University Of Miami School Of Medicine

NIH Research Projects · FY 2025 · 2024-02

Project Summary Diabetes is a leading cause of end-stage kidney disease (ESKD), with about 55-600% of patients on dialysis having diabetes. Glucose management in the context of dialysis is challenging. In recent studies, we reported that patients with diabetes and ESKD have increased risk of severe hypoglycaemia and hyperglycemic crises. However, there is lack of contemporary, nationwide data about how diabetes is managed among these patients, and how each treatment regimen may impact the risks of acute preventable complications (e.g., severe hypoglycaemia and hyperglycemia) and health outcomes (e.g., readmissions, mortality). Similarly, it is unknown whether there is a relationship between specific anti-diabetic agents or insulin formulations and the rates of diabetes crises among minority groups, such as African Americans and Hispanics, which are disproportionately affected by diabetes and ESKD. In this observational study, we propose to use the most comprehensive database of patients with ESKD, the United States Renal Data System to (aim 1) characterize contemporary, real-world patterns of glucose-lowering therapy and diabetes technologies in this population, which may diverge from what is recommended in clinical guideless or explicitly studied in clinical trials. We will also (aim 2) examine health-related outcomes, specifically all-cause and cause-specific (i.e., for hypoglycemia, hyperglycemia) hospitalizations, readmissions, and mortality, as a function of glucose-lowering regimens (e.g., human and analog insulin, GLP-1 receptor agonist, DPP-4 inhibitor). Our analyses will also evaluate for differences by diabetes type and for disparities based on patient’s race/ethnicity and sex. We anticipate identifying the most vulnerable subgroups and treatment agents associated with higher risk of acute glycemic complications, which will have downstream impacts such as translational research trials and management recommendations. With support from a K23 grant (5-K23DK123384-04), Dr. Galindo is currently assessing if (aim 1) a novel, factory-calibrated, continuous glucose monitoring (CGM) system can provide better assessment of glycemic excursions compared to the standard-of-care using capillary glucose testing, and (aim 2) if using real-time CGM will prevent hypoglycemic events among patients with diabetes and ESKD on dialysis using insulin therapy, and provide better overall glycemic control. Findings from this study R03 study, coupled with findings from my K23 prospective study, will have strong clinical and translational impact, and will be used to design randomized trials focusing on de-escalating or tailoring patient-centered efficacious and safer anti- diabetic regimens, using an improved glycemic monitoring systems (e.g., CGM) for this high-risk group in subsequent R01 applications.

NIH Research Projects · FY 2025 · 2024-02

Project Summary/Abstract Penumbral salvage is a central dogma of acute stroke care. Radiological characterization of the penumbra has transformed acute ischemic stroke care by carefully selecting patients who may benefit from acute intervention. No such innovation has taken place in intracerebral hemorrhage (ICH). Our central hypothesis is that there is a “mechanical penumbra” surrounding the hemorrhage - an area that is mechanically compressed and at risk of becoming irreversibly damaged – but still salvageable if the compression is relieved in a timely manner. Randomized clinical trials of cerebral hematoma evacuation have not consistently shown a benefit. A potential explanation for these results is that we have failed to radiologically select patients who may still have salvageable tissue and are most likely to benefit from hematoma evacuation. Most ICHs occur in the putamen and thalamus, which border the internal capsule (IC). The internal capsule consists of the long axons of the corticospinal tract connecting the cortex with the spinal cord. Structural damage to the IC is the major determinant of functional outcomes after ICH. Preserving the structural integrity of the IC should therefore be the therapeutic goal. Diffusion-tensor imaging (DTI) is a magnetic resonance-based imaging technique which allows for the structural assessment of axons in the white matter, such as the IC. DTI detected damage to the IC highly correlates with long-term functional outcome after ICH. We hypothesize that this mechanical penumbra in ICH can be evaluated using DTI imaging (using fractional anisotropy (FA) values, and other measures). Our preliminary data support the feasibility of using DTI imaging and the ability to identify spatial and temporal shifts of FA values at baseline and in the early hours after ICH. Identifying mechanical penumbra may facilitate the selection of patients for surgical intervention in future clinical trials. In this study, we propose serial MR DTI imaging at the time of ICH presentation (< 12h from symptom onset), at 24h, and at 10-14 days after injury in 24 patients. We will analyze DTI sequences for radiological markers of permanent injury. The IC will be segmented into three zones emanating from the hematoma to determine if there is a gradient of DTI measures (including FA values). We hypothesize that axons closest to the hemorrhage have the lowest (damaged axons) and those furthest away from the hemorrhage have the highest (preserved axons) FA values. Demonstrating such a gradient of FA values around the ICH suggests the presence of axons with variable degrees of injury, some of which may still be salvageable. (Aim 1) We will then investigate the temporal evolution of these axons and correlate the impact of this evolution on functional recovery at 3-month follow-up (Aim 2). We hypothesize that an improved DTI measures is associated with good functional outcomes whereas a deterioration of FA values over time is associated with poor outcomes.

NIH Research Projects · FY 2025 · 2024-02

Exhausted T cells (TEX) differentiate in cancer and chronic infections. TEX do not function optimally and they do not provide protection. Immunotherapy reinvigorates TEX by antagonizing surface inhibitory receptors, such as PD-1. However, the epigenetic stability of TEX inhibits complete reversal of exhaustion, therefore significantly limiting the clinical benefits of immunotherapy. We recently proposed the use of microRNAs to attenuate exhaustion. MicroRNAs regulate cell differentiation by targeting multiple mRNAs. Indeed, microRNA-29a (miR- 29a) regulates CD8 T cell differentiation, enhances CD8 T cell persistence and function and promotes memory- like responses in chronic infection by epigenetically altering TEX differentiation. Therefore, we suggest the use of miR-29a to enhance TEX reinvigoration. However, delivering microRNAs to TEX in vivo is a major challenge. The goal of this proposal is to reinvigorate TEX and improve anti-viral and anti-tumor responses using a novel method for delivery of miR-29a in vivo. Our preliminary data suggest that miR-29a synergizes with αPD-1 to enhance CD8 T cell responses and promote durable, functional CD8 T cell reinvigoration. However, our results are based on retroviral overexpression in CD8 T cells, which does not allow for delivery of miR-29a to TEX later when exhaustion has progressed. Therefore, we developed a novel method to deliver miR-29a to already differentiated TEX in vivo. We conjugated miR-29a to αPD-1 (miR-29a/αPD-1). MiR-29a/αPD-1 increased miR- 29a expression and decreased miR-29a target expression in TEX, suggesting that miR-29a delivery is efficient and functional. Therefore, our overall hypothesis is that miR-29a/αPD-1 alters T cell differentiation, reinvigorates TEX, and enhances anti-viral and anti-tumor T cell responses. Aim 1. Reinvigorate TEX with miR-29a/αPD-1 in vivo. MiR-29a regulates key transcriptional circuits and synergizes with αPD-1 to promote TEX reinvigoration. Therefore, we hypothesize that miR-29a delivery by miR- 29a/αPD-1 can promote TEX reinvigoration both early and late during the differentiation of exhaustion. We will administer miR-29a/αPD-1 to mice with chronic viral infection (LCMV) or tumors (B16 melanoma or KPC-PDAC). We will elucidate the role of miR-29a in TEX reinvigoration during the early and late exhaustion phase. Aim 2. Delineate targeting of immune cells and tumor cells by miR-29a/αPD-1. PD-1 is highly expressed in TEX, but also in other immune cells, and tumor cells. Interestingly, miR-29a can suppress tumorigenesis and inhibit tumor cell proliferation. Therefore, we hypothesize that miR-29a/αPD-1 can provide significant clinical benefits by targeting both immune and tumor cells. To test this, we will use a fluorescence labelled miR-29a/αPD- 1 to elucidate miR-29a delivery and identify other immune cells and tumor cells targeted by miR-29a/αPD-1. At the completion of these studies we will a) demonstrate the ability of miR-29a to prevent and/or reverse exhaustion, b) develop a novel immunotherapeutic to provide immune reinvigoration with a dual effect on immune cells and tumor cells, and c) develop a novel platform for in vivo delivery of miRNAs to TEX.

