Auranofin Induces Heme Oxygenase-1-mediated Ferroptosis in Vestibular Schwannoma cells = 청신경종양 세포에서 Auranofin의 HO-1 매개 페롭토시스 유도 연구

다국어 초록 (Multilingual Abstract)

Vestibular schwannoma (VS) is a benign neoplasm originating from Schwann cells of the vestibulocochlear nerve. Although non-malignant, VS can cause progressive hearing loss, dizziness and imbalance. Despite these clinical manifestations, no FDA-approved pharmacological treatments are currently available. In this study, we performed high-throughput screening of an FDA-approved drug library to identify compounds capable of reducing the viability of VS cells. Through this screening, auranofin (AF), an antirheumatic agent, emerged as a compound that markedly inhibited VS cell viability. Importantly, AF suppressed VS cell proliferation more potently than aspirin or mifepristone, both of which have been reported previously as effective therapeutic agents. In addition, AF significantly inhibited the growth of patient-derived primary VS cells. AF increased the expression of multiple ferroptosis-related genes in VS cells. AF also promoted intracellular iron accumulation and lipid peroxidation, characteristic markers of ferroptosis, both of which were attenuated by co-treatment with the ferroptosis inhibitor ferrostatin-1. AF increased the expression of heme oxygenase-1 (HO-1), the enzyme responsible for converting heme into biliverdin, ferrous ion, and carbon monoxide. The upregulation of HO-1 expression was closely linked to AF-mediated suppression of VS cell viability. However, pre-treatment with HO-1 inhibitors significantly attenuated the inhibitor effects of AF on cellular proliferation and iron accumulation. In in vivo models, AF suppressed tumor progression and increased the levels of multiple ferroptosis-related markers, including HO-1, in tumor tissues. AF exhibited enhanced antitumor efficacy when combined with hemin, an HO-1 inducer, but this effect was reversed by zinc protoporphyrin, an HO-1 inhibitor, treatment. Taken together, these results demonstrate that AF induces ferroptosis in VS cells via HO-1 upregulation and may serve as a viable therapeutic strategy for VS.

목차 (Table of Contents)

• List of Tables ⅲ
• List of Figures ⅳ
• ABSTRACT ⅴ
• Ⅰ. Introduction 1
• Ⅱ. Materials and Methods 4
• 1. Cell culture and reagents 4
• 2. Human primary VS cells isolation and culture 4
• 3. High-throughput screening (HTS) 4
• 4. Cell viability and EdU labeling assay 5
• 5. Annexin V apoptosis assay 5
• 6. Establishment of stable cell line 5
• 7. Thioredoxin reductase assay 6
• 8. Hydrogen peroxide assay and DCF-DA staining 6
• 9. Reverse transcription-quantitative polymerase chain reaction(RT-qPCR) 6
• 10. Intracellular iron assay and FerroOrange staining 7
• 11. Lipid peroxidation assay 8
• 12. Western blot analysis 8
• 13. Immunofluorescence staining 8
• 14. Allograft model 9
• 15. Statistical analysis 9
• Ⅲ. Results 10
• 1. HTS analysis of FDA-approved drug libraries reveals AF as a potential agent with therapeutic efficacy in VS cells 10
• 2. AF potently suppresses VS cell proliferation and promotes cell death 10
• 3. AF enhances ferroptosis in VS cells 11
• 4. HO-1 overexpression markedly facilitates AF-induced ferroptosis in VS cells 13
• 5. The TrxR-ROS-Nrf2/HO-1 axis plays a critical role in mediating AF-induced ferroptosis in VS cells 14
• 6. HO-1 expression plays an essential role in mediating the antitumor activity of AF against VS cells in vivo 14
• Ⅳ. Discussion 29
• Ⅴ. References 32
• 국문요약 37