Omarigliptin (MK-3102): A novel long-acting DPP-4 inhibitor for once-weekly treatment of type 2 diabetes

Article abstract of DOI:10.1021/jm401992e

In our effort to discover DPP-4 inhibitors with added benefits over currently commercially available DPP-4 inhibitors, MK-3102 (omarigliptin), was identified as a potent and selective dipeptidyl peptidase 4 (DPP-4) inhibitor with an excellent pharmacokinetic profile amenable for once-weekly human dosing and selected as a clinical development candidate. This manuscript summarizes the mechanism of action, scientific rationale, medicinal chemistry, pharmacokinetic properties, and human efficacy data for omarigliptin, which is currently in phase 3 clinical development.

Full text of DOI:10.1021/jm401992e

Drug Annotation
pubs.acs.org/jmc
Omarigliptin (MK-3102): A Novel Long-Acting DPP‑4 Inhibitor for
Once-Weekly Treatment of Type 2 Diabetes
Tesfaye Biftu,*,† Ranabir Sinha-Roy,‡ Ping Chen,† Xiaoxia Qian,† Dennis Feng,† Jeffrey T. Kuethe,†
Giovanna Scapin,† Ying Duo Gao,† Youwei Yan,† Davida Krueger,⊥ Annette Bak,⊥ George Eiermann,§
Jiafang He,† Jason Cox,† Jacqueline Hicks,† Kathy Lyons,∥ Huaibing He,∥ Gino Salituro,∥ Sharon Tong,∥
Sangita Patel,† George Doss,† Aleksandr Petrov,§ Joseph Wu,‡ Shiyao Sherrie Xu,∥ Charles Sewall,#
Xiaoping Zhang,‡ Bei Zhang,‡ Nancy A. Thornberry,‡ and Ann E. Weber†
‡
§
∥
†Department of Discovery Chemistry, Department of Metabolic Disorders, Department of Pharmacology, Department of Drug
⊥
#
Metabolism, Basic Pharmaceutical Sciences, and Department of Safety Assessment and Animal Resources, Merck & Co., Inc.,
Whitehouse Station, New Jersey 08889, United States
ABSTRACT: In our effort to discover DPP-4 inhibitors with added benefits over currently
commercially available DPP-4 inhibitors, MK-3102 (omarigliptin), was identified as a potent
and selective dipeptidyl peptidase 4 (DPP-4) inhibitor with an excellent pharmacokinetic
profile amenable for once-weekly human dosing and selected as a clinical development
candidate. This manuscript summarizes the mechanism of action, scientific rationale, medicinal
chemistry, pharmacokinetic properties, and human efficacy data for omarigliptin, which is
currently in phase 3 clinical development.
shown to inhibit glucagon release and decrease in gastric
emptying. In rodent studies, it promotes the regeneration and
differentiation of islet β-cells.2a,b
However, GLP-1 is not a stable peptide and is cleaved rapidly
to its inactive form by DPP-4. Inhibition of DPP-4 prolongs the
circulating half-life of glucagon-like peptide 1 and glucose-
dependent insulinotropic polypeptide.2c By increasing the
circulating concentration of GLP-1 and GIP, DPP-4 inhibitors
improve glucose control in patients with type 2 diabetes.2d−f
STRUCTURE
■
Omarigliptin (MK-3102) (2R,3S,5R)-2-(2,5-difluorophenyl)-5-
[2-(methylsulfonyl)-2,6-dihydropyrrolo[3,4-c]pyrazol-5(4H)-
yl]tetrahydro-2H-pyran-3-amine,
SCIENTIFIC RATIONALE FOR BRINGING
OMARIGLIPTIN TO CLINIC
■
is a 2,3,5-substituted tetrahydropyran analogue currently in
phase 3 clinical trial for type 2 diabetes mellitus (T2DM).
Given the clinical success of sitagliptin (1), a once-daily
antidiabetic agent, our laboratories have been interested in
generating structurally diverse, best-in-class DPP-4 inhibitors.
In particular, we were interested in discovering analogues with
long half-lives that are amenable for once-weekly dosing.
Medication adherence in chronic diseases such as T2DM may
be low, averaging approximately 50%.3
Side effects associated with antihyperglycaemic agents
(AHA), including hypoglycaemia, weight gain, and gastro-
intestinal intolerance, are common patient-reported reasons for
poor medication adherence. Adherence to AHAs is also directly
related to the number of pills prescribed or complexity in
dosing regimens. Thus, the convenience of an effective, well-
tolerated, weekly oral AHA has the potential to improve
medication adherence, which may translate into better
outcomes for patients with T2DM. The currently marketed
DISEASE TARGET
■
Type 2 diabetes mellitus is a chronic, progressive disease that
affects 366 million people globally, including nearly 26 million
people in the United States. T2DM is a growing epidemic.1
Commercial proof of concept for DPP-4 inhibitors has been
established with sitagliptin, Merck’s lead DPP-4 inhibitor (1,
Figure 1), which was the first DPP-4 inhibitor approved by the
United States Food and Drug Administration in October 2006.
Subsequently, additional DPP-4 inhibitors have entered the
market. These include vildagliptin (2), saxagliptin (3),
linagliptin (4), and alogliptin (5).
MECHANISM OF ACTION
■
The incretin hormones, glucagon-like peptide 1 (GLP-1) and
glucose-dependent insulinotropic polypeptide (GIP), are
released by the gut in response to food intake and an increase
in blood glucose. These hormones stimulate insulin secretion in
a glucose-dependent manner. Additionally, GLP-1 has been
Received: January 6, 2014
© XXXX American Chemical Society
A
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Drug Annotation
Figure 1. DPP-4 inhibitors.
drugs (1−5) including sitagliptin are prescribed as a once-daily
oral medication.
reduced selectivity against related proteases, QPP (IC50 = 160
nM).
