Who Are We?

Molecular Testing Labs is a cutting-edge molecular and genetics testing laboratory focused on pharmacogenomics, also called Personalized Medicine.Our primary goal is to provide the physician, healthcare practitioner and patient with the information necessary to understand how a patient’s unique gene families interact and impact specific drug responses. • Molecular Testing Labs is a CLIA approved (#50D2050397) Molecular

/ Genetic Testing Laboratory.

Our Test Groups

Molecular Testing Labs currently offers molecular pharmacogenomics testing in 6 specific areas:

1. Pain Management Drug Response – tests to help determine the patient’s response to common pain medications and opioids.

2. Cardiac Risk Factors – tests to determine the genetic risk factors for thrombosis or prothrombin deficiency. Includes evaluating patient’s response to commonly used cardiovascular drugs.

3. Psychotropic/Neurotropic Drug Response – tests to help understand the patient’s response to neuroactive drugs. These drugs include antidepressants, antipsychotics and anxiolytics.

4. Hormone Interaction Response – tests to understand the patient’s response to common hormone replacement therapies.

5. MTHFR Testing – testing for a gene related to folic acid metabolism, which is related to a variety of birth defects and neural tube defects.

6. Infectious Disease Testing – molecular-based tests for common infectious organisms, particularly STDs such as Chlamydia, Gonorrhea, Human Papillomavirus, as well as Hepatitis viruses.

Cytochrome P450

The cytochrome P450 superfamily (officially abbreviated as CYP) is a large and diverse group of enzymes that catalyze the oxidation of organic substances. The substances of CYP enzymes include metabolic intermediates such as lipids and steroidal hormones, as well as xenobiotic substances such as drugs and other toxic chemicals. CYPs are the major enzymes involved in drug metabolism and bioactivation, accounting for about 75% of the total number of different metabolic reactions.The most common reaction catalyzed by cytochromes P450 is a monooxygenase reaction, e.g., insertion of one atom of oxygen into an organic substrate (RH) while the other oxygen atom is reduced to water:

RH + O2 + NADPH + H+ → ROH + H2O + NADP+

Cytochromes P450 (CYPs) belong to the superfamily of proteins containing a heme cofactor and, therefore, are hemoproteins. CYPs use a variety of small and large molecules as substrates in enzymatic reactions. Often, they form part of multi-component electron transfer chains, called P450 containing systems. The letter in P450 represents the word pigment as these enzymes are red because of their heme group. The number 450 reflects wavelength of the absorption maximum of the enzyme when it is in the reduced state and complexed with CO.

The cytochrome P450 (CYP450) enzymes are involved in the metabolism and processing many drugs and chemicals in our body. CYP450 is a large enzyme family mostly found in the liver. Since these enzymes have more than 50 variations, their activity may vary from person to person. Individual differences of cytochrome P450 activity can mean that certain drugs aren’t metabolized at all, are metabolized slowly, or are metabolized very quickly. This can lead to adverse drug reactions or a lack of therapeutic effect under standard therapy conditions.

Cytochrome P450

Pharmacogenomics

Pharmacogenomics (PGx) Phar-ma-co-ge-no-mics

Pharmacogenomics - the influence of genetic variation on drug response in patients. The goal of Pharmacogenomics is to develop optimized drug therapy with respect to the patients' unique drug response.

Pharmacogenomics is the study of how an individual's genetic inheritance affects the body's response to drugs. It may be possible to predict therapeutic failures or severe adverse drug reactions in individual patients by testing for important DNA sequence variations or polymorphisms (genotyping) in key drug-metabolizing enzymes, receptors, transporters, etc. Potentially, test results could be used to optimize drug choice and/or dose earlier for more effective therapy, avoid serious adverse effects, and decrease medical costs

The cytochrome P450 superfamily (CYP450) is a large and diverse group of enzymes that are responsible for drug metabolism and drug biotransformation.

Molecular Testing Labs’ CYP450 tests provide information on how quickly or slowly the patient will metabolize a certain drug (ex: warfarin or clopidogrel). This allows one to assess the likely risk of adverse drug reactions, drug toxicity, decreased effectiveness or drug failure when using a specific drug in one particular individual.

