What Is Addiction?

Methadone and Buprenorphine Maintenance: Effective Treatments for Opiate Addiction

Mary Jeanne Kreek, M.D. Patrick E. and Beatrice M. Haggerty Professor Head of Laboratory The Laboratory of the Biology of Addictive Diseases The Rockefeller University Senior Physician The Rockefeller University Hospital

June 11, 2019 International Conference on Opioids Boston, MA

funded primarily by NIH-NIDA, NIH-NCRR and the Adelson Medical Research Foundation

What is Addiction?

Compulsive drug seeking behavior and drug self-administration, without regard to negative consequences to self or others.

(adapted from WHO)

“drug” = nicotine, alcohol – legal drugs prescription opiates, marijuana (cannabis) – medicines (?) heroin, cocaine – illegal drugs 2019

Natural History of Drug and Alcohol Abuse and Addictions relapse to addiction without pharmacotherapy 90% - opiate; Primary Possible Utility of Vaccines Medications Useful 60% - cocaine, alcohol Prevention and Selected Medications and Needed

Initial Sporadic Regular Addiction Early Protracted Self-Administration Intermittent Use Withdrawal Abstinence of Drug of Abuse Use (abstinence)

Progression sustain abstinence with no specific medications ADDICTION: Compulsive drug seeking behavior 10% - opiate; and drug self-administration, without regard to 40% - cocaine, alcohol negative consequences to self or others (adapted from WHO). Adapted from Kreek et al., Nature Reviews Drug Discovery, 1:710, 2002; 2019

1 Prevalence of Specific Drug Abuse and Vulnerability to Develop Addictions – 2019 National Household Survey and Related Surveys – 2007 – 2016 Heroin Use – ever ~ 5.2 million Heroin Addiction ~ 626,000 Illicit Use of Opiate Medication – ever ~ 37.1 million (i.e., 14.2% of the population 12 and over) Dependence on Opiate Medication ~ 2.1 million Opiate (heroin, fentanyl, other) Overdose Deaths ~ 72,300 (in 2017)*

Cocaine Use – ever ~ 40.5 million Cocaine Addiction ~ 966,000

Alcohol Use – ever ~ 216 million Alcoholism ~ 14.5 million

Marijuana Use – ever ~ 123 million Marijuana Daily Use ~ 4 million Development of Addiction After Self-Exposure to Specific Drugs Opiate Addiction ~ 1 in 5 to 1 in 15 (20% to 6.5%) Alcoholism, Marijuana, and Cocaine Dependency ~ 1 in 8 to 1 in 15 (12.5% to 6.5%) SAMHSA Nat’l Survey on Drug Use and Health, 2017; Others, 2007-18; *Nat’l Center for Health Statistics (CDC), 2019

• Drug overdoses, primarily opioids, killed more than 72,300 Americans in 2017, a record and a rise of approximately 10% over 2016 • Drug overdose deaths in 2017 were higher than the peak yearly deaths from HIV, car accidents, or gun Overdose Deaths in Thousands in Preceding 12 months deaths 30 thousand Synthetic opioids • Overdose deaths have begun to fall in Massachusetts, Vermont, and Rhode 20 Heroin Island following major Other public health opioids campaigns, including Cocaine increased access to 10 treatment, in response Other Psychostimulants to the early arrival of fentanyl in those states Methadone 0 Sanger-Katz, NY Times, 2015 2016 2017 Aug 15, 2018

RECOMMENDATIONS FOR ADDRESSING THE CRISIS OF OPIATE OVERDOSE AND ADDICTION IN THE UNITED STATES prepared by Miriam Adelson, MD and Mary Jeanne Kreek, MD (November 2017; Updated April 2019) PROBLEM AND PREVENTION

• Doctors, nurse practitioners, and dentists should not prescribe opiates for acute pain (short-term use) for longer than 7 days.

• Further, they should encourage the patient to return any unused opiates to the physician, dentist, or pharmacy immediately.

HISTORY OF PROBLEMS OF OPIOID OVERPRESCRIPTION AND MISUSE

• Guidelines surrounding treatment of pain need to be revised to include a clear definition of acute pain versus chronic pain, with additional clarification of what kinds of chronic pain may respond to treatment with opiates (e.g., cancer-related pain and some types of neuropathic pain).

• Types of pain which do not respond to chronic opiate treatment include pain related to essentially all types of arthritis, lower back pain, and many types of neurodegenerative diseases).

• There is urgent need for development of new pain medications with novel neurobiological targets to treat the disorders in which pain is not relieved by opiate medications.

Kreek, Adelson, 2017, 2019

2 Update on Laws Governing Prescribing of Opioids In Various States

“In 2016, Massachusetts became the first state in the nation to pass a law limiting first time opioid prescriptions to 7 days.”

(Of interest, at this meeting in June 2015, MJK, an early-morning speaker, had an invited public dialogue on this topic with newly elected governor, Charlie Baker, after his talk.)

“Since then, over half of all states have enacted laws that restrict the prescribing or dispensing of opioids for acute pain. To date, there is no data on whether and to what extent these laws mediate opioid-related morbidity and mortality.”

Subsequently, “fifteen states have passed laws limiting opioid prescribing for acute pain in an opioid naive patient to a 7-day supply. These states include Alaska, Hawaii, Colorado, Utah, Oklahoma, Louisiana, Missouri, Indiana, West Virginia, South Carolina, Pennsylvania, New York, Maine, Connecticut, Massachusetts. In addition, Arizona, North Carolina, and New Jersey limit initial prescribing to 5 days.”

