1-Arylsulfonyl indoline-benzamides as a new antitubulin agents, with inhibition of histone deacetylase

Article abstract of DOI:10.1016/j.ejmech.2018.10.066

We report structure-activity relationships of 1-arylsulfonyl indoline based benzamides. The benzamide (9) exhibits striking tubulin inhibition with an IC₅₀ value of 1.1 μM, better than that of combretastain A-4 (3), and substantial antiproliferative activity against a variety of cancer cells, including MDR-positive cell lines with an IC₅₀ value of 49 nM (KB), 79 nM (A549), 63 nM (MKN45), 64 nM (KB-VIN10), 43 nM (KB-S15), and 46 nM (KB-7D). Dual inhibitory potential of compound 9 was found as it demonstrated significant inhibitory potential against HDAC1, 2 and 6 in comparison to MS-275 (6). Some key interactions of 9 with the amino acid residues of the active site of tubulin and with amino acid residues of HDAC 1 isoform have been figured out by molecular modeling. Compound 9 also demonstrated significant in vivo efficacy in the human non-small cell lung cancer A549 xenograft model as well as B-cell lymphoma BJAB xenograft tumor model.

Full text of DOI:10.1016/j.ejmech.2018.10.066

Accepted Manuscript
1-Arylsulfonyl indoline-benzamides as a new antitubulin agents, with inhibition of
histone deacetylase
Mei-Jung Lai, Ritu Ojha, Mei-Hsiang Lin, Yi-Min Liu, Hsueh-Yun Lee, Tony Eight Lin,
Kai-Cheng Hsu, Chi-Yen Chang, Mei-Chuan Chen, Kunal Nepali, Jang-Yang Chang,
Jing-Ping Liou
PII:
S0223-5234(18)30943-7
DOI:
https://doi.org/10.1016/j.ejmech.2018.10.066
EJMECH 10852
Reference:
To appear in: European Journal of Medicinal Chemistry
Received Date: 15 August 2018
Revised Date: 27 October 2018
Accepted Date: 30 October 2018
Please cite this article as: M.-J. Lai, R. Ojha, M.-H. Lin, Y.-M. Liu, H.-Y. Lee, T.E. Lin, K.-C. Hsu, C.-Y.
Chang, M.-C. Chen, K. Nepali, J.-Y. Chang, J.-P. Liou, 1-Arylsulfonyl indoline-benzamides as a new
antitubulin agents, with inhibition of histone deacetylase, European Journal of Medicinal Chemistry
(2018), doi: https://doi.org/10.1016/j.ejmech.2018.10.066.
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Graphical Abstract
Docking pose of compound 9 in tubulin

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European Journal of Medicinal Chemistry
1
-Arylsulfonyl Indoline-Benzamides as a New Antitubulin Agents, with Inhibition of
Histone Deacetylase
c,1
a,1
a
c
a
d
Mei-Jung Lai, Ritu Ojha, Mei-Hsiang Lin, Yi-Min Liu, Hsueh-Yun Lee, Tony Eight Lin, Kai-
d
f
b
a
*,e,f
Cheng Hsu, Chi-Yen Chang, Mei-Chuan Chen, Kunal Nepali, Jang-Yang Chang, and Jing-Ping
*
,a,c
Liou
*
To whom correspondence should be addressed. Chang J. Y. (Phone) 886-6-2353-535 ext 5002. (e-
mail) jychang@nhri.org.tw.; J. P. Liou: (Phone) 886-2-2736-1661 ext 6130, (e-mail) jpl@tmu.edu.tw
1
Contributed equally to this work.
a. School of Pharmacy, College of Pharmacy, Taipei Medical University, 250 Wuxing Street, Taipei
1
1031, Taiwan.
b. Ph.D. Program for Clinical Drug Development of Chinese Herbal Medicine, College of Pharmacy,
Taipei Medical University.
c. TMU Biomedical Commercialization Center, Taipei Medical University
d. The Ph.D. Program for Cancer Biology and Drug Discovery, College of Medical Science and
Technology, Taipei Medical University, Taipei, Taiwan.
e. Division of hematology/Oncology ,Department of Internal Medicine, National Cheng King
University Hospital, College of Medicine, National Cheng King University , Tainan, Taiwan.
f. National Institute of Cancer Research, National Health Research Institutes, Tainan 704, Taiwan.

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ABSTRACT. We report structure-activity relationships of 1-arylsulfonyl indoline based benzamides.
The benzamide (9) exhibits striking tubulin inhibition with an IC value of 1.1 µM, better than that of
5
0
combretastain A-4 (3), and substantial antiproliferative activity against a variety of cancer cells,
including MDR-positive cell lines with an IC value of 49 nM (KB), 79 nM (A549), 63 nM (MKN45),
5
0
6
4 nM (KB-VIN10), 43 nM (KB-S15), and 46 nM (KB-7D). Dual inhibitory potential of compound 9
was found as it demonstrated significant inhibitory potential against HDAC1, 2 and 6 in comparison to
MS-275 (6). Some key interactions of 9 with the amino acid residues of the active site of tubulin and
with amino acid residues of HDAC 1 isoform have been figured out by molecular modeling. Compound
9
also demonstrated significant in vivo efficacy in the human non-small cell lung cancer A549 xenograft
model as well as B-cell lymphoma BJAB xenograft tumor model.
Keywords: Tubulin, Indoline, benzamide, HDAC, cancer

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1
. Introduction
Chemotherapy is one of the major approaches to cancer treatment. The high risk of toxicity, drug
resistance, and lack of specificity limit the use of traditional cytotoxic agents in the clinic and this, in
turn has triggered the continuing search for new anticancer agents [1]. ABT751 (1), colchicine (2),
combretastatin A-4 (3), combretastatin A-4P (4), and AVE-8062 (5) are some examples of antimitotic
agents (Figure 1) which inhibit not only the dynamics of tubulin polymerization but also serve as
structural scaffolds for novel potent lead anti- cancer molecules [2,3].
Indolines are structurally essential elements of biologically active natural compounds and as they are
widely used as pharmacophores in drug discovery [4-6] they are extremely important in medicinal
chemistry. As a part of our drug discovery program, we are actively involved in the design of indoline-
based antiproliferative agents. In our previous work, 7-aroyl-aminoindoline-sulfonamides were designed
in an effort to rigidify compound 1, an oral antimitotic agent and vascular disrupting agent [6].
Compound 1 binds competitively to the colchicine site of tubulin and inhibits tubulin polymerization
with resultant G2/M arrest and apoptosis. The promising findings of the study motivated us to
investigate indoline based constructs exerting anticancer effects via diverse mechanisms.
Recent reports on the antiproliferative potential of benzamides operating with diverse mechanisms
[7-15] also prompted us to design N-aryl-4-(((1-(arylsulfonyl)-indolin-7-yl)amino)methyl)benzamides
as a new class of antiproliferative agents and this is reported in this study. Literature survey reveals that
tubulin inhibitors have demonstrated significant flexibility towards the inclusion of diverse antitumor
pharmacophores that enables the modulation of multiple targets [16, 17]. Indoline and indole based
scaffolds have been widely employed as structural motifs for the design of tubulin inhibitors as well as
flexible components of HDAC inhibitory model [6, 18-19]. The benzamide group represents a well-
accepted pharmacophore for HDAC inhibition as a zinc binding group that chelates the zinc atom in its
active site [10]. MS-275 (6) [12 - 13] and Chidamide (7) approved by China’s FDA represents HDAC
inhibitor with a benzamide group [20]. Keeping this in view, fusion of 7-aroyl-aminoindoline-

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sulfonamide scaffold with a benzamide functionality appeared to be a rational approach for designing
tubulin inhibitors capable of modulating the activity of HDAC enzymes. To add on, the strategy of
activating tubulin inhibitory framework towards the HDAC inhibition has been attempted previously by
several research groups and optimistic results were attained [17].
Furthermore, number of attempts were made to gain better insight into the SAR of these rigid
benzamide analogs and included: a) the influence of electronic effects was observed by placing a
differently substituted aryl sulfonyl group at the N1 position of the indoline moiety, b) the 7-anilino/7-
aroyl group in previously synthesized analogs was replaced by N- and O-benzyl functionalities, and an
ethyl or vinyl group was introduced as a linker for benzamide functionality to study the influence of
induced flexibility in the designed compounds, c) appropriate comparisons were made by synthesizing
various indole-based target compounds to evaluate the effect of planarity on the antiproliferative activity,
d) regio effects of the benzamide group and comparisons of the designed compounds with the
biarylamine type compound were analyzed.
Thus the present study describes the first exploration of the antiproliferative effects of 1-
(arylsulfonyl)indolines possessing a benzamide group. The synthesized compounds were evaluated for
the in vitro cytotoxic activity against a panel of human cancer cell lines including multidrug resistant
cancer cell lines and also for their effect on tubulin polymerization. The in-vivo efficacy along with
HDAC inhibitory potential of the most potent antiproliferative agent and tubulin polymerization
inhibitor has also been explored.
2
2
. Results and Discussion
.1 Chemistry
In Schemes 1-3 the synthetic routes to the designed compounds (8-28) are shown. Reaction of
compounds 29a-e with various substituted benzenesulfonyl chlorides yielded 5-bromo-1-
benzenesulfonyl)-7-nitroindolines which were subsequently reduced to provide the corresponding
amines. The treatment of the amines (in the case of compounds synthesized from 29a) with tributyltin
(

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hydride (Bu
3
SnH) and 2,2’-azobis-(2-methylpropionitrile) (AIBN) initiated a free radical cascade
leading to the debrominated compounds (30a-30g). Compounds 30a-30k were then subjected to
reductive amination with 3- and 4-carboxybenzaldehyde and NaBH CN to afford the corresponding
3
acids (31a-g and 31i-n). For the synthesis of compound 31h, 30b was treated with 4-
(
methoxycarbonyl)-phenyl)boronic acid in the presence of Cu(OAc) , myristic acid, and 2,6-lutidine
2
followed by LiOH-mediated ester hydrolysis. The resulting carboxylic acids was further reacted with o,
m, p-phenylenediamine and 2-aminophenol using PyBOP as the coupling reagent and triethylamine as
the base, to obtain the desired benzamides (8-25) (Scheme 1). The synthetic route employed in scheme 1
afforded the target benzamides in moderate to good yields (43 - 76 %). Significant variations were
observed in the yields of the benzamides and effect of differently substituted benzene sulfonyl
1
functionality (at N position) along with the different placement of benzamide functionality could not be
ascertained on the reaction yields.
The
synthetic
route
to
N-(2-aminophenyl)-4-(2-(1-((4-methoxyphenyl)sulfonyl)indolin-7-
yl)ethyl)benzamide (26) is shown in Scheme 2. 1H-Indole-7-carboxaldehyde was subjected to a Wittig
reaction with 4-methoxycarbonylbenzyltriphenylphosphonium chloride to yield methyl (E)-4-(2-(1H-
indol-7-yl)vinyl)benzoate. Hydrogenation with Pd/C and subsequent reduction of the indole to an
indoline with sodium cyanoborohydride gave methyl 4-(2-(indolin-7-yl)ethyl)-benzoate (34). The
benzamide (26) was produced from compound 35 by a synthetic route similar to that shown in Scheme
1
and involving sulfonylation, ester hydrolysis, and amidation with o-phenylenediamine. The benzamide
(27) with a vinyl linker, was synthesized in a manner similar to that employed for compound 26 keeping
the double bond at C7 intact. The synthetic route employed afforded the target benzamides in
moderately good yields (65 %). The synthetic route to compound 28 is shown in Scheme 3. tert-
Butyldimethylsilyl-protected 7-hydroxyindoline was treated with 4-methoxybenzenesulfonyl chloride in
pyridine followed by TBAF mediated deprotection of the tert-butyldimethylsilyl group in THF to afford
the 7-hydroxy-1-(4-methoxybenzene-sulfonyl)indoline (40). Compound 40, on reaction with methyl 4-

