler ve Kanser Tedavisinde Sıklıkla Kullanılan İlaçları Metabolize Eden Enzimler

1. Pürin-pirimidin analoglarını metabolize eden enzimler

2. Folat metabolizmasında görevli enzimler,

3. Sitokrom P450 enzimleri (CYP 450)

4. Transferazlar,

5. Taşıyıcı proteinler,

6. Reseptörler.

Pürin-pirimidin analoglarını metabolize eden enzimler

Tiopurinler: losemi tedavisinde yaygın olarak kullanılan ilaclardır.

- merkaptopurin: ALL (akut lenfoblastik lösemi)

- Tioguanin: AML (akut myeloblastik lösemi)

- Azotiopürin:immün baskılayıcı (Crohn hastalığının tedavisi)

Tiopurin S-metil transferaz (TPMT)

Tiopürinlrin terapötik etkinliği ve toksisitesinden sorumlu

TPMT genindeki polimorfizmler TPMT enzim aktivitesini azaltır

TPMT aktivite azalması : ciddi, hayatı tehdit eden toksisiteler

TPMT ve Merkaptopurin Tedavisi

6-Mercaptopurin

inaktif ön-ilaç

TPMT 6-MeMP

(inactive)

HP

RT

Thioguanin nucleotids

•Antitumor effect

•Myelosupression

TPMT = Tiyopürin metiltransferaz /

6-MeMP = 6-metil merkaptopürin

HPRT = Hipoksantin fosforibozil transferaz : Biochimica et Biophysica Acta, 1603 (2003)

*Polymorp.

6-MP; ALL tedavisinde kullanılan ön ilaç

6-MP transferaz hypoxantine guanin phosphorybosyl (6 thioguanine oluşur) tarafından aktive olur

TPMT inactivates 6-MP by S methylation

6-MP TPM tarafından S metilasyon ile inaktive olur

Tiopurin metil transferaz

Doğal tipi TPMT1

Alelleri :5 adet aleli mevcut TPMT2,TPMT3A,TPMT3B,TPMT3C, TPMT4

Alellerin 3’ü %95’i kapsar

Kişilerin %10’unda aktivite az, %3’ünde hiç yok

heterozigotlar dozun %65’ini tolere edebilir, homozigotlar toksisiteden ölür.

Bu varyantlar nedeniyle doz indirimi yapılan hastalarda sağkalım= doğal tip alel taşıyıcıları

Genotype-Guided 6-MP Dosing

Pharmacogenomics 2002;3(1):89-98.

Genetic polymorphism of TPMT activity and thiopurine therapy

Krynetski & Evans, Am J Hum Genet 1998 63:11-16

Prevalence of TPMT Genotypes-Three Major SNPs (single gene) Define Mutant Alleles

Newly Diagnosed ALL Patients per Year

30,000

Homozygotes

100

None or Low

TPMT Activity

Heterozygotes

3,300

Intermediate

TPMT Activity

Wild Type

26,600

High

TPMT Activity

Two Mutant Alleles

v/v

One Mutant Allele

wt/v

No Known Mutation

wt/wt

ALL, 6MP and polymorphic TPMT

Children TPMT Toxicity

Children TPMT Poor response

6MP

6MP

Pharmacogenetics: Use in drug delivery

HERCEPTIN

Oncology

Breast Cancer – 25-30% of breast and ovarian tumors overexpress HER2

– Overexpression is correlated with prognosis

– Patients positive for HER2 can be treated with Herceptin

– Herceptin is a monoclonal antibody against the HER2 receptor

– Improved survival

Examples……

pharmacodynamics

Breast Cancer abnormally high amounts of HER2 protein in 30% of patients

Herceptin binds to HER2 slowing tumour growth, 70% of patients do not respond

HER-2 Protein and Herceptin

Herceptin (trastuzumab): – Metastatic breast cancer

– Targets tumor cells that over-express the human epidermal growth factor receptor 2 (HER2) protein

– Best response attained in women who over-express the HER2 protein

– HER-2 over-expression in breast cancer cells should be done before patients receive the drug

Herceptin: Prescribing Information

HERCEPTIN (Trastuzumab) as a single agent is indicated for the treatment of patients with metastatic breast cancer whose tumors overexpress the HER2 protein and who have received one or more chemotherapy regimens for their metastatic disease.

