Process Development for the Synthesis of a Selective M1 and M4 Muscarinic Acetylcholine Receptors Agonist

Article abstract of DOI:10.1021/acs.oprd.7b00236

A practical and chromatography-free synthetic process to selective M₁ and M₄ muscarinic acetylcholine receptors agonist was developed and demonstrated on a several hundred gram scale. The key feature of this route is N,N-dimethylcarb

Full text of DOI:10.1021/acs.oprd.7b00236

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Full Paper
Process Development for the Synthesis of a Selective
M1 and M4 Muscarinic Acetylcholine Receptors Agonist
Yoshiharu Uruno, Kazuki Hashimoto, Yoichi Hiyama, and Takaaki Sumiyoshi
Org. Process Res. Dev., Just Accepted Manuscript • DOI: 10.1021/acs.oprd.7b00236 • Publication Date (Web): 05 Sep 2017
Downloaded from http://pubs.acs.org on September 5, 2017
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Process Development for the Synthesis of a Selective M1
and M4 Muscarinic Acetylcholine Receptors Agonist
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Yoshiharu Uruno,†, * Kazuki Hashimoto,‡ Yoichi Hiyama,§ and Takaaki Sumiyoshi†,¶
†Chemistry Research Laboratory, Sumitomo Dainippon Pharma Co. Ltd., 3ꢀ1ꢀ98, Kasugadeꢀnaka,
Konohanaꢀku, Osaka 554ꢀ0022, Japan
‡Process Chemistry Research & Development Laboratories, Sumitomo Dainippon Pharma Co. Ltd.,
3ꢀ1ꢀ98, Kasugadeꢀnaka, Konohanaꢀku, Osaka 554ꢀ0022, Japan
§Genomic Science Laboratories, Sumitomo Dainippon Pharma Co. Ltd., 3ꢀ1ꢀ98, Kasugadeꢀnaka,
Konohanaꢀku, Osaka 554ꢀ0022, Japan
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TABLE OF CONTENTS GRAPHIC
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ABSTRACT
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A practical and chromatographyꢀfree synthetic process to selective M1 and M4 muscarinic acetylcholine
receptors agonist was developed and demonstrated on a several hundred grams scale. The key feature of
this route is N,Nꢀdimethylcarbamoylation of the anilinic nitrogen on the spiro 7ꢀazaindoline structure via
intermolecular migration of the N,Nꢀdimethylcarbamoyl group. The resulting compound 1 was prepared
in 43% overall yield with a chemical purity >99% via six steps starting with
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(2ꢀchloropyridinꢀ3ꢀyl)acetonitrile.
Keywords: 7ꢀAzaindoline, N,NꢀDimethylcarbamoylation, Intermolecular migration
,
Muscarinic
acetylcholine receptors agonist, Xꢀray crystal structure
INTRODUCTION
Many of the important central actions of acetylcholine (ACh) are mediated by the muscarinic ACh
receptors (mAChRs).1 To date, five mAChR subtypes (M1−M5) have been identified and are considered
to play important roles in the peripheral and central nervous systems (CNS).2 Clinical trials with
xanomeline, an M1 and M4 preferring orthosteric agonist, demonstrated that M1 and M4 mAChRs are
attractive therapeutic targets for Alzheimer’s disease and schizophrenia.3 During one of our drug
discovery program,4 we have found the 7ꢀazaindoline derivative 1 as a lead candidate with selective M1
(EC50 = 133 nM) and M4 (EC50 = 47 nM) mAChRs agonistic activity. Further studies led to the selection
of this compound as a potential therapy for CNS disorders. However, to support preclinical studies, we
had to find a practical synthetic route to 1 suitable for kilogram scale. This is because the initial chemical
synthesis of 1 presents not only safety concerns, but also reproducibility and purification challenges. In
this report, we describe a successful synthetic route to 1 that overcomes conventional issues and
provides a practical process for scaleꢀup to several hundred grams.
Figure 1. Structure of a selective M1 and M4 mAChRs agonist 1
RESULTS AND DISCUSSION
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Initial Chemical Synthesis of 1.
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Various effective synthetic approaches of azaspiroindoline5 or spiroindoline6 derivatives were reported.