Pharmacy Times, February 5, 2019

RECOMMENDATIONS FOR ADDRESSING THE CRISIS OF OPIATE OVERDOSE AND ADDICTION IN THE UNITED STATES prepared by Miriam Adelson, MD and Mary Jeanne Kreek, MD (November 2017; Updated April 2019)

ACUTE INTERVENTION FOR OVERDOSE.*

• Naloxone (Narcan) is the only available effective treatment for narcotic overdose. It should be widely communicated that naloxone is effective for only 60-90 minutes; repeated doses of naloxone must be given when a long-acting opiate (or large dose of short-acting opiate) has caused the overdose.

• For public health reasons, the cost of naloxone formulations should be kept appropriately low.

*NB – According to SysGalaxy Market Research report Global Naloxone Market 2013-2018 and Forecast 2019-2024 ($2500 to purchase report), parenteral or nasal naloxone in different ampules or total doeses available from ADAPT Pharma, Amphastar Pharmaceuticals, Pfizer, kaleo, Sandoz, Amneal Pharmaceuticals, West Ward Pharmaceuticals, and Mylan. Kreek, Adelson, 2017, 2019

RECOMMENDATIONS FOR ADDRESSING THE CRISIS OF OPIATE OVERDOSE AND ADDICTION IN THE UNITED STATES prepared by Miriam Adelson, MD and Mary Jeanne Kreek, MD (November 2017; Updated April 2019)

CHRONIC EFFECTIVE TREATMENT FOR OPIATE ADDICTION: MAINTENANCE TREATMENT WITH METHADONE OR BUPRENORPHINE-NALOXONE

• Access to methadone maintenance treatment and buprenorphine-naloxone maintenance treatment should be expanded throughout the nation. Also, more long-term medical maintenance clinics are needed in all States.

• The Federal Regulations surrounding criteria for entry into methadone maintenance treatment, which currently require one year of daily self- administration of a short accent opiate, should be appropriately reduced to match the criteria for entry into buprenorphine-naloxone treatment (i.e., when a physician identifies that opiate addiction exists, usually after three months of daily self-administration of any short-acting opiate).

• Appropriate counseling, or other psychiatric or psychological care, should be strongly encouraged to augment buprenorphine-naloxone treatment (as is already required for methadone maintenance treatment).

• Major medical centers should be encouraged to house onsite treatment programs so that patients may have access to top-level medical, psychiatric, and psychological health care. Kreek, Adelson, 2017, 2019

3 RECOMMENDATIONS FOR ADDRESSING THE CRISIS OF OPIATE OVERDOSE AND ADDICTION IN THE UNITED STATES prepared by Miriam Adelson, MD and Mary Jeanne Kreek, MD (November 2017; Updated April 2019)

ELIMINATION OF STIGMA

• Efforts should be made to educate the public that addictions are diseases of the brain, and not simply anti-social or criminal behaviors (as has been well- established by the scientific and medical community for over 50 years).

• Further, efforts should be made to educate the public that excellent treatments are available, and that opiate addiction usually requires long- term, chronic treatment with medications like methadone or buprenorphine- naloxone, in the same way that diseases like hypertension and diabetes require long-term treatment with appropriate medications.

Kreek, Adelson, 2017, 2019

Initial Development of Methadone Maintenance Treatment and Current Use of Maintenance Treatment with Methadone or Buprenorpine-Naloxone for Opiate Addiction

The Beginning: 1963-1964 Treatment of Heroin Addiction: Identification of Need, Formulation of Hypothesis, Basic Clinical and Related Laboratory Research, Translational Research, and Evaluations Autumn Vincent P. Dole, Jr., MD recruitment of two new staff 1963 members to his Laboratory of Physiology and Metabolism at The Rockefeller Institute for Medical Research:

1) Marie Nyswander, MD – psychiatrist, years of work with heroin addicts in New York City and Lexington, KY. Author of book The Addict as a Patient.

2) Mary Jeanne Kreek, MD – second year Resident in (Internal) Medicine (“PGY-2”) at Cornell-New York Hospital with experience in clinical and laboratory based research at NIH and Columbia P&S.

January – New team formed and first patients admitted to the July 1964 Rockefeller Hospital; initial research accomplished by July. AATOD 2013; EUROPAD 2014; 2019

4 55th Anniversary of Development of Methadone Maintenance Treatment

Vincent P. Dole, Jr., MD; Marie Nyswander, MD; and Mary Jeanne Kreek, MD

1964: Initial clinical research at The Rockefeller Hospital on development of treatment using methadone maintenance pharmacotherapy and on elucidating mechanisms of efficacy.