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2 3
(chloromethyl)-benzoate in the presence of K CO followed by the synthetic strategy employed in
previous schemes yielded the benzamide 28 (Scheme 3). Benzamide 28 was obtained in 52 % yield via
synthetic strategy employed in scheme 3.
2
.2 Biological Evaluation.
The synthesized compounds were evaluated for growth inhibitory effects in vitro against three human
cancer cell lines, oral epidermoid carcinoma (KB cells), stomach carcinoma (MKN45 cells) and lung
adenocarcinoma (A549 cells). Compounds 1, 2, and 6 were used as standards and the results, presented
in Table 1 revealed that the oral epidermoid carcinoma KB cells were the most sensitive to these
synthetic compounds. Compound 9 exhibited significant inhibitory effects against all the cell lines (IC50
=
50 to 80 nM) with the most substantial activity against KB cells (IC = 50 nM). The inhibitory effects
50
of 9 were more potent than those of 1 and 6. A decrease in the inhibitory potential was observed with
compound 8 which bears an unsubstituted phenyl ring at the N1 position as compared to compound 9
possessing a 4-methoxyphenyl ring at the same position. Use of a dimethoxybenzenesulfonyl ring (10)
does not favor the activity and a remarkable decrease in the cytotoxicity was observed with 10 as
compared to compounds 8 and 9. Comparison of the cytotoxic effects of 11 with 9 further confirmed
that the 4-methoxybenzenesulfonyl group, a structural feature in compound 1, is required for significant
activity. Compound 11 with a 3-methoxyphenyl ring at N1 was found to be 12-16 fold less active than
compound 9. A clear dependence of the cell growth inhibitory activity on the electronic factors could
not be established however as compound 12 with a 4-hydroxy substituted benzene sulfonyl group failed
to show any potential comparable to that of compound 9 and was even less active than 13 which bears
chlorine at the para position of the phenyl ring, the N1 position. Beneficial effects of the benzyl amino
functionality at C7 position were clearly demonstrated by the relatively inferior potential of compound
1
6, which possesses the biaryl amino moiety, when compared to compound 9. However, 16 was still
more active than the control compounds 1 and 6 against all the cell lines examined. The significant
potential of compound 9 further supported our attempt to design relatively flexible chemical

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architectures by using N-benzyl functionality in place of the biaryl amine and amide groups at C7 of the
indoline. A drastic decline in the activity was observed when the benzyl amino group was moved from
C7 to the C5 or C6 position of indoline (compare compound 9 with 21 and 22). The influence of
planarity could be easily observed by comparison of the decreased inhibitory effects of 23 and 24,
possessing the indole moiety, with 9 and 22 bearing a non-planar indoline ring. Replacement of –NH-
th
CH functionality (7 position) with –NH-CO- functionality (compound 25) resulted in drastic decline
2
in the cytotoxic potential against A549 cell lines. Compounds with ethylene and vinyl linkers (26 and
2
7) for the N-phenylbenzamide at C7 position were also evaluated. Compound 26 has inhibitory effects
comparable to those of the biaryl amino compound (16). Better activity induced via an ethylene linker in
compound 26 as compared to compound 27 bearing a vinyl linker, further confirmed the favorable trend
observed with increased flexibility at the C7 position. Further insights into the structure activity
relationships were established by the observation of inhibitory potential of 17, 18, and 19. An interesting
revelation was that the shift of the 2-NH group to the 3- or 4-position showed that the amino group at
2
the 4-position was most favorable for the activity as 17 and 18 were found to have reduced activity
compared to 9. The replacement of the amino group with a hydroxy group on N-phenylbenzamide ring
(19) was not useful as the resulting compound displayed complete loss of activity. Compound 20 with a
m-substituted benzamide group showed weak inhibitory activity. Conversion of the connecting linker at
C7 of the indoline from NH-CH to O-CH resulted in inhibitory activities comparable to those of the
2
2
most potent benzamide (compare 9 and 28). The structure activity relationships are presented in Figure
. The efficacy of compounds 9, 16, and 28 against P-gp170/MDR (KB-VIN10 and KB-S15) and MRP-
3
overexpressing (KB-7D) drug-resistant cell lines is shown in Table 2. Benzamides 9, 16, and 28 were
equally effective towards the KB-derived MDR-positive cell lines, even in the presence of high level
expression of drug-resistant efflux protein (MDR-P-gp or MRP) in KB-VIN10, KB-S15, and KB-7D.
Further mechanistic studies were performed as per the previous studies [18, 21, 22, 44] to evaluate
whether the benzamides inhibits the tubulin assembly. The results are presented in Table 3 and Figure 4.

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Among the compounds, compound 9, 16, 17, 26, 27 and 28 induced significant inhibition of tubulin
polymerization with IC50 < 3 µM. Compound 9 exhibited the best tubulin inhibitory activity with an
IC50 value of 1.1 µM (Table 3). The inhibitory effects of compound 9 on the tubulin polymerization
were found to be comparable to CA-4 (3). Overall, the results indicates that the compounds
substoichometrically inhibit tubulin assembly except those inactive. We further compared the binding
affinity potency of compound 9, 16, 28 at concentration of 1 and 5 µM with reference compounds (1, 2
and 3) using the H3-colchicine competition scintillation proximity assay due to their substantial
cytotoxic effects against the cancer cell lines and MDR cell lines along with remarkable inhibition of the
tubulin polymerization.The results indicated all the three compounds 9, 16 and 28 exhibited better
affinity for the colchicine binding site than colchicine itself.
To confirm the inhibition by the HDAC isoforms, the effect of compound 9 and the reference
compound (6) was observed against HDAC isoforms. (Table 4) The benzamide (9) possesses
remarkable inhibitory effects against all the cell lines and significant tubulin polymerization inhibition.
It was also found to possess HDAC inhibitory potential with IC50 values of 0.221 µM (HDAC1) and
0
.662 µM (HDAC2). Compound 9 was 2.5-fold more potent than 6 (IC50 = 0.544 µM) against the
HDAC1 isoform and displayed comparable efficacy against the HDAC2 isoform. The benzamide (9)
also inhibited HDAC6 isoform (IC50 = 0.314 µM), however no inhibition against HDAC6 was
demonstrated by 6. This effect was further shown in western blot analysis where treatment with
compound 9 resulted in upregulation of acetyl α-tubulin levels. (Figure 5) The results confirmed the
dual inhibitory effects of compound 9.
A molecular docking analysis to elucidate interactions between the synthesized compounds and
tubulin was performed. First, to test the accuracy of the docking program, the co-crystallized inhibitor,
colchicine, was docked into the binding site of the tubulin structure (PDB code: 1SA0) using LeadIT
[23]. The docking results produced similar docking conformations to the co-crystallized colchicine (Fig.
S1) indicating that the docking program is reliable. Next, the synthesized compounds were docked into

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the colchicine binding site (Fig. S2). The results of the docking study revealed that the binding site can
be separated into four sites (S1-S4) according to the interacting residues and structure of the most potent
compound 9 (Fig. 6). It was found that the N‐(2‐aminophenyl)‐4‐methylbenzamide moiety occupies
sites S1 and S3 where site S1 contains residues Ser178, Thr179 and Ala180 that produces a hydrophobic
pocket with the aromatic ring of compound 9 (Fig. 6B) and site S3 consists of residues Leu255 and
Asn258 that create hydrophobic contacts with the middle aromatic ring. Hydrogen bonding interaction
were observed with S1 residues Asn101, Thr179, and Glu183. Previous research has identified potential
inhibitors involved in similar hydrogen-bonding interactions with these residues [24-25]. The second
portion of compound 9 (1‐(4‐methoxybenzenesulfonyl)‐2,3‐dihydro‐1H‐indol‐7‐amine) is located at
sites S2 and S4 (Fig. 6B). A hydrogen bond between residue Ser178 of site S2 and the methoxy group is
observed. Site S2 contains residues Leu248, Lys352, Thr353, and Ala354 that create a hydrophobic
pocket occupied by the anisole moiety. The carbon atoms in the side chain of Lys352, the backbone
atoms of Thr353 and hydrophobic residues Leu248 and Ala354 create hydrophobic interactions with
anisole ring (Fig.6B). Site S4 creates another hydrophobic pocket with residues Cys241, Ala250,
Lys254 and Leu255. Thus, the docking analysis of compound 9 reveals hydrogen bonds and
hydrophobic interactions with residues that stabilize the compound within the tubulin binding site.
We further examined the structure activity relationship (SAR) between the colchicine binding site
and the designed compounds using the computational software Forge [26]. Using Forge, we identified
the “average field of actives”, which highlight common features between the active compounds (Fig.
7
A). Forge also produced an activity cliff for each compound using compound 9 as a reference. The
results indicates the impact of structural changes in the structure of the compounds on the activity within
the tubulin binding site. An “activity cliff summary” summarizes the activity cliff data across the
compounds in this study (Fig. 7B).
The average field of actives was produced using the active compound in this study. This model
indicates the electrostatic and hydrophobic interactions of compounds in the binding site. The most

