Using Imaging to Assess Effects of THC on Brain Activity

Marijuana has become the most popular illicit drug in the western world (Substance Abuse and
Mental Health Services Administration, 2004). Both acute and chronic exposure to cannabis is
associated with changes in brain activity in frontal, limbic and also cerebellar regions,
with generally increased brain activity following acute exposure and generally decreased
activity during abstinence (Quickfall, 2006). Regions showing differences in brain activity
involve the extended dopamine reward pathways and the frontocerebellar network.

Most brain imaging studies of acute marijuana exposure have demonstrated global cortical
activity increases during administration of smoked cannabis or infused THC. Several
techniques have been used to track alterations of the brain associated with marijuana
intoxication (Mathew and Wilson, 1992, Levin, 2001 and Mathew et al., 1997). Using positron
emission tomography (PET), studies have shown that marijuana intoxication is associated with
global and rCBF increases that are most notable over the frontal regions and, in right-handed
subjects, in the right hemisphere, (Mathew et al., 1997). Ratings of intoxication and
euphoria correlated with the right hemispheric rCBF increase (Mathew et al., 1997). In
another PET study, Volkow et al. (1991a) showed that increased cerebellar metabolism during
intoxication, correlated with degree of intoxication and plasma THC levels (Volkow et
al.,1991b). FMRI studies have also shown increases in brain activity when individuals are
intoxicated. In two pharmacological fMRI studies, Bossong and colleagues investigated the
acute effects of THC, administered with a Volcano vaporizer (6 mg), and found that activity
during a memory task was significantly increased in a network of memory-related brain
regions, particularly when workload increased.

An alternative technology to PET and fMRI that has not been used to assess the effects of THC
on brain function is functional near-infrared spectroscopy (fNIRS). fNIRS is a neuroimaging
technology for mapping the functioning human cortex that exploits the principles of
near-infrared spectroscopy (NIRS). An fNIRS headset can be quickly fixed to the forehead and
enables measurements of the prefrontal cortex (PFC) within minutes. NIRS is based on the fact
that: 1) human tissues are relatively transparent to light in the NIR spectral window
(650-1000 nm); 2) NIR light is either absorbed by pigmented compounds (chromophores) or
scattered in tissues; 3) NIR light is able to penetrate human tissues (Delpy and Cope, 1997);
and 4) the relatively high attenuation of NIR light in tissue is due to hemoglobin, the
oxygen transport red blood cell protein, located in small vessels of the microcirculation,
such as capillary, arteriolar and venular beds. NIRS is weakly sensitive to blood vessels >1
mm because they completely absorb the light. The absorption spectrum of hemoglobin depends on
its level of oxygenation. NIRS is a non-invasive, safe technique that utilizes 1) laser diode
and/or light emitting diode light sources spanning the optical window between 650 and 1000
nm, and 2) flexible fiber optics to carry the NIR light to (source) and from (detector)
tissues. Fiber optics are suitable for any head position and posture. Adequate depth of NIR
light penetration (almost one half of the source-detector distance) can be achieved using a
source-detector distance around 3 cm. fNIRS has been used in several fields in which fMRI is
impractical due to the constraints of the fMRI scanning environment. fNIRS can be utilized,
for example, in infant and children developmental studies, in neurorehabilitation assessment,
and in simultaneous brain activation studies on multiple subjects. In this study, we will use
fNIRS to assess the effects of THC intoxication on hemodynamic response and connectivity in
the prefrontal cortex.

Dronabinol (marinol) is an FDA approved, synthetic, orally active cannabinoid used for
appetite stimulation that has been shown to be safe for multi dose use at doses up to 210 mg
per day, although after dosing of 210 mg per day for 16 days, a withdrawal syndrome was
described with abrupt discontinuation. Doses of 2.5-20 mg per day are used for appetite
stimulation and antiemetic indications. Doses of 0.5 mg/kg body weight reliably produce
intoxication (easy laughing, elation and subjective heightened awareness)

II. SPECIFIC AIMS Aim 1. Assess the effects of THC intoxication using dronabinol (synthetic
THC) on the HbO signal during resting state and task-based activation in the prefrontal
cortex (PFC) and resting state connectivity, as well as on neurocognitive task performance
and correlations between these measurements and clinical signs of intoxication.

Aim 2. Investigate at which doses of dronabinol an effect on neurocognitive task performance
or an effect on PFC activity can be observed.

Aim 3. Examine potential interaction following co-administration of THC with oral ethanol
exposure in healthy volunteers.

