Pennsylvania State Univ Hershey Med Ctr

NIH Research Projects · FY 2026 · 2023-06

ABSTRACT Hispanic/Latino (H/L) children and adolescents have a 40% higher death rate than non-Hispanic whites (NHW) after correcting socioeconomic factors. Recently, we identified a highly increased incidence of deletion of one IKZF1 allele in H/L children, which provided a biological rationale for the worse prognosis of B-cell acute lymphoblastic leukemia (B-ALL) in this population. The very high incidence (29%) makes the deletion of one IKZF1 allele the most frequent genetic alteration that confers adverse prognosis in B-ALL in H/L children. Thus, understanding the biological basis for resistance to chemotherapy and developing novel therapies to treat B-ALL with deletion of one IKZF1 allele will reduce the health disparity in survival for Hispanic/Latino children with ALL. The IKZF1 gene encodes a DNA-binding protein, IKAROS, which functions as a transcriptional regulator. Our published data showed that, in B-ALL, IKAROS phosphorylation by the oncogenic Casein Kinase II (CK2) reduces IKAROS DNA-binding affinity and abolishes its function as a transcriptional regulator. Our new preliminary data suggest that CK2 impairs the ability of IKAROS to repress transcription of two key genes involved in the thiopurine pathway, resistance to 6-mercaptopurine (6-MP) treatment, and relapse: TPMT and NT5C2. Treatment with CK2-specific inhibitor, CX-4945, restores IKAROS-mediated repression of the TPMT and NT5C2 genes in primary B-ALL cells from H/L children. In vitro treatment of B-ALL with CX-4945 in combination with 6-MP exhibits a strong synergistic cytotoxic effect. Our overall hypothesis is that overexpression of CK2 in high-risk B-ALL of H/L children impairs IKAROS’ ability to transcriptionally repress the genes that regulate resistance to treatment with 6-MP, and that CK2 inhibition will restore IKAROS function as a transcriptional repressor of these genes, resulting in increased sensitivity to 6-MP. This hypothesis will be tested in vivo, using various biological models. We will define the mechanism through which IKAROS regulates the thiopurine pathway in primary B-ALL cells from H/L children. The effect of increased CK2 expression on thiopurine pathway and IKAROS function will be analyzed in patient-derived xenografts (PDXs), generated from B-ALL from H/L children. Finally, we will establish the therapeutic efficacy of combination treatment with CK2 inhibitor (CX-4945) and 6-mercaptopurine (6-MP) in a preclinical model of B-ALL from H/L children. Results of the project will provide a novel insight into the regulation of the thiopurine pathway and help develop novel treatments for high-risk B-ALL in H/L children and reduce childhood cancer health disparities.

NIH Research Projects · FY 2026 · 2023-05

Project Summary The gut microbiota is a critical determinant of human health and development. Diets rich in the refined sugars glucose and fructose can modulate gut microbial abundance and metabolism thereby increasing disease susceptibility. However, the mechanisms governing microbial responses to dietary sugar in the gut remain poorly understood. We have demonstrated that host dietary sugar consumption silences expression of a colonization factor called Roc in Bacteroides thetaiotaomicron, an abundant gut bacterium associated with lean, healthy individuals. We determined that a conserved transcription factor is necessary to synthesize Roc and regulates additional bacterial factors that mediate critical host-microbial interactions, including an immunomodulatory protein and fucose utilization genes. We hypothesize that the activity of this transcription factor is governed by a putative intracellular metabolite that is differentially synthesized according to host diet composition and dramatically reduced upon sugar-rich diet consumption. We have identified critical molecular players governing the activity of this transcription factor and propose to 1.) elucidate the mechanisms governing its activation, 2.) determine how dietary sugar consumption silences its activity, and 3.) characterize the consequences of host sugar-rich diet consumption on microbial product synthesis in vivo. We believe that this work will reveal a conserved pathway that is disrupted by host sugar consumption resulting in aberrant gut microbial activities. Furthermore, this work has uncovered a molecular target for modulation of microbial abundance and metabolism in the host and can be exploited for rational manipulation of the gut microbiota to treat diseases.

NIH Research Projects · FY 2026 · 2023-05

PROJECT SUMMARY Stroke is a leading cause of long-term disability in U.S. and worldwide. Post-stroke cognitive impairment (PSCI), a common sequela after stroke, is a decisive determinant of the quality of life for stroke survivors. Clinical studies have indicated that PSCI is common in both young and old stroke patients, even in cases of relative mild stroke and victims with successful thrombolysis and endovascular reperfusion therapies. However, the underlying mechanisms of PSCI remains poorly understood and no FDA approved treatment is available for PSCI. In this application, we propose to investigate the roles and therapeutic potential of DKK3 in PSCI and the underlying mechanisms using an experimental stroke model of transient middle cerebral artery occlusion (MCAO) followed by reperfusion. Previous studies (including ours) have demonstrated that mice subjected to transient MCAO developed long-term cognitive deficits that correlate with secondary damage to the hippocampus. Based on our promising pilot data, we hypothesize that DKK3 plays an important role not only in acute brain damage but also in secondary hippocampal damage and thereby represents a novel promising therapeutic target for treating cognitive impairment after ischemic stroke. First, we will determine the tempo-spatial regulation of DKK3 (and miR-125a) expression in the normal and ischemic brains at different time points after stroke, and evaluate the efficacy of early treatment versus delayed treatment by intranasal administration of recombinant DKK3 protein to ameliorate acute stroke injury and to improve long-term neurologic and cognitive outcomes after ischemic stroke (Aim1). Next, we will determine the mechanistic roles of DKK3 in neuropathology with the focus on the hippocampal mechanisms of PSCI after ischemic stroke (Aim 2). To test this hypothesis, the loss-of-function and gain- of-function experiments will be performed, in which conventional and conditional DKK3 knockout mice and functional reconstitution study with recombinant DKK3 via intranasal drug delivery will be utilized. Based on pilot data, we further hypothesize that ischemic stroke induces increase of miR-125a expression hence down-regulates DKK3 expression in the hippocampus, which contributes to PSCI induced by transient MCAO (Aim 3). We will determine the DKK3-dependent effects of miR-125a inhibition to improve PSCI after ischemic stroke. Both young adult and aged mice will be studied. The proposed studies may reveal previously unappreciated mechanisms underlying PSCI and provide a novel therapeutic approach to improve neurologic and cognitive outcomes following ischemic stroke.

