Design, synthesis and biological evaluation of novel thiazole-based derivatives as human Pin1 inhibitors

Article abstract of DOI:10.1016/j.bmc.2020.115878

Pin1 is a peptidyl prolyl cis-trans isomerase (PPIase) and inhibiting Pin1 is a potential way for discovering anti-tumor agents. With an aim to find potent Pin1 inhibitors with a novel scaffold, a series of thiazole derivatives with an alicyclic heterocycles on the 2-position were designed, synthesized and tested against human Pin1. Compound 9p bearing a 2-oxa-6-azaspiro [3,3] heptane moiety on the thiazole scaffold was identified as the most potent Pin1 inhibitor of this series with an IC₅₀ value of 0.95 μM. The structure-activity relationship (SAR) and molecular modeling study indicated that introducing an alicyclic ring with an H-bond acceptor would be a viable way to improve the binding affinity.

Full text of DOI:10.1016/j.bmc.2020.115878

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Design, synthesis and biological evaluation of novel thiazole-based deriva-
tives as human Pin1 inhibitors
Lifei Du, Xiaoyu Wang, Guonan Cui, Bailing Xu
PII:
S0968-0896(20)30708-2
https://doi.org/10.1016/j.bmc.2020.115878
BMC 115878
DOI:
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11 November 2020
Please cite this article as: L. Du, X. Wang, G. Cui, B. Xu, Design, synthesis and biological evaluation of novel
thiazole-based derivatives as human Pin1 inhibitors, Bioorganic & Medicinal Chemistry (2020), doi: https://
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Design, synthesis and biological evaluation of novel thiazole-based
derivatives as human Pin1 inhibitors
Lifei Du, Xiaoyu Wang, Guonan Cui, Bailing Xu*
Beijing Key Laboratory of Active Substance Discovery and Druggability Evaluation, Institute of
Materia Medica, Chinese Academy of Medical Sciences＆Peking Union Medical College,
Beijing, 100050, China
Abstract
Pin1 is a peptidyl prolyl cis-trans isomerase (PPIase) and inhibiting Pin1 is a
potential way for discovering anti-tumor agents. With an aim to find potent Pin1
inhibitors with a novel scaffold, a series of thiazole derivatives with an alicyclic
heterocycles on the 2-position were designed, synthesized and tested against human
Pin1. Compound 9p bearing a 2-oxa-6-azaspiro [3,3] heptane moiety on the thiazole
scaffold was identified as the most potent Pin1 inhibitor of this series with an IC value
5
0
of 0.95 μM. The structure-activity relationship (SAR) and molecular modeling study
indicated that introducing an alicyclic ring with an H-bond acceptor would be a viable
way to improve the binding affinity.
Keywords: PPIase, Pin1, thiazole derivatives, Pin1 inhibitor
1
. Introduction
Proline-directed phosphorylation on Ser or Thr is a common regulation mechanism
in diverse physiological processes. Pin1(protein interacting with NIMA1) is the only
enzyme, functioning as a conformational switch, specifically isomerizing
phosphorylated Ser/Thr-Pro peptide bonds in a variety of substrate proteins,1-5 and its
dysregulation is implicated in a number of diseases, including cancer and Alzheimer
disease.6, 7 It was observed that Pin1 is overexpressed in various human cancer cells
including breast, prostate, colon, cervical cancer cells.8, 9 It was reported that Pin1
*
Corresponding author. Bailing Xu, Tel: +86 10 63166764
E-mail address: xubl@imm.ac.cn (B. Xu).

activates numerous oncogenic proteins or growth-promoting proteins and also
1
0
inactivates multiple tumor suppressors or growth-inhibitory proteins. Thus, inhibiting
Pin1 is expected to be a potential strategy for the treatment of cancer.
During the past more than two decades, many efforts have been made on the
discovery of Pin1 inhibitors and a number of structurally distinct small molecules have
been reported, and these inhibitors can be divided into non-covalent inhibitors and
covalent inhibitors.11-47 The research on non-covalent inhibitors was represented by
Pfizer, which developed several structurally novel Pin1 inhibitors using structure
guided design strategies.34-36 In 2009, Pfizer disclosed a phosphate-containing
derivative, which was the most potent Pin1 inhibitor to date (Fig. 1 compound A, K =
i
0
.006 μM). However, its membrane permeability was supposed to be poor and there
was no activity at the cellular level due to the presence of a phosphate group. Therefore,
Pfizer designed novel derivatives containing carboxylic acid moiety instead of the
3
4
phosphate group in the subsequent structural optimization. In 2010, Compound B was
reported to be active against Pin1 (Fig. 1 K = 0.89 μM), however, it was still not active
i
3
5
at the cellular level. In 2014, Pfizer took advantage of scaffold hopping approach and
designed compound C, which was active at both enzymatic and cellular levels (Fig. 1,
3
6
K = 0.58 μM，HT29 cells IC = 1.9 μM). Obviously, it is challenging to develop
i
50
drug-like Pin1 inhibitors with high potency. Interestingly, all-trans retinoic acid (Fig.
, ATRA) is a drug for the treatment of acute promyelocytic leukemia (APL). In 2015,
1
Kunping Lu’s team utilized a mechanism-based high-throughput screening method and
found that ATRA can directly bind to Pin1, inhibiting Pin1 with a K value of 0.82
i
3
7
μM. This result further indicated that Pin1 was involved in cancer development and
was a potential anticancer target.
Recently, the research on Pin1 covalent inhibitors was gradually increasing. Juglone
(Fig. 1) was the first covalent binding inhibitor of Pin1, and it irreversibly inhibited
3
8
Pin1. The SH of Cys113 in the catalytic domain can undergo a Michael addition with
Juglone, which led to the partial opening of the binding cavity and inhibited Pin1, but

3
9
Juglone is a non-specific inhibitor. Different from Juglone, KPT-6566, reported in
4
0
2
016, is a specific covalent inhibitor of Pin1 (Fig. 1, IC = 0.64 μM). The results of
50
molecular docking showed that the electrophilic thioglycolic acid fragment was close
to Cys113, forming a disulfide bond with Cys113, and releasing the quinone structure
which induced apoptosis of tumor cells by alkylation of DNA. These two mechanisms
could synergistically kill the tumor cells. In 2019, Ieda et. al successfully engineered
the covalent binding inhibitor E of Pin1 by introducing a 2-naphthalene acryl group to
4
1
the lead compound D and this inhibitor could covalently bind to Cys113. Based on a
highly potent peptide inhibitor, D-peptide (Ac-Phe-D-phosThr-Pip-Nal-Gln-NH2; K =
i
2
0 nM), a selective and potent covalent inhibitor BJP-06-005-3 without a phosphate
group was achieved by using α-chloroacetamide fragment as an electrophile to interact
with Cys113. However, it had poor mouse liver microsome stability. 42 Although
various chemical entities were disclosed as Pin1 inhibitors, discovering of drug-like
new Pin1 inhibitors with distinct scaffolds is still in quest for the development of
potential anti-cancer drugs.
O
S
HN
O
O
HN
O
O
O
F
P
F
O
HO OH
OH
Juglone IC = 5 µM
A. K = 0.006 µM
B. K = 0.89 µM
5
0
i
i
F
O
S
COOH
Cl
HN
COOH
O
N
N
S
O
Cl
S
COOH
C. K = 0.58 µM
ATRA K = 0.82 µM
KPT6566 IC = 0.64 µM
50
i
i
IC = 1.9 µM ( HT29 cells)
5
0

H
N
O
O
O
O
O
Cl
O
N
N
N
O
O
H
O
O
N
N
N
N
O
H
O
S
N
N
H
COOH
H
COOH
NH
NH2
O
D. IC = 5.4 µM
E. IC = 3.2 µM
BJP-06-005-3 K = 48 nM
i
5
0
50
Figure 1. Chemical structures of several reported Pin1 inhibitors
The crystal structures of some small molecule inhibitors complexed with Pin1
revealed that the binding site of catalytic domain consisted of three sub-pockets, namely,
a prolyl pocket, a slightly shallow hydrophobic shelf and a unique phosphate binding
pocket.43-46 The prolyl site is a characteristic region of Pin1 and was exploited by
numerous inhibitors. In our previous work, a series of thiazole-based inhibitors (Figure
4
7
2
. structure I) were constructed and showed micromolar inhibitory activity. In order
to improve the potency and drug-like properties of the thiazole derivatives, we
tentatively used the alicyclic moieties to replace the aromatic ring on the 2-position to
fully exploit the prolyl pocket, while employing the acetic acid substituted for oxalic
acid for creating hydrogen binding within phosphate binding pocket. We conjectured
that the designed structure II (Figure 2) was somewhat less rigid than structure I and
this would be beneficial to the physicochemical properties. In addition, an appropriate
substitution on the alicyclic ring might bring about hydrogen bindings to enhance the
inhibition. Herein, the synthesis and enzymatic activity of the newly designed thiazole
derivatives were described, and the structure-activity relationship analysis and the
molecular docking were performed for guiding further structural optimizations.

Figure 2. The design of new thiazole-based Pin1 inhibitors
2
. Chemistry
The designed compounds were prepared according to Schemes 1-3 and their
structures were listed in Tables 1-5. As shown in Scheme 1, 2-naphthylamine was first
acylated with bromoacetyl bromide to form compound 2, which was reacted with
potassium cyanocarbonimidodithiate 4 obtained from cyanamide to give rise to the key
thiazole intermediate 5 in 76% yield. The reductive amination reaction between 4-
amino subsituted thiazole 5 and ethyl glyoxylate provided compound 6, which was
oxidized in the presence of Na WO .2H O and H O to give the 2-methylsulfonyl
2
4
2
2
2
thiazole derivatives 7. Treatment of compound 7 with a variety of alicyclic amines
resulted in the compounds 8, which were hydrolyzed to afford the corresponding target
compounds 9. In addition, removal the Boc group of compounds 9e and 9f using TFA
gave compounds 9g and 9h.

O
O
O
a
b
Br
Br
Ar
Br
N
H
O
NH2
N
1
2
NH
c
H
d
S
N
N
S
Ar
H
S
N
O
S
Ar
NC
SK
SK
NC NH2
N
O
5
6
3
4
O
O
O
O
OH
O
e
f
g
NH
O
NH
h
h
NH
N
9e
9f
9g
9h
O
S
N
N
H
R2
H
N
R2
H
N
S
Ar
N
S
Ar
S
Ar
O
O
O
9
7
8
9
9
a-9q, 9a-1-9a-6, 9b-1-9b-16: Ar = Naphthalen-2-yl; 9r: Ar = 4-PhPh; 9s: Ar = 3,5-Dichlorobenzyl;
t: Ar = 3-Isoproplybenzyl
Scheme 1. Reagents and conditions (a) ArNH , pyridine, DCM, 53-99%; (b) KOH, CS , ethanol,
2
2
8
5%; (c) i: H O, acetone; ii: NaH, ethanol; iii: MeI, ethanol, 46-81%; (d) Ethyl glyoxalate, NaBH ,
2 4
toluene, 40-55%; (e) Na WO .2H O, H O , ethanol, 74-88%; (f) Alicyclic amine, TEA, dioxane,
2
4
2
2
2
2
0-90%; (g) NaOH, H O, ethanol, 90%; (h) TFA, DCM, 94%.
2
In order to explore the SAR on the 4-position of thiazole scaffold, a number of
derivatives were prepared as outlined in Schemes 2 and 3. The carboxyl amide
derivative 10 was readily obtained through a condensation reaction of compound 9b-
1
5 with NH .H O. The elimination reaction of compound 10 was conducted in the
3 2
presence of ethyl dichlorophosphate and DBU to generate the cyano substituded
compound 11.
Intermediate 12 was obtained by ring closure of compound 4 and ethyl bromoacetate
in a 3-step one-pot reaction. Using Sandmeyer reaction, compound 12 was converted
into 4-bromo substituted thiazole derivative 13, which was subjected to hydrolysis and
condensation reaction to give compound 15. Then the key intermediate 16 was prepared
by Suzuki-Miyaura reaction, which was further transformed into compound 17 by the
reduction reaction in the presence of NaBH and NiCl . Upon subsequent oxidation of the
4
2
methylthio group and the nucleophilic substitution with 2-oxa-6-azaspiro[3.3]heptane,
compound 17 was smoothly converted into intermediate 19, which was hydrolyzed to
afford the target compound 20.

