Stanford University

NIH Research Projects · FY 2025 · 2023-04

Project Summary/Abstract The mechano-electrical transduction (MET) process allows the transduction of mechanical information from sound into electrical signals, and it is a fundamental step in cochlear system function. Failures in this process lead to hearing loss and deafness. Understanding the basic properties of MET will lead to a better understanding of deafness, leading to targeted treatments and therapies. MET takes place at the level of the hair bundle and is mediated by tip links, extracellular proteins connecting shorter stereocilia to adjacent taller stereocilia. Deflections of the hair bundle towards the tallest stereocilia row increase tip-link tension and open MET channels that reside at the top of the shorter stereocilia. Although there is a large body of work regarding lipid membrane modulation of mechanosensitive ion channels, there is a limited but growing body of data on lipid modulation of cochlear hair cell MET. The lipid environment can affect channels indirectly through changes in membrane mechanics, or directly through individual lipid/protein interactions. PIP2, an endogenous phospholipid, modulates MET channel properties, potentially through a direct interaction or indirectly by altering membrane mechanics. A stretch activated channel modifier, GsMTx4 reduces the resting open probability (Po) of MET channel while also blocking the increase in Po induced by lowering external calcium or depolarizing the hair cell, suggesting the lipid membrane may be involved in modulating MET channel Po. The effect of voltage and calcium could be mediated through changes in lipid packing due to multivalent ions interacting between adjacent lipids. Our recent direct assessment of membrane diffusivity of individual stereocilium at a time using two-photon Fluorescent Recovery after Photobleaching (FRAP) demonstrated that stereocilia membrane is sensitive to calcium and voltage but not the soma, and MET channel Po co-varies with membrane diffusivity, supporting the hypothesis that the MET channel can be modulated by membrane mechanics. However, due to spatial and temporal limitations of FRAP, we were unable to monitor stereocilia membrane locally and dynamically. To further test this hypothesis and overcome current technological limitations, I will combine electrophysiology with live-cell fluorescence lifetime imaging (FLIM) of a novel viscosity sensor to examine the membrane viscosity with improved spatio-temporal resolution for the first time in mammalian cochlea. I will assess local and temporal changes in the stereociliary membrane viscosity with voltage, calcium, and membrane components like cholesterol and PIP2 and correlate these effects to changes in MET channel Po. These studies will enhance our basic understanding of the importance of lipid membrane in hair cell mechanotransduction. Understanding the crucial components in the mechanical underpinnings of the stereocilia are both biophysically and biologically relevant. The development and use of these new technologies will greatly advance my career as an independent investigator and likely have broader applications in the auditory field and beyond.

NIH Research Projects · FY 2025 · 2023-03

Project Summary/Abstract Positron emission tomography (PET) is a method of medical imaging that employs positron emitting radionuclides attached to probe molecules (tracers) for non-invasively interrogating biological processes in vivo. Each radionuclide decay emits a positron, which then combines with an electron and creates two oppositely directed, colinear 511 keV annihilation photons. These annihilation photons are detected in opposing elements of a photon detector ring forming lines of response (LOR) along which each positron emission originated. After collecting millions of such photon pair events and positioning them along system LORs, an image can be reconstructed to visualize and quantify the 3D distribution of tracer probe molecules within the body. Up to now, PET systems detect only one tracer per study. However, more complete understanding of the disease biology often requires the study of multiple biological processes simultaneously. Alzheimer’s Disease specifically is characterized by presence of neuroinflammation, β-amyloid, phosphor-τ, and neurodegeneration. This project aims to enable simultaneous imaging of multiple tracers by strategically choosing at least one PET tracer that emits gamma photons in cascade with their positron. These gamma photons can be differentiated from annihilation photons through their higher energy measured by the photon detector. Thus, LORs can be associated with this positron + gamma tracer when a high energy photon arrives nearly the same time as a pair of 511 keV photons. Challenges associated with using prompt gamma emitters come from the lower probability of detecting both the annihilation photons and gamma photon within the appropriate timing and energy windows. Another challenge associated with using multiple tracers is misclassification of events among tracers due to missed, tissue scattered, or random photons detected. Low detection efficiency and misclassification will reduce image quality and accuracy of the associated multi-tracer images; thus, methods must be developed to mitigate these issues. This project proposes to develop and characterize a position-sensitive endcap detector that will cover the open end of an existing PET ring system to increase the detection efficiency for 3-photon events through increasing the solid angle coverage of the system photon detectors. Signal processing algorithms will also be employed using multiple temporal and energy windows to mitigate misclassification of photons coming from the two emitters in addition to compensating for sensitivity differences between two- and three-photon emitters. These techniques will include use of delayed time windows to estimate the different random coincidence rates and joint maximum likelihood estimation of coincidence events based on system geometry. We will also use deep learning to improve the image quality of the low dose images and to accurately separate the images obtained. Through these techniques, this project aims to develop the first system capable of simultaneous multi-tracer PET imaging.

NIH Research Projects · FY 2026 · 2023-03

Project Summary/Abstract Non-alcoholic steatohepatitis (NASH) is a major cause of HCC. It is clinically recognized that HCC in NASH often arises at a pre-cirrhotic stage, however the pathomechanism of HCC in non-cirrhotic livers is not well understood. As T2DM with poor glycemic control is an independent risk factor for HCC it is plausible that there are distinct pathways that create a pro- carcinogenic niche in non-cirrhotic T2DM/NASH. We showed that the accumulation of advanced glycation end products (AGEs) in patients with T2DM/NASH and in an animal model are key to necroinflammation and oxidative liver injury. Downregulation of the AGE clearance receptor AGER1 accelerated AGE deposition, and correcting AGER1 in vivo improved NASH. To study how high AGE environment creates permissive conditions for transformed cells, we modulated diet/AGE content prior to hydrodynamic injection of hMET/mutant β-catenin: 1) High AGE background induced an earlier and more invasive HCC, 2) AGE accumulation was linked to significant changes in matrix dynamics-with an increase in energy dissipation or loss and faster stress relaxation in response to a deformation - in an AGE and receptor for AGEs (RAGE)-dependent manner, 3) Inhibiting AGE production reversed changes in matrix viscoelasticity in vivo and lowered tumor burden. We will test the hypothesis that in non- cirrhotic T2DM/NASH accumulation of AGEs contribute to an increase in matrix viscoelasticity and matricellular changes creating a pro-invasive environment. Aim 1: We propose to investigate the link between diet/AGE content, matrix viscoelasticity, sex and HCC phenotypes and outcomes in a novel HCC model. In particular we will focus on how AGE-mediated collagen crosslinks on specific amino acids can alter intermolecular recognition and interaction with proteoglycans thereby affecting matrix dynamics. Aim 2: We will dissect the effects of RAGE and AGER1 signals and AGE accumulation on matrix viscoelastic parameters and HCC phenotypes. In Aim 3 we propose to develop a 3D hydrogel system with tunable viscoelasticity and the impact of AGEs-modified matrix on cell behavior will be studied. Based on the RNAseq data we will focus on the pathomechanism of how viscoelastic changes are sensed by cells, and the key matricellular signals that confer invasive and migratory properties. These studies will demonstrate the impact of AGEs on the liver matrix in non-cirrhotic NASH and outline the pathomechanism for a pro-invasive environment. Defining the key matricellular signals that drive invasion will enable us to pursue translational studied in the future.

