Discovery of 6,7-dihydro-5H-pyrrolo[2,3-a]pyrimidines as orally available g protein-coupled receptor 119 agonists

Article abstract of DOI:10.1021/jm301404a

GPR119 is a 7-transmembrane receptor that is expressed in the enteroendocrine cells in the intestine and in the islets of Langerhans in the pancreas. Indolines and 6,7-dihydro-5H-pyrrolo[2,3-a]pyrimidines were discovered as G protein-coupled receptor 119

Full text of DOI:10.1021/jm301404a

Article
pubs.acs.org/jmc
Discovery of 6,7-Dihydro‑5H‑pyrrolo[2,3‑a]pyrimidines as Orally
Available G Protein-Coupled Receptor 119 Agonists
Subba R. Katamreddy,*,† Andrew J. Carpenter,* Carina E. Ammala, Eric E. Boros, Ron L. Brashear,
Celia P. Briscoe, Sarah R. Bullard, Richard D. Caldwell, Christopher R. Conlee, Dallas K. Croom,
Shane M. Hart, Dennis O. Heyer, Paul R. Johnson, Jennifer A. Kashatus, Doug J. Minick,
Gregory E. Peckham, Sean A. Ross, Shane G. Roller, Vicente A. Samano, Howard R. Sauls,
Sarva M. Tadepalli, James B. Thompson, Yun Xu, and James M. Way
GlaxoSmithKline Research & Development, Five Moore Drive, Research Triangle Park, North Carolina 27709, United States
ABSTRACT: GPR119 is a 7-transmembrane receptor that is expressed in the
enteroendocrine cells in the intestine and in the islets of Langerhans in the pancreas.
Indolines and 6,7-dihydro-5H-pyrrolo[2,3-a]pyrimidines were discovered as G protein-
coupled receptor 119 (GPR119) agonists, and lead optimization efforts led to the
identification of 1-methylethyl 4-({7-[2-fluoro-4-(methylsulfonyl)phenyl]-6,7-dihydro-
5H-pyrrolo[2,3-d]pyrimidin-4-yl}oxy)-1-piperidinecarboxylate (GSK1104252A) (3), a
potent and selective GPR119 agonist. Compound 3 showed excellent pharmacokinetic
properties and sufficient selectivity with in vivo studies supporting a role for GPR119 in glucose homeostasis in the rodent. Thus,
3 appeared to modulate the enteroinsular axis, improve glycemic control, and strengthen previous suggestions that GPR119
agonists may have utility in the treatment of type 2 diabetes.
(NAPEs) and N-oleoyldopamine12 (OLDA), and, most recently,
INTRODUCTION
■
5-hydroxyeicosapentaenoic acid (5-HEPE)13 have all been
Type 2 diabetes (T2D) is a chronic disease characterized by
insulin resistance, insufficient insulin secretion from pancreatic
β-cells, and hyperglycemia. Glycemic control is the key for
metabolic homeostasis, and the uncontrolled hyperglycemia
suggested as endogenous ligands. However, given that the
potency is relatively low or there are selectivity issues, it is
difficult to conclusively identify any one of these as the only or
true endogenous ligand.
leads to long-term complications including organ failure and
Evidence that activation of GPR119 may play a role in glucose
amputation.1 Recent estimates suggest that the incidence of
diabetes in the U.S. may double over the next 20 years.2
homeostasis mainly comes from selective, small-molecule
agonists. Specifically, GPR119 agonists have been shown to
Presently, less than one-third of T2D patients will achieve the
elicit nutrient-independent incretin and GI hormone release as
targeted plasma glucose levels with the currently available
well as nutrient-stimulated insulin secretion,8b,c,14 thereby
antidiabetic agents.3 Furthermore, the currently available
lowering plasma glucose levels during an oral glucose tolerance
treatments either as monotherapies or combination therapies
test.14a Interestingly, there is some evidence to support a role for
have their own limitations and potential risks.4 This has
prompted the scientific community to search for novel agents
GPR119 agonists in reducing food intake and promoting weight
loss, possibly through the release of GLP-1, which is known to
for the treatment and prevention of diabetes with novel
delay gastric emptying.10 However, Arena Pharmaceuticals have
reported that anorectic effects of their GPR119 agonists are still
observed in knockout mice.15 While the details of the mechanism
of action of this receptor are not yet fully understood, GPR119
agonists may have the potential to deliver superior glycemic
control versus current gold standard oral therapies given that no
current antidiabetic medication triggers the incretin and GI
hormone release as well as having direct actions on nutrient-
stimulated insulin secretion mechanisms for glucose control.16
The disclosure by Arena Pharmaceuticals that pyrazolopyr-
imidines,17 represented by example 1, were potent GPR119
agonists18 led us to surmise that alternative bicyclic structures
might be equally active (Figure 1). In this article, we disclose our
mechanisms of action.5
G Protein-coupled receptor 119 (GPR119) is a class A
(rhodopsin-like) 7-transmembrane, G protein-coupled receptor
(GPCR) with little homology to other receptors.6 The human
GPR119 receptor consists of 335 amino acids and is highly
conserved in mouse and rat.7 GPR119 is expressed in human
insulin-secreting pancreatic islets and incretin (GIP and GLP-1)
secreting K- and L-cells of the gastrointestinal (GI) tract.8 There
are some contradictory reports on its cellular subtype localization
in islets;8a,b however, there seems to be some consensus that the
receptor is expressed in the β-cells.8b,9
The physiological role of GPR119 remains unknown.
Elucidating the role of GPR119 signaling has been hampered
by the lack of endogenous ligands that signal solely through
GPR119. Lysophosphatidylcholine6a (LPC), oleoylethanola-
mide10 (OEA), N-acylated ethanolamine phospholipids11
Received: September 27, 2012
© XXXX American Chemical Society
A
dx.doi.org/10.1021/jm301404a | J. Med. Chem. XXXX, XXX, XXX−XXX

Journal of Medicinal Chemistry
Article
Figure 1. Representative GPR119 agonists.
a
Scheme 1. Preparation of Indolines 2, 9, and 10
a
Reagents and conditions: (a) Ph3P, DIAD, THF, RT, 77%; (b) CuI, K3PO4, trans-1,2-cyclohexanediamine, toluene, reflux, 8%; (c) CuI, L-proline,
NaOH, CH3SO2Na, DMSO, 110 °C, 98% for 6→7; (d) NaCNBH3, HOAc, RT, 71% for 7→9; (e) (i) TFA, CH2Cl2, RT, (ii) i-PrOCOCl, Et3N,
CH2Cl2, RT, 54%.
initial discoveries19 of indolines, such as 2, and 6,7-dihydro-5H-
pyrrolo[2,3-d]pyrimidines (DHPPs), such as 3
(GSK1104252A), as GPR119 agonists.14a The structure−activity
RESULTS
■
Indoline-based agonists were prepared as shown in Scheme 1. A
Mitsunobu reaction between substituted 4-hydroxyindole (4)
relationships (SAR) of the DHPP chemotype, in addition to in
vitro and in vivo activities of the lead molecule 3, will be
highlighted.
and N-Boc-4-hydroxypiperidine afforded the intermediate 5.
Compound 5, upon N-arylation with 1,4-diiodobenzene using
Buchwald conditions,20 yielded indole derivative 6. A copper-
B
dx.doi.org/10.1021/jm301404a | J. Med. Chem. XXXX, XXX, XXX−XXX

Journal of Medicinal Chemistry
Article
a
Scheme 2. Attempted Preparation of DHPP Chemotype
a
Reagents and conditions: (a) CuI, K3PO4, toluene, reflux, 30%; (b) NaH, THF, reflux, 70%; (c) CuI, L-proline, NaOH, CH3SO2Na, DMSO, 110
°C, 84%; (d) NaCNBH3, HOAc or NaCNBH3, BF3·OEt2, MeOH, reflux or NiCl2·6H2O, NaBH4, MeOH or 10% Pd/C, H2, MeOH.
a
Scheme 3. Preparation of the DHPP Chemotype, Exemplifying 3 and 23.
a
Reagents and conditions: (a (i) 0.5M NaOMe in MeOH, (ii) POCl3, 86% yield from 16; (b) NaH, THF, reflux; (c) (i) O3, −78 °C, CH2Cl2, (ii)
NaBH4, MeOH; (d) Ms2O, Et3N, CH2Cl2, 0 °C to RT, 46% overall yield from 17; (e) NaH, THF, reflux, 71%.
mediated sulfonylation21 was used to install the methyl sulfone
group to afford 7, and then the indole was reduced with
NaCNBH3 in acetic acid22 to afford indoline 9. Alternatively,
indole 6 could first be reduced to afford indoline 8, then
sulfonylated to give compound 9. Either scheme could be used to
provide good yields of the desired indoline. The tert-
butylcarbamate of compound 9 was removed using 20%
trifluoroacetic acid in dichloromethane and then converted to
isopropylcarbamate derivative 2 using isopropyl chloroformate
in good overall yield. Substituting 4-bromo-2-fluoro-1-iodoben-
zene in the N-arylation step and following the rest of the
sequence afforded compound 10.
7H-pyrrolo[2,3-d]pyrimidine (11) with 1-bromo-4-iodobenzene
afforded N-aryl derivative 12. Introduction of the piperidine
moiety was accomplished with treatment of the chloro derivative
12 with the sodium alkoxide in THF at reflux to afford
intermediate 13. Conversion of the bromide to the sulfone using
sodium methanesulfinate afforded the pyrrolo[2,3-d]pyrimidine
intermediate necessary to try the reduction step. Unfortunately,
attempts to reduce the pyrrolo[2,3-d]pyrimidine of 14 to the
desired dihydro derivative 15 using a variety of conditions
(NaCNBH3, HOAc; NaCNBH3, BF3·OEt2, MeOH; Pd/C,
EtOH; NiCl2·6H2O, NaBH4, MeOH) were unsuccessful.
Our first successful synthesis in the DHPP series is illustrated
in Scheme 3.23 2-Allyldiethylmalonate (16) and formamidine
hydrochloride, under basic conditions, gave the 4,6-dihydrox-
Preparation of DHPP-based agonists was attempted as shown
in Scheme 2. Transition metal-mediated coupling of 4-chloro-
C
dx.doi.org/10.1021/jm301404a | J. Med. Chem. XXXX, XXX, XXX−XXX

Journal of Medicinal Chemistry
Article
a
Scheme 4. Synthesis of Phenyl Ring Derivatives 25−27
a
Reagents and conditions: (a) CuCN, NMP, 150 °C, 85% for 25, or PdCl2(Ph3P)2, 2-(tributylstannanyl)-1,3-thiazole, THF, 78 °C, 61% for 26, or
PdCl2(Ph3P)2, 1-methyl-2-(tributylstannanyl)-1H-pyrrole, THF, 78 °C, 12% for 27.
a
Scheme 5. Synthesis of Piperidine Derivatives 29−44
a
Reagents and conditions: (a) NaH, THF, reflux, 71%; (b) (i) 20% TFA, CH2Cl2, RT, (ii) RCO2Cl, Et3N (29−36); or RC(O)SCl, Et3N (37,
38); or 1,1′-carbonyldiimidazole; then MeI in CH3CN, then i-PrSH, Et3N, CH2Cl2 (39); RCS2Ph, Et3N (40); or RSO2Cl, Et3N (41−43); or
ClCH2CONEt2, Et3N (44).
a
Scheme 6. Synthesis of Oxadiazole, Pyrimidine, and Guanidine Intermediates 47, 50, 51, and 54
a
Reagents and conditions: (a) (i) CNBr, aqueous NaHCO3, CH2Cl2; (b) (i) 1N ZnCl2, Et2O/EtOAc, (ii) 4N HCl, EtOH, ∼30% from 45; (c) i-
Pr2NEt, CH3CN, reflux, 83% for 50, 50% for 51, and 51% for 53; (d) H2, 10% Pd/C, HOAc/aqueous HCl, 84%.
ypyrimidine derivative that, upon POCl3 treatment, afforded
dichloro compound 17. Nucleophilic displacement reaction
between 17 and 2-fluoro-4-methylsulfonylaniline (18) using
NaH in THF delivered the pyrimidine derivative 19. The allyl
group of 19 was oxidatively cleaved using ozone, then reduced in
situ with NaBH4 to afford alcohol 20. Selective activation of the
alcohol moiety as a mesylate was achieved with Ms2O and Et3N
in CH2Cl2 to afford an intermediate, which spontaneously
cyclized to afford the desired key bicyclic core 21. Introduction of
the hydroxypiperidine portion was accomplished as before,
D
dx.doi.org/10.1021/jm301404a | J. Med. Chem. XXXX, XXX, XXX−XXX

Journal of Medicinal Chemistry
Article
a
Scheme 7. Synthesis of Oxadiazole, Pyrimidine, and Guanidine Derivatives 55−58
a
Reagents and conditions: (a) NaH, THF, reflux, 55% for 55, 39% for 56, 50% for 57, and 6% for 58.
Scheme 8. Preparation of C2-Methyl Analogues 60−62
under basic conditions (NaH, THF, reflux), to provide desired
compound 3.
coupling with the appropriate aniline or adding in the
appropriately functionalized piperidine, respectively. A prom-
inent feature found on the piperidine ring of many GPR119
agonists17 is the 1,2,4-oxadiazole. This could be prepared
following Scheme 6.17 4-Hydroxypiperidine (45) was treated
with cyanogen bromide in the presence of NaHCO3 to afford the
intermediate carbonitrile 46. Treatment of 46 with N-hydroxy-2-
methylpropanimidamide in the presence of ZnCl2 afforded the
coupled intermediate. Cyclization-dehydration occurred when
the intermediate was treated with acid (4N HCl) in EtOH at
reflux for 1 h to provide 47. 1-(5-Fluoropyrimidin-2-yl)piperidin-
4-ol (50) and 1-(5-ethylpyrimidin-2-yl)piperidin-4-ol (51) were
prepared from 45 and the corresponding chloropyrimidines 48
and 49, respectively, in acetonitrile with diisopropylethylamine.
1-(Pyrimidin-2-yl)piperidin-4-ol (52) was prepared in a similar
manner, and then the pyrimidine ring of 53 was partially reduced
with hydrogen and Degussa-type palladium on carbon in a
mixture of acetic acid and aqueous hydrochloric acid to afford 54.
Analogues 55−58 were prepared from the corresponding
hydroxypiperidine intermediates with 21 and NaH in refluxing
THF as shown in Scheme 7.
Substitution at the C2 position of the pyrimidine ring was
explored. Compounds (60−62) with a methyl substituent at the
C2 position of the DHPPs could be prepared in a similar manner
to that described for the unsubstituted analogues shown in
Scheme 3 but starting with ethanimidamide hydrochloride (59)
and 16 (Scheme 8).
Compounds bearing an amino substituent at the C2 position
of the pyrimidine ring of the DHPPs were prepared as described
in Scheme 9, which shows that the five-membered ring is built
Compound 23 was prepared by reacting 4-bromoaniline with
17 and then converting the bromide to the methylsulfonyl (CuI,
L-proline, NaOH, CH3SO2Na, DMSO, 110 °C) in the final step
in the synthesis. An improved synthesis of the DHPPs,
particularly for key intermediate 21, employing a reductive
amination reaction as the key step, has recently been reported.24
Exploration of the SAR of the phenyl region of the DHPPs was
most easily accomplished with intermediate 24, which provided
an opportunity to examine the left-hand side of these molecules
in a divergent manner (Scheme 4). Compound 24 was prepared
from 17 and commercially available 4-bromo-2-fluoroaniline as
described for the synthesis of 3 in Scheme 3. The bromide of 24
served as a handle to access a nitrile 25 via Rosenmund−von
Braun conditions,25 or heteroaromatic groups (compounds 26
and 27) via Suzuki conditions.26
On the other end of the molecules, the N-Boc protected
piperidine 28 served as a convenient spot for analogue synthesis
through deprotection, to afford the unprotected piperidine,
which could be functionalized. Carbamates 29−36 were
prepared from the corresponding chloroformates. The thio-
carbamates 37−39 and dithiocarbamate 40 were prepared from
S-alkyl chloridothiocarbonates27a or carbamoylimidazolium
salts27b and alkyl chlorodithiocarbonates,27a respectively
(Scheme 5). Sulfonamides 41−43 were prepared from sulfonyl
chlorides. 2-Aminoacetamide 44 was prepared from the
corresponding 2-chloroacetamide.
While additional substituents on the left- and right-hand side
were evaluated, they were prepared by an alternative route:
E
dx.doi.org/10.1021/jm301404a | J. Med. Chem. XXXX, XXX, XXX−XXX

Journal of Medicinal Chemistry
Article
a
Scheme 9. Preparation of C2-Amino Analogues 73 and 74
a
Reagents and conditions: (a) CH3CN, 60 °C; (b) catalytic amount of concentrated H2SO4, MeOH, 43% for 2 steps; (c) P2S5, THF, 70 °C, 50%;
(d) guanidine hydrochloride or N-methyl guanidine hydrochloride, 1M NaOMe, MeOH, 60 → 90 °C; (e) POCl3, Et3N, 70 °C; (f) NaH, THF, 60
°C, 2% from 61 and 15% from 62.
a
Scheme 10. Preparation of Analogues 77 and 78 Bearing a Nitrogen-Atom Linker
a
Reagents and conditions: (a) K2CO3, DMF, μw, 200 → 220 °C, 12% for 77 and 9% for 78.
a
Scheme 11. Preparation of Analogues 80 and 81 Bearing a Sulfur-Atom Linker
a
Reagents and conditions: (a) K2CO3, acetone, 60 °C, 71%; (b) m-CPBA, CH2Cl2, 0 °C, 18%.
first, then construction of the pyrimidine follows. Reaction28 of
6,6-dimethyl-5,7-dioxaspiro[2.5]octane-4,8-dione (63) with ani-
line 19 afforded the 2-oxo-1-aryl-3-pyrrolidinecarboxylic acid 64,
which could be esterified using Fischer protocol (acid, methanol)
to provide 65. Conversion of the carbonyl moiety to a
thiocarbonyl using P2S5 or Lawesson’s reagent afforded
intermediate 66, which can be reacted with guanidine derivatives
(R = H, guanidine hydrochloride, (67) or R = Me, N-
F
dx.doi.org/10.1021/jm301404a | J. Med. Chem. XXXX, XXX, XXX−XXX

Journal of Medicinal Chemistry
Article
a
Scheme 12. Preparation of C6-Methyl Analogues 86 and 87
a
Reagents and conditions: (a) (i) Boc2O, cat. DMAP, CH2Cl2, RT, (ii) K2OsO4·2H2O, 3:2 acetone/H2O, NaIO4, RT; (b) 1.4M MeMgBr, THF,
39% from 19; (c) 20% TFA, CH2Cl2, RT; (d) Ms2O, Et3N, CH2Cl2, RT, 47% from 83; (e) (i) NaH, THF, reflux; (ii) chiral separation, 82%.
a
Scheme 13. Preparation of Pyridyl Analogues 97 and 98
a
Reagents and conditions: (a) NaCNBH3, HOAc, MeOH, −15 °C → RT; (b) KO-t-Bu, MeOH, 11% for 93; (c) NaH, THF, reflux, 88% for 95; (d)
Oxone, MeOH/acetone/H2O, RT, 81% for 97 and 7% overall yield for 98 from 17.
methylguanidine hydrochloride (68)) in the presence of sodium
methoxide to afford bicyclic intermediates 69 (R = H) and 70 (R
= Me). Chlorination with POCl3 afforded intermediates 71 and
72, which was followed by introduction of the hydroxypiperidine
portion via the alkoxide to give the desired targets 73 and 74.
Changes to the linker between the bicyclic core and the
piperidine were investigated. The nitrogen-linked analogue 77
was prepared by treating the chloro intermediate 21 with 1-
methylethyl 4-({[(1,1-dimethylethyl)oxy]carbonyl}amino)-1-
piperidinecarboxylate (75) and heating in a microwave in the
presence of potassium carbonate, as shown in Scheme 10. The N-
methyl derivative 78 was prepared using 1-methylethyl 4-
[{[(1,1-dimethylethyl)oxy]carbonyl}(methyl)amino]-1-piperi-
dinecarboxylate (76) under conditions described above.
temperature (60 °C) afforded desired compound 80 in good
yield (71%). Oxidation of the sulfur to the sulfone oxidation state
was accomplished with m-CPBA at 0 °C, which afforded
compound 81.
Introduction of a methyl group at the 2-position of the five-
membered ring was accomplished by intercepting the N-Boc
protected aldehyde 82, prepared from allyl 19, with methyl
Grignard in THF at room temperature in 39% yield to afford
racemic alcohol 83 (Scheme 12). After N-Boc deprotection to
afford intermediate 84, activation of the secondary alcohol as its
mesylate was achieved with methanesulfonyl anhydride in
CH2Cl2 with triethylamine. The presence of triethylamine likely
facilitated cyclization as the isolated product was the
pentultimate intermediate 85. Introduction of the piperidine
portion occurred as has been reported for other substrates. The
resulting racemate was separated by chiral chromatography to
afford individual enantiomers (+)-86 and (−)-87. The assign-
ment of R- or S-stereochemistry for this enantiomeric pair was
not made.
Thio- or sulfonyl-linked analogues were prepared by the
following sequence (Scheme 11), which employed 1-methylethyl
4-mercapto-1-piperidinecarboxylate (79)19 as the nucleophile.
Treatment of chloro intermediate 21 with mercaptan 79 in
acetone in the presence of potassium carbonate at elevated
G
dx.doi.org/10.1021/jm301404a | J. Med. Chem. XXXX, XXX, XXX−XXX

Journal of Medicinal Chemistry
Article
Figure 2. Pyridyl (99−101) and substituted phenyl (102−108) analogues.
a
Scheme 14. Preparation of Sulfone Analogue 113
a
Reagents and conditions: (a) NaSMe, DMF, 85 °C, 88%; (b) SnCl2, conc HCl, EtOH, 80 °C, 81%; (c) (i) NaCNBH3, HOAc, MeOH, −15 °C →
RT, (ii) 4% TFA, 1,2-DCE, MeOH, RT, 31%; (d) (i) NaH, THF, reflux, 72%, (ii) Oxone, MeOH/acetone/H2O, RT, 29%.
Replacement of the N-phenyl group with pyridyl rings was
accomplished using the sequence shown in Scheme 13. Using the
reductive amination strategy with the appropriate aminopyridine
derivative (90 and 91) and aldehyde 88, prepared from allyl 17,
the uncyclized intermediates 89 and 90 were prepared,
respectively. Unlike the phenyl congeners, which cyclized
under acidic conditions (TFA), the pyridyl analogues cyclized
more smoothly in the presence of a base such as potassium t-
butoxide or a trialkylamine (Hunig’s base). The resultant chloro
intermediates 93 and 94 were then reacted with the alkoxide of
the 4-hydroxypiperidine derivative to afford sulfides 95 and 96.
Oxidation of the sulfides to sulfones 97 and 98 was accomplished
with Oxone or m-CPBA.
Employing a similar strategy as shown in Scheme 13, other
pyridyl analogues (compounds 99, 100) and substituted phenyl
derivatives (102−105) were prepared (Figure 2). For analogues
99 and 100, the initial reductive amination step with 88 and
commercially available 6-aminonicotinonitrile or 5-fluoropyr-
idin-2-amine, respectively, employed Na(OAc)3BH and TFA in
CH2Cl2. In addition, the acidic conditions in the reduction
amination step facilitated the ring closure of the five-membered
ring, so a separate cyclization step was not needed for each
analogue. The synthesis of 99 and 100 were completed by
introducing the functionalized hydroxypiperidine 22 as
previously shown for other analogues. The synthesis of 101
began with a reduction amination reaction between 88 and
commercially available 2-methyl-3-pyridinamine. The subse-
quent key five-membered ring closing step occurred using a Pd-
mediated (PdCl2(Ph3P)2, KO-t-Bu, toluene, reflux) cyclization
reaction. Synthesis of 101 was completed by introducing the
functionalized hydroxypiperidine 22 as previously shown for
other analogues, in 55% overall yield. Intermediate bromide 102
was prepared from 88 and 4-bromo-3-fluoroaniline in a manner
similar to that described for 99. Bromide 102 was converted to
nitrile 103 using CuCN in NMP at 150 °C (7% overall yield from
89). Sulfone 104 was prepared from 102 using CH3SO2Na, CuI,
L-proline, and NaOH in DMSO at 110 °C (38% overall yield
from 88). Sulfonamide 105 was prepared from 88 and 4-amino-
N,N-dimethylbenzenesulfonamide in a manner similar to that
described for 99 in 17% overall yield. Sulfide 106 was prepared
from 88 and 2-fluoro-4-(methylthio)aniline in 34% yield using
the same strategy used in the synthesis of 99. 2-Fluoro-4-
(methylthio)aniline was prepared from the commercially
available 2-fluoro-4-iodoaniline and NiBr2 (Zn dust, 2,2′-
dipyridyl, DMF, 75 °C) in 73% yield. Racemic sulfoxide was
prepared from sulfide 106 via oxidation with hydrogen peroxide
in hexafluoroisopropanol,29 to prevent overoxidation. The
resultant racemate was separated by chiral chromatography to
H
dx.doi.org/10.1021/jm301404a | J. Med. Chem. XXXX, XXX, XXX−XXX

