Short-term cyclophosphamide therapy in a cohort of over 200 Patients

Investigators:
Alexandra Villa-Forte, MD and Gary S. Hoffman, MD

Location:
Center for Vasculitis Care and Research, Cleveland Clinic Foundation Cleveland, Ohio

Timeline:
February 1, 2002 – February 28, 2003

Abstract
Background: Cyclophosphamide (CP) is the most effective therapy for Wegener’s Granulomatosis (WG). Drug toxicity limits long-term CP therapy. Dr. Hoffman and others have stressed the limitations of CP. Since becoming Chairman of Rheumatic and Immunologic Diseases (1992) at the Cleveland Clinic Foundation (CCF), Dr. Hoffman has provided care for over 200 patients with WG. The protocol employed for severe WG has included induction of remission with CP, followed by maintenance therapy with either Methotrexate (MTX) or Azathioprine (Aza).

Objective
To utilize a single practitioner’s large patient cohort to determine whether short-term CP therapy, followed by either MTX or Aza, provides less morbidity and mortality than conventional approaches for WG.

Specific aims
1. Determine efficacy and toxicity of short-term (3-6 months) CP, followed by an alternate remission-maintenance therapy. Comparisons with conventional therapy will be facilitated because of the senior investigator’s (GSH) role as the principal investigator for the NIH WG-CP and MTX protocols.

2. Determine whether different initial clinical presentations of WG correlate with ong-term prognosis.

3. To test the hypothesis that sustained lymphopenia is associated with increased treatment effectiveness. The finding of a significant association between the presence of lymphopenia and efficacy would result in new guidelines for monitoring and modifying drug therapy.

 

What this means for patients

This study will help determine answers to the following questions:

1. Will the milder agent, MTX, be associated with fewer long-term remissions?

2. If relapses are more common after MTX has been started, will patients still do better because of fewer side effects from less prolonged use of CP (3-6 months vs. 18 or more months)?
Treatment and outcomes-related data gathered from this study will be compared to results that have been obtained by others, who have utilized similar and dissimilar treatment approaches.

3. Will only certain patients with specific types (subsets) of WG benefit from this approach? A related goal in this analysis is to determine whether certain initial features (or combinations of features) of disease, such as persistent sinusitis vs. joint, muscle, skin, lung or kidney disease, are associated with different ultimate outcomes.

4. Does the effect of therapy on reducing the numbers of certain types of white blood cells (lymphocytes) determine the likelihood of staying in remission?

It is known that one of the most striking effects of CP, MTX or other “chemotherapy”-type drugs is in reducing lymphocyte counts and suppressing lymphocyte functions. These effects may persist more than 1 year after the treatment has been discontinued. It is very possible that the risk of WG relapse is increased if the lymphocyte count is never adequately reduced or if a patient’s counts return to normal soon after remission.

Depending on the answers, the knowledge derived may change and improve therapy.

Host-microbial interactions in Wegener’s Granulomatosis: The role of ANCA and S. aureus persistence

Investigator:
Robert D. Inman, MD

Location:
University of Toronto, Toronto Western Hospital, Toronto, Ontario Canada

Timeline:
September 1, 2002 – August 31, 2003 (extended to Spring 2004)

Abstract
There is compelling circumstantial evidence to implicate a role for infection in the pathogenesis of Wegener’s Granulomatosis (WG). Recently there has been confirmation of a higher nasal carriage of Staphylococcus aureus in WG patients. Yet it is not known whether this finding is a cause or an effect of the disease process. Secondly, there continues to be mounting evidence of the efficacy of antibiotic therapy for WG. It is unresolved however whether agents such as ulfonamide or trimethoprim function as antibiotics or by an alternative mechanism. Third, neutrophils represent the first line of defense against bacterial pathogens. The ANCA antibody response that is the hallmark of WG may influence host defenses against such infections in a fundamental way.

Specific Aims
The aims of the proposed project are to analyze the relationship between ANCA and clearance of pathogens. This will shed new light on the relationship between infection and autoantibody formation. This is one of the fundamental problems in all autoimmune disease, and the insights from this study will have important implications for other chronic inflammatory conditions, which may have an infectious trigger.

1. To define the impact of ANCA on the internalization and intracellular killing of S. aureus by neutrophils and by endothelial cells.
2. To examine the impact of ANCA on the parameters of innate immunity, particularly TLR2 and TLR4 signaling and activation of NFkB, as marker of gene transcription and cytokine production.

What this means for patients
There is compelling reason to resolve the possible role of infection in WG:
1. As has just been confirmed, WG patients have a higher prevalence of nasal carriage in S. aureus. The reason for this microbial persistence has not been resolved.
2. Antibiotics form an important part of the treatment regimen of WG, and it has been assumed that the primary mode of action of these agents is their antimicrobial effect.
3. Since the neutrophil functions as a first line of defense against pathogens such as S. aureus, it is important to resolve whether ANCA may be significantly impairing host defenses against infection by altering neutrophil function.

The role of shear stress in neutrophil proteinase-3 expression and its importance in vascular injury sites in WG

Investigator: Deborah J. Stearns-Kurosawa, Ph.D.
Location: Oklahoma Medical Research Foundation, Oklahoma City, Oklahoma
Timeline: March 1, 2003 – February 29, 2004 (extended to June 2004)

Abstract
Vascular inflammation is a hallmark of Wegener’s Granulomatosis (WG) and other primary systemic vasculitides. Vascular inflammation is also characteristic of atherosclerosis, and WG patients have increased intimal thickening, more frequent cardiovascular disease and accelerated atherosclerosis. The concurrent endothelial dysfunction and altered flow hemodynamics will induce homing and transmigration of pro-inflammatory cells, including neutrophils.

Enzymes, cytokines and receptors expressed by emigrating neutrophils contribute to the ongoing inflammation. Neutrophils express proteinase-3, an enzyme located in neutrophil granules and recognized by the autoantibody in WG patients (PR3-ANCA). In vitro experiments have demonstrated that this PR3-autoantibody interaction induces a myriad of neutrophil inflammatory events. The in vivo importance of PR3-ANCA remains controversial in part because it is not clear how PR3 becomes accessible to the autoantibodies on largely unactivated neutrophils.

However, few investigators have considered potential contributions from flow-induced shear stress. The current data demonstrate that low level shear stress results in unique surface expression of PR3 on neutrophils; other neutrophil granule markers are largely unaffected.

Our hypothesis is that low shear stress levels are important for expression of neutrophil PR3, so that it becomes accessible to pro-inflammatory PR3-ANCA. We also hypothesize that this low level shear-induced neutrophil PR3 expression plays a major role in the localization of endothelial injury, since Wegener’s granulomatosis selectively affects small-to-medium sized vessels. Experiments also are proposed to identify shear-induced signaling mechanisms to provide mechanistic insight into PR3 expression under rheologic conditions.

Collectively, these events would bring neutrophils and their expressed PR3 and PR3/PR3-ANCA complexes in close proximity to endothelial cells, providing all the players to initiate or propagate inflammatory events.

Results from proposed studies will form the foundation of future studies identifying the molecular mechanisms underlying this unique influence of shear stress on neutrophil PR3 expression.

Specific aims
These aims are designed as a 1 year project to identify the rheologic conditions that result in surface PR3 expression on neutrophils exposed to laminar shear forces and on neutrophils rolling on endothelial cells under flow. Signaling mechanisms relevant to shear-induced neutrophil PR3 expression will be explored. Additionally, the ability of macromolecules known to bind to neutrophil PR3 under static conditions, including PR3-ANCA, will be determined under flow conditions.

