2-Aminooxazole as a Novel Privileged Scaffold in Antitubercular Medicinal Chemistry

Article abstract of DOI:10.1021/acsmedchemlett.0c00173

To obtain effective eradication of numerous infectious diseases such as tuberculosis, it is important to supply the medicinal chemistry arsenal with novel chemical agents. Isosterism and bioisosterism are widely known concepts in the field of early drug d

Full text of DOI:10.1021/acsmedchemlett.0c00173

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Letter
2-Aminooxazole as a Novel Privileged Scaffold
in Antitubercular Medicinal Chemistry
Elisa Azzali, Miriam Girardini, Giannamaria Annunziato, Marialaura Pavone, Federica
Vacondio, Giorgia Mori, Maria Rosalia PASCA, Gabriele Costantino, and Marco Pieroni
ACS Med. Chem. Lett., Just Accepted Manuscript • DOI: 10.1021/
acsmedchemlett.0c00173 • Publication Date (Web): 08 Jun 2020
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Table of contents
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2-Aminooxazole as a Novel Privileged Scaffold in Antitubercular Medicinal Chemistry
Elisa Azzali,a,b,¥ Miriam Girardini,a,b Giannamaria Annunziato,a,b Marialaura Pavone,a,b Federica
Vacondio,b,d Giorgia Mori,c Maria Rosalia Pasca,c Gabriele Costantino,a,b,d,e Marco Pieroni,a,b,d*
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aP4T group, and Department of Food and Drug, University of Parma, 43124 Parma, Italy;
Department of Biology and Biotechnology “Lazzaro Spallanzani”, University of Pavia, 27100 Pavia;
dCentro Interdipartimentale “Biopharmanet-tec”, Università degli Studi di Parma, Parma, Italy;
eCentro Interdipartimentale Misure (CIM) ’G. Casnati’, University of Parma, Parma, Italy.
¥
Present Address: Aptuit (Verona) Srl, an Evotec Company, Via Alessandro Fleming, 4, 37135
Verona, Italy
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Abstract
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To obtain effective eradication of numerous infectious diseases such as tuberculosis, it is important to supply
the medicinal chemistry arsenal with novel chemical agents. Isosterism and bioisosterism are widely known
concepts in the field of early drug discovery, and, in several cases, rational isosteric replacements have
contributed to improve efficacy and physicochemical characteristics throughout the hit-to-lead optimization
process. However, sometimes the synthesis of isosteres might not be as straightforward as that of the parent
compounds, therefore novel synthetic strategies must be elaborated. In this regard, we herein report the
evaluation of a series of N-substituted 4-phenyl-2-aminooxazoles that, despite being isosteres of a widely used
nucleus such as the 2-aminothiazole, have been only seldom explored. After elaborating a convenient synthetic
strategy, a small set of 2-aminothiazoles and the 2-aminooxazole counterparts were compared regarding their
antitubercular activity and physicochemical characteristics.