Dole, V.P., Nyswander, M.E. and Kreek, M.J.: Narcotic blockade. Arch. Intern. Med., 1966

Development of Methadone Maintenance Treatment – 1964 Onward 1964 HYPOTHESIS –NOW EVIDENCE-BASED FACT: Heroin (opiate) addiction is a disease – a “metabolic disease” – of the brain with resultant behaviors of “drug hunger” and drug self-administration, despite negative consequences to self and others. Heroin addiction is not simply a criminal behavior or due alone to antisocial personality or some other personality disorder. First publications describing methadone maintenance treatment research 1) 1964: Initial clinical research on development of treatment using methadone maintenance pharmacotherapy and on elucidating mechanisms of efficacy performed at The Rockefeller Hospital of The Rockefeller Institute for Medical Research: Dole, V.P., Nyswander, M.E. and Kreek, M.J.: Narcotic blockade. Arch. Intern. Med., 118:304-309, 1966. (also recorded in the Association of American Physicians meeting transcription of discussion) 2) 1965: Translational applied clinical research performed at Manhattan General Hospital: Dole, V.P. and Nyswander, M.E.: A medical treatment for diacetylmorphine (heroin) addiction. JAMA, 193:646-650, 1965. AATOD 2013; EUROPAD 2014; MACAU 2015

1964-1966: Early Expansion of Methadone Maintenance Treatment 1964-1967 – Initially all patients admitted as active or recently abstinent heroin addicts to the Rockefeller Hospital, unlocked research ward for inpatient induction into methadone maintenance treatment (minimum 6-8 weeks). Follow-up in Rockefeller Hospital Outpatient Department.

1965-1966 – Replication of methadone maintenance treatment research model in an inpatient unit of the Manhattan General Hospital, a fee-for-service proprietary hospital (later to become the Bernstein Institute of Beth Israel Hospital), which had been engaged in 10-14 day detoxification “treatment” for opiates (revolving door).

Kreek, AATOD 2013; EUROPAD 2014

5 1967-1972: Continued Early Expansion of Methadone Maintenance Treatment 1967-1968 – Introduction of outpatient induction into methadone maintenance treatment (at the Rockefeller Hospital clinic, Dr. MJ Kreek).

1967-1972 – Development of multiple satellite outpatient methadone maintenance treatment programs at Harlem, Beth Israel, St. Luke’s, and Roosevelt Hospitals in New York City.

1967-1972 – First replication of methadone maintenance program outside of New York City – Department of Psychiatry, University of Uppsala, Uppsala, Sweden (Dr. Lars Gunne and later Drs. Leif Grönbladh and Olof Blix).

1971 – Development of “Adolescent Development Program” in the Department of Public Health of Cornell Medical College – New York Hospital (Drs. Elizabeth Khuri and R. Millman).

1972 – Development of “Adult Clinic” in the Department of Medicine of Cornell Medical College – New York Hospital (Dr. Aaron Wells).

Kreek, AATOD 2013 ; EUROPAD 2014

Two Earliest Papers on Pharmacology and Medical Safety of Methadone as Used in Maintenance Treatment of Addiction

Impact of Short-Acting Heroin versus Long-Acting Methadone Administered on a Chronic Basis in Humans (1964 through 1978 Studies): Opioid Agonist Pharmacokinetics – Heroin Versus Methadone

Apparent Plasma Terminal

"High" Drug or Half-life and Duration of

Medication Desired Effects

HEROIN 3 min for prodrug "Straight"

30 min for active (Heroin)

"Sick" compound, mono-acetyl

Functional State Functional morphine (fast on-set and

AM PM AM PM AM off-set) Days

6 hours for active

metabolites (morphine "High" and others)

"Straight" METHADONE 24h for racemic (rs) medication (slow on-set

and off-set – steady-state (Methadone)

Functional State Functional "Sick" achieved)

48h for active (r) AM PM AM PM AM Days H enantiomer

Dole, Nyswander and Kreek, 1966; Kreek et al., 1973; 1976; 1977; 1979; 1982; Inturrisi et al, 1973; 1984; 2018

6 “On-Off” versus “Steady-State”: Relationship Between Blood (and Brain) Levels of Drugs of Abuse and Their Effects

Disruption versus Normalization • levels of gene expression • receptor mediated events • physiology • behaviors

Rates of rise of blood (and presumable brain) levels of drugs of abuse are related positively to their reinforcing effects Rates of fall of blood (and presumably brain) levels of drugs of abuse are related positively to the onset of withdrawal symptoms and/or acute “craving”

Kreek, 1978;1987;1991;1992; 2001

35th Anniversary of Identification of HIV-1 Infection in Drug Users in New York City and Protective Value of Methadone Maintenance Treatment: (1969 –) 1978 – 1992; 1983 – 1984 Study and Beyond 100

Percent of IV Drug Users Infected with HIV-1 75

% 50

0 1978 1979 1980 1981 1982 1983 1984 1985 1986 1987 1992

50% – 60% Untreated, street heroin addicts: positive for HIV-1 antibody

9% Methadone maintained since <1978 (beginning of AIDS epidemic): less than 10% positive for HIV-1 antibody

Kreek with Des Jarlais and others, 1984

Methadone Maintenance Treatment for Opiate (Heroin) Addiction – 2019

Number of patients currently in treatment: ~ 1.4 million worldwide

 USA: ~ 360,000  Europe: ~ 600,000  Rest of world: ~ 400,000 Efficacy in “good” methadone treatment programs using adequate doses (80 to 150mg/d): Voluntary retention in treatment (1 year or more) 60 – 80% Continuing use of illicit heroin 5 – 20% Actions of methadone treatment: • Prevents withdrawal symptoms and “drug hunger” • Blocks euphoric effects of short-acting narcotics • Allows normalization of disrupted physiology Mechanism of action: Long-acting medication (24h half-life for racemate in humans) provides steady levels of opioid at specific receptor sites. • methadone found to be a full mu opioid receptor agonist which internalizes like endorphins (beta-endorphin and enkephalins) • methadone also has modest NMDA receptor complex antagonism Kreek, 1972; 1973; 2019