NIH Research Projects · FY 2025 · 2023-04

ABSTRACT Autism Spectrum Disorder (ASD) is a lifelong, neurodevelopmental disorder with significant consequences for individuals and their families. Over 95% of individuals with ASD have at least one comorbid psychiatric, neurologic, or physical health condition. Many also struggle with other developmental challenges such as intellectual disabilities and/or complex communication needs and often receive inadequate care. These unmet needs, which often begin in childhood, can have negative impacts on the health and well-being of those with ASD through their lives. The incidence rates of adverse outcomes have steadily increased for youth with ASD over the last decade. While consistent management of mental health and other comorbid conditions might reduce episodes of emergencies, the heterogeneity of ASD and health disparities resulting from race/ethnicity, gender, and/or socioeconomic status (SES) potentially impede the effectiveness of evidence- based treatments for ASD in the “real world”. In addition, the increasing practice of excessive polypharmacy of psychotropic medication is likely to have negative impact on utilization and effectiveness of non- pharmacological treatments, though little is known about its specific effect on uptake as well as, intensity and duration of behavioral therapies. The COVID-19 pandemic has increased stress on individuals with mental health illnesses, and, subsequently, incidence of psychiatric crises. Understanding treatment utilization patterns and variations across demographic, geographic and socioeconomic factors will provide guidance to improve the efficiency of health care delivery and quality of health services for individuals with ASD. Leveraging large, national, longitudinally constructed Medicaid claims databases (2008-2022), we will examine the interplay between polypharmacy of psychotropic medications and key ASD behavioral therapies among youth aged 5-26 with ASD and evaluate their impact on preventing behavioral health crises. In particular, we will (1) examine the trends and multi-level factors associated with utilization and quality of behavioral health and pharmacological treatments among youth with ASD; (2) examine the trends and health disparities in the rates of psychiatric adverse events among youth with ASD; (3) examine the impact of quality of behavioral health treatments and their interplay with polypharmacy on psychiatric adverse events among youth with ASD. The proposed study will provide a comprehensive assessment of the quality of ASD-related treatments and services in a real-world setting and shed light on disparities in service use, quality of care, and health outcomes, particularly in regards to the risk of behavioral health crises among youth with ASD. This information will be valuable to families exploring treatment options, as well as to providers in determining treatment options to maximize benefit. Identifying barriers to accessing services and implementation of evidence-based practice will help guide policies at the payer, state, and national levels.

NIH Research Projects · FY 2025 · 2023-04

Project Summary/Abstract: Electronic cigarette (e-cig) usage is on the rise, particularly among youths; however, their potential for harm is not understood, complicating development of informed regulatory strategies. The lack of data on e-cig related harm is, in part, due to the lack of specific biomarkers for exposure to e-cig aerosols. We found that e-cig aerosol contains highly reactive free radicals that can cause oxidative damage to the user. Free radicals can damage numerous cellular pathways possibly contributing to the progression of cancers and other diseases. Detection of e-cig free radicals can be accomplished by trapping with spin traps (e.g., DMPO) and analysis by electron paramagnetic resonance (EPR) spectroscopy. Preliminary research with EPR shows that free radicals produced in the e-cig aerosol by e-liquid solvents, propylene glycol (PG) and glycerin (GLY), common to all e-cigs, display unique structural characteristics. Our objective is to identify these free radical structures and utilize their unique structure to develop an e-cig specific biomarker of exposure. The specific aims of the proposed research are: (Aim 1) To determine the structures of the free radicals produced by PG and GLY in e-cigs; (Aim 2) To determine the primary targets of and adducts formed from free radical assault in the tissue of e-cig exposures in rodent models and possible metabolites formed from the these radical adducts in the serum of e-cig exposed rodent. This project represents an important research direction where a chemical/biological approach can inform tobacco regulatory science. As such, an important aspect of this application it to extend my background in areas relevant to translational science in addition to providing specific training in new biomarker-relevant research areas including metabolomics and free radical structural identification. To this end, my training will occur through a series of courses, relevant mentorship, and practical experience, each geared to ensure my transition to an independent researcher in the fields of biomarker development and regulatory science. Coursework, mentorship, conference participation, and practical training/experience will be completed during the K99 phase. During this phase, Aim 1 of the research plan will be completed and Aim 2 will be initiated (for completion during the R00 phase of the award). To accomplish the research aims, advanced pulsed EPR and mass spectroscopy approaches will be utilized for radicals produced by PG and GLY in e-cig aerosols. In a mouse exposure model, free radical exposure and targets of attack in the lung will be determined using a novel in vivo DMPO/anti-DMPO antibody approach. This will allow for the identification of specific protein and DNA adducts by TOF-MS. A post-DMPO exposure study will consist of an untargeted pairwise metabolomics approach to look at changes in metabolite profiles before and after e-cig exposure. By leveraging the unique structures of e-cig produced free radicals and their targets of attack in the lung, biomarkers of exposure specific to e-cig aerosols can be identified and used to develop regulatory strategies aimed at reducing harm from e-cig exposure.

NIH Research Projects · FY 2026 · 2023-03

Project Summary/Abstract How neurons guide their processes to the correct binding partner is a complicated task, but is critical during development and recovery from injury. It involves the highly coordinated action of many cytoskeletal proteins and their binding partners within the growth cone at the tips of extending neurites, as they feel their way through the neuropil. There is a lot known about the signaling pathways that regulate neurite outgrowth and turning, but the details of how molecular structures come together to achieve growth cone behavior are still unclear. This project will initially focus on the structure of bundled cofilactin filaments (cofilin-decorated F-actin) in situ, and how this novel filament structure and fascin cross-linking determine filopodial dynamics. Here we propose that filopodial behavior is governed partially by the transition from the fascin cross-linked to a cofilin cross-linked filaments, that makes filopodial bundles more pliable. Experiments are focused around three aims: 1) to study the high-resolution structure of fascin- and cofilin-linked actin bundles to determine their impact on actin structure, 2) to determine how changes in fascin and cofilin concentration regulate filopodial dynamics and structure, and 3) determine how LIMKI and SSH1 form the core of a bidirectional regulatory mechanism for regulating actin architecture via tuning the phosphorylation state of Ser3 on cofilin.

NIH Research Projects · FY 2026 · 2023-02

ABSTRACT There is tremendous interest in cannabis (as smoked marijuana or CBD- or THC-dominant extracts) as a therapeutic modality for a variety of health indications (including chronic pain). Given the complexity of cannabis, however, we have little insight into its mechanisms of action in complementary and integrative health approaches. Specifically, there is a prevailing notion that the 100+ cannabinoids and the various terpenoids/flavonoids that comprise cannabis act in concert to create an “Entourage Effect”. A comprehensive analysis is required to better understand the potential of cannabis agents as complementary medicines. We herein propose a novel artificial intelligence-driven approach to address this gap in our knowledge. Not surprisingly, a natural product (e.g., cannabis) that is active in an organism typically works because it acts like endogenous ligands or those known to the organism. We hypothesize that deconstructing ligand structures into specific fragments will allow us to identify targets that bind endogenous tergets containing such fragments. Moreover, we believe that disparate compounds acting in concert will maximally engage selective pathways. We have developed an artificial intelligence (AI)-driven platform, DRIFT (drug-target identification based on chemical similarity), to map ligand compounds (cannabinoids and terpenoids) to molecular targets. Thus, we can illuminate involved cellular pathways, and predict physiological response. We will use DRIFT to profile compounds in a number of different cannabis extracts (e.g., high in CBD, CBG, or THC) with varying analgesic properties to identify therapeutic combinations and their relevant targets. We will undertake three specific aims. In Specific Aim 1, use DRIFT for massive mapping of cannabis constituents to corresponding target proteins. Then, in Specific Aim 2, we will extend the DRIFT platform by evaluating binding affinities between metabolites and their proteins using a new neural network paradigm (NeuralDock). We will use text mining techniques to mine compound-protein relationships from PubMed. Finally, in Specific Aim 3, we will experimentally validate the outputs of the DRIFT platform to predict mechanisms of cannabis on pain. We will test the AI-based results using traditional pharmacology tools and a variety of preclinical animal models of pain. These same models will then be employed to test mechanisms through the complementary use of agonists, antagonists, inhibitors and (where appropriate) gene knockouts to validate mechanisms. When these studies are successful, we will have validated DRIFT as a new and valuable AI tool for studying natural products. Moreover, we will provide important insights into the growing use of cannabis in complementary and integrative health.

