Synthesis and evaluation of (S,S)-N,N′-bis-[3-(2,2′,6,6′-tetramethylbenzhydryloxy)-2-hydroxy-propyl]-ethylenediamine (S2824) analogs with anti-tuberculosis activity

Article abstract of DOI:10.1016/j.bmcl.2009.09.035

In order to identify new and potent candidate drugs to treat tuberculosis, a library of compounds was screened, and (S,S)-N,N′-bis-[3-(2,2′,6,6′-tetramethylbenzhydryloxy)-2-hydroxy-propyl]-ethylenediamine (S2824) was identified as a hit in the screen. Thi

Full text of DOI:10.1016/j.bmcl.2009.09.035

Bioorganic & Medicinal Chemistry Letters 19 (2009) 6074–6077
Contents lists available at ScienceDirect
Bioorganic & Medicinal Chemistry Letters
journal homepage: www.elsevier.com/locate/bmcl
Synthesis and evaluation of (S,S)-N,N0-bis-[3-(2,20,6,60-tetramethylbenzhydryloxy)-
2-hydroxy-propyl]-ethylenediamine (S2824) analogs with
anti-tuberculosis activity
Xuelian Zhang a, , Yanwei Hu b,c, , Shudan Chen a, Rusong Luo a, Jun Yue d, Ying Zhang e, Wenhu Duan f,
Honghai Wang a,
*
a State Key Laboratory of Genetic Engineering, Institute of Genetics, School of Life Sciences, Fudan University, Shanghai, China
b Department of Medicinal Chemistry, Fudan University, Shanghai, China
c School of Pharmacy, Medical College of Soochow University, Suzhou, China
d Department of Clinical Laboratory, Shanghai Pulmonary Hospital, Shanghai, China
e Department of Molecular Microbiology and Immunology, Bloomberg School of Public Health, Johns Hopkins University, Baltimore, United States
f Department of Medicinal Chemistry, Shanghai Institute of Materia Medica, Chinese Academy of Sciences, Shanghai, China
a r t i c l e i n f o
a b s t r a c t
Article history:
Received 25 May 2009
Revised 27 August 2009
Accepted 11 September 2009
Available online 17 September 2009
In order to identify new and potent candidate drugs to treat tuberculosis, a library of compounds was
screened, and (S,S)-N,N0-bis-[3-(2,20,6,60-tetramethylbenzhydryloxy)-2-hydroxy-propyl]-ethylenedia-
mine (S2824) was identified as a hit in the screen. This research discusses our efforts to synthesize
and test 30 analogs of this hit for activity against Mycobacterium tuberculosis. Two compounds with
homopiperazine ring possess high in vitro activity against drug sensitive and resistant M. tuberculosis
with MICs 0.78–3.13 lg/mL (or 1.22–4.88 lM).
Keywords:
Ó 2009 Elsevier Ltd. All rights reserved.
Antitubercular drug
Resistance
MIC
Tuberculosis is one of the leading causes of death worldwide
due to an infectious agent. The WHO estimates that there are about
9.2 million new cases of active TB disease each year and about 1.7
million deaths.1 The rising incidence of tuberculosis, especially the
multidrug resistant tuberculosis (MDR-TB), has complicated the
global fight against TB. Consequently there is an urgent need to call
for the development of new drugs with potent antimycobacterial
activity.
and screened from diamine analogues of EMB.2–5 Compound
S2824 was then selected for further investigation. This Letter
describes our efforts to synthesize and test a series of compounds
based on the (S,S)-N,N0-bis-[3-(2,20,6,60-tetramethylbenzhydryloxy)-
2-hydroxy-propyl]-ethylenediamine for activity against M. tubercu-
losis in vitro. The effects of the most potent hit compounds in drug
sensitive and drug resistant strains from clinical isolates of M. tuber-
culosis and cytotoxicity were also investigated.
