University Of Nebraska Medical Center

NIH Research Projects · FY 2025 · 2021-04

Project Summary Multiple myeloma (MM) is an incurable bone marrow (BM) cancer characterized by the production of monoclonal protein (MP). Development of drug resistance and off-target effects limits the efficacy of currently available agents. Therefore, novel therapeutic strategies, including drug delivery strategies, are urgently needed. We have focused on the novel strategy of targeting the trafficking of MP in MM cells by inhibiting the enzyme geranylgeranyl diphosphate synthase (GGDPS). GGDPS inhibitors (GGSIs) disrupt Rab geranylgeranylation, which results in intracellular MP accumulation, ER stress, induction of all three arms of the unfolded protein response pathway and ultimately MM cell death. Our GGSI development efforts have focused on isoprenoid triazole bisphosphonates and our structure-function studies have determined that isoprenoid chain length and stereochemistry impact inhibitor potency as well as in vivo biodistribution. Preclinical studies with our lead GGSIs have demonstrated key drug-like properties, including metabolic stability, prolonged plasma half-life, systemic distribution, in vivo disruption of protein geranylgeranylation and anti-tumor efficacy in a mouse MM xenograft model. Dose-finding and toxicology studies revealed hepatic toxicity as dose-limiting with no effects on hematologic, renal, cardiac or neurologic function. Our preliminary studies revealed that altering the molecular weight of a hyaluronic acid (HA) polymer can limit hepatic uptake and enhance BM uptake and that MM cells readily take up HA. We therefore hypothesize that the therapeutic potential of GGSIs can be optimized via linkage of our GGSIs to HA polymers, thus enhancing delivery of GGSI to the BM and minimizing hepatic uptake and our preliminary studies support this hypothesis. To this end, we will synthesize novel GGSIs that are are based upon our lead compound but either modified at the α- position to allow conjugation to HA or linked to HA through phosphonate prodrug forms (Aim 1) We will perform detailed structural studies, including protein crystallography studies with bound inhibitors, to clarify the mechanisms by which these GGSIs interact with the target enzyme (Aim 1) which will aid in the design of future generations of GGSIs. We will determine the pharmacokinetic/pharmacodynamic profiles and biodistribution patterns of the novel linkable GGSIs and the corresponding HA polymer conjugates (Aim 2). We will investigate the mechanisms regulating GGSI hepatic uptake and toxicity. The efficacy of the lead GGSI-HA conjugates will be assessed in xenograft studies which model extramedullary and medullary disease involvement (Aim 3). In vitro and in vivo studies evaluating the combination of GGSI therapy with clinically used anti-MM agents will be performed. While our current focus is to develop GGSI therapy for treatment of MM, these studies have added significance because this novel approach of drug conjugation to HA to alter biodistribution could be applied to the delivery of other clinically relevant agents for treatment of MM.

NIH Research Projects · FY 2025 · 2021-04

PROJECT SUMMARY/ABSTRACT The SWI/SNF family ATP-dependent chromatin remodelers are multi-subunit protein complexes that dynamically position chromosomal nucleosomes to modulate DNA accessibility, transcription-factor binding, and cell-type-specific gene expression. SWI/SNF function is crucial at several stages of mammalian development, and recent human whole-genome and exome sequencing studies revealed striking mutational frequencies in genes encoding SWI/SNF subunits across a range of diseases - from neurologic disorders to over 20% of all cancers. Disease-associated SWI/SNF mutations often cause the loss of a protein subunit that further destabilizes the complex and results in altered subunit composition and functions. Of note, heterogeneity in SWI/SNF subunit composition is also observed naturally within and across cell types, hypothesized to result in complex- and tissue-specific SWI/SNF functions. Despite their importance, how these subunits determine the substrate preference and mechanistic functions of SWI/SNF complexes are not well understood, largely due to the lack of robust approaches to characterize where they bind within the genome and how they interact with chromatin genome-wide. Thus, the main goal of this proposal is to address this key knowledge gap. Since chromatin remodelers directly interact with nucleosomes, it is important to clarify remodeler-bound nucleosome compositions and structures. Standard biochemical characterization of soluble components purified from cellular extracts cannot determine the nucleosome structures associated with chromatin-bound complexes. It is also important to understand the kinetic parameters of remodeler-chromatin interactions, such as how fast a remodeler is recruited to its target sites, and how long does it remain bound at its genomic sites. The candidate proposes to address these questions by using structural and functional epigenomics approaches and live-cell single-molecule imaging, to characterize remodeler-chromatin interactions in the context of the complex and dynamic chromatin environment inside cells. Aim 1 is to determine the genome-wide occupancy of SWI/SNF complexes with distinct subunit compositions. Aim 2 is to determine a) the structures and histone composition of remodeler-bound nucleosomes genome-wide, and b) the kinetic parameters of remodeler-chromatin interactions in live cells. Aim 3 is to study the interaction of SWI/SNF with repressive chromatin. The mentored phase of this project will be completed under the sponsorship of Dr. Steven Henikoff at the Fred Hutchinson Cancer Research Center. The candidate will acquire training in live-cell single-molecule imaging under the supervision of Dr. Sheila Teves at the University of British Columbia. The proposed research and training will provide a strong foundation for the candidate to develop as an independent investigator, studying chromatin remodeling mechanisms and dynamics in the regulation of fundamental cellular processes.

NIH Research Projects · FY 2025 · 2021-03

Name of Principal Investigator: Sean N. Avedissian Project Summary I am applying for a Mentored Patient-Oriented Research Career Development Award (NIMH-K23) for training in patient- oriented research with the goal of development into a productive and independent clinical and translational researcher. As part of this award, I will complete a PhD in Clinical Translational Research (CTR) through the mentored scholar’s program at my institution, University of Nebraska Medical Center (UNMC). This PhD program does not compete with the K23. The members of my mentoring team are Drs. Courtney V. Fletcher, Howard Fox, Kimberly Scarsi and Susan Swindells, all recognized leaders in HIV with expertise in all aspects of this research proposal. Dr. Fletcher will serve as my primary mentor and will guide and facilitate continued training in HIV pharmacology as well as in responsible conduct of research (RCR). The overall aim of the didactic training/PhD program is to provide multidisciplinary didactic education and practical research grant training to junior faculty like myself, who intend to develop a career in CTR, so they may acquire the skills to design, implement, analyze and report ethically sound, extramurally funded CTR. The PhD program is intended to serve as a pathway to accelerate the career development of junior faculty members. The director of the CTR-MSP is Dr. Lani Zimmerman, Professor in the College of Nursing at UNMC. My research will focus on investigation of fundamental characteristics of antiretroviral penetration into the central nervous system (CNS) using in vitro and animal models, with confirmation of penetration properties in people with HIV, followed by development and translation of a new approach to assess the anti-HIV activity of an entire antiretroviral regimen (as opposed to just that of an individual drug). The long-term goal is to develop novel strategies to achieve complete viral cure in all reservoirs of people with HIV. This proposal is clinically significant because HIV is known to persist in the brain, even in patients receiving potent antiretroviral regimens, who are highly adherent and have undetectable plasma HIV viral load. Given advancements in HIV therapeutics, people with HIV are older due to longer life expectancies(1, 2). However, even with advancements in HIV therapeutics, the rates of HIV-associated brain disturbances continue to remain high(3). Emerging evidence suggests that older patients with HIV may be at an even higher risk of developing HIV-associated neurocognitive dysfunction due to age and viral burden in the CSF(4, 5). Thus, there is an unmet need for improved personalized medicine approaches directed towards maximizing antiviral exposures to decrease viral burden throughout the whole body. Importantly, the goals of my research address the neurologic complications focus component of the current NIH Priorities for HIV and HIV-Related Research which is to “Address HIV-Associated Comorbidities, Coinfections and Complications”. On completion of the award period, I will have acquired the expertise to conduct independent research in CNS focused HIV pharmacology. I will have completed in-depth training in HIV and CNS translational and clinical research, allowing me to become an independent clinical scientist and leader in the field of HIV viral reservoir related pharmacology.