1. Define the laminar flow conditions that result in neutrophil PR3 expression. Neutrophils will be exposed to varying levels of laminar shear stress in a parallel plate flow chamber. Surface PR3 expression will be evaluated by flow cytometry using anti-PR3 monoclonal antibodies and patient PR3-ANCA. These results will give insight into the influence of shear stress on PR3 expression and accessibility. Other granule markers will be evaluated concurrently. The binding of molecules known to interact with PR3 (e.g., alpha 1-antitrypsin, soluble endothelial protein C receptor) will be determined similarly under flow conditions for comparison with known static systems.
2. Evaluate the impact of shear stress and activated endothelium on PR3 expression on rolling neutrophils. Neutrophils will be rolled over TNF-α activated endothelial cells at varying physiological flow velocities and PR3 expression quantitated by flow cytometry.
3. The influence of modulators (patient PR3-ANCA, a1-antitrypsin, etc.) on adhesion, rolling velocities and transmigration will be determined. Neutrophils will be harvested for evaluation of signaling events using western blotting and ribonuclease protection assays. These studies provide a physiological context in which to evaluate the influence of laminar flow forces on PR3 expression, PR3 accessibility to PR3-ANCAs, and subsequent neutrophil adhesion events.

What it means for patients
No analysis of the type proposed has ever been attempted and a body of important new information will be developed as a result of this study. The approaches described in this proposal are new and unique ways to look at the fundamental mechanisms underlying the pathogenesis of WG and related vasculitides. The long-term consequences are impossible to predict, but knowledge of mechanisms regulating our response to inflammation may provide insight into novel therapies.

Analysis of NKG2D expression on CD28

Investigators: Peter Lamprecht, MD, and Wolfgang L. Gross, MD

Location: Department of Rheumatology, University Hospital of Schleswig-Holstein, Campus Luebeck and Rheumaklinik Bad Bramstedt Luebeck and Bad Bramstedt, Germany

Timeline: January 1, 2004 – December 31, 2004

Abstract
Wegener’s Granulomatosis (WG) is a chronic inflammatory disease characterized by granulomatous lesions and an autoimmune vasculitis. Previously we have shown that circulating Th1-type so-called “late” differentiated or “effector memory” cells lacking the co-stimulatory molecule CD28 are expanded. Expansion of CD28 ¯ T-cells starts early during the disease process and correlates with organ involvement. In granulomatous lesions T-cells also generally lack CD28. Moreover, CD28¯ T-cells are the Th1-type cytokine producing T-cell population within these lesions. Data from in vitro studies suggest that CD28¯ T-cells express the co-stimulatory activating receptor NKG2D under inflammatory conditions. NKG2D interacts with the ligands MICA and MICB which are non-classical MHC-class I antigens expressed on antigen-presenting cells upon infection and cellular distress.
We hypothesize that in WG CD28¯ T-cells express NKG2D and interact with MICA and MICB expressed on antigen-presenting within granulomatous lesions. Therefore we want to determine the expression of NKG2D on peripheral blood CD28¯ T-cells by flow cytometry (n=25) and on CD28¯ T-cells within granulomatous lesions by immunohistochemistry (n=5). MICA and MICB expression on antigen-presenting cells within granulomatous lesions will be shown by immunohistochemistry (n=5).
Detection of NKG2D+CD28¯ T-cells interacting with MICA+MICB+ cells would shed new light on the nature of T-cell activation in WG and allow future detailed analysis of possible antigens involved in this process. The analysis might reveal new potential targets for a more specific therapy, for instance DAP 10, which is an adaptor protein necessary for NKG2D signaling.
What this means for patients
We see an expansion of a population of specialized, proinflammatory immune cells in WG. These specialized, proinflammatory immune cells are coined CD28¯ T-cells, because they lack a molecule, namely CD28, on their cell surface. We want to analyze whether these CD28¯ T-cells express another molecule, NKG2D, on their cell surface. The NKG2D molecule is important for the activation of CD28¯ T-cells and enables them to sense cellular distress and/or infection on surrounding cells. In that case the surrounding cells express the NKG2D ligands (counterparts) MICA and MICB on their cell surface to communicate with the CD28¯ T-cells. We want to analyze whether peripheral blood CD28¯ T-cells and CD28¯ T-cells within inflammatory lesions in nasal tissue express the NKG2D molecule on their surface and whether their ligands MICA and MICB are expressed on the cell surface of surrounding cells.
The proposed analysis will shed new light on the nature of the activation of proinflammatory immune cells in WG and might also reveal new potential targets for a more specific therapy.

Identification of Novel Wegener’s Granulomatosis Susceptibility Genes

Investigator: Katherine A. Siminovitch, MD

Location: Mount Sinai Hospital, Toronto, Ontario, Canada

Timeline: March 1, 2004 – February 28, 2005

PATIENTS MAY STILL ENROLL IN STUDY.

For more information please contact:

Alida Pokoradi, Clinical Research Coordinator

Phone: 416-946-4501 x. 3297

Fax: 416-946-4531

Email: clinicalgenet.research@gmail.com

Abstract
Wegener’s granulomatosis (WG) is a multisystem, necrotizing, granulomatous vasculitis that affects about 3 in every 100,000 people in the United States and is equally prevalent in males and females. Diagnosis of WG is difficult, and many physicians are unfamiliar with the condition. Thus, there is a significant unmet medical need for improved diagnostic tests. Development of new diagnostic and therapeutic approaches to WG has been substantially impeded by lack of understanding of the causes of WG.

Many lines of evidence suggest that a major cause of WG is infectious insult of genetically susceptible individuals. A handful of susceptibility genes have been examined as possible etiologic factors, but to date both the genetic and infectious factors underlying WG remain unknown. Identification of WG susceptibility genes would, however, provide a very promising avenue to the development of improved diagnostic and prognostic tests as well as new targets for therapeutic management. Although technologies for disease gene discovery are radically improving, the application of such technologies is highly dependent on the availability of collections of DNA samples from affected individuals and their family members.

This proposal describes a strategy to collect blood samples and prepare DNA from individuals with WG and their immediate family members, and to conduct comprehensive clinical chart review in order to obtain powerful data for results stratification and analysis. The obtained samples and clinical information will lay the foundation, and be absolutely required for, future studies to identify WG susceptibility genes.

Specific aims and goals contained within this grant:
a) To collect blood samples for DNA analysis from 500 WG patients and family members where available;
b) To review and document clinical data from these WG patients, including demographic data;
c) To prepare DNA samples for banking from the blood samples;
d) To lay the groundwork for future genetic analysis of the WG patient/family cohort

Larger goals within the framework of our WG research program (anticipated to be funded by additional national granting programs)
e) To understand the fundamental causes of WG by using genetic association and family-based linkage studies (where possible) to focus on identification of the full complement of WG susceptibility genes;
f) To develop gene-based (DNA) diagnostic assays for early identification of susceptible individuals, in conjunction with established ANCA blood testing and other diagnostic procedures;
g) To elucidate the molecular pathophysiology of WG and the nature of environmental-genetic interactions that trigger disease, and;
h) To design improved strategies for prevention and treatment of WG, including the diagnostic applications mentioned above.

Relevance to WG:
The exact cause(s) of WG are unknown, but it is generally thought to result from infection of individuals with an underlying genetic susceptibility. A variety of gene variants have been shown to be associated with WG susceptibility, including the immunologically relevant IL-10 (Interlukin 10), transforming growth factor-beta (TGFB) and the cytoxic T lymphocyte associated antigen-4 (CTLA-4) genes. Additionally, some other genes have been associated with WG, including those for the angiotensin-converting enzyme (ACE), proteinase-3 and alpha-1-antitrypsin. The tendency to relapse among WG patients has also been associated with variants of the Fcgamma receptor gene. Despite these data, the molecular and biochemical pathways underlying WG onset and progression remain largely uncharacterized.

Recent studies of the genetics of Crohn’s Disease, an inflammatory disease of the gastrointestinal tract, and of the rare Blau syndrome, have demonstrated an association with variants of the NOD2 gene bacterial species. Because both Crohn’s Disease and Blau syndrome are associated with granuloma development, and because granulomatous disease in Crohn’s Disease patients are specifically associated with NOD2 gene variants, we recently undertook a genetic study in WG patients (Newman et al. (2003)., J. Rheuma. 30(2):305-7. While we did not detect any association between WG and the NOD2 gene, our results illustrate our general strategy in examining candidate genes that may be involved in WG, and further emphasize the necessity of much larger, family-based genetic studies in research into the genetic underpinnings of this disease. Accordingly, we propose to greatly expand our current collection of DNA samples from WG patients and their family members, in order to lend greater power to future genetic association and family-based linkage studies. We anticipate working closely with the WGA to achieve these goals.

