Dartmouth College

NIH Research Projects · FY 2026 · 2022-05

PROJECT SUMMARY One in nine people aged 65 or older in the US lives with Alzheimer's disease (AD) or Alzheimer's disease-related dementia (ADRD). Patients living with dementia (PLWD) and their care partners rely on primary care clinic visits for dementia information, management, and community referrals. Quality interpersonal communication is associated with improved health-related outcomes. Models of triadic interactions purport that information exchange, rapport, and patient and care partner engagement in goal setting and decision-making are key to effective interpersonal communication. However, the degree to which effective interpersonal communication is achieved during triadic visits is unknown, and few interventions to support it exist. Using audio recordings of clinic visits is a novel, evidence-based strategy with the potential to support triadic interactions, yet its application is unexplored in dementia. The objective of this proposal is to design an intervention that enhances interpersonal communication in triadic visits using visit recordings. Applicants will follow the NIH Stage Model to redesign their visit recording platform, HealthPAL, which leverages natural language processing to structure visit information. The specific aims are: Aim 1 (Stage 0): Conduct a prospective observational study, with outpatient clinic visits of 200 triads (PLWD/care partner/clinician) audio recorded for 12 months; 1.a. Examine the association between interpersonal communication in triadic AD/ADRD visits and health-related outcomes; 1.b. Identify barriers and enablers to interpersonal communication in triadic AD/ADRD visits; Aim 2 (Stage 1A): Adapt HealthPAL to enhance interpersonal communication in triadic AD/ADRD visits; and Aim 3 (Stage 1B): Demonstrate the usability, feasibility, acceptability, and potential effectiveness of HealthPAL in AD/ADRD. Applicants hypothesize: 1) Constructs from models of interpersonal communication will be associated with health-related outcomes; 2) HealthPAL will surpass usability, feasibility and acceptability metrics for dyads and clinicians. In Aim 1 applicants will use an explanatory sequential mixed methods design. Informed by the Behavior Change Wheel, targets for behavior change will be identified using quantitative assessment of interpersonal communication during triadic visits (200 dyads, 3 visits annually; ∼600 visits), supplemented by semi-structured interviews with a purposive sample of 1a triads (n=42); In Aim 2, we will use participatory design methods (n=60) to redesign HealthPAL using findings from Aim 1; and in Aim 3 we will use an open label, single-arm, multi-site pilot trial (n=30) to determine usability, feasibility and acceptability of HealthPAL and gather preliminary data on its impact on interpersonal communication in triadic AD/ADRD visits. This work is a necessary first step to improving PLWD triadic care by identifying behaviors that impact interpersonal communication and their associations with health- related outcomes. The intervention developed, and the extensive data collected, will serve as a powerful resource that can be leveraged to address other gaps in clinical knowledge related to the care of PLWD.

NIH Research Projects · FY 2025 · 2022-04

ABSTRACT Significant improvement in the effectiveness of radiation therapy (RT) now seems possible because of recent exciting research results using ultra-high dose rates (UHDR), indicating that normal tissue damage can be reduced (the `FLASH' effect), compared to conventional radiation for the same total dose. FLASH RT delivery has shown reduced morphological and functional damage to normal tissues such as brain, colon, lung, and skin. Although significant research remains, early indications are that the normal tissue sparing effect may as high 50% at some dose levels. If proven in translation, this effect would be the most significant improvement in RT therapeutic ratio since the advent of treatment planning. While intriguing mechanisms for this result have been postulated, they remain only well-reasoned speculations because of a lack of direct in vivo data on the physico- chemical mechanisms including oxygen depletion and free radical species alterations. Dartmouth has created the first reversible MeV FLASH beam on a clinically commissioned linac with >100 Gy/s at the patient treatment bed, and prototyped an open-source treatment planning system, expanding research access. Additionally, the team has invented unique technological capabilities to directly measure the highest dose rates and in vivo tissue oxygen transients. The key technological barriers solved in the proposed research with preliminary data are: 1) demonstration of conversion to a UHDR irradiator within a clinically commissioned linac, 2) verification of per- pulse and per-fraction dose rates, 3) direct in vivo observation of oxygen transients, 4) direct measurements of free radical species changes in vitro, and 5) access to functional tissue assays and genetic and proteomic assays. Single-pulse and single-fraction dose rates will be quantified by high frame rate imaging. Additionally, the in vivo oxygen changes will be quantified by two independent methods for co-validation, including electron paramagnetic resonance oximetry and optical luminescence oximetry. Free radical species changes produced from transient hypoxia will also be assessed through systematic in vitro analyses, and the potential linkages to functional, proteomic and DNA damage examined. The FLASH beam conditions that minimize normal tissue damage for a fixed dose will be established with this baseline data. The work is pre-clinical but can be readily adapted to ongoing NIH sponsored spontaneous canine cancer studies has relevance to future large animal and first-in-human translation. Multiple Dartmouth centers partnered to initiate and support this FLASH program. Taken altogether, this bioengineering research project will advance the state of the art in high dose rate radiation therapy using tools that have been uniquely developed by our 3 research groups, and the project results will build the basic science needed to support proposed human translation of this ground-breaking field. The team has leading expertise in radiation physics, in vivo molecular measurement, and radiation genetics and immunomolecular biology and pathology. The work is supported by an External Advisory Board of international expert FLASH consultants, as well as an internal Radiation Oncology Advisory Group.

NIH Research Projects · FY 2026 · 2022-03

Abstract: We study the molecular mechanisms and signaling, from environmental input to physiological output, that control bacterial biofilm formation, a key bacterial lifestyle linked to host-microbe interactions and infectious disease. In particular, the capability of bacteria to sense and respond to various microenvironments, particularly during the transition from a free-swimming to a sessile biofilm lifestyle, contributes to the establishment of chronic infections. The underlying mechanisms are equally important for non-pathogenic bacteria that can live in commensal relationship with their host(s). Understanding the architecture and regulation of signaling systems that control bacterial cell adhesion and biofilm formation is critical in the development of novel therapies and preventative interventions, while providing fundamental insight into bacterial signaling processes. Here, we propose mechanistic studies on a broadly conserved cell adhesion system that is crucial for biofilm formation in a variety of pathogenic and commensal organisms. We will focus on fundamental unanswered questions concerning how cyclic-di-GMP networks control localization of these adhesins as well as how two such adhesins contribute to formation of biofilms. We will also enhance the impact of our studies by investigating a newly identified, analogous signaling system in an important commensal sulfate- reducing bacterium. The central hypothesis of this proposal is that cyclic-di-GMP signaling via a network of ligand-responsive DGCs regulates biofilm formation across a number of microbes of importance in pathogenic, host-associated, and environmental contexts. We propose the following Specific Aims to test this hypothesis: AIM 1. Test the hypothesis that small-molecule ligands are critical for regulating the localized cdG network via receptor complexes in P. fluorescens. AIM 2. Test the hypothesis that discrete domain structures of LapA and MapA in Pfl contribute to their differential impacts on biofilm formation. AIM 3. Test the hypothesis that the sulfate-reducing bacterium LapD/LapG-like system controls localization of this intestinal bacterium’s LapA homolog.