Over the course of our studies, several lead classes have
evolved. Critical to this work was the development of
cyclohexylamine (6),4 a structurally distinct rigid analogue of
sitagliptin (1). The cyclohexylamine analogues are potent and
selective against other related proteases with excellent
pharmacokinetic profiles; however, they block the human
potassium channel hERG (human-ether-a-go-go related gene)
at low micromolar concentrations. This has pharmacological
result of prolonging QTc (a measure of the time between the
start of the Q wave and the end of the T wave in the heart’s
electrical cycle) in a cardiovascular dog model (CV-dog).5 The
corresponding tetrahydropyran (7) analogues4c have reduced
basicity compared to the cyclohexylamines and thereby reduced
hERG potency. In addition, they do not affect QTc in the
cardiovascular CV-dog model. Following systematic structure−
activity-relationship (SAR) studies of diverse tetrahydropyran
analogues, omarigliptin was selected as a clinical candidate.
Figure 2. Formation of pyrrolopyrimidine metabolite.
Several attempts were made to circumvent the metabolism
on the right-hand-side amine. Attempts to block the oxidation
sites with gem-dimethyl groups gave compounds with much
reduced potency.
We then considered evaluation of several 5,5-heterocycles.4c
Pyrrolooxazole is less potent. Pyrroloimidazole has good
potency, but selectivity against DPP8 is poor. However,
pyrrolopyrazole 7b gave compounds with good potency and
selectivity (Figure 3). In addition, 7b was found to be stable
and no oxidation product was observed when dosed in rats or
dogs.
STRUCTURE−ACTIVITY RELATIONSHIP LEADING
TO PRODUCT: KEY CHALLENGES
■
Various substituted tetrahydropyran analogues were made and
examined for their in vitro inhibition of human DPP-4 and
related enzymes6 such as QPP,7 DPP8,8 and FAP.9 Inhibition of
DPP8 and DPP910 has been associated with toxicity in
preclinical species.11 DPP9 inhibition typically tracks closely
with DPP8 and thus was not reported for this series. The
compounds were also evaluated in the hERG assay.
The cyclohexylamine analogue 6a reported in our earlier
publications4 is a potent DPP-4 inhibitor (IC50 = 0.5 nM).
However, selectivity against IKr (IC50 = 4.8 μM) was below the
desired standard (IC50 > 30 μM). In addition, in the CV-dog
model, 6a was found to prolong QTc (>5% at 3 mpk).
Replacement of the cyclohexylamine with tetrahydropyran
(e.g., 7a) reduced the pKa of the primary amine from 8.6 to 7.3,
and the hERG selectivity improved accordingly (IC50 = 23
μM). In addition, pyran 7a is devoid of any QTc prolongation
in the CV dog model at doses up to 30 mg/kg iv. However, 7a
had stability concerns, producing an undesirable circulating
pyrrolopyrimidine upon dosing 7a in both rats (∼50%) and
dogs (∼30%) (Figure 2). The observed pyrrolopyrimidine
metabolite has weaker potency in DPP-4 (IC50 = 140 nM) and
Replacement of the left-hand-side trifluorophenyl group of
7b with other substituents resulted in reduced potency with the
exception of the difluorophenyl group which has comparable
DPP-4 inhibition activity.
Since the best analogue in the series 7b has three chiral
centers, all of the possible eight stereoisomers were made and
evaluated in the DPP-4 and counterassays. The 2R,3S,5R
isomer was found to be the most potent isomer. The trans-2,5-
difluorophenyl-3-amino group is essential for binding to the
DPP-4 protein active site, and the 2R,3S isomer fits better than
the 2S,3R isomer. Replacement of the 2-pyrazole hydrogen with
methylsulfonyl group gave omarigliptin (MK-3102), a potent,
selective analogue with a long duration of action amenable for
once-weekly human dosing. In preclinical pharmacokinetic
studies, the 1-H pyrazole has a shorter half-life compared to the
corresponding methylsulfone analogue.4c
As shown in Figure 4, sitagliptin (in yellow stick) and
fluoroomarigliptin (in green stick) bind very similarly in the
DPP-4 active site and share the same key interactions. The 2-F
B
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Drug Annotation
AUC) to 0.3 mg/kg (51% reduction). The efficacy of glucose
lowering in this model was similar to that achieved with
sitagliptin. In the corresponding pharmacodynamic (PD) assay,
omarigliptin-mediated plasma DPP-4 inhibition and plasma
compound concentrations were dose-dependent. At the 0.3
mg/kg dose (corresponding to maximum acute glucose
lowering efficacy), plasma DPP-4 activity was inhibited by
85% (uncorrected for assay dilution), which exceeds the target
inhibition (80%) associated with maximal glucose lowering
efficacy. The observed plasma DPP-4 inhibition was consistent
with the measured plasma inhibitor concentration (521 nM)
and the potency of the compound against murine plasma DPP-
4 (IC50 = 43.9 nM in 50% mouse plasma). In addition, the
administration of omarigliptin dose-dependently increased
plasma concentrations of active GLP-1 (GLP-1[7−36]amide
and GLP-1[7−37]) in this study, with the maximal increase in
active GLP-1 observed at the 0.3−1 mg/kg dosages. The
augmentation of active GLP-1 levels achieved at these doses
(>10-fold) was in the range of elevation in circulating hormone
observed in DPP-4-deficient (Dpp4−/−) mice (3- to 8-fold)
relative to wild type animals.12
Figure 3. Improvement of metabolic stability.
PHARMACOKINETICS (PK) IN PRECLINICAL
SPECIES
■
PK experiments were generally conducted as follows: All
species were fasted overnight before dosing, provided water ad
libitum, and fed 4 h following drug treatment. Blood was
collected at predetermined intervals for all species into EDTA-
containing tubes and centrifuged. Plasma was harvested and
stored at −70 °C until analysis.
Test compounds were typically formulated as solutions in
saline. Fasted male Sprague−Dawley rats were given either an
iv dose of test compound solution via a cannula implanted in
the femoral vein (n = 2) or a po dose by gavage (n = 3). Serial
blood samples were collected at 5 (iv only), 15, and 30 min and
at 1, 2, 4, 6, 8, 24, and 48 h postdose. Plasma was collected by
centrifugation, and plasma concentrations of test compound
were determined by LC−MS/MS following protein precip-
itation with acetonitrile.
