Br?nsted acid-promoted ‘on–water’ C(sp3)-H functionalization for the synthesis of isoindolinone/[1,2,4]triazolo[1,5-a]pyrimidine derivatives targeting the SKP2-CKS1 interaction

Article abstract of DOI:10.1016/j.cclet.2019.07.019

The isoindolinone and biaryl scaffolds are prevalent in natural products and drug molecules, which have showed broad and interesting biological activities. The efficient construction of such hybridized molecules and biological evaluation are of great interest to medicinal chemistry community. In this communication, we report an efficient Br?nsted acid-promoted C(sp³)-H functionalization approach that enables the rapid construction of biologically important isoindolinone/[1,2,4]triazolo[1,5-a]pyrimidine hybrids from 5-methyl-7-(2,4,6-trimethoxyphenyl)-[1,2,4]triazolo[1,5-a]pyrimidine, 2-formylbenzoic acid and various anilines. The title compounds were generated in high to excellent yields (up to 96%) regardless of the electronic nature and steric effects of the substituents. In this reaction, an isoindolinone scaffold, one C[sbnd]C single bond, and two C[sbnd]N bonds were formed simultaneously with high atom economy. In this work, we have envisioned that the methyl group linked to the electron-deficient N-heterocycles could be used as a new synthetic handle for late-state diversification and may have broad applications in the field of organic and medicinal chemistry. Besides, the title compounds have exhibited promising activity against the SKP2-CKS1 interaction.

Full text of DOI:10.1016/j.cclet.2019.07.019

Accepted Manuscript
Title: Brønsted acid-promoted ‘on–water’ C(sp3)-H
functionalization for the synthesis of
isoindolinone/[1,2,4]triazolo[1,5-a]pyrimidine derivatives
targeting the SKP2-CKS1 interaction
Authors: Shuo Yuan, Sixi Wang, Min Zhao, Danqing Zhang,
Jinjie Chen, Jian-Xin Li, Jingya Zhang, Yihui Song, Jinyi
Wang, Bin Yu, Hongmin Liu
PII:
DOI:
Reference:
S1001-8417(19)30401-2
https://doi.org/10.1016/j.cclet.2019.07.019
CCLET 5102
To appear in:
Chinese Chemical Letters
Accepted date:
8 July 2019
Please cite this article as: Yuan S, Wang S, Zhao M, Zhang D, Chen J, Li J-Xin,
Zhang J, Song Y, Wang J, Yu B, Liu H, Brønsted acid-promoted ‘on–water’ C(sp3)-
H functionalization for the synthesis of isoindolinone/[1,2,4]triazolo[1,5-a]pyrimidine
derivatives targeting the SKP2-CKS1 interaction, Chinese Chemical Letters (2019),
https://doi.org/10.1016/j.cclet.2019.07.019
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Communication
Brønsted acid-promoted ‘on–water’ C(sp3)-H functionalization for the synthesis of
isoindolinone/[1,2,4]triazolo[1,5-a]pyrimidine derivatives targeting the SKP2-CKS1
interaction
Shuo Yuan, Sixi Wang, Min Zhao, Danqing Zhang, Jinjie Chen, Jian-Xin Li, Jingya Zhang, Yihui Song,
Jinyi Wang, Bin Yu, Hongmin Liu*
School of Pharmaceutical Sciences, Zhengzhou University, Zhengzhou 450001, China
Corresponding authors.
E-mail addresses: yubin@zzu.edu.cn (B. Yu); liuhm@zzu.edu.cn (H.M. Liu)
ARTICLE INFO
Article history:
Received 19 May 2019
Received in revised form 1 July 2019
Accepted 5 July 2019
Available online
Graphical abstract
New isoindolinone/[1,2,4]triazolo[1,5-a]pyrimidine derivatives were efficiently synthesized in good to excellent yields
through the Brønsted acid-promoted ‘on–water’ C(sp3)-H functionalization and capable of inhibiting the SKP2-CKS1
interaction. The title compounds could be used as hit compounds to design more potent small–molecule inhibitors targeting
the SKP2-CKS1 interaction.
ABSTRACT
The isoindolinone and biaryl scaffolds are prevalent in natural products and drug molecules, which have showed broad and interesting biological activities. The
efficient construction of such hybridized molecules and biological evaluation are of great interest to medicinal chemistry community. In this communication, we
report an efficient Brønsted acid-promoted C(sp3)-H functionalization approach that enables the rapid construction of biologically important
isoindolinone/[1,2,4]triazolo[1,5-a]pyrimidine hybrids from 5-methyl-7-(2,4,6-trimethoxyphenyl)-[1,2,4]triazolo[1,5-a]pyrimidine, 2-formylbenzoic acid and
various anilines. The title compounds were generated in high to excellent yields (up to 96%) regardless of the electronic nature and steric effects of the
substituents. In this reaction, an isoindolinone scaffold, one C-C single bond, and two C-N bonds were formed simultaneously with high atom economy. In this
work, we have envisioned that the methyl group linked to the electron-deficient N-heterocycles could be used as a new synthetic handle for late-state
diversification and may have broad applications in the field of organic and medicinal chemistry. Besides, the title compounds have exhibited promising activity
against the SKP2-CKS1 interaction.
———

