Genetic and Environmental Risk Factors for Psoriatic Arthritis among patients with Psoriasis

Transcription

1 Genetic and Environmental Risk Factors for Psoriatic Arthritis among patients with Psoriasis by Lihi Eder A thesis submitted in conformity with the requirements for the degree of PhD Institute for Medical Science University of Toronto Copyright by Lihi Eder 2011

2 Genetic and Environmental Risk Factors for Psoriatic Arthritis among Patients with Psoriasis Lihi Eder PhD Institute of Medical Science University of Toronto 2011 Abstract Aim: Most of the patients with Psoriatic Arthritis (PsA) develop arthritis following the onset of psoriasis. The aim of the project is to identify genetic and environmental risk factors for PsA among psoriasis patients. Methods: PsA and psoriasis patients from two prospective cohorts were analyzed. The incidence of PsA among a prospective cohort of psoriasis patients was assessed. The distribution of Human Leukocyte Antigen (HLA) alleles and Killer Cell Immunoglobulin like Receptors (KIRs) and their combinations was compared between PsA, psoriasis and healthy controls. In addition, the association between a wide range of environmental exposures and PsA was evaluated by comparing the frequencies of exposed individuals among patients with recent onset PsA and psoriasis. The association between smoking and PsA was further investigated. The prevalence of smoking was in PsA, psoriasis and the general population. The interaction between HLA-C*06 and smoking was also tested. ii

3 Results: The genetic analysis revealed several HLA-B alleles and HLA haplotypes that are associated with PsA compared to psoriasis and can potentially serve as independent markers for PsA. Furthermore, several combinations of KIR genes and their respective HLA ligands were also found to be associated with PsA compared to psoriasis. The incidence of PsA among psoriasis patients was found to be higher than previously reported and its rate was constant over time. HLA-C*06 was associated with increased interval between psoriasis onset and PsA. Several environmental factors including occupational exposures, infections, injuries and smoking were associated with development of PsA. The prevalence of smoking was decreased among PsA patients compared to psoriasis. The interaction between HLA- C*06 and smoking was found to be significant. Conclusions: Genetic and environmental factors are associated with the development of PsA in patients with psoriasis. These factors may serve as specific markers to identify psoriasis patients at increased risk for PsA. iii

4 Acknowledgments I would like to express my gratitude and appreciation to Dr. Dafna Gladman, my thesis supervisor for introducing me to the great world of research. Her leadership, support, hard work, and scholarship have set an example I hope to match some day. She is a true role model to me as a researcher, physician and as a person. I would like to thank the other members of my committee, Dr. Cheryl Rosen, for the insights and patience particularly in reviewing my thesis, and Dr. Shelley Bull for her direction and guidance in analyzing the data and insightful comments. Gratitude is also expressed to all of the people whom without their help this project would not have been possible. I would like to thank Dr. Vinod Chandran who helped me understand better the world of genetic epidemiology. My sincere thanks also go to Sutha Shanmugarajah for her immeasurable help in recruiting patients and managing the data for this project. I would also like to thank Fawnda Pellet and Remy Pollock for their hard work in the laboratory performing the HLA and KIR genotyping. I wish to thank Dr. Richard Cook, Hua Shen and Arane Thavaneswaran for the meaningful analysis and guidance in performing the statistical analysis of this project. Special thanks to Anne MacKinnon for her help behind the scene in getting things done. I would like to thank the Abbott PsA Fellowship and the Canadian Arthritis Network for their financial support. Last but not least, I would like to thank my family, Ben, Adi and Shani Eder, who joined me on this journey. Without their encouragement and understanding it would have been impossible for me to finish this work. iv

