Grants

NIDDK - National Institute of Diabetes and Digestive and Kidney Diseases

PROJECT SUMMARY Autophagy, the process of lysosomal degradation of cellular components, is a conserved homeostatic mechanism with crucial roles in the maintenance of protein homeostasis (proteostasis) and in the cellular response to stress through rapid organelle and proteome remodeling. A selective type of autophagy found in mammalian cells, chaperone-mediated autophagy (CMA), facilitates the timely and selective degradation of key soluble, cytosolic proteins involved in processes like the regulation of hepatic metabolism, the cell cycle, cell death, and the cellular stress response. Basal and inducible levels of CMA decline with age, resulting in widespread dysregulation of key cellular processes and aggravation of age-related diseases. Metabolic dysfunction-associated steatotic liver disease (MASLD) is the most common chronic liver disease in western countries. It represents the beginning of a spectrum of disease that can lead to hepatocellular carcinoma, the third leading cause of cancer death globally. Progressive forms of MASLD are now the leading etiology of liver transplantation in hepatocellular carcinoma patients. There is only one FDA-approved drug targeting this pathology, despite its high prevalence and impact. Because of CMA’s established roles in the regulation of hepatic metabolism, we hypothesized that CMA failure may result in proteostatic and metabolic alterations that favor the development of MASLD. This proposal contains exciting data supporting this hypothesis. The studies described leverage several already-established, well-characterized genetically modified mice with deficiencies in CMA and a fluorescent reporter of CMA activity, as well as small molecule chemical activators of CMA suitable for in vivo use. We aim to further elucidate CMA’s hepatoprotective role, as well as the consequences of CMA failure and their mechanisms (Aim 1), and the impact of pharmacologic CMA upregulation (Aim 2) during MASLD development and progression. This proposal will synergize (via a co-mentorship) the Arias Laboratory’s expertise in liver physiology and pathobiology and the Cuervo Laboratory’s expertise in CMA to discover previously unknown molecular mechanisms of chronic liver disease. It may further identify CMA as a novel therapeutic target to prevent liver disease progression. This fellowship funding will enable me to advance toward my goal of becoming an independent physician scientist investigator by strengthening my technical abilities; expanding my background on liver pathophysiology, metabolism, and autophagy; and improving my ability to formulate and test scientific hypotheses in a robust and reproducible manner, as well as my scientific communication and presentation skills. The group of exceptional mentors and collaborators assembled for this application will aid in assuring timely completion of the proposed project and a smooth transition into my clinical training.

NIDDK - National Institute of Diabetes and Digestive and Kidney Diseases

Project Summary/ Abstract Glucagon-like peptide-1 (Glp1) receptor (Glp1r) agonists are a promising new class of weight loss drugs for treating obesity, yet weight loss outcomes to Glp1r agonists range from 0% to over 20% of starting body weight which raises the critical question of why some individuals are more responsive than others. Previous findings in my lab suggest that dietary carbohydrate composition influences the weight loss effects induced by Glp1r agonists. Specifically, we observe greater Glp1r agonist-induced weight loss in mice fed a high carbohydrate diet compared to mice fed a calorically matched low carbohydrate diet. Investigating the factors contributing to this effect, multiple groups have observed that Glp1r agonists increase plasma levels of the liver-derived hormone fibroblast growth factor 21 (FGF21). FGF21 levels are increased by the intake of high carbohydrate diets and feed back to suppress subsequent carbohydrate intake. We show that Glp1r agonists increase FGF21 levels by engaging neuronal Glp1r. Furthermore, we show that deleting either liver FGF21 or the obligate FGF21 co-receptor b-klotho (Klb) in neurons impairs Glp1r agonist-induced weight loss only in mice fed a high carbohydrate diet. This suggests that dietary carbohydrate composition plays a crucial role in mediating the effectiveness of Glp1r agonists in promoting weight loss, and FGF21 is a key molecule behind this interaction via crosstalk between the brain and the liver. These data prompt my investigation to determine whether proopiomelanocortin (POMC) neurons mediate the ability of liraglutide to increase FGF21 levels and whether Klb in the ventral medial hypothalamus (VMH) is required for liraglutide-induced FGF21 to reduce body weight in mice fed high carbohydrate diets (Aim 1), and b) how the Glp1r agonist-FGF21 relationship interacts with dietary carbohydrate composition to influence the weight loss effects of Glp1r agonists (Aim 2). In Aim 1, I will test the hypothesis that Glp1r agonists engage Glp1r in POMC neurons to induce liver FGF21, and in turn, I will test the hypothesis that the ventral medial hypothalamus Klb is the target of Glp1r agonist-induced FGF21 to regulate body weight. In Aim 2, I will test the hypothesis that liver FGF21 contributes to enhanced Glp1r agonist-induced weight loss in a manner that not only involves dietary carbohydrate content (low vs. high carbohydrate) but also carbohydrate source (starch vs. sucrose). I will also test whether these effects on weight loss are due to changes in absolute carbohydrate/sucrose intake and/or preference. Findings from these studies will not only provide novel insight into the mechanism of action of Glp1r agonists but will also identify key factors that contribute to the variability in the response to these therapeutics. Upon completion of the experiments proposed in this application will provide me with extensive training in experimental design, data analysis, and interpretation as well as in mastering concepts and methods relevant to neuroendocrinological regulation of metabolic phenotypes and clinical applications. My Sponsor and co- Sponsor are committed to my training and to helping me develop into an academic researcher.

