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Synthesis and biological evaluation of 3-carbamate smilagenin derivatives as potential neuroprotective agents
Bioorganic & Medicinal Chemistry Letters 29 (2019) 126622
Contents lists available at ScienceDirect
Bioorganic & Medicinal Chemistry Letters journal homepage: www.elsevier.com/locate/bmcl
Synthesis and biological evaluation of 3-carbamate smilagenin derivatives as potential neuroprotective agents Cong Zhang, Yan Wu, Jie Li, Gui-Xiang Yang, Lin Su, Yan Huang, Rui Wang , Lei Ma ⁎
T
⁎
Shanghai Key Laboratory of New Drug Design, School of Pharmacy, East China University of Science and Technology, 130 Meilong Road, Shanghai 200237, China
ARTICLE INFO
ABSTRACT
Keywords: Smilagenin derivatives Alzheimer’s disease Oxidative stress Neuroprotective activity
Studies indicated that smilagenin, isolated from Anemarrhena asphodeloides Bunge, could improve cognitive impairment and exhibit neuroprotective activity. On the basis of the structure of smilagenin, a series of derivatives were synthesized and evaluated for their neuroprotective effects of H2O2-induced, oxygen glucose deprivation-induced neurotoxicity in SH-SY5Y cells and LPS-induced NO production in RAW264.7 cells. Structure activity relationship of derivatives revealed that benzyl-substituted piperazine formate derivatives showed the potent neuroprotective activity such as A12. These findings may provide new insights for the development of neuroprotective agents against Alzheimer’s disease.
Alzheimer’s disease (AD), one of the most familiar neurodegenerative disorders, is a clinical syndrome characterized by memory loss and cognitive impairment.1 There is about 47 million people suffering from AD worldwide, and the number is predicted to grow to 114 million by 2050.2 In recent decades, a lot of efforts in AD research have been made, but the pathological mechanism of AD is still far from being elucidated.3 However, evidences indicated that oxidative stress (OS) played a pivotal role in AD.4–7 OS can cause oxidative modification of proteins, lipids and DNA/RNA, leading to mitochondrial dysfunction and cell damage. Therefore, antioxidant stress may contribute to the treatment of AD. In recent years, the search for treatments of neurodegenerative diseases has attracted widespread attention.8,9 It was found that mickle natural products, isolated from a series of medicinal plants, exhibited potential neuroprotective activity, such as steroids,10 alkaloids,11 polysaccharide12 and polyphenols.13,14 Smilagenin (SMI), a kind of lipid-soluble small molecule steroidal sapogenin isolated from Anemarrhena asphodeloides Bunge, showed positive effects on cognition.15 In addition, SMI had antioxidant activity16 and improved memory impairment.17 Due to the low content of SMI in natural products, in this study, SMI was first synthesized with diosgenin as a raw material,18 via a few steps. The synthesis of SMI was outlined in Scheme 1. The compounds A1–A8 (Scheme 2) were synthesized from SMI by
⁎
two steps. In the first step, SMI was treated with 4-nitrophenyl chloroformate in the presence of pyridine for 4 h to give the intermediate product S1. Then S1 was reacted with the related amine in dichloromethane (DCM) and triethylamine (TEA) to form the carbamate derivatives (A1–A8). The synthesis of compounds A9–A14 (Scheme 3) was prepared from SMI by three steps. The preparation of compound S2 was the same in first two steps staring from compounds A1–A8. Then, compound S2 was reacted with NaBH3CN, aldehydes and acetic acid in MeOH to obtain the target products (A9–A14). In order to evaluate the neuroprotective activity of SMI derivatives, cell viabilities of A1–A14 against oxidative stress induced by H2O2 in SH-SY5Y cells were carried out by MTT assay. As shown in Table 1, most target compounds could improve cell viabilities against H2O2-induced damage.19 Compared with A1 and A2, the compounds with aromatic ring (A3–A5) had relatively strong antioxidant activity. But the compound A6 with benzene ring didn’t exhibit good activity. It might hint that the antioxidant activity of compound was improved when the p-position of benzene ring was substituted. Besides, A7 and A8 didn’t show good activity. Interestingly, when the p-position of piperazine was substituted by benzyl groups, the compounds of A9, A10, A12 and A13 displayed satisfactory activities from 34.8 ± 1.8% to 40.5 ± 2.3%. We could conclude that derivatives with N-benzyl piperazine could improve the antioxidant activity.
