University Of Alabama At Birmingham

NIH Research Projects · FY 2026 · 2023-04

This proposal examines the functional role of Sec20, a BH3 and Secretory (Sec) domain protein, in neurons and its relevance to a motor neuron disease using Drosophila. It also examines the interaction between sec20 and GGGGCC (G4C2) repeats of the gene C9orf72 (Chromosome9 open reading frame), a well- known mutation that causes Amyotrophic Lateral Sclerosis (ALS) and Frontotemporal dementia (FTD) in humans. ALS-FTD are two progressive, adult-onset neurodegenerative diseases, often occurring together. The particular focus of this proposal is neuronal loss and mitochondrial and autophagy defects in the brain. Dysfunctional mitochondria and autophagic failures have emerged as important factors in neurodegenerative diseases. They may actively mediate these diseases or exacerbate them. sec20 is the ortholog of vertebrate Bnip1 gene, which is a member of the Bcl2 interacting protein family. Bnip1 has been tentatively identified as a risk factor for ALS and FTD in humans. We found that loss of function for sec20 in the CNS in Drosophila caused severe motor neuron disease and death. There was neuronal loss, mitochondrial dysfunction and autophagic failures in these flies. The disease and these molecular features had similarities to the motor deficits disease caused by the expression of G4C2 repeats. The phenotypes caused by the G4C2 repeats were upstream of sec20 in flies as well as in humans. Thus, our specific aims are: 1) Determine the molecular basis for the loss of motor neurons in sec20 mutant flies, 2) Determine if defective mitophagy and autophagy in sec20 mutants contribute to the disease, and 3) Delineate the interaction between sec20 and the c9orf72-G4C2-R in the CNS. In Drosophila, we go from phenotypes to genes and then molecular underpinnings. There is always a bottom line with this system. Our aims investigate the basics of the phenotypes caused by these genes/mutations. These studies will help understand the function of Sec20 in the CNS and how it relates to G4C2 repeats in the biology and in the diseases of the brain.

NIH Research Projects · FY 2026 · 2023-04

Project Summary Bronchopulmonary dysplasia (BPD) is common in extremely low birth weight (ELBW) infants. Recently, we discovered that the strongest biomarker signal was of microRNA let-7b-5p, with a 46-fold increase (p<0.001) at birth in the blood of infants who subsequently developed severe BPD (versus no BPD) many weeks later at 36w post-menstrual age. We also found a 14-fold increase of let-7b-5p on day 1 in the tracheal aspirate of infants who subsequently developed BPD. In cell culture, airway epithelial cells were the primary source of let-7b-5p, that increased with hyperoxia. We found that excessive let-7b inhibits angiogenesis, and that let-7b inhibition during hyperoxia improves lung development in newborn mice. In the “Let-7b in BPD” project, we will build upon our exciting discovery of let-7b-5p as a robust biomarker of BPD, and determine its relevance to lung development and BPD. We will test the central hypotheses that miRNA let-7b-5p is (a) a valuable biomarker for staging, monitoring disease progression and response to therapy, (b) is released from airway epithelial cells by oxidative stress, (c) is a contributor to dysregulated angiogenesis in bronchopulmonary dysplasia, and (d) that inhibition of let-7b signaling improves lung angiogenesis and attenuates the BPD phenotype. We will test the hypotheses by the following Specific Aims: Specific Aim 1 – Determine if plasma let-7b-5p concentrations in extremely preterm infants track with lung disease progression and correlate with response to therapy. Let-7b-5p will be measured in serial plasma samples from a well characterized prospective cohort of 150 extremely preterm infants. We will define the temporal changes in let-7b-5p with respiratory illness severity, BPD staging and lung mechanics at 36w PMA, and with clinical therapies. Specific Aim 2 – Determine the mechanisms of Let-7b release by newborn mouse lung airway epithelium To confirm that the let-7b-5p release by oxidative stress is the key upstream mechanism, we will use novel transgenic mice. We will test the hypothesis that reduction of mitochondrial ROS reduces let-7b-5p and the BPD phenotype, and determine the role of Nrf2 and NF-kB signaling using specific inhibitors/modulators in cell culture models. Specific Aim 3 – Determine effects of excessive let-7b-5p signaling on lung microvascular development. We will test the hypothesis that over-expression of let-7b-5p induces impaired lung microvascular development, inducing a BPD phenotype in newborn mice even in normoxia, and that inhibition of let-7b-5p improves lung development in hyperoxia- exposed newborn mouse lung (BPD model).

NIH Research Projects · FY 2025 · 2023-04

SUMMARY: Our understanding of the mechanisms used by the host immune system to fight growing tumors has increased exponentially in recent decades, ushering in the development of new therapeutic approaches such as immune checkpoint blockade (ICB) that have revolutionized cancer treatment. Despite these advances, many patients fail to respond to even the most promising immunotherapies. Thus, there persists an urgent need to identify the host factors responsible for the limited efficacy of these otherwise potent immunotherapies in cancer patients. Not surprisingly, identifying mechanisms of immunotherapy resistance is an active area of research. However, the vast majority of pre-clinical and clinical studies in this area do not take into account the effects of common patient co-morbidities like obesity. This is notable because it is estimated that by 2050 half of all adults in the U.S. will have obesity (defined as a Body Mass Index (BMI) of >30 kg/m2). Studies focused on understanding the impact of obesity on anti-tumor immunity and cancer immunotherapy outcomes have grown dramatically in number. However, the field still lacks a clear understanding of when obesity is beneficial for cancer patients, when it is not, and why. Renal cancer is one of 13 tumor types whose prevalence is increased by obesity. Multiple independent studies have found that even in advanced renal cancer patients, 50-60% display BMI-defined overweight or obesity at ICB treatment initiation. Thus, identifying the biological drivers of obesity-associated ICB resistance versus susceptibility in renal cancer is a critically important area of research that could directly impact the majority of patients battling this disease. We have been studying ICB resistance in the context of host obesity. We retrospectively examined outcomes in a cohort of renal cancer patients who received standard of care anti-PD-1. In patients with BMI-defined obesity at treatment initiation, 67% exhibited cancer progression at 12 months, whereas the remaining 33% were progression-free, illustrating the highly variable effects of host obesity on patient outcomes. This trend was reflected in our pre-clinical model of orthotopic renal cancer, in which we found that host obesity is linked to cancer progression in 56% of mice that received an anti-PD-1-based combinatorial immunotherapy. Here we will combine pre-clinical murine and prospective human subject studies to test the hypothesis that sustained high levels of IL-1b inflammation during ICB therapy are associated with poor outcomes in renal cancer patients who have obesity. We further predict that blockade of intratumoral IL-1b will result in a favorable remodeling of the renal TME and improve ICB outcomes. Our findings will permit a more nuanced understanding of the obesity-associated mediators of ICB resistance in renal cancer patients, thereby: 1) providing new metrics to use for identifying patients who are less likely to respond to ICB and 2) facilitating the development of new therapeutic and clinical management strategies for patients with RCC or other solid tumors that are treated with ICB therapeutics.