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potent compound in this study, compound 9, was used to describe the model. The model contains a
negative electrostatic field at site S3 (Cyan, Fig. 7A). Residue Asn101 occupies this region and creates a
hydrogen bond with the carbonyl oxygen of compound 9, which acts as a hydrogen bond acceptor. In
contrast, the positive electrostatic potential occupies the S1 site, which consists of residues Ser178,
Thr179 and Ala180 (Red, Fig. 7A). Favorable moieties to occupy this site include an amino group to
serve as a donor for hydrogen bonds. The aromatic rings of compound 9 occupies the hydrophobic areas
within the tubulin binding site (Yellow, Fig. 7A). As a result, compound 9 occupies favorable
electrostatic fields within the tubulin binding site.
The “activity cliff summary” of the tubulin binding site combined the activity cliff obtained from all
pairs of compounds employing compound 9 as the reference (Fig. 7B). Compounds 19, 20, 21 and 22
were selected to further describe the model. At the S1 site, compound 19 (yellow) contains a hydroxyl
group on the ortho position in place of the amino group present in compound 9 (grey). In addition,
compound 19’s terminal phenol ring in the S1 region is rotated roughly 90° when compared to
compound 9 and is located within an unfavorable hydrophobic region. This may be due to the different
characteristics of the aniline and the phenol moiety of compound 9 and 19, respectively (Fig. 7C). In
addition, the hydroxyl group on the terminal ring of compound 19 form a hydrogen bond with residue
Glu183.
Compounds 20, 21 and 22 differ from compound 9 either on the location of the amine linker (at
position 7 in compound 9) or the linkage of the benzene-1,2-diamine (at para position in compound 9).
Compound 20 differs from compound 9 on the linkage of the benzene-1,2-diamine and was found to be
rotated away from favorable hydrophobic regions found in sites S2 and S4. The terminal anisole of
compound 20 is also positioned outside of the favorable hydrophobic region of site S2. Due to this, the
hydrophobic region in S2 repels the secondary amine linker (7-position) and the indole of compound 20
is positioned in an unfavorable hydrophobic region in site S4 (Fig. 7D). Compound 21 contains the
amine linker at the 5 position. Unlike compound 9, the terminal anisole of compound 21 is sandwiched

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by unfavorable hydrophobic fields at site S1. The substitution at the indole (5 position) prevents the
structure from reaching the favorable hydrophobic field. Therefore, the terminal anisole moiety does not
interact with site S2 (Fig. 7E). Finally, compound 22 contains an indole substituted at position 6 by the
amine linker. The amine linker of compound 22 is positioned within a field that is favorable for
hydrophobic interactions in site S4. This slight difference occupies the hydrophobic region and positions
the indole in a more unfavorable region. As a result, the anisole of compound 22 is positioned adjacent
to the favorable hydrophobic region in sites S2 and S4 (Fig. 7F). The “activity cliff summary” details the
structural differences that have resulted in a variable activity profile of the compounds. Our analysis
showed that compound 9 occupies many favorable fields when compared to compounds 19, 20, 21 and
2
2.
To elucidate binding interactions with HDAC, compound 9 was docked into HDAC 1 (PDB ID:
ICN) (Fig. 8A). Compound 9 contains typical features of a HDAC inhibitor – a zinc binding group
5
(ZBG), a linker, and a cap to block the entrance to the binding site [27-28]. The ZBG of compound 9
consists of the N-(2-aminophenyl)formamide moiety. This moiety contains a nitrogen and a carbonyl
oxygen that coordinates the zinc ion in the binding site (Fig. 8A). The nitrogen that links the two
aromatic structures in this moiety forms a hydrogen bond with residue G149. The hydrophobic tunnel is
occupied by the benzyl ring. The benzyl ring creates a pi-pi interaction with the hydrophobic F150
residue. Moreover, compound 9 consists of a 1‐(4‐methoxybenzenesulfonyl)‐2,3‐dihydroindole moiety
that functions as a cap. The cap creates pi-pi stacking interaction with F205 and pi interactions with
Y204 and L271. The interactions formed between the cap and the enzyme surface may increase HDAC
specificity [27-29]. In contrast, HDAC8 has a M274 residue in place of L271 in HDAC1. The HDAC8
isoform is known to form a specific hydrophobic subpocket [30] and compound 9 could not properly
exploit this subpocket. Therefore, the cap construct not only forms less interactions, but is also rotated
away from the surface residues when compared to its docking pose in HDAC1. (Fig. 8B). Moreover,
docking poses of compound 9 in HDAC1 (PDB ID: 5ICN), 2 (PDB ID: 5IX0), and 6 (PDB ID: 5EF8)

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were observed to be similar on superimposition (Fig. 8C). Briefly, the ZBG coordinate the zinc ion and
the linker occupies the hydrophobic tunnel, as seen in HDAC1. The cap also creates similar interactions
with the surface residue leucine (L271, L276, and L712 for HDAC1, 2, and 6, respectively) when
compared to HDAC1 (Fig. 8C). Thus, the interactions with the surface residue leucine observed in
HDAC1, 2 and 6 are important for compound 9 to effectively inhibit HDAC function.
The anticancer evaluation of compound 9 was further extended to in vivo studies in human non-small
cell lung cancer A549 xenograft model as well as BJAB B-cell lymphoma xenograft tumor model. Both
tubulin inhibitors as well as HDAC inhibitors have demonstrated clinical benefits in advanced NSCLC
[31-32] and lymphomas [33-35]. Of particular mention are the implications of US FDA approved
HDAC inhibitors such as SAHA, FK-228, PXD101 and CFDA approved Chidamide (7) in the treatment
of lymphoma [36-40]. To add on, an indoline sulfonamide based HDAC inhibitor previously
synthesised by our research group displayed significant antiproliferative effects against B-cell
lymphoma [41]. These factors collectively led us investigate compound 9 in NSCLC and BJAB
xenograft tumor models. As shown in Fig. 9a, compound 9 caused significant reduction of the tumor
volume in the A549 tumor bearing nude mice (TGI = 62.9 %, 50 mg/kg, ip, qd, **p < 0.01). In addition,
there was no significant change in body weight of tested animals after treatment with compound 9.
Furthermore, in vitro studies on B-cell lymphoma cells (BJAB) revealed that compound 9 was endowed
with potent cellular activity with an IC50 value of 40 nM (results of MTT assay placed in supporting
information). The in vivo evaluation results of compound 9 (intravenous injection) in subcutaneous
xenograft tumor model of BJAB also revealed remarkable inhibition of the tumor growth by benzamide
9
. No significant change in body weight of tested animals was observed in this study also (Fig 10 and
1). Thus the in vivo animal model experiments demonstrated that compound 9 possess significant in
1
vivo potential against NSCLC and B-Cell lymphoma. The in vivo evaluation of compound 9 in oral
epidermoid carcinoma (KB cells) is under progress.
3
. Conclusion

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A series of 1-arylsulfonyl indoline-based benzamides has been synthesized and evaluated against a
panel of human cancer cell lines. The results of in vitro cytotoxicity studies indicated that compound 9,
1
6 and 28 were found to have promising antiproliferative activity with compound 9 displaying striking
cell killing effects against the KB cells with an IC value of 48 nM. Among them, compounds 9 and 28
5
0
were found to be more potent tubulin polymerization inhibitors with an IC50 of 1.1 and 1.9 µM,
respectively, than the reference compounds 2 and 3. The highlight of the present study demonstrated by
these compounds was the unaltered ability to inhibit the KB-derived MDR-positive cell lines, even in
the presence of high level expression of drug-resistant efflux protein (MDR-P-gp or MRP) in KB-
VIN10, KB-S15, and KB-7D cells. The most potent antiproliferative compound (9) possessing the best
tubulin polymerization inhibitory activity also inhibited HDAC 1, 2 and 6 confirming its dual inhibitory
potential. Some of the important interactions of the active compounds with the amino acid residues of
tubulin and HDAC 1 isoform have been figured out by molecular modeling which supports the results
of the in vitro assays. Compound 9 also demonstrated promising significant in vivo efficacy in the
human non-small cell lung cancer A549 xenograft model as well as xenograft tumor model of BJAB.
These promising findings clearly indicate the need for detailed preclinical and clinical investigation as
these scaffold could emerge as templates for further development of potent anticancer agents.
4
. Experimental Section
A) Chemistry. Nuclear magnetic resonance ( H and C NMR) spectra were obtained with Bruker
Fourier 300 and DRX-500 NMR spectrometers, and are reported as chemical shifts in parts per million
ppm, δ) downfield from TMS as an internal standard. High-resolution mass spectra (HRMS) were
1
13
(
(
recorded with a Finnigan MAT 95S mass spectrometer. The purities of the final compounds were
determined using a Hitachi 2000 series HPLC system using an Agilent Zorbax Eclipse XDB-C18 column
(5 µm, 4.6 mm × 150 mm) with the solvent system consisting of acetonitrile (mobile phase A) and water
containing 0.1% formic acid and 10 mmol NH OAc (mobile phase B), and were found to be ≥95%.
4

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Flash column chromatography was performed using silica gel (Merck Kieselgel 60, no. 9385, 230−400
mesh ASTM). All reactions were carried out under an atmosphere of dry nitrogen.
N-(2-Aminophenyl)-4-(((1-(phenylsulfonyl)indolin-7-yl)amino)methyl)benzamide (8)
To a solution of 30a (0.21 g, 0.51 mmol), PyBOP (0.26 g, 0.51 mmol), triethylamine (0.16 ml, 1.15
mmol) in DMF 1.5 mL, benzene-1,2-diamine (0.05 g, 0.48 mmol) was added and stirred at room
temperature. After being stirred for 2 h, the reaction was quenched with water, followed by extraction
4
with EtOAc (15 mL × 3). The combined organic layer was dried over anhydrous MgSO and
concentrated under reduced pressure. The residue was purified by silica gel chromatography (EtOAc) to
1
give 8 as a brown solid in 71% yield; t
R
= 43.65. mp: 192-193 °C. H NMR (300 MHz, DMSO-d
6
) : δ
2
.09 (t, J = 7.5 Hz, 2H), 4.01 (t, J = 7.2 Hz, 2H), 4.54 (d, J = 4.8 Hz, 2H), 6.31 (t, J = 5.7 Hz, 1H), 6.36
d, J = 6.9 Hz, 1H), 6.47 (d, J = 8.1 Hz, 1H), 6.64 (m, 1H), 6.82 (dd, J = 1.5 and 8.1 Hz, 1H), 6.91 (t, J
7.8 Hz, 1H), 7.00 (m, 2H), 7.20 (dd, J = 1.2 and 7.8 Hz, 1H), 7.50 - 7.74 (m, 7H), 8.01 (d, J = 8.1 Hz,
(
=
1
3
2
1
1
H). C NMR (75 MHz, CDCl ) δ 29.01, 46.71, 53.72, 111.01, 113.14, 116.79, 117.03, 124.05, 126.93,
3
27.17, 127.37, 127.45, 127.86, 128.51, 128.55, 129.75, 133.72, 134.41, 136.47, 138.86, 141.49,
＋
26 4 3
43.19, 144.04, 165.67. HRMS (ESI) for C28H N O S (M ): calcd, 498.1726; found, 498.1723.
N-(2-Aminophenyl)-4-(((1-((4-methoxyphenyl) sulfonyl)indolin-7-yl)amino)methyl)benzamide (9)
To a solution of 30b (0.22 g, 0.51 mmol), PyBOP (0.27 g, 0.51 mmol), Et N (0.17 mL, 1.15 mmol)
3
in DMF (1.5 mL), benzene-1,2-diamine (0.05 g, 48 mmol) was added and stirred at room temperature.
After being stirred for 2h, the reaction was quenched with H2O, followed by extraction with EtOAc (15
mL × 3). The combined organic layer was dried over anhydrous MgSO4 and concentrated under
reduced pressure. The residue was purified by silica gel chromatography (EtOAc) to give 9 as a white
1
solid (0.18 g, 67% yield); tR = 43.65 min. mp: 171-172 °C. H NMR (300 MHz, DMSO-d ) δ 2.14 (t, J
6
=
7.2 Hz, 2H), 3.82 (s, 3H), 3.98 (t, J = 7.5 Hz, 2H), 4.53 (d, J = 6.0 Hz, 2H), 4.91 (s, 2H), 6.28 (t, J =
.0 Hz, 1H), 6.39 (d, J = 6.6 Hz, 2H), 6.44 (d, J = 8.1 Hz, 1H), 6.61 (m, 1H), 6.79 (dd, J = 1.5 and 8.1
Hz, 1H), 6.91 (t, J = 7.8 Hz, 1H), 6.98 (m, 1H), 7.05 (d, J = 6.9 Hz, 2H), 7.18 (dd, J = 1.2 and 7.8 Hz,
6