H2N
NH
HO
NH
O
CN
NH
O
i
N
j
N
N
H3C
HO
H
H
N
H C
H3C
HO
H
N
3
N
N
N
S
N
S
HO
S
O
O
O
9b-15
10
11
Scheme 2. Reagents and conditions (i) NH .H O, EDCI, HOBt, THF, 58%; (j) Ethyl
3
2
dichlorophosphate, DBU, DCM, 85%.
NH2
Br
O
Br
O
NC
SK
a
N
b
N
S
c
N
S
N
S
S
S
O
O
OH
S
SK
O
4
12
13
14
O
O
O
O
Br
O
d
e
f
N
S
N
N
S
H
H
H
N
S
S
N
N
S
S
O
O
15
16
17
O
O
O
O
O
OH
O
S
N
N
N
g
h
i
H
H
H
N
N
N
O
N
O
N
S
S
S
O
O
O
O
18
19
20
Scheme 3. Reagents and conditions (a) i: Ethyl bromoacetate, H O, acetone; ii: NaH, ethanol; iii:
2
MeI, ethanol, 78%; (b) CuBr , t-BuONO, acetonitrile, 73%; (c) NaOH, H O, ethanol, 90%; (d) 2-
2
2
Naphthylamine, oxalyl chloride, Py, DCM, 50%; (e) 2-Ethoxylcarbonylvinyl boronic ester,
Pd(OAc) , K PO , 1,1'-bis(di-tert-butylphosphino)ferrocene, toluene, 82%; (f) NaBH , NiCl ,
2
3
4
4
2
MeOH, 34%; (g) Na WO .2H O, H O , ethanol, 64%; (h) 2-Oxa-6-azaspiro[3.3]heptane
2
4
2
2
2
hemioxalate, TEA, dioxane, 38%; (i) NaOH, H O, ethanol, THF, 67%.
2
3
. Biological results and discussion
All target compounds were tested for their inhibitory potency against hPin1 and the
corresponding results were expressed as IC values and presented in Tables 1-5.
5
0
In this work, we mainly focused on the exploration of the SAR on the 2-position of
the thiazole ring. Initially, the impact of alicyclic ring size on the inhibition was
evaluated as shown in Table 1. It was observed that the inhibitory activity gradually
increased from the five-membered ring (compound 9a, IC = 16.61 µM) to the seven-
5
0
membered ring (compound 9c, IC = 5.30 µM). In contrast, compound 9d (IC = 13.70
5
0
50
µM) with an eight-membered ring gave reduced inhibition, suggesting that the binding

site had a limited space. With an aim to guide the further variation of alicyclic ring,
molecular docking was performed with compounds 9a-9d. These compounds showed
similar binding poses, which were illustrated by compound 9b (Figure 3). The acetic
acid extended into the phosphate binding pocket and formed H-bonds and charge-
charge interactions with lys63, Arg69 and Ser114. As expected, the piperidine ring
occupied the proline pocket composed of Leu122, Gln131, Phe134 and Met130. The
bulky naphthalene ring was bound to a hydrophobic region consisting of Leu122 and
Cys113. It was reported that these hydrophobic interactions were beneficial for the
binding affinity.35, 48 As the proline pocket is one of the key binding regions within
Pin1, and the results in Table 1 demonstrated that it could be exploited by the alicyclic
ring with an appropriate size. Considering the diversity and availability of the ring
derivatives, we further explored the SAR of pyrrolidine and piperidine derivatives.
Table 1
The chemical structures and inhibitory activities against hPin1 of compounds 9a - d
O
OH
NH
N
R2
H
N
S
O
R2
IC ± SD (µM)
50
a
Compd.
9
a
16.61 ± 0.99
10.79 ± 2.33
5.30 ± 0.92
13.70 ± 3.09
N
9
b
N
9
c
N
N
9
d
a
The measured IC50 for compound B was 0.09 µM, the reported K for compound B was 0.006
i
µM; SD, standard deviation of two independent assays.

Figure 3. Predicted binding mode of compound 9b (carbon atoms colored blue) was shown in
comparison with a carboxylate Pin1 inhibitor (carbon atoms colored pink) within the co-crystal
4
9
structure (3JYJ in PDB) using CDOCKER protocol integrated in Accelrys Discovery Studio. The
binding pocket of Pin1 was shown in grey. The residues interacting with compound 9b through
hydrogen bonds (green lines) and charge-charge interactions (blue lines) were shown as sticks with
yellow carbon atoms. The image was created by MOE 2019.
The enzymatic activity of the 3-substitued pyrrolidine derivatives were presented in
Table 2. Most of tested compounds exhibited inhibition with IC values of 3.75 µM -
5
0
2
8.52 µM. It was observed that the configuration of the substituted pyrrolidine ring had
a pronounced effect on the inhibition. When a 3-hydroxyl or 3-methoxyl substituted
pyrrolidine was incorporated, the R-enatiomer was more active than the S-enatiomer,
and compound (R)-9a-2 showed more than 5-fold increase than compound (S)-9a-2.
However, when a 3-ethoxyl pyrrolidine was placed on the thiazole ring, the S-enatiomer
was more active than the R-enatiomer. When a F atom or a ketal moiety was used as a
3
-substituent on the pyrrolidine ring, the resultant compounds 9a-5 and 9a-6 exhibited
inhibition as well. In contrast to other compounds in Table 2, compound 9a-4 with a
hydroxylmethyl group had no inhibition at all.Collectively, an H-bond acceptor linked
to pyrrolidine ring was beneficial to the binding and a key feature of this series of
inhibitors.
Table 2
The chemical structures and inhibitory activities against hPin1 of compounds 9a-1 - 9a-
6

O
OH
NH
N
R2
H
N
S
O
R2
IC ± SD (µM) Compd.
50
a
R2
IC ± SD (µM)
a
Compd.
50
(
R)-9a-
(
R)
(R)
N
N
N
N
(
R)-9a-1 HO
3.75 ± 0.64
O
O
23.18 ± 2.62
3
(
S)
(S)
S)
(
S)-9a-1
HO
O
8.46 ± 1.36
7.15 ± 0.06
5.04 ± 0.08
28.52 ± 1.94
(S)-9a-3
9a-4
5.26 ± 0.18
>100
(
N
N
9
a-2
HO
F
(
R)
(S)
N
N
N
(R)-9a-2
O
O
9a-5
10.11 ± 0.38
12.78 ± 1.65
O
(
S)
N
(
S)9a-2
9a-6
O
a
The measured IC50 for compound B was 0.09 µM, the reported K for compound B was 0.006
i
µM; SD, standard deviation of two independent assays.
A variety of piperidine derivatives were evaluated as well and the results were listed
in Table 3. Except for compounds 9b-11, all other compounds displayed inhibitory
activity with IC50 values ranging from 3.04 µM to 19.73 µM. It was noted that
compounds 9b-3 and 9b-4 bearing a hydroxyl group on the 3- or 4-position of
piperidine ring showed stronger inhibition than the corresponding compounds with a
methyl or methoxyl group (compounds 9b-1, 9b-2, 9b-5, and 9b-6), these two
molecules had IC values at a low micromolar level, suggesting that the OH group
5
0
might create an H-bond to benefit the binding. Interestingly, compound 9b-14 (IC =
5
0
3
.63 µM) with a gem-dimethyl group on the piperidine, compound 9b-15 (IC = 3.78
50
µM) with a 4-hydroxyl-4-methyl on the piperidine ring, and compound 9b-16 (IC =
5
0
4
.71 µM) with a tetrahydropiperidine on the 2-position of thiazole scaffold displayed

potent inhibition as well. Presumably, these compounds might possess an appropriate
shape matching with the proline pocket and thus the hydrophobic interactions bolstered
the binding. It was also observed that a somewhat bulky group on the 4-position of
piperidine ring was not preferred, as compounds 9b-7, 9b-12 and 9b-13 showed lower
potency with IC values of > 10 µM. In addition, different from 3-methoxyl substituted
5
0
pyrrolidine derivatives, the R-enatiomer of 3-methoxyl substituted piperidine
derivative ((R)-9b-6) showed similar potency to the S-enatiomer ((S)-9b-6). Taken
together, the results demonstrated that a 4-hydroxyl, 4-hydroxyl-4-methyl, or a gem-
dimethyl group substituted on the 4-position of piperidine ring would be beneficial to
the binding.
Table 3
The chemical structures and inhibitory activities against hPin1 of compounds 9b-1 -
9
b-16
O
OH
NH
N
R2
H
N
S
O
a
R2
a
Compd.
R2
IC ± SD (µM)
Compd.
9b-8
IC ± SD (µM)
5
0
50
O
N
N
9
b-1
7.64 ± 1.59
O
6.78 ± 0.05
N
N
9
9
9
9
b-2
b-3
b-4
b-5
16.42 ± 0.42
3.04 ± 0.00
5.07 ± 0.14
16.12 ± 1.16
9b-9
9b-10
9b-11
9b-12
8.30 ± 0.59
7.43 ± 0.93
>100
HO
HO
N
N
HO
HO
N
N
N
O
N
13.98 ± 0.38
O
NC

O
O
O
N
N
N
9
b-6
8.74 ± 0.65
9b-13
9b-14
9b-15
9b-16
19.73 ± 2.19
3.63 ± 1.37
3.78 ± 0.54
4.71 ± 0.67
F3C
(
S)-9b-
N
(
S)
8
9
.50 ± 0.42
.09 ± 1.85
6
(
R)-9b-
N
N
N
(R)
3
H C
6
HO
N
9
b-7
O
12.91 ± 3.79
O
a
The measured IC50 for compound B was 0.09 µM, the reported K for compound B was 0.006
i
µM; SD, standard deviation of two independent assays.
The piperazine derivatives and its close isosteres were also synthesized and tested
(
Table 4). Compound 9g bearing a piperazine moiety had a weak inhibition (IC =
50
4
6.89 µM). Introducing a 3-methyl piperazine ring on the thiazole ring produced a 7-
fold enhancement in potency (compound 9h, IC = 6.55 µM), whereas incorporating a
5
0
3
,3-dimethyl piperazine, 3,5-dimethyl piperazine or N-methyl piperazine fragment
resulted in a loss of activity (compounds 9i, 9j, 9k), and compound 9l bearing a 3-
methyl-N-methyl piperazine showed similar potency to compound 9g. These results
suggested that very limited modifications could be allowed on the piperazine ring and
the proline pocket had a restricted space. Compounds 9n with a morpholine ring and
9
o with a thiomorpholine ring, the close analogs of compound 9g, displayed a 2.6-fold
and 5-fold increase in potency, respectively, suggesting that an H-bond acceptor (O or
S) benefited to the binding. Remarkably, it was found that compound 9p with a 2-oxa-
6
0
-azaspiro[3,3]heptane fragment presented a strong inhibition with an IC value of
50
.95 µM, which is the most potent inhibitor of this series. In contrast, compound 9q
bearing a 2-azaspiro[3,3]heptane fragment produced a 5.6-fold reduction in potency,
suggesting that the oxygen atom in compound 9p played a key role in the binding, and
it was supposed to be a critical H-bond acceptor.
With an aim to probe the binding features of compound 9p, molecular docking was
conducted and the binding mode was illustrated in Figure 4. As expected, the binding
pose of compound 9p was the same as that of compound 9b, however, the oxygen atom

on the ring formed a hydrogen bond with the backbone NH of Gln131, and this
hydrogen binding greatly contributed to the binding affinity.
Table 4
The chemical structures and inhibitory activities against hPin1of compounds 9g-9q
O
OH
NH
N
R2
H
N
S
O
R2
IC ± SD (µM) Compd.
a
R2
IC ± SD (µM)
a
Compd.
5
0
50
O
N
N
9
9
g
h
46.89 ± 14.17
9m
11.38 ± 0.47
HN
HN
HN
N
N
N
6.55 ± 0.91
>100
9n
9o
9p
9q
17.71 ± 0.92
9.32 ± 0.82
0.95 ± 0.11
5.34 ± 0.35
O
S
N
N
9
9
i
j
HN
HN
N
>100
O
N
N
N
9
k
>100
N
N
9
l
37.07 ± 9.96
a
The measured IC50 for compound B was 0.09 µM, the reported K for compound B was 0.006
i
µM; SD, standard deviation of two independent assays.

