Synthesis of nucleo aminooxy acid derivatives

Article abstract of DOI:10.1055/s-0032-1317689

Nucleobase-functionalized peptides have attracted increasing interest because of their well-ordered secondary structures and stability toward enzymatic degradations. We have designed and synthesized nucleo aminooxy acids as novel building blocks for nucle

Full text of DOI:10.1055/s-0032-1317689

134▌
PAPER
paper
Synthesis of Nucleo Aminooxy Acid Derivatives
Synthesis of Nucleo Aminooxy Acid Derivatives
Olivier Noel, Juan Xie*
PPSM, ENS Cachan, Institut d’Alembert, CNRS, UMR 8531, 61 av. Président Wilson, 94235 Cachan cedex, France
Fax +33(1)47402454; E-mail: joanne.xie@ens-cachan.fr
Received: 15.10.2012; Accepted after revision: 02.11.2012
H
O
N
NHCbz
Abstract: Nucleobase-functionalized peptides have attracted in-
creasing interest because of their well-ordered secondary structures
and stability toward enzymatic degradations. We have designed and
synthesized nucleo aminooxy acids as novel building blocks for nu-
cleopeptides. Four nucleo aminooxy acid derivatives with cytosine
or thymine in the side chain linked by an amide or a triazole moiety
have been synthesized from L-serine.
O
N
O
N
N
O
NH
O
NH
R1R2N
O
R1R2N
O
CO2R3
CO2R3
1a R1 = H, R2 = Boc, R3 = Me
1b R1 = R2 = Boc, R3 = Me
1c R1 = R2 = R3 = H
2a R1, R2 = Phth, R3 = Me
2b R1 = R2 = Boc, R3 = Me
2c R1 = R2 = R3 = H
Key words: nucleo aminooxy acids, nucleopeptides, N-oxy nu-
cleopeptides, cytosine, thymine, amides, triazoles, L-serine, elimi-
nations
H
N
NHCbz
O
N
O
O
N
Nucleo amino acids are synthetic amino acids bearing nu-
cleobases covalently linked to their side chains. Various
peptides containing nucleo α- and β-amino acids have
been reported as being able to form rigid and helical struc-
tures as well as well-defined double strands with comple-
mentary sequences.1–7 Moreover, nucleopeptides have
recently emerged as a promising alternative to peptide nu-
cleic acids,8,9 able to penetrate into a cell nucleus without
cytotoxic effects.10
N
N
N
N
N
N
N
O
O
NH
NH
PhthN
O
PhthN
O
CO2Me
CO2Me
3
4
Aminooxy acids are analogues of amino acids bearing an
oxyamine function (O–NH2) in the place of amine. Pep-
tides of aminooxy acids have an ease of forming well-
defined structures like α-, β- and γ-turns or helices thanks
to intramolecular hydrogen-bond formation.11,12 It would
therefore be interesting to synthesize nucleo aminooxy
acids containing nucleobases on the side chain of amino-
oxy acids in order to study the secondary structure and
DNA/RNA binding properties of the corresponding N-
oxy nucleopeptides, since both the N-oxy peptide and the
nucleobases could contribute to structure organization. As
part of a continuing program on the synthesis of sugar-
and nucleoside-derived aminooxy acids,13–17 we report
herein the synthesis of nucleo aminooxy acid derivatives
with thymine or cytosine connected to the side chain of a
β-aminooxy acid through either an amide or a triazole
linkage (Figure 1). To the best of our knowledge, nucleo
aminooxy acids have not been previously reported in the
literature.
Figure 1 Structures of the target nucleo aminooxy acid derivatives
lation and Mitsunobu reactions, and purified compounds
6 and 7 by simple precipitation, without column chroma-
tography. Treatment of 7 with 36% hydrochloric acid in
dichloromethane as reported led, however, to a mixture of
compounds. Removal of the trityl group was then
achieved with acetyl chloride in methanol, leading to the
amine salt 8 in 65% yield. Coupling of 8 with N4-Cbz-pro-
tected cytosin-1-ylacetic acid 919 using BOP reagent fur-
nished the cytosin-1-yl-substituted aminooxy ester 2a in
53% yield; however, reaction of 8 with thymin-1-ylacetic
acid (14)20 using EDC/HOAt led to the corresponding po-
lar thymin-1-yl-substituted aminooxy ester which proved
to be difficult to purify.
We then decided to replace the phthaloyl protecting group
of the oxyamine in 8 by Boc, through Cbz protection of
the amine function, hydrazinolysis and treatment with
Boc2O;18 however, saponification of 10 led to a mixture of
the desired carboxylic acid 1118 and the elimination prod-
uct 1221 in a 1:1 ratio (Scheme 1), showing that the N-Boc-
protected aminooxy ester 10 is sensitive to basic condi-
tions. Deprotection of the Cbz group under hydrogenoly-
sis conditions was also troublesome. In fact, prolonged
reaction induced homolytic cleavage of the N–O bond.15
The target nucleo aminooxy acid derivatives are accessi-
ble from the β-phthalimidooxy ester 8 (Scheme 1). This
compound has been previously prepared from L-serine by
Burke and co-workers.18 We have synthesized the
phthalimidooxy ester 7 from L-Ser–OMe (5) by N-trity-
SYNTHESIS 2013, 45, 0134–0140
Advanced online publication: 23.11.2012
0
0
3
9
-
7
8
8
1
1
4
3
7
-
2
1
0
X
DOI: 10.1055/s-0032-1317689; Art ID: SS-2012-Z0807-OP
© Georg Thieme Verlag Stuttgart · New York

PAPER
Synthesis of Nucleo Aminooxy Acid Derivatives
135
NHTr
CO2Me
NHTr
NH3+Cl–
CO2Me
NH3+Cl–
CO2Me
AcCl, MeOH
PhthNOH, Ph3P
TrCl, Et3N
HO
PhthN
O
PhthN O
HO
DIAD, PhMe,
0 °C to r.t., 18 h
58.5%
CH2Cl2, r.t., 16 h
84%
0 °C, 1 h, 65%
CO2Me
6
7
5
8
O
N
NHCbz
NHCbz
N
BOP, DIPEA, DMF
r.t., 18 h, 53%
N
8 +
O
N
O
NH
PhthN
O
CO2H
2a
9
CO2Me
NHCbz
CO2H
NHCbz
NHCbz
CO2Me
1. CbzCl, NaHCO3, r.t., 98%
LiOH, THF, H2O
0 °C
8
BocHN
O
+
BocHN
O
2. MeNHNH2, CH2Cl2, 98%
3. Boc2O, Et3N, THF, 76%
CO2H
11
12
10
O
H
N
NH
O
O
N
O
N
NH2
CO2Me
HO2C
H2, 10% Pd/C
14
BocHN
O
10
MeOH, r.t., 10 min
EDC, HOAt
O
NH
CO2Me
13
1a
CH2Cl2, r.t., 43%
BocHN
O
Boc
Cbz
N
NHCbz
Boc2O (1.2 equiv), DMAP
THF, r.t.
