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©The Author(s) 2022. Published by Baishideng Publishing Group Inc. All rights reserved.
World J Gastrointest Oncol. Feb 15, 2022; 14(2): 450-477
Published online Feb 15, 2022. doi: 10.4251/wjgo.v14.i2.450
Frankincense myrrh attenuates hepatocellular carcinoma by regulating tumor blood vessel development through multiple epidermal growth factor receptor-mediated signaling pathways
Piao Zheng, Zhen Huang, Dong-Chang Tong, Qing Zhou, Sha Tian, Bo-Wei Chen, Di-Min Ning, Yin-Mei Guo, Wen-Hao Zhu, Yan Long, Wei Xiao, Zhe Deng, Yi-Chen Lei, Xue-Fei Tian
Piao Zheng, Zhen Huang, Dong-Chang Tong, Sha Tian, Di-Min Ning, Wen-Hao Zhu, Zhe Deng, Yi-Chen Lei, Xue-Fei Tian, College of Integrated Traditional Chinese and Western Medicine, Hunan University of Chinese Medicine, Changsha 410208, Hunan Province, China
Piao Zheng, Department of Integrated Traditional Chinese and Western Medicine, The Second Xiangya Hospital, Central South University, Changsha 410011, Hunan Province, China
Qing Zhou, Bo-Wei Chen, Yan Long, The First Affiliated Hospital, Hunan University of Chinese Medicine, Changsha 410021, Hunan Province, China
Yin-Mei Guo, Hunan Key Laboratory of Translational Research in Formulas and Zheng of Traditional Chinese Medicine, Hunan University of Chinese Medicine, Changsha 410208, Hunan Province, China
Wei Xiao, Institute of Integrative Medicine, Department of Integrated Traditional Chinese and Western Medicine, Xiangya Hospital, Central South University, Changsha 410008, Hunan Province, China
Author contributions: Tian XF provided the conceptual and technical guidance, designed the study, and revised the manuscript critically for important intellectual content; Zheng P, Tian S and Zhou Q coordinated the study; Zheng P, Huang Z, Tong DC, Ning DM, Guo YM, Zhu WH, Long Y, Deng Z and Lei YC performed the experiments; Zheng P, Chen BW and Xiao W interpreted data; Zheng P wrote the manuscript; all authors approved the final version of the article.
Supported by the National Natural Science Foundation of China, No. U20A20408 (Major Program) and No. 82074450 (General Program); Natural Science Foundation of Hunan Province, No. 2020JJ4066; Hunan Province Research and innovation projects for Postgraduates, No. CX20190541; Hunan Province "domestic first-class cultivation discipline" Integrated Traditional Chinese and Western medicine open fund project, No. 2018ZXYJH03; Hunan University Undergraduate Research Learning and Innovative Experiment Project, No. 201609030114.
Institutional animal care and use committee statement: This research was reviewed and approved by the Committee on the Ethics of Animal Experiments of Hunan University of Chinese Medicine, No. LL2020102801. All animal experiments were conducted in accordance with international standards and under the guidelines for animal care and use formulated by the Animal Experiment Center of Hunan University of Chinese Medicine.
Conflict-of-interest statement: All authors declare no financial or commercial conflict of interest.
Data sharing statement: All data generated or analyzed during this study are included in this published article.
ARRIVE guidelines statement: The authors have read the ARRIVE guidelines, and the manuscript was prepared and revised according to the ARRIVE guidelines.
Open-Access: This article is an open-access article that was selected by an in-house editor and fully peer-reviewed by external reviewers. It is distributed in accordance with the Creative Commons Attribution NonCommercial (CC BY-NC 4.0) license, which permits others to distribute, remix, adapt, build upon this work non-commercially, and license their derivative works on different terms, provided the original work is properly cited and the use is non-commercial. See:
https://creativecommons.org/Licenses/by-nc/4.0/ Corresponding author: Xue-Fei Tian, PhD, Professor, College of Integrated Traditional Chinese and Western Medicine, Hunan University of Chinese Medicine, No. 300 Xueshi Road, Yuelu District, Changsha 410208, Hunan Province, China. 003640@hnucm.edu.cn
Received: July 22, 2021
Peer-review started: July 22, 2021
First decision: November 8, 2021
Revised: November 19, 2021
Accepted: January 14, 2022
Article in press: January 14, 2022
Published online: February 15, 2022
Processing time: 202 Days and 23.7 Hours
BACKGROUND
In traditional Chinese medicine (TCM), frankincense and myrrh are the main components of the antitumor drug Xihuang Pill. These compounds show anticancer activity in other biological systems. However, whether frankincense and/or myrrh can inhibit the occurrence of hepatocellular carcinoma (HCC) is unknown, and the potential molecular mechanism(s) has not yet been determined.
