Cincinnati Childrens Hosp Med Ctr

NIH Research Projects · FY 2024 · 2021-09

Healthcare workers are consistently exposed to hazards with widespread consequences for themselves, patients, and the organization. In a 2018 Bureau of Labor Statistics report, healthcare continues to have one of the highest rates of non-fatal occupational injury relative to other industries. Musculoskeletal injuries, violence, falls, and needle-sticks are most common. It is essential that healthcare systems employ effective and user- friendly surveillance to allocate resources for injury prevention. However, there is extensive evidence that reported injury rates significantly underestimate the true risk, and existing injury surveillance systems do not capture other adverse outcomes that impact health and performance. Other subjective health complains (SHCs), such as stress and fatigue, are major reasons for sickness and absence from work. Furthermore, near-misses have long been integrated into injury surveillance of high-risk industries such as aviation, nuclear power, and the military, but healthcare has yet to systematically assess the incidence of these important sentinel events as part of injury prevention. Finally, despite federal law and institutional policies promoting employee safety, employees are reluctant to report injuries due to time constraints, symptom self-management, peer pressure, perceived `normalcy' of injury, and fear of reprisal. Valid data on the incidence of near-misses, injuries, and time trends are essential for identifying high-risk areas and deploying resources to mitigate harm. This study will address the Healthcare & Social Assistance sector and the cross-sector programs of Musculoskeletal Health and Healthy Work Design & Well-Being by addressing the weaknesses in current injury surveillance methods using a proactive approach. Our long-term goal is to improve healthcare injury surveillance systems and significantly reduce work-related injuries. Our specific aims are to: integrate passive injury reporting with active injury reporting to improve detection of worker harm; leverage unit-based risk metrics, passive and active injury reports to continuously monitor the work environment; and develop an early-warning system that will more accurately trigger interventions to decrease the risk of and prevent worker injuries. Randomly selected patient care providers will verbally record injuries, near-misses, and SHCs on a mobile application for 800 two-week periods. Using advanced statistical methods (e.g. machine learning) and data visualization we will aggregate injury data and unit-based metrics to produce an injury risk identification and prediction model; develop a digital heat map to identify high-risk units in near real-time, in a real-world setting, and provide hospital leaders with notifications with established risk-specific mitigation strategies. We build upon our R21OH010035 project, which established the feasibility of active injury surveillance and documented its superiority to current surveillance practice. In response to PAR-18-812, our project can shift current practice toward a more rigorous integration of injury surveillance and prevention.

NIH Research Projects · FY 2025 · 2021-09

Neurodevelopmental processes are shaped by dynamic interactions between genes and environments. Maladaptive experiences early in life can alter developmental trajectories, leading to harmful and enduring developmental sequelae. Pre- and postnatal hazards include maternal substance exposure, toxicant exposures in pregnancy and early life, maternal health conditions, parental psychopathology, maltreatment, and excessive stress. To elucidate how various environmental hazards impact child development, it is imperative that a normative template of developmental trajectories over the first 10 years of life be established based on a sufficiently large and demographically heterogeneous sample of the US population. To accomplish this, the Healthy Brain and Child Development (HBCD) Consortium has been formed to deploy a harmonized, optimized, and innovative set of neuroimaging (MRI, EEG) measures complemented by an extensive battery of behavioral, physiological, and psychological tools, and biospecimens to understand neurodevelopmental trajectories in a sample of 7,200 mothers and infants enrolled at 27 sites across the United States (US). The HBCD Study will carry out a common research protocol under direction of the HBCD Consortium Administrative Core (HCAC) and will assemble and distribute a comprehensive and well-curated research dataset to the scientific community at large under the direction of the HBCD Data Coordinating Center (HDCC). The overarching goal of the HBCD Study is to create a comprehensive, harmonized, and high-dimensional dataset that will characterize typical neurodevelopmental trajectories in US children and that will assess how biological and environmental exposures affect those trajectories. A special emphasis will be placed on understanding the impact of pre- and postnatal exposure to opioids, marijuana, alcohol, tobacco and/or other substances. To address these broad objectives, the sample of women enrolled will include: 1) a varied cohort that is representative of the US population; 2) pregnant woman with use of targeted substances (opioids, marijuana, alcohol, tobacco); and 3) demographically and behaviorally similar women without substance use in pregnancy to enable valid causal inferences. In addition, the HBCD Study will identify key developmental windows during which both harmful and protective environments have the most influence on later neurodevelopmental outcomes. The large, multi-modal, longitudinal, and generalizable dataset that will be produced for the first time by this study will provide novel insights into child development using state-of-the-art methods. The HBCD Study will inform public policy to improve the health and development of children across the nation.

NIH Research Projects · FY 2025 · 2021-09

ABSTRACT Asthma is a major public health problem in the United States, affecting 11 million children. Despite advances in asthma care, African Americans (AAs) are 4 times more likely to be hospitalized and 5 times more likely to die from asthma than European Americans (EAs). Several factors could be responsible for the observed asthma racial disparities including genetic and non-genetic factors. While epigenetics appear to serve as a critical biological switch between genetic vulnerability and socio-environmental exposures, limited studies are available that directly map the socio- environmental exposures with asthmatic epigenome/genome information. In addition, current approach do not leverage existing geospatial data such as environmental exposure and neighborhood socioeconomic conditions to improve asthma risk prediction. In this proposal, we will utilize comprehensive geocoding algorithms, novel statistical methods to integrate social, clinical, environmental, genetic, and epigenetic data into a composite score for asthma risk stratification and prediction. The overall objective of this research is to conduct genome-wide Methyl-Seq analysis and leverage existing well-phenotyped AA pediatric asthma cohort with extensive socio-environmental exposures and ancestry-tailored multi-ethnic genotyping array (MEGA) data from Cincinnati Pediatrics Repository to accurately determine and develop ancestry- specific asthma risk stratification and prediction models. The objective of this application is to undertake an epigenome-wide association study (EWAS), incorporating geocoded neighborhood- and individual-level socio-environmental predictors, and novel analytical strategies to create a composite risk score incorporating methylation risk score (MRS), ancestry, environmental exposures and social characteristics to predict asthma. We will accomplish these objectives through the following Specific Aims: 1) Develop an ancestry-specific methylation risk score (MRS) for asthma and test its association with socio-environmental exposures contributing to asthma risk. 2) Determine the mediation effects of MRS between genetic ancestry and asthma risk. 3) Develop a multivariable risk predictive model for asthma incorporating MRS, genetic ancestry, clinical, and socio-environmental risk factors. The proposed research is innovative because this will be the first time a MRS approach will be used to develop a population-based risk profile in asthmatics. The study will provide insights in the use of risk stratification for screening and targeted interventions. This work is significant because it can serve as a model to study the composite effect of MRS, ancestry, socio-environmental, and clinical risk factors on racial disparities in other well-documented common complex diseases beyond asthma.