Fasted dogs were administered intravenous doses via the
cephalic vein (dogs, n = 2). Oral doses were administered via
gastric gavage (n = 2). Serial blood samples were collected at 5
(iv only), 15, and 30 min and at 1, 2, 4, 6, 8, 24, 30, 48, and 72
h postdose. Plasma was collected by centrifugation, and plasma
concentrations of test compound were determined by LC−
MS/MS following protein precipitation with acetonitrile.
Pharmacokinetic parameters were calculated by established
noncompartmental methods.
The pharmacokinetics of omarigliptin in male Sprague−
Dawley rat and beagle dog were characterized by a low plasma
clearance (0.9−1.1 mL min−1 kg−1), a volume of distribution at
steady state of 0.8−1.3 L/kg, and a long terminal half-life
(∼11−22 h) (Table 1). The oral bioavailability of omarigliptin
was good in both dogs and rats (∼100%). The mean
percentage of unbound [3H]omarigliptin (1, 10, and 100
μM) in CD-1 mouse, Sprague−Dawley rat, beagle dog, and
human plasma was 38%, 15%, 43%, and 68%, respectively. The
blood-to-plasma concentration ratio in these species ranged
from 0.6 to 1.2.
Figure 4. Superposition of sitagliptin and fluoroomarigliptin in the
DPP-4 active site using their cocrystal structures of DPP-4 (PDB codes
1X70 and 4PNZ). The image was generated using PyMol.
atom on the trifluorophenyl group makes a H-bond with the
side chain of R125. The basic amine forms salt bridges with
E205 and E206, respectively. On the right-hand side, the fused
ring π−π stacks with the side chain of F347. One unique
interaction in sitagliptin is the H-bond between the amide
carbonyl and the side chain of Y547 through a water molecule.
IN VITRO PHARMACOLOGY
■
Omarigliptin is a competitive, reversible inhibitor of DPP-4
(IC50 = 1.6 nM, Ki = 0.8 nM) and is more potent than
sitagliptin (IC50 = 18 nM). It is highly selective over all
proteases tested (IC50 > 67 μM), including QPP, FAP, PEP,
DPP8, and DPP9. The compound has weak ion channel activity
(IC50 > 30 μM at IKr, Cav1.2, and Nav1.5). An expansive
selectivity counterscreen (168 radioligand binding or enzymatic
assays) was carried out at MDS Pharma. An IC50 > 10 μM was
obtained in all assays.
IN VIVO PHARMACOLOGY IN PRECLINICAL
SPECIES
■
Omarigliptin was evaluated for its ability to improve glucose
tolerance in lean mice. When orally administered 1 h prior to
dextrose challenge in an oral glucose tolerance test (OGTT), it
significantly reduced blood glucose excursion in a dose-
dependent manner from 0.01 mg/kg (7% reduction in glucose
Omarigliptin has a long half-life (rat, 11 h; dog, 22 h) and
lower clearance (rat, 1.1 mL min−1 kg−1; dog, 0.9 mL min−1
kg−1) in preclinical species. On the basis of the human PK
prediction, omarigliptin is projected to be amenable for once-
C
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Drug Annotation
cholesterol. The AUC(0−24h), Cmax, and Tmax were 5003 μM·h,
371 μM, and 2 h, respectively.
Table 1. Mean Pharmacokinetic Parameters of Omarigliptin
in Nonclinical Species
a
parameter
rat
dog
PHARMACEUTICAL PROPERTIES
■
dose iv/po (mg/kg)
1/2
1.1
1/2
0.9
Omarigliptin used for clinical trial is a white material.
Crystallinity was confirmed by optical microscopy and XRPD.
Differential scanning calorimetry (DSC) showed a melting
endotherm at 176.0 °C (heat of fusion, 89.68 J/g). The glass
transition temperature of the amorphous material was found to
be 58 °C. An anhydrous crystalline free base of omarigliptin is
chemically and physically stable at 40 °C/75% RH for up to 4
weeks. Omarigliptin was shown to be photostable as a bulk
material under 100 000 lx·h of cool white fluorescent light.
After a 24 h equilibration in aqueous buffer, the
concentration of omarigliptin is 7.1 mg/mL (pH 2), 8.7 mg/
mL (pH 6), and 3.1 mg/mL (pH 8). After a 24 h equilibration
of omarigliptin in buffer, the concentration of omarigliptin was
>20 mg/mL (pH 2−6) and 6.2 mg/mL at pH 8. Omarigliptin
has two pKa values measured at 3.5 and 7.1.
Cl (mL min−1 kg−1
Vdss (L/kg)
T1/2 (h)
)
0.8
1.3
11
22
Cmax (μM)
Tmax (h)
9.0
5.9
1.0
1.3
Foral (%)
∼100
47.8
∼100
54.0
nAUC (μM·h/(mg/kg))
a
Clp, plasma clearance; Vdss, volume of distribution at steady state;
T1/2, terminal half-life; Cmax, maximum plasma concentration following
oral dosing; Tmax, time to maximum concentration; Foral, oral
bioavailability; nAUC, dose-normalized area under the plasma
concentration vs time curve following oral dosing (1 mg/kg iv and
2.0 mg/kg po). Pharmacokinetic parameters were obtained following
administration of the TFA salt in saline (solution) to rats and beagle
dogs.
CLINICAL DATA
■
Dose-dependent efficacy in 2 h post meal glucose reduction,
fasting plasma glucose reduction, and HbA1c and safety profile
similar to that seen with once daily DPP-4 inhibitor such as
sitagliptin was observed in a 12-week phase IIB dose-range-
finding study.13
weekly dosing. This is recapitulated in the clinical studies,
where omarigliptin is shown to have a biphasic PK profile with
a terminal half-life of 120 h.
SAFETY PHARMACOLOGY
■
Omarigliptin is negative in the Ames mutagenicity assay.