Boc2N
O
Boc2N
O
+
10
15
CO2Me
CO2Me
15 (70%)
16 (15%)
H
O
O
N
N
O
NHCbz
1. H2, 10% Pd/C
MeOH, r.t., 10 min
N
N
or
2. 14 or 9, EDC, HOBt,
CH2Cl2
O
O
O
NH
NH
CO2Me
Boc2N
Boc2N O
2b (65%)
CO2Me
1b (55%)
H
O
N
O
N
N
N
O
NHCbz
1. AcCl, MeOH
1. AcCl, MeOH
1b
2b
2. LiOH, THF, H2O
97%
2. LiOH, THF, H2O
100%
O
O
NH
NH
H2N
O
H2N O
CO2H
CO2H
1c
2c
Scheme 1 Synthesis of nucleo aminooxy acid derivatives 1 and 2
Nevertheless, it was possible to obtain the amine 13 in ac- in 55% and 65% yield, respectively. To prepare the fully
ceptable yield when the reaction time did not exceed 10 deprotected nucleo aminooxy acids 1c and 2c, it is prefer-
minutes. Coupling of amine 13 with thymin-1-ylacetic able to remove the Boc groups before saponification in or-
acid (14) promoted by EDC/HOAt furnished the thymin- der to avoid the elimination reaction of 1b and 2b under
1-yl-substituted aminooxy ester 1a in 43% yield (two basic conditions. Compounds 1c and 2c were obtained in
steps), along with a small quantity of over-acylation prod- quantitative yield (Scheme 1).
uct of the HN–O nitrogen.22 Boc protection of the HN–O
nitrogen in 10 was then realized with Boc2O in the pres-
ence of 4-(dimethylamino)pyridine to afford a 70% yield
of 15 and a 15% yield of the tris-Boc derivative 16. Fast
Triazole-linked nucleo aminooxy acid derivatives 3 and 4
could be prepared by click reaction between the azido in-
termediate 19 and the alkyne derivatives 22 and 2123
(Scheme 2). Compound 8 was firstly acylated with chlo-
hydrogenolysis of 15 followed by coupling with 14 or 9
roacetyl chloride to give compound 17; however, subse-
gave the corresponding nucleo aminooxy esters 1b and 2b
© Georg Thieme Verlag Stuttgart · New York
Synthesis 2013, 45, 134–140

136
O. Noel, J. Xie
PAPER
N3
Cl
O
NH3+Cl–
CO2Me
ClCH2COCl, Et3N
THF, r.t., 53%
NaN3, DMF
HN
O
NH
PhthN
O
PhthN
O
0 °C, 150 min, 100%
CO2Me
8
CO2Me
17
18
N3
EDC, HOBt, base
CH2Cl2, r.t.
O
O
NH
+
N3CH2CO2H + 8
18
PhthN
Et3N
DTBMP
CO2Me
19
base:
24%
95%
>50%
0%
NHCbz
NHCbz
N
Br
N
NaH, MeCN
reflux, 18 h
30%
O
N
O
N
H
20
21
H
O
N
O
N
O
NH
N
O
N
22
N
N
19
CuSO4, sodium ascorbate
DMF, r.t., overnight, 68%
O
NH
23
CO2Me
H
N
O
NHCbz
N
O
N
O
N
N
22 or 21, CuSO4, ascorbic acid
or
19
N
N
DMF, r.t., overnight
N
N
N
O
O
NH
NH
PhthN
O
PhthN
O
CO2Me
CO2Me
3 (87%)
4 (77%)
Scheme 2 Synthesis of nucleo aminooxy esters 3 and 4; DTBMP = 2,6-di(tert-butyl)-4-methylpyridine
quent substitution with sodium azide at 0 °C leading to compound 19 in 95% yield. Click reaction of 19
quantitatively afforded the elimination product 18. We with 1-propargylthymine (22) catalyzed by copper(II) sul-
then decided to prepare 19 by condensation of amine salt fate and sodium ascorbate accomplished the cycloaddi-
8 with azidoacetic acid24 promoted by EDC/HOBt in the tion reaction, followed, however, by elimination of the
presence of 1 equivalent of triethylamine. Once again, the phthalimidooxy moiety to give compound 23 in 68%
elimination reaction mainly occurred: the desired com- yield. Fortunately, the use of ascorbic acid25 avoided the
pound 19 was isolated in only 24% yield. To avoid this elimination reaction and afforded the desired nucleo ami-
side reaction, hindered 2,6-di-tert-butyl-4-methylpyridine nooxy acid derivatives 3 and 4 in 87% and 77% yield, re-
was chosen to neutralize the amine salt 8, successfully spectively.
Synthesis 2013, 45, 134–140
© Georg Thieme Verlag Stuttgart · New York

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Synthesis of Nucleo Aminooxy Acid Derivatives
137
[α]D27 +2.6 (c 0.5, MeOH).
1H NMR (CD3OD): δ = 7.89–7.86 (m, 4 H, H-Phth), 4.65–4.63 (m,
2 H, CH2), 4.61–4.58 (m, 1 H, CH), 3.88 (s, 3 H, OCH3).
13C NMR (CD3OD): δ = 170.6, 167.2 (CO), 138.9 (CH-Phth), 132.7
In summary, two series of nucleo aminooxy acid deriva-
tives have been synthesized by linking thymine or cyto-
sine to the side chain of an α-amino-β-aminooxy acid
prepared from L-serine methyl ester. A more convenient
procedure for the synthesis of phthaloyl-protected β-ami- (Cq), 127.3 (CH-Phth), 78.3 (CH2), 56.5 (OCH3), 55.3 (CH).
