Pyrrolamide DNA gyrase inhibitors: Optimization of antibacterial activity and efficacy

Article abstract of DOI:10.1016/j.bmcl.2011.10.010

The pyrrolamides are a new class of antibacterial agents targeting DNA gyrase, an essential enzyme across bacterial species and inhibition results in the disruption of DNA synthesis and subsequently, cell death. The optimization of biochemical activity an

Full text of DOI:10.1016/j.bmcl.2011.10.010

Bioorganic & Medicinal Chemistry Letters 21 (2011) 7416–7420
Contents lists available at SciVerse ScienceDirect
Bioorganic & Medicinal Chemistry Letters
journal homepage: www.elsevier.com/locate/bmcl
Pyrrolamide DNA gyrase inhibitors: Optimization of antibacterial activity
and efficacy
⇑
Brian A. Sherer , Kenneth Hull, Oluyinka Green, Gregory Basarab, Sheila Hauck, Pamela Hill,
James T. Loch III, George Mullen, Shanta Bist, Joanna Bryant, Ann Boriack-Sjodin, Jon Read,
Nancy DeGrace, Maria Uria-Nickelsen, Ruth N. Illingworth, Ann E. Eakin
Infection Innovative Medicines Unit, AstraZeneca R&D Boston, 35 Gatehouse Drive, Waltham, MA 02451, USA
a r t i c l e i n f o
a b s t r a c t
Article history:
The pyrrolamides are a new class of antibacterial agents targeting DNA gyrase, an essential enzyme
across bacterial species and inhibition results in the disruption of DNA synthesis and subsequently, cell
death. The optimization of biochemical activity and other drug-like properties through substitutions to
the pyrrole, piperidine, and heterocycle portions of the molecule resulted in pyrrolamides with improved
cellular activity and in vivo efficacy.
Received 8 August 2011
Revised 3 October 2011
Accepted 4 October 2011
Available online 12 October 2011
Ó 2011 Elsevier Ltd. All rights reserved.
Keywords:
Antibacterial
Gyrase
Pyrrolamide
The emergence of drug resistance in both community- and hos-
pital-acquired infections has outpaced development and delivery
of new antibacterial drugs to the clinic. As a result, there is a seri-
ous global health threat that currently available therapies will no
longer be effective in treating bacterial infections.1 One approach
to combating the emergence of resistance to current antibacterial
drugs is to discover novel agents that inhibit known drug targets
through a unique binding site, chemistry or mechanism of inhibi-
tion; thereby evading existing resistance mechanisms developed
by bacteria.
Previous efforts at AstraZeneca identified a novel class of DNA
gyrase inhibitors—the pyrrolamides—using fragment-based
a
NMR screening approach followed by a structure-guided lead iden-
6
tification strategy. The pyrrolamide series inhibits the enzyme by
binding to the ATP pocket of GyrB, as demonstrated by an initial
lead compound 1 (Fig. 1) with submicromolar enzyme potency
but weak antibacterial activity. Compound 2 (Fig. 1) is a subse-
quent compound in the pyrrolamide series that displayed promis-
ing antibacterial activity and demonstrated in vivo efficacy against
7
S. pneumoniae in a mouse pneumonia model. Described herein is
DNA gyrase has long been known as an attractive target for
antibacterial drugs. DNA gyrase is a member of the type II family
the elaboration of SAR for the pyrrolamide series that led to 2
and further optimization of the antibacterial potency to provide
26, which resulted in improved in vivo efficacy over that of 2.7
The synthesis of substituted pyrrolamide analogs is illustrated in
Scheme 1. The previously reported ethyl 2-methyl-5-carboxylate
2
of topoisomerases that control the topological state of DNA in
3
cells. It is a tetrameric enzyme comprised of two subunits each
of GyrA and GyrB. DNA gyrase couples ATP hydrolysis by the GyrB
subunit to negative supercoiling of DNA, which is required for
maintenance of DNA topology during the replication process.
