Important Genotypes in Drug- Enzyme Interactions Nissa Abbasi-Shaffer University of Washington School of Pharmacy Doctoral Candidate, 2009 August 1, 2008.

Important Genotypes in Drug-Enzyme Interactions

Important Genotypes in Drug-Enzyme Interactions

Nissa Abbasi-ShafferUniversity of Washington

School of PharmacyDoctoral Candidate, 2009

August 1, 2008

Nissa Abbasi-ShafferUniversity of Washington

School of PharmacyDoctoral Candidate, 2009

August 1, 2008

OutlineOutline

• Introduction

• Genotypes in GeneMedRx

• Additions to GeneMedRx

• Introduction

• Genotypes in GeneMedRx

• Additions to GeneMedRx

Drug-Enzyme InteractionsDrug-Enzyme Interactions

The activity an enzyme will have on a substrate is determined in part by the physical enzyme-substrate interaction

(Nonsynonymous) Mutations in genes can affect the protein product interaction: Conformational changes pH changes H-bonding changes

Patient response to drugs can be altered by their genotype

The activity an enzyme will have on a substrate is determined in part by the physical enzyme-substrate interaction

(Nonsynonymous) Mutations in genes can affect the protein product interaction: Conformational changes pH changes H-bonding changes

Patient response to drugs can be altered by their genotype

Genotypes in GeneMedRxGenotypes in GeneMedRx

Function in GeneMedRx for genotype selection

Users can select appropriate genotype and medication profile to determine if an “interaction” exists: Metabolic enzymes:

Reduced efficacy: eg. CYP2D6 UM Increased toxicity: eg. CYP2C9 PM

Target sites: Efficacy: eg. Her2/Neu Toxicity?

Function in GeneMedRx for genotype selection

Users can select appropriate genotype and medication profile to determine if an “interaction” exists: Metabolic enzymes:

Reduced efficacy: eg. CYP2D6 UM Increased toxicity: eg. CYP2C9 PM

Target sites: Efficacy: eg. Her2/Neu Toxicity?

Updated GenesUpdated Genes

CYP2D6 CYP2C9 CYP2C19 NAT2 TPMT UGT1A1

CYP2D6 CYP2C9 CYP2C19 NAT2 TPMT UGT1A1

Each phenotype was given a descriptor so that if selected, regardless of whether a known drug interaction exists, the significance of the genotype is displayed

e.g. CYP2D6 Poor Metabolizer updated note

e.g. CYP2D6 Poor Metabolizer updated note

Gene: 2D6 Poor MetabolizerCytochrome P450 2D6 Poor Metabolizers (PM) have absent or

greatly reduced CYP2D6 activity. They are identified by DNA testing and comprise up to 12% of the population. Drugs that are substrates of 2D6 will reach higher levels in 2D6 PMs and may cause toxicity. The increase is often substantial, roughly equal to adding a strong 2D6 inhibitor drug. Poor metabolizers may need to be started at 20-60% of standard doses for drugs metabolized by 2D6.

Prodrugs that require activation by CYP2D6 may be less effective in PMs, including: tamoxifen, codeine, tramadol, risperidone and encainide.

Recommendations for 2D6 genotype-based changes in dosage for many psychotropic drugs can be found at: http://www.healthanddna.com/professional/depression.html

Gene: 2D6 Poor MetabolizerCytochrome P450 2D6 Poor Metabolizers (PM) have absent or

greatly reduced CYP2D6 activity. They are identified by DNA testing and comprise up to 12% of the population. Drugs that are substrates of 2D6 will reach higher levels in 2D6 PMs and may cause toxicity. The increase is often substantial, roughly equal to adding a strong 2D6 inhibitor drug. Poor metabolizers may need to be started at 20-60% of standard doses for drugs metabolized by 2D6.

Prodrugs that require activation by CYP2D6 may be less effective in PMs, including: tamoxifen, codeine, tramadol, risperidone and encainide.

