Discovery of highly functionalized scaffolds: Pyrroloimidazolediones as P2X7 receptor antagonists

Article abstract of DOI:10.1016/j.tet.2017.07.036

A broad range of pyrroloimidazolediones, new highly functionalized bicyclic heterocycles, was obtained by a cycloaddition reaction between L-pyroglutamide derivatives and Bredereck's reagent. Methanolysis of subsequent pyrroloimidazolediones provided antagonists of P2X7 receptor, with retention of initial stereoconfiguration, with potential applications in the treatment of inflammatory and neurological diseases. We have thus developed a new synthesis of lactamic nitrogen free enaminones which is currently the easiest method cited in the literature to access these compounds.

Full text of DOI:10.1016/j.tet.2017.07.036

Accepted Manuscript
Discovery of highly functionalized scaffolds: Pyrroloimidazolediones as P2X7 receptor
antagonists
Germain Homerin, Emmanuelle Lipka, Benoît Rigo, Régis Millet, Xavier Dezitter,
Christophe Furman, Alina Ghinet
PII:
S0040-4020(17)30778-0
10.1016/j.tet.2017.07.036
TET 28868
DOI:
Reference:
To appear in: Tetrahedron
Received Date: 3 May 2017
Revised Date: 20 July 2017
Accepted Date: 21 July 2017
Please cite this article as: Homerin G, Lipka E, Rigo Benoî, Millet Ré, Dezitter X, Furman C, Ghinet
A, Discovery of highly functionalized scaffolds: Pyrroloimidazolediones as P2X7 receptor antagonists,
Tetrahedron (2017), doi: 10.1016/j.tet.2017.07.036.
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Discovery of highly functionalized scaffolds:
Pyrroloimidazolediones as P2X7 receptor
antagonists.
Leave this area blank for abstract info.
Germain Homerina,b, Emmanuelle Lipkaa,c, Benoît Rigoa,b, Régis Milleta,d, Xavier Dezittera,d, Christophe
Furmana,d and Alina Ghineta,b,e,
*

1
ACCEPTED MANUSCRIPT
Tetrahedron
journal homepage: www.elsevier.com
Discovery of highly functionalized scaffolds: Pyrroloimidazolediones as P2X7
receptor antagonists.
Germain Homerina,b, Emmanuelle Lipkaa,c, Benoît Rigoa,b, Régis Milleta,d, Xavier Dezittera,d, Christophe
Furmana,d and Alina Ghineta,b,e,
*
aInserm U995, LIRIC, Université de Lille, CHRU de Lille, Faculté de médecine – Pôle recherche, Place Verdun, F-59045 Lille Cedex, France
bHautes Etudes d’Ingénieur (HEI), Groupe Yncréa Hauts-de-France, UCLille, Laboratoire de pharmacochimie, 13 rue de Toul, F-59046 Lille, France
cFaculté des Sciences Pharmaceutiques et Biologiques de Lille, Laboratoire de Chimie Analytique, F-59006 Lille Cedex, France
dInstitut de Chimie Pharmaceutique Albert Lespagnol, IFR114, 3 rue du Pr Laguesse, F-59006 Lille, France
e‘Al. I. Cuza’ University of Iasi, Faculty of Chemistry, Bd. Carol I, nr. 11, 700506 Iasi, Romania
ARTICLE INFO
ABSTRACT
Article history:
Received
Received in revised form
Accepted
A broad range of pyrroloimidazolediones, new highly functionalized bicyclic heterocycles, was
obtained by a cycloaddition reaction between L-pyroglutamide derivatives and Bredereck’s
reagent. Solvolysis in methanol of subsequent pyrroloimidazolediones provided antagonists of
P2X7 receptor, with retention of initial stereoconfiguration, with potential applications in the
treatment of inflammatory and neurological diseases. We have thus developed a new synthesis
of lactamic nitrogen free enaminones which is currently the easiest method cited in the literature
to access these compounds.
Available online
Keywords:
Pyroglutamic acid
Pyroglutamide
Pipecolic acid
P2X7 receptor
Pyrroloimidazoledione
Enaminone
2009 Elsevier Ltd. All rights reserved
.
1. Introduction
enamine unit next to a carbonyl function could be obtained by
using dimethylformamide acetals (such as DMF dimethyl
acetal),8 Vilsmeier’s reagent (DMF/POCl3),9 Gold’s reagent,10 or
Bredereck’s reagent (tert-butoxy bis(dimethylamino)methane)
(BR).11
P2X7 receptors (P2X7R) are ion channels implicated in the
processing and release of pro-inflammatory cytokines such as
interleukine-1β.1 They are activated by high concentrations of
adenosine 5’-trisphosphate (ATP), and are acting as a danger
signal, where an immune response is needed, by initiating an
inflammation process.2 These receptors are involved in many
diseases such as neurodegenerative diseases (Parkinson’s and
Alzheimer’s diseases),3 and chronic inflammation (multiple
sclerosis and inflammatory bowel diseases (IBD)).2 To date, no
antagonist of P2X7R succeeded in clinical trial, mostly due to a
lack of efficiency during phase II studies; nevertheless, the
results observed on animal models are raising the hope of new
treatments targeting these receptors.4
Me
Me
Cl
N
R
H
SAR
N
N
R1
O
O
N
Cl
N
O
O
Me
R2
1
2a-d
Target compounds
GSK1370319A
Figure 1. Pharmacomodulations envisaged on P2X7R Antagonist
GSK1370319A (1).
2. Results and Discussion
Our group is interested in pyroglutamic acid derivatives as
potential drug candidates.5 Starting from L-pyroglutamic-based
P2X7R ligands described in the literature (especially from
compound 1, GSK1370319A),6 and from previous results of our
laboratory,7 we designed new potential antagonists of P2X7R.
The aim of this work was to explore the conformational space
available for P2X7R ligands, and eventually to find usable drugs
for the treatment of IBD. Thus we wanted to obtain enaminones 2
bearing a dimethylenamine moiety in position 3 of the lactam
ring (Figure 1). According to literature, the addition of an
In our hands, DMFDMA and Gold’s reagent were not reactive
enough to lead to β-(dimethylamino)-enones 2a, 2b and 2c from
pyroglutamides 1, 3 (synthesized according to a previously
described method),12 and 4a (Scheme 1). Interestingly, BR
furnished the N-aryl lactam 2b but not the N-methyl lactam 2a,
and it was though that these compounds could later be obtained
by N-methylation of dimethylaminoenamides unsubstituted on
the lactamic nitrogen (R2 = H) 2c. In order to obtain these
compounds, a β-(dimethylamino)-enone moiety needs to be
placed in position 3 of the lactam ring of pyroglutamides 4.

2
Tetrahedron
ACCEPTED MANUSCRIPT
NMe2
NMe2
NMe2
R
R
N
N
tBuOCH(NMe2)2
BR (5 equiv)
H
R1
R1
OMe
N
O
O
O
O
H2NR1
BR (5 equiv)
O
O
N
N
N
R1
N
O
O
N
H
N
O
O
120°C
24-48h
(Lit. 12)
50-110°C,
2-24h
110°C, 1.5-24h
N
R2
R2
H
N
O
O
Me2N
Cl
Me2N
R1
1
R = H, R1 = 2,4-dicholorobenzyl, R2 = methyl
R = H, R1 = 2,4-dicholorobenzyl, R2 = 2-thienyl
4a R, R2 = H, R1 = 2,4-dicholorobenzyl
2a (0%)
2b (45%)
2c (0%)
6
4a-q (16-96%)
5a-q
3
5a (75%)
Cl
NMe2
NMe2
NMe2
(isomeric excess 97%)
Scheme 1. Introduction of β-(dimethylamino)-enone moiety in position 3.
O
O
O
O
O
O
N
N
N
However, a mixture of lactam 4a and BR (5 equiv), stirred at
110°C for 1.5 hours provided unexpected pyrroloimidazoledione
5a as the unique reaction product in 75% isolated yield (Scheme
1). To the best of our knowledge, no 3-aminotetrahydro-1H-
pyrrolo[1,2-c]imidazole-1,5-dione was ever described.13 As this
scaffold could be perceived as constrained analog of P2X7R
antagonist 1, the synthetic pathway was first optimized, and
initial conditions proved to be the most efficient (Table 1). Then
the generalization to L-pyroglutamides 4b-q (synthesized from
L-methyl pyroglutamate 6, according to a method previously
described),7 afforded pyrroloimidazoledione 5b-f and 5k-m
(Scheme 2). The investigation of compounds 5 by chiral
N
N
N
Me2N
Cl
Me2N
Me2N
Me
5a (75%)
5b (47%)
5c (14%)
Cl
O
NMe2
NMe2
NMe2
O
O
O
O
O
N
N
N
N
N
N
OMe
Me2N
Me2N
Me2N
5f (47%)
5d (29%)
5e (52%)
MeO
MeO
Cl
NMe2
NMe2
NMe2
O
O
O
O
O
O
N
N
N
Cl
N
N
OMe
N
supercritical fluid chromatography (SFC) indicated
a
Me2N
Me2N
Me2N
5g (0%)
5h (0%)
5i (0%)
stereoisomeric mixture of two diastereoisomers except for
compound 5c for which an asymmetric induction was observed
(see ‘Supporting Information’ section).
Cl
NMe2
NMe2
NMe2
OMe
O
O
O
O
O
O
N
N
N
The results obtained by varying amounts of BR, temperature,
duration of the reaction, and solvent-free versus toluene
conditions are reported in Table 1. Because two equivalents of
BR are necessary – one for cyclization to imidazolone, and one
for the introduction of the β-(dimethylamino)-enone moiety – and
since BR has autoreactive behavior,11c a large excess was
privileged to provide target pyrroloimidazoledione 5a (Table 1,
entries 1-5 versus entries 6-8). Raising the temperature
accelerates the reaction (Table 1, entry 4 versus entry 5) without
degrading the medium up to 120 °C (data not shown), while
dilution in toluene gradually decreases the reaction progress
(Table 1, entry 6 versus entry 7). Finally, the most suitable
conditions for the synthesis of 5a were to perform a solvent-free
reaction in presence of 5 equiv of BR at 110 °C for 1.5h (Table 1,
entry 8).
N
N
N
Me2N
Me2N
Me2N
OMe
5j (0%)
5k (6%)
5l (55%)
MeO
N
OMe
OMe
NMe2
NMe2
NMe2
O
O
O
O
O
O
N
N
N
N
N
N
Cl
Cl
Me2N
5m (7%)
Me2N
Me2N
5o (0%)
5n (0%)
NMe2
NMe2
O
O
O
O
N
N
N
N
Me2N
Me2N
nC6H13
5p (0%)
5q (50%)
Table 1. Optimization of pyrroloimidazoledione 5a synthesis
Scheme 2. Synthesis of pyrroloimidazolediones (5a-q).
NMe2
Cl
Our optimized pyrroloimidazoledione ring closure protocol
was further investigated on N-benzylpyroglutamides 4b-f
(Scheme 2). Overall, yields for this procedure were generally
good. The moderate yields observed for isolated compounds are
not in accordance to quantitative conversions of 4b-f into 5b-f,
observed in the 1H NMR of the crudes, and were due to
O
O
H
N
BR
Cl
N
N
O
Solvent, temp, t
N
H
Me2N
Cl
O
4a
5a
Cl
1H NMR
conversion
rate of 4a
(%)
Equiv
of BR
Entry
Solvent Temp (°C) t (h)
chromatographic
purification
difficulties.
In
the
(hetero)arylpyroglutamides series 5g-l, as it could be expected,
electron-rich arene groups improved reactions yields, while steric
hindrance or low nucleophilicity of starting amides has prevented
the preparation of pyrroloimidazoledione 5g-j, 5o and 5p. For
unexplained reasons, the extension from N-benzyl to N-(2-
phenyl)ethyl resulted in poor yield for pyrroloimidazoledione 5m
and traces for 5n. Finally, the alkylpyroglutamide 4q engaged
1
2
3
4
5
6
7
8
1
1
-
RT
50
6
6
30
25
-
1.5
2
-
50
3
57
-
110
50
10
24
12
16
1.5
75
effectively
in
the
cyclization
reaction
to
afford
2.5
5
-
65
pyrroloimidazoledione 5q in 50% isolated yield.