NIA - National Institute on Aging

Summary. Von Willebrand factor (VWF) is a protein produced in endothelial cells and megakaryocytes. It is best known for its roles in hemostasis and thrombosis; however, an emerging literature has recently linked VWF with development of demenAa. In clinical studies, elevated plasma level of VWF and reduced acAvity of its protease (ADAMTS13) were shown to correlate with increased prevalence and risk for demenAa. In addiAon, elevated VWF is found in cerebrospinal fluid (CSF) of paAents with certain subtypes of Alzheimer’s disease, parAcularly those with vascular dysfuncAon. Normally, VWF within blood vessels is restricted to the endothelium and the immediately underlying basal lamina. However, the presence of VWF ‘deeper’ into the vessel wall (i.e. in the smooth muscle layer) is found in certain vascular pathologies and is referred to as “intramural VWF”. One contribuAng source of intramural VWF is thought to derive from the luminal side, resulAng from dysfuncAonal or damaged endothelial cells. But given that CSF bathes the outer surface of the leptomeningeal vasculature, elevated VWF in CSF may provide an addiAonal (perivascular) route by which VWF can reach smooth muscle cells (SMC) within the vascular wall. Our preliminary data demonstrate intramural VWF in cerebral vasculature in condiAons of amyloidosis, including arterioles of paAents with confirmed cerebral amyloid angiopathy (CAA) and in leptomeningeal arteries of Tg2576 mice (mouse model of amyloidosis). To define the funcAonal roles of intramural VWF, we performed experiments to increase and decrease perivascular VWF. We showed that perivascular VWF promotes abnormal vascular morphology and potenAates mechanisms of vascular remodeling, whereas VWF depleAon aZenuates vascular remodeling, specifically by reducing SMC proliferaAon. In separate experimental approaches, we showed that VWF promotes increased SMC migraAon and transcripAonal changes consistent with a more proliferaAve SMC phenotype. We will now test the overall hypothesis that intramural VWF amplifies pathological remodeling of leptomeningeal arteries in condiAons of amyloidosis, thereby contribuAng to cerebrovascular dysfuncAon, the development of CAA, and worsening of brain pathology. Aim 1 will define how VWF modifies cerebral smooth muscle funcAon and phenotype in the absence and presence of amyloidosis. MechanisAc studies will be performed with human brain SMC and validated in human Assues and a mouse model of amyloidosis (Tg2576). Aim 2 will test the hypothesis that intramural VWF impairs cerebrovascular funcAon and accelerates the development of CAA in the leptomeningeal vasculature. Studies will be performed in Tg2576 mice, which develop leptomeningeal CAA. Aim 3 will determine if reducing intramural VWF in Tg2576 mice can preserve or rescue cerebrovascular funcAon in different stages of vascular Ab pathology. CompleAon of these studies will define the role of intramural VWF in potenAaAng amyloid-driven cerebrovascular pathology and determine the potenAal of targeAng these VWF-mediated mechanisms as a means to reduce cerebrovascular dysfuncAon and slow progression of Alzheimer’s Disease and Alzheimer’s Disease Related DemenAas (AD/ADRD).

NIDA - National Institute on Drug Abuse

Project Summary/Abstract African Americans living with HIV experience accelerated neurocognitive aging despite effective antiretroviral therapy (ART). This project, "Contribution of Neuronal Mitochondrial Dysfunction and Microglial HIV Reservoirs to Neurocognitive Aging in African Americans with Nicotine Addiction and Viral Suppression," investigates how persistent microglial HIV reservoirs, exacerbated by nicotine exposure, disrupt neuronal mitochondria and accelerate cognitive decline. Smoking increases oxidative stress and inflammation, compounding mitochondrial dysfunction and neuroinflammation, leading to greater neuronal injury. Microglia sustain these pathological processes despite systemic viral suppression, contributing to HIV-associated neurocognitive disorders (HAND). Our long-term goal is to mitigate HAND by targeting the core drivers of neurocognitive decline—neuronal mitochondrial dysfunction and persistent microglial HIV reservoirs—particularly in African Americans with nicotine addiction. We hypothesize that chronic microglial HIV reservoir activity accelerates neuronal mitochondrial dysfunction and that nicotine amplifies these effects, leading to earlier and more severe neurocognitive deficits. To test this hypothesis, we propose the following specific aims: Aim 1: Quantify mitochondrial DNA (mtDNA) in neuron-derived extracellular vesicles (NEVs) from blood plasma to assess neuronal mitochondrial dysfunction and its link to neurocognitive impairment in African Americans with nicotine addiction and viral suppression. Aim 2: Measure HIV RNA in microglia-derived extracellular vesicles (MEVs) from blood plasma to assess microglial reservoir activity and its contribution to neurocognitive aging in African Americans with nicotine addiction and viral suppression. We will recruit African American participants stratified by HIV status, smoking behavior, age, and sex. NEVs and MEVs will be isolated from blood plasma using neuron- and microglia-specific antibodies. Biomarkers— mtDNA (neuronal mitochondrial dysfunction) and HIV RNA (microglial reservoirs)—will be quantified using droplet digital PCR (ddPCR) for mtDNA and reverse transcription-ddPCR (RT-ddPCR) for HIV RNA. Neurocognitive function will be assessed with standardized tests, and statistical analyses will correlate biomarker levels with demographic variables and cognitive outcomes. Our preliminary findings indicate increased mtDNA in NEVs and detectable HIV RNA in MEVs among African American men with HIV who smoke, supporting the study’s hypothesis. By integrating biomarker assays with neurocognitive testing, this project will elucidate how microglial HIV reservoirs and nicotine disrupt neuronal mitochondrial integrity and contribute to cognitive decline. Findings will inform precision-medicine strategies to reduce HAND-related health disparities in African Americans living with HIV and nicotine addiction.

NIAID - National Institute of Allergy and Infectious Diseases

SUMMARY Staphylococcus aureus is a major human pathogen that is a common cause of hospital and community acquired pneumonia, both of which lead to significant morbidity and mortality. Increasing antibiotic resistance and prevalence of methicillin resistant strains is of critical concern. The long-term goal of our research is to better understand the host-pathogen interaction between S. aureus and the host innate immune system. It is hoped that an improved understanding of this interaction could lead to novel therapies targeting the bacterium or modulating the host to thwart this multidrug resistant pathogen. The objective of this proposal is to understand how type III IFN contributes to the pathogenesis of S. aureus infection. The rationale for this approach is that new targets are needed to either develop antibiotics/inhibitors or vaccines against S. aureus. Studies investigating type III IFN to-date are largely focused on viral infections where activation of signaling is important to control the infection. We have shown that type III IFN signaling contributes to the pathogenesis of acute S. aureus pneumonia. We have demonstrated alterations to the alveolar macrophage population can improve bacterial clearance by manipulating the pathways identified from our RNA-seq analysis. Investigation of both differential transcripts and proteins indicates changes to the epithelial barrier occur when type III IFN signaling is present. We also have data that shows that S. aureus strains can activate type I and III IFN signaling with variable intensity and independent of each pathway. Precise mechanisms for this involvement of type III IFN signaling are still lacking. This contribution is significant as it will provide a deeper understanding on how the type III IFN pathway influences the clearance of S. aureus from the airway. Completion of our objectives will be accomplished by pursuing two specific aims: 1) Determining the role of type III IFN in pathogenesis through studies on MID1 in alveolar macrophages, the role of IL-1β in immunopathology its effect in the airway barrier and 2) defining how S. aureus activates type III IFN, its comparison to type I IFN signaling and the utility in targeting this pathway. The innovation of this research is both conceptual and technical. The concept that the pathway is inhibitory to bacterial clearance through the epithelial response to type III IFN signaling is unique. We will examine both direct effects on airway epithelial cells as well as the effects IFN signaling has on professional phagocytes. It is also thought activation of type I&III IFN is linked, and we provide evidence this is not entirely the case. We will utilize techniques across multiple fields, microbiology, immunology, cell and molecular biology, utilizing a well-tested animal model of pneumonia, in vitro functional and primary cell culture assays and global approaches to understand the host response such as RNA-seq. At the conclusion of these studies, we will have expanded our knowledge on how an innate signaling pathway can impact negatively on acute bacterial infection in the airway, how this pathway can be activated and how it can coordinate multiple cells in response to its activation.