Long-range objective:
The long-range goal of our WG research program is to identify genes that confer susceptibility to WG.

Progress to date:
Over the past three years, our group has focused on collection of patient and family samples required to map and isolate genes that cause and/or modulate WG. To date, patients have been ascertained mainly from the Canadian Wegener’s Granulomatosis Support Group (WGSG), and the relevant family members have been ascertained via telephone contact initiated with the probands’ consent. To allow for case-control and family-based association as well as linkage analysis, we have collected the following types of materials: individual patients for whom no other family members are available (singletons) and who can be used for case-control association studies; patients and their parents and/or unaffected siblings (trio families), who can be used for family-based association studies; and multicase families such as affected siblings and their parents (sib-pair family) or patient and affected as well as unaffected parent (relative-pair family), who can be used for linkage analysis.
Collection involves obtaining blood samples for DNA and lymphoblastoid cell line preparation and detailed demographic data on all study participants and detailed clinical data on all affected individuals.

Research Plan
The objective of this proposal is to collect sufficient numbers of WG patients and family members to lay the groundwork for identifying WG susceptibility genes. This proposal is therefore entirely focused on patient recruitment. However, in the future we will use these patient materials to carry out a genome wide scan of WG families and to examine and analyze any other genes that are discovered in the future to be associated with granulomatous disease and/or granuloma formation. Data from these studies will provide the framework for further studies aimed at identifying candidate and ultimately the relevant WG susceptibility genes.

As noted above, the patient collections possible in eastern Canada are largely completed. However, significant additional patient sample resources are required to ensure that sufficient clinical materials are available, for example, for large-scale case-control association and linkage analyses. We propose to work closely with, and seek the assistance of, the WGA in ascertaining and recruiting volunteer patients and family members interested in participating in this study.

To allow for the use of several complementary methods for WG gene identification, we will continue to collect WG patients/families of varying pedigree structures. These include:

• Multicase families: These families, which will be used for linkage analysis, contain a minimum of two affected siblings and their parents (sib-pair family) or an affected parent-child combination and an unaffected parent and/or sibling (relative-pair family).
• Trio families: These families include patients and their parents (or minimally, a patient and a single parent) as well as patients and unaffected siblings.
• Singletons: Individual patients, for whom family members cannot be recruited are also being collected and will be used for case-control association studies to facilitate candidate gene analysis once chromosomal regions of interest are identified. As most of the patients collected represent singletons, we anticipate that at least 500 such individuals will be recruited to the study by the end of year 1 and these individuals, together with any individuals drawn from the trio and multicase families, will provide a substantive resource for analysis of the association between selected SNPs/genes and WG.

Conclusion:
The identification of disease susceptibility genes has attracted considerable interest in recent years, an interest that builds upon the wealth of genetic information and improved genotyping technologies emanating from the Human Genome Project research. Definition of WG susceptibility genes will not only elucidate the combination of genes which influence disease progression, severity and response to treatment, but will also pave the way for identifying individuals at risk, delineating the specific environmental factors which help convert genetic susceptibility to disease, and designing strategies for intervention and disease prevention. Even in advance of development of new therapeutic drugs, our investigations should provide DNA-based diagnostic tools that will complement and enhance current ANCA and other clinical testing procedures, resulting in earlier diagnosis of WG and the opportunity to widen the opportunity for therapy.

Environmental Factors in the Pathogenesis of Wegener’s Granulomatosis

Investigator:
Daniel A. Albert, MD, Professor of Medicine

Location:
University of Pennsylvania School of Medicine, Division of Rheumatology Philadelphia, Pennsylvania

Timeline:
January 1, 2005 – December 31, 2005

Abstract:
Wegener’s Granulomatosis (WG) is an autoimmune inflammatory disease of the sinus pulmonary tract and other organs of unknown etiology. Both allergic (sinusitis) and infectious (staphylococcus aureus) factors are thought to play a role, but other factors remain to be elucidated. Previous studies have focused on environmental exposures that might provide further stimulation for this autoimmune process. These studies suggest that exposure to heavy metals (mercury and possibly lead), aromatic hydrocarbons, and silica may precede the onset of Wegener’s and that one possible source is through contaminated ground water in wells. This study will assess these risk factors in greater detail through a questionnaire survey of patients with Wegener’s versus controls with either gout or osteoarthritis. In addition, we will directly measure heavy metal exposure through a blood test that will quantify body burden of lead, mercury, cadmium, and arsenic and other heavy metals. These data will provide new information regarding environmental exposures that may be casually linked to Wegener’s Granulomatosis.

This is a pilot study that will provide preliminary data necessary to justify an RO1 application. We plan to utilize this data to support a proposal to examine the role of these factors in animal models of vasculitis and in future studies on patients with Wegener’s Granulomatosis.

Localized Wegener’s Granulomatosis: Epidemiological, clinical and histopathological characteristics

Investigators: Peer Malte Aries, MD and Wolfgang L. Gross, MD

Location: Department of Rheumatology, University Hospital of Schleswig- Holstein, Campus Lübeck and Rheumaklinik Bad Bramstedt, Germany

Timeline: January 1, 2005 – December 31, 2005

Abstract
The existence of different clinical phenotypes within Wegener’s granulomatosis (WG) has been recognized for several decades. So far, it is unknown why some patients remain in the early, localized phase of the disease with the predominance of granulomatous lesions of the upper and/or lower respiratory tract, whereas in most patients disease progresses to early systemic or generalized WG.

Specific aim
The aim of this study is to examine the clinical, histological and immunological (phenotypes of leukocytes subsets, activation markers, proteinase 3-expression and Th1/Th2 cytokine profile) features of localized compared to generalized WG. We hypothesize, that a Th1-type cytokine expression in serum, together with a lower PR3 expression on T cells will help us to distinguish localized from generalized WG. These findings may help us to establish diagnosis of WG earlier, even if symptoms are restricted to the upper and/or lower respiratory tract like in localized WG.

Detection of distinguishing features of localized WG will give us the opportunity to start treatment early in the course of disease; even before major organ involvement has been established. Apart from diagnostical assistance the analysis might reveal new potential targets for a more specific and stage adopted therapy.

Funding for the proposed project will enable us to get a closer insight into the mechanisms of the initial course of the disease -also called localized WG- and give us new support in the clinical management of those patients. Early disease detection and prevention of disease progression is a major interest of the WGA. This data are essential and necessary for the following application for a larger grant funded by the Deutsche Forschungsgemeinschaft (German Research Society – the NIH equivalent in Germany) to further support.

The ultimate goal of this research is to identify immunological and histological features distinguishing the early granulomatous phase of WG from its subsequent generalized vasculitic stage. A closer understanding of the immunological characteristics distinguishing localized versus generalized WG will help us to develop a more stage related therapy that may prevent further progression to the generalized vasculitic disease.

Wegener’s Granulomatosis and Microscopic Polyarteritis Case Control Study in Western Montana

Investigators: Helen Emery, MD, Andrew Zeft, MD, and Peggy Schlesinger, MD

Location: University of Washington/Children's Hospital and Regional Medical Center Section of Rheumatology, Seattle, Washington

Timeline: August 1, 2004 – July 30, 2005

Abstract:
Etiologic factors for Wegener’s Granulomatosis (WG) and Microscopic Polyarteritis (MPA) have not been identified, although it is possible that specific environmental exposures may trigger disease onset in a susceptible host. Previous studies have shown an increased risk of these disorders in patients with certain environmental exposures, the clearest association being with occupational silica.
In the last ten years, the Montana rheumatology community has observed an increased incidence and case clustering of both WGA and MPA in certain counties of western Montana. The mining industry and areas with known high heavy metal concentrations make this region unique.