-
50
85
5
toluene
-
50
80
With the BR ring closure methodology in our hands, we
extended it to other amides such as N-(2,4-dichlorobenzyl)-6-
oxopiperidine-2-carboxamide 7, synthesized from commercially
5
110
>95
available (2S)-6-oxopipecolic acid
8 by peptide coupling
(Scheme 3). The ring closure was effective, with total conversion
of the initial amide to the corresponding imidazopyridine-1,5-
dione 9 (observed as the major product by 1H NMR of the crude).

3
However, only the ring-opened (5E)-N-(2,4-dichlorobenzyl)-5-
[(dimethylamino)methylene]-6-oxopiperidine-2-carboxamide 10
was isolated after flash chromatography purification.
(scheme 7) which is based on the low reactivity of
ACCEPTED MANUSCRIPT
DMFDMA which was however systematically observed in the
reaction medium while performing 1H NMR analysis.
OH
O
N
H
O
8
H2NR1, EDCI
rt, 24 h
NMe2
NMe2
Cl
Cl
BR
H
N
H
O
O
N
N
O
N
O
N
110°C, 24 h
H
H
N
O
Cl
O
Cl
Me2N
Cl
7 (62%)
9 (0%)
10 (16%)
Cl
Scheme 3. Generalization of the protocol.
We have then performed additional experiments to propose a
mechanism for this ring closure reaction: when mild conditions
were utilized with amide 4q, by using only one equivalent of BR
at 50 °C, a mixture of intermediate 11 (20% yield) and final
product 5q (14% yield) was obtained (Scheme 4); moreover, the
Vilsmeier-Haak reaction of pyroglutamide 4a resulted in N-
formyl lactam 12 as the single product isolated in a low 5% yield
(Scheme 4), confirming that position 1 is the most reactive
towards electrophiles. Consequently, the first step in the
synthesis of pyrroloimidazolediones 5 seems to pass through the
formation of an N-acyliminium ion A which cyclizes to bicyclic
imidazolone B, and then a classical Bredereck’s reaction
Scheme 7. Proposed mechanism for methanol assisted ring opening of
pyrroloimidazolediones 5.
This methanol-assisted opening of the imidazolone ring opens
the way to potential P2X7R ligands modified in position 3 of the
lactam scaffold. Indeed, substitution of the dimethylamino group
of pyrroloimidazoledione 5a by primary and secondary amines
thus provided L-pyroglutamides 13a-e (Scheme 8). Interestingly,
the pyrroloimidazoledione scaffold and the dimethylamino group
of compound 5a were hydrolyzed by refluxing in acidified water,
thus affording enol 14. In addition, the reaction of compound 5a
with nitrous acid,11a provided the ring-opened oxime 15. We then
successfully performed sodium periodate-assisted oxidation of
the dimethylenamine moiety of compound 5a. In this case, the
ring opening was not observed thus providing the
pyrroloimidazoletrione 16 (in equilibrium with its enol form, see
‘Supporting Information section).
1
pathway1 a leads to the introduction of the β-(dimethylamino)-
enone moiety in α-position of the lactam ring (Scheme 5).
NMe2
H
N
BR (1 equiv)
O
O
O
O
nC6H13
N
N
O
+
N
50°C
6h
N
H
N
O
Me2N
nC6H13
Me2N
nC6H13
Cl
4q
11 (20%)
5q (14%)
Cl
Noteworthy, the chiral chromatography of compound 2c
confirmed that ring-closure to condensed imidazolones 5 then
opening to pyroglutamides 2 and 13, occurred with retention of
chirality.
+
Me2N=CHCl Cl
(1 equiv)
H
H
N
N
Cl
Cl
O
O
DMF, RT
48h
N
H
N
O
O
O
OH
4a
12 (5%)
Cl
O
N
NaNO2, CH3CO2H
THF, 0°C, 3h
NaIO4
Scheme 4. Isolation of intermediates.
H
H2O / EtOAc / CH3CN
rt, 1h
N
O
O
NMe2
O
N
N
NMe2
H
Cl
O
N
15 (77%)
H
Me2N
Cl
H
N
N
BR
BR
O
O
R
R
N
O
O
16 (34%)
O
O
O
O
N
N+
N
N
Cl
H
N
O
O
N
N
Me2N
Me2N
Cl
Me2N
Me2N
R
R
5a
4
A
B
5
R1
Cl
N
R2
Scheme 5. Possible reaction mechanism.
Cl
OH
Cl
MeSO3H
H
HNR1R2
H
N
N
H2O,
reflux, 24h
toluene,
reflux, 24h
O
Now, the target L-pyroglutamide 2c (72%) was obtained by
stirring pyrroloimidazoledione 5a for 8 hours in refluxing
methanol (Scheme 6), and a similar opening of the imidazole ring
was observed after elution of crude pyrroloimidazoledione 5k
(Scheme 2) on a silica gel column by using methanol as the co-
solvent with dichloromethane. Indeed, this purification allowed
isolation of only 5% of 5k, accompanied by 30% of lactam 2d
(Scheme 6). To the best of our knowledge, we provided here the
easiest synthetic method to access pyroglutamic enaminones
unsubstituted on the lactamic nitrogen.
N
O
N
H
Cl
H
Cl
O
O
O
14 (40%)
13a-e
Cl
O
O
O
Cl
NH
O
Cl
13c (65%)
H
NH
N
Cl
O
13a (53%)
N
H
Cl
O
H
N
MeO
MeO
O
O
N
H
Cl
O
N
NH
Cl
13d (21%)
13b (82%)
Cl
N
MeO
NMe2
NMe2
H
N
H
N
O
N
H
O
H
N
Cl
i) MeOH / Reflux
N
O
R
Me
H
Cl
O
O
O
O
N
or ii) MeOH / SiO2 / rt
N
H
O
N
Me2N
R
N
5a R = 2,4-dichlorobenzyl
5k R = 4-methoxyphenyl
2c R = 2,4-dichlorobenzyl (72%)
2d R = 4-methoxyphenyl (30%)
13e (67%)
Cl
H
N
Scheme 6. Ring opening by methanolysis.
O
N
H
Cl
O
Scheme 8. Reactivity of pyrroloimidazoledione 5a.
A potential mechanistic pathway has been proposed for the
methanol assisted ring opening of pyrroloimidazolediones 5

4
Tetrahedron
ACCEPTED MANUSCRIPT
3. Conclusions
21
13e
49.2 ± 2.5
48.3 ± 3.8
52.0 ± 0.4
22
23
14
15
Finally the search for potential P2X7R ligands resulted in a
quite general synthesis of scaffolds 2 and 13 and of their
constrained analogues 5. An in vitro anti-proliferative activity
assay was realized for some compounds selected by the National
Cancer Institute (NCI).14 None of the compounds inhibited the
NCI 60 cell lines panel, suggesting an absence of cytotoxicity
(results available in the ‘Supporting information’ section). Newly
synthesized compounds were screened for the antagonistic
properties on the P2X7R at a 10 µM concentration (Table 2).15
The introduction of an enamine group in position 3 of the lactam
ring in compounds 2c proves to be deleterious for the activity on
the receptor (Table 2, entries 1-4). Constrained analogs 5 and 11
also exhibited diminished antagonistic activity against P2X7R
(Table 2, entries 5-15). Finally, replacing the β-(dimethylamino)-
enone function by other amines did not improve the activity
(Table 2, entries 17-21). This new SAR study permitted to
conclude that the conformational space available in the position 3
of pyroglutamide-like P2X7R antagonists is limited.
Consequently, further chemical efforts on different points of
compound 1 might be necessary to obtain molecules targeting
P2X7R with improved biological properties.
In addition to their good stability in some conditions, these
newly synthesized highly functionalized bicyclic heterocycles are
reactive platforms on different positions, providing good tools to
access new chemical entities, and give way to advances in the
field.
4. Experimental Section
Supplementary data (general experimental procedure, full
characterization data including 1H and 13C NMR spectra of
original compounds, supercritical fluid chromatograms for
compounds 2c, 5a and 5c, h-P2X7R screening procedure, cell
proliferation assay procedure and one dose mean graphics for the
NCI-cell proliferation assay) associated with this article can be
found, in the online version, at …
Materials and methods for synthesis and characterizations
Starting materials are commercially available and were used
without further purification (suppliers: Carlo Erba Reagents
S.A.S., Thermo Fisher Scientific Inc., and Sigma-Aldrich Co.).
Intermediates were synthesized according to the methods
described in the literature (references are given in figure S1of the
supporting information). Melting points were measured on a
MPA 100 OptiMelt® apparatus and are uncorrected. Nuclear
magnetic resonance (NMR) spectra were acquired at 400 MHz
for 1H NMR and at 100 MHz for 13C NMR on a Varian 400-MR
spectrometer with tetramethylsilane (TMS) as internal standard,
at room temperature (RT). Chemical shifts (δ) are expressed in
ppm relative to TMS. Splitting patterns are designed: s, singlet;
d, doublet; dd, doublet of doublet; t, triplet; m, multiplet; sym m,
symmetric multiplet; br s, broaden singlet; br t, broaden triplet.
Coupling constants (J) are reported in Hertz (Hz). Thin layer
chromatographies (TLC) were realized on Macherey Nagel silica
gel plates with fluorescent indicator and were visualized under a
UV-lamp at 254 nm and 365 nm. Column chromatographies were
performed with a CombiFlash Rf Companion (Teledyne-Isco
System) using RediSep packed columns. IR spectra were
recorded on a Varian 640-IR FT-IR Spectrometer. Elemental
analyses (C, H, N, S) of new compounds were determined on a
Thermo Electron apparatus by ‘Pôle Chimie Moléculaire-
Welience’, Faculté de Sciences Mirande, Université de
Bourgogne, Dijon, France. LC-MS was accomplished using an
HPLC combined with a Surveyor MSQ (Thermo Electron)
equipped with APCI source. Mass spectra were acquired on a
LTQ Orbitrap XL (Thermo Scientific) apparatus equipped with
electrospray ionization source in positive and negative mode.
Mass spectra samples were first dissolved in DMSO then diluted
in methanol to obtain a final concentration of approximatively 10-
5 M.
In this study, ring closure of L-pyroglutamides 4a-q has been
shown to proceed under the action of BR to provide
unprecedented heterocyclic moieties, namely 3-aminotetrahydro-
1H-pyrrolo[1,2-c]imidazole-1,5(6H)-diones. The efforts made to
propose an unequivocal synthesis of these bicyclic structures 5a-
q showed that the reaction performance was highest for
substrates bearing benzylic amides. Preliminary mechanistic
experiments have provided evidence for a pathway dealing with
an N-acyliminium species. Noteworthy, while direct synthesis of
L-pyroglutamides 2c and 2d (bearing a hydrogen in position 1 of
the lactam ring) is not possible, they can be obtained by
methanolysis of condensed intermediates 5.
Table 2. Inhibition of P2X7R assay at 10-5M concentration
P2X7R inhibition at 10 µM ± SD
Entry
Compd
(%)
1
2
1
2b
2c
100
9.8 ± 21.9
3
53.3 ± 8.7
4
2d
5a
0
5
27.3 ± 13.6
6
5b
5c
0
7
0
8
5d
5e
3.0 ± 0.4
9
0
10
11
12
13
14
15
16
17
18
19
20
5f
0
5k
5l
0
General Method A. Aminolyses of methyl pyroglutamate:
A mixture of (S)-methyl pyroglutamate (1 equiv), amine
derivative (1 equiv) and PTSA (0.05 equiv) was stirred for 24
hours at 120°C under nitrogen atmosphere. The resulting mixture
was purified by direct crystallization in CH3CN to afford pure
compound.
0
0
5m
5q
11
0
2.5 ± 1.7
25.4 ± 1.6
31.4 ± 10.0
16.2 ± 1.6
0
12
General Method B. synthesis of tetrahydro-1H-
pyrrolo[1,2-c]imidazole-1,5(6H)-dione
derivatives
using
13a
13b
13c
13d
Bredereck’s reagent: Pyroglutamide derivative (1 equiv) was
added to Bredereck’s reagent (1-5 equiv). The mixture was
stirred for 2-24 hours at 50-110°C under nitrogen atmosphere.
