The present invention relates to an improved process for the preparation of a compound of
Formula 1, used in the synthesis of Semaglutide.
10
O
O
O
NH
O
O O
NH
O
O
O
NH O
O
O
HN
O
OH
HN O
O
Formula 1
The present invention also relates to an improved process for the preparation of a compound
15 of Formula G, used in the synthesis of the compound of Formula 1.
O
O
O
NH
O O
O
O
X
Formula G
Wherein, COOX is an activated ester
20
The present invention further relates to an improved process for the preparation of a
compound of Formula I, used in the synthesis of the compound of Formula 1.
O
O
O
O
X
3
Formula I
Wherein, COOX is an activated ester
The present invention further relates to an improved process for the preparation of a
5 compound of Formula E, used in the synthesis of compound of Formula 1.
NH O
O
O
O
X
PG1
Formula E
wherein PG1 is a protecting group, and COOX is an activated ester
10
BACKGROUND OF THE INVENTION
Semaglutide, a human GLP-1 receptor agonist (or GLP-1 analog) is represented by the
Formula 2,
15
Formula 2
Semaglutide (marketed under the trade name Ozempic®, Novo Nordisk) is indicated as an
adjunct to diet and exercise to improve glycemic control in adults with type 2 diabetes
mellitus.
20
Semaglutide, as represented by Formula 2 is disclosed in WO 2006/097537. WO
2006/097537 provides a process wherein the Lys20 side chain is attached to full-chain peptide
4
in one step in solution phase. The process for the preparation of Lys20 side chain or
Semaglutide side chain of Formula 1 is reported in CN 104356224 and CN 110041219.
The process disclosed in CN 104356224 involve tedious work up procedures such as pH
5 adjustment, filtration, washings, back extraction etc. for the isolation of intermediate
compounds prepared during the synthesis. The process also involves recrystallization steps
using ethanol. Moreover, the process disclosed in CN 104356224 is a less efficient process,
as some of the steps require overnight stirring.
10 The process disclosed in CN 110041219, requires stringent operations such as column
chromatography for the purification / isolation of reaction intermediates. This process
requires Pd/c at various reaction steps, which makes this process less suitable for the
commercial purposes.
15 From the foregoing, it is apparent that the reported methods for the preparation of side chain
of Semaglutide require stringent operational conditions, which are not only tedious but also
result in significant yield loss. The processes require long reaction time for the completion at
several stages including tedious work up procedures and purification steps.
Thus, there remains the need to formulate an efficient, simple and industrially viable
20 synthetic process which can overcome the drawbacks of the prior art.
OBJECT OF THE INVENTION
It is an object of the present invention to overcome the above-mentioned drawbacks of the
prior art.
25
It is another objective of the present invention to avoid tedious and long work-up procedures
during the synthesis.
It is a further objective of the present invention to provide an improved and commercially
30 viable process for the synthesis of Semaglutide side chain.
5
SUMMARY OF THE INVENTION
In a first aspect, the present invention relates to a process for the preparation of a compound
5 of Formula 1,
O
O
O
NH
O
O O
NH
O
O
O
NH O
O
O
HN
O
OH
HN O
O
Formula 1
comprising the steps of:
a) coupling a compound of Formula A;
10
NH O
O
O
OH
PG1
Formula A
wherein PG1 is a protecting group, cleavable under non-basic conditions,
15
by activation of the carboxylic acid, with a compound of Formula B;
O
O
HN
HOOC
NH2
Formula B
20
to obtain a compound of Formula C;
6
NH O
O
O
NH
O
OH
HN
O
O
PG1
Formula C
b) de-protecting the compound of Formula C to obtain a compound of Formula D or its
salt;
O
O
O
NH
O
OH
HN
O
O
H2N
5
Formula D
c) reacting the compound of Formula D or its salt with a compound of Formula E;
NH O
O
O
O
X
PG1
10 Formula E
Wherein COOX is an activated ester
in the presence of a base, to obtain a compound of Formula F;
NH O
O
O
NH O
O
O
HN
O
OH
HN O
O
PG1
15
Formula F
7
d) de-protecting the compound of Formula F;
e) reacting the reaction mixture of step d) with a compound of Formula G;
O
O
O
NH
O O
O
O
X
5
Formula G
Wherein, COOX is an activated ester
in the presence of a base to obtain the compound of Formula 1;
10 Wherein:
i) the compound of Formula E used in step c) is prepared by a process
comprising the reaction of a compound of Formula E1;
NH O
O
O
OH
PG1
15 Formula E1
with a carboxylic acid activator;
and,
20 ii) the compound of Formula G used in step e) is prepared by a process comprising the
steps of:
f1) coupling a compound of Formula H;
O
O
O
OH
25 Formula H
by activation of the carboxylic acid, with a compound of Formula J
8
O OH
O O
NH2
Formula J
to obtain a compound of Formula K;
5
O
O
O
NH
O
O
HO O
Formula K
f2) reacting the compound of Formula K with a carboxylic acid activator to obtain the
compound of Formula G.
10
In another aspect, the present invention relates to a process for the preparation of a
compound of Formula C;
NH O
O
O
NH
O
OH
HN
O
O
PG1
Formula C
15 wherein PG1 is a protecting group, cleavable under non-basic conditions,
comprising the steps of:
a) reacting a compound of Formula A;
NH O
O
O
OH
PG1
20
9
Formula A
with a carboxylic acid activator in the presence of N-(3-Dimethylaminopropyl)-N′-
ethylcarbodiimide hydrochloride in a polar aprotic solvent selected from N,N5 dimethylformamide, dimethyl sulfoxide and tetrahydrofuran, preferably
dimethylformamide;
b) reacting the reaction mixture of step a) with a compound of Formula B;
O
O
HN
HOOC
NH2
10
Formula B
in the presence of N′N-diisopropylethylamine to obtain a compound of Formula C.
In another aspect, the present invention relates to a process for the preparation of a
15 compound of Formula G;
O
O
O
NH
O O
O
O
X
Formula G
Wherein, COOX is an activated ester
20 comprising a reaction of a compound of Formula K;
10
O
O
O
NH
O
O
HO O
Formula K
with a carboxylic acid activator in the presence of N-(3-Dimethylaminopropyl)-N′-
ethylcarbodiimide hydrochloride.
5
The aspect of the invention provides a process for the preparation of a compound of
Formula I;
O
O
O
O
X
10 Formula I
Wherein, COOX is an activator ester
comprising the reaction of a compound of Formula H;
O
O
O
OH
Formula H
15 with a carboxylic acid activator in the presence of N-(3-Dimethylaminopropyl)-N′-
ethylcarbodiimide hydrochloride to obtain a compound of Formula I.