Journal of Medicinal Chemistry
Article
a
afford the individual sulfoxide enantiomers 107 (R enantiomer)
and 108 (S enantiomer). The absolute stereochemistry was
determined based on VCD spectroscopy.30
Table 1. Potency of Selected Analogues
compd
av pEC50
StDev
av % max
StDev
n
2
6.3
7.3
6.2
6.8
6.0
6.9
7.2
7.1
6.8
7.7
6.5
6.6
7.1
7.3
7.6
7.4
6.3
6.5
6.6
7.5
8.0
7.9
5.8
5.9
6.4
<5
0.25
0.15
0.31
0.13
0.19
0.38
0.11
0.16
0.09
0.10
0.26
0.14
0.09
0.29
0.09
0.15
0.21
0.07
0.15
0.07
0.09
0.24
0.10
0.08
0.17
70
91
6.3
15.9
5.6
3
6
3
6
6
12
6
6
5
6
6
6
6
8
6
6
6
5
6
6
6
4
4
5
4
3
6
6
6
2
4
6
4
4
6
6
6
5
6
6
6
6
6
6
6
6
6
6
4
9
10
14
9
The synthesis of sulfone 113 began by converting 4-fluoro-2-
methyl-1-nitrobenzene (109) to the methylthio derivative 110
with sodium thiomethoxide. Reduction of the nitro group led to
intermediate 111 that underwent the reduction amination/
cyclization sequence with 88 to afford chloride 112. The
hydroxypiperidine portion was appended using NaH in THF,
and the sulfide was oxidized with Oxone to complete the
synthesis of 113.
3
7
28
9
92
12.2
15.5
26.4
6.4
10
23
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
40
41
42
43
44
46
47
55
58
60
61
62
73
74
77
78
80
81
86
87
97
98
99
100
101
103
104
105
106
107
108
113
62
154
134
74
10.8
5.2
44
139
119
135
111
131
94
18.3
34.1
12.1
13.5
22.4
5.3
DISCUSSION
■
In relation to their bicyclic counterparts (represented by 1), the
indolines and DHPPs were initially designed to provide novel
compounds with potentially better: (1) potency, (2) systemic
exposure after oral administration, due to better solubility from
the reduction of aromaticity, and (3) selectivity.
80
5.3
The potency of the indolines and DHPPs were measured using
a reporter assay with the human GPR119 receptor stably
expressed in CHO-K1 6CRE-luciferase cells.31 All compounds
are referenced to a small molecule GPR119 agonist32 for
evaluation of intrinsic efficacy (% max response) because this
reporter assay is not sensitive enough to demonstrate robust
activity with the putative endogenous ligands (OEA, LPC, etc.).
The results are summarized in Table 1.
It was gratifying to observe that indoline 9 not only showed
activity against human GPR119 with a pEC50 = 6.8 (92%), it was
markedly better than its indole counterpart 7 (pEC50 = 6.2
(28%)). In addition, the intrinsic activity of the indoline was
much improved over the indole, 92% versus 28%. This served as
another impetus to continue to examine compounds with the
DHPP scaffold. As anticipated, good activity was also observed
with compounds 3 and 23 (pEC50 = 7.3 (91%) and pEC50 = 6.7
(154%), respectively), bearing the pyrimidine ring nitrogens of
the DHPP scaffold.
Substituents on the aryl or heteroaryl ring attached to the
dihydropyrrole nitrogen were investigated. Substitution at the 4-
position of the N-aryl ring was found to be critical for activity. For
example, compounds that lacked a functional group that could
act as a hydrogen bond acceptor were usually about a log unit less
potent than methylsulfone or nitrile. Thiazole 26 and pyrrole 27,
with a heterocycle at the 4-position, were found to retain much of
the potency, but not as much as the sulfone. Sulfide 106 also
showed intermediate potency with a pEC50 = 6.4, typical of a
compound that lacked a hydrogen-bond acceptor. The oxidation
state of the sulfur turned out to be less important than the
stereochemistry. For example, (R)-sulfoxide 107 had similar
potency to the sulfide and was a log less potent than the (S)-
isomer 108, which was equipotent with sulfone 3.
Substitution at the 2- and 3-positions of the phenyl ring was
also examined. A substituent, such as fluoro or methyl, was found
to increase potency over their unsubstituted versions, such as 23
(pEC50 = 6.9). A very slight preference for a fluoro atom at the 2-
position of 3 over the other possible iterations (3-F, 104, and 2-
Me, 113) was observed.
86
10.2
13.5
25.9
13.9
9.7
86
135
128
138
103
51
19.1
10.8
7.4
62
68
4.1
6.5
7.3
7.1
5.7
6.9
7.0
6.6
6.1
<5
0.22
0.14
0.15
0.02
0.33
0.12
0.18
0.07
129
105
124
66
16.8
6.0
19.3
19.0
21.1
3.1
69
45
44
7.0
33
3.4
5.7
6.2
7.1
<5
0.22
0.07
0.19
64
88
69
8.0
8.8
12.2
<5
<5
6.9
6.6
6.1
<5
0.10
0.13
0.14
111
97
6.4
4.8
105
14.4
<5
6.4
6.8
6.3
6.4
6.2
7.3
7.0
0.38
0.11
0.07
0.24
0.31
0.22
0.24
121
109
128
121
155
122
80
15.3
14.7
22.6
15.3
45.1
16.2
26.8
a
av StDev = average standard deviation; av %max = average percent
Similar findings with respect to optimal 4-substitution were
observed in the analogues bearing a pyridyl ring in place of the
phenyl ring. The methylsulfone in 97 (and 98) was preferred
over the nitrile from 99, which was better than a fluoro atom
found in 100. The placement of the pyridyl nitrogen at the 2- or
3-position did not affect activity in 97 and 98, respectively, as
each had a pEC50 ≥ 6.6.
maximum response; n = number of runs.
Structure−activity relationships of the bicyclic scaffold
revealed that further substitution on the rings at the 2- and 6-
positions did not produce compounds with greater potency.
Quite the contrary, these changes produced compounds with
I
dx.doi.org/10.1021/jm301404a | J. Med. Chem. XXXX, XXX, XXX−XXX

Journal of Medicinal Chemistry
Article
Table 2. Pharmacokinetic Properties of 3 in Rat and Mouse
species
t1/2 (h)
Cmax (μM)
Cl (mL/min/kg)
DNAUC0−24 h (μg·h/mL/dose)
Vss (L/kg)
%F
brain:plasma
rat
8.7
2.8
9
8.4
7.2
2035
1701
1.5
0.7
66
40
0.7
nd
mouse
20
diminished activity. Methyl-substituted analogues 60−62 were
about a log unit less potent, as was amino 73. The
monomethylated analogue, 74, did not agonize the receptor at
measurable levels. The 6-substituted analogues, 86 and 87, also
showed diminished activity. In addition, compared to the
oxygen-linked analogues, the nitrogen-linked analogues 77 and
78 were less active. The sulfur-linked analogue 80 was
comparable to oxygen analogue 3; however, the potency of
sulfone derivative 81 was not.
A general trend for the requirement of a lipophilic moiety was
observed during exploration of the right-hand side. This is most
easily seen in the carbamate series. As the size of the alkyl group
grew, so did potency. Methyl carbamate 29 was the weakest of
the group, while the larger tert-butyl carbamate 28, iso-butyl
carbamate 33, and neopentyl carbamate 34 were more potent.
When lipophilic portions were substituted with more hydrophilic
groups, activity was lost. Replacing a methylene in the n-butyl
derivative of 32 (pEC50 = 7.3) with oxygen (3-methoxypropyl
carbamate 36, pEC50 = 6.5) resulted in loss of potency. Similarly,
the fluoroethyl derivative 35 was less potent than the
corresponding n-propyl 31.
S-Thiocarbamates were also found to be potent. However,
these analogues did not seem to offer any advantage over their
oxygen congeners. For example, methyl S-thiocarbamate 37 had
a pEC50 = 6, which was about equipotent to methylcarbamate 29
(pEC50 = 6.5). Briefly, dithiocarbamates were also investigated.
Phenyldithiocarbamate 40 was potent, with a pEC50 = 7.9.
Unfortunately, these sulfur-containing compounds resulted in
low plasma exposure (dose normalized area under the curve,
DNAUC0−24h <0.03 μg·h/mL/dose), possibly due to extensive
metabolism. In addition, there was the potential for reactive
metabolites; thus, this class was deprioritized.
Sulfonamides were also investigated. Alkyl sulfonamides 41
and 42 were active, but a direct comparison between the
corresponding carbamate 30 to 41 demonstrates that carbamates
are superior (30,pEC50 = 7.1 versus 41, pEC50 = 5.8). The N,N-
dimethylsulfonamide derivative 43 also showed activity but was
still less potent than similar sized carbamates.
Having established which arrangement of functional groups on
the DHPP scaffold conferred reasonable potency, we turned our
attention to identifying compounds possessing the proper
combination of potency and plasma exposure. We had ruled
out the S-thiocarbamates and dithiocarbamates due to putative
extensive metabolism. As the size of the alkyl groups on the
carbamates grew, potency went up; however, we were mindful to
keep MW down in order to identify compounds with desirable
developability parameters. Among the carbamates, the best
balance of potency, MW, and exposure was the isopropylcarba-
mate. From a potency perspective, the isopropyloxadiazole and
4-ethylpyrimidine analogues were comparable and attractive.
Unfortunately, the AUC0−24h for isopropyloxadiazole 55 in the
rat35 after a 10 mg/kg dose was 0.5 μg·h/mL. 4-Ethylpyrimidine
57 was equally poor in the mouse with a DNAUC0−24h = 0.04
μg·h/mL/dose. Nitrile 25, however, did show good levels in rat
plasma (DNAUC >1 μg·h/mL/dose). Ultimately, methylsulfone
3 emerged as possessing the best balance of potency and
solubility. In fasted simulated intestinal fluid, the equilibrium
solubility of 3 was 4 μg/mL. Importantly, sulfone 3 was found to
have a DNAUC0−24h value of 2 μg·h/mL/dose. In addition, in the
rat, it possessed a moderate half-life (t1/2 = 8.7 h) and good
bioavailability (%F = 66). Additional pharmacokinetic properties
of 3, dosed 30 mg/kg in the rat and mouse, are shown in Table 2.
The selectivity profile of 3 revealed few issues. In a panel
against 50 aminergic receptors, 3 showed <20% binding at all
receptors at a concentration of 1 μM. In a separate panel of 68
receptors, ion channels, and enzymes, the only activity that
registered >50% inhibition or stimulation at 10 μM was
monoamine oxidase-B (MAO-B) inhibition (54%). Sulfone 3
did inhibit the cytochrome P450 subtype 2C9 with an IC50 = 3
μM. Overall, though, we concluded that the selectivity profile was
sufficiently clean to carry forward into target validation studies.
GPR119 agonist 3 was evaluated for its ability to increase
incretin and insulin secretion in vitro and ex vivo. Compound 3
caused GLP-1 secretion in GluTag36 cells with a pEC50 = 7.7,
which closely approximates its receptor potency (Figure 3). In
primary colonic crypt cells harvested from mice, 3 stimulated
GLP-1 release in a statistically significant manner over the vehicle
treated group (Figure 4) (p < 0.001). The lack of a dose response
Heterocyclic isosteres of the carbamates were examined. The
two best replacements that were identified were the oxadiazoles
and pyrimidines. For example, oxadiazole 55 and 4-ethyl-
pyrimidine 57 were as potent as carbamate 3.
While other classes of functional groups (amides, ureas,
substituted benzyls, alkyls; data not shown) were explored on the
right-hand side, most of them did not show desirable potency
(pEC50 > 6). One example is acetamide 44, in which a methylene
has been inserted between the carbonyl and the piperidine
nitrogen. Although it does preserve the lipophilic substitution
with the N,N-diethyl moiety, this analogue did not agonize the
receptor at concentrations lower than 10 μM. The SAR suggests
that the more basic piperidine nitrogen is not well tolerated.
In attempts to prepare analogues with increased solubility
while preserving the lipophilic alkyl groups, and while not
increasing molecular weight34 significantly, guanidine-containing
58 was prepared. Unfortunately, this analogue was not very
potent (pEC50 = 5.7).
Figure 3. GLP-1 release in GluTag cells with 3.
J
dx.doi.org/10.1021/jm301404a | J. Med. Chem. XXXX, XXX, XXX−XXX

Journal of Medicinal Chemistry
Article
(0−5 min) and second phase (10−90 min) C-peptide profile
(Figure 7), a direct reflection of insulin secretion.37
Figure 4. GLP-1 secretion in primary colonic crypt (PCC) cells with 3.
Figure 6. Glucose Infusion Rate with 3.
in this study may be due to the inherent variability in an
experiment working with primary cells or possibly that
compound 3 had reached the upper plateau on the dose
response curve at those doses. In isolated rat islets, 3, at 0.3 μM,
augmented insulin release in the presence of 12 mM glucose (p <
0.01). No insulin secretion was observed at 3 mM glucose for
either the treated or untreated cells.
To determine if the effects observed in vitro were GPR119-
mediated, 3 (30 mg/kg) was administered to wild-type and
knockout mice that had been fasted overnight (Figure 5). A near
4-fold increase in GLP-1 and 2-fold increase in GIP, when
compared to vehicle (n = 10, p < 0.01 and p < 0.01, respectively),
was observed in the WT mice, whereas no increase was observed
in GPR119 KO mice. This suggests that the effects on incretin
hormone secretion are mediated through GPR119.
Interestingly, there was also a 3-fold increase in PYY levels and
a 2-fold increase in glucagon in WT mice, compared to vehicle (n
= 8−10, p < 0.01 and p < 0.01, respectively; data not shown). Like
the incretins, no effect of 3 on PYY or glucagon secretion was
observed in KO mice.
To assess the effects of 3 on islet glucose response in vivo, C-
peptide levels were assessed in rats using the hyperglycemic
clamp. The plasma glucose levels were elevated and maintained
at similar (clamped) levels in both treatment groups throughout
the glucose infusion. The glucose infusion rate required to
maintain the clamp was significantly increased in the rats treated
with 3 (Figure 6), correlating with an increase in both first phase
Figure 7. Hyperglycemic clamp with 3.
With 3 having shown the ability to promote incretin secretion
in vitro and in vivo, in addition to demonstrating its effects on
insulin secretion in the whole animal, we sought to establish
whether or not 3 would impact glucose levels as a reflection of
these responses.19b In normal rats, 3, administered orally at 10
mg/kg, decreased the glucose excursion curve by 43% in a
glucose tolerance test (data not shown) when glucose was
administered intravenously and 38% when glucose was
administered orally (Figure 8) (n = 7, p < 0.01 for each). This
Figure 5. Incretin secretion in WT and KO mice.
K
dx.doi.org/10.1021/jm301404a | J. Med. Chem. XXXX, XXX, XXX−XXX

Journal of Medicinal Chemistry
Article
effect was similar to LAF23738 (vildagliptin), a DPP-IV inhibitor
used as a positive control.
While the details of the mechanism of action of this receptor
are not yet fully understood, small molecule 3 was used to
support a role for GPR119 in the control of glucose levels in the
rodent. These findings support preclinical reports of others that
have identified GPR119 agonists and suggest that GPR119 may
be an attractive target for the treatment of type 2 diabetes.
Preliminary communications from clinical studies with a few
GPR119 agonists, however, show mixed results. As observed in
preclinical studies in animals, MBX-2982 showed dose-depend-
ent reductions in glucose and increases in GLP-1 following a
mixed meal.40 Recently, Katz et al. demonstrated that a single-
dose of a GPR119 agonist, JNJ-38431055, in healthy subjects
increased postprandial GLP-1, GIP, and PYY but did not
decreased glucose excursion or increase insulin.41 Similarly,
GSK1292263,42 a selective GPR119 agonist that comes from a
structurally different class than the DHPPs, did not reduce
glucose AUC (0−24 h) when administered alone or when
codosed with metformin or sitagliptin.43 Whether GPR119
agonists are able to deliver sustained glucose lowering in type 2
diabetics is at present unclear and awaits longer term studies.
Figure 8. Rat OGTT with 3 and a DPP-IV inhibitor.
EXPERIMENTAL SECTION
■
While there is a statistically significant increase of GLP-1 upon
administration of 3, we wondered whether the magnitude was
sufficient to inhibit gastric emptying, as elevated GLP-1 levels are
known to cause a delay. Compound 3, dosed orally (10 mg/kg)
to normal male, SD rats that had been fasted overnight resulted in
a 27% reduction of the acetaminophen AUC (0−2 h) when
compared to vehicle. It has been shown that acetaminophen is
relatively unabsorbed in the stomach while rapidly absorbed and
detected in the circulation upon gastric release to the small
intestine, making it a suitable marker for determining the rate of
gastric emptying.39
In a separate species (mouse), we evaluated whether or not
compound 3 would affect food intake. Individually housed male,
C57BL/6 mice were fasted overnight, then treated with 10 mg/
kg of 3, and food intake was measure at 1, 2, 4, 6, and 24 h post
dose (data not shown). There was no effect relative to vehicle-
treated animals.
Unless otherwise noted, all nonaqueous reactions were carried out
under a nitrogen atmosphere using commercial grade solvents and
reagents. 1H NMR spectra were taken on a Varian (Agilent) Inova 400
NMR spectrometer. Chemical shifts are reported in parts per million
(ppm, δ) using the residual solvent line as a reference. Splitting patterns
are designated using the following abbreviations: s, singlet; d, doublet; t,
triplet; dd, doublet of doublet; m, multiplet; br, broad. Coupling
constants (J) are reported in hertz (Hz) where relevant. Mass
spectrometric analyses were conducted on a Waters Acquity UPLC
system and Waters Acquity SQD with alternating positive/negative
electrospray ionization scanning from 125 to 1000 amu, with a scan time
of 105 ms and an interscan delay of 20 ms. High resolution mass
spectrometric analysis was performed on a Waters qTOF Premiere mass
spectrometer operating in W mode. Reverse phase HPLC was
performed on a Gilson preparative system. The purity of the compounds
was established through analytical HPLC analysis on an Agilent
HP1100, and all compounds reported in the manuscript have a chemical
purity ≥95%.
1-Methylethyl 4-({1-[4-(Methylsulfonyl)phenyl]-2,3-dihydro-
1H-indol-4-yl}oxy)-1-piperidinecarboxylate (2). A solution of 4-
hydroxyindole (6.65 g, 50 mmol) in THF (600 mL) at 5 °C under N2
was treated with 1,1-dimethylethyl 4-hydroxy-1-piperidinecarboxylate
(20.1 g, 100 mmol) and triphenylphosphine (26.2 g, 100 mmol).
Diisopropyl azodicarboxylate (19.4 mL, 100 mmol) was added dropwise
over a 20 min period, and then the mixture was allowed to stir and warm
to RT for 8 h. The reaction was concentrated to give an orange oil that
was purified by flash chromatography (10→50% EtOAc/hexanes) to
afford 1,1-dimethylethyl 4-(1H-indol-4-yloxy)-1-piperidinecarboxylate
(5) (12.3 g, 77%) as a solid. 1H NMR (400 MHz, DMSO-d6): δ 11.03
(br s, 1H), 7.19 (t, J = 2.8 Hz, 1H), 6.99−6.93 (m, 2H), 6.54 (dd, J = 7.2,
1.2 Hz, 1H), 6.41 (app t, J = 2.4 Hz, 1H), 4.63 (m, 1H), 3.67−3.61 (m,
2H), 3.26−3.21 (m, 2H), 1.94−1.87 (m, 2H), 1.64−1.52 (m, 2H), 1.40
(s, 9H). LCMS (ESI): m/z 339 [M + Na]+. A solution of 5 (4.0 g, 12.6
mmol) in 1,4-dioxane (80 mL) at RT under N2 was treated with CuI
(240 mg, 1.3 mmol), K3PO4 (4.0 g, 19 mmol), and trans-1,2-
diaminocyclohexane (0.3 mL, 2.5 mmol), followed by 1,4-diiodoben-
zene (6.3 g, 19 mmol), and the mixture was heated to reflux for 15 h. The
reaction mixture was allowed to cool to RT, diluted with EtOAc (250
mL), and then filtered through a plug of silica (EtOAc wash). The filtrate
was concentrated, and the resultant residue was purified by flash
chromatography (0→20% EtOAc/hexanes) to afford 524 mg of 1,1-
dimethylethyl 4-{[1-(4-iodophenyl)-1H-indol-4-yl]oxy}-1-piperidine-
CONCLUSION
■
A novel series of indolines were found to be potent and selective
GPR119 agonists. Optimization of this series led to the 6,7-
dihydro-5H-pyrrolopyrimidines, best exemplified by compound
3. SAR analysis of this novel series suggests that a hydrogen-bond
acceptor group appended to the N-aryl ring significantly boosts
potency, while lipophilic groups as extensions from the
piperidine ring are preferred. The central region can accom-
modate a variety of scaffolds, and it has been shown that the
bicyclic, 6,7-dihydro-5H-pyrrolopyrimidine ring system produ-
ces compounds that are potent and selective while possessing
favorable pharmacokinetic properties.
Accordingly, compound 3 was used in functional in vitro and
in vivo studies. The data suggest that compound 3-mediated
GPR119 activation promotes in vitro (GluTag cells) GLP-1
secretion. It increases GLP-1 and GIP levels in vivo, and this
effect is nutrient independent and GPR119 dependent. In vitro,
compound 3 augments glucose-stimulated insulin secretion and,
in vivo, enhances both first and second phase insulin secretion.
Compound 3 also reduces glucose excursions and improves
glucose tolerance. When administered orally, 3 has been shown
to reduce gastric emptying, with no commensurate effect on food
intake.
1
carboxylate (6). H NMR (400 MHz, DMSO-d6): δ 7.88 (t, J = 8.8
Hz, 2H), 7.51 (d, J = 3.2 Hz, 1H), 7.39 (d, J = 8.8 Hz, 2H), 7.12 (t, J = 5.6
Hz, 1H), 7.09 (t, J = 8.0 Hz, 1H), 6.72 (d, J = 7.2 Hz, 1H), 6.70 (d, J = 3.2
L
dx.doi.org/10.1021/jm301404a | J. Med. Chem. XXXX, XXX, XXX−XXX