Bredereck’s reagent was then evaporated under low pressure and
56.5 ± 5.6

5
the resulting mixture was purified by direct crystallization in
diethyl ether or purified by column chromatography on silica gel
to afford pure compound.
silica gel (CH2Cl2/MeOH: 1/0 to 9/1), and washing of the
ACCEPTED MANUSCRIP
T
obtained solid with Et2O resulted in the isolation of pure
compound 2d in 30% yield (1.43 g, 4.96 mmol). mp (Et2O) 169-
170 °C; TLC Rf (CH2Cl2/MeOH: 9/1) 0.2; IR (neat): ν cm-1 3274,
2933, 1662, 1634, 1246, 1109; 1H NMR (DMSO-[d6], 400 MHz):
δ ppm 2.90 (s, 7H, NCHN(CH3)2 and CCH2CH), 3.28 (d, J = 6.2
Hz, 1H, CCH2CH), 3.72 (s, 3H, OCH3), 4.03-4.14 (m, 1H,
CCH2CH), 6.63 (s, 1H, CCHN), 6.88 (d, J = 8.2 Hz, 2H, ArH),
7.22 (s, 1H, NHCH), 7.56 (d, J = 8.2 Hz, 2H, ArH), 9.77 (s, 1H,
NHC); 13C NMR (DMSO-[d6], 100 MHz): δ ppm 29.1 (CH2),
41.1 (2 CH3), 53.7 (CH), 55.0 (CH3), 93.2 (C), 113.7 (2 CH),
120.7 (2 CH), 131.9 (C), 141.4 (C), 155.2 (CH), 171.3 (C), 176.6
(C). Anal. Calcd for C15H19N3O3: C, 62.27; H, 6.62; N, 14.52.
Found: C, 62.63; H, 6.25; N, 14.57%.
General Method C. Substitution of enamine moiety by
other amines: Compound 5a (1 equiv) was added to the
corresponding amine (1 equiv) in toluene. The mixture was
stirred for 2-24 hours at reflux under nitrogen atmosphere. The
resulting mixture was purified by direct crystallization in ethanol
or chloroform, or diethyl ether, or purified by column
chromatography on silica gel to afford pure compound.
(2S)-(4E)-N-(2,4-dichlorobenzyl)-4-
[(dimethylamino)methylidene]-5-oxo-1-(thiophen-2-
yl)pyrrolidine-2-carboxamide (2b): Pyroglutamide derivative 3
(1.00 g, 2.71 mmol) was added to Bredereck’s reagent (2.36 g,
13.54 mmol). The mixture was stirred for 24 hours at 110°C
under nitrogen atmosphere. Bredereck’s reagent was then
evaporated under reduced pressure. After cooling down to RT,
diethyl ether (50 ml) and ethanol (5 ml) were added. The desired
product crystalized and was filtered to afford compound 2b in
45% yield (0.52 g, 1.22 mmol). mp (Et2O) 233-236 °C; TLC Rf
(CH2Cl2/MeOH: 95/5) 0.6; IR (neat): ν cm-1 3259, 1668, 1605,
1527, 1338, 695; 1H NMR (CDCl3, 400 MHz): δ ppm 2.91 (d, J =
16.0 Hz, 1H, CCH2CH), 2.99 (s, 6H, N(CH3)2), 3.42 (dd, J =
14.8, 10.1 Hz, 1H, CCH2CH), 4.34 (d, J = 5.3 Hz, 2H,
NHCH2Ar), 4.64 (dd, J = 10.1, 2.9 Hz, 1H, CCH2CH), 6.29 (d, J
= 3.8 Hz, 1H, ArH), 6.81 (dd, J = 5.3, 3.8 Hz, 1H, ArH), 6.89 (s,
1H, CCHN(CH3)2), 6.92 (d, J = 5.3 Hz, 1H, ArH), 7.27 (d, J =
8.3 Hz, 1H, ArH), 7.38 (dd, J = 8.3, 1.8 Hz, 1H, ArH), 7.61 (d, J
=1.8 Hz, 1H, ArH), 8.88 (br t, J = 5.3 Hz, 1H, NHCH2); 13C
NMR (CDCl3, 100 MHz): δ ppm 28.3 (CH2), 40.3 (CH2), 40.6
(CH3), 41.9 (CH3), 59.0 (CH), 91.8 (C), 108.5 (CH), 117.3 (CH),
123.9 (CH), 127.7 (CH), 129.1 (CH), 130.9 (CH), 132.8 (C),
133.6 (C), 135.7 (C), 141.5 (C), 144.1 (CH), 169.2 (C), 171.6
(C). Anal. Calcd for C19H19Cl2N3O2S: C, 53.78; H, 4.51; N, 9.90.
Found: C, 53.59; H, 4.82; N, 10.04%.
(2S)-N-(2,4-dichlorobenzyl)-5-oxo-1-(thiophen-2-
yl)pyrrolidine-2-carboxamide (3):
A stirred mixture of
pyroglutamide derivative 4a (5,00 g, 17,41 mmol), 2-
iodothiophene (3.66 g, 17.41 mmol), copper iodide (1.66 g, 8.71
mmol), cesium carbonate (11.33 g, 34.82 mmol) and N,N’-
dimethylethylenediamine (1.53 g, 17.41 mmol) was heated at
reflux of dioxane for 5 hours under a nitrogen atmosphere. After
cooling down to RT, copper salts and carbonates were filtered
off. The solvent was then evaporated. The resulting mixture was
solved in CH2Cl2 and washed twice with water. The organic layer
was separated and dried over MgSO4. After evaporation of the
solvent, the resulting solid was washed with diethyl ether to
afford compound 3 in 50% yield (3.14 g, 8.50 mmol). mp (Et2O)
156-159 °C; TLC Rf (CH2Cl2/MeOH: 95/5) 0.6; IR (neat): ν cm-1
3278, 1687, 1647, 1537, 1455, 1228, 823, 675; 1H NMR (CDCl3,
400 MHz): δ ppm 2.24-2.35 (m, 1H, CH2CH2CH), 2.52-2.66 (m,
2H, CH2CH2CH), 2.66-2.80 (m, 1H, CH2CH2CH), 4.39 (dd, J =
15.7, 5.8 Hz, 1H, NHCH2), 4.52 (dd, J = 15.7, 5.8 Hz, 1H,
NHCH2), 4.63 (dd, J = 9.2, 2.5 Hz, 1H, CH2CH2CH), 6.43 (dd, J
= 3.9, 1.4 Hz, 1H, ArH), 6.50 (br t, J = 5.9 Hz, 1H, CONH), 6.80
(dd, J = 5.5, 3.9 Hz, 1H, ArH), 6.91 (dd, J = 5.5, 1.4 Hz, 1H,
ArH), 7.08 (d, J = 8.1 Hz, 1H, ArH), 7.14 (dd, J = 8.3, 2.0 Hz,
1H, ArH), 7.31 (d, J = 2.0 Hz, 1H, ArH); 13C NMR (CDCl3, 100
MHz): δ ppm 23.9 (CH2), 29.9 (CH2), 41.1 (CH2), 63.5 (CH),
112.2 (CH), 119.2 (C), 123.9 (CH), 127.3 (CH), 129.4 (CH),
130.8 (CH), 133.4 (CH), 134.1 (C), 134.2 (C), 138.8 (C), 170.6
(C), 172.6 (C). Anal. Calcd for C16H14Cl2N2O2S: C, 52.04; H,
3.82; N, 7.59; S, 8.68. Found: C, 52.31; H, 4.10; N, 7.70; S,
8.38%.
(2S)-(4E)-N-(2,4-dichlorobenzyl)-4-
[(dimethylamino)methylidene]-5-oxopyrrolidine-2-
carboxamide (2c): Compound 4a (1.00 g, 2.52 mmol) was
stirred in methanol (10 ml) for 8 hours under reflux and under
nitrogen atmosphere. The solvent was evaporated. The resulting
solid was washed with diethylether to afford compound 2c in
72% yield (0.62 g, 1.81 mmol). mp (Et2O) 165-166 °C; TLC Rf
(CH2Cl2/MeOH: 9/1) 0.7; IR (neat): ν cm-1 3257, 2934, 2358,
1683, 1615, 1246, 1032; 1H NMR (CDCl3, 400 MHz): δ ppm 2.98
(s, 6H, NCHN(CH3)2), 3.06 (dd, J = 15.0, 4.6 Hz, 1H, CCH2CH),
3.50 (dd, J = 15.1, 10.4 Hz, 1H, CCH2CH), 4.11-4.17 (m, 1H,
CCH2CH), 4.50 (t, J = 5.5 Hz, 2H, NHCH2), 6.14 (s, 1H,
CCHN), 6.90 (s, 1H, NH), 7.12 (br s, 1H, NHCH2), 7.20 (d, J =
8.1 Hz, 1H, ArH), 7.27 (d, J = 8.5 Hz, 1H, ArH), 7.32 (s, 1H,
ArH); 13C NMR (CDCl3, 100 MHz): δ ppm 30.1 (CH2), 40.8 (2
CH3), 41.7 (CH2), 54.6 (CH), 91.0 (C), 127.2 (CH), 129.3 (CH),
130.7 (CH), 133.9 (C), 134.1 (C), 134.2 (C), 143.9 (CH), 173.5
(C), 176.4 (C). Anal. Calcd for C15H17Cl2N3O2: C, 52.65; H, 5.01;
N, 12.28. Found: C, 52.90; H, 5.06; N, 12.62%.
(2S)-5-oxo-N-(pyridin-4-yl)pyrrolidine-2-carboxamide
(4j): The general procedure
A was followed using 4-
pyridylamine (3.29 g, 34.96 mmol). The desired product
crystallizes in CH3CN as a pink powder, which was filtered to
afford pure 4j in 38% yield (2.73 g, 13.28 mmol). mp (CH3CN)
152-154 °C; TLC Rf (CH2Cl2/MeOH: 9/1) 0.2; IR (neat): ν cm-1
1
3318, 3141 3093, 1668, 1634, 1567, 1525, 1287, 836; H NMR
(DMSO[d6], 400 MHz): δ ppm 1.90-1.98 (m, 1H, CH2CH2CH),
2.07-2.12 (m, 2H, CH2CH2CH), 2.22-2.31 (m, 1H, CH2CH2CH),
4.17 (dd, J = 8.0, 4.4 Hz, 1H, CH2CH2CH), 6.48 (br s, 2H, NH),
6.53 (d, J = 6.6 Hz, 1H, ArH), 7.78 (s, 1H, ArH), 8.00 (d, J = 6.2
Hz, 2H, ArH); 13C NMR (DMSO[d6], 100 MHz): δ ppm 30.0
(CH2), 34.5 (CH2), 60.74 (CH), 113.9 (2 CH), 152.1 (C), 160.8 (2
CH), 180.1 (C), 182.1 (C). Anal. Calcd for C10H11N3O2: C, 58.53;
H, 5.40; N, 20.48. Found: C, 58.83; H, 5.58; N, 20.31%.
(2S)-(4E)-4-[(dimethylamino)methylidene]-N-(4-
methoxyphenyl)-5-oxopyrrolidine-2-carboxamide
(2d):
Pyroglutamide derivative 4k (3.87 g, 16.52 mmol) was added to
Bredereck’s reagent (8.64 g, 49.58 mmol). The mixture was
stirred for 9 hours at 50°C under nitrogen atmosphere.
Bredereck’s reagent was then evaporated under reduced pressure.
1H NMR of the crude reveals that it was containing compound 5k
but not compound 2d. Compound 5k was purified by direct
crystallization in diethyl ether (see page 10 for compound 5k).
