Discovery of N-Aryl-2-acylindole human glucagon receptor antagonists

Article abstract of DOI:10.1016/j.bmcl.2011.09.105

A novel class of N-aryl-2-acylindole human glucagon receptor (hGCGR) antagonists is reported. These compounds demonstrate good pharmacokinetic profiles in multiple preclinical species. One compound from this series, indole 33, is orally active in a transgenic murine pharmacodynamic model. Furthermore, a 1 mg/kg oral dose of indole 33 lowers ambient glucose levels in an ob/ob/hGCGR transgenic murine diabetes model. This compound was deemed suitable for preclinical safety studies and was found to be well tolerated in an 8-day experimental rodent tolerability study. The combination of preclinical efficacy and safety observed with compound 33 highlights the potential of this class as a treatment for type 2 diabetes.

Full text of DOI:10.1016/j.bmcl.2011.09.105

Bioorganic & Medicinal Chemistry Letters 21 (2011) 7124–7130
Contents lists available at SciVerse ScienceDirect
Bioorganic & Medicinal Chemistry Letters
journal homepage: www.elsevier.com/locate/bmcl
Discovery of N-Aryl-2-acylindole human glucagon receptor antagonists
⇑
Christopher Sinz , Amy Bittner, Ed Brady, Mari Candelore, Qing Dallas-Yang, Victor Ding, Guoqiang Jiang,
Zhen Lin, Sajjad Qureshi, Gino Salituro, Richard Saperstein, Jackie Shang, Deborah Szalkowski,
Laurie Tota, Stella Vincent, Michael Wright, Shiyao Xu, Xiaodong Yang, Bei Zhang, James Tata,
Ronald Kim, Emma R. Parmee
Discovery and Preclinical Sciences, Merck Research Laboratories, PO Box 2000, Rahway, NJ 07065, USA
a r t i c l e i n f o
a b s t r a c t
Article history:
Available online 5 October 2011
A novel class of N-aryl-2-acylindole human glucagon receptor (hGCGR) antagonists is reported. These
compounds demonstrate good pharmacokinetic profiles in multiple preclinical species. One compound
from this series, indole 33, is orally active in a transgenic murine pharmacodynamic model. Furthermore,
a 1 mg/kg oral dose of indole 33 lowers ambient glucose levels in an ob/ob/hGCGR transgenic murine dia-
betes model. This compound was deemed suitable for preclinical safety studies and was found to be well
tolerated in an 8-day experimental rodent tolerability study. The combination of preclinical efficacy and
safety observed with compound 33 highlights the potential of this class as a treatment for type 2
diabetes.
Keywords:
Glucagon
Diabetes
Glucose
Antagonist
Indole
Ó 2011 Elsevier Ltd. All rights reserved.
Glucagon and insulin are the primary counter-regulatory
hormones responsible for maintaining glucose homeostasis.1 In re-
sponse to falling plasma glucose levels, glucagon is secreted from
core structure. We thus synthesized 2-acylindole 2, a non-basic
heterocycle designed to display the critical pharmacophores in a
similar fashion as compound 1. This compound class indeed
showed improved selectivity over the hERG channel, but provided
compounds with poor solubility. Varying the display of the hydro-
phobic groups on the acylindole scaffold and replacing the amino-
tetrazole amide moiety with a b-alanine amide led to indole 3.
Compounds of this type showed enhanced functional activity
against the human glucagon receptor relative to indole 2, and also
generally displayed improved rat pharmacokinetic profiles relative
to compounds typified by 1. A further improvement in rat pharma-
cokinetic profile was realized upon conversion of the N-benzyl in-
dole to an N-aryl indole, as in compound 4. While this change led
to a small loss in hGCGR binding potency, the promising rodent PK
profile prompted further development of this novel series and ulti-
mately led to the discovery of a highly potent, selective hGCGR
antagonist with a profile suitable for evaluation in preclinical
safety studies.
