Title Page

Contents

Abstract 10

Ⅰ. Introduction 12

Ⅱ. Method and Materials 15

1. Cell line 15

2. Mouse models 15

3. Cell proliferation assay 16

4. Colony formation assay 16

5. Plasmids and shRNA Retroviral transduction 17

6. Mass spectrometry analysis 17

7. Site directed mutagenesis 18

8. In Vitro Acetylation and Deacetylation Assays 18

9. Immunoprecipitation assay 19

10. Immunofluorescence assay 19

11. Protein lysate preparation and western blotting 20

12. Isolation of RNA and qRT-PCR 21

13. RNA sequencing 21

14. Mouse xenografts-SIRT7i treatment with sorafenib 21

15. Mouse xenografts-DDX3Xi (RK-33) treatment with sorafenib 22

16. Immunohistochemical staining 22

17. Statistical Analysis 23

18. Data and code availability 24

Ⅲ. Results 25

1. TCGA liver HCC data implied that SIRT7 expression may be associated with sorafenib resistance 25

2. Loss-of-SIRT7 effectively re-sensitized HCC to sorafenib 30

3. SIRT7 inhibitors combined with sorafenib drove potent synergistic cell death by suppressing pERK1/2 signaling 35

4. Suppression of SIRT7 inhibited tumor growth in vivo 43

5. DDX3X was found to be a deacetylation target of SIRT7 46

6. SIRT7-mediated DDX3X modification was responsible for regulating ERK1/2 signaling 51

7. Loss-of-SIRT7 suppressed pERK1/2 through DDX3X-mediated NLRP3 inflammasome 55

Ⅳ. Discussion 60

References 71

국문초록 80

Table 1. Antibody list. 65

Table 2. Cell line list. 67

Table 3. Primer sequences. 68

Table 4. Other (e.g., drugs, proteins, vectors etc.). 69

Table 5. Organisms. 70

Table 6. Deposited data. 70

Table 7. Software. 70

Fig. 1. Association analysis of the transcriptional expressions in sorafenib-resistant HCC samples. 27

Fig. 2. Expression profiles of the protein deacetylase family. 29

Fig. 3. SIRT7 silencing inhibits the proliferation of sorafenib-resistant liver cancer cells. 32

Fig. 4. SIRT7 silencing inhibits proliferation of sorafenib-resistant HCC cells. 34

Fig. 5. Development of SIRT7 - targeted chemical inhibitors. 37

Fig. 6. Modeled structure of SIRT7-targeted chemical inhibitors. 39

Fig. 7. Chemical inhibitor of SIRT7 sensitizes sorafenib resistance. 40

Fig. 8. SIRT7 chemical inhibitors combined with sorafenib treatment reduce resistant cell viability. 42

Fig. 9. Combination of SIRT7i and sorafenib displays potent antiproliferative effects in a xenograft mouse model. 44

Fig. 10. DDX3X is a deacetylation target of SIRT7. 48

Fig. 11. DDX3X as the deacetylation target of SIRT7 by interactome analysis. 50

Fig. 12. SIRT7 is required to activate pERK signaling through stabilising DDX3X. 53

Fig. 13. Loss-of-SIRT7 inhibits NLRP3 inflammasome assembly to enable HCC cells to perform pro-survival signaling through DDX3X modification 57

Fig. 14. Loss-of-SIRT7 inhibits interleukin-1β expression. 59