Synthesis of bis-aryl phosphates based on triazine scaffold

Article abstract of DOI:10.3987/COM-09-11877

In view of targeting the Grb2-SH2 signaling protein, the synthesis of potential ligands based on the 1,3,5-triazine scaffold bearing two phenylphosphate groups is described. The triazine functionality was introduced using an orthogonal strategy via the su

Full text of DOI:10.3987/COM-09-11877

HETEROCYCLES, Vol. 81, No. 4, 2010
867
HETEROCYCLES, Vol. 81, No. 4, 2010, pp. 867 - 882. © The Japan Institute of Heterocyclic Chemistry
Received, 2nd December, 2009, Accepted, 8th February, 2010, Published online, 9th February, 2010
DOI: 10.3987/COM-09-11877
SYNTHESIS OF BIS-ARYL PHOSPHATES BASED ON TRIAZINE
SCAFFOLD
Caroline Courme,a,b Nohad Gresh,c,d Christine Lenoir,c,d Michel Vidal,c,d
Christiane Garbay,c,d Jean-Claude Florent,a,b and Emmanuel Bertounesque*a,b
aCNRS UMR 176, 26 rue d’Ulm, 75005 Paris, France. Institut Curie, Centre de
b
c
Recherche, 26 rue d’Ulm, 75005 Paris, France. Université Paris Descartes, UFR
d
Biomédicale, 45 rue des Saints-Pères, 75006 Paris, France. INSERM U648,
Laboratoire de Pharmacochimie Moléculaire et Cellulaire, Paris, France. E-mail:
emmanuel.bertounesque@curie.fr
Abstract – In view of targeting the Grb2-SH2 signaling protein, the synthesis of
potential ligands based on the 1,3,5-triazine scaffold bearing two
phenylphosphate groups is described. The triazine functionality was
introduced using an orthogonal strategy via the sulfone chemistry.
INTRODUCTION
The 1,3,5-triazine scaffold has occupied an important position in drug discovery programs, especially
since Fesik identified it as a privileged structure using screening by NMR.1 Recent examples illustrate the
value of 1,3,5-triazine derivatives for cytotoxic activity,2 antitumoral activity,3 inhibition of angiogenesis,4
antimicrobial activity5 and anti-HIV activity.6 The use of this molecular scaffold would be also of interest
to design inhibitors of the Grb2 (growth factor receptor bound-protein 2) adaptor protein. Grb2 is
involved in RTK signal transduction via the binding of the SH2 domain to phosphotyrosines on the
C-terminal end of the activated receptors. Thus, it is implicated in Ras activation but can also play a role
in other signaling pathways in mammalian cells (e.g., PI3K signaling pathway). There has been
tremendous interest in the development of inhibitors of the SH2 domain of Grb2 as therapeutic agents in
treating cancers.7 Much of the effort in designing active site directed Grb2-SH2 inhibitors has focused on
the synthesis of peptidic compounds containing phosphotyrosine (pTyr) or phosphotyrosine mimics.8 To
date, only a limited number of non-peptidic antagonists of Grb-SH2 was reported from the low
micromolar range to the high micromolar range, i.e., urea 1,9 thiazole 2,10 tetrahydropyrimidinone 311 and
triazine 412 (Figure 1). The discovery of the latter class of compounds should facilitate the development of
more drug-like inhibitors for the treatment of cancers.

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HETEROCYCLES, Vol. 81, No. 4, 2010
OPO3H2
O
NH2
NH
H
N
NH2
N
O
N
NH
O
N
H
N
HN
N
CONH2
O
O
N
N
H
S
N
N
N
H
HN
O
O
H2O3PO
H2O3PO
OPO3H2
H2O3PO
1 IC50 = 6.19 μM
2 IC50 = 25.9 μM
4 IC50 = 265 μM
3 (UCM104) IC50 = 174 μM
Figure 1. Non-peptidic antagonists 1-4
In the context of our ongoing research on Grb2 targeting,12-14 we report herein the synthesis of novel
triazine scaffold-based bis-aryl phosphates 5-7 (Figure 2), the design of which was inspired by the
structure of triazine 4, using a flexible synthetic strategy. We have sought to replace the 4-phenoxyphenyl
substructure by the simpler 4-aminomethylphenyl one. For the three compounds, MD simulations showed
that this modification enabled to dock, on the one hand, its linked phosphate group into the pTyr-binding
pocket encompassing residues Arg 86, Ser 88, Ser 90, Ser 96 and Lys 109; and, on the other hand, the
aniline-bound phosphate into the second pTyr pocket, involving residues Arg 142, Asn 143, and Ser 141.
In particular, it appeared interesting to introduce a urea functionality on the triazine framework so as to
obtain hydrogen-bonding interactions with the main-chain carbonyl of Leu 120 and amide of Lys 109 in
the specificity pocket.
OPO3H2
HN
N
R
N
N
HN
OPO3H2
5 R = NH2
6 R = NHCONH2
7 R = CONH2
Figure 2. Triazine scaffold-based bis-aryl phosphates 5-7
RESULTS AND DISCUSSION
The bis-aryl phosphates 5-7 were synthetized using the orthogonal strategy developped by Chang et al.15
from cyanuric chloride (Scheme 1), wherein the last substitution step takes place via an
oxidation-activation of the benzylthioether group of 2-(benzylthio)-4,6-dichloro-1,3,5-triazine 9.

HETEROCYCLES, Vol. 81, No. 4, 2010
869
OPO3H2
OR
O
O
HN
N
R
HN
N
S
Cl
N
SBn
Bn
N
N
N
N
N
N
HN
HN
Cl
9
OPO3H2
OR
5-7
8
Scheme 1. Orthogonal strategy via sulfone chemistry
The synthesis of aminotriazine 5 began with a one-pot sequential procedure, involving addition of benzyl
mercaptan to cyanuric chloride followed by that of 4-aminophenol to give 10 in 82% yield (Scheme 2).
OR
OPO(OBn)2
OPO(OBn)2
OH
O
O
HN
N
SBn
HN
N
S
HN
N
NH2
Cl
N
Cl
HN
N
SBn
a
b
d
e
Bn
N
N
N
N
N
N
N
N
N
N
HN
HN
HN
Cl
Cl
10
OPO3R2
OR
OPO(OBn)2
OPO(OBn)2
11 R = H
12 R = PO(OBn)2
13
14
c
HN
N
NH2
f
N
N
HN
OPO3R2
5 R = H
15 R = Na
g
Scheme 2. Reagents and conditions: (a) BnSH (0.94 eq), DIEA (0.95 eq), THF, 0 °C, 1.5 h then
4-HOC6H4NH2 (1 eq), DIEA (1 eq), THF, 25 °C, 3 h, 82%; (b) 4-HOC6H4CH2NH2 (1.09 eq), DIEA (1.09
eq), THF, reflux, 36 h, 92%; (c) HPO(OBn)2 (4 eq), DIEA (2 eq), DMAP (1 eq), MeCN/CCl4 (5:1),
–15 °C, 1.5 h, 54%; (d) MCPBA (2.5 eq), THF 0 °C, 2 h, 25 °C, 15 min, 79%; (e) NH3(g), CH2Cl2, 25 °C,
15 h, 72%; (f) H2, 10% Pd/C (50% w/w), MeOH, 25 °C, 15 h, 36%; (g) MeONa (4 eq), MeOH, 0 °C, 2 h,
quant.
Subsequent addition of 4-hydroxybenzylamine furnished 11 (75% overall yield). The latter has also
been synthesized in a one-pot synthesis from cyanuric chloride but in a lower overall yield (61%).
Phosphorylation of 11 with dibenzylchlorophosphate (in situ prepared from dibenzylphosphite) gave

