Synthesis and biological evaluation of tryptophan-derived rhodanine derivatives as PTP1B inhibitors and anti-bacterial agents

Article abstract of DOI:10.1016/j.ejmech.2019.03.059

Several series of novel tryptophan-derived rhodanine derivatives were synthesized and identified as potential competitive PTP1B inhibitors and antibacterial agents. Among the compounds studied, 10b was found to have the best in vitro inhibition activity against PTP1B (IC₅₀ = 0.36 ± 0.02 μM). In addition, the compounds also showed potent inhibition against other PTPs, especially CDC25B. Molecular docking analysis demonstrated that compounds 7c and 10b could occupy both the catalytic site and the adjacent pTyr binding site simultaneously. The compounds also showed higher levels of activity against gram-positive strains, the gram-negative strain Escherichia coli 1924, and multidrug-resistant gram-positive bacterial strains. Compounds 7c, 8c, 9e, 10a, and 10c had comparable or more potent antibacterial activity than the positive controls.

Full text of DOI:10.1016/j.ejmech.2019.03.059

Accepted Manuscript
Synthesis and biological evaluation of tryptophan-derived rhodanine derivatives as
PTP1B inhibitors and anti-bacterial agents
Hongyan Liu, Danwen Sun, Hang Du, Changji Zheng, Jingya Li, Huri Piao, Jia Li,
Liangpeng Sun
PII:
S0223-5234(19)30287-9
DOI:
https://doi.org/10.1016/j.ejmech.2019.03.059
EJMECH 11229
Reference:
To appear in: European Journal of Medicinal Chemistry
Received Date: 14 December 2018
Revised Date: 6 March 2019
Accepted Date: 25 March 2019
Please cite this article as: H. Liu, D. Sun, H. Du, C. Zheng, J. Li, H. Piao, J. Li, L. Sun, Synthesis
and biological evaluation of tryptophan-derived rhodanine derivatives as PTP1B inhibitors and anti-
bacterial agents, European Journal of Medicinal Chemistry (2019), doi: https://doi.org/10.1016/
j.ejmech.2019.03.059.
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ACCEPTED MANUSCRIPT
Synthesis and biological evaluation of tryptophan-derived rhodanine
derivatives as PTP1B inhibitors and anti-bacterial agents
Hongyan Liu a, 1, Danwen Sun b, 1, Hang Du a, Changji Zheng a, Jingya Li c,
Huri Piao a, **, Jia Li c, **, Liangpeng Sun d, *
Several series of novel tryptophan-derived rhodanine derivatives were synthesized and
identified as potential competitive PTP1B inhibitors and antibacterial agents.

ACCEPTED MANUSCRIPT
Synthesis and biological evaluation of tryptophan-derived rhodanine
derivatives as PTP1B inhibitors and anti-bacterial agents
Hongyan Liu a, 1, Danwen Sun b, 1, Hang Du a, Changji Zheng a, Jingya Li c,
Huri Piao a, **, Jia Li c, **, Liangpeng Sun d, *
a
Key Laboratory of Natural Resources of Changbai Mountain & Functional Molecules, Ministry of
Education, Yanbian University College of Pharmacy, Yanji, 133000, PR China
b College of Chemistry and Molecular Engineering, East China of Normal University, 3663 Zhongshan
North Road, Shanghai, 200062, China
c National Center for Drug Screening, Shanghai Institute of Materia Medica, Shanghai Institutes for
Biological Science, Chinese Academy of Sciences, Shanghai 201203, China
d College of Medicine, Yanbian University, Yanji, 133000, PR China
* Corresponding author. Tel: + 86 433 2436596
E-mail address: lpsun@ybu.edu.cn (L.P. Sun)
** Co-corresponding author. Tel/Fax: + 86 21 50800721
E-mail address: jli@simm.ac.cn (J. Li)
** Co-corresponding author. Tel: + 86 433 2436149
E-mail address: piaohr@ybu.edu.cn (H.R. Piao)
1 These authors contributed equally to this work.

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Abstract Several series of novel tryptophan-derived rhodanine derivatives were
synthesized and identified as potential competitive PTP1B inhibitors and antibacterial
agents. Among the compounds studied, 10b was found to have the best in vitro
inhibition activity against PTP1B (IC50 = 0.36 ± 0.02 µM). In addition, the
compounds also showed potent inhibition against other PTPs, especially CDC25B.
Molecular docking analysis demonstrated that compounds 7c and 10b could occupy
both the catalytic site and the adjacent pTyr binding site simultaneously. The
compounds also showed higher levels of activity against gram-positive strains, the
gram-negative strain Escherichia coli 1924, and multidrug-resistant gram-positive
bacterial strains. Compounds 7c, 8c, 9e, 10a, and 10c had comparable or more potent
antibacterial activity than the positive controls.
Keywords: Tryptophan-derived rhodanine; PTP1B inhibitor; Anti-bacterial; SAR
1. Introduction
Protein tyrosine phosphatases (PTPs) play an important regulatory role in the
intracellular phosphorylation state of proteins [1-3]. Among the PTPs, protein tyrosine
phosphatase 1B (PTP1B) acts as a negative regulator of the insulin and leptin
signaling pathways, which are responsible for dephosphorylating the activated insulin
receptor, insulin receptor substrate (IRS)-1 and IRS-2, and the leptin receptor [4–8].
PTP1B-knockout mice exhibit increased insulin sensitivity, improved glucose
tolerance, and resistance to diet-induced obesity [9–10]. Therefore, PTP1B is a
potential therapeutic target for the treatment of type 2 diabetes mellitus and obesity.
Following elucidation of the basic structural requirements for PTP1B–substrate and
PTP1B–inhibitor interactions, a large variety of potent PTP1B inhibitors have been
identified through high-throughput screening, and structure- and fragment-based drug
design. However, the development of clinically relevant PTP1B inhibitors is still
problematic, with compounds often exhibiting poor selectivity and unfavorable
pharmacokinetics [11-12].

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Because of the electrostatic properties of the PTP1B enzyme active site [13], it has
proven difficult to develop effective uncharged phosphotyrosine (pTyr) mimetics.
Replacing the phosphate group of pTyr with bioisosteric monoanionic groups, such as
carboxylates, is considered a valid method to obtain inhibitors with low polarity
[14-18]. Indeed, several carboxylic acids with good PTP1B inhibitory profiles, and
cellular activity or oral bioavailability have been reported [19-20]. For example, as
shown in Fig. 1, compounds I were identified as inhibitors of PTP1B, in which a
4-((2,4-dioxothiazolidin-3-yl)methyl)benzoic acid moiety can act as a monoanionic
pTyr mimetic group and replicate the interactions of pTyr with the catalytic site of the
enzyme [21]. Subsequent studies showed that compounds II, where a 2-phenylimino
moiety was introduced into the 4-thiazolidinone ring, could further enhance the
affinity between the inhibitor and enzyme by means of favorable interactions with
active site residues and the surrounding loops [22].
Rhodanine derivatives have emerged as important target molecules because of their
therapeutic and pharmacological properties, such as anti-bacterial [23-27], antifungal
[28], antidiabetic [29], antitubercular [30], anticancer [31], anti-infective [32], and
anti-HIV [33] activities. As reported recently, rhodanine-3-acetic acid derivatives (III)
have been identified as PTP1B inhibitors, in which the rhodanine-3-acetic acid moiety
can act as a monoanionic pTyr mimetic group and interact with the catalytic site of the
enzyme. However, they showed weaker inhibitory activities against PTP1B [34]. All
these findings prompted us to expand the range of available pharmacophores from
which new PTP1B inhibitors can be developed. Inspired by compounds II, we
assumed that the introduction of suitable substituted groups at the 2-position of the
carboxyl group of the rhodanine-3-acetic acid might offer a possible alternative
skeleton which could bind simultaneously at both the catalytic site and the adjacent
pTyr binding site (Fig. 2), and will improve not only the binding affinity but also the
selectivity for PTP1B. Indole is one of the most important privileged scaffolds in drug
discovery, with a wide range of biological activities. Indole derivatives can mimic the
structure of peptides and bind reversibly to enzymes, which provide tremendous
opportunities to discover novel lead compounds with different modes of action [35].