NHLBI - National Heart Lung and Blood Institute

PROJECT SUMMARY/ABSTRACT Hematopoietic stem cells (HSCs) can self-renew and generate all mature blood cells, allowing them to be used for transplantation to treat hematological disorders. However, as many as 5-27% of patients experience poor graft function, or incomplete hematopoietic recovery after transplant, which has only 6% 2-year overall survival. This can be a direct result of defective HSC self-renewal and multilineage differentiation potential. However, the specific pathways that regulate HSC function remain poorly understood. HSCs are mostly quiescent with low metabolic and mitochondrial activity. They are activated into cell cycle and their mitochondrial activity increases following transplant or 5-fluorouracil (5-FU)-induced myeloablation. Prior work in our lab has shown that after replicative stress, HSCs have decreased regenerative ability, dysfunctional mitochondrial dynamics, and decreased expression of metabolic genes such as stearoyl-Coa desaturase 1 (Scd1). SCD1 is an enzyme that catalyzes the formation of monounsaturated fatty acids (MUFAs) in the de novo lipid synthesis pathway. These products are used to form triglycerides and phospholipids that alter cell membrane structure, signaling, and mitochondrial homeostasis. Specifically, lipids are necessary for proper mitochondrial membrane architecture, dynamics, and oxidative metabolism. Our preliminary data demonstrate that pharmacological inhibition of SCD1 in HSCs in vitro significantly decreases their regenerative ability. SCD1 inhibition also leads to changes in mitochondrial network structure. However, the contribution of MUFA synthesis to HSC function in vivo and why HSCs depend on this pathway is unknown. Our central hypothesis is that MUFA synthesis is important for maintaining HSC regenerative function by supporting mitochondrial function, and loss of this pathway after replicative stress causes HSC functional decline. We will address this hypothesis by determining the role of MUFA synthesis in HSC function in vivo (Aim 1) using an inducible SCD1 knockout mouse model. We will then assess HSC regenerative function by serial competitive transplantation and recovery after 5-FU-induced myeloablation. We will also determine the mechanism by which MUFA synthesis maintains HSC function (Aim 2) by assessing mitochondrial function and performing lipidomic and metabolomic analysis in SCD1-deficient conditions. Using oleic acid, a product of the SCD1-catalyzed reaction, to restore MUFAs, we will determine whether this improves HSC regenerative and mitochondrial function after replicative stress. These studies will lead to a global understanding of the importance of MUFA synthesis to HSC function and reveal pathways that can be targeted to improve HSC regenerative functions and clinical outcomes of HSC-based therapies.

NHLBI - National Heart Lung and Blood Institute

Project Summary/Abstract Myocardial infarction (MI) is one of the most prevalent diseases in the United States and worldwide. A characteristic of the post-MI tissue response includes a rapid inflammatory response. The intensity and duration of this inflammatory response are critical in determining extent of death of cardiomyocytes, progression of the immune response towards anti-inflammatory and pro-repair type, and the extent of fibrosis. Hence, inflammation and its timely resolution can affect organism survival, tissue damage and heart function post-injury. Clinically, excessive inflammation has also been shown to be directly correlated to long-term heart failure and mortality after MI. Inflammation-targeting therapies have been tried in the clinical trials, yet we can do better. Following cardiac injury, the inflammatory response includes infiltration of neutrophils and macrophages (Mφ) into the injured tissue. The resolution of this inflammatory response is an active process in which Mφs play a vital role by engulfing dying cells and apoptotic inflammatory immune cells by a process called efferocytosis. This leads to the production of anti-inflammatory cytokines like IL-10, TGF-β, and bioactive lipids like lipoxins. Mφ metabolism is also intertwined with its functional response as inflammatory Mφs prioritize glycolysis while anti-inflammatory Mφs appear to utilize oxidative phosphorylation to a higher degree. The efferocytic Mφ can utilize fatty acids derived from engulfed cells for oxidative metabolism in mitochondria and facilitate repair functions post injury. Peroxisomes are cellular organelles that have important functions in cellular fatty acid sensing and metabolism. These organelles have the unique ability to metabolize very-long-chain fatty acids (VLCFA) in cells. Our preliminary data suggest a spatial localization of peroxisomes surrounding apoptotic cells, as well as induction of peroxisomal β-oxidation and accumulation of VLCFAs in Mφs during efferocytosis. My experiments further imply that VLCFA metabolism regulates cytokine production and polarization of Mφs following efferocytosis. I hypothesize that Mφ peroxisomal VLCFA metabolism regulates the tissue repair response after injury through altering immunometabolic signaling that controls cytokine production, and in turn, the efficiency of organ repair. My hypothesis will be tested through molecular mechanistic studies that identify causal determinants of peroxisomal signaling in Mφs, and consequences on tissue damage, organ function, and inflammation. My studies also have the potential to uncover new targets for the enhancement of the tissue repair.

NIDDK - National Institute of Diabetes and Digestive and Kidney Diseases

Project Summary/Abstract The long-term goal of this proposal is to modernize clinical methods for assessing fluid excess for removal during in-center hemodialysis treatment using newer assessment techniques. The experience of trial-and-error assessment of a “dry weight,” a weight at which a patient can tolerate fluid removal without the development of symptoms of hypovolemia or hypervolemia, a common practice for over 50 years, that may have a lasting effect on the ability to function for nearly 500,000 Americans who depend on hemodialysis for survival. Newer techniques for quantifying excess extracellular volume for removal during hemodialysis do exist. Small studies have suggested the potential utility of multi-frequency bioelectrical impedance spectroscopy (BIS) as a means to improve on clinical dry weight estimates to guide intradialytic ultrafiltration. Moreover, the rate at which fluid is removed during hemodialysis has been limited to not exceed 13ml//kg/hr based on observational studies on mortality. However, this association may differ relative to an individual's body size and may be more of a reflection of persistent volume overload rather than the just the rate of removal itself. The crude imprecise practice of dry weight assessment together with limitations in ultrafiltration rate may contribute to adverse patient experiences including persistent volume overload with peripheral edema and dyspnea on one hand, and intradialytic hypotension, fatigue, and muscle cramping on the other. Dialysis treatment day BIS estimation of excess extracellular fluid to direct ultrafiltration has the potential to improve patient-centered outcomes, the overarching premise of this proposal. We will gather evidence on the potential utility of BIS to improve the patient experience immediately following and during the post dialysis period. Our findings will inform future studies testing whether use of BIS to guide fluid removal during dialysis can improve how patients feel, function, and survive. We propose to compare dry weight estimation by BIS to usual care including estimate discordance on patient centered outcomes (Aim 1). We will also quantify the relative impact of high ultrafiltration rate and achievement (or non-achievement) of BIS-estimated target weight on patient physical function, sleep and quality of life (Aim 2) and examine clinical decision-making and patient perspectives on how UF management influences patients' dialysis experience using qualitative methods (Aim 3) among adults receiving hemodialysis in two metropolitan areas. The National Kidney Foundation's Kidney Disease Outcomes Quality Initiative (KDOQI) and the Kidney Disease Improving Global Outcomes (KDIGO) organizations are actively examining patient centered ways to improve care quality, the proposed work aligns with these efforts.

