Urine Drug Screening: The Essentials of Interpretation

Urine Drug Screening: The Essentials of Interpretation Loralie J Langman, PhD DABCC (CC, MD, TC), F-ABFT Director Clinical and Forensic Toxicology Laboratory, Mayo Clinic Professor, Mayo Clinic College of Medicine Rochester, MN September 30, 2016

DISCLOSURE Relevant Financial Relationship(s) None Off Label Usage None

Case #1 50 y old male Prescribed: Percocet 5/325 You want to see if he is taking it.

Drug Testing What specimen to collect? Is the individual impaired/under the influence now? BLOOD / SERUM Was an individual using or exposed to a drug? URINE

Focus on urine drug testing (UDT)

Case #1 Urine Immunoassay Screen Drug Result Cut-off (ng/ml) amphetamines Presumptive Positive 500 barbiturates negative 200 benzodiazepines negative 200 cannabinoids negative 20 cocaine metabolite negative 150 opiates negative 300

Drug Screening How do we do what we do?!

Traditional Drug Screening Classically Two Stages: Stage 1 Screen Stage 2 Confirmation/Definitive Testing

How do Immunoassay Screens work? Usually immunoassay-based method Detect compounds that look like the drug you are testing for Purpose is to identify negative samples.

Opiate Screening Immunoassay Designed to look for illicit opiate use HEROIN morphine More practically thought of as a morphine immunoassay Therefore, detection of other opiates (ex. codeine, hydrocodone, hydromorphone, oxycodone, oxymorphone ) depends on antibody specificity.

N CH 3 N CH 3 HO HO O OH H 3 C O O O morphine oxycodone

Opiate Screening Immunoassay Assay 1 Oxycodone ~ 20% cross reactivity Need ~ 5X more oxycodone to give a positive Opiate Screen result than morphine ~ 1500 ng/ml oxycodone to give a positive screening test when a 300 ng/ml cut-off is used Assay 2 Oxycodone ~ 3% cross reactivity Need ~ 33X more oxycodone to give a positive Opiate Screen result than morphine ~ 9900 ng/ml oxycodone to give a positive screening test when a 300 ng/ml cut-off is used

Definitive Testing Identify what is causing the positive result in the immunoassay screening assay OR What to order when you are looking for a specific drug May or may not be quantitative.

Laboratory Medicine Practice Guideline: Using Clinical Laboratory Tests to Monitor Drug Therapy in Pain Management Patients Recommendation: First-line definitive testing (qualitative or quantitative) is recommended for detecting the use of relevant over-thecounter medications, prescribed and non-prescribed drugs, and illicit substances in pain management patients. Definitive urine drug testing specifically identifies or quantifies a drug and/or its metabolites. Strength of recommendation: A Quality of evidence: II https://www.aacc.org/community/national-academy-of-clinical-biochemistry

Targeted Opioid Screen

Definitive Testing Done in the Lab Gas Chromatography Mass Spectrometry (GC/MS) Liquid Chromatography Mass Spectrometry (LC/MS, LC/MS-MS, TOF, QTOF) Pro Identify the drug(s) Con Looks for specific drugs

Case #1 Opiate Confirmation (LC-MS/MS) Drug LOD oxycodone 6600 ng/ml 100 ng/ml oxymorphone 8152 ng/ml 100 ng/ml morphine negative 100 ng/ml codeine negative 100 ng/ml hydrocodone 250 ng/ml 100 ng/ml hydromorphone negative 100 ng/ml

Drug Metabolism Phase 1 Reactions Oxidation CYP450 Reduction Phase 2 Reactions Conjugation Reactions: Glucuronidation Sulfate Glutathione Glucine Acetylation Methylation Generally these reactions generate compounds that are more water soluble - aid in excretion

Metabolism Parent Drug Major/Active Metabolite (s) morphine codeine morphine heroin 6-mono-acetylmorphine morphine hydromorphone hydrocodone hydromorphone oxymorphone oxycodone oxymorphone Baselt, RC. Disposition of Toxic Drugs and Chemicals in Man, 9th Edition Ed, 2011, Chemical Toxicology Institute, Foster City, CA. Boerner, U. The metabolism of morphine and heroin in man. Drug Metab Rev, 1975; 4(1), p 39-73. Goodman & Gilman's The Pharmacological Basis of Therapeutics. 12th ed. New York: McGraw-Hill..