HRMS (ESI): m/z [M + H]+ calcd for C12H13N2O5: 265.0824;
found: 265.0820.
nooxy ester 8 has been developed. The high polarity of
phthaloyl-protected amide-linked nucleo β-aminooxy es-
ters led us to prepare Boc-protected derivatives 1a, 1b and
2b which were fully deprotected to the free nucleo β-ami-
nooxy acids 1c and 2c. Triazole-linked nucleo aminooxy
esters 3 and 4 have also been successfully synthesized via
click chemistry. During our synthesis, we also observed
the instability of phthaloyl- or Boc-protected β-aminooxy
esters under basic conditions, leading to the correspond-
ing acrylate elimination products. This side reaction could
be avoided by the use of a hindered base or nonbasic con-
ditions. These newly synthesized nucleo aminooxy acid
derivatives might constitute useful building blocks for the
synthesis of N-oxy nucleopeptides for investigation of
their secondary structure and DNA/RNA binding proper-
ties.
Methyl (S)-2-[2-(4-{[(Benzyloxy)carbonyl]amino}-2-oxopyrimi-
din-1(2H)-yl)acetamido]-3-[(1,3-dioxoisoindolin-2-yl)oxy]pro-
panoate (2a)
To a soln of {N4-[(benzyloxy)carbonyl]cytosin-1-yl}acetic acid (9;
1.31 g, 4.33 mmol) in DMF (20 mL) were added DIPEA (1.1 mL,
6.66 mmol) and BOP reagent (1.91 g, 4.33 mmol) at r.t. After 10
min, compound 8 (1.00 g, 3.33 mmol) was added. The reaction mix-
ture was stirred for 18 h, then concentrated and dissolved in EtOAc
(100 mL). The organic layer was washed with sat. aq NH4Cl (1 × 30
mL), sat. aq NaHCO3 (2 × 30 mL) and brine (1 × 30 mL), then dried
over MgSO4 and concentrated under reduced pressure. The crude
product was purified by column chromatography (CH2Cl2 to
CH2Cl2–MeOH, 96:4) to give 2a as white crystals; yield: 863 mg
(53%); mp 118 °C.
[α]D27 +13.3 (c 0.5, CHCl3); Rf = 0.40 (CH2Cl2–MeOH, 95:5).
1H NMR (CDCl3): δ = 8.01 (d, J = 7.8 Hz, 1 H, NH), 7.85–7.79 (m,
2 H, H-Phth), 7.75–7.72 (m, 2 H, H-Phth), 7.69 (d, J = 7.3 Hz, 1 H,
H-Ar), 7.37 (m, 5 H, CH), 7.26 (d, J = 7.3 Hz, 1 H, CH), 5.20 (s, 2
H, OCH2), 4.88–4.86 (m, 1 H, CH), 4.85 (dd, J = 3.2, 7.3 Hz, 1 H,
OCH2), 4.73 (s, 2 H, NCH2), 4.38 (dd, J = 3.2, 7.3 Hz, 1 H, OCH2),
3.68 (s, 3 H, OCH3).
13C NMR (CDCl3): δ = 169.1, 166.8, 163.5, 163.0, 156.0, 149.3
(Cq), 149.5, 135.2 (CH), 134.9 (Cq), 128.8, 128.7, 128.6 (CH),
128.4 (Cq), 123.9, 95.8 (CH), 77.7, 68.0 (OCH2), 53.6 (NCH2), 53.1
(OCH3), 52.2 (CH).
Commercially available solvents and reagents were used without
further purification, except DMF which was distilled over CaH2.
Melting points were measured on a Kofler bench. Optical rotations
were measured using a JASCO P-2000 polarimeter. Column chro-
matography was performed on Carlo Erba silica gel 60A (40–63
μm). Analytical thin-layer chromatography was performed on E.
Merck aluminum precoated plates of silica gel 60F-254 with detec-
tion by UV light and by spraying with 10% H2SO4 in EtOH or nin-
hydrin soln (3 g·L–1) and heating for about 20 seconds at 400 °C. 1H
and 13C NMR spectra were recorded on a Jeol ECS-400 spectrome-
ter. ESI-HRMS data were recorded on a Bruker micrOTOF-Q II or
a Bruker maXis spectrometer using standard conditions.
HRMS (ESI): m/z [M + Na]+ calcd for C26H23N5NaO9: 572.1393;
found: 572.1387.
Methyl (S)-3-{[(tert-Butoxycarbonyl)amino]oxy}-2-{2-[5-meth-
yl-2,4-dioxo-3,4-dihydropyrimidin-1(2H)-yl]acetamido}pro-
panoate (1a)
N-Trityl-L-serine Methyl Ester (6)18
To a soln of L-Ser–OMe·HCl (5; 9.29 g, 59.9 mmol) in CH2Cl2 (250
mL) were added Et3N (23 mL, 163.7 mmol) and TrCl (19.98 g, 71.9
mmol). The resulting mixture was stirred for 16 h at r.t. and then
concentrated under reduced pressure. The crude material was tritu-
rated in EtOAc and compound 6 was isolated by precipitation as a
white solid; yield: 18.64 g (84%); mp 138 °C.
To a soln of methyl (S)-2-{[(benzyloxy)carbonyl]amino}-3-{[(tert-
butoxycarbonyl)amino]oxy}propanoate18 (10; 204 mg, 0.55 mmol)
in MeOH (10 mL) was added 10% Pd/C (30 mg). H2 was bubbled
into the mixture for 10 min. The mixture was filtered and the filtrate
was concentrated under reduced pressure. The resultant oily residue
in CH2Cl2 (4 mL) was added to a mixture of 2-[5-methyl-2,4-dioxo-
3,4-dihydropyrimidin-1(2H)-yl]acetic acid (14; 131 mg, 0.72
mmol), EDC (136 mg, 0.72 mmol) and HOAt (97 mg, 071 mmol)
in CH2Cl2 (26 mL) at r.t. After 18 h of stirring, the mixture was
washed with sat. aq NaHCO3 (2 × 10 mL) and brine (1 × 10 mL).
The organic layer was dried over MgSO4 and concentrated. The res-
idue was purified by column chromatography (CH2Cl2–MeOH,
96:4) to obtain 1a as a white foam; yield: 94 mg (43%); mp 110 °C.
Rf = 0.5 (EtOAc–PE, 1:1).