NIH Research Projects · FY 2026 · 2023-04

PROJECT SUMMARY/ABSTRACT Cardiovascular disease is the leading cause of death in the world. Changes in cardiac metabolic substrate utilization underlie, and may play a causative role in, the development of heart failure. A critical point of regulation in the ability of the heart to fully oxidize glucose is that of the pyruvate dehydrogenase (PDH) complex that, in the heart, is negatively regulated by phosphorylation mediated by two PDH kinases (PDK2 and PDK4). PDKs are differentially regulated in response to physiological stimuli (e.g., exercise) and pathological stimuli (e.g., heart failure). Therefore, they represent likely candidates for therapeutic intervention. However, early attempts to regulate the PDKs have not fully tested the individual isoforms. A long-term goal of our laboratory is to understand the role of these different isoforms in the progression of heart failure. Our preliminary data using germline knockout mice for either of these isoforms in a pressure-overload induced model of heart failure, provides compelling data for a protective role for loss of Pdk2, and an exacerbated role for loss of Pdk4. Furthermore, our initial characterization identified several PDK isoform specific molecular differences, including higher protein acetylation in Pdk2-/- hearts and a higher mortality in Pdk4-/- mice. We also provide evidence to support a mechanism by which the differential acetylation is targeted to the nucleus and may play a role in the histone code, linking it to epigenetic regulation of gene expression. A critical barrier in determining the molecular mechanisms of these PDK isoforms has been the ability to examine each individually. To overcome this barrier, we have generated inducible cardiomyocyte specific knockouts to test the tissue-specific roles of these kinases in adult hearts. The objective of the current proposal is to test the hypothesis that loss of PDK2, but not PDK4, is cardioprotective through differential epigenetic (histone protein acetylation) and transcriptional regulation in response to pressure-overload induced heart failure. We have developed two novel mouse models to test this hypothesis. In this proposal we will: determine the role of each PDK isoform in heart failure progression (Aim 1) and identify molecular mechanisms of these differences (Aim 2). Collectively, the completion of these studies will provide fundamental insights into the mechanistic basis for individual PDK isoforms in the regulation of cardiac gene expression contributing to the development of heart failure.

NIH Research Projects · FY 2026 · 2023-04

ABSTRACT Acute kidney injury (AKI) affects up to half of critically ill patients admitted to intensive care units (ICU). In patients with AKI and hemodynamic instability, continuous renal replacement therapy (CRRT) is the preferred dialysis modality. ICU mortality in this vulnerable population is high but kidney recovery occurs in up to two-thirds of survivors. Universally accepted and accurate approaches for predicting survival or kidney recovery in these patients do not exist currently. This is clinically relevant as prediction of key outcomes could guide decision-making of CRRT delivery, goals of acute care, and personalized post-ICU care according to kidney recovery prognosis. Since there are no proven interventions to improve outcomes in these patients, identification of modifiable risk factors and sub-phenotypes is necessary to develop precision medicine approaches in CRRT. Due to advances in artificial intelligence (AI) and availability of multi-modal data, deep learning (DL) –a subset of AI– is a valuable approach that allows construction of accurate and reliable risk prediction models. Further, the use of novel algorithms such as the Feasible Solution Algorithm (FSA) could help identify patient sub-phenotypes and model applications. We propose to develop and validate innovative and reproducible DL approaches to predict RRT-free survival at actionable timepoints and use FSA to identify patient sub-phenotypes with differing RRT-free survival risk according to multi- modal data. Our published preliminary data demonstrated superiority of DL models compared to optimized logistic regression for RRT-free survival prediction. Prediction of 24-hour mortality was improved by incorporating time-series data during CRRT. We hypothesize that time-series multi-modal data (including EHR and CRRT machine data) will generate accurate and generalizable risk prediction to guide clinical interventions and identify sub-phenotypes for model interpretation and clinical utility testing. We will utilize datasets from 9 institutions that encompass multi-modal EHR clinical data and programmatic and therapy data from CRRT machines for model and sub-phenotyping development, testing, and independent validation. This innovative research will 1) assist development of clinical decision support platforms to guide informed CRRT delivery and improve clinical outcomes and 2) identify sub-phenotypes of patients that could benefit from more personalized and testable novel CRRT interventions.

NIH Research Projects · FY 2026 · 2023-04

Complexities in treating breast cancer (BCa) with bone metastasis are aggravated by a vicious protumorigenic pathology involving a shift in skeletal homeostasis towards aggressive osteoclast activity and polarization of myeloid cells, favoring M2 macrophage (MФ) and myeloid-derived suppressor cell (MDSC) accumulation as key mediators of immunosuppression. In addition to the tumor cells, protumorigenic myeloid cells contribute to the cascade by expressing checkpoint ligands, blunting antitumor functions of effector T cells. Hence, a better understanding of key signaling mechanisms that alter skeletal and immune homeostasis towards protumorigenic functions will enable the designing of new combination therapies targeting this biphasic effect. In this pursuit, we have identified that in addition to robust activation of osteoclast precursors, receptor activator of nuclear factor kappa-Β ligand (RANKL) plays an important role as an osteoimmune link in MФ polarization and programmed death-ligand 1 (PD-L1) expression. We identified that elevated RANKL from BCa cells induce paracrine effects on differentiation of monocytes to immunosuppressive M2 MФ in a spatiotemporal manner. Preliminary studies presented in this application indicate that whereas RANKL canonical autocrine signaling via RANK activates a feed-forward loop in BCa cells, non-canonical RANKL signaling enhances PD-L1 expression in M2 MФ and MDSCs via the leucine-rich repeat containing G-protein coupled receptor (Lgr4). Based on our published and preliminary findings, the overarching goal of this proposal is to expand our understanding on the pleiotropic mechanisms of RANKL in BCa immunosuppression and bone damage, and to test the potential of combining a novel osteoprotegerin (OPG) cell therapy without interfering in TNF-related apoptosis-inducing ligand (TRAIL) function, with checkpoint blockade and chemotherapies, to reverse tumor- associated pathology in the immune and skeletal systems. We recently adopted a protein structure-based engineering approach and identified a critical domain on OPG for TRAIL binding and successfully developed and validated in vivo an OPG variant (OPGY49R) that retains RANKL binding, but lacks TRAIL binding. Preliminary studies, directly comparing a cell-based, single-application OPGY49R treatment with multiple applications of a neutralizing RANKL mAb therapy indicated systemically stable levels of OPGY49R from a single injection and a significant decrease in CD8+ T cell exhaustion, compared to RANKL mAb treatment. More importantly, OPGY49R greatly decreased metastasis of primary tumors in vivo, demonstrating its potential advantage over the RANKL mAb, denosumab, which failed to delay bone metastasis or disease recurrence in patients with high-risk early- stage BCa in a recent international double-blinded randomized placebo-controlled, phase 3 study (D-CARE). This proposal will test this novel, biologically driven combination therapy approach by using immunocompetent mouse models of BCa, as applicable to both pre-metastatic and metastatic disease.

NIH Research Projects · FY 2025 · 2023-04

PROJECT SUMMARY/ABSTRACT The major objective of this proposal is to identify Tiam1-mediated synaptic plasticity as the molecular mechanism underlying opioid tolerance and validate Tiam1 as a promising therapeutic target in the relief of tolerance. Opioid pain medications remain the gold standard for the treatment of moderate to severe perioperative and chronic pain. However, over time, opioid use can result in tolerance, which is a primary driver for opioid misuse and overdose that directly contribute to increased morbidity and mortality. Opioid action at µ opioid receptors (MORs) expressed by nociceptors not only acutely depresses nociceptive transmission, but can induce glutamate release and brain-derived neurotrophic factor (BDNF) release in the spinal dorsal horn, which initiate downstream events that trigger the molecular, synaptic, and network-level adaptations that drive tolerance. Among these, synaptic plasticity is assumed to be the key determinant in opioid tolerance. However, the molecular mechanisms that trigger synaptic plasticity remain unclear. Rho GTPases, activated by guanine nucleotide exchange factors (GEFs) and inhibited by GTPase-activating proteins (GAPs), play important roles in dendritic spine morphogenesis and synaptic plasticity by controlling actin cytoskeleton remodeling in response to extracellular cues. We and others previously identified the Rac1-GEF Tiam1 as a critical regulator of dendrite, spine, and synapse development, which couples synaptic N-methyl-D-aspartate receptors (NMDARs) and TrkB receptors to Rac1 signaling-mediated actin cytoskeleton remodeling during brain development. In preliminary studies, we found that Tiam1 is activated in the spinal dorsal horn in response to chronic morphine treatment and it modulates synaptic remodeling by promoting chronic morphine-induced actin polymerization and synaptic NMDAR expression. Genetic deletion of Tiam1, deletion of Tiam1 from spinal dorsal horn neurons, or pharmacological blockade of Tiam1 signaling prevents the development of morphine tolerance. Moreover, combination morphine and Tiam1 inhibitor therapy reduce morphine tolerance in completer Freund’s adjuvant (CFA) inflammatory pain management. In this proposal, we will use a multidisciplinary approach to test our central hypothesis that Tiam1 links opioid-induced activation of synaptic NMDARs and/or TrkB receptors to Rac1 signaling in spinal dorsal horn neurons, resulting in synaptic structural and functional plasticity via actin cytoskeleton reorganization and NMDAR stabilization, which together underlies opioid tolerance. Moreover, we will determine whether blocking Tiam1-mediated synaptic plasticity with Tiam1 inhibitor or antisense oligonucleotides (ASOs) produces the long- lasting relief of opioid tolerance. The contribution of this proposed research is significant because it will uncover a previously unknown mechanism that underlies opioid tolerance and will provide a promising therapeutic target for the long-lasting relief of opioid tolerance.