In order to identify new and potent candidate drug to treat tuber-
culosis, we used high-throughput screening from our chemical li-
brary to discover potential new antituberculosis agents. One
compound S2824, (S,S)-N,N0-bis-[3-(2,20,6,60-tetramethylbenzhyd-
ryloxy)-2-hydroxy-propyl]-ethylenediamine (1) (Fig. 1) was identi-
fied as a lead compound, showing interesting activity against
The modification of compounds S282401–S282430 focused on
replacement of benzyl and ethylenediamine in compound 1 by var-
ious aromatic moieties (substituted aryl groups, bicycloaryl
groups, heteroaryl) and piperazine or homopiperazine group,
respectively. Derivatives S282401-S282420 was synthesized by
diarylmethanol 2 with (R)-2-(chloromethyl) oxirane or (S)-2-(chlo-
Mycobacterium tuberculosis with an MIC = 6.25 lg/mL. According
to the structure research, we find that it is structurally similar to eth-
ambutol (EMB) shown in Figure 1; EMB is commonly used as one of
the first-line drugs for antituberculosis therapy. In addition, SQ109
(N-geranyl-N0-(2-adamantyl)ethane-1,2-diamine), which has excel-
lent in vitro and vivo activity against M. tuberculosis, is synthesized
* Corresponding author. Tel.: +86 21 65643777; fax: +86 21 65648376.
E-mail address: hhwang@fudan.edu.cn (H. Wang).
Figure 1. Chemical structures of (S,S)-N,N0-bis-[3-(2,20,6,60-tetramethylbenzhyd-
These authors contributed equally to this work.
ryloxy)-2-hydroxy-propyl]-ethylenediamine (S2824) and ethambutol (EMB).
0960-894X/$ - see front matter Ó 2009 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2009.09.035

X. Zhang et al. / Bioorg. Med. Chem. Lett. 19 (2009) 6074–6077
6075
romethyl) oxirane to give intermediate 3, followed by alkylation
of ethylenediamine generated S282401–S282420 (Scheme 1).
Moreover, there is an inversion of configuration in the reaction of
alkoxides with chloromethyloxirane (Scheme 2). In order to
synthesize derivatives S282421–S282424, coupling reaction was
carried out with furan or thiophene 4–5 and 2-furaldehyde or
2-thiophenecarboxaldehyde 6–7 to obtain difuran-2-ylmethanol
or dithiophen-2-ylmethanol 8–9.6,7 Condensation of 8 or 9 with
(R)-2-(chloromethyl) oxirane or (S)-2-(chloromethyl) oxirane gave
intermediate 10–13, followed by alkylation of ethylenediamine
which generated S282421–S282424 (Scheme 3). S282425–
S282430 were formed by reaction of compound 3 and piperazine
or homopiperazine (Scheme 4). All the compounds described in
this Letter were purified using silica gel column chromatogra-
phy, and the structures were confirmed with 1H NMR and HRMS.
We tested 30 analogs for their in vitro activity against H37Ra at
first. In general, all the compounds showed anti-tuberculosis
activity. Results showed that the most moderate activity was
Scheme 4. Reagent and condition: CH3OH, 60 °C.
[3-(4,40-dimethoxybenzhydryloxy)-2-hydroxy-propyl]-ethylenedia-
mine (S282402), (R,R)-N,N0-bis-[3-(4,40-dimethylbenzhydryloxy)-
2-hydroxy-propyl]-ethylenediamine (S282405), (S,S)-N,N0-bis-[3-
(4,40-dimethylbenzhydryloxy)-2-hydroxy-propyl]-ethylenediamine
(S282406) and (S,S)-N,N0-bis-[3-(4,40-dibromobenzhydryloxy)-2-hy-
observed with MICs 25.0 l
g/mL. The MICs for (S,S)-N,N0-bis-
droxy-propyl]-ethylenediamine (S282408) were 3.13
lg/mL. The
best MICs of 0.78 g/mL (or 1.22 M) were found for compounds
l
l
S282429 and S282430. The data, summarized in Table 1, indicated
that small groups such as bromo (S282407 and S282408) or methyl
(S282405 and S282406) can further enhance potency. However, lar-
ger groups such as tertiary butyl (S282419 and S282420) or oxyiso-
propyl (S282416 and S282417) caused reduction in potency. When
benzene ring was replaced by heteroaromatic, the compounds
showed less antimycobacterial activity. Activity could be improved
when ethylenediamine was replaced by homopiperazine ring. The
data showed in Table 1 indicate that the MIC of S282428 was
1.56 lg/mL (or 2.5 lM) and S282427 displayed relatively moderate
activity, although they were only different in configuration.