ACCEPTED MANUSCRIPT
1
3
1
H), 7.55 (d, J = 9.0 Hz, 2H), 7.59 (d, J = 8.4 Hz, 2H), 7.99 (d, J = 8.4 Hz, 2H), 9.61 (s, 1H). C NMR
(
75 MHz, DMSO-d ) δ 26.55, 44.14, 51.04, 53.56, 108.34, 110.58, 112.27, 114.05, 121.29, 124.51,
6
1
25.85, 127.45, 131.14, 136.30, 138.94, 140.93, 141.41, 161.17, 163.08. HRMS (ESI) for C H N O S
29 29 4 4
+
(
M + H ): calcd, 529.1910; found, 529.1905.
N-(2-Aminophenyl)-4-(((1-((3,4-dimethoxyphenyl)sulfonyl)indolin-7-yl)amino)methyl)benzamide
(10)
The title compound 10 was obtained as a white solid in 50% yield in a similar manner as described
1
R 6
for the preparation of 9; t = 33.78 min. mp: 199-200 °C. H NMR (300 MHz, DMSO-d ) δ 2.09 (t, J =
7
6
2
1
.2 Hz, 2H), 3.43 (s, 3H), 3.83 (s, 3H), 3.96 (t, J = 7.5 Hz, 2H), 4.54 (d, J = 6.3 Hz, 2H), 4.90 (s, 2H),
.31 (t, J = 6.3 Hz, 1H), 6.39 (d, J = 6.6 Hz, 1H), 6.46 (d, J = 8.1 Hz, 1H), 6.61 (m, 1H), 6.76 – 6.81 (m,
H), 6.90 (d, J = 7.5 Hz, 1H), 6.95 - 7.01 (m, 1H), 7.11 (d, J = 8.7 Hz, 1H), 7.18 (dd, J = 1.2 and 7.5 Hz,
H), 7.34 (dd, J = 2.1 and 8.4 Hz, 1H), 7.61 (d, J = 8.4 Hz, 2H), 7.99 (d, J = 8.1 Hz, 2H), 9.61 (s, 1H).
1
3
C NMR (125 MHz, CDCl ) δ 26.53, 44.68, 50.87, 53.24, 53.50, 107.29, 107.85, 108.14, 110.61,
3
1
1
15.71, 117.11, 118.81, 121.96, 122.66, 124.57, 124.83, 125.06, 125.18, 125.49, 125.59, 130.31,
＋
36.10, 138.09, 138.74, 141.37, 145.87, 150.57, 163.14. HRMS (ESI) for C30
58.1937; found, 558.1934.
30 4 5
H N O S (M ): calcd,
5
N-(2-Aminophenyl)-4-(((1-((3-methoxyphenyl)sulfonyl)indolin-7-yl)amino)methyl)benzamide (11)
The title compound 11 was obtained as a light brown crystalline solid in 52% yield in a similar
1
manner as described for the preparation of 9; t = 36.12 min. mp: 102-103 °C. H NMR (500 MHz,
R
CDCl ) δ 2.13 (t, J = 7.5 Hz, 2H), 3.56 (s, 3H), 4.00 (t, J = 7.5 Hz, 2H), 4.54 (d, J = 5.5 Hz, 2H), 6.27 (t,
3
J = 6.0 Hz, 1H), 6.39 (d, J = 7.5 Hz, 1H), 6.44 (d, J = 8.0 Hz, 1H), 6.83 - 6.86 (m, 2H), 6.91 - 6.96 (m,
2
2
1
H), 7.07 -7.11 (m, 2H), 7.26 - 7.35 (m, 4H), 7.60 (d, J = 8.0 Hz, 2H), 7.81 (s, 1H), 7.89 (d, J = 8.0 Hz,
1
3
H). C NMR (75 MHz, CDCl
3
) δ 26.46, 44.65, 50.99, 52.85, 108.24, 109.03, 110.61, 115.33, 116.62,
17.12, 117.91, 121.71, 123.20, 124.59, 124.65, 125.22, 125.60, 127.41, 130.25, 134.58, 136.00,

ACCEPTED MANUSCRIPT
+
1
5
38.62, 138.69, 141.09, 156.79, 163.46, 168.67. HRMS (ESI) for C29
29.1910; found, 529.1912.
29 4 4
H N O S (M + H ): calcd,
N-(2-Aminophenyl)-4-(((1-((4-hydroxyphenyl)sulfonyl)indolin-7-yl)amino)methyl)benzamide (12)
To a solution of 9 (0.1 g, 0.19 mmol) in dichloromethane (10 mL), bromotribromide (0.024 mL, 0.57
mmol) was added dropwise and the reaction mixture was stirred at room temperature for overnight. The
mixture was quenched with water and extracted with dichloromethane. The combined organic layer was
4
dried over anhydrous MgSO and concentrated under reduced pressure to afford a residue which was
R
purified by silica gel chromatography (EtOAc) to give 12 as a white solid in 76% yield; t = 30.95 min.
1
mp: 197-198 °C, H NMR (300 MHz, DMSO-d
6
): δ 2.14 (t, J = 7.2 Hz, 2H), 3.94 (t, J = 7.5 Hz, 2H),
4
.52 (d, J = 6.0 Hz, 2H), 4.92 (s, 2H), 6.27 (t, J = 6.3 Hz, 1H), 6.39 (d, J = 7.2 Hz, 1H), 6.44 (d, J = 8.1
Hz, 1H), 6.62 (m, 1H), 6.79 - 6.84 (m, 3H), 6.90 (t, J = 7.8 Hz, 1H), 6.96 – 7.01 (m, 1H), 7.18 (m, 1H),
7
.43 (d, J = 9.0 Hz, 2H), 7.59 (d, J = 8.1 Hz, 2H), 8.00 (d, J = 8.4 Hz, 2H), 9.64 (s, 1H), 10.87 (bs, 1H);
C NMR (300 MHz, CD OD + DMSO-d ): δ 30.15, 48.58, 54.85, 112.32, 114.52, 117.37, 117.67,
1
3
3
6
1
1
18.57, 119.30, 125.30, 127.98, 128.59, 128.69, 128.91, 129.35, 129.62, 131.90, 132.03, 134.17,
＋
34.58, 140.04, 142.81, 144.16, 145.60, 168.47, 168.93. HRMS (ESI) for C28
14.1675; found, 514.1678.
26 4 4
H N O S (M ): calcd,
5
N-(2-Aminophenyl)-4-(((1-((4-chlorophenyl)sulfonyl)indolin-7-yl)amino)methyl)benzamide (13)
The title compound 13 was obtained as a white solid in 56% yield in a similar manner as described
1
for the preparation of 9; t = 38.21 min. mp: 101-102 °C. H NMR (500 MHz, CD OD) δ 2.17 (t, J =
R
3
7
.5 Hz, 2H), 4.01 (t, J = 7.2 Hz, 2H), 4.53 (d, J = 6.0 Hz, 2H), 4.91 (s, 2H), 6.24 (t, J = 6.3 Hz, 1H)
.40 - 6.48 (m, 2H), 6.61 (m, 1H), 6.79 (dd, J = 1.5 Hz and 8.1 Hz, 1H), 6.93 - 6.98 (m, 2H), 7.18 (dd,
6
13
J = 1.2 and 7.8 Hz, 1H), 7.56 - 7.69 (m, 5H), 7.98 (d, J = 8.0 Hz, 2H), 9.63 (s, 1H). C NMR (125
MHz, CDCl ) δ 26.49, 44.70, 50.95, 108.32, 110.71, 115.51, 116.85, 121.80, 122.97, 124.59, 124.74,
25.20, 125.38, 125.83, 126.35, 126.60, 130.31, 132.39, 135.60, 137.48, 137.52, 138.39, 138.69,
3
1
1
＋
41.07, 141.31, 163.78. HRMS (ESI) for C H ClN O S (M ): calcd, 532.1336; found, 532.1339.
2
8
25
4
3