5 Table of Contents Abstract... ii Table of Contents... v List of Tables... vii List of Figures... x List of Appendices... xi Chapter 1. Literature Review The Phenotype - Psoriasis The Phenotype Psoriatic Arthritis The Etiology of Psoriasis and PsA Environmental risk factors for Psoriasis and PsA Genetics of Psoriasis and PsA Chapter 2. Hypothesis and Aims Rationale Hypothesis Aims Chapter 3. Study population and data collection Study population Data collection Chapter 4. The incidence of arthritis in a prospective cohort of psoriasis patients Background Methods Results Discussion Chapter 5. HLA risk alleles for PsA among psoriasis patients Background Methods Population stratification Results Discussion Chapter 6. The effect of HLA risk alleles on the rate of development of PsA among psoriasis patients v

6 6.1. Background Methods Results Discussion Chapter 7. The association of KIRs and their HLA ligands with PsA Background Methods Results Discussion Chapter 8. The association between environmental factors and onset of PsA in patients with psoriasis Background Methods Study population Data collection Statistical analysis Results Study population characteristics Proportion of exposure to the different factors Smoking and alcohol consumption Discussion Chapter 9. Smoking is inversely associated with development of psoriatic arthritis among psoriasis patients Background Methods Results Discussion Chapter 10. General Discussion Conclusions Future directions References Appendices Appendix 1 - Questionnaire for the assessment of exposure to environmental factors 196 vi

7 List of Tables Table Characteristics of the study population and the incident cases of PsA Table Population stratification within the Caucasian study sample Table Demographic and clinical characteristics of the study population Table HLA-A allele distribution Psoriatic disease vs. Controls Table HLA-B allele distribution Psoriatic disease vs. Controls Table HLA-C allele distribution Psoriatic disease vs. Controls Table HLA-DRB1 allele distribution Psoriatic Disease vs. Controls Table HLA-DQ allele distribution Psoriatic Disease vs. Controls Table The association of HLA alleles and PsD compared to healthy controls using multivariate logistic regression analysis Table HLA-A allele distribution PsA vs. Psoriasis Table HLA-B allele distribution PsA vs. Psoriasis Table HLA-C allele distribution PsA vs. Psoriasis Table HLA-DRB1 allele distribution PsA vs. Psoriasis Table HLA-DQ allele distribution PsA vs. Psoriasis Table Odds ratios comparing HLA allele frequencies in PsA to psoriasis using logistic regression analysis Table Odds ratios comparing HLA-C*06 allele frequencies in PsA to psoriasis by age at onset of psoriasis Table Odds ratios comparing HLA allele frequencies in Axial PsA vs. psoriasis using logistic regression analysis Table Odds ratios comparing HLA allele frequencies in peripheral PsA vs. psoriasis using logistic regression analysis Table Odds ratios comparing extended HLA haplotype frequencies in PsD to healthy controls using logistic regression analysis Table Linkage Disequilibrium of selected HLA-B and C alleles Table Odds ratios comparing HLA-B/HLA-C haplotype frequencies in PsA to psoriasis using logistic regression analysis Table Detailed family structure of the study population vii

8 Table Family based association test Affected (PsA probands) - Unaffected (psoriasis) sib-pairs and Trios Table Family based association test Affected (PsD)-Unaffected (Healthy controls) sibpairs and Trios Table Characteristics of the study population Table The association between HLA alleles and the risk of PsA in analysis of time from onset of psoriasis by parametric proportional hazard model Table The association between HLA alleles and risk of PsA in analysis of time from birth by parametric proportional hazard model Table KIR ligand specificities Table KIR gene distribution Psoriatic disease vs. Healthy controls Table KIR gene distribution PsA vs. Psoriasis Table The frequency of HLA-C group 1/HLA-C group 2 alleles in PsA, psoriasis and healthy controls Table 7.5 The association between combinations of KIR2D/HLA based on activation model and PsA vs. Psoriasis Table The frequencies of KIR2DS2 and KIR2DL2 in PsA and psoriasis Table The association between combinations of KIR2D/HLA based on activation model and PsD vs. Controls Table The frequency of KIR3DS1 in PsA and psoriasis Table The association between combinations of KIR3D/HLA based on activation model and PsA vs. Psoriasis Table Clinical characteristics of the study population Table 8.2 The association between environmental exposures and PsA Table 8.3 The Association between female hormonal exposures and PsA Table Full regression model adjusted for age, sex, education level, duration and severity of psoriasis Table 9.1 Demographic and clinical characteristics of the study population Table 9.2 Smoking characteristics among psoriasis and PsA patients Table 9.3 Smoking status at the time of the diagnosis by matched pairs viii