However, there are few examples about 7ꢀazaspiroindoline.7 Based on these literatures, we chose the
synthetic route in scheme 1 for construction of 7ꢀazaspiroindoline derivative 1 in terms of short synthetic
process. Compound 1 was first prepared from the commercially available (2ꢀChloropyridinꢀ3ꢀyl)
acetonitrile 2 in six steps and 30 % yield (Scheme 1). Deprotection at the benzylic position of compound
2 by NaH followed by reaction with 7 in DMSO gave the spiro compound 3 in 61% yield. Reduction of
3 with LiAlH(OtꢀBu)3 and subsequent cyclization provided the 7ꢀazaindoline derivative 4 in 84 % yield.
Next, N,Nꢀdimethylcarbamoylation of the anilinic nitrogen of 4 was accomplished in 68% yield by use
of excess amounts of N,Nꢀdimethylcarbamoyl chloride (4.0 equiv.) and triethylamine (8.0 equiv.).
Deprotection of the benzyl group of 5 with HCOONH4 in MeOH, followed by reductive amination with
the aldehyde 8 afforded the free form of 1 in 86% yield. Finally, salt formation with fumaric acid and
reꢀcrystallization using acetoneꢀethyl acetate gave the desired product 1 in 85% yield. The initial
synthesis enabled the production of gram quantities of 1. However, further development was necessary
to address largeꢀscale safety concerns and multiple chromatography purification. The initial synthesis of
1 presented three majors issues. Namely, 1) the potential of explosion in the steps that transform
compound 2 to 3 and compound 5 to 6, 2) the poor reproducibly in the step that provides compound 5
from 4, and 3) the need for column chromatography purification in the preparation of compounds 3 and
5.
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Scheme 1. Initial Synthesis of the Selective M1 and M4 mAChRs Agonist 1
Bn
Bn
N
Cl
Cl
N
N
7
NaH
LiAlH(Ot-Bu)3
CN
CN
THF
reflux, 16 h
84%
N
Cl
DMSO, 80 °C, 2.5 h
61%
N
N
Cl
N
H
2
3
4
CO2Et
N
O
1)
N
OHC
Bn
N
O
Cl
N(CH3)2
(4.0 equiv.)
H
N
8
CH3
N
NaBH(OAc)3, CH3COOH
CH2Cl2, rt, 2 h, 86%
O
10% Pd-C
HCO2NH4
Et3N (8.0 equiv.)
HOOC
N(CH3)2
2) fumaric acid
acetone-AcOEt
60 °C, 1 h, 85%
MeOH
reflux
3 h, quant.
N
N
toluene, 90 °C
3 h, 68%
N
N
COOH
N
N
N(CH3)2
N(CH3)2
O
O
O
5
6
1
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The preparation of compound 3 required the use of NaH, compound 7 in DMSO, and a temperature of
80 °C. These reaction conditions can cause explosion at scaleꢀup.8 In addition, the commercially
available 7·HCl had to be desalted with 10% K2CO3 solution before use and, as such, could not be stored
for long time. Furthermore, compound 3 could not be purified without column chromatography due to
the production of multi undesired byꢀproducts. To overcome these challenges, we considered
optimization of the reaction conditions.
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Table 1. Optimization of the Reaction Conditions
entry reagent base (equiv.) solvent temp. yield
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4
5
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7
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7
7
NaH (2.0) DMSO 80 °C 61%
KOH (2.0) DMSO 80 °C 67%
KOH (2.0) DMSO 30 °C 70%
KOH (3.0) DMSO 30 °C 82%
NaOH (3.0) DMSO 30 °C 65%
7
7·HCl
7·HCl
7·HCl K2CO3 (3.0) DMSO 30 °C 14%
7·HCl
9·HCl
KOH (3.0) DMF 30 °C trace
KOH (3.0) DMSO 30 °C trace
Construction of spiro ring structure at the benzylic position of compound 2 is usually conducted by NaH
in DMSO condition at high temperature. In previous literatures, spiro structure at benzylic position was
constructed by hexadecyltributylphosphonium bromide in 50% NaOH solution at reflux condition.9 We
tried to adapt and simplify this reaction condition. First, we changed the base used for deprotonation of
compound 2 from NaH to KOH (entry 2, Table 1). This change allowed the preparation of compound 3
in 67% yield as compared to 61% with NaH. Lowering the temperature from 80 °C to 30 °C (entry 3)
resulted in 70% yield. Next, we tried to use the commercially available 7·HCl without preꢀdesaltation
and instead used KOH at 3 equiv. to achieve inꢀsitu desaltation. This modification afforded 3 in 82%
yield (entry 4). Using NaOH or K2CO3 as a base provided 3 in 65% or 14%, respectably (entries 5 and 6).