ACCEPTED MANUSCRIPT
N-(2-Aminophenyl)-4-(((1-((4-fluorophenyl)sulfonyl)indolin-7-yl)amino)methyl)benzamide (14)
The title compound 14 was obtained as a white solid in 49% yield in a similar manner as described
1
for the preparation of 9; t = 32.32 min. mp: 199-200 °C. H NMR (300 MHz, DMSO-d ) δ 2.15 (t, J =
R
6
7
6
.2 Hz, 2H), 4.02 (t, J = 7.2 Hz, 2H), 4.54 (d, J = 6.00 Hz, 2H), 4.92 (s, 2H), 6.25 (t, J = 6.0 Hz, 1H),
.40 (d, J = 7.2 Hz, 1H), 6.47 (d, J = 7.8 Hz, 1H), 6.61 (m, 1H), 6.80 (dd, J = 1.2, 7.8 Hz, 1H), 6.91 (d, J
=
7.8 Hz, 1 H), 6.96 (m, 1H), 7.19 (dd, J = 1.2, 7.8 Hz, 1H), 7.40 - 7.43 (m, 2H), 7.60 (d, J = 8.1 Hz,
1
3
2
4
1
3
H), 7.67 - 7.70 (m, 2H), 8.00 (d, J = 8.1 Hz, 2H), 9.61 (s, 1H). C NMR (75 MHz, CDCl ) δ 26.47,
4.71, 50.89, 108.28, 110.65, 113.47, 113.77, 115.58, 116.96, 121.86, 122.85, 124.59, 124.79, 124.89,
＋
25.16, 129.86, 129.90, 135.64, 138.28, 138.74, 141.16, 163.26. HRMS (ESI) for C H FN O S (M +
2
8
26
4
3
H): calcd, 517.1710, found, 517.1708.
N-(2-Aminophenyl)-4-(((1-((4-cyanophenyl)sulfonyl)indolin-7-yl)amino)methyl)benzamide (15)
The title compound 15 was obtained as a pale yellow solid in 59% yield in a similar manner as
1
described for the preparation of 9; t = 43.32 min. mp: 113-114 °C. H NMR (300 MHz, DMSO-d ) δ
R
6
2
.16 (t, J = 7.2 Hz, 2H), 4.06 (t, J = 7.2 Hz, 2H), 4.54 (d, J = 6.0 Hz, 2H), 4.92 (s, 2H), 6.25 (d, J = 6.0
Hz, 1H), 6.39 (d, J = 6.9 Hz, 1 H), 6.48 (d, J = 8.1 Hz, 1H), 6.61 (m, 1H), 6.79 (dd, J = 1.2 and 7.8 Hz,
H), 6.91 - 6.98 (m, 2H), 7.18 (dd, J = 1.2 and 7.8 Hz, 1H), 7.59 (d, J = 8.1 Hz, 2H), 7.81 (d, J = 8.4
1
1
3
Hz, 2 H), 7.98 - 8.05 (m, 4H), 9.63 (s, 1H). C NMR (75 MHz, CDCl
3
) δ 26.57, 44.71, 51.04, 108.46,
1
10.77, 114.44, 115.65, 117.02, 121.87, 122.79, 124.17, 124.61, 124.81, 125.19, 125.56, 126.15,
+
1
29.96, 130.43, 135.35, 138.05, 138.61, 140.99, 163.17. HRMS (ESI) for C H N O S (M + H ):
29 26 5 3
calcd, 524.1756; found, 524.1758.
N-(2-Aminophenyl)-4-((1-((4-methoxyphenyl)sulfonyl)indolin-7-yl)amino)benzamide (16)
The title compound 16 was obtained as a brown solid in 51% yield in a similar manner as described
1
for the preparation of 9; t
R
= 34.89 min. mp: 100-101 °C. H NMR (500 MHz, CDCl
3
): δ 2.23 (t, J = 7.5
Hz, 2H), 3.81 (s, 3H), 4.02 (t, J = 7.5 Hz, 2H), 6.70 (d, J = 7.0 Hz, 1H), 6.84 - 6.86 (m, 4 H), 7.08 - 7.14
m, 4H), 7.31 (d, J = 7.5 Hz, 1H), 7.37 (d, J = 8.0 Hz, 2H), 7.52 (d, J = 9.0 Hz, 1H), 7.73 (s, 1H), 7.81
(

ACCEPTED MANUSCRIPT
1
3
(
d, J = 8.5 Hz, 2 H), 8.33 (s, 1H). C NMR (75 MHz, CD
3
OD + DMSO-d
6
): δ 48.91, 55.72, 95.46,
1
09.59, 112.50, 114.74, 117.02, 117.51, 121.41, 121.92, 123.23, 123.89, 126.92, 127.21, 127.28,
+
1
28.30, 129.12, 133.23, 136.55, 143.24, 144.87, 146.73, 164.02. HRMS (ESI) for C H N O S (M ):
28 26 4 4
calcd, 514.1675; found, 514.1677.
N-(3-Aminophenyl)-4-(((1-((4-methoxyphenyl)sulfonyl)indolin-7-yl)amino)methyl)benzamide (17)
The title compound 17 was obtained as a brown solid in 51% yield in a similar manner as described
1
for the preparation of 9; t
R
= 34.86 min. mp: 171-172 °C. H NMR (300 MHz, DMSO-d
6
) δ 2.14 (t, J =
7
.2 Hz, 2H), 3.82 (s, 3H), 3.97 (t, J = 7.2 Hz, 2H), 4.52 (d, J = 6.00 Hz, 2H), 5.08 (s, 2H), 6.30 - 6.34
(
(
m, 2H), 6.38 - 6.46 (m, 2H), 6.85 - 6.99 (m, 3H), 7.05 (d, J = 9.0 Hz, 2H), 7.12 (t, J = 1.8 Hz, 1H), 7.55
1
3
d, J = 8.7 Hz, 2H), 7.59 (d, J = 8.1 Hz, 2H), 7.93 (d, J = 8.4 Hz, 2H), 9.91 (s, 1H). C NMR (75 MHz,
CDCl5 ) δ 26.57, 44.67, 50.85, 52.98, 104.28, 107.53, 108.15, 108.63, 110.56, 111.48, 125.34, 125.41,
3
1
25.50, 131.26, 135.86, 136.46, 138.80, 141.11, 144.65, 160.93, 163.15. HRMS (ESI) for C H N O S
29 29 4 4
+
(
M + H ): calcd, 529.1910; found, 529.1905.
N-(4-Aminophenyl)-4-(((1-((4-methoxyphenyl)sulfonyl)indolin-7-yl)amino)methyl)benzamide (18)
The title compound 18 was obtained as a brown solid in 47% yield in a similar manner as described
1
for the preparation of 9; t
R
= 32.62 min. mp: 101-102 °C. H NMR (500 MHz, CDCl
3
) δ 2.14 (t, J = 7.5
Hz, 2H), 3.82 (s, 3H), 4.12 (t, J = 7.5 Hz, 2H), 4.51 (d, J = 6.0 Hz, 2H), 6.26 (t, J = 6.0 Hz, 1H), 6.37 (d,
J = 7.5 Hz, 1H), 6.42 (d, J = 8.0 Hz, 1H), 6.69 (d, J = 8.5 Hz, 2H), 6.83 (d, J = 8.5 Hz, 2H), 6.91 (t, J =
7
7
1
1
5
.5 Hz, 1H), 7.39 (d, J = 8.0 Hz, 2 H), 7.50 (d, J = 8.0 Hz, 2H), 7.57 (d, J = 8.5 Hz, 2H), 7.65 (s, 1H),
1
3
.83 (d, J = 8.0 Hz, 2H). C NMR (75 MHz, CDCl ) δ 26.57, 44.69, 50.85, 52.98, 108.16, 110.53,
3
11.48, 112.80, 119.77, 124.73, 124.79, 125.32, 125.50, 126.76, 127.04, 131.30, 135.84, 138.82,
+
40.80, 140.84, 160.93, 163.05. HRMS (ESI) for C29
29.1907.
H
28
N
4
O
4
S (M + H ): calcd, 529.1910; found,
N-(2-Hydroxyphenyl)-4-(((1-((4-methoxyphenyl)sulfonyl)indolin-7-yl)methyl)amino)benzamide
19)
(

ACCEPTED MANUSCRIPT
The title compound 19 was obtained with 2-aminophenol as a yellow solid in 49% yield in a similar
1
manner as described for the preparation of 9; t = 43.42 min. mp: 75-76 °C. H NMR (500 MHz,
R
CDCl ) δ 2.15 (t, J = 7.5 Hz, 2H), 3.83 (s, 3H), 4.00 (t, J = 7.5 Hz, 2H), 4.57 (d, J = 6.0 Hz, 2H), 6.33 (t,
3
J = 6.0 Hz, 1H), 6.38-6.41 (m, 2H), 6.84 (d, J = 9.0 Hz, 2H), 6.92 (t, J = 7.5 Hz, 1H), 7.38 (dd, J = 1.5,
8
1
1
1
.0 Hz, 1H), 7.43 (m, 1H), 7.52 (d, J = 8.5 Hz, 2H), 7.63 (d, J = 8.5 Hz, 2H), 7.70 (m, 1H), 8.13 (dd, J =
1
3
.5, 8.0 Hz, 1H), 8.18 (d, J = 8.0 Hz, 2H). C NMR (75 MHz, CDCl ) δ 26.59, 44.77, 50.87, 52.99,
3
08.16, 110.65, 111.48, 122.81, 123.19, 123.96, 124.51, 124.71, 125.40, 125.48, 125.52, 127.07,
28 4 4
28.30, 132.05, 135.89, 138.67, 139.40, 141.76, 144.19, 160.93, 161.64. HRMS (ESI) for C29H N O S
+
(
M ): calcd, 529.1671; found, 529.1675.
N-(2-Aminophenyl)-3-(((1-((4-methoxyphenyl)sulfonyl)indolin-7 yl)amino)methyl)benzamide (20)
The title compound 20 was obtained as a white solid in 43% yield in a similar manner as described
1
for the preparation of 9; t = 35.73 min. mp: 99-100 °C. H NMR (500 MHz, CDCl ) : δ 2.12 (t, J = 7.0
R
3
Hz, 2H), 3.77 (s, 3H), 3.96 (t, J = 7.0 Hz, 2H), 4.55 (d, J = 6.5 Hz, 2H), 6.18 (t, J = 6.0 Hz, 1H), 6.37 (d,
J = 7.5 Hz, 1H), 6.45 (d, J = 8.5 Hz, 1H), 6.67 - 6.75 (m, 4H), 6.92 - 6.99 (m, 2H), 7.17 (d, J = 7.5 Hz,
1
8
1
1
H), 7.44 (d, J = 8.5 Hz, 2H), 7.49 (t, J = 8.0 Hz, 1H), 7.62 (d, J = 7.5 Hz, 1H), 7.88 (d, J = 8.0 Hz, 1H),
1
3
3
.00 (s, 1H), 8.12 (s, 1H). C NMR (75 MHz, CDCl ): δ 29.23, 47.33, 53.51, 55.64, 107.07, 110.34,
10.81, 111.39, 113.22, 114.15, 127.44, 127.99, 129.75, 133.90, 138.52, 139.13, 141.46, 143.76,
+
63.59, 165.81. HRMS (ESI) for C H N O S (M + H ): calcd, 529.1910; found, 529.1912.
2
9
29
4
4
N-(2-Aminophenyl)-4-(((1-(4-methoxyphenylsulfonyl)indolin-5-yl)amino)methyl)benzamide (21)
The title compound 21 was obtained as a brown solid in 68% yield in a similar manner as described
1
for the preparation of 9; t = 32.33 min. mp: 173-174 °C. H NMR (300 MHz, DMSO-d ) : δ 2.59 (t, J =
R
6
8
.1 Hz, 2H), 3.77 (t, J = 8.7 Hz, 2H), 3.80 (s, 3H), 4.31 (d, J = 5.7 Hz, 2H), 4.89 (s, 2H), 6.27 (t, J = 6.0
Hz, 1H), 6.37 (d, J = 3.1 Hz, 1H), 6.46 (dd, J = 2.1 and 8.7 Hz, 1H), 6.61 (m, 1H), 6.79 (dd, J = 1.2 and
8
7
.1 Hz, 1H), 6.95 - 7.05 (m, 3H), 7.16 - 7.23 (m, 2H), 7.46 (d, J = 8.4 Hz, 1H), 7.61 (d, J = 9.0 Hz, 2H),
1
3
.93 (d, J = 8.1 Hz, 2H), 9.63 (s, 1H). C NMR (75 MHz, CD OD + DMSO-d ): δ 28.84, 53.39, 55.16,
3
6

