Synthesis and biological evaluation of piperazinyl carbamates and ureas as fatty acid amide hydrolase (FAAH) and transient receptor potential (TRP) channel dual ligands

Article abstract of DOI:10.1016/j.bmcl.2009.09.033

The evaluation of a series of piperazinyl carbamates and ureas, designed on the basis of previously reported TRPV1 antagonists and FAAH inhibitors, led to the identification of some 'dual-action' compounds targeting both FAAH and TRPV1 or TRPA1 receptors.

Full text of DOI:10.1016/j.bmcl.2009.09.033

Bioorganic & Medicinal Chemistry Letters 19 (2009) 6806–6809
Contents lists available at ScienceDirect
Bioorganic & Medicinal Chemistry Letters
journal homepage: www.elsevier.com/locate/bmcl
Synthesis and biological evaluation of piperazinyl carbamates and ureas as
fatty acid amide hydrolase (FAAH) and transient receptor potential (TRP)
channel dual ligands
Enrico Morera a, Luciano De Petrocellis b, Ludovica Morera a, Aniello Schiano Moriello b, Alessia Ligresti c,
Marianna Nalli a, David F. Woodward d, Vincenzo Di Marzo c, Giorgio Ortar a,
*
a Dipartimento di Chimica e Tecnologie del Farmaco, Sapienza Università di Roma, piazzale Aldo Moro 5, 00185 Roma, Italy
b Endocannabinoid Research Group, Istituto di Cibernetica, Consiglio Nazionale delle Ricerche, Via Campi Flegrei 34, 80078, Pozzuoli (NA), Italy
c Endocannabinoid Research Group, Istituto di Chimica Biomolecolare, Consiglio Nazionale delle Ricerche, Via Campi Flegrei 34, 80078, Pozzuoli (NA), Italy
d Department of Biological Sciences, Allergan, Inc., 2525 Dupont Drive, Irvine, CA 92612, USA
a r t i c l e i n f o
a b s t r a c t
Article history:
Received 20 July 2009
Revised 9 September 2009
Accepted 10 September 2009
Available online 20 October 2009
The evaluation of a series of piperazinyl carbamates and ureas, designed on the basis of previously
reported TRPV1 antagonists and FAAH inhibitors, led to the identification of some ‘dual-action’ com-
pounds targeting both FAAH and TRPV1 or TRPA1 receptors.
Ó 2009 Elsevier Ltd. All rights reserved.
Keywords:
Fatty acid amide hydrolase
FAAH
Transient receptor potential channels
TRPV1
TRPA1
Dual-ligands
Piperazinyl carbamates
The endocannabinoid system, which comprises at least two
G-protein coupled receptors (CB1 and CB2), their endogenous
ligands (endocannabinoids), and a series of proteins involved in
endocannabinoid metabolism,1 has emerged as a particularly
promising target for the treatment of wide array of human disor-
ders, in particular neuropathic and inflammatory pain.2 Indeed,
the endocannabinoid N-arachidonoylethanolamine (anandamide,
AEA), and the endocannabinoid-like molecules, N-arachidonoylgly-
cine (NAGly) and N-palmitoylethanolamine (PEA), exert potent
analgesic and anti inflammatory activities in vivo.3
AEA, NAGly, and PEA are all inactivated by enzymatic hydrolysis
catalyzed by amidases, the best characterized of which is the fatty
acid amide hydrolase (FAAH).4 The past 10 years have witnessed
the discovery of an ever increasing number of FAAH inhibitors with
analgesic activity in a variety of pain models through mechanisms
that are not uniquely mediated by cannabinoid receptors.5
The transient receptor potential vanilloid type-1 (TRPV1)
channel represents another possible new answer to the ever
increasing demand for analgesic drugs.6 TRPV1 is a nonselective
cation channel localized in neurons of C- and Ad-sensory fibers
and considered to be a key polymodal integrator of noxious stim-
uli.7 Its activation by noxious heat, acidic pH, plant toxins and
endogenous lipid mediators such as AEA (which can thus be con-
sidered also a true endovanilloid)8 causes calcium influx, neuron
depolarization, and an initial sensation of pain. The analgesic pro-
file generated in animal pain models by TRPV1 receptor genetic
or pharmacological blockade has provided compelling data for
the therapeutic use of TRPV1 antagonists, the identification of
which has become a focus of major attention within the pharma-
ceutical industry.9
A popular and logic approach to developing inhibitors of FAAH
has been the modification of its substrates, and early work in this
area identified a number of such compounds including N-arachid-
onoylserotonin (AA-5-HT).5a AA-5-HT was recently found to also
antagonize TRPV1 receptors, a property that might explain the high
efficacy of this compound against neuropathic pain and anxiety,
thus prompting the development of additional therapeutically use-
ful dual FAAH/TRPV1 blockers.10 Simultaneous blockade of FAAH
and TRPV1 channels might cause analgesic effects stronger then
the targeting of each single system, due to the different respective
roles in the control of nociception.11
* Corresponding author. Tel.: +39 6 49913612; fax: +39 6 491491.