Figure 4. Predicted binding mode of compound 9p using CDOCKER protocol integrated in
Accelrys Discovery Studio. 49 The enzyme was shown in grey, compound 9p was shown as sticks
with blue carbon atoms. The residues that interacted with compound 9p were shown as sticks with
yellow carbon atoms, and hydrogen bonds were indicated with green lines and charge-charge
interactions (blue lines) were shown as sticks with yellow carbon atoms. The image was created by
MOE 2019.
We also preliminarily investigated the SAR of 4- and 5-substitution of the thiazole
ring. Changing the carboxylic acid group of compound 9b-15 to the corresponding
carboxylic amide (compound 10) or acetonitrile analog (compound 11) led to loss of
potency, demonstrating that the carboxylic acid group is a characteristic structural
moiety as well for this series of inhibitors, and this result is consistent with the predicted
binding mode (Figure 4). The replacement of the NH group on the side chain of
compound 9p with a CH moiety afforded compound 20, which displayed a 12-fold
2
reduction in potency, demonstrating that this NH group was critical for the binding. As
shown in Figure 4, presumably, partly because the intramolecular hydrogen bond
between the NH and carbonyl group on the ortho position was favorable for the binding
through a less entropy cost. When the naphthyl fragment in compound 9p was
substituted by other aromatic segments including the biphenyl, 3,4-dichlorophenyl, or
3
-isopropylphenyl group, the inhibition of the resulted compounds (9r, 9s and 9t)

declined drastically, indicating that the fused aromatic ring was preferred, and this
hydrophobic interaction made a pronounced contribution to the inhibition.
Table 5
The chemical structures and inhibitory activities against hPin1 of compounds 9b-15,
1
0, 11, 9p, 20, 9r-9t
R4
N
O
R2
S
R5
R2
R4
R5
IC ± SD (µM)
50
a
Compd
O
H
N
9
b-15
3.77±0.54
H
N
N
N
N
OH
H3C
HO
O
H
N
1
1
9
2
0
1
p
0
>100
H
N
NH2
H3C
HO
H
N
H
N
>100
CN
O
H3C
HO
H
N
0.95±0.11
11.69±3.16
13.35±1.29
28.00±4.40
16.83±1.83
N
H
N
OH
O
O
O
O
O
O
O
O
O
H
N
N
N
N
N
OH
H
9
r
H
N
N
OH
OH
OH
H
9
s
N
Cl
Cl
H
N
H
9
t
H
N
N
a
The measured IC50 for compound B was 0.09 µM, the reported K for compound B was 0.006
i
µM; SD, standard deviation of two independent assays.
4
. Conclusion
In summary, a series of novel thiazole derivatives bearing alicyclic heterocycles on

the 2-position were prepared to explore the proline binding pocket of Pin1. As a result,
compound 9p was disclosed as the most potent inhibitor with an IC value of 0.95 µM,
5
0
and it possessed drug-like properties with ClogP value of 3.20 and tPSA of 103.26. It
provides a new template for further structural modifications. The SAR and molecular
docking results indicated that introducing an H-bond acceptor to form an H-bond with
Gln131 was accessible and greatly benefited to the binding affinity. The proline pocket
could accommodate various hydrophobic alicyclic heterocycles, however, the space
was somewhat restricted. In addition, it has been demonstrated that the carboxylic acid
and the bulky aromatic moiety were also the key pharmacophoric groups, which were
consistent with the known SAR. The above results would facilitate to discover new
Pin1 inhibitors for probing the potential of Pin1 as an anticancer target.
5
.Experimental
5
.1. General
1H NMR spectra on a Varian Mercury 400 or 500 spectrometer were recorded using
tetramethylsilane (TMS) as the internal standard in Acetone-d , DMSO-d , MeOD or
6
6
CDCl . High resolution mass spectra (HRMS) were obtained on an Agilent
3
Technologies LC/MSD TOF spectrometer. Melting points were measured on a Yanaco
micro melting point apparatus and were uncorrected. All chemicals and solvents used
were of reagent grade without further purification or dried before used. All the reactions
were monitored by thin-layer chromatography (TLC) under a UV lamp at 254 nm.
Column chromatography separations were performed with silica gel (200-300 mesh).
5
5
.2. Synthesis of compounds
.2.1 (5-(Naphthalen-2-ylcarbamoyl)-2-(pyrrolidin-1-yl)thiazol-4-yl)glycine (9a)
To a stirred solution of 8a (79 mg, 0.19mmol) in ethanol (2 mL) was added NaOH
(
22 mg, 0.56 mmol) dissolved in H O (1 mL) dropwise, the reaction mixture was then
2
stirred at room temperature for 3 h. Ethanol was removed under reduced pressure, then
adjusting pH of the residue to 2-3 with dilute hydrochloric acid. After filtration and
drying under high vacuum, compound 9a was obtained as a white solid (70 mg, 93%);
1
mp 162-164 °C; H NMR (400 MHz, DMSO-d ) δ (ppm) 8.97 (s, 1H), 8.30 (d, J = 1.9
6
Hz, 1H), 8.09 (t, J = 5.7 Hz, 1H), 7.83 - 7.71 (m, 4H), 7.46 - 7.31 (m, 2H), 4.09 (d, J =
1
3
5
.6 Hz, 2H), 3.42 (brs, 4H), 2.03 - 1.95 (m, 4H); C NMR (150 MHz, DMSO-d ) δ
6
(ppm) 172.54, 166.07, 163.74, 163.29, 138.47, 134.04, 129.66, 128.14, 127.81, 127.55,
1
26.56, 124.40, 121.57, 115.73, 81.96, 49.47, 46.06, 25.59. HRMS (ESI): m/z, Calcd.
+
for C H O N S [M+H] : 397.1329, Found 397.1345.
2
0
21
3
4

5
.2.2 (5-(Naphthalen-2-ylcarbamoyl)-2-(piperidin-1-yl)thiazol-4-yl)glycine (9b)
Following the preparation protocol of compound 9a, starting from compound 8b (50
mg, 0.11 mmol), LiOH instead of NaOH was used, the title compound 9b was obtained
1
as a white solid (10 mg, 25%); mp 132-134 °C; H NMR (400 MHz, DMSO-d ) δ (ppm)
6
9
-
.00 (s, 1H), 8.28 (d, J = 2.0 Hz, 1H), 8.04 (t, J = 5.9 Hz, 1H), 7.84 - 7.69 (m, 4H), 7.47
7.31 (m, 2H), 4.12 (d, J = 5.8 Hz, 2H), 3.50 (brs, 4H), 1.62 (brs, 6H); C NMR (150
1
3
MHz, DMSO-d ) δ (ppm) 172.59, 169.28, 163.49, 163.27, 138.33, 134.01, 129.73,
6
1
2
28.18, 127.83, 127.56, 126.60, 124.48, 121.60, 115.87, 82.08, 48.90, 45.61, 25.21,
+
3.90; HRMS (ESI): m/z, Calcd. for C H O N S [M+H] : 411.1485, Found 411.1469.
2
1
23
3
4
5
.2.3 (2-(Azepan-1-yl)-5-(naphthalen-2-ylcarbamoyl)thiazol-4-yl)glycine (9c)
Following the preparation protocol of compound 9a, starting from compound 8c (130
mg, 0.33 mmol), the title compound 9c was obtained as a grayish white solid (93 mg,
1
7
6%); mp 131-133 °C; H NMR (400 MHz, Acetone-d ) δ (ppm) 8.36 (s, 1H), 8.18 (t,
6
J = 5.2 Hz, 1H), 8.11 (s, 1H), 7.82 - 7.72 (m, 4H), 7.45 - 7.30 (m, 2H), 4.30 (d, J = 4.7
1
3
Hz, 2H), 3.76 - 3.48(m, 4H), 1.89 - 1.78 (m, 4H), 1.65 - 1.55 (m, 4H); C NMR (150
MHz, DMSO-d ) δ (ppm) 172.54, 168.63, 163.54, 163.21, 138.32, 133.96, 129.65,
6
1
1
28.11, 127.77, 127.50, 126.53, 124.40, 121.52, 115.76, 81.90, 45.31, 27.32, 27.26,
+
5.61; HRMS (ESI): m/z, Calcd. for C H O N S [M+H] : 425.1642, Found 425.1670.
2
2
25
3
4
5
.2.4 (2-(Azepan-1-yl)-5-(naphthalen-2-ylcarbamoyl)thiazol-4-yl)glycine (9d)
Following the preparation protocol of compound 9a, starting from compound 8d (50
mg, 0.11 mmol), the title compound 9d was obtained as a white solid (24 mg, 76%);
1
mp 133-135 °C; H NMR (400 MHz, DMSO-d ) δ (ppm) 8.97 (s, 1H), 8.29 (d, J = 2.0
6
Hz, 1H), 8.04 (t, J = 5.8 Hz, 1H), 7.83 - 7.69 (m, 4H), 7.46 - 7.30 (m, 2H), 4.11 (d, J =
5
.7 Hz, 2H), 3.55 (brs, 4H), 1.79 (brs, 4H), 1.60 - 1.40 (m, 6H); HRMS (ESI): m/z,
+
Calcd. for C H O N S [M+H] : 439.1798, Found 439.1801.
2
3
27
3
4
5
.2.5
(R)-(2-(3-Hydroxypyrrolidin-1-yl)-5-(naphthalen-2-ylcarbamoyl)thiazol-4-
yl)glycine ((R)-9a-1)
Following the preparation protocol of compound 9a, starting from compound 8a-1R
(
100 mg, 0.22 mmol), LiOH was used, the title compound (R)-9a-1 was obtained as a
1
white solid (50 mg, 53%); mp 158-160 °C; H NMR (400 MHz, DMSO-d ) δ (ppm)
6
9
-
.00 (s, 1H), 8.29 (d, J = 2.0 Hz, 1H), 8.09 (t, J = 5.9 Hz, 1H), 7.84 - 7.69 (m, 4H), 7.47
7.31 (m, 2H), 4.43 (brs, 1H), 4.15 (d, J = 5.0 Hz, 2H), 3.59 - 3.45 (m, 4H), 2.17 - 1.88
1
3
(m, 2H); C NMR (150 MHz, DMSO-d ) δ (ppm) 172.56, 166.35, 163.72, 163.30,
6
1
6
4
38.33, 133.99, 129.70, 128.16, 127.82, 127.55, 126.58, 124.46, 121.57, 115.85, 82.24,
+
9.73, 57.85, 47.69, 45.27, 34.08; HRMS (ESI): m/z, Calcd. for C H O N S [M+H] :
2
0
21
4
4
13.1278, Found 413.1314.
5
.2.6 (S)-(2-(3-Hydroxypyrrolidin-1-yl)-5-(naphthalen-2-ylcarbamoyl)thiazol-4-
yl)glycine ((S)-9a-1)
Following the preparation protocol of compound 9a, starting from compound 8a-1S
(100 mg, 0.22 mmol), LiOH was used, the title compound (S)-9a-1 was obtained as a