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Herein,
a
series of tryptophan-derived rhodanine derivatives, where
a
(1H-indol-3-yl)methyl group was inserted onto the 2-position of the carboxyl group,
were designed, synthesized, and evaluated for their PTP1B inhibitory activity.
Interestingly, all of the tested compounds showed higher levels of PTP1B inhibitory
activity compared with compound III, as expected. Here, we report a series of
tryptophan-derived rhodanine compounds as novel and potent inhibitors of PTP1B.
The mechanism of inhibition of PTP1B by these compounds, along with
structure–activity relationships (SARs) were investigated.
In our previous work [24-27], a variety of rhodanine derivatives were synthesized
and identified as potential antibacterial agents, including rhodanine-3-acetic acid (IV),
phenylalanine-derived rhodanine (V) and leucine-derived rhodanine (VI) derivatives
(Fig. 1). Most compounds bearing a rhodanine-3-acetic acid scaffold showed
moderate to strong activity against several gram-positive bacterial strains, including
multidrug-resistant clinical isolates. Unfortunately, none of these compounds were
active against gram-negative bacteria. As a part of our ongoing investigations into the
development of rhodanine antibacterial agents, these tryptophan-derived rhodanine
derivatives were also evaluated for their in vitro antibacterial activity, although it is
not clear that there is any correlation between PTP1B inhibition and antimicrobial
activity.
2. Chemistry
A group of tryptophan-derived rhodanine derivatives (7–10) were synthesized, as
shown in Scheme 1. A key intermediate 3-(1H-indol-3-yl)-2-(2-thioxothiazolidin
-3-yl)propanoic acid (6) was prepared using a previously reported method [36].
Compounds 2–5 were obtained in high yield through a nucleophilic substitution
reaction of substituted benzyl chlorides with hydroxyaromatic aldehydes in refluxing
acetone in the presence of potassium carbonate. The resulting 2, 3, 4, and 5 series of
compounds were subjected to Knoevenagel condensation with intermediate 6 to form
the target compounds 7–10.

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3. Results and discussion
3.1. Evaluation of PTP1B inhibitory activity
The inhibitory activities of all the compounds against PTP1B were measured using
p-nitrophenyl phosphate (pNPP) as a substrate, and the results are summarized in
Tables 1. The known PTP1B inhibitor, oleanolic acid (IC50 = 2.69 ± 0.15 ꢀM), was
used as the positive control [37-38]. As shown in Table 1, all of the compounds tested
showed good PTP1B inhibitory activity at 20 ꢀg/mL and dose-dependently inhibited
PTP1B with IC50 values ranging from 0.36 ± 0.02 to 8.23 ± 0.45 ꢀM. Of these, 10b
was the most potent among the series with an IC50 value of 0.36 ± 0.02 ꢀM, about
8-fold better than that of the positive control.
The following SARs were obtained by analyzing the activities of compounds 7–10.
First, to investigate the SAR between PTP1B inhibitory activity and the position of
the different benzyloxy groups on the phenylidene group, compounds substituted at
the 4- (7) and the 3- (8) positions were synthesized. However, the compounds in
series 7 showed comparable activity against PTP1B as the compounds in series 8.
These results suggest that changing the position of the substituents on the phenylidene
group has no remarkable impact on PTP1B inhibition. Second, a comparison of the
compounds in series 7 and series 9 indicated that the introduction of a methoxy group
to the phenylidene group did not result in an obvious improvement in PTP1B
inhibitory activity. Third, interestingly, compared with compounds in series 7,
compounds with a naphthalene ring (series 10) had more impact on PTP1B inhibitory
activity with a 6–9-fold increase in potency, which might be attributed to the ability of
the naphthalene ring to form steric interactions with Ile261, Leu260, Leu227, Gly223
and Ile219 as suggested by the docking study (Fig. 4D). These results agree with
earlier studies in which lipophilic moieties stabilized an enzyme–compound complex
via hydrophobic interactions with the active site and surrounding subpockets, a
common feature of PTP1B–inhibitor complexes [18, 20-22, 39-40]. Among
compounds 10a–k, the halogen-substituted derivatives (10b–g) showed better activity
than compounds 10h–j containing electron-donating substituted groups (except for

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10a). The impact of the substituent position on the benzene ring on the efficacy was
not predictable. These results indicate that electron-withdrawing groups might
contribute more to the inhibitory activity toward PTP1B than electron-donating
groups.
Representative compounds (10a–d) were investigated for their selectivity toward
other PTPs, using a previously described method [34-35], including homogeneous
T-cell protein tyrosine phosphatase (TCPTP), cell division cycle 25 homolog B
(CDC25B), leukocyte antigen-related phosphatase (LAR), src homology
phosphatase-1 (SHP-1) and src homology phosphatase-2 (SHP-2). As shown in Table
2, these compounds showed better inhibition toward TCPTP, CDC25B, SHP1, and
SHP2, with no visible inhibitory effects on LAR. We determined that these
compounds showed 2–3-fold greater selectivity for PTP1B than for TCPTP,
approximately 3–6-fold selectivity for PTP1B over SHP-1, and approximately
9–10-fold selectivity for PTP1B over SHP-2. The compounds possessed comparable
inhibitory activity against PTP1B and CDC25B. CDC25B, a dual-specificity
phosphatase, is an attractive potential therapeutic antitumor target for small molecule
intervention due to its central role in controlling malignant cell proliferation by
regulating cyclin dependent kinases and because it is highly expressed in many human
tumors [41]. These results provide information toward developing suitable lead
compounds to aid in the design of other PTP inhibitors.
A kinetic study was performed in order to shed light on the inhibitory mechanism
of compound 10b. As also elucidated in Fig. 3, 10b demonstrated a time-independent
inhibition of PTP1B, which showed 10b was a fast-binding inhibitor of PTP1B (Fig.
3A). The time-independent inhibition of 10b towards PTP1B may also exclude that
10b is a nonspecific inhibitor, because nonspecific inhibitors always show
time-dependent behavior and steep inhibition curve [42]. As shown in Fig 3B, we
further determined the inhibition modality of 10b which inhibited PTP1B with the
characteristics typical of a competitive inhibitor, as indicated by increased Km values
and unchanged Vmax values when the inhibitor concentration was increased.
Meanwhile, the result of the Lineweaver-Burk plot confirmed 10b as a competitive

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inhibitor of PTP1B for intersecting at the y-axis of a nest of lines with increased
inhibitor concentration (Fig. 3C). The results indicate that 10b bind the catalytic
pocket of PTP1B and behave as a competitor to the substrate. The Ki values was
calculated as 0.07 µM from Figure 3D.
To understand the inhibitory activities of 7c and 10b against PTP1B, we subjected
them to molecular docking analysis using MOE Dock in MOE v2014.0901 [43] and
predicted their binding affinity with PTP1B (PDB code: 2vey). The binding modes of
the two compounds revealed by the docking studies are shown in Fig. 4. These results
indicated that their interaction modes were similar to that of compounds I and II
[21-22], in which the tryptophan-derived rhodanine group situated in the catalytic site
of PTP1B and the lipophilic 5-arylidene moiety extended into the hydrophobic
subpocket near the catalytic site as expected.
The binding mode of 7c with PTP1B is illustrated in Fig. 4B. The oxygen atom of
the carboxyl and the nitrogen atom of the indole ring, regarded as hydrogen bond
acceptor and donor, respectively, form two hydrogen bonds with the sidechain of
Asp181 and Cys215 in the catalytic site of PTP1B, respectively. The pyrrole ring of
the indole ring forms a H-π conjugation with the backbone of Ala217 in PTP1B. The
bromine atom of the terminal benzene ring, regarded as a hydrogen bond acceptor,
forms one hydrogen bond with the backbone of Ser28 in PTP1B. Additionally, the
phenylidene group provides good van der Waals interactions with Leu260 and Ile219
in PTP1B.
As shown in Fig. 4D, the oxygen atom of carboxyl and the sulfur atom of the
carbon-sulfur double bond of the thiazole ring of 10b, regarded as a hydrogen bond
acceptor, forms two hydrogen bonds with the backbone of Arg221 and Phe182 in the
catalytic site of PTP1B. The nitrogen atom of the indole ring, regarded as a hydrogen
bond acceptor, forms one hydrogen bond with the sidechain of Ser216 in PTP1B.
Some van der Waals interactions were also observed, such as interactions between the
naphthalene ring and Ile261, Leu260, Leu227, Gly223 and Ile219, the indole ring and
Tyr46.
The active site of PTP1B includes the residues Cys215, Asp181, Arg221, Ser216

