Solid and solution phase syntheses of the 2-cyanopyrrolidide DPP-IV inhibitor NVP-DPP728

Article abstract of DOI:10.1016/S0040-4020(02)00536-7

DPP-IV inhibitors have been suggested as potential new treatments for type-II diabetes and 2-cyanopyrrolidides have been reported as potent DPP-IV inhibitors. Alternative synthetic approaches to one such compound, NVP-DPP728, are investigated here. One strategy is based in solution phase and is amenable to scale-up. The other is based on solid phase and is appropriate for the rapid analoging of the structural series.

Full text of DOI:10.1016/S0040-4020(02)00536-7

TETRAHEDRON
Pergamon
Tetrahedron ꢀ8 52002) ꢀ741±ꢀ746
Solid and solution phase syntheses of the 2-cyanopyrrolidide
DPP-IV inhibitor NVP-DPP728
a
Nicolas Willand, Jurgen Joossens, Jean-Claude Gesqui eÁ re, Andr e L. Tartar, D. Michael Evans
a
a
a
b
b,p
and Michael B. Roe
a
Laboratoire de chimie organique, UMR 8525, Facult e des sciences pharmaceutiques et biologiques, 3 rue du Pr. Laguesse,
F-59006 Lille Cedex, France
b
Ferring Research Limited, Chilworth Research Centre, Southampton SO16 7NP, UK
Received 22 January 2002; revised 22 April 2002; accepted 10 May 2002
AbstractÐDPP-IV inhibitors have been suggested as potential new treatments for type-II diabetes and 2-cyanopyrrolidides have been
reported as potent DPP-IV inhibitors. Alternative synthetic approaches to one such compound, NVP-DPP728, are investigated here. One
strategy is based in solution phase and is amenable to scale-up. The other is based on solid phase and is appropriate for the rapid analoging of
the structural series. q 2002 Elsevier Science Ltd. All rights reserved.
Dipeptidyl peptidase IV 5DPP-IV, CD26, EC.3.4.14.ꢀ) is a
membrane-bound and circulating serine protease. It
catalyses the hydrolysis of peptides after a penultimate
in phase II clinical trials as a potential new drug therapy for
type-II diabetes.
1
N-terminal proline or alanine residue. One such peptide
The preparation of NVP-DPP728 1 is described in Scheme
1. In our hands, this procedure gave a very poor yield. This
ꢀ
is glucagon-like peptide-1 5GLP-1). Hence, inhibition of
DPP-IV has been suggested as a novel treatment for type-
II diabetes. A series of 2-cyanopyrrolidides has been
is mainly due to the number of reactive amine functional
groups present in both the starting substrate 5 and in the
product 1. Therefore, we investigated alternative routes
that would avoid formation of by-products and/or allow
easier preparation of analogues. We have developed two
different strategies to the synthesis of NVP-DPP728 1.
One makes use of an original protection and deprotection
2
reported by Ferring Research Limited as very potent
inhibitors of DPP-IV. More recently one 2-cyanopyrroli-
3
dide, NVP-DPP728 1, has been reported to increase plasma
GLP-1 57-36 amide) concentrations and improve oral
4
glucose tolerance in obese Zucker rats. This compound is
ꢀ
Scheme 1. The solution phase synthesis of NVP-DPP728 as reported in patent application WO 98/19998.
Keywords: NVP-DPP728; DPP-IV inhibitor; cyanopyrrolidide.
Corresponding author. Tel.: 144-23-8076-3400; fax: 144-23-8076-62ꢀ3; e-mail: michael.roe@ferring-research.co.uk
p
0040±4020/02/$ - see front matter q 2002 Elsevier Science Ltd. All rights reserved.
PII: S0040-4020502)00ꢀ36-7

ꢀ742
N. Willand et al. / Tetrahedron 58 ꢀ2002) 5741±5746
strategy. This is a solution phase synthesis and is suitable for
scale-up. The other, performed on solid phase, is amenable
to the fast preparation of structural analogues.
the crystallisation remains to be optimised, the chemistry is
convenient to carry out and is amenable to scale-up. The
free base of 8 was liberated and coupled to bromide 3 to give
6
9
in 61% yield. It was necessary to remove the formalde-
hyde group. The literature precedent was unsuitable because
the high polarity of the product made isolation from malonic
acid and pyridine dif®cult. Instead, because of the reversi-
1
.Solution phase strategy
8
The major drawback of the existing synthesis is the presence
of several nucleophilic groups during the alkylation step. In
selecting an appropriate protection of the reactive amine
functionalities, an important consideration is the suscepti-
bility for cyclisation under strongly basic conditions.
