Periodic Reporting for period 4 - METAPTPs (PROTEIN TYROSINE PHOSPHATASES IN METABOLIC DISEASES: OXIDATION, DYSFUNCTION AND THERAPEUTIC POTENTIAL)
Período documentado: 2023-10-01 hasta 2025-03-31
The hypothesis of the ERC Consolidator project METAPTPs is that specific signalling pathways are dysregulated by obesity, inflammation and oxidative stress in the pancreas and liver. The oxidative stress caused by inflammation and/or obesity affects the activity of important metabolic molecules known as protein tyrosine phosphatases, which could trigger the processes that cause metabolic diseases such as diabetes and liver dysfunction. To test our hypothesis, we developed tools which will allow us to identify major inactivated metabolic proteins and their role in cellular responses. We specifically focussed on the role of protein tyrosine phosphatases in the pancreas in type 1 and type 2 diabetes and in the liver for obesity-associated hepatic dysfunction. We have shown that inactivation of phosphatases by inflammatory-mediated oxidative stress severely affects pancreatic β-cell function and survival in the context of type 1 diabetes (Figure 1). Testing our hypothesis that protein tyrosine phosphatase dysregulated activity is a common mechanism in autoimmunity will increase our understanding of the pathogenesis and postulate novel targets for future clinical interventions in type 1 diabetes. For instance, identification of the mechanisms by which protein tyrosine phosphatases modulate cytokine activity may provide relevant and realistic targets for the design of pharmacological agents and diets. The present project has the potential to explain many of the “unknowns” that still linger regarding the disease. In the METAPTPs project, we take advantage of human stem cells and CRISPR/Cas gene editing to directly elucidate mechanisms modulated by aberrant signalling in differentiated metabolic cells. The differentiation method has been successfully implemented in our lab with the generation of insulin-producing β-cells and albumin-producing hepatocytes. Our knowledge accumulated in recent years enable us to study the mechanisms by which different subcomponents of the JAK-STAT signalling influence obesity and type 2 diabetes, i.e. acting as regulators of β-cell function (Figure 2). In addition, based on our previous data and unpublished observations, we predict that research strategies based on a family of proteins tyrosine phosphatases will allow us to better understand the transition from fat accumulation in the hepatocytes to hepatic cancer development (Figure 3). This information will be critical for new pharmacology screening. We aim to restore protein tyrosine phosphatase activity and associated dysfunctional signalling in patients.
Due to the COVID-19 pandemic, the recruitment and starting of the project was significantly affected. Nonetheless, the ERC Consolidator grant allowed to develop the research line of the Signal Transduction and Metabolism Laboratory at the ULB-Faculty of Medicine. The research group has currently 5 Postdoctoral Fellows, 5 PhD students, 4 technicians and 1 secretary.
Here are the advances and problems in the different work packages of the project:
WP1: Identify the major oxidised PTPs in metabolic relevant tissues/cells in both obesity and diabetes: We have set up the technique to measure PTP expression in liver biopsies by mass spectrometry (under patent) and obtained preliminary results of dysregulated PTPs in the human liver samples (under patent).
WP2: Determine the contribution of PTP inactivation in cellular responses to metabolic signalling in human samples. We have set up the differentiation of Stem Cells into hepatocyte-like and beta-like cells and obtained preliminary results. It was needed to develop a novel Stem Cell facility to develop our research project. We have obtained important mechanistic results about the role of protein tyrosine phosphatases in the control of inflammatory-mediated beta cell dysfunction and obesity-induced liver dysfunction.
WP3: Assess the impact of tissue-specific PTP deficiency on the development of obesity and diabetes. We have generated novel flox mice for inactivation of different protein tyrosine phosphatases identified by mass spectrometry, the breeding with the respective CRE lines was delayed (due to the pandemic) and started in early 2021. In addition, we set up the metabolic studies in the facility allocated for our project and performed pilot studies. It was not possible to run big numbers of mouse studies due to the ongoing construction of the new ULB animal facility (the studies are thus programmed for early 2022).
WP4: Test novel therapeutic approaches targeting PTPs to prevent/reverse metabolic disorders. Due to the problems in the animal facility, we established a collaboration with colleagues from the KU Leuven to develop our studies with special diets in the autoimmune model of NOD mice in their animal facility. We have preliminary results of the effects of special diets (tested in a pilot clinical trial) on diabetes development, additional experiments are currently ongoing.
Here are the advances and problems in the different work packages of the project:
WP1: Identify the major oxidised PTPs in metabolic relevant tissues/cells in both obesity and diabetes: We have set up the technique to measure PTP expression in liver biopsies by mass spectrometry (under patent) and obtained preliminary results of dysregulated PTPs in the human liver samples (under patent).
