Structure-function studies on a synthetic guanosine receptor that simultaneously binds Watson-Crick and Hoogsteen sites

Article abstract of DOI:10.1021/jo0501689

A series of receptors (11-16) designed to simultaneously bind the Watson-Crick and Hoogsteen sites of guanosine were synthesized, and their binding of guanosine tri-O-pentanoate (32) was probed via ¹H NMR complexation studies in 5% DMSO-d₆-chloroform-d. The guanosine receptors were synthesized with aminonaphthalene or aminoquinoline auxiliary groups tethered to N-4 of cytosine via a methylene or carbonyl group. A structure-function relationship was established allowing energetic contributions made by components of nucleoside analogues to be probed and more general design rules formulated that may guide the development of more efficacious DNA bases.

Full text of DOI:10.1021/jo0501689

VOLUME 70, NUMBER 19
SEPTEMBER 16, 2005
© Copyright 2005 by the American Chemical Society
Structure-Function Studies on a Synthetic Guanosine Receptor
That Simultaneously Binds Watson-Crick and Hoogsteen Sites
Jordan R. Quinn and Steven C. Zimmerman*
Department of Chemistry, University of Illinois at Urbana-Champaign, Urbana, Illinois 61801
sczimmer@uiuc.edu
Received January 26, 2005
A series of receptors (11-16) designed to simultaneously bind the Watson-Crick and Hoogsteen
sites of guanosine were synthesized, and their binding of guanosine tri-O-pentanoate (32) was probed
1
via H NMR complexation studies in 5% DMSO-d6-chloroform-d. The guanosine receptors were
synthesized with aminonaphthalene or aminoquinoline auxiliary groups tethered to N-4 of cytosine
via a methylene or carbonyl group. A structure-function relationship was established allowing
energetic contributions made by components of nucleoside analogues to be probed and more general
design rules formulated that may guide the development of more efficacious DNA bases.
Introduction
forming multiple hydrogen-bonding contacts, a number
of these receptors are able to complex nucleobases in
competitive media, including some in water. However,
there are few accounts of nucleoside analogues, particu-
larly those incorporated into oligonucleotides, having
enhanced hydrogen-bonding capabilities.6
The high specificity with which oligonucleotides form
duplexes has been exploited in areas ranging from
nanotechnology1 to genetic medicines.2 In an effort to
improve the performance of some of these DNA- and
RNA-based systems, considerable effort has focused on
novel nucleotides with enhanced hydrogen-bonding and
base-stacking abilities.3 Considerably more attention has
been given to increasing base stacking especially through
a dangling-end base. In contrast, many synthetic recep-
tors for nucleobases4 have expanded their hydrogen-
bonding contacts beyond the Watson-Crick edge.5 By
Umezawa et al.7 reported that 1, a cytosine analogue
with an 8-ureido-2-naphthoyl group attached at N-4,
complexes guanine (G) with five hydrogen bonds and
(4) Selected reviews: Fan, E.; Hamilton, A. D. In Crystal Engineer-
ing: The Design and Application of Functional Solids; Seddon, K. R.,
Zaworotko, M., Eds.; Kluwer: Dordrecht, 1999; pp 357-369. Hamilton,
A. D.; Fan, E.; Van Arman, S.; Vicent, C.; Tellado, F. G.; Geib, S. J.
Supramol. Chem. 1993, 1, 247-252. Rebek, J., Jr. Top. Curr. Chem.
1988, 149, 189-210. Wintner, E. A.; Conn, M. M.; Rebek, J., Jr. Acc.
Chem. Res. 1994, 27, 198-203. Hartley, J. H.; James, T. D.; Ward, C.
J. J. Chem. Soc., Perkin Trans. 1 2000, 19, 3155-3184. Beer, P. D.;
Schmitt, P. Curr. Opin. Chem. Biol. 1997, 1, 475-482. Sessler, J. L.;
Sansom, P. I.; Andrievsky, A.; Kral, V. In Supramolecular Chemistry
of Anions; Bianchi, A., Bowman-James, K., Garcia-Espana, E., Eds.;
Wiley-VCH: New York, 1997; pp 355-419. Davis, A. P. In Supra-
molecular Science: Where It Is and Where It Is Going; Ungaro, R.,
Dalcanale, E., Eds.; Kluwer: Dordrecht, 1999; pp 125-146. Zimmer-
man, S. C.; Corbin, P. S. Struct. Bonding 2000, 96, 63-94.
(1) (a) Seeman, N. C. Nature 2003, 421, 427-431. (b) Storhoff, J.
J.; Mirkin, C. A. Chem. Rev. 1999, 99, 1849-1862. (c) Niemeyer, C.
M. Angew. Chem., Int. Ed. Engl. 1997, 36, 585-587.
(2) (a) Scherer, L. J.; Rossi, J. J. Nature Biotechnol. 2003, 21, 1457-
1465. (b) Therapeutic Oligonucleotides: Antisense, RNAi, Triple-Helix,
Enhancer-Decoy, CpG, and DNA Chips; Cho-Chung, Y. S., Gewirtz,
A. M., Stein, C. A., Eds. Ann. N.Y. Acad. Sci. 2003, 1002, 274 pp.
(3) Verma, S.; Eckstein, F. Annu. Rev. Biochem. 1998, 67, 99-134.
Kool, E. T. Chem. Rev. 1997, 97, 1473-1487. Herdewijn, P. Antisense
Nucleic A. 2000, 10, 297-310. Luyten, I.; Herdewijn, P. Eur. J. Med.
Chem. 1998, 33, 515-576.
10.1021/jo0501689 CCC: $30.25 © 2005 American Chemical Society
Published on Web 05/12/2005
J. Org. Chem. 2005, 70, 7459-7467
7459

Quinn and Zimmerman
SCHEME 1
SCHEME 2
an association constant (Kassoc) ) 170 M-1 in 4:1 (v/v)
CDCl3/DMSO-d6 (Scheme 1). To our knowledge, this
nucleoside analogue has not been incorporated into
oligonucleotides. In this regard, the N-4 amido group
presents a special challenge because it may lack sufficient
stability to withstand the standard DNA deprotection
conditions. Herein, we explore analogues of Umezawa’s
receptor designed both to be more stable to hydrolysis
and to probe the importance of the various associated
structural features in 1. By examining the effect of
structural modifications on complex stability within
nucleoside-synthetic receptor complexes it was hoped
that the energetic contributions made by components of
nucleoside analogues would be probed and ultimately
more general design rules formulated.
Results and Discussion
Receptor Design. As noted above, one of the appeal-
ing features of 1 is the tethering of the cytosine nucleus
to a recognition unit capable of hydrogen bonding to the
Hoogsteen site of guanosine. Modeling of DNA oligo-
nucleotides further suggested that runs of base 1 (amino
in place of ureido group, vide infra) in one strand would
position the naphthalene units so that they can stack
efficiently in the major groove. Modeling also indicated
that a similar complex could be formed analogously with
2. N-4-Substituted cytidines exist in either the imino or
amino tautomeric form (Scheme 2). Both acetyl and alkyl
substitution at N-4 produce the amino tautomer exclu-
sively, provided C-5 is unsubstituted.8,9 In addition, N-4
monosubstitution gives rise to syn and anti rotamers.10,11
In N,N′-1,4-dimethylcytosine the syn form is energetically
favored by 1.6 kcal/mol at -19 °C in DMF-d7.10 Steric
repulsion between H-5 and the alkyl substituent is
proposed to lead to the observed preference for the syn
(5) Selected examples: Adrian, J. C.; Wilcox, C. S. J. Am. Chem.
Soc. 1989, 111, 8055-8057. Seel, C.; Vo¨gtle, F. Angew. Chem., Int. Ed.
Engl. 1991, 30, 442-444. Deslongchamps, G.; Gala´n, A.; de Mendoza,
J.; Rebek, J., Jr. Angew. Chem., Int. Ed. Engl. 1992, 31, 61-63. Gu¨ther,
R.; Nieger, M.; Vo¨gtle, F. Angew. Chem., Int. Ed. Engl. 1993, 32, 601-
603. Kato, Y.; Conn, M. M.; Rebek, J., Jr. J. Am. Chem. Soc. 1994,
116, 3279-3284. Bell, T. W.; Hou, Z.; Zimmerman, S. C.; Thiessen, P.