To evaluate whether the anti-TB activity shown by compounds
was bacteriostatic or cidal, MBCs and MICs of active compounds
Scheme 1. Reagents and conditions: (a) (R)-2-(chloromethyl)oxirane or (S)-2-
(chloromethyl)oxirane/50% NaOH–H2O, rt; (b) ethylenediamine/CH3OH, 60 °C.
with MICs equivalent to and lower than 3.13 lg/mL against
H37Ra strain were determined for strain H37Rv. A compound is
generally considered to be bactericidal if the ratio of MIC to MBC
is 64.8 As shown in Table 2, S282405, S282406 and S282430 could
be considered to be bactericidal due to the low ratios obtained. On
the other hand, S282428 and S282429 induced moderate bacterici-
dal activity at concentrations equivalent to their MIC. However,
S282402 and S282408 appeared to be primarily bacteriostatic.
Three of the most potent hit compounds with good antimyco-
bacterial activity were subjected to the determination of MICs
against drug-sensitive and MDR clinical isolates of M. tuberculosis
by BACTEC 460 system.9 Data in Table 3 showed that S282429
and S282430 were highly active against all the isolates tested
and were equally active against the drug-sensitive and MDR-TB,
Scheme 2. Inversion of configuration in the reaction of alkoxides with
chloromethyloxirane.
with MICs ranging from 0.78 to 3.13 lg/ml (or 1.22–4.88 lM).
Compared with the activity of EMB, the hit compounds were
more active against drug-resistant strains of M. tuberculosis. More-
over, the MICs range of S282429 and S282430 do not change for
MDR strains compared to drug-sensitive strains of M. tuberculosis.
The susceptibilities of MDR strains can be indicated by the radios of
MICs against resistance and sensitive strains,8 which were gener-
ally ꢀ2 for S282429 and S282430. This indicated that there is a lit-
tle cross-resistance with current anti-TB drugs thereby supporting
a novel mechanism of action. Four strains tested were resistant to
isoniazid, rifampicin, streptomycin and ethambutol, indicating that
S282429 and S282430 will maintain activity on MDR strains. How-
ever, S282428 showed moderate activity against drug-resistant
strains compared with S282429 and S282430 for the activity on
the drug sensitive stains and the drug-resistant strains varied
2–4-fold.
Scheme 3. Reagents and conditions: (a) n-BuLi/THF, ꢁ78 °C; (b) (R)-2-(chloro-
methyl) oxirane or (S)-2-(chloromethyl) oxirane/50% NaOH–H2O, rt; (c) ethylene-
diamine/CH3OH, 60 °C.