1
white solid (66 mg, 70%); mp 149-151 °C; H NMR (400 MHz, DMSO-d ) δ (ppm)
6
9
4
.00 (s, 1H), 8.29 (s, 1H), 8.20 - 7.98 (m, 1H), 7.94 - 7.69 (m, 4H), 7.53 - 7.30 (m, 2H),
.43 (s, 1H), 4.15 (s, 2H), 3.53 (s, 4H), 2.19 - 1.85 (m, 2H); C NMR (150 MHz,
1
3
DMSO-d ) δ 172.57, 166.37, 163.74, 163.32, 138.34, 134.01, 129.72, 128.17, 127.83,
6
1
27.56, 126.59, 124.48, 121.58, 115.87, 82.25, 69.74, 57.87, 47.71, 45.29, 34.09;
+
HRMS (ESI): m/z, Calcd. for C H O N S [M+H] : 413.1278, Found 413.1255.
2
0
21
4
4
5
.2.7
((2-(3-Methoxypyrrolidin-1-yl)-5-(naphthalen-2-ylcarbamoyl)thiazol-4-
yl)glycine (9a-2)
Following the preparation protocol of compound 9a, starting from compound 8a-2
(
9
50 mg, 0.11 mmol), the title compound 9a-2 was obtained as a yellow solid (40 mg,
1
4%); mp 136-137 °C; H NMR (400 MHz, DMSO-d ) δ (ppm) 12.576 (s, 1H), 9.02
6
(s, 1H),8.29 (d, J = 2.0 Hz, 1H), 8.09 (t, J = 6.0 Hz, 1H), 7.84 - 7.69 (m, 4H), 7.48 -
7
2
.29 (m, 2H), 4.15 (d, J = 5.9 Hz, 2H), 4.11 (m, 1H), 3.61 - 3.84 (m, 4H),3.28 (s, 3H),
+
.18-2.05 (m, 2H); HRMS (ESI): m/z, Calcd. for C H O N S [M+H] : 427.1435,
2
1
23
4
4
Found 427.1409.
5
.2.8
(R)-(2-(3-Methoxypyrrolidin-1-yl)-5-(naphthalen-2-ylcarbamoyl)thiazol-4-
yl)glycine ((R)-9a-2)
Following the preparation protocol of compound 9a, starting from compound 8a-2R
(
6
80 mg, 0.18 mmol), the title compound (R)-9a-2 was obtained as a white solid (52 mg,
1
9%); mp 126-128 °C; H NMR (400 MHz, DMSO-d ) δ (ppm) 9.02 (s, 1H), 8.30 (d,
6
J = 2.4 Hz, 1H), 8.10 (brs, 1H), 7.87 - 7.69 (m, 4H), 7.47 - 7.32 (m, 2H), 4.23 - 4.07
1
3
(
m, 3H), 3.60 - 3.36 (m, 4H), 3.28 (s, 3H), 2.19 - 2.07 (m, 2H); C NMR (150 MHz,
DMSO-d ) δ (ppm) 172.55, 166.30, 163.65, 163.29, 138.31, 133.99, 129.72, 128.16,
6
1
4
4
27.82, 127.56, 126.58, 124.48, 121.57, 115.88, 82.38, 79.51, 56.38, 54.52, 47.65,
+
5.29, 30.65; HRMS (ESI): m/z, Calcd. for C H O N S [M+H] : 427.1435, Found
2
1
23
4
4
27.1405.
5
.2.9
(S)-(2-(3-Methoxypyrrolidin-1-yl)-5-(naphthalen-2-ylcarbamoyl)thiazol-4-
yl)glycine ((S)-9a-2)
Following the preparation protocol of compound 9a, starting from compound 8a-2S
(
100 mg, 0.22 mmol), the title compound (S)-9a-2 was obtained as a light pink solid
1
(88 mg, 95%); mp 126-128 °C; H NMR (400 MHz, DMSO-d ) δ (ppm) 9.03 (s, 1H),
6
8
3
1
1
.29 (s, 1H), 7.96 - 7.68 (m, 4H), 7.54 - 7.29 (m, 2H), 4.15 (s, 2H), 4.12 (s, 1H), 3.63 -
1
3
.38 (m, 4H), 3.28 (s, 3H), 2.14 (s, 2H); C NMR (150 MHz, DMSO-d ) δ (ppm)
6
72.54, 166.30, 163.64, 163.29, 138.30, 133.98, 129.71, 128.16, 127.82, 127.56,
26.58, 124.48, 121.57, 115.88, 82.38, 79.51, 56.38, 54.51, 47.65, 45.29, 30.65; HRMS
+
(ESI): m/z, Calcd. for C H O N S [M+H] : 427.1435, Found 427.1407.
2
1
23
4
4
5
.2.10 (R)-(2-(3-Ehoxypyrrolidin-1-yl)-5-(naphthalen-2-ylcarbamoyl)thiazol-4-
yl)glycine ((R)-9a-3)
Following the preparation protocol of compound 9a, starting from compound 8a-3R
(70 mg, 0.18 mmol), the title compound (R)-9a-3 was obtained as a light pink solid (31

1
mg, 48%); mp 130-132 °C; H NMR (500 MHz, DMSO-d ) δ (ppm) 8.92 (s, 1H), 8.33
6
(
2
s, 1H), 8.11 (brs, 1H), 7.82 - 7.69 (m, 3H), 7.46 - 7.29 (m, 2H), 4.21 (s, 1H), 3.71 (m,
H), 3.64 - 3.33 (m, 6H), 2.12 (s, 2H), 1.12 (t, J = 7.1 Hz, 3H); C NMR (150 MHz,
1
3
DMSO-d ) δ (ppm) 172.23, 166.22, 163.55, 163.07, 138.80, 134.11, 129.50, 128.04,
6
1
4
27.77, 127.51, 126.48, 124.20, 121.50, 115.30, 81.07, 77.73, 64.03, 54.84, 48.98,
+
7.59, 31.13, 15.79. HRMS (ESI): m/z, Calcd. for C H O N S [M+H] : 441.1591,
2
2
25
4
4
Found 441.1561.
5
.2.11
(S)-(2-(3-Ethoxypyrrolidin-1-yl)-5-(naphthalen-2-ylcarbamoyl)thiazol-4-
yl)glycine ((S)-9a-3)
Following the preparation protocol of compound 9a, starting from compound (S)-
8
a-3 (100 mg, 0.21 mmol), the title compound (S)-9a-3 was obtained as a white solid
1
(79 mg, 84%); mp 131-133 °C; H NMR (400 MHz, CD OD) δ (ppm) δ 8.04 (d, J = 2.1
3
Hz, 1H), 7.77 (t, J = 7.6 Hz, 3H), 7.65 - 7.57 (m, 1H), 7.53 - 7.32 (m, 2H), 4.26 (s, 3H),
1
3
3
.70 - 3.39 (m, 6H), 2.25 - 2.09 (m, 2H), 1.21 (t, J = 7.0 Hz, 3H); C NMR (150 MHz,
DMSO-d ) δ (ppm) 172.57, 166.28, 163.67, 163.31, 138.32, 134.00, 129.73, 128.17,
6
1
4
27.83, 127.56, 126.59, 124.49, 121.58, 115.89, 82.37, 77.72, 64.04, 54.95, 47.69,
+
5.29, 31.09, 15.78; HRMS (ESI): m/z, Calcd. for C H O N S [M+H] : 441.1591,
2
2
25
4
4
Found 441.1633.
5
.2.12
(S)-(2-(3-(Hydroxymethyl)pyrrolidin-1-yl)-5-(naphthalen-2-
ylcarbamoyl)thiazol-4-yl)glycine (9a-4)
Following the preparation protocol of compound 9a, starting from compound 8a-4
(
7
60 mg, 0.13 mmol), the title compound 9a-4 was obtained as a white solid (34 mg,
1
4%); mp 183-185 °C; H NMR (400 MHz, DMSO-d ) δ(ppm) 8.87 (s, 1H), 8.32 (d, J
6
=
1.8 Hz, 1H), 8.06 (t, J = 4.0 Hz, 1H), 7.82 - 7.69 (m, 4H), 7.46 - 7.27 (m, 2H), 4.81
(brs, 1H), 3.66 (d, J = 4.3 Hz, 2H), 3.55 - 3.39 (m, 5H), 3.27 - 3.19 (m, 1H), 2.48 - 2.42
(
m, 1H), 2.13 - 1.99 (m, 1H), 1.86 - 1.74 (m, 1H); HRMS (ESI): m/z, Calcd. for
+
C H O N S [M+H] : 427.1435, Found 424.1478.
2
1
23
4
4
5
.2.13
(
(
S)-(2-(3-Fluoropyrrolidin-1-yl)-5-(naphthalen-2-ylcarbamoyl)thiazol-4-yl)glycine
9a-5)
Following the preparation protocol of compound 9a, starting from compound 8a-5
(
8
1
50 mg, 0.11 mmol), the title compound 9a-5 was obtained as a white solid (39 mg,
1
3%); mp 176-178 °C; H NMR (400 MHz, DMSO-d ) δ (ppm) 12.59 (s, 1H), 9.07 (s,
6
H), 8.30 (d, J = 2.0 Hz, 1H), 8.10 (t, J = 5.9 Hz, 1H), 7.85 - 7.70 (m, 4H), 7.48 - 7.31
(
-
m, 2H), 5.47 (d, J = 52.8 Hz, 1H), 4.16 (d, J = 5.8 Hz, 2H), 3.83 - 3.47 (m, 4H), 2.44
1
3
2.17 (m, 2H); C NMR (150 MHz, DMSO-d ) δ (ppm) 172.54, 166.17, 163.54,
6
1
1
63.28, 138.25, 133.98, 129.74, 128.18, 127.82, 127.56, 126.59, 124.51, 121.57,
15.95, 94.10, 92.95, 82.64, 56.04, 55.89, 47.38, 45.30, 32.26, 32.12; HRMS (ESI):
+
m/z, Calcd. for C H O N SF [M+H] : 415.1235, Found 415.1242.
2
0
20
3
4
5
.2.14
(5-(Naphthalen-2-ylcarbamoyl)-2-(1,4-dioxa-7-azaspiro[4.4]nonan-7-

yl)thiazol-4-yl)glycine (9a-6)
Following the preparation protocol of compound 9a, starting from compound 8a-6
(
76 mg, 0.16 mmol), the title compound 9a-6 was obtained as a yellow solid (59 mg,
1
8
1
4
2%); mp 135-137 °C; H NMR (400 MHz, DMSO-d ) δ (ppm) 9.07 (s, 1H), 8.29 (s,
6
H), 8.08 (brs, 1H), 7.84 - 7.67 (m, 4H), 7.39 (m, 2H), 4.14 (s, 2H), 3.98 - 3.95 (m,
1
3
H), 3.53 (t, J = 6.4 Hz, 2H), 3.49 (s, 2H), 2.21 (t, J = 7.2 Hz, 2H); C NMR (150
MHz, DMSO-d ) δ (ppm) 172.54, 166.49, 163.50, 163.29, 138.24, 133.98, 129.75,
6
1
4
28.18, 127.82, 127.57, 126.59, 124.52, 121.58, 115.96, 113.35, 82.62, 64.95, 56.22,
+
7.75, 45.29, 34.36; HRMS (ESI): m/z, Calcd. for C H O N S [M+H] : 455.1384,
2
2
23
5
4
Found 455.1382.
5
(
.2.15 (2-(4-Methylpiperidin-1-yl)-5-(naphthalen-2-ylcarbamoyl)thiazol-4-yl)glycine
9b-1)
Following the preparation protocol of compound 9a, starting from compound 8b-1
(
4
80 mg, 0.18 mmol), the title compound 9b-1 was obtained as a white solid (33 mg,
1
4%); mp 148-150 °C; H NMR (400 MHz, DMSO-d ) δ (ppm): 8.99 (s, 1H), 8.28 (d,
6
J = 2.1 Hz, 1H), 8.04 (brs, 1H), 7.82 - 7.70 (m, 4H), 7.48 - 7.30 (m, 2H), 4.12 (s, 2H),
3
.95 - 3.84 (m, 2H), 3.19 - 3.05 (m, 2H), 1.79 - 1.58 (m, 3H), 1.29 - 1.08 (m, 2H), 0.94
+
(d, J = 6.4 Hz, 4H); HRMS (ESI): m/z, Calcd. for C H O N S [M+H] : 425.1642,
2
2
25
3
4
Found 425.1634.
5
(
.2.16 (2-(3-Methylpiperidin-1-yl)-5-(naphthalen-2-ylcarbamoyl)thiazol-4-yl)glycine
9b-2)
Following the preparation protocol of compound 9a, starting from compound 8b-2
(
4
80 mg, 0.18 mmol), the title compound 9b-2 was obtained as a white solid (33 mg,
1
4%); mp 134-135 °C; H NMR (400 MHz, Acetone-d ) δ (ppm) 8.35 (s, 1H), 8.16 (t,
6
J = 6.0 Hz, 1H), 8.11 (brs, 1H), 7.82 - 7.74 (m, 4H), 7.45 - 7.31 (m, 2H), 4.28 (d, J =
.4 Hz, 2H), 4.00 - 3.84 (m, 2H), 3.13 - 3.02 (m, 1H), 2.84- 2.72 (m, 1H), 1.92 - 1.82
m, 1H), 1.82 - 1.67 (m, 2H), 1.66 - 1.50 (m, 1H), 1.31 - 1.16 (m, 1H), 0.96 (d, J = 6.6
4
(
1
3
Hz, 3H); C NMR (150 MHz, DMSO-d ) δ (ppm) 172.57, 169.17, 163.47, 163.25,
6
1
4
4
38.28, 133.98, 129.72, 128.18, 127.82, 127.55, 126.59, 124.49, 121.54, 115.86, 82.12,
+
5.27, 32.31, 30.67, 24.54, 19.17; HRMS (ESI): m/z, Calcd. for C H O N S [M+H] :
2
2
25
3
4
25.1642, Found 425.1637.
5
.2.17 (2-(4-Hydroxypiperidin-1-yl)-5-(naphthalen-2-ylcarbamoyl)thiazol-4-
yl)glycine (9b-3)
Following the preparation protocol of compound 9a, starting from compound 8b-3
(
8
100 mg, 0.22 mmol), the title compound 9b-3 was obtained as a white solid (84 mg,
1
9%); mp 158-160 °C; H NMR (400 MHz, DMSO-d ) δ (ppm) 9.03 (s, 1H), 8.29 (d,
6
J = 2.1 Hz, 1H), 8.05 (t, J = 4.8 Hz, 1H), 7.82 - 7.69 (m, 4H), 7.46 - 7.32 (m, 2H), 4.12
(
-
d, J = 4.6 Hz, 2H), 3.81 - 3.68 (m, 3H), 3.32 - 3.25 (m, 2H), 1.92 - 1.77 (m, 2H), 1.53
1
3
1.39 (m, 2H); C NMR (150 MHz, DMSO-d ) δ (ppm) 172.52, 169.09, 163.42,
6
1
1
63.21, 138.20, 133.94, 129.70, 128.14, 127.78, 127.51, 126.55, 124.47, 121.55,
+
15.89, 82.35, 65.29, 45.23, 33.57; HRMS (ESI): m/z, Calcd. for C H O N S [M+H] :
2
1
23
4
4