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HETEROCYCLES, Vol. 81, No. 4, 2010
12 in 54% yield.16 Oxidation of the benzylsulfanyl group with MCPBA led to the desired sulfone 13
in 79% yield. Then, a two-step sequence (i. e., substitution of ammonia and hydrogenation over Pd/C)
allowed to obtain amino-triazine 5 of very low solubility which was finally converted into the
water-soluble sodium salt 15. In order to access to urea-triazine 6, preliminary investigation showed
that the formation of the urea group from 14 failed, due to its unreactivity, under classical conditions
(KOCN 1-2 eq, AcOH/H2O or MeOH/1M HCl, rt or 50 °C, 2 h). The clivage of a dibenzylphosphate
group was observed when prolonging the reaction time (4 h) with 2.5 eq of KOCN.
OBn
OBn
H
N
KOCN
HN
N
NH2
HN
N
NH2
OR
OBn
N
N
N
N
O
or
O
HN
HN
d
NO2
H2N
N
H
HN
N
SBn
HN
N
SO2Bn
a
c
10
N
N
N
N
e
OBn
19
OBn
20
HN
HN
OBn
OR
OBn
OBn
h
H
N
H
N
16 R = H
17 R = Bn
18
HN
N
HN
N
NH2
b
f
N
N
N
N
N
O
HN
HN
OR
OBn
O
OBn
21
OR
HN
N
HN
N
CN
i
NH2
20 R = Bn
22 R = H
N
N
N
N
g
HN
HN
OR
OBn
23
24 R = Bn
25 R = H
j
Scheme 3. Reagents and conditions: (a) 4-BnOC6H4CH2NH2 (2 eq), DIEA (2 eq), THF/MeOH (5:1),
reflux, 15 h, 92%; (b) BnBr (1.5 eq), K2CO3 (1.5 eq), DMF, 0 °C, 1 h, 25 °C, 1 h, 75%, (c) MCPBA (3
eq), THF, 0 °C, 2 h, 79%; (d) NH3(g), CH2Cl2, 25 °C, 15 h, 72%; (e) H2NCN (10 eq), DIEA (2 eq), THF,
reflux, 16 h; (f) 4 N HCl/EtOH (1:1), 90 °C, 16 h; (g) H2, 10% Pd/C, MeOH/AcOH, 25 °C, 16 h, 42%
overall yield (three steps); (h) KCN (3 eq), DMF, 80 °C, 3 h, 96% (crude yield); (i) 1N NaOH/ EtOH/
30% H2O2 (6:3:1) 60 °C, 15 h, 64%; (j) H2, 10% Pd/C (50% w/w), MeOH/AcOH (1:1), 25 °C, 36 h, 63%.

HETEROCYCLES, Vol. 81, No. 4, 2010
871
Additionally, benzylation prior to the phosphorylation reaction led us to the synthesis of
N2-(4-(benzyloxy)benzyl)-N4-(4-(benzyloxy)phenyl)-1,3,5-triazine-2,4,6-triamine 19 (Scheme 3). The
latter was accessible via a four-step sequence from 10, involving the preparation of
(4-(benzyloxy)phenyl)methanamine.17 Triazine 19 was also found unaffected, even under strong
conditions (KOCN 2.4 eq, AcOH/H2O, reflux, 16 h). Moreover, formation of the urea 20 from this
protected aminotriazine was also unsuccessful using 1-nitrourea (H2SO4 cc 0.05%, EtOH, reflux, 2 h
or H2SO4 cc 4-8%, EtOH, 80 °C, 16 h). Finally, the sulfone derivative 18 was allowed to react with
cyanamide in refluxing tetrahydrofuran, affording the desired compound 21. Subsequent hydrolysis
using sulfuric acid at high temperature led to the corresponding urea 20 which underwent deprotection
of the two benzyl groups by hydrogenolysis to give 22 as a precursor of 6.
The introduction of the carboxamide group was also carried out starting from sulfonyle 18 (Scheme 3).
Treatment of 18 with potassium cyanide in dimethylformamide at 80 °C led to nitrile 23. Note that
cyanation from 13 was possible but much less satisfying in terms of yield (KCN 2.5 eq, DMF, 80 °C,
3 h, 43%). Hydrolysis of 23 using the method described by Dragovich et al.18 (sodium hydroxide and
hydrogen peroxide at 60 °C) gave the corresponding carboxamide derivative 24. Hydrogenation over
Pd/C then gave compound 25.
Phosphorylation of both derivatives 22 and 25 gave 26 and 27, respectively (Scheme 4). Following
catalytic hydrogenation, the resulting bis-phosphates (6 and 7, resp.) of very low solublity were
directly transformed into the sodium salts 28 and 29, respectively.
OPO(OBn)2
OPO3H2
OPO3Na2
HN
N
R
HN
N
R
HN
N
R
a
b
c
22, 25
N
N
N
N
N
N
HN
HN
HN
OPO(OBn)2
OPO3H2
OPO3Na2
26 R = NHCONH2
27 R = CONH2
6 R = NHCONH2
7 R = CONH2
28 R = NHCONH2
29 R = CONH2
Scheme 4. Reagents and conditions: (a) HPO(OBn)2 (5 eq), DIEA (8 eq), DMAP (1 eq), MeCN/CCl4
(5:1), –15 °C, 1.5 h, 26 (61%), 27 (72%); (b) H2, 10% Pd/C (50% w/w), MeOH, 25 °C, 15 h, 6 (61%), 7
(43%); (c) MeONa (4 eq), MeOH, 0 °C, 2 h, 28 (quant.), 29 (quant.).