ACCEPTED MANUSCRIPT
1
1
10.83, 113.13, 114.12, 117.18, 117.97, 123.90, 126.58, 127.19, 127.31, 127.95, 128.01, 129.72,
＋
33.12, 138.77, 141.43, 142.73, 144.22, 163.95, 167.04. HRMS (EI) for C H N O S (M ): calcd,
29
28
4
4
5
28.1831; found, 528.1829.
N-(2-Aminophenyl)-4-(((1-(4-methoxyphenylsulfonyl)indolin-6-yl)amino)methyl)benzamide (22)
The title compound 22 was obtained as a brown solid in 65% yield in a similar manner as described
1
for the preparation of 9; t = 38.08 min. mp: 173-174 °C. H NMR (300 MHz, DMSO-d ) δ 2.72 (t, J =
R
6
8
2
7
.4 Hz, 2H), 3.75 (t, J = 8.4 Hz, 2H), 3.79 (s, 3H), 4.38 (d, J = 6.3 Hz, 2H), 4.88 (s, 2H), 6.25 (dd, J =
.4 and 8.4 Hz, 1H), 6.56 - 6.64 (m, 2H), 6.77 - 6.81 (m, 3H), 6.96-6.99 (m, 3H), 7.16 (dd, J = 1.2 and
1
3
.8 Hz, 1H), 7.47 (d, J = 9.0 Hz, 2H), 7.51 (d, J = 8.4 Hz, 2H), 8.02 (d, J = 8.1 Hz, 2H). C NMR (75
OD + DMSO-d ): δ 29.94, 47.49, 57.02, 111.77, 114.08, 115.71, 116.14, 116.33, 117.27,
17.52, 117.68, 117.79, 127.29, 128.51, 128.76, 129.86, 130.76, 130.88, 132.02, 135.72, 136.68,
MHz, CD
3
6
1
1
＋
39.76, 142.28, 145.46, 150.69, 150.91, 164.59, 166.59. HRMS (EI) for C H N O S (M ): calcd,
29
28
4
4
5
28.1831; found, 528.1832.
N-(2-Aminophenyl)-4-(((1-((4-methoxyphenyl)sulfonyl)-1H-indol-7-yl)amino)methyl)benzamide
(23)
The title compound 23 was obtained as brown solid in 62% yield in a similar manner as described for
1
R 6
the preparation of 9; t = 36.87 min. mp: 169-170 °C. H NMR (300 MHz, DMSO-d ) δ 3.81 (s, 3H),
4
7
.57 (d, J = 5.4 Hz, 2H), 4.92 (s, 2H), 6.49 (d, J = 7.5 Hz, 1H), 6.62 (m, 1H), 6.80 - 6.81 (m, 3H), 6.93-
.10 (m, 4H), 7.20 (d, J = 1.2 Hz, 1H), 7.48 (d, J = 8.4 Hz, 2H), 7.69 – 7.72 (m, 3H), 7.99 (d, J = 8.1
＋
Hz, 2H), 9.66 (s, 1H). HRMS (EI) for C H N O S (M ): calcd, 526.1675; found, 526.1677.
2
9
26
4
4
N-(2-Aminophenyl)-4-(((1-((4-methoxyphenyl)sulfonyl)-1H-indol-6-yl)amino)methyl)benzamide
24)
(
The title compound 24 was obtained as a brown solid in 58% yield in a similar manner as described for
1
the preparation of 9; t = 43.62 min. mp: 179-180 °C. H NMR (300 MHz, DMSO-d ) δ 3.75 (s, 3H),
R
6
4
.45 (d, J = 5.7 Hz, 2H), 4.90 (s, 2H), 6.55 – 6.82 (m, 5H), 6.92 - 7.01 (m, 4H), 7.17 (dd, J = 1.2 and 6.6

ACCEPTED MANUSCRIPT
Hz, 1H), 7.24 (d, J = 8.4 Hz, 2H), 7.36 (d, J = 6.6 Hz, 2H), 7.48 (d, J = 9.0 Hz, 2H), 7.56 (d, J = 8.4 Hz,
1
3
2
1
1
H), 8.05 (d, J = 8.1 Hz, 2H). C NMR (75 MHz, CDCl ): δ 46.97, 56.24, 95.42, 110.02, 112.55,
3
15.14, 116.68, 116.80, 121.07, 122.11, 123.46, 123.84, 127.00, 127.17, 127.28, 128.51, 128.92,
＋
29.25, 133.49, 136.51, 143.64, 144.56, 146.93, 163.91, 165.69. HRMS (EI) for C H N O S (M ):
2
9
26
4
4
calcd, 526.1675; found, 526.1677.
1
4
N -(2-aminophenyl)-N -(1-((4-methoxyphenyl)sulfonyl)indolin-7-yl)terephthalamide (25)
The title compound 25 was obtained as a yellow solid in 58% yield in a similar manner as described for
1
the preparation of 9. H NMR (300 MHz, DMSO-d
6
) : δ 2.31 (t, J = 6.6 Hz, 2H), 3.83 (s, 3H), 4.08 (t, J
=
7.5 Hz, 2H), 4.98 (s, 2H), 6.62 (t, J = 7.2 Hz, 1H), 6.80 (dd, J = 1.2 and 6.9 Hz, 1H), 6.98 – 7.06 (m,
4
8
5
1
H), 7.20 - 7.27 (m, 2H), 7.57 (d, J = 8.7 Hz, 2H), 7.99 (d, J = 8.1 Hz, 1H), 8.10 (d, J = 8.4 Hz, 2H),
1
3
.17 (d, J = 8.4 Hz, 2H), 9.82 (s, 1H), 10.24 (s, 1H). C NMR (75 MHz, DMSO-d ) : δ 29.05, 53.69,
6
6.28, 115.09, 116.58, 116.70, 121.84, 122.80, 123.45, 127.29, 127.61, 127.87, 128.76, 130.16, 131.16,
33.28, 137.24, 138.17, 139.30, 143.73, 164.06, 164.53, 165.18.
N-(2-Aminophenyl)-4-(2-(1-((4-methoxyphenyl)sulfonyl)indolin-7-yl)ethyl)benzamide (26)
Compound 37 (1 g, 2.2 mmol) was dissolved in dioxane (15 mL) and 1M LiOH(aq) (12 mL) was added
to the solution. The reaction mixture was stirred at 40 ℃for 3h and then was evaporated under reduced
pressure. The reaction mixture was quenched with water and the pH was adjusted with 3N HCl(aq). The
reaction mixture was extracted with ethyl acetate (50 mL x 3). The combined organic layer was dried
over anhydrous MgSO , concentrated under reduced pressure to give acid. The obtained acid was
4
subjected to the amide formation in a similar manner to give 25 as described for the preparation of 8 as a
1
brown solid in 65% yield; t = 36.87 min. mp: 157-158 °C. H NMR (500 MHz, CDCl ) δ 2.11 (t, J =
R
3
7
.5 Hz, 2H), 3.09 (t, J = 7.5 Hz, 2H), 3.37 (t, J = 7.5 Hz, 2H), 3.82 (s, 3H), 3.88 (t, J = 8.0 Hz, 2H), 6.80
(d, J = 8.5 Hz, 2H), 6.88 (d, J = 7.5 Hz, 1H), 7.07 (t, J = 7.5 Hz, 2H), 7.15 (d, J = 7.5 Hz, 1H), 7.30 (d, J
1
3
=
3
8.0 Hz, 1H), 7.39 (d, J = 8.5 Hz, 2H), 7.92 (d, J = 8.0 Hz, 2H). C NMR (75 MHz, CDCl ) δ 26.26,
3
1.33, 33.34, 50.08, 52.96, 111.35, 115.57, 116.94, 119.89, 122.02, 122.79, 124.41, 124.47, 124.73,

ACCEPTED MANUSCRIPT
1
26.31, 126.35, 127.10, 128.97, 133.06, 135.31, 138.25, 138.74, 143.98, 160.74, 163.44. HRMS (ESI)
+
C H N O S (M + H ): calcd, 528.1957; found, 528.1956.
30
29
3
4
(E)-N-(2-Aminophenyl)-4-(2-(1-((4-methoxyphenyl)sulfonyl)indolin-7-yl)vinyl)benzamide (27)
The title compound 27 was obtained as a brown solid in 65% yield in a similar manner as described
1
for the preparation of 26; t = 43.62 min. mp: 121-122 °C. H NMR (500 MHz, CDCl ) δ 2.22 (t, J =
R
3
7
.5 Hz, 2H), 3.82 (s, 3H), 4.04 (t, J = 7.5 Hz, 2H), 6.81 (d, J = 9.0 Hz, 2H), 6.85 - 6.87 (m, 2H), 6.98 (d,
J = 7.5 Hz, 1H), 7.08 - 7.11 (m, 1H), 7.16 - 7.19 (m, 2H), 7.36 (d, J = 8.0 Hz, 1H), 7.41 (d, J = 9.0 Hz,
＋
2
H), 7.67 - 7.71 (m, 3H), 7.85 - 7.90 (m, 3H). HRMS (EI) for C H N O S (M - H ): calcd, 524.1644;
30 28 3 4
found,. 524.1648.
N-(2-Aminophenyl)-4-(((1-((4-methoxyphenyl)sulfonyl)indolin-7-yl)oxy)methyl)benzamide (28)
The title compound 28 was synthesized as a light brown solid in 52% yield using compound 41 in a
1
similar manner as that described for the synthesis of 9; t = 33.81 min. mp: 179-180 °C. H NMR (300
R
MHz,DMSO-d ) δ 2.37 (t, J = 7.5 Hz, 2H), 3.82 (s, 3H), 4.01 (t, J = 7.5 Hz, 2H), 4.93 (s, 2H), 5.27 (s,
6
2
H), 6.62 (t, J = 6.6 Hz, 1H), 6.80 (d, J = 8.1 Hz, 2H), 6.96 - 7.02 (m, 4H), 7.10 (t, J = 7.8 Hz, 2H), 7.20
(d, J = 6.9 Hz, 1H), 7.55 (d, J = 9.0 Hz, 2H), 7.66 (d, J = 8.1 Hz, 2H), 8.01 (d, J = 8.4 Hz, 2H), 9.67 (s,
1
3
1
1
3
H). C NMR (75 MHz, CDCl ): δ 26.70, 50.29, 52.91, 67.54, 111.25, 111.28, 124.73, 124.99, 125.05,
28 3 5
26.81, 127.21, 128.59, 130.71, 136.42, 138.45, 148.30, 160.55. HRMS (ESI) for C29H N O S (M +
+
H ): calcd, 530.1750; found, 530.1752.
1
-(Phenylsulfonyl)indolin-7-amine (30a)
The title compound 30a was obtained in 80% yield from compound 29a in a manner similar to that
1
described for the synthesis of compound 30b. H NMR (500 MHz, CDCl ) δ 2.11 (t, J = 7.5 Hz, 2H),
3
3
7
1
.98 (t, J = 7.5 Hz, 2H), 4.71 (s, 2H), 6.39 - 6.41 (m, 1H), 6.59 (d, J = 7.5 Hz, 1H), 6.89 - 6.92 (m, 1H),
.36 - 7.39 (m, 2H), 7.54-7.57 (m, 1H), 7.61-7.64 (m, 2H).
-((4-Methoxyphenyl)sulfonyl)indolin-7-amine (30b).

