Emerging Target Discovery Strategies Drive the Decoding of Therapeutic Power of Natural Products and Further Drug Development: A Case Study of Celastrol

Abstract

Celastrol (CEL) is a natural pentacyclic triterpenoid demonstrating significant therapeutic properties against various diseases. However, the ambiguity of target information poses a significant challenge in transitioning CEL from a traditional remedy to a modern pharmaceutical agent. Recently, the emerging target discovery approaches of natural products have broadened extensive avenues for uncovering comprehensive target information of CEL and promoting its drug development. Herein, diverse target discovery strategies are overviewed for the pharmacological and toxicological studies of CEL, including chemical proteomics, protein microarray, degradation-based protein profiling, proteome-wide label-free approaches, network pharmacology, target-based drug screening, multi-omics analysis, and hypothesis-driven target confirmation. Dozens of CEL targets have been identified, which significantly suggests that CEL functions as a multi-target therapeutic agent. Further network interaction analysis and frequency analysis of collected targets reveal that PRDXs, HMGB1, HSP90, STAT3, and PKM2 may serve as key targets for CEL. Additionally, this review highlights the positive role of target discovery in facilitating CEL-based combination therapy and drug delivery, which is essential for further advancing the clinical applications of CEL. Efforts in CEL target identification not only aid in unraveling the scientific underpinnings of its multiple pharmacological effects but also offer crucial insights for further drug development of CEL-based drugs.

Keywords

celastrol / combination therapy / drug delivery / natural product / target identification / target validation

References

[1] a) A. G. Atanasov, S. B. Zotchev, V. M. Dirsch, and C. T. Supuran, “Natural Products in Drug Discovery: Advances and Opportunities,” Nature Reviews Drug Discovery 20 (2021): 200-216. b) L. Zhang, J. Song, L. Kong, et al., “The Strategies and Techniques of Drug Discovery from Natural Products,” Pharmacology and Therapeutics 216 (2020): 107686.

[2] D. J. Newman and G. M. Cragg, “Natural Products as Sources of New Drugs over the Nearly Four Decades from 01/1981 to 09/2019,” Journal of Natural Products 83 (2020): 770-803.

[3] a) X. Chen, Y. Wang, N. Ma, et al., “Target Identification of Natural Medicine with Chemical Proteomics Approach: Probe Synthesis, Target Fishing and Protein Identification,” Signal Transduction and Targeted Therapy 5 (2020): 72. b) J. Chang, Y. Kim, and H. Kwon, “Advances in Identification and Validation of Protein Targets of Natural Products Without Chemical Modification,” Natural Product Reports 33 (2016): 719-730.

[4] Y. Tu, L. Tan, H. Tao, Y. Li, and H. Liu, “CETSA and Thermal Proteome Profiling Strategies for Target Identification and Drug Discovery of Natural Products,” Phytomedicine 116 (2023): 154862.

[5] T. W. Corson and C. M. Crews, “Molecular Understanding and Modern Application of Traditional Medicines: Triumphs and Trials,” Cell 130 (2007): 769-774.

[6] J. Liu, J. Lee, M. A. S. Hernandez, R. Mazitschek, and U. Ozcan, “Treatment of Obesity with Celastrol,” Cell 161 (2015): 999-1011.

[7] J. Bao and S.-M. Dai, “A Chinese Herb Tripterygium Wilfordii Hook F in the Treatment of Rheumatoid Arthritis: Mechanism, Efficacy, and Safety,” Rheumatology International 31 (2011): 1123-1129.

[8] a) C. Wang, S. Dai, X. Zhao, et al., “Celastrol as an Emerging Anticancer Agent: Current Status, Challenges and Therapeutic Strategies,” Biomedicine and Pharmacotherapy 163 (2023): 114882. b) S. W. Ng, Y. Chan, D. K. Chellappan, et al., “Molecular Modulators of Celastrol as the Keystones for Its Diverse Pharmacological Activities,” Biomedicine and Pharmacotherapy 109 (2019): 1785-1792.

[9] a) Y. Lu, Y. Liu, J. Zhou, D. Li, and W. Gao, “Biosynthesis, Total Synthesis, Structural Modifications, Bioactivity, and Mechanism of Action of the Quinone-Methide Triterpenoid Celastrol,” Medicinal Research Reviews 41 (2021): 1022-1060. b) R. Cascão, J. E. Fonseca, and L. F. Moita, “Celastrol: A Spectrum of Treatment Opportunities in Chronic Diseases,” Frontiers in Medicine 4 (2017): 69. c) S. Xu, Y. Feng, W. He, et al., “Celastrol in Metabolic Diseases: Progress and Application Prospects,” Pharmacological Research 167 (2021): 105572.

