Preparation of a set of 4,5-dihydro-3H-pyrrolo[3,2-d]pyrimidin-4-ones as potential Hsp90 ligands

Article abstract of DOI:10.1016/j.tetlet.2008.07.158

A synthetic route for the preparation of 4,5-dihydro-3H-pyrrolo[3,2-d]pyrimidin-4-ones characterized by a decorated benzyl moiety at different positions of the five-membered ring has been developed, and some compounds have been tested as Hsp90 ligands. One of them displayed IC₅₀ = 50 μM representing an interesting starting point for further investigations.

Full text of DOI:10.1016/j.tetlet.2008.07.158

Tetrahedron Letters 49 (2008) 5965–5967
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Tetrahedron Letters
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Preparation of a set of 4,5-dihydro-3H-pyrrolo[3,2-d]pyrimidin-4-ones
as potential Hsp90 ligands
*
*
Teresa Semeraro, Claudia Mugnaini , Federico Corelli
Dipartimento Farmaco Chimico Tecnologico, Università degli Studi di Siena, Via A. Moro, snc 53100 Siena, Italy
a r t i c l e i n f o
a b s t r a c t
Article history:
A synthetic route for the preparation of 4,5-dihydro-3H-pyrrolo[3,2-d]pyrimidin-4-ones characterized by
Received 13 June 2008
Revised 24 July 2008
Accepted 29 July 2008
Available online 31 July 2008
a decorated benzyl moiety at different positions of the five-membered ring has been developed, and some
compounds have been tested as Hsp90 ligands. One of them displayed IC50 = 50
interesting starting point for further investigations.
lM representing an
Ó 2008 Elsevier Ltd. All rights reserved.
Keywords:
Hsp90 ligands
Pyrrolo[3,2-d]pyrimidin-4-ones
Deazapurines
The 90 kDa heat shock protein (Hsp90) is emerging as an impor-
tant target for cancer chemotherapy because of its role in
chaperoning proteins involved in signal transduction pathways
that control tumor cell proliferation.1–4 Among those disclosed
being able to inhibit Hsp90, the purine (PU) class of inhibitors
(Fig. 1)5–9 represents the most investigated one, exhibiting
biological activities in the nanomolar range and high levels of
selectivity for tumor vs normal cell Hsp90.
Compounds 1a–c were prepared starting from ethyl 3-amino-4-
cyano-1H-pyrrole-2-carboxylate (2) (Scheme 1), easily accessible
by condensation of ethoxymethylenemalononitrile and diethyl
aminomalonate. Reaction of 2 with DMF dimethyl acetal gave the
protected derivative 3, which was alkylated with the appropriate
benzyl halide to give the corresponding benzyl derivatives 4a,b.
MW-aided cyclization of 4a,b in the presence of ammonium
Based on these observations and taking advantage of our expe-
rience in the synthesis of deazaguanine derivatives,10 we planned
to develop a synthetic route to access 4,5-dihydro-3H-pyrrol-
o[3,2-d]pyrimidin-4-ones of general structure 1, characterized by
the presence of a substituted benzyl moiety at different positions
of the pyrrole ring, with the aim of exploring the structure–activity
relationships for this class of compounds (Fig. 1).
H
N
EtOOC
N
H
N
EtOOC
H2N
a
CN
N
90%
CN
3
2
b
R3
R1
O
R2
O
N
NH2
3
HN
4
R1
7
N
6
N 5
5
N
c
N
1N
2
EtOOC
HN
R3
6
X
8
2
R1
N
1
7 R4
N
4
R1
N
3
CN
N
9R2
CN
N
1
PU-class
5a R1 = 4-OMe (78%)
5b R1 = 3,4-methylenedioxy (47%)
4a R1 = 4-OMe (75%)
4b R1 = 3,4-methylenedioxy (82%)
Figure 1. Purine and deazapurine Hsp90 ligands.
* Corresponding authors. Tel.: +39 (0)577 234308 (F.C.); tel.: +39 (0)577 234318
(C.M.).