A synthetic route which allows for late stage introduction of a
variety of N-aryl groups is depicted below (Scheme 1). Commer-
cially available indole 5 was converted to the corresponding
Weinreb amide. Exposure of this amide to an excess of an alkyllith-
ium reagent such as n-butyllithium provided ketone 6. A sequence
involving protection of the indole nitrogen with a benzyl group to
give indole 7, ketone enolate alkylation and indole deprotection5
gave the key intermediate 8. From this intermediate, a wide variety
of aryl and heteroaryl groups could be introduced by Buchwald’s
method,6 giving rise to N-arylindoles exemplified by 9. Saponifica-
tion followed by EDC-mediated amide bond formation with a
pancreatic a-cells. This hormone activates its receptor, a G-protein
coupled receptor located predominately in the liver, leading to in-
creased hepatic glucose production via glycogenolysis and gluco-
neogenesis. In patients with type 2 diabetes, inappropriately
elevated glucagon levels result in excessive hepatic glucose output,
a key contributor to hyperglycemia. This has stimulated efforts to
develop various strategies targeting reduced glucagon action. To-
ward this end, both antisense oligonucleotides2 and small mole-
cule glucagon receptor antagonists3,4 have been reported. Herein
we report a novel class of acyl indole human glucagon receptor
(hGCGR) antagonists which evolved from our earlier work on cyclic
guanidine hGCGR antagonists.4
We recently disclosed a cyclic guanidine class of human gluca-
gon receptor (hGCGR) antagonists, exemplified by compound 1
(Fig. 1).4b Compound 1 showed excellent efficacy in a murine glu-
cagon challenge pharmacodynamic model, but displayed poor
pharmacokinetic (PK) profiles across preclinical species. Many ana-
logs also carried unacceptable off-target activities, often displaying
potent binding of IKr, the ion channel derived from hERG. As we at-
tempted to address these issues within the cyclic guanidine series,
we initiated a parallel effort to identify a novel scaffold based on
the cyclic guanidine design. We hypothesized that the suboptimal
off-target profile could be attributed to Bronsted basicity of the
⇑
Corresponding author.
E-mail address: christopher_sinz@merck.com (C. Sinz).
0960-894X/$ - see front matter Ó 2011 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2011.09.105

C. Sinz et al. / Bioorg. Med. Chem. Lett. 21 (2011) 7124–7130
7125
OCF3
OCF3
Cl
Cl
N
N
N
N
O
F3C
H
N
H
N
HN
HN
N
O
N
N
O
N
N
N
2
1
hGCGR IC50 = 51 nM
Rat PK Profile:
Cl = 10 ml/min/kg
F = 40%
hGCGR IC50 = 19 nM
Rat PK Profile:
Cl = 16 ml/min/kg
F = 8%
O
O
NH
HO
NH
O
O
Cl
Cl
HO
N
O
N
O
3
4
hGCGR IC50 = 52 nM
Rat PK Profile:
Cl = 4.7 ml/min/kg
F = 38%
hGCGR IC50 = 16 nM
Rat PK Profile:
Cl = 13 ml/min/kg
F = 24%
Figure 1. Evolution of N-aryl acylindole glucagon receptor antagonists.
protected b-alanine gave amides such as 10. At this point, enantio-
pure compounds could be obtained via chiral HPLC.7 Finally, TFA-
mediated ester deprotection provided the target compounds such
as 4.
the a-carbonyl substituent held constant. Compound 4, which
bears a sterically bulky t-butyl group at the 4-position of the aro-
matic ring, showed moderate activity in both the binding and func-
tional assays. Replacing the t-butyl substituent with a 4-chloro
substituent, as in 14, led to an improvement in binding affinity
but a concomitant decrease in functional activity. Modest increases
in both binding affinity and functional activity were realized upon
introduction of sterically bulky, electron withdrawing trifluoro-
methoxy and trifluoromethyl groups as in 15 and 16. Shifting these
substituents to the 3-position of the aromatic ring (compounds 17
and 18) provided compounds of comparable activity.
While 3,5-disubstituted N-aryl groups led to decreased activity
(e.g., compounds 20 and 21), certain 3,4-disubstituted N-aryl
groups such as in compound 23 provided enhanced potency.
Replacing the N-phenyl groups with substituted naphthyl rings
or other [6.6]-systems as in compounds 24 and 25, failed to pro-
vide an increase in activity. While the quinolyl group of 26 was
poorly tolerated, the less basic 3-trifluoromethyl-2-pyridyl group
present in compound 27 led to significantly enhanced binding
affinity.