Another aspect of the invention is to provide a compound of Formula EB;
NH O
O
O
O
N
N
N
PG1
20
Formula EB
wherein PG1 is a protecting group.
In another aspect, the present invention provides a process for the preparation of a
25 compound of Formula E;
11
NH O
O
O
O
X
PG1
Formula E
wherein PG1 is a protecting group, and COOX is an activated ester
5
comprising a reaction of a compound of Formula E1;
NH O
O
O
OH
PG1
Formula E1
10
with a carboxylic acid activator in the presence of N-(3-Dimethylaminopropyl)-N′-
ethylcarbodiimide hydrochloride.
Another aspect of the present invention provides the use of the compounds of Formula C,
15 G, I and E for the preparation of the Semaglutide side chain of Formula 1.
DEFINITIONS
The following definitions are used in connection with the present application unless the
20 context indicates otherwise.
The term “Coupling” refers to a reaction where an amide linkage is formed by reacting
carboxylic acid with an amine. Preferably, the coupling is carried out in the presence of a
coupling agent. Examples of coupling agent include but are not limited to benzotriazol-1-yl25 oxytripyrrolidinophosphonium hexafluorophosphate (PyBOP), 3-
[Bis(dimethylamino)methyliumyl]-3H-benzotriazol-1-oxide hexafluorophosphate (HBTU),
2-(1H-Benzotriazole-1-yl)-1,1,3,3-tetramethylaminium tetrafluoroborate (TBTU), N-EthylN'-(3-dimethylaminopropyl)carbodiimide hydrochloride, N,N′-Dicyclohexylcarbodiimide
and N,N′-Diisopropylcarbodiimide. The preferred coupling agent is N-Ethyl-N'-(3-
30 dimethylaminopropyl)carbodiimide hydrochloride.
12
The term “carboxylic acid” is an organic compound that contains a -COOH group.
The term “activation of carboxylic acid” refers to conversion of –OH of the carboxylic acid
5 into a good leaving group by reacting with a carboxylic acid activator to result in an
activated ester. Examples of carboxylic acid activators include but are not limited to Nhydroxysuccinimide (HOSu), N-hydroxy-5-norbornene2,3-dicarboximide (HONB), 1-
hydroxybenzotriazole (HOBt), 6-chloro-1-hydroxybenzotriazole, (6-Cl-HOBt), 1-hydroxy-7-
azabenzotriazole (HOAt), 3-hydroxy-4-oxo-3,4-dihydro-1,2,3-benzotriazine (HODhbt), ethyl
10 1-hydroxy-1H-1,2,3-triazole-4-carboxylate (HOCt) and N-hydroxytetrazole (HOt). The
preferred carboxylic acid activator is selected from N-hydroxysuccinimide (HOSu) and 1-
hydroxybenzotriazole (HOBt).
The term “activated ester” refers to an ester functional group that is highly susceptible toward
15 nucleophilic attack. An activated ester undergoes the same reactions as their inactivated
analogues but do so more rapidly. An activated ester can be prepared by a reaction of
carboxylic acid functional with a carboxylic acid activator.
The term “protecting group” refers to a temporarily attached group to decrease reactivity of
20 a functional group so that the protected functional group does not react under synthetic
conditions to which the molecule is subjected in one or more subsequent steps. For the
purpose of the present invention, examples of protecting group PG1 include but are not
limited to Carbobenzyloxy (Cbz), p-Methoxybenzyl carbonyl (Moz or MeOZ), tertButyloxycarbonyl (BOC), 9-Fluorenylmethyloxycarbonyl (Fmoc), Acetyl (Ac), Benzoyl
25 (Bz), Benzyl (Bn), p-Methoxybenzyl (PMB), Tosyl (Ts) and Troc (trichloroethyl
chloroformate). The preferred protecting group is tert-Butyloxycarbonyl (BOC).
The term “a protecting group cleavable under non-basic conditions” is a protecting group
which can be cleaved under acidic and/or neutral conditions. Examples of protecting group
cleavable under non-basic conditions include but are not limited to tert-butoxycarbonyl (t30 BOC), triphenylmethyl, methanesulfonyl, acetyl and benzoyl. The preferred protecting group
cleavable under non-basic conditions is tert-Butyloxycarbonyl (BOC). The protection and
deprotection methods are well known in the art (see notably "Protective groups in organic
synthesis", Greene T. W. and Wuts P. G. M., Wiley-Interscience, 1999).
13
The term “ambient temperature” refers to a temperature ranging from about 15 °C to 35 °C,
preferably to a temperature ranging from about 20 °C to 30 °C, more preferably to a
temperature of 20 - 25 °C.
5
DETAILED DESCRIPTION OF THE INVENTION
The present invention relates to a process for the preparation of the side chain of Semaglutide
of Formula I;
O
O
O
NH
O
O O
NH
O
O
O
NH O
O
O
HN
O
OH
HN O
10 O
Formula 1
In an embodiment, the process comprises a step for the preparation of a compound of
Formula C;
NH O
O
O
NH
O
OH
HN
O
O
PG1
15
Formula C
In one embodiment, the compound of Formula C is prepared by coupling a compound of
Formula A;
NH O
O
O
OH
PG1
20
14
Formula A
wherein PG1 is a protecting group, cleavable under non-basic conditions
by activation of the carboxylic acid, with a compound of Formula B;
5
O
O
HN
HOOC
NH2
Formula B
The activation of the carboxylic acid is carried out in the presence of a carboxylic acid
activator, preferably selected from the N-hydroxysuccinimide (HOSu), N-hydroxy-5-
10 norbornene2,3-dicarboximide (HONB), 1-hydroxybenzotriazole (HOBt), 6-chloro-1-
hydroxybenzotriazole, (6-Cl-HOBt), 1-hydroxy-7-azabenzotriazole (HOAt), 3-
hydroxy-4-oxo-3,4-dihydro-1,2,3-benzotriazine (HODhbt), ethyl 1-hydroxy-1H-1,2,3-
triazole-4-carboxylate (HOCt) and N-hydroxytetrazole (HOt). In a preferred
embodiment, the compound of Formula A is reacted with a carboxylic acid activator
15 1-hydroxybenzotriazole (HOBt). The resulting reaction mixture is reacted with a
compound of Formula B to obtain a compound of Formula C.
PG1 is preferably selected from tert-butoxycarbonyl (t-BOC), triphenylmethyl,
methanesulfonyl, methanesulfonyl, acetyl and benzoyl. The coupling is carried out in
20 the presence of a coupling agent selected from benzotriazol-1-yloxytripyrrolidinophosphonium hexafluorophosphate (PyBOP), 3-
[Bis(dimethylamino)methyliumyl]-3H-benzotriazol-1-oxide hexafluorophosphate
(HBTU), 2-(1H-Benzotriazole-1-yl)-1,1,3,3-tetramethylaminium tetrafluoroborate
(TBTU), N-Ethyl-N'-(3-dimethylaminopropyl)carbodiimide hydrochloride, N,N′-
25 Dicyclohexylcarbodiimide and N,N′-Diisopropylcarbodiimide, preferably, N-(3-
Dimethylaminopropyl)-N’-ethylcarbodiimide hydrochloride .