In the PatchXpress cardiac ion channel panel, omarigliptin
exhibited minimal functional inhibition of hERG current up to
the highest tested concentration of 30 μM. In the nonfunctional
MK-499 displacement binding studies the compound had an
IC50 of >30 μM, and there were no remarkable effects on IKs,
INa, and ICaL up to 30 μM.
CHEMISTRY
■
The synthesis of omarigliptin 12b is illustrated in Schemes 1−3.
Omarigliptin is made by reductive amination of tetrahy-
dropyranone 8b with methylsulfonylpyrrolopyrazole 10 in the
presence of triacetoxyborohydride in dimethylacetal to give
intermediates 11, which was deprotected, neutralized with
ammonium hydroxide, and crystallized from ethyl acetate to
provide omarigliptin.
Omarigliptin was also evaluated in an exploratory 14-day oral
safety study in male rats at 100 mg kg−1 day−1. The compound
was well tolerated over the duration of the study, with no
mortality or physical signs noted. Clinical pathology findings
were limited to slight decreases in glucose, triglycerides, and
The tetrahydropyranone intermediates 8a and 8b were made
as follows. The nitroketone 15 was made by reacting aldehyde
13 with nitromethane in the presence of a catalytic amount of
Scheme 1. Synthesis of Omarigliptin
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Scheme 2. Synthesis of Tetrahydropyranones
Scheme 3. Synthesis of Pyrrolopyrazoles
sodium hydroxide as a base and oxidizing the resulting nitro
alcohol 14 with Jones or Dess−Martin reagent. Heating the
nitroketone 15 with 3-iodo-2-(iodomethyl)prop-1-ene gave the
pyran 16, which when reduced with sodium borohydride gave a
mixture of racemic trans-17 and racemic cis-18 isomers
separated by flash column chromatography. The cis-isomer 18
could easily be converted to the trans isomer 17 by treatment
with 1,8-diazabicyclo[5.4.0]undec-7-ene (DBU). The nitro-
substituted pyrans 17 was then reduced using zinc and
hydrochloric acid. The resulting amines were protected as
Boc derivatives by treatment with di-tert-butyl dicarbonate and
separated by chiral column chromatography to give 19a and
19b. Treatment of each of the two isomers of 19a and 19b with
osmium tetroxide and N-methylmorpholine N-oxide followed
by treatment with sodium periodate gave ketones 8a and 8b,
respectively.
The pyrrolopyrazole intermediate 9 was made by heating a
solution of the Boc-protected ketone 20 with N,N-dimethyl-
formamide−dimethylacetal (DMF−DMA) to form the enam-
ine 21, which was subsequently dissolved in ethanol and heated
under pressure with hydrazine and then heated with hydrogen
chloride in dry ethyl acetate. Neutralization with aqueous
sodium hydroxide gave the desired pyrrolopyrazole 9.
E
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(iodomethyl)prop-1-ene (4.8 g). The mixture was heated at 60 °C
for 2 h. After completion of reaction (monitored by LC−MS), the
contents were diluted with ethyl acetate (50.0 mL) and washed with
water (2 × 20 mL). Combined organic layers were dried over sodium
sulfate, filtered, and solvent was removed under reduced pressure. The
crude residue was purified by flash chromatography on a silica gel
column with hexane/ethyl acetate (80:20) as eluent to afford the
desired compound 16 (1.0 g, 30.0% yield). 1H NMR (300 MHz,
CDCl3) δ = 7.18−6.99 (m, 3 H), 5.40−5.24 (m, 2 H), 4.62 (s, 2 H),
3.61 (d, J = 1.3 Hz, 2 H). MS C12H9F2NO3. LC−MS: 254.5 (M + 1).
trans-5-Methylene-3-nitro-2-(2,5-difluorophenyl)-
tetrahydro-2H-pyran (17) and cis-5-Methylene-3-nitro-2-(2,5-
difluorophenyl)tetrahydro-2H-pyran (18). To a solution of 3-
methylene-5-nitro-6-(2,5-difluorophenyl)-3,4-dihydro-2H-pyran (1.0
g, 3.9 mmol) in chloroform (52.0 mL) and isopropyl alcohol (9.8
mL) were added silica gel (6.3 g) and sodium borohydride (0.553 g,
14.6 mmol), and the reaction mixture was stirred for 2 h at room
temperature. The reaction mixture was then quenched by addition of
hydrochloric acid (7.0 mL, 1.5 N) and filtered. The resulting solid
residue was washed with ethyl acetate (50.0 mL). The combined
filtrate was washed successively with saturated sodium bicarbonate
solution and brine, dried over anhydrous sodium sulfate and solvent
removed under reduced pressure. Flash chromatography afforded the
two diastereoisomers.
The methylsulfonylpyrrolopyrazole intermediate 10 was
made from the Boc-protected pyrrolopyrazole intermediate
22 by sequential treatment with sodium hydride, methane-
sulfonyl chloride, and deprotection with benzene sulfonic acid.
EXPERIMENTAL SECTION
■
General. All commercial chemicals and solvents are reagent grade
and were used without further purification unless otherwise specified.
1H NMR spectra were recorded on a Varian InNova 500 MHz
instrument in CDCl3 or CD3OD solutions unless otherwise noted.
Low-resolution mass spectra were determined on a Micromass
Platform liquid chromatography−mass spectrometer (LC−MS).
High-resolution mass spectra were acquired from a Micromass Q-
TOF quadrupole time-of-flight mass spectrometer. All MS experi-
ments were performed using electrospray ionization in positive ion
mode.