Nucleophilic attack by the secondary amine onto the nitrile
in 1 affords a six-membered cyclic amidine that can
hydrolyse to a diketopiperazine. Under non-basic con-
ditions, the secondary amine is less prone to cyclisation.
Depending on the example or the conditions employed,
amino acid 2-cyanopyrrolidides demonstrate half-lives
bility of the formaldehyde condensation under aqueous
conditions, it was hypothesised that simple acid catalysed
hydrolysis under dilute conditions would be suf®cient.
Therefore, the formaldehyde adduct 9 was dissolved in a
10% TFA 5aq.) solution and stirred for 24 h. Basic work-
up and ¯ash chromatography afforded the title compound 1
in a ꢀ0% yield. The remaining material was identi®ed as
unreacted 9. There was no evidence of intramolecular cycli-
sation. Under these conditions, the ®nal deprotection step is
reversible and an equilibrium mixture is obtained. In order
to drive the reaction to completion, high dilution conditions
were considered. However, such conditions are not
amenable to scale-up. Therefore, we decided to use a
supported scavenging reagent. Excess 3-54-5hydrazinosul-
fonyl)phenyl)propionyl AM resin was suspended in a
solution of intermediate 9 in a mixture of 10% TFA/90%
CH Cl and stirred for an hour at room temperature.
3
,7
greater than 24 h. Therefore, we were prompted to investi-
gate protecting groups that may be removed under neutral or
acidic conditions.
We were attracted by a strategy used for the selective
protection of spermidine derivatives. This involved the
8
2
2
condensation of spermidine with formaldehyde yielding a
cyclic product. In this way, the primary amino group is
converted to a secondary amine, while the secondary
amine is converted to a tertiary amine. Subsequent removal
of the methylene group was carried out with malonic acid
and pyridine in methanol. Scheme 2 shows the synthetic
route to NVP-DPP728 1 using a strategy similar to this.
The amine´TFA salt 5 was prepared according to Scheme
After work-up and chromatography the free base of NVP-
DPP728 1 was obtained in 7ꢀ% yield. Thus the hydrazine
scavenging reagent was successful at improving the yield of
the ®nal deprotection step, reducing the reaction time
dramatically and eliminating the need for high-dilution
conditions.
2
and condensed with aqueous formaldehyde solution. This
proceeded smoothly to yield the cyclic formaldehyde adduct
´TFA salt which was isolated by crystallisation from a 1:1
mixture of diethyl ether and ethanol. Although the yield of
2.Solid phase strategy
8
First of all we prepared the Rink resin bound key
Scheme 2. The solution phase preparation of NVP-DPP728 1 using a formaldehyde derived protecting group.

N. Willand et al. / Tetrahedron 58 ꢀ2002) 5741±5746
ꢀ743
Scheme 3. The synthesis of NVP-DPP728 1 using a solid phase approach via the key BOC protected intermediate 8.
intermediate 15 in high yield using similar chemistry to that
already reported 5Scheme 3). This would allow rapid
group strategy that would allow speci®c addition of
6-chloronicotinonitrile 6 and other electrophiles to the
terminal nitrogen under forcing conditions. Initial attempts
to use formaldehyde, as in the solution phase synthesis,
proved unsuccessful. Complex mixtures were obtained
after cleavage from the resin. Accordingly, 2-acetyl-
dimedone was added in DMF to the resin 15 to afford the
6
derivatisation of the primary amino group. Our initial
attempts to exemplify this with the preparation of NVP-
DPP728 were not successful. Scheme 3 illustrates three
different sets of experimental conditions, all employing a
large excess of 6-chloronicotinonitrile 6 510 equiv.). Using
potassium carbonate as base gave no reaction products.
Using diisopropylethylamine in either THF or N-methyl-
pyrrolidone yielded mixtures of the required product 16
and the double addition product 17 as well as other
unidenti®ed products. This led us to investigate a protecting
9
Dde protected primary amine. Addition of BOC anhydride
followed by deprotection of the Dde group with hydrazine
in DMF afforded the BOC protected compound 19.