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Keywords
2-aminooxazole; 2-aminothiazoles; bioisosterism; Buchwald-Hartwig coupling; isosterism
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Introduction
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Chemotherapy is nowadays the only means to eradicate those bacterial infections for which an effective
vaccine is not available. However, the emergence of antimicrobial resistance has significantly narrowed the
scope of a number of chemotherapeutics, making them obsolete and sometimes devoid of efficacy.1,2 In this
regard, the exploitation of novel chemical tools to support the drug discovery process is highly desirable. This
applies to tuberculosis (TB), which causes ill-health among millions of people each year and ranks alongside
the HIV as the leading cause of death worldwide.3–5 Specific medicinal chemistry efforts undertaken over a
number of years have led to a nourished pipeline of novel chemical antitubercular entities, which are now
either in the clinical or in the preclinical/hit-to-lead optimization phases.6 With the aim to further enrich the
antitubercular arsenal, in the recent years we have developed a series of 2-aminothiazole derivatives endowed
with strong antitubercular activity coupled to irrelevant cytotoxicity toward eukaryotic cells (compounds 1–4,
Figure 1), high selectivity toward other microorganisms, and little tendency to be substrate of Mtb efflux
pumps.7–11
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[Insert Figure 1 here]
Encouraged by these studies, and with the aim of further improving the antitubercular efficacy, we deemed of
interest to define a reliable set of Structure-Activity Relationships (SAR) for this series of molecules. First, we
wanted to investigate whether the 2-aminothiazole moiety, supposed to be the pharmacophore for molecules
1–4, could be successfully replaced. The sulfur atom of the 2-aminothiazole can be easily oxidized and, as a
result, the molecule metabolically inactivated. In addition, 2-aminothiazoles, in some cases, have been referred
to as Pan-Assay Interference Compounds (PAINS),12–14 therefore the evaluation of different structural motifs
might be worthwhile. The most obvious substitution of the 2-aminothiazole moiety is represented by the 2-
aminooxazole,15,16 where the sulfur atom of the thiazole ring is substituted by oxygen, a sulfur isostere.16 In
this regard, Hinsberg has extended the concept of isosterism and bioisosterism to heterocycles according to the
“ring equivalence” theory, on the basis of which benzene, thiophene, pyridine and furan can be considered ring
equivalents. Moreover, the 2-aminooxazoles could possess some advantages over 2-aminothiazoles such as a
decreased ClogP, and thus improved solubility, and a slower metabolism rate since the lack of the oxidizable
sulfur atom. These findings prompted us to plan the synthesis of a series of substituted 2-aminooxazoles to
evaluate their biological activity as compared with the corresponding 2-aminothiazoles 1–4. Despite the
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synthesis of the oxazole core has been addressed by a number of groups,17–19 however the preparation of
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substituted 2-aminooxazoles has been only marginally considered. Surprisingly, the Hantzsch protocol,20
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highly versatile method for the straightforward preparation of substituted 2-aminothiazoles starting from the
suitable α-bromoketones and N-substituted thioureas, did not allow us to obtain 2-aminooxazoles when N-
substituted ureas were used. Even more surprising was the fact that procedures to prepare 2-aminooxazole
analogues presenting the same substituent pattern as our antitubercular 2-aminothiazoles (i.e. an aromatic ring
at position C-4 and one at the nitrogen atom, Figure 1) are scarcely reported in the scientific literature. For
instance, the synthesis of a structure as simple as that of N,4-diphenyl-2-aminooxazole 37 (Figure 2) is rarely
reported,21–24 and in all cases the synthetic procedures suffer from narrow feasibility and poor yields. In one
case, upon reproduction of the conditions reported,22 we failed to obtain the desired compound.
[Insert Figure 2 here]
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The poor reactivity of N-substituted ureas might be due to the lower nucleophilicity of the oxygen atom
compared to the sulfur, however, this does not seem to apply to unsubstituted urea, that is reported to plainly
react with different α-bromoacetophenones to give N-unsubstituted 2-aminooxazoles. Given these findings,
the development of a new and versatile synthetic strategy to efficiently obtain N-substituted 4-aryl-2-
aminooxazoles might be of interest for the medicinal chemistry community.
Results and discussion
Considering what is reported above, we evaluated the exploitation of an optimized two-step method based on:
a) the condensation of the proper α-bromoacetophenones with urea and b) the Buchwald-Hartwig cross
coupling of the 2-aminooxazole with an aryl halide. Finally, the versatility of the method was evaluated
through the use of variously substituted α-bromoacetophenones and aryl halides. In this initial phase, α-bromo-
4'-methylacetophenone 5 was used as the starting building block to react with urea 6 (Table 1). This reaction
is reported in six studies,21,25–29 and the yields for the formation of 7 is reported in only one case (21%).25 As
for the first step, we focused our interest in the optimization of parameters such as the solvent, stoichiometric
ratio between α-bromoacetophenone and urea, and whether microwave (MW) irradiation was more efficient
than conventional heating. Regarding the solvents, first attempts were made using acetonitrile, ethanol,
dimethoxyethane and PEG-400, as already reported in the literature for this kind of reaction. Reactions were
performed overnight at 80°C, with two different stoichiometric ratios of urea that is 1:2 and 1:10 (Table 1).