ACCEPTED MANUSCRIPT
A mixture of 5-bromo-7-nitro-indoline (29a) (2.18 g, 9.0 mmol) and 4-methoxybenzene-1-sulfonyl
chloride (1.85 g, 9.0 mmol) in pyridine (6 mL) was refluxed overnight. After cooling, the reaction
mixture was quenched with H O and extracted with CH Cl . The combined organic layer was dried over
2
2
2
anhydrous MgSO , then concentrated under reduced pressure. The residue was purified by silica gel
4
chromatography (EtOAc: n-hexane = 1: 4) to give a yellow solid (3.47 g, 93% yield). The solid residue
was dissolved in i-PrOH (45 mL) and H O (5 mL). To this solution, Fe powder (2.31 g, 41.5 mmol) and
2
ammonium chloride (887 mg, 16.6 mmol) was added and the reaction mixture was refluxed for 6 h. The
progress of the reaction was monitored by TLC. Upon completion of the reaction, the reaction was
2
filtered over celite. The filtrate was concentrated and H O was added. Extraction was with EtOAc (50
mL x 3). The combined organic layer was dried over anhydrous MgSO and concentrated under reduced
4
pressure to yield the amine (2.73 g, 85% yield). The mixture of the obtained residue,
azobisisobutyronitrile (1.19 g, 7.3 mmol) and tributyltin hydride (6.43 g, 22.1 mmol) were refluxed in
toluene (30 mL) overnight. After cooling, the reaction mixture was quenched with H O and extracted
2
2 2 4
with CH Cl . The combined organic layer was dried over anhydrous MgSO , concentrated under
reduced pressure and dried under vacuum. The residue was purified by silica gel chromatography
1
(
EtOAc: n-hexane = 1: 1) to give 30b (1.82 g, 84% yield). HNMR (500 MHz, CDCl
3
) δ 2.15 (t, J = 7.5
Hz, 2H), 3.82 (s, 3H), 3.95 (t, J = 8.0 Hz, 2H), 4.70 (s, 2H, D O exchangeable protons), 6.41 (d, J = 7.5
2
Hz, 1H), 6.58 (d, J = 8.0 Hz, 1H), 6.81-6.84 (m, 2H), 6.89 - 6.92 (m, 1H), 7.52 -7.55 (m, 2H).
1
-((3,4-Dimethoxyphenyl)sulfonyl)indolin-7-amine (30c)
The title compound 30c was obtained in 78% yield from compound 29a in a manner similar to that
1
described for the synthesis of compound 30b. H NMR (500MHz, CDCl ) δ 2.16 (t, J = 7.5 Hz, 2H),
3
3
7
1
2
.60 (s, 3H), 3.91 (s, 3H), 3.95 (t, J = 7.5 Hz, 2H), 4.39 (s, 2H, D O exchangeable protons), 6.54 (d, J =
.5 Hz, 1H), 6.82 - 6.87 (m, 3H), 6.95 - 6.96 (m, 1H), 7.37 (dd, J = 2.0, 8.0 Hz, 1H).
-((3-Methoxyphenyl)sulfonyl)indolin-7-amine (30d)

ACCEPTED MANUSCRIPT
The title compound 30d was obtained in 73% yield from compound 29a in a manner similar to that
1
described for the synthesis of compound 30b. H NMR (500 MHz, CDCl ): δ 2.13 (t, J = 7.5 Hz, 2H),
3
3
6
1
.60 (s, 3H), 3.96 (t, J = 7.5 Hz, 2H), 4.70 (s, 2H, D O exchangeable protons), 6.42 (d, J = 7.0 Hz, 1H),
2
.60 (d, J = 8.0 Hz, 1H), 6.91 - 6.97 (m, 3H), 7.07 (m, 1H), 7.42 (m, 1H).
-((4-Chlorophenyl)sulfonyl)indolin-7-amine (30e)
The title compound 30e was obtained in 71% yield from compound 29a in a manner similar to that
1
described for the synthesis of compound 30b. H NMR (500 MHz, CDCl
3
) δ 2.23 (t, J = 7.5 Hz, 2H),
3
8
1
2
.99 (t, J = 7.5 Hz, 2H), 4.49 (s, 2H, D O exchangeable protons), 6.54 (d, J = 7.0 Hz, 1H), 6.80 (d, J =
.0 Hz, 1H), 6.96 - 6.99 (m, 1H), 7.35 (d, J = 8.5 Hz, 2H), 7.56 (d, J = 8.5 Hz, 2H).
-((4-Fluorophenyl)sulfonyl)indolin-7-amine (30f)
The title compound 30f was obtained in 72% yield from compound 29a in a manner similar to that
1
described for the synthesis of compound 30b. H NMR (500 MHz, CDCl ) δ 2.16 (t, J = 7.5 Hz, 2H),
3
3
8
4
.99 (t, J = 7.5 Hz, 2H), 4.69 (s, 2H, D O exchangeable protons), 6.42 (d, J = 7.5 Hz, 1H), 6.60 (d, J =
2
.0 Hz, 1H), 6.90 (t, J = 7.5 Hz, 1H), 7.05 (m, 2H) 7.63 (m, 2H).
-((7-Aminoindolin-1-yl)sulfonyl)benzonitrile (30g)
The title compound 30g was obtained in 75% yield from compound 29a in a manner similar to that
1
described for the synthesis of compound 30b. H NMR (500 MHz, CDCl ) δ 2.22 (t, J = 7.5 Hz, 2H),
3
4
8
1
.09 (t, J = 7.5 Hz, 2H), 4.23 (s, 2H, D O exchangeable protons), 6.43 (d, J = 7.5 Hz, 1H), 6.54 (d, J =
2
.0 Hz, 1H), 6.97 (t, J = 7.5 Hz, 1H), 7.89 (d, J = 8.5 Hz, 2H), 8.03 (d, J = 8.0 Hz, 2H).
-((4-Methoxyphenyl)sulfonyl)indolin-5-amine (30h)
The mixture of 5-nitro-indoline (29b) (1 g, 6.09 mmol) and benzenesulfonyl chloride (1.07 g, 6.09
mmol) in pyridine (3 mL) was refluxed overnight. After cooling, the reaction mixture was quenched
with water and extracted with dichloromethane. The combined organic layer was dried over anhydrous
4
MgSO and concentrated under reduced pressure. The residue obtained was purified by silica gel
chromatography (EtOAc: n-hexane = 1: 4). A mixture of the obtained solid (1.83 g, 5.47 mmol), iron

ACCEPTED MANUSCRIPT
powder (611 mg, 10.9 mmol), ammonium chloride (1.46 g, 27.3 mmol) in isopropanol (45 mL) and
water (5 mL) was refluxed for 6 h. The progress of the reaction was monitored on TLC. On completion
of the reaction, the reaction was filtered over celite. The filtrate was concentrated and water was added
to it. Extraction was done with ethyl acetate (50 mL x 3). The combined organic layer was dried over
anhydrous MgSO and concentrated under reduced pressure. The resulting residue was purified by silica
4
.
1
gel chromatography (EtOAc: n-hexane = 1: 1) to give compound 30h in 75% yield H NMR (500 MHz,
CDCl ) δ 2.66 (t, J = 8.5 Hz, 2H), 3.81 (s, 3H), 3.85 (t, J = 8.5 Hz, 2H), 6.49 (s, 1H), 6.59 (d, J = 8.5
Hz, 1H), 6.85 (d, J = 9.0 Hz, 2H), 7.45 (d, J = 8.5 Hz, 1H), 7.62-7.64 (m, 2H).
3
1
-((4-Methoxyphenyl)sulfonyl)indolin-6-amine (30i)
The title compound 30i was obtained in 71% yield from compound 29c in a manner similar to that
1
described for the preparation of 30h. H NMR (500 MHz, DMSO-d ) δ 2.61 (t, J = 8.0 Hz, 2H), 3.73 (t,
6
J = 8.0 Hz, 2H), 3.81 (s, 3H), 6.74 (d, J = 8.0 Hz, 1H), 6.93 (d, J = 8.00 Hz, 2H), 7.12 (s, 1H), 7.41 (d, J
=
7.5 Hz, 1H), 7.67 (d, J = 8.5 Hz, 2H).
1
-((4-Methoxyphenyl)sulfonyl)-1H-indol-7-amine (30j)
The title compound 30j was obtained in 71% yield from compound 29d in a manner similar to that
1
described for the preparation of 30h. H NMR (500 MHz, CDCl
3
) δ 3.79 (s, 3H), 6.61 (d, J = 4.5 Hz,
1
7
1
H), 6.85 - 6.89 (m, 3H), 7.01 (d, J = 7.5 Hz, 1H), 7.08 - 7.11 (m, 1H), 7.55 (d, J = 4.5 Hz, 1H), 7.79 -
.81 (d, J = 7.5 Hz, 2H).
-((4-Methoxyphenyl)sulfonyl)-1H-indol-6-amine (30k)
The title compound 30k was obtained in 69% yield from compound 29e in a manner similar to that
1
described for the preparation of 29i. H NMR (500 MHz, CDCl ) δ 3.80 (s, 3H), 6.51 (d, J = 3.0 Hz,
3
1
H), 6.74 (dd, J = 1.5, 8.5 Hz, 1H), 6.86 (d, J = 8.5 Hz, 2H), 7.29 (d, J = 8.5 Hz, 1H), 7.36 (d, J = 3.0
Hz, 1H), 7.48 (s, 1H), 7.79 (d, J = 8.5 Hz, 2H).
-(((1-(Phenylsulfonyl)indolin-7-yl)amino)methyl)benzoic acid (31a)
4