7 Limited Targeted Pharmacotherapies Available for Specific Addictive Diseases I. Opiate Addiction (Heroin and Illicit Use of Opiates) a. METHADONE (oral; 60-80%)** b. BUPRENORPHINE (+/- NALOXONE) (sublingual; 40-50%)**; BUPRENORPHINE 30 DAY IMPLANT (no data; approved November 2017) [c. NALTREXONE / SUSTAINED RELEASE NALTREXONE (<15%)*]

II. Alcohol Addiction and Excessive Alcohol Use a. NALTREXONE (30-40%)* b. NALMEFENE (approved in Europe only, 2012) c. ACAMPROSATE (low in USA)

III. Cocaine, Amphetamines and Other Stimulants NONE (%) is % of unselected persons with specific addictions who can be retained voluntarily in treatment for 3 months (*) or 12 months (**), with success in eliminating specific drug use. According to the National Institute on Drug Abuse, every year drug and alcohol misuse costs the United States $64 billion in healthcare and a total of $600 billion in healthcare, crime and criminal justice, and loss of productivity costs. By comparison, cancer costs $172 billion annually.

Effective treatment saves around $12 for every $1 spent. Kreek, 2019

Methadone synthetic mu opioid receptor full agonist NMDA partial antagonist

Buprenorphine thebaine-derived mu opioid receptor partial agonist kappa opioid receptor partial agonist

Naltrexone thebaine-derived mu opioid receptor full antagonist kappa opioid receptor partial agonist

Kreek 2019

Status of Methadone, Buprenorphine, and Extended Release Naltrexone Treatments for Opioid Addiction in the United States: Decrease, then Increase, of Numbers in Treatment 2015-2017 (2015, 2016, and 2017 data, SAMHSA, 2018)* US Patients in Treatment

Treatment 2015 2016 2017 Methadone 356,843 345,443 382,867 Maintenance (-11,400; -3.2%) (+37,424; +10.8%)

Buprenorphine 75,723 61,486 112,223 Maintenance (-14,237; -18.8%) (+50,737; +82.5%)

Extended 7,035 10,128 23,065 Release (+3,093; +44.0%) (+12,937; +128.7%) Naltrexone

Source: Center for Behavioral Health Statistics and Quality, Substance Abuse and Mental Health Services Administration (SAMHSA), National Survey of Substance Abuse Treatment Services (N-SSATS), 2018; Kreek 2019

8 Targets of Currently Approved Treatments for Addictive Disorders

Kreek et al, Journal of Clinical Investigation, 12: 3387, 2012

Opiate (Opioid) Analgesic Agents ~2008-2019 • Numerous – all mu opioid receptor-directed • Diverse formulations – many long-acting • Some available in oral, sublingual, spray, film, or patch preparations Examples – mostly short-acting Sustained Release/Long-Acting(*) Morphine (e.g., MS Contin®) Oxycodone (e.g., OxyContin®) Hydromorphone (e.g., HydromorphoneContin®)** Oxymorphone Acetaminophen and Hydrocodone (e.g., Vicodin®) Fentanyl (and related compounds) (e.g., Duragesic®) Meperidine Codeine Methadone* (long-acting) (e.g., Dolophine®) – full mu agonist Buprenorphine* (long-acting) (e.g., Subatex® sl) – partial mu agonist Buprenorphine-Naloxone* (long-acting) (e.g., Suboxone® sl) – partial mu agonist ** Removed from market by FDA shortly after approval Kreek, 2008; 2015; 2019

Use of Methadone as an Analgesic: Doses Usually Range from 0.5mg tid to 10mg tid – Rarely Over 60 mg/d • Very few indications for use of methadone in management of acute pain – because of long-acting pharmacokinetic properties, accumulation of medication must be anticipated over the first four days or longer. Accidental overdose deaths frequently occur when methadone is so prescribed.

• Methadone is a superb (and often superior) opioid analgesic for chronic pain, including neoplastic and non-neoplastic pain – because tolerance develops more slowly than to other opioid analgesics, steady doses may be used over long periods of time and escalation of dose may be gradual; no drowsiness, no dulled responses.

Kreek, 1975 on; 2003; 2008; 2015

9 Neurobiologic Basis of Opiate and Other Addictive Diseases

Factors Contributing to Vulnerability to Develop a Specific Addiction

use of the drug of abuse essential (100%)

Genetic Environmental (25-60%) (very high) • DNA • prenatal • SNPs • postnatal • other • contemporary polymorphisms • cues • comorbidity • stress-responsivity

• mRNA levels • neurochemistry • peptides • synaptogenesis • proteomics Drug-Induced Effects • behaviors (very high) Kreek et al., 2000; 2005

Development of an Addiction

•Drugs alter normal brain networks and chemicals

•“Rewarding” or “pleasurable” effects of drugs (the so-called “reinforcing effects”) involve: – Dopamine – Endorphins (acting at Mu Opioid Receptors)

•“Countermodulatory” response to reward involves: – Dynorphins (acting at Kappa Opioid Receptors)

10 Frontal Striatum cortex Substantia DOPAMINE nigra SYNAPSE

Nucleus accumbens VTA

National Institute on Drug Abuse

Heroin (or other opiates) increases dopamine release by acting at mu opioid receptors to inhibit GABAergic inhibition in the SNc and VTA Cocaine blocks dopamine re-uptake at synapse

Kreek, 2016 Maisonneuve, 1994

Development of an Addiction: Stress and Atypical Responsivity to Stressors – HPA Axis

Atypical responsivity to stress and stressors hypothalamus may contribute to the – Arginine persistence of, and CRF Vasopressin relapse to, self- + administration of drugs Anterior of abuse and thus to Endogenous – pituitary + Opioids addictive diseases. POMC (mu – inhibition) (kappa – ? activation) Such atypical stress b-End responsivity in some Cortisol ACTH individuals may exist prior to use of addictive adrenal + drugs on a genetic or acquired basis, and increase the vulnerability to develop

Kreek, 1972; 1981; 1982; 1984 … 2019 an addictive disease.