HERCEPTIN should be used in patients whose tumors have been evaluated with an assay validated to predict HER2 protein overexpression

Pharmacogenetics = molecular DD Case Study: Herceptin®

Low HER2

High HER2

Bimodal response:

2/3 of patients: addition of Herceptin® to chemoRx

no benefit

1/3 of patients: addition of Herceptin® to chemoRx

50% survival time increased by factor 1.5 (20 29 weeks)

5 fluorouracil and DPD

5 fluorouracil and DPD

5-FU may cause serious gastrointestinal and

hematologic side effects (3-5%).

5-FU causes grade 4 neutropenia in pts with DPD

deficiency (40-50%)

Polymorphism in 5-fluorouracil (5-FU) Metabolism

(detoksification)

5-FU (activation)

Active metabolit

MTHFR polymorphism

• low MTHFR activity

• 5-FU efficacy increase

TS polymorphism

• TS activity increases

• 5-FU efficacy decreases

DPD polymorphism

• neurologic toxicity

• GI toxicity

• hematologic toxicity

İnactive metabolite

TRENDS in Pharmacological Sciences, 25 (2004)

(TP) thymidin phosphorilase

DPD

MTHFR

TS

prodrug 5-floro 2 deoksiuridin

monofosfat (FdUMP)

Tamoxifen

Pro-drug

widely used treatment in breast cancer patients

inhibits estrogen from binding to its receptor, thus inhibiting estrogen-regulated genes (growth factors and angiogenic factors)

metabolized by cytochrome P450s into N-desmethyltamoxifen (NDM), 4-hydroxytamoxifen (4-OH tamoxifen) and endoxifen

efficacy of tamoxifen therapy among women is heterogeneous due to genetic variants of CYP2D6

On October 18 (2006), a presantation of the study related to Tamoxifen by Dr. Goetz to the FDA led to an advisory committee unanimously recommending a label chance for tamoxifen. This change would include information about the incresaded risk both from genetic factors and drug interactions affecting CYP2D6. The majority of the committee also recommended that CYP2D6 genotype testing as an option for women before they are prescribed tamoxifen.

Jin et al., J. Natl. Cancer Inst. 97:20-39, 2005.

Tamoxifen Biotransformation

Relapse–Free Survival, % Disease–Free Survival

Tamoxifen Pharmacogenomics

Goetz et al., Breast Cancer Res. Treat. 101:113-121, 2007.

Breast Cancer (190 Patients)

Relapse-Free (RF) Probability

by CYP2D6 Genotype

Schroth et al., JCO 25:5187-93, 2007

No Tamoxifen Tamoxifen Tamoxifen

Selective Serotonin Reuptake Inhibitors (SSRIs) & Tamoxifen

antidepressants that are often prescribed to treat hot flashes in women who take tamoxifen

paroxetine, sertraline, citalopram, fluoxetine and venlafaxine

inhibition of CYP2D6 by SSRIs likely to affect metabolism of tamoxifen use of SSRIs associated with a reduced mean plasma concentration of endoxifen (lower response to tamoxifen therapy)

Results

with inhibitor

without inhibitor

Jin et al., 2005

Objectives

examine association between genotype (CYP2D6) and plasma concentrations of tamoxifen and its metabolites

examine effect of CYP2D6 inhibitors on plasma concentrations of endoxifen

examine SSRIs and their association with plasma concentrations of endoxifen

Mary 55 years

PHARMACOGENETICS

Tamoxifen+CYP2D6

CYP2D6 PM CYP2D6*4/*4

!!!

CYP2D6 EM – OK CYP2D6*1/*1

Noeffects, risk of disease relapse

Drug – gene interactions

Improved benefit from tamoxifen

Carol 55 years

Conclusion & Significance

higher plasma endoxifen concentration than 4-OH tamoxifen may mean it has a more important role

efficacy of tamoxifen therapy among women is heterogeneous due to genetic variants of CYP2D6

use of SSRIs associated with a reduced mean plasma concentration of endoxifen (lower response to tamoxifen therapy)

PROJECT Pharmacogenomic and Nutrigenomic Enhancement of CYP2D6 Activity in

Tamoxifen-Resistant Breast Cancer Patients in Turkish, French &Canadian Patients

Presley JF1, Bernard-Gallon D2, Genç E3, Kilic U3, Dejgaard SY1, Yeliz Demirci3, Bignon YJ2, Hizel C 1,

2, 3 ,4

Study Objective/ Hypothesis

to develop an integrated pharmacogenomic & nurigenomic approach to tamoxifen therapy

to gain the capability to significantly improve this outcome with targeted and inexpensive nutritional modification which can alter the expression level of CYP2D6 in IMs.

