University Of California At Davis

NIH Research Projects · FY 2026 · 2024-02

PROJECT SUMMARY/ABSTRACT Homologous recombination is an essential DNA repair process that facilitates accurate chromosome segregation during meiosis, the specialized cell division that produces sperm and eggs. A fundamental gap in our understanding of recombination during meiosis in mammals is the role the breast cancer associated protein, BRCA2, which loads the central recombination enzyme RAD51 onto the ends of DNA breaks. This knowledge gap persists because essential factors like BRCA2 are inherently hard to study, and reagents to detect and analyze BRCA2 are limiting. These impediments are overcome in this proposal with the development of genetic, genomic, and biochemical reagents and approaches to study the meiotic functions of BRCA2. These include a fully functional epitope-tagged allele of the native Brca2 gene in mouse; high- resolution mapping approaches to define BRCA2 binding sites genome wide; germ-cell specific Cre-lox mouse lines to ablate Brca2 gene function at different stages of meiosis; and purified full-length human BRCA2 protein and its partner proteins for biochemical analysis. Exploiting this unique constellation of tools, the long-term objective of this project, to understand the roles and regulation of BRCA2 during meiosis in mammals, will be pursued through three aims. Aim 1 will define the spatial-temporal binding patterns of BRCA2 and related recombination factors using super-resolution immunofluorescence imaging, complemented by genome-wide ChIP-Seq mapping. Aim 2 will determine the roles of BRCA2 throughout meiosis by inactivating the Brca2 gene in mouse spermatocytes and oocytes before, during and after meiotic prophase. Mutant phenotypes will be analyzed using a battery physiological, histological, cell biology, and molecular assays. Preliminary data support the hypothesis that BRCA2 functions at multiple stages of meiosis, and in the maintenance of ovarian reserves. Aim 3 will define how a meiosis-specific chromosomal protein, SYCP3, modulates meiotic functions of BRCA2 to optimize meiotic HR. Super-resolution imaging and genome-wide ChIP-Seq mapping of BRCA2 and related recombination factors will be performed in Sycp3 mutant mice. Biochemical assays with purified human proteins will test the hypothesis that SYCP3 binds BRCA2 to facilitate the balanced assembly of RAD51 and its meiotic homolog DMC1 at recombination sites to promote DMC1-catalyzed recombination between homologous chromosomes. The results of these aims will provide unprecedented insights into the missing biology of BRCA2 function during meiosis in mammals. These fundamental discoveries will be germane to understanding pathologies associated with human meiosis, including infertility, miscarriage, congenital disease, and premature ovarian insufficiency.

NIH Research Projects · FY 2026 · 2024-01

Diabetic foot ulcers (DFUs) are the leading cause of lower extremity amputations in the US and are responsible for more hospitalizations than any other complication of diabetes. The sheer number of diabetic ulcers that progress to amputation underscores the inadequacy of conventional therapies and the need for novel approaches. Phagocytic leukocytes - particularly neutrophils - play a major role defending wounds from invading pathogens. Yet, despite excessive neutrophil influx and persistent non-resolving inflammation, DFUs are highly vulnerable to infection with pathogenic bacteria, such as Pseudomonas aeruginosa and Staphylococcus aureus, which further contribute to their impaired healing. Although, impairments in the diabetic neutrophil’s bactericidal functions have been blamed for this major co-morbidity, what causes these impairments and whether they can be corrected or overcome, remain poorly understood. Recently, we demonstrated that neutrophil trafficking is delayed in diabetic wounds, and this delay in neutrophil response, renders diabetic wounds vulnerable to infection early after injury, which in turn exacerbates wound damage and impairs healing. We further showed that reduction in the formyl peptide chemokine receptors (FPR) in diabetic neutrophils (due to high glucose) is responsible for this delay. We found some auxiliary receptors (e.g., CCR1) that remained functional under diabetic conditions but they were not functioning because of their ligands were not adequately expressed in diabetic wounds during the acute phase of healing early after injury. Importantly, we showed that one-time topical treatment with CCL3 (a ligand for CCR1) restored the dynamics of neutrophil and inflammatory responses in diabetic wounds, which in turn reduced infection by >99%. CCL3 treatment also substantially improved wound healing in diabetic mice. Diabetic neutrophils are known to have impairments in their bactericidal functions, although it remains unclear what causes these impairments. It remains unclear how neutrophils (recruited into diabetic wounds by CCL3 treatment) could destroy bacteria and reduce infection if they have bactericidal functional impairments. In this proposal, we will determine the molecular mechanism(s) underlying impaired antimicrobial functions in diabetic neutrophils (Aim 1). We will also evaluate CCL3 therapeutic doses and assess their efficacies and safety profiles in diabetic mice and diabetic pig models (Aim 2). If we are successful, our studies will fill crucial scientific gaps and reveal mechanistic insights regarding defective mechanisms underlying impaired bactericidal functions in diabetic neutrophils. They will also determine the optimum dose(s) and delivery mechanism of CCL3 for topical clinical use to control infection and to stimulate healing in diabetic wounds. Aim 2 will also enable us to obtain an IND for topical therapeutic application of CCL3 in human diabetic patients with non-healing ulcers and has the potential to be applicable to all wounds.

NIH Research Projects · FY 2025 · 2024-01

Interferometric near-infrared spectroscopy for transabdominal fetal oximetry The outcomes and cost of childbirth is an important issue that almost every family faced or will face. Compared to vaginal delivery, Cesarean section (C-section) has a higher cost, and may increase the health risk to both the baby and mother. About 32% of the deliveries are via C-section, which is much higher than the 10-15% ideal rate published by World Health Organization. A major triggering factor of C-section is a conservative prediction of fetal hypoxia during labor, which unfortunately has a high false-positive rate using the existing, widely-adopted intrapartum fetal monitoring technique of cardiotocography (CTG). Strikingly, since the introduction of CTG in the early 1970s, the rate of C-section deliveries in the US has risen five folds, while the rates of conditions associated with hypoxia remain unchanged. A noninvasive measurement of the fetal oxygen saturation during labor and delivery could provide obstetricians specific indicator of the necessity of emergency C-section. This may help to reduce the rate of unnecessary C-sections. Here, we propose a new transabdominal fetal oximetry (TFO) technique based on an innovative interferometric near-infrared spectroscopy (iNIRS) approach that could noninvasively measure the fetal arterial blood oxygen saturation. The iNIRS TFO measures the time-resolved reflectance of near-infrared light shining on the maternal abdomen. Our new method is distinctly different from the conventional pulse oximetry, and offers critical advantage to detect weak fetal signals buried in strong overwhelming maternal signal through the abdomen while greatly reducing the required light power on the tissue. Our approach distinguishes photons traversing different tissue depth, thus facilitating the separation of signal between the shallow maternal layer and the deep fetal layer. It greatly increases sensitivity to deep tissue signals and assures both an accurate and safe measurement. We conducted preliminary experiment on pregnant ewe and verified our technique could detect the fetal heartbeat through a single optical wavelength and a single detector, which presages the feasibility of iNIRS to measure fetal oxygen saturation. Here, we aim to build a two-wavelength iNIRS with a significantly improved measurement sensitivity to perform transabdominal fetal oximetry. The two optical wavelengths could extract the relative change of the oxygenated and deoxygenated hemoglobin during a cardiac cycle, and we will use time- division-multiplexing to switch between two optical wavelengths in the iNIRS setup. To improve the tissue penetration depth, we will increase the frequency tuning rate of the laser, use an innovative detection fiber and increase the number of detectors. All these approaches could increase the measurement sensitivity and thus the accessible tissue depth. Finally, we will develop advanced machine learning algorithms to extract the fetal oxygen saturation level from the raw measurements. We will perform the experiment in pregnant ewe model and validate the results with the arterial blood gas (ABG) test. The success of our project will make a significant impact on both the outcomes and cost of childbirth.

