Molecular Genetics of Inflammatory Bowel Disease [2nd ed. 2019] 978-3-030-28702-3, 978-3-030-28703-0

Table of contents :
Front Matter ....Pages i-x
Front Matter ....Pages 1-1
A Primer on IBD: Phenotypes, Diagnosis, Treatment, and Clinical Challenges (Katherine Falloon, Mark Lazarev)....Pages 3-24
Genetic Epidemiology of Inflammatory Bowel Disease, Early Twin and Family Studies (Jonas Halfvarson)....Pages 25-45
Experimental Models of Intestinal Inflammation: Lessons from Mouse and Zebrafish (Oscar E. Diaz, Rodrigo A. Morales, Srustidhar Das, Eduardo J. Villablanca)....Pages 47-76
Front Matter ....Pages 77-77
Complex Disease Genes and Their Discovery (Jeffrey C. Barrett, Mark J. Daly)....Pages 79-89
IBD Genomic Risk Loci and Overlap with Other Inflammatory Diseases (Fatemeh Hadizadeh, Charlie W. Lees, Catherine Labbé, John D. Rioux, Miles Parkes, Alexandra Zhernakova et al.)....Pages 91-115
Sequencing and Mapping IBD Genes to Individual Causative Variants and Their Clinical Relevance (Aleixo Muise, Hailiang Huang)....Pages 117-139
Genetic Risk Prediction in IBD (Urko M. Marigorta)....Pages 141-156
Molecular Profiling of IBD Subtypes and Therapy Responses (Ho-Su Lee, Isabelle Cleynen)....Pages 157-182
Inflammatory Bowel Disease and Epigenetics (Antonella Fazio, Dora Bordoni, Philip Rosenstiel)....Pages 183-201
MicroRNAs and Inflammatory Bowel Disease (Matthias Hübenthal, Andre Franke, Simone Lipinski, Simonas Juzėnas)....Pages 203-230
IBD Genetics and the Gut Microbiome (Shixian Hu, Alexander Kurilshikov, Alexandra Zhernakova, Rinse Weersma)....Pages 231-248
Front Matter ....Pages 249-249
NOD1 and NOD2 and the Immune Response to Bacteria (Maria Kaparakis-Liaskos, Ashleigh Goethel, Dana J. Philpott)....Pages 251-280
The IL-23/Th17 Axis in Intestinal Inflammation (Kevin J. Maloy)....Pages 281-303
Inflammatory Bowel Disease at the Intersection of Autophagy and Immunity: Insights from Human Genetics (Natalia Nedelsky, Petric Kuballa, Adam B. Castoreno, Ramnik J. Xavier)....Pages 305-328
The Epithelial Barrier (Celia Escudero-Hernández, Stefan Koch)....Pages 329-345
The Gut Microbiome in Inflammatory Bowel Disease (Aonghus Lavelle, Harry Sokol)....Pages 347-377
Front Matter ....Pages 379-379
Toward Personalized Therapy in Inflammatory Bowel Disease (Ryan C. Ungaro, Jean-Frederic Colombel)....Pages 381-389
Back Matter ....Pages 391-402

Citation preview

Charlotte Hedin · John D. Rioux  Mauro D’Amato Editors

Molecular Genetics of Inflammatory Bowel Disease Second Edition

Molecular Genetics of Inflammatory Bowel Disease

Charlotte Hedin  •  John D. Rioux Mauro D’Amato Editors

Molecular Genetics of Inflammatory Bowel Disease Second Edition

Editors Charlotte Hedin Gastroenterology unit Patient Area Gastroenterology Dermatovenereology and Rheumatology Karolinska University Hospital Stockholm, Sweden

John D. Rioux Montreal Heart Institute and Université de Montréal Montréal, QC, Canada

Mauro D’Amato School of Biological Sciences Monash University Clayton, VIC, Australia

ISBN 978-3-030-28702-3    ISBN 978-3-030-28703-0 (eBook) https://doi.org/10.1007/978-3-030-28703-0 © Springer Nature Switzerland AG 2013, 2019 This work is subject to copyright. All rights are reserved by the Publisher, whether the whole or part of the material is concerned, specifically the rights of translation, reprinting, reuse of illustrations, recitation, broadcasting, reproduction on microfilms or in any other physical way, and transmission or information storage and retrieval, electronic adaptation, computer software, or by similar or dissimilar methodology now known or hereafter developed. The use of general descriptive names, registered names, trademarks, service marks, etc. in this publication does not imply, even in the absence of a specific statement, that such names are exempt from the relevant protective laws and regulations and therefore free for general use. The publisher, the authors, and the editors are safe to assume that the advice and information in this book are believed to be true and accurate at the date of publication. Neither the publisher nor the authors or the editors give a warranty, express or implied, with respect to the material contained herein or for any errors or omissions that may have been made. The publisher remains neutral with regard to jurisdictional claims in published maps and institutional affiliations. This Springer imprint is published by the registered company Springer Nature Switzerland AG The registered company address is: Gewerbestrasse 11, 6330 Cham, Switzerland

Preface

In 2013, when the first edition of this book was published, the field of inflammatory bowel disease (IBD) was being galvanized by the ever-expanding range of new pathogenic pathways and potential molecular treatment targets discovered through genetic research. In the intervening 6 years, this knowledge base has been further enriched with new genes, pathways, epigenetic mechanisms, and a better understanding of the interaction between genotype and environment. Now, the web of intersecting and converging mechanisms is leading us away from classifying disease according to clinical phenotypes – Crohn’s disease vs. ulcerative colitis, for example – toward a new molecular taxonomy of IBD. Such a classification has its roots in genetic surveys that are the result of multinational networks of large groups of scientists and clinicians, collaborating to deepen our knowledge of the genetic foundations of IBD. The genetic and molecular overlap between different chronic inflammatory conditions has brought the promise of a future, comprehensive molecular taxonomy spanning all chronic inflammatory diseases, which may facilitate highly personalized treatment. Scientists from a range of disciplines are engaging with IBD as the archetypal disease at the interface between the human immune system and the environment. Accumulating evidence suggests that the complex interactions between humans and the microenvironment that forms the gut may even be an arena for the indoctrination of the developing immune system cells. As such, the interface of the gut mucosa may have a role in defining an individual’s lifelong immunological disposition. Understanding what happens when this interface is dysfunctional, such as in IBD, has the potential to illuminate its fundamental role in human health. With eight entirely new chapters and additional updates and revisions, this book aims to cover the full spectrum of the contribution of genetics to the current understanding of IBD.  Here, we review how the familial epidemiology of IBD makes evident what is known and what is unknown of the heritability of IBD. We introduce the utility of the available animal models of IBD for picking apart individual components of IBD pathogenesis. We detail the genes associated with IBD and examine the overlap with other chronic inflammatory disorders. The available methods for the genetic prediction of IBD risk are discussed, as well as the potential to i­ mplement v

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Preface

molecular profiling of IBD subtypes and therapy responses. The field of epigenetics gives insights into the mechanisms behind non-genetic but still heritable phenotypic traits. Epigenetic factors and posttranscriptional regulation of gene expression through, for example, micro-RNA, also underpin the influence of environment on the expression of the IBD risk genotype. Genetically driven variation in epithelial homeostasis alters gut barrier function and is causatively involved in IBD pathogenesis. Alterations in gut barrier function disturb the interaction between the gut microbiome and the human immune system, leading to imbalances on both sides. Ultimately, the cumulation of these insights, arising out of the newly identified genetic architecture of IBD, may provide the knowledge we need to better tailor therapy to the patient’s specific genetic and molecular IBD signature. This book should serve as a guide for navigating the growing complexity of the molecular genetics of IBD. Stockholm, Sweden Montréal, QC, Canada Clayton, VIC, Australia

Charlotte Hedin John D. Rioux Mauro D’Amato

Contents

Part I The Foundation of IBD Genetics: Human and Animal Models A Primer on IBD: Phenotypes, Diagnosis, Treatment, and Clinical Challenges ����������������������������������������������������������������������������������    3 Katherine Falloon and Mark Lazarev Genetic Epidemiology of Inflammatory Bowel Disease, Early Twin and Family Studies����������������������������������������������������������������������   25 Jonas Halfvarson Experimental Models of Intestinal Inflammation: Lessons from Mouse and Zebrafish������������������������������������������������������������������������������   47 Oscar E. Diaz, Rodrigo A. Morales, Srustidhar Das, and Eduardo J. Villablanca Part II The Genetic and Molecular Makeup of IBD Complex Disease Genes and Their Discovery ����������������������������������������������   79 Jeffrey C. Barrett and Mark J. Daly IBD Genomic Risk Loci and Overlap with Other Inflammatory Diseases������������������������������������������������������������������������������������   91 Fatemeh Hadizadeh, Charlie W. Lees, Catherine Labbé, John D. Rioux, Miles Parkes, Alexandra Zhernakova, Andre Franke, Charlotte Hedin, and Mauro D’Amato Sequencing and Mapping IBD Genes to Individual Causative Variants and Their Clinical Relevance����������������������������������������������������������  117 Aleixo Muise and Hailiang Huang Genetic Risk Prediction in IBD����������������������������������������������������������������������  141 Urko M. Marigorta Molecular Profiling of IBD Subtypes and Therapy Responses ������������������  157 Ho-Su Lee and Isabelle Cleynen vii

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Contents

Inflammatory Bowel Disease and Epigenetics����������������������������������������������  183 Antonella Fazio, Dora Bordoni, and Philip Rosenstiel MicroRNAs and Inflammatory Bowel Disease���������������������������������������������  203 Matthias Hübenthal, Andre Franke, Simone Lipinski, and Simonas Juzėnas IBD Genetics and the Gut Microbiome���������������������������������������������������������  231 Shixian Hu, Alexander Kurilshikov, Alexandra Zhernakova, and Rinse Weersma Part III Pathogenetic Pathways in IBD NOD1 and NOD2 and the Immune Response to Bacteria ��������������������������  251 Maria Kaparakis-Liaskos, Ashleigh Goethel, and Dana J. Philpott The IL-23/Th17 Axis in Intestinal Inflammation������������������������������������������  281 Kevin J. Maloy Inflammatory Bowel Disease at the Intersection of Autophagy and Immunity: Insights from Human Genetics��������������������������������������������  305 Natalia Nedelsky, Petric Kuballa, Adam B. Castoreno, and Ramnik J. Xavier The Epithelial Barrier ������������������������������������������������������������������������������������  329 Celia Escudero-Hernández and Stefan Koch The Gut Microbiome in Inflammatory Bowel Disease ��������������������������������  347 Aonghus Lavelle and Harry Sokol Part IV Concluding Remarks and Future Perspective Toward Personalized Therapy in Inflammatory Bowel Disease������������������  381 Ryan C. Ungaro and Jean-Frederic Colombel Index������������������������������������������������������������������������������������������������������������������  391

