Mayo Clinic Rochester

NIH Research Projects · FY 2026 · 2023-01

PROJECT SUMMARY/ABSTRACT Chemotherapy-induced peripheral neuropathy (CIPN) is a serious side effect that causes morbidity and limits the dose of chemotherapy allowed to treat cancers. In most classes of neurotoxic chemotherapeutics, CIPN manifests as damage to dorsal root ganglia sensory neurons. The neuronal damage is thought to be due to an unelucidated combination of dysfunctional microtubules, axonal transport and mitochondria that ultimately leads to programmed axonal degeneration via a nicotinamide mononucleotide adenylyl transferase 2 (NMNAT2) and Sterile Alpha and TIR Motif Containing 1 (SARM1) pathway. The link between these processes, however, remains entirely unknown. Using a human induced pluripotent stem cell (iPSC)-derived dorsal root ganglia sensory neuronal culture system (iSN), we have identified microtubule-associated protein 2 (MAP2) as a key determinant of CIPN. When exposed to clinically-relevant doses of bortezomib, paclitaxel or vincristine, MAP2 levels drop and there is subcellular MAP2 mislocalization in iSN that occurs prior to axonal degeneration. Critically, overexpression of MAP2 is protective for bortezomib-induced neurotoxicity in iSN. In this grant application, we propose to further dissect the role of MAP2 in CIPN by utilizing an innovative CRISPR-ErCas12a system to develop genetically-engineered iPSC lines that fluorescently-tag either MAP2 or NMNAT2. These novel iPSC lines will allow for detailed analyses of the time course of altered subcellular localization and axonal transport, and how these processes link to SARM1 activation and axonal degeneration. Furthermore, we will investigate the overlapping mechanisms in CIPN due to bortezomib, paclitaxel, and vincristine using our previously successful proteomics analysis approach, which now will employ a state-of-the-art quantitative proteomic and phosphoproteomic technology. The proposed Specific Aims build on existing understanding of the pathomechanisms of CIPN while innovating into unexplored novel areas that are potential targets for preventative therapeutic development.

NIH Research Projects · FY 2026 · 2022-12

Project Summary/Abstract Heart transplantation is recognized as the gold standard therapy for end-stage heart failure. However, demand for donor hearts currently far outstrips supply due to multiple factors. An important limitation is primary graft dysfunction (PDG) in 10-20% of transplants and is an important contributor to adverse clinical outcomes and increased resource utilization. PGD occurs when donor heart function is inadequate for end organ perfusion and the risk increases once cold preservation time is >4 hours. Mineralocorticoid receptor (MR) signaling has a key role in many cardiovascular diseases including heart failure, and cardiac hypertrophy. It mediates harmful processes such as oxidative stress, inflammation, and fibrosis. Using murine models with MR deletions in cardiomyocytes as well as clinically utilized mineralocorticoid receptor antagonists (e.g. canrenone), we show that MR antagonism can greatly improve donor heart function following preservation. We also demonstrate that cardiac preservation is associated with increased MR protein expression coupled with organization of MR into molecular condensates with organizational structures that are known to augment protein transcription and translation. Importantly we find this occurs both in murine models as well as human hearts suggesting conserved events during evolution. We also confirm that MR contains intrinsic disordered regions which mediate phase separation and condensate formation. Our plan is to: (1) Identify the MR domains that can mediate condensate formation at differing temperatures. We will also determine if MR ligands, MR response elements and histone deacetylases (HDAC) can facilitate MR condensate formation. This will be performed by in-vitro evaluation of condensate formation by cloning truncated MR constructs missing specific domains and then adding different mediators of interest (e.g. HDAC) in the presence of a crowding agent. (2) We will examine the effects of MR condensate formation by first deleting MR in cardiomyocytes and then use adeno- associated virus to reconstitute cardiomyocytes with a MR mutant that has an impaired ability to form condensates because it is missing the intrinsic disordered region. We will then compare the inflammatory, oxidative stress and cell death responses associated with full length MR and the MR mutant. This will be evaluated in-vivo in a transplant model which incorporates recipient responses, ex-vivo in a perfusion model consisting only of the native cardiac cells and in an in-vitro neonatal cardiomyocyte preservation-reperfusion culture model. (3) We will determine the efficacy of canrenone (a MR antagonist) with or without valproic acid (a HDAC inhibitor) for improving donor heart function in pigs and humans. Single-cell RNA sequencing will be performed in human hearts to determine the cell population in which MR signaling is important for preservation. Our findings will contribute towards decreasing PGD occurrence, increasing donor heart utilization, and improving transplant outcomes. This is expected to have broad clinical implications such as transplantation of other solid organs (e.g. liver, kidneys) and for warm cardiac ischemia settings (e.g. myocardial infarction).

NIH Research Projects · FY 2026 · 2022-12

ABSTRACT / SUMMARY Background: Breast cancer, the most common cancer in women, involves the axillary lymph nodes in 25-30% of cases. Treatment usually involves systemic therapy before surgery. Complete response to neoadjuvant systemic therapy (NST) with normalization of the lymph nodes translates to less extensive surgery, lower related morbidity, and sometimes less intense radiation therapy. To mark a positive lymph node, a biopsy marker is placed in the positive node pre-NST. Post-NST, the marked node is identified with ultrasound, mammography, or computed tomography to implant a localizer, which the surgeon uses for node removal. Ultrasound is the first- line imaging modality for this task. Still, the marker cannot be found sonographically in ~25% of cases, resulting in suboptimal or aborted localizations, delays to surgery, longer procedural times, increased patient discomfort or inconvenience, and increased absorbed costs by the clinical practice. Errors in identifying the proper node(s) may lead to false-negative results, over/undertreatment, and potential cancer recurrence. Marker migration remains another concern that can lead to misguided localization, directing the surgeon to the incorrect node. This research project aims to address these unmet and critical needs to develop reliable, readily ultrasound-conspicuous markers that are also resistant to migration. Methods: Our preliminary data suggest that physical features of markers, like surface roughness, can cause an ultrasound twinkling artifact or “twinkling signature” classically associated with kidney stones using color Doppler flow imaging. With the collective expertise of a diverse research team, we have developed markers using polymethyl methacrylate and additive manufacturing (three-dimensional printing). These markers generate robust twinkling signatures with ultrasound, are visible with multiple imaging modalities, and are resistant to marker migration. We will test the efficacy and safety of these markers in a porcine model over a six-month period to mimic the duration of NST treatment with chemotherapy. Lastly, we will conduct a Phase 1 clinical trial in breast cancer patients with our optimized marker to evaluate the long-term strength of the twinkling signature and the degree of marker migration. To accomplish the goals of this project, we propose these Specific Aims: Aim 1 Optimize ultrasound acquisition characteristics and the signal processing chain to robustly detect twinkling. Aim 2 Optimize physical characteristics of breast procedure markers that enhance Doppler-based twinkling and reduce marker migration. Aim 3 Assess long-term safety, twinkling persistence, and marker migration of candidate markers in a porcine animal model. Aim 4 Evaluate twinkling and migration of optimized markers in a Phase 1 clinical trial using markers implanted in a positive axillary lymph node prior to NST in patients with breast cancer.

