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Abstract or Project Description

Inappropriate and chronic inflammatory responses underly the development of Inflammatory Bowel Diseases (IBD), including both ulcerative colitis and Crohn’s disease. This process is an intertwined, inter-cellular cascade among diverse immune cells and tissue stroma. The interactions between immune and stromal cells not only propagate inflammatory responses but also regulate resolution of inflammation. Yet, we do not understand how immune cells interact with tissue stroma when intestinal inflammation occurs and resolve. We propose to first develop cutting-edge technologies that discover interacting partners of immune cells in vivo in a high-throughput and unbiased manner. We will then apply these new tools to identify the interactomes of immune cells over the course of intestinal inflammation. Funding from the Kenneth Rainin Foundation will support our program to map out the path of immune-stromal interactions from a healthy intestine to an inflamed tissue to recovery and repair. Targeting immune-stromal interactions that cause inflammatory propagation as well as the resolution will establish a new strategy to alleviate inflammation. Our long-term goal is to leverage this knowledge to switch “inflammatory interactions” to “resolving interactions” in order to restore intestinal homeostasis, which will ultimately curtail IBD and its devastating health implications.

Abstract or Project Description

The epithelial cells lining the intestine control responses to gut bacteria, limiting intestinal damage in Crohn’s disease. A critical gene in this pathway is DUOX2, which is up-regulated in response to bacteria, and helps maintain intestinal health.  We discovered that Crohn’s disease patients who carry genetic variants in DUOX2 are more likely to progress to fibrotic disease complications requiring surgery. We tested an intestinal gene signature for refractory pediatric Crohn’s disease in a database containing 2000 small molecules, identifying 140 which might be more effective therapies. Our studies will now utilize a novel Human Intestinal Organoid, or “mini-gut” model system for refractory pediatric Crohn’s disease that we have developed informed by genetic factors like DUOX2. We have prepared stem cells from pediatric Crohn’s disease patients who did or did not have DUOX2 genetic variants, and will differentiate them into white blood cells and human intestinal organoids. This novel model system will be used to test ways to modulate pathways linked to fibrosis in patients, by screening the top small molecules which we have identified as potentially beneficial. Our studies will advance precision medicine, by developing a platform to screen new therapies in patient-derived intestinal organoids with clinically important genetic variants.

Abstract or Project Description

NOD2 was the first and to date the most frequent gene associated with Inflammatory Bowel Disease (IBD). The NOD2 protein functions as a sensor of bacteria by binding molecules (peptidoglycans) shed from the bacterial cell wall. The IBD mutations in NOD2 result in its inactivation or partial loss of function. In 2019 we reported that NOD2 requires the attachment of 2 fatty acids to be functional. This modification termed palmitoylation is mediated by the protein zDHHC5 and is missing from many of the mutant forms of NOD2 associated with Crohn’s disease. Palmitoylation is a reversible modification and while zDHHC5 is the “writer” that adds the modification, the “eraser” that removes it has not been identified. We believe that by blocking the eraser that we can restore function to at least some of the NOD2 mutants. Our unpublished work has identified the relevant acyl thioesterase which removes palmitate and inactivates NOD2. Through the use of chemical inhibitors we are able to rescue the function of several NOD2 mutant proteins. This proposal represents the next logical steps in our goal to characterize the relevant acyl thioesterase and to determine if any FDA approved drugs can restore function to NOD2 by inhibiting this eraser.

Abstract or Project Description

Our gut microbiome has a strong influence on a variety of diseases, including Inflammatory Bowel Disease (IBD). Fungi form an integral part the human gut microbiome, and studies in mice showed that fungi may have a substantial influence on inflammation and IBD development, while antibody responses against fungi constitute important clinical IBD diagnostic tools. However, the functional roles of fungi in modulating IBD, and whether they can be harnessed towards new treatments, remain elusive to date. In this study, we plan to uncover the role of fungi in IBD by taking biopsy samples of the gastrointestinal tract from IBD patients. From the biopsies, we plan to grow mini-guts, called organoids, in petri dishes. We have previously shown that it is possible to use these organoids cultured with microbes to study the interactions between bacteria and the gut cells. Thus, using IBD patient-derived gut organoids and specific IBD-relevant fungi, we will be able to identify mechanisms of the interactions between the diseased gut cells and the fungi. Interesting fungal candidates will be further validated in IBD-like mouse models. This combination of different approaches will allow us to better understand the influence of fungi on IBD progression and develop new intervention strategies for treating IBD.