9 Table 9.4 The association between smoking status and PsA compared to psoriasis alone by logistic regression analysis ix

10 List of Figures Figure Genes within the MHC region Figure Follow-up summary of the study population Figure PsA incidence over time estimated with a Kaplan-Meir curve from time of first clinic visit to diagnosis of PsA Figure 6.1- The probability of developing PsA among psoriasis patients from the onset of psoriasis assuming an exponential model Figure Illustration of the structure of the different type of KIRs Figure The 10 most frequent KIR genotypes in PsA, psoriasis and healthy controls Figure Smoking status among PsA and psoriasis patients by HLA-C*06 status x

11 List of Appendices Appendix 1 - Questionnaire for the assessment of exposure to environmental factors xi

12 List of abbreviations AS - Ankylosing Spondylitis BSA - Body Surface Area CASPAR Classification of Psoriatic Arthritis CDSN - Corneodesmosin CI Confidence Interval DMARDs - Disease Modifying Anti-Rheumatic Drugs FDR - False Discovery Rate GWAS - Genome Wide Association Study HLA - Human Leukocyte Antigen IBD Inflammatory Bowel Disease KIR - Killer Immunoglobulin-like Receptor LR Likelihood Ratio LD - Linkage Disequilibrium msasss - modified Stoke Ankylosing Spondylitis Spine Score mnapsi - modified Nail Psoriasis Severity Index NK - Natural Killer NSAIDs - Non-Steroidal Anti-Inflammatory Drugs OR - Odds Ratio PsA Psoriatic Arthritis PASI Psoriasis Area and Severity Index PSORS - PSORiasis Susceptibility locus PsD - Psoriatic Disease ReA - Reactive arthritis SNP - Single Nucleotide Polymorphisms SpA Spondyloarthropathies SPR Standardized Prevalence Ratio TNF - Tumor Necrosis Factor ToPAS - Toronto Psoriatic Arthritis Screen xii

13 Chapter 1. Literature Review 1

14 The Phenotype - Psoriasis Psoriasis is a common skin disease affecting 1-3% of the population [1-3]. The antiquity of the disease dates back to biblical times. It has long been recognized that most cases of biblical leprosy were actually psoriasis [4]. Lepers were considered unclean and were subject to regulation and segregation from the normal population. Only in the 18 th century a detailed description that distinguished these skin lesions by the British dermatologist Robert Willan, ended hundreds of years of confusion and laid the foundation for establishing psoriasis as an independent disease [5] Epidemiology of psoriasis Psoriasis affects men and women equally, and is seen in all races. Although psoriasis can begin at any age, there seem to be two peaks in onset: one between ages 20 and 30 and another between ages 50 and 60 [6]. Psoriasis has been described as two types depending on the age of onset. Patients with early-onset or Type I psoriasis (before the age of 40) tend to have more severe disease with a familial history. Patients with late-onset or Type II psoriasis (after the age of 40) tend to have a milder disease [7] Clinical manifestations of psoriasis Psoriasis is usually manifested as raised, erythematous plaques with adherent silvery scales. It is usually easily recognized, but atypical or non-classic forms are more difficult to identify. There are several clinical types of psoriasis; the most common one is chronic plaque psoriasis or psoriasis vulgaris that affects 85-90% of all patients with the disease. This type usually presents in young adults with symmetrically distributed plaques involving the scalp, extensor elbows, knees, and back. Other types include flexural psoriasis, guttate psoriasis, pustular psoriasis and erythroderma [8]. Approximately 40% of patients with psoriasis have nail lesions. One of the typical nail abnormalities in psoriasis is pitting, consisting of a few to multiple tiny pits scattered over the nail plate. The pits reflect abnormal nail plate growth resulting from psoriatic involvement of the nail matrix. These changes produce friable areas of nail plate that erode away with normal friction. Another