NIH Research Projects · FY 2025 · 2021-09

Project Summary Polycomb group (PcG) proteins play critical roles in maintaining epigenetic memory of gene silencing in normal development and human diseases. PcG proteins function in two enzymatic multi-subunit complexes: Polycomb repressive complex 1 and 2 (PRC1 and 2). PRC1 deposits monoubiquitin to lysine 119 on histone H2A (H2Aub) whereas PRC2 methylates all states of lysine 27 on histone H3 (H3K27me1/2/3). How Polycomb domains are reprogrammed during mammalian preimplantation development remains largely unknown. Recent advances on low input epigenomic profiling techniques make it feasible to investigate chromatin dynamics in mammalian preimplantation embryos. The discovery that non-canonical H3K27me3 in oocytes can mediate germline DNA methylation-independent genomic imprinting has raised several important questions on Polycomb domain regulation in early development. For example, whether H2Aub follows a similar reprogramming dynamic as H3K27me3, what’s the role of PRC1/2 in regulating 3D chromatin, and whether PRC1/2 form a positive feedback loop to reinforce each other during preimplantation development. To address these questions, I have generated preliminary data showing that, in contrast to conventional view that H2Aub and H3K27me3 are largely co- localized, H2Aub and H3K27me3 undergo genome-wide distinct reprogramming dynamics after fertilization. In addition, H2Aub deposition by PRC1 is independent of PRC2 in oocytes and preimplantation embryos, suggesting a more critical role of PRC1 than PRC2 during this developmental window. Built on the unexpected observations, I propose to use a combination of low input epigenomics, bioinformatics, and rapid protein degradation approach to understand mechanisms and functions of PRC1/2 as well as the chromatin modifications they respectively deposit in mouse preimplantation development. In Aim 1 (K99 phase), I will identify mechanisms underlying the distinct reprogramming dynamics of H2Aub and H3K27me3 after fertilization. In Aim 2 (K99 phase), I will rapidly degrade PRC1 in zygotes to assess its impact on zygotic genome activation, PRC2 recruitment, and 3D chromatin structures. I will be trained by Drs. Yi Zhang (Boston Children’s Hospital/Harvard Medical School), Peter Park (Harvard Medical School), and Bin Gu (Michigan State University) to establish low input epigenomic profiling tools, biochemical assays, computational pipelines, and the rapid protein degradation technique. In Aim 3 (R00 phase), I will take advantage of the techniques and computational pipelines established during the K99 phase to study the role of variant PRC1 subcomplexes in preimplantation development. The NIH K99/R00 Pathway to Independence Award, together with the outstanding research environment at BCH/HMS will facilitate my completion of the proposed work and transition to an independent investigator. Collectively, completion of these aims will reveal mechanisms underlying PcG recruitment, define the role of PcG-mediated gene silencing in mouse preimplantation development, and uncover new paradigms on chromatin reprogramming during mammalian gamete-to-embryo transition.

NIH Research Projects · FY 2025 · 2021-09

PROJECT SUMMARY/ABSTRACT The number of infants exposed to opioids prenatally in the US remains high, with one infant born exposed to opioids every 15 minutes. Optimal treatment for exposed infants who develop neonatal opioid withdrawal syndrome (NOWS) has not been established, and randomized comparative effectiveness trials of medications for NOWS are critically needed. Cincinnati Children’s Hospital Medical Center (CCHMC) is uniquely suited to participate as a clinical site in such a multicenter randomized comparative effectiveness trial. The CCHMC Perinatal Institute is a large regional perinatal center, providing population-based clinical service for approximately 25,000 newborns each year. Intrauterine exposure to opioids in the Cincinnati region was approximately 26 per 1000 live births in 2020, with 10.3 per 1000 developing NOWS requiring pharmacologic treatment. The proposed PI for this study Stephanie Merhar, proposed alternate PI Scott Wexelblatt, and proposed follow-up PI Jennifer McAllister all have extensive experience in clinical care, quality improvement, and clinical research in the NOWS population. The Cincinnati team will provide significant intellectual contributions to the design of the study, recruit a large number of infants with NOWS to participate, and continue our tradition of excellent follow-up rates to retain infants in the study to age 2. The expertise and experience of the investigators and support staff, along with the facilities and population available for study at CCHMC, make CCHMC a strong site for this proposed study.

NIH Research Projects · FY 2025 · 2021-09

PROJECT SUMMARY: Comprehensive regulation of transcription is a major mechanism by which immature cells functionally determine their identity. The epigenetic factors responsible for this transcriptional regulatory activity are some of the most potent, and most commonly mutated, factors implicated in tissue homeostasis and disease. We understand the role of many epigenetic factors responsible for activating transcription, but we have a limited understanding of the factors that repress transcription associated with differentiation. Additionally, we know many of the epigenetic transcriptional regulators present on the chromatin before and after replication, but we have a limited understanding of how these factors interact during replication. The relative lack of understanding of how the replication machinery regulates transcription associated with cell identity is a key knowledge gap in the field of epigenetics. Our studies suggest that the Chromatin Assembly Factor 1 (CAF1) complex, functioning in its canonical role of replication-linked chromatin assembly, is the critical factor responsible for directly regulating fate-specific transcription during replication. CAF1 is a heterotrimeric protein complex responsible for facilitating histone H3/H4 heterodimer assembly at the replication fork during the S phase of cell cycle. Our lab focuses on the functional contributions of CAF1 through studying its p60 subunit: Chromatin Assembly Factor 1B (CHAF1B). We are using CHAF1B as a model for CAF1 function because of the abundance of tools and expertise we have generated to study CHAF1B and the overwhelming evidence that readouts of CHAF1B and the CAF1 complex are functionally identical. CHAF1B is highly expressed in uncommitted stem cells, and its expression decreases as cells differentiate. We recently reported that CHAF1B directly binds chromatin at promoters and enhancers of differentiation genes, repressing their expression by blocking transcription factor binding. Depletion of CHAF1B in independent models led to massive upregulation of gene activation caused by transcription factors flooding regions of chromatin previously occupied by CHAF1B. This led to our hypothesis: CHAF1B is a master regulator of cell identity through a novel role as transcriptional repressor of fate genes in immature cells. Our long-term goal is to understand how cells regulate transcription associated with cell fate throughout differentiation. Our short-term goal is to understand how the chromatin assembly machinery affects transcriptional repression of fate genes in immature cells. In this proposal we will study the functional roles of the factors responsible for CHAF1B direct binding to chromatin on transcription and phenotype (Focus 1), and the contributions of the DNA/protein complexes recruited by CHAF1B to the chromatin on transcription and phenotype (Focus 2). Discoveries about CHAF1B, the CAF1 complex, and the larger DNA/protein chromatin landscape it promotes, have the potential for driving sweeping changes in the way we understand functional epigenetics and treat disease.