Corresponding authors. E-mail addresses: [email protected] (R. Wang), [email protected] (L. Ma).
https://doi.org/10.1016/j.bmcl.2019.08.026 Received 30 June 2019; Received in revised form 27 July 2019; Accepted 12 August 2019 Available online 13 August 2019 0960-894X/ © 2019 Elsevier Ltd. All rights reserved.
Bioorganic & Medicinal Chemistry Letters 29 (2019) 126622
C. Zhang, et al.
Scheme 1. Preparation of smilagenin. Reagents and conditions: (a) cyclohexanone, aluminium isopropoxide, tolunene, 100 °C, 6 h; (b) H2, Pd/C, MeOH, 60 °C, 4 h; (c) NaBH4, THF, r.t, 2 h.
Scheme 2. Synthesis of SMI derivatives A1–A8. Reagents and conditions: (a) 4-nitrophenyl chloroformate, CH2Cl2, pyridine, 0 °C to r.t, 4 h; (b) R1R2NH, CH2Cl2, Et3N, r.t, 3 h.
2
Bioorganic & Medicinal Chemistry Letters 29 (2019) 126622
C. Zhang, et al.
Scheme 3. Synthesis of SMI derivatives A9–A14. Reagents and conditions: (a) 4-nitrophenyl chloroformate, CH2Cl2, pyridine, 0 °C to r.t, 4 h; (b) piperazine, CH2Cl2, Et3N, r.t, 3 h; (c) NaBH3CN, aldehydes, acetic acid, MeOH, r.t, 2 h.
Table 1 Neuroprotective activities against H2O2-induced cytotoxicity of compound A1–A14. Compound
% Cell viability (10 μM)
Compound
NAC
37.1 ± 2.5
A7
NA
SMI
3.5 ± 1.6
A8
NA
A1
9.2 ± 1.1
A9
36.9 ± 3.5
A2
6.3 ± 2.9
A10
34.8 ± 1.8
A3
45.5 ± 2.1
A11
4.8 ± 0.2
A4
29.2 ± 3.1
A12
40.5 ± 2.3
a
R
R
% Cell viability (10 μM)
(continued on next page)
3
Bioorganic & Medicinal Chemistry Letters 29 (2019) 126622
C. Zhang, et al.
Table 1 (continued) Compound
% Cell viability (10 μM)
Compound
A5
17.9 ± 1.5
A13
38.4 ± 1.2
A6
5.8 ± 0.5
A14
10.3 ± 1.3
a
R
R
% Cell viability (10 μM)
N-Acetyl-L-Cysteine (1 mM).
In summary, there were seven compounds exhibited good cell viabilities (A3, A4, A5, A9, A10, A12 and A13). Further studies would be focused on these compounds. Oxidative stress was a state caused by an imbalance between the production and removal of reactive oxygen species (ROS)/reactive nitrogen species (RNS) in the body.20 NO, a kind of RNS, is one of the important causes of oxidative stress. Overproduction of NO could lead to oxidative stress damage in RAW264.7 cells, and inhibition of NO production was beneficial to neuroprotection.21 For further explore the neuroprotective activity of SMI derivatives, the NO production inhibitory activity of some derivatives were studied. The results of NO inhibitory activity were illustrated in Table 2. As shown in Table 2, the results revealed that A3, A9, A10 and A12 performed potential NO inhibitory activities. Compared with the activities of A3, A4, and A5, the substituents at p-position of benzene ring had an effect on NO production inhibitory activity. In addition, the
activity of compounds A9, A10, and A12 was significantly improved compared with A4 and A5. The result indicated that the introduction of piperazine ring had a huge impact on activity. When the p-position of benzene ring was substituted by cyanide like compound A12, the NO inhibitory activity would increase markedly. In summary, there were four compounds showed good NO production inhibitory activity (A3, A9, A10, and A12). Further studies would be focused on these compounds. In addition, the oxygen glucose deprivation (OGD) model is also frequently used to investigate the neuroprotective activity of drugs against oxidative stress damage.22 Herein, we selected some semisynthesis derivatives of SMI with relatively better neuroprotective activity against H2O2-induced cytotoxicity and NO inhibitory activity to evaluate their neuroprotective activity in the OGD model in SH-SY5Y cells. The results were shown in Fig. 1. In Fig. 1, the activity of A12 was significantly better than that of A3, A9 and A10 in OGD model. The result revealed again that benzylsubstituted piperazine formate derivatives like A12 exhibited the most potent neuroprotective activity. In conclusion, based on the structure of SMI, a series of carbamate smilagenin derivatives at 3-position were synthesized and their neuroprotective effects of H2O2-induced, oxygen glucose deprivation-
Table 2 NO inhibitory activities of the compounds A3–A13. Compound
R
% NO inhibition (10 μM)
A3
17.6 ± 1.5
A4
6.8 ± 0.8
A5
9.2 ± 0.8
A9
18.3 ± 1.4
A10
15.2 ± 1.6
A12
23.5 ± 1.2
A13
7.02 ± 1.5
Fig. 1. Neuroprotective activity of compounds A3, A9, A10 and A12 were evaluated via the OGD model in SH-SY5Y cell lines. ###P < 0.001 versus control group, ***P < 0.001 compared to the model group. 4