DNA gyrase is an essential enzyme across bacterial species and
inhibition results in a disruption of DNA synthesis and ultimately,
cell death. Two classes of antibiotics have clinically validated DNA
gyrase as a viable target—the fluoroquinolones and aminocouma-
rins. However, increasing prevalence of resistant bacterial strains
is eroding the utility of these quite successful fluoroquinolone
8
pyrrole was treated with either N-bromo or N-chlorosuccinimide
followed by ester hydrolysis to obtain the 3-halosubstituted pyr-
roles 3 and 4. Ethyl 5-methyl-1H-pyrrole-2-carboxylate was also
subjected to sulfuryl chloride and followed by ester hydrolysis to
provide the 3,4-dichlorosubstituted pyrrole 5. A three step proce-
dure from ethyl 5-methyl-1H-pyrrole-2-carboxylate to provide the
corresponding aldehyde followed by formation of the hydroxyli-
mine and dehydration gave the 3-cyanopyrrole 6. Pyrroles 3, 4, 5,
and 6 were subjected to an amide coupling with commercially
available N-Boc-4-aminopiperidine. Finally, Boc-deprotection was
followed by nucleophilic substitution with commercially available
4
drugs. The coumarin class of antibiotics have failed to achieve
widespread utility because of poor pharmacokinetics and issues
5
of safety.
2
,6-dichloropyridine-4-carboxamide to yield compounds 1, 7–10.
The synthetic sequence used to prepare the pyrrolamides 2
⇑
Corresponding author. Tel.: +781839 4143.
E-mail address: brian.sherer@astrazeneca.com (B.A. Sherer).
and 12–16 is described in Scheme 2. Amide coupling of
0
960-894X/$ - see front matter Ó 2011 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2011.10.010

B. A. Sherer et al. / Bioorg. Med. Chem. Lett. 21 (2011) 7416–7420
7417
Br
O
Cl
Cl
H
N
H
NH2
N
N
N
N
N
H
O
N
N
H
O
S
OH
Cl
1
2
O
Figure 1. Structure of early lead compounds in the pyrrolamide series.
Cl
Cl
X
b
OEt
OH
a
OH
N
N
H
N
H
H
O
O
O
5
3
4
: X = Br
: X = Cl
c
NC
OH
R1
N
H
O
6
R1
R
2
R2
O
H
OH
N
d
NH2
N
N
N
H
O
H
O
N
3
- 5
Cl
2
1
7
8
9
1
: R1 = Br; R = H
: R = Cl; R2 = H
1
1
2
: R = CN; R = H
1
2
: R = R = H
0: R1 = R = Cl
2
Scheme 1. Synthesis of substituted pyrrole compounds 1, 7–10. Reagents and conditions: (a) (1) N-bromosuccinimide or N-chlorosuccinimide 0 °C, 38–50%; (2) NaOH,
MeOH, water, 70–92%; (b) (1) SO Cl , carbon tetrachloride, 77%; (2) NaOH, MeOH, water, 60–75%; (c) (1) trimethoxymethane, TFA, 45%; (2) hydroxylamine, ethanol then
2
2
thionyl chloride, DMF, 0 °C 80%; (3) NaOH, MeOH, water, 70% (d) (1) HATU, DIPEA, DMF, N-Boc-4-aminopiperidine, 70–95%; (2) HCl, dioxane, 98%; (3) 2,6-dichloropyridine-4-
carboxamide, 80 °C, 60–80%.
N-Boc-4-aminopiperidine and removal of the protecting group
provided intermediate 11. Nucleophilic substitution of the
corresponding halo-heterocycle with piperidine 11 followed by
hydrolysis of the ester yielded acids 2, 12–13. Compound 14
was analogously prepared by nucleophilic displacement of
chloro-oxadiazole. A reductive amination of intermediate 11
and trans-hydroxyproline ester followed by basic hydrolysis pro-
vided pyrrolidine 15. Formation of the amide from acid 2 under
standard conditions yielded amide 16.
Shown in Table 1 are the results of efforts to improve both the
1
2
enzyme and antibacterial activity of lead compounds in the pyr-
rolamide series by changing the substituents on the pyrrole por-
1
3
tion of the molecule. The 3,4-dichloropyrrole 10 provided the
best in vitro potency of 0.03 M while the methyl pyrrolamide
9 had the weakest activity with an IC50 of 7
3-substituted pyrroles 1, 7–9 had similar potencies of ꢀ0.15
M. The pyrrole group binds in the adenine binding pocket of
l
8
b
14
lM. All of the
l
GyrB forming hydrogen bond interactions with a conserved
aspartic acid 82 residue and water molecule as shown in the X-
ray co-crystal structure of 26 with the 24 kDa N-terminal ATP
binding domain of GyrB that had previously been used for analyz-
The synthesis of 3-fluoropiperdine analogs is described in
Scheme 3. Racemic cis- and trans-N-Boc-3-fluoro-4-benzylamin-
9
opiperidines (18a, 18b) were prepared as described previously.