NIH Research Projects · FY 2026 · 2021-02

ABSTRACT Our research program develops and applies innovative mass spectrometry technologies, bioinformatics tools, and methodologies to transform our understanding of cell surface proteins and glycans and answer outstanding questions in stem cell biology and cardiac pathology. Our analytical platforms promote the development of new reagents and strategies to improve the quality and homogeneity of stem cell-derived cardiomyocytes for research and clinical applications and the discovery of strategies to monitor and treat patients with advanced heart failure. Specifically, our approaches enable the identification, characterization, and quantification of cell surface glycoproteins and glycans from small numbers of human cells. To date, we have developed new markers for identifying cardiomyocytes with high specificity and selecting maturation stage-specific stem cell derived cardiomyocytes to enable reproducible assessment and isolation of functionally-defined cells. Applying our technologies to primary human heart tissue, we have begun to develop cell-type specific views of the cell surface proteome and glycome within normal and failing hearts. We have also developed innovative bioinformatics tools to inform our next level of technology development to enhance our capabilities and improve the speed and accuracy with which we analyze our mass spectrometry data. The proposed studies build on these experiences to: 1) develop the next generation technology that will provide unparalleled specificity regarding the molecular phenotypes presented at the cell surface, information that is not possible to obtain by any current method, 2) develop marker panels that enable the assessment and selection of chamber- and maturation-stage specific stem cell derived cardiomyocytes without genetic editing, 3) define the cell-type specific receptors, membrane- bound ligands and secreted factors present in the normal human heart and how they change in disease to provide new understanding of intercellular signaling and inform the development of remote sensing markers to benefit the care of patients with advanced heart failure. The impact from the proposed studies lies within future applications and mechanistic studies that will be possible based on the approaches and data that we generate. As our program evolves to pursue mechanistic and translational studies of the molecules revealed by our discovery efforts, we expect the outcomes of these studies will broadly impact the development of strategies to improve the quality of stem cell derivatives to promote their utility for drug testing, disease modeling, and therapeutic applications, inform the development of cell-type directed payload delivery systems and drugs that avoid cardiotoxic effects, and yield new strategies to assess and treat advanced heart failure.

NIH Research Projects · FY 2025 · 2021-02

Abstract Breast cancer (BC) brain/central nervous system (CNS) metastasis is associated with poor survival among U.S. women, with 30-40% of these cases reported as triple receptor-negative (TN) and epidermal growth factor receptor-positive (ErbB2+) BC subtypes. Despite the progress in the diagnostic and therapeutic management of BC, there is still a significant increase in brain metastasis (BM) cases, and the survival rate among these BCBM patients is very bleak. Therefore, there is an urgent need to identify primary molecular drivers of BCBM and novel predictive biomarker(s) for the early detection of BM. In this regard, our global transcriptomic analysis showed that levels of a secretory gel-forming mucin, MUC5AC, is significantly higher in the brain tropic (BT) cells than the parental BC cells. Additionally, an in silico analysis revealed significantly higher levels of MUC5AC in the archived BCBM tissues compared to the primary tumors. Most importantly, the augmented levels of MUC5AC were detected in the serum of BCBM patients compared to non-BM BC patients. These studies strongly suggest that MUC5AC could be a potential predictive biomarker for the early detection of BCBM. Furthermore, MUC5AC knockdown (KD) resulted in reduced motility, cell adhesion, and blood-brain barrier (BBB) transmigration in BT cell lines relative to controls. Importantly, MUC5AC KD cells showed diminished BM potential in an intracardiac mouse model. Our initial mechanistic studies on the MUC5AC-mediated BM showed an important role of the CD44 and cMET pathways in BT cells. MUC5AC interacted with CD44v6, a co-receptor for cMET, and co- localized with the activated form of cMET to establish BCBM. CD44v6 and cMET have been shown to preferentially enhance BM through a feed-forward loop using hyaluronic acid and hepatocyte growth factor pathways. We also observed robust expression of MUC5AC in BT cells in the presence of microglia/astrocyte conditioned media. Targeting MUC5AC with PLB-1001 reduces MUC5AC expression in BT cells. We hypothesize that “MUC5AC enhances BCBM through CD44v6/cMET-axis” and could thus be a useful marker to predict BCBM. In Aim 1, we will establish MUC5AC as a novel predictive biomarker for BM in high-risk BC patients, and examine whether high MUC5AC expression in primary tumors predicts BM, and correlates with response to therapy, overall survival, and relapse. Aim 2 studies will define the regulation of MUC5AC-mediated BM through the cMET/CD44v6/NF-κB-axis using preclinical mouse models. In Aim 3, we will use a BBB penetrable phase 1 tested cMET inhibitor alone or in combination with cisplatin or neratinib as novel therapeutic strategy for TN and ErbB2+ brain metastatic BC. Altogether, the proposed studies will establish MUC5AC as a novel predictive biomarker for high-risk BM and will help in developing preventive strategies for BCBM, which currently has no cure.

NIH Research Projects · FY 2025 · 2021-01

ABSTRACT Erythropoiesis, the production of red blood cells (RBCs), is fine-tuned to meet physiological demands. During regeneration or “stress erythropoiesis” caused by anemia, a subset of genes and proteins are upregulated in association with an increased rate of RBC production. A critical gap in understanding erythroid regeneration hinges on whether the activation of gene regulatory networks in erythroid progenitors (driven by transcription factors) are stress-specific or represent broader control mechanisms in other contexts. Moreover, the underlying etiologies in which the erythroid system loses its ability to regenerate in chronic anemias are often unclear. We discovered that the Sterile Alpha Motif Domain-14 (Samd14) gene is elevated in models of acute/chronic anemia. Samd14 is regulated by the transcription factor GATA2, which coordinates a network of genes with critical functions in hematopoiesis and hematologic disease. The GATA2-occupied Samd14 cis-element (Samd14-Enh) is required for survival in a model of hemolytic anemia, but dispensable for steady state erythropoiesis. Erythroid progenitors lacking Samd14-Enh have impaired c-Kit signaling, a quintessential pathway regulating hematopoiesis and erythropoiesis. These results reveal the involvement of a GATA2-Samd14-c-Kit regulatory axis in erythroid regeneration. Whereas SAM domain-containing proteins are involved in hematopoiesis and cell signaling, and several are upregulated in anemia, their mechanisms of action are not well understood. Our data suggests additional cohorts of enhancers with properties mimicking the Samd14-Enh are anemia-regulated. We hypothesize that Samd14 and additional GATA2 and Regeneration-Activated (G2R) enhancers control erythroid regeneration. In Aim 1, mechanistic analyses in human and mouse will define Samd14 requirements for cell signaling and survival of erythroid progenitors. Aim 2 will delineate a GATA2 and anemia-regulated (G2R) gene network governing a sector of the complex biology surrounding anemia responses. Approaches using primary human/mouse cells and innovative mouse genetic model approaches will test SAMD14 mechanisms (and other SAM domain proteins) in c-Kit signaling and erythroid regeneration. These aims will establish valuable contrasts between homeostatic and regenerative erythropoietic mechanisms, enhancer knockout and gene knockout phenotypes, and between functionally-distinct cis-elements which contain similar sequence and molecular properties. By elucidating a GATA2-Samd14-c-Kit axis in acute anemia, and global/locus-specific GATA2 mechanisms of Samd14-Enh-like cis-elements, we expect these studies will reveal fundamental gene regulatory mechanisms in erythroid regeneration with implications in hematologic disease, including anemias.