NIH Research Projects · FY 2026 · 2024-01

Astrocytes, the most populous glial cell in the central nervous system (CNS), play essential role in brain development and function including ion/neurotransmitter homeostasis, synapse formation/removal and synaptic transmission modulation. These crucial functions are molecularly determined by their gene signatures which are controlled by nuclear transcription factors. In response to neurological diseases and injuries, astrocytes become reactive astrocytes and adopt neurotoxic and/or neuroprotective functions which modulate CNS pathophysiology. The ultimately functional output of reactive astrocytes is determined by their differential gene expression signature which is controlled transcriptional factors. Our long-term goal has been centered on unveiling mechanisms underlying CNS glial cell homeostasis and pathophysiology with a focus on the transcription factor SOX2. We found that SOX2 is highly enriched in adult quiescent astrocytes and retained in reactive astrocytes in response to inflammatory demyelinating injury. However, little is known about the functional significance of SOX2 in regulating astrocyte biology and pathology. Recently, we reported a crucial role of SOX2 in astrocyte maturation and animal behavior. As a transcription factor, SOX2 binds to the regulatory elements of a cohort of astrocytic signature and functional genes and modulates their expression. These data highlight the importance of SOX2 in astrocyte development and function. We employed a “top- down” approach to investigate functional significance of SOX2 in adult astrocytes. Astrocyte-specific SOX2 conditional knockout (cKO) results in hyperactive locomotor yet normal motor coordination and cognition functions. Furthermore, astrocyte SOX2 cKO mice develop worse clinical symptoms in response to EAE injury. In this project, we will test the hypothesis that SOX2 is essential for adult astrocyte homeostasis and function and its deficiency in reactive astrocytes exacerbates inflammatory demyelinating injury. This project, upon completion, will provide the first conceptual landscape of the functional significance of SOX2 in quiescent astrocytes and reactive astrocytes.

NIH Research Projects · FY 2026 · 2024-01

PROJECT SUMMARY Changes in physiological stimuli reprogram the translation machinery to alter preferential recruitment of mRNAs to the ribosome and which start site on a mRNA is selected. Genome wide analysis using ribosome profiling has revealed far more extensive regulation of these events than previously appreciated, yet the mechanism of mRNA recruitment and its regulation remains poorly understood. This knowledge gap persists mainly because of difficulties in determining which intermediate step(s) in the initiation pathway function as kinetic checkpoints to control mRNA recruitment. To date, initiation pathway intermediates have largely been identified on the basis of their thermodynamic stability which must withstand traditional assays (e.g. sucrose gradients and immunoprecipitations). These approaches take minutes to hours to perform, and/or require cross-linking agents to stabilize them, but most intermediates prior to initiation codon selection occur on the sub-second to second time scale. To determine how mRNAs are selected for translation, one must develop and use assays that can precisely monitor the formation of pathway intermediates in real-time. To overcome this bottleneck and move the field forward, we have developed innovative ensemble and single-molecule fluorescence-based assays that can monitor the rate of mRNA recruitment to the ribosome in real-time. Our highly purified reconstituted system will enable us to successfully test and build models with which to understand mRNA recruitment and its regulation. Models that we generate will be tested using translation assays in cell-free extracts and intact cells. Our long-term objective is to understand the mechanism by which alterations in initiation factor availability and their post-translational modification reprograms the translational apparatus to control which mRNAs are translated in response to physiological stimuli.

NIH Research Projects · FY 2026 · 2024-01

SUMMARY: Circadian rhythm is important for human physiology and health. Human body temperature increases during wakefulness and decreases during sleep. This body temperature rhythm (BTR) is a robust output of the circadian clock and is fundamental for maintaining homeostasis and its related processes, such as sleep and metabolism. The long-term goal of our research is to understand the molecular and neural mechanisms by which BTR is regulated and how BTR is related to sleep regulation. To understand the mechanisms of BTR, we use Drosophila. This model has provided many strong contributions to studying the circadian clock, including the discovery of conserved mammalian circadian clock genes and mechanisms. We demonstrated that Drosophila exhibits a circadian rhythm of temperature preference, referred to as temperature preference rhythms (TPR). While mammals regulate BTR by generating or losing internal heat, small ectotherms, including Drosophila, regulate BTR via selecting an environmental temperature. As flies are small ectotherms, their body temperature is close to that of the surrounding environment. Thus, Drosophila TPR produces BTR in a similar pattern as mammals. Furthermore, our study provides the first evidence that fly DH31R and its mammalian homolog, Calcitonin receptor, CALCR, regulate BTR. Thus, understanding fly TPR will provide fundamental insights into the molecular and neural mechanisms that control BTR in mammals. Using fly TPR, in this study, we will address an outstanding knowledge gap regarding how temperature fluctuations are determined and how the preferred temperature is set at a specific time of day. We will elucidate the molecular and neural mechanisms underlying TPR. We will examine the distinct functions of each pacemaker by manipulating their activities and neuropeptide-receptor signals which could control temperature fluctuation or temperature setpoint. We will clarify the mechanistic difference between TPR and locomotor activity, focusing on a noncanonical pathway that we anticipate to be a specific regulator of TPR. This study will unveil the unique neural circuits controlling fly TPR. Further, we will take advantage of fly TPR and determine the mechanistic link between temperature change and sleep; their relationship has been well known, but the underpinning molecular mechanism has been a big mystery. This study will facilitate an understanding the regulatory mechanisms underlying BTR and its relationship to sleep. We have shown that parallel mechanistic functions between mammalian BTR and fly TPR; thus, the basic foundation of BTR is aligned with fly TPR. The outcome of this R35 proposal will facilitate to reveal BTR mechanisms in mammals and lend actionable insights into the treatment of circadian clock diseases, sleep problems, and the health of night-shift workers.

NIH Research Projects · FY 2025 · 2024-01

PROJECT SUMMARY Heart failure (HF) with reduced ejection fraction is a leading cause of death in the US. HF is associated with autonomic dysregulation and significant remodeling of calcium handling, excitation-contraction coupling, and electrophysiology, which collectively lead to contractile dysfunction and increased risk of ventricular arrhythmia. HF incidence and outcomes are strongly sex-dependent; men have a higher incidence of HF, and in patients with non-ischemic cardiomyopathy, women display a lower arrhythmia propensity than men. The precise mechanisms underlying these sex differences and the protection in female hearts remain unclear. Our initial studies discovered regional- and sex-dependent differences in b-adrenergic receptor (b-AR)- cyclic AMP (cAMP) signaling that lead to previously unrecognized functional electrophysiological differences in male and female mice, and could act as a potential anti-arrhythmic mechanism in female hearts. By combining whole- heart, cellular, and subcellular approaches, this project aims to uncover novel mechanisms that underlie sex differences in b-AR-cAMP signaling, which may play a role in sex-dependent outcomes in HF. A multi-level experimental approach will be employed, investigating at the nanoscale, cellular, and whole heart level, to examine: 1) the structural and functional mechanisms underlying sex differences in b-AR-cAMP signaling in the intact heart, and 2) how sex- and region-dependent sympathetic remodeling in HF impacts b-AR-cAMP signaling and arrhythmias. We propose to collaborate with a multidisciplinary advisory team to use novel whole-heart FRET + optical mapping, isolated myocyte experiments, biochemical approaches, and super resolution microscopy to assess cellular and subcellular mechanisms underlying regional- and sex-differences in b-AR- cAMP signaling. How these signaling cascades and functional outputs are remodeled in a rodent model of HF will also be tested. Completion of this proposal will significantly advance our understanding of sex differences in cardiac patho-physiology and may provide insight into new gender-specific therapeutic approaches. UC Davis offers an exceptional training environment for the mentored phase of the award to achieve these goals. Moreover, the proposed research and training plan will significantly contribute to the applicant's personal and professional growth. The mentored phase of this award will provide an invaluable training opportunity to develop a unique scientific and professional skillset necessary to address the goals of this proposal, as well as prepare for independence and make the applicant a competitive candidate for faculty positions. The independent phase of this award will provide time, and funds, to create an independent research program in cardiovascular physiology.

