Neamine-heterocycle conjugates as potential anti-HIV agents

Article abstract of DOI:10.1016/j.cclet.2013.03.009

A series of neamine-heterocycle conjugates were designed and synthesized. All new compounds displayed more potent inhibitory effect on HIV replication than neamine, among them two compounds displayed stronger anti-HIV activity than neomycin B. The results

Full text of DOI:10.1016/j.cclet.2013.03.009

Chinese Chemical Letters 24 (2013) 273–278
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Chinese Chemical Letters
journal homepage: www.elsevier.com/locate/cclet
Original article
Neamine-heterocycle conjugates as potential anti-HIV agents
a,
Ri-Bai Yan a, Yan-Fen Wu a, Jun Liu b, Xiao-Lian Zhang b, , Xin-Shan Ye
*
*
a State Key Laboratory of Natural and Biomimetic Drugs, School of Pharmaceutical Sciences, Peking University, Beijing 100191, China
b State Key Laboratory of Virology, Department of Immunology, Hubei Province Key Laboratory of Allergy and Immunology, Wuhan University School of Medicine,
Wuhan 430071, China
A R T I C L E I N F O
A B S T R A C T
Article history:
A series of neamine-heterocycle conjugates were designed and synthesized. All new compounds
displayed more potent inhibitory effect on HIV replication than neamine, among them two compounds
displayed stronger anti-HIV activity than neomycin B. The results suggested that it might be a promising
approach to modify neamine for the discovery of new anti-HIV agents.
Received 9 January 2013
Received in revised form 16 February 2013
Accepted 21 February 2013
ß 2013 Xiao-Lian Zhang, Xin-Shan Ye. Published by Elsevier B.V. on behalf of Chinese Chemical
Society. All rights reserved.
Keywords:
Aminoglycoside
Anti-HIV
Neamine
Synthesis
1. Introduction
whether these modifications benefit the actual bioactivity remains
unknown. Herein we report the design and synthesis of some new
Aminoglycosides, usually binding to a variety of RNAs, are a
large family of clinically important antibiotics effective against a
broad range of bacteria [1]. They are multiply charged compounds
of high flexibility. The positive charges are attracted to the
negatively charged RNA backbone. The flexibility of aminoglyco-
sides facilitates accommodation into a binding pocket for making
specific contacts. Among them, neomycin B (Fig. 1) has been
proved to display anti-HIV activity [2]. Further investigations
suggested that neamine, a component of neomycin B, is the key
moiety in the interaction with HIV target RNAs [3]. Neamine is a
hydrophilic pseudo-disaccharide possessing several amino func-
tionalities that are mostly protonated under physiological condi-
tions and have good cellular uptake ability [4]. By means of
electrostatic and H-bond effects, it can bind to RNAs in a sequence-
specific fashion [5]. So far many neamine derivatives have been
prepared, and their affinities to target RNAs were evaluated by
fluorescence titration, surface plasmid resonance (SPR), or other
methods. Considerable new compounds exhibited stronger affini-
ties to target RNAs of HIV than neamine [6]. However, previous
research on antibacterial evaluation revealed that there is no good
correlation between affinity and bioactivity [7], and corresponding
data on anti-HIV potency have been less reported, therefore
neamine-heterocycle conjugates and their bioassay on anti-HIV. In
order to avoid the specious results in bioactivity assessment, we
observed the potency of these compounds to inhibit the replication
of HIV on cell level.
2. Experimental
Non-aqueous reactions were performed under nitrogen atmo-
sphere at room temperature, unless otherwise noted. All commer-
cial reagents were purchased from Aldrich or Acros and were used
without further purification. Anhydrous pyridine and dichloro-
methane were obtained by distilling commercial ones over CaH2
and anhydrous DMF by distilling over P2O5 under reduced
pressure. Routine 1H and 13C nuclear magnetic resonance spectra
were recorded. Samples were dissolved in deuterated chloroform
(CDCl3) or deuterium oxide (D2O) and tetramethylsilane (TMS) was
used as reference. Elemental analyses were performed for key
compounds. Elemental analyses were performed in PE-2400C
analyzer. Mass spectra were recorded on PE SCLEX QSTAR mass
spectrometer (for low resolution ESI-MS) or Bruker APEX IV mass
spectrometer (for high resolution ESI-MS). Analytical thin-layer
chromatography (TLC) was performed on Merck silica gel 60 F254
.
Compounds were visualized by UV light (254 nm) and/or by
staining with a yellow solution containing Ce(NH4)2(NO3)6 (0.5 g)
and (NH4)6Mo7O24ꢀ4H2O (24.0 g) in 6% H2SO4 (500 mL) (for
intermediates) or ninhydrin solution in ethyl acetate (5%) followed
by heating (for final compounds).
* Corresponding authors.
E-mail addresses: zhangxiaolian@whu.edu.cn (X.-L. Zhang),
xinshan@bjmu.edu.cn (X.-S. Ye).