Keywords:
Isoindolinone
[1,2,4]Triazolo[1,5-a]pyrimidine
Biaryl scaffold
C(sp3)-H activation
Molecular hybridization
SKP2-CKS1 interaction
N-Heterocyclic scaffolds, which exist widely in pharmaceuticals, agrochemicals and natural products, are considered as one of the
most abundant and integral motifs due to their interesting biological activities [1-4]. Among these N-heterocyclic compounds, the
isoindolinone frameworks are a privileged class of building blocks found in many pharmaceutically relevant molecules, such as
Pazinaclone (acting as a partial agonist at GABAA benzodiazepine receptors for the treatment of anxiety) [5], (R)-PD172938 (an
antagonist of the dopamine D4 receptor for the treatment of psychosis) [6], and Lenalidomide (an immunomodulatory agent for the
treatment of multiple myeloma) [7] (Fig. 1a). In addition, the [1,2,4]triazolo[1,5-a]pyrimidine fragment is another biologically important
scaffold, compounds bearing this scaffold have been found to possess diverse bioactivities [8-10]. Of particular interest are the
[1,2,4]triazolo[1,5-a]pyrimidine containing biaryl compounds, which possess unique structural features (e.g., chiral center) and
interesting biological profiles [11-13]. For instance, Cevipabulin (binding to the vinca domain of tubulin for the treatment of tumors)
[14], AF64394 (a selective GPR3 inverse agonist for the treatment of neuropsychiatric disorders) [15], compound A (a PA-PB1
interaction inhibitor for the treatment of influenza) [16] (Fig. 1b). Our group has also identified some biologically important triazole
fused pyrimidine derivatives with anticancer potential (Fig. 1c) [17-22]. For example, compound B effectively and reversibly inhibits
histone lysine specific demethylase 1 (LSD1/KDM1A) with an IC50 value of 49 nmol/L, and exhibits high selectivity to LSD1 over
monoamine oxidases (MAO-A/B) [18]. Compound C inactivates LSD1 (IC50 = 154 nmol/L) and also significantly inhibits migration of
lung cancer cells A549 and PC-921. Very recently, we have found that WS-383 effectively inhibits neddylation of Cul3 and Cul1 in cells
by interrupting the DCN1-UBC12 protein-protein interaction (IC50 = 11 nmol/L) [22].
In view of the biological importance of the isoindolinone and [1,2,4]triazolo[1,5-a]pyrimidine containing biaryl substructures, it would
be interesting to generate structurally new molecular entities (NMEs) possessing these two substructures through the molecular
hybridization approach, which is widely employed in drug discovery [23, 24]. It is generally believed that these hybridized molecules
have improved and/or new biological profiles. However, the molecular hybridization approach always relies on pluripotent functional
groups (e.g., –COOH, -NH2, -OH, etc.) to form hybridized molecules. Therefore, the question on how to efficiently combine these two
frameworks without these functional groups into one molecule has not been fully addressed.
Fig. 1. Bioactive molecules having isoindolinone and triazole-fused pyrimidine cores.
The C(sp3)-H functionalization is an emerging strategy in organic synthesis and has found broad applications in late-state
diversification of complex natural products and advanced drug molecules [25,26]. The methyl group attached to the electron-deficient
N-heterocycles have been found to undergo diverse transformations depending on the reaction conditions [27-32]. Following this work,