15
In a preferred embodiment, the compound of Formula A [wherein PG1 is tertbutoxycarbonyl (t-BOC)] is reacted with 1-hydroxybenzotriazole (HOBt). The
resulting reaction mixture is reacted with a compound of Formula B to obtain a
compound of Formula C.
5
Preferably, the coupling is carried out in the presence of a base selected from
triethylamine, N′N-diisopropylethylamine, sodium bicarbonate and sodium carbonate,
preferably N′N-diisopropylethylamine.
The reaction may be carried out in a solvent. The solvent may be selected from the
10 group comprising N-dimethylformamide, dimethylsulfoxide, aromatic hydrocarbons,
such as toluene, benzene, xylene or the like, saturated hydrocarbons, such as
cyclohexane, hexane or the like, halogenated hydrocarbons, such as dichloromethane,
chloroform, 1,2-dichloroethane or the like, ethers, such as tetrahydrofuran,
diethylether, dioxane, 1,2-dimethoxyethane or the like, esters, such as methyl acetate,
15 ethyl acetate or the like, ketones, such as acetone, diisobutyl ketone, cyclohexanone,
methylcyclohexanone, methyl ethyl ketone, methyl isobutyl ketone, acetylacetone or
the like, nitriles, such as acetonitrile or the like, alcohols, such as methanol, ethanol, tbutanol, isopropanol or the like, water, ethers, such as tetrahydrofuran, diethylether,
dioxane or the like, and mixtures thereof. Preferably, the solvent is polar aprotic
20 solvent selected from N,N-dimethylformamide, dimethyl sulfoxide and
tetrahydrofuran. More preferably, the solvent is N-dimethylformamide.
In a preferred embodiment, the compound of Formula A [wherein PG1 is tertbutoxycarbonyl (t-BOC)] is reacted with 1-hydroxybenzotriazole (HOBt) in the
25 presence of N-dimethylformamide. The resulting reaction mixture is reacted with a
compound of Formula B in the presence of N′N-diisopropylethylamine to obtain a
compound of Formula C. More preferably, the resulting reaction mixture is reacted
with a compound of Formula B in the presence of a base and water to obtain a
compound of Formula C.
30
In one aspect of the invention, the compound of Formula A is reacted with a
carboxylic acid activator in the presence of N-(3-Dimethylaminopropyl)-N’-
16
ethylcarbodiimide hydrochloride in a polar aprotic solvent selected from N,Ndimethylformamide, dimethyl sulfoxide and tetrahydrofuran. The resulting reaction
mixture is reacted with a compound of Formula B in the presence of N′Ndiisopropylethylamine and water.
5
In a preferred embodiment, the compound of Formula A (wherein PG1 is t-BOC) is
reacted with 1-hydroxybenzotriazole (HOBt) in the presence of N-(3-
Dimethylaminopropyl)-N’-ethylcarbodiimide hydrochloride in N,Ndimethylformamide at an ambient temperature. More preferably, the reaction mixture
10 is stirred for 2-3 hours at an ambient temperature. Subsequently, the reaction mixture
is reacted with a compound of Formula B in the presence of N′Ndiisopropylethylamine and water at an ambient temperature.
Surprisingly, the inventors of the present invention found that the compound of
15 Formula C can be isolated in a very simple and commercially viable method. The
compound of Formula C can be isolated by adjusting the pH of the reaction mixture
from 3 to 4 and evaporating the solvent. The pH of the solution can be adjusted using
diluted HCl or diluted phosphoric acid. The compound can be extracted using ethyl
acetate or dichloromethane.
20
This highly efficient procedure overcomes the disadvantages of the prior art
procedures described above. The process of the present invention avoids tedious and
time-consuming steps of work up procedure for the isolation of the intermediate
during the synthesis of the compound of Formula C. The reaction step is completed in
25 a significant shorter time (appx. 2-4 hours) compared to the prior art process which
requires overnight stirring at one stage. Moreover, the work up procedure for the
isolation of the compound of Formula C is easy to operate at commercial scale. The
use of ethanol as needed in the prior art procedure is disadvantageous in commercial
manufacturing and this disadvantage is also overcome by the present invention.
30
The present invention has the advantage of providing the desired compound of
Formula C which is very low in impurities.
17
The compound of Formula C can be converted to a compound of Formula 1 by any
method known in the art, for example, using a process as reported in CN104356224.
In one embodiment, the compound of Formula C is converted to the compound of
Formula 1 by following the process, which is another aspect of the invention. The
5 process comprises a step for the preparation of a compound of Formula D or its salt;
O
O
O
NH
O
OH
HN
O
O
H2N
Formula D
by de-protection of the compound of Formula C. The formation of corresponding salts
10 depends on the corresponding acid used for de-protection of compound of Formula C.
In a preferred embodiment, the compound of Formula D or its salt is prepared by
treating a compound of Formula C with an acid, optionally in the presence of a
solvent. Preferably, the deprotection is carried out using 1 to 10 equivalents of acid,
15 more preferably 1 to 3 equivalents at ambient temperature for at least 1 minute,
preferably over a time period ranging from 1 minute to 5 hours, more preferably from
1 to 2 hours.
The acid may be selected from the group comprising mineral acids, such as
hydrochloric acid, hydrobromic acid, phosphoric acid, metaphosphoric acid, nitric
20 acid and sulphuric acid; and organic acids, such as tartaric acid, acetic acid,
trifluoroacetic acid, citric acid, malic acid, lactic acid, fumaric acid, benzoic acid,
glycolic acid, gluconic acid, succinic acid, alkylsulphonic acids, such as
methanesulphonic, ethanesulphonic acids, ethane-1, 2-disulfonic acid and 2-
hydroxyethanesulfonic acid, arylsulphonic acids, such as benzene sulfonic acid, 2-
25 naphthalenesulfonic acid, p-toluenesulphonic acid and naphthalene-1,5-disulfonic
acid, preferably p-toluenesulphonic acid.
18
The solvent may be selected from the group comprising non-polar solvents, such as
hexane, benzene, toluene, dioxane, ethers, halogenated hydrocarbons, such as
dichloromethane, chloroform, 1,2-dichloroethane or the like, polar aprotic solvents,
such as tetrahydrofuran, ethyl acetate, acetone, dimethylformamide, acetonitrile,
5 dimethyl sulfoxide or the like, polar protic solvents, such as methanol, ethanol, tbutanol, n-butanol, isopropanol, formic acid, acetic acid, nitromethane or the like,
water, esters, such as methyl acetate, ethyl acetate or the like, ketones, such as
acetone, diisobutyl ketone, cyclohexanone, methylcyclohexanone, methyl ethyl
ketone, methyl isobutyl ketone, acetylacetone or the like, preferably dichloromethane.