1001-8417/$ – see front matter ß 2013 Xiao-Lian Zhang, Xin-Shan Ye. Published by Elsevier B.V. on behalf of Chinese Chemical Society. All rights reserved.
http://dx.doi.org/10.1016/j.cclet.2013.03.009

[
R.-B. Yan et al. / Chinese Chemical Letters 24 (2013) 273–278
NH2
O
HO
HO
NH2
O
HO
HO
H2N
NH2
O
H2N
HO
NH2
NH2
O
NH2
OH
O
O
HO
OH
NH2
OH
O
OH
O
OH
H2N
Neamine, 2
Fig. 1. The structures of neomycin B and neamine.
Neomycin B, 1
2.1. epi-5-(2-Aminoethyl)-amino-1,3,20,60-tetraazido-6,30,40-tri-O-
43.64, H 4.65, N 29.36; found: C 43.56, H 4.87, N 29.19. HRESI-MS
calcd. for C26H33N15O10 [M+H]+: 716.2608; found: 716.2617.
acetyl neamine (14)
To a stirring mixture of 13 (300 mg, 0.54 mmol), pyridine
(0.2 mL, 2.48 mmol) and dichloromethane (10 mL) was added
triflic anhydride (0.18 mL, 1.08 mmol) solution in dichloro-
methane (1 mL) dropwise by a syringe in a period of 10 min at
room temperature. After stirring for 3.5 h, TLC assay showed that
compound 13 was consumed. After removal of the solvent of the
reaction mixture, the residue was dissolved in 20 mL of ethyl
acetate and the resulting solution was washed with brine (2ꢁ
60 mL). The organic layer was collected, dried over anhydrous
sodium sulfate, and concentrated to foam in vacuo. Then it was
dissolved in 5 mL of acetonitrile and to the solution was added
ethylene diamine (1 mL) in one portion while stirring. Half an hour
later, this reaction mixture was poured into cold water (8 mL) and
extracted with dichloromethane (3ꢁ 30 mL), and the combined
organic layers were dried over anhydrous sodium sulfate, filtered
and concentrated in vacuum. The residue was purified by column
chromatography on silica gel (chloroform/methanol = 30/1) to
provide a white foam (200 mg, 63% yield). 1H NMR (400 MHz,
2.3. epi-5-[2-(6-Hydroxynicotinyl)aminoethyl)-amino neamine (5)
To a solution of 15a (78 mg, 0.11 mmol) in methanol (10 mL)
was added sodium methoxide solution in methanol (30%, 0.1 mL)
while stirring. When TLC showed the reaction was completed, the
solvent was removed. After the residue was purified by a short
column chromatography on silica gel (eluent: chloroform/metha-
nol = 10/1), the resulting product was dissolved in a mixture of
pyridine (3 mL), triethylamine (2 mL), and water (1 mL). Then the
mixture was bubbled with H2S until it turned deep green. After
stirring overnight, the solvent was removed in vacuum. The
residue was suspended in 1.5 mL of methanol and the resulting
mixture was absorbed in a proper quantity of silica gel. After
removal of the volatile component, the mixture was transferred
onto a short silica gel column. The column was washed with
eluents as following: petroleum/ethyl acetate (25 mL/25 mL),
ethyl acetate (50 mL), ethyl acetate/methanol (25 mL/25 mL),
methanol (50 mL), and methanol/concentrated aqua ammonia
(100 mL/10 mL). Finally the desired product was obtained as a
white powder (42 mg, 0.09 mmol) in 79% yield over two steps.
CDCl3):
d 5.48 (t, 1H, J = 10.0 Hz), 5.10–5.03 (m, 2H), 4.70 (dd, 1H,
J1 = 2.5 Hz, J2 = 10.0 Hz), 4.32–4.19 (m, 2H), 4.03–3.94 (m, 1H),
3.70–3.61 (m, 2H), 3.47–3.41 (m, 2H), 3.30 (dd, 1H, J1 = 5.0 Hz,
J2 = 13.5 Hz), 2.98–2.83 (m, 3H), 2.67–2.64 (m, 1H), 2.36 (ddd, 1H,
J1 = 13.5 Hz, J2 = J3 = 5.0 Hz), 2.15 (s, 3H), 2.09 (s, 3H), 2.06 (s, 3H),
2.4. epi-5-Amino-1,3,20,60-tetraazido-6,30,40-tri-O-benzyl neamine
(17)
1.35 (ddd, 1H, J1 = J2 = J3 = 13.5 Hz); 13C NMR (75 MHz, CDCl3):
d
169.9, 169.7, 94.3, 77.9, 76.0, 70.9, 69.5, 68.9, 61.3, 57.4, 56.1, 50.6,
50.5, 41.6, 32.2, 20.9, 20.6. HRESI-MS calcd. for C20H30N14O8
[M+H]+: 595.2443; found: 595.2431.
