Delineation of a fundamental α-ketoheterocycle substituent effect for use in the design of enzyme inhibitors

Article abstract of DOI:10.1021/ja064522b

The synthesis and examination of a systematic series of 5-substituted 2-keto oxazoles as inhibitors of fatty acid amide hydrolase (FAAH) defined a fundamental substituent effect that led to the discovery of inhibitors with K_i's as low as 400 pM. The intrinsic basis of the relationship (-log K_i vs σ_p), which relates K_i with the Hammett σ_p constant of the substituent, the magnitude of the effect (ρ = 3.01), and its predictive value (R² = 0.91) suggest a widespread applicability in studies beyond FAAH. Copyright

Full text of DOI:10.1021/ja064522b

Published on Web 10/11/2006
Delineation of a Fundamental r-Ketoheterocycle Substituent Effect for Use in
the Design of Enzyme Inhibitors
F. Anthony Romero, Inkyu Hwang, and Dale L. Boger*
Department of Chemistry and the Skaggs Institute for Chemical Biology,
The Scripps Research Institute, 10550 North Torrey Pines Road, La Jolla, California 92037
Received June 26, 2006; E-mail: boger@scripps.edu
Anandamide1 (1a) and oleamide2-4 (1b) have emerged as the
prototypical members of a class of endogenous fatty acid amides
that serve as chemical messengers (Figure 1). Anandamide binds
and activates the central (CB1) and peripheral (CB2) cannabinoid
receptors where it has been implicated in the modulation of
nociception, feeding, and anxiety.5 Oleamide was found to ac-
cumulate in the cerebrospinal fluid of animals under conditions of
sleep deprivation and induces physiological sleep in a dose-
dependent manner.3,4 The pharmacological action of both ananda-
mide and oleamide is terminated by the enzyme fatty acid amide
hydrolase (FAAH, Figure 1).6-9 It degrades neuromodulating fatty
acid amides at their site of action and is currently the only
Figure 1. Substrates of fatty acid amide hydrolase (FAAH).
characterized mammalian enzyme that is in the amidase signature
family bearing an unusual catalytic Ser-Ser-Lys triad.6-9 As such,
FAAH has emerged as an exciting therapeutic target for a range of
clinical disorders.5,10
Scheme 1
One class of FAAH inhibitors that exhibit potent and selective
enzyme inhibition and in vivo efficacy is the R-ketoheterocycles.11-16
Since their introduction by Edwards,17-19 the use of such R-keto-
heterocycles has emerged as a powerful design concept for the
development of inhibitors of serine and cysteine proteases and
hydrolases.20 Possessing electrophilic carbonyls, they reversibly
form enzyme-stabilized covalent hemiketals or hemithioketals with
the enzyme catalytic nucleophile analogous to more conventional
aldehyde,21 trifluoromethyl ketone,22 or R-keto ester23 and amide
inhibitors.24 By virtue of the ability of the heterocycle to hydrogen
bond to the adjacent hemiketal of the enzyme adduct17-19 and
because of interactions of the heterocycle itself with the enzyme
active site independent of its role in activating the carbonyl,13 they
offer advantages over the simpler predecessors. Although the
potency of such R-ketoheterocycles has been anticipated to be
related to the intrinsic electron-withdrawing properties of the
heterocycles,17-19 attempts to draw such correlations are weak,
accompanied by deviations from expectations, and the more potent
heterocycles in a series are empirically derived.25
Herein, we report the synthesis and evaluation of a series of
5-substituted 7-phenyl-1-(oxazol-2-yl)heptan-1-ones that define an
alternative and fundamental R-ketoheterocycle substituent effect that
led to the discovery of FAAH inhibitors with Ki’s as low as 400
pM. Its intrinsic basis, which relates Ki with the Hammett σp
constant of the substituent, as well as the magnitude of the effect
(F ) 3.01), and its predictive value (R2 ) 0.91) suggest a
widespread applicability in studies beyond FAAH inhibition.
Key to the divergent synthesis of the inhibitors was the
preparation of intermediate 311 from which all the compounds could
be derived (Scheme 1). Intermediate 3 was obtained by Vedejs
oxazole metalation,26 condensation with 7-phenylheptanal, and TBS
protection of the resulting alcohol. Selective C5 lithiation27 of 3
followed by treatment with various electrophiles (CO2(g), CF3-
CONMe2, CH3CONMe2, DMF, I2, Br2, NCS, N-fluorobenzene-
sulfonimide, CH3I, (MeS)2) afforded 4b, 4f-h, 4j, and 4l-p, many
of which served as precursors to additional candidate inhibitors
bearing further modified C5 substituents. Carboxylic acid 5b was
directly converted to its corresponding methyl ester 5c by treatment
with TMSCHN2. The ester 5c was converted to the carboxamide
5d by treatment with methanolic ammonia, which, in turn, was
dehydrated with TFAA and pyridine to provide nitrile 5i. Using a
method developed by Chen et al., iodide 4l was transformed to 4k
(FSO2CF2CO2CH3, CuI) bearing a C5 trifluoromethyl substitu-
ent.28,29 In each case, deprotection of the TBS ether followed by
Dess-Martin periodinane oxidation30 of the liberated alcohol
yielded the corresponding R-ketoheterocycles.
This series, which constitutes a set of relatively small substituents
that can occupy accessible space in the FAAH active site, exhibited
FAAH inhibition that tracked with the electron-withdrawing proper-
ties of the substituents (Figure 2). A plot of the inhibition (-log
Ki) versus the Hammett σp constant for the substituents (Figure 3)
was found to follow a well-defined correlation (F ) 3.01, R2 )
0.91). In addition, this substituent effect was established to be large
(F ) 3.01), resulting in a 1000-fold increase in Ki per unit change
in σp and indicating that the electronic character of the substituent
is the dominant factor contributing to the differences in binding
affinity. Presumably, this arises from the increased electrophilic
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10.1021/ja064522b CCC: $33.50 © 2006 American Chemical Society

C O M M U N I C A T I O N S
(-log Ki) and the magnitude of the effect (F ) 3.01) provides a
useful new predictive tool for the rational design of serine and
cysteine protease and hydrolase inhibitors.
Acknowledgment. We gratefully acknowledge the financial
support of the National Institutes of Health (DA15648) and the
Skaggs Institute for Chemical Biology, and the postdoctoral
fellowship support for F.A.R. (American Cancer Society).
Supporting Information Available: Full experimental details and
characterization, and FAAH assay measurement errors of the inhibitors
disclosed herein. This material is available free of charge via the Internet
at http://pubs.acs.org.
Figure 2. FAAH inhibition. Ki measurement errors are provided in
Supporting Information.
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Figure 3. Value of -log Ki (µM) versus σp.
character of the C2 carbonyl imparted by the electron-withdrawing
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1
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(33) An example is provided in Supporting Information.
The delineation of a fundamental correlation that relates the
Hammett σp constant of a substituent with its enzyme inhibition
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