Title of Invention

N-SUBSTITUTED CARBAMATE META-BIPHENYL ESTER AS NOVEL FATTY ACID AMIDE HYDROLASE INHIBITORS

Abstract There is disclosed an N-substituted carbamate meta-biphenyl ester of the formula: wherein: X is O Q is O; n is 1, 2, or 3; wherein each Ra member is independently selected from the group consisting of unsubstituted saturated alkyl, unsubstituted alkenyl, unsubstituted alkynyl, unsubstituted at least partially saturated cycloalkyl, unsubstituted ketoalkyl, unsubstituted hydroxyalkyl, unsubstituted aminoalkyl, -CH2-NR3R4, unsubstituted alkoxy, halo, unsubstituted haloalkyl, cyano, hydroxy, nitro, amino, -NR3R4, unsubstituted carboxamido, -CONR3R4, sulfonamido, -SO2NR3R4; -SR5, -SO2R6, -S(O)R6, -NR6C(O)R7, and -NR6SO2R7; wherein R3 and R4 are independently selected from the group consisting of H, unsubstituted saturated alkyl, unsubstituted alkenyl, unsubstituted alkynyl, unsubstituted at least partially saturated cycloalkyl, unsubstituted hydroxyalkyl and imino-methylamino and wherein optionally R3 and R4 may together with the N atom to which they are attached combine to form a 5-7 membered ring; and R5, R6 and R7 are independently selected from the group consisting of hydrogen, unsubstituted alkyl, and unsubstituted heteroalkyl; wherein R1 and R2 are independently selected from the group consisting of hydrogen, substituted or unsubstituted straight or branched alkyl, substituted or unsubstituted heteroalkyl, substituted or unsubstituted at least partially saturated cycloalkyl, and substituted or unsubstituted cycloheteroalkyl, with the proviso that R1 and R2 are not both hydrogen; and the pharmaceutically acceptable salts thereof.
Full Text N-SUBSTITUTED CARBAMATE META-BIPHENYL ESTER AS NOVEL
FATTY ACID AMIDE HYDROLASE INHIBITORS
STATEMENT AS TO RIGHTS TO INVENTIONS MADE UNDER
FEDERALLY SPONSORED RESEARCH OR DEVELOPMENT
This research was made, in part, with government support under Grant Nos.
DA12413, DA 12447, and DA 12653 awarded by the National Institutes of Health which
may have certain rights to the invention.
RELATED APPLICATIONS
This application claims benefit of U.S. Provisional Application No.
60/417,008 filed October 7,2002. This application is also related to U.S. Application No.
10/112,509 filed March 27,2003 and U.S. Patent Application No. 10/642,462 filed August
15,2003. The disclosures of each of which are incorporated in its entirety.
BACKGROUND OF THE INVENTION
[01] Anxiety is a pathological counterpart of fear and is often accompanied by
disturbances of mood, thinking, behavior, and physiology. Whereas fear is typically
triggered by a perception of a threat in the environment, anxiety disorders typically involve a
fearfulness which is either unprovoked by an environmental threat or highly disproportionate
to an environmental threat.
[02] Anxiety disorders are among the most common mental disorders and can greatly limit
the quality of life. In an anxiety disorder, an extreme or pathological anxiety is generally the
principal disturbance of mood or emotional tone. Such disorders include, but are not limited
to, panic disorder (with and without a history of agoraphobia), agoraphobia (with and without
a history of panic disorder), generalized anxiety disorder, specific phobia, social phobia,
obsessive-compulsive disorder, acute stress disorder, and post-traumatic stress disorder. In
addition, there are adjustment disorders with anxious features, anxiety disorders due to
general medical conditions, substance-induced anxiety disorders, and the residual category of
anxiety disorder not otherwise specified (See DSM-TV).
[03] Cognitive-behavioral therapies can be beneficial for many patients with anxiety
disorders (Chambless et al., 1998). Benzodiazepines, antidepressants, selective serotonin

reuptake inhibitors and the novel compound buspirone (Lydiard et al., 1996) have been used
with some success in the treatment of anxiety and anxiety disorders. Treatments combining
psychotherapy and pharmacotherapy are also beneficial (March et al, 1997; American
Psychiatric Association, 1998). One limitation of such psychotherapeutic treatments has
been their cost and the reluctance of patients to enter such treatment. Many of the
pharmaceutical treatments rely upon drugs, most particularly the important class of
benzodiazepines, which have some addiction or abuse potential. As the anti-anxiety
pharmacopeia is relatively bare, there is a need for additional therapeutic agents to treat
anxiety and anxiety disorders.
[04] Anxiety is one of the few mental disorders for which animal models are available.
Researchers can reproduce symptoms of human anxiety in test animals by manipulating
physical or psychosocial stressors. These animal models provide a means for screening
compounds for anti-anxiety activity. In light of increasing awareness of numerous
neurochemical alterations in anxiety disorders, many new classes of drugs are likely to be
developed through such screening.
[05] The psychoactive constituent of Cannabis, ∆9-tetrahydrocannabinol (∆9-THC),
produces in humans subjective emotional responses mediated by CB1 cannabinoid receptors,
suggesting that endogenous cannabinoids may contribute to the regulation of mood and
emotions. But the variable effects of ∆9-THC—which, depending on dosage, range from
relaxation and euphoria to anxiety and panic attacks—obscure the interpretation of these
results and limit the therapeutic potential of direct-acting cannabinoid agonists.
[06] Anandamide, the naturally occurring amide of arachidonic acid with ethanolamine,
meets all key criteria of an endogenous cannabinoid substance (Devane, WA et al. Science,
258,1946-1949 (1992)): it is released upon demand by stimulated neurons (Di Marzo, V. et
al., Nature, 372, 686-691 (1994); Giumida, A. et al., Nat. Neurosci., 2, 358-363 (1999)); it
activates cannabinoid receptors with high affinity (Devane, W.A. et al. Science, 258, 1946-
1949 (1992)) and it is rapidly eliminated through a two-step process consisting of carrier-
mediated transport followed by intracellular hydrolysis (Di Marzo, V. et al., Nature, 372,
686-691 (1994); Beltramo, M. et al., FEBS Lett, 403, 263-267 (1997)). Anandamide
hydrolysis is catalyzed by theyenzyme fatty acid_amide hydrolass(FAAH), a membrane-
bound serine hydrolase (Cravatt, B.F. et al., Nature, 384, 83-87 (1996); Patricelli, M.P. et al.,
Biochemistry, 38, 9804-9812 (1999)) (WO 98/20119) (U.S. Patent No. 6,271,015) that also
cleaves other bioactive fatty ethanolamides, such as oleoylethanolamide (cis-9-

octadecenamide)) (Rodriguez de Fonseca, F. et al. Nature, 414, 209-212 (2001)) and
palmitoylethanolamide (Calignano, A. et al., Nature, 394, 277-281 (1998)). Mutant mice
lacking the gene encoding for FAAH cannot metabolize anandamide (Cravatt, B.F. et al.,
Proc. Natl. Acad. Sci. U. S. A., 98, 9371-9376 (2001)) and, though fertile and generally
normal, show signs of enhanced anandamide activity at cannabinoid receptors, such as
reduced pain sensation (Cravatt, B.F. et al., Proc. Natl. Acad. Sci. U. S. A., 98, 9371-9376
(2001)). This suggests the possibility that drugs targeting FAAH may heighten the tonic
actions of anandamide, while possibly avoiding the multiple, often unwanted effects
produced by ∆9-THC and other direct-acting cannabinoid agonists (Hall, W., et al., Lancet,
352,1611-1616 (1998 ); Chaperon, F., et al., Crit. Rev. Neurobiol, 13,243-281 (1999)).
[07] Most current inhibitors of the FAAH enzyme lack the target selectivity and biological
availability needed for in vivo studies (Koutek, B. et al., J. Biol. Chern., 269, 22937-22940
(1994); De Petrocellis, L. et al., Biochem. Biophys. Res. Commun., 231, 82-88 (1997);
Deutsch, D.G. et al., Biochem. Biophys. Res. Commun., 231,217-221 (1997); Beltramo, M. et
al., FEBS Lett., 403:263-267 (1997)), while newer compounds, though promising, have not
yet been characterized (Boger, D.L. et al. Proc. Natl. Acad. Sci. U. S. A., 97, 5044-5049
(2000)). Thus, the therapeutic potential of FAAH inhibition with respect to the endogenous
cannabinoid system activity remains essentially unexplored.
[08] The present invention expands the pharmacopeia for the inhibition of FAAH and the
treatment of anxiety and other conditions by providing a new class of FAAH inhibitors and
new methods for treating anxiety and anxiety disorders, or conditions by administering FAAH
inhibitors.
BRIEF SUMMARY OF THE INVENTION
[09] The invention provides novel compounds for inhibiting Fatty Acid Amide Hydrolase
(FAAH) and methods of treating anxiety or pain, and other neurological, sleep, or
psychological disorders, for inducing sleep, for treating glaucoma, and controlling appetite or
treating appetive disorders by administering FAAH inhibitors to a subject. In one of its
aspects the invention discloses the use of FAAH inhibitors as useful in treating anxiety and
depression. In another of its aspects the invention provides FAAH inhibitory compounds of
the following Formula:


in which X is CH2, NH, O, or S; Q is O or S; Z is 0 or N, with the proviso that when Z is 0,
one of R1 and R2 is absent; and R is an aromatic or alkyl or lipophilic moiety selected from
the group consisting of substituted or unsubstituted aryl; substituted or unsubstituted
biphenylyl, substituted or unsubstituted naphthyl, and substituted or unsubstituted phenyl;
substituted or unsubstituted terphenylyl; substituted or unsubstituted cycloalkyl, substituted
or unsubstituted heteroaryl, substituted or unsubstituted alkyl; substituted or unsubstituted
heteroalkyl, and

wherein p is a number from from 0 to 3; m is a number from 0 to 4, and n is a number from 0
to 5, Z1 and Z2 are same or different and are independently a divalent radical selected from
the group consisting of -O-, -S-, -N(R5)-, -C(R6)-, C(R7)-,-C(R6)-=N- and -N=C(R6)- wherein
R5 is selected from H, alkyl, heteroalkyl, alkenyl, alkynyl, cycloalkyl, aryl, acyl and aroyl; R6
and R7 are H or R6 and R7 optionally may combine to form a saturated or unsaturated
carbocyclic or heterocyclic ring, optionally substituted with one or more Ra and Rb groups; Y
is a bond, or a divalent radical selected from the group consisting of -O-, -S-, -N(R5)-, C1-C4
alkylene, (Z)- or (E)-ethylene, and cycloalkyl with 3 to 6 carbon atoms; Ra and Rb are
independently selected from the group consisting of H, alkyl, heteroalkyl, alkenyl, alkynyl,
cycloalkyl, aryl, substituted aryl, arylalkyl, substituted arylalkyl, ketoalkyl, hydroxyalkyl,
aminoalkyl, -CH2-NR3R4. alkoxy, aryloxy, arylalkyloxy, halo, haloalkyl, cyano, hydroxy,
nitro, amino, -NR3R4, -SR5, carboxamido, -CONR3R4, -O-carboxamido, -O-CO-NR3R4,
sulfonamido, and -SO2NR3R4. wherein R3 and R4 are selected from H, alkyl, alkenyl, alkynyl,
cycloalkyl, hydroxyalkyl and imino-methylamino and wherein optionally R3 and R4 together
with the N atom to which they are attached to form a 5-7 membered cyclic ring.

[10J In addition, R1 and R2 are independently selected from the group consisting of H,
substituted or unsubstituted alkyl, substituted or unsubstituted heteroalkyl, substituted or
unsubstituted cycloalkyl, and substituted or unsubstituted cycloheteroalkyl, and substituted or
unsubstituted phenyl, and substituted or unsubstituted aryl or heteroaryl, and wherein
optionally, when X is N, if taken together with the N atom to which they are attached, R1 and
R2, form a substituted or unsubstituted N-heterocycle or substituted or unsubstituted
heteroaryl with the atom to which they are each attached.
[11] In one aspect, the invention provides compounds in which R and not -NR1R2 provides
the majority the hydrophobic bulkiness needed to occupy a hydrophobic channel in FAAH as
set forth in Example 18. In particular embodiments, for instance, when R1 and/or R2 is
substituted aryl or heteroaryl or comprises a bulky aromatic ring, R represents a substituent
which has a greater mass or weight than -NR1R2. Other embodiments include, but are not
limited to, compounds in which R has a greater mass or molecular weight than -NR1R2 when
R1 and R2 taken alone or together do not comprise any aryl, heteroaryl, or aromatic rings. In
some embodiments, a compound with a FAAH IC50 of less than 1 micromolar can have a
surprisingly small -NR1R2 moiety. For instance, such an inhibitor can have a molecular
weight of less than 200 or 100 Daltons or be no larger in bulkiness than a C7 cycloalkyl or C6
or C10 alkyl group when R is as described above. In another aspect, the compounds of the
invention include compounds which interact more with the hydrophobic channel of FAAH
instead of the catalytic site of the enzyme to support their binding. Exemplary such
compounds have a hydrophilic moiety distal to the carbamate end of the molecule and
separated mere from by a lipophilic moiety occupying the hydrophilic channel of the
enzyme.
[12] In one embodiment, the inhibitor of Formula I has an IC50 of less than 1 µM. In one
embodiment the inhibitor Formula I has an IC50 of less than 0.01 µM. In another
embodiment, the inhibitor Formula I has an IC50 of from about 1 uM to about 0.01 uM, or
from about 0.01 to about 0.001 µM.
[13] In one embodiment the FAAH inhibitor is a compound of Formula II:


in which R1, R2 and R are defined as recited above.
[14] In one embodiment of a compound of Formula II, R1 is H and R2 is cyclohexyl. In
another embodiment, R is substituted or unsubstituted biphenylyl. In a further embodiment,
R1 is H, R2 is cyclohexyl and R is substituted or unsubstituted biphenylyl or substituted or
unsubstituted phenyl.
[15] In one embodiment, such an inhibitor or compound according to Formula I or II has a
FAAH IC50 of less than 1 µM. In another embodiment the inhibitors have a FAAH IC50 of
less than 0.01 µM. In another embodiment, the inhibitors have a FAAH IC50 of from about
1 µM to about 0.01 µM, or from about 0.01 to about 0.001 µM.
[16] In yet another embodiment of a compound in which X is O, Q is O; and Z is N; R1 is
H and R2 is C1 to C8 alkyl. In a further embodiment, R is substituted or unsubstituted
biphenylyl, terphenylyl, or stilbyl.
[17] In another aspect, the invention provides FAAH inhibitors and compounds of the
following general formula:

In Formula m, R1 and R2 are independently selected from the group consisting of H,
unsubstituted or substituted homoalkyl, substituted or unsubstituted heteroalkyl, substituted
or unsubstituted cycloalkyl, substituted or unsubstituted heterocycloalkyl, and optionally R1
and R2, may be taken together to form a substituted or unsubstituted heterocycle with N; U
is a hydrophobic spacer, wherein the spacer comprises substituted or unsubstituted aryl; and
V is a hydrophilic moiety having at least one functional group capable of forming a hydrogen
bond. In addition, the hydrophobic spacer is at least 9 angstroms in length; and the
hydrophilic moiety is attached to the spacer at a point from 8 to 12 angstroms distant from a
point at which the hydrophobic spacer is covalently attached to the rest of the inhibitor.

[18] In a further embodiment, the inhibitor or compound of Formula III has a
hydrophobic spacer comprising a first and a second aromatic ring, wherein the first and
second aromatic rings are covalently attached by a linker selected from the group consisting
of a bond, a single heteroatom, and substituted or unsubstituted C1 to C4 alkylene.
[19] In another embodiment, V is selected from the group consisting of ketoalkyl,
hydroxyalkyl, aminoalkyl, -CH2-NR3R4, alkoxy, aryloxy, halo, haloalkyl, cyano, hydroxy,
nitro, amino, -NR3R4, carboxamido, -CONR3R4, -0-carboxamido, -O-CO-NR3R4,
sulfonamido, and -SO2NR3R4; in which R3 and R4 are selected from H, alkyl, heteroalkyl,
alkenyl, alkynyl, cycloalkyl, hydroxyalkyl and imino-methylamino or R3 and R4 may
optionally combine with the N atom to which they are attached to form a 5-7 membered
cyclic ring.
[20] In another embodiment, U and V are each set forth as described above and the
hydrophilic moiety is attached to the spacer at a point about 9 to 11 angstroms from a point
at which the spacer is covalently attached to the rest of the inhibitor. In a further
embodiment, the hydrophilic moiety is attached to the spacer at a point about 10 angstroms
from a point at which the spacer is covalently attached to the rest of the inhibitor.
[21] In one embodiment, U and V taken together have a greater mass than the remainder of
the molecule or than the -NR1R2 moiety.
[22] In yet another embodiment, the hydrophobic spacer occupies the central hydrophobic
channel of the FAAH enzyme as set forth in Example 18 and is of sufficient length to allow
an appropriately positioned V group to engage in hydrogen bonding at a hydrophilic site at
the indicated hydrophilic wall of the channel and distal to the carbamate moiety binding site.
[23] In another embodiment, the compound of Formula I or Formula II or Formula HI is a
FAAH inhibitor which is selective for FAAH as compared to the neurotoxic esterase (NTE)
or acetylcholinesterase (ACHE). In one embodiment, the compound has an FAAH IC50
which is one-tenth to one-hundredth that for the NTE or ACHE IC50.
[24] In one embodiment, such an inhibitor according to Formula I, H, or HI has an IC50 of
less than 1 µM. In still another embodiment, the inhibitor according to Formula I, II, or III
has an IC50 of less than 0.01 µM. In yet another embodiment, the inhibitor according to
Formula I, n, or in has an IC50 of from about 1 µM to about 0.01 µM, or from about 0.01 to
about 0.001 µM.
[25] In one embodiment, the inhibitor of Formula I, II or III is a selective inhibitor of
FAAH over any one of electric eel ACHE, rat brain monoglyceride lipase, or horse plasma

butyryl cholinesterase. In further embodiments, the selective inhibitor has an IC50 for
inhibition of FAAH mat is no more than one-fifth, one-tenth, or one-twentieth its IC50 for
inhibition of electric eel ACHE, rat brain monoglyceride lipase, or horse plasma butyryl
cholinesterase.
[26] In one embodiment, the inhibitor of Formula I, II or III is a selective inhibitor of
FAAH which does not substantially directly interact with the CB1 or the CB2 cannabinoid
receptor at the IC50 concentrations for inhibition of FAAH. In further embodiments, the
binding IC50 for inhibiting the binding of an agonist of the CB1 or CB2 cannabinoid receptor
is at least 10 or 20 or 100 times greater than the IC50 for inhibiting FAAH.
[27] In one embodiment, a compound of the Formula I or II or III is contacted with FAAH
to inhibit the FAAH enzyme. In a further embodiment, the enzyme is contacted in vivo. In a
second further embodiment, the enzyme is contacted in vitro with a compound of Formula I,
II, or III.
[28] In one aspect, the invention provides novel carbamate inhibitors of FAAH and then-
use in inhibiting FAAH. In one embodiment, a compound of the Formula I, n, or in is
contacted with a FAAH to inhibit the enzyme. In a further embodiment, the enzyme is
contacted in vivo. In another embodiment, the enzyme is contacted in vitro with a compound
of Formula I, n, or III.
[29] In another aspect, the invention provides a method of treating anxiety, an anxiety
disorder, or a psychological disorder associated with anxiety by administering an inhibitor of
a FAAH inhibitor to a subject having one or more of such conditions. In another
embodiment, such a subject is not otherwise in need of treatment with a FAAH inhibitor. In
another embodiment, the subject with one of such conditions is not in need of a sleep-
inducing agent or pain relief. In another embodiment, the FAAH inhibitor is selective for
FAAH as compared to the neurotoxic esterase (NTE) or acetylcholinesterase (ACHE).
[30] In one embodiment, a FAAH inhibitor is administered to a subject to treat anxiety or
an anxiety disorder. In a further embodiment, a compound according to Formula I or
Formula II or Formula III is administered to treat anxiety or an anxiety disorder. In a further
embodiment, the compound to be administered to treat anxiety or an anxiety disorder is
UCM532 or UCM597. In a further embodiment, the subject is not otherwise in need of
treatment with a FAAH inhibitor. In another embcnliment, the subject to be treated is not in
need of a sleep-inducing agent or pain relief, including but not limited to neuropathic pain. In
another embodiment, the FAAH inhibitor is formulated with an antianxiety compound which

is not a FAAH inhibitor and/or administered as part of a combination therapy with such an
antianxiety compound.
[31] In one embodiment, a FAAH inhibitor is administered to a subject to treat depression
or a depressive disorder. In a further embodiment, a compound according to Formula I, n,
or HI is administered to treat such depression or disorder. In a further embodiment, the
compound to be administered to treat such depression or disorder is UCM532 or UCM597. In
another embodiment, such a subject is not otherwise in need of treatment with a FAAH
inhibitor. In another embodiment, the subject is not in need of sleep-inducing agent or pain
relief. In another embodiment, the FAAH inhibitor is formulated with a second
antidepressant which is not a FAAH inhibitor or administered as part of a combination
therapy with such an antidepresssant.
[32] In another aspect, the invention provides a method of treating epilepsy by
administering a FAAH inhibitor to a subject with epilepsy. In one embodiment, a compound
according to Formula I, II, or HI is administered to treat epilepsy. In another embodiment,
the subject is also treated with an additional anti-epilepsy compound which is not an FAAH
inhibitor. In another embodiment, the compound according to Formula I is UCM532 or
UCM597. In another embodiment, the FAAH inhibitor is administered or formulated with a
second compound which is not a FAAH inhibitor.
[33] In one aspect of the instant invention, methods are provided for reducing appetite,
body fat or body weight, or for treating or preventing obesity or overweight, or for reducing
food intake, or treating an appetency disorder in a mammal by administering to the mammal a
FAAH inhibitor to reduce an appetite or the consumption of an appetizing substance such as
food. In some embodiments, the compound is a compound of Formula I, n, or HI.
[34] In another aspect the invention provides methods of identifying anxiolytic FAAH
inhibitors by administering the inhibitor to a test subject; exposing the subject to an anxiety-
causing stimulus; and measuring the degree of anxiety in the exposed test subject. In one
embodiment, the invention provides a method of determining whether a compound is an
anxiolytic fatty acid amide hydrolase inhibitor by contacting the compound with a fatty acid
amide hydrolase enzyme under enzymatic assay conditions and selecting the compound if it
is a fatty acid amide hydrolase inhibitor, and then determining if the selected compound is
an anxiolytic fatty acid amide hydrolase enzyme inhibitor by administering the inhibitor to a
test subject; exposing the subject to an anxiety-causing stimulus; and measuring the degree
of anxiety in the exposed test subject In one embodiment, such an inhibitor is a compound
of Formula I, n, or III.