1
5
The benzyl protecting group was removed from 18a and 18b and
the resulting amine was coupled to acid 5. Deprotection of the
piperidine, followed by nucleophilic substitution on methyl
ing inhibitor complexes (Fig. 2). The enhanced enzyme activity
of compound 10 relative to 1, 7–9 can be explained by the pres-
ence of two lipophilic electron withdrawing groups on the pyr-
role, which increase hydrophobic interactions in the adenine
2
-bromothiazole-5-carboxylate, and ester hydrolysis led to race-
mic cis- and trans-acids 19 and 20. Piperidine 18 was also subjected
to chiral chromatography10 to provide the pure cis-enantiomers 24
and 25, which were then independently subjected to the reaction
conditions described above to provide the enantiomerically pure
analogs 26 and 27. The absolute configuration of compound 26
was determined via small molecule X-ray crystallography to be
a
pocket and decrease the pK of the pyrrole nitrogen resulting in
a stronger hydrogen bond donation to the aspartate residue.
The decreased activity of the pyrrole 9 confirms the hypothesis
that an electron withdrawing group is preferred for increasing po-
tency. The enzyme potency correlated well with the cellular
activity as compound 10 displayed the best antibacterial activity
particularly against Gram-positive bacteria. All the compounds
1
1
(
3S,4R).

7
418
B. A. Sherer et al. / Bioorg. Med. Chem. Lett. 21 (2011) 7416–7420
Cl
Cl
Cl
Cl
Cl
Cl
H
N
H
N
a
b
OH
N
O
NH
N
H
N
H
Het
N
H
O
O
O
1
1
5
c
Het
d
OH
Cl
Cl
Cl
Cl
12
N
H
H
O
N
N
N
Cl
N
N
N
N
N
N
H
OH
H
O
O
O
N
H
S
2
OH
1
4
15
O
O
N
13
OH
Cl
Cl
S
H
Cl
Cl
N
O
2
H
N
N
N
e
N
N
H
N
N
S
H
O
OH
S
H
N
O
1
6
O
O
Scheme 2. Synthesis of heterocyclic compounds 2, 12–16. Reagents and conditions: (a) (1) HATU, DIPEA, N-Boc-4-aminopiperidine, DMF, 70–90%; (2) HCl, dioxane, 98%; (b)
1) DIPEA, DMF, methyl 2,6-dichloropyridine-4-carboxylate or methyl 2-bromothiazole-4-carboxylate or 2-bromothiazole-5-carboxylate, 80 °C, 63–79%; (2) NaOH, water,
(
MeOH, 88%; (c) preparation according to ref 7a; (d) (1) N-Boc-trans-4-hydroxy-L-proline methyl ester, NaCNBH
3
, 64% (2) HCl, dioxane, 95%; (3) NaOH, MeOH, water, 90%; (e)
HATU, DIPEA, DMF, methoxylamine, 78%.
Cl
Cl
NHBn
NBoc
NHBn
NBoc
H
N
F
F
a
+
N
N
N
H
O
F
S
1
8a
1
8b
19: rel(3R,4S)
OH
2
0: rel(3R,4R)
O
b
Cl
Cl
NHBn
NBoc
NHBn
a
H
F
F
N
+
N
N
N
H
NBoc
O
F
S
2
4
25
OH
2
7
O
a
Cl
Cl
H
N
N
N
N
H
O
F
S
OH
2
6
O
Scheme 3. Synthesis of 3-fluoropiperidines 19, 20, 26, 27. Reagents and conditions: (a) (1) 5, H
2
, Pd/C, MeOH, 82%; (2) HATU, DIPEA, DMF, 60%; (3) HCl, 1,4-dioxane, 95%; (4)
methyl 2-bromothiazol-5-carboxylate, 65%; (5) NaOH, MeOH, water, 87%; (b) chiral chromatograp.
displayed significantly lower activity against the Gram-negative
bacteria, Haemophilus influenzae and Escherichia coli. However,
all of the 3-substituted pyrroles except 9 displayed activity
an efflux mechanism limits the Gram-negative potency of the
more active DNA gyrase inhibiting compounds.
