Syntheses and biological evaluation of new fluoroquinolone antibacterials containing chiral oxiimino pyrrolidine

Article abstract of DOI:10.1016/j.bmcl.2003.12.044

The design and syntheses of new fluoroquinolone antibacterial agents having pyrrolidine ring at C-7 position are described. The pyrrolidine ring is optically active and possesses methyloxime functional group. Two of them have excellent in vitro antibacterial activities and pharmacokinetic profiles.

Full text of DOI:10.1016/j.bmcl.2003.12.044

Bioorganic & Medicinal Chemistry Letters 14 (2004) 1273–1277
Syntheses and biological evaluation of new fluoroquinolone
antibacterials containing chiral oxiimino pyrrolidine
Dong Rack Choi,* Jung Han Shin, Jin Yang, Sue Hye Yoon and Yong Ho Jung
Central Research Laboratory, Dong-Wha Pharm. Ind. Co. Ltd, Anyang, Kyunggi, 430–017, South Korea
Received 16 October 2003;revised 9 December 2003;accepted 9 December 2003
Abstract—The design and syntheses of new fluoroquinolone antibacterial agents having pyrrolidine ring at C-7 position are
described. The pyrrolidine ring is optically active and possesses methyloxime functional group. Two of them have excellent in vitro
antibacterial activities and pharmacokinetic profiles.
# 2003 Elsevier Ltd. All rights reserved.
Quinolone antibacterial agents are among the most
attractive drugs in the anti-infective chemotherapy field.
To date many quinolone antibacterials have been
introduced into clinical use, and significant improve-
ments in antibacterial spectrum and activity have been
achieved. And since the discovery of Norfloxacin,1
fluoroquinolones have changed the landscape of anti-
microbial chemotherapy. Although the current fluoro-
quinolones are generally characterized by a broad
antimicrobial spectrum, their activities against clinically
important Gram-positive pathogens (including Staphy-
lococcus aureus, Streptococcus pyogens, Streptococcus
pneumoniae and Enterococcus) are relatively moderate.2
In addition, extensive use of fluoroquinolones has
brought increasing quinolone resistance to many
pathogens and majority of Methicillin-Resistant Sta-
phylococcus aureus (MRSA) have abtained resistance
to Ciprofloxacin (CPFX),3 a representative fluoro-
quinolone, as well as other antibacterials.
tiomer (R)-8 were chosen as C-7 substituents and
various quinolone and naphthyridone nuclei were
introduced.
In this report, we describe the synthesis and anti-
bacterial activity of a series of fluoroquinolones having
optically active oxiimino pyrrolidine at C-7 position.
Initially, optically inactive pyrrolidine derivative (8) was
prepared as depicted in Scheme 1. Methylation and
subsequent ketalization with neopentylglycol of readily
available compound 18 gave properly protected com-
pound 3. But at this stage, we had to swich N-protecting
group Cbz to Bn for the next steps harsh reaction con-
dition. And then, the conversion of ester group of com-
pound 4 to hydroxymethyl moiety was attempted by
several methods using borohydride and aluminium
hydride, thus lithium aluminium hydride and tetra-
hydrofuran turned out to be the best reaction condition.
Amination of the alcohol (5) was carried out by succes-
sive mesylation, azidation and azide reduction to fur-
nish compound 6. The primary amine of 6 was
protected with Boc and then selective N-deprotection of
compound 7 was performed by catalytic hydrogenation
to give compound 8.
On the basis of these considerations, we have focused
on development of compounds with good activity
against both Gram-positive and resistant organisms.
Inspired by three groups previous research results,4À6 we
planned to synthesize oxime-incorporated pyrrolidine
rings as C-7 substituents. And methyl group was intro-
duced into 3-position of pyrrolidine ring for the purpose
of increasing Gram-positive antibacterial activity.7 In
the course of our research, Compound 8 and its enan-
Condensation of 9a–e with 8 was carried out according
to the reactivities of 9a–e (Scheme 2). In case of 9a, a
condition under acetonitrile and triethylamine at 45 ꢀC
was the most suitable, but for 9b–d, additional heating
was needed, and for 9e, 1,8-diazabicyclo[5.4.0]-7-unde-
cene (DBU) was used instead of triethylamine. In the
next step, 10a–e were subjected to deprotection of ketal
Keywords: Antibacterial activity;fluoroquinolone;oxiimino pyrrolidine.