STRESS RESPONSIVITY – Heroin, Cocaine, and Alcohol Profoundly Alter Stress Responsive Hypothalamic-Pituitary-Adrenal (HPA) Axis: Normalization During Methadone Treatment • Acute effects of opiates Suppression of HPA Axis • Chronic effects of short-acting (decrease levels of HPA opiates (e.g., heroin addiction) hormones)

• Opiate withdrawal effects * Activation of HPA Axis • Opioid antagonist effects (increase levels of HPA • Cocaine effects * Hormones) • Alcohol effects

• Chronic effects of long-acting opiate (e.g. methadone in Normalization of HPA Axis maintenance treatment)

* Our challenge studies have shown that a relative and functional “endorphin deficiency” develops.

Kreek, 1972; 1973; 1987; 1992 … 2010

11 STRESS RESPONSIVITY – Normalization of Heroin Disrupted Physiology During Methadone Maintenance Treatment: PET Studies of mu Opioid Receptors in Human Brain Regions Using

the Antagonist [18F] Cyclofoxy

Normal volunteers n=14 16 MMTP volunteers n=14 (~20-30 percent reduction in 14 mu-opioid receptor binding due to methadone occupancy) 12

2 Specific Binding Binding (ml Specific plasma/mltissue) 0 Thl Amy Caud Ins ACg Put MT MFr Par Crb IT Hip WMt Area related to pain response Region of Interest Kling et al., . J. Area with dopamine terminals involved in “reward”, mood, and decision-making Pharmacol. Exp. Area with dopamine terminals involved in memory, learning, and movement Ther., 295, 2000

Countermodulation of Reward – Reversal or Modulation of Drug or Task-induced Stress:

Kappa Opioid Receptor-Dynorphin Neurobiology

COUNTERMODULATION – KAPPA OPIOID RECEPTOR-DYNORPHIN SYSTEM: Cocaine Increases Kappa Opioid Receptor Density in Rat, But Kappa Opioid Receptor Directed Dynorphins Also Increase Persistently (Opiates and Alcohol Also Increase Dynorphins)

7 1 2 Saline 14 day cocaine (15mg/kg x 3) 6

Control Cocaine 5

3 4

pg ppDyn mRNA RNA /total µg mRNA ppDyn pg Control Cocaine 0 Caudate Putamen Nucleus Accumbens Dynorphin Acting at the Kappa Opioid Receptor Lowers Dopamine Levels and Prevents Surge After Cocaine

Spangler… and Kreek, Brain Res. Mol. Brain Res., 19:323-327, 1993; Unterwald, Rubenfeld, and Kreek , NeuroReport, 5:1613, 1994; Spangler, Ho, Zhou, Maggos, Yuferov, and Kreek , Mol. Brain Res., 38:71, 1996

12 COUNTERMODULATION OF REWARD: Natural Dynorphin A1-17 (Kappa Opioid Receptor Agonist) Infusion into Mouse Striatum Lowers Basal and Cocaine Induced Dopamine Levels

10 10

8 8

6 6

4 4

Dopamine in Dopamine

Dopamine in Dopamine Dialysate (nM) Dialysate 2 (nM) Dialysate 2

0 0 60 120 180 60 120 180 Infusion Infusion Injection 20-min Sample 20-min Sample Dynorphin Dose (nmol) Infusion and Injection 0 2.0 Control 1.0 4.4 Cocaine (15mg/kg) 4.4+nBNI Dynorphin (4.4nmol) (antagonist) + Cocaine (15mg/kg)

Zhang, Butelman, Schlussman, Ho, and Kreek, Psychopharmacology, 172:422, 2004

Stress Manifested by Immobility During Forced Swim in Rats is Due To Increased Dynorphin Levels: Effect Reversed by Kappa Opioid Receptor Antagonist (nor-BNI) in Dose-Dependent Manner

60 nor-BNI Dose

50 0 mg/kg (n=6) * 5 mg/kg (n=6)

40 10 mg/kg (n=6)

Swim 15 Minutes Day 1; 30 1 hour later pre-treatment Score nor-BNI Swim 5 Minutes Day 2 20 (* - p<0.05)

Immobility

Reed et al., Neuroscience, 220:109-118, 2012

COUNTERMODULATION – KAPPA OPIOID RECEPTOR – DYNORPHIN SYSTEM: “BIOMARKER” Dynorphin A Lowers Tuberoinfundibular Dopaminergic Tone, which Tonically Inhibits Prolactin Release

Dose-Response Effects of Dynorphin A1-13 on

Prolactin Levels (BIOMARKER) in Normal Volunteers

Dynorphin A1-13 35 500 µ g/kg 120 µg/kg – Hypothalamus 30 Placebo (n=10) + TIDA 25

10 –

anterior pituitary 5 lactotropes (ng/ml) Levels Prolactin 0

-10 0 10 20 30 40 50 60 75 90 120 150 180 240

Time after injection (min)