Metabolism and more Parent Drug Minor metabolite morphine hydromorphone <2.5 codeine hydrocodone <11 High doses for long period of time % of parent drug References Cone, EJ et al. J Anal Toxicol, 2006; 30(1), p 1-5 Oyler, JM et al. J Anal Toxicol, 2000; 24(7), p 530-5. Parent Drug Contaminant % of parent drug heroin codeine NOT metabolite Very low levels compared to morphine References Baselt, RC. Disposition of Toxic Drugs and Chemicals in Man, 8th Edition Ed, 2008, Chemical Toxicology Institute, Foster City, CA.

Acceptable Impurities in Commercial Drug Preparations Active Pharmaceutical Ingredient (API) Process impurity* (%) Allowable limit (%) Typical observed (%) hydrocodone codeine 0.15 0.00 0.10 hydromorphone morphine hydrocodone 0.15 0.10 0.00 0.025 0.00 0.025 morphine codeine 0.50 0.01 0.05 oxycodone hydrocodone 1.00 0.02 0.12 oxymorphone hydrocodone oxycodone 0.15 0.50 NOTE: All contaminants when detected in greater amounts than the allowable limits stated above would be considered evidence of use. *These are NOT metabolites. 0.03 0.10 0.05 0.40 Pesce, A., et al., Interpretation of urine drug testing in pain patients. Pain medicine, 2012. 13(7): p. 868-85

Case #1 cont d Patient is also prescribed Ritalin (methyl phenidate) Urine Immunoassay Screen Drug Result Cut-off (ng/ml) amphetamines Presumptive Positive 500 barbiturates negative 200 benzodiazepines negative 200 cannabinoids negative 20 cocaine metabolite negative 150 opiates negative 300 oxycodone Presumptive Positive 100

Case #1 cont O OH NH 2 CH 3 amphetamine HN methylphenidate NH CH 3 CH 3 methamphetamine Baselt, RC. Disposition of Toxic Drugs and Chemicals in Man, 9th Edition Ed, 2011, Chemical Toxicology Institute, Foster City, CA. Goodman & Gilman's: The Pharmacological Basis of Therapeutics, 12th Ed, 2011, McGraw-Hill.

Drug Screens are good a finding things you didn t know you were looking for.. Not good at finding things you are. Whaaaaaaaaaaaaaat?!?!?!?!?!?!?! Screening methods try to pick up as much as possible but to do that you sacrifice of the detection limit and the ability to identify exactly which drug it is.

Drug Screens One could say that: drug screening is like looking for a needle in a haystack when you don t know what the needle looks like or if there is a needle at all!!!! Getty images

If you know what you are looking for order a test that looks for it specifically (Definitive Testing)

Case #2 50 y old female Prescribed: methadone 60 mg/d

Case #2 Urine Immunoassay Screen Drug Result Cut-off (ng/ml) amphetamines negative 500 barbiturates negative 200 benzodiazepines negative 200 cannabinoids negative 20 cocaine metabolite negative 150 opiates Presumptive Positive 300 oxycodone negative 100

Opiate vs Opioid Opiate: The term is often incorrectly used to refer to all drugs with morphine-like pharmacological action. More properly classified under the broader term opioid. http://dictionary.reference.com/browse/opiate http://en.wikipedia.org/wiki/opiate#cite_note-0

Opiate vs Opioid An opioid is a chemical substance that has a morphine-like action in the body. Natural opiates - alkaloids contained in the resin of the opium poppy morphine, codeine Semi-synthetic opiates - created from the natural opiates hydromorphone, hydrocodone, oxycodone, oxymorphone, diacetylmorphine (Heroin) Fully synthetic opioids fentanyl, meperidine (pethidine), methadone, tramadol Endogenous opioid peptides - produced naturally in the body endorphins, enkephalins, dynorphins, and endomorphins. Goodman & Gilman's The Pharmacological Basis of Therapeutics. 12th ed. New York: McGraw-Hill..