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Bis-aryl phosphate sodium salts 15, 28 and 29 were assayed for binding to Grb2 using an ELISA
test.19 The phosphorylated peptide PSpYVNVPQ (Kd = 9 nM) was used for this competitive binding
assay. None of the designed compounds were found to be antagonists of the SH2 domain of Grb2
contrary to earlier predictions from MD simulations of their complexes (unpublished) in the Grb2
SH2 domain structure with implicit solvent. These results reassert the importance of accounting for
explicit water molecules in the MD simulations. Water probably plays a critical role in the stability of
such Grb2 complexes with peptide and non-peptidic inhibitors as shown recently by Leroux et al.20
We have synthesized novel bis-aryl phosphates based on the privileged structure triazine, using an
orthogonal strategy from cyanuric chloride, via the sulfone chemistry. In order to strengthen the
1,3,5-trisubstituted triazine scaffold-based ligand design strategy that exploits the use of mimics of
phosphates (e.g., keto-acid, IZD),12 further molecular modeling of such compounds through MD
simulations in water should be also considered.
EXPERIMENTAL
All commercial reagents were used without purification and all solvents were reaction grade. When
necessary, solvents were previously dried over molecular sieves. Tetrahydrofuran was also dried over
molecular sieves 4 Å unless otherwise stated (distilled from sodium/benzophenone under argon). All
reactions were performed under an inert atmosphere of argon unless otherwise stated. All reaction
mixtures were stirred magnetically and monitored by thin-layer chromatography using Merck silica
gel 60 F254, visualized with UV light. Flash chromatography were performed using SDS silica gel 60
(35-70 μm). Melting points (uncorrected) were determined on a Kofler bench. Mass spectra were
recorded on a ZQ 2000 MS (ES) or a spectrometer (LC) HRMS ESI/TOF (LCT, Waters).
13
31
1H, C and P NMR spectra were recorded on a Bruker AM 300 spectrometer. Chemical shift values
are reported in parts per million (ppm) and coupling constants in Hertz (Hz). In order to simplify the
reading of the NMR spectra interpretations (COSY, HMQC, HMBC), the following attributions were
chosen (Scheme 5): Letter "A" will always refer to the heterocyclic core – whether it is a pyrimidine
or a triazine – and letter "B" to the aryl group linked to this heterocycle through an NH bridge. "C"
and "D" will refer to the diaryl ether and finally "E" will refer to the benzyl groups, either benzyloxy
or dibenzylphosphates. Two AA’BB’ systems were observed for the two para-substituted aromatic
rings B and D. Compounds 11, 12 and 17 are described below as examples.

HETEROCYCLES, Vol. 81, No. 4, 2010
873
O
2
E
5
2
E
5
1
1
P
O
O
3
4
3
4
OH
4
O
4
BnO
6
5
4
6
5
6
3
2
5
3
2
3
2
5
5
C
5
B
2
C
C
6
6
6
1
4
3
6
1
4
3
6
1
4
3
1
HN
1
1
HN
B
B
HN
N
S
N
S
N
S
4
2
N
4
2
N
4
2
N
2
2
A
A
A
N
N
N
6
6
6
HN
HN
HN
1
1
1
2
3
6
2
3
6
5
2
3
6
D
D
D
5
5
4
OH
4
OPO(OBn)2
4
OBn
11
12
17
Scheme 5
4-(4-Benzylthio-6-chloro-1,3,5-triazin-2-ylamino)phenol (10)
Benzylthiol (6 mL, 51.0 mmol) and diisopropylethylamine (9 mL, 51.7 mmol) were added to a
solution of cyanuric chloride (10 g, 54.2 mmol) in dry THF (200 mL) at 0 °C. After 2 h of stirring at
0 °C, 4-aminophenol (5.9 g, 54.1 mmol) and diisopropylethylamine (9.5 mL, 54.5 mmol) were added.
The mixture was then stirred at room temperature for 3 h and the solvent was removed under reduce
pressure. Purification by flash chromatography with cyclohexane/EtOAc (4:1) gave 10 as a white
solid (14.4 g, 82%). Mp 179 °C; 1H NMR (300 MHz; MeOD): ꢀ 4.28 (2H, s, CH2), 6.75-6.78 (2H, m,
13
3C, 5C), 7.21-7.23 (3H, m, 3B, 4B, 5B), 7.28-7.31 (2H, m, 2C, 6C), 7.35-7.43 (2H, m, 2B, 6B); C NMR
(75 MHz; MeOD): ꢀ 34.8 (CH2), 115.6 (3C, 5C), 123.5 (4B), 124.1 (3B, 5B), 127.5 (1C), 128.6 (2C, 6C),
129.0 (2B, 6B), 136.4 (1B), 154.6 (4C), 162.7 (4A), 168.2 (6A), 183.2 (2A); MS (ES+) m/z 345 [M+H]+,
367 [M+Na]+; MS (ES–) m/z 343 [M–H]–; HRMS (ES+) m/z: calcd for C16H13ClN4OS 345.0577
[M+H]+, 367.0396 [M+Na]+; found 345.0583, 367.0404.
4-[4-Benzylthio-6-(4-hydroxybenzylamino)-1,3,5-triazin-2-ylamino]phenol (11)
Compound 10 (234 mg, 0.679 mmol) and 4-(aminomethyl)phenol (91 mg, 0.739 mmol) were
dissolved in dry THF. Diisopropylethylamine (140 μL, 0.804 mmol) was added and the mixture was
refluxed for 36 h. The solvent was removed under reduce pressure. Purification by flash
chromatography with cyclohexane/EtOAc (gradient from 4:1 to 1:1) gave 11 as a white solid (270 mg,
92%). Mp 94-97 °C; 1H NMR (300 MHz; MeOD): ꢀ 4.31 (2H, s, S-CH2), 4.42 (2H, s, NH-CH2), 6.72
(4H, br s, 3C, 5C, 3D, 5D), 7.11-7.14 (2H, m, 2D, 6D), 7.20-7.28 (5H, m, 2B, 3B, 4B, 5B, 6B), 7.31-7.33 (2H,
13
m, 2C, 6C); C NMR (75 MHz; MeOD): ꢀ 34.7 (S-CH2), 44.8 (NH-CH2), 116.1 (3C, 5C, 3D, 5D), 124.5
(2C, 6C), 129.4, 129.7, 130.0 (2B, 3B, 4B, 5B, 6B, 2D, 6D), 131.5 (1C), 132.1 (1D), 139.5 (1B), 154.8 (4C),
157.4 (4D), 164.2 (4A), 165.6 (6A), 180.9 (2A); MS (ES+) m/z 432 [M+H]+, 454 [M+Na]+; MS (ES–) m/z
430 [M–H]–.