A. Angew. Chem., Int. Ed. Engl. 1995, 34, 2163-2165. Lonergan, D.
G.; Deslongchamps, G.; Toma`s, S. Tetrahedron Lett. 1998, 39, 7861-
7864. Alkorta, I.; Elguero, J.; Goswami, S.; Mukherjee, R. J. Chem.
Soc., Perkin Trans. 2 2002, 894-898. Faraoni, R.; Blanzat, M.; Kubicek,
S.; Braun, C.; Schweizer, W. B.; Gramlich, V.; Diederich, F. Org.
Biomol. Chem. 2004, 2, 1962-1964.
(6) A notable exception is the G-clamp, which forms two hydrogen
bonds to the Hoogsteen side of G in addition to the standard three
Watson-Crick hydrogen bonds: Lin, K.-Y.; Matteucci, M. D. J. Am.
Chem. Soc. 1998, 120, 8531-8532. Flanagan, W. M.; Wolf, J. J.; Olson,
P.; Grant, D.; Lin, K.-Y.; Wagner, R. W.; Matteucci, M. D. Proc. Natl.
Acad. Sci. U.S.A. 1999, 96, 3513-3518. Wilds, C. J.; Maier, M. A.;
Tereshko, V.; Manoharan, M.; Egli, M. Angew. Chem., Int. Ed. 2002,
41, 115-117. Wilds, C. J.; Maier, M. A.; Manoharan, M.; Egli, M. Helv.
Chim. Acta 2003, 86, 966-978.
(7) Amemiya, S.; Bu¨hlmann, P.; Umezawa, Y. Chem. Commun.
1997, 1027-1028. Tohda, K.; Amemiya, S.; Ohki, T.; Nagahora, S.;
Tanaka, S.; Bu¨hlmann, P.; Umezawa, Y. Isr. J. Chem. 1997, 37, 267-
275.
(8) (a) Busson, R.; Kerremans, L.; Van Aerschot, A.; Peeters, M.;
Blaton, N.; Herdewijn, P. Nucleosides Nucleotides 1999, 18, 1079-
1082. (b) Wada, T.; Kobori, A.; Kawahara, S.; Sekine, M. Tetrahedron
Lett. 1998, 39, 6907-6910.
7460 J. Org. Chem., Vol. 70, No. 19, 2005

Structure-Function Studies on a Synthetic Guanosine Receptor
such as 1 would represent a significant synthetic chal-
lenge, but there was interest in the quinoline analogue
of 1. Hunter and co-workers reported a p-quinone recep-
tor with a binding pocket containing 4 that was superior
to its analogue with 3.14c The increased stability may
originate in a degree of preorganization owing to the
intramolecular hydrogen bonding between the amido NH
groups and pyridine nitrogen of 4.13,21 However, electro-
static repulsion of the pyridine nitrogen with the gua-
nosine carbonyl oxygen might offset the energy gains
from preorganization.15
Considering the design issues outlined above and the
goal of establishing a structure activity relationship that
would produce design rules for tethered bases such as 1,
synthetic receptors 9-16 were synthesized and model
complexation studies performed with cytosine in chloro-
form solution.
FIGURE 1. Structural diversity in carbonyl complexation
units. (a) General motif with NH groups oriented toward
carbonyl “rabbit ears.” Selected examples: (b) isophthaloyl and
pyridine-2,6-dicarboxamide, (c) and (d) chromenone, (e) pyrido-
dipyrimidinedione, (f) pyridobisindole.
Synthesis of Nucleoside Receptors. Cytidine ana-
logues with N-4-amido and alkyl substituents are shown
in Chart 1. Acetylcytosine 9 was prepared by treating
2′-deoxy-3′,5′-bis-O-(tert-butyldimethylsilyl)cytidine (17)
with acetic anhydride, whereas the corresponding benzoyl
derivative 10 was prepared by treating 17 with benzoyl
chloride. Analogous to the procedure reported by
Umezawa,7b amido cytidines 11-13 were synthesized by
coupling of TBDMS-protected deoxycytidine 17 with
carboxylic acids 18, 19, and 20,7b respectively, using
1-ethyl-3-(3′-dimethylaminopropyl)carbodiimide hydro-
chloride (EDCI) (Scheme 3). Naphthalene carboxylic acids
19 and 20 were available from 2-naphthalene carboxylic
acid in three and four steps, respectively.7b Quinoline
carboxylic acid 18 was prepared from commercially
available 2-methylquinoline by tribromination and hy-
drolysis.22 Thus, coupling of 17 and 20 directly gave 13
in 81% yield, whereas 17 coupled with 18 and 19 to give
rotamer in N-4-alkylated cytosines.11 Syn-anti isomer-
ization has not been reported for N-4-acylcytidines which
may reflect rapid rotation on the NMR time-scale or more
likely a bias toward the anti form as a result of the
interaction between the carbonyl group and N-3 shown
in Scheme 2 and a favorable CH‚‚‚O contact with H-5.8b,12
In considering structural modifications to guanosine
receptor 1 attention was focused on: (1) the group linking
the cytosine nucleus to the major groove-binding naph-
thalene unit, (2) the carbonyl binding cleft formed by the
amide and urea groups, and (3) the Hoogsteen pairing
unit, the urea group itself. As indicated above, there was
considerable interest in the methylene linker because of
its expected stability during oligonucleotide synthesis.
With regard to the carbonyl binding cleft, there is a
remarkable diversity in the units designed to spatially
position two N-H groups to hydrogen bond the “rabbit
ear” lone pairs of a carbonyl group. As seen in Figure 1,
the simplest is 3 and 4, pioneered by Hamilton and co-
workers in the context of receptors for barbiturates,13 and
extended to hosts for quinone,14 diamides,15 and urea16
and a variety of other applications.17 More rigid ana-
logues of 3 and 4 that provide a similar spatial arrange-
ment of two NH groups are seen in Mora´n’s chromenone-
based receptors 5 (6),18 our dimethylacetamide complexing
tecton 7,19 and Thummel’s urea binder 8.20 Integrating
a highly preorganized unit such as 6-8 into a receptor
(14) (a) Carver, F. J.; Hunter, C. A.; Shannon, R. J. J. Chem. Soc.,
Chem. Commun. 1994, 1277-1280. (b) Brooksby, P. A.; Hunter, C. A.;
McQuillan, A. J.; Purvis, D. H.; Rowan, A. E.; Shannon, R. J.; Walsh,
R. Angew. Chem., Int. Ed. Engl. 1994, 33, 2489-2491. (c) Hunter, C.
A.; Pervis, D. H. Angew. Chem., Int. Ed. Engl. 1992, 31, 792-795.
(15) Chang, S.-Y.; Kim, H. S.; Chang, K.-J.; Jeong, K.-S. Org. Lett.
2004, 6, 181-184.
(16) Goswami, S.; Mukherjee, R. Tetrahedron Lett. 1997, 38, 1619-
1622.
(17) For example: Lukin, O.; Recker, J.; Bo¨hmer, A.; Mu¨ller, W. M.;
Kubota, T.; Okamoto, Y.; Nieger, M.; Fro¨hlich, R.; Vo¨gtle, F. Angew.
Chem., Int. Ed. 2003, 42, 442-445. Kidd, T. J.; Leigh, D. A.; Wilson,
A. J. J. Am. Chem. Soc. 1999, 121, 1599-1600.
(18) Raposo, C.; Crego, M.; Mussons, M. L.; Caballero, M. C.; Mora´n,
J. R. Tetrahedron Lett. 1994, 35, 3409-3410. Crego, M.; Raposo, C.;
Caballero, M. C.; Garc´ıa, E.; Saez, J. G.; Mora´n, J. R. Tetrahedron Lett.
1992, 33, 7437-7440. Crego, M.; Partearroyo, A.; Raposo, C.; Mussons,
M. L.; Lo´pez, J. L.; Alca´zar, V.; Mora´n, J. R. Tetrahedron Lett. 1994,
35, 1435-1438. Martin, M.; Raposo, C.; Almaraz, M.; Crego, M.;
Caballero, C.; Grande, M.; Mora´n, J. R. Angew. Chem., Int. Ed. Engl.
1996, 35, 2386-2388.