Journal of Medicinal Chemistry
Article
Hz, 1H), 4.70 (m, 1H), 3.66−3.60 (m, 2H), 3.28−3.24 (m, 2H), 1.94−
1.89 (m, 2H), 1.66−1.58 (m, 2H), 1.39 (s, 9H). LCMS (ESI): m/z 518
[M + H]+. A solution of 6 (355 mg, 0.69 mmol) in DMSO (4 mL) in a
10 mL glass tube was treated with CuI (13 mg, 0.07 mmol), MeSO2Na
(103 mg, 1.0 mmol), L-proline (16 mg, 0.14 mmol), and NaOH (6 mg,
0.14 mmol). The tube was sealed and then heated at 110 °C for 18 h.
The reaction mixture was allowed to cool and then poured onto EtOAc.
The mixture was washed with water and brine and was dried over
Na2SO4 and concentrated. The resultant residue was purified by flash
SiO2 column chromatography to afford 314 mg (98%) of 7 as an off-
white solid. 1H NMR (400 MHz, DMSO-d6): δ 8.08 (d, J = 8.5 Hz, 2H),
7.87 (d, J = 8.5 Hz, 2H), 7.65 (d, J = 3.4 Hz, 1H), 7.27 (d, J = 8.4 Hz,
1H), 7.15 (t, J = 8.0 Hz, 1H), 6.78 (m, 1H), 6.77 (s, 1H), 4.72 (m, 1H),
3.68−3.60 (m, 2H), 3.33−3.22 (m, 2H), 3.28 (s, 3H), 1.97−1.88 (m,
2H), 1.70−1.58 (m, 2H), 1.40 (s, 9H). HRMS (ESI): m/z calcd for
C25H30N2O5S [M + Na]+ 493.1773, obsd 493.1771. A solution of 7 (310
mg, 0.66 mmol) in acetic acid (4 mL) at RT was treated with NaCNBH3
(0.62 g, 9.9 mmol), and the mixture was stirred at RT for 36 h. The
mixture was then diluted with H2O (40 mL) and stirred for 30 min. The
resultant precipitant was filtered (water wash) and recrystallied from
methanol to afford 1,1-dimethylethyl 4-({1-[4-(methylsulfonyl)-
phenyl]-2,3-dihydro-1H-indol-4-yl}oxy)-1-piperidinecarboxylate 9
d6): δ 8.08 (s, 1H), 5.90−5.80 (m, 1H), 5.17−5.09 (app m, 2H), 3.65
(app d, J = 6.4 Hz, 2H). LCMS (ESI): m/z 190 [M + H]+. A slurry of
NaH (7.94 g, 60% dispersion in mineral oil, washed with anhydrous
toluene) in THF (50 mL) under N2 at RT was treated with 2-fluoro-4-
(methylsulfonyl)aniline (12.5 g, 66.1 mmol) in THF (150 mL). The
mixture was stirred at RT for 1 h and then treated with a solution of 17
(12.5 g, 66.1 mmol) in THF (50 mL). The resulting reaction mixture
was heated at reflux for 3 h, and then the mixture was allowed to cool to
room temperature, poured slowly onto H2O (600 mL), and then
extracted with EtOAc. The combined organic extracts were washed with
brine, dried over Na2SO4, and the filtrate concentrated. The crude
product was crystallized from EtOAc to give 10.3 g of solid material. The
mother liquor was concentrated and purified by flash chromatography to
give an additional 9.35 g of product. A total yield of 19.7 g (87%) of 6-
chloro-N-[2-fluoro-4-(methylsulfonyl)phenyl]-5-(2-propen-1-yl)-4-
pyrimidinamine (19) was obtained as an off-white solid. 1H NMR (400
MHz, DMSO-d6): δ 9.10 (s, 1H), 8.29 (s, 1H), 7.83 (dd, J = 9.2, 1.6 Hz,
1H), 7.78−7.72 (m, 2H), 5.96−5.86 (m, 1H), 5.10 (dd, J = 10.4, 1.2 Hz,
1H), 5.05 (dd, J = 16.0, 1.6 Hz, 1H), 3.58 (d, J = 5.6 Hz, 2H), 3.27 (s,
3H). LCMS (ESI): m/z 342 [M + H]+. A stream of ozone was bubbled
into a solution of 19 (15 g, 44 mmol) in CH2Cl2 (600 mL) and MeOH
(150 mL) at −78 °C for 4 h, at which time the solution turned light-blue.
A stream of N2 was bubbled through the solution until the blue
disappeared. Solid NaBH4 (6.7 g, 176 mmol) was added portionwise
over 10 min, and the reaction was stirred at −78 °C for 4 h and allowed
to warm to RT overnight. The reaction mixture was carefully poured
onto 1N HCl (1 L) and then was extracted with EtOAc (4 × 500 mL).
The organic layer was dried over MgSO4, filtered, and concentrated to
afford crude 2-(4-chloro-6-((2-fluoro-4-(methylsulfonyl)phenyl)-
amino)pyrimidin-5-yl)ethanol (20, 15 g). 1H NMR (400 MHz,
DMSO-d6): δ 9.57 (s, 1H), 8.38 (s, 1H), 8.03 (t, J = 8.2 Hz, 1H),
7.82 (dd, J = 10.4, 2.0 Hz, 1H), 7.74 (dd, J = 8.6, 1.8 Hz, 1H), 3.73 (t, J =
5.8 Hz, 2H), 3.25 (s, 3H), 2.97 (t, J = 5.8 Hz, 2H). LCMS (ESI): m/z
346 [M + H]+. A solution of 20 (15 g, 44 mmol) in CH2Cl2 (250 mL)
was cooled to 5 °C and treated with Et3N (36.4 mL, 263 mmol) and
Ms2O (15.3 g, 88 mmol), and the mixture was stirred and allowed to
warm to RT over a 15 h period. The reaction was diluted with CH2Cl2
(500 mL), quenched by the addition of water, and washed with water
and brine. The organic layer was dried over Na2SO4, filtered, and
concentrated. Purification by flash chromatography (0→2% MeOH/
CHCl3) afforded 4-chloro-7-[2-fluoro-4-(methylsulfonyl)phenyl]-6,7-
dihydro-5H-pyrrolo[2,3-d]pyrimidine (21, 6.6 g, 46%) as a light-yellow
solid. 1H NMR (400 MHz, DMSO-d6): δ 8.29 (s, 1H), 7.94 (app t, J =
7.6 Hz, 1H), 7.90 (dd, J = 15.6, 1.6 Hz, 1H), 7.79 (dd, J = 8.4, 2.0 Hz,
1H), 4.20 (t, J = 8.4 Hz, 2H), 3.27 (s, 3H), 3.23 (t, J = 8.8 Hz, 2H).
LCMS (ESI): m/z 328 [M + H]+. A slurry of NaH (0.73 g, 60%
dispersion in mineral oil, washed with anhydrous toluene) in anhydrous
THF (15 mL) was treated with a solution of 1-methylethyl 4-hydroxy-1-
piperidinecarboxylate (22, 1.09 g, 5.80 mmol) in THF (15 mL). The
mixture was heated at reflux for 0.5 h and then allowed to cool to RT. A
solution of 21 (2.0 g, mmol) in THF (30 mL) was added, and the
reaction was heated at reflux for 1.5 h. The reaction mixture was allowed
to cool to RT, poured onto H2O (150 mL), and then extracted with
CH2Cl2. The combined organic layer was dried over Na2SO4, filtered,
and then concentrated. The crude product was purified by flash
chromatography (0→2% MeOH/CHCl3) to afford 3 (1.96 g, 71%) as a
white solid. 1H NMR (400 MHz, DMSO-d6): δ 8.22 (s, 1H), 7.96 (app t,
J = 8.4 Hz, 1H), 7.84 (dd, J = 11.2, 2.0 Hz, 1H), 7.74 (dd, J = 8.4, 2.0 Hz,
1H), 5.30 −5.24 (m, 1H), 4.76 (septuplet, J = 6.0 Hz, 1H), 4.14 (t, J =
8.4 Hz, 2H), 3.70−3.64 (m, 2H), 3.25 (s, 3H), 3.40−3.21 (m, 2H), 3.05
(t, J = 8.8 Hz, 2H), 1.95−1.90 (m, 2H), 1.67−1.51 (m, 2H), 1.17 (d, J =
6.0 Hz, 6H). HRMS (ESI): m/z calcd for C22H27FN4O5S [M + H]+
479.1764, obsd 479.1765.
1
(218 mg, 71%) as a white solid. H NMR (400 MHz, DMSO-d6): δ
7.79 (t, J = 8.8 Hz, 2H), 7.35 (d, J = 8.8 Hz, 2H), 7.08 (app t, J = 8.4 Hz,
1H), 6.93 (d, J = 8.0 Hz, 1H), 6.60 (d, J = 8.0 Hz, 1H), 4.60−4.52 (m,
1H), 4.01 (t, J = 8.8 Hz, 2H), 3.61−3.55 (m, 2H), 3.30−3.21 (m, 2H),
3.13 (s, 3H), 3.01 (t, J = 8.4 Hz, 2H), 1.89−1.84 (m, 2H), 1.39 (s, 9H).
LCMS (ESI): m/z 495 [M + Na]+. HRMS (ESI): m/z calcd for
C25H32N2O5S [M + Na]+ 495.1930, obsd 495.1926. A solution of 9 (140
mg, 0.3 mmol) in CH2Cl2 (8 mL) at RT under N2 was treated with
CF3CO2H (2 mL), and the reaction mixture was stirred at RT for 2.5 h
and then concentrated. The resultant crude material was redissolved in
CH2Cl2 (4 mL) and treated with DIPEA (0.26 mL, 1.5 mmol), followed
by the dropwise addition of a 1M solution of isopropyl chloroformate in
toluene (3 mL, 3 mmol). The mixture was stirred at RT for 8 h. The
reaction mixture was diluted with CH2Cl2 (40 mL) and then washed
with water (10 mL) and brine (10 mL), dried over Na2SO4, and filtered,
and the filtrate was concentrated to afford the crude material. The
material was purified by flash chromatography (0→20% EtOAc/
hexanes) to afford 74 g (54%) of 2 as white solid. 1H NMR (400 MHz,
DMSO-d6): δ 7.79 (t, J = 8.8 Hz, 2H), 7.35 (d, J = 8.8 Hz, 2H), 7.08 (app
t, J = 8.4 Hz, 1H), 6.93 (d, J = 8.0 Hz, 1H), 6.60 (d, J = 8.4 Hz, 1H), 4.75
(septuplet, J = 6.0 Hz, 1H), 4.60 (m, 1H), 4.01 (t, J = 8.8 Hz, 2H), 3.60−
3.55 (m, 2H), 3.2−3.26 (m, 2H), 3.13 (s, 3H), 3.01 (t, J = 8.4 Hz, 2H),
1.89−1.60 (m, 2H), 1.59−1.52 (m, 2H), 1.17 (d, J = 6.4 Hz, 6H).
HRMS (ESI): m/z calcd for C24H30N2O5S [M + H]+ 459.1954, obsd
459.1954.
1-Methylethyl 4-({7-[2-Fluoro-4-(methylsulfonyl)phenyl]-
6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-4-yl}oxy)-1-piperidi-
necarboxylate (3). A solution of sodium methoxide (0.5 M in MeOH,
1.89 L, 0.95 mol) was treated with formamidine hydrochloride (39.0 g,
0.48 mol) under N2. The mixture was stirred at RT for 1 h. Diethyl
allylmalonate (91.5 mL, 0.46 mol) was added to the above mixture, and
the resulting reaction mixture was stirred at RT for 36 h. The reaction
mixture was filtered and the filtrate was concentrated to afford a crude
solid material. The crude material was dissolved in H2O (1000 mL) and
acidified with concentrated HCl. The solid was filtered and dried to
afford 62.8 g (90%) of 6-hydroxy-5-(2-propen-1-yl)-4(1H)-pyrimidi-
none as a white solid. 1H NMR (400 MHz, DMSO-d6): δ 11.65 (br s,
2H), 7.90 (s, 1H), 5.79−5.71 (m, 1H), 4.92 (app dd, J = 17.2, 2.0 Hz,
2H), 2.96 (d, J = 6.0 Hz, 2H). LCMS (ESI): m/z 153 [M + H]+. Under
N2, 6-hydroxy-5-(2-propen-1-yl)-4(1H)-pyrimidinone (19.0 g, 125.0
mmol) was treated with phosphorus oxychloride (200 mL). The
mixture heated at reflux for 4 h and then allowed to cool to RT. The
reaction mixture was poured very slowly onto ice-cold (5 °C) water
(2000 mL) with vigorous stirring. The mixture was extracted with
CH2Cl2. The combined organic layer was washed with saturated
aqueous NaHCO3 and brine, dried over Na2SO4, filtered, and the filtrate
concentrated to afford 22.7 g (96%) of 4,6-dichloro-5-(2-propen-1-
yl)pyrimidine (17) as a light-yellow oil. 1H NMR (400 MHz, DMSO-
1-Methylethyl 4-({1-[2-Fluoro-4-(methylsulfonyl)phenyl]-
2,3-dihydro-1H-indol-4-yl}oxy)-1-piperidinecarboxylate (10).
Compound 10 was prepared as a white solid in 10% overall yield
from 5 in a similar manner to that described for 2 using 4-bromo-2-
fluoro-1-iodobenzene. 1H NMR (400 MHz, DMSO-d6): δ 7.79 (dd, J =
11.6, 2.0 Hz, 1H), 7.68 (dd, J = 8.4, 2.0 Hz, 1H), 7.59 (app t, J = 8.4 Hz,
1H), 7.03 (t, J = 8.0 Hz, 1H), 6.56 (d, J = 8.4 Hz, 2H), 6.36 (dd, J = 7.6,
M
dx.doi.org/10.1021/jm301404a | J. Med. Chem. XXXX, XXX, XXX−XXX

Journal of Medicinal Chemistry
Article
4.4 Hz, 1H), 4.76 (septuplet, J = 6.4 Hz, 1H), 4.58 (m, 1H), 4.0 (t, J = 8.4
Hz, 1H), 3.68−3.57 (m, 2H), 3.30−3.21 (m, 2H), 3.22 (s, 3H), 3.02 (t, J
= 8.8 Hz, 2H), 1.92−1.82 (m, 2H), 1.61−1.52 (m, 2H), 1.17 (d, J = 6.4
Hz, 6H). HRMS (ESI): m/z calcd for C24H29FN2O5S [M + H]+
461.1859, obsd 461.1860.
mL) under N2 was treated with PdCl2(Ph3P)2 (16 mg, 0.023 mmol) and
2-(tributylstannanyl)-1,3-thiazole (85 mg, 0.23 mmol), and the reaction
mixture was refluxed for 12 h. The mixture was quenched by the addition
of water and extracted with EtOAc. The organic extracts were washed
with brine, dried over MgSO4, and concentrated. Purification by flash
1
chromatography afforded 26 (66 mg, 61%) as an off-white solid. H
tert-Butyl 4-((7-(4-(Methylsulfonyl)phenyl)-7H-pyrrolo[2,3-
d]pyrimidin-4-yl)oxy)piperidine-1-carboxylate (14). 7-(4-Bro-
mophenyl)-4-chloro-7H-pyrrolo[2,3-d]pyrimidine (12) was prepared
as an off-white solid in 30% yield from 4-chloro-7H-pyrrolo[2,3-
d]pyrimidine in a similar manner to that described for 6 using 4-bromo-
1-iodobenzene. 1H NMR (400 MHz, DMSO-d6): δ 8.70 (s, 1H), 8.15 (s,
1H), 7.93 (d, J = 8.5 Hz, 1H), 7.80 (m, 2H), 7.68 (d, J = 8.5 Hz, 1H),
6.89 (m, 1H). LRMS (ESI): m/z 310/308 [M + H]+. A solution of 12
(420 mg, 1.36 mmol) in DMF (5 mL) was treated with tert-butyl 4-
hydroxy-1-piperidinecarboxylate (1.2 g, 3.6 mmol) and Cs2CO3 (0.72 g,
3.6 mmol), and the mixture was stirred at heated at 110 °C for 18 h
under N2. The mixture was allowed to cool and then was diluted with
EtOAc and water. The organic layer was washed with water and brine
dried over Na2SO4, and filtered. The filtrate was concentrated and
purified by flash chromatography to give tert-butyl 4-((7-(4-
bromophenyl)-7H-pyrrolo[2,3-d]pyrimidin-4-yl)oxy)piperidine-1-car-
NMR (400 MHz, DMSO-d6): δ 8.18 (s, 1H), 7.92 (d, J = 3.2 Hz, 1H),
7.84−7.79 (m, 2H), 7.78−7.74 (m, 2H), 5.26 (m, 1H), 4.76 (septuplet, J
= 6.0 Hz, 1H), 4.10 (t, J = 8.8 Hz, 1H), 3.71−3.65 (m, 2H), 3.25−3.21
(m, 2H), 3.05 (t, J = 8.8 Hz, 2H), 1.95−1.91 (m, 2H), 1.61−1.53 (m,
2H), 1.17 (d, J = 6.4 Hz, 6H). HRMS (ESI): m/z calcd for
C24H26FN5O3S [M + H]+ 484.1819, obsd 484.1817.
1-Methylethyl 4-({7-[2-Fluoro-4-(1-methyl-1H-pyrrol-2-yl)-
phenyl]-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-4-yl}oxy)-1-
piperidinecarboxylate (27). Compound 27 was prepared as a white
solid in 12% yield from 24 in a similar manner to that described for 26
using 1-methyl-2-(tributylstannanyl)-1H-pyrrole. 1H NMR (400 MHz,
DMSO-d6): δ 8.12 (s, 1H), 7.58 (t, J = 8.4 Hz, 1H), 7.36 (dd, J = 12.4, 2.0
Hz, 1H), 7.27 (dd, J = 8.4, 1.6 Hz, 1H), 6.84 (t, J = 2.0 Hz, 1H), 6.23 (dd,
J = 3.6, 2.0 Hz, 1H), 6.04 (t, J = 3.2 Hz, 1H), 5.28−5.22 (m, 1H), 4.76
(septuplet, J = 6.0 Hz, 1H), 4.04 (t, J = 9.2 Hz, 2H), 3.67 (s, 3H), 3.71−
3.65 (m, 2H), 3.23 (app t, J = 10.0 Hz, 2H), 3.04 (t, J = 9.2 Hz, 2H),
1.95−1.90 (m, 2H), 1.61−1.52 (m, 2H), 1.17 (d, J = 6.0 Hz, 6H).
HRMS (ESI): m/z calcd for C26H30FN5O3 [M + H]+ 480.2411, obsd
480.2411.
1,1-Dimethylethyl 4-({7-[2-Fluoro-4-(methylsulfonyl)-
phenyl]-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-4-yl}oxy)-1-
piperidinecarboxylate (28). Compound 28 was prepared as a light-
yellow solid in 73% yield from 21 in a similar manner to that described
for 3 using 1,1-dimethylethyl 4-hydroxy-1-piperidinecarboxylate. 1H
NMR (400 MHz, DMSO-d6): δ 8.22 (s, 1H), 7.96 (app t, J = 7.6 Hz,
1H), 7.84 (dd, J = 11.2, 2.0 Hz, 1H), 7.74 (dd, J = 8.4, 2.0 Hz, 1H), 5.26
(m, 1H), 4.14 (t, J = 8.0 Hz, 2H), 3.68−3.62 (m, 2H), 3.25 (s, 3H),
3.18−3.16 (m, 2H), 3.05 (t, J = 9.2 Hz, 2H), 1.94−1.89 (m, 2H), 1.59−
1.53 (m, 2H), 1.39 (s, 9H). HRMS (ESI): m/z calcd for C23H29FN4O5S
[M + H]+ 493.1921, obsd 493.1921.
Methyl 4-({7-[2-Fluoro-4-(methylsulfonyl)phenyl]-6,7-dihy-
dro-5H-pyrrolo[2,3-d]pyrimidin-4-yl}oxy)-1-piperidinecarbox-
ylate (29). A solution of 28 (1.0 g, 2.0 mmol) in CH2Cl2 (40 mL) was
treated with CF3CO2H (10 mL) at RT and stirred for 2 h. The reaction
mixture was concentrated and then redissolved in CH2Cl2 (20 mL) and
treated with Et3N (1.7 mL, ∼6 equiv). A portion of the material (2.2 mL,
∼0.20 mmol) was treated with methyl chloroformate (200 μL) at RT for
18 h. The reaction mixture was diluted with CH2Cl2 (30 mL) and then
washed with water and brine, dried over Na2SO4, and then concentrated
to afford the crude product, which was purified by flash chromatography
to afford 29 (76 mg, 85%) as a white solid. 1H NMR (400 MHz, DMSO-
d6): δ 8.22 (s, 1H), 7.96 (t, J = 8.0 Hz, 1H), 7.84 (dd, J = 11.2, 2.0 Hz,
1H), 7.73 (dd, J = 8.4, 2.4 Hz, 1H), 5.30−5.22 (m, 1H), 4.14 (t, J = 8.4
Hz, 2H), 3.58 (s, 3H), 3.70−3.64 (m, 2H), 3.28−3.24 (app br m, 2H),
3.25 (s, 3H), 3.05 (t, J = 8.8 Hz, 2H), 1.96−1.91 (m, 2H), 1.64−1.54 (m,
2H). HRMS (ESI): m/z calcd for C20H23FN4O5S [M + H]+ 451.1451,
obsd 451.1452.
1
boxylate (13, 0.45 g, 70%) as a white foam. H NMR (400 MHz,
DMSO-d6): δ 8.46 (s, 1H), 7.89 (d, J = 8.8 Hz, 1H), 7.84 (m, 2H), 7.74
(d, J = 8.9 Hz, 1H), 7.69 (d, J = 8.8 Hz, 1H), 6.75 (m, 1H), 5.48 (m, 1H),
3.70 (m, 2H), 3.23 (m, 2H), 1.98 (m, 2H), 1.66 (m, 2H), 1.40 (s, 9H).
LRMS (ESI): m/z 475/473 [M + H]+. tert-Butyl 4-((7-(4-
(methylsulfonyl)phenyl)-7H-pyrrolo[2,3-d]pyrimidin-4-yl)oxy)-
piperidine-1-carboxylate (14) was prepared as an off-white solid in 84%
yield from 13 in a similar manner to that described for 7. 1H NMR (400
MHz, DMSO-d6): δ 8.51 (s, 1H), 8.21 (d, J = 8.8 Hz, 2H), 8.09 (d, J =
8.8 Hz, 2H), 7.88 (d, J = 3.7 Hz, 1H), 6.83 (d, J = 3.7 Hz, 1H), 5.50 (m,
1H), 3.74−3.66 (m, 2H), 3.27 (s, 3H), 2.48 (m, 2H), 2.03−1.99 (m,
2H), 1.70−1.62 (m, 2H), 1.40 (s, 9H). LRMS (ESI): m/z 473 [M + H]+.
1-Methylethyl 4-({7-[4-(Methylsulfonyl)phenyl]-6,7-dihydro-
5H-pyrrolo[2,3-d]pyrimidin-4-yl}oxy)-1-piperidinecarboxylate
(23). Compound 23 was prepared as an off-white solid in 34% overall
yield from 17 in a similar manner to that described for 3 using 4-
bromoaniline. 1H NMR (400 MHz, CDCl3): δ 8.39 (s, 1H), 7.98−7.90
(m, 4H), 5.38 (m, 1H), 4.93 (septuplet, J = 6.4 Hz, 1H), 4.15 (t, J = 8.8
Hz, 2H), 3.83 (br m, 2H), 3.37−3.30 (m, 2H), 3.12 (t, J = 8.8 Hz, 2H),
3.04 (s, 3H), 2.12−1.96 (m, 2H), 1.80−1.69 (m, 2H), 1.25 (d, J = 6.4
Hz, 6H). HRMS (ESI): m/z calcd for C22H28N4O5S [M + H]+
477.1859, obsd 477.1857.
1-Methylethyl 4-{[7-(4-Cyano-2-fluorophenyl)-6,7-dihydro-
5H-pyrrolo[2,3-d]pyrimidin-4-yl]oxy}-1-piperidinecarboxylate
(25). 1-Methylethyl 4-{[7-(4-bromo-2-fluorophenyl)-6,7-dihydro-5H-
pyrrolo[2,3-d]pyrimidin-4-yl]oxy}-1-piperidinecarboxylate (24) was
prepared as an off-white solid in 19% overall yield from 17 in a similar
manner to that described for 3 using 4-bromo-2-fluoroaniline. 1H NMR
(400 MHz, DMSO-d6): δ 8.13 (s, 1H), 7.63 (dd, J = 10.8, 2.0 Hz, 1H),
7.56 (app t, J = 8.4 Hz, 1H), 7.42 (dd, J = 8.8, 2.0 Hz, 1H), 5.25 (m, 1H),
4.75 (septuplet, J = 6.4 Hz, 1H), 4.00 (t, J = 9.2 Hz, 2H), 3.72−3.68 (m,
2H), 3.22 (m, 2H), 3.02 (t, J = 8.8 Hz, 2H), 1.94−1.89 (m, 2H), 1.60−
1.55 (m, 2H), 1.17 (d, J = 6.4 Hz, 6H). LCMS (ESI): m/z 481 (M + H)+.
A solution of 24 (100 mg, 0.209 mmol) in NMP (2 mL) was treated with
CuCN (42 mg, 0.47 mmol) under N2. The resulting mixture was heated
to 150 °C for 15 h. The mixture was cooled to RT, poured onto H2O (25
mL), and the mixture extracted with EtOAc. The combined organic
layer was washed with brine, dried over Na2SO4, filtered, and
concentrated. The crude product was purified by flash SiO2 column
chromatography to afford 25 (76 mg, 85%) as a white solid. 1H NMR
(400 MHz, DMSO-d6): δ8.23 (s, 2H), 7.95−7.89 (m, 2H), 7.68 (dd, J =
8.0, 2.0 Hz, 1H), 5.27 (m, J = 4.0 Hz, 1H), 4.76 (septuplet, J = 6.4 Hz,
1H), 4.14 (t, J = 8.0 Hz, 2H), 3.70−3.64 (m, 2H), 3.25−3.21 (m, 2H),
3.04 (t, J = 8.4 Hz, 2H), 1.95−1.90 (m, 2H), 1.61−1.53 (m, 2H), 1.17
(d, J = 6.0 Hz, 6H). HRMS (ESI): m/z calcd for C22H24FN5O3 [M + H]+
426.1942, obsd 426.1942.
Ethyl 4-({7-[2-Fluoro-4-(methylsulfonyl)phenyl]-6,7-dihydro-
5H-pyrrolo[2,3-d]pyrimidin-4-yl}oxy)-1-piperidinecarboxylate
(30). Compound 30 was prepared as a white solid in 91% yield from 28
in a similar manner to that described for 29 using ethyl chloroformate.
1H NMR (400 MHz, DMSO-d6): δ 8.22 (s, 1H), 7.96 (t, J = 7.6 Hz, 1H),
7.83 (dd, J = 11.2, 2.0 Hz, 1H), 7.74 (dd, J = 8.4, 2.0 Hz, 1H), 5.28−5.24
(m, 1H), 4.14 (t, J = 8.4 Hz, 2H), 4.02 (q, J = 7.2 Hz, 2H), 3.71−3.65 (m,
2H), 3.26−3.24 (br s, 2H), 3.25 (s, 3H), 3.05 (t, J = 8.8 Hz, 2H), 1.96−
1.90 (br m, 2H), 1.62−1.54 (m, 2H), 1.17 (t, J = 6.8 Hz, 3H). HRMS
(ESI): m/z calcd for C21H25FN4O5S [M + H]+ 465.1605, obsd
465.1605.
n-Propyl 4-({7-[2-Fluoro-4-(methylsulfonyl)phenyl]-6,7-dihy-
dro-5H-pyrrolo[2,3-d]pyrimidin-4-yl}oxy)-1-piperidinecarbox-
ylate (31). Compound 31 was prepared as a white solid in 75% yield
from 28 in a similar manner to that described for 29 using n-propyl
chloroformate. 1H NMR (400 MHz, DMSO-d6): δ 8.22 (s, 1H), 7.96 (t,
J = 8.0 Hz, 1H), 7.84 (dd, J = 10.8, 2.0 Hz, 1H), 7.74 (dd, J = 8.4, 2.4 Hz,
1H), 5.30−5.24 (m, 1H), 4.14 (t, J = 8.4 Hz, 2H), 3.94 (t, J = 6.8 Hz,
1-Methylethyl 4-({7-[2-Fluoro-4-(1,3-thiazol-2-yl)phenyl]-
6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-4-yl}oxy)-1-piperidi-
necarboxylate (26). A solution of 24 (108 mg, 0.23 mmol) in THF (4
N
dx.doi.org/10.1021/jm301404a | J. Med. Chem. XXXX, XXX, XXX−XXX