O-Phthalimido-N-trityl-L-serine Methyl Ester (7)18
To a soln of 6 (10.02 g, 27.76 mmol) in toluene (150 mL) were add-
ed N-hydroxyphthalimide (6.33 g, 38.86 mmol) and Ph3P (10.18 g,
38.86 mmol). At 0 °C, DIAD (7.65 mL, 41.64 mmol) was then add-
ed dropwise. The resulting mixture was stirred for 18 h at r.t. and
then washed with 1 N NaOH (2 × 50 mL) and brine (50 mL), dried
over MgSO4 and concentrated under reduced pressure. To the or-
ange crude material dissolved in Et2O (100 mL), ice (100 g) was
added. After 2 h of vigorous stirring, the precipitate was collected
by filtration to obtain 7 as a white solid; yield: 8.23 g (58.5%); mp
113 °C.
[α]D27 +2.0 (c 0.5, CHCl3); Rf = 0.40 (CH2Cl2–MeOH, 9:1).
1H NMR (CDCl3): δ = 9.57 (s, 1 H, NH), 8.07 (d, J = 8.2 Hz, 1 H,
NH), 7.85 (s, 1 H, NH), 7.11 (s, 1 H, CH), 4.81–4.78 (m, 1 H, CH),
4.50 (s, 2 H, NCH2), 4.30 (dd, J = 3.7, 11.0 Hz, 1 H, OCH2), 4.02
(dd, J = 3.7, 11.0 Hz, 1 H, OCH2), 3.71 (s, 3 H, OCH3), 1.91 (s, 3 H,
CH3), 1.45 (s, 9 H, t-Bu).
Rf = 0.74 (EtOAc–PE, 1:1).
13C NMR (CDCl3): δ = 170.3, 167.2, 164.5, 157.5, 151.5 (CO),
141.0 (CH-Ar), 111.2 (Cq), 82.6 (Cq-t-Bu), 75.6 (OCH2), 53.0
(OCH3), 51.7 (CH), 50.2 (NCH2), 28.2 (t-Bu), 12.5 (CH3).
HRMS (ESI): m/z [M + Na]+ calcd for C16H24N4NaO8: 423.1492;
found: 423.1488.
O-Phthalimido-L-serine Methyl Ester Hydrochloride (8)
To a soln of 7 (11.12 g, 21.98 mmol) in MeOH (300 mL) was added
AcCl (1.73 mL, 24.18 mmol) at 0 °C. After 1 h of stirring, MeOH
was evaporated under reduced pressure to give a white crude mate-
rial which precipitated in CH2Cl2 to give 8 as a white solid; yield:
4.28 g (65%); mp 154 °C.
© Georg Thieme Verlag Stuttgart · New York
Synthesis 2013, 45, 134–140

138
O. Noel, J. Xie
PAPER
Methyl (S)-2-{[(Benzyloxy)carbonyl]amino}-3-{[bis(tert-bu-
toxycarbonyl)amino]oxy}propanoate (15)
CH2Cl2 (10 mL) was added to a mixture of 9 (623 mg, 2.06 mmol),
EDC (600 mg, 2.02 mmol) and HOBt (278 mg, 2.06 mmol) in
CH2Cl2 (70 mL) at r.t. After 18 h of stirring, the mixture was washed
with sat. aq NaHCO3 (2 × 50 mL) and brine (1 × 50 mL). The or-
ganic layer was dried over MgSO4 and concentrated. The residue
was purified by column chromatography (EtOAc–PE, 4:6 to 6:4) to
obtain 2b as a white solid; yield: 587 mg (65%); mp 97 °C.
To a soln of 10 (106 mg, 0.29 mmol) in THF (3 mL) were added a
soln of Boc2O (75 mg, 0.35 mmol) in THF (3 mL) and DMAP (70
mg, 0.58 mmol) at r.t. After 17 h of stirring, the reaction mixture
was concentrated. The crude product was purified by column chro-
matography (EtOAc–PE, 1:9 to 2:8) to give compound 15 as a
colorless oil [yield: 94 mg (70%)] and methyl (S)-2-{[(benzyl-
oxy)carbonyl](tert-butoxycarbonyl)amino}-3-{[bis(tert-butoxycarb-
onyl)amino]oxy}propanoate (16) as a colorless oil [yield: 25 mg
(15%)].
[α]D27 +4.1 (c 0.5, CHCl3); Rf = 0.33 (EtOAc).
1H NMR (CDCl3): δ = 8.16 (s, 1 H, NH), 7.94 (d, J = 7.3 Hz, 1 H,
NH), 7.67 (d, J = 7.4 Hz, 1 H, CH), 7.32–7.28 (m, 5 H, H-Ph), 7.20
(d, J = 7.4 Hz, 1 H, CH), 5.14 (s, 2 H, OCH2), 4.68–4.65 (m, 1 H,
CH), 4.63 (s, 2 H, NCH2), 4.52 (dd, J = 3.2, 10.1 Hz, 1 H, OCH2),
3.98 (dd, J = 3.7, 10.1 Hz, 1 H, OCH2), 3.62 (s, 3 H, OCH3), 1.40
(s, 18 H, 2 × t-Bu).
13C NMR (CDCl3): δ = 169.3, 168.0, 166.8, 162.9, 155.9, 152.4,
150.2 (Cq), 149.6 (CH), 135.2 (Cq), 128.7, 128.6, 128.3, 95.3 (CH),
84.8 (Cq-t-Bu), 75.4, 67.9 (OCH2), 52.9 (OCH3), 52.0 (CH), 50.8
(NCH2), 28.0 (t-Bu).
Compound 15
[α]D27 –2.0 (c 0.5, CHCl3); Rf = 0.61 (EtOAc–PE, 3:7).
1H NMR (CDCl3): δ = 7.36–7.33 (m, 5 H, H-Ph), 6.16 (d, J = 8.2
Hz, 1 H, NH), 5.14 (s, 2 H, OCH2), 4.57–4.54 (m, 1 H, OCH2),
4.57–4.54 (m, 1 H, CH), 4.02 (dd, J = 3.2, 9.2 Hz, 1 H, OCH2), 3.78
(s, 3 H, OCH3), 1.52 (s, 18 H, 2 × t-Bu).