Bioorganic & Medicinal Chemistry Letters 29 (2019) 126622
C. Zhang, et al.
induced neurotoxicity and LPS-induced NO production were evaluated by MTT assays. The results indicated that the benzyl-substituted piperazine formate derivatives were effective against cell damage induced by H2O2 and lipopolysaccharide. The study on structure activity relationship of these derivatives revealed that benzyl-substituted piperazine formate derivatives showed the potent neuroprotective activity such as A10 and A12. In addition, compound A12 displayed the most potential neuroprotective activity in the OGD model. These findings may provide new insights for the development of neuroprotective agents against Alzheimer’s disease.
doi.org/10.1016/j.bmcl.2019.08.026. References 1. 2. 3. 4. 5. 6. 7. 8. 9. 10. 11. 12. 13. 14. 15. 16. 17. 18. 19. 20. 21. 22.
Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper. Acknowledgments This work was supported by National Natural Science Foundation of China (81673318) and Shanghai Biomedical Technology Support Program (15401901100). Appendix A. Supplementary data Supplementary data to this article can be found online at https://
5
Fang Y, Wang R, Wang Q, et al. Bioorg Med Chem Lett. 2018;28:668. Yang G-X, Ge S-L, Wu Y, et al. Eur J Med Chem. 2018;156:206. Pan H, Van Khang P, Dong D, et al. Bioorg Med Chem Lett. 2017;27:662. Nunomura A, Perry G, Aliev G, et al. J Neuropathol Exp Neurol. 2001;60:759. Castegna A, Lauderback CM, Mohmmad-Abdul H, et al. Brain Res. 2004;1004:193. Granold M, Moosmann B, Staib-Lasarzik I, et al. Redox Biol. 2015;4:200. Su JH, Deng G, Cotman CW. Brain Res. 1997;774:193. Riedel G, Bergman J, Vanderschuren L, et al. Behav Pharmacol. 2017;28:91. Barnett R, Richard. Lancet. 2019;393:1589. Wang Z-D, Yao G-D, Wang W, et al. Steroids. 2017;125:93. Zhou L-Y, Zhu Y, Jiang Y-R, et al. Bioorg Med Chem Lett. 2017;27:4180. Liu Q, Wang S-C, Ding K. Chin J Nat Med. 2017;15:641. Ringman JM, Frautschy SA, Cole GM, et al. Curr Alzheimer Res. 2005;2:131. Cole GM, Lim GP, Yang F, et al. Neurobiol Aging. 2005;26:133. Kim HK, Mendonca KM, Howson PA, et al. Eur J Pharmacol. 2015;764:379. Murali A, Ashok P, Madhavan V. PhOL. 2010:121. Zhang Y, Xia Z, Hu Y, et al. FEBS Lett. 2008;582:956. Gunning PJ, Tiffin PD. US 2006041119 (A1). EP; 2006. Bao D, Wang J, Pang X, et al. Molecules. 2017;22:1122. Swomley AM, Förster S, Keeney JT, et al. BBA-Mol Basis Dis. 2014;1842:1248. Hariharan A, Jing Y, Collie ND, et al. Nitric Oxide-Biol Chem. 2019;87:60. Li Y, Zhang W, Chen C, et al. J Appl Biomed. 2018;16:126.