Journal of Medicinal Chemistry
Article
2H), 3.72−3.66 (m, 2H), 3.30−3.24 (br m, 2H), 3.05 (t, J = 8.4 Hz, 2H),
1.98−1.88 (m, 2H), 1.62−1.52 (m, 4H), 0.87 (t, J = 7.2 Hz, 2H). HRMS
(ESI): m/z calcd for C22H27FN4O5S [M + H]+ 479.1764, obsd
479.1764.
3H), 1.96 (br s, 2H), 1.62 (m, 2H). HRMS (ESI): m/z calcd for
C20H23FN4O4S2 [M + H]+ 467.1222, obsd 467.1222.
S-Ethyl 4-({7-[2-Fluoro-4-(methylsulfonyl)phenyl]-6,7-dihy-
dro-5H-pyrrolo[2,3-d]pyrimidin-4-yl}oxy)-1-piperidinecarbo-
thioate (38). Compound 38 was prepared as a white solid in 87% yield
from 28 in a similar manner to that described for 29 using S-ethyl
chloroformate. 1H NMR (400 MHz, DMSO-d6): δ 8.23 (s, 1H), 7.96 (t,
J = 8.0 Hz, 1H), 7.84 (dd, J = 11.2, 2.0 Hz, 1H), 7.74 (dd, J = 8.4, 2.0 Hz,
1H), 5.36−5.29 (m, 1H), 4.14 (t, J = 8.4 Hz, 2H), 3.76 (app br s, 2H),
3.41−3.36 (m, 2H), 3.25 (s, 3H), 3.06 (t, J = 8.8 Hz, 2H), 2.81 (q, J = 7.2
Hz, 2H), 1.96 (br s, 2H), 1.62 (br s, 2H), 1.18 (t, J = 7.6 Hz, 2H). HRMS
(ESI): m/z calcd for C21H25FN4O4S2 [M + H]+ 481.1380, obsd
481.1380.
n-Butyl 4-({7-[2-Fluoro-4-(methylsulfonyl)phenyl]-6,7-dihy-
dro-5H-pyrrolo[2,3-d]pyrimidin-4-yl}oxy)-1-piperidinecarbox-
ylate (32). Compound 32 was prepared as a white solid in 77% yield
from 28 in a similar manner to that described for 29 using n-butyl
chloroformate. 1H NMR (400 MHz, DMSO-d6): δ 8.22 (s, 1H), 7.96 (t,
J = 8.0 Hz, 1H), 7.84 (dd, J = 10.8, 2.0 Hz, 1H), 7.74 (dd, J = 8.4, 2.4 Hz,
1H), 5.30−5.24 (m, 1H), 4.14 (t, J = 8.0 Hz, 2H), 3.98 (t, J = 6.4 Hz,
2H), 3.71−3.66 (m, 2H), 3.26−3.24 (br s, 2H), 3.25 (s, 3H), 3.05 (t, J =
8.8 Hz, 2H), 1.96−1.90 (m, 2H), 1.62−1.49 (m, 4H), 1.36−1.26 (m,
2H), 0.88 (t, J = 7.6 Hz, 3H). HRMS (ESI): m/z calcd for
C23H29FN4O5S [M + H]+ 493.1921, obsd 493.1920.
2-Methylpropyl 4-({7-[2-Fluoro-4-(methylsulfonyl)phenyl]-
6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-4-yl}oxy)-1-piperidi-
necarboxylate (33). Compound 33 was prepared as a white solid in
83% yield from 21 in a similar manner to that described for 3 using 2-
methylpropyl 4-hydroxy-1-piperidinecarboxylate. 1H NMR (400 MHz,
DMSO-d6): δ 8.22 (s, 1H), 7.96 (app t, J = 8.0 Hz, 1H), 7.84 (dd, J =
10.8, 1.6 Hz, 1H), 7.74 (dd, J = 8.8, 2.0 Hz, 1H), 5.28 (m, 1H), 4.14 (t, J
= 8.0 Hz, 2H), 3.78 (d, J = 6.8 Hz, 2H), 3.71−3.68 (m, 2H), 3.25 (s, 3H),
3.30−3.27 (m, 2H), 3.05 (t, J = 8.8 Hz, 2H), 1.97−1.91 (m, 2H), 1.88−
1.80 (m, 1H), 1.63−1.54 (m, 2H), 0.87 (d, J = 6.8 Hz, 6H). HRMS
(ESI): m/z calcd for C23H29FN4O5S [M + H]+ 493.1921, obsd
493.1920.
S-(1-Methylethyl) 4-({7-[2-Fluoro-4-(methylsulfonyl)phenyl]-
6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-4-yl}oxy)-1-piperidi-
necarbothioate (39). A solution of 28 (2.0 g, 4.1 mmol) was treated
with a 20% solution of CF3CO2H in CH2Cl2 (50 mL) at RT, and the
mixture was stirred for 3 h. The reaction mixture was concentrated and
then redissolved in CH2Cl2 (150 mL) and treated with 1,1′-
(oxomethanediyl)bis-1H-imidazole (725 mg, 4.47 mmol) at 5 °C.
The reaction mixture was allowed to warm to ambient temperature and
allowed to stir for 24 h. The reaction mixture was diluted with CH2Cl2
(200 mL) and washed with water, and the organic layer was dried over
Na2SO4 and then concentrated to afford crude product, which was
recrystallized from CH2Cl2 to give (1.9 g, 97%) of 7-[2-fluoro-4-
(methylsulfonyl)phenyl]-4-{[1-(1H-imidazol-1-ylcarbonyl)-4-
piperidinyl]oxy}-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidine as a white
solid. 1H NMR (400 MHz, DMSO-d6): δ 8.24 (s, 1H), 8.02 (s, 1H), 7.96
(app t, J = 8.0 Hz, 1H), 7.84 (dd, J = 11.2, 2.4 Hz, 1H), 7.74 (dd, J = 8.4,
2.0 Hz, 1H), 7.47 (s, 1H), 7.02 (s, 1H), 5.40−5.28 (m, 1H), 4.15 (t, J =
8.4 Hz, 2H), 3.71 (m, 2H), 3.48−3.42 (m, 2H), 3.25 (s, 3H), 3.07 (t, J =
9.2 Hz, 2H), 2.10−2.02 (m, 2H), 1.83−1.75 (m, 2H); LRMS (ESI): m/z
487 (M + H)+. A solution of 7-[2-fluoro-4-(methylsulfonyl)phenyl]-4-
{[1-(1H-imidazol-1-ylcarbonyl)-4-piperidinyl]oxy}-6,7-dihydro-5H-
pyrrolo[2,3-d]pyrimidine (1.9 g, 3.90 mmol) in CH3CN (30 mL) and
CH2Cl2 (30 mL) was treated with methyl iodide (1.46 mL, 23.5 mmol)
at RT under N2. The reaction mixture was stirred at RT for 24 h and then
concentrated to afford 1-{[4-({7-[2-fluoro-4-(methylsulfonyl)phenyl]-
6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-4-yl}oxy)-1-piperidinyl]-
carbonyl}-3-methyl-1H-imidazol-3-ium iodide (2.46 g, 100%) as an off-
2,2-Dimethylpropyl 4-({7-[2-Fluoro-4-(methylsulfonyl)-
phenyl]-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-4-yl}oxy)-1-
piperidinecarboxylate (34). Compound 34 was prepared as a white
solid in 72% yield from 21 in a similar manner to that described for 3
1
using 2,2-dimethylpropyl 4-hydroxy-1-piperidinecarboxylate. H NMR
(400 MHz, DMSO-d6): δ 8.23 (s, 1H), 7.96 (app t, J = 8.0 Hz, 1H), 7.84
(dd, J = 10.8, 1.6 Hz, 1H), 7.74 (dd, J = 8.8, 2.0 Hz, 1H), 5.29 (m, 1H),
4.14 (t, J = 8.4 Hz, 2H), 3.70 (s, 2H), 3.73−3.64 (m, 2H), 3.25 (s, 3H),
3.30−3.27 (m, 2H), 3.06 (t, J = 8.8 Hz, 2H), 1.97−1.90 (m, 2H), 1.62−
1.52 (m, 2H), 0.89 (s, 9H). HRMS (ESI): m/z calcd for C24H31FN4O5S
[M + H]+ 507.2077, obsd 507.2078.
2-Fluoroethyl 4-({7-[2-Fluoro-4-(methylsulfonyl)phenyl]-6,7-
dihydro-5H-pyrrolo[2,3-d]pyrimidin-4-yl}oxy)-1-piperidinecar-
boxylate (35). Compound 35 was prepared as a white solid in 58%
yield from 28 in a similar manner to that described for 29 using 2-
fluoroethyl chloridocarbonate. 1H NMR (400 MHz, DMSO-d6): δ 8.23
(s, 1H), 7.96 (t, J = 8.0 Hz, 1H), 7.84 (dd, J = 10.8, 2.4 Hz, 1H), 7.74 (dd,
J = 8.8, 2.0 Hz, 1H), 5.31−5.25 (m, 1H), 4.65 (app t, J = 4.0 Hz, 1H),
4.53 (app t, J = 3.6 Hz, 1H), 4.27 (app t, J = 4.0 Hz, 1H), 4.19 (app t, J =
4.0 Hz, 1H), 4.14 (t, J = 8.0 Hz, 2H), 3.70 (m, 2H), 3.28−3.25 (br s,
2H), 3,25 (s, 3H), 3.06 (t, J = 8.8 Hz, 2H), 1.98−1.90 (m, 2H), 1.65−
1.56 (m, 2H). HRMS (ESI): m/z calcd for C21H24F2N4O5S [M + H]+
483.1514, obsd 483.1514.
2-(Methyloxy)ethyl 4-({7-[2-Fluoro-4-(methylsulfonyl)-
phenyl]-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-4-yl}oxy)-1-
piperidinecarboxylate (36). Compound 36 was prepared as a white
solid in 65% yield from 28 in a similar manner to that described for 29
using 2-(methyloxy)ethyl chloroformate. 1H NMR (400 MHz, DMSO-
d6): δ 8.22 (s, 1H), 7.96 (t, J = 7.6 Hz, 1H), 7.84 (dd, J = 11.2, 2.0 Hz,
1H), 7.74 (dd, J = 8.4, 2.0 Hz, 1H), 5.30−5.26 (m, 1H), 4.16−4.09 (m,
2H), 3.72−3.66 (m, 2H), 3.50 (t, J = 4.4 Hz, 2H), 3.24 (s, 3H), 3.28−
3.24 (br s, 2H), 3.05 (t, J = 8.8 Hz, 2H), 5.30−5.26 (m, 1H), 4.16−4.09
(m, 2H), 3.72−3.66 (m, 2H), 3.50 (t, J = 4.4 Hz, 2H), 3.28−3.24 (br s,
2H), 3.24 (s, 6H), 3.05 (t, J = 8.8 Hz, 2H), 1.98−1.90 (m, 2H), 1.63−
1.54 (m, 2H). HRMS (ESI): m/z calcd for C22H27FN4O6S [M + H]+
495.1714, obsd 495.1715.
1
white solid. H NMR (400 MHz, DMSO-d6): δ 9.55 (s, 1H), 8.24 (s,
1H), 8.03 (app t, J = 3.6 Hz, 1H), 7.96 (t, J = 8.0 Hz, 1H), 7.86−7.82 (m,
2H), 7.75 (dd, J = 8.4, 2.0 Hz, 1H), (app t, J = 8.0 Hz, 1H), 5.42 (m, 1H),
4.16 (t, J = 8.4 Hz, 2H), 3.90 (s, 3H), 3.71 (br s, 2H), 3.50 (br s, 2H),
3.25 (s, 3H), 3.07 (t, J = 9.2 Hz, 2H), 2.12−2.04 (br m, 2H), 1.88−1.80
(m, 2H). A suspension of the above salt (315 mg, 0.5 mmol) in CH2Cl2
(3 mL) was treated with Et3N (70 μL, 0.5 mmol) and isopropylthiol (38
mg, 0.5 mmol) at RT under N2. The reaction mixture was stirred at RT
for 18 h and then quenched by the addition of water. The aqueous was
extracted with EtOAc, and the organic layer was dried over MgSO4 and
concentrated. Purification by flash chromatography afforded 39 (156
mg, 63%) as a white solid. 1H NMR (400 MHz, DMSO-d6): δ 8.23 (s,
1H), 7.96 (app t, J = 8.0 Hz, 1H), 7.84 (app br d, J = 10.8 Hz, 1H), 7.74
(app br d, J = 6.8 Hz, 1H), 5.30−5.28 (m, 1H), 4.14 (t, J = 8.4 Hz, 2H),
3.46 (m, 1H), 3.37−3.30 (br m, 2H), 3.25 (s, 3H), 3.06 (t, J = 8.4 Hz,
2H), 1.94 (app br s, 2H), 1.61 (app br s, 2H), 1.26 (d, J = 6.4 Hz, 6H).
HRMS (ESI): m/z calcd for C22H27FN4O4S2 [M + H]+ 495.1536, obsd
495.1536.
Phenyl 4-({7-[2-Fluoro-4-(methylsulfonyl)phenyl]-6,7-dihy-
dro-5H-pyrrolo[2,3-d]pyrimidin-4-yl}oxy)-1-piperidinecarbodi-
thioate (40). Compound 40 was prepared as a white solid in 55% yield
from 28 in a similar manner to that described for 29 using phenyl
S-Methyl 4-({7-[2-Fluoro-4-(methylsulfonyl)phenyl]-6,7-di-
hydro-5H-pyrrolo[2,3-d]pyrimidin-4-yl}oxy)-1-piperidinecar-
bothioate (37). Compound 37 was prepared as a white solid in 85%
yield from 28 in a similar manner to that described for 29 using S-methyl
1
chloridodithiocarbonate. H NMR (400 MHz, DMSO-d6): δ 8.25 (s,
1H), 7.96 (t, J = 8.0 Hz, 1H), 7.85 (dd, J = 11.2, 2.0 Hz, 1H), 7.75 (dd, J =
8.4, 1.6 Hz, 1H), 7.48−7.40 (m, 5H), 5.48−5.42 (m, 1H), 4.46 (app br s,
1H), 4.22 (br s, 2H), 4.16 (t, J = 8.4 Hz, 2H), 4.06 (br s, 2H), 3.26 (s,
3H), 3.09 (t, J = 8.8 Hz, 2H), 2.20−2.06 (br s, 2H), 1.92−1.72 (br s,
2H). HRMS (ESI): m/z calcd for C25H25FN4O3S3 [M + H]+ 545.1152,
obsd 545.1152.
1
chloroformate. H NMR (400 MHz, DMSO-d6): δ 8.22 (s, 1H), 7.96
(app t, J = 8.4 Hz, 1H), 7.84 (dd, J = 10.8, 2.0 Hz, 1H), 7.74 (dd, J = 8.4,
2.0 Hz, 1H), 5.36−5.28 (m, 2H), 4.14 (t, J = 8.0 Hz, 2H), 3.74 (br s,
2H), 3.42−3.34 (m, 2H), 3.25 (s, 3H), 3.06 (t, J = 8.8 Hz, 2H), 2.24 (s,
O
dx.doi.org/10.1021/jm301404a | J. Med. Chem. XXXX, XXX, XXX−XXX