6-Chloronicotinitrile 6 was added to the terminal amine
under forcing conditions 548 h at 808C). After cleavage

ꢀ744
N. Willand et al. / Tetrahedron 58 ꢀ2002) 5741±5746
from the resin and concomitant removal of the BOC group,
the carboxamide was dehydrated under standard conditions.
All steps in the solid phase synthesis proceeded cleanly such
to be quantitative and the material was used directly in the
next step.
1
that H NMR showed the crude material obtained upon
4.1.3. 6-Imidazolidin-1-yl-nicotinonitrile )8ꢀ. 6-52-Amino-
ethylamino)nicotinonitrile tri¯uoroacetate 5 5120 mmol)
was dissolved in water 51300 ml). Aqueous 37% formalde-
hyde solution 51ꢀ.ꢀ g, 12.ꢀ ml, 168 mmol) was added and
the mixture was stirred for 3 days. The mixture was concen-
trated then azeotroped with toluene twice and with
cleavage from the resin and dehydration to be predomi-
nantly the required tri¯uoroacetamide 21. After chromato-
graphy, treatment with methanolic ammonia afforded the
free base of NVP-DPP728 1. This procedure illustrates an
orthogonal protecting group strategy that affords the BOC
protected resin bound intermediate 19. This is a common
intermediate that may be utilized for the rapid analoging of
the terminal N-substituent using large sets of acylating and
alkylating agents.
petroleum ether once. The residue was taken up in Et O/
2
EtOH 5ꢀ0:ꢀ0, 200 ml) and scratched to initiate crystallisa-
tion. The mixture was cooled in an ice/water bath for 4 h and
®ltered to afford 6-imidazolidin-1-yl-nicotinonitrile
tri¯uoroacetate 511.4 g) as a pale yellow crystalline solid.
H NMR 5d -DMSO, 270 MHz): d 3.ꢀ8±3.78 54H, m), 4.7ꢀ
1
6
5
8
2H, s), 6.78 51H, d, Jˆ8.9 Hz), 8.01 51H, dd, Jˆ2.2,
3.Summary
.9 Hz), 8.ꢀ8 51H, d, Jˆ2.2 Hz) ppm. This was dissolved
in water and saturated NaHCO solution was added until the
3
We have presented two new chemistries for the preparation
of NVP-DPP728 1. The solid phase approach utilises an
orthogonal protecting group strategy to afford a BOC
protected intermediate. This may be used to prepare the
title compound or may be used as a common intermediate
for the rapid synthesis of a series of compounds for SAR
analysis. Complementary to this, the solution phase
approach utilises a new and ef®cient protecting group
strategy that is amenable to scale-up. Considering the
perceived importance of DPP-IV inhibitors in the ®eld of
diabetes these chemistries will be of bene®t to any groups
active in the ®eld.
solution was basic. The mixture was extracted with CH Cl
2
2
four times and the combined organic extracts were dried
over Na SO and concentrated in vacuo to afford 6-imida-
zolidin-1-yl-nicotinonitrile free base 8 56.7 g, 33%) as a
2
4
1
yellow oil. H NMR 5CDCl , 270 MHz): d 3.26±3.48
3
5
4H, m), 4.46 52H, s), 6.28 51H, d, Jˆ8.7 Hz), 7.ꢀ7 51H,
dd, Jˆ2.2, 8.7 Hz), 8.37 51H, d, Jˆ2.2 Hz) ppm.