About the Editors

Charlotte  Hedin, MD, PhD  is a Specialist in Luminal Gastroenterology at the Karolinska University Hospital in Stockholm. She completed her PhD on Crohn’s disease pathogenesis at King’s College London and Queen Mary University of London. In 2017, she was awarded a UEG Rising Star Award for her research in IBD and, in 2018, the Karolina Prize for “Exemplary Patient Flow and Quality Work” at the Karolinska University Hospital. She holds a clinical postdoctorate from the Karolinska University Hospital. She is a Member of the Swedish Organisation for the Study of IBD (SOIBD)) and is Committee Member for the European Crohn’s and Colitis Organisation (ECCO). In addition, she collaborates in international IBD research projects, including the GEM project. Dr. Hedin’s research focusses on delineating pathogenic pathways in IBD through studying at-­risk individuals (families of IBD patients) and defining the process of mucosal healing. John  D.  Rioux, PhD  is a Full Professor of Medicine at Université de Montréal (UdeM) and Senior Researcher at the Montreal Heart Institute (MHI) and holder of the Canada Research Chair in Genetics and Genomic Medicine. He is a Founding Member of multiple international consortia and currently co-leads the International IBD Genetics Consortium, is Chair of the Steering Committee of the NIDDK IBD Genetics Consortium and the Leader of the IBD Genomic Medicine (iGenoMed) Consortium. Dr. Rioux’s research focuses on three main areas: (1) genetic studies to identify risk factors for common and rare diseases, (2) functional studies to understand how these genetic risk factors protect or predispose to disease, and (3) integrative human studies to identify predictive biomarkers of important clinical outcomes. His work has led to over 200 publications, cited over 30,000 times. Mauro D’Amato, PhD  is Professor of Genetics and Genomics and Head of the Gastrointestinal Genetics Unit at the School of Biological Sciences, Faculty of Science, Monash University, Melbourne, Australia. He conceived and coordinates the largest gene mapping effort in irritable bowel syndrome, the bellygenes initiative exploiting data from >800,000 individuals, has served in the Management Committee of several consortia including the International IBD Genetics Consortium, and is ix

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About the Editors

Member of the European Microscopic Colitis Group. His team combines genomic, computational, and preclinical expertise to elucidate the pathogenetic mechanisms predisposing to inflammatory and functional gastrointestinal diseases. The druggable genome, nutrigenetics, and host (genome)-microbiota interactions are also new research lines within the group. His research has resulted in more than 150 publications and over 13,000 citations.

Part I

The Foundation of IBD Genetics: Human and Animal Models

A Primer on IBD: Phenotypes, Diagnosis, Treatment, and Clinical Challenges Katherine Falloon and Mark Lazarev

Abstract  Inflammatory bowel disease (IBD) is a chronic inflammatory condition of the gastrointestinal tract, most commonly divided into ulcerative colitis (UC) and Crohn’s disease. We have seen an increase in incidence in IBD over the last few decades worldwide. UC and Crohn’s disease have strong genetic underpinnings which have been steadily elucidated over the past 20 years. Additionally, there are number of environmental factors that have been recognized as triggers of disease, including dietary/microbiome and smoking. In this chapter, we lay out the primary phenotypes observed in Crohn’s and UC. We next discuss the approach to diagnosis, which is generally multifactorial, including blood and stool testing, abdominal imaging, and colonoscopy with biopsy. Next, we summarize the treatment algorithms for both disease, including the pre- and post-biologic era. Greater concentration is given to the discussion of the anti-tumor necrosis factor (TNF) alpha, which to date has been the greatest game changer in IBD management. We also discuss the newer pharmacologic mechanisms of targeting the disease including lymphocyte adhesion blockers, anti-IL 12/23 inhibitors, and drugs that target the JAK/STAT pathway. Some of the newer agents in the pipeline are also briefly discussed. In the final section, we explore clinical correlates of the genetic findings to date. We delve into some of the key findings including the discovery of the NOD2 risk gene, as well as the most up-to-date genome wide association studies (GWAS) findings. Through this we explore how these genetic findings correlate with specific disease phenotypes, and how the findings have helped choose targets for pharmacologic therapy.

K. Falloon · M. Lazarev (*) Johns Hopkins University School of Medicine, Department of Medicine, Division of Gastroenterology, Baltimore, MD, USA e-mail: [email protected] © Springer Nature Switzerland AG 2019 C. Hedin et al. (eds.), Molecular Genetics of Inflammatory Bowel Disease, https://doi.org/10.1007/978-3-030-28703-0_1

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Introduction Inflammatory bowel disease encompasses a group of diseases that involve chronic, relapsing inflammation of the GI tract, divided into three main categories—ulcerative colitis (UC), Crohn’s disease (CD), and indeterminate colitis (IC). Another form of chronic colitis includes microscopic colitis (lymphocytic colitis, collagenous colitis); this group will not be covered in this review. There appears to be a geographical pattern to IBD, with a North–South gradient both across the hemispheres and within individual countries [1, 2]. IBD predominates in the Western world, with rates highest in Europe and North America, where approximately 0.4% of the population lives with IBD [2, 3]. In the USA alone, this leads to a quarter million doctor visits, 30,000 hospitalizations, the loss of over a million workdays, and direct medical costs of over 4 billion annually [4–7]. Incidence of the disease around the world has either stabilized at a high rate or is increasing, suggesting the emergence of IBD as a global disease [8]. IBD is most prevalent among Caucasians, followed by African Americans, though incidence rates in Asian Americans and Hispanics are growing [9]. It is also more prevalent among the Ashkenazi Jewish population [10]. Peak onset is between 15 and 30 years, although the disease can begin at any age [8]. The pathogenesis of IBD is only partially understood. The prevailing theory posits that IBD is the result of an excessive inflammatory response to an environmental trigger (e.g., infection or medication) in a genetically predisposed individual. The “hygiene hypothesis” has been proposed to explain the geographic and demographic tendencies of IBD, with the thought that factors such as increased industrialization, sanitation, and quality of health care systems may help explain the increased risk of IBD in the Western world as well as in higher social strata [10–13]. In support of this hypothesis are studies demonstrating lower risk of IBD in larger or poorer families with lack of access to clean water as well as the increasing incidence rates among immigrants who move from low to high incidence regions, though understanding of the development of disease remains incomplete and further study is needed [14–16]. Genetics also play a role, with a positive family history still the greatest risk factor for IBD and a number of genes already implicated in development of the disease [10, 17]. Other factors of interest include diet, the microbiome, and smoking. There have been associations with specific dietary components or patterns and the development of IBD, but the quality of these studies is mixed and so it is challenging to draw any meaningful conclusions [10, 11, 18–20]. It is possible that dietary intake may be most important for the development of IBD in the way that it affects the gut microbiome. Numerous studies have demonstrated that dysbiosis is prominent in IBD, and when compared to healthy controls, patients with IBD have decreased numbers and reduced diversity of Firmicutes and Bacteroides [21, 22]. However, whether alterations in gut flora are the result of the disease or lead to it is still undetermined [18, 23–25]. Smoking worsens the course of CD but has been associated with more benign disease course in UC [26, 27]. The use of nonsteroidal a­ nti-­inflammatory drugs has also been shown to increase the frequency of flares in patients with IBD [28].

A Primer on IBD: Phenotypes, Diagnosis, Treatment, and Clinical Challenges

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Clinical Manifestations and Phenotypes The two main forms of IBD include UC and Crohn’s disease. This distinction is important as the two diseases vary in both their pathogenesis and their treatment. UC is characterized by diffuse inflammation that is typically confined to the mucosa. It involves the rectum in the vast majority of cases and can extend in uninterrupted fashion to the rest of the large intestine but does not involve the small bowel [5]. Disease course is commonly characterized by the gradual onset of bloody diarrhea with urgency, tenesmus, and cramping abdominal pain [5, 29] There are various classification schemes for UC. The Montreal consensus classifies UC based on anatomic extent—ulcerative proctitis (E1), distal or left-sided UC (E2), and extensive UC involving the colon proximal to the splenic flexure (E3) [30]. Severity of disease can also be defined by the Mayo system, which takes into account factors such as stool pattern, rectal bleeding, and endoscopic findings [31]. Long-term complications may include medically refractory disease and rarely toxic megacolon, a dilation of the colon which left untreated could result in perforation. Crohn’s disease, on the other hand, involves transmural inflammation and can occur anywhere in the gastrointestinal tract [32]. Crohn’s is most frequently classified according to disease location (terminal ileal or L1, colonic or L2, ileocolic or L3, isolated upper GI or L4) and behavior (non-stricturing and non-penetrating or B1, stricturing or B2, penetrating or B3) [32]. Fifty percent of patients present with ileocolonic disease, 80% will have ileitis, and 20% will have isolated colonic disease [33, 34]. Disease presentation is dependent on disease location but is typically characterized by diarrhea, abdominal pain, and weight loss. Involvement of the upper GI tract can lead to aphthous ulcers, nausea, and vomiting. Involvement of the small bowel can lead to malabsorption of bile acids, iron, vitamin B12, and fat soluble vitamins. Rectal bleeding is less typical except in those patients with Crohn’s localized only to the colon. In CD, perianal disease is present in about 30–35% of patients [35–37]. Complications of perianal disease include perirectal abscesses, anorectal fistulas, and anal fistulas. Over time, left untreated, over 80% of patients with CD will develop some complication of disease, which includes strictures, fistulas, or abdominal abscesses [38]. Both Crohn’s and UC can also have extra-intestinal manifestations. These include ocular complications (episcleritis, scleritis, uveitis), arthropathies and sacroiliitis, and dermatologic complications (erythema nodosum, pyoderma gangrenosum) [5, 32]. Primary sclerosing cholangitis (PSC) is more likely in patients with UC and is associated with a risk of chronic liver disease, and an increased risk of colorectal cancer [39, 40]. Anxiety and depression are also more common among patients with IBD, especially those with greater disease activity [41, 42]. Of note, approximately 5–10% of patients with IBD are unclassifiable and thus labeled as indeterminate colitis [43]. This is more common in children than in adults [44]. Other chronic colitides include microscopic colitis, in which the colon appears endoscopically normal but is characterized histologically by lymphocyte infiltrates (lymphocytic colitis), or a subepithelial collagen band (collagenous colitis) [45].

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Diversion colitis can occur if part of the colon is excluded from the fecal stream [46]. Diverticular colitis, which is limited to portions of the colon with diverticula present, and pouchitis can occur in patients with UC who have undergone total proctocolectomy with ileal pouch-anal anastomosis [47]. Other diseases that can present similarly to IBD and may need to be considered based on clinical context include infections (infectious colitis, amebic colitis, schistosomiasis, intestinal TB), NSAID enteropathy, Behcet’s, celiac disease, radiation colitis, ischemic colitis, intestinal lymphoma, eosinophilic enteritis, bile salt malabsorption, bacterial overgrowth, over-use or misuse of laxatives, irritable bowel syndrome (IBS), and colon cancer.