For each drug test requested, Molecular Testing Labs provides an analysis of the patient’s genetic response to that specific drug, classifying them as:

Normal Metabolizers (NM)Poor Metabolizers (PM)Intermediate Metabolizers (IM)Rapid Metabolizers (RM)Ultra Rapid Metabolizers (URM)

Key Point: Each person responds to drug treatments differently.

Molecular Testing Labs is a strong advocate for this type of genetic testing. We are not alone! The U.S. Department of Health and Human Services (HHS) and the Food and Drug Administration (FDA) have also released directives and drug label warnings in support of genetic pharmacological testing.

http://www.fda.gov/drugs/scienceresearch/researchareas/pharmacogenetics/ucm083378.htm

U.S. Government Support of Personalized Medicine

This Statement was Provided by the U.S. Department of Health and Human Services:

“The Personalized Health Care Initiative will improve the safety, quality and effectiveness of healthcare for every patient in the U.S. By using ‘genomics,’ or the identification of genes and how they relate to drug treatment, personalized healthcare will enable medicine to be tailored to each person’s needs.

“Healthcare that is proactive, instead of reactive, gives the patient the opportunity to become more involved in their own wellness. The U.S. Department of Health and Human Services seeks to advance this Initiative through two guiding principles:

• Provide federal leadership supporting research addressing individual aspects of disease and disease prevention with the ultimate goal of shaping preventive and diagnostic care to match each person’s unique genetic characteristics.

• Create a ‘network of networks’ to aggregate anonymous healthcare data to help researchers establish patterns and identify genetic ‘definitions’ to existing diseases.”

Personalized Medicine

Personalized medicine is a medical model that proposes the customization of healthcare, with decisions and practices being tailored to the individual patient by use of genetic or other information.

CYP450 2C19, 2C9, 2D6, 3A4, 3A5:

Key Factors in Effective Pain Management

Pain Management

Incidence of Pain

More than 116 million Americans suffer from acute or chronic pain each yearMore than 2/3 of all emergency room patients are present for pain management

Condition No. Sufferers Source

Chronic Pain 116 million Americans(>1.5 billion Worldwide)

Institute of Medicine of The National Academies

Diabetes 25.8 million Americans American Diabetes Association

Coronary Heart Disease 16.3 million Americans American Heart Association

Stroke 7.0 million Americans American Heart Association

Cancer 11.9 million Americans American Cancer Society

Source: American Academy of Pain Medicine

State of Pain Management

Drugs are the “first line” of treatment for most forms of pain.

Goal of successful pain management is to effectively control patient pain without causing excess side effects from the medication prescribed.

However, only 58% of those who took prescription medication received any form of pain relief.

Less than 41% of patients taking over-the-counter pain medication reported relief.

Source: Peter D. Hart Research Associates – Americans Talk About Pain: Survey Among Adults Nationwide, Aug. 2003

Dosing Management

The incidence of unintentional (and preventable) overdose- related deaths is growing exponentially.

Of all drug-related deaths in the U.S., 43% are due to pain relief medication.

80% of all pain management drugs are prescribed by the general practitioner or the internist

Sources: American Academy of Pain Medicine; CDC; IMS Institute of Healthcare Informatics

The CYP2D6 gene metabolizes 25% of all prescribed drugs, such as codeine, tricyclic antidepressants (eg, nortriptyline), SSRI fluvoxamine, SSNRI venlafaxine, classical antipsychotics (eg, haloperidol), and beta-blockers.

Analgesic Related Deaths

The rising death toll is attributed to an increased use of a class of drugs known as Opioid Pain Relievers.

Sources: CDC; IMS Institute of Healthcare Informatics

Opioids 101

Opioids are a class of narcotic analgesics used to control pain sensations. These drugs are often indicated for the relief of chronic, and moderate to severe post-surgical pain, in addition to recovering cancer patients.Commonly encountered members of this class include:

HydrocodoneCodeineOxycodoneMorphine

FACT: Hydrocodone was the most frequently prescribed drug in the U.S. in 2010, with 131.9 million prescriptions

Sources: IMS Institute of Healthcare Informatics

Opioids – Symptoms

Common side effects and symptoms of opioid toxicity include:DizzinessNauseaVomitingPhysical DependenceToleranceSedation/DrowsinessWeaknessRespiratory Depression

Prolonged duration of these symptoms will lead to addiction, overdose, and death.