NIH Research Projects · FY 2026 · 2022-12

PROJECT SUMMARY Efficacy of CAR T cell therapies against solid tumors has been limited by a lack of expansion in vivo, low levels of tumor trafficking, poor functionality/persistence and by suppression in the tumor micro-environment (TME). Conventionally prepared CAR T cells are unable to retain sufficient differentiational plasticity to undergo natural pathways of T cell proliferation/differentiation/persistence in vivo for sustained, effective cytotoxic anti-tumor activity. Oncolytic viruses (OV) selectively replicate in tumor cells to generate a highly inflammatory TME which may enhance CAR T cell recruitment to, and function within, solid tumors. Our goal is to develop a novel regimen by which oncolytic virotherapy can be used as a potent immunological adjuvant to improve the efficacy of CAR T cell therapy against solid tumors. By loading the OV on the CAR T cells ex vivo, we achieved highly significant improvements in tumor therapy compared to virus or CAR T cells alone. Mice treated with completely systemically delivered CAR T and VSV developed a population of CD8+ CAR T cells with T Cell Receptor (TCR) specificity for the immunodominant H2Kb VSV N52-59 epitope of VSV which selectively expanded in vivo to very high frequency. This population of dual-specific (DS) (virus-specific and CAR T antigen-specific) memory-like (TM) CAR T cells 1). persisted in mice for much longer than conventional CAR T, 2). was significantly more functional against tumor, and 3). could be rapidly re-activated in vivo against tumor by a secondary systemic boost with homologous virus, resulting in long-term tumor cures. These therapeutic effects were observed with two different OV (VSV and reovirus) and across tumor sites (subcutaneous and intra-cranial). Therefore, here we propose a completely novel approach to expand the scope of CAR T cell therapy against solid tumors in which dual specific CAR T cells with improved activity against tumors are generated by using co-administered OV to induce a recapitulation of the physiological pathways that lead to optimal (CAR) T cell activation, proliferation and differentiation in vivo in response to virus infection. We have formulated three Specific Aims: 1) To define the molecular mechanisms by which TCR engagement of DS CAR T cells determines their phenotype, improved persistence and function; 2) To generate in vivo expanded populations of DS CAR T cells with TCR specificity against either tumor associated, or viral recall, antigens and to determine their therapeutic activity against established tumors; 3) To optimize in vivo expansion, persistence/longevity and re-activation of DS CAR T cells through novel boost and rest strategies targeting either the TCR, CAR or both. This will lead to implementation of fully systemic protocols for the combination of CAR T cells with OV which do not require any access to tumors.

NIH Research Projects · FY 2026 · 2022-12

PROJECT SUMMARY/ABSTRACT MLL-rearranged (MLL-r) leukemias account for 5-10% of human acute leukemia and is associated with poor prognosis. The unmet clinical needs and the lack of an effective targeted therapy to the MLL-r leukemias emphasize the need for novel regimens. Recent cancer epigenetics studies discovered a central role for the histone H3 lysine 79 (H3K79) methyltransferase DOT1L in MLL-r leukemogenesis. Important clinical responses have been noted with DOT1L inhibitor treatment as a single agent, however, it is expected that combination treatments will be necessary. Our preliminary studies based on a Tudor domain focused CRISPR screen in MLL-r leukemia identified SGF29 as a novel vulnerability in MLL-r leukemia. The objective of this application is to determine the critical epigenetic mechanisms that mediate the availability of KAT2A/B to maintain H3K9ac and oncogene expression in MLL-r leukemia. Our central hypothesis is that SGF29, an H3K4me3 reader protein, mediates recruitment of KAT2A/B to maintain histone H3K9ac and MYC oncogenic program in MLL-r leukemia. We will dissect the SGF29-mediated epigenetic mechanisms (Aim 1) and investigate the efficacy of SGF29 targeting (alone or in combination with DOT1L inhibition) as a novel therapy in MLL-r leukemia (Aim 2). This study is innovative because (1) it introduces a novel concept of simultaneously targeting multiple components of an epigenetic network to efficiently suppress the cancer programs, and (2) it establishes a brand new genetic screen approach for a sub-protein level functional pocket and drug discovery. The impact of this research will be of significance because (1) it immediately provides novel therapeutic opportunities against the difficult-to-treat MLL-r leukemias, and (2) it will help identify novel functional elements in epigenetic regulators for future pharmaceutical targeting.

NIH Research Projects · FY 2026 · 2022-12

Project Summary Interstitial lung diseases (ILDs) including idiopathic and other forms of pulmonary fibrosis represent a major and growing medical burden. While FDA-approved therapeutics limit progression of fibrotic ILDs, they do not fundamentally alter the course of these diseases. A central element of fibrosis progression in ILDs is the persistent activation of fibroblasts to a fibrogenic state; whereas transient fibroblast activation promotes wound healing, aberrant and prolonged fibroblast activation promotes fibrotic ECM deposition and hinders restoration of cellular homeostasis in epithelial and hematopoietic compartments. Hence, understanding how fibroblasts become activated and then locked in fibrogenic states is central to the development more effective therapies for fibrotic ILDs. Based on extensive preliminary data demonstrating a key role for the transcription factor Runx1 in fibroblast activation and fibrogenic memory in mouse and human fibrotic lung tissue, we will test the central hypothesis that fibroblasts gain and maintain a memory of fibrogenic activation that primes them for amplified activation upon repeated injury, and that Runx1 plays a central role in this activation and fibrogenic memory. We propose to test this hypothesis in three specific aims. In the first aim we will use a mouse model to identify the location, abundance and specific transcriptional targets of Runx1 engagement during fibrosis initiation, resolution and persistence. In the second aim we will test whether conditional deletion of Runx1 attenuates fibrosis and fibroblast memory, diminishes persistent fibrosis in a repeated bleomycin injury model, and restores homeostatic states in mesenchymal and other lung compartments. In the final aim we will analyze human lung tissue and fresh sorted fibroblasts to delineate Runx1 engagement, targets and functional effects relevant to human disease. In both mouse and human tissue, we will seek to identify the role of mechanical and biochemical signals in conferring fibrogenic memory and Runx1 activation and will test established and investigational therapeutics for their capacity to erase fibrogenic memory. Together these studies will test the function and regulation of Runx1 in fibrogenic cell activation and memory in mouse models and human tissue, potentially identifying a novel targetable mechanism underlying fibrotic ILD progression.