E-mail address: giorgio.ortar@uniroma1.it (G. Ortar).
0960-894X/$ - see front matter Ó 2009 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2009.09.033

E. Morera et al. / Bioorg. Med. Chem. Lett. 19 (2009) 6806–6809
6807
R1
NH
As a continuation of our efforts to identify new molecules able
R1
to target simultaneously both FAAH and TRPV1 receptors,12 we
have selected as lead compounds BCTC,9c the most studied mem-
ber of the piperazine carboxamide class of TRPV1 antagonists,
and PF-622,13 a piperazinyl urea recently described as the proto-
type of a novel mechanistic class of FAAH inhibitors (Fig. 1).
In view of the well documented efficacy of O-arylcarbamates as
FAAH inhibitors,5 a first series of compounds was synthesized in
which the ureic function of BCTC and PF-622 was replaced by a car-
bamate moiety (compounds 1–18). According to the pharmaco-
phore model developed for pyridyl piperazine carboxamides and
related structures that can be traced back to the prototypical
TRPV1 antagonist capsazepine,7b compounds 1–18 should retain
the ability to interact with TRPV1 channels. In fact, two carbamates
structurally related to compounds 1–18 have been claimed in the
patent literature to exhibit TRPV1 antagonism.14 A second series
of compounds was designed through the combination of the pyrid-
inyl (or quinolinyl) piperazine moiety of BCTC and PF-622 with the
serotonin portion of AA-5-HT (compounds 19–22).
N
Cl
a
N
N
23: R1 = Cl
25: R1 = Cl
24: R1 = CF3
26: R1 = CF3
O
R2
R1
N
O
N
b,c
R2
N
OH
1-14
27-33
R1 = Cl, CF3
R2 = 4-t-Bu, 3-t-Bu, 4-CF3, 3-CF3, 4-Cl, 3-Cl, 3-Ph
Scheme 1. Synthesis of compounds 1–14. Reagents and conditions: (a) piperazine,
n-butanol, reflux, 20 h, 91–94%; (b) 20% COCl2 in toluene, Et3N, toluene, 0 °C, then
2 h at 25 °C; (c) 25 or 26, Et3N, DMF–CH2Cl2 1:1, 15 h, 25 °C, 32–63%.
The synthesis of compounds 1–14 is shown in Scheme 1. The
piperazinyl moieties 25 and 26, obtained by condensation of the
commercially available pyridines 23 or 24 with an excess of piper-
azine,15 were reacted with the appropriately substituted phenyl
chloroformates, generated by treatment of the corresponding phe-
nols with phosgene. Compounds 15–18 were obtained by reacting
1-Cbz-piperazine16 with substituted phenyl chloroformates.
Deprotection of the Cbz group by hydrogenolysis afforded the
piperazine derivatives, which were subsequently coupled with
the appropriate 2-chloromethylheterocycles (Scheme 2). Ureas 19
and 20 were synthesized by treatment of the 1-pyridin-2-ylpiper-
azines 25 or 26 with phosgene followed by reaction of the resulting
carbamoyl chlorides with serotonin hydrochloride (Scheme 3).