4
27.1435, Found 427.1433.
.2.18 (2-(3-Hydroxypiperidin-1-yl)-5-(naphthalen-2-ylcarbamoyl)thiazol-4-
5
yl)glycine (9b-4)
Following the preparation protocol of compound 9a, starting from compound 8b-4
(
9
8
100 mg, 0.22 mmol), the title compound 9b-4 was obtained as a white solid (87 mg,
1
2%); mp 159-161 °C; H NMR (400 MHz, DMSO-d ) δ 12.55 (s, 1H), 8.99 (s, 1H),
6
.28 (s, 1H), 8.05 (t, J = 6.0 Hz, 1H), 7.85 - 7.68 (m, 4H), 7.48 - 7.29 (m, 2H), 5.06 (d,
J = 4.2 Hz, 1H), 4.13 (d, J = 5.8 Hz, 2H), 3.87 - 3.71 (m, 1H), 3.62 (brs, 2H), 3.28 -
3
.19 (m, 1H), 3.10 - 2.97 (m, 1H), 1.95 - 1.71 (m, 2H), 1.47 (m, 2H); HRMS (ESI):
+
m/z, Calcd. for C H O N S [M+H] : 427.1435, Found 427.1454.
2
1
23
4
4
5
.2.19
(2-(4-Methoxypiperidin-1-yl)-5-(naphthalen-2-ylcarbamoyl)thiazol-4-
yl)glycine (9b-5)
Following the preparation protocol of compound 9a, starting from compound 8b-5 (140
mg, 0.30 mmol), the title compound 9b-5 was obtained as a white solid (47 mg, 36%);
1
mp 163-165 °C; H NMR (400 MHz, DMSO-d ) δ (ppm) 9.06 (s, 1H), 8.29 (d, J = 2.0
6
Hz, 1H), 8.04 (t, J = 5.9 Hz, 1H), 7.83 - 7.71 (m, 4H), 7.47 - 7.31 (m, 2H), 4.12 (d, J =
5
2
1
1
.8 Hz, 2H), 3.76 - 3.63 (m, 2H), 3.53 - 3.44 (m, 1H), 3.42 - 3.37 (m, 2H), 3.29 (s, 3H),
1
3
.00 - 1.85 (m, 2H), 1.62 - 1.48 (m, 2H); C NMR (150 MHz, DMSO-d ) δ (ppm)
6
72.51, 169.16, 163.39, 163.26, 138.24, 133.97, 129.74, 128.16, 127.82, 127.55,
26.58, 124.50, 121.61, 115.95, 82.52, 74.72, 55.61, 45.33, 29.95; HRMS (ESI): m/z,
+
Calcd. for C H O N S [M+H] : 441.1591, Found 441.1592.
2
2
25
4
4
5
.2.20
(2-(3-Methoxypiperidin-1-yl)-5-(naphthalen-2-ylcarbamoyl)thiazol-4-
yl)glycine (9b-6)
Following the preparation protocol of compound 9a, starting from compound 8b-6
(
100 mg, 0.22 mmol), LiOH instead of NaOH was used, the title compound 9b-6 was
1
obtained as a white solid (24 mg, 50%); mp 107-109 °C; H NMR (400 MHz, DMSO-
d6) δ (ppm) 12.54 (s, 1H), 8.99 (s, 1H), 8.28 (d, J = 2.0 Hz, 1H), 8.04 (t, J = 5.9 Hz,
1
2
H), 7.83 - 7.68 (m, 4H), 7.47 - 7.31 (m, 2H), 4.12 (d, J = 5.9 Hz, 2H), 3.70 - 3.53 (m,
H), 3.51 - 3.44 (m, 2H), 3.42 - 3.37 (m, 1H), 3.29 (s, 3H), 1.92 - 1.45 (m, 4H); C
1
3
NMR (150 MHz, DMSO-d ) δ (ppm): 172.57, 169.38, 163.40, 163.24, 138.26, 133.98,
6
1
4
29.73, 128.17, 127.82, 127.54, 126.59, 124.49, 121.57, 115.91, 82.08, 73.96, 56.12,
+
5.26, 28.97, 21.34; HRMS (ESI): m/z, Calcd. for C H O N S [M+H] : 441.1591,
2
2
25
4
4
Found 441.1590.
5
.2.21
(S)-(2-(3-Methoxypiperidin-1-yl)-5-(naphthalen-2-ylcarbamoyl)thiazol-4-
yl)glycine ((S)-9b-6)
Following the preparation protocol of compound 9a, starting from compound (S)-
8
b-6 (50 mg, 0.11 mmol), the title compound (S)-9b-6 was obtained as a white solid
1
(40 mg, 91%); mp 115-117 °C; H NMR (400 MHz, DMSO-d ) δ (ppm) 8.99 (s, 1H),
6
8
.28 (d, J = 2.0 Hz, 1H), 8.04 (brs, 1H), 7.85 - 7.66 (m, 4H), 7.46 - 7.32 (m, 2H), 4.12
(s, 2H), 3.66 - 3.60 (m, 2H), 3.49 - 3.36 (m, 3H), 3.29 (s, 3H), 1.91 - 1.73 (m, 2H), 1.73

1
3
-
1.59 (m, 1H), 1.58 - 1.47 (m, 1H); C NMR (150 MHz, DMSO-d ) δ (ppm) 172.56,
6
1
1
69.38, 163.40, 163.24, 138.26, 133.98, 129.73, 128.17, 127.82, 127.54, 126.58,
24.49, 121.57, 115.91, 82.08, 73.96, 56.12, 50.65, 48.45, 45.25, 28.97, 21.34; HRMS
+
(ESI): m/z, Calcd. for C H O N S [M+H] : 441.1591, Found 441.1585.
2
2
25
4
4
5
.2.22
(R)-(2-(3-Methoxypiperidin-1-yl)-5-(naphthalen-2-ylcarbamoyl)thiazol-4-
yl)glycine ((R)-9b-6)
Following the preparation protocol of compound 9a, starting from compound (R)-
8
b-6 (45 mg, 0.10 mmol), the title compound (R)-9b-6 was obtained as a white solid
1
(40 mg, 95%); mp 123-125 °C; H NMR (400 MHz, DMSO-d ) δ (ppm) 8.99 (s, 1H),
6
8
2
1
1
1
.32 - 8.24 (m, 1H), 8.05 (brs, 1H), 7.83 - 7.69 (m, 3H), 7.48 - 7.32 (m, 2H), 4.12 (s,
H), 3.67 - 3.59 (m, 2H), 3.51 - 3.36 (m, 3H), 3.29 (s, 3H), 1.93 - 1.72 (m, 2H), 1.69 -
1
3
.60 (m, 1H), 1.57 - 1.45 (m, 1H); C NMR (150 MHz, DMSO-d ) δ (ppm) 172.56,
6
69.39, 163.40, 163.24, 138.26, 133.98, 129.73, 128.17, 127.82, 127.54, 126.59,
24.49, 121.58, 115.91, 82.08, 73.96, 56.12, 50.66, 48.42, 45.26, 28.97, 21.34; HRMS
+
(ESI): m/z, Calcd. for C H O N S [M+H] : 441.1591, Found 441.1584.
2
2
25
4
4
5
.2.23
(5-(Naphthalen-2-ylcarbamoyl)-2-(1,4-dioxa-8-azaspiro[4.5]decan-8-
yl)thiazol-4-yl)glycine (9b-7)
Following the preparation protocol of compound 9a, starting from compound 8b-7
(
100 mg, 0.20 mmol), the title compound 9b-7 was obtained as a white solid (85 mg,
1
9
1
2
4
1
6
0%); mp 131-133 °C; H NMR (400 MHz, DMSO-d ) δ (ppm) 12.56 (s, 1H), 9.05 (s,
6
H), 8.28 (d, J = 2.1 Hz, 1H), 8.09 - 8.01 (m, 1H), 7.85 - 7.67 (m, 4H), 7.48 - 7.31 (m,
H), 4.13 (d, J = 5.8 Hz, 2H), 3.94 (s, 4H), 3.59 (t, J = 5.6 Hz, 4H), 1.75 (t, J = 5.9 Hz,
1
3
H); C NMR (150 MHz, DMSO-d ) δ (ppm) 172.56, 169.34, 163.31, 163.22, 138.25,
6
33.98, 129.74, 128.17, 127.82, 127.55, 126.59, 124.50, 121.57, 115.91, 105.51, 82.12,
4.72, 49.38, 47.73, 45.29, 33.68, 22.68; HRMS (ESI): m/z, Calcd. for C H O N S
2
3
25
4
5
+
[M+H] : 469.1540, Found 469.1529.
5
.2.24
(5-(Naphthalen-2-ylcarbamoyl)-2-(1,4-dioxa-7-azaspiro[4.5]decan-7-
yl)thiazol-4-yl)glycine (9b-8)
Following the preparation protocol of compound 9a, starting from compound 8b-8
(
9
100 mg, 0.20 mmol), the title compound 9b-8 was obtained as a white solid (88 mg,
1
3%); mp 143-145 °C; H NMR (400 MHz, DMSO-d ) δ (ppm) 8.99 (s, 1H), 8.28 (d,
6
J = 2.1 Hz, 1H), 8.06 (s, 1H), 7.83 - 7.67 (m, 4H), 7.47 - 7.29 (m, 2H), 4.12 (s, 2H),
1
3
3
.98 - 3.92 (m, 4H), 3.51 (s, 4H), 1.84 - 1.71 (m, 4H); C NMR (150 MHz, DMSO-d )
6
δ (ppm) 172.53, 169.01, 163.33, 163.23, 138.19, 133.96, 129.76, 128.19, 127.82,
1
27.56, 126.60, 124.53, 121.58, 115.97, 106.44, 82.73, 64.42, 46.36, 45.28, 34.11;
+
HRMS (ESI): m/z, Calcd. for C H O N S [M+H] : 469.1540, Found 469.1538.
2
3
25
4
5
5
.2.25 (2-(4-(Hydroxymethyl)piperidin-1-yl)-5-(naphthalen-2-ylcarbamoyl)thiazol-4-
yl)glycine (9b-9)
Following the preparation protocol of compound 9a, starting from compound 8b-9
(100 mg, 0.21 mmol), the title compound 9b-9 was obtained as a yellow solid (85 mg,