[10] a) Z. Cui, C. Li, P. Chen, and H. Yang, “An Update of Label-Free Protein Target Identification Methods for Natural Active Products,” Theranostics 12 (2022): 1829-1854. b) X.-X. Guo, S. An, F. Bao, and T.-R. Xu, “Challenges and Perspectives in Target Identification and Mechanism Illustration for Chinese Medicine,” Chinese Journal of Integrative Medicine 29 (2023): 644-654.

[11] M. Schenone, V. Dančík, B. K. Wagner, and P. A. Clemons, “Target Identification and Mechanism of Action in Chemical Biology and Drug Discovery,” Nature Chemical Biology 9 (2013): 232-240.

[12] a) Y. Gao, M. Ma, W. Li, and X. Lei, “Chemoproteomics, a Broad Avenue to Target Deconvolution,” Advancement of Science 11 (2024): 2305608. b) G. Li, X. Peng, Y. Guo, S. Gong, S. Cao, and F. Qiu, “Currently Available Strategies for Target Identification of Bioactive Natural Products,” Frontiers in Chemistry 9 (2021): 761609.

[13] H.-W. Zhang, C. Lv, L.-J. Zhang, et al., “Application of Omics- and Multi-Omics-Based Techniques for Natural Product Target Discovery,” Biomedicine and Pharmacotherapy 141 (2021): 111833.

[14] a) M. Kibble, N. Saarinen, J. Tang, K. Wennerberg, S. Mäkelä, and T. Aittokallio, “Network Pharmacology Applications to Map the Unexplored Target Space and Therapeutic Potential of Natural Products,” Natural Product Reports 32 (2015): 1249-1266. b) S.-Q. Yang, Q. Ye, J.-J. Ding, et al., “Current Advances in Ligand-based Target Prediction,” Wiley Interdisciplinary Reviews: Computational Molecular Science 11 (2021): e1504. c) H. J. Jung and H. J. Kwon, “Target Deconvolution of Bioactive Small Molecules: The Heart of Chemical Biology and Drug Discovery,” Archives of Pharmacal Research 38 (2015): 1627-1641.

[15] a) H. Deng, Q. Lei, Y. Wu, Y. He, and W. Li, “Activity-Based Protein Profiling: Recent Advances in Medicinal Chemistry,” European Journal of Medicinal Chemistry 191 (2020): 112151. b) Y. Zhu, Z. Ouyang, H. Du, et al., “New Opportunities and Challenges of Natural Products Research: When Target Identification Meets Single-Cell Multiomics,” Acta Pharmaceutica Sinica B 12 (2022): 4011-4039.

[16] J. Wang, L. Gao, Y. M. Lee, et al., “Target Identification of Natural and Traditional Medicines with Quantitative Chemical Proteomics Approaches,” Pharmacology and Therapeutics 162 (2016): 10-22.

[17] a) P. Luo, D. Liu, Q. Zhang, et al., “Celastrol Induces Ferroptosis in Activated HSCs to Ameliorate Hepatic Fibrosis via Targeting Peroxiredoxins and HO-1,” Acta Pharmaceutica Sinica B 12 (2022): 2300-2314. b) D.-D. Liu, P. Luo, L. Gu, et al., “Celastrol Exerts a Neuroprotective Effect by Directly Binding to HMGB1 Protein in Cerebral Ischemia-Reperfusion,” Journal of Neuroinflammation 18 (2021): 174. c) Y. Zhou, M. Li, T. Shen, et al., “Celastrol Targets Cullin-Associated and Neddylation-Dissociated 1 to Prevent Fibroblast-Myofibroblast Transformation against Pulmonary Fibrosis,” ACS Chemical Biology 17 (2022): 2734-2743. d) Z. Hong, J. Cao, D. Liu, et al., “Celastrol Targeting Nedd4 Reduces Nrf2-Mediated Oxidative Stress in Astrocytes After Ischemic Stroke,” Journal of Pharmaceutical Analysis 13 (2023): 156-169. e) P. Luo, Q. Zhang, T.-Y. Zhong, et al., “Celastrol Mitigates Inflammation in Sepsis by Inhibiting the PKM2-Dependent Warburg Effect,” Military Medical Research 9 (2022): 22. f) P. Luo, Q. Zhang, S. Shen, et al., “Mechanistic Engineering of Celastrol Liposomes Induces Ferroptosis and Apoptosis by Directly Targeting VDAC2 in Hepatocellular Carcinoma,” Asian Journal of Pharmaceutical Sciences 18 (2023): 100874.