15 3 typical psoriatic nail lesion is onycholysis, that occurs as a result of a separation of the nail plate from its underlying attachment to the nail bed [6] Diagnosis of psoriasis Although the differential diagnosis of psoriasis is broad, a skin biopsy is rarely needed. The diagnosis is usually made by history and physical examination. There are no laboratory tests that confirm or exclude the diagnosis. A detailed physical examination should focus on typical sites of involvement such as knees and elbows with a special attention to subtle findings in the scalp, umbilicus, intergluteal cleft, and nails Clinical course and co-morbidities of psoriasis Psoriasis tends to be a chronic disease. However, its course is unpredictable. There may be marked variability in severity over time, and remissions at some stage are seen in up to 40% of cases [9]. Although generally not life threatening, psoriasis may be associated with important morbidity and disability. It can range from a very mild disease with few small hidden plaques that do not interfere with daily life to severe wide-spread skin lesions that may lead to disability and poor quality of life. Patients with psoriasis, like those with other major medical disorders, have reduced levels of employment as well as decreased quality of life [10, 11]. Psoriasis is also associated with other co-morbidities including obesity and other related metabolic abnormalities such as diabetes mellitus and dyslipidemia [12, 13]. These in turn lead to increased cardiovascular morbidity and mortality [14]. In addition, approximately 30% of the patients with psoriasis develop an inflammatory arthritis termed Psoriatic Arthritis (PsA) [15].

16 Measurement tools in Psoriasis The Psoriasis Area and Severity Index (PASI) is the most widely used tool for the measurement of severity of psoriasis. PASI combines the assessment of the severity of lesions and the area affected into a single score in the range of 0 (no disease) to 72 (maximal disease) [16]. Another measurement tool for psoriasis severity is the Body Surface Area (BSA) which is an estimate of the percentage of the body surface affected by psoriasis. Patients with severe psoriasis are those with BSA>10% or PASI>10 [17] Treatment of psoriasis Treatment in psoriasis can generally be categorized into topical and systemic therapies. The treatment choice is dictated by the severity, type, and location of psoriasis. Patients with mild psoriasis can usually be managed with topical agents including: corticosteroids, tar, retinoids and vitamin D derivates. Moderate to severe psoriasis requires phototherapy or systemic therapies such as methotrexate, retinoids, cyclosporine or the biologic immune modifying agents including alefacept, the anti- Tumor Necrosis Factor (TNF) agents or anti-il12/23 monoclonal antibody (ustekinumab) [18].

17 The Phenotype Psoriatic Arthritis Psoriatic arthritis has been defined as an inflammatory arthritis associated with psoriasis, usually seronegative for rheumatoid factor [19]. The association between psoriasis and arthritis was first described in 1818 by the French physician, Baron Jean Louis Alibert [20]. However, only in 1964 did the American Rheumatology Association recognize PsA as a unique disease entity that is separate from rheumatoid arthritis [21]. Furthermore, only in 2006, were classification criteria for PsA developed and allowed a better definition of cases for research purposes [22]. Psoriatic arthritis is classified among the seronegative spondyloarthropathies (SpA). This term refers to a family of diseases that share certain clinical features. The most distinguishing features are inflammation of the axial joints, asymmetric oligoarthritis, and enthesitis (inflammation at sites of ligamentous or tendon attachment to bone). Additional features are genital and skin lesions, eye and bowel inflammation, an association with preceding or ongoing infectious disorders, and a strong association with the Human Leukocyte Antigen (HLA) -B*27. The SpA group consists of the following disorders: Ankylosing Spondylitis (AS), Reactive arthritis (ReA), PsA, Undifferentiated spondyloarthritis, SpA associated with Inflammatory Bowel Disease (IBD) and Juvenile onset spondyloarthritis [23] Epidemiology of PsA The most recent estimate of the prevalence of PsA in North America is 0.25% (95% CI: 0.18%, 0.31%) [24]. The reported incidence of PsA in the general population ranges from per 100,000 [25-28]. PsA has been reported in 7-42% of patients with psoriasis, with a recent estimate being approximately 30% [15, 29, 30]. The marked variability in reported prevalence and incidence rates is probably related to different definitions of the disease as well as variable sources of populations. Approximately 67% of the patients develop psoriasis before arthritis and in 16% arthritis and psoriasis present within 12 month of each other [31]. There is limited information about the incidence of PsA among patients with