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and Gly220, which were reported in previous studies as important in modulating its
activities [44]. Taken together, the docking simulation studies indicate that 7c
interacts with Asp181, Cys215, Ser28 and Ala217 of PTP1B through hydrogen bonds
and H-π conjugation, and 10b interacts with Ser216, Arg221 and Phe182 of PTP1B
through hydrogen bonds. However, the naphthalene ring of 10b provides more van
der Waals interactions than the benzene ring of 7c, which might explain the activity
difference between compound 7 and 10. The different binding models between
ligands and PTP1B result in different binding abilities. The docking scores of 7c and
10b are shown in Table 3, which are −11.29 kcal/mol and −13.94 kcal/mol for PTP1B,
respectively. The computational results indicate that 7c and 10b interact with PTP1B
and their binding ability with PTP1B is in the order 10b >7c.
3.2. Evaluation of antibacterial activity
The in vitro antibacterial activities of the synthesized compounds (7-10) were
evaluated using a 96-well microtiter plate and a serial dilution method to obtain the
minimum inhibitory concentrations (MICs) for different strains (including
multidrug-resistant clinical isolates). The compounds were screened against four
Gram-positive strains (S. aureus RN 4220, S. aureus KCTC 209, S. aureus KCTC 503
and S. Mutans KTCT 3289), two Gram-negative strains (Escherichia coli 1924 and P.
aeruginosa 2742) and one fungus (Candida albicans 7535). Gatifloxacin,
moxifloxacin, norfloxacin and oxacillin were used as positive controls for
antibacterial activity, and fluconazole was used as positive controls for antifungal
activity.
The in vitro antibacterial activity of the synthesized compounds is summarized in
Table 4. All the synthesized compounds showed potent inhibition against four types of
gram-positive bacteria with MIC values in the range of 1–8 µg/mL. For S. aureus RN
4220, compounds 7c and 10c (MIC = 1 µg/mL) were slightly less active than the
positive controls gatifloxacin and moxifloxacin (MIC = 0.25 µg/mL). For S. aureus
KCTC 209, the compounds 8c, 9b, 9e, 10a, and 10b (MIC = 2 µg/mL) had
comparable activity to that of gatifloxacin and moxifloxacin (MIC = 2 µg/mL). For S.

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aureus KCTC 503, the compounds 7c, 8c, 9e, 10a, and 10c (MIC = 2 µg/mL) had
more potent activity (2-fold) than gatifloxacin (MIC = 4 µg/mL) and comparable
activity to that of moxifloxacin (MIC = 2 µg/mL). For S. Mutans KTCT 3289,
compound 7c exhibited good inhibitory activity with a MIC value of 1 µg/mL, but
was less active than gatifloxacin and moxifloxacin (MIC = 0.25 µg/mL). All the
synthesized compounds showed potent inhibition against the gram-negative strain
Escherichia coli 1924 with MIC values in the range of 1–16 µg/mL. Among them,
compound 7c (MIC = 1 µg/mL) was 2-fold more potent than the positive controls
gatifloxacin and moxifloxacin (MIC = 2 µg/mL). Compounds 8c, 9b, 9e, 10a, 10c,
10f and 10g (MIC = 2 µg/mL) had comparable activity to that of gatifloxacin and
moxifloxacin (MIC = 2 µg/mL). Disappointingly, none of the compounds synthesized
in the current study exhibited activity against P. aeruginosa 2742 or Candida albicans
7535 at 64 µg/mL.
The synthesized compounds were also evaluated for their inhibitory activity against
the clinical isolates of several different multidrug-resistant, gram-positive bacterial
strains (methicillin-resistant S. aureus (MRSA) CCARM 3167 and 3506, and
quinolone-resistant S. aureus (QRSA) CCARM 3505 and 3519). As shown in Table 5,
all the compounds showed high levels of activity with MIC values of 2–16 µg/mL.
For MRSA 3167 and 3506, most compounds had comparable or more potent activity
than the positive controls. Among them, compounds 7c, 9e, 10c and 10g with MIC
values of 2 µg/mL, were 4-fold more potent than norfloxacin (MIC = 8 µg/mL) and
64-fold more potent than oxacillin (MIC >64 µg/mL), comparable with gatifloxacin
(MIC = 2 µg/mL), and slightly less active than moxifloxacin (MIC = 1 µg/mL). For
QRSA 3505 and 3519, compounds 7c, 9e, 10a, and 10c also showed high levels of
activity (MIC = 2 µg/mL), and were slightly less active than oxacillin (MIC = 1
µg/mL) but more potent than norfloxacin (MIC >64 µg/mL), gatifloxacin (MIC = 8 or
4 µg/mL), and moxifloxacin (MIC = 4 µg/mL).
Several SARs could be determined from these results. First, there was no
remarkable difference in antibacterial activity between compounds 7, 8, 9, and 10.
Second, a comparison of compounds 10b–g with compounds 10h–j indicated that the

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presence of electron-withdrawing groups might favor antibacterial activity more than
that of electron-donating groups. The impact of the position of the substituents on the
benzene ring on the efficacy was not predictable. Third, all the synthesized
compounds exhibited high levels of activity against E. coli 1924 with MIC values in
the range of 1–16 µg/mL. Compared with previously reported rhodanine-3-acetic acid
derivatives (such as compound IV, with no activity against E. coli 1924) that were
prepared in our laboratory [25], the introduction of a (1H-indol-3-yl)methyl group
appeared to be critical for the inhibition of both gram-negative and gram-positive
strains of bacteria.
4. Conclusion
In the current study, a series of tryptophan-derived rhodanine derivatives were
synthesized and evaluated for their PTP1B inhibitory and antibacterial activity. The
compounds were shown to be competitive PTP1B inhibitors. Of the compounds, 10b
was found to have the best in vitro inhibitory activity against PTP1B (IC50 = 0.36 ±
0.02 µM). The compounds also showed potent inhibition against other PTPs,
especially CDC25B. A preliminary SAR study highlighted that the introduction of a
(1H-indol-3-yl)methyl group at the 2-position of the carboxyl group remarkably
improved the PTP1B inhibitory activity compared with compound III. Molecular
docking studies of two compounds (7c and 10b) indicated that the tryptophan-derived
rhodanine moiety situated in the catalytic site of PTP1B and the lipophilic 5-arylidene
moiety extended into the hydrophobic subpocket near the catalytic site, which showed
a possible binding mode and provided insights for further optimization of the scaffold.
Additionally, the compounds showed higher levels of activity against gram-positive
strains, the gram-negative strain E. coli 1924, and multidrug-resistant gram-positive
bacterial strains compared with the lead compound (IV). Compounds 7c, 8c, 9e, 10a,
and 10c had comparable or more potent antibacterial activity compared with the
positive controls. Unfortunately, none of the compounds synthesized exhibited
activity against P. aeruginosa 2742 or C. albicans 7535 at 64 µg/mL. These results

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provide the basis for the further design and development of novel PTP1B inhibitors
and antibacterial agents.
5. Experimental protocols
5.1. Chemistry
Melting points were determined in open capillary tubes and are uncorrected.
Reaction courses were monitored by TLC on silica gelprecoated F254 Merck plates.
1
13
Developed plates were examined with UV lamps (254 nm). H NMR and C NMR
spectra were measured on a Bruker AV-300 spectrometer using TMS as the internal
standard. ESI-HRMS spectra were recorded on a Thermo scientific LTQ Orbitrap XL
mass spectrometer. The major chemicals were purchased from Sigma-Aldrich and
Fluka.
5.2 General procedure for the synthesis of 7–10
Compound 6 was synthesized as previously reported [36].
A mixture of hydroxyaromatic aldehydes (1.0 mmol), appropriate benzyl chlorides
(1.2 mmol), and anhydrous sodium carbonate was stirred for 4 h in refluxing acetone.
The solvent was evaporated under reduced pressure and the residue was dissolved in
dichloromethane, washed using water and brine, dried over anhydrous MgSO4, and
the solvent was removed under reduced pressure. The resulting residue was purified
through silica gel column chromatography (dichloromethane/PE, 5:1 v/v) to afford
compounds 2–5 as white solids. To a solution of each of compounds 2–5 (1.0 mmol)
and compound 6 (1.0 mmol) in absolute ethanol (5 mL), was added drops of glacial
acetic acid and piperidine. The reaction mixture was stirred at 80°C for 2–6 h, until
the completion of the reaction as evidenced through TLC. The resulting reaction
mixture was concentrated to dryness, and purified through silica gel column
chromatography (dichloromethane/methanol/acetic acid, 100:1:1) to afford the pure
products 7–10. The yield, melting point and spectral data of each compound are given
below.