Abstract or Project Description

Besides the brain, the gastrointestinal tract has the highest number of neurons in the entire body. The enteric nervous system (ENS) is often referred to as the second brain as it commands all manner of intestinal function, from nutrient absorption to defecation control. In patients suffering from Inflammatory Bowel Disease (IBD), the ENS undergoes major disruption resulting in many debilitating complications impacting the quality of life. We have recently uncovered that the ENS also controls intestinal inflammation and regulates the balance of “good” vs. “bad” bacteria in the gut. As the ENS has the unique capacity to rapidly relay signals from the gut directly to the brain and, by extension, to all other organs, we hypothesized that ENS might act as front-line sensors of gut inflammation to warn the body of impending infections. Our preliminary data indicate that when intestinal neurons are activated, they can trigger inflammation in other organs to protect against bacterial invasion. This novel protective mechanism has never been reported before and in this project, we aim to elucidate this unknown pathway of inflammation. Going forward, we will further explore if this axis is aberrantly activated during chronic intestinal inflammation and if it directly contributes to non-intestinal inflammatory complications exhibited in patients with IBD to aid in novel therapeutic design.

Abstract or Project Description

Ulcerative colitis (UC) is characterized by non-resolving pathological inflammation leading to erosion of the colon tissue. Recent studies analyzing gene expression signatures in UC patients highlighted specific colonic fibroblast populations that may contribute to inflammation and disease progression. While these studies emphasize the proinflammatory function of activated intestinal fibroblasts, it is unknown whether subsets of intestinal fibroblasts may fulfill protective regulatory functions required to limit intestinal inflammation. Our studies leverage a novel spatial transcriptomics method (MERFISH) to generate a detailed cellular and molecular atlas of the colon, with an emphasis on elucidating the function of fibroblasts in mouse and human colitis. We discovered a new population of regulatory fibroblasts (Fregs) that differentiate during colitis and suppress intestinal inflammation. In particular, we found that Fregs appear in non-inflammatory regions and promote the differentiation of regulatory B cells (Bregs) and tolerogenic dendritic cells (tolDCs). The goal of this project is to define the molecular framework governing Freg, Breg and tolDC interactions during colitis, and establish the nature of these interactions in human UC. These studies will demonstrate how Fregs orchestrate immune tolerance to colitis and whether failure of this mechanism may contribute to tolerance breakdown in UC.

Abstract or Project Description

Pain is a major component of Inflammatory Bowel Disease (IBD), increasing the suffering of patients and decreasing their quality of life. However, the underlying cellular basis for pain in IBD and the role of pain-mediating neurons in driving gut dysfunction are not well known. Our goal is to characterize the role of neurons that mediate pain in IBD, and to determine whether inhibiting or activating these neurons leads to changes in the gut that can treat pain and inflammation.

Abstract or Project Description

Accumulating data supports the concept that Inflammatory Bowel Disease (IBD) has a preclinical period that may be amenable to early diagnosis and intervention. Understanding the molecular underpinnings of this preclinical period in disease pathogenesis offers new opportunities in risk prediction and prevention. We have recently reported the first data from a large study using longitudinal pre-diagnostic samples from the US Department of Defense Serum repository, the PREDICTS study, where we assessed the predictive value of a panel of antimicrobial markers and a proteomic panel (>1100 proteins). Additional –omics data including glycan profiles, metabolome and exposome are currently generated on these same samples that span from up to 5 years before diagnosis to first clinical symptoms and confirmed diagnosis. We will integrate these massive multi-omics datasets to develop machine-learning based models predictive of future progression of Crohn’s disease and ulcerative colitis years before diagnosis, and to discover driver biomarkers and biological processes responsible for disease initiation. This will be achieved by constructing time-varying disease-specific networks capturing the associations across all biomarkers measured at different years before diagnosis available through the PREDICTS initiative. We will make all results publicly available via a user-friendly web-platform to facilitate the easy query, exploration and utilization of these data.

