Synthesis and biological evaluation of purine derivatives incorporating metal chelating ligands as HIV integrase inhibitors

Article abstract of DOI:10.1016/j.bmc.2006.04.011

Because of its essential role in HIV replication and lack of human counterpart, HIV integrase is an attractive target for the development of novel anti-AIDS agents. Among the recently developed integrase inhibitors, only the α,γ-diketo acid (DKA) compound

Full text of DOI:10.1016/j.bmc.2006.04.011

Bioorganic & Medicinal Chemistry 14 (2006) 5742–5755
Synthesis and biological evaluation of purine derivatives
incorporating metal chelating ligands as HIV integrase inhibitors
Xingnan Li and Robert Vince*
Center for Drug Design, Academic Health Center, and Department of Medicinal Chemistry, College of Pharmacy,
University of Minnesota, 8-123A WDH, 308 Harvard St SE, Minneapolis, MN 55455, USA
Received 3 February 2006; revised 3 April 2006; accepted 6 April 2006
Available online 5 June 2006
Abstract—Because of its essential role in HIV replication and lack of human counterpart, HIV integrase is an attractive target for
the development of novel anti-AIDS agents. Among the recently developed integrase inhibitors, only the a,c-diketo acid (DKA)
compounds were biologically validated as potent and selective integrase inhibitors. The general structure of DKAs contains a diketo
acid moiety as the Mg2+ chelating pharmacophore, and an adjacent aryl group to provide selectivity. Numerous structure–activity
relationship (SAR) studies on DKAs have been conducted, which generally involved substituting the carboxylate group or the aryl
group. Our objective was to investigate the SARs of the DKA molecule by incorporating a purine ring in the aryl moiety and replac-
ing the labile diketo acid moiety with other divalent metal (Me2+) chelating ligands. A series of amide substituted purine derivatives
were synthesized via palladium-catalyzed amidation reactions, and their biological activities against HIV integrase were evaluated.
These purine derivatives showed anti-integrase activity at low micromolar range. The biological results indicated that the type of
Me2+ ligands, two-point ligand picolinamide or three-point ligand 8-hydroxy-quinoline-7-carboxamide, affected inhibitory potency
depending on the substitution position of the para-fluorobenzyl group. The C6-,C8-dipicolinamide substituted purine (32) exhibited
the best potency among this series.
Ó 2006 Elsevier Ltd. All rights reserved.
1. Introduction
HIV integrase has recently attracted attention as a
promising anti-HIV target because of its necessity in vir-
al replication and its lack of human counterpart (see re-
views5–7). HIV integrase catalyzes the insertion of viral
DNA into host genome, a critical step for viral matura-
tion that includes a 30-processing step and a subsequent
strand transfer step.5 Recent studies using recombinant
integrase and high throughput screening assays have
led to the identification of the a,c-diketo acid (DKA)
type of compounds as potent and selective HIV inte-
grase inhibitors (Fig. 1A, 1 and 2).8,9 DKAs compete
with host DNA for the binding sites on HIV integrase,
thus selectively inhibiting the strand transfer process.8
This inhibition is dependent on the presence of divalent
magnesium ion, or manganese ion in in vitro assays.10 It
is believed that the a,c-diketo acid moiety chelates with
two divalent metal ions on the active site of the integrase
to form a tertiary ligand–Me2+–integrase complex, thus
blocking substrate DNA binding.10
The acquired immunodeficiency syndrome (AIDS) has
become a worldwide pandemic since its first diagnosis
in the early 1980s. It has claimed the lives of more than
25 million people globally and over 40 million people are
still living with HIV.1 Currently, a standard treatment
strategy for AIDS is referred to as the highly active anti-
retroviral therapy (HAART), which is a combination of
three or more anti-HIV drugs inhibiting two viral en-
zymes (i.e., reverse transcriptase and protease) and virus
fusion. Although HAART regimens have greatly
improved the life span and life quality of AIDS
patients,2 their application and effectiveness is hampered
by related toxicity issues and the ever-increasing
multi-drug resistance of HIV.3,4 New anti-HIV agents
with alternative target sites, therefore, are greatly
desired either as replacements or as supplements to the
current HAART regimens.
The general structure of DKAs contains an a,c-diketo
acid moiety as the key pharmacophore,10 and an adja-
cent aryl group providing efficacy and selectivity11–13
(Fig. 1B). Numerous structure–activity relationship
(SAR) studies on DKAs have been conducted to search
Keywords: HIV integrase inhibitor; Purine; Metal chelating ligand;
Pd-catalyzed amidation.
*
Corresponding author. Tel.: +1 612 624 9911; fax: +1 612 624
0139; e-mail: vince001@umn.edu
0968-0896/$ - see front matter Ó 2006 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmc.2006.04.011

X. Li, R. Vince / Bioorg. Med. Chem. 14 (2006) 5742–5755
5743
A
OH
O
O
N
NH
OH
N
N
O
O
OH
F
F
2
1
S-1360
(L-731,988)
B
O
O
COOH
R1
R2
R1=α,γ– diketo acid
R2= Aromatic or heteroaromatic moieties
Figure 1. (A) Representative a,c-diketo acid compounds (1 and 2) as selective HIV integrase inhibitors, (B) general structure of a,c-diketo acid
(DKA).
for a clinical candidate. Many of the reported SARs
focused on substituting the aryl group with aromatic12
or heteroaromatic groups.14–18 Among these heterocy-
cles, pyrimidine scaffolds were used to mimic the
nucleobases, which have led to significant increases in
anti-integrase activity.15,19
with a para-fluorobenzyl group to improve potency.11,12
Two types of metal chelating ligands were selected to re-
place the a,c-diketo acid moiety, including the two-point
ligand picolinamide (Fig. 2, a), which possesses a mod-
erately hard amide O- and a softer pyridine N-binding
site that can form a bidentate ligand–Me2+ complex
with many divalent metal ions,21,22 and the three-point
ligand 8-hydroxy-quinoline-7-carboxamide which is
known as a strong Mg2+ chelating ligand and mimics
the diketo acid group23 (Fig. 2, b). The spatial relation-
ships of the purine substituents were also investigated to
look for the optimal geometry of the inhibitor molecule,
and the chelating ligands were substituted either at 8- (4)
Because the a,c-diketo acid group is biologically labile,
we were interested in replacing it with alternative, more
stable, metal chelating ligands. In this study, we describe
the synthesis and biological evaluation of new DKA
analogs with purine scaffolds.20 The purine core was
used to mimic the nucleobases and it was substituted
7
6
5
1
N
N
8
2
N 9
4
N
3
X
R
N
N
N
N
F
N
F
R
C8 substitution
C6 substitution
Purine core
N
N
N
a: Two-point ligand
F
HN
N
4a, 4b
5a, 5b
X= NH2, N(CH3)2
O
Me2+
b: Three-point ligand
H
N
N
Me2+
O
O
Me2+
Me2+ chelating ligand
R =
Figure 2. Design of purine derivatives incorporating metal chelating ligands as HIV integrase inhibitors.