Recommendations for 2D6 genotype-based changes in dosage for many psychotropic drugs can be found at: http://www.healthanddna.com/professional/depression.html

New Genotypes AddedNew Genotypes Added

MTHFR DPYD HLA-B*5701 HER2/NEU 5HTT

MTHFR DPYD HLA-B*5701 HER2/NEU 5HTT

MTHFRMTHFR Methylene tetrahydrofolate reductase is an

important enzyme in folate metabolism Allows for methyl group transfer needed for DNA

synthesis and methylation Reduced activity is associated with homocysteine

accumulation

Folate status may be associated with malignancy (reduced -CH3)

Position C677T mutation results in loss-of-function of reductase enzyme T/T 70% reduction: seen in ~10% of North Americans C/T 40% reduction: seen in ~40% of North Americans One study reports 677T allele frequencies of:

Caucasians 0.34 Japanese 0.42 Africans 0.08

Methylene tetrahydrofolate reductase is an important enzyme in folate metabolism Allows for methyl group transfer needed for DNA

synthesis and methylation Reduced activity is associated with homocysteine

accumulation

Folate status may be associated with malignancy (reduced -CH3)

Position C677T mutation results in loss-of-function of reductase enzyme T/T 70% reduction: seen in ~10% of North Americans C/T 40% reduction: seen in ~40% of North Americans One study reports 677T allele frequencies of:

Caucasians 0.34 Japanese 0.42 Africans 0.08

Methotrexate Methotrexate

Folate analog that inhibits dihydrofolate reductase (also important in folate metabolism)

MTX used to treat many cancers, rheumatoid arthritis & psoriasis

T/T genotype associated with increased toxicity: Fever, malaise, GI, skin, pharyngeal, pulmonary mucositis & extended thrombocytopenia reported in

leukemia pts

Genotype is important in predicting response

Folate analog that inhibits dihydrofolate reductase (also important in folate metabolism)

MTX used to treat many cancers, rheumatoid arthritis & psoriasis

T/T genotype associated with increased toxicity: Fever, malaise, GI, skin, pharyngeal, pulmonary mucositis & extended thrombocytopenia reported in

leukemia pts

Genotype is important in predicting response

Folate pathwayFolate pathway

Image downloaded from http://www.pharmgkb.org

MTX

XX

DPYDDPYD Dihydropyrimidine dehydrogenase is the

initial and rate-limiting enzyme in pyrimidine (uracil & thymine) catabolism

Exon 14-skipping mutation GT to AT mutation located in the 5’-splicing

consensus sequence Causes exon 14 of DPYD to be spliced out

and not expressed in protein product, DPD 165 bp deletion = 55 amino acids absent Complete loss-of-function

Mutation found in ~1% of European-based population

Dihydropyrimidine dehydrogenase is the initial and rate-limiting enzyme in pyrimidine (uracil & thymine) catabolism

Exon 14-skipping mutation GT to AT mutation located in the 5’-splicing

consensus sequence Causes exon 14 of DPYD to be spliced out

and not expressed in protein product, DPD 165 bp deletion = 55 amino acids absent Complete loss-of-function

Mutation found in ~1% of European-based population

FluorouracilFluorouracil Analog of uracil

DPD is the initial enzyme in its metabolism Reduced DPD activity causes increase 5FU levels

5FU is widely used in cancer therapy (breast, colon, head & neck, etc.)

Toxicity from 5FU is reported to occur in those who have even 50% of DPD activity--one null allele can be enough Toxicity can be severe, including mucositis,

granulocytopenia In one study 24% of patients with WHO grade IV toxicity were

found to have the exon 14-skipping mutation PM will need dose reduction

Analog of uracil DPD is the initial enzyme in its metabolism Reduced DPD activity causes increase 5FU levels

5FU is widely used in cancer therapy (breast, colon, head & neck, etc.)