For certain
a-keto groups either incompatible with the Lewis
acid promoted indole deprotection (Scheme 1, step e) or not ame-
nable to alkyllithium formation (Scheme 1, step b), an alternative
approach was employed (Scheme 2). In one example, Claisen con-
densation of t-butyl acetate with indole 11 followed by C-alkyl-
ation with bromomethyl cyclopropane provided b-keto ester 12.
Acidic cleavage of the t-butyl ester and thermal decarboxylation
gave ketone 13, which could be processed to the target hGCGR
antagonist as described in Scheme 1.
The in vitro biological activities of these compounds were eval-
uated in both binding and functional assays. Binding affinity was
determined by a scintillation proximity assay measuring the inhi-
bition of I125-labelled glucagon binding to the hGCGR expressed on
CHO cell membranes (hGCGR BND IC50).8 Compounds demonstrat-
ing good affinity (IC50 <100 nM) were tested for functional inhibi-
tion of glucagon-induced cAMP production in hGCGR-transfected
CHO cells (hGCGR cAMP IC50).8
These chiral hGCGR antagonists showed considerable differ-
ences in activity as a function of absolute configuration. For exam-
ple, while compound 27 was a highly potent antagonist (hGCGR
Table 1 summarizes hGCGR binding and functional data for a
series of compounds displaying a range of N-aryl substituents, with

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C. Sinz et al. / Bioorg. Med. Chem. Lett. 21 (2011) 7124–7130
a, b
Cl
c, d
OH
O
Cl
Cl
N
H
N
H
O
N
O
5
6
7
CO2Me
CO2Me
e
f
Cl
Cl
N
O
N
O
H
8
9
O
O
NH
NH
O
O
O
HO
f, g
h, i, j
Cl
Cl
N
O
N
O
10
4
Scheme 1. Reagents and conditions: (a) MeONHOMeÁHCl, EDC, HOBt, DIEA, DMF, 40 °C; (b) n-BuLi, THF, À78 °C to 0 °C; (c) NaH, BnBr, DMF, rt; (d) KHMDS, methyl 4-
bromomethylbenzoate, À78 °C to rt; (e) AlCl3, benzene, 0 °C to rt; (f) 4-tBu-iodobenzene, CuI, K3PO4, trans,N,N-dimethyl-1,2-cyclohexanediamine, toluene, 110 °C; (g) LiOH,
dioxane/water; (h) b-alanine-OtBuÁHCl, EDC, HOBt, DIEA; (i) chiral separation; for further details, see Ref. 7; (j) TFA, CH2Cl2.
O
Cl
a, b
Cl
O
O
Cl
c, d
O
N
N
O
N
O
F3C
F3C
F3C
12
11
13
Scheme 2. Reagents and conditions: (a) tBuOAc, LHMDS, THF, À78 °C to 0 °C; (b) NaH, bromomethylcyclopropane, DMF, rt; (c) TFA, CH2Cl2, rt; (d) benzene, 90 °C.
IC50 = 9.2 nM), the corresponding enantiomer 28 was nearly fifty-
fold less potent (Scheme 3). This prompted us to elucidate the
absolute configuration of the more potent enantioseries. Toward
this end, hydrolysis of ester 8, EDC coupling with b-alanine t-butyl
ester and chiral separation gave rise to compound 29, a single
enantiomer of unknown configuration (Scheme 3). Buchwald ary-
lation6 with 2-bromo-6-trifluoromethylpyridine followed by
deprotection of the b-alanine t-butyl ester gave compound 27,
indicating that indole 29 belongs to the more potent enantioseries.
Treatment of 29 with excess mCPBA gave rise to Baeyer–Villiger
type oxidation9 with in situ hydrolysis, providing a carboxylic acid
which could be coupled with (R,R)-pseudoephedrine to give amide
30. Intermediate 30 made possible the elucidation of stereochem-
istry by chemical correlation, as this compound could be synthe-
sized by a well precedented route (Scheme 4). Thus, Myers
alkylation10 of (R,R)-pseudoephedrine valeramide (31) with methyl
4-bromomethylbenzoate, saponification and amide bond forma-
tion with b-alanine t-butyl ester gave compound 30, thereby estab-
lishing the (S)-configuration for the more potent enantioseries.11
Since installation of an N-3-trifluoromethylphenyl group led to
potent compounds with favorable pharmacokinetic profiles (vide
infra), this substituent was held constant in a survey of a-carbonyl
alkyl groups (Table 2). While increasing the alkyl chain length from
n-propyl to n-butyl had no discernible effect (cf., compounds 18 and
32), the n-pentyl homologue 33 showed improved binding and
functional activity. Replacing the n-pentyl group with its isopentyl
isomer as in 34 led to a three-fold reduction in functional activity.
