University Of Chicago

NIH Research Projects · FY 2025 · 2023-07

Abstract Breast cancer is the leading cause of cancer death for women globally, with over 2.3 million cases diagnosed each year. Most cases are hormone receptor positive and effectively treated with anti-estrogen therapy, but some patients have aggressive disease and are at risk for recurrence and death without chemotherapy. Gene expression based recurrence assays, such as OncotypeDX, were designed to predict recurrence on hormonal therapy and are used to select patients for chemotherapy. However, these assays are expensive (> $3,000 per test), take considerable time to perform leading to treatment delays, and testing is underutilized or frankly unavailable in low resource settings in the US and globally. Conversely, every patient with breast cancer has a biopsy to confirm the diagnosis, which is routinely analyzed by pathologist to determine subtype of breast cancer and grade. Deep learning is an emerging technique for quantitative image analysis, and can identify non-intuitive features from pathology, including gene expression patterns. In preliminary work, I have demonstrated that deep learning on pathology samples can provide rapid and cost-effective prediction of OncotypeDX score using readily available data, and can identify patients at low risk of recurrence on hormonal therapy. However, OncotypeDX remains an imperfect predictor of chemotherapy benefit, as it was developed to predict recurrence on hormonal therapy. By refining my deep learning biomarker to incorporate clinical and immune features of breast cancer, I can improve accuracy in prediction of chemotherapy benefit and thus the ability to personalize treatment. First, I will capitalize on the recent expansion of clinical data in the National Cancer Data Base to develop a more accurate clinical models of prognosis and chemotherapy benefit. Next, I will use multiplex immunofluorescence to better characterize spatial and cell density features associated with chemotherapy benefit, and use deep learning models to infer these features from standard hematoxylin and eosin stained digital pathology. Finally, I will integrate these clinical and immune models with my existing deep learning pathologic model and validate the integrated model in a multi-institutional cohort. The result of this work will result in a prognostic and predictive deep learning biomarker that makes accurate predictions from readily available clinical, pathologic, and inferred immune features. This approach has the potential to reduce chemotherapy delays due to rapid turnaround time, combat healthcare disparities through improved availability of testing, and improve personalization of treatment by tailoring a biomarker for prediction of chemotherapy benefit.

NIH Research Projects · FY 2025 · 2023-07

Summary A favorable target for radiation synthetic combinations would be a feature of cancer cells critically involved in growth, signaling, repair, or survival that can be blocked with an otherwise non-toxic drug, leaving tumors vulnerable to radiation without adverse effects on normal tissue. This project is directed at validating inhibition of telomerase reverse transcriptase (TERT) as a means to enhance the therapeutic index of radiation and achieving key progress toward translating this strategy to the clinic. While TERT is not expressed in most normal cells, approximately 90% of cancers display reactivation of TERT expression, supporting the catalytic activity of telomerase to maintain telomere integrity despite deregulated growth. While drugs targeting TERT have displayed sufficient safety in patients to evaluate effects of blocking telomere repeat synthesis, this has failed in solid tumors, as telomere erosion is too slow to affect tumor progression. Beyond its essential role in cancer cell immortality, TERT also contributes to pathways that support multiple cancer hallmarks. By limiting oxidative stress, accelerating double strand break repair and supporting cell survival, TERT expression in cancer cells may confer clinically significant resistance to radiation. This raises the question whether transiently targeting TERT during radiotherapy to enhance the toxicity of the resulting DNA damage to the cancer cells might significantly improve the therapeutic index of radiation. In recently published work, our groups described a novel class of TERT inhibitors inspired by the antibiotic chrolactomycin. Like the natural product, our streamlined natural product analogs react with an active site cysteine in the TERT reverse transcriptase active site. The optimized inhibitor, NU-1, is otherwise nontoxic in vitro or in vivo, but inhibits telomerase activity at low micromolar concentrations. NU-1 confers sensitivity to radiation to TERT-expressing cancer cells. Our data suggest that TERT may promote non-homologous end- joining repair, thereby affecting repair pathway choice. Finally, using a syngeneic tumor model in BALB/c mice, we have demonstrated marked sensitization to radiation in vivo, apparently mediated by persistent DNA damage and increased anti-tumor immune response. With these preliminary studies in hand, we propose to 1) Dissect the roles of TERT in double strand break repair and immune evasion, and 2) Improve the drug-like properties of NU-1 and use these novel compounds to understand how best to obtain radiation sensitization and an effective anti-tumor immune response.

NIH Research Projects · FY 2025 · 2023-07

Environmental temperature dictates biology. Animals use thermal environments to guide vital behaviors such as migration, circadian rhythms, growth, and feeding. Heat is also an important influence on human health. About 1 in 100 deaths globally stem from heat-related causes, and beyond lethal heatstroke, more mild heat < 40°C can alter and dysregulate human physiology down to the cellular level, particularly in immune cells. Such sublethal heat shocks occur in hyperthermia and heat illness, as well as frequently in fever: a systemic heat shock which regulates the immune system during infection. Yet even in the well-known context of fever, we lack understanding of how human cells sense sublethal heat shock. The cell biology of extreme heat shock > 40°C is well-characterized, but far less is understood about sublethal, fever-range temperatures < 40°C. However, we do know that certain immune cells upregulate heat shock protein expression in response to fever. The induction of heat shock proteins, or the heat shock response, occurs in eukaryotes when heat activates transcription factor Hsf1, via titration of its repressor (heat shock protein Hsp70) away from Hsf1. This titration is caused by the generation of new, heat shock-induced substrates for Hsp70 to bind. These substrates, i.e., the upstream sensors of heat, are unidentified in sublethal heat shock. We hypothesize biomolecular condensates are these substrates which help cells sense sublethal heat shock. Condensation, or reorganization of proteins and RNA into larger foci, occurs in response to environmental stimuli across species from yeast to humans. Our group showed recently that heat-induced condensates are Hsp70 substrates in yeast. We hypothesize that sublethal heat shock-induced condensates are Hsp70 substrates in humans, enabling cells to sense and respond to such fever-range temperatures. It is not known what proteins condense in human cells at these temperatures, nor if such condensates might be Hsp70 substrates. Moreover, in any species, we lack molecular-scale understanding of how condensates and Hsp70 interact. We are poised to unlock exactly this knowledge using a complement of biochemical, microscopic, and molecular-level approaches. First, we will uncover protein condensation in human cell lines at fever-range temperature, using the established sedimentation-mass spectrometry method of our group. Second, we will observe directly how condensates and Hsp70 interact at the molecular scale, using single-molecule microscopy. Together, these aims will help us elucidate fundamentally how cells sense and respond to sublethal heat shock.