Finally, the synthesis of compounds 21–22 is outlined in
Scheme 4. The 1-Cbz-piperazinyl chloroformate, obtained by
reaction of 1-Cbz-piperazine with phosgene, was condensed with
serotonin hydrochloride; deprotection of the Cbz group by hydrog-
enolysis and alkylation of the resulting piperazine derivative with
2-(chloromethyl)pyridine and 2-(chloromethyl)quinoline afforded
21 and 22, respectively.17
All compounds were evaluated in assays of FAAH and TRPV1
activity as well as on transient receptor potential ankyrin type-1
(TRPA1). TRPA1 is another member of the transient receptor poten-
tial family of ion channels and is restrictively expressed in sensory
neurons of dorsal root and trigeminal ganglia, and in hair cells of
the inner ear. The TRPA1 channel is activated by a variety of
noxious stimuli including cold temperatures, pungent natural
compounds, and environmental irritants and, like TRPV1, with
which it is very often co-localized, plays an important role in pain
R1
R1
O
a,b,c
d
N
O
OH
N
HN
R1
R1 = CF3, Cl
O
O
R2
N
R2
=
N
15-18
N
Scheme 2. Synthesis of compounds 15–18. Reagents and conditions: (a) 20% COCl2
in toluene, Et3N, toluene, 0 °C, then 2 h at 25 °C; (b) 1-Cbz-piperazine, Et3N, DMF–
CH2Cl2 1:1, 15 h, 25 °C, 47–54%; (c) 10% Pd/C, H2 (1 atm), MeOH, 2 h, 25 °C, 63–78%;
(d) 2-(chloromethyl)pyridine hydrochloride or 2-(chloromethyl)quinoline hydro-
chloride, KI, Et3N, DMF, 80 °C, 15 h, 35–72%.
H
N
O
R
N
N
H
N
a,b
OH
25 or 26
N
19: R = Cl
20: R = CF3
Scheme 3. Synthesis of compounds 19, 20. Reagents and conditions: (a) 20% COCl2
in toluene, Et3N, CH2Cl2, 0 °C, then 3 h at 25 °C; (b) serotonin hydrochloride, Et3N,
DMF–CH2Cl2 1:1, 15 h, 25 °C, 74–77%.
H
N
O
sensing.18 The effects of carbamates 1–18 and ureas 19–22 on
[
N
H
14C]anandamide hydrolysis by rat brain membranes (which ex-
OH
press FAAH as the only AEA hydrolyzing enzyme) and on intracel-
lular Ca2+ elevation mediated by TRPV1 and TRPA1 channels
overexpressed in HEK293 cells, are shown in Table 1.19 Data for
the lead compounds, namely AA-5-HT, BCTC, and PF-622, are also
included in the Table as controls.
AA-5-HT
O
O
The most important results were as follows. Almost all carba-
mates 1–18 showed fairly good FAAH inhibitory activities (2–7
and 9–18). Compounds with meta substituted aromatic rings were
more potent than para substituted analogues, with only one excep-
tion (3 and 4), independent of the nature of the substituent and
similar to previously reported results for AA-5-HT analogues12
and alkylcarbamic acid aryl esters.20 The presence of a methylene
N
N
H
Cl
N
N
H
N
N
N
N
BCTC
PF-622
Figure 1. Structures of some reported FAAH and/or TRPV1 ligands.

6808
E. Morera et al. / Bioorg. Med. Chem. Lett. 19 (2009) 6806–6809
onist than AA-5-HT,12 and compounds 2, 3, 5, 6, 13 and 14, which
H
N
O
exhibited dual inhibitory and stimulatory activity at FAAH and
TRPA1, respectively, and are potentially able to covalently bind
to electron donor aminoacid residues in TRPA1 channels,21 thereby
desensitising them to the action of other nociceptive stimuli. It is
worth mentioning in this context that URB597, a potent FAAH
inhibitor, was also reported to activate TRPA1 channels with a po-
tency 6.6–40.8-fold lower than that of 2, 3, 5, 6, 13 and 14,22 and
that most of TRPA1 agonists so far identified are either highly reac-
tive, nonselective, or not potent or efficacious and, therefore, do
not represent optimal tools for pharmacological studies. Con-
versely, none of the serotonin ureic derivatives (19–22) showed
significant activity on the selected targets. This result was rather
unexpected in view of the potent and selective FAAH inhibitory
activity of PF-622 and of the hypothesis that the aniline moiety
of PF-622 is accommodated in the hydrophilic cytoplasmatic ac-
cess channel of FAAH,13 which should have allowed for a stronger
interaction with the enzyme following introduction of serotonin
moiety.
