Antitubercular Nitroimidazoles Revisited: Synthesis and Activity of the Authentic 3-Nitro Isomer of Pretomanid

Article abstract of DOI:10.1021/acsmedchemlett.7b00356

A published study of structural features associated with the aerobic and anaerobic activities of 4- and 5-nitroimidazoles had found that the 3-nitro isomer of pretomanid, 8, displayed interesting potencies, including against nitroreductase mutant Mycobacterium tuberculosis. However, recent nuclear magnetic resonance analyses of two trace byproducts, isolated from early process optimization studies toward a large-scale synthesis of pretomanid, raised structural assignment queries, particularly for 8, stimulating further investigation. Following our discovery that the reported compound was a 6-nitroimidazooxazole derivative, we developed a de novo synthesis of authentic 8 via nitration of the chiral des-nitro imidazooxazine alcohol 26 in trifluoroacetic or acetic anhydride, and verified its identity through an X-ray crystal structure. Unfortunately, 8 displayed no antitubercular activity (MICs > 128 μM), whereas the second byproduct (3′-methyl pretomanid) was eight-fold more potent than pretomanid in the aerobic assay. These findings further clarify target specificities for bicyclic nitroimidazoles, which may become important in the event of any future clinical resistance.

Full text of DOI:10.1021/acsmedchemlett.7b00356

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Letter
Antitubercular Nitroimidazoles Revisited: Synthesis and
Activity of the Authentic 3-Nitro Isomer of Pretomanid
Andrew M. Thompson, Muriel Bonnet, Ho Huat Lee, Scott G. Franzblau,
Baojie Wan, George S. Wong, Christopher B. Cooper, and William A. Denny
ACS Med. Chem. Lett., Just Accepted Manuscript • DOI: 10.1021/acsmedchemlett.7b00356 • Publication Date (Web): 13 Nov 2017
Downloaded from http://pubs.acs.org on November 16, 2017
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ACS Medicinal Chemistry Letters
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Antitubercular Nitroimidazoles Revisited: Synthesis and Activity of
the Authentic 3-Nitro Isomer of Pretomanid
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Andrew M. Thompson,*,† Muriel Bonnet,† Ho H. Lee,† Scott G. Franzblau,§ Baojie Wan,§ George S.
Wong,# Christopher B. Cooper,‡ and William A. Denny†
†Auckland Cancer Society Research Centre, School of Medical Sciences, The University of Auckland, Private Bag 92019,
Auckland 1142, New Zealand
§Institute for Tuberculosis Research, College of Pharmacy, University of Illinois at Chicago, 833 South Wood Street, Chicaꢀ
go, Illinois 60612, United States
#Summit CMC Alliance LLC, 61 Hawthorne Place, Summit, New Jersey 07901, United States
‡Global Alliance for TB Drug Development, 40 Wall St, New York 10005, United States
KEYWORDS: Tuberculosis, pretomanid, drug resistance, nitroimidazole, nitration, silyl migration
ABSTRACT: A published study of structural features associated with the aerobic and anaerobic activities of 4ꢀ and 5ꢀ
nitroimidazoles had found that the 3ꢀnitro isomer of pretomanid, 8, displayed interesting potencies, including against nitroreducꢀ
taseꢀmutant Mycobacterium tuberculosis. However, recent NMR analyses of two trace byproducts, isolated from early process opꢀ
timization studies toward a largeꢀscale synthesis of pretomanid, raised structural assignment queries, particularly for 8, stimulating
further investigation. Following our discovery that the reported compound was a 6ꢀnitroimidazooxazole derivative, we developed a
de novo synthesis of authentic 8 via nitration of the chiral desꢀnitro imidazooxazine alcohol 26 in trifluoroacetic or acetic anhyꢀ
dride, and verified its identity through an Xꢀray crystal structure. Unfortunately, 8 displayed no antitubercular activity (MICs >128
µM), whereas the second byproduct (3’ꢀmethyl pretomanid) was 8ꢀfold more potent than pretomanid in the aerobic assay. These
findings further clarify target specificities for bicyclic nitroimidazoles, which may become important in the event of any future clinꢀ
ical resistance.
