The study on structure-activity relationship between chromone derivatives and inhibition of superoxide anion generating from human neutrophils

Article abstract of DOI:10.1016/j.bmcl.2021.127822

Over activation of neutrophils has been linked to many inflammatory diseases; one of critical pathologic mechanisms is that generation and exocellular release of superoxide anion from neutrophils results in peripheral tissues damage. Besides, in this study, 2-(3,5-dimethoxyphenoxy)-5,7-dimethoxy-chromen-4-one (4), a 2-phexnoychromone from our compound bank, was demonstrated to have the moderate inhibitory effect on superoxide anion generating. Therefore, serial chromones substituted with phenols or 3-flourothiophenol were designed, synthesized, and examined for suppression of superoxide anion generation. In accordance with the results, the methoxy group at 7 position (R³) of the chromone, as well as a hydrogen bond donor at a meta site of the phenyl ring greatly impacted on the activity. 2-(3-fluorophenyl)sulfanyl-7-methoxy-chromen-4-one (16), a successful example of bioisosteres from a phenol to a thiophenol, exhibited prominent anti-inflammatory effects with the IC₅₀ value against superoxide anion generation of 5.0 ± 1.4 μM.

Full text of DOI:10.1016/j.bmcl.2021.127822

Bioorg. Med. Chem. Lett. 36 (2021) 127822
Contents lists available at ScienceDirect
Bioorganic & Medicinal Chemistry Letters
journal homepage: www.elsevier.com/locate/bmcl
The study on structure-activity relationship between chromone derivatives
and inhibition of superoxide anion generating from human neutrophils
,
c
,
,*
Yi-Han Changa, Fang Shu-Yenb, Hsuan-Yu Laia, Tsong-Long Hwangb d, Hsin-Yi Hunga
a School of Pharmacy, National Cheng Kung University Hospital, College of Medicine, National Cheng Kung University, Tainan 701, Taiwan, ROC
b Graduate Institute of Biomedical Sciences and Graduate Institute of Natural Products, College of Medicine, Chang Gung University, Taoyuan 333, Taiwan, ROC
c Research Center for Chinese Herbal Medicine, Graduate Institute of Health Industry Technology, College of Human Ecology, Chang Gung University of Science and
Technology, Taoyuan 333, Taiwan, ROC
d Department of Anesthesiology, Chang Gung Memorial Hospital, Taoyuan 333, Taiwan, ROC
A R T I C L E I N F O
A B S T R A C T
Keywords:
Over activation of neutrophils has been linked to many inflammatory diseases; one of critical pathologic
mechanisms is that generation and exocellular release of superoxide anion from neutrophils results in peripheral
tissues damage. Besides, in this study, 2-(3,5-dimethoxyphenoxy)-5,7-dimethoxy-chromen-4-one (4), a 2-phex-
noychromone from our compound bank, was demonstrated to have the moderate inhibitory effect on superox-
ide anion generating. Therefore, serial chromones substituted with phenols or 3-flourothiophenol were designed,
synthesized, and examined for suppression of superoxide anion generation. In accordance with the results, the
methoxy group at 7 position (R3) of the chromone, as well as a hydrogen bond donor at a meta site of the phenyl
ring greatly impacted on the activity. 2-(3-fluorophenyl)sulfanyl-7-methoxy-chromen-4-one (16), a successful