NIH Research Projects · FY 2026 · 2022-12

1 PROJECT SUMMARY 2 Alzheimer’s disease (AD) is the leading cause of dementia in the United States and its impact is only growing with 3 shifting demographics. The development of powerful biomarkers, measuring amyloid deposition, tau accumulation, 4 and neurodegeneration, has provided important insights into the pathophysiology of AD and AD-related dementias 5 (ADRD). Nonetheless, given the large variability across individuals, our understanding of the link between pathology 6 and cognitive dysfunction remains incomplete. Structural factors contribute significantly to this pathology-cognition 7 disconnect and are termed as “brain reserve.” There is a critical need for objective measures of reserve that will 8 improve the assessment of individual prognosis and guide therapy. 9 Brain biomechanics are an understudied structural feature of the brain, due in large part to the difficulty in measuring 10 relevant biomechanical states in vivo. Magnetic resonance elastography (MRE) is currently unmatched for 11 noninvasive measurement of brain mechanical properties. We have previously demonstrated that brain stiffness is 12 reduced due to AD, and our group and others have demonstrated in multiple studies that brain stiffness is a significant 13 reporter of cognitive function. However, previous studies face important limitations, namely technologies that were 14 optimized for reliability over resolution, and incomplete pathological assessment. Therefore, we will investigate two 15 aims with the overall goals to (1) advance MRE technology in order to (2) evaluate of the role of biomechanics in 16 brain reserve. 17 In Aim 1, we will optimize our machine learning-based MRE inversion framework by incorporating new a priori 18 information into the model that is specific to the brain. These advances to the model include the incorporation of 19 partial volume effects to reduce atrophy-related bias, and mechanical anisotropy to accurately model the coherent 20 structure of white matter tracts. Each advance will be tested in simulation and phantom experiments, and finally in 21 vivo for its ability to boost sensitivity to key AD/ADRD processes. 22 In Aim 2, we will use these tools to simultaneously map the mechanical signature of 4 pathophysiological processes 23 including amyloid, tau, white matter hyperintensities, and cardiometabolic conditions. Using first a discovery data set, 24 we will extract the mechanical feature that best reports cognitive performance, both globally and in specific domains. 25 These MRE-based features will then be evaluated in an independent test set for their ability to predict concurrent and 26 future cognitive performance. Finally, we will assess the unique value of mechanical biomarkers to predict cognitive 27 performance, using a parallel analysis but controlling for existing biomarkers derived from anatomical, functional, and 28 diffusion MRI. 29 In sum, the success of this proposal will shed new light on alterations to brain biomechanics with respect to 30 AD/ADRD processes, and their role as a buffer between pathology and cognition.

NIH Research Projects · FY 2025 · 2022-09

PROJECT SUMMARY Knowledge about lupus is biased towards the severe end of the disease spectrum due to difficulty assembling population-based cohorts of persons with lupus. An urgent need to expand the basic epidemiologic understanding of cutaneous and systemic lupus in four content areas (i.e., natural history, treatment, health care access, and disparities) was identified by the Centers for Disease Control and Prevention. Our project addresses these needs using the unique research infrastructure of the Rochester Epidemiology Project (REP), which contains longitudinal data on residents of a 27-county region in Minnesota and Wisconsin. We have assembled population-based cohorts comprised of persons with prevalent and incidence systemic lupus erythematosus (SLE) and cutaneous lupus erythematosus (CLE) matched to general population comparators without SLE/CLE by sex, age, race/ethnicity, and county of residence. These cohorts comprise the Lupus Midwest Network (LUMEN). Through the REP infrastructure, LUMEN follows individuals regardless of insurance status and encompasses the full spectrum of care, from primary care clinics to tertiary specialty care, and maintains an electronic index of diagnoses and procedures, including all inpatient and outpatient encounters, laboratory measures, and drug prescription data. In this competitive renewal of a highly successful grant investigating multimorbidity in SLE and CLE, opioid pain therapy, and healthcare access and gaps, our team of researchers and clinicians with decades of experience conducting population-based research will now focus on new research questions designed to have a high public health impact and to leap forward in our understanding of the natural history, treatment, healthcare access, and disparities across the full spectrum of CLE and SLE. Using the LUMEN population-based cohorts, we aim to 1) determine the contributions of lupus nephritis (LN) and anti-phospholipid syndrome (APS) to the excess mortality and unplanned healthcare utilization (i.e., emergency department visits and hospitalizations) among patients with SLE; 2) determine the contribution of LN to the excess risk of cardiovascular disease (myocardial infarction and stroke), infections, and osteoporosis among patients with SLE; 3) determine access to SLE-specific screening services and patterns of glucocorticoid use by social determinants of health, age, sex, and race/ethnicity; and 4) determine the transition rates over time and risk factors for transition from CLE to SLE over four decades to test the hypothesis that a) most transitions from CLE to SLE occur within five years of the CLE diagnosis, and b) oral contraceptives or postmenopausal hormone use increase the risk of transitioning from CLE to SLE. These projects will inform the individualized prognostication of SLE in clearly defined disease phenotypes and inform interventions to optimize patient care, prevent or reduce adverse outcomes in patients with lupus, and will identify a window of opportunity in CLE for therapeutic interventions to prevent the transition to SLE.

NIH Research Projects · FY 2025 · 2022-09

Project Summary / Abstract Modern biomedical science and medicine, with large varied and complex datasets, requires a paradigm shift in our educational models to train the next generation of scientists and physicians capable of performing and interpreting computational analyses. Current educational models have created a critical gap of expertise between computational data scientists and disease-oriented physiologists. The Specific Aim of this proposal is to establish an innovative educational model to promote the training of computational skills for immediate and direct application to research experiences and create the next generation of biomedical scientists to lead new discoveries in diabetes, digestive and kidney disease-oriented research. The Mayo Clinic Research Education Program in Computational Autonomic Neurobiology is a unique new program designed to ensure foundational understanding of both computational science and disease-oriented physiology, prior to learning practical skills in computational methods that are directly applied to data from mentored research experiences. Participants in the program, including graduate students, post-doctoral fellows, biomedical engineers, medical residents, and clinical fellows, are recruited widely from existing educational, training and research programs via the international reputations and productivity of the highly invested participating faculty. The program leverages institutional resources for program administration and robust program evaluation to remain adaptive and responsive to the needs of participants. The short-term objectives of the program are to: 1) provide foundational knowledge in physiology and computational science; 2) provide practical skill in the use creation and dissemination of computational workflows; and 3) provide a research environment to enrich skill honing and productivity. The long-term objectives of the program are to: 4) expand the use and access of computational analyses to impact diabetes, digestive and kidney disease-oriented research; and 5) attract and retain diverse scientists that impact the fields of diabetes, digestive and kidney disease- oriented research. By completing the program, we anticipate these scientists will form a bridge between the often distant physiological and computational approaches to understand human health and form a generation of diverse scientists that impact the fields of diabetes, digestive and kidney diseases.

NIH Research Projects · FY 2024 · 2022-09

PROJECT SUMMARY/ABSTRACT Shared decision making (SDM) is a process in which patients and clinicians work together to understand the patient’s problematic situation and collaboratively develop a plan of care that makes intellectual, practical, and emotional sense as a response to the situation and the individuality of the person. The practice of SDM is widely promoted through clinical guidelines, organizational and governmental policies and initiatives, and is a feature of patient-centered care. A goal of SDM is to improve health and behavioral outcomes through enhancing patients’ participation in decision making, however it has proven hard to establish this impact, in part due to limitations in measuring SDM. Researchers currently do not have the measures they need to determine if, in any patient-clinician encounter, a SDM process was used to collaboratively deliberate and form a plan of care. The ability to determine the occurrence of SDM is highly limited by a lack of conceptual and operational clarity around SDM, narrow measurement frameworks, and limitations to existing observer and self-reported measures, particularly in the context of chronic multimorbidity. Therefore, in this three-year study, the team—comprising experts in SDM, measure development and validation, and user-centered design—propose to develop a theoretically based, expert-informed, user-tested, observer-based measure of SDM occurrence (“SDMo”) and to estimate its reliability and validity. Specifically, the objectives of this application are to develop a comprehensive set of items using existing literature and theory to capture observable processes and behaviors indicative of SDM (Aim 1); using candidate items derived from Aim 1, to deploy a user-centered design process to develop a usable and feasible measure of SDM occurrence through an iterative process of field testing and refinement of the SDMo measure on a previously collected corpus of video-recorded clinical encounters (Aim 2); and to estimate the reliability and validity of the SDMo measure (Aim 3). Completion of these three aims will result in a usable measure of SDM occurrence in clinical encounters with patients with chronic multimorbidity, estimates of the measure’s reliability and validity, and a user-tested Users’ Manual that will include a detailed explanation of each item, illustrations for its use, and a training procedure with video clips that will promote highly reliable use of the instrument by other researchers. In the long-term, the new measure, SDMo, will contribute to uncovering the real prevalence of SDM in the care of patients with multiple chronic health conditions and improve the quality and consistency of research on SDM in this priority population. Ultimately, a comprehensive, valid, and reliable measure of SDM will allow clinical researchers to rigorously test associations between the practice of SDM and behavioral and physical health outcomes and facilitate improvements to clinical practice based on new evidence.