When the solvent was changed to DMF, 3 was obtained only in trace amount (entry 7). In case of the
Boc protected reagent 9·HCl, multi products were afforded (entry 8). Based on these findings, we
selected the conditions of entry 4 for a safe reaction at ambient temperature.
Preparation of the 7ꢀAzaindoline Derivative 4 and N,NꢀDimetylcarbamoylation of Anilinic
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Nitrogen.
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Compound 4 was synthesized in acceptable yield from compound 3 using LiAlH(OtꢀBu)3 as a reductive
agent.10 However N,Nꢀdimethylcarbamoylation of the anilinic nitrogen of 4 required the use of excess
amounts of N,Nꢀdimethylcarbamoyl chloride (4.0 equiv.) and triethylamine (8.0 equiv.), giving 5 in 68%
yield (entry 2, Table 2). Without these excess amounts (entry 1, Table 2), compound 5 was obtained in
only 28% isolated yield. Although the excess conditions provide 5 in 68% yield (entry 2), the
reproducibility of this yield is poor, especially in case of large scale production (>10 g). Furthermore,
some of the starting material 4 remained after the reaction (entries 1 and 2), necessitating column
chromatography for separation of 5. Thus, perfect conversion will be needed to avoid column
chromatography. To avoid chromatography, we conducted further investigation and found that this
reaction proceeds in two steps via intermolecular migration of the N,Nꢀdimethycarbamoly group11 as
depicted in Scheme 2. Details of an experimental and theoretical study of this reaction indicate that the
concentration, temperature and procedure are important to accomplish complete conversion of 4 to 5.12
Therefore, optimized reaction conditions as described in entry 3 afforded the desired compound 5 in
99% yield (entry 3). Details of the procedure are described in the experimental section.
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Table 2. Effects of Base, Concentration and Temperature on N,NꢀDimethylcarbamoylation of 4
entry A (equiv.)
base (equiv.)
Et3N (2.0)
conc. temp. yield
0.2 M 90 °C
0.2 M 90 °C
2.0 M reflux
1
2
3
1.5
4.0
28%
68%
99%
DIPEA (8.0)
1.5 + 0.2 DIPEA (2.0 + 0.6)
Scheme 2. Reaction Mechanism of N,NꢀDimethylcarbamoylation of 4
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Deprotection of the benzyl group of 5, giving compound 6.
In initial synthesis of 1, deprotection of benzyl group of 5 was achieved by 10%Pd/C and HCOONH4 in
methanol under reflux condition, affording 6 in quantitative yield (Table 3, entry 1). However, at
kilogram scale, it is possible that HCOONH4 decomposition material clogs the reflux condenser. To
avoid such risk, hydrogen was used as reducing agent and 10% Pd/C as catalyst (entry 2). Using 20%
Pd(OH)2/C (entry 3) reduced the reaction time by half to 8 h (entry 3).
Table 3. Optimization of Deprotection of the Benzyl Group of 5
entry
reagents
temp. time conversion
1
2
3
HCOONH4
10%Pd/C
reflux 3 h
>99%
H2
H2
10%Pd/C
20%Pd(OH)2/C 40 °C 8 h
40 °C 16 h
70%
>99%
Reductive Amination of 6 with 8.