(9) Gas-phase calculations by Wallis et al. suggest that N-4 benzoyl
cytosine (CBz) prefers the intramolecularly hydrogen bonded syn-imino
form (Wallis, M. P.; Schwalbe, C. H.; Fraser, W. Nucleosides Nucleo-
tides 1997, 16, 2053-2068); however, solution data (ref 8a) and solid-
state structures indicate that the anti-amino form dominates. Selected
examples of crystal structures of CBz derivatives: Scho¨ning, K.-U.;
Scholz, P.; Wu, X.; Guntha, S.; Delgado, G.; Krishnamurthy, R.;
Eschenmoser, A. Helv. Chim. Acta 2002, 85, 4111-4153. Rennenberg,
D.; Leumann, C. J. J. Am. Chem. Soc. 2002, 124, 5993-6002. Woz´niak,
L. A.; Wieczorek, M.; Pyzowski, J.; Majzner, W.; Stec, W. J. J. Org.
Chem. 1998, 63, 5395-5402.
(19) Petersen, P. M.; Wu, W.; Fenlon, E. E.; Kim, S.; Zimmerman,
S. C. Bioorg. Med. Chem. 1996, 4, 1107-1112.
(20) van Straaten-Nijenhuis, W. F.; de Jong, F.; Reinhoudt, D. N.;
Thummel, R. P.; Bell, T. W.; Liu, J. J. Membrane Sci. 1993, 82, 277-
283. Hegde, V.; Hung, C.-Y.; Madhukar, P.; Cunningham, R.; Ho¨pfner,
T.; Thummel, R. P. J. Am. Chem. Soc. 1993, 115, 872-878. Hegde, V.;
Madhukar, P.; Madura, J. D.; Thummel, R. P. J. Am. Chem. Soc. 1990,
112, 4549-4550.
(10) Shoup, R. R.; Miles, H. T.; Becker, E. D. J. Phys. Chem. 1972,
76, 64-70.
(11) Engel, J. D.; von Hippel, P. H. Biochemistry 1974, 13, 4143-
4158.
(12) Wada, T.; Kobori, A.; Kawahara, S.; Sekine, M. Eur. J. Org.
Chem. 2001, 4583-4593. Parthasarathy, R.; Ginell, S. L.; De, N. C.;
Chheda, G. B. Biochem. Biophys. Res. Commun. 1978, 83, 657-663.
(13) Chang, S.-K.; Hamilton, A. D. J. Am. Chem. Soc. 1988, 110,
1318-1319. Chang, S.-K.; Van Engen, D.; Fan, E.; Hamilton, A. D. J.
Am. Chem. Soc. 1991, 113, 7640-7645. Valenta, J. N.; Dixon, R. P.;
Hamilton, A. D.; Weber, S. G. Anal. Chem. 1994, 66, 2397-2403.
(21) Geib, S. J.; Hirst, S. C.; Vincent, C.; Hamilton, A. D. J. Chem.
Soc., Chem. Soc. 1991, 1283-1285. Hamuro, Y.; Geib, S. J.; Hamilton,
A. D. J. Am. Chem. Soc. 1997, 119, 10587-10593. Lo´pez de la Paz,
M.; Ellis, G.; Penade´s, S.; Vicent, C. Tetrahedron Lett. 1997, 38, 1659-
1662. Crisp, G. T.; Jiang, Y.-L. Tetrahedron 1999, 55, 549-560.
(22) Hammick, D. L. J. Chem. Soc. 1926, 1302-1304. Roth, R.;
Erlenmeyer, H. Helv. Chim. Acta 1954, 37, 1064-1068.
J. Org. Chem, Vol. 70, No. 19, 2005 7461

Quinn and Zimmerman
CHART 1
SCHEME 4
SCHEME 5
SCHEME 6
SCHEME 3
sium cyanide and benzoyl chloride gave quinoline-2-
carbonitrile 27 and 8-nitroquinoline-2-carbonitrile 28.
The 2,8-substitution of carbonitrile 28 was confirmed by
single-crystal X-ray analysis (see the Supporting Infor-
mation). Reduction of the cyano group in 27 and 28
produced amines 23 and 24, respectively. Uracil 29 was
activated for nucleophilic substitution by conversion into
triazolylpyrimidinone 30.24 Reaction with amines 23 and
24 at 100 °C in DMF produced nucleoside analogues 14
and 15 (Scheme 5). Because of the difficulty in preparing
differentially substituted 1,7-naphthalenes, compound 16
was synthesized by reduction of 22. Thus, the amide
group was reduced with borane-diisopropylethylamine
and the nitro group subsequently reduced by hydrogena-
tion (Scheme 6).
nitro intermediates 21 and 22, which were then reduced
to 11 and 12, respectively.
N-4-Alkylated cytidines shown in Chart 1 were pre-
pared either by nucleophilic substitution of an activated
uracil with the appropriate amine or by reduction of an
N-4-amidocytidine (vide infra). The synthesis of the
aminomethylquinoline auxiliary groups 23 and 24 is
outlined in Scheme 4. Both began with oxidation of
quinoline to quinoline N-oxide (25) followed by nitration
according to Yokoyama et al.23 to give 26 and the 5-nitro
(minor) regioisomer. Treatment of 25 and 26 with potas-
Complexation Studies in Organic Solvents. Initial
studies focused on establishing intermolecular contact
between receptors 11-16 and guanosine tri-O-pen-
tanoate (32). It is well established that NH protons
experience large downfield shifts in their 1H NMR spectra
(23) Yokoyama, A.; Ohwada, T.; Saito, S.; Shudo, K. Chem. Pharm.
Bull. 1997, 45, 279-283.
(24) For related derivatives, see: Sung, W. L. J. Org. Chem. 1982,
47, 3623-3628.
7462 J. Org. Chem., Vol. 70, No. 19, 2005

Structure-Function Studies on a Synthetic Guanosine Receptor
TABLE 1. Association Constants (Kassoc) Measured for
Receptor Complexes with G′a
Kassoc
(M-1
-∆G°
-∆∆G°
298
complex
)
(kcal m2o9l8-1
)
(kcal mol-1
)
9‚G′
1610
210
4.3
3.1
3.2
4.6
4.8
3.0
3.2
3.5
4.4
0.1
1.3
10‚G′
11‚G′
12‚G′
13‚G′
14‚G′
15‚G′
16‚G′
17‚G′
230
1.2
2530
4070
170
-0.2
-0.4
1.4
250
1.2
440
0.9
2040
0.0
a The association constants are typically averages of two experi-
ments in 5% DMSO-d6/CDCl3 at 20 °C in which the values agreed
within 15%.
in the magnitude of the complexation shifts in Figure 2
may reflect differences both in the uncomplexed and
complexed receptor structures. A reverse titration was
performed by holding guanosine at a fixed concentration
between 0.1 and 0.3 mM in 5% (v/v) DMSO-d6/chloro-
form-d and adding receptors 11-16. Each receptor
induced downfield shifts in NH-1 and CH-8 of G′. These
resonances were often visible throughout the titration;
however, the NH-2 group usually broadened into the
baseline with relatively few equivalents of added recep-
tor. Overall, the chemical shift changes observed in both
G′ and receptors 10-16 indicate Watson-Crick hydrogen
bonding and, minimally, one or two additional hydrogen
bonds to the Hoogsteen edge of G′ from the pendant
amino or ureido group, respectively.
1
Quantitative H NMR complexation studies were ini-
tially performed in chloroform-d but ultimately in 5% (v/
v) DMSO-d6/chloroform-d to more readily measure and
FIGURE 2. (a) Chemical shift change (red: upfield, black:
downfield) for protons observed for 0.3 mM solution of receptor
in the presence of 5 equiv of 32 (G′). In the case of 16, shifts
represent ∆δmax values from a titration with 32 (0-19.2 mM).
Protons without shifts listed were obscured. An asterisk (*)
indicates protons whose assignments might be switched. See
text for an explanation of the red dot. (b) Structure of
guanosine derivative 32. All measurements were made in
CDCl3 at 20 °C.
compare binding constants. Association constants (Kassoc
)
were determined by monitoring the downfield shifts of
the NH protons on 32 (G′) with an increase in the
concentration of the receptor 11-16. Dilution of receptors
11-16 monitored by 1H NMR revealed concentration
independent chemical shifts indicating that receptor
dimerization could be ignored in determination of Kassoc
values. Guanosine is well-known to form dimers and
higher order aggregates26,27 but at the concentrations
used in this study the Kassoc values were negligibly
affected.