NIH Research Projects · FY 2026 · 2023-02

PROJECT SUMMARY/ABSTRACT A rotator cuff tendon tear affects up to 64% of adults, resulting in muscular changes that drive functional disability and poor clinical outcomes. Rotator cuff tears are associated with muscle atrophy and fatty infiltration (accrual of fat in the muscle bulk), non-homogenous distribution of fatty infiltration within the muscle, and reduced muscle strength and contractile function. Changes to muscle morphology likely have a negative effect on the muscle’s architecture (number and organization of muscle fibers) and the mechanical properties dictating the muscle’s overall force-generating capacity. However, little is known about the mechanisms linking muscle structure and function or how these associations change over time, exposing a notable knowledge gap. Establishing the structure-function associations of muscle after rotator cuff tear will expose the targets and key time points for design of treatments to improve outcomes for patients with a rotator cuff tear. Thus, the objective of this project is to determine the spatial and temporal changes to muscle morphology, architecture, and multi-scale mechanics after rotator cuff tear and surgical repair, and develop a predictive model to identify the mechanisms driving biomechanical function. To achieve this goal, longitudinal experimental assessments will be performed in an established rabbit model; a novel computational model will also be developed based on experimental structural measurements of muscle to further probe the mechanisms of muscle function. Specifically, this project aims to: 1) Establish the structure-function relationships of muscle after rotator cuff tear and surgical repair; and 2) Develop and test a predictive model to examine mechanisms driving reduced biomechanical function after rotator cuff tear. A rotator cuff tear will be surgically introduced in rabbits by blunt dissection of the supraspinatus tendon, with assessments at 2, 4, 6, and 8 weeks after injury; surgical repair will be performed 8 weeks after injury, with a final assessment 8 weeks after repair. At each time point, muscle morphology and architecture, and intra- versus extra-cellular location of lipid accumulation will be quantified using magnetic resonance imaging (MRI). Multi-scale mechanics will be assessed at whole muscle and single fiber levels. Structure-function associations and how these associations change over time will be established. Structural measures of muscle morphology, architecture, and mechanics will be used to develop and test a predictive model to probe the mechanistic role of each structural parameter on biomechanical function. Models will be used to determine the mechanism and timing that should be targeted by treatment to improve functional outcomes. This work is significant because it will establish the structure-function relationship of muscle after rotator cuff tear and surgical repair and identify the mechanisms underpinning biomechanical function. Little work has examined both spatial and temporal changes to muscle structure and examined their influence on function, making this work innovative. Study outcomes will expand our understanding of the impact of rotator cuff tear and drive development of novel treatments for the rotator cuff tear patient population.

NIH Research Projects · FY 2026 · 2023-01

ABSTRACT Acquired von Willebrand Syndrome (AvWS) characterized by the loss of high molecular weight multimers (HMWMs) of von Willebrand factor (vWF) is often associated with nonphysiologic blood flows. AvWS is found in patients with severe aortic stenosis (AS) or continuous-flow left ventricular assist devices (cf-LVADs). Interestingly, this hemostatic abnormality associated with severe AS is fully corrected on the first day after surgery. For AvWS associated with cf-LVADs, it disappears quickly after removal of the device, strongly suggesting that the device itself is responsible for the syndrome. It is widely believed that the supraphysiologic shear stress and/or long exposure time in severe AS and cf-LVADs are responsible for the loss of HMWM. Although the destruction of HMWM is believed to be a combination of mechanical and enzymatic cleavage, the complete mechanism still remains unclear. Also notable is that AvWS is rarely observed in pulsatile blood flow devices. There is a need to understand the degradation mechanism of vWF and exposure time especially when nonphysiologic blood flows are expected. The objective of this proposal is to characterize the degradation of HMWM under fully controlled laminar through turbulent blood flow conditions in terms of power density (energy dissipation rate per unit mass) and clinically relevant exposure times. The central hypothesis is that the degradation of HMWM of vWF is a time-sensitive mechanoenzymatic event that occurs primarily under turbulent flow conditions by exposing the ADAMTS13 to cleavage sites on vWF. Based on our previous publications as well as evidence in the literature, we believe that the majority of vWF multimer degradation is ADAMTS13 mediated. However, we recognize that there are other potential sinks and sources of vWF including adsorption to the surfaces of a device, binding to platelets, and release of vWF from ⍺-granules of activated platelets. We will test and account for these sinks and sources in these experiments. Successful completion of the proposed study will allow us to i) determine the relationship between nonphysiologic blood flow and exposure time involved in the loss of HMWM in cf-LVADs, ii) evaluate the degradation mechanism of HMWM, and iii) characterize the mechanism in terms of mechanoenzymatic, mechanistic, and enzymatic sensitivity through comprehensive vWF biology. Ultimately, the prospective model is expected to be a tool for device design optimization leading to a next-generation blood pump and better clinical outcomes for patients with AvWS.

NIH Research Projects · FY 2026 · 2022-12

Project Summary/Abstract Dilated cardiomyopathy (DCM) is the second most common cause of heart failure world-wide, and inherited forms of DCM make up 30% of non-ischemic cases. MYH7, which encodes beta-cardiac myosin (M2β), is one of the more commonly mutated genes and is the molecular motor that powers contraction in ventricular cardiomyocytes. This proposal is focused on examining the structural and functional impact of DCM mutations in human M2β, with an overall goal of determining molecular mechanisms of contractile defects and developing a foundation for therapeutic strategies. The force, velocity, and power generating capacity of muscle is related to the ability to recruit myosin molecules in the thick filament to interact with actin in the thin filament of the muscle sarcomere. The recruited myosin molecules generate force by utilizing a conserved ATPase cycle in which myosin generates a power stroke while interacting with actin. Cardiac myosin can exist in the auto-inhibited state with slow ATP turnover (super relaxed state, SRX) in which head-head and head-tail interactions prevent it from interacting with actin (interacting heads motif, IHM) or the uninhibited state (disordered relaxed state, DRX) that is readily available to produce force. The recruited myosin also impacts the calcium sensitivity of the myofilaments because myosin binding cooperatively activates the actin thin filament. We will test the central hypothesis that DCM mutations impair systolic contraction in the heart by altering the intrinsic force producing ability of individual cardiac myosin molecules, stabilizing the SRX/IHM state, and/or altering cooperative activation of the actin thin filament. In the first Aim we will examine the impact of the DCM mutations on the myosin ATPase cycle, duty ratio, and formation of the SRX state. The structural impact of the mutations will be examined by using a FRET biosensor that monitors the myosin power stroke and another FRET sensor that examines IHM state formation. Electron microscopy will also be used to evaluate the formation of the IHM state which will be directly compared to the fluorescence spectroscopy and biochemical analysis. Aim 2 will examine the impact of DCM mutations on the single molecule mechanical properties of human M2β, including step size and load-dependent detachment, using a load clamped optical trap. In Aim 3 we will utilize a computational model of muscle contraction to predict how the parameters measured in Aims 1&2 will impact ensemble force, velocity, and power. We will then directly examine the impact of the mutations on the force generating properties by incorporating the human M2β constructs into DNA-based “designer” thick filaments, and examining their ability to interact with regulated thin filaments in a calcium dependent manner. The force, velocity, and power measurements will be performed in the “designer” thick filaments, which contain native thick filament-like geometric spacing of myosin molecules. Overall, the completion of the specific aims of this proposal will enhance our understanding of the molecular mechanisms of disease pathogenesis in DCM and provide a foundation for developing therapies for treating DCM.