NIH Research Projects · FY 2026 · 2023-04

PROJECT SUMMARY The thesis of this R35 Program is that immune cell derived “pathogenic exosomes” are key players in chronic obstructive pulmonary disease (COPD) and that their role can be modeled in a smoking mouse model of exosome transfer to attain novel understanding of the disease which can in turn be applied to COPD. This R35 Program is at the cutting edge of innovation having discovered the existence of alpha-1 antitrypsin (α-1AT) resistant neutrophil elastase (NE)+ neutrophil (PMN)-derived exosomes in COPD patients. In doing so, the Program has challenged the concept that protease activity is solely solution phase and shown extracellular matrix (ECM) proteolytic activity is hugely enhanced by protease attachment to the extracellular vesicle (EV) surface. Consequently, we: i) describe what appears to be the first EV that can transfer a COPD-like phenotype from humans to mice; ii) elucidate a new mechanism by which proteases escape anti-protease inactivation, leading to ECM degradation and cell death via receptor-interacting protein kinase 3 (RIPK3); iii) describe a new model that transfers a COPD phenotype from cigarette smoke (CS) exposed mice to naïve mice via immune cell- derived exosomes; iv) use the mouse model to discover a new CS induced protective mechanism against exosomal damage as well as a mechanism for exosomal self-propagation; v) uncover new therapeutic targets for exosomal damage and; vi) translate the new findings to better understand disease in COPD patients and smokers. Another highly significant aspect of this research Program is training/mentoring. In the past ten years, which includes the PI's inaugural R35 grant, the PI has been or is currently mentor to two PhD students, five medical students, an MD/PhD student, six K awardees, four postdoctoral trainees, a pulmonary fellow, and a cardiology fellow. The PI currently mentors seven junior faculty members. Overall, our Program is both multidisciplinary – employing chemistry, biochemistry, molecular and cell biology, animal physiology, etc. – and translational in pairing basic scientists with physician scientists. Our team and trainees fulfill the NHLBI/NIH mission to have diversity by including African American, Latina, Asian, and Caucasian members. Our program fulfills all four goals of the NHLBI Strategic Vision by: a) elucidating a new exosomal aspect of human biology; b) reducing human disease by new therapeutic development against the exosomal pathway; c) developing a work force and an animal model exosome resource and; d) translating the exosome research to human disease.

NIH Research Projects · FY 2026 · 2023-04

PROJECT SUMMARY The overarching goal of this project is to quantify the temporal kinetics of T-cell activation and infiltration during novel combination intratumoral oncolytic herpes simplex virus (oHSV) immunotherapy with advanced molecular immuno- positron emission tomography (PET) imaging in preclinical models of glioblastoma (GBM). Novel targeted oHSV immunotherapy has been shown to directly kill GBM tumor cells as the virus selectively replicates within and lyses malignantly transformed cells, however there is a knowledge gap in understanding the kinetics of the immune cell changes and how to harness those changes to improve therapeutic efficacy. As inflammation and pseudo progression are key characteristics of immunotherapy-induced tumor changes, standard imaging methods fail to provide reliable response assessments, which can sometimes take up to six months to reveal through clinical changes in tumor size. Advanced quantitative imaging strategies can provide spatial and temporal information on biological alterations prior to the clinically observed, downstream changes in tumor size. Immuno- PET imaging could stratify selection of patients who pursue immunotherapy and guide the timing and sequencing of combinatorial therapies. We have preliminary data showing that there are differential responses in CD8 expression in gliomas during oHSV therapy and that oHSV increases CD8 infiltration, as we can image these changes with PET. Further, preliminary evidence shows that granzyme B increases (as measured by GZP-PET) are related to overall tumor response. The overarching hypothesis is PET molecular imaging can guide targeted IL-12 and IL-18 oHSV to enhance therapeutic efficacy and extend survival in preclinical models of GBM. There are three specific aims to answer this hypothesis: Aim 1. Using advanced molecular immunoPET in GBM murine models with biological validation, determine if early T-cell kinetics of infiltration and activation are increased following oHSV therapy alone, or with oHSV expressing IL-12 and/or IL- 18. We will quantify longitudinal alterations in T-cell infiltration with [89Zr]-CD8-PET imaging of CD8+ cells and quantify longitudinal alterations in T-cell activation (granzyme B) with [68Ga]-GZP-PET imaging. Aim 2. Using advanced molecular immunoPET in GBM murine models, determine if secondary therapeutics (radiation therapy, or immunomodulatory immune checkpoint inhibitors) used in combination with oHSV IOT produce increased intermediate term T cell infiltration and activation and improve long-term tumor response. Aim 3. Determine if secondary boosts of oHSV IOT during imaging-identified timing windows of decreased T cell infiltration and activation can salvage therapeutic response. ImmunoPET imaging allows longitudinal quantification of underlying immunological kinetics during oncolytic herpes simplex virus (oHSV) immunotherapy (IOT) that will allow optimization of therapeutic regimens on a personalized basis.

NIH Research Projects · FY 2026 · 2023-03

PROJECT SUMMARY Individuals with rare genetic diseases do not receive attention from the medical and research community. One out of two patients diagnosed with a rare disease is a child, meaning that patients and their families must endure long battles relating to disease progress throughout their child’s lifetime. Therefore, characterization of clinical/molecular aspects of rare diseases will greatly benefit young patients and their families. SON is a DNA- and RNA-binding protein that plays dual roles as an RNA splicing factor and a transcriptional repressor. Our research team recently identified Zhu-Tokita-Takenouchi-Kim syndrome (ZTTK syndrome), a rare genetic disease with multi-organ abnormalities caused by heterozygous loss-of-function mutations in the SON gene (SON haploinsufficiency). Our research and publication played a key role in documenting this syndrome in major public databases to facilitate clinical diagnosis. As a first step in supporting ZTTK families and promoting awareness, we recently launched an official foundation, the ZTTK SON-Shine Foundation. Our recent efforts revealed that many children with ZTTK syndrome experience various hematopoietic disorders and immune dysfunction, which sometimes leads to life-threatening sepsis. To understand the hematopoietic abnormalities associated with ZTTK syndrome, we have generated mouse models of Son knockout (KO) and Son haploinsufficiency. Our preliminary data demonstrated that complete Son KO in hematopoietic cells causes hematopoietic stem cell (HSC) expansion and embryonic lethality. Furthermore, we found that Son haploinsufficiency leads to abnormal proportions of lineage-primed multipotent progenitors (MPPs), with an expansion of megakaryocyte-erythroid lineage-primed MPPs and a shrinkage of lymphoid lineage-primed MPPs, which is already evident during fetal liver hematopoiesis and persists in adult hematopoiesis in the bone marrow. Importantly, our RNA-sequencing analyses revealed that critical chromatin modifier genes were the major targets dysregulated by Son haploinsufficiency in early stage hematopoietic stem and progenitor cells (HSPCs). Based on these preliminary data, our central hypothesis is that Son haploinsufficiency directly and indirectly alters the expression/splicing of key chromatin modifiers, which collectively reshapes chromatin status at the level of HSCs and MPPs, and this leads to skewed lineage bias and impaired functional output of HSCs/MPPs. To test these hypotheses, we will investigate how Son haploinsufficiency affects functional output of HSCs and lineage primed MPPs in vivo (Aim 1), and will dissect the underlying molecular mechanisms by which Son haploinsufficiency leads to HSPC abnormalities (Aim 2). Successful completion of this proposed study will significantly advance our knowledge about ZTTK syndrome-associated abnormal hematological features and will serve as a valuable resource to identify therapeutic strategies. Most importantly, this study will bring hope to children and their families battling this rare disease.

