Objective
The development of T1D is associated with a rise in inflammatory landscape and oxidative stress. These processes induce the death of beta-cells, which leads to the onset of T1D. We study the role of lipid signaling on beta-cell survival and find that T1D development is associated with increases in a number of harmful lipids. Among them, we identified two particular lipids whose production by beta-cells and inflammatory macrophages were increased during the pre-diabetic phase. While the beneficial effects of inhibiting the signaling of these lipids, thereby mitigating inflammation and oxidative stress, are recognized, the potential benefits of inhibiting their signaling to prevent or delay T1D onset have, to date, not been examined. Therefore, our objective will be to determine if blocking their lipid signaling using multiple (chemical and genetic) approaches will prevent or delay the onset of T1D. Our studies will include (a) characterizing the impact of their signaling on beta-cells and immune cells, (b) assessing the effects of inhibiting their signaling on inflammatory and oxidative stress outcomes in these cells, and (c) employing novel approaches (CRISPR-Cas9), that can be translated to humans, to reduce their signaling in the whole animal and monitoring the impact of such interventions on T1D incidence.
We propose to exploit pharmacological small molecule antagonists and inhibitors that can selectively block the lipid signaling. The selected antagonists and inhibitors are potent and beneficial in mitigating inflammation and oxidative stress in pre-clinical and clinical studies of non-T1D disorders. We will repurpose them to prevent or delay T1D onset by assessing the impact of interfering with the signaling of either of the two lipids or both lipids. This will be achieved by targeting the receptors that bind the lipids and transduce their signaling to trigger downstream pathways. Our proposed studies are expected to identify the impact of the selective lipid signaling on inducing inflammation or oxidative stress and provide the impetus to translate the findings to target the onset of T1D in humans. These efforts will be favored by the advantages of the agents to be tested being well-tolerated and orally-active.
Additionally, we will use novel molecular biology approaches (i.e., CRISPR-Cas9) to eliminate the expression of the receptors that the lipids signal through in immune cells and examine if such modifications can prevent or delay T1D onset. Our analyses will include monitoring blood sugar levels and the ability of the body to remove sugar from the blood (reflecting sugar tolerance), assessing beta-cell insulin content, quantifying damage to the islets, and determining the abundances of inflammatory lipids in the plasma. We will compare these outcomes in an autoimmune rodent model of T1D treated with vehicle (no drug) or with interventions targeting the specific receptors of the lipids. The most effective regimen will be then studied in greater detail, with the intention of moving it forward to clinical trials. The signaling pathways of the select lipids are novel targets to the T1D field and could feasibly be targeted in normoglycemic subjects at high-risk for developing diabetes in their lifetime.
Background Rationale
Type 1 diabetes (T1D) results from a loss in the ability of pancreatic islets to generate enough insulin to control the blood glucose levels. More specifically, the beta-cells in the islets, which synthesize and secrete insulin into the blood, are destroyed over time. Humans who are prone to developing T1D generate autoantibodies that can be detected in the blood. There are four major antibodies and when two or more of these antibodies are detected, these individuals are considered to be at high-risk for developing T1D at some point in their lifetime. Whereas the causes of the initiation of the process are not well-understood, they are nevertheless thought to recruit similar downstream pathways that eventually lead to the same end, death of the beta-cells.
Currently, the only effective T1D treatment is the supply of the deficient insulin via daily insulin administrations. Other therapies, including islet transplantation, are successfully being used to treat T1D, but they are not permanent solutions. In addition, immunotherapies or stem-cell derived beta-cell transplantation is a promising alternative, but limited at this time due to immuno-rejection and issues with preserving optimal function. Of note, these therapies are implemented after the subject has been diagnosed with T1D.
Our pre-clinical studies in a T1D rodent model, in which the progression to T1D development can be feasibly monitored, identified previously unrecognized sequential events that occur prior to the onset of T1D. These events include the development and progression of inflammation and oxidative stress. We found that production of certain lipids that are considered causative factors of inflammation or oxidative stress were increased prior to the onset of T1D. This raises the intriguing possibility that targeting the signaling pathways of these lipids during the pre-diabetic phase (i.e., before T1D onset) would be a feasible approach to prevent or delay the onset of T1D in humans.
To address this possibility, we have assembled a team of experts in beta-cell biology/lipid signaling (Ramanadham, PD/PI), development of lipid chemical mediators/intervention studies (Hammock), and lipid analyses in cells and blood (Chalfant). Together, we propose to determine whether (a) inhibiting the signaling mediated by these lipids mitigates inflammation and oxidative stress in beta-cells and immune cells, (b) inhibiting their signaling can reduce T1D incidence, and (c) whether novel approaches to eliminate the activation or expression of the receptors to which these lipids bind to can prevent or delay the onset of T1D.
To date, the impact of reducing the signaling of these select lipids has been investigated in non-T1D disorders, and such interventions have been reported to be effective in mitigating inflammatory/oxidative stress. However, studies to examine the involvement of these select lipids or the beneficial effects of inhibiting their signaling on the onset of T1D have not been performed. Therefore, we will exploit pharmacological and novel molecular biology approaches to target and intervene with the signaling of these lipids, as a means to prevent or delay the onset of T1D. Positive findings from these studies will facilitate our long-term goal of achieving beneficial outcomes in subjects that are still normoglycemic, but, at high-risk for developing T1D in their lifetime.