NIH Research Projects · FY 2026 · 2022-09

PROJECT SUMMARY Delaying the onset of Alzheimer’s disease and related dementias (ADRD) by even a few years could amass substantial improvements in quality of life for individuals and families, independent living, and the cost of specialized healthcare. Positive cognitive outcomes associated with physical activity interventions conducted in middle-aged and older adults have increased the promise that primary prevention of ADRD may be achievable via lifestyle change. It may take years to decades of adherence to health promoting behaviors to realize appreciable gains in cognitive health span. Critically, while lifestyle interventions are supportive of behavior change in the short-term, longer-term maintenance of cognitive health promoting behaviors (i.e. continued enactment of these behaviors 6+ months after completion of the intervention) has proven difficult to achieve. The current project will leverage an mHealth approach using commercially-available technology (smart phones, wrist-worn activity monitors) to promote long-term maintenance of light intensity physical activity (LIPA) in middle-aged adults at increased risk for ADRD (adults with obesity). Following a health education session with a certified exercise physiologist, our approach will deliver the other intervention components: adaptive daily step goal setting (both arms), self-monitoring (both arms), and interim goal setting every 2-3 hours (treatment arm-only), on study smart phones, over the course of participants daily lives. We aim to demonstrate that the interim goal setting manipulation to the intervention can lead to greater long-term maintenance of LIPA by keeping LIPA goals active in the focus attention during each day of the intervention period and increasing LIPA self-efficacy through regular goal attainment. Goal maintenance (indicated by self- monitoring frequency) will be quantified via participant interactions with custom-integrated Fitbit wrist-worn activity monitors and the Mobile Monitoring of Cognitive Change (M2C2) component of the forthcoming NIH Mobile Toolbox. Fitbits will be configured with a custom-designed clockface that displays accumulated step counts when a ‘check my steps’ button is pressed. Logging and timestamping these interactions will allow for quantification of self-monitoring frequency. We propose that development of these adherence-promoting mechanisms (goal maintenance and self-efficacy) can act as a self-regulatory scaffold from which long-term health promotion gains can be realized. The current project will: 1) demonstrate the ability of interim goal setting to engage our proposed adherence-promoting targets (Aim 1); 2) test the efficacy of the interim goal setting manipulation to increase short- and long-term LIPA maintenance following the intervention through goal maintenance and increased self-efficacy (Aim 2); 3) examine how variation in cognition influences the ability of the intervention to engage goal maintenance (Aim 3); 4) explore the long-term salutary effects of the intervention on cognitive (change in ADRD risk) and physical health (change in weight, V02-max; Aim 4).

NIH Research Projects · FY 2024 · 2022-09

Abstract: The current COVID-19 pandemic creates many challenges for individuals and health care systems in the United States. To make matters worse, the pandemic is occurring in the midst of a public health crisis of opioid use disorder and overdose during pregnancy. Between 2010 and 2017, the incidence of maternal opioid-related diagnoses increased from 3.5 to 8.2 per 1000 hospital live births per year. Maternal opioid use during pregnancy is associated with increased risks of prolonged hospital stay, placental abruption, poor fetal growth, preterm labor, premature delivery, stillbirth, neonatal abstinence syndrome, and maternal death. State responses regarding maternal opioid use during pregnancy prior to the COVID-19 pandemic have varied widely and included: 1) creation of funded drug-treatment programs specifically for pregnant women, 2) priority access to state-funded treatment programs, 3) mandated reporting and drug screening by healthcare professionals, and 4) criminalization of opioid use during pregnancy or grounds for commitment. However, these policies are unlikely to remain static, especially during the current COVID-19 pandemic. The need for social distancing has led to policy changes related to treatment of opioid use disorder more generally, such as easing methadone dispensing rules and making it easier for patients to initiate buprenorphine treatment from an opioid treatment center. These changes are not mandatory, however, and system-level gains in access to medication assisted treatment (MAT) may be eliminated as the pandemic subsides. In addition, little is known about the extent to which policies related to maternal opioid use during pregnancy are changing during the COVID-19 pandemic, or the extent to which such changes affect maternal and child healthcare treatment, outcomes and costs. Thus, the aims of this study are to 1) examine how state policies related to maternal opioid use during pregnancy have changed in response to the COVID-19 pandemic, and 2) examine the effects of COVID-19-related changes in state policies regarding maternal opioid use on patterns of healthcare service use, maternal and child outcomes, and healthcare costs. By thoroughly examining the policy responses to the COVID-19 pandemic and linking these data to detailed claims data describing healthcare service use of women who use opioids during pregnancy and their newborns, we will be able to provide critical data to providers, insurers, health systems, and policymakers as they design treatment processes and policies to provide adequate pregnancy and substance use care to this vulnerable population to mitigate these two public health crises.

NIH Research Projects · FY 2026 · 2022-09

ABSTRACT Individuals with Type 1 Diabetes (T1D) are susceptible to repeated episodes of hypoglycemia, which can result in impaired awareness of hypoglycemia (IAH). As a consequence, IAH contributes to diminished recognition of the need for external glucose and the blunting of counter-regulatory responses that are required to increase blood glucose. This can be a very risky situation for T1D individuals with IAH, possibly leading to coma and in some circumstances, death. This can be a very risky situation for T1D individuals with IAH, possibly leading to coma and in some circumstances, death. Many T1D studies exclude IAH individuals because of confounding issues. Therefore, the National Institute for Diabetes and Digestive and Kidney Diseases (NIDKK) has issued two RFAs to form the IAH Consortium that will investigate factors which contribute to the heterogeneity of restoration of awareness of hypoglycemia in adult T1D individuals with IAH. RFA-DK-21-020 and RFA-DK-21- 036 request applications for Clinical Centers and the sole Biostatistics Research Center (BRC), respectively. The objectives of the IAH Consortium will be to • determine if the most up-to-date management strategies using diabetes technology to optimize HbA1c while minimizing hypoglycemia can restore awareness of hypoglycemia and improve counter-regulatory responses in individuals with T1D and IAH • identify the magnitude and duration of time in range, time spent in hypoglycemia or other metrics for continuous glucose monitors that are associated with restoration of awareness of hypoglycemia • determine the association of current or newly developed self-reported measures of IAH with counter- regulatory responses to elucidate the heterogeneity in restoration of hypoglycemia awareness The Department of Public Health Sciences at The Pennsylvania State University College of Medicine proposes to serve as the BRC for the IAH Consortium, and will pursue seven specific aims. Specific Aim 1: Collaborate on the development and conduct of studies, and lead statistical analyses Specific Aim 2: Lead the Consortium in developing measures of self-reported IAH Specific Aim 3: House a Central Laboratory Specific Aim 4: Lead the Consortium in stakeholder engagement Specific Aim 5: Develop data management and website processes Specific Aim 6: Maintain regulatory compliance Specific Aim 7: Provide communication and project management support