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Reaction progress was evaluated by TLC, and when no significant change could be observed, either by
persistence of the starting material or lack of novel spots, the reaction was stopped, and the starting material
recovered. Unfortunately, it was not possible to obtain compound 7 from any of the reported conditions (Table
1, entries I-VI). We reasoned that temperature might have played a role in promoting the condensation
reaction, therefore high-boiling aprotic polar solvents such as dimethylformamide (DMF) and
dimethylsulfoxide (DMSO) were investigated to carry out the reaction. Using DMF as the solvent, we were
pleased to notice that no more starting material could be detected in the TLC after 3 hours, despite the
temperature was kept at 80°C (Table 1, entry IX, yield 37%).
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[Insert Table 1 here]
In the same conditions, but using DMSO as the solvent, the reaction failed to show any conversion of the
starting material in the desired compound (Table 1, entry VII). As such, we focused on the use of DMF as the
solvent for the condensation reaction. If the stoichiometric amount of urea used is reduced (from 1:10 to 1:2),
a longer reaction time, coupled to a decrease in the yield (18%), was observed upon complete consumption of
the starting material (Table 1, entry VIII). Conversely, raising the temperature up to 120°C led to a higher
yield (45%) and a reduction of the reaction time, which resulted in 30 minutes (Table 1, entry X). We reasoned
that the reaction time could be shortened even further by using a microwave reactor, as it is known to reduce
the reaction time due to the higher pressure reached in the reaction vessel. Indeed, we were pleased to note
that, at 1:10 stoichiometric ratio, the desired 4-(p-tolyl)oxazol-2-amine 7 could be obtained in fairly similar
yields either reacting at 80°C for 15 minutes or 3 minutes at 120°C (table 1, entries XI and XII, yields 53%
and 56%, respectively). Using another high-boiling aprotic polar solvent such as N-methyl pyrrolidone (NMP)
at the same conditions, led to similar results (Table 1, entries XIII and XIV, yields 50% and 45%, respectively),
although the purification of the reaction mixtures with NMP are notoriously more problematic than those in
DMF. This led us to consider the use of DMF as the most convenient conditions under which to carry out the
condensation reaction, with 1:10 stoichiometric ratio of urea, in a microwave reactor for 3 minutes and 120°C
(Table 1, entry XII).
The Buchwald-Hartwig reaction is a cross-coupling reaction of an aryl halide with an amine using a palladium
source as the catalyst and a strong base. We have taken advantage from this well-established synthetic protocol
especially in view of the fact that this procedure has been already applied to many amino-heterocycles,
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including 2-aminooxazoles,30,31 although this is the first time that this reaction is used systematically to prepare
4-aryl substituted 2-aminooxazoles. On the other hand, in the case of 2-aminothiazoles, one palladium-
catalysed reaction has been described making use of biaryl-phosphorinanes.32,33 We have investigated 3
different catalytic systems to obtain the title compound 9 from 4-(p-tolyl)oxazol-2-amine 7 and 4-
bromobenzene 8 (Table 2): X-Phos Pd G2, S-Phos Pd G2, which are second-generation Buchwald precatalysts,
with Pd and ligand linked together, and DavePhos/Pd(OAc)2. Each of them was used in combination with 4
bases, sodium tert-butoxide (tBuONa) , Cs2CO3, K2CO3 and K3PO4, and the reactions were carried out in
toluene in a microwave reactor at 130 °C for 10 minutes, since microwave irradiation has already proven its
effectiveness in high-temperature reactions. The catalytic system constituted from DavePhos/Pd(OAc)2, in
general, gave the worst yields (Table 2, entries IX-XII, yields 8-11%), regardless the base used. In the other
cases, conversion of the 4-(p-tolyl)oxazol-2-amine 7 in the desired adduct 9 was appreciable, except when
K2CO3 was used as the base (Table 2, entries III and VII).