NIH Research Projects · FY 2025 · 2023-07

PROJECT SUMMARY The mission of The University of Chicago (UChicago) Medical Scientist Training Program (MSTP) is to train the next generation of clinician-scientists for impactful careers in the biomedical sciences. Impact is broadly defined as: providing fundamental insights into disease and relevant biological mechanisms, leading or making substantial contributions to translational efforts to identify better therapies and/or filling leadership roles that will determine how limited resources can best be used to improve human health. Our overall objective is to train outstanding and self-directed clinician scientists who can formulate and answer biologically and medically- important questions. To ensure the greatest impact, our program must reflect the diversity of the U.S. population in both who it trains and who it serves. To accomplish our mission and overall objective, we defined 15 specific and measurable objectives that stress the technical, operational and professional skills that support science identity and self-efficacy and are critical for a successful career as an independent biomedical investigator. Two unique features of the program enable our mission and provide an optimal training environment for the development of clinician-scientists. First, we are our own graduate program, the Interdisciplinary Scientist Training Program (ISTP). The ISTP allows integration of graduate and medical school classes in MS1, development of tailored curricula for each trainee, coordination across training transitions and uniform oversight and expectations during the graduate phase. Second, most trainees pursue a 1-4-3 progression of study rather than the traditional 2-4-2 progression. While trainees may pursue a 2-4-2, the 1-4-3 provides a continuous and in-depth graduate experience where trainees become well-trained scientists who can propose and rigorously test hypotheses. The 1-4-3 also provides a seamless transition between the preclinical and clinical phases which facilitates overall success in medical school. Multiple programmatic features ensure that fundamental scientific and clinical knowledge are interwoven to foster the development of impactful clinician-scientists. The UChicago MSTP enjoys sustained and substantial University financial backing including generous direct investment and support of facilities that cultivate innovative science. We are requesting 25 training slots to supplement University investments to support trainees for six of the typical eight years they spend in the MSTP. This requested investment by the NIH would constitute only 13% of total anticipated cost for our MSTP. We anticipate a program size of approximately 80 trainees and expect, based on our track record, that our trainees will contribute significantly to the U.S. biomedical enterprise.

NIH Research Projects · FY 2024 · 2023-07

PROJECT SUMMARY/ABSTRACT Inflammatory Bowel Diseases (IBD) are a growing concern in industrialized and newly developing countries as the environment, lifestyles, and diets change. IBD involves a complex interplay between the host's environment, diet, genetics, and gut microbiota, each necessary but insufficient to cause disease. However, it is unclear what initiates these diseases, what the relative contributions of these factors play in the pathogenesis of IBD, and what makes these diseases chronic. Current therapies are still imprecise, and preventative disease strategies for high-risk patients are non-existent. Some evidence suggests that changes in the gut microbiota can influence the risk and trigger IBD. Our lab uses a human model involving ulcerative colitis(UC) patients who undergo colectomy followed by creating an ileal pouch-anal anastomosis (IPAA). In theory, IPAA should be curative, but nearly half of these patients develop an inflammatory condition of the ileal pouch called pouchitis, whereas non-IBD patients undergoing the same procedure rarely develop pouchitis. These patients were followed and sampled longitudinally, and metagenomic data shows blooms of pathobionts such as Bacteroides fragilis before and during the development of pouchitis. Upon antibiotic-induced remission, the relative abundance of B. fragilis decreased, implicating it as a potential trigger and sustainer of pouchitis. The proposed research strategy tests the hypothesis that pouchitis-associated pathobionts (PAP) like Bacteroides fragilis cause in IPAA-associated pouchitis. Notably all the B. fragilis strains cultivated from UC pouchitis patients are non-enterotoxigenic strains that were present even before the IPAA was performed. Thus, we believe that insight gained through these studies will directly relate to the etiopathogenesis of human IBD. As a counterpart to the human pouchitis model, we employ germ-free (GF) and SPF (specific pathogen-free) wild-type (WT) and IL10 knockout (IL10-/-) mice where engraftment of PAP B. fragilis in the latter is associated with increased incidence and severity of colitis. I will determine whether B. fragilis promotes colitis directly and/or indirectly by causing perturbations in the membership and function of the indigenous colonic microbiota. SPF, but not GF, IL10-/- mice are genetically predisposed to developing IBD. Human PAP B. fragilis strains readily engraft into SPF IL10-/- mouse microbiota, but not that of WT mice, and promote the development of colitis. Using Bulk-RNA Sequencing analysis and flow cytometry, I will evaluate how the presence of B. fragilis alters the host's immune responses and gene expression in the GI tract of GF and SPF IL-10-/- mice. Fecal samples, regional intestinal luminal, and mucosal content will be subjected to 16S rRNA, metagenomic, and metabolomic analyses to assess if B. fragilis leads to dysbiosis–promoting chronic colitis in the IL10-/- mouse model. This study provides critical insight into the events prior to the development of chronic inflammation in an IBD-susceptible host. In addition, these studies will shed light on potential biomarkers that can predict risk, help define preventative strategies, and guide therapeutic management.

NIH Research Projects · FY 2025 · 2023-06

Randomized controlled trials are the gold standard for measuring the effect of a treatment or intervention. Unfortunately, it is not feasible to conduct a randomized controlled trial to test all research questions, whether due to cost, achieving sufficient subject sizes or when administering an arm of the trial would be unethical. To understand the effects of therapeutics, policy changes, and other interventions where it is not possible to administer a clinical trial, researchers have developed approaches that attempt to simulate clinical trials in observational data. Despite sophisticated statistical methodologies, it is not clear whether it is possible to reliably simulate a randomized controlled in observational data. We aim to quantify one potential driver of these different results, differences between the clinical trial and real-world populations. In Aim 1, we compare trials that have individual level data available to three real- world data sources. In Aim 2, we develop methodologies to infer most likely individual-level statistics from aggregate trial statistics using real world data. Finally, in Aim 3 we compare neurological trials that do not release individual level data to real world data. We then estimate the transportability of treatment estimates across different populations including: the population eligible for the trial in RWD and the population ineligible for the trial but receiving the treatment in RWD. This allows for the study of indication drift and treatment heterogeneity. By uncovering differences between these groups, we may be able to identify groups that are underrepresented in clinical trials to help reduce healthcare disparities. The K99/R00 award will allow me to gain expertise in using regulatory sciences (with mentor Dr. Florence Bourgeois and advisor Dr. Deborah Schrag) for biologic discovery (with mentors Dr. Tianxi Cai and Dr. Isaac Kohane) within neurology (with mentors Dr. Page Pennell and Dr. Clemens Scherzer). My background in statistics, informatics, genetics, and machine learning with clinical data sources ideally positions me for the proposed project. The proposed training plan, mentoring and project will provide a strong foundation for a successful transition to independent research.