NIH Research Projects · FY 2026 · 2023-03

Abstract Kidney cancer is increasing in prevalence and is one of the top 10 most common cancers world-wide. Clear cell renal cell carcinoma (ccRCC) is the most common and aggressive type of kidney cancer. While primary ccRCC is treated with surgery, 30% of patients are diagnosed with regionally advanced or metastatic disease that requires systemic therapy. Despite current treatments that target the tumor microenvironment, the 5-year survival rate for advanced ccRCC remains 11%. HIF-2 plays an important oncogenic role in ccRCC, which has led to the recent development of the HIF-2 inhibitor belzutifan. However, innate and acquired resistance limits durable responses in the majority of patients. Thus, there is a need to identify additional therapeutic targets that directly inhibit the growth and survival of ccRCC cancer cells. The von Hippel Lindau (VHL) tumor suppressor gene is lost in ccRCC tumors associated with the VHL disease and in 90% of sporadic ccRCC tumors. Hallmark features of VHL deficient ccRCC include constitutive activation of hypoxic (HIF) signaling, angiogenesis and metabolic reprogramming. We recently discovered a synthetic lethal interaction between the RNA demethylase FTO and VHL in ccRCC. Importantly, FTO knockdown reduces ccRCC growth and survival independent of HIF-2. Yet, actionable mechanistic insights into 1) how FTO promotes VHL deficient ccRCC growth and survival and 2) the therapeutic potential of FTO-based therapy in ccRCC are critical gaps in knowledge addressed in this application. Our central hypothesis is that the m6A RNA demethylase, fat-mass and obesity-associated protein (FTO), promotes glutamine reprogramming to support the growth of belzutifan sensitive and resistant ccRCC. Our specific aims will test the following hypotheses: (Aim1) FTO promotes VHL deficient ccRCC glutamine reprogramming by increasing the expression of the glutamine transporter SLC1A5 via m6A demethylation; (Aim 2) FTO inhibitors create a metabolic vulnerability in VHL deficient ccRCC that can be exploited therapeutically to treat belzutifan sensitive and resistant tumors. Upon conclusion, we will understand the role of m6A RNA methylation and FTO as epitranscriptomic regulators of glutamine reprogramming in ccRCC. This contribution is significant since it will establish that SLC1A5, an actionable and prognostic cancer therapy target, is regulated by FTO through m6A modifications. Additionally, insight into the mechanisms of FTO- mediated growth, survival and glutamine reprogramming is important for the rapid translation of precision medicine approaches as FTO inhibitors are currently in clinical development.

NIH Research Projects · FY 2026 · 2023-03

PROJECT SUMMARY 3,4-Methylenedioxymethamphetamine (MDMA) a Phase 3 clinical trials as an adjunct to psychotherapy for Post-Traumatic Stress Disorder (PTSD). Published data show that MDMA therapy has a rapid onset and a ability to foster feelings of social connection, empathy and trust. However, MDMA itself may not be an ideal therapeutic, as it has a well-known potential for abuse and is associated with cardiovascular and neuro- psychiatric toxicity. Despite these and other limitations, the apparent efficacy of MDMA suggests that directly enhancing sociability and social reward sensitivity are feasible, potentially powerful therapeutic strategies. In mouse models we can use MDMA as a unique probe to understand evolutionarily conserved social behaviors with potential therapeutic relevance. Conventional approaches to understanding the mechanism of MDMA and other psychiatric drugs, focusing on high affinity receptor interactions and select brain areas, have had limited success at developing novel therapeutics for psychiatric disease. My lab has developed a way to define with few assumptions about pharmacology or brain areas involved. This method, in mice, maps brain-wide activity evoked du for MDMA-linked behaviors. Combining social behavioral testing and imaging could be used to screen novel therapeutics for MDMA-like profiles and provides testable hypotheses for human imaging studies with MDMA. MDMA releases supraphysiological levels of serotonin (5-HT) and dopamine (DA) among other neuromodulators and evokes acute social preference, social reward learning, and nonsocial drug reward in humans and mice. Mechanistically similar drugs that primarily release 5-HT (d-fenfluramine, FEN) or DA (d- methamphetamine, METH) recapitulate selective components of the total MDMA effect, but neither induces social reward learning. Here, we propose to take advantage of the overlapping yet distinct behavioral and unique prosocial effects. First, we compare brain-wide Fos expression maps between groups of mice under drug/environmental conditions that on brain-wide neural activity -like behavioral effects. Second, we test whether activity in identified regions is required for expression of four MDMA-evoked behaviors: acute social preference, drug craving, social reward craving and social operant conditioning. My preliminary data demonstrates proof-of-concept: we have discovered that the dorsal endopiriform nucleus/ ventral claustrum (DEn/VC) has an obligate role in MDMA-evoked acute social preference. Third, we detail the anatomy and connectivity of the DEn/VC, and test whether its activity can suffice to drive prosocial behaviors.

NIH Research Projects · FY 2025 · 2023-03

Dr. Sutha’s long-term career goal is to be a physician-scientist who improves the lives of his patients through basic science and translational research directed at developing novel therapies for kidney disease, including acute kidney injury and allograft rejection. A detailed understanding of both acute and chronic kidney disease pathogenesis and advancement of targeted therapies to treat them remains limited. Development of novel therapeutics has been hampered by lack of holistic, in vitro experimental models that fully preserve the complex interactions between epithelial, stromal, and immune components of the kidney. In this work, Ken Sutha, MD, PhD, a pediatric nephrology physician-scientist, builds upon his expertise establishing a patient-derived, adult kidney organoid culture preserving endogenous stroma and immune cells, in addition to his primary mentor Dr. Calvin Kuo’s experience with organoids from other tissues for studying immune processes and diseases. First, using his established adult kidney organoids as a platform, Aim 1 will delineate interactions occurring between kidney epithelia and accompanying stroma in the setting of nephrotoxic injury induced by cisplatin. These studies will yield insights into epithelial and fibroblast activation and interactions after injury and will further identify protective versus maladaptive signals in the progression of acute kidney injury to chronic kidney disease. This knowledge will enable the development of novel strategies to disrupt fibrosis while promoting repair, which will subsequently be further tested within adult kidney organoids. Aim 2 will then translate the use of his adult kidney organoids to kidney transplant, using organoids to model the allograft immune microenvironment, critically while preserving host immune cells within allograft tissue. These organoids will then be applied to model transplant rejection and treatment response, including functional assessments of immune activation. Initial feasibility studies will serve as proof of principle for future prospective studies using transplant organoid treatment response to inform patient therapeutic decisions. Dr. Sutha will benefit from the interdisciplinary environment at Stanford University, along with resources and support available to him through the Department of Pediatrics. With mentorship from Drs. Kuo, Vivek Bhalla, and Jonathan Maltzman, Dr. Sutha will expand upon his expertise in nephrology and adult kidney organoid models by developing new, complementary skills in bioinformatic analysis and transplant immunology, critical to successful advancement of his project. This training will enable the novel application of his established kidney organoid model to study kidney injury and the transplant immune microenvironment, eventually enabling him to develop and test new, targeted kidney disease therapies. This approach will be further applied to model other kidney diseases and treatments using biopsy samples, and results generated from the completion of his K08 Aims will serve as preliminary data for Dr. Sutha’s future R01 investigating cellular interactions and crosstalk in the progression of chronic kidney disease.