[35] In another aspect the invention provides methods for identifying an anti-depressant or
an anti-epilepsy FAAH inhibitor by administering the inhibitor to a test subject in an animal
model for depression or epilepsy, respectively, and measuring the performance of the test
subject according to the test. In one embodiment, the invention provides a method of
determining whether a compound is an anti-depressant or an anti-epileptic fatty acid amide
hydrolase inhibitor by contacting the compound with a fatty acid amide hydrolase enzyme
under enzymatic assay conditions and selecting the compound if it is a fatty acid amide
hydrolase inhibitor; and then detennining if the selected compound is an anti-depressant or
anti-epileptic fatty acid amide hydrolase enzyme inhibitor by administering the inhibitor to a
test subject in an animal model for depression or epilepsy, respectively. In one embodiment,
such an inhibitor is a compound of Formula I, II, or III.
[36] In one embodiment, a FAAH inhibitor is administered to a subject to treat
schizophrenia or paranoia or a related disorder or a disorder of dopamine transmission. In a
further embodiment, a compound according to Formula I, II, or III is administered to treat
such diseases or conditions. In a further embodiment, the compound to be administered is
UCM532 or UCM597. In one embodiment, the FAAH inhibitor is formulated or
administered with or given as part of a combination therapy with a second anti-psychotic
agent which is not a FAAH inhibitor.
[37] In still another aspect, the invention provides a pharmaceutical composition
comprising a compound of Formula I, II, or III and a pharmaceutically acceptable excipient.
In another aspect, the invention provides methods of treating depression, anxiety, insomnia,
pain, schizophrenia, epilepsy, glaucoma, or an appetite disorder by administering such a
composition to a subject. In one embodiment, the invention provides pharmaceutical doses in
unit dosage format comprising a therapeutically effective amount of the FAAH inhibitor. In
some embodiments, the therapeutically effect is in an amount sufficient to treat one of the
above psychological conditions or disorders. In one embodiment, the therapeutically effect is
in an amount sufficient to treat anxiety or an anxiety disorder in a subject. In other
embodiments, the treated subject is a human with acute anxiety, chronic anxiety, or an
anxiety disorder. In other embodiments, the unit dosage of the FAAH inhibitor is in an
amount sufficient to treat a human with depression or a depressive disorder.
[38] In still other aspects the invention provides a method of modulating endogenous fatty
acid amide levels in a subject by administering a compound of Formula I, II, or III to a
subject. In one such embodiment, the modulating reduces anxiety in said subject. In another
embodiment, the modulating reduces sensitivity to pain in the subject, In another

embodiment, the modulating does not induce catalepsy. In another embodiment, the
modulating does not induce hyperphagia or affect appetite.
[39] In another aspect, the invention also provides methods for increasing the levels of
endogenous anandamide, endogenous oleoylethanolamide, and other endogenous fatty acid
amides in a subject by administering a compound of Formula I, II, or in. The invention also
provides methods for increasing the levels (e.g., blood, plasma, brain or other tissue
concentrations) or biological activity (e.g., therapeutic activity, FAAH inhibitory activity) of
administered or exogenous anandamide, oleoylethanolamide, and fatty acid amides in a
subject by administering a compound of Formula I, II, or III.
BRIEF DESCRIPTION OF THE ACCOMPANYING DRIVINGS
[40] Fig. 1. The FAAH inhibitors UCM532 and UCM597 block [3H]anandamide
degradation in intact brain neurons. a, Concentration-dependent inhibition of
[3H]anandamide hydrolysis by UCM597 (open squares) and UCM532 (closed circles) in
primary cultures of rat cortical neurons, b, Unlike UCM532 (FB, 3 µM) or UCM597 (CF, 10
nM), the UCM532 analog 7 (10 µM) has no effect on [3H]anandamide degradation, c,
UCM532 (FB, 3 (µM) and UCM597 (CF, 10 nM) promote accumulation of non-metabolized
[3H]anandamide in neurons, whereas the anandamide transport inhibitor AM404 (AM, 10
µM) reduces it. d, Release of non-metabolized [3H] anandamide from UCM597-treated (10
nM) neurons during a 15-min incubation in the absence or presence of AM404 (AM, 10 µM).
e, Time course of [3H] anandamide release from UCM597-treated (10 nM) neurons. One
asterisk, P Tukey's post-hoc test (n - 4-8).
[41] Fig. 2. In vivo inhibition of FAAH activity by UCM532 and UCM597. a, Dose-
dependent inhibition of brain FAAH activity by UCM532 (closed circles) and UCM597
(open squares), but not by the inactive analog 7 (closed diamond), after systemic (i.p.)
administration in the rat. b, Time-course of the inhibition of brain FAAH activity after a
single injection of UCM597 (0.3 mg kg-1, i.p.). Brain levels of anandamide (c) and 2-AG (d)
2 h after injections of vehicle (V) or UCM597 (CF, 0.3 mg kg-1, i.p). One asterisk, P two asterisks, P anandamide-induced hypothermia by UCM597. Effects of UCM597 (0.3 mg kg-1; open
squares), anandamide (5 mg kg-1; closed circles), anandamide (15 mg kg-1; open circles), and

anandamide (5 mg kg-1) plus UCM597 (0.3 mg kg-1, 30 min before anandamide) (closed
diamonds). One asterisk (P differences between anandamide and anandamide/UCM597; {-test with Bonferroni's
correction (« = 6-7).
[42] Fig- 3. Antinociceptive actions of UCM597. a, Effects of UCM597 (CF, 0.5 mg kg-1,
i.p.) on response latencies in the mouse hot-plate test, in the absence or presence of the CB1
antagonist rimonabant (Ri, 0.2 mg kg-1, i.v.). b, Effects of UCM597 (0.5 mg kg-1) on the
early phase (open bars) and late phase (closed bars) of formalin-evoked pain, in the absence
or presence of rimonabant. UCM597 and rimonabant were injected 60 min and 40 min
before tests, respectively. One asterisk, P 12).
[43] Fig. 4 Anxiolytic-like actions of UCM532 and UCM597. Dose-dependent effects of
(a) UCM532 (FB, 5 and 10 mg kg-1, i.p.) or (b) UCM597 (CF, 0.05 and 0.1 mg kg-1, i.p.) on
the percent time spent by adult rats in the open quadrants of a zero maze (percent time open).
c, Effect of the CB1 antagonist rimonabant (2 mg kg-1, i.p.) on the change in percent time
open produced by UCM532 (5 mg kg"1), d, Effects of UCM532 (5 and 10 mg kg-1) on
ambulation time in adult rats (20 min session), e, Effects of UCM532 (5 and 10 mg kg-1) on
isolation-induced ultrasonic vocalizations in rat pups. One asterisk, P [44] Fig. 5 sets forth the chemical structures of anandamide (1) and URB524 (2).
[45] Fig. 6 provides a plot of FAAH inhibition potency (pIC50) vs lipophilicity (n) for
URB524 and its meta-substituted derivatives.
[46] Fig. 7 is a drawing of the 3-D surface of the catalytic channel inside FAAH, colored
by residue lipophilicity. Hydrophilic regions are colored in blue, lipophilic ones in brown.
[47] Fig 8 represents a superposition of the biphenyl fragment of URB524 to the
arachidonyl chain of MAPF co-crystallyzed with FAAH.
[48] Fig. 9 illustrates docking of URB597 into the FAAH binding site. The hydrogen
bonds of the carbamoyl group with the enzyme are evidenced in yellow.
[49] Table 1. Values reported are the concentrations required to inhibit FAAH activity by
50% (ICso, nM), and are expressed as the mean ± SEM of at least three independent
experiments. They were calculated from concentration-response curves, by using non-linear
regression analysis as implemented in the Prism 2.0 software package.

[50J Table 2. Values indicate the maximal concentrations of FAAH inhibitor tested on
each target (in µM) and their corresponding selectivity index (SI). The SI is the ratio of
maximal inhibitor concentration tested/IC50 for FAAH (from Table 1).
[51] Table 3. FAAH and acetylcholinesterase IC50 values are reported for 18 compounds,
including carbaryl and physostigmine.
152] Table 4. FAAH IC50 values are reported for over 50 compounds of Formula I.
[53] Table 5. FAAH IC50 values are reported for over 20 meta biphenyl compounds.
[54] Table 6. Observed and calculated pIC50 values for FAAH inhibition of the meta
substituted derivatives included in the QSAR analysis.
DETAILED DESCRIPTION OF THE INVENTION
[55] In one aspect, the invention provides novel inhibitors of fatty acid amide hydrolase
(FAAH), the enzyme responsible for the intracellular degradation of the endogenous
cannabinoid anandamide. The inventors have surprisingly discovered compounds that inhibit
FAAH in vivo with a low IC50. Exemplary compounds according to the invention can be
potent, selective, systemically active inhibitors of FAAH. FAAH inhibitors can be useful for
a variety of purposes such as the induction of sleep, treatment of insomnia, and the alleviation
of pain. The invention also provides a means of treating anxiety by administering FAAH
inhibitors. Like clinically used anti-anxiety drugs, such inhibitors surprisingly exhibit
benzodiazepine-like properties in the elevated zero-maze test and suppress isolation-induced
vocalizations in rats. Furthermore, they reduce nocifensive (pain-avoiding) behaviors in
models of acute pain. These effects have been accompanied by augmented brain levels of
anandamide, but not of the other endogenous cannabinoid 2-aracbidonoylglycerol, and are
prevented or antagonized by CB1 cannabinoid receptor blockade. The results show that
anandamide participates in the modulation of emotional states, and point to FAAH inhibition
as an innovative mechanistic approach to anti-anxiety therapy.
[56] FAAH inhibitors can be also useful in the treatment of a variety of other neurological
psychological disorders and conditions, including but not limited to pain, depression,
attention deficit hyperactivity disorders, jet lag, insomnia, schizophrenia, pain, muscle
spasticity, epilepsy, and seizure disorders as well as glaucoma.
[57] The invention also provides methods for increasing the levels of endogenous
anandamide, endogenous oleoylethanolamide, and other endogenous fatty acid amides in a
subject by administering a compound of Formula I or Formula n. The invention also

provides methods for increasing the levels and biological activity of administered
anandamide, oleoylethanolamide, and fatty acid amides in a subject by also administering
(e.g., prior administration, contemporaneous administration, co-administration) to the subject
a FAAH inhibitor of Formula I or Formula n. Thus, the FAAH inhibitors of Formula I and II
can be useful in potentiating the biological activity of anandamide or oleoylethanolamide.
[58] The invention provides a new class of agents that prevent anandamide or
oleoylethanolamide inactivation by targeting the intracellular enzymatic activity of FAAH.
[59] The invention provides further a novel class of inhibitors of FAAH activity, which
enhance endogenous anandamide signaling. The behavioral profile of these agents—
characterized by anxiolysis and mild analgesia—reveal a key role for anandamide in the
regulation of emotional states and provide a new mechanistic approach to anti-anxiety
therapy in particular.
[60] The invention also provides a means of inhibiting FAAH by contacting the enzyme in
vitro or in vivo with an inhibitor or compound according to the invention (e.g., compounds of
Formula I, Ia-c, I, IIa-b, and III). The enzyme is preferably mammalian (e.g., rat, human,
mouse, dog, cat, domesticated species of mammals).
Definitions
[61] It is noted here that, as used in this specification, the singular forms "a," "an," and
"the" include plural reference unless the context clearly dictates otherwise.
[62] Unless otherwise defined, all technical and scientific terms used herein have the same
meaning as commonly understood by one of ordinary skill in the art to which this invention
belongs. Each publication, patent application, patent, and other reference cited herein is
incorporated by reference in its entirety to the extent that it is not inconsistent with the
present disclosure.
[63] In the present description and in the claims, "appetency disorders" are understood as
meaning disorders associated with a substance and especially abuse of a substance and/or
dependency on a substance, disorders of food behaviors, especially those liable to cause
excess weight, irrespective of its origin, for example: bulimia, appetency for sugars, non-
insulin-dependent diabetes. Appetizing substances are therefore understood as meaning
substances to be taken into the body and for which an appetite or craving for such
consumption is present by any route of entry or self-administration. Appetizing substances
includes, foods, and their appetizing ingredients such as sugars, carbohydrates, or fats, as
well as drinking alcohol or drugs of abuse or excess consumption. An "appetite' may be

directed toward such substances as foods, sugars, carbohydrates, fats, as well as ethanol or
drugs of abuse or addiction or excess consumption (e.g., tobacco, CNS depressants, CNS
stimulants).
[64] Appetite refers to the desire to consume an appetizing substance or the behavior of
consuming appetizing substances. An appetizing substance may be a food or sugar or other
substance. In one embodiment, the appetizing substance is a food. In some embodiments,
the appetizing substance is a drug of abuse such as ethanol, nicotine, cocaine, an opioid, a
CNS stimulant or a CNS depressant.
[65] Anxiety is a state of fearfulness which is unprovoked by an environmental threat or
highly disproportionate to an environmental threat. Anxiety may be acute and short term
lasting hours to days; or chronic and lasting from many days to weeks or longer.
[66] The term clinical anxiety refers to any form of anxiety for which treatment is
necessary or indicated in order to alleviate it. Such clinical anxiety may be persistent or
recurrent and typically severe.
[67] Anxiety disorders include, but are not limited to, any of the anxiety disorders as
provided in the Diagnostic and Statistical Manual of Mental Disorders, Fourth Edition.
(Copyright 1994 American Psychiatric Association) which is hereby incorporated by
reference. Such disorders include, but are not limited to, panic disorder, agoraphobia,
generalized anxiety disorder, specific phobia, social phobia, obsessive-compulsive disorder,
acute stress disorder, and post-traumatic stress disorder, and adjustment disorders with
anxious features, anxiety disorders due to general medical conditions, substance-induced
anxiety disorders, and the residual category of anxiety disorder not otherwise specified.
[68] Depressive disorders and conditions include, but are not limited to, any of the
depressive disorders and conditions as provided in the Diagnostic and Statistical Manual of
Mental Disorders, Fourth Edition (Copyright 1994 American Psychiatric Association). These
disorders include major depressive disorder (unipolar depression), dysthymic disorder
(chronic, mild depression), and bipolar disorder (manic-depression). Clinical depression
refers to any form of depression that requires some form of treatment in order to alleviate it.
Such clinical depression may persist for months and last for most of every day and seriously
impairs the quality of life.
[69] A "major depressive episode" is defined as at least two weeks of depressed mood or
loss of interest, which may be accompanied by other symptoms of depression. The symptoms
must persist for most of the day (i.e. for at least two thirds of the patients' waking hours),
nearly every day (i.e. for at least ten out of fourteen days) for at least two consecutive weeks.

A "depressed mood" is often described by the patient as feeling sad, hopeless, helpless or
worthless. The patient may also appear sad to an observer, for example, through facial
expression, posture, voice and tearfulness. In children and adolescents, the mood may be
irritable. A "loss of interest" is often described by the patient as feeling less interested in
hobbies or not feeling any enjoyment in activities that were previously considered to be
pleasurable.
[70] A major depressive episode may be accompanied by other symptoms of depression
including significant weight loss when not dieting or weight gain (e.g. a change of more than
5% body weight in one month), or decrease or increase in appetite; insomnia or hypersomnia;
psychomotor agitation or retardation; fatigue or loss of energy; feelings of worthlessness or
excessive or inappropriate guilt; diminished ability to think or concentrate; or indecisiveness;
and recurrent thoughts of death, recurrent suicidal ideation with or without a specific plan, or
a suicide attempt
[71] The term "composition", as in pharmaceutical composition, is intended to encompass
a product comprising the active ingredient(s), and the inert ingredient(s) that make up the
carrier, as well as any product which results, directly or indirectly, from combination,
complexation or aggregation of any two or more of the ingredients, or from dissociation of
one or more of the ingredients, or from other types of reactions or interactions of one or more
of the ingredients. Accordingly, the pharmaceutical compositions of the present invention
encompass any composition made by admixing a compound of the present invention and a
pharmaceutically acceptable carrier. The term "pharmaceutical composition" indicates a
composition suitable for pharmaceutical use in a subject, including an animal or human. A
pharmaceutical composition generally comprises an effective amount of an active agent and a
pharmaceutically acceptable carrier.
[72] The term "modulate" means to induce any change including increasing or decreasing.
(e.g., a modulator of fatty acid oxidation increases or decreases the rate of fatty oxidation.)
[73] The term "pharmaceutically acceptable carrier" encompasses any of the standard
pharmaceutical carriers, buffers and excipients, including phosphate-buffered saline solution,
water, and emulsions (such as an oil/water or water/oil emulsion), and various types of
wetting agents and/or adjuvants. Suitable pharmaceutical carriers and their formulations are
described in Remington's Pharmaceutical Sciences (Mack Publishing Co., Easton, 19th ed.
1995). Preferred pharmaceutical carriers depend upon the intended mode of administration
of the active agent. Typical modes of administration are described below.

(74] The term "effective amount" means a dosage sufficient to produce a desired result
with respect to the indicated disorder, condition, or mental state. The desired result may
comprise a subjective or objective improvement in the recipient of the dosage. With respect
to anxiety, the improvement may be decreased sign or symptom of anxiety.
[75] The terms "treatment", "therapy" and the like include, but are not limited to, methods
and manipulations to produce beneficial changes in a recipient's status. The changes can be
either subjective or objective and can relate to features such as symptoms or signs of the
disease, disorder or condition being treated. For example, if the patient notes decreased
fearfulness, anxiety or worry, then successful treatment of anxiety or an anxiety disorder has
occurred. For example, if a decrease in the frequency or severity of seizures is noted, then a
beneficial treatment of epilepsy has occurred. For example, if depressive ideation is reduced,
a beneficial change in depression or a depressive disorder has been achieved. Similarly, if
the clinician notes objective changes, such as decreases in tremulousness or agitation, then
treatment for anxiety has also been beneficial or successful. Preventing the deterioration of a
recipient's status is also included by the term. Therapeutic benefit includes any of a number
of subjective or objective factors indicating a response of the condition being treated as
discussed herein.
[76] "Drug", "pharmacological agent", "pharmaceutical agent", "active agent", and
"agent" are used interchangeably and are intended to have their broadest interpretation as to
any therapeutically active substance which is delivered to a living organism to produce a
desired, usually beneficial effect.
[77] "Pharmaceutically-acceptable" or "therapeutically-acceptable" refers to a substance
which does not interfere with the effectiveness or the biological activity of the active
ingredients and which is not toxic to the hosts, which may be either humans or animals, to
which it is administered.
[78] "Therapeutically-effective amount" refers to the amount of an active agent sufficient
to induce a desired biological or clinical result. That result may be alleviation of the signs,
symptoms, or causes of a disease, or any other desired alteration of a biological system. The
term "therapeutically effective amount" is used herein to denote any amount of the
formulation which causes a substantial improvement in a disease, disorder or condition when
administered to a subject over a period of time. The amount will vary with the condition
being treated, the stage of advancement of the condition, and the type and concentration of
formulation applied. Appropriate amounts in any given instance will be readily apparent to
those skilled in the art or capable of determination by routine experimentation.

[79] A "prophylactic treatment" is a treatment administered to a subject who does not
exhibit signs of a neurological or psychological disorder or condition or exhibits only early or
slight signs of such a disorder or condition, wherein treatment is administered for the purpose
of decreasing the risk of developing a pathology or worsening of disorder or condition. The
compounds of the invention may be given as a prophylactic treatment to prevent undesirable
or unwanted anxiety or panic attacks, or to reduce the level of anxiety should worsening
occur.
[80] The term "subject" as used herein includes any animal, including, but not limited to,
mammals (e.g., rat, mouse, cat, dog) including humans to which a treatment is to be given.
[81] Schizophrenia and related disorders include, but are not limited to the following
types: Catatonic Type; Disorganized Type; Paranoid Type; Residual Type; Undifferentiated
Type; and Schizophreniform Disorder as provided in the Diagnostic and Statistical Manual of
Mental Disorders, Fourth Edition. TEXT REVISION Copyright 2000 American Psychiatric
Association which is hereby incorporated be reference.
[82] "FAAH" denotes a mammalian Fatty Acid Amide Hydrolase and includes, but is not
limited to, the human, rat, mouse forms of the enzyme. U.S. Patent No. 6,271,015 discloses
isolated and purified forms of FAAH. In one set of embodiments, the FAAH IC50 of the
subject compounds is defined according to inhibition of the rat enzyme under physiologically
relevant conditions. Fatty Amide Hydrolases (FAAHs) (Deutsch, D.G., et al., Prostaglandins
Leukot. Essent. Fatty Acid, 66, 201-210 (2002)) are enzymes responsible for the degradation
of lipid ethanolamides, (Fowler, C. J., et al., Biochem. Pharmacol 62, 517-526 (2001);
Patricelli, M. P., et al. Vitam. Horm., 62, 663-674 (2001)) e.g. anandamide (AEA, 1, Figure
1), (Devane, W. A., et al., Science 258,1946-1949 (1992)) oleoylethanolamide, (Rodriguez
de Fonseca, F., et al. Nature (London) 414, 209-212 (2001); Fu, J., et al., Nature (London)
425, 90-93 (2003)) and palmitoylethanolamide, (Calignano, A., et al. Nature (London) 394,
277-281 (1998); Lambert, D.M., et al., Curr. Med. Chem. 9, 663-674 (2002)) a biochemical
process which, along with selective trasport into cells in the case of AEA, (Di Marzo, V.,
Nature (London) 372, 686-691 (1994); Beltrama, M., et al., Science 277, 1094-1097 (1997);
Piomelli, D., et al., Proc. Natl. Acad. Sci. U.S.A. (2002)) brings about the cessation of the
cellular effects of these autacoids. Owing to the various and important physiological roles of
fatty acid ethanolamides, classes of small-molecule compounds able to block FAAH or
FAAHS but not bind to other endocannabinoid-metabolizing enzymes, e.g. monoglyceride
lipase (MGL), (Dinh, T.P., et al., Proc Natl. Acad. Sci. U.S.A. 99, 10819-10824 (2002)) or
cannabinoid receptors, would be advantageous both as pharmacological tools and as

prototypes for drug development projects (Piomelli, D., et al. Trends Pharmacol. Sci. 21,
218-224 (2000); Bisogno, T., et al., Curr. Phartn. Des. 8,533-547 (2002); Yarnell, A., Chem.
Eng. News 80(49), 32 (2002); Smith, A., Nat. Rev. DrugDiscov. 2,92 (2003); Wendeler, M.,
et al. Angew. Chem. Int. Ed. Al, 2938-2941 (2003)).
Compounds of the Invention Generally.
[83] Compounds of the invention may contain one or more asymmetric centers and can
thus occur as racemates and racemic mixtures, single enantiomers, diastereomeric mixtures
and individual diastereomers. The present invention is meant to comprehend all such
isomeric forms of the inventive compounds.
[84] Compounds of the invention include the diastereoisomers of pairs of enantiomers.
Diastereomers for example, can be obtained by fractional crystallization from a suitable
solvent, for example methanol or ethyl acetate or a mixture thereof. The pair of enantiomers
thus obtained may be separated into individual stereoisomers by conventional means, for
example by the use of an optically active acid as a resolving agent.
[85] Alternatively, any enantiomer of such a compound of the invention may be obtained
by stereospecific synthesis using optically pure starting materials of known configuration.
[86] The compounds of the present invention may have unnatural ratios of atomic isotopes
at one or more of their atoms. For example, the compounds may be radiolabeled with
isotopes, such as tritium or carbon-14. All isotopic variations of the compounds of the
present invention, whether radioactive or not, are within the scope of the present invention.
[87] The instant compounds may be isolated in the form of their pharmaceutically
acceptable acid addition salts, such as the salts derived from using inorganic and organic
acids. Such acids may include hydrochloric, nitric, sulfuric, phosphoric, formic, acetic,
trifluoroacetic, propionic, maleic, succinic, malonic and the like. In addition, certain
compounds containing an acidic function can be in the form of their inorganic salt in which
the counterion can be selected from sodium, potassium, lithium, calcium, magnesium and the
like, as well as from organic bases. The term "pharmaceutically acceptable salts" refers to
salts prepared from pharmaceutically acceptable non-toxic bases or acids including inorganic
bases or acids and organic bases or acids.
[88] The invention also encompasses prodrugs of the present compounds, which on
administration undergo chemical conversion by metabolic processes before becoming active
pharmacological substances. In general, such prodrugs will be derivatives of the present
compounds that are readily convertible in vivo into a functional compound of the invention.