Toxicity from 5FU is reported to occur in those who have even 50% of DPD activity--one null allele can be enough Toxicity can be severe, including mucositis,

granulocytopenia In one study 24% of patients with WHO grade IV toxicity were

found to have the exon 14-skipping mutation PM will need dose reduction

HLA-B*5701HLA-B*5701 The HLA class I molecules present antigens to killer T cells

to induce immunity Some allotypes within this class have a propensity to induce

allergic-type reactions The HLA-B*5701 genotype is associated with allergic reactions to

abacavir Found in ~5% of the European-based population (less common in

African Americans)

Abacavir Hypersensitivity Syndrome (AHS) Graft vs host-like response, generally appears within 6 wks Symptoms can include rash, fever, malaise, nausea, vomiting,

diarrhea, respiratory problems, and can be fatal A reaction requires immediate discontinuation of the drug; do not

rechallenge

The HLA class I molecules present antigens to killer T cells to induce immunity Some allotypes within this class have a propensity to induce

allergic-type reactions The HLA-B*5701 genotype is associated with allergic reactions to

abacavir Found in ~5% of the European-based population (less common in

African Americans)

Abacavir Hypersensitivity Syndrome (AHS) Graft vs host-like response, generally appears within 6 wks Symptoms can include rash, fever, malaise, nausea, vomiting,

diarrhea, respiratory problems, and can be fatal A reaction requires immediate discontinuation of the drug; do not

rechallenge

FDA AlertFDA Alert

Last week (7/24/2008) the FDA issued an alert recommending all patients be genotyped before starting or restarting abacavir due to the high correlation of AHS to this drug, the potential severity of the symptoms and the ease of preventing the reaction by DNA testing.

HER2/NeuHER2/Neu HER2 belongs to the Epidermal Growth Factor Receptor

family, associated w/ cellular growth and differentiation

Overexpression of the HER2 protein found in 18-20% of breast cancer resulting in more aggressive tumors

Clinical testing (IHC and FISH) to screen overexpressors determines course of therapy: targeted or not better response to anthracyclines

Current targeted therapies: Trastuzumab (Herceptin) efficacy associated w/ overexpression—

std treatmant for early stage Her2/Neu positive breast cancer (given in combo w/ chemo)

Lapatinib is an oral TKI targeted to Her2/Neu in patients who are overexpressors who failed Herceptin therapy (given in combo with capecitabine)

HER2 belongs to the Epidermal Growth Factor Receptor family, associated w/ cellular growth and differentiation

Overexpression of the HER2 protein found in 18-20% of breast cancer resulting in more aggressive tumors

Clinical testing (IHC and FISH) to screen overexpressors determines course of therapy: targeted or not better response to anthracyclines

Current targeted therapies: Trastuzumab (Herceptin) efficacy associated w/ overexpression—

std treatmant for early stage Her2/Neu positive breast cancer (given in combo w/ chemo)

Lapatinib is an oral TKI targeted to Her2/Neu in patients who are overexpressors who failed Herceptin therapy (given in combo with capecitabine)

5HTT/SLC6A45HTT/SLC6A4 The serotonin transporter (5HTT)

reuptakes serotonin from the synaptic cleft into the presynaptic terminus

5HTT is the target of SSRIs--limiting reuptake allows for more serotonin transmission

Variability in SSRI efficacy within the general population has lead the search for a genetic explanation Discovery of 5HTTLPR polymorphism

The serotonin transporter (5HTT) reuptakes serotonin from the synaptic cleft into the presynaptic terminus

5HTT is the target of SSRIs--limiting reuptake allows for more serotonin transmission

Variability in SSRI efficacy within the general population has lead the search for a genetic explanation Discovery of 5HTTLPR polymorphism