NIH Research Projects · FY 2025 · 2020-12

PROJECT SUMMARY Medulloblastoma (MB) is a pediatric brain tumor arising from the cerebellum. MB treatment is challenging due to diverse genetic make-up, resistance to chemotherapy, inefficient drug transport across the blood brain barrier (BBB) and drug induced neurotoxicity. Hedgehog (Hh) and IGF/PI3K signaling pathways regulate cell growth, cancer stem cell (CSC) proliferation, and tumorigenicity in MB patients. Hh inhibitors are effective initially to treat SHH-MB, but their repeated use develops chemoresistance due to mutations in SMOothened (SMO) but can be overcome by modulating GLI, which is downstream of SMO using SF2523, which is a BRD4/PI3K dual inhibitor and inhibits MYCN expression. In our preliminary studies, we synthesize SMO inhibitor 2-chloro-N1-[4-chloro-3-(2-pyridinyl) phenyl]-N4, N4-bis (2-pyridinyl methyl)-1, 4-benzene- dicarboxamide (MDB5). MDB5 and SF2523 effectively inhibited the proliferation of ONS-76 and HD-MB03 cells in a dose dependent manner, with significantly higher cell killing when these drugs were used in combination. Treatment of HD-MB03 cells with the combination of these two drugs showed significantly higher decrease in colony formation and cyclin D1 expression but higher increase in Bax expression, compared to individual drugs. We synthesized mPEG-b-PCC-g-DC copolymer, with 5.1±0.21 and 6.5±0.1% loading for MDB5 and SF2523 when formulated into nanoparticles (NPs). There was sustained drug release from NPs, wherein 100% of MDB5 was released in 50 h, but only 60% of SF2523 was released in 80 h. Targeted NPs were prepared by mixing COG-133-PEG-b-PCC-g-DC and mPEG-b-PCC-g-DC at 10/90, 20/80 and 30/70 ratios, with the highest cellular uptake at 30/70 ratio. Systemic administration of COG-133-NPs loaded SF2523 into orthotopic SHH-MB tumor bearing NSG mice resulted in significantly higher drug concentration in the brain at 6 and 24h post administration compared to non-targeted NPs loaded with this drug while systemic injection of free drug showed negligible drug concentration in the brain. Moreover, systemic administration of COG-133-NPs loaded with MDB5 and SF2523 resulted in decreased tumor burden compared to non-targeted NPs of MDB5 and SF2523 as determined by IVIS imaging, with no hepatic toxicity. Our hypothesis is that BRD4/PI3K and Hh signaling pathways exert control over CSC proliferation in SHH-MB and hence represents a target for therapeutic exploitation with small molecules which can inhibit these signaling pathways. Our specific aims are to i) evaluate the resensitization effects of MDB5 and SF2523 on SHH and MYC driven MB and patient derived xenograft (PDX) cells; ii) formulate MDB5 and SF2523 into NPs with COG-133 conjugation and determine biodistribution and systemic/organ toxicity; and iii) determine effects of targeted NPs loaded with SF2523 and MDB5 in SHH and MYC driven orthotopic, patient derived xenograft (PDX) and transgenic SmoA1 MB mouse models. Long- term significance. Successful completion of this project will provide a platform technology for treating SHH- MB and other brain tumors using this innovative NP-based combination therapy of Hh and BRD4/PI3K inhibitors.

NIH Research Projects · FY 2024 · 2020-08

Project Summary Despite the ability of combination antiretroviral therapy to dramatically suppress viremia, the brain continues to be a reservoir of HIV low-level replication. Adding further complexity to this is the co-morbidity of drug abuse with HIV-associated neurocognitive disorders and neuroHIV. Among several abused drugs, the use of opiates is highly prevalent in HIV-infected individuals, both as an abused drug as well as for pain management. It has been shown that both HIV/HIV proteins and abused drugs such as morphine contribute to neuroinflammation, which, in turn, involves activation of the inflammasomes. Mounting evidence has also shown that noncoding RNAs function as epigenetic and post-transcriptional regulators controlling vital cellular functions, including activation, thus altering the dynamics of various biological processes. While both HIV proteins and opioids have been shown to change the gene expression profiles of CNS cells, there is a gap of knowledge on the detailed molecular mechanism(s) underlying the co-operative effects of HIV Transactivator of transcription (Tat) protein and morphine on astrocyte activation. The central hypothesis of this proposal is that HIV Tat and morphine, via distinct regulatory mechanism(s) augment astrocyte activation leading, in turn, to exacerbated neuroinflammation. Our preliminary data on whole-genome bisulfite sequencing in the frontal cortices of SIV- infected rhesus macaques show increased DNA hypomethylation of the NLRP6 (NOD-like receptor family, pyrin domain-containing protein 6) promoter with a concomitant upregulation in NLRP6 expression. Additionally, exposure of human and mouse primary astrocytes to HIV Tat and morphine also resulted in decreased expression of microRNA-152 that was accompanied by increased expression of NLRP6. Interestingly, we also found that exposure of these primary astrocytes to either HIV Tat or morphine resulted in increased cellular activation with increased expression of proinflammatory cytokines (IL1β and IL18) via pyroptosis. Based on the central hypothesis and the reliable preliminary data, this proposal led to the following specific aims: Specific Aim 1: Determine the molecular mechanism(s) involved in microRNA-152 mediated NLRP6 inflammasome activation in HIV Tat and morphine-exposed astrocytes in vitro; Specific Aim 2: Determine the epigenetic mechanism(s) involved in promoter DNA hypomethylation of the NLRP6 in HIV Tat and morphine-exposed astrocytes in vitro; Specific Aim 3: Validate the combinatorial effects of HIV Tat and morphine on NLRP6 inflammasome mediated astrocyte activation, in vivo. Understanding the mechanisms responsible for astrocyte activation induced by HIV and morphine will set the stage for the future development of novel therapeutics aimed at dampening HIV and opiate-mediated neuroinflammatory responses.

NIH Research Projects · FY 2026 · 2020-07

Project Summary/Abstract: Over one million Americans suffer from Systemic Lupus Erythematosus, an autoimmune disease for which there is no cure. Systemic lupus erythematosus (SLE or lupus) is a chronic autoimmune disease. The disease presentation is heterogenous, women are nine times more likely to develop SLE than men, and lupus is significantly more prevalent in people of Asian, Hispanic, Native American, and African ancestry than people of European ancestry. Lupus is a significantly understudied disease. Monozygotic twin concordance is found to be as low as 25% and familial aggregation studies suggest that lupus results at least in part from genetic predisposition. Most experts in the field agree that gene-environment interactions are important for lupus development. Recent work from our laboratory suggests that aberrant DNA repair leads to the development of lupus in a mouse model of the disease. We originally showed that a mutation in the POLB gene in mice results in development of lupus as a result of defective VDJ recombination and somatic hypermutation. In collaboration with Dr. Lindsey Criswell we have now identified a large number of coding germline variants that are enriched in individuals with lupus. In preliminary research, we have demonstrated that mice harboring one of these variants within the mismatch repair pathway develop high levels of antinuclear antibodies and lupus-associated lung disease. We have shown that somatic hypermutation is abnormal in these mice and results in the production of autoantibodies. Our RIVER project is focused on providing mechanistic insights into the development of lupus as a result of gene-environment interactions. A challenge in the field is understanding how environmental exposures influence lupus development. We suggest that many previous analyses may be underpowered because the genetic predisposition factors of the individuals studied are likely to differ, and that genetic factors play a significant role in the response of the organism to the environment. Our approach to address this challenge is to construct mouse models of coding genetic variants in DNA repair genes that are significantly enriched in individuals with lupus. This will be followed by characterization of the disease pathologies emerging in these mice in the absence and presence of environmental exposures that are known to be linked to lupus development. We will then take a combined genetic, molecular, and biochemical approach to elucidate underlying mechanistic insights into the development of lupus. Our project has significant potential to uncover the genetic and environmental bases of lupus development and to yield paradigm-shifting results that will impact the treatment of this devastating disease.