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HETEROCYCLES, Vol. 81, No. 4, 2010
Dibenzyl 4-[4-benzylthio-6-(4-dibenzylphosphoryloxybenzylamino)-1,3,5-triazin-2-ylamino]-
phenyl phosphate (12)
To a solution of 11 (4 g, 9.27 mmol) in dry MeCN (200 mL) cooled at –15 °C was added dry CCl4
(40 mL) and the mixture was stirred at –15 °C for 5 min. Diisopropylethylamine (12.9 mL, 74.1
mmol) and N,N-dimethylaminopyridine (1.13 g, 9.27 mmol) were then added and the mixture was
stirred at –15 °C for 30 min. Dibenzylphosphite (8.2 mL, 37.1 mmol) was added dropwise and the
mixture was stirred at –15 °C for 1 h 30. A 0.5 M solution of potassium dihydrogen phosphate
(50 mL) was added, the mixture was warmed up to room temperature and extracted with EtOAc. The
combined extracts were then washed with water and brine, dried over magnesium sulfate and
evaporated under reduced pressure. Purification by flash chromatography with CH2Cl2/MeOH (99:1)
1
afforded 12 as a colourless oil (4.74 g, 54%). H NMR (300 MHz; CDCl3): ꢀ 4.33 (2H, s, S-CH2),
4.55 (2H, m, NH-CH2), 5.10 (m, 4H, 2 ꢀ P-O-CH2), 5.12 (m, 4H, 2 ꢀ P-O-CH2), 7.04-7.11 (4H, m, 3C,
5C, 3D, 5D), 7.20-7.26 (2H, m, 2D, 6D), 7.24-7.31 (25H, m, 4 ꢀ 2E, 4 ꢀ 3E, 4 ꢀ 4E, 4 ꢀ 5E, 4 ꢀ 6E, 2B, 3B,
13
4B, 5B, 6B), 7.40-7.43 (m, 2H, 2C, 6C); C NMR (75 MHz; CDCl3): ꢀ 34.1 (S-CH2), 44.1 (NH-CH2),
69.9 (P-O-CH2), 120.1, 120.2 (3C, 5C, 3D, 5D), 121.7 (2C, 6C), 127.1, 128.0, 128.4, 128.5, 128.6, 128.7,
128.8 (2E, 3E, 4E, 5E, 6E, 2B, 3B, 4B, 5B, 6B, 2D, 6D), 135.2, 135.3, 135.4 (1C, 1D, 1E), 137.4 (1B), 146.0
(4C), 149.7 (4D), 162.7 (4A), 164.5 (6A), 180.3 (2A).
Dibenzyl 4-[4-benzylsulfonyl-6-(4-dibenzylphosphoryloxybenzylamino)-1,3,5-triazin-2-ylamino]-
phenyl phosphate (13)
MCPBA (430 mg, 2.49 mmol) was added portionwise over 20 min to a solution of thioether 12 (1 g,
1.05 mmol) in THF (50 mL) at 0 °C under air. The mixture was stirred at 0 °C for 2 h and at room
temperature for 15 min and a saturated aqueous sodium hydrogen carbonate solution was added
(50 mL). The product was extracted with EtOAc, the combined extracts were then washed with water
and brine, dried over magnesium sulfate and evaporated under reduced pressure. Purification by flash
chromatography with CH2Cl2/MeOH (100:1.5) afforded 13 as a beige powder (820 mg, 79%). Mp
47 °C; 1H NMR (300 MHz; CDCl3): ꢀ 4.54 (2H, s, NH-CH2), 4.61 (2H, s, SO2-CH2), 5.10 (4H, s, 2 ꢀ
P-O-CH2), 5.12 (4H, s, 2 ꢀ P-O-CH2), 7.11 (4H, m, 3C, 5C, 3D, 5D), 7.17 (2H, m, 2D, 6D), 7.31 (25H, br
13
s, 4 ꢀ 2E, 4 ꢀ 3E, 4 ꢀ 4E, 4 ꢀ 5E, 4 ꢀ 6E, 2B, 3B, 4B, 5B, 6B), 7.37-7.45 (4H, m, 2C, 6C); C NMR (75
MHz; CDCl3): ꢀ 44.6 (NH-CH2), 56.7 (SO2-CH2), 70.0 (P-O-CH2), 70.1 (P-O-CH2), 120.4, 120.5 (3C,
5C, 3D, 5D), 122.0 (2C, 6C), 126.6 (2B, 6B), 128.0 (2E, 6E), 128.6, 128.7, 128.8 (3E, 4E, 5E, 2D, 6D, 3B, 4B,
5B), 131.4 (1B), 134.0 (1D), 134.3 (1C), 135.4 (1E), 147.0 (4C), 150.0 (4D), 163.7 (4A), 165.5 (6A), 172.4
(2A); MS (ES+) m/z 1001 [M+NH4]+, 1006 [M+Na]+.

