Inhibition of SIRT7 overcomes sorafenib acquired resistance by suppressing ERK1/2 phosphorylation via the DDX3X-mediated NLRP3 inflammasome in hepatocellular carcinoma = SIRT7 의 억제는 간세포암종에서 DDX3X 매개의 NLRP3 염증조절복합체를 통한 ERK1/2 인산화를 억제함으로써 sorafenib 의 후천성 저항성을 극복할 수 있다|RISS 상세보기

다국어 초록 (Multilingual Abstract)

Sirtuin 7 (SIRT7) plays an important role in tumor development and it has been characterized as a potent regulator of cellular stress which has a critical role in stress responses and tumor cell death. However, little is known for the effect of SIRT7 on sorafenib acquired resistance and a possible anti-tumor mechanism beyond this process in HCC has not been clarified. In this study, we examined the therapeutic potential of SIRT7 and determined whether it functions synergistically with sorafenib to overcome chemoresistance. Here, we show that SIRT7 is up-regulated in sorafenib resistant HCC and its inhibition combined with sorafenib effectively restores sorafenib sensitivity. Further, acquired sorafenib resistance is associated with a high level of ERK1/2 phosphorylation and hyperactivate ERK1/2 is controlled by SIRT7-mediated deacetylation of DDX3X regulating NLRP3 inflammasome produced il-1β signaling. Mechanistically, SIRT7 inhibition mediated DDX3X depletion can re-sensitize acquired sorafenib resistance by disrupting NLRP3 inflammasome assembly, ultimately suppressing hyperactive ERK1/2 signaling in response to NLRP3 inflammasome-mediated IL-1β inhibition.
This preclinical study provides the functional role of SIRT7 that acts as a key effector molecule to regulate ERK1/2 signaling activation by mediating DDX3X deacetylation in sorafenib acquired resistant HCC cells. Suppression of SIRT7 combined with sorafenib could effectively restore sorafenib sensitivity. Thus, the combination therapy identified here may represent a promising strategy for patients with advanced HCC with sorafenib resistance.

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목차 (Table of Contents)

1. Introduction 7
2. Method and Materials 10
2.1. Cell line 10
2.2. Mouse models 10
2.3. Cell proliferation assay 11

1. Introduction 7
2. Method and Materials 10
2.1. Cell line 10
2.2. Mouse models 10
2.3. Cell proliferation assay 11
2.4. Colony formation assay 11
2.5. Plasmids and shRNA Retroviral transduction 12
2.6. Mass spectrometry analysis 12
2.7. Site directed mutagenesis 13
2.8. In Vitro Acetylation and Deacetylation Assays 13
2.9. Immunoprecipitation assay 14
2.10. Immunofluorescence assay 15
2.11. Protein lysate preparation and western blotting 15
2.12. Isolation of RNA and qRT-PCR 16
2.13. RNA sequencing 16
2.14. Mouse xenografts-SIRT7i treatment with sorafenib 17
2.15. Mouse xenografts–DDX3Xi (RK-33) treatment with sorafenib 17
2.16. Immunohistochemical staining 18
2.17. Statistical Analysis 18
2.18. Data and code availability 19
3. Results 20
3.1. TCGA liver HCC data implied that SIRT7 expression may be associated with sorafenib resistance 20
3.2. Loss-of-SIRT7 effectively re-sensitized HCC to sorafenib 25
3.3. SIRT7 inhibitors combined with sorafenib drove potent synergistic cell death by suppressing pERK1/2 signaling 30
3.4. Suppression of SIRT7 inhibited tumor growth in vivo 38
3.5. DDX3X was found to be a deacetylation target of SIRT7 41
3.6. SIRT7-mediated DDX3X modification was responsible for regulating ERK1/2 signaling 46
3.7. Loss-of-SIRT7 suppressed pERK1/2 through DDX3X-mediated NLRP3 inflammasome 50
4. Discussion 55
5. References 66

참고문헌 (Reference)

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