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postgraduate thesis: Transketolase (TKT), a critical enzyme in the pentose phosphate pathway (PPP), counteracts oxidative stress to drive hepatocellular carcinoma (HCC) progression
Title | Transketolase (TKT), a critical enzyme in the pentose phosphate pathway (PPP), counteracts oxidative stress to drive hepatocellular carcinoma (HCC) progression |
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Authors | |
Issue Date | 2016 |
Publisher | The University of Hong Kong (Pokfulam, Hong Kong) |
Citation | Xu, M. [徐明婧]. (2016). Transketolase (TKT), a critical enzyme in the pentose phosphate pathway (PPP), counteracts oxidative stress to drive hepatocellular carcinoma (HCC) progression. (Thesis). University of Hong Kong, Pokfulam, Hong Kong SAR. |
Abstract | Hepatocellular carcinoma (HCC) is the sixth most common malignancy in the world, causing approximately 745,000 deaths annually. Sorafenib is the only effective targeted therapy for advanced HCC to date. HCC progression is sustained by the alteration of a series of metabolic reactions. Understanding the metabolic networks of HCC cells will facilitate the design of novel therapeutic strategy.
The pentose phosphate pathway (PPP) is a pivotal metabolic pathway in glucose metabolism, which diverges from glycolysis for the production of NADPH, the major cellular antioxidant, and ribose-5-phosphate, the nucleotide precursor. Transketolase (TKT), the reversible enzyme bridges the PPP with glycolysis, is also the key enzyme controlling the metabolic flux and direction of PPP. In this study, we demonstrated that TKT was highly up-regulated in HCC tumors and other solid cancers. Overexpression of TKT was closely associated with poor clinical outcomes in HCC patients, including larger tumor size, venous invasion, microsatellite formation and absence of tumor encapsulation, suggesting that TKT played a critical role in HCC development.
At the regulatory level, we found that TKT was regulated by NRF2/KEAP1/BACH1 pathway. NRF2 and BACH1 are two critical transcription regulators in oxidative stress defense. NRF2 and BACH1 control the expression of a variety antioxidant genes by binding with the antioxidant response element (ARE). By chromatin immunoprecipitation, we found that NRF2 and BACH1 competitively bound to two identical AREs in TKT. Expression studies further showed that NRF2 worked as a positive regulator of TKT whereas BACH1 worked as a negative regulator of TKT.
Functionally, genetic ablation of TKT resulted in an evident reduction in the HCC cell growth both in vitro and in vivo. Metabolic assays revealed that stable knockdown of TKT reduced glucose uptake and NADPH generation, escalated intracellular reactive oxygen species (ROS) level and caused ROS-induced cell cycle delay. By utilizing metabolomics and metabolic flux analysis, we disclosed that loss of TKT disrupted the connection between glycolysis and the PPP, leading to the subsequent decrease in NADPH production.
Therapeutically, genetic knockdown and pharmacological inhibition of TKT by shRNA and oxythiamine (OT), respectively, sensitized HCC cells to sorafenib. Moreover, our results showed that knockdown or inhibition of TKT enhanced the sorafenib induced ROS production, whereas treatment with antioxidant diminished the efficiency of sorafenib.
Furthermore, knockdown of TKT in normal hepatocyte cells did not affect proliferation, NADPH production, ROS level or their sensitivity towards sorafenib treatment, suggesting that co-treatment of OT and sorafenib was a potent and specific therapeutic strategy for HCC.
Taken together, we demonstrated that TKT counteracted oxidative stress to promote HCC progression, and suggested that blockage of the PPP by inhibiting TKT represents a potential therapeutic strategy for the treatment of HCC.