NIH Research Projects · FY 2026 · 2021-09

Common variable immunodeficiency (CVID) affects approximately 10,000 people in the USA and is caused by an expanding array of genetic lesions. It is defined by international consensus criteria that may be summarized as loss of B cell function developing after infancy with or without a well-described constellation of autoimmune or lymphoproliferative complications. The development of a distinctive form of interstitial lung dis- ease, termed granulomatous-lymphocytic interstitial lung disease (GLILD), is particularly problematic in CVID because it is relatively common, associated with significant mortality and morbidity, and has an unmet need for adequate treatments. We found that the biologic drug abatacept showed good efficacy for reversal of severe, often refractory GLILD in a small series of patients with CVID. Abatacept is a recombinant fusion protein incor- porating CTLA-4 (cytotoxic T lymphocyte–associated protein 4) that blocks T cell activation by binding to CD80/CD86, thereby preventing CD28 engagement. Our findings suggest that abatacept would be an effec- tive therapy for GLILD in CVID. However, rigorous clinical trials are needed to prospectively define the risks and benefits of abatacept as a therapy for GLILD. To fill this critical gap, we have designed the ABCVILD trial. In this trial, we plan to treat with either pla- cebo or abatacept in a double-blinded fashion. Following a six-month blinded treatment phase, patients will enter a six-month open-label phase in which they will start or continue abatacept. On the basis of our clinical experience to date, we hypothesize that 6 months of abatacept therapy (compared to placebo) will clinically improve GLILD (as assessed by objective and QOL measures) across CVID genotypes and improve sCD25 and other exploratory biomarkers of T cell activation. We will test this hypothesis by pursuing these aims: Aim 1: Determine the response rate of GLILD after six months of abatacept therapy (versus placebo). Our readouts for the ABCVILD trial include quantitative assessment of chest CT scans, pulmonary function tests, quality of life measures, and radiation-free measure 129Xe magnetic resonance image (MRI). Aim 2: Determine whether genotype and/or lung histology predict GLILD responsiveness to abatacept therapy. As CD4+ T cells are often the most prominent infiltrating cells in GLILD and T follicular helper (Tfh) cells have been shown experimentally to be sensitive to CD28/CTLA4 manipulation, we hypothesize that GLILD is largely a disease of Tfh cells and that responsiveness will correlate with these cells in lesions. Aim 3: Define the utility of sCD25, abatacept pharmacokinetics, or exploratory biomarkers of T cell acti- vation for predicting response to therapy. We will serially assess sCD25, various exploratory biomarkers, and abatacept levels to test the predictive value of each, understand the effects of our adaptive dosing regimen on these parameters, and further test our hypothesize that GLILD is a consequence of inappropriate T cell activa- tion in patients with CVID.

NIH Research Projects · FY 2025 · 2021-09

SUMMARY/ABSTRACT Epilepsy is very prevalent and highly refractory to currently available medical treatments in Tuberous Sclerosis Complex (TSC), a genetic disorder affecting 1:6000 live births. Medically-refractory epilepsy in TSC is associated with lifelong intellectual disability and neurodevelopmental deficits. Everolimus and sirolimus, pharmacological inhibitors of the mechanistic target of rapamycin complex 1 (mTORC1), have been successfully repurposed to treat may clinical manifestations of TSC, including focal-onset epilepsy. However, few patients become seizure- free following treatment with mTORC1 inhibitors and 60% of patients with TSC are still in need of effective treatment. Mouse models of TSC and human clinical trials indicate early treatment with mTORC1 inhibitors, before the onset of seizures, may be a more effective treatment strategy against epilepsy and epilepsy- associated deficits in neurodevelopment in patients diagnosed with TSC. The current study proposes a Phase IIb multicenter, randomized, double-blind, placebo-controlled clinical trial with sirolimus to test this hypothesis. The primary aims of the clinical trial are (1) to demonstrate that sirolimus prevents or delays seizures in infants with TSC that are 0-12 months of age; and (2) to demonstrate that sirolimus is safe and well-tolerated in infants with TSC that are 0-12 months of age. Additional (secondary) aims of the trial are: (1) to demonstrate that early sirolimus treatment improves developmental delay, language impairment, adaptive skills, and autism risk; (2) to assess the utility of EEG and MRI biomarkers for measuring mTORC1 inhibition in the brain; and (3) to validate precision dosing of sirolimus in infants with TSC.

NIH Research Projects · FY 2025 · 2021-09

Project Summary Hematopoietic stem cell transplant (HSCT) is increasingly used for non-malignant hematologic disorders such as sickle cell anemia and marrow failure, and complications of HSCT are more prevalent and important as causes of long term morbidity. Improved diagnosis and treatment and treatment are urgent issues. Lung injury is frequent after HSCT, with as many as 15% of children transplanted having reduced FEV1, later leading to overt bronchiolitis obliterans (BO) which is associated with high morbidity and mortality. Diagnosis of BO in adults depends on spirometry, and evidence suggests that early diagnosis may improve chances of recovery of lung function and survival. Small children, and even older children and teenagers who feel unwell and are uncooperative, are unable to perform spirometry, meaning that lung impairment is “invisible” until severe and clinically manifest. Progress in improving lung outcomes of HSCT in children has been hindered by lack of diagnostic strategies, and by the need for a large group of cases to allow testing of biological, pathological and diagnostic hypotheses to advance the field. We propose assembling a prospective cohort study of all children receiving HSCT at 6 major US transplant centers. We will perform standardized prospective testing of lung function and clinical data collection in all children in the cohort, to determine the frequency, clinical phenotypes and risk factors for BO (Specific Aim 1). BO is a rare disease so progress will not be made absent a multi-center cohort study that uniformly defines clinical phenotype and prospectively collects samples, allowing retrospective study of early molecular events that predict later disease. In Specific Aim 2 we will collect calendar and event-driven biological samples, including serum, plasma, peripheral blood mononuclear cells and bronchiolar lavage fluid, which will be stored and used to identify biomarkers of onset of BO and of response to therapy. Moreover, we will collect pathological samples that will be analyzed using a pre- existing rare lung disease pathology platform to identify genetic and anatomic changes that define BO. Lastly, in Specific Aim 3 we will test and disseminate novel lung testing strategies, including innovative imaging, to address the critical clinical challenge of late diagnosis of lung impairment in children. We provide examples of essential biological studies that can only be performed using the resources of the cohort, for which we will seek additional R01 funding. Taken together, this cohort study can fundamentally change the paradigm of lung injury after transplant and provide a platform for testing preventive and therapeutic treatments.

NIH Research Projects · FY 2025 · 2021-09

Abstract/summary Leveraging the electronic health record to characterize and optimize care delivery for children with cerebral palsy: Cerebral palsy (CP) is the most common physical disability of childhood, but it is highly heterogeneous with respect to its severity, response to therapy, care needs, and impact on wellness for the child and family. To optimize health and wellness throughout life and enable new research avenues to be effectively tested, it is critical to develop a comprehensive clinical care and biopsychosocial data model. Development of a comprehensive model would both accelerate and improve understanding, care, and further research, including the identification of novel targets for interventions. The overriding objective of this proposal is to develop a precision health model for CP-related phenotypes, health status, care activities, and psychosocial well-being that will individualize care. To accomplish this objective, we will automate the collection, cleaning, and integration of multi-dimensional, multi-domain and multi-cohort based "big data" extracted from the electronic health record (EHR), and combine this EHR data with prospectively collected, high-resolution clinical, functional, environmental and psychosocial data. The focus of this proposal will be on children between ages 6 and 12 years. Preliminary work indicates that the medical center provides care for approximately 1,800 patients with CP who are between the ages of 6 and 12 years and have at least three years of EHR data. From this EHR cohort, we will prospectively recruit 200 children and their families for detailed phenotyping. Recruitment will be stratified by Gross Motor Functional Classification System (GMFCS) Levels: 60% GMFCS I, II, or III (able to walk) and 40% GMFCS IV or V (use a wheelchair). Using this multi-cohort design will allow for robust characterization of multi-dimensional factors that impact care receipt, functional outcomes, quality of life and participation. Aim 1 will focus on creating a diverse and comprehensive data repository using both retrospective and prospective data to characterize actual versus optimal care (defined by current evidence-based literature). Aim 2 will lead to development of a receipt of care coefficient score and characterize how the degree of optimal care relates to function, quality of life and participation, controlling for functional status and age. [Models developed in Aim 2 will be translated into a clinical decision tool prototype. Aim 3 will demonstrate proof of concept for scalability of machine learning algorithms with the PEDSnet Learning Health System. This project innovatively combines retrospective EHR data with prospective clinical data to elucidate individual, treatment, family, and environmental factors associated with greater receipt of evidence-based care and/or better outcomes. This project will move the field toward precision medicine for CP and create a foundation for development of clinical dashboards to optimize practice.]