NIH Research Projects · FY 2025 · 2024-02

PROJECT SUMMARY: Glioblastoma (GBM) is an uncurable form of primary brain tumor with extremely poor prognosis. Despite multi- modal therapy including surgery, irradiation and chemotherapy, all patients experience tumor progression. No standard of care is established in recurrent or progressive GBM. The identification of a neuronal cellular state of GBM as a more differentiated state enriched at recurrence and periphery of the tumor provides new insights into how neuronal activity regulates tumor invasion. Deconvolution of normal cell types from single-cell RNAseq and bulk tumors revealed that neuronal state of GBM is associated with high infiltration of non-malignant cells. The intricate synaptic communications between neurons and brain tumor cells are crucial for glioma progression and resistance to standard therapies, which is supported by ample data from the rapidly emerging field of “cancer neuroscience”. However, the molecular mechanisms driving enhanced neuronal activity at recurrence remains to be understood. Dissecting the spatiotemporal dynamics of glioma eco-system during evolution will be necessary to identify therapeutic vulnerability of recurrent GBM. Here, proteogenomics and single-nuclei RNAseq profiling of matched primary and recurrent GBM IDH wild-type suggest that the evolutionary transition from a more proliferative-progenitor towards the neuronal state in GBM is regulated by both genetic and post- genetic molecular events and potential functional interactions between malignant and non-malignant cells. In Aim 1, the development of a multiomics-based network diffusion approach will enlighten subnetworks of proteins/phospho-proteins significantly affected by upstream genetic events driving activation of neuronal programs during progression. Generation of in silico knock-out networks screen and integrative analysis of proteomics and pharmacological data of cancer cell lines will prioritize lethal and essential proteins in the subnetworks to identify potential therapeutic vulnerabilities in neuronal-recurrent GBM. In Aim 2, single-nuclei and spatial transcriptomics profiling of matched primary and recurrent GBM IDH wild-type will identify the functional connections between neurodevelopmental tumor cellular states and cell types in tumor microenvironment. The development of a spatial informed cell–cell communications algorithm and the reconstruction of intercellular signaling networks will infer the key functional interactions between neurodevelopmental tumor cellular states and cell types in tumor microenvironment along with the potential effect of these interactions on downstream regulatory molecular pathways. These studies lay the foundation for my future research program and will advance the understanding of the molecular mechanisms mediating glioma connectomes and driving glioma invasion. These studies will advance the neuroscience field through discovery of targetable pathways and proteins providing therapeutic opportunities for recurrent GBM.

NIH Research Projects · FY 2026 · 2024-02

The autogenous arteriovenous (A-V) fistula represents the most important lifeline of >600,000 Americans currently on hemodialysis. However, ~40 percent of newly created fistulas cannot be used for dialysis without a salvage procedure because stenosis (narrowing) prevents them from reaching the necessary blood flow. This problem represents significant comorbidity for the end-stage renal disease (ESRD) population and a substantial economic burden that currently surpasses $2 billion annually. Most troubling about this glaring statistic is that it has plateaued over the past five decades, highlighting a pressing need for revamped efforts to improve vascular access outcomes using novel and innovative therapeutic approaches. This translational proposal establishes a new modality to treat post-operative A-V fistula failure using an emerging anti-inflammatory concept. We have built this proposal upon the premise that a newly discovered agonist of Mac-1 integrin prevents stenosis in A-V fistulas by controlling post-operative inflammatory activity. Macrophages and other myeloid inflammatory cells control the underlying causes of stenosis, such as the development of post-operative intimal hyperplasia (IH) and fibrosis. Herein we hypothesize that pharmacological and genetic activation of the Mac-1 integrin is sufficient to control myeloid- derived inflammatory cell infiltration in the fistula without compromising access maturation. We also hypothesize that activation of the Mac-1 integrin attenuates the RAGE signaling in macrophages that leads to inward remodeling in newly created A-V fistulas. Our scientific premise is supported by published and preliminary data that demonstrate our experience with animal models to study fistula maturation biology. We will test our hypothesis in three specific aims and five experimental layouts to: 1) demonstrate that increased monocyte adhesion following Mac-1 activation protects experimental A-V fistulas from failure; 2) demonstrate the anti- inflammatory mechanism following Mac-1 activation in the fistula wall; and 3) demonstrate that Mac-1 activation attenuates post-operative inflammation, IH, and stenosis in preclinical A-V fistulas. We will combine fine microsurgical techniques and transgenic mice to achieve our goals successfully. In conclusion, successful completion of this proposal will pave the way to the design of new drugs using a novel mode of action to effectively target A-V fistula fibrosis and IH and reduce vascular access complications.

NIH Research Projects · FY 2025 · 2024-01

PROJECT SUMMARY/ABSTRACT: Over 700,000 Americans live with end-stage kidney disease (ESKD) and more than 1 million are projected for 2030. Ninety percent of these patients depend on a functional vascular access for hemodialysis (HD) treatments. However, the high rate of failure and complications of HD accesses is one of the most common causes of morbidity in this vulnerable population. Approximately 70% of new ESKD patients initiate HD with a central venous catheter (CVC), and most continue on a catheter for the next 6 months despite the high risk of life- threatening bloodstream infections. One of the factors contributing to prolonged CVC use is the high rate of arteriovenous fistula (AVF) maturation failure. Up to 50% of newly created AVFs fail to mature independently due to significant stenoses (narrowing) in the venous segment. Interestingly, prior CVC use is significantly associated with AVF maturation failure. In this proposal, I will challenge the high-risk, high-reward hypothesis that AVF maturation failure after CVC use has a preventable immunological basis. Namely, this discovery and mechanistic proposal aims to demonstrate the mechanistic link between CVC-derived molecular patterns, exaggerated activation of neutrophils, and increased vascular damage in newly created fistulas. This proposal is built on strong scientific premises, including the increased expression of at least 20 neutrophil enriched genes in human veins that failed compared to veins that remodeled successfully after AVF creation. I hypothesize that activation of the FPR1 receptor by catheter-derived bacterial peptides primes circulating neutrophils prior to AVF creation, increases vascular infiltration, and causes an overresponse to postoperative vascular injury that contributes to AVF failure. The proposal is organized in three complementary Specific Aims (SA). A discovery SA will look for CVC-related changes in peripheral neutrophils of patients undergoing surgery for AVF creation using single-cell RNA sequencing. A mechanistic SA will dissect the role of catheter derived N-formylated peptides in neutrophil activation, and the consequences of this inflammatory pathway for postoperative AVF remodeling using genetically modified animal models. Lastly, a pre-clinical translational SA will create a relevant animal model that combines CVC insertion, biofilm formation, and AVF creation, and test the efficacy of FPR1 activation blockers and neutrophil-depleting therapies to improve AVF remodeling. Considering the frequent use of catheters in HD patients, results from this grant may not only be significant for AVF maturation outcomes but also for other postsurgical scenarios such as transplant injury. From the career development point of view, this application responds to Funding Opportunity Announcement PAR-21-313, which aims at facilitating the transition of new investigators of diverse and underrepresented backgrounds such as myself to research independence. This proposal will support the development of a novel scientific niche in vascular access research and will generate state-of-the-art data for competitive R01 applications.