NIH Research Projects · FY 2026 · 2023-06

Project Summary/Abstract This is an application for a K08 Career Development Award for Jonathan Trujillo, MD PhD, who is a clinical instructor in the Section of Hematology/Oncology at the University of Chicago. He is building a career as a physician-scientist focused on identifying immunotherapy resistance mechanisms to improve outcomes for patients with cancer. His proposed project will determine the role of tumor cell-intrinsic hypoxia-inducible factor (HIF)-1α and HIF-2α activation in mediating tumor immune evasion and immunotherapy resistance. Cancer remains the second leading cause of mortality in the United States in spite of intensive treatments. Immune checkpoint inhibitors have shown impressive and durable clinical responses in some patients, yet the majority of patients fail to respond to these immunotherapies. Emerging data indicate that increased hypoxia-induced signaling, which is mediated by the hypoxia inducible factor (HIF)-1α and HIF-2α pathways, within the tumor microenvironment is associated with reduced T cell-based inflammation and resistance to anti-PD-1 blockade. The impact of tumor cell-intrinsic HIF-1α and HIF-2α activation on the anti-tumor T cell response has yet to be determined. The overarching hypothesis of this proposal is that cancer cell-intrinsic HIF-1α or HIF-2α activation leads to defective T cell priming and ineffective T cell infiltration, thereby promoting tumor immune evasion and immunotherapy resistance. Novel genetically engineered mouse models of melanoma with conditional expression of a stabilized variant of HIF-1α or HIF-2α have been generated to determine whether cancer cell- intrinsic HIF-1α or HIF-2α activation limits the degree of T cell accumulation within tumor tissue and reduces tumor sensitivity to immune checkpoint inhibitors. The proposed studies will determine whether HIF-1α or HIF- 2α functions by impairing T cell priming and/or by limiting effector T cell infiltration and function in the tumor microenvironment. Mechanistic studies will be performed to determine whether HIF-1α or HIF-2α pathway activation results in failure to accumulate and activate dendritic cells required to generate T cell responses, failure to upregulate chemokines needed for DC or T cell recruitment, aberrant tumor vasculature which can limit T cell infiltration, or induction of immunosuppressive factors that can impair T cell responses. These data may provide rationale for the combination of novel HIF inhibitors and immune checkpoint blockade therapy. Dr. Trujillo has devised a career development plan to accomplish the following goals during this award: 1) develop expertise in immunologic and genomics techniques and mouse models of anti-tumor immunity; 2) become proficient in bioinformatics; 3) to expand knowledge in renal cell carcinoma with a focus on immunotherapeutics and novel HIF inhibitors. He has developed a strong mentoring committee led by his primary mentor Dr. Thomas Gajewski, an internationally renowned expert in cancer immunology. Thus, upon completion of this proposal, Dr. Trujillo will emerge as an independent physician-scientist focused on cancer immunology and clinical immunotherapy.

NIH Research Projects · FY 2026 · 2023-06

All (+) RNA viruses modify cytoplasmic membranes, such as the endoplasmic reticulum (ER) to establish replication compartments (RCs). These RCs are thought to form a platform for membrane- associated replicases, in addition to protecting the viral RNAs from cytosolic RIG-I-like receptors that trigger innate immune signaling and RNA-degradation machinery. We and others have shown that a key component in the viral mechanism of RC formation is the modulation of RC membrane lipid composition. We previously published that at least 3 (+) RNA virus families (Bromoviridae, Picornaviridae, and Flaviviridae) share the property of stimulating phosphatidyl choline (PC) accumulation at RCs. This suggests that understanding viral modulation of PC synthesis may have broad implication as a conserved mechanism in RC formation. We have extended this observation to gain significant mechanistic insight into this process. The specific aims are: Aim 1. Define the contribution of PC synthesis for viral replication. We hypothesize the activation of PC synthesis aids the formation of viral RCs and may also impact virion infectivity via altered ER lipid composition. Aim 2. Define the mechanism by which HCV modulates PC synthesis. Aim 3. Define the significance of the ASCL enzymes in HCV replication. ASCLs localize to RCs and are required for HCV replication. We hypothesize that a viral protein recruits them to RCs and that they are required to provide the long chain fatty acids for phospholipids, such as PI and PC.

NIH Research Projects · FY 2026 · 2023-06

PROJECT SUMMARY Across many species, the evolutionary processes that underlie genetic variation are structured by the geographical distribution of individuals and how geography impacts patterns of reproduction and dispersal. In human genetics, geographic patterning in allele frequencies has important practical consequences for genome-wide association studies, as it can produce a type of confounding with other spatially varying factors impacting traits. It also has relevance for the discovery of rare loss-of-function variant carriers. In infectious disease, the arrival and spread of novel adaptive variants is mediated by geographic dispersal patterns. In this project, we will develop theory, methods, and resources that incorporate an explicitly geographic component. In the first research area, we will develop new theoretical models for investigating the impacts of varying spatial sampling strategies on the detection of deleterious alleles, such as loss-of-function alleles. We will also study the spread of advantageous alleles in populations with super-spreaders and long-distance dispersal, as well as graph-based dispersal dynamics, in a series of analyses that is relevant for understanding the spread of adaptive variants in human-dispersed pathogens. In the second research area, we will expand a set of methods for understanding spatial structure in genetic data. Specifically, we hope to build models that more accurately capture directional and long-distance migration. In the third research area, we will continue to maintain and develop resources for visualizing geographic distributions in allele frequencies. These three research areas are synergistic and ideally will help advance our understanding of genetic variation in numerous species, including humans.