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[Insert Table 2 here]
This could be due to the fact that K2CO3, as the base, is too weak to allow the reaction to proceed. However,
it must be noticed that in the case of DavePhos/Pd(OAc)2, K2CO3 allowed for formation of the desired final
compound, although in low yields (Table 2, entry XI, yield 11%). Using X-Phos Pd G2 and S-Phos Pd G2 as
the catalysts, tBuONa gave the best results (Table 2, entries I and V), allowing for formation of the desired
coupling product in 50% and 49% yield, respectively. The use of bases such as Cs2CO3 and K3PO4 (Table 2,
entries II, IV, VI, VIII)allowed for formation of the desired compound in moderate yields (20-42%), although
the use of X-Phos Pd G2 provided a better overall yield compared to S-Phos Pd G2 (Table 2, entries II and IV
vs VI and VIII), whereas in the case of tBuONa the difference is negligible.
The final step was represented by the assessment of the versatility of the approach. It was evaluated how the
BH coupling reaction was affected by the different substitution pattern of the bromobenzene, that was adorned
with electron-donor groups (EDGs) and electron-withdrawing groups (EWGs) at different positions of the
aromatic ring, that is the ortho, the meta and the para position (Table 3, reaction B). In this preliminary
investigation, the methoxy group was chosen as representative of the EDGs, whereas the electronegative
fluorine atom was the representative of the EWGs. All of the reactions were carried out in a microwave reactor
using X-Phos Pd G2 as the catalyst and tBuONa as the base, in toluene at 130 °C until consumption of the
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limiting reagent, that usually occurred in 10 minutes, as shown by TLC. First, it must be noticed that, in most
cases, a substituted bromobenzene was more reactive than the unsubstituted counterpart. Although it can’t be
a general rule of thumb, in our case an EDG such as the methoxy group resulted in better yields (27–29, yields
48–71%) than an EWG such as the fluorine (24–26, yields 37–59%). In both cases, the ortho substitution led
to the best results, whereas at this point it would be too speculative to establish whether the meta or the para
position are detrimental for the yields (Table 3). In a similar way, we wanted to evaluate whether the
condensation with urea, using the conditions described above, could be applied to several substituted
acetophenones (10–16). For this preliminary evaluation, we used acetophenones substituted at the ortho, meta
and para position with EDGs and EWGs. As expected, the effect of the substituent is negligible (Table 3), and
the yield for this reaction is always around 50%. Once the versatility of the procedure was demonstrated, we
applied it to the synthesis of 2-aminooxazoles 30, 31, 34 and 36, which are structurally related to the 2-
aminothiazoles 1–4,7,10 respectively. These derivatives were successfully synthesized following the procedure
above reported, that further confirmed its versatility. It should also be noted that, also in these cases, the
standard Hantzsch protocol with substituted ureas did not lead to the title compounds, despite performing the
reaction in different conditions (data not shown).
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The final step of this research was both to evaluate the actual isosterism and bioisosterism of the 2-
aminooxazoles compared to the corresponding 2-aminothiazole. First, the synthesized compounds were
evaluated for their effect toward Mtb, and we were pleased to note that compounds 30, 34, 36, bearing a 2-
aminooxazole moiety, showed promising activity toward Mtb, with a trend similar to that of 2-aminothiazoles
1–4. Indeed, as for the case of 2-aminothiazoles, it is possible to appreciate that isoxazole derivatives 34 and
36 were generally more active than the aryl analogues, with negligible cytotoxicity toward VERO cells (data
not shown).
[Insert Table 3 here]
Afterward, as the final proof of the isosterism of the novel molecules, we evaluated other physicochemical and
in vitro ADME properties, such as (i) kinetic solubility in water and Phosphate Buffered Saline (PBS) pH 7.4,
(ii) metabolic stability in human liver microsomes (HLM), and (iii) reactivity toward glutathione (GSH). In
particular, the latter was considered important in order to evaluate the tendency of these compounds to be
PAINS.