NIGMS - National Institute of General Medical Sciences

Project Summary/Abstract Phenotype control, an active area of research in network control, is applicable to interacting biomolecular systems and can identify targeted interventions that lead to desired cell phenotypes. Phenotype control distinguishes itself from classical control theory in that (i) its objectives are related to dynamical attractors (e.g., stable states), and (ii) its interventions don’t need to be continuously adjusted based on the state of the system. This type of control is well-suited for guided cell differentiation, inducing cell fate changes, or inducing apoptosis of a target cell population. This research program will further develop two phenotype control methods. Feedback vertex set (FVS) control is based on the interaction network that underlies a biological system, and stable motif (SM) control is based on a dynamic model of the system. The PI has participated in the establishment of both of these methods, and has a track record of collaborative construction of experimentally validated dynamic models of biological systems. This research program will overcome the remaining challenge to the wide implementation of each phenotype control method. The barrier to wide application of FVS control is that in many systems the characterization of the target cell phenotype (e.g., the known state of a few biomarkers) is not sufficient to specify the desired state of all FVS nodes. This barrier will be eliminated by identifying the most parsimonious and sufficiently discerning characterization of each phenotype and extrapolating the existing biological knowledge to achieve this characterization. The bottleneck to the wide application of SM control is the long time needed for the development and verification of dynamic models. Automating the key steps of model construction and refinement, building on the recently developed BOOLean MOdel REfiner (boolmore) tool, will substantially decrease this time. The FVS and SM control methods will be implemented on networks constructed from curated pathway databases, on ensembles of dynamic models generated for each network, as well as on select dynamic models constructed in collaboration with domain experts. Integration of the two methods will identify multiple high-confidence interventions for each system. The predicted interventions will be validated via experiments done by the PI’s collaborators (see letters of Prof. Nobile and Prof. O’Rourke). The outcomes of the next five years will include a publicly shared suite of computational methods for efficient control of cell phenotypes as well as novel control protocols for multiple specific systems.

NIA - National Institute on Aging

ABSTRACT: The accumulation of waste and neurotoxic proteins, such as amyloid-β and tau, is a hallmark of aging and neurodegenerative disorders like Alzheimer’s disease. The glymphatic system plays a critical role in brain-waste clearance. It facilitates the perfusion of cerebrospinal fluid (CSF) into the brain, which interchanges with interstitial fluid to flush out harmful waste, nanoplastics, and neurotoxic proteins. Our recent findings indicate that conditions impeding CSF flow dynamics—such as craniosynostosis and normal aging—disrupt this clearance system, exacerbating amyloid plaque buildup. Importantly, we have also shown that pharmacological activation of Piezo1, a mechanosensitive ion channel, using Yoda1 agonist enhances CSF-mediated waste clearance in both craniosynostosis and aged wild-type mice. However, a major gap remains in our understanding of how waste clearance is regulated at the cellular level. This obscures the cellular mechanism(s) by which Piezo1 acts to control brain waste clearance. Key preliminary data suggest that mechanical forces exerted by CSF flow may activate Piezo1 in brain border macrophages (BBMs). These cells are essential for brain waste clearance as they degrade extracellular matrix (ECM) proteins along arterial basement membranes, enabling proper arterial motion to drive CSF flow. This proposal aims to determine whether Piezo1 functions in BBMs to control brain-waste clearance, leveraging our team’s expertise in mouse genetics, high-resolution multi-photon imaging, and computational modeling. Aim 1 uses a pharmacological approach to investigate whether Yoda1 enhances brain-CSF perfusion by acting on BBMs to help regulate ECM remodeling. Aim 2 uses genetic approaches to directly test whether Piezo1 is required in BBMs to maintain CSF hydrodynamics and waste clearance by facilitating ECM degradation and arterial motion. Aim 3 examines the impact of Piezo1 manipulation in BBMs on amyloid pathology in mouse models of Alzheimer’s disease, including assessing whether stimulating Piezo1 activation can enhance amyloid clearance. By defining the role of Piezo1 in CSF- driven waste clearance, this work will establish a mechanistic framework for preserving glymphatic function across the lifespan. Targeting Piezo1 in BBMs may represent a novel therapeutic strategy to counteract age- related and disease-associated declines in brain-waste clearance, addressing a critical unmet need in Alzheimer’s disease and related dementias. As such, we expect our findings will lay the groundwork for translational efforts to develop new therapies aimed at sustaining cognitive health in aging populations. Our approaches may also be leveraged in the future to investigate how environmental pollutants (e.g., nanoplastic accumulation) may affect waste clearance and BBM functions, leading to cognitive impairment.

NSF

This award supports research that seeks to enable new theory and algorithms for modeling and control of deformable systems with an emphasis on soft robots, thereby advancing the national health, promoting the progress of science, advancing prosperity and welfare, and securing the national defense. Deformable systems are infinite-dimensional systems that can undergo continuous deformations while moving in their configuration space. Control methods for deformable systems are scarce and their applicability is limited to certain types of systems (e.g., rod-like soft robots). In principle, one can design (model-based) controllers for deformable systems by using Finite Element Models (FEMs). Model reduction methods and/or local linearization techniques can reduce the complexity of FEMs but also lead to the design of unreliable controllers. This project looks to solve this challenge by developing a coarse deterministic FEM whereas its state is represented by a multi-modal density model and synthesizing control methods leveraging multi-modal density steering problems and Koopman operator theory. The special property of deformable systems allows them to find many applications in manufacturing (e.g., handling fragile objects during assembly and packaging), healthcare (e.g., soft robotic prosthetics), search and rescue (e.g., reaching locations under the ruins of collapsed buildings), and agriculture (e.g., soft gripping and harvesting of fruits and vegetables), to name but a few. In addition, the combined educational, research and outreach activities in this project will attract talented and motivated students to learn more about deformable systems and work on relevant research problems. This research aims to establish the theoretical foundations for the creation of novel modeling and control methods for deformable (continuum) systems which can strike the right balance between model complexity and controller accuracy/performance depending on the requirements of the application at hand. In this project, the dynamics of a deformable system will be described by a reduced-order coarse deterministic FEM whereas its shape will be represented by multi-modal density models that adhere to the spatial structure of the FEM but their expressiveness is not necessarily limited by the latter. To handle the nonlinearities in the dynamics of our model, Koopman Operator theory plans to be used to characterize linear predictors which globally approximate the original nonlinear dynamics. Subsequently, control algorithms look to be created by solving relevant multi-modal density steering problems. The research will be verified and validated with the aid of specialized simulation tools. This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.