Hypothesis

nutritional interventions that improve metabolism of tamoxifen will significantly improve survival rates of IM breast cancer patients.

1)Department of Anatomy and Cell Biology, McGill University, Montreal, Quebec, Canada H3A2B2, 2) Département d'Oncogénétique du Centre Jean Perrin, 63011 Clermont-Ferrand Cedex 01, France 3)Department of Biochemistry, Faculty of Medicine, Yeditepe University, Istanbul,Turkey. 4) C2H-Vichy Genomics, Vichy,France

References

Garber, K. (2005) J. Natl. Cancer Inst. 97: 412-413.

Jin, Y. et al. (2005) J. Natl. Cancer Inst. 97: 30-39.

Osborne, C.K. (1998) N. Eng. J. Med. 339: 1609-1618.

Stearns, V. et al. (2003) J. Natl. Cancer Inst. 95: 1758-1764.

Irinotecan (CPT-11)

Pro-drug

FDA approved & widely used in treatment of metastatic colorectal cancer

Topoisomerase I inhibitor

Complex metabolic pathway

Wide interindividual variability in drug response and in occurrence of toxic side-effects (severe diarrhea, neutropenia, myelosuppression).

Polymorphisms that affect irinotecan therapy

Relling & Dervieux, Nature Cancer Reviews 1:99-108

Genotype Versus Phenotype

Genotype and Phenotype

_______________________ Genotype Glucuronidation

6/6 851±545

6/7 699±361

7/7 199±118

_______________________

Mean ± Standard Deviation

Fisher et al. Pharmacogenetics, 2000

UGT1A1*28 Genotype and Estradiol

Glucuronidation

0

200

400

600

800

1000

1200

1400

1600

6/6 6/7 7/7

Genotype G

lucu

ron

ide F

orm

ati

on

Ra

te

Current Understanding of PGx and Neutropenia

Group Prevalence Risk of Toxicity

All Patients ----- 10%

Patients That Are 7/7

10% 50%

Patients That Are 6/7

40% 12.5%

Patients That Are 6/6

50% 0%

Based on data from Innocenti et al (2004)

Potential of UGT Testing to Inform Dosing

All Patients with Same Diagnosis

PGx profile for high risk (50%): treat with

alternative drug or dose

PGx Profile for moderate risk (12.5%): treat with alternative drug or dose

PGx Profile for low risk (0%): treat with

conventional dose

Conclusions “UGT1A1 genotype (UGT1A1*28)and total bilirubin levels are strongly associated with

severe diarrhea & neutropenia.”

UGT 1A1*28 is a variant allele

» Variation in the TA repeats in the promoter region Normal allele: 6 TA repeats (6/6)

Variant allele: 7 TA repeats (7/7)

UGT 1A1*28 is associated with reduced gene expression and reduced glucuronidation in human liver microsomes.

Grade 4 neutropenia is more common (p=0.001) in patients with 7/7 genotype patients

A strong correlation between genotype and diarrhea (P=0.01)

Utility of UGT1A1 Polymorphism in the Use of Irinotecan

Excluding 7/7 patients from standard dose of irinotecan treatment will reduce the overall incidence of grade 4 neutropenia from 10.1% to 5.7%.

7/7 genotype patients may get a alternate irinotecan regimen?

Genotypic testing combined with bilirubin levels will allow better patient selection for Irinotecan therapy. Physicians will be better informed about use of Irinotecan as a single agent or in combination therapy.

Genotypic testing will allow 7/7 genotype patients with a choice for equally efficacious alternate therapy.

6-Mercaptopurine (6MP) and Childhood Acute Lymphoblastic Leukemia (ALL)

ALL is a life-threatening disease and 6MP can cause life-threatening toxicities

Dose titration (dosing size, duration and intensity) is major determinant of long-term EFS and myelosuppressive effects

6MP is metabolized to pharmacologically active thiopurine nucleotides by thiopurine methyltransferase (TPMT)

TPMT activity shows trimodal variation in the general population

TPMT Genetic Polymorphism

Well-documented, causal link between TPMT polymorphism and clinical effects, including toxicity

Genotypes with reduced (10% of the population) or no (0.3% of the population) activity are at a substantially increased risk of myelosuppression and secondary cancer

Pharmacogenetic tests are available and feasible to use for identifying these patients and guide optimal dosing

CYP2D6 and Metabolism of Tamoxifen

CYP2D6 enzyme activity directly

related to:

• Common genetic polymorphisms

• Concomitant medications that

inhibit CYP2D6 activity