5HTTLPR5HTTLPR The serotonin transporter-linked polymorphic

region is located in the promoter region of 5HTT 44-bp insertion/deletion produces the long (L = 16 repeat

units) and short (S = 14 repeat units) alleles S allele results in 50% of the transcriptional activity as

the L allele ~40% of North Americans carry at least one S allele

(varies from 10-70% globally) Other lengths have been discovered, but very rare in the

population: 15, 18-20, 22 repeated units

S allele is associated with reduced efficacy to SSRIs Hypothesis: if less transporter made, less target for SSRIs Carriers of the S allele often take longer to respond or do

not respond at all Genotyping can be used to identify candidates

The serotonin transporter-linked polymorphic region is located in the promoter region of 5HTT 44-bp insertion/deletion produces the long (L = 16 repeat

units) and short (S = 14 repeat units) alleles S allele results in 50% of the transcriptional activity as

the L allele ~40% of North Americans carry at least one S allele

(varies from 10-70% globally) Other lengths have been discovered, but very rare in the

population: 15, 18-20, 22 repeated units

S allele is associated with reduced efficacy to SSRIs Hypothesis: if less transporter made, less target for SSRIs Carriers of the S allele often take longer to respond or do

not respond at all Genotyping can be used to identify candidates

Thank you Genelex!

Any questions???

Thank you Genelex!

Any questions???

ReferencesReferences MTHFR:Kremer JM. Methotrexate pharmacogenomics. Ann Rheum Dis. 2006 Sep;65(9):1121-3. Hughes LB, Beasley TM, Patel H, et al.Racial or ethnic differences in allele frequencies of single-nucleotide

polymorphisms in the methylenetetrahydrofolate reductase gene and their influence on response to methotrexate in rheumatoid arthritis. Ann Rheum Dis. 2006 Sep;65(9):1213-8

Frosst P, Blom HJ, Milos R, Goyette P, Sheppard CA, Matthews RG, Boers GJ, den Heijer M, Kluijtmans LA, van den Heuvel LP, et al. A candidate genetic risk factor for vascular disease: a common mutation in methylenetetrahydrofolate reductase.Nat Genet. 1995 May;10(1):111-3

DPYP:Tomalik-Scharte D, Lazar A, Fuhr U, Kirchheiner J. The clinical role of genetic polymorphisms in drug-metabolizing

enzymes. Pharmacogenomics J. 2008 Feb;8(1):4-15Maitland ML, Vasisht K, Ratain MJ. TPMT, UGT1A1 and DPYD: genotyping to ensure safer cancer therapy? Trends

Pharmacol Sci. 2006 Aug;27(8):432-7Wei X, McLeod HL, McMurrough J, Gonzalez FJ, Fernandez-Salguero P. Molecular basis of the human dihydropyrimidine

dehydrogenase deficiency and 5-fluorouracil toxicity. J Clin Invest. 1996 Aug 1;98(3):610-5 HLA-B*5701:Saag M, Balu R, Phillips E, et al; Study of Hypersensitivity to Abacavir and Pharmacogenetic Evaluation Study Team.

High sensitivity of human leukocyte antigen-b*5701 as a marker for immunologically confirmed abacavir hypersensitivity in white and black patients. Clin Infect Dis. 2008 Apr 1;46(7):1111-8.

Mallal S, Phillips E, Carosi G, et al; PREDICT-1 Study Team. HLA-B*5701 screening for hypersensitivity to abacavir. N Engl J Med. 2008 Feb 7;358(6):568-79.

Chessman D, Kostenko L, Lethborg T, et al. Human leukocyte antigen class I-restricted activation of CD8+ T cells provides the immunogenetic basis of a systemic drug hypersensitivity. Immunity. 2008 Jun;28(6):822-32.

www.fda.gov/cder/drug/InfoSheets/HCP/abacavirHCP.htm HER2/Neu:Lin A, Rugo HS.The role of trastuzumab in early stage breast cancer: current data and treatment recommendations.

Curr Treat Options Oncol. 2007 Feb;8(1):47-60.Yamauchi H, Hayes D. HER2 and predicting response to therapy in breast cancer. Downloaded from:

www.uptodate.com [accessed 2008 July 29]. 5HTT:Lesch KP, Gutknecht L. Pharmacogenetics of the serotonin transporter. Prog Neuropsychopharmacol Biol Psychiatry.