NIH Research Projects · FY 2024 · 2022-09

ABSTRACT Systemic lupus erythematosus (SLE) is a heritable autoimmune disease that primarily affects young females. SLE symptoms can be very heterogeneous, which posts great challenges in early diagnosis. In its advanced stages, SLE can lead to multiple organ failures and even fatality. Early diagnosis is critical for controlling and mitigating the symptoms and improving the quality of life. Genome-wide association studies of SLE to date have identified >150 loci. Yet, the causal variants remain elusive for most loci. There is great interest to integrate datasets from diverse human populations to further empower discovery, refine causal variants, and improve the risk prediction accuracy. In this proposal, we seek to aggregate available GWAS summary statistics from a myriad of autoimmune diseases. We will develop improved meta-analysis methods that effectively integrate data from multiple traits and ancestries. We will also develop better fine-mapping methods that can integrate statistical approaches and experimental validations. Finally, we will develop more accurate genetic risk scores using datasets from multiple ancestries and traits, and combine them with commonly used lab values for improved disease risk predictions. We will release useful software packages implementing these methods to benefit studies of other traits and maximize the impact of the proposed research.

NIH Research Projects · FY 2024 · 2022-09

Project Summary/Abstract Although breast cancer is the most frequently diagnosed cancer in the world, recently, the survival rate has greatly improved. With this improved survival rate, there is a growing population of obese, older adult breast cancer survivors facing unique challenges and late effects on brain health. Breast cancer survivors suffer cognitive impairments before, during, and post- treatment; more than one third experience persistent cognitive impairment lasting decades. Anxiety is the most common psychological symptom among cancer survivors, and frequently unrecognized and undertreated in health care settings. These survivors are also at increased risk of Alzheimer’s disease or dementia. There is unanimous consensus that physical activity and alcohol consumption are key modifiable behaviors to improve the physical and mental health of cancer survivors. Further, the American Cancer Society promotes increasing physical activity, and avoiding alcohol to manage treatment-related cognitive impairment. Although the benefits of physical activity for cancer survivors are well-established, the evidence is predominantly derived from guidelines-based, moderate-to-vigorous-intensity physical activity. However, survivors often do not meet guidelines for moderate-to-vigorous-intensity physical activity, do not adhere to physical activity interventions set at this intensity, and enjoy and prefer lighter-intensity activities. Much less is known about the effects of light-intensity physical activity on cognitive function and anxiety symptoms, and the 2018 United States Physical Activity Guidelines Advisory Committee Scientific Report identified the need to determine the role and contribution of light-intensity activity to diverse health outcomes as an overarching research need. Additionally, less is known about the effects of physical activity interventions on changes in alcohol consumption. Recent findings indicate physical activity may unintentionally increase alcohol consumption among cancer survivors. Increasing alcohol consumption places survivors at increased risk of recurrence, and may confound the positive effects of physical activity. Therefore, in the process of evaluating the effects of light-intensity physical activity on brain health outcomes among obese, older adult breast cancer survivors, the effects on alcohol consumption should be evaluated as well, to avoid unintended consequences. The light- intensity physical activity-breast cancer survivors (LIPA-BCS) trial addresses multiple knowledge gaps, with a primary and secondary focus on improving cognitive function and anxiety symptoms. The proposed work builds on this existing randomized controlled trial that randomizes obese, older adult breast cancer survivors 1-10 years post-breast cancer treatment, to either 15 weeks of light-intensity physical activity, or usual care. I will take advantage of this existing clinical trial to quantify and describe the effects on cognitive function, anxiety symptoms, and observe associated changes in alcohol consumption. To achieve this aim, we will assess cognitive function, anxiety symptoms, and alcohol consumption both pre- post, and utilizing ecological momentary assessments in 14-day bursts throughout the 15 week intervention. Through this research experience, and proposed post-doctoral fellowship, I will learn about the conduct of high quality human clinical trials among obese, older adult cancer survivors, and receive mentored training in ecological momentary assessments, biostatistics, and becoming an independent investigator in exercise oncology.

NIH Research Projects · FY 2025 · 2022-09

Penn State Cancer Institute’s (PSCI) PROMISE program (Penn State Research training in Oncology and Medicine to Inspire Student Engagement) is a holistic approach to inspire, attract, and retain cancer researchers for the 21st century biomedical workforce. The program engages STEM college undergraduates and medical students in an immersive program that provides authentic individual and team research experiences, demonstrating first-hand how new cancer research discoveries are translated into clinical and societal practice. Additional program workshops provide support training in both professional and personal skills needed for sustained research careers. PROMISE emphasizes the interdisciplinary nature of modern cancer research. We will recruit motivated and talented students from a variety of disciplines and create an environment in which cancer research is approachable and accessible for all participants. The PROMISE design is built upon a relational pedagogical model with peers, near-peers, and faculty to promote participants’ sense of belonging in, and contributing to, the cancer research community. The long-term goals of PROMISE are to instill in participants a comprehensive understanding of cancer as a disease (how it is prevented, controlled, and treated), while stimulating interest in oncology careers and providing skills needed for durable research careers. To achieve these goals, Specific Aim 1 will provide integrated training in basic, clinical, and populations science approaches through mentored research experiences. The PROMISE: Research for Life training focus will show students how interdisciplinary research is conducted to solve current oncology problems and to save lives through effective cancer management. Under the experienced mentorship of PSCI scientists and clinicians, the cohort will learn the language, techniques, strategies, and concepts used every day by cancer researchers. Specific Aim 2 will enhance trainee effectiveness through intensive support activities and rigorous program outcomes evaluations. Workshop activities will provide the opportunity to experience peer acceptance, identify role models, and overcome potential roadblocks to a successful research career. Events with stakeholders highlight how advances in clinical oncology and societal practice are driven by research discoveries. Specific Aim 3 will create a cohort from varied disciplines and inspire them to pursue advanced training in cancer research and/or clinical oncology. STEM students who do not traditionally enter biomedical careers will learn basic cancer biology and realize they can contribute to advancing cancer health outcomes through collaborative research. Peer and near-peer mentors will support self-identification as a cancer researcher and faculty will provide authentic role models for future career paths. Participation in PROMISE will be life-changing for our cohort, as students will have experienced the critical importance of modern cancer research to saving the lives of individuals, a perspective that will inspire them to pursue cancer research careers and sustain them during their advanced training years.