NSF

Efficient memory processes rely on our brain's ability to store similar experiences as distinct memories. This process, performed in part by a structure in the brains of mammals called hippocampus, facilitates memory discrimination and is affected in brain disorders like dementia and post-traumatic stress disorders. As a result, animals and people suffering from these disorders experience difficulties recalling or accurately retrieving memories. Our understanding of how the hippocampus encodes similar experiences as distinct memories, allowing for easy discrimination during recall, is still limited. Recent advancements in neuroscience suggest that coordinated activity between excitatory and inhibitory neurons enables a specific computation known as pattern separation within the hippocampal circuit. This computation transforms similar input patterns into highly dissimilar output patterns and is thought to be the basis for memory discrimination. In the proposed research, the investigators aim to determine the contribution of a specific type of inhibitory neurons called cholecystokinin-expressing (CCK) neurons to the hippocampal circuit’s function. Their long-term goal is to enhance our understanding of critical computations, like pattern separation, performed by this brain region, which are crucial for memory discrimination. As part of this project, the investigators also provide a strong educational and training environment to students and new generations of scientists, and they participate in outreach programs, such as the Big Brother Big Sister event organized by the Reno Museum of Natural History, which they use as a platform to share their research with the general public and students of all ages. The investigators aim to determine the contribution of CCK neurons to hippocampal circuit function, focusing on the plasticity of CCK neurons during the developmental period of juvenile mice. Their previous work demonstrates that environmental enrichment (EE) leads to increased synapses formed by CCK neurons in the dentate gyrus, suggesting their involvement in the maturation of hippocampal function and pattern separation. For the proposed work, they use an EE paradigm in mice and take advantage of viral mediated gene delivery technique combined with optogenetic approaches to manipulate the activity of the CCK neurons. They determine how the remodeling of the inhibitory CCK network adjusts the computation in the DG. In the first Objective of the project, they determine the contribution of CCK neurons to feedback and feedforward inhibitory microcircuits, both of which can impact pattern separation. In the second Objective, they measure the change in pattern separation and filtering performance of the DG induced by the EE using a novel in-vitro electrophysiological protocol. Together their research aims to unravel the mechanisms underlying pattern separation by exploring the involvement of CCK neurons in the hippocampal circuit. By increasing our understanding of these fundamental computations, they will shed light on the processes that support efficient memory functioning and potentially contribute to the development of novel therapeutic approaches for memory-related disorders. This project is jointly funded by the Modulation Program of the Neural Systems Cluster in the BIO Directorate and the Established Program to Stimulate Competitive Research (EPSCoR). This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.

NIAID - National Institute of Allergy and Infectious Diseases

7. PROJECT SUMMARY/ABSTRACT How apicomplexan parasites recognize host cells to invade them remains a mystery. Since these parasites only replicate intracellularly, invading host cells is essential to their survival. Invasion relies on the coordinated secretion of proteins from two distinct types of organelles: micronemes and rhoptries. Rhoptries are exclusively discharged upon host-cell contact; however, the molecular events that trigger this process remain completely unknown. Understanding the factors required for rhoptry discharge can help us uncover the basis of host-cell recognition. Two conserved microneme complexes—the CLAMP and CRMP complexes—have recently been shown by the Lourido and Lebrun labs to be required for rhoptry discharge, making them likely mediators of host-cell recognition. Our preliminary studies have identified N-glycosylation pathways as key host requirements for rhoptry discharge. Loss of host N-glycosylation significantly reduced rhoptry discharge, while buffering any additional effect from knocking down the CLAMP or CRMP complexes. Based on these observations, we hypothesize that the CLAMP and CRMP complexes recognize host cells through N-glycans to mediate rhoptry discharge. This proposal brings together the two teams that discovered these critical complexes to examine how they recognize host N-glycosylation and mediate rhoptry discharge. Aim 1 will determine the glycan specificity of the CLAMP and CRMP complexes through traditional or liquid glycan arrays. Aim 2 will characterize how N-glycosylation alters the various stages of invasion using high-speed live-cell imaging to dissect the multi-stage invasion process and pinpoint defects to rhoptry discharge. Aim 3 will characterize the signaling events triggered by host-cell recognition through phosphoproteomics and proximity-labeling. Taken together the proposed experiments will uncover how apicomplexans respond to host contact to discharge rhoptries and initiate invasion. Beyond its fundamental importance, this information can be used to design inhibitors of host cell entry that would block parasite infection.

NIDDK - National Institute of Diabetes and Digestive and Kidney Diseases

SUMMARY There is an urgent need for new therapies for metabolic disorders, including obesity, which is associated with a significant decrease in life expectancy by 5–10 years and increased mortality resulting from diabetes and cardiovascular diseases. Peptide hormones comprise a class of small (<100 amino acids), low abundance, bioactive molecules involved in regulating physiological processes such as glucose and insulin levels, food intake, and energy expenditure, making them attractive targets for modulation of energy metabolism. However, the functional identification and characterization of small, proteolytically cleaved peptides have traditionally been challenging because of their low abundance and the difficulties in distinguishing bioactive from inactive fragments or degradation products. Recently, my lab discovered a 12-mer secreted non-incretin human peptide called BRP that controls obesity by lowering food intake through the central activation of Fos. This innovative proposal will explore the mechanisms and biology of BRP to validate the receptor target for this promising cleavage product. Based on preliminary data, we hypothesize that BRP is endogenously cleaved from the precursor BRINP2 by the proconvertase PCSK1, and degraded by proteolytic processing. Based on a receptor screen, we also hypothesize that its function is linked to its binding to a GPCR, thereby activating a CREB signaling pathway leading to Fos transcription. Our hypotheses are based on robust preliminary data generated using in vitro models, structure- activity relationships, and in vivo studies. In Aim 1 of this proposal, we will test the hypothesis that BRP is generated by cleavage of a proconvertase and that BRP is regulated by degradation. These studies will be important to understand the endogenous mechanism of regulation. In Aim 2, we determine the causal hypothalamic region and cell type activated by BRP. In Aim 3, we will validate the target receptor candidate and the signaling mechanism for BRP that is responsible for its appetite-suppressing effects. In conclusion, these studies will determine the endogenous regulation and mechanistic action of BRP, which will open up a new pathway of peptide signaling in body weight regulation.