NIH Research Projects · FY 2025 · 2022-09

PROJECT ABSTRACT Southeast Asian (SEA) Americans are diagnosed with hepatocellular cancer (HCC) at 8 times the rate of the general population and at twice the rate of other Asian Americans. Poignantly, most of these patients are never treated for their cancer. These troubling observations – coupled with the candidate/her mentoring team’s preliminary data which point to persistent, contemporaneous disparities in cancer care among SEA Americans – provide the impetus for this K23 mentored career development application. This proposal is the logical next step to support Dr. Tran’s long-term goal of becoming an independent investigator in disparity research in HCC. The current proposal includes one of the first comprehensive approaches that focuses on social determinants of health; builds on this team’s prior data on transportation/distance to cancer care to explain the foregoing disparities; leverages resources from within the multi-site Mayo Clinic (Minnesota, Arizona, Florida) and the 3-state (Minnesota, Wisconsin, Iowa), community-based Mayo Clinic Health System; and capitalizes on Dr. Tran’s passionate interests and growing expertise in HCC and disparities and on the capabilities of her 5 well-established mentors. This 2-aim proposal first seeks to characterize the social determinants of health – specifically travel-limiting access to cancer care as well as other determinants including joblessness, trouble finding childcare, limited education, and others – among Hmong and Vietnamese Americans and non-Asian Americans. Dr. Tran will create and maintain a prospective cohort of patients with HCC – enriched with those of SEA American heritage – to investigate the relationships between social determinants of health, ethnicity/race, the rendering of cancer therapy, and cancer outcomes. Then, in aim 2, to understand barriers to access of cancer care among SEA Americans with HCC, she will conduct one-on-one qualitative interviews to learn directly from patients how to overcome barriers to the receipt of cancer care. She will also use these interviews to explore low rates of viral hepatitis treatment and low rates of HCC screening. During the 5-year grant period, Dr. Tran will acquire new skills in qualitative methods, statistics, and survey research through coursework and hands-on experience. She will work closely with her outstanding multi-disciplinary team of mentors (a medical oncologist, health disparity researchers, statistician, and qualitative researcher); develop and hone skills in managing a multi-site team; acquire greater expertise in writing manuscripts, grants and become more adept in presenting her data. The long-term impact of this research is to further understand and obtain strong preliminary data on the social determinants of health in SEA Americans with liver cancer and to position Dr. Tran to implement novel and feasible interventions in a future R01 application with the goal of eliminating these disparities in SEA Americans with HCC.

NIH Research Projects · FY 2024 · 2022-09

Project Summary/Abstract: Urinary stone disease (USD) is third most common and painful urological disease in men and women. Prevention of USD and its associated costs and morbidity requires an understanding of early and late USD pathogenesis. Emerging evidence suggests interactions between intrarenal crystal nucleation, growth, and phagocytic cellular responses plays a key but unrecognized role in USD. Studies in vitro demonstrate that calcium phosphate (CaP) and calcium oxalate (CaOx) crystals induce renal tubular and phagocytic cell secretion of cytokines, chemokines, and extracellular vesicles (EVs; exosomes and microvesicles). These biomarkers can attract blood or residential monocytes and convert monocytes into pro (M1) or anti (M2)- inflammatory macrophages (Mφ’s). Observations in experimental animal models and human tissues suggests that renal tissue monocytes and Mφ’s can phagocytose and metabolize crystals, and urinary stone formers appear to have increased medullary M1 and decreased M2 Mφ populations. In a hyperoxaluric mouse model, suppression of monocyte to M2 Mφ conversion significantly increased intrarenal CaOx deposition. Our studies also demonstrated that urinary excretion of EVs bearing inflammatory markers derived from specific segments of renal tubules were significantly lower in idiopathic calcium stone formers (ICSFs) compared to controls. Thus, multiple lines of evidence suggest that tubular and monocyte derived Mφ populations can phagocytose and degrade crystals as a crystal clearance mechanism, and defects in these clearance mechanisms could result in interstitial Randall’s plaque (RP) and collecting duct plugs (CDP) or even grow directly into USD. The proposed research project is designed to evaluate the role of Mφ’s in RP and CDP formation using a novel hypercalciuric claudin-2 global knockout mouse model (over 3-24 months age) that resembles the phenotype of patients with idiopathic hypercalciuria and USD (Aim 1), and to define the frequency and spatial distribution of monocyte/ Mφ populations in carefully phenotyped ICSFs (20-70 years) with hydroxyapatite, brushite, and calcium oxalate stones and varying amounts of RP (Aim 2). The proposed innovative study will elucidate the role of renal medullary pro-and anti-inflammatory phagocytic cells in the development of RP, CDP, and USD and whether urinary cytokines, chemokines or EVs carrying biomarkers of pro-/anti-inflammatory phagocytic cells can be used to non-invasively monitor intrarenal crystal deposition. Completion of this study will also facilitate the formation of a skilled multidisciplinary team including a promising early-stage surgeon-scientist (Dr Kevin Koo) under the mentorship of an experienced and skilled USD clinical and researcher (Dr. Lieske). The resulting preliminary data will provide evidence of the effectiveness of our team. This work will also enable submission of future detailed grant or center proposals that will extend these mechanistic studies, and has great potential to elucidate underlying pathogenic steps in USD genesis and identify novel therapeutic targets.

NIH Research Projects · FY 2025 · 2022-09

ABSTRACT Osteoarthritis (OA) is a chronically painful condition and a leading cause of disability around the world. OA is characterized by articular cartilage deterioration and other joint pathologies including meniscus and ligament damage, synovitis, bone spurs and joint pain. None of the currently approved treatments alter disease progression by slowing articular cartilage degradation or repairing damage to joint tissues. We identified Phlpp phosphatases as modifiable targets for cartilage regeneration and joint pain. Phlpp1 and Phlpp2 are abnormally expressed in human OA cartilage. Phlpp1 knockout (KO) mice are protected from cartilage degradation and pain-related behaviors (allodynia and reduced mobility) after surgery that destabilizes the medial meniscus (DMM), but mice where Phlpp1 is conditionally depleted in just aggrecan (Agc)-expressing cells (Phlpp1 CKOAgc) are only protected from cartilage degradation, not reduced mobility. These results indicate an intrinsic role for Phlpp1 in chondrocytes, as well as a role for Phlpp1/2 in other tissues or cells within articulating joints. Of interest, small molecule inhibitors of Phlpp1 and Phlpp2 further slow cartilage degradation in Phlpp1 CKOAgc mice and increase mobility. Phlpp1/2 inhibitors also stimulate chondrocyte proliferation and matrix production and prevent neurite outgrowth and expression of sensory neuron genes in vitro and in vivo. Together these data indicate that Phlpp1/2 activity in multiple cell types/tissues within articulating joints contributes to OA pathogenesis and that Phlpp2 may contribute to OA phenotypes by compensating for Phlpp1 deficiency in cartilage and/or by regulating joint innervation and pain-related behaviors. The overall goals of this project are to define how Phlpp1 and Phlpp2 modulate cartilage quantity and quality (i.e. stiffness) and how Phlpp1/2 inactivation contributes to injury-induced innervation. The specific aims are to: 1) Elucidate the roles of Phlpp1 and Phlpp2 in articular chondrocytes following a joint injury, 2) Determine how Phlpp1 and Phlpp2 affect biomechanical and structural properties of articular cartilage and other joint tissues after injury and throughout aging, and 3) Define activities of Phlpp1/2 in sensory neurons and OA-associated pain behaviors. These studies will provide new information about the relationship between joint damage, joint innervation and pain. Such insights will enable the development of much needed disease- modifying drugs for osteoarthritis.