Contents lists available at ScienceDirect
Bioorganic & Medicinal Chemistry Letters journal homepage: www.elsevier.com/locate/bmcl
Synthesis and biological evaluation of 3-carbamate smilagenin derivatives as potential neuroprotective agents Cong Zhang, Yan Wu, Jie Li, Gui-Xiang Yang, Lin Su, Yan Huang, Rui Wang , Lei Ma ⁎
T
⁎
Shanghai Key Laboratory of New Drug Design, School of Pharmacy, East China University of Science and Technology, 130 Meilong Road, Shanghai 200237, China
ARTICLE INFO
ABSTRACT
Keywords: Smilagenin derivatives Alzheimer’s disease Oxidative stress Neuroprotective activity
Studies indicated that smilagenin, isolated from Anemarrhena asphodeloides Bunge, could improve cognitive impairment and exhibit neuroprotective activity. On the basis of the structure of smilagenin, a series of derivatives were synthesized and evaluated for their neuroprotective effects of H2O2-induced, oxygen glucose deprivation-induced neurotoxicity in SH-SY5Y cells and LPS-induced NO production in RAW264.7 cells. Structure activity relationship of derivatives revealed that benzyl-substituted piperazine formate derivatives showed the potent neuroprotective activity such as A12. These findings may provide new insights for the development of neuroprotective agents against Alzheimer’s disease.
Alzheimer’s disease (AD), one of the most familiar neurodegenerative disorders, is a clinical syndrome characterized by memory loss and cognitive impairment.1 There is about 47 million people suffering from AD worldwide, and the number is predicted to grow to 114 million by 2050.2 In recent decades, a lot of efforts in AD research have been made, but the pathological mechanism of AD is still far from being elucidated.3 However, evidences indicated that oxidative stress (OS) played a pivotal role in AD.4–7 OS can cause oxidative modification of proteins, lipids and DNA/RNA, leading to mitochondrial dysfunction and cell damage. Therefore, antioxidant stress may contribute to the treatment of AD. In recent years, the search for treatments of neurodegenerative diseases has attracted widespread attention.8,9 It was found that mickle natural products, isolated from a series of medicinal plants, exhibited potential neuroprotective activity, such as steroids,10 alkaloids,11 polysaccharide12 and polyphenols.13,14 Smilagenin (SMI), a kind of lipid-soluble small molecule steroidal sapogenin isolated from Anemarrhena asphodeloides Bunge, showed positive effects on cognition.15 In addition, SMI had antioxidant activity16 and improved memory impairment.17 Due to the low content of SMI in natural products, in this study, SMI was first synthesized with diosgenin as a raw material,18 via a few steps. The synthesis of SMI was outlined in Scheme 1. The compounds A1–A8 (Scheme 2) were synthesized from SMI by
⁎
two steps. In the first step, SMI was treated with 4-nitrophenyl chloroformate in the presence of pyridine for 4 h to give the intermediate product S1. Then S1 was reacted with the related amine in dichloromethane (DCM) and triethylamine (TEA) to form the carbamate derivatives (A1–A8). The synthesis of compounds A9–A14 (Scheme 3) was prepared from SMI by three steps. The preparation of compound S2 was the same in first two steps staring from compounds A1–A8. Then, compound S2 was reacted with NaBH3CN, aldehydes and acetic acid in MeOH to obtain the target products (A9–A14). In order to evaluate the neuroprotective activity of SMI derivatives, cell viabilities of A1–A14 against oxidative stress induced by H2O2 in SH-SY5Y cells were carried out by MTT assay. As shown in Table 1, most target compounds could improve cell viabilities against H2O2-induced damage.19 Compared with A1 and A2, the compounds with aromatic ring (A3–A5) had relatively strong antioxidant activity. But the compound A6 with benzene ring didn’t exhibit good activity. It might hint that the antioxidant activity of compound was improved when the p-position of benzene ring was substituted. Besides, A7 and A8 didn’t show good activity. Interestingly, when the p-position of piperazine was substituted by benzyl groups, the compounds of A9, A10, A12 and A13 displayed satisfactory activities from 34.8 ± 1.8% to 40.5 ± 2.3%. We could conclude that derivatives with N-benzyl piperazine could improve the antioxidant activity.