Tuberculosis (TB) currently ranks alongside HIV/AIDS as
one of the leading causes of global mortality, claiming about
1.4 million lives every year.1 The spread of multiꢀ and extenꢀ
sively drugꢀresistant (MDR/XDR) TB is widely regarded as a
burgeoning health crisis, placing great strain on limited
healthcare resources for only modest treatment outcomes (cure
rates of 20ꢀ50% following complex therapy for up to 30
months with several more costly and toxic agents).1ꢀ3 The reꢀ
cent emergence of programmatically incurable tuberculosis
has led to many patients being discharged back into the comꢀ
munity, further threatening control efforts.4,5 In this context,
the conditional approval of two new MDRꢀTB drugs,
delamanid (OPCꢀ67683, 1; see Figure 1) and bedaquiline
(TMCꢀ207, 2), is a tremendous advance, although access to
these and repurposed agents such as linezolid (3) remains very
limited in many countries.4,6 To further address this urgent
Figure 1. Drugs included in novel regimens for MDR/XDRꢀTB
need, the TB Alliance has been developing shorter acting novꢀ
el regimens involving the nitroimidazooxazine pretomanid
(PAꢀ824, 4).7,8 Initial phase III clinical results for the combinaꢀ
tion of 2, 3, and 4 (NixꢀTB) in XDRꢀTB patients look highly
encouraging, with 29 of 31 patients who completed the treatꢀ
ment and 6 month followꢀup being cured.9 Another regimen
that combines 2 and 4 with pyrazinamide (5) and moxifloxacin
(6) is also demonstrating excellent bactericidal efficacy
against both drugꢀsensitive and MDRꢀTB.10
The mechanism of action of 4 involves bioreductive activaꢀ
tion by a deazaflavin (F420) dependent nitroreductase (Ddn),
leading to several metabolites arising from reduction of the
imidazole ring at Cꢀ3, including a desꢀnitro derivative.11 Forꢀ
mation of the latter correlates with the release of nitric oxide, a
crucial element in the anaerobic bactericidal activity of 4,
which is considered important for treatment shortening.12 In
contrast, the aerobic killing effects of 4 are attributed to the
inhibition of cell wall mycolic acid biosynthesis.13 The 5ꢀ
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nitroimidazole metronidazole also has some antitubercular THP deprotection
Page 2 of 6
and
installation
of
the
4ꢀ
1
2
3
4
activity under hypoxic conditions (vide infra) but a clinical
study in MDRꢀTB patients showed that it was too neurotoxic
for long term use.14
(trifluoromethoxy)benzyl ether, Kim et al. finally obtained 20
(instead of 8), equivalent to the 2S enantiomer of compound
10 in our nitroimidazooxazole paper,23 as verified by their
1
identical H and 13C NMR data. We note that while our comꢀ
In the past decade, several SAR studies of the nitroimidꢀ
azooxazine class have been reported by ourselves and othꢀ
ers,15,16 seeking more effective secondꢀgeneration analogues of
4. While the predominant focus has centered on optimising the
aryl side chain (e.g., clinical candidate TBAꢀ35417), a few
studies examined alternative linkages at Cꢀ618 or more fundaꢀ
mental modifications to the nitroheterobicyclic “warhead”.19,20
Here, replacement of the nitroimidazole portion by nitrotriaꢀ
zole or nitropyrazole abolished activity, whereas the 8ꢀoxygen
could be exchanged for sulfur or nitrogen, and substitution at
Cꢀ7 was tolerated.21 Notably, Kim et al.22 also described the
synthesis and biological evaluation of the 3ꢀnitro isomer of 4,
which was recorded as being “only slightly less active” (5ꢀ to
10ꢀfold) than 4 itself, whereas its precursor alcohol derivative
was 16ꢀ to 31ꢀfold less active than 4. Furthermore, the anaeroꢀ
bic killing of mutant Mycobacterium tuberculosis (M. tb) by