NIH Research Projects · FY 2026 · 2023-06

PROJECT SUMMARY/ABSTRACT Patients with cancer represent one of the most vulnerable populations in our healthcare system. Not only do such individuals face a life-threatening diagnosis, but the toxicities of treatment regimens place cancer patients at additional risk for adverse outcomes. Additionally, most cancer patients experience inadequately treated pain, despite pharmacologic interventions by clinicians to address pain. The use of germline pharmacogenomic information offers a potential solution to better inform therapeutic decision-making. Many consider it an ideal discipline by which to advance and examine the clinical application of genomic medicine. Nevertheless, preemptive germline pharmacogenomic testing does not currently constitute the standard of care before utilization of most anti-cancer therapies, nor for pain prescribing. One of the most frequently cited reasons is the lack of prospective randomized data demonstrating its utility. We posit that preemptive genotyping and the upfront use of pharmacogenomic information offers the potential to improve cancer care. We hypothesize that providing germline pharmacogenomic information along with clinical decision support will guide personalized chemotherapy and pain medication choices and dosing decisions, reduce toxicity, and result in improved patient outcomes. We propose to leverage our existing institutional infrastructure and two strengths of the University of Chicago (pharmacogenomics, oncology) by performing the first known broad prospective randomized studies of germline pharmacogenomics in oncology. We will recruit patients preparing to initiate cancer chemotherapy or pain treatment who have one of three major cancer types (gastrointestinal, head and neck, or breast malignancies), and randomize to upfront pharmacogenomic testing versus no upfront pharmacogenomic information (usual care). Chemotherapy dosing and the incidence of severe toxicities, as well as opioid selection for pain treatment and pain medication response will be compared between arms. Additionally, we will evaluate whether sharing of pharmacogenomic results with patients in an informed manner improves patients’ knowledge about and perceptions of treatment choices and alignment with treatment goals. In summary, this proposal will examine and advance the idea that widespread efficacious clinical translation of genomic discoveries will be actualized both by systems/technology changes as well as the strategic assessment of genomic knowledge applications within key clinical settings and stakeholder populations. Future effectiveness and impact on public health will result not just from the findings we generate but from an improved understanding of decision-making processes involved in promoting and adopting risk- reductive, customized treatment practices.

NIH Research Projects · FY 2025 · 2023-06

Project Summary: New approaches, such as the emergence of immune checkpoint blockade over the past decade, hold great promise for patients with metastatic cancer. Indeed, PD-1/PD-L1 blockade has enjoyed remarkable clinical success for immunogenically “hot” tumors such as subsets of melanoma and non-small cell lung cancer patients; however, for patients with immunologically “cold” tumors, such as most advanced colorectal cancers, patient response rates can be as low as 5%. With the advent of whole genome sequencing and sophisticated bioinformatics techniques, patient-specific neoantigens in tumors can now be identified and provide the basis for many therapeutic vaccines in preclinical and clinical development. Neoantigen-based cancer vaccines can be tailored to individual patients by identifying their unique neoantigens. We have pioneered the development of nanoscale coordination polymers (NCPs), which are a class of hybrid nanoparticles formed by the self-assembly of metal ions and polydentate bridging ligands. NCPs preferentially accumulate in tumor tissues by taking advantage of the enhanced permeability and retention effect and possess several advantages over existing nanocarriers. The long-term goal of our collaborative research is to establish a new treatment paradigm for metastatic colorectal cancer, through the development and characterization of effective NCPs that can be delivered systemically. The overall goal of the proposed studies is to develop robust NCPs, namely, ZnCDN, for the systemic delivery of a neoantigen and hydrophilic cyclic dinucleotide (CDN) STING agonists, including CDA, ADU-S100, and MK-1454, to potentiate the antitumor immune effect of anti-PD-L1 immunotherapy for the effective treatment of mCRC. Increased understanding of the mechanisms involved in this combination therapy will provide critical insights to enhance the response rates and durability of immunotherapies for mCRC. We have designed ZnCDN NCP with a core of CDA and Zn2+ ions and a hydrophilic shell of PEG2000 to resist plasma protein absorption and clearance by the monocytic phagocytic system. As a result, ZnCDN can be administered to mice via intravenous injection to significantly accumulate in the TME. We will study three ZnCDN formulations with CDA, ADU-S100, and MK-1454 to obtain the best ZnCDN and evaluate its effects on tumor vasculature and intratumoal retention. We will evaluate immune activation in mouse models of mCRC, and elucidate the mechanisms of ZnCDN-mediated STING activation in the tumor microenvironment. In order to evaluate the delivery of tumor-specific antigens, we will incorporate tumor antigen peptides into NCPs to facilitate cross-priming of CD8+ T cells and enhance antitumor efficacy. Finally, we will elucidate the mechanism of STING activation with ZnCDN-antigen to overcome resistance to PD-1/PD-L1 blockade. Our labs have been working together on this project in the Ludwig Center for Metastasis Research for most of the past decade. This interdisciplinary endeavor could lead to a transformation in the treatment of mCRC.

NIH Research Projects · FY 2026 · 2023-06

PROJECT SUMMARY/ABSTRACT My group develops and uses chemical and biochemical tools to investigate the structure and function of noncoding RNAs (ncRNA) and their complexes with proteins (RNPs) and small molecules (smRNAs). In the context of this project, we will specifically focus on two general areas: 1) engineering fragment antigen binding (Fabs) as chaperones for RNA/RNP/smRNA crystallography, cryoelectron microscopy (cryoEM) and as enabling tools for RNA biology, and 2) elaborating mechanisms of RNA function, especially catalytic RNAs (ribozymes). The terrestrial transcriptome contains vast numbers of structured RNAs within both coding and noncoding transcripts that act alone and in concert with proteins to carry out critically important roles in nearly every facet of cellular function. Understanding how these RNAs mediate biological function in health and disease requires knowledge of their three-dimensional architectures. We will deploy a variety of structural biology, biophysical, biochemical, computational and chemical approaches, including kinetic isotope effect analysis and the use of nucleotides from an artificially expanded genetic system. The data that emerge from our recombinant Fab work will engender a robust platform to generate Fabs that bind structured RNA with high affinity and specificity, establish Fabs as powerful reagents in structural and mechanistic RNA biology and yield new macromolecular structures, which have profound impact on our understanding of the underlying biology and serve as a framework to develop and test functional hypotheses. In addition, our in-depth work on catalytic RNA will define for the first time the relationship between catalytic structure and transition state structure, a missing link that has plagued efforts to engineer macromoleclules with novel catalytic functions. Beyond the conceptual advances that emerge, this work promises to deliver to the RNA community powerful Fab reagents for RNA/RNP structural and functional studies, nucleotide analogs to support novel approaches to define RNA catalytic mechanisms, and strategies to render large RNAs more tractable by increasing the density of information.