The Kassoc values are collected in Table 1. Perhaps the
most striking comparison is between the 17‚32 (C‚G′)
base pair and the 13‚G′ complex. Despite the latter
having as many as four additional hydrogen bonds to the
Hoogsteen side of G′, it is more stable by only 0.4 kcal
mol-1. As indicated above, the amido group could affect
the amino-imino tautomeric equilibrium or it might
lower the pairing strength if the syn conformation is
significantly populated (Figure 1). However, the 4-amido
group on the cytosine unit does not by itself present any
when engaged in hydrogen bonds. For example, upon
addition of 5 equiv of 32 to a 0.3 mM solution of cytosine
derivative 17, its NH2 group shifted downfield by ca. 4.2
ppm (Figure 2 shows representative examples). Under
the same conditions, similar although less dramatic shifts
were seen in the 4-NH groups of 4-benzoylcytosine 10,
Umezawa’s receptor 1, and its simpler analogues 11 and
16. The amino group on the quinoline unit in 11 and both
urea NH groups on receptor 1 showed significant down-
field shifts consistent with hydrogen bonding to the
Hoogsteen side of 32. The amino group in 16 was
obscured during its titration and could not be followed.
In general, most of the protons not engaged in hydro-
gen bonding showed only small upfield or downfield
shifts. Protons residing in positions labeled with a red
dot in Figure 1 become localized in the deshielding cone
of a carbonyl if complexes such as 1‚G are formed. If
alternative conformations are significantly populated
prior to complexation, then significant (ca. 0.1-0.4 ppm)
downfield shifts can be expected after binding to 32.25 In
a few cases, downfield shifts of this magnitude are seen
but in others it appears that the receptor is largely
preorganized. In comparing receptors, small differences
(25) Zimmerman, S. C.; Schmitt, P. J. Am. Chem. Soc. 1995, 117,
10769-10770. Mertz, E.; Mattei, S.; Zimmerman, S. C. Org. Lett. 2000,
2, 2931-2934. Mertz, E.; Mattei, S.; Zimmerman, S. C. Bioorg. Med.
Chem. 2004, 12, 1517-1526.
(26) Williams, L. D.; Chawla, B.; Shaw, B. R. Biopolymers 1987, 26,
591-603. Williams, N. G.; Williams, L. D.; Shaw, B. R. J. Am. Chem.
Soc. 1989, 111, 7205-7209.
(27) The self-association constant for guanosine could not be deter-
mined in this solvent system because nonlinear regression analysis
did not converge on a solution.
J. Org. Chem, Vol. 70, No. 19, 2005 7463

Quinn and Zimmerman
cost to binding as the 9‚G′ complex is just 0.1 kcal mol-1
less stable, a value within experimental error. This is
consistent with previous work by Sekine and co-workers
with oligonucleotides wherein 4-N-acetylcytosine paired
with guanosine as well as did cytosine. These authors
further noted that the imino tautomer is significantly less
stable that the amino tautomers. It is also true that the
amido group can alter the strength of the hydrogen bond
donor and acceptor sites along the Watson-Crick edge.
Acylation makes the cytidine N-4-H a better hydrogen
bond donor; however, the electron-withdrawing nature
of the amido group decreases the hydrogen bond acceptor
ability of N3 and O2. Thus, these electronic effects
partially offset.
In contrast to the small effect of the acetyl group in 9,
the N-4-benzoyl group in 10 significantly depresses the
complex stability (Kassoc ) 210 M-1, ∆∆G° ) 1.3 kcal
298
mol-1). Molecular modeling indicates a close contact
between the guanosine carbonyl group and the H2 proton
of the phenyl group, but it is unclear if steric repulsion
alone can account for the entire drop in stability or
whether electronic or conformational effects contribute.
In comparing the quinoline system 11 to receptor 10, the
proton in question is no longer present and there is the
possibility of additional hydrogen bonding from the
pendant amino group. However, the Kassoc ) 230 M-1 for
11‚G′ is essentially unchanged, suggesting an unfavor-
able electrostatic interaction between the quinoline ni-
trogen lone pair and the guanosine carbonyl group (vide
supra).
One of the key goals of this investigation was to
determine whether the added flexibility and, particularly,
the preferred syn orientation of the more robust meth-
ylene linker in 14-16 would significantly compromise its
complexation efficiency. The stability of their complexes
with G′ suggest that indeed the energy price for adopting
the required anti orientation is high. Whatever the origin
of the weak binding shown by benzoyl derivative 10, it
is clear that the pendant hydrogen bonding group in 1
(12 and 13) compensates to a significant extent. How
much do these additional Hoogsteen-side hydrogen bonds
contribute? In the series 10 f 12 f 13, the Kassoc values
increase from 210 M-1 f 2530 M-1 f 4070 M-1 reflecting
the progression from no hydrogen bonding functionality
to an amino group to a ureido group. Although the ureido
group has been shown to be effective in Hoogsteen-side
recognition of guanosine,26 its advantage in 13 relative
to the amino group in 12 is only 0.2 kcal mol-1 and just
outside of experimental error.
Molecular Modeling. Preliminary molecular model-
ing provides an explanation for the similar stability of
G′ complexes observed for 12 and 13. In the 12‚G′
complex, the lengths of the Watson-Crick hydrogen
bonds (labeled a-c in Figure 3a) are close to those
observed in the X-ray structure of GC base pairs.28 The
average difference in heteroatom to heteroatom distance
is <3%, and whereas two receptor-ligand hydrogen
bonds are longer, one is shorter. This same minimum
found in the modeling is very nearly planar. The ad-
ditional, Hoogsteen-side hydrogen bond is close to being
linear (NsHsO angle ) 175.9°), and its length (d, 3.06
FIGURE 3. MMFF- and HF/3-21G*-minimized structure of
(a) complex 12‚G and (b) complex 13‚G. In both cases, the
ribose has been replaced by a proton or methyl group.
Hydrogen-bonding distances are shown between heteroatoms.
Parenthetical values shown in (a) are for the G‚C base pair.
The program Spartan was used.
Å) is only slight longer than the a-c lengths. In contrast,
all of the hydrogen bond lengths within the 13‚G′ complex
are longer and the complex adopts a marked nonplanar
conformation to accommodate two additional hydrogen
bonds between the ureido group and the Hoogsteen edge
of G. These hydrogen bonds to N-7 and O-6 of G are long
and further bent from linearity with an NsHsN angle
) 173.2° and NsHsO angle ) 164.9°. More important
than the NsHsN bond angle is the fact that the ureido
N-H group is directed toward the π-system more than
to the N-7 lone pair. Thus, this ureido N-H bond is
rotated 33.2° out of the plane of the G aromatic system.
Although each ureido N-H group of 13 may form a
bifurcated hydrogen bond to N-7 and O-6 of G for a total
of four Hoogsteen-side hydrogen bonds (see Scheme 1),
the nonplanar geometry of the complex makes these
distances (H‚‚‚N distance ) 3.18 Å, H‚‚‚O distance ) 3.26
Å) too long to be considered as traditional hydrogen
bonds. The overall picture that emerges from the model-
ing, and particularly the data shown in Figure 3, is that
significant reorganization is needed for receptor 13 to
form five hydrogen bonds to G.
(28) Rosenberg, J. M.; Seeman, N. C.; Day, R. O.; Rich, A. J. Mol.
Biol. 1976, 104, 145-167.
7464 J. Org. Chem., Vol. 70, No. 19, 2005

Structure-Function Studies on a Synthetic Guanosine Receptor
-5.0, -5.5, -5.6; [R]21D ) +44.9 (c ) 0.55, CHCl3); ESI-HRMS
calcd for (C23H43N3O5Si2‚H)+ 498.2820, found 498.2816. Anal.
Calcd for C23H43N3O5Si2: C, 55.50; H, 8.71; N, 8.44. Found:
C, 55.26; H, 8.74; N, 8.41.
Conclusions
The structure-activity relationship examined here
provides a number of general design principles that may
be applied to nucleoside recognition as well as some
immediate guidance on how to develop novel nucleosides
that will show enhanced recognition of guanosine in
duplex DNA. Specifically, in considering hydrogen bond-
ing alone it is not easy to surpass the base-pairing ability
of the natural bases. Receptor 13, an analogue of the
Umezawa receptor 1, and receptor 12 bind G′ in 5% (v/
v) DMSO-d6/chloroform-d with free energies that are just
0.4 and 0.2 kcal mol-1 more favorable, respectively, than
C (17). However, the stability of the duplex DNA is
heavily dependent on aromatic stacking, a feature that
is absent in these model studies of base pairing.