Abstract or Project Description

The interleukin-2 (IL-2) protein plays an essential role in regulating the immune response and preventing the development of autoimmune disorders by supporting the growth and activity of regulatory T cells (Tregs). Promising preclinical and clinical work has demonstrated the potential for IL-2 to mitigate autoimmune diseases, including Inflammatory Bowel Disease (IBD). However, in addition to stimulating Tregs, IL-2 also stimulates effector T cells and natural killer (NK) cells, which drive inflammation and lead to autoimmune disease progression. Thus, developing a biased version of IL-2 that activates Tregs but not effector T or NK cells would represent a monumental advance for autoimmune disease therapy. A recently discovered antibody known as F5111 specifically directs the activities of IL-2 towards Tregs, while also extending its persistence in the bloodstream. We built on this work and designed a single-agent fusion protein that combines IL-2 with F5111 and demonstrated that our engineered therapeutic, called the F5111 immunocytokine (IC), selectively activated Tregs and protected mice against disease development in an IBD model. This project aims to advance the translation of F5111 IC, performing vital studies in animal models and patient samples that will catapult this molecule into clinical trials to directly benefit patients with IBD.

Abstract or Project Description

Despite medical advances, a significant proportion of Inflammatory Bowel Disease (IBD) patients are resistant to therapies, with CD4 T cells being consistently activated. Analysis of inflamed tissues from a cohort of refractory IBD patients reveals an enrichment of metabolism gene transcripts in CD4 T cells. Exploring the mechanisms, our preliminary data demonstrate that several proinflammatory cytokines can independently trigger metabolic phenotypes which we believe reinvigorates the pathogenic function of CD4 T cells. This led us to formulate the hypothesis that IBD CD4 T cells, governed by metabolic cues, utilize common intracellular metabolites to drive inflammatory response that results in disease pathogenesis. Thus, our research goal is to identify the metabolic vulnerabilities of IBD CD4 T cells, which will consequently guide strategies for developing efficacious therapies.

Abstract or Project Description

Recently there has been increased evidence that patients with Crohn’s disease (CD) commonly have disrupted sleep, and disrupted sleep correlates with the risk of disease flare. The sleep/wake cycle, immune function, and multiple biological processes are orchestrated by the circadian clock. The central circadian clock in the brain entrains these processes to the environment by light exposure, and CD subjects have evidence of misaligned circadian rhythms that correlate with subclinical inflammation. The long-term goal of this research proposal is to show that bright light therapy in CD improves barrier integrity, reduces markers of subclinical inflammation and modifies the microbial structure/function to an anti-inflammatory state. To test this hypothesis, we will conduct a randomized crossover trial of morning bright light therapy (60 min) given by a wearable device (Re-timer™). We will do three assessments in inactive CD subjects at: 1) baseline; 2) after four weeks of wearable glasses with bright light therapy; and 3) after four weeks of wearable glasses without bright light therapy. The results of this innovative proposal will improve quality of life in CD subjects, reduce the risk of disease flare and identify novel circadian regulated targets for the treatment of CD.

Abstract or Project Description

Diet plays a unique role in Crohn’s disease (CD) pathogenesis and therapy. In pediatric CD, commercial formulas used as exclusive enteral nutrition (EEN) are effective at inducing both clinical and biochemical remission. This study assesses the impact of a whole-food blended smoothie as EEN on CD activity and the intestinal microbiome.

The primary aims of this study are: 1) To evaluate the tolerability of a whole-food based smoothie as EEN therapy, and 2) To evaluate the ability of a whole-food based smoothie to induce remission of active pediatric CD. 

This study will compare the tolerability and efficacy of conventional formula EEN and whole-food blended smoothie EEN by enrolling a total of 40 participants with newly diagnosed pediatric CD. Participants will be provided either commercial formula or guided on the preparation of the home-blended smoothie. These participants will be given a specific recipe, Vitamix blender and be provided the food components to the smoothie. The study will total 8 weeks and will assess tolerance, clinical outcomes, the stool microbiome and quality of life.