NIH Research Projects · FY 2026 · 2023-06

Chronic pain conditions plague more than 20% of adults in the United States, emphasizing the need for a more comprehensive understanding of pain control mechanisms and identification of better pain therapies. This need is amplified further by the current opioid crisis, which killed over 60,000 Americans in 2020. While opioids are remarkably effective treatments for acute and chronic pain, adverse side effects, including abuse liability and tolerance to the analgesic effects with repeated use, highlight the need for non-opioid pain therapies. This need motivates our investigations of pain modulation by acetylcholine (ACh) and its receptors. Ascending nociceptive signaling from periphery to central nervous system is modulated by the descending pain modulatory pathway, including the ventrolateral periaqueductal grey (vlPAG) and its projections to rostral ventromedial medulla (RVM). This pathway is a crucial site of action of opioids and endogenous pain control. While much is known about this circuitry, the impact of neuromodulators like ACh on this circuit, particularly under chronic pain conditions have not been investigated. Using a fluorescent ACh sensor, we will investigate how pain and other behaviorally relevant stimuli alter ACh release dynamics in the vlPAG. Using brain slice electrophysiology, we will identify the cellular mechanisms behind alterations in ACh release. And finally, we will attempt to reverse these maladaptive ACh dynamics using chemogenetics and pharmacological approaches. Our preliminary data show that cholinergic projections from the pedunculopontine tegmentum (PPTg), modulate excitability of vlPAG neurons. Under chronic pain conditions, we noted a decrease in ACh release in vlPAG and reduced neuronal activity in PPTg. We also found that M2 muscarinic AChRs are expressed on vlPAG neurons, and that pain increases the activity of these neurons. These observations lead us to hypothesize that chronic pain lowers cholinergic signaling from PPTg and reduces M2 activity in vlPAG, contributing to chronic pain symptoms. We further hypothesize that reversing these changes will relieve somatic and affective symptoms of chronic pain. We will use an ACh sensor with in vivo fiber photometry to assess the impact of acute and chronic pain on ACh release in the vlPAG. We will complement these assays using in vivo microdialysis for ACh. Brain slice electrophysiology will be used to explore the pain-induced changes in synaptic drive and intrinsic excitability of PPTg cholinergic neurons that project to vlPAG. In vivo imaging will be used to assess chronic pain induced changes in vlPAG neuronal activity. Then, using chemogenetics and M2 mAChR agonists we will attempt to reverse the pain-induced changes in excitability of vlPAG. Behavioral testing will confirm reversal of the somatic and affective components of chronic pain. Better understanding of these modulatory inputs to descending pain pathways will help identify novel targets for treating chronic pain.

NIH Research Projects · FY 2026 · 2023-06

Project Summary Genetic deletion (haploinsufficiency) or duplication events are often associated with a variety of diseases, including: cancers, cardiovascular and metabolic disease, and neuropathies. Because many genes are sensitive to both over- and under-expression, with imbalances in gene product in either direction leading to disease, tight regulatory control is a therapeutic necessity. RNA-targeting technologies, however, provide a mechanism to appropriately and reversibly modulate gene expression at the transcript level, resulting in tunable, cell-specific remediation of disease-causing shifts in gene dosage. Our recently developed CRISPR- Cas- inspired RNA targeting system (CIRTS), integrates small, human-derived proteins and guide RNA into an easily programmed and packaged technology for tunable gene expression manipulation at the transcript level. Notably, CIRTS imparts a major clinical advantage over that of functionally-related, but bacteria-derived CRISPR/Cas systems: minimized immunogenic risk stemming from the use of human protein parts. This proposal seeks to use protein engineering and cell-based screens to create highly efficient, optimized CIRTS- based technologies – both for gene activation and deactivation – and assess their generality across a panel of disease-relevant targets, and to test the potential of the technology in vivo. Initial screening will focus on the gene PMP22, an exemplar gene for which both over- and under-activation result in distinct, but related, genetic disorders. This provides a fertile testbed for optimization of both CIRTS degraders (Aim 1) and activators (Aim 2). In Aim 1, the human proteome will be mined to identify and characterize functional domains that can be integrated into CIRTS to programmably degrade PMP22 RNA. How target RNA landing sites correlate with the functional outcomes of each successful design, as well as generality against another target, SNCA, will be simultaneously pursued. Aim 2 of the proposal focuses on developing CIRTS activators, mining the human proteome for novel functional domains and using novel reporter systems to efficiently screen and characterize them, and finally, assessing lead designs in two other target genes, SCN1a (brain) and JAG1 (liver). Aim 3 will assess selectivity and specificity constraints of the system, and the generality of the CIRTS technology across a range of biological contexts. Here, lead CIRTS architectures will be benchmarked against state-of-the-art Cas13-based systems, comparing immunogenicity in vivo and testing the efficacy of CIRTS-based regulatory systems in “real world” contexts, namely mouse models of PMP22-based gene dosing disorders. Critically, data from all three aims feed off of and inform one another, resulting in a tripartite design-build-test optimization cycle. Completion of this project will generate a multi-modal, programmable CIRTS toolkit for the manipulation of gene expression at the RNA level. In addition to providing proof-of-principle validation of this system’s potential in one exemplar clinical application, this work will also produce a unique suite RNA-targeting effectors with broad utility across a wide variety of biomedical research and synthetic biology fields.

