Judith & Stewart Colton Center for Autoimmunity Research Projects & Lead Investigators
At NYU Langone’s Judith and Stewart Colton Center for Autoimmunity, our researchers conduct pilot projects and collect data for biomedical research to better understand the immunological basis of autoimmune disorders.
We pioneer basic science and translational research that drives advances in the diagnosis, treatment, and prevention of autoimmune diseases, such as ankylosing spondylitis, antiphospholipid syndrome, arthritis, lupus, Sjogren’s syndrome, and type 1 diabetes.
We also select a Colton Family Scholar, an early career investigator, to participate in the Department of Medicine’s Young Scholars Program and develop innovative projects that expand the scope of autoimmune research.
2021–22 Pilot Projects
Validation of a New Ion Channel as a Critical Modulator of Inflammatory Macrophage Function in Rheumatoid Arthritis
Principal investigator: Stefan Feske, MD
Rheumatoid arthritis (RA) is one of the most common autoimmune diseases affecting 0.5-1% of the population. The disease primarily involves the inflammation and destruction of joints but can frequently also cause inflammation and damage of internal organs such as lungs and blood vessels. RA starts with the inflammation of the synovium, the tissue lining the inner surface of joints, and then progresses by causing the destruction of cartilage, the rubber-like padding that covers and protects the ends of bones and that forms the joints, and progresses to erosion of bones themselves. The end stage of RA is the severe deformation and loss of function of joints, which results in significant disability.
Many components of the immune system are involved in the disease mechanisms causing RA. One of the most critical immune cells causing inflammation in RA is a white blood cell type called macrophage. These cells not only ingest pathogens and cell debris, they also secrete messenger molecules, called cytokines, that recruit and activate other immune cells and cells in tissues including the joints. In RA, an increase in the number of macrophages in affected joints is an early mark of disease, and the degree of macrophage infiltration correlates with the degree of joint damage. The important role of macrophages in the development of RA has been demonstrated using animal models of RA. Macrophages are the main producers of cytokines, which cause joint inflammation, and neutralization of these cytokines has revolutionized RA treatment in the past two decades. Although these drugs have dramatically improved treatment options for people with RA, many patients do not respond to or become resistant to therapy, necessitating new therapeutic approaches.
In this project, the Feske Lab is investigating how the function of macrophages and their ability to cause inflammation in RA is regulated by a new ion channel. Ion channels form pores that can be opened and closed in the membrane of cells. They transport inorganic ions such as sodium and calcium or small organic molecules into or out of cells. There are hundreds of different ion channels. Some of them have been intensively studied and are important drug targets for the treatment of neuropsychiatric or cardiovascular diseases.
We have observed that a so far little studied ion channel is highly and selectively enriched in inflammatory macrophages. Importantly, the channel is strongly upregulated in macrophages present in the joints of RA patients. In further experiments, we found that genetic deletion of the channel almost completely suppresses the proinflammatory function of macrophages.
The overall goal of our pilot project is to understand (1) if suppression of the function of this new ion channel ameliorates the severity of RA in animal models of the disease, and (2) how the channel regulates proinflammatory macrophage function, as well as joint inflammation and destruction. Because this channel, like many other ion channels, is present at the surface of cells and therefore accessible to drugs, the long-term goal of this project is to determine if the channel is a new drug target for the treatment of RA.
Targeting MPZL1 in Autoimmune Disease
Multiple sclerosis (MS) is caused by immune-mediated demyelination and inflammation in the central nervous system (CNS), which results in debilitating symptoms. Current therapies only partially inhibit disease progression, and patients with progressive MS have limited benefit. Patients with systemic lupus erythematosus (SLE) exhibit varying clinical presentations but frequently have damage to critical organs. Conventional SLE treatments (e.g., corticosteroids, cyclophosphamide) can have serious side effects, while biologics, such as abatacept, have limited efficacy.
New approaches to modify immune regulatory pathways are needed to improve patient outcomes in both of these multifactorial disorders. Immune cell activation is regulated by transmembrane immune inhibitory receptors, also known as checkpoint receptors, that contain immunoreceptor tyrosine-based inhibitory motifs (ITIMs; consensus sequence S/I/V/LxYxxI/V/L). Myelin protein zero like 1 (MPZL1, a.k.a. PZR) is a single-pass transmembrane protein with two ITIMs.
The MPZL1 structure suggests a regulatory role, yet little is known about its physiological function or ligands. Preliminary data indicate that MPZL1 plays a role in regulating autoimmune responses. We hope to define the mechanism by which MPZL1 regulates immune responses and explore the therapeutic utility of biologic modulators of MPZL1 function in mouse MS and SLE models. Our results could yield novel insights into pathogenesis and yield new therapeutic options for these disorders.
Identification of Putative Autoantigen in Hidradenitis Suppurativa
Hidradenitis suppurativa (HS) is a severe chronic inflammatory skin disease that occurs in human apocrine sweat gland-bearing skin regions, with an estimated prevalence of 1%. It is characterized by extremely painful nodules, odorous suppuration, ulceration, extensive formation sinus tracts, and permanent scarring. Current treatments, including antibiotics and TNF-α inhibitors, are not effective, with surgical excision being the last resort. Even after surgical excision, patients often develop painful nodules again and disease continues to affect neighboring skin regions, which severely and negatively impact on patients’ quality of life. Clinically, it is still extremely difficult to treat due to the lack of understanding of the disease pathogenesis and the molecular pathways driving disease progression. Thus, there is a high clinical need to better characterize the disease and to identify novel targets for efficacious therapeutics for HS.