4.1.4. 6-{3-[2-)2-S-Cyanopyrrolidin-1-ylꢀ-2-oxo-ethyl]-
imidazolidin-1-yl}-nicotinonitrile )9ꢀ. A solution of 52S)-
ꢀ
-5bromoacetyl)pyrrolidine-2-carbonitrile 3, 53.94 g, 18.1
1
mmol) in THF 520 ml) was added to an ice-cold solution
of 6-imidazolidin-1-yl-nicotinonitrile 8 53.16 g, 18.1 mmol)
and triethylamine 52.8 ml, 2.0 g, 20 mmol) in THF 520 ml)
over 3 min. The resulting cloudy mixture was allowed to
warm to room temperature and stirred for 6 h. The mixture
4.Experimental
4
4
.1. Data for compounds
was added to dilute NaHCO
CH Cl four times. The combined organic extracts were
solution and extracted with
3
2
2
.1.1. tert-Butyl )2-)5-cyano-2-pyridylaminoꢀethylꢀcar-
dried over Na
SO
2
4
and concentrated in vacuo. Flash chro-
Cl )
2
bamate )7ꢀ. A solution of 6-chloronicotinonitrile 56)
matography on silica gel 52.ꢀ% MeOH/97.ꢀ% CH
2
5
5
5
21.7 g, 160 mmol), potassium hydrogen carbonate
17.6 g, 176 mmol) and tert-butyl 52-aminoethyl)carbamate
2ꢀ.8 g, 160 mmol) in DMF 580 ml) was heated to 908C
afforded 6-{3-[2-52-S-cyanopyrrolidin-1-yl)-2-oxo-ethyl]-
imidazolidin-1-yl}-nicotinonitrile 9 53.4 g, 61%) as a
, 270 MHz): d 2.04±2.42
1
white foam. H NMR 5CDCl
3
under N for 2.ꢀ h. The mixture was cooled, added to satu-
2
rated NaHCO solution and extracted with EtOAc. The
3
54H, m), 3.18 52H, t, Jˆ6.7 Hz), 3.34±3.76 56H, m) 4.26±
4.42 52H, m), 4.74±4.80 and 4.9ꢀ±ꢀ.04 5total 1H in the ratio
3:1, each m), 6.30 51H, d, Jˆ8.9 Hz), 7.60 51H, dd, Jˆ2.0,
8.9 Hz), 8.38 51H, d, Jˆ2.0 Hz) ppm. MS 5ESI) m/z 311.0
organic phase was dried over Na SO and concentrated in
2
4
vacuo until the product started to precipitate. Further
product was precipitated by adding petroleum ether. The
combined precipitates were collected and washed with
cold EtOAc to afford tert-butyl 52-5ꢀ-cyano-2-pyridyl-
1
5MH ).
4.1.5. )6-{2-[2-)2-S-Cyanopyrrolidin-1-ylꢀ-2-oxo-ethyl-
amino]-ethylamino}-nicotinonitrile )1ꢀ, by removal of
formaldehyde group using aqueous tri¯uoroacetic acid.
A solution of 6-{3-[2-52-S-cyanopyrrolidin-1-yl)-2-oxo-
ethyl]-imidazolidin-1-yl}-nicotinonitrile 9 53.1ꢀ g, 10.2
mmol) in 10% tri¯uoroacetic acid 52ꢀ0 ml) was stirred for
amino)ethyl)carbamate 7 527.2 g, 6ꢀ%) as a white solid.
H NMR 5CDCl , 270 MHz): d 1.41 59H, s), 3.30±3.42
1
3
5
2H, m), 3.44±3.ꢀ6 52H, m), 4.92 51H, br s), ꢀ.72 51H, br
s), 6.39 51H, dd, Jˆ8.9, 0.7 Hz), 7.ꢀ1 51H, dd, Jˆ8.9,
2
.2 Hz), 8.30±8.36 51H, m) ppm.
2
4 h. The mixture was cooled in an ice/water bath and
potassium carbonate was added cautiously until saturated,
then extracted with CH Cl seven times. The combined
extracts were dried over Na SO and concentrated in
vacuo. Flash chromatography on silica gel 510% MeOH/
90% CH Cl ) afforded 56-{2-[2-52-S-cyanopyrrolidin-1-
4
.1.2. 6-)2-Aminoethylaminoꢀnicotinonitrile tri¯uoro-
acetate )5ꢀ. Tri¯uoroacetic acid 512ꢀ ml) was added to an
ice-cold stirred suspension of tert-butyl 52-5ꢀ-cyano-2-
pyridylamino)ethyl)carbamate 7 532.0 g, 120 mmol) in
CH Cl 512ꢀ ml) to give a clear solution. After gas evolution
2
2
2
4
2
2
2
2
had ceased, the cooling bath was removed. After 1.ꢀ h the
mixture was concentrated in vacuo and azeotroped with
toluene three times to afford 6-52-aminoethylamino)-
nicotinonitrile tri¯uoroacetate 5. The yield was assumed
yl)-2-oxo-ethylamino]-ethylamino}-nicotinonitrile 1 51.ꢀ3 g,
ꢀ0%) as a colourless gum. H NMR at 2ꢀ8C was consistent
with the presence of two rotameric isomers in a ratio of
1
1
,8ꢀ:1ꢀ. H NMR 5d
-DMSO, 270 MHz): d 1.84±2.30
6

N. Willand et al. / Tetrahedron 58 ꢀ2002) 5741±5746
ꢀ74ꢀ
5
4H, m), 2.71 52H, t, Jˆ6.2 Hz), 3.22±3.48 5ꢀH, m), 3.ꢀ0±
yellow solution was shaken at 08C for 30 min then added
to the resin. The reaction was shaken for 3 h at 208C until a
ninhydrin test was negative. Washing sequentially with
DMF, CH Cl and DMF afforded Fmoc-l-Pro-resin 12.