Compound 16 (300 mg, 0.43 mmol), pyridine (1.2 mL), and
dichloromethane (15 mL) were mixed in one flask. To the solution
was added triflic anhydride (1.2 mL, 7.15 mmol) solution in
dichloromethane (10 mL) dropwise in a period of 2 h while
stirring at room temperature. After completion of the addition of
triflic anhydride, the reaction was stirred for another 4 h. Then the
reaction mixture was poured into the mixture of ice-water (40 mL)
and was stirred for 20 min. The organic layer was separated out,
dried over anhydrous sodium sulfate, and then concentrated to
residue. After running column chromatography on silica gel with
petroleum/ethyl acetate (10/1) as eluent, the crude product was
dissolved in acetone (15 mL). Then the solution was cooled down
with an ice bath and bubbled with ammonia for 1 h. After stirring
for another 1 h, TLC assay showed that all of the triflate was
consumed. Then the reaction mixture was concentrated and the
resulting residue was subjected to column chromatography on
silica gel with petroleum/ethyl acetate (8/1) as eluent. Finally the
desired product was obtained (155 mg, 52% yield from 16) as a
2.2. epi-5-[2-(6-Hydroxynicotinyl)aminoethyl]-amino-1,3,20,60-
tetraazido-6,30,40-tri-O-acetyl neamine (15a)
Compound 14 (100 mg, 0.17 mmol), 6-hydroxyl-nicotinic acid
(35 mg, 0.26 mmol), HBTU (115 mg, 0.39 mmol), DIPEA (68 mg,
0.5 mmol) and DMF (15 mL) were mixed in one portion. After
stirring for 12 h, TLC assay showed the reaction was completed.
The solvent was removed in vacuo, and the residue was purified by
column chromatography on silica gel with chloroform/methanol
(30/1) as eluent to afford the desired product (92 mg, 76% yield) as
a white powder. 1H NMR (300 MHz, CDCl3):
d 8.53 (d, 1H,
J = 2.1 Hz), 7.90 (dd, 1H, J1 = 2.4 Hz, J2 = 8.4 Hz), 6.52 (d, 1H,
J = 8.4 Hz), 6.44 (br, 1H), 5.48 (t, 1H, J = 9.9 Hz), 5.09–5.03 (m, 2H),
4.77 (s, 2H), 4.72 (dd, 1H, J1 = 2.5 Hz, J2 = 10.1 Hz), 4.32–4.26 (m,
1H), 4.12–4.03 (m, 1H), 3.93–3.84 (m, 1H), 3.70–3.61 (m, 3H),
3.48–3.37 (m, 3H), 3.30 (dd, 1H, J1 = 5.1 Hz, J2 = 13.5 Hz), 2.98–2.91
(m, 2H), 2.37 (ddd, 1H, J1 = 13.2 Hz, J2 = J3 = 5.0 Hz), 2.13 (s, 3H),
2.09 (s, 3H), 2.05 (s, 3H), 1.39 (ddd, 1H, J1 = J2 = J3 = 12.5 Hz). 13C-
white semisolid. 1H NMR (500 MHz, CDCl3):
d 7.39–7.25 (m, 15H),
4.95 (d, 1H, J = 4.0 Hz), 4.90–4.88 (m, 3H), 4.70–4.61 (m, 3H), 4.12–
4.02 (m, 4H), 3.80 (t, 1H, J = 3.0 Hz), 3.63–3.50 (m, 3H), 3.41 (dd, 1H,
J1 = 3.5 Hz, J2 = 10.0 Hz), 3.36 (dd, 1H, J1 = 4.5 Hz, J2 = 13.5 Hz), 3.27
(dd, 1H, J1 = 3.5 Hz, J2 = 9.5 Hz), 2.23 (ddd, 1H, J1 = 13.5 Hz,
J2 = J3 = 5.0 Hz), 1.22 (ddd, 1H, J1 = J2 = J3 = 12.5 Hz). 13C NMR
NMR (75 MHz, CDCl3):
d 170.0, 169.9, 169.7, 160.3, 147.5, 137.0,
120.2, 107.9, 94.4, 77.8, 75.8, 70.9, 69.6, 69.0, 61.3, 57.4, 56.2, 55.9,
50.6, 49.5, 40.4, 32.0, 20.9, 20.6. Anal. calcd. for C26H33N15O10: C
(125 MHz, CDCl3):
d 137.6, 137.5, 137.3, 128.6, 128.5 (2 C),

R.-B. Yan et al. / Chinese Chemical Letters 24 (2013) 273–278
275
128.1, 128.0, 127.9, 127.7, 93.9, 81.5, 80.4, 78.6, 78.1, 75.7, 75.0,
72.2, 71.3, 63.5, 57.6, 57.3, 50.9, 47.9, 32.6. Anal. calcd. for
dissolved in 5 mL of methanol and pH of the resulting solution was
adjusted to 3–4 with hydrochloric acid (1 mol/L). Then catalytic
amount of Pd/C was added. The mixture was stirred for 5 days
under H2 atmosphere. After TLC showed that the reaction was
completed, the solvent was evaporated to give the product as
white amorphous powder in 67% yield.