5744
X. Li, R. Vince / Bioorg. Med. Chem. 14 (2006) 5742–5755
or 6- (5) (Fig. 2) positions on the purine ring. Based on
these considerations, a new series of DKA analogs were
synthesized.
compound 10 with picolinamide 11 under the reported
amidation conditions for 6-halo or 6-arylsulfonate
nucleoside substrates25,26 using palladium catalyst was
unsuccessful. Subsequent attempts using Cu-catalyzed
amidation28 or other palladium ligand29 still could not
give any products. Interestingly, changing the substrate
from N9-benzylated 10 to N3-benzylated 8-bromopurine
12 readily afforded the amidation product 13 under Pd-
catalyzed conditions. The Boc groups were subsequently
removed in trifluoroacetic acid (TFA) to give the 6-ami-
no compound 14 (Scheme 1). The difference in amida-
tion reactivity of N9- (10) and N3- (12) benzylated
purines could be due to the steric hindrance brought
by the benzyl group.
2. Results and discussion
2.1. Chemistry
Palladium-catalyzed amination methodologies for the
formation of C–N bonds have been successfully applied
to the synthesis of nucleosides derivatives.24,25 More
recently, amides were used as coupling substrates in
the Pd-catalyzed amidations with 6-halopurines.26 In
this study, we applied this methodology for the synthesis
of the target C8- (4) and C6- (5) amide substituted
purines.
The N3-benzylated 8-bromopurine 12 was next coupled
with the three-point ligand 8-hydroxy-quinoline-7-car-
boxamide 17. The amide 17 was prepared from the car-
boxylic acid 15 via esterification30 and subsequent
aminolysis (Scheme 2). Under previous Pd-catalyzed
conditions, however, amide 17 did not react with com-
pound 12. We surmised that the palladium catalyst
may lose activity by chelating to the 8-hydroxy-quino-
line moiety (17b). Therefore, the phenol group in com-
pound 15 was protected as a benzyl ether via ester 18
(Scheme 2), and the resulting amide 19 readily reacted
with compound 12 under previous conditions to give
the coupled product 21 (Scheme 3). Subsequent removal
of the Boc groups of 21 in TFA afforded not only 50% of
de-Boc product 22, but also 30% of de-Boc and
The C8-amide substituted purine derivative 4 was syn-
thesized from adenine 6, which was first converted to
the amidation substrate 8-bromopurine 8 by bromina-
tion27 and subsequent alkylation with para-fluorobenzyl
bromide (Scheme 1). Two products were isolated from
the alkylation step in a 1 (8) to 1.3 (9) ratio, and they
were determined as N9- (8) and N3- (9) benzylated pur-
ines by 13C and HMBC NMR analysis (Fig. 3A). The
N9-benzylated 8-bromo adenine 8 was further double
Boc protected (10) to improve solubility and to avoid
the competition between the amino and amide nitrogens
in the next amidation step. Our first attempt to couple
NH2
NH2
N
N
NH2
N
N
NH2
N
Br
Br
N
+
N
a
b
N
N
N
N
N
Br
Method A
N
H
N
H
N
N
7
6
F
F
9
8
Boc
N
Boc
N
N
Pd-catalyzed Amidation
N
c
Br
H2N
+
X
8
N
Method B
or Cu-catalyzed Amidation
N
O
11
F
10
H2N
Boc
N
Boc
N
Boc
N
Boc
NH2
N
N
N
O
N
N
11
N
NH
N
N
e
N
NH
c
d
N
9
Br
N
Method D
O
O
Method C
N
N
N
13
14
12
F
F
F
Scheme 1. Reagents and conditions: (a) Br2, rt, overnight; (b) NaH, DMF, 4-fluorobenzyl bromide, 70 °C, 2 h; (c) (Boc)2O, DMAP, Et3N, DMF, rt,
2 h; (d) Pd2dba3ÆCHCl3, dppf, Cs2CO3, toluene, 90 °C, 4 h; (e) TFA, DCM, rt, overnight.

X. Li, R. Vince / Bioorg. Med. Chem. 14 (2006) 5742–5755
5745
A
NH2
NH2
N
NH2
154.2
122.1
NH2
119.6N
155.4
N
N
N
139.9
N
N
N
N
N
126B.9r
Br
Br
Br
144.5
N
150.2
N
154
N
H
N
H
N
N151.4
46.7
CH2
C1'
52.1
CH2
C1'
132.6
130.9
132.8
130.1
116.4
164.1
163.8
116.3
F
F
F
F
8
9
N9 substitution
N3 substitution
B
Cl
N
F
164.8
F
Cl
131.7N
116.8
129.2
151.3
N
N
N
144.9
N
CH2
C1'
130.6
152.3
151.9N
N
C1'
Cl
Cl
N
50.4
47.5
CH2
143.3
N
122.6
130.6
130.1
116.6
N
N
N
149.1
164.6
152.8
N
N
162.6
F
N
F
26
N7 substitution
25
N9 substitution
Figure 3. 13C and HMBC correlation between C01 proton and purine carbons of (A) N3/N9-benzylated 8-bromo adenine, (B) N9/N7-benzylated
6-chloropurine.
a
b or c
H2N
f
H3CO
N
HO
N
Polymerized acid
N
O
OH
O
OH
O
OH
16
17
15
f
d
H2N
N
e
a
H2N
BnO
N
HO
O
O
N
Pd
N
O
OBn
O
OBn
18
O
OBn
19
17b
20
Scheme 2. Reagents and conditions: (a) liquid NH3, bomb, 70 °C, overnight; (b) H2SO4, MeOH, reflux, 5%; (c) HClSO3, MeOH, reflux, 10%;
(d) BnBr, K2CO3, acetone/DMF, reflux, overnight; (e) 1 N NaOH, MeOH, H2O, rt; (f) SOCl2, heating 1 h.
de-benzyl product 23. Further debenzylation of com-
pound 22 by catalytic hydrogenation only led to the re-
duced quinoline, therefore the benzyl group was
removed in TFA to afford the final product 23 (Scheme 3).
under previous conditions did not afford any product.
We further investigated these amidation conditions with
palladium catalysts (Table 1). We were able to obtain
the amidation product 29 in 29% yield (entry 4), but
we could not get any coupled product using the bigger
amide 19.
Using the same Pd-catalyzed amidation conditions, the
C6-amide substituted purine derivatives 5a,b were suc-
cessfully synthesized (Scheme 4). 6-Chloropurine 24
was alkylated with para-fluorobenzyl bromide to give
N9- (25) and N7- (26) alkylated purines in a 3:1 ratio
(Fig. 3B). Subsequent Pd-catalyzed amidation of N9-
benzylated 6-chloropurine 25 with amide 11 and 19
afforded the corresponding coupled products 5a and
27. The O-benzyl group in compound 27 was removed
in TFA to give the target compound 5b. Amidation of
the N7-benzylated 6-chloropurine 26 with amide 11
As previously described, the N9-benzyl 8-bromopurine
10 did not undergo amidation, therefore, we switched
to amide coupling to prepare the N9-benzyl C8-amide
purines. The coupling intermediate N9-benzylated
8-amino adenine 30 was synthesized from 8-bromo com-
pound 8 by azido replacement followed by reduction31
(Scheme 5). Interestingly, the 8-amino compound 30
did not react with the carboxylic acid 15 under conven-
tional EDC or CDI conditions, and the more reactive

5746
X. Li, R. Vince / Bioorg. Med. Chem. 14 (2006) 5742–5755
Boc
N
Boc
Boc
N
Boc
N
N
N
N
b
N
a
N
NH
N
Method D
H2N
Br
+
Method C
O
N
N
N
OBn
O
OBn
19
21
12
F
F
NH2
NH2
N
N
N
N
c
NH
N
N
NH
N
N
O
N
N
OH
O
OR
23
22 R=Bn
23 R=H
F
F
Scheme 3. Reagents and conditions: (a) Pd2dba3ÆCHCl3, dppf, Cs2CO3, toluene, 90 °C, 4 h; (b) TFA, DCM, rt, 2 days, 22 (50%), 23 (30%); (c) TFA,
DCM, rt, 1 day.
Scheme 4. Reagents and conditions: (a) NaH, DMF, 4-fluorobenzyl bromide, 70 °C, 1 h, 25 (63%), 26 (23%); (b) Pd2dba3ÆCHCl3, dppf, Cs2CO3,
toluene, 90 °C, 5 h; (c) TFA, DCM, rt, 3 days, 84%.
picolinoyl chloride 31 readily reacted with compound 30
to give the C6- and C8-double amide substituted purine
32. In order to avoid the free 6-amino group in the
coupling step, the Boc protected compound 10 was sub-
jected to the sodium azide treatment. Nonetheless, only
6,8-diaminopurine was isolated after the reduction step.
Changing the substrate to 6-chloropurine 25 also result-
ed in the 6,8-diamino compound due to the lack of selec-
tivity between 6-chloro and 8-bromo (33) toward azide
replacement (Scheme 5).