Our preliminary data reveals massive B and plasma cell infiltration in the proximity of sweat glands and sinus tracts in HS lesional skin, suggesting a possibility that HS may be an autoimmune disease. We plan to use single-cell RNA sequencing and VDJ-sequencing to examine the clonality of the B cells and plasma cells, and to characterize the antibodies and their antigen(s). We also hope to expand our collection of HS patient samples to acquire a more comprehensive understanding of the role of plasma cells in HS pathogenesis and how these cells may lead to other autoimmune comorbidities.
Impact of Type I Interferons on Rheumatoid Arthritis Patient Monocytes, Monocyte-Derived Macrophages and Fibroblast-Like Synoviocytes Using a Novel Vascularized Pannus-on-a-Chip Model
In rheumatoid arthritis (RA), monocytes from the blood invade joint tissue (synovium/pannus) and transform into inflammatory monocyte and macrophage populations. This is influenced by environmental cues in the tissue, including fibroblast-like synoviocytes (FLS). Our preliminary data suggest that circulating type I interferons affect monocyte activation. The relative contributions of blood versus tissue signals in the invasion and differentiation of monocytes to inflammatory monocytes and macrophages is not known.
We hypothesize that increased type I interferon signaling in monocytes alters monocyte invasion into the joint tissue and that interferon-stimulated FLS promote a pro-inflammatory macrophage phenotype. We plan to test this hypothesis using a novel vascularized pannus-on-a-chip model. This bioengineered perfusable organ-on-chip device is ideal for examining effects of type I interferons on infiltrating and tissue cells and can be further adapted to include additional cell types in the future.
The bioengineered human pannus-on-a-chip system allows longitudinal analysis of cells to understand how the joint tissue environment changes the way cells act during immunomodulation. Manipulation of the RA-chip environments and monitoring the effects by multimodal means provides a new way to interrogate and examine human RA pathobiology. In our work, we plan to test type I interferons in this system. We will then test tumor necrosis factor inhibitors.
Ultimately, this bioengineered human synovium-on-a-chip model has the potential to achieve a so-called "clinical trial on a chip" that could prescreen patients suitable for specific immunotherapies. This platform could be used in preclinical drug testing for RA to support early-stage drug discovery work at the Colton Center.
2020–21 Pilot Projects
Artificial Intelligence to Understand Tissue Inflammation in Autoimmune Diseases
Artificial intelligence in autoimmunity project applies computational deep learning (DL) approaches to study pathophysiology of autoimmune diseases such as myositis and lupus nephritis. Tissue biopsies are commonly used for diagnosis and research purposes in myositis and lupus nephritis. Only certain features are used for diagnosis, which represent a tiny fraction of the information available on the slide. While this strategy creates a manageable workload and aids standardization across different pathologists, it also leaves much unstudied and unknown when considering human disease pathogenesis.
Multiomics approaches have been useful in extending our knowledge, but the tissue has to be dissociated and the critical architecture of the tissue inflammatory reaction is lost. Machine learning (ML) approaches to analyze tissue histopathology are revolutionizing our understanding of disease in oncology. ML algorithms have been published that can identify tumors and improve cancer diagnosis. However, such algorithms have not been applied to autoimmune disease. In autoimmune disease, we still have major outstanding research questions regarding which cell types participate in inflammation at the level of the tissue, and how they interact with each other in space. While these questions could be partially answered using traditional methods, we hypothesize that the ML approach will provide a much more efficient and unbiased way to understand the architecture of inflammation in autoimmune disease, holding great promise for novel insights. To address this, we setup a collaborative effort between algorithm developers, biologists, and bioimage analysts to understand pathology of complex autoimmune diseases.
We are proposing to develop a novel DL algorithm that will handle multiple parameters across multiple slides and stains that could be trained to study multiple different disease states and questions. This would provide an invaluable tool as well as novel insight into the pathogenesis of the two disease states we will study. Our team is ideally suited to this work, with extensive expertise in artificial intelligence (Dr. Razavian) and immunofluorescence and histology (Dr. Puranik).
Multi-Modal Single-Cell Analysis to Reveal the Genesis of Inflammation
Inflammatory bowel diseases (IBDs), including Crohn’s disease (CD) and ulcerative colitis (UC), are chronic inflammatory conditions of the gastrointestinal tract that affects more than three million people in the United States. Approximately 20 percent of IBD patients require surgery over the course of their disease. A restorative proctocolectomy with ileal pouch-anal anastomosis (RPC-IPAA), also known as a J-pouch, is a common surgical procedure performed on individuals who undergo a colectomy for complications of IBD. The procedure involves removal of the entire colon with the creation of a pouch by looping healthy small intestine into a novel J-shape organ that serves as functional colon.
