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Molecular Characterization of 3β-Hydroxy-∆5-C27-Steroid Oxidoreductase and Discovery of Novel Inhibitors for Clear Cell Renal Cell Carcinoma Sarah M Gardner
- Format:
- Book
- Thesis/Dissertation
- Author/Creator:
- Gardner, Sarah M., author.
- Language:
- English
- Subjects (All):
- Biochemistry.
- Biophysics.
- Cellular biology.
- Molecular biology.
- 0487.
- 0786.
- 0379.
- 0307.
- Local Subjects:
- Biochemistry.
- Biophysics.
- Cellular biology.
- Molecular biology.
- 0487.
- 0786.
- 0379.
- 0307.
- Physical Description:
- 1 electronic resource (155 pages)
- Contained In:
- Dissertations Abstracts International 86-07B
- Place of Publication:
- Ann Arbor : ProQuest Dissertations and Theses, 2024
- Language Note:
- English
- Summary:
- Bile acid metabolism is an essential regulator of cholesterol homeostasis and dysregulation of these pathways can result in nutrient malabsorption and bile acid toxicity. 3β-hydroxy-5-C27-steroid oxidoreductase (HSD3B7) catalyzes essential steps in the conversion of cholesterol to bile acids, converting the characteristic 3β-hydroxyl group of cholesterol on route to the canonical 3α-hydroxy group of all bile acids. Genetic defects in HSD3B7 severely impair the production of primary bile acids and result in the build up of hepatotoxic oxysterols. Additionally, we have found that diseases that rely heavily on cholesterol may be uniquely susceptible to modulators of cholesterol catabolism enzymes, including HSD3B7. Unfortunately, limited knowledge of the catalytic mechanism and substrate specificity of HSD3B7 hinders our ability to develop small molecule inhibitors. Here, we investigated the mechanism of HSD3B7 through a combination of biophysical tools, including kinetic and structural modeling and gain insight into the substrate binding pockets and the necessity of membrane association for activity. These experiments set the stage for us to investigate HSD3B7 in the context of clear cell Renal Cell Carcinoma (ccRCC). Bile acid metabolic genes were upregulated within ccRCC as a means to detoxify the oxysterol pool, highlighting a key vulnerability. Upon genetic knockdown of HSD3B7, we saw an impact on cell viability both in vitro and vivo. This led us to screen for small molecule inhibitors of HSD3B7 where we identified a potent inhibitor, celastrol. Celastrol phenocopied genetic knockdown, and resulted in significant decreases in tumor growth in vivo. Unfortunately, celastrol has many off target effects, which led us to perform a more diverse high throughput screen to identify unique chemical scaffolds for HSD3B7 inhibition. This screen identified 3 unique backbones, some of which have selectivity for HSD3B7 over other 3β-HSD enzymes. These compounds can be further developed for ideal pharmacological properties and higher potency with systematic structure activity relationship studies. Together, this work details a comprehensive biochemical characterization of HSD3B7, unveils its role in bile acid metabolism within ccRCC, and initiates the development of small molecule inhibitors as potential therapeutics
- Notes:
- Source: Dissertations Abstracts International, Volume: 86-07, Section: B.
- Advisors: Marmorstein, Ronen Committee members: Penning, Trevor M.; Murphy, Maureen E.; Christianson, David W.; Schultz, David C.
- Ph.D. University of Pennsylvania 2024
- Local Notes:
- School code: 0175
- ISBN:
- 9798302184337
- Access Restriction:
- Restricted for use by site license
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