X. Li, R. Vince / Bioorg. Med. Chem. 14 (2006) 5742–5755
Table 1. Amidation conditions of N7-alkylated 6-chloropurine 26 with picolinamide 11
5747
F
F
O
N
P
N
NH
Cl
N
H2N
Pri
iPr
N
O
N
N
N
N
N
11
a
N
iPr
28
29
26
Entry
Catalyst
Ligand
dppf
Base
Solvent
Temp (°C)
Time
Yield (%)
1
2
3
4
Pd2dba3
Pd2dba3
Pd(OAc)2
Pd(OAc)2
Cs2CO3
Cs2CO3
K2CO3
K3PO4
Toluene
Toluene
tBuOH
tBuOH
110
110
110
110
Overnight
18 h
0
0
Ligand 28
Ligand 28
Ligand 28
Overnight
2 days
<10
29
HO
N
O
OH
15
X
NH2
N
NR2
b or c
N
N
N
N
N
N
NH2
Br
a
O
N
Cl
N
N
Method E
O
NH
31
N
N
N
F
e
NH
N
Method G
O
F
F
N
30
8
R= H
10 R= Boc
a
Cl
Cl
N
32
N
N
N
d
N
Br
Method F
N
N
N
F
F
33
25
Scheme 5. Reagents and conditions: (a) i—NaN3, DMF, 90 °C, overnight; ii—10% Pd/C, H2 balloon, MeOH, rt, overnight; (b) EDC, HOBt,
N-methyl morphiline, DMF, rt, overnight; (c) CDI, DMF, rt, overnight; (d) LDA, 1,2-dibromotetrachloroethane, THF, ꢀ70 °C, 2 h; (e) 31, TEA,
DMF, 60 °C, 2 h.
The N,N-dimethyl 8-amide purine derivative 4 was pre-
pared from N,N-dimethyl adenine 34 (Scheme 6). Com-
pound 34 was first benzylated and the N9-benzylated
compound 35 was 8-brominated32 and subsequently con-
verted to 8-amino compound 38. Treatment of compound
38 with picolinoyl chloride 31afforded the desiredmonoa-
mide 4a. Unfortunately, the acyl chloride of carboxylic
acid 15 cannot be obtained by thionyl chloride or oxalyl
chloride treatment, and only polymerized product was
isolated. Changing to the benzyl protected acid 20 also
afforded the polymerized products (Scheme 2). Unable
to get the acyl chloride, the target compound 4b was pre-
pared by aminolysis of the benzyl ester 18 with 8-amino
compound 38 in the presence of sodium hydride. The
resulting coupled product 39 was debenzylated in TFA
to give target compound 4b (Scheme 6).
2.2. Structure–activity relationship studies
All the final compounds were tested for their activities
against HIV integrase in microtiter and radioactive gel as-
says (Table 2). These DKA analogs with purine core
inhibited recombinant HIV integrase at low micromolar
range, but are less potent than the known a,c-diketo acid
inhibitors (Table 2, 1). Our SAR results indicated that the
two types of metal chelating ligands used in this study did
not lead to significant difference in potency; the three-
point Mg2+ ligand 8-hydroxy-quinoline-7-carboxamide,
which mimics diketo acid, and the two-point Me2+ ligand
picolinamide showed 10–45% and 20–50% inhibition at
10 lM, respectively. Our results also indicated that the
spatial relationship between the two purine substituents
was important for potency, purines containing the

5748
X. Li, R. Vince / Bioorg. Med. Chem. 14 (2006) 5742–5755
H3C
N
CH3
N
N
H3C
CH3
F
N
N
N
H3C
N
CH3
N
N
a
N
N
N
+
N
Method A
N
H
N
34
36
F
35
Method F
b
Cl
H3C
N
CH3
N
H3C
N
CH3
H3C
N
CH3
N
N
N
N
N
O
N
31
N
N
NH
d
N
c
NH2
Br
O
N
Method G
N
Method E
N
N
F
F
F
4a
37
38
BnO
e
N
OBn
O
18
H3C
N
CH3
H3C
CH3
N
N
N
f
N
NH
N
N
N
NH
N
N
N
O
OH
O
N
OBn
4b
F
39
F
Scheme 6. Reagents and conditions: (a) NaH, DMF, 4-fluorobenzyl bromide, rt, 2 h; (b) LDA, 1,2-dibromotetrachloroethane, THF, ꢀ70 °C;
(c) i—NaN3, DMF, 80 °C, 8 h; ii—H2 balloon, 10% Pd/C, MeOH, overnight; (d) 31, TEA, DMF, 60 °C, 3 h; (e) NaH, rt, DMF; (f) TFA, rt, 2 days.
three-point ligand were slightly more active than the two-
point ligand containing purines when the para-
fluorobenzyl group was N3-substituted (14 and 23); but
were 2-fold less active when the para-fluorobenzyl group
was N9-substituted (Table 2, 4a,b and 5a,b). The results
also demonstrated that the position of para-fluorobenzyl
group was crucial for potency, and the N7-substituted
purine (29) was 2- to 3-fold less active than the N9
(4a and 5a) and N3 (14) substituted ones. Interestingly,
the C6,C8-dipicolinamide substituted purine 32 showed
very potent inhibitory activity against integrase at
10 lM, and is 2- to 3-fold more active than the C8- (4a)
or C6- (5a) mono-picolinamide substituted purines. This
result suggested that an additional metal chelating ligand
could contribute to the increase in potency, and the spatial
arrangement of the ligand could be crucial.
results were shown as the combined activity of both
30-processing and strand transfer steps given as percent-
age of inhibition at 10 lM. Each assay reactions was
done in duplicate, and the data were given as an average
of the duplicate.
3.2. Chemistry
All reactions were performed in septum-sealed flasks un-
der argon atmosphere. Progress of the reaction was fol-
lowed by TLC. Organic solutions were dried over
MgSO4 and evaporated on a rotary evaporator under
reduced pressure. Melting points were determined on a
1
Mel-Temp II apparatus and are uncorrected. H and
13C NMR spectra were obtained using Varian 300 and
600 spectrometers operating at field strength of 300
and 600 MHz. Chemical shifts (d) are reported in parts
per million (ppm) and coupling constants (J) in hertz.
Standard and peak multiplicities are designated as fol-
lows: s, singlet; d, doublet; dd, doublet of doublets; t,
triplet; q, quartet; p, pentet; br s, broad singlet; and
m, multiplet. Mass spectra were determined using Bru-
ker BioTOF II and Agilent LC-TOF spectrometers.
Flash chromatography was performed using 200–300
mesh silica gel. All commercial solvents were of reagent
3. Experimental
3.1. Biological assays
All the synthesized compounds were tested in a dual
microtiter assay and radioactive gel assay (data
provided by Southern Research Institute33,34). The

X. Li, R. Vince / Bioorg. Med. Chem. 14 (2006) 5742–5755
Table 2. Anti-HIV integrase activities of purine derivatives in microtiter and gel assays
5749
Compound
Structure
Inhibition at 10 lMa (%)
Microtiter assay (SD)
Gel assay (SD)
O
OH
N
OH
O
1
79.0 (1.2)
61.4 (1.2)
F
NH2
N
H
N
N
N
N
O
N
14
27.0 (3.3)
31.0 (1.2)
F
NH2
N
N
N
N
NH
N
O
OH
23
41.4 (3.6)
44.6 (2.4)
F
H3C
CH3
N
N
N
NH
N
N
4a
4b
5a
5b
N
40.2 (8.9)
51.4 (9.0)
O
F
H3C
N
CH3
N
N
N
OH
O
N
NH
N
26.1 (5.6)
20.2 (4.3)
11.2 (4.3)
19.3 (4.9)
41 (12.5)
30.1 (2.6)
F
O
N
NH
N
N
N
N
F
OH
O
N
N
NH
N
N
N
F
(continued on next page)