Specific aim
To evaluate the relative risk of Wegener’s Granulomatosis and Microscopic Polyarteritis in a defined population in Western Montana, focusing on residential and occupational exposure to silica, heavy metals, and pesticides.

We will identify and characterize all WG and MPA cases in western Montana onset from January 1993 to June 2004 by case report from Montana rheumatologists, pulmonologists, nephrologists, and otolaryngology physicians. Using hospital records, we will identify two controls matched by age, gender, race, and distance to the hospital of index WG or MPA cases. We will interview index and control cases for residential and occupational exposures to silica, heavy metals, and pesticides.

Development of a Measure to Assess Patients’ Adherence to Vasculitis Treatment Regimens

Investigator: Robert F. DeVellis, Ph.D.

Location: Arthritis Multidisciplinary Clinical Research Center; Department of Health Behavior & Health Education, School of Public Health; Department of Psychology, University of North Carolina at Chapel Hill, North Carolina

Timeline: September 1, 2004 – August 31, 2005

Abstract:
Individuals who are diagnosed with Wegener’s Granulomatosis (WG) and other forms of ANCA-associated systemic vasculitis (AASV) face a daunting array of psychosocial challenges. One challenge not previously studied is patients’ adherence to treatment regimens. The lack of research is troublesome, given that non-adherence to treatment regimens is common across all chronic illnesses, and could lead to ineffective treatment, serious complications, and even death in patients with these illnesses.
Complicating the study of treatment non-adherence in these patients is the lack of an existing self-report measure that can reliably, validly, and conveniently measure adherence to the self-care behaviors specifically recommended by physicians to AASV patients.

This study will characterize the behavioral demands of WG treatment regimens and the barriers individuals face in trying to follow them, develop and evaluate a self-report measure of WG treatment adherence, and describe the tendencies of patients to adhere to the different behavioral recommendations in WG treatment regimens.

Toward this end, we will survey a total of 335 adults living with WG or other forms of AASV. In Phase 1, we will interview 20 patients about the self-care behaviors required by their treatment regimens and the barriers and facilitators they experience in trying to perform them. The Phase 1 data will help inform the development of a self-report adherence measure to be piloted with clinical experts and 15 patients in Phase 2. In Phase 3, 300 patients will complete a revised draft of the adherence measure and their data will be analyzed to determine the measure’s psychometric properties.

What this means for patients
Through this research, we hope to gain insight into the challenges that patients face in trying to adhere to their treatment regimens, and facilitate future investigations by developing a convenient measure of treatment adherence specific to WG and related disorders.

A Pilot Project towards Establishment of a US/Canadian Diagnostic Registry of Children with WG and Related Vasculitides

Investigator: David A. Cabral, MBBS

Location: BC Children’s Hospital and University of British Columbia, Vancouver, Canada

Timeline: March 1, 2005 – February 28, 2006

Abstract
The presentation and natural history of pediatric Wegener’s Granulomatosis (pWG) may differ significantly from adult disease; and pWG may be more difficult to distinguish from other related types of chronic vasculitis. One-third of pWG does not fulfill adult-derived classification criteria and two-thirds of chronic vasculitis in children is unclassifiable. Because study of pWG is limited by rarity of the disease, we propose to set up a pilot pediatric registry of all chronic small-medium vessel vasculitis to determine the validity, and specificity of adult classification criteria for use in pWG; feasibility of establishing an international (US/Canada) registry will thus be assessed during operation of the pilot registry and by survey of potential physician participants. In the long term, a registry will permit detailed characterization of pWG and identify homogeneous subgroups of patients to enhance pathophysiological understanding and treatment of the disease.

The first phase of the study will electronically survey pediatric rheumatologists throughout US/Canada who are members of CARRA (Childhood Arthritis and Rheumatology Research Alliance) to determine feasibility of establishing and maintaining an international registry for the short or long term; assessment will include estimates of patient numbers, local resources and commitment to participation.

Secondly, by Delphi survey of the membership we will determine optimal diagnostic features and disease markers for pWG, to be included in the pilot, and proposed international registry; this is a well established consensus formation methodology to assimilate ‘expert’ opinion (weighted for experience) through the use of sequential, iterative questionnaires rationalized by interim analysis.

Finally, information on all cases of pWG and related vasculitides over the preceding five years will be collected from CARRA centers in the Pacific-Northwest, entered into the ‘electronic’ pilot registry, and analyzed against adult-derived criteria. Demands on participating centers will be simultaneously evaluated as part of the feasibility assessment.

Specific aim
The ultimate aim of this project is to identify the need and feasibility for a valid, secure, web-based international (US/Canadian) registry of children with Wegener’s Granulomatosis and related small-medium vessel chronic vasculitis that has the ongoing commitment of all pediatric rheumatology centers to ensure reliable collection of diagnostic and monitoring data.

Renal transplantation in WG: mortality, allograft outcomes, and incidence of malignancies from the United States

Investigator: Nadine D. Tanenbaum, MD

Location: School of Medicine, Duke University Medical Center Durham, North Carolina

Timeline: July 1, 2005 - July 1, 2006

Abstract
Background: Wegener's Granulomatosis (WG) is a rare systemic vasculitis that can affect almost any organ. Renal involvement occurs in at least 80% of patients. Kidney transplantation is a treatment option for those who reach end-stage renal disease. Small studies have demonstrated the success of transplantation in this population. Multiple aspects, however, regarding the long-term course of WG patients who undergo renal transplantation remain unknown. Information on the long-term allograft and patient survival in large numbers of patients, as well as on the incidences of acute allograft rejection and posttransplant malignancies, is lacking.
Hypotheses:
1. Patient and allograft survival may be different between WG transplant recipients and transplant recipients without WG.
2. WG transplant recipients and transplant recipients without WG may have different incidences of acute allograft rejection.
3. Patients with WG may be at greater risk than patients without WG for the development of posttransplant malignancies.
4. Specific risk factors can be identified that predispose certain WG transplant recipients to poorer outcomes.

Methods and Aims: Data will be extracted from the United Network for Organ Sharing database on 883 patients with WG who have undergone a total of 929 renal transplantations from January 1, 1988 through February 28, 2005. A group of 2787 patients matched for age, ethnicity, history of prior transplant, time of transplant, and center will serve as the control group. The aims of this study are to compare long-term patient and allograft survival rates, as well as to compare the incidences of acute allograft rejection and posttransplant malignancies between these two groups. Another aim is to identify predictors of adverse outcomes in the WG cohort.

Significance: Characterizing the course of WG transplant recipients and identifying risk factors for adverse outcomes may help us better define how to care for this population of patients.
The goal of this study is to examine the clinical course of a large population of patients with Wegener's granulomatosis (WG) who have undergone renal transplantation. In a preliminary data search on the United Network for Organ Sharing (UNOS) website, 883 WG patients in the United States from January 1, 1988 through February 28, 2005 have been identified who have received a total of 929 kidney transplants.
Specific aims
1. To compare long-term patient and renal allograft survival in renal transplant recipients with and without WG.
2. To compare the incidence of acute allograft rejection in renal transplant recipients with and without WG.
3. To determine the incidence and types of posttransplant malignancies in WG transplant recipients and to evaluate whether this risk is elevated compared to transplant recipients without WG.
4. To identify predictors for patient and renal allograft death, malignancy, and WG allograft recurrence in WG transplant recipients.

Characterizing the clinical course of WG transplant recipients and identifying risk factors for adverse outcomes may help us better define how to follow and care for this population of patients.