NIH Research Projects · FY 2024 · 2021-09

Prematurity is a public health crisis impacting 10% of all births. There is significant risk for neurocognitive impairment, including language. This can adversely impact quality of life. Current prognostic tools leave a significant proportion of variance in later language scores unexplained, focus on risk factors which are often not modifiable, and fail to identify resiliency, or positive outcome despite risk. There is a critical need for advanced neuroimaging studies of well-performing extremely preterm (EPT, <28 weeks gestation) children at school-age and beyond to identify brain-based markers of resiliency while explicitly accounting for socioenvironmental factors. The proposed experiments will address this need by leveraging an existing cohort of 45 EPT children with no overt brain injury and 45 term comparison children. These EPT children were found to have increased functional and structural connectivity--including an extracallosal interhemispheric pathway--versus term children, which positively correlated with language at 4 to 6 years of age for EPT children exclusively. The central hypothesis is that EPT children performing within normal limits on language tasks do so via adaptive neural networks--which are extracallosal and bypass areas vulnerable to white matter injury of prematurity--and the environment in which EPT children are developing drives this adaptive hyperconnectivity. The objective of the proposed research is to determine if this hyperconnectivity remains a brain-based biomarker of resiliency at 8- 11 years and analyze the influence of the child’s social environment on hyperconnectivity. The first aim is to determine if the extracallosal hyperconnectivity (versus term children) we observed at 4-6 years of age in the EPT group persists to 8-11 years of age. The second aim is to determine the extent to which language scores at 8-11 years relate to hyperconnectivity for EPT children. The third aim is to evaluate the extent to which environmental factors drive brain connectivity. Children will be assessed longitudinally with magnetoencephalography, functional magnetic resonance imaging (MRI), and diffusion MRI to validate unique neuroimaging signatures previously identified. Environment-based factors will be assessed at birth (through linkage with birth records), 4-6 years, and 8-11 years to evaluate the extent to which they influence preterm brain connectivity. The candidate’s long-term goal is to reduce the burden of neurodevelopmental impairment for children born EPT by uncovering adaptive, neuroprotective mechanisms and developing imaging markers that can subsequently be used to improve prediction of outcomes before language can be reliably assessed. Through formal coursework, workshops, and hands-on training during the proposed research with her interdisciplinary committee of expert advisors and mentors, the candidate will gain expertise in epidemiological methods to evaluate socioenvironmental risk across generations and in the science of adversity and resiliency. This is a critical next step to establish scientific independence as an investigator and a leader in the study of socioenvironmental resiliency, language development, and neuroprotective mechanisms in the preterm brain.

NIH Research Projects · FY 2025 · 2021-09

PROJECT SUMMARY ABSTRACT- MANDATORY CORE The purpose for Mandatory Core Component A is to support an Enhanced Surveillance Network of US pediatric institutions to develop and implement standard research protocols to conduct prospective active population-based surveillance in defined inpatient and emergency department (ED) settings for a) acute gastroenteritis (AGE) due to norovirus, rotavirus and other enteric pathogens, b) acute respiratory infection (ARI) due to respiratory viruses including influenza, RSV, parainfluenza viruses, human metapneumovirus, rhinoviruses, enteroviruses (including EV-D68), adenoviruses, coronaviruses (including SARS-CoV-2) and other respiratory viruses, c) and healthy controls (HC) and d) Acute Flaccid Myelitis (AFM) syndrome among pediatric patients seeking healthcare at these pediatric medical institutions. The results from this program will be used to inform vaccine recommendations and assess the public health impact of the US rotavirus, influenza and SARS-CoV-2 vaccination programs and provide epidemiologic data for other infectious diseases with therapeutics and vaccines in development. There are two objectives for Mandatory Core Component A:: For Objective 1 we will conduct population-based active surveillance for respiratory and enteric viral pathogens in pediatric inpatient and ED settings and enroll asymptomatic healthy controls in Hamilton Co. children <18 years of age. Through our already established ARI, AGE and HC Surveillance Platforms we will 1) perform active surveillance to determine the etiology and burden of inpatient and ED acute viral enteric and respiratory diseases in our defined population, 2) characterize the clinical and epidemiologic factors of infections including asymptomatic children and 3) evaluate VE and impact of vaccines and other interventions available or projected to become available during the study agreement period for rotavirus and influenza vaccines using a test- negative design and for RSV and SARS-CoV-2, when available and recommended for children and adolescents. For Objective 2 we will conduct surveillance activities for acute flaccid myelitis (AFM in hospitalized children <18 years of age). Through our already established AFM Surveillance Platform, we will 1) define baseline rates of AFM in our pediatric institution through active case finding in collaboration with our Neurology and Neuroradiology Co-Investigators using the CDC's case definition for patients meeting the clinical criterion for AFM and a spinal MRI showing at least some gray matter involvement by conducting active surveillance and establishing incident rates for AFM among hospitalized children within our catchment area, 2) compare rates of AFM to rates of circulating respiratory and enteric pathogens at our site from our population- based active surveillance program and 3) characterize the clinical spectrum of pediatric AFM and compare with the clinical spectrum seen with other similar neurologic conditions.

NIH Research Projects · FY 2024 · 2021-08

Abstract: Mounting evidence indicates that lineage decisions are controlled by local structures that support differentiation in distinct regions of the bone marrow. Our ability to manipulate hematopoiesis to treat disease has been hampered by our lack of understanding of these anatomical cues. We have found that HPC are regionally organized in the BM and that progenitors and immature cells belonging to the same lineage segregate to different sinusoids. We have also found that there is regional organization to CSF1 production and that specific BM vessels are critical sources of CSF1 that regulate MDP, monocytes and dendritic cells but not HSC or macrophages. The central hypothesis is that different subsets of sinusoids function as assembly lines or hubs that organize and maintain different hematopoietic lineages through the production of lineage-specific cytokines. We want to understand how these cellular assembly lines function in the steady-state and after perturbation by acute myeloid leukemia (AML). We will test this hypothesis in two aims. In Aim 1 we will determine the physiology of these assembly lines using clonal fate mapping in situ; identifying the components of each assembly line; and conditionally deleting specific cytokines from key components of each assembly line. In Aim 2 we will investigate how AML inhibits normal hematopoiesis by perturbing the microenvironment that supports each assembly line.