Despite the quality of life and functional improvements provided by a J-pouch, approximately 20 to 50 percent of patients will develop an inflammatory disease in the pouch referred to as pouchitis. Pouchitis is associated with poor long-term function, often requiring long-term medical therapy, surgical redo, or a permanent ileostomy. It is not known why certain individuals develop pouchitis and what are the initiating events of this progressive inflammatory response. In this study, we will use a newly developed next-generation sequencing technique to define the molecular features of a healthy and inflamed pouch with unprecedented resolution. By examining this surgically created organ before and after the disease originates, we have a unique opportunity to understand the genesis of immune-mediated disorders and to identify the events that predetermine, establish, and maintain inflammation.
Molecular Engineering of FGL1 for the Treatment of Systemic Lupus Erythematosus
Systemic lupus erythematosus (SLE) is a common autoimmune disease with little effective therapies options, underlining an urgent need for the development of therapeutic agents that more effectively modulate human immunity, especially the T-cells responses. Recently, we discovered fibrinogen-like protein 1 (FGL1) as a novel functional ligand for T-cell checkpoint inhibitory receptor lymphocyte-activating gene 3 (LAG-3).
FGL1 is a soluble protein normally produced by hepatocytes, and our data suggest it can suppress antigen-specific T-cell activation in a LAG-3 dependent manner. Blockade of the FGL1–LAG-3 interaction by monoclonal antibodies potentiates anti-tumor T-cell immunity and eradicates established mouse tumors. In addition to anti-cancer applications, we also found that soluble recombinant FGL1 has immune suppressive activity in vivo. This proposal describes the engineering of novel FGL1-based therapeutics tailored for the treatment of SLE by generating hyperactive recombinant FGL1 mutants that strongly agonize LAG-3 signaling.
FGL1 mutants will be designed through a rational, structure-based mutagenesis based on the CryoEM structure of the FGL1 ligand/receptor complex (in collaboration with Dr. Kong), and selected through a novel functional screening system that couples FGL1/LAG-3 signaling to T-cell activity. Ultimately, and most importantly, candidate FGL1 therapeutics developed through the methods above will be assessed for efficacy in the treatment of SLE in mouse models (in collaboration with Boris Reizis, PhD). Thus, through these multidisciplinary and translational studies, we will advance new candidate SLE therapies for clinical development.
Microbial-Host Interaction in Response to COVID-19 Infection
The New York City area was at one point the hot spot of the 2019 coronavirus disease (COVID-19) pandemic in the United States. COVID-19 begins as an infection of the respiratory tract, but we know very little about how a patient’s immune system engages the coronavirus in the lungs leading to severe respiratory damage. Besides uncontrollable viral replication, data from other viral respiratory infections have supported that acquisition of a secondary pathogen leading to lower airway dysbiosis can also be contributing to the inflammatory process and poor outcome of these patients.
Studying the microbial and host immune environment in the lung of patients affected with this virus can provide insights about viral viability, acquisition of secondary pathogens, and development of hyper-aggressive host-immune responses. Here, we hypothesize that among subjects with SARS-CoV-2 and poor outcome, lower airway dysbiosis (virome and microbiome) induces an exacerbated proinflammatory state in the lung.
During the surge of cases with severe COVID-19 infection at NYU Langone, we performed clinically indicated bronchoscopies to alleviate hypoxemia in more than 100 patients and started a biorepository of samples from the lower airways paired with blood samples. Using these samples, we will evaluate the lower airway microbial environment (both viral and bacterial fraction) of subjects with SARS-CoV-2 and investigate the role of lung resident macrophage subsets, lymphocyte populations, and cytokine production on the clinical outcome. The identified biomarkers for predicting the severity of viral illness will provide new therapeutic targets for controlling damaging inflammatory mediators.
2019–20 Pilot Projects
Investigating the Role of the Long Noncoding RNA CHROME in Lupus
For decades, researchers have ignored the “dark matter” of the genome—RNAs that are transcribed but that do not code for proteins, also called long noncoding RNAs (lncRNAs). lncRNAs comprise the majority of transcriptionally active regions of the human genome, yet fewer than 1 percent of human lncRNAs are functionally characterized.
Recent studies have shown that lncRNAs are potent regulators of gene expression and have important functions in both health and disease. We have discovered an lncRNA, called CHROME, that is expressed specifically in primates and is dysregulated in systemic lupus erythematosus (SLE). In SLE, immune complex formation and toll-like receptor ligation trigger the expression of interferon (IFN), which propagates autoimmune inflammation. Notably, genetic susceptibility to higher IFN production is linked to SLE development, and is associated with higher serum autoantibodies and increased disease activity.
Our preliminary data indicate that CHROME is elevated in the plasma of SLE patients compared with healthy controls. Mechanistic studies have revealed that CHROME regulates IFN-stimulated gene expression in monocytes and macrophages. Using chromatin isolation by RNA purification (ChIRP) to map RNA–protein and RNA–DNA interactions, we found that CHROME binds at the promoters of IFN-stimulated genes and interacts with the DNA helicase topoisomerase to promote gene expression.
We hypothesize that CHROME contributes to SLE activity through transcriptional regulation of IFN-stimulated genes. To test this hypothesis, we propose investigating the expression of CHROME in patients with SLE to identify the mechanisms by which CHROME is induced in SLE and to identify how CHROME regulates IFN-stimulated gene expression.
Understanding the role of primate-specific lncRNAs such as CHROME in orchestrating IFN signaling may identify mechanisms underlying SLE-characteristic immune dysregulation and novel therapeutic targets for intervention.