Journal of Medicinal Chemistry
Article
4-{[1-(Ethylsulfonyl)-4-piperidinyl]oxy}-7-[2-fluoro-4-
(methylsulfonyl)phenyl]-6,7-dihydro-5H-pyrrolo[2,3-d]-
pyrimidine (41). Compound 41 was prepared as a tan solid in 70%
yield from 28 in a similar manner to that described for 29 using
ethanesulfonyl chloride. 1H NMR (400 MHz, CDCl3): δ 8.27 (s, 1H),
7.86 (app t, J = 8.5 Hz, 1H), 7.78−7.68 (m, 2H), 5.42−5.25 (m, 1H),
4.26 (t, J = 8.5 Hz, 2H), 3.69−3.52 (m, 2H), 3.35−3.23 (m, 2H), 3.19 (t,
J = 8.5 Hz, 2H), 3.06 (s, 3H), 2.99 (q, J = 7.8 Hz, 2H), 2.20−2.05 (m,
2H), 1.98−1.81 (m, 2H), 1.38 (t, J = 8.5 Hz 3H). HRMS (ESI): m/z
calcd for C20H25FN4O5S2 [M + H]+ 485.1329, obsd 485.1328.
7-[2-Fluoro-4-(methylsulfonyl)phenyl]-4-({1-[(1-
methylethyl)sulfonyl]-4-piperidinyl}oxy)-6,7-dihydro-5H-
pyrrolo[2,3-d]pyrimidine (42). Compound 42 was prepared as a tan
solid in 54% yield from 28 in a similar manner to that described for 29
using 2-propanesulfonyl chloride. 1H NMR (400 MHz, CDCl3): δ 8.31
(s, 1H), 7.91 (app t, J = 8.5 Hz, 1H), 7.87−7.60 (m, 2H), 5.49−5.20 (m,
1H), 4.31 (t, J = 8.5 Hz, 2H), 3.80−3.52 (m, 2H), 3.42−3.26 (m, 2H),
3.24−3.12 (m, 4 H), 2.23−1.99 (m, 2H), 1.98−1.81 (m, 2H), 1.97−1.75
(m, 2H), 1.32 (d, J = 6.8 Hz, 6H). HRMS (ESI): m/z calcd for
C21H27FN4O5S2 [M + H]+ 499.1485, obsd 499.1484.
1H), 7.87 (app br d, J = 10.8 Hz,1H), 7.78 (app br t, J = 8.4 Hz, 1H), 5.36
(br m, 1H), 4.18 (t, J = 8.8 Hz, 2H), 3.82−3.78 (m, 2H), 3.56−3.50 (m,
2H), 3.28 (s, 3H), 3.10 (t, J = 8.4 Hz, 2H), 2.83 (septuplet, J = 6.8 Hz,
1H), 2.09 (m, 2H), 1.78 (m, 2H), 1.20 (d, J = 6.8 Hz, 6H). HRMS (ESI):
m/z calcd for C23H27FN6O4S [M + H]+ 503.1877, obsd 503.1879.
7-[2-Fluoro-4-(methylsulfonyl)phenyl]-4-{[1-(5-fluoro-2-pyri-
midinyl)-4-piperidinyl]oxy}-6,7-dihydro-5H-pyrrolo[2,3-d]-
pyrimidine (56). A solution of 4-hydroxypiperidine (45, 1.53 g, 15.1
mmol) in acetonitrile (50 mL) was treated with diisopropylethylamine
(7.9 mL, 45.3 mmol) and 5-fluoro-2-chloropyrimidine (48, 2.0 g, 15.1
mmol) under N2, and the reaction mixture was refluxed for 10 h. The
reaction was concentrated and then purified by flash chromatography to
afford 1-(5-fluoro-2-pyrimidinyl)-4-piperidinol (50, 2.46 g, 83%) as a
white solid. 1H NMR (400 MHz, DMSO-d6): δ 8.18 (s, 2H), 4.37−4.31
(m, 2H), 3.97−3.91 (m, 1H), 3.32−3.25 (m, 2H), 1.98−1.92 (m, 2H),
1.77 (br s, 1H), 1.57−1.48 (m, 2H). LCMS (ESI): m/z 198 [M + H]+.
Compound 56 was prepared as a white solid in 39% yield from 21 in a
similar manner to that described for 3 using 1-(5-fluoro-2-pyrimidinyl)-
1
4-piperidinol (50). H NMR (400 MHz, DMSO-d6): δ 8.44 (s, 2H),
8.24 (s, 1H), 7.96 (app t, J = 7.6 Hz, 1H), 7.84 (dd, J = 10.8, 2.0 Hz, 1H),
7.74 (dd, J = 8.4, 1.6 Hz, 1H), 5.36 (m, 1H), 4.16−4.10 (m, 2H), 4.14 (t,
J = 8.4 Hz, 2H), 3.55−3.44 (m, 2H), 3.25 (s, 3H), 3.05 (t, J = 8.8 Hz,
2H), 2.03−1.98 (m, 2H), 1.66−1.60 (m, 2H). HRMS (ESI): m/z calcd
for C22H22F2N6O3S [M + H]+ 461.1859, obsd 461.1859
4-{[1-(5-Ethyl-2-pyrimidinyl)-4-piperidinyl]oxy}-7-[2-fluoro-
4-(methylsulfonyl)phenyl]-6,7-dihydro-5H-pyrrolo[2,3-d]-
pyrimidine (57). 1-(5-Ethyl-2-pyrimidinyl)-4-piperidinol (51) was
prepared in a similar manner to that described for 1-(5-fluoro-2-
pyrimidinyl)-4-piperidinol (50) using 5-ethyl-2-chloropyrimidine (49).
1H NMR (400 MHz, DMSO-d6): δ 8.19 (s, 1H), 4.67 (br s, 1H), 4.24−
4-({7-[2-Fluoro-4-(methylsulfonyl)phenyl]-6,7-dihydro-5H-
pyrrolo[2,3-d]pyrimidin-4-yl}oxy)-N,N-dimethyl-1-piperidine-
sulfonamide (43). Compound 43 was prepared as a tan solid in 54%
yield from 28 in a similar manner to that described for 29 using N,N-
1
dimethylsulfamoyl chloride. H NMR (400 MHz, CDCl3): δ 8.28 (s,
1H), 8.00 (app t, J = 8.5 Hz, 1H), 7.76−7.67 (m, 2H), 5.38−5.29 (m,
1H), 4.19 (t, J = 8.5 Hz, 2H), 3.57−3.48 (m, 2H), 3.29−3.20 (m, 2H),
3.13 (t, J = 8.5 Hz, 2H), 3.04 (s, 3H), 2.84 (s, 6H), 2.13−2.02 (m, 2H),
1.96−1.85 (m, 2H). HRMS (ESI): m/z calcd for C20H26FN5O5S2 [M +
H]+ 500.1438, obsd 500.1438.
N,N-Diethyl-2-[4-({7-[2-fluoro-4-(methylsulfonyl)phenyl]-
6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-4-yl}oxy)-1-
piperidinyl]acetamide trifluoroacetate (44). Compound 44 was
prepared as a white solid in 37% yield from 28 in a similar manner to that
described for 29 using 2-chloro-N,N-dimethylacetamide, diisopropyle-
thylamine and a catalytic amount of sodium iodide. 1H NMR (400 MHz,
CDCl3): δ 8.28 (s, 1H), 7.96 (app t, J = 8.5 Hz, 1H), 7.76−7.73 (m, 2H),
5.58−5.39 (m, 1H), 4.22 (t, J = 8.5 Hz, 2H), 4.08 (s, 2H), 3.74−3.58 (m,
2H), 3.56−3.43 (m, 2H), 3.41−3.16 (m, 6H), 3.06 (s, 3H), 2.43−2.30
(m, 2H), 2.29−2.17 (m, 2H), 1.18−1.12 (m, 6H). HRMS (ESI): m/z
calcd for C24H32FN5O4S [M + H]+ 506.2237, obsd 506.2236.
4.18 (m, 2H), 3.68 (m, 2H), 3.19−3.12 (m, 2H), 2.38 (q, J = 7.6 Hz,
2H), 1.75−1.69 (m, 2H), 1.31−1.22 (m, 2H), 1.09 (t, J = 7.2 Hz, 3H).
LRMS (ESI): m/z 207 (M + H)+. Compound 57 was prepared as a
white solid in 50% yield from 21 in a similar manner to that described for
1
3 using 1-(5-ethyl-2-pyrimidinyl)-4-piperidinol (51). H NMR (400
MHz, DMSO-d6): δ8.24 (s, 3H), 7.96 (app t, J = 8.4 Hz, 1H), 7.84 (dd, J
= 10.8, 2.0 Hz, 1H), 7.74 (dd, J = 8.4, 2.0 Hz, 1H), 5.36 (m, 1H), 4.20−
4.18 (m, 2H), 4.14 (t, J = 8.4 Hz, 2H), 3.51−3.45 (m, 2H), 3.25 (s, 3H),
3.05 (t, J = 8.8 Hz, 2H), 2.41 (q, J = 7.6 Hz, 2H), 2.01−1.97 (m, 2H),
1.65−1.48 (m, 2H), 1.11 (t, J = 7.6 Hz, 3H). HRMS (ESI): m/z calcd for
C24H27FN6O3S [M + H]+ 499.1928, obsd 499.1932.
7-[2-Fluoro-4-(methylsulfonyl)phenyl]-4-({1-[3-(1-methyl-
ethyl)-1,2,4-oxadiazol-5-yl]-4-piperidinyl}oxy)-6,7-dihydro-5H-
pyrrolo[2,3-d]pyrimidine (55). Sodium bicarbonate (41 g, 0.494
mol) was dissolved in water (20 mL) and cooled to 0 °C. A solution of 4-
hydroxypiperidine (45, 25 g, 0.25 mol) in CH2Cl2 (75 mL) was added to
the above solution with vigorous stirring. A solution of cyanogen
bromide (28 g, 0.27 mol) in CH2Cl2 (25 mL) was then added and the ice
bath was removed, and the mixture was stirred at ambient temperature
for 18 h. Solid sodium carbonate (75 g) was then added until the pH was
adjusted to ∼7, and then MgSO4 (25 g) was added to remove water. The
solids were removed via filtration and were washed several times with
CH2Cl2. The filtrate was concentrated, and the resultant 4-hydroxy-1-
piperidinecarbonitrile 46 (24.8 g) was used without purification. A
solution of 46 (12.9 g, 0.1 mol) and N-hydroxy-2-methylpropanimida-
mide (12.8 g, 0.13 mol) in a mixture of EtOAc (500 mL) and ether (100
mL) was treated with a 1N solution of ZnCl2 (120 mL, 0.12 mol) in
ether. The mixture was stirred for 15 min, and the supernatant was
decanted off. The residue was washed 2 times with 250 mL of ether,
which was decanted off after each wash. The residue was then dissolved
in a mixture of 4N HCl (50 mL) and EtOH (100 mL). The resulting
solution was refluxed for 1 h, the solvent was reduced to about 25 mL,
and 25 g of Na2CO3 was added along with 100 mL of CH2Cl2. The solids
were removed via filtration. The organic phase was dried over MgSO4,
filtered, and concentrated to give 1-[3-(1-methylethyl)-1,2,4-oxadiazol-
5-yl]-4-piperidinol 47 (16 g) as a tan oil that was used without
purification. 1H NMR (400 MHz, CDCl3): δ 4.04−3.74 (m, 3H), 3.44−
3.20 (m, 2H), 2.78 (m, 1H), 2.05−1.80 (m, 2H), 1.74−1.49 (m, 2H),
1.38−1.13 (m, 6H). Compound 55 was prepared as a white solid in 55%
yield from 21 in a similar manner to that described for 3 using 47. 1H
NMR (400 MHz, DMSO-d6): δ 8.27 (s, 1H), 7.99 (app t, J = 7.6 Hz,
7-[2-Fluoro-4-(methylsulfonyl)phenyl]-4-{[1-(1,4,5,6-tetrahy-
dro-2-pyrimidinyl)-4-piperidinyl]oxy}-6,7-dihydro-5H-pyrrolo-
[2,3-d]pyrimidine (58). 1-(2-Pyrimidinyl)-4-piperidinol (53) was
prepared in 81% yield as a white solid in a similar manner to that
described for 50 using 2-chloropyrimidine. 1H NMR (400 MHz,
CDCl3): δ 8.28 (d, J = 4.6 Hz, 2H), 6.49 (t, J = 4.8 Hz, 1H), 4.49−4.33
(m, 2H), 4.01−3.88 (m, 1H), 3.37−3.21 (m, 2H), 2.02−1.90 (m, 2H),
1.60−1.45 (m, 2H). LRMS (ESI): m/z 180 (M + H)+. A solution of 1-
(2-pyrimidinyl)-4-piperidinol (53, 300 mg, 1.67 mmol) in acetic acid
(10 mL) was treated with 10% palladium on carbon (Degussa type, 100
mg) and concentrated hydrochloric acid (0.5 mL), and the mixture was
placed under an atmosphere of hydrogen (40 psi) for 18 h at room
temperature. The reaction mixture was then filtered and the solvent was
removed to give 1-(1,4,5,6-tetrahydro-2-pyrimidinyl)-4-piperidinol 54
(304 mg, 84%). 1H NMR (400 MHz, DMSO-d6): δ 8.25 (m, 2H), 4.03−
3.41 (m, 3H), 2.95−2.86 (m, 4H), 2.09−1.55 (m, 6H), 1.55−1.18 (m,
2H). Compound 58 was prepared as a white solid in 6% yield from 21 in
a similar manner to that described for 3 using 54 and DMF as solvent. 1H
NMR (400 MHz, CD3OD): δ 8.46 (s, 1H), 8.17 (s, 1H), 7.95 (app t, J =
8.5 Hz, 1H), 7.88−7.66 (m, 2H), 5.54−5.37 (m, 1H), 4.20 (t, J = 8.5 Hz,
2H), 3.75−3.57 (m, 2H), 3.49−3.33 (m, 6H), 3.19−3.07 (m, 5H),
2.21−2.06 (m, 2H), 2.01−1.76 (m, 4H). HRMS (ESI): m/z calcd for
C22H27FN6O3S [M + H]+ 475.1928, obsd 475.1927.
1,1-Dimethylethyl 4-({7-[2-Fluoro-4-(methylsulfonyl)-
phenyl]-2-methyl-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-4-
yl}oxy)-1-piperidinecarboxylate (60). Compound 60 was prepared
as a white solid in 15% overall yield from ethanimidamide hydrochloride
and 16 in addition to 2-fluoro-4-(methylsulfonyl)aniline and 1,1-
dimethylethyl 4-hydroxy-1-piperidinecarboxylate in a similar manner to
that described for 3. 1H NMR (400 MHz, CDCl3): δ 8.13 (m, 1H), 7.69
P
dx.doi.org/10.1021/jm301404a | J. Med. Chem. XXXX, XXX, XXX−XXX

Journal of Medicinal Chemistry
Article
(m, 2H), 5.38 (m, 1H), 4.25 (t, J = 7.3 Hz, 2H), 3.87 (m, 2H), 3.32 (m,
2H), 3.04 (m, 5H), 1.97 (m, 2H), 1.73 (m, 2H), 1.46 (s, 9H). HRMS
(ESI): m/z calcd for C24H31FN4O5S [M + H]+ 507.2077, obsd
507.2078.
oxy}-1- piperidinecarboxylate (74). Compound 74 was prepared as
an off-white foam in 15% overall yield from N-methylguanidine
hydrochloride (68) and 66 in a similar manner to that described for
73. 1H NMR (400 MHz, CDCl3): δ 8.15−8.05 (br s, 1H), 7.70−7.65
(m, 2H), 5.30 (br s, 1H), 4.90 (septuplet, J = 6.0 Hz, 1H), 4.85−4.60 (m,
1H), 4.15 (br t, J = 7.3 Hz, 2H), 3.85−3.70 (m, 2H), 3.40−3.30 (m, 2H),
3.05 (s, 3H), 2.95 (t, J = 8.5 Hz, 2H), 2.92 (d, J = 4.9 Hz, 3H), 2.00−1.95
(m, 2H), 1.85−1.80 (m, 2H), 1.24 (d, J = 6.0 Hz, 6H). HRMS (ESI): m/
z calcd for C23H30FN5O5S [M + H]+ 508.2030, obsd 508.2030.
1-Methylethyl 4-({7-[2-Fluoro-4-(methylsulfonyl)phenyl]-2-
methyl-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-4-yl}oxy)-1-pi-
peridinecarboxylate (61). Compound 61 was prepared as a white
solid in 98% overall yield from 56 in a similar manner to that described
1
for the Boc deprotection and isopropylcarbamate formation of 2. H
NMR (400 MHz, CDCl3): δ 8.15 (m, 2H), 7.69 (m, 2H), 5.38 (m, 1H),
4.93 (m, 1H), 4.19 (t, J = 7.3 Hz, 2H), 3.75 (m, 2H), 3.35 (m, 2H) 3.05
(m, 5H), 2.47 (s, 3H), 1.95 (m, 2H), 1.71 (m, 2H), 1.17 (d, J = 6.8 Hz,
6H). HRMS (ESI): m/z calcd for C23H29FN4O5S [M + H]+ 493.1921,
obsd 493.1920.
1-Methylethyl 4-({7-[2-Fluoro-4-(methylsulfonyl)phenyl]-
6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-4-yl}amino)-1-piperi-
dinecarboxylate trifluoroacetate (77). A solution of 1,1-dimethy-
lethyl 4-piperidinylcarbamate (512 mg, 2.6 mmol) in CH2Cl2 (25 mL)
was treated with Et3N (0.71 mL, 5.1 mmol) followed by the dropwise
addition of isopropylchloroformate (1.0M in toluene, 2.8 mL, 2.8
mmol) via addition funnel at RT. The reaction mixture was stirred at RT
for 17 h, then washed with water, dried over MgSO4, filtered, and
concentrated to afford 1-methylethyl 4-({[(1,1-dimethylethyl)oxy]-
carbonyl}amino)-1-piperidinecarboxylate 75 (585 mg, 80%) as a
1-Methylethyl 4-({2-Amino-7-[2-fluoro-4-(methylsulfonyl)-
phenyl]-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-4-yl}oxy)-1-
piperidinecarboxylate (73). A solution of 2-fluoro-4-
(methylsulfonyl)aniline (19, 3.3 g, 17.3 mmol) in acetonitrile (10
mL) under N2 was treated with 6,6-dimethyl-5,6-dioxaspiro[2.5]octane
(63, 9.7 g, 57 mmol) and stirred and heated at 60 °C for 3 h. The
reaction mixture was allowed to cool to RT and was concentrated. The
residue was heated at 70 °C for 5 h to afford 64. Methanol (15 mL) and
concentrated H2SO4 (0.2 mL) were added, and the mixture was heated
to reflux for 2 h. The mixture was concentrated, and the residue was
purified by flash chromatography to give methyl 1-[2-fluoro-4-
(methylsulfonyl)phenyl]-2-oxo-3-pyrrolidinecarboxylate 65 (2.3 g,
1
colorless oil. H NMR (400 MHz, CDCl3): δ 4.85 (septuplet, J = 6.4
Hz, 1H), 4.63−4.48 (m, 1H), 4.13−3.89 (m, 2H), 3.55 (br s, 1H), 2.82
(t, J = 11.7 Hz, 2H), 1.88−1.85 (m, 2H), 1.39 (s, 9H), 1.29−1.22 (m,
2H), 1.19 (d, J = 6.1 Hz, 6H). A solution of 21 (50 mg, 0.15 mmol) in
DMF (1.5 mL) was treated with 75 (87 mg, 0.31 mmol) and K2CO3 (42
mg, 0.31 mmol), and the reaction mixture was sealed and heated in a
microwave for 20 min at 200 °C. The reaction mixture was then heated
in a microwave an additional 60 min at 220 °C. The reaction mixture was
then diluted with water and extracted with EtOAc. The combined
organic layer was dried over MgSO4, filtered, and concentrated. The
crude product was purified by reverse phase HPLC to give 77 (11 mg,
1
43%) as an off-white solid. H NMR (400 MHz, DMSO-d6): δ 7.89
(br d, J = 10.0 Hz, 1H), 7.81−7.73 (m, 2H), 3.85 (br t, J = 7.2 Hz, 2H),
3.75 (t, J = 8.7 Hz, 1H), 3.68 (s, 3H), 3.30−3.25 (br s, 3H), 2.45−2.35
(m, 2H). LRMS (ESI): m/z 316 (M + H)+. A solution of 65 (2.3 g, 7.2
mmol) in THF (100 mL) under N2 was treated with P2S5 (3.9 g, 8.7
mmol) and was heated at 70 °C for 12 h. The mixture was filtered and
the solid washed twice with EtOAc. The filtrate was concentrated and
the residue was purified by flash chromatography to afford methyl 1-[2-
fluoro-4-(methylsulfonyl)phenyl]-2-thioxo-3-pyrrolidinecarboxylate 66
(1.2 g, 50%) as a pale-yellow solid. 1H NMR (400 MHz, DMSO-d6): δ
7.98 (br d, J = 9.3 Hz, 1H), 7.89 (br d, J = 8.3 Hz, 1H), 7.81 (app t, J = 7.5
Hz, 1H), 4.12−4.08 (m, 3H), 3.70 (s, 3H), 3.30 (s, 3H), 2.60−2.50 (m,
2H). LRMS (ESI): m/z 332 [M + H]+. A solution of guanidine
hydrochloride (67, 0.3 g, 3.2 mmol) in anhydrous MeOH (3 mL) was
treated with freshly prepared 1M NaOMe/MeOH (3.2 mL, 3.2 mmol),
and the mixture was stirred for 30 min at ambient temperature under N2.
The mixture was filtered and the filtrate added to 62 (0.22 g, 0.64
mmol). The yellow mixture was stirred for 5 min then concentrated and
heated stepwise from 60 to 90 °C with a slight vacuum for 1 h. The
mixture was allowed to cool to ambient temperature, and H2O was
added. The pH was adjusted to 6 with 1 M HCl/H2O, and the mixture
was stirred at ambient temperature for 18 h. The solid was filtered and
dried in vacuum for 2 h to provide 2-amino-7-[2-fluoro-4-
(methylsulfonyl)phenyl]-1,5,6,7-tetrahydro-4H-pyrrolo[2,3-d]-
pyrimidin-4-one 69 (0.12 g) as an yellow solid. LRMS (ESI): m/z 325
[M + H]+. A suspension of the crude 69 (0.12 g, 0.36 mmol) in POCl3
(1.2 mL) was carefully treated with (i-Pr)2NEt (0.13 mL, 0.1 g, 0.72
mmol), and the mixture was slowly heated to 70 °C. After 1 h, the
mixture was concentrated and the residue was partitioned between
EtOAc/CH2Cl2 and saturated NaHCO3/H2O. The organic phase was
filtered, dried over Na2SO4, and concentrated to afford 4-chloro-7-[2-
fluoro-4-(methylsulfonyl)phenyl]-6,7-dihydro-5H-pyrrolo[2,3-d]-
pyrimidin-2-amine 71 (51 mg) as a dark-green solid. LRMS (ESI): m/z
343 [M + H]+. Compound 73 was prepared as an off-white solid in 7%
yield from 71 in a similar manner to that described for 3 using 1-
1
12%) as a yellow solid. H NMR (400 MHz, CDCl3): δ 8.09 (s, 1H),
7.81−7.77 (m, 2H), 7.72−7.69 (m, 1H), 4.91 (septuplet, J = 6.4 Hz,
1H), 4.29 (t, J = 9.0 Hz, 2H), 4.23−4.12 (m, 2H), 3.79−3.68 (m, 1H),
3.43−3.38 (m, 2H), 3.09 (s, 3H), 2.96−2.86 (m, 2H), 1.97−1.93 (m,
2H), 1.77−1.67 (m, 2H), 1.25 (d, J = 6.4 Hz, 6H). HRMS (ESI): m/z
calcd for C22H28FN5O4S [M + H]+ 478.1924, obsd 478.1924.
1-Methylethyl 4-[{7-[2-Fluoro-4-(methylsulfonyl)phenyl]-
6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-4-yl}(methyl)amino]-
1-piperidinecarboxylate trifluoroacetate (78). Compound 76 was
prepared as a colorless oil in 87% yield from from 1,1-dimethylethyl
methyl(4-piperidinyl)carbamate in a similar manner to that described
for 75. 1H NMR (400 MHz, CDCl3): δ 4.88 (septuplet, J = 6.4 Hz, 1H),
4.28−4.16 (m, 3H), 2.77−2.71 (m, 2H), 2.68 (s, 3H), 1.62−1.57 (m,
2H), 1.55−1.50 (m, 2H), 1.44 (s, 9H), 1.21 (d, J = 6.4 Hz, 6H).
Compound 78 was prepared as a yellow solid in 9% yield from 21 and 75
in a similar manner to that described for 77. HRMS (ESI): m/z calcd for
C23H30FN5O4S [M + H]+ 492.2081, obsd 492.2083.
1-Methylethyl 4-({7-[4-(Methylsulfonyl)phenyl]-6,7-dihydro-
5H-pyrrolo[2,3-d]pyrimidin-4-yl}thio)-1-piperidinecarboxylate
(80). A solution of 4-bromopiperidine hydrochloride (5.0 g, 20.4 mmol)
in dichloromethane (100 mL) under N2 at 0 °C was treated with
diisopropylethylamine (7.8 mL, 44.9 mmol) followed by the dropwise
addition of a 1.0 M solution of isopropyl chloroformate in toluene (20.4
mL, 20.4 mmol). The mixture was stirred and allowed to warm to 25 °C
over a 3 h period. The reaction was quenched by the addition of 1N HCl
and diluted with EtOAc. The aqueous layer was extracted with EtOAc,
and the combined organic extracts were washed with saturated aqueous
NaHCO3 and brine, dried (MgSO4), and concentrated to give 1-
methylethyl 4-bromo-1-piperidinecarboxylate as a colorless oil that was
used without further purification. 1H NMR (400 MHz, CDCl3): δ4.91
(septuplet, J = 6.2 Hz, 1H), 4.35 (m, J = 3.8 Hz, 1H), 3.71 (m, 2H), 3.36
(m, 2H), 2.09 (m, 2H), 1.93 (m, 2H), 1.24 (d, J = 6.2 Hz, 6H). A
solution of 1-methylethyl 4-bromo-1-piperidinecarboxylate (4.0 g, 16
mmol) in DMF (40 mL) under N2 at 25 °C was treated with potassium
thioacetate (3.4 g, 29.8 mmol), and the mixture was stirred and heated at
100 °C for 3.5 h. The reaction mixture was allowed to cool to 25 °C and
was quenched by the addition of water and diluted with EtOAc. The
aqueous layer was extracted with EtOAc, and the combined organic
extracts were washed with water, saturated aqueous NaHCO3, and brine,
dried (Na2SO4), and concentrated to dark-brown oil. Purification by
1
methylethyl 4-hydroxy-1-piperidinecarboxylate. H NMR (400 MHz,
CDCl3): δ 8.10−8.00 (br s, 1H), 7.70−7.65 (m, 2H), 5.30 (br s, 1H),
4.90 (septuplet, J = 6.0 Hz, 1H), 4.85−4.60 (m, 2H), 4.15 (br t, J = 7.7
Hz, 2H), 3.85−3.70 (m, 2H), 3.40−3.30 (m, 2H), 3.05 (s, 3H), 2.95 (t, J
= 8.5 Hz, 2H), 2.00−1.95 (m, 2H), 1.85−1.80 (m, 2H), 1.24 (d, J = 6.0
Hz, 6H). HRMS (ESI): m/z calcd for C22H28FN5O5S [M + H]+
494.1873, obsd 494.1872.
1-Methylethyl 4-{[7-[2-Fluoro-4-(methylsulfonyl)phenyl]-2-
(methylamino)-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-4-yl]-
Q
dx.doi.org/10.1021/jm301404a | J. Med. Chem. XXXX, XXX, XXX−XXX