3
6
7
.66 51H, m), 4.73 and ꢀ.13 51H, ratio ,8ꢀ:1ꢀ, dd, Jˆ4.7,
.4 Hz and dd, Jˆ2.ꢀ, 6.7 Hz), 6.ꢀ6 51H, d, Jˆ8.9 Hz),
.ꢀ0±7.72 52H, m), 8.36 51H, d, Jˆ2.0 Hz) ppm. At higher
2
2
temperatures, the NMR signals of the two compounds
appeared to coalesce as is typical of rotameric isomers. H
NMR 5d -DMSO, 270 MHz): d 1.98±2.30 54H, m), 2.77
5
6
Resin 12 was then shaken at room temperature with a
20% piperidine solution in DMF 5ꢀ0 ml) for ꢀ min. The
mixture was ®ltered and shaken with 20% piperidine in
DMF 5ꢀ0 ml) for another 20 min. Washing sequentially
with DMF, CH Cl and DMF afforded l-Pro-resin 13
1
6
2H, t, Jˆ6.2 Hz), 3.30±3.62 56H, m), 4.62±4.88 51H, m),
.ꢀ7 51H, d, Jˆ8.9 Hz), 7.26 51H, br s), 7.61 51H, dd, Jˆ2.2,
.9 Hz), 8.33 51H, d, Jˆ2.2 Hz) ppm.
2
2
8
which was used directly in the next step. A few beads of
the resin 13 were cleaved with tri¯uoroacetic acid to afford
1
l-Pro-NH . MS 5ESI) m/z 11ꢀ 5MH , 100%).
4
.1.6. )6-{2-[2-)2-S-Cyanopyrrolidin-1-ylꢀ-2-oxo-ethyl-
2
amino]-ethylamino}-nicotinonitrile )1ꢀ, by removal of
formaldehyde group using solid supported reagent.
4.1.9. N-Bromoacetyl-l-Pro-resin )14ꢀ. A solution of
bromoacetic acid 53.3 g, 24 mmol) in DMF 530 ml) was
added to l-Pro-resin 13 52.4 mmol) followed by a solution
of DIC 54.8 ml, 3.9 g, 31 mmol) in DMF 5ꢀ ml) and the
mixture was shaken at room temperature for 30 min. The
resin was ®ltered and the procedure repeated once. Washing
sequentially with DMF, CH Cl and DMF afforded
6
-{3-[2-52-S-Cyanopyrrolidin-1-yl)-2-oxo-ethyl]-imidazo-
lidin-1-yl}-nicotinonitrile 9 5223 mg, 0.72 mmol) and 3-54-
hydrazinosulfonyl)phenyl) propionyl AM resin 51.3 g,
.9 mmol) were dissolved in CH Cl 520 ml). Tri¯uoro-
5
1
2
2
acetic acid 53 ml) was added and the reaction was stirred
at room temperature for 1 h. The organic layer was ®ltered
and the resin was washed with CH Cl 5100 ml) and EtOH
2
2
N-bromoacetyl-l-Pro-resin 14 which was used directly in
the next step. A few beads of the resin 14 were cleaved
with tri¯uoroacetic acid to afford N-bromoacetyl-l-Pro-
2
2
5
vacuo. The residue was dissolved in CH Cl 5100 ml) and
200 ml). The combined organic layers were evaporated in
2
2
1
washed with potassium carbonate solution 530 ml). The
aqueous layer was extracted with CH Cl three times and
NH . MS 5ESI) m/z 236 5MH , 100%).
2
2
2
the combined organic layers were evaporated in vacuo.