Kreek et al., 1994; 1999; 2016

13 TRANSLATIONAL RESEARCH: IDENTIFICATION OF POTENTIAL TARGET FOR NOVEL PHARMACOTHERAPIES FOR COCAINE ADDICTION OR ALCOHOLISM – KAPPA OPIOID RECEPTOR / DYNORPHIN SYSTEM • Novel target for possible treatment of cocaine addiction, alcoholism, and stimulant or alcohol co-dependence with opioid addiction. (Our laboratory and others have shown that heroin, morphine, cocaine, and alcohol and also stress (e.g., forced swim test) increase dynorphin gene expression in rodents.) • Kappa Opioid Receptor Full Agonist – lowers basal dopaminergic tone and decreases drug-induced dopaminergic surge – ? reduction of reward after cocaine, alcohol, or opiate use – ? but possibly with persistent dysphoric side-effects (? avoid by use of biased or partial kappa agonist) • Kappa Opioid Receptor Antagonist – reduction of depressive symptoms, stress-related, spontaneous, or drug use-induced increase in dynorphin levels (shown in rats), ? decreased relapse to drug use but possible increases in dopamine surge after drug or alcohol self-administration • Kappa Opioid Receptor Partial Agonist (antagonist and agonist activity) – ? reduces dopaminergic surge and modulates dopaminergic tone – ? thus reduction of reward after cocaine, alcohol, or opiate use, and ? reduction of depressive symptoms – ? reduction of drug self-administration Kreek, AD Dunn, AM Dunn, Butelman, Reed, in preparation, 2019

Kappa Opioid Receptor Agonists or Partial Agonists: Unbiased or Biased Agonism

•Kappa Opioid Receptor member of 7-transmembrane GPCR family

•Prototype unbiased kappa opioid receptor agonists G-protein β-arrestin U50,488 and U69,593 ? ? •Biased agonism: differentially Analgesia Sedation activate intracellular Motor incoordination

pathways ?

(e.g., G-protein vs. β-arrestin) Anhedonia Reed, AD Dunn, Butelman, Kreek, CPDD 2017, in preparation, 2019

Compounds Approved for Use in Human Therapeutics with KOPr Partial Agonism in Addition to Mu-Opioid Receptor Antagonism or Partial Agonism; Binding Affinity in Cloned Human Receptors

Naloxone Naltrexone Nalmefene Buprenorphine MOP-r affinity 0.66 0.11 0.24 0.21 Ki (nM) KOP-r affinity 1.2 0.19 0.083 0.62 Ki (nM) DOP-r affinity 120 60 16 2.1 Ki (nM) Bidlack Laboratory (U Rochester); Bart… Bidlack et al, Neuropsychopharmacol, 30:2254, 2005; Kreek, Reed, Butelman 2014

14 REVERSAL OF DYNORPHIN-KAPPA INDUCED STRESS – Short-Acting Selective Kappa Opioid Receptor Antagonists

LY2444296 – “Tool” compound LY2456302 – “OpraKappa” (Lilly – Laboratory) (Lilly – Clinical)

CERC-501 (now owned by Jansen of Johnson & Johnson and under study for treatment of depression) Kreek, 2019

Pretreatment with “tool compound” (LY2444296) reduces anxiety- and depression-like behaviors in rats 30h in withdrawal from extended-access (18h/d, 14d) cocaine self-administration

FORCED SWIM TEST ELEVATED PLUS MAZE 150 30h Vehiclepost 14d cocaine SA: 50 30h post 14d cocaine SA: VehicleLY2444296 3 mg/kg vehicle 120 LY2444296 3 mg/kg 40 LY2444296 3 mg/kg 90 30 * 60 20 * Time 30 10

Immobility (s) Immobility

0 arms (s) open in 0 300300 300

225225 225

150 150 ** 150 75 75 75

Latency to Latency

immobility (s) immobility ***

immobility (s) immobility 0 0 enter to Latency arms (s) open in 0 Valenza et al., Psychopharmacology, 234: 2219, 2017

“OpraK” Clinical Study (LY2456302): kappa opioid receptor antagonist We have hypothesized that a selective kappa opioid receptor (KOPr) antagonist might be helpful in managing the dysphoric and depressive symptoms of early and protracted abstinence from cocaine or alcohol. However, we also have hypothesized that a KOPr antagonist might increase the reward of cocaine or alcohol by increasing baseline dopaminergic tone and drug-induced dopamine surges. There is only limited data available on the impact of selective KOP-r antagonism in humans. We have conducted an in-patient basic research study that examined neuroendocrine and behavioral effects of a novel short-acting selective KOP-r antagonist, LY2456302 (which we call “OpraK”) in normal volunteers (n=40) and in volunteers diagnosed with cocaine dependence (DSM IV) (n=30). Four days of drug administration caused no adverse effects. Reed et al, Neuropsychopharmacology, 43:739, 2017

15 “OpraK”: Prolactin Levels – Male Normal Volunteers vs. Early Abstinence Cocaine Dependent Volunteers – No Evidence of Kappa Partial Agonism 25 Baseline, Day 1 20 1st OpraK (10mg), Day 2 th 15 4 OpraK (10mg), Day 5

10 Normal Volunteers

5 (n=24)

-30 0 30 60 90 120 150 180 210 240 270 300 330 360 390 420 450 480 serum prolactin (ng/mL) 25 Time (min) Baseline, Day 1 20 1st OpraK (10mg), Day 2 4th OpraK (10mg), Day 5 15