NIH Research Projects · FY 2025 · 2022-09

Project Summary Detecting various compositionally similar but structurally distinct glycans present in heterogenous mixtures has traditionally been challenging due to the limitations facing current glycomics approaches. These limitations have hindered our understanding of the availability and abundance of individual glycans in the mammalian gut environment, which is replete with diverse mixtures of microbial, mammalian, and plant-derived oligo- and polysaccharides. Moreover, the intestinal glycan composition is a primary driver of gut microbiome composition and metabolism, which represents an increasingly important human health determinant, and necessitates a deep understanding of the glycomic-microbial interface to identify important biological interactions and putatively develop glycan-derived therapeutics to target specific microbial activities. Therefore, new tools are necessary to detect and measure the relative abundance of individual glycan substrates present in the heterogenous mixtures prepared from biological sources such as mammalian intestinal contents. We have harnessed the glycan detection machinery employed by dominant members of the gut microbiota to detect, measure and isolate individual glycan substrates present in heterogenous mixtures extracted from the mammalian intestine. Herein, we demonstrate robust, specific, and scalable approaches by which engineered microbes report the presence of individual glycan substrates with incredible sensitivity. Furthermore, we demonstrate that this approach can accurately measure the abundance of individual glycans present in mixtures across wide linear ranges and that the specificity and sensitivity of these measurements can be tuned by modifying particular microbial glycan utilization genes. Finally, we demonstrate that microbially-encoded glycan-binding proteins can be used to isolate individual target glycans from mixtures for downstream compositional and structural determination. We propose to 1.) develop arrayed libraries of distinct gut microbial species, each engineered to report the presence of unique target glycans, 2.) develop a rapid glycan isolation pipeline to purify individual substrates of interest for downstream structural and functional characterization, and 3.) develop genetically modified microbial strains with enhanced sensitivity or target specificity. In addition to offering a rapid and inexpensive alternative to quantifying known glycans, we believe that further development of these tools will reveal the presence and abundance of previously undetectable glycans and dramatically enhance our understanding of the interactions between gut microbes and their mammalian hosts.

NIH Research Projects · FY 2025 · 2022-09

Project Summary/Abstract Neonatal abstinence syndrome (NAS) is characterized by central nervous system hyperactivity that occurs when an infant experiences withdrawal from maternal opioid use at birth. Rates of NAS in the United States have skyrocketed amidst the opioid epidemic. Assessment and clinical management of NAS rely on subjective symptom scales, which may contribute to the prolonged hospital stays and poor neuro- developmental outcomes associated with NAS. Enhancing our knowledge about the molecular factors that regulate the biologic response to opioid withdrawal in developing infants will provide an opportunity to create objective clinical tests and novel therapies for this significant medical problem. To date, there is no biologic tool to determine the necessary morphine dose in withdrawing infants, or to predict which infants will experience neurodevelopmental delays. This is partly due to the fact that prior studies have largely focused on the molecular response to opioid administration in adults, rather than the response to opioid withdrawal in infants. Serum levels of certain micro-ribonucleic acids (miRNAs) are impacted by opioid administration in adults (e.g., let-7a, miR-146a, miR-192). These short, non-coding nucleic acids also regulate neurogenesis, neuronal progenitor cell (NPC) maturation, and cell survival by repressing target messenger RNAs in the Argonaute complex. My sponsor, Dr. Steve Hicks, MD, PhD has pioneered the use of salivary miRNAs as non-invasive markers for pediatric neurodevelopmental conditions. Most salivary miRNAs are derived from exosomes, which can arise from the cranial nerves that densely innervate the oropharynx. My preliminary data shows that several “opioid-responsive” miRNAs are perturbed in the saliva of infants with NAS relative to healthy infants. Further, I have developed an in vitro system of opioid withdrawal utilizing human NPCs that displays dose- related perturbations in miR-146a along with disruptions in NPC fate. Based on these findings, the central hypothesis of my fellowship application is that morphine withdrawal alters miRNA expression in the developing brain, which impairs neuronal maturation by mRNA translation via Argonaute binding. I also hypothesize that salivary levels of exosomal miRNAs from infants with NAS will be directly related to both the maximal dose of morphine required for symptom control, and neurodevelopmental outcomes at six months. I will test these hypotheses in two specific aims. First, I will perform a longitudinal cohort study of 50 infants with NAS to determine whether salivary miRNA levels within brain-related exosomes can be measured at admission to predict the maximum morphine dose required for symptom control, and again at discharge to predict neurodevelopmental outcomes at six months (Aim 1). Second, I will transfect miRNA mimics in my in vitro NPC design of opioid withdrawal, and assess the mechanism by which miRNAs impact the response to opioid withdrawal with immunocytochemistry, single-cell RNA sequencing, and an Argonaute pull-down assay (Aim 2).

NIH Research Projects · FY 2025 · 2022-09

PROJECT SUMMARY The slow growing nature of most prostate cancers (PCa) provides multiple windows of opportunity to block or delay disease progression to significantly reduce patient suffering and mortality. This is especially true following the failure of radical prostatectomy (RP) and radiation therapy (RT) of PCa prior to androgen deprivation therapy (ADT; i.e., medical or surgical castration). Biochemical recurrent PCa is sensitively indicated by rising blood prostate specific antigen (PSA). ADT is not curative and causes many serious adverse effects. Our goal is to develop Angelica gigas Nakai (AGN, Korean Angelica) root ethanolic extract or its constituent(s) as a safer and practical modality for PCa interception akin to secondary prevention to delay ADT or avoid it entirely. We hypothesize that (1) the PCa interception efficacy of AGN and its pyranocoumarins in animal models is extendable into human PCa patients, given sufficient AGN dosage and exposure duration; (2) multiple mechanisms including immune surveillance and suppression of inflammation contribute to the clinical cancer interception activity. The scientific premise is based on the presence of novel active pyranocoumarin compounds distinct from those in soy, tea, fish oil, raspberries, mushrooms, and cannabis and reported broad spectrum anti- cancer efficacy in animal cancer models. Moreover, we have demonstrated a) Oral bioavailability and favorable pharmacokinetic (PK) metrics in rodents and in humans; b) Cytochrome P450 (CYP) 2C19 and 3A4 first-pass conversion of pyranocoumarins decursin (D) and decursinol angelate (DA) to their metabolite decursinol (DOH); c) Proficient tissue retention of decursinol in mouse target prostate; d) Animal modeling of AGN and decursinol showing independence of the androgen receptor axis, avoiding side effects of ADT drugs and making blood PSA a reliable readout for cancer burden; e) AGN enhanced immune and decreased inflammatory gene signatures in an animal PCa model; and f) A single AGN dose in human subjects increased natural killer [NK] mRNA signature and decreased IL-8 chemokine mRNA in their peripheral blood mononuclear cells. We propose 3 specific aims in post-RP and post-RT patients with rising plasma PSA that is “clean” (due to prostate already removed) and indicative of the biochemical recurrent PCa burden. Aim 1. Characterize pyranocoumarin PK dose response proportionality to AGN supplement and food effects in 12 patients to probe any metabolic ceiling and minimize food-herbal interaction. Aim 2. Evaluate safety and recurrent PCa (PSA)-interception efficacy of twice daily AGN supplement in a Phase I/II trial with 36 patients. Aim 3. Measure acute- and repeated-dose PK metrics, CYP 2C19 and 3A4 metabolizer status and changes of immune and inflammation biomarkers to correlate to PSA outcomes. Impacts: The safety information and a favorable PSA response will guide future randomized control trials to prevent or treat PCa and other cancers. The novel knowledge on PK dose response, repeated dose PK behavior, CYP pharmacogenetics and the immune biomarkers not only fills in critical gaps for AGN supplements in cancer patients, but also is exportable to disease prevention and therapy beyond oncology.