Osteopontin as a Target in Lupus Nephritis
Principal investigators: Amrutesh S. Puranik, PharmD, and Timothy Niewold, MD
Long-term lupus results in accumulation of immune complexes in the kidney, which in turn leads to cell death and decline in kidney function. This condition is called lupus nephritis. Earlier studies from the Niewold Lab have shown that high levels of osteopontin are associated with lupus nephritis.
Recently, we observed that osteopontin is natively expressed by certain types of macrophages in the kidney. These macrophages reside in the kidney at birth and perform functions that include consuming dead cells and debris generated in the kidney, thereby keeping other cells safe. Similarly, macrophages also preferentially take up the immune complexes that reach the kidney and prevent them from damaging other cells.
In this study, we evaluate the role of these osteopontin-expressing macrophages (OPN-macrophages) in lupus nephritis. We expect that in lupus nephritis, OPN-macrophages will have an inflammatory phenotype and be localized to sites of damage in nephritis. Because these macrophages are involved in the early immune complex–mediated stages of nephritis, these cells could be attractive targets for early intervention or prevention of nephritis. Our studies could suggest OPN-macrophages as novel therapeutic targets.
Optimizing Immunoglobulin-Like Transcript 7 and BDCA-2 Regulation of Plasmacytoid Cells in Systemic Lupus Erythematosus
Principal investigators: Mark A. Jensen, PhD, and Timothy Niewold, MD
Systemic lupus erythematosus (SLE) is a prototype systemic autoimmune disease characterized by rash, arthritis, nephritis, serosal inflammation, cytopenias, and autoantibodies that are directed against nuclear antigens.
A hyperactive, dysregulated type I interferon signaling pathway is thought to contribute to disease pathogenesis, and high levels of type I interferons can be detected in the blood of a subset of patients. Levels of type I interferons are stably increased in these patients and are associated with worse clinical disease.
Plasmacytoid dendritic cells (PDCs) are a type of white blood cell that becomes activated in SLE and are thought to contribute to disease pathogenesis. Activated PDCs produce type I interferons and inflammatory cytokines and become potent antigen-presenting cells that promote the autoreactive immune response. PDCs express several different inhibitory surface receptors and intracellular signaling pathways that regulate their activation, as well as blunt production of interferons and inflammatory cytokines.
We studied PDCs from a cohort of SLE patients and unaffected controls. We found that PDCs from patients with more severe disease express lower levels of a subset of regulatory receptors. This suggests that PDCs in these patients have become dysregulated, leading to their chronic activation.
This research project focuses on using combinatorial approaches to identify two or more different signaling pathways that together optimally regulate PDCs in SLE patients that have more severe disease. This strategy has the potential to define an optimal combination of agents for regulating PDC activation that could lead to a novel treatment approach for SLE and rapidly advance to the clinic.
Defining Gene Regulatory Networks in Autoimmune Disease
Recent genetic studies have identified hundreds of gene variations that are associated with risk for autoimmune disease. Despite great strides in mapping the regions of the genome linked to autoimmune diseases, we know little about the biology or functions of these risk factors.
Many of the gene variants identified do not change the structure of proteins, but instead are likely to change the expression of nearby genes. Thus, the inherited gene variants associated with autoimmune disease result in a regulatory network that changes gene expression and alters immune cell function.
In this project, we seek to systematically define the gene regulatory network in human immune cells and to understand how sequence polymorphisms, or other changes in regulatory elements, perturb this network. Combining our expertise in human molecular genetics, human immunology, and autoimmune disease pathogenesis, our team is uniquely poised to accomplish these goals.
Development of a Novel Light-Inducible Cre Recombinase to Track and Manipulate Migration of Self-Reactive T Cells
It is increasingly clear that an immune response in one part of the body can shape a distinct immune response at a distant site. We know that interactions between the immune system and bacteria in the gut can have powerful effects on the course of autoimmune diseases that are spatially separated from the intestine, such as damage to the brain associated with multiple sclerosis or joint damage associated with arthritis.
Yet it remains far from clear how this occurs. A basic obstacle to understanding this problem is that we lack the tools to label cells in one part of the body and follow their movement to other sites, as well as the ability to manipulate cells in one part of the body and assess how this ultimately affects courses of disease.
To address these problems, we propose using a mouse that expresses a light-inducible Cre recombinase, which will allow us to alter gene expression in cells at a specific time and place using blue light to track the cells. As a proof of principle, we plan to use these mice to identify immune cells in the intestine and trace cell migration upon the induction of a murine model of multiple sclerosis.
We hypothesize that inflammation induces the release of immune cells from the gut, that these cells traffic to the central nervous system, and that their exit from the intestine requires expression of a homing receptor, sphingosine-1-phosphate receptor 1 (S1P1). The U.S. Food and Drug Administration (FDA)–approved drug, Gilenya®, inhibits S1P1.
Ultimately, we hope our study enables investigators to ask a wide range of fundamental questions about cell migration in autoimmune disease and to identify novel targets for therapy.