Diagnosis The diagnosis of IBD is made based on clinical suspicion in combination with laboratory, radiologic, endoscopic, and histologic findings [48]. The clinical symptoms of IBD have been described above. Physical exam findings may include tachycardia, cachexia, abdominal distension, tenderness, rebound, or guarding, skin tags, fissures, or fistulas in the perianal region, or extra-intestinal manifestations such as aphthous ulcers or erythema nodosum. Patients in whom there is suspicion for IBD should be ruled out for infectious causes of symptoms via stool studies, including testing for C. difficile, which is also found at increased rates in patients with IBD [49, 50]. Pertinent blood work includes complete blood count (CBC) to look for anemia, ferritin, erythrocyte sedimentation rate (ESR), and C-reactive protein (CRP) to look for inflammation, and albumin and electrolytes to look for signs of malnutrition. Tissue transglutaminase IgA and quantitative IgA may also be performed to rule out celiac disease. Serologic testing includes perinuclear anti-­ neutrophil cytoplasmic antibodies (p-ANCA) and anti-saccharomyces cerevisiae antibody (ASCA)—a positive p-ANCA and negative ASCA suggest UC while a negative p-ANCA and positive ASCA suggest CD—however, such testing is usually unnecessary if further endoscopic work-up is planned [29]. Fecal calprotectin and lactoferrin can be indicative of intestinal inflammation, though they are not specific for IBD associated intestinal inflammation. From a radiologic perspective, plain abdominal films have little utility. Computerized tomography (CT) and magnetic resonance enterography (MRE) employ both a water-based oral contrast and intravenous IV contrast, and have become the standard for evaluating the small bowel for active disease, strictures, and fistulas. MRE has the added benefit of avoiding ionizing radiation. Barium studies are generally falling out of favor, largely secondary to lack of expertise in ­interpreting these studies, and the fact that they do not evaluate extraluminal regions (such as ruling out an abdominal abscess). Ultrasound is also emerging as an imaging modality that can be used to detect CD without exposing patients to radiation; however, its use has been limited as it is very operator dependent and

A Primer on IBD: Phenotypes, Diagnosis, Treatment, and Clinical Challenges

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requires specific expertise in study interpretation [51, 52].With regard to evaluation for perianal disease, MRI of the pelvis is the study of choice to better characterize the anatomy of these lesions [51]. Endoscopy with biopsy remains the gold standard for diagnosis and is used to assess severity of disease and response to treatment, monitor for development of dysplasia, and provide endoscopic therapies [52–54]. Colonoscopy with ileal intubation and biopsy from the colon and ileum is usually satisfactory in making a diagnosis. A lack of endoscopic inflammation does not preclude histologic inflammation, and so if there is a clinical suspicion biopsies should be performed regardless of colonic appearance [54]. The classic endoscopic findings in UC include loss of the typical vascular pattern with a granular texture to the mucosa, friability, superficial ulceration, and a line of demarcation separating the affected from the unaffected colon [55]. In UC, the inflammation typically involves the rectum with variable stretches of continuously diffusely inflamed mucosa more proximally, though patients with UC can also have an isolated cecal patch. By contrast, Crohn’s disease is typified by skip lesions, segmental colitis, rectal sparing, serpiginous and irregular stellate ulcers, cobblestoning (ulceration with tissue edema), as well as perianal disease, fistulous tracts, and stricturing disease [54]. Ileal involvement is classic for CD, though backwash ileitis (a mild non-specific terminal ileitis) may also be seen in patients with UC who have pancolitis [56]. When used in conjunction with other available diagnostic testing, colonoscopy can differentiate UC from CD in approximately 90% of cases [54, 57]. Other endoscopic modalities include esophagogastroduodenoscopy (EGD), capsule endoscopy, and single and double balloon enteroscopy. EGD can be useful for evaluating involvement of the upper GI tract in patients with Crohn’s disease and is recommended for children suspected of having CD, though routine EGD is not indicated for adults unless symptoms dictate the need (including upper abdominal pain, nausea/vomiting, or dysphagia) [54, 58]. Capsule endoscopy can directly visualize the small bowel mucosa and demonstrate erythema, villous atrophy, erosions, and ulcerations typical of CD when other modalities have failed to diagnose disease but suspicion remains high [59]. However, capsule endoscopy does not allow for tissue sampling. Additionally, if there is a concern for a stricture, there is an elevated risk of capsule retention, and thus a dissolvable patency capsule is recommended prior to performing the study. If the capsule reveals a small bowel target, balloon enteroscopy may be needed for tissue sampling and to make a diagnosis. Histologically, findings seen in IBD include atrophy and architectural distortion of the crypts (feature of chronicity), and inflammation in the lamina propria [5, 60]. Basal plasamacytosis and basal lymphoid hyperplasia may also be seen. It is not always possible to distinguish Crohn’s from UC based on histopathology [61, 62]. Epithelioid granulomas are seen in Crohn’s, but not UC. That said, they are not typically found in biopsy samples and are not pathognomonic for Crohn’s [63–65]. Also, early in the disease course the inflammation can be non-specific and without features of chronicity.

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Treatment The primary goals of therapy in IBD are to reduce symptoms, control inflammation, improve quality of life, induce and maintain steroid-free remission, and prevent complications and hospitalizations [5, 32, 66]. Endoscopic mucosal healing has also become an important goal of management for patients with IBD [67–71]. Therapies for IBD are evolving, with new agents targeting different mechanisms continually emerging. For both UC and CD, the choice of therapy and mode of delivery depends on anatomic location of disease as well as extent of disease and extra-intestinal manifestations. There is no universal treatment strategy, and there is debate in the field regarding optimal therapeutic timing, with data suggesting that starting immunomodulators and biologics early (“top down therapy”), as opposed to the traditional course of starting with standard agents and escalating if patients fail to respond (“step up therapy”), may be beneficial [72, 73] (Tables 1). Mild to moderately active UC can be treated with oral and/or topical mesalamine, depending on disease location and patient preference; some may achieve maximum benefit from combination therapy [74, 75]. For treatment of mild CD, mesalamines are no longer recommended as they have not demonstrated consistent benefit [32, 76]. For patients with moderate to severe UC or CD, thiopurines (azathioprine, 6-mercaptopurine (6-MP)) have traditionally been the steroid sparing agent of choice prior to the approval of anti-TNF agents. Thiopurines can induce remission; however, prescribers must take into account their slow onset of action (up to 3–6 months) as well as their side effect profile, which includes dose-dependent bone marrow suppression, infection, hepatotoxicity, pancreatitis, and malignancies, particularly non-Hodgkin’s lymphoma and non-melanomatous skin cancer [5, 32, 77]. Today, thiopurines are rarely started as monotherapy in IBD. Agents targeting tumor necrosis factor (TNF) alpha have become the most important class of therapeutics for both the induction and maintenance of remission for both Crohn’s and UC. These agents are monoclonal antibodies targeting circulating tumor necrosis factor, one of the primary inflammatory mediators. Infliximab was the first to be approved for Crohn’s in 1998 as an IV infusion, followed by adalimumab and certolizumab pegol (both subcutaneous injectables) [78–81]. Infliximab, adalimumab, and golimumab have been approved for UC [82–84]. In a pivotal study, the combination of infliximab with azathioprine was more effective in the treatment of moderate to severe Crohn’s than either infliximab or azathioprine alone [85]. Overall response rates have hovered around 65–70% and remission rates of 35–40% [86]. These agents have been shown to lead to mucosal healing as well as decreased rates of hospitalization and surgery [71, 87, 88]. Additionally, anti-­TNFs are more effective when administered early in the disease course before c­ omplications have developed; this is especially true for Crohn’s disease [89]. Anti-TNFs are also useful for complex perianal fistulizing CD and in preventing post-operative recurrence [90, 91].

Biologics

Immunomodulators Azathioprine (Imuran) Mercaptopurine (6-MP)

Topical 5-asa (Canasa, Rowasa)

Induction and maintenance Oral of remission in moderate to Antagonizes purine severe UC/Crohn’s metabolism to de-activate T lymphocytes and suppress the mucosal immune system

(continued)

Prior to initiation: TPMT level or genotype While on therapy: Complete blood counts (CBCs), liver function tests (LFTs), skin cancer screening, PAP smears

Yearly creatinine

Headache, nausea/vomiting, abdominal pain, diarrhea, rash, fever paradoxical worsening of disease, pancreatitis, acute interstitial nephritis As above, and reversible oligospermia. Of note, typically less well tolerated than mesalamine but more affordable Typically fewer side effects than oral formulations secondary to less systemic absorption

Induction and maintenance of remission in mild to moderately active UC, alone or in conjunction with topical 5-ASA

Dose-dependent myelosuppression, increased risk of infection, hepatotoxicity, pancreatitis, rash, malignancy (particularly non-Hodgkin’s lymphoma and non-­ melanomatous skin cancer)

Monitoring

Notable side effects

Indications for use

Rectal (suppository or enema) Induction and maintenance See above of remission in mild to moderately active UC, alone or in conjunction with oral 5-ASA

Route of administration Drug class mechanism of action Aminosalicylates (ASA) Mesalamine Oral Acts through a variety of receptors, including synthetic nuclear receptors, that exert anti-inflammatory effects on Sulfasalazine (Azulfidine, EN-tabs) the colon

Table 1  Therapeutic agents for the treatment of IBD

A Primer on IBD: Phenotypes, Diagnosis, Treatment, and Clinical Challenges 9

Natalizumab (Tysabri)

Golimumab (Simponi)

Certolizumab (Cimzia)

Adalimumab (Humira)

Drug class Infliximab (Remicade)

Table 1 (continued)

Route of administration mechanism of action Intravenous (IV) infusion Chimeric (mouse/human) IgG monoclonal anti-tumor necrosis factor (TNF) alpha antibody Subcutaneous injectable Fully human anti-tumor necrosis factor (TNF) alpha Subcutaneous injectable Humanized pegylated anti-tumor necrosis factor (TNF) alpha antibody IV infusion Human monoclonal antibody to tumor necrosis factor (TNF) alpha IV infusion Monoclonal antibody that blocks alpha-4-integrin, inhibiting attachment of activated leukocyte in the capillary at site of inflammation Induction and maintenance of remission in moderate to severe Crohn’s (typically after failure to respond to anti-TNF)

Induction and maintenance or remission in moderate to severe UC

Induction and maintenance or remission in moderate to severe Crohn’s

Indications for use Induction and maintenance of remission in moderate to severe UC/Crohn’s