NIH Research Projects · FY 2025 · 2022-08

ABSTRACT Large scale genetic datasets have revolutionized human genetic research. In the past decade, genome-wide association studies have identified numerous genetic variants associated with various complex traits. These discoveries have informed new biology and led to novel therapeutics. Yet, most studies focused on European samples and most of the studies were conducted by pooling both sexes together. Sex-specific genetic architecture in diverse populations remains largely elusive. As the next step, consortia efforts have begun to aggregate datasets from diverse non-European populations. Meta-analysis is often employed in large consortium studies There is a compelling need to develop methods that can effectively conduct trans-ancestry meta- analysis. To this end, we will develop a series of innovative methods to improve the power for association mapping, the precision of fine mapping and the accuracy of polygenic predictions in trans -ancestry analysis. Resulting methods will be applied to a series of heart, lung, and blood related traits, in collaboration with a number of consortia studies. This is a continuation of our strong track record of method development in statistical genetics. We will implement the methods into a few widely used tools, including RVTESTS, SEQMINER, and RAREMETAL, which will not only benefit our own study but also other researchers in the field.

NIH Research Projects · FY 2026 · 2022-08

Project Summary/Abstract The overarching goal of my research program is to understand how the human genome is faithfully replicated when it is constantly damaged by covalent modifications that can alter its coding properties. A focus is on translesion DNA synthesis (TLS), the predominant DNA damage tolerance (DDT) pathway utilized in humans to replicate damaged DNA. TLS utilizes specialized DNA polymerases that can accommodate an array of DNA damages, albeit with lowered fidelities, and promotes cell survival in the face of DNA damage by allowing replication of a damaged genome to continue. However, tight regulation is critical as aberrant TLS can selectively propagate cells with increased mutagenesis and chromosomal rearrangements, which can lead to cancer. Furthermore, because TLS promotes cell survival after exposure to DNA damaging agents, aberrant TLS can also afford cancer cells the ability to overcome common chemotherapies that aim to trigger cell death by acutely damaging DNA. Hence, TLS is a promising candidate for targeted cancer co-therapy. Despite the established links between human health and TLS, the progression and regulation of TLS is unclear and many key gaps persist in our fundamental knowledge that cloud our understanding of the contribution of TLS to genetic inheritance and carcinogenesis. For example, human DDT can occur by at least three pathways but what is the interplay between TLS and other DDT pathways? My long-standing interests and vast expertise in human DNA damage repair and replication and my established ability to utilize a multi-faceted approach integrating molecular biology, biochemistry, and biophysics provide a unique opportunity to fill these gaps. This proposal will address the progression and regulation of TLS by tackling two broad areas that are each a cornerstone of our fundamental understanding of TLS and DDT in general; 1) Activation of TLS and; 2) The interplay between DDT pathways. To do so, we design and apply unique, quantitative approaches that utilize kinetic techniques and can be adapted to many biological scenarios. The PCNA sliding clamp is an essential DNA replication factor and is monoubiquitinated at sites of DNA damage by the Rad6(Rad18)2 complex. PCNA monoubiquitination is imperative for human TLS and activates this critical pathway by recruiting TLS factors. Area 1 will investigate how the activity of the human Rad6(Rad18)2 complex is regulated to efficiently monoubiquitinate PCNA at DNA damage sites, activating TLS. Human DDT can occur by at least three pathways including TLS and all emanate from a common intermediate. Area 2 will investigate the interplay between TLS and other DDT pathways and decipher functional relationships between key events in each pathway. This proposal investigates TLS in broad contexts that consider the complexities and dynamics of cellular environments and will significantly advance our fundamental understanding of how the human genome is faithfully replicated in the face of DNA damage and inspire new avenues of research in cancer etiology and treatment.

NIH Research Projects · FY 2024 · 2022-08

Project Summary Heart failure is a leading cause of mortality and morbidity in patients with diabetes and obesity, yet much remains unknown regarding the molecular events whereby metabolic disease causes myocardial dysfunction. At the cellular level, oxidative stress and inflammation are considered hallmarks of myocardial impairment. The overarching hypothesis for this F31 fellowship proposal is that the stress response protein regulated in development and DNA damage 1 (REDD1) plays a maladaptive role in the pathogenesis of heart disease by augmenting the development of oxidative stress and inflammation in cardiomyocytes. Preliminary data support that REDD1 expression is enhanced in the heart of mice fed a Western (i.e., high fat, high sucrose) diet and in cardiomyocyte cultures exposed to the saturated fatty acid palmitate. Recent studies from our laboratory demonstrate a key role for REDD1 in the development of diabetes-induced oxidative stress. More specifically, REDD1 acts via a GSK-3β-dependent signaling axis to suppress the nuclear factor erythroid-2-related factor 2 (Nrf2) antioxidant response in the retina of diabetic mice. Additionally, REDD1 was found to promote inflammatory signaling in macrophages by direct sequestration of inhibitor of κB (IκB), leading to nuclear factor kappa B (NF-κB) activation. A role for REDD1 in activation of these two non-conventional signaling axes for Nrf2 repression and NF-κB activation has never been interrogated in the heart. Nor is it know if REDD1 contributes to the development of cardiac dysfunction. To test the hypothesis, I will pursue an experimental protocol involving model systems ranging from intact mice to cardiomyocyte cell cultures. Aim 1 will investigate the mechanism whereby consumption of a Western diet promotes transcriptional upregulation of REDD1 in the heart. Aim 2 will investigate the role of REDD1 in the development of oxidative stress and inflammation in cardiomyocytes. Aim 3 will determine the impact of REDD1 deletion on the development of cardiac dysfunction. In addition to my continued graduate training in molecular biology and cellular physiology at Penn State College of Medicine, this fellowship will provide key training opportunities with experts in cardiovascular dysfunction. Specifically, I will learn to properly culture and manipulate adult ventricular cardiomyocytes and develop the technical expertise in echocardiography to assess the impact of REDD1 on cardiac function in transgenic mice. With respect to outcomes, the project will not only expand my skills and systems of analysis beyond those of my primary Sponsor, but will also potentially identify and characterize a unifying regulatory mechanism whereby metabolic disease limits the endogenous antioxidant response and upregulates inflammation in heart. Identification of such a mechanism is significant because it will validate new targets for the development of preventative and/or therapeutic interventions aimed at addressing the molecular basis of heart failure.