2005 Jul;29(6):1062-73.Gelernter J, Cubells JF, Kidd JR, Pakstis AJ, Kidd KK. Population studies of polymorphisms of the serotonin transporter

protein gene. Am J Med Genet. 1999 Feb 5;88(1):61-6.Heils A, Teufel A, Petri S, Stöber G, Riederer P, Bengel D, Lesch KP. Allelic variation of human serotonin transporter

gene expression. J Neurochem. 1996 Jun;66(6):2621-4.

MTHFR:Kremer JM. Methotrexate pharmacogenomics. Ann Rheum Dis. 2006 Sep;65(9):1121-3. Hughes LB, Beasley TM, Patel H, et al.Racial or ethnic differences in allele frequencies of single-nucleotide

polymorphisms in the methylenetetrahydrofolate reductase gene and their influence on response to methotrexate in rheumatoid arthritis. Ann Rheum Dis. 2006 Sep;65(9):1213-8

Frosst P, Blom HJ, Milos R, Goyette P, Sheppard CA, Matthews RG, Boers GJ, den Heijer M, Kluijtmans LA, van den Heuvel LP, et al. A candidate genetic risk factor for vascular disease: a common mutation in methylenetetrahydrofolate reductase.Nat Genet. 1995 May;10(1):111-3

DPYP:Tomalik-Scharte D, Lazar A, Fuhr U, Kirchheiner J. The clinical role of genetic polymorphisms in drug-metabolizing

enzymes. Pharmacogenomics J. 2008 Feb;8(1):4-15Maitland ML, Vasisht K, Ratain MJ. TPMT, UGT1A1 and DPYD: genotyping to ensure safer cancer therapy? Trends

Pharmacol Sci. 2006 Aug;27(8):432-7Wei X, McLeod HL, McMurrough J, Gonzalez FJ, Fernandez-Salguero P. Molecular basis of the human dihydropyrimidine

dehydrogenase deficiency and 5-fluorouracil toxicity. J Clin Invest. 1996 Aug 1;98(3):610-5 HLA-B*5701:Saag M, Balu R, Phillips E, et al; Study of Hypersensitivity to Abacavir and Pharmacogenetic Evaluation Study Team.

High sensitivity of human leukocyte antigen-b*5701 as a marker for immunologically confirmed abacavir hypersensitivity in white and black patients. Clin Infect Dis. 2008 Apr 1;46(7):1111-8.

Mallal S, Phillips E, Carosi G, et al; PREDICT-1 Study Team. HLA-B*5701 screening for hypersensitivity to abacavir. N Engl J Med. 2008 Feb 7;358(6):568-79.

Chessman D, Kostenko L, Lethborg T, et al. Human leukocyte antigen class I-restricted activation of CD8+ T cells provides the immunogenetic basis of a systemic drug hypersensitivity. Immunity. 2008 Jun;28(6):822-32.

www.fda.gov/cder/drug/InfoSheets/HCP/abacavirHCP.htm HER2/Neu:Lin A, Rugo HS.The role of trastuzumab in early stage breast cancer: current data and treatment recommendations.

Curr Treat Options Oncol. 2007 Feb;8(1):47-60.Yamauchi H, Hayes D. HER2 and predicting response to therapy in breast cancer. Downloaded from:

www.uptodate.com [accessed 2008 July 29]. 5HTT:Lesch KP, Gutknecht L. Pharmacogenetics of the serotonin transporter. Prog Neuropsychopharmacol Biol Psychiatry.

2005 Jul;29(6):1062-73.Gelernter J, Cubells JF, Kidd JR, Pakstis AJ, Kidd KK. Population studies of polymorphisms of the serotonin transporter

protein gene. Am J Med Genet. 1999 Feb 5;88(1):61-6.Heils A, Teufel A, Petri S, Stöber G, Riederer P, Bengel D, Lesch KP. Allelic variation of human serotonin transporter

gene expression. J Neurochem. 1996 Jun;66(6):2621-4.