Methods
WG is a rare systemic anti-neutrophil cytoplasmic antibody (ANCA)-associated small vessel vasculitis that can affect almost any organ. Untreated, the one year mortality rate is 80% [1]. Renal involvement, characterized by a pauci-immune necrotizing crescentic glomerulonephritis (GN), occurs in at least 80% of patients at some point in their course [2-4]. Between 11-43% of patients with WG or ANCA-associated GN develop end-stage renal disease (ESRD) and require chronic hemodialysis [3,5-8]. Among patients with ANCA-associated GN, the need for dialysis at diagnosis is a predictor of mortality with a significant difference in 5-year survival rates between patients with ESRD (61%) and those with preserved renal function (87%) [7].

In WG patients who reach ESRD, renal transplantation is treatment option. Case reports and small observational studies have demonstrated the success of transplantation in this population [9-13]. Multiple important aspects, however, regarding the course of WG patients who undergo renal transplantation remain unknown. Information is lacking regarding the long-term renal and patient survival in large numbers of patients, the risk of posttransplant malignancies, and the optimal immunosuppressive regimens. A better characterization of the incidence of disease recurrence and acute allograft rejection in this population is also needed.

Patient and renal allograft survival
One aim of this study is to compare differences in mortality and allograft survival between WG patients and patients without WG who receive a kidney transplant. The age of transplant recipients in this country is increasing. Patients older than 50 year now receive 47% of cadaveric transplants [14]. It is therefore likely that in the future clinicians will be confronted more frequently with the decision of whether renal transplantation in the often elderly WG patient is appropriate and whether WG patients will fare as well as patients without vasculitis. In the general transplant recipient population among patients age 60-74 years old, the estimate of additional years of life gained with a transplant compared to dialysis is about four years [15]. Small studies with fairly short follow-up in both WG patients and in patients with other types of vasculitides such as lupus suggest similar patient and graft survival when compared to patients without vasculitis [16-21]. In a group of 115 WG patients transplanted between 1982-1990, patient and allograft survival at 3 years of follow-up was similar between those with and without WG, although there was a trend towards a lower survival in the WG group [16]. Smaller studies that have followed patients up to 5 years have shown similar equivalent results [17-19]. However, the latter studies have consisted of less than 20 patients with either microscopic polyangiitis or WG [17-19]. Given its large cohort size and long length of follow-up, the proposed investigation is uniquely suited to address this issue in more detail. Furthermore, unlike most prior studies, this proposal includes patients only with WG. The course of renal disease in WG may differ from that associated with microscopic polyangiitis (MPA) or renal-limited vasculitis (RLV) [7].

Acute allograft rejection
Another aim of this study is to evaluate whether WG patients are at increased risk for acute allograft rejection. Acute allograft rejection is a major risk factor for chronic allograft nephropathy and ultimate graft loss [22-24]. Established risk factors for acute allograft rejection are the following: Black race, cadaveric transplant, second transplant recipients, elevated panel-reactive antibodies, increasing degree of HLA antigen mismatches, and possibly viral infections most notably CMV [25]. It is possible that ANCAs may cause subclinical neutrophil activation in WG transplant recipients, causing subsequent elevated levels of cytokines that might augment allograft rejection. Alternatively, the occurrence of rejection may predispose to recurrent allograft WG by cytokine-induced priming of ANCA antigens in the allograft. Demonstrating that WG transplant recipients have an elevated risk of acute allograft rejection in adjusted analyses would lend support to the idea that ANCA antibodies might be pathogenic in the rejection process and further studies, such as ANCA titer monitoring, could be proposed.

Malignancy in transplantation and WG
Patients with WG and ANCA-associated vasculitis are at increased risk for malignancies, especially bladder, skin, hematological, and possibly renal cell malignancies [26-33]. Much of this risk has been attributed to cyclophosphamide use. Of 1065 WG patients from a Swedish registry, 13% of deaths were secondary to a malignancy [27]. In some studies, the risk of bladder cancer in patients treated with cyclophosphamide has been about 30-fold greater than expected [26,31]. Other studies without available history on cyclophosphamide use have reported a 5-fold increase in bladder cancer risk [28]. The link between vasculitis and malignancies in general is incompletely understood. WG patients not treated with cyclophosphamide may also be at increased risk for malignancies. For example, Tatsis et. al. reported either preceding or concurrent diagnosis of malignancy in 23 patients also diagnosed with WG [32]. It is conceivable that there is a common genetic susceptibility for development of both WG and malignancy. It is also possible that the chronic immune stimulation of vasculitis predisposes to malignancy or inversely, that neoantigen exposure in malignancy may trigger the development of WG.

Malignancy is more common in all renal transplant recipients compared to age- and gender-matched controls, in large part secondary to the intensity of the immunosuppressive regimen. Skin cancers, particularly squamous cell skin cancers, are most common. There is also an increased risk of lymphomas, especially non-Hodgkin’s lymphomas, as well as renal cell carcinomas, Kaposi sarcoma, anogenital and hepatocellular carcinomas. Interestingly, there is only a mild increase in the risk of more common cancers such as lung, prostate, and colorectal cancers [34,35]. Clear risk factors for the development of malignancies after transplantation are sun exposure (for skin cancers), history of pretransplant malignancies, increasing age, Epstein Barr virus (EBV) negative to positive seroconversion at the time of transplantation and exposure to antilymphocytic therapy (the latter two are risk factors mainly for lymphomas) [34,35]. Higher maintenance levels of cyclosporine and tacrolimus, lower CD4 counts (for skin cancer) and increasing length of dialysis pretransplant have also been implicated as risk factors [34,35]. Recent studies have suggested that the immunosuppressive medicine sirolimus, which binds to FK binding protein and modulates the mammalian target of rapamycin (mTOR) resulting in eventual cell cycle arrest in the G1-S phase, has antiproliferative properties and has been associated with less posttransplant malignancies. Unlike tacrolimus and cyclosporine, sirolimus blocks cytokine-driven T- and B-cell activation [36-38].

Understanding the risk of posttransplant malignancy in WG transplant recipients is another goal of this study. It is important to define this risk in order to identify whether specific posttransplant screening is warranted in this population. To the best of the principal investigator's knowledge, only one prior study with 8 WG and 7 MPA transplant recipients has investigated whether patients with ANCA-associated vasculitis are more prone to malignancies [39]. The results of this study suggest that the incidence of malignancy may be elevated. Risks of malignancies in patients with other types with vasculitis who may have received cyclophosphamide (eg. systemic lupus erythematosus (SLE) patients) have not been specifically investigated in the transplant setting.

Predictors of adverse outcomes: patient and renal allograft death, WG recurrent allograft disease, and malignancy
A final aim is to identify independent clinical predictors of four specific outcomes (patient and renal allograft death, WG recurrent allograft disease, and malignancy) in the group of WG transplant recipients. While many of these predictors are likely to be the same as in the general transplant population at large, attention here is drawn to several specific questions. In nontransplanted WG patients with newly-diagnosed GN, serum creatinine at diagnosis of renal dysfunction is the single most important predictor of renal outcome [40-42]. Presumably, this is true as well in the transplant population. Therefore, it is important to identify risk factors for WG allograft recurrence in this population in order to highlight those at risk so that an early diagnosis can be made. Recurrent disease in the allograft has been estimated to be 16%, with 40% of these cases resulting in graft failure [43]. Recurrence has been described up to 13 years after transplantation [44]. ANCA titers at transplant have not been shown to be predictive of allograft outcome or disease relapse [19,21,45]. Whether ANCA monitoring in a high risk subgroup is not known. It is conceivable but unproven that different immunosuppressive regimens may affect disease relapse rates. In one of the largest studies of 127 WG renal transplant recipients, Nachman et. al. reported a roughly 20% systemic disease relapse rate which was similar between patients treated with cyclosporine and without cyclosporine [45].

Whether disease relapse rates are lower in the era of tacrolimus, mycophenalate mofetil, and sirolimus use has not been studied. We intend to examine what is the incidence of recurrent allograft disease and whether certain immunosuppressive regimens may be associated with lower rates of recurrence. We also intend to examine whether occurrence of acute allograft rejection is an independent risk factor for recurrent disease (see also section above entitled acute allograft rejection).