NIDA - National Institute on Drug Abuse

Project Summary Opioids play a frontline role in acute pain management in the clinical setting. Prolonged use of opioids causes multiple adaptations including tolerance—a phenomenon in which increasing opioid dosage is required to maintain drug efficacy. The pharmacological component of opioid tolerance is largely driven on the cellular level and is the result of cellular protein networks terminating the activity of the mu opioid receptor (µOR). A critical step in this regulatory process is membrane trafficking. High efficacy opioids cause the µOR to rapidly traffic from the cell surface to internal cellular structures called endosomes. At endosomes, the µOR is either sorted back to the cell surface for continued activity (resensitization) or sent to the lysosome for destruction (downregulation). A long-standing model is that downregulation can drive pharmacological opioid tolerance through proteolysis of opioid receptors outpacing new receptor synthesis. One challenge in testing this model is that we don’t know the identity of key cellular protein networks regulating mu opioid receptor trafficking at endosomes. Here we leverage our recently developed proteomic and genomic methods for identifying cellular proteins which regulate opioid receptors. In Aim 1 we focus on the trafficking of the µOR at endosomes and, using biochemical and cellular techniques, test the hypothesis that the endosomal Retromer complex rescues mu opioid receptor from lysosomal degradation in a process called recycling. We will use functional genomics to identify potential druggable targets—kinases and ubiquitin-ligases—that regulate µOR through endosomes, and test the related hypothesis that opioid-induced downregulation of the µOR can be blocked by engineered “super-recycling” variants of the receptor. In Aim 2 we test the hypothesis that endosomal recycling and the endosomal Retromer complex controls three different phases of µOR signaling: endosomal signaling, resensitization, and the development of pharmacological opioid tolerance at the cellular level. Together, these studies seek to define a critical regulatory process controlling µOR function and identify cellular proteins which could be targeted to control the development pharmacological opioid tolerance.

NCI - National Cancer Institute

PROJECT SUMMARY In cancer and chronic infection, T cells develop into a dysfunctional state called exhaustion because of persistent antigen stimulation. Exhausted T cells upregulate immune checkpoints, display dysregulated metabolism, and progressively lose effector function and the ability to persist and develop immune memory. T cell exhaustion hinders the clearance of pathogens and malignant cells by the immune system and limits the effectiveness of immunotherapies such as chimeric antigen receptor (CAR) T cell therapy. Therefore, understanding how T cells adapt to chronic antigen receptor signaling is critical for developing more effective immunotherapies. A stem-like CD8 T cell subset has been identified in chronic infections and cancers. Stem-like CD8 T cells mediate long-term immunity by self-renewal and replenishing other CD8 T cell subsets. Stem-like properties in T cells are essential for the efficacy of immunotherapies. We have identified key transcription factors that regulate the differentiation of stem-like CD8 T cells. However, the molecular program underlying the adaptation of stem-like CD8 T cells to chronic antigen receptor signaling is incompletely understood. Our recent findings show that the redox sensing KEAP1-NRF2 pathway is critical for the differentiation of stem-like CD8 T cells and adaptation of CD8 T cells to chronic antigen receptor signaling. In the proposed study, we will determine how the KEAP1-NRF2 pathway regulates the adaptation of CD8 T cells to persistent antigen receptor signaling through preventing TCR hyperactivation and promoting metabolic fitness. Our study will shed important new light on the development of more potent and efficacious immunotherapies for cancers and chronic infections. Furthermore, KEAP1 and NRF2 have been extensively investigated as potential drug targets for chronic diseases. Therefore, our findings hold significant promises for informing interventions that aim at modulating this pathway.

NINDS - National Institute of Neurological Disorders and Stroke

Project Summary/Abstract The tongue mediates behaviors fundamental to survival, such as vocal communication and food manipulation. Cerebellar dysfunction disrupts normal tongue muscle activation patterns, leading to significant clinical pathologies in swallowing and speech that severely impact quality of life and can be fatal. Despite this clinical significance, the neural mechanisms of cerebellar tongue control in non-human primates remain largely unexplored, with no standardized behavioral tasks for measuring lingual motor learning in these models. Pathologically, cerebellar dysfunction is characterized by deficits in Purkinje cells (P-cells), the sole output neurons of the cerebellar cortex. P-cells produce two types of action potential: the simple spike, which is their primary output, and the complex spike, which is triggered by climbing fiber inputs to the P-cells. Complex spikes are often in response to error, and are always followed by complete suppression of the simple spikes. Though we know that P-cells sculpt the computational output of the cerebellar cortex, the way in which they enact population-level control of tongue movements remains mysterious. Here, I propose to establish the marmoset (Callithrix jacchus) as a model for cerebellar tongue control. Marmosets present several key advantages: 1) their long and protrusive tongue is easy to track with computer vision, 2) their cerebellum demonstrates developmental alignment with non-human primates while remaining accessible with recording instruments designed for mice, and 3) their survival depends on harvesting tree sap via skilled licks, suggesting robust cerebellar involvement in control of targeted tongue movements. Our recent work in marmosets has yielded a breakthrough: when a P-cell is briefly but completely suppressed by a complex spike, the result is a small displacement which serves as a proxy for that cell's causal contribution to behavioral output. I hypothesize that grouping P-cells by their downstream effects on licking will reveal population-level computations that the cerebellum performs to aid in control of the tongue. However, testing this hypothesis requires mapping downstream effects of P-cell suppression across cerebellar lingual regions, as it is a prerequisite for functional classification of P-cells and subsequent analysis of population-level dynamics during targeted licking. Here, my aim is to first develop an experimental setup that will allow for a wide range of errors during licking, thus driving complex spikes. Next, I will record from various cerebellar lingual regions and use spike-triggered averaging to measure the change in tongue kinematics caused by the complex spike-induced suppression of each P-cell. To test specificity of the effects due to suppression, and not error, I will consider movements in which there was no error, but a suppression occurred. This will test whether downstream effects of P-cells are regionally organized in the cerebellum. Overall, my proposed work will bridge the gap in our understanding of cerebellar control over tongue movements and will aid lingual neuroscience by developing a new non-human primate model in marmosets.