WP2: Determine the contribution of PTP inactivation in cellular responses to metabolic signalling in human samples. We have set up the differentiation of Stem Cells into hepatocyte-like and beta-like cells and obtained preliminary results. It was needed to develop a novel Stem Cell facility to develop our research project. We have obtained important mechanistic results about the role of protein tyrosine phosphatases in the control of inflammatory-mediated beta cell dysfunction and obesity-induced liver dysfunction.
WP3: Assess the impact of tissue-specific PTP deficiency on the development of obesity and diabetes. We have generated novel flox mice for inactivation of different protein tyrosine phosphatases identified by mass spectrometry, the breeding with the respective CRE lines was delayed (due to the pandemic) and started in early 2021. In addition, we set up the metabolic studies in the facility allocated for our project and performed pilot studies. It was not possible to run big numbers of mouse studies due to the ongoing construction of the new ULB animal facility (the studies are thus programmed for early 2022).
WP4: Test novel therapeutic approaches targeting PTPs to prevent/reverse metabolic disorders. Due to the problems in the animal facility, we established a collaboration with colleagues from the KU Leuven to develop our studies with special diets in the autoimmune model of NOD mice in their animal facility. We have preliminary results of the effects of special diets (tested in a pilot clinical trial) on diabetes development, additional experiments are currently ongoing.
Our preliminary and published results and published data indicate that changes in protein tyrosine phosphatases induced by obesity and inflammation in the pancreas and liver affect several pathways that are altered in both metabolic and inflammatory diseases suggesting common pathogenic mechanisms. Indeed, many human pathologies are characterized by inflammation, oxidative stress and potential phosphatase inactivation. Thus, results of METAPTPs might have wide-ranging implications for the diagnostic and treatment of several metabolic diseases.
Expected results of METAPTPs until the end of the project include:
-We will clarify for the first time the expression and oxidation of protein tyrosine phosphatases during the pathogenesis of autoimmune diabetes in the pancreas and obesity-associated liver dysfunction in human samples. The experiments will provide information about the degree of expression/oxidation of the proteins at different stages of the disease development. The results will provide important knowledge for mechanistic fundamental research.
-We will establish whether differential expression and activity of protein tyrosine phosphatases in autoimmunity and obese livers or pancreas allow stratification and risk diagnostic of disease progression. In addition, gene expression studies and functional analysis are currently developed in the METAPTPs project to determine the networks of modulated genes in inflammation and obesity.
-We expect to develop a risk diagnostic assay intended for identifying metabolic-associated disease subtypes and monitoring patients in response to therapy. The assay will lead to a better understanding of the disorder and implementation of personalized therapeutic strategies to improve survival. The Technology Transfer Office of the ULB is supporting Dr Gurzov to assess commercially knowledge generated by the project and will provide legal assistance. We are on track to demonstrate that inactivation of candidate proteins for metabolic disorders by inflammatory/obesity-mediated oxidative stress in beta and liver cells is a key contributing factor to the development of the pathogenesis. We will determine the exact molecular pathways leading to inactivation of these candidate proteins and how to prevent it. Accordingly, we will establish a solid mechanistic background to test different protocols that will ultimately improve therapeutics in diabetes and obesity.
Expected results of METAPTPs until the end of the project include:
-We will clarify for the first time the expression and oxidation of protein tyrosine phosphatases during the pathogenesis of autoimmune diabetes in the pancreas and obesity-associated liver dysfunction in human samples. The experiments will provide information about the degree of expression/oxidation of the proteins at different stages of the disease development. The results will provide important knowledge for mechanistic fundamental research.
-We will establish whether differential expression and activity of protein tyrosine phosphatases in autoimmunity and obese livers or pancreas allow stratification and risk diagnostic of disease progression. In addition, gene expression studies and functional analysis are currently developed in the METAPTPs project to determine the networks of modulated genes in inflammation and obesity.
-We expect to develop a risk diagnostic assay intended for identifying metabolic-associated disease subtypes and monitoring patients in response to therapy. The assay will lead to a better understanding of the disorder and implementation of personalized therapeutic strategies to improve survival. The Technology Transfer Office of the ULB is supporting Dr Gurzov to assess commercially knowledge generated by the project and will provide legal assistance. We are on track to demonstrate that inactivation of candidate proteins for metabolic disorders by inflammatory/obesity-mediated oxidative stress in beta and liver cells is a key contributing factor to the development of the pathogenesis. We will determine the exact molecular pathways leading to inactivation of these candidate proteins and how to prevent it. Accordingly, we will establish a solid mechanistic background to test different protocols that will ultimately improve therapeutics in diabetes and obesity.