Flash chromatography on silica gel 590% CH Cl /10%
MeOH and 80% CH Cl /20% MeOH) afforded 56-{2-[2-
4.1.10. N-)2-Aminoethylꢀ-Gly-l-Pro-resin )15ꢀ. A solu-
tion of ethylenediamine 52.3 ml, 2.1 g, 3ꢀ mmol) in
DMSO 530 ml) was added to the resin 14 52.4 mmol) and
the mixture was shaken for 2 h. Washing sequentially with
DMF, CH Cl and DMF afforded N-52-aminoethyl)-Gly-l-
2
2
2
2
5
2-S-cyanopyrrolidin-1-yl)-2-oxo-ethylamino]-ethylamino}-
1
nicotinonitrile 1 5162 mg, 7ꢀ%) as a colourless gum. H
NMR data was identical to that obtained by the aqueous
tri¯uoroacetic acid route above.
2
2
Pro-resin 15 which was used directly in the next step. A few
beads of the resin 15 were cleaved with tri¯uoroacetic acid
to afford N-52-aminoethyl)-Gly-l-Pro-NH . MS 5ESI) m/z
2
1
4
.1.7. )6-{2-[2-)2-S-Cyanopyrrolidin-1-ylꢀ-2-oxo-ethyl-
21ꢀ 5MH , 100%).
amino]-ethylamino}-nicotinonitrile )1ꢀ, hydrochloride
salt. 4N HCl/dioxan 51.3 ml, ꢀ.1 mmol) was added to a
solution of 56-{2-[2-52-S-cyanopyrrolidin-1-yl)-2-oxo-
ethylamino]-ethylamino}-nicotinonitrile 1 51.ꢀ3 g, ꢀ.13
mmol) to afford a white precipitate. The solid was ®ltered,
4.1.11. N-)N-[1-{4,4-Dimethyl-2,6-dioxo-cyclohexylidene}-
ethylamino]-2-aminoethylꢀ-Gly-l-Pro-resin )18ꢀ. A solu-
tion of 2-acetyldimedone 50.69 g, 3.8 mmol) in DMF
510 ml) was added to a proportion of the resin 15 5ꢀ.1 g,
1.9 mmol) in DMF 520 ml). The mixture was shaken at
room temperature for 3 h until a negative ninhydrin test.
Washing sequentially with DMF, CH Cl and DMF
washed and triturated with Et O and dried in vacuo over
2
P2Oꢀ to 56-{2-[2-52-S-cyanopyrrolidin-1-yl)-2-oxo-ethyl-
amino]-ethylamino}-nicotinonitrile 1 hydrochloride salt
2
2
1
1.40 g, 82%) as a white powder. H NMR 5d -DMSO,
5
afforded
N-5N-[1-{4,4-dimethyl-2,6-dioxo-cyclohexyl-
6
2
5
70 MHz): d 1.88±2.30 54H, m), 3.13 52H, s), 3.32±3.ꢀ0
1H, m), 3.ꢀ2±3.98 54H, m), 4.00±4.22 51H, m), 4.84 51H, t,
Jˆꢀ.6 Hz), 6.64 51H, d, Jˆ8.9 Hz), 7.76 51H, dd, Jˆ8.9,
.1 Hz), 8.00 51H, br s), 8.43 51H, d, Jˆ2.1 Hz), 9.2ꢀ 52H,
idene}-ethylamino]-2-aminoethyl)-Gly-l-Pro-resin 18 which
was used directly in the next step. A few beads of the resin
18 were cleaved with tri¯uoroacetic acid to afford N-5N-[1-
2
{4,4-dimethyl-2,6-dioxo-cyclohexylidene}-ethylamino]-2-
aminoethyl)-Gly-l-Pro-NH tri¯uoroacetate salt. H NMR
1
3
1
br s) ppm. C NMR 5D O, 68 MHz): d 24.7, 29.6, 38.2,
2
2
4
1
1
6.3, 47.0, 47.1, 48.2, 97.3, 111.ꢀ, 117.9, 118.7, 141.6,
49.ꢀ, 1ꢀ6.8, 16ꢀ.0 ppm. MS 5ESI) m/z 299 5MH ,
00%). An analytical sample was recystallised from
5d -DMSO, 300 MHz): d 0.99 56H, s), 1.82±2.06 54H, m),
6
1
2.29 54H, s), 2.ꢀ0 53H, s), 3.1ꢀ±3.23 52H, m), 3.40±3.ꢀ0
52H, m), 3.73±3.81 52H, m), 4.03 52H, s), 4.26 51H, m), 4.ꢀ0
52H, s), 9.00 52H, s), 13.13±13.21 51H, m) ppm. MS 5ESI)
MeOH, mp 1ꢀ8±1638C 5reported mp from WO98/19998,
example no. 3ˆ1ꢀꢀ±1ꢀ78C).