NIH Research Projects · FY 2026 · 2023-03

Youth-onset type 2 diabetes (YO-T2D) is increasingly prevalent in parallel with the obesity epidemic, yet effective treatment and prevention strategies are limited. The physiologic increase in insulin resistance occurring during puberty, in combination with obesity-related insulin resistance, enhances the risk of T2D. Yet, it remains unclear why some youth progress through puberty with intact β-cell function, while others do not, despite similar phenotypic and metabolic characteristics. More information is needed regarding the unique events during puberty to better understand 1) the basic pathophysiology of glucose control, insulin sensitivity, β-cell function, and T2D risk in youth, 2) differences among girls and boys, populations at highest risk, and urban and rural geographies, and 3) the potential contribution of other risk factors including psychological, behavioral, and social and external contexts. Importantly, this research needs to address the timeline of pathophysiology and progression from normoglycemia or prediabetes to YO-T2D. The DISCOVERY of Risk Factors for Type 2 Diabetes in Youth (DISCOVERY) study provides a unique opportunity to characterize the risk progression profile and mechanisms underlying the development of YO-T2D, and evaluate the effects of modifiable and non-modifiable risk factors. Ultimately, the results of this study will establish a basic pathophysiology to inform future studies aimed at achieving target glycemia, improving insulin sensitivity, preserving β-cell function, and/or preventing YO-T2D. To address this goal, DISCOVERY will recruit, enroll, and follow a nationally-representative cohort of 3,600 at-risk obese youth in early puberty; extensively phenotype them as they transition through puberty; and characterize the course of decline and dysfunction in pathophysiological indicators that lead to YO-T2D. The expected duration of the DISCOVERY is 5 years, including planning, recruitment, follow-up, analysis, and reporting. In addition, DISCOVERY will store longitudinal biospecimens and genetic material with the intention of acquiring additional ancillary funding to pursue analysis of emerging indicators. The University of Alabama at Birmingham (UAB) has experience in multicenter and diabetes-related investigations and will contribute to DISCOVERY through the recruitment of approximately 240 at-risk youth, implementation of the IRB-approved consensus protocol, participation on DISCOVERY committees, and collaboration on the analyses and dissemination of the findings from DISCOVERY.

NIH Research Projects · FY 2026 · 2023-03

Abstract: Non-melanoma skin cancer (NMSC), which include basal cell carcinoma (BCC) and squamous cell carcinoma (SCC) is one of the most common types of cancers in the US. Due to compromised immunity, solid organ transplant recipients (SOTRs) are at a much higher risk for NMSC and it often becomes the cause of death in SOTRs. Prevention of NMSC is an ideal strategy, particularly for immune compromised populations. Rexinoids are small molecule drugs and are able to prevent skin cancer. However, some of the adverse effects may lead to non-compliance in their use. We have developed two distinct classes of structures of rexinoids: UAB30 and UAB20, which are highly selective, non-toxic and orally bioavailable RXR agonists. These agents are also highly effective in preventing skin cancer as shown in our preliminary data. Both UAB30 and UAB20 do not cause hyperlipidemia, an effect usually associated with clinically approved rexinoids. Dampening cancer associated inflammatory biomarkers is also an attractive property of these agents. Thus, our enthusiasm for further developing highly effective analogs of these agents as cancer chemopreventive agents has prompted us to submit this proposal. Guided by x-ray crystallography and biophysical studies, we propose to develop novel analogs of UAB30 & UAB20 with enhanced potency than parent compounds, with suitable pharmacokinetics for chronic administration and without any overt toxicity. Low energy molecular conformation of UAB30 fit well into the RXR ligand-binding pocket (LBP). Our strategy is to improve the potency significantly without distorting the molecular conformations of these two agents. Therefore, we propose to substitute a hydrogen atom with a halogen and/or a heteroatom. Because of the electronegativity of fluorine and the strength of carbon-fluorine bond, we have reasoned that strategic introduction of fluorine will improve the potency, oral bioavailability, metabolic stability and pharmacokinetics of the newly synthesized analogs. Heteroatoms such as nitrogen can modulate the polarity (logP) value significantly. Therefore, we have proposed to substitute a single carbon of tetralone ring of the UAB30 with a nitrogen. Individually and collectively, these modifications will significantly contribute to the potency of UAB30 analogs and will make them ideal agents for pre-clinical evaluation. Similarly, for UAB20, x-ray crystal structures reveal that a five membered heterocyclic ring is accommodated more favorably and make interactions within the LBP. Therefore, we have proposed to substitute the phenyl ring with heterocyclic rings. These modifications will also modulate the logP and improve pharmacokinetics of these new analogs. Our co-investigator in this application has developed unique murine models of the NMSC, which recapitulate human pathobiology of the disease both in normal population and in SOTRs. These models will be employed in the proposed investigations to define the most effective and non-toxic analogs suitable for chronic administration for NMSC chemoprevention. During this grant period, we expect to develop at least one new rexinoid ready for clinical development after GLP toxicity evaluation.

NIH Research Projects · FY 2026 · 2023-03

ABSTRACT: To develop treatments, it is vital to identify neural mechanisms underlying relapse in opioid use disorder. To dissect neural activity underlying relapse, we developed a novel assay allowing for in vivo two- photon calcium imaging while head-fixed mice engage in heroin self-administration. Using this approach, we find global excitatory activity in the prelimbic-prefrontal cortex (PrL) decreases with acquisition of heroin seeking, an effect that persists through extinction, but then resurges during reinstatement. These dysregulated global PrL dynamics mirror clinical observations that are considered hallmarks of substance use disorder. Importantly, our approach has identified, for the first time, that PrL activity dynamics emerging during acquisition, extinction, and reinstatement are heterogenous, with distinct ensembles exhibiting unique excitatory and inhibitory activity dynamics aligned with behavioral epochs. How these unique activity dynamics guide drug seeking, however, is unknown. This implies greater resolution of PrL activity dynamics is necessary to determine how they functionally regulate behavior. Here, under the expert guidance of Drs. Jim Otis and Peter Kalivas, my K99 training in advanced computational neural analyses and single-cell optogenetics paired with in vivo calcium imaging will resolve the function of discrete PrL ensemble dynamics for relapse (Aim 1). Notably, there are subpopulations of individuals with substance use disorder in whom relapse may emerge through different neurobiological mechanisms. Females form one such subpopulation, as they exhibit enhanced relapse vulnerability and greater prefrontal activity during craving and relapse compared to males. However, this vulnerability covaries with the ovarian hormone cycle, such that peak circulating levels of progesterone (PROG) appear protective. These effects are mediated by its 5alpha-reductase neuroactive steroid metabolite, allopregnanolone (ALLO), which can influence prefrontal circuitry and promote adaptive responding in females. As ALLO acts as a positive allosteric modulator at GABA-A receptors, it likely constrains PrL neuronal activity to suppress drug seeking. As I find ALLO can act directly within PrL to suppress heroin-seeking reinstatement, I hypothesize that PROG and ALLO can disrupt PrL activity dynamics which functionally guide reinstatement. During the R00 phase, my career goal to be an independent investigator will involve building a research program wherein I resolve the influence of PROG and ALLO on PrL activity dynamics during reinstatement using in vivo two-photon calcium imaging. As activation of PrL projections to the nucleus accumbens core (NAcC) is necessary for reinstatement, I will retro-label PrL-NAcC neurons to enable simultaneous monitoring of global PrL and PrL-NAcC ensembles (Aim 2). Using single-cell optogenetic and circuit labelling approaches, I will assess the functional influence of PROG and ALLO on PrL and PrL-NAcC ensembles for reinstatement (Aims 2-3). As I find ALLO but not PROG suppresses reinstatement in males, I will assess steroidal effects on PrL and PrL-NAcC ensemble activity dynamics as a function of biological sex.