Likewise, introduction of the cyclopropylmethyl group in com-
pound 35 resulted in decreased activity. Thus, the combination of
N-3-trifluoromethylphenyl and
a-keto n-pentyl moieties in com-
pound 33 appeared optimal. Efforts to further optimize in vitro
activity by varying substitution at the 5-position of the indole
served to confirm the 5-chloroindole fragment as providing the
optimal combination of binding and functional activity (Table 3).

C. Sinz et al. / Bioorg. Med. Chem. Lett. 21 (2011) 7124–7130
7127
Table 1
SAR of N-aryl substituenta
O
NH
OH
O
Cl
N
R
O
Compound
R
hGCGR BNDb IC50 (nM) (n)
hGCGR cAMPb IC50 (nM) (n)
4
58
52 11 (2)
100 11 (2)
45 6.5 (2)
14
15
32
Cl
28 4.7 (2)
F3CO
16
17
31 3.7 (2)
24 3.5 (2)
30 13 (2)
F3C
35
OCF3
18
17 6.4 (2)
55 7.0 (2)
CF3
F
19
20
21
47
150 32 (2)
Cl
110
120
n/d
n/d
Cl
Me
Me
F3C
22
23
24
25
26
23 8.5 (2)
12 5.4 (2)
34
39 4.5 (2)
17 1.1 (2)
95 55 (2)
49 5.7 (2)
MeO
F3C
Cl
OMe
O
O
28 9.2 (2)
58 19 (3)
9.2 0.8 (2)
718
N
F3C
N
27
34 5.4 (5)
a
All compounds are single enantiomers which were resolved via chiral HPLC (see footnote 7 for details); data shown are for the enantiomer which is the more potent
hGCGR antagonist.
b
Each IC50 in this Letter is reported as an average rounded to two significant figures standard error of the mean when more than one measurement was made.

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C. Sinz et al. / Bioorg. Med. Chem. Lett. 21 (2011) 7124–7130
O
O
NH
NH
O
O
Cl
HO
Cl
HO
N
O
N
O
N
N
CF3
28
hGCGR IC50 = 450 nM
CF3
27
hGCGR IC50 = 9.2 nM
O
Me
O
NH
N
a, b
OH Me
O
O
H
N
Cl
O
30
N
H
O
29
O
O
O
NH
O
c, d
Cl
HO
N
O
N
CF3
27
Scheme 3. Reagents and conditions: (a) mCPBA, CH2Cl2, rt; (b) (R,R)-pseudoephedrineÁHCl, EDC, HOBt, DIEA, DMF; (c) 2-bromo-6-trifluoromethylpyridine, CuI, K3PO4,
trans,N,N-dimethyl-1,2-cyclohexanediamine, toluene, 110 °C; (d) TFA, CH2Cl2, rt.
Me
O
Me
O
N
a, b, c
OH Me
N
H
N
OH Me
O
30
31
O
O
Scheme 4. Reagents and conditions: (a) n-BuLi, LiCl, methyl 4-bromomethyl benzoate, THF; (b) LiOH, dioxane/water; (c) b-alanine-OtBu, EDC, HOBt, DIEA, DMF.
A number of compounds were selected for determination of
their PK profiles in the rat (Table 4). These analogs, though struc-
turally similar, showed diverse PK profiles. Compound 18 showed
moderate plasma clearance, a moderate half-life, and good bio-
availability (F = 42%). Compound 23, which differs from 18 by the
presence of an additional 4-chloro group on the N-aryl ring,
showed lower plasma clearance, but decreased oral bioavailability.
Pyridine analog 27 showed relatively high plasma clearance and
low bioavailability. While increased lipophilicity on the N-aryl ring
had a detrimental effect on rat PK (cf., compounds 18 and 23)
pound 33 was a weak binder of the hERG gene product (IKr
IC50 = 3700 nM),12 and showed minimal inhibition of the major
CYP isoforms (CYP3A4, 2C9, 2D6: IC50 >11000 nM).