NIH Research Projects · FY 2026 · 2024-01

PROJECT SUMMARY (ABSTRACT) As the population ages, the prevalence and risk for Alzheimer’s disease and related dementia (ADRD) is expected to increase with even greater risk for Blacks and Hispanic/Latinx groups. Historical and contemporary structural racism in the form of residential segregation is a fundamental cause of health disparities that may contribute to ADRD inequities. However, residential segregation in some communities may inadvertently lead to formation of ethnic enclaves where higher concentration and proportion of some ethnic groups may be protective against the negative effects of segregation. Some studies found that higher residential segregation is associated with greater cognitive decline and higher risk for ADRD, but other found that it may be protective. It is unclear how lifecourse residential segregation is associated with cognitive decline and risk for ADRD. This project will determine the relationship of lifecourse residential segregation with ADRD using measures of incident ADRD, cognition and neuroimaging brain biomarkers. Furthermore, we will explore how education and poverty rates may mediate the relationship of segregation and ADRD risk. We will use data from the KHANDLE and STAR cohort studies, two longitudinal cohort studies of racial/ethnically diverse participants of older-adults residing in the Northern California area. Measures of residential segregation indices of evenness, exposure, concentration, centralization, and clustering will be derived from decennial census data from 1970s to 2010s and linked with historical addresses of participants across 7 timepoints (birth, ages 5, 10, 18, 30, 40, 65). Causal inferences models will be used to (Aim 1) examine the association of lifecourse residential segregation and cognition for approximately 2476 participants and (Aim 3) examine 614 subset of participants with MRI markers of brain health. Mediation analysis (Aim 2) will be conducted for educational quality and attainment, and poverty rates to assess the impact of proximal and distal social determinants of health on the relationship between residential segregation and late-life cognition. As of current, this is the first study to our knowledge that will examine the lifecourse timing and duration of residential segregation with cognitive decline and risk for ADRD in multiple racial groups. This study present opportunities to comprehensively assess the effects of lifecourse residential segregation, significantly improving our understanding of structural racism on late-life cognitive outcomes and ADRD. Findings from this study may inform decision-making policies on equitable home lending, and neighborhood reform for healthy cognitive aging. With the support of an exemplary team of experts, the applicant will be trained and enhanced with skills, research proficiency, and rigorous methodology needed to carry out this project and advanced his career as a researcher on health disparities.

NIH Research Projects · FY 2026 · 2023-12

PROJECT SUMMARY/ABSTRACT Extended-spectrum beta-lactamase (ESBL)-producing Enterobacterales (ESBL-PE) have become widespread in recent years, posing a serious threat to the treatment of common bacterial infections. Increased use of "last resort" carbapenem antibiotics to treat ESBL-PE infections has fueled the emergence of carbapenem-resistant Enterobacterales. Whole genome sequencing (WGS)-based prediction of antibiotic resistance phenotypes may enable early initiation of appropriate treatment options for ESBL-PE infections, including selection of carbapenem-sparing beta-lactam regimens. However, existing genomic prediction models have had relatively poor accuracy for predicting resistance to beta-lactams in ESBL-PE. The genetic environment of ESBL genes, including their location in the bacterial genome and association with mobile genetic elements (MGEs), is unaccounted for in these models. These structural factors may contribute to beta-lactam resistance by providing promotors modulating the expression of ESBL genes and enabling their duplication and mobilization within and between Gram-negative bacteria. We hypothesize that including MGEs in genomic prediction models will improve detection of beta-lactam resistance phenotypes in ESBL-PE. The objectives of this proposal are to 1) determine how MGEs modulate beta-lactam minimum inhibitory concentrations (MICs) in ESBL-PE and 2) develop and optimize multivariate regression models for predicting beta-lactam resistance from WGS data in ESBL-PE. Our multidisciplinary research team has the necessary expertise in bacterial genomics, molecular biology, infectious diseases epidemiology, and biostatistics to ensure successful completion of the proposed studies. To characterize the genetic environment of ESBL genes, we will take advantage of the ability of nanopore WGS to resolve structurally complex genomic regions including MGEs and plasmids. We will test our hypothesis through the following three aims: 1) we will comprehensively genotype and phenotype a large collection of clinical ESBL-PE isolates (n=450) to test the association between specific MGE genotypes and beta-lactam MICs; 2) we will evaluate potential mechanisms whereby MGEs may affect beta-lactam resistance phenotypes, focusing on the role that MGE-associated promotors play in modulating ESBL gene expression and the impact of ESBL gene copy number on beta-lactam MICs; 3) we will use machine learning to build and optimize multivariate regression models incorporating MGE genotypes and other genetic factors to predict beta- lactam resistance phenotypes and validate our resulting models in a prospective collection of 200 ESBL-PE isolated from urine cultures. The development of rapid diagnostic methods that predict antibiotic treatment options to ESBL-PE should be a research priority. In addition to determining how MGEs contribute to ESBL-PE resistance phenotypes, the proposed research will help enable integration of nanopore WGS into surveillance and diagnostic approaches to detect beta-lactam resistance in ESBL-PE and facilitate the selection of appropriate carbapenem-sparing regimens for ESBL-PE infections using genomic prediction models.

NIH Research Projects · FY 2026 · 2023-12

Abstract . Vascular complications associated with lifestyle-related diseases, such as diabetes, contribute to an increased risk of hypertension, coronary artery disease, and heart failure associated with preserved ejection fraction (HFpEF). These vascular complications may involve, at least in part, vascular smooth muscle (VSM) dysfunction leading to impaired vasomotor function, but mechanisms remain poorly understood. We recently identified a novel AKAP5/P2Y11/AC5/PKA/CaV1.2 nanocomplex that is activated by the release of extracellular nucleotides, such as ATP. This complex controls VSM contractility, vasoconstriction, and altered blood flow (BF) and blood pressure (BP) in response to elevated extracellular glucose (e.g. hyperglycemia; HG – a major metabolic abnormality in diabetes). Yet, whether HG stimulates cellular nucleotide release, the precise molecular entity underlying this process, and its potential role in diabetes and other diseases (e.g. HFpEF) are unknown. The overarching goal of this MPI proposal is to provide insight into these issues. Our preliminary data offers a unique window into these queries and uncover an essential role for the large-pore channel pannexin 1 (Panx1) as a key member and upstream protein activating the nanocomplex and thus contributing to altered vascular reactivity during diabetes and perhaps other diseases such as HFpEF. We will address the novel central hypothesis that Panx1 is upstream and part of the AKAP5/P2Y11/AC5/PKA/CaV1.2 signaling axis that promotes VSM contractility and altered BF/BP in response to diabetic hyperglycemia. The proposal has high significance as it defines Panx1 as 1) a key upstream pathway triggering pathological signaling resulting in VSM hypercontractility during diabetic hyperglycemia and 2) a potential therapeutic target to treat vascular complications in diabetes (and perhaps HFpEF). The influence of Panx1 in contributing to the ATP release in response to HG, and in strengthening/weakening the formation of nanocomplexes to mediate functional responses, which may have broader implications in any Panx1-dependent signaling process, is an emerging and innovative concept. Our innovative multi-scale approach using state-of-the-art approaches will be implemented to explore the following aims. Aim 1 is to elucidate the role of Panx1 on HG signaling in VSM. Aim 2 is to determine the impact of Panx1 on VSM dysfunction during diabetes and HFpEF. The impact of the proposal is in establishing Panx1 as a key contributor to defective VSM function and vascular complications in diabetes and HFpEF.