NIH Research Projects · FY 2026 · 2024-01

PROJECT SUMMARY ABSTRACT The exocrine pancreas of patients with type 1 diabetes is smaller than the pancreas of healthy subjects and shows histological anomalies such as fibrosis, fatty degeneration, inflammatory cell infiltration and atherosclerosis. The histological features associated with diabetes are so specific that they have been defined as a distinct entity called diabetic exocrine pancreatopathy, which can be readily distinguished from chronic pancreatitis. The prevalence is high: 35 to 77% of adult patients with type 1 diabetes present with pancreatic exocrine dysfunction. Multiple etiological factors have been proposed, but it is inescapable that type 1 diabetes is defined by the autoimmune attack destroying the beta cells. Our goals are (a) to understand how the beta cell coordinates pancreas function and (b) to determine how the progressive demise of the beta cell impacts this coordination. In preliminary studies, we found that local insulin signaling affects the function of the acinar tissue and the islet microvasculature. We thus propose that locally delivered insulin orchestrates pancreas function by acting on three functional effectors: (1) adjacent acinar tissues, (2) vascular units comprised of pericytes and endothelial cells, and (3) intrapancreatic neurons that provide local cholinergic input. We hypothesize that beta cell control of these effectors deteriorates during the progression of T1D, leading to inadequate coordination of pancreas activity. The long term loss of this coordination produces pathological sequelae across compartments. The rationale for the proposed research is that if we want to develop therapies it is imperative to elucidate the mutual interactions and mechanism that promote the disease state. The proposed research is therefore relevant to the mission of the NIH that pertains to the pursuit of fundamental knowledge about beta cell function and its demise in diabetes. Guided by strong preliminary data, our hypothesis will be tested by pursuing three specific aims (1) Determine the impact of insulin on exocrine tissue function during diabetes development, (2) Determine the impact of insulin on the pancreatic microvasculature, and (3) Determine how insulin impacts neural coordination of pancreas function. We will determine the trophic role of insulin by manipulating insulin signaling and measuring the structural and functional consequences in exocrine, vascular, and neural compartments. The manipulation will be performed in the mouse in vivo, using genetic tools or in diabetes models. We will study the healthy and the diseased state in the human pancreas by using living pancreas slices provided by the network of pancreatic organ donors. The proposed research is significant because it could generate mechanistic insight into how beta cells influence surrounding tissues to coordinate activity across pancreas compartments. By identifying the effectors and principles governing this local regulation, we will be able to understand how the progressive loss of beta cell function produces pathological changes across pancreas compartments in people with type 1 diabetes.

NIH Research Projects · FY 2026 · 2024-01

Project summary Eukaryotic protein kinases regulate important cellular processes through their ability to phosphorylate themselves and substrate proteins. One of the phosphorylation events common to most protein kinases is the phosphorylation that occurs at the activation loop. This phosphorylation event often occur via auto- phosphorylation although how an inactive kinase achieves the phospho-transfer reaction on its own activation loop site is still unclear [2]. We recently reported that activation of dual specificity tyrosine-phosphorylation- regulated kinases 1A and 1B (DYRK1A and DYRK1B) requires prolyl-hydroxylation by the oxygen sensing prolyl hydroxylase PHD1. DYRK1 activation by prolyl-hydroxylation instigates a sequence of events whereby phosphorylation of ID2 by DYRK1 releases ID2 mediated constraints on VHL ubiquitin ligase tumor suppressor complex, thus regulating the degradation of HIF proteins in brain tumors and cancer stem cells. Our most recent work identified prolyl hydroxylation by PHD1 as the general mechanism required in trans to prime protein kinases of the CMGC family for autophosphorylation and activation. Beside DYRK1, CMGC kinases includes Cyclin dependent kinases (C), Mitogen activated protein kinases (M), Glycogen synthase kinases (G) and CDC-like kinases (C). In this proposal we will follow the long-standing interest of the lab on the molecular pathways that favor neural cell self-renewal during brain development and are aberrantly recruited during gliomagenesis and investigate the mode of regulation of glycogen synthase kinase 3 (GSK3), a central hub in the control of brain functions and oncogenesis. GSK3 shares with other members of the CMGC kinase family the highly conserved CMGC insert domain. We found that this domain harbors a L/xGxP motif and the highly conserved proline residue, Pro-276, which is targeted by hydroxylation by the proline hydroxylase enzyme PHD1 and is necessary for kinase activation. These observations led us to propose a combination of mechanistic and genetic studies to define the dynamics of GSK3a/b kinase maturation induced by PHD1 and the interaction with other signaling mechanisms that regulate GSK3 kinase activity. The significance of proline hydroxylation for gliomagenesis will be investigated in knock-in mouse models of Pro to Ala mutation of GSK3a/b (Pro-339 and Pro-276, respectively). Mouse tumors will be analyzed using proteomics and phosphoproteomics to reconstruct the activity of wild type and mutant GSK3 kinases. Given the uncertain role of GSK3 kinases in cancer and glioblastoma (promoter or suppressor), findings will lead to a better understanding of the potential benefit/harm of GSK3 inhibitors that are currently proposed for the treatment of glioblastoma.

NIH Research Projects · FY 2026 · 2024-01

PROJECT SUMMARY The cellular plasticity of the human pancreas is implied from multiple scRNAseq studies. However, these have been invariably based on static datasets from which fate trajectories can only be inferred using pseudotemporal estimations. Furthermore, the reliance on isolated islet preparations for the conduct of these analyses has resulted in a drastic underrepresentation of other non-endocrine cell types, hindering our ability to accurately interrogate exocrine-endocrine interactions. The long-term culture of human pancreatic slices (HPSs) has presented the field with an opportunity to sidestep these limitations by longitudinally tracking tissue plasticity at the single- cell level. Combining single-cell transcriptomic datasets from same-donor HPSs at different time points, with or without a known regenerative stimulus (BMP signaling), has led to the integration of dynamic datasets that store true temporal or treatment-dependent information. This novel approach (Dynamic SliceSeq, or DSSeq for short) has revealed population shifts consistent with the BMP-mediated progenitor cell activation, the blurring of ductal/acinar boundaries, the formation of clear ducto-acinar-endocrine differentiation axes and, notably, the appearance of transitional insulin+ cell populations ‘caught in the act’ of adopting endocrine fates. Our research is the first to unveil human pancreatic plasticity at the single cell level as a function of treatment and time. In vitro lineage tracing indicates that BMP signaling also elicits the formation of functional glucose-responsive insulin+ cells within the exocrine compartment of slices from type 1 diabetic (t1D) donors. Our first hypothesis is that the path through which these cells arise in samples from non-diabetic donors will be largely preserved in those with autoimmune diabetes, and potentially even reinforced as a result of compensatory responses. We further hypothesize that the ductoacinar-endocrine differentiation axis identified by DSSeq (where progenitor populations of BMP-stimulated ductal cells differentiate into endocrine cells through an intermediate acinar-like stage) may mirror the process of embryonic ductal delamination. In particular, we expect BMP signaling to induce such progenitors to migrate into the acinar parenchyma prior to their coalescence into islets. To test these hypotheses, we will pursue the following specific aims: (1) DSSeq analysis of BMP-induced endocrine regeneration in HPSs from t1D donors; (2) Longitudinal resolution of ductal tissue remodeling and neogenesis of insulin+ cells within their native histological microenvironment; and (3) Spatial transcriptomics analysis of islet- duct interfaces in histological samples from nPOD’s t1D collection. The development of long-term HPS culture techniques, and especially DSSeq, has enabled the dissection of regenerative responses directly in live tissue from t1D donors with an unprecedented degree of resolution. The completion of our research objectives is expected to correlate the dynamic compartmental plasticity revealed by DSSeq with real-time histological changes induced by BMP signaling in HPSs from diabetic donors. Beyond the significance of these findings from a basic science perspective, our research will have a direct impact on the design of therapeutic approaches to regenerate β-cells in diabetic patients.