Journal of Medicinal Chemistry
Article
flash chromatography (80 g silica column, 10→20% EtOAc/hexane)
afforded 1-methylethyl 4-(acetylthio)-1-piperidinecarboxylate (2.3 g,
59%) as a brown oil. 1H NMR (400 MHz, CDCl3): δ 4.91 (septuplet, J =
6.2 Hz, 1H), 4.35 (m, J = 3.8 Hz, 1H), 3.71 (m, 2H), 3.36 (m, 2H), 2.09
(m, 2H), 1.93 (m, 2H), 1.24 (d, J = 6.2 Hz, 6H). A solution of 1-
methylethyl 4-(acetylthio)-1-piperidinecarboxylate (2.3 g, 9.4 mmol) in
THF:water (1:1, 50 mL) under N2 at 0 °C was treated with 1N NaOH
(11.2 mL, 11.2 mmol), and the mixture was stirred at 0 °C for 2 h. The
reaction mixture was quenched by the addition of aqueous citric acid,
and the aqueous layer was extracted with EtOAc. The combined organic
extracts were washed with saturated aqueous NaHCO3 and brine and
were dried (MgSO4) and concentrated to give a brown oil. Purification
by flash chromatography (40 g silica column, 10→25% EtOAc/hexane)
afforded 1-methylethyl 4-mercapto-1-piperidinecarboxylate 79 (1.6 g,
89%) as an amber oil. 1H NMR (400 MHz, DMSO-d6): δ 4.72
(septuplet, J = 6.4 Hz, 1H), 3.80 (overlapping br s, 2H), 2.96−2.79 (br
m, 3H), 2.64 (d, J = 7.1 Hz, 1H), 1.90−1.82 (m, 2H), 1.38−1.27 (m,
2H), 1.15 (d, J = 6.4 Hz, 6H). A solution of 79 (82 mg, 0.43 mmol) in
acetone (2 mL) was treated with 21 under N2 at 25 °C followed by
K2CO3 (89 mg, 0.64 mmol), and the mixture was stirred and heated in a
sealed vial at 60 °C for 62 h. The reaction mixture was allowed to cool to
25 °C and was concentrated to ∼1/3 volume and purified by flash
chromatography (12 g silica column, 10→60% EtOAc/hexane, 35 min
gradient) to afford 80 (150 mg, 71%) as a white solid. 1H NMR (400
MHz, DMSO-d6): δ 8.35 (s, 1H), 7.96 (t, J = 8.1 Hz, 1H), 7.85 (d, J =
11.2 Hz, 1H), 7.75 (d, J = 8.5 Hz, 1H), 4.74 (septuplet, J = 6.2 Hz, 1H),
4.15 (app t, J = 8.5 Hz, 2H), 4.11 (obscured m, 1H), 3.82 (br d, J = 12.7
Hz, 2H), 3.25 (s, 3H), 3.07 (br m, 2H), 2.99 (app t, J = 8.5 Hz, 2H), 2.01
(br d, J = 12.7 Hz, 2H), 1.52 (m, 2H), 1.16 (d, J = 6.2 Hz, 6H). HRMS
(ESI): m/z calcd for C22H27FN4O4S2 [M + H]+ 495.1536, obsd
495.1537.
1-Methylethyl 4-({7-[2-Fluoro-4-(methylsulfonyl)phenyl]-
6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-4-yl}sulfonyl)-1-piper-
idinecarboxylate (81). A solution of 80 (145 mg, 0.29 mmol) in
CH2Cl2 (5 mL) was treated with m-CPBA (77% maximum by weight,
105 mg, 0.47 mmol) under N2 at 0 °C, and the mixture was stirred at 0
°C for 1.5 h. The reaction mixture was quenched by the addition of
methanol, concentrated to ∼1/3 volume, and purified by reverse phase
HPLC (C18 column, 10→100% CH3CN/H2O + 0.5% trifluoroacetic
acid, 10 min gradient) to give 81 (28 mg, 18%) as a white solid after
lyophilization. 1H NMR (400 MHz, DMSO-d6): δ 8.57 (s, 1H), 8.02−
7.97 (overlapping m, 2H), 7.88 (d, J = 8.2 Hz, 1H), 4.79 (septuplet, J =
6.0 Hz, 1H), 4.26 (app t, J = 8.0 Hz, 2H), 4.08 (br m, 2H), 3.84 (br m,
2H), 3.55 (app t, J = 8.0 Hz, 2H), 3.34 (s, 3H), 2.89 (br m, 2H), 1.95 (br
d, J = 11.4 Hz, 2H), 1.54 (m, 2H), 1.20 (d, J = 6.0 Hz, 6H). HRMS
(ESI): m/z calcd for C22H27FN4O6S2 [M + H]+ 527.1432, obsd
527.1435.
6.2 Hz, 3H), 1.25 (s, 9H). LRMS (ESI): m/z 460 (M + H)+. A solution
of 83 (0.8 g, 1.7 mmol) in CH2Cl2 (20 mL) was treated with CF3CO2H
(5 mL) at RT under N2. The reaction mixture was stirred at RT for 3 h
and then concentrated to afford ( )-1-(4-chloro-6-{[2-fluoro-4-
(methylsulfonyl)phenyl]amino}-5-pyrimidinyl)-2-propanol 84, which
was used without further purification. 1H NMR (400 MHz, DMSO-d6):
δ 9.84 (s, 1H), 8.42 (s, 1H), 8.16 (app, t, J = 8.0 Hz, 1H), 7.82 (dd, J =
10.4, 2.0 Hz, 1H), 7.73 (dd, J = 8.8, 1.6 Hz, 1H), 5.87 (d, J = 2.8 Hz, 1H),
4.07 (app br s, 1H), 3.24 (s, 3H), 2.94−2.83 (m, 2H), 1.20 (d, J = 6.4 Hz,
3H). LRMS (ESI): m/z 360 (M + H)+. A solution of 84 (1.7 mmol) in
CH2Cl2 (30 mL) was added Et3N (0.72 mL, 5.2 mmol) followed by
methanesulfonic anhydride (0.61 g, 3.5 mmol) at RT for 24 h. The
reaction was diluted with CH2Cl2 and quenched by the addition of
water. The organic layer was washed with brine, dried over Na2SO4, and
concentrated. Purification by flash chromatography afforded ( )-4-
chloro-7-[2-fluoro-4-(methylsulfonyl)phenyl]-6-methyl-6,7-dihydro-
5H-pyrrolo[2,3-d]pyrimidine 85 (0.27 g, 47% overall for two steps) as
an off-white solid. 1H NMR (400 MHz, DMSO-d6): δ 8.23 (s, 1H), 7.93
(app dd, J = 10.8, 1.2 Hz, 1H), 7.87−7.81 (m, 2H), 4.80−4.72 (m, 1H),
3.45 (dd, J = 17.2, 9.6 Hz, 1H), 3.30 (s, 3H), 2.87 (dd, J = 17.2, 7.2 Hz,
1H), 1.21 (d, J = 6.0 Hz, 3H). LRMS (ESI): m/z 342 (M + H)+. A
racemic mixture of 86 and 87 was prepared as a white solid in 82% yield
from 85 and 1-methylethyl 4-hydroxy-1-piperidinecarboxylate in a
similar manner to that described for 3. 1H NMR (400 MHz, DMSO-d6):
δ 8.16 (s, 1H), 7.87 (app br d, J = 10.4 Hz, 1H), 7.82 (app br d, J = 7.6
Hz, 1H), 7.77 (dd, J = 8.4, 1.6 Hz, 1H), 5.30−5.22 (m, 1H), 4.78−4.66
(m, 2H), 3.68−3.64 (m, 2H), 3.32−3.28 (m, 1H), 3.28 (s, 3H), 3.28−
3.22 (m, 2H), 2.68 (app dd, J = 16.0, 7.2 Hz, 1H), 1.96−186 (m, 2H),
1.62−1.50 (m, 2H), 1.19 (app d, J = 5.6 Hz, 3H), 1.17 (d, J = 6.0 Hz,
6H). LRMS (ESI): m/z 493 (M + H)+. The racemic mixture (200 mg)
was subjected to Chiral HPLC [column: Chiralpak OJ (analytical),
Chiralpak OJ (prep); mobile phase, 95% CO2, 5% MeOH:CHCl3
(90:1); 2 mL/min; pressure 140 bar, temperature 30 °C, 215 nm, and
280 nm) ] analysis and then separated into two (+ and −) enantiomers.
Enantiomer 1, (+)-86: Retention time = 19.96 min, 46 mg (%ee >98%).
HRMS (ESI): m/z calcd for C23H29FN4O5S [M + H]+ 493.1921, obsd
493.1920. Specific rotation in 50:50 DCM:methanol vol/vol, 25 °C, 589
nm = +119.17°. Enantiomer 2, (−)-87: Retention time = 21.0 min, 66
mg (%ee >98%). HRMS (ESI): m/z calcd for C23H29FN4O5S [M + H]+
493.1921, obsd 493.1921. Specific rotation in 50:50 DCM:methanol
vol/vol, 25 °C, 589 nm = −97.06°.
1-Methylethyl 4-({7-[5-(Methylsulfonyl)-2-pyridinyl]-6,7-di-
hydro-5H-pyrrolo[2,3-d]pyrimidin-4-yl}oxy)-1-piperidinecar-
boxylate Hydrochloride (97). A solution of 2-amino-5-iodopyridine
(11.6 g, 52.6 mmol) in MeOH (250 mL) was treated with sodium
thiomethoxide (5.16 g, 73.6 mmol) and copper powder (1.07 g, 16.8
mmol) at RT. The reaction mixture was heated in a sealed tube at 120 °C
for 68 h. The reaction mixture was then cooled to RT and filtered
through a pad of Celite. The pad was rinsed with MeOH, and the
combined organics were concentrated and redissolved in EtOAc. The
organic layer was washed with water, and the aqueous layer was
extracted with EtOAc. The combined organic layer was dried over
MgSO4, filtered, and concentrated to afford 5-(methylthio)-2-pyridin-
(
)-1-Methylethyl 4-({7-[2-Fluoro-4-(methylsulfonyl)-
phenyl]-6-methyl-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-4-
yl}oxy)-1-piperidinecarboxylate (86 and 87). A solution of allyl 19
(3.42 g, 10 mmol) in 3:2 acetone/H2O (250 mL) was treated with
K2OsO4·2H2O (147 mg, 0.4 mmol). Solid NaIO4 (8.56 g, 40 mmol) was
added portionwise, and the reaction was stirred for 4 h. The workup gave
1,1-dimethylethyl [6-chloro-5-(2-oxoethyl)-4-pyrimidinyl][2-fluoro-4-
(methylsulfonyl)phenyl]carbamate (82, 5.0 g, 100%) as an off-white
solid. 1H NMR (400 MHz, DMSO-d6): δ 9.63 (s, 1H), 8.82 (s, 1H), 7.95
(dd, J = 9.6, 2.0 Hz, 1H), 7.77 (dd, J = 8.4, 1.6 Hz, 1H), 7.59 (app t, J =
7.6 Hz, 1H), 3.93 (s, 2H), 3.30 (s, 3H), 1.38 (s, 9H). A solution of 82
(2.0 g, 4.51 mmol) in anhydrous THF (50 mL) was treated with a 1.4 M
solution of methylmagnesium bromide in THF (6.5 mL, 9.1 mmol) at
RT under N2. The resultant mixture was stirred at RT for 3 h. The
reaction mixture was poured onto 10% aqueous HCl (200 mL), and the
mixture was extracted with EtOAc. The combined organic layer was
washed with brine, dried over Na2SO4, and then concentrated to afford
the crude product, which was purified by flash chromatography to afford
( )-1,1-dimethylethyl [6-chloro-5-(2-hydroxypropyl)-4-pyrimidinyl]-
[2-fluoro-4-(methylsulfonyl)phenyl]carbamate (83, 0.82 g, 39%) as a
1
amine 89 (6.23 g, 84%) as an off-white solid. H NMR (400 MHz,
CDCl3): δ 8.09 (s, 1H), 7.51−7.48 (m, 1H), 6.48−6.45 (m, 1H), 4.35
(br s, 2H), 2.38 (s, 3H). A solution of 17 (18.9 g, 0.1 mol) in acetone/
water (1:1, 1 L) was treated with K2OsO4·2H2O (1.47 g, 4 mmol)
followed by the portiowise addition of NaIO4 (64.2 g, 0.4 mol). The
reaction mixture was stirred at RT for 4 h and then filtered through a
plug of silica (CH2Cl2 wash). The filtrate was extracted with CH2Cl2,
and the extracts were concentrated. Purification by flash chromatog-
raphy afforded (4,6-dichloro-5-pyrimidinyl)acetaldehyde 88 (10 g,
45%). A solution of 88 (3.4 g, 17.8 mmol) in MeOH (180 mL) was
treated with 89 (3.0 g, 21.4 mmol) at RT. The reaction mixture was
cooled to −15 °C with an ice/methanol bath and glacial acetic acid (3.1
mL, 53.4 mmol) and NaBH3CN (3.4 g, 53.4 mmol) were added. The
reaction mixture was stirred at −15 °C for 15 min and then allowed to
warm to RT and stir for 19 h. The reaction mixture was then diluted with
water (50 mL) and extracted with CH2Cl2. The combined organic layer
was dried over MgSO4, filtered, and concentrated to afford N-[2-(4,6-
1
white solid. H NMR (400 MHz, DMSO-d6): δ 9.15 (s, 1H), 8.28 (s,
1H), 7.84 (dd, J = 9.6, 2.0 Hz, 1H), 7.80−7.68 (m, 2H), 5.06−4.99 (m,
1H), 3.28 (s, 3H), 3.14−3.10 (m, 1H), 3.05−3.02 (m, 1H), 1.33 (d, J =
R
dx.doi.org/10.1021/jm301404a | J. Med. Chem. XXXX, XXX, XXX−XXX

Journal of Medicinal Chemistry
Article
dichloro-5-pyrimidinyl)ethyl]-5-(methylthio)-2-pyridinamine 91 as a
yellow oil. The crude oil was dissolved in THF (500 mL) and then
treated with t-BuOK (5.99 g, 53.4 mmol) at RT. The reaction mixture
immediately changed to a brown color and was stirred at RT for 22 h.
The reaction mixture was quenched with water (50 mL), concentrated,
and then redissolved in EtOAc (200 mL). The organic layer was washed
with water, and the aqueous layer was extracted with EtOAc. The
combined organic layer was dried over MgSO4, filtered, and
concentrated to a brown oil. The crude oil was purified by flash
chromatography (20→40% EtOAc/hexanes; monitoring at 319 nm) to
afford 4-chloro-7-[5-(methylthio)-2-pyridinyl]-6,7-dihydro-5H-
pyrrolo[2,3-d]pyrimidine 93 (550 mg, 11%) as a colorless oil. 1H
NMR (400 MHz, CDCl3): δ 8.57 (d, J = 9.0 Hz, 1H), 8.43 (s, 1H), 8.29
(d, J = 2.4 Hz, 1H), 7.67 (dd, J = 8.8, 2.4 Hz, 1H), 4.37−4.33 (m, 2H),
3.16 (t, J = 8.8 Hz, 2H), 2.47 (s, 3H). 1-Methylethyl 4-({7-[5-
(methylthio)-2-pyridinyl]-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-4-
yl}oxy)-1-piperidinecarboxylate (95) was prepared as a off-white solid
in 88% yield from 93 and 1-methylethyl 4-hydroxypiperidinecarboxylate
(100). Compound 100 was prepared as a white solid in 15% overall yield
from 88 and 2-amino-5-fluoropyridine in a similar manner to that
described for 99. 1H NMR (400 MHz, CDCl3): δ 8.66 (dd, J = 9.3, 4.0
Hz, 1H), 8.35 (s, 1H), 8.17 (d, J = 3.11 Hz, 1H), 7.47−7.40 (m, 1H),
5.41−5.30 (m, 1H), 5.00−4.89 (m, 1H), 4.30 (t, J = 8.6 Hz, 2H), 3.89−
3.73 (m, 2H), 3.42−3.30 (m, 2H), 3.04 (t, J = 8.6 Hz, 2H), 2.07−1.93
(m, 2H), 1.84−1.67 (m, 2H), 1.26 (d, J = 6.2 Hz, 6H). HRMS (ESI): m/
z calcd for C20H24FN5O3 [M + H]+ 402.1942, obsd 402.1942.
1-Methylethyl 4-{[7-(2-Methyl-3-pyridinyl)-6,7-dihydro-5H-
pyrrolo[2,3-d]pyrimidin-4-yl]oxy}-1-piperidinecarboxylate
(101). N-[2-(4,6-Dichloro-5-pyrimidinyl)ethyl]-2-methyl-3-pyridin-
amine was prepared from 88 and 2-methyl-3-pyridinamine in a similar
1
manner to that described for 91. H NMR (400 MHz, DMSO-d6): δ
8.81 (s, 1H), 7.95−7.94 (m, 1H), 7.71−7.66 (m, 2H), 6.63 (app t, J = 5.6
Hz, 1H), 3.59 (app q, J = 6.0 Hz, 2H), 3.08 (t, J = 6.4 Hz, 2H), 2.40 (s,
3H), LRMS (ESI): m/z 285 (M + H)+. A solution of N-[2-(4,6-
dichloro-5-pyrimidinyl)ethyl]-2-methyl-3-pyridinamine (∼11.0 g) in
toluene (200 mL) was treated with PdCl2(Ph3P)2 (1.36 g, 2 mmol),
potassium tert-butoxide (8.7 g, 77.7 mmol), and K2CO3 (10.7 g, 77.7
mmol) at RT under N2. The reaction mixture was stirred and refluxed
for 18 h. The reaction mixture was allowed to cool to RT and then
filtered through a pad of Celite. The filtrate was concentrated and
purified by flash chromatography to afford 4-chloro-7-(2-methyl-3-
pyridinyl)-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidine (0.85 g). 1H NMR
(400 MHz, DMSO-d6): δ 8.41 (dd, J = 4.8, 1.2 Hz, 1H), 8.11 (s, 1H),
7.75 (dd, J = 8.0, 1.6 Hz, 1H), 7.31 (dd, J = 8.0, 4.8 Hz, 1H), 4.03 (t, J =
8.8 Hz, 2H), 3.20 (t, J = 8.4 Hz, 2H), 2.34 (s, 3H), LRMS (ESI): m/z
247 (M + H)+. Compound 101 was prepared as a white solid in 55%
overall yield from 4-chloro-7-(2-methyl-3-pyridinyl)-6,7-dihydro-5H-
pyrrolo[2,3-d]pyrimidine and 1-methylethyl 4-hydroxypiperidinecar-
boxylate in a similar manner to that described for 3. 1H NMR (400 MHz,
DMSO-d6): δ 8.36 (dd, J = 4.8, 1.2 Hz, 1H), 8.05 (s, 1H), 7.69 (dd, J =
8.0, 1.6 Hz, 1H), 7.26 (dd, J = 8.0, 4.8 Hz, 1H), 5.27−5.20 (m, 1H), 4.27
(septuplet, J = 6.4 Hz, 1H), 3.94 (t, J = 8.8 Hz, 2H), 3.71−3.66 (m, 2H),
3.21 (app br t, J = 10.0 Hz, 2H), 3.04 (t, J = 9.2 Hz, 2H), 2.35 (s, 3H),
1.95−1.89 (m, 2H), 1.60−1.50 (m, 2H), 1.17 (d, J = 6.0 Hz, 6H).
HRMS (ESI): m/z calcd for C21H27N5O3 [M + H]+ 398.2191, obsd
398.2191.
1
in a similar manner to that described for 3. H NMR (400 MHz,
CDCl3): δ 8.73−8.66 (m, 1H), 8.42−8.39 (m, 1H), 8.30 (d, J = 2.4 Hz,
1H), 8.77−8.70 (m, 1H), 5.39−5.32 (m, 1H), 4.92 (septuplet, J = 6.1
Hz, 1H), 4.46−4.36 (m, 2H), 3.85−3.76 (m, 2H), 3.36−3.30 (m, 2H),
3.12−3.05 (m, 2H), 2.48 (s, 3H), 2.03−1.95 (m, 2H), 1.79−1.70 (m,
2H), 1.25 (d, J = 6.4 Hz, 6H). LRMS (ESI): m/z 430 (M + H)+. A
solution of 95 (186 mg, 0.43 mmol) in MeOH (4 mL) and acetone (3
mL) was treated with a solution of Oxone (798 mg, 1.3 mmol) in water
(4 mL) via pipet at RT. A white solid precipitated immediately, and the
reaction mixture was stirred at RT for 1 h. The reaction mixture was
quenched with saturated aqueous Na2SO3 and extracted with CH2Cl2.
The combined organic layer was dried over MgSO4, filtered, and
concentrated. The resulting solid was dissolved in a mixture of Et2O (8
mL) and acetone (3 mL) warmed with a heat gun and was treated with
HCl (1.0M in Et2O, 0.88 mL), during which time a white solid
precipitated. The solid was filtered, washed with Et2O, and dried under
high vacuum to afford 97 (190 mg, 81%) as a white solid. 1H NMR (400
MHz, CDCl3): δ 9.01 (d, J = 2.4 Hz, 1H), 8.83 (s, 1H), 8.18 (dd, J = 8.8,
2.2 Hz, 1H), 7.70 (d, J = 8.8 Hz, 1H), 5.56 (septuplet, J = 4.2 Hz, 1H),
4.93 (app quintuplet, J = 6.4 Hz, 1H), 4.57 (t, J = 8.8 Hz, 2H), 3.89−3.83
(m, 2H), 3.38−3.29 (m, 4H), 3.05 (s, 3H), 2.10−2.04 (m, 2H), 1.84−
1.76 (m, 2H), 1.26 (d, J = 6.4 Hz, 6H). HRMS (ESI): m/z calcd for
C21H27FN5O5S [M + H]+ 462.1811, obsd 462.1810.
1-Methylethyl 4-({7-[6-(Methylsulfonyl)-3-pyridinyl]-6,7-di-
hydro-5H-pyrrolo[2,3-d]pyrimidin-4-yl}oxy)-1-piperidinecar-
boxylate Hydrochloride (98). 6-(Methylthio)-3-pyridinamine 90
was prepared as a brown solid in 26% yield from 5-amino-2-
bromopyridine in a similar manner to that described for 89. 1H NMR
(400 MHz, CDCl3): δ 8.00 (d, J = 2.9 Hz, 1H), 7.06−7.00 (m, 1H),
6.92−6.89 (m, 1H), 3.50 (br s, 2H), 2.51 (s, 3H). Compound 98 was
prepared as a white solid in 7% overall yield from 88 and 90 in a similar
manner to that described for 97. 1H NMR (400 MHz, CDCl3): δ 9.02
(d, J = 2.4 Hz, 1H), 8.57 (dd, J = 8.8, 2.7 Hz, 1H), 8.37 (s, 1H), 8.04 (d, J
= 8.8 Hz, 1H), 5.36 (septuplet, J = 3.9 Hz, 1H), 4.91 (app quintuplet, J =
6.4 Hz, 1H), 4.14 (t, J = 8.8 Hz, 2H), 3.84−3.76 (m, 2H), 3.35−3.28 (m,
2H), 3.18 (s, 3H), 3.15 (t, J = 8.8 Hz, 2H), 2.02−1.95 (m, 2H), 1.77−
1.68 (m, 2H), 1.24 (d, J = 6.4 Hz, 6H). HRMS (ESI): m/z calcd for
C21H27FN5O5S [M + H]+ 462.1811, obsd 462.1811.
1-Methylethyl 4-{[7-(4-Cyano-3-fluorophenyl)-6,7-dihydro-
5H-pyrrolo[2,3-d]pyrimidin-4-yl]oxy}-1-piperidinecarboxylate
(103). 1-Methylethyl 4-{[7-(4-bromo-3-fluorophenyl)-6,7-dihydro-5H-
pyrrolo[2,3-d]pyrimidin-4-yl]oxy}-1-piperidinecarboxylate (102) was
prepared as a white solid in 38% overall yield from 88 and 4-bromo-3-
fluoroaniline in a similar manner to that described for 97. 1H NMR (400
MHz, CDCl3): δ 8.33 (s, 1H), 7.84 (dd, J = 12.0, 2.4 Hz, 1H), 7.47 (t, J =
8.0 Hz, 1H), 7.34 (dd, J = 8.8, 2.8 Hz, 1H), 5.37−5.32 (m, 1H), 4.93
(septuplet, J = 6.0 Hz, 1H), 4.04 (t, J = 8.8 Hz, 2H), 3.82−3.76 (br m,
2H), 3.36−3.30 (m, 2H), 3.07 (t, J = 8.8 Hz, 2H), 2.02−1.95 (m, 2H),
1.77−1.69 (m, 2H), 1.25 (d, J = 6.4 Hz, 6H). LRMS (ESI): m/z 481 [M
+ H]+. Compound 103 was prepared as a white solid in 19% yield from
1-methylethyl 4-{[7-(4-bromo-3-fluorophenyl)-6,7-dihydro-5H-
pyrrolo[2,3-d]pyrimidin-4-yl]oxy}-1-piperidinecarboxylate in a similar
1
manner to that described for 25. H NMR (400 MHz, DMSO-d6): δ
8.42 (s, 1H), 8.11 (dd, J = 13.6, 1.6 Hz, 1H), 7.84 (app t, J = 8.0 Hz, 1H),
7.74 (dd, J = 8.8, 2.0 Hz, 1H), 5.32−5.25 (m, 1H), 4.75 (septuplet, J =
6.4 Hz, 1H), 4.10 (t, J = 8.8 Hz, 2H), 3.70−3.64 (m, 2H), 3.24 (app J =
9.6 Hz, 2H), 3.03 (t, J = 8.8 Hz, 2H), 1.96−1.89 (m, 2H), 1.61−1.53 (m,
2H), 1.16 (d, J = 6.0 Hz, 6H). HRMS (ESI): m/z calcd for C22H24FN5O3
[M + H]+ 426.1941, obsd 426.1943.
1-Methylethyl 4-{[7-(5-Cyano-2-pyridinyl)-6,7-dihydro-5H-
pyrrolo[2,3-d]pyrimidin-4-yl]oxy}-1-piperidinecarboxylate
(99). Compound 99 was prepared as a white solid in 5% overall yield
from 88 and 2-amino-5-cyanopyridine in a similar manner to that
described for 97, except that Na(OAc)3BH and CF3CO2H was used in
the reduction amination step in place of NaCNBH3 and acetic acid. 1H
NMR (400 MHz, CDCl3): δ 8.90−8.85 (m, 1H), 8.61−8.57 (m, 1H),
8.42 (s, 1H), 7.87−7.82 (m, 1H), 5.42−5.33 (m, 1H), 4.99−4.90 (m,
1H), 4.35 (t, J = 8.61 Hz, 2H), 3.89−3.74 (m, 2H), 3.42−3.29 (m, 2H),
3.07 (t, J = 8.1 Hz, 2H), 2.08−1.94 (m, 2H), 1.83−1.70 (m, 2H), 1.27
(d, J = 6.2 Hz, 6H). HRMS (ESI): m/z calcd for C21H24FN6O3 [M + H]+
409.1988, obsd 409.1989.
1-Methylethyl 4-({7-[3-Fluoro-4-(methylsulfonyl)phenyl]-
6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-4-yl}oxy)-1-piperidi-
necarboxylate (104). 1-Methylethyl 4-{[7-(4-bromo-3-fluorophen-
yl)-6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-4-yl]oxy}-1-piperidinecar-
boxylate was prepared as a white solid in 38% overall yield from 4-
bromo-3-fluoroaniline and 88 in a similar manner to that described for
97, except the cyclization occurred spontaneously. Compound 104 was
prepared from 1-methylethyl 4-{[7-(4-bromo-3-fluorophenyl)-6,7-
dihydro-5H-pyrrolo[2,3-d]pyrimidin-4-yl]oxy}-1-piperidinecarboxylate
and sodium methanesulfinate using the sulfonylation reaction
1-Methylethyl 4-{[7-(5-Fluoro-2-pyridinyl)-6,7-dihydro-5H-
pyrrolo[2,3-d]pyrimidin-4-yl]oxy}-1-piperidinecarboxylate
1
conditions described for 2. H NMR (400 MHz, CDCl3): δ 8.40 (s,
S
dx.doi.org/10.1021/jm301404a | J. Med. Chem. XXXX, XXX, XXX−XXX