NIH Research Projects · FY 2025 · 2022-09

Project Summary/Abstract Neurological and psychiatric disorders affect millions of people in the United States and worldwide, and produce a third of all health care costs. Recent research has produced encouraging evidence that adaptive neuromodulation can induce nervous system plasticity that produces long-lasting improvements in certain neurological disorders such as stroke. At the same time, it is becoming increasingly clear that the technologies that support these demonstrations remain painfully inadequate and inaccessible for both research and clinical application: current non-invasive technologies are typically imprecise; and current invasive technologies, which are more precise, are currently only available with serious restrictions for human use. Moreover, all of the few existing neuromodulation platforms for human use require substantial expertise in diverse areas of engineering, physiology, and regulatory domains that is not available to most groups. This lack of availability of sufficiently capable and readily useable neuromodulation technologies greatly impedes the development, application, and optimization of new adaptive protocols for improving symptoms of devastating neurological and psychiatric disorders. The purpose of the project proposed here is to address this critical issue by developing, validating, and widely sharing with the community an easy-to-use adaptive neuromodulation ecosystem (comprised of technology and protocols) that is optimized for the needs of invasive basic and clinical research. We will validate this ecosystem in a canine model, and disseminate it with appropriate documentation to other scientists and clinicians through three project-related test sites and three workshops. In accord with this objective, we will: 1. Develop a general-purpose hardware/software neuromodulation platform for invasive neuromodulation research 2. Develop and validate an ecosystem for adaptive neuromodulation research and clinical application 3. Disseminate this ecosystem of technologies and protocols Achieving these three aims will create, validate, and disseminate the first comprehensive ecosystem that facilitates the conception, development, and clinical application of invasive adaptive neuromodulation protocols. We expect that the availability of this ecosystem will greatly increase activities in basic and clinical neuromodulation research that will lead to new understanding of the neural underpinnings of normal and abnormal function and will thereby accelerate the development of novel adaptive neuromodulation protocols to improve treatment for many devastating neurological disorders.

NIH Research Projects · FY 2025 · 2022-09

PROJECT SUMMARY/ ABSTRACT Myofascial pain syndrome (MPS) is a common public health problem. Knowledge of MPS injury mechanisms and treatment of this condition is currently limited by a lack of objective assessment tools. Efforts to better understand the origin and pathology of MPS have increasingly focused on impairments involving myofascial connective tissue and the function of fascial interfaces. Studies using ultrasound imaging technology to evaluate the function at sliding myofascial interfaces have provided insights into the underlying mechanisms of the syndrome. Researchers suggest that alterations in the viscoelastic properties of fascial structures may contribute to the MPS etiology. This may be perceived by patients as an increase in fascial stiffness and pain with restricted motion. However, major knowledge gaps remain in the understanding of myofascial biomechanics and how changes in these structures contribute to myofascial pain. Development of technology capable of providing biomarkers that quantitatively characterize the viscoelastic properties of myofascial tissue and the state of adhesion at interfaces would address these gaps and contribute to the assessment of therapeutic modalities. Currently, a noninvasive tool for quantifying fascia mechanical properties is very limited. Our goals are to (1) develop an MRE-based imaging technique for quantifying the mechanical properties of myofascial tissue and (2) establish new quantitative biomarkers for assessing impaired myofascial tissue and treatment efficacy. In Aim 1 (R61 phase), we will build an MRE-based framework to integrate multiple driving systems inducing desired shear motion in the lower back, upper and lower legs, respectively; an advanced pulse sequence to measure the corresponding full-volume dynamic 4D muscle displacement fields; and a post-processing approach to assess resultant mechanical parameters of the myofascial interface in those regions in vivo. This will create a foundation to characterize the myofascial interface mobility, stiffness, viscosity, and loading sensitivity. In Aim 2 (R61 phase), we will evaluate the repeatability and reproducibility of the MRE-assessed fascia mechanical properties in healthy volunteers using a test-retest strategy. A pilot clinical study will also be performed to evaluate and compare MRE-assessed fascia mechanical properties in age-/sex-matched normal and patients with conditions in the MPS spectrum. The transition milestone we are looking for is imaging biomarker(s) that demonstrate a statistically significant (p < 0.05) group difference. In Aim 3 (R33 phase), we will assess the abilities of the quantitative biomarker(s) developed in the R61 phase to monitor treatment responses to a physical force-based manipulation treatment (Tuina) and predict outcomes in a longitudinal study. Taken together, these aims will provide innovative methods and unique datasets for studying myofascial biomechanics, novel imaging biomarkers to distinguish healthy versus abnormal myofascial tissue and interfaces, and new imaging biomarkers to aid clinicians in developing effective approaches to myofascial pain, and helping to address one of the most important conditions that has led to overuse of opioid analgesics.

NIH Research Projects · FY 2025 · 2022-09

PROJECT SUMMARY Osteoporotic fractures threaten the health, independence, and survival of millions of people nationwide. An estimated 40% of women and 30% of men will suffer a hip, spine, or wrist fracture in their lifetime. Two million Americans experience a fracture annually, resulting in more than 430,000 hospital admissions, 2.5 million medical office visits, and 180,000 nursing home admissions. Due in part to an aging population, the cost of osteoporotic fracture-related care will exceed $25 billion by 2025. This suffering and cost is preventable. Large randomized controlled trials (RCT) have demonstrated the substantial benefit of osteoporosis drug therapies (ODT) in reducing the risk of osteoporotic fractures. Yet, fewer than 50-80% of patients at risk of fracture will receive ODT and half will discontinue them prematurely. Underuse of, and poor adherence to, ODTs stems in part from the lack of evidence about the effectiveness and safety long-term use of anti-resorptive ODT (i.e., bisphosphonates and denosumab), particularly with respect to rare side effects such as atypical femur fracture (AFF) and osteonecrosis of the jaw (ONJ). While interruption of long-term ODT (‘drug holiday’) and use of sequential therapies (i.e., anabolic ODT followed by anti-resorptive ODT) have been proposed as ways to limit risks of AFF and ONJ, it remains unknown whether these strategies actually reduce risk of harm without compromising ODT effectiveness with respect to fracture prevention. We will address these knowledge gaps by emulating a series of target RCTs examining the comparative effectiveness and safety of ODT regimens with respect to fragility fractures (primary outcome), AFF, ONJ, and other safety endpoints. We will use claims and electronic health record data from OptumLabs Data Warehouse, a dataset of privately-insured and Medicare Advantage beneficiaries, linked to a 100% sample of Medicare fee-for-service claims, to allow for an unprecedented evaluation of ODT over time and across populations, geographies, health systems, and health plans. We will emulate the following target RCTs (eRCTs): Aim 1) eRCT 1 comparing ≤3 years vs. >3 years non- interrupted anti-resorptive therapy. Aim 2) eRCT 2 comparing non-interrupted long-term biphosphonate ODT (>3 years) vs. short-term (≤3 years) ODT followed by either brief (≤3 years) or prolonged (>3 years) drug holiday. Aim 3) eRCT 3 comparing sequential therapy with anabolic ODT followed by >3 years bisphosphonate vs. denosumab treatment. Within each eRCT, we will assess for heterogeneity of treatment effects as a function of gender and risk profile (e.g. steroid use, etc). Finally, to address the gaps in translation of evidence to patient care decisions, Aim 4 will update and field test an effective but outdated shared decision-making tool (Osteoporosis Choice) with data produced by Aims 1-3. At the conclusion of this work, we will generate both the evidence to inform high quality osteoporosis care and a ready-to-use shared decision-making tool to support the implementation of this evidence into practice to improve the health of patients with osteoporosis.