[18] L. Klaić, R. I. Morimoto, and R. B. Silverman, “Celastrol Analogues as Inducers of the Heat Shock Response. Design and Synthesis of Affinity Probes for the Identification of Protein Targets,” ACS Chemical Biology 7 (2012): 928-937.

[19] C. Zhu, J. Yang, Y. Zhu, et al., “Celastrol Alleviates Comorbid Obesity and Depression by Directly Binding Amygdala HnRNPA1 in a Mouse Model,” Clinical and Translational Medicine 11 (2021): e394.

[20] X. Zhang, J. Zhou, Y. Zhu, et al., “Quantitative Chemical Proteomics Reveals Anti-Cancer Targets of Celastrol in HCT116 HHuman Colon Cancer Cells,” Phytomedicine 101 (2022): 154096.

[21] Y. Zhou, Y.-Z. Wei, T.-M. Yang, and T.-T. Zhang, “Celastrol Ameliorates Metabolic Disorders Through Inhibition of PTP1B and TCPTP,” Acta Pharmacologica Sinica 58 (2023): 688.

[22] H. Guo, Y. Yang, Q. Zhang, et al., “Integrated Mass Spectrometry Reveals Celastrol as a Novel Catechol-O-Methyltransferase Inhibitor,” ACS Chemical Biology 17 (2022): 2003-2009.

[23] C. Zhu, X. Yao, D. Liu, et al., “Neuroinflammation Attenuation Effects by Celastrol and PDIA3 in the Amygdala, Hippocampus and Dorsal Raphe Nucleus of Obese Mice,” Journal of Pharmaceutical Analysis 14 (2024): 287-290.

[24] P. Gao, J. Wang, H. Tang, et al., “Chemoproteomics-Based Profiling Reveals Potential Antimalarial Mechanism of Celastrol by Disrupting Spermidine and Protein Synthesis,” Cell Communication and Signaling 22 (2024): 139.

[25] Y. Zhou, W. Li, M. Wang, et al., “Competitive Profiling of Celastrol Targets in Human Cervical Cancer HeLa Cells via Quantitative Chemical Proteomics,” Molecular Biosystems 13 (2017): 83-91.

[26] N. Fan, X. Zhang, W. Zhao, et al., “Covalent Inhibition of Pyruvate Kinase M2 Reprograms Metabolic and Inflammatory Pathways in Hepatic Macrophages Against Non-Alcoholic Fatty Liver Disease,” International Journal of Biological Sciences 18 (2022): 5260-5275.

[27] a) G.-D. Syu, J. Dunn, and H. Zhu, “Developments and Applications of Functional Protein Microarrays,” Molecular and Cellular Proteomics 19 (2020): 916-927. b) J. G. Duarte and J. M. Blackburn, “Advances in the Development of Human Protein Microarrays,” Expert Review of Proteomics 14 (2017): 627-641.

[28] a) H. Xiao-Pei, C. Ji-Kuai, W. Xue, et al., “Systematic Identification of Celastrol-Binding Proteins Reveals That Shoc2 is Inhibited by Celastrol,” Bioscience Reports 38 (2018): BSR20181233. b) X. Chen, Y. Zhao, W. Luo, et al., “Celastrol Induces ROS-Mediated Apoptosis via Directly Targeting Peroxiredoxin-2 in Gastric Cancer Cells,” Theranostics 10 (2020): 10290-10308. c) S. Ye, W. Luo, Z. A. Khan, et al., “Celastrol Attenuates Angiotensin II-Induced Cardiac Remodeling by Targeting STAT3,” Circulation Research 126 (2020): 1007-1023.

[29] Z. Deng, S. Sun, N. Zhou, et al., “PNPO-Mediated Oxidation of DVL3 Promotes Multiple Myeloma Malignancy and Osteoclastogenesis by Activating the Wnt/β-Catenin Pathway,” Advancement of Science 12 (2024): 2407681.

[30] M. He, C. Cao, Z. Ni, et al., “PROTACs: Great Opportunities for Academia and Industry (an Update From 2020 to 2021),” Signal Transduction and Targeted Therapy 7 (2022): 181.

[31] Y. Wu, Y. Yang, W. Wang, et al., “PROTAC Technology as a Novel Tool to Identify the Target of Lathyrane Diterpenoids,” Acta Pharmaceutica Sinica B 12 (2022): 4262-4265.

[32] Z. Ni, Y. Shi, Q. Liu, L. Wang, X. Sun, and Y. Rao, “Degradation-Based Protein Profiling: A Case Study of Celastrol,” Advancement of Science 11 (2024): 2308186.