NIH Research Projects · FY 2025 · 2020-03

The CoNDA Center at the University of Nebraska Medical Center (UNMC) is dedicated to advancing our understanding of neurocognitive impairments in both development and aging. As the prevalence of disorders like Autism Spectrum Disorder and Alzheimer’s disease continues to increase, the Center's mission becomes increasingly relevant. Building on the success of Phase I, Phase II of the Center's efforts aims to further bolster multidisciplinary research and collaboration. This will be achieved by building upon the following efforts: (1) recruiting highly qualified faculty, as top-tier talent is essential for driving innovative research forward. By bringing in exceptional researchers, the Center ensures a wide range of expertise to tackle complex questions in neurocognitive impairments; (2) maintaining a robust Administrative Core and mentoring program, which is crucial for supporting the development and success of faculty members, particularly our recruits. The mentoring infrastructure provides guidance, resources, and support to help researchers thrive in their roles; and (3) continuing to enhance CoNDA’s Research Cores. Investing in state-of-the-art scientific Research Cores is essential for expanding the Center's research capacity. By providing access to cutting-edge technologies for neuroimaging and behavioral assessment in both human subjects and animal models, the Center empowers researchers to conduct high-quality translational research. Through these initiatives, the CoNDA Center aims to advance scientific knowledge and support its researchers' career advancement. By fostering a collaborative and supportive environment, the Center positions itself as a leader in the field of cognitive neuroscience, with the potential to make significant contributions to our understanding of neurocognitive impairments across the lifespan.

NIH Research Projects · FY 2026 · 2019-09

Abstract The direct elimination of integrated replication silent human immunodeficiency virus type one (HIV-1) proviral DNA has not been achieved in an infected human host. A pathway toward success rests in surgical delivery to tissues and cell viral reservoirs. We posit that this can be realized through optimal viral suppression (by ultra- long-acting antiretrovirals, ULA ARV) coupled with tissue-cell targeted lipid nanoparticles (LNPs) for CD4+ T and myeloid cell delivery and enhancements of viral excision efficacy. Our goals are to suppress, target, deliver, and then eliminate integrated proviral DNA in target cells and tissue reservoirs by a novel delivery platform. This platform will be optimized in HIV-1 infected cells and then in infected animals (H. Gendelman). We will achieve maximal viral suppression by ULA ARV (B. Edagwa). The viral cell and tissue reservoirs targeting will be achieved by compositionally unique CRISPR-Cas9 and guide RNA (gRNA) encased LNP (CRISPR LNP) formulation (S. Panja). The brain delivery of CRISPR LNP will be achieved by microbubble- enhanced focused ultrasound (FUS) (L. Chang). With this objective in mind, we will optimize the LNP composition to achieve lymphoid tissue-specific biodistribution. The biodistribution will be accessed by radiomagnetic labeling of LNP using positron emission tomography and magnetic resonance imaging (MRI) bioimaging (Y. Liu). We will build on the tissue-specific LNP and establish a tissue and cell reserve targeted CRISPR LNP by decorating with C- X-C chemokine receptor type 4 (CXCR4) and/or C-C motif chemokine receptor 5 (CCR5) targeting ligands. FUS will ensure the CRISPR LNP biodistribution, most notably to the central nervous system. The research, taken together, seeks to determine how, where, and to what extent the viral editing CRISPR cargo will reach its destination (i.e., lymph nodes, gut, spleen, and brain). Therefore, we now have the tools in place for therapeutic viral excision to precisely eliminate integrated latent HIV-1 from the host genome (P. Dash and S. Gorantla). Our multi-exon gRNAs recognize a broad swath of viral strains, and CRISPR LNPs can bypass the viral vector delivery and eliminate induced immunogenicity and consequent toxicity. We posit that viral suppression by combinations of ULA ARV followed by tissue-cell targeted delivery of CRISPR cargos can achieve viral “elimination” in live infected mice. The cross-disciplinary LNP delivery platform will ensure the efficacy, safety, and rigor of HIV-1 viral elimination approaches.

NIH Research Projects · FY 2024 · 2019-09

Project Abstract While environmental entities instigate T1D development, their identity is unknown. Genetics greatly increases risk, but the discordance between identical twins proves an equally critical role for exogenous factors. To identify causal entities, sampling must occur before clinical disease develops. Although labor intensive and costly, prospective longitudinal testing is the only way to illuminate the mysterious events that precede disease. Results from our recent collaboration with the TrialNet Pathway to Prevention study point to a role for CMV infection in T1D development. We found a striking expansion of terminally-differentiated short lived effector CD8 T cells (SLEC) in seroconverted (AA+) at-risk subjects. The frequencies of SLEC are highest in seroconverted subjects that progress to disease. This SLEC subset is identical to the well-characterized CD8 response to CMV. Importantly, we found the SLEC expansion is strongly associated with CMV seropositivity. Our findings demonstrate that an expanded, exhausted antiviral response occurs one year before autoimmune T1D develops. We also determined that SLEC levels correlate strongly with IA-2 autoantibodies, demonstrating a connection between CMV and autoimmunity. T1D results from autoreactive CD8 T cell-mediated attack on pancreatic islets. These autoreactive T cells may arise as a byproduct of the anti- viral response. Our overarching hypothesis is that CMV is an important environmental factor accelerating T1D development in children with genetic risk. The objective of this application is to longitudinally probe the preclinical disease process and unravel the mechanisms connecting CMV and T1D. In Aim 1 we propose to derive kinetic measurements through a longitudinal study. We hypothesize that genetically susceptible pre-T1D children show increased levels of exhausted CD8 T cells prior to the development of autoantibodies and/or clinical autoimmunity. In aim 2 we will measure CMV replication and persistence in at risk subjects that develop autoimmunity. We hypothesize that the accumulation of exhausted CD8 T cells causes viral persistence. In aim 3 we will test mechanisms whereby the virus activates islet specific CD8 T cells. We hypothesize that CMV infection induces the expansion of islet antigen-specific CD8 T cells. An established connection between CMV replication and T1D will point to antiviral therapies as efficacious in subjects with high genetic risk. Furthermore, levels of CMV replication and SLEC represent interlocking biomarkers for stratifying subjects for antiviral and/or vaccine trials. Several CMV vaccines are currently being tested in the clinic. Our results will also facilitate the understanding of other CMV-related human diseases.