C
33H37N13O5: C 56.97, H 5.36, N 26.17; found: C 57.20, H 5.13, N
25.89. ESI-MS calcd. for C33H37N13O5 [M+H]+: 696; found: 696.
2.5. epi-5-[(2-Amino-6-benzyloxy-purine-9-yl) acetyl]-amino-
1,3,20,60-tetraazido-6,30,40-tri-O-benzyl neamine (18a)
2.8. epi-5-(2-Aminoethyl)-amino neamine (3)
Compound 17 (52 mg, 0.075 mmol), (2-amino-6-(benzyloxy)-
purine-9-yl) acetic acid (27 mg, 0.090 mmol), TBTU (35 mg,
0.108 mmol), triethylamine (45 mg, 0.45 mmol) and DMF (8 mL)
were mixed in one flask. After stirring for 12 h, another portion of
A mixture of 19a (21 mg, 0.028 mmol), triphenylphosphine
(60 mg, 0.228 mmol), THF (10 mL) and water (0.5 mL) was heated
under reflux for 24 h. Then the solvent was removed and the
residue was subjected to a short column chromatography on silica
gel with eluents as following: EtOAc (40 mL), EtOAc/MeOH (1/1,
60 mL), MeOH/NH3(aq) (40/1, 80 mL). The fraction containing
amine was collected and concentrated to residue. The residue was
dissolved in the mixture of THF (5.6 mL), acetic acid (1.4 mL) and
water (1.4 mL). Then catalytic amount of Pd/C was added. The
mixture was stirred under H2 atmosphere for 72 h. The mixture
was filtered, and the filtrate was concentrated to afford the desired
product in the form of acetate as yellowish amorphous powder
(18 mg, 95% yield).
(2-amino-6-(benzyloxy)-purine-9-yl)
acetic
acid
(14 mg,
0.045 mmol) and TBTU (18 mg, 0.054 mmol) was added to the
reaction mixture. The reaction was stirred for another 24 h. After
removal of the solvent followed by purification with gradient
column chromatography on silica gel (eluent: petroleum/ethyl
acetate 2/1 to 1.5/1), the desired product was obtained as white
solid (36 mg, 49% yield). 1H NMR (500 MHz, CDCl3):
d 7.60 (s, 1H),
7.55 (d, 1H, J = 5.0 Hz), 7.46 (d, 2H, J = 6.0 Hz), 7.36–7.22 (m, 20H),
5.59 (d, 1H, J = 12.5 Hz), 5.55 (d, 1H, J = 12.0 Hz), 5.24 (d, 1H,
J = 4.0 Hz), 5.10 (m, 1H), 4.90 (s, 2H), 4.86 (d, 1H, J = 11.0 Hz), 4.80–
4.69 (m, 4H), 4.59 (t, 1H, J = 11.5 Hz), 4.41 (d, 1H, J = 11.0 Hz), 3.93
(dd, 1H, J1 = 9.0 Hz, J2 = 10.0 Hz), 3.88 (m, 1H), 3.58–3.34 (m, 7H),
3.29 (dd, 1H, J1 = 5.5 Hz, J2 = 13.0 Hz), 2.14 (ddd, 1H, J1 = 13.5 Hz,
J2 = J3 = 4.5 Hz), 1.23 (ddd, 1H, J1 = J2 = J3 = 13.0 Hz). 13C NMR
3. Results and discussion
It is known that neamine is a weak antibiotic and can bind to
RRE RNA with a low affinity [6c], we wonder whether the
introduction of an additional heterocyclic moiety can enhance its
activity. So a series of new neamine derivatives, which are
characterized by the neamine moiety conjugated with a heterocy-
cle, typically a nucleobase or its mimics, by a linker, were designed
(Fig. 2). Except for the inherent electrostatic and H-bond effects of
neamine, we anticipated the additional base or its mimics can
exert the base–base-like interaction with target RNA, just like it is
observed in classic nucleic acids, thus enhancing the anti-HIV
activities.
(125 MHz, CDCl3):
d 167.5, 161.5, 159.2, 153.7, 139.8, 137.5,
136.6, 136.0, 128.5 (2C), 128.4, 128.3, 128.1, 128.0, 127.9, 127.8,
115.6, 93.6, 79.9, 78.7, 78.4, 75.5, 75.0, 74.6, 71.7, 71.4, 68.7, 63.0,
58.3, 57.9, 51.0, 47.8, 44.9, 31.9. Anal. calcd. for C47H48N18O7: C
57.78, H 4.95, N 25.81; found: C 57.83, H 5.17, N 25.63. ESI-MS
calcd. for C47H48N18O7 [M+H]+: 977; found: 977.
2.6. epi-5-(2-Aminoethyl)-amino-1,3,20,60-tetraazido-6,30,40-tri-O-
benzyl neamine (19a)
The synthesis of 5-O-triflyl-1,3,20,60-tetraazido- 6,30,40-tri-O-
benzyl neamine was the same as described in synthesis of 17. To
the triflate (600 mg, 0.72 mmol) solution in acetonitrile (20 mL)
was added ethylene diamine (1 mL) while stirring. When TLC
showed that all of the triflate was consumed, the solvent was
removed and the residue was purified by column chromatography
on silica gel. The final product was obtained as a yellow gel
To introduce heterocycles onto neamine, two synthetic routes
were employed. The first route used perazidotriacetyl neamine
derivative 14 as the key intermediate, as depicted in Scheme 1.
Perazidotriacetyl neamine (13) [8] was prepared from neamine by
an improved diazotransfer procedure [9]. Treatment of compound
13 with triflic anhydride in the presence of pyridine in
dichloromethane gave its triflate, which reacted with ethylene
diamine to yield 14. Then heterocyclic carboxylic acids were
coupled with 14 to form the corresponding amides 15a–d.