Potential Causes

Doctors know approximately 50% of all patients undergoing pain management will not receive adequate relief at first dosing and/or are at a higher risk of experiencing adverse, potentially life threatening events.

Explanations:Under/over dosingDrug interactionsGenetic variations

CYP450 2C19, 2C9, 2D6, 3A4, 3A5

CYP2D6 and 3A4

CYP2D6 and CYP3A4 involvement in the primary oxidative metabolism of hydrocodone by human liver microsomes.

Hutchinson, et al. – Brit J Clin Pharm, Sep. 2003Objective: To identify the CYP450 enzymes involved in the metabolism of hydrocodone to its active metabolites hydromorphone and norhydrocodoneConclusions:CYP2D6 is almost exclusively responsible for the formation of hydromorphone.

CYP2D6 Poor Metabolizers formed significantly less hydromorphone than Extensive Metabolizers (~1% of normal).

CYP3A4 is mainly responsible for the formation of norhydrocodone.

Role of CYP2D6 in Opioids

Utilization of Pharmacogenomics and Therapeutic Drug Monitoring for Opioid Pain Management

Jannetto, et al. – Pharmacogenomics, Oct. 2009Objective: To determine whether there is a relation between patient CYP2D6 genotype and observed response to opioid based analgesic therapy.Conclusions:54% of the study population were CYP2D6 extensive metabolizers; 41% were intermediate metabolizers; and 5% were poor metabolizers.80% of patients exhibiting adverse drug events from opioid therapy had impaired CYP2D6 function (remainder 20% of ADEs were attributed to other drug-drug interactions).A relationship exists between CYP2D6 genotype and observed opioid pain management outcomes. Genotyping information for patient CPY2D6 status will enhance analgesic therapy.

Impact of Combined Enzyme Inhibition

Effect of inhibition of CYP450 enzymes 2D6 and 3A4 on the pharmacokinetics of intravenous oxycodone: a randomized, three phase, cross-over, placebo-

controlled studyGronlund, et al. – Clin Drug Investig, Mar. 2011

Objective: To determine whether the inhibition of CYP2D6 alone or along with CYP3A4 would alter the metabolism of oxycodone.Conclusions:Inhibition of just the CYP2D6 enzyme did not yield any significant change to the plasma concentration of oxycodone.However, inhibition of both CYP2D6 and CYP3A4 increased the half-life of oxycodone from 3.8 hrs to 6.6 hrs and increased exposure to the drug 2-fold.While isolated enzyme function inhibition is insignificant, the combined effect of non-functional CYP2D6 and CYP3A4 substantially increases exposure to intravenous oxycodone.The presence of this mutation, by increasing the half-life of the active drug, clues the provider he/she should decrease the dose, extend the time between doses, or both when prescribing this drug, or choose a different drug altogether.

Utility of CYP3A4 & 3A5 Genotyping

Pharmacogenomics as Molecular Autopsy for Forensic Toxicology: Genotyping Cytochrome P450 3A4* 1B and 3A5* 3 for 25 Fentanyl Cases

Jin, et al. – J Anal Toxicol, Oct. 2005Objective: To identify the role of CYP3A4 and 3A5 in cases of fatal overdosing of the opioid, fentanyl.Conclusions:6% of the study cases had the CYP3A4*1B mutation; 92% had the CYP3A5*3 mutation.Subjects with these mutations had significantly lower concentrations of the opioid fentanyl than wild type subjects (1.4 ug/L vs. 7.3 ug/L).Homozygous CYP3A5*3 mutation severely impairs opioid metabolism.Genotyping of both CYP3A4*1B and CYP3A5*3 can help certify and prevent fentanyl toxicity.The presence of this mutation, by increasing the half-life of the active drug, clues the provider he/she should decrease the dose, extend the time between doses, or both when prescribing this drug, or choose a different drug altogether.