Participants: 150 adults who use marijuana at least monthly (aged 18-55) will be recruited to
participate in this Phase IIa study of dronabinol vs placebo. Subjects will be asked to
refrain from using marijuana within 24 hours of the start of study procedures. Since THCCOOH,
the main secondary metabolite of tetrahydrocannabinol (THC), can be detected in urine several
weeks after last use, a negative urine toxicology screen for THC-COOH will not be required,
participants will be admitted to the inpatient research unit of Massachusetts General
Hospital the night prior to start of study procedures on each study day after screening, and
participants who arive exhibiting overt signs of intoxication (e.g. congestion of the
conjunctival blood vessels (red eyes), slowed speech response, giddiness) will not be tested
the next morning.

Study Drug and Dosing: Participants will be given up to 50 mg of dronabinol, an FDA-approved
synthetic form of THC that is used to treat loss of appetite that causes weight loss in
people with AIDS. THC is the principle psychoactive drug in marijuana. The study physician,
Dr. A. Eden Evins MD, as well as physicians on study staff at the Center for Addiction
Medicine, will be responsible for administering the drug.

Dronabinol will be dosed to 50mg per study visit. The rationale for this dose is based on
several factors. The study will be conducted in regular cannabis users who present at their
first study visit with a positive urine screen for THC metabolites. Chronic cannabis use is
associated with significant tachyphylaxis and tolerance to pharmacologic effects of THC and
other cannabinoids. In healthy males who received 210 mg/day dronabinol for 16 days,
tolerance developed to the CNS effects of dronabinol (including the euphoric 'high' effect)
within 12 days. At doses of 5 mg/day for appetite stimulation in 139 patients, only 18%
experienced ay CNS effect (feeling high, dizziness, confusion, somnolence). These CNS effects
are the target symptoms we wish to assess in study participants. Because of tolerance, the
rate of CNS effects is expected to be lower in our study population than in non-cannabis
users who participated in dronabinol studies for anti-emetic and appetite stimulation
indications.

Dronabinol was evaluated in 454 patients with cancer at doses up to 40 mg/day. At doses
greater than 0.175 mg/kg approximately 58% reported feeling 'high'. The starting recommended
antiemetic dose is 15-20mg/day, noting that dosage may be escalated during a chemotherapy
cycle or at subsequent cycles based upon initial results.

For appetite stimulation, the recommended dose range is up to 20 mg/day, and at this dose
approximately half of patients tested have reported CNS symptoms targeted in this study
(feeling high, dizziness, confusion, somnolence).

Thus, we will dose at up to 50mg and will assess for CNS symptoms frequently during the study
visit. The exact dronabinol dose will be clinically determined by a study physician. The dose
will be designed to minimize adverse psychologic and physiologic effects of dronabinol and
will be primarily based on the degree of tolerance expected (based on dose and frequency of
cannabis use), self report of degree of intoxication experienced with each use, and any
regular or prior negative psychological reactions to cannabis use or binges. This is an
inexact determination as potency of cannabis used is almost never known. Other factors that
will be considered when determining dose are height, weight, BMI, and baseline blood
pressure. For example: Heavy daily cannabis users will generally receive 50 mg. Daily
cannabis users who use 1/2-1 gram per day and who weigh over approximately 140 pounds will
generally receive 50 mg, while those who use less than half a gram per day, use less than
daily or who weigh less than 120 pounds will receive between 30 and 45 mg based on the
physician's best judgment.

Urine Drug Screen: Before study procedures are initiated, we will perform a qualitative urine
drug screen (Multi Drug 6 Panel Urine Test; Medimpex United Inc., Bensalem, PA) that will
test for marijuana, amphetamines, methamphetamines, cocaine, and opiates. These rapid tests
are simple, can be done at home, and are CLIA-waived. The test will be administered by a
trained research coordinator.

Urine Pregnancy: Before study procedures are initiated, we will perform a qualitative urine
pregnancy test for females. The test will be administered by a trained research coordinator.

Overview of Procedures Phase 2A. Investigate the effect of THC on fNIRS brain signature and
its association with self-reported intoxication, laboratory measures of impairment, and the
gold-standard behavioral field test of driving impairment used by law enforcement, the
primary classifier.