Conventional procedures for the selection and preparation of suitable prodrug derivatives are
described, for example, in "Design of Prodrugs", ed. H. Bundgaard, Elsevier, 1985. The
invention also encompasses active metabolites of the present compounds.
[89] Some of the compounds described herein contain olefinic double bonds, and unless
specified otherwise, are meant to include both E and Z geometric isomers.
[90] Some of the compounds described herein may exist with different points of
attachment of hydrogen, referred to as tautomers. Such an example may be a ketone and its
enol form known as keto-enol tautomers. The individual tautomers as well as mixture thereof
are encompassed by the inventive Formulas.
[91] As used herein, the term "heteroatom" is meant to include oxygen (0), nitrogen (N),
sulfur (S).
[92] Where substituent groups are specified by their conventional chemical Formulae,
written from left to right, they equally encompass the chemically identical substituents which
would result from writing the structure from right to left, e.g., -CH2O- is intended to also
recite-OCH2-.
[93] The term "alkyl," by itself or as part of another substituent, means, unless otherwise
stated, a straight or branched chain, or cyclic hydrocarbon radical, or combination thereof,
which may be fully saturated, mono- or polyunsaturated and can include di- and multivalent
radicals, having the number of carbon atoms designated (i.e. C1-C10 means one to ten
carbons). Examples of saturated hydrocarbon radicals include, but are not limited to, groups
such as methyl, ethyl, n-propyl, isopropyl, n-butyl, /-butyl, isobutyl, sec-butyl, cyclohexyl,
cyclohexylmethyl, cyclopropylmethyl, homologs and isomers of, for example, n-pentyl, n-
hexyl, n-heptyi, n-octyi, and the like. An unsaturated alkyl group is one having one or more
double bonds or triple bonds. Examples of unsaturated alkyl groups include, but are not
limited to, vinyl, 2-propenyl, crotyl, 2-isopentenyl, 2-butadienyl, 2,4-pentadienyl, 3-(l,4-
pentadienyl), ethynyl, 1- and 3-propynyl, 3-butynyl, and the higher homologs and isomers.
The term "alkyl," unless otherwise noted, is also meant to include those derivatives of alkyl
defined in more detail below, such as "heteroalkyl." Alkyl groups which are limited to
hydrocarbon groups are termed "homoalkyl".
[94] The term "alkylene" by itself or as part of another substituent means a divalent radical
derived from an alkane, as exemplified, but not limited, by -CH2CH2CH2CH2-, and further
includes those groups described below as "heteroalkylene." Typically, an alkyl (or alkylene)
group will have from 1 to 24 carbon atoms, with those groups having 10 or fewer carbon

atoms being preferred in the present invention. A "lower alkyl" or "lower alkylene" is a
shorter chain alkyl or alkylene group, generally having eight or fewer carbon atoms.
[95] The terms "alkoxy," "alkylamino" and "alkylthio" (or thioalkoxy) are used in their
conventional sense, and refer to those alkyl groups attached to the remainder of the molecule
via an oxygen atom, an amino group, or a sulfur atom, respectively.
[96] The term "heteroalkyl," by itself or in combination with another term, means, unless
otherwise stated, a stable straight or branched chain, or cyclic hydrocarbon radical, or
combinations thereof, consisting of the stated number of carbon atoms and at least one
heteroatom selected from the group consisting of O, N, and S, and wherein the nitrogen and
sulfur atoms may optionally be oxidized and the nitrogen heteroatom may optionally be
quaternized. The heteroatom(s) 0, N and S may be placed at any interior position of the
heteroalkyl group or at the position at which the alkyl group is attached to the remainder of
the molecule. Examples include, but are not limited to, -CH2-CH2-O-CH3, -CH2-CH2-NH-
CH3, -CH2-CH2-N(CH3)-CH3, -CH2-S-CH2-CH3, -CH2-CH2,-S(O)CH3, -CH2-CH2-S(O)2-
CH3, -CH=CH-O-CH3, -Si(CH3)3, -CH2-CH=N-OCH3, and -CH=CH-N(CH3)-CH3. Up to
two heteroatoms may be consecutive, such as, for example, -CH2-NH-OCH3 and -CH2-O-
Si(CH3)3. Similarly, the term "heteroalkylene" by itself or as part of another substituent
means a divalent radical derived from heteroalkyl, as exemplified, but not limited by, -CH2-
CH2-S-CH2-CH2- and -CH2-S-CH2-CH2-NH-CH2-. For heteroalkylene groups, heteroatoms
can also occupy either or both of the chain termini (e.g., alkyleneoxy, alkylenedioxy,
alkyleneamino, alkylenediamino, and the like). Still further, for alkylene and heteroalkylene
linking groups, no orientation of the linking group is implied by the direction in which the
Formula of the linking group is written. For example, the Formula -C(O)2R'- represents both
-C(O)2R'-and-R'C(O)2-.
[97] The terms "cycloalkyl" and "heterocycloalkyl", by themselves or in combination with
other terms, represent, unless otherwise stated, cyclic versions of "alkyl" and "heteroalkyl",
respectively. Additionally, for heterocycloalkyl, a heteroatom can occupy the position at
which the heterocycle is attached to the remainder of the molecule. Examples of cycloalkyl
include, but are not limited to, cyclopentyl, cyclohexyl, 1-cyclohexenyl, 3-cyclohexenyl,
cycloheptyl, and the like. Examples of heterocycloalkyl include, but are not limited to, 1 -
(1,2,5,6-tetrahydropyridyl), 1-piperidinyl, 2-piperidinyl, 3-piperidinyl, 4-morpholinyl, 3-
morpholinyl, tetrahydroturan-2-yl, tetrahydroturan-3-yl, tetrahydrothien-2-yl,
tetrahydrothien-3-yl, 1 -piperazinyl, 2-piperazinyl, and the like.

[98] The terms "halo" or "halogen," by themselves or as part of another substituent, mean,
unless otherwise stated, a fluorine, chlorine, bromine, or iodine atom. Additionally, terms
such as "haloalkyl," are meant to include monohaloalkyl and polyhaloalkyl. For example, the
term "halo(C1-C4)alkyl" is mean to include, but not be limited to, trifiuoromethyl, 2,2,2-
trifluoroethyl, 4-chlorobutyl, 3-bromopropyl, and the like.
[99] The term "aryl" means, unless otherwise stated, a polyunsaturated, aromatic,
hydrocarbon substituent which can be a single ring or multiple rings (preferably from 1 to 3
rings) which are fused together or linked covalently. The term "heteroaryl" refers to aryl
groups (or rings) that contain from one to four heteroatoms selected from N, O, and S,
wherein the nitrogen and sulfur atoms are optionally oxidized, and the nitrogen atom(s) are
optionally quatemized. A heteroaryl group can be attached to the remainder of the molecule
through a heteroatom. Non-limiting examples of aryl and heteroaryl groups include phenyl,
1-naphthyl, 2-naphthyl, 4-biphenyl, 1-pyrrolyl, 2-pyrrolyl, 3-pyrrolyl, 3-pyrazolyl, 2-
imidazolyi, 4-imidazolyl, pyrazinyl, 2-oxazolyl, 4-oxazolyl, 2-phenyl-4-oxazolyl, 5-oxazolyl,
3-isoxazolyl, 4-isoxazolyl, 5-isoxazolyl, 2-thiazolyl, 4-thiazolyl, 5-thiazolyl, 2-furyl, 3-furyl,
2-thienyl, 3-thienyl, 2-pyridyl, 3-pyridyl, 4-pyridyl, 2-pyrimidyl, 4-pyrimidyl, 5-
benzothiazolyl, purinyl, 2-benzimidazolyl, 5-indolyl, 1-isoquinolyl, 5-isoquinolyl, 2-
quinoxalinyl, 5-quinoxalinyl, 3-quinolyl, and 6-quinolyl. Substituents for each of the above
noted aryl and heteroaryl ring systems are selected from the group of acceptable substituents
described below.
[100] For brevity, the term "aryl" includes both aryl and heteroaryl rings as defined above.
Thus, the term "arylalkyl" is meant to include those radicals in which an aryl group is
attached to an alkyl group (e.g., benzyl, phenethyl, pyridylmethyi and the like) including
those alkyl groups in which a carbon atom (e.g., a methylene group) has been replaced by, for
example, an oxygen atom (e.g., phenoxymethyl, 2-pyridyloxymethyl, 3-(l-
naphthyloxy)propyl, and the like).
[101] Each of the above terms (e.g., "alkyl," "heteroalkyl," "aryl" and "heteroaryl") are
meant to include both substituted and unsubstituted forms of the indicated radical. Preferred
substituents for each type of radical are provided below.
[102] Substituents for the alkyl and heteroalkyl radicals (including those groups often
referred to as alkylene, alkenyl, heteroalkylene, heteroalkenyl, alkynyl, cycloalkyl,
heterocycloalkyl, cycloalkenyl, and heterocycloalkenyl) can be one or more of a variety of
groups selected from, but not limited to: -OR', O, =NR', =N-OR', -NR'R", -SR', -halogen,
-OC(O)R', -C(O)R', -CO2R', -CONR'R", -OC(O)NR'R", -NR"C(O)R', -NR'-C(O)NR"R'",

-NR"C(O)2R', -NR-C(NR'R"R"')=NR"", -NR-C(NR'R")NR"', -S(O)R', -S(O)2R', -
S(O)2NR'R", -NRSO2R', -CN and -NO2 in a number ranging from zero to (2m'+l), where
m' is the total number of carbon atoms in such radical. R', R", R'" and R"" each preferably
independently refer to hydrogen, substituted or unsubstituted heteroalkyl, substituted or
unsubstituted aryl, e.g., aryl substituted with 1-3 halogens, substituted or unsubstituted alkyl,
alkoxy or thioalkoxy groups, or arylalkyl groups. When a compound of the invention
includes more than one R group, for example, each of the R groups is independently selected
as are each R', R", R'" and R"" groups when more than one of these groups is present. When
R' and R" are attached to the same nitrogen atom, they can be combined with the nitrogen
atom to form a 5-, 6-, or 7-membered ring. For example, -NR'R" is meant to include, but not
be limited to, 1-pyrrolidinyl and 4-morpholinyl. From the above discussion of substituents,
one of skill in the art will understand that the term "alkyl" is meant to include groups
including carbon atoms bound to groups other than hydrogen groups, such as haloalkyl (e.g.,
-CF3 and -CH2CF3) and acyl (e.g., -C(O)CH3, -C(O)CF3, -C(O)CH2OCH3, and the like).
[103] Similar to the substituents described for the alkyl radical, substituents for the aryl and
heteroaryl groups are varied and are selected from, for example: halogen, -OR', =0, =NR',
=N-OR', -NR'R", -SR', -halogen, -OC(O)R', -C(O)R', -CO2R', -CONR'R", -OC(O)NR'R",
-NR"C(O)R', -NR'-C(O)NR"R'", -NR"C(O)2R', -NR-C(NR'R"R"')=NR"",
-NR-C(NR'R")=NR"', -S(O)R', -S(O)2R', -S(O)2NR'R", -NRS02R', -CN and -NO2, -R', -
N3, -CH(Ph)2, fluoro(C1-C4)alkoxy, and fiuoro(C1-C4)alkyl, in a number ranging from zero to
the total number of open valences on the aromatic ring system; and where R', R", R'" and
R"" are preferably independently selected from hydrogen, (C1-C8)alkyl and heteroalkyl,
unsubstituted aryl and heteroaryl, (unsubstituted aryl)-(C1-C4)alkyl, and (unsubstituted
aryl)oxy-(C1-C4)alkyl. When a compound of the invention includes more than one R group,
for example, each of the R groups is independently selected as are each R', R", R'" and R""
groups when more than one of these groups is present.
Novel Fatty Acid Amide Hydrolase Inhibitory Compounds of the Invention
[104] The invention provides fatty acid amide hydrolase inhibitors of the Formula:


in which X is CH2, NH, O, or S; Q is 0 or S; Z is O or N, with the proviso that if Z is O then
one of R1 or R2 is absent; and R is an aromatic or alkyl or lipophilic moiety selected from the
group consisting of substituted or unsubstituted aryl; substituted or unsubstituted biphenylyl,
substituted or unsubstituted naphthyl, and substituted or unsubstituted phenyl; substituted or
unsubstituted terphenylyl; substituted or unsubstituted cycloalkyl, substituted or unsubstituted
heteroaryl, substituted or unsubstituted alkyl, substituted or unsubstituted heteroalkyl, and

in which p is a number from 0 to 3; m is a number from 0 to 4, and n is a number from 0 to
5, Z1 and Z2 are same or different and are independently a divalent radical selected from the
group consisting of -O-, -S-, -N(R5)-, -C(R6)=C(R7)-, -CR6,-C(R6)=N- and -N=C(R6)-
wherein R5 is selected from H, alkyl, heteroalkyl, alkenyl, alkynyl, cycloalkyl, aryl, acyl and
aroyl; R6 and R7 are independently Hor R6 and R7 optionally may combine to form a
saturated or unsaturated carbocyclic or heterocyclic ring, optionally substituted with one or
more Ra and Rb groups; Y is a linker, including but not limited to, a bond, -O-, -S-, -N(R5)-,
C1-C4 alkylene, (Z) or (E)-ethylene, and cycloalkyl with 3 to 6 carbon atoms; Ra and Rb are
independently selected from the group consisting of H, alkyl, heteroalkyl, alkenyl, alkynyl,
cycloalkyl, aryl, substituted aryl, aryialkyl, substituted arylalkyl, ketoalkyl, hydroxyalkyl,
aminoalkyl, -CH2-NR3R4, alkoxy, aryloxy, arylalkyloxy, halo, haloalkyl, cyano, hydroxy,
nitro, amino, -NR3R4, -SR5, carboxamido, -CONR3R4, -O-carboxamido, -O-CO-NR3R4,
sulfonamido, and -SO2NR3R4; and R3 and R4 are selected from H, alkyl, alkenyl, alkynyl,
cycloalkyl, hydroxyalkyl and mimo-memylamino and optionally R3 and R4 together with the
N atom to which they are attached to form a 5-7 membered cyclic ring. When Z\ is
-C(R6)=C(R7)- or -N=C(R6)-, and p is O, the aromatic ring of which Z2 is a member is

preferably in the meta or para position with respect to Y. More preferably, the position is
meta.
[105] In addition, R1 and R2 are independently selected from the group consisting of H,
substituted or unsubstituted alkyl, substituted or unsubstituted heteroalkyl, substituted or
unsubstituted cycloalkyl, and substituted or unsubstituted cycloheteroalkyl, and substituted or
unsubstituted phenyl, and substituted or unsubstituted aryl or heteroaryl, and wherein
optionally, when X is N, if taken together with the N atom to which they are attached, R1 and
R2, form a substituted or unsubstituted N-heterocycle or substituted or unsubstituted
heteroaryl with the atom to which they are each attached. In some embodiments, R1 and R2 is
H or hydrocarbyl selected from alkyl, alkenyl, alkynyl, cycloalkyl in which optionally one or
more carbons of these hydrocarbyl groups may be substituted with a heteroatom selected
from O, N-R5 ,and S-R5, aryl, acyl and aroyl and in which, optionally, when X is N, if taken
together with the N atom to which they are attached, R1 and R2, form a substituted or
unsubstituted N-heterocycle or substituted or unsubstituted heteroaryl with the atom to which
they are each attached.
[106] In some embodiments, in compounds of the above formula, X is O or S; Q is O or S;
and R is selected from the group consisting of substituted or unsubstituted aryl, substituted or
unsubstituted biphenylyl, substituted or unsubstituted naphthyl, and substituted or
unsubstituted phenyl, substituted or unsubstituted terphenylyl, substituted or unsubstituted
heteroaryl, and

in which p is a number from 0 to 3; m is a number from 0 to 4, and n is a number from 0 to
5, Z1 and Z2 are same or different and are independently a divalent radical selected from the
group consisting of -O-, -S-, -N(R5), -C(R6)=C(R7), and -N=C(R6)- wherein R5 is selected
from H, alkyl, heteroalkyl, alkenyl, alkynyl, cycloalkyl, aryl, acyl and aroyl; R6 and R7 are
independently selected from the group consisting of H, alkyl, heteroalkyl, alkenyl, alkynyl,
cycloalkyl, aryl, acyl and aroyl, wherein R6 and R7 optionally may combine to form a
saturated or unsaturated carbocyclic or heterocyclic ring, optionally substituted with one or
more Ra and Rb groups; and Y is a linking member. Y may be a bond or selected from the
group consisting of -O-, -S-, -N(R5>, -S(R5> , -C1-C4 alkylene, (Z> or (E)-ethylene, and

cycloalkyl with 3 to 6 carbon atoms; each Ra and each Rb are independently selected from
the group consisting of H, alkyl, heteroalkyl, alkenyl, alkynyl, cycloalkyl, aryl, substituted
aryl, arylalkyl, substituted arylalkyl, ketoalkyl, hydroxyalkyl, aminoalkyl, -CH2-NR3R4,
alkoxy, aryloxy, arylalkyloxy, halo, haloalkyl, cyano, hydroxy, nitro, amino, -NR3R4, -SR5,
carboxamido, -CONR3R4, -O-carboxamido, -O-CO-NR3R4, sulfonamido, and -SC^NRsRi,
wherein R3 and R4 are selected from H, alkyl, alkenyl, alkynyl, cycloalkyl, hydroxyalkyl and
imino-methylamino and optionally R3 and R4 together with the N atom to which they are
attached to form a 5-7 membered cyclic ring; and Ri and R2 are independently selected from
the group consisting of H, substituted or unsubstituted alkyl, substituted or unsubstituted
heteroalkyl, substituted or unsubstituted cycloalkyl, and substituted or unsubstituted
cycloheteroalkyl, and optionally Ri and R2, may be taken together with the N atom to which
they are attached to form a substituted or unsubstituted ring.
[107] In another embodiment, in the compound of Formula I, X is O and Q is O. In a
further embodiment of such compounds Z is N. In a still further embodiment, p is 0, m is 1
and n is 0,1,2, or 3. In still another embodiment, m is 0 and n is 1,2, or 3.
[108] In still another embodiment of the compound of Formula I, R is selected from the
group consisting of substituted or unsubstituted biphenylyl, substituted or unsubstituted
naphthyl, substituted or unsubstituted terphenylyl, and substituted or unsubstituted cis-stilbyl
((Z>C6H5-CH=CHC6H5-). In a further embodiment of such R compound, X is also O and
QisOandZisN.
[1091 hi an exemplary embodiment, R is substituted or unsubstituted biphenylyl. In a
further embodiment of such biphenyl compounds, X is also O and Q is O and Z is N. In a
still further embodiment, at least one of Ri and R2 is H.
[1101 hi yet another embodiment, in the compound of Formula I, Ri is Ci-Cghomoalkyl, Ci-
C8 heteroalkyl, or Ci-Cs cycloalkyl. In a further embodiment, the Ci-Cg alkyl is methyl,
ethyl, H-propyl, i-propyl, n-butyl, sec-butyl, te/t-butyl, pentyl, hexyl, cyclopentyl, cyclohexyl,
cycloheptyl, cyclopentenyl, cyclohexenyL or cycloheptenyl. A particularly preferred RI
group is Ri is cyclohexyl. In a further embodiment of such compounds, X is O; Q is O and
Z is N. In an exemplary further embodiment, R is substituted or unsubstituted biphenyl,
terphenyl, or stilbyl in which up to 3 atoms of the aromatic ring may be substituted with a
heteroatom selected from the group consisting of NR5, O, or S wherein R5 is as defined in
any of the above.
[Ill] In a further embodiment, the compound of Formula I has an R, group which is a
piperidinyi, furyl, furfuryl, furanyl, and morpholinyl and may be substituted or unsubstituted.

In a further embodiment of such compounds, X is preferably O; Q is O and Z is N. In a still
further embodiment, the R group is substituted or unsubstituted biphenyl, terphenyl, or
stilbyl in which up to 3 atoms of the aromatic ring may be substituted with a heteroatom
selected from the group consisting of NR5, O, or S wherein R5 is selected from H, alkyl,
alkenyl, alkynyl, cycloalkyl, aryl, acyl and aroyl.
[112] In one set of embodiments, the compound of Formula I is of the formula:

In the above formula, m is a number from 0 to 4 and n is a number from 0 to 5. In some
embodiments, m is 0 or 1 and n is 2 or 3. In the above formula, each R* and each Rb are
independently selected from the group consisting of H, alkyl, alkenyl, alkynyL cycloalkyl,
aryl, substituted aryl, arylalkyl, substituted arylalkyl, ketoalkyl, hydroxyalkyl, aminoalkyl, -
CH2-NR3R4, alkoxy, aryloxy, halo, haloalkyL cyano, hydroxy, nitro, amino, -NR3R4,
carboxamido, -ONR3R4, -O-carboxamido, -O-CO-NR3R4, sulfonamido, -SO2NR3R4. R3 and
R4 are selected from the group consisting of H, alkyl, heteroalkyl, alkenyl, alkynyl,
cycloalkyl, hydroxyalkyl and imino-methylamino or R3 and R4 may combine with the N atom
to which they are attached to form a 5-7 membered cyclic ring. In some further
embodiments, each R, and each Rb are independently selected from H, ketoalkyl,
hydroxyalkyl, aminoalkyl, -CH2-NR3R4, alkoxy, halo, haloalkyl, cyano, hydroxy, nitro,
amino, -NR3R4, carboxamido, -ONR3R4, -O-carboxamido, -O-CO-NR3R4, sulfonamido, and -
SO2NR3R4.
[113] Yet another embodiment is represented by a compound of the following formula:


In the above formula, Rai and Ra2 are independently selected from the group consisting of
H, alkyl, alkenyl, alkynyl, cycloalkyl, aryl, substituted aryl, arylalkyl, substituted arylalkyl,
ketoalkyl, hydroxyalkyl, aminoalkyl, CH2-NR3R4, alkoxy, aryloxy, halo, haloalkyl, cyano,
hydroxy, nitro, amino, NR3R4, carboxamido, CONR3R4,0-carboxamido, O-CO-NR3R4,
sulfonamido, and SO2NR3R4; and R3 and R4 are selected from H, alkyl, alkenyl, alkynyl,
cycloalkyl, hydroxyalkyl and miino-methylamino. In addition, R3 and R4 may together with
the N atom to which they are attached combine to form a 5-7 membered cyclic ring. In a
particular set of embodiments, each R« and each Rb is independently selected from H,
ketoalkyl, hydroxyalkyl, aminoalkyl, -CH2-NR3R4, alkoxy, halo, haloalkyl, cyano, hydroxy,
nitro, amino, -NR3R4, carboxamido, -ONR3R4, -O-carboxamido, -O-CO-NR3R4,
sulfonamido, and -SO2NR3R4. In a further set of such embodiments, at least one of Rai and
Ra2 is H. In another of such embodiments, Rai is selected from the group consisting of
-C(O)NH2. -C(O)CH3, or -(ClfckOH and Ra2 is H. In another set of embodiments, Rai and
Raj are each H. In still further embodiments of such compounds Ri is Q-Cs alkyl and R2 is
H.
[114] In other embodiments, the compound of Formula 1 has an Ri of Ci-Cs alkyl an X of
O, a Q of S and a Z of N. In some such embodiments, R is substituted or unsubstituted
phenyl, biphenyl, terphenyl, or stilbyl.
1115] In other embodiments, the compound of Formula 1 has an X of 0, a Q of 0 and a Z of
N, and R is substituted or unsubstituted biaryl or heterobiaryl. m some further embodiments,
the heterobiaryl has up to 3 members of the heterobiaryl rings selected from the group
consisting of O, N, or S. In still other embodiments, the heterobiaryl is bipyridyl or
phenylpyridyl.
[116] In yet another embodiment, the compound of Formula I is n-butyl 4-benzyloxyphenyl
carbamate or N-cyclohexyl 3'-carboxamido-biphenyl-3-yl carbamate.