NIH Research Projects · FY 2026 · 2023-03

The retinal pigment epithelium (RPE) nourishes and promotes survival of photoreceptors. The RPE contains abundant mitochondria, consistent with a metabolically active tissue with a variety of energy intensive tasks. Our characterization of the retinal phenotype of mice with postnatal RPE-selective ablation of Tfam (RPEΔTfam) demonstrates the necessity of RPE mitochondrial function for the integrity of this epithelium, and for the well being of photoreceptors. RPE-selective knockout of Tfam results in RPE-cell autonomous and non- cell autonomous effects including a progressive loss of photoreceptor function and numbers. Our findings complement studies implicating the RPE as the site of ocular pathology in individuals with inherited mitochondrial defects, and support a causal role for for RPE mitochondrial dysfunction in age-related macular degeneration (AMD). Our preliminary studies have uncovered a signaling pathway that is quiescent in normal RPE cells and induced by diverse stressors; striking upregulation of FGF21 in the RPE of RPEΔTfam mice and dispersion to the neural retina implicates this secreted molecule as a critical signal capable of propagating the negative effects of RPE mitochondrial distress. We propose to test this hypothesis and understand the mechanisms by which FGF21 affects the stressed mouse retina. In Aim 1, we will use mouse models to determine the consequences of loss and gain of FGF21 function on retinal phenotype in the context of RPE mitochondrial dysfunction. In Aim 2, we will determine the FGF21 autocrine and paracrine contributions to the retinal phenotype in this context, including cellular transcriptional responses. In Aim 3, we will develop molecular inhibitors of FGF21 and test their efficacy in mouse models of RPE distress. Given the centrality of RPE mitochondrial function to retinal homeostasis and the relevance of chronic stress responses to human diseases, including AMD, a mechanistic understanding of the consequences of this RPE-derived mitochondrial distress signal could have a substantial long term impact from both basic and applied perspectives.

NIH Research Projects · FY 2026 · 2023-03

PROJECT SUMMARY/ABSTRACT The opioid crisis remains a major health concern with millions of Americans addicted to opioid drugs and thousands of opioid related deaths per year. The sad reality is that while medication assisted therapies are highly effective, relapse rates remain high. More understanding is needed into the neurobiology and circuits of opioid addiction to identify new therapies. Opioids exert their rewarding and addictive effects through action at the mu opioid receptor (MOR). The MOR is expressed widely throughout the nervous system including regions associated with drug reward such as the nucleus accumbens. It is present in a peculiar neuroanatomic organization referred to as “patch” or “striosome,” with dense regional expression situated in a network of islands throughout dorsal striatum and nucleus accumbens. The region outside of these islands is referred to as matrix. The functional relevance of this level of neuroanatomic organization is mysterious and its consequence for opioid use disorders is almost completely unknown. While the direct and indirect pathway of striatal organization has revealed critical insights into motivated behavior and pathologic changes associated with substance use disorders, it remains incomplete especially in regions of the ventral striatum such as the nucleus accumbens. The neuroanatomy of “patch” vs “matrix,” and the cell types contained within each compartment, opens the possibility for a revived lens through which to look at the functional organization of the nucleus accumbens in motivated behavior and addiction. Recently, the power of mouse genetics revealed two separate populations of direct pathway medium spiny neurons housed within MOR positive patch networks. Further work has shown that while one population encodes positive valence and positive reinforcement, the other encodes negative valence and negative reinforcement, challenging the traditional dogma of the direct pathway. This proposal resubmission begins to define a role for these cell populations in preclinical models of opioid abuse, investigating the properties of patches in the valence of opioids and withdrawal, opioid consumption, maintenance, extinction and reinstatement. Input and output circuitry will be defined in each patch cell type within the nucleus accumbens. The work will combine opioid self-administration, behavioral economic analysis, viral neuroanatomic techniques, optogenetic and chemogenetic manipulations and cell type neuroimaging with fiber photometry. This work will be among the first to study MOR (+) patch circuits in the context of opioid use disorder. Through this new lens of functional organization, insights can be revealed that could lead to new therapies in treating the devastating health and societal impact of opioid use disorders.

NIH Research Projects · FY 2026 · 2023-03

Project Summary/Abstract The goal of our research is to understand the visual processing performed by the human retina. In the proposed work, we will explore the striking and novel visual properties of numerous new cell types we have recently discovered in the human retina – including more than 10 retinal ganglion cell (RGC) types that send visual information to the brain, and more than 10 polyaxonal amacrine cell (PAC) types that modulate RGC activity over large areas. We will develop new methods to understand their role in natural vision. We will leverage our large, unique database of recordings from macaque monkeys, the animal model closest to the human, to direct our experiments with precious human tissue. This work is made possible by several technical advances: novel large-scale high-density recordings from human donor retinas, novel machine-learning methods for mining of similar large-scale recordings from macaque monkey retinas performed over the last 15 years, and novel analytical methods to identify, model and compare new cell types and their function in the two species. Preliminary results in both species reveal a striking variety of new RGC and PAC types, some of which are likely homologs, and others that may be unique to humans. We will pursue two aims: (1) identify many novel RGC and PAC types in the macaque retina, probe their unusual light response properties, and develop models that allow us to further probe their role in natural vision, and (2) identify and characterize many RGC and PAC types in the human retina, and then use novel large-scale analysis methods to identify homologous types in the macaque retina. At the conclusion of this work, we will have identified and thoroughly characterized many of the most important visual pathways at the output of the human retina, developed and tested models of their function in natural vision, quantitatively compared to homologs in the most important animal model, and identified retinal cell types and visual signals that are likely unique to humans.