NIH Research Projects · FY 2025 · 2021-08

Abstract Children with acquired brain injuries (ABI) treated on an inpatient rehabilitation unit are at significant risk for long term functional impairment, highlighting the importance of maximizing the effectiveness and utilization of inpatient rehabilitation therapies. The proposed crossover trial seeks to determine the effect of animal-assisted therapy (AAT) during inpatient rehabilitation following pediatric acquired brain injury. To accomplish this objective, we will employ a within subjects cross-over trial. Data will be collected during four therapy sessions across two weeks (2 in week 1 of admission and 2 in week 2 of admission). AAT, via integration of dogs into patient's physical therapy (PT) and occupational therapy (OT) session, will occur during one of the two sessions in the first week and one of the two sessions the second week. The non-AAT condition will be treatment as usual (TAU) as defined by the patient's treatment team, resulting in 2 AAT sessions and 2 TAU sessions total. The order of condition will be randomized separately for each week. We will recruit 90 patients ages 4-21 being treated for ABI on the inpatient rehabilitation unit at CCHMC. We have engaged a highly qualified multidisciplinary team to ensure the safety, feasibility, and effectiveness of the methods. We will accomplish the following aims: 1) determine the effect of AAT on patient engagement in inpatient rehabilitation therapies, 2) examine potential mechanisms of action for AAT-associated improvement in patient engagement (mood and physiological distress), and 3) determine factors that moderate the effect of AAT on patient engagement (patient anthropomorphism, patient-animal closeness, time course in admission, and demographic and injury characteristics). We hypothesize that a greater level of patient engagement will be observed during AAT sessions compared to treatment as usual (TAU) sessions. Additionally, AAT will be associated with more positive mood and lower levels of distress, which in turn will be related to greater patient engagement. Finally, a greater effect of AAT will be noted earlier during patient admissions, and greater levels of anthropomorphism and patient-animal closeness will be associated with greater levels of patient engagement during AAT sessions. We propose the use of novel methodology and innovative technology in data collection designed to address concerns related to bias and reliance of subjective outcomes in the AAT evidence base. This innovation positions the project to fill an important gap in the existing AAT literature. Findings will provide critical new insights into why pediatric inpatients benefit from AAT and who is most likely to benefit. We will translate our findings to ultimately improve functional outcomes following pediatric brain injury, a leading cause of disability.

NIH Research Projects · FY 2025 · 2021-08

SUMMARY/Abstract: The long-term goal of the Cincinnati Cooperative Center of Excellence in Hematology (CCCEH) is to understand and correct, at the molecular level, hematological diseases of various lineages. To attain this goal, we must understand basic biological processes that affect behaviors of hematopoietic stem cells and their various differentiated progenies in vitro and in vivo. We envision this long-term goal will be best accomplished by interdisciplinary and collaborative approaches using state-of-the-art methods, along with timely and rational use of translational studies into the preclinical and clinical settings. Cincinnati Children's Hospital Medical Center (CCHMC) has developed a rich and stimulating intellectual environment that brings together outstanding expertise and basic research in stem cell biology, virus vector technology, immunohematologic cell biology, genomics and genetics, and hematopoiesis; in addition, the CCCEH has world-class expertise in translational and clinical research in hematology, immunology and hematopoietic stem cell transplantation. This focus has been greatly strengthened by the recruitment of over 30 faculty members in these areas of hematology research over the last 15 years at CCHMC, and the inclusion of over 15 external hematology researchers around US. Basic research is linked with Cincinnati NIH-funded Clinical and Translational Science Award (CTSA), which has facilitated interdisciplinary interactions and developed an infrastructure that supports state-of-the-art clinical trials in cell and gene therapy. Here we seek to solidify the exciting and systemic growth of research in hematology at CCHMC to support a national center of excellence, using the shared services and administrative structure of the center grant mechanism. The CCCEH Administrative Core supports an exceptional array of research cores, which function together to support innovative basic and translational research both locally and nationally, in non-malignant hematology. Our three proposed research cores include (1) Xenotransplant and Genome Editing Core, which maintains specialized mouse strains and provides mouse transplant and transgenic services; (2) Single Cell Characterization and Procurement Core that includes state- of-the-art imaging as well as single cell capture and gene expression capabilities; and (3) Gene Delivery Core that covers services such as ex vivo manipulation of blood cells by lentivirus and retrovirus transduction and shRNA/sgRNA screening. Finally, this center seeks support for an innovative Enrichment Program that fosters a collaborative and educational environment in the center and for outreach to the broad hematology research community around US.

NIH Research Projects · FY 2025 · 2021-07

Abstract Bronchiolitis obliterans syndrome (BOS) is an obstructive lung disease caused by a combination of inflammation and immune response that is irreversible in its late stage. Children with BOS are typically diagnosed late because they are unable to perform spirometry, and morbidity and mortality are high. The long-term goal of this work is to improve survival and reduce morbidity from BOS by identifying strategies for accurate screening and prediction of BOS in children and young adults after HSCT and using these tools to identify novel drug targets for early intervention or prevention of BOS. The objective of this application is to validate novel predictive plasma protein biomarkers and establish a dynamic prediction model for BOS for early diagnosis, risk stratification and disease trajectory prediction for BOS after HSCT. We will achieve these goals through the following specific aims: 1) Validate longitudinal predictive performance of newly discovered plasma biomarkers of BOS risk in samples from banked and prospective studies by mass spectrometry and ELISA in both pediatric and adult cohorts. 2) Optimize and validate our dynamic prediction algorithm using pulmonary function and clinical data as well as biomarker levels (needed when no spirometry can be obtained) as covariates to project risks of BOS and rapid BOS lung-function decline to inform treatment decisions. There are currently no biomarkers or predictive tools for BOS so this work is entirely novel. The use of a dynamic prediction algorithm in this clinical setting is innovative allowing for the first time the ability to predict and diagnose early lung disease in HSCT subjects prior to the clinical diagnosis of BOS. Identification of BOS risk and stratification of screening and treatment procedures according to risk and predicted disease course would allow us to modify post-transplant care and reduce morbidity and mortality. We will use our data to inform prospective clinical trials of both early active treatment prior to development of early fibrosis and to test novel prophylactic therapies to reduce incidence in high risk individuals. Our studies will provide blood biomarkers that can be used as frequently as necessary without requiring active participation from small and often very sick children. Our preliminary biomarker and HSCT specific algorithm data demonstrate detection of BOS as soon as 2 weeks to 6 months prior to clinical diagnosis of BOS. This work is both significant and vital because improvements in HSCT techniques and supportive care have led to improved survival. Improved survival increases the number of children at risk for late complications of HSCT that are associated with life-changing morbidity and late mortality and there is urgent need to address these issues. This work will advance prediction and early diagnosis of BOS, as well as providing the framework for future prevention and treatment trials.