Deacetylation of compounds 15a–d which was followed by
reduction of azido groups with H2S provided the target compounds
5–8 [10].
For the synthesis of neamine-heterocycle conjugates 9–12, the
second approach was used, as outlined in Scheme 2. The
benzylated neamine derivative 17 served as the intermediate.
Firstly, 30,40,6-tribenzylperazido neamine (16) was prepared
according to the literature procedure [6b]. The triflation of 5-
hydroxyl in 16 which was followed by nucleophilic substitution
with ammonia afforded intermediate 17, which was coupled with
various heterocyclic carboxylic acids [8,11], yielding the neamine-
heterocycle conjugates 18a–d. The target compounds 9–12 [12]
were accomplished successfully by reduction of azido groups with
H2S which was followed by removal of benzyl groups with
palladium-catalyzed hydrogenolysis. By a similar way, two simple
neamine derivatives, epi-5-(2-aminoethyl)-amino neamine (3)
and epi-5-(2-hydroxyethyl)-amino neamine (4) were also
obtained [13].
(471 mg, 67% overall yield from 16). 1H NMR (300 MHz, CDCl3):
d
7.38–7.26 (m, 15H), 5.28 (d, 1H, J = 12.0 Hz), 5.02 (d, 1H, J = 3.6 Hz),
4.89 (d, 1H, J = 13.2 Hz), 4.86 (s, 2H), 4.71–4.60 (m, 3H), 4.11–3.98
(m, 4H), 3.59–3.45 (m, 5H), 3.34 (dd, 1H, J1 = 4.5 Hz, J2 = 13.5 Hz),
3.26 (dd, 1H, J1 = 2.4 Hz, J2 = 9.9 Hz), 2.96–2.89 (m, 1H), 2.82–2.72
(m, 3H), 2.25 (ddd, 1H, J1 = J2 = 5.1 Hz, J3 = 13.2 Hz), 1.19 (ddd, 1H,
J1 = J2 = J3 = 13.2 Hz). 13C NMR (75 MHz, CDCl3):
d 137.6, 137.5,
137.3, 128.5, 128.1, 128.0, 127.9, 127.7, 93.8, 82.8, 80.2, 78.5, 77.7,
77.2, 75.6, 75.0, 72.4, 71.3, 63.5, 58.0, 54.2, 52.3, 50.9, 42.3, 32.9.
HRESI-MS calcd. for
739.3543.
C
35H42N14O5 [M+H]+: 739.3535; found:
2.7. epi-5-[(Guanine-9-yl)acetyl]-amino neamine (9)
Compound 18a (100 mg) was dissolved in the mixture of
pyridine (3 mL), triethylamine (2 mL) and water (0.5 mL). Then,
H2S gas was passed into the mixture until it turned deep green. The
reaction mixture was stirred for 10 h at room temperature. The
mixture was concentrated and passed through a short silica
column with eluents as following: petroleum/ethyl acetate
(25 mL/25 mL), ethyl acetate (50 mL), ethyl acetate/methanol
(25 mL/25 mL), methanol (50 mL), and methanol/concentrated
aqua ammonia (100 mL/10 mL). The crude product was then
In order to test the effects of these compounds on the HIV
replication, different compounds were added to each HIV pseudo-
type virus (HIVpp) infected cells, respectively. HIV pseudo-type

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R.-B. Yan et al. / Chinese Chemical Letters 24 (2013) 273–278
R =
4
6
3
5
NH2
O
OH
NH2
O
N
N
O
N
HO
HO
OH
OCH3
OH
H2N
NH
O
NH
O
NH2
O
NH2
N
7
9
8
N
NH2
OH
NH
NHR
NH
OH
NH2
NH
O
O
N
10
N
N
N
N
O
O
NH2
NH2
O
O
N
N
N
O
N
12
11
NH
NH2
N
Fig. 2. Structures of the designed neamine derivatives.
virus was produced by co-transfection of HEK 293T cells with a
plasmid phRL-null vector (Promega) and a pNL4-3.luc.RꢂEꢂ (NIH)
containing the env deficient HIV proviral genome and an intact
luciferase gene. The phRL-null vector [hRluc/SV40] (Promega) was
used as the internal control Renilla luciferase reporter. The
replication of the HIVpp in 293T cells was measured through
normalizing the transfection efficacy with Luciferase and Renilla
luciferase activities. The relative luciferase activities represented
the viral replication efficacy of HIVpp. For better understanding the
potency of these compounds on anti-HIV, AZT, a well-known anti-
HIV drug, was used as a positive control during evaluation.
As shown in Fig. 3, all of the tested compounds displayed
inhibitory effects on the HIV replication when comparing with the
control group without drugs. Joyfully, all new compounds
exhibited stronger potency than neamine, the parent compound.