10 Early Abstinence Cocaine Dependent 5 Volunteers 0 (n=19)

-30 0 30 60 90 120 150 180 210 240 270 300 330 360 390 420 450 480 serum prolactin (ng/mL) TimeTime (min)(min) Reed et al, Neuropsychopharmacology, 43:739, 2017

“OpraK”: Serum ACTH and Cortisol Levels AUC – Normal Volunteers vs. Early Abstinence Cocaine Dependent Volunteers: Evidence of Partial Mu Opioid Receptor Antagonism

ACTH p<0.05 Cortisol p<0.0005 18000 p<0.005 6000 p<0.0005

16000 5000 14000

12000 4000

10000 3000 8000

6000 2000

4000 1000 2000

0 Area Under the Curve, 0-480 minutes, cortisol 0 Area Under the Curve, 0-480 minutes, ACTH Baseline-DayBaseline 1 1st OpraKappa-Day1st OpraK 2 4th OpraKappa-Day4th OpraK 5 Baseline-DayBaseline 1 1st OpraKappa-Day1st OpraK 24th OpraKappa-Day4th OpraK 5 Day 1 (10mg)Day (10mg) Day 1 (10mg)Day (10mg) Day 2 Day 5 Day 2 Day 5 HV (n=16) HV (n=39) EACD (n=13) EACD (n=23) Reed et al, Neuropsychopharmacology, 43:739, 2017

Genetic Contributions to the Vulnerability to Develop Opioid Addiction

16 Genetic Vulnerability to (or Protection from) Development of an Addiction

Genetic vulnerability to develop an addiction once self-exposed probably due to: •Multiple variants (SNPs and other types) of •Multiple genes (as with any complex disorder, e.g., hypertension, diabetes) •Probably both shared and also unique variants for each specific addiction •Genetic contributions of comorbid conditions and personality types may play a role No gene variant or group of variants causes an addictive disease alone; self-administration of a

drug is essential. Kreek, 2008

Genetic Variants of the Human Mu Opioid Receptor: Single Nucleotide Polymorphisms in the Coding Region Including the Functional A118G (N40D) Variant

HYPOTHESIS (C17T)

Gene variants: (A118G) • Alter physiology “PHYSIOGENETICS”

• Alter response to medications “PHARMACOGENETICS”

• Are associated with specific addictions

Bond, LaForge… Kreek, Yu, PNAS , 95:9608, 1998; Kreek, Yuferov and LaForge, 2000

FUNCTIONAL MOP-r (A118G) VARIANT – Enhanced Binding and Coupling to G Protein-Activated, Inwardly Rectifying K+(GIRK) Channels by Beta-Endorphin Acting at A118G Variant Compared with Prototype A118A

100

A118G A118G 1.0 Prototype Prototype 80

0.5 40

Percent Bound Percent 20

Current Response Current Fraction Maximum Fraction 0 0

-11 -10 -9 -8 -7 -9 -8 -7 -6 Log [b Endorphin (M)] Log [b Endorphin (M)]

Bond, LaForge… Kreek, Yu, PNAS , 95:9608, 1998; Kreek, Yuferov and LaForge, 2000

17 FUNCTIONAL MOP-r (A118G) VARIANT – “Physiogenetics” Related to A118G Variant of Human Mu Opioid Receptor Gene – Alters Stress Responsivity in Healthy Control Volunteers

P = Placebo

N = Naloxone

2500 Cortisol 24 A/A (n=29)

22 A/G (n=7) N N

2000 P < 0.05 AUC

20 - 1500 18 16 N 1000 14

10:30am + + 90min) 10:30am P - 500 40 19 12 I N

Serum Cortisol (ug/dl) Cortisol Serum 10 (no food for hours) 9 food (no

Cortisol Levels Levels Cortisol 0 8 (9:30am Prototype A118G 50 0 50 100 150 200

Time (min)

Bart et al. Neuropsychopharmacology, Wand et al., Neuropsychopharmacol, 26:106, 2002 31:2313-2317, 2006 Chong…Wand, Neuropsychopharmacology, 31:204, 2006

Association Between a Functional Polymorphism (SNP) in the Mu Opioid Receptor Gene (A118G) and Opiate Addiction and Also Alcoholism in Central Sweden

Opiate Dependent (n=139) Control (n=170)

118G Allele Frequency 0.155 0.074 (15.5%) (7.4%) Odds Ratio=2.86 p=0.00025 In the entire study group in this central Swedish population: Attributable Risk due to genotypes with a G allele: 18% Bart et al., Molecular Psychiatry, 9:547-549, 2004

Alcohol Dependent (n=389) Control (n=170) 118G Allele Frequency * 0.125 0.074 (12.5%) (7.4%) Odds Ratio=1.92 p=0.0074 * Overall 118G Allele Frequency = 0.109 (10.9%) In the entire study group in this central Swedish population: Attributable Risk due to genotypes with a G allele: 11.1%

Bart et al., Neuropsychopharmacology, 30:417, 2005

Genetically Modified A112G Mice, A Model of the Human A118G Mu Opioid Receptor Functional Variant: Microdialysis in Striatum of Wild-Type AA ( ) versus Genetically Modified GG ( ) Mice: Absolute Dopamine Levels (Three Baseline Samples) and Levels of Dopamine after Heroin Injections