4-N-Benzoyl-2′-deoxy-3′,5′-bis-O-(tert-butyldimethyl-
silyl)cytidine (10). To a solution of 104 mg (0.229 mmol) of
17 in 0.5 mL of pyridine was added 50 µL (60 mg, 0.43 mmol)
of benzoyl chloride dropwise at room temperature. The result-
ing suspension was stirred for 10 min and then concentrated
in vacuo, dissolved in 25 mL of CH2Cl2, and successively
washed with 15 mL of saturated aqueous NaHCO3 and 15 mL
of brine. The organic layer was dried over MgSO4, filtered,
concentrated in vacuo, and purifed by column chromatography
(SiO2, Rf ) 0.3, 5% MeOH/CH2Cl2) to give 73 mg (57%) of a
white foam spectroscopically similar to published data.30 Anal.
Calcd for C28H45N3O5Si2: C, 60.07; H, 8.10; N, 7.51. Found:
C, 59.97; H, 8.19; N, 7.49.
4-N-(8-Aminoquinolin-2-ylformyl)-2′-deoxy-3′,5′-bis-O-
(tert-butyldimethylsilyl)cytidine (11). A mixture of 144 mg
(0.220 mmol) of 21 and 6 mL of 1:1 (v/v) MeOH/EtOAc was
hydrogenated at atmospheric pressure for 4 h over 75 mg of
Pd/C. The mixture was filtered through Celite and the Celite
washed with 2 × 15 mL of EtOAc. The filtrate was concen-
trated in vacuo and purified by column chromatography (SiO2,
Rf ) 0.35, 5% MeOH/CH2Cl2). The product was precipitated
from 3 mL of 10:1 pentane/EtOAc to give 71 mg (52%) of an
orange powder: 1H NMR (500 MHz, DMSO-d6) δ 11.67 (s, 1H),
8.35 (d, 1H, J ) 8.5), 8.32 (d, 1H, J ) 7.6), 8.14 (d, 1H, J )
8.5), 7.53 (d, 1H, J ) 7.4), 7.41 (t, 1H, J ) 7.8), 7.06 (d, 1H, J
) 8.1), 6.88 (d, 1H, J ) 7.5), 6.87 (br s, 2H), 6.14 (t, 1H, J )
5.9), 4.40 (dt, 1H, J ) 5.0, 4.8), 3.91 (dt, 1H, J ) 3.7, 3.4), 3.86
(dd, 1H, J ) 11.5, 3.9), 3.76 (dd, 1H, J ) 11.5, 3.1), 2.35 (dt,
1H, J ) 12.6, 6.0), 2.21 (dt, 1H, J ) 13.1, 6.0), 0.90 (s, 9H),
0.87 (s, 9H), 0.104 (s, 3H), 0.098 (s, 3H), 0.082 (s, 6H); 13C NMR
(125 MHz, DMSO-d6) δ 164.9, 162.6, 154.5, 147.3, 144.6, 144.6,
137.6, 135.5, 130.6, 130.3, 118.9, 112.4, 109.2, 96.1, 87.2, 86.0,
70.8, 61.9, 40.9, 25.8, 25.7, 18.0, 17.7, -4.8, -5.0, -5.5, -5.6;
In considering the possibility of incorporating receptors
such as 9-16 into oligonucleotides to test the importance
of stacking the tethered aromatic system in the DNA
major groove, the results above provide the following
guidance. The additional hydrogen bonding provided by
12 and 13 should enhance the selectivity of these bases
for G, however, the highly nonplanar complex seen with
13 suggests that it will not be accommodated well into
duplex DNA. In general, the amide linker group appears
to be the best candidate despite the difficulty in incor-
porating such nucleosides into DNA as a result of their
hydrolytic lability. Although the more robust methylene
linker groups produced very poor binders of G it is
possible that the energy gained from aromatic stacking
might offset the energy price required to shift these bases
from the syn to anti form (Scheme 2). Efforts to test some
of these novel nucleoside-based receptors in duplex DNA
are underway and will be reported in due course.
[R]21 ) +53.4 (CHCl3, c ) 0.51); ESI-HRMS calcd for
D
(C31H47N5O5Si2‚H)+ 626.3194, found 626.3164. Anal. Calcd for
C31H47N5O5Si2‚H2O: C, 57.82; H, 7.67; N, 10.88. Found: C,
57.85; H, 7.34; N, 10.62.
Experimental Section
1H NMR Binding Studies. CDCl3 was distilled from CaCl2,
passed through a column of activated (flame-dried) basic
alumina, and stored over 4 Å molecular sieves. DMSO-d6 was
dried over 4 Å molecular sieves. NMR tubes and volumetric
flasks were dried in a 110 °C oven and cooled in a desiccator
prior to sample preparation. Samples (8-12) were prepared
by adding aliquots of stock solutions of guanosine and receptor
via syringe directly into NMR tubes and diluting to 0.700 mL.
Spectra were acquired with a 500 MHz spectrometer using
temperature control at 20 °C. Data were fit to a 1:1 binding
isotherm using standard methods.29
4-N-Acetyl-2′-deoxy-3′,5′-bis-O-(tert-butyldimethylsilyl)-
cytidine (9). A solution of 113 mg (0.249 mmol) of 17 and 0.5
mL (5.3 mmol) of acetic anhydride in 1.5 mL of pyridine was
stirred at room temperature for 20 h and then concentrated
in vacuo. The residue was dissolved in 25 mL of CH2Cl2 and
successively washed with 10 mL of saturated aqueous NaHCO3
and 10 mL of brine. The separated organic layer was dried
over MgSO4, filtered, concentrated in vacuo, and purified by
column chromatography (SiO2, Rf ) 0.35, 10% MeOH/CH2Cl2)
to give 120 mg (97%) of a white foam: 1H NMR (500 MHz,
CDCl3) δ 8.41 (d, 1H, J ) 7.6), 8.27 (s, 1H), 7.34 (d, 1H, J )
7.6), 6.24 (t, 1H, J ) 5.5), 4.38 (q, 1H, J ) 5.6), 3.98-3.95 (m,
2H), 3.78 (dd, 1H, J ) 12.0, 2.7), 2.53 (dt, 1H, J ) 13.4, 6.3),
2.23 (s, 3H), 2.13 (dt, 1H, J ) 12.7, 5.9), 0.93 (s, 9H), 0.88 (s,
9H), 0.123 (s, 3H), 0.111 (s, 3H), 0.058 (s, 3H), 0.055 (s, 3H);
13C NMR (125 MHz, CDCl3) δ 171.1, 162.8, 155.1, 144.7, 96.4,
87.8, 86.7, 69.8, 61.6, 42.2, 25.8, 25.6, 24.8, 18.3, 17.9, -4.7,
4-N-(8-Aminonaphthalen-2-ylformyl)-2′-deoxy-3′,5′-bis-
O-(tert-butyldimethylsilyl)cytidine (12). A mixture of 36
mg (0.0550 mmol) of 22 in 4 mL of 1:1 (v/v) MeOH/EtOAc was
hydrogenated at atmospheric pressure over 30 mg of Pd/C for
15 min. The mixture was filtered through Celite and the Celite
washed with 15 mL of EtOAc. The filtrate was concentrated
in vacuo and purified by column chromatography (SiO2, Rf )
0.15, 1:1 CHCl3/EtOAc) to give 34 mg (100%) of a yellow solid:
1H NMR (500 MHz, DMSO-d6) δ 10.99 (s, 1H), 8.95 (s, 1H),
8.30 (d, 1H, J ) 7.5), 7.91 (d, 1H, J ) 8.8), 7.81 (d, 1H, 8.6),
7.47 (d, 1H, J ) 6.9), 7.34 (t, 1H, J ) 7.8), 7.11 (d, 1H, J )
7.9), 6.74 (d, 1H, J ) 7.6), 6.14 (t, 1H, J ) 6.0), 6.07 (br s, 2H),
4.40 (dt, 1H, J ) 5.1, 4.6), 3.91 (q, 1H, J ) 3.6), 3.86 (dd, 1H,
J ) 11.5, 3.8), 3.76 (dd, 1H, J ) 11.4, 3.0), 2.34 (dt, 1H, J )
12.9, 5.6), 2.20 (dt, 1H, J ) 13.4, 5.9), 0.90 (s, 9H), 0.87 (s,
9H), 0.102 (s, 3H), 0.096 (s, 3H), 0.081 (s, 6H); 13C NMR (125
MHz, DMSO-d6) δ 166.9, 163.0, 154.4, 146.6, 144.6, 136.1,
129.9, 128.2, 127.2, 124.1, 124.0, 121.1, 114.8, 108.3, 95.8, 87.1,
86.0, 70.8, 61.9, 40.9, 25.7, 25.7, 18.0, 17.7, -4.8, -5.0, -5.6,
-5.6; FD-LRMS m/z 624 (M+). Anal. Calcd for C32H48N4O5Si2:
C, 61.50; H, 7.74; N, 8.97. Found: C, 61.72; H, 7.78; N, 8.97.