The purification of the filtrate by column chromatography on a
(2S)-5-oxo-N-(3,4,5-trimethoxyphenyl)pyrrolidine-2-
carboxamide (4l): The general procedure A was followed using
3,4,5-trimethoxyaniline (1.28 g, 6.99 mmol). The desired product
crystallizes in CH3CN as a white powder, which was filtered to
afford pure 4l in 44% yield (0.91 g, 3.08 mmol). mp (CH3CN)
190-195 °C; TLC Rf (CH2Cl2/MeOH: 9/1) 0.5; IR (neat): ν cm-1

6
Tetrahedron
ACCEPTED MANUSCRIPT
1
3439, 3295, 3189, 3103, 2940, 1670, 1230, 1130; H NMR
(CH), 173.3 (C), 179.1 (C). Anal. Calcd for C18H22Cl2N4O2: C,
54.42; H, 5.58; N, 14.10. Found: C, 54.81; H, 5.56; N, 14.34%.
(CDCl3, 400 MHz): δ ppm 2.23-2.52 (m, 3H, CH2CH2CH), 2.56-
2.67 (m, 1H, CH2CH2CH), 3.77 (s, 3H, OCH3), 3.81 (s, 6H,
OCH3), 4.27 (dd, J = 9.1, 5.0 Hz, 1H, CH2CH2CH), 6.73 (br s,
1H, NHCOCH2), 6.91 (s, 2H, ArH), 8.26 (br s, 1H, NHCOCH);
13C NMR (CDCl3, 100 MHz): δ ppm 26.0 (CH2), 29.3 (CH2),
56.1 (2 CH3), 57.8 (CH), 61.0 (CH3), 97.5 (2 CH), 133.5 (C),
134.7 (C), 153.2 (2 C), 170.1 (C), 179.3 (C). Anal. Calcd for
C14H18N2O5: C, 57.14; H, 6.16; N, 9.52. Found: C, 56.91; H,
6.00; N, 9.51%.
(8S)-(6E)-2-benzyl-3-(dimethylamino)-6-
[(dimethylamino)methylidene]tetrahydro-1H-pyrrolo[1,2-
c]imidazole-1,5(6H)-dione (5b): The general procedure B was
followed using pyroglutamide 4b (1.75 g, 8.02 mmol) and 5
equivalents of Bredereck’s reagent at 110°C for 2 hours. Diethyl
ether was added at the end of the reaction. The desired product
crystallizes as a white powder, which was filtered to afford pure
5b in 47% yield (1.23 g, 3.77 mmol). mp (Et2O) 113-114 °C;
TLC Rf (CH2Cl2/MeOH: 95/5) 0.4; IR (neat): ν cm-1 2941, 2900,
2858, 1689, 1597, 1198, 1026, 706; 1H NMR (CDCl3, 400 MHz):
δ ppm 2.26 (s, 6H, NCHN(CH3)2), 3.01 (s, 6H, CCHN(CH3)2),
3.22 (dd, J = 15.1, 3.4 Hz, 1H, CCH2CH), 3.34 (dd, J = 15.1,
10.8 Hz, 1H, CCH2CH), 4.06 (d, J = 14.5 Hz, 1H, NCH2), 4.16
(dd, J = 10.2, 4.1 Hz, 1H, CCH2CH), 4.87 (d, J = 14.5 Hz, 1H,
NCH2), 5.40 (s, 1H, NCHN(CH3)2), 7.01 (br s, 1H,
CCHN(CH3)2), 7.20-7.32 (m, 5H, ArH); 13C NMR (CDCl3, 100
MHz): δ ppm 27.1 (CH2), 38.0 (2 CH3), 40.1 (2 CH3), 43.6
(CH2), 58.4 (CH), 88.9 (CH), 91.7 (C), 127.7 (CH), 128.4 (2
CH), 128.7 (2 CH), 135.7 (C), 145.7 (CH), 173.0 (C), 179.0 (C).
Anal. Calcd for C18H24N4O2: C, 65.83; H, 7.37; N, 17.06. Found:
C, 65.65; H, 7.55; N, 17.06%.
(2S)-N-[2-(2,4-dichlorophenyl)ethyl]-5-oxopyrrolidine-2-
carboxamide (4n): The general procedure A was followed using
2-(2,4-dichlorophenyl)ethylamine (0.85 g, 6.99 mmol). The
desired product crystallizes in CH3CN as a white powder, which
was filtered to afford pure 4n in 92% yield (1.93 g, 6.43 mmol).
mp (CH3CN) 141-143 °C; TLC Rf (CH2Cl2/MeOH: 95/5) 0.5; IR
(neat): ν cm-1 3235, 3073, 1662, 1249, 809; H NMR (CDCl3,
1
400 MHz): δ ppm 2.06-2.17 (m, 1H, CH2CH2CH), 2.30-2.37 (m,
2H, CH2CH2CH), 2.47-2.59 (m, 1H, CH2CH2CH), 2.96 (t, J = 7.0
Hz, 2H, CH2CH2Ar), 3.55 (dd, J = 6.4, 2.7 Hz, 2H, CH2CH2Ar),
4.12 (ddd, J = 9.1, 5.0, 1.2 Hz, 1H, CH2CH2CH), 6.24 (s, 1H,
NHCH), 6.31 (s, 1H, NHCH2), 7.16 (d, J = 8.3 Hz, 1H, ArH),
7.21 (dd, J = 8.5, 2.1 Hz, 1H, ArH), 7.39 (d, J = 2.1 Hz, 1H,
ArH);); 13C NMR (CDCl3, 100 MHz): δ ppm 26.1 (CH2), 29.1
(CH2), 32.7 (CH2), 39.1 (CH2), 56.8 (CH), 127.4 (CH), 129.5
(CH), 131.7 (CH), 133.3 (C), 134.7 (C), 134.8 (C), 172.0 (C),
179.0 (C). Anal. Calcd for C13H14Cl2N2O2: C, 51.85; H, 4.69; N,
9.30. Found: C, 51.88; H, 4.61; N, 9.70%.
(3S,
8S)-(6E)-3-(dimethylamino)-6-
[(dimethylamino)methylidene]-2-[(1S)-1-
phenylethyl]tetrahydro-1H-pyrrolo[1,2-c]imidazole-1,5(6H)-
dione (5c): The general procedure B was followed using
pyroglutamide 4c (1.90 g, 8.18 mmol) and 5 equivalents of
Bredereck’s reagent at 110°C for 3 hours. Diethyl ether was
added at the end of the reaction. The desired product crystallizes
as a white powder, which was filtered to afford pure 5c in 14%
yield (0.39 g, 1.15 mmol). mp (Et2O) 188-192 °C; TLC Rf
(CH2Cl2/MeOH: 95/5) 0.5; IR (neat): ν cm-1 2972, 2939, 2865,
(2S)-N-(2-norbornyl)-5-oxopyrrolidine-2-carboxamide
(4p): The general procedure
A was followed using 2-
norbornylamine (0.78 g, 6.99 mmol). The desired product
crystallizes in CH3CN as a white powder, which was filtered to
afford pure 4p in 84% yield (1.31 g, 5.87 mmol). mp (CH3CN)
175-178 °C; TLC Rf (CH2Cl2/MeOH: 9/1) 0.5; IR (neat): ν cm-1
3249, 2963, 2873, 2727, 2629, 1695, 1664, 1622, 1562, 1531,
1
1683, 1597, 1315, 1198, 1021; H NMR (CDCl3, 400 MHz): δ
ppm 1.67 (d, J = 7.2 Hz, 3H, CHCH3), 2.20 (s, 6H,
NCHN(CH3)2), 3.00 (s, 6H, CCHN(CH3)2), 3.17-3.31 (m, 2H,
CCH2CH), 4.13 (dd, J = 10.1, 4.2 Hz, 1H, CCH2CH), 5.11 (s,
1H, NCHN(CH3)2), 5.23 (q, J = 7.2 Hz, 1H, CHCH3), 6.96 (br t,
J = 1.6 Hz, 1H, CCHN(CH3)2), 7.24-7.37 (m, 5H, ArH); 13C
NMR (CDCl3, 100 MHz): δ ppm 17.6 (CH3), 26.9 (CH2), 38.2 (2
CH3), 41.8 (2 CH3), 51.5 (CH), 58.6 (CH), 89.1 (CH), 91.8 (C),
127.9 (CH), 128.2 (2 CH), 128.5 (2 CH), 138.7 (C), 145.6 (CH),
172.7 (C), 178.7 (C). Anal. Calcd for C19H26N4O2: C, 66.64; H,
7.65; N, 16.36. Found: C, 66.42; H, 7.82; N, 16.35%.
1
1403, 1299; H NMR (CDCl3, 400 MHz): δ ppm 1.04-1.24 (m,
3H, norbornyl-H), 1.39-1.59 (m, 3H, norbornyl-H), 1.63-1.72 (m,
2H, norbornyl-H), 2.02-2.12 (m, 1H, CH2CH2CH), 2.24-2.40 (m,
5H, CH2CH2CH and norbornyl-H), 3.02-3.08 (m, 1H,
CH2CH2CH), 3.98-4.06 (m, 1H, CH2CH2CH), 7.92 (br s, 1H,
NHCOCH2), NHCOCH (not seen); 13C NMR (CDCl3, 100 MHz):
δ ppm 25.5 (CH2), 26.7 (CH2), 27.8 (CH), 30.8 (CH2), 34.7 (CH),
36.1 (CH2), 37.6 (CH2), 41.0 (CH2), 53.5 (CH), 58.6 (CH), 178.4
(C), 179.4 (C). Anal. Calcd for C12H18N2O2: C, 64.84; H, 8.16; N,
12.60. Found: C, 64.54; H, 8.10; N, 12.57%.
(8S)-(6E)-3-(dimethylamino)-6-
(8S)-(6E)-2-(2,4-dichlorobenzyl)-3-(dimethylamino)-6-
[(dimethylamino)methylidene]tetrahydro-1H-pyrrolo[1,2-
c]imidazole-1,5(6H)-dione (5a): The general procedure B was
followed using pyroglutamide 4a (4.04 g, 14.07 mmol) and 5
equivalents of Bredereck’s reagent at 110°C for 24 hours. Diethyl
ether was added at the end of the reaction. The desired product
crystallizes as a white powder, which was filtered to afford pure
5a in 75% yield (4.19 g, 10.55 mmol). mp (Et2O) 175-179 °C;
TLC Rf (CH2Cl2/MeOH: 95/5) 0.8; IR (neat): ν cm-1 2948, 2870,
[(dimethylamino)methylidene]-2-(2-
methoxybenzyl)tetrahydro-1H-pyrrolo[1,2-c]imidazole-
1,5(6H)-dione (5d): The general procedure B was followed
using pyroglutamide 4d (2.50 g, 10.07 mmol) and 5 equivalents
of Bredereck’s reagent at 110°C for 17 hours. Diethyl ether and
ethanol were added at the end of the reaction. The desired
product crystallizes as a beige powder, which was filtered to
afford pure 5d in 29% yield (1.05 g, 2.92 mmol). mp (Et2O) 213-
217 °C; TLC Rf (CH2Cl2/MeOH: 95/5) 0.3; IR (neat): ν cm-1
2947, 1702, 1685, 1618, 1242, 1204, 1035; 1H NMR (CDCl3, 400
MHz): δ ppm 2.28 (s, 6H, NCHN(CH3)2), 3.00 (s, 6H,
CCHN(CH3)2), 3.19 (dd, J = 15.3, 4.6 Hz, 1H, CCH2CH), 3.32
(dd, J = 15.3, 10.5 Hz, 1H, CCH2CH), 3.80 (s, 3H, OCH3), 4.14
(dd, J = 10.5, 4.3 Hz, 1H, CCH2CH), 4.24 (d, J = 14.9 Hz, 1H,
NCH2), 4.75 (d, J = 14.9 Hz, 1H, NCH2), 5.53 (s, 1H,
NCHN(CH3)2), 6.83 (d, J = 8.7 Hz, 1H, ArH), 6.88 (d, J = 7.5
Hz, 1H, ArH), 7.00 (br s, 1H, CCHN(CH3)2), 7.15 (d, J = 7.5 Hz,
1H, ArH), 7.22 (m, 1H, ArH); 13C NMR (CDCl3, 100 MHz): δ
ppm 27.2 (CH2), 38.1 (2 CH3), 39.0 (CH2), 41.8 (2 CH3), 55.2
1
1707, 1682, 1616, 1204, 1033; H NMR (CDCl3, 400 MHz): δ
ppm 2.14 (s, 6H, NCHN(CH3)2), 2.99 (s, 6H, CCHN(CH3)2), 3.02
(m, 1H, CCH2CH), 3.30 (m, 1H, CCH2CH), 4.18 (dd, J = 10.5,
3.5 Hz, 1H, CCH2CH), 4.19 (d, J = 15.0 Hz, 1H, NCH2), 4.68 (d,
J = 15.0 Hz, 1H, NCH2), 5.24 (s, 1H, NCHN(CH3)2), 6.91 (br t, J
= 1.7 Hz, 1H, CCHN(CH3)2), 7.22 (d, J = 8.4 Hz, 1H, ArH), 7.44
(dd, J = 8.4, 1.9 Hz, 1H, ArH), 7.63 (d, J = 1.9 Hz, 1H, ArH); 13
C
NMR (CDCl3, 100 MHz): δ ppm 27.1 (CH2), 38.0 (2 CH3), 40.9
(CH2), 41.8 (2 CH3), 58.2 (CH), 89.1 (CH), 91.4 (C), 127.4 (CH),
129.6 (CH), 130.9 (CH), 132.1 (C), 134.2 (C), 134.5 (C), 145.9

