NIAMS Roundtable on Arthritis Biology Research

Wednesday, December 7, 2011

NIH Campus, Building 45, Room D
9:00 AM – 4:00 PM

Background and Approach

The goal of this roundtable was to discuss scientific and clinical needs in arthritic diseases research. Discussion was aimed at identifying research directions that could enable a better understanding of the molecular mechanisms underlying disease pathogenesis, and, ultimately, lay the groundwork for future development of interventions that improve patient outcomes.

In advance of the meeting, participants were encouraged to consult with colleagues colleagues and provide feedback on the following key questions:

• What are new scientific questions that can be addressed in the short-term with currently available knowledge/resources?
• What opportunities exist for the application of knowledge gained from arthritis biology research (e.g., disease diagnosis, sub-phenotyping, therapeutic development, preventive interventions)?
• What genetic/molecular indicators (e.g., gene expression patterns, biomarkers) involved in the development of arthritic diseases hold the greatest promise?
• How can we better understand the impact of human translational studies on arthritic diseases, particularly as they relate to fundamental biology and the identification of biomarkers?
• What are some of the key mechanisms in the progression of arthritic diseases, both locally and systemically? What is their potential as therapeutic targets? How do they intersect with each other, or with other organ systems?
• What are the most significant gaps in knowledge that need to be addressed to fully understand the processes leading to clinical stages of arthritic diseases?
• What cutting-edge approaches can be applied to the study of arthritis biology? Are there strategies that can accelerate research and make it more productive and efficient? What novel methodologies could be used to address questions regarding early pathogenic events?
• How can involvement in arthritis biology research in the United States be expanded and sustained?

The common themes that emerged from the feedback were used to guide the discussion. These themes were: the microbiome; mechanisms of disease onset and progression; biomarkers; interactions of tissues; and integration of the data obtained with common tools into a biosignature.

Time was also reserved to discuss strategies to maintain the vitality of the scientific enterprise in an era of fiscal challenges.

Although only a subset of topics were discussed in depth and are summarized here, NIAMS leadership and the appropriate program staff read each comment. The NIAMS greatly appreciates the community’s input on these questions.

Summary

Which basic and translational research areas in arthritis biology are critical to advance the field?

The Microbiome

Current genetic discoveries resulting from genome-wide association studies (GWAS) explain only about 20% of the disease variance of rheumatoid arthritis (RA), while the heritability of RA is estimated to be at approximately 60%. Although genes contribute to RA susceptibility, interaction between gene effects and environmental factors contribute to disease development. It has long been postulated that autoimmune disorders, including RA, are triggered by microorganisms, and recent evidence suggests that microorganisms inhabiting the human body, particularly those residing in the oral and gut cavities, have a role in the development of RA.

In the human body, microbes outnumber human cells by about tenfold, and contribute to healthy physiological development. However, up to now, many microbes could not be sequenced. To understand the role of the microbiome in disease, the complex interactions between microbes and host, as well as those among different microbe species, need to be studied. The collected information also needs to be placed in the context of environment and diet. Moreover, it is critical to correlate the understanding of the microbiome, immunophenotype, and immune response for both patients and “normal” individuals. One of the challenges in such studies is that the diversity of the microbiome among healthy individuals is much greater than anticipated, and probably greater than the difference between healthy and “diseased” individuals. In order to facilitate these studies, the NIH’s Common Fund Human Microbiome Project (HMP) has enabled the cataloguing of microbiota distribution at several body sites in normal individuals. Data have emerged that, despite the huge number of species and great variation among individuals, the microbiota composition of the human gut can be classified into at least three distinct groups, or enterotypes. There is evidence that a single species change can tip the balance between healthy and diseased, and that this shift could be due to the genetic predisposition of an individual.

Emerging technologies, including transformative advances in imaging and DNA sequencing, continue to facilitate advances in the field. For instance, enhancements in cytotechnology have enabled the phenotyping of cells in a much more granular way than ever before, which allows fingerprinting of exposures to assess how environmental challenges shape the immune system. Clearly, a coordinated effort among researchers is necessary to share information obtained from studies on host/microbiome interactions among the research community. To this end, it may be possible to apply technology used in other large studies such as the Cooperative Centers for Translational Research on Human Immunology and Biodefense program, which supports a centralized infrastructure for multi-disciplinary research on human immunological responses. Similar to studies on the human microbiome, a large challenge right now with understanding the human immune response at the level of integrating immune cell signatures is that an individual’s immunosignature varies tremendously even over short periods of time, making it difficult to integrate this information throughout an individual’s lifetime.