NIH Research Projects · FY 2026 · 2023-03

ABSTRACT Gastric cancer is a major global disease burden and leading cause of cancer mortality worldwide. Current treatment decision is made primarily on the basis of staging, which divides patients into several prognostic groups. For patients with localized and locally advanced disease, curative-intent surgery with chemotherapy is the standard treatment. However, survival outcomes vary widely, even among patients with disease of the same stage. Certain patients with early-stage disease have a sufficiently low risk of recurrence and may not benefit from, or could even be harmed by, chemotherapy given the associated toxicity and side effects. Conversely, many patients with aggressive tumors do not respond well to standard chemotherapy and still recur despite receiving extensive but ineffective treatment. Therefore, current one-size-fits-all approach is suboptimal, leading to over- and under-treatment in many patients. There is an unmet need for reliable prognostic and predictive models to guide personalized treatment of gastric cancer. To address this unmet need, we propose robust radiomics features of tumor morphology and spatial heterogeneity and establish their prognostic value. In addition, we will incorporate pathobiological knowledge into the design of deep learning models for predicting prognosis. Further, we will develop novel deep learning architecture to analyze longitudinal images for predicting pathologic response to neoadjuvant therapy. Finally, by leveraging the complementary value of imaging data, clinicopathologic variables and serial serum markers, we will construct integrative models to further improve prediction. If successful, the proposed models will be useful in two ways: (1), identify which patients with early gastric cancer may safely forego chemotherapy and avoid toxicity; (2), select the most effective chemotherapy regimen for a given patient. Further, the models can also identify patients with advanced disease who do not respond to standard chemotherapy and may benefit from novel targeted therapy or immunotherapy. The proposed computational imaging approaches are generally applicable for response monitoring and disease surveillance in many solid tumor types. Finally, the AI-based imaging technology developed here can bring benefit to underserved populations in minority groups and community settings. Progress made in gastric cancer will not only improve outcomes for patients in the US but also have global impact given its high incidence and mortality worldwide.

NIH Research Projects · FY 2025 · 2023-02

PROJECT SUMMARY: New strategies to targeted difficult-to-drug diseases such as cancers, neurodegeneration, and genetic disorders are urgently needed. RNA manipulation is an emerging, therapeutically attractive paradigm which allows target intervention orthogonal to drugging proteins and without the permanence of gene editing. A variety of tools for RNA manipulation have been developed, but they rely on ribonuclear proteins, which are difficult to deliver in vivo or are limited in scope of effect. This proposal aims to develop RNA-based bifunctional molecules (RBMs) which will consist of an RNA oligonucleotide liked to a small molecule which recruits an effector protein, and will enable modular, programmable targeting of RNA with a variety of manipulations. The proposed mechanism for RBMs is based on small interfering RNA (siRNA) oligonucleotides which are widely used as research tools and have resulted in multiple approved therapeutics. In cells, siRNAs are loaded into Argonaute (AGO) proteins which are part of the RNA silencing complex (RISC). AGO then guides RISC to mRNA targets complementary to its loaded siRNA guide, which are cleaved upon binding, resulting in translational silencing. The oligonucleotide portion of RBMs will function much like siRNAs but will feature key mismatches with the target RNA. Such mismatches have been shown to oblate the cleavage activity of RISC while maintaining target binding. Target binding will induce proximity between the target RNA and an effector protein recruited by the small molecule ligand of the RBM, allowing the effector to act on the target. I will synthesize a small library of RBMs with variable linker lengths and positions, and verify that they can interact with AGO and a model effector protein in vitro (Aim 1). Next, I will use AGO pulldown to show that these interactions can be recapitulated in cells, then use two model systems to show that RBMs can enable post-transcriptional control of mRNA targets (Aim 2). Finally, I will show that RBMs targeting nuclear, long non-coding RNAs can enable control of gene expression (Aim 3). Overall, this will create a platform in which the siRNA paradigm is expanded to enable a much wider variety of manipulations, which will enable novel research tools and therapies. This project will use my existing skills in synthetic chemistry as a foundation and then allow me to branch out in the field of chemical biology. The laboratory of my sponsor, Prof. Steven Banik, is a supportive research environment which will enable me to successfully learn the new skills required to execute this proposal. Prof. Banik is a member of Stanford's Chemistry Engineering and Medicine for Human Health (ChEM-H), a highly collaborative and interdisciplinary institute. Stanford and ChEM-H will afford me all necessary research resources, a variety of opportunities for professional development, and the opportunity to work with, and learn from many, different scientists.

NIH Research Projects · FY 2026 · 2023-02

Project abstract Maintaining the junctional epithelium (JE) is of primary importance if preservation of the cementum, periodontal ligament (PDL), and alveolar bone is to be achieved. New insights into JE barrier functions came with our discovery of a Wnt- responsive stem cell niche in the JE. In this proposal, our goal is to characterize the cellular players, niche signals, and regulatory mechanisms that control and maintain the JE stem cell niche in health, and after damage or disease. AIM 1 experiments will first test whether Wnt/β-catenin signaling is necessary for JE maintenance. Pathway inhibition will be achieved in Axin2LacZ/+ mice using adenovirus expressing the soluble Wnt inhibitor Dkk1; controls will receive adenovirus encoding the Fc portion of immunoglobulin G. At defined intervals, quantitative analyses will assess endogenous Wnt/β- catenin signaling via Xgal staining; JE hemidesmosomal gene and protein distribution via quantitative immunohistochemistry (qIHC); JE and GE cell cycle kinetics by EdU/BrdU labeling; and inflammatory cell infiltration in connective tissues underlying the JE and GE by FACS. Second, whether Wnt/β-catenin signaling is necessary for JE regeneration will be determined by subjecting Wnt lineage tracer e.g., Axin2CreERT2/+;R26RmTmG/+ mice to partial gingivectomy followed by Ad-Dkk1/Ad-Fc delivery. Lineage-tracing and quantitative analyses will establish a relationship between Wnt- responsive cell progeny, cell cycle kinetics, hemidesmosomal gene and protein distribution, and regeneration of JE barrier functions. AIM 2 experiments will evaluate the ability of a stabilized formulation of WNT protein to regenerate a functional JE. In one injury-repair model the JE will be surgically excised; in a second model, JE breakdown will be triggered via a ligature-induced periodontitis; both will be carried out in Axin2LacZ/+ and Axin2CreERT2/+;R26RmTmG/+ mice. Delivery of the WNT therapeutic will be followed at multiple timepoints by quantitative analyses to assess re-establishment of JE barrier functions. AIM 3 experiments will characterize Wnt-responsive JE stem cells and their progeny. Wnt-responsive stem cell pools from adjacent gingival epithelium (GE) will serve as control. Axin2CreERT2/+;R26RmTmG/+ mice will be exposed to tamoxifen, followed by harvest of JE and GE tissues at defined timepoints. GFP+ cells will be sorted by flow cytometry. Gene expression profiling of GFP+ cells will focus stem cell and differentiation markers. Fluorescent in situ hybridization will confirm gene expression patterns using RNA probe libraries corresponding to stem cell markers, components of Wnt/β-catenin, Notch, and Bone Morphogenetic Protein (BMP) pathways. Collectively, this proposal promises to provide important new insights into the requirement for Wnt/β-catenin signaling in maintaining the JE stem cell niche; regulating JE and GE cell proliferation and differentiation; and influencing hemidesmosomal-mediated attachment to the tooth surface. In addition, it should resolve the current debate over the molecular identities of JE v. GE stem cell pools and their differentiation potential. The proposed work also has the potential to identify an innovative therapeutic strategy for rebuilding a damaged JE and thus open new avenues for the restoration of the soft tissue attachment following periodontal diseases.