4-N-[8-(3-Methylureido)naphthalen-2-ylformyl]-2′-
deoxy-3′,5′-bis-O-(tert-butyldimethylsilyl)cytidine (13).
To a suspension of 75 mg (0.307 mmol) of 20, 62 mg (0.323
mmol) of 1-[3-(dimethylamino)propyl]-3-ethylcarbodiimide hy-
drochloride, 46 mg (0.340 mmol) of 1-hydroxybenzotriazole,
and 2 mL of THF was added a solution of 149 mg (0.327 mmol)
of 17 in 1 mL of THF at rt. The solution was stirred at rt for
22 h, diluted with 25 mL of CH2Cl2, and successively washed
(29) Connors, K. A. Binding Constants: The Measurement of Mo-
lecular Complex Stability; Wiley: New York, 1987.
(30) Igolen, J.; Morin, C. J. Org. Chem. 1980, 45, 4802-4804.
J. Org. Chem, Vol. 70, No. 19, 2005 7465

Quinn and Zimmerman
with 15 mL of saturated aqueous NaHCO3 and 15 mL of brine.
The separated organic layer was dried (Na2SO4), filtered,
concentrated in vacuo, and purified by column chromatography
(SiO2, Rf ) 0.6, 10% MeOH/CH2Cl2) using 8% MeOH/CH2Cl2
to give 170 mg (81%) of a yellow foam: 1H NMR (500 MHz,
DMSO-d6) δ 11.34 (s, 1H), 8.792 (s, 1H), 8.746 (s, 1H), 8.325
(d, 1H, J ) 7.2), 8.03-7.97 (m, 3H), 7.62 (d, 1H, J ) 8.0), 7.56
(t, 1H, J ) 7.9), 7.44 (d, 1H, J ) 7.2), 6.52 (q, 1H, J ) 4.4),
6.14 (t, 1H, J ) 5.9), 4.40 (dt, 1H, J ) 5.1, 4.9), 3.91 (dt, 1H,
J ) 3.8, 3.6), 3.86 (dd, 1H, J ) 11.3, 4.0), 3.76 (dd, 1H, J )
11.4, 2.8), 2.72 (d, 3H, J ) 4.7), 2.35 (m, 1H), 2.20 (dt, 1H, J
) 13.1, 6.1), 0.91 (s, 9H), 0.88 (s, 9H), 0.11 (s, 3H), 0.105 (s,
3H), 0.09 (s, 6H); 13C NMR (125 MHz, DMSO-d6) δ 167.7,
163.1, 156.1, 154.4, 144.6, 136.8, 135.6, 129.7, 128.7, 128.5,
124.4, 124.2, 123.8, 121.6, 117.2, 95.8, 87.1, 86.0, 70.7, 61.9,
2H), 4.34 (m, 1H), 3.79-3.75 (m, 2H), 3.69 (dd, 1H, J ) 10.9,
2.8), 2.13 (ddd, 1H, J ) 13.4, 6.4, 4.5), 2.04 (dt, 1H, J ) 13.1,
6.7), 0.88 (s, 9H), 0.86 (s, 9H), 0.067 (s, 12H); 13C NMR (125
MHz, DMSO-d6) δ 163.3, 155.0, 144.6, 139.6, 133.9, 133.4,
128.2, 126.6, 125.6, 122.5, 121.0, 115.3, 107.8, 94.7, 86.5, 84.6,
71.4, 62.3, 43.9, 40.3, 25.7, 25.7, 17.9, 17.7, -4.8, -5.0, -5.6,
-5.6; ESI-HRMS calcd for (C32H50N4O4Si2‚H)+ 611.3449, found
611.3462.
4-N-(8-Nitroquinolin-2-ylformyl)-2′-deoxy-3′,5′-bis-O-
(tert-butyldimethylsilyl)cytidine (21). To a suspension of
106 mg (0.509 mmol) of 18, 102 mg (0.532 mmol) of 1-[3-
(dimethylamino)propyl]-3-ethylcarbodiimide hydrochloride, 76
mg (0.562 mmol) of 1-hydroxybenzotriazole, and 3 mL of THF
was added a solution of 247 mg (0.542 mmol) of 17 in 2 mL of
THF at rt. The mixture was stirred for 21 h at rt and then
diluted with 50 mL of CH2Cl2 and successively washed with
25 mL of saturated aqueous NaHCO3 and 25 mL of brine. The
separated organic layer was dried over Na2SO4, filtered,
concentrated in vacuo, and purified by column chromatography
(SiO2, Rf ) 0.25, 3% MeOH/CH2Cl2) to give 277 mg (83%) of a
yellow foam: 1H NMR (500 MHz, CDCl3) δ 10.33 (s, 1H), 8.55-
8.50 (m, 3H), 8.40 (dd, 1H, J ) 8.3, 1.2), 8.22 (dd, 1H, J ) 7.5,
1.1), 7.79 (t, 1H, J ) 7.9), 6.59 (d, 1H, J ) 7.4), 6.29 (dd, 1H,
J ) 6.5, 4.4), 4.41 (dt, 1H, J ) 6.2, 5.3), 4.01 (dd, 1H, J )
11.3, 2.5), 3.98 (m, 1H), 3.81 (dd, 1H, J ) 11.3, 1.6), 2.58 (dt,
1H, J ) 13.6, 6.4), 2.21 (ddd, 1H, J ) 13.5, 6.4, 4.4), 0.97 (s,
9H), 0.89 (s, 9H), 0.16 (s, 3H), 0.14 (s, 3H), 0.074 (s, 3H), 0.067
(s, 3H); 13C NMR (100 MHz, CDCl3) δ 162.9, 161.4, 155.3,
150.0, 147.9, 145.0, 138.6, 137.7, 132.1, 130.2, 127.7, 125.6,
120.5, 95.8, 87.7, 86.9, 69.7, 61.6, 42.3, 25.9, 25.7, 18.4, 17.9,
-4.6, -5.0, -5.5, -5.5; [R]21D ) +37.2 (c ) 0.54, CHCl3); ESI-
HRMS calcd for (C31H45N5O7Si2‚H)+ 656.2936, found 656.2944.
Anal. Calcd for C31H45N5O7Si2: C, 56.77; H, 6.92; N, 10.68.
Found: C, 56.46; H, 6.86; N, 10.61.
4-N-(8-Nitronaphthalen-2-ylformyl)-2′-deoxy-3′,5′-bis-
O-(tert-butyldimethylsilyl)cytidine (22). To a suspension
of 205 mg (0.944 mmol) of 19, 196 mg (1.02 mmol) of 1-[3-
(dimethylamino)propyl]-3-ethylcarbodiimide hydrochloride, 145
mg (1.07 mmol) of 1-hydroxybenzotriazole, and 5 mL of THF
was added a solution of 445 mg (0.977 mmol) of 17 in 3 mL of
THF at rt. The suspension was stirred at rt for 25 h, diluted
with 50 mL of CH2Cl2, and successively washed with 25 mL
of saturated aqueous NaHCO3 and 25 mL of brine. The
separated organic layer was dried (Na2SO4), filtered, concen-
trated in vacuo, and purified by column chromatography (SiO2,
Rf ) 0.65, 10% MeOH/CH2Cl2) using 8% MeOH/CH2Cl2 to give
592 mg (96%) of a yellow foam: 1H NMR (500 MHz, DMSO-
d6) δ 11.67 (s, 1H), 9.03 (s, 1H), 8.45 (d, 1H, J ) 8.2), 8.40 (d,
1H, J ) 7.5), 8.34 (d, 1H, J ) 7.5), 8.30 (d, 1H, J ) 8.7), 8.18
(d, 1H, J ) 8.6), 7.86 (t, 1H, J ) 7.9), 7.43 (d, 1H, J ) 7.3),
6.13 (t, 1H, J ) 5.9), 4.40 (q, 1H, J ) 4.9), 3.92 (m, 1H), 3.87
(dd, 1H, J ) 11.5, 3.6), 3.77 (dd, 1H, J ) 11.4, 2.8), 2.36 (m,
1H), 2.21 (dt, 1H, J ) 13.5, 5.6), 0.91 (s, 9H), 0.88 (s, 9H),
0.11 (s, 3H), 0.105 (s, 3H), 0.09 (s, 6H); 13C NMR (100 MHz,
DMSO-d6) δ 167.0, 162.9, 154.1, 147.0, 144.5, 135.3, 134.4,
133.4, 129.1, 127.2, 126.2, 124.7, 123.9, 123.1, 95.9, 87.1, 86.0,
70.6, 61.8, 40.9, 25.7, 25.6, 18.0, 17.7, -4.8, -5.1, -5.6, -5.6;
[R]21D ) +33.3 (c ) 0.81, CHCl3); FD-LRMS m/z 655 (M+), 597
(100, -tBu). Anal. Calcd for C32H46N4O7Si2: C, 58.69; H, 7.08;
N, 8.55. Found: C, 58.70; H, 7.13; N, 8.50.