[33] a) L. Dai, Z. Li, D. Chen, et al., “Target Identification and Validation of Natural Products With Label-Free Methodology: A Critical Review From 2005 to 2020,” Pharmacology and Therapeutics 216 (2020): 107690. b) J. Sun, N. Prabhu, J. Tang, et al., “Recent Advances in Proteome-Wide Label-Free Target Deconvolution for Bioactive Small Molecules,” Medicinal Research Reviews 41 (2021): 2893-2926.

[34] F. Yang, H. Zhou, P. Luo, et al., “Celastrol Induces DNA Damage and Cell Death in BCR-ABL T315I-Mutant CML by Targeting YY1 and HMCES,” Phytomedicine 134 (2024): 155937.

[35] Y. Tian, N. Wan, H. Zhang, et al., “Chemoproteomic Mapping of the Glycolytic Targetome in Cancer Cells,” Nature Chemical Biology 19 (2023): 1480-1491.

[36] Y. Zhu, N. Wan, X. Shan, et al., “Celastrol Targets Adenylyl Cyclase-associated Protein 1 to Reduce Macrophages-Mediated Inflammation and Ameliorates High Fat Diet-Induced Metabolic Syndrome in Mice,” Acta Pharmaceutica Sinica B 11 (2021): 1200-1212.

[37] A. L. Hopkins, “Network Pharmacology,” Nature Biotechnology 25 (2007): 1110-1111.

[38] a) R. Zhang, X. Zhu, H. Bai, and K. Ning, “Network Pharmacology Databases for Traditional Chinese Medicine: Review and Assessment,” Frontiers in Pharmacology 10 (2019): 123. b) L. Shao and B. Zhang, “Traditional Chinese Medicine Network Pharmacology: Theory, Methodology and Application,” Chinese Journal of Natural Medicines 11 (2013): 110. c) Y. Su, Q. Bai, H. Tao, and B. Xu, “Prospects for the Application of Traditional Chinese Medicine Network Pharmacology in Food Science Research,” Journal of the Science of Food and Agriculture 103 (2023): 5183-5200.

[39] a) J. Zhao, H. Liu, M. Xia, et al., “Network Pharmacology and Experimental Validation to Explore That Celastrol Targeting PTEN Is the Potential Mechanism of Tripterygium Wilfordii (Lév.) Hutch Against IgA Nephropathy,” Drug Design, Development and Therapy 17 (2023): 887-900. b) F. Qian, P. Ren, L. Zhao, D. Zheng, W. He, and J. Jin, “Study of Molecular Mechanisms Underlying the Medicinal Plant Tripterygium Wilfordii-Derived Compound Celastrol in Treating Diabetic Nephropathy Based on Network Pharmacology and Molecular Docking,” Biocell 47 (2023): 1853-1867.

[40] S. Xiang, J. Chen, M. Deng, et al., “Celastrol Ameliorates Experimental Autoimmune Uveitis Through STAT3 Targeting and Gut Microenvironment Reprofiling,” International Immunopharmacology 127 (2024): 111339.

[41] S. Dai, H. Wang, M. Wang, Y. Zhang, Z. Zhang, and Z. Lin, “Comparative Transcriptomics and Network Pharmacology Analysis to Identify the Potential Mechanism of Celastrol Against Osteoarthritis,” Clinical Rheumatology 40 (2021): 4259-4268.

[42] a) M. Hu, Q. Luo, G. Alitongbieke, et al., “Celastrol-Induced Nur77 Interaction with TRAF2 Alleviates Inflammation by Promoting Mitochondrial Ubiquitination and Autophagy,” Molecular Cell 66 (2017): 141-153.e6. b) D. Zhang, Z. Chen, C. Hu, et al., “Celastrol Binds to Its Target Protein via Specific Noncovalent Interactions and Reversible Covalent Bonds,” Chemical Communications 54 (2018): 12871-12874.

[43] K. Nouri, T. Azad, M. Ling, et al., “Identification of Celastrol as a Novel YAP-TEAD Inhibitor for Cancer Therapy by High Throughput Screening with Ultrasensitive YAP/TAZ-TEAD Biosensors,” Cancers 11 (2019): 1596.

[44] T. Shirai, A. Nakai, E. Ando, et al., “Celastrol Suppresses Humoral Immune Responses and Autoimmunity by Targeting the COMMD3/8 Complex,” Science Immunology 8 (2023): eadc9324.

[45] O. Cho, J.-W. Lee, Y.-J. Jeong, L. K. Kim, B.-K. Jung, and T.-H. Heo, “Celastrol, Which Targets IL-2/CD25 Binding Inhibition, Induces T Cell-Mediated Antitumor Activity in Melanoma,” European Journal of Pharmacology 962 (2024): 176239.