NIAID - National Institute of Allergy and Infectious Diseases

Controlled Delivery of Adjuvanted Multivalent Fusion Peptide Focused HIV Vaccines Project Summary The development of a safe and effective vaccine against HIV-1 continues to be a global health priority and has posed a formidable scientific challenge. Recent advances in vaccinology and immunology highlight great promise in enhancing potency of vaccine-induced immunity using prolonged delivery of vaccine immunogens and adjuvants. Striking increases in the induction and persistence of germinal centers produced by such regimens in rodents and non-human primates highlight the enormous benefit in affinity maturation of vaccine-specific B cells, which is critical for HIV-1 targeted vaccines. Notably, we have recently found that the SOSIP+3M-052 combination induced antigen-specific long-lived plasma cells that successfully trafficked to the bone marrow in non-human primates (NHPs) and were found to be stable for 2-3.5 years in independent studies. We seek to capitalize on these insights with five mutually reinforcing components: (i) safe and injectable hydrogels, based on unique chemistry of tunable bond formation and disconnection, that provide uniquely tunable extended release properties; (ii) safe and highly immunogenic virus-like particle platforms for multivalent antigen display; (iii) platform-switching heterologous immunization to focus the immune response on the HIV-1 fusion peptide; (iv) state-of-the-art SOSIP-based envelope glycoprotein (Env) immunogens; and (v) a nanoparticle-based form of the TLR7/8 targeted adjuvant 3M-052. These tools will be used to generate vaccine-induced durable and protective HIV-1 specific immunity, and to study responses at the cellular, molecular, and antibody-epitope levels in NHPs to maximize the changes of clinical success. Our three specific aims blend innovative chemistry, immunogen display, and delivery (Finn lab) with optimization of vaccination strategies coupled with deep immunological and vaccinological analyses in rodents and NHPs (Kasturi Lab). Upon successful completion of the program, we will have validated a modular class of vaccine formulations and identified key determinants of vaccine-induced durable and protective humoral immunity. This work will enable rapid future optimization of candidate vaccines against HIV-1, and potentially other pathogens, to elicit broad and long-lasting protection.

NSF

With support from the Chemical Mechanism, Function, and Properties (CMFP) program in the Division of Chemistry, Professors Andrew Musser and Phillip Milner of Cornell University are using ultrafast laser techniques to study how molecular coherence can be controlled. Coherence can arise when the electronic or vibrational states of two or more molecules synchronize, and it plays in central role in how light interacts with matter, from the physics of vision to photosynthesis to new systems for generating electricity from light. Proper control of such coherences could lead to devices that more efficiently harvest and transport energy, or to advanced applications of quantum mechanics in sensing and information science. And yet, despite its ubiquity, very little is known about how to tune such coherences or identify their unique contribution to photochemical processes. In this project, the teams of Musser and Milner will tackle this question in the context of singlet fission, a process in which one photoexcited state splits in two, with major implications for technologies such as quantum information science and light harvesting devices. Professor Milner and his students will prepare systematic libraries of crystalline sponge-like materials known as metal-organic frameworks – effectively molecular tinker toys whose modularity lets the team dial in specific interactions between photoactive molecules. Professor Musser and his students will use cutting-edge ultrafast laser-based measurements to watch the molecules in these frameworks share energy and move in real time and identify conditions where they achieve coherent synchronization. These studies will lead to new design principles to control and steer coherent singlet fission dynamics for next-generation devices. The team will additionally train multiple graduate and undergraduate students in interdisciplinary research spanning organic chemistry to spectroscopy, and they will develop middle-school outreach programs on how light interacts with matter. Recent advances in spectroscopic techniques have made it ever easier to identify the presence of coherence in a host of processes driven by light-matter interactions, and it is associated with ultrafast, high-efficiency transfer and conversion dynamics. But it has proved more challenging to identify the unique impact of coherence itself: the general conditions that enable superpositions of quantum states (e.g. energetic resonance) also favor efficient incoherent processes. By studying coherent dynamics in well-defined solids where the interactions are tuned with molecular-level precision, the team aims to establish structural guidelines to harness and tune intermolecular coherence. The team will probe this behavior in the context of singlet fission, in which an excited singlet separates into two low-energy triplets. The initial ultrafast conversion into an intermediate triplet-pair state is often described in terms of vibronic coherence, while the product triplets exhibit persistent spin coherence. Both types of coherence present potential control knobs in terms of vibrational environment, orbital overlap, and intermolecular transport pathways. By incorporating fission-active molecules (such as monomeric or dimeric pentacenes, rylene diimides) into modular crystalline frameworks with varied structure, the team will systematically tune these parameters. The project will use ultrafast electronic spectroscopy to track coherent vibronic wavepackets through the initial fission process and monitor variation in long-time spin coherence through magnetic-field effects. Comparing these measurements across the structural library, the team will develop empirical guidelines to enhance intermolecular coherence. The insights generated will enable the design of more effective singlet fission materials for optoelectronic devices. This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.

NSF

This research project aims to advance fundamental understanding of the deformation behavior of amorphous solids with many-body interactions among constituent particles. Unlike crystalline materials, amorphous solids such as metallic glasses and granular systems exhibit complex, non-crystalline structures and deformation mechanisms that are not captured by conventional models based on binary interactions. In many-body systems, the interaction between two particles can be influenced by the presence of a third — a phenomenon common in biological tissues and social networks but not yet well understood in the context of amorphous solids. This project seeks to extract governing physics from simulations and experiments of amorphous systems with many-body interactions and develop new solid mechanics models to predict their behavior. The outcomes seek to to impact a wide range of applications, including adaptive metamaterials, two-dimensional materials, and damage-tolerant structural materials. Educational activities include hands-on demonstrations, curriculum integration, and public outreach aimed at fostering future scientists and engineers. The technical objective is to quantify how many-body interactions influence deformation through the lens of energy landscape complexity. Atomistic simulations and machine learning–assisted analyses will be used to study emergent features in the energy landscape while systematically tuning many-body interactions. These findings look to be validated through two granular experimental systems with fluid-mediated interactions arising from air-fluidization and capillarity. Realistic interactions will be modeled using graph neural networks and integrated into atomistic simulations. A key focus is to uncover how many-body effects alter the synchrony between particle-level and system-level energy responses during deformation. The results will inform a meso-scale elastoplastic finite element model capable of capturing strain localization and shear band formation. Ultimately, the project aims to enable the design of advanced amorphous materials by engineering the morphology of their energy landscapes via tailored particle interactions. This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.