NIH Research Projects · FY 2026 · 2023-12

Title: Two-way Magnetic Resonance Tuning Nanoprobe Enhanced Subtraction Imaging for Precision Diagnosis of Brain Metastasis Project Summary/Abstract Metastasis from systemic cancers to the brain is a leading cause of cancer mortality. The current diagnostic method is sensitive only to larger tumors when therapeutic options are limited. Visualizing early brain metastases by non-invasive imaging approaches with high sensitivity and spatial resolution followed by timely treatment is crucially important to reduce their high mortality rate. While effective interventions (e.g., surgery, radiation, targeted therapy, and immunotherapy) strongly depend on our ability to detect brain metastases at an early stage, imaging small brain metastases hidden in a large population of normal cells presents a unique challenge. It is essential to design novel imaging approaches to detect small brain metastases with the highest possible tumor-to-normal tissue ratios (TNRs). The goal of this application is to develop a new molecular nanoprobe with activatable magnetic resonance contrast integrated with a new computational subtraction approach to improve the TNR of imaging for small brain metastases. We recently developed a new two-way magnetic resonance tuning (TMRET) nanoprobe with dually activatable T1&T2 magnetic resonance signals coupled with dual-contrast enhanced subtraction imaging (DESI) to dramatically enhance contrast in targeted tissues and suppress the background signal from normal tissue. This integrated platform could sensitively detect very small tumors in the brain by magnetic resonance imaging (MRI) in patient-derived xenograft (PDX) models with a TNR >10. We also developed a Sequential Targeting In CrosslinKing (STICK) nano-delivery strategy to “stick in” central nervous system (CNS) tumors and metastases, which will be applied to our TMRET nanoplatform to enhance its specific delivery to brain metastases. In this project, we will develop novel blood-brain barrier (BBB)-traversing and deep tumor-penetrating TMRET (bt-TMRET) nanoprobes with superior TNR for sensitive and specific detection of brain metastases. The STICK strategy will be used to improve the CNS pharmacokinetics (PK) of TMRET nanoprobes by pHe-cleavable crosslinkers to maximize the time window for transcytosis through the BBB. Our STICK strategy will further enhance the efficiency of BBB traversal by manipulation of glucose transporter 1 (GLUT1) on the BBB by optimization of the polyvalent interaction of nanoprobes with GLUT1 via fine-tuning the surface targeting moieties. The STICK strategy optimizes the pH-responsive size transformation for improved tumor tissue penetration and sialic acid-targeting selectivity for enhanced tumor cell specificity. The MRI signal can be turned ON specifically at the brain metastases after BBB traversal and tumor penetration via size transformation in acidic tumor microenvironment. Our hypothesis is that the proposed nanoprobes can improve the TNR for MRI detection of small brain metastases through a synergized strategy of background suppression, signal amplification via deep penetration and specific targeting in brain metastases and activation at tumor sites, and DESI technology will further enhance the TNR. This new imaging platform is expected to improve cancer detection in the clinic and serve as a great tool for biomarker detection in preclinical research.

NIH Research Projects · FY 2026 · 2023-12

Pulmonary arterial hypertension (PAH), a progressive fatal disease, manifests by remodeling of pulmonary ar- teries (PA), leading to increased PA pressure, right heart failure and death. The key component of PA remodeling is the progressive vessel wall thickening due to hyper-proliferation of PA smooth muscle cells (PASMC), endo- thelial cells (PAEC), and adventitial fibroblasts (PAAF) the mechanisms of which are not completely understood. PASMC in PAH switch to unique disease-specific phenotype, characterized by metabolic shift to glycolysis, au- tonomous proliferation, and apoptosis resistance. PAH pulmonary vascular cells are also highly secretory and support pro-proliferative microenvironment, further amplifying PA remodeling and PAH. Published and our new preliminary data strongly suggest that metabolite L-lactate acts as a central regulator of the molecular and met- abolic processes responsible for pulmonary vascular cell hyper-proliferation, remodeling, and PAH. Our pilot data show that lactate, over-produced by PAH PASMC due to over-expression of lactate dehydrogenase A (LDHA), promotes aberrant lactylation of DNA topoisomerase 1 (TOP1) and EMILIN1, leading to TOP1 up-reg- ulation and EMILIN1 deficiency, consequent up-regulation of pro-proliferative Akt-mTOR, Yap/Taz, TGFβ, in- creased proliferation and survival. Our data also suggest that lactate over-production in PAH PASMC is self- supported via glycolysis and EMILIN1-TGFβ1-HIF1α-LDHA circuit, and that lactate is secreted by PAH PASMC and promotes proliferation of PA endothelial cells (PAEC) and adventitial fibroblasts (PAAF). We further report that suppression of LDHA-lactate axis reduces proliferation and induces apoptosis in human PAH PASMC, re- verses pulmonary vascular remodeling and experimental PH in mice. We propose to elucidate the role and mechanisms of regulation and function of LDHA-lactate signaling in PAH pulmonary vasculature and explore the benefits of targeting this pathway to correct mechanistic abnormalities and reverse PA remodeling and PH. Spe- cifically, we will: (1) critically test the role of LDHA-lactate in PAH PASMC proliferation and survival, pulmonary vascular remodeling and PH using human PAH and non-diseased PASMC and lung tissue samples and SM- specific Ldhaknock-out mice; determine the relationship among LDHA-lactate, lactylation of TOP1 and EMILIN1, and Yap/Taz, Akt-mTOR, and TGFβ1 in regulating PASMC proliferation and survival; (2) investigate whether lactate over-production is supported through up-regulation of glycolysis and EMILIN-TGFβ1-HIF1α-LDHA feed- forward loop, evaluate the metabolic consequences of lactate over-production, and determine the role of PASMC-secreted lactate in the proliferation of PAEC and PAAF; and (3) examine whether targeting lactate sig- naling by LDHA inhibitor oxamate and TOP1 inhibitor indotecan selectively inhibits proliferation and induces apoptosis in vitro in human PAH PASMC, reverses or attenuates experimental pulmonary vascular remodeling and PH in rats. Proposed study will identify new critical mechanism of pulmonary vascular remodeling and dis- sect new remodeling-focused molecular target(s) for therapeutic intervention. .