Challenging questions to be addressed include: at what stage, and how does, the microbiome contribute to disease? Infection is frequently associated with the onset of arthritis, yet only a small number of patients progress to chronic arthritis. It is not clear why these organisms hone in on the joint, and how chronicity of infection impacts the immune (both innate and adaptive) responses. The inability to measure environmental stressors is a huge roadblock in understanding arthritis pathogenesis in humans; measuring the microbiota may be a way to quantitate environmental exposures. Although not easy to do, studying the microbiome of a “birth cohort” (sampling over several body sites starting at birth and continuing for several years) may prove to be extraordinarily valuable in understanding early microbiome acquisition and environmental influences, including diet and geographical location, in the microbiome’s development.

Recent studies have indicated that microbes residing in the oral cavity and gut may be drivers in the development of RA. For example, P. gingivalis, a bacterium implicated in periodontal disease, has the capacity to produce citrullinated proteins, leading to the production of anti-CCP antibodies that contribute to RA. Also, certain gut bacteria are able to induce inflammatory Th17 cells that lead to arthritis in a mouse model of RA. In an ongoing study on the role of the oral and gut microbiome in RA, American Recovery and Reinvestment Act (ARRA) funded researchers aim to correlate the microbes present in oral and stool samples with immune cell subsets (Th17 and Treg cells) in RA patients. Understanding the mechanisms of these arthritogenic organisms, and the regulation of Th17 cells in RA, may help in developing new therapies for RA. In regard to the gut microbiome, it would be interesting to study arthritis patients who develop inflammatory bowel disease (IBD).

The role of the microbiome in juvenile idiopathic arthritis (JIA) has not yet been explored. Although the triggers of JIA are unknown, infections are postulated to be a component of the disease. Many questions remain about the difference in the age of onset for JIA versus RA, and why JIA patients can spontaneously outgrow their disease, providing fertile ground for future research on the interplay between genetics, the microbiome, and environmental factors in JIA.

Mechanisms of Disease Onset and Progression

Increased understanding of the pathogenesis of inflammatory arthritides at the cellular and molecular levels has resulted in much progress in the treatment of these diseases. Research continues on both immune cells (T cells, B cells, macrophages, neutrophils, dendritic cells, etc.) and non-immune lineages (such as synovial fibroblasts and platelets) to understand their roles in joint inflammation and tissue destruction. In addition, recent studies on the role of antibodies to citrullinated protein antigens (ACPAs), smoking, and periodontal disease in RA point to the important relationship between potential genetic and environmental risk factors in the development of RA. Discovery of ACPAs has transformed our thinking on RA pathogenesis; however, it is still not clear whether ACPAs are a primary driver of disease, or an amplification event. While it is known that ACPAs and certain blood markers can precede disease by several years, it is not well-understood what triggers a change from the sub-clinical presence of autoantibodies to an arthritic disorder. Further studies are needed to better understand how ACPAs develop and their mechanism of action in RA.

The link between systemic autoimmunity and/or inflammation and local joint disease remains quite elusive. Some evidence shows that systemic immunity acts locally in the joint; for example, a normal joint has waves of citrullination that occur due to micro-trauma, which could result in an autoimmune response. Additional evidence from mouse models demonstrates that circulating antigens can be deposited in the joint, which becomes the focus of immune complexes. Specific characteristics of the cartilage surface, such as charge, may contribute to such deposition. Moreover, local changes in the vasculature of the joint gives immune cells access to the joint. Currently, there is still a very limited understanding of the unique characteristics of the joint structure which may play a role in the immune process. Furthermore, we need a better understanding of how innate immune variability and adaptive immune dysfunction work together in the progression of arthritis, and the biological process differences between acute and chronic synovitis. A confounding issue in arthritis research is that arthritis is a common final pathway of several processes; one has to consider this heterogeneity when studying the pathogenic mechanisms.