1
9
1
0%); mp 155-157 °C; H NMR (400 MHz, DMSO-d ) δ (ppm) 12.59 (s, 1H), 8.99 (s,
6
H), 8.28 (d, J = 2.0 Hz, 1H), 8.04 (t, J = 5.9 Hz, 1H), 7.86 - 7.66 (m, 4H), 7.49 - 7.30
(m, 2H), 4.55 (brs, 1H), 4.12 (d, J = 5.9 Hz, 2H), 4.02 - 3.85 (m, 2H), 3.31 - 3.26 (m,
2
H), 3.19 - 3.02 (m, 2H), 1.85 - 1.73 (m, 2H), 1.73 - 1.59 (m, 1H), 1.29 - 1.13 (m, 2H);
1
3
C NMR (150 MHz, DMSO-d ) δ (ppm) 172.56, 169.21, 163.47, 163.27, 138.26,
6
1
4
4
33.98, 129.74, 128.17, 127.82, 127.55, 126.59, 124.50, 121.58, 115.91, 82.23, 65.80,
+
8.12, 45.29, 38.32, 28.26; HRMS (ESI): m/z, Calcd. for C H O N S [M+H] :
2
2
25
4
4
41.1591, Found 441.1600.
5
.2.26 (2-(3-Hydroxy-8-azabicyclo[3.2.1]octan-8-yl)-5-(naphthalen-2-
ylcarbamoyl)thiazol-4-yl)glycine (9b-10)
Following the preparation protocol of compound 9a, starting from compound 8b-10
(
7
1
80 mg, 0.17 mmol), the title compound 9b-10 was obtained as a yellow solid (54 mg,
1
4%); mp 159-161 °C; H NMR (400 MHz, DMSO-d ) δ (ppm) 12.53 (s, 1H), 8.99 (s,
6
H), 8.28 (d, J = 2.0 Hz, 1H), 8.06 (t, J = 6.0 Hz, 1H), 7.82 - 7.68 (m, 4H), 7.49 - 7.28
(
2
m, 2H), 4.76 (brs, 1H), 4.23 - 4.08 (m, 4H), 3.92 (brs, 1H), 2.36 - 2.24 (m, 2H), 2.16 -
.04 (m, 2H), 2.03 - 1.91 (m, 2H), 1.79 - 1.67 (m, 2H); C NMR (150 MHz, DMSO-
1
3
d6) δ (ppm) 172.55, 165.20, 163.61, 163.28, 138.29, 133.98, 129.72, 128.16, 127.81,
1
27.54, 126.58, 124.48, 121.57, 115.87, 82.25, 63.21, 57.12, 45.32, 36.95, 28.28;
+
HRMS (ESI): m/z, Calcd. for C H O N S [M+H] : 453.1591, Found 453.1571.
2
3
25
4
4
5
(
.2.27
9b-11)
To a stirred solution of 8b-11 (193 mg, 0.39 mmol) in actone (2 mL) was added 1M
(5-(Naphthalen-2-ylcarbamoyl)-2-(4-oxopiperidin-1-yl)thiazol-4-yl)glycine
HCl aqueous solution (2 mL), the reaction mixture was then refluxed for 2 h. Then the
mixture was adjusted pH to 7 with saturated Na CO solution. The aqueous layer was
2
3
washed with EA (5 mL × 3), crystallized and filtered to obtain 9b-11 as a light pink
1
solid (81 mg, 49%); mp 175-177 °C; H NMR (400 MHz, DMSO-d ) δ (ppm) 9.02 (s,
6
1
2
H), 8.32 (d, J = 2.0 Hz, 1H), 8.04 (t, J = 4.4, 1H), 7.81 - 7.69 (m, 4H), 7.46 - 7.29 (m,
H), 3.86 (t, J = 6.3 Hz, 4H), 3.68 (d, J = 4.3 Hz, 2H), 2.57 (t, J = 6.2 Hz, 4H); C
1
3
NMR (150 MHz, DMSO-d ) δ (ppm) 207.12, 171.92, 168.70, 163.16, 163.05, 138.67,
6
1
4
4
34.09, 129.58, 128.08, 127.79, 127.53, 126.51, 124.28, 121.52, 115.44, 81.87, 48.94,
+
5.97, 40.54; HRMS (ESI): m/z, Calcd. for C H O N S [M+H] : 425.1278, Found
2
1
21
4
4
25.1259.
5
.2.28 (2-(4-Cyanopiperidin-1-yl)-5-(naphthalen-2-ylcarbamoyl)thiazol-4-yl)glycine
(9b-12)
Following the preparation protocol of compound 9a, starting from compound 8b-12
(
6
50 mg, 0.11 mmol), the title compound 9b-12 was obtained as a white solid (28 mg,
1
0%); mp 146-148 °C; H NMR (400 MHz, DMSO-d ) δ (ppm) 9.07 (s, 1H), 8.28 (d,
6
J = 2.0 Hz, 1H), 7.83 - 7.67 (m, 3H), 7.47 - 7.31 (m, 2H), 4.13 (s, 2H), 3.77 - 3.62 (m,
2
H), 3.50 - 3.37 (m, 2H), 3.23 - 3.12 (m, 1H), 2.08 - 1.95 (m, 2H), 1.90 - 1.76 (m, 2H);
1
3
C NMR (150 MHz, DMSO-d ) δ (ppm) 172.52, 169.22, 163.23, 163.20, 138.15,
6
1
33.95, 129.78, 128.19, 127.83, 127.56, 126.61, 124.55, 122.12, 121.59, 116.03, 82.76,

+
4
4
6.25, 45.28, 27.63, 25.43; HRMS (ESI): m/z, Calcd. for C H O N S [M+H] :
22 22 3 5
36.1438, Found 436.1441.
5
.2.29 (5-(Naphthalen-2-ylcarbamoyl)-2-(4-(trifluoromethyl)piperidin-1-yl)thiazol-4-
yl)glycine (9b-13)
Following the preparation protocol of compound 9a, starting from compound 8b-13
(
7
100 mg, 0.20 mmol), the title compound 9b-13 was obtained as a yellow solid (71 mg,
1
5%); mp 160-162 °C; H NMR (400 MHz, DMSO-d ) δ (ppm) 9.02 (s, 1H), 8.29 (d,
6
J = 2.0 Hz, 1H), 8.03 (t, J = 5.2 Hz, 1H), 7.84 - 7.68 (m, 4H), 7.47 - 7.30 (m, 2H), 4.09
-
(
3.97 (m, 2H), 3.94 (d, J = 5.1 Hz, 2H), 3.24 - 3.11 (m, 2H), 2.67 (s, 1H), 2.01 - 1.88
1
3
m, 2H), 1.60 - 1.43 (m, 2H); C NMR (150 MHz, DMSO-d ) δ (ppm) 172.48, 169.21,
6
1
1
63.21, 163.18, 138.35, 134.01, 127.93 (q), 129.70, 128.15, 127.80, 127.55, 126.56,
24.44, 121.57, 115.80, 82.25, 46.67, 46.60, 38.85 (q), 23.87; HRMS (ESI): m/z, Calcd.
+
for C H O N SF [M+H] : 479.1359, Found 479.1353.
2
2
22
3
4
3
5
.2.30
(2-(4,4-Dimethylpiperidin-1-yl)-5-(naphthalen-2-ylcarbamoyl)thiazol-4-
yl)glycine (9b-14)
Following the preparation protocol of compound 9a, starting from compound 8b-14
(
8
1
100 mg, 0.21 mmol), the title compound 9b-14 was obtained as a white solid (78 mg,
1
3%); mp 172-174 °C; H NMR (400 MHz, DMSO-d ) δ (ppm) 12.58 (s, 1H), 9.01 (s,
6
H), 8.29 (d, J = 2.0 Hz, 1H), 8.05 (t, J = 5.9 Hz, 1H), 7.85 - 7.68 (m, 4H), 7.48 - 7.31
(
m, 2H), 4.12 (d, J = 5.9 Hz, 2H), 3.51 (t, J = 5.9 Hz, 4H), 1.43 (t, J = 5.9 Hz, 4H), 0.99
1
3
(s, 7H); C NMR (150 MHz, DMSO-d ) δ (ppm) 172.55, 169.21, 163.47, 163.26,
6
1
4
4
38.26, 133.98, 129.73, 128.17, 127.81, 127.54, 126.58, 124.49, 121.57, 115.89, 82.22,
+
5.26, 37.56, 29.25, 27.77; HRMS (ESI): m/z, Calcd. for C H O N S [M+H] :
2
3
27
3
4
39.1798, Found 439.1791.
5
.2.31 (2-(4-Hydroxy-4-methylpiperidin-1-yl)-5-(naphthalen-2-ylcarbamoyl)thiazol-4-
yl)glycine (9b-15)
Following the preparation protocol of compound 9a, starting from compound 8b-15
(
9
100 mg, 0.21 mmol), the title compound 9b-15 was obtained as a white solid (85 mg,
1
0%); mp 180-182 °C; H NMR (400 MHz, DMSO-d ) δ (ppm) 8.99 (s, 1H), 8.28 (d,
6
J = 2.0 Hz, 1H), 8.04 (s, 1H), 7.83 - 7.69 (m, 4H), 7.48 - 7.31 (m, 2H), 4.12 (d, J = 3.6
1
3
Hz, 2H), 3.75 - 3.59 (m, 2H), 3.47 - 3.41 (m, 2H), 1.64 - 1.50 (m, 4H), 1.18 (s, 3H); C
NMR (150 MHz, DMSO-d ) δ (ppm) 172.37, 169.06, 163.46, 163.17, 138.49, 134.04,
6
1
4
29.63, 128.11, 127.79, 127.53, 126.53, 124.35, 121.54, 115.62, 81.68, 66.37, 46.93,
+
4.85, 37.72, 30.06; HRMS (ESI): m/z, Calcd. for C H O N S [M+H] : 441.1591,
2
2
25
4
4
Found 425.1604.
5
.2.32 (2-(3,6-Dihydropyridin-1(2H)-yl)-5-(naphthalen-2-ylcarbamoyl)thiazol-4-
yl)glycine (9b-16)
Following the preparation protocol of compound 9a, starting from compound 8b-16
(
7
50 mg, 0.11 mmol), the title compound 9b-16 was obtained as a white solid (34 mg,
1
4%); mp 131-133 °C; H NMR (400 MHz, DMSO-d ) δ (ppm) 12.53 (s, 1H), 9.06 (s,
6