Significance
• It is important to determine in a large study with long follow-up whether WG transplant recipients have equivalent patient and allograft survival to patients without WG in order to make the most appropriate decisions regarding transplantation.
• If WG is shown to be a risk factor for acute allograft rejection, this would lend support to the idea that ANCA antibodies might be pathogenic in the rejection process.
• As the age of patients receiving transplants increases and improving allograft survival necessitates longer duration of chronic immunosuppresion, it is essential to understand what, if any, the increased risk over others WG patients are at for the development of posttransplant malignancy.
• Finally, identifying independent predictors of poor outcomes in WG transplant recipients could be helpful in developing future predictive models to assist with patient care decisions.

Identification and characterization of immunodominant, conformational epitopes of anti-neutrophil cytoplasmic antibodies

Investigators: Antje Mueller, Ph.D., Elena Csernok, Ph.D. and Prof. Peter Lamprecht, MD

Location: Department of Rheumatology, University Hospital of Schleswig-Holstein, Campus Luebeck, and Rheumaklinik Bad Bramstedt, Germany

Timeline: January 1, 2006 – December 31, 2006

Abstract
Autoantibodies against human neutrophilic cytoplasmic antigens (ANCA) have been a major diagnostic hallmark for small vessel vasculitides such as Wegener`s granulomatosis (WG) since their discovery more than 20 years ago. Animal models demonstrated that ANCA targeting either myeloperoxidase or proteinase 3 (PR3 = “Wegener`s autoantigen”) induce vasculitis. Binding of ANCA to an exposed epitope of the autoantigen PR3 is one of the determining steps in their mechanism of action. Therefore, knowledge of binding epitopes on the autoantigen is necessary to understand subsequent pathologic events and develop inhibitory strategies. The aim of this study is to identify and characterize immunodominant, conformational epitopes binding to ANCA directed against proteinase 3 and/or related molecules such as other neutrophilic enzymes and complementary PR3 (cPR3). A random peptide (12-mers) library expressed in E. coli will be employed to screen for antigen-antibody interactions. We hypothesize that by screening immunoglobulin G (IgG) preparations of patients with PR3-ANCA+ WG we will find and characterize novel antigenic peptide molecules with homologies to PR3 and/or other antigens, e.g. cross-reactive antigens. This approach differs from previous experiments, because the ANCA antibodies can bind conformational, i.e. non-linear epitopes. We shall isolate peptide clones that bind to PR3-ANCA-IgG pooled from WG patients (n=25) and synthesize and further characterize such peptides. The analysis allows to detect immunodominant, conformationally expressed antigenic peptide epitopes of PR3. Blocking such epitopes and thereby subsequent events might provide a future therapeutic approach of selective intervention in WG.

Specific aims
The aim of the study is to identify and characterize immunodominant, conformational epitopes binding to anti-neutrophil cytoplasmic antibodies in WG. Such an analysis will also disclose cross-reactive antigens and related molecules (cPR3), respectively. The study might generate new tools for the detection of disease-inducing epitopes and therapeutic intervention. The study will also provide new clues for the analysis of the impact of PR3-specific T cells (reacting with linear peptides) in the pathogenesis of WG.

1. Rationale of proposal and relevance to Wegener’s Granulomatosis.
a) Background

Several linear antigenic epitopes/areas of PR3 and, recently, complementary PR3 (cPR3) binding to antibodies have been characterized, mainly by looking at proliferative responses.

b) Own studies

As inferred by renowned immunologist R.M. Zinkernagel, many autoimmune diseases display lymphoid tissue-like structures in their inflamed target organs, where autoantigen recognition and presentation is thought to be maintained. We have shown that PR3+ cells, B cells, plasma cells, T cells and potential autoantigen-presenting cells are seen in granulomatous lesions of WG patients (11,12,13), thus permitting and sustaining an ongoing (auto)immune response.

Further, we know that monoclonal antibodies directed against PR3, like WGM2, generated by our group (14), recognize conformational epitopes of PR3. In contrast, T cells recognize linear peptide sequences as shown by incubation with four linear HLA-restricted PR3 peptides resulting in the detection of TNFα-producing PR3-specific T cells in WG (15). To detect conformational epitopes binding to PR3-ANCA+ IgG we searched for an approach that allows for antibody binding to non-linear epitopes (16). By now, we have established a random peptide library screening, which can identify binding to linear and/or non-linear antigenic epitopes. Preliminary findings yielded a potentially PR3-ANCA-binding epitope that shows some consensus sequence to PR3 (depicted in bold), “SVSTVHNHETVR” (one-letter-code of amino acids). When compared to the crystal structure of PR3 (17), the contiguous area seems to be accessible for antibody binding (Fig. 2). These findings warrant further analyses of a larger patient group.
The necessity and the potential of finding relevant ANCA-binding epitopes is also supported by a study showing that the interaction of PR3 and PR3-ANCA can be targeted in an interventional approach by inhibition of PR3-ANCA binding and destruction of ANCA-producing B cells by a fusion molecule of PR3 with a toxin (18).

c) Rationale and relevance

The pathogenetic role and relevance of a PR3-specific immune response in WG is still not fully elucidated. One major open question relates to (an) antigenic motif(s) on PR3 or other potential antigens, which via binding to ANCA induce(s) the well-described chain of inflammatory events leading eventually to subsequent endothelial destruction and full-blown small vessel vasculitis, i.e. WG (19). Once this question is answered, a more precise examination of a corresponding PR3-specific T cell response should be possible, as well as a monitoring of a potential epitope change. For systemic sclerosis, the use of a random peptide library not only led to the detection of an immunodominant epitope with homologies to a viral protein as well as to an autoantigen, but this recognition also initiated a potential pathologic mechanism, namely apoptosis of endothelial cells (20). Therefore, we hypothesize that such an approach will allow us to confirm known and identify still unknown immunodominant antigenic epitopes binding to ANCA IgG. Thus, the rationale of this proposed study is to obtain information about WG-relevant and less-WG-relevant peptide epitopes binding to ANCA, which could then be used for elucidating (auto)antigen presentation and antigenic T cell responses in WG and even for PR3-PR3-ANCA binding inhibition experiments in search for specific therapeutic agents.

2. Research methods and procedures including, if applicable, proposed statistical analysis.

IgG from PR3-ANCA+ sera of WG patients (n=25) will be purified by protein G column liquid chromatography. IgG preparations will be examined for PR3-ANCA titer (U/ml) by ELISA. Screening of potentially ANCA binding epitopes will be done employing a commercially available random peptide library, based upon a fusion protein consisting of a flagellar protein and thioredoxin, which is expressed at the surface of E. coli. In brief, the pooled IgG preparations will be coated onto a tissue culture plate at 20 µg/ml and then an incubation with the peptide-expressing E. coli will follow. After washing the bound bacteria are eluted, multiplied and incubated again with the IgG preparation to select for frequent epitopes. Following each so-called panning step the bound bacteria are seeded on agar plates as well to identify single clones. These clones are then grown in liquid culture and the plasmid DNA is isolated, purified and subjected to sequencing. Follwowing sequencing, data will be analysed using appropriate alignment and structure tools from NCBI (Blast etc., 21) to identify potential peptides, i.e. if mimics can be found. Then, a number of peptides will be synthesized and tested for binding to the patient’s IgG in ELISA, in comparison to PR3. Statistics will be analyzed using the SPSS software (Munich, Germany).