this isomeric compound was suggested to imply a different
biological target. This result could be of particular significance
in the event of future clinical resistance to 4. Nevertheless, in
the related nitroimidazooxazole class (cf. 1), we discovered
that relocating the nitro group from C6 to C5 reduced the aerꢀ
obic MIC against M. tb by at least 2ꢀ3 orders of magnitude.23
pound had been prepared by an independent and unequivocal
method, we have rigorously confirmed its structure by 2D
NMR, including a NOESY experiment, where an NOE effect
was observed between the imidazole proton resonance (Hꢀ5)
and resonances from the adjacent methylene protons at the 3ꢀ
position in the oxazole ring. This revised structure (20) for the
compound claimed by Kim et al. as 8 is also consistent with
the much smaller than expected optical rotation value obtained
([α]D20 +7.4 cf. ꢀ44.7 for 4 in MeOH25) as well as the moderate
antitubercular potency recorded (vide infra), as S enantiomers
are known to be an order of magnitude less effective than R
forms in the 2H nitroimidazooxazole class.23
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Figure 2. 2D NMR evidence for structure 16 over the reported 13
In an alternative approach to the 3ꢀnitro isomer of 4, we exꢀ
plored the novel nitration of known22 imidazooxazine alcohol
26 (Scheme 1B). This strategy was based on previous success
in our laboratory with the nitration of a related 5,6,7,8ꢀ
tetrahydroimidazo[1,2ꢀa]pyridinꢀ6ꢀol in acetic anhydride,19 as
well as literature precedence for the regioselective Cꢀ5 nitraꢀ
tion of both 1ꢀalkylꢀ2ꢀ(alkylthio)imidazoles26 and 2,3ꢀ
dihydroimidazo[2,1ꢀb][1,3]thiazole.27 During our initial synꢀ
thesis of 26, we again encountered a problem with migration
of the TBS group when glycidyl ether 11 was reacted with 2ꢀ
nitroimidazole (21).22 Fortunately, this was easily overcome
by switching the protecting group to the more stable TIPS
(22).24,25 We also made improvements to the subsequent THP
protection and deprotection steps by adopting methods from
Orita et al.,25 enabling the synthesis of alcohol 26 in 65% yield
over 4 steps. Surprisingly, attempted preparation of its acetate
ester (30) using acetyl chloride in pyridine gave only ring
opened Nꢀacetyl imidazolꢀ2ꢀone products, 27 and 28 (having a
During initial attempts to optimize a largeꢀscale synthesis of
4, two trace byproducts were isolated (<0.5%), and one of
these was postulated to be the 3ꢀnitro isomer, although its
NMR data did not match those provided by Kim et al.22 To
establish its identity, we sought to make this 3ꢀ
nitroimidazooxazine analogue of 4 via an unambiguous chiral
synthesis. Concurrently, we also targeted the second comꢀ
pound, thought to be a 3’ꢀmethyl derivative of 4. We now
report the intriguing findings from this study, including some
preliminary biological assessments.
To prepare the 3ꢀnitro isomer of 4, we first investigated the
method of Kim22 (Scheme 1A). Reaction of 2ꢀchloroꢀ4ꢀ
nitroimidazole (10) with the TBS ether derivative of Rꢀ
glycidol (11) gave two products in a ratio of ~3:1, with the
major one being the expected 4ꢀnitroimidazole derivative 12,
as described. However, after low temperature crystallization of
the more polar oily minor product (reportedly the 5ꢀ
1
nitroimidazole isomer of 12, i.e. 13), we found that its H
1
distinctive H NMR methyl resonance28 at δH 2.64 ppm); this
NMR spectrum in CDCl3 contained a D2Oꢀexchangeable hyꢀ
droxyl proton resonance at δH 1.83 ppm, which appeared as a
sharp “dd” (J = 6.7, 5.1 Hz), and coupled to the two proton
resonance of a methylene group in a COSY experiment.