7
(CH3), 58.2 (CH), 89.3 (CH), 92.0 (C), 110.4 (CH), 120.5 (CH),
124.1 (C), 128.8 (CH), 129.6 (CH), 145.5 (CH), 157.7 (C), 173.1
(C), 179.1 (C). Anal. Calcd for C19H26N4O3: C, 63.67; H, 7.31; N,
15.63. Found: C, 63.58; H, 7.19; N, 15.37%.
27.1 (2 CH3), 38.2 (2 CH3), 55.4 (CH3), 58.8 (CH), 91.2 (CH),
ACCEPTED MANUSCRIPT
91.5 (C), 114.3 (2 CH), 125.1 (2 CH), 129.0 (C), 146.0 (CH),
157.8 (C), 172.6 (C), 179.3 (C). Anal. Calcd for C18H24N4O3: C,
62.77; H, 7.02; N, 16.27. Found: C, 62.79; H, 7.02; N, 16.53%.
(8S)-(6E)-3-(dimethylamino)-2-(2,4-dimethoxybenzyl)-6-
[(dimethylamino)methylidene]tetrahydro-1H-pyrrolo[1,2-
c]imidazole-1,5(6H)-dione (5e). The general procedure B was
followed using pyroglutamide 4e (0.38 g, 1.37 mmol) and 5
equivalents of Bredereck’s reagent at 110°C for 3 hours. Diethyl
ether was added at the end of the reaction. The desired product
crystallizes as a white powder, which was filtered to afford pure
5e in 52% yield (0.28 g, 0.71 mmol). mp (Et2O) 115-116 °C;
TLC Rf (CH2Cl2/MeOH: 95/5) 0.4; IR (neat): ν cm-1 2942, 2924,
2835, 1706, 1632, 1609, 1585, 1265, 1029, 838; 1H NMR
(CDCl3, 400 MHz): δ ppm 2.27 (s, 6H, NCHN(CH3)2), 3.00 (s,
6H, CCHN(CH3)2), 3.17 (dd, J = 15.4, 2.8 Hz, 1H, CCH2CH),
3.30 (dd, J = 14.3, 10.7 Hz, 1H, CCH2CH), 3.77 (s, 3H, OCH3),
3.78 (s, 3H, OCH3), 4.12 (dd, J = 9.8, 4.0 Hz, 1H, CCH2CH),
4.14 (d, J = 14.2 Hz, 1H, NCH2), 4.70 (d, J = 14.4 Hz, 1H,
NCH2), 5.49 (s, 1H, NCHN), 6.40 (m, 1H, ArH), 6.40 (m, 1H,
CCHN), 6.99 (t, J = 1.7 Hz, 1H, ArH), 7.10 (d, J = 8.9 Hz, 1H,
ArH); 13C NMR (CDCl3, 100 MHz): δ ppm 27.2 (CH2), 38.1 (2
CH3), 38.6 (CH2), 41.7 (2 CH3), 55.3 (CH3), 55.4 (CH3), 58.3
(CH), 89.3 (CH), 92.2 (C), 98.6 (CH), 104.1 (CH), 116.7 (C),
130.7 (CH), 145.4 (CH), 158.5 (C), 160.5 (C), 172.9 (C), 179.1
(C). Anal. Calcd for C20H28N4O4: C, 61.84; H, 7.27; N, 14.42.
Found: C, 61.63; H, 7.45; N, 14.51%.
(8S)-(6E)-3-(dimethylamino)-6-
[(dimethylamino)methylidene]-2-(3,4,5-
trimethoxyphenyl)tetrahydro-1H-pyrrolo[1,2-c]imidazole-
1,5(6H)-dione (5l): The general procedure B was followed using
pyroglutamide 4l (0.80 g, 2.72 mmol) and 5 equivalents of
Bredereck’s reagent at 110°C for 4 hours. Diethyl ether was
added at the end of the reaction. The desired product crystallizes
as a dark blue powder, which was filtered to afford pure 5l in
55% yield (0.61 g, 1.50 mmol). mp (Et2O) 143-147 °C; TLC Rf
(CH2Cl2/MeOH: 95/5) 0.4; IR (neat): ν cm-1 2941, 1711, 1682,
1591, 1507, 1126, 1034; 1H NMR (DMSO[d6], 400 MHz): δ ppm
2.22 (s, 6H, NCHN(CH3)2), 3.01 (s, 6H, CCHN(CH3)2), 3.04-
3.11 (m, 1H, CCH2CH), 3.34-3.40 (m, 1H, CCH2CH), 3.65 (s,
3H, OCH3), 3.75 (s, 6H, OCH3), 4.34 (dd, J = 10.4, 3.4 Hz, 1H,
CCH2CH), 6.01 (s, 1H, NCHN(CH3)2), 6.87 (s, 2H, ArH), 6.98
(br s, 1H, CCHN(CH3)2); 13C NMR (DMSO[d6], 100 MHz): δ
ppm 26.3 (CH2), 37.7 (2 CH3), 40.1 (2 CH3), 55.9 (2 CH3), 57.9
(CH), 59.9 (CH3), 90.1 (CH), 90.5 (C), 101.2 (2 CH), 132.2 (C),
135.3 (C), 145.5 (CH), 152.6 (2 C), 172.0 (C), 177.7 (C). Anal.
Calcd for C20H28N4O5: C, 59.39; H, 6.98; N, 13.85. Found: C,
59.53; H, 7.22; N, 13.72%.
(8S)-(6E)-3-(dimethylamino)-6-
[(dimethylamino)methylidene]-2-(2-phenylethyl)tetrahydro-
1H-pyrrolo[1,2-c]imidazole-1,5(6H)-dione (5m): The general
procedure B was followed using pyroglutamide 4m (0.85 g, 3.66
mmol) and 5 equivalent of Bredereck’s reagent at 110°C for 3
hours. Diethyl ether and ethanol were added at the end of the
reaction. The desired product crystallizes as a white powder,
which was filtered to afford pure 5m in 7% yield (0.09 g, 0.26
mmol). mp (Et2O) 139-142 °C; TLC Rf (CH2Cl2/MeOH: 95/5)
(8S)-(6E)-2-(2-chlorobenzyl)-3-(dimethylamino)-6-
[(dimethylamino)methylidene]tetrahydro-1H-pyrrolo[1,2-
c]imidazole-1,5(6H)-dione (5f): The general procedure B was
followed using pyroglutamide 4f (2.00 g, 7.91 mmol) and 5
equivalents of Bredereck’s reagent at 110°C for 3 hours. Diethyl
ether was added at the end of the reaction. The desired product
crystallizes as a white powder, which was filtered to afford pure
5f in 47% yield (1.34 g, 3.71 mmol). mp (Et2O) 172-173 °C;
TLC Rf (CH2Cl2/MeOH: 9/1) 0.4; IR (neat): ν cm-1 2937, 2259,
1709, 1681, 1619, 1203, 1036, 757; 1H NMR (CDCl3, 400 MHz):
δ ppm 2.26 (s, 6H, NCHN(CH3)2), 3.02 (s, 6H, NCHN(CH3)2),
3.20 (sym m, J = 16.2 Hz, 1H, CCH2CH), 3.30 (dd, J = 16.2,
10.6 Hz, 1H, CCH2CH), 4.17 (dd, J = 10.6, 5.0 Hz, 1H,
CCH2CH), 4.30 (d, J = 15.1 Hz, 1H, NCH2), 4.91 (d, J = 15.0 Hz,
1H, NCH2), 5.44 (s, 1H, NCHN), 7.03 (s, 1H, CCHNH), 7.19-
7.22 (m, 3H, ArH), 7.31-7.36 (m, 1H, ArH); 13C NMR (CDCl3,
100 MHz): δ ppm 27.2 (CH2), 38.0 (2 CH3), 41.4 (2 CH3), 41.8
(CH2), 58.2 (CH), 89.1 (CH), 91.7 (C), 127.0 (CH), 129.0 (CH),
129.8 (CH), 129.9 (CH), 133.4 (C), 133.7 (C), 145.8 (CH), 173.2
(C), 179.1 (C). Anal. Calcd for C18H23ClN4O2: C, 59.58; H, 6.39;
N, 15.44. Found: C, 59.38; H, 6.39; N, 15.41%.
0.5; IR (neat): ν cm-1 2942, 1678, 1612, 1313, 1203, 1033; H
1
NMR (DMSO[d6], 400 MHz): δ ppm 2.12 (s, 6H, NCHN(CH3)2),
2.65-2.87 (m, 3H, CCH2CH and NCH2CH2), 2.98 (s, 6H,
CCHN(CH3)2), 3.10-3.24 (m, 2H, CCH2CH and NCH2CH2),
3.56-3.64 (m, 1H, NCH2CH2), 4.03 (dd, J = 10.5, 3.7 Hz, 1H,
CCH2CH), 5.38 (s, 1H, NCHN(CH3)2), 6.92 (br t, J = 1.6 Hz, 1H,
CCHN(CH3)2), 7.10-7.26 (m, 5H, ArH); 13C NMR (DMSO[d6],
100 MHz): δ ppm 26.7 (CH2), 33.3 (CH2), 38.2 (2 CH3), 41.3
(CH2), 41.8 (2 CH3), 57.6 (CH), 89.2 (CH), 91.4 (C), 126.7 (CH),
128.7 (2 CH), 129.0 (2 CH), 139.1 (C), 147.7 (CH), 172.9 (C),
178.6 (C). Anal. Calcd for C19H26N4O2: C, 66.64; H, 7.65; N,
16.36. Found: C, 66.93; H, 7.66; N, 15.96%.
(8S)-(6E)-3-(dimethylamino)-6-
[(dimethylamino)methylidene]-2-hexyltetrahydro-1H-
(8S)-(6E)-3-(dimethylamino)-6-
pyrrolo[1,2-c]imidazole-1,5(6H)-dione
(5q)
and
3-
[(dimethylamino)methylidene]-2-(4-
(dimethylamino)-2-hexyl-tetrahydro-1H-pyrrolo[1,2-
methoxyphenyl)tetrahydro-1H-pyrrolo[1,2-c]imidazole-
1,5(6H)-dione (5k): The general procedure B was followed
using pyroglutamide 4k (3.87 g, 16.52 mmol) and 5 equivalents
of Bredereck’s reagent at 50°C for 9 hours. Diethyl ether was
added at the end of the reaction. The desired product crystallizes
as a white powder, which was filtered to afford pure 5k in 6%
yield (0.34 g, 0.99 mmol). mp (Et2O) 148-149 °C; TLC Rf
(CH2Cl2/MeOH: 95/5) 0.6; IR (neat): ν cm-1 1679, 1620, 1511,
c]imidazole-1,5(6H)-dione (11): The general procedure B was
followed using pyroglutamide 4q (2.00 g, 9.42 mmol) and one
equivalent of Bredereck’s reagent at 50°C for 6 hours. The
1
reaction was followed by H NMR. We observed compound 11
forming after 2 hours, then compound 5q was produced while
compound 11 was reacting. The resulting mixture was purified
on a silica gel by column chromatography (EtOAc/n-heptane: 0/1
to 1/0) to afford a mixture of compound 5q and compound 11
(proportion: 1/3). A second chromatography (Et2O/MeOH: 0/1 to
9/1) resulted in a first yellow oil, compound 5q (yield: 14%, 0.43
g, 1.31 mmol), and in a second yellow oil, compound 11 (yield:
20%, 0.50 g, 1.88 mmol) in two separated fractions of the eluent.