Novel Hematopoietic Stem Cell Transplant Strategies for the Treatment of Autoimmune Disease
Principal investigators: Christopher Y. Park, MD, PhD and Doyun Park, MD
The cornerstone of therapy for autoimmune disorders is the elimination, or crippling, of immune cells that cause these disorders. However, current strategies frequently provide only temporary control of disease. Blood stem cell transplant holds the promise of resetting the immune system by allowing complete replacement of the blood system with normal cells.
This strategy has been explored as a treatment for severe and refractory autoimmune diseases. Although these studies strongly suggest that blood stem cell transplant can provide a significant clinical benefit, even for patients with advanced autoimmune disease, this approach has been limited to the investigational setting. Risks associated with transplant include short-term mortality from the procedure and long-term consequences, such as infertility and secondary cancers.
Developing strategies for blood stem cell transplant that reduce short- and long-term side effects and preserve, or even improve, immunologic remodeling after the procedure would likely promote its use as a treatment for lupus and other autoimmune diseases.
We plan to use animal models of autoimmune disease to test whether safer methods of performing transplants and providing specific subtypes of stem cells may better treat diseases with the goal of moving these approaches into the clinic.
Understanding Lupus Nephritis
Lupus nephritis is one of the most feared complications of systemic lupus erythematosus (SLE). It arises without warning or symptoms and can rapidly progress to renal failure. Clinically, we monitor urine tests and kidney function, but this can only detect disease after it has begun. Treatment is still nonspecific immunosuppression, which carries a high risk of side effects.
We seek to address the challenges of lupus nephritis with the goal of developing a prognostic model and identifying novel targets for therapy. Our multidisciplinary team includes outstanding expertise in genetics, murine and human immunology, and clinical lupus pathogenesis.
We propose three main projects to address the genetic basis of the nephritis, immunological pathogenesis, and the prediction of disease in the clinic. These projects seek to accomplish the following objectives:
- understand the functional genetics of SLE-associated alleles that confer risk of nephritis
- examine the role of type I interferon in SLE nephritis
- develop a predictive model for lupus nephritis
These studies are designed to work together and leverage a number of previous research findings with the goal of bringing our research to the clinic. Our team of primary investigators have complementary areas of expertise to drive these projects to completion.
Defining the Antigenic Landscape of Muscle-Specific Kinase Autoantibodies to Design Therapies for Myasthenia Gravis
Myasthenia gravis is an autoimmune disease of the neuromuscular system that causes muscle weakness and fatigue. The condition can result in difficulty swallowing; altered facial, neck, and limb movements; and impaired breathing.
In approximately 20 percent of patients with myasthenia gravis, the disease is caused by autoantibodies that bind a protein—muscle-specific kinase (MuSK)—that is critical for synaptic differentiation and to maintain the connection between motor neurons and muscles. For this subset of patients, the disease can be severe, and the standard of care is less successful.
We are developing a pipeline to study how autoantibodies bind the MuSK protein. We hope that this knowledge may lead to the design of protein-based therapies to sequester autoantibodies and develop agonist antibodies to stimulate MuSK and allow the protein to carry out its normal function.
In the long term, if the project leads to an understanding of how autoantibodies recognize MuSK, this information could be used to design therapeutic antibodies, other proteins, or small molecules that block the interaction and protect patient tissues. The same approaches could be applied to any other autoimmune disease mediated by antibodies, including lupus, rheumatoid arthritis, Guillain-Barré syndrome, and more.
2018–19 Pilot Projects
Evaluation of the Transcriptome of Nonlesional, Non–Sun-exposed Skin to Provide Insights into the Pathogenesis of Lupus Nephritis and Response to Therapy
Lupus nephritis that leads to acute or end-stage renal failure is a major factor in the association of systemic lupus erythematosus (SLE) with mortality. Given the intense focus on new biologics for the treatment of lupus nephritis, the research community is eager for new pathologic insights and predictors of treatment unresponsiveness. Identification of a biomarker associated with poor prognosis would be useful for stratifying patients in clinical trials.
Efforts to identify biomarkers in serum and urine from people with lupus nephritis have not yet yielded sufficiently robust markers to replace renal biopsy. Previous studies have shown that endothelial changes, such as increased levels of membrane protein C receptor, predict poor responses to therapy. Furthermore, increased levels of the protein were noted in nonlesional, non–sun-exposed skin biopsies from people with lupus nephritis, suggesting that alterations in the microvasculature are widespread and extend to the dermal vasculature.
Analysis of this more readily accessible tissue, even distant from the primary affected organ, may provide an opportunity to explore surrogates for renal tissue analyses. Although ongoing studies are examining single-cell RNA sequencing to link phenotype to biotype and to identify cell-specific pathways in the kidney, this proposal addresses the hypothesis that these pathways may be reflected in uninvolved skin that is more likely to be serially biopsied.
The aim of this pilot study is to evaluate single-cell transcriptomes from biopsies of nonlesional, non–sun-exposed skin that has been temporally aligned with renal biopsies from people with lupus nephritis. We plan to explore associations in the context of biopsy class, activity and chronicity indices, and extent of tubulointerstitial disease; and renal outcome at 12 months.
This project leverages skin samples collected from patients with lupus nephritis, who are enrolled in the National Institutes of Health–funded Accelerating Medicines Partnership Rheumatoid Arthritis/Systemic Lupus Erythematosus (AMP RA/SLE) Program. The current proposal is to evaluate these cryopreserved skin biopsies on the 10× Genomics Chromium scRNA-seq platform, which offers droplet-based cell capture that is nearly 10-fold higher than previous approaches. This platform employs reverse transcription adapters equipped with unique molecular identifiers that more accurately count transcripts captured per cell.