4
1
m/z 379 5MH , 100%).
4
0
.1.8. l-Pro-resin )13ꢀ. Rink amide resin 10 5ꢀ g,
.47 mmol/g, 2.4 mmol) was shaken at room temperature
4.1.12. N-)2-Aminoethylꢀ-Boc-Gly-l-Pro-resin )19ꢀ. A
solution of di-tert-butyl dicarbonate 52.0 g, 9.2 mmol) and
i
EtN Pr 50.6ꢀ ml, 3.8 mmol) in dioxan 530 ml) was added to
with a 20% piperidine solution in DMF 5ꢀ0 ml) for ꢀ min.
The mixture was ®ltered and shaken with 20% piperidine in
DMF 5ꢀ0 ml) for another 20 min. The resin was then
washed sequentially with DMF, CH Cl and DMF. TBTU
2
resin 18 51.9 mmol) in dioxan 510 ml). The reaction was
shaken for 2 h at room temperature. The resulting resin
was washed sequentially with DMF, CH Cl and DMF.
2
2
2
2
i
51.9 g, ꢀ.9 mmol), HOBT 50.79 g, ꢀ.9 mmol) and EtN Pr
53.0 ml, 17 mmol) were added to a solution of Fmoc-l-
The resin was treated with 2% hydrazine/DMF solution
under continuous ¯ow conditions 53 ml/min) for 20 min.
Washing sequentially with DMF, CH Cl and DMF
2
Proline 52.0 g, ꢀ.9 mmol) in DMF 530 ml). The resulting
2
2

ꢀ746
N. Willand et al. / Tetrahedron 58 ꢀ2002) 5741±5746
afforded N-52-aminoethyl)-Boc-Gly-l-Pro-NH2 resin 19
which was used directly in the next step.
ethyl]-acetamide 21 520 mg, 0.0ꢀ mmol) in CH Cl 52 ml).
2 2
The reaction was stirred for 3 h. The solvent was removed
under vacuum to give 56-{2-[2-52-S-cyanopyrrolidin-1-yl)-
4
.1.13. N-)N-[5-Cyanopyridin-2-yl]-2-aminoethylꢀ-Boc-
Gly-l-Pro-resin )20ꢀ. 6-Chloronicotinonitrile 51.3 g,
2-oxo-ethylamino]-ethylamino}-nicotinonitrile
colourless gum 512 mg, 80%). Analytical data 5 H,
NMR and MS) for this free base and after conversion to
the hydrochloride salt were identical to those obtained
from the solution phase synthesis.
1
as 13
a
C
1
i
.ꢀ mmol) and EtN Pr 51.6 ml, 1.2 g, 9.ꢀ mmol) were
9
2
added to the resin 19 51.9 mmol) in N-methylpyrrolidinone
30 ml). The mixture was heated at 808C and shaken for
8 h. Washing sequentially with DMF, CH Cl and DMF
5
4
2
2
afforded N-5N-[ꢀ-cyanopyridin-2-yl]-2-aminoethyl)-Boc-
Gly-l-Pro-resin 20 which was used directly in the next step.
References
1
2
. Mentlein, R. Regul. Pept. 1999, 85, 9±24.
4.1.14. N-[2-)5-Cyanopyridin-2-ylaminoꢀ-ethyl]-N-[2-)2-
S-cyanopyrrolidin-1-ylꢀ-2-oxo-ethyl]-acetamide )21ꢀ.
. Deacon, C. F.; Nauck, M. A.; Toft-Nielsen, M-B.; Pridal, L.;
Willms, B.; Holst, J. J. Diabetes 1995, 44, 1126±1131.
. Ashworth, D. M.; Atrash, B.; Baker, G. R.; Baxter, A. J.;
Jenkins, P. D.; Jones, D. M.; Szelke, M. Bioorg. Med. Chem.
Lett. 1996, 6, 1163±1166.
The resin 20 51.9 mmol) was added to a mixture of tri¯uoro-
acetic acid and CH Cl 51:1) solution and stirred for 2 h at
3
2
2
room temperature. The supernatant was added to Et O
2
to precipitate 1-{2-[2-5ꢀ-cyanopyridin-2-ylamino)-ethyl-
amino]-acetyl}-pyrrolidine-2-S-carboxylic acid amide
4
ꢀ
6
. Balkan, B.; Kwasnik, L.; Miserendino, R.; Holst, J. J.; Li, X.