* Corresponding author. Tel.: +82-3144-52485;fax: +82-3144-69556;
e-mail: org301@iris.dwcrc.co.kr
0960-894X/$ - see front matter # 2003 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2003.12.044

1274
D. R. Choi et al. / Bioorg. Med. Chem. Lett. 14 (2004) 1273–1277
and Boc group simultaneously by means of concd. HCl
to provide 11a–e. Finally, Introducing methyloxime to
ketone of 11a–e gave 12a–e as biologically applicable
form. In addition, we also provided 14 (piperidine ring
instead of pyrrolidine), 15 (replaced methyloxime with
benzyloxime) and 16 (bulky ketal group in place of
oxime) to prove the necessity of the functional groups in
the C-7 substituent of 12a–e.
The antibacterial activities of the synthesized com-
pounds along with the reference agents CPFX and
Gemifloxacin (GMFX) against general strains are listed
in Table 1.
When we compared 12a with 14–16,10 piperidine ring
(14) showed significantly low Gram-negative activity
and exchanging methyloxime to bulky ketal group (16)
Scheme 1. Reagents and conditions: (a) Methyl iodide, K2CO3, acetone, 45 ꢀC, 3 h, 89%;(b) Neopentyl glycol, p-TSA, n-heptane, 120 ꢀC, 6 h, 97%;
(c) (i) H2(g), 10% Pd/C, MeOH, rt, 3 h;(ii) Benzyl chloride, K 2CO3, acetonitrile, reflux, overnight, 90% (two-step yield);(d) Lithium aluminium
hydride, THF, À5 ꢀC, 1.5 h, 93%;(e) (i) Methanesulfonyl chloride, triethylamine, MC, 0 ꢀC–rt, 2 h;(ii) Sodium azide, DMF, reflux, overnight;
(iii) Triphenylphospine, H2O, THF, reflux, 5 h, 73% (three-step yield);(f) Di- tert-butyl dicarbonate, MC, 0 ꢀC–rt, 1 h, 94%;(g) H (g), 10% Pd/C,
2
MeOH, rt, 4 h, 91%.
Scheme 2. Reagents and conditons: (a) 9a: Triethylamine, acetonitrile, 45 ꢀC, 1.5 h, 89%; 9b–d: Triethylamine, acetonitrile, reflux, overnight, 80–83%;
9e;1,8-diazabicyclo[5.4.0]-7-undecene, acetonitrile, reflux, overnight, 78%;(b) concd HCl, rt, overnight, 70–89%;(c) methoxylamine hydrochlor ide,
pyridine, 45 ꢀC, 4 h, 81–85%.

D. R. Choi et al. / Bioorg. Med. Chem. Lett. 14 (2004) 1273–1277
Table 1. In vitro antibacterial activities against general strains9
1275
Compd
Minimum inhibitory concentration (mg/mL)
S.p. 308A
S.p. 77A
S.a. SG511
S.a. 285
E.c. 078
E.c. DC 2
P.a. 1592E
P.a. 1771M
S.t.
E.cl. 1321 E
12a
12b
12c
12d
12e
14
15
16
(R)-12a
(S)-12a
(R)-12e
(S)-12e
CPFX
GMFX
0.098
0.098
0.098
0.195
0.013
6.250
0.195
0.391
0.098
0.195
0.004
0.049
3.125
0.195
0.025
0.013
0.025
0.025
<0.002
6.250
0.025
0.098
0.013
0.098
<0.002
0.013
0.391
0.025
0.004
0.007
0.013
0.007
<0.002
0.781
<0.002
0.013
0.004
0.007
0.013
0.013
0.007
<0.002
1.563
<0.002
0.025
0.007
0.004
0.013
0.025
0.007
<0.002
1.563
0.049
0.195
0.025
0.025
<0.002
0.004
0.004
0.007
0.049
0.025
0.049
0.025
0.007
12.500
0.098
0.195
0.049
0.098
<0.002
0.013
0.098
0.025
0.781
0.781
1.563
0.781
0.391
12.500
6.250
6.250
1.563
0.781
0.391
0.391
0.195
0.391
0.391
0.391
0.781
0.195
0.195
6.250
1.563
3.125
0.781
0.391
0.195
0.195
0.098
0.195
0.007
0.013
0.025
0.004
0.013
0.013
0.025
0.004
<0.002
0.781
0.098
0.195
0.013
0.013
<0.002
<0.002
0.013
<0.002
0.781
0.049
0.098
0.007
0.007
<0.002
<0.002
<0.002
0.004
0.049
0.049
<0.002
<0.002
0.195
<0.002
0.004
0.781
0.013
0.025
0.004
S.p.: Streptococcus pyogenes, S.a.: Staphylococcus aureus, E.c.: Escherichia coli, P.a.: Pseudomonas aeruginosa, S.t.: Salmonella typhimurium, E.cl.:
Enterobacter cloacae.