18 6 psoriasis. A retrospective study from Germany reported that the cumulative incidence of PsA among psoriasis patients reached 20.5% after 30 years from the diagnosis of psoriasis [32]. Another retrospective study from Rochester, Minnesota has found a lower cumulative incidence of 3.1% cases of PsA among psoriasis patients after 10 years from the onset of the skin disease [33]. No study to date has prospectively assessed the incidence of PsA among psoriasis patients. The prevalence of the disease is equal among males and females. PsA usually occurs in the third or fourth decade of life [15]. There is very little information about racial and ethnic associations as most epidemiological studies has been performed in Caucasians Clinical manifestations Five patterns of PsA have been described: the symmetric polyarticular pattern being the most common [34], distal arthritis that involves the distal interphalangeal joints, asymmetric oligoarthritis in which less than 5 joints are affected, arthritis mutilans that is characterized by deforming and destructive arthritis, and spondyloarthritis that includes sacroiliitis and spondylitis [35]. Some of the patients present with more than one pattern or change their pattern during the course of their disease. Another common feature of PsA is enthesitis, an inflammation at the site of tendon insertion into the bone, that often affects the Achilles tendon, plantar fascia and pelvis bones [36]. Dactylitis is characterized by diffuse swelling of the entire finger or toe; it affects about half of the patients and is associated with radiographic joint damage [37]. Similarly to the other spondyloarthropathies, PsA is also associated with inflammation in other extra-articular sites including the eye (uveitis) and the gastrointestinal tract (inflammatory bowel disease) Relationship between skin and joint disease PsA may be considered as a disease within a disease as most of the patients with PsA also have psoriasis, although according to the new classification criteria, patients with PsA do not have to have established psoriasis [22]. Most of the patients develop PsA after or at the

19 7 same time as the skin disease, however, 15% of patients with PsA present with arthritis before psoriasis [35]. Since most patients develop PsA after the onset of psoriasis, the skin disease serves as a marker for the development of PsA. The paradigm that patients with severe psoriasis are the ones who develop arthritis is controversial. Several studies reported a higher prevalence of PsA among patients with severe psoriasis [24, 38]. However, the fact that PsA can present before psoriasis as well as recent observations of no relationship between the severity of the skin and joint manifestations [39, 40], suggest there is no direct link between psoriasis severity and arthritis. Nail lesions are more common among PsA patients than in psoriasis and the presence of nail lesions has been suggested as marker of increased risk for PsA among psoriasis patients [41, 42] Diagnosis of PsA The diagnosis of PsA is based on a typical combination of clinical, laboratory and radiographic findings. The ClASsification of Psoriatic ARthritis (CASPAR) criteria are a set classification criteria for PsA that were published in They may be used for diagnosis and allow a uniform definition of cases for research purposes. The CASPAR criteria showed high sensitivity and specificity for PsA (91.4% and 98.7%, respectively) [22]. The CASPAR criteria consist of the following terms: Required: The presence of an inflammatory arthritis, enthesitis, or spondylitis. Plus 3 points from the following: 1. Skin psoriasis (present) (2 points), previously present by history (1 point), or a family history of psoriasis (1 point) 2. Psoriatic nail lesions (1 point) 3. Dactylitis (1 point) 4. Negative rheumatoid factor (1 point)