The ketone moiety in compound 33 raised the theoretical po-
tential for racemization of the
a-chiral center and covalent trap-
ping of nucleophiles. To address the first concern, compound 33
was incubated at 40 °C for 24 h in 1:1 mixtures of dioxane and buf-
fered solutions from pH 2 to pH 10. In all cases, chiral HPLC anal-
ysis indicated no racemization. Furthermore, compound 33
demonstrated no racemization or irreversible covalent binding
when incubated with human liver microsomes, or when adminis-
tered orally in the rat.13
increasing the
a-carbonyl chain length as in compound 33 gave
rise to an improved pharmacokinetic profile relative to compound
18, with lower plasma clearance, increased oral exposure, and
moderate oral bioavailability. Compound 33 also displayed a favor-
able PK profile in the dog (t1/2 = 2.7 h, F = 60%).
In addition to providing favorable PK profiles in the rat and the
dog, compound 33 was found to have acceptable selectivity over
related GPCRs and other ancillary targets. When tested for func-
tional antagonism of the human GLP-1 and human GIP receptors,
which play roles in glucose dependent insulin secretion, compound
33 was >100-fold selective for the human glucagon receptor
(hGLP-1 cAMP IC50 >10000 nM; hGIP cAMP IC50 = 1400 nM). Com-
Indole 33, a highly potent hGCGR antagonist (cAMP IC50
=
12 nM) with desirable PK profiles and a good off-target profile,
was further evaluated for activity in vivo. When administered oral-
ly to mice expressing the human glucagon receptor, followed after
60 min by an IP injection of glucagon,8 compound 33 effected a sig-
nificant blockade of the glucagon-induced glucose excursion at all
doses (3, 10, and 30 mg/kg, Fig. 2). Compound 33 was then tested
for its ability to lower ambient glucose levels in ob/ob mice
expressing only the human glucagon receptor. In this hyperglyce-
mic rodent model (baseline glucose levels ꢀ275 mg/dl), oral

C. Sinz et al. / Bioorg. Med. Chem. Lett. 21 (2011) 7124–7130
7129
Table 2
Effect of
a
-keto substitutiona
O
NH
OH
R
O
Cl
N
O
CF3
Compound
R
hGCGR BNDb IC50
hGCGR cAMPb IC50
(nM) (n)
(nM) (n)
18
32
33
17
55
16 0.6 (2)
10 1.9 (4)
42
12 1.6 (4)
34
35
17 3.0 (2)
37
36
55
Figure 2. Effect of compound 33 on glucagon-induced glucose excursion in hGCGR
mice. Compound was administered orally in 0.25% methylcellulose.
a
All compounds are single enantiomers which were resolved via chiral HPLC (see
footnote 7 for details); data shown are for the enantiomer which is the more potent
hGCGR antagonist.
b
Each IC50 in this Letter is reported as an average rounded to two significant
figures standard error of the mean when more than one measurement was made.
Table 3
Optimization of indole 5-substituenta
O
NH
OH
O
R
N
O
CF3
Figure 3. Acute lowering of ambient glucose levels by compound 33 in an hGCGR/
ob/ob mouse model. Compound was administered orally in 0.5% methylcellulose.
Compound
R
hGCGR BNDb IC50 (nM) (n) hGCGR cAMPb IC50 (nM)
(n)
N-Aryl-2-acylindoles are a novel class of potent human gluca-
gon receptor antagonists. These chiral compounds displayed a con-
siderable preference for a single enantiomer. Optimization of the
33
36
37
38
39
40
Cl
F
10 1.9 (4)
10 5.8 (3)
12 1.7 (4)
20
35
17 0.8 (2)
44 19 (2)
24
OMe 26 3.7 (2)
OCF3 5.7 1.5 (2)
CF3
Me
indole C5-substituent, substitution on the N-aryl ring, and the
a-
16 6.9 (3)
12 4.7 (3)
carbonyl group led to the discovery of indole 33. This compound
displayed desirable PK profiles in the rat and the dog, as well as
good activity in both an acute hGCGR murine pharmacodynamic
assay and an acute ob/ob/hGCGR murine efficacy model. Based on
this overall profile, compound 33 was selected for further evalua-
tion in preclinical safety studies. Toward this end, compound 33
was found to be well tolerated at 100 mg/kg/day in an 8-day exper-
imental rodent tolerability study, lending further support for the
potential utility of N-aryl-2-acylindoles as a treatment for type 2
diabetes.
a
All compounds are single enantiomers which were resolved via chiral HPLC (see
footnote 7 for details); data shown are for the enantiomer which is the more potent
hGCGR antagonist.