Gene Expression Profile of Temporal Arteritis in Giant Cell Arteritis

Investigator:

Rula, A. Hajj-Ali, MD and Gary S. Hoffman, MD

Location:

Center for Vasculitis Care and Research, Cleveland Clinic Foundation Cleveland, Ohio

Timeline:

March 1, 2006 - February 28, 2007

Abstract:

Giant cell arteritis (GCA) is the most common type of vasculitis and affects people with mean age of about 70.Complications include ischemic–occlusive disease leading to blindness, strokes, neurologic dysfunction, and lesions in major arteries. The pathogenesis of vasculitis is incompletely understood. It is uncertain whether selective injury to specific vascular sites is due to primary abnormalities of immune function or abnormalities that first appear within the once normal vessel that “invites” an immunoinflammatory response. Observations in man and experimental models indicate that the vessel wall is not a passive participant in the process of injury, and injured sites are not randomly selected. We hypothesize that the tissue substrate (vessel wall) in GCA plays an important role in disease vulnerability and pathogenesis and that gene expression profile of uninvolved segments of temporal arteries from patients with GCA, will reveal the initial changes that predispose to vasculitis, before overt inflammation occur. Our goals are to characterize the molecular signature that defines vessel targeting and susceptibility for different vascular disease and to identify pathogenic mechanisms in giant cell arteritis. We propose to use microarray techniques to specifically aim to identify differential gene expression patterns of uninvolved segments of temporal arteries from patients with GCA compared to age-, gender and ethnicity- matched control specimens, to validate the data obtained from microarray analysis by using Real Time Quantitative PCR and to Identify the microenvironment where the genes of interest are expressed using RNA in situ hybridization techniques. Differences in gene expression that we identify in the temporal arteries of GCA-affected vs. control temporal arteries should lead to a better understanding of pathogenesis of GCA. These findings will aid to understanding qualities of vascular substrate that influence susceptibilities to other type of systemic vasculitis including Wegener Granulomatosis.

Specific AimsTo determine whether acquired abnormalities within the vessel wall precede and are required for inflammatory injury (i.e. vasculitis) in giant cell arteritis (GCA) of the elderly.

Relevance to Wegener's Granulomatosis

It is uncertain whether selective injury to specific vascular sites is due to primary abnormalities of immune function or abnormalities that first appear within the once normal vessel that “invites” an immunoinflammatory response. The fact that giant cell arteritis affects patients >50 years of age (mean 74) suggests that acquired abnormalities of either the targeted vessels and/or of immune function play a role in pathogenesis. Observations in man and experimental models indicate that the vessel wall is not a passive participant in the process of injury, and injured sites are not randomly elected. For example, GCA has a predilection for the aortic arch, its primary branches, and the cranial vessels that originate from the external carotid artery (e.g. temporal, occipital, maxillary and facial arteries). In contrast, the abdominal aorta and its distal branches are rarely involved. These observations about site predilection argue most strongly for initial change in tissue substrate playing an important role in pathogenesis.

However such patterns and onset in the elderly would not eliminate a role for genetic factors. Indications of genetic factors playing a role in GCA include the observations that women are affected 2-3 times more often than men and individuals of Northern European decent are more commonly affected than others, such as African Americans and Asians, among whom GCA is rare. Patients with GCA also have an ncreased requency of HLA-DR4 alleles. Prior studies, that have evaluated the pathogenesis of GCA, have primarily focused mostly on description of the inflammatory events within the vessel wall. These studies have added a great insight to understanding cell populations and cytokine profiles that play a role in injury. However, the question of why selective vessel targeting occurs or what acquired vessel changes lead to triggering disease in specific vessels has not been addressed. Knowledge of gene expression patterns within the vessel wall in GCA will enhance our understanding of the initial events in the pathogenesis of GCA and will have implications in the pathogenesis of not only of giant cell vasculitis but also in different systemic vasculitides including Wegener Granulomatosis.

Future Plans

The pathophysiology of GCA is poorly understood and efforts to understand this disease are highly appreciated. We are proposing to study the pathophysiology of giant cell arteries in a novel way that address the organ targeting and challenge the long standing paradigm that the initial insult in GCA is immune mediated. The advent of new methodologies like the microarray provide unprecedented opportunities to study the wall of vessels affected by GCA in a comprehensive manner. In addition, complementing these studies with In situ hybridization will reveal the topography of the identified genes. Such methods have not been applied previously in studying this disease. We expect to find differential gene expression profile at initial stage of the disease and complement the data in the future by the differential protein profile which will add a great insight into the understanding of the functional role of the expressed genes. We have taken a careful step in analyzing the data, to include ways to check for PI Name: Hajj-Ali , Rula, A. reproducibility, validity and reliability of the measurements, in order to interpret the results in a meaningful way. In this design we aimed to further validate the data and most importantly to localize the microenvironment of the genes of interest. This proposal represents a pilot study of our future research plans. Once the differentially expressed genes have been identified and organized into functional groups or “clusters”, the implications of these gene expression changes need to be addressed. The second tier of studies will allow us to determine whether the gene expression changes identified are initial triggering events, secondary changes or late compensatory changes. We will employ functional proteomic analysis to investigate proteins that interacts with specific genes of interest.

The localization of the cellular microenvironment or structure where the differentially expressed genes will emerge will serve as the basis for specific evaluation of this structure or cellular element in the pathogenesis of GCA.

Rituxan in Churg Strauss Syndrome - Pilot Study at Mayo

The Mayo Clinic Nephrology Collaborative Group (MNCG) announces a new pilot study on the use of Rituximab in the treatment of Churg Strauss Syndrome with renal involvement.

The plan is to enroll five patients in the open label study using four weekly doses of Rituximab. Patients will be followed for one year after beginning of the remission induction regimen. Disease relapses, as defined in the protocol will be treated according to the standard of practice for patients with this disease.

To learn more:
Study Coordinators:
Lori Riess, 507.266.1047
Shirley Jennison, 507.255.0231
Email: MNCG@mayo.edu

Rituximab (Anti-CD20) for the Treatment of Hepatitis C Associated Cryoglobulinemic Vasculitis

Summary: This study will examine the safety and effectiveness of the drug Rituximab in treating hepatitis C-associated cryoglobulinemic vasculitis.

About 5 percent of patients with hepatitis C develop cryoglobulinemic vasculitis. This syndrome, characterized by inflammation of blood vessels (vasculitis), may involve the skin, joints, kidneys, nerves and other sites, and cause skin rashes, joint pain, weakness, fatigue, and numbness.

About 10 to 30 percent of patients develop kidney disease, which, in some cases, can lead to kidney failure. Although the cause of cryoglobulinemic vasculitis is not known, a critical component is the presence of cryoglobulins-abnormal proteins that white blood cells called B lymphocytes produce in response to the chronic hepatitis C infection. Rituximab decreases the number of B cells.

The Food and Drug Administration approved Rituximab in 1997 for the treatment of B-cell non-Hodgkin's lymphoma.

Patients between 18 and 75 years of age with hepatitis C and signs and symptoms of cryoglobulinemic vasculitis may be eligible for this study. They must have failed, or been unable to tolerate, treatment with IFN-a and ribavirin. Candidates will be screened with a history and physical examination, electrocardiogram (ECG), blood and urine tests, 24-hour urine collection and chest X-ray, if clinically indicated.

Participants will be randomly assigned to receive Rituximab upon entering the study or 6 months after entering the study. Those whose treatment is delayed 6 months will be followed once a month at NIH for disease evaluation and blood tests during that time.

Patients will be given Rituximab intravenously (through a vein) once a week for 4 weeks. For the first dose, patients will be admitted to the hospital for at least 24 hours after the infusion for monitoring. Subsequent infusions will be given on an inpatient or outpatient basis, depending on how the infusion is tolerated. The day before each infusion they will have a history and physical examination, blood work, and other tests, such as X-rays, as clinically indicated. After the four infusions, patients will be followed for drug side effects and response to treatment. They will have blood tests every week for 4 weeks and will then return to NIH for 1 day every month for 12 months for a physical examination, blood tests, and X-rays, if medically indicated. Visits may be more frequent, if necessary, and patients may be asked to stay longer than a day if test findings require further investigation.

Patients whose cryoglobulinemic vasculitis improves and remains inactive for 12 months after completing Rituximab therapy will have completed the study. Those who benefit from treatment but have a later worsening of disease may receive a second course of four infusions, with follow-up for 12 months after the last infusion. Patients who do not respond to treatment and whose disease worsens will be advised of other treatment options, but will be asked to return to NIH monthly for 12 months for follow-up.