Scheme 1. Synthesis of compounds 5a,b. Reagents and conditions: (a)
(MeO)2CHNMe2, DCM, rt, 3 h; (b) benzyl halide, TBACl, NaI, 20% NaOH, DCM, rt,
1 h; (c) NH4OH, EtOH, THF, MW, 100 °C, 20 min, sealed tube.
E-mail addresses: mugnaini11@unisi.it (C. Mugnaini), corelli@unisi.it (F. Corelli).
0040-4039/$ - see front matter Ó 2008 Elsevier Ltd. All rights reserved.
doi:10.1016/j.tetlet.2008.07.158

5966
T. Semeraro et al. / Tetrahedron Letters 49 (2008) 5965–5967
O
N
H
O
N
H
N
N
a
R1
EtOOC
HN
a
N
HN
63%
5a,5b
H2N
10
11
CHO
6a R1 = 4-OMe (35%)
6b R1 = 3,4-methylenedioxy (30%)
b
20%
O
N
H
N
b
c
(from 6a)
HN
45%
N
O
OMe
R1
O
N
1f
HN
N
HN
N
Scheme 4. Synthesis of compound 1f. Reagents and conditions: (a) NH@CHNH2
AcOH, EtOH, reflux, 5 h; (b) N-methylallylamine, CH2O, CH3COOH, H2O, 95 °C, 24 h.
N
NH
COOMe
1a R1 = 4-OMe (35%)
1c
presence of formamidine acetate to afford the key intermediates
9a,b. Treatment of 9a,b with the appropriate amine in the presence
of Ti(OiPr)4 gave directly the final amides 1d,e.
Compound 1f was obtained starting from ethyl 3-amino-5-
benzyl-1H-pyrrole-2-carboxylate (10)11 by cyclization with form-
amidine acetate and subsequent Mannich reaction with formalde-
hyde and N-methylallylamine of the deazapurine derivative 11
(Scheme 4).
1b R1 = 3,4-methylenedioxy (40%)
Scheme 2. Synthesis of compounds 1a–c. Reagents and conditions: (a) LiAlH4, THF,
rt, 2 h; (b) allylamine, Na(AcO)3BH, AcOH, DCE, rt, 48 h; (c) Ph3P@CHCOOMe, MeOH,
reflux, 20 h then H2, Pd/C, EtOAc, rt, 4 h.
hydroxide afforded the N5-benzylated pyrrolo[3,2-d]pyrimidines
5a,b functionalized at C7.
Finally, compounds 1g,h were prepared according to Scheme 5.
Aldehyde 12 was condensed with methyl cyanoacetate to give the
intermediate 13. Simultaneous reduction of the double bond and
the ester function by means of NaBH4 afforded the alcohol 14,
which was transformed into the pyrrole derivative 15 by oxidation
to the corresponding aldehyde, reaction with diethyl aminomalo-
nate, and subsequent cyclization in the presence of NaOMe.
Elaboration of the CN group of 5a,b (Scheme 2) to afford alde-
hydes 6a,b failed under different experimental conditions using
selective reducing agents, such as DIBAL, but was surprisingly
accomplished by means of the strong reductant LiAlH4. Subsequent
reductive amination of the aldehydes 6a,b gave the final com-
pounds 1a,b in an acceptable overall yield. Compound 1c was ob-
tained, starting from 6a, through a Wittig olefination using methyl
triphenylphosphoranylideneacetate followed by catalytic hydroge-
nation of the double bond.
A similar approach was followed for the synthesis of com-
pounds 1d,e which were prepared starting from diethyl 3-amino-
1H-2,4-pyrroledicarboxylate (7) (Scheme 3).