NIH Research Projects · FY 2025 · 2023-12

Myalgic encephalomyelitis/chronic fatigue syndrome (ME/CFS) is a complex, debilitating disease with inconclusive causes and no laboratory-based diagnostic tests. Oxygen tension (PO2)-regulated red blood cell (RBC) capillary velocity has emerged recently as a new mechanism regulating cerebral capillary perfusion and measures RBC responses to local hypoxia. Such intrinsic RBC responses to PO2 changes may change distinctly under different pathological conditions and thus represent a new RBC-based approach for disease diagnosis. Indeed, our preliminary results showed that RBCs collected from ME/CFS patients exhibited impaired velocity in a microfluidic capillary in response to reduced PO2 and PO2-regulated RBC capillary velocity was improved significantly when patients received craniocervical instability surgery, strongly suggesting that PO2-regualted RBC capillary velocity may represent a new characteristic of ME/CFS that might be used to diagnose ME/CFS and measure ME/CFS progression. Here, we extend our preliminary study and propose an interdisciplinary approach combing microfluidics, machine learning and RBC cytokine assay to examine rigorously the accuracy of PO2-regulated RBC capillary velocity as a new laboratory test for ME/CFS and investigate mechanistically the roles of RBC cytokine signaling in ME/CFS. Aim 1 will measure PO2-regulated RBC capillary velocity using RBCs from 96 participants and compare the results of ME/CFS patients with age, gender and race-matched healthy controls. Furthermore, we will develop machine-learning algorithms to establish a diagnostic classifier to validate PO2-regulated RBC capillary velocity as a new laboratory test for ME/CFS and assess its feasibility to differentiate ME/CFS patients with different disease severity. Mechanistically, we will examine hemoglobin-band 3 interactions in RBCs from ME/CFS patients and examine their correlations with increased oxidative stress in ME/CFS. Aim 2 will quantify the cytokine profile in RBCs from ME/CFS patients and compare them with age, gender and race-matched healthy controls. Furthermore, we will alter the RBC cytokine profile and measure the corresponding changes of PO2-regulated RBC capillary velocity to examine whether RBC cytokine profile plays a role in the modulation of PO2-regulated RBC capillary velocity. These experiments will provide a previously unrecognized RBC cytokine signaling in ME/CFS and add new insights to the immune dysregulation in ME/CFS. Together, the proposed studies exploit new tools and technology to develop a new laboratory test for ME/CFS and reveal mechanistims underlying PO2-regulated RBC capillary velocity in ME/CFS, which we believe will directly improve the diagnosis and treatment of ME/CFS.

NIH Research Projects · FY 2025 · 2023-11

Recent studies support the hypothesis that the host plays an active role in maintaining homeostatic symbiosis with the gut microbiota. However, the host-specific mechanisms employed to sculpt the microbiota remain to be fully elucidated, in part, due to shortcomings of current in vivo models. Data from our group and others support that -defensins, abundant antimicrobial effector molecules of small-intestinal Paneth cells, help shape the composition of the gut microbiota. Perturbations in Paneth cell function contribute to pathogenesis of disease by disturbing the homeostatic balance of the microbiota (dysbiosis). Yet, little is known regarding the role of Paneth cell effectors (e.g., -defensins) in mediating changes in the gut microbiota during dynamic physiological (non-disease) processes. This proposal aims to develop two innovative mouse models valuable for delineating host mechanisms that shape gut microbiota during processes intrinsic to mammalian biology. During lactation, arguably one of the most strenuous physiological processes for mammals, intestinal biology must adapt to the demands of reproductive biology. Our preliminary data demonstrate that in lactating dams, rapid and dramatic changes in the small intestine at both the gross and molecular levels accompany a major restructuring of gut-associated microbial communities. Concomitantly, during lactation a striking and unsynchronized change in the expression of individual Paneth cell -defensins is observed at a scale far greater than previously observed in other experimental settings, including enteric infection and inflammation. We will address the hypothesis that during lactation, a period of extraordinary energy demand, a dramatic alteration in α-defensin production drives changes in the microbiota that serve to benefit the dam (effective energy harvest from foodstuffs). A corollary hypothesis that changes in the microbiota of the lactating dam benefit the offspring (e.g., facilitation of vertical microbiota transmission) will be addressed in the future using the model developed herein. Aim #1 will develop the lactating dam as a novel model to interrogate dynamic and physiological host-microbiota interactions in mammals. Factors impacting dynamics of the host-microbe interface will be systematically delineated to create a robust and reproducible model for study. Aim #2 will develop a novel Paneth cell -defensin knockout mouse to explore the necessity of -defensins in driving microbiota changes during lactation; this knockout model will also be invaluable to the field for other studies of intestinal innate immunity. The paradigm of pronounced maternal intestinal remodeling during lactation and its potential long-term influence on both maternal and offspring health has been largely unexplored. The dynamically adapting intestine of the lactating dam may represent an ecosystem of host-microbe interactions fundamental to mammalian life. Successful completion of these Aims will establish valuable models to better understand host-mediated mechanisms to shape the gut microbiota, contributing insights on intestinal innate mucosal immunity, as well as providing foundations for application to other disciplines.

NIH Research Projects · FY 2026 · 2023-11

ABSTRACT Autism spectrum disorder (ASD) is typically described as a disorder of childhood; however, challenges in socioemotional behavior and cognition persist across the lifespan. Nearly 6 million adults in the United States currently live with ASD, with prevalence more than doubling in the last 10 years. Recent studies suggest that adults with ASD are twice as likely to develop dementia, and have an increased prevalence of age-related physical and psychiatric conditions. While brain and behavioral differences are well researched in children with ASD, a major gap in knowledge exists for the adult brain. The brains of individuals with ASD undergo an atypical developmental trajectory, marked by initial excess in volume, neuron density, and connectivity in childhood that is followed by a progressive reduction in cell number, myelin thickness, and synapses into adulthood. We propose that early neuronal excess and local over-connectivity, in concert with systemic neuroimmune dysregulation, may render the ASD brain vulnerable to age-related pathological and pro- inflammatory processes beginning in early adulthood that may ultimately lead to cellular dysfunction and later cognitive decline. Here we aim to determine the cell types affected as individuals with ASD age through adulthood and identify neuroinflammatory markers specific to those cells. We will utilize a unique sample of clinically and genetically characterized human postmortem adult brains, examining regions implicated in the socioemotional and neurobiological impairments of ASD and aging. We hypothesize that aberrant immune activation of pro-inflammatory processes contribute to the altered trajectory of brain development, as well as deleterious neuropathogenic protein aggregations associated with cognitive decline. We will utilize unbiased transcriptomic methods to determine which genes may be differentially expressed in specific cell groups, and assays to determine which proteins, including cytokines, may be disproportionately affected with age in ASD (Aim 1). Informed by these results, we will carry out a systematic examination of the expression of high-priority markers in specific cell types with preserved spatial information in brain tissue sections (Aim 2). Finally, we will examine the effects of neuroinflammation and aging in ASD on the synaptic connections between neurons (Aim 3). These critical, discovery-driven studies will serve as an essential first step to characterize cellular and molecular processes associated with brain aging in ASD, and provide a fundamental platform upon which mechanistic studies and animal models can be built. This project will define the neuropathological features associated with brain aging in ASD and correlate these variables with extensive clinical and genetic information for each subject, in support of precision medicine. Ultimately, we aim to shift the mindset of autism research toward the view of adulthood not as an endpoint, but as part of a trajectory of brain development across the lifespan, with the goal of promoting healthy aging in the rapidly growing population of adults living with ASD.