Sponsoring Institute: National Institute of Allergy and Infectious Diseases (NIAID)

Recruitment Detail Type: Participants currently recruited/enrolled

Gender: Male & Female Referral Letter Required: No Population Exclusion(s): Children Eligibility Criteria:

INCLUSION CRITERIA

Diagnosis of HCV-CV: must have all of the following HCV infection documented by serology and/or plasma HCV RNA.

One or more organ system with objective evidence of active vasculitis such as: Palpable purpura; Glomerulonephritis (defined by the presence of glomerular hematuria and/or new or worsening proteinuria); Acute peripheral neuropathy. Detectable cryoglobulins and/or RF.

Failure of treatment with IFN-alpha and ribavirin to control manifestations of HCV-CV OR intolerance to IFN-alpha/ribavirin regimen.

Patients must have a personal physician responsible for the care of their HCV.

Ages of 18 and 75 years

Willingness to use effective contraception during and for 12 months following Rituximab treatment. Effective contraception methods include abstinence, surgical sterilization of either partner, barrier methods such as diaphragm, condom, cap or sponge, or hormonal contraception.

EXCLUSION CRITERIA

Recent (within 4 weeks) initiation of or increase in immunosuppressive therapy.

Active systemic infection (other than hepatitis C).

Pregnancy or breast feeding.

Prior treatment with Rituximab.

Known allergy to murine proteins.

Significant renal insufficiency (creatinine clearance less than 30 ml/min).

Presence of life-threatening HCV-CV; defined as rapidly progressive glomerulonephritis (defined as a doubling of the serum creatinine over a 3 month period), CNS vasculitis, cardiac disease due to active vasculitis, or GI vasculitis (defined by ischemic bowel, perforation, or infarction).

Significant hepatic insufficiency as manifested by Child-Pugh classification of B or C.

History of variceal bleeding, encephalopathy.

History of liver transplantation.

Co-infection with either HBV or HIV.

Any underlying medical condition that in the judgment of the investigator would put the patient at increased risk for serious infusion-related adverse events. Special Instructions: Currently Not Provided Keywords: Hepatitis Vasculitis Rituximab Cryoglobulinemia Recruitment Keyword(s): Hepatitis C Vasculitis HcV-cV Cryoglobulinemic Vasculitis Cryoglobulinemia Condition(s): Hepatitis C

Vasculitis Investigational Drug(s): Rituximab Investigational Device(s): None Intervention(s): Drug: Rituximab Supporting Site: National Institute of Allergy and Infectious Diseases

Contact:

National Institutes of Health Building 10 Room 8C416 10 Center Drive

Bethesda, Maryland 20892

Phone: 1-(800) 411-1222

VCRC Longitudinal Studies and Patient Registry

Vasculitis Clinical Research Consortium (VCRC)

The VCRC was established in 2004 to address the challenges inherent in diagnosing and treating rare vasculitic diseases. Dr. Peter Merkel, of Boston University School of Medicine, is the Principal and Overall Lead Investigator for the VCRC, along with Co-Investigators from Johns Hopkins, Cleveland Clinic and the Mayo Clinic. The VCRC is conducting Longitudinal Studies (now enrolling patients) in different forms of vasculitis to identify biomarkers for disease risk, disease severity and activity, and clinical outcome. The VCRC is also planning to conduct experimental trials of new drugs for vasculitis. The VCRC Patient Contact Registry is a method for patients with vasculitis to register themselves with the VCRC so that they will be notified of new clinical research studies conducted by the VCRC. The contact registry is free of charge and anonymous, and patients can register by using a paper form, by calling a toll-free number (866-313-9879), or online by clicking “Join the VCRC Contact Registry” at http://rarediseasesnetwork.org/vcrc. By taking action, patients can help researchers develop a better understanding about vascultis. Patients who participate in research make it possible for researchers to create new studies and to find new treatments.

The Genetics Study of Systemic Vasculitis

Dear Interested Participant:

You are invited to participate in a new research study on the genetics of vasculitis. Some examples of the diseases classified as vasculitis include Wegener's granulomatosis, Takayasu's arteritis, microscopic polyangiitis, the Churg-Strauss syndrome, and polyarteritis nodosa.

Vasculitis is a group of diseases about which much remains unknown. This is particularly true about risk factors for the disease: who gets the disease and why?

We believe that genes contribute to a person's risk of getting vasculitis. This study is designed to identify some of the genes that may be important in leading to the condition of your specific form of vasculitis.

Your participation in the study would involve the following:

(1) You must have a diagnosis of vasculitis and be willing to answer a series of questions regarding your health and medications. We may also ask your doctor some questions regarding your health and test results. Before starting this study, we will have you sign a form that tells you about this study and how we plan to keep your information private during the study.

(2) You must have two living biological parents or one parent and two siblings who are also willing to participate in the study.

(3) You would provide a blood sample that could be drawn at your doctor's office using materials we send in the mail. The sample will be mailed back to us, using a pre-packaged mailing kit.

(4) After completing the consent process, your parents and/or siblings would provide a swab from inside their cheek (this can be done at home, without requiring a visit to their doctor's office or drawing blood) using materials we would send in the mail. The swab would then be mailed back to us, using a pre-packaged mailing kit.

If you are interested in participating in this study, you may contact us in one of two ways:

Telephone (on a toll-free number): 1-800-226-6023

Email: jhvc@jhmi.edu

We appreciate your interest in the study.

Sincerely,

Dr. Stuart Levine

Assistant Professor of Medicine

Co-Director, The Johns Hopkins Vasculitis Center

Johns Hopkins Bayview Medical Center

Division of Rheumatology

Rituximab for ANCA-Associated Vasculitis (RAVE) Study

Enrollment is Open: Participants Needed

Sponsored by The National Institute of Allergy and Infectious Disease and The Immune Tolerance Network

A randomized, multicenter trial for patients with severe ANCA-associated vasculitis.

Participants will be randomized to either a treatment of cyclophosphamide (CYC) and prednisone or rituximab and prednisone.

The primary objective of this trial is to determine the efficacy of rituximab and glucocorticoids in the induction of remission of ANCA-associated vasculitis.

For further detailed information: http://vasculitis.med.jhu.edu

Study Coordinators:

Lourdes Sejismundo (410) 550-6818 or

Yavette Goldsborough (410) 550-4580

Email: jhvc@jhmi.edu

Lead Investigators:

Ulrich Specks, M.D.,

Division of Pulmonary and Critical Care, Mayo Clinic

Co-directors: Philip Seo, M.D./Stuart Levine, M.D.,

The Johns Hopkins Vasculitis Center

Participating Medical Centers

Boston University

Cleveland Clinic

Duke University

Hospital for Special Surgery, New York City

Mayo Clinic

The Johns Hopkins Vasculitis Center

University of Alabama at Birmingham

Identification of Novel Wegener’s Granulomatosis Susceptibility Genes

Investigator:

Katherine A. Siminovitch, MD Location: Mount Sinai Hospital, Toronto, Ontario, Canada

Timeline: March 1, 2004 – ongoing

Dr. Katherine Siminovitch’s study “Identification of Novel Wegener’s granulomatosis Susceptibility Genes” is an ongoing project to collect blood samples from Wegener's patients and their family members.

The goal of the study is to collect blood samples and prepare DNA from 500 patients and their immediate family members, and to conduct a comprehensive clinical chart review in order to obtain powerful data for results stratification and analysis.

The study needs at least 250 more WG patients to donate samples to have sufficient data to develop definitive results. It is very easy to participate in the study. Patients enroll in the study and have a blood sample drawn at their local doctor’s office and shipped to Dr. Siminovitch with the materials provided.

For more information please contact:

Alida Pokoradi, Clinical Research Coordinator

Phone: 416-946-4501 x3297

Fax: 416-946-4531

Email: clinicalgenet.research@gmail.com

To read the complete abstract for the study:

Identification of Novel Wegener’s granulomatosis Susceptibility Genes