Design, synthesis and binding affinity of 3′-fluoro analogues of Cl-IB-MECA as adenosine A3 receptor ligands

Article abstract of DOI:10.1016/S0960-894X(03)00027-1

Several 3′-fluoro analogues, 1a, 1b, and 1c of selective and potent adenosine A₃ receptor agonist, Cl-IB-MECA were synthesized from D-xylose via highly regioselective opening of lyxo-epoxides, 8a and 8b with fluoride anion. Compared to the high binding affinity of Cl-IB-MECA to the A₃ adenosine receptor, the corresponding 3′-fluoro derivative showed remarkably decreased binding affinity, indicating that 3′-hydroxyl group acts as hydrogen bonding acceptor, not hydrogen bonding donor like fluorine atom in binding to the A₃ adenosine receptor.

Full text of DOI:10.1016/S0960-894X(03)00027-1

Bioorganic & Medicinal Chemistry Letters 13 (2003) 817–820
Design, Synthesis and Binding Affinity of 30-Fluoro Analogues of
Cl-IB-MECA as Adenosine A3 Receptor Ligands
Moo Hong Lim,a Hea Ok Kim,b Hyung Ryong Moon,c Seung Jin Lee,c
Moon Woo Chun,a,* Zhan-Guo Gao,d Neli Melman,d Kenneth A. Jacobson,d
Joong Hyup Kime and Lak Shin Jeongc,*
aCollege of Pharmacy, Seoul National University, Seoul 151-742, South Korea
bDivision of Chemistry andMolecular Engineering, Seoul National University, Seoul 151-742, South Korea
cLaboratory of Medicinal Chemistry, College of Pharmacy, Ewha Womans University, Seoul 120-750, South Korea
dMolecular Recognition Section, Laboratory of Bioorganic Chemistry, National Institute of Diabetes,
and Digestive and Kidney Disease, National Institutes of Health, Bethesda, MA 20892, USA
eKorea Institute of Science andTechnology, Seoul 136-791, South Korea
Received 1 November 2002; accepted 26 December 2002
Abstract—Several 30-fluoro analogues, 1a, 1b, and 1c of selective and potent adenosine A3 receptor agonist, Cl-IB-MECA were
synthesized from d-xylose via highly regioselective opening of lyxo-epoxides, 8a and 8b with fluoride anion. Compared to the high
binding affinity of Cl-IB-MECA to the A3 adenosine receptor, the corresponding 30-fluoro derivative showed remarkably decreased
binding affinity, indicating that 30-hydroxyl group acts as hydrogen bonding acceptor, not hydrogen bonding donor like fluorine
atom in binding to the A3 adenosine receptor.
# 2003 Elsevier Science Ltd. All rights reserved.
Introduction
A number of compounds have been synthesized and
evaluated for binding affinity to adenosine A3 receptor
for the development of therapeutically useful agents.
Four major subtypes of adenosine receptors, A1, A2A
,
A2B, and A3 have been identified so far,1,2 among which
A3 adenosine receptors3 were mostrecenlty idenitfied.
The A3 adenosine receptors may play an important role
in the regulation of CNS, cardiac, inflammatory, and
reproductive functions. Chronic administration of an
A3 agonist showed high cerebroprotection in a model of
global cerebral ischemia in gerbrils4 and significant
protection against chemically induced seizures5 in mice.
Since A3 receptors are expressed on the cardiac ven-
tricular cell, the activation of A3 receptor is related to
the cardioprotective preconditioning response,6 and
thus the A3 adenosine receptor agonists show a power-
ful protection against myocardial ischemia. Moreover,
adenosine A3 receptor antagonists may have potential
use in treating inflammation since A3 receptor stimu-
lates the release of histamine on mast cells.7 The ade-
nosine A3 receptor antagonists also may be developed
as anti-asthma agent since the A3 receptor is highly
expressed on eosinophils in the lung.8
Among these compounds, 2-chloro-N6-(3-iodobenzyl)-
adenosine-50-methylcarboxamide (Cl-IB-MECA) was
discovered to be one of the most selective agonists
(Ki=0.33 nM)9 atratA
adenosine receptor from the
3
structure–activity relationship study for N6- and 50-sub-
stituted adenosine derivatives (Fig. 1).
*Corresponding author. Tel.: +82-2-3277-3466; fax: +82-2-3277-
2851; e-mail: lakjeong@ewha.ac.kr
Figure 1. The rationale for the design of the desired nucleosides.
0960-894X/03/$ - see front matter # 2003 Elsevier Science Ltd. All rights reserved.
doi:10.1016/S0960-894X(03)00027-1