The structure activity relationships from efforts to further
optimize enzyme and cellular potency by replacement of the
À
against an efflux deficient strain of E. coli (tolC ), suggesting that

B. A. Sherer et al. / Bioorg. Med. Chem. Lett. 21 (2011) 7416–7420
7419
Table 1
Structure–activity relationship of pyrrole substitutions
Compd
S. aureus IC50
(lM)
Minimum Inhibitory Concentration (lg/mL)
S. aureus MRQRb
M. catarrhalis
E. faecium
H. influenzae
E. coli
E. coli tolC
À
S. pneumoniae
S. aureus
1
7
8
9
0.10
0.11
0.15
6.9
4
4
4
>64
1
32
32
32
>64
>64
4
8
ND
ND
>64
2
>64
>32
ND
>64
4
64
>64
>64
ND
>64
>64
4
8
ND
>64
2
a
>32
ND
>64
4
>64
>64
>64
>64
1
0
0.03
a
ND = Not determined.
MRQR = methicilin-resistant, quinolone resistant.
b
Figure 2. An X-ray structure of a 3-fluorosubstituted piperidine analog 26 with S. aureus GyrB displayed with surface (left) and stick (right) representations.
pyridine of 10 with other heterocycles are shown in Table 2. Five-
membered ring heterocycles, such as thiazoles, displayed the best
combination of physical properties and potency. Heterocycles 2,
pocket generally show promising cellular activity against Gram-
positive bacteria, including methicilin- and quinolone-resistant S.
aureus and fastidious Gram-negative bacteria, such as H. influenzae
and M. catarrhalis. Broader potency against enteric Gram-negative
pathogens such as E. coli remains hampered by efflux, as demon-
1
2–14, and 16 all maintained enzyme potency in the low nanomo-
lar range while compound 15 had weaker activity at the M level.
l
À
The heterocycles attached to the piperidine nitrogen extends out-
side of the ATP binding region, forming stacking and hydrogen
bonding interactions with protein residues as shown in Figure 2.
The aromatic nature of these heterocycles is key for optimizing
pi-stacking with Arg84. The aromatic heterocycles of all the com-
pounds except 15 are additionally substituted with carboxylate
or carboxamide groups which form hydrogen bond interactions
with Arg144 in the GyrB active site (Fig. 2). The lower enzyme po-
tency for 15 is due in part from the lack of capability for the pi-
stacking with Arg144 and in part from there being a poor align-
ment for the hydrogen bond between the carboxylate and Arg84.
An enhancement in antibacterial spectrum of compounds 2 and
strated by improved potency against an efflux-deficient (tolC )
strain.
The piperidine moiety of the pyrrolamides was investigated to
determine if any further improvements could be obtained in en-
zyme potency and antibacterial activity by incorporating a single
1
6
fluorine substituent at the 3-position (Table 3). The resultant ste-
reochemical relationship between the 3-F and the 4-amide substit-
uents on the piperidine proved to be crucial for improvement in
potency with the racemic cis-piperidine 19 being 10–20 times
more potent than the racemic trans-piperidine 20 in both enzyme
and anti-bacterial activity. Furthermore, the enantiomerically pure
(3S,4R) analog 26 displayed approximately eightfold greater po-
tency over the (3R,4S) enantiomer 27 in both enzyme and cellular
activities. Overall, the single fluorine atom incorporation in 26
improves activity 4–8 fold. The fluorine atom in 26 is directed to-
wards a hydrophobic pocket in the GyrB active site (Fig. 2), which
could account for the improvement in potency compared to unsub-
stituted analogs. Additionally, the fluorine atom could adjust the
piperidine conformation to better align the pyrrole carboxamide
1
2–13 to include fastidious Gram-negative bacteria, such as H.