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[Insert Table 4 here]
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Kinetic solubility measurements
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The kinetic solubility of selected 2-aminothiazoles was assessed starting from DMSO stock solutions of 1-4,
compared to that of corresponding isosteres, the 2-aminooxazoles 30, 31, 34, 36. This approach was preferred
to the measurement of equilibrium solubility starting from powder, because the latter is significantly influenced
by the nature of the solid state (amorphous vs. crystalline, polymorphic forms), which, generally, is not
thoroughly investigated at the discovery stage.34 In Table 4, the values of kinetic solubility, in water and in
PBS buffer pH 7.4, expressed as µM, are reported. In general, kinetic solubility values in water ranged between
0.7 µM (Log S = –6.15) of the least soluble 31 to 55 µM (Log S = –4.26) of the most soluble 3, spanning two
orders of magnitude, while in PBS buffer at physiological pH they were between 1.1 µM (Log S = –5.95) for
31 and 27.7 µM (Log S = –4.56) for 3. The two scales of log S values in water and PBS were positively
correlated (Log Swat = 1.40(0.18) Log SPBS + 2.26(0.96); n = 7; r2 = 0.921; s = 0.19; F = 59), with the sole
exclusion of 2-aminothiazole 2 for which solubility value in PBS was – 1.2 log unit lower than that in water.
Comparing the 2-aminothiazole/2-aminooxazole isosteric couples 1-31, 2-30, 3-34, 4-36, we did not observe
any significant difference between the average solubility values measured either in water [average LogS (2-
aminothiazoles) = –4.78 ( 0.54); mean  SD (n=4); average LogS (2-aminooxazoles) = –5.27 ( 0.54); mean
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 SD (n=4); p>0.05] or in PBS buffer [mean LogS (2-aminothiazoles) = – 5.30 ( 0.59); mean LogS (2-
aminooxazoles) = –5.34 ( 0.41); p>0.05]. Compounds which showed the two best ranking in kinetic solubility
values were: the already cited 2-aminothiazole 3 (55 µM equal to 18 g/mL in water; 27 µM equal to 9 g/mL
in PBS) and the 2-aminooxazole 36 (24 µM corresponding to 8.7 g/mL in water; 9.3 µM equal to 3.4 g/mL
in PBS).
Metabolic stability in human liver microsomes
We next determined the metabolic stability of the set of 2-aminothiazoles and 2-aminooxazoles in human liver
microsomes (HLM). Although liver microsomes are the simplest in vitro model for phase I metabolism, a first
assessment of metabolic stability by this subcellular model is useful since compounds presenting short in vitro
half-lives (t1/2) in this assay are those most likely to encounter later in vivo pharmacokinetic issues (i.e. high in
vivo clearance and, as a consequence, reduced oral bioavailability).35 An “intrinsic clearance” (Cl’int) can also
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be calculated based on the in vitro microsomal stability data (Table 4). As it can be seen from Table 4, two
compounds of the set (i.e. the two ethyl esters 3 and 34) were characterized by the shortest half-lives in HLM
(i.e. t1/2 = 3.9 min for 3; 2.7 min for 34), due to their rapid cleavage to the corresponding acids. Indeed, the
corresponding acids showed a reduced clearance in comparison to esters (Cl’int = 160.7 ml/min/kg for ester 3
and 226.1 ml/min/kg for 34 if compared to a Cl’int  371 ml/min/kg for both the corresponding acids). For
all the remaining compounds, we could not see a clear dependence of metabolic stability on the substitution of
the core 2-aminothiazole ring with the isostere 2-aminooxazole. Indeed, even if the 2-aminooxazole 30 had a
half-life significantly longer than that of the corresponding 2-aminothiazole 2 (t1/2 = 22.6  1.2 min vs. 14.5 
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0.5 min; mean  SD; n=3; p<0.01), for the other two couples of isosteres 31-1 and 36-4 half-lives were
comparable, suggesting that the preferred metabolic pathways in HLM of compounds 1 and 4 did not involve
the sulfur atom of the thiazole core ring.