Corresponding authors. E-mail addresses: [email protected] (R. Wang), [email protected] (L. Ma).
https://doi.org/10.1016/j.bmcl.2019.08.026 Received 30 June 2019; Received in revised form 27 July 2019; Accepted 12 August 2019 Available online 13 August 2019 0960-894X/ © 2019 Elsevier Ltd. All rights reserved.
Bioorganic & Medicinal Chemistry Letters 29 (2019) 126622
C. Zhang, et al.
Scheme 1. Preparation of smilagenin. Reagents and conditions: (a) cyclohexanone, aluminium isopropoxide, tolunene, 100 °C, 6 h; (b) H2, Pd/C, MeOH, 60 °C, 4 h; (c) NaBH4, THF, r.t, 2 h.
Scheme 2. Synthesis of SMI derivatives A1–A8. Reagents and conditions: (a) 4-nitrophenyl chloroformate, CH2Cl2, pyridine, 0 °C to r.t, 4 h; (b) R1R2NH, CH2Cl2, Et3N, r.t, 3 h.
2
Bioorganic & Medicinal Chemistry Letters 29 (2019) 126622
C. Zhang, et al.
Scheme 3. Synthesis of SMI derivatives A9–A14. Reagents and conditions: (a) 4-nitrophenyl chloroformate, CH2Cl2, pyridine, 0 °C to r.t, 4 h; (b) piperazine, CH2Cl2, Et3N, r.t, 3 h; (c) NaBH3CN, aldehydes, acetic acid, MeOH, r.t, 2 h.
Table 1 Neuroprotective activities against H2O2-induced cytotoxicity of compound A1–A14. Compound
% Cell viability (10 μM)
Compound
NAC
37.1 ± 2.5
A7
NA
SMI
3.5 ± 1.6
A8
NA
A1
9.2 ± 1.1
A9
36.9 ± 3.5
A2
6.3 ± 2.9
A10
34.8 ± 1.8
A3
45.5 ± 2.1
A11
4.8 ± 0.2
A4
29.2 ± 3.1
A12
40.5 ± 2.3
a
R
R
% Cell viability (10 μM)
(continued on next page)
3
Bioorganic & Medicinal Chemistry Letters 29 (2019) 126622
C. Zhang, et al.
Table 1 (continued) Compound
% Cell viability (10 μM)
Compound
A5
17.9 ± 1.5
A13
38.4 ± 1.2
A6
5.8 ± 0.5
A14
10.3 ± 1.3
a
R
R
% Cell viability (10 μM)
N-Acetyl-L-Cysteine (1 mM).
In summary, there were seven compounds exhibited good cell viabilities (A3, A4, A5, A9, A10, A12 and A13). Further studies would be focused on these compounds. Oxidative stress was a state caused by an imbalance between the production and removal of reactive oxygen species (ROS)/reactive nitrogen species (RNS) in the body.20 NO, a kind of RNS, is one of the important causes of oxidative stress. Overproduction of NO could lead to oxidative stress damage in RAW264.7 cells, and inhibition of NO production was beneficial to neuroprotection.21 For further explore the neuroprotective activity of SMI derivatives, the NO production inhibitory activity of some derivatives were studied. The results of NO inhibitory activity were illustrated in Table 2. As shown in Table 2, the results revealed that A3, A9, A10 and A12 performed potential NO inhibitory activities. Compared with the activities of A3, A4, and A5, the substituents at p-position of benzene ring had an effect on NO production inhibitory activity. In addition, the
activity of compounds A9, A10, and A12 was significantly improved compared with A4 and A5. The result indicated that the introduction of piperazine ring had a huge impact on activity. When the p-position of benzene ring was substituted by cyanide like compound A12, the NO inhibitory activity would increase markedly. In summary, there were four compounds showed good NO production inhibitory activity (A3, A9, A10, and A12). Further studies would be focused on these compounds. In addition, the oxygen glucose deprivation (OGD) model is also frequently used to investigate the neuroprotective activity of drugs against oxidative stress damage.22 Herein, we selected some semisynthesis derivatives of SMI with relatively better neuroprotective activity against H2O2-induced cytotoxicity and NO inhibitory activity to evaluate their neuroprotective activity in the OGD model in SH-SY5Y cells. The results were shown in Fig. 1. In Fig. 1, the activity of A12 was significantly better than that of A3, A9 and A10 in OGD model. The result revealed again that benzylsubstituted piperazine formate derivatives like A12 exhibited the most potent neuroprotective activity. In conclusion, based on the structure of SMI, a series of carbamate smilagenin derivatives at 3-position were synthesized and their neuroprotective effects of H2O2-induced, oxygen glucose deprivation-
Table 2 NO inhibitory activities of the compounds A3–A13. Compound
R
% NO inhibition (10 μM)
A3
17.6 ± 1.5
A4
6.8 ± 0.8
A5
9.2 ± 0.8
A9
18.3 ± 1.4
A10
15.2 ± 1.6
A12
23.5 ± 1.2
A13
7.02 ± 1.5
Fig. 1. Neuroprotective activity of compounds A3, A9, A10 and A12 were evaluated via the OGD model in SH-SY5Y cell lines. ###P < 0.001 versus control group, ***P < 0.001 compared to the model group. 4