Liquid chromatography−mass spectrometry (LC−MS) involved
use of a Waters Xterrra MSC18 3.5 μm, 50 mm × 3.0 mm column
with a binary solvent system where solvent A = water, 0.06%
trifluoroacetic acid (by volume) and solvent B = acetonitrile, 0.05%
trifluoroacetic acid (by volume). The LC method used a flow rate of
1.0 mL/min with the following gradient: t = 0 min, 90% solvent A; t =
3.75 min, 2.0% solvent A; t = 4.75 min, 2% solvent A; t = 4.76 min,
90% solvent A; t = 5.5 min, 90% solvent. Purity determination was
performed by HPLC/DAD analysis under two conditions. Both
conditions used the Zorbax SB-18 (4.6 mm × 150 mm, 5 μm) at 40
°C, flow rate 1 mL/min. Mobile phase first condition was the
following: (A) 0.1% H3PO4, (B) ACN. Mobile phase second condition
was the following: (A) 10 mM potassium phosphate buffer pH 7, (B)
MeOH. All final compounds show a single peak (>95% purity) using
this analytical method.
1
Less polar diastereoisomer (17): 0.60 g, 59.5% yield; H NMR (400
MHz, CDCl3) δ = 7.18−7.11 (m, 1 H), 7.09−7.01 (m, 2 H), 5.16−
5.04 (m, 3 H), 4.82−4.72 (m, 1 H), 4.43−4.20 (m, 2 H), 3.13 (d, J =
8.2 Hz, 2 H). MS C12H11F2NO3. LC−MS: 254.4 (M − 1).
More polar diastereoisomer (18): 0.30 g, 29.8% yield; 1H NMR (300
MHz, CDCl3) δ = 7.18 (ddd, J = 2.9, 5.6, 8.6 Hz, 1 H), 7.08−6.92 (m,
2 H), 5.15 (s, 1 H), 5.12−5.01 (m, 3 H), 4.62 (d, J = 13.0 Hz, 1 H),
4.31 (d, J = 12.8 Hz, 1 H), 3.13−2.85 (m, 2 H). MS C12H11F2NO3.
LC−MS: 254.4 (M − 1).
1-(2,5-Difluorophenyl)-2-nitroethanol (14). To sodium hydrox-
ide (1 N, 42.8 mL) and methanol (20.0 mL) at 5 °C was added a
solution of 2,5-difluorobenzaldehyde (5.0 g, 35.18 mmol) and
nitromethane (2.26 mL, 41.4 mmol) in methanol (5.0 mL) dropwise
over a period of 1 h. The reaction mixture was then neutralized with
glacial acetic acid (3.0 mL). The mixture was diluted with water (150
mL) and extracted with dichloromethane (2 × 100 mL). Combined
organic extracts were dried over anhydrous sodium sulfate and solvent
was removed under reduced pressure to give the desired nitro alcohol
tert-Butyl [(2S,3R)-5-Methylene-2-(2,5-difluorophenyl)-
tetrahydro-2H-pyran-3-yl]carbamate (19a) and tert-Butyl
[(2R,3S)-5-Methylene-2-(2,5-difluorophenyl)tetrahydro-2H-
pyran-3-yl]carbamate (19b). To a vigorously stirred suspension of
nitro compound 17 (1.0 g, 3.92 mmol) and zinc powder (2.99 g, 45.88
mmol) in ethanol (20.0 mL) was added acetic acid (4.58 mL, 74.5
mmol). After 1 h of stirring, the contents were filtered through Celite
pad and the pad was washed with ethanol (10.0 mL). The filtrate was
adjusted to pH 10 using aqueous sodium hydroxide solution (2 N, 7.0
mL) and extracted with dichloromethane (2 × 30 mL). The organic
layer was washed with saturated brine and dried over anhydrous
sodium sulfate, and solvent was removed under reduced pressure to
furnish the crude amine (1.2 g) which was used for next step without
further purification. To a solution of amine in dichloromethane (20.0
mL) was added di-tert-butyl dicarbonate (1.75 g, 8.0 mmol), and the
mixture was stirred for 20 h at room temperature. After completion of
reaction (as monitored by TLC), the mixture was diluted with
dichloromethane (50.0 mL) and the combined organic layers were
washed with water (2 × 20.0 mL) and dried over anhydrous sodium
sulfate. The solution was evaporated under reduced pressure. The
crude product was purified by flash chromatography on a silica gel
column with hexane/ethyl acetate (85:15) to furnish racemic product
19. Compound 19 was further resolved by employing preparative
HPLC conditions given below using Chiralpak IA (250 mm × 4.6
mm) column.
1
14 (7.2 g, 100% yield). H NMR (400 MHz, CDCl3) δ = 7.37−7.30
(m, 1 H), 7.12−7.00 (m, 2 H), 5.74 (dd, J = 1.8, 9.4 Hz, 1 H), 4.70−
4.50 (m, 2 H). MS C8H7F2NO3. LC−MS: 202.4 (M − 1).
2-Nitro-1-(2,5-difluorophenyl)ethanone (15). A solution of
Dess−Martin periodinane (17.7 g, 41.73 mmol) in dichloromethane
(100 mL) was added to a solution of the nitro alcohol 14 (6.3 g, 31.0
mmol) at 10 °C over a period of 30 min. Stirring was continued for 2
h, and the reaction mixture was then poured onto a mixture of sodium
bicarbonate (50 g) and sodium thiosulfate (50 g) in water (500 mL).
The desired product was extracted with dichloromethane (2 × 200
mL). The combined organic layers were dried over anhydrous sodium
sulfate. The contents were filtered, and solvent was removed under
reduced pressure. The residue was purified by flash chromatography
on a silica gel column with hexane/ethyl acetate (75:25) as eluent to
1
afford the desired nitroketone 15 (5.0 g, 80.2% yield). H NMR (400
MHz, CDCl3) δ = 7.73 (ddd, J = 3.2, 5.2, 8.1 Hz, 1 H), 7.43−7.35 (m,
1 H), 7.29−7.18 (m, 1 H), 5.81 (d, J = 3.2 Hz, 1 H), 4.79 (br s, 1 H).
MS C8H5F2NO3. LC−MS: 200.4 (M − 1).