Monitoring Prior to initiation: Screen for tuberculosis, Hepatitis B While on therapy: Screening for skin cancer

JC virus— > 50% are Headache, nausea, abdominal pain, JC seropositive in general virus induced progressive multifocal leukoencephalopathy (PML) (almost never population used secondary to this side effect)

Notable side effects Opportunistic infections (bacterial and fungal), lymphoma (especially in combination with thiopurines), demyelinating processes, psoriatic and lupus-like reactions, infusion/injection reactions

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Metronidazole (Flagyl)

Antibiotics Ciprofloxacin

Budesonide (Uceris, Entocort)

Steroids Prednisone/ Methylprednisolone

Tofacitinib (Xeljanz)

Ustekinumab (Stelara)

Drug class Vedolizumab (Entyvio)

None required

None required

Disulfuram-like reaction with alcohol

None required

Less systemic effects compared to prednisone/methylprednisolone

Treatment of flares/ induction of remission in UC/Crohn’s

Increased risk of aortic dissection/rupture, hypoglycemia, tendon rupture, CNS effects

Dependent on duration of use, dosage

Cushingoid features, emotional and psychiatric disturbances, skin striae, metabolic bone disease, and accelerated atherogenesis

Higher all cause infections (most mild to moderate—nasopharyngitis), increased rates of herpes zoster increased cholesterol

Induction and maintenance of remission in moderate to severe UC (typically after failure to respond to anti-TNF)

Monitoring Prior to initiation: Screen for tuberculosis

Treatment of flares/ induction of remission in UC/Crohn’s

Notable side effects Nasopharyngitis. Lower risk of infection compared to anti-TNF agents. No known increased risk for malignancy Nasopharyngitis

Indications for use Induction and maintenance of remission in moderate to severe UC/Crohn’s Induction and maintenance of remission in moderate to severe Crohn’s

Septic complications in Oral or IV severe colitis, peri-anal Inhibits DNA gyrase and disease, pouchitis topoisomerase IV to halt bacterial cell division Oral or IV Inhibits bacterial nucleic acid synthesis

Oral/IV Binds to cytoplasmic receptors to inhibit DNA synthesis and suppress the immune system Oral As above but undergoes extensive first pass hepatic metabolism so with less systemic absorption

Route of administration mechanism of action IV infusion Blocks gut specific alpha-4-­ beta-7 integrin IV induction followed by maintenance subcutaneous injection Blocks p40 subunit of IL-12 and IL-23 Oral Small molecule that targets JAK1 and JAK3 in the JAK STAT pathway A Primer on IBD: Phenotypes, Diagnosis, Treatment, and Clinical Challenges 11

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Patients who do not have a response to induction with an anti-TNF are termed primary non-responders. By contrast, secondary non-responders are patients who initially respond to therapy but then lose response. Mechanisms of non-response or loss of response include (1) development of autoantibodies targeting the drug, (2) inflammatory response not mediated by a TNF-driven pathway, and (3) high rate of fecal wasting of the anti-TNF owing to large burden of active disease [92, 93]. Therapeutic drug monitoring can be helpful in sorting out if the dose should be increased, if an alternate anti-TNF agent should be tried, or if a drug employing a different mechanism of action should be attempted. Currently, it is recommended to check drug and antibody levels in a reactive approach where a patient is not responding adequately [94]. Generally, anti-TNFs are well tolerated. Potential side effects of anti-TNFs include infusion reactions, increased risk of infection (the drug is contraindicated in those with untreated latent TB and also places patients at increased risk for bacterial and fungal opportunistic infections), lupus-like illness, hepatotoxicity, development or exacerbation of multiple sclerosis, worsening of congestive heart failure, and hepatosplenic T cell lymphoma, particularly in young males treated with concomitant immunomodulators [95]. A minority of patients can also develop a psoriaform-­ like rash—this is usually an anti-TNF class effect, although it can usually be managed with topical therapy. In the last decade we have seen a number of other biologics approved that act by different mechanisms. Natalizumab, a monoclonal antibody that blocks alpha-4-­ integrin and thus attachment of the activated leukocyte in the capillary at the site of inflammation, was approved for moderate to severe Crohn’s. However, its use has all but ceased owing to concerns with development of progressive multifocal leukoencephalopathy (PML), which is rare but frequently fatal brain infection [96, 97]. In 2014, vedolizumab was widely approved for induction and maintenance of remission in both Crohn’s and UC. It blocks alpha-4-beta-7-integrin which is gut specific and has not been associated with PML [98]. In 2016, ustekinumab was approved in many countries for the induction and maintenance of remission in Crohn’s. Ustekinumab blocks the p40 subunit of IL-12 and IL-23 [99]. In 2018, tofacitanib was approved in a number of countries for the induction and maintenance of moderate to severe UC. This small molecule, administered orally, targets the JAK-STAT pathway, specifically binding to JAK1 and JAK3 [100]. Overall, these newer biologics have had favorable tolerability and safety profiles. They are also less likely to provoke auto-antibody reaction compared to anti-TNF agents. The main potential adverse events continue to be increased risk for infection [95]. Where to position these newer biologics in relation to the anti-TNFs is a matter of considerable debate and is an area of intense study. For treatment of flares, steroids have long been a mainstay of therapy for both UC and CD, and are given in IV form to those hospitalized with severe colitis. They are effective at inducing remission but not in maintaining remission, uncommonly achieve endoscopic remission, and are associated with significant side effects, including Cushingoid features, emotional and psychiatric disturbances, insomnia, skin striae, metabolic bone disease, and accelerated atherogenesis [101, 102]. For

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these reasons they are not recommended for long-term use. Budesonide is an alternative to traditional steroids. Because of extensive first pass hepatic metabolism, this agent does not have as much systemic absorption and thus side effects compared to prednisone [103]. Antibiotics are recommended for treatment of septic complications in severe colitis, perianal disease, and postoperative recurrence of ileocolitis, and pouchitis but are otherwise not indicated for treatment of diseases [104]. There are a number of other exciting new agents in the pipeline for IBD. Phase 2 trials of etrolizumab look encouraging for UC. Similar to vedolizumab, this compound blocks lymphocyte cell adhesion by binding the beta-7 subunit, thus blocking adherence to MAdCAM-1 and E-cadherin [105]. By contrast, AJM300 is an oral formulation of an alpha-4 subunit antagonist that has a similar target to natalizumab and theoretically carries a lower risk of developing PML, although this is still a potential safety concern [106]. This agent is promising for UC, and phase 3 studies are underway. Looking at promising anti-interleukin agents, risankizumab is an IgG1 monoclonal antibody that targets the p19 subunit of IL-23; IL-23 selectivity without affecting IL-12 may potentially offer the same efficacy with an even better safety profile [107]. With regard to JAK/STAT targets, filgotinib is a JAK1 inhibitor that has encouraging phase 2 data for Crohn’s patients [108]. Finally, ozanimod works by a unique mechanism as an agonist of sphingosine-1-phosphate 1 and 5. This oral agent interferes with the migration of the lymphocyte from lymphoid organs to the blood and gut. Ozanimod upregulates receptor internalization and degradation and is currently in phase 3 study for UC [109]. Other areas of active research include the use of diet and fecal microbiota transplant to treat disease. In pediatric IBD, there is some data to suggest that exclusive enteral nutrition may induce remission [110]. Nutritional support is indicated for all patients with IBD, with the enteral route preferred when possible [111]. TPN has not shown benefit as primary therapy. With respect to FMT, the data remains mixed and further trials are ongoing [112]. Despite all attempts, some IBD patients will need surgery. Indications for surgery can include (1) medically refractory disease, (2) finding of low or high grade dysplasia that cannot be endoscopically resected, or finding of colitis associated cancer, (3) obstruction that is not amenable to endoscopic balloon dilation (Crohn’s only), and penetrating complications, particularly intraabdominal abscess formation (Crohn’s only) [32, 101]. In UC, surgery is curative with total proctocolectomy; it is usually performed with ileal pouch anal anastomosis (IPAA) [113]. In Crohn’s, surgery is not curative and disease predictably returns (usually recurrence occurs just proximal to the anastomosis in the case of ileocolonic resections) [114]. Crohn’s patients are either administered post-operative prophylaxis to prevent recurrence, or are aggressively monitored for recurrence on the basis of ileocolonoscopy [90]. Given the complicated nature of disease and the high risk of disease progression with complications, IBD management requires coordinated care from specialists across multiple disciplines, including gastroenterology, surgery, and nutrition.

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IBD Genetics: Clinical Correlates The goal of understanding the genetics of IBD is several folds, including diagnosis, predicting disease severity, evaluating response to therapy, and disease prevention [115, 116]. IBD is a genetically complex disease and does not fit within the framework of a classical Medelian disorder. Genetic facts play an important role in the pathogenesis of disease, as evidenced by the increased risk of IBD in Ashkenazi Jews. In monozygotic twin studies, the IBD concordance rate is only 15–30%, and 3–4% in dizygotic twins, thus emphasizing the important contribution of environment or epigenetic factors of disease development [117]. As more genome-wide single nucleotide polymorphism (SNP) data has become disseminated, a hypothesis free analysis of allelic frequency between cases and controls has become possible through genome wide association studies (GWAS). GWAS have identified now 232 genetic loci for IBD in Caucasians, the majority of which are shared between CD and UC, and one-third of IBD loci are found in other autoimmune diseases [118, 119]. GWAS generally is effective in targeting common disease common variants (minor allele frequency > 0.05). The genes involve an array of functions, including microbial recognition and innate immunity, lymphocyte activation and proliferation, cytokine production (interferon gamma, IL-12, TNF-alpha and IL-10 signaling), JAK/STAT signaling, autophagy, and intestinal epithelial defense [115, 120]. In the latest landmark paper based on a total sample size of almost 60,000 individuals, 25 new susceptibility loci were identified, including three in the integrin genes [119]. Integrins are critical for leukocyte homing to an area of inflammation attributed to IBD. No individual gene is determining; from the latest estimates, GWAS findings only explain 26% and 19% of heritability in Crohn’s and ulcerative colitis, respectively [118]. Notably, a majority of variants discovered in GWAS are likely not functional in contributing to IBD, but rather are correlated to the causal variant through linkage disequilibrium [121]. For Crohn’s disease, NOD2 represents the strongest CD risk gene in Caucasians; three common loss of function mutations are known (R702W, G908R, and L1007fsinsC) representing 81% of disease causing mutations [122]. NOD2 was identified through a combination of linkage analysis and fine mapping in 2001. NOD2 is expressed in epithelial Paneth cells, neutrophils, and macrophages, and activates immunity against bacterial cell wall peptidoglycan. Thus, mutations in NOD2 cause a failure to protect the gut from luminal bacteria. Ten percent of healthy Caucasians carry NOD2 major variants, while 70% of CD patients have no NOD2 major variants. NOD2 confers a three-fold risk of developing Crohn’s in heterozygotes and a 20-fold increased risk for homozygotes with a 5% penetrance. By contrast mutation in IL23R provides a 2–3-fold protection against developing Crohn’s [123]. For UC, HLA is the strongest UC risk region with a 2–3 fold increased risk for carriers of HLA DRB1∗0103 [124]. Notably, it is critical to understand that many of these important mutations including NOD2, IL23R, as well as the autophagy gene ATG16L1, are important in the patients of European descent, but not in non-­ European populations. In Asians, mutations in TNFSF15 are known to confer an increased risk of Crohn’s [125]. In African Americans, a GWAS study of over 2000

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African American IBD patients recapitulated the findings from the European population, but also identified African specific SNPs at ZNF649 (transcriptional repressor) and LSAMP (axonal neuronal adhesion molecule) among UC patients [126]. By contrast, gene penetrance is much higher in the setting of very early onset IBD (VEOIBD), age 1.2. Crohn’s disease (as will be described in future chapters) benefited from a number of early GWAS discoveries, increasing the number of confirmed loci to a dozen [18–20].