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5.2.1 (Z)-2-(5-(4-(Benzyloxy)benzylidene)-4-oxo-2-thioxothiazolidin-3-yl)-3-(1H-
indol-3-yl)propanoic acid (7a)
Yield 30%, m.p. 60-68 °C. 1H NMR (300 MHz, DMSO-d6, ppm) δ 3.71-3.74 (m, 2H,
CH2), 5.19 (s, 2H, CH2), 5.76 (br s, 1H, CH), 6.89 (t, 1H, J = 7.5 Hz, Ar–H),
6.97-7.01 (m, 2H, Ar–H), 7.18 (d, 2H, J = 9.0 Hz, Ar–H), 7.25 (d, 1H, J = 6.0 Hz),
7.34-7.47 (m, 6H, Ar–H), 7.55-7.58 (d, 2H, J = 9.0 Hz), 7.70 (s, 1H, Ar–H), 10.75 (s,
1H, NH), 13.90 (s, 1H, COOH). 13C NMR (75 MHz, DMSO-d6, ppm) δ 193.55,
169.97, 167.28, 161.04, 136.86, 136.43, 133.36 (2C), 128.97 (2C), 128.52, 128.32
(2C), 127.68, 126.13, 123.64, 121.29, 118.73, 118.41, 116.37 (2C), 111.78, 110.69,
70.03, 64.05, 23.65. ESI-HRMS calcd for C28H22N2O4S2 [M + Na] +: 537.0913, found:
537.0918.
5.2.2 (Z)-2-(5-(4-(4-Methylbenzyloxy)benzylidene)-4-oxo-2-thioxothiazolidin-3-
yl)-3-(1H-indol-3-yl)propanoic acid (7b)
1
Yield 55%, m.p. 158-160 °C. H NMR (300 MHz, DMSO-d6, ppm) δ 2.30 (s, 3H,
CH3), 3.57-3.77 (m, 2H, CH2), 5.19 (s, 2H, CH2), 5.69 (br s, 1H, CH), 6.89 (t, 1H, J =
7.5 Hz, Ar–H), 6.96-7.01 (m, 3H, Ar–H), 7.13-7.26 (m, 4H, Ar–H), 7.34 (d, 2H, J =
6.0 Hz, Ar–H), 7.45 (d, 1H, J = 6.0 Hz), 7.55 (d, 2H, J = 9.0 Hz), 7.68 (s, 1H, Ar–H),
10.75 (s, 1H, NH). ESI-HRMS calcd for C29H24N2O4S2 [M + Na] +: 551.1070, found:
551.1062.
5.2.3 (Z)-2-(5-(4-(4-Bromobenzyloxy)benzylidene)-4-oxo-2-thioxothiazolidin-3-
yl)-3-(1H-indol-3-yl)propanoic acid (7c)
1
Yield 55%, m.p.128-130 °C. H NMR (300 MHz, DMSO-d6, ppm) δ 3.61-3.79 (m,
2H, CH2), 5.17 (s, 2H, CH2), 5.59 (br s, 1H, CH), 6.86-7.01 (m, 4H, Ar–H), 7.16 (d,
2H, J = 9.0 Hz, Ar–H), 7.24 (d, 1H, J = 6.0 Hz), 7.43 (br s, 1H, Ar–H), 7.52-7.61 (m,
+
6H, Ar–H), 10.68 (s, 1H, NH). ESI-HRMS calcd for C28H21BrN2O4S2 [M + Na] :
615.0018, found: 615.0017.

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5.2.4 (Z)-2-(5-(4-(4-Chlorobenzyloxy)benzylidene)-4-oxo-2-thioxothiazolidin
-3-yl)-3-(1H-indol-3-yl)propanoic acid (7d)
Yield 42%, m.p. 58-60 °C. 1H NMR (300 MHz, DMSO-d6, ppm) δ 3.62-3.78 (m, 2H,
CH2), 5.19 (s, 2H, CH2), 5.63 (br s, 1H, CH), 6.89 (t, 1H, J = 7.5 Hz, Ar–H),
6.96-7.01 (m, 3H, Ar–H), 7.17 (d, 2H, J = 9.0 Hz, Ar–H), 7.25 (d, 1H, J = 6.0 Hz,
Ar–H), 7.43-7.51 (m, 4H, Ar–H), 7.56 (d, 2H, J = 9.0 Hz, Ar–H), 7.66 (br s, 1H,
Ar–H), 10.73 (s, 1H, NH). 13C NMR (75 MHz, DMSO-d6, ppm) δ 193.24, 169.46,
166.89, 159.03, 136.41, 136.12, 134.58, 133.94, 133.02, 131.17, 130.02, 128.96,
127.55, 124.23, 123.50, 121.95, 121.41, 118.87, 118.53, 118.34, 117.01, 111.85,
+
109.42, 69.00, 58.59, 23.53. ESI-HRMS calcd for C28H21ClN2O4S2 [M + Na] :
571.0523, found: 571.0526.
5.2.5 (Z)-2-(5-(4-(2,4-Dichlorobenzyloxy)benzylidene)-4-oxo-2-thioxothiazolidin
-3-yl)-3-(1H-indol-3-yl)propanoic acid (7e)
1
Yield 36%, m.p. 125-130 °C. H NMR (300 MHz, DMSO-d6, ppm) δ 3.60-3.72 (m,
2H, CH2), 5.20 (s, 2H, CH2), 5.80 (br s, 1H, CH), 6.9-7.03 (m, 3H, Ar–H), 7.19-7.25
(m, 3H, Ar–H), 7.40-7.45 (m, 4H, Ar–H), 7.58 (br s, 2H, Ar–H), 7.72 (br s, 1H,
+
Ar–H), 10.77 (s, 1H, NH). ESI-HRMS calcd for C28H20Cl2N2O4S2 [M + Na] :
605.0134, found: 605.0124.
5.2.6 (Z)-2-(5-(3-(Benzyloxy)benzylidene)-4-oxo-2-thioxothiazolidin-3-yl)-3-(1H
-indol-3-yl)propanoic acid (8a)
1
Yield 20%, m.p. 120-122 °C. H NMR (300 MHz, DMSO-d6, ppm) δ 3.61-3.66 (m,
2H, CH2), 5.16 (s, 2H, CH2), 5.84 (br s, 1H, CH), 6.90 (t, 1H, J = 7.5 Hz, Ar–H),
7.00 (t, 1H, J = 7.5 Hz, Ar–H), 7.05 (s, 1H, Ar–H), 7.17 (dd, 2H, J = 9.0, 3.0 Hz,
Ar–H), 7.22 (s, 1H, Ar–H), 7.24 (d, 1H, J = 9.0 Hz, Ar–H), 7.28-7.48 (m, 7H, Ar–H),
13
7.75 (s, 1H, Ar–H), 10.81 (s, 1H, NH), 13.57 (s, 1H, COOH). C NMR (75 MHz,
DMSO-d6, ppm) δ 193.31, 169.43, 166.92, 159.21, 137.07, 136.40, 134.58, 133.88,
131.14, 128.96 (2C), 128.44, 128.23 (2C), 127.55, 124.14, 123.38, 121.93, 121.37,
118.83, 118.55, 118.34, 116.97, 111.83, 109.58, 69.84, 58.85, 23.56. ESI-HRMS calcd