Bioorganic & Medicinal Chemistry Letters 29 (2019) 126622
C. Zhang, et al.
induced neurotoxicity and LPS-induced NO production were evaluated by MTT assays. The results indicated that the benzyl-substituted piperazine formate derivatives were effective against cell damage induced by H2O2 and lipopolysaccharide. The study on structure activity relationship of these derivatives revealed that benzyl-substituted piperazine formate derivatives showed the potent neuroprotective activity such as A10 and A12. In addition, compound A12 displayed the most potential neuroprotective activity in the OGD model. These findings may provide new insights for the development of neuroprotective agents against Alzheimer’s disease.
doi.org/10.1016/j.bmcl.2019.08.026. References 1. 2. 3. 4. 5. 6. 7. 8. 9. 10. 11. 12. 13. 14. 15. 16. 17. 18. 19. 20. 21. 22.
Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper. Acknowledgments This work was supported by National Natural Science Foundation of China (81673318) and Shanghai Biomedical Technology Support Program (15401901100). Appendix A. Supplementary data Supplementary data to this article can be found online at https://
5
Fang Y, Wang R, Wang Q, et al. Bioorg Med Chem Lett. 2018;28:668. Yang G-X, Ge S-L, Wu Y, et al. Eur J Med Chem. 2018;156:206. Pan H, Van Khang P, Dong D, et al. Bioorg Med Chem Lett. 2017;27:662. Nunomura A, Perry G, Aliev G, et al. J Neuropathol Exp Neurol. 2001;60:759. Castegna A, Lauderback CM, Mohmmad-Abdul H, et al. Brain Res. 2004;1004:193. Granold M, Moosmann B, Staib-Lasarzik I, et al. Redox Biol. 2015;4:200. Su JH, Deng G, Cotman CW. Brain Res. 1997;774:193. Riedel G, Bergman J, Vanderschuren L, et al. Behav Pharmacol. 2017;28:91. Barnett R, Richard. Lancet. 2019;393:1589. Wang Z-D, Yao G-D, Wang W, et al. Steroids. 2017;125:93. Zhou L-Y, Zhu Y, Jiang Y-R, et al. Bioorg Med Chem Lett. 2017;27:4180. Liu Q, Wang S-C, Ding K. Chin J Nat Med. 2017;15:641. Ringman JM, Frautschy SA, Cole GM, et al. Curr Alzheimer Res. 2005;2:131. Cole GM, Lim GP, Yang F, et al. Neurobiol Aging. 2005;26:133. Kim HK, Mendonca KM, Howson PA, et al. Eur J Pharmacol. 2015;764:379. Murali A, Ashok P, Madhavan V. PhOL. 2010:121. Zhang Y, Xia Z, Hu Y, et al. FEBS Lett. 2008;582:956. Gunning PJ, Tiffin PD. US 2006041119 (A1). EP; 2006. Bao D, Wang J, Pang X, et al. Molecules. 2017;22:1122. Swomley AM, Förster S, Keeney JT, et al. BBA-Mol Basis Dis. 2014;1842:1248. Hariharan A, Jing Y, Collie ND, et al. Nitric Oxide-Biol Chem. 2019;87:60. Li Y, Zhang W, Chen C, et al. J Appl Biomed. 2018;16:126.