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It quickly became clear that data quality control of GWAS data was essential to producing interpretable and reproducible results [21]. While the genotyping platforms produced data that were extremely high quality on average, the sheer size of the datasets compared to earlier studies meant that even very low error rates could produce spurious associations. A suite of quality control metrics, including missing data rates, Hardy-Weinberg equilibrium, and overall heterozygosity quickly became standards in GWAS analysis, and geneticists became familiar with Q-Q plots and other statistical tools as a rapid transition from genetic studies where each genotyping assay was manually inspected and scored to automated genome-wide typing technologies took place. It was also recognized that, even if genotyping data were perfect, false inference of association could arise if the ancestries of cases and controls were not well matched and the frequency differences characteristic of different populations were confounded with case-control status. Here a parallel set of methods emerged [22, 23] to measure and control for population structure in association studies that, like the QC standards, are still in wide use. Furthermore, the genetics community insisted on stringent statistical significance thresholds (p  50% Variant Chr Position Ns Signals mapped to a single variant rs7307562 12 40724960 2 rs2066844 16 50745926 10 rs2066845 16 50756540 10 rs6017342 20 43065028 2

Phe AF

Prob

INFO Func Annotation

CD CD CD UC

0.999 0.999 0.999 0.999

1 0.8 1 1

0.398 0.063 0.022 0.544

LRRK2(intronic) NOD2(R702W) NOD2(G908R) HNF4A (downstream), Gut_H3K27ac rs61839660 10 6094697 2 CD 0.094 0.999 1 E IL2RA(intronic), Immune_H3K4me1 rs5743293 16 50763781 10 CD 0.964 0.999 1 C NOD2(fs1007insC) rs6062496 20 62329099 1 IBD 0.587 0.996 1 T RTEL1-­ TNFRSF6B(ncRNA_ intronic), EBF1 TFBS rs141992399 9 139259592 3 IBD 0.005 0.995 1 C CARD9(1434+1G>C) rs35667974 2 163124637 1 UC 0.021 0.994 1 C IFIH1(I923V) rs74465132 7 50304782 3 IBD 0.034 0.994 1 T,E IKZF1(upstream), EP300 TFBS, Immune_H3K4me1 rs4676408 2 241574401 1 UC 0.508 0.994 0.99 GPR35(downstream) rs5743271 16 50744688 10 CD 0.007 0.993 1 C NOD2(N289S) rs10748781 10 101283330 2 IBD 0.55 0.990 1 E NKX2-3 (upstream), Gut_H3K27ac rs35874463 15 67457698 2 IBD 0.054 0.989 1 C,E SMAD3(I170V), Gut_H3K27ac rs72796367 16 50762771 10 CD 0.023 0.983 1 NOD2(intronic) rs1887428 9 4984530 1 IBD 0.603 0.974 0.97 JAK2(upstream) rs41313262 1 67705900 5 CD 0.014 0.973 1 C IL23R(V362I) rs28701841 6 106530330 2 CD 0.116 0.971 1 PRDM1 (upstream) Signals mapped to 2–50 variants and the lead variant has posterior probability >50% rs76418789 1 67648596 5 CD 0.006 0.937 0.59 C IL23R(G149R) rs7711427 5 40414886 3 CD 0.633 0.919 1 rs1736137 21 16806695 2 CD 0.407 0.879 1 rs104895444 16 50746199 10 CD 0.003 0.865 1 C NOD2(V793M) rs56167332 5 158827769 2 IBD 0.353 0.845 1 IL12B rs104895467 16 50750810 10 CD 0.002 0.833 1 C NOD2(N852S) rs630923 11 118754353 2 CD 0.153 0.820 0.98 rs3812565 9 139272502 3 IBD 0.402 0.815 1 Q eQTL of INPP5E in CD4 and CD8; CARD9 in CD14 rs4655215 1 20137714 3 UC 0.763 0.784 1 E Gut_H3K27ac rs145530718 19 10568883 3 CD 0.023 0.762 0.97 rs6426833 1 20171860 3 UC 0.555 0.752 1 chr20: 20 43258079 2 CD 0.041 0.736 0.88 43258079 rs17229679 2 199560757 2 UC 0.028 0.716 1 C C E

(continued)

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Table 2 (continued) Variant rs4728142 rs2143178

Chr Position Ns Phe AF Prob INFO Func Annotation 7 128573967 1 UC 0.448 0.664 1 E Immune_H3K4me1 22 39660829 2 IBD 0.157 0.662 1 T,E NFKB TFBS, Gut_H3K27ac rs34536443 19 10463118 3 CD 0.038 0.649 1 C TYK2(P1104A) rs138425259 16 50663477 10 UC 0.009 0.648 0.92 rs146029108 9 139329966 3 CD 0.036 0.643 0.92 rs12722504 10 6089777 2 CD 0.26 0.615 1 rs60542850 19 10488360 3 IBD 0.17 0.591 0.89 rs2188962 5 131770805 1 CD 0.44 0.590 1 E,Q Gut_H3K27ac, eQTL of SLC22A5 in CD14, CD15 and IL rs2019262 1 67679990 5 IBD 0.4 0.586 1 rs3024493 1 206943968 2 IBD 0.171 0.537 1 E Immune_H3K4me1 rs7915475 10 64381668 3 CD 0.304 0.528 1 rs77981966 2 43777964 1 CD 0.077 0.521 1 rs9889296 17 32570547 1 CD 0.264 0.512 1 rs2476601 1 114377568 1 CD 0.908 0.508 1 C PTPN22(W620R)

Table adapted from Nature volume 547, pages 173–178 (13 July 2017) Ns number of independent signals in the locus, Phe phenotype, AF allele frequency, Prob posterior probability for being a causal variant, INFO imputation, Func functional annotations – coding (C), disrupting transcription factor binding sites (T), overlapping epigenetic peaks (E), and colocalization with eQTL (Q)

 ucleotide-Binding Oligomerization Domain-Containing N Protein 2 (NOD2) NOD2 is a member of the Nod1/Apaf-1 family encoding a protein with 2 caspase recruitment domains (CARDs) and 11 leucine-rich repeats (LRRs). It is a key player in the immune system in mounting a reaction against infections [93]. NOD2 is also known as caspase recruitment domain-containing protein 15 (CARD15) or inflammatory bowel disease protein 1 (IBD1). As the latter name implies, NOD2 is the first gene found to be associated with polygenic IBD. In 2001, Ogura et al. [94] and Hugot et  al. [95] independently reported associations between NOD2 and CD.  The frameshift variant in NOD2 (at the second nucleotide of codon 1007) reported in both studies is under balancing selection [3]. The NOD2-centric regulation of expression network revealed several IBD genes implicated in the Mycobacterium tuberculosis response, suggesting that the host–microbe interactions might have shaped the IBD genetic architecture [3]. To date, NOD2 also hosts the greatest number of disease-causal variants among genes underlying human polygenic disorders, including missense variants R702W, G908R, N289S, V793M, and N852S. It is also worth mentioning that despite the pervasive sharing of causal variants between CD and UC, causal variants in NOD2 have a stronger penetrance in CD [85].

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Caspase Recruitment Domain Family Member 9 (CARD9) CARD9 is a member of the CARD protein family. It plays an important role in cell apoptosis and controls various innate immune responses against infections [96, 97]. A splice variant, which results in skipping of exon 11, was found to confer a protective effect toward IBD [98], possibly through the NF-κB-mediated cytokine production. CARD9 carrying this variant loses a functional C-terminal domain which is required for its recruitment of the E3 ubiquitin ligase TRIM62 [99]. Unlike risk variants, protective variants are especially helpful in designing therapeutics as new strategies can be designed by mimicking the variants’ protective effects. A study by Leshchiner et  al. [99] designed compounds that bind CARD9 and disrupt the TRIM62 recruitment, demonstrating a blueprint that CARD9 can be a path toward IBD therapeutics.

Interleukin 23 Receptor (IL23R) The protein encoded by IL23R pairs with the protein encoded by IL12RB1 to form the IL23 receptor. IL23R plays a role in the innate and adaptive immunity through activation of the Jak-Stat signaling cascade [100]. Multiple IBD causal variants were found in IL23R including R381Q, V362I, and G149R [85, 101]. These causal variants also confer protective effects. A study by Sivanesan et al. [102] suggested that IL23R carrying these variants has reduced protein stability and a reduced phosphorylation of the downstream STAT proteins.

Interferon Induced with Helicase C Domain (IFIH1) IFIH1 gene encodes Melanoma Differentiation-Associated Protein 5 (MDA5) which is part of the RIG-I-Like receptor family and is a cytoplasmic viral RNA receptor involved in activating type I interferon signaling. IFIH1 plays an important role in sensing the viral infection and mounting the antiviral response [103]. The V923I variant in IFIH1 was found to be the IBD causal variant. This variant was also found to be strongly protective in type-1 diabetes [104] with a strong effect (odds ratio close to 0.5), possibly because of the reduced production of interferon-­beta in the variant carriers [105]. Peisley et al. found the interferon production was reduced because the mutant form of this protein has a shorter binding domain for the formation of the dimer and is less stable and can be more quickly degraded [106].