NIH Research Projects · FY 2025 · 2023-02

Project Summary While a growing body of work has contributed to our understanding of synapse assembly, less is known about how synapses are maintained throughout life. This long life of synapses is crucial for the sustained function of neural circuits, including those supporting cognition, movement, and other vital functions. However, maintaining long-lived synaptic connections presents a cell biological challenge, as synaptic proteins have finite lifetimes, synaptic vesicles turnover rapidly, and protein synthesis is scarce in the presynaptic compartment. This project will study the mechanisms of presynaptic maintenance, using C. elegans as a model system. In Aim 1, I will investigate the proteins involved in maintaining synaptic structures, using the auxin-inducible degron system to remove the candidate proteins SYD-2, SYD-1, SAD-1, CDK-5, and PCT-1 from the mature nervous system. Changes in synapse organization will be assessed using endogenous, cell-type specific markers of synaptic vesicles and active zone proteins. In Aim 2, I will identify regulators of the presynaptic scaffolding protein SYD-2. I will implement a visual forward genetic screen to identify candidates that regulate SYD-2 stability, using a pulse-chase SYD-2 HaloTag approach to visualize SYD-2 turnover. In Aim 3, I will identify regulators of SYD-2 through the use of Split-TurboID proximity biotinylation to detect interacting partners of SYD-2 in the presynaptic compartment. Candidate regulators of SYD-2 turnover will be assessed using the SYD-2 HaloTag system. This work will identify key synaptic maintenance proteins and their regulators. Understanding the mechanisms that maintain stable synapses provides therapeutic avenues for preserving synapses in aging and neurodegenerative diseases. This project, performed in the laboratory of Dr. Kang Shen at Stanford University, provides a strong training opportunity for me in the fields of cell biology and neuroscience, and I will gain new experience with the C. elegans model system, genetic manipulations, and microscopy.

NIH Research Projects · FY 2026 · 2023-02

Metabolic mechanisms of cognitive decline in aging and AD mediated by inflammatory PGE2 signaling Project Summary Aging is characterized by the development of maladaptive immune responses that promote cognitive decline and Alzheimer’s disease (AD). We recently identified the inflammatory lipid messenger prostaglandin E2 (PGE2), signaling through its EP2 receptor, as a major driver of age-associated inflammation and cognitive decline. Genetic deletion of the EP2 receptor in myeloid cells was sufficient to prevent systemic and brain inflammation and cognitive decline in aging mice. Myeloid EP2 deletion rescued healthy immune cell responses by restoring glucose flux and downstream mitochondrial respiration in aging macrophages and microglia. We also made the surprising observation that peripheral inhibition of EP2 signaling with a non-brain penetrant EP2 antagonist phenocopied the effect of pan-myeloid EP2 genetic deletion. These data suggest that peripheral inhibition of pro-inflammatory PGE2 signaling is sufficient to restore healthy hippocampal function in aging mice. In this proposal, we will build on these initial findings and define how metabolically reprogrammed myeloid cells in the periphery can elicit effects beyond the blood-brain barrier (BBB) that reverse changes in hippocampal function in models of aging and AD pathology. We will test the hypothesis that the beneficial immune-metabolic effects of EP2 inhibition on myeloid cells in the periphery are transmitted from the blood to the cerebral endothelium and then to astrocytes, leading to improved astrocytic support of neurons. We will employ preclinical models of aging and mutant APP lines, targeted metabolomics and transcriptomics to understand how improving peripheral myeloid energy metabolism leads to beneficial effects beyond the blood brain barrier. We will test whether peripheral EP2 immune blockade, by reprogramming circulating blood, will improve endothelial function. We will then test whether astrocytes, whose foot processes envelop the capillary bed are in turn functionally improved. As astrocytes support neuronal metabolism, we hypothesize that peripheral EP2 inhibition will improve astrocytic support of neurons, leading to improved cognitive function in models of aging and AD.

NIH Research Projects · FY 2026 · 2023-02

Abstract Deuterium metabolic imaging (DMI) is an emerging MRI technique whereby deuterated substrates and their metabolic products are imaged in vivo. A primary application is the study of energy metabolism, a fundamental process for virtually all cells in the body. In particular, glucose (Glc) metabolism plays a critical role in cancer, with two key metrics of tumor metabolism being total glucose consumption and the relative fraction of Glc undergoing glycolysis (GLY) versus oxidative phosphorylation (OXPHOS). In contrast to normal tissues, most cancers exhibit a preponderance of GLY over OXPHOS. Known as the Warburg effect or, more generally metabolic reprogramming, these alterations are particularly pronounced in glioma and other brain tumors. Elevated GLY in high-grade brain tumors has been shown to be a marker of tumor growth and aggressiveness. From a therapeutic perspective, studies strongly support that this Warburg phenotype is necessary and sufficient for the cancer process, which provides the framework of a highly novel therapeutic strategy targeted at affecting these metabolic pathways. We contend that clinical translation is presently impeded not so much by a lack of agents, but by the difficulty in measuring these fundamental aspects of tumor metabolism in vivo. Of the available imaging techniques, 18F-FDG-PET is well-established for imaging glucose uptake, whereas robust in vivo measurements of GLY and OXPHOS are considerably more challenging. Triple 15O-PET can be used to assess oxygen consumption (and hence OXPHOS) but is clinically problematic due to the 2-min half- life of 15O and the challenges of coordinating multiple inhaled radioactive gases. MRI of hyperpolarized 13C- labeled pyruvate has been shown capable of assessing tumor GLY/OXPHOS ratios; however, this technique is very expensive with limited availability and unique challenges. More recently, the feasibility of using conventional 2H MRSI of deuterated glucose to measure both GLY and OXPHOS has been successfully demonstrated. Given the ubiquity of 3T scanners, we contend that 3T DMI would have maximal clinical impact, and initial results for the human brain reported at 4T, in combination with our own 3T DMI data, indicate limited spatial resolution, low SNR, and correspondingly long scan times are the primary limitations. This technical development project will address these challenges by enhancing DMI via the incorporation of multimodal information. Noting that 1H MRI and FDG-PET share significant mutual anatomic and metabolic information with DMI, we propose to significantly enhance 3T DMI using signal processing and machine learning approaches analogous to techniques using MRI to enhance FDG-PET resolution and SNR. The overall goal is to demonstrate enhanced DMI acquisitions and image processing pipelines for maximal clinical impact, with the initial application being the imaging of the Warburg effect in brain tumors.