In conclusion, we have synthesized and assayed a series of pip-
erazinyl carbamates and ureas designed on the basis of previously
reported TRPV1 antagonists and FAAH inhibitors, and have identi-
fied some piperazinyl carbamates that inhibit FAAH with fairly
good potency and interact also with excitatory TRP ionotropic
receptors involved in nociception. Promising hits proved to be in
particular: (a) carbamates 10 and 12, able to act as dual FAAH/
TRPV1 blockers with a potency slightly lower than that of the pro-
totypical ‘hybrid’ blocker AA-5-HT, but with likely higher chemical
stability than this arachidonic acid-containing compound; and (b)
FAAH/TRPA1 ligands 2, 3, 5, 6, 13 and 14, which could conceivably
NH
a,b
N
N
H
BzO
N
BzO
N
OH
O
O
H
N
O
c,d
N
N
H
R
N
N
OH
21: R =
22: R =
N
Scheme 4. Synthesis of compounds 21, 22. Reagents and conditions: (a) 20% COCl2
in toluene, Et3N, CH2Cl2, 0 °C, then 3 h at 25 °C; (b) serotonin hydrochloride, Et3N,
DMF–CH2Cl2 1:1, 15 h, 25 °C, 100%; (c) 10% Pd/C, H2 (1 atm), MeOH/AcOEt 1:1, 3 h,
25 °C, 72%; (d) 2-(chloromethyl)pyridine hydrochloride or 2-(chloromethyl)quino-
line hydrochloride, KI, Et3N, DMF, 80 °C, 15 h, 27–47%.
group between the piperazine and the pyridine (or quinoline) rings
did not affect critically the FAAH inhibitory activity. Some of the
carbamates also exhibited significant TRPV1 antagonist (1, 4, 8,
10, 12) and/or TRPA1 agonist (1–6, 13, 14) activity. A stronger
TRPV1 antagonist activity was observed with para substituted
derivatives (in particular 1 and 8). Among carbamates, particularly
interesting appears to be compound 10, which was 2.4-fold more
potent as FAAH inhibitor and 3.7-fold less potent as TRPV1 antag-
Table 1
Results of FAAH, TRPV1, and TRPA1 assays of carbamates 1–18 and ureas 19–22a
H
O
N
R2
O
N
O
N
N
H
N
R1
N
R1
OH
19-22
1-18
Compound
R1
R2
AEA hydrolysis (IC50
,
l
M)
% Inhibition (c = 50
lM)
TRPV1 (IC50
,
l
M)
TRPA1 (EC50, lM)
1
2
3
4
5
6
7
8
9
10
11
12
13
14
15
16
17
18
19
20
21
22
AA-5-HT
BCTC
PF-622
3-Chloropyridin-2-yl
3-Chloropyridin-2-yl
3-Chloropyridin-2-yl
3-Chloropyridin-2-yl
3-Chloropyridin-2-yl
3-Chloropyridin-2-yl
3-Chloropyridin-2-yl
3-Trifluoromethylpyridin-2-yl
3-Trifluoromethylpyridin-2-yl
3-Trifluoromethylpyridin-2-yl
3-Trifluoromethylpyridin-2-yl
3-Trifluoromethylpyridin-2-yl
3-Trifluoromethylpyridin-2-yl
3-Trifluoromethylpyridin-2-yl
Pyridin-2-ylmethyl
Quinolin-2-ylmethyl
Pyridin-2-ylmethyl
Quinolin-2-ylmethyl
3-Chloropyridin-2-yl
4-t-Bu
3-t-Bu
4-CF3
3-CF3
4-Cl
19.19
1.27
3.90
6.56
6.44
0.91
0.75
30.11
0.77
3.36
0.61
2.19
0.74
0.38
2.9
9.3
4.1
5.9
>50
>50
>50
>50
8.0b
>50
0.50
72.9
97.5
97.4
98.3
78.6
97.7
97.7
63.1
97.0
69.4
98.2
97.5
98.5
97.4
92.4
98.0
90.1
90.0
21.5
35.4
34.0
32.6
92.0
24.6
98.8
0.48
>10
>10
3.9
>10
>10
>10
0.26
>10
1.0
1.6
1.8
3.1
2.0
0.6
1.0
>25
>25
>25
>25
>25
>25
3.7
3-Cl
3-Ph
4-t-Bu
3-t-Bu
4-CF3
3-CF3
4-Cl
3-Cl
3-Ph
4-CF3
4-CF3
4-Cl
>10
4.6
>10
>10
>10
>10
>10
>10
>10
>10
>10
>10
0.27b
0.0067
>10
1.0
>25
>25
>25
>25
>25
>25
>25
>25
>25
4.5
4-Cl
3-Trifluoromethylpyridin-2-yl
Pyridin-2-ylmethyl
Quinolin-2-ylmethyl
>25
a
Data are means of n = 4 separate determinations. Standard errors are not shown for the sake of clarity and were never higher than 10% of the means.