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Interleukin-2 Receptor Alpha-Chain (IL2RA) IL2RA encodes CD25, a trans-membrane protein on activated T cells and B cells. Together with IL2RB and IL2RG, IL2RA form the receptor for IL2 [107]. An intronic variant, rs61839660, was found to be causal to IBD. This variant confers protection to type-1 diabetes [108] and increases the levels of CD25 expressing T cells [109].

 others Against Decapentaplegic, Drosophila Family Member 3 M (SMAD3) SMAD3 is a member of the SMAD family and a signal transducer mediating multiple signaling pathways [110]. Two SMAD3 variants were found to be causal to IBD: a coding variant at I170V and a variant disrupting the AP-1 binding motif that is highly conserved across vertebrates [84]. ChIP-seq data from HeLa cells from ENCODE [111], which are heterozygous at this site, showed major allelic imbalance with almost non-existent binding to the CD risk allele.

Functional Landscape of Polygenic IBD Causal Variants As discussed in the previous section, a large number of IBD causal variants disrupt protein-coding as frameshift, splicing, or missense variants. In aggregation, we saw more than tenfold enrichment of such variants as IBD causal variants comparing with the synonymous variants (Fig. 5a). Despite the strong contribution of coding variants, the majority of IBD causal variants are in the noncoding genome. Previous studies [85] found significant contributions from the variants disrupting the binding sites of transcription factors (Fig.  5a). An example was discussed as the SMAD3 causal variant disrupting the AP-1 binding in the previous section [84, 85]. In addition, Huang et al. [85] also found genetic contributions to IBD from regions marked by the histone H3K4me1 and H3K27ac (Fig. 5b–d). The IBD causal variants implicate the histone marked regions in a tissue-specific manner, with the causal variants preferentially implicating the H3K4me1 marked regions in the immune cell lines and the H3K27ac marked regions in the GI tissues. Huang et al. [85] also looked into variants regulating the gene expression levels (expression quantitative trait loci – eQTLs) from large-scale peripheral blood samples and found, surprisingly, a quite modest enrichment of such variants in the IBD causal variants. While it has been speculated that eQTL variants might account for complex disease genetics [112, 113], it does not appear to be the case for the eQTL inferred from the peripheral blood in IBD. However, when eQTLs from specific tissues

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Fig. 5  Functional annotation of causal variants. (a) Proportion of credible variants that are protein-­coding disrupt/create TFBS or are synonymous, sorted by posterior probability. (b). Epigenetic peaks overlapping credible variants in cell and tissue types from the Roadmap Epigenomics Consortium39. Significant enrichment has been marked with asterisks. Proportion of credible variants that overlap (c) core immune peaks for H4K4me1 or (d) core gut peaks for H3K27ac (Methods). In (a), (c), and (d), the vertical dotted lines mark 50% posterior probability, and the horizontal dashed lines show the background proportions of each functional category. (Figure and legend adapted from Nature volume 547, pages 173–178 (13 July 2017))

were used, the enrichment improved even if the sample size dropped. For example, significant enrichment was found for eQTL variants from the CD4+ cell lineage and ileum and rectum tissues using 100–400 samples (comparing with 2000–8000 samples from whole blood). This suggests that the path forward to elucidate the contribution of eQTLs to IBD has to be construed in a tissue or cell line-­specific manner with larger numbers of samples and more complete catalog of tissues and cell lines.

Causal Variants of Low Frequency The strategy we described typically works well for variants that have greater than 1% AF. For IBD causal variants in the EUR population, because of the large sample size in the study and reference panel, and the ultra-high-density chip (e.g., ImmunoChip), several causal variants can be detected and tested at as low AF as 0.1%. Alleles that have large effects on disorders are under strong natural selection pressure; therefore, such alleles are less prevalent than variants that have smaller effects. Rare variants also tend to have less variants in LD so it is naturally easier to tell causal variant from other variants in LD. To identify causal variants below 0.1% AF, large-scale sequencing efforts are inevitably the best strategy. Using targeted sequencing, Rivas et al. found IBD causal variants in NOD2 and CARD9 in 2011 [98] and in RNF186 in 2016 [114]. An ongoing IBD exome sequencing effort has accumulated over 10,000 IBD cases and 20,000 controls. Preliminary results are available through a portal at: https://ibd.broadinstitute.org. Such studies need

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large samples for statistical power. One solution to increase the power to find disease-­causal rare variants is to perform the study in bottlenecked populations [115] such as the Finnish or Jewish populations. Due to the bottleneck effect, variants that have low frequency in other population can be more prevalent in these populations. This strategy has identified additional IBD causal variants in the Ashkenazi Jewish population [116].

Concluding Remarks We have discussed the causal variants for polygenic IBD from the angle of cutting-­ edge genomic technologies and large-scale samples accumulated in the past years. Among many genes, NOD2, IL23R, and CARD9 host the strongest IBD causal variants disrupting the corresponding protein-coding sequences. Although we observed a clear excess of coding variants among IBD causal variants, this observation does not imply that pathogenic mechanism in IBD mostly involves this type of DNA changes. Possibly, coding variants in IBD have a much stronger effect compared to noncoding variants (Fig. 6) because changes in the protein structure are more functionally disruptive than changes in the regulation (level of expression) of proteins. As a result, coding variants are easier to be mapped due to the statistical power. Although the effect is weaker, noncoding IBD causal variants are important because

Fig. 6  Distribution of OR for coding and noncoding IBD causal variants

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they are more numerous than coding causal variants, and in aggregation, the noncoding variants account for a greater heritability of IBD. Huang et al. resolved many noncoding IBD loci to a small subset of causal variants and leveraged comprehensive functional resources to understand their functional implications. Still, close to half of these well-resolved IBD loci have unknown function, highlighting our lack of knowledge of the noncoding genome. To complete our understanding of the IBD causal variants, cell line- or tissue-specific resources are needed at largescale level. Finally, the IBD causal variants discussed in this chapter were all derived from samples of Caucasian ancestry. Therefore, these findings may be restricted from patients in the rest of the world. For genetics to benefit everyone, and to complete our knowledge of the IBD genetics (as other populations may host causal variants not present in Europeans), IBD genetics resources from Africa, Asia, South America, and other populations are needed [117]. The diversity of the genomes from additional populations can also help to achieve a better resolution in mapping variants in LD to smaller subsets [90].

References 1. Uhlig HH, Schwerd T, Koletzko S et al (2014) The diagnostic approach to monogenic very early onset inflammatory bowel disease. Gastroenterology 147:990–1007.e3 2. Liu JZ, van Sommeren S, Huang H et al (2015) Association analyses identify 38 susceptibility loci for inflammatory bowel disease and highlight shared genetic risk across populations. Nat Genet 47:979–986 3. Jostins L, Ripke S, Weersma RK et  al (2012) Host-microbe interactions have shaped the genetic architecture of inflammatory bowel disease. Nature 491:119–124 4. Franke A, McGovern DPB, Barrett JC et al (2010) Genome-wide meta-analysis increases to 71 the number of confirmed Crohn’s disease susceptibility loci. Nat Genet 42:1118–1125 5. Anderson CA, Boucher G, Lees CW et  al (2011) Meta-analysis identifies 29 additional ulcerative colitis risk loci, increasing the number of confirmed associations to 47. Nat Genet 43:246–252 6. Uhlig HH, Muise AM (2017) Clinical genomics in inflammatory bowel disease. Trends Genet 33:629–641 7. Uhlig HH, Muise AM (2017) Clinical genomics in inflammatory bowel disease. Trends Genet: TIG 33:629–641 8. Crowley E, Muise A (2018) Inflammatory bowel disease: what very early onset disease teaches us. Gastroenterol Clin N Am 47:755–772 9. Ruemmele FM, El Khoury MG, Talbotec C et  al (2006) Characteristics of inflammatory bowel disease with onset during the first year of life. J Pediatr Gastroenterol Nutr 43:603–609 10. Paul T, Birnbaum A, Pal DK et al (2006) Distinct phenotype of early childhood inflammatory bowel disease. J Clin Gastroenterol 40:583–586 11. Griffiths AM (2004) Specificities of inflammatory bowel disease in childhood. Best Pract Res Clin Gastroenterol 18:509–523 12. Heyman MB, Kirschner BS, Gold BD et al (2005) Children with early-onset inflammatory bowel disease (IBD): analysis of a pediatric IBD consortium registry. J Pediatr 146:35–40 13. Muise AM, Xu W, Guo CH et al (2012) NADPH oxidase complex and IBD candidate gene studies: identification of a rare variant in NCF2 that results in reduced binding to RAC2. Gut 61:1028–1035

Sequencing and Mapping IBD Genes to Individual Causative Variants and Their…

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14. Dhillon SS, Fattouh R, Elkadri A et al (2014) Variants in nicotinamide adenine dinucleotide phosphate oxidase complex components determine susceptibility to very early onset inflammatory bowel disease. Gastroenterology 147:680–689 e2 15. Dhillon SS, Mastropaolo LA, Murchie R et al (2014) Higher activity of the inducible nitric oxide synthase contributes to very early onset inflammatory bowel disease. Clin Transl Gastroenterol 5:e46 16. Hayes P, Dhillon S, O’Neill K et al (2015) Defects in NADPH oxidase genes and in very early onset inflammatory bowel disease. Cell Mol Gastroenterol Hepatol 1:489–502 17. Ruel J, Ruane D, Mehandru S, Gower-Rousseau C, Colombel JF (2014) IBD across the age spectrum: is it the same disease? Nat Rev Gastroenterol Hepatol 11:88–98 18. Benchimol EI, Mack DR, Nguyen GC et al (2014) Incidence, outcomes, and health services burden of very early onset inflammatory bowel disease. Gastroenterology 147:803–813 e7; quiz e14–5 19. Kotlarz D, Marquardt B, Baroy T et al (2018) Human TGF-beta1 deficiency causes severe inflammatory bowel disease and encephalopathy. Nat Genet 50:344–348 20. Parlato M, Charbit-Henrion F, Pan J et al (2018) Human ALPI deficiency causes inflammatory bowel disease and highlights a key mechanism of gut homeostasis. EMBO Mol Med (2018)10:e8483 21. Leung G, Muise AM (2018) Monogenic intestinal epithelium defects and the development of inflammatory bowel disease. Physiology (Bethesda) 33:360–369 22. Samuels ME, Majewski J, Alirezaie N et al (2013) Exome sequencing identifies mutations in the gene TTC7A in French-Canadian cases with hereditary multiple intestinal atresia. J Med Genet 50:324–329 23. Chen R, Giliani S, Lanzi G et al (2013) Whole exome sequencing identifies TTC7A mutations for combined immunodeficiency with intestinal atresias. J  Allergy Clin Immunol 132:656–664 e17 24. Avitzur Y, Guo C, Mastropaolo LA et al (2014) Mutations in tetratricopeptide repeat domain 7A result in a severe form of very early onset inflammatory bowel disease. Gastroenterology 146:1028–1039 25. Agarwal NS, Northrop L, Anyane-Yeboa K, Aggarwal VS, Nagy PL, Demirdag YY (2014) Tetratricopeptide repeat domain 7A (TTC7A) mutation in a newborn with multiple intestinal atresia and combined immunodeficiency. J Clin Immunol 34:607–610 26. Bigorgne AE, Farin HF, Lemoine R et al (2014) TTC7A mutations disrupt intestinal epithelial apicobasal polarity. J Clin Invest 124:328–337 27. Lemoine R, Pachlopnik-Schmid J, Farin HF et  al (2014) Immune deficiency-related enteropathy-­ lymphocytopenia-alopecia syndrome results from tetratricopeptide repeat domain 7A deficiency. J Allergy Clin Immunol 134:1354–1364 e6 28. Guana R, Garofano S, Teruzzi E et al (2014) The complex surgical management of the first case of severe combined immunodeficiency and multiple intestinal atresias surviving after the fourth year of life. Pediatr Gastroenterol Hepatol Nutr 17:257–262 29. Woutsas S, Aytekin C, Salzer E et al (2015) Hypomorphic mutation in TTC7A causes combined immunodeficiency with mild structural intestinal defects. Blood 125:1674–1676 30. Yang W, Lee PP, Thong MK et al (2015) Compound heterozygous mutations in TTC7A cause familial multiple intestinal atresias and severe combined immunodeficiency. Clin Genet 88:542–549 31. Kammermeier J, Lucchini G, Pai SY et al (2016) Stem cell transplantation for tetratricopeptide repeat domain 7A deficiency: long-term follow-up. Blood 128:1306–1308 32. Lien R, Lin YF, Lai MW et al (2017) Novel mutations of the tetratricopeptide repeat domain 7A gene and phenotype/genotype comparison. Front Immunol 8:1066 33. Lawless D, Mistry A, Wood PM et al (2017) Bialellic mutations in tetratricopeptide repeat domain 7A (TTC7A) cause common variable immunodeficiency-like phenotype with enteropathy. J Clin Immunol 37:617–622