Compound 7 was initially alkylated to give the corresponding
benzyl derivatives 8a,b which, in turn, were cyclized in the
Br
Br
MeO
MeO
MeO
MeO
CN
a
O
COOCH3
80%
OMe
OMe
12
13
b
94%
R1
EtOOC
H2N
H
N
H
Br
a
N
EtOOC
OMe
OMe
EtOOC
H2N
N
MeO
MeO
CN
OH
Br
c
H2N
COOEt
55%
COOEt
OMe
OMe
8a R1 = 4-OMe (34%)
7
15
14
8b R1 = 3,5-di-OMe (30%)
e
d
50%
15%
b
O
N
O
N
H
N
H
N
O
HN
H2N
HN
O
N
R1
R1
Br
Br
N
OMe
OMe
c
N
HN
HN
N
OMe
OMe
OMe
OMe
COOEt
CONHR2
1g
1h
9a R1 = 4-OMe (91%)
1d R1 = 4-OMe, R2 = butyl (97%)
9b R1 = 3,5-di-OMe (90%)
1e R1 = 3,5-di-OMe, R2 = allyl (72%)
Scheme 5. Synthesis of compounds 1g,h. Reagents and conditions: (a)
CNCH2COOCH3, piperidine, toluene, reflux, 1 h, (b) NaBH4, EtOH, rt, 1 h; (c) (i)
(COCl)2, DMSO, DIPEA, DCM, À78 °C, 2 h; (ii) NH2CH(COOEt)2ÁHCl, CH3COONa, H2O,
EtOH, rt, 20 h; (iii) NaOMe, MeOH, rt, 3 h; (d) NH@CHNH2ÁAcOH, 2-methoxyethanol,
reflux, 1 h; (e) 50% NH2CN, 50% NaOH, rt, 16 h.
Scheme 3. Synthesis of compounds 1d,e. Reagents and conditions: (a) benzyl
halide, TBACl, NaI, 20% NaOH, DCM, rt, 1 h; (b) NH@CHNH2ÁAcOH, EtOH or 2-
methoxyethanol, reflux, 48 h; (c) R2NH2, Ti(OiPr)4, reflux, 24 h.

T. Semeraro et al. / Tetrahedron Letters 49 (2008) 5965–5967
5967
3. Solit, D. B.; Chiosis, G. Drug Discov. Today 2008, 13, 38–43.
Condensation of 15 with formamidine acetate or cyanamide gave
the target compounds 1g and 1h, respectively.
All the newly synthesized compounds were submitted to a
rHsp90 competitive binding assay in order to evaluate their ability
to bind Hsp90.12 Compounds 1a,b and 1d–f showed
4. Kamal, A.; Kasibhatla, S.; Biamonte, M.; Zhang, H.; Zhang, L.; Lunsgren, K.;
Boehm, M. F.; Burrows, F. J. In Heat Shock Proteins in Cancer; Calderwood, S. K.,
Sherman, M. Y., Ciocca, D. R., Eds.; Springer, 2007; pp 73–92.
5. Chiosis, G.; Kang, Y.; Sun, W. Expert Opin. Drug Discov. 2008, 3, 99–114.
6. Biamonte, M. A.; Shi, J.; Hong, K.; Hurst, D. C.; Zhang, L.; fan, J.; Busch, D. J.;
Karjian, P. L.; Maldonado, A. A.; Sensintaffar, J. L.; Yang, Y.-C.; Kamal, A.; Lough,
R. E.; Lundgren, K.; Burrows, F. J.; Timony, G. A.; Boehm, M. F.; Kasibhatla, S. R. J.
Med. Chem. 2006, 49, 817–828.
7. He, H.; Zatorska, D.; Kim, J.; Aguirre, J.; Llauger, L.; She, Y.; Wu, N.; Immormino,
R. M.; Gewirth, D. T.; Chiosis, G. J. Med. Chem. 2006, 49, 381–390.
8. Kasibhatla, S. R.; Hong, K.; Biamonte, M. A.; Busch, D. J.; Karjian, P. L.;
Sensintaffar, J. L.; Kamal, A.; Lough, R. E.; Brekken, J.; Lundgren, K.; Grecko, R.;
Timony, G. A.; Ran, Y.; Mansfield, R.; Fritz, L. C.; Ulm, E.; Burrows, F. J.; Boehm,
M. F. J. Med. Chem. 2007, 50, 2767–2778.
IC50 > 10,000
compounds of the same series only 1c elicited some activity
(IC50 = 364 M), while 1a and 1d, although bearing the same 4-
lM and thus were considered inactive. Among the
l
OMe benzyl group as 1c, but differing for the nature and length
of the C7 chain, proved to be completely inactive. Interestingly,
the activity was restored with compound 1h13 endowed with
IC50 = 50 lM.