NIH Research Projects · FY 2024 · 2022-08

PROJECT SUMMARY Hepatitis B virus (HBV) is a DNA virus belonging to the hepadnavirus family and is responsible for chronically infecting over 250 million individuals worldwide, resulting in nearly a million annual deaths due to severe liver diseases such as cirrhosis and hepatocellular carcinoma. Unfortunately, anti-viral treatments fail to cure HBV infection due to the tenacity of covalently closed circular DNA (cccDNA), which is the template for viral transcription and subsequent translation of all viral products necessary to establish and sustain HBV replication. Much is learnt about HBV replication events post cccDNA production, however little is known about the intracellular trafficking steps that are required for the initial cccDNA production to establish productive infection. My PhD dissertation research aims to define the role of the viral capsid in endocytic trafficking during HBV infection (Aim 1). We have identified a series of HBV capsid mutants which have a unique phenotype in that during infection, they do not lead to productive infection (i.e., no cccDNA formation), however cccDNA can form during transfection via intracellular amplification, which bypasses the viral entry steps during infection, indicating that there is a block in the viral entry steps during infection with the mutant viruses that are not required during transfection. We plan to monitor post-entry trafficking events of WT and mutant HBV (Aim 1.1) and will further define the role of the viral capsid in directing endosomal trafficking through use of capsid inhibitors that induce a similar effect to our capsid mutants (Aim 1.2). These capsid inhibitors are in active clinical development; therefore, it is crucial that we understand what entry steps are modulated by them. Addressing questions related to HBV entry has been made possible by the identification of the viral entry receptor, hNTCP, however its expression cannot confer susceptibility to HBV infection in HEK293 cells, which can support cccDNA formation during transfection but cannot support cccDNA formation during infection, phenocopying the effects of the HBV capsid mutations and inhibitors. The second portion of this project will focus on defining the role of host cells in HBV trafficking during infection (Aim 2). We will address the endocytic trafficking of HBV in HEK293-hNTCP cells to identify where the block(s) in infection occurs, rendering these cells non-susceptible (Aim 2.1). To help identify the host determinants, beyond the entry receptor hNTCP, that may play a role in HBV entry, we have designed a split GFP-based HBV reporter system that will be used to isolate a subpopulation of HepG2-hNTCP cells that are highly susceptible to HBV infection (Aim 2.2) and perform RNA sequencing analysis to identify differentially expressed genes between susceptible vs. non-susceptible cells. These proposed studies will increase our understanding of HBV entry and identify the essential endocytic trafficking steps that lead to productive infection. Altogether, the successful accomplishment of my dissertation will have profound impacts on (1) understanding the essential events of HBV endosomal trafficking and entry and (2) identifying the HBV capsid and novel host determinants that are critical for HBV entry and productive infection.

NIH Research Projects · FY 2025 · 2022-07

SUMMARY The overall incidence of HPV+ head and neck squamous cell carcinoma (HNSCC) has exhibited an increasing trend over the last few decades, and has now in the U.S. overtaken cervical cancer as the most common site of HPV related cancer. The majority of patients present with advanced-stage HNSCC are without a clinical history of pre-malignancy. Because there is a paucity of pre-cancer clinical samples the studying the steps of cancer progression that would provide an understanding of the carcinogenic we have developed an in vitro progression model. Interestingly, HPV+ patients compared to HPV negative (HPV-) patients have an improved overall prognosis and greater disease-free survival, attributed partly to a better response to radio- and/or chemotherapy. The mechanisms and natural history of oral HPV infection and carcinogenic progression are poorly understood. Men and women infected with human immunodeficiency virus (HIV) have higher rates of HPV16 infection, longer persistence of HPV16, increased incidence of HNSCC, poorer prognosis, as well as more recurrences after definitive therapy. There have been substantial changes in the burden of cancer affecting HIV-infected individuals in the U.S. during the period spanning the introduction of highly active anti- retroviral therapy (HAART). Scaling-up of HAART therapy has dramatically increased the life expectancy of people living with HIV (PLHIV). As a result of these temporal changes, non-AIDS-defining cancers have come to comprise the majority of cancers in HIV-infected persons during the HAART era. Among these emerging malignancies, HPV-associated cancers of the oral cavity/pharynx increased significantly following an AIDS diagnosis. Most patients with HIV/HPV co-infections on HAART therapy have reduced risk of developing HPV-associated cancers such as cervical cancer but have a higher risk of developing HPV-associated oral cancers the risk is suggested to be due to longer life expectancy as a results of HAART therapy. But it is unclear why HAART therapy may have different effects on HPV+ HNCSCC versus HPV+ cervical cancers. Our overarching hypothesis is that HAART therapy affects the progression of HPV+ HNC. This increases the numbers of PLHIV who have persistent oral HPV infections that progress to HNCSCC, poorer prognosis, and death. We propose to focus on three areas to test this hypothesis. Specific Aim 1. Analyze HAART induced differences of HPV carcinogenic propensity in oral epithelial cells. Specific Aim 2: Investigate the effect of HAART on carcinogenic markers and metabolic pathways associated with HPV+ HNSCC. Specific Aim 3: Study the effect of HAART on glycolysis versus mitochondrial respiration.

NIH Research Projects · FY 2026 · 2022-07

PROJECT SUMMARY/ABSTRACT Traumatic burn injuries generate excruciating pain resulting from tissue and nerve damage and the accompanying exaggeration of inflammation. Necessary wound care procedures such as debridement, burn excision, grafting, and mobilization only add to this pain. For these reasons, pain from traumatic burn injury is notoriously difficult to manage; hence current reports suggest that burn-related pain is currently undertreated. Traumatic injuries have a dramatic increase in pain sensitivity (hyperalgesia) and accelerated development of tolerance to opioid analgesics compared with non-traumatic injuries. Unfortunately, the mechanisms driving increased sensitivity and tolerance following trauma are unclear. Furthermore, over half of all burn survivors develop chronic pain, making the transition from acute to chronic pain a significant concern following burn injury. Research has primarily focused on the innate immune response to injury as a leading candidate; however, the recent inclusion of female subjects has uncovered sex-specific differences in immune-modulated pain sensitivity. These findings have led to reconsideration of the influence of sex on pain processing and whether the body of male-only research is truly applicable. It is well known that females are more likely to report more severe pain and to experience chronic pain following such injuries; however, the mechanisms behind these sex-specific differences are currently unknown. Additionally, stress-induced adaptations in the neuroendocrine system are believed to play an essential role in the pathogenesis of acute and chronic pain outcomes. Dysregulation of the hypothalamic-pituitary-adrenal and sympathoadrenal axes following traumatic injury has been shown to reduce the activity of descending pathways that modulate endogenous pain inhibition and opioid analgesia. We hypothesize that the immune response involved in the pathogenesis of pain following traumatic burn injury is regulated in a sex-specific manner through involvement of the neuroendocrine system. Exploiting these sex- specific differences may be key to understanding how to treat pain generated from traumatic burn injuries. Sex- specific genes, hormones, and signaling mechanisms may shed light on novel targets that have been previously overlooked, giving great hope for future sex-specific interventions. Accordingly, the proposed research program will address this critical gap in our understanding by systematically investigating the role of sex-specific immune reactivity and neuroendocrine responses in an animal model of full-thickness thermal burn injury. This program of research will lead us closer to understanding mechanisms that make pain from traumatic burn injuries so challenging to treat and identify sex-related drivers behind them. Our goal over the duration of this award is to identify sex-specific mechanisms underlying the accelerated development of tolerance and hyperalgesia and to provide insight to potential targets for sex-specific interventions to treat acute pain and prevent the transition to chronic pain. Conduct of these studies will build a robust program of research positioned to investigate any promising targets we may uncover.