[117] Other embodiments are drawn to those compounds of Formula I which have an IC50
for inhibiting the human fatty acid amide hydrolase of less than 1 micromolar. In such
embodiments, compounds which have an IC50 for inhibiting the human fatty acid amide
hydrolase of from 100 to 10 nanomolar, or 10 to 1 nanomolar, or less than 10 nanomolar are
exemplary.
[118] In other embodiments, the compounds of Formula I are compounds in which the
molecular weight of the R-X- group is greater than the molecular weight of the -NR1R2
group. In other embodiments, the bulk of the R-X- group is greater than that for the -NR1R2
group. In further embodiments of such compounds, X is O; Q is O and Z is N. In still
further embodiments, R is substituted or unsubstituted aryl, including but not limited to
biphenyl, terphenyl, and cis-stilbyl compounds.
[119] Li another embodiment of the compound of Formula I, X is O; Q is O; Z is N and
[120] R is substituted or unsubstituted biaryl and substituted or unsubstituted heterobiaryl.
In a further embodiment, R is substituted or unsubstituted biaryl or heterobiaryl having up to
3 members of the heterobiaryl rings selected from the group consisting of O, N, or S. In still
further embodiments, Ri and R2 are independently selected from the group consisting of H,
unsubstituted or substituted homoalkyl, substituted or unsubstituted heteroalkyl, substituted
or unsubstituted cycloalkyl, substituted or unsubstituted heterocycloalkyl, and optionally Ri
and R2 may be taken together to form a substituted or unsubstituted heterocycle with N; and
Rai and Ra2 are independently selected from the group consisting of H, alkyl, alkenyl,
alkynyl, cycloalkyl, ketoalkyl, hydroxyalkyl, aminoalkyl, -CH2-NR3R4, alkoxy, aryloxy, halo,
haloalkyl, cyano, hydroxy, nitro, amino, -NR3R4, carboxamido, -CONR3R4, -O-
carboxamido, -O-CO-NR3R4, sulfonamido, and -SO2NR3R4; and R3 and R4 are selected from
H, alkyl, heteroalkyl, alkenyl, alkynyl, cycloalkyl, hydroxyalkyl and imino-methylamino or
R3 and R4 may combine together with the N to which they are attached to form a 5-7
membered cyclic ring. In a further such embodiment, Ri is Ci-Cs homoalkyl, Ci-Cs
heteroalkyl, or Ci-Cg cycloalkyl. In a still further embodiment, R2 is H. In a still further
embodiment, Ri is cyclohexyl and R2 is H.
[121] The invention also provides fatty acid amide hydrolase inhibitors of the Formula:


in which X is NH, 0, or S; Q is 0 or S; Z is 0 or N; and R is an aromatic or alkyl or
lipophilic moiety selected from the group consisting of substituted or unsubstituted aryl;
substituted or unsubstituted biphenylyl, substituted or unsubstituted naphthyl, and substituted
or unsubstituted phenyl; substituted or unsubstituted terphenylyl; substituted or unsubstituted
cycloalkyl, heteroaryl, or alkyl; and wherein Ri and R2 are independently selected from the
group consisting of H, substituted or unsubstituted alkyl, substituted or unsubstituted
heteroalkyl, and substituted or unsubstituted phenyl; with the provision that if Z is 0, one of
Ri or R2 is absent, and further with the proviso that if Z is N, Ri and R2 may be optionally
taken together to form a substituted or unsubstituted AT-heterocycle or substituted or
unsubstituted heteroaryl with the N atom to which they are each attached. In some
embodiments, Ri and R2 is H or hydrocarbyl selected from alkyl, alkenyl, alkynyl, cycloalkyl
in which optionally one or more carbons of these hydrocarbyl groups may be substituted
with a heteroatom selected from 0, N-R5 ,and S-R5, aryl, acyl and aroyl; R$ and R7
[122] In one embodiment, the FAAH inhibitory compound of Formula la has an IC50 of less
than 10 uM or 1 uM. In another, embodiment the compound of Formula I is an FAAH
inhibitor with an IC50 of less than 0.01 pM. In another embodiment, the compound of
Formula I is an inhibitor of FAAH with an IC50 of from about 1 uM to 0.01 pM, or from
about 0.01 to 0.001 uM.
1123] In one embodiment of a compound of Formula la, X is O, Q is O; and Z is N. In
another embodiment, X is O; Q is O; and Z is N; Ri is H and R2 is cyclohexyl. In another
embodiment, X is O, Q is O; Z is N and R is substituted or unsubstituted biphenylyl. In a
further embodiment, X is O, Q is O; Z is N; Ri is H, R2 is cyclohexyl and R is substituted or
unsubstituted biphenylyl or substituted or unsubstituted phenyl.
[124] In another embodiment of a compound of Formula la, X is 0; Q is 0; and Z is N; Ri
is H and R2 is cyclohexyl. In another embodiment, R is substituted or unsubstituted
biphenylyl. In another embodiment, Ri is H and R2 is cyclohexyl and R is substituted or
unsubstituted biphenylyl or substituted or unsubstituted phenyl.
[125] In one embodiment the compound of Formula I is of the Formula


in which R is an aromatic or alkyl or lipophilic moiety selected from the group consisting of
substituted or unsubstituted aryl; substituted or unsubstituted biphenylyl, substituted or
unsubstituted naphthyl, and substituted or unsubstituted phenyl; substituted or unsubstituted
terphenylyl; substituted or unsubstituted cycloalkyl, heteroaryl, or alkyl; and wherein Ri and
R2 are independently selected from the group consisting of H, substituted or unsubstituted
alkyl, substituted or unsubstituted heteroalkyl, and substituted or unsubstituted phenyl,
substituted or unsubstituted aryl, substituted or unsubstituted heteroaryl, and optionally
wherein if Ri and R2 are taken together, form a substituted or unsubstituted AT-heterocycle or
substituted or unsubstituted heteroaryl with the atom to which they are each attached. In
some embodiments, the compound of Formula I or II has an R group with a greater molecular
weight or mass than the -NR1R2 portion of the compound.
[126] In another embodiment, the FAAH inhibitory compound of Formula II has an IC50 of
less man 10 pM or 1 pM. In another? embodiment the compound of Formula II is an FAAH
inhibitor with an IC50 of less than 0.01 uM. In another embodiment, the compound of
Formula II is an inhibitor of FAAH with an an IC50 of from about 1 uM to 0.01 uM, or from
about 0.01 to 0.001 pM.
[127] In one embodiment, R is naphthyl in the compound of Formula I, Ia-c, n, or Ila-b, and
m.
[128J In another embodiment, Ri is H and R2 is cyclohexyl in a compound of Formula I, Ia-
c, n, or Ila-b. In another embodiment, Ri is H and R2 is alkyl in the compound of Formula I,
Ia-c, n, or Ila-b.
[129] In another embodiment, Rj is H and R2 is alkyl and R is alkyl in the compound of
Formula I, Ia-c, n, or Ila-b.
[130] In another embodiment, Ri is H and R2 is cyclohexyl and R is substituted or
unsubstituted aryl in the compound of Formula Ic or Formula H In a further embodiment, in
the compound of Formula Ic or Formula n, R is substituted or unsubstituted biphenylyl. m a
still further embodiment, the biphenyl is substituted with halogen (e.g., F) or alkyl (e.g.,
methyl) or amino or amido or trifluromethyl.
[131] Exemplary compounds according to Formula I, Ia-c, n,or Ila-b include, but are not
limited to, the following compounds from Table 3: 2-naphthyl 7V-cyclohexylcarbamate; 4-
(benzyloxy) phenyl N-butyl carbamate; 6-bromo-2-naphthylyl N-butylcarbamate; 4-
biphenylyl JV-cyclohexylcarbamate; hexyl Af-cyclohexyl carbamate; p-tolyl-N-

cyclohexylcarbamate; 0-butyl 4-(4'methoxyphenoxycarbonyl)-phenyl carbonate; and 4-
fluorophenyl iV-butyl carbamate.
[132] In another embodiment, the compound of Formula I is UCM532 or UCM597.
[133] In another embodiment, the inventive FAAH inhibitor is a compound of Formula II in
which Ri is H: R2 is C1-C10 alkyl; and R is substituted aryl. In a further embodiment, R2 is a
tert-butyl, sec-butyl, or n-butyl moiety. In another embodiment of the compounds according
to the above Formula H, R2 is cyclohexyl.
[134] In another embodiment, the inventive FAAH inhibitor is a compound of Formula I, n,
or Ha in which Ri is H; R2 is C1-C10 alkyl; and R is one of the following:

[135] In other embodiments, wherein R is one of Rf, Rg, Re, Rd, Re, or Rh, R2 is a tert-
butyl, sec-butyl, n-butyl moiety, or cyclohexyl. In other embodiments, wherein R is one of
Rf, Rg, Re, Rd, Re, or Rh, each of the benzene rings are further independently and optionally

substituted with one, two, three, or four substituents, other than a sole H atom, as defined
above for aryl or heteroaryl groups.
[136] In another embodiment, the compound is of Formula Ha:

in which R2 is C1-C10 alkyl. In a further embodiment, R2 is a cyclohexyl or tert-butyl, sec-
butyl, or n-butyl moiety. In another embodiment of the compounds according to the above
Formuala HA, R2 is cyclohexyl.
1137] In other embodiments, the inventive compounds are compounds of Formula Ha which
are additionally further independently and optionally substituted on each of the benzene rings
with one, two, three, or four other substituents, other than a sole H atom, as defined above for
aryl or heteroaryl groups
[138] In another embodiment, compound or the FAAH inhibitor is a compound of Formula
lib:

[139] in which Ri is alkyl. In a further embodiment, Ri is lower Ci-Q alkyl. In a still
further embodiment, Ri is a embodiments, the inventive compounds are compounds of Formula lib which are additionally
further independently and optionally substituted on each of the benzene rings with one, two,
three, or four additional substituents, other than a sole H atom, as defined above for aryl or
heteroaryl groups.

[140] In some embodiments, a compound according to the invention is any compound of
Table I through Table VI which has a FAAHIC50 of less than 1 micromolar.
[141] In the embodiments, the compounds (e.g., the compounds of Formulae I, la-Ic, II, Ila-
Ilb, IE), include their pharmaceutically acceptable salts and biologically active isomers and
conformers.
[142] In another aspect, the invention provides FAAH inhibitors and compounds or the
following general formula:

in Formula IE, R\ and R2 are independently selected from the group consisting of H,
unsubstituted or substituted homoalkyl, substituted or unsubstituted heteroalkyl, substituted
or unsubstituted cycloalkyl, substituted or unsubstituted heterocycloalkyl, and optionally Ri
and R2, may be taken together to form a substituted or unsubstituted heterocycle with N; U
is a hydrophobic spacer, wherein the spacer comprises substituted or unsubstituted aryl; and
V is a hydrophilic moiety having at least one functional group capable of forming a hydrogen
bond. In addition, the hydrophobic spacer is at least 9 angstroms in length; and the
hydrophilic moiety is attached to the spacer at a point from 8 to 12 angstroms distant from a
point at which the hydrophobic spacer is covalently attached to the rest of the inhibitor.
[143] In a further embodiment, the inhibitor or compound of Formula HI has a
hydrophobic spacer comprising a first and a second aromatic ring, wherein the first and
second aromatic rings are covalently attached by a linker selected from the group consisting
of a bond, a single heteroatom, and substituted or unsubstituted Ci to C4 alkylene.
[144] In another embodiment, V is selected from the group consisting of ketoalkyl,
hydroxyalkyl, aminoalkyl, -CH2-NR3R4, alkoxy, aryloxy, halo, haloalkyl, cyano, hydroxy,
nitro, amino, -NR3R4, carboxamido, -CONR3R4, -O-carboxamido, -O-CO-NR3R4,
sulfonamido, and -SO2NR3R4; wherein R3 and R4 are selected from H, alkyl, heteroalkyl,
alkenyl, alkynyl, cycloalkyl, hydroxyalkyl and immo-methylamino or R3 and R4 may
combine with the N atom to which they are attached to form a 5-7 membered cyclic ring.
[145] In another embodiment, U and V are each set forth as described above and the
hydrophilic moiety is attached to the spacer at a point about 9 to 11 angstroms from a point

at which the spacer is covalently attached to the rest of the inhibitor. In a further
embodiment, the hydrophilic moiety is attached to the spacer at a point about 10 angstroms
from a point at which the spacer is covalently attached to the rest of the inhibitor.
[146] In another embodiment, U and V taken together have a greater molecular weight or
bulk than the remainder of the molecule or than the -NR1R2 moiety.
[1471 In some embodiments, in Formula I, Ia-c, n, Ila-b, or HI, Ri and R2 are independently
selected from H or hydrocarbyl selected from alkyl, alkenyl, alkynyl, and cycloalkyl in which
optionally one or more carbons of these hydrocarbyl groups may be substituted with a
heteroatom selected from 0, N-R5 ,and S-R5, and in which, optionally, when X is N, if taken
together with the N atom to which they are attached, Ri and R2 form a substituted or
unsubstituted AT-heterocycle or substituted or unsubstituted heteroaryl. In these embodiments,
R5 is a H or a hydrocarbyl selected from alkyl, alkenyl, alkynyl, cycloalkyl in which
optionally one or more carbons of these hydrocarbyl groups may be substituted with a
heteroatom selected from O, NH ,and SH.
[148] In some embodiments, in Formula I, Ia-c, n, Ila-b, or HI, Ri and R2 are
independently selected from H or hydrocarbyl selected from alkyl, alkenyl, alkynyl, and
cycloalkyl in which optionally one or more carbons of these hydrocarbyl groups may be
substituted with a heteroatom selected from O, N-R5 ,and S-R5, and in which, optionally,
when X is N, if taken together with the N atom to which they are attached, Ri and R2 form a
substituted or unsubstituted JV-heterocycle or substituted or unsubstituted heteroaryl. In these
embodiments, R5 is a H or a hydrocarbyl selected from alkyl, alkenyl, alkynyl, cycloalkyl in
which optionally one or more carbons of these hydrocarbyl groups may be substituted with a
heteroatom selected from O, NH ,and SH. In still further embodiments in which p is a
number from 0 to 3; m is a number from 0 to 4, and n is a number from 0 to 5, Z\ and Z2 are
same or different and are independently a divalent radical selected from the group consisting
of -O-, -S-, -N(R5)-, -CCReKXR?)-, -CR6 ,-N=C(R independently selected from H or hydrocarbyl in which the hydrocarbyl is alkyl, alkenyl,
alkynyl, cycloalkyl and optionally one or more carbons of these hydrocarbyl groups may be
substituted with a heteroatom selected from O, N-R5 ,and S-R5, aryl, acyl and aroyi. In
preferred sets of these embodiments, the hydrocarbyl groups are each independently a Ci to
C10 hydrocarbyl group which may be optionally substituted with O, N-R5, and S-R5 in which
R5 is also a Ci to C10 hydrocarbyl. In a more preferred set of embodiments, the R« and R« are
each independently a d to Cg hydrocarbyl group which may be optionally substituted with
O, N-R5, and S-R5 in which the R5 hydrocarbyl groups are Ci to C6.

[149] In some preferred embodiments, in Formula I, Ia-c, n, Ila-b, or in, Ri is selected
from the group consisting of piperidinyl, furyl, furfuryl, furanyl, morpholinyl, is 2-,3-,4-
piperidinyl, 2- and 3-morpholinyl, 2- and 3-furyl, furfuryl, 2- and 3-pyrryl or 2- or 3-thienyL.
[150] In some preferred embodiments, in Formula I, Ia-c, n, Ha-b, or HI,, Ri is Q-Cs
hydrocarbyl selected from alkyl and cycloalkyl and optionally one or more carbons of these
hydrocarbyl groups may be substituted with a heteroatom selected from O, N-R5 ,and S-R5.
In still further such embodiments, R2 is H. In still further preferred embodiments, R is
substituted or unsubstituted biphenylyl.
[151] m some embodiments, in Formula I, Ia-c, H, na-b, or m, R* and Rb are independently
selected from the H or a hydrocarbyl group consisting of alkyl, alkenyl, alkynyl, cycloalkyl,
wherein optionally one or more carbons of these hydrocarbyl groups may be substituted with
a heteroatom selected from O, N-R5 ,and S-Rs, aryl; substituted aryl; aralkyl; substituted
aralkyl; ketoalkyl; hydroxyalkyl; aminoalkyl; -CH2-NR3R4, alkoxy, aryloxy, aralkyloxy, halo,
haloalkyl, cyano, hydroxy, nitro, amino, -NR3R4, SR5, carboxamido, -CONR3R4,0-
carboxamido, -O-CO-NR3R4, sulfonamido, and -SO2NR3R4, wherein R3 and R4 are selected
from H or a hydrocarbyl group selected from alkyl, alkenyl, alkynyl, cycloalkyl,
hydroxyhomoalkyl and imino-methylamino and optionally R3 and R4 may combine to form a
5-7 membered cyclic ring with the N to which they are attached. In these embodiments, R5 is
a H or a hydrocarbyl selected from alkyl, alkenyl, alkynyl, cycloalkyl in which optionally one
or more carbons of these hydrocarbyl groups may be substituted with a heteroatom selected
from O, NH ,and SH. In preferred sets of these embodiments, the hydrocarbyl groups are
each independently a Ci to C10 hydrocarbyl group which may be optionally substituted with
O, N-R5, and S-R5 in which R5 is also a Ci to C10 hydrocarbyl. In a more preferred set the
R» and Rb are each independently a Ci to Ck hydrocarbyl group which may be optionally
substituted with O, N-R5, and S-R5 in which the R5 hydrocarbyl groups are Ci to Q.
[152] In some embodiments, in Formula I, Ia-c, n, na-b, or m, Ri and R2 are
independently selected from H or hydrocarbyl selected from alkyl, alkenyl, alkynyl, and
cycloalkyl in which optionally one or more carbons of these hydrocarbyl groups may be
substituted with a heteroatom selected from O, N-R5 ,and S-R5, and in which, optionally,
when X is N, if taken together with the N atom to which they are attached, Ri and R2 form a
substituted or unsubstituted iV-heterocycle or substituted or unsubstituted heteroaryl. In these
embodiments, R5 is a H or a hydrocarbyl selected from alkyl, alkenyl, alkynyl, cycloalkyl in
which optionally one or more carbons of these hydrocarbyl groups may be substituted with a
heteroatom selected from O, NH ,and SH. In still further embodiments, in which p is a

number from 0 to 3; m is a number from 0 to 4, and n is a number from 0 to 5, Zi and Zt are
same or different and are independently a divalent radical selected from the group consisting
of -O-, -S-, -N(R5)-, -C(R6)=€(R?)-, -CR*, -C(R independently selected from H or hydrocarbyl in which the hydrocarbyl is alkyl, alkenyl,
alkynyl, cycloalkyl and optionally one or more carbons of these hydrocarbyl groups may be
substituted with a heteroatom selected from O, N-R5 ,and S-Rs, aryl, acyl and aroyl. In
preferred sets of these embodiments, the hydrocarbyl groups are each independently a Ci to
Cio hydrocarbyl group which may be optionally substituted with O, N-R5, and S-R5 in which
R5 is also a Ci to Cio hydrocarbyl. In a more preferred set of such compounds, the R« and R$
are each independently a d to C6 hydrocarbyl group which may be optionally substituted
with O, N-R5> and S-R5 in which the R5 hydrocarbyl groups are Ci to C6. In still further
embodiments, R« and Rb are independently selected from the H or a hydrocarbyl group
consisting of alkyl, alkenyl, alkynyl, cycloalkyl, wherein optionally one or more carbons of
these hydrocarbyl groups may be substituted with a heteroatom selected from O, N-R5 ,and S-
R5, aryl; substituted aryl; aralkyl; substituted aralkyl; ketoalkyl; hydroxyalkyl; aminoalkyl; -
CH2-NR3R4, alkoxy, aryloxy, aralkyloxy, halo, haloalkyl, cyano, hydroxy, nitro, amino, -
NR3R4, SR5, carboxamido, -CONR3R4,0-carboxamido, -O-CO-NR3R4, sulfonamido, and -
SO2NR3R4, wherein R3 and R4 are selected from H or a hydrocarbyl group selected from
alkyl, alkenyl, alkynyl, cycloalkyl, hydroxyhomoalkyl and imino-methylamino and optionally
R3 and R4 may combine to form a 5-7 membered cyclic ring with the N to which they are
attached. In these embodiments, R5 is a H or a hydrocarbyl selected from alkyl, alkenyl,
alkynyl, cycloalkyl in which optionally one or more carbons of these hydrocarbyl groups
may be substituted with a heteroatom selected from O, NH ,and SH. In preferred sets of
these embodiments, the hydrocarbyl groups are each independently a Ci to Cio hydrocarbyl
group which may be optionally substituted with O, N-R5, and S-R5 in which R5 is also a Ci to
Cio hydrocarbyl. In a more preferred set the R* and Rb are each independently a Ci to C6
hydrocarbyl group which may be optionally substituted with O, N-R5, and S-R5 in which the
Rs hydrocarbyl groups are Q to Q.
Synthesis of inhibitors of Formula I or Formula II
1153] The compounds of the present invention can be made with commercially available
starting materials using straightforward chemistry. Carbamates are well known in the art.
The following procedures are exemplary synthetic routes, which are intended to illustrate, but

not to limit the present invention. One of ordinary skill in the art will recognize other
variations, modifications, and alternatives.
[154] In one example, n-butylcarbamic acid 4-benzyloxyphenyl ester (UCM532) (4) and 4-
fluorophenylcarbamic acid 4-benzyloxyphenyl ester (8) were obtained by treatment of 4-
benzyloxyphenol with n-butylisocyanate, and 4-fluorophenylisocyanate, respectively, with a
catalytic amount of triethylamine in refluxing toluene. The resulting products were obtained
in good yields.
[155] Similarly, cyclohexylcarbamic acid biphenyl-3-yl ester (5), cyclohexylcarbamic acid
5-phenylpentyl ester (7), and cyclohexylcarbamic acid 3'-carbamoylbiphenyl-3-yl ester
(UCM597) (6) were synthesized by reacting cyclohexylisocyanate with 3-phenylphenol, 5-
phenylpentan-1-ol, and 3'-hydimybiphenyl-3-carboxylic acid amide, respectively. Again, the
resulting products were obtained in good yield.
[156] The latter reactant was prepared as follows: 3-bromobenzoic acid amide, obtained by
reaction of 3-bromobenzonitrile and sodium perborate, was coupled with
methoxyphenylboronic acid to give 3'-methoxybiphenyl-3-carboxylic acid amide, which was
hydrolized with BBr3 to generate the desired 3Mby&oxybiphenyl-3-carboxylic acid amide.
Detailed synthetic procedures and physicochemical data will be reported elsewhere.
[157] Other methods suitable for making the subject compounds are disclosed in Tarzia et
al. J. Med. Chem. 46:2352-2360 (2003) and Kathuria et al. Nature Medicine 9(1): 76 (2003)
which are incorporated herein by reference.
[158] Other FAAH Inhibitors for Use in the Treatment of Anxiety
[159] Trifluoroketone inhibitors such as the compound of Formula IV are also contemplated
for use in inhibiting FAAH to raise endogenous levels of anandamide or treat the subject
conditions and disorders.

[160] Such compounds are taught in U.S. Patent Application No. 6,096,784 herein
incorporated by reference.

[161] Other compounds for use according to the invention include octylsulfonyl and
octylphosphonyl compounds. See Quistand et al. in Toxicology and Applied Pharmacology
179: 57-63 (2002). See also Quistand et al. in Toxicology and Applied Pharmacology 173:
48-55 (2001).
[162J Other compounds for use according to the invention include the alpha-keto-
oxazolpyridines which are reversible and extremely potent inhibitiors of FAAH. See Boger
et al., PNAS USA 97:5044-49 (2000). Exemplary compounds include compounds of the
Formula:

[163] wherein R is an alpha-keto oxazolopyridinyl moiety such as

[164] Boger et al. teach other exemplary compounds of the invention including substituted
alpha-keto-heterocycle analogs of fatty acid amides. In particular, wherein R is an alpha-keto
oxazolopyridinyl moiety and the fatty acid moiety is a homolog of oleic acid or arachidonic
acid.
[165] Other FAAH inhibitors for use according to the invention include fatty acid sulfonyl
fluorides such as compound AM374 which irreversibly binds FAAH. See Deutsch et al.,
Biochem. Biophys Res Commun. 231:217-221 (1997).
Methods of Screening Compounds for FAAH Inhibitory Activity.
[166] Methods for screening compounds for FAAH inhibitory activity in vitro are well
known to one of ordinary skill in the art. Such methods are taught in Quistand et al. in