FDA Relabels Opioids

The FDA had relabeled several prominent opioids to include pertinent pharmacogenomics information.Example: OxyContin:(From OxyContin Package Insert)

Inhibitors of CYP3A4:Since the CYP3A4 isoenzyme plays a major role in the metabolism of oxycodone, co-administration of drugs that inhibit CYP3A4 activity… may cause decreased clearance of oxycodone which could lead to an increase in oxycodone plasma concentrations.

Inducers of CYP3A4: CYP450 inducers… may induce the metabolism of oxycodone and, therefore, may cause increased clearance of the drug, which could lead to a decrease in oxycodone plasma concentrations, lack of efficacy, or, possibly, development of abstinence syndrome in a patient who had developed physical dependence to oxycodone.

Economics of Testing

The U.S. Government understands the cost of molecular pharmacological testing and fully reimburses the cost of these tests through Medicare. The superordinate goal is to save monies on the overall cost of the treatment by mitigating the cost consequences of

inappropriate dosing and use of pain management drugs.

Source: 1: INMAR: Relieving Pain in America: A Blueprint for Transforming Prevention, Care, Education, and Research, 2011; 2: Castro, et. al. “Pharmacogenomics in the Clinic”, American Society of Clinical Oncology, Journal of Oncology Practice, 2006

116 million people suffering from pain each

year in U.S.1

$250-$500 per patient for

adverse event prevention

testing2

$29-$58 billion in total testing

costs each year in U.S.

X =

Dosing Guidelines from the CPIC

Clinical Pharmacogenetics Implementation Consortium Guidelines for Codeine Therapy in Context of CYP450 2D6 Genotype

Phenotype Implications for Codeine Metabolism Recommendations for Codeine Therapy

Ultra rapid metabolizer Increased formation of morphine following codeine administration, leading to higher risk of toxicity

Avoid codeine use due to potential fortoxicity. Consider alternative analgesics such as morphine or a nonopioid. Consider avoiding tramadol.

Extensive metabolizer Normal morphine formation 15–60 mg every 4 hr as needed for pain (label recommendation)

Intermediate metabolizer Reduced morphine formation Begin with 15–60 mg every 4 hr as needed for pain. If no response, consider alternative analgesics such as morphine or a nonopioid. Monitor tramadol use for response.

Poor metabolizer Greatly reduced morphine formation following codeine administration, leading to insufficient pain relief

Avoid codeine use due to lack ofefficacy. Consider alternative analgesics such as morphine or a nonopioid. Consider avoiding tramadol.

Metabolism of Common Opioids

Class Drug MetabolismOpium alkaloids Codeine 10% CYP3A4 (to norcodeine); 5% CYP2D6 (to morphine); 80% UGT2B7

Hydrocodone CYP2D6 (to hydromorphone) and CYP3A4 (to norhydrocodone); other minor non-CYP oxidative enzymes; UGTs (CYP-metabolized products)

Oxycodone CYP2D6 (to oxymorphone) and CYP3A4 (to noroxycodone); CYP-metabolized product(s) by UGTs

Semisynthetic derivatives

Dihydrocodeine 5%-10% CYP2D6 (to dihydromorphine) and CYP3A4 (to nordihydrocodeine); 85% UGT2B7

Hydromorphone Hepatic glucuronide conjugation via UGT1A3, UGT2B7; dihydromorphinone ketone reductase

Phenylheptylamine Methadone N-demethylation by CYP3A4

Oripavine derivatives Buprenorphine CYP3A4 (65%), CYP2C8 (30%), CYP3A5, CYP3A7, CYP2C9, CYP2C19, and CYP2C18; CYP-metabolized product(s) further cleared by UGTs

Phenylpiperidines Fentanyl N-dealkylation by CYP3A4 Meperidine CYP3A4, CYP2B6, and CYP2C19 Sufentanil N-dealkylation by CYP3A4Diphenylpropylamine Loperamide N-demethylation by CYP2B6, CYP2C8, CYP2D6, and CYP3A4derivatives Propoxyphene N-demethylation by CYP3A4

Source: Kadiev E, Patel V, Rad P, et al. Role of pharmacogenetics in variable response to drugs: focus on opioids. Expert Opin Drug Metab Toxicol. 2008;4(1):77-91

Our Summary

43% of drug-related deaths in the U.S. are related to pain medication.