10
In a preferred embodiment, the de-protection is carried out by treating a compound of
Formula C with p-toluenesulphonic acid in the presence of a solvent such as
dichloromethane. The reaction mixture is stirred for 1 to 2 hours, at ambient
temperature to obtain a compound of Formula D or its salt.
15 The compound of Formula D or its salt can be converted to a compound of Formula 1
by any method known in the art, for example, using a process as reported in
CN104356224.
In one embodiment, the compound of Formula D or its salt is converted to the
20 compound of Formula 1 by the process steps of the present invention.
The process comprises a step for the preparation of a compound of Formula F;
NH O
O
O
NH O
O
O
HN
O
OH
HN O
O
PG1
25 Formula F
The compound of Formula F can be prepared by reacting a compound of Formula D
or its salt with a compound of Formula E,
19
NH O
O
O
O
X
PG1
Formula E
Wherein COOX is an activated ester and PG1 is a protecting group, cleavable under
non-basic conditions; in the presence of a base.
5
X is selected from Succinimide, 5-norbornene2,3-dicarboximide, Benzotriazole, 6-
chloro-benzotriazole, Azabenzotriazole, Oxo-3,4-dihydro-1,2,3-benzotriazine, Ethyl
1,2,3-triazole-4-carboxylate and tetrazole, preferably Benzotriazole. The base may be
selected from triethylamine, N′N-diisopropylethylamine, sodium bicarbonate and
10 sodium carbonate, preferably sodium bicarbonate.
In a preferred embodiment, the compound of Formula D or its salt, preferably the para
toluenesulphonic acid addition salt of Formula D, is reacted with a compound of
Formula E in the presence of sodium bicarbonate in water. Preferably, the reaction is
15 carried out at -10 to 30 degree Celsius, more preferably at 20 to 30 degree Celsius.
The reaction mixture is stirred for 2 to 3 hours.
The present invention has an advantage of providing the desired compound of
Formula F which is very low in impurities.
20 The compound of Formula F can be converted to a compound of Formula 1 by any
method known in the art, for example, using a process as reported in CN104356224.
In one embodiment, the compound of Formula F is converted to the compound of
Formula 1 by following the process, which is another aspect of the invention.
25
The compound of Formula E as used in the present invention can be prepared by
using methods known in the art, for example, using a process as reported in
CN104356224.
20
In one embodiment, the compound of Formula E is prepared by the process steps as
per the present invention. The process comprises a step for the preparation of a
compound of Formula E by reacting a compound of Formula E1;
NH O
O
O
OH
PG1
5
PG1 is a protecting group
Formula E1
with a carboxylic acid activator. The carboxylic acid activator may be selected from
the N-hydroxysuccinimide (HOSu), N-hydroxy-5-norbornene2,3-dicarboximide
10 (HONB), 1-hydroxybenzotriazole (HOBt), 6-chloro-1-hydroxybenzotriazole, (6-ClHOBt), 1-hydroxy-7-azabenzotriazole (HOAt), 3-hydroxy-4-oxo-3,4-dihydro-1,2,3-
benzotriazine (HODhbt), ethyl 1-hydroxy-1H-1,2,3-triazole-4-carboxylate (HOCt)
and N-hydroxytetrazole (HOt).
15 In a preferred embodiment, the compound of Formula E is prepared by reacting a
compound of Formula E1 with 1-hydroxybenzotriazole. More preferably, the reaction
is carried out in the presence of a coupling agent, optionally in the presence of a
solvent. The coupling agent may be selected from benzotriazol-1-yloxytripyrrolidinophosphonium hexafluorophosphate (PyBOP), 3-
20 [Bis(dimethylamino)methyliumyl]-3H-benzotriazol-1-oxide hexafluorophosphate
(HBTU), 2-(1H-Benzotriazole-1-yl)-1,1,3,3-tetramethylaminium tetrafluoroborate
(TBTU), N-Ethyl-N'-(3-dimethylaminopropyl)carbodiimide hydrochloride, N,N′-
Dicyclohexylcarbodiimide and N,N′-Diisopropylcarbodiimide, preferably, N-(3-
Dimethylaminopropyl)-N’-ethylcarbodiimide hydrochloride. The solvent may be a
25 nitrile solvent selected from acetonitrile and propionitrile or mixture thereof,
preferably acetonitrile.
In an embodiment, the compound of Formula E1 is reacted with 1-
Hydroxybenzotriazole in the presence of N-(3-Dimethylaminopropyl)-N′-
30 ethylcarbodiimide hydrochloride in acetonitrile at an ambient temperature to obtain a
compound of Formula EB;
21
NH O
O
O
O
N
N
N
PG1
Formula EB
wherein PG1 is a protecting group.
The reaction mixture is stirred for 2 to 3 hours. The compound of Formula EB can be
5 obtained by evaporating the solvent. Thus, one aspect of the present invention
provides a novel compound of Formula EB.
Surprisingly, the inventors of the present invention have found that the compound of
Formula E prepared by the present invention does not need to be purified as described
10 in the prior art, but can be isolated by evaporation of the solvent, if present. It has
been also found that the compound of Formula E prepared by the present invention
can directly be used (i.e. without isolating) for the preparation of the compound of
Formula F. This avoids stringent operations of the isolation and makes the process
for the preparation of compound of Formula I very cost effective.
15
Thus, the present invention has an advantage to provide a process for the preparation
of a compound of Formula E, where reaction mixture can directly be used for the
preparation of a compound of Formula 1.
20 The compound of Formula F can be converted to the compound of Formula 1 by the
process steps of the present invention. The process comprises a step of de-protection
of the compound of Formula F.
The de-protection of the compound of Formula F can be carried out in the presence of
25 an acid, optionally in the presence of a solvent. Preferably, the deprotection is carried
out using 1 to 10 equivalents of acid, more preferably 1 to 3 equivalents at ambient
temperature for at least 1 minute, preferably over a time period ranging from 1 minute
to 5 hours, more preferably from 1 to 2 hours.