Data from Ref. 12.
b

E. Morera et al. / Bioorg. Med. Chem. Lett. 19 (2009) 6806–6809
6809
16. Löwik, D. W. P. M.; Lowe, C. R. Eur. J. Org. Chem. 2001, 2825.
17. Data for selected compounds: Compound 6: yield 63%; oil; IR (CHCl3) 2925,
2847, 1717, 1578, 1421, 1238 cmÀ1 1H NMR (200 MHz, CDCl3) d 3.38–3.43 (m,
represent useful pharmacological tools for the study of TRPA1
channels.
;
4H), 3.74–3.82 (m, 4H), 6.89 (dd, J = 7.6, 4.6 Hz, 1H), 7.02–7.08 (m, 1H), 7.17–
7.21 (m, 2H), 7.27 (d, J = 8 Hz, 1H), 7.62 (dd, J = 7.8, 1.6 Hz, 1H), 8.20 (dd, J = 4.8,
1.6 Hz, 1H); 13C NMR (50 MHz, CDCl3) d 44.13, 44.51, 48.89, 118.54, 120.11,
122.43, 122.86, 125.58, 129.94, 134.53, 138.92, 145.93, 151.93, 153.23, 158.05.
Compound 10: yield 43%; mp 106–109 °C; IR (KBr) 2921, 2869, 1717, 1591,
References and notes
1. (a) De Petrocellis, L.; Di Marzo, V. Best Pract. Clin. Endocrinol. Metab. 2009, 23, 1;
(b) Alexander, S. P. H.; Kendall, D. A. Br. J. Pharmacol. 2007, 152, 602; (c) Fride,
E.; Gobshtis, N. Immunol. Endocr. Metabol. Agents Med. Chem. 2007, 7, 157; (d)
Lambert, D. M.; Fowler, C. J. J. Med. Chem. 2005, 48, 5059.
2. (a) Di Marzo, V. Nat. Rev. Drug Disc. 2008, 7, 438; (b) Jhaveri, M. D.; Richardson,
D.; Chapman, V. Br. J. Pharmacol. 2007, 152, 624.
3. Bradshaw, H. B.; Walker, J. M. Br. J. Pharmacol. 2005, 144, 459.
4. (a) Bracey, M. H.; Hanson, M. A.; Masuda, K. R.; Stevens, R. C.; Cravatt, B. F.
Science 2002, 298, 1793; (b) Patricelli, M. P.; Cravatt, B. F. Vitam. Horm. 2001, 62,
95; (c) Giang, D. K.; Cravatt, B. F. Proc. Natl. Acad. Sci. U.S.A. 1997, 94, 2238; (d)
Cravatt, B. F.; Giang, D. K.; Mayfield, S. P.; Boger, D. L.; Lerner, R. A.; Gilula, N. B.
Nature 1996, 384, 83.
5. (a) Seierstad, M.; Breitenbucher, G. J. Med. Chem. 2008, 51, 7327; (b)
Vandervoorde, S. Curr. Top. Med. Chem. 2008, 8, 247; (c) Maccarrone, M. Curr.