NINDS - National Institute of Neurological Disorders and Stroke

PROJECT SUMMARY/ABSTRACT Temporal lobe epilepsy (TLE) is a complex neurological disorder that is characterized by spontaneous, reoccurring seizures and often presents with debilitating cognitive comorbidities including learning and memory deficits. Current treatments for TLE are ineffective at managing cognitive comorbidities and often exacerbate these symptoms. The mechanisms underlying cognitive impairments in TLE are poorly understood, but learning and memory deficits are thought to arise when TLE pathologies disrupt the normal function of hippocampal circuits that support these cognitive processes. Circuits in the hippocampal dentate gyrus (DG) are known to support learning and memory and have been shown to be disrupted in TLE patients and in animal models of TLE. Recently, Dr. Shuman showed that the spike timing of DG interneurons is disrupted in TLE. Additionally, he showed that restoring proper spike timing of DG interneurons with closed-loop optogenetic stimulation decreased seizure susceptibility in TLE mice. Because proper spike timing in the hippocampus is an important mechanism that supports learning and memory computations, I hypothesize that DG interneuron spike timing is also important for behavior. In aim 1, I will test this hypothesis by using closed- loop optogenetic stimulation to alter the spike timing of DG interneurons relative to ongoing theta oscillations. I predict that disrupting DG interneuron spike timing in non-epileptic mice will impair learning and memory while restoring the disrupted spike timing of DG interneurons in TLE mice will restore learning and memory deficits. While optogenetic approaches are great for assessing the causal relationships between physiological processes and behavior, there are a number of barriers that limit the translational potential of optogenetics. In aim 2, I will employ a novel circuit-editing tool to increase connectivity between DG interneurons and their upstream inputs in the medial entorhinal cortex (MEC) with the goal of increasing feedforward inhibition in the DG. I hypothesize that increasing connectivity to DG interneurons will decrease seizure susceptibility. Additionally, I predict that this approach will restore the proper spike timing of DG interneurons because inputs fire at the same preferred phase of theta as DG interneurons and are not disrupted in TLE. Circuit-editing is a novel technique that has never been applied to epilepsy. Successful implementation of this approach could result in a transformative gene therapy for epilepsy patients and will inform how feedforward inhibition in the DG normally supports hippocampal physiology. The following research project and training plan were designed to help me grow both scientifically and professionally, and support from this grant will be invaluable as I continue to work towards my goal of becoming an independent researcher. The outstanding opportunities provided by the Shuman lab and Mount Sinai provide the ideal training environment for me as I pursue my postdoctoral fellowship and ultimately a career in academic research.

NSF

This award supports a joint effort between the University of Nebraska-Lincoln and the University of Texas at Austin to study the underlying physics of how a plasma responds to energy inputs. The ultimate goal of the project is to develop techniques to actively control the evolution of the plasma state and energy flow. The control of plasma dynamics is central to a wide range of applications. Knowledge gained in this project is expected to help realize the technological promise of many plasma-based systems, including laser-plasma accelerators with applications ranging from high-energy physics, astrophysics, and nuclear science to medicine, biology, and chemistry. This work will also have an educational component that will serve to help renew the plasma science and engineering workforce at all levels by providing training in both the analytical and numerical treatment of plasma dynamics. Plasma-based systems are the subject of much investigation as energy converters. In these systems, electromagnetic radiation impinges on the plasma, driving a complex internal state which is generally allowed to evolve freely, ultimately forming beams of particle and or radiation. In this project, linear theory will be used to instantaneously predict the short time evolution of the full nonlinear system and this may be used to apply external fields to steer the evolution on a desirable path to obtain a sought after state. Controlling the dynamics in plasma-based systems, beyond simply suppressing instabilities, could open the door to releasing significant amounts of energy on queue. Such control would also allow the plasma to be engineered to enable energy conversion from electromagnetic frequencies that are easily produced to those parts of the spectrum that currently do not have ready sources. This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.

NIDDK - National Institute of Diabetes and Digestive and Kidney Diseases

Title: Controlling NETosis to Treat Diabetic Wounds. Project Summary This Phase 1 STTR proposal is focused on establishing proof-of-concept studies aimed at developing a novel humanized monoclonal antibody for the treatment of diabetic wounds. NETosis is a unique form of cell death characterized by neutrophils releasing net-like structures known as "neutrophil extracellular traps (NETs)" to combat bacterial pathogens. Initially lauded for its role in innate immunity, emerging evidence suggests the adverse effects of excessive NETosis in both infectious and non-infectious diseases. Diabetic individuals manifest a notable elevation in NETs-releasing products in their bloodstream, and uncontrolled NETosis has been linked to impaired wound healing in diabetes and increased infection susceptibility. Citrullination of histone H3 (CitH3) plays a pivotal role in initiating NETs-induced immune cell death and tissue damage. Elevated CitH3 has been identified as a risk factor for wound healing impairment and amputation in patients with diabetic foot ulcer. To translate our basic findings on CitH3 into practical applications for humans, we have recently developed a humanized anti-CitH3 monoclonal antibody (hCitH3-mAb). We have completed developability studies and established a protocol for scaled-up production and purification of hCitH3-mAb, demonstrating superior binding capabilities to CitH3 surpassing commercially available antibodies. In vivo studies have shown that hCitH3-mAb improves survival in septic shock mouse models and enhances wound healing in diabetic mice. Our proposal aims to formulate a topical delivery system for hCitH3-mAb, enabling sustained release for the treatment of diabetic ulcers. Following this, we will conduct proof-of-concept studies utilizing hCitH3-mAb to treat diabetic wounds in animal models. The successful completion of these studies brings us one step closer to submitting an IND application to the FDA, potentially revolutionizing the treatment of diabetic ulcers.

NSF

With the support of the Chemical Mechanism, Function, and Properties Program of the Division of Chemistry, Professor Liviu M. Mirica of the Department of Chemistry at the University of Illinois Urbana-Champaign aims to develop new odd-electron reaction processes for palladium metal, which normally favors even-electron processes. The proposed research will lead to better fundamental understanding of the electronic properties and reactivity profiles of the uncommon paramagnetic (odd-electron) palladium systems, which impacts how chemists build complex molecules more efficiently. The proposed studies comprise synthetic inorganic and organometallic chemistry and physical inorganic spectroscopy, thus providing interdisciplinary training and the opportunities students to develop a broad range of scientific skills. Palladium (Pd) complexes play an important role organometallic catalysis of a wide range of synthetically useful organic transformations such as C-H functionalization, C-C coupling, and C-heteroatom bond formation reactions. The vast majority of these catalytic processes formally involve Pd(0)/Pd(II) oxidation states, while the Pd(IV) oxidation state has been invoked more recently in several chemical transformations. By contrast, the chemistry of odd-electron Pd(I) and Pd(III) oxidation states is less appreciated and understood. Herein, the PI aims to develop (1) a fundamental understanding of the electronic properties and reactivity profiles of paramagnetic Pd(I) and Pd(III) systems, and (2) methods to control one- vs. two-electron processes at these Pd centers. Spectroscopic techniques including EPR, low temperature UV-vis, low temperature electrochemistry, X-ray crystallography, solution X-ray absorption spectroscopy, and computational studies will be applied to study these uncommon paramagnetic Pd systems. The specific activities include: (1) perform ligand-enabled C-H activation/functionalization studies; (2) investigate reactivity of photogenerated Pd(I) species; and (3) perform mechanistic studies of reactions at Pd(I) and Pd(III) centers. This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.