Reactivity of selected 2-aminothiazoles and 2-aminooxazoles with GSH
Compounds defined as PAINS generally give false positive results in HTS assays as they tend to react non-
specifically. Thus, the investigation of the reactivity of the set of 2-aminothiazoles and, in parallel, of
corresponding 2-aminooxazoles was carried out. Test compounds were incubated with GSH, the most
abundant thiol in cells, for 24 h before analysing the formation of the GSH-conjugates by HPLC-MS/MS. At
full scan MS analysis only the molecular ion [M+H]+ was observed for all the compounds tested, while for the
GSH-conjugates the presence of both the singly charged ([M+H]+ conjugate) either the doubly charged ion
([M+2H]2+ conjugate) were detected (e.g. Supplementary Fig. S1). The percentage ratios of the GSH conjugate
formed, was thus calculated as: (Peak Area of [M+H]+ conjugate + Peak Area ([M+2H]2+ conjugate)/ Peak
Area ([M+H]+ compound) and reported in Table 4. In general, at 24 h, despite the fact that GSH had been
employed in a large molar excess (1:100), only a small percentage of the starting compounds reacted with
GSH. In fact, percentages of compound-GSH conjugates ranged between a 2.1% for 36 to 16% for 34. Given
the overall poor reactivity of these compounds towards GSH, it could be deduced that they do not act like
PAINS and thus their activity can be considered the result of a specific interaction with a molecular target.
In conclusion, we have investigated the possibility of exploiting the 2-aminooxazole scaffold as a novel
privileged structure in medicinal chemistry. Whereas the 2-aminothiazole is widely used in many bioactive
molecules, either in the early drug-discovery phases or in the market, the 2-aminooxazole, an isostere of 2-
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aminothiazole, has been poorly investigated. We have first set up a synthetic protocol for the preparation of N-
4-disubstituted 2-aminooxazoles, and then we have evaluated a small set of 2-aminooxazoles and
corresponding 2-aminothiazoles with regard to their biological and pharmacokinetic characteristics. We
noticed that the antitubercular activity is maintained in the 2-aminooxazole derivatives, with a trend similar to
that of the 2-aminothiazoles, confirming how the two nuclei can be considered bioisosteres. Moreover, the 2-
aminooxazole structure does not negatively affect other physico-chemical or in vitro PK parameters such as
solubility, metabolic stability, and unselective reactivity with thiols. A larger set of molecules and assays are
needed to confirm this and are currently under investigation in our laboratories.
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Supporting Information The Supporting Information is available free of charge athttps://pubs.acs.org/.
Description of the chemical and biological methods, evaluation of MICs, and the analytical data related to
the compounds synthesized are reported in the supporting information.
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Author information marco.pieroni@unipr.it
Author Addresses Food and Drug Department, University of Parma, Parco Area delle Scienze 27, 43124,
Parma
Author Contributions EA, GA, MG, MLP: synthesis, data collection, data analysis; MRP, GM: MIC
determination, manuscript revision; FV: Study on solubility and microsomal stability, manuscript revision;
GC: resources, supervision, manuscript revision. MP: study design, supervision, manuscript writing and
revision;
Acknowledgments: The “Centro Interdipartimentale Misure “G. Casnati” is kindly acknowledged for the
contribution in the analytical determination of the molecules synthesized. This research was also supported by
the Italian Ministry of Education, University and Research (MIUR): Dipartimenti di Eccellenza Program
(2018–2022) - Dept. of Biology and Biotechnology "L. Spallanzani", University of Pavia (MRP, GM).
Abbreviations ADME, absorption, distribution, metabolism and excretion; DMF, dimethylformamide;
DMSO, dimethylsulfoxide; EDC, N-(3-Dimethylaminopropyl)-N′-ethylcarbodiimide; EDG, electron-donor
group; EWG, electron-withdrawing group; HLM, human liver microsomes; GSH, glutathione; MDR,
multidrug resistant; Mtb, Mycobacterium tuberculosis; MW, microwave; PAINS, pan-assay interfering
compounds; SAR, Structure-Activity Relationships; TB, tuberculosis; TBTU, O-(Benzotriazol-1-yl)-
N,N,N′,N′-tetramethyluronium tetrafluoroborate; TDR, totally drug resistant; TEA, triethylamine; THF,
tetrahydrofuran; TLC, thin-layer chromatography; XDR, extensively drug resistant.