NIH Research Projects · FY 2026 · 2022-01

ABSTRACT Durable immunity to cancer is sustained by memory T cells. In contrast to circulating memory subsets, which traffic in and out of the blood, tissue-resident memory (TRM) cells are transcriptionally programed for prolonged residence and recall function within tissue. Collaborative studies between our laboratories were among the first to identify a requirement for TRM cells in immunity to cancer. Using a melanoma-associated vitiligo (MAV) mouse model that closely mimics the vitiligo that develops in immune checkpoint inhibitor-treated melanoma patients who benefit from prolonged disease-free survival, we showed that skin TRM cells are necessary and sufficient for long term protective immunity against melanoma in the dermis. However, mechanisms for controlling TRM cell persistence and identity as well as the contribution of TRM cells to tumor immunity at sites of frequent metastasis remain unclear. In this application, we examine an unexpected mechanism for TRM cell maintenance in the skin and reveal a new subset of vitally important TRM cells that persist in tumor-draining lymph nodes. In the skin of mice with MAV, as well as melanoma patients with vitiligo, immunofluorescent imaging revealed that TRM cells form lymphoid aggregates containing large populations of CD11c-expressing myeloid cells. While prior work indicates that CD11c+ dendritic cells (DCs) are critical for initiating immune responses but dispensable for reactivating TRM we find that depletion of CD11c-expressing cells results in rapid disaggregation and loss of CD8 TRM cells in the skin. We further show that the CXCR6/CXCL16 axis is required for TRM cell persistence and tumor protection in the skin. These findings identify a critical requirement for CXCL16-expressing myeloid cells in coordinating the organization and retention of CXCR6-expressing TRM in the tissue, which will be examined in Specific Aim 1. The importance of the CXCR6/CXCL16 axis and persisting self antigen in controlling TRM cell function and plasticity will be tested in Specific Aim 2. Finally, parallel mechanisms will be explored in lymph nodes (LNs) where our preliminary studies led us to discover a novel population of tumor-specific T cells that is crucial for protection against melanoma growth in lymph nodes. The presence of LN TRM cells has not previously been shown in the setting of cancer. A role for APCs and chemokines in maintaining such responses is essentially unknown, and will be the focus of Specific Aim 3. This proposal will thus test the overarching hypothesis that tumor-specific TRM cells— both in skin and draining lymph nodes—rely on key interactions with APCs and chemokines for their proper positioning, maintenance, and anti-tumor function.

NIH Research Projects · FY 2025 · 2021-12

In the US, ~24 million persons live with COPD, half undiagnosed, and ~150,000 die of COPD annually. COPD causes over 700,000 US hospitalizations and costs nearly $50 billion per year. The human and financial burdens of COPD could likely be reduced if disease progression and other adverse events could be anticipated, enabling caregivers to focus finite resources on at-risk patients. We propose to create a decision-support tool that integrates biomedical informatics with advanced machine learning (ML) and deep learning (DL) algorithms to predict acute and chronic healthcare encounters (hospital admissions, readmissions, and ED encounters) and major disease progression events (home oxygen therapy) for outpatients with COPD. Such a tool would confer immediate clinical benefits and accelerate research on COPD disease progression and treatment. Predictive modeling is widely used to identify high-risk patients for care management in COPD and other disorders, with a strong emphasis on readmission risk. However, extant techniques are not sufficiently accurate and do not identify the specific nature of likely future medical events, estimate time-to-event, and specifically forecast medical encounters and disease progression events for individuals with COPD. Recent research in disease progression modeling support the application of DL and other ML methods to electronic health records (EHRs) to predict aspects of health history. EHRs contain both readily accessible structured data (e.g., lab results in well-defined fields) and unstructured texts such as physician’s notes. Unstructured texts contain a great deal of clinical information, but this information is laborious to access; impeding its routine use in research and the clinic. This has motivated attempts to use natural language processing (NLP) methods to automate annotation. We will apply NLP to identify symptoms, treatments, procedures, diagnoses, social risk factors, and functional status from clinical notes, expanding the data available from EHRs far beyond the usual coded variables. Also, and distinctively, we will carry out a stepped-wedge clinical implementation of the proposed predictive tool and evaluate its performance, a first for ML and DL prediction of COPD health events. Therefore, we propose four Specific Aims: AIM 1: Transform EHR data streams to provision patient-level feature sets for ML and DL consumption. AIM 2: Develop a set of ML and DL models to predict the time-to-event for home oxygen therapy initiation and healthcare encounters among patients with COPD. AIM 3: To develop and implement a prospective performance surveillance and calibration maintenance system to maintain the final Aim 2 model for each outcome. AIM 4: Evaluate adoption and usability of the DeepCOPD toolkit in near-realtime clinical use in two healthcare systems. The application is responsive to the NHLBI IDEA2Health (NOT-HL-19-712).

NIH Research Projects · FY 2024 · 2021-09

PROJECT SUMMARY Obesity-related cancers, type 2 diabetes mellitus and cardiovascular disease, are characterized by a chronic breakdown in metabolic functioning that impacts quality of life, physical functioning and longevity. Though obesity plays a pivotal role in the etiology of at least 13 cancer types, the traditional metric for measuring obesity, body mass index (BMI), is imperfect and may fail to identify a third of individuals at risk of these cancers owing to metabolic dysfunction. While accumulated cellular damage and abrogated resilience mechanisms are part of the natural aging process, damage accumulation and dysregulation of homeostasis mechanisms, potentially driven by metabolic dysfunction, may lead to accelerated biological aging that has recently been linked to cancer risk and survival. A better understanding of the relationship between metabolic health, regardless of BMI, with accelerated aging and cancer is needed to inform who to target for prevention efforts. The long-term goal of this application is to understand how metabolic dysfunction influences biological aging and risk of cancer, at all levels of adiposity, to inform interventions that prevent or delay these deadly diseases. The central hypothesis is that metabolic dysfunction, independent of obesity, is associated with accelerated biological aging and obesity-related cancers. Aim 1 (F99 phase) will leverage data from the Utah Obesity Study to measure the association between metabolic dysfunction (metabolic syndrome and diabetes) across BMI categories (i.e., “metabolic health phenotype”) and risk of developing obesity-related cancer (esophageal, gastric, colorectal, liver, gallbladder, pancreas, uterus, ovary, thyroid, meningioma, kidney, and breast cancers, and multiple myeloma). In the Women’s Health Initiative (WHI), diabetes status at cancer diagnosis will be measured in relation to cancer-specific and overall survival. This research will be extended in Aim 2 (K00 phase) where metabolic health phenotype will be studied in relation to accelerated biological aging and obesity-related cancer risk. In Aim 2a, data from the prospective WHI, Jackson Heart Study, Health and Retirement Study, Framingham Heart Study and others will be used to measure the extent to which accelerated biological age explains the association of metabolic health phenotype with obesity-related cancer risk. In Aim 2b, using data from The Cancer Genomic Atlas (TCGA) cohort, accelerated biological aging will be evaluated in relation to survival after obesity-related cancer diagnosis. The pre-doctoral to post-doctoral candidate will expand upon her didactic and experiential training in biostatistics, epidemiology, aging and epigenetics research both at the University of Utah, Huntsman Cancer Institute, and Yale School of Medicine. Practical training will be obtained in human metabolism, biostatistics, epidemiology, aging and epigenetics research. The proposed project will help to better identify those at risk of obesity-related cancers and support changes in clinical cancer management to support diabetes and accelerated aging prevention.