1
1248, 1206, 1036; H NMR (CDCl3, 400 MHz): δ ppm 2.32 (s,
6H, NCHN(CH3)2), 3.28-3.34 (m, 2H, CCH2CH), 3.02 (s, 6H,
CCHN(CH3)2), 3.77 (s, 3H, OCH3), 4.27 (dd, J = 10.2, 3.8 Hz,
1H, CCH2CH), 5.99 (s, 1H, CCHN(CH3)2), 6.88 (dd, J = 12.8,
9.0 Hz, 1H, ArH), 7.08 (t, J = 1.7 Hz, 2H, ArH), 7.39 (d, J = 8.9
Hz, 2H, ArH); 13C NMR (CDCl3, 100 MHz): δ ppm 27.1 (CH2),
(6E)-3-(dimethylamino)-6-[(dimethylamino)methylidene]-
2-hexyltetrahydro-1H-pyrrolo[1,2-c]imidazole-1,5(6H)-dione

8
Tetrahedron
ACCEPTED MANUSCRIPT
(5q). TLC Rf (CH2Cl2/MeOH: 9/1) 0.7; IR (neat): ν cm-1 2929,
NHCH2), 4.74 (d, J = 15.4 Hz, 1H, NHCH2), 5.66 (d, J = 1.2
2860, 1850, 1606, 1308, 1204, 1031; 1H NMR (CDCl3, 400
MHz): δ ppm 0.86 (br s, 3H, CH2CH3), 1.18-1.36 (m, 6H,
CH2CH2CH2CH3), 1.40-1.59 (m, 2H, NCH2CH2), 2.27 (s, 6H,
NCHN(CH3)2), 2.81 (s, 1H, NCH2), 3.00 (s, 6H, CCHN(CH3)2),
3.19 (sym m, J = 14.9, 4.1 Hz, 1H, CCH2CH), 3.29 (sym m, J =
15.1, 10.4 Hz, 1H, CCH2CH), 3.51 (dq, J = 9.3, 6.3 Hz, 1H,
NCH2), 4.08 (dd, J = 10.2, 3.8 Hz, 1H, CCH2CH), 5.56 (s, 1H,
NCHN), 7.05 (s, 1H, CCHN); 13C NMR (CDCl3, 100 MHz): δ
ppm 14.0 (CH3), 22.5 (CH2), 26.5 (CH2), 27.0 (CH2), 27.2 (CH2),
31.4 (CH2), 38.1 (2 CH3), 40.3 (2 CH3), 58.0 (2 CH), 89.4 (CH),
91.8 (C), 145.7 (CH), 173 (C), 179.5 (C). Anal. Calcd for
C17H30N4O2: C, 63.32; H, 9.38; N, 17.38. Found: C, 63.07; H,
9.42; N, 17.42%.
Hz, 1H, CCHN(CH3)2), 7.32 (d, J = 8.7 Hz, 1H, ArH), 7.44
(dd, J = 8.7, 2.0 Hz, 1H, ArH), 7.51 (br s, 1H, NHCH), 7.64
(d, J = 2.0 Hz, 1H, ArH), 10.38 (br s, 1H, NHCH2); 13C NMR
(DMSO[d6], 100 MHz): δ ppm 20.0 (CH2), 24.9 (CH2), 37.8
(2 CH3), 57.6 (CH2), 85.5 (CH), 105.0 (CH), 127.5 (CH),
128.9 (CH), 131.3 (CH), 132.5 (C), 132.9 (C), 133.4 (C),
151.4 (C), 164.9 (C), 169.7 (C). Anal. Calcd for
C16H19Cl2N3O2: C, 53.94; H, 5.38; N, 11.79. Found: C, 53.95; H,
5.38; N, 11.76%.
(2S)-N-(2,4-dichlorobenzyl)-1-formyl-5-oxopyrrolidine-2-
carboxamide (12): Compound 4a (1.00 g, 3.48 mmol) and
Vielsmeier’s reagent (0.45 g, 3.48 mmol) were introduced in
DMF (5 mL). The mixture was stirred at RT for 48 hours. DMF
was evaporated without heating under low pressure. Chloroform
was added (30 mL) and the desired product crystalizes as a
salmon colored solid, which was filtered to afford pure 12 in 5%
yield (0.05 g, 0.17 mmol). mp (CHCl3) 155-160 °C; TLC Rf
(CH2Cl2/MeOH: 95/5) 0.6; IR (neat): ν cm-1 3273, 3089, 1742,
(8S)-3-(dimethylamino)-2-hexyl-tetrahydro-1H-
pyrrolo[1,2-c]imidazole-1,5(6H)-dione
(11):
TLC
Rf
(CH2Cl2/MeOH: 9/1) 0.6; IR (neat): ν cm-1 2928, 2858, 1699,
1
1235, 1033; H NMR (CDCl3, 400 MHz): δ ppm 0.79-0.91 (m,
3H, CH2CH3), 1.14-1.35 (m, 6H, CH2(CH2)3CH3), 1.38-1.60 (m,
2H, NCH2CH2), 1.83-2.00 (m, 1H, CH2CH2CH), 2.14 (s, 6H,
N(CH3)2), 2.19-2.28 (m, 1H, CH2CH2CH), 2.31-2.42 (m, 1H,
CH2CH2CH), 2.62-2.74 (m, 1H, CH2CH2CH), 2.91-3.00 (m, 1H,
NCH2), 3.34-3.38 (m, 1H, NCH2), 4.27 (t, J = 8.4 Hz, 1H,
CCH2CH), 5.50 (s, 1H, NCHN); 13C NMR (CDCl3, 100 MHz): δ
ppm 14.0 (CH3), 22.5 (CH2), 25.3 (CH2), 26.6 (CH2), 27.2 (CH2),
31.4 (CH2), 32.8 (CH2), 37.9 (2 CH3), 40.1 (CH2), 60.5 (CH),
86.9 (CH), 171.4 (C), 180.27 (C). Anal. Calcd for C14H25N3O2: C,
62.89; H, 9.42; N, 15.72. Found: C, 62.50; H, 9.71; N, 15.53%.
1
1695, 1651, 1557, 1237; H NMR (CDCl3, 400 MHz): δ ppm
2.19-2.27 (m, 1H, CH2CH2CH), 2.45-2.59 (m, 2H, CH2CH2CH),
2.82-2.93 (m, 1H, CH2CH2CH), 4.49 (d, J = 2.5 Hz, 1H,
NHCH2), 4.51 (d, J = 2.5 Hz, 1H, NHCH2), 4.61 (dd, J = 9.2, 1.2
Hz, 1H, CCH2CH), 6.66 (br s, 1H, NHCH2), 7.24 (dd, J = 8.2, 2.2
Hz, 1H, ArH), 7.29 (d, J = 8.2 Hz, 1H, ArH), 7.40 (d, J = 2.2 Hz,
1H, ArH), 9.10 (s, 1H, CHO); 13C NMR (CDCl3, 100 MHz): δ
ppm 22.0 (CH2), 31.1 (CH2), 41.3 (CH2), 56.2 (CH), 127.5 (CH),
129.5 (CH), 130.9 (CH), 133.4 (C), 134.2 (C), 134.3 (C), 160.7
(CH), 168.9 (C), 176.2 (C). Anal. Calcd for C13H12Cl2N2O3: C,
49.44; H, 3.84; N, 8.89. Found: C, 49.44; H, 3.78; N, 8.59%.
N-(2,4-dichlorobenzyl)-6-oxopiperidine-2-carboxamide
(7): A mixture of (S)-6-oxopiperidine-2-carboxylic acid (1.00 g,
6.99 mmol), 2,4-dichlorobenzylamine (1.48 g, 8.38 mmol) and 3-
{[(ethylimino)methylene]amino}-N,N-dimethylpropan-1-
aminium chloride (EDCI) (1.61 g, 8.38 mmol), in CH2Cl2 was
stirred at r.t. for 24 h. Solvents were evaporated and the mixture
was solved in EtOH (40 mL), then Et2O was added (60 mL). The
desired product crystalized overnight. It was filtered to give a
white solid in 62 % yield (1.30 g, 4.32 mmol). mp (EtOH/Et2O)
142-144 °C; TLC Rf (CH2Cl2/MeOH: 96/4) 0.3; 1H NMR
(2S)-(4E)-4-[(4’-aminobenzo-15-crown-5)methylidene]-N-
(2,4-dichlorobenzyl)-5-oxopyrrolidine-2-carboxamide (13a):
The general procedure C was followed using 4’-aminobenzo-15-
crown-5 (0.08 g, 0.29 mmol) and 3 hours of reaction. Ethanol
was added at the end of the reaction. The desired product
crystalizes as a black powder, which was filtered to afford pure
13a in 53% yield (0.09 g, 0.15 mmol). mp (EtOH) 109-110 °C;
TLC Rf (CH2Cl2/MeOH: 9/1) 0.2; IR (neat): ν cm-1 3295, 2926,
2361, 1644, 1613, 1222, 1126; 1H NMR (DMSO[d6], 400 MHz):
δ ppm 2.56 (ddd, J = 17.0, 4.4, 2.1 Hz, 1H, CCH2CH), 2.94 (dd, J
= 6.7, 2.2 Hz, 1H, CCH2CH), 3.51-3.60 (m, 8H, OCH2CH2O),
3.66-3.74 (m, 4H, OCH2CH2O), 3.90-3.93 (m, 2H, OCH2CH2O),
3.98-4.01 (m, 2H, OCH2CH2O), 4.08 (dd, J = 9.5, 4.2 Hz, 1H,
CCH2CH), 4.24 (dd, J = 15.4, 5.7 Hz, 1H, NHCH2), 4.31 (dd, J =
15.4, 5.7 Hz, 1H, NHCH2), 6.47 (dd, J = 8.8, 2.5 Hz, 1H, ArH),
6.60 (d, J = 2.7 Hz, 1H, ArH), 6.79 (d, J = 8.7 Hz, 1H, ArH),
7.20 (dt, J = 12.7, 1.9 Hz, 1H, NHCH2), 7.30 (d, J = 8.4 Hz, 1H,
ArH), 7.35 (dd, J = 8.4, 2.3 Hz, 1H, ArH), 7.52 (br s, 1H,
CONHCH), 7.57 (d, J = 2.3 Hz, 1H, ArH), 8.24 (d, J = 12.6 Hz,
1H, CHNHAr), 8.48 (br t, J = 5.8 Hz, 1H, CHNHAr); 13C NMR
(DMSO[d6], 100 MHz): δ ppm 29.2 (CH2), 40.6 (CH2), 53.8
(CH), 68.5 (CH2), 69.2 (CH2), 69.5 (CH2), 69.8 (CH2), 70.1
(CH2), 70.4 (CH2), 70.6 (CH2), 70.7 (CH2), 101.5 (CH), 101.6
(C), 106.2 (CH), 116.5 (CH), 127.7 (CH), 129.0 (CH), 129.9
(CH), 130.6 (C), 132.7 (C), 133.4 (C), 135.8 (CH), 137.6 (C),
143.1 (C), 150.1 (C), 172.8 (C), 173.6 (C). Anal. Calcd for
C27H31Cl2N3O7: C, 55.87; H, 5.38; N, 7.24. Found: C, 55.96; H,
5.58; N, 7.57%.