The FDA has issued numerous bulletins and updates to product labels informing clinicians and the public of these dangers, and how they can be assessed with genetic testing.

Dosing algorithms are/soon will be updated to account for genetic guidance.

A percentage of deaths from opioid abuse is attributable to outright addiction and abuse, and not directly related to genetically predictable adverse reactions to the drug. However, Molecular Testing Labs notes a possible correlation between the individuals addicted to opioids and their genetic susceptibility to their effects.

The goal of Molecular Testing Labs is to provide physicians and their patients with insight into their unique response to pain medications in order to decrease the number of adverse reactions and to better manage their pain medication strategy.

Cardiac Molecular Tests

CYP2C19 and its Relationship to the Cardiac ResponseCYP450 includes a gene (CYP2C19) that codes for numerous liver enzymes involved in the metabolism of toxins, metabolic intermediates, lipids, sterols and xenobiotic substances such as drugs. These enzymes account for 75% of the metabolic processes involved in the metabolism and bioactivation of all drugs. Approximately 15% of all prescribed drugs, including clopidogrel (Plavix), propafenone, omeprazole, sertraline, citalopram, and diazepam are affected by base pair substitutions in this gene. These mutations affect the speed and ability of the body to metabolize and utilize these drugs. Detecting these genetic variations helps clinicians predict how a patient will respond to these types of drugs.Clopidogrel (Plavix) is a common anticoagulant used to prevent and treat blood clots. Clopidogrel is absorbed in the intestines and converted into its active form by enzymes in the liver. Genetic studies indicate that some people have genetic variations that reduce the activity of these critical enzymes. Patients who need clopidogrel but have these variations are at increased risk of heart attacks, strokes and death from cardiovascular causes compared to those whose genetic makeup enables them to normally metabolize the drug. Five phenotypes are identified when using the 2C19 genetic test from Molecular Testing Labs: normal metabolizers (NM), poor metabolizers (PM), intermediate metabolizers (IM), rapid metabolizers (RM) and ultra rapid metabolizers (URM).Patients with reduced function alleles have ~3.5x greater risk for major adverse cardiovascular events – with the greatest risk for poor metabolizers of CYP2C19.1

People of African and Asian ancestry have a greatly increased prevalence of Poor Metabolizer status of CYP2C19.2

Source: 1 Tabassome Simon, MD Genetic Determinents to Response and Cardiovaascular Events ( English Journal of Medicine 2011 p.300-365 ) 2.Hansten PO ( The top 100 Drug Interactions ) H&H Publicatins 2011

Cardiac Molecular Tests

Factor II and its Relationship to Cardiac EventsThe prothrombin gene G20210A mutation differs from the gene for normal prothrombin or factor II by a single nucleotide. This point mutation causes the body to produce excess amounts of prothrombin.

In individuals with the prothrombin gene mutation, prothrombin levels are higher, which in turn contributes to the formation of blood clots.

Individuals who have inherited one copy of the gene mutation (heterozygotes) have a risk of DVT or PE similar to the risk of individuals who have inherited one copy of the factor V Leiden gene.

Individuals who have inherited: 1) two copies of the prothrombin gene mutation (homozygotes), or 2) individuals who have inherited one copy of the prothrombin gene mutation and

one copy of the factor V Leiden (compound heterozygotes) . . .. . . have a risk of DVT or PE similar to the risk of individuals who have inherited

two copies of the factor V Leiden gene.

Cardiac Molecular Tests

Factor II and its Relationship to Cardiac Events

The increased risk of venous thrombosis in patients who are heterozygous for the prothrombin gene polymorphosis is 3X greater than normal.

Patients with a previous or current thrombotic event that have the prothrombin gene polymorphism are at increased risk for a reoccurence.

