Pharmsci Exam 3

Factors Affecting Drug Activity/Toxicity

Age

Sex

Health

Underlying disease status (hepatic/renal function, etc.)

Drug-drug interactions

Inherited differences!

Pharmacogenetics (PGt)

 Pharmacology + genetics

 Defined as the effect of variations in a single gene on drug therapy

Pharmacogenomics (PGx)

 Pharmacology + genomics

 Defined as the effect of variations in an entire genome on drug therapy

Traditional Medicine

Treatment is based on a standard regimen

 “One dose fits all”

 Drug/dose may be based on age, weight, etc.

 Adjust dose depending on patient’s response e.g., warfarin

Precision (Personalized) Medicine

• Treatment decisions also reflect a person’s unique genetic profile to improve their response to drugs

Goals

• The right drug

• The right dose

Precision Medicine Benefits to patients

• Lower number of meds needed to reach desire therapeutic response

• Reduced treatment time

Fewer adverse drug reactions (ADRs)

Reduced effect of disease on the body

Improved health!

precision medecine benefits to industry

Fewer failed drug trials

Reduced time for drug development/approval

Lower costs!

Precision Medicine and the FDA

FDA is responsible for monitoring drug safety

• FDA Precision Medicine website

• PGx info included in labels of many drugs (can incl. recommendations for dosing, genetic testing, etc.)

• Maintains database of drug-gene interactions (includes drugs, genes, phenotypes & description of interactions)

Roles of Pharmacists in Precision Medicine before testing

• ID eligible patients for testing

• Patient education & informed consent

• Obtain authorization for testing from HC provider

• Obtain patient samples

Roles of Pharmacists in Precision Medicine after testing

• Counsel patients on test results

• Consult with HC provider & discuss recommendations

• Review PGx results for new meds

Genotype

Individual’s genetic makeup

Phenotype

Observable features

Eye/hair color, height, sex

Certain disease (e.g. cancers)

Drug response, etc.

Transcription

DNA (genotype)→ RNA

Translation

RNA → protein (phenotype)

Changes in DNA

Individual bases (SNP)

Insertions/deletions (Indels)

Copy number variants

GENOTYPE ALTERED

Chnages in protein

Alterations in amino acid seuence increase or decrease…

• expression levels

• activity

• stability

Phenotype may or may not be altered

Mendelian Genetics

One copy of an allele (alternate form of a gene) is inherited from each parent

Both alleles contribute to phenotype in PGx!

Allele

Versions of a gene

• e.g., D, E, E*, F, F*

Genotype

Both copies of an allele

( D/D, E/E*, F/F*)

Haplotype

Variants on one chromosome that are inherited together

Nucleic acid bases

Purines- adenine (A), guanine (G)

Pyrimidines- cytosine (C), thymine (T) Uracil (U)

Human genome consists of

3 billion base pairs

• 99.9% identical!

Genes spread out over 46 chromosomes

• Humans are diploid

• 44 autosomes (22 × 2) + XX (female) or XY (male)

How many genes do humans have?

20 k

Exons (region of a gene)

expressed sequences → coding (determines amino acid sequence)

Introns (region of a gene)

= intervening sequences → non-coding

Promoter (region of a gene)

is non coding

 5’-UTR & 3’-UTR=

 untranslated regions → non-coding

at beginning (5) and at end (3) of gene

Role of RNA Splicing in Genetics

• DNA transcribed to hnRNA (pre-mRNA)

• Introns spliced out to produce mRNA (exons left)

• Poly A tail added to end of mRNA

• mRNA undergoes translation to protein

Alternative RNA Splicing

 Exons recombined to form alternative mRNAs during splicing

• Multiple proteins from a single gene! (e.g., UGT1A gene)

mRNA Codons (Translation)

64 codons

• 4 RNA bases used in combos of 3 (UUU, ACU, GUU)

64 codons but translate to only 20 amino acids

Start Codon

AUG

Stop Codon

UAA, UAG, UGA

Genetic Polymorphisms

DNA sequence variation (mutation)that occurs in a population

Types of genetic variants

• Single nucleotide polymorphisms (SNPs)

• Insertions-deletions (Indels)

• Copy number variants (CNVs)

Single Nucleotide Polymorphisms (SNPs)

Most common genetic variant

 Est’d to occur every 1,000-1,500 base pairs

SNP (DNA) may alter codon sequence (RNA)

Synonymous SNP

 Unaltered amino acid sequence

Non-synonymous SNP

Altered amino acid sequence

Truncated protein - stop codon variant

Wild type (wt)

most common DNA gene sequence in a population

Intronic SNP’s

Base alteration occurs in intron sequence

• Copied into hnRNA

• Splicing error may occur

• Intron may be translated

• Protein seq. altered

Promoter SNPs

Base alteration occurs in promoter sequence (start of gene)

• Gene transcription may be affected

• Protein expression altered

Indels

Insertion or deletion of bases

insertion adds base and pushes over rest of sequence

Deletion removes a base and pulls back rest of sequence

Protein sewuence alteres

CNVs (Gene Duplication)

Multiple copies of a gene are present

 e.g. CYP2D6 gene

Protein is overexpressed

CNVs (Gene Deletion)

Section of DNA with gene is deleted

e.g., CYP2D6 gene

No protein expressed!