Compounds 9–12 with heterocycle-acetyl moieties showed
relatively weak inhibitory activities. To our surprise, compounds
3 and 4, two simply-modified derivatives of neamine, showed a
remarkable increase in anti-HIV potency, even beyond some
compounds containing heterocycle moiety. Compounds 5–7
contain the nicotine moiety substituted by the hydroxyl, methoxy,
and amino groups, respectively. Among these three compounds,
compound 6 with the methoxy substituent exhibited the strongest
potency. Compound 8 with the aminoethyl linkage also showed
potent anti-HIV activity. At the same concentration, the suppres-
sion effects of 6 and 8 exceeded that of neomycin B. In fact,
compound 6 showed the best inhibitory effect on anti-HIV among
all compounds with nucleobase-like heterocycles. Generally, our
experiments demonstrated that for neamine-heterocycle conju-
gates, compounds with the aminoethyl linkage showed better anti-
HIV activities than the compounds with heterocycle-acetyl
moieties.
[(Scheme_1)TD$FIG]
N3
N3
NH2
N3
O
O
O
O
AcO
AcO
AcO
AcO
HO
HO
AcO
AcO
N3
N3
b
H2N
a
N
c
3 O
N
NH2
3 O
N3
N3
O
R
N3
R
N3
O
NH2
OAc
OAc
HO
N3
OH
O
O
HN
OAc
H2N
NH
NH
HN
NH
13
14
15a, 15b, 15c, 15d
5, 6, 7, 8
R
R
15a
15b
HO
N
HO
5
6
N
H3CO
Cl
N
N
15c
15d
7
8
H2N
HO
H2N
N
N
N
HO
N
N
N
HO
HO
Scheme 1. Synthesis of neamine-heterocycle conjugates 5, 6, 7, and 8. Reagents and conditions: (a) (i) Tf2O, Py, CH2Cl2; (ii) ethylene diamine and CH3CN, 63%; (b) heterocyclic
acids, HOBT/HBTU or DCC, DMF, 76% for 15a, 75% for 15b, 43% for 15c, 33% for 15d; (c) (i) NaOMe, MeOH; (ii) H2S, pyridine/Et3N/H2O, 79% for 5, 81% for 6, 85% for 7, 68% for 8.

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R.-B. Yan et al. / Chinese Chemical Letters 24 (2013) 273–278
277
NH2
O
N3
N3
O
O
HO
HO
BnO
BnO
N3
BnO
BnO
a
c
O
b
BnO
BnO
N3
N3
N3
O
N3
H2N
NH2
N3
N3
O
O
O
N3
N3
O
NH2
O
OBn
OBn
OH
HO
N3
NH2
NH
NH
OBn
R
R
16
N3
17
9, 10, 11, 12
d
18a, 18b, 18c, 18d
O
BnO
BnO
O
N
OBn
N
N3
N3
N
HN
N
O
9: R=
N3
18a: R=
N
H2N
OBn
N
R
H2N
N
NH
19a: R= NH2
19b: R= OH
H2N
N
O
10: R=
11: R=
12: R=
CbzHN
NHCbz
N
18b: R=
N
NH2
N
O
e
NH2
O
N
N
N
N
HO
HO
N
18c: R=
18d: R=
H2N
N
NH2
N
N
O
O
O
NH2
OH
R
NH
HN
HN
N
N
3: R= NH2
4: R= OH
H2N
H2N
Scheme 2. Synthesis of neamine-heterocycle conjugates 9, 10, 11, 12 and compounds 3, 4. Reagents and conditions: (a) (i) Tf2O, pyridine, CH2Cl2; (ii) NH3, acetone, 52%; (b)
heterocyclic acids, HBTU, DIPEA, DMF, 49% for 18a, 80% for 18b, 50% for 18c, 67% for 18d; (c) (i) H2S, pyridine/Et3N/H2O; (ii) H2, Pd/C, 67% for 9, 72% for 10, 77% for 11, 61% for
12; (d) (i) Tf2O, pyridine, CH2Cl2; (ii) 2-amino-ethanol or ethylene diamine, CH3CN, 67% for 19a, 63% for 19b; (e) (i) Ph3P, THF, H2O; (ii) H2, Pd/C, THF/HOAc/H2O, 95% for 3, 89%
for 4.
[(Fig._3)TD$FIG]
2012CB822100, and 2012CB720604) and the National Natural
Science Foundation of China (Nos. 81025008 and 30921001).