we herein report a Brønsted acid-promoted three-component cascade reaction for the synthesis of structurally complex
isoindolinone/[1,2,4]triazolo[1,5-a]pyrimidine hybrids on water based on the C(sp3)-H functionalization of the methyl group (Scheme
1).
Scheme 1. Our present work on C(sp3)-H functionalization.
The trimethoxyphenyl (TMP) group is prevalent in natural products and drug candidates, particularly widely found in tubulin inhibitors
[33]. Till date, a large number of TMP-based tubulin inhibitors have been identified, and some of them such as ZD6126, BNC-105p,
CKD-516, CA-1P, CA-4P, and AVE8062 are currently being evaluated in clinical trials for cancer therapy. Given the prevalence and
biological importance of the TMP moiety, 5-methyl-7-(2,4,6-trimethoxyphenyl)-[1,2,4]triazolo[1,5-a]pyrimidine (1a) was utilized with
high priority as a model substrate coupled with 2-formylbenzoic acid (2a) and aniline (3a), expecting to obtain biologically interesting
TMP containing isoindolinone/[1,2,4]triazolo[1,5-a]pyrimidine hybrids (Table 1). Firstly, several commonly used solvents were
examined (entries 1-7), the results indicated that water was the best solvent for this transformation, forming compound 4a in 43% yield
(entry 7). Other organic solvents were less efficient. Based on the possible reaction mechanism, we speculate that addition of acidic
additive may promote this conversion. We next examined the effects of different acidic additives on the reactivity (entries 8-14), and
found that acetic acid (AcOH) significantly improved the reaction efficiency, compound 4a was formed in 93% yield (entry 14). Other
Brønsted acids such as benzoic acid, trifluoroacetic acid (TFA), p-toluenesulfonic acid (TsOH) and trifluoromethanesulfonic acid (TfOH)
were less efficient (entries 9-12). Besides, decrease of additive (AcOH) loading led to decreased yield, compound 4a was obtained in
68% yield when using 5 equiv. of AcOH in H2O (entry 13). Considering the high reactivity, the reaction was not further optimized. Based
on above optimizations, the best reaction condition was that 1a (1.0 mmol), 2a (1.0 mmol) and aniline (3a, 1.0 mmol) were stirred in
H2O (2 mL) at 100 oC for 12 h in the presence of AcOH (10 mmol).
Table 1
Optimization of the reaction conditions. a
Entry
1
Solvent
MeOH
THF
Temp.(oC) Additive
Yield (%)b
70
-
N.D.
15
7
2
70
-
3
MeCN
DMF
DMSO
Dioxane
H2O
90
-
-
4
100
100
100
100
100
100
100
100
100
100
100
6
5
-
39
38
43
56
36
37
45
39
68c
93d
6
-
7
-
8
H2O
AcOH
PhCO2H
TFA
TsOH
TfOH
AcOH
AcOH
9
H2O
10
11
12
13
14
H2O
H2O
H2O
H2O
H2O
N.D.: Not detected.
a Reaction conditions: 1a (1 mmol), 2a (1 mmol), 3a (1 mmol), additive (1 mmol), solvent (2 mL), 12 h, 100 oC.
b NMR yields determined by 1H NMR using the triphenylmethane as an internal standard.
c 5 equiv. of AcOH was added.
d 10 equiv. of AcOH was added.

