PYRAZOLE COMPOUNDS
This invention relates to pyrazole compounds or pharmaceutically acceptable salts
thereof. Compounds of this invention are inhibitors of methionine aminopeptidase 2
(MetAP2) and dipeptidyl peptidase-4 (DPP-4).
MetAP2 is a metalloproteinase that cleaves initiator methionine from nascent
peptide emerging from the ribosomes. WO 2010/065879 reports small molecule MetAP2
inhibitors for obesity treatment.
DPP-4 inhibitors are an established drug class to improve glycemic control in
patients with type 2 diabetes mellitus. Compounds with dual inhibitory activity in both
MetAP2 and DPP-4 are desired.
The present invention provides novel compounds with dual MetAP2 and DPP-4
inhibitition. These dual inhibitor compounds can be useful in the treatment of a MetAP2
and DPP-4 mediated condition.
The present invention provides a compound of the Formula I
or a pharmaceutically acceptable salt thereof.
In an embodiment of the invention, the compound is
or a pharmaceutically acceptable salt thereof.
In an embodiment of the invention the compound is (3R,4S)-l-[5-[[3-(4,4-
Difluorocyclohexen-l-yl)-lH-pyrazol-4-yl]oxy]pyrimidin-2-yl]-4-(2,4,5-
trifluorophenyl)pyrrolidin-3 -amine.
The invention provides a pharmaceutical composition comprising a compound of
Formula I, or a pharmaceutically acceptable salt thereof, and at least one selected from
the group consisting of a pharmaceutically acceptable carrier, diluent, and excipient.
The invention provides a method for treating type II diabetes in a mammal in need
thereof, comprising administering to the mammal an effective amount of a compound of
Formula I, or a pharmaceutically acceptable salt thereof. The invention provides a
method for treating obesity in an mammal in need thereof, comprising administering to
the mammal an effective amount of a compound of Formula I. In another embodiment, a
compound of Formula I, or a pharmaceutically acceptable salt thereof for use in therapy.
Further, provided is a compound of Formula I, or a pharmaceutically acceptable salt
thereof, for use in the manufacture of a medicament.
Compounds of the present invention can be provided as a pharmaceutically
acceptable salt. "Pharmaceutically-acceptable salt" refers to salts of the compound of the
invention considered to be acceptable for clinical and/or veterinary use. Pharmaceutically
acceptable salts and common methodology for preparing them are well known in the art.
See, e.g., P. Stahl, etal., Handbook of Pharmaceutical Salts: Properties, Selection and
Use, (VCHA/Wiley-VCH, 2002); S.M. Berge, et al, "Pharmaceutical Salts," Journal of
Pharmaceutical Sciences, Vol. 66, No. 1, January 1977.
Additionally, certain intermediates described in the following preparations may
contain one or more nitrogen protecting groups. The variable protecting group may be
the same or different in each occurrence depending on the particular reaction conditions
and the particular transformations to be performed. The protection and deprotection
conditions are well known to the skilled artisan and are described in the literature (See for
example "Greene's Protective Groups in Organic Synthesis", Fourth Edition, by Peter
G.M. Wuts and Theodora W. Greene, John Wiley and Sons, Inc. 2007).
Individual isomers, enantiomers, and diastereomers may be separated or resolved
by one of ordinary skill in the art at any convenient point in the synthesis of compounds
of the invention, by methods such as selective crystallization techniques or chiral
chromatography (See for example, J. Jacques, et al., "Enantiomers, Racemates, and
Resolutions" , John Wiley and Sons, Inc., 1981, and E.L. Eliel and S.H. Wilen,"
Stereochemistry of Organic Compounds", Wiley-Interscience, 1994).
The compounds of the present invention, or salts thereof, may be prepared by a
variety of procedures known in the art, some of which are illustrated in the Preparations
and Examples below. The specific synthetic steps for each of the routes described may be
combined in different ways, or in conjunction with steps from different schemes, to
prepare compounds of the invention, or salts thereof. The products of each step in the
schemes below can be recovered by conventional methods well known in the art,
including extraction, evaporation, precipitation, chromatography, filtration, trituration,
and crystallization. In the schemes below, all substituents unless otherwise indicated, are
as previously defined. The reagents and starting materials are readily available to one of
ordinary skill in the art. Others may be made by standard techniques of organic and
heterocyclic chemistry which are analogous to the syntheses of known structurally-similar
compounds and the procedures described in the Preparations and Examples which follow
including any novel procedures.