NIH Research Projects · FY 2025 · 2024-01

SUMMARY Esophageal adenocarcinoma (EAC) is the most prevalent histological type of esophageal malignancy in the US and many Western nations. This tumor remains deadly as approximately 80% of patients are diagnosed at advanced stages and have a low five-year survival rate. Gastroesophageal reflux disease (GERD) is one of the strongest risk factors for EAC. In GERD patients, the epithelial lining of the esophagus is exposed to the gastroesophageal reflux (GER) that contains gastric acid frequently mixed with duodenal bile. The esophageal epithelial cells undergoes severe damage from exposure to acid and bile salts. This exposure also promotes inflammation, which can exacerbate tissue damage and lead to the development of Barrett's esophagus (BE). BE is a preneoplastic condition that is disposed to malignant transformation. The molecular mechanisms of esophageal tumorigenesis in conditions of esophageal reflux injury remain poorly understood. We have developed an innovative hypothesis to investigate how isolevuglandin (isoLG) lipid derivatives that adduct multiple proteins in conditions of esophageal reflux facilitate tumorigenic processes by protein adduction. IsoLGs are formed from the free radicals induced peroxidation of lipids and cyclooxygenase (COX) and are highly reactive for lysine as well as other cellular amines. IsoLGs bind covalently with the protein molecules to inflict damage before being recognized by cellular defense mechanisms. In our experimental conditions of esophageal reflux, p63 is found to be one of the most adducted proteins by isoLGs. P63 is a master regulator of esophageal epithelial development, which also regulate a broad spectrum of genes involved in different cellular processes such as DNA repair, stemness, proliferation and differentiation. Our preliminary data strongly support the hypothesis by providing evidence of the alteration of p63 protein by adduction. In aim 1, using in vitro cell systems, this proposal will examine the unique mechanisms regulating p63 signaling pathway by protein adduction and its biological impact in conditions of esophageal reflux injury. In aim 2, we will study the p63 protein adduction in in vivo mice model and test various pharmacological options to reverse this process. If successful, this study will provide a new therapeutic approach to prevent the pro-tumorigenic alterations of esophageal cancer.

NIH Research Projects · FY 2025 · 2023-12

Abstract Critical limb ischemia (CLI) is the end stage of peripheral artery disease (PAD) and can be an underlying cause of ischemic rest pain, gangrene, and amputation. Primary amputation is often required for the 30% of CLI patients who are not eligible for limb revascularization; thus, an effective therapy to improve neovascularization is urgently needed. During hindlimb ischemia, monocytes are among the first cells to hone in on the ischemia site and contribute to neovascularization. Our recent publication revealed that ischemia training, performed by 24 hours of unilateral femoral artery ligation, led to functional reprogramming of bone marrow-derived monocytes (BM-Mono), enabling them to protect against the outcomes of limb ischemia by increasing perfusion and neovascularization. Mechanistically, this reprogramming resulted in the downregulation of 24-Dehydrocholesterol Reductase (Dhcr24, an important enzyme that converts desmosterol into cholesterol), and a consequent accumulation of desmosterol in those cells. Interestingly, our preliminary data have shown that ischemic-trained monocytes control vascular proliferation, and both vascular cell types, endothelial cells (ECs) and smooth muscle cells (SMCs), have increased expression of IL- 10 and its receptors. Moreover, the macrophages differentiated from the ischemic-trained monocytes possess an anti-inflammatory phenotype. Here our primary scientific goal is to target Dhcr24 in monocytes as a novel and unique strategy to improve neovascularization in the setting of PAD/CLI. First, we will assess whether downregulation of Dhcr24 in BM-Mono improves hindlimb ischemia outcomes, such as perfusion and neovascularization; and second, we will identify the mechanisms by which those BM-Mono with low Dhcr24 expression control vascular proliferation. Our long-term goal is to translate these findings using a Dhcr24 inhibitor into a new therapeutic approach to treat PAD/CLI. We hypothesize that the low expression of Dhcr24 in BM-Mono regulates anti-inflammatory pathways in vascular cells that controls their proliferation, leading to a proper neovascularization in the ischemic limb. We will test our hypothesis in two specific aims. SA1: Determine whether monocyte Dhcr24 regulation alters hindlimb ischemia outcomes. We will determine the dependence of loss- or gain-of-function of monocyte Dhcr24 on limb perfusion, function, and neovascularization using a pre-clinical model of hindlimb ischemia. We expect to demonstrate that low expression of Dhcr24 in BM-Mono plays a beneficial role against hindlimb ischemia. SA2: Identify the mechanism by which monocytes with low Dhcr24 control vascular proliferation We will dissect the underlying mechanisms by which monocytes with low Dhcr24 expression regulate vascular proliferation. We will also inhibit Dhcr24 in human monocytes and assess their inflammatory phenotype. We expect to find that the underlying mechanisms are based on anti-inflammatory pathways.

NIH Research Projects · FY 2025 · 2023-09

The burden of hepatocellular carcinoma (HCC) due to non-alcoholic fatty liver disease (NAFLD) has increased substantially over the past two decades. Although both NAFLD and HCC disproportionately affect Latino individuals, few Latinos were included in genetic and epidemiologic studies evaluating HCC risk. Prior genetic studies examining NAFLD risk that did include Latinos were limited in that they used candidate-gene approaches which cannot detect novel genetic associations. Other studies limited inclusion to individuals from one country, and thus could not consider how the incredible diversity among Latinos might drive genetic risk or differences in NAFLD phenotype, risk of cirrhosis, or HCC risk. In the proposed study, Drs. Jones and Flores will work collaboratively as multiple principal investigators. Along with co-investigators, they will collaborate with other members of the Liver Cirrhosis Network (LCN) to develop precise, personalized approaches to NAFLD prognostication and HCC risk stratification targeted specifically to Latinos, a vulnerable population with excess disease burden. In Aim 1, Drs. Flores and Jones will leverage existing data from two studies (UCLA ATLAS Community Health Initiative and NIH All of Us Research Program) to conduct a genome wide association study (GWAS), phenome-wide association study, and create polygenic risk scores in persons with NAFLD. All of Us and ATLAS aim to enroll diverse participants to ensure inclusivity and generalizability in Precision Medicine research. As such, the proposed study represents the largest, most racially diverse GWAS in NAFLD with 21,199 individuals with NAFLD already identified. We will define the relationship between known and novel single nucleotide polypmorphisms (SNPs) and risk of NAFLD, NASH, NAFLD-cirrhosis, and NAFLD-HCC, stratified by race, region of origin and genetic ancestry. In collaboration with LCN investigators, Drs. Flores and Jones will enroll Latino participants with NAFLD into a prospective case-control study that aims to characterize gene- environment interactions between known and novel genetic NAFLD-associated SNPs and environmental risk factors including HIV, diabetes, and metabolic syndrome. They will engage new and existing LCN Cohort participants as well as participants enrolled in existing cohorts at the University of Miami (UM), the University of Los Angeles California (UCLA) and the University of Puerto Rico Comprehensive Cancer Center (UPRCCC). All sites will identify and recruit new cases with NAFLD and healthy controls. We will develop polygenic risk scores that incorporate genetic, clinical, sociocultural, behavioral, and environmental characteristics to predict (1) risk of cirrhosis in persons with NAFLD, (2) hepatic decompensation in NAFLD patients with cirrhosis, and (3) HCC risk in NAFLD patients with or without cirrhosis. NAFLD is the fastest growing cause of cirrhosis in the US and disproportionately impacts Latinos who also have the highest HCC burden. By identifying the strongest risk factors that drive differences in NAFLD phenotype, cirrhosis decompensation, and HCC risk in a large, diverse Latino sample, we can identify those at greatest risk and intervene on modifiable risk factors.