22
The acid may be selected from the group comprising mineral acids, such as
hydrochloric acid, hydrobromic acid, phosphoric acid, metaphosphoric acid, nitric
acid and sulphuric acid, and organic acids, such as tartaric acid, acetic acid,
trifluoroacetic acid, citric acid, malic acid, lactic acid, fumaric acid, benzoic acid,
5 glycolic acid, gluconic acid, succinic acid, alkylsulphonic acids such as
methanesulphonic, ethanesulphonic acids, ethane-1, 2-disulfonic acid and 2-
hydroxyethanesulfonic acid, arylsulphonic acids, such as benzene sulfonic acid, 2-
naphthalenesulfonic acid, p-toluenesulphonic acid and naphthalene-1,5-disulfonic
acid; preferably p-toluenesulphonic acid is used in a solvent such as tetrahydrofuran,
10 ethyl acetate, dichloromethane and acetonitrile. In a preferred embodiment, the deprotection is carried out by treating a compound of Formula F with ptoluenesulphonic acid in the presence of a solvent such as dichloromethane. The
reaction mixture is stirred for 1 to 2 hours. The solvent is evaporated to obtain a white
solid compound. In one aspect of the invention, a solid compound is obtained after
15 evaporation of the solvent.
The obtained compound can be converted to a compound of Formula 1 by any method
known in the art, for example, using a process as reported in CN104356224.
20 In one embodiment, the obtained compound is converted to the compound of Formula
1 by the process steps as per the present invention. The process comprises a step of
reacting the solid compound obtained after de-protection of compound of Formula F
with a compound of Formula G;
O
O
O
NH
O O
O
O
X
25
Formula G
Wherein, COOX is an activated ester
in the presence of a base.
23
The base may be selected from the group comprising triethylamine, N′Ndiisopropylethylamine, sodium bicarbonate and sodium carbonate, preferably the base
is sodium bicarbonate.
5 The reaction may be carried out in a solvent. The solvent may be selected from the
group comprising N-dimethylformamide, dimethylsulfoxide, ethers, such as
tetrahydrofuran, dioxane, 1,2-dimethoxyethane or the like, esters, such as methyl
acetate, ethyl acetate or the like, ketones, such as acetone, diisobutyl ketone,
cyclohexanone, methylcyclohexanone, methyl ethyl ketone, acetylacetone or the
10 like,nitriles, such as acetonitrile or the like, alcohols, such as methanol, ethanol, tbutanol, isopropanol or the like, water and mixtures thereof, preferably acetonitrile
and water.
In a preferred embodiment, the compound obtained after de-protection of the
15 compound of Formula F is reacted with a compound of Formula G in acetonitrile and
water in the presence of sodium carbonate at an ambient temperature. The compound
of Formula 1 can be isolated by adjusting pH of the solution 3-4 using diluted HCl or
phosphoric acid. Solvent was evaporated and compound can be extracted using
ethylacetate. The compound can be recrystallized using diisopropylether to give a
20 white solid. The process of the present invention provides compound of Formula 1
with high yield.
The compound of Formula G used in the process can be prepared by any method
known in the art, for example, using a process as reported in CN104356224.
25
In one embodiment, the compound of Formula G is prepared by the process steps as
per the present invention. The process comprises a step of reacting a compound of
Formula K,
O
O
O
NH O
O
HO O
30
24
Formula K
with a carboxylic acid activator.
5 The carboxylic acid activator may be selected from the N-hydroxysuccinimide
(HOSu), N-hydroxy-5-norbornene2,3-dicarboximide (HONB), 1-
hydroxybenzotriazole (HOBt), 6-chloro-1-hydroxybenzotriazole, (6-Cl-HOBt), 1-
hydroxy-7-azabenzotriazole (HOAt), 3-hydroxy-4-oxo-3,4-dihydro-1,2,3-
benzotriazine (HODhbt), ethyl 1-hydroxy-1H-1,2,3-triazole-4-carboxylate (HOCt)
10 and N-hydroxytetrazole (HOt).
In a preferred embodiment, the compound of Formula G is prepared by reacting a
compound of Formula K with N-hydroxysuccinimide (HOSu). More preferably, the
reaction is carried out in the presence of a reagent selected from benzotriazol-1-yl15 oxytripyrrolidinophosphonium hexafluorophosphate (PyBOP), 3-
[Bis(dimethylamino)methyliumyl]-3H-benzotriazol-1-oxide hexafluorophosphate
(HBTU), 2-(1H-Benzotriazole-1-yl)-1,1,3,3-tetramethylaminium tetrafluoroborate
(TBTU), N-Ethyl-N'-(3-dimethylaminopropyl)carbodiimide hydrochloride, N,N′-
Dicyclohexylcarbodiimide and N,N′-Diisopropylcarbodiimide, preferably, N-(3-
20 Dimethylaminopropyl)-N’-ethylcarbodiimide hydrochloride, optionally in the
presence of a solvent.
The solvent may be selected from the group comprising of N-dimethylformamide,
dimethylsulfoxide, aromatic hydrocarbons such as toluene, benzene, xylene or the
like or saturated hydrocarbons such as cyclohexane, hexane or the like or halogenated
25 hydrocarbons such as dichloromethane, chloroform, 1,2-dichloroethane or the like or
ethers such as tetrahydrofuran, diethylether, dioxane, 1,2-dimethoxyethane or the like
or esters such as methyl acetate, ethyl acetate or the like or ketones such as acetone,
diisobutyl ketone, cyclohexanone, methylcyclohexanone, methyl ethyl ketone, methyl
isobutyl ketone, acetylacetone or the like or nitriles such as acetonitrile or the like or
30 alcohols such as methanol, ethanol, t-butanol, isopropanol or the like or water or
ethers such as tetrahydrofuran, diethylether, dioxane or the like or in a mixed solvent
thereof or the like, preferably dichloromethane.
25
In a preferred embodiment, the compound of Formula K is reacted with NHydroxybenzotriazole in the presence of N-(3-Dimethylaminopropyl)-N′-
ethylcarbodiimide hydrochloride in dichloromethane at an ambient temperature.
5 The compound of Formula G can be isolated by washing the reaction mixture with
sodium bicarbonate and brine solution. The residue is concentrated under vacuum and
crystallized using diisopropyl ether.
The inventors of the present invention surprisingly found that the process of the
10 present invention avoids long and tedious work up procedure for the isolation and
crystallization of the compound of Formula G.
The compound of Formula K used in the process can be prepared by using methods
known in the art, for example, using a process as reported in CN104356224.
15
The compound of Formula K is prepared by coupling a compound of Formula H;
O
O
O
OH
Formula H
by activation of the carboxylic acid, with a compound of Formula J
20
O OH
O O
NH2
Formula J
The activation of the carboxylic acid is carried out in the presence of a carboxylic acid
25 activator selected from N-hydroxysuccinimide (HOSu), N-hydroxy-5-norbornene2,3-
dicarboximide (HONB), 1-hydroxybenzotriazole (HOBt), 6-chloro-1-
hydroxybenzotriazole, (6-Cl-HOBt), 1-hydroxy-7-azabenzotriazole (HOAt), 3-
hydroxy-4-oxo-3,4-dihydro-1,2,3-benzotriazine (HODhbt), ethyl 1-hydroxy-1H-1,2,3-
triazole-4-carboxylate (HOCt) and N-hydroxytetrazole (HOt).