The abbreviations used herein are defined according to Aldrichimica Acta, Vol.
17, No. 1, 1984. Other abbreviations are defined as follows: "AMC" refers to 7-amido-4-
methylcoumarin hydrobromide; "BSA" refers to Bovine Serum Albumin; "DIO" refers to
diet induced obese; "EDTA" refers to ethylenediaminetetraacetic acid; "HEC" refers to
hydroxy ethyl cellulose; "HEPES" refers to 4-(2-hydroxyethyl)-lpiperazineethanesulfonic
acid; "HFD" refers to high fat diet; "HOAc" refers to acetic
acid; "IC 50" refers to the concentration of an agent that produces 50% of the maximal
inhibitory response possible for that agent; "MeOH" refers to methyl alcohol or methanol;
"THF" refers to tetrahydrofuran and "Tris" refers to tris(hydroxymethyl)aminomethane.
Preparation 1
2-(2-Chloropyrimidin-5-yl)oxyacetonitrile
To a mixture of bromoacetonitrile (2.94 mL, 42.18 mmol), 2-chloropyrimidin-5-ol
(5 g, 38.30 mmol) in CH3CN (100 mL) is added potassium carbonate (7.94 g, 57.45
mmol). The reaction mixture is heated to 60 °C for 2 hours. After cooling to room
temperature, the solid is filtered and the filtrate is concentrated under reduced pressure.
The residue is purified by silica gel flash chromatography eluting with a gradient of 100%
ether to 1/1 ether/ethyl acetate to give the title compound (6 g, 35.38 mmol, 92.37%) as a
white solid. Mass spectrum (m/z): 170 (M+H).
Preparation 2
2-(2-Chloropyrimidin-5-yl)oxy-3-oxo-propanenitrile
To a solution of 2-(2-chloropyrimidin-5-yl)oxyacetonitrile (6.00 g, 35.4 mmol)
and ethyl formate (26.2 g, 354 mmol) in 1,2-dimethoxy ethane ( 200 mL, 1930 mmol) is
added potassium i30 , demonstrating selective MetAP2 inhibition as
compared with MetAPl.
Enzymatic activity assay of DPP-4
Human DPP-4 ((39-766)-His) and mouse DPP-4 ((29-760)-His) are purified for
use in the assay. The final concentration of hDPP-4 and mDPP-4 in the assay is 0.04 nM
and 0.22 nM respectively.
Inhibition of the catalytic activity of human and mouse DPP-4 by the compound
in the present invention is monitored by the formation of product fluorescence AMC (7-
amido-4-methylcoumarin hydrobromide) from substrate Gly-Pro-AMC (Sigma, G2761)
on an Envision plate reader. The reaction is typically conducted by incubating the
enzyme, test compound, and substrate (10 ) in a 75 ΐ assay buffer (0.01% BSA, 0.1
mM EDTA, 50 Tris-HCl, 0.01% Triton™-X100, 0.1 M NaCl at pH 7.5) for 30
minutes. After the reaction is stopped by the addition of 25 ΐ ZnS0 4 (10 mM), the
formation of fluorescent product AMC is measured on an Envision plate reader with the
excitation wavelength at 355 nm and emission wavelength at 460 nm. The IC50 value is
calculated typically from a 10-point dose titration curve using the 4-parameter logistic
equation.
The IC50 for Example 1 is lower than 1000 nM in the human and mouse DPP-4
assay and the results are shown in Table 1.
Table 1. Enzymatic activity of Example 1.
Mean + SEM (n); SEM = standard error of the mean; n = number of determination.