NIH Research Projects · FY 2025 · 2021-09

Project Summary /Abstract Intrinsically disordered proteins (IDPs), which lack a fixed three-dimensional structure under physiological conditions, represent ~40% of the human proteome, have crucial functional roles in a variety of biological pathways and biomolecular assemblies and are implicated in a large number of human diseases. As IDPs populate a dynamic conformational ensemble of rapidly interconverting structures in solution, and cannot be represented by a single dominant conformation, or even a small number of substantially populated conformations, they are not suitable targets for conventional structure-based drug design methods. If it becomes possible to target IDPs with small molecule drugs, the druggable proteome will be dramatically expanded and therapeutic interventions may become accessible for currently untreatable diseases The PI’s laboratory utilizes an integrated computational and experimental research strategy to combine state-of-the-art all-atom molecular simulations with experimental measurements from NMR spectroscopy and other biophysical experiments to obtain atomistic descriptions of the dynamic binding mechanisms of IDPs and uses insights form these binding mechanisms to predict and rationally design novel binding interactions. This proposal focuses on applying this integrated computational and experimental approach to elucidate the binding mechanisms of small molecule drug candidates that target that intrinsically disordered domain of the androgen receptor and have entered clinical trials for castration resistant prostate cancer (CRPC). These binding mechanisms will be used to inform the rational design more potent and selective androgen receptor inhibitors and more effective CRPC therapeutics . This proposal describes a remarkable opportunity to draw connections between molecular binding mechanisms studied by molecular simulations and NMR, biological activity observed in cellular assays, and clinical results from human CRPC drug trials. This proposal will initiate a sustainable long-term research effort to combine computational and experimental methods to study the dynamic interactions of IDPs in a variety of cellular and pharmaceutical contexts. This research effort will stimulate the development of robust platforms to integrate computational and experimental methods that will dramatically increase the number of proteins amenable to structural and mechanistic characterization and pharmaceutical targeting and will provide new avenues to therapeutic interventions in diseases associated with aberrant biological interactions of IDPs such as those mediated by biomolecular condensate formation and protein misfolding.

NIH Research Projects · FY 2025 · 2021-09

Abstract Cortical circuits, comprising specialized neuron subpopulations and their selective synaptic connections, send output to long-distance cortical and subcortical targets via two distinct classes of projection neurons: intratelencephalic (IT) neurons that project primary within and across cortical hemispheres, and pyramidal tract (PT) neurons that project to deep subcortical targets (e.g., the thalamus and brainstem). Cortical circuits are regulated by a variety of modulatory neurotransmitters, such as serotonin (5-HT) and acetylcholine (ACh), that optimize circuit performance for different cognitive tasks. Indeed, disruption of serotonergic or cholinergic signaling in the cortex impairs cognition and normal behavior, and both transmitter systems are implicated in a range of psychiatric diseases. Therefore, revealing the physiological mechanisms by which 5-HT and ACh influence cortical processing will enhance our understanding of normal cognition, and will advance the development of novel therapeutic strategies for psychiatric patients. In the mouse prefrontal cortex (PFC), 5-HT and ACh act via G-protein-coupled receptors to reciprocally regulate the postsynaptic excitability of IT and PT neurons. 5-HT promotes IT neuron output, but suppresses PT neurons. Conversely, ACh preferentially excites PT neurons, but has limited impact on IT neurons. Regardless of neuromodulatory state, action potential generation in IT and PT neurons requires excitatory synaptic drive that may also be regulated, at the presynaptic level, by 5-HT and/or ACh. Our pilot studies suggest that 5-HT and ACh act differentially to regulate key excitatory afferents to IT and PT neurons. This current project will test the overarching hypothesis that 5-HT and ACh bias the “throughput” of cortical circuits via coordinated pre- and postsynaptic regulation of specific combinations of excitatory afferent and cortical projection target neuron subtype. Our first aim is to map the relative targeting, and functional excitatory drive, of IT and PT neurons by key extrinsic excitatory afferents to the mouse medial PFC. Our second aim is to test for afferent-specific presynaptic regulation by 5-HT and ACh, thereby determining whether pre- and postsynaptic neuromodulation is coordinated to facilitate specific combinations of afferent input and cortical projection output. Our third aim is to test whether 5-HT and/or ACh regulate local circuit communication between IT and PT neurons in a manner consistent with the opposing postsynaptic impacts of these modulators on excitability. Completion of these aims will establish a rigorous, circuit-based framework for understanding cholinergic and serotonergic regulation of cognitive function, and will provide insight into how disruptions of neuromodulatory circuits contribute to psychiatric disease.

NIH Research Projects · FY 2025 · 2021-09

PROJECT SUMMARY Myelin has evolved to speed up, finely tune, and increase the metabolic efficiency of electrical signal transmission in the brain. In numerous human diseases however, myelin degenerates, ultimately resulting in devastating motor and cognitive impairment. Importantly, in order for tissue repair to proceed after myelin damage has occurred, the many layers of compacted cell membrane that constitute the myelin sheath must be rapidly and efficiently removed by resident phagocytic cells in the brain. Defective removal of these debris has been implicated in a number of degenerative conditions, including but not limited to, multiple sclerosis and aging, yet we know little about the cellular dynamics and molecular mechanisms governing these processes. In order to study these critical cellular events and answer questions centered on which cell populations are involved and what roles these different cell types play, we have developed advanced techniques for imaging and manipulating these discrete events in the live animal over a wide range of temporal scales from seconds to months. These techniques include intravital imaging of new combinations of fluorophore-based multicolor transgenic labels of distinct populations of neurons and glia together with label-free imaging modalities specific for compact myelin. In addition to these powerful labeling and optical imaging strategies, we have also developed a new technique for targeted induction of single-cell death, which we have recently established as a model of on-demand and titratable demyelination in the mouse cortical gray matter. Combining these techniques now allows dynamic investigation of demyelination and remyelination in the context of targeted genetic manipulations and animal models of human disease. Using these powerful tools this project will investigate three central aims. First, there is increasing evidence that in addition to microglia, the primary phagocytes of the brain, other resident glial cell types, namely astrocytes and NG2 glia, are also involved and play important roles in the phagocytosis and repair process. We will determine the precise contribution of each glial cell type in the dynamic detection and clearance of degenerating myelin debris. Next, we and others have shown the importance of phosphatidylserine receptors in the efficient detection and clearance of dying neurons and other cells in different organs. We will determine the role and consequences of both defective phagocytic receptors and debris digestion signaling on the dynamic response by NG2 glia to cortical demyelination and the resulting remyelination success and myelin patterning. Finally, there is evidence that neuronal activity and/or sensory experience can modify remyelination, but less is known about the roles of neuronal activity on phagocytic function in the context of demyelination. We will determine the consequences of bidirectional neuronal activity changes on the response by phagocytic cells to single-cell demyelination. Ultimately, these studies will reveal which cells are involved in myelin debris clearance, the role of major cell debris recognition pathways in successful clearance and repair, and how neuronal activity and sensory experience modify the response of phagocytic glia to a demyelinating event.