could not maintain activity against both Gram-positive
and negative. And benzyloxime group (15) also did not
improve Gram-negative activity. Compared with GMFX,
12a demonstrated improved Gram-positive antibacterial
activity as we expected. When it comes to quinolone
nuclei, all members of the series (12a–e) had more
potent activities than CPFX and GMFX, and 8-halo-
genated quinolone nuclei (12c and 12d) showed slightly
better antibacterial efficacy than 12b.
them have excellent advantages in maximal concen-
tration in blood, half-life period and area under curve
over GMFX and CPFX used as control.
It was reported that introduction of methyl group into
3-position of pyrrolidine could cause increment of
cytotoxicity against mammalian cell as well as Gram-
positive activity.7 (R)-12a showed similar cytotoxity
with GMFX, but, (R)-12e was more cytotoxic than
GMFX (Table 4).
Especially, 5-amino-8-fluoro derivative (12e) demon-
strated extremely high activity and naphthyridone
nucleus derivative (12a) also gave strong and broad
antibacterial spectrum. So at this point, we selected 12a
and 12e for further study and tried to get chiral sub-
stituent for biological evaluation. Simply, we attatched
N-tosyl protected l-proline to the primary amine of 6
and provided diastereomer 13 as separable form by col-
umn chromatography (Scheme 3). Through column
chromatography and base-catalyzed hydrolysis, opti-
cally active (R)-6 and (S)-6 were obtained and we
assigned their absolute configuration by X-ray crystal-
lography.11 Following the each chemical modification
described in Scheme 1 and 2, we could also easily pro-
vide (R)-12a(DW-286a), (S)-12a, (R)-12e and (S)-12e
from (R)-6 and (S)-6.12
In summary, new optically active quinolone anti-
bacterial agent (R)-12a and (R)-12e were designed and
synthesized. They displayed far more potent anti-
bacterial activity than CPFX and GMFX on important
The MIC data against general strains were also listed in
Table 1 and there were slight improvements in Gram-
positive antibacterial activities for (R)-12a and (R)-12e.
But for resistant strains including MRSA and Oflox-
acin13 resistant strains, (R)-12a and (R)-12e exhibited
far superior antibacterial activities to CPFX and
GMFX (Table 2). Generally, R-form enantiomers [(R)-
12a and (R)-12e] were more potent than the S-form
counterparts [(S)-12a and (S)-12e] by 10-fold and their
racemates (12a and 12e) had intermediate activities in
MIC.
Scheme 3. Reagents and conditions: (a) Ethyl chloroformate, triethyl-
amine, MC, 0 ꢀC–rt, 2 h, 86%;(b) column chromatography (eluent:
ethyl acetate/n-Hexane=2/3);(c) Potassium hydroxide, iso-propyl
alcohol, reflux, overnight, quant.