Toxicology and Applied Pharmacology 179: 57-63 (2002); Quistand et al. in Toxicology and
Applied Pharmacology 173, 48-55 (2001); Boger et al., Proc. Natl. Acad. Sci. U.S.A. 97,
504449(2000).
(1671 Methods for screening compounds for FAAH inhibitory activity in vivo and increased
endogenous cannabinoid levels or activity are known to one of ordinary skill in the art. Such
methods include measurement of fatty acid ethanolamides in tissue and are taught in
Quistand et al. in Toxicology and Applied Pharmacology 179, 57-63 (2002); Quistand et al.
in Toxicology and Applied Pharmacology 173,48-55 (2001); Boger et al., Proc. Natl. Acad.
Sci. U.S.A. 97:5044-49 (2000). See U.S. Patent No. 6,096,784. See also PCT Publication
WO 98/24396. See Cravatt et al. Proc. Natl. Acad. Sci. U.S.A. 98,9371-9376 (2001).
[168] Methods for Assaying ACHE and NTE Inhibitory Activity.
[169] One of ordinary skill in the art would know how to screen a substance for an
inhibitory effect on ACHE or NTE. See for instance Quistand et al. in Toxicology and
Applied Pharmacology 179, 57-63 (2002); and Quistand et al. in Toxicology and Applied
Pharmacology 173,48-55 (2001).
[170] Cannabinoid CB1 Receptor Activity
[171] A variety of means may be used to screen cannabinoid CB1 receptor binding activity
in order to identify the compounds according to the inventioa A variety of such methods are
taught in U.S. Patent No. 5,747,524 and U.S. Patent No. 6,017,919.
[172] CB1 receptor binding assays are well known to one of ordinary skill in the art. For
instance, see, U.S. Patent Application No. US 2001/0053788 published on December 20,
2001, U.S. Patent No. 5,747,524, and U.S. Patent No. 5,596,106 and (see: Felder, C. C, et al.,
Proa Natl. Acad. Sci., 90, 7656-7660 (1993) each of which is incorporated herein by
reference. The affinity of an agent for cannabinoid CB1 receptors can be determined using
membrane preparations of Chinese hamster ovary (CHO) cells in which the human cannabis
CB1 receptor is stably transfected in conjunction with [3H]CP-55,940 as radioligand. After
incubation of a freshly prepared cell membrane preparation with the [3H]-ligand, with or
without addition of compounds of the invention, separation of bound and free ligand can be
performed by filtration over glass fiber filters. Radioactivity on the filter was measured by
liquid scintillation counting.
[173] The cannabinoid CB1 activity of a candidate compound for use according to the
invention can also be determined by functional studies using CHO cells in which human

cannabinoid CB1 receptors are stably expressed. Adenylyl cyclase can be stimulated using
forskolin and measured by quantifying the amount of accumulated cyclic AMP. Concomitant
activation of CB1 receptors by CB1 receptor agonists (e.g., CP-55,940 or (i?)-WIN-55,212-2)
can attenuate the forskolin-induced accumulation of cAMP in a concentration-dependent
manner. This CB1 receptor-mediated response can be antagonized by CB1 receptor
antagonists. See, U.S. Patent Application No. US 2001/0053788 published on December 20,
2001.
[174] Samples rich in cannabinoid CB1 receptors and CB2 receptors, rat cerebellar
membrane fraction and spleen cells can be respectively used (male SD rats, 7-9 weeks old). A
sample (cerebellar membrane fraction: 50 (i.g/ml or spleen cells: l(xl07 cells/ml), labeled
ligand ([3 H]Win55212-2, 2 nM) and unlabeled Win55212-2 or a test compound can be
plated in round bottom 24 well plates, and incubated at 30°C for 90 min in the case of
cerebellar membrane fraction, and at 4°C for 360 min in the case of spleen cells. As the assay
buffer, 50 mM Tris solution containing 0.2% BSA can be used for cerebellar membrane
fraction, and 50 mM Tris-HBSS containing 0.2% BSA can be used for spleen cells. After
incubation, the samples are filtrated through a filter (Packard, Unifilter 24 GF/B) and dried.
A scintillation solution (Packard, Microsint-20) can be added, and the radioactivity of the
samples determined (Packard, Top count A9912V). The non-specific binding can be
determined by adding an excess Win55212-2 (1 uM), and calculating specific binding by
subtracting non-specific binding from the total binding obtained by adding the labeled ligand
alone. The test compounds can be dissolved in DMSO to the final concentration of DMSO of
0.1%. IC50 can be determined from the proportion of the specifically-bound test compounds,
and the Kj value of the test compounds can be calculated from IC50 and K H]WIN55212-2. See U.S. Patent No. 6,017,919.
[1751 in one embodiment, the IC50 for cannabinoid receptor binding is determined
according to the method of Devane et al. Science 258:1946-1949 (1992) and Devane et al. J.
Med. Chem. 35:2065 (1992). In this method, the ability of a compound to competitively
inhibit'the binding of an radiolabeled probe (e.g., 3H-HU-2430) is determined.
[1761 ui other embodiments, the IC50 of an inventive compound for the CB1 receptor is
determined according to any one of the above ligand binding assay methods. In another
embodiment, the IC50 is according to any assay method which studies binding at
physiological pH or physiologically relevant conditions. In another embodiment, the IC50 is

determined according to any assay method which studies binding at physiological pH and
ionic strength.
Cannabinoid CB2 Receptor Binding Assay.
[177] Methods of studying CB2 receptor binding are well known to one of ordinary skill in
the art. For instance, binding to the human cannabinoid CB2 receptor can be assessed using
the procedure of Showalter, et al., /. Pharmacol Exp Ther. 278(3), 989-99 (1996), with minor
modifications as taught for instance in U.S. Patent Application No. 20020026050 published
February 28,2002. Each of which is incorporated herein by reference.
[178] In other embodiments, the ICso of an inventive compound for the CB2 receptor is
determined according to any one of the above CB2 receptor ligand binding assay methods. In
another embodiment, the ICso is according to any assay method which studies binding at
physiological pH or physiologically relevant conditions. In another embodiment, the ICso is
determined according to any assay method which studies binding at physiological pH and
ionic strength.
Combinatorial chemical libraries.
[179] Recently, attention has focused on the use of combinatorial chemical libraries to assist
in the generation of new chemical compound leads. A combinatorial chemical library is a
collection of diverse chemical compounds generated by either chemical synthesis or
biological synthesis by combining a number of chemical "building blocks" such as reagents.
For example, a linear combinatorial chemical library such as a polypeptide library is formed
by combining a set of chemical building blocks called amino acids in every possible way for
a given compound length (i.e., the number of amino acids in a polypeptide compound).
Millions of chemical compounds can be synthesized through such combinatorial mixing of
chemical building blocks. For example, one commentator has observed that the systematic,
combinatorial mixing of 100 interchangeable chemical building blocks results in the
theoretical synthesis of 100 million tetrameric compounds or 10 billion pentameric
compounds (Gallop et al., /. Med. Chem. 37(9), 1233(1994)).
[180] Preparation and screening of combinatorial chemical libraries are well known to those
of skill in the art. Such combinatorial chemical libraries include, but are not limited to
diversomers such as hydantoins, benzodiazepines and dipeptides (Hobbs et al., Proc. Natl.
Acad. Sci. U.SA. 90,6909 (1993)), analogous organic syntheses of small compound libraries
(Chen et al., J. Amer. Chem. Soc. 116: 2661 (1994)), oligocarbamates (Cho, et al., Science

261, 1303(1993)), and/or peptidyl phosphorates (Campbell et aL, J. Org. Chem. 59: 658
(1994)), and small organic molecule libraries (see, e.g., benzodiazepines (Baum C&EN, Jan
18, 33(1993)), thiazolidinones and metathiazanones (U.S. Patent 5,549,974), pyrrolidines
(U.S. Patents 5,525,735 and 5,519,134), benzodiazepines (U.S. Patent 5,288,514), and the
like.
[181] Devices for the preparation of combinatorial libraries are commercially available (see,
e.g., 357 MPS, 390 MPS, Advanced Chem Tech, Louisville KY, Symphony, Rainin,
Woburn, MA, 433A Applied Biosystems, Foster City, CA, 9050 Plus, Millipore, Bedford,
MA).
[182] A number of well known robotic systems have also been developed for solution phase
chemistries. These systems include automated workstations like the automated synthesis
apparatus developed by Takeda Chemical Industries, LTD. (Osaka, Japan) and many robotic
systems utilizing robotic arms (Zymate IL Zymark Corporation, Hopkinton, Mass.; Orca,
HewletfPackard, Palo Alto, CA) which mimic (he manual synthetic operations performed by
a chemist. Any of the above devices are suitable for use with the present invention. The
nature and implementation of modifications to these devices so that they can operate as
discussed herein will be apparent to persons skilled in the relevant art. In addition, numerous
combinatorial libraries are themselves commercially available (see, e.g., ComGenex,
Princeton, N.J., Asinex, Moscow, Ru, Tripos, Inc., St. Louis, MO, ChemStar, Ltd., Moscow,
RU, 3D Pharmaceuticals, Exton, PA, Martek Biosciences, Columbia, MD, etc.).
High throughput FAAH Inhibition Assays of Chemical Libraries of Compounds
according to Formula I or n.
[183] The assays for compounds described herein are amenable to high throughput
screening. Preferred assays thus detect binding of the inhibitor to FAAH or the release of a
reaction product (e.g., fatty acid amide or ethanolamine) produced by the hydrolysis of a
substrate such as oleoylethanolamide or ananadamide. The substrate may be labeled to
facilitate detection of the released reaction products. High throughput assays for die
presence, absence, or quantification of particular reaction products are well known to those of
skill in die art Thus, for example, U.S. Patent 5,559,410 discloses high throughput screening
methods for proteins, and U.S. Patents 5,576,220 and 5,541,061 disclose high fliroughput
methods of screening for ligand/antibody binding.
[184] In addition, high throughput screening systems are commercially available (see, e.g.,
Zymark Corp., Hopkinton, MA; Air Technical Industries, Mentor, OH; Beckman
Instruments, Inc. Fullerton, CA; Precision Systems, Inc., Natick, MA, etc.). These systems

typically automate entire procedures including all sample and reagent pipetting, liquid
dispensing, timed incubations, and final readings of the microplate in detectors) appropriate
for the assay. These configurable systems provide high throughput and rapid start up as well
as a high degree of flexibility and customization. The manufacturers of such systems provide
detailed protocols the various high throughput. Thus, for example, Zymark Corp. provides
technical bulletins describing screening systems for detecting the modulation of gene
transcription, ligand binding, and the like.
Screening for anxiolytic activity
[185] One of ordinary skill in the art would appreciate that there are a number of animal
models available for assessing the antianxiety effects of a compound. Two
pharmacologically validated animal models of anxiety are the elevated zero maze test, and
the isolation-induced ultrasonic emission test. The zero maze consists of an elevated annular
platform with two open and two closed quadrants and is based on the conflict between an
animal's instinct to explore its environment and its fear of open spaces, where it may be
attacked by predators (Bickerdike, MJ. et al., Eur. J. Pharmacol, 271, 403-411 (1994);
Shepherd, J.K. et al., Psychopharmacology, 116, 56-64 (1994)). Clinically used anxiolytic
drugs, such as the benzodiazepines, increase the proportion of time spent in, and the number
of entries made into, the open compartments.
[186] A second test for an antianxiety compound is the ultrasonic vocalization emission
model, which measures the number of stress-induced vocalizations emitted by rat pups
removed from their nest (Insel, T.R. et al., Pharmacol Biochem. Behav., 24, 1263-1267
(1986); Miczek, K.A. et al., Psychopharmacology, 121, 38-56 (1995); Winslow, J.T. et al.,
Biol Psychiatry, 15,745-757 (1991).
[187] A large number of animal models have been developed in the attempt to predict the
anxiolytic activity of novel compounds in man. Many of these paradigms evaluate animal
behavior in a so-called "conflict" situation, i.e. a behavioral response is simultaneously under
the influence of two opposing motivational states such as approach and avoidance tendencies.
Probably the best known model is the conditioned punishment conflict paradigm in which
animals are trained to voluntarily exhibit a certain response (e.g. pressing a lever) in order to
receive a reward (e.g. food for a hungry animal). Once the animals exhibit a constant rate of
lever-press responding, then short periods are introduced (usually signaled by visual or
acoustic signals) during which lever pressing is simultaneously rewarded by food and
punished by mild electrical foot shock. Animals exhibit a markedly reduced response rate

during these conflict periods, which are also characterized by various overt signs of
emotionality. The characteristic effect of benzodiazepine receptor agonists, for example the
anxiolytic diazepam, is the disinhibition of punished behavior (resulting in an increase in the
rate of responding under punishment) at doses that fail to disrupt unpunished responding.
Furthermore, these same active drugs produce an anxiolytic-like effect in the absence of
actual punishment, i.e. when the rate of lever pressing is reduced by conditioned fear of
punishment. The conflict task does not require conditioned behavioral responses: naive
thirsty animals can be offered the opportunity to drink, with drinking punished via contact
with an electrified spout. Such punishment-suppressed drinking is disinhibited dose-
dependently by benzodiazepine receptor agonists (e.g., diazepam). Exploratory activity can
likewise be decreased by contingent punishment and released by treatment with known
anxiolytics. Conflict models without punishment are based on the presence of the natural
opposing motivational states, on the one hand the tendency to explore and, on the other hand,
fear of a novel environment (e.g. dark-light chamber task, elevated plus-maze, consumption
of unfamiliar food or normal food in an unfamiliar environment, social interaction between
animals unfamiliar with each other). While it is obvious to ascribe the behavioral
disinhibitory effect of benzodiazepine receptor agonism in these experimental situations to an
anxiolytic-like action, their effect can also be interpreted as a general reduction of the
influence of aversive factors or even to an impaired ability to withhold innate or conditioned
responses. An anti-frustration effect resulting from benzodiazepine receptor agonism is
suggested by the increase of responding which is maintained by response-contingent reward
in the situation in which the reward is reduced or omitted. Electrical stimulation of the
periaqueductal gray area of the midbrain via chronically implanted electrodes in animals is
aversive and elicits a number of emotional reactions; benzodiazepine receptor agonists
increase the aversive threshold. States of acute anxiety characterised by behavioral and
physiological symptoms (cardiovascular, endocrine) can be induced by chemicals known to
be anxiogenic in man, e.g. convulsants such as pentylenetetrazol, inverse agonists at the
benzodiazepine receptor agonists administered in subconvulsive doses, or even abrupt drug
withdrawal after chronic treatment with high doses of sedatives. Ultrasonic distress cries by
rat pups acutely separated from their mothers are decreased by benzodiazepine receptor
agonists.
Screening for Antidepressant Activity

[188] Animal models for depression are also well known to those of ordinary skill in the art.
For instance, the effect of the compound of the invention in the treatment of depression can
be tested in the model of chronic mild stress induced anhedonia in rats. This model is based
on the observation that chronic mild stress causes a gradual decrease in sensitivity to rewards,
for example consumption of sucrose, and that this decrease is dose-dependently reversed by
chronic treatment with antidepressants. The method has previously been described and more
information with respect to the test appears from Willner, Paul, Psychopharmacology, 1997,
134,319-329.
11891 Another test for antidepressant activity is the forced swimming test (Nature 266, 730-
732, 1977) In this test, animals are administered an agent preferably by the intraperitoneal
route or by the oral route 30 or 60 minutes before the test. The animals are placed in a
crystallizing dish filled with water and the time during which they remain immobile is
clocked. The immobility time is then compared with that of the control group treated with
distilled water, Imipramine 25 mg/kg. can beused as the positive control. The antidepressant
compounds decrease the immobility time of the mice thus immersed.
[190] Another test for antidepressant activity is the caudal suspension test on the mouse
(Psychopharmacology, 85, 367-370, 1985) In this test, animals are preferably treated with
the study compound by the intraperitoneal route or by the oral route 30 or 60 minutes before
the test The animals are then suspended by the tail and their immobility time is automatically
recorded by a computer system. The immobility times are then compared with those of a
control group treated with distilled water. Imipramine 25 mg/kg can be used as the positive
control. Antidepressant compounds decrease the immobility time of the mice.
[191] Another test for screening antidepressants is the DRL-72 TEST. This test, carried out
according to the protocol of Andrews et al ["Effects of imipramine and mirtazapine on
operant performance in rateF-Drug Development Research 32, 58-66 (1994)], gives an
indication of antidepressant-like activity. See also U.S. Patent No. 6,403,573.
[192] Additional animal models for screening are well known to one of ordinary skill in the
art. For instance, see U.S. Patent No. 5,952,315.
Methods for Screening for Anticonvulsant and Antiepilepsy Activity
[193] Animals models are available to one of ordinary skill in the art for studying
anticonvulsant activity of test compounds. See for instance, U.S. Patent No. 6,309,406 and
U.S. Patent No. 6,326,156 which describe methods for performing such tests. In addition, the
compounds may be administered to humans suffering from epilepsy or other convulsive

conditions and the effect on the frequency or severity or onset of convulsions clinically
assessed.
Methods for Screening for Sleep Promoting or Soporific Properties.
[194] Inhibition of FAAH has been reported to induce sleep in test animals (U.S. Patent No.
6,096,784). Methods for studying sleep inducing compounds are well known to one of
ordinary skill in the art. In particular, methods for testing the ability of a test FAAH
inhibitory compound to induce sleep or treat insomnia are also disclosed in U.S. Patent No.
6,096,784 and U.S. Patent No. 6,271,015. Most obviously, the compounds can be
administered to a test animal (e.g., rat or mouse) or a human and the subsequent time (e.g.,
onset, duration) spent sleeping (e.g., eyes closed, motor quiescence) can be monitored. See
also WO 98/24396.
Methods for Screening Compounds which Induce Catalepsy or Affect Motor Activity
[195] Methods for screening FAAH inhibitors which induce catalepsy are also well known
to one of ordinary in the art. See Quistand et al. in Toxicology and Applied Pharmacology
173:48-55 (2001). See Cravatt et al. Proc. Natl. Acad. Sci. U.S.A. 98:9371-9376 (2001).
Methods for Screening Compounds for Antinociceptive Activity.
[196] Methods for screening FAAH inhibitors for an antinociceptive effect are well known
to one of ordinary in the art. For instance, the test compounds can be administered to the
subject animals in the mouse hot-plate test and the mouse formalin test and the nociceptive
reactions to thermal or chemical tissue damage measured. See also U.S. Patent No.
6,326,156 which teaches methods of screening for antinociceptive activity. See Cravatt et al.
Proc. Natl Acad. Sci. U.S.A. 98:9371-9376 (2001).
Methods for assessing the effect of a FAAH inhibitor on an Appetite.
[197] Compounds of the invention can be administered to an animal to determine whether
they affect food intake and body weight, body fat, appetite, food seeking behavior, or
modulate modulator fatty acid oxidation. Method of conducting such tests are known to one
of ordinary skill in the art. For instance, see U.S. Patent Application No. 60/336,289
assigned to the same assignee and herein incorporated by reference in its entirety.
[198] Animals can be, for example, obese or normal guinea pigs, rats, mice, or rabbits.
Suitable rats include, for example, Zucker rats. Suitable mice include, for example, normal

mice, ALS/LtJ, C3.SW-#-2b/SnJ, (NON/LtJ x NZO/HLQF1, NZO/HU, ALR/LtJ, NON/LtJ,
KK.Cg-AALR/LtJ, NON/LtJ, KK.Cg-Ay/J, B6.HRS(BKS)- B6.V-Lepob, BKS.Cg-m +/+ Lep% and C57BL/6J with Diet Induced Obesity.
[199] Administration of an appropriate amount the candidate compound may be by any
means known in the art such as, for example, oral or rectal, parenteral such as, for example,
intraperitoneal, intravenous, subcutaneous, subdermal, intranasal, or intramuscular.
Preferably administration may be intraperitoneal or oral. An appropriate effective amount of
the candidate compound may be determined empirically as is known in the art.
[200] Other methods of assessing appetitive behavior are known to one of ordinary skill in
the art. For instance, Maruani et al. (U.S. Patent No. 6,344,474 ) teach two such assays. One
method of assessing the effect on appetite behavior is to administer a FAAH inhibitor to a rat
and assess its effect on the intake of a sucrose solution. This method is taught in W. C.
Lynch et al., Physiol Behav., 1993,54,877-880. Male Sprague-Dawley rats weighing 190 to
210 g are under a normal light cycle (from 7 am to 7 pm) and receive water and food ad
libitum. For 6 days, between 11 am and 3 pm, the food and the water bottles are withdrawn
and the rats are given a 5% sucrose solution to drink. Rats drinking less than 3 g of sucrose
solution are eliminated. On the seventh day the test is carried out according to the following
procedure: 9 am: withdrawal of food, 10 am: administration of the inhibitor or vehicle to the
test animals; 11 am=T0: introduction of bottles containing a weighed sucrose solution, T0+1
hour, T0+2 hours, T0+3 hours, T0+4 hours: measurement of the sucrose consumption by
weighing of the bottles. Followed by comparison of the experimental and control groups'
intake of the sucrose solution.
[201] In another test, the effect of a FAAH inhibitor on the consumption of an alcohol
solution can be assessed in mice. For instance, male C 57 BL 6 mice are isolated on the day
of their arrival in an animal housing under a reverse cycle (night from 10 am to 10 pm) with 2
bottles filled with water. After 1 week, one of the bottles of water is replaced with a bottle
filled with a 10% alcohol solution for 6 hours of the test. Each day, 30 minutes before the
bottle of alcohol is introduced, the mice are treated with a FAAH inhibitor. The amounts of
alcohol and water consumed are measured after 6 hours. The test is repeated for 4 days.
[202] The results for an experimental and a control or vehicle are compared.
[203] Methods for Screening for Antipsychotic or Antischizophrenic or Dopamine-
Modulating Activity

[204] Without being wed to theory, it is believed that excessive dopamine transmission in
the CNS may contribute to schizophrenia and other mental disorders. Approximately one-
third of all schizophrenic patients manifest obvious dopamine transmitter and/or receptor
increases. Others who do not overtly manifest this abnormality still show improvement of
symptoms with the pharmacological blockade of dopamine receptors. These dopamine
receptor antagonists ultimately result in overall reductions in dopamine concentrations due to
depolarization block and dopamine receptor antagonism. Thus, malfunction of neural circuits,
many of which dopamine has a direct and/or indirect role in activating, appears to be
involved in schizophrenic symptoms. As has been shown above, blocking dopamine
receptors in subcortical areas of the brain substantially reduces schizophrenic symptoms.
Generalized reduction of dopamine production in these areas provides similar relief to
patients suffering from this disease. Cannabinoids have been found to modulate dopamine
activity in the CNS.
[205] Methods for screening compound for their effects on dopaminergic transmission and
systems in the CNS are well known to one of ordinary skill in the art. Methods for conducting
clinical trials of candidate agents in any of the above neurological diseases, disorders and
conditions are well known to one of ordinary skill in the art.
Glaucoma
[206] Methods of measuring the intraocular pressure of the eye with respect to the
treatment of glaucoma are routine in the medical arts and may be readily and safely
performed used human or animal subjects. The effect of a FAAH inhibitor on subject eye
pressure can be readily assessed by applying the compound directly to the eye and monitoring
eye pressure over the next several hours or day. The alternate eye may be used as a control.
Alternatively the FAAH inhibitor may be given systemically and another vehicle treated
subject used as the control.
Methods of Use, Pharmaceutical Compositions, and their Administration
Methods of Use
Anxiety and Anxiety related disorders.
[207] In some embodiments, the FAAH inhibitory compounds, including the compounds of
Formula I and H, and their pharmaceutical compositions and methods of administering them
are useful in treating anxiety and anxiety disorders or conditions. The compounds and
compositions are useful, for example in treating anxiety, clinical anxiety, panic disorder,

agoraphobia, generalized anxiety disorder, specific phobia, social phobia, obsessive-
compulsive disorder, acute stress disorder, and post-traumatic stress disorder; and adjustment
disorders with anxious features, anxiety disorders due to general medical conditions,
substance-induced anxiety disorders, and the residual category of anxiety disorder not
otherwise specified. The treatment may be prophylactic or therapeutic. The treatment may
be administered to a human subject. The compounds may be used in otherwise healthy
individuals who are not otherwise in need of any pharmaceutical intervention for a disease or
condition such as insomnia or for pain relief.
[208] In some embodiments, the compounds methods, and compositions of the invention
may also be administered to treat anxiety in mammals, including cats, dogs, and humans. In
some embodiments, the compounds may be used in otherwise healthy individuals who are not
in need of pharmaceutical interventions for any other disease or disorder than anxiety or an
anxiety disorder. In some embodiments, the subject is not otherwise in need of a FAAH
inhibitor.
[209] The compounds and compositions of the invention may be administered solely for the
purposes of reducing the severity or frequency of anxiety or an anxiety disorder.
[210] Preferred inhibitors for such uses are UCM532 and UCM597.
Depression and Depressive disorders
[211] In some embodiments, the FAAH inhibitory compounds of Formula I and n, their
pharmaceutical compositions and methods of administering them are useful in treating
depression and depressive disorders or conditions. The compounds and compositions are
useful, for example in treating major depressive disorders (unipolar depression), dysthymic
disorders (chronic, mild depression), and bipolar disorders (manic-depression). The
depression may be clinical or subclinical depression. The treatment may be prophylactic or
therapeutic. The treatment may be administered to a human subject. The compounds may be
used in otherwise healthy individuals who are not otherwise in need of any pharmaceutical
intervention for a disease such as insomnia or for pain relief.
[212] In some embodiments, the compounds methods, and compositions of the invention
may also be administered to treat depression in mammals, including cats, dogs, and humans.
In some embodiments, the compounds may be used in otherwise healthy individuals who are
not in need of pharmaceutical interventions for any other disease or disorder than depression
or a depressive disorder, hi some embodiments, the subject is not otherwise in need of a
FAAH inhibitor.

[213] The compounds and compositions of the invention may be administered solely for the
purposes of reducing the severity or frequency of depression or a depressive disorder.
[214] Preferred inhibitors for such uses are UCM532 and UCM597.
Seizure Disorders
[215[ In some embodiments, the FAAH inhibitory compounds, their pharmaceutical
compositions and methods of administering them are useful in treating epilepsy and
convulsive disorders or seizures. The treatment may be prophylactic or therapeutic. The
treatment may be administered to a human subject. The compounds may be used in otherwise
healthy individuals who are not otherwise in need of any pharmaceutical intervention for a
disease such as insomnia or pain relief.
[216] In some embodiments, the compounds methods, and compositions of the invention
may also be administered to treat such diseases and disorders in mammals, including cats,
dogs, and humans. In some embodiments, the compounds may be used in otherwise healthy
individuals who are not in need of pharmaceutical interventions for any other disease or
disorder than a seizure disorder. In some embodiments, the subject is not otherwise in need
of a FAAH inhibitor.
[217] The compounds and compositions of the invention may be administered solely for the
purposes of reducing the severity or frequency of convulsions or seizures.
[218] Preferred inhibitors for such uses are UCM532 and UCM597.
Use of FAAH Inhibitors to Control of Appetite and Treatment of Appetite Disorders
[219] In some embodiments, the invention provides pharmaceutical compositions and
methods of using FAAH inhibitory compound to reduce appetite(s), reduce body fat and for
treating or preventing obesity or overweight in a mammal and for preventing or treating the
diseases associated with these health conditions. In one aspect of the instant invention,
methods are provided for reducing appetite, body fat or body weight, or for treating or
preventing obesity or overweight, or for reducing food intake, or treating an appetency
disorder in a mammal by administering to the mammal a FAAH inhibitor, including
inhibitors according to Formula I and Formula H In a further embodiment, the inhibitor is
administered in a combination therapy with oleoylethanolamide (OEA) or another fatty acid
alkanolamide compound, or a homologue or analog of oleylethanolamide or the fatty acid
alkanolamide compound, which reduces appetite or food consumption and is subject to
hydrolysis by FAAH.