Pharm. Des. 2006, 12, 759.
6. (a) Patapoutian, A.; Tate, S.; Woolf, C. J. Nat. Rev. Drug Disc. 2009, 8, 55; (b)
Westway, S. M. J. Med. Chem. 2007, 50, 2589.
7. (a) Vennekens, R.; Owsianik, G.; Nilius, B. Curr. Pharm. Des. 2009, 14, 18; (b)
Szallasi, A.; Cortright, D. N.; Blum, C. A.; Eid, S. R. Nat. Rev. Drug Disc. 2007, 6,
357.
8. Starowicz, K.; Nigam, S.; Di Marzo, V. Pharmacol. Ther. 2007, 114, 13.
9. (a) Gunthorpe, M. J.; Szallasi, A. Curr. Pharm. Des. 2008, 14, 32; (b) Broad, L. M.;
Keding, S. J.; Blanco, M. J. Curr. Top. Med. Chem. 2008, 8, 1431; (c) Gharat, L.;
Szallasi, A. Drug Dev. Res. 2007, 68, 477.
10. Maione, S.; De Petrocellis, L.; de Novellis, V.; Schiano Moriello, A.; Petrosino, S.;
Palazzo, E.; Rossi, F.; Woodward, D. F.; Di Marzo, V. Br. J. Pharmacol. 2007, 150,
766.
1450, 1417, 1324, 1212 cmÀ1 1H NMR (400 MHz, CDCl3) d 3.33 (br s, 4H),
;
3.73–3.83 (m, 4H), 7.09 (dd, J = 7.8, 4.8 Hz, 1H), 7.26 (d, J = 8.2 Hz, 4H), 7.64 (d,
J = 8.4 Hz), 7.92 (dd, J = 7.8, 1.6 Hz, 1H), 8.48 (dd, J = 4.6, 1.6 Hz, 1H); 13C NMR
(100 MHz, CDCl3) d 44.19, 44.76, 50.68, 117.83 (q, J = 31 Hz), 118.10, 122.14,
123.59 (q, J = 271 Hz), 123.82 (q, J = 271 Hz), 126.68, 127.69 (q, J = 33 Hz),
137.26, 151.30, 153.03, 153.94, 159.78. Compound 12: yield 55%; mp 103–
105 °C; IR (KBr) 2904, 2865, 1715, 1590, 1577, 1444, 1417, 1249, 1218,
1121 cmÀ1 1H NMR (400 MHz, CDCl3) d 3.32–3.33 (m, 4H), 3.71–3.81 (m, 4H),
;
7.07 (d, J = 8.8 Hz, 2H), 7.33 (d, J = 8.8 Hz, 2H), 7.91 (dd, J = 7.8, 1.6 Hz, 1H), 8.47
(dd, J = 4.6, 1.6 Hz, 1H); 13C NMR (100 MHz, CDCl3) d 44.13, 44.66, 50.67,
117.76 (q, J = 31 Hz), 118.00, 123.12, 123.82 (q, J = 271 Hz), 129.33, 130.70,
137.26, 149.89, 151.26, 153.45, 159.79. Compound 13: yield 45%; oil; IR
(CHCl3) 2927, 2850, 1718, 1592, 1445, 1421, 1311, 1238 cmÀ1 1H NMR
;
(400 MHz, CDCl3) d 3.32 (br s, 4H), 3.72–3.80 (m, 4H), 7.04–7.30 (m, 5H), 7.91
(dd, J = 7.8, 1.6 Hz, 1H), 8.47 (dd, J = 4.6, 1.6 Hz, 1H); 13C NMR (100 MHz, CDCl3)
d 44.12, 44.68, 50.66, 117.72 (q, J = 31 Hz), 118.02, 120.15, 122.45, 123.68 (q,
J = 271 Hz), 125.63, 130.00, 134.52, 137.21, 151.27, 151.89, 153.24, 159.75.