The phamacokinetic profiles of (R)-12a and (R)-12e
were examined to know whether they could be applied
as useful drugs to the body (Table 3). In rats, both of

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D. R. Choi et al. / Bioorg. Med. Chem. Lett. 14 (2004) 1273–1277
Table 2. In vitro antibacterial activities against drug-resistant strains8
Strain
Minimum inhibitory concentration (mg/mL)
12a
(R)12a
(S)12a
12e
(R)12e
(S)12e
CPFX
GMFX
MRSA
690 E
692 E
697 E
701 E
705 E
707 E
0.004
<0.002
0.007
0.007
0.013
<0.002
<0.002
<0.002
<0.002
<0.002
0.004
0.025
0.025
0.025
0.049
0.049
0.013
<0.002
<0.002
<0.002
<0.002
0.004
<0.002
<0.002
<0.002
<0.002
<0.002
<0.002
0.004
0.004
0.004
0.007
0.007
<0.002
0.391
0.391
0.391
0.391
0.391
0.195
0.098
0.049
0.049
0.098
0.098
0.025
<0.002
<0.002
Ofloxacin Resistant
S.a. 179
S.a. 293
S.a. 303
S.e. 291
S.e. 319
E.k. 101
P.a. 279II
K.s. 30-92
0.195
0.195
0.195
0.391
0.391
0.098
6.250
1.563
0.098
0.098
0.098
0.195
0.391
0.098
6.250
1.563
1.563
1.563
0.781
3.125
6.250
0.391
6.250
3.125
0.025
0.025
0.025
0.025
0.049
0.025
6.250
0.781
0.013
0.013
0.013
0.049
0.025
0.013
3.125
0.781
0.195
0.195
0.195
0.391
0.781
0.195
3.125
1.563
12.500
12.500
12.500
50.000
100.00
0.781
0.781
0.781
0.781
1.563
3.125
0.098
6.250
0.781
6.250
1.563
S.a.: Staphylococcus aureus, S.e.: Staphylococcus epidermidis, E.k.: Enterococcus knothe, P.a.: Pseudomonas aeruginosa, K.s.: Klebsiella species.
Table 3. Pharmacokinetics of (R)-12a and (R)-12e in rats
Parameter
(R)-12a
(R)-12e
GMFX
CPFX
Cmax (mg/mL)
Tmax (h)
T1/2 (h)
9.06Æ2.040
2.00
4.50
6.67Æ3.327
1.00
6.94
6.63
1.00
2.12
25.79
4.39Æ1.220
0.50
2.07
.
AUC (mg h/mL)
90.82
68.77
12.72
SD rat, administered at a dose of 50mg/kg
Overview and Conclusions Ibid. pp 527–535. (c) Cecchetti,
V.;Fravolini, A.;Lorenzini, M. C.;Tabarrini, O.;Terni,
P.;Xin, T. J. Med. Chem. 1996, 39, 436.
Table 4. Cytotoxicity of (R)-12a and (R)-12e (IC50, mg/mL)
(R)-12a
(R)-12e
GMFX
Trovafloxacin
CPFX
3. (a) Jaynes, B. H.;Dirlam, J. P.;Hecker, S. J. Annu. Rep.
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V79-4
HepG2
5.49
37.26
0.21
11.59
10.08
13.49
3.77
11.16
58.55
37.47
V79-4 and HepG2 were incubated in the presence of each sample for
72 h. Cell density was determined by MTT assay or by staining the
cells with crystal violet.
Gram-positive organisms, MRSA and Ofloxacin resis-
tant organisms. And with excellent pharmacokinetic
profiles, both (R)-12a and (R)-12e are expected to be the
best drug candidates near future. Now, we make efforts
to optimize the synthetic scheme of them and develop
better C-7 substituents.
5. Nakano, J.;Fukui, H.;Haigoh, H.;Senda, H.;Iwatani,
W.;Arika, T. E.P. 0541086, Dec 5, 1993.
6. (a) Hong, C. Y.;Kim, Y. K.;Lee, Y. H.;Kwak, J. H.
Bioorg. Med. Chem. Lett. 1998, 8, 221. (b) Hong, C. Y.;
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8. Coupling glycine ethylester hydrochloride with ethyl
acrylate in the presence of triethylamine, Cbz protection
and potassium tert-butoxide assisted cyclization can give
the indicated starting material 1 (1-benzyl 3-ethyl 4-oxo-
1,3-pyrrolidinedicarboxylate).
Acknowledgements
This study was supported by a grant of the Korea
Health 21 R&D Project, Ministry of Health & Welfare,
Republic of Korea (01-PJ1-PG4-01PT01-0013).
9. All cultures were obtained from Hoechst and MICs were
determined by the agar dilution method as recommended by
the National Committee for Clinical Laboratory Standards.