ACCEPTED MANUSCRIPT
for C28H22N2O4S2 [M + Na] +: 537.0913, found: 537.0919.
5.2.7 (Z)-2-(5-(3-(4-Bromobenzyloxy)benzylidene)-4-oxo-2-thioxothiazolidin-3-
yl)-3-(1H-indol-3-yl)propanoic acid (8b)
1
Yield 25%, m.p. 162-163 °C. H NMR (300 MHz, DMSO-d6, ppm) δ 3.59-3.79 (m,
2H, CH2), 5.15 (s, 2H, CH2), 5.60 (br s, 1H, CH2), 6.89 (t, 1H, J = 7.5 Hz, Ar–H),
6.96-7.02 (m, 3H, Ar–H), 7.12-7.19 (m, 3H, Ar–H), 7.25 (d, 1H, J = 6.0 Hz, Ar–H),
7.41-7.47 (m, 4H, Ar–H), 7.59 (d, 1H, J = 6.0 Hz, Ar–H), 7.65 (s, 1H, Ar–H), 10.72
+
(s, 1H, NH). ESI-HRMS calcd for C28H21BrN2O4S2 [M + Na] : 615.0018, found:
615.0022.
5.2.8 (Z)-2-(5-(3-(4-Chlorobenzyloxy)benzylidene)-4-oxo-2-thioxothiazolidin-3-
yl)-3-(1H-indol-3-yl)propanoic acid (8c)
1
Yield 60%, m.p. 130-134 °C. H NMR (300 MHz, DMSO-d6, ppm) δ 3.58-3.69 (m,
2H, CH2), 5.17 (s, 2H, CH2), 5.87 (br s, 1H, CH2), 6.90 (t, 1H, J = 7.5 Hz, Ar–H),
7.00 (t, 1H, J = 7.5 Hz, Ar–H), 7.06 (s, 1H, Ar–H), 7.18 (t, 3H, J = 7.5 Hz, Ar–H),
7.27 (d, 1H, J = 9.0 Hz, Ar–H), 7.44-7.48 (m, 6H, Ar–H), 7.76 (s, 1H, Ar–H), 10.81 (s,
1H, NH), 13.48 (s, 1H, COOH). 13C NMR (75 MHz, DMSO-d6, ppm) δ 193.21,
169.56, 167.02, 150.96, 149.73, 136.83, 136.41, 134.40, 128.96 (2C), 128.55, 128.43
(2C), 127.60, 126.21, 125.31, 124.06, 121.37, 118.82, 118.48, 118.39, 114.27, 114.01,
111.83, 109.69, 70.38, 56.05, 23.61. ESI-HRMS calcd for C28H21ClN2O4S2 [M + Na]
+: 571.0523, found: 571.0521.
5.2.9 (Z)-2-(5-(3-(2,4-Dichlorobenzyloxy)benzylidene)-4-oxo-2-thioxothiazolidin-3-
yl)-3-(1H-indol-3-yl)propanoic acid (8d)
Yield 48%, m.p. 86-88 °C. 1H NMR (300 MHz, DMSO-d6, ppm) δ 3.56-3.69 (m, 2H,
CH2), 5.21 (s, 2H, CH2), 5.87 (br s, 1H, CH), 6.90 (t, 1H, J = 6.0 Hz, Ar–H), 7.00 (t,
1H, J = 7.5 Hz, Ar–H), 7.06 (s, 1H, Ar–H), 7.20 (d, 2H, J = 9.0 Hz, Ar–H), 7.27 (d,
2H, J = 9.0 Hz, Ar–H), 7.46-7.50 (m, 3H, Ar–H), 7.63 (d, 1H, J = 9.0 Hz, Ar–H),
7.71 (d, 1H, J = 3.0 Hz, Ar–H), 7.78 (s, 1H, Ar–H), 10.83 (s, 1H, NH), 13.46 (s, 1H,

ACCEPTED MANUSCRIPT
+
COOH). ESI-HRMS calcd for C28H20Cl2N2O4S2 [M + Na] : 605.0134, found:
605.0137.
5.2.10 (Z)-2-(5-(4-(Benzyloxy)-3-methoxybenzylidene)-4-oxo-2-thioxothiazolidin-
3-yl)-3-(1H-indol-3-yl)propanoic acid (9a)
1
Yield 33%, m.p. 136-140 °C. H NMR (300 MHz, DMSO-d6, ppm) δ 3.57-3.73 (m,
2H, CH2), 3.81 (s, 3H, OCH3), 5.18 (s, 2H, CH2), 5.85 (br s, 1H, CH2), 6.90 (t, 1H, J
= 7.5 Hz, Ar–H), 6.98-7.04 (m, 2H, Ar–H), 7.16-7.28 (m, 4H, Ar–H), 7.34-7.49 (m,
6H, Ar–H), 7.73 (s, 1H, Ar–H), 10.81 (s, 1H, NH), 13.59 (s, 1H, COOH).
ESI-HRMS calcd for C29H24N2O5S2 [M + H] +: 545.1199, found: 545.1195.
5.2.11 (Z)-2-(5-(4-(4-Methylbenzyloxy)-3-methoxybenzylidene)-4-oxo-2-
thioxothiazolidin-3-yl)-3-(1H-indol-3-yl)propanoic acid (9b)
1
Yield 29%, m.p. 126-130 °C. H NMR (300 MHz, DMSO-d6, ppm) δ 2.30 (s, 3H,
CH3), 3.58-3.72 (m, 2H, CH2), 3.82 (s, 3H, OCH3), 5.22 (s, 2H, CH2), 5.80 (br s, 1H,
CH), 6.91 (t, 1H, J = 7.5 Hz, Ar–H), 7.01 (t, 1H, J = 6.0 Hz, Ar–H), 7.06 (s, 1H,
Ar–H), 7.18-7.28 (m, 5H, Ar–H), 7.49 (t, 2H, J = 7.5 Hz, Ar–H), 7.61 (d, 1H, Ar–H),
7.74 (d, 2H, J = 9.0 Hz, Ar–H), 10.80 (s, 1H, NH), 13.41 (s, 1H, COOH).
ESI-HRMS calcd for C30H26N2O5S2 [M + Na] +: 581.1175, found: 581.1179.
5.2.12 (Z)-2-(5-(4-(4-Bromobenzyloxy)-3-methoxybenzylidene)-4-oxo-2-
thioxothiazolidin-3-yl)-3-(1H-indol-3-yl)propanoic acid (9c)
1
Yield 24%, m.p. 148-152 °C. H NMR (300 MHz, DMSO-d6, ppm) δ 3.58-3.82 (m,
2H, CH2), 3.83 (s, 3H, OCH3), 5.16 (s, 2H, CH2), 5.59 (br s, 1H, CH), 6.90-6.99 (m,
3H, Ar–H), 7.17-7.26 (m, 4H, Ar–H), 7.42 (br s, 3H, Ar–H), 7.61 (d, 3H, J = 9.0 Hz,
+
Ar–H), 10.68 (s, 1H, NH). ESI-HRMS calcd for C29H23BrN2O5S2 [M + Na] :
645.0124, found: 645.0133.
5.2.13 (Z)-2-(5-(4-(4-Chlorobenzyloxy)-3-methoxybenzylidene)-4-oxo-2-
thioxothiazolidin-3-yl)-3-(1H-indol-3-yl)propanoic acid (9d)

