Study on identification of RIPK1 inhibitor for therapeutic potential : Identification of novel inhibitor of receptor-interacting protein kinase 1 (RIPK1)
- 주제(키워드) RIPK1
- 주제(DDC) 570
- 발행기관 아주대학교 일반대학원
- 지도교수 You-Sun Kim
- 발행년도 2024
- 학위수여년월 2024. 8
- 학위명 석사
- 학과 및 전공 일반대학원 의생명과학과
- 실제URI http://www.dcollection.net/handler/ajou/000000033970
- 본문언어 영어
- 저작권 아주대학교 논문은 저작권에 의해 보호받습니다.
초록/요약
Study on identification of RIPK1 inhibitor for therapeutic potential RIPK1 stands out as a pivotal player controlling cell death and inflammation. Moreover, given that inhibition of RIPK1 kinase activity has been shown to be effective in animal models of human diseases such as autoimmune and neurodegenerative disorders, RIPK1 has been considered as an attractive therapeutic target. Specifically, owing to its unique allosteric pocket, RIPK1 kinase is ripe for small-molecule inhibitors. As a representative example, necrostatin-1 (Nec-1), a first small-molecule inhibitor of RIPK1 kinase, has been extensively used to exploring the function of RIPK1. Herein, we screened a collection of approved drug and ongoing clinical trial drugs with structural similarity to Nec-1 to assess their ability to regulate RIPK1-mediated cell death. Through this small-scale screen, we discovered that Phensuximide, already FDA approved drug as anti-epilepsy, can prevent with necroptosis by targeting RIPK1 kinase. Importantly, we identified that phensuximide effectively prevents RIPK1-dependent necroptosis in human and murine cell lines without affecting the NF-κB and MAPK pathway or apoptosis. Additionally, we show that phensuximide can protect against LPS-induced systemic inflammatory response syndrome (SIRS), which is an animal model of sepsis, and it involves RIPK1 kinase activity. Overall, our findings suggest that the already FDA- approved phensuximide may offer a new strategy for targeting RIPK1-mediated diseases. Keywords: RIPK1, Inflammation, Allosteric pocket, SIRS, Phensuximide
more목차
I. INTRODUCTION 1
II. MATERIALS AND METHODS 4
A. Antibodies and chemical reagents 4
B. Cell lines and culture conditions 5
C. Immunoblot analysis and immunoprecipitation 5
D. Plasmid construction and transfection 6
E. Cytotoxicity assays 6
F. Immunofluorescence staining 6
G. Primary culture and activation of BMDMs 6
H. Flow cytometry 7
I. Quantitative RT-PCR 7
J. Histology analysis 7
K. Serum biochemistry 8
L. LPS-induced SIRS model 8
M. Structural similarity calculation 8
N. Molecular docking and dynamics simulation study 8
O. Statistical analysis 9
III. RESULTS 10
A. Identification of phensuximide as a potential RIPK 1 inhibitor. 10
B. Phensuximide shows potent inhibitory effect on TNF-induced necroptotic cell death. 15
C. Inhibition of RIPK1 kinase activity by phensuximide treatment. 23
D. Phensuximide does not involved in receptor-mediated complex I signal. 33
E. Inhibition of RIPK1 kinase activity-dependent cell death and inflammatory response by phensuximide in macrophages. 38
F. Phensuximide is a potential therapeutic candidate for necroptosis related diseases. 46
IV. DISCUSSION 52
V. REFERENCES 54
국문요약 59
