Objective
The overall goal of this research plan is to understand whether ZBED3 is a gene that controls beta cell vulnerability and alters beta cell response to immune cell killing and inflammatory conditions. We will characterize the role of ZBED3 in beta cells with three research aims: (1). Define how ZBED3 affects immune cell killing: We will perform experiments to examine any immune cells’ behavior change in response to the ZBED3 gene deletion in beta cells. (2). Determine the activity of ZBED3 in controlling the profile of beta cell antigens: Antigens are a small fraction of protein presented on the cell membrane surface to be recognized by immune cells. The presence of antigens on the beta cell membrane surface will tell immune cells that this beta cell is “self-origin” and send a “do not kill me” signal. However, this signal is failed in individuals with T1D, and that triggers immune cell killing targeting beta cells. Therefore, in this research Aim 2, we will perform a series of experiments to understand how ZBED3 affects beta cell antigens’ presentation. This may further explain the reasons behind the outcomes we observed in Aim 1. (3). Define how ZBED3 alters beta cell response under inflammation and stress conditions: Immune cells release a group of small proteins, which induces beta cell inflammation and cellular stress, which promotes beta cell death. In this Aim3, we will examine whether the ZBED3 gene deletion makes beta cells more capable of enduring inflammatory conditions induced by immune cells. Taken together, we will establish a comprehensive view of the role of ZBED3 in protecting beta cells against immune cell killing.
Background Rationale
Type 1 diabetes (T1D) is an autoimmune disease, which means the immune system targets and destroys insulin-producing beta cells in the pancreatic specialized cell structure, islets. Loss of beta cells leads to insulin insufficiency and subsequently uncontrollable high blood sugar levels and the occurrence of diabetes. In principle, beta cells sense high glucose levels in circulation and release insulin into the blood for making organs, like the liver and skeletal muscle, absorb glucose from circulation, thereby reducing blood glucose levels. Since the discovery of insulin in 1921, insulin has been the only effective treatment for T1D. In 2000, islet transplantation surgery was first approved for its efficacy in treating T1D. However, the source of transplantable islets is limited. Nowadays, stem cell technology has advanced, which allows scientists to generate beta cells from stem cells in laboratories. However, this new therapeutic approach for T1D remains a critical challenge – autoimmunity. Without drugs that inhibit the activity of the immune system, the transplanted beta cells in individuals with T1D will be rapidly destroyed.
Our strategy to tackle this key challenge was based on the bold assumption that single gene modifications may exist to protect beta cells against immune cell killing without inhibiting the immune system in the body. We applied the state-of-art technology, CRISPR-Cas9, to search for genes that make beta cells withstand autoimmunity. We found eleven potential genes that may protect beta cells against immune cell killing. In this research project, we will examine one identified gene, ZBED3. Indeed, we found that beta cells without ZBED3 gene expression were able to sustain immune cell attacks. Deleting the ZBED3 gene also made beta cells more resistant to stress conditions. Stress has been thought to be a pathway that compromises beta cells' integrity, making beta cells more susceptible to death signals. In this research project, we will characterize the role of ZBED3 in beta cells and understand how this gene deletion protects beta cells against autoimmunity.
Description of Project
Type 1 diabetes (T1D) is an autoimmune disease in which the body’s immune system destroys insulin-producing beta cells in the pancreas. T1D is still incurable, and individuals living with T1D need daily insulin injections to maintain their blood glucose levels. Currently, scientists may use stem cells to re-direct them to become beta cells in laboratories. With the potentially unlimited supply of beta cells, the significant challenge that remained in the T1D cure is the uncontrollable immune cell-triggered beta cell killing. Without any drug inhibition on immune cell activity, the surgery-implanted beta cells will be rapidly destroyed by immune cells. However, inhibiting the whole body’s immune cell response will weaken the immunity for combatting any infections. Therefore, we used a cutting-edge genomic editing technology, CRISPR-Cas9, to search for genes that can make beta cells withstand immune cell killing. Indeed, we found that deleting a gene named ZBED3 made beta cells sustain the attack from immune cells. In this research proposal, we have designed a thorough plan to characterize and understand why this ZBED3 gene deletion may protect beta cells against immune cell killing. The research results will deliver the possibility of using ZBED3 gene modification for developing T1D therapy.
Anticipated Outcome
This research project will produce new concepts and knowledge on beta cell autoimmune protection that will contribute to beta cell biology and benefit T1D research, including (1) Zbed3, a new master gene regulator in beta cell antigen presentation. Deletion of the ZBED3 gene enables beta cells to escape from immune cell attacks. Creating immune-invisible beta cells not only requires multiple gene modifications but also is challenged by diabetes researchers due to the loss of immune cell activity in fighting infections and tumor formation. This may not be a safe option for T1D treatment. Here we provide a solution with a single modification that delivers beta cell autoimmune protection without compromising other immune reactivities. (2) Discovery of new gene networks in regulating beta cell response to stress and inflammation. Our research design will identify the truthful dependent signals of ZBED3. These results will address and demonstrate how beta cell accommodates stress condition and the fundamental knowledge in understanding the beta cell biology and how beta cell progresses to diseased condition during T1D.
Once the project is complete, we plan to use the data to leverage future funding opportunities in the form of significant grant applications. Future funding will be used to extend the information we collected from the current project. Our short-term goal will be to search for ZBED3 inhibitors. We will work with our institute's pharmaceutical biotechnology center to facilitate the identification of ZBED3 inhibitors, aiming to reduce the treatment cost and become a more practical therapeutic option for people with T1D. Our long-term goals include using ZBED3 gene deleted beta cells or ZBED3 inhibitors to reverse established T1D and examining the possibility of ZBED3 inhibitors for T1D prevention in high-risk individuals. Ultimately, our goal is to translate our research outcome from the pre-clinical stage to Phase 1 clinical trial.
Relevance to T1D
Although daily insulin injections can manage blood sugar levels in the majority of people with T1D, there is no cure for this lifelong disease. Scientists have spent decades searching for T1D cures; however, the prevalence of T1D is rising. Currently, approximately 1.6 million Americans are living with T1D. JDRF expects that 5 million Americans will be diagnosed with T1D by 2050. Therefore, new research approaches for developing new T1D therapy are urgently needed. CRISPR-Cas9 technology was introduced in 2011, and the two scientists who discovered the CRISPR-Cas9 system won the 2020 Nobel Prize in Chemistry. Using this cutting-edge technology, we searched for genes that protect beta cells against autoimmunity. Indeed, we found that a single gene deletion made beta cells survive under immune cell killing. Combining with stem cell technology, we aim to deliver a potential T1D therapeutic strategy through a genetic modification for curing T1D.