Moreover, a NOESY experiment revealed a strong NOE effect
between the imidazole proton resonance and the proton resoꢀ
nances from the directly attached Nꢀmethylene. HMBC correꢀ
lations between Hꢀ5 and this NCH2 carbon and between the
NCH2 protons and both Cꢀ2 and Cꢀ5 (Figure 2) confirmed that
the correct structure of this minor product was the 4ꢀ
nitroimidazole 16. This result is in good accordance with the
known tendency of a TBS group to migrate under basic condiꢀ
tions, as employed here.24
was largely circumvented by employing acetic anhydride.
Several reagent systems were then explored for the nitration
of 26 or 30 (Table 1). Neither heating in aqueous nitric acid
nor treatment with acetyl nitrate in acetic anhydride gave the
desired product, while a reaction of 30 with nitronium tetraꢀ
fluoroborate was lowꢀyielding and accompanied by minor side
products. Overall, the best conditions were 70% nitric acid in
trifluoroacetic anhydride29 or a mixture of concentrated nitric
and sulfuric acids in acetic anhydride, which enabled yields of
~25ꢀ30% of the 3ꢀnitro derivatives, 7 and 33, respectively.
Intriguingly, only one nitro isomer was formed in trifluoroaceꢀ
tic anhydride, but this reaction proved to be more capricious
than the acetic anhydride alternative, where the isomer ratio
was ~9:1 in favour of nitration at Cꢀ3 over Cꢀ2 (for confirmaꢀ
tion, known25 2ꢀnitro acetate ester 32 was prepared from alcoꢀ
hol 3130). Isomers 32 and 33 were separable by careful column
chromatography and cleavage of acetate 33 to alcohol 7 was
cleanly achieved through the use of a mild base (NaHCO3).
A critical consequence of this structural assignment error is
that subsequent THP protection of the primary hydroxyl of 16
and treatment with TBAF (as reported) would induce cyclizaꢀ
tion to a 6-nitroimidazooxazole (18), rather than the 3ꢀ
nitroimidazooxazine 15 (Scheme 1A). Therefore, following
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Scheme 1. Synthesis of the target compounds 7-9, highlighting a key nitration stepa
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aReagents and conditions: (i) K2CO3, EtOH, 70 °C, 20 h (for 12/16), or 75ꢀ82 °C, 23 h (for 23); (ii) 3,4ꢀdihydroꢀ2Hꢀpyran, PPTS,
toluene, 20 °C, 5 d; (iii) TBAF, THF, 124 °C, 24 h (sealed tube); (iv) concd HCl (1.1 equiv), MeOH, 20 °C, 15 h; (v) AcCl, pyridine, 0ꢀ20
°C, 4 h; vi) Ac2O, pyridine, 0ꢀ20 °C, 7ꢀ19 h; (vii) TFAA, ꢀ5 to 0 °C, 15 min, then concd HNO3, ꢀ50 to 20 °C, 3 h, then ice, NaHCO3; (viii)
Ac2O, 0ꢀ20 °C, 5ꢀ80 min, then concd HNO3, concd H2SO4, ꢀ50 to 0 °C, 1.5ꢀ2.5 h, then ice, NaHCO3; (ix) NaHCO3, aq MeOH, 20 °C, 5 h;
(x) 4ꢀOCF3BnBr or 37, NaH, DMF, 0ꢀ20 °C, 2.5ꢀ3.3 h; (xi) nBuLi, THF, ꢀ78 °C, 1 h, then DMF, ꢀ78 to 20 °C, 1.5 h, and then aq citric
acid; (xii) NaBH4, MeOH, 0ꢀ20 °C, 1.5 h; (xiii) HBr, AcOH, 20 °C, 13 h.