(DMSO[d6], 400 MHz):
δ ppm 1.56-1.81 (m, 3H,
CH2CH2CH2CH), 1.86-1.96 (m, 1H, CH2CH2CH2CH), 2.15 (t,
J = 6.8 Hz, 2H, CH2CH2CH2CH), 3.95 (dd, J = 5.9, 3.0 Hz,
1H, CH2CH2CH2CH), 4.28 (dd, J = 15.8, 5.6 Hz, 1H,
NHCH2), 4.37 (dd, J = 15.8, 5.6 Hz, 1H, NHCH2), 7.38 (d, J
= 8.3 Hz, 1H, ArH), 7.42 (dd, J = 8.3, 2.1 Hz, 1H, ArH), 7.57
(br d, J = 2.5 Hz, 1H, NHCH), 7.60 (d, J = 2.1 Hz, 1H, ArH),
8.52 (br t, J = 5.8 Hz, 1H, NHCH2); 13C NMR (DMSO[d6],
100 MHz): δ ppm 18.9 (CH2), 26.4 (CH2), 31.7 (CH2), 40.2
(CH2), 55.4 (CH), 127.7 (CH), 128.9 (CH), 130.6 (CH), 132.6
(C), 133.3 (C), 135.9 (C), 170.9 (CH), 172.7 (C). Anal. Calcd
for C13H14Cl2N2O2: C, 51.85; H, 4.69; N, 9.30. Found: C, 51.82;
H, 4.70; N, 9.29%.
(6E)-2-(2,4-dichlorobenzyl)-3-(dimethylamino)-6-
[(dimethylamino)methylene]hexahydroimidazo[1,5-
a]pyridine-1,5-dione (10): Compound 7 (1.00 g, 3.32 mmol) and
Bredereck’s reagent (2.89 g, 16.60 mmol) were heated at 110°C
for 3 hours. The solvent was evaporated and diethylether was
added to the crude with a few drops of ethanol. The mixture was
purified by column chromatography (CH2Cl2/MeOH: 1/0 to 9/1)
to give a white solid in 16% yield (0.19 g, 0.53 mmol). mp
(MeOH) 119-120 °C; TLC Rf (CH2Cl2/MeOH: 96/4) 0.8; 1H
NMR (DMSO[d6], 400 MHz) δ (ppm) 2.14-2.25 (m, 8H,
CCH2CH2CH and N(CH3)2), 2.37-2.42 (m, 1H,
CCH2CH2CH), 2.63-2.71 (m, 1H, CCH2CH2CH), 4.11 (dd, J
= 12.1, 3.0 Hz, 1H, CCH2CH2CH), 4.22 (d, J = 15.4 Hz, 1H,
(2S)-(4E)-N-(2,4-dichlorobenzyl)-5-oxo-4-{[(3,4,5-
trimethoxybenzyl)amino]methylidene}pyrrolidine-2-
carboxamide (13b): The general procedure C was followed
using 3,4,5-trimethoxybenzylamine (1.00 g, 5.07 mmol) and 24
hours of reaction. The resulting mixture was purified on a silica
column (EtOAc/MeOH: 9/1) and washed with diethyl ether to

9
afford pure 13b as a white powder in 82% yield (2.06 g, 4.16
mmol). mp (Et2O) 125-126 °C; TLC Rf (CH2Cl2/MeOH: 95/5)
(CH2Cl2/MeOH: 9/1) 0.2; IR (neat): ν cm-1 3293, 2932, 2362,
1
ACCEPTED MANUSCRIPT
1699, 1666, 1615; H NMR (CDCl3, 400 MHz): δ ppm 2.31 (s,
3H, NCH3), 2.35-2.43 (m, 4H, N(CH2)2), 2.96 (ddd, J = 15.1, 4.8,
1.6 Hz, 1H, CCH2CH), 3.32-3.43 (m, 5H, N(CH2)2 and
CCH2CH), 4.18 (ddd, J = 10.2, 4.7, 1.7 Hz, 1H, CCH2CH), 4.47
(dd, J = 15.3, 6.1 Hz, 1H, NHCH2), 4.52 (dd, J = 15.3, 6.1 Hz,
1H, NHCH2), 6.29 (s, 1H, NCH), 6.88 (br t, J = 2.0 Hz, 1H,
NHCH), 7.15 (br t, J = 5.8 Hz, 1H, NHCH2), 7.19 (dd, J = 8.3,
2.1 Hz, 1H, ArH), 7.30 (d, J = 8.3 Hz, 1H, ArH), 7.37 (d, J = 2.1
Hz, 1H, ArH); 13C NMR (CDCl3, 100 MHz): δ ppm 30.5 (CH2),
40.8 (CH2), 46.1 (CH3), 49.7 (2 CH2), 54.5 (CH), 54.8 (2 CH2),
91.6 (C), 127.3 (CH), 129.3 (CH), 130.8 (CH), 133.9 (C), 134.0
(C), 134.2 (C), 142.1 (CH), 173.3 (C), 176.3 (C). Anal. Calcd for
C18H22Cl2N4O2: C, 54.42; H, 5.58; N, 14.10. Found: C, 54.06; H,
5.68; N, 13.96%.
1
0.1; IR (neat): ν cm-1 3278, 1687, 1647, 1537, 1455; H NMR
(CD3OD, 400 MHz): δ ppm 2.61 (ddd, J = 15.7, 4.8, 2.0 Hz, 1H,
CCH2CH), 3.04 (ddd, J = 15.7, 10.3, 2.0 Hz, 1H, CCH2CH), 3.71
(s, 3H, OCH3), 3.80 (s, 6H, 2 OCH3), 4.20 (dd, J = 9.8, 4.5 Hz,
1H, CCH2CH), 4.24 (s, 2H, NHCH2), 4.43 (s, 2H, NHCH2), 4.57
(m, 1H, CH2CH), 6.58 (s, 2H, 2 ArH), 7.06 (br s, 1H, NHCH2),
7.24 (dd, J = 8.3, 2.0 Hz, 1H, ArH), 7.29 (d, J = 8.3 Hz, 1H,
ArH), 7.42 (d, J = 2.0 Hz, 1H, ArH), (NH lactam and NH amide
not seen); 13C NMR (CD3OD, 100 MHz): δ ppm 30.1 (CH2), 41.5
(CH2), 52.5 (CH), 55.9 (CH2), 56.5 (2 CH3), 61.1 (CH3), 92.2
(C), 105.5 (2 CH), 128.3 (CH), 130.1 (CH), 131.2 (CH), 134.7
(C), 135.0 (C), 135.8 (C), 137.4 (C), 138.1 (C), 142.3 (2 C),
154.7 (CH), 176.0 (C), 177.4 (C). Anal. Calcd for
C23H25Cl2N3O5: C, 55.88; H, 5.10; N, 8.50. Found: C, 55.50; H,
5.01; N, 8.25%.
(2S)-(4E)-N-(2,4-dichlorobenzyl)-4-(hydroxymethylidene)-
5-oxopyrrolidine-2-carboxamide (14): Compound 5a (2.00 g,
5.03 mmol) was added to a solution of MeSO3H (0.024 g, 0.25
mmol) in water (20 mL, pH ≈ 4). The mixture was stirred under
reflux for 24 hours. The desired product crystalizes in water at
RT as pink crystals, which were filtered and washed with CH2Cl2
to afford pure 14 in 40% yield (0.64 g, 2.03 mmol). mp (CH2Cl2)
167-173 °C; TLC Rf (CH2Cl2/MeOH: 95/5) 0.5; IR (neat): ν cm-1
3317, 3158, 2925, 1697, 1659, 1627, 1548, 1346; 1H NMR
(DMSO[d6], 400 MHz): δ ppm 2.85 (dd, J = 15.3, 4.8 Hz, 1H,
CCH2CH), 3.24-3.30 (m, 1H, CCH2CH), 4.00 (dd, J = 9.4, 4.6
Hz, 1H, CCH2CH), 4.25-4.37 (m, 2H, NCH2), 6.66 (br t, J = 1.8
Hz, 1H, CCHOH), 7.21 (br s, 1H, NH), 7.35 (d, J = 8.3 Hz, 1H,
ArH), 7.41 (dd, J = 8.3, 1.9 Hz, 1H, ArH), 7.60 (d, J = 1.9 Hz,
1H, ArH), 8.41 (br t, J = 6.4 Hz, 1H, NH), (OH not seen); 13C
NMR (DMSO[d6], 100 MHz): δ ppm 29.1 (CH2), 41.2 (CH2),
53.3 (CH), 93.1 (CH), 127.1 (CH), 128.4 (CH), 130.0 (CH),
132.8 (C), 135.4 (2 C), 141.5 (CH), 173.2 (C), 173.7 (C); HRMS
(ESI): negative mode [M - H]- (C13H12O3N2Cl2 - H): calcd.
313.01412, found 313.01378 g/mol.
(2S)-(4E)-N-(2,4-dichlorobenzyl)-4-{[(2,4-
dichlorobenzyl)amino]methylidene}-5-oxopyrrolidine-2-
carboxamide (13c): The general procedure C was followed
using 2,4-dichlorobenzylamine (0.09 g, 0.51 mmol) and 2 hours
of reaction. Chloroform was added at the end of the reaction. The
desired product crystalizes as a white powder, which was filtered
to afford pure 13c in 65% yield (0.16 g, 0.33 mmol). mp (CHCl3)
156-161 °C; TLC Rf (CH2Cl2/MeOH: 95/5) 0.3; IR (neat): ν cm-1
1
3305, 1700, 1627, 1188, 1104, 816; H NMR (DMSO[d6], 400
MHz): δ ppm 2.43-2.46 (m, 1H, CCH2CH), 2.82-2.91 (m, 1H,
CCH2CH), 4.05 (dd, J = 9.6, 4.4 Hz, 1H, CCH2CH), 4.24-4.38
(m, 4H, NHCH2), 6.38-6.46 (m, 1H, NHCHC), 6.78 (d, J = 12.4
Hz, 1H, CNHCH2), 7.25 (s, 1H, NHCHCH2), 7.32 (d, J = 8.6 Hz,
1H, ArH), 7.36-7.40 (m, 2H, 2ArH), 7.46 (dd, J = 8.6, 2.3 Hz,
1H, ArH), 7.61 (m, 2H, 2ArH), 8.42 (br t, J = 5.9 Hz, 1H,
CCHNH); 13C NMR (DMSO[d6], 100 MHz): δ ppm 29.2 (CH2),
40.8 (CH2), 48.2 (CH2), 54.0 (CH), 96.8 (C), 127.8 (CH), 128.0
(CH), 129.1 (CH), 129.3 (CH), 130.7 (CH), 131.0 (CH), 132.8
(C), 133.0 (C), 133.4 (C), 135.5 (C), 136.1 (CH), 137.5 (C),
139.0 (C), 173.4 (C), 173.9 (C). Anal. Calcd for C20H17Cl4N3O2:
C, 50.77; H, 3.62; N, 8.88. Found: C, 50.94; H, 3.83; N, 8.48%.
(2S)-(4E)-N-(2,4-dichlorobenzyl)-4-
[(hydroxyimino)methylidene]-5-oxopyrrolidine-2-
carboxamide (15): Compound 5a (0.83 g, 2.09 mmol) was
introduced in THF (20 mL). Glacial acetic acid (2 mL) and
NaNO2 (0.60 g, 8.70 mmol) were added to the mixture which
was then stirred at 0°C for 3 hours. Water (50 mL) and CH2Cl2
(50 mL) were added and the mixture was filtered. The organic
layer was separated and the solvent was evaporated. The desired
product crystalizes in diethyl ether and ethanol. The resulting
precipitate was filtered to afford pure 15 as a white solid in 77%
yield (0.51 g, 1.61 mmol). mp (EtOH) 137-139 °C; TLC Rf
(CH2Cl2/MeOH: 95/5) 0.1; IR (neat): ν cm-1 3265, 2397, 1693,
1652, 981; 1H NMR (DMSO[d6], 400 MHz): δ ppm 2.59 (dd, J =
19.1, 3.1 Hz, 1H, CCH2CH), 3.00 (dd, J = 19.1, 9.2 Hz, 1H,
CCH2CH), 4.22 (dd, J = 9.0, 3.3 Hz, 1H, CH2CH2CH), 4.27-4.39
(m, 2H, NHCH2), 7.38 (d, J = 8.6 Hz, 1H, ArH), 7.43 (dd, J =
8.6, 2.3 Hz, 1H, ArH), 7.62 (d, J = 2.3 Hz, 1H, ArH), 7.73 (br t, J
= 5.5 Hz, 1H, NHCH2), 8.83 (br s, 1H, NHCH), NOH (not seen);
13C NMR (DMSO[d6], 100 MHz): δ ppm 27.5 (CH2), 40.0 (CH2),
51.5 (CH), 127.2 (CH), 128.5 (CH), 130.3 (CH), 132.3 (C), 132.9
(C), 135.0 (C), 150.5 (C), 165.3 (C), 171.6 (C). Anal. Calcd for
C12H11Cl2N3O3: C, 45.59; H, 3.51; N, 13.29. Found: C, 45.83; H,
3.45; N, 12.86%; HRMS (ESI): positive mode [M + Cl]-
(C12H11O3N3Cl2 + Cl): calcd. 349.98605, found 349.98600 g/mol.