136

A. Muise and H. Huang

34. Neves JF, Afonso I, Borrego L et al (2018) Missense mutation of TTC7A mimicking tricho-­ hepato-­enteric (SD/THE) syndrome in a patient with very-early onset inflammatory bowel disease. Eur J Med Genet 61:185–188 35. Mandiá N, Perez-Muñuzuri A, Lopez-Suarez O Congenital intestinal atresias with multiple episodes of sepsis. undefined 2018 36. Fayard J, Collardeau S, Bertrand Y et  al (2018) TTC7A mutation must be considered in patients with repeated intestinal atresia associated with early inflammatory bowel disease: two new case reports and a literature review. Arch Pediatr 25:334–339 37. Fullerton BS, Velazco CS, Hong CR, Carey AN, Jaksic T (2018) High rates of positive severe combined immunodeficiency screening among newborns with severe intestinal failure. JPEN J Parenter Enteral Nutr 42:239–246 38. Jardine S, Dhingani N, Muise AM (2019) TTC7A: steward of intestinal health. Cell Mol Gastroenterol Hepatol 7:555–570 39. Glocker EO, Kotlarz D, Boztug K et al (2009) Inflammatory bowel disease and mutations affecting the interleukin-10 receptor. N Engl J Med 361:2033–2045 40. Glocker EO, Frede N, Perro M et al (2010) Infant colitis—it’s in the genes. Lancet 376:1272 41. Kotlarz D, Beier R, Murugan D et  al (2012) Loss of interleukin-10 signaling and infantile inflammatory bowel disease: implications for diagnosis and therapy. Gastroenterology 143:347–355 42. Glocker EO, Kotlarz D, Klein C, Shah N, Grimbacher B (2011) IL-10 and IL-10 receptor defects in humans. Ann N Y Acad Sci 1246:102–107 43. Engelhardt KR, Shah N, Faizura-Yeop I et al (2013) Clinical outcome in IL-10- and IL-10 receptor-deficient patients with or without hematopoietic stem cell transplantation. J Allergy Clin Immunol 131:825–830 e9 44. Neven B, Mamessier E, Bruneau J et al (2013) A Mendelian predisposition to B-cell lymphoma caused by IL-10R deficiency. Blood 122:3713–3722 45. Shouval DS, Ebens CL, Murchie R et  al (2016) Large B-cell lymphoma in an adolescent patient with interleukin-10 receptor deficiency and history of infantile inflammatory bowel disease. J Pediatr Gastroenterol Nutr 63:e15–e17 46. Marlow GJ, van Gent D, Ferguson LR (2013) Why interleukin-10 supplementation does not work in Crohn’s disease patients. World J Gastroenterol 19:3931–3941 47. Nauseef WM (2008) Biological roles for the NOX family NADPH oxidases. J Biol Chem 283:16961–16965 48. Heyworth PG, Cross AR, Curnutte JT (2003) Chronic granulomatous disease. Curr Opin Immunol 15:578–584 49. Marks DJ, Miyagi K, Rahman FZ, Novelli M, Bloom SL, Segal AW (2009) Inflammatory bowel disease in CGD reproduces the clinicopathological features of Crohn’s disease. Am J Gastroenterol 104:117–124 50. Schappi MG, Smith VV, Goldblatt D, Lindley KJ, Milla PJ (2001) Colitis in chronic granulomatous disease. Arch Dis Child 84:147–151 51. Werlin SL, Chusid MJ, Caya J, Oechler HW (1982) Colitis in chronic granulomatous disease. Gastroenterology 82:328–331 52. Bennett CL, Christie J, Ramsdell F et al (2001) The immune dysregulation, polyendocrinopathy, enteropathy, X-linked syndrome (IPEX) is caused by mutations of FOXP3. Nat Genet 27:20–21 53. Okou DT, Mondal K, Faubion WA et al (2014) Exome sequencing identifies a novel FOXP3 mutation in a 2-generation family with inflammatory bowel disease. J Pediatr Gastroenterol Nutr 58:561–568 54. Rigaud S, Fondaneche MC, Lambert N et al (2006) XIAP deficiency in humans causes an X-linked lymphoproliferative syndrome. Nature 444:110–114 55. Worthey EA, Mayer AN, Syverson GD et al (2011) Making a definitive diagnosis: successful clinical application of whole exome sequencing in a child with intractable inflammatory bowel disease. Genet Med 13:255–262

Sequencing and Mapping IBD Genes to Individual Causative Variants and Their…

137

56. Aguilar C, Lenoir C, Lambert N et al (2014) Characterization of Crohn disease in X-linked inhibitor of apoptosis-deficient male patients and female symptomatic carriers. J Allergy Clin Immunol 134:1131–1141 57. Zeissig Y, Petersen BS, Milutinovic S et al (2015) XIAP variants in male Crohn’s disease. Gut 64:66–76 58. Ashton JJ, Andreoletti G, Coelho T et al (2016) Identification of variants in genes associated with single-gene inflammatory bowel disease by whole-exome sequencing. Inflamm Bowel Dis 22:2317–2327 59. Schubert D, Bode C, Kenefeck R et al (2014) Autosomal dominant immune dysregulation syndrome in humans with CTLA4 mutations. Nat Med 20:1410–1416 60. Gamez-Diaz L, August D, Stepensky P et al (2016) The extended phenotype of LPS-responsive beige-like anchor protein (LRBA) deficiency. J Allergy Clin Immunol 137:223–230 61. Serwas NK, Kansu A, Santos-Valente E et al (2015) Atypical manifestation of LRBA deficiency with predominant IBD-like phenotype. Inflamm Bowel Dis 21:40–47 62. Lo B, Zhang K, Lu W et  al (2015) Autoimmune disease. Patients with LRBA deficiency show CTLA4 loss and immune dysregulation responsive to abatacept therapy. Science 349:436–440 63. van de Geer A, Nieto-Patlan A, Kuhns DB et al (2018) Inherited p40phox deficiency differs from classic chronic granulomatous disease. J Clin Invest 28:3957–3975 64. Lehle AS, Farin HF, Marquardt B et  al (2019) Intestinal inflammation and dysregulated immunity in patients with inherited caspase-8 deficiency. Gastroenterology 156:275–278 65. Cuchet-Lourenco D, Eletto D, Wu C et al (2018) Biallelic RIPK1 mutations in humans cause severe immunodeficiency, arthritis, and intestinal inflammation. Science (New York, NY) 361:810–813 66. Li Y, Fuhrer M, Bahrami E et al (2019) Human RIPK1 deficiency causes combined immunodeficiency and inflammatory bowel diseases. Proc Natl Acad Sci U S A 116:970–975 67. Kammermeier J, Dziubak R, Pescarin M et al (2017) Phenotypic and genotypic characterisation of inflammatory bowel disease presenting before the age of 2 years. J Crohns Colitis 11:60–69 68. Kammermeier J, Drury S, James CT et al (2014) Targeted gene panel sequencing in children with very early onset inflammatory bowel disease--evaluation and prospective analysis. J Med Genet 51:748–755 69. Ostrowski J, Paziewska A, Lazowska I et al (2016) Genetic architecture differences between pediatric and adult-onset inflammatory bowel diseases in the Polish population. Sci Rep 6:39831 70. Murugan D, Albert MH, Langemeier J et al (2014) Very early onset inflammatory bowel disease associated with aberrant trafficking of IL-10R1 and cure by T cell replete haploidentical bone marrow transplantation. J Clin Immunol 34:331–339 71. Charbit-Henrion F, Jeverica AK, Begue B et al (2017) Deficiency in mucosa-associated lymphoid tissue lymphoma translocation 1: a novel cause of IPEX-Like syndrome. J  Pediatr Gastroenterol Nutr 64:378–384 72. Booth C, Gilmour KC, Veys P et al (2011) X-linked lymphoproliferative disease due to SAP/ SH2D1A deficiency: a multicenter study on the manifestations, management and outcome of the disease. Blood 117:53–62 73. Rohr J, Pannicke U, Doring M et al (2010) Chronic inflammatory bowel disease as key manifestation of atypical ARTEMIS deficiency. J Clin Immunol 30:314–320 74. Cuvelier GD, Rubin TS, Wall DA, Schroeder ML (2016) Long-term outcomes of hematopoietic stem cell transplantation for ZAP70 deficiency. J Clin Immunol 36:713–724 75. Ngwube A, Hanson IC, Orange J et al (2018) Outcomes after allogeneic transplant in patients with Wiskott-Aldrich syndrome. Biol Blood Marrow Transplant 24:537–554 76. Chiriaco M, Salfa I, Di Matteo G, Rossi P, Finocchi A (2016) Chronic granulomatous disease: clinical, molecular, and therapeutic aspects. Pediatr Allergy Immunol 27:242–253 77. Kato K, Kojima Y, Kobayashi C et al (2011) Successful allogeneic hematopoietic stem cell transplantation for chronic granulomatous disease with inflammatory complications and severe infection. Int J Hematol 94:479–482