26
Preferably, the coupling is carried out in the presence of a base selected from
triethylamine, N′N-diisopropylethylamine, sodium bicarbonate and sodium carbonate,
preferably sodium bicarbonate.
5
The reaction may be carried out in a solvent. The solvent may be selected from the
group comprising of N-dimethylformamide, dimethylsulfoxide, aromatic
hydrocarbons such as toluene, benzene, xylene or the like or saturated hydrocarbons
such as cyclohexane, hexane or the like or halogenated hydrocarbons such as
10 dichloromethane, chloroform, 1,2-dichloroethane or the like or ethers such as
tetrahydrofuran, diethylether, dioxane, 1,2-dimethoxyethane or the like or esters such
as methyl acetate, ethyl acetate or the like or ketones such as acetone, diisobutyl
ketone, cyclohexanone, methylcyclohexanone, methyl ethyl ketone, methyl isobutyl
ketone, acetylacetone or the like or nitriles such as acetonitrile or the like or alcohols
15 such as methanol, ethanol, t-butanol, isopropanol or the like or water or ethers such as
tetrahydrofuran, diethylether, dioxane or the like or in a mixed solvent thereof or the
like.
In one embodiment of the process, an intermediate compound of Formula I is isolated
20 during the process for the preparation of a compound of Formula K. The process for
the preparation of the compound of Formula I comprises a reaction of the compound
of Formula H with a carboxylic acid activator in the presence of N-(3-
Dimethylaminopropyl)-N’-ethylcarbodiimide hydrochloride.
25 The reaction may be carried out in a solvent. The solvent is selected from
dichloromethane, chloroform, acetone, acetonitrile, tetrahydrofuran,
dimethylsulphoxide, dimethylformamide and mixtures thereof, preferably
dichloromethane.
30 The inventors of the present invention found that the compound of Formula I prepared
by the process of present invention can be isolated by following a simple procedure.
27
The resulting compound of Formula I can be isolated using dichloromethane and
concentrated under vacuum.
O
O
O
O
X
5 Formula I
Wherein, COOX is an activator ester
Preferably, the compound of Formula I can be crystallized using isopropyl alcohol.
The process known in the art for the isolation of the compound of Formula I require
10 tedious work-up procedure which require several solvent washings and subsequently
water washings, which results in the loss of yield of the product.
Thus, one aspect of the invention provides a process for the preparation of a
compound of Formula I.
15
The compound of Formula I is reacted with a compound of Formula J in the presence
of sodium bicarbonate.
The reaction may be carried out in a solvent. The solvent may be selected from the
20 group comprising of N-dimethylformamide, dimethylsulfoxide or ethers such as
tetrahydrofuran, dioxane, 1,2-dimethoxyethane or the like or esters such as methyl
acetate, ethyl acetate or the like or ketones such as acetone, diisobutyl ketone,
cyclohexanone, methylcyclohexanone, methyl ethyl ketone, acetylacetone or the like
or nitriles such as acetonitrile or the like or alcohols such as methanol, ethanol, t25 butanol, isopropanol or the like or water or ethers such as tetrahydrofuran, dioxane or
the like or in a mixed solvent thereof or the like or mixture thereof, preferably
tetrahydrofuran and water.
28
In a preferred embodiment, the compound of Formula I is reacted with a compound of
Formula J in the presence of sodium bicarbonate in tetrahydrofuran and water to
obtain a compound of Formula K. The reaction mixture is allowed to stir for 1 to 2
hours.
5
Thus, one aspect of the invention provides a process for the preparation of a
compound of Formula K.
EXPERIMENTAL
10 Detailed experimental parameters according to the present invention are provided by
the following examples, which are intended to be illustrative and not limiting of all
possible embodiments of the invention.
Examples
15 Example-1
Preparation of N6
-(2,2-dimethyl-4,13-dioxo-3,8,11-trioxa-5-azatridecan-13-yl)-
N2
-{[(9H-fluoren-9-yl)methoxy]carbonyl}-L-lysine
O NH O
O
O
O
OH
O
O
HN
HOOC
NH2
O NH O
O
O
O
NH
O
OH
HN
O
O
2. DIPEA/DM Water
1. ACN, EDC.HCl,HOBt
20
EDC.HCl (9.46 g) was added to the mixture of 2,2-dimethyl-4-oxo-3,8,11-trioxa-5-
azatridecan-13-oic acid (10.0 g) and N-Hydroxy benzotriazole (5.13 g) in 50 ml of Ndimethylformamide at ambient temperature. The resulting mixture was stirred for 3
hours at ambient temperature. This reaction mass was added drop-wise to the mixture
of N
2
25 -{[(9H-fluoren-9-yl)methoxy]carbonyl}-L-lysine (14.0 g) and N,N-diisopropyl
ethyl amine (10 g) in water (50 ml). After reaction completion, pH of the reaction
mixture was adjusted to 3.0-3.5 with diluted phosphoric acid. Solvent was removed
29
by evaporation and reaction mixture was extracted in ethyl acetate. Solid was isolated
by addition of Diisopropyl ether followed by filtration and drying (18.5 g). Reaction
yield is ca. 80% (Purity 99%).
Example-2:
Preparation of N6 5 -(2,2-dimethyl-4,13,22-trioxo-3,8,11,17,20-pentaoxa-5,14-
diazadocosan-22-yl)-N2
-{[(9H-fluoren-9-yl)methoxy]carbonyl}-L-lysine
O NH
O
O
O
O
NH
O
OH
HN
O
O
H3N
+ O
O
O
NH
O
OH
HN
O
O
O NH
O
O
O
O
OH O NH
O
O
O
O
O
N
N
N
O NH O
O
O
O
NH O
O
O
HN
O
OH
HN O
O
Dichloromethane
N
6
-(2,2-dimethyl-4,13,22-trioxo-3,8,11,17,20-pe
ntaoxa-5,14-diazadocosan-22-yl)-N
2
-{[(9H-fluor
en-9-yl)methoxy]carbonyl}-L-lysine
Trifluoroacetic acid
Sodium
bicarbonate/Water
O
F
F
F
O
–
2,2-dimethyl-4-oxo-3,8,11-trioxa
-5-azatridecan-13-oic acid
Acetonitrile/EDC.HCl
HOBT
Solution B
Solution A
A: 10.0 g of N
6
-(2,2-dimethyl-4,13-dioxo-3,8,11-trioxa-5-azatridecan-13-yl)-N
2
10 -
{[(9H-fluoren-9-yl)methoxy]carbonyl}-L-lysine was dissolved in 25 ml of
dichloromethane. The solution was treated with Trifluoro acetic acid (25 mL) at
ambient temperature. Reaction mixture was stirred for 2 hours at ambient temperature
followed by removal of solvent to give white solid. To this white solid, 25 ml of water
15 and Sodium bicarbonate (2.74 g) was added to make solution A.