NIH Research Projects · FY 2024 · 2021-07

We normally think of the neural retina as the only light-responsive tissue of mammals. Recently, however, it has been shown that the atypical opsins Opsin 3 (OPN3, encephalopsin) and Opsin 5 (OPN5, neuropsin) are expressed in other tissues. Investigating this further, the Lang lab has shown that OPN5 mediates light reception in skin and brain and that OPN3 does so in adipose tissue. In all cases, these opsins mediate direct, acute light responsiveness of the non-retinal tissue. Here we identify the ocular lens as an intrinsically light sensitive tissue. Lenses isolated in culture show an acute response to blue (470 nm) light that changes their optical properties. According to preliminary data, OPN3 is required for the intrinsic light response of the lens and we further show that light stimulation of the lens suppresses the phosphorylation of the water channel Aquaporin 0, a substrate of protein kinase A. Since Aquaporin 0 regulates lens transparency, we hypothesize that an OPN3-dependent, intrinsic light response regulates aquaporin activity via the cAMP-PKA pathway and thus the optical performance of the lens. This hypothesis implies that the lens has distinct optical states adapted to daytime and nighttime vision. Seeking a deeper understanding of this unique lens biology, we propose three aims, (Aim 1), To determine which G-protein OPN3 uses for signaling in the lens, (Aim 2), To confirm the OPN3 signaling mechanism, and the influence of that signaling on aquaporins and lens transparency including cataractogenesis, and (Aim 3), To assess the influence of lens OPN3 on lens refractive power and eye refractive development. This application builds on a long history of interest in the ocular lens from the Lang lab and combines expertise in lens cell biology from Steve Bassnett and eye refractive development from Rafael Grytz and Machelle Pardue. Identification of a direct, light- and opsin-dependent optical change in the ocular lens is unexpected and so when complete, this work will fundamentally change the way we think of the eye as a light sensing organ. There is also the possibility that this work will identify the lens as a new target for myopia treatment.

NIH Research Projects · FY 2025 · 2021-07

Project Summary Initiation of innate immune responses depends on cognate interaction between germline-encoded pattern recognition receptors and their ligands (expressed by microbes). Following recognition, the receptors initiate activation of downstream signal transduction pathways that often involve recruitment of downstream adapters and kinases. The Toll-like receptor family of PRRs, which are the subject of current investigation, are expressed both on the plasma membrane and in the endosomes. Several recent studies have demonstrated that endocytosis of the plasma membrane TLRs (especially TLR4) plays a critical role in regulating both quality and magnitude of inflammatory responses in a responding macrophage. Other studies have demonstrated that TLR signaling enhances phagocytosis of microbial cargo but not of apoptotic cell cargo suggesting a degree of specificity that is not understood. In addition, although endocytosis of TLR4 and the events following endocytosis of TLR4 that influence signal transduction are very well studied, it is not entirely clear if and how endocytosis influences signaling downstream of other plasma membrane and endosomal TLRs. In our studies, we find that a protein called Vps33B regulates handling of the phagocytic and endocytic cargo following pattern recognition receptor activation. More importantly, Vps33B directly influences the outcome of signaling downstream of TLRs in mice and Toll- and IMD pathways in Drosophila. Mutations in the genes VPS33B and VPS16B are linked to a rare human disease called ARC (Arthrogryposis-renal dysfunction-cholestasis) syndrome. Both of these ARC genes encode paralogs of HOPS complex subunits suggesting a role in membrane fusions but how perturbation of function of these proteins results in a diverse spectrum of disease symptoms in ARC patients is not entirely clear. It has however been documented that ARC patients suffer from sepsis and recurrent bacterial infections and we were therefore investigated the role of these proteins in influencing immune responses. We find that in the absence of VPS33B, Drosophila respond vigorously to microbial insult. Exaggerated immune responses are generated in response to live or dead bacteria and purified ligands of the Toll and IMD pathway results in death of Vps33B mutant, but not wild-type flies. This function of Vps33B is conserved in vertebrates and we find that mouse macrophages lacking Vps33B secrete very high quantities of inflammatory cytokines, when stimulated by either plasma membrane or endosomal TLR ligands. We therefore hypothesize that activation of pattern recognition receptors and specifically TLRs leads to formation of specialized endosomes that depend on Vps33B for lysosomal fusion. Lack of Vps33B is likely to affect several aspects of innate and adaptive immunity and to test this hypothesis, we propose to 1. Define the molecular events that regulate Vps33B function in endosomal maturation, 2. Define the role of Vps33B-regulated TLR trafficking and signaling 3. Investigate the role of Vps33B in regulating cargo handling by DCs and 4. Investigate the role of Vps33B in regulating antigen presentation and adaptive immunity.

NIH Research Projects · FY 2025 · 2021-07

ABSTRACT Hematopoietic stem cell transplantation (HSCT) is a highly effective treatment, but serious virus infections occur in 82% of children undergoing HSCT at our institution and others. Current anti- viral drugs have inadequate response rates, prolong hospitalizations, and are frequently associated with organ toxicity. An alternative cellular therapy approach uses engineered viral specific T-lymphocytes (VSTs) manufactured from blood donated by a patient’s stem cell donor. VST therapy is highly effective when cells are infused in response to viremia, with response rates of over 80%. We seek to make a critical advance in this technology by testing whether scheduled administration of bone marrow donor derived VSTs 21 days after HSCT will be safe and at least as effective as our current pre-emptive treatment approach of only administering VSTs once viral reactivation or infection has occurred. We propose two specific aims. Specific Aim 1: Randomized comparison of donor-derived scheduled vs treatment VSTs in HSCT recipients to prevent viral infections. Hypothesis: Recipients of scheduled VSTs given 21 days after stem cell infusion will have a significantly lower frequency of viremia and invasive viral infections 100 days after HSCT than patients randomized to treatment use of VST’s. Specific Aim 2: Identify product characteristics that predict response to therapy and compare responses in those with and without exposure to viral ligand. Hypothesis: Specific product characteristics can be established that will identify VST products likely to be clinically effective, and VSTs will persist in scheduled VST recipients without stimulation from viral ligand. Aim 2a: We will study the T cell response to adenovirus, EBV, CMV or BK virus, and product persistence by ELISpot testing, TCR clonogram and VST persistence using TCR sequencing to define more and less effective VST products, and to determine if VSTs expand and persist if they are infused on a schedule into a person with no active viral replication to provide ligand. Aim 2b: We will systematically assess HLA restriction of presentation of viral antigens, using peptide mapping and single antigen cell lines (SALs). These data are vital for future “third party” use of VSTs in persons without donor derived product. This clinical trial may change the paradigm of treatment for viral infections if we are able to show that prophylactic VST infusion prevents infections.