NIH Research Projects · FY 2025 · 2021-08

The impact of the gut microbiota on human health depends on the identity of the species therein. The mechanisms that lead to differences in microbiota composition between people is not well understood. We focus on interbacterial interactions between members of the dominant taxon in the gut of Western adults, the order Bacteroidales. These bacteria compete for space and nutrients via a molecular nanoweapon known as the type VI secretion system (T6SS). Toxic protein effectors delivered between adjacent Bacteroidales cells by the T6SS result in cell statis or lysis, and we and others have previously revealed that this competition results in strain-level differences in the microbiota through exclusion of target bacteria via killing. Since effectors can be delivered indiscriminately to kin cells, T6SS-encoding bacteria produce immunity factors that specifically neutralize cognate effectors. We have previously identified the pervasive presence of “orphan” immunity factors encoded within large genomic arrays within Bacteroidales genomes that lack cognate effectors. These acquired interbacterial defense (AID) systems render the T6SS ineffective through neutralization of cognate effectors and facilitate strain- exclusion from microbiomes. In this proposal, we seek to understand the mechanism by which orphan immunity genes are captured aggregated into the most common type of AID system, the recombinase-associated AID (rAID) system, using a powerful combination of bacterial genetics, biochemistry, metagenomics, and gnotobiology. We further seek to understand the regulation and biogeographical role of rAID systems through a hypothesized “competition-sensing” mechanism that involves a hybrid sensor-kinase pathway. Together, we aim to understand the impact of Bacteroidales defense against the T6SS on the gut microbiome.

NIH Research Projects · FY 2025 · 2021-08

Project Summary/Abstract The removal of toxic or unwanted cellular structures – such as protein aggregates, intracellular pathogens and damaged organelles – presents a fundamental challenge for eukaryotic cells. To eliminate these structures, which are too large and too complex for the proteasome, cells require an alternative degradation pathway, autophagy. The predominant view in the field is that all autophagy targets are degraded by a common mechanism. However, this understanding comes primarily from studies of only a few model substrates, with the unsubstantiated assumption that other cargoes behave similarly. To test this model, I developed a panel of cargo-specific autophagy reporters amenable to genetic screening, quantitative microscopy, and biochemical manipulation. By this approach, I discovered that autophagy cargo (and their receptors) are instructive for autophagy, such that different cargoes can induce autophagy via different mechanisms. In the process, I discovered a new cohort of proteins that facilitate cargo selection and autophagosome initiation. Going forward, the broad goal of our work is to apply our receptor-centric paradigm to reveal new autophagy mechanisms, dissect the physiological impact(s) of autophagy, and elucidate heretofore unknown lysosomal trafficking pathways. We expect our studies to reshape the traditional view of how autophagy factors function into a more complex web of overlapping mechanisms that ensure robust cytoplasm-to-lysosome delivery. In doing so, we will reveal conserved principles of autophagy and inform our understanding of autophagy dysregulation in disease.

NIH Research Projects · FY 2025 · 2021-06

We propose to establish a best practice for implementation of a multifaceted approach designed to attenuate perioperative ESKAPE (Enterococcus, S. aureus, Klebsiella, Acinetobacter, Pseudomonas, Enterobacter spp.) transmission and associated surgical site infection (SSI) development. Perioperative ESKAPE transmission (inoculum) contributes to the development of surgical site infections (SSIs) which affect 3-5% of patients undergoing surgery. SSIs increase patient morbidity, prolong hospitalization, increase the risk of intensive care unit admission, and increase the risk of death 2-fold. ESKAPE pathogens are particularly problematic. Sustained reductions in epidemiologically-related, perioperative S. aureus transmission events achieved via a multi-faceted approach including surveillance feedback optimization resulted in substantial SSI reductions (88% decrease). An evidence-based approach for attenuation of the perioperative bacterial inoculum2 must integrate improvements in provider hand hygiene, intravascular catheter design/handling, environmental cleaning/organization, and patient decolonization. In this application, we propose a type 1 hybrid effectiveness-implementation using a 2x2 factorial cluster-randomized design guided by RE-AIM. We aim to identify a best practice for addressing the perioperative ESKAPE inoculum. We will examine the relative effectiveness of increased site awareness and commitment to generating improvements via technical assistance (TA), team coaching implementation of an evidence-based set of interventions (EBIP), and TA or EBIP with ESKAPE transmission surveillance feedback. Our strong preliminary data from a randomized trial implementing a multifaceted program with surveillance has demonstrated substantial and statistically significant reductions in transmission of S. aureus and 90-day SSIs and recently reproduced our randomized trial findings in an additional external site. Therefore, our prior research strongly suggests that the proposed research should be done and justifies scaling up to dissemination and implementation. Our exceptional multidisciplinary team is well equipped to successfully complete the proposed trial and aims. In the proposed trial guided by RE-AIM, we will advance scientific knowledge and inform future dissemination and implementation by investigating how best to scale-up an already successful multifaceted approach to national dissemination through either TA or EBIP with or without surveillance. We will conduct a rigorous cost- effectiveness analysis including evaluation of net cost savings. The proposed trial guided by RE-AIM (Aim 1), the addition of 1-year follow-up for sustainability (Aim 2), and cost-effectiveness analysis (Aim 3) will provide the essential scientific knowledge to adopters and organizers to be able to reproduce the most effective delivery method of our interventions to their local setting in addition to informing our investigative team which approach to scale-up to reach national dissemination.

NIH Research Projects · FY 2025 · 2021-06

Abstract. Cystic fibrosis (CF) is a fatal genetic disease characterized by overproduction of mucus in the lungs followed by chronic lung infections. Conventional wisdom has been that most CF lung infections involve a single dominant organism, most commonly the pathogenic bacterium Pseudomonas aeruginosa. Advances in culture-independent techniques have revealed that CF lung infections are rarely mono-microbial and instead usually involve complex microbial communities, yet the interspecies interactions that drive these communities are poorly understood. Furthermore, numerous studies have demonstrated that polymicrobial infections are more difficult than mono-microbial infections to eradicate with antibiotics, leading to the concept of recalcitrant communities. The mechanisms underlying recalcitrance are thought to involve synergistic interactions between community members, but very little data are available to understand this phenomenon. Combined with the realization that many CF patients respond poorly to available antibiotic regimens compels a more detailed understanding of interspecies interactions and their impacts on antibiotic recalcitrance to improve the treatment of CF infections, as well as other polymicrobial diseases. Here, we combine big-data bioinformatics, in silico computational modeling and in vitro culture experiments to gain insights into the metabolic interactions that drive CF disease outcomes and antibiotic recalcitrance. The research will leverage an available data set of hundreds of CF patient samples that provide both bacterial composition data and clinical metadata, including measures of lung function. These samples will be clustered according to their measured compositions and metabolic capabilities predicted through computational metabolic modeling to test the hypothesis that the vast complexity of these many bacterial communities can be collapsed into a small number of model communities that capture most of the observed metabolic variability. These computational predictions will be tested by developing in vitro cell culture models that recapitulate the most important metabolic features of the in vivo polymicrobial communities (Aim 1). By applying bioinformatics and modeling to the same clinical data, we will test the hypothesis that community metabolic features drive disease outcomes and the virulence potential of these communities (Aim 2). Finally, we will interrogate the clinical data and in vitro communities to test the hypothesis that community metabolic features drive antibiotic recalcitrance and differentiate community responsiveness to antibiotics according to these metabolic features (Aim 3). Our research will yield novel insights into how complex polymicrobial communities are compositionally structured, interact metabolically, contribute to disease and respond to antibiotics. Moreover, the research will validate in vitro models that offer the potential for development of novel antimicrobial strategies to better treat chronic, polymicrobial infections in CF and other diseases. Our transdisciplinary team offers the necessary expertise in bioinformatics, computational modeling, microbial physiology and CF polymicrobial infections to tackle this complex problem.