B: In another flask, EDC.HCl (9.46 g) was added to the mixture of 2,2-dimethyl-4-
oxo-3,8,11-trioxa-5-azatridecan-13-oic acid (10.0 g) and N-Hydroxy benzotriazole
30
(5.13 g) in 50 ml of acetonitrile at ambient temperature. The resulting mixture was
stirred for 3 hours at ambient temperature.
This solution B was added to the solution A at ambient temperature. The reaction
5 mixture was stirred 1-2 hours and pH of the reaction mixture was adjusted to 3.0-3.5
with diluted phosphoric acid. Solvent was removed by evaporation and reaction
mixture was extracted in ethyl acetate. Solid was isolated by addition of diisopropyl
ether followed by filtration and drying (10.0 g). Reaction yield is ca. 90% (Purity 98.5
%).
10
Example-3:
Preparation of tert-butyl 18-[(2,5-dioxopyrrolidin-1-yl)oxy]-18-oxooctadecanoate
EDC.HCl O
O
O
OH
O
O
O
O
N
O
O
18-(tert-butoxy)-18-oxooctadecanoic acid
HO
N
O
O
Dichloromethane
tert-butyl 18-[(2,5-dioxopyrrolidin-1-yl)oxy]-18-
oxooctadecanoate
15 EDC.HCl (20.2 g) was added to a mixture of 18-(tert-butoxy)-18-oxooctadecanoic
acid (30.0 g) and N-Hydroxy succinimide (12.1 g) in dichloromethane (300 mL). The
mixture was stirred at ambient temperature. The reaction mixture was washed with
sodium bicarbonate solution. After separation, the aqueous layer was extracted with
dichloromethane (150 mL). Organic layer was washed with brine solution and
20 concentrated under vacuum. The residue was crystallized with Isopropyl alcohol to
afford tert-butyl 18-[(2,5-dioxopyrrolidin-1-yl)oxy]-18-oxooctadecanoate (36.0 g).
Reaction yield is ca. 95% (Purity 99.5 %).
Example-4:
25 Preparation of (4S)-5-tert-butoxy-4-[(18-tert-butoxy-18-oxooctadecanoyl)amino]-
5-oxopentanoic acid
31
O
O
O
O
N
O
O
O OH
O O
NH2
Sodium
bicarbonate
+
O
O
O
NH O
O
HO O
(4S)-4-amino-5-tert-butoxy-5-
oxopentanoic acid
tert-butyl 18-[(2,5-dioxopyrrolidin-1-yl)oxy]-
18-oxooctadecanoate
THF/Water
(4S)-5-tert-butoxy-4-[(18-tert-butoxy-18-oxooctadecanoyl)amino]
-5-oxopentanoic acid
Sodium bicarbonate (1.8 g) was added to a mixture of tert-butyl 18-[(2,5-
dioxopyrrolidin-1-yl)oxy]-18-oxooctadecanoate (5.0 g) and (4S)-4-amino-5-tertbutoxy-5-oxopentanoic acid (2.8 g) in THF and water mixture. The mixture was
5 stirred at ambient temperature for 1-2 hours. THF was removed by evaporation and
residue was diluted with water (50 mL) and pH was adjusted to 2.0-3.0. Product was
extracted with ethyl acetate. The resulting mixture was concentrated under vacuum.
The residue was crystallized with n-Heptane to afford (4S)-5-tert-butoxy-4-[(18-tertbutoxy-18-oxooctadecanoyl)amino]-5-oxopentanoic acid (5.5 g). Reaction yield is ca.
10 92% (Purity 99%).
Example-5:
Preparation of 1-tert-butyl 5-(2,5-dioxopyrrolidin-1-yl) N-(18-tert-butoxy-18-
oxooctadecanoyl)-L-glutamate
EDC.HCl
O
O
O
NH O
O
HO O HO
N
O
O
N
O
O
O
O
O
NH
O O
O
O
Dichloromethane
(4S)-5-tert-butoxy-4-[(18-tert-butoxy-18-oxooctadecanoyl)
amino]-5-oxopentanoic acid
1-tert-butyl 5-(2,5-dioxopyrrolidin-1-yl) N-(18-tert-butoxy18-oxooctadecanoyl)-L-glutamate
15
EDC.HCl (0.448 g) was added to a mixture of (4S)-5-tert-butoxy-4-[(18-tert-butoxy18-oxooctadecanoyl)amino]-5-oxopentanoic acid (1.0 g) and N-Hydroxy succinimide
(0.269 g) in dichloromethane. The mixture was stirred at ambient temperature till
20 reaction completion. Reaction mixture was washed with sodium bicarbonate solution
32
followed by brine washing and concentration under vacuum. The residue was
crystallized in diisopropyl etherto afford 1-tert-butyl 5-(2,5-dioxopyrrolidin-1-yl) N-
(18-tert-butoxy-18-oxooctadecanoyl)-L-glutamate (1.0 g). Reaction yield is ca. 94%
(Purity 99.5 %).
5 Example-6:
Preparation of Semaglutide Side chain
NH
O
O
O
NH O
O
O
NH
O
OH
NH
O
O O
O
N
O
O
O
O
O
NH
O O
O
O
O
O
O
NH
O
O O
NH
O
O
O
NH O
O
O
HN
O
OH
HN O
O
+
Semaglutide side chain
1-tert-butyl 5-(2,5-dioxopyrrolidin-1-yl) N-(18-
tert-butoxy-18-oxooctadecanoyl)-L-glutamate
1.TFA/DCM
2.Sodium bicarbonate
Acetonitrile
DM water
S
S
S
S
N
6
-(2,2-dimethyl-4,13,22-trioxo-3,8,11,17,20-pentaoxa-5,14-diaz
adocosan-22-yl)-N
2
-{[(9H-fluoren-9-yl)methoxy]carbonyl}-L-lysine
10.0 g of N
6
-(2,2-dimethyl-4,13,22-trioxo-3,8,11,17,20-pentaoxa-5,14-diazadocosan22-yl)-N
2
10 -{[(9H-fluoren-9-yl)methoxy]carbonyl}-L-lysine was dissolved in 25 ml of
dichloromethane. Solution was treated with Trifluoroacetic acid (25 mL) at ambient
temperature. After completion of reaction, solvent was removed by evaporation, to
give white solid. To this white solid, 25 ml of water and 2.21 g of Sodium bicarbonate
was added followed by addition of 1-tert-butyl 5-(2,5-dioxopyrrolidin-1-yl) N-(18-
15 tert-butoxy-18-oxooctadecanoyl)-L-glutamate (8.6 g) in 50 ml of acetonitrile at
ambient temperature. After completion of reaction, pH of reaction mixture was
adjusted to 3.0-3.5 with diluted phosphoric acid. Acetonitrile was removed by
evaporation and reaction mixture was extracted with ethyl acetate. To this diisopropyl
ether was added to give a white solid, (12.6 g). Reaction yield is ca. 90% (Purity 98.5
20 %).