10. 14: 7-(3-Aminomethyl-4-methoxyimino-3-methyl-piperidin-
1-yl)-1-cyclopropyl-6-fluoro-4-oxo-1,4-dihydro-[1,8]naph-
thyridine-3-carboxylic acid hydrochloride, 15: 7-(3-Ami-
nomethyl-4-benzyloxyimino-3-methyl-pyrrolidin-1-yl)-1-
cyclopropyl-6-fluoro-4-oxo-1,4-dihydro-[1,8]naphthyridine-
3-carboxylic acid hydrochloride, 16: 7-(4-Aminomethyl-
4,8,8-trimethyl-7,9-dioxa-2-aza-spiro[4.5]dec-2-yl)-1-cyclo-
References and notes
1. Koga, H.;Itoh, A.;Murayama, S.;Suzue, S.;Irikura, T.
J. Med. Chem. 1980, 23, 1358.
2. (a) Peterson, L. P. Quinolone Resistance in Clinical Prac-
tice: Occurrence and Importance. In Quinolone Anti-
bacterial Agents, 2nd ed.;Hooper, D. C., Ed. American
Society for Microbiology: Washington, DC, 1993;p 119.
(b) Moellering, R. C., Jr. Quinolone Antimicrobial Agents;

D. R. Choi et al. / Bioorg. Med. Chem. Lett. 14 (2004) 1273–1277
1277
propyl-6-fluoro-4oxo-1,4-dihydro-[1,8]naphthyridine-3-
carboxylic acid hydrochloride.
1.20 (2H, d, J=7.3 Hz), 1.34 (3H, s), 3.08 (1H, d, J=13.2
Hz), 3.14 (1H, d, J=13.2 Hz), 3.15 (2H, m), 3.66 (1H, bs),
3.86 (4H, bs), 4.08 (1H, d, J=12.7 Hz), 4.61 (2H, s), 8.99
(1H, d, J=12.4 Hz), 8.56 (1H, s), [a]D=+5.77 (c=2.33,
H2O, 25 ꢀC), (S)-12a: [a]D=À3.74 (c=1.32, H2O, 25 ꢀC),
(R)-12e: (R)-(À)-5-Amino-7-(3-aminomethyl-4-methoxy-
imino-3-methyl-pyrrolidin-1-yl)-1-cyclopropyl-6,8-difluoro-
4-oxo-1,4-dihydro-quinoline-3-carboxylic acid hydrochlo-
11. We could obtain single crystal of Cbz substituted (S)-13
derivative [(S)-4,8,8-trimethyl-4-({[1-(toluene-4-sulfonyl)-
pyrrolidine-2-carbonyl]-amino}-methyl)-6,10-dioxa-2-aza-
spiro[4.5]decane-2-carboxylic acid benzyl ester, the chiral
centre of this compound has same absolute configuration
to that of (S)-13] to assign absolute configuration of the
chiral centre of (R)-13 and (S)-13. Crystal data:
C31H14O7N3S from Ethanol and Butanol solution, F.W.=
599.75, triclinic, a=10.1966; b=12.2398; c=13.6633;
1
ride, H NMR (DMSO-d6+CF3COOD, ppm) 0.98 (2H,
bs), 1.03 (2H, d, J=6.8 Hz), 1.28 (3H, s), 3.00 (1H, d,
J=13.2 Hz), 3.05 (1H, d, J=13.2 Hz), 3.59 (1H, d,
J=10.8 Hz), 3.79 (4H, bs), 3.91 (1H, bs), 4.25 (1H, d,
J=17.3 Hz), 4.41 (1H, d, J=17.3 Hz), 8.45 (1H, s),
[a]D=À15.89 (c=0.54, H2O, 25 ꢀC), (S)-12e: [a]D=+17.12
(c=1.10, H2O, 25 ꢀC).
a=69.007; b=88.535; g=86.991; V=1589.823, dcalc
=
1.253g/cm3, Radiation=Mo Ka(l=0.71069), Space
group=P1, Z=2, Final R=0.0793.
12. (R)-12a: (R)-(+)-7-(3-Aminomethyl-4-methoxyimino-3-
methyl-pyrrolidin-1-yl)-1-cyclopropyl-6-fluoro-4-oxo-1,4-
dihydro-[1,8]naphthyridine-3-carboxylic acid hydrochlo-
ride, 1H NMR(DMSO-d6+CF3OOD, ppm) 1.05 (2H, bs),
13. Sato, K.;Matsuura, Y.;Inone, M.;Une, T.;Osada, Y.;
Ogawa, H.;Mitsuhashi, S. Antimicrob. Agents. Chemother.
1982, 22, 548.