NIH Research Projects · FY 2025 · 2021-05

Up to 60 chemical elements can be detected in the human body, about 25 of which are essential to life. Elements such as iron (Fe), zinc (Zn), copper (Cu), and selenium (Se) are constituents of enzyme cofactors required for a wide range of processes from cell cycle regulation, reproduction, structural repair, and immunity. Dysregulation of these elements is a hallmark of disease. For instance, aluminum (Al), Fe, Cu and Zn accumulation in amyloid plaques in the brain are associated with Alzheimer’s disease and dysregulation of Cu causes Menkes and Wilson’s disease. Chemical elements are also diagnostic tools and chemotherapeutics; gadolinium (Gd) is the most common magnetic resonance imaging contrast agent, metal-base nanoparticles show promise for targeted drug delivery and platinum (Pt) and arsenic (As) compounds are used in cancer treatment. Invariably, knowing the spatial distribution of an element in an organ, tissue or cell is essential to fully investigate mechanisms of disease or the efficacy of drug delivery. Hence, elemental imaging at the micron scale is essential to further the NIGMS goal to support basic research that increases our understanding of biological processes and lays the foundation for advances in disease diagnosis, treatment, and prevention. Current analytical resources for biomedical elemental imaging are either over-subscribed (synchrotron X-ray fluorescence), cover only a limited range of elements, are performed under a vacuum or not readily scalable to imaging at the mm scale. Laser ablation inductively coupled plasma mass spectrometry (LA-ICP-MS) is a laboratory-based elemental imaging (elemental imaging) technique that it is increasingly being applied to biomedical applications with comparable or lower detection limits than other elemental imaging techniques and, potentially, easier access and operability. To date there are no elemental imaging LA-ICP-MS user resources in the US. The Dartmouth Trace Element Analysis Core is an established and highly regarded shared resource that supports researchers throughout the US, is staffed by experienced researchers in elemental imaging and ICP-MS, uses state-of-the-art instrumentation and has excellent links to industry partners. Our aims are to expand upon our current ad hoc elemental imaging service to establish an elemental imaging user facility; the Biomedical National Elemental Imaging Resource (BNEIR), which will accelerate and simplify access for biomedical researchers to instrumentation, expertise, web-based and in-person training, after-visit support, software and will foster a dynamic community for elemental imaging users. We will actively promote the elemental imaging within the larger NIGMS community of scientists through webinars, seminars, attendance at national and international meetings and actively seeking out NIH researchers currently pursuing relevant research projects.

NIH Research Projects · FY 2025 · 2021-04

PROJECT SUMMARY A research program will be undertaken to study SMARCB1-regulated SWI/SNF complex function in malignant rhabdoid tumors. Over the past few years, major cancer genome sequencing studies have identified frequent inactivating mutations of mammalian SWI/SNF chromatin remodeling complex in over 25% of human cancers. Despite the prevalence of SWI/SNF mutations, the contributions of these alterations to tumorigenesis remain unclear. SMARCB1, a core subunit of the complex, was the first subunit linked to cancer when it was found to be recurrently mutated in almost all cases of malignant rhabdoid tumors (RT). While studies using genetically engineered mouse models have firmly established SMARCB1 as a tumor suppressor, the epigenetic mechanism by which SMARCB1 mutation drives tumorigenesis remains elusive. The central hypothesis is that the biochemical diversity of the SWI/SNF complex composition determines its function in proper chromatin targeting to maintain discrete chromatin landscapes and transcriptional programs. Mutation of SMARCB1 leads to aberrant intra- and inter-complex protein-protein interactions that influence chromatin targeting, which consequently alters chromatin landscapes and higher-order structures, and promote an oncogenic epigenome and transcriptome. The objective is to determine the role of SMARCB1 in SWI/SNF subcomplex formation and the impact of these dynamics on chromatin targeting and higher-order chromatin structure. Our ultimate goal is to determine the fundamental epigenetic mechanism(s) by which loss of this chromatin regulatory subunit drives rhabdoid tumor development. To test this hypothesis, we will use a combination of cutting edge biochemical, molecular, and high-throughput sequencing technologies to move forward the program through the following specific aims: Aim 1, Determine SMARCB1 regulated SWI/SNF subcomplex assembly dynamics and identify SWI/SNF interactome in rhabdoid tumors; Aim 2, Determine the chromatin targeting activities of SWI/SNF subcomplexes and their relationship with other epigenetic regulators in rhabdoid tumors; Aim 3, Define higher- order chromatin structures regulated by the SWI/SNF complex in rhabdoid tumors.

NIH Research Projects · FY 2025 · 2021-04

PROJECT SUMMARY The goal of this proposal for the Biobehavioral Research Awards for Innovative New Scientists (BRAINS) program is to understand typical and atypical adolescent development of reward processing and impulsive behavior. These complex phenotypes are found in many psychiatric disorders including attention deficit hyperactivity disorder, borderline personality disorder, and schizophrenia. Adolescence is a sensitive period for the emergence of dysregulated reward processing and disordered impulsivity, and for the development of underlying neural circuits thought to be responsible. However, it is unclear what factors push development towards pathological trajectories and it is unknown how pathology is encoded by changes in neural circuits. The Adolescent Brain Cognitive Development (ABCD) longitudinal study of human brain and behavior is underway to identify factors in adolescence that predict impulsivity and other reward-related phenotypes. Similar longitudinal data from mice are necessary to allow molecular, cellular, and circuit-level interrogation of adolescent development. This knowledge is critical for targeted interventions to alter and prevent developmental pathology. This proposal develops a framework for mouse ABCD studies. We use an innovative approach to measure complex behavioral phenotypes in the homecage that allows for testing on a timescale compatible with assessing the dynamic changes during adolescence. This proposal focuses on the role of serotonin modulation of corticostriatal projections in driving adolescent maturation of reward processing and impulsivity. Using transgenic mouse lines for cell-type and time period-specific manipulations, we will investigate circuit-level mechanisms of serotonin modulation of adolescent developmental trajectories. The high-risk, high-reward use of in vivo calcium imaging in adolescents will uncover the single cell and ensemble- level changes occurring in the adolescent brain that supports adolescent behavioral maturation. Using microendoscope technology we will identify neural changes at the cellular level throughout adolescence, and define a trajectory of pathological development. These studies will point to a timeframe and mechanism for targeted prevention and treatment of developmental pathology related to reward processing and impulsivity. Our results will inform refinement of pharmacotherapies aimed at modulating serotonin signaling in adolescents for the treatment of depression, anxiety, and obsessive compulsive disorder. This research project is highly appropriate for BRAINS funding because it directly addresses two key objectives in the NIMH Strategic Plan and applies novel methods and techniques to advance our understanding of the major conceptual question of what drives adolescent maturation. As an early career investigator, funding for this ambitious proposal, which I’m uniquely equipped to carry out, would allow me to launch an innovative basic research program aimed at understanding the atypical development of reward processing and impulsivity.