Compound 19a: 0.28 g, 22.0% yield. tR = 5.28 min (Chiralpak IA,
hexane/ethanol (90:10), flow rate 1.0 mL/min). 1H NMR (400 MHz,
CDCl3) δ = 7.30−7.18 (m, 2 H), 6.96 (br s, 2 H), 4.96 (d, J = 10.1 Hz,
2 H), 4.49 (d, J = 7.5 Hz, 1 H), 4.32 (d, J = 12.3 Hz, 1 H), 4.08 (d, J =
12.8 Hz, 1 H), 3.71 (br s, 1 H), 2.85 (d, J = 9.6 Hz, 1 H), 2.29 (t, J =
11.5 Hz, 1 H), 1.38−1.13 (m, 9 H). MS C17H21F2NO3. LC−MS:
3-Methylene-5-nitro-6-(2,5-difluorophenyl)-3,4-dihydro-2H-
pyran (16). A mixture of 3-chloro-2-(chloromethyl)prop-1-ene (1.85
mL, 16.0 mmol) and sodium iodide (13.19 g, 88.0 mmol) in acetone
(10.0 mL) was stirred at room temperature for 20 h. The solvent was
removed under reduced pressure, and the contents were partitioned
between dichloromethane (150.0 mL) and water (50.0 mL). The
organic layer was dried over sodium sulfate, filtered, and evaporated to
yield 3-iodo-2-(iodomethyl)prop-1-ene (4.8 g) as a reddish oil.
N,N-Diisopropylethylamine (5.1 mL, 30.86 mmol) was added to a
solution of 2-nitro-1-(2,5-difluorophenyl)ethanone (2.67 g, 13.27
mmol) in N,N-dimethylformamide (5.0 mL) and 3-iodo-2-
23
226.2 (M − Boc). Optical rotation [α]D +0.6° (c 0.95, CHCl3)
Compound 19b: 0.3 g, 23.5% yield. tR = 8.69 min (Chiralpak IA,
hexane/ethanol (90:10), flow rate 1.0 mL/min). 1H NMR (400 MHz,
CDCl3) δ = 7.29−7.18 (m, 1 H), 6.96 (br s, 2 H), 4.96 (d, J = 10.7 Hz,
3 H), 4.49 (d, J = 8.0 Hz, 1 H), 4.32 (d, J = 12.3 Hz, 1 H), 4.08 (d, J =
12.8 Hz, 1 H), 3.71 (br s, 1 H), 2.84 (d, J = 10.1 Hz, 1 H), 2.29 (t, J =
F
dx.doi.org/10.1021/jm401992e | J. Med. Chem. XXXX, XXX, XXX−XXX

Journal of Medicinal Chemistry
Drug Annotation
11.7 Hz, 1 H), 1.37−1.14 (m, 9 H). MS C17H21F2NO3. LC−MS:
(23b). A suspension of N-Boc-pyrazolopyrolidine 22 (27.2 g, 130
mmol) in anhydrous acetonitrile (1.0 L) was charged in a 2.0 L three-
neck flask fitted with a thermometer and an addition funnel and then
treated with sodium hydride (60% dispersion in oil, 6.23 g, 156 mmol)
while under a nitrogen atmosphere in one portion. The reaction
mixture was stirred at room temperature for 2 h. The resulting white
suspension was then cooled in an ice bath, and methanesulfonyl
chloride (25.2 mL, 324 mmol) was slowly added via addition funnel.
The ice bath was then removed, and the mixture was stirred for 1 h at
room temperature. The reaction mixture was quenched with water
(500 mL), and the layers were separated. The aqueous layer was then
extracted with dichloromethane (2 × 500 mL). The combined organic
layers were dried over sodium sulfate and concentrated under reduced
pressure to give a mixture of products 2-methylsulfonyl 23a and 1-
methylsulfonyl substituted analogues 23b as colorless syrups. NMR in
CD3OD indicated a 1:1 mixture of two products, in which the proton
on the pyrazole ring in product 23a appeared at 7.70 ppm while the
proton in product 23b appeared at 7.95 pm. Yield: 37.3 g, 100%. LC−
MS: 288.08 (M + 1).
23
226.2 (M − Boc). Optical rotation [α]D −0.6° (c 0.96, CHCl3).
tert-Butyl [(2S,3R)-5-Oxo-2-(2,5-difluorophenyl)tetrahydro-
2H-pyran-3-yl]carbamate (8a). To a solution of 19a (0.25 g,
0.769 mmol) in 1,4-dioxane (8.0 mL) was added a solution of sodium
metaperiodate (0.49 g, 2.31 mmol dissolved in 8.0 mL of water)
followed by addition of osmium tetroxide (0.095 mL, 1 M solution in
water) while maintaining the temperature at 0 °C. Stirring was
continued for 2 h. The mixture was diluted with ethyl acetate (10.0
mL) and washed with water (2 × 5.0 mL). Organic layer was dried
over anhydrous sodium sulfate. The solvent was removed under
reduced pressure and crude product purified by flash chromatography
(silica gel, gradient 20% ethyl acetate in hexanes) to yield the product
as a white solid (0.12 g, 47.7% yield). 1H NMR (400 MHz, CDCl3) δ
= 7.23 (d, J = 3.2 Hz, 1 H), 7.08−6.96 (m, 2 H), 4.85 (d, J = 7.5 Hz, 1
H), 4.64 (br s, 1 H), 4.35−4.26 (m, 1 H), 4.17−4.02 (m, 2 H), 3.06
(dd, J = 5.3, 16.5 Hz, 1 H), 2.76 (d, J = 10.7 Hz, 1 H), 1.39−1.19 (m, 9
H). MS C16H19F2NO4. LC−MS: 228.3 (M − Boc). Optical rotation
25
[α]D −2.63° (c 1.12, CHCl3).