HETEROCYCLES, Vol. 81, No. 4, 2010
875
4-[4-Amino-6-(4-dibenzylphosphoryloxybenzylamino)-1,3,5-triazin-2-ylamino]phenyl dibenzyl
phosphate (14)
Ammonia was bubbled for 1 h into a solution of compound 13 (298 mg, 0.303 mmol) in CH2Cl2
(15 mL) at room temperature. The flask was then sealed and the mixture was stirred for 15 h. The
solvent was removed under reduce pressure and purification by flash chromatography with
CH2Cl2/MeOH (100:4) gave 14 as a white solid (184 mg, 72%). Mp 114-117 °C; 1H NMR (300 MHz;
CDCl3): ꢀ 4.53 (2H, s, NH-CH2), 5.09 (4H, s, 2 ꢀ P-O-CH2), 5.11 (4H, s, 2 ꢀ P-O-CH2), 7.04-7.10
(4H, m, 3C, 5C, 3D, 5D), 7.21-7.24 (2H, m, 2D, 6D), 7.31 (20H, br s, 4 ꢀ 2E, 4 ꢀ 3E, 4 ꢀ 4E, 4 ꢀ 5E, 4 ꢀ 6E),
7.42-7.45 (2H, m, 2C, 6C); 13C NMR (75 MHz; CDCl3): ꢀ 44.1 (NH-CH2), 70.0 (P-O-CH2), 70.1
(P-O-CH2), 120.3, 120.4 (3C, 5C, 3D, 5D), 121.5 (2C, 6C), 128.1 (2E, 6E), 128.7 (3E, 4E, 5E), 128.8 (2D, 6D),
135.5 (1E), 135.9, 136.1 (1D, 1C), 146.0 (4C), 149.8 (4D), 164.7 (4A), 166.5 (6A, 2A); MS (ES+) m/z 844
[M+H]+, 867 [M+Na]+.
4-[4-Amino-6-(4-phosphoryloxybenzylamino)-1,3,5-triazin-2-ylamino]phenyl phosphate (5)
10% palladium on carbon (60 mg) was added to a solution of compound 14 (121 mg, 0.143 mmol) in
MeOH (50 mL). The mixture was stirred under H2 at room temperature for 15 h. The catalyst was
filtered using two filter papers and washed with hot MeOH, affording 5 as a white solid (25 mg, 36%).
Mp 216-218 °C with decomposition; MS (ES–) m/z 241 [M–2H]2–.
Tetrasodium 4-[4-amino-6-(4-phosphonatooxybenzyl amino)-[1,3,5]-triazin-2-ylamino]phenyl
phosphate (15)
A fresh 0.539 M sodium methoxide solution (276 μL, 149 μmol) was added to a suspension of
compound 5 (18 mg, 37.2 μmol) in dry MeOH (2.5 mL) at 0 °C. The mixture was stirred at 0 °C for
2 h and the MeOH was removed under reduced pressure to afford 15 as a water-soluble white powder
1
(21 mg, 99%). H NMR (300 MHz; D2O): ꢀ 4.37 (2H, s, NH-CH2), 7.06 (4H, br s, 3C, 5C, 3D, 5D),
13
7.16-7.19 (4H, m, 2C, 6C, 2D, 6D); C NMR (75 MHz; D2O): ꢀ 43.3 (NH-CH2), 120.5, 120.6 (3C,
5C, 3D, 5D), 124.4 (2C, 6C), 127.9 (2D, 6D), 132.0 (1C), 132.7 (1D), 150.8, 152.9 (4C, 4D), 164.6
(4A), 165.6 (6A), 166.8 (2A); 31P NMR (121 MHz; D2O): ꢀ 0.51; HRMS (ES–) m/z: calcd for
C16H14N6Na4O8P2 527.0222 [M–2Na+H]–, 505.0403 [M–3Na+2H] –; found 527.0247, 505.0428.
4-[4-(4-Benzyloxybenzylamino)-6-benzylthio-1,3,5-triazin-2-ylamino]phenol (16)
Compound 10 (664 mg, 1.93 mmol) and (4-benzyloxyphenyl)methanamine (821 mg, 3.85 mmol)
were dissolved in a THF/MeOH 5:1 mixture (18 mL). Diisopropylethylamine (670 μL, 3.85 mmol)
was added and the mixture was refluxed for 15 h. The solvent was removed under reduce pressure and

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HETEROCYCLES, Vol. 81, No. 4, 2010
purification by flash chromatography with cyclohexane/EtOAc (4:1) gave 16 as a white solid (926 mg,
1
92%). Mp 117 °C; H NMR (300 MHz; CDCl3): ꢀ 4.32 (2H, s, S-CH2), 4.51 (2H, s, NH-CH2), 5.03
(2H, s, O-CH2), 5.60 (1H, br s, NH-CH2), 6.71 (2H, s, 3C, 5C), 6.60-6.92 (2H, m, 3D, 5D), 7.17-7.19
13
(2H, m, 2D, 6D), 7.23-7.40 (12H, m, 2C, 6C, 2B, 3B, 4B, 5B, 6B, 2E, 3E, 4E, 5E, 6E); C NMR
(75 MHz; CDCl3): ꢀ 34.1 (S-CH2), 44.3 (NH-CH2), 70.1 (O-CH2), 115.0 (3D, 5D), 115.6 (3C, 5C),
123.0 (2C, 6C), 127.1 (4B), 127.5 (2E, 6E), 128.0 (4E), 128.5, 128.6 (3B, 5B, 3E, 5E), 128.8, 128.9
(2D, 6D, 2B, 6B), 130.7 (1D), 131.0 (1C), 136.9 (1E), 136.6 (1B), 152.2 (4C), 158.1 (4D), 162.9 (4A),
164.2 (6A), 179.5 (2A); MS (ES+) m/z 522 [M+H]+, 544 [M+Na]+, MS (ES–) m/z 520 [M–H]–.
N2-(4-Benzyloxybenzyl)-N4-(4-benzyloxyphenyl)-6-benzylthio-1,3,5-triazine-2,4-diamine (17)
Potassium carbonate (1.9 g, 13.7 mmol) was added to a solution of 16 (4.72 g, 9.05 mmol) in dry
DMF (200 mL). Benzyl bromide (1.6 mL, 13.5 mmol) was then added dropwise at 0 °C and the
mixture was stirred at 0 °C for 1 h and then at room temperature for 1 h. Water was added and the
mixture was extracted with Et2O. The combined extracts were washed with water and brine, dried
over magnesium sulfate and evaporated under reduced pressure. Purification by flash chromatography
with cyclohexane/EtOAc (5:1) gave 17 as a white powder (4.18 g, 75%). Mp 45-47 °C; 1H NMR (300
MHz; CDCl3): ꢀ 4.33 (2H, s, S-CH2), 4.54 (2H, s, NH-CH2), 5.04 (4H, s, O-CH2), 5.61 (1H, br s,
NH-CH2), 6.89-6.94 (4H, m, 3C, 5C, 3D, 5D), 7.02 (1H, br s, NH), 7.23-7.26 (5H, m, 2D, 6D, 3B, 4B,
5B), 7.35-7.41 (14H, m, 2C, 6C, 2B, 6B, 2 ꢀ 2E, 2 ꢀ 3E, 2 ꢀ 4E, 2 ꢀ 5E, 2 ꢀ 6E); 13C NMR (75 MHz;
CDCl3): ꢀ 34.3 (S-CH2), 44.5 (NH-CH2), 70.2 (O-CH2), 115.2 (3C, 5C, 3D, 5D), 122.8 (2C, 6C),
127.2 (4B), 127.6 (2E, 6E), 128.1 (4E), 128.6, 128.7 (2B, 6B, 2D, 6D, 3E, 5E), 129.1 (3B, 5B), 130.9
(1D), 131.8 (1C), 137.1 (1E), 137.5 (1B), 155.3 (4C), 158.3 (4D), 162.9 (4A), 164.6 (6A), 178.7 (2A).
N2-(4-Benzyloxybenzyl)-N4-(4-benzyloxyphenyl)-6-benzylsulfonyl-1,3,5-triazine-2,4-diamine (18)
Sulfone 18 was prepared as sulfone 13, from compound 17 (4.2 g, 6.87 mmol) and MCPBA (3.6 g,
20.9 mmol). Purification via flash chromatography with cyclohexane/EtOAc (5:1) afforded 18 as an
1
off-white solid (3.5 g, 79%). Mp 144-151 °C; H NMR (300 MHz; CDCl3): ꢀ 4.53 (2H, s, NH-CH2),
4.62 (2H, s, SO2-CH2), 5.06 (4H, s, O-CH2), 5.86 (1H, br s, NH-CH2), 6.95 (4H, s, 3C, 5C, 3D, 5D),
7.19-7.22 (2H, m, 2D, 6D), 7.31-7.41 (17H, m, 2C, 6C, 2B, 3B, 4B, 5B, 6B, 2 ꢀ 2E, 2 ꢀ 3E, 2 ꢀ 4E, 2 ꢀ 5E, 2
ꢀ 6E); 13C NMR (75 MHz; CDCl3): ꢀ 44.9 (NH-CH2), 56.4 (SO2-CH2), 70.1 (O-CH2), 115.1, 115.2 (3C,
5C, 3D, 5D), 122.5 (2C, 6C), 126.6 (1B), 127.4 (2E, 6E), 128.0 (4E), 128.6, 128.8 (3B, 4B, 5B, 2D, 6D, 3E, 5E),
129.1, 129.5 (1C, 1D), 131.4 (2B, 6B), 136.8 (1E), 156.0 (4C), 158.4 (4D), 163.8 (4A), 165.6 (6A), 172.0
(2A); MS (ES+) m/z 666 [M+Na]+.