Diabetologia 1999, 42, 1324±1331.
1
tri¯uoroacetate salt 50.70 g, 87%). H NMR 5d -DMSO,
6
. Villhauer, E. B. International Patent Application Number PCT/
EP97/0612ꢀꢀ, published as WO98/19998 1998.
3
3
4
7
00 MHz): d 1.82±2.0ꢀ 54H, m), 3.08±3.16 52H, m),
.40±3.ꢀ0 52H, m), 3.ꢀ9±3.67 52H, m), 4.02 52H, s),
.22±4.30 51H, m), ꢀ.40 52H, s), 6.60 51H, d, Jˆ8.8 Hz),
. While this work was in progress an oral communication was
presented that described the synthesis of NVP-DPP728 on solid
phase. It differs from the one reported here in the order that the
steps were carried out and in the choice of protecting groups.
The published work is amenable to analoging of the ethylene
sidechain while the one reported here is more suitable for
analoging of the terminal aromatic group. Villhauer, E. B.;
Anderson, R. C.; Balkan, B.; Barilla, D.; Brinkman, J. A.;
Dunn, E.; Dunning, B.; Graham, E. D.; Gu, H. H.; Gutierrez,
C. M.; Hamilton, B. H.; Kwasnik, L. A.; Li, X.; Mangold, B. L.;
Maniara, W. M.; Miserendino-Molteni, R.; Mone, M.; Naderi,
G. B.; Ramos, K. L.; Russell, M. E.; Rothenberg, P. L.;
Tullman, R. H.; Valentin, M.; Walter, R. E.; Weldon, S. C.;
Hughes, T. E. Abstracts of Papers, #343, 221st National Meet-
ing of the American Chemical Society, San Diego, CA, 2001.
. Hughes, T. E.; Mone, M. D.; Russell, M. E.; Weldon, S. C.;
Villhauer, E. B. Biochemistry 1999, 38, 11ꢀ97±11603.
. Almeida, M. L. S.; Grehn, L.; Ragnarsson, U. J. Chem. Soc.,
Perkin Trans. 1 1988, 190ꢀ±1911.
.74 51H, dd, Jˆ8.8, 2.1 Hz), 7.81 51H, s), 8.42 51H, d,
1
Jˆ2.1 Hz), 9.00 52H, s) ppm. MS 5ESI) m/z 317 5MH ,
1
00%). This precipitate 50.ꢀ4 g, 1.3 mmol) was dissolved
in anhydrous THF 52ꢀ ml) at 08C. Tri¯uoroacetic anhydride
0.88 ml, 6.3 mmol) was added. The reaction was stirred at
8C for 2 h. The solvent was removed under vacuum. Flash
chromatography on silica gel 52% MeOH/98% CH Cl and
5
0
2
2
ꢀ
ylamino)-ethyl]-N-[2-52-S-cyanopyrrolidin-1-yl)-2-oxo-
% MeOH/9ꢀ% CH Cl ) afforded N-[2-5ꢀ-cyanopyridin-2-
2 2
1
ethyl]-acetamide 21 as a white solid 50.3ꢀ g, 70%). H NMR
5
5
5
5
d6-DMSO, 300 MHz) d 1.82±2.0ꢀ 54H, m), 3.40±3.ꢀ0
2H, m), 3.ꢀ9±3.67 52H, m), 4.06±4.14 52H, m), 4.29
2H, s), 4.68±4.76 51H, m), 6.60 51H, d, Jˆ8.8 Hz), 7.74
1H, dd, Jˆ8.8, 2.1 Hz), 7.81 51H, s), 8.42 51H, d,
1
7
8
9
Jˆ2.1 Hz) ppm. MS 5ESI) m/z 39ꢀ 5MH , 100%).
4.1.15. )6-{2-[2-)2-S-Cyanopyrrolidin-1-ylꢀ-2-oxo-ethyl-
amino]-ethylamino}-nicotinonitrile )1ꢀ, by removal of
. Nash, I.; Bycroft, B. W.; Chang, W. C. Tetrahedron Lett. 1996,
3
tri¯uoroacetyl group. Aqueous ammonia solution 533%,
.ꢀ0 ml) was added to a solution of N-[2-5ꢀ-cyanopyridin-
-ylamino)-ethyl]-N-[2-52-S-cyanopyrrolidin-1-yl)-2-oxo-
7, 262ꢀ±2628.
0
2