Table 1. Summary of nitration methods explored for the synthesis of alcohol 7 or acetate 33
Compd
26
Solvent
H2O
Reagents (equiv)
3M HNO3 (9)
Temp range
80 to 97 °C
90 °C
Time/temp
3 h/97 °C
Products (% yield)
ꢀ
ꢀ
H2O
50% HNO3 (>100)
2 h/90 °C
26
26a
26a
26a
26a
26a
30
Ac2O
Ac2O
Ac2O
Ac2O
TFAA
CH3CN
Ac2O
70% HNO3 (3.1)b
ꢀ15 to 20 °C
ꢀ55 to 0 °C
ꢀ50 to 0 °C
ꢀ35 to ꢀ30 °C
ꢀ50 to 20 °C
ꢀ48 to 20 °C
ꢀ50 to 0 °C
16 h/20 °C
60 min/0 °C
75 min/0 °C
20 min/ꢀ30 °C
2ꢀ3 h/20 °C
90 min/20 °C
30 min/0 °C
ꢀ
96% H2SO4 (1.7), 100% HNO3 (3.0)
96% H2SO4 (1.7), 70% HNO3 (3.0)
96% H2SO4 (2.5), KNO3 (1.7)
70% HNO3 (2.5ꢀ2.8)
32 (<3), 33 (20)c
32 (<3), 33 (26)c
32 (<2), 33 (14)c
7 (12ꢀ35)
NO2BF4 (1.5)
30 (15), 32 (1), 33 (14)c
32 (<3), 33 (28)c
96% H2SO4 (1.9), 70% HNO3 (3.2)
30
aAcetylated or trifluoroacetylated in situ, prior to nitration. bPreformed acetyl nitrate. cYields after chromatography and crystallization.
Importantly, all of the successful nitration methods gave a
complete retention of stereochemistry (100% ee by chiral
HPLC analysis, using the analogously synthesized racemic
form of 7 as a reference standard). This indicated that the in
situ formed trifluoroacetate or acetate esters were sufficiently
stable to protect the chiral alcohol from potential racemisation,
e.g., through the formation of a chiral nitrate ester and subseꢀ
quent hydrolysis, as this can induce inversion of configuraꢀ
tion.31 Alkylation of pure 7 with 4ꢀ(trifluoromethoxy)benzyl
bromide (NaH/DMF) then gave the desired target 8 in excelꢀ
lent yield (95%) and, gratifyingly, this proved to be identical
(by NMR, mp, HPLC, and optical rotation) to the first byꢀ
product derived from optimization studies for a largeꢀscale
process route to 4. As expected from findings in the nitroimidꢀ
24
azooxazole class, the rotation value for 8 {[α]D ꢀ150.7 (c
1.002, CHCl3)} was indeed much larger than the one recorded
for 4.25 Conclusive structural proof was gained through a sinꢀ
gle crystal Xꢀray structure (Figure 3); of note, the benzyloxy
side chain adopted a pronounced pseudoaxial conformation at
C6, the same as that observed in the previously reported crysꢀ
tal structure of 4.21
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As expected from our studies in the nitroimidazooxazole
class,23 3ꢀnitro compounds 7 and 8 were completely inactive in
both M. tb assays (MICs >128 µM; Table 2). For 8, this indiꢀ
cates a >256ꢀfold loss in activity in comparison to its 2ꢀnitro
isomer, 4. Conversely, the 3’ꢀmethyl derivative of 4 (9) was 7ꢀ
to 8ꢀfold more active than both 4 and moxifloxacin (6) in
MABA (similar to rifampicin and bedaquiline 2) and ~3ꢀfold
better than both 4 and linezolid (3) in LORA (comparable to
rifampicin). These results for 9 were in accordance with the
findings reported by Cherian et al.16 for the 3’ꢀmethoxy derivaꢀ
tive of 4, which was 5ꢀfold superior to 4 in the aerobic assay
and 2ꢀfold more effective than 4 under hypoxic conditions.