Description of Project
Type 1 diabetes (T1D), also known as insulin-dependent or juvenile diabetes, is a consequence of beta-cell death. The beta-cells, located in the pancreatic islets, synthesize and store insulin. When blood sugar levels increase (e.g., after a meal), the beta-cells are stimulated and secrete insulin, which increases sugar uptake, utilization, and storage by various organs, and the blood sugar levels return to normal levels. In T1D, destruction of beta-cells leads to a loss in the source of insulin. Thus, a meal is not followed by the needed increases in insulin secretion and this leads to elevations in the levels of blood sugar, and subsequent accumulation of sugar in various organs. Progressive increases in the organ levels of sugar can lead to various complications including abnormal heart function, kidney failure, blindness, and nerve damage in the feet and fingers.
Current regimens are aimed at treating patients that have been diagnosed with TID, with insulin therapy being the most effective in controlling the complications associated with T1D. Other approaches include islet transplantation, which may require several donors per transplantation and the need for multiple procedures over a lifetime. Developing immunotherapy or stem-cell derived beta-cell replacement protocols has been challenging and fraught with rejection issues. Thus, there is continued need to explore new avenues to manage T1D.
Many factors (e.g., genetic, environmental, and lifestyles) can trigger T1D onset. In all cases, they induce an autoimmune attack of the beta-cells, where specialized cells in one’s own body kill the beta-cells. Studies to identify the mechanisms that lead to beta-cell death have largely focused on the immune system. However, our work has identified critical roles for lipids (or fat) signaling in modulating beta-cell function and survival. The Principal Investigator of this proposal started on this journey with a JDRFI award in 1994 that led to the identification of an enzyme, whose activation leads to the generation of these lipids. Our continued work on this enzyme revealed that inhibiting the production of these lipids by inactivating the enzyme resulted in a reduced incidence of T1D. In the current proposal, we aim to test whether we can modify the signaling of selective lipids that arise through activation of this enzyme, to prevent or delay the onset of T1D. Our motivation for this proposal is based on the following: (a) these lipids are important contributors to inflammation and oxidative stress, which are critical that inducers of beta-cell death, leading to T1D; (b) inhibition of their lipid signaling has been reported to mitigate inflammatory and stress landscapes in non-T1D disorders; (c) these lipids are elevated during the pre-diabetic phase, suggesting that they could be considered as potential biomarkers of T1D; and (d) signaling through these lipids can be inhibited by commercially-available orally-active drugs that we are proposing to use as probe compounds that could be repurposed to prevent or delay T1D onset. Importantly, the goal of our study aligns with the mission of this RFA in that the chosen lipids for study, to date, have not been targeted, in the context of preventing or delaying the onset of T1D.
While current therapeutics can alleviate complications associated with T1D, they cannot prevent or delay T1D onset. Therefore, our novel perspective of targeting selective lipid signaling could be a means to prevent or delay T1D onset. Such therapeutics would also preclude development of autoimmunity following transplantations. Importantly, targeting lipid signaling that are elevated during the pre-diabetic phase when inflammation and oxidative stress are evolving, will offer novel strategies to prevent or delay the course of T1D onset in the human population, and lead to a paradigm shift in T1D therapy.
Anticipated Outcome
Our proposal is designed to address the impact of select lipids, that to date, have not been considered as contributors to T1D onset. The chosen lipids have previously been targeted to reduce inflammation and oxidative stress in non-T1D disorders. However, while inflammation and oxidative stress are critical contributors to the eventual demise of beta-cells, the possibility that these lipids may play a role in the evolution of inflammation and oxidative stress that can lead to T1D-associated beta-cell death and T1D onset has not been tested.
Our Aims are designed to address the consequences of triggering the signaling of these select lipids at the cell level and the whole animal level, in the context of preventing or delaying the onset of T1D. At the whole animal level, we will also consider the feasibility of employing novel molecular biology approaches to eliminate the lipid signaling in immune cells alone and the assessment of such modifications on the progression and onset of T1D. After first establishing the inhibition of inflammation and oxidative stress with antagonism of these lipids signaling, we will initiate intervention protocols (in an autoimmune mouse model of T1D) at 4 weeks of age, which coincides with the start of the inflammatory process. Each protocol will include a cohort that will be treated with only the vehicle (control group).
We expect that targeting signaling of the identified select lipids will mitigate inflammatory and oxidative stress landscapes in beta-cells and immune cells. We also predict that intervention of individual lipid signaling will lead to a decreased inflammatory profile or oxidative stress in the autoimmune T1D mouse model. Moreover, we anticipate that intervening with the lipid signaling of either or both lipids will lead to mitigation of T1D outcomes and prevent or delay T1D onset.
Beneficial positive impacts of the proposed interventions should be viewed as paradigm shifting in the field of T1D therapy and be expected to provide motivation to examine in greater detail the extent of lipid signaling contribution to the onset of T1D. They would also provide strong rationale to translate the regimen of including the blocking of select lipid signals to clinical trials of subjects diagnosed at high-risk for developing T1D.
Relevance to T1D
Current therapeutic strategies for T1D encompass approaches to treat subjects diagnosed with diabetes with agents to reduce the severity of complications associated with the disease. As such, treatment regimens are initiated following the diagnosis of T1D. Irrespective of the triggers of T1D onset, all of the downstream processes lead to the induction of inflammation and stress responses that cause beta-cell death and frank T1D. Therefore, our goal here is to target these downstream events (i.e., those that lead to T1D onset), so that the onset of T1D is prevented or delayed. If our approach is successful, the need to develop or employ protocols to reduce autoimmune responses that are triggered by transplantation of preparations donated by foreign sources could be re-evaluated. Importantly, it would highlight the potential for modulating novel targets during the pre-diabetic phase with classes of bio-available agents, that are already available and are constantly being improved, as a means to prevent or delay the onset of T1D. As such, our work would have a significant impact in the T1D field and contribute critical insights towards the preventing or delaying the onset of T1D.