NIH Research Projects · FY 2025 · 2023-02

PROJECT SUMMARY The goals of this study are to target the neurobiology underlying restricted and repetitive behaviors (RRB) in children with autism spectrum disorder (ASD) using N-acetylcysteine (NAC), a well-tolerated nutritional supplement and glutamatergic modulator that has exhibited efficacy for reducing RRB severity in recent preliminary trials. The goals of this career developmental award are to learn the theoretical principles and develop practical application techniques for proton spectroscopy (1H MRS) and electroencephalography (EEG) approaches in children with neurodevelopmental disorders, learn clinical trial methodologies and develop skills for examining treatment efficacy in controlled trials, and learn advanced statistical modeling techniques for assessing complex treatment-related outcomes in pediatric populations. The ultimate overarching goal is to support a promising early career investigator in transitioning to an independent research position. To achieve these goals, we will (Aim 1) acquire 1H MRS and EEG data from children with ASD who exhibit severe RRB and examine the ability of NAC to modulate excitatory signaling in cortico-striatal circuits (CSC) in a single dose challenge study (NAC and placebo). We will also (Aim 2) examine the efficacy of NAC for improving RRB in a 12-week randomized controlled trial and (Aim 3) assess the ability of neurobiological measures of excitatory signaling (1H MRS and EEG) to predict treatment response. CSC are a salient treatment target because CSC contribute to RRB in mouse models of ASD and exhibit relationships with RRB severity in children with ASD. Altered excitatory signaling in CSC regions have also been reported in children/adults with ASD, and most importantly, NAC can modulate glutamatergic signaling in CSC regions. Thus, excitatory (i.e. glutamatergic) signaling in CSC in ASD may contribute to the severity of RRB, at least for some individuals, and modulation with NAC may confer some clinical benefits, especially for children with elevated levels at baseline. We expect that children with ASD who receive NAC will exhibit a larger reduction in 1H MRS and EEG measures of glutamatergic signaling compared to children who receive placebo, which will be associated with a larger reduction in RRB severity following 12 weeks of treatment with NAC compared to placebo. We expect that baseline measures will also be able to accurately predict which children respond to NAC and will explore the effects of NAC on different subtypes of RRB, which may be due to different neurobiological alterations. The findings from this research will support the efficacy of NAC for the treatment of RRB and shed light on the mechanisms of action underlying NAC-mediated improvements. This is particularly important because there are currently no drug treatments for the core symptoms of ASD, including RRB, and severe RRB are associated with management challenges and barriers to adaptive learning. With this training, I plan to develop a programmatic line of research to identify the neurobiology of specific symptoms in children with ASD and develop objective biological markers that can be used to improve treatment-related research and help advance precision medicine.

NIH Research Projects · FY 2026 · 2023-02

Development of AI-Augmented quality assurance tools for radiation therapy Project summary Quality assurance (QA) is an essential part of radiation therapy (RT) workflow and critically determines the success of patient care. However, current treatment plan QA methods and tools are deficient in multiple aspects and suffer from problems such as limited accountability, labor intensive, and costly. In this project we will leverage the emerging deep learning techniques to create clinically translatable solutions for robust and efficient QA of modern RT. Specifically, we aim to (i) establish a novel framework for using deep learning to verify the machine delivery parameters of an RT treatment plan; (ii) investigate the use of deep learning for RT dose verification; and (iii) evaluate the performance of the QA system and show its potential clinical impact . This research presents the first-of-its-kind treatment plan QA strategy capable of providing both machine delivery parameters (MLC apertures and MUs of the involved IMRT/VMAT fields) and dosimetric distribution on the patient’s treatment geometry. The research will also make it possible to take advantage of the useful features of both deep learning models from Aims 1 and 2 and check the cycle-consistency of a treatment plan (i.e., from the beam parameters of the plan to the corresponding 3D dose distribution, and then from the 3D dose to the beam parameters) for enhanced plan QA. Successful completion of the project will provide urgently needed plan QA tools for safe, efficient and high-quality RT practice, and enable patients to truly benefit from modern RT modalities. Finally, the proposed strategy is quite broad and can be readily generalized for QA of other treatment modalities, such as proton therapy and high-dose rate (HDR) brachytherapy.

NIH Research Projects · FY 2026 · 2023-01

Aneuploidy is a ubiquitous but poorly-understood feature of tumor genomes. For instance, approximately 25% of human cancers harbor extra copies of the “q” arm of chromosome 1, making this amplification more common across cancer types than mutations in KRAS, PIK3CA, RB1, and many other widely-studied cancer driver genes. Despite the prevalence of 1q aneuploidy in cancer, we have little insight into its role in tumorigenesis. While evolutionary studies have defined consistent patterns in which single nucleotide substitutions occur in oncogenes and tumor suppressors during cancer development, the relative timing of most copy number alterations remains unknown. Additionally, while multiple approaches have been developed to experimentally manipulate single genes in cancer, our ability to alter and study chromosome-scale dosage changes is extremely limited. Thus, we lack genetic strategies that would allow us to develop a mechanistic understanding of how aneuploidies like Chr1q gains influence cancer biology. We hypothesize that certain commonly observed aneuploidies like Chr1q-amplifications may function as cancer “addictions”, in the same way that some cancers can be addicted to oncogenes like KRAS and PIK3CA. Eliminating these aneuploidy “addictions” could therefore block cancer growth and suppress various malignant phenotypes. To investigate this hypothesis, and to uncover the role of 1q-gains in cancer biology more broadly, we have developed novel computational and functional approaches to study cancer aneuploidy. In preliminary work, we discovered that Chr1q gains are commonly the first arm-scale copy number change that occurs during tumor development, and we found that genetically eliminating Chr1q aneuploidy prevents malignant growth in human cancers. To build on these findings, in Aim 1, we will optimize and apply a strategy to reconstruct the evolutionary timing of somatic copy number alterations from multi-sample sequencing studies of human tumors. In Aim 2, we will apply a novel chromosome-engineering approach to eliminate Chr1q aneuploidy from human cancers, and then we will characterize how aneuploidy-loss affects various malignant phenotypes. In Aim 3, we will identify the dosage-sensitive driver genes encoded on Chr1q that contribute to this “aneuploidy addiction” phenotype. In total, these aims will shed light on the functional consequences of an enigmatic genomic alteration found in many cancers. As 1q gains commonly arise during malignant growth but are extremely rare in normal tissue, a greater understanding of this aneuploidy could point toward treatment strategies that are effective against a wide range of tumors but that have little effect on normal diploid tissue.