NIH Research Projects · FY 2025 · 2021-02

ABSTRACT Surgical resection is a fixture in the treatment of intracranial tumors, and there is mounting data indicating that overall and progression-free survival improve for gross total resection compared to subtotal resection. The current standard of care for managing intracranial tumors relies heavily on MRI of gadolinium-based contrast agents (Gd-MRI), which plays a central role in diagnosis, surgical planning, intra-surgical guidance, and follow-up monitoring. During surgery, patients are spatially registered to the pre-operative MRI and MRI-derived tumor contours projected over the visual field within the surgical microscope to guide resection. Despite the widespread deployment of these sophisticated tools in surgery, subtotal resection rates remain stubbornly high. The primary culprits include difficulty in identifying tumor visually and the diminishing accuracy of the pre-op registration due to brain deformation as the surgery progresses. In this context, expansive efforts have sought to alleviate these shortcomings, including the use of intra-operative stereovision and/or ultrasound with brain deformation models to update the pre-op MRI and the use of fluorescent agents to label tumor in the visual field. Although promising, both of these approaches have known shortcomings. Specifically, the data sources used for updating pre-op MRI are only surrogate correlates with MRI, and most current fluorescence guided surgery (FGS) efforts focus on targeted agents designed to mark molecular features of tumor cells, which have shown high intra-patient/tumoral heterogeneity. This project aims to solve both of these shortcomings directly by leveraging the existing clinical understanding of Gd-MRI in managing intracranial tumors. Specifically, we will identify and evaluate fluorescent agents that mimic the kinetic behavior of conventional MRI-based contrast agents to guide intracranial tumor surgery. This approach will transfer the well-understood behavior of Gd-MRI directly into the visual field, enable rapid, intra-surgical administration of the agent, and provide an ideal data input for updating of pre-op MRI during surgery. Our approach is premised on compelling preliminary data in small animal glioma models showing highly correlative uptake between Gd-MRI and several untargeted optical agents. To advance this strategy we will, (1) rigorously validate these results and examine additional optical agent candidates using MRI and our custom hyperspectral whole-body imaging cryomacrotome, (2) establish concordance between candidate optical agents and Gd-MRI in a new porcine glioma model using our intra-operative MRI facility and FGS instruments, and (3) assess the capacity to use the optical agent data to update the pre-op MRI. We will also quantitatively compare uptake of the candidate agents, Gd-MRI and ALA-PPIX, the current standard for FGS of glioma. Completing the aims of this project will establish the optical analog strategy as a compelling approach for surgical guidance and lay the groundwork for clinical translation.

NIH Research Projects · FY 2026 · 2020-12

Project Summary/Abstract The mononuclear phagocyte (MP) system plays a fundamental role in both innate and adaptive immunity. It includes three broad classes of MPs extensively characterized in the mouse: (1) macrophages, including alveolar macrophages, Langerhans cells, and three distinct subtypes of interstitial macrophages; (2) tissue-trafficking monocytes; and (3) dendritic cells (DCs), which fall into two main types (DC1 and DC2), though DC2 can be further subdivided. All these MPs, except AMs and LCs, which are unique to lung and skin, reside in multiple organs, including the heart, skin, liver, and gut. MP subtypes demonstrate a clear division of labor during innate and adaptive immunity with little to virtually no functional redundancy, which means that specific interactions among them are crucial for optimal immune responses against viral, bacterial, and fungal infections. Currently, however, multiple fundamental gaps for the identification and understanding of how these MPs function in human organs limit our ability to develop prevention and treatment strategies across diseases. This project will investigate cross-species and cross-tissue homologies at the cellular, gene expression and functional levels. We will obtain fresh human and mouse tissue from multiple organs (lung, skin, and their draining lymph nodes), and employ three broad approaches. First, we will use both bulk RNA sequencing (RNA-seq) and single-cell RNA sequencing (scRNA-seq) to identify cross-species and cross-tissue homology. RNA-seq provides sequencing depth (i.e., whole-transcriptome coverage), and scRNA-seq provides the ability to confirm bulk homologous MP subtypes and examine the heterogeneity within previously defined MP subtypes. Thus, bioinformatics analyses will identify clusters of homologous MP cell types and align them across species. Second, for each cluster identified, we will identify genes conserved across species and tissues, and those that are unique to a given homologous MP subtype, termed marker genes. The results of these analyses will provide specific genetic markers for human MP subtypes and genetic treatment targets. Broadly speaking, there are two categories of key marker genes we will functionally investigate: those conserved in human-mouse MP counterparts that (1) have been well-defined in mice, but not previously investigated in their human counterparts; and (2) not well-defined or extensively studied in either species. Third, we will undertake a rigorous functional validation of the key genes identified in human-mouse MP counterparts. This includes (a) in-vivo murine models with selective depletion of specific genes using transgenic and conditional knockout (KO) mice; (b) in vitro model systems for human MPs, including assays for antigen acquisition and processing, cellular interactions, and induction of adaptive immune responses; and (c) create time-lapse videos with cellular-level microscopy for functional and morphological characterization.