example of bioisosteres from a phenol to a thiophenol, exhibited prominent anti-inflammatory effects with the
2-Phenxoychromone
Neutrophils
Inflammatory diseases
Superoxide anion
Structure-activity relationship
IC50 value against superoxide anion generation of 5.0 ± 1.4 μM.
Neutrophils, members of innate immune system, are the first white
blood cells recruited to infectious sites to protect the host from further
invasion of various pathogens.1 Neutrophils eliminate pathogens
through primary four mechanisms: (1.) phagocytosis, (2.) respiratory
burst with generation of highly oxidant chemical molecules like reactive
elastic tissues, collagen and basement membranes are also detrimental
to these inflammatory disorders.5 As a consequence, suppressing
vigorous neutrophils may be a strategy to ameliorate inflammatory
diseases.
Several compounds with different skeletons in our compound bank
were examined for inhibition of dihydronicotinamide-adenine dinucle-
–
•
oxygen species (ROS), Nitric oxide ( NO), and peroxynitrite (ONOO ),
(3.) degranulation of cytotoxic proteases and (4.) neutrophil extracel-
lular traps (NET).2,3 However, overactivation of neutrophils would lead
to inflammatory diseases. Generally, after phagocytosis, phagosomes
merge with granules including ROS or cytotoxins to eradicate patho-
gens.1 Nonetheless, severe infection or over production of pro-
inflammatory mediators trigger formation of enormous complexes
compromised of pathogens, complements and antibodies. Since these
complexes are too large that neutrophils are unable to swallow in a
mouthful, incomplete phagosomes form and fuse with granules during
phagocytosis. Extracellular leakage of ROS and cytotoxins from the
imperfect phagosomes would end up damaging peripheral tissues.2,4
This pathogenic mechanism has been linked to several inflammatory
diseases, such as glomerulonephritis,5 thrombohemorrhagic vasculop-
athy.6 Furthermore, other pathologic mechanisms like digestion of
•
otide phosphate (NADPH) oxidase-mediated superoxide anion (Oꢀ2 )
generation in human neutrophils, which is the primary stage for the
synthesis of different ROS.7 Compound 4, a 2-phenoxychromoe, was
shown to moderately inhibit the generation of superoxide anion at a
concentration of 10 μM, and the IC50 value was 27.5 ± 3.5 μM (Table 1).
2-Phenoxychromones are uncommon in natural products, one of which,
capillarisin (Fig. 1), isolated from Artemisia capillaris Thunb., was also
reported to have anti-inflammatory effects.8 Therefore, this study aimed
at design and synthesis of 2-phenoxychromoe analogues. Furthermore,
structure-activity relationship (SAR) between these compounds and
suppression on superoxide anion generation from neutrophils was also
investigated.
As there are four methoxy groups in compound 4, this functional
group may play an important role in activity. Thus, compounds with
* Corresponding author.
E-mail address: z10308005@email.ncku.edu.tw (H.-Y. Hung).
https://doi.org/10.1016/j.bmcl.2021.127822
Received 26 November 2020; Received in revised form 11 January 2021; Accepted 19 January 2021
Available online 27 January 2021
0960-894X/© 2021 Elsevier Ltd. All rights reserved.