[220] In some embodiments, the FAAH inhibitor is administered to a subject in amounts
sufficient to reduce body fat, body weight, or prevent body fat or body weight gain or to
reduce appetite(s). In another aspect of the invention, pharmaceutical compositions are
provided which comprise a first compound which is a FAAH inhibitor and a second
compound which is oleylethanolamide or a fatty acid alkanolamide compound, or a
homologue or analog of oleylethanolamide or the fatty acid alkanolamide compound which
reduces appetite or which has an effect to reduce appetite. In other aspects, the invention is
drawn to such pharmaceutical compositions and their methods of use to reduce or control
appetite or to treat appetite disorders.
[221] In some aspects, the invention provides method of treating an appetency disorder
comprising administration of a first compound which is a FAAH inhibitor and a second
compound which is a fatty acid alkanolamide compound, homologue or OEA analog which is
not a significant antagonist of the cannabinoid CB1 receptor and is administered in an amount
which does not by itself significantly activate or inhibit the CB1 receptor. In another aspect
of the invention, pharmaceutical compositions are provided which comprise a first compound
which is a FAAH inhibitor and a second compound which is oleylethanolamide (OEA) or a
fatty acid alkanolamide compound, or a homologue or analog of oleylethanolamide or the
fatty acid alkanolamide compound, which is not a significant CB1 cannabinoid receptor
antagonist and which reduces appetite or which has an effect to reduce appetite which is not
substantially mediated by binding of the second compound to the CB1 cannabinoid receptor.
In other aspects, the invention is drawn to such pharmaceutical compositions and then-
methods of use to reduce or control appetite and to treat appetite disorders.
[222] Preferred inhibitors for such uses are UCM532 and UCM597.
Schizophrenia and dopamine related disorders
[223] Is some embodiments, the FAAH inhibitory compounds according to Formula I or
Formula n, their pharmaceutical compositions and methods of administering them are useful
in treating schizophrenia and dopamine related disorders. The treatment may be prophylactic
or therapeutic. The treatment may be administered to a human subject The compounds may
be used in otherwise healthy individuals who are not otherwise in need of any pharmaceutical
intervention for a disease such as insomnia or hyperalgesia. In some embodiments, the
compounds may be used in otherwise healthy individuals who are not in need of
pharmaceutical interventions for any other disease or disorder than a seizure disorder. In
some embodiments, the subject is not otherwise in need of a FAAH inhibitor.

[224] The compounds and compositions of the invention may be administered solely for the
purposes of reducing the severity or frequency of the schizophrenia or dopamine related
disorder. They may be administered to reduce paranoid ideation and flat affect.
[225] Preferred inhibitors for such uses are UCM532 and UCM597.
Use to Induce Sleep
[226] In some embodiments, the compounds of Formula I and II may be administered to
induce or promote sleep in a mammalian subject. The treatment may be prophylactic or
therapeutic. The treatment may be administered to a human subject. The compounds and
compositions of the invention may be administered solely for the purposes of reducing the
severity or frequency or extent of sleeplessness.
[227] Another aspect of the invention is directed to a method for inhibiting oleamide
hydrolysis by FAAH. The method employs the act of contacting or combining the FAAH
with an inhibitor. The inhibitor is according to Formula I or Formula n.
[228] Another aspect of the invention is directed to a method for inducing sleep within an
oleamide sensitive animal. More particularly, this aspect of the invention is directed to the
administration to an oleamide sensitive animal of an effective dose of an agonist of oleamide
hydrolase. Preferred inhibitors for such uses are UCM532 and UCM597.
Control of Pain
[229] In some embodiments, the compounds of Formula I and II may be administered to
alleviate pain in a subject The treatment may be prophylactic or therapeutic. The treatment
may be administered to a human subject. The compounds and compositions of the invention
may be administered solely for the purposes of reducing the severity or frequency or extent of
pain. The treatment may be administered in a combination therapy with another pain reliever
or antiinflammatory agent.
Glaucoma
[230] In some embodiments, FAAH inhibitors may be administered to treat or prevent
glaucoma or to reduce intraocular eye pressure. In some embodiments, the compounds may
be given systemically. In other embodiments, the FAAH inhibitors are direct applied to the
surface of the eye (e.g., via eye drops).
Pharmaceutical Compositions.

[231] Another aspect of the present invention provides pharmaceutical compositions which
comprise compounds of the invention and a pharmaceutically acceptable carrier.
[2321 The pharmaceutical compositions of the present invention comprise a compound of
the instant invention as an active ingredient or a pharmaceutically acceptable salt thereof, and
may also contain a pharmaceutically acceptable carrier and optionally other therapeutic
ingredients. In some embodiments, the compositions comprise a compound of Formula I or
Formula E.
[233] The compositions include compositions suitable for oral, rectal, topical, parenteral
(including subcutaneous, intramuscular, and intravenous), ocular (ophthalmic), pulmonary
(nasal or buccal inhalation), or nasal administration, although the most suitable route in any
given case will depend in part on the nature and severity of the conditions being treated and
on the nature of the active ingredient. An exemplary route of administration is the oral route.
The compositions may be conveniently presented in unit dosage form and prepared by any of
the methods well-known in the art of pharmacy.
[234] In practical use, the compounds of the invention can be combined as the active
ingredient in intimate admixture with a pharmaceutical carrier according to conventional
pharmaceutical compounding techniques. The carrier may take a wide variety of forms
depending on the form of preparation desired for administration, e.g., oral or parenteral
(including intravenous). In preparing the compositions for oral dosage form, any of the usual
pharmaceutical media may be employed, such as, for example, water, glycols, oils, alcohols,
flavoring agents, preservatives, coloring agents and. the like in the case of oral liquid
preparations, such as, for example, suspensions, elixirs and solutions; or carriers such as
starches, sugars, microcrystalline cellulose, diluents, granulating agents, lubricants, binders,
disintegrating agents and the like in the case of oral solid preparations such as, for example,
powders, hard and soft capsules and tablets, with the solid oral preparations being preferred
over the liquid preparations.
[235] Because of their ease of administration, tablets and capsules represent the most
advantageous oral dosage unit form in which case solid pharmaceutical carriers are obviously
employed. If desired, tablets may be coated by standard aqueous or nonaqueous techniques.
Such compositions and preparations can contain at least 0.1 percent of active compound. The
percentage of active compound in these compositions may, of course, be varied and may
conveniently be between about 2 percent to about 60 percent of the weight of the unit. The
amount of active compound in such therapeutically useful compositions is such that a

therapeutically effective dosage will be obtained. The active compounds can also be
administered intranasally as, for example, liquid drops or spray.
[236] The tablets, pills, capsules, and the like may also contain a binder such as gum
tragacanth, acacia, corn starch or gelatin; excipients such as dicalcium phosphate; a
disintegrating agent such as corn starch, potato starch, alginic acid; a lubricant such as
magnesium stearate; and a sweetening agent such as sucrose, lactose or saccharin. When a
dosage unit form is a capsule, it may contain, in addition to materials of the above type, a
liquid carrier such as a fatty oil.
[237] Various other materials may be present as coatings or to modify the physical form of
the dosage unit. For instance, tablets may be coated with shellac, sugar or both. A syrup or
elixir may contain, in addition to the active ingredient, sucrose as a sweetening agent, methyl
and propylparabens as preservatives, a dye and a flavoring such as cherry or orange flavor.
To prevent breakdown during transit through the upper portion of the GI tract, the
composition may be an enteric coated formulation.
[238] With respect to formulations with respect to any variety of routes of administration,
methods and formulations for the administration of drugs are disclosed in Remington's
Pharmaceutical Sciences, 17th Edition, (Gennaro et al. Eds., Mack Publishing Co., 1985).
Remington's Pharmaceutical Sciences, Gennaro AR ed. 20th edition, 2000: Williams &
WilkinsPA,USA.
Administration
[239] The compounds of the invention may also be administered parenterally. Solutions or
suspensions of these active compounds can be prepared in water suitably mixed with a
surfactant such as hydroxypropylcellulose. Dispersions can also be prepared in glycerol,
liquid polyethylene glycols and mixtures thereof in oils. Under ordinary conditions of storage
and use, these preparations contain a preservative to prevent the growth of microorganisms.
[240] The pharmaceutical forms suitable for injectable use include sterile aqueous solutions
or dispersions and sterile powders for the extemporaneous preparation of sterile injectable
solutions or dispersions. In all cases, the form must be sterile and must be fluid to the extent
that easy syringability exists. It must be stable under the conditions of manufacture and
storage and must be preserved against the contaminating action of microorganisms such as
bacteria and fungi. The carrier can be a solvent or dispersion medium containing, for
example, water, ethanol, polyol (e.g. glycerol, propylene glycol and liquid polyethylene
glycol), suitable mixtures thereof, and vegetable oils.

[241] The compounds of the invention can be effective over a wide dosage range. For
example, in the treatment of adult humans, dosages from about 10 to about 1000 mg, about
100 to about 500 mg or about 1 to about 100 mg may be needed. Doses of the 0.05 to about
100 mg, and more preferably from about 0.1 to about 100 mg, per day may be used. A most
preferable dosage is about 0.1 mg to about 70 mg per day. In choosing a regimen for patients,
it may frequently be necessary to begin with a dosage of from about 2 to about 70 mg per day
and when the condition is under control to reduce the dosage as low as from about 0.1 to
about 10 mg per day. For example, in the treatment of adult humans, dosages from about 0.05
to about 100 mg, preferably from about 0.1 to about 100 mg, per day may be used. The exact
dosage will depend upon the mode of administration, on the therapy desired, form in which
administered, the subject to be treated and the body weight of the subject to be treated, and
the preference and experience of the physician or veterinarian in charge.
[242] Generally, the compounds of the present invention can be dispensed in unit dosage
form comprising preferably from about 0.1 to about 100 mg of active ingredient together with
a pharmaceutically acceptable carrier per unit dosage. Usually, dosage forms suitable for oral,
nasal, pulmonary or transdermal administration comprise from about 0.001 mg to about 100
mg, preferably from about 0.01 mg to about 50 mg of the compounds admixed with a
pharmaceutically acceptable carrier or diluent. For storage and use, these preparations
preferably contain a preservative to prevent the growth of microorganisms.
[243] Administration of an appropriate amount the candidate compound may be by any
means known in the art such as, for example, oral.or rectal, parenteral, intraperitoneal,
intravenous, subcutaneous, subdermal, intranasal, or intramuscular, in some embodiments,
administration is transdermal. An appropriate amount or dose of the candidate compound
may be determined empirically as is known in the art. An appropriate or therapeutic amount
is an amount sufficient to effect a loss of body fat or a loss in body weight in the animal over
time. The candidate compound can be administered as often as required to effect a loss of
body fat or loss in body weight, for example, hourly, every six, eight, twelve, or eighteen
hours, daily, or weekly
[244] Formulations suitable for oral administration can consist of (a) liquid solutions, such
as an effective amount of the packaged nucleic acid suspended in diluents, such as water,
saline or PEG 400; (b) capsules, sachets or tablets, each containing a predetermined amount
of the active ingredient, as liquids, solids, granules or gelatin; (c) suspensions in an
appropriate liquid; and (d) suitable emulsions. Tablet forms can include one or more of
lactose, sucrose, mannitol, sorbitol, calcium phosphates, corn starch, potato starch,

microcrystalline cellulose, gelatin, colloidal silicon dioxide, talc, magnesium stearate, stearic
acid, and other excipients, colorants, fillers, binders, diluents, buffering agents, moistening
agents, preservatives, flavoring agents, dyes, disintegrating agents, and pharmaceutically
compatible carriers. Lozenge forms can comprise the active ingredient in a flavor, e.g.,
sucrose, as well as pastilles comprising the active ingredient in an inert base, such as gelatin
and glycerin or sucrose and acacia emulsions, gels, and the like containing, in addition to the
active ingredient, carriers known in the art.
[245] Injection solutions and suspensions can be prepared from sterile powders, granules,
and tablets of the kind previously described. Formulations suitable for parenteral
administration, such as, for example, by intraarticular (in the joints), intravenous,
intramuscular, intradermal, intraperitoneal, and subcutaneous routes, include aqueous and
non-aqueous, isotonic sterile injection solutions, which can contain antioxidants, buffers,
bacteriostats, and solutes that render the formulation isotonic with the blood of the intended
recipient, and aqueous and non-aqueous sterile suspensions that can include suspending
agents, solubilizers, thickening agents, stabilizers, and preservatives.
[246] With respect to transdermal routes of administration, methods for transdermal
administration of drugs are disclosed in Remington's Pharmaceutical Sciences, Gennaro AR
ed. 20th edition, 2000: Williams & Wilkins PA, USA. Dermal or skin patches are a preferred
means for transdermal delivery of the compounds of the invention. Patches preferably
provide an absorption enhancer such as DMSO to increase the absorption of the compounds.
Other methods for transdermal drug delivery are disclosed in U.S. Patents No. 5,962,012,
6,261,595, and 6,261,595. Each of which is incorporated by reference in its entirety.
[247] Preferred patches include those that control the rate of drug delivery to the skin.
Patches may provide a variety of dosing systems including a reservoir system or a
monolithic system, respectively. The reservoir design may, for example, have four layers: the
adhesive layer that directly contacts the skin, the control membrane, which controls the
diffusion of drug molecules, the reservoir of drug molecules, and a water-resistant backing.
Such a design delivers uniform amounts of the drug over a specified time period, the rate of
delivery has to be less than the saturation limit of different types of skin.
[248] The monolithic design, for example, typically has only three layers: the adhesive
layer, a polymer matrix containing the compound, and a water-proof backing. This design
brings a saturating amount of drug to the skin. Thereby, delivery is controlled by the skin.
As the drug amount decreases in the patch to below the saturating level, the delivery rate
falls.

[249] Compounds of the invention may be used in combination with other compounds of
the invention or with other drugs that may also be useful in the treatment, prevention,
suppression of a neurological or psychological disorder. In one embodiment, the second drug
is not a FAAH inhibitor and is directed toward the same disorder as the fatty acid amide
inhibitor. Such other drugs may be administered, by a route and in an amount commonly
used therefor, contemporaneously or sequentially with a compound of the invention. When a
compound of the invention is used contemporaneously with one or more other drugs, a
pharmaceutical composition in unit dosage form containing such other drugs and the
compound is preferred. When used in combination with one or more other active ingredients,
the compound of the present invention and the other active ingredients may be used in lower
doses man when each is used singly. Accordingly, the pharmaceutical compositions of the
present invention include those that contain one or more other active ingredients, in addition
to the compounds disclosed above. For example, a FAAH inhibitor according to Formula I
or Formula II may be formulated with an anxiolytic agent which is not a FAAH inhibitor. For
example, a FAAH inhibitor according to Formula I or Formula II may be formulated with an
antidepressant.
[250] In the pharmaceutical compositions of the present invention for oral, sublingual,
subcutaneous, intramuscular, intravenous, transdermal, local or rectal administration, the
active principle, by itself or in association with another active principle, can be administered
to animals and humans in unit forms of administration mixed with conventional
pharmaceutical carriers. The appropriate unit forms of administration include oral forms such
as tablets, gelatin capsules, powders, granules and solutions or suspensions to be taken orally,
sublingual and buccal forms of administration, aerosols, implants, subcutaneous,
intramuscular, intravenous, intranasal or intraocular forms of administration and rectal forms
of administration.
[251] In other embodiments, the pharmaceutical compositions of the present invention, the
active principle or active principles are generally formulated in dosage units. The dosage unit
contains from 0.5 to 1000 mg, advantageously from 1 to 500 mg and preferably from 2 to 200
mg of FAAH inhibitor per dosage unit for daily administration.
[252] When used to treat glaucoma, direct application to the eye is preferred. Ocular carrier
formulations for such ocular application are taught in Remington's Pharmaceutical Sciences,
Gennaro AR ed. 20th edition, 2000: Williams & Wilkins PA, USA.
[253] The following examples are provided for illustrative purposes, and are not intended
to limit the scope of the invention as claimed herein. Any variations in the exemplified

articles and/or methods which occur to the skilled artisan are intended to fall within the scope
of the present invention.
EXAMPLES
Example 1: Subjects
[254J We used male and female Wistar rats (200-350 g) and male Swiss mice (20 g). All
procedures met the National Institutes of Health guidelines for the care and use of laboratory
animals, and those of the Italian Ministry of Health (D.L. 116/92). We prepared primary
cultures of cortical neurons from 18-day-old Wistar rat embryos, and maintained them as
described (Stella, N. et al, Eur. J. Pharmacol, 425,189-196 (2001)); we purchased human
astrocytoma cells from American Type Culture Collection (Manassas, VA).
Example 2: Chemicals
[255] Anandamide and related lipids were synthesized in the laboratory (Giuffrida, A. et al,
Anal. Biochem., 280, 87-93 (2000)). SR141716A (rimonabant) was provided by RBI
(Natick, MA) as part of the Chemical Synthesis Program of the National Institutes of Health;
AM404 was from Tocris (Avonmouth, UK) and other drugs from Sigma (St Louis, MO).
All chemicals necessary for the preparation of inhibitors were from Aldricb,
Example 3: Synthesis of inhibitors
UK] n-Butylcarbamic acid 4-benzyloxyphenyl ester (UCM532) (4) and 4-
fluorophenylcarbamic acid 4-benzyloxyphenyl ester (8) were obtained by treatment of 4-
benzyloxyphenol with n-butylisocyanate, and 4-fluorophenylisocyanate, respectively, with a
catalytic amount of triethylamine in remixing toluene. Similarly, cyclohexylcarbamic acid
biphenyl-3-yl ester (5), cyclohexylcarbamic acid 5-phenylpentyl ester (7), and
cyclohexylcarbamic acid 3'-carbamoyIbiphenyl-3-yl ester (UCM597) (6) were synthesized by
reacting cyclohexylisocyanate with 3-phenylphenol, 5-phenylpentan-l-ol, and 3-
hydroxybiphenyl-3-carboxylic acid amide, respectively. The latter reactant was prepared as
follows: 3-bromobenzoic acid amide, obtained by reaction of 3-bromobenzonitrile and
sodium perborate, was coupled with methoxyphenylboronic acid to give 3'-methoxybiphenyl-
3-carboxylic acid amide, which was hydrolized with BBr3 to generate the desired 3 -
hydroxybiphenyl-3-carboxylic acid amide.

3-Bromobenzoic acid amide. To a solution of 3-bromobenzonitrile (0.91 g, 5 mmol) in
dioxane (19 mL), NaB03-4H20 (2.12 g, 13.78 mmol) and H20 (19 mL) were added. The
mixture was stirred at 80 °C 16 hours (h), cooled, added of H20 and extracted with CH2C12.
The combined organic layers were dried over Na2S04 and evaporated. Purification of the
residue by column chromatography (hexane/EtOAc 2:8) and recrystallization gave the
desired product as colorless tablets. Yield 80% (EtOH). Mp: 156-7 °C (lit. 156 °C) (Pearson,
D.E. et at, J. Org. Chem., 28:3147-3149, (1963)). MS (EI): m/z 199 (M+); 183 (100%).
3'-Methoxybiphenyl-3-carboxylic acid amide. To a stirred mixture 3-bromobenzoic
acid amide (0.76 g; 3.8 mmol) and toluene (25 mL), Pd(PPh3)4 (0.180 g; 0.16 mmol), a
solution of Na2C03 (2.543 g; 24 mmol) in H20 (10 mL), and one of 3-methoxyphenylboronic
acid (1.132 g; 7.45 mmol) in EtOH (10 mL) were added. The mixture was refiuxed for 1 h
under vigorous stirring, cooled, and the acqueous phase extracted with AcOEt. The combined
organic layers were dried over Na2S04 and concentrated. Purification of the residue by
column chromatography (cyclohexane/EtOAc 1:1 then 4:6) and recrystallization gave the
desired product (0.64 g) as white solid. Yield 74%. Mp: 138-40 °C (EtOH). MS (EI): m/z
227 (M+, 100%). »H NMR (CDC13): 5 3.88 (s, 3H); 5.71 (br s, 1H); 6.11 (br s, 1H); 6.94 (m,
1H); 7.18 (m, 2H); 7.39 (t, 1H); 7.53 (t, 1H); 7.77 (m, 2H); 8.05 (t, 1H) ppm. IR (Nujol):
3327,3148,1676,1640,1613,1584 cm-».
3'-Hydroxybiphenyl-3-carboxylic acid amide. To a stirred, cooled (0 °C) solution of
3'-methoxybiphenyl-3-carboxylic acid amide (0.57 g; 2.5 mmol) in dry CH2C12 (28 mL),
under N2 atmosphere, a 1M solution of BBr3 in CH2C12 (6.4 mL) was added. The mixture was
stirred at room temperature for 1 h, quenched with 2N Na2C03 and extracted with AcOEt
The combined organic layers were washed with brine,, dried over Na2S04 and concentrated.
Purification of residue by column chromatography (cyclohexane/EtOAc 2:8) gave the desired
product as an amorphous solid. Yield 91%. Mp: 148-51 °C (after digestion with i-Pr20). MS
(EI): m/z 213 6.85 (m, 2H); 7.01 (t, 1H); 7.23 (t, 1H); 7.35 (br s, 1H); 7.45 (m, 1H); 7.60 (m, 1H) 7.88 (s,
1H); 8.80 (s, 1H) ppm. IR (Nujol): 3314,3141,1669,1630,1607,1577 cm-'.
Cyclohexylcarbamic acid 3'-carbamoylbiphenyl-3-yI ester. To a stirred mixture of
3Miydroxybiphenyl-3-carboxylic acid amide (0.43 g, 2 mmol) in toluene (12 mL), Et^
(0.012 g, 0.016 mL, 0.12 mmol), and cyclohexyl isocyanate (0.28 g, 0.28 mL, 2.2 mmol)
were added. After refluxing for 20 h, the mixture was cooled and concentrated. Purification
of the residue by column chromatography (cyclohexane/EtOAc 4:6) and recrystallization
gave 5a as a white solid. An amount of unreacted 4a (0.07 g, 17%) was also recovered.
Yield: 33%. Mp: 178 °C (EtOH) (sealed capillar tube). MS (EI): m/z 213 (100%). >H NMR
(CDCI3): 5 1.17-1.43 (m, 6H); 1.76 (m, 2H); 2.04 (m, 2H); 3.57 (m, 1H); 4.97 (br d, 1H);
5.63 (br s, 1H); 6.14 (br s, 1H); 7.16 (m, 1H); 7.39-7.56 (m, 4H); 7.77 (m, 2H); 8.03 (s, 1H)

ppm. IR (CHCI3): 3301, 3142, 1693, 1666, 1627, 1604, 1573 cm'1. Anal, calcd for
QoH^NA (338.41): C, 70.99; H, 6.55; N, 8.28. Found: C, 70.83; H, 6.65; N, 8.17.
Example 4: Molecular Modeling
[257] Molecular modeling calculations, including conformational analysis, energy
minimization and compound superposition were performed using a Sybyl 6.8 software
(Tripos), employing the MMFF94s force field.
Example 5: Biochemical assays
[258] We prepared cell fractions from rat brain homogenates, and assayed membrane
FAAH activity and cytosol MGL activity using anandamide [ethanolamine-3!!] (American
Radiolabeled Chemicals, ARC (St Louis MO), 60 Ci/mmol) and 2-mono-oleoyl-glycerol-
[glycerol-U3-3H] (ARC, St. Louis MO, 20 Ci/mmol), respectively, as substrates (Dinh, T.
Proc Natl. Acad. Set U. S. A. (2002)). We conducted [^anandamide transport assays in
human astrocytoma cells (Piomelli, D. et aL, Proc. Natl. Acad. Sci. U. S. A, 96,5802-5807
(1999)); CB1 and CB2 binding assays in rat cerebellar membranes and CB2-overexpressing
Chinese hamster ovary cells (Receptor Biology-Perkin Elmer, Wellesley, MA), respectively,
using [3H]WIN-55212-2 (NEN-Dupont, Boston, MA, 40-60 Ci/mmol) as a ligand (Devane,
W.A. et al. Science, 258, 1946-1949 (1992)); cholinesterase assays with a commercial kit
(Sigma, St. Louis, MO), using purified enzymes (electric eel acetylcholinesterase type V-S
and horse serum cholinesterase; bom from Sigma, St. Louis, MO) and following
manufacturer's instructions. To measure anandamide transport and hydrolysis in rat cortical
neurons, we preincubated the cells with FAAH inhibitors at appropriate concentrations for 10
min at 37°C, prior to exposure to [3H]anandamide for 4 min. hi some experiments, we
stopped the reactions with cold Tris-Krebs' buffer containing 0.1% bovine serum albumin
(Type V, fatty acid free, Sigma, St. Louis, MO), removed the cells by trypsin-EDTA
treatment, and extracted cell lipids with chloroform/methanol (1/1, vol/vol). We measured
non-metabolized [^anandamide in the organic phase of the extracts, and metabolized
[3H]anandamide (as [3H]ethanolamine) in the aqueous phase. In other experiments, after
having exposed the neurons to [3H]anandamide for 4 min, we replaced the medium, rinsed
the cells and measured [3H]anandamide release into the medium as described above.
Example 6: High-performance liquid chromatography/mass spectrometry
(HPLC/MS)