In the initial synthesis of 11, compound 6 was reacted with the aldehyde 8 and NaBH(OAc)3 in CH2Cl2
to afford compound 11 in 86% yield (entry 1, Table 4). In this reaction, the isolated yield was acceptable,
however, a halogenated solvent (dichloromethane) was used. Generally, a replacement for this solvent is
desired for pharmaceutical manufacturing. We found that this reaction could be conducted in THF with
the desired compound 11 obtained in >99% yield (entry 2, Table 4). By using toluene, the yield of
compound 11 decreased to 91%. Furthermore compound 11 was obtained in >99% yield without
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addition of acetic acid (entry 3, Table 4). These optimization steps allowed the preparation of compound
11 in 99% yield without the use of a halogenated solvent, which is suitable pharmaceutical
manufacturing. Single crystal Xꢀray structure of the obtained compound 11 is depicted in Figure 2, and
the details are described in the supporting information.13
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Table 4. Reductive Amination of compound 6
entry
AcOH
1.0 equiv. CH2Cl2 2 h 86%
1.0 equiv. THF 2 h >99%
1.0 equiv. toluene 2 h 91%
THF 2 h >99%
solvent time yield
1
2
3
4
ꢀ
Figure 2. Xꢀray crystal structure of 11
Improved Process for Preparation of 1.
Based on all the optimizations described above, we attempted the synthesis of 1 at several hundred
grams scale. Starting with 477 g of compound 2, we were able to generate compound 11 (free form of 1)
in good yield (Scheme 3). Finally, transformation into a salt using fumaric acid in ethanol and
reꢀcrystallization with acetoneꢀethyl acetate at 60 °C afforded the target compound 1 in 43% over all
yield and >99% HPLC area purity.
Scheme 3. Improved Process for the Preparation of 1
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CONCLUSION
In conclusion, we described in this study the initial synthesis of compound 1, a selective M1 and M4
mAChRs agonist, and carried various optimizations to suit production at several hundred grams scale.
Our work addressed safety and methodology concerns and simplified the original synthesis. We also
accomplished a higher yield than the initial synthetic route.
EXPERIMENTAL SECTION
General. All reagents and solvents were purchased from commercial suppliers and used without further
purification. Nuclear Magnetic Resonance (NMR) spectra were recorded at ambient temperature on a
JEOL JNMꢀAL400 FT NMR spectrometer, operating at 400 MHz for 1H NMR and 100 MHz for 13C
NMR. ESI mass spectra were measured on a SHIMADZU LCMSꢀ2010EV. ESI mass spectra (for
HRMS) were measured on a Thermo Fischer Science LTQ Orbitrap Discovery MS equipment. Infrared
(IR) spectra were measured on a SHIMADZU IRAffinityꢀ1 equipped with Smiths Detection DuraScope.
Melting points were determined on a YANACO MPꢀJ3 without correction. Elemental analysis was
performed on a CE Instruments EA1110 and a Yokokawa analytical system IC7000. HPLC analysis was
performed on Agilent 1100 Series. NMR spectra of unknown compounds are included within the
supporting information. Characterization of compound 4, 5, 10 and 11 have been described in our
previous paper (Tetrahedron 2013, 69, 9675ꢀ9681).
1ꢀBenzylꢀ4ꢀ(2ꢀchloropyridinꢀ3ꢀyl)piperidineꢀ4ꢀcarbonitrile (3). To a solution of 2 (477 g, 3.13 mol) in
DMSO (9.5 L) was added KOH pellet (544 g, 9.69 mol) portion wise at room temperature.
N-Benzylꢀbis(2ꢀchloroethyl)amine hydrochloride (840 g, 3.13 mol) was added to reaction mixture
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portion wise and reaction was continued at 30 °C for 2.5 h. The reaction mixture was cooled to 20 °C
and poured on water and extracted with ethyl acetate. The organic extracts were washed with brine, dried
over Na2SO4, filtered and concentrated under reduced pressure to give brown solid (959 g). Ethyl acetate
(190 mL) was added to the brown solid and stirred at 65 °C for 3 h. The reaction mixture was cooled to
5 °C and stirred for 2 h. The reaction mixture was filtrated to give pale brown solid and the solid was
washed by nꢀhexane to afford 3 (627 g, 64%). Brown solid; mp 144 °C; 1 H NMR (400 MHz, CDCl3 ) δ
2.10 (dt, J = 12.9, 3.7 Hz, 2H), 2.51 (dd, J = 12.9, 2.4 Hz, 2H), 2.61 (dt, J = 12.9, 2.0 Hz, 2H), 3.05 (d, J
= 12.9 Hz, 2H), 3.62 (s, 2H), 7.26−7.34 (m, 6H), 7.75 (dd, J = 8.1, 1.7 Hz, 1H), 8.40 (dd, J = 4.6, 1.7 Hz,
1H); 13C NMR(100 MHz, CDCl3) δ 33.8, 40.4, 50.0, 62.7, 119.7, 122.9, 127.3, 128.4, 129.1, 133.4,
136.0, 138.0, 149.0, 150.5; IR 702, 741, 795, 991, 1400, 1562 cmꢀ1; MS (ESI, positive) m/z 312 (M+H)+;
HRMS (ESI, positive) m/z calcd for C18H19N3Cl (M+H)+ 312.1262, found 312.1270.