NIH Research Projects · FY 2026 · 2023-05

Project Summary Dynamic mRNA modifications, such as the m6A-dependent regulation at the mRNA level, add a critical new dimension to post-transcriptional regulation of gene expression. The rapid development of sequencing technologies has transformed the field of epitranscriptomics studies by resulting in the successful profiling transcriptome-wide RNA modifications under different states and conditions. They hold the promise to reveal regulatory machinery of RNA modifications, which contributes to almost every phase of mRNA metabolism and function, thereby impacting diverse biological processes. How- ever, analytical developments in epitranscriptomics lag far behind the pace of technological discovery, and the bioinformatic infrastructure available for epitranscriptomic studies remains limited. The overar- ching goal of this proposal is to address three most pressing challenges facing profiling and interpreting epitranscriptomics. Specifically, we will achieve the following aims: Aim1. Develop statistical methods for RNA modification detection at single nucleotide resolution. Aim 2. Develop computational meth- ods for cell type-specific methylation analysis. Aim 3. Develop web servers that enable integrating RNA modification with a rich catalog of genomics features. All the methods will be implemented in user-friendly software and disseminated to the scientific community. Successful achievement of all aims will dramatically increase the power of epitranscriptomes analysis, leading to better understand- ing of regulatory mechanisms in RNA modifications and their implications in phenotypes and human diseases.

NIH Research Projects · FY 2025 · 2023-05

PROJECT SUMMARY Outcomes for patients with relapsed/refractory diffuse large B cell lymphoma (DLBCL) remain poor despite recent therapeutic advances, particularly in the area of immunotherapy. Classification of solid tumor microenvironments as immune “inflamed” or “non-inflamed” through bulk transcriptional profiling enriches for a subset that is checkpoint blockade therapy (CBT) responsive. Conversely, the DLBCL immune landscape has not been as well-characterized, and the extent to which immune environmental features can predict for immunotherapy response in DLBCL patients is unknown. Given that a growing number of immunotherapies are being explored in the relapsed/refractory DLBCL space, a deeper understanding of the DLBCL immune landscape might uncover clues that aid in identifying patients likely to benefit from checkpoint blockade and/or CAR T cell therapies. Additionally, growing evidence indicates that cancer cell-intrinsic alterations can profoundly affect the tumor immune environment, which directly impacts immunotherapy sensitivity. How recurring genetic alterations and related pathways in malignant B cells contribute to shaping the DLBCL immune environment is unclear. Additional research is clearly needed in order to address these gaps in knowledge. Toward that end, we incorporated curated immune- and cell-of-origin (COO)-related gene sets into a gene set variation analysis (GSVA) on transcriptomes of 874 DLBCL specimens. Among the four clusters that emerged (germinal center B cell (GCB) hot, GCB cold, activated B cell (ABC) hot, and ABC cold), analysis of whole exome sequencing data revealed significantly enriched genetic alterations in each. For instance, loss- of-function (LOF) mutations in SOCS1, a negative regulator of JAK/STAT signaling, were enriched in GCB hot DLBCLs, suggesting these lymphomas may be particularly sensitive to IFNγ signaling and vulnerable to anti- PD-1 therapy. Conversely, LOF alterations in TMEM30A, which regulates phosphatidylserine (PS) orientation in the plasma membrane, were common among ABC cold DLBCLs, which may render these lymphomas sensitive to immunotherapies that enhance macrophage phagocytosis. These observations support the central hypothesis that lymphoma cell-intrinsic mechanisms contribute significantly to shaping unique DLBCL immune environments, the characterization of which will identify patients who will or will not benefit from CBT or CAR T cell therapy. The main objectives of the proposal are: 1) to determine mechanisms by which select genetic alterations in lymphoma cells shape the DLBCL immune environment, and 2) to develop a DLBCL “immune score” and determine its utility in identifying patients for whom PD-1 blockade or CAR T cell therapy will be effective. The principal investigator, a physician-scientist with clinical expertise in lymphoma and a research background in cancer immunology, is well-suited to oversee the experiments proposed in this application. In order to execute computational aspects of the proposal, the principal investigator has recruited a rising star in the field. Together, their complementary skills will ensure successful completion of the planned research.

NIH Research Projects · FY 2025 · 2023-05

PROJECT SUMMARY Anxiety and depression are highly debilitating mental health disorders with origins in early life, making research in children and adolescents a critical public health need. The prevalence of both disorders in this age group has rapidly increased over the past decade, particularly in urban areas. Though the etiologies of child and adolescent mental health disorders remain poorly understood, increasing trends over mere decades point to environmental causes more than genetics. We propose that non-chemical stressors (noise, violence, negative life events, and neighborhood environment) in the urban environment play a major role and interact with environmental factors that show strong urban-rural gradients, such as air pollution. The neurocognitive toxicity of air pollution has been intensely studied in animal and epidemiologic research but its role in anxiety and depression is poorly understood, with only a few studies in adults, and even less in younger populations. Further, the mechanisms by which air pollution impacts the brain are poorly understood, although altered functioning of the hypothalamic-pituitary-adrenal (HPA) axis and its role in regulating cortisol secretion is a prime candidate. This study will determine whether early life PM2.5 and non- chemical stressors impact symptoms of depression and anxiety in preadolescence/ late childhood and whether cortisol mediates and/or modifies these relationships. We will leverage resources from an established longitudinal birth cohort in Mexico City - Programming Research in Obesity, Growth, Environment and Social Stress (PROGRESS). Specifically, we will examine time-specific and cumulative PM2.5 exposure in relation to mental health outcomes in 8-11 year olds (Aim 1), the role of individual and combined urban non-chemical stressors in relation to mental health symptoms (Aim 2), and the role of hair cortisol levels on the biological pathway from PM2.5 to mental health symptoms (Aim 3). In order to more comprehensively characterize urban stressors in environmental epidemiology and assess their impacts on mental health in preadolescents, I will cross-train in child and adolescent psychopathology and enhance my skills in geospatial modeling. I will additionally train in advanced statistical mixtures and causal mediation to better characterize biological pathways from PM2.5 and stress to mental health outcomes. I will develop these skills through didactic training, independent study, and mentorship from experts in developmental psychology, pediatrics, geography, social epidemiology, and biostatistics, specifically - Drs. Rosalind Wright, Robert Wright, Itai Kloog, Daniel Klein, and Brent Coull. At the end of this training period, I will be uniquely positioned to more comprehensively examine the effectsof multiple urban stressors on mental health outcomes in future research. Further, I will use the knowledge gained and the noise model I develop in future grants, setting the stage for my long-term goal of studying the effects of chemical and non-chemical stressors in the urban environment on brain development and mental health outcomes across the life course.