Opiates N CH 3 N CH 3 N CH 3 HO HO O morphine OH HO O hydromorphone O HO O oxymorphone O N CH 3 N CH 3 N CH 3 HO C H 3 O O codeine OH C H 3 O O hydrocodone O H 3 C O O oxycodone O

Opioids in the Opiate assay Opioids No chemical similarity to opiates CH 3 H 3 C CH 3 N N CH 3 HO tapentadol CH 3 N CH 3 CH 3 H 3 C O HO tramadol HO O morphine OH CH 3 O CH 3 H 3 C N O O CH 3 N N O N CH 3 H 3 C methadone meperidine fentanyl CH 3

Case #2 cont Urine Methadone Confirmation (GC-MS) Methadone 175 ng/ml Screening assay cut off 300 ng/ml EDDP (methadone metabolite) 12062 ng/ml Confirmatory methods typically have limits of quantitation lower than the screening method. & Why to order it when you know you are looking for it

Methadone Patients who are taking methadone for therapeutic purposes usually excrete both parent methadone and EDDP in their urine. BUT Methadone levels in urine are widely variable depending on factors such as dose, timing of collection, metabolism, and urine ph (can even be undetectable). EDDP levels, in contrast, are relatively unaffected by the influence of ph and are therefore preferable for assessing compliance with therapy. measure levels of both methadone and EDDP Principles of Forensic Toxicology. 2nd ed. Washington DC: AACC Press, 2003

Another complicated class of drugs.

Benzodiazepines General Structure H 3 C N O Cl N diazepam

Drug Metabolism Benzodiazepine Drug Phase I Phase II Lorazepam glucuronidation clonazepam 7-amino-clonzaepam glucuronidation Flunitrazepam 7-amino-flunitrazepam glucuronidation Triazolam Hydroxy-traizolam glucuronidation Alprazolam Hydroxy-alprazolam glucuronidation Midazolam Hydroxy-midazolam glucuronidation Flurazepam Hydrox-fluazepam glucuronidation Baselt, RC. Disposition of Toxic Drugs and Chemicals in Man, 7th Edition Ed, 2005, Chemical Toxicology Institute, Foster City, CA. Boerner, U. The metabolism of morphine and heroin in man. Drug Metab Rev, 1975; 4(1), p 39-73. Goodman & Gilman's: The Pharmacological Basis of Therapeutics, 10th Ed, 2001, McGraw-Hill.

Drug Metabolism Benzodiazepine Chlordiazepoxide Diazepam oxidation CYP450 oxidation CYP450 Demoxepam oxidation CYP450 Nordiazepam Temazepam Oxazepam glucuronidation glucuronidation EXCRETION Baselt, RC. Disposition of Toxic Drugs and Chemicals in Man, 7th Edition Ed, 2005, Chemical Toxicology Institute, Foster City, CA. Boerner, U. The metabolism of morphine and heroin in man. Drug Metab Rev, 1975; 4(1), p 39-73. Goodman & Gilman's: The Pharmacological Basis of Therapeutics, 10th Ed, 2001, McGraw-Hill.

Benzodiazepine Immunoassay The cross-reactivity of the different manufactures immunoassays to the over 30 prescribed benzodiazepines are even more variable than the opiates one Clonazepam cross-reactivity is almost universally poor

Tip If you are looking for a specific drug Tell us The more information the lab has the better!

Definitive methods.. Identify the drug(s) indicated with certainty.. So can those results be false positives too?

Definitive methods.. Identify the drug(s) indicated with certainty.. So can those results be false positives too? But you just said they were definitive!!!!!!!!!!