HETEROCYCLES, Vol. 81, No. 4, 2010
877
N2-(4-Benzyloxybenzyl)-N4-(4-benzyloxyphenyl)-1,3,5-triazine-2,4,6-triamine (19)
Ammonia was bubbled for 1 h into a solution of sulfone 18 (2 g, 3.11 mmol) in CH2Cl2 (100 mL) at
room temperature. The flask was then sealed and the mixture was stirred for 15 h. The solvent was
removed under reduce pressure and purification by flash chromatography with CH2Cl2/MeOH
1
(gradient from 100:2 to 100:5) gave 19 as a white powder (1.12 g, 72%). Mp 77 °C; H NMR (300
MHz; CDCl3): ꢀ 4.48 (2H, s, NH-CH2), 5.02 (4H, s, O-CH2), 5.81 (1H, br s, NH-CH2), 6.87-6.93 (4H,
m, 3C, 5C, 3D, 5D), 7.19-7.22 (2H, m, 2D, 6D), 7,29-7,43 (12H, m, 2C, 6C, 2 ꢀ 2E, 2 ꢀ 3E, 2 ꢀ 4E, 2 ꢀ 5E, 2
ꢀ 6E); 13C NMR (75 MHz; CDCl3): ꢀ 44.1 (N-CH2), 70.0 (O-CH2), 114.9, 115.0 (3C, 5C, 3D, 5D), 122.4
(2C, 6C), 127.4 (2E, 6E), 127.9 (4E), 128.5 (3E, 5E), 128.7 (2D, 6D), 131.2 (1D), 132.0 (1C), 136.9 (1E),
154.9 (4C), 158.0 (4D), 164.5 (4A), 166.1 (6A, 2A); MS (ES+) m/z 505 [M+H]+, 527 [M+Na]+; MS (ES–)
m/z 503 [M–H]–.
N-[4-(4-Benzyloxybenzylamino)-6-(4-benzyloxyphenylamino)-1,3,5-triazin-2-yl]cyanamide (21)
Sulfone 18 (707 mg, 1.10 mmol) was dissolved in THF (100 mL). Cyanamide (461 mg, 11.0 mmol)
and diisopropylethylamine (380 μL, 2.18 mmol) were added and the mixture was refluxed overnight.
THF was removed under reduce pressure, water was added (50 mL) and the product was extracted
with EtOAc. The combined extracts were then washed with water and brine, dried over magnesium
sulfate and evaporated under reduced pressure to give an off-white powder (1.05 g). This crude
product (21) proved to be highly insoluble and was hence used in the next step without purification.
MS (ES+) m/z 530 [M+H]+, 552 [M+Na]+, MS (ES–) m/z 528 [M–H]–.
1-[4-(4-Benzyloxybenzylamino)-6-(4-benzyloxyphenylamino)-1,3,5-triazin-2-yl]urea (20)
The crude product 21 (826 mg) was placed into a 4 N HCl/EtOH mixture (1:1, 50 mL). The mixture
was heated at 90 °C for 16 h then neutralized with 1 N NaOH. The product was extracted with EtOAc.
The combined extracts were then washed with water and brine, dried over magnesium sulfate and
evaporated under reduced pressure to give a white powder (477 mg). As in the previous reaction,
compound 20 was insoluble and therefore used in the next step without purification. MS (ES+) m/z
548 [M+H]+.
1-[4-(4-Hydroxybenzylamino)-6-(4-hydroxyphenylamino)-1,3,5-triazin-2-yl]urea (22)
10% Palladium on carbon (230 mg) was added to a suspension of crude compound 20 (477 mg) in a
MeOH/AcOH mixture (1:1, 250 mL). The mixture was stirred under an hydrogen atmosphere at room
temperature for 16 h. The catalyst was filtered and the product was purified by flash chromatography
with CH2Cl2/MeOH (100:8) to give 22 as a white powder (133 mg, 42% yield over 3 steps). Mp