138

A. Muise and H. Huang

78. Freudenberg F, Wintergerst U, Roesen-Wolff A et al (2010) Therapeutic strategy in p47-phox deficient chronic granulomatous disease presenting as inflammatory bowel disease. J Allergy Clin Immunol 125:943–946 e1 79. Uzel G, Orange JS, Poliak N, Marciano BE, Heller T, Holland SM (2010) Complications of tumor necrosis factor-alpha blockade in chronic granulomatous disease-related colitis. Clin Infect Dis 51:1429–1434 80. Canna SW, Girard C, Malle L et al (2017) Life-threatening NLRC4-associated hyperinflammation successfully treated with IL-18 inhibition. J Allergy Clin Immunol 139:1698–1701 81. Weinacht KG, Charbonnier LM, Alroqi F et al (2017) Ruxolitinib reverses dysregulated T helper cell responses and controls autoimmunity caused by a novel signal transducer and activator of transcription 1 (STAT1) gain-of-function mutation. J  Allergy Clin Immunol 139:1629–1640 e2 82. Schaid DJ, Chen W, Larson NB (2018) From genome-wide associations to candidate causal variants by statistical fine-mapping. Nat Rev Genet 19:491–504 83. Goode EL (2011) Linkage disequilibrium. In: Schwab M (ed) Encyclopedia of cancer. Springer Berlin Heidelberg, Berlin, Heidelberg, pp 2043–2048 84. Farh KK-H, Marson A, Zhu J  et  al (2015) Genetic and epigenetic fine mapping of causal autoimmune disease variants. Nature 518:337–343 85. Huang H, Fang M, Jostins L et al (2017) Fine-mapping inflammatory bowel disease loci to single-variant resolution. Nature 547:173–178 86. Mahajan A, Taliun D, Thurner M et al (2018) Fine-mapping type 2 diabetes loci to single-­ variant resolution using high-density imputation and islet-specific epigenome maps. Nat Genet 50:1505–1513 87. Davey JW, Hohenlohe PA, Etter PD, Boone JQ, Catchen JM, Blaxter ML (2011) Genome-­ wide genetic marker discovery and genotyping using next-generation sequencing. Nat Rev Genet 12:499 88. Marchini J, Howie B (2010) Genotype imputation for genome-wide association studies. Nat Rev Genet 11:499–511 89. Jostins L (2012) Using next-generation genomic datasets in disease association. The University of Cambridge 90. Lam M, Chen C-Y, Li Z et al (2018) Comparative genetic architectures of schizophrenia in East Asian and European populations. bioRxiv:445874 91. Kichaev G, Pasaniuc B (2015) Leveraging functional-annotation data in trans-ethnic fine-­ mapping studies. Am J Hum Genet 97:260–271 92. Hormozdiari F, van de Bunt M, Segrè Ayellet V et al (2016) Colocalization of GWAS and eQTL signals detects target genes. Am J Hum Genet 99:1245–1260 93. Caruso R, Warner N, Inohara N, Nunez G (2014) NOD1 and NOD2: signaling, host defense, and inflammatory disease. Immunity 41:898–908 94. Ogura Y, Bonen DK, Inohara N et al (2001) A frameshift mutation in NOD2 associated with susceptibility to Crohn’s disease. Nature 411:603 95. Hugot JP, Chamaillard M, Zouali H et al (2001) Association of NOD2 leucine-rich repeat variants with susceptibility to Crohn’s disease. Nature 411:599–603 96. Zhong X, Chen B, Yang L, Yang Z (2018) Molecular and physiological roles of the adaptor protein CARD9 in immunity. Cell Death Dis 9:52 97. Cao Z, Conway KL, Heath RJ et  al (2015) Ubiquitin ligase TRIM62 regulates CARD9-­ mediated anti-fungal immunity and intestinal inflammation. Immunity 43:715–726 98. Rivas MA, Beaudoin M, Gardet A et  al (2011) Deep resequencing of GWAS loci identifies independent rare variants associated with inflammatory bowel disease. Nat Genet 43:1066–1073 99. Leshchiner ES, Rush JS, Durney MA et al (2017) Small-molecule inhibitors directly target CARD9 and mimic its protective variant in inflammatory bowel disease. Proc Natl Acad Sci U S A 114:11392–11397

Sequencing and Mapping IBD Genes to Individual Causative Variants and Their…

139

100. Buonocore S, Ahern PP, Uhlig HH et al (2010) Innate lymphoid cells drive interleukin-23-­ dependent innate intestinal pathology. Nature 464:1371–1375 101. Duerr RH, Taylor KD, Brant SR et  al (2006) A genome-wide association study identifies IL23R as an inflammatory bowel disease gene. Science 314:1461 102. Sivanesan D, Beauchamp C, Quinou C et  al (2016) IL23R (interleukin 23 receptor) variants protective against inflammatory bowel diseases (IBD) display loss of function due to impaired protein stability and intracellular trafficking. J Biol Chem 291:8673–8685 103. Takeuchi O, Akira S (2008) MDA5/RIG-I and virus recognition. Curr Opin Immunol 20:17–22 104. Nejentsev S, Walker N, Riches D, Egholm M, Todd JA (2009) Rare variants of IFIH1, a gene implicated in antiviral responses, protect against type 1 diabetes. Science 324:387–389 105. Chistiakov DA, Voronova NV, Savost’Anov KV, Turakulov RI (2010) Loss-of-function mutations E6 27X and I923V of IFIH1 are associated with lower poly(I:C)-induced interferon-beta production in peripheral blood mononuclear cells of type 1 diabetes patients. Hum Immunol 71:1128–1134 106. Peisley A, Lin C, Wu B et  al (2011) Cooperative assembly and dynamic disassembly of MDA5 filaments for viral dsRNA recognition. Proc Natl Acad Sci U S A 108:21010–21015 107. Liao W, Lin JX, Leonard WJ (2013) Interleukin-2 at the crossroads of effector responses, tolerance, and immunotherapy. Immunity 38:13–25 108. Huang J, Ellinghaus D, Franke A, Howie B, Li Y (2012) 1000 Genomes-based imputation identifies novel and refined associations for the Wellcome Trust Case Control Consortium phase 1 Data. Eur J Hum Genet 20:801–805 109. Orru V, Steri M, Sole G et al (2013) Genetic variants regulating immune cell levels in health and disease. Cell 155:242–256 110. Roberts AB, Russo A, Felici A, Flanders KC (2003) Smad3: a key player in pathogenetic mechanisms dependent on TGF-beta. Ann N Y Acad Sci 995:1–10 111. Consortium EP (2012) An integrated encyclopedia of DNA elements in the human genome. Nature 489:57–74 112. Nica AC, Montgomery SB, Dimas AS et al (2010) Candidate causal regulatory effects by integration of expression QTLs with complex trait genetic associations. PLoS Genet 6:e1000895 113. Lappalainen T, Sammeth M, Friedlander MR et  al (2013) Transcriptome and genome sequencing uncovers functional variation in humans. Nature 501:506–511 114. Rivas MA, Graham D, Sulem P et al (2016) A protein-truncating R179X variant in RNF186 confers protection against ulcerative colitis. Nat Commun 7:12342 115. Hatzikotoulas K, Gilly A, Zeggini E (2014) Using population isolates in genetic association studies. Brief Funct Genomics 13:371–377 116. Kenny EE, Pe’er I, Karban A et al (2012) A genome-wide scan of Ashkenazi Jewish Crohn’s disease suggests novel susceptibility loci. PLoS Genet 8:e1002559 117. Popejoy AB, Fullerton SM (2016) Genomics is failing on diversity. Nature 538:161–164

Genetic Risk Prediction in IBD Urko M. Marigorta

Abstract  An emerging objective in the IBD community is to gear the findings from genome-wide association studies towards achieving precision medicine. The main goal of this new model of disease management is to adapt clinical practice to the needs of each patient. This includes, among others, obtaining more detailed characterizations of disease presentation at diagnosis, better prediction of disease prognosis that permits to anticipate and adapt management to variations in symptomatology, and tailoring treatment to individual needs. In this chapter, we will introduce the methodology for estimation of disease risk using genetic data from individuals and discuss the potential and main challenges for using genomic risk profiling as a predictive tool that can pave the way for adoption of precision medicine-based approaches in the clinical management of IBD.

Calculation of Risk Scores for Genomic Profiling in IBD The main goal of genomic risk profiling lies in estimating genetic liability for disease in each person. As multifactorial traits, the architecture of common diseases such as IBD is composed by large yet undetermined numbers of risk variants. This idea is the inspiration for the liability threshold model, the main paradigm used in current genetics research on complex traits [1, 2]. This model posits that causal factors involved in disease susceptibility form a continuous distribution of risk that is at the root of binary disease outcomes such as, for instance, suffering from IBD. In most cases, the inherent physiological robustness of biological systems outweighs the ill action of genetic and environmental risk factors, and hence most individuals do not develop disease. However, in a small fraction of individuals, there is an excess of risk factors whose effects add up beyond some critical threshold that leads to disease. U. M. Marigorta (*) Integrative Genomics Lab, CIC bioGUNE, Bizkaia Science and Technology Park, Derio, Biscay, Spain IKERBASQUE, Basque Foundation for Science, Bilbao, Spain e-mail: [email protected] © Springer Nature Switzerland AG 2019 C. Hedin et al. (eds.), Molecular Genetics of Inflammatory Bowel Disease, https://doi.org/10.1007/978-3-030-28703-0_7

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Under the threshold liability paradigm, actual risk of disease in each individual can be summarized into a single numeric value that sums up the joint effects of all the myriad of factors associated with IBD risk [3]. In practice, genetic liability is estimated through genetic risk scores (GRSs) that pool together the effects of susceptibility single nucleotide polymorphisms (SNPs) discovered by GWAS.  In its simplest form, GRSs are calculated by summing up the total risk allele dosage that each individual genome harbours. The resulting score is labelled as ‘unweighted GRS’ because all variants are assumed to contribute equally and no differential weighting is applied. In contrast, ‘weighted GRSs’ incorporate information about the effect size by multiplying the allele dosage at each variant by the logarithm of the odds ratio from the discovery of genome-wide association study (GWAS). Although the latter method is more appropriate considering that risk effects can vary among susceptibility variants, in practice most genome-wide significant discoveries have similar odds ratios, and therefore unweighted and weighted estimates tend to be comparable. Of note, given that the majority of susceptibility alleles discovered by GWAS are common (i.e. minor allele frequency >5%), all individuals inevitably carry a number of risk alleles, and therefore GRS distributions are continuous and normally distributed. A key decision for GRS calculation lies in deciding which risk variants are included in the calculation. The most frequent practice consists of incorporating only SNPs that pass strict genome-wide significance thresholds in large discovery GWAS (i.e. P