Researchers have used various approaches, including animal models and patient tissues, to study arthritis; it is critical to have access to patient tissues at early stages of disease, as well as normal tissues for comparison. Although patients’ (including pediatric patients in the CARRA registry) willingness to participate in studies has increased over time, collection of enough samples will continue to be a limiting factor. Alternative non-invasive techniques like intra-vital imaging should be further developed for phenotyping, as well as for pathogenesis studies, to complement conventional invasive methods. While mouse models have provided invaluable insights to the processes of the immune system and disease pathogenesis, they clearly have shortcomings in the study of human arthritis. For instance, HLA associations are not easily elucidated with outbred populations. Also, it has recently been demonstrated that, in contrast to what was seen with transgenic mice, healthy humans seem to all have autoreactive T cells, calling into question accepted tenets on mechanisms of centralized tolerance. Thus, merely measuring the frequency of autoreactive cells may not yield differences between healthy and diseased populations. Whether phenotypic differences exist in T cells of patients who have disease and those who do not also needs to be determined. Use of tetramers, a transforming tool in immunology research, may lead the way in addressing these questions.

Studies in ankylosing spondyloarthritis (AS) have been hampered due to the lack of an animal model that adequately recapitulates disease, as well as the difficulty in obtaining spinal tissue from AS patients. Studies of AS patients have shown that suppressing inflammation, through a TNF inhibitor, for instance, does not necessarily slow disease. Ongoing studies are assessing whether disease course and/or bone destruction can be altered by inhibiting TNF much earlier in disease (even before the condition meets disease criteria). There has been emerging interest in the concept of a synovio-entheseal complex, emphasizing the interdependence between synovial membrane and entheses. These studies require interdisciplinary collaborations between bioengineers and biologists.

There are several avenues of research on arthritic disease onset and progression that could be addressed in the short-term. One is the study of immune regulatory mechanisms, such as those associated with regulatory T and B cells, which have been shown to be important in other autoimmune diseases like multiple sclerosis. IL-10 production has been shown to be the key mechanism of Breg cells in humans, but more work is needed to understand the role of Breg cells in disease. Another fertile ground is the study of the role of vascularization in the RA joint. Several cell types are implicated in promoting angiogenesis; for example, both macrophages and synovial fibroblasts have the ability to sense hypoxia in the joint and produce pro-inflammatory cytokines, as well as angiogenic factors, and patients with elevated levels of Th17 cells have more vascularized disease. In addition, migration of monocytes into the arthritic joint likely plays a role in the chronic stage of arthritis. Furthermore, understanding how cytokines really signal, and whether the signaling cascades in the target cells are amenable to interference through small molecule drugs, presents a great opportunity in the field of therapeutics, particularly because of the high cost of biologics. Finally, what do we learn from treated patients (e.g., those treated with anti-cytokine therapies) is a fundamental question that needs to be addressed, not only to improve our understanding of disease mechanisms, but also to help in the development of new therapies.

What are the impediments to translation of basic research to patients/the public?

Biomarkers

Biomarkers can be employed to diagnose patients early in the disease process, to improve the understanding of the mechanism of action of treatments used, and to predict responsiveness to therapy and thereby guide clinicians in establishing a treatment course. IFN signatures, serologic markers like anti-CCP, and ANA, are some examples of biomarkers that have taught us about disease pathogenesis. From a clinical perspective, molecular guidance on how and when to use biologic therapies is lacking; reliable biomarkers allowing prediction of therapeutic responsiveness would increase treatment success and reduce safety concerns. Some obstacles to the use of biomarkers in arthritic disease treatment include the enormous heterogeneity when comparing individual patients, and the lack of an accurate quantification of the biology of disease. For instance, pain, an important feature of arthritis, is extremely difficult to measure quantitatively. In addition, flare, relapse, and disease remission all need to be defined on a molecular level. Only after these parameters are in place can one begin to link the changes in biomarker(s) to the disease outcome. Another consideration for arthritis treatment is that even if joint symptoms are halted, systemic modifications/co-morbidities (e.g., cardiovascular disease associated with RA) also need to be monitored. In distinguishing important, reliable biomarkers from a large number of candidates, applied mouse studies should be coupled with human studies. There is still a need for the discovery of new biomarkers, as well for the re-examination of “old” biomarkers that may have been previously dismissed due to lack of specificity or sensitivity. Emerging technologies in genetic techniques and imaging will promote these studies; for example, novel imaging techniques may allow for inflammation to be assessed on the single-cell level in individuals. On the other hand, clinical trials have already created sub-phenotypes of patients (e.g., responders vs. non-responders), which are valuable resources to further investigate and evaluate disease biomarkers.