NIH Research Projects · FY 2025 · 2023-09

Abstract: More than 40,000 women die each year of metastatic breast cancer. The majority of these tumors are hormone receptor positive (HR+) that are treated with a cyclin dependent kinase 4/6 (CDK4/6) inhibitor in combination with an aromatase inhibitor or fulvestrant. With these treatments, women with advanced breast cancer are living longer, but treatment related toxicities inevitably occur, and quality of life is limited by side effects of cancer treatment which may result in dose reductions and delays. Fatigue is the most commonly cited adverse side effect reported for women taking CDK4/6 inhibitors and mechanistically may be associated with inflammation. When unmanaged, fatigue is debilitating directly impacting both psychological and physical quality of life and a key driver in discontinuation of therapy. Lifestyle interventions targeting diet and exercise have evidence for improving fatigue in early stage breast cancer, however whether these strategies are efficacious for improving outcomes in women with advanced breast cancer remains unknown. Research regarding the impact of these strategies on high grade fatigue induced by CDK4/6 inhibitors in advanced disease from a representative patient population are needed. The prolonged overnight Fasting and/or Exercise on fatigue and other patient reported outcomes in women with hormone Receptor positive advanced breast cancer (FastER) study, will evaluate a phase II, 2 x 2 randomized controlled trial testing the effects of a prolonged overnight fasting (POF) intervention alone, moderate-intensity exercise alone, or in combination, on fatigue in 260 women with advanced breast cancer initiating treatment with hormonal therapy in combination with a CDK4/6 inhibitor. Participants would undergo assessment of fatigue and associated inflammatory biomarkers, as well as assessment of physical activity, diet, physical function, body composition and patient reported outcomes at baseline (prior to the initiation of CDK4/6 inhibitor) and 12 weeks (post-intervention), 6 and 12 months after study enrollment. The primary outcome of the study is to evaluate the impact of the interventions (vs control) on fatigue in women at 12 weeks. Secondary outcomes include inflammatory biomarkers, patient-reported outcomes, physical function and body composition. The FastER study will also explore the impact of the intervention on circadian rhythms, fatigue, sleep, depression, anxiety, quality of life, physical function and body composition. intensity exercise alone or in combination can mitigate the adverse consequences of treatment and improve fatigue and other outcomes, in women with advanced breast cancer treated with CDK4/6 inhibitors to ultimately improve both quantity and quality of life for this growing population of women.

NIH Research Projects · FY 2025 · 2023-09

Breast cancer patients represent fast-growing medical cannabis and cannabinoid (MCC) users in this country; a recent survey indicates that about 42% of breast cancer patients use MCC to alleviate treatment-related symptoms, and many of these patients do not discuss their use with their oncologists. Despite being considered safe and well-tolerated, MCC may result in potential interactions with cancer treatments, adverse reactions, and tumor progression. Unfortunately, despite the increase in MCC use, research on its health effects, including the potential benefits and harms, remains limited. Therefore, we propose a prospective cohort study of breast cancer patients to address gaps in knowledge and build evidence of the types of products in use, frequency, dosage of use, and the benefits and harms of MCC use during and after cancer treatment. Our central hypothesis is that MCC may improve treatment-related symptoms and clinical outcomes in some patients by targeting the inflammasome/inflammatory pathway. Therapeutic strategies targeting inflammasome may prevent treatment-related symptoms and improve clinical outcomes. We will test a new paradigm that the inhibition of inflammasome-mediated inflammatory responses by MCC plays a role in its biological effects. We plan to enroll 600 breast cancer patients (300 MCC users and 300 non-MCC users) and collect data on patient characteristics, treatment plans, clinician-reported outcomes, adverse reactions, and patient-reported outcomes. We will monitor MCC use through in-person visits and technology-based assessments such as smartphone and sensor-based measurement bursts. We will also collect blood samples for MCC quantitation/characterization and inflammasome/inflammatory biomarkers. Aim 1 will evaluate the benefits and harms of MCC in breast cancer patients during and after treatment. Aim 2 will elucidate whether inflammasome/inflammatory biomarkers mediate the benefits and harms of MCC in breast cancer therapies and other medications used. Aim 3 will investigate how MCC product characteristics impact benefits and harms (Aim 1) and inflammasome and inflammatory biomarkers (Aim 2). Leveraging our promising preliminary data, state-of-the-art high-throughput technologies, and a multidisciplinary research team with complementary expertise, we are well-positioned to conduct this study and address the scientific knowledge gaps surrounding the benefits and harms of MCC in breast cancer patients. We will provide scientific evidence on the impact of MCC on breast cancer patients during and after treatments, the biological effects of MCC during treatments, potential interactions between MCC and cancer therapies and other medications, and the types and patterns of MCC use that present the best benefits/harms ratio. Our results will be of great value to physicians and cancer patients, as they will inform decision-making regarding MCC to enhance therapies, improve quality of life, and minimize adverse effects. If funded, we will work closely with the coordinating center and NCI in sharing assessment measures and disseminating study findings.

NIH Research Projects · FY 2025 · 2023-09

Abstract Itch is a global health problem, affecting tens of millions of people. The brain plays a crucial role in itch perception; thus the specific circuit for itch processing in the brain has the potential to become a therapeutic target for itch in a wide range of pruritic diseases. Our understanding about the brain mechanism of itch has advanced in the past decades. Previous studies identified a key brain circuit related to itch, which includes the ascending pathway projecting from the thalamus to the posterior insular cortex (pIC) and networks originating from the pIC. In addition, dopamine was found to be a key neurotransmitter associated with itch. These achievements have also identified two important topics that must be understood to develop effective itch treatment targeting the brain. First, the brain circuit of itch is similar to that of pain, though itch and pain are clearly distinct sensations. An itch-specific ascending pathway and network have not yet been identified within the circuit (an itch-specific circuit). Second, there could be dopaminergic gene polymorphisms that determine individual differences in itch perception. If such polymorphisms are identified, brain imaging studies focusing on these genes can identify key loci regulating itch perception within the itch-specific circuit. This line of work will eventually enable us to develop brain-based, tailored itch treatment. To this end, it is crucial to identify an itch- specific circuit and dopaminergic gene polymorphisms influencing individual differences in itch perception. Our research proposal will address this significant gap. Aims 1 and 2 will use fMRI to determine an itch-specific ascending pathway and network by comparing brain activity between itch and pain stimuli. Aim 3 will use our novel method for quantitatively assessing genetic impacts on itch to identify dopaminergic gene polymorphisms that influence individual differences in itch perception. Success of our project will eventually lead to the development of brain-based tailored itch treatment, which will advance the treatment of chronic itch regardless of the underlying etiology.