NIH Research Projects · FY 2025 · 2021-05

PROJECT SUMMARY: It is being increasingly recognized that changes in chromatin state are associated with a wide spectrum of lung diseases, ranging from bronchopulmonary dysplasia (BPD) to chronic obstructive pulmonary disease (COPD). However, the mechanisms by which these changes contribute to the pathogenesis of these diseases, and how to manipulate the epigenome for therapeutic benefit, remains largely unknown. Modulation of chromatin accessibility is an important epigenetic mechanism by which gene expression is controlled, even across repeated cell divisions. However, as a prerequisite to understanding how altered chromatin accessibility contributes to disease, the mechanisms by which chromatin accessibility patterns first establish and maintain cellular identity within the lung must be defined. This proposal is based on studies from our group that identified the SWI/SNF proteins Arid1a and Arid1b as key mediators of the chromatin accessibility changes that occur during development of the SOX9+ lung epithelial stem/progenitor cell population. Our data demonstrate that loss of Arid1a or Arid1b led to persistence of the SOX9+ progenitor cell population, impaired alveolar differentiation, and neonatal death due to respiratory distress. In addition, ARID1A directly interacts with NKX2-1 and SOX9. The central hypothesis of the present proposal is that ARID1A and ARID1B interact with key lung developmental TFs to direct the SWI/SNF complex to remodel chromatin at specific loci, silencing progenitor cell gene expression programs and promoting the maturation and function of the mature alveolar epithelium. The proposed studies will: A) Define the role that Arid1a/Arid1b, and the larger SWI/SNF complex, play in establishment of mature alveolar cell type identify in mouse and human. B) Identify the mechanism(s) by which the SWI/SNF complex remodels chromatin, in conjunction with key lung transcription factors, to establish and maintain gene expression modules controlling type I & II AEC identity and function. C) Determine how Arid1a/Arid1b-mediated chromatin remodeling contributes to the lung epithelial repair response following influenza infection. These studies will provide conceptual advances in our understanding of how mature alveolar epithelial cells are established and maintained, how the chromatin accessibility landscape interacts with previously well- defined transcription factor networks, and how chromatin remodeling directs the normal repair process after lung injury. Emerging epigenomic tools and systems biology approaches will be applied to the epithelium for the first time. Taken together, these data will inform future translational studies seeking to understand how alterations in the epigenome contribute to lung disease, and will provide a foundation for future efforts to manipulate the lung’s epigenomic code to restore normal lung structure and function.

NIH Research Projects · FY 2025 · 2021-05

ABSTRACT Gene complementation and pulmonary macrophage transplantation (PMT Therapy) is a promising potential therapy of hereditary pulmonary alveolar proteinosis (hPAP) – a disorder of progressive of alveolar surfactant accumulation and respiratory failure – for which no pharmacotherapy therapy exists. We defined the patho- genesis, presentation, diagnosis, and management of hPAP, showed it is caused by the loss of GM-CSF re- ceptor signaling and disruption of alveolar macrophage (AM) functions including the removal of surfactant from alveoli. We demonstrated lentiviral vector-mediated complementation of function-disrupting CSF2RA mutations restored GM-CSF receptor signaling in human AMs including rescue of surfactant clearance. Despite outstand- ing progress, including demonstration of the safety, tolerability, efficacy, and durability of PMT Therapy in two validated hPAP animal models, lack of clinical studies of PMT Therapy in humans is a critical barrier to its fur- ther therapeutic development. Our long-term goal is to develop PMT Therapy as the an effective, disease- specific therapy of hPAP (and possibly other diseases). The objective here is to complete preparations for, and then to conduct, a Phase I trial to establish the safety of PMT in human patients with hPAP and also identify useful clinical and biological outcome measures informing the design of a future Phase II efficacy trial. The central Hypothesis is that after PMT of autologous CD34+ cell-derived CSF2RA gene-corrected macro- phages without myeloablation, the transplanted cells will survive, engraft, adopt the phenotype and function of normal AMs, replace endogenous AMs, reestablish a normal-sized AM population that remains in the lungs and results in a safe, well-tolerated, effective, and durable treatment benefit. The rationale for the proposed research is that a ‘first-in-human’ study establishing the safety of PMT Therapy in humans will unblock further clinical development of PMT Therapy including preparation for conduct of a future phase 2 clinical efficacy trial. We plan to address our hypothesis by pursuing four specific aims in the R61 phase and three aims in the R33 phase: 1) finalize stability testing of CSF2RA gene-corrected macrophages; complete 2) trial-related and 3) IND-related documents; 4) obtain regulatory approvals (Institutional Review Board and Biosafety Committee, Data, Safety, and Monitoring Board, FDA); 5) assess the safety, 6) measure the pharmacokinetics and phar- macodynamics, and 7) identify useful measures of the clinical outcomes and biological signature of PMT Ther- apy in hPAP patients. The proposed research is innovative because it represents a marked departure from the current treatment approach, whole lung lavage (an invasive, inefficient, procedure to physically remove surfac- tant) by establishing, in hPAP patients, the feasibility of a new approach to restore a GM-CSF-responsive, functional AM population. The proposed research is significant because establishing the safety, pharmacoki- netics and pharmacodynamics, and identifying useful clinical outcome measures of PMT Therapy in humans is the next step in our clinical research program to develop PMT as the first specific therapy of hPAP.

NIH Research Projects · FY 2026 · 2021-04

ABSTRACT Asthma is a common, complex and costly pediatric chronic condition in the U.S., resulting in nearly 2 million acute-care visits and $82 billion in overall costs each year. Children of low-income families in urban communities have the highest prevalence of asthma and the greatest disparities in asthma morbidity and mortality. In Hamilton County, OH, over 36,000 children have asthma, >14,000 of whom are Medicaid- insured with a 10-fold higher rate of hospitalization. Asthma exacerbations, acute episodes of increased asthma symptoms and deteriorations in lung function, are a major cause of stress for patients and families. While exacerbations are a prominent feature of poorly controlled and severe asthma, exacerbation rates can be high even in patients with mild asthma and asthma exacerbation frequency can remain unchanged despite intensive controller therapy. Indeed, pediatric studies have revealed that exacerbations continue to occur despite good baseline symptom control. A frequent exacerbator phenotype of asthma (defined by 2 or more exacerbations/year), has been described, but little is known about this asthma subgroup, which disproportionately affects urban populations and is responsible for considerable asthma morbidity and healthcare costs. No specific clinical characteristics can reliably discriminate between frequent and nonfrequent exacerbators highlighting the need for systems level approaches. Our center-specific project will fill this gap in understanding. Our preliminary data reveal differentially expressed and differentially methylated genes in the nasal epithelial of children who are frequent exacerbators and highlight biologic pathways that implicate distinct mechanisms that underlie the frequent exacerbator phenotype. We will test the hypothesis: the frequent exacerbator is a distinct endotype of asthma that is characterized by host nasal epithelial transcriptomic and/or DNA methylomic patterns that distinguish the frequent exacerbator from asthmatic children who are not frequent exacerbators, and that these patterns are, in part, triggered by distinct nasal microbial patterns. We propose an innovative strategy utilizing multiple systems-based approaches that layer host nasal biome and methylome patterns with nasal epithelial transcriptomics taken during an acute exacerbation (before administration of steroids). We are uniquely poised to conduct this study due to the tremendous infrastructure that we have established as part of the Ohio Pediatric Asthma Repository, and our multidisciplinary team. The overall objective of our studies is to improve the lives of children with asthma from low-income families who live in urban communities. We propose to 2 aims: (1) To contribute to the overall CAUSE goals by conducting network-wide CAUSE clinical research projects and participating in the CAUSE Steering Committee and other network functions; (2) To conduct a center- specific project aligned with CAUSE goals focused on the frequent exacerbator asthma phenotype.