Journal of Medicinal Chemistry
Article
1H), 8.15 (d, J = 13.5 Hz, 1H), 7.88 (dt, J = 8.7, 2.2 Hz, 1H), 7.43 (d, J =
9.0 Hz, 1H), 7.26 (d, J = 2.2 Hz, 1H), 5.35 (m, 1H), 4.93 (septuplet, J =
6.2 Hz, 1H), 4.11 (t, J = 8.0 Hz, 2H), 3.80 (m, 2H), 3.37−3.31 (m, 2H),
3.20 (s, 3H), 3.12 (t, J = 8.4 Hz, 2H), 2.94 (m, 2H), 2.02−1.90 (m, 2H),
1.80−1.70 (m, 2H), 1.25 (d, J = 6.3 Hz, 6H). HRMS (ESI): m/z calcd
for C22H27FN4O5S [M + H]+ 479.1764, obsd 479.1764.
necarboxylate (113). A solution of 4-fluoro-2-methyl-1-nitrobenzene
(109, 2.4 g, 19.3 mmol) in DMF was treated with sodium methoxide
(1.5 g, 21.3 mmol) and stirred and heated at 85 °C for 1 h. The mixture
was allowed to cool to RT and was treated with water and extracted with
EtOAc (3×). The combined organic extracts were dried over MgSO4,
filtered, and concentrated to afford 2-methyl-4-(methylthio)-1-nitro-
benzene (110, 3.1 g, 88%) as an orange residue. 1H NMR (400 MHz,
DMSO-d6): δ 7.95 (d, J = 8.6 Hz, 1H), 7.32 (d, J = 8.6 Hz, 1H), 7.27 (dd,
J = 8.6, 2.0 Hz, 1H), 2.54 (s, 3H). LRMS (ESI): m/z 184 (M + H)+.
Tin(II) chloride (10.7 g, 56.4 mmol) was added portionwise to
concentrated HCl (20 mL) at RT. The mixture was treated with a
solution of 2-methyl-4-(methylthio)-1-nitrobenzene (3.1 g, 17.1 mmol)
in ethanol (10 mL). The mixture was stirred and heated at 80 °C for 1 h.
The mixture was allowed to cool to RT, the pH was adjusted to ∼10 with
10N NaOH, and extracted with EtOAc (3×). The combined organic
extracts were dried over MgSO4, filtered, and concentrated to afford 2-
methyl-4-(methylthio)aniline (111, 2.1 g, 81%) as a dark oil. 1H NMR
(400 MHz, DMSO-d6): δ 7.00 (s, 1H), 6.96 (dd, J = 8.3, 2.0 Hz, 1H),
6.71 (d, J = 8.2 Hz, 1H), 2.34 (s, 3H), 2.07 (s, 3H). LRMS (ESI): m/z
154 (M + H)+. Compound 104 was prepared as a white solid in 5%
overall yield from 111 and 88 in a similar manner to that described for
97, with the exception that CF3CO2H was used for the cyclization step.
1H NMR (400 MHz, CDCl3): δ 8.19 (s, 1H), 7.86 (s, 1H), 7.79 (dd, J =
8.4, 1.7 Hz, 1H), 7.40 (d, J = 8.4 Hz, 1H), 5.37−5.29 (m, 1H), 5.00−4.83
(m, 1H), 3.99 (t, J = 8.8 Hz, 2H), 3.80 (d, J = 11.4 Hz, 2H), 3.40−3.23
(m, 2H), 3.13 (t, J = 8.8 Hz, 2H), 3.03 (s, 3H), 2.34 (s, 3H), 1.92 (dt, J =
6.3, 3.0 Hz, 2H), 1.23 (d, J = 6.2 Hz, 6H). LRMS (ESI): m/z 475 (M +
H)+. HRMS (ESI): m/z calcd for C23H30N4O5S [M + H]+ 475.2015,
obsd 475.2014.
1-Methylethyl 4-[(7-{4-[(Dimethylamino)sulfonyl]phenyl}-
6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-4-yl)oxy]-1-piperidi-
necarboxylate (105). Compound 105 was prepared as a white solid in
17% overall yield from 4-amino-N,N-dimethylbenzenesulfonamide and
1
88 in a similar manner to that described for 99. H NMR (400 MHz,
CDCl3): δ 8.38 (s, 1H), 7.96 (d, J = 9.0 Hz, 2H), 7.77 (d, J = 8.8 Hz, 2H),
5.42−5.30 (m, 1H), 4.99−4.86 (m, 1H), 4.14 (t, J = 8.8 Hz, 2H), 3.91−
3.75 (m, 2H), 3.41−3.27 (m, 2H), 3.12 (t, J = 8.8 Hz, 2H), 2.70 (s, 6H),
2.09−1.92 (m, 2H), 1.83−1.66 (m, 2H), 1.26 (d, J = 6.2 Hz, 6H).
HRMS (ESI): m/z calcd for C23H31N5O5S [M + H]+ 490.2124, obsd
490.2124.
1-Methylethyl 4-({7-[2-Fluoro-4-(methylthio)phenyl]-6,7-di-
hydro-5H-pyrrolo[2,3-d]pyrimidin-4-yl}oxy)-1-piperidinecar-
boxylate (106). A slurry of nickel(II) bromide (9.12 g, 42 mmol), 2,2′-
dipyridyl (6.56 g, 42 mmol), and zinc dust (55 g, 844 mmol) in
anhydrous DMF (300 mL) were treated with 2-fluoro-4-iodoaniline
(100 g, 0.42 mol) and dimethyldisulfide (96 mL, 1.06 mol) under N2.
The resultant mixture was stirred at 75 °C for 1.5 h. The reaction
mixture was concentrated. The crude residue was dissolved in Et2O (500
mL), filtered through a pad of Celite, and washed with Et2O. The filtrate
was concentrated and purified by flash chromatography to afford [2-
fluoro-4-(methylthio)phenyl]amine (48.7 g, 73%) as a brown liquid. 1H
NMR (400 MHz, DMSO-d6): δ 9.72 (s, 1H), 8.86 (s, 1H), 7.00 (dd, J =
11.2, 2.0 Hz, 1H), 6.93 (dd, J = 8.4, 4.0 Hz, 1H), 6.70 (t, J = 8.8 Hz, 1H),
4.21 (s, 2H), 3.69 (br s, 2H), 2.40 (s, 3H). LRMS (ESI): m/z 158 (M +
H)+. Compound 106 was prepared as an off-white solid in 34% overall
yield from [2-fluoro-4-(methylthio)phenyl]amine in a similar manner to
that described for 99. 1H NMR (400 MHz, DMSO-d6): δ 8.10 (s, 1H),
7.46 (t, J = 8.4 Hz, 1H), 7.21 (dd, J = 12.0, 2.0 Hz, 1H), 7.09 (dd, J = 8.0,
2.0 Hz, 1H), 5.26−5.20 (m, 1H), 4.75 (septuplet, J = 6.4 Hz, 1H), 3.96
(t, J = 8.8 Hz, 2H), 3.70−3.64 (m, 2H), 3.22 (app t, J = 9.6 Hz, 2H), 3.01
(t, J = 8.8 Hz, 2H), 1.94−1.88 (m, 2H), 1.59−1.50 (m, 2H), 1.17 (d, J =
6.4 Hz, 6H). HRMS (ESI): m/z calcd for C22H27FN4O3S [M + H]+
447.1866, obsd 447.1865.
( )-1,1-Dimethylethyl 4-({1-[4-(Methylsulfinyl)phenyl]-2,3-
dihydro-1H-indol-4-yl}oxy)-1-piperidinecarboxylate (107 and
108). A solution of 106 (200 mg, 0.454 mmol) and hexafluoroisopro-
panol (2.5 mL) was treated with 30% aqueous H2O2 (100 μL). The
mixture was stirred at RT for 0.5 h. A saturated sodium sulfide solution
(8 mL) was introduced to the above mixture and stirred for 5 min.
Layers were separated, and the organic layer was concentrated and
purified by flash chromatography (0→20% EtOAc/hexanes) to afford a
racemic mixture of 107 and 108 (178 mg, 86%) as an off-white solid. 1H
NMR (400 MHz, DMSO-d6): δ 7.61 (t, J = 8.8 Hz, 2H), 7.34 (d, J = 8.8
Hz, 2H), 7.04 (app t, J = 8.0 Hz, 1H), 6.83 (d, J = 8.0 Hz, 1H), 6.54 (d, J
= 8.4 Hz, 1H), 4.56 (septuplet, J = 3.2 Hz, 1H), 3.98 (t, J = 8.8 Hz, 2H),
3.60−3.55 (m, 2H), 3.25−3.22 (m, 2H), 2.99 (t, J = 8.4 Hz, 2H), 2.69 (s,
3H), 1.88−1.83 (m, 2H), 1.58−1.49 (m, 2H), 1.39 (s, 9H). LRMS
(ESI): m/z 457 (M + H)+.
GLP-1 Release from GluTag Cells with 3. GLUTag cells were
cultured in Dulbecco’s Modified Eagles Medium (Sigma-Aldrich)
containing 5.5 mM glucose, 10% fetal bovine serum, and 2 mM L-
glutamine.
GLUTag cells were plated in 384-well cell culture plates coated with
Matrigel (BD Biosciences) and grown to 80% confluence in Dulbecco’s
Modified Eagle’S Medium (Sigma-Aldrich) containing 1000 mg/L
glucose, pyroxidine HCl, and sodium bicarbonate and supplemented
with 2 mM L-glutamine and 10% fetal bovine serum. On the day of the
experiment, cell media was removed and cells were preincubated for 30
min in Kreb’s Ringers assay buffer (120 mM NaCl, 5 mM KCl, 2 mM
CaCl2, 1 mM MgCl2, 22 mM NaHCO3, 10 mM Hepes, 0.1% BSA, 10
mM D-glucose, and DPP-IV inhibitor (Millipore DPP4−010).
Following preincubation, media was again removed and replaced with
KRB assay buffer containing GSK252 at varying concentrations. Cells
were incubated for 2 h in a 37 °C, 5% CO2 incubator. After the
incubation period, media was collected and centrifuged to remove any
floating cells and assayed for GLP-1.
Capture antibody (Abcam ab23468) was biotinylated using the
QuickPlex Biotin NHS Ester kit (MSD R91DS-1) according to the
manufacturer’s protocol. Detection antibody (Abcam ab26278) was
sulfo-tagged using a MSD SULFO-TAG NHS Ester kit according to the
manufacturer’s protocol (MSD R91AN-1). Active GLP-1 was measured
using a chemiluminescent detection assay specific for amidated forms of
GLP-1 (1−36 amide, 7−36 amide, and 9−36 amide). Briefly, 15 μL of
cell supernatant plus 5 μL of assay buffer containing 2 μg/mL
biotinylated anti-GLP-1 capture antibody, 2 μg/mL sulfo-tagged anti-
GLP-1 detection antibody, 8% BSA Fraction V (Sigma-Aldrich), 4×
DPP-IV inhibitor (Millipore) in PBS pH = 7.4 were added to each well
of a 384 Standard avidin-coated plate (MSD L21AA-1). Plates were
sealed, shaken briefly, and incubated overnight at 40 °C. The following
day, samples were aspirated from wells and washed 3 times with 1× Tris
wash buffer (MSD R61TX-1). Wash buffer was aspirated, and 35 μL 1×
Read T buffer (MSD R92TC-2) was added. Plates were read on an MSD
Sector 6000 instrument.
(R)-1,1-Dimethylethyl 4-({1-[4-(Methylsulfinyl)phenyl]-2,3-
dihydro-1H-indol-4-yl}oxy)-1-piperidinecarboxylate (107). The
racemic sulfoxide (150 mg) was subjected to Chiral HPLC (column:
Chiralpak AS (analytical), Chiralpak (prep); mobile phase, 78% CO2,
22% MeOH (14 mL/min), pressure 140 bar, temperature 40 °C, 240
nm)) analysis and then was separated into its (R and S) enantiomers.
Retention time was 11.57 min for the first eluting enantiomer, which
provided 33 mg (%ee >98%). HRMS (ESI): m/z calcd for
C22H27FN4O4S [M + H]+ 463.1815, obsd 463.1816.
(S)-1,1-Dimethylethyl 4-({1-[4-(Methylsulfinyl)phenyl]-2,3-di-
hydro-1H-indol-4-yl}oxy)-1-piperidinecarboxylate (108). Reten-
tion time was 14.04 min for the second eluting isomer, 38 mg (%ee
>97%). HRMS (ESI): m/z calcd for C22H27FN4O4S [M + H]+
463.1815, obsd 463.1815.
Primary Colonic Crypt Secretion of GLP-1 with 3. Primary
colonic crypts were prepared from 6−8 week old C57BL/6 male mice.
Intestine was removed, cecum to rectum, filled and flushed three times
with cold PBS, and then slit open longitudinally and cut into 2−3 mm
lengths. Tissue sections from five animals were pooled and washed with
Hank’s Balanced Salt Solution (Gibco) five times, allowing sections to
1-Methylethyl 4-({7-[2-Methyl-4-(methylsulfonyl)phenyl]-
6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidin-4-yl}oxy)-1-piperidi-
T
dx.doi.org/10.1021/jm301404a | J. Med. Chem. XXXX, XXX, XXX−XXX