NIH Research Projects · FY 2023 · 2023-09

Abstract Interstitial cystitis/ bladder pain syndrome (IC/BPS) is one of the most debilitating chronic pelvic pain (CPP) conditions that negatively impacts the quality of life and sexual activities in 2.7% to 6.5% of women in the US. Pelvic floor muscle (PFM) overactivity, characterized by an increase in the tonic muscle activity, is a condition related to myofascial pain that presents in the majority of CPP conditions, including up to 85% of women with IC/BPS. However, pelvic floor pain is intrinsically a multifactorial dysfunction that is attributed to postural issues, myofascial trigger points, and abnormal muscle tone. Myofascial therapy, including specific pelvic floor muscle soft tissue mobilization and muscle stretching, is standard treatment for patients with IC/BPS and concomitant PFM tenderness. Unfortunately, even among IC/BPS patients with PFM tenderness on exam, only 59% of patients report symptom improvement after myofascial therapy. Pelvic floor myofascial therapy, however, does not address movement impairments of the trunk and hips, which are also associated with pelvic pain. A pelvic floor muscle phenotyping framework would allow IC/BPS patients to be categorized to facilitate individualized treatment. Unfortunately, no technology is currently available for quantitatively and objectively assessing PFM etiologic factors associated with IC/BPS, which, otherwise, would advance the understanding of the underlying mechanisms and allow for phenotyping patients for appropriate intervention. Our team has successfully 1) developed a novel intra-vaginal high-density surface electromyography (HD-sEMG) technique to reliably and quantitatively assess PFM overactivity in women with IC/BPS and 2) developed a novel muscle network analysis technique to reveal, for the first time, the inter-muscular connectivity pattern alterations among patients with neuromuscular conditions, and 3) demonstrated the feasibility to cluster patients with IC/BPS into phenotypic subgroups, depending on the underlying mechanism. This study aims to comprehensively assess the PFM overactivity, hip/trunk muscle activity alteration, PFM-to-Hip/ Trunk inter-muscular connectivity, and distinct PFM phenotypic subtypes in IC/BPS. This research represents the first effort to comprehensively assess the PFM overactivity, hip/trunk muscle activity alteration, PFM-to-Hip/Trunk inter-muscular connectivity, and distinct PFM phenotypic subtypes in IC/BPS. The integration of these multifactorial assessments will advance our understanding of the multifactorial pathology of IC/BPS. The quantification of relative importance of these pathological contributors will allow for IC/BPS patient phenotyping. Identification of PFM phenotypic subtypes may facilitate personalized physical therapy treatments.

NIH Research Projects · FY 2025 · 2023-09

PROJECT SUMMARY ABSTRACT: The broad goal of the University of Miami's Advanced Workshops and Coaching Network (AWACN) is to enhance the stage- specific career development skills of newly funded NIDDK researchers at the postdoctoral and junior faculty levels. Continued development of professional skills will foster early career success and lay the foundation for a strong and rewarding career in academic research. Program scholars will participate in a 3-day workshop hosted by the University of Miami's Miller School of Medicine. Workshop curricula will be determined by pre-workshop needs assessment surveys allowing each workshop to target needs of the scholar cohort and avoid overlap with strong programs at the scholars' home institutions. Workshops will include didactic instruction, experiential activities and provide scholars with tangible outputs. All workshops will be led by NIDDK-funded researchers at the University of Miami, or faculty with expertise in transferable skills development. The latter will include effective negotiation tactics, strategies to combat microaggression, and work/life balance. During the workshop each scholar will also meet their two coaches, who, over the next two years, will provide guidance and support to the scholar. Coaches are all University of Miami faculty with current research funded by NIDDK and who are trained mentors. One coach will match the research type of the scholar (basic science, translational, or clinical research) and the other coach will provide a broader perspective (e.g. a translational scientist will bring experience in moving basic science discoveries into potential therapies). Coaches will meet remotely with their scholars approximately once a month by phone or video conference to provide guidance on topics of the scholar's choice. Coaches will also strongly promote the scholar's engagement in productive, substantive collaborations that lead to joint publications that then support their recognition in their field and promote their career progression. The effectiveness of the workshops (Aim 1) and extended coaching (Aim 2) will be evaluated using a outcome logic model. This model will assess the effectiveness of the program components and resources on the scholars' career progression and satisfaction. The evaluation process will also address whether an external workshop program and/or coaching network can enhance career development

NIH Research Projects · FY 2025 · 2023-09

PROJECT SUMMARY Glioblastoma is the most common primary brain cancer worldwide. Novel treatment strategies are urgently needed since glioblastoma is nearly universally fatal with a median overall survival of only 1.5- 2 years. A frustrating aspect of glioblastoma is that approximately half of all patients will have what looks to be tumor growth on their post-treatment MRI, termed progression. Although, half of patients with progression will turn out to have pseudoprogression, which is a not-fully understood phenomenon believed to be edema and inflammation caused by the immune system and represents a good response to treatment. In fact, patients with pseudoprogression tend to do better than the general glioblastoma population and have a median overall survival of up to 3 years. On the other hand, patients with true progression of disease (tumor growth and poor/nonresponse to treatment) tend to do worse than the general glioblastoma population and have a medial overall survival of only 10 months. The frustrating part for clinical team, and the patients themselves, is that true progression and pseudoprogression are not discernable from one another during treatment, or even on initial post-treatment imaging (1-month post-treatment). Instead, the current gold-standard to distinguish between true and pseudoprogression is to “watch and wait” – continue monitoring with serial imaging and see if the patient clinically worsens or stabilizes. Thus, there is an unmet need for techniques that reliably and accurately determine if tumor growth/progression is occurring during treatment and predict/determine which sub-type of progression (true progression or pseudoprogression) a patient has. My laboratory focuses on responding to this unmet need through a variety of methods: serial multiparametric MRI (anatomic, perfusion, diffusion, spectroscopic, etc.), quantitative MRI analysis, machine learning, and molecular research including analyzing blood samples of glioblastoma patients to look for circulating tumor cells and other molecular markers. This proposal focuses on auto-detection of tumors on MRI based on machine learning (Aim 1) and analysis of anatomic and physiologic changes (Aim 2) from daily multiparametric MRI to address this issue by creating techniques that can detect enlarging tumors during treatment and predict between true and pseudoprogression months earlier than current methods. The goal of this proposal is to develop tools that identify and monitor patients with significant anatomic and/or physiologic tumor changes much earlier than current methods, so that in the future, prompt, aggressive, and early therapy adaption can be implemented. This project will translate directly to the practice of clinical medicine and advance the field of glioblastoma treatment. Additionally, it will allow me to gain hands-on skills and expertise in machine learning, radiomics, MRI, neuroimaging, neuro-anatomy, radiation therapy, and oncology, and aid in preparing me for a career as an academic physician scientist in the field of radiation oncology.

NIH Research Projects · FY 2025 · 2023-09

The goal of the University of Miami CTSI K12 Career Development Program is to prepare 10 scholars (MD or PhD) for independent and sustained careers in clinic.al translational science and research (CTSR). Integrated with the Miami CTSI, the program will develop a robust, broad-based CTSR workforce of scientists who possess both deep scientific domain expertise and broader research systems understanding. Driven by this goal, we have set three objectives 1) develop core competences and knowledge in CTSR; 2) provide evidence informed mentoring and 3) accelerate transition to independent and sustained careers in CTSR. Our program is designed to ensure that a pool of highly trained CTSR scientists can address the needs of South Florida and beyond. Following the NCATS vision of the ideal translational scientist the program's approach to CTSR training is global. where scholars produce discoveries that are simultaneously important for their disciplines, contribute to other disciplines~ and of benefit to patients and communities, advancing the translational process as a whole. Our program's conceptual approach integrates principles of team science and dissemination & implementation to help scholars successfully move their research products into real-world use relevant to public health. Given our track record of ten years with a successful CTSI KL2 program and a strong institutional commitment to faculty development and a supportive research environment, we offer unique training opportunities for addressing complex CTSR challenges. The program is organized around five. major activities: (1) recruitment and selection of highly qualified candidates; (2) developing key competencies in CTSR; (3) collaborate mentorship by CTSR dou1ain experts, and clinician, peer and community mentors; (4) accelerating scholar transition to an independent and sustained career in CTSR; and (5) program evaluation. In the five years of the Kl2 award cycle, the program will provide up to two years of funding to each scholar for personalized training. intense scientific practice, and research career development experience under the guidance of collaborative mentorships and partnerships outside of their areas of expertise, both within their research domains and other relevant stakeholders. Our rigorous evaluation plan assesses the. quality and effectiveness of the training program and scholar capacity to effectively lead, communicate, collaborate. and break down barriers across multidisciplinary teams and the translational process. The program is led by two directors who are translational MD/PhD researchers with complementary laboratory-to-clinical and clinical-to population health research. and guidance committees composed of experienced scientists and training program leaders from our Hub and other CTSAs.