1
H), 8.29 (d, J = 2.0 Hz, 1H), 8.06 (t, J = 6.0 Hz, 1H), 7.84 - 7.69 (m, 4H), 7.50 - 7.31
(
4
m, 2H), 5.93 (d, J = 10.4 Hz, 1H), 5.82 (d, J = 10.4 Hz, 1H), 4.14 (d, J = 5.5 Hz, 2H),
.00 - 3.92 (m, 2H), 3.64 (t, J = 5.8 Hz, 2H), 2.26 (brs, 2H); C NMR (150 MHz,
1
3
DMSO-d ) δ (ppm) 172.56, 169.22, 163.28, 163.26, 138.22, 133.97, 129.75, 128.18,
6
1
4
4
27.82, 127.55, 126.59, 125.72, 124.52, 123.93, 121.60, 115.97, 82.23, 47.02, 45.27,
+
4.53, 24.40; HRMS (ESI): m/z, Calcd. for C H O N S [M+H] : 409.1329, Found
2
1
21
3
4
09.1311.
5
.2.33 (5-(Naphthalen-2-ylcarbamoyl)-2-(piperazin-1-yl)thiazol-4-yl)glycine (9g)
To a stirred solution of 9e (46 mg, 0.08 mmol) in DCM (2 mL) was added TFA (0.1
mL) dropwise, the reaction mixture was then allowed to stir at room temperature for 5
h. DCM and excess TFA were removed under reduced pressure, diethyl ether (2 mL)
were then added to the residue. After filtration and drying under high vacuum,
1
compound 9g was obtained as a yellow solid (35 mg, 94%); mp 131-133 °C; H NMR
(
400 MHz, DMSO-d ) δ (ppm) 9.23 (s, 1H), 8.29 (d, J = 2.0 Hz, 1H), 8.06 (t, J = 5.9
6
Hz, 1H), 7.84 - 7.70 (m, 4H), 7.47 - 7.33 (m, 2H), 4.14 (d, J = 6.0 Hz, 2H), 3.85 - 3.65
m, 4H), 3.29 - 3.18 (m, 4H).
(
5
(
.2.34 (2-(3-Methylpiperazin-1-yl)-5-(naphthalen-2-ylcarbamoyl)thiazol-4-yl)glycine
9h)
Following the preparation protocol of compound 9g, starting from compound 9f (50
mg, 0.11 mmol), the title compound 9h was obtained as a yellow solid (30 mg, 64%);
1
mp 139-140 °C; H NMR (400 MHz, DMSO-d ) δ (ppm) 9.18 (s, 1H), 8.28 (d, J = 2.0
6
Hz, 1H), 8.06 (t, J = 5.1 Hz, 1H), 7.84 - 7.79 (m, 2H), 7.77 (d, J = 8.2 Hz, 1H), 7.72
(dd, J = 8.9, 2.1 Hz, 1H), 7.48 - 7.32 (m, 2H), 4.16 (d, J = 4.9 Hz, 2H), 3.97 (t, J = 12.7
Hz, 2H), 3.51 - 3.43 (m, 2H), 3.41 - 3.35 (m, 1H), 3.27 - 3.14 (m, 2H), 1.28 (d, J = 6.5
+
Hz, 3H); HRMS (ESI): m/z, Calcd. for C H O N S [M+H] : 426.1594, Found
2
1
24
3
5
4
26.1607.
5
.2.35
(2-(3,3-Dimethylpiperazin-1-yl)-5-(naphthalen-2-ylcarbamoyl)thiazol-4-
yl)glycine (9i)
Following the preparation protocol of compound 9a, starting from compound 8i (79
mg, 0.19 mmol), the title compound 9i was obtained as a white solid (70 mg, 94%); mp
1
1
1
7
79-181 °C; H NMR (400 MHz, DMSO-d ) δ (ppm) 8.98 (s, 1H), 8.28 (d, J = 2.0 Hz,
6
H), 8.04 (t, J = 5.8 Hz, 1H), 7.79 (dd, J = 8.5, 3.1 Hz, 2H), 7.75 (d, J = 8.2 Hz, 1H),
.71 (dd, J = 8.9, 2.1 Hz, 1H), 7.48 - 7.29 (m, 2H), 4.07 (d, J = 5.7 Hz, 2H), 3.50 - 3.42
m, 2H), 3.26 (s, 2H), 2.89 (t, J = 5.1 Hz, 2H), 1.10 (s, 6H); C NMR (150 MHz,
1
3
(
DMSO-d ) δ (ppm) 172.54, 169.60, 163.36, 163.23, 138.27, 133.98, 129.72, 128.17,
6
1
4
4
27.81, 127.54, 126.59, 124.48, 121.54, 115.84, 81.97, 58.27, 50.46, 48.02, 45.50,
+
0.53, 25.23 ; HRMS (ESI): m/z, Calcd. for C H O N S [M+H] : 440.1751, Found
2
2
26
3
5
40.1764.
5
4
.2.36 (2-((3S,5R)-3,5-Dimethylpiperazin-1-yl)-5-(naphthalen-2-ylcarbamoyl)thiazol-
-yl)glycine (9j)

Following the preparation protocol of compound 9a, starting from compound 8j (140
mg, 0.30 mmol), the title compound 9j was obtained as a white solid (47 mg, 36%); mp
1
1
1
80-182 °C H NMR (400 MHz, DMSO-d ) δ (ppm) 8.98 (s, 1H), 8.29 (d, J = 2.0 Hz,
6
H), 8.03 (t, J = 5.5 Hz, 1H), 7.82 - 7.69 (m, 4H), 7.46 - 7.31 (m, 2H), 4.04 (d, J = 5.4
Hz, 2H), 3.73 (brs, 2H), 2.86 - 2.76 (m, 2H), 2.69 - 2.59 (m, 2H), 1.04 (d, J = 6.2 Hz,
1
3
6
H); C NMR (150 MHz, DMSO-d ) δ (ppm) 172.45, 169.18, 163.30, 163.19, 138.36,
6
1
5
34.00, 129.68, 128.16, 127.80, 127.55, 126.56, 124.42, 121.51, 115.73, 81.94, 54.13,
+
0.26, 46.28, 19.19; HRMS (ESI): m/z, Calcd. for C H O N S [M+H] : 440.1751,
2
2
26
3
5
Found 440.1752.
5
(
.2.37 (2-(4-Methylpiperazin-1-yl)-5-(naphthalen-2-ylcarbamoyl)thiazol-4-yl)glycine
9k)
Following the preparation protocol of compound 9a, starting from compound 8k
(
7
130 mg, 0.30 mmol), the title compound 9k was obtained as a white solid (91 mg,
1
7%); mp 215-217 °C; H NMR (400 MHz, DMSO-d ) δ (ppm) 9.08 (s, 1H), 8.29 (d,
6
J = 2.0 Hz, 1H), 8.05 (t, J = 5.9 Hz, 1H), 7.85 - 7.78 (m, 2H), 7.76 (d, J = 8.1 Hz, 1H),
7
=
.72 (dd, J = 8.9, 2.1 Hz, 1H), 7.47 - 7.32 (m, 2H), 4.12 (d, J = 5.8 Hz, 2H), 3.49 (t, J
4.8, 4H), 2.44 (t, J = 5.1 Hz, 4H), 2.24 (s, 3H); C NMR (150 MHz, DMSO-d ) δ
6
1
3
(ppm) 172.52, 169.45, 163.27, 163.24, 138.20, 133.97, 129.76, 128.18, 127.82, 127.56,
1
26.59, 124.52, 121.60, 115.98, 82.56, 53.99, 47.65, 46.09, 45.44; HRMS (ESI): m/z,
+
Calcd. for C H O N S [M+H] : 426.1594, Found 426.1594.
2
1
24
3
5
5
.2.38
(2-(3,4-Dimethylpiperazin-1-yl)-5-(naphthalen-2-ylcarbamoyl)thiazol-4-
yl)glycine (9l)
Following the preparation protocol of compound 9a, starting from compound 8l (130
mg, 0.28 mmol), the title compound 9l was obtained as a white solid (112 mg, 97%);
1
mp 187-189 °C; H NMR (400 MHz, DMSO-d ) δ 9.04 (s, 1H), 8.28 (d, J = 2.0 Hz,
6
1
=
H), 8.04 (t, J = 5.9 Hz, 1H), 7.84 - 7.77 (m, 2H), 7.76 (d, J = 8.1 Hz, 1H), 7.72 (dd, J
8.8, 2.1 Hz, 1H), 7.48 - 7.31 (m, 2H), 4.13 (d, J = 5.8 Hz, 2H), 3.83 - 3.61 (m, 2H),
3
.27 - 3.21 (m, 1H), 2.96 - 2.78 (m, 2H), 2.29 - 2.11 (m, 5H), 1.05 (d, J = 6.1 Hz, 3H);
1
3
C NMR (150 MHz, DMSO-d ) δ (ppm) 172.55, 169.20, 163.29, 163.24, 138.19,
6
1
5
33.97, 129.76, 128.20, 127.82, 127.56, 126.60, 124.53, 121.56, 115.96, 82.58, 56.85,
3.91, 53.83, 47.67, 45.32, 42.35, 16.47 ; HRMS (ESI): m/z, Calcd. for C H O N S
2
2
26
3
5
+
[M+H] : 440.1751, Found 440.1751.
5
.2.39
(5-(Naphthalen-2-ylcarbamoyl)-2-(3-oxopiperazin-1-yl)thiazol-4-yl)glycine
(9m)
Following the preparation protocol of compound 9a, starting from compound 8m (30
mg, 0.06 mmol), the title compound 9m was obtained as a yellow solid (22 mg, 78%);
1
mp 108-110 °C; H NMR (400 MHz, DMSO-d ) δ(ppm)δ 9.13 (s, 1H), 8.65 (brs, 1H),
6
8
2
3
1
.29 (t, J = 2.6 Hz, 2H), 8.06 (q, J = 5.6 Hz, 1H), 7.84 - 7.75 (m, 3H), 7.73 (dd, J = 8.9,
.0 Hz, 1H), 7.47 - 7.31 (m, 2H), 4.16 (m, 2H), 4.01 (s, 1H), 3.82 (s, 2H), 3.67 (m, 1H),
1
3
.40 - 3.35 (m, 1H), 3.16 (m, 1H); C NMR (150 MHz, DMSO-d ) δ (ppm) 168.15,
6
68.10, 165.90, 163.22, 163.05, 138.17, 133.96, 129.77, 128.19, 127.82, 127.56,