Y.-H. Chang et al.
Bioorganic & Medicinal Chemistry Letters 36 (2021) 127822
under basic condition, which were then attacked by 1,2,4-triazole at
C-2 position concomitantly with dehydroiodination at C-3 position to
give 2-(1H-1,2,4-triazol-1-yl)-chromen-4-ones (3a–3e).9 Functioning as
a leaving group, the triazole group at 3a–3e was attacked by various
phenols under basic condition to afford final compound 4–13. The
inhibitory activity of superoxide anion generation (Table 1) and elastase
release (Supplementary Information) for all final compounds was eval-
Table 1
Effects of compound 4–18 on superoxide anion generation in fMLF/CB-induced
human neutrophils.
Compound Inhibition (%)a IC50
(
μ
M) Compound Inhibition (%)a IC50
(μM)
4
27.5 ± 3.5 **
7.8 ± 7.4
>10
>10
>10
>10
>10
>10
12
13
14
15
16
17
18
6.9 ± 4.1
>10
5
71.3 ± 2.5 *** 5.2 ± 0.5
61.0 ± 2.4 *** 7.0 ± 0.6
6
22.4 ± 4.3 **
2.5 ± 6.3
7
10.6 ± 4.6
>10
uated at a concentration of 10
μM. Nonetheless, none of the tested
8
ꢀ 0.4 ± 2.3
12.8 ± 7.7
70.9 ± 3.1 *** 5.0 ± 1.4
17.3 ± 8.0 >10
compounds exhibited remarkable inhibition on elastase release.
Since two meta-methoxy groups on compound 4 enhanced the elec-
tron density, compound 5, with two methoxy groups at the meta and
para positions of the same ring, was also predicted to have activity.
However, inferior activity of 5 to 4 indicated that electron density was
not the principal factor. In contrast, 6 maintained similar activity with 4,
implying that meta substitutions rather than electron density dominated
this inhibitory effect. This was also supported with the loss of activity of
7 and 11, whose only methoxy group was at the ortho position. Next, we
concentrated on the functional groups at chromone ring. 8 and 9 almost
lost activity, compared to 4, implicating that the methoxy group at R1 of
the chromone ring deteriorated activity; instead, 10 was superior in
activity to 4 and 8, suggesting that the methoxy group at R3 of the same
ring was essential. Moreover, a hydrogen bond acceptor instead of a
hydrogen bond donor at the meta position of the phenyl ring was much
more crucial, as 12 almost totally lost the activity in comparison to 10.
Interestingly, inserting a hydroxyl group to 10 at para position of the
phenyl ring drastically refrained from generating superoxide anion (13);
nonetheless, the hydroquinone moiety in 13 might cause cytotoxicity.
As a result, the phenyl ring was bioisosterically replaced with a thio-
phenol substituted with a fluorine at the meta position, which was also a
potent hydrogen bond acceptor.
9
10
11
46.8 ± 3.4 *** >10
47.5 ± 1.4 *** >10
3.8 ± 6.9
>10
Ly294002b 93.4 ± 2.7 *** 1.3 ± 0.4
a
Percentage of inhibition at 10 μM. Results are presented as mean ± S.E.M. (n
= 3 ~ 5). ** P < 0.01, *** P < 0.001 compared with the control (DMSO).
b
Positive control. A phosphoinositide 3-kinase (PI3K) inhibitor that interferes
degranulation pathway.10
Fig. 1. The structure of capillarisin.
various numbers of methoxy groups at different positions were prepared
(5–13).
The general synthesis procedure is demonstrated in Fig. 2. In brief, 2-
hydorxyacetophenones (1a–1e) reacted with N,N-dimethylformamide
dimethyl acetal (DMF-DMA) under reflux condition to afford enamino-
ketone intermediates; sequentially HCl-mediated ring closure of enam-
inoketones gave chromones (2a–2e). Catalyzed by 1,2,4-triazole,
chromones reacted with molecular iodine to form 3-iodo-chromones
The general synthesis procedure of 2-(3-fluorophenyl)sulfanylchro-
men-4-ones substituted with methoxy groups (14–18) was the same as
Fig. 2. Synthesis of target compounds 4–18:(a) DMF-DMA, 100 ◦C, 2 h; (b) HCl, DCM, 40 ◦C, 20 mins; (c) I2, 1,2,4-triazole, K2CO3, DMF, 80 ◦C, 5 h; (d) Cs2CO3, 1,4-
dioxane, 80 ◦C, 30 mins.
2

Y.-H. Chang et al.
Bioorganic & Medicinal Chemistry Letters 36 (2021) 127822
that of 4–13 (Fig. 2).
Acknowledgments
Successfully, 16 retained similar activity to 13. However, 14 and 18,
with methoxy groups at R1 and/or R2 position, worse suppressed su-
peroxide anion generation than 16 did. In addition, 15 and 17, with a
methoxy group at R3 and R2 position receptively, had little inhibitory
activity, in accordance with aforementioned hypothesis that the
methoxy at R3 was important.
The investigation was supported by research grants from Ministry of
Science and Technology, Taiwan awarded to H.-Y.H.
Appendix A. Supplementary data
To sum up, in this study, a series of chromones substituted with
phenols or 3-flourothiophenol were designed, synthesized, and investi-
gated the anti-inflammatory effect against superoxide anion generation.
According to SAR, a methoxy group at R3 was essential while that at R1
or R2 was deleterious to activity; in addition, phenyl or thiophenyl ring
with a hydrogen bond acceptor at the meta site potentiated the inhibi-
tory effect. Compound 16, which prominently suppressed superoxide
Supplementary data to this article can be found online at https://doi.
org/10.1016/j.bmcl.2021.127822.
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