NIH Research Projects · FY 2025 · 2023-09

Project Summary Retinitis pigmentosa (RP) and related inherited photoreceptor dystrophies are characterized by progressive photoreceptor loss, resulting in poor vision or even blindness. While these disorders are caused by the mutations of various genes, mutations in RHO, USH2A, PRPH2, RP1, CNGB1, EYS, PDE6A, PDE6G, PDE6C, PDE6H, GNAT1, and NR2E3 account for a substantial number of cases. We recently performed a genome-wide DNA methylation analysis of human and murine fetal retinas (which mostly contain retinal progenitor cells [RPCs]), postnatal murine RPCs, and mature photoreceptors. We discovered that the promoters of all of the genes above were highly methylated (hypermethylated) in DNA isolated from fetal retinas and RPCs. The methylation of these promoters was significantly reduced during RPC differentiation into photoreceptors and accompanied by an increased expression of the corresponding genes. It is generally accepted that DNA methylation in promoter regions silences gene expression, while DNA demethylation should occur to allow gene expression. Unsuccessful demethylation of the promoters of the genes above during RPC differentiation into photoreceptors may reduce or even eliminate their activity, leading to photoreceptor dystrophies without any mutations in the genomic DNA. Thus, not only mutations in DNA but also retina- specific epigenetic changes in the DNA may contribute to the pathogenesis of RP and related diseases, indicating the importance of understanding the DNA demethylation pathway during photoreceptor development. The ten–eleven translocation (TET) protein family has a vital role in DNA demethylation and regulates eye development and neurogenesis in various species. Our data and the results of other laboratories indicate that the TET-dependent DNA demethylation pathway controls photoreceptor development. The objectives of this project are to gain a detailed understanding of how the TET-driven DNA demethylation pathway specifies the differentiation of RPCs into photoreceptors, and to investigate how irregularities in its activity lead to photoreceptor death and retinal degeneration. Using a rigorous experimental design, we will explore this pathway in accordance to our specific aims: 1) determine whether the TET-dependent DNA demethylation pathway acts as a “vertical” epigenetic “switch” between progenitor and photoreceptor precursor fates in the developing retina; 2) determine whether the TET-dependent DNA demethylation pathway functions as a “horizontal” epigenetic “switch” between rod and cone photoreceptor phenotypes; 3) determine whether TET enzymes require transcription factors with DNA binding domains acting as TET binding partners to specify target genes for demethylation and activation during photoreceptor development. To reach these objectives, we will employ animal models and a wide range of biochemical, molecular, and epigenetic approaches, striving to obtain robust and unbiased results. Upon the completion of the project, we expect the results to reveal the role of epigenetic mechanisms in photoreceptor normal and pathological development.

NIH Research Projects · FY 2025 · 2023-08

Located in South Florida (SoFL), the goal of University of Miami’s Clinical and Translational Science Institute (CTSI) are to improve health outcomes in South Florida (SoFL) and beyond. The CTSI accomplishes these goals by catalyzing the development, demonstration, and dissemination of scientific and operational innovations that improve the efficiency, effectiveness, and quality of clinical and translational research. This is done through bidirectional engagement of patient advocates and community collaborators, to help identify gaps, challenges, and chokepoints in the translational research process. In response, the CTSI develops and test stakeholderdriven resources, tools, and interventions ultimately promoting the adoption and integration of those that are demonstrated to be successful, through dissemination and implementation efforts. This work is shaped by two primary factors: 1) SoFL embodies the evolving face of our nation, which enables CTSI Hub investigators to explore research questions difficult to conceptualize and evaluate elsewhere; and, 2) CTSI has a strong, successful and sustained record of authentic community and stakeholder engagement which facilitates multidisciplinary research that is grounded in the lived realities of the people that it is intended to serve. The CTSI’s focus, scope of activities and aims are grounded in an understanding of these factors and their complex interplay with one another. Together with community and patient stakeholders, the CTSI will: 1) support the development and dissemination of innovative resources and services to increase the quality, efficiency, and effectiveness of research across the entire translational research spectrum; 2) promote research collaborations aimed at facilitating and accelerating research to improve community health; 3) develop innovative training programs to support a team science-oriented CTS workforce, from translational scientists to clinical research professionals and key stakeholders; and 4) support CTS research responsive to these overarching goals. Through these efforts, the CTSI expects to achieve significant improvements in the quality, safety, efficiency, effectiveness, and informativeness of its Clinical and Translational Enterprise. The CTSI will then broadly disseminate its acquired knowledge regionally and nationally to maximize impact and opportunity, both now and in the future. By doing so, the CTSI accelerates bringing the benefits of translational science (more effective treatments, drugs, devices, behavioral interventions, and medical procedures) to all people.

NIH Research Projects · FY 2025 · 2023-08

Project Summary The focus of research in the Taylor laboratory is on nuclear export in eukaryotic cells, which we study by perturbing the normal function of the main exporter of proteins, XPO1 or Exportin-1. We use molecular biology, genomics, proteomics, and mouse modeling to determine the mechanisms of XPO1-mediated nuclear export function and the role of XPO1 in disease pathogenesis. We also study how XPO1 interacts with other proteins and molecules such as ribonucleic acid (RNA). In the next five years, the goal of the lab is to define the role of XPO1 in the nuclear export of RNA. The laboratory is currently pursuing the following projects. Defining the molecular effects of wildtype and mutant XPO1 on gene expression, splicing, and translation. Given the known role of XPO1 in the export of small nuclear RNA (snRNA) and ribosomal RNA (rRNA) via the binding of RNA binding proteins, we hypothesize that alterations in XPO1 may impact RNA splicing and/or mRNA translation. We have generated several genetically engineered models to allow endogenous expression of the XPO1 E571K or R749Q mutation, including conditional knockin mice. We have demonstrated through a variety of proteomic, biochemical, structural, and molecular studies that XPO1 E571K affects recognition of its cargo’s nuclear export signal (NES) and results specifically in altered export of of NFκB and NFAT transcription factor proteins. The aim of my first project is to continue to study the extent of how the XPO1 E571K alters nuclear export and to explore whether RNA export, and consequent mRNA splicing and translation, is affected through the mislocalization of RNA binding proteins. We have recently discovered another mutation, XPO1 R749Q, that affects nuclear export by increasing the export of proteins out of the nucleus. We plan to study which proteins this affects and how this affects cell growth, cell cycle and other homeostatic processes. We are also modeling overexpression of XPO1 and will compare how this overactive mutant compares to overexpression in relation to protein and RNA export. Drugs that selectively inhibit XPO1 will allow us to isolate the effects of XPO1- dependent export. Furthermore, our preliminary data show that cells with abnormal splicing due to SF3B1 mutations undergo apoptosis upon exposure to XPO1 inhibitors. We will also investigate the effects of XPO1 inhibition in mutant Sf3b1 expressing cells using genetically engineered cell lines and an Sf3b1 knockin mouse model. We hypothesis that XPO1 inhibition perturbs RNA export and affects gene expression and mRNA splicing given the known role of XPO1 to export small nuclear RNA via RNA-binding proteins. This work will serve as foundational data for future grant submissions investigating the biology of XPO1 and its role in nuclear export of RNA. The overarching goal of this research is aligned with the NIGMS’ mission to support new basic discovery science that can eventually culminate in new medical therapies. By better understanding the role of XPO1 in RNA export, we can better design therapies that modulate nuclear export or RNA splicing for a wide range of diseases where RNA regulation may play a role.