818
M. H. Lim et al. / Bioorg. Med. Chem. Lett. 13 (2003) 817–820
On the basis of its high binding affinity to adenosine A3
receptor, it was interesting to determine whether 20- or
30-hydroxyl group of 2-Cl-IB-MECA is essential for the
binding affinity to the receptor or compatible with bioi-
sosteric fluorine for the binding affinity to this receptor.
For this purpose, we substituted the 30-hydroxyl group
of Cl-IB-MECA with bioisosteric fluorine. In this
communication, we wish to report the synthesis of 30-
fluoro analogue 1c of Cl-IB-MECA and its related
compounds, 1a and 1b (Fig. 1) and their binding
affinities to adenosine A3 receptor.
synthetic route to utilize the regioselective opening of
lyxo-epoxides 8a and 8b and directS N2 displacementof
10a and 10b for inversion of stereochemistry at C2
position as key steps (Scheme 2).
d-Xylose was converted to anomeric methoxides 6a and
6b according to Baker’s method.10 Tosylation of 6a and
6b gave 7a and 7b, which were converted to lyxo-epox-
ides 8a and 8b, respectively, by treating with 80% acetic
acid followed by addition of sodium methoxide. Regio-
selective formation of 3-deoxy-3-fluoro derivatives, 9a
(82%) and 9b (96%) was cleanly obtained by refluxing
of 8a and 8b with potassium hydrogen fluoride and
sodium fluoride in 1,2-ethylene glycol, respectively.
Higher yield of 9b than 9a was attributed to the difficulty
in attack of the fluoride anion at the C2 position due to
the steric effect by the a-methoxy substituent. Interest-
ingly, the same ring-opening reaction on lyxo-epoxide
with its 5-hydroxyl group protected as benzyl ether11 or
benzoyl group afforded the desired 3-deoxy-3-fluoro
analogue in moderate yield. This might be due to many
side reactions as well as the separation problem caused
by ethylene glycol during silica gel column chromato-
graphy. For the conversion of arabino configuration to
ribo configuration, silyl protection of 9a and 9b as TBS
ethers followed by tosylation afforded 10a and 10b,
respectively. When b-methoxide 10a was treated with
sodium benzoate in DMSO at 200 ꢀC, the desired 11b
(38%) and its debenzoylated compound 11a (52%) were
obtained as major products. Interestingly, 5-TBS group
was transformed to the benzoyl group under the reac-
tion conditions. It is believed that silicon-oxygen bond
was first cleaved on thermal conditions to form 5-TBS
cation and oxygen anion. TBS cation was then reacted
with sodium benzoate to give 5-TBS-benzoate, which
was finally attacked by oxygen anion to yield the 5-
benzoate derivative. However, under the same reaction
conditions, SN2 displacementof a-methoxide 10b by
sodium benzoate was greatly hindered by the presence of
a-methoxy substituent at C1 position, yielding the SN2
displaced products, 13a (17%) and 13b (10%) in low
yields with recovered starting material 10b (16%) and its
desilylated compound 12 (29%). Compounds 11b and
13b were each converted to the same glycosyl donor 14.
Results and Discussion
Our original strategy to synthesize the desired 30-deoxy-
30-fluoroadenosine analogues was directly to fluorinate
on carbohydrates 2 or nucleosides 4 with xylo configur-
ation, as shown in Scheme 1, but many attempts to
fluorinate them under the various reaction conditions
failed to give the desired fluorinated compounds, 3 and
5, respectively.
Thus, synthetic difficulties in fluorinating the com-
pounds with xylo configuration made us take alternative
Scheme 1.
Scheme 2. Reagents and conditions: (a) TsCl, pyridine, rt, 1–3 days; (b) (i) 80% AcOH 50 ꢀC, 2 h; (ii) NaOMe, MeOH, 0–10 ꢀC, 2–3 days; (c) KHF2,
NaF, 1,2-ethylene glycol, reflux, 0.5–1 h; (c) TBSCl, imidazole, DMF, 0–10 ꢀC, 15 h; (e) TsCl, pyridine, rt, 13 h; (f) NaOBz, 18-crown-6, DMSO,
200 ꢀC, 13 h (g) BzCl, pyridine, rt, 5 h; (h) Ac2O, AcOH, H2SO4, rt, 15 min.