[46] X. Jiang, S. Chen, Q. Zhang, et al., “Celastrol is a Novel Selective Agonist of Cannabinoid Receptor 2 With Anti-Inflammatory and Anti-Fibrotic Activity in a Mouse Model of Systemic Sclerosis,” Phytomedicine 67 (2020): 153160.

[47] D. V. Titov and J. O. Liu, “Identification and Validation of Protein Targets of Bioactive Small Molecules,” Bioorganic and Medicinal Chemistry 20 (2012): 1902-1909.

[48] M. Cassotta, T. Y. Forbes-Hernandez, D. Cianciosi, et al., “Nutrition and Rheumatoid Arthritis in the ‘Omics’ Era,” Nutrients 13 (2021): 763.

[49] X. Feng, D. Guan, T. Auen, et al., “IL1R1 is Required for Celastrol's Leptin-Sensitization and Antiobesity Effects,” Nature Medicine 25 (2019): 575-582.

[50] M. Dai, W. Peng, L. Lin, et al., “Celastrol as an Intestinal FXR Inhibitor Triggers Tripolide-Induced Intestinal Bleeding: Underlying Mechanism of Gastrointestinal Injury Induced by Tripterygium Wilfordii,” Phytomedicine 121 (2023): 155054.

[51] X. Li Sr., W. Liu, G. Jiang, J. Lian, et al., “Celastrol Ameliorates Neuronal Mitochondrial Dysfunction Induced by Intracerebral Hemorrhage via Targeting cAMP-Activated Exchange Protein-1,” Advancement of Science 11 (2024): 2307556.

[52] Z. Yuan, J. Wang, Q. Qu, et al., “Celastrol Combats Methicillin-Resistant Staphylococcus aureus by Targeting Δ1 -Pyrroline-5-Carboxylate Dehydrogenase,” Advancement of Science 10 (2023): 2302459.

[53] P. Zhang, D. Zhang, W. Zhou, et al., “Network Pharmacology: Towards the Artificial Intelligence-Based Precision Traditional Chinese Medicine,” Briefings in Bioinformatics 25 (2024): bbad518.

[54] H. Xu, H. Zhao, C. Ding, et al., “Celastrol Suppresses Colorectal Cancer via Covalent Targeting Peroxiredoxin 1,” Signal Transduction and Targeted Therapy 8 (2023): 51.

[55] a) H. Wang, X. Ning, F. Zhao, H. Zhao, and D. Li, “Human Organoids-On-Chips for Biomedical Research and Applications,” Theranostics 14 (2024): 788-818. b) X.-Y. Tang, S. Wu, D. Wang, et al., “Human Organoids in Basic Research and Clinical Applications,” Signal Transduction and Targeted Therapy 7 (2022): 168.

[56] a) T. Zhang, Y. Li, Y. Yu, P. Zou, Y. Jiang, and D. Sun, “Characterization of Celastrol to Inhibit Hsp90 and Cdc37 Interaction,” Journal of Biological Chemistry 284 (2009): 35381-35389. b) T. Zhang, A. Hamza, X. Cao, et al., “A Novel Hsp90 Inhibitor to Disrupt Hsp90/Cdc37 Complex Against Pancreatic Cancer Cells,” Molecular Cancer Therapeutics 7 (2008): 162-170.

[57] S.-R. Chen, Z.-Q. Li, J. Xu, et al., “Celastrol Attenuates hepatitis C Virus Translation and Inflammatory Response in Mice by Suppressing Heat Shock Protein 90β,” Acta Pharmacologica Sinica 44 (2023): 1637-1648.

[58] E. Kyriakou, S. Schmidt, G. T. Dodd, et al., “Celastrol Promotes Weight Loss in Diet-Induced Obesity by Inhibiting the Protein Tyrosine Phosphatases PTP1B and TCPTP in the Hypothalamus,” Journal of Medicinal Chemistry 61 (2018): 11144-11157.

[59] R. Fan, H. Chen, B. Lai, Z. Li, W. Xu, and G. Yan, “Celastrol Exerts Anti-Colorectal Cancer Effect via STAT3 Inhibition,” Chinese Pharmacological Bulletin 38 (2022): 1673.

[60] D. Zhou, X. Li, X. Xiao, et al., “Celastrol Targets the ChREBP-TXNIP Axis to Ameliorates Type 2 Diabetes Mellitus,” Phytomedicine 110 (2023): 154634.

[61] C. Liu, N. Li, M. Peng, et al., “Celastrol Directly Binds with VAMP7 and RAB7 to Inhibit Autophagy and Induce Apoptosis in Preadipocytes,” Frontiers in Pharmacology 14 (2023): 1094584.