ACCEPTED MANUSCRIPT
1
Yield 42%, m.p. 156-160 °C. H NMR (300 MHz, DMSO-d6, ppm) δ 3.60-3.80 (m,
2H, CH2), 3.81 (s, 3H, OCH3), 5.18 (s, 2H, CH2), 5.67 (br s, 1H, CH), 6.90 (t, 1H, J =
7.5 Hz, Ar–H), 6.97-7.02 (m, 2H, Ar–H), 7.14-7.19 (m, 3H, Ar–H), 7.25 (d, 1H, J =
6.0 Hz, Ar–H), 7.44-7.47 (m, 5H, Ar–H), 7.66 (s, 1H, Ar–H), 10.73 (s, 1H, NH). 13C
NMR (75 MHz, DMSO-d6, ppm) δ 193.37, 169.54, 167.13, 161.13, 136.82, 136.40,
134.25, 133.46, 132.04, 129.47, 128.97 (2C), 128.53, 128.32 (2C), 127.61, 126.04,
123.93, 121.33, 118.78, 118.39, 116.38, 113.68, 111.81, 110.01, 70.03, 59.26, 23.67.
ESI-HRMS calcd for C29H23ClN2O5S2 [M + Na] +: 601.0629, found: 601.0620.
5.2.14 (Z)-2-(5-(4-(2,4-Dichlorobenzyloxy)-3-methoxybenzylidene)-4-oxo-2-
thioxothiazolidin-3-yl)-3-(1H-indol-3-yl)propanoic acid (9e)
1
Yield 48%, m.p. 108-112 °C. H NMR (300 MHz, DMSO-d6, ppm) δ 3.60-3.71 (m,
2H, CH2), 3.81 (s, 3H, OCH3), 5.22 (s, 2H, CH2), 5.88 (br s, 1H, CH), 6.90 (t, 1H, J
= 7.5 Hz, Ar–H), 7.01 (t, 1H, J = 6.0 Hz, Ar–H), 7.06 (s, 1H, Ar–H), 7.23-7.28 (m,
4H, Ar–H), 7.49 (t, 2H, J = 7.5 Hz, Ar–H), 7.61 (d, 1H, J = 9.0 Hz, Ar–H), 7.74 (d,
2H, J = 9.0 Hz, Ar–H), 10.81 (s, 1H, NH), 13.44 (s, 1H, COOH). ESI-HRMS calcd
for C29H22Cl2N2O5S2 [M + H] +: 613.0420, found: 613.0424.
5.2.15(Z)-2-(5-((6-(4-Methylbenzyloxy)naphthalen-2-yl)methylene)-4-oxo-2-
thioxothiazolidin-3-yl)-3-(1H-indol-3-yl)propanoic acid (10a)
1
Yield 32%, m.p. 156-158 °C. H NMR (300 MHz, DMSO-d6, ppm) δ 2.31 (s, 3H,
CH3), 3.58-3.77 (m, 2H, CH2), 5.20 (s, 2H, CH2), 5.72 (br s, 1H, CH), 6.90 (t, 1H, J =
7.5 Hz, Ar–H), 6.97-7.02 (m, 2H, Ar–H), 7.22 (d, 2H, J = 6.0 Hz, Ar–H), 7.27-7.32
(m, 2H, Ar–H), 7.40 (d, 2H, J = 6.0 Hz, Ar–H), 7.46-7.50 (m, 2H, Ar–H), 7.62 (d, 1H,
J = 9.0 Hz, Ar–H), 7.83 (s, 1H, Ar–H), 7.91 (d, 1H, J = 9.0 Hz, Ar–H), 8.00 (d, 1H, J
13
= 9.0 Hz, Ar–H), 8.13 (s, 1H, Ar–H), 10.75 (s, 1H, NH). C NMR (75 MHz,
DMSO-d6, ppm) δ 193.76, 169.85, 167.28, 158.79, 137.75, 136.47, 135.66, 133.99,
132.35, 131.20, 129.50 (2C), 128.70, 128.49 (2C), 127.72, 127.43, 123.63, 121.28,
118.73, 118.44, 111.79, 110.84, 107.86, 69.91, 56.51, 22.72. ESI-HRMS calcd for
C33H26N2O4S2 [M + Na] +: 601.1226, found: 601.1230.

ACCEPTED MANUSCRIPT
5.2.16 (Z)-2-(5-((6-(4-Bromobenzyloxy)naphthalen-2-yl)methylene)-4-oxo-2-
thioxothiazolidin-3-yl)-3-(1H-indol-3-yl)propanoic acid (10b)
1
Yield 26%, m.p. 126-130 oC. H NMR (300 MHz, DMSO-d6, ppm) δ 3.62-3.82 (m,
2H, CH2), 5.24 (s, 2H, CH2), 5.61 (br s, 1H, CH), 6.90 (t, 1H, J = 7.5 Hz, Ar–H),
6.96-7.02 (m, 2H, Ar–H), 7.25 (d, 1H, J = 6.0 Hz, Ar–H), 7.32 (d, 1H, J = 9.0 Hz,
Ar–H), 7.47-7.49 (m, 4H, Ar–H), 7.62 (d, 3H, J = 9.0 Hz, Ar–H), 7.77 (s, 1H, Ar–H),
7.91 (d, 1H, J = 9.0 Hz, Ar–H), 8.00 (d, 1H, J = 9.0 Hz, Ar–H), 8.10 (s, 1H, Ar–H),
+
10.71 (s, 1H, NH). ESI-HRMS calcd for C32H23BrN2O4S2 [M + Na] : 665.0175,
found: 665.0172.
5.2.17 (Z)-2-(5-((6-(4-Fluorobenzyloxy)naphthalen-2-yl)methylene)-4-oxo-2-
thioxothiazolidin-3-yl)-3-(1H-indol-3-yl)propanoic acid (10c)
1
Yield 26%, m.p. 102-104 °C. H NMR (300 MHz, DMSO-d6, ppm) δ 3.63-3.74 (m,
2H, CH2), 5.25 (s, 2H, CH2), 5.83 (br s, 1H, CH), 6.90 (t, 1H, J = 7.5 Hz, Ar–H), 7.00
(t, 1H, J = 7.5 Hz, Ar–H), 7.06 (d, 1H, J = 3.0 Hz, Ar–H), 7.22-7.28 (m, 4H, Ar–H),
7.48-7.60 (m, 5H, Ar–H), 7.88-7.94 (m, 2H, Ar–H), 8.02 (d, 1H, J = 9.0 Hz, Ar–H),
13
8.15 (s, 1H, Ar–H), 10.79 (s, 1H, NH), 13.58 (s, 1H, COOH). C NMR (75 MHz,
DMSO-d6, ppm) δ 193.43, 169.55, 167.11, 162.35 (1C, d, J = 242.25 Hz), 158.73,
136.44, 135.73, 134.16, 133.27 (1C, d, J = 3.0 Hz), 132.52, 131.30, 130.71 (1C, d, J =
8.25 Hz), 128.73, 128.65, 128.47, 127.62, 127.50, 124.01, 121.34, 120.55, 120.34,
118.80, 118.39, 115.79 (1C, d, J = 21.0 Hz), 111.83, 109.91, 107.88, 69.27, 59.22,
23.69. ESI-HRMS calcd for C32H23FN2O4S2 [M + Na] +: 605.0975, found: 605.0968.
5.2.18 (Z)-2-(5-((6-(2,4-Dichlorobenzyloxy)naphthalen-2-yl)methylene)-4-oxo-2-
thioxothiazolidin-3-yl)-3-(1H-indol-3-yl)propanoic acid (10d)
1
Yield 36%, m.p. 172-174 °C. H NMR (300 MHz, DMSO-d6, ppm) δ 3.64-3.79 (m,
2H, CH2), 5.30 (s, 2H, CH2), 5.74 (br s, 1H, CH), 6.90 (t, 1H, J = 7.5 Hz, Ar–H),
6.97-7.02 (m, 2H, Ar–H), 7.26 (d, 1H, J = 9.0 Hz, Ar–H), 7.33 (dd, 1H, J = 9.0, 3.0
Hz, Ar–H), 7.46-7.55 (m, 3H, Ar–H), 7.63 (d, 1H, J = 6.0 Hz, Ar–H), 7.69-7.73 (m,