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Figure 3. Xꢀray crystal structure of compound 8
In their original investigation, Kim et al.22 reported an aeroꢀ
bic MIC99 value of 4ꢀ8 µM and weak anaerobic activity
(MIC90 31 µM) for the compound they believed to be 8
(shown here to be the 2H nitroimidazooxazole 20). Intriguingꢀ
ly, the same compound also yielded an anaerobic MIC90 value
of 62.5ꢀ125 µM against M. tb having mutations in the nitroreꢀ
ductase Ddn, suggesting the participation of a different biologꢀ
ical target. While delamanid (1) was shown to be primarily
triggered by Ddn, several Ddn homologues have been impliꢀ
cated in the cellular activation of simple 2H nitroimidazooxaꢀ
zoles (e.g., 2ꢀEt, 2ꢀPh).34 However, in the case of 20, the lack
of significant potency against this Ddn mutant under aerobic
conditions (MIC99 >100 µM) renders this explanation unsatisꢀ
factory. Overall, the results of the current investigation support
findings from previous studies of resistance to 4 that the actiꢀ
vation of nitroimidazooxazines relies exclusively on Ddn.35 In
this class, relocation of the nitro group from Cꢀ2 to Cꢀ3 deꢀ
stroys all antitubercular activity, implying that, like the Rꢀ
enantiomer of 4,34 8 is not a substrate for Ddn or its homoꢀ
logues and does not release nitric oxide.
Preparation of the second byproduct from the synthesis of 4
was straightforward (Scheme 1C). Lithiation of bromotoluene
34 and quenching with DMF gave the required aldehyde 35,
which was easily reduced (NaBH4) and brominated to give 37.
Alkylation of chiral alcohol 3130 with bromide 37 then providꢀ
ed the expected compound 9, which was also identical to that
derived from the nonꢀoptimized process chemistry route for 4.
This byproduct (<0.2%) was postulated to arise from traces of
37 in the bulk commercial 4ꢀ(trifluoromethoxy)benzyl broꢀ
mide. Full experimental procedures and characterization data
for all compounds have been provided in the Supporting Inꢀ
formation.
The comparative effects of 7ꢀ9 and several reference drugs
against M. tb (strain H37Rv) were assessed in two in vitro
assays, MABA32 and LORA,33 conducted under aerobic and
hypoxic conditions, respectively. The latter assay employed
bacteria preadapted to low oxygen conditions and represented
a targeted screen for the identification of agents with better
sterilizing ability against nonꢀreplicating persistent bacteria.
Recorded minimum inhibitory concentrations (MICs) correꢀ
sponding to growth inhibitions of ≥90% (Table 2) were mean
values obtained from replicate measurements (± standard deꢀ
viation). Compounds 7ꢀ9 were also evaluated for cytotoxicity
against mammalian cells (VERO) in a 72 h assay32 and found
to be nonꢀtoxic (IC50 >128 µM).
In summary, we set out to establish the identities of two
novel byproducts from optimization studies around a manufacꢀ
turing route to 4 through a combination of 2D NMR analysis
and de novo chiral synthesis. In the case of 3ꢀnitro isomer 8,
this entailed the development of an innovative nitration route,
following our discovery of a critical structural assignment
error in the published method, and we obtained an Xꢀray crysꢀ
tal structure of this compound for final confirmation. Prelimiꢀ
nary in vitro assessments indicated that whereas the 3’ꢀmethyl
derivative of 4 (9) was markedly more effective than 4, both 8
and its alcohol precursor 7 were completely inactive, overturnꢀ
ing previous misconceptions regarding their aerobic and anꢀ
aerobic activities and the suggested involvement of another
target facilitating their activity against Ddn mutant M. tb. Theꢀ
se results provide further clarity of fundamental SAR for
pretomanid and of the structural features of antitubercular
nitroimidazoles that are more likely to overcome any future
clinical resistance to 4.
Table 2. In vitro activities of 7-9 versus other TB drugs
MICa (µM)
MABA
IC50b (µM)
VERO
>10
Compd
LORA
0.11 ± 0.03
2.8 ± 0.3
2.6 ± 1.4
>128
2
0.070 ± 0.018
2.9 ± 1.2
3
4c
>128
0.50 ± 0.30
0.42 ± 0.13
>128
6
>128
>128
>128
>128
7
>128
>128
8
ASSOCIATED CONTENT
Supporting Information
9
0.063 ± 0.003
>512
1.0 ± 0.1
79 ± 40
0.64 ± 0.35
>128
MET
RMP
INH
The Supporting Information is available free of charge on the
ACS Publications website.