NIH Research Projects · FY 2026 · 2019-07

Abstract Long acting (LA) therapeutics offer real potential for improved regimen adherence and disease outcomes for chronic infectious diseases. For human immunodeficiency virus (HIV) infections only cabenuva has emerged as a sole complete LA regimen to affect long-term viral suppression. However, the requirements for every two- months dosing, injection site reactions, health care practitioner administration, high dosing volumes, and prolonged “tail phase” pharmacokinetic (PK) profiles are challenges for broad use. Ultra-LA (ULA) therapies are therefore obvious next step needs. Such ULA antiretroviral therapies (ART) would increase dose intervals to simplify treatment, limit emergence of viral mutations through improved adherence to therapy, and synchronize regular six-month lab tests clinic visits with dosing. In our prior funding cycle we developed then licensed one of the “first” ULA antiretrovirals. It is a monomeric stearate ester of cabotegravir (CAB, named XVIR-110), a potent HIV integrase strand transfer inhibitor. We are pleased to affirm its clinical candidate selection status for HIV-1 prevention. XVIR-110 has a projected dosing interval of beyond every six months. The formulation has successfully completed IND-enabling GLP toxicity studies for a planned Phase I clinical trial and found to be safe. With this success as a foundation we now present our next research goal. It is to develop a next generation ULA ART that favorably addresses the needs for combinatorial ULA regimens, shorten the PK tail in existing LA formulations and XVIR-110, and further decrease injection volumes. In support of each goal is extensive preliminary data offered through the selection of homodimeric bictegravir (BIC) ester prodrugs demonstrating superiority over dolutegravir (DTG) and CAB. We posit that process chemistry design and development for the lead BIC dimer prodrug and prodrug design for rilpivirine (RPV) and tenofovir (TFV) will produce a complete six month dual therapy (BIC/RPV and BIC/TFV) for treatment of HIV-1 infection. Drug choices, prodrug and formulation development, in vitro and in vivo screening and mechanisms, toxicology, PK, and pharmacodynamic profiles will follow comprehensive Go-No Go criteria that advanced XVIR-110. To accomplish our research goals, a partnership was made between a medicinal and polymer chemist (B Edagwa) and a virologist, cell biologist, and immunologist (H Gendelman). Our complementary team has a strong history of collaboration and will continue to leverage on technical and scientific support from Exavir Therapeutics Inc. and Gilead Sciences. We posit that harnessing of natural viral target cells (monocyte-macrophages and CD4+ T cells) as drug carriers and depots will improve treatment outcomes. We will use a milestone-driven approach to identify and advance the most promising six month complete two drug regimen towards clinical translation.

NIH Research Projects · FY 2025 · 2019-06

PROJECT SUMMARY This Pathway to Independence Award will equip the candidate with the key skills to study interpersonal connectedness by examining behavioral and physiological linkage (e.g., how closely people’s heart rates and emotional behaviors are synchronized) in individuals with frontotemporal dementia (FTD) and their spousal caregivers (CGs). FTD is associated with profound neurodegeneration in the frontal and temporal regions of the brain and severe socioemotional symptoms (e.g., apathy, disinhibition) that are difficult not only for the person with dementia (PWD) but also for the CG. Caring for a loved one with FTD can be a meaningful part of family life; however, disruptive symptoms can produce deleterious effects and undermine CGs’ health and well-being. Given the strong associations established between close interpersonal connectedness and health/well-being, the candidate plans to take novel steps to examine (a) how interpersonal connectedness of the PWD and CG is altered in FTD (Aims 1 and 3), and (b) how altered PWD-CG connectedness provides a bridge that links PWD’s socioemotional symptoms to CG’s health declines (Aim 2). Importantly, the candidate plans to study PWD-CG connectedness by assessing both behavioral and physiological indicators of PWD-CG connectedness. This novel approach will provide more objective, less cognitive-demanding, and more continuous measures for PWD-CG connectedness, as compared to single self-report measures that have typically been used in past research. The candidate also plans to develop the optimal methodological approach for quantifying PWD-CG connectedness in both laboratory and naturalistic settings (e.g., identify the measure of physiological linkage that best predicts poor CG health; Aim 4). The candidate has received solid training in cognitive neuroscience and has a strong track record using integrative methods to study emotion and social functioning in healthy and clinical populations. To support the candidate in conducting the proposed research, training in 3 areas is planned: (a) dyadic behavioral and physiological time series data analysis, (b) diagnosis and clinical assessment of FTD, and (c) dementia caregiving. Training will occur under the mentorship of renowned experts in each field (including a clinical psychologist, a behavioral neurologist, and a nurse specialist). The training environment at the Department of Psychology, University of California, Berkeley (primary), and the Memory and Aging Center, University of California, San Francisco (secondary), will be ideal for the proposed training and research, as well as for developing the candidate’s professional skills. Together, the award will help the candidate launch his research career as an independent scientist with unique expertise in dyadic emotion and psychophysiology, neurodegenerative disease, and caregiving. The proposed research will help advance the understanding of the pathways through which FTD yields collateral damage on CGs, and provide a novel, objective, continuous, and potentially portable measure to quantify PWD-CG connectedness for future research and clinical use.

NIH Research Projects · FY 2025 · 2018-07

ABSTRACT Despite advances in basic and translational neuroscience research, effective therapeutics for neurodegenerative, neuroinflammatory, psychiatric, developmental, and neuroinfectious disorders remain in want. The National Institutes of Health has recognized bench to bedside research to improve disease outcomes and as such initiated programs to train researchers who can conceptualize new disease approaches to aide in effective treatments for neurological disease. One critical “in need” area is neuroimmunity which remains understudied despite its close linkage to the pathobiology of a broad spectrum of neurological disorders. The tradition of training separate groups of students in the disciplines of neuroscience and immunology hinders field growth. Following notable training successes, we seek continuance in training of predoctoral students in studies of neuroimmunology with linkages to neuronal injury, differentiation, regeneration, and protection. The program is designed to train 4 predoctoral students at a time in 1 or 2 year appointments with broad exposures to research methods facilitating best approach, technical, and outcome proficiency. And that each student acquires a broad interdisciplinary field knowledge. This allows for critical thinking in how inflammation affects disease pathogenesis and treatment. Several approaches are proposed to achieve these goals. First, is providing students training opportunities for multidisciplinary studies that intersect immunity and neural function. Second, is the use of our textbook Neuroimmune Pharmacology designed specifically as a guide in the intersections between neuroscience, immunology, and pharmacology. This is taught in our Neuroimmunology course. Third, we will provide a unique conceptual framework to integrate approaches relevant to neuroimmunology research. This contains, but is not limited to, systems biology, cell signaling, glial and neuronal biology, relevant rodent and laboratory models, and synaptic-network physiology. Fourth, are our bi-monthly workshops with student presentations to the program’s neuroscience and immunology faculty and a statistician to acquire feedback in research design, uses and limits of quantitative approaches, statistical interpretations, and conceptualization of ongoing research activities. These are highly interactive, diving into the mechanics of research and complement the formal student research presentations at the program’s annual retreat. Fifth, are clinical neurological experiences to gain the big-picture of a real-world perspective of neurological disease. Sixth, are “cross- disciplinary internships” where students will complete thesis component(s) in another laboratory using a different research approach and mentor. Seventh, are sustained community, university, and logistical support. By coordinating the training efforts of divergent research groups linked by common interests in neuroimmunity, trainees will develop deeper understandings of innate and adaptive immunity in relationship to neurologic disease. Such trainees will be better prepared to develop successful careers in studies of disease pathobiology and therapeutic interventions for human nervous system disorders.