NIH Research Projects · FY 2026 · 2023-01

Project Summary The spatial organization of cells and molecules in biological tissues plays a critical role in pathophysiology. For example, spatial heterogeneity in the tumor microenvironment determines tumor initiation, metastasis, and drug response. Despite advances in spatial transcriptomics to map RNA in tissues, it is proteins, rather than RNA, that drive most cellular processes and determine disease state. As protein abundance cannot be inferred precisely from transcriptomic data, it is important to measure protein abundance and their spatial distribution to better predict pathophysiological phenomena, as well as to identify biomarkers and therapeutic targets. Previous work on spatial proteomics is based on antibody recognition, mass spectrometry imaging, or physical dissection of the tissue followed by liquid chromatography-tandem mass spectrometry (LC-MS/MS). Antibody-based and mass spectrometry imaging approaches have low proteome coverage (<100 proteins). The only approach with deep coverage (>3000 proteins) is tissue dissection followed by LC-MS/MS. This method leverages the power of state-of-the-art LC-MS/MS to achieve in-depth quantification of thousands of proteins along with their post-translational modifications. However, this approach is limited by current dissection methods. Manual dissection has low throughput and poor spatial resolution. Laser capture microdissection (LCM) has high resolution, but the isolation of many pixels, required for tissue mapping, is tedious and suffers from sample loss. The goal of this project is to develop a high throughput and scalable technology to perform tissue microdissection that preserves tissue spatial information and couples directly to established LC-MS/MS workflow for deep and unbiased spatial mapping of the proteome. We will demonstrate our technology on tumor slices of cutaneous squamous cell carcinoma. Our approach integrates a novel tissue micro-dicing device (“µDicer”), a nanodroplet sample preparation platform (“nanoPOTS”) for LC-MS/MS analysis with single-cell sensitivity, and a microfluidic device (“µMapper”) to transfer the diced tissue pixels from the µDicer to the nanoPOTS array while preserving their spatial order. Our approach is innovative because no technology currently exists to perform tissue micro-dissection and their transfer to macroscopic wells in parallel for LC-MS/MS while preserving spatial information. The specific aims are to optimize the µDicer for dicing fixed tissue slices into 10-100 µm micro-tissue pixels, develop and validate the µMapper to transfer tissue pixels from the µDicer onto nanoPOTS chips, and develop a high throughput and integrated spatial proteomics workflow and apply it to map human tumor slices. The project is significant because it will accelerate mass spectrometry-based spatial proteomics, thereby advancing our understanding of the role of tissue heterogeneity in pathophysiology, such as the role of the tumor microenvironment on cancer progression, and will enable the identification of novel protein biomarkers and therapeutic targets to facilitate the early detection, diagnosis, and intervention of diseases.

NIH Research Projects · FY 2026 · 2023-01

PROJECT SUMMARY/ABSTRACT Infection with Epstein-Barr Virus (EBV) has been epidemiologically demonstrated to be a pre-requisite for developing multiple sclerosis (MS), as essentially 100% of MS patients become infected with EBV prior to MS onset. We recently identified molecular mimicry between the EBV transcription factor EBNA1 and the glial cellular adhesion molecule GlialCAM in 20 - 25% of MS patients, and this is likely a critical mechanism underlying the development of MS in this subset of patients. This R01 will take our investigation of the role of EBV in mediating the pathogenesis of MS to the next level, through: (i) analysis of dysregulated transcriptional pathways in EBV- transformed B cells derived from MS patients, (ii) investigation of the ability of EBV-transformed B cells to mediate autoreactive T cell activation, and (iii) characterization of molecular mimicry-related EBV and CNS autoantibodies in MS. We will test the hypothesis that in MS, EBV activates B cells to circumvent B cell tolerance, and thereby drive autoantibody production, autoreactive T cell activation, and MS pathology. We further hypothesize that different reactivities to self-antigens, to EBV, and to molecular mimics are pathognomonic for distinct MS disease subtypes with different HLA/genetic backgrounds and disease characteristics. Aim 1 will analyze dysregulated transcriptional pathways in EBV+ B cells derived from MS vs. controls. Aim 2 will express MS EBV+ B cell-encoded monoclonal antibodies and identify their EBV and CNS targets. Aim 3 will investigate the ability of EBV-transformed B cells to mediate autoreactive T cell activation. Aim 4 will perform serologic analysis to characterize molecular mimicry-related EBV and CNS autoantibodies in sera from large cohorts of MS and comparator patients. Success of the proposed studies would elucidate the mechanisms by which EBV causes MS, which would transform our understanding of MS and could lead to fundamental therapies.

NIH Research Projects · FY 2026 · 2023-01

Abstract This project addresses the current lack of quantifiable and clinically relevant imaging endpoints for use in patient-derived organoid models. Microphysiological tumor models (μPTMs) are tissue-engineered 3D tumors that can be grown inside microfluidic devices to form multicellular tissue-like constructs that retain the biological and functional characteristics of the tissue of origin. These μPTMs provide a powerful model of individual patients’ tumor and are used in drug discovery, cancer research, and personalized medicine. However, a critical hurdle remains that, unlike xenotransplanted tumors, μPTMs are incompatible with positron emission tomography (PET) and other diagnostic imaging tools used in oncology. There is a dearth of quantitative imaging methods that can be applied seamlessly across physical scales, ranging from in vitro cell cultures to animal models and cancer patients. Drawing from extensive preliminary work, we will bridge this gap by harnessing the ability of radioluminescence microscopy (RLM) to image clinical radionuclides in organoids with ultra-high spatial resolution. Upon completion, this project will enable routine imaging of in vitro tumor models using the growing array of diagnostic and therapeutic radiopharmaceuticals, many of which are used as clinical standard of care. This goal will be achieved by pursuing three specific aims. First, we will demonstrate that quantitative image-based metrics can be acquired using PET tracers in patient-derived organoids. Validation will be conducted for three PET tracers against mouse xenograft models derived from the same set of cancer patients. Second, we will refine the μPTMs by incorporating functional (perfusable) human microvascular networks within the 3D matrix and, using imaging and other assays, determine the effect of the vasculature on image-based endpoints. Third, as a pilot translational study, we will develop patient-specific μPTMs (n=10) and compare fluorodeoxyglucose (FDG) metabolic activity in these organoids against biomarkers derived from clinical FDG-PET. In sum, this project will enhance the ability of researchers to run clinical trials “on a chip”, using the patient’s own tumor and clinically approved radiopharmaceuticals. Ultimately, these advances could be translated to predict the efficacy of new drugs, test biological hypotheses, and individualize patient therapy.

NIH Research Projects · FY 2026 · 2023-01

PROJECT SUMMARY Ventricular arrhythmias remain the leading cause of death in patients with cardiomyopathy and account for up to 300,000 deaths per year in the United States. However, the current classifications of these rhythms is based largely on whether the cardiomyopathy is due to obstructed coronary arteries and poorly stratifies patient response to therapy, arrhythmic risk, and pathophysiology. The goal of this project is to develop an actionable classification scheme for ventricular arrhythmias in patients with cardiomyopathy that is based on the interplay between both structural and electrical abnormalities measured from each patient’s heart. Such a classification, based in measurements of pathophysiology, would inform the clinical approach to risk assessment, interventional therapies, and medications. The proposal outlines three Specific Aims: 1) To identify electrical fingerprints of endocardial, mid-myocardial and epicardial scar using machine learning of endocardial high-density contact electrograms trained to the ground truth of regional delayed gadolinium fibrosis on magnetic resonance imaging, from our large patient registry. 2) To develop and validate a mapping strategy that could be used at clinical electrophysiology to measure ventricular refractory period, a measure of electrical remodeling that indicates ability to sustain VA, by machine learning of high density electrical signals from the heart of a porcine model labeled by repolarization indices from the gold standard, simultaneously recorded, monophasic action potentials. And 3) To derive novel phenotypes of arrhythmogenic cardiomyopathy in patients with VA based on regional distributions of fibrosis and electrical remodeling, and associate these with acute response to ablation and recurrence in a well- characterized patient registry. To successfully complete the proposed project, training objectives include 1) advanced MRI processing and segmentation, 2) machine learning models for multimodal data analysis, 3) translational interventional procedures, and 4) translational clinical electrophysiology. The proposed NHLBI K23 award will provide protected time for the candidate to obtain this advanced training, to disseminate new knowledge via written and spoken communication, and to build the foundation for an independent research program focused on ventricular arrhythmia diagnosis, prevention, and therapy in a supportive environment of established mentorship, collaborators, and interdisciplinary experts spanning engineering and medicine.