NIH Research Projects · FY 2025 · 2023-05

PROJECT SUMMARY/ABSTRACT Humans have highly advanced cognitive abilities and motor skills, characteristics which are reflected in the enlarged size and cell diversity of our central nervous system (CNS). My overall goal is to profile and compare progenitor cell diversity in humans, non-human primates and rodents, and thereby identify the origins of increased cell diversity and size of the human CNS. As part of my postdoctoral research, I collected and analyzed high-temporal-resolution single-cell RNA-seq data of the well characterized human and mouse spinal motor neuron (MN) lineage, which led to the identification of a molecularly distinct, human-specific (i.e., not found in mouse) motor neuron progenitor (hsPMN) cell type. I found that hsPMNs undergo delayed and protracted neurogenesis, increasing total MN output by ~2-fold. The proposed study aims to characterize how they contribute to the population size and subtype diversity of the MN lineage by combining single-cell RNA-seq with long-term astrocyte co-culture or xeno-transplantation of lineage-labeled human cells. Second, the proposed study will further investigate the evolution of hsPMN cells and their gene expression program by a) determining whether an orthologous cell type is found in old-world monkeys (i.e., macaque), and b) identifying and functionally testing newly evolved gene regulatory elements that give rise to hsPMN-specific gene expression patterns. Finally, I will develop a 3D spinal organoid in vitro differentiation system that harbors spatially patterned ventral and dorsal spinal lineages, using a standard set of differentiation conditions that can be applied to human, macaque, and mouse cells. This last aim will broaden the scope of comparative investigations to encompass multiple spinal lineages, thus opening up new avenues and hypotheses for future research, as well as providing a more comprehensive in vitro model system of spinal cord development. The insights, protocols, and data generated from this study will enable and provide valuable comparative analyses into human neural progenitor diversity and how human-specific progenitors contribute to the size and cell diversity of the CNS during development. Furthermore, these studies and data will form the foundation of my research agenda as an independent investigator, while allow me to broaden my research training to include a diverse host of species, model systems and a wide array of experimental as well as computational techniques.

Fonds de recherche du Québec – Société et culture · FY 2023-2024 · 2023-04

Volet: Bourses postdoctorales; Domaine: Arts, littérature et société; Objet: Systèmes religieux; Objet: Droit canonique; Application: Structures et relations sociales; Application: Droits et justice; Mots-clés: LEGAL TRADITION, CHURCH OF THE EAST, SYRIAC CHRISTIANITY, HISTORY OF LAW IN THE MIDDLE EAST, JEWISH LAW, ZOROASTRIAN LAW

NIH Research Projects · FY 2026 · 2023-04

PROJECT SUMMARY Synaptic transmission and plasticity are fundamental to neuronal functions, and dysregulations of synaptic protein expression are direct causes of neurodevelopmental disorders such as autism. Over one hundred de novo loss-of-function mutations in SYNGAP1 have been unambiguously associated with autism spectral disorders and intellectual disability. Recent success in splice-switching oligonucleotides (SSOs) suggests that redirecting splicing through genetic and SSO-mediated ablations is a promising approach to rescue haploinsufficiency. We have identified an alternative splicing event in SYNGAP1 that leads to nonsense- mediated mRNA decay (NMD) during mouse and human development. To determine whether the SYNGAP1 NMD exon is a viable therapeutic target, we investigate the regulatory mechanism and its functions using genetic approaches, and determine whether genetic deletion and SSO suppression of the SYNGAP1 NMD exon can rescue heterozygous knockout phenotypes in mouse mutants and patient-iPSC-derived neurons. Upon completion, this project will provide genetic insights into the physiological functions of this SYNGAP1 NMD exon and generate critical preclinical reagents to restore SYNGAP1 protein expression from haploinsufficiency.

NIH Research Projects · FY 2026 · 2023-04

When a US hospital system is overwhelmed by disaster, Crisis Standards of Care guide the triage teams forced to choose which patients receive scarce life support treatments. Analogous to an organ allocation system, these algorithms convert ethical principles into a concrete rank ordering of candidates for Intensive Care Unit (ICU) treatments with life support allocation scores. Disasters that produce scarcity tend to fall hardest on specific geographical areas and communities. When designing algorithms to allocate scarce life support, public health officials should take this context into account. In an attempt to identify the critically ill patients with the highest likelihood of benefit from treatment, most US states would prioritize those with low Sequential Organ Failure Assessment (SOFA) scores. But SOFA was designed for patients already on life support in the ICU, using routinely measured laboratory values, drug doses, and vital signs to monitor response to treatment. Most patients have low SOFA scores when critical illness is first recognized, and SOFA cannot accurately predict the risk of death using data before life support was allocated. We demonstrated how the poor predictive performance of SOFA-based triage protocols is partially explained by a miscalibrated renal component of the SOFA score. There is a clear need to develop and validate a novel life support allocation protocol designed to debias existing scores and save more lives. Place-based disadvantage indices, such as the Area Deprivation Index (ADI) and the Social Vulnerability Index, offer a potential solution. Using these validated geographical measures of neighborhood deprivation to allocate scarce healthcare resources counteracts the geographic concentration of the disaster. We hypothesize that a well-designed life support allocation score using place-based disadvantage indices can save more lives. The overall objective of this project is to develop a life support allocation algorithm that accurately and equitably allocates scarce ICU treatments in a crisis. In Aim 1, we will use structural equation modeling to create an Equitable Life Support Allocation (ELSA) score, using place-based disadvantage indices to debias SOFA. In Aim 2, develop the ICU Crisis Simulation Model (ICSM), a discrete event simulation that models patient flow and survival, as a testing and evaluation environment for life support allocation protocols. In Aim 3, we will externally validate ELSA and ICSM in the National COVID Cohort Collaborative Data Enclave, which currently contains geocoded records from 14 million patients from 74 sites. Our project will address one of the most pressing challenges in applied public health ethics, producing 1) an empirically derived score to distribute life support more accurately and equitably in a crisis and 2) open-source simulation software to evaluate the efficiency and equity of life support allocation protocols.