In sum, analysis of more readily accessible skin tissue is expected to provide insight regarding keratinocyte, fibroblast, endothelial, and other tissue–resident cell dysfunction. We hope this novel approach may be better able to serially follow cell type–resolved gene expression signatures that may better correlate with renal outcomes than whole tissue or peripheral blood in patients with lupus nephritis.
Functional Genetics of Interferon Regulatory Factor 5 in Human Lupus
We study the lupus risk gene interferon regulatory factor 5 (IRF5). In our earlier work, we have shown that the risk variant of IRF5 is gain-of-function downstream of endosomal toll-like receptors. The role of IRF5 gene variants in causing SLE is not currently clear.
There are four common functional elements in the IRF5 gene. The SLE risk variant is a Neanderthal-derived haplotype that contains all four of these functional elements. These elements typically come together in strong linkage disequilibrium, so the contribution of each element cannot be determined individually.
This presents a major limitation of genetic epidemiology—because the variants are not observed in isolation on human chromosomes, the causal element or elements cannot be determined.
In our pilot project, we have used a novel DNA synthesis method in collaboration with the Boeke Laboratory and the Institute for Systems Genetics at NYU Langone to solve this problem. We are creating synthetic IRF5 haplotypes, which are not found in nature, that contain each risk-associated variant in isolation, as well as in novel combinations. Our goal is to elucidate the molecular function of each element and potential synergy or interaction. We have made these elements and are working to put them into stem cells, which would then allow for testing of the different IRF5 alleles in various immune cell types.
Identifying the Source of Pathogenic Type I Interferon in Experimental Lupus
Principal investigator: Boris Reizis, PhD
Elevated levels of type I interferons (IFNs α and β) are thought to contribute to the pathogenesis of systemic lupus erythematosus (SLE) and represent an attractive target of emerging therapies for the disease. Despite the readily detectable expression of IFN-stimulated genes (also called IFN signature), the actual cellular origin of IFN in human SLE patients and in SLE-prone animals remains poorly defined.
The overall goal of this pilot project is to establish the cellular source of IFN in experimental models of SLE, specifically in SLE-driven kidney inflammation. We use genetic reporters to visualize the ongoing production of IFN in experimental SLE and identify the IFN-producing cells in the lymphoid organs, as well as in inflamed tissues such as the kidneys.
Defining the Antigenic Landscape of Muscle-Specific Kinase Autoantibodies to Design Therapies for Myasthenia Gravis
Myasthenia gravis is an autoimmune disease of the neuromuscular system that causes muscle weakness and fatigue, resulting in difficulty swallowing; altered facial, neck, and limb movements; and impaired breathing.
In approximately 20 percent of patients, the disease is caused by autoantibodies binding to a protein—muscle-specific kinase (MuSK)—that is critical for synaptic differentiation and maintaining the connection between motor neurons and muscles. For this subset of patients, the disease can be severe, and the standard of care—immunosuppressants and blood plasma replacement—is less successful.
Dr. Bhabha, Dr. Burden, and Dr. Ekiert are studying how the autoantibodies bind to MuSK protein in the hopes that this knowledge may lead to the design of protein-based therapies to sequester the autoantibodies, allowing MuSK to carry out its normal function.
In the long term, if the project leads to an understanding of how autoantibodies recognize MuSK, this information could be used to design therapeutic proteins or small molecules that block the interaction and protect tissue. The same approaches could be applied to any other autoimmune disease mediated by antibodies, such as lupus, rheumatoid arthritis, Guillain-Barré, and others.
Understanding the Functional Role of Purine Nucleoside Phosphorylase NP Polymorphism in Human Systemic Lupus Erythematosus
Dr. Ghodke was a lead investigator of the first study to identify a variant of purine nucleoside phosphorylase (PNP) as a lupus risk gene. PNP is essential to cellular DNA synthesis.
Additional studies have demonstrated that the variant of the PNP gene associated with lupus disrupts DNA synthesis. In this pilot project, we hypothesize that this disruption causes fragments of the DNA to leak out from the cell, which triggers the immune system and eventually results in lupus.
Because the PNP gene is also critical for metabolizing a chemical that helps the immune system to maintain balance, we also hypothesize that a variant of the gene may also lead to the overactive immune response that is the hallmark of lupus.
If correct, both hypotheses would yield novel mechanisms of lupus pathogenesis. Moreover, each has the potential for personalized therapeutic development. One of the Colton Center for Autoimmunity’s overarching goals in lupus research is to address the fact that the disease’s origins and development can be completely different from patient to patient, yet standard care is “one size fits all.”
One of the implications of this project is that some patients with lupus might be receiving drugs that actually exacerbate the disease. For example, the medication azathioprine interferes with DNA synthesis in the same way as the variant of the PNP gene. This project could help move the field toward diagnosing and treating patients based on their molecular and immunological profiles.