5750
X. Li, R. Vince / Bioorg. Med. Chem. 14 (2006) 5742–5755
Table 2 (continued)
Compound
Structure
Inhibition at 10 lMa (%)
Microtiter assay (SD)
Gel assay (SD)
O
N
NH
N
O
N
N
N
NH
F
32
29
72.2 (8.9)
83.6 (6.5)
N
F
O
N
13.1 (5.5)
17.1 (7.8)
NH
N
N
N
N
a Mean and standard deviation (SD) were calculated from duplicate experiments.
grade or better and used directly as supplied. The vials
and caps for the amidation reactions were purchased
from Biotage, VA. Yields refer to purified products
and were not optimized.
3.1 mmol) were dissolved in 10 mL DMF and treated
with triethylamine (650 lL, 4.65 mmol) and DMAP
(20 mg). The mixture was stirred at room temperature
for 2 h. After the reaction was complete, the mixture
was concentrated and purified by flash chromatography
(EtOAc/hexanes 1:10–1:5) to give the title compound as
white crystal. Yield: 95%; mp 176–178 °C; 1H NMR
(CDCl3): d 8.85 (1H, s), 7.36 (2H, dd, J = 4.5, 9 Hz),
7.03 (2H, t, J = 9 Hz), 5.44 (2H, s), 1.45 (18H, s); 13C
NMR (CDCl3): d 164.4, 161.1, 154.5, 152.5, 150.4,
149.1, 133.5, 130.5, 130.1, 129.1, 116.3, 116.0, 84.3,
47.6, 36.8, 28.1; MS (ESI): m/z 544.09, 546.09 [M+Na]+.
3.3. 8-Bromo adenine (7)
The title compound was prepared as described.27 Yield:
70%; mp >260 °C; NMR (DMSO-d6): d 8.08 (1H, s),
7.39 (2H, s); 13C NMR (DMSO-d6): d 154.5, 152.7,
150.6, 120.6; MS (ESI): m/z 214, 216 [M+H]+.
3.4. Method A: Preparation of 8-bromo 9-(4-fluorobenzyl)
adenine (8) and 8-bromo 3-(4-fluorobenzyl) adenine (9)
3.6. N,N-(di-tert-Butoxycarbonyl) 8-bromo 3-(para-
fluorobenzyl) adenine (12)
A suspension of 8-bromo adenine (7, 1.07 g, 5 mmol) in
25 mL anhydrous DMF was treated with sodium
hydride (220 mg, 5.5 mmol) and heated at 70 °C for
30 min. 4-Fluorobenzyl bromide (1.04 g, 685 lL,
5.5 mmol) was added dropwise at the same temperature.
The reaction was complete after 2 h of heating. The solid
precipitate was filtered and the filtrate was concentrated
in vacuo. The crude product was purified by flash
chromatography (CHCl3/MeOH 40:1–20:1) to give 8
The title compound was prepared from 8-bromo 3-(4-
fluorobenzyl) adenine (9) according to method B. Yield:
80%; mp 155–157 °C; 1H NMR (CDCl3): d 7.97 (1H, s),
7.08 (2H, t, J = 8 Hz), 6.74 (2H, t, J = 8 Hz), 5.33 (2H,
s), 1.12 (18H, s); 13C NMR (CDCl3): d 161.8, 156.9,
152.6, 150.1, 145.4, 143.7, 139.2, 135.5, 131.2, 117.0,
116.7, 110.0, 84.9, 54.1, 28.1; MS (ESI): m/z 522.1,
524.1 [M+H]+.
1
as white crystal. Yield: 37%; mp 230–231 °C; H NMR
(DMSO-d6): d 8.14 (1 H, s), 7.44 (2H, br s), 7.27 (2H,
m), 7.16 (2H, m), 5.32 (2H, s); 13C NMR (DMSO-d6):
d 163.8, 160.6, 155.4, 153.8, 151.5, 132.8 (2), 130.1 (2),
126.9, 119.6, 116.4, 116.1, 46.7; MS (ESI): m/z 322.0,
324.0 [M+H]+, 344.0, 346.0 [M+Na]+, and 9 as white
crystal. Yield: 48%; mp 216–217 °C; 1H NMR
(DMSO-d6): d 8.53 (1H, s), 8.24 (1H, br s), 8.08 (1H,
br s), 7.47 (2H, dd, J = 6.0, 9.0 Hz), 7.17 (2H, t,
J = 9.0 Hz), 5.42 (2H, s); 13C NMR (DMSO-d6): d
164.1, 160.8, 154.2, 150.2, 144.5, 140.0, 132.6 (2), 130.9
(2), 122.2, 116.3, 116.1, 52.2; MS (ESI): 322.0, 324.0
[M+H]+, 344.0, 346.0 [M+Na]+.
3.7. Method C: Preparation of N,N-(di-tert-butoxycar-
bonyl)-3-(4-fluorobenzyl) 8-picolinamide adenine (13)
A
10 mL vial was charged with Pd2dba3ÆCHCl3
(20.7 mg, 0.02 mmol) and dppf (33.3 mg, 0.06 mmol).
The vial was sealed with Teflon cap and filled with argon
to evacuate air. A solution of N,N-(di-tert-butoxycar-
bonyl) 8-bromo 3-(para-fluorobenzyl) adenine (12,
208.8 mg, 0.4 mmol) in 8 mL dry toluene was added
via cannula, followed by the addition of picolinamide
(11, 53.7 mg, 0.44 mmol) and cesium carbonate
(182.5 mg, 0.56 mmol). The air was again evacuated
with argon. The resulting mixture was heated at 80 °C
for 4 h. After cooling down, the mixture was concentrat-
ed and purified by flash chromatography (CHCl3/
MeOH 40:1) to give the title compound as white solid.
Yield: 93%; mp 196–198 °C; 1H NMR (CDCl3): d
11.01 (1H, s), 8.65 (1H, m), 8.36 (1H, dt, J = 0.9,
3.5. Method B: Preparation of N,N-(di-tert-butoxycar-
bonyl) 8-bromo 9-(4-fluorobenzyl) adenine (10)
8-Bromo 9-(4-fluorobenzyl) adenine (8, 500 mg,
1.55 mmol) and di-tert-butyl dicarbonate (470 mg,