NIH Research Projects · FY 2026 · 2023-03

Summary Abstract This application puts for the notion of a brain-gut- retinal axis that becomes dysfunctional in diabetic retinopathy (DR). The innate immune system has been strongly implicated in the pathogenesis of DR, but less is known about the role of the adaptive immune system. At the interface of these two systems is a critical population of cells, Th17 cells, that typically reside in the gut during health. Th17 cells have homeostatic properties, mediating host defense against bacterial and fungal infections; however, it remains unclear how intestinal Th17 cells integrate diverse signals into a set of cellular programs that allow them to maintain tissue homeostasis yet also become pathogenic, serving as primary drivers of tissue inflammation. We have identified a critical role of somatostatinergic neurons in the paraventricular nucleus of the hypothalamus (PVN) in regulation of immune function through “loss of function” studies and “gain of function studies. SST expression is dramatically reduced in the PVN of diabetic animals. Hypothalamic dysfunction, as seen in diabetes, has the capacity to induce injury directly through hyperactivation of sympathetic nerves. Based on preliminary and published data, we put for the hypothesis that: Diabetes-induced loss of inhibitory SST neurons in the PVN drives increased autonomic input to the intestine shifting Th17 cells from a homeostatic to a pathologic state. Pathologic Th17 cells leave the intestine and traffic to areas of tissue injury such as the retina in DR. In the retina the pathologic Th17 cells secrete proinflammatory cytokines that recruitment innate immune cells into the retina exacerbating DR. To examine this hypothesis, we propose the following aims: Aim 1: To test if impaired function of hypothalamic SST neurons in diabetes contributes to hyperactivity of autonomic efferents to the gut and increases activation of enteric neurons. Aim 2: To selectively ablate SST PVN neurons (in the absence of diabetes) and evaluate if this results in increased autonomic input to the gut and a shift from “homeostatic” Th17 cells to a “pathogenic” Th17 cells that migrate to the retina and recruit circulating immune cells. Aim 3: To determine if maintaining hypothalamic SST levels at nondiabetic levels in diabetic mice will preserve the function of homeostatic Th17 cells in the gut and prevent their recruitment to the retina delaying the development of DR. Impact: SST analogues may provide an important complementary strategy for DR management by preventing increased sympathetic drive to the gut and Th17 cell dysfunction.

NIH Research Projects · FY 2025 · 2023-03

PROJECT SUMMARY Although much progress has been made in the treatment and prevention of diarrheal illness over the last century, the developing world is still plagued by diarrheal pathogens, with young children being particularly susceptible to diarrhea-related deaths. The long-term goal is to better understand how the coordinated responses of both immune and non-immune cells contribute to intestinal barrier protection during enteric infection. Our group recently discovered that Th17/22 cells are uniquely charged with protection of the intestinal crypts from enteric pathogens; however, the mechanisms by which these T cells are recruited to and sustain activation of crypt intestinal epithelial cells (cIECs) are unknown. The overall objectives in this application are to (i) elucidate the molecular mechanisms involved in recruiting T cells to cIECs and (ii) determine how recruited T cells interact with cIECs to deliver protective IL-22 signals. The central hypothesis is that IFN–induced upregulation of IEC- derived T cell-recruiting chemokines and MHCII expression promotes T cell recruitment to colonic crypts and targets pathogen-specific T cell-derived IL-22 signals to cIECs, respectively. The rationale for this project is that molecules involved in both recruitment and localized delivery of IL-22 to IECs will likely yield novel targets for therapies in diarrheal illnesses and inflammatory bowel disease (IBD), since responses to enteric pathogens and etiology of IBD share overlapping immune mechanisms. The central hypothesis will be tested by pursuing two specific aims: 1) IFN signaling in Lgr5+ IECs is critical for recruiting host-protective Cxcr3+ Th1 and Th17/22 cells to the colonic crypts; and 2) pathogen-specific T cells protect intestinal crypts via MHCII-dependent, sustained IL-22 delivery to IECs. The Citrobacter rodentium (C.r) model of infectious colitis, which closely models human E. coli infections, will be used to test both aims in this proposal. In Aim I, mice with deletion of the IFNR1 in Lgr5+ IECs will be used to to test the hypothesis that IFN signaling in Lgr5+ IECs is required for upregulation of T cell-recruiting chemokines Cxcl9 and Cxcl10 and adoptive transfer of Cxcr3-deficient T cells will be used to determine if the Cxcl9/10-Cxcr3 axis is involved in recruiting CD4+ T cells to cIECs. In Aim II, mice deficient for MHCII expression on IECs will be utilized to test the requirement of antigen presentation by IECs for IL-22- mediated protection of the intestinal crypts. In addition, mice that report TCR stimulation and adoptive transfer of C.r-specific T cells will be used to test the hypothesis that antigen-specific T cell–IEC interactions are important for protection from C.r infection. The research proposed in this application is innovative because it utilizes a newly engineered C.r strain to track antigen-specific T cells in the colon, and it offers unique insights into previously unrecognized roles of Lgr5+ intestinal stem cells. The proposed research is significant because it is expected to provide novel mechanisms by which T cells and IECs cooperate to protect the host from enteric pathogens. Ultimately, such knowledge has the potential for innovative therapies for infectious colitis and IBD.

NIH Research Projects · FY 2026 · 2023-02

PROJECT SUMMARY Mechanisms Controlling the Development and Function of Intestinal Effector Treg Cells. The pathogenesis of a spectrum of disorders referred to as inflammatory bowel disease (IBD) is characterized by immune dysregulation to components of the enteric microbiota. Findings from mouse and man highlight a critical, non-redundant role for the immunoregulatory cytokine IL-10 in maintenance of intestinal immune homeostasis. Our labs have shown that Foxp3+ regulatory T (Treg) cells are, overwhelmingly, the major source of IL-10 in the intestines, where many of these cells co-express the canonical “Th17” transcription factor, RORγt—particularly in large intestine (LI). However, IL-10 is only produced by a subset of Treg cells—defined as ‘effector’ (e)Treg cells. Mechanisms that control the development of eTreg cells are incompletely understood. The premise of this application, founded on recent discoveries from, and synergy between, the two PIs (Weaver, Hatton) is that heretofore unappreciated interplay between signaling pathways of the IL-2 and TNF superfamilies has a central role in regulating the development and function of eTreg cells in the intestines. Specifically, we have identified the TNF receptor superfamily signaling pair, TNFSF15-TNFRSF25 (TL1A-DR3), as an important amplifier of the transition of “central” (c)Treg cells into eTreg cells and propose that this pathway plays an important role in calibrating IL-2–driven control of the Treg cell program via multiple mechanisms. We hypothesize that the TL1A- DR3 pathway is non-redundant in its regulation of IL-2 receptor (IL-2R) signaling to modulate the size and cTreg– eTreg balance of the LI Treg cell pool both at homeostasis and under inflammatory conditions. Further, we posit that DR3 acts through multiple mechanisms to alter output of the IL-2R to regulate development of IL-10– producing eTreg cells. We propose that a major driver of eTreg cell development is increased STAT3 output of the IL-2R, which is enhanced by actions of DR3 to: (i) increase sensitivity of the IL-2R; and (ii) decrease pSTAT3 degradation, thereby altering the IL-2–induced STAT5/STAT3 ratio. Finally, we propose that other TNFRSF members contribute to eTreg cell maintenance after DR3 expression declines. We test this hypothesis through complementary, but not inter-dependent, Aims, leveraging: a novel method for generating stable, colitis-curing Treg cells ex vivo; new approaches for efficient gene knock-downs in primary T cells; and new gene-targeted mouse models to define mechanisms governing the convergence of these signaling pathways in controlling the transcriptional regulation of Il10 and the eTreg program. Successful completion of these Aims will establish new biological paradigms and inform novel therapeutic approaches by which endogenous IL-10 can be up-regulated and the differentiation and function of eTreg cells enhanced to treat human IBD.