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1ꢀBenzylꢀ1',2'ꢀdihydrospiro[piperidineꢀ4,3'ꢀpyrrolo[2,3ꢀb]pyridine] (4). To a solution of
LiAlH(OtꢀBu)3 (2.55 kg, 10.0 mol) in THF (3.1 L) was added to a THF solution of 3 (625 g, 2.00 mol)
in THF solution (4.4 L) portion wise at 65 °C. The mixture was stirred under reflux for 23 h. The
resulting mixture was cooled to 0 ºC and ice water was poured on ice water. The reaction mixture was
extracted with ethyl acetate and washed with 20% NaCl aqueous solution, dried over Na2SO4, filtered
and concentrated under reduced pressure to give brown solid (551 g). The brown solid was dissolved in
ethyl acetate (1.1 L) at 75 °C and the reaction mixture was cooled to 25 °C and stirred for 1.5 h.
nꢀHexane was added the mixture and filtrated to give pale brown solid. The pale brown solid was
washed by nꢀhexane to afford 4 (406 g, 73%).
1ꢀBenzylꢀN,Nꢀdimethylspiro[piperidineꢀ4,3'ꢀpyrrolo[2,3ꢀb]pyridine]ꢀ1'(2'H)ꢀcarboxamide (5).
N, NꢀDiisopripylethylamine (105 mL, 752 mmol) was added to a stirred solution of 4 (105 g, 376 mmol)
in toluene (247 mL) at room temperature. N, NꢀDimetylcarbamoyl chloride (52 mL, 564 mmol) was
added to the reaction mixture and reaction was continued at reflux condition. After 3 h,
N,Nꢀdiisopripylethylamine (32 mL, 226 mmol) and N,Nꢀdimetylcarbamoyl chloride (10 mL, 113 mmol)
were added to reaction mixture. Additional 3 h, the reaction mixture was cooled to room temperature
and quenched by water and separated. The water layer was extracted with AcOEt and combined. The
organic extracts were washed with saturated aqueous NaHCO3 solution and brine, dried over Na2SO4,
filtered and concentrated under reduced pressure to give 5 (132 g).
N, NꢀDimethylspiro[piperidineꢀ4,3'ꢀpyrrolo[2,3ꢀb]pyridine]ꢀ1'(2'H)ꢀcarboxamide (6).
A mixture of 5 (132 g, 376 mmol), 20% Pd(OH)2 on carbon (23.8 g) and MeOH (1.1 L) was stirred and
stirred at 40 °C under H2 atmosphere for 8 h. Filtrated Pd on carbon and concentrated under reduced
pressure to give 6 (96 g). White solid; mp 104 °C; 1 H NMR(400 MHz, CDCl3 ) δ 1.68−1.71 (m, 2H),
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1.79−1.86 (m, 2H), 2.76−2.83 (m, 2H), 3.01 (s, 6H), 3.03−3.10 (m, 2H), 3.80 (s, 2H), 6.74 (dd, J = 7.3,
5.1 Hz, 1H), 7.35 (dd, J = 7.3, 1.7 Hz, 1H), 8.03 (dd, J = 5.1, 1.7 Hz, 1H); 13C NMR(100 MHz, CDCl3)
δ 36.2, 38.1, 40.3, 43.1, 57.7, 116.2, 130.3, 132.3, 146.5, 157.2, 57.6; IR 771, 1219, 1366, 1381,1419,
1651 cmꢀ1; MS (ESI, positive) m/z 260 (M+H)+; HRMS (ESI, positive) m/z calcd for C14H21ON4 (M+H)+
261.1710, found 261.1718.