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[12] Compound 9: 1H NMR (500 MHz, D2O): d 8.40 (s, 1H), 5.53 (d, 1H, J = 3.0 Hz), 5.26
(d, 1H, J = 17.5 Hz), 5.18 (d, 1H, J = 17.5 Hz), 5.08 (br, 1H), 4.26 (dd, 1H, J1 = 3.0 Hz,
J2 = 11.0 Hz), 4.09 (dd, 1H, J1 = 4.0 Hz, J2 = 11.0 Hz), 4.03 (t, 1H, J = 11.0 Hz), 3.91–
3.87 (m, 1H), 3.79 (ddd, 1H, J1 = 12.0 Hz, J2 = J3 = 4.0 Hz), 3.66 (ddd, 1H,
J1 = 12.0 Hz, J2 = J3 = 4.5 Hz), 3.53–3.49 (m, 2H), 3.45 (dd, 1H, J1 = 3.5 Hz,
J2 = 11.0 Hz), 3.32 (dd, 1H, J1 = 6.0 Hz, J2 = 14.0 Hz), 2.63 (ddd, 1H, J1 = 13.0 Hz,
J2 = J3 = 4.5 Hz), 1.94 (ddd, 1H, J1 = J2 = J3 = 12.5 Hz). 13C NMR (125 MHz, D2O): d
171.3, 158.0, 155.5, 140.3, 112.8, 92.6, 72.8, 71.1, 70.1, 69.1, 69.0, 53.7, 50.1, 49.3,
47.9, 46.8, 40.6, 29.0. HRESI-MS calcd. for C19H32N10O7 [M+H]+: 513.2528; found:
513.2521. Compound 10: 1H NMR (500 MHz, D2O): d 5.48 (d, 1H, J = 3.5 Hz), 5.12
(t, 1H, J = 4.0 Hz), 4.54 (d, 1H, J = 18.0 Hz), 4.28 (dd, 1H, J1 = 4.0 Hz, J2 = 11.0 Hz),
4.09–4.02 (m, 3H), 3.94–3.90 (m, 1H), 3.78 (ddd, 1H, J1 = 12.5 Hz, J2 = J3 = 3.0 Hz),
3.72–3.67 (m, 1H), 3.62–3.49 (m, 5H), 3.32 (dd, 1H, J1 = 2.0 Hz, J2 = 14.0 Hz), 2.87–
2.76 (m, 2H), 2.60 (ddd, 1H, J1 = 13.0 Hz, J2 = J3 = 4.5 Hz), 1.95 (ddd, 1H,
J1 = J2 = J3 = 13.0 Hz). 13C NMR (125 MHz, D2O): d 174.9, 173.7, 155.6, 92.0,
72.0, 71.2, 69.9, 69.1, 69.0, 53.9, 51.6, 49.3, 49.1, 48.1, 44.8, 40.8, 30.8, 28.9.
HRESI-MS calcd. for C18H34N8O7 [M+H]+: 475.2623; found: 475.2622. Compound
11: 1H NMR (500 MHz, D2O): d 8.48 (s, 1H), 8.35 (s, 1H), 5.52 (d, 1H, J = 3.5 Hz),
5.42 (d, 1H, J = 17.5 Hz), 5.38 (d, 1H, J = 17.5 Hz), 5.09 (t, 1H, J = 4.0 Hz), 4.28 (dd,
1H, J1 = 3.0 Hz, J2 = 11.0 Hz), 4.11–4.03 (m, 2H), 3.92–3.88 (m, 1H), 3.81 (ddd, 1H,
J1 = 12.0 Hz, J2 = J3 = 4.5 Hz), 3.54–3.45 (m, 3H), 3.32 (dd, 1H, J1 = 7.0 Hz,
J2 = 14.0 Hz), 2.63 (ddd, 1H, J1 = 13.0 Hz, J2 = J3 = 4.5 Hz), 1.94 (ddd, 1H,
J1 = J2 = J3 = 12.5 Hz). 13C NMR (125 MHz, D2O): d 171.3, 150.7, 149.8, 146.1,
145.6, 118.8, 92.6, 72.7, 71.2, 70.1, 69.1, 68.9, 53.8, 50.1, 49.3, 48.0, 47.0, 40.7,
29.0. HRESI-MS calcd. for C19H32N10O6 [M+H]+: 497.2579; found: 497.2583.
Compound 12: 1H NMR (500 MHz, D2O): d 5.51 (d, 1H, J = 4.0 Hz), 5.50 (d, 1H,
J = 3.5 Hz), 5.06 (t, 1H, J = 3.5 Hz), 5.04 (t, 1H, J = 3.5 Hz), 4.36–4.24 (m, 4H), 4.22–
4.02 (m, 4H), 3.93–3.89 (m, 2H), 3.82–3.74 (m, 2H), 3.62–3.44 (m, 8H), 3.34–3.28
(m, 2H), 3.04–2.71 (m, 10H), 2.62–2.54 (m, 2H), 2.02–1.88 (m, 2H). 13C NMR
(125 MHz, D2O): d 174.6, 170.4, 154.2, 92.5, 92.3, 72.6, 72.4, 71.4, 70.1, 70.0, 53.8,
49.5, 49.3, 47.9, 46.2, 45.9, 40.8, 40.1, 39.8, 35.0, 34.8, 28.9. HRESI-MS calcd. for
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[10] Compound 5: 1H NMR (500 MHz, D2O, in the hydrochloride form): d 8.35 (d, 1H,
J = 2.0 Hz), 8.20 (dd, 1H, J1 = 2.0 Hz, J2 = 9.5 Hz), 7.10 (d, 1H, J = 9.5 Hz), 5.73 (d, 1H,
J = 3.5 Hz), 4.50 (dd, 1H, J1 = 3.5 Hz, J2 = 11.0 Hz), 4.38 (t, 1H, J = 4.0 Hz), 4.34 (dd,
1H, J1 = 4.0 Hz, J2 = 11.0 Hz), 4.12 (dd, 1H, J1 = 9.0 Hz, J2 = 10.0 Hz), 4.03 (m, 1H),
3.89 (ddd, 1H, J1 = J2 = 4.5 Hz, J3 = 12.0 Hz), 3.84–3.73 (m, 4H), 3.67–3.61 (m, 3H),
3.52 (dd, 1H, J1 = 3.5 Hz, J2 = 13.5 Hz), 3.41 (dd, 1H, J1 = 7.0 Hz, J2 = 13.5 Hz), 2.66
(ddd, 1H, J1 = 12.5 Hz, J2 = J3 = 4.5 Hz), 2.02 (ddd, 1H, J1 = J2 = J3 = 12.5 Hz). 13C-
NMR (125 MHz, D2O, in the hydrochloride form): d 168.0, 155.6, 142.2, 137.6,
119.1, 114.7, 92.7, 72.0, 71.3, 70.5, 68.9, 68.4, 58.9, 53.6, 51.8, 48.9, 47.7, 40.4,
38.7, 28.7. HRESI-MS calcd. for C20H35N7O7 [Mꢂe]+: 485.2593; found: 485.2598.