13C NMR (CDCl3): δ = 169.9, 156.2, 149.8 (CO), 136.4 (Cq-Ph),
128.5, 128.2, 128.1 (CH-Ph), 84.6 (Cq-t-Bu), 75.6, 67.1 (CH2), 53.3
(CH), 52.8 (OCH3), 28.1 (t-Bu).
HRMS (ESI): m/z [M + H]+ calcd for C28H38N5O11: 620.2568;
found: 620.2561.
HRMS (ESI): m/z [M + Na]+ calcd for C22H32N2NaO9: 491.2006;
found: 491.2015.
(S)-3-(Aminooxy)-2-{2-[5-methyl-2,4-dioxo-3,4-dihydropyrimi-
din-1(2H)-yl]acetamido}propanoic Acid (1c)
Compound 16
[α]D27 –16.4 (c 0.5, CHCl3); Rf = 0.68 (EtOAc–PE, 3:7).
To a soln of 1b (46 mg, 0.092 mmol) in MeOH (2 mL) was added
AcCl (200 μL, 2.88 mmol) at 0 °C. After 4 h of stirring, the mixture
was concentrated under reduced pressure. The residue was dis-
solved in THF (1.5 mL) and H2O (1.5 mL). LiOH (6.6 mg, 0.28
mmol) was added to the solution at r.t. After an overnight stirring,
the mixture was neutralized with H+ resin (Dowex) and concentrat-
ed under reduced pressure to give 1c as a white solid; yield: 26 mg
(100%); mp 124 °C.
1H NMR (CDCl3): δ = 7.35–7.30 (m, 5 H, H-Ph), 5.37–5.34 (m, 1
H, CH), 5.21 (s, 2 H, OCH2), 4.62–4.59 (m, 1 H, OCH2), 4.25–4.23
(m, 1 H, OCH2), 3.64 (s, 3 H, OCH3), 1.46 (s, 18 H, 2 × t-Bu), 1.41
(s, 9 H, t-Bu).
13C NMR (CDCl3): δ = 168.5 (CO), 153.5, 151.1, 149.9, 135.2 (Cq),
128.6, 128.5, 128.4 (CH-Ph), 84.1, 83.8 (Cq-t-Bu), 75.1, 69.0
(OCH2), 57.5 (CH), 52.6 (OCH3), 28.2, 28.1 (t-Bu).
[α]D27 +3.4 (c 0.5, MeOH).
IR (KBr): 3481.6, 3329.1, 2976.3, 1724.2, 1670.0, 1625.5, 1618.3,
1557.3 cm–1.
1H NMR (DMSO-d6): δ = 11.29 (s, 1 H, OH), 8.80 (d, J = 7.3 Hz, 1
H, NH), 7.43 (s, 1 H, CH), 4.77–4.70 (m, 1 H, CH), 4.51 (m, 1 H,
OCH2), 4.38–4.28 (m, 2 H, NCH2), 3.91–3.86 (m, 1 H, OCH2), 1.73
(s, 3 H, CH3).
HRMS (ESI): m/z [M + Na]+ calcd for C27H40N2NaO11: 591.2530;
found: 591.2520.
Methyl (S)-3-{[Bis(tert-butoxycarbonyl)amino]oxy}-2-{2-[5-
methyl-2,4-dioxo-3,4-dihydropyrimidin-1(2H)-yl]acetami-
do}propanoate (1b)
To a soln of 15 (100 mg, 0.21 mmol) in MeOH (2 mL) was added
10% Pd/C (40 mg). H2 was bubbled into the mixture for 10 min. The
mixture was filtered and the filtrate was concentrated under reduced
pressure. The resultant oily residue in CH2Cl2 (2 mL) was added to
a mixture of 14 (55 mg, 0.29 mmol), EDC (57 mg, 0.29 mmol) and
HOBt (41 mg, 0.29 mmol) in CH2Cl2 (8 mL) at r.t. After 18 h of stir-
ring, the mixture was washed with sat. aq NaHCO3 (2 × 10 mL) and
brine (1 × 10 mL). The organic layer was dried over MgSO4 and
concentrated. The residue was purified by column chromatography
(EtOAc–PE, 4:6 to 6:4) to obtain 1b as a white foam; yield: 59 mg
(55%); mp 100 °C.
13C NMR (DMSO-d6): δ = 168.0, 165.0, 151.5 (Cq), 142.8 (CH),
108.5 (Cq), 73.0 (OCH2), 51.8 (CH), 49.6 (NCH2), 12.5 (CH3).
HRMS (ESI): m/z [M – H2O + Na]+ calcd for C10H12N4NaO5:
291.0705; found: 291.0701.
(S)-3-(Aminooxy)-2-{2-[4-{[(benzyloxy)carbonyl]amino}-2-
oxopyrimidin-1(2H)-yl]acetamido}propanoic Acid (2c)
To a soln of 2b (49 mg, 0.079 mmol) in MeOH (2 mL) was added
AcCl (200 μL, 2.88 mmol) at 0 °C. After 4 h of stirring, the mixture
was concentrated under reduced pressure. The residue was dis-
solved in THF (1.5 mL) and H2O (1.5 mL). LiOH (6.6 mg, 0.28
mmol) was added to the solution at r.t. After an overnight stirring,
the mixture was neutralized with H+ resin (Dowex) and concentrat-
ed under reduced pressure to give 2c as a white solid; yield: 31 mg
(97%); mp 113 °C.
[α]D27 +11.0 (c 0.5, CHCl3); Rf = 0.43 (CH2Cl2–MeOH, 9:1).
1H NMR (CDCl3): δ = 9.00 (s, 1 H, NH), 7.86 (d, J = 7.8 Hz, 1 H,
NH), 7.05 (s, 1 H, CH), 4.70–4.67 (m, 1 H, CH), 4.61 (dd, J = 2.8,
9.6 Hz, 1 H, OCH2), 4.48 (s, 2 H, NCH2), 3.97 (dd, J = 3.7, 10.1 Hz,
1 H, OCH2), 3.73 (s, 3 H, OCH3), 1.88 (d, J = 0.9 Hz, 3 H, CH3),
1.49 (s, 18 H, 2 × t-Bu).
13C NMR (CDCl3): δ = 169.2, 166.8, 164.2, 151.0, 150.4 (Cq),
140.6 (CH-Ar), 111.2, 85.1 (Cq), 75.6 (OCH2), 53.0 (OCH3), 51.9
(CH), 49.8 (NCH2), 28.1 (t-Bu), 12.4 (CH3).