2-(Methylsulfonyl)-2,4,5,6-tetrahydropyrrolo[3,4-c]pyrazole
(10). In a 300 mL flask was placed 16.18 g (56.3 mmol) of the Boc-
protected mesylated pyrazoles (23a and 23b) and 110 mL of isopropyl
acetate. To the solution was added 16.0 g (101.4 mmol) of
benzenesulfonic acid in 40 mL of isopropyl acetate. The resulting
solution was heated to 30 °C for 2 h and cooled to room temperature
and the slurry stirred for 14 h. The slurry was filtered and washed with
40 mL of isopropyl acetate and dried under vacuum/N2 sweep for 6 h
tert-Butyl [(2R,3S)-5-Oxo-2-(2,5-difluorophenyl)tetrahydro-
2H-pyran-3-yl]carbamate (8b). This compound was made from
1
19b by following the same method described for intermediate 8a. H
NMR (400 MHz, CDCl3) δ = 7.23 (d, J = 3.2 Hz, 1 H), 7.08−6.96 (m,
2 H), 4.85 (d, J = 7.5 Hz, 1 H), 4.64 (br s, 1 H), 4.34−4.26 (m, 1 H),
4.17−4.01 (m, 2 H), 3.06 (dd, J = 5.3, 16.5 Hz, 1 H), 2.76 (d, J = 11.2
Hz, 1 H), 1.38−1.19 (m, 9 H). MS C16H19F2NO4. LC−MS: 228.3 (M
25
− Boc). Optical rotation [α]D +5.49° (c 1.05, CHCl3).
1
to afford 10 (19.5 g, 99%) as a colorless solid: H NMR (DMSO-d6,
tert-Butyl (3Z)-3-[(Dimethylamino)methylene]-4-oxopyrroli-
dine-1-carboxylate (21). A solution of tert-butyl 3-oxopyrrolidine-1-
carboxylate 20 (40 g, 216 mmol) was treated with DMF−DMA (267
g, 2241 mmol) and heated at 105 °C for 40 min. The solution was
cooled and evaporated under reduced pressure, and the resulting
orange solid was treated with hexane (200 mL) and cooled in a
refrigerator for 3 days. The resulting brownish-yellow solid obtained as
such was collected by filtration, dried, and used in the next step
without further purification.
400 mHz) δ = 9.80 (s, 2H), 8.13 (s, 1H), 7.63−7.57 (m, 2H), 7.35−
7.28 (m, 2H), 4.43 (s, 2H), 4.36 (s, 2H), 3.58 (s, 3H). LC−MS:
188.20 (M + 1).
(2R,3S,5R)-2-(2,5-Difluorophenyl)-5-[2-(methylsulfonyl)-2,6-
dihydropyrrolo[3,4-c]pyrazol-5(4H)-yl]tetrahydro-2H-pyran-3-
amine, Omarigliptin. In a 500 mL three-neck flask equipped with a
mechanical stirrer, nitrogen inlet, and a thermocouple were placed 8.25
g (25.2 mmol) of the ketone 8b, 9.80 g (28.4 mmol) of the pyrazole
salt 10, and 124 mL of dimethylacetamide. The resulting
homogeneous solution was cooled to −10 °C, and 6.94 g (32.7
mmol) of sodium triacetoxyborohydride was added portionwise. The
resulting mixture was stirred for 2 h and quenched by the slow
addition of a mixture of NH4OH (8.3 mL) and water (16.5 mL). The
slurry was heated to 50 °C and then cooled to 22 °C and filtered. The
wet cake was washed with 5:1 dimethylacetamide/water (65 mL) and
then water (65 mL) and dried under vacuum/N2 sweep to afford (10.6
g, 94%) of the title compound as an off white solid (11). LC−MS:
499.10 (M + 1). The product was used in the next step without further
purification.
In a 200 mL three-neck jacketed flask equipped with a mechanical
stirrer, nitrogen inlet, and thermocouple were placed 10.35 g (20.8
mmol) of the above compound (11), 31 mL of dimethylacetamide,
and 42 mL of water. To the resulting slurry was added 12.2 mL of
H2SO4 in 21 mL of water over 3.5 h, and the slurry was stirred for 15 h
at room temperature. To the reaction mixture was added NH4OH
until the pH was 10.2. The slurry was filtered and the wet cake was
washed with water (2 × 17.5 mL) and dried under vacuum/N2 sweep
to give the title compound as a colorless solid (6.73 g, 89%).
Crystallization from ethyl acetate gave a compound with greater than
99% purity. Optical rotation [α]D20 −12.0° (c 1.0, CH3OH). 1H NMR
(CD3OD, 500 MHz) δ = 1.71 (q, 1H, J = 12 Hz), 2.56−2.61 (m, 1H),
3.11−3.18 (m, 1H), 3.36−3.40 (m, 1H), 3.48 (t, 1H, J = 12 Hz),
3.88−3.94 (m, 4H), 4.30−4.35 (m, 1H), 4.53 (d, 1H, J = 12 Hz),
7.14−7.23 (m, 2H), 7.26−7.30 (m, 1H), 7.88 (s, 1H). LC−MS:
399.04 (M + 1).
1,4,5,6-Tetrahydropyrrolo[3,4-c]pyrazole (9). A solution of
hydrazine (3 mL) and tert-butyl (3Z)-3-[(dimethylamino)methylene]-
4-oxopyrrolidine-1-carboxylate 21 obtained above (19.2 g) in ethanol
(40 mL) was heated at 85 °C in a sealed tube for 4 h. Solvent was
removed under reduced pressure, and the residue was triturated with
dichloromethane (160 mL) and ethyl acetate (15 mL). The resulting
solid was filtered. The filtrate was concentrated, and the resulting solid
was triturated again and filtered. The combined solids were treated
with 4 N hydrochloric acid (250 mL) in methanol and stirred for 6 h.