Subjects will be scheduled for 2 study sessions. In a double-blind, placebo-controlled,
random order cross-over study of single dose THC (dronabinol) at physician determined doses
of 20-50mg, participants will receive THC capsules or identical placebo. The MGH research
pharmacy will generate a blinded randomization code for order of dosing and will dispense
blinded drug in the dose ordered and identical placebo for use on separate study days. In
order for a complete washout of effects, and to minimize the effect of repeat testing for the
neuropsychology tasks, study visits for each subject will be conducted at least 1 week apart.
On each study day, participants will be admitted to the MGH inpatient research unit, and will
have a light standardized breakfast after overnight fast and sleep. They will be asked not to
deviate from normal behaviors (e.g. if they drink caffeinated beverages or smoke cigarettes
in the mornings, they will be asked to do so on the study day so that no symptoms of caffeine
or nicotine withdrawal will appear during the scanning period. On each study day,
participants will complete tests of motor function/spatial navigation and fNIRS data
collection before study drug is administered. cognitive tasks will be repeated approximately
90 minutes after study drug administration, fNIRS will be repeated at approximately 120
minutes post dose, and the field sobriety test will be conducted immediately following the
second fNIRS assessment. Blood pressure, heart rate and ratings of subjective drug effects
will be monitored at approximately 15-minute intervals throughout the study. A third fNIRS
assessment will be conducted when intoxication has resolved, at approximately 220 minutes
post dose.

Participants: 150 healthy adults, aged 18-55, who use cannabis at least weekly, will be
enrolled. Participants must test positive for cannabis at the screening visit and have no
serious unstable medical illness to participate (See Participant Selection Section).

Phase 2B. Examine potential interaction following co-administration of THC with oral ethanol
exposure in healthy volunteers.

[Design adapted from recently published alcohol interaction studies (Modi et al., 2007;
Trevisan et al., 2008).]

Phase 2B is a randomized, double-blind, placebo-controlled, 2 by 2 crossover study of effect
of dronabinol, ethanol, and combined dronabinol and ethanol on brain activation and
connectivity as measured by fNIRS. Participants will be assigned randomly for order to
receive single doses of each of the following study medications on 4 separate study visits
conducted at least one week apart:

- Placebo THC + Ethanol

- THC + Placebo Ethanol

- THC + Ethanol

- Placebo THC + Placebo Ethanol

Participants will be admitted in the evening, have an overnight fast, be provided a standard
breakfast of approximately 400 calories (e.g. juice, cereal, bagel), approximately 45 minutes
before administering study medication.

Ethanol will be dosedby weight to obtain a breath alcohol concentration (BrAC) of
approximately 0.05, consistent with BrAC following 1-2 standard drinks, using established
methods of dosing based on previously published procedures (e.g. King 2005, Alcohol Clin Exp
Res, Vol 29, No 4, 2005: pp 547-552).

The required volume of 95% ethanol will be calculated from the nomogram published by Watson
(1989), and diluted to a final concentration of 20% by volume with diet soda. The nomogram,
based on total body water, estimated for each subject using standard equations based on
gender, age, height, and weight (Watson et al., 1980) will be used to prepare the ethanol for
dosing by study staff who are not otherwise involved in study procedures.

In an attempt to standardize the drinking time to 8 minutes, the total dose will be split
into 4 equal aliquots and placed in cups with lids. The subject will be given a cup every 2
minutes with instructions to sip the drink using a straw.

• Placebo Ethanol drink will be made as an equal volume of diet soda placed in identical cups
with lids and straws. 0.25 ml of 95% ethanol will be floated on the top of the soda in each
cup in an attempt to blind the participant and study staff who are involved with assessments
/ data acquisition.

The alcohol administration procedure will take place approximately 45 min before the start of
the second fNIRS session.

Breathalyzer measurements will be taken approximately every 5-10 minutes after the alcohol
administration procedure, until BrACs have returned to <0.01. Participants will be required
to stay in the laboratory until BrACs have returned to <0.01. A member of our study staff
will then (1) arrange cab service, and (2) escort the participant to the cab at the end of
the study visit. Alternatively, participants may arrange travel home with a family member or
friend. If they choose to do this, a study staff member will ensure that someone is there to
bring them home prior to departure.

Inclusion Criteria:

- Weekly or greater Marijuana users

- Aged 21-55

- Participants must test positive for cannabis and have no serious unstable medical
illness

Exclusion Criteria:

- Any unstable, serious medical illness, eg. unstable angina, or cardiovascular event in
the past 6 months eg. myocardial infarction or stroke.

- Diabetes, seizure disorder, cirrhosis, schizophrenia, bipolar disorder, clinically
significant cardiac conduction disorder, uncontrolled hypertension, tachycardia, renal
failure;

- History of syncope, HIV, Hepatitis C, migraines, head injury with prolonged
unconsciousness (> 24 hours);

- Allergy to sesame oil (contained in Marinol pills) or marinol capsules

- Current regular use of benzodiazepines or barbiturates, antihistamines, atropine,
scopolamine, other anticholinergic agents;

- Current pregnancy or lactation, or trying to become pregnant (confirmed by urine
pregnancy test)

- Currently seeking treatment, in treatment, or in recovery from an alcohol use
disorder.

- In the opinion of the investigator, not able to safely participate in this study.