Males Females

14 14

GG (6) 12 GG (6) 12 10 10 8 8 6 6 4 4 AA (6) AA (7)

2 2

Dopamine (nM) Dopamine (nM) Dopamine 0 0

10mg/kg 20mg/kg 10mg/kg 20mg/kg Heroin Heroin Heroin Heroin

Zhang, Blendy…Kreek et al., Neuropsychopharmacology, 40:1091, 2015

18 Genetically Modified A112G Mice (Asparagine to Aspartic Acid), A Model of the Human A118G Mu Opioid Receptor Functional Variant: Heroin versus Oxycodone Self-Administration (10d, 4h/d) by

Wild-Type AA ( ) versus Genetically Modified GG ( ) versus Mice )

4 0 1 0 ) 4 0 1 0

) g

Males – Heroin Females – Heroin )

g m

8 8 g 5

3 0 5 3 0

(

A ,

6 6 A

= n

R 2 0

2 0 n

(

r 4

1 0 o 1 0 o A A (12)

A A (1 5 ) y

y p

2 l p

2 l

i e

e G G (1 2 )

a a

s G G (1 2 )

D N 0 0 N 0 0

1 2 3 4 5 6 7 8 9 1 0 1 2 3 4 5 6 7 8 9 1 0

)

) )

g Day (4h/d) Day (4h/d)

g g 6 0 1 5 g

k 6 0 1 5

/ g

Males – Oxycodone g Females – Oxycodone

(

A 0 4 0 1 0 0

S 4 0 1 0

(

c s

2 0 5 2 0 5 c

O O

O x y -A A (7 ) O x y -A A (8 )

i s

O x y -G G (6 ) s O x y -G G (8 )

a o

0 0 o 0 0

D N 1 2 3 4 5 6 7 8 9 1 0 N 1 2 3 4 5 6 7 8 9 1 0 Day (4h/d) Day (4h/d) Zhang, Blendy…Kreek et al., Neuropsychopharm, 40:1091, 2015; unpublished data presented at CPDD 2018

ASSOCIATION WITH OPIATE ADDICTION IN CAUCASIANS, AFRICAN AMERICANS, AND BOTH Neuro- Opioid Stress transmitters Total Genes 5/0 13/11 42/34 60/45 SNPs in Genes 11/0 22/12 48/54 81/66 Genes Replicated 3/0 6/3 13/17 22/20 SNPs Replicated 6/0 7/1 2/30 15/31 OPIOID SYSTEM (selected genes) STRESS SYSTEM (selected genes) OPRM1 AVPR1A (mu opioid receptor) (arginine vasopressin receptor 1A)

OPRD1 FKBP5 (delta opioid receptor) (FK506-binding protein 51/ corticosterone chaperone) GAL OPRK1 (galanin) (kappa opioid receptor) CSNK1E PDYN (casein kinase 1, epsilon) (dynorphin peptide) CRHBP (Corticotropin Releasing Hormone Binding Protein)

updated after Reed et al., Current Psychiatry Reports, 16(11): 504, 2014 Kreek Lab: Bond, Yu, LaForge, Nielsen, Levran, Randesi, Yuferov, and others; Kreek 2019

COMT GABRG1 (catechol-o-methyltransferase) (gamma-aminobutyric acid (GABA) A receptor) HTR1B (serotonin receptor 1B) GRIN2A (glutamate receptor, ionotropic, N- BDNF methyl D-aspartate 2A) (brain-derived neurotrophic factor) GAD1 (glutamate decarboxylase 1)

updated after Reed et al., Current Psychiatry Reports, 16(11): 504, 2014 Kreek Lab: Bond, Yu, LaForge, Nielsen, Levran, Randesi, Yuferov, and others; Kreek 2019

19 Recent Published Results from Our Laboratory on Association of Specific Gene Variants with Opiate and/or Cocaine Addiction (over 145) Using AIMS Markers to Define Ethnicity Circadian Adrenergic Rhythms Stress Dopamine Cholinergic Serotonin

Signal Transduction GABA Opioid

African European 12 African African cluster sample genes sample Descent (24 genes) (35 genes)

European Descent European cluster Shared SNPs, e.g.: DRD2: rs1076563 rs2587546 Kreek 2019 after Levran 2015

The Laboratory of the Biology of Addictive Diseases – 2019 Laboratory Scientists Postdoctoral Fellows Guest Investigators Eduardo Butelman Kyle Windisch Miriam Adelson Yan Zhou Devon Collins Gavin Bart Orna Levran Lawrence Brown Yong Zhang Graduate Students Don Des Jarlais Vadim Yuferov Amy Dunn David Novick Brian Reed Einat Peles Assistants for Research Ellen Unterwald Laboratory Manager Ariel Ben-Ezra Matthew Randesi Jose Erazo Other Rockefeller Michelle Morochnik University Collaborators Research Nurse Carina Chen Brian Chait Practitioner Bryan Elroy Jeff Friedman Kate Brown Paul Greengard (d. 2019) Statistics & Informatics Bruce McEwen Administrative Team Collaborators Don Pfaff Kitt Lavoie Jurg Ott Sid Strickland Abigail Sintim Yupu Liang Tom Tuschl Funded primarily by Dr. Miriam and Sheldon Adelson Medical Research Foundation, NIH-NIDA, NIH-NIAAA, NIH-CRR, Tri-Institutional Therapeutics Discovery Institute, Robertson Therapeutic Development Fund, and others

Kreek Laboratory 2019

20 21