NIH Research Projects · FY 2026 · 2020-09

U.S. adult rates of cannabis use and DSM-5 Cannabis Use Disorder (CUD) have now reached historic peaks, medical cannabis use for pain is legal in 40 states, and the FDA has recommended re-classifying cannabis from Schedule 1 (no medical use, high abuse potential) to Schedule 3 (use for pain, moderate-low dependence potential). These changes have prompted calls to re-examine and improve the DSM-5-TR CUD criteria, which have not been updated in over 10 years. Opioid use disorder (OUD) is a useful reference for re-examining CUD because pain is the most common medical reason for using opioids or cannabis. Earlier, we showed excellent reliability and validity of 3 conceptual models of prescription opioid use disorder (OUD-P), with strongest validity evidence for the ‘fully-adjusted’ model that did not count DSM-5 criteria as positive if they occurred only when opioids were used as prescribed (under medical supervision; i.e., therapeutic use). This framework is our starting point to study potential improvements in CUD diagnostic criteria. However, unlike opioids, cannabis is ‘authorized’, not prescribed; medical supervision is rare; and dosing seldom specified. Knowledge gaps about key medical cannabis behaviors (e.g., efforts to get medical supervision, how therapeutic doses are determined) impede creating guidelines to assess therapeutic cannabis use. In R01DA050032, we used cognitive interviews and online surveys to produce the Cannabis Exposure Index (CEI), a new measure of mg/THC (i.e., cannabis dose). We propose a 5-year renewal to use these methods to compare reliability and validity of 3 CUD models. 1) Unadjusted (DSM-5-TR); 2) Physiologically-adjusted (tolerance, withdrawal criteria not counted towards a CUD diagnosis if they arose solely from therapeutic cannabis use) and 3) Therapeutically-adjusted (novel, fully- adjusted model: any CUD criteria occurring solely during therapeutic use not counted towards a diagnosis). To test the models, we must first fill knowledge gaps on medical cannabis supervision/dosing in order to create guidelines to assess therapeutic use. We will then adapt our diagnostic measure, PRISM-5-OP, into a cannabis version (PRISM-CUD) to test if physiologically- or therapeutically-adjusted CUD models are improvements over DSM-5-TR CUD. Given FDA recommendations, we focus on cannabis for pain, but also explore sleep and anxiety. All participants will have chronic pain and use cannabis daily/near-daily. AIM 1. Fill knowledge gaps about medical cannabis (e.g., efforts to obtain medical supervision or dose information; self-efforts to determine right dose) via online survey (n=1,000). AIM 2. Use findings to create guidelines to differentiate therapeutic from other use for PRISM-CUD measures of the CUD models; pilot and conduct cognitive interviews; refine items. AIM 3. Using PRISM-CUD, compare convergent and discriminant validity of the 3 CUD diagnostic models in online sample (n=3,000), re-testing 400 to determine reliability. Hypothesis: Validity in capturing true cases of CUD will be ordered as: Therapeutically-adjusted>Physiologically-adjusted>Unadjusted. We aim to update and improve validity of CUD diagnoses, thereby improving public health in many areas that require such diagnoses.

NIH Research Projects · FY 2024 · 2020-09

A primary driver of immune deficiency caused by HIV is the destruction of T cells, which if left untreated results in AIDS. Depression of cellular immunity results in a failure to control pre-existing virus infections, such as those by the human gammaherpesviruses: the Epstein-Barr virus and the Kaposi's sarcoma-associated herpesvirus. In some AIDS patients this results in severe disease, due to a failure to control virus-infected cells. Disease is a consequence of infection of B cells that harbor latent infection in the absence of virus replication. Previous work has shown the memory CD8 T cell response is the most important component of immune surveillance that controls latently infected cells in healthy patients. Therefore deeper understanding of CD8 T cell-mediated immune surveillance can help us understand how this response fails in AIDS patients, promoting development of strategies to restore immune surveillance to prevent gammaherpesvirus-associated diseases. This proposal will build on the novel finding that the BTB-ZF family transcription repressor Zbtb20 is essential for effective immune surveillance against murine gammaherpesvirus-68 (MHV-68). This rodent virus has proven to be an excellent model for virus-immune interactions, recapitulating many of the immune mechanisms used to control AIDS-relevant gammaherpesviruses. Preliminary data show the absence of Zbtb20 prevents the generation of cells with an effector / effector memory transcriptional signature. In addition rates of both glycolytic and mitochondrial metabolism were aberrantly elevated in Zbtb20-deficient CD8 T cells, indicating an important role for Zbtb20 in regulating immunometabolic status appropriate for the differentiation state of the T cell. This is critical, as it is clear that the metabolic state of the T cell is a critical driver of differentiation to memory cells, but very little is known about the metabolic state required for long-term immune surveillance. Our transcriptomic data identify key genes in glycolytic and mitochondrial respiratory pathways that are elevated in the absence of Ztbtb20. We will test whether dysregulation of these genes leads to attrition of immune surveillance, and if gene knockdown restores appropriate T cell differentiation and immunometabolism. Further experiments test the extent to which Zbtb20 is necessary for protection from disease associated with gammaherpesvirus infection in mice lacking endogenous T cell immunity, to mimic AIDS-defining immunodeficiency. These parameters are also tested using T cells genetically modified to restore effector memory differentiation or normalize metabolic rates. In summary, the significance is a mechanistic understanding of what is required for effective immune surveillance against an important class of AIDS-associated pathogen. Armed with this knowledge, we can design improved immune-based therapies to prevent serious disease in AIDS patients.

NIH Research Projects · FY 2024 · 2020-09

Engineered tissue holds tremendous promise for improving health and quality of life of patients suffering from trauma, illness, or organ failure. Realizing the full benefits in tissue engineering requires improved fundamental understanding of homeostasis, metabolism, inflammation, and nutrient transport in engineered tissue, coupled with reliable and integrated quality control during the manufacturing process. By virtue of being modular, portable, capable of operating in real-time environments, as well as being amenable to non-invasive and label-free formats, a chemical quality control based on electroanalysis offers one plausible solution to this challenge. However, current electroanalytical devices do not allow for selective in-situ continuous chemical monitoring and reporting of performance in engineered 3D tissue scaffolds within enclosed bioreactors. Enabling the study of chemical processes of engineered tissue requires radically new sensing materials with improved chemical sensitivity, selectivity, chemical stability capable of straightforward integration with 3D tissue scaffolds. The overarching goal of this research is to develop conductive metal-organic frameworks (MOFs) and covalent organic frameworks (COFs) as multifunctional sensing materials with broad potential utility in electroanalysis. The proposed technological approach to chemical detection offers unprecedented ability to generate atomically-precise electronic materials and devices with chemically-tunable electroanalytical performance. This MIRA application leverages bottom-up synthesis and self-assembly to develop sensitive and selective non-enzymatic porous working electrodes for gasotransmitters (CO, NO, H2S), nutrients and metabolites (glucose and lactate), and neurochemicals (ascorbic acid, uric acid, dopamine, and serotonin). The research plan implements a multidisciplinary approach comprising chemical synthesis, spectroscopic characterization, device integration, and electroanalysis to achieve three hierarchical levels of chemical control in molecular engineering of framework materials for chemical detection: (1) Atomic-level control of host-guest interactions through solvothermal synthesis and self-assembly; (2) Nanoscale control through morphological tuning of surface electrocatalysis; (3) Epitaxial control of electrochemical interfaces within solid-state, porous, and flexible devices. Conceptual and technological advances emerging from this work will serve as a vehicle to develop the proposed materials into novel components of future electroanalytical devices with transformative potential in tissue engineering, biomedical analysis, and patient- centered mobile healthcare.

NIH Research Projects · FY 2024 · 2020-09

Abstract POWERED The proposed Program for Oncology Workforce Education and Research at Dartmouth (POWERED), will identify, recruit and train underrepresented minority undergraduates for independent careers in cancer research with specific attention to related scientific and life/career skills. Qualified students will be from groups underrepresented in biomedical research. They will receive a holistic multi-modal two-year intensive training experience spanning the summer following sophomore year through their senior year. We propose building on the foundation we have developed with three years of an NCI-funded P30 “CURE” supplement that introduced undergrads from underrepresented minorities to oncology research at Dartmouth. POWERED is a two-year program including two 8-week summers (16 weeks total) of full-time mentored research at Dartmouth's Norris Cotton Cancer Center (NCCC) boosted by 4 semesters (16 weeks each) of part-time coordinated scientific research at their home institutions. Considerable effort will be made to customize each participant's program according to their scientific interests in cancer. Each student will have a mentor at their home laboratory and at NCCC. Together, with the PI, they will form Individual Mentoring Team(s) to track the progress of each student. Faculty from colleges and universities throughout New Hampshire that make up New Hampshire INBRE, (NH- INBRE Idea Network of Biomedical Research Excellence), will recruit and nominate promising underrepresented minority students to apply and subsequently arranged for mentoring the selected students as research trainees at the home institution. A specific emphasis in the senior year is securing post-graduation work or training in cancer research or a related biomedical field. A set of Educational Tools to achieve the Specific Aims will be developed and evaluated using formative and summative measures to allow for dynamic change as appropriate to meet the Specific Aims and dissemination to other educators.