>100
0.049 ± 0.027
0.34 ± 0.18
aMinimum inhibitory concentration against M. tb, determined
under aerobic (MABA)32 or hypoxic (LORA)33 conditions. Each
value is the mean of ≥2 independent determinations (7ꢀ9 were
Further background, scheme for preparation of racemic 7, experꢀ
imental procedures and characterizations for compounds, combusꢀ
tion analytical data, packing diagram for Xꢀray structure of 8,
crystallographic data, NMR spectra for key compounds, chiral
HPLC trace for 7 (PDF)
tested
3 times). The controls were metronidazole (MET),
b
rifampicin (RMP), and isoniazid (INH). IC50 values for cytotoxiꢀ
city toward VERO cells. cMIC data from ref. 15.
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(10) Dawson, R.; Harris, K.; Conradie, A.; Burger, D.; Murray, S.;
AUTHOR INFORMATION
Mendel, C.; Spigelman, M. Efficacy of bedaquiline, pretomanid,
moxifloxacin & PZA (BPAMZ) against DSꢀ & MDRꢀTB. Abstracts
of Papers, Conference on Retroviruses and Opportunistic Infections
(CROI 2017), Seattle, Washington, February 13ꢀ16, 2017; 724LB.
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Corresponding Author
*Phone: (+649) 923 6145. Fax: (+649) 373 7502. Email:
am.thompson@auckland.ac.nz.
See:
http://www.croiconference.org/sessions/efficacyꢀbedaquilineꢀ
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pretomanidꢀmoxifloxacinꢀpzaꢀbpamzꢀagainstꢀdsꢀmdrꢀtb (accessed 30
April, 2017).
Notes
The authors declare no competing financial interest.
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J.; Zia, N.; Lee, M.; Lee, J.; Via, L. E.; Lee, S.; Eum, S.ꢀY.; Lee, S.ꢀJ.;
Goldfeder, L. C.; Cai, Y.; Jin, B.; Kim, Y.; Oh, T.; Chen, R. Y.;
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Franzblau, S. G.; Wan, B.; Wang, Y.; Ma, Z.; Denny, W. A.; Thompꢀ
son, A. M. Synthesis and structureꢀactivity relationships for extended
side chain analogues of the antitubercular drug (6S)ꢀ2ꢀnitroꢀ6ꢀ{[4ꢀ
(trifluoromethoxy)benzyl]oxy}ꢀ6,7ꢀdihydroꢀ5Hꢀimidazo[2,1ꢀ
b][1,3]oxazine (PAꢀ824). J. Med. Chem. 2015, 58, 3036ꢀ3059.
(16) Cherian, J.; Choi, I.; Nayyar, A.; Manjunatha, U. H.; Mukherꢀ
jee, T.; Lee, Y. S.; Boshoff, H. I.; Singh, R.; Ha, Y. H.; Goodwin, M.;
Lakshminarayana, S. B.; Niyomrattanakit, P.; Jiricek, J.; Ravindran,
S.; Dick, T.; Keller, T. H.; Dartois, V.; Barry, C. E., III. Structureꢀ
activity relationships of antitubercular nitroimidazoles. 3. Exploration
of the linker and lipophilic tail of ((S)ꢀ2ꢀnitroꢀ6,7ꢀdihydroꢀ5Hꢀ
imidazo[2,1ꢀb][1,3]oxazinꢀ6ꢀyl)ꢀ(4ꢀtrifluoromethoxybenzyl)amine (6ꢀ
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ACKNOWLEDGMENT
9
The authors thank the Global Alliance for Tuberculosis Drug
Development (TB Alliance) for financial support through a colꢀ
laborative research agreement. The TB Alliance gratefully
acknowledges funding from the Bill & Melinda Gates Foundation
(Investment ID: OPP1129600).
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ABBREVIATIONS
TB, tuberculosis; M. tb, Mycobacterium tuberculosis; MDR, mulꢀ
tidrugꢀresistant; XDR, extensively drugꢀresistant; Ddn, deazaflaꢀ
vinꢀdependent nitroreductase
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