influenzae and Moraxella catarrhalis, can potentially be attributed
to an increase in cell permeability imparted by the presence of a
carboxylate substituent on the heterocycle. Compounds such as
1
2 that maximize both stacking and hydrogen bond interactions
outside of the ATP pocket and, additionally, that optimize the pyr-
role hydrogen bond to Asp81 and that are hydrophobic in the ATP
Table 2
Structure–activity relationship of heterocycle substitutions
Compd
S. aureus IC50
(l
M)
Minimum Inhibitory Concentration (lg/mL)
À
S. pneumoniae
S. aureus
S. aureus MRQR
M. catarrhalis
E. faecium
H. influenzae
E. coli
E. coli tolC
1
1
2
1
1
1
1
0
2
0.03
0.01
0.0 2
0.02
0.05
0.23
129
1
4
2
8
8
8
32
>64
4
2
8
8
8
32
>64
2
4
1
2
4
8
32
>64
>64
4
2
>64
>64
>64
>64
>64
>64
>64
2
0.5
0.5
0.5
1
4
>64
0.13
0.5
0.5
1
4
>64
0.25
0.25
0.25
0.25
4
3
6
4
5
2
16
64
>64
>64

7
420
B. A. Sherer et al. / Bioorg. Med. Chem. Lett. 21 (2011) 7416–7420
Table 3
Structure–activity relationship of unsubstituted and 3-F substituted piperidines
Compd
S. aureus IC50
(l
M)
Minimum Inhibitory Concentration (lg/mL)
À
S. pneumoniae
S. aureus
S. aureus MRQR
M. catarrhalis
E. faecium
H. influenzae
E. coli
E. coli tolC
2
0.02
<0.01
0.11
0.004
0.035
0.5
0.13
8
0.06
2
8
2
>64
2
16
8
4
>64
2
32
0.25
<0.06
16
0.06
1
2
2
>64
>64
>64
>64
>64
0.25
0.13
4
0.06
0.5
1
2
2
2
9
0
6
7
0.5
32
0.25
4
0.5
64
0.25
4
Acknowledgments
8
7
6
5
4
3
2
The authors thank the following individuals for valuable techni-
cal contributions to this project: Barbara Arsenault, John Breed,
Amanda Doucette, Elise Gorseth, Grant Walkup, Bryan Prince, Lin-
da Otterson, Lena Grosser, Susie Hopkins, Larry MacPherson, and
Dedong Wu.
References and notes
1.
(a) Boucher, H. W.; Talbot, G. H.; Bradley, J. S.; Edwards, J. E., Jr.; Gilbert, D.;
Rice, B.; Scheld, M.; Spellberg, B.; Bartlett, J. Clin. Infect. Dis. 2009, 48, 1; (b)
Talbot, G. H.; Bradley, J.; Edwards, J. E., Jr.; Gilbert, D.; Scheld, M.; Bartlett, J. Clin.
Infect. Dis. 2006, 42, 657; (c) Alanis, A. J. Arch. Med. Res 2005, 36, 697.
Pre-tx
40 mg/kg
60 mg/kg
80 mg/kg
2. Maxwell, A. Trends Micro. 1997, 5, 102.
3.
Wang, J. C. Untangling the double helix; DNA entanglement the action of the DNA
topoisomerases; Cold Spring Harbor Laboratory Press: Cold Spring Harbor, NY,
vehicle
2009.
2
3
0 mg/kg
0 mg/kg
4.
(a) Biedenbach, D. J.; Farrell, D. J.; Mendes, R. E.; Ross, J. E.; Jones, R. N. Diagn.
Microbiol. Infect. Dis. 2010, 68, 459; (b) Rhomberg, P. R.; Jones, R. N. Diagn.
Microbiol. Infect. Dis. 2009, 65, 414; (c) Boyd, L. B.; Atmar, R.; Randall, G.; Hanill,
R.; Steffen, D.; Zechiedrich, L. B. M. C. Infect. Dis. 2008, 8, 4.
Figure 3. Dose–response effect on the treatment of mice with compound 26 in a S.
pneumoniae lung infection model. Error bars represent standard error in the CFU
measurements.
5
6
.
.
Maxwell, A. Mol. Microbiol. 1993, 9, 101.
Green, O.; Ni, H.; Singh, A.; Walkup, G.; Hales, N.; Breeze, A.; Eakin, A.E. 48th
Annual Meeting of the Interscience Conference on Antimicrobial Agents and
Chemotherapy, Washington, D.C. October 25–28, 2008, F1-2025.
Illingworth, R.N.; Uria-Nickelsen, M; Eakin, A.E. 48th Annual Meeting of the
Interscience Conference on Antimicrobial Agents and Chemotherapy,
Washington, D.C. October 25–28, 2008, F1-2028.
7
.
with Asp81 and the thiazole carboxylate with Arg84. The racemic
trans-piperidine 20 showed weak activity relative to the unsubsti-
tuted analog 2 consistent with the trans-isomer changing the con-
formation of the piperidine ring and disrupting optimal stacking
and hydrogen bonding interactions with the protein. The fluorine
atom of the (3R, 4S) enantiomer 27 is orientated towards a less
favorable interaction with the water molecule associated with
Asp81 rather than towards the hydrophobic pocket as for the more
active (3S, 4R) enantiomer.