NIH Research Projects · FY 2024 · 2020-08

PROJECT SUMMARY Comorbidities between different mood disorders, as well as those with other syndromes such as schizophrenia, PTSD, and OCD suggest the possibility of shared underlying mechanisms that could inform development of new treatments. However, a rate-limiting step in uncovering such mechanisms continues to be the difficulty of studying these disorders in laboratory animals: it may never be possible to look at the behavior of an animal and know that it is experiencing a mood disorder. To side-step this fundamental challenge, we take a different approach: we will establish a neural assay of negatively biased thought. Many mood disorders are characterized by biased information processing, including recall biases for negative past events (in major depression) and biases towards generating negative future scenarios (in generalized anxiety disorder). Cognitive process theories of learning, memory and decision-making indicate that such biased thought may not only be a signature of mood disorders, but in fact causally contribute to it. This work will develop a neural assay of negatively biased thought in mice by focusing on sharp wave-ripples (SWRs): bursts of internally generated, synchronous neural activity in the hippocampus that can depict past or upcoming experiences. SWRs are an attractive preclinical target because (1) SWRs are highly conserved across mammals including rodents and humans, making it likely that results will translate, and (2) SWR activity can be decoded to reveal their content, such as retrieval of specific prior experiences. This implies that we can establish an objective neural assay for negatively biased thought: given a positive and a negative experience, are both experiences retrieved equally often, equally effective downstream, or is there a bias? Aim 1 will optimize a behavioral task and neural assay to provide positive and negative experiences that can be neurally distinguished, yielding for individual subjects and individual recording sessions a SWR content bias for negative compared to positive events. Next, Aim 2 will administer drugs known to generate affectively biased information processing in rodents in humans (amphetamine and pramipexole) and determine their effects on SWR bias and behavior. Finally, we will probe the mechanisms that link SWR content to motivational/affective systems at the neural circuit level through their interaction with dopamine in Aim 3. Taken together, the proposed work is expected to establish SWR content as a candidate transdiagnostic process that is of broad relevance to a number of mental health disorders.

NIH Research Projects · FY 2024 · 2020-07

ABSTRACT The primary objective of surgical therapy for the treatment of patients with cancer is to remove all cancer cells from within the body, with the secondary objective of maintaining organ function. The primary pathological metric used to rate the success of a surgical procedure is evaluation of the surgical margin of the resected tissue specimen, post-operatively. This typically involves cutting the tissue into sections and microscopically exploring these tissue samples for the presence of cancer cells at the margins. Cancer cells noted at the margins represent Positive Surgical Margins (PSMs) and suggest that cancer cells were left in the body following the procedure. As a result, patients with PSMs are often exposed to noxious additional procedures to eradicate the cancer cells left behind including radiation, chemical, hormonal, and additional surgical therapy; these all have adverse morbidities that decrease a patient's quality of life. No clinical protocols are routinely used to intraoperatively assess surgical margin status during surgical procedures. Instead, margins are evaluated through microscopic assessment of the tissue following the procedure, when it is too late to provide additional surgical intervention. We aim to develop an intraoperative device able to assess surgical margin status so that the surgeons can extract additional tissues in real-time and ultimately decrease the rates of PSMs. While our technology can be applied for most cancer surgeries, we are focusing our efforts on prostate cancer as these are the highest incidence and cause of death for men and because patients with PSMs following these procedures have a much higher rate of recurrence than patients that have negative surgical margins. We have previously shown that the electrical impedance (a property that describes how easily electrical current passes through a tissue) of tissue is sensitive to a tissue's cellular arrangement and can be used to distinguish cancer from benign tissue in prostate. We have developed a prototype flexible endoscopic device capable of imaging the electrical impedance tissue during radical prostatectomy procedures using Electrical Impedance Tomography (EIT) techniques. This device makes focal measurements of margin status. Here we aim to take the significant step of constructing an optimized EIT device that can be deployed laparoscopically (e.g. prostate surgery) to provide an accurate method of intraoperatively identifying positive surgical margins. We aim to develop this device, develop intraoperative visualization strategies to help guide surgeons, evaluate the technology in an in vivo study, and validate the technology intraoperatively. By the end of this program we intend to have developed a low-cost, single use probe that can be deployed in a multi- center clinical trial to evaluate the efficacy of this technology for intraoperative surgical margin assessment.

NIH Research Projects · FY 2024 · 2020-07

Project Summary Inhibition of Wnt Receptor Activation by the Tumor Suppressor Adenomatous Polyposis Coli The long-term objective of this study is to investigate how the tumor suppressor Adenomatous polyposis coli (APC) inhibits the Wnt signal transduction pathway by regulating the Wnt receptor complex (signalosome) and to demonstrate how this can be exploited to target APC mutant colorectal cancers (CRCs). Wnt signaling is essential for intestinal stem cell maintenance, whereas aberrant activation of this pathway, which occurs most frequently through mutational inactivation of APC, triggers the development of the vast majority of CRCs. In the classical model for Wnt signaling, the sole role of APC is to destabilize the key transcriptional activator in the Wnt pathway, beta-catenin. However, our recently published findings reveal an additional and entirely new function – APC prevents the internalization and consequent activation of the signalosome, a novel role that is evolutionarily conserved. We have shown that: 1) inducible loss of APC is rapidly followed by ligand-independent signalosome activation; 2) depletion or antibody-mediated inhibition of LRP6 (a signalosome component) inhibits the stabilization of beta-catenin, the transcriptional activation of Wnt target genes, and the proliferation of APC mutant cells; and 3) in APC mutant cells, endocytosis of Wnt receptors is required for the aberrant activation of Wnt signaling. The goal of this project is to use in vitro, ex vivo, and in vivo approaches to gain a better understanding of how APC inhibits signalosome activation under physiological conditions and to determine how aberrant activation of the signalosome underlies the consequences of APC inactivation in tumors. The three specific aims are to: 1) elucidate the mechanism by which APC loss promotes signalosome assembly in CRC cells; 2) identify the APC mutant CRC cells most susceptible to LRP6 inactivation; and 3) test the efficacy of LRP6 inactivation on CRC tumorigenicity in vivo. Because the molecular mechanisms by which APC prevents the aberrant activation of Wnt signaling are important for our understanding of colorectal carcinogenesis, the knowledge gained from this study will aid in the development of new therapeutic strategies for the treatment of CRC and other Wnt-driven cancers.