[α]D27 –6.8 (c 0.5, DMSO).
IR (KBr): 3403.3, 3319.1, 1734.4, 1709.2, 1666.0, 1644.7, 1606.9
cm–1.
1H NMR (DMSO-d6): δ = 11.28 (s, 1 H, OH), 9.82 (s, 1 H, NH),
8.88 (d, J = 7.8 Hz, 1 H, NH), 7.96 (d, J = 7.8 Hz, 1 H, CH), 7.38–
7.30 (m, 5 H, H-Ph), 6.95 (d, J = 7.8 Hz, 1 H, CH), 5.14 (s, 2 H,
HRMS (ESI): m/z [M + Na]+ calcd for C21H32N4NaO10: 523.2016;
found: 523.2007.
OCH2), 4.74–4.68 (m,
1 H, CH), 4.55–4.44 (m, 3 H,
NCH2 + OCH2), 3.90–3.86 (m, 1 H, OCH2).
Methyl (S)-2-{2-[4-{[(Benzyloxy)carbonyl]amino}-2-oxopyrim-
idin-1(2H)-yl]acetamido}-3-{[bis(tert-butoxycarbonyl)ami-
no]oxy}propanoate (2b)
Compound 15 (676 mg, 1.44 mmol) was hydrogenolyzed in the
presence of 10% Pd/C (270 mg). The resultant oily residue in
13C NMR (DMSO-d6): δ = 167.8, 163.7, 160.5, 155.5, 153.7 (Cq),
151.5 (CH), 136.5 (Cq), 129.0, 128.7, 128.5, 92.3 (CH), 73.0, 67.0
(OCH2), 51.9 (CH), 51.6 (NCH2).
Synthesis 2013, 45, 134–140
© Georg Thieme Verlag Stuttgart · New York

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Synthesis of Nucleo Aminooxy Acid Derivatives
139
HRMS (ESI): m/z [M – H2O + H]+ calcd for C17H18N5O6:
388.1257; found: 388.1250.
the reaction mixture was washed with brine (2 × 15 mL). The or-
ganic layer was dried over MgSO4 and concentrated. The residue
was purified by column chromatography (EtOAc–PE, 5:5 to 8:2) to
give 21 as a pale yellow solid; yield: 192 mg (30%); mp 154 °C.
Methyl (S)-2-(2-Chloroacetamido)-3-[(1,3-dioxoisoindolin-2-
yl)oxy]propanoate (17)
Rf = 0.60 (EtOAc).
To a soln of 8 (209 mg, 0.70 mmol) in THF (10 mL) were added
Et3N (196 μL, 1.40 mmol) and chloroacetyl chloride (83 μL, 1.05
mmol) at 0 °C. After 17 h of stirring at r.t., the reaction mixture was
washed with brine (2 × 10 mL). The aqueous layer was extracted
with EtOAc (2 × 10 mL). The combined organic layers were dried
over MgSO4 and concentrated. The crude product was purified by
column chromatography (EtOAc–PE, 2:8 to 6:4) to give 17 as a
white solid; yield: 126 mg (53%); mp 138 °C.
1H NMR (CD3OD): δ = 8.10 (d, J = 7.3 Hz, 1 H, =CH), 7.38–7.30
(m, 6 H, H-Ar), 5.19 (s, 2 H, OCH2), 4.67 (s, 2 H, NCH2), 2.96 (s, 1
H, ≡CH).
13C NMR (CD3OD): δ = 163.9, 156.4, 153.2 (Cq), 147.9 (CH),
135.8 (Cq), 128.3, 128.1, 128.0, 95.9 (CH), 75.1 (≡CH), 70.3 (≡Cq),
67.2 (OCH2), 38.6 (NCH2).
[α]D27 +62.5 (c 0.5, CHCl3); Rf = 0.37 (EtOAc–PE, 1:1).
Methyl 2-[2-(4-{[5-Methyl-2,4-dioxo-3,4-dihydropyrimidin-
1(2H)-yl]methyl}-1H-1,2,3-triazol-1-yl)acetamido]acrylate (23)
To a soln of 19 (230 mg, 0.66 mmol) in DMF (3 mL) were added 5-
methyl-1-(prop-2-yn-1-yl)pyrimidine-2,4(1H,3H)-dione (22; 109
mg, 0.66 mmol), sodium ascorbate (66 mg, 0.33 mmol) and
CuSO4·5H2O (41 mg, 0.17 mmol) at r.t. After an overnight stirring,
EtOAc (50 mL) was added to the mixture. The solution was washed
with sat. aq NaHCO3 (1 × 30 mL) and brine (2 × 30 mL). The or-
ganic layer was dried over MgSO4 and concentrated under reduced
pressure to give 23 as a white solid; yield: 157 mg (68%); mp
150 °C.
1H NMR (CDCl3): δ = 7.98 (d, J = 5.0 Hz, 1 H, NH), 7.88–7.83 (m,
2 H, H-Phth), 7.80–7.77 (m, 2 H, H-Phth), 4.89–4.87 (m, 1 H, CH),
4.89–4.87 (m, 1 H, OCH2), 4.42 (dd, J = 4.6, 11.9 Hz, 1 H, OCH2),
4.18 (s, 2 H, CH2Cl), 3.74 (s, 3 H, OCH3).
13C NMR (CDCl3): δ = 168.9, 166.6, 163.4 (CO), 135.0 (CH-Phth),
128.7 (Cq), 123.9 (CH-Phth), 77.1 (OCH2), 53.2 (OCH3), 52.0
(CH), 42.5 (CH2Cl).
HRMS (ESI): m/z [M + H]+ calcd for C14H14ClN2O6: 341.0540;
found: 341.0535.
Rf = 0.66 (CH2Cl2–MeOH, 9:1).
Methyl 2-(2-Azidoacetamido)acrylate (18)
1H NMR (DMSO-d6): δ = 11.29 (s, 1 H, NH), 9.88 (s, 1 H, NH),
8.01 (s, 1 H, H-triazole), 7.60 (s, 1 H, =CH), 6.22 (s, 1 H, =CH),
5.75 (s, 1 H, =CH), 5.31 (s, 2 H, CH2N), 4.87 (s, 2 H, CH2N), 3.73
(s, 3 H, OCH3), 1.71 (s, 3 H, CH3).