The reaction mixture was concentrated and dried. The resulting solid
was treated again for 6 h with 4 N hydrochloric acid (250 mL) in
methanol. After concentration and drying, the resulting hydrochloride
salt was treated with ammonia in methanol (2 N, 300 mL) and
ammonium hydroxide solution in water (28%, 30 mL) and
concentrated to dryness. The solid obtained was treated with methanol
(70 mL) and water (5 mL) and purified in three batches on Biotage
Horizon system (silica, gradient 5−17% methanol containing 10%
concentrated ammonium hydroxide in ethyl acetate) to yield 1,4,5,6-
tetrahydropyrrolo[3,4-c]pyrazole 9: 7.0 g, 62.2%. 1H NMR (500 MHz,
CD3OD): δ = 4.04 (d, 4H), 7.39 (s, 1H). LC−MS: 109.85 (M + 1)
N-Boc-pyrazolopyrrolidine (22). To a suspension of 1,4,5,6-
tetrahydropyrrolo[3,4-c]pyrazole 9 (41.0 g) in dichloromethane (1.0
L) was added triethylamine (147 mL) followed by addition of liquid
(Boc)2O (95 mL) via an addition funnel over a period of 45 min. The
mixture was stirred at room temperature overnight, transferred to a
separation funnel, and washed successively with 2.0 N HCl (3 × 500
mL), 10% NaOH (2 × 500 mL), and brine (400 mL), dried over
anhydrous sodium sulfate, and evaporated to give clean 22 as a brown
solid (82.1 g, 96.1%). The material is ready for the next step without
purification. 1H NMR (500 MHz, CD3OD): δ 1.51 (s, 9H), 4.39−4.42
(m, 4H), 7.42 (d, 1H, J = 12 Hz). LC−MS: 210.10 (M + 1).
tert-Butyl 1-(Methylsulfonyl)]-4,6-dihydropyrrolo[3,4-c]-
pyrazole-5(1H)-carboxylate (23a) and tert-Butyl 2-(Methylsul-
fonyl)]-2,6-dihydropyrrolo[3,4-c]pyrazole-5(4H)-carboxylate
AUTHOR INFORMATION
■
Corresponding Author
*Phone: 732-594-6514. E-mail: tesfaye_biftu@merck.com.
Notes
The authors declare no competing financial interest.
G
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Journal of Medicinal Chemistry
Drug Annotation
(7) Collins, P. J.; McMahon, G.; O’Brien, P.; O’Connor, B.
Purification, identification and characterisation of seprase from bovine
serum. Int. J. Biochem. Cell Biol. 2004, 36, 2320−2333.
ACKNOWLEDGMENTS
■
We acknowledge Eric Streckfuss and Dorothy Levorse for
purification and analysis of intermediates, respectively. We also
thank Dr. Phil Eskola, Mark Levorse, Bob Frankshun, Amanda
Makarewicz, Daniel Kim for scale-up of key intermediates used
for the synthesis of omarigliptin and other closely related
analogues.
(8) Abbott, C. A.; Yu, D. M.; Woollatt, E.; Sutherland, G. R.;
McCaughan, G. W.; Gorrell, M. D. Cloning, expression and
chromosomal localization of a novel human dipeptidyl peptidase
(DPP) IV homolog, DPP8. Eur. J. Biochem. 2000, 267, 6140−6150.
(9) Scanlan, M. J.; Raj, B. K.; Calvo, B.; Garin-Chesa, P.; Sanz-
Moncasi, M. P.; Healey, J. H.; Old, L. J.; Retting, W. J. Molecular
cloning of fibroblast activation protein alpha, a member of the serine
protease family selectively expressed in stromal fibroblasts of epithelial
cancers. Proc. Natl. Acad. Sci.U.S.A. 1994, 91, 5657−5661.
(10) Olsen, C.; Wagtmann, N. Identification and characterization of
human DPP9, a novel homologue of dipeptidyl peptidase IV. Gene
2002, 299, 185−195.
(11) Lankas, G. R.; Leiting, B.; Roy, R. S.; Wiermann, G. J.; Beconi,
M. G.; Biftu, T.; Chan, C.; Edmondson, S.; Feeney, W. P.; He, H.;
Ippolito, D. E.; Kim, D.; Lyons, K. A.; Ok, H. O.; Patel, R. A.; Petrov,
A. N.; Pryon, K. A.; Qian, X.; Reigle, L.; Woods, A.; Wu, J. K.; Zaller,
D.; Zhang, X.; Zhu, L.; Weber, A. E.; Thornberry, N. A. Dipeptidyl
peptidase IV inhibition for the treatment of type 2 diabetes: potential
importance of selectivity over dipeptidyl peptidases 8 and 9. Diabetes
2005, 54, 2988−2944.
(12) Conarello, S. L.; Li, Z.; Ronan, J.; Sinha Roy, R.; Zhu, L.; Jiang,
G.; Liu, F.; Woods, J.; Zycband, E.; Moller, D. E.; Thornberry, N. A.;
Zhang, B. B. Mice lacking dipeptidyl peptidase IV are protected against
obesity and insulin resistance. Proc. Natl. Acad. Sci. U.S.A. 2003, 100,
6825.
(13) Gantz, I.; Chen, M.; Mirza, A.; Suryawanshi, S.; Davies, M. J.;
Goldstein, B. J. Effect of MK-3102, a novel once-weekly DPP-4
inhibitor, over 12 weeks in patients with type 2 diabetes mellitus.
Presented at the 48th Annual Meeting of the European Association for
the Study of Diabetes (EASD), Berlin, Germany, October 2012;
Abstract 101 (Clinical Research, Metabolism, Merck Research
Laboratories).
ABBREVIATIONS USED
■
DPP-4, dipeptidyl peptidase 4; GLP-1, glucacon-like peptide 1;
GIP, glucose-dependent insulinotropic polypeptide; DPP8,
dipeptidyl peptidase 8; DPP9, dipeptidyl peptidase 9; FAP,
seprase or fibroblast activation protein; QPP, quiescent cell
proline dipeptidase; DPPII, dipeptidyl peptidase II; APP,
aminopeptidase P; F, oral bioavailability; val-pyr, valine
pyrrolidide; SAR, structure−activity relationship; MRT, mean
residence time; hERG, human ether-a-go-go related gene,
which is a human cardiac potassium ion channel; OGTT, oral
glucose tolerance test; PD, pharmacodynamic; AMC, amino-
methylcoumarin; DIO, diet-induced obese; AUC, area under
the curve; Clp, clearance; DAD, diode array detector
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