Genetic Testing info

 Unknown genotype may be assigned as “wild- type”

 Info is “permanent” in patient’s history

Genetic testing Clinical tests

 Ordered by a healthcare provider

• Specific medical reason

• Response to medication = “pharmacogenomics” testing

• Results interpreted by healthcare professional

Genetic testing Direct-to-consumer tests

• Ordered by the consumer

• Info about responses to medications, disease risk, or ancestry

• Okay for decisions about lifestyle choices

• Not suited for decisions about medical care alone

FDA Table of Pharmacogenomic Biomarkers

List of drugs with pharmacogenomic info in the drug label

• PGx info can be found in Adverse Effects, Clinical Pharmacology, Dosage (may be general or specific) & other sections

• May not be up to date

Pharmacogene Variation Consortium

 Database for pharmacogene nomenclature (mostly CYPs)

• Lists known allelic variations

• Provides standard nomenclature

• Links to other nomenclature databases (Resources)

ClinGen

Central database of genes and their variant

ClinPGx

comprehensive web resource

Formerly (PharmGKB) and (CPIC) websites

Database with searchable info about how genetic variation affects drug response

• Includes therapeutic dosing guidelines for many drugs

• National Institutes of Health (NIH) and Stanford University partnership

Pharmacokinetics

effect of body on the drug

• Drug metabolizing enzymes

• Active transporters

Pharmacodynamics

effect of drug on the body

• Receptors

• Enzymes

Factors that Affect Clinical Importance of Genetic Polymorphisms

Type of genetic change

• E.g., synonymous vs. non-synonymous SNPs, Indels, CNVs

Therapeutic index of a drug

• E.g., wide (e.g., statins) vs. narrow (e.g., warfarin)

Effect of metabolism on a drug

Presence of multiple metabolism or elimination pathways

• Participation of different metabolic enzymes

In which scenario is a genetic polymorphism more likely to result in a clinically relevant effect?

A narrow therapeutic index drug that is metabolized by a single drug metabolizing enzyme

Star Gene Nomenclature System

Gene names are italicized (convention)

Variants identified by “star” nomenclature

• *1 = ref. sequence (usually 1st discovered and/or wild-type)

• *2, *3, *5, *9, etc. = variants (could be SNPs, indels, etc.)

• *1xN = additional copies (CNVs)

many copies of a gene

Why are narrow therapeutic index drugs important in pharmacogenomics?

Small changes in drug concentration can cause toxicity or treatment failure.

What can genetic polymorphisms do to active transporters?

Change transporter expression, drug transport, specificity, or stability.

Human Genome Variant Society (HGVS) Gene Nomenclature System

Variants identified by type, position and change in gene sequence (c = coding, # = position in gene)

HGVS substitution

c.4375C>T, C in position 4375 of gene replaced by T

HGVS Deletion

c.4375_4379del, bases between position 4375 to 4379 are missing

HGVS insertion

c.4375_4376insACGT, new sequence ACGT inserted between positions 4375 and 4376

HGVS genotype homo vs hetero

c.[2376G>C]; [2376G>C] is a homozygote, c.[2376G>C]; [2376=] is a heterozygote

Active Drugs metabolism

Metabolism usually INACTIVATES them.

If metabolism decreases:

• Drug levels ↑

• Toxicity risk ↑

Examples

• Warfarin

• Nortriptyline

• Tacrolimus

War No Tac

Prodrugs metabolism

Metabolism ACTIVATES them.

If metabolism decreases:

• Active metabolite ↓

• Efficacy ↓

Examples

• Codeine

• Irinotecan

• Thiopurines

Cod I Thio

warfarin genes

Genes

• CYP2C9

• VKORC1

• CYP4F2

CYP2C9 (warfarin)

Metabolizes S-warfarin (MOST potent form).

Variants

• *2

• *3

Effect

↓ CYP2C9 activity

Clinical result

• Less warfarin metabolism

• More warfarin exposure

• Bleeding risk ↑

• Dose usually ↓

VKORC1 (warfarin)

Warfarin target enzyme.

Variant

• c.-1693G>A

Effect

↓ VKORC1 activity

Clinical result

• Increased warfarin sensitivity

• Lower dose needed

CYP4F2 (warfarin)

Breaks down vitamin K.

Variant

• *3

Effect

↓ CYP4F2 activity

Clinical result

• More vitamin K available

• More clotting tendency

• Higher warfarin dose may be needed

CODEINE

Gene

• CYP2D6

Important concept

Codeine is a PRODRUG.