NIH Research Projects · FY 2026 · 2023-01

Cardiovascular (CV) diseases are rising causes of morbidity and mortality worldwide. There is excitement that computational medicine, an emerging field combining engineering disciplines with the life sciences, will enable scientific and clinical breakthroughs and accelerate bench-to-bedside translation. However, few training programs exist in this field, and so trainees often learn ad hoc. We seek funding for a new multidisciplinary T32 program in Computational medicine in the Heart: Integrated training Program (CHIP) at Stanford. CHIP will provide cutting-edge training for 3 post-PhD, -MD or -MD/PhD fellows annually, each undergoing 2 years of training at the intersection of engineering, CV physiology and medicine. Trainees will pursue a cutting-edge research project mentored by faculty with complementary expertise in engineering and the life sciences, and select didactic courses to build expertise, grow professionally, and develop community. The forward-looking vision of CHIP addresses key priorities of several National Agencies and fills current gaps in interdisciplinary training. Stanford CHIP leverages faculty and resources at top-ranked Schools of Engineering, Medicine and Humanities and Sciences. The T32 is co-directed by a physician-engineer and an engineer-physiologist, bringing 38 faculty from 13 Departments and Divisions. Key support is provided by the inter-disciplinary Cardiovascular Institute (CVI) and the Institute for Computational and Mathematical Engineering (ICME) at Stanford. Faculty will provide trainees with research opportunities in CV science spanning cell-to-organ and bench-to-bedside, as well as computational science, clinical care, and therapeutic innovation. The faculty are highly collaborative and have exceptional track records of launching trainees into independent scientific careers. Trainees will not be required to have backgrounds in both engineering and life sciences. The T32 will provide tailored teaching of CV science to engineers, engineering to life-science fellows, and advanced cross-disciplinary topics to each. Core didactics also include ethics, the responsible conduct of science, methods to ensure reproducibility. Evaluation will be both constructive and bidirectional between trainees and faculty. In summary, the CHIP T32 at Stanford provides post-MD, post-PhD and post-MD/PhD graduates with world-class training at the intersection of bioengineering, CV science and medicine. The T32 is well positioned for success due to the co-location of these top-tier resources on a single campus in Silicon Valley, a hub of innovation in data science, artificial intelligence and therapy. Our aspirational goal is that CHIP graduates will become global scientific leaders at the cusp of engineering, physiology and medicine.

NIH Research Projects · FY 2026 · 2023-01

Myelin—the structure that encapsulates axons—is integral to efficient transduction of electrical signals and metabolic support of neurons. Myelin deficits have been commonly identified in a wide range of brain disorders—from neurodevelopmental to neurodegenerative—implicating dysmyelination as a prominent, but often underappreciated, feature of many neurological disorders. Similar myelin deficiencies cause decrements in attention, memory, learning, social behaviors, and motor function in preclinical models. To ultimately understand how myelination fails in these brain disorders, we first must have a comprehensive understanding of the mechanisms driving the proliferation and maintenance of myelin-forming precursor cells. Developmental myelination during pre- and post-natal life and neuronal activity-dependent adaptive myelination in adulthood both depend on oligodendrocyte precursor cells (OPCs) and their progeny, myelin-forming oligodendrocytes. There remains an unmet need to define the mechanisms mediating OPC dynamics during these two types of myelination and to reveal their roles in defining and refining circuits and behavior in development and disease. The OPC is the most abundantly mitotic cell in the brain with 70- 90% of all dividing cells at a given time being OPCs. Based on published work showing that the circadian (~24 hour) system—driven by the principal circadian molecular regulator Bmal1—regulates cell proliferation of numerous neural precursor and stem cell populations and our preliminary data confirming the necessity of Bmal1 in OPC dynamics, we aim to investigate how the circadian system regulates OPCs and consequent myelination. In addition to dysmyelination, circadian phase-shifts and polymorphisms in circadian clock genes like BMAL1 have been documented in individuals with autism, attention deficit/hyperactivity disorder, multiple sclerosis, and Alzheimer’s disease, linking disruptions in circadian machinery with pathophysiology in these disorders. We posit that circadian disruption of myelin-forming cells during development will lay the foundation for a broad range of brain pathologies. In this proposal, we aim to elevate the current biological understanding of myelination. The proposed work will investigate how the circadian system regulates myelination through 1) the development of two distinct but complimentary genetic mouse models targeting Bmal1 knock down in OPCs to probe developmental myelination 2) application of the environmental chronodisruptive chronic jet lag (CJL) model to developmental myelination, and 3) diurnal changes in neuronal activity-induced adaptive myelination. Our preliminary data establish that genetic knock down of the Bmal1-driven circadian clock in OPCs during embryonic and post-natal development results in a reduction in 1) OPC density, 2) OPC proliferation, 3) myelination, and 4) myelin-associated behaviors. A comprehensive understanding of the interplay between circadian modulation and OPC maintenance and myelination will not only inform on mechanisms of brain health but will also establish insights into potential therapeutic strategies targeting myelin-specific circadian regulatory processes in numerous brain disorders.

NIH Research Projects · FY 2026 · 2023-01

Project Summary Ribosomes are macromolecular machines that decode the genome. Quality control mechanisms that ensure the fidelity of this process are thus of paramount importance cellular and organismal viability. Ribosomes move at variable rates, slowing down or even pausing to facilitate organelle targeting, domain folding and co-translational assembly, but prolonged stalls are deleterious to cells because they can deplete functional ribosomes and produce highly toxic truncated nascent chains. Stalls that occur on endoplasmic reticulum (ER) ribosomes are even more damaging because they additionally obstruct the translocons through which all secreted and membrane proteins must transit en route to the secretory pathway. Ribosome quality control (RQC) is a conserved and essential process that rescues stalled 60S subunits by extracting the obstructing nascent chain and degrading it by the UPS. Despite immense progress in the past decade in defining RQC for cytosolic ribosome, the how RQC operates for ER-stalled ribosomes is almost completely uninvestigated. Recently my lab discovered that UFMylation – the process by which UFM1, a small ubiquitin-like protein is conjugated to ribosomes — plays a central and essential role specifically in adapting RQC to proteins synthesized at the ER. The three specific aims described in this proposal seek to build on the foundation provided by our extensive preliminary data to define the mechanism by which nascent chains that obstruct ribosomes that stall on ER translocons are extracted and degraded. In aim 1 we will conduct a systematic dissection of the RQC machinery to define the role of known RQC required to resolve stalled ribosomes at the ER. In aim 2 we will define the interplay between ribosome UFMylation and RQC and dentify the readers of UFMylated ribosomes at the ER. In aim 3 we will determine the structure of UFMyulated ribosomes and the E3 ligase that conjugates UFM1 to the ribosome.