NIH Research Projects · FY 2026 · 2022-09

PROJECT SUMMARY/ABSTRACT A major breakthrough in cancer immunotherapy in the last decade is the development of chimeric antigen receptor (CAR) T cell therapy. Although anti-B cell maturation antigen (BCMA) directed CART cell therapy demonstrated unprecedented initial responses in patients with relapse/refractory B-cell malignancies or multiple myeloma (MM), respectively, durable responses are limited. The mechanisms of relapse after CART cell therapy are not fully unveiled yet. The long-term goal of my proposal is to uncover mechanisms of resistance to CART cell therapy and to develop an independent research career focused on targeting the tumor microenvironment simultaneously with CART cells. The unifying objective of this application is to design novel strategies for the treatment and prevention of tumor immunoescape based on my clinically relevant preliminary data, using BCMA CART cell as a model. The central hypothesis is cancer-associated fibroblast (CAFs) induce CART dysfunction through complex mechanisms and that dual targeting of tumor cells and CAFs is safe and enhances the efficacy of CART cell therapy. To test this hypothesis, I have designed three specific aims: Aim #1) Determine the mechanisms of CAF-induced BCMA CART cell dysfunction in MM patients; Aim #2) Study the potential toxicity of dual targeting CART cells in syngeneic humanized MM-tumor microenvironment (TME) models; Aim #3) Utilize samples from patients who were treated with BCMA CART cell therapy to study the interactions between CAFs and CART cells. The rationale for this proposal is that in my established MM-TME model, CART cell trafficking to the tumor site is significantly inhibited. My preliminary data also showed that CAFs suppressed CART cell functions by secreting TGF-β and inhibitory cytokines as well as altering the PD-1/PD-L1 axis. Dual targeting tumor cells and CAFs significantly improved CART cell functions. This research will be significant because it will contribute depth (of understanding the CART cell resistance) and breadth (of novel potentially curative therapy) to the immunotherapeutic strategy against not just MM but also solid tumors. Ultimately, such discoveries have the potential to vertically advance the field of CART cell immunotherapy as well as other targeted immunotherapies. This project is innovative because it combines high throughput screening techniques to study the possible mechanisms of resistance after immunotherapy and, therefore, to generate novel treatments. The proposed research activities are crucial to the development of the applicant as an independently funded scientist with a focus on cellular immunotherapy. I will receive further training in molecular biology, translational medicine, and immunology from my mentors and training in cancer biology and bioinformatics from experienced collaborators at the Mayo Clinic. Therefore, at the conclusion of the training period, I will have acquired a unique set of intellectual and technical skills that will allow me to promote myself to become an independent translational researcher.

NIH Research Projects · FY 2025 · 2022-09

Chronic lymphocytic leukemia (CLL) is the most common leukemia in the United States and still remains incurable. Due to its impaired immune response, CLL has high number of morbidity and mortality complications including increased risk of infections and second cancers. Therefore, identifying individuals who are at markedly higher risk of developing this disease may enable future prevention strategies. Monoclonal B-cell lymphocytosis (MBL) is the precursor state to CLL. The prevalence of MBL increases with age and is found in almost a third of Caucasians older than 60 years. Although all individuals with CLL pass through the MBL precursor state, the reason why some individuals with MBL progress to CLL yet many do not is unclear. Therefore, there is a need to identify biomarkers that differentiate those MBL who will progress versus those who will remain asymptomatic. The overall goal of this application is to address this knowledge gap by evaluating genetic and epigenetic factors associated with risk of progression to CLL among an established cohort of 1,729 individuals with MBL. In Aim 1 we will analyze the polygenetic risk score (PRS) comprised of a weighted average of 41 inherited single nucleotide polymorphisms (SNPs) that have been previously identified through genome wide association studies (GWAS) of CLL with risk of progression to CLL. In Aim 2 we will perform deep targeted sequencing of 59 putative CLL driver genes to investigate if individual genes with high-impact mutations or the aggregate number of mutated genes leads to an increased risk of progression from MBL to CLL. Finally, in Aim 3 we will evaluate whether methylation signatures that classify MBL individuals into low-, intermediate-, and high-risk predict progression to CLL. At the completion of this application, we will have identified three complementary yet independent genetic and epigenetic factors that we hypothesize will be strong predictors of progression to CLL among a cohort of individuals with MBLs. Our preliminary data support our hypotheses. We have the largest cohort of individuals with MBL collected in US, putting us in an unsurpassed situation to prospectively evaluate the effect of known CLL risk factors at the precancer phase. Our integrative predictive model of all three biomarkers may change current practice guidelines and ultimately improve quality of life by reducing anxiety and distress for individuals in pre-malignant phase.

NIH Research Projects · FY 2025 · 2022-09

Limb regeneration after injury is a sophisticated and energetically expensive process. In this process, progenitor proliferation, osteoblast differentiation and mineral deposition all require sufficient supplies of biological building blocks and ATP(1-5). However, the contribution of cell metabolism and its genetic control of skeletal regeneration is largely unknown, representing a major knowledge gap. Using the established mouse digit tip amputation model, we found that mice exhibit impaired regeneration during aging, and that this impairment is linked to increased expression of glycolysis and oxidative phosphorylation (OxPhos) genes, compared to young mice. These exciting preliminary results have led us to investigate the metabolic and genetic mechanisms that underlie skeletal regeneration. Our preliminary findings support that skeletal regeneration is metabolism-dependent and can be manipulated by exogenous metabolites and gene expression, respectively. These data suggest that administration of oxaloacetate (OAA), a pro-glycolytic and pro-respiratory metabolite, increases regenerated bone volume and thickness in a mouse model. Alternatively, modulation of collagen triple helix repeat containing 1 (Cthrc1) also alters skeletal regeneration. Cthrc1 is specifically expressed in the blastema, the dedifferentiated tissue structure central to regeneration, and Cthrc1-/- mice demonstrated impaired regeneration and dysregulated cell metabolism. Moreover, our preliminary data show that treatment with OAA increases Cthrc1 expression, reinforcing a direct link between metabolism and genetic control. We hypothesize that a finely tuned interaction between cell metabolism and genetic control synergistically regulates cell function, and that this interaction can be manipulated both exogenously (OAA) and at a gene level (Cthrc1) to modulate regenerative outcomes.