False Positives Clinical False Positive Methodological False Positive

Clinical False Positive

Amphetamines Amphetamine and methamphetamine have limited legitimate pharmacological uses including narcolepsy, obesity, and attention-deficit hyperactivity disorders. There are also prescription drugs that are metabolised to amphetamine or methamphetamine. Principles of Forensic Toxicology. 2nd Ed, 2003, p 245-264. AACC Press, Washington DC. Logan, BK. Forensic Sci Rev, 2002; 14(p133-151. Hornbeck, CL et al. J Anal Toxicol, 1993; 17(1), p 23-5.

Clinical False Positive Drug uses Metabolites Adderall amphetamine Dexedrin d-amphetamine ADHD Narcolepsy ADHD Desoxyn methamphetamine ADHD amphetamine Deprenyl selegiline Geowdin famprofazone Parkinson s Disease Analgesic, antipyretic methamphetamine amphetamine methamphetamine amphetamine

How to approach the case A thorough review of medical history to determine what medications the patient is prescribed. Can t rule out they DIDN T use amphetamine or methamphetamine But does explain a positive test

Point to Remember An awareness of the patients complete/recent medication history is critical to prevent the individual from being wrongly accused of using illicit drugs.

Methodological False Positive A drug is confirmed/identified when the drug is absent.

Amphetamine Type Stimulants cont. Amphetamine-type stimulants are often chiral compounds. NH 2 NH 2 H 3 C H H CH 3 Generally: S(+) enantiomers stimulate the central nervous system R(-) enantiomers act peripherally Illicit amphetamine-type stimulants are mixtures of S(-) and R(-) enantiomers. Principles of Forensic Toxicology. 2nd Ed, 2003, p 245-264. AACC Press, Washington DC.

Methamphetamine NH CH3 CH 3 CNS effects of S(+) methamphetamine [d-methamphetamine] are ~ 10X greater. Vasoconstrictive effects of R( ) methamphetamine [lmethamphetamine] are greater. Principles of Forensic Toxicology. 2nd Ed, 2003, p 245-264. AACC Press, Washington DC. Fitzgerald, RL et al. J Anal Toxicol, 1988; 12(5), p 255-9. Hornbeck, CL et al. J Anal Toxicol, 1993; 17(1), p 23-5. Poklis, A et al. Ther Drug Monit, 1995; 17(1), p 89-94.

Methamphetamine Because of the minimal CNS activity and, therefore low abuse potential, R( ) methamphetamine [l-methamphetamine] was included in some nonprescription nasal decongestants because of its vasoconstrictive properties. They have largely been replaced with other decongestant medications. Principles of Forensic Toxicology. 2nd Ed, 2003, p 245-264. AACC Press, Washington DC. Fitzgerald, RL et al. J Anal Toxicol, 1988; 12(5), p 255-9. Hornbeck, CL et al. J Anal Toxicol, 1993; 17(1), p 23-5. Poklis, A et al. Ther Drug Monit, 1995; 17(1), p 89-94.

Some prescription drugs either have chiral amphetamine or methamphetamine as the active ingredient, or are metabolised to chiral methamphetamine or amphetamine. Drug Drugs detected Dexedrin d-amphetamine S(+) amphetamine Deprenyl selegiline R(-) methamphetamine R(-) amphetamine Vyvanse lisdexamfetamine ( L-lysine-dextroamphetamine) S(+) amphetamine Logan, BK. Forensic Sci Rev, 2002; 14, p 133-151. Romberg, RW et al. J Forensic Sci, 1995; 40(6), p 1100-2.

How to approach the case Routine confirmatory testing does not distinguish between the enantiomers Chiral discrimination of methamphetamine isomers may be necessary to distinguish: use of nonprescription nasal inhalants chiral prescription medications from illicit use of methamphetamine Principles of Forensic Toxicology. 2nd Ed, 2003, p 245-264. AACC Press, Washington DC. Cody, JT. J Chromatogr, 1992; 580(1-2), p 77-95. Fitzgerald, RL et al. J Anal Toxicol, 1988; 12(5), p 255-9. Hughes, RO et al. J Anal Toxicol, 1991; 15(5), p 256-9. Rasmussen, LB et al. J Chromatogr B Analyt Technol Biomed Life Sci, 2006; 842(2), p 136-41.