X. Li, R. Vince / Bioorg. Med. Chem. 14 (2006) 5742–5755
5751
7.2 Hz), 8.25 (1H, s), 7.93 (1H, td, J = 1.5, 7.5 Hz), 7.55–
7.46 (3H, m), 7.07 (2H, t, J = 6.0 Hz), 5.75 (2H, s), 1.47
(18H, s); 13C NMR (CDCl3): d 165.0, 163.5, 161.6,
158.4, 150.4, 149.2, 148.4, 142.6, 137.9, 137.6, 135.1,
131.1 (2), 129.3 (2) 127.2, 123.1, 116.8 (2), 84.3, 53.7,
28.2; MS (ESI): m/z 564.3 [M+H]+.
0.5 mL DMSO was heated at 70 °C for 30 min. Benzyl
bromide (180 lL, 1.5 mmol) was then added and the
resulting mixture was heated at 70 °C overnight. After
cooling down, the undissloved solid was filtered, and
the filtrate was concentrated and purified by flash chro-
matography (hexanes/EtOAc 15:1) to give the title com-
1
pound as yellow oil. Yield: 98%; H NMR (CDCl3): d
3.8. Method D: Preparation of 3-(4-fluorobenzyl)
8-picolinamide adenine (14)
9.03 (1H, s), 8.16 (1H, d, J = 9 Hz), 7.89 (1H, d,
J = 9 Hz), 7.58 (2H, m), 7.45 (2H, m), 7.37 (8H, m),
5.54 (2H, s), 5.39 (2H, s); MS (ESI): m/z 370.14 [M+H]+.
A solution of N,N-(di-tert-butoxycarbonyl)-3-(4-fluo-
robenzyl) 8-(picolinamide) adenine (13, 210 mg,
0.37 mmol) in 15 mL DCM was treated with 1 mL
TFA and stirred at room temperature. The reaction
was complete after two days and the mixture was con-
centrated. The resulting solid was redissolved in 20 mL
DCM, poured into 10 mL ice-water, and neutralized
with saturated NaHCO3 to pH 8–9. The aqueous layer
was extracted with DCM. The organic portions were
combined and dried over MgSO4. After concentration,
the crude product was purified by flash chromatography
(CHCl3/MeOH 30:1) to give the title compound as white
solid. Yield: 63%; mp >160 °C (dec); 1H NMR (DMSO-
d6): d 10.46 (1H, br s), 8.71 (1H, d, J = 4.5 Hz), 8.53 (1H,
s), 8.18 (1H, d, J = 7.8 Hz), 8.08 (1H, t, J = 7.8 Hz), 7.80
(2H, br s), 7.69 (1H, m), 7.56 (2H, m), 7.19 (2H, t,
J = 9 Hz), 5.48 (2H, s); 13C NMR (DMSO-d6): d
160.9, 153.6, 150.1, 149.2, 143.2, 139.0, 133.0 (2), 131.1
(2), 127.8, 122.8, 116.3, 116.0, 52.0; MS (ESI): m/z
364.2 [M+H]+, 386.2 [M+Na]+; HRMS (ESI): Calcd
for (C18H14FN7O)H+: 364.1317, found: 364.1317 MH+.
3.12. 8-Benzyloxy-quinoline-7-carboxamide (19)
8-Benzyloxy-quinoline-7-carboxylic acid benzyl ester
(18, 800 mg) was treated with 10 mL liquid ammonia.
The mixture was heated in a sealed steel bomb at
70 °C overnight. The excess ammonia was evaporated
in the hood, and the crude product was purified by flash
chromatography (hexanes/EtOAc 5:1–2:1) to give the ti-
tle compound as cream-colored crystal. Yield: 80%; mp
1
128–130 °C; H NMR (CDCl3): d 9.02 (1H, dd, J = 1.8,
4.5 Hz), 8.24 (1H, d, J = 9 Hz), 8.19 (1H, dd, J = 1.8,
9 Hz), 8.05 (1H, br s), 7.64 (1H, d, J = 9 Hz), 7.57–
7.49 (3H, m), 7.42–7.37 (3H, m), 5.96 (1H, br s), 5.61
(2H, s); 13C NMR (CDCl3): d 167.1, 154.6, 150.0,
142.8, 136.5 (2), 132.0, 128 (m), 127.8, 124.7, 123.6,
122.9, 78.7; MS (ESI): m/z 279.10 [M+H]+.
3.13. 8-Benzyloxy-quinoline-7-carboxylic acid (20)
8-Benzyloxy-quinoline-7-carboxylic acid benzyl ester
(18, 1.8 g, 4.8 mmol) was dissolved in 10 mL methanol
and 25 mL THF. The solution was treated with 6 mL
of 1 N NaOH and stirred at room temperature for 5 h.
The organic solvent was removed in vacuo and the crude
product was redissloved in 15 mL water. The aqueous
portion was washed with DCM and subsequently acidi-
fied to pH 4–5 with 1 N HCl under ice bath. The crystal
precipitate was collected by filtration and washed with
DCM to give the title compound as cream-colored crys-
3.9. 8-Hydroxy-quinoline-7-carboxylic acid methyl ester
(16)
The title compound was prepared as described.30 Yield:
1
10%; H NMR (CDCl3): d 11.92 (1H, br s), 8.96 (1H,
dd, J = 1.8, 4.2 Hz), 8.09 (1H, dd, J = 1.5, 8.4 Hz),
7.87 (1H, d, J = 8.7 Hz), 7.51 (1H, dd, J = 4.2, 8.4 Hz),
7.26 (1H, d, J = 8.7 Hz), 4.01 (3H, s); 13C NMR
(CDCl3): d 170.9, 160.1, 149.9, 139.9, 135.9, 132.5,
125.6, 124.1, 117.9, 109.4, 52.9.
1
tal. Yield: 73%; mp 176–180 °C; H NMR (DMSO-d6):
d 13.18 (1H, s), 9.01 (1H, d, J = 2.7 Hz), 8.43 (1H, d,
J = 8 Hz), 7.77 (2H, s), 7.66–7.58 (3H, m), 7.41–7.32
(3H, m), 5.42 (2H, s); 13C NMR (DMSO-d6): d 168.3,
154.4, 151.1, 143.1, 138.4, 137.1, 131.5, 128 (m), 126.4,
124.2, 123.6, 77.1; MS (ESI): m/z 280.0 [M+H]+, 302.0
[M+Na]+.
3.10. 8-Hydroxy-quinoline-7-carboxamide (17)
8-Hydroxy-quinoline-7-carboxylic acid methyl ester (16,
80 mg, 0.39 mmol) was treated with 10 mL liquid
ammonia. The mixture was heated in a sealed steel
bomb at 70 °C overnight. The excess ammonia was
evaporated in the hood. The crude product was recrys-
tallized with DCM to give the title compound as faint
3.14. N,N-(di-tert-Butoxycarbonyl)-3-(4-fluorobenzyl)-8-
(8-benzyloxy-quinoline-7-carboxamide) adenine (21)
1
yellow crystal. Yield: 55%; mp 190–192 °C; H NMR
Compound 21 was prepared from N,N-(di-tert-butoxy-
carbonyl) 8-bromo 3-(para-fluorobenzyl) adenine (12)
and 8-benzyloxy-quinoline-7-carboxamide (19) accord-
ing to method C and purified by flash chromatography
(hexanes/EtOAc 5:1–1:1) to give the title compound as
white solid. Yield: 74%; mp 185–187 °C; 1H NMR
(CDCl3): d 11.27 (1H, s), 9.07 (1H, dd, J = 1.8,
4.5 Hz), 8.35 (1H, d, J = 9 Hz), 8.23 (1H, d,
J = 1.8 Hz), 8.21 (1H, s), 7.67 (1H, d, J = 9 Hz), 7.60–
7.53 (3H, m), 7.47 (2H, m), 7.24 (4H, m), 7.08 (2H,
m), 5.76 (2H, s), 5.70 (2H, s), 1.48 (18H, s); 13C NMR
(CDCl3): d 164.0, 163.6, 162.4, 162.1, 158.2, 153.7,
(CDCl3): d 8.86 (1H, dd, J = 1.5, 4.5 Hz), 8.19 (2H,
dd, J = 1.5, 7.5 Hz), 8.15 (1H, d, J = 9 Hz), 7.7 (1H, br
s), 7.55 (1H, dd, J = 4.5, 9 Hz), 7.39 (1H, d, J = 9 Hz);
MS (ESI): m/z 211.04 [M+Na]+.
3.11. 8-Benzyloxy-quinoline-7-carboxylic acid benzyl
ester (18)
A suspension of 8-hydroxy-quinoline-7-carboxylic acid
(15, 95 mg, 0.5 mmol) and anhydrous potassium car-
bonate (194 mg, 1.4 mmol) in 20 mL acetone and