NIH Research Projects · FY 2026 · 2023-04

PROJECT SUMMARY Mucosal healing is the primary goal of all therapies for inflammatory bowel disease (IBD). No currently approved therapy directly promotes wound healing of the bowel epithelium, which forms a single-cell barrier between the host and lumen and regulates the immune response. The cellular and molecular mechanisms that define the healing process remain to be defined. The intestinal crypt maintains the normal turnover of epithelial cells within a vertical lineage hierarchy of basal stem and terminally differentiated luminal cells. During wound healing, this lineage hierarchy is suspended, and epithelial cells can undergo dedifferentiation to mediate re- epithelialization. In recent work, we have identified an alternate source of wound healing in the distal colon of mice. In acute and chronic models of colitis, a skin-like cell population at the anal transition zone (ATZ), bordering the sharp squamocolumnar anorectal junction, migrates into the colon and forms a permanent hybrid epithelial structure called squamous neo-epithelium of colon (SNEC). SNEC represents the end-product of wound healing by cells of the ATZ. The ATZ is an anatomically small region that is composed of a unique population of epithelial stem cells that have a mixed colonic/epidermal phenotype and are capable of wound- healing plasticity. Moreover, these stem cells are partially resistant to the damage of colitis. The study of these cells could highlight new pathways for colonic epithelial regeneration in the context of IBD. However, the lineage diversity and relationships within the myriad tissue types of the human ATZ are not known, and furthermore it remains to be defined whether ATZ stem cells might provide enduring intestinal function, protect from potential oncogenic sequela, or repair ulcers throughout the colon. In the proposed work, we will test whether ATZ-derived stem cells could be suitable reagents to mediate colonic epithelial wound healing and identify key pathways that contribute to their proliferative potential. The Specific Aims of the project are: 1) to define regenerative cell populations in the human ATZ, 2) to define long-term outcomes of endogenous colonic wound healing by ATZ cells, and 3) to identify mechanisms of functional ATZ plasticity in wound healing. A deeper investigation of the mechanisms, implications, and potential translation of this noncanonical form of colonic wound healing could reveal new therapeutic targets to direct mucosal healing in IBD.

NIH Research Projects · FY 2026 · 2023-04

ABSTRACT Diabetes mellitus (DM) affects 30 million people in the U.S. African-Americans and Hispanics are 1.4 and 1.2 times more likely to have DM compared to non-Hispanic whites. Diabetes distress—stress, fear, and guilt related to managing diabetes—is linked to poor glycemic control and disproportionately affects African Americans and Hispanic adults with type 2 diabetes mellitus (T2DM). The American Diabetes Association (ADA) has published guidelines promoting screening for and addressing diabetes distress (DD) as a critical part of clinical care. However, only 24% of adults with diabetes report their health care team asked them how diabetes affected their lives. Efforts to systematically identify and address DD could be an important strategy to improve diabetes outcomes among disadvantaged populations and address diabetes disparities. Community health centers (CHCs) can be important partners in this effort; CHCs provide primary care for 2.5 million adults with diabetes. More than 70% of CHC patients have income below 100% of the federal poverty level and 57% are people of color. No studies have systematically implemented DD screening and treatment interventions into a real-world primary care setting or used a guideline based approach. To fill this gap, we developed the ARISE (Achieving Routine Intervention and Screening for Emotional health) intervention. Based on published guidelines, ARISE incorporates validated screening instruments, draws from interventions shown to improve DD and is individualized to patients’ domains of DD. ARISE includes a standardized process for screening adult patients with T2DM for DD, training for health center staff on how to address distress in the patient encounter, and an algorithm for action steps and referrals based on the domains identified as sources of distress. This study aims to compare ARISE to enhanced usual care (didactic training for health care teams on DD) in CHCs using a type I hybrid effectiveness-implementation design via a cluster randomized pragmatic trial. First, we will adapt ARISE into clinical workflows in two CHCs (one urban and one rural) using the Form and Function domains of the Complex Health Intervention Framework. Using the lessons learned from the adaptation, we will conduct a cluster randomized pragmatic trial across 12 CHCs (six ARISE and six enhanced usual care) to test ARISE vs. enhanced usual care among adult patients with T2DM and A1c>8%. Primary outcome will be change in A1C from baseline to 12-months between arms. Secondary outcomes will include change in DD from baseline to 6-months within the ARISE arm and change in patients’ systolic blood pressure, low density lipoprotein (LDL), and body mass index (BMI) across the two arms. We will assess the adoption, implementation, and maintenance of the ARISE intervention. We will use knowledge gained to develop best practices for CHCs across the country that are charged with caring for the largest share of America’s medically vulnerable patients with T2DM.

Fonds de recherche du Québec – Société et culture · FY 2023-2024 · 2023-04

Volet: Bourses de doctorat en recherche; Domaine: Développement et fonctionnement des personnes et des communautés, et vie sociale; Objet: Rôles familiaux; Objet: Économie familiale; Application: Structures et relations sociales; Application: Populations; Mots-clés: SIGNIFICATION SOCIALE DE L'ARGENT, ROLES DE GENRE, PLANIFICATION FINANCIERE, TRANSMISSIONS INTERGENERATIONNELLES, DROIT DE LA FAMILLE, HERITAGE

NIH Research Projects · FY 2026 · 2023-04

Metabolism is required for life and as such impinges on almost every physiological and disease state. Currently, there is a strong need for training in metabolism-based research, as we face alarming increases in type 2 diabetes and metabolic syndrome stemming from the obesity epidemic, as well as rising rates of type 1 diabetes. Now, there is a realization that regulation of metabolism is much more complex when considered at the molecular, cellular and/or systemic levels. Yet, despite this gap in knowledge, there are relatively few predoctoral training programs specializing in the study of the metabolic and molecular basis of type 1 and type 2 diabetes, obesity and other metabolic diseases such as the proposed Molecular Metabolism Training Program (MMTP). There is a need to bring through the next generation of metabolism researchers, so that these diseases can be better understood, alleviated, treated, and even prevented. The overall goal of the MMTP is to harness unique training capabilities at the University of Chicago through the graduate Committee system, to identify and train motivated, creative and thoughtful predoctoral students from a variety of biological backgrounds to receive intensive training to better understand and address the causes of metabolic diseases. The MMTP will be administratively housed in the established PhD granting Committee on Molecular Metabolism and Nutrition (CMMN), but will be able to draw from multiple other graduate programs throughout the Biological Sciences Division (BSD) with a common and sole focus on the study of diabetes, obesity and metabolic disease. The MMTP will play a critical part in the orchestra training predoctoral students throughout the BSD at the University of Chicago to become the next generation of metabolic researchers.