Journal of Medicinal Chemistry
Article
sediment under gravity each time, and then transferred to a Petri dish
and diced to a very fine mince. The tissue slurry was incubated in 0.1%
Collagenase II−S (Sigma), 0.2% Dispase I (Becton Dickenson), 2000
Kunitz DNase-I (Sigma), and 0.1% Hyaluronidase I−S (Sigma) in
culture media (Dulbecco’s Modified Eagles Medium Hi Glucose
(Gibco), 10% heat inactivated FBS (Gibco), 1× Pen/Strep (Gibco))
for 15 min at 37 °C while gently rocking. First digest was triturated 50
times with a 10 mL pipet, and the contents allowed to sediment under
gravity for 1 min. Then 20 mL of the supernatant was carefully removed .
Two additional digests were performed on the remaining tissue
fragments with fresh enzyme solution as described above, saving the
supernatant each time. All digest fractions were pooled for gentle spins
(3 min, 300 rpm in swinging bucket centrifuge) and washed in Hank’s
Balanced Salt Solution containing 5 mM β-mercaptoethanol until the
supernatant was clear. The pellet of crypts was resuspended in culture
media and seeded on 384-well cell culture plates coated with Matrigel
(BD Biosciences) and allowed to attach for 24 h. GLP-1 levels were
measured as described above.
min, food hoppers containing normal chow (Purina) were weighed and
reintroduced. Hopper wights were recorded after 1, 2, 4, 6, and 24 h
Rat Gastric Emptying with 3. Male Sprague−Dawley rats (Charles
River Laboratories, USA) were fasted overnight and administered
vehicle (dose volume of 10 mL/kg, 0.5% HPMC/0.1% Tween80) or
compound 3 (10 mg/kg in dose volume of 10 mL/kg) orally at time =
−120 min. At time = −15 min, the positive control rat cohort was
administered Pramlintide (15 μg in dose volume of 10 mL/kg,
Polypeptide). At time = 0, baseline plasma samples were collected,
followed by gavage of acetominophen (65 mg in vehicle at a dose
volume of 10 mL/kg). The concentration time profile of acetominophen
was measured over the next 3 h.
AUTHOR INFORMATION
Corresponding Author
■
*For S.R.K.: phone, (919) 314-5528; fax, (919) 314-5529; E-
mail, skatamreddy@nc.rr.com. For A.J.C.: phone, (919) 483-
9868; fax, (919) 315-6923; E-mail, andrew.j.carpenter@gsk.com.
Incretin Secretion in WT and KO Mice with 3. First, 17-week-old
transgenic male (WT) and GPR119 knockout (KO) mice (Charles
River Laboratories, USA) were fasted overnight and then randomly
distributed into treatment groups (n = 8 and 10 per group, repsectively).
Animals received wither vehicle (0.5% HPMC/0.1% Tween80) or
compound 3 (30 mg/kg) by oral gavage at a dose volume of 10 mL/kg.
Then 60 min after the oral gavage, mice were anesthetized using
isofluorane and a blood sample was collected by cardiac puncture. All
mice were euthanized by cervical dislocation immediately following
cardiac puncture. Blood samples were placed in chilled K2-EDTA coated
capiject tubes (cat. no. T-MQK, Terumo Medical Corp., Elkton, MD)
supplemented with a DPP-4 inhibitor and Aprotenin (Trasylol; Bayer
Pharmaceuticals, West Haven, CT), yielding a final concentration of 30
μL and 250 KIU/mL, respectively. The tubes were immediately
centrifuged at 4 °C and the plasma collected and stored at −80 °C for
subsequent hormone measurements. Total GLP-1 levels were
determined using a Multi-Array electrochemoluminescence assay from
Meso Scale Discovery (cat. no. K110-FAC-2; Gaithersburg, MD). Total
GIP levels were determined using an ELISA assay (cat. no. EZRMGIP-
55K, Linco Diagnostic Services, St. Charles, MO). Data are represented
as box plots summarizing the distribution of data for each treatment
group. Horizontal lines represent the mean data.
Hyperglycemic Clamp in Rats with 3. Male Wistar rats (Taconic
Farms, USA), weighing 300−350 g, had catheters surgically implanted
into the jugular and femoral veins and allowed to recover for one week
following the surgery. Animals were fasted overnight. Animals were then
randomly assigned to treatment groups, vehicle (n = 19), compound 3
(1 mg/kg, n = 8), or compound 3 (10 mg/kg, n = 8), and baseline blood
samples were collected from the jugular vein. Animals were
administered vehicle or compound 3 by oral gavage. A second blood
sample was collected approximately 90 min after the oral gavage and
immediately prior to starting the hyperglycemic clamp. The clamp was
initiated by infusing an empirically determined glucose bolus over 15−
30 s into the femoral vein, followed by a variable rate glucose infusion
adjusted to achieve a stable plasma glucose level of 190−210 mg/dL.
After infusion of the glucose bolus, blood samples were collected every 5
min for glucose measurements. Samples for C-peptide levels were
collected at 2, 5, 10, 15, 20, 25, 30, 45, 60, 90, and 120 min during the
hyperglycemic clamp. All blood samples were placed in individual tubes
containing a gel clot activator (cat. no. T-MG, Terumo Medical
Corporation, Elkton, MD) and allowed to clot at room temperature for
20 min before centrifugation and collection of serum. Glucose levels
were measured using an Olympus AU640 analyzer (Olympus America
Inc., Melville, NY) and C-peptide levels by an RIA (Linco Diagnostic
Services, St. Charles, MO).
Present Address
†Vijaya Pharmaceuticals, 104 T. W. Alexander Drive, Building #5,
Research Triangle Park, North Carolina 27709, United States.
Notes
The authors declare no competing financial interest.
ACKNOWLEDGMENTS
■
We thank the Analytical Chemistry group of Matt Lochansky and
Iris Scherer for performing HRMS analysis for the compounds
reported and Manon Villeneuve, Yang Zhao, Eric Pullen, Eric
Wentz, and Bill Hall for performing chiral separations. We also
thank Brad Henke for preparing compound 114 to be used as the
standard in the in vitro assay.
ABBREVIATIONS USED
■
T2D, type 2 diabetes; GPR119, G protein-coupled receptor 119;
GIP, glucose-dependent insulinotropic peptide; DPP-4, dipep-
tidyl peptidase-4; LPC, lysophosphatidylcholine; OEA, oleoyle-
thanolamide; NAPE, N-acylethanolamide; OLDA, N-oleoyldop-
amine; DHPP, 6,7-dihydro-5H-pyrrolopyrimidine; DIAD, diiso-
propyl azodicarboxylate; 1,2-DCE, 1,2-dichloroethane; pEC50,
negative logarithm of the EC50; EC50, effective concentration at
half-maximal response; av, average; StDev, standard deviation;
AUC0−24h, area under the curve from 0 to 24 h; DNAUC0−24h
dose normalized area under the curve from 0 to 24 h; Cmax
maximum concentration; Clb, blood clearance; Vss, volume of
distribution; KO, knockout; PYY, peptide YY; SD, Sprague−
Dawley; nd, not determined; HPMC, hydroxypropylmethylcel-
lulose; HP-β-CD, hydroxypropyl-β-cyclodextrin; FBS, fetal
bovine serum
,
,
REFERENCES
■
(1) National Diabetes Fact Sheet, 2011; Centers for Disease Control and
Prevention: Atlanta, GA, 2011; http://www.cdc.gov/diabetes/pubs/
pdf/ndfs_2011.pdf (accessed November 7, 2012).
(2) Wild, S.; Roglic, G.; Green, A.; Sicree, R.; King, H. Global
prevalence of diabetes: Estimates for the year 2000 and projections for
2030. Diabetes Care 2004, 27, 1047−1053.
(3) Koro, C. E.; Bowlin, S. J.; Bourgeois, N.; Fedder, D. O. Glycemic
Control From 1988 to 2000 among U.S. Adults Diagnosed with Type 2
Diabetes. Diabetes Care 2004, 27, 17−20.
(4) Philippe, J.; Raccah, D. Treating type 2 diabetes: how safe are
current therapeutic agents? Int. J. Clin. Pract. 2009, 63, 321−332.
(5) (a) Hansen, K. B.; Knop, F. K.; Holst, J. J.; Vilsbøll, T. Treatment of
type 2 diabetes with glucagon-like peptide-1 receptor agonists. Int. J.
Clin. Pract. 2009, 63, 1154−1160. (b) Mohler, M. L.; He, Y.; Wu, Z.;
Mouse Food Intake with 3. Individually housed, male C57BL/6
mice were fasted overnight. Body weights were recorded, and
compound 3 (10 mg/kg in dose volume of 10 mL/kg, 10%
DMSO:30% solutol:60% HP-β-CD) was delivered by oral gavage at
time = −60 min. Control (Exendin-4, 100 μg/kg in dose volume of 10
mL/kg, Polypeptide) was delivered IP at time = −30 min. At time = 0
U
dx.doi.org/10.1021/jm301404a | J. Med. Chem. XXXX, XXX, XXX−XXX

Journal of Medicinal Chemistry
Article
Hwang, D. J.; Miller, D. D. Recent and Emerging Anti-Diabetes Targets.
Med. Res. Rev. 2009, 29, 125−195. (c) Ashiya, M.; Smith, R. E. Non-
insulin therapies for type 2 diabetes. Nature Rev. Drug Discovery 2007, 6,
777−778.
(6) (a) Soga, T.; Ohishi, T.; Matsui, T.; Saito, T.; Matsumoto, M.;
Takasaki, J.; Matsumoto, S.; Kamohara, M.; Hiyama, H.; Yoshida, S.;
Momose, K.; Ueda, Y.; Matsushime, H.; Kobori, M.; Furuichi, K.
Lyophosphatidylcholine enhances glucose-dependent insulin secretion
via orphan G-protein-coupled receptor. Biochem. Biophys. Res. Commun.
1066. (c) Flock, G.; Holland, D.; Seino, Y.; Drucker, D. J. GPR119
Regulates Murine Glucose Homeostasis Through Incretin Receptor-
Dependent and Independent Mechanisms. Endocrinology 2011, 152,
374−383. (d) Parker, H. E.; Habib, A. M.; Rogers, G. J.; Gribble, F. M.;
Reimann, F. Nutrient-dependent secretion of glucose-dependent
insulinotropic polypeptide from primary murine K cells. Diabetologia
2009, 52, 289−298. (e) Ammala, C.; Bullard, S.; Kashatus, J.;
Katamreddy, S.; Way, J.; Carpenter, A. GPR119 dependent hormone
secretion: insulin, GLP-1 and more. Keystone Symposium, Islet and β-
Cell Biology, Snowbird, UT, April 6-11, 2008, Abstract 102.
(15) Jones, R. M.; Leonard, J. N.; Buzard, D. J.; Lehmann, J. GPR119
agonists for the treatment of type 2 diabetes. Expert Opin. Ther. Pat.
2009, 19 (10), 1340−1359.
2005, 326, 744−751. (b) Fredriksson, R.; Hoglund, P. J.; Gloriam, D. E.
̈
I.; Lagerstrom, M. C.; Schioth, H. B. Seven evolutionary conserved
̈
̈
human rhodopsin G protein-coupled receptors lacking close relatives.
FEBS Lett. 2003, 554, 381−388.
(7) (a) Bonini, J. A.; Borowsky, B. E.; Adham, N.; Boyle, N.;
Thompson, T. O. DNA encoding SNORF25 receptor. US Patent
6,221,660-B1, 2001. (b) Bonini, J. A.; Borowsky, B. E.; Adham, N.;
Boyle, N.; Thompson, T. O. Methods of identifying compounds that
bind to SNORF25 receptors. US Patent 6,468,756-B1, 2002.
(8) (a) Sakamoto, Y.; Inoue, H.; Kawakami, S.; Miyawaki, K.;
Miyamoto, T.; Mizuta, K.; Itakura, M. Expression and distribution of
Gpr119 in the pancreatic islets of mice and rats: predominant
localization in pancreatic polypeptide-secreting cells. Biochem. Biophys.
Res. Commun. 2006, 351, 474−480. (b) Chu, Z.-L.; Jones, R. M.; He, H.;
Carroll, C.; Gutierrez, V.; Lucman, A.; Moloney, M.; Gao, H.; Mondala,
H.; Bagnol, D.; Unett, D.; Liang, Y.; Demarest, K.; Semple, G.; Behan, D.
P.; Leonard, J. A Role for β-Cell-Expressing G Protein-Coupled
Receptor 119 in Glycemic Control by Enhancing Glucose-Dependent
Insulin Release. Endocrinology 2007, 148, 2601−2609. (c) Chu, Z.-L.;
Carroll, C.; Alfonso, J.; Gutierrez, V.; He, H.; Lucman, A.; Pedraza, M.;
Mondala, H.; Gao, H.; Bagnol, D.; Chen, R.; Jones, R. M.; Behan, D. P.;
Leonard, J. A Role for Intestinal Endocrine Cell-Expressing G Protein-
Coupled Receptor 119 in Glycemic Control by Enhancing Glucagon-
Like Peptide-1 and Glucose-Dependent Insulinotropic Peptide Release.
Endocrinology 2008, 149, 2038−2647. (d) Parker, H. E.; Habib, A. M.;
Rogers, G. J.; Gribble, F. M.; Reimann, F. Nutrient-dependent secretion
of glucose-dependent insulinotropic polypeptide from primary murine
K cells. Diabetologia 2009, 52, 289−298.
(9) Jones, R. M.; Leonard, J. N. The Emergence of GPR119 Agonists as
Anti-Diabetic Agents. In Annual Reports in Medicinal Chemistry, Vol. 44;
Macor, J. E.; Stamford, A. W., Eds.; Academic Press: New York, 2009; pp
149−170.
(10) Overton, H. A.; Babbs, A. J.; Doel, S. M.; Fyfe, M. C. T.; Gardner,
L. S.; Griffin, G.; Jackson, H. C.; Procter, M. J.; Rasamison, C. M.; Tang-
Christensen, M.; Widdowson, P. S.; Williams, G. M.; Reynet, C.
Deorphanization of a G protein-coupled receptor for oleoylethanola-
mide and its use in the discovery of small-molecule hypophagic agents.
Cell Metab. 2006, 3, 167−175.
(11) Gillum, M. P.; Zhang, D.; Zhang, X.-M.; Erion, D. M.; Jamison, R.
A.; Choi, C.; Dong, J.; Shanabrough, M.; Duenas, H. R.; Frederick, D.
W.; Hsiao, J. J.; Horvath, T. L.; Lo, C. M.; Tso, P.; Cline, G. W.;
Shulman, G. I. N-Acylphosphatidylethanolamine, a Gut-Derived
Circulating Factor Induced by Fat Ingestion, Inhibits Food Intake.
Cell 2008, 135, 813−824.
(12) Chu, Z.-L.; Carroll, C.; Chen, R.; Alfonso, J.; Gutierrez, V.; He, H.;
Lucman, A.; Xing, C.; Sebring, K.; Zhou, J.; Wagner, B.; Unett, D.; Jones,
R. M.; Behan, D. P.; Leonard, J. N-Oleoyldopamine Enhances Glucose
Homeostasis through the Activation of GPR119. Mol. Endocrinol. 2010,
24, 161−170.
(13) Kogure, R.; Toyama, K.; Hiyamuta, S.; Kojima, I.; Takeda, S. 5-
Hydroxy-eicosapentaenoic acid is an endogenous GPR119 agonist and
enhances glucose-dependent insulin secretion. Biochem. Biophys. Res.
Commun. 2011, 416, 58−63.
(14) (a) Carpenter, A.; Ammala, C.; Briscoe, C.; Bullard, S.; Kashatus,
J.; Katamreddy, S.; Mertz, R.; Ross, S. The in vitro and in vivo effects of a
GPR119 agonist. Keystone Symposium, Diabetes Mellitis, Insulin
Action and Resistance, Breckenridge, CO, January 22−27, 2008.
(b) Lauffer, L. M.; Iakoubov, R.; Brubaker, P. L. GPR119 Is Essential
for Oleoylethanolamide-Induced Glucagon-Like Peptide-1 Secretion
From the Intestinal Enteroendocrine L-Cell. Diabetes 2009, 58, 1058−
(16) (a) Overton, H. A.; Fyfe, M. C. T.; Reynet, C. GPR119, a novel G
protein-coupled receptor target for the treatment of type 2 diabetes and
obesity. Br. J. Pharmacol. 2008, 153, S76−S81. (b) Fyfe, M. C. T.;
McCormack, J. G.; Overton, H. A.; Procter, M. J.; Reynet, C. GPR119
agonists as potential new oral agents for the treatment of type 2 diabetes
and obesity. Expert Opin. Drug Discovery 2008, 3, 403−413.
(17) Jones, R. M.; Semple, G.; Xiong, Y.; Shin, Y.-J.; Ren, A. S.;
Calderon, I.; Fioravanti, B.; Choi, J. S. K.; Sage, C. R. Fused-aryl and
heteroaryl derivatives as modulators of metabolism and the prophylaxis
and treatment of disorders related thereto. WO 2005/007658A2, 2005.
(18) Semple, G.; Fioravanti, B.; Pereira, G.; Calderon, I.; Uy, J.; Choi,
K.; Xiong, Y.; Ren, A.; Morgan, M.; Dave, V.; Thomsen, W.; Unett, D. J.;
Xing, C.; Bossie, S.; Carroll, C.; Chu, Z.-L.; Grottick, A. J.; Hauser, E. K.;
Leonard, J.; Jones, R. M. Discovery of the First Potent and Orally
Efficacious Agonist of the Orphan G-Protein Coupled Receptor 119. J.
Med. Chem. 2008, 51, 5172−5175.
(19) (a) Katamreddy, S. R.; Caldwell, R. D.; Heyer, D.; Samano, V.;
Thompson, J. B.; Carpenter, A. J.; Conlee, C. R.; Boros, E. E.;
Thompson, B. D. Preparation of pyrrolo[2,3-d]pyrimidine derivatives as
GPR119 agonists. WO 2008/008887A2, 2008. (b) Ammala, C.; Briscoe,
C. GPR119 agonists for the treatment of diabetes and related disorders.
WO 2008/008895A1, 2008.
(20) For a review, see: Yang, B. H.; Buchwald, S. L. Palladium-catalyzed
amination of aryl halides and sulfonates. J. Organomet. Chem. 1999, 576,
125−146.
(21) Zhu, W.; Ma, D. Synthesis of aryl sulfones via L-proline-promoted
CuI-catalyzed coupling reaction of aryl halides with sulfinic acid salts. J.
Org. Chem. 2005, 70, 2696−2700.
(22) Gribble, G. W.; Hoffman, J. H. Reactions of sodium borohydride
in acidic media; VI. Reduction of indoles with cyanoborohydride in
acetic acid. Synthesis 1977, 12, 859−860.
(23) For a previously reported synthesis of 6,7-dihydro-5H-
pyrrolo[2,3-d]pyrimidines, see: Di Fabio, R.; Marchionni, C.;
Micheli, F.; Pasquarello, A.; Perini, B.; St.-Denis, Y. Preparation of
substituted 6,7-dihydro-5H-pyrrolo[2,3-d]pyrimidines as corticotropin
releasing factor antagonists. WO 2002/100863A1, 2002.
(24) Boros, E.; Thompson, J. B.; Katamreddy, S. R.; Carpenter, A. J.
Facile Reductive Amination of Aldehydes with Electron-Deficient
Anilines by Acyloxyborohydrides in TFA: Application to a Diazaindo-
line Scale-Up. J. Org. Chem. 2009, 74, 3587−3590.
(25) Friedman, L.; Shechter, H. Dimethylformamide as a useful solvent
in preparing nitriles from aryl halides and cuprous cyanide; improved
isolation techniques. J. Org. Chem. 1961, 26, 2522−2524.
(26) Hermann, W. The Suzuki Cross-Coupling. In From Applied
Homogeneous Catalysis with Organometallic Compounds, 2nd ed.; Cornils,
B., Herrmann, W. A., Eds.; Wiley-VCH Verlag GmbH: Weinheim,
Germany, 2002, pp 591−598.
(27) Preparation of the thiocarbamates was accomplished by two
different methods. (a) An example of a secondary, cyclic amine with S-
methyl chlorothioformate in the presence of triethylamine: Fishbein, P.
L.; Kohn, H. Synthesis and Antineoplastic Activity of 1a-Formyl and 1a-
Thioformyl Derivatives of Mitomycin C and 2-Methylaziridine. J. Med.
Chem. 1987, 30, 1767−1773. (b) Grzyb, J. A.; Batey, R. A.
Carbamoylimidazolium salts as diversification reagents: an application
to the synthesis of tertiary amides from carboxylic acids. Tetrahedron
Lett. 2003, 44, 7485−7488.
V
dx.doi.org/10.1021/jm301404a | J. Med. Chem. XXXX, XXX, XXX−XXX

Journal of Medicinal Chemistry
Article
(28) Danishefsky, S.; Singh, R. K. A Highly Activated Cyclopropane for
Homoconjugate Reactions. J. Am. Chem. Soc. 1975, 97, 3239−3241.
Results of Single & Multiple Dose Studies. American Diabetes
Association Meeting, New Orleans, LA, 2009, 164-OR.
(29) Ravikumar, K. S.; Beg
Conversion of Sulfide to Sulfoxide in Hexafluoro-2-propanol.
Tetrahedron Lett. 1998, 39, 3141−3144.
́
ue,
́
J.-P.; Bonnet-Delpon, D. A Selective
(41) Katz, L. B.; Gambale, J. J.; Rothenberg, P. L.; Vanapalli, S. R.;
Vaccaro, N.; Xi, L.; Polidori, D. C.; Vets, E.; Sarich, T. C.; Stein, P. P.
Pharmacokinetics, Pharmacodynamics, Safety, and Tolerability of JNJ-
38431055, a Novel GPR119 Receptor Agonist and Potential
Antidiabetes Agent, in Healthy Male Subjects. Clin. Pharmacol. Ther.
2011, 90 (5), 685−692.
(30) Stephens, P. J.; Devlin, F. J.; Pan, J.-J. The Determination of the
Absolute Configurations of Chiral Molecules Using Vibrational Circular
Dichroism (VCD) Spectroscopy. Chirality 2008, 20, 643−663.
(31) The assay consists of CHO-K1 6CRE-luciferase cells that stably
express human GPR119 receptor plated at 15000 cells/well in
Dulbecco’s Modified Eagle Medium: Nutrient Mixture F12 (DMEM/
F12), 5% fetal bovine serum (FBS), and 2 mM L-glutamine in black 384-
well assay plates. On the following day, the media is removed by
aspiration and replaced with 20 μL of DMEM/F12, 2 mM L-glutamine
(no FBS) utilizing a Matrix Multidrop. Test compounds (20 μL) are
then pipetted into the assay plate using a Packard Minitrak. The plates
are then incubated for 5 h at 37 °C. Under subdued light conditions, 15
μL of a 1:1 solution containing SteadyLite and Dulbecco’s Phosphate
Biffered Saline with 1 mM CaCl2 and 1 mM MgCl2 is added to the plates
using a Matrix Multidrop Plates are then sealed with self-adhesive clear
plate seals and the amount of luciferase generated is quantified in a
Wallac Viewlux. Compounds are also tested in the same manner against
cells without the GPR119 receptor so as to check for false positives.
(32) All compounds are referenced to a small molecule GPR119
agonist for evaluation of intrinsic efficacy (%max response) because this
reporter assay is not sensitive enough to demonstrate robust activity of
putative endogenous ligands (OEA, LPC, etc.). Our experience with the
∼3000 compounds made for the program is that compounds with ∼80−
200% max response are likely full agonists. Compounds with consistent
levels of receptor activation below 80% may be partial agonists. The
degree of variability in the assay accounts for the large range considered
as full agonists. The structure of our standard compound 114, with an
EC50 = 0.2 μM (max response = 100%), is shown below. This compound
is reported33 to have an EC50 = 0.13 μM in a cAMP assay.
(42) Carpenter, A. J. Discovery of GSK1292263A, a GPR119 Agonist
for the Treatment of T2D. National Medicinal Chemistry Symposium,
Minneapolis, MN, 2010, OS2−2.
(43) Nunez, D. J.; Bush, M. A.; Collins, D. A.; McMullen, S. L.;
̃
Feldman, P. L.; Jackson, T. D.; Ross, S. A. Evaluation of GSK1292263, a
Novel GPR119 Agonist, in Type 2 Diabetes Mellitus (T2DM): Safety,
Tolerability, Pharmacokinetics (PK) and Pharmacodynamics (PD) of
Single and Multiple Doses. American Diabetes Association Meeting, San
Diego, CA, 2011, 996-P.
(33) Jones, R. M.; Semple, G.; Fioravanti, B.; Pereira, G.; Calderon, I.;
Uy, J.; Duvvuri, K.; Choi, J. S. K.; Xiong, Y.; Dave, V. 1,2,3-Trisubstituted
Aryl and Heteroaryl Derivatives as Modulators of Metabolism and the
Prophylaxis and Treatment of Disorders Related Thereto Such as
Diabetes and Hyperglycemia. WO 2004/065380 A1, 2004.
(34) Leeson, P. D.; Springthorpe, B. The influence of drug-like
concepts on decision-making in medicinal chemistry. Nature Rev. Drug
Discovery 2007, 6, 881−890.
(35) All studies were conducted after review by the GSK Institutional
Animal Care and Use Committee and in accordance with the GSK
Policy on the Care, Welfare and Treatment of Laboratory Animals.
(36) Lee, Y. C.; Asa, S. L.; Drucker, D. J. Glucagon Gene 5′-Flanking
Sequences Direct Expression of Simian Virus 40 Large T Antigen to the
Intestine, Producing Carcinoma of the Large Bowel in Transgenic Mice.
J. Biol. Chem. 1992, 267, 10705−10709.
(37) Kaneko, T.; Kaneto, A.; Yanaihara, N.; Yasuda, H.; Suzuki, S.;
Oka, H.; Oda, T. Insulin and C-peptide secretion in animals. Int. Congr.
Ser., 1979; Vol. 468, Proinsulin, Insulin, C-Pept. , pp 138−147.
(38) Barlocco, D. LAF-237 Novartis. Curr. Opin. Investig. Drugs 2004,
5, 1094−1100.
(39) Hatanaka, S.; Kondoh, M.; Kawarabayashi, K.; Furuhama, K. The
Measurement of Gastric Emptying in Conscious Rats by Monitoring
Serial Changes in Serum Acetaminophen Level. J. Pharmacol. Toxicol.
Methods 1994, 31, 161−165.
(40) Roberts, B.; Gregoire, F. M.; Karpe, D. B.; Clemens, E.; Lavan, B.;
Johnson, J.; McWherter, C. A.; Martin, R.; Wilson, M. MBX-2982, a
Novel Oral GPR119 Agonist for the Treatment of Type 2 Diabetes:
W
dx.doi.org/10.1021/jm301404a | J. Med. Chem. XXXX, XXX, XXX−XXX