With the optimized reaction conditions in hand, we next examined the scope and generality of this three-component reaction (Scheme
2). The title compounds 4a-q were obtained in moderate to excellent yields (67%-96%) regardless of the electronic nature and steric
effects of the substituents. It should be noted that halogen atoms such as fluoro, chloro and bromo were well tolerated, and could be
potentially used for further functionalization. However, for aliphatic amine substrate, the yield decreased. Compound 4o was generated
in 67% yield when glycine ethyl ester was used, suggesting that other amino esters could also be employed to construct structurally
diverse and complex compounds for biological screening. Inspired by the reactivity observed, we then examined the reactivity of other
electron-deficient N-heterocycles bearing the methyl group. To our delight, compounds 4p and 4q were obtained in 81% and 86% yields,
respectively. Of note, heteroaromatic amines were incompatible with this protocol and failed to generate the corresponding products. All
1
new products were fully characterized by H and 13C NMR (please see the Supporting information for associated NMR spectra). The
current work has provided an example regarding the synthesis of complex molecules based on the C(sp3)-H functionalization.
Conceivably, other aliphatic amines and electron-deficient N-heterocycles bearing the activated methyl group could also be used under
the optimal condition to generate structurally new compounds.
Scheme 2. Substrate scope. Reaction conditions: 1a (1 mmol), 2a (1 mmol), 3 (1 mmol), AcOH (10 mmol), H2O (2 mL), 12 h, 100 oC.
Based on the above observation, we proposed a plausible mechanistic pathway as depicted in Scheme 3. Initially, the imine
intermediate A was formed through the acetic acid promoted condensation of aniline 3a with 2-formylbenzoic acid 2. Followed by
protonation, the activated imine B was afforded, the acidity of C-H bond in protonated compound 1a increased, forming the nucleophilic
intermediate C, the intermediate C then attacked the activated imine bond, followed by intramolecular cyclization reaction to generate
compound 4a. It should be noted that in this process, an isoindolinone framework was constructed, accompanying by formation of
consecutive C-C and C-N bonds.

Scheme 3. Proposed mechanism for the formation of 4a.
With these compounds in hand, we next tested the inhibitory activity of the synthesized compounds against the SKP2-CKS1 interaction
based on our established HTRF method. As shown in Fig. 2, some compounds showed acceptable inhibitory activity. Particularly,
compounds 4b, 4j, and 4n exhibited moderate inhibition against the SKP2-CKS1 interaction at 50 µmol/L with the inhibitory rates of
45%, 57%, and 54%, respectively. We believe that these compounds could be used as hit compounds to perform further structure-based
design for more potent inhibitors.
Fig. 2. The inhibitory activity of selected compounds against the SKP2-CKS1 interaction. GST-SKP2 and His6-CKS1 were treated with the
compounds at 50 µmol/L, and the inhibition rates were determined by the HTRF binding assay.
In summary, we have developed an efficient Brønsted acid-promoted C(sp3)-H functionalization approach that enables rapid access
to the structurally new isoindolinone/[1,2,4]triazolo[1,5-a]pyrimidine hybrids on water from 5-methyl-7-(2,4,6-trimethoxyphenyl)-
[1,2,4]triazolo[1,5-a]pyrimidine, 2-formylbenzoic acid and various anilines. The title compounds were generated in high to excellent
yields (up to 96%) regardless of the electronic nature and steric effects of the substituents. In this reaction, an isoindolinone scaffold, one
C-C single bond, and two C-N bonds were formed simultaneously with high atom economy. It is worth noting that the biologically
important biaryl, [1,2,4]triazolo[1,5-a]pyrimidine, and isoindolinone substructures were efficiently incorporated into one molecule
through the Brønsted acid-promoted C(sp3)-H functionalization approach. Biological evaluation indicated that some of these compounds
showed promising inhibitory activity against the SKP2-CKS1 interaction and therefore could be used to design more potent small-
molecule inhibitors targeting the SKP2-CKS1 interaction.
Acknowledgment
This work was supported by the National Natural Science Foundation of China (Nos. 81773562 and 81703326), China Postdoctoral
Science Foundation (Nos. 2018M630840 and 2019T120641), and Scientific Program of Henan Province (No. 182102310123). We
sincerely thank Dr. Erqing Li (Zhengzhou University) for analyzing the NMR spectra of some compounds.

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