NIH Research Projects · FY 2025 · 2022-09

Differences in cancer pain are profound and uniquely harmful among some cancer survivors as they may undermine their already limited ability to access, tolerate, and/or receive treatment for their cancer. Differences relate to poor care, needless persistence and intense pain, as well as the over- and under-prescribing of opioids. Multi-modal pain care (MMPC), a robustly validated, safer, and more effective alternative to a solely medication-based approach has proven challenging to implement broadly, and virtually impossible in resource limited and isolated settings. The factors that impede delivery of MMPC; provider bias, patients’ reluctance to report pain, and lack of patient-centered MMPC options, also mediate differences making them key targets for improvement. The Collaborative Care Model (CCM) provides a well-established and validated framework that can neutralize factors that perpetuate differences, guide MMPC delivery, and improve pain detection and treatment. However, as currently configured the CCM’s single symptom emphasis needs to be modified to address the multi-level drivers of pain. Our team has developed and tested CCM iterations that integrate elements of team-based care (TBC) to improve the CCM’s monitoring of patients’ needs, as well as to accommodate MMPC’s multi-disciplinary care requirements. In addition, we have leveraged electronic health records (EHRs) to enable care teams to link symptomatic cancer patients with MMPC providers and resources. Our prior research deploying CCM-TBC hybrid interventions with patient-and-care team-centered EHR-reengineering has also significantly improved patient symptom reporting and deployment of MMPC. These efforts, while fruitful, have also shown us that a broader EHR retrofitting is required to address the breadth of patients’ needs and the requirements of real-world clinical workflows. This experience suggests that a flexible, modular CCM-TBC hybrid system, supported by EHR enablement, can deliver high fidelity MMPC in a manner that improves care and pain symptom experiences at multiple levels among cancer survivors. We plan to evaluate the effectiveness of this approach in a clinical trial entitled “Advancing Safe, Comprehensive, Digitally-Enabled Cancer Pain managemeNT (ASCENT).” More specifically, we will partner with our community stakeholders during an initial, 1-year R61 development phase to refine a patient-centered version of our CCM-TBC hybrid that addresses the needs and preferences of cancer survivors most susceptible to poor pain outcomes (Aim 1). After confirming the functionality of the intervention’s components, we plan to transition to a 4-year R33 execution phase with a 2-arm, parallel group randomized clinical trial. This trial (Aim 2) will be conducted in 4 semi-autonomous Health Care Systems and is designed to assess whether our tailored/personalized CCM-TBC hybrid intervention improves pain outcomes relative to usual care among 578 survivors susceptible to poor pain outcomes. Primary (pain) and secondary (mood, sleep, physical function, work status, and healthcare utilization) outcomes will be assessed at 0, 3, and 6 months. All data, excepting patient reported outcome measures, will be extracted from the EHR for main effects, as well as exploratory mediator and machine learning analyses; the latter to identify characteristics associated with positive responses. Aim 3 will evaluate implementation strategies to support multistakeholder adoption and use of intervention components.

NIH Research Projects · FY 2025 · 2022-09

PROJECT SUMMARY/ABSTRACT Progressive supranuclear palsy (PSP) is a fatal, progressive, incurable neurodegenerative tauopathy. Falls resulting from gait and balance impairments are a cardinal feature of the disease and occur in all PSP syndromes. Currently, detection of these abnormalities relies on largely subjective clinical examination and scales. There are no standardized ways to quantify specific features of gait and balance impairments in PSP. Diagnostic, and prognostic tests focusing on patient relevant outcomes such as gait related disability are needed. Motion analysis technology can be used for objective qualification of gait and balance variables. This offers an opportunity to develop patient-centric diagnostic tools that can measure features of gait impairment and its impact on risk for falls. Laboratory-based video motion capture utilizes multiple high frequency cameras in a well calibrated controlled environment to generate a three-dimensional model of human locomotion and balance. This is considered the most accurate and reliable method of motion analysis. Our project will utilize laboratory-based motion capture to identify gait metrics that best characterize PSP Richardson syndrome, PSP-cortical and PSP-subcortical syndromes and distinguish PSP from normal controls. However, accredited motion analysis labs are not readily accessible, and require an elaborate usually expensive set-up. Body-worn motion-sensing devices offer a feasible alternative. They are cost-effective, portable, easy to use in ambulatory settings. However, it is essential to establish reliability and accuracy of these devices in specific patient populations and assess clinical significance of the data captured. Therefore, in this project we will employ a body-worn motion sensing device or inertial monitoring unit (IMU) to detect gait and balance metrics that best characterize PSP. The data from the IMU will then be compared to the motion analysis lab to establish its accuracy. Gait metrics have the potential to serve as patient-centric outcome measures in PSP. In this project we will also use these variables to analyze neurobiological mechanisms underlying gait and balance impairment in PSP. We will correlate the identified gait and balance metrics with volumetric magnetic resonance imaging (MRI), diffusion MRI (dMRI) and tractography to determine patterns of disruption of cortical and subcortical motor control systems in PSP. The effect of sex as a biologically significant variable will be assessed. Age will be a covariate to identify gait abnormalities that are specifically related to disease state and not secondary to normal aging. The outcomes of this research project will contribute to development of patient centric outcome measures, assessment of clinical heterogeneity and understanding of mechanisms of gait impairment in PSP syndromes. Future directions will include longitudinal assessment and comparative analyses across related neurodegenerative diseases to advance diagnosis and development of patient centric outcome measures for clinical trials.

NIH Research Projects · FY 2025 · 2022-09

This NIA K07 Academic Leadership Career Award resubmission entitled, “Older Sexual Minority Patients with Serious Illness: Program-Building to Identity and Address Needs,” seeks to help these understudied and underserved groups of patients, who, in the face of advanced age and serious illness, are more likely to be single, living alone, without children, lacking in social support, and at risk for and fearful of untoward health issues and outcomes. The applicant is a senior physician at Mayo Clinic and has a long track record of research in studying symptoms and psychosocial issues in ill patients (400+ publications), mentoring (50+ mentees + principal investigator/program director of career development program K12CA090628-20), and serving as a leader in both her home institution and nationally. This track record and these experiences are brought to this application with the goal of understanding and addressing the unmet needs of these groups of older sexual minority (SM) patients – defined (revised) by the NIH as “individuals who identify as lesbian, gay, and bisexual,.” A uniquely rich institutional environment that includes the Kogod Center on Aging; the Mayo Clinic Cancer Center (an NCI-designated comprehensive cancer center); a robust Center for Clinical and Translational Science (CCATS); a nationally renowned statistics and data center; the robust Mayo Clinic practice, which provides depth and breadth of clinical expertise in the management of a vast array of serious illnesses to over 1 million patients per year; and, last but not least, the 120,000+ (and growing) social media platform, Mayo Clinic Connect -- perhaps the largest such patient-centric electronic medical resource in the world -- will set the stage for the development of this programmatic infrastructure. Novel, ground-breaking research early in the grant-cycle will consist of a needs-assessment that uses both qualitative and quantitative methodology to learn the unremitting symptomatology and unrelieved psychosocial hardships of these older patients and that relies on Mayo Clinic Connect (an estimated 30% of users self-report as 65+ years of age) to reach older SM patients anonymously and to learn confidentially about their concerns. This needs assessment will give rise to a series of annual requests for pilot grant applications aimed at young investigators. Young investigators who are awarded a pilot project will acquire mentoring from the applicant, engage in coursework, participate in a weekly seminar series, learn from a biannual visiting professor series, begin to learn to contribute to CCATS course development on older SM patients, and further launch their careers as investigators in this field. National/international experts in their respective fields and a patient advocate will serve on an advisory committee, providing programmatic guidance over the 5-year grant cycle. These efforts are destined to establish a sustainable, funded research program to improve the lives of older SM patients with serious illness.