NIH Research Projects · FY 2025 · 2023-08

Alcohol-associated liver disease (ALD), which includes alcohol-associated cirrhosis (AAC) and alcohol- associated hepatitis (AH), is now the leading indication for liver transplant (LT) in the US. Early LT (eLT), defined as LT evaluation with <6 months of alcohol abstinence, is associated with acceptable outcomes for AH in retrospective studies. However, prospective, multi-center data including biopsychosocial factors on eLT for all advanced ALD are lacking. It is known that alcohol cessation is the most important factor influencing survival in ALD, and integrated alcohol use disorder (AUD)/ALD care is critical to help patients achieve abstinence, yet the degree of care integration and how this influences post-LT outcomes has not been systematically studied. Knowledge gaps in eLT for ALD include: a) limited data on who gets referred for eLT and referral barriers; b) lack of standardized biopsychosocial measures and outcomes; and c) minimal stakeholder involvement beyond LT providers. There is an urgent need to (1) define factors influencing eLT referral, (2) develop risk prediction models of key patient-centered outcomes, (3) incorporate validated biopsychosocial measures into models, and (4) evaluate the impact of integrated care on outcomes following eLT. For example, The INTEGRATE collaborative, comprised of multidisciplinary clinicians and researchers from the University of Texas Southwestern Medical Center, University of Michigan, University of Miami, and Columbia University-Weill Cornell Medicine, is ideally positioned to address these urgent research needs. Collectively, we have developed a distinctive investigator team with clinical and methodological expertise in ALD, AUD, LT, behavioral research, risk modeling, data harmonization, health disparities, causal inference, and mixed-methods research, and (4) documented track record of NIH funding in LT access, organ allocation, LT outcomes and healthcare disparities, and NIAAA funding in ALD/AUD. Our large volume transplant centers with established protocols for eLT for ALD will facilitate the following aims: 1) characterize and develop risk prediction models for transplant-free survival among those with limited access to LT to define those in greatest need of eLT referral and listing; 2) evaluate barriers and facilitators to referral for eLT in ALD; 3) apply causal inference approaches to observational data to evaluate biopsychosocial factors and develop risk models predictive of outcomes at key timepoints in eLT for ALD; 4) define stakeholder perceptions and preferences for selection and outcomes in eLT for ALD; and 5) evaluate how integrated care processes influence outcomes in eLT for ALD. At the conclusion of this work, we will have collaboratively: (1) defined factors for referral and waitlisting for eLT in ALD (selection), (2) identified which biopsychosocial factors are causally related and predictive of outcomes most important to stakeholders (outcomes) and (3) determined how integrated care influences stakeholder-relevant outcomes in eLT for ALD (management).

NIH Research Projects · FY 2025 · 2023-08

Amyloid-bodies and the Evolution of Malignancies Project Summary The ability of cancer cells to adapt to a wide variety of stress conditions plays a critical role in various physiological facets of tumorigenesis. We recently reported the discovery of stress-induced low complexity noncoding RNA derived from stimuli-specific loci of the ribosomal intergenic spacer (rIGSRNA); an enigmatic region of the human genome historically dismissed as “junk” DNA. We showed that low complexity rIGSRNA activate a physiological amyloidogenic program that converts nucleoli into Amyloid-bodies: reversible nuclear membrane-less compartments composed of immobilized proteins in an amyloid-like state. While many cellular bodies have been described as liquid-like (e.g., stress granules, P-bodies, germ cell granules), the discovery of Amyloid-bodies provided evidence of an amyloidogenic program that can physiologically transition biological matter to a solid state. Amyloid-bodies are found in sub-populations of cells in normal tissues, the core of low- grade human tumors and cells responding to various stimuli highlighting their ubiquitous nature. Proteomic analysis revealed that Amyloid-bodies immobilize participants of the DNA synthesis machinery and cell cycle control, amongst many other metabolic regulators. Intriguingly, Amyloid-bodies share many biophysical properties with the amyloidogenic, solid-like Balbiani-bodies involved in metabolic suppression in Xenopus. Likewise, yeast solidify elements of their proteome to sporulate and arrest growth in non-permissive conditions. This raises the fascinating possibility that stressed cancer cells assemble Amyloid-bodies to enter a spore-like state of extreme metabolic depression. In this grant proposal, we will show preliminary data that low complexity rIGSRNA coordinate unusual RNA tailing programs to drive system-wide amyloidogenic phase transition. This post-translational pathway enables cancer cells to immobilize elements of the DNA synthesis machinery and halt oncogenic signaling in an adaptive response to severe environmental insults. Based on these preliminary and published results, we hypothesize that “Nucleolar phase transition programs temporarily suspend oncogenicity”. We plan to test this hypothesis by: 1- Uncovering mechanisms of physiological phase transition; 2- Examining how low complexity rIGSRNA activate RNA tailing programs; 3- Demonstrating a role for RNA tailing-mediated phase transition in tumorigenesis. The discovery of dedicated enzymatic programs that drive physiological amyloidogenesis provides a unique opportunity to study the role of liquid-to-solid phase transition in human clinical samples and in vivo tumor assays. By studying clinical samples, in culture and orthotopic animal models, we will test if phase transition induces a unique and yet uncharacterized cancer cell state of extreme metabolic depression, while highlighting biochemical functions for low complexity RNA typically discarded as useless nucleic acids.

NIH Research Projects · FY 2025 · 2023-08

The goal of this application is to develop a research training program in the US Virgin Islands (USVI) to inspire the next generation of USVI students to pursue research careers in cardiovascular disease (CVD) and related chronic diseases. We propose an eight-week intensive summer research institute facilitated through a partnership between the University of Miami Miller School of Medicine (UMMSM), the Virgin Islands Department of Health (VIDOH), and University of the Virgin Islands (UVI). Designated by HRSA as a high-priority health professional shortage area, USVI is a U.S. Territory in which 80% of residents report having at least one major CVD risk factor.  VIDOH is the leading provider of health care in the region but faces numerous challenges including a health care workforce shortage. Over 30% of federally funded VIDOH positions are vacant and 42% of funded public health positions are unfilled. Lack of data, limited research capacity, and well-trained research workforce are additional challenges to tackling CVD on the island. In response, we propose the Cardiovascular Research Empowerment Workforce (CREW) program aimed at increasing knowledge, skills, and motivation of USVI scholars to pursue medical research careers. UMMSM has long been a national leader in healthcare training with extensive research infrastructure and training expertise targeting learners at all academic levels. CREW will leverage UMMSM’s rich, interdisciplinary research environment which includes the Clinical and Translational Science Institute, Florida Stroke Center, and the Sleep Disorders Training Institute.  Aligned with the USVI’s healthcare context, the proposed program seeks to prepare a cohort of emerging student researchers to address CVD and related comorbidities in the USVI.  The aims of the program are to: 1) Facilitate immersive research experiences to USVI college students addressing CVD and related comorbidities; 2) Implement an engaging curriculum that supports meaningful research experiences, and nurtures trainees’ potential to envision a CVD-focused research science career; and 3) Establish a support network among participants, UMMSM, VIDOH, and UVI scientists to provide trainees with ongoing research and graduate school preparation mentorship. CREW curricula are adapted from UMMSM’s existing programs and structured as an eight-week summer fellowship. It includes hands-on exposure to CVD-related research, professional development, and continuous research training through immersion in a specific USVI-based research project based. In the five-year funding period, ~50 undergraduate scholars from USVI will receive interdisciplinary research training and mentorship and learn strategies to create locally-tailored cardiovascular interventions from evidence-based models in clinical and basic sciences. By doing so, the program will further develop USVI’s undergraduate research training infrastructure and promote locally designed research addressing CVD outcomes and related chronic diseases, serving as a catalyst to increase the representation of USVI scientists in the health care workforce.