Type I Interferon as a Predictor of Treatment Response in Rheumatoid Arthritis
Principal investigator: Theresa L. Wampler Muskardin, MD
For patients with rheumatoid arthritis, receiving the right kind of treatment as early as possible is crucial. Remission within the first three months of therapy is the best predictor of remission at one year, so a delay in getting the correct treatment can significantly change the likelihood of a positive outcome.
Because there are currently no biomarkers to predict which treatment is best for a given patient, physicians try a therapy and take the “watch and wait” approach to judge whether it’s working. Because of this, patients are often at risk of permanent joint damage.
Dr. Wampler Muskardin has demonstrated that measuring levels of type I interferon may help predict which patients will respond to the inhibitors commonly used to treat rheumatoid arthritis. Her work to date suggests there is a precise degree of interferon activity that correlates with outcomes. This could enable physicians to make objective, data-based decisions about which therapy to recommend.
Investigating Cellular and Molecular Triggers of Disease Progression in Psoriatic Arthritis
Approximately 30 percent of patients with psoriasis will eventually develop psoriatic arthritis, which presents a unique opportunity for potential treatments. If researchers are able to find biomarkers that indicate exactly when and how psoriasis turns into psoriatic arthritis, the discovery could lead to early intervention and prevention in a susceptible and readily identifiable population.
So far, however, researchers have made little progress in understanding the pathology that links the two conditions due to a limited understanding of the phenotypic, molecular, and cellular events underlying the transition from psoriasis to psoriatic arthritis.
One of the major questions, and the subject of this study by Dr. Koralov and Dr. Scher, is to understand how cells of the adaptive immune system promote the progression from psoriasis to psoriatic arthritis. The two are working closely with colleagues at the New York Genome Center to take advantage of new high-throughput technology.
This technology enables researchers to simultaneously analyze the molecules on the surface of cells and the RNA molecules being expressed inside the cell, as well as to sequence the antigen specificity of T and B cells. These insights provide unprecedented insight into the immune landscape of inflamed tissue.
This research could provide a complete map of the cells that are pivotal in the inflammatory process of various tissues and how these may interact with each other to break tolerance and promote disease.
Exposure to Phthalates in Pregnant Women with Systemic Lupus Erythematosus: A Proof-of-Concept Study
Phthalates are a group of organic chemicals that are widely used in consumer products and are considered nonpersistant, meaning that environmental damage attributed to these chemicals is reversible. Animal and in vitro studies have associated alterations in the immune system and susceptibility to autoimmunity with exposure to phthalates.
A recent comparison of 58 people with systemic lupus erythematous systemic (SLE) and 78 controls has shown that those with SLE had substantially higher levels of phthalate metabolites measured in banked serum than healthy controls. Phthalate exposure represents a special concern in pregnant women because of the evidence suggesting that phthalates can influence intrauterine fetal growth and birth outcomes.
The objective of this pilot project is to measure concentrations of phthalate metabolites in urine samples collected through pregnancy in a group of pregnant women with SLE and to compare their levels with existing data from a cohort of pregnant women from the general population, who had no underlying conditions.
2017–18 Pilot Projects
Microbiome and Its Metabolites in Psoriatic Arthritis Pathogenesis
Our team examines how the intestinal and skin microbiomes and the metabolites these microbes produce contribute to the chronic inflammation that characterizes psoriatic arthritis (PsA). We study how these microbiomes and microbial metabolites differ between patients with PsA and healthy individuals and also changes in the microbiome and metabolites that take place during progression from psoriasis to PsA.
We also examine changes in circulating immune cells among patients with PsA and control cohorts to correlate the changes in microbiota and metabolites to functional changes in the immune compartment. We recently characterized a novel small animal model of PsA and are using this model to demonstrate and probe the causal link between changes in microbiota and the metabolites they produce and the chronic inflammatory response in the skin and joints of these animals.
Our hope is that these studies will produce novel insight into PsA pathogenesis and identify novel targets for new therapeutic approaches.
Translational Regulation of Autoimmune-Suppressive Regulatory T Cells
Principal investigators: Robert J. Schneider, PhD, and Adam Mor, MD, PhD
Regulatory T cells (Tregs) suppress the activation of other immune cells by both contact-dependent and -independent mechanisms, thereby maintaining immune system homeostasis, inhibiting effector T cells in the periphery, controlling excessive responses to foreign antigens, and preventing autoimmune disease. When Treg function is impaired, autoimmune diseases arise both in patients and in mouse models.
Although it is well established that Treg production increases when the enzyme mTOR is inhibited—in fact, mTOR inhibitor drugs are sometimes used in kidney transplant recipients to prevent organ rejection—there are still many questions about the exact nature of the connection between mTOR and Treg production.
Our research has helped us better understand this connection with data showing a novel translational reprogramming mechanism that promotes Treg development and differentiation. We have found that disrupting the metabolic processes of mTOR is the key to Treg development. This disruption promotes the messenger RNA that “tells” undifferentiated, or naive, T cells to become Tregs, while inhibiting messenger RNA that tells undifferentiated T cells to become other types.
We seek to understand, and ultimately target with drugs, the crucial intracellular molecular translational control signals that are essential for promoting and impairing Treg development and function.