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HETEROCYCLES, Vol. 81, No. 4, 2010
264 °C; 1H NMR (300 MHz; DMSO-d6): ꢀ 4.32 (2H, s, NH-CH2), 6.64-6.67 (2H, m, 3C, 5C), 6.68-6.71
(2H, m, 3D, 5D), 7.09-7.12 (2H, m, 2D, 6D), 7.38-7.41 (2H, m, 2C, 6C), 9.13 (1H, br s, 4C-OH), 9.28 (1H,
13
br s, 4D-OH); C NMR (75 MHz; DMSO-d6): ꢀ 43.1 (NH-CH2), 114.9 (3C, 5C, 3D, 5D), 121.7 (2C, 6C),
128.5 (2D, 6D), 129.8 (1D), 131.0 (1C), 152.9 (4C), 156.0 (CO), 156.2 (4D), 163.0, 164.9, 165.1 (2A, 4A,
6A); MS (ES+) m/z 368 [M+H]+, 390 [M+Na]+.
1-[4-(4-Dibenzylphosphoryloxybenzylamino)-6-(4-dibenzylphosphoryloxyphenylamino)-1,3,5-
triazin-2-yl]urea (26)
To a solution of 22 (60 mg, 0.163 mmol) in dry MeCN (10 mL) cooled at –15 °C was added dry CCl4
(2 mL) and the mixture was stirred at –15 °C for 5 min. Diisopropylethylamine (230 μL, 1.32 mmol)
and N,N-dimethylaminopyridine (20 mg, 0.164 mmol) were then added and the mixture was stirred at
–15 °C for 30 min. Dibenzylphosphite (180 μL, 0.815 mmol) was added dropwise and the mixture
was stirred at –15 °C for 2 h. A 0.5 M solution of potassium dihydrogen phosphate (5 mL) was added,
the mixture was warmed up to room temperature and extracted with EtOAc. The combined extracts
were then washed with water and brine, dried over magnesium sulfate and evaporated under reduced
pressure. Purification by flash chromatography with CH2Cl2/MeOH (gradient from 100:2 to 100:5)
1
afforded 26 as a colorless oil (89 mg, 61%). H NMR (300 MHz; CDCl3): ꢀ 4.44 (2H, s, NH-CH2),
5.08 (4H, s, P-O-CH2), 5.10 (4H, s, P-O-CH2), 7.03-7.10 (4H, m, 3C, 5C, 3D, 5D), 7.22-7.26 (2H, m, 2D,
13
6D), 7.30 (20H, br s, 4 ꢀ 2E, 4 ꢀ 3E, 4 ꢀ 4E, 4 ꢀ 5E, 4 ꢀ 6E), 7.56-7.67 (m, 2H, 2C, 6C); C NMR (75
MHz; CDCl3): ꢀ 44.3 (NH-CH2), 69.9 (P-O-CH2), 70.0 (P-O-CH2), 120.1, 120.5 (3C, 5C, 3D, 5D), 122.4
(2C, 6C), 128.0, 128.3, 128.6 (2E, 3E, 4E, 5E, 6E, 2D, 6D), 135.4 (1C, 1D, 1E), 147.0 (4C), 149.9 (4D), 157.1
(CO), 163.0, 163.3 (2A, 4A, 6A); MS (ES+) m/z 888 [M+H]+, 910 [M+Na]+.
1-[4-(4-Phosphoryloxybenzylamino)-6-(4-phosphoryloxyphenylamino)-1,3,5-triazin-2-yl]urea (6)
10% Palladium on carbon (41 mg) was added to a solution of compound 26 (83 mg, 93.4 μmol) in
MeOH (30 mL). The mixture was stirred under an hydrogen atmosphere at room temperature
overnight. The catalyst was filtered using two filter papers, then washed with both hot MeOH and hot
water. Purification by flash chromatography on reverse phase silica gel C18 with water/MeCN (95:5)
gave 6 as a white solid (30 mg, 61%). Mp 209-211 °C with decomposition; MS (ES–) m/z 262
[M–2H]2–.
Sodium 1-[4-(4-phosphonatooxybenzylamino)-6-(4-phosphonatooxyphenylamino)-1,3,5-triazin-
2-yl]urea (28)
A fresh 0.539 M sodium methoxide solution (205 μL, 111 μmol) was added to a suspension of

HETEROCYCLES, Vol. 81, No. 4, 2010
879
compound 6 (14 mg, 27.7 μmol) in dry MeOH (3 mL) at 0 °C. The mixture was stirred at 0 °C for 2 h
and the MeOH was removed under reduced pressure to afford 28 as a water-soluble white powder (17
1
mg, 100%). H NMR (300 MHz; D2O): ꢀ 4.37 (2H, s, NH-CH2), 7.03 (4H, br s, 3C, 5C, 3D, 5D), 7.15
(2H, br s, 2D, 6D), 7.19-7.22 (2H, m, 2C, 6C); 13C NMR (75 MHz; D2O): ꢀ 43.5 (NH-CH2), 120.6 (3C, 5C,
3D, 5D), 124.1 (2C, 6C), 127.4 (2D, 6D), 130.6 (1C), 132.6 (1D), 150.4 (4C), 152.9 (4D), 157.5, 159.0 (CO,
2A, 4A), 164.4 (6A) P NMR (121 MHz; D2O): ꢀ 0.52; HRMS (ES–) m/z: calcd for C17H15N7Na4O9P2
31
592.0100 [M–Na]–; found 592.0108.
4-(4-Benzyloxybenzylamino)-6-(4-benzyloxyphenylamino)-1,3,5-triazine-2-carbonitrile (23)
Potassium cyanide (455 mg, 6.99 mmol) was cautiously added to a solution of compound 18 (1.5 g,
2.33 mmol) in dry DMF (80 mL). The mixture was heated at 80 °C for 3 h and the solvent was
removed under high vacuum. Water was added and the mixture was extracted with Et2O. The
combined extracts were washed with water and brine, dried over magnesium sulfate and evaporated
under reduced pressure to afford 23 as a yellow powder (1.15 g, 96%). This product was used in the
next step without purification. Mp 147 °C; 1H NMR (300 MHz; CDCl3): ꢀ 4.29 (2H, s, NH-CH2), 5.06
(4H, s, O-CH2), 6.95 (4H, s, 3C, 5C, 3D, 5D), 7.22-7.24 (2H, m, 2D, 6D), 7.36-742 (12H, m, 2C, 6C, 2 ꢀ 2E,
2 ꢀ 3E, 2 ꢀ 4E, 2 ꢀ 5E, 2 ꢀ 6E); 13C NMR (75 MHz; CDCl3): ꢀ 44.7 (NH-CH2), 70.2 (O-CH2), 115.3 (3C,
5C, 3D, 5D), 123.0, 123.3 (2C, 6C, CN), 127.6 (2E, 6E), 128.2 (4E), 128.8 (3E, 5E), 129.0 (2D, 6D), 129.4
(1D), 130.3 (1C), 136.9 (1E), 156.0 (4C), 158.5 (4D), 163.5 (4A), 165.1 (6A), 176.4 (2A) ; MS (ES+) m/z
515 [M+Na]+. HRMS (ES+) m/z: calcd for C31H26N6O2 537.2015 [M+Na]+; found 537.2014.
4-(4-Benzyloxybenzylamino)-6-(4-benzyloxyphenylamino)-1,3,5-triazine-2-carboxamide (24)
A solution of compound 23 (150 mg, 0.291 mmol) in a 1 N NaOH/ EtOH/ 30% H2O2 mixture (6:3:1,
10 mL) was heated at 60 °C overnight. EtOH was removed under reduce pressure, the mixture was
neutralized with 1 N HCl and the product was extracted with CH2Cl2. The combined extracts were
washed with water and brine, dried over magnesium sulfate and evaporated under reduced pressure.
Purification by flash chromatography with CH2Cl2/MeOH (gradient from 100:2 to 100:9) gave 24 as a
1
beige solid (100 mg, 64%). Mp 222 °C; H NMR (300 MHz; CDCl3): ꢀ 4.43 (2H, m, NH-CH2), 5.07
(4H, s, O-CH2), 6.94 (4H, m, 3C, 5C, 3D, 5D), 7.22-7.43 (12H, m, 2D, 6D, 2 ꢀ 2E, 2 ꢀ 3E, 2 ꢀ 4E, 2 ꢀ 5E, 2
ꢀ 6E), 7.57 (2H, m, 2C, 6C); 13C NMR (75 MHz; CDCl3): ꢀ 42.9 (NH-CH2), 69.2 (O-CH2), 114.6 (3C, 5C,
3D, 5D), 121.4 (2C, 6C), 127.6 (2E, 6E), 127.7 (4E), 128.4 (3E, 5E), 128.8 (2D, 6D), 131.6 (1D), 132.5 (1C),
137.1 (1E), 153.9 (4C), 157.2 (4D), 163.9, 165.0 (4A, CONH2), 165.7 (6A), 166.1 (2A); MS (ES+) m/z
532 [M+H]+, 555 [M+Na]+. HRMS (ES+) m/z: calcd for C31H28N6O3 555.2121 [M+Na]+; found
555.2115.