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Figure 1: 2-aminothiazoles with anti-TB activity.7,10
5
6
7
8
S
S
F
NH
NH
N
N
9
O
10
11
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CF3
1
2
O
S
O
S
N
H
O
N
N
N
N
N
N
O
H
H
N
O
3
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Figure 2: attempts to synthesize N,4-diphenyl-2-aminooxazole21–24
Deep eutectic
O
solvent, Ultrasound, rt
O
NH
H
PEG-400, rt, 3h
N
H2N
N
+
O
Br
Glycerol, rt, 1h
Ethanol, 70°C
37
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Scheme 1a,b
4
5
6
7
8
O
O
O
O
R
b
a
NH2
Br
+
N
N
R
N
H2N
NH2
R
H
9
10
11
12
13
14
15
16
17
18
19
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5, 10 - 16
6
7, 17 - 23
24 - 31
aReagent and conditions: a) DMF, MW (120C, 300W), 3 min, 25-38%; b) 4-bromotoluene, tBuONa, X-Phos
Pd G2, tBuOH, toluene, MW (130C, 300W), 15 min, 10-31%. bFor complete structures see Table 3.
Scheme 2a
O
O
O
a
b
O
O
O
O
O
O
N
N
O
N
N
O
N
N
O
Br
NH2
H
32
33
34
c
O
O
d
HO
N
O
N
H
O
N
N
O
N
O
N
N
N
H
H
36
35
aReagent and conditions: a) urea, DMF, MW (120C, 300W), 3 min, 49%; b) 4-bromotoluene, tBuONa, X-
Phos Pd G2, t-BuOH, toluene, MW (130C, 300W), 15 min, 16%; c) LiOH, THF/H2O/MeOH 3:1:1, rt, 3 h,
quantitative; d) 2-aminopyridine, TBTU, EDC, TEA, anhydrous DMF, rt, 5 h, 57%.
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Table 1: Optimization of the condensation reaction.
O
5
6
7
8
O
O
NH2
Br
+
N
H2N
NH2
9
5
6
7
10
11
12
13
14
15
16
17
18
19
20
21
22
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Entry
I
II
III
IV
V
VI
VII
VIII
IX
X
XI
XII
XIII
XIV
Solvent
EtOH
EtOH
PEG400
CH3CN
CH3CN
DME
DMSO
DMF
DMF
M/Ma
1/2
1/10
1/2
Time
18h
18h
18h
18h
18h
18h
3h
8h
Temp
Yield (%)
80°C
80°C
20°C
80°C
80°C
80°C
80°C
80°C
80°C
120°C
0
0
0
0
0
1/2
1/10
1/10
1/10
1/2
1/10
1/10
1/10
1/10
1/10
1/10
0
0
18
37
45
53
56
50
45
3h
DMF
DMF
DMF
NMP
30 min
15 min
3 min
15 min
3 min
80°C μW
120°C μW
80°C μW
120°C μW
NMP
astoichiometric ratio between 5 and 6
Table 2: Optimization of the Buchwald-Hartwig reaction conditions.