Understanding the Role of Calcium Ion Signaling in the Pathogenesis of Sjogren’s Syndrome
In Sjogren’s syndrome, a condition that affects as many as 4 million people in the United States, the moisture-producing glands of the body are impaired, leading to symptoms such as excessive dryness of the salivary and tear ducts, profound fatigue, chronic pain, major organ involvement, and neuropathies and lymphomas. Approximately 50 percent of patients develop complications, including non-Hodgkin’s B-cell lymphoma.
Although Sjogren’s syndrome is considered an autoimmune disease, the reason why autoimmunity develops—for instance, in response to a viral infection—is not understood. However, recent discoveries indicate that genetic and environmental factors precipitate Sjogren’s syndrome and that subsequent activation of the adaptive immune system starts the cycle of inflammation and gland destruction.
In preliminary data, we found an unexpected link between the onset of Sjogren’s syndrome and the way that T cells process calcium signals. Calcium ions are crucial to the biological processes of all cell types. In T cells, calcium ions flow into the cell through a channel that is regulated by several proteins: ORAI1, STIM1, and STIM2.
When any of these proteins is deleted, T cells cannot properly process calcium ions, which leads to the autoimmune inflammation of the salivary glands that characterizes Sjogren’s syndrome.
2016–17 Pilot Projects
Protein Engineering of a Nuclease for the Rational Immunotherapy of Systemic Lupus Erythematosus
A collaboration between the Reizis Lab, which specializes in experimental immunology, and the Cardozo Lab, which focuses on protein engineering, our project focuses on DNase IL3, a unique secreted deoxyribonuclease associated with rare familial cases of human systemic lupus erythematosus (SLE).
With a model of the DNase IL3 structure generated by the Cardozo Lab, our team has characterized unique properties and endogenous DNA substrate of DNase IL3. We have created a proof of principle for the delivery of exogenous DNase IL3 to ameliorate anti–double-stranded DNA response. Because DNase IL3 is an endogenous human serum protein, we hypothesize that its delivery as a biologic would be a safe and specific therapeutic strategy in SLE.
Isolation of Novel Members of the Gut Microbiota That Mediate Resistance to Autoimmunity
Our team is investigating the hypothesis that an increase in the incidence of autoimmune diseases is associated with changes in the environment, such as decreased exposure to helminths and alterations to the gut microbiota.
Previously, we found that infecting mice that have NOD2 mutations with helminths induces expansion of Clostridiales, which reverse intestinal disease by inhibiting Bacteroides vulgatus colonization. Helminth infections in the indigenous people of Malaysia have been associated with similar alterations in gut microbiota and suggest an antagonistic relationship between Clostridiales and inflammatory bacteria in humans.
Our research will isolate novel Clostridiales strains from a cohort of individuals with helminth infection and test their ability to suppress inflammation. This information may reveal strategies to improve the safety and efficacy of ongoing efforts to therapeutically target the microbiota in patients with autoimmune diseases.
2014–16 Pilot Projects
The Examination of Microbiota-Triggered Adaptive Immune Responses in Autoimmune Disease
Principal investigator: Dan R. Littman, MD, PhD
The central hypothesis of our studies is that an altered gastrointestinal microbiome, or dysbiosis, is an important initiating factor in the development of inflammatory arthritis in genetically susceptible individuals. Study findings may lead to novel treatment strategies to prevent or treat diseases such as rheumatoid arthritis, psoriatic arthritis, and ankylosing spondylitis.
In research conducted in collaboration with Steven Abramson, MD, and Jose U. Scher, MD, our group previously reported a correlation of new-onset rheumatoid arthritis with fecal microbiota colonization with Prevotella copri. Results from a mouse model showed that colonization with the Th17 cell–inducing, segmented filamentous bacteria spontaneously triggered arthritis, which has prompted us to study whether P. copri triggers disease.
We have cultured multiple strains of P. copri from patients with new-onset rheumatoid arthritis and healthy controls and have prepared sequencing libraries from 96 strains, which are currently being processed.
Our group is collaborating with rheumatology groups at the University of Oxford and University College London to sequence the microbiota of patients with rheumatoid arthritis, ankylosing spondylitis, and juvenile inflammatory arthritis.
The Examination of Microbiota-Triggered Immune Responses in Systemic Lupus Erythematosus and Antiphospholipid Antibody Syndrome
Principal investigator: Gregg Silverman, MD
The overarching goal of these studies is to investigate whether specific bacterial isolates in the intestines of people with systemic lupus erythematosus (SLE) contribute to their autoimmune pathogenesis and disease flares.
Working with Jill P. Buyon, MD, our group has uncovered several important new findings in studies of whether specific bacterial isolates in the intestines of patients with SLE contribute to their autoimmune pathogenesis and disease flares. One such finding has led to a project invention for antimicrobial fecal immunoglobulin A as a diagnostic for SLE.
The Role of DNase IL3 and Microparticle-Associated DNA in Human Systemic Lupus Erythematosus
Our group is investigating the role of DNase IL3 in systemic lupus erythematosus (SLE) and has developed a new assay to measure DNase IL3 activity in human serum or plasma, as well as a new method to measure DNA concentration in circulating microparticles in plasma.
We have developed three novel readouts of anti-DNA reactivity in SLE, which may represent novel, pathogenetically relevant parameters of the disease and have diagnostic or prognostic potential (or both). Based on these findings, we have developed a provisional project invention for DNase IL3 as a therapeutic agent.