5752
X. Li, R. Vince / Bioorg. Med. Chem. 14 (2006) 5742–5755
150.4, 150.0, 142.6, 142.2, 137.1, 136.3, 135.4, 134.8,
132.1, 130.9, 129.7, 129.3, 128.8, 128.6, 127.8, 125.1,
123.7, 122.9, 116.5, 116.3, 83.9, 78.8, 53.4, 27.8; MS
(ESI): m/z 720.29 [M+H]+.
7.06 (2H, t, J = 8.4 Hz), 5.46 (2H, s); 13C NMR
(CDCl3): d 163.6, 161.9, 161.4, 153.1, 152.0, 149.2,
149.0, 148.3, 142.6, 137.8, 131.0, 129.8 (2), 127.2,
123.1, 122.8, 116.3, 116.1, 46.8; MS (ESI): m/z 349.12
[M+H]+; HRMS (ESI): Calcd for (C18H13FN6O)Na+:
371.1027, found: 371.1021 [M+Na]+.
3.15. 3-(4-Fluorobenzyl) 8-(8-benzyloxy-quinoline-7-
carboxamide) adenine (22) and 3-(4-fluorobenzyl)
8-(8-hydroxy-quinoline-7-carboxamide) adenine (23)
3.18. 6-(8-Benzyloxy-quinoline-7-carboxamide)-9-
(4-fluorobenzyl) purine (27)
The title compounds were prepared from compound 21
according to method D. After the reaction was com-
plete, the mixture was concentrated and the resulting
semi-solid was dissolved in 25 mL DCM and washed
with saturated NaHCO3. Some yellow solid was precip-
itated out while mixing with the basic solution. The solid
was collected by filtration and recrystallized with meth-
anol to give 23 as bright yellow crystal. Yield: 30%;
Compound 27 was prepared from compound 25 and 8-
benzyloxy-quinoline-7-carboxamide (19) according to
method C. Yield: 68%; mp 276–278 °C; 1H NMR
(CDCl3): d 11.42 (1H, s), 9.06 (1H, dd, J = 1.8, 4.2 Hz),
8.84 (1H, s), 8.34 (1H, d, J = 8.4 Hz), 8.21 (1H, d,
J = 8.4 Hz), 7.92 (1H, s), 7.66 (1H, d, J = 9 Hz), 7.54
(1H, dd, J = 4.2, 8.4 Hz), 7.41 (2H, d, J = 6.6 Hz), 7.34
(2H, dd, J = 4.8, 8.4 Hz), 7.11–7.06 (5H, m), 5.89 (2H,
s), 5.42 (2H, s); 13C NMR (CDCl3): d 163.6, 162.5,
161.9, 154.0, 152.9, 151.9, 149.9, 149.5, 142.6, 142.2,
136.3, 135.6, 132.2, 131.2 (2), 129.8 (2), 129.7, 128.6,
128.2, 127.8, 124.9, 123.6, 123.2, 122.9, 116.2, 116.1,
78.7, 46.7; MS (ESI): m/z 505.1 [M+H]+, 527.1 [M+Na]+.
1
mp >250 °C (dec); H NMR (DMSO-d6): d 13.09 (1H,
br s), 8.89 (3H, s), 8.82 (1H, d, J = 7.8 Hz), 8.68 (1H,
s), 8.14 (1H, d, J = 9 Hz), 7.95 (1H, m), 7.57 (2H, m),
7.23 (2H, t, J = 9 Hz), 7.16 (1H, d, J = 7.8 Hz), 5.57
(2H, s); 13C NMR (DMSO-d6): d 170.8, 168.4, 161.8,
158.5, 158.3, 148.1, 131.8, 130.0 (2), 117.0, 116.5,
116.4, 110.6, 52.0; HRMS (ESI): Calcd for (C22H17
FN7O2)+: 430.1422, found: 430.1435 MH+. The concen-
trated organic portion was purified by flash chromatog-
raphy (hexanes:EtOAc 5:1–1:2) to give compound 22 as
white solid. Yield: 49%; mp 228–230 °C; 1H NMR
(DMSO-d6): d 10.82 (1H, br s), 9.02 (1H, s), 8.43 (2H,
d, J = 9 Hz), 7.79 (2H, m), 7.68 (2H, m), 7.63 (1H, m),
7.57 (2H, m), 7.48 (1H, br s), 7.24 (3H, m), 7.112 (1H,
br s), 5.53 (2H, br s), 5.44 (2H, br s); 13C NMR
(DMSO-d6): d 163.3, 161.7, 153.5, 150.9, 150.1, 142.8,
137.2, 130.9 (2), 129.2, 128.8, 128.6, 126.9, 123.2,
116.1, 115.9, 51.7; MS (ESI): m/z 520.2 [M+H]+; 542.1
[M+Na]+. Compound 22 was further hydrolyzed with
TFA in DCM to give compound 23, yield 30%.
3.19. 6-(8-Hydroxy-quinoline-7-carboxamide)-9-
(4-fluorobenzyl) purine (5b)
A solution of compound 27 (40 mg, 0.08 mmol) in
15 mL DCM was treated with 1 mL TFA. The mixture
was stirred at room temperature for 3 days. The excess
TFA was removed in vacuo, and the resulting semi-oil
was crystallized with DCM to give the title compound
as bright yellow crystals. Yield: 62%; mp 238–240 °C;
1H NMR (DMSO-d6): d 8.90 (1H, d, J = 4.2 Hz), 8.76
(1H, d, J = 9 Hz), 8.72 (2H, s), 8.13 (1H, d,
J = 8.4 Hz), 7.89 (1H, dd, J = 4.8, 7.8 Hz), 7.44 (2H,
dd, J = 5.4, 7.8 Hz), 7.25 (1H, d, J = 8.4 Hz), 7.18 (2H,
t, J = 9 Hz), 5.51 (2H, s); 13C NMR (DMSO-d6): d
163.3, 161.7, 161.3, 151.7, 149.3, 148.7, 145.9, 144.8,
142.8, 136.9, 136.9, 133.2, 133.1, 129.6, 124.8, 123.4,
116.4, 116.2, 114.8, 46.8; MS (ESI): m/z 415.05
[M+H]+; 437.03 [M+Na]+, 453.00 [M+K]+; HRMS
(ESI): Calcd for (C22H15FN6NaO2)+: 437.1133, found:
4378.1143 [M+Na]+.
3.16. 9-(4-Fluorobenzyl) 6-chloropurine (25) and
7-(4-fluorobenzyl) 6-chloropurine (26)
The title compounds were prepared from 6-chloropurine
(24) according to method A. Compound 25 was obtained
1
as white crystal. Yield: 63%; mp 131–132 °C; H NMR
(CDCl3): d 8.79 (1H, s), 8.10 (1H, s), 7.33 (2H, dd,
J = 6, 9 Hz), 7.07 (2H, t, J = 9 Hz), 5.43 (2H, s); 13C
NMR (CDCl3): d 164.6, 161.3, 152.4, 151.9, 151.3,
144.9, 131.7, 130.6 (2), 130.0, 116.7, 116.4, 47.6; MS
(ESI): m/z 285.1 [M+Na]+. Compound 26 was obtained
3.20. 6-Picolinamide-7-(4-fluorobenzyl) purine (29)
A 10 mL vial was charged with Pd(OAc)2 (3.8 mg,
0.017 mmol) and ligand 28 (14.2 mg, 0.03 mmol). The vial
was filled with argon to evacuate air. About 4 mL of
t-BuOH (solid) was added and the tube was filled with ar-
gon and the air evacuated. The mixture was heated at
30–40 °C for 10 min for t-BuOH to melt. of the red-
brownish solution were added compound 27 (105 mg,
0.4 mmol), picolinamide (11, 73 mg, 0.6 mmol), phenyl
boronic acid (3.2 mg, 0.026 mmol), and potassium phos-
phate (150 mg, 0.71 mmol). The vial was sealed with Tef-
lon cap, filled with argon, and heated at 100–110 °C. The
reaction was complete after 3 days of heating (starting
materials were all consumed, several new spots observed
on TLC). After cooling down, the mixture was concen-
trated and purified by flash chromatography (hexanes/
EtOAc 1:1–CHCl3/MeOH 50:1) to give the title
1
as white crystal. Yield: 23%; mp 167–168 °C; H NMR
(CDCl3): d 8.89 (1H, s), 8.26 (1H, s), 7.18 (2H, m), 7.09
(2H, m), 5.68 (2H, s); 13C NMR (CDCl3): d 164.6,
162.2, 161.3, 152.8, 149.1, 143.3, 130.6 (2), 129.2 (2),
122.6, 116.8, 116.5, 50.4; MS (ESI): m/z 263.04 [M+H]+.
3.17. 6-Picolinamide 9-(4-fluorobenzyl) purine (5a)
Compound 5a was prepared from compound 25 and
picolinamide 11 according to Method C. Yield: 68%;
mp 222–224 °C; 1H NMR (CDCl3): d 11.24 (1H, s),
8.89 (1H, s), 8.70 (1H, d, J = 4.8 Hz), 8.39 (1H, d,
J = 7.8 Hz), 8.01 (1H, s), 7.95 (1H, t, J = 7.8 Hz), 7.54
(1H, dd, J = 4.8, 7.0 Hz), 7.34 (2H, dd, J = 6, 8.4 Hz),