40.9, 26.3, 25.7, 25.6, 18.0, 17.7, -4.8, -5.0, -5.6, -5.6; [R]21
D
) +89.2 (c ) 0.57, CHCl3); FD-LRMS m/z 681 (M+), 651 (100,
-NHMe). Anal. Calcd for C34H51N5O6Si2: C, 59.88; H, 7.54;
N, 10.27. Found: C, 59.73; H, 7.54; N, 10.25.
4-N-(Quinolin-2-ylmethyl)-2′-deoxy-3′,5′-bis-O-(tert-butyl-
dimethylsilyl)cytidine (14). A solution of 1.10 g (2.17 mmol)
of 30 and 0.50 g (3.2 mmol) of 2331 in 5 mL of DMF was stirred
at 100 °C for 23 h. The solvent was removed via vacuum, and
the residue was purified by column chromatography using 2%
MeOH-CH2Cl2 (SiO2, Rf ) 0.25, 5% MeOH/CH2Cl2) to give
1.22 g (95%) of an off-white foam: 1H NMR (400 MHz, DMSO-
d6) 8.41 (t, 1H, J ) 5.9), 8.32 (d, 1H, J ) 8.3), 7.97-7.54 (m,
5H), 7.46 (d, 1H, J ) 8.5), 6.12 (t, 1H, J ) 6.3), 5.92 (d, 1H, J
) 7.3), 4.75 (m, 2H), 4.34 (m, 1H), 3.77-3.67 (m, 3H), 2.11-
2.02 (m, 2H), 0.87 (s, 9H), 0.84 (s, 9H), 0.062 (s, 6H), 0.048 (s,
6H); 13C NMR (100 MHz, DMSO-d6) δ 163.5, 158.9, 154.8,
147.0, 139.9, 136.7, 129.7, 128.4, 127.9, 127.0, 126.2, 119.9,
94.6, 86.5, 84.6, 71.4, 62.3, 45.7, 40.4, 25.8, 25.7, 18.0, 17.7,
-4.8, -5.0, -5.5, -5.6; [R]21 ) +3.3 (c ) 1.00, DMSO); ESI-
D
HRMS calcd for (C31H48N4O4Si2‚H)+ 597.3292, found 597.3287.
Anal. Calcd for C31H48N4O4Si2: C, 62.38; H, 8.11; N, 9.39.
Found: C, 61.98; H, 8.09; N, 9.30.
4-N-(8-Aminoquinolin-2-ylmethyl)-2′-deoxy-3′,5′-bis-O-
(tert-butyldimethylsilyl)cytidine (15). A solution of 886 mg
(1.74 mmol) of 30 and 324 mg (1.87 mg) of 24 in 8 mL of DMF
was stirred at 100 °C for 19 h. The mixture was concentrated
in vacuo and purified by column chromatography (SiO2, Rf )
0.10, 3% MeOH/CH2Cl2) to afford 890 mg (84%) of an orange
foam: 1H NMR (500 MHz, DMSO-d6) δ 8.32 (t, 1H, J ) 5.1),
8.13 (d, 1H, J ) 8.6), 7.74 (d, 1H, J ) 7.7), 7.40 (d, 1H, J )
8.5), 7.24 (t, 1H, J ) 7.8), 7.01 (d, 1H, J ) 7.6), 6.84 (d, 1H, J
) 7.8), 6.16 (t, 1H, J ) 6.4), 6.06 (d, 1H, J ) 7.2), 6.05 (br s,
2H), 4.76 (m, 2H), 4.35 (m, 1H), 3.81-3.70 (m, 3H), 2.14 (ddd,
1H, J ) 13.2, 6.1, 4.2), 2.05 (dt, 1H, J ) 13.1, 6.2), 0.89 (s,
9H), 0.86 (s, 9H), 0.079 (s, 6H), 0.064 (s, 6H); 13C NMR (100
MHz, DMSO-d6) δ 163.4, 154.9, 154.0, 145.0, 139.6, 136.6,
136.2, 127.5, 127.2, 119.8, 113.4, 108.9, 94.8, 86.5, 84.6, 71.4,
62.3, 45.5, 40.4, 25.8, 25.7, 18.0, 17.7, -4.8, -5.0, -5.5, -5.6;
[R]21D ) +16.7 (c ) 0.52, CHCl3); FD-LRMS m/z 611 (M+). Anal.
Calcd for C31H49N5O4Si2: C, 60.85; H, 8.07; N, 11.44. Found:
C, 60.67; H, 8.19; N, 11.40.
4-N-(8-Aminonaphthalen-2-ylmethyl)-2′deoxy-3′,5′-bis-
O-(tert-butyldimethylsilyl)cytidine (16). A mixture of 64
mg (0.0999 mmol) of 31 and 4 mL of 1:1 (v/v) MeOH/EtOAc
was hydrogenated at atmospheric pressure for 14 h over 50
mg of Pd/C. The mixture was filtered through Celite, and the
Celite was washed twice with 10 mL of 5% (v/v) MeOH/EtOAc.
The filtrate was concentrated in vacuo and purified by column
chromatography (SiO2, Rf ) 0.30, 100:50:1 EtOAc/CHCl3/Et3N)
to give 29 mg (48%) of an orange foam: 1H NMR (500 MHz,
DMSO-d6) δ 8.15 (t, 1H, J ) 5.3), 7.97 (s, 1H), 7.70 (d, 1H, J
) 7.6), 7.69 (d, 1H, J ) 8.6), 7.34 (d, 1H, J ) 8.2), 7.16 (t, 1H,
J ) 7.8), 7.04 (d, 1H, J ) 8.1), 6.65 (d, 1H, J ) 7.1), 6.16 (t,
1H, J ) 6.5), 5.82 (d, 1H, J ) 7.4), 5.66 (br s, 2H), 4.60 (m,
2-Aminomethylquinolin-8-ylamine (24). A solution of 28
(0.943 g, 4.74 mmol) in 100 mL of 5% (v/v) trifluoroacetic acid/
MeOH was hydrogenated (Parr apparatus) at 55 psi for 2.5 h
over Pd/C. The mixture was filtered through Celite, concen-
trated in vacuo, and made slightly basic with 10 mL of water
and 7 mL of saturated aqueous Na2CO3. The mixture was
extracted twice with 70 mL of CH2Cl2, and the combined
organic layers were successively washed with 15 mL of water
and 25 mL of brine. The organic layer was dried over K2CO3,
filtered, and concentrated in vacuo to afford 630 mg (77%) of
a yellow oil that solidified upon standing. The product was of
sufficient purity to carry forward without further purification
(31) Yousif, M. M.; Saeki, S.; Hamana, M. J. Heterocycl. Chem. 1980,
17, 1029. Langry, K. C. Org. Prep. Proc. Int. 1994, 26, 429-438.
7466 J. Org. Chem., Vol. 70, No. 19, 2005

Structure-Function Studies on a Synthetic Guanosine Receptor
(see the Supporting Information): 1H NMR (500 MHz, CDCl3)
δ 8.08 (d, 1H, J ) 8.4), 7.31 (d, 1H, J ) 8.4), 7.29 (dd, 1H, J
) 7.9, 7.7), 7.14 (dd, 1H, J ) 8.1, 1.3), 6.93 (dd, 1H, J ) 7.5,
1.3), 4.98 (br s, 2H), 4.13 (s, 2H), 1.81 (br s, 2H); 13C NMR
(125 MHz, CDCl3) δ 159.7, 144.0, 138.0, 137.0, 128.2, 127.2,
120.5, 116.5, 110.8, 48.6; FD-LRMS m/z 173 (M+).