880
HETEROCYCLES, Vol. 81, No. 4, 2010
4-(4-Hydroxybenzylamino)-6-(4-hydroxyphenylamino)-1,3,5-triazine-2-carboxamide (25)
10% Palladium on carbon (77 mg) was added to a solution of compound 24 (770 mg, 1.45 mmol) in a
MeOH/AcOH mixture (1:1, 150 mL). The mixture was stirred under H2 at room temperature for 36 h.
The catalyst was filtered and the product was purified by flash chromatography with MeCN/water
1
(100:2.5) to afford 25 as pale yellow oil (322 mg, 63%). H NMR (300 MHz; MeOD): ꢀ 4.46 (2H, s,
13
NH-CH2), 6.71-6.75 (4H, m, 3C, 5C, 3D, 5D), 7.14-7.19 (2H, m, 2D, 6D), 7.37-7.40 (2H, m, 2C, 6C); C
NMR (75 MHz; MeOD): ꢀ 45.1 (NH-CH2), 116.2, 116.3 (3C, 5C, 3D, 5D), 123.5 (2C, 6C), 129.8 (2D, 6D),
131.0, 132.0 (1D, 1C), 154.8 (4C), 157.6 (4D), 165.7 (4A, CONH2), 167.5 (6A), 167.8 (2A); MS (ES+) m/z
353 [M+H]+, 375 [M+Na]+.
Dibenzyl 4-[4-(4-(dibenzylphosphoryloxy)benzylamino)-6-carbamoyl-1,3,5-triazin-2-ylamino]-
phenyl phosphate (27)
To a solution of 25 (322 g, 0.914 mmol) in dry MeCN (50 mL) cooled at –15 °C was added dry CCl4
(10 mL) and the mixture was stirred at –15 °C for 5 min. Diisopropylethylamine (1.4 mL, 8.04 mmol)
and N,N-dimethylaminopyridine (111 mg, 0.909 mmol) were then added and the mixture was stirred
at –15 °C for 30 min. Dibenzylphosphite (1 mL, 4.53 mmol) was added dropwise and the mixture was
stirred at –15 °C for 2 h. A 0.5 M solution of potassium dihydrogen phosphate (20 mL) was added, the
mixture was warmed up to room temperature and extracted with EtOAc. The combined extracts were
then washed with water and brine, dried over magnesium sulfate and evaporated under reduced
pressure. Purification by flash chromatography with CH2Cl2/MeOH (gradient from 100:2 to 100:8)
1
afforded 27 as a colorless oil (574 mg, 72%). H NMR (300 MHz; CDCl3): ꢀ 4.57 (2H, m, NH-CH2),
5.06-5.10 (8H, m, P-O-CH2), 7.08-7.11 (4H, m, 3C, 5C, 3D, 5D), 7.21-7.24 (2H, m, 2D, 6D), 7.30 (20H,
13
br s, 4 ꢀ 2E, 4 ꢀ 3E, 4 ꢀ 4E, 4 ꢀ 5E, 4 ꢀ 6E), 7.45-7.48 (2H, m, 2C, 6C); C NMR (75 MHz; CDCl3): ꢀ
44.3 (NH-CH2), 69.9 (P-O-CH2), 70.0 (P-O-CH2), 120.2 (3C, 5C, 3D, 5D), 121.7 (2C, 6C), 128.0 (2E, 6E),
128.5 (3E, 4E, 5E), 128.9 (2D, 6D), 134.9 (1C, 1D), 135.2 (1E), 146.3 (4C), 149.8 (4D), 164.3, 165.1
(CONH2, 4A), 166.2 (6A), 169.6 (2A); MS (ES+) m/z 873 [M+H]+, 895 [M+Na]+.
4-[4-(4-Phosphoryloxybenzylamino)-6-carbamoyl-1,3,5-triazin-2-ylamino]phenyl phosphate (7)
10% Palladium on carbon (75 mg) was added to a solution of compound 27 (153 mg, 0.175 mmol) in
MeOH (50 mL). The mixture was stirred under an hydrogen atmosphere at room temperature
overnight. The catalyst was filtered on two filter papers and washed with a hot MeOH/AcOH 98:2
mixture. Purification by column chromatography on reverse phase silica gel C18 using a water/MeCN
95:5 mixture gave 7 as a yellow solid (39 mg, 43%). Mp 196-199 °C; MS (ES–) m/z 255 [M–2H]2–.

HETEROCYCLES, Vol. 81, No. 4, 2010
881
Tetrasodium 4-[4-(4-phosphonatooxybenzylamino)-6-carbamoyl[1,3,5]triazin-2-ylamino]phenyl
phosphate (29)
A fresh 0.539 M sodium methoxide solution (304 μL, 164 μmol) was added to a suspension of
compound 7 (21 mg, 41.0 μmol) in dry MeOH (3 mL) at 0 °C. The mixture was stirred at 0 °C for 2 h
and the MeOH was removed under reduced pressure to afford 29 as a watersoluble beige powder (24
1
mg, 98%). H NMR (300 MHz; D2O): ꢀ 4.40 (2H, s, NH-CH2), 7.03-7.05 (4H, m, 3C, 5C, 3D, 5D),
13
7.15-7.17 (2H, m, 2D, 6D), 7.27-7.29 (2H, m, 2C, 6C); C NMR (75 MHz; D2O): ꢀ 43.4 (NH-CH2),
120.4, 120.6 (3C, 5C, 3D, 5D), 123.7 (2C, 6C), 128.3 (2D, 6D), 131.7 (1C), 132.0 (1D), 150.8 (4C), 152.8
(4D), 164.3, 164.4 (4A, CONH2), 165.4 (6A), 166.8 (2A); 31P NMR (121 MHz; D2O): ꢀ 0.43; HRMS
(ES–) m/z: calcd for C17H14N6Na4O9P2 576.9991 [M–Na]–, 533.0352 [M–3Na+2H]–; found 576.9979,
533.0339.
ACKNOWLEDGEMENTS
We thank CNRS, Institut Curie and Ministère de la Recherche (ACI “Molécules et Cibles
Thérapeutiques” 2002 N° 02L0521) for financial support. Fondation pour la Recherche Médicale
(FRM) is gratefully acknowledged for a fellowship granted to Caroline Courme.
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