CYP2D6 converts:

Codeine → Morphine

codeine poor metabolizers CYP2D6

Examples:

• 4/4

Effect

• Little morphine formed

• Poor pain control

Recommendation

Avoid codeine

Ultrarapid Metabolizers (UM)

(codeine)

Examples:

• 1/1xN CVP

Effect

• Too much morphine formed

• Toxicity risk ↑

Recommendation

Avoid codeine

Intermediate Metabolizers (IM) (codeine)

• Reduced morphine formation

• Monitor patient

normal + 1 low or no function (e.g. *1/*4)

Extensive (EM) Codeine

~77-92% patients

2 normal function alleles (e.g. *1/*1)

Full CYP2D6

Full Morphine

Dose As Recommended

NORTRIPTYLINE gene

Gene

• CYP2D6 metabolized

Important concept

Nortriptyline is ACTIVE.

Reduced CYP2D6 activity

• Drug levels ↑

• Toxicity risk ↑

Increased CYP2D6 activity

• Drug levels ↓

• Reduced efficacy

TACROLIMUS gene

Gene

• CYP3A5

Important concept

Tacrolimus is inactivated by CYP3A5.

CYP3A5*1 (tacro)

Functional enzyme present.

Effect

• More tacrolimus metabolism

• Lower drug levels

Clinical result

Higher dose needed

CYP3A5*3/*3 (tacro)

No functional enzyme.

Effect

• Less metabolism

• Higher tacrolimus levels

Clinical result

Standard/lower dosing

ethanol genes

Genes

• ADH1B

• ALDH2

ADH1B (ethanol)

Converts:
Ethanol → Acetaldehyde

ADH1B*2

Higher enzyme activity

Result

• Faster acetaldehyde buildup

• More unpleasant effects

• Lower alcoholism risk

ALDH2 (ethanol)

Inactive enzyme

Result

• Acetaldehyde accumulates

• Facial flushing

• Nausea/vomiting

• Tachycardia

IRINOTECAN gene

Gene

• UGT1A1

Important concept

Irinotecan is a PRODRUG.

Active metabolite:

• SN-38

UGT1A1 inactivates SN-38.

UGT1A1*28 (Irinotecan)

Effect

↓ UGT1A1 activity

Result

• Less SN-38 breakdown

• Toxicity ↑

Toxicities

• Diarrhea

• Neutropenia

Clinical result

Dose reduction may be needed

THIOPURINES (6-MP) gene

Gene

• TPMT

Important concept

TPMT INACTIVATES thiopurines.

TPMT variants (THIOPURINES)

• *2

• *3A

• *3C

Effect

↓ TPMT activity

Result

• More active metabolites

• Severe bone marrow suppression risk

Clinical result

Major dose reduction needed

STATINS gene/transporter

Gene/Transporter

• SLCO1B1

Important concept

SLCO1B1 transports statins into liver.

SLCO1B1 variants ( Statins)

Variant

• c.521T>C

Effect

↓ Transport into hepatocytes

Result

• Plasma statin levels ↑

• Myopathy risk ↑

Strongest evidence

• Simvastatin

GPCR POLYMORPHISMS

Variants may alter receptor response.

ADRB1 variants (GPCR)

Affect beta-blockers:

• Metoprolol

• Atenolol

• Bisoprolol

Result

Altered antihypertensive response

ADRB2 variants (GPCR)

Affect long-acting beta agonists.

Result

↑ Asthma exacerbations

OPM1 variants (GPCR)

Affect opioids like buprenorphine.

Result

↓ Drug efficacy

Metabolism of Small vs large drugs

Small

Phase I & II metabolism

Oxidation (e.g., CYPs)

Glucuronidation, etc.

Large

Proteolysis

Proteolysis

 Definition- degradation of a peptide drug or mAb into smaller fragments

 Enzymes (peptidases/proteases) widespread in body

 May be selective or non-selective

 Involves stepwise hydrolysis of peptide bonds → add water

→ bond breaks

Metabolism of IgG/mAbs

 IgG/mAb is taken up by endocytosis

• IgG/mAb transferred to endosome

• May bind to FcRn

• “Sorting” occurs

• Involves FcRn binding

• Unbound IgG/mAb sent to

lysosome→ proteolysis 

Lysosomes

 Intracellular organelles

 Multiple cellular functions

 Contain 50+ “digestive” enzymes

 Degrade peptide/proteins (via proteolysis), lipids polysaccharides, etc.

 Active at acidic pHs

What is Ethics?

Ethics is a generic term for various ways of understanding and examining the moral life. (Beauchamp and Childress, 1979)

○ Morality refers to norms about right and wrong human conduct that are so widely shared that they form a stable (although usually incomplete) social consensus.

○ Morality, as a social institution, encompasses many standards of conduct, including moral principles, rules, rights, and virtues.

Ethical Principles

● Autonomy

● Beneficence

● Justice

● Non-maleficence

Autonomy

• Self-rule free from

■ controlling interference by others and

■ limitations, such as inadequate understanding.

• The autonomous individual acts freely in accordance with a self-chosen plan

Two conditions required for autonomy to exist:

■ Liberty (independence from controlling influences) and

■ Agency (capacity for intentional action)

Beneficence

principle that refers to a moral obligation to act for the benefit of others

Justice

Justice is fair, equitable, and appropriate treatment

considering what is due or owed to persons

Distributive justice

refers to distribution of all rights and responsibilities in society

Criminal justice

which refers to the just infliction of punishment