NIH Research Projects · FY 2025 · 2022-09

PROJECT SUMMARY/ABSTRACT There is a fundamental gap in understanding the immunological paradox by which solid tumors first metastasize via lymphatic channels into the immune-rich lymph node. During this process, immune surveillance is disabled, ultimately allowing metastatic dissemination to the precise location where it should be first eliminated—the tumor-draining, sentinel lymph node (SLN). While notably prognostic, it is clear the metastatic status of the SLN alone is not sufficient in determining patient’s risk or relapse. Therefore, understanding the mechanisms underlining this chronic process of tumor mediated regional immunosuppression, commonly referred to as pre-metastatic niche (PMN) formation in the SLN will lend significant insights for developing improved prognostic and therapeutic tools to detect and reverse cancer dissemination early in the natural history of metastatic progression. Our long term goal is to develop therapeutic strategies capable of overcoming the immune compromise of the SLN PMN and thereby disrupt the first stage of cancer metastasis. Therefore, the objective of the current work is to mechanistically interrogate the process by which the subcellular component of the primary tumor lymphatic effluent directly mediates PMN formation. The central hypothesis proposes that in solid tumors, subcellular mediators derived from the primary tumor microenvironment actively traffic through the lymphatics and in a cargo-dependent manner create a PMN in the tumor-draining SLN. This hypothesis has been formulated on the basis of preliminary data produced in the applicant’s laboratory; namely the discovery and characterization of human lymphatic extracellular vesicles (L-EV) which have a demonstrated capacity to modulate immune function. The rationale asserts that in elucidating the factors and signatures that define PMN formation in the SLN, the knowledge gained will be significant as it will identify histopathologic biomarkers that could aid in patient risk stratification beyond the presence of melanoma cells in the SLN. Guided by strong preliminary data, this hypothesis will be tested in two specific aims: 1) identify mechanisms whereby lymphatic subcellular factors promote immune dysfunction in the pre-metastatic SLN beyond those already identified (i.e. S100A9); 2) evaluate the prognostic utility of these immune-modulating factors in predicting risk of recurrence by considering the comprehensive, interactive cellular landscape that defines the immunologically compromised SLN. The approach is innovative as it will use a mechanism-driven model to identify subcellular factors from a previously uncharacterized biological fluid, human lymph (Aim 1), complemented by a novel, multiplexed biomarker imaging approach in order to survey the SLN immunological landscape in a quantitative and spatially preserved manner to ultimately translate predictive features into clinically amenable platforms (Aim 2). Such findings will result in a refined definition of early-stage patients at risk of relapse and in need of earlier interventions.

NIH Research Projects · FY 2025 · 2022-09

ABSTRACT Urinary stone disease (USD) affects up to 12% of individuals in developed countries, and is associated with significant morbidity, pain, and annual US health care costs of >$10 billion. There is strong evidence of heritability, however risk factors remain relatively obscure. Our prior work in the Rare Kidney Stone Consortium (RKSC) developed a targeted nephrology gene capture panel (tNGS) with 144 known and candidate USD genes and resolved ~30% of >500 patients suspected of monogenic USD due to early onset nephrolithiasis (NL), nephrocalcinosis (NC), chronic kidney disease (CKD), and/or multiple stones, with ~15 different genes implicated. In addition to making a definitive diagnosis, in many cases the resulting information revealed new treatment options or directed patients to specific clinical trials. Here we propose extending our understanding of the spectrum of genes and variants associated with monogenic NL/NC in the RKSC cohort, including functional in vitro analyses of variants of uncertain significance (VUS; Aim #1). In parallel, we will study a cohort of >50,000 Mayo Clinic biobank patients with detailed linked medical record information (~12.7% with stones) (Aim #2). Findings in this Biobank cohort will be validated and extended in a Prospective Kidney Stone Cohort of more than 550 first time USD patients and 480 matched controls (Aim #3). This third community-based cohort has detailed clinical and biochemical data, including comprehensive assessment of symptomatic and radiographic recurrence, and will thus allow detailed study of potential mechanism(s) of implicated genes. This proposal takes advantage of available genetic analyses in Aim #2 and Aim #3 that includes whole exome sequence (WES) as well as genomewide single nucleotide polymorphism (SNP) data for association studies. Studies will determine if coding, noncoding, and expression related variants of monogenic USD genes are associated with NL/NC risk and correlate with biochemical determinants of stones. Our Specific aims are: Aim #1: Genotype patients suspected of monogenic forms of USD using global NGS approaches; Aim #2: Validate identified genetic USD risks in a Mayo Clinic Biobank cohort including first time and recurrent calcium stone patients and controls with WES and SNP association studies; Aim #3: Characterize genetic USD risks and pathways in a prospective chart-validated cohort of first-time symptomatic calcium stone patients with age-sex- matched controls with WES and SNP association studies. Our overall hypothesis is: variants in monogenic USD genes not only explain Mendelian disease, but are risk factors for the common (predominately calcium) USD that is found in the general community. This study will improve understanding of the etiology and pathogenesis of monogenic USD and impact patient treatments and clinical trial recruitment. Risk factors for common US and its recurrence will also be identified.

NIH Research Projects · FY 2025 · 2022-09

PROJECT SUMMARY Norepinephrine (NE)-containing neuronal cells in the locus coeruleus (LC) are thought to co-release dopamine (DA) from many of its projections, including into the hippocampus. Through DA and NE release, the LC is thought to play a major role in modulating hippocampal memory encoding through the maintenance of long-term potentiation (LTP), an integral mechanism for memory consolidation. Understanding the mechanisms by which the LC modulates memory is of fundamental importance to investigations into hippocampal memory circuitry and dynamics. However, because of their structural similarity, it has been difficult to ascertain the precise roles that DA and NE each have on LTP maintenance. In vivo microdialysis has been traditionally used in the past to measure tonic extracellular concentrations of molecules in the brain, but damage to tissue because of the size of the sampling probe and low spatiotemporal resolution make real-time tracking of tonic neurotransmitter concentrations, and their biological functions, problematic. Our lab has developed state- of-the-art voltammetric techniques capable of measuring tonic concentrations of neurotransmitters in real-time with very high spatial resolution. We hypothesize that by altering the voltammetric waveform applied in vivo and by developing a novel artificial intelligence based post-processing pipeline, very similar analytes such as DA and NE can be reliably resolved. Through accurate neurotransmitter identification and pharmacologic manipulation, we aim to ascertain the specific effects NE and DA have on hippocampal LTP. Additionally, we hypothesize that electrical stimulation of the LC will lead to increased hippocampal DA and NE release and enhanced LTP induction compared to no stimulation. The ability to resolve individual analytes based on their voltammetric signals has been an unsolved problem in electrochemistry and will enable tracking of their relative real-time contributions to LTP induction and maintenance in the hippocampus with high accuracy. A greater understanding of LTP induction and maintenance mechanisms is of vital importance to garnering an increased understanding of memory circuitry and physiology.