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Ethyl4ꢀ{[1'ꢀ(dimethylcarbamoyl)ꢀ1',2'ꢀdihydroꢀ1Hꢀspiro[piperidineꢀ4,3'ꢀpyrrolo[2,3ꢀb]pyridin]ꢀ1ꢀy
l]methyl}piperidineꢀ1ꢀcarboxylate fumarate (1). To a solution of 6 (96 g, 370 mmol) and
4ꢀformylpiperidineꢀ1ꢀcarboxylate 8 (69 g, 370 mmol) in THF (542 mL) was added sodium
triacetoxyborohydride (118 g, 555 mmol) at room temperature. After 2 h, 10% aqueous K2CO3 solution
(963 mL) was added to the mixture drop wise at 0 ºC. The mixture was extracted with toluene. Organic
layer was separated and concentrated under reduced pressure to afford 11 as white solid (164 g). The
white solid 11 (130 g, 301 mmol) was dissolved in ethanol (1.2 L) and fumaric acid (35 g, 301 mmol)
was added to the solution. The mixture was stirred for 1 h and solvent was evaporated under reduced
pressure to give colorless amorphous (119 g, 99%). The colorless amorphous was dissolved in acetone
(1.2 L), ethyl acetate (1.0 L) was added to the solution and wormed up to 60 °C. After 1 h, reaction
mixture was cooled to 25 °C and white precipitate was collected by suction filtration. The white
precipitate was washed by ethyl acetate (431 mL) and dried to give 1 (150 g, 91% in four steps, 99.7 %
1
HPLC area purity). White solid; H NMR (400 MHz, DMSO−d6) δ 0.94−1.06 (m, 2H), 1.15 (t, J = 7.1
Hz, 3H), 1.65−1.78 (m, 5H), 1.88−1.98 (m, 2H), 2.30ꢀ2.42 (m, 4H), 2.70−2.89 (m, 4H), 2.90 (s, 6H),
3.65 (s, 2H), 3.89ꢀ3.99 (m, 2H), 4.00 (q, J = 7.1 Hz, 2H), 6.58 (dd, J = 7.3, 5.1 Hz, 1H), 7.54 (d, J = 6.6
Hz, 1H), 7.99 (d, J = 5.1 Hz, 1H); 13C NMR (100 MHz, DMSO−d6) δ 14.6, 30.2 (×2), 32.1, 33.9 (×2)
,
37.3 (×2), 43.2 (×2), 49.7 (×2), 57.0, 60.5, 63.0, 116.3, 130.7, 131.4, 134.4 (×3), 146.2, 154.6, 156.6,
157.2, 166.7 (×2); IR 1273, 1377, 1423, 1663, 1680 cmꢀ1; MS (ESI, positive) m/z 430 (M+H)+; HRMS
(ESI, positive) m/z calcd for C23H36O3N5 (M+H)+ 430.2813, found 430.2824; Anal. calcd for
C23H35N5O3·C4H4O4·1/2 H2O: C, 58.47; H, 7.27; N, 12.63. Found: C, 58.28; H, 7.25, N, 12.63.; HPLC
analysis conditions (Column: Eclipse Plus C18 4.6 × 100 mm, 3.5 µm; flow 1.0 mL/min; Eluent:
Gradient 10−90% methanol in water containing 0.01% TFA).
ASSOCIATED CONTENT
Supporting Information
1H NMR and 13C NMR of compounds 1, 3 and 6. Detail data of a single crystal Xꢀray structure analysis.
This material is available free of charge via the Internet at http://pubs.acs.org.
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AUTHOR INFORMATION
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* Corresponding author. Tel.: +81ꢀ6ꢀ6466ꢀ3104; fax: +81ꢀ6ꢀ6337ꢀ5287;
Eꢀmail: yoshiharuꢀuruno@dsꢀpharma.co.jp
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Present Adresses
¶ Current address: Faculty of Chemistry, Materials and Bioengineering, Department of Life Science and
Biotechnology, Kansai University, 3ꢀ3ꢀ35 Yamateꢀcho, Suita, Osaka 564ꢀ8680, Japan.
Notes
The authors declare no competing financial interest.
ACKNOWLEDGMENTS
We really thank Keiko Bando for carrying out elemental analyses, Naohiro Nishimura and Sayo
Takayama for HRMS analyses and Kenjiro Hira for crystallization of compound 11.
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at room temperature for 1 day. Single crystal of 11 was formed.
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