9. Legraverend, M.; Grierson, D. S. Bioorg. Med. Chem. 2006, 14, 3987–
4006.
10. Semeraro, T.; Lossani, A.; Botta, M.; Ghiron, C.; Alvarez, R.; Manetti, F.;
Mugnaini, C.; Valensin, S.; Focher, F.; Corelli, F. J. Med. Chem. 2006, 49, 6037–
6045.
In summary, we have described the synthesis of new 4,5-dihy-
dro-3H-pyrrolo[3,2-d]pyrimidin-4-ones substituted at different
positions of the pentatomic ring with benzyl groups.
Some of the new compounds proved to be able to mimic mem-
bers of the PU class in binding to Hsp90 protein. The synthetic ap-
proach described herein will be exploited for the preparation of
several families of potential Hsp90 ligands.
11. Synthesis of compound 10. Acetic acid (21
aminomalonate hydrochloride (184 mg, 0.84 mmol) and sodium acetate
(71 mg, 0.87 mmol) were added to stirred solution of 3-oxo-4-
lL, 0.37 mmol), diethyl
a
phenylbutyronitrile (200 mg, 1.3 mmol) in dry EtOH (1 mL), and the mixture
was stirred for 2 days, after which it was concentrated under vacuum. The
residue was partitioned between DCM and water, and the organic phase was
dried over Na2SO4. Removal of the solvent afforded a syrup that was dissolved
in an ethanolic solution of EtONa (0.87 mmol, 1.8 mL); after 3 days of stirring,
Acknowledgments
acetic acid (52 lL) was added and EtOH was removed under vacuum. The
residue was dissolved in DCM and washed with NaHCO3. The organic phase
was evaporated to dryness. The crude product was purified by flash
chromatography (eluent: EtOAc/PE, 2/3) to give 10 as a yellow solid (50%
yield); mp 85–86 °C (EtOAc); 1H NMR (200 MHz, CDCl3): d 7.31–7.15 (m, 5H),
5.48 (s, 1H), 4.24 (q, J = 7.1 Hz, 2H), 3.82 (s, 2H), 1.26 (t, J = 7.1 Hz, 3H); MS m/z
We thank Dr. Mario Varasi from Genextra Group S.p.A., Milan
(Italy) for helpful discussions and Dr. Thomas Hesterkamp from
Evotec AG, Hamburg (Germany) for the biological assay. Financial
support from Fondazione Monte dei Paschi di Siena (n 30041,
2007), Ministero dell’Università e della Ricerca (PRIN 2006—Prot.
n 2006030948_002), and Università di Siena (PAR Progetti 2006)
is gratefully acknowledged.
245 (M+H)+; IR (Nujol):
m
3448, 2337, 1673 cmÀ1. Anal. Calcd for C14H16N2O2:
C, 68.83; H, 6.60; N, 11.47. Found: C, 68.98; H, 6.58; N, 11.46.
12. Schilb, A.; Riou, V.; Schoepfer, J.; Ottl, J.; Müller, K.; Chene, P.; Mayr, L. M.;
Filipuzzi, I. J. Biomol. Screen. 2004, 9, 569–577.
13. Analytical data for compound 1h: white solid; mp 289–290 °C; 1H NMR
(200 MHz, Me2SO-d6): d 11.20 (br s, 1H), 10.32 (br s, 1H), 6.86 (s, 1H), 6.82
(s, 1H), 5.79 (s, 2H), 3.81 (s, 2H), 3.74 (s, 3H), 3.71 (s, 3H), 3.66 (s, 3H); MS m/z
References and notes
411 (M+H)+; IR (Nujol):
m
3423, 3134, 2752, 1671, 1629 cmÀ1. Anal. Calcd for
C
16H17BrN4O4: C, 46.96; H, 4.19; N, 13.69. Found: C, 46.75; H, 4.27; N,
1. Blagg, B. S. J.; Kerr, T. D. Med. Res. Rev. 2006, 26, 310–338.
2. Chiosis, G.; Rodina, A.; Moulick, K. Anti-Cancer Agents—Med. Chem. 2006, 6, 1–8.
13.91.