O
BH
conditions
Br
O
NH2
+
NH
N
N
7
8
9
Entry
I
II
Pd source
Base
t-BuONa
Cs2CO3
K2CO3
K3PO4
t-BuONa
Cs2CO3
K2CO3
K3PO4
t-BuONa
Cs2CO3
K2CO3
K3PO4
Reaction conditionsa
130°C, 10 min, μW
130°C, 10 min, μW
130°C, 10 min, μW
130°C, 10 min, μW
130°C, 10 min, μW
130°C, 10 min, μW
130°C, 10 min, μW
130°C, 10 min, μW
130°C, 10 min, μW
130°C, 10 min, μW
130°C, 10 min, μW
130°C, 10 min, μW
Yield (%)
X-Phos Pd G2
X-Phos Pd G2
X-Phos Pd G2
X-Phos Pd G2
S-Phos Pd G2
S-Phos Pd G2
S-Phos Pd G2
S-Phos Pd G2
50
42
0
37
49
20
0
28
8
8
III
IV
V
VI
VII
VIII
IX
X
Dave Phos Pd(OAc)2
Dave Phos Pd(OAc)2
Dave Phos Pd(OAc)2
Dave Phos Pd(OAc)2
XI
XII
11
11
asee experimental part for details. 7 (1 equiv), 8 (0,5 equiv), base (1 equiv), catalyst (0,1 equiv), toluene
(2,5 mL/mmol), t-BuOH (0,5 mL/mmol).
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Table 3: Synthesis of variously substituted 2-aminooxazoles.
5
6
7
8
O
O
A
O
NH2
Br
+
N
R1
R1
DMF, 120°C
MW 3 min
H2N
NH2
9
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
40
41
42
43
44
45
46
47
48
49
50
51
52
53
54
55
56
57
58
59
60
5, 10 - 16
6
7, 17 - 23
Entry
5, 7
R1
Yield passage A (%)
p-CH3
o-CH3
m-CH3
o-Br
m-Br
p-F
56
52
52
40
57
41
25
38
10, 17
11, 18
12, 19
13, 20
14, 21
15, 22
16, 23
2-F, 5-CF3
p-OCH3
33
49
O
B
O
NH2
R2
N
R1
X-Phos Pd G2
-BuONa, 130°C
MW, 10 min,
toluene
N
R1
N
H
t
7, 22, 23
24 - 31
Entry
24
25
26
27
28
29
30
31
R1
p-CH3
p-CH3
p-CH3
p-CH3
R2
Yield passage B (%)
o-F
m-F
p-F
59
37
53
71
59
48
31
10
o-OCH3
m-OCH3
p-OCH3
p-CH3
p-CH3
p-CH3
2-F, 5-CF3
p-OCH3
p-CH3
34
16
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Table 4: Biological and physicochemical evaluation of 2-aminothiazoles and 2-aminooxazoles
Kinetic
solubility solubility (μM)
Kinetic
Mean GSH-
conjugate
area
303
Ratio (%)
GSH-
conjugate area
10.4±1.7
13.9±0.2
3.7±0.5
5
6
7
8
MIC
(μM)
HLM t1/2 HLM Cl’int
Entry
mind
(mL/min/kg)e
(μM)
24±2.7
20±2.4
54.9±15
2.9±0.5
5.2±0.6
0.7±0.2
9.1±2
PBS pH 7.4
7.5±1.1
1.2±0.2
27.7±8
1
2
3
4
30
27.5a
13a
22.4±1.6
14.5±0.5
3.3±0.3
16.1±0.2
22.6±1.2
20.9±1.4
2.7±0.8
15±0.1
27.9
42.9
188.8
38.8
27.6
29.9
226.1
41.5
9
616
34
111
2117
467
1.5b
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
40
41
42
43
44
45
46
47
48
49
50
51
52
53
54
55
56
57
58
59
60
0.16b
12.3a
2.4±0.5
5.9±0.5
1.1±0.3
6.8±1.1
9.3±1.2
7.1±0.9
14.2±5
14.2±1.5
15.8±1.3
2.1±0.9
31 >100a
34
36
2.3a
6.9c
120
82
24±4
MIC values determined by MABA are the mean of replicated experiments (SD < 15%);10 MIC determined by microdilution;7 cMetodo
PASCA dMeasured in human liver microsomes and expressed in minutes; eIntrinsic clearance was calculated as (0.693/in vitro t1/2)
× (mL incubation/mg microsomes) × (45 mg microsomes/g liver) × (20 g liver/kg body weight).
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Evaluation of
Antitubercular
activity
Solubility
PAINS
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
40
41
42
Metabolic
stability
Buchwald-Hartwig
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