Compound 26 was evaluated in vivo against S. pneumoniae in a
mouse model of pneumonia (Fig. 3).17 Compound 26 or vehicle was
dosed orally starting 18 h post-infection and activity was quanti-
fied by viable counts on serial dilutions of lung homogenates
8. (a) Curran, T. P.; Keaney, M. T. J. Org. Chem. 1996, 61, 9068; (b) Breeze, A.L.;
Green, O. M.; Hull, K. G.; Hauck, S. I.; Ni, H.; Mullen G. B.; Hales, N. J.; Timms, D.
WO2005026149.
9.
Van Niel, M. B.; Collins, I.; Beer, M. S.; Broughton, H. B.; Cheng, S. K. F.;
Goodacre, S. C.; Heald, A.; Locker, K. L.; MacLeod, A. M.; Morrison, D.; Moyes, C.
R.; O’Connor, D.; Pike, A.; Rowley, M.; Russell, M. G. N.; Sohal, B.; Stanton, J. A.;
Thomas, S.; Verrier, H.; Watt, A. P.; Castro, J. L. J. Med. Chem. 1999, 42, 2087.
1
0. Chiral chromatography conditions: Chiralpak AD 500 Â 50 mm, 20
hexanes, 5% methanol, 5% ethanol, 0.1% diethylamine; flow rate: 118 ml/min.
11. X-ray coordinates deposited in Cambridge Crystallographic Data Centre
835685.
l, 90%
#
1
2. Minimum Inhibitory Concentration measurements performed according to
Clinical and Laboratory Standards Institute, Wayne, PA on the following
bacterial strains from the AstraZeneca collection: Streptococcus pneumoniae
ARC548, Staphylcoccus aureus ARC516, S. aureus MRQR ARC517, Moraxella
catarrhalis ARC445, Enterococcus faecium ARC073, Haemophilus influenzae
ARC446, Escherichia coli ARC523, and E. coli tolC- ARC524.
2
4 h after start of the treatment and presented in Figure 3 as mean
log CFU/lung (+/- standard error) for each group of mice. Com-
pound 26 was found to be efficacious against S. pneumoniae in this
mouse model of pneumonia, causing a dose-dependent decrease in
viable bacterial counts in the lung. A dose of 80 mg/kg resulted in
the maximum response seen in this experiment and an ED50 was
estimated to be 54 mg/kg. Overall, considerable improvement in
efficacy was demonstrated with 26 relative to 2 (4Â by dose for
maximal effect), as reported previously,7 in line with what would
be anticipated with the greater antibacterial activity against the
pathogen.
In conclusion, this Letter has presented the efforts at optimiza-
tion of biochemical and cellular activity of a novel class of antibac-
terial agents, the pyrrolamides. Substitution on the piperidine
moiety resulted in improvements in potency, which translated into
in vivo activity in a lung model of infection. Further efforts to im-
prove potency and decrease the dose required for efficacy are
ongoing.
13. Hull, K.; Green, O.; Singh, A.; Bist, S.; Demeritt, J.; Loch,J.; Mullen, G.; Hauck, S.;
Sherer, B.; Ni, H.; Eakin, A. E., 48th Annual Meeting of the Interscience
Conference on Antimicrobial Agents and Chemotherapy, October 24–28, 2008,
F1-2027.
14. Inhibition of DNA gyrase by test compounds was measured using ammonium
molybdate/malachite green-based phosphate detection assays performed in
the presence of a range of concentrations of test compound. Assays of the S.
aureus GyrB ATPase utilized a hybrid tetramer enzyme reconstituted in vitro
from recombinant S. aureus GyrB and E. coli GyrA. The IC50 value is the
concentration of compound required for 50% inhibition of the gyrase ATP-
hydrolysis activity.
15. X-ray coordinates deposited in RSCB Protein Data Bank; PDB ID Code: 3TTZ.
16. Sherer, B.; Basarab, G.; Hull, K.; Hauck, S.; Bist, S.; Eakin, A. E. 49th Annual
Meeting of the Interscience Conference on Antimicrobial Agents and
Chemotherapy, San Francisco, CA, September 12–15, 2009, F1-1977.
1
7. The animals used for the mouse infection model were maintained in
accordance with the criteria for the American Association for Accreditation
of Laboratory Animal Care. The entire study was conducted using an IACUC-
approved protocol in accordance with Title
9 of the Code of Federal
Regulations. Groups of 10 CD1 mice (Charles River Laboratories, MA) were
infected intratracheally with 105 CFU/lung of S. pneumonia ARC548, whilst
under anesthesia with ketamine/xylazine.