NIH Research Projects · FY 2026 · 2023-02

Project Summary/Abstract For the more than 5.5 million people in the U.S. who use wheelchairs as their primary mode of ambulation, there is a pervasive lack of research on reducing cardiometabolic risk through structured exercise. Confirmatory analyses have revealed that wheelchair users live predominantly sedentary lifestyles and have substantially higher cardiometabolic risk factors compared to the general population. Although few exercise training studies have investigated cardiometabolic risk in certain subgroups with a disability, small sample sizes and homogeneous groups limit the translatability of this knowledge into clinical practice risk reduction strategies for wheelchair users. To address these issues, we propose to use a robust remote training system with built-in videoconferencing and real-time monitoring of vital sign data (e.g., heart rate, respiratory rate). This procedure will allow us to examine the efficacy of an evidence-based Movement-to-Music (M2M) program adapted for telehealth delivery and cardio emphasis (M2M-C). We are proposing a two-arm randomized controlled trial including 132 wheelchair users with a poor cardiometabolic profile (e.g., elevated triglycerides) recruited through our established network of clinics serving this population. The long-term goal of this proposal is to develop an effective and enjoyable modality for promoting health-enhancing exercise for wheelchair users by confirming exercise dose requirements for this underrepresented group. The primary aim will examine the average treatment effects of a 24-week M2M-C program on core indicators of CMH in wheelchair users with ≥ 2 cardiometabolic risk factors. CMH outcomes will be measured via blood tests (i.e., high sensitivity C-reactive protein, hemoglobin, fasting insulin, triglycerides, and cholesterol) and DEXA scan (i.e., body composition) at baseline and after 12 and 24 weeks of synchronous M2M-C training. The secondary aim will explore the beneficial effects of M2M-C on cardiovascular capacity, physical activity, and quality of life. Cardiovascular capacity will be measured via peak oxygen consumption. Physical activity and quality of life will be measured by self-report instruments validated for this population. The tertiary aim 1 is to evaluate the sustained effects of M2M-C (24 to 36 weeks) on physical activity. After M2M participants complete the 24-week program, they will be instructed to perform asynchronous exercise training using guided M2M online videos for an additional 12 weeks. The tertiary aim 2 is to examine the heterogeneity of treatment effect (HTE), which aims to understand whom the intervention is most effective.

NIH Research Projects · FY 2026 · 2023-02

The NLRP3 inflammasome is unique in that it requires a two-step activation process: the priming and the activating. The priming step involves the induction of NLRP3 and pro-IL-1β, whereas the activating step results in the full inflammasome activation triggered by a NLRP3 activator. Although the mechanism leading to the activation of the NLRP3 inflammasome has been increasingly clear, the regulation of this process remains poorly defined. Dysregulation of NLRP3 inflammasome has been frequently implicated in the bacterial pneumonia caused acute lung injury (ALI). MafB is a member of the large MAF transcription factor subfamily and has been implicated in immune disorders, which mainly concerns its role in macrophage apoptosis, phagocytosis and the complement system. In our preliminary studies, we made an unexpected discovery that uncovers a novel function of MafB in regulating the NLRP3 inflammasome activation. Our findings suggest that MafB is a new negative regulator of the NLRP3 inflammasome in vitro and in vivo. We hypothesize that LPS, and P. aeruginosa induced MafB downregulation is a crucial step for the NLRP3 inflammasome priming; MafB is a key player that regulates the NLRP3 inflammasome activation; MafB plays an important role in the pathogenesis of ALI; as well as targeting MafB is an effective therapeutics for ALI. We aim to comprehensively delineate the regulation of MafB expression at the transcriptional and post-translational levels during the NLRP3 inflammasome priming; to delineate the mechanism by which MafB inhibits the NLRP3 inflammasome activation; and to determine the role of MafB in LPS and P. aeruginosa induced acute lung injury (ALI).

NIH Research Projects · FY 2026 · 2023-02

OVERALL SUMMARY Early life stress (ELS) is defined as stressful and traumatic events, such as household dysfunction, neglect, sexual or physical abuse, economic hardship, and exposure to violence, experienced up to 18 years. ELS was identified as a cardiovascular disease (CVD) risk factor over 20 years ago, but mechanistic insights into its effects remain very limited. Exposure to ELS is pervasive in the US with ~50% of children and adolescents having one or more major ELS experiences. ELS exposure increases the risk traditional CVD risk factors - by the 3rd-4th decade of life. A recent analysis of healthcare burden in Europe and North America attributed $748 billion in annual costs to the effects of ELS, with 75% of those costs in people with multiple ELS exposures. The significance of our work aligns with the NHLBI mission to promote the prevention and treatment of cardiovascular diseases enhancing the health of all individuals to live longer and fulfilling lives. Among individuals with a history of ELS exposure, vascular dysfunction (elevated peripheral vascular resistance, increased vascular stiffness) and elevated diastolic blood pressure are already evident in early adulthood. The overall goal of our program project grant (PPG) is to define mechanisms by which ELS leads to CVD risk and inform strategies for prevention and effective treatment of CVD consequences in individuals exposed to ELS. This PPG will address two critical barriers to fulfill our goal: 1) Need for ELS-specific in-depth mechanistic and translational studies; and, 2) Identify modifiable protective factors that can reduce ELS-induced CVD risk. The overarching hypothesis of our PPG is that ELS induces immune cell activation leading to vascular dysfunction with increased hypertension risk and CVD risk that are exacerbated by later life stressors or moderated by resilience/protective factors. This PPG with both basic science and clinical projects utilizes a synergistic and integrative approach translating concepts from clinically relevant rodent models to humans. Our group is extremely synergistic, with each leader bringing unique expertise from different scientific backgrounds focused on our overall goal to understand mechanisms of ELS induced indicators of CVD risk and resilience. Over the past several years, our team built strong collaborations translating discoveries between basic and clinical labs with several pilot grants, co-mentoring trainees, and multiple jointly authored abstracts, manuscripts, and publications. The four projects and three cores are integrated in their goals and impact such that much more will be achieved together than separately. This PPG utilizes a range of approaches to investigate in-depth molecular mechanisms of ELS-induced hypertension and vascular disease risk as well as to delineate protective factors mediating resiliency to this risk. The results will have important translational potential pointing to new intervention or prevention strategies for the health consequences of CVD and reducing the healthcare burden of CVD.