We Claim:

1. Process for the preparation of the side chain of Semaglutide of Formula 1,
O
O
O
NH
O
O O
NH
O
O
O
NH O
O
O
HN
O
OH
HN O
O
5 Formula 1
comprising the steps of:
a) coupling a compound of Formula A;
NH
O
O
O
OH
PG1
10
Formula A
wherein PG1 is a protecting group, cleavable under non-basic conditions
by activation of the carboxylic acid, with a compound of Formula B;
O
O
HN
HOOC
NH2
15
Formula B
to obtain a compound of Formula C;
NH
O
O
O
NH
O
OH
HN
O
O
PG1
34
Formula C
b) de-protecting the compound of Formula C to obtain a compound of Formula D or its
salt;
O
O
O
NH
O
OH
HN
O
O
H2N
5
Formula D
c) reacting the compound of Formula D or its salt with a compound of Formula E;
NH O
O
O
O
X
PG1
Formula E
10 Wherein COOX is an activated ester
in the presence of a base, to obtain a compound of Formula F;
NH O
O
O
NH O
O
O
HN
O
OH
HN O
O
PG1
Formula F
15 d) de-protecting the compound of Formula F;
e) reacting the reaction mixture of step d) with a compound of Formula G;
35
O
O
O
NH
O O
O
O
X
Formula G
Wherein, COOX is an activated ester
in the presence of a base to obtain the compound of Formula 1;
5 Wherein:
i) the compound of Formula E used in step c) is prepared by a process
comprising the reaction of a compound of Formula E1;
NH O
O
O
OH
PG1
10
Formula E1
with a carboxylic acid activator;
and,
ii) the compound of Formula G used in step e) is prepared by a process comprising the
15 steps of:
f1) coupling a compound of Formula H;
O
O
O
OH
Formula H
20 by activation of the carboxylic acid, with a compound of Formula J
O OH
O O
NH2
Formula J
to obtain a compound of Formula K;
36
O
O
O
NH
O
O
HO O
Formula K
f2) reacting the compound of Formula K with a carboxylic acid activator to obtain the
compound of Formula G.
5
2. The process according to claim 1, wherein the base of step c) and step e) is
independently selected from triethylamine, N′N-diisopropylethylamine, sodium
bicarbonate and sodium carbonate, preferably sodium bicarbonate.
10 3. Process for the preparation of a compound of Formula C;
NH O
O
O
NH
O
OH
HN
O
O
PG1
Formula C
wherein PG1 is a protecting group, cleavable under non-basic conditions
comprising the steps of:
15 a) reacting a compound of Formula A;
NH
O
O
O
OH
PG1
Formula A
20 with a carboxylic acid activator in the presence of N-(3-Dimethylaminopropyl)-N′-
ethylcarbodiimide hydrochloride in a polar aprotic solvent selected from N,N-
37
dimethylformamide, dimethyl sulfoxide and tetrahydrofuran, preferably
dimethylformamide;
b) reacting the reaction mixture of step a) with a compound of Formula B;
5
O
O
HN
HOOC
NH2
Formula B
in the presence of N′N-diisopropylethylamine to obtain a compound of Formula C.
10 4. Use of the compound of Formula C prepared according to claim 3 for the preparation of
side chain of Semaglutide of Formula 1;
O
O
O
NH
O
O O
NH
O
O
O
NH O
O
O
HN
O
OH
HN O
O
Formula 1.
15 5. Process for the preparation of a compound of Formula G;
O
O
O
NH
O O
O
O
X
Formula G
Wherein, COOX is an activated ester
20 comprising a reaction of a compound of Formula K;
38
O
O
O
NH
O
O
HO O
Formula K
with a carboxylic acid activator in the presence of N-(3-Dimethylaminopropyl)-N′-
ethylcarbodiimide hydrochloride.
5
6. The process according to claim 5, wherein the reaction is carried out in the presence of
a solvent selected from the group consisting of dichloromethane, chloroform, acetone,
acetonitrile, tetrahydrofuran, dimethylsulphoxide and dimethylformamide or mixture
thereof, preferably dichloromethane.
10
7. Use of the compound of Formula G prepared according to claim 5 or 6 for the
preparation of side chain of Semaglutide of Formula 1;
O
O
O
NH
O
O O
NH
O
O
O
NH O
O
O
HN
O
OH
HN O
O
Formula 1.
15 8. Process for the preparation of a compound of Formula I;
O
O
O
O
X
Formula I
Wherein, COOX is an activator ester
20 comprising the reaction of a compound of Formula H;
O
O
O
OH
Formula H
39
with a carboxylic acid activator in the presence of N-(3-Dimethylaminopropyl)-N′-
ethylcarbodiimide hydrochloride in a solvent to obtain a compound of Formula I.
9. The process according to claim 8, wherein the solvent is selected from the group
5 consisting of dichloromethane, chloroform, acetone, acetonitrile, tetrahydrofuran,
dimethylsulphoxide and dimethylformamide or mixture thereof, preferably
dichloromethane.
10. Use of the compound of Formula I prepared according to claim 8 or 9 for the
10 preparation of a compound of Formula 1;
O
O
O
NH
O
O O
NH
O
O
O
NH O
O
O
HN
O
OH
HN O
O
Formula 1.
11. A compound of Formula EB;
15
NH O
O
O
O
N
N
N
PG1
Formula EB
wherein PG1 is a protecting group.

20 12. Process for the preparation of a compound of Formula E;
NH O
O
O
O
X
PG1
Formula E
wherein PG1 is a protecting group, and COOX is an activated ester
25 comprising a reaction of a compound of Formula E1;
40
NH O
O
O
OH
PG1
Formula E1
5 with a carboxylic acid activator in the presence of N-(3-Dimethylaminopropyl)-N′-
ethylcarbodiimide hydrochloride.
13. The process according to claim 12, wherein the reaction is carried out in the presence
of a solvent selected from the group consisting of dichloromethane, chloroform,
10 acetone, acetonitrile, tetrahydrofuran, dimethylsulphoxide and dimethylformamide or
mixture thereof, preferably dichloromethane.
14. Use of the compound of Formula E according to claim 12 or 13 for the preparation of
side chain of Semaglutide of Formula 1;