ACCEPTED MANUSCRIPT
2H, Ar–H), 7.84 (s, 1H, Ar–H), 7.95 (d, 1H, J = 9.0 Hz, Ar–H), 8.02 (d, 1H, J = 9.0
Hz, Ar–H), 8.14 (s, 1H, Ar–H), 10.00 (s, 1H, NH). ESI-HRMS calcd for
C32H22Cl2N2O4S2 [M + Na] +: 655.0290, found: 655.0296.
5.2.19 (Z)-2-(5-((6-((2-Chlorobenzyl)oxy)naphthalen-2-yl)methylene)-4-oxo-2-
thioxothiazolidin-3-yl)-3-(1H-indol-3-yl)propanoic acid (10e)
1
Yield 26%, m.p. 100-102 °C. H NMR (300 MHz, DMSO-d6, ppm) δ 3.61-3.72 (m,
2H, CH2), 5.25 (s, 2H, CH2), 5.78 (br s, 1H, Ar–H), 7.04 (d, 1H, J = 6 Hz, Ar–H),
7.05-7.11 (m, 1H, Ar–H), 7.11-7.16 (m, 1H, Ar–H), 7.20-7.38 (m, 5H, Ar–H), 7.41
(dd, 1H, J = 6, 3 Hz, Ar–H), 7.56 (dd, 1H, J = 6, 3 Hz, Ar–H), 7.58-7.85 (m, 5H,
Ar–H), 8.04 (s, 1H, Ar–H), 9.96 (s, 1H, NH). 13C NMR (75 MHz, DMSO-d6, ppm) δ
192.58, 173.43, 167.22, 158.41, 138.56, 136.38, 135.16, 132.71, 132.14, 130.68,
129.46, 129.22 (2C), 128.81, 128.74, 128.65, 127.95, 127.30, 127.02, 126.85, 123.16,
122.09, 120.60, 120.14, 119.60, 118.47, 111.12, 110.20, 107.26, 67.21, 57.50, 23.64.
ESI-HRMS calcd for C32H24ClN2O4S2 [M + H] +: 599.0860, found: 599.0866.
5.2.20 (Z)-2-(5-((6-((3-Chlorobenzyl)oxy)naphthalen-2-yl)methylene)-4-oxo-2-
thioxothiazolidin-3-yl)-3-(1H-indol-3-yl)propanoic acid (10f)
1
Yield 28%, m.p. 110-112 °C. H NMR (300 MHz, DMSO-d6, ppm) δ 3.65-3.83 (m,
2H, CH2), 5.15 (s, 2H, CH2), 5.69 (br s, 1H, CH), 7.09 (dd, 3H, J = 18, 6 Hz, Ar–H),
7.20-7.37 (m, 4H, Ar–H), 7.39-7.57 (m, 5H, Ar–H), 7.62 (d, 1H, J = 12 Hz, Ar–H),
7.78 (dd, 3H, J = 9, 6 Hz, Ar–H), 8.08 (s, 1H, Ar–H). 13C NMR (75 MHz, DMSO-d6,
ppm) δ 192.76, 170.01, 167.26, 158.20, 138.20, 136.59, 135.87, 135.20, 134.30,
133.24, 131.90, 131.74, 130.38, 129.71, 128.59, 127.99, 127.70, 127.70, 126.62,
125.20, 122.81, 121.30, 120.75, 119.90, 118.75, 118.01, 110.92, 109.85, 106.94, 68.98,
+
57.80, 23.38. ESI-HRMS calcd for C32H24ClN2O4S2 [M + H] : 599.0860, found:
599.0863.
5.2.21 (Z)-2-(5-((6-((4-Chlorobenzyl)oxy)naphthalen-2-yl)methylene)-4-oxo-2-

ACCEPTED MANUSCRIPT
thioxothiazolidin-3-yl)-3-(1H-indol-3-yl)propanoic acid (10g)
1
Yield 16%, m.p. 150-152 °C. H NMR (300 MHz, DMSO-d6, ppm) δ 3.67-3.76 (m,
2H, CH2), 5.14 (s, 2H, CH2), 5.84 (br s, 1H, CH), 7.06 (d, 1H, J = 6 Hz, Ar–H), 7.09 –
7.19 (m, 3H, Ar–H), 7.26 (t, 2H, J = 9 Hz, Ar–H), 7.39 (d, 5H, J = 3 Hz, Ar–H), 7.62
(d, 1H, J = 6 Hz, Ar–H), 7.73 (t, 2H, J = 9 Hz, Ar–H), 7.80 (d, 2H, J = 6 Hz, Ar–H),
13
8.06 (s, 1H, Ar-H). C NMR (75 MHz, DMSO-d6, ppm) δ 192.84, 173.09, 167.46,
158.76, 136.25, 135.69, 135.07, 134.29, 134.19, 132.37, 130.95, 129.12 (2C), 129.08
(2C), 129.00, 128.97, 128.19, 127.57, 127.18, 123.37, 122.39, 120.97, 120.51, 119.89,
118.74, 111.36, 110.52, 107.48, 69.63, 57.73, 23.92. ESI-HRMS calcd for
C32H24ClN2O4S2 [M + H] +: 599.0860, found: 599.0862.
5.2.22 (Z)-2-(5-((6-((2-Methoxybenzyl)oxy)naphthalen-2-yl)methylene)-4-oxo-2-
thioxothiazolidin-3-yl)-3-(1H-indol-3-yl)propanoic acid (10h)
1
Yield 23%, m.p. 115-116 °C. H NMR (300 MHz, DMSO-d6, ppm) δ 3.51-3.67 (m,
2H, CH2), 3.89 (s, 3H, CH3), 5.34 (s, 2H, CH2), 5.76 (br s, 1H, CH), 6.99-7.16 (m, 2H,
Ar–H), 7.19 (s, 1H, Ar–H), 7.23-7.34 (m, 3H, Ar–H), 7.35-7.53 (m, 3H, Ar–H),
7.56-7.89 (m, 7H, Ar–H), 8.11 (s, 1H, Ar–H). 13C NMR (75 MHz, DMSO-d6, ppm) δ
192.32, 172.75, 166.94, 157.76, 156.98, 143.52, 138.89, 135.74, 132.70, 130.55,
129.75, 128.94, 128.85, 128.76, 128.62, 128.36, 127.78, 127.05, 126.44, 123.65,
122.89, 121.85, 119.75, 118.22, 115.89, 111.53, 110.92, 109.78, 107.12, 67.31, 63.23,
+
56.45, 23.39. ESI-HRMS calcd for C33H26N2O5S2 [M + H] : 595.1355, found:
595.1357.
5.2.23 (Z)-2-(5-((6-((3-Methoxybenzyl)oxy)naphthalen-2-yl)methylene)-4-oxo-2-
thioxothiazolidin-3-yl)-3-(1H-indol-3-yl)propanoic acid (10i)
1
Yield 31%, m.p. 121-122 °C. H NMR (300 MHz, DMSO-d6, ppm) δ 3.53-3.70 (m,
2H, CH2), 3.90 (s, 3H, CH3), 5.36 (s, 2H, CH2), 5.82 (br s, 1H, CH), 6.91-7.09 (m, 2H,
Ar–H), 7.23 (s, 1H, Ar–H), 7.32-7.45 (m, 3H, Ar–H), 7.59-7.72 (m, 3H, Ar–H),
7.81-7.98 (m, 7H, Ar–H), 8.12 (s, 1H, Ar–H). 13C NMR (75 MHz, DMSO-d6, ppm) δ
192.40, 172.65, 166.87, 161.39, 157.69, 143.57, 142.02, 138.97, 135.69, 132.61,

ACCEPTED MANUSCRIPT
130.51, 128.97, 128.58, 128.35, 127.86, 127.19, 126.32, 122.98, 121.79, 120.39,
119.87, 119.26, 118.34, 115.92, 113.45, 111.56, 110.98, 109.72, 107.09, 71.93, 67.28,
+
56.08, 23.92. ESI-HRMS calcd for C33H26N2O5S2 [M + H] : 595.1355, found:
595.1359.
5.2.24 (Z)-2-(5-((6-((4-Methoxybenzyl)oxy)naphthalen-2-yl)methylene)-4-oxo-2-
thioxothiazolidin-3-yl)-3-(1H-indol-3-yl)propanoic acid (10j)
1
Yield 22%, m.p. 104-106 °C. H NMR (300 MHz, DMSO-d6, ppm) δ 3.68-3.80 (m,
2H, CH2), 3.91 (s, 3H, CH3), 5.23 (s, 2H, CH2), 5.78 (br s, 1H, CH), 6.86-7.18 (m, 4H,
Ar–H), 7.20-7.35 (m, 3H, Ar–H), 7.41 (d, 1H, J = 9 Hz, Ar–H), 7.60 (dd, 3H, J = 12,
9 Hz, Ar–H), 7.70 (dd, 3H, J = 9, 3 Hz, Ar–H), 7.78 (d, 2H, J = 9 Hz, Ar–H), 8.01 (s,
1H, Ar–H). 13C NMR (75 MHz, DMSO-d6, ppm) δ 192.38, 172.68, 166.89, 158.95,
157.71, 143.51, 138.93, 135.72, 132.63, 130.48, 130.15 (2C), 129.36, 128.94, 128.53,
128.32, 127.85, 127.39, 126.38, 122.93, 121.98, 119.80, 119.23, 118.29, 115.87,
113.98, 111.55, 111.02, 107.16, 70.92, 67.35, 56.23, 24.04. ESI-HRMS calcd for
C33H26N2O5S2 [M + H] +: 595.1355, found: 595.1357.
5.2.25 (Z)-2-(5-((6-(Benzyloxy)naphthalen-2-yl)methylene)-4-oxo-2-thioxothiazolidin
-3-yl)-3-(1H-indol-3-yl)propanoic acid (10k)
1
Yield 39%, m.p. 116-118 °C. H NMR (300 MHz, DMSO-d6, ppm) δ 3.62-3.75 (m,
2H, CH2), 5.25 (s, 2H, CH2), 5.81 (br s, 1H, CH), 6.90-7.16 (m, 4H, Ar–H), 7.28-7.42
(m, 3H, Ar–H), 7.50 (d, 1H, J = 9 Hz, Ar–H), 7.66 (dd, 3H, J = 12, 9 Hz, Ar–H), 7.81
(dd, 3H, J = 9, 3 Hz, Ar–H), 7.92 (d, 2H, J = 9 Hz, Ar–H), 8.03 (s, 1H, Ar–H). 13C
NMR (75 MHz, DMSO-d6, ppm) δ 192.65, 170.84, 166.48, 157.34, 148.34, 142.63,
138.92, 136.52, 136.18, 133.64, 132.61, 132.41, 131.18, 129.36, 128.54 (2C), 128.33,
127.47, 127.69 (2C), 123.36, 121.78, 120.64, 119.54, 118.86, 118.65, 111.76, 110.54,
107.47, 69.45, 63.38, 24.14. ESI-HRMS calcd for C32H24N2O4S2 [M + H] +: 565.1250,
found: 565.1253.
5.3. Biological assay