NIH Research Projects · FY 2026 · 2023-02

Project Summary IgA nephropathy (IgAN) is the most common primary glomerulonephritis in the world, with 30-50% of patients progressing to end-stage kidney disease. Diagnosis is biopsy-based, with routine immunofluorescence showing IgA (co)dominant immunodeposits usually with C3 and often with IgG co-deposits. IgA in the immunodeposits is of IgA1 subclass and enriched for galactose-deficient IgA1 glycoforms (Gd-IgA1). There is currently no disease specific therapy for IgAN due to our limited understanding of the underlying mechanisms of disease progression. Recent data indicate that IgG is present in immunodeposits of virtually all IgAN patients and that it is enriched for Gd-IgA1-specific autoantibodies. Furthermore, Gd-IgA1 and anti-Gd-IgA1 IgG autoantibodies are elevated in the circulation of IgAN patients and their levels predict disease progression. Based on these, and other data, we have proposed a multi-hit hypothesis explaining the autoimmune nature of IgAN: Gd-IgA1 is elevated in circulation of IgAN patients and is recognized by anti-Gd-IgA1 IgG autoantibodies, resulting in the formation of circulating immune complexes, some of which deposit in the glomeruli, inciting renal injury. The origin of the autoantigen and the characteristics of specific cell population(s) producing Gd-IgA1 remain unknown. Recently, we showed that immortalized IgA1-secreting cells from IgAN patients produce more Gd-IgA1 autoantigen compared to the cells from healthy controls. Furthermore, only these IgAN-derived cells increase Gd-IgA1 production in response to cytokine stimulation (e.g., IL-6). Follow up analysis found enhanced STAT3 and STAT1 activation only in IgAN B cells, and that it was necessary for Gd-IgA1 production following cytokine stimulation. Single-cell transcriptome analysis of cytokine stimulated B cells from IgAN patients found IgHA1-subpopulations with abnormal expression of genes responsible for regulation of multiple cytokine signaling pathways. These IgHA1-subpopulations also exhibited abnormal alterations in the expression level of glycosyltransferase enzymes relevant to IgA1 glycosylation. Transcriptional data alone does not identify Gd-IgA1 producers. To address this, we developed a novel glycophenotyping method using lectins and recombinant anti-Gd-IgA1 IgG that can target cell surface presentation of Gd-IgA1. This process enriched for low and high Gd-IgA1-producing subpopulations, which had differential activation of transcriptional factors before and after cytokine stimulation. Using this glycophenotyping method with nucleotide conjugation, we can target Gd-IgA1-producing cells for transcriptome analysis, intracellular signaling staining, chromatin remodeling, and specific autoantigen production rates. Together, these studies will elucidate the mechanisms of autoantigen production within specific subpopulations of IgA1-producing cells and potentially lead to the development of new disease therapies for IgAN.

NIH Research Projects · FY 2026 · 2023-02

Project Summary. Despite the numerous pharmacological and immunological approaches for multimodal glioma treatments proposed in recent years, glioma phenotypic and genotypic spatial profiles remain heterogeneous and, therefore, represent the biggest disadvantage for patient outcomes due to the development of treatment resistance. Cell fusions through permanent cell-fusion and temporal tunneling nanotube (TNT) formations are novel, recently discovered sources of intercellular gene transfers and glioma heterogeneity. In our grant, we will provide a detailed analysis of cellular mechanisms essential for intercellular gene transfer via cell fusion and TNT formations for different glioma subtypes in patient-derived tissue, mimicked glioma microenvironment in vitro, and in mouse glioma models in vivo. Cell fusion and TNT formations in the context of tumor heterogeneity will be detailed by following techniques: i) the high-resolution spatial cell transcriptional and histological profiling in tissue slices (Visium platform); ii) the RNA-Seq and WES profiling (Illumina platform) on a single-cell level after cell-type-specific enrichment by flow cytometry technique from dissociated tissue; iii) the proteomics (HCP, mass spectrometry) analysis of molecular complexes involved in gene transfer at different stages of cell interaction. Cell-specific Cas9/gRNA- directed gene knockdown in combination with the target rescue experiments and quantitative Cre/fluorescence-based reporters of cell fusion and TNT formations will be utilized in vitro and in vivo to complement RNA-Seq, WES, and proteomics data, allowing identification of new targets and providing seeds for the development of novel pharmacological inhibitors of cell fusion and TNT formations. Our preliminary data identified horizontal gene transfer via cell fusion and TNT formations between glioma cells themselves and glioma/normal host cells in the hypoxic, inflammatory, mechanically stressed, cytotoxic, and amino acid deprived microenvironmental glioma loci. We analyzed the transcriptomic and proteomic signature of five PDGx cell lines of different molecular subtypes and confirmed that a central node of the cellular stress response, the mRNA-binding protein HuR, is an essential regulator of cell fusion and TNT formations in numerous stress conditions. The key biomarkers of TNT formations (TNFAIP2, GJA1) and numerous endogenous fusogenes are direct HuR mRNA targets and overexpressed in gliomas in a HuR- dependent manner. Therefore, we propose that pharmacological inhibitors of HuR function may serve as suppressors of intercellular gene transfers evoked by cell fusion and TNT formations. In our grant, the impact of the recently developed and patented by our group inhibitors of HuR dimerization (SRI42127 is a lead compound) will be assessed in the regulation of glioma heterogeneity in vitro and in vivo. The high throughput genome-wide in vivo screening of PDGx cell lines transduced with a Human GeCKOv2 CRISPR genome-wide knockout library will be employed to reveal and compensate potential mechanisms of cell fusion tolerance to the HuR inhibitors.

NIH Research Projects · FY 2026 · 2023-02

Colorectal cancer (CRC) is the second-leading cause of cancer-related mortality in the United States. Chronic intestinal inflammation is a major risk factor for the development and progression of sporadic and colitis- associated colorectal cancer (CAC). While the pathogenic effects of tumor-promoting inflammation are widely recognized, the mechanisms driving this process, including the cellular and molecular mediators are poorly understood. This has therapeutic implications especially in CRC where the currently available T-cell checkpoint inhibitors have a limited benefit. In preliminary studies, we used the azoxymethane/ dextran sulfate sodium model of inflammation-driven colon carcinogenesis that allows interrogation of the initiation-promotion events underpinning the development of CAC. We observed that compared to the surrounding non-tumor-bearing colon, the tumor microenvironment is enriched with tumor-infiltrating myeloid (TIM) cells, including neutrophils (PMNs), monocytes, and tumor-associated macrophages (TAMs). PMNs are the predominant population and are recruited through the Cxc-Cxcr2 axis while the monocytes/TAMs (M) appear to rely on the Cc-Ccr2 pathway. Both subsets express pro-inflammatory and immune-suppressive genes thereby suggesting their pro- tumorigenic role. Additionally, the tumor microenvironment has an increased abundance of interleukin (IL)-17 producing CD4+ T (Th17) and gamma-delta T cells. IL-17 is a pro-inflammatory cytokine that promotes CRC growth and increased expression is associated with poor prognosis. Previous studies indicate that TIMs promote Th17 development through the production of IL-23 (IL-23A/IL-12B heterodimer). However, our single-cell RNA sequencing studies revealed that while the PMNs expressed Il23a, they did not express Il12b thereby suggesting an inability to produce functional IL-23. Instead, the PMNs expressed Epstein-Barr virus-induced gene 3 (Ebi3) which has recently been shown to dimerize with IL-23A to form IL-39, a pro-inflammatory cytokine. We hypothesize that the myeloid-Th17 axis is critical for the maintenance of an inflammatory microenvironment that helps sustain CAC progression. We propose the existence of a feed-forward loop in the tumor microenvironment whereby the PMNs and M work synergistically to promote Th17 development, and the resulting IL-17, in turn, recruits and maintains more PMNs and M in the tumor thereby reinforcing an inflammatory microenvironment which fuels CAC progression. Mechanisms to disrupt this loop will be the focus of our proposal. Our first aim will attempt to define the role of TIMs in the pathogenesis of CAC by exploring the effects of blocking colonic recruitment of PMNs and M on tumor development (Aim IA) or their deletion in established tumors (Aim IB). In our second aim, we will explore the role of IL-39 as an alternative mechanism of Th17 development (Aim IIA) and assess its role in tumor development (Aim IIB), including the effects of cell-specific deletion of the Ebi3 in PMNs and CD4+T cells (Aim IIC). These studies will have implications in treatment strategies for patients with CAC and other immune-mediated diseases like inflammatory bowel disease.