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Degree | Doctor of Philosophy |
Subject | Liver - Cancer - Pathogenesis Transketolase |
Dept/Program | Pathology |
Persistent Identifier | http://hdl.handle.net/10722/249189 |
DC Field | Value | Language |
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dc.contributor.author | Xu, Mingjing | - |
dc.contributor.author | 徐明婧 | - |
dc.date.accessioned | 2017-11-01T07:38:41Z | - |
dc.date.available | 2017-11-01T07:38:41Z | - |
dc.date.issued | 2016 | - |
dc.identifier.citation | Xu, M. [徐明婧]. (2016). Transketolase (TKT), a critical enzyme in the pentose phosphate pathway (PPP), counteracts oxidative stress to drive hepatocellular carcinoma (HCC) progression. (Thesis). University of Hong Kong, Pokfulam, Hong Kong SAR. | - |
dc.identifier.uri | http://hdl.handle.net/10722/249189 | - |
dc.description.abstract | Hepatocellular carcinoma (HCC) is the sixth most common malignancy in the world, causing approximately 745,000 deaths annually. Sorafenib is the only effective targeted therapy for advanced HCC to date. HCC progression is sustained by the alteration of a series of metabolic reactions. Understanding the metabolic networks of HCC cells will facilitate the design of novel therapeutic strategy. The pentose phosphate pathway (PPP) is a pivotal metabolic pathway in glucose metabolism, which diverges from glycolysis for the production of NADPH, the major cellular antioxidant, and ribose-5-phosphate, the nucleotide precursor. Transketolase (TKT), the reversible enzyme bridges the PPP with glycolysis, is also the key enzyme controlling the metabolic flux and direction of PPP. In this study, we demonstrated that TKT was highly up-regulated in HCC tumors and other solid cancers. Overexpression of TKT was closely associated with poor clinical outcomes in HCC patients, including larger tumor size, venous invasion, microsatellite formation and absence of tumor encapsulation, suggesting that TKT played a critical role in HCC development. At the regulatory level, we found that TKT was regulated by NRF2/KEAP1/BACH1 pathway. NRF2 and BACH1 are two critical transcription regulators in oxidative stress defense. NRF2 and BACH1 control the expression of a variety antioxidant genes by binding with the antioxidant response element (ARE). By chromatin immunoprecipitation, we found that NRF2 and BACH1 competitively bound to two identical AREs in TKT. Expression studies further showed that NRF2 worked as a positive regulator of TKT whereas BACH1 worked as a negative regulator of TKT. Functionally, genetic ablation of TKT resulted in an evident reduction in the HCC cell growth both in vitro and in vivo. Metabolic assays revealed that stable knockdown of TKT reduced glucose uptake and NADPH generation, escalated intracellular reactive oxygen species (ROS) level and caused ROS-induced cell cycle delay. By utilizing metabolomics and metabolic flux analysis, we disclosed that loss of TKT disrupted the connection between glycolysis and the PPP, leading to the subsequent decrease in NADPH production. Therapeutically, genetic knockdown and pharmacological inhibition of TKT by shRNA and oxythiamine (OT), respectively, sensitized HCC cells to sorafenib. Moreover, our results showed that knockdown or inhibition of TKT enhanced the sorafenib induced ROS production, whereas treatment with antioxidant diminished the efficiency of sorafenib. Furthermore, knockdown of TKT in normal hepatocyte cells did not affect proliferation, NADPH production, ROS level or their sensitivity towards sorafenib treatment, suggesting that co-treatment of OT and sorafenib was a potent and specific therapeutic strategy for HCC. Taken together, we demonstrated that TKT counteracted oxidative stress to promote HCC progression, and suggested that blockage of the PPP by inhibiting TKT represents a potential therapeutic strategy for the treatment of HCC. | - |
dc.language | eng | - |
dc.publisher | The University of Hong Kong (Pokfulam, Hong Kong) | - |
dc.relation.ispartof | HKU Theses Online (HKUTO) | - |
dc.rights | The author retains all proprietary rights, (such as patent rights) and the right to use in future works. | - |
dc.rights | This work is licensed under a Creative Commons Attribution-NonCommercial-NoDerivatives 4.0 International License. | - |
dc.subject.lcsh | Liver - Cancer - Pathogenesis | - |
dc.subject.lcsh | Transketolase | - |
dc.title | Transketolase (TKT), a critical enzyme in the pentose phosphate pathway (PPP), counteracts oxidative stress to drive hepatocellular carcinoma (HCC) progression | - |
dc.type | PG_Thesis | - |
dc.description.thesisname | Doctor of Philosophy | - |
dc.description.thesislevel | Doctoral | - |
dc.description.thesisdiscipline | Pathology | - |
dc.description.nature | published_or_final_version | - |
dc.identifier.doi | 10.5353/th_991043962678203414 | - |
dc.date.hkucongregation | 2017 | - |
dc.identifier.mmsid | 991043962678203414 | - |