Compound 14: yield 45%; oil; IR (CHCl3) 2927, 2850, 1709, 1592, 1446, 1311,
1240, 1183 cmÀ1 1H NMR (400 MHz, CDCl3) d 3.32 (br s, 4H), 3.73–3.83 (m,
;
4H), 7.02 (dd, J = 7.6, 4.6 Hz, 1H), 7.11–7.13 (m, 1H), 7.31–7.42 (m, 6H), 7.57–
7.59 (m, 2H), 7.88 (dd, J = 7.8, 1.6 Hz, 1H), 8.45 (dd, J = 4.6, 1.6 Hz, 1H); 13C NMR
(100 MHz, CDCl3) d 44.07, 44.66, 50.69, 117.77 (q, J = 31 Hz), 117.90, 120.53,
122.01, 123.88 (q, J = 271 Hz), 124.11, 127.15, 127.57, 128.75, 129.57, 137.21,
140.31, 142.69, 151.24, 151.77, 153.79, 159.77.
18. (a) Karashima, Y.; Talavera, K.; Everaerts, W.; Janssens, A.; Kwan, K. Y.;
Vennekens, R.; Niluis, B.; Voets, T. Proc. Natl. Acad. Sci. U.S.A. 2009, 106, 1273;
(b) Tai, C.; Zhu, S.; Zhou, N. J. Neurosci. 2008, 28, 1019.
19. Detailed procedures for assays of FAAH, TRPV1, and TRPA1 activities were
previously reported. See Ref. 12 and: Ortar, G.; Schiano Moriello, A.; Cascio, M.
G.; De Petrocellis, L.; Ligresti, A.; Morera, E.; Nalli, M.; Di Marzo, V. Bioorg. Med.
Chem. Lett. 2008, 18, 2820.
11. (a) Fowler, C. J.; Naidu, P. S.; Lichtman, A.; Onnis, V. Br. J. Pharmacol. 2009, 156,
412; For a review on ‘designed multiple ligands’, see: (b) Morphy, R.; Rankovic,
Z. J. Med. Chem. 2005, 48, 6523.
12. Ortar, G.; Cascio, M. G.; De Petrocellis, L.; Morera, E.; Rossi, F.; Schiano Moriello,
A.; Nalli, M.; de Novellis, V.; Woodward, D. F.; Maione, S.; Di Marzo, V. J. Med.
Chem. 2007, 50, 6554.
13. Ahn, K.; Johnson, D. S.; Fitzgerald, L. R.; Liimatta, M.; Arensde, A.; Stevenson, T.;
Lund, E. T.; Nugent, R. A.; Nomanbhoy, T. K.; Alexander, J. P.; Cravatt, B. F.
Biochemistry 2007, 46, 13019.
20. Tarzia, G.; Duranti, A.; Tontini, A.; Piersanti, G.; Mor, M.; Rivara, S.; Plazzi, P. V.;
Park, C.; Kathuria, S.; Piomelli, D. J. Med. Chem. 2003, 46, 2352.
21. (a) Macpherson, L. J.; Dubin, A. E.; Evans, M. J.; Marr, F.; Schultz, P. G.; Cravatt,
B. F.; Patapoutian, A. Nature 2007, 445, 541; (b) Hinman, A.; Chuang, H. H.;
Bautista, D. M.; Julius, D. Proc. Natl. Acad. Sci. U.S.A. 2006, 103, 19564.
22. Niforatos, W.; Zhang, X.-F.; Lake, M. R.; Walter, K. A.; Neelands, T.; Holzman, T.
F.; Scott, V. E.; Faltynek, C. R.; Moreland, R. B.; Chen, J. Mol. Pharmacol. 2007, 71,
1209.
14. Bakthavatchalam, R. WO Patent 0208221, 2002.
15. Swanson, D. M.; Dubin, A. E.; Shah, C.; Nasser, N.; Chang, L.; Dax, S. L.; Jetter,
M.; Breitenbucher, J. G.; Liu, C.; Mazur, C.; Lord, B.; Gonzales, L.; Hoey, K.;
Rizzolio, M.; Bogenstaetter, M.; Codd, E. E.; Lee, D. H.; Zhang, S.-P.; Chaplan, S.
R.; Carruthers, N. I. J. Med. Chem. 2005, 48, 1857.