13C NMR (DMSO-d6): δ = 165.9, 164.8, 164.0, 151.2, 142.9 (Cq),
141.7 (CH-Ar), 132.6 (Cq), 125.8, 110.9 (CH), 109.4 (Cq), 53.3
(OCH3), 52.5, 42.7 (NCH2), 12.5 (CH3).
To a soln of 17 (211 mg, 0.62 mmol) in DMF (3 mL) was added
NaN3 (61 mg, 0.93 mmol) at 0 °C. After 150 min at 0 °C, EtOAc (20
mL) was added to the mixture and the resultant solution was washed
with brine (2 × 15 mL). The aqueous layer was extracted with
EtOAc (1 × 20 mL). The combined organic layers were dried over
MgSO4 and concentrated. The residue was purified by column chro-
matography (EtOAc–PE, 2:8 to 3:7) to give 18 as a colorless oil;
yield: 155 mg (100%).
HRMS (ESI): m/z [M + H]+ calcd for C14H17N6O5: 349.1260;
found: 349.1255.
Rf = 0.47 (EtOAc–PE, 3:7).
1H NMR (CDCl3): δ = 8.47 (s, 1 H, NH), 6.54 (s, 1 H, =CH), 5.87
(s, 1 H, =CH), 4.04 (s, 2 H, CH2N3), 3.78 (s, 3 H, OCH3).
13C NMR (CDCl3): δ = 165.5, 164.1 (CO), 130.5 (Cq), 109.8
Methyl (S)-3-[(1,3-Dioxoisoindolin-2-yl)oxy]-2-[2-(4-{[5-meth-
yl-2,4-dioxo-3,4-dihydropyrimidin-1(2H)-yl]methyl}-1H-1,2,3-
triazol-1-yl)acetamido]propanoate (3)
To a soln of 19 (200 mg, 0.58 mmol) in DMF (4 mL) were added 22
(95 mg, 0.58 mmol), ascorbic acid (51 mg, 0.29 mmol) and
CuSO4·5H2O (36 mg, 0.14 mmol) at r.t. After 18 h of stirring,
EtOAc (40 mL) was added to the solution. The precipitate formed
was filtered and the filtrate was washed with brine (2 × 20 mL). The
organic layer was dried over MgSO4 and concentrated under re-
duced pressure to give 3 as a white solid; yield: 255 mg (87%); mp
156 °C.
(=CH2), 53.1 (OCH3), 52.9 (CH2N3).
Methyl (S)-2-(2-Azidoacetamido)-3-[(1,3-dioxoisoindolin-2-
yl)oxy]propanoate (19)
To a soln of 2-azidoacetic acid (111 mg, 1.0 mmol) in CH2Cl2 (5
mL) were added EDC (83 mg, 0.43 mmol), HOBt (58 mg, 0.43
mmol) and 2,6-di-tert-butyl-4-methylpyridine (68 mg, 0.33 mmol)
at r.t. After 10 min, compound 8 (100 mg, 0.33 mmol) was added.
After 17 h of stirring, the reaction mixture was washed with sat. aq
NaHCO3 (1 × 10 mL) and brine (1 × 10 mL). The organic layer was
dried over MgSO4 and concentrated. The resulting residue was pu-
rified by column chromatography (EtOAc–PE, 5:5 to 10:0) to give
19 as a white solid; yield: 110 mg (95%); mp 158 °C.
[α]D27 +3.9 (c 0.5, DMSO); Rf = 0.14 (EtOAc).
1H NMR (DMSO-d6): δ = 11.31 (s, 1 H, NH), 9.08 (d, J = 7.8 Hz, 1
H, NH), 8.03 (s, 1 H, H-triazole), 7.85–7.79 (m, 4 H, H-Phth), 7.60
(d, J = 1.4 Hz, 1 H, CH), 5.23 (s, 2 H, CH2N), 4.91 (s, 2 H, CH2N),
4.79–4.77 (m, 1 H, CH), 4.51–4.42 (m, 2 H, OCH2), 3.68 (s, 3 H,
OCH3), 1.74 (s, 3 H, CH3).
13C NMR (DMSO-d6): δ = 169.6, 166.4, 164.8, 163.4, 162.8, 151.3,
142.8 (Cq), 141.7, 135.4 (CH), 129.1 (Cq), 125.6, 123.9 (CH),
109.4 (Cq), 76.8 (OCH2), 53.0 (OCH3), 52.2 (CH), 51.9, 42.6
(NCH2), 12.5 (CH3).
[α]D27 +45.9 (c 0.5, CHCl3); Rf = 0.38 (EtOAc–PE, 1:1).
1H NMR (CDCl3): δ = 7.85–7.83 (m, 2 H, H-Phth), 7.79–7.75 (m, 2
H, H-Phth), 4.90–4.88 (m, 2 H, OCH2), 4.39–4.36 (m, 1 H, CH),
4.10 (s, 2 H, CH2N3), 3.73 (s, 3 H, OCH3).
13C NMR (CDCl3): δ = 169.0, 167.3, 163.4 (CO), 135.0 (CH-Phth),
128.6 (Cq), 123.9 (CH-Phth), 77.3 (OCH2), 53.1 (OCH3), 52.7
(CH2N3), 51.6 (CH).
HRMS (ESI): m/z [M + H]+ calcd for C22H22N7O8: 512.1530;
found: 512.1522.
HRMS (ESI): m/z [M + H]+ calcd for C14H14N5O6: 348.0944;
found: 348.0939.
Methyl (S)-2-(2-{4-[(4-{[(Benzyloxy)carbonyl]amino}-2-oxopy-
rimidin-1(2H)-yl)methyl]-1H-1,2,3-triazol-1-yl}acetamido)-3-
[(1,3-dioxoisoindolin-2-yl)oxy]propanoate (4)
To a soln of 19 (50 mg, 0.14 mmol) in DMF (1 mL) were added 21
(41 mg, 0.14 mmol), ascorbic acid (13 mg, 0.07 mmol) and
CuSO4·5H2O (9 mg, 0.035 mmol) at r.t. After 18 h of stirring,
EtOAc (15 mL) was added to the solution. The precipitate formed
was filtered and the filtrate was washed with brine (2 × 10 mL). The
Benzyl [2-Oxo-1-(prop-2-yn-1-yl)-1,2-dihydropyrimidin-4-
yl]carbamate (21)
To a soln of benzyl (2-oxo-1,2-dihydropyrimidin-4-yl)carbamate26
(20; 556 mg, 2.27 mmol) in MeCN (25 mL) was added 60% NaH
(108 mg, 4.53 mmol) at 0 °C. Propargyl bromide (80% in toluene;
401 μL, 2.72 mmol) was then added. After 18 h of stirring at reflux,
© Georg Thieme Verlag Stuttgart · New York
Synthesis 2013, 45, 134–140

140
O. Noel, J. Xie
PAPER
organic layer was dried over MgSO4 and concentrated under re-
duced pressure to give 4 as a white solid; yield: 70 mg (77%); mp
126 °C.