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5.3.1. PTP1B and related PTPs biological assay
The enzymatic assays of PTP1B and related PTPs were referred [37-38].
5.3.2. Characterization of the inhibitors’ enzyme kinetics
The enzyme kinetics assay of compound 10b was referred [37-38].
5.3.3. Molecular docking
MOE Dock in MOE v2014.0901 was used for molecular docking simulations of 7c
and 10b and predicting the binding affinity with the PTP1B (PDB code: 2vey). The
2D structures of the molecules were drawn in ChemBioDraw 2014 and converted to
3D in MOE v2014.0901 through energy minimization. And then, the protonation state
of the enzyme and the orientation of the hydrogens were optimized by LigX, at the pH
of 7 and temperature of 300 K. Prior to docking, the force field of AMBER12: EHT
and the implicit solvation model of Reaction Field (R-field) were selected. The
docking workflow followed the “induced fit” protocol, in which the side chains of the
receptor pocket were allowed to move according to ligand conformations, with a
constraint on their positions. The weight used for tethering side chain atoms to their
original positions was 10. All docked poses of molecules were ranked by London dG
scoring first, and then a force field refinement was carried out on the top 30 poses
followed by a rescoring of GBVI/WSA dG.
5.4. Evaluation of antibacterial activity in vitro
In vitro antibacterial activity assay method was referred [27].
Acknowledgments
We wish to thank the National Natural Science Foundation of China (No. 81460524,
81360473 and 81125023) and the Education Department of Jilin Province Scientific
Research Fund Project (2015-50) for financial support. We thank Renee Mosi, PhD,
from Liwen Bianji, Edanz Group China (www.liwenbianji.cn/ac), for editing the
English text of a draft of this manuscript.

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Table 1. Inhibitory activity of compounds 7-10 against PTP1B
Compound
R
IC50 (µM) a
Compound
R
IC50 (µM)
7a
H
4-CH3
4-Br
4-Cl
2,4-Cl2
H
5.52 ± 0.23
5.93 ± 1.23
2.60 ± 0.49
2.44 ± 0.29
3.38 ± 0.64
8.23 ± 0.45
2.70 ± 0.61
5.76 ± 0.82
1.70 ±0.19
5.77 ± 1.05
3.60 ± 0.32
3.18 ± 1.30
4.36 ± 0.81
2.69 ± 0.15
9e
2,4-Cl2
4-CH3
4-Br
0.70 ± 0.28
0.64 ± 0.10
0.36 ± 0.02
0.45 ± 0.15
0.54 ± 0.08
7b
10a
10b
10c
10d
10e
10f
10g
10h
10i
7c
7d
4-F
7e
2,4-Cl2
2-Cl
8a
0.80
0.72
0.72
1.53
1.67
1.45
0.82
±
±
±
±
±
±
±
0.12
0.09
0.02
0.07
0.05
0.19
0.00
8b
4-Br
4-Cl
2,4-Cl2
H
3-Cl
8c
4-Cl
8d
2-OCH3
3-OCH3
4-OCH3
H
9a
9b
4-CH3
4-Br
4-Cl
10j
10k
9c
9d
Oleanolic acidb
a The pNPP assay. IC50 values were determined by regression analyses and expressed as means ±
SD of three replications.
b Positive control.

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Table 2. Inhibitory activity of selected compounds against PTP1B and related PTPs
IC50 (µM) a
Compound
PTP1B
TCPTP
CDC25B
SHP-1
SHP-2
LAR
NA b
10a
10b
0.64 ± 0.10 1.69 ± 0.03 0.83 ± 0.32 3.99 ± 0.21 5.81 ± 0.99
0.36 ± 0.02 0.78 ± 0.03 0.32 ± 0.04 1.11 ± 0.20 3.80 ± 0.20 29.12 ± 1.36
0.45 ± 0.15 1.12 ± 0.00 0.38 ± 0.02 1.52 ± 0.24 4.26 ± 0.20 28.52 ± 0.81
10c
10d
0.54 ± 0.08 1.83 ± 0.17 0.49 ± 0.09 2.95 ± 0.10 6.93 ± 0.06
NA
d
c
Na3VO4
—
—
0.21 ± 0.00 1.25 ± 0.05 2.00 ± 0.32 17.98 ± 4.70
Oleanolic
acid d
2.69 ± 0.15 6.42 ± 0.11
—
—
—
—
aThe pNPP assay. IC50 values were determined by regression analyses and expressed as means ±
SD of three replications.
b Not active at 20 µg/mL concentration.
c Not tested.
d Positive control.

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Table 3. The docking score of molecules binding with PTP1B.
Ligands
Receptor
Docking score (kcal/mol)
7c
PTP1B
PTP1B
-11.29
-13.94
10b

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Table 4. Inhibitory activities (MIC a, µg/mL) of compounds 7-10 against various bacteria.
Gram-positive strains
Gram-negative strains
Fungus
7535 h
Compound
4220 b 209 c
503 d
3289 e
1924 f
2742 g
7a
4
4
8
4
8
8
4
4
8
8
＞64
＞64
＞64
＞64
＞64
＞64
＞64
＞64
＞64
＞64
＞64
＞64
＞64
＞64
＞64
＞64
＞64
＞64
＞64
＞64
＞64
＞64
＞64
＞64
＞64
＞64
1
＞64
＞64
＞64
＞64
＞64
＞64
＞64
＞64
＞64
＞64
＞64
＞64
＞64
＞64
＞64
＞64
＞64
＞64
＞64
＞64
＞64
＞64
＞64
＞64
＞64
＞64
0.50
7b
7c
1
4
2
1
1
7d
2
4
4
2
4
7e
4
4
8
8
8
8a
4
8
8
8
8
8b
2
4
4
4
8
8c
2
2
2
2
2
8d
4
4
8
4
8
9a
8
8
8
8
16
2
9b
2
2
4
2
9c
4
4
4
8
8
9d
2
4
4
4
4
9e
10a
2
2
2
2
2
2
2
2
2
2
10b
2
2
4
8
8
10c
1
4
2
2
2
10d
2
4
4
4
8
i
10e
4
—
—
—
—
—
—
—
—
2
—
—
—
—
—
—
—
4
2
4
10f
4
2
2
10g
2
2
2
10h
8
8
16
16
16
8
10i
8
8
10j
8
8
10k
8
8
4
—
0.25
0.25
—
＞64
2
IV
Gatifloxacin
Moxifloxacin
Fluconazole
0.25
0.25
—
4
2
2
2
1
0.50
—
—
—
—
1
a The antibacterial tests was carried out three times, and the average values were taken as the
MICs.
b Staphylococcus aureus 4220.
c Staphylococcus aureus 209.
d Staphylococcus aureus 503.
e Streptococcus mutans 3289.
f Escherichia coli 1924.
g Pseudomonas aeruginosa 2742.
h Candida albicans 7535.