Each IC50 in this Letter is reported as an average rounded to two significant
figures standard error of the mean when more than one measurement was made.
b
administration of 33 at 1 mg/kg led to a significant reduction in
ambient glucose levels over a 6 h period (Fig. 3).
Table 4
Pharmacokinetic profiles of human glucagon receptor antagonists in rata and dogb
Compound
Animal
Clp (ml/min/kg)
Vd (L/kg)
t1/2 (h)
PO AUCN
(lM.h/dose)
Cmax/dose (lM)
%F
18
23
27
33
33
Rat
Rat
Rat
Rat
Dog
14.2
9.2
24
6.8
3.8
1.5
1.7
1.3
1.1
1.9
2.7
0.85
0.52
0.15
1.46
4.2
0.25
0.15
0.07
0.33
0.84
42
17
9.1
33
60
0.84
0.90
0.98
0.32
a
Compounds were dosed at 1 mg/kg IV, 2 mg/kg PO in a 15:35:55 mixture of DMSO, water, and PEG400.
Compound was dosed at 0.5 mg/kg IV, 2 mg/kg PO in a 10:50:40 mixture of EtOH, PEG400, and water.
b

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C. Sinz et al. / Bioorg. Med. Chem. Lett. 21 (2011) 7124–7130
MEDI-294.; (b) Sinz, C. J., et al, Bioorg. Med. Chem. Lett. 2011, 21.; (c) Chang, J.,
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of the American Chemical Society, New Orleans, LA, April 6, 2008; American
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Christensen, I. T.; Jørgensen, A. S.; Ynddal, L.; Brand, C. L.; Bagger, M. A.; Lau, J. J.
Med. Chem. 2009, 52, 2989–3000.
11. Compound 28 prepared by the degradation route described in Scheme 3 was
spectroscopically identical (HPLC, NMR) to that prepared by the asymmetric
alkylation route shown in Scheme 4. Furthermore, the enantiomer of indole 29
was carried through the route shown in Scheme 3, leading to a compound
which is diastereomeric with respect to pseudoephedine amide 30, and which
is spectroscopically distinct by NMR.
12. IKr binding data were obtained by measuring displacement of
[
35S]-
radiolabeled MK-499 from HEK cells stably expressing hERG: Lynch, J. J., Jr.;
Wallace, A. A.; Stupienski, R. F., III; Baskin, E. P.; Beare, C. M.; Appleby, S. D.;
Salata, J. J.; Jurkiewicz, N. K.; Sanguinetti, M. C.; Stein, R. B.; Gehret, J. R.;
Kothestein, T.; Claremon, D. A.; Elliott, J. M.; Butcher, J. W.; Remy, D. C.;
Baldwin, J. J. J. Pharmacol. Exp. Ther. 1994, 269, 541–553.
13. For in vitro experiments, a radiolabelled (3H) analog of compound 33 (10
lM,
60 min, 37 °C) was incubated with human and rat liver microsomes and
irreversible protein binding was measured. For in vivo covalent binding
experiments, a radiolabelled analog of compound 33 was administered PO
(20 mg/kg) to Sprague-Dawley rats and irreversible binding was measured to
plasma and liver proteins 2, 6 and 24 h post dose; for analysis of racemization
in vivo, radiolabelled 33 was administered IV (2 mg/kg) to bile duct cannulated
rats, and the excreted parent compound was assayed via chiral HPLC.
4. (a) Kim, R. M.; Chang, J.; Lins, A. R.; Sinz, C. J.; Beconi, M.; Brady, E. J.; Candelore,
M.; Ding, V.; Jiang, G.; Lin, Z.; Qureshi, S.; Salituro, G.; Saperstein, R.; Shang, J.;
Tota, L.; Wright, M.; Yang, X.; Zhang, B. B.; Parmee, E. R. Abstracts of Papers,
232nd National Meeting of the American Chemical Society, San Francisco, CA,
September 12, 2006; American Chemical Society: Washington, DC, 2008;