Interpretation Urine Concentrations Levels in urine vary widely depending on factors such as: dose hydration status timing of collection metabolism. CANNOT relate a urine concentration back to a dose? But within a given patient the pattern is may be stable.

Another way to look at urine results drug ng/ml 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 sample

Another way to look at urine results drug ng/ml creatinine g/dl 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 sample

Another way to look at urine results drug/creatinine 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 sam ple What this tells you is that something changed: Disease Concomitant medications Altered compliance...

The Key to understanding and interpreting drugs screens is:

Points to Take Home Know what the lab can detect and let them know what you need. Drug screen interpretation requires two-way communication.

Points to Take Home A positive result: They were exposed to the drug(s) identified. Cant tell the dose or when it was taken. A negative result: Doesn t exclude the possibility they were exposed to drug(s).

Points to Take Home The lab can tell you what active drug(s) was taken, NOT the formulation. The lab CANNOT test for diversion We can help identify if a patient has not taken the drug but we can t tell you what else the patient did with it

Points to Take Home Is the right test ordered? Be specific in what you order you will only get what you ask for Reminder: Opiate vs opioids

Points to Take Home Relating a urine drug concentration to dose is difficult because: kidney s concentrating ability hydration of patient timing of collection (in relation to dose) Intuitively higher levels ~ recent use and/or large dose

Points to Take Home Are any additional tests required? Do you require creatinine or specific gravity? Do you require additional tests to screen for adulterants? ph, oxidants Do you require Chain of Custody?

langman.loralie@mayo.edu

Interpretation Detection Times Focus on urine drug testing (UDT)

Interpretation Detection Times Stimulants Substance Amphetamine Methamphetamine Amphetamine variants (MDA, MDMA) Cocaine (benzyolecgonine) Period of Detection 1-3 days 1-3 days 1-3 days 1-4 days These are approximate detection times for the drug or metabolites in urine. The actual detection time depends on individual metabolism and the dose of the drug. www.nida.nih.gov

Interpretation Detection Times Opioids and Morphine Derivatives Substance Morphine Codeine Oxycodone Methadone and metabolite (EDDP) Period of Detection 1-3 days 1-3 days 1-3 days 1 day - 1 week These are approximate detection times for the drug or metabolites in urine. The actual detection time depends on individual metabolism and the dose of the drug. www.nida.nih.gov

Interpretation Detection Times Depressants Substance Alcohol Barbiturates Short acting Intermediate acting Long acting Period of Detection 6-10 hours 1 day 2-3 days 7-10 days These are approximate detection times for the drug or metabolites in urine. The actual detection time depends on individual metabolism and the dose of the drug. www.nida.nih.gov

Interpretation Detection Times Depressants Substance Benzodiazepines Short acting Intermediate acting Long acting Period of Detection 1 day 2-4 days 1 week These are approximate detection times for the drug or metabolites in urine. The actual detection time depends on individual metabolism and the dose of the drug. www.nida.nih.gov

Interpretation Detection Times Hallucinogens Substance LSD Mescaline Phencyclidine Psilocybin Amphetamine variants DOB, DOM, MDA, MDMA Period of Detection 8 hours 2-3 days 2-8 days 8 hours 1-3 days These are approximate detection times for the drug or metabolites in urine. The actual detection time depends on individual metabolism and the dose of the drug. www.nida.nih.gov

Interpretation Detection Times Hallucinogens Substance Marijuana Single use Several times a week daily Period of Detection 1-3 days 2 weeks 3-6 weeks These are approximate detection times for the drug or metabolites in urine. The actual detection time depends on individual metabolism and the dose of the drug. www.nida.nih.gov