NIH Research Projects · FY 2026 · 2023-02

PROJECT SUMMARY/ABSTRACT Primary cilia are present on most renal epithelial cells and their disruption leads to cyst formation. Despite their clinical importance, the function of the primary renal cilium remains poorly understood. In previous studies, we and others demonstrated that when cilia assembly (Ift88 mutant) or function (Pkd1 or Pkd2) is disrupted in adult mice, cysts form slowly in focal areas. This occurs despite nearly all epithelial cells in the kidney being mutant for these genes. A hint as to why these cysts develop in focal locations in the adult-induced mutants has emerged from data indicating that renal injury promotes cyst formation and results in cyst formation throughout the kidney. Based on these data, we propose that PKD2 and cilia regulate renal injury and repair responses and that the focal cysts in the adult-induced mutants result from a few cells that have experienced an injury. In the absence of normal cilia function, these cells enter a state of persistent maladaptive repair (as marked by persistent SOX9 expression) and progress to cystogenesis. While cysts are formed in Ift88 and Pkd2 mutants, the cystic kidney phenotype is always more severe with loss of Pkd2. In double mutants, however, the cystic pathology resembles the milder Ift88 mutant alone. Our analysis further shows that the inflammatory response following injury in the Ift88 and Pkd2 mutant mice is altered and as observed with cyst formation; it is exacerbated in Pkd2 mutants compared to Ift88 or Ift88/Pkd2 double mutants. Collectively, these data suggest that IFT88/cilia and PKD2 regulate signaling activities between the ciliated tubule epithelium and the responding non-ciliated immune cells for initiation and resolution of an injury response and that cyst formation is promoted when these signaling pathways are incorrectly controlled. The goals of this proposal are to define the cellular and then molecular mechanism(s) responsible for the focal and widespread injury-induced cyst formation and how IFT88/cilia and PKD2 are involved in this process. From this proposal, we will determine whether cilia loss/dysfunction alters the kidney's sensitivity to injury and results in an increase in the state of epithelial cell maladaptive repair. We will utilize lineage tracing approaches to determine if descendants of injured cells contribute directly to cyst formation. We will define new IFT88/cilia and PKD2-dependent intercellular signaling networks involved in the injury and repair process and how they are dysregulated when cilia are disrupted (Ift88 mutant) or cilia function is perturbed (Pkd2 mutant). Finally, we will define mechanisms by which loss of Ift88 in a Pkd2 mutant background suppresses cyst severity through changes in these injury and repair signaling pathways. The data from the proposal are needed to: 1) elucidate novel functions of the renal cilium, 2) define cellular mechanisms involved in formation of the sporadic and widespread cysts, 3) understand how IFT88/cilia and PKD2 are involved in regulating injury/repair responses, 4) assess how disrupting this injury response promotes cyst formation and expansion, and 5) explore mechanisms involved in the epistatic relationship between Ift88 and Pkd2 mutations.

NIH Research Projects · FY 2025 · 2023-02

PROJECT SUMMARY Lynch syndrome (LS) is an inherited cancer predisposition syndrome that substantially elevates lifetime risks for multiple cancers in both men and women. Identifying people with LS enables more frequent and earlier cancer surveillance and prevention, which can effectively reduce LS-related cancer morbidity and mortality. However, most people with LS in the United States are not diagnosed. One cost-effective way to identify individuals with LS is by conducting genetic testing on blood relatives of patients diagnosed with LS (or “cascade testing”), but the testing rate is low. Pre-test genetic counseling is a promising means to address many of the barriers to testing and to increase testing uptake. However, pre-test genetic counseling uptake is low in relatives at risk of LS, and the barriers and facilitators have not been systematically studied. Although informational resources for LS genetic counseling exist, they are unlikely to be sufficient for motivating behavior change. Aligned with the National Cancer Institute’s focus on cancer research to advance scientific knowledge and help all people live longer, healthier lives, the proposed research will address these critical gaps through three aims: 1) Identify barriers and facilitators to pre-test genetic counseling among relatives with no personal history of cancer but at risk of LS; 2) Develop a theory-based behavioral intervention to increase pre-test genetic counseling uptake in this population; and 3) Evaluate and optimize feasibility of the trial methods and the behavioral intervention to prepare for a fully powered randomized controlled trial and explore the intervention’s preliminary efficacy. I will apply the Behavior Change Wheel, a well-established behavior change framework, and use mixed methods (i.e., focus groups, usability testing, and a randomized controlled pilot trial) to develop a behavioral intervention to increase pre-test genetic counseling uptake. The proposed research will lead to an R01 proposal to test the efficacy of the intervention. My study team consists of outstanding mentors who have recognized expertise in the methodologies and topic areas of the proposed research, and a genetic counselor who will ensure that the research is clinically grounded. In coordination with my research activities, my career development plan includes structured training and one-on-one mentoring in behavioral medicine and intervention design and development, design and conduct of clinical trials, qualitative methods, and grant writing and management. Along with the abundant research and training resources and the supportive environment for transitioning early-career researchers to independence at Northwestern University Feinberg School of Medicine and Department of Medical Social Sciences, this K99/R00 will enable me to become an independent investigator working to facilitate decision making in patients and other stakeholders who face difficult cancer-related decisions, including those involving genetic risks.

NIH Research Projects · FY 2026 · 2023-02

The etiology for late-onset Alzheimer’s disease (LOAD), which accounts for >95% of AD cases, is unknown. Aging is the greatest risk factor for LOAD, whereas APOE4 is believed to be a major genetic risk factor in acquiring LOAD, with female APOE4 carriers at the greatest risk. Yet, not all of APOE4 carriers, even older female carriers, acquire AD, suggesting that other factors including environmental exposure must play a role. Ozone (O3) is a highly reactive oxidant and one of the most abundant urban pollutants. Recent epidemiology studies show that exposure to high levels of O3 is associated with an increased incidence of AD in the elderly and that APOE4 carriers are more sensitive to O3-induced memory decline than non-APOE4 carriers are, although no study has addressed sex-differences in response to O3. In an R21 project, we tested the hypothesis that O3 exposure synergies with aging and APOE4 leading to AD, using male apoE4 target replacement (TR) mice (only male mice were proposed due to time and budget limitation). We found, surprisingly, that O3 exposure impaired memory in old apoE3 (represents the majority of human population who carry the APOE3 allele) male mice, while old apoE4 or young apoE3 and apoE4 male mice were spared 6. Associated with memory loss, old apoE3 male mice exhibit increased protein oxidative modifications (glutathionylation) and neuroinflammation, compared to other groups6. Old apoE4 male mice, on the other hand, have significantly increased expression/activities of several antioxidant enzymes and diminished protein oxidation as well as neuroinflammation upon O3 exposure6. Our data suggest that an elevated antioxidant capacity may underlie the increased resistance of old male apoE4 mice to O3-induced memory loss. Our data also suggest that APOE4 may affect the sensitivity to O3-induced memory loss in a sex-dependent manner, just as it does to the toxicities of other toxicants. This R01 will continue our R21 project to further test sex-dependent effects of ApoE4 and O3. As oxidative stress plays a critical role in AD pathophysiology and plasma level of estrogen, an inducer of many antioxidant enzymes, decreases with age in females, our new data-supported hypothesis is that O3 exposure synergizes with aging and APOE4 leading to LOAD in sex-dependent manner, which is to promote AD in females but not in males. We will test this hypothesis in two specific Aims, using apoE3 and apoE4 TR mice and a cyclic O3 exposure protocol, which mimics human exposure scenarios. In Aim 1, we will test the hypothesis that O3 exposure induces AD-like pathophysiology in female apoE4 TR mice compared to their male counterparts and this is exacerbated by aging. In Aim 2, we will test the hypothesis that restoration of brain antioxidant capacity with tert-butyl hydroquinone (TBHQ), a canonical activator of nuclear factor erythroid 2-related factor 2 (Nrf2), will eliminate sex-APOE genotype-aging-dependent sensitivity to O3-induced neuropathophysiology. The results from these studies will not only shed new light on the etiology of LOAD but may also lead to the development of new strategies for the prevention and treatment of this devastating disease.