X. Li, R. Vince / Bioorg. Med. Chem. 14 (2006) 5742–5755
5753
compound. Yield: 29%; mp 201–204 °C; 1H NMR
(CDCl3): d 10.28 (1H, br s), 8.96 (1H, s), 8.63 (1H, dt,
J = 4.2, 0.9 Hz), 8.30 (1H, dt, J = 6.9, 0.9 Hz), 8.21 (1H,
s), 7.97 (1H, dt, J = 1.8, 7.5 Hz), 7.59 (1H, ddd, J = 1.2,
4.8, 7.8 Hz), 7.02 (2H, m), 6.94 (2H, t, J = 9 Hz), 5.55
(2H, s); 13C NMR (DMSO-d6): d 162.9, 161.3, 152.5,
150.9, 149.3, 138.8, 133.4, 129.3 (2), 128.2, 123.4, 116.0,
115.9, 49.8; MS (ESI): m/z 349.2 [M+H]+, 371.2
[M+Na]+; HRMS (ESI): Calcd for (C18H13FN6ONa)+:
371.1027, found: 371.1053 [M+Na]+.
um solution (844 lL, 2.1 mmol) in 2 mL anhydrous
THF at ꢀ70 °C under argon for 1 h. To the LDA
solution was added a solution of 9-(4-fluorobenzyl)
6-chloropurine (25, 394 mg, 1.5 mmol) in 4.5 mL
THF at ꢀ70 °C. The mixture turned dark after add-
ing 25. After stirring for 1 h at ꢀ70 °C, a solution
of dibromotetrachloroethane (977 mg, 3 mmol) in
3 mL THF was added dropwise. The reaction was
complete after stirring at ꢀ70 °C for 1 h. The mixture
was gradually warmed up to room temperature, and
2 mL of saturated NH4Cl was added. The mixture
was concentrated to remove THF. The resulting resi-
due was redissolved in DCM and washed with water.
The organic layer was separated and dried over
MgSO4. After concentration, the crude product was
purified by flash chromatography (hexanes/EtOAc
5:1) to give the title compound as white crystal. Yield:
3.21. Method E: Preparation of 8-amino 9-(4-fluoro-
benzyl) adenine (30)
8-Bromo 9-(4-fluorobenzyl) adenine (8, 450 mg,
1.4 mmol) and sodium azide (182 mg, 2.8 mmol) were
suspended in 10 mL DMF and heated at 100 °C. The
reaction was complete after 9 h of heating. The reaction
mixture was concentrated in vacuo to nearly dryness
and co-evaporated with toluene. The crude product
was then dissolved in 40 mL methanol and treated with
200 mg 10% Pd/C. The mixture was stirred under hydro-
gen balloon at room temperature overnight. After the
reaction was complete, the catalyst was filtered and the
concentrated filtrate was purified by flash chromatogra-
phy (CHCl3/MeOH 20:1–10:1) to give the title com-
pound as faint yellow solid. Yield: 70%; mp >232 °C
1
55%; mp 143–144 °C; H NMR (CDCl3): d 8.74 (1H,
s), 7.38 (2H, dd, J = 3.0, 9 Hz), 7.01 (2H, t, J = 9 Hz),
5.45 (2H, s); 13C NMR (CDCl3): d 164.5, 161.2,
153.0, 152.4 (2), 149.7, 134.2, 132.0, 130.2 (m),
116.4, 116.1, 48.0; MS (ESI): m/z 362.8, 364.8
[M+Na]+.
3.24. N,N-Dimethyl 9-(4-fluorobenzyl) adenine (35) and
N,N-dimethyl 7-(4-fluorobenzyl) adenine (36)
1
(dec); H NMR (DMSO-d6): d 7.93 (1H, s), 7.30 (2H,
The title compounds were prepared from N,N-dimethyl
adeine (34) at room temperature according to Method
A. The reaction was complete after stirring at room tem-
perature for 8 h. Compound 35 was obtained as white
crystal. Yield: 68%; mp 129–130 °C; 1H NMR (CDCl3):
d 8.38 (1H, s), 7.69 (1H, s), 7.26 (2H, m), 7.02 (2H, t,
J = 9 Hz), 5.32 (2H, s), 3.53 (6H, br s); 13C NMR
(CDCl3): d 164.2, 160.9, 155.0, 152.7, 150.6, 137.9,
131.9, 129.6, 120.1, 116.2 (2), 46.5, 38.8; MS (ESI): m/
z 272.0 [M+H]+, 294.0 [M+Na]+, and 36 as white crys-
dd, J = 3.0, 9.0 Hz), 7.12 (2H, t, J = 9 Hz), 6.62 (2H,
s), 6.59 (2H, s), 5.18 (2H, s); 13C NMR (DMSO-d6): d
163.6, 160.4, 152.6 (2), 150.3, 149.5, 133.8, 130.0 (2),
117.6, 116.1, 457.8, 43.6; MS (ESI): m/z 259.1
[M+H]+, 281.1 [M+Na]+.
3.22. Method G: Preparation of 6,8-dipicolinamide
9-(4-fluorobenzyl) purine (32)
1
The picolinoyl chloride 31 was prepared by refluxing
picolinic acid (95 mg, 0.77 mmol) in 1 mL thionyl chlo-
ride for 1 h. The excess thionyl chloride was removed in
vacuo and co-evaporated with toluene. The crude product
(31, white solid) was directly used for the next step. In a
10 mL flask were added 8-amino 9-(4-fluorobenzyl) 6-
aminopurine (30, 80 mg, 0.3 mmol) and 2 mL DMF. To
the solution were added triethylamine (100 lL) and a
solution of compound 31 in 2 mL DMF. The resulting
mixture was heated at 60 °C for 2 h. After concentration,
the crude product was purified by flash chromatography
(CHCl3/MeOH 50:1) to give the title compound as faint
yellow solid. Yield: 43%; mp 241–243 °C; 1H NMR
(DMSO-d6, d ppm): (hard to dissolve in DMSO) 12.9
(0.5H, br s); 11.5 (0.5H, br s); 11.1 (0.5H, br s); 10.9
(0.5H, br s); 8.7 (3H, m); 8.3 (1H, m); 8.1 (2H, m); 7.9
(1H, m); 7.7 (2H, s); 7.6 (2H, s); 7.2–7.06 (3H, m); 5.4
(2H, s); MS (ESI): m/z 469.0 [M+H]+, 491.0 [M+Na]+;
HRMS (ESI): Calcd for (C24H17FN8O2Na)+ 491.1351
[M+Na]+, found: 491.1356 [M+Na]+.
tal. Yield: 30%; mp 124–126 °C; H NMR (CDCl3): d
8.03 (1H, s), 7.95 (1H, s), 7.40 (2H, m), 7.03 (2H, m),
5.49 (2H, s), 3.94 (3H, s), 3.34 (3H, s); 13C NMR
(CDCl3): d 164.5, 161.3, 153.3, 152.8, 150.6, 140.8,
130.7, 130.3, 121.9, 116.4, 116.1, 52.5, 40.1, 38.3; MS
(ESI): m/z 272.0 [M+H]+, 294.0 [M+Na]+.
3.25. N,N-Dimethyl 8-bromo 9-(4-fluorobenzyl) adenine
(37)
The title compound was prepared from N,N-dimethyl 9-
(4-fluorobenzyl) adenine (35) according to Method F.
Compound 37 was obtained as white crystal. Yield:
1
73%; mp 154–155 °C; H NMR (CDCl3): d 8.28 (1H,
s), 7.28 (2H, dd, J = 6, 9 Hz), 6.96 (2H, t, J = 9 Hz),
5.30 (2H, s), 3.45 (6H, br s); 13C NMR (CDCl3): d
164.2, 160.9, 153.8, 152.8, 152.1, 131.4, 129.8 (2),
124.5, 116.0 (2), 46.9, 38.8; MS (ESI): m/z 350.0, 352.0
[M+H]+, 372.0, 374.0 [M+Na]+.
3.26. N,N-Dimethyl 8-amino 9-(4-fluorobenzyl) adenine
(38)
3.23. Method F: Preparation of 8-Bromo 9-(4-fluoro-
benzyl) 6-chloropurine (33)32
Compound 38 was prepared from N,N-dimethyl
8-bromo 9-(4-fluorobenzyl) adenine (37) according to
method E. Compound 38 was obtained as white crys-
LDA solution was freshly made by stirring diisopro-
pylamine (295 lL, 2.1 mmol) and 2.5 M n-butyl lithi-