NIH Research Projects · FY 2025 · 2018-03

Project Summary: Overall The goal of the Nebraska Center for Molecular Target Discovery and Development (CMTDD) is to establish and expand physical and intellectual resources at the University of Nebraska Medical Center (UNMC) and the University of Nebraska system to catalyze the identification, validation, and development of approaches for manipulation of molecular targets implicated in clinically important diseases. We perceive an increasing expectation by journals and granting agencies for progressively sophisticated model systems and data sets. Acquisition of these systems and generation of high-content data often requires a substantial investment in time and money that taxes limited research resources. To enhance the competitiveness of CMTDD Research Project Leaders (RPLs) and other university faculty members during Phase 2, the CMTDD recently invested in the development of technologies to enhance the sophistication of investigator research while minimizing the cost of access. These technologies: single-cell and spatial transcriptomic analysis, organoid generation and manipulation, and inhibitor and proteolysis-targeting chimera (PROTAC) synthesis will be integrated into three Phase 2 CMTDD cores. These investments will enhance the research capabilities of the institution, expand the translational capacity of its faculty, facilitate the training, mentoring, and career advancement of promising new faculty, and efficiently drive discovery and development for the improvement of human health in Nebraska and the nation. Phase 2 RPLs bring a breadth of cutting-edge expertise to the CMTDD. With mentoring and ready access to CMTDD-supported cores, our RPLs will become self-supporting faculty members invaluable to the future success of UNMC. Our Phase 2 RPLs share both a passion for exploration of the basic biologic principles that underly cell homeostasis and a belief that many diseases can be effectively classified and characterized through detailed genomic, genetic, and molecular analyses that identify drivers and vulnerabilities from which will emerge unique therapeutic approaches. To realize this vision, the CMTDD will: 1) maintain an Administrative Core and mentoring program that successfully graduated all but one of its initial RPLs in the first four years of support; 2) increase research capacity through newly established state-of-the-art scientific cores for single-cell and spatial transcriptomics, organoid development, and inhibitor and PROTAC synthesis and validation; and 3) provide low- or no-cost access to these technologies and reagents as an affordable means for increasing the impact of investigator research.

NIH Research Projects · FY 2025 · 2017-09

PROJECT SUMMARY The University of Nebraska Medical Center (UNMC) College of Medicine and the University of Nebraska Omaha's Biology Department and its programs in Native American Studies and in Medical Humanities, will leverage the trust and cooperative spirit that has been garnered working with tribal schools and communities in Nebraska, South Dakota and Arizona to develop, implement, and evaluate cancer-related science curricula and research experiences aimed to support Native American students. The UNMC in community-based cancer prevention combined with the Munroe- excellence in educational outreach and the first-rate basic and translational cancer research in state-of-the-art laboratories at the Eppley Institute in the NCI-designated Fred & Pamela Buffett Cancer Center are consolidated in the UNMC Youth Enjoy Science program (UNMC-YES) for the benefit of Native American students' education in all aspects of cancer research. The long-term goals of this project are to promote student interest in the sciences, foster a more science-literate public, and increase the number of Native Americans entering health and science careers with a particular emphasis in cancer research. UNMC-YES aims to: 1) Cultivate and sustain trusting community partnerships to expand outreach initiated in the original grant cycle to rural and urban Native American students, 2) Develop, disseminate and implement culturally relevant, technology-based cancer education curricula with Native American populations, and 3) Expand Native American student horizons through hands-on, culture- and standards-based science activities, research experiences and exposure to health science careers. These aims will be accomplished through innovative and evidence-based 1) outreach and engagement, 2) curriculum development and dissemination, and 3) provision of research experiences. UNMC-YES will generate and deliver cancer-related activities, lessons and research experiences to Native American students in grades 6 through undergraduate. UNMC-YES will also provide teachers with research experiences, workshops, mentoring, and in-service education to facilitate implementation of project curriculum with Native American students. Student participants will experience a range of highly intensive activities such as summer research camps to higher frequency activities such as school-based science clubs, mentored cancer research internships, and ongoing research talking circles. Professional evaluations will be made at all stages, with major emphasis on the educational, professional and community impact of the project. Pre- and post-test measures will analyze participant outcomes. Stakeholder perspectives and community systems for reducing cancer will be assessed to inform continuous improvement. Our objective is not only to increase Native American representation in cancer-related health professions and research but also to advance the health of Native Peoples through improvements in cancer prevention, treatment, and research.

NIH Research Projects · FY 2026 · 2017-07

Project Summary Heart failure (HF) is a major public health problem worldwide, especially myocardial infarction-induced HF with reduced ejection fraction (HFrEF) accounting for 50% of all HF cases. Malignant ventricular arrhythmia accounts for nearly 50-60% of mortality in HF patients. Cardiac sympathetic overactivation, a major feature of HF, could trigger malignant ventricular arrhythmias and sudden cardiac death. My research laboratory is continually focusing on the regulatory role of the peripheral nervous system in peripheral tissues (such as myocardium) in pathophysiological conditions including HF. Our recent research project found that cardiac sympathetic neuronal dysfunction contributes to cardiac sympathetic overactivation and malignant ventricular arrhythmias in advanced HF. To extend our current work, the overall vision of this R01 application is to test whether satellite glia-modulated macrophages can drive cardiac sympathetic functional and structural remodeling and be the therapeutic target for improving cardiac sympathetic function and reducing malignant ventricular arrhythmogenesis in advanced HF. To accomplish this vision, the proposal will still use myocardial infarction-induced advanced HF and sham (sham surgery) animals as the primary experimental tool to pioneer discovery in 3 Specific Aims. We will also use multifaceted approaches in this project, from conscious and anesthetized animals to cellular-molecular-genetic levels, along with some especially cutting-edge techniques (e.g., optogenetic satellite glial silencing, 3D reconstruction with tissue-maker software, cardiac slice electrochemistry recording, and a hyaluronic acid-based hydrogel delivery system). Specific Aim 1 will determine activation of satellite glia and macrophages as well as relations between satellite glia and macrophages in cardiac sympathetic ganglia in advanced HF, because there is limited information about relations among satellite glia, macrophages, and sympathetic ganglionic neurons in advanced HF. Specific Aim 2 will determine the involvement of satellite glia and macrophages in cardiac sympathetic remodeling including structural and functional alterations in cardiac sympathetic neurons located in stellate ganglia and their nerve terminals in advanced HF, because most published studies (including our work) reported the scattered information about HF-triggered cardiac sympathetic remodeling. Specific Aim 3 will provide a tight link with other Specific Aims to establish optogenetic satellite glial silencing and hydrogel-encapsulated (also macrophage-related) therapeutic approaches against sympathetic overactivation, ventricular arrhythmogenesis, and cardiac contractile dysfunction in advanced HF. Accomplishing these goals will provide major conceptual, technical, and translational advances for our understanding of the pathophysiology and related therapeutic targets of cardiac sympathetic overactivation and malignant ventricular arrhythmogenesis in advanced HF.