Cardiac Molecular Tests

Factor V and its Relationship to Cardiac EventsFactor V is the most common variant associated with inherited thrombosis.Has a high prevalence in the U.S. population (4% - 6% and accounts for 85% - 95% of activate protein C resistance cases (APCR).Enhanced risk of venous thrombosis in the presence of the Factor V variant, with odds of 3% – 8% in heterozygotes and 30% – 40% in homozygotes.The risk of thrombosis is greatly increased for patients with multiple genetic risk factors.Molecular Testing Labs recommends that the Factor II and Factor V genetic tests are completed in conjunction with the 2C19 genetic test.

MTHFR

MTHFR is an abbreviation for the gene that codes for an enzyme called methyltetrahydrofolate reductase. The MTHFR enzyme helps the body to manufacture proteins. Two mutations are the most studied: 677 and 1298, although there are more than 50 known MTHFR variants.Mutations in the 677T variant is associated with thromboembolic and cardiovascular disease, which can lead to blood clots, strokes, embolisms, and heart attacks. Mutations in the 1298C variant are associated with a number of chronic diseases, including fibromyalgia, irritable bowel syndrome, migraines, and other conditions. Approximately 30%-40% of the population carries a mutation at position C677T, and 20% of the population carries a mutation at position C1298T. Individuals with these mutations are at increased risk for thromboembolic and cardiovascular disease. They are also linked with recurrent miscarriages and neutral tube defects (NTD) in the children of the women carrying the mutations.Neural tube defects are common birth defects that result in incomplete development of the spinal cord or brain. The most common NTDs are anencephaly and spina bifida. Anencephaly results in major portions of the brain and skull not developing. Spina bifida results in incomplete development of the spine, which causes the spinal cord and the meninges to poke out of the child’s back. At least 40 of the MTHFR gene mutations result in elevated levels of homocysteine in the blood (homocystinuria). Homocystinuria is linked to a wide range of disorders, including nearsightedness, osteoporosis, failure to thrive, seizures, megaloblastic anemia and movement disorders.Individuals with both mutations, 677T/1298C are more prone to fibromyalgia, IBS, migraine, and autism. Studies have found that 98% of autistic children have an MTHFR abnormality.

MTHFR

Molecular Testing Labs considers the MTHFR genetic test one of the most important to your health. MTHFR is involved in processing folic acid. Small changes in this chemical process can be associated with any one of the following conditions: • Depression• Anxiety• Chronic pain• Chronic fatigue• Nerve pain• Migraines• Elevated homocysteine levels• Fibromyalgia• Irritable Bowel Syndrome• Alzheimer’s• Bipolar disorder• Schizophrenia• Parkinson’s• Stroke• Heart Disease• Multiple miscarriages• Stillbirths, as well as other congenital anomalies (birth defects)• Down syndrome• Autism

Infectious Diseases - HPV

Molecular Testing Labs provides a test for Human Papillomavirus (HPV) that detects the presence of thirteen (13) high-risk HPV types (Types 16, 18, 31, 33, 35, 39, 45, 51, 52, 56, 58, 59, and 68) and two low risk types (Types 6 and 11). The test also identifies high-risk HPV types by specific types (Types 16, 18, 31, 33 and 45).HPV is the most common sexually transmitted virus in the U.S. and is associated with several types of cancer: cervical, vulvar, vaginal, penile, anal and oropharyngeal.

Almost every sexually active person will acquire HPV at some point in their lives.Each year over 21,000 HPV associated cancers are diagnosed in women.Each year over 12,000 HPV associated cancers are diagnosed in men.

High-risk HPV infection accounts for approximately 5% of all cancers worldwide.Persistent HPV infection can result in precancerous cervical lesions as well as invasive cervical and/or head and neck cancers. With regular HPV Molecular screening and appropriate follow-up, most cancer precursors can be identified and treated to interrupt progression to invasive disease.Some clinicians recommend that women who are non-symptomatic have an HPV Molecular Screening test as part of their vaginal birth discussion.Molecular Testing Labs provides HPV screening through a buccal swab technique in the cheek or by a cervical sample in conjunction with cytology.

MOLECULAR TESTING LABS

Helping You Help Yourself

Our facility is located in Vancouver, Washington.

For more information, visit our website:

WWW.MOLECULARTESTINGLABS.COM