[62] M. Luo, Y. Wang, Y. Ma, J. Li, J. Wang, and C. Liu, “Celastrol Stabilizes Glycolipid Metabolism in Hepatic Steatosis by Binding and Regulating the Peroxisome Proliferator-Activated Receptor γ Signaling Pathway,” Metabolites 14 (2024): 64.

[63] A. Park and T.-H. Heo, “Celastrol Regulates Psoriatic Inflammation and Autophagy by Targeting IL-17A,” Biomedicine and Pharmacotherapy 172 (2024): 116256.

[64] Y. Zhu, Y. Meng, J. Zhang, et al., “Recent Trends in Anti-tumor Mechanisms and Molecular Targets of Celastrol,” International Journal of Biological Sciences 20 (2024): 5510-5530.

[65] F. Wen, D. Liu, M. Wang, et al., “Celastrol Induces Premature Ovarian Insufficiency by Inducing Apoptosis in Granulosa Cells,” Biomedicine and Pharmacotherapy 169 (2023): 115815.

[66] X. Liu, Q. Zhang, P. Wang, et al., “Dissection of Targeting Molecular Mechanisms of Celastrol-Induced Nephrotoxicity via a Combined Deconvolution Strategy of Chemoproteomics and Metabolomics,” International Journal of Biological Sciences 20 (2024): 4601-4617.

[67] Z. Ding, X. Wang, Y. Zhang, et al., “Altered Iron-Mediated Metabolic Homeostasis Governs the Efficacy and Toxicity of Tripterygium Glycosides Tablets Against Rheumatoid Arthritis,” Engineering 39 (2024): 166-179.

[68] Y. Fang, “Ligand-Receptor Interaction Platforms and Their Applications for Drug Discovery,” Expert Opinion on Drug Discovery 7 (2012): 969-988.

[69] C. J. Wienken, P. Baaske, U. Rothbauer, D. Braun, and S. Duhr, “Protein-Binding Assays in Biological Liquids Using Microscale Thermophoresis,” Nature Communications 1 (2010): 100.

[70] Y. Liu, C. Han, T. Lu, et al., “Investigation of the Interaction between Chrysoeriol and Xanthine Oxidase Using Computational and In Vitro Approaches,” International Journal of Biological Macromolecules 190 (2021): 463-473.

[71] Y.-S. Ren, H.-L. Li, X.-H. Piao, Z.-Y. Yang, S.-M. Wang, and Y.-W. Ge, “Drug Affinity Responsive Target Stability (DARTS) Accelerated Small Molecules Target Discovery: Principles and Application,” Biochemical Pharmacology 194 (2021): 114798.

[72] J. Fan, A. Fu, and L. Zhang, “Progress in Molecular Docking,” Quantitative Biology 7 (2019): 83-89.

[73] V. Salmaso and S. Moro, “Bridging Molecular Docking to Molecular Dynamics in Exploring Ligand-Protein Recognition Process: An Overview,” Frontiers in Pharmacology 9 (2018): 923.

[74] a) M. Wu, C. Deng, T.-H. Lo, K.-Y. Chan, X. Li, and C.-M. Wong, “Peroxiredoxin, Senescence, and Cancer,” Cells 11 (2022): 1772. b) M. Szeliga, “Peroxiredoxins in Neurodegenerative Diseases,” Antioxidants 9 (2020): 1203.

[75] a) R. Kang, R. Chen, Q. Zhang, et al., “HMGB1 in Health and Disease,” Molecular Aspects of Medicine 40 (2014): 1-116. b) J. Xue, J. S. Suarez, M. Minaai, et al., “HMGB1 as a Therapeutic Target in Disease,” Journal of Cellular Physiology 236 (2021): 3406-3419.

[76] A. Hoter, M. E. El-Sabban, and H. Y. Naim, “The HSP90 family: Structure, Regulation, Function, and Implications in Health and Disease,” International Journal of Molecular Sciences 19 (2018): 2560.

[77] S. Arshad, M. Naveed, M. Ullia, et al., “Targeting STAT-3 Signaling Pathway in Cancer for Development of Novel Drugs: Advancements and Challenges,” Genetics and Molecular Biology 43 (2020): e20180160.

[78] Z. Zhang, X. Deng, Y. Liu, Y. Liu, L. Sun, and F. Chen, “PKM2, Function and Expression and Regulation,” Cell and Bioscience 9 (2019): 52.

[79] P. Chen, L. Lou, B. Sharma, et al., “Recent Advances on PKM2 Inhibitors and Activators in Cancer Applications,” Current Medicinal Chemistry 31 (2024): 2955-2973.