NIH Research Projects · FY 2024 · 2017-05

ABSRTACT Despite major efforts to keep operating rooms sterile, surgical site infections (SSIs) remain a serious and stub- born problem, killing up to 8,200 patients a year in the U.S. Our long-term goal is to develop novel therapies that effectively minimize risk of SSIs and promote wound healing. During the previous funding cycles of this award, we have demonstrated that i) successful encapsulation and sustained release of vitamin D and other immuno- modulating compounds induced higher cathelicidin antimicrobial peptide (CAMP) gene and protein (hCAP18/LL- 37) levels in immune cells and keratinocytes in cell culture, a human CAMP transgenic mouse wound model and human skin explants; ii) topical vitamin D increased killing of Staphylococcus aureus in a skin wound-infection model using our human CAMP transgenic mouse; and iii) exosomes secreted from immune cells treated with vitamin D contained higher levels of hCAP18/LL-37. Furthermore, we developed a novel gas-foaming expansion technique to fabricate improved 3D nanofiber scaffolds eluting vitamin D that promoted immune cell infiltration, induced hCAP18/LL-37, decreased inflammation, and promoted neovascularization and collagen deposition in human immune system-engrafted mice. Building on these findings, and our preliminary data, the goal of our proposal focuses on the development of nanofiber-based dressings for enhancing innate immunity. Our over- arching hypothesis is that co-encapsulating immunomodulating compounds with exosomes secreted from treated immune cells in 3D nanofiber scaffolds will synergistically enhance protection against SSIs and promote wound healing better than either component alone. To accomplish this, there are three specific aims: 1) Demon- strate efficient encapsulation and elution of immunomodulating compounds from our improved 3D nanofiber scaffolds; 2) Determine the antimicrobial and wound-healing efficacy of exosomes derived from primed or human CAMP transfected immune cells; and 3) Demonstrate the efficacy of immunomodulating compounds and exo- somes - co-incorporated 3D nanofiber scaffolds to promote healing and prevent infection in our humanized trans- genic mouse model and ex vivo human skin explants. Building on work from our prior grant, we expect successful completion of the aims in this renewal will lay a strong foundation for developing the next generation of novel therapeutic anti-infective wound dressings that could greatly speed healing, reduce rates of SSIs and minimize development of antibiotic resistance. We also expect these dressings could serve as effective treatments for traumatic and combat-related injuries.

NIH Research Projects · FY 2025 · 2017-05

Abstract This proposal to continue in the Pancreatic Cancer Detection Consortium will further build and enhance our biospecimen resource that collects longitudinal blood samples on patients at risk of developing pancreatic cancer and unique tissue resources that include rare pancreatic premalignant lesions. We propose to continue and enhance our discovery and validation of blood-based and imaging-based biomarkers that have the potential to detect and differentiate the earliest possible stages of pancreatic lesions that are likely to progress to cancer. This proposal proceeds from an ongoing effort that was initiated 3.5 years ago in which we have enrolled approximately 469 patients with known germline mutations or with significant inherited risk of unknown etiology (more than 2 affected with no defined mutations) for pancreatic cancer. We prospectively collect biological specimens (100 mL blood) and detailed clinical data every six months from an at-risk cohort that represents the target population envisioned for clinical application of biomarkers with potential to detect pancreatic cancer. In addition to patients with inherited risk, we will continue to collect longitudinal samples on groups of patients with increased risk of pancreatic cancer: those with pancreatic cystic neoplasms or chronic pancreatitis, and new onset diabetes (NOD). We also propose three biomarker research specific aims that have grown out of progress during the previous grant award. One aim will build upon our previous results with mucin type biomarkers and related glycoproteins by examining the potential of a set of additional biomarkers to improve detection cancer prior to clinical diagnosis in the earliest stages of disease progression. A second aim will examine the capacity of newly identified antibodies to CEACAM6 to identify earliest lesions in the pancreas by immunohistochemistry and state of the art imaging modalities. A third aim will determine the performance characteristics of a panel of exosome-based biomarkers detecting early pancreatic cancer and differentiating benign cystic lesions from those that progress to cancer.

NIH Research Projects · FY 2025 · 2016-09

Although clinical trials are the highest standard of evidence for the effectiveness and safety of medical interventions, children often receive medical therapies that have never been studied in clinical trials involving pediatric subjects. Knowledge of the etiology, prevention, and/or treatment of these illnesses often must be extrapolated from adult-derived data. Children often respond to medications differently from adults due to differences in physiology and drug metabolism, placing them at increased risk of experiencing adverse, even life-threatening events from off-label medical treatment use. Clinical trials furnish patients opportunities to receive well-supervised treatments and procedures before they are widely available, gain improved understanding of their health conditions, and contribute to knowledge that benefits others. Importantly for IDeA states like Nebraska, children from communities with lower access to advanced medicine (e.g., rural regions) are less likely to have access to clinical trials. During the first two cycles of the ECHO IDeA States Pediatric Clinical Trials Network (ISPCTN), the Nebraska Pediatric Clinical Trials Unit (NPCTU) has led the first network-developed randomized controlled trial, enrolled participants in almost all ISPCTN studies, and expanded human and organizational capital in Nebraska to conduct clinical trials statewide. The NPCTU is poised to build upon this success to ensure that all Nebraska children have access to clinical trials. We propose the following Specific Aims to drive continued growth in clinical trials access and participation for Nebraska children: (1) Develop, conduct, and disseminate findings from multicenter clinical trials research, assuring the participation of children living in IDeA states. The NPCTU will solicit trial proposals from community stakeholders and faculty and propose at least three multicenter clinical trial concepts to the ISPCTN, participate in implementing all five anticipated multicenter network trials, including our proposed trial to test the effectiveness of a multi-level mHealth-based intervention to increase adherence to recommended infant immunizations and lab screenings. (2) Build pediatric clinical trial research capacity at the clinical site. The NPCTU will continue to expand clinical trials performance capabilities by enacting a multifaceted training and mentoring plan for developing early-stage faculty into proficient leaders of multicenter clinical trials and train clinical and research staff to conduct trials. (3) Engage interested parties such as community members, nonprofit organizations, and professional societies to enhance ECHO ISPCTN clinical trial impact, transferability, rigor, and feasibility. The NPCTU will sustain and build on our robust engagements with professional societies that issue pediatric clinical practice guidelines and with a wide range of individual- and organization-level community stakeholders. Existing and new collaborations will provide rich insight into communities’ research priorities, help us deploy trials in ways that more fully engage Nebraska children, and disseminate results back to communities.

NIH Research Projects · FY 2025 · 2016-09

Project Summary The highly innovative Rapid Autopsy Program (RAP) addresses a major limitation in pancreatic and prostate cancer research: the lack of quality human specimens of cancer representing all stages of disease. This program, led by Dr. Paul M. Grandgenett, which includes rapid collection of large quantities of all organs and fluids within 5 hours of death, is a unique program in the world and is designed to maintain cellular and molecular integrity of the tissue. The RAP is a core component of the University of Nebraska Medical Center (UNMC) Pancreatic Cancer Research program and the GU Oncology Focus Group, that is tasked with the collection, archiving and distribution of tissues and fluids from patients that have recently passed with pancreas or prostate cancer. The Unit Director of the Pancreatic Cancer Research program at the Fred and Pamela Buffett Cancer Center is Dr. Michael A. Hollingsworth, while the prostate group is led by Dr. Surinder Batra and Dr. Shawna Boyle. Dr. Grandgenett, a Research Associate Professor under Dr. Hollingsworth's aegis, is the director of the RAP and the applicant for this Research Specialist Award. The rapid autopsy program operates within the Pancreatic Cancer Research Program at UNMC and is important to and currently funded in part by: a pancreatic cancer SPORE p50 grant; ongoing and pending P01 and U01 grants and several R01 grants to individual investigators. Dr. Grandgenett personally directs the collection of primary tumor, residual unaffected tissues, distant metastases, all unaffected tissues, as well as fluids such as blood and ascites for all PDAC and Prostate cancer patients. He has also established an extensive normal tissue acquisition program that is highly utilized. Dr. Grandgenett determines in advance how specific samples will be prioritized based on the needs of funded projects both at UNMC and at other institutions. Currently, Dr. Grandgenett provides fresh samples to multiple laboratories, at UNMC and outside-UNMC, conducting research on blood-based biomarkers (proteins, RNA, DNA), circulating tumor cells, patient-derived modeling including organoids and xenografts, and other processes. The remaining samples are snap frozen, maintained in long-term storage and made available through request and collaboration for local, national and international cancer studies. Key roles of Dr. Grandgenett is to coordinate the training and participation of the thirty RAP volunteers and to work with collaborators to determine the samples necessary for their research project, to modify our SOP as needed to obtain those samples, confirm specimen quality and then to distribute those samples to investigators. It is essential for the leadership of this resource to be a highly trained Research Specialist with extensive experience through participation in the autopsies, a comprehensive knowledge of patient sample availability, as well as the ability to interact on a scientific level with each investigator. Dr. Grandgenett has a major role in all aspects of the collection, processing, annotation and distribution of these biospecimens and plays a vital role in expanding collaborative projects with investigators at UNMC and on a national and international level.