[α]D27 +0.9 (c 0.5, CHCl3); Rf = 0.15 (EtOAc).
1H NMR (CDCl3): δ = 8.18 (s, 1 H, H-triazole), 7.99 (d, J = 7.8 Hz,
1 H, NH), 7.89 (d, J = 8.0 Hz, 1 H, CH), 7.73–7.70 (m, 2 H, H-
Phth), 7.68–7.65 (m, 2 H, H-Phth), 7.31–7.28 (m, 5 H, H-Ph), 7.13
(d, J = 8.0 Hz, 1 H, CH), 5.32 (s, 2 H, CH2N), 5.12 (s, 2 H, CH2N),
5.10 (s, 2 H, OCH2), 4.86–4.84 (m, 1 H, CH), 4.73–4.71 (m, 1 H,
NOCH2), 4.39–4.36 (m, 1 H, NOCH2), 3.62 (s, 3 H, OCH3).
13C NMR (CDCl3): δ = 169.1, 165.9, 163.3, 155.9, 152.5 (Cq),
148.8 (CH), 142.0 (Cq), 134.9, 128.7 (CH), 128.6 (Cq), 128.5,
128.3, 126.3, 123.9, 95.5 (CH), 77.3, 67.8 (OCH2), 53.1 (CH), 52.5
(CH2N), 51.9 (OCH3), 45.3 (NCH2).
(8) Garner, P.; Dey, S.; Huang, Y. J. Am. Chem. Soc. 2000, 122,
2405.
(9) Huang, Y.; Dey, S.; Zhang, X.; Sönnichsen, F.; Garner, P.
J. Am. Chem. Soc. 2004, 126, 4626.
(10) Geotti-Bianchini, P.; Beyrath, J.; Chaloin, O.; Formaggio,
F.; Bianco, A. Org. Biomol. Chem. 2008, 6, 3661.
(11) Yang, D.; Qu, J.; Li, B.; Ng, F.-F.; Wang, X.-C.; Cheung, K.-
K.; Wang, D.-P.; Wu, Y.-D. J. Am. Chem. Soc. 1999, 121,
589.
(12) Li, X.; Wu, Y.-D.; Yang, D. Acc. Chem. Res. 2008, 41, 1428.
(13) Malapelle, A.; Ramozzi, R.; Xie, J. Synthesis 2009, 888.
(14) Gong, Y. C.; Sun, H. B.; Xie, J. Eur. J. Org. Chem. 2009,
6027.
(15) Gong, Y.; Peyrat, S.; Sun, H.; Xie, J. Tetrahedron 2011, 67,
7114.
HRMS (ESI): m/z [M + H]+ calcd for C29H27N8O9: 631.1901;
found: 631.1891.
(16) Song, Z.; He, X.-P.; Chen, G.-R.; Xie, J. Synthesis 2011,
2761.
(17) Peyrat, S.; Xie, J. Synthesis 2012, 44, 1718.
(18) Liu, F.; Thomas, J.; Burke, T. R. Jr Synthesis 2008, 2432.
(19) Katritzky, A. R.; Narindoshvili, T. Org. Biomol. Chem.
2008, 6, 3171.
(20) Schwergold, C.; Depecker, G.; Di Giorgio, C.; Patino, N.;
Jossinet, F.; Ehresmann, B.; Terreux, R.; Cabrol-Bass, D.;
Condom, R. Tetrahedron 2002, 58, 5675.
(21) Srinivasan, A.; Stephenson, R. W.; Olsen, R. K. J. Org.
Chem. 1977, 42, 2253.
(22) Decostaire, I. P.; Lelièvre, D.; Zhang, H.; Delmas, A. F.
Tetrahedron Lett. 2006, 47, 7057.
(23) Lindsell, W. E.; Murray, C.; Preston, P. N.; Woodman, T. A.
J. Tetrahedron 2000, 56, 1233.
(24) Haridas, V.; Sharma, Y. K.; Sahu, S.; Verma, R. P.;
Sadanandan, S.; Kacheshwar, B. G. Tetrahedron 2011, 67,
1873.
(25) Maisonneuve, M.; Xie, J. Synlett 2009, 2977.
(26) Dueholm, K. L.; Egholm, M.; Behrens, C.; Christensen, L.;
Hansen, H. F.; Vulpius, T.; Petersen, K. H.; Berg, R. H.;
Nielsen, P. E.; Buchardt, O. J. Org. Chem. 1994, 59, 5767.
References
(1) Diederichsen, U. Angew. Chem., Int. Ed. Engl. 1996, 35,
445.
(2) Diederichsen, U.; Schmitt, H. W. Angew. Chem. Int. Ed.
1998, 37, 302.
(3) Chakraborty, P.; Diederichsen, U. Chem.–Eur. J. 2005, 11,
3207.
(4) McCarthy, O. K.; Schipani, A.; Buendía, A. M.; Ruiz-Perez,
L. M.; Kaiser, M.; Brun, R.; Pacanowska, D. G.; Gilbert, I.
H. Bioorg. Med. Chem. Lett. 2006, 16, 3809.
(5) Srivastava, R.; Ray, A. K.; Diederichsen, U. Eur. J. Org.
Chem. 2009, 4793.
(6) Geotti-Bianchini, P.; Crisma, M.; Peggion, C.; Bianco, A.;
Formaggio, F. Chem. Commun. 2009, 3178.
(7) Geotti-Bianchini, P.; Moretto, A.; Peggion, C.; Beyrath, J.;
Bianco, A.; Formaggio, F. Org. Biomol. Chem. 2010, 8,
1315.
Synthesis 2013, 45, 134–140
© Georg Thieme Verlag Stuttgart · New York