NIH Research Projects · FY 2025 · 2021-04

PROJECT SUMMARY Acts of school violence have increased over the past decade and over 20% of students report being bullied at school. School violence has a far-reaching impact on the entire school population, including staff, students and families. It was noted that the largest crime-prevention results occurred when youth at elevated risk were given effective prevention programs. As such, there is a critical need for developing a rapid and accurate approach to interview students, assess their risk characteristics, and provide supportive evidence for prevention. Our study focuses on detecting and preventing youth aggression, the predominant form of school violence. Several risk assessment scales, ranging from simple clinical impressions to structured professional judgments, have been proposed to identify youth violence. However, these assessments heavily rely on clinicians' subjec- tive impressions and their predictive validities remain a major issue. In addition, none of the risk assessments include direct analysis of the words (language) used by students and hence, provide little information to sup- port subsequent prevention. Our long-term goal is to develop an Automated RIsk Assessment (ARIA) system to analyze participant interviews, detect elevated-risk students, and provide risk characteristics (e.g., impul- sivity, negative thoughts) to assist prevention. In our earlier study we developed a risk assessment approach to interview students and evaluate their risk of aggression. The overall objective of this study is to validate our risk assessment approach with real-world evidence, and to develop an AIRA system to automate the assessment process. We hypothesize that our risk assessment approach will have sufficient predictive validity in predicting aggression at school, and a computerized system leveraging machine learning and natural language pro- cessing (NLP) will be able to detect high-risk students, identify violence-related predictors from linguistic con- tent, and improve subsequent prevention by assisting recommendations. The hypothesis will be tested by pur- suing three specific aims: 1) Evaluate the predictive validity and generalizability of our risk assessment approach with prospectively collected school-based outcomes; 2) Develop a high-performing ARIA system to identify risk characteristics and predict risk of school violence; and 3) Compare actionable recommenda- tions and school outcomes with and without using the ARIA system in a prospective observational study. The study is highly innovative in that it will be among the first efforts that leverage NLP and machine learning to analyze interviews, identify risk characteristics from student language, and predict violence outcomes. The study will have a significant impact on several fronts. Successful validation of our risk assessment approach on multiple sites (Aim 1) will provide a valid mechanism to detect youth aggression at school. The AIRA system developed in Aim 2 will enable accurate and scalable risk screening for individual students. Aim 3 is a bench-to-practice translational aim to rapidly transfer our findings to clinical practice. The study will help establish a nationwide solution for school violence risk assessment, which will benefit healthcare institutions, schools, and students.

NIH Research Projects · FY 2025 · 2021-03

Proposal Summary/Abstract: Though outcomes for pediatric acute leukemia have improved dramatically over the last few decades, infant leukemia is a particularly aggressive disease which remains difficult to cure even with chemotherapy intensification and bone marrow transplant. Most of these leukemias carry a rearrangement of the MLL gene, a phenomenon also seen in some de novo acute myeloid leukemias (AML) as well as therapy-related AML, and studies have demonstrated that a common gene expression program underlies these leukemias regardless of lineage. Through preliminary studies, we have found that MLL-fusion leukemias express high levels of MBNL1, an RNA binding protein that regulates alternative RNA splicing. We have also identified an alternative splicing (AS) signature unique to MLL-fusion leukemia, and show that MBNL1 loss causes reversion of this signature and impairs MLL-fusion leukemia cell growth. We thus hypothesize that MBNL1 is a critical regulator of an MLL-fusion specific AS program, and that disruption of this program via MBNL1 inhibition leads to leukemia cell death. To test these hypotheses, our specific aims are 1) to determine the mechanism underlying MLL-fusion leukemia dependence on MBNL1, and 2) optimize characteristics of a small-molecule MBNL1 inhibitor as a treatment for MLL-fusion leukemia. To achieve the first aim, we will characterize changes in key cell growth and death pathways which we hypothesize are responsible for the effects seen with genetic knockdown. We will also identify novel MBNL1-mRNA interactions in MLL-fusion leukemia using CLIP-seq (cross-linking immunoprecipitation with RNAseq). For the second aim, we have shown as a proof of concept that a small molecule inhibitor of MBNL1 can induce MLL-fusion leukemia cell death. This compound requires optimizations for potency, which we will achieve by applying medicinal chemistry principles. Furthermore, we have used its structural characteristics and published crystal structures to initiate an in silico screen of a proprietary compound library. This proposal will advance our understanding of the role of AS in the pathogenesis of MLL-fusion leukemia, while also directly leading to a first-in-class therapy for this disease. The applicant, who is currently an instructor in the Division of Oncology at Cincinnati Children’s Hospital Medical Center, will execute this research plan while simultaneously engaging in structured didactics and receiving close individual guidance from a panel of mentors as described in the application. These scientists possess significant expertise in the molecular pathogenesis of leukemia. The experiments, mentoring, and structured classwork described in this career development plan will position the applicant to successfully transition into an independent researcher and physician-scientist, with expertise in the role of RNA binding proteins in leukemia pathogenesis.

NIH Research Projects · FY 2025 · 2021-03

An experimentally-refined, dynamic gene regulatory network model of T-cell memory Summary T cell memory induced by prior exposure to a pathogen or vaccination provides enhanced protection against a subsequent infection with the same pathogen. Enhanced protection is partially driven by clonal expansion, which leads to an increased number of T cells capable of recognizing the antigen. Additionally, memory T cells possess a “rapid recall ability” that allows them to fight pathogens by producing cytokines and other effector molecules within minutes of re-exposure (as opposed to days, upon initial exposure). We recently showed that rapid recall correlates with the epigenetic poising of enhancers and promoters of the “rapid-recall genes” in memory T cells. Importantly, the sites of epigenetic change significantly overlap with the risk loci for autoimmune and atopic disease, suggesting that this mechanism is important for human health. However, it is still unclear if and how the epigenetic poising causes enhanced expression of rapid recall genes. Furthermore, memory T cells persist for a lifetime; yet the mechanisms that maintain the memory epigenome – for decades– are not known. Our preliminary data suggest that rapid recall is coordinated by several families of transcription factors (TFs) and thousands of putative DNA regulatory elements. This complexity requires a systems-level, engineering approach. Thus, this proposal is a collaboration between the groups of Artem Barski, a T cell biologist, and Emily Miraldi, a mathematical modeler, to create experimentally validated, genome-scale models of memory immune responses across heterogeneous T cell populations. Aim 1. Using single-cell genomics, we will characterize the gene expression and chromatin dynamics of T cell activation in naïve and memory cells and build mathematical models that integrate these data (along with relevant existing genomics resources) into a dynamic gene regulatory network (GRN). Our GRN model will predict the molecular drivers (TFs) and regulatory elements that orchestrate rapid recall. Aim 2. Although T-cell activation in naïve and memory cells similarly promotes nuclear translocation of inducible TFs, our data lead us to hypothesize that chromatin remodeling upon initial pathogen exposure alters the occupancy of inducible TFs in memory T cells and that this is the basis of rapid recall. We will combine dynamic TF perturbation and occupancy experiments to establish the molecular interactions driving rapid recall. Aim 3. We will identify the mechanisms by which memory T cells maintain the epigenome conducive for rapid recall – over the human lifespan. We hypothesize that constitutive TFs maintain the epigenome poised for rapid recall. We propose dynamic TF perturbation experiments to uncover the identities of these regulators. This study will help uncover basic mechanisms of T cell memory and identify potential targets for manipulating immunologic memory responses. Because rapid recall is the basis for vaccination and central to allergy, asthma, and cancer immunity, this study will have a broad impact on human health.