NIH Research Projects · FY 2026 · 2023-11

Toxoplasma gondii is an obligate intracellular parasite that can cause severe disease in congenitally infected infants and in immunosuppressed people. Toxoplasma co-opts host cells by secreting effector proteins, called ROPs and GRAs, into the host cell. In mice, the cytokine interferon gamma (IFNγ) is essential to control Toxoplasma by inducing a variety of parasiticidal mechanisms while in humans the cytokine tumor necrosis factor (TNF)α can compensate for the absence of IFNγ. The IFNγ-induced mechanisms that control Toxoplasma in rodents are largely absent in humans and most of the parasite’s ROPs and GRAs that determine virulence in rodents play no role in counteracting the human IFNγ response. Thus, despite Toxoplasma’s enormous health implications, there is currently a lack of mechanistic knowledge on (i) how IFNγ- or TNFα-stimulated primary human cells inhibit Toxoplasma growth; and (ii) how Toxoplasma secreted effectors allow parasite growth in stimulated primary human cells. Our long-term goal is to determine how Toxoplasma secreted effectors mediate its survival in humans, even in the presence of a fully functioning immune system, which could help the development of targeted interventions to minimize the adverse health effects of toxoplasmosis in at-risk individuals. The overall objective for this application is to determine the mechanisms by which activated primary human cells detect and destroy Toxoplasma and how Toxoplasma counteracts these mechanisms. Our hypotheses are (i) that mechanisms of Toxoplasma growth inhibition in stimulated human cells are guanylate-binding protein (GBP1)-mediated breakage of the vacuole followed by endolysosomal fusion with the parasitophororous vacuole, which is Toxoplasma’s replication niche, and induction of early parasite egress; and (ii) that the novel Toxoplasma GRA effectors identified from our genome-wide parasite loss-of-function screen in IFNγ-stimulated human cells determine parasite resistance to IFNγ by preventing early parasite egress. The hypotheses will be tested by pursuing two specific aims: 1) Determine the mechanism by which IFNγ/TNFα-stimulated human cells inhibit Toxoplasma growth; and 2) Determine the mechanism by which Toxoplasma effectors affect parasite fitness in IFNγ/TNFα-stimulated human cells. The research proposed in this application is innovative because it is using results from an unbiased innovative CRISPR/Cas9 loss-of-function screen that identified Toxoplasma genes that determine resistance to IFNγ-mediated parasite growth inhibition in primary human cells. These results are expected to have an important positive impact because they will provide a science-based framework for the future development of novel targets for therapy, e.g., by stimulating host parasite killing mechanism or by inhibiting parasite virulence determinants. Our results will also aid prediction of Toxoplasma strain virulence in humans and identification of human toxoplasmosis susceptibility loci.

NIH Research Projects · FY 2025 · 2023-11

The varied and complex symptoms of neurodevelopmental disorders (NDDs) present significant challenges for the development of effective treatments. To provide targeted interventions, an understanding of neurobiological etiology is imperative. Studies have uncovered strong single gene risk factors for NDDs, prominent among which are genes coding for chromatin remodelers. The chromatin remodeler CHD8 has emerged among such genes with one of the highest frequency of de novo mutations in ASD cohorts. Intriguingly, CHD8 mutations have also been discovered in other neurological disorder cohorts, including schizophrenia and obsessive compulsive disorder (OCD). Thus, CHD8 represents a model for rare monogenic NDDs, with a common core set of symptoms associated with ASD, but overall broad and complex expressivity spanning early development through adulthood. Various Chd8 mouse models have been generated, with constitutive mutants exhibiting NDD-relevant phenotypes. However, in these models the wide expression of Chd8 and its involvement in early development have hampered dissociation of developmental effects from effects on mature neurons. Moreover, experimental emphasis on the cerebral cortex and hippocampus has limited advancement from genetic findings to causal neurobiology. The cerebellum (CB) has been consistently and causally associated with NDDs, including in emerging studies of developmental roles of Chd8, and has been implicated in schizophrenia and OCD. Even though roles of the CB have expanded from a motor control center to include cognitive and affective (i.e., limbic) functionality, CB non-motor contributions and pathology remain understudied in neuropsychiatric and NDD research compared to forebrain regions. Further, CB studies in NDDs have primarily investigated developmental impacts. However, Chd8 expression persists in the adult CB. This raises the novel hypothesis that conditional Chd8 deletion in the adult CB contributes to neuropsychiatric and NDD-relevant pathology. Here we propose initial work toward testing this model, defining the impact of conditional Chd8 ablation on adult CB function across behavioral, genomic and electrophysiological dimensions. In two aims, we will manipulate Chd8 expression in CB output nuclei and test CB-specific vulnerabilities in limbic behaviors; investigate CB transcriptomic phenotypes; and begin to probe pathology in the functional connectivity of CB output circuits to the limbic system. These experiments will establish impacts of conditional Chd8 ablation on the developed CB and output circuits; pinpoint molecular underpinnings of CB dysfunction; and begin to build a circuit-level understanding of how Chd8 ablation impacts CB connectivity with the rest of the brain. If successful, this work will lead to new avenues of research on CB dysfunction in NDDs by linking a high confidence risk gene with behavioral, cellular, molecular, and circuit deficits in the CB. By pinpointing biological processes and circuits that remain vulnerable to Chd8 ablation in the developed CB, this line of research has the potential to identify novel therapeutic targets over a time-extended treatment window.

NIH Research Projects · FY 2024 · 2023-09

PROJECT SUMMARY Polychlorinated biphenyls (PCBs) remain a significant risk to human health, and a primary target of concern is the developing brain. Epidemiological studies have reported positive associations between developmental exposures to PCBs and increased risk for neurodevelopmental disorders (NDD); however, experimental studies designed to assess the strength of these associations and identify biological mechanisms underlying PCB DNT have focused almost exclusively on the higher chlorinated (HC)-PCBs, the predominant congeners found in the legacy commercial PCB mixtures. In contrast, data regarding the potential for lower chlorinated (LC)-PCBs to interfere with neurodevelopment is extremely limited. This is a troubling gap considering recent reports that environmental levels of LC-PCBs are increasing worldwide and that the LC-PCB congeners 11 and 28 were found to comprise >70% of PCBs in the serum of pregnant women at increased risk for having a child with an NDD. We previously reported that PCB 11 and its metabolites formed via cytochrome P450 (CYP)-mediated oxidation promoted dendritic and axonal growth in vitro, and these effects were observed at concentrations relevant to the human gestational environment. Interestingly, the potency of the metabolites varied from that of the parent. Our in vitro studies also suggested that PCB 11 enhanced dendritic growth via activation of CREB- dependent signaling pathways, but whether the metabolites alter neurodevelopment via the same molecular mechanism is not known. We also do we know whether (1) other LC-PCBs found in human tissues have DNT activity; (2) LC-PCBs or their metabolites modulate other neurodevelopmental outcomes known to be regulated by CREB-dependent signaling, specifically axonal growth and neuronal apoptosis; or (3) the contribution of cytochrome P450-mediated metabolism to LC-PCB DNT. My central hypothesis is that LC-PCBs and their metabolites formed via human CYP2A6 and CYP2B6 alter neurodevelopment in primary neurons via CREB-dependent mechanisms. To test this hypothesis, I will be characterizing the in vitro DNT profile of human-relevant LC-PCBs and their metabolites, assessing how the metabolism of LC-PCBs by specific human CYPs influences DNT, and evaluating the role of CREB in LC-PCB DNT. This research will generate data critically needed to inform risk assessments of the potential for LC-PCBs to exert neurotoxic effects on the developing brain. Data from these studies will also provide novel mechanistic insights regarding the role of CREB and CYPS in LC-PCB DNT. Given the association of gain-of-function mutations in CREB with NDDs, and the well-known functional polymorphisms in human CYPs, data implicating CREB and/or CYP-mediated metabolism in LC-PCB DNT would suggest testable hypotheses regarding gene-environment interactions that influence NDD risk and possible dietary and/or pharmacological strategies for reducing LC-PCB DNT in at-risk populations.