Abstract or Project Description

Patients with Inflammatory Bowel Disease (IBD) can experience many years of inflammation and remission in the gastrointestinal tract. As a result, the intestines can undergo a structural change known as fibrosis, which narrows the inside. This narrowing is a major, and sometimes life-threatening, health issue for patients because it can block the passage of digested food and stool. Unfortunately, the only treatment for this condition consists of invasive surgical procedures to either widen the opening or remove the affected areas of the intestine. Therefore, there is an urgent need to develop less invasive therapies such as drug-based treatments. Preliminary studies carried out in our laboratory have pointed to a potential role for the molecule serine, an amino acid, in the development of intestinal fibrosis. For example, we have found that serine is produced in areas of fibrosis in the intestines of IBD patients and in our mouse model of intestinal fibrosis, suggesting that it may play a role in the development of this condition. The work proposed in this application will determine if in fact serine is largely responsible for intestinal fibrosis and establish methods to limit its production and thus prevent fibrosis development. Carrying out the proposed work can lead to the development of novel treatments for intestinal fibrosis and significantly improve the lives of patients with IBD.

Abstract or Project Description

The microbes that flourish within our guts are central to many aspects of human health. Which microbes are present can dictate susceptibility to a number of acute and chronic diseases, including IBD. This proposal will use a mouse model with a defined and stable set of bacteria to probe how mice interact with specific members of their microbiota in health and during disease. First, each microbe will be tested for its ability to elicit an immune response in healthy mice. Once we have established the baseline response in normal healthy mice, we will use chemicals to disrupt the gut lining to elicit an inflammatory response, or we will disrupt genes known to limit intestinal inflammation. We will then investigate how immune recognition of the microbiota changes under these altered conditions. This system will enable us to track responses to the microbiota with unprecedented precision and will provide clues regarding which microbes drive immune activation during dysregulated responses to the microbiota. Such studies will pave the way for future work aimed at manipulating responses to key members of the microbiota responsible for inflammatory diseases of the gut.

Abstract or Project Description

In Inflammatory Bowel Disease (IBD), inflammation of the digestive tract lining results in the development of chronic ulcers appearing throughout the intestines, colon or rectum. Although symptoms of IBD can be managed through the use of corticosteroids, complete disease remission is rare, often leading to surgical intervention. High expression of the transcription factor HIF-1_ is a key feature of regeneration in the Murphy Roths Large mouse, an inbred laboratory strain displaying remarkable regenerative capabilities. Recently, our team demonstrated enhanced regeneration of acute and chronic tissue wounds in adult mice following transient therapeutic upregulation of HIF-1_ using the experimental drug DPCA. Previous studies of HIF-1_ in IBD have been limited to preventative therapy and have not explored fibrosis inhibition or regeneration. Therefore, we hypothesize that transient upregulation of HIF-1_ can inhibit fibrosis of IBD lesions. We propose to study the effects of DPCA on 1) fibrosis in a human intestinal organoid (HIO) model; and 2) lesion development in a murine TGFb-induced model of intestinal fibrosis. This project will contribute to the development of a new clinical treatment for IBD, as an alternative or adjunct to existing therapies that focus on resolving inflammation, but do not address fibrosis.

Abstract or Project Description

Various immune cells in the gut recognize and respond to microbes that live there, which are collectively known as the gut microbiota. This project focuses on a subset of immune cells termed B cells, which are important for functions such as antibody production. In particular, we are investigating how certain kinds of beneficial gut bacteria interact with B cells to improve gut health. Moreover, we are testing whether harmful bacteria found in the gut of patients with Inflammatory Bowel Disease do not interact with B cells, which may partly explain the altered immune responses observed during gut inflammation in these patients. Collectively, our work will have a significant impact on Inflammatory Bowel Disease by identifying and investigating new ways by which beneficial and harmful bacteria interact with B cells in the gut, which could have important implications for the future treatment of these diseases.