ACCEPTED MANUSCRIPT
The title compound 31a was obtained in 66% yield from compound 30a in a manner similar to that
1
described for the preparation of 31b. H NMR (500 MHz, DMSO-d ) δ 2.15 (t, J = 7.5 Hz, 2H), 3.99 (t,
6
J = 7.5 Hz, 2H), 4.61 (d, J = 6.0 Hz, 2H), 6.25 - 6.29 (m, 1H), 6.39 (d, J = 7.5 Hz, 1H), 6.45 (d, J = 8.5
Hz, 1H), 6.89 - 6.93 (m, 1H), 7.23-739 (m, 2H), 7.54 - 7.59 (m, 2H), 7.62 (d, J = 8.0 Hz, 2H), 7.98 (d, J
=
8.5 Hz, 2H).
4
-(((1-((4-Methoxyphenyl)sulfonyl)indolin-7-yl)amino)-methyl)benzoic acid (31b)
To a stirred solution of 30b (1.80 g, 5.94 mmol) and 4-carboxybenzaldehyde (0.885 g, 5.94 mmol) in
MeOH (10 mL), few drops of glacial AcOH were added. Sodium cyanoborohydride (560 mg, 8.92
mmol) was added and the reaction mixture was stirred at room temperature overnight. The reaction was
quenched with H O then extracted with EtOAc. The combined organic layer was dried over anhydrous
2
MgSO and concentrated under reduced pressure. The residue was purified by silica gel chromatography
4
1
(
EtOAc: n-hexane = 1: 0.95 with 0.5% AcOH) to give compound 31b (1.76 g, yield 68%). H NMR
6
(500 MHz, DMSO-d ) δ 2.10 (t, J = 7.5 Hz, 2H), 3.79 (s, 3H), 3.93 (t, J = 7.5 Hz, 2H), 4.48 (d, J = 6.0
Hz, 2H), 6.35 (d, J = 7.5 Hz, 1H), 6.38 (d, J = 8.5 Hz, 1H), 6.85 - 6.88 (m, 1H), 7.00 (d, J = 9 Hz, 2H),
7
.49 - 7.51 (m, 2H), 7.54 (d, J = 8.0 Hz, 2H), 7.91 (d, J = 8.5 Hz, 2H).
4
-(((1-((3,4-Dimethoxyphenyl)sulfonyl)indolin-7-yl)amino)methyl)benzoic acid (31c)
The title compound 31c was obtained in 71% yield from compound 30c in a manner similar to that
1
described for the preparation of 31b. H NMR (500 MHz, CD OD) δ 2.10 (t, J = 7.5 Hz, 2H), 3.36 (s,
3
3
1
1
4
H), 3.85 (s, 3H), 3.96 (t, J = 7.5 Hz, 2H), 4.53 (s, 2H), 6.36 (d, J = 7.5 Hz, 1H), 6.48 (d, J = 7.5 Hz,
H), 6.70 (d, J = 2.0 Hz, 1H), 6.88-6.91 (m, 1H), 7.02 (d, J = 7.5 Hz, 1H), 7.38 (dd, J = 2.0, 7.5 Hz,
H), 7.60 (d, J = 8.5 Hz, 2H), 7.97 (d, J = 8.5 Hz, 2H).
-(((1-((3-Methoxyphenyl)sulfonyl)indolin-7-yl)amino)methyl)benzoic acid (31d)
The title compound 31d was obtained in 73% yield from compound 30d in a manner similar to that
1
described for the preparation of 31b. H NMR (500 MHz, DMSO-d ) δ 2.08 (t, J = 7.5 Hz, 2H), 3.53 (s,
6

ACCEPTED MANUSCRIPT
3
6
8
4
H), 3.96 (t, J = 7.5 Hz, 2H), 4.50 (d, J = 6.0 Hz, 2H), 6.34 (d, J = 7.5 Hz, 1H), 6.40 (d, J = 8.5 Hz, 1H),
.86 - 6.89 (m, 2H), 7.21 - 7.25 (m, 2H), 7.43 (t, J = 8.0 Hz, 1H), 7.60 (d, J = 8.5 Hz, 2H), 7.96 (d, J =
.5 Hz, 2H).
-(((1-((4-Chlorophenyl)sulfonyl)indolin-7-yl)amino)methyl)benzoic acid (31e)
The title compound 31e was obtained in 62% yield from compound 30e in a manner similar to that
1
described for the preparation of 31b. H NMR (500 MHz, DMSO-d ) δ 2.13 (t, J = 7.5 Hz, 2H), 3.98 (t,
6
J = 7.5 Hz, 2H), 4.48 (d, J = 6.0 Hz, 2H), 6.36 (d, J = 7.0 Hz, 1H), 6.40 (d, J = 8.5 Hz, 1H), 6.87 - 6.90
(m, 1H), 7.53 (d, J = 8.0 Hz, 2H), 7.55 - 7.57 (m, 4H), 7.91 (d, J = 8.0 Hz, 2H).
4
-(((1-((4-Fluorophenyl)sulfonyl)indolin-7-yl)amino)methyl)benzoic acid (31f)
The title compound 31f was obtained in 64% from compound 30f in a manner similar to that
1
described for the preparation of 31b. H NMR (500 MHz, DMSO-d ) δ 2.12 (t, J = 7.5 Hz, 2H), 3.98 (t,
6
J = 7.5 Hz, 2H), 4.49 (d, J = 6.0 Hz, 2H), 6.36 (d, J = 7.0 Hz, 1H), 6.40 (d, J = 8.5 Hz, 1H), 6.89 (m,
1
H), 7.36 (t, J = 9.0 Hz, 2H), 7.54 (d, J = 8.5 Hz, 2H), 7.64 - 7.67 (m, 2H), 7.93 (d, J = 8.0 Hz, 2H).
4
-(((1-((4-Cyanophenyl)sulfonyl)indolin-7-yl)amino)methyl)benzoic acid (31g)
The title compound 31g was obtained in 65% from compound 30g in a manner similar to that
1
described for the preparation of 31b. H NMR (500 MHz, DMSO-d
6
) δ 2.22 (t, J = 7.5 Hz, 2H), 4.09 (t,
J = 7.5 Hz, 2H), 4.23 (d, J = 6.0 Hz, 2H), 6.43 (d, J = 7.5 Hz, 1H), 6.53 (d, J = 8.0 Hz, 1H), 6.97 (t, J =
.0 Hz, 1H), 7.60 (d, J = 8.0 Hz, 2H), 7.81 (d, J = 8.0 Hz, 2H), 7.89 (d, J = 8.0 Hz, 2H), 8.03 (d, J = 8.0
Hz, 2H).
8
4
-((1-((4-Methoxyphenyl)sulfonyl)indolin-7-yl)amino)benzoic acid (31h)
To a flame-dried 100 mL pear shaped flask was added (4-(methoxycarbonyl)phenyl)boronic acid
(
0.18 g, 1.0 mmol), 20 mol % of solid Cu(OAc)
2
(0.03 g, 0.16 mmol), crystalline myristic acid (0.75 g,
0
.33 mmol) and dry toluene (5 mL).The resulting suspension was stirred for 10 min and 2,6-lutidine
(0.20 mL, 1.73 mmol) was added by syringe. After 30 min, 30b (0.20 g, 0.66 mmol) was added and
stirred vigorously at ambient temperature for 24 h. The reaction mixture was filtered and washed by

ACCEPTED MANUSCRIPT
2 2 4
CH Cl and EtOAc. The organic layers were dried over MgSO , concentrated and purified by flash
column chromatography (EtOAc: n-hexane = 1: 3) to give the residue. 1M LiOH(aq) (0.37 mL) was
added to the solution of the residue (0.08 g, 0.18 mmol) in dioxane (5 mL) and the reaction mixture was
o
heated to 40 C for 18 h. After cooling to room temperature, the reaction mixture was neutralized to pH
=
7, extracted with water (10 mL) and chloroform/IPA (chloroform/ IPA = 3/1; 10mL x 3). The organic
layers were dried over MgSO and the residue was recrystallized from methanol to afford 31h (0.07 g,
4
1
9
7
7
3
6%) as a white solid. H NMR (500 MHz, CDCl
3
) δ 2.26 (t, J = 7.0 Hz, 2H), 3.83 (s, 3H), 4.02 (t, J =
.5 Hz, 2H), 6.75 (d, J = 7.0 Hz, 1H), 6.93 (d, J = 9.0 Hz, 2H), 7.01 (d, J = 8.5 Hz, 2H), 7.11 (m, 1H),
.34 (d, J = 8.5 Hz, 1H), 7.50 (d, J = 9.0 Hz, 2H), 7.79 (br, 1H), 7.88 (d, J = 9.0 Hz, 2H).
-(((1-((4-Methoxyphenyl)sulfonyl)indolin-7-yl)amino)methyl)benzoic acid (31i)
The title compound 31i was obtained in 69% yield from compound 30b using 3-formylbenzoic acid
1
in a manner similar to that described for the preparation of 31b. H NMR (500 MHz, CD OD) δ 2.18 (t,
3
J = 7.0 Hz, 2H), 3.88 (s, 3H), 3.99 (t, J = 7.0 Hz, 2H), 4.58 (d, J = 6.5 Hz, 2H), 6.19 (t, J = 6.0 Hz, 1H),
6
.39 (d, J = 7.5 Hz, 1H), 6.48 (d, J = 8.5 Hz, 1H), 6.69 - 6.79 (m, 2H), 7.21 (d, J = 7.5 Hz, 1H), 7.57 (d,
J = 8.5 Hz, 2H), 7.59 (t, J = 8.0 Hz, 1 H), 7.69 (d, J = 7.5 Hz, 1H), 7.91 (d, J = 8.0 Hz, 1H), 8.17 (s,
1
H).
4
-((1-((4-methoxyphenyl)sulfonyl)indolin-7-yl)carbamoyl)benzoic acid (31j)
To a solution of 30b (1 g, 3.28 mmol) in dry pyridine (5 mL), methyl 4-(chlorocarbonyl)benzoate
(0.652 g, 3.28 mmol) was added. The reaction mixture was stirred at room temperature for 2h. The
reaction was then quenched with H O and extracted with EtOAc (3 x 50 mL). The combined organic
2
layer was dried over anhydrous MgSO and concentrated under reduced pressure. The mixture was
4
purified by silica gel chromatography (EtOAc: n-hexane = 1: 1) to give residue (1.52 g, 65 % yield).
The resulting residue was dissolved in dioxane and 10 mL LiOH (aq, 1M) was added to it. The reaction
mixture was stirred for 5 h at 40 ⁰ C . The reaction was concentrated under reduced pressure and then
was added water. The mixture was acidified with 3 N HCl and extracted with ethyl acetate. The