M. H. Lim et al. / Bioorg. Med. Chem. Lett. 13 (2003) 817–820
819
For the synthesis of N6-substituted adenosine deriva-
tives 1a and 1b (Scheme 3), glycosyl donor 14 was con-
densed with silylated 2,6-dichloropurine to give the
protected nucleoside 15. Treatment of 15 with 3-iodo-
benzylamine and methylamine in ethanol afforded N6-
(3-iodobenzyl)- and N6-methyladenosine derivatives 16
and 17, respectively. The final nucleosides, 1a and 1b
were obtained after debenzoylation of 16 and 17,
respectively.
the conversion of anomeric methoxide to anomeric ace-
tate 22 under acidic conditions, TBS group in 20 was
changed to the benzoyl group, giving 21.
The glycosyl donor 22 was condensed with silylated 2-
chloro-N6-(3-iodobenzyl)adenine in the presence of
TMSOTf to give the protected nucleoside 23 (Scheme 5).
Even on treatment of methyl ester 23 with 2 M methy-
lamine solution in THF for only 4 min, substantial
amounts of elimination product 24 (25%) was obtained
along with the final nucleoside 1c (12%), its benzoylated
compound 25 (35%) and recovered starting material 23
(20%). Treatment of 25 with 2 M methylamine solution
for the removal of benzoyl group afforded the 30-fluoro
analogue of Cl-IB-MECA, 1c (47%) with concomitant
formation of elimination product 24 (38%).
For the synthesis of 30-fluoro analogue of Cl-IB-MECA,
another glycosyl donor 22 was synthesized as shown in
Scheme 4.
Compound 11a was protected as TBS ether 18, in which
benzoyl group was removed to give 19. Oxidation of the
primary hydroxyl group of 19 followed by esterification
using DCC and methanol afforded methyl ester 20. For
The final nucleosides 1a–1c were tested in radioligand
binding assays12ꢁ15 for affinities at rat brain A112 and
13
2A
14,15
A
and human A3
adenosine receptors (Table 1).
As shown in Table 1, substitution of 30-hydroxyl group
with fluorine resulted in dramatic decrease in binding
affinities to adenosine receptors, despite of their bioi-
sosteric relationship. Compared to the high binding
affinity (Ki=1.0 nM) of Cl-IB-MECA to the human A3
adenosine receptor, binding affinities (Ki=75 and 326
nM) of compounds, 1a and 1b t o A receptor were
3
remarkably diminished. Moreover, conversion of 40-
hydroxymethyl derivative 1a into its methyl amide 1c
resulted in ca. 5-fold further decrease in binding affinity
(Ki=406 nM) to the A3 receptor, although 40-urona-
mide moiety has been generally reported to show better
affinity in binding to adenosine receptors than the cor-
responding 40-hydroxymethyl moiety. It is attributed
that hydrogen bonding capacity of the 40-uronamide
was greatly diminished due to the presence of the
strongly electronegative fluorine. However, all tested
compounds did notshow any binding affiniyt ( Ki>10
Scheme 3. Reagents and conditions: (a) silylated 2,6-dichloropurine,
TMSOTf, 0–50 ꢀC, 4 h; (b) 3-iodobenzylamine hydrochloride, Et3N,
EtOH, 50 ꢀC, 5 h for 16; methylamine hydrochloride, Et3N, EtOH, rt,
4 h for 17; (c) NaOMe, MeOH, rt, 1 h.
Scheme 4. Reagents and conditions: (a) TBSCl, imidazole, rt, 6 h; (b)
NaOMe, MeOH, rt, 2 h; (c) RuCl3, NaIO4, MeCN/CCl4/H2O (1/1/
1.5), rt, 2 h, then DCC, DMAP, MeOH/CH2Cl2, rt, 10 h; (d) TBAF/
AcOH, THF, rt, 3 days, then BzCl, pyridine, rt, 3 h; (e) Ac2O, AcOH,
H2SO4, 0ꢀC to rt, 30 min.
Scheme 5. Reagents and conditions: (a) silylated 2-chloro-N6-(3-iodo-
benzyl)adenine, TMSOTf, ClCH2CH2Cl, 0–55 ꢀC; (b) 2 M MeNH2,
THF, rt, 4 min.

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M. H. Lim et al. / Bioorg. Med. Chem. Lett. 13 (2003) 817–820
Table 1. Binding affinities of the 30-fluoronucleoside analogues to adenosine receptors
Compd R, X
Ki (nM)
a
b
c
rA1
rA2A
hA3
Cl-IB-MECA R=3-iodobenzyl X=CONHMe
Compound 1a R=3-iodobenzyl X=CH2OH
Compound 1b R=CH3 X=CH2OH
820ꢂ570
1,350ꢂ350
17,100ꢂ5300
780ꢂ280
470ꢂ365
>10,000
>10,000
>10,000
1.0ꢂ0.2
75ꢂ7
326ꢂ112
406ꢂ60
Compound 1c R=3-iodobenzyl X=CONHMe
aDisplacementof specific binding of [ 3H]PIA, unless noted, in rat brain membranes expressed as KiꢂSEM in nM (n=3–6).
bDisplacementof specific binding of [ 3H]CGS 21680, unless noted, in rat brain membranes expressed as KiꢂSEM in nM (n=3–6).
cDisplacementof specific binding of [ 125I]-AB-MECA binding, unless noted, in CHO cells expressing the recombinant receptor as KiꢂSEM in nM
(n=3–5).
2. Daly, J. W.; Jacobson, K. A. In Adenosine Receptors:
mM) to A2A receptor and exhibited similar binding affi-
Selective Agonists andAntagonists ; Bellardinelli, L., Pelleg, A.,
nities to A1 receptor except 1b. This biological results
indicate that 30-hydroxyl group plays an essential role in
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fluorine via regioselective opening of the lyxo-epoxide
with fluoride anion and evaluated them for binding
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have found very important and essential role of 30-
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This work was supported by Korea Research Found-
ation Grant (KRF-2000-042-F00121).
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