AIM
To predict and determine latent anti-HCC therapeutic targets and molecular mechanisms of frankincense and myrrh in vivo.
METHODS
In the present study, which was based on the Traditional Chinese Medicine Systems Pharmacology Database and Analysis Platform (http://tcmspw.com/tcmsp.php), Universal Protein database (http://www.uniprot.org), GeneCards: The Human Gene Database (http://www.genecards.org/) and Comparative Toxicogenomics Database (http://www.ctdbase.org/), the efficacy of and mechanism by which frankincense and myrrh act as anti-HCC compounds were predicted. The core prediction targets were screened by molecular docking. In vivo, SMMC-7721 human liver cancer cells were transplanted as xenografts into nude mice to establish a subcutaneous tumor model, and two doses of frankincense plus myrrh or one dose of an EGFR inhibitor was administered to these mice continuously for 14 d. The tumors were collected and evaluated: the tumor volume and growth rate were gauged to evaluate tumor growth; hematoxylin-eosin staining was performed to estimate histopathological changes; immunofluorescence (IF) was performed to detect the expression of CD31, α-SMA and collagen IV; transmission electron microscopy (TEM) was conducted to observe the morphological structure of vascular cells; enzyme-linked immunosorbent assay (ELISA) was performed to measure the levels of secreted HIF-1α and TNF-α; reverse transcription-polymerase chain reaction (RT-qPCR) was performed to measure the mRNA expression of HIF-1α, TNF-α, VEGF and MMP-9; and Western blot (WB) was performed to determine the levels of proteins expressed in the EGFR-mediated PI3K/Akt and MAPK signaling pathways.
RESULTS
The results of the network pharmacology analysis showed that there were 35 active components in the frankincense and myrrh extracts targeting 151 key targets. The molecular docking analysis showed that both boswellic acid and stigmasterol showed strong affinity for the targets, with the greatest affinity for EGFR. Frankincense and myrrh treatment may play a role in the treatment of HCC by regulating hypoxia responses and vascular system-related pathological processes, such as cytokine-receptor binding, and pathways, such as those involving serine/threonine protein kinase complexes and MAPK, HIF-1 and ErbB signaling cascades. The animal experiment results were verified. First, we found that, through frankincense and/or myrrh treatment, the volume of subcutaneously transplanted HCC tumors was significantly reduced, and the pathological morphology was attenuated. Then, IF and TEM showed that frankincense and/or myrrh treatment reduced CD31 and collagen IV expression, increased the coverage of perivascular cells, tightened the connection between cells, and improved the shape of blood vessels. In addition, ELISA, RT-qPCR and WB analyses showed that frankincense and/or myrrh treatment inhibited the levels of hypoxia-inducible factors, inflammatory factors and angiogenesis-related factors, namely, HIF-1α, TNF-α, VEGF and MMP-9. Furthermore, mechanistic experiments illustrated that the effect of frankincense plus myrrh treatment was similar to that of an EGFR inhibitor with regard to controlling EGFR activation, thereby inhibiting the phosphorylation activity of its downstream targets: the PI3K/Akt and MAPK (ERK, p38 and JNK) pathways.
CONCLUSION
In summary, frankincense and myrrh treatment targets tumor blood vessels to exert anti-HCC effects via EGFR-activated PI3K/Akt and MAPK signaling pathways, highlighting the potential of this dual TCM compound as an anti-HCC candidate.
Core Tip: Hepatocellular carcinoma (HCC) is a serious threat to human health, and its pathological process is closely related to blood vessels. Currently, powerful targeted drugs for HCC treatment are lacking. The discovery of new drugs is an urgent task. As a complementary and alternative therapy, traditional Chinese medicine has played an increasingly prominent role. This study introduces frankincense and myrrh derived from the classic anticancer Chinese patent medicine Xihuang Pill. Through prediction and verification, the findings confirmed that this frankincense and myrrh treatment targets tumor blood vessels through EGFR-activated PI3K/Akt and MAPK signaling pathways to exert anti-HCC effects. Frankincense and/or myrrh treatment might be a potential anti-HCC drug candidate.