5754
X. Li, R. Vince / Bioorg. Med. Chem. 14 (2006) 5742–5755
tal. Yield: 50%; mp 210–212 °C; 1H NMR (DMSO-
d6): d 7.97 (1H, s), 7.26 (2H, dd, J = 6, 9 Hz), 7.12
(2H, t, J = 9 Hz), 6.57 (2H, s), 5.16 (2H, s), 3.32
124.4, 116.2, 116.0, 53.7, 38.9; MS (ESI): m/z 458.2
[M+H]+; HRMS (ESI): Calcd for (C24H21N7O2F)+
458.1735, found 458.1741 MH+.
(6H, s); 13C NMR (DMSO-d6):
d 151.8, 151.2,
150.9, 148.9, 133.9, 130.0 (2), 118.1, 116.0 (2), 43.5,
38.5; MS (ESI): m/z 287.07 [M+H]+, 309.05
[M+Na]+. The reduced starting material (same as 35)
was also isolated, yield 50%.
References and notes
1. UNAIDS; WHO. AIDS epidemic update, December 2005.
Special Report on HIV Prevention. Geneva, Switzerland:
Joint United Nations Programme on HIV/AIDS and
World Health Organization, 2005.
3.27. N,N-Dimethyl 8-picolinamide-9-(4-fluorobenzyl)
adenine (4a)
2. Richman, D. D. Nature 2001, 410, 995–1001.
3. Mocroft, A.; Lundgren, J. D. J. Antimicrob. Chemother.
2004, 54, 10–13.
4. Shehu-Xhilaga, M.; Tachedjian, G.; Crowe, S. M.;
Kedzierska, K. Curr. Med. Chem. 2005, 12, 1705–
1719.
5. Pommier, Y.; Neamati, N. Adv. Virus Res. 1999, 52,
427–458.
6. Asante-Appiah, E.; Skalka, A. M. Adv. Virus Res. 1999,
52, 351–369.
7. Pommier, Y.; Johnson, A. A.; Marchand, C. Nat. Rev.
Drug Disc. 2005, 4, 236–248.
8. Hazuda, D. J.; Felock, P.; Witmer, M.; Wolfe, A.;
Stillmock, K.; Grobler, J. A.; Espeseth, A.; Gabryelski,
L.; Schleif, W.; Blau, C.; Miller, M. D. Science 2000, 287,
646–650.
9. Billich, A. Curr. Opin. Investig. Drugs 2003, 4, 206–209.
10. Grobler, J. A.; Stillmock, K.; Hu, B.; Witmer, M.;
Felock, P.; Espeseth, A. S.; Wolfe, A.; Egbertson, M.;
Bourgeois, M.; Melamed, J.; Wai, J. S.; Young, S.;
Vacca, J.; Hazuda, D. J. Proc. Natl. Acad. Sci. U.S.A.
2002, 99, 6661–6666.
11. Wai, J. S.; Egbertson, M. S.; Payne, L. S.; Fisher, T. E.;
Embrey, M. W.; Tran, L. O.; Melamed, J. Y.; Langford,
H. M.; Guare, J. P., Jr.; Zhuang, L.; Grey, V. E.; Vacca, J.
P.; Holloway, M. K.; Naylor-Olsen, A. M.; Hazuda, D. J.;
Felock, P. J.; Wolfe, A. L.; Stillmock, K. A.; Schleif, W.
A.; Gabryelski, L. J.; Young, S. D. J. Med. Chem. 2000,
43, 4923–4926.
12. Pais, G. C. G.; Zhang, X.; Marchand, C.; Neamati, N.;
Cowansage, K.; Svarovskaia, E. S.; Pathak, V. K.; Tang,
Y.; Nicklaus, M.; Pommier, Y.; Burke, T. R., Jr. J. Med.
Chem. 2002, 45, 3184–3194.
13. Sechi, M.; Derudas, M.; Dallocchio, R.; Dessi, A.; Bacchi,
A.; Sannia, L.; Carta, F.; Palomba, M.; Ragab, O.; Chan,
C.; Shoemaker, R.; Sei, S.; Dayam, R.; Neamati, N.
J. Med. Chem. 2004, 47, 5298–5310.
14. Fujishita, T.; Yoshinaga, T. WO Patent 9950245, 1999.
15. Kawasuji, T.; Yoshinaga, T. WO Patent 2001017968,
2001.
16. Selnick, H. G.; Hazuda, D. J.; Egbertson, M.; Guare, J. P.,
Jr.; Wai, J. S.; Young, S. D.; Clark, D. L.; Medina, J. C.
WO Patent 9962513, 1999.
17. Young, S. D.; Egbertson, M.; Payne, L. S.; Wai, J. S.;
Fisher, T. E.; Guare, J. P., Jr.; Embrey, M. W.; Tran, L.;
Zhuang, L.; Vacca, J. P.; Langford, M.; Melamed, J.;
Clark, D. L.; Medina, J. C.; Jaen, J. WO Patent 9962520,
1999.
18. Fujishita, T.; Yoshinaga, T.; Sato, A. WO Patent 0039086,
2000.
19. Nair, V.; Chi, G.; Ptak, R.; Neamati, N. J. Med. Chem.
2006, 49, 445–447.
Compound 4a was prepared from compound 38 (30 mg,
0.1 mmol) according to method G. Yield: 54%; mp 172–
173 °C; H NMR (CDCl3): d 10.04 (1H, s), 8.59 (1H, d,
J = 4.2 Hz), 8.38 (1H, s), 8.28 (1H, d, J = 7.8 Hz), 7.94
(1H, t, J = 7.8 Hz), 7.54 (1H, dd, J = 4.8, 6.6 Hz), 7.18
(2H, m), 6.91 (2H, t, J = 8.4 Hz), 5.45 (2H, s), 3.58 (6H,
br s); 13C NMR (CDCl3): d 163.2, 161.6, 152.3, 150.8,
148.4, 148.1, 137.8, 131.7, 129.4 (2), 127.3, 122.8, 115.8,
115.6, 45.5, 38.5; MS (ESI): m/z 392.0 [M+H]+, 414.0
[M+Na]+; HRMS (ESI): Calcd for (C20H18FN7NaO)+
414.1449, found: 414.1471 [M+Na]+.
1
3.28. N,N-Dimethyl 9-(4-fluorobenzyl) 8-(8-benzyloxy-
quinoline-7-carboxamide) adenine (39)
A solution of N,N-dimethyl 8-amino 9-(4-fluorobenzyl)
adenine (38, 30 mg, 0.105 mmol) in 1 mL DMF was
treated with sodium hydride (5 mg, 0.125 mmol) in a
5 mL flask at room temperature for 20 min. A solution
of 8-benzyloxy-quinoline-7-carboxylic acid benzyl ester
(18, 60 mg, 0.162 mmol) in 1 mL DMF was added to
the flask and continued stirring at room temperature
overnight. The excess DMF was removed in vacuo
and the crude product was purified by flash chromatog-
raphy (CHCl3/MeOH 100:1) to give the title compound
1
as white solid. Yield: 53%; mp 216–220 °C; H NMR
(CDCl3): d 10.42 (1H, br s), 9.07 (1H, d, J = 3.6 Hz),
8.37 (1H, s), 8.29 (1H, d, J = 9 Hz), 8.24 (1H, d,
J = 8.4 Hz), 7.70 (1H, d, J = 8.4 Hz), 7.58 (1H, dd,
J = 4.8, 8.4 Hz), 7.55 (2H, d, J = 4.2 Hz), 7.36 (3H, m),
7.045 (2H, m), 6.81 (2H, t, J = 8.4 Hz), 5.63 (2H, s),
5.29 (2H, s), 3.53 (6H, br s); 13C NMR (CDCl3): d
164.2, 161.4, 154.5, 152.2, 150.9, 150.2, 139.9, 136.3,
135.6, 132.3, 129.4, 129.2, 129.1, 129.0, 128.7, 127.3,
123.8, 123.2, 115.6, 115.5, 78.8, 45.3, 38.4; MS (ESI):
m/z 548.2 [M+H]+, 570.1 [M+Na]+.
3.29. N,N-Dimethyl 9-(4-fluorobenzyl) 8-(8-hydroxy-
quinoline-7-carboxamide) adenine (4b)
A solution of compound 39 (25 mg, 0.045 mmol) in
5 mL DCM was treated with 1 mL TFA. The mixture
was stirred at room temperature for 3 days. After con-
centration, the semi-solid was dissolved in 1 mL EtOAc
and crystallized to give the title compound as bright yel-
low crystal. Yield: 50%; mp 265–267 °C; 1H NMR
(DMSO-d6): d 8.91 (1H, d, J = 4.2 Hz), 8.61 (1H, d,
J = 8.1 Hz), 8.28 (1H, s), 8.02 (1H, d, J = 9 Hz), 7.80
(1H, dd, J = 8.1, 4.8 Hz), 7.31 (3H, m), 7.09 (2H, t,
J = 9 Hz), 5.38 (2H, s), 4.01 (6H, br s); 13C NMR
(DMSO-d6): d 161.4, 158.7, 133.0, 132.4, 130.3, 128.6,
20. After we submitted our manuscript, the following refer-
ence came to our attention Nair, V.; Uchil, V.; Neamati,
N. Bioorg. Med. Chem. Lett. 2006, 16, 1920–1923.
21. Baaden, M.; Berny, F.; Madic, C.; Schurhammer, R.;
Wipff, G. Solvent Extr. Ion Exch. 2003, 21, 199–220.

X. Li, R. Vince / Bioorg. Med. Chem. 14 (2006) 5742–5755
5755
22. Conley, H. L., Jr.; Martin, R. B. J. Phys. Chem. 1965, 69,
2914–2923.
28. Klapars, A.; Huang, X.; Buchwald, S. L. J. Am. Chem.
Soc. 2002, 124, 7421–7428.
23. Zhuang, L.; Wai, J. S.; Embrey, M. W.; Fisher, T. E.;
Egbertson, M. S.; Payne, L. S.; Guare, J. P., Jr.; Vacca, J.
P.; Hazuda, D. J.; Felock, P. J.; Wolfe, A. L.; Stillmock,
K. A.; Witmer, M. V.; Moyer, G.; Schleif, W. A.;
Gabryelski, L. J.; Leonard, Y. M.; Lynch, J. J., Jr.;
Michelson, S. R.; Young, S. D. J. Med. Chem. 2003, 46,
453–456.
29. Huang, X.; Anderson, K. W.; Zim, D.; Jiang, L.; Klapars,
A.;Buchwald, S. L. J. Am. Chem.Soc. 2003, 125, 6653–6655.
30. Eckstein, Z.; Kanik, B.; Leszczynska, E.; Lipczynska-
Kochany, E.; Pajak, J.; Zawadzka, M. Pol. J. Chem. 1979,
53, 2373–2377.
31. Holmes, R. E.; Robins, R. K. J. Am. Chem. Soc. 1965, 87,
1772–1776.
24. Lakshman, M. K. Curr. Org. Synth. 2005, 2, 83–112.
25. Gunda, P.; Russon, L. M.; Lakshman, M. K. Angew.
Chem., Int. Ed. 2004, 43, 6372–6377.
32. Laufer, S. A.; Domeyer, D. M.; Scior, T. R. F.;
Albrecht, W.; Hauser, D. R. J. J. Med. Chem. 2005,
48, 710–722.
26. Terrazas, M.; Ariza, X.; Farras, J.; Guisado-Yang, J. M.;
Vilarrasa, J. J. Org. Chem. 2004, 69, 5473–5475.
27. Meszarosova, K.; Holy, A.; Masojidkova, M. Collect.
Czech. Chem. Commun. 2000, 65, 1109–1125.
33. John, S.; Fletcher, T. M., III; Jonsson, C. B. J. Biomol.
Screen. 2005, 10, 606–614.
34. Wang, T.; John, S.; Archuleta, S.; Jonsson, C. B. Protein
Expr. Purif. 2004, 33, 232–237.