NIH Research Projects · FY 2025 · 2023-09

OCT-based functional biomarkers for degenerative diseases of the photoreceptor-RPE-choroid neurovascular unit Abstract: Important diseases of the photoreceptor-RPE-choroid neurovascular unit include age-related macular degeneration (AMD), retinitis pigmentosa (RP), and other inherited retinal degenerations (IRDs). The number of people with AMD worldwide is nearly 200 million, and expected to approach 300 million over the next twenty years, while IRDs affect more than 5 million more. AMD is the leading cause of blindness in the industrialized world, and also the leading cause of blindness among people over the age of 60. Standard treatments exist for neither the more prevalent dry form of AMD nor IRDs, and key obstacles to discovering them are 1) limited understanding of the pathogenic steps leading to vision loss; and 2) lack of biomarkers for disease progression and recovery. In recent years our team has pioneered the emerging field of optoretinography (ORG) using adaptive optics (AO) and OCT. The ORG is an all-optical, noninvasive, objective measure of neural function in the retina, and has the potential to yield new functional biomarkers of retinal disease. Due to the cost and complexity of AO-OCT imaging systems and their subsequent scarcity, ORG measurements have been made only in small numbers of volunteers, mostly without retinal disease. We propose to develop a next-generation, proto-clinical ORG system and characterize its sensitivity, dynamic range, and spatial resolution. First, we will test a number of modifications to the protocols for imaging and signal processing, with a focus on improving the clinical utility of the method. Second, in healthy subjects and several patient populations we will measure ORG responses, along with other OCT-based measurements such as structural OCT imaging and OCTA. The patient populations to be studied are: 1) IRD patients including those with Stargardt’s disease and retinitis pigmentosa; 2) intermediate AMD patients with drusen, subretinal drusenoid deposits, and/or hyperreflective foci; and 3) late AMD patients with regions of geographic atrophy. The motivations for these patient populations are 1) to test hypotheses about the mechanisms underlying the ORG response, and 2) to demonstrate the ORG’s capacity to detect and quantify disease-related dysfunction. The proposed work will result in 1) a clinically useful ORG platform (also capable of structural and angiographic imaging), and 2) new knowledge that will permit us to design ORG- and OCT-based biomarkers for retinal disease that relate to mechanisms of action and have characterized sensitivity to photoreceptor dysfunction.

NIH Research Projects · FY 2025 · 2023-09

PROJECT SUMMARY/ABSTRACT Prior studies demonstrate that 25-50% of adolescents with mild traumatic brain injuries (mTBIs) have substantial mental health sequelae during their recoveries. Moreover, 31-78% of children with TBI who have new or worsening mental health concerns are not receiving appropriate mental health care, with racial, ethnic, and economic inequities existing in post-TBI management and outcomes. While a substantial number of children experience ongoing or worsening mental health concerns in the first three months after mTBI, there are no validated prognostic tools to assess risk of ongoing or worsening mental health concerns in these patients. Our objective is to develop and validate a clinical tool to predict mental health sequelae in adolescents after mTBI. We will conduct a multicenter, prospective observational study in six PECARN (Pediatric Emergency Care Applied Research Network) emergency departments (EDs). We will enroll a derivation cohort (n=1512) at four sites and a validation cohort (n=1080) at two sites. Patients will be enrolled in EDs and have follow-up evaluations at 1 to 2 weeks, 1 month, and 3 months after the index ED visit. The primary outcome is a composite self-report measure of new or worsening mental health sequelae (based on minimal clinically important difference) 1 to 3 months after mTBI as measured by the Generalized Anxiety Disorder-7 and the Patient Health Questionnaire-8. We will also measure unmet mental health care needs, defined as not receiving any mental or behavioral health care in patients with new or worsening anxiety or depression. We selected sites with large numbers of children with mTBI and a high proportion of children from diverse backgrounds. Our study has the potential to impact the health and wellbeing of injured children worldwide. The results will be immediately significant, affecting both clinical practice, guidelines, and policy. The study will be conducted at PECARN sites, with the PECARN data coordinating center, and a centralized mental health outcome core, leveraging existing resources and investigators with multicenter, pediatric emergency care clinical prediction tool development experience.

NIH Research Projects · FY 2024 · 2023-09

PROJECT SUMMARY/ABSTRACT We propose to evaluate a novel αvβ6 integrin molecularly targeted theranostic approach for the detection and treatment of metastatic cancers. The αvβ6 integrin is a cell surface receptor that is low or undetectable in normal adult epithelium but is widely expressed on numerous carcinomas. There is a statistically significant association between the high expression of the αvβ6 integrin, distant spread, and poor survival. We previously developed an αvβ6 -Binding Peptide (BP) with nanomolar affinity and high selectivity for the integrin αvβ66. Our prior clinical data demonstrates that using [18F]-αvβ6 -BP PET/CT imaging we can detect both primary tumors and metastases. PET images showed low background uptake in normal brain, lungs, liver, and osseous skeleton which are common sites of metastatic disease. Furthermore, sub-centimeter metastases to these organs were detected using [18F]αvβ6 -BP PET/CT. We further developed our αvβ6-BP into a novel theranostic pair, [68Ga]Ga DOTA-5G and [177Lu]Lu DOTA-ABM-5G in which [68Ga]Ga DOTA-5G is being developed as a diagnostic and [177Lu]Lu DOTA-ABM-5G is being developed as a radiotherapy. We now propose a prospective clinical trial to test the [68Ga]Ga DOTA-5G and [177Lu]Lu DOTA-ABM-5G in patients with metastatic disease. Patients will undergo [68Ga]Ga DOTA-5G PET/CT scans to confirm eligibility for the [177Lu]Lu DOTA-ABM-5G therapy and follow up [68Ga]Ga DOTA-5G PET/CT scans to evaluate treatment response. We hypothesize that a) [68Ga]Ga DOTA-5G will detect lesions in patients with metastatic cancers b) the theranostic pair [68Ga]Ga DOTA-5G/ [177Lu]Lu DOTA-ABM-5G will be safe and well tolerated; and c) a therapeutic response will be achieved with a single dose of [177Lu]Lu DOTA-ABM-5G. This proposal leverages the complimentary expertise of Dr. Sutcliffe to develop and translate the theranostic agents, and Dr. Foster to treat patients and interpret images for treatment response. The ability of the Sutcliffe lab to generate the theranostic agents in this study for patients treated at UC Davis maximizes existing resources and a well-established successful collaboration between the PIs, increasing the speed of translation of these theranostic agents from research to Phase II trials and ultimately, a standard of care option for patients with metastatic disease. Given the role of the αvβ6 integrin receptor in the processes of invasion and metastasis, αvβ6 is a very attractive target for the detection and treatment of metastatic cancers and could have broad impact across multiple cancers. [68Ga]Ga DOTA-5G will more accurately and non-invasively detect disease and the promising pharmacokinetic profile of the theranostics proposed suggests that off-target toxicity of this [177Lu]Lu DOTA-ABM-5G therapy is not expected. This poses a major improvement over current treatments that are known to cause bone marrow toxicity, hepatobiliary toxicity, and cardiotoxicity. Collectively, this proposal will significantly help transform cancer treatment for patients with advanced disease.

NIH Research Projects · FY 2024 · 2023-09

PROJECT SUMMARY/ABSTRACT Despite significant efforts to identify genes important in human neurodevelopment and disease, a large proportion of genes and variants remain undiscovered. Duplicated parts of the genome are largely understudied due to historical errors in the reference and bioinformatic pipelines that filter reads mapping to multiple locations in the genome. With the recent publication of a complete telomere-to-telomere human genome, genes and variants can be more effectively assayed across complex loci, but modified computational approaches are necessary. The proposed study will leverage diverse expertise in functional genomics and human genetics to test the hypothesis that a subset of human duplicated genes both contribute to neurological features and cause disorders exclusive to modern-day humans. Duplicated genes have previously been shown to play a role in early brain development and are enriched at genomic hotspots where recurrent copy-number variants are associated with neurodevelopmental disorders. Starting with a comprehensive list of thousands of human duplicated genes, functions of a subset of genes expressed during human corticogenesis will be tested using CRISPR knockout of orthologs and expression of human paralogs in zebrafish to determine their effects on general morphology, synaptic function, and brain development. The ability to test tens to hundreds of genes in parallel and conservation of basic developmental processes—such as neural proliferation, axonal guidance, and synaptogenesis—make zebrafish an ideal model to test these genes. Second, a genetic screen will be performed in human population cohorts to identify conserved duplicated genes. Since standard methods filter variants across many complex genomic loci, an improved bioinformatics approach leveraging short-read data will be devised and optimized using available sequencing benchmarks. Further, conserved genes will be screened for de novo and rare variants in autistic individuals using published datasets. Leveraging this multifaceted approach will enable systematic assessment of duplicated genes and their putative roles in human neurological traits and disorders. The zebrafish toolkit will be generally applicable to assaying functions of additional (non-duplicated) genes important in brain development, while the improved bioinformatics approach will enable additional screens of duplicated genes in other disease cohorts. This project will not only provide important insights into what it means to be human, but also it has the capability to discover missing genetic risk and elucidate the etiology of complex genetic neural traits and disorders.