[259] We extracted lipids from tissues with a methanol-chloroform mixture and fractionated
them by silica gel column chromatography (Giuffrida, A. et al., Anal. Biochem., 280, 87-93
(2000)). Anandamide and other fatty acid derivatives were quantified by HPLC/MS, using
an isotope dilution method (Giuffrida, A. et al., Anal. Biochem., 280,87-93 (2000)).
Example 7: Body temperature and catalepsy
[260] We dissolved all compounds in saline/Tween 80/polyethylenglycol (90/S/5) and
administered them by i.p. injection immediately before tests. We measured body
temperature with a rectal probe (Type T, Copper-Costantan Thermocouple-Physitemp
Instruments INC, Clifton, NJ) connected to a digital thermometer (model BAT-12 Physitemp
Instruments INC, Clifton, NJ); and catalepsy using the procedure described in Tseng and
Craft (Tseng, A.H. et al., Eur. J. Pharmacol., 430,41-47 (2001)).
Example 8: Food intake
[261] We dissolved UCM597 in DMSO/saline (7/3) and administered it by i.p. injection 45
minutes before the test. We recorded food intake in free-feeding rats by using an automated
system (Scipro Inc., New York). Rats were acclimated to the test cages for three days prior
to the tests. Each test began at the onset of the dark phase and lasted for 24 hours.
Example 9: Antinociception
[262] We dissolved FAAH inhibitors in polyethyleneglycol/water (1/1) and rimonabant in
saline. Formalin and hot-plate assays were carried out in the mouse, as described (Beltramo,
M. et al., FEBSLett., 403,263-267 (1997)).
Example 10: Anxiety and motor activity
[263] We dissolved FAAH inhibitors and rimonabant in dimethylsulfoxide (DMSO)/saline
(7/3 and 9/1, respectively); we administered FAAH inhibitors by i.p. injection 30 min before
tests and rimonabant 30 min before UCM532. The elevated zero maze was comprised of a
black Perspex annular platform (105 cm diameter, 10 cm width) elevated to 65 cm above
ground level, divided equally into four quadrants (Bickerdike, M.J. et al., Eur. J. Pharmacol,
271,403-411 (1994); Shepherd, J.K. et al., Psyche-pharmacology, 116, 56-64 (1994)). Two
opposite quadrants were enclosed by black walls (27 cm high) on both the inner and outer
edges of the platform, while the other two were surrounded only by a shallow edge (1 cm
high). The apparatus was illuminated by uniform dim red light (40-60 lux). We placed the

rats in a closed quadrant and video recorded them for 5-min periods, thoroughly cleaning the
maze between sessions. Rats were considered to be in an open quadrant when their four
paws were within the quadrant. Results are expressed as percent time in open quadrant/total
time (percent time open). Results were analyzed by one-way ANOVA followed by Tukey's
test We recorded motor activity of each rat in an Opto-Varimex cage (Columbus
Instruments, Columbus, OH) linked on-line to a computer and placed in a sound-attenuated
room illuminated by a 20-W white light. The amount of time spent in ambulatory activity
was analyzed using an Auto-Track software (Columbus Instruments, Columbus, OH) as
described elsewhere (Ali, MM. et al., Neurosci. Lett., 284,57-60 (2000); Wedzony, K. et al.,
Neuropsychopharmacology, 23, 547-559 (2000)). Session duration was 20 min for 10-days
old adult rats and 60 s for 10-days old pups. We analyzed data by overall one-way ANOVA
followed by Tukey's test for individual between-group comparisons. We recorded 10-days
old pup ultrasonic vocalizations in a sound-attenuating chamber, following the procedure
described by Cuomo et al. (Cuomo, V. et al., Neuropharmacology, 26, 701-705 (1987)).
Tests were conducted between 900 and 1400 h and lasted for 15 s. Drugs were administered
after baseline values collection (15 s) and pups were tested again 30 min after drug
administration. Data were expressed as percent change from baseline and analysed by overall
one-way ANOVA followed by Tukey's test for individual between-group comparisons.
Example 11: Lead identification and optimization
[264] Despite its unusual catalytic mechanism (Patricelli, M.P. et al., Biochemistry, 38,
9804-9812 (1999)), FAAH is blocked by a variety of serine hydrolase inhibitors, including
compounds with activated carbonyls (Boger, D.L. et al. Proc Natl Acad. Set U. S. A, 97,
5044-5049 (2000)). Therefore we examined whether esters of carbamic acid such as the
anticholinesterase agent carbaryl (Table I, compound 1) may inhibit FAAH activity in rat
brain membranes. Although 1 was ineffective, its positional isomer 2 produced a weak
inhibition of FAAH (half-maximal inhibitory concentration, ICso = 18.6 ± 0.7 uM; mean ±
SEM, n - 3), which was enhanced by replacing the A'-methyl substituent with a cyclohexyl
group (3, IC50- 324 ± 31 nM). The aryl ester 4, the benzyloxyphenyl group of which can be
regarded as an elongated bioisosteric variant of the naphthyl moiety of 2, inhibited FAAH
with a potency (IC50 = 396 ± 63 nM) equivalent to 3. A conformational analysis of 4
revealed families of accessible conformers differing mainly in the torsion angle around the O-
CH2 bond, with substituents in anti or gauche conformations (data not shown). As the latter

conformations more closely resembled the shape of the naphthyl derivative 3, we
hypothesized that they might be responsible for the interaction of 4 with the active site of
FAAH. Testing this hypothesis led to the design of the biphenyl derivative 5 (IC5 0 = 63 ± 9
nM), which was further optimized by systematic modifications of the distal phenyl group,
resulting in the potent inhibitor 6 (IC50=4.6 ±1.6 nM) (Table 1).
[265] Kinetic analyses and dialysis experiments suggest that compounds 4 and 6 may inhibit
FAAH activity through an irreversible interaction with the enzyme (data not shown), possibly
due to a nucleophilic attack of an active serine residue on the carbamate group. This
mechanism sets the present compounds apart from the a-keto heterocycle derivatives
described by Boger et al. (D.L. Boger et al., Proa Natl. Acad. Set U. S. A., 97, 5044-5049
(2000)), which act as competitive FAAH inhibitors. A further indication of such distinction is
that in the a-keto heterocycle series potency is strongly dependent on the hydrophobicity of
the flexible acyl chain, whereas in the carbamate series potency is modulated by the shape of
the rigid aromatic moiety. Accordingly, when we replaced the biphenyl of 5 with a 5-
phenylpentyl group, representing the most effective acyl chain in the a-keto heterocycle
series, the inhibitory activity was lost (compound 7, Table 1).
Compounds 4 (UCM532) and 6 (UCM597) blocked the FAAH-catalyzed hydrolysis of
exogenous [3H]anandamide by intact cortical neurons in primary cultures, with IC50 values
that paralleled those obtained in membrane preparations (UCM532,214 ± 79 nM; UCM597,
0.50 ± 0.05 nM; n = 8) (Fig. la). By contrast, compound 7, a UCM532 analog that does not
inhibit FAAH in membranes (Table 1), had no such effect (Fig. lb). Moreover, UCM532
and UCM597 selectively impaired the breakdown of [3H]anandamide without reducing its
carrier-mediated uptake, causing non-metabolized [^HJanandamide to accumulate in, and
eventually exit from, the neurons. Thus, after a 4-min incubation with [3H]anandamide, the
intracellular content of non-metabolized [3H]anandamide was markedly higher in inhibitor-
treated man in control neurons (Fig. lc). As expected, the anandamide transport blocker N-
(4-hydroxyphenyl)arachidonamide (AM404) had an opposite effect, significantly reducing
[^anandamide internalization (Beltramo, M. et al., FEBSLett., 403, 263-267 (1997)) (Fig.
lc). When UCM597-treated neurons were exposed for 4 min to [3H]anandamide and then
incubated for 15 min in an [3H]anandamide-free solution, 42.6 ± 8.7% of the accumulated
[3H]anandamide was released back into the medium (n - 3) (Fig. Id). This process was
linear with time (Fig. lc) and was not inhibited by AM404 (Fig.l occurred through passive diffusion rather man reverse transport No such time-dependent

release was observed in control neurons, the medium of which only contained residual levels
of [^anandamide carried over from the preincubation period. Together, these studies
identify a novel class of carbamate inhibitors of FAAH activity, which potently block
anandamide breakdown in intact brain neurons.
Example 12: Target selectivity
[266] UCM532 and UCM597 inhibited FAAH, but did not affect the activities of three
additional serine hydrolases: electric eel acetylcholinesterase, horse plasma butyryl
cholinesterase, and rat brain monoglyceride lipase (MGL) (Table 2). The lack of MGL
inhibition is particularly noteworthy in light of the proposed role of this enzyme in the
biological inactivation of 2-aracbidonoylglycerol (2-AG) (Dinh, T. Proc. Natl. Acad. Sci. U.
S. A. (2002)), another endogenous cannabinoid present in the brain (Mechoulam, R. et al.
Biochem. Pharmacol., 50, 83-90 (1995); Sugiura, T. et al., Biochem. Biophys. Res. Commun.,
215, 89-97 (1995); Stella, N. et al., Nature, 388, 773-778 (1997)). Furthermore, UCM532
and UCM597 had no effect on anandamide transport in human astrocytoma cells or on the
binding of a high-affinity ligand to CB1 and CB2 receptors (Table 2). Even further,
UCM532 (10 fiM) did not significantly interact with a panel of 21 receptors, ion channels and
neurotransmitter transporters, which included adenosine Ai, AM and A2B; adrenergic ajA,
a2A, Pi and P2; dopamine Di and D2; glutamate JV^methyl-(D)-aspartate; y-amino-butyric acid
(GABA)A agonist site; histamine Hi; opiate n; muscarinic M2; and brain nicotinic receptors
(data not shown). This high selectivity for FAAH encouraged us to examine the effects of
UCM532 and UCM597 in live animals.
Example 13: FAAH inhibition in vivo
[267] Intraperitoneal (i.p.) injections of either UCM532 or UCM597, but not the inactive
analog 7, produced a profound, dose-dependent inhibition of brain FAAH activity (Fig. 2a).
In six experiments, half-maximal inhibition was reached at 0.60 ± 0.09 mg kg'1 UCM532 and
0.150 ± 0.007 mg kg*1 UCM597. After injection of a maximal dose of UCM597 (0.3 mg kg"
\ i.p.), FAAH inhibition was rapid in onset ( 6 h) (Fig. 2b), and
accompanied by significant elevations in the brain content of anandamide (Fig. 2c) and other
fatty acid ethanolamides that are substrates for FAAH (in pmol g'1 of tissue at 2 h after
injection; oleoylethanolamide: vehicle, 137.0 ± 14.3; UCM597 0.3 mg kg"1, 725.3 ± 28.6;
palmitoylethanolamide: vehicle, 259.1 ± 15.0; UCM597, 1324 ± 395; n = 8-15). Parallel

changes in FAAH activity and fatty acid ethanolamide levels were also measured in various
peripheral tissues (data not shown). In agreement with the lack of MGL inhibition noted in
our in vitro experiments (Table 2), UCM597 did not change the brain content of 2-AG (Fig.
Id).
[268] As previously observed in mutant FAAH"'" mice (Cravatt, B.F. et al., Proc. Natl.
Acad. Sci. U. S. A., 98, 9371-9376 (2001)), FAAH inhibition was associated with increased
sensitivity to the administration of exogenous anandamide. Accordingly, UCM597 (0.3 mg
kg'1, i.p.) intensified and prolonged the decrease in body temperature elicited by a
subthreshold dose of anandamide (5 mg kg'1, i.p.), whereas it had no effect when injected
alone (Fig. 2e) {Ftnctnmis = 38.36, df = 1/143, P 0.0001; Ftunxtreatmrns = 2.64, df = 12/143, P anandamide plus UCM597).
Example 14: Antinociceptive effects of FAAH inhibitors
[269] Though UCM532 and UCM597 increased brain anandamide levels, they did not
overtly mimic the spectrum of pharmacological responses produced by exogenous
anandamide. Systemic doses of UCM532 (10 mg kg"1, i.p.) or UCM597 (0.3 mg kg"1, i.p.)
that maximally blocked FAAH activity produced no catalepsy (rigid immobility),
hypothermia or hyperphagia (increased food intake), three typical signs of CB1 receptor
activation (Chaperon, F. et al., Crit. Rev. Neurobiol, 13,243-281 (1999)) (data not shown).
The compounds exerted, however, moderate antinociceptive effects in two models of acute
pain, m the mouse hot-plate test, which measures the animal's response to noxious thermal
stimuli, UCM597 significantly lengthened response latencies at a dose of 0.5 mg kg'1 (Fig.
3a), but not at a lower dose (0.1 mg kg"1; data not shown). Moreover, in the mouse formalin
test, which measures nocifensive reactions to chemical tissue damage, UCM597 (0.5 mg kg"1)
attenuated the early phase of pain behavior, with little or no change in the late phase (Fig.
36). Both effects were abrogated by the CB1 antagonist SR1417161A (rimonabant) (02 mg
kg*1, intravenous, i.v.) (Fig. 3a and b) and mimicked, albeit less effectively, by UCM532
(data not shown). Our results corroborate those obtained in mutant FAAH"7" mice (Cravatt
Bj; et al., Proc. Natl. Acad Sci. U. S. A., 98, 9371-9376 (2001), indicating that acute
disruption of FAAH activity results in moderate CB1 -mediated antinociception, but no
hypothermia or catalepsy.
Example 15. Anxiolytic effects of FAAH inhibitors

[270] To identify other intrinsic actions of anandamide that might be magnified by FAAH
inhibition, we turned to the regulation of emotional reactivity, for three reasons. First, CB1
receptors are expressed at high levels in brain regions, such as the amygdala, which are
implicated in the control of anxiety and fear (Herkenham, M. et al., Proc. Natl. Acad. Sci. U.
S. A, 87,1932-1936 (1990); Glass, M. et al., Neuroscience, 77,299-318 (1997); Katona, I. et
al., J. Neurosci., 21, 9506-9518 (2001)). Second, acute administration of cannabinoid drugs
produces marked emotional responses in rodents (Chaperon, F. et al., Crit. Rev. Neurobiol,
13,243-281 (1999)) and humans (Hall, W. et al., Lancet, 352,1611-1616 (1998); Robson, P.
Br. J. Psychiatry, 178, 107-115 (2001)). Third, the CB1 antagonist rimonabant elicits
anxiety-like behaviors in rats, suggesting the existence of an intrinsic anxiolytic tone
mediated by endogenous cannabinoids (Rodriguez de Fonseca, F. et al., Science, 276, 2050-
2054 (1997); C. Arevalo, R. et al., Pharmacol. Biochem. Behav., 70,123-131 (2001)).
[271] We used two pharmacologically validated animal models of anxiety, the elevated zero
maze test, and the isolation-induced ultrasonic emission test. The zero maze consists of an
elevated annular platform with two open and two closed quadrants and is based on the
conflict between an animal's instinct to explore its environment and its fear of open spaces,
where it maybe attacked by predators (Bickerdike, M.J. et al., Eur. J. Pharmacol., 271,403-
411 (1994); Shepherd, J.K. et al., Psychopharmacology, 116, 56-64 (1994)). Clinically used
anxiolytic drags, such as the benzodiazepines, increase the proportion of time spent in, and
the number of entries made into, the open compartments. Similarly, UCM532 (5 and 10 mg
kg"1, i.p.) and UCM597 (0.05-0.1 mg kg'1, i.p.) evoked anxiolytic-like responses at doses mat
corresponded to those required to inhibit FAAH activity in vivo (Fig. 4 a and b) (F - 38.58,
df - 2/27, P endogenous anandamide, the anxiolytic-like effects of UCM532 were attenuated by a non-
anxiogenic dose of the CB1 antagonist rimonabant (2 mg kg"1, i.p.) (Fig. 4c) (F= 14.87, df=
3/31, P motor behavior. Indeed, although UCM532 elicited, in adult rats, a modest decrease in
ambulation (which was also antagonized by rimonabant, data not shown), it did so at doses
that were higher than those needed to cause anxiolysis (>10 mg kg"1) (Fig. Ad) (F - 3.57, df
■ 2/22, P vocalization emission model, which measures the number of stress-induced vocalizations
emitted by rat pups removed from their nest (Insel, TiL et al., Pharmacol. Biochem. Behav.,
24, 1263-1267 (1986); Miczek, K.A et al., Psychopharmacology, 121, 38-56 (1995);
Winslow, J.T. et al., Biol. Psychiatry, 15:745-757 (1991)). As seen with anxiolytic drugs,

UCM532 strongly reduced ultrasonic calls (Fig. 4c) (F - 12.27; df - 2/18, P dose (5 mg kg"1) that had no effect on pup movement (data not shown) (F - 3.23, df = 2/18,
n.s.).
Example 16. Overall Pharmacological Activity
[272] The fatty acid amide hydrolase inhibitors of Formula I and Formula II represent a new
class of agents mat prevent anandamide inactivation by targeting the intracellular enzymatic
activity of FAAH. UCM597, the most potent member of mis class, inhibited FAAH activity
with an IC50 value of 4 nM in brain membranes and 0.5 nM in intact neurons, and an ID50
value of 0.15 mg kg'1 following systemic administration in the rat This compound had much
greater selectivity for FAAH than other cannabinoid-related targets, including cannabinoid
receptors (selectivity index: > 25,000) and MGL, an enzyme involved in the deactivation of
the endogenous cannabinoid ester, 2-AG (selectivity index: > 7,500). Such a remarkable
target discrimination was matched by a lack of overt cannabimimetic effects in vivo. Thus, at
doses that almost abolished FAAH activity and substantially raised brain anandamide levels,
UCM597 and its analog UCM532 did not evoke catalepsy, reduce body temperature or
stimulate feeding, three key symptoms of cannabinoid intoxication in the rodent (Chaperon,
F. et al., Crit. Rev. Neurobiol., 13,243-281 (1999)).
[273] Nevertheless, the compounds did elicit marked anxiolytic-like responses, which
paralleled their ability to inactivate FAAH and were attenuated by the CB1 receptor
antagonist rimonabant.
[274] Without being wed to theory, UCM597 and UCM532 selectively modulate anxiety-
like behaviors by enhancing the tonic actions of anandamide on a subset of CB1 receptors,
which may normally be engaged in controlling emotions. Forebrain sites that might be
implicated in such actions include the basolateral amygdala, the anterior cingulate cortex and
the prefrontal cortex, key elements of an "emotion circuit" (Cahill, L. et al., Trends
Neurosci., 21,294-299 (1998)) that contains high densities of CB1 receptors (Herkenham, M.
et al., Proc Natl. Acad. Set U. S. A., 87,1932-1936 (1990); Glass, M. et al., Neuroscience,
77, 299-318 (1997)). Interestingly, CB1 receptors in these structures are exclusively
localized to the axon terminals of a subpopulation of GAB A-ergic interneurons, which also
express me peptide cholecystokinin (CCK) (Katona, L et al., J. Neurosci., 21, 9506-9518
(2001); McDonald, AJ. et al., Neuroscience, 107,641-652 (2001)).
[275] In addition to their anxiolytic-like actions, UCM597 and UCM532 exerted moderate,
but significant antinociceptive effects, which also were sensitive to CB1 receptor blockade.

These findings are strikingly similar to those reported for mutant FAAIT'- mice (Cravatt, B.F.
et al., Proc. Natl. Acad. Sci. U. S. A., 98:9371-9376 (2001)) and underscore the emerging
roles of anandamide in the intrinsic modulation of pain (Iversen, L. et al., Curr. Opin.
Pharmacol, 2:50-55 (2002)). Since emotional states may strongly influence pain sensation,
it is possible that anxiolysis might have contributed to the antinociceptive effects of the
FAAH inhibitors. Distinguishing the roles of these two components will require, however,
further experimentation.
[276] UCM597 and TJCM532 increased brain anandamide levels without modifying those
of the second endogenous cannabinoid, 2-AG. It is likely, therefore, that the pharmacological
actions of these compounds, which are sensitive to the CB1 antagonist rimonabant, are
primarily due to anandamide accumulation But the FAAH inhibitors also produced large
elevations in the levels of two anandamide analogs, palmitoylethanolamide and
oleoylethanolamide, whose recently discovered biological effects are independent of CB1
receptors (Calignano, A- et al., Nature, 394:277-281 (1998); Rodriguez de Fonseca, F. et al.
Nature, 414:209-212 (2001)).
Example 17
[277] This example provides the FAAH inhibitory IC50 values for over 50 compounds
according to Formula I or Formula II. The results of testing the compounds are shown in
Table 4.
Example 18
[278] This example provides a more detailed 3D-QSAR analysis of O-aryl N-alkylcarbamic
acid aryl esters and then relates such to the 3-dimensiohal structure of FAAH.
[279] Recently, starting from the assumption that carbamic acid esters could act as active
site-directed inhibitors of FAAH, a series of O-aryl-i^-alkylcarbamic acid aryl esters were
developed that irreversibly inhibit FAAH activity with good in vitro and in vivo potency, and
as a result, exert anxiolytic effects in rats. The methods and results of this work are published
in (Kathuria, S, et al., Nat Med. 9,76-81 (2003); Tarzia, G., et al., J. Med. Chem. 46,2352-
2360 (2003)) and incorporated herein by reference in its entirety. Notably, most of the
compounds block FAAH, but not several other serine hydrolases, e.g. acetylcholinesterase
and MGL, and do not bind to cannabinoid receptors. A preliminary S AR investigation, aimed
at the definition of shape requirements for the lipophilic O-aryl moiety, showed that
structures characterized by a non-linear shape led to an improvement of potency. The

methods and results of this work are described in Tarzia, et al., Med. Client. 46, 2352-2360
(2003) which is incorporated by reference in its entirety. More precisely, the curved
molecules of the most potent inhibitors resembled that observed for the folded conformations
of fatty acids in complexes with different proteins and for the so-called U-shaped
conformation of anandamide, (Reggio, P.H., et al. Chem. Phys. Lipids 108, 15-35 (2000)
which had been recently proposed as one possibly assumed at the CBi receptor binding site.
(Bamett-Norris, J., et al., J. Med. Chem. 451, 3649-3659 (2002)). Moreover, the recently
published crystal structure of a complex of FAAH and the inhibitor methyl arachidonyl
fluorophosphonate (MAPF)(Bracey, M. H., Science 298,1793-1796 (2002) revealed a folded
conformation of the arachidonyl chain.
[280] The 3D-QSAR analysis of O-aryl-JV-alkylcarbainic acid aryl esters indicates that the
space occupancy of a region corresponding to the meta position of an O-phenyl ring was
positively correlated with inhibitor potency, thus suggesting its beneficial interaction with the
enzyme binding site. The most potent compound found in this series was URB524 (2, Figure
5), having an IC50 value of 63 nM; To explore QSARs of carbamate FAAH inhibitors and
optimizing their activity, compound based upon this compound as the starting point were
tested to provide for a systematic exploration of the effect of phenyl substitution. As in the
previous series only lipophilic groups had been introduced at the O- position of the
carbamate, the series provided information only on the steric interactions occurring within the
binding site. To further characterize the nature of possible interaction, a balanced set of
substituents, with variation of their lipophilic and electronic properties, was introduced at the
meta and para positions of the distal phenyl ring of URB524, since mis moiety had been
indicated as crucial by the cited 3D-QSAR model.
[281] The experimental design for the exploration of substituent effects was set up in two
steps. Firstly, a small set of substituents, having moderate size, was employed to test the
sensibility of the meta and the para positions to lipophilic and electronic properties; the four
substituents methyl, trifluoromethyl, amino and carbamoyl represented the four combinations
of positive and negative levels for the n and a (am or ap) descriptors; (van de Waterbeemd,
H., et aL J. Comput-Aided Des. 3,111-132 (1989)); furthermore, a small (fluoro) and a big
(cyclohexylcarbamoyloxy) substituents were added to this first explorative set. Having
identified the more responsive position of the phenyl ring, as a second optimization step the
series of subsitutents were expanded seeking to maintain significant and independent
variation among the variables describing lipophilic, electronic and steric properties of the

substituents, which is a prerequisite to investigate QSARs by multiple regression analysis
(MRA) (Box, G. E. P.; et al. Statistics for Experimenters, Wiley: New York 306-373 (1978);
van de Waaterbeemd, H., et al. Chemometric Methods in Molecular Design, VCH Publishers
Inc.: New York 49-62 (1995)).
[282] Half-maximal concentrations (IC50) for inhibition of FAAH activity in rat brain
membranes, using [3H]anandamide as a substrate,18 by compounds 3a-z are reported in Table
5.
[283] The first limited set of substituents at the ending phenyl ring (3a-l) revealed that the
meta position was much more promising for potency optimization. In fact, though the 3'-
methyl (3h) and 3'-amino (3j) derivatives resulted in a compound as potent as the parent
compound for FAAH inhibition, the 3'-carbamoyl one (3i) was more potent by an order of
magnitude; on the other hand, all the para-substituted compounds were less potent than the
parent compound, only the 4'-fluoro derivative (3e) having a comparable IC50 value.
[284] The set of substituents at the meta position was therefore expanded to explore
statistical relationships between substituent properties and inhibitor potency. Therefore, 12
additional substituents (3m-z in Table 5) were selected to enlarge the space representing
lipophilic, steric and electronic properties; some of them were chosen for their similarity to
the carbamoyl group (i.e., the sulphamoyl group in 3r) or to parts of it (i.e., the acetyl,
aminomethyl, or hydroxymethyl group in 3s, 3z, or 3v, respectively). The 19 substituents,
including H, reported in Table 6, had large variation in lipophilicity (almost 4 n unities) and
steric bulk, boui practically uncorrelated to electronic effects (r with am of -0.19 and -0.16,
respectively); they still had some correlation between lipophilic (it) and steric (MR)
descriptors (r=0.63), due to the known difficulty to obtain big hydrophilic substituents.