Therapeutic Weight Loss Effect Measurement of Compounds
To determine the therapeutic weight loss effects and improvement of metabolic
parameters, the compound from the invention is tested in the high fat diet (HFD) feeding
induced obese mouse model (DIO mice). In this model, C57/B16J male mouse is fed with
the 60% HFD (D12492i, Research Diets) for 16 ~ 28 weeks to establish obesity with
body weight reaching around 50 g. The mice will gradually increase the body weight to
about 50 g and maintain that weight in this obese state. Test compound (via the vehicle
of 0.5% HEC plus 0.25% Tween®-80 at 5 mL/kg) is administered orally to the obese
DIO mice once or twice daily throughout the study duration. The dose-dependent weight
loss of obese DIO mice for Example 1 of the oral treatment at 20 mg/kg and 60 mg/kg
once daily is about 4% and 17% weight loss compared to the vehicle group at day 14,
respectively. The data support that the compound of Example 1 is associated with desired
weight loss and could offer a therapeutic weight loss effect.
DPP-4 pharmacodynamics Assay in Mouse
To determine the in vivo DPP-4 inhibition by MetAP2 plus DPP-4 dual inhibitor
compounds, C57B/L6 lean mice are administrated with the compound in fed states and
then DPP-4 target engagement in plasma is measured.
Animals are weighed and randomized by body weight. Each mouse is dosed via
oral gavage with vehicle or testing compound formulated with vehicle for up to 3 times.
The first dose is administrated at 9-10 am on day 1. The second dose is administrated at
16:30-17:30 on day 1. The third dose is administrated at 9-10 am on day 2. The mice
are fasted for 6 hours after the last dose before termination at - 3 pm on day 2. Blood
samples are collected at 1 hour after the first dosing and upon termination. EDTA-K2 at
final concentration of 5 mM is used as an anticoagulant. Plasma, isolated from the blood
samples, is used to determine the plasma DPP-4 enzyme activity.
Plasma DPP-4 enzyme activity in the present invention is monitored by the
formation rate of fluorescence AMC from substrate Gly-Pro-AMC (Sigma, G2761) via
Envision plate reader. The reaction is typically conducted by incubating the plasma (20
ΐ) and substrate (10 ) in a 40 assay buffer (0.01% BSA, 0.1 mM EDTA, 50 
Tris-HCl, 0.01% Triton™-X100, 0.1 M NaCl at pH 7.5). Fluorescence signal is read
immediately after the start of the reaction in kinetic model in Envision plate reader. The
excitation wavelength is set at 355 nm and emission wavelength is set at 460 nm. The
plasma DPP-4 activity is calculated from reaction velocity. The percentage plasma DPP-
4 inhibition is normalized against plasma DPP-4 activity in the vehicle group, which is set
as 0% inhibition.
The plasma DPP-4 inhibition for Example 1 under the assay condition is 86% and
83% for 1 hour after the first dosing and upon termination, respectively. The data support
that the compound of Example 1 is associated with desired DPP-4 inhibition that could
yield therapeutic glycemic control.
WE CLAIM:
1. A compound of the Formula
or a pharmaceutically acceptable salt thereof.
or a pharmaceutically acceptable salt thereof.
3. A compound or salt as claimed by Claim 1 wherein the compound is (3R,4S)-1-
[5-[[3-(4,4-Difluorocyclohexen-l-yl)-lH-pyrazol-4-yl]oxy]pyrimidin-2-yl]-4-(2,4,5-
trifluorophenyl)pyrrolidin-3-amine.
4. A pharmaceutical composition comprising a compound as claimed by any one of
Claims 1 to 3, or a pharmaceutically acceptable salt thereof, and at least one of a
pharmaceutically acceptable carrier, diluent, or excipient.
5. A method for treating type II diabetes in a mammal in need thereof, comprising
administering to the mammal an effective amount of a compound, or a pharmaceutically
acceptable salt thereof, as claimed by any one of Claims 1 to 3.
6. A method for treating obesity in a mammal in need thereof, comprising
administering to the mammal an effective amount of a compound, or pharmaceutically
acceptable salt thereof, as claimed by any one of Claims 1 to 3.
7. A compound, or a pharmaceutically acceptable salt thereof, as claimed by any one
of Claims 1 to 3 for use in the manufacture of a medicament.