NIH Research Projects · FY 2025 · 2023-09

PROJECT SUMMARY/ABSTRACT Despite significant efforts to identify genes important in human neurodevelopment and disease, a large proportion of genes and variants remain undiscovered. Duplicated parts of the genome are largely understudied due to historical errors in the reference and bioinformatic pipelines that filter reads mapping to multiple locations in the genome. With the recent publication of a complete telomere-to-telomere human genome, genes and variants can be more effectively assayed across complex loci, but modified computational approaches are necessary. The proposed study will leverage diverse expertise in functional genomics and human genetics to test the hypothesis that a subset of human duplicated genes both contribute to neurological features and cause disorders exclusive to modern-day humans. Duplicated genes have previously been shown to play a role in early brain development and are enriched at genomic hotspots where recurrent copy-number variants are associated with neurodevelopmental disorders. Starting with a comprehensive list of thousands of human duplicated genes, functions of a subset of genes expressed during human corticogenesis will be tested using CRISPR knockout of orthologs and expression of human paralogs in zebrafish to determine their effects on general morphology, synaptic function, and brain development. The ability to test tens to hundreds of genes in parallel and conservation of basic developmental processes—such as neural proliferation, axonal guidance, and synaptogenesis—make zebrafish an ideal model to test these genes. Second, a genetic screen will be performed in human population cohorts to identify conserved duplicated genes. Since standard methods filter variants across many complex genomic loci, an improved bioinformatics approach leveraging short-read data will be devised and optimized using available sequencing benchmarks. Further, conserved genes will be screened for de novo and rare variants in autistic individuals using published datasets. Leveraging this multifaceted approach will enable systematic assessment of duplicated genes and their putative roles in human neurological traits and disorders. The zebrafish toolkit will be generally applicable to assaying functions of additional (non-duplicated) genes important in brain development, while the improved bioinformatics approach will enable additional screens of duplicated genes in other disease cohorts. This project will not only provide important insights into what it means to be human, but also it has the capability to discover missing genetic risk and elucidate the etiology of complex genetic neural traits and disorders.

NIH Research Projects · FY 2025 · 2023-09

ABSTRACT Prenatal exposure to viral or bacterial infections during pregnancy is associated with an increased risk of offspring neurodevelopmental disorders, including autism and schizophrenia. Gestational biomarkers indicate that the maternal immune response is the critical link between maternal infection and altered offspring neurodevelopment. However, our ability to mitigate the deleterious impact of maternal infection on offspring brain development is severely restricted by our limited mechanistic understanding of the underlying neurobiological changes. Although preclinical rodent models have provided foundational evidence of alterations in brain and behavioral development resulting from MIA exposure that mirror some changes in human disorders, translational limitations provide a need to extend this program of research into a species more closely related to humans. Nonhuman primates (NHPs) provide the closest model to human development, sharing similarities in placental structure and pregnancy physiology, maternal-fetal interface, gestational timeline, fetal and postnatal brain development, and complex social behavior and cognition. Our laboratory has developed the first viral-mimic based NHP MIA model and demonstrated that MIA-exposed NHPs exhibit alterations in brain and behavioral development implicating selective vulnerability to socioemotional amygdala-prefrontal circuitry. Here we propose to leverage the entire biorepository of brain tissue from previous NHP MIA models to determine, at the single cell level, the transcriptomic, cellular, and connectomic alterations triggered by prenatal immune challenge. We have developed a novel pipeline for a genes-to-circuitry approach that maximizes the yield of information from this precious tissue resource. Here, we will target key brain regions in the amygdala-prefrontal network mediating socioemotional behaviors implicated in human neurodevelopmental and mental health disorders at two critical age time points for the pathophysiology of mental illness: juvenile (18 month) and adolescent (4 year). We will generate single-nuclei transcriptomic profiles and quantify differentially expressed genes (DEGs) in specific cell types (Aim 1), spatially map and quantify high-priority transcripts in specific cell types and within-cell transcriptomic colocalization (Aim 2), and map spatial distribution of synaptic composition, receptors, and direct inputs onto specific cell types (Aim 3). These data, in combination with the extensive, longitudinal characterization of offspring brain and behavioral development, build a comprehensive picture of MIA-induced changes in NHP brain circuitry, toward the ultimate goal of identifying pathways of vulnerability and critical periods for novel, targeted interventions and biotherapeutics to reduce the number of children adversely affected by prenatal exposure to maternal infection.

NIH Research Projects · FY 2024 · 2023-09

ABSTRACT Prenatal exposure to viral or bacterial infections during pregnancy is associated with an increased risk of offspring neurodevelopmental disorders, including autism and schizophrenia. Gestational biomarkers indicate that the maternal immune response is the critical link between maternal infection and altered offspring neurodevelopment. However, our ability to mitigate the deleterious impact of maternal infection on offspring brain development is severely restricted by our limited mechanistic understanding of the underlying neurobiological changes. Although preclinical rodent models have provided foundational evidence of alterations in brain and behavioral development resulting from MIA exposure that mirror some changes in human disorders, translational limitations provide a need to extend this program of research into a species more closely related to humans. Nonhuman primates (NHPs) provide the closest model to human development, sharing similarities in placental structure and pregnancy physiology, maternal-fetal interface, gestational timeline, fetal and postnatal brain development, and complex social behavior and cognition. Our laboratory has developed the first viral-mimic based NHP MIA model and demonstrated that MIA-exposed NHPs exhibit alterations in brain and behavioral development implicating selective vulnerability to socioemotional amygdala-prefrontal circuitry. Here we propose to leverage the entire biorepository of brain tissue from previous NHP MIA models to determine, at the single cell level, the transcriptomic, cellular, and connectomic alterations triggered by prenatal immune challenge. We have developed a novel pipeline for a genes-to-circuitry approach that maximizes the yield of information from this precious tissue resource. Here, we will target key brain regions in the amygdala-prefrontal network mediating socioemotional behaviors implicated in human neurodevelopmental and mental health disorders at two critical age time points for the pathophysiology of mental illness: juvenile (18 month) and adolescent (4 year). We will generate single-nuclei transcriptomic profiles and quantify differentially expressed genes (DEGs) in specific cell types (Aim 1), spatially map and quantify high-priority transcripts in specific cell types and within-cell transcriptomic colocalization (Aim 2), and map spatial distribution of synaptic composition, receptors, and direct inputs onto specific cell types (Aim 3). These data, in combination with the extensive, longitudinal characterization of offspring brain and behavioral development, build a comprehensive picture of MIA-induced changes in NHP brain circuitry, toward the ultimate goal of identifying pathways of vulnerability and critical periods for novel, targeted interventions and biotherapeutics to reduce the number of children adversely affected by prenatal exposure to maternal infection.