6076
X. Zhang et al. / Bioorg. Med. Chem. Lett. 19 (2009) 6074–6077
Table 1
Structures and in vitro activity against M. tuberculosis H37Ra
Compound
R
R1
R2
Iso
MICa
lg/mL (or lM)
S282401
S282402
S282403
S282404
S282405
S282406
S282407
S282408
S282409
S282410
S282411
S282412
S282413
S282414
S282415
S282416
S282417
S282418
S282419
S282420
OCH3
OCH3
CH3OCH2O
CH3OCH2O
CH3
CH3
Br
Br
OCH3
H
H
H
H
H
H
H
H
OCH3
OCH3
H
H
H
H
H
H
H
H
H
H
H
4R,11R
4S,11S
4R,11R
4S,11S
4R,11R
4S,11S
4R,11R
4S,11S
4R,11R
4S,11S
12.5
4R,11R
4S,11S
12.5
4S,11S
4R,11R
4S,11S
4R,11R
4R,11R
4S,11S
6.25 (9.46)
3.13 (4.73)
6.25 (8.0)
6.25 (8.0)
3.13 (5.24)
3.13 (5.24)
6.25 (7.34)
3.13 (3.67)
12.5 (16.0)
12.5 (16.0)
12.5 (16.17)
6.25 (8.72)
6.25 (8.72)
12.5 (17.44)
6.25 (8.72)
6.25 (8.09)
12.5 (16.18)
12.5 (13.87)
25.0 (32.67)
12.5 (16.34)
H
H
OCH3
OCH2CH2O
OC2H5
OC2H5
OCH2O
OCH2O
OC3H7-i
OC3H7-i
OCH3
4R,11R
H
H
4R,11R
H
H
H
OCH3
H
H
H
H
H
OCH3
H
H
t-C4H9
t-C4H9
S282421
S282422
S282423
S282424
—
—
—
—
—
—
—
—
4R,11R
4S,11S
4R,11R
4S,11S
25.0 (49.95)
12.5 (24.97)
12.5 (22.13)
12.5 (22.13)
S282425
S282426
S282427
S282428
S282429
S282430
Br
Br
CH3
CH3
CH3
CH3
—
—
—
—
—
—
—
—
—
—
—
—
4S,11S
4R,11R
4S,11S
4R,11R
4R,11R
4S,11S
25.0 (28.33)
25.0 (28.33)
25.0 (40.14)
1.56 (2.50)
0.78 (1.22)
0.78 (1.22)
a
MICs were determined for H37Ra as described in Supplementary data. MIC of ethambutol (EMB) was 1.56 lg/mL or 7.64 lM.
According to the research, anti-TB activity was improved when
tic window for the selective killing of M. tuberculosis with these
three compounds.
ethylenediamine was replaced by homopiperazine ring, which sug-
gest a synthetic direction for further improvement of antitubercu-
losis activity. Furthermore, active compounds including S282428,
S282429 and S282430 were also assessed for cytotoxicity using
an in vitro assay with the THP-1 cell line. In generally, the selectiv-
ity indices (SI) obtained by dividing the cytotoxic IC50 values by the
broth MICs were higher than 10, which suggests that the com-
pounds may be considered suitable for further screening. The com-
pounds S282428, S282429 and S282430 with SIs were 12, 26.92,
and 30.77, respectively, suggesting that there is a larger therapeu-
In summary, high-throughput screening of a chemical library
identified a hit compound with anti-tuberculosis activity and an
unknown mechanism of action. Based on this hit compound, a
series of compounds were synthesized and the in vitro antituber-
culosis activity investigated. S282429 and S282430 with homopip-
erazine ring displayed better efficacies for M. tuberculosis in vitro
including activity for MDR-TB clinical isolates (MIC = 0.78–
3.13
lg/mL) and higher SI values (>10). Further development of
these compounds includes modification of S282429 and S282430,

X. Zhang et al. / Bioorg. Med. Chem. Lett. 19 (2009) 6074–6077
6077
Table 2
clear indication that the promising activities of these compounds
may be important for development of much needed drugs for im-
proved treatment of MDR/XDR-TB.10,11
MICs and MBCs against M. tuberculosis H37Rv
Compound
H37Rv
MICa
(
lg/mL)
MBCb
(
lg/mL)
MBC/MIC
S282402
S282405
S282406
S282408
S282428
S282429
S282430
EMB
3.13
3.13
3.13
3.13
1.56
0.78
0.78
1.56
25
8
2
2
8
4
4
2
2
Acknowledgements
6.25
6.25
25
6.25
3.13
1.56
3.13
This work was supported by the 863 high-tech project
(2007AA02Z316), the national natural science foundation of China
(30901828) Shanghai science and technology funding (06JC14012)
and the severe infectious disease project (2008ZX10003-006-2).
a
MICs were determined for H37Rv as described in Supplementary data.