Understanding Interactions of Tissues

Arthritic diseases involve multiple organs and tissues; there is a great need to better understand the interactions of tissues and cells, as well as the cross-talk between local and systemic manifestations. Almost all arthritides have co-morbidities, such as RA with cardiovascular disease, but the pathogenesis of these co-morbidities is not well-understood. Furthermore, arthritis therapies may have different impacts on co-morbidities.

One of the major areas of research has focused on immune cells - how they act towards each other, but also with other cells and tissues in the settings of normal and pathogenic immune responses. The immune response in primary and secondary lymphoid tissue may be very different than that in other tissues, where immune cells utilize microvasculature to migrate to sites of inflammation. Furthermore, it has been recognized that age affects the human immune system, which changes dramatically as we go through life. Some clinical consequences of an aging immune system (immunosenescence) include poor responses to new antigens or vaccinations, increased infection rates with higher morbidity and mortality, and increased incidence of autoimmune diseases. It has been shown that telomere shortening, a chromosomal feature associated with cellular aging, is accelerated in immune cells from RA patients; these and other studies have placed telomere dynamics as a focus of interest in understanding the relationship among aging, immunosenescence, and autoimmunity. Premature immunosenescence is suggested to play a role in the pathogenesis of JIA and adult RA. Currently, it is not known whether premature immunosenscence drives the development of autoimmune conditions, or is secondary to chronic inflammatory processes. Neither is it known if an aging immune system can be repaired, or if interference with the inflammatory process will then interfere with aging. These questions, among others, highlight the need for population-based longitudinal studies on immune-risk factors for autoimmune diseases.

What critical resources and new technologies are needed to move the field forward? What are the best ways to promote interdisciplinary collaborations?

Data Integration into Biosignatures

The discussants identified some significant knowledge gaps in fully understanding the processes leading to clinical stages of arthritic diseases:

• What do we need to know about initiators of disease?
• What are the functional roles of genetic variants in RA identified by GWAS?
• What is the impact of environmental risk factors?
• What are the temporal components of disease progression in humans?
• What factors specify phenotype of disease?
• What are appropriate methodologies to use for identification of molecular targets for therapy?
• What is the impact of inhibiting abnormal vascularization on inflammation in RA?
• What is the impact of age and the microbiome on the outcome of RA treatment?

To approach such complex, inter-related issues, it is necessary to integrate data obtained through the use of common tools, including next-generation sequencing, mRNA and protein profiling, single cell analyses, etc., into large-scale collaborations to generate biosignatures, in order to understand environmental effects and disease mechanisms. Towards these goals, several new collaboration-intensive research approaches and technologies have emerged recently, including epigenetics, systems biology, functional genomics, and mass flow cytometry. In this post-fluorescence era, mass flow cytometry technology now enables interrogation of up to 45 parameters on a single cell. This new technology allows use of smaller samples from humans, and increases understanding of cell expression and differentiation in functional studies. One could use a tandem approach to first identify cell phenotypes using this technology, and then sort out the desired cells for further studies. Finally, through the use of these and other technological advances, the idea of “synovitis on a chip” may be achievable in the near future.

Overall, these cutting-edge approaches can be readily applied to the study of arthritis biology. In addition, the strategies of integrating human disease studies and clinical trials, forming global partnerships in establishing biorepositories, and making databases easier to create, obtain, and share, will definitely accelerate arthritic diseases research.

Maintaining the vitality of the scientific enterprise in an era of fiscal challenges

Given the interest in maintaining robust research programs and the scientific work force during the current budget climate, time was reserved for participants to discuss ideas outlined in the NIH Office of Extramural Research “Rock Talk” blog and a presentation on “Ways of Managing NIH Resources.”

Arthritis Biology Roundtable Discussion participants raised several community concerns and suggestions that may lead to cost savings. Ultimately, the group agreed that there were several priorities that must be protected during tight fiscal times. In addition to the investigator-initiated R01, it was generally felt that the pipeline of new investigators should be protected. This takes the form of training and mentoring awards, but also partnerships with outside organizations that provide bridge funding.

Core facilities with shared infrastructure were also seen as critical because no single investigator can afford these resources individually. Throughout the day, many expressed the need for greater collaboration and coordination between researchers across disciplines and organizations.