[80] X. Liu, X. Xiao, X. Han, L. Yao, and W. Lan, “A New Therapeutic Trend: Natural Medicine for Ameliorating Ischemic Stroke via PI3K/Akt Signaling Pathway,” Molecules (Basel, Switzerland) 27 (2022): 7963.

[81] H. Hu, L. Guo, J. Overholser, and X. Wang, “Mitochondrial VDAC1: A Potential Therapeutic Target of Inflammation-Related Diseases and Clinical Opportunities,” Cells 11 (2022): 3174.

[82] Y. Zhang and T. Hunter, “Roles of Chk1 in Cell Biology and Cancer Therapy,” International Journal of Cancer 134 (2014): 1013-1023.

[83] S. Xiao, M. Zhang, Y. Liang, and D. Wang, “Celastrol Synergizes with Oral Nifedipine to Attenuate Hypertension in Preeclampsia: A Randomized, Placebo-Controlled, and Double Blinded Trial,” Journal of the American Society of Hypertension 11 (2017): 598-603.

[84] a) J.-Y. Ning, B. Ma, J.-Y. Huang, L. Han, Y.-H. Shao, and F.-Y. Wang, “Integrated Network Pharmacology and Metabolomics Reveal the Action Mechanisms of Vincristine Combined with Celastrol against Colon Cancer,” Journal of Pharmaceutical and Biomedical 239 (2024): 115883. b) M.-T. Moradi, D. Altememy, M. Asadi-Samani, et al., “The Effect of Celastrol in Combination With 5-Fluorouracil on Proliferation and Apoptosis of Gastric Cancer Cell Lines,” Oncology Research 32 (2024): 1231-1237. c) B. Qi, Q. Yan, P. Zhang, et al., “Celastrol Enhances Tamoxifen Sensitivity in the Treatment of Triple Negative Breast Cancer via Mitochondria Mediated Apoptosis Pathway,” American Journal of Translational Research 15 (2023): 2703.

[85] C. A. Deane and I. R. Brown, “Induction of Heat Shock Proteins in Differentiated human Neuronal Cells Following Co-Application of Celastrol and Arimoclomol,” Cell Stress and Chaperones 21 (2016): 837-848.

[86] S. Zhu, F. Sun, P. Zhao, et al., “Brain-Targeting Biomimetic Nanoparticles for Codelivery of Celastrol and LY2157299 for Reversing Glioma Immunosuppression,” International Journal of Pharmaceutics 619 (2022): 121709.

[87] J. H. Kim, Y. Lee, I. Kim, et al., “Reducing Peptidoglycan Crosslinking by Chemical Modulator Reverts β-Lactam Resistance in Methicillin-Resistant Staphylococcus aureus,” Advancement of Science 11 (2024): 2400858.

[88] B. Ouyang, C. Shan, S. Shen, et al., “AI-Powered Omics-Based Drug Pair Discovery for Pyroptosis Therapy Targeting Triple-Negative Breast Cancer,” Nature Communications 15 (2024): 7560.

[89] L. Guo, Y. Zhang, and K. T. Al-Jamal, “Recent Progress in Nanotechnology-Based Drug Carriers for Celastrol Delivery,” Biomaterials Science 9 (2021): 6355-6380.

[90] D. Wang, T. Wang, Z. Zhang, et al., “Recent Advances in the Effects of Dietary Polyphenols on Inflammation In Vivo: Potential Molecular Mechanisms, Receptor Targets, Safety Issues, and Uses of Nanodelivery System and Polyphenol Polymers,” Current Opinion in Food Science 48 (2022): 100921.

[91] a) G. Fang and B. Tang, “Current Advances in the Nano-Delivery of Celastrol for Treating Inflammation-Associated Diseases,” Journal of Materials Chemistry B 8 (2020): 10954-10965. b) Y. Sun, C. Wang, X. Li, J. Lu, and M. Wang, “Recent Advances in Drug Delivery of Celastrol for Enhancing Efficiency and Reducing the Toxicity,” Frontiers in Pharmacology 15 (2024): 1137289.

[92] Y. Zhao, Y. Tan, T. Meng, et al., “Simultaneous Targeting Therapy for Lung Metastasis and Breast Tumor by Blocking the NF-κB Signaling Pathway Using Celastrol-Loaded Micelles,” Drug Delivery 25 (2018): 341-352.

[93] Z. Li, R. Liu, X. Zhang, et al., “An Injectable Thermoresponsive-Hydrogel for Lamellar Keratoplasty: In-Situ Releases Celastrol and Hampers Corneal Scars,” Journal of Controlled Release 369 (2024): 604-616.