(2S)-(4E)-N-(2,4-dichlorobenzyl)-4-(morpholin-4-
ylmethylidene)-5-oxopyrrolidine-2-carboxamide (13d): The
general procedure C was followed using morpholine (0.53 g,
6.08 mmol) and 24 hours of reaction. The resulting mixture was
purified on a silica column (CH2Cl2/MeOH: 1/0 to 9/1) and
washed with diethyl ether to afford pure 13d as a white powder
in 21% yield (0.49 g, 1.28 mmol). mp (Et2O) 165-169 °C; TLC
Rf (CH2Cl2/MeOH: 9/1) 0.4; IR (neat): ν cm-1 3265, 2922, 2852,
1653, 1614, 1110; 1H NMR (CDCl3, 400 MHz): δ ppm 2.93-3.00
(m, 1H, CCH2CH), 3.31-3.44 (m, 5H, CCH2CH and N(CH2)2),
3.69 (t, J = 5.0 Hz, 4H, O(CH2)2), 4.15-4.21 (m, 1H, CCH2CH),
4.40-4.55 (m, 2H, NHCH2), 6.67 (d, J = 13.9 Hz, 1H, NHCH),
6.80 (s, 1H, CCHN), 7.15-7.22 (m, 2H, ArH and NHCH2), 7.29
(d, J = 8.4 Hz, 1H, ArH), 7.36 (d, J = 2.2 Hz, 1H, ArH); 13C
NMR (CDCl3, 100 MHz): δ ppm 30.5 (CH2), 40.9 (CH2), 49.9 (2
CH2), 54.5 (CH), 66.6 (2 CH2), 92.4 (C), 127.3 (CH), 129.3
(CH), 130.8 (CH), 130.9 (C), 133.9 (C), 134.0 (C), 142.1 (CH),
173.2 (C), 176.1 (C). Anal. Calcd for C17H19Cl2N3O3: C, 53.14;
H, 4.98; N, 10.94. Found: C, 53.37; H, 5.09; N, 10.64%.
(2S)-(4E)-N-(2,4-dichlorobenzyl)-4-[(4-methylpiperazin-1-
yl)methylidene]-5-oxopyrrolidine-2-carboxamide (13e): The
general procedure C was followed using N-methylpiperazine
(0.03 g, 0.30 mmol) and 3 hours of reaction. Diethyl ether was
added at the end of the reaction. The desired product crystalizes
as a white powder, which was filtered to afford pure 13e in 67%
yield (0.08 g, 0.20 mmol). mp (Et2O) 136-137 °C; TLC Rf
2-(2,4-dichlorobenzyl)-3-(dimethylamino)tetrahydro-1H-
pyrrolo[1,2-c]imidazole-1,5,6-trione (15) in equilibrium with
enol form: Compound 5a (1.00 g, 2.52 mmol) and NaIO4 (2.69
g, 12.58 mmol) were stirred vigorously in EtOAc/CH3CN (40
mL, 1/1) and water (40 mL) for 1 hour. The organic layer was

10
Tetrahedron
ACCEPTED MANUSCRIPT
5. For selected examples, see: (a) Rigo, B.; Lespagnol, C.; Pauly, M.
collected and washed with brine. The aqueous layer was
J. Heterocycl. Chem. 1988, 25, 49-57. (b) Rigo, B.; Laruelle, C. J.
Heterocycl. Chem. 1997, 34, 279-283. (c) Millet, R.; Meulon, E.;
Goossens, L.; Houssin, R.; Hénichart, J.-P.; Rigo, B. J.
Heterocycl. Chem. 2000, 37, 1491-1494. (d) Bourry, A.; Pitard, F.;
Rigo, B.; Sanz, G.; Camus, F.; Norberg, B.; Durant, F.; Couturier,
D. J. Heterocycl. Chem. 2002, 39, 109-118. (e) Boisse, T.; Gavara,
L.; Gautret, P.; Baldeyrou, B.; Lansiaux, A.; Goossens, J.-F.;
Hénichart, J.-P.; Rigo, B. Tetrahedron Lett. 2011, 52, 1592-1596.
6. Chambers, L. J.; Gleave, R.; Senger, S.; Walter, D. S. WO
2008003697A1 (2008) Chem. Abstr. 2008, 148, 145026.
washed two times with CH2Cl2. The organic phases were
gathered, evaporated and the mixture was purified by column
chromatography (EtOAc/n-heptane: 0/1 to 1/0) to give a yellow
solid in 34% yield (0.30 g, 0.84 mmol). mp (EtOAc) 128-134 °C;
TLC Rf (CH2Cl2/MeOH: 96/4) 0.1; 1H NMR (DMSO[d6], 400
MHz): δ ppm 2.05 (dd, J = 12.7, 9.4 Hz, 1H, CCH2CH), 2.15
(s, 6H, N(CH3)2), 2.21 (s, 6H, N(CH3)2), 2.55 (s, 1H,
CH(OH)), 3.27-3.33 (m, 1H, CHCH2CH), 4.11-4.17 (m, 1H,
CH2CH), 4.26 (dd, J = 18.9, 15.6 Hz, 2H, NCH2), 4.70 (t, J =
15.0 Hz, 2H, NCH2), 4.90 (s, 1H, CH2CH), 5.28 (s, 1H,
NCHN), 5.44 (s, 1H, NCHN), 7.35 (d, J = 8.8 Hz, 2H, ArH),
7.42-7.47 (m, 2H, ArH), 7.59-7.67 (m, 2H, ArH), 10.51 (br s,
1H, OH); 13C NMR (DMSO[d6], 100 MHz): δ ppm 34.8
(CH2), 37.8 (2 CH3), 37.8 (CH2), 37.9 (2 CH3), 40.8 (CH2),
41.8 (CH2), 55.2 (CH), 61.1 (CH), 77.0 (CH), 86.4 (CH), 87.2
(CH), 123.2 (CH), 128.1 (2 CH), 129.4 (CH), 132.0 (CH),
132.4 (C), 132.6 (C), 132.9 (C), 133.6 (2 C), 134.0 (2 C),
144.3 (C), 167.5 (C), 170.8 (C), 174.0 (C), 177.2 (C). Anal.
Calcd for C15H15Cl2N3O3: C, 50.58; H, 4.24; N, 11.80%. Found:
C, 50.55; H, 4.27; N, 11.82%.
7. Homerin, G.; Baudelet, D.; Dufrénoy, P.; Rigo, B.; Lipka, E.;
Dezitter, X.; Furman, C.; Millet, R.; Ghinet, A. Tetrahedron Lett.
2016, 57, 1165-1170.
8. Batcho, A. D.; Leimgruber, W. Org. Synth. 1985, 63, 214-225.
9. Guzman, A.; Romero, M.; Maddox, M. L.; Muchowski, J. M. J.
Org. Chem. 1990, 55, 5793-5797.
10. For selected publications, see: (a) Gold, H., Angew. Chem. 1960,
24, 956-959. (b) Schuppe, A. W.; Cabrera, M. J.; McGeoch, C. L.
B.; Newhouse, T. R., Tetrahedron, 2017, 73, 3646-3651.
11. For selected reviews, see: (a) Stanovnik, B.; Svete, J. Chem. Rev.
2004, 104, 2433-2480. (b) Rosso, G. B. Synlett. 2006, 5, 809-810.
(c) Kantlehner, W. J. prakt. Chem. 1995, 337, 418-421.
12. Baudelet, D.; Daich, A.; Rigo, B.; Lipka, E.; Gautret, P.; Homerin,
G.; Claverie, C.; Rousseau, J.; Abuhaie, C.-M.; Ghinet, A.
Synthesis 2016, 48, 2226-2244.
h-P2X7R screening procedure:
13. Search on the 3-aminotetrahydro-1H-pyrrolo[1,2-c]imidazole-1,5-
dione scaffold using SciFinder® (5/03/2017) resulted only in 3
commercial compounds, with no description of synthesis or
physical data.
14. Cell proliferation assay: The cell proliferation assay was realized
with compounds 2d, 5a, 5e, 5f, 5k, 5q, 11, and 13b-e on a panel
of 60 human cancer cell lines at the National Cancer Institute,
Germantown, MD, USA and was conducted using a 48 h exposure
using the sulfurhodamine B assay.
15. h-P2X7R screening procedure: HEK-293 cells overexpressing
human P2X7R were stained with the calcium indicator Fluo-3 AM
and then incubated with the non-permeant DNA intercalants TO-
PRO-3 (large pore opening) and propidium iodide (cell viability)
in presence of different molecules at 10-5M concentration for 15
min before addition of the P2X7R agonist BzATP (100 ꢀM).
Compound 1 was used as a reference antagonist for human
P2X7R. Cells were then analyzed after 1 hour by flow cytometry
using Cyan cytometer (Beckman). Percentages of cells displaying
P2X7R activation (Fluo-3/TO-PRO-3 double positive cells) in
presence of the molecules were calculated.
HEK-293 cells overexpressing human P2X7R were stained
with the calcium indicator Fluo-3 AM and then incubated with
the non-permeant DNA intercalants TO-PRO-3 (large pore
opening) and propidium iodide (cell viability) in presence of
different molecules at 10-5 M concentration for 15 min before
addition of the P2X7R agonist BzATP (100 ꢀM). Compound 1
was used as a reference antagonist for human P2X7R. Cells were
then analysed after 1 hour by flow cytometry using Cyan
cytometer (Beckman). Percentages of cells displaying P2X7R
activation (Fluo-3/TO-PRO-3 double positive cells) in presence
of the molecules were calculated.
Cell proliferation assay:
The cell proliferation assay was realized with compounds 2d,
5a, 5e, 5f, 5k, 5q, 11 and 13b-e on a panel of 60 human cancer
cell lines at the National Cancer Institute, Germantown, MD,
USA and was conducted using a 48 h exposure using the
sulfurhodamine B assay. One dose mean graphics are given in
supporting information.
Acknowledgements
The authors gratefully acknowledge the “Métropole
Européenne de Lille” (MEL) for G.H.’s PhD scholarship, the
DigestScience Foundation (Foundation on digestive tract diseases
and nutrition) and the INCa (Institut National du Cancer, France)
for financial support. The authors also acknowledge the National
Cancer Institute (NCI) for biological evaluation of compounds on
their 60-cell panel: the testing was performed by the
Developmental Therapeutics Program, Division of Cancer
Treatment and Diagnosis (the URL to the Program's website:
http://dtp.cancer.gov).
References and notes
1. Di Virgilio, F. Biomed. J. 2016, 39, 326-338, and references cited
therein.
2. Baudelet, D.; Lipka, E.; Millet, R.; Ghinet, A. Curr. Med. Chem.
2015, 22, 713-729, and references cited therein.
3. Guile, S. D.; Alcaraz, L.; Birkinshaw, T. N.; Bowers, K. C.;
Ebden, M. R.; Furber, M.; Stocks, M. J. J. Med. Chem. 2009, 52,
3123-3141.
4. Barlett, R.; Stokes, L.; Sluyter, R. Pharmacol. Rev. 2014, 66, 638-
675, and references cited therein.