Compound 6: 1H NMR (300 MHz, D2O): d 8.34 (d, 1H, J = 2.1 Hz), 7.91 (dd, 1H,
J1 = 2.4 Hz, J2 = 9.0 Hz), 6.79 (d, 1H, J = 9.0 Hz), 4.91 (d, 1H, J = 3.6 Hz), 3.80 (s, 1H),
3.56–3.12 (m, 9H), 2.97–2.82 (m, 4H), 2.64 (dd, 1H, J1 = 6.6 Hz, J2 = 13.5 Hz), 2.55
(dd, 1H, J1 = 3.6 Hz, J2 = 10.5 Hz), 1.84 (ddd, 1H, J1 = 12.9 Hz, J2 = J3 = 4.5 Hz), 0.90
(ddd, 1H, J1 = J2 = J3 = 12.3 Hz). 13C NMR (75 MHz, D2O): d 171.2, 168.6, 149.1,
141.4, 126.1, 113.0, 98.0, 81.6, 78.6, 76.2, 75.0, 73.9, 60.9, 57.7, 57.1, 52.7, 50.6,
49.7, 44.1, 42.7, 38.5. HRESI-MS calcd. for C21H37N7O7 [M+H]+: 500.2827; found:
500.2829. Compound 7: 1H NMR (500 MHz, D2O, in the hydrochloride form): d
8.14 (d, 1H, J = 2.5 Hz), 7.99 (dd, 1H, J1 = 2.5 Hz, J2 = 9.5 Hz), 6.67 (d, 1H,
J = 9.5 Hz), 5.73 (d, 1H, J = 3.5 Hz), 4.51 (dd, 1H, J1 = 3.5 Hz, J2 = 11.0 Hz), 4.44
(t, 1H, J = 4.0 Hz), 4.37 (dd, 1H, J1 = 4.5 Hz, J2 = 11.0 Hz), 4.11 (dd, 1H, J1 = 8.5 Hz,
J2 = 10.0 Hz), 4.04 (m, 1H), 3.91–3.59 (m, 8H), 3.52 (dd, 1H, J1 = 3.5 Hz,
J2 = 14.0 Hz), 3.41 (dd, 1H, J1 = 7.0 Hz, J2 = 14.0 Hz), 2.67 (ddd, 1H, J1 = 13.0 Hz,
C
18H34N8O7 [M+H]+: 475.2623; found: 475.2631.
[13] Compound 3: 1H NMR (500 MHz, D2O): d 5.32 (d, 1H, J = 3.5 Hz), 3.94–3.90 (m,
1H), 3.87–3.79 (m, 3H), 3.75–3.71 (m, 1H), 3.66–3.58 (m, 3H), 3.47–3.41 (m, 3H),
3.22 (dd, 1H, J1 = 3.0 Hz, J2 = 13.5 Hz), 3.17 (dd, 1H, J1 = 3.5 Hz, J2 = 10.5 Hz), 3.09–
3.05 (m, 1H), 2.94–2.89 (m, 1H), 2.35 (ddd, 1H, J1 = 12.5 Hz, J2 = J3 = 4.5 Hz), 1.90
(s, 15H), 1.55 (ddd, 1H, J1 = J2 = J3 = 12.5 Hz). 13C NMR (75 MHz, D2O): d 184.2,
95.5, 77.8, 74.6, 73.7, 73.4, 71.5, 63.0, 59.1, 56.6, 54.6, 51.3, 49.4, 43.0, 33.3, 25.9.
HRESI-MS calcd. for C14H32N6O5 [M]+: 364.2429; found: 364.2413. Compound 4:
1H NMR (500 MHz, D2O, in the form of acetate): d 5.27 (d, 1H, J = 3.5 Hz), 3.90–
3.73 (m, 4H), 3.67–3.60 (m, 1H), 3.56 (s, 1H), 3.49–3.38 (m, 3H), 3.23–2.99 (m,
6H), 2.34 (ddd, 1H, J1 = 12.5 Hz, J2 = J3 = 4.5 Hz), 1.89 (s, 15H), 1.54 (ddd, 1H,
J1 = J2 = J3 = 12.5 Hz). 13C NMR (75 MHz, D2O, in the form of acetate): d 182.0, 93.8,
75.2, 72.1, 71.8, 71.6, 69.4, 57.2, 54.6, 49.0, 47.3, 40.8, 40.2, 30.7, 23.8. HRESI-MS
calcd. for C14H31N5O6 [M+H]+: 366.2347; found: 366.2346.