[285] Multiple regression analysis (MRA) applied to the data set composed by maximum 3
active variable, selected among 8 common physico-chemical descriptors (p, om F, R, MR, L,
Bi, Bs) and the square of it, gave no significant model. However, a simple plot of the potency
variable, pICso, vs lipophilicity (Figure 6) revealed that a clear relationship was observable,
but the methylamino derivative 3z resulted an outlier, this could be attributed to its basicity,
making it the only compound with a large prevalence of protonated, cationic species at
neutral pH. Omitting 3z from the regression set, the negative correlation between lipophilicity
and potency could be described by the linear model reported in equation 1; the pIQo values
calculated by equation 1 are reported in Table 6.


[286] This negative correlation is characteristic of the meta position, since the limited set of
para substituents showed, on the same scales, a parabolic relationships described by the
regression equation 2.

[287] While the set of para-substituted compounds was too small to consider more
complicated models, the 18-compound set of meta-substituted derivatives represented a good
set for a detailed QSAR investigation; moreover, the negative correlation with lipophilicity
appeared perplexing, as our previous results supported the hypothesis that the biphenyl
moiety would mimic the arachidonoyl chain in its lipophilic FAAH binding site.Em>rl B,okmark
not denned. However, no MRA model including up to 5 variables had statistical quality better or
comparable to that of equation 1; only the inclusion of an hydrogen bond indicator variable
(HB), set to 1 for substituents able to give hydrogen bonds and to 0 in other cases, allowed
the detection of an alternative model (equation 3) with comparable descriptive (r2) and
predictive (q2) power, indicating that potency increase was negatively correlated to the steric
bulk (MR) of the substituents, but the polar ones had an average 0.8 pICso units more than the
apolar ones.

[288] Although this model was not statistically better than equation 1, it could provide a
possible interpretation for the positive effect of substituent hydrophilicity within a putative
lipophilic binding pocket, attributing this behaviour to the formation of hydrogen bonds
between the meta substituent and some polar atoms of the enzyme, not available for the para
substituents.
[289] The crystal structure for the FAAH bound to a covalent arachidonyl-phosphonate
inhibitor22 has recently been reported by others. Without being wed to theory, the docking of
our inhibitors within the enzyme active site and molecular dynamics simulations can explain
the role of the U and V groups or R groups with respect to the subject compounds surprising

ability to inhibit the enzyme. It is thought that the binding site of the aforementioned
arachidonyl inhibitor is part of a channel spanning the whole enzyme, represented in Figure
7; the catalytic Ser241 is placed in the middle of this channel, which extends towards the
membrane on one side (bottom in Figure 7, left) and the cytosol on the other. This channel
has a complex topography, with a hydrophilic surface in correspondence of the catalytic site,
surrounded by lipophilic surfaces in the two directions. That pointing towards the membrane,
which is occupied by the arachidonyl chain of the phosphonate inhibitor, forms a lipophilic
bulge allocating the terminal atoms of this inhibitor and a narrow tunnel having a hydrophilic
"ridge," which could be used by the hydrophilic head of OEA to approach the catalytic site,
while moving along from the membrane. Our molecular docking showed that the biphenyl
moiety of URB524 could, after deletion of the phosphonate inhibitor, occupy the space of the
arachidonyl chain, with the meta position of the distal phenyl ring pointing exactly towards
the hydrophilic ridge (see Figure 7). The superposition of a minimum-energy conformation
of the biphenyl moiety to the arachidonyl chain, in the conformation found for the
MAPF/FAAH complex, highlights the steric similarity with the first two double bonds, thus
supporting our hypothesis (see Figure 8). Interestingly, all the polar meta substituents were
able to undertake hydrogen bonds with some polar residue of the hydrophilic ridge. The
heuristic model described by of equations 1 and 3 indicates that the inhibitor potency of meta
substituted biphenylyl carbamates is negatively correlated to lipopbilicity because, although
the biphenyl scaffold can be located in the lipophilic region of space generally occupied by
the first atoms of fatty acid chain, substituents at the meta position of the distal phenyl ring
can interact to form hydrogen bonds with a hydrophilic wall of a narrow tunnel within the
enzyme. These results indicate that moieties interacting via hydrogen bonding with the
subject hydrophilic ridge can contribute importantly to the activity of a FAAH inhibitor.
Consistent with this prediction from our model, docking of the most potent compound in the
biphenyl carbamate series, 3i (URB 597), led to a solution which showed the possibility that
the carbamoyl group of URB 597 undertakes two hydrogen bonds with the enzyme: one as an
HB-acceptor, with the hydroxyl group of Thr488, and the second as an HB-donor, with the
backbone carbonyl of Leul92 (Figure 9). Altogether our results show that compounds
primarily interacting with the hydrophobic channel rather than the catalytic site can be high
potency inhibitors of FAAH.
[2901 Each publication, patent application, patent, and other reference cited in any part of
the specification is incorporated by reference in its entirety to the extent mat it is not
inconsistent with the present disclosure.

[291] Although the foregoing invention has been described in some detail by way of
illustration and example for purposes of clarity of understanding, it will be readily apparent to
those of ordinary skill in the art in light of the teachings of this invention that certain changes
and modifications may be made thereto without departing from the spirit or scope of the
appended claims.

WE CLAIM:
1. An N-substituted carbamate meta-biphenyl ester of the formula:

wherein:
X is O
Q is O;
n is 1, 2, or 3;
wherein each Ra member is independently selected from the group consisting of
unsubstituted saturated alkyl, unsubstituted alkenyl, unsubstituted alkynyl,
unsubstituted at least partially saturated cycloalkyl, unsubstituted
ketoalkyl, unsubstituted hydroxyalkyl, unsubstituted aminoalkyl, -CH2-
NR3R4, unsubstituted alkoxy, halo, unsubstituted haloalkyl, cyano,
hydroxy, nitro, amino, -NR3R4, unsubstituted carboxamido, -CONR3R4,
sulfonamide -SO2NR3R4; -SR5, -SO2R6, -S(O)R6, -NR6C(O)R7, and
-NR6SO2R7;
wherein R3 and R4 are independently selected from the group consisting of H,
unsubstituted saturated alkyl, unsubstituted alkenyl, unsubstituted alkynyl,
unsubstituted at least partially saturated cycloalkyl, unsubstituted
hydroxyalkyl and imino-methylamino and wherein optionally R3 and R4
may together with the N atom to which they are attached combine to form
a 5-7 membered ring; and R5, R6 and R7 are independently selected from
the group consisting of hydrogen, unsubstituted alkyl, and unsubstituted
heteroalkyl;
wherein R1 and R2 are independently selected from the group consisting of
hydrogen, substituted or unsubstituted straight or branched alkyl,
substituted or unsubstituted heteroalkyl, substituted or unsubstituted at
least partially saturated cycloalkyl, and substituted or unsubstituted
cycloheteroalkyl, with the proviso that R1 and R2 are not both hydrogen;
and the pharmaceutically acceptable salts thereof.

2. The compound as claimed in claim 1, wherein the compound is of a formula
selected from the group consisting of:

wherein Ra1 and Ra2 are independently selected from the group consisting
of unsubstituted saturated alkyl, unsubstituted alkenyl, unsubstituted alkynyl,
unsubstituted cycloalkyl, unsubstituted ketoalkyl, unsubstituted hydroxyalkyl,
unsubstituted aminoalkyl, -CH2-NR3R4, unsubstituted alkoxy, halo, unsubstituted
haloalkyl, cyano, hydroxy, nitro, amino, -NR3R4, unsubstituted carboxamido, -CONR3R4,
sulfonamide -SO2NR3R4; -C(O)R6, -SR5, -SO2R6, -S(O)R6, -NR6C(O)R7, and -NR6SO2R7;
and
wherein R3 and R4 are independently selected from the group consisting of H,
unsubstituted saturated alkyl, unsubstituted alkenyl, unsubstituted alkynyl, unsubstituted
at least partially saturated cycloalkyl, unsubstituted hydroxyalkyl and imino-methylamino

and wherein optionally R3 and R4 may together with the N atom to which they are
attached combine to form a 5-7 membered cyclic ring; and R5, R6 and R7 are
independently selected from the group consisting of hydrogen, unsubstituted alkyl, and
unsubstituted heteroalkyl.
3. The compound as claimed in claim 2, wherein R2 is selected from the group
consisting of substituted or unsubstituted straight or branched alkyl, substituted or
unsubstituted heteroalkyl, substituted or unsubstituted at least partially saturated
cycloalkyl, and substituted or unsubstituted cycloheteroalkyl.
4. The compound as claimed in claim 3, wherein each Ra member is independently
selected from the group consisting of unsubstituted saturated alkyl, unsubstituted
heteroalkyl, unsubstituted alkenyl, unsubstituted alkynyl, unsubstituted at least partially
saturated cycloalkyl, unsubstituted ketoalkyl, unsubstituted hydroxyalkyl, unsubstituted
aminoalkyl, -CH2-NR3R4, unsubstituted alkoxy, halo, unsubstituted haloalkyl, cyano,
hydroxy, nitro, amino, -NR3R4, -SR5, unsubstituted carboxamido, -CONR3R4,
sulfonamido, and -SO2NR3R4
5 . The compound as claimed in claim 4, wherein R1 is hydrogen; and R2 is an isomer
of propyl, butyl, pentyl, hexyl, heptyl, octyl, heteroalkyl, cycloheteroalkyl, or at least
partially saturated cycloalkyl.
6. The compound as claimed in claim 2, wherein R2 is selected from the group
consisting of unsubstituted straight or branched alkyl, unsubstituted heteroalkyl,
unsubstituted at least partially saturated cycloalkyl, and unsubstituted cycloheteroalkyl
7 . The compound as claimed in claim 5, wherein R2 is fully saturated and an isomer
of pentyl, hexyl, heptyl, or octyl.
8. The compound as claimed in claim 5, wherein the R2 member is a fully saturated
cycloalkyl.
9 . The compound as claimed in claim 2, wherein at least one of R1 and R2 is H.
10. The compound as claimed in claim 1, wherein each Ra is independently selected
from the group consisting of -C(O)NH2,-C(O)CH3, -(CH2)2OH, SO2NH2, CH2OH, - CF3,
-CH3, -CH2CH3, n-C3H7, -NH2, -F, -NO2, -CN and -OH.

11. The compound as claimed in claim 2, wherein the compound is of the formula:

12. The compound as claimed in claim 11, wherein Ra1 is selected from the group
consisting of-C(O)NH2,-C(O)CH3, -(CH2)2OH, -SO2NH2, -CH2OH, -CF3, -CH3,
-CH2CH3, n-C3H7, -NH2, -F, -NO2, -CN and -OH.
13 . The compound as claimed in claim 11, wherein R2 is selected from the group
consisting of unsubstituted straight or branched alkyl, unsubstituted heteroalkyl,
unsubstituted at least partially saturated cycloalkyl, and unsubstituted cycloheteroalkyl.
14. The compound as claimed in claim 1, wherein n is 2.
15. The compound as claimed in claim 1, wherein R1 and R2 are independently
selected from the group consisting of hydrogen, substituted or unsubstituted at least
partially saturated cycloalkyl, and substituted or unsubstituted cycloheteroalkyl.
16. The compound as claimed in claim 1, wherein the compound has an IC50 for
inhibiting the human fatty acid amide hydrolase of less than 1 micromolar.
17. The compound as claimed in claim 1, wherein the compound has an IC50 for
inhibiting the human fatty acid amide hydrolase of less than 10 nanomolar.
18. An N-substituted carbamate meta-biphenyl ester of the formula

wherein
X is O or S;
Q is O or S;

R is substituted or unsubstituted meta-biphenyl;
R1 and R2 are independently selected from the group consisting of hydrogen,
substituted or unsubstituted at least partially saturated cycloalkyl, and substituted or
unsubstituted cycloheteroalkyl, with the proviso that R1 and R2 are not both hydrogen.-
19 . The compound as claimed in claim 18, wherein X is O and Q is O and R is
unsubstituted meta biphenyl.
20 . The compound as claimed in claim 19, wherein one of R1 and R2 is hydrogen.
21 . The compound as claimed in claim 20, wherein R1 is at least partially saturated
unsubstituted C3-C8 cycloalkyl or unsubstituted C3-C8 cycloheteroalkyl.
22 . The compound as claimed in claim 20, wherein R1 is an at least partially saturated
cyclopentyl, cyclohexyl, cycloheptyl, cyclopentenyl, cyclohexenyl, cycloheptenyl,
cyclooctyl, or cyclooctenyl.
23 . The compound as claimed in claim 22, wherein R1 is a saturated cyclopentyl,
cyclohexyl, cycloheptyl, or cyclooctyl.
24 . The compound as claimed in claim 20, wherein R1 is unsubstituted and is a
saturated cyclopentyl, cyclohexyl, or cycloheptyl.
25 . The compound as claimed in claim 18, wherein the compound is N-cyclohexyl
3'-carboxamido biphenyl-3-yl carbamate.
26. The compound as claimed in claim 18 of the formula:

wherein m is a number from 0 to 1 and n is a number from 0 to 2;
each Ra member is independently selected from the group consisting of H,

halogen, nitro, cyano, hydroxy, -C(O)C1-C4alkyl, sulfonamido, unsubstituted C1-C4alkyl,
hydroxyC1-C4alkyl, unsubstituted phenoxy, and unsubstituted phenyl, carboxamido,
amino, and trihalomethyl; and
Rb is selected from the group consisting of C1-C4alkyl, amino, and carboxamido;
wherein the C1-C4alkyl subject matter of the Ra and Rb Markush groups are
independently methyl, ethyl, propyl, or butyl;
and the pharmaceutically acceptable salts thereof..

27. The compound as claimed in claim 26, wherein Rb is selected from the group
consisting of methyl, amino, and carboxamido.
28. The compound as claimed in claim 26, wherein the compound is of the formula:

wherein Ra1 and Ra2 are independently selected from the group consisting of H,
halogen, unsubstituted C1-C4alkyl, unsubstituted hydroxyC1-C4 alkyl, unsubstituted
phenoxy, and unsubstituted phenyl, carboxamido, amino, sulfonamido, cyano, hydroxy,
nitro, and trihalomethyl, and
wherein the C1-C4alkyl is methyl, ethyl, propyl, or butyl.
29. The compound as claimed in claim 28, wherein at least one of Ra1 and Ra2 is H.
30. The compound as claimed in claim 28, wherein Ra1 selected from the group
consisting of halogen, unsubstituted C1-C4 alkyl, unsubstituted phenoxy, and
unsubstituted phenyl, carboxamido, amino, and trihalomethyl and Ra2 is H.
31. The compound as claimed in claim 29, wherein Ra1 and Ra2 are each H.
32. The compound as claimed in claim 29, wherein Ra2 is selected from the group
consisting of trihalomethyl, carboxamido, amino, C1-C4 alkyl, and halogen.

33. The compound as claimed in claim 28, wherein Ra2 is selected from the group
consisting of trifluoromethyl, amino, carboxamido, methyl, and fluoro.
34. The compound as claimed in claim 28, wherein Ra1 selected from the group
consisting of fluoro, methyl, propyl, sulfonamido, cyano, nitro, carboxamido, amino,
phenoxy, phenyl, hydroxy, hydroxymethyl, hydroxyethyl,and trifluoromethyl and Ra2 is H.
35. The compound as claimed in claim 18, wherein the compound has an IC50 for
inhibiting the human fatty acid amide hydrolase of less than 1 micromolar.
36. The compound as claimed in claim 18, wherein the compound has an IC50 for
inhibiting the human fatty acid amide hydrolase of less than 10 nanomolar.
37. A pharmaceutical composition comprising the compound as claimed in claim 1,
and a pharmaceutically acceptable carrier, such as herein described.
38. A pharmaceutical composition comprising the compound as claimed in claim 2,
and a pharmaceutically acceptable carrier, such as herein described.
39. A pharmaceutical composition comprising the compound as claimed in claim 3,
and a pharmaceutically acceptable carrier, such as herein described.
40. A pharmaceutical composition comprising the compound as claimed in claim 4,
and a pharmaceutically acceptable carrier, such as herein described.
41. A pharmaceutical composition comprising the compound as claimed in claim 5,
and a pharmaceutically acceptable carrier, such as herein described.
42. A pharmaceutical composition comprising the compound as claimed in claim 6,
and a pharmaceutically acceptable carrier, such as herein described.
43. A pharmaceutical composition comprising the compound as claimed in claim 7,
and a pharmaceutically acceptable carrier, such as herein described.
44. A pharmaceutical composition comprising the compound as claimed in claim 8,
and a pharmaceutically acceptable carrier, such as herein described.
45. A pharmaceutical composition comprising the compound as claimed in claim 9,
and a pharmaceutically acceptable, carrier such as herein described.

46. A pharmaceutical composition comprising the compound as claimed in claim 10,
and a pharmaceutically acceptable carrier, such as herein described.
47. A pharmaceutical composition comprising the compound as claimed in claim 11,
and a pharmaceutically acceptable carrier, such as herein described.
48. A pharmaceutical composition comprising the compound as claimed in claim 12,
and a pharmaceutically acceptable carrier, such as herein described.
49. A pharmaceutical composition comprising the compound as claimed in claim 13,
and a pharmaceutically acceptable carrier, such as herein described.
50. A pharmaceutical composition comprising the compound as claimed in claim 14,
and a pharmaceutically acceptable carrier, such as herein described.
51. A pharmaceutical composition comprising the compound as claimed in claim 15,
and a pharmaceutically acceptable carrier, such as herein described.
52. A pharmaceutical composition comprising the compound as claimed in claim 16,
and a pharmaceutically acceptable carrier, such as herein described.
53. A pharmaceutical composition comprising the compound as claimed in claim 17,
and a pharmaceutically acceptable carrier, such as herein described.
54. A pharmaceutical composition comprising the compound as claimed in claim 18,
and a pharmaceutically acceptable carrier, such as herein described.
55. A pharmaceutical composition comprising the compound as claimed in claim 19,
and a pharmaceutically acceptable carrier, such as herein described.
56. A pharmaceutical composition comprising the compound as claimed in claim 20,
and a pharmaceutically acceptable carrier, such as herein described.
57. A pharmaceutical composition comprising the compound as claimed in claim 21,
and a pharmaceutically acceptable carrier, such as herein described.
58. A pharmaceutical composition comprising the compound as claimed in claim 22,
and a pharmaceutically acceptable carrier, such as herein described.
59. A pharmaceutical composition comprising the compound as claimed in claim 23,

and a pharmaceutically acceptable carrier, such as herein described.
60. A pharmaceutical composition comprising the compound as claimed in claim 24,
and a pharmaceutically acceptable carrier, such as herein described.
61. A pharmaceutical composition comprising the compound as claimed in claim 25,
and a pharmaceutically acceptable carrier, such as herein described.
62. A pharmaceutical composition comprising the compound as claimed in claim 26, f
and a pharmaceutically acceptable carrier, such as herein described.
63. A pharmaceutical composition comprising the compound as claimed in claim 27,
and a pharmaceutically acceptable carrier, such as herein described.
64. A pharmaceutical composition comprising the compound as claimed in claim 28,
and a pharmaceutically acceptable carrier, such as herein described.
65. A pharmaceutical composition comprising the compound as claimed in claim 29,
and a pharmaceutically acceptable carrier, such as herein described.
66. A pharmaceutical composition comprising the compound as claimed in claim 30,
and a pharmaceutically acceptable carrier, such as herein described.
67. A pharmaceutical composition comprising the compound as claimed in claim 31,
and a pharmaceutically acceptable carrier, such as herein described.
68. A pharmaceutical composition comprising the compound as claimed in claim 32,
and a pharmaceutically acceptable carrier, such as herein described.
69. A pharmaceutical composition comprising the compound as claimed in claim 33,
and a pharmaceutically acceptable carrier, such as herein described.
70. A pharmaceutical composition comprising the compound as claimed in claim 34,
and a pharmaceutically acceptable carrier, such as herein described.

71. A pharmaceutical composition comprising the compound as claimed in claim 35,
and a pharmaceutically acceptable carrier, such as herein described.
72. A pharmaceutical composition comprising the compound as claimed in claim 36,
and a pharmaceutically acceptable carrier, such as herein described.


































There is disclosed an N-substituted carbamate meta-biphenyl ester of the formula:

wherein:
X is O
Q is O;
n is 1, 2, or 3;
wherein each Ra member is independently selected from the group consisting of
unsubstituted saturated alkyl, unsubstituted alkenyl, unsubstituted alkynyl,
unsubstituted at least partially saturated cycloalkyl, unsubstituted ketoalkyl,
unsubstituted hydroxyalkyl, unsubstituted aminoalkyl, -CH2-NR3R4,
unsubstituted alkoxy, halo, unsubstituted haloalkyl, cyano, hydroxy, nitro,
amino, -NR3R4, unsubstituted carboxamido, -CONR3R4, sulfonamido,
-SO2NR3R4; -SR5, -SO2R6, -S(O)R6, -NR6C(O)R7, and -NR6SO2R7;
wherein R3 and R4 are independently selected from the group consisting of H,
unsubstituted saturated alkyl, unsubstituted alkenyl, unsubstituted alkynyl,
unsubstituted at least partially saturated cycloalkyl, unsubstituted hydroxyalkyl
and imino-methylamino and wherein optionally R3 and R4 may together with the
N atom to which they are attached combine to form a 5-7 membered ring; and
R5, R6 and R7 are independently selected from the group consisting of hydrogen,
unsubstituted alkyl, and unsubstituted heteroalkyl;
wherein R1 and R2 are independently selected from the group consisting of hydrogen,
substituted or unsubstituted straight or branched alkyl, substituted or
unsubstituted heteroalkyl, substituted or unsubstituted at least partially saturated
cycloalkyl, and substituted or unsubstituted cycloheteroalkyl, with the proviso
that R1 and R2 are not both hydrogen;
and the pharmaceutically acceptable salts thereof.

Documents:

811-kolnp-2005-assignment.pdf

811-kolnp-2005-correspondence.pdf

811-kolnp-2005-examination report.pdf

811-kolnp-2005-form 18.pdf

811-kolnp-2005-form 3.pdf

811-kolnp-2005-form 5.pdf

811-KOLNP-2005-FORM-27.pdf

811-kolnp-2005-gpa.pdf

811-kolnp-2005-granted-abstract.pdf

811-kolnp-2005-granted-claims.pdf

811-kolnp-2005-granted-description (complete).pdf

811-kolnp-2005-granted-drawings.pdf

811-kolnp-2005-granted-form 1.pdf

811-kolnp-2005-granted-specification.pdf

811-kolnp-2005-others.pdf

811-kolnp-2005-reply to examination report.pdf


Patent Number 247104
Indian Patent Application Number 811/KOLNP/2005
PG Journal Number 13/2011
Publication Date 01-Apr-2011
Grant Date 28-Mar-2011
Date of Filing 04-May-2005
Name of Patentee THE REGENTS OF THE UNIVERSITY OF CALIFORNIA
Applicant Address 1111 FRANKLIN STREET, 12TH FLOOR, OAKLAND CA
Inventors:
# Inventor's Name Inventor's Address
1 PIOMELLI, DANIELE 3 VALLEY VIEW, IRVINE CA 92612
2 DURANTI, ANDREA I-URBINO
3 TONTINI, ANDREA I-PESARO
4 MOR, MARCO I-GHEDI
5 TARZIA, GEORGIA I-PETRIANO
PCT International Classification Number C07C 271/00
PCT International Application Number PCT/US2003/031844
PCT International Filing date 2003-10-07
PCT Conventions:
# PCT Application Number Date of Convention Priority Country
1 60/417,008 2002-10-07 U.S.A.