1H, J ) 7.3), 8.11 (s, 1H), 6.97 (d, 1H, J ) 7.3), 6.25 (dd, 1H,
J ) 6.4, 4.3), 4.39 (dt, 1H, J ) 6.2, 5.1), 3.99 (m, 2H), 3.80 (m,
1H), 2.58 (dt, 1H, J ) 13.7, 6.4), 2.19 (ddd, 1H, J ) 13.7, 6.4,
4.5), 0.93 (s, 9H), 0.87 (s, 9H), 0.13 (s, 3H), 0.11 (s, 3H), 0.05
(s, 3H), 0.04 (s, 3H); 13C NMR (125 MHz, CDCl3) δ 159.1, 154.4,
153.9, 147.0, 143.2, 94.0, 88.0, 87.5, 69.5, 61.5, 42.3, 25.9, 25.6,
18.3, 17.9, -4.6, -5.0, -5.5; [R]21D ) +56.5 (c ) 1.00, CHCl3);
ESI-HRMS calcd for (C23H41N5O4Si2‚H)+ 508.2775, found
508.2787. Anal. Calcd for C23H41N5O4Si2: C, 54.40; H, 8.14;
N, 13.79. Found: C, 54.39; H, 8.25; N, 13.51.
8-Nitroquinoline-2-carbonitrile (28). To a suspension of
15 g (0.23 mol) of KCN and 400 mL of MeOH was added a
solution of 22.0 g (0.116 mol) of 2632 in 500 mL of CH3CN.
Benzoyl chloride (27 mL, 33 g, 0.23 mol) was added over 5
min. The mixture was stirred for 20 h at rt and then treated
with 300 mL of H2O and 400 mL of CH2Cl2. The organic layer
was removed, and the aqueous layer was successively ex-
tracted with 400 mL and 100 mL of CH2Cl2. The combined
organic layers were washed with 200 mL of brine, dried over
MgSO4, filtered, concentrated in vacuo, purified by column
chromatography (SiO2, Rf ) 0.4, 1:1 PE:EtOAc), and recrystal-
lized from CH3CN/CHCl3 to give 9.3 g (40%) of light brown
needles. A total of 8.2 g of 26 was recovered from chromatog-
raphy. Crystals suitable for X-ray crystallographic analysis
were obtained by slow evaporation (see the Supporting Infor-
mation for the CIF file): mp 194-195 °C; 1H NMR (500 MHz,
CDCl3) δ 8.45 (d, 1H, J ) 8.6), 8.17 (dd, 1H, J ) 7.5, 1.3), 8.13
(dd, 1H, J ) 8.4, 1.5), 7.87 (d, 1H, J ) 8.4), 7.81 (dd, 1H, J )
8.4, 7.5); 13C NMR (125 MHz, CDCl3) δ 148.5, 139.9, 138.4,
136.2, 132.3, 129.5, 128.8, 125.8, 125.7, 117.1; IR (KBr) 3413,
3075, 2245, 1538, 1498, 1378, 1310; FD-LRMS m/z 199 (M+).
Anal. Calcd for C10H5N3O2: C, 60.31; H, 2.53; N, 21.10.
Found: C, 60.25; H, 2.40; N, 20.72.
1-[2-Deoxy-3,5-bis-O-(tert-butyldimethylsilyl)-â-D-ribo-
furanosyl]-4-[1,2,4]triazol-1-yl-1H-pyrimidin-2-one (30).33
To a suspension of 3.9 g (56 mmol) of 1,2,4-1H-triazole, 1.1
mL (1.8 g, 11.8 mmol) of POCl3, and 32 mL of CH3CN at 0 °C
was added a solution of 1.87 g (4.09 mmol) of 2934 in 17 mL of
CH3CN over 15 min. The suspension was stirred at 0 °C for
20 min and then at ambient temperature for 1 h. The reaction
was quenched with 4.5 mL of Et3N and 1.7 mL of water and
the mixture concentrated in vacuo. The residue was dissolved
in 75 mL of CH2Cl2 and washed with 50 mL of 0.5 M aqueous
NaHCO3. The organic layer was removed, and the aqueous
layer was extracted three times with 75 mL each of CH2Cl2.
The combined organic layers were dried over Na2SO4, filtered,
concentrated in vacuo, and purified by column chromatography
(SiO2, Rf ) 0.40, 5% MeOH/CH2Cl2) using 100:1f100:3 CH2Cl2/
MeOH to give 2.03 g (98%) of a colorless oil that solidifies on
standing: 1H NMR (500 MHz, CDCl3) δ 9.26 (s, 1H), 8.71 (d,
4-N-(8-Nitronaphthalen-2-ylmethyl)-2′-deoxy-3′,5′-bis-
O-(tert-butyldimethylsilyl)cytidine (31).35 To a solution of
95 mg (0.145 mmol) of 22 in 2 mL of THF was added 0.177
mL (147 mg, 0.725 mmol) of N,O-bis(trimethylsilyl)acetamide
dropwise at rt. The solution was treated dropwise with 0.252
mL (207 mg, 1.45 mmol) of borane-N,N-diisopropylethylamine
complex and the solution stirred at rt for 15 min. The reaction
was quenched with 10 mL of methanol, and the mixture was
concentrated in vacuo. The resulting residue was dissolved in
80 mL of a 1:1 (v/v) mixture of methanolic ammonia (17 wt
%)/aqueous ammonia (28 wt %) and incubated for 13 h at 50
°C. The mixture was cooled to room temperature, diluted with
100 mL of water, and partially neutralized with 25 mL of 1 N
aqueous HCl. The mixture was extracted with chloroform (2
× 50 mL), and the combined extracts were washed with brine,
dried over MgSO4, filtered, concentrated in vacuo, and purified
by column chromatography (SiO2, Rf ) 0.15, 5% MeOH/
CH2Cl2) to give 69 mg (74%) of an orange foam: 1H NMR (500
MHz, DMSO-d6) δ 8.37 (t, 1H, J ) 5.9), 8.34 (d, 1H, J ) 8.3),
8.31 (d, 1H, J ) 7.6), 8.23 (s, 1H), 8.14 (d, 1H, J ) 8.5), 7.73
(d, 1H, J ) 7.3), 7.67 (t, 1H, J ) 7.9), 7.65 (d, 1H, J ) 8.5),
6.13 (t, 1H, J ) 6.2), 5.85 (d, 1H, J ) 7.3), 4.72 (m, 2H), 4.34
(m, 1H), 3.79-3.68 (m, 3H), 2.12 (m, 1H), 2.04 (dt, 1H, J )
12.9, 6.8), 0.88 (s, 9H), 0.85 (s, 9H), 0.067 (s, 6H), 0.58 (s, 6H);
ESI-HRMS calcd for (C32H48N4O6Si2‚H)+ 641.3191, found
641.3174. Anal. Calcd for C32H48N4O6Si2: C, 59.97; H, 7.55;
N, 8.74. Found: C, 59.87; H, 7.58; N, 8.55.
Acknowledgment. Funding of this work by the NIH
(GM65249) is gratefully acknowledged.
Supporting Information Available: General experimen-
tal procedures, X-ray analysis data for compound 28 (CIF file),
nonlinear regression plots for complexation studies, and NMR
spectra. This material is available free of charge via the
Internet at http://pubs.acs.org.
(32) Yokoyama, A.; Ohwada, T.; Saito, S.; Shudo, K. Chem. Pharm.
Bull. 1997, 45, 279-283.
JO0501689
(33) Procedure adapted from: Trafelet, H.; Stulz, E.; Leumann, C.
Helv. Chim. Acta 2001, 84, 87-105.
(34) Ogilvie, K. K.; Beaucage, S. L.; Schifman, A. L.; Theriault, N.
Y.; Sadana, K. L. Can. J. Chem. 1978, 56, 2768-2780.
(35) Procedure adapted from: Sergueeva, Z. A.; Sergueev, D. S.;
Shaw, B. R. Nucleosides and Nucleotides 2000, 19, 275-282.
J. Org. Chem, Vol. 70, No. 19, 2005 7467