The MBC was determined for H37Rv strain by sub-culturing onto drug-free
Supplementary data
b
solid medium and enumeration of CFU.
Supplementary data (the detailed synthetic and microbiological
procedures, as well as full spectral characterization of the new
compounds) associated with this article can be found, in the online
version, at doi:10.1016/j.bmcl.2009.09.035.
Table 3
MICs of compounds for drug sensitive and resistant clinical isolates of M. tuberculosis
Isolate resistance profilea
MIC (
lg/mL)
S282428
S282429
S282430
EMB
References and notes
Sensitive
Sensitive
Sensitive
INH,RIF
INH,RIF
INH,RIF
INH,RIF,EMB,STR
INH,RIF,EMB,STR
INH,RIF,EMB,STR
INH,RIF,EMB,STR
3.13
3.13
3.13
6.25
12.5
6.25
6.25
12.5
6.25
6.25
0.78
1.56
1.56
1.56
3.13
3.13
1.56
3.13
3.13
3.13
0.78
1.56
1.56
3.13
1.56
1.56
1.56
3.13
0.78
1.56
3.13
3.13
3.13
6.25
3.13
3.13
12.5
>12.5
>12.5
>12.5
1. http://www.who.int/tb/en/. In World Health Organization Tuberculosis
Programme, 2007.
2. Nikonenko, B. V.; Protopopova, M.; Sarnala, R.; Einck, L.; Nacy, C. A. Antimicrob.
Agents Chemother. 2007, 51, 1563.
3. Lee, R. E.; Protopopova, M.; Crooks, E.; Slayden, R. A.; Terrot, M.; Barry, C. E., III J.
Comb. Chem. 2003, 5, 172.
4. Jia, L.; Tomaszewski, J. E.; Noker, P. E.; Gorman, G. S.; Glaze, E.; Protopopova, M.
J. Pharm. Biomed. Anal. 2005, 37, 793.
5. Protopopova, M.; Hanrahan, C.; Nikonenko, B.; Samala, R.; Chen, P.; Gearhart, J.;
Einck, L.; Nacy, C. A. J. Antimicrob. Chemother. 2005, 56, 968.
6. Hodgson, R.; Majid, T.; Nelson, A. J. Chem. Soc., Perkin Trans. 1 2002, 1631.
7. Gupta, I.; Ravikanth, M. J. Org. Chem. 2004, 69, 6796.
8. Villar, R.; Vicente, E.; Solano, B.; Perez-Silanes, S.; Aldana, I.; Maddry, J. A.;
Lenaerts, A. J.; Franzblau, S. G.; Cho, S. H.; Monge, A.; Goldman, R. C. J.
Antimicrob. Chemother. 2008, 62, 547.
a
Susceptibility of drug-resistant isolates to the following compounds were tes-
ted at Shanghai Pulmonary Hospital where the isolates were collected: The resis-
tance cutoffs for the following drugs were: isoniazid (INH) at 0.2 g/mL, rifampin
(RIF) at 2.0 g/mL, ethambutol (EMB) at 7.5 g/mL, streptomycin (STR) at 4.0 g/
l
l
l
l
9. Jagannath, C.; Allaudeen, H. S.; Hunter, R. L. Antimicrob. Agents Chemother. 1995,
39, 1349.
10. Lawn, S. D.; Wilkinson, R. Br. J. Med. 2006, 333, 559.
11. Goldman, R. C.; Plumley, K. V.; Laughon, B. E. Infect. Disord. Drug Targets 2007, 7,
73.
evaluation of the most active compounds in the mouse model of
tuberculosis infection. The data presented in this paper provide a