[94] M. A. Abdelmoneem, M. M. Abd Elwakil, S. N. Khattab, et al., “Lactoferrin-Dual Drug Nanoconjugate: Synergistic Anti-Tumor Efficacy of Docetaxel and the NF-κB Inhibitor Celastrol,” Materials Science and Engineering, C: Biomimetic and Supramolecular Systems 118 (2021): 111422.

[95] P. Hua, R. Liang, S. Yang, Y. Tu, and M. Chen, “Microneedle-Assisted Dual Delivery of PUMA Gene and Celastrol for Synergistic Therapy of Rheumatoid Arthritis through Restoring Synovial Homeostasis,” Bioactive Materials 36 (2024): 83-95.

[96] M. Liu, Z. Luo, Z. Li, et al., “Engineered Celastrol and Plasmid Co-Delivery for In Situ Expression and Targeted Mitochondrial Relocation of Nur77 Protein Towards Effective Drug Resistance Reversion,” Chemical Engineering Journal 453 (2023): 139879.

[97] Z. Li, Z. Guo, D. Chu, et al., “Celastrol-Based Nanoporous Membranes Prevent Subconjunctival Fibrosis by Activating Autophagy,” Materials Today Advances 18 (2023): 100356.

[98] H. Zeng, X. Zeng, C. Wang, et al., “Combination Therapy Using Cel-CSO/Taxol NPs for Reversing Drug Resistance in Breast Cancer Through Inhibiting PI3K/AKT/NF-κB/HIF-1α Pathway,” Drug Delivery and Translational Research 15 (2024): 992-1010.

[99] a) X. Guo, W. Luo, L. Wu, et al., “Natural Products From Herbal Medicine Self-Assemble Into Advanced Bioactive Materials,” Advancement of Science 11 (2024): 2403388. b) Y. L. Wang, Y. Mu, Y. L. Zhang, et al., “Accessible and Effective Nanomedicines: Self-Assembly Products From Chinese Herbal Medicines (CHMs),” Advanced Functional Materials 35 (2024): 2416151.

[100] J. Tian, K. Chen, Q. Zhang, et al., “Mechanism of Self-Assembled Celastrol-Erianin Nanomedicine for Treatment of Breast Cancer,” Chemical Engineering Journal 499 (2024): 155709.

[101] L. Wu, W. Pi, X. Huang, et al., “Orchestrated Metal-Coordinated Carrier-Free Celastrol Hydrogel Intensifies T Cell Activation and Regulates Response to Immune Checkpoint Blockade for Synergistic Chemo-immunotherapy,” Biomaterials 312 (2025): 122723.

[102] A. Mateus, N. Kurzawa, J. Perrin, G. Bergamini, and M. M. Savitski, “Drug Target Identification in Tissues by Thermal Proteome Profiling,” Annual Review of Pharmacology 62 (2022): 465-482.

[103] Q.-W. Luo, L. Yao, L. Li, et al., “Inherent Capability of Self-Assembling Nanostructures in Specific Proteasome Activation for Cancer Cell Pyroptosis,” Small 19 (2023): 2205531.

[104] M. Kurata, K. Yamamoto, B. S. Moriarity, M. Kitagawa, and D. A. Largaespada, “CRISPR/Cas9 Library Screening for Drug Target Discovery,” Journal of Human Genetics 63 (2018): 179-186.

[105] X. Zhang, K. Wang, S. Wu, et al., “Highly Effective Identification of Drug Targets at the Proteome Level by pH-dependent Protein Precipitation,” Chemical Science 13 (2022): 12403-12418.

[106] L. Yang, C.-W. Guo, Q.-M. Luo, et al., “Thermostability-assisted Limited Proteolysis-Coupled Mass Spectrometry for Capturing Drug Target Proteins and Sites,” Analytica Chimica Acta 1312 (2024): 342755.

[107] a) Y. You, X. Lai, Y. Pan, et al., “Artificial Intelligence in Cancer Target Identification and Drug Discovery,” Signal Transduction and Targeted Therapy 7 (2022): 156. b) N. E. Thomford, D. A. Senthebane, A. Rowe, et al., “Natural Products for Drug Discovery in the 21st Century: Innovations for Novel Drug Discovery,” International Journal of Molecular Sciences 19 (2018): 1578.

[108] A. Koeberle and O. Werz, “Multi-Target Approach for Natural Products in Inflammation,” Drug Discovery Today 19 (2014): 1871-1882.

[109] H. Yao, J. Liu, S. Xu, Z. Zhu, and J. Xu, “The Structural Modification of Natural Products for Novel Drug Discovery,” Expert Opinion on Drug Discovery 12 (2017): 121-140.

RIGHTS & PERMISSIONS

2025 The Author(s). Exploration published by Henan University and John Wiley & Sons Australia, Ltd.