NIH Research Projects · FY 2025 · 2016-09

Biomedical Informatics, Bioinformatics, and Cyberinfrastructure Enhancement Project Summary: The Biomedical Informatics, Bioinformatics, and Cyberinfrastructure Enhancement (BIBCE) Core offers a robust and agile biomedical informatics infrastructure for the Great Plains (GP) IDeA-CTR members with cutting-edge support tools, expertise and training across bio-, clinical-, and public health informatics realms. Extensive reliance on genetic and molecular data in clinical practice has become a game-changer in health care practice, especially in the emerging field of precision medicine. Hence, clinical and translational research (CTR) needs a range of support, from data collection and analysis to data integration and correlation, to make clinically relevant inferences effectively. However, it is extremely challenging for individual institutions to locally develop and maintain sophisticated infrastructure and technical expertise in this rapidly changing field. The primary objective of the BIBCE Core is to develop expertise, tools, and services and make them accessible across all institutions of the GP IDeA-CTR network. Collecting and connecting data from the molecular level to population level, the BIBCE Core will provide access to a robust suite of tools for regional CTR endeavors while building the overall CTR capacity of the GP IDeA-CTR network. While Nebraska has an extensive telehealth network that links all public educational facilities and hospitals, informatics tools and expert support are essential to generate multi-institutional collaborations in CTR, and having a centrally supported core will be pivotal to accomplish the mission of GP IDeA-CTR. The BIBCE Core will support this mission with four specific aims: 1) Augment CTR capacity by leveraging existing infrastructure, networking, and creative collaborations among GP IDeA-CTR, institutional, and other IDeA-funded projects; 2) Integrate and streamline access to genomics-based and health data, biobanks & registries, electronic health records, and decision support systems across the network to promote translational precision health research; 3) Connect the clinical informatics resources with CTSA’s National Center for Data to Health (CD2H) and other CTR centers to facilitate easy access and stimulate feasibility studies, outcomes research, and CTR; and 4) Support data sharing, knowledge and resource sharing, and training using cyberinfrastructure enhancement to reach out to the populations in the GP IDeA-CTR network. The BIBCE Core will have a profound impact on the success of GP IDeA-CTR through augmenting the overall research capacity in the realms of clinical, translational, and precision health; bolstering collaborations through team science within as well as across the IDeA networks; streamlining informatics resources; aligning the regional CTR with national efforts for interoperability; and sharing knowledge with new courses and training activities.

NIH Research Projects · FY 2026 · 2012-03

In the US, 15 million people have alcohol use disorders (AUD), and 95,000 people die per year of alcohol attributable causes, making alcohol misuse the third-leading preventable cause of death in the United States. Despite the overwhelming need for better prevention and treatment of the health consequences of AUD, the biomedical science workforce devoted to this problem is inadequate. The University of Nebraska Medical Center has a history of performing quality alcohol-related research since the 1960s. This history, existing infrastructure, and organization of multiple enthusiastic, federally funded alcohol researchers make us well-positioned to continue to provide superior training and inspire the next generation of alcohol researchers. Our program’s goal is to provide undergraduate students with early, immersive exposures to alcohol research, with the hope of launching some of these students on their way to careers in biomedical and alcohol-focused research. We have a strong track record of supporting the development of young scientists with our Summer Undergraduate Alcohol Research Program (SUARP). The SUARP was initiated in 2012 with funding through the NIAAA and has trained 77 undergraduate students. Our program is already making progress in achieving its major long-term objective of adding to the biomedical research workforce. Of the 77 students participating in the 10-week program since 2012, we have follow-up data on 71 (92%). Of those, 72% reported participating in a research project after completing our program, 7% are in graduate school in a science field, and 58% are in health professions school or are practicing clinicians. The SUARP program will provide a 10-week, hands-on, mentored research experience for undergraduate students to stimulate their interest in careers in biomedical research. As a part of the program, we will teach SUARP students the pathophysiological consequences of alcohol misuse on human health. We will also introduce students to the scientific skills needed to build a successful scientific career. This is a comprehensive, intense program of scientific training dedicated to developing the next generation of scientists engaged in examining the challenging issue of alcohol intake and its impact on health. We have built a strong program and remain enthusiastic to continue to serve students and contribute promising researchers to the alcohol research community.

NIH Research Projects · FY 2025 · 2011-08

ABSTRACT This proposal is a competitive renewal of a multiple-PI R01 funded through NIAAA. The goal of the application is to examine how ethanol (EtOH) exposure contributes to fat accumulation in the liver due to altered dynamic properties of the lipid droplet (LD), a fat storage organelle. Almost all heavy drinkers develop fatty liver, which is marked by the aberrant and significant accumulation of intrahepatocellular triglycerides stored within LDs. Understanding the cellular processes contributing to this fat accumulation will provide essential information for preventing further injury progression, as it is known that alcoholic fatty liver is the initial but reversible stage of liver injury. Here, we propose to provide molecular insights into these critical questions, aimed at potential treatments that reverse or prevent EtOH-induced steatosis. This proposal combines the expertise of two senior investigators recognized for their contributions to the study of EtOH-induced cell injury and hepatic LD biology. We believe this collaborative effort has been beneficial to the field of ALD and will result in outcomes not attainable by individual efforts alone. This proposal comprises two well-integrated aims. Aim One: Blockage of ER-Associated Lipophagy by EtOH. We have made the interesting observation that nascent LDs of a conserved size (170nm) accumulate at the ER surface of EtOH-damaged hepatocytes and that lysosomal/autophagic compartments subsequently catabolize these ER-associated LDs. Notably, both chronic EtOH exposure and pharmacologic disruption of lysosome function attenuate this process leading to hepatocyte steatosis. Aim Two: Disruptive Alterations of the LD Proteome by EtOH. We found that many components of the LD “surface proteome” are post-translationally modified by ubiquitin, a pathway we posit directs removal of select proteins to the proteasome for degradation or directs the entire LD to the lysosome for catabolism. Importantly, we have found that chronic EtOH exposure markedly increases the ubiquitination of the LD proteome and thus disrupts normal LD degradation/catabolism leading to steatosis. These observations and specific aims will allow us to pursue the CENTRAL HYPOTHESIS of this study that EtOH promotes accumulation of both mature and nascent, ER-associated, LDs through alterations of the LD proteome inhibiting lipophagy and lipid catabolism leading to hepatocyte steatosis. The proposed investigation will utilize state-of- the-art imaging and proteomic technologies to quantify specific molecular events that contribute to alcohol- induced fatty liver. Successful completion of these studies will provide novel insights as to how EtOH affects LD dynamics in liver cells and important information for therapeutic strategies aimed at reducing or eliminating the severity of steatosis and blocking its further progression to alcoholic steatohepatitis, fibrosis, and cirrhosis.