1
26.60, 124.55, 121.60, 116.03, 82.70, 63.27, 50.26, 45.82, 44.62; HRMS (ESI): m/z,
+
Calcd. for C H O N S [M+H] : 426.1231, Found 426.1232.
2
0
20
4
5
5
.2.40 (2-Morpholino-5-(naphthalen-2-ylcarbamoyl)thiazol-4-yl)glycine (9n)
Following the preparation protocol of compound 9a, starting from compound 8n
(
4
130 mg, 0.29 mmol), the title compound 9n was obtained as a white solid (55 mg,
1
5%); mp 174-176 °C; H NMR (400 MHz, DMSO-d ) δ (ppm) 9.03 (s, 1H), 8.31 (d,
6
J = 2.0 Hz, 1H), 8.01 (t, J = 4.4, 1H), 7.82 - 7.73 (m, 3H), 7.72 (dd, J = 8.9, 2.0 Hz,
1
4
H), 7.47 - 7.28 (m, 2H), 3.72 (t, J = 4.9 Hz, 4H), 3.67 (d, J = 4.2 Hz, 2H), 3.48 (t, J =
1
3
.9 Hz, 4H); C NMR (150 MHz, DMSO-d ) δ (ppm) 172.53, 169.82, 163.27, 163.14,
6
1
6
38.13, 133.95, 129.79, 128.19, 127.83, 127.57, 126.61, 124.57, 121.62, 116.08, 82.72,
+
5.76, 47.78, 45.27; HRMS (ESI): m/z, Calcd. for C H O N S [M+H] : 413.1278,
2
0
21
4
4
Found 413.1276.
5
.2.41 (5-(Naphthalen-2-ylcarbamoyl)-2-thiomorpholinothiazol-4-yl)glycine (9o)
Following the preparation protocol of compound 9a, starting from compound 8o (70
mg, 0.15 mmol), the title compound 9o was obtained as a white solid (63 mg,96%); mp
1
1
31-133 °C; H NMR (400 MHz, DMSO-d ) δ (ppm) 12.60 (s, 1H), 9.03 (s, 1H), 8.31
6
(
4
d, J = 2.0 Hz, 1H), 8.05 (t, J = 6.0 Hz, 1H), 7.81 - 7.70 (m, 4H), 7.46 - 7.29 (m, 2H),
.12 (d, J = 5.6 Hz, 2H), 3.89 - 3.72 (m, 4H), 2.80 - 2.66 (m, 4H); C NMR (150 MHz,
1
3
DMSO-d ) δ (ppm) 172.51, 169.04, 163.24, 163.19, 138.17, 133.96, 129.78, 128.20,
6
1
27.83, 127.56, 126.61, 124.55, 121.57, 115.99, 82.65, 50.61, 45.30, 26.21; HRMS
+
(ESI): m/z, Calcd. for C H O N S [M+H] : 429.1050, Found 429.1044.
2
0
21
3
4 2
5
.2.42 (5-(Naphthalen-2-ylcarbamoyl)-2-(2-oxa-6-azaspiro[3.3]heptan-6-yl)thiazol-4-
yl)glycine (9p)
Following the preparation protocol of compound 9a, starting from compound 8p (51
mg, 0.11 mmol), the title compound 9p was obtained as a yellow solid (36 mg, 75%);
1
mp 181-183 °C; H NMR (400 MHz, DMSO-d ) δ (ppm) 9.05 (s, 1H), 8.28 (d, J = 2.0
6
Hz, 1H), 8.08 (s, 1H), 7.84 - 7.78 (m, 2H), 7.76 (d, J = 8.2 Hz, 1H), 7.72 (dd, J = 8.9,
2
.1 Hz, 1H), 7.47 - 7.31 (m, 2H), 4.73 (s, 4H), 4.27 (s, 4H), 4.12 (s, 2H); HRMS
+
(ESI): m/z, Calcd. for C H O N S [M+H] : 425.1278, Found 425.1276.
2
1
21
4
4
5
.2.43
(5-(Naphthalen-2-ylcarbamoyl)-2-(2-azaspiro[3.3]heptan-2-yl)thiazol-4-
yl)glycine (9q)
Following the preparation protocol of compound 9a, starting from compound 8q (50
mg, 0.11 mmol), the title compound 9q was obtained as a yellow solid (46 mg, 95%);
1
mp 144-146 °C; H NMR (400 MHz, DMSO-d ) δ (ppm) 9.03 (s, 1H), 8.28 (d, J = 2.1
6
Hz, 1H), 8.08 (t, J = 5.5 Hz, 1H), 7.84 - 7.77 (m, 2H), 7.76 (d, J = 8.1 Hz, 1H), 7.72
(
2
dd, J = 8.9, 2.1 Hz, 1H), 7.47 - 7.31 (m, 2H), 4.11 (d, J = 4.5 Hz, 2H), 4.06 (s, 4H),
.22 (t, J = 7.6 Hz, 4H), 1.80 (p, J = 7.7 Hz, 2H); C NMR (150 MHz, DMSO-d ) δ
6
1
3
(ppm) 172.46, 169.35, 163.57, 163.25, 138.19, 133.96, 129.76, 128.18, 127.82, 127.56,
1
26.60, 124.53, 121.59, 115.99, 83.17, 64.94, 45.30, 32.73, 32.11, 16.10; HRMS (ESI):
+
m/z, Calcd. for C H O N S [M+H] : 423.1485, Found 423.1488.
2
2
23
3
4

5
.2.44
(5-([1,1'-Biphenyl]-4-ylcarbamoyl)-2-(2-oxa-6-azaspiro[3.3]heptan-6-
yl)thiazol-4-yl)glycine (9r)
Following the preparation protocol of compound 9a, starting from compound 8r (75
mg, 0.16 mmol), the title compound 9r was obtained as a white solid (64 mg, 90%);
1
mp 144-146 °C; H NMR (400 MHz, DMSO-d ) δ (ppm) 8.94 (s, 1H), 8.03 (s, 1H),
6
7
7
.74 (d, J= 8.4 Hz, 2H), 7.64 (d, J = 7.7 Hz, 2H), 7.57 (d, J = 8.4 Hz, 2H), 7.43 (t, J =
.6 Hz, 2H), 7.31 (t, J = 7.4 Hz, 1H), 4.73 (s, 4H), 4.26 (s, 4H), 4.11 (s, 2H); C NMR
1
3
(
150 MHz, DMSO-d ) δ (ppm) 172.44, 169.19, 163.41, 163.06, 140.37, 140.03, 134.29,
6
1
3
29.34, 127.32, 127.28, 126.97, 126.79, 126.59, 120.72, 83.30, 79.89, 62.02, 45.30,
+
0.51; HRMS (ESI): m/z, Calcd. for C H O N S [M+H] : 451.1435, Found 451.1408.
2
3
23
4
4
5
.2.45
(5-((3,4-Dichlorophenyl)carbamoyl)-2-(2-oxa-6-azaspiro[3.3]heptan-6-
yl)thiazol-4-yl)glycine (9s)
Following the preparation protocol of compound 9a, starting from compound 8s (75
mg, 0.16mmol), the title compound 9s was obtained as a white solid (69 mg, 97%); mp
1
1
1
4
47-149 °C; H NMR (400 MHz, DMSO-d ) δ (ppm) 9.08 (s, 1H), 8.06 (d, J = 2.4 Hz,
6
H), 8.03 (t, J = 5.8 Hz, 1H), 7.60 (dd, J = 8.9, 2.5 Hz, 1H), 7.48 (d, J = 8.8 Hz, 1H),
.72 (s, 4H), 4.26 (s, 4H), 4.07 (d, J = 5.8 Hz, 2H); C NMR (150 MHz, DMSO-d ) δ
1
3
6
(ppm) 172.26, 169.32, 163.96, 162.91, 140.83, 130.99, 130.64, 123.72, 121.15, 120.03,
8
2.75, 79.85, 61.97, 45.52, 30.18; HRMS (ESI): m/z, Calcd. for C H O N SCl
1
7
17
4
4
2
+
[M+H] : 443.0342, Found 443.0341.
5
.2.46
(5-((3-Isopropylphenyl)carbamoyl)-2-(2-oxa-6-azaspiro[3.3]heptan-6-
yl)thiazol-4-yl)glycine (9t)
Following the preparation protocol of compound 9a, starting from compound 8t (100
mg, 0.23 mmol), the title compound 9t was obtained as a white solid (76 mg, 81%); mp
1
1
36-138 °C; H NMR (400 MHz, DMSO-d ) δ (ppm) 12.52 (s, 1H), 8.75 (s, 1H), 7.98
6
(t, J = 6.0 Hz, 1H), 7.52 (t, J = 1.9 Hz, 1H), 7.45 (dd, J = 7.6, 2.1 Hz, 1H), 7.14 (t, J =
7
2
.8 Hz, 1H), 6.85 (d, J = 7.6 Hz, 1H), 4.72 (s, 4H), 4.25 (s, 4H), 4.09 (d, J = 5.9 Hz,
H), 2.82 (hept, J = 6.8 Hz, 1H), 1.19 (d, J = 6.9 Hz, 6H); C NMR (150 MHz, DMSO-
1
3
d6) δ (ppm) 172.48, 169.11, 163.21, 163.06, 148.93, 140.41, 128.61, 120.84, 118.43,
1
18.04, 83.21, 79.89, 62.01, 45.47, 34.01, 28.90, 24.39; HRMS (ESI): m/z, Calcd. for
+
C H O N S [M+H] : 417.1591, Found 417.1597.
2
0
25
4
4
5
.2.47
4-((2-Amino-2-oxoethyl)amino)-2-(4-hydroxy-4-methylpiperidin-1-yl)-N-
(naphthalen-2-yl)thiazole-5-carboxamide (10)
To a stirred solution of 9b-15 (367 mg, 0.80 mmol) in THF (20 mL) was added EDCI
335 mg, 1.75 mmol) and HOBt (237 mg, 1.75 mmol), the reaction mixture was then
(
allowed to stir at room temperature for 1 h. Ammonia (145 mg,2.4 mmol) was added
and stirred at room temperature for 2 h. The reaction solution was diluted with EA (5
mL) and washed with saturated NaCl solution (10 mL × 2) and water (10 mL × 2). The
organic layer was concentrated and the crude product was purified by column
chromatography (D/M = 50:1-20:1) to give the title compound 10 (218 mg, 58%) as a

1
white solid; mp 121-123 °C; H NMR (400 MHz, Methanol-d ) δ (ppm) 8.04 (s, 1H),
4
7
.81 - 7.72 (m, 3H), 7.64 - 7.58 (m, 1H), 7.46 - 7.32 (m, 2H), 4.13 - 4.09 (m, 2H), 3.79
(
d, J = 13.1 Hz, 2H), 3.53 - 3.45 (m, 3H), 1.73 - 1.60 (m, 4H), 1.27 (s, 3H); HRMS
+
(ESI): m/z, Calcd. for C H O N S [M+H] : 440.1751, Found 440.1728.
2
2
26
3
5
5
2
.2.48 2-(4-Amino-4-hydroxypiperidin-1-yl)-4-((cyanomethyl)amino)-N-(naphthalen-
-yl)thiazole-5-carboxamide (11)
To a stirred solution of 10 (112 mg, 0.25 mmol) in DCM (5 mL) was added DBU
(197 mg, 1.27 mmol) and stirred at room temperature for 10 min. Ethyl
dichlorophosphate (210 mg, 1.27 mmol) was added. The reaction mixture was then
allowed to stir at room temperature for 3 h. The reaction solution was diluted with DCM
(5 mL) and washed with saturated NaCl solution (10 mL × 2) and water (10 mL × 2).
The organic layer was concentrated and the crude product was purified by column
chromatography (P/E= 1:1) to give the title compound 11 (25 mg, 23%) as a white
1
solid; mp 149-151 °C; H NMR (500 MHz, CDCl ) δ (ppm) 8.08 (s, 1H), 7.92 - 7.67
3
(
-
m, 4H), 7.52 - 7.34 (m, 3H), 6.66 (s, 1H), 4.37 (d, J = 6.2 Hz, 2H), 3.80 (s, 2H), 3.61
1
3
3.46 (m, 2H), 1.71 (s, 4H), 1.33 (s, 3H); C NMR (150MHz, DMSO-d ) δ (ppm)
6
1
1
69.13, 162.95, 161.91, 137.94, 133.87, 129.82, 128.20, 127.81, 127.53, 126.62,
24.63, 121.55, 119.15, 116.20, 84.04, 66.30, 44.97, 37.63, 32.11, 30.02; HRMS (ESI):
+
m/z, Calcd. for C H O N S [M+H] : 422.1645, Found 422.1652.
2
2
24
2
5
5
4
.2.49 3-(5-(Naphthalen-2-ylcarbamoyl)-2-(2-oxa-6-azaspiro[3.3]heptan-6-yl)thiazol-
-yl)propanoic acid (20)
Following the preparation protocol of compound 9a, starting from compound 19 (30
mg, 0.07 mmol), the title compound 20 was obtained as a yellow solid (19 mg, 67%);
1
mp 143-145 °C; H NMR (400 MHz, DMSO-d ) δ (ppm) 10.17 (s, 1H), 8.28 (d, J = 2.1
6
Hz, 1H), 7.90 - 7.77 (m, 3H), 7.69 (dd, J = 8.9, 2.1 Hz, 1H), 7.52 - 7.36 (m, 2H), 4.73
(
s, 4H), 4.26 (s, 4H), 3.07 (t, J = 7.2 Hz, 2H), 2.68 (t, J = 7.1 Hz, 2H); HRMS (ESI):
+
m/z, Calcd. for C H O N S [M+H] : 424.1326, Found 424.1340.
2
2
22
4
3
5
5
.3. Biological evaluation
.3.1. Protein expression and purification
The pET-28a-Pin1 plasmid was a gift from Professor Joseph P. Noel (The Salk
Institute for Biological Studies, La Jolla, California). The Pin1 gene fragment was
amplified with PCR and inserted into pET-28a plasmid after a His-tag and a Prescission
protease cleavage site (His-tag-Leu-Phe-Gln-Gly-Pro), enabling to remove the N-
terminal His-tag by Prescission protease. The Pin1 was expressed in E.coli BL21(DE3)
induced by IPTG. Cells were harvested via centrifugation and resuspended in 25 mM
Tris-HCl, 200 mM NaCl, pH 7.8. Following sonication at 4 °C, the crude protein
solution was sequencially purified with HisTrapTM HP, HitrapTM Q HP and Superdex
G75 (GE health care). After first purified with HisTrapTM HP, the collected Pin1 was
incubated with Precission at 4 °C for 16 h.