20 8 5. Juxta-articular bone formation on radiographs (1 point) Clinical course of PsA In most patients PsA runs a course of a chronic, progressive disease, although some patients can achieve a complete remission. In our cohort, 17.6% of the patients achieved a remission, however, periods of remission lasted on average 2.6 years and most patients experienced a relapse [43]. PsA is more severe than previously thought [44, 45]. It can lead to severe joint damage and disability that are comparable to those that occur in rheumatoid arthritis [46].Patients with PsA demonstrate clinical and radiographic progression in the course of follow-up [47, 48] and have an increased mortality risk compared to the general population, although this risk has decreased over the past two decades [49, 50]. In addition to the disability that is related to the joint disease, PsA patients can suffer from the same co-morbidities as psoriasis patients [51]. Thus, PsA poses a major health burden, in addition to that caused by psoriasis alone Treatment of PsA The treatment in PsA is aimed at controlling both skin and joint inflammation in order to reduce the symptoms and to prevent joint damage. Treatment usually begins with Non- Steroidal Anti-Inflammatory Drugs (NSAIDs) that can control mild arthritis, enthesitis and spondylitis. Second line therapies are indicated when arthritis does not respond to NSAIDs. Many of the Disease Modifying Anti-Rheumatic Drugs (DMARDs), such as methotrexate, leflunomide, sulfasalazine, cyclosporine and azathioprine, were borrowed from rheumatoid arthritis for the treatment of peripheral arthritis in PsA. Although there are limited clinical trials that evaluated their efficacy in PsA, they are often effective in controlling articular symptoms. The new targeted biologic therapies particularly the anti- TNF agents, are the most effective treatments currently available to control all aspects of the disease and to prevent the progression of joint damage [52].

21 The Etiology of Psoriasis and PsA Immunologic mechanisms in Psoriasis and PsA The role of immunologic mechanisms is suggested by the inflammatory response in the psoriatic skin lesions and by the synovial lesions, which are at times indistinguishable from rheumatoid arthritis [53]. T cells and pro-inflammatory cytokines have important roles in the pathogenesis of both diseases. CD8+ T cells clones from the synovial fluid and skin lesions of patients expand in a presumptive autoantigen-driven manner [54-56]. Patients with PsA have a pattern of pro-inflammatory cytokines in the synovium, particularly from the Th1 milieu including: TNF-alpha, IL-1, IL-6, IL-8 and more [57-59]. The effectiveness of T cell-targeting and cytokine modulating treatments including the anti TNF-α agents, alefacept and efalizumab in psoriasis and PsA support the important role of T cells in the pathogenesis of these conditions. The innate immune system provides an early response mechanism against external threats, mainly infectious agents and includes dendritic cells, keratinocytes, macrophages and Natural Killer (NK) cells. Dysregulation of the innate immune system is thought to play a role in the pathogenesis of psoriasis and PsA [60]. Activation of dendritic cells through antimicrobial peptides leads to an increased production of interferon alpha which is a major inducer of psoriasis. This mechanism provides a potential explanation for the link between environmental triggers that may break the immunologic tolerance and lead to psoriasis and PsA [61]. Natural Killer cells and NK-T cells, which are part of the innate immune system, have been implicated in the pathogenesis of psoriasis [62] and PsA [63-65]. Both NK and NK-T cells have been described in increased numbers in psoriatic plaques [66] and in synovial tissues from PsA patients [63]. NK cell activity is partially controlled through interaction between Killer Immunoglobulin-like Receptors (KIRs) on NK cells and their respective HLA class I ligands [67]. It has been proposed that direct activation of NK cells bearing KIRs for MHC class I may play a role in the pathogenesis of PsA [68, 69]. The concept of synovio-entheseal complex is a proposed theory that can provide a link between mechanical stress and the development of PsA. The synovium provides nourishment and lubrication to the entheseal fibrocartilage. During mechanical stress and

22 10 injury to the enthesis, the associated inflammatory reaction would be manifested in the juxtaposed synovium. The damaged tissue triggers the previously discussed immune mechanisms [70]. Microanatomical studies provide support for the theory by demonstrating an anatomical connection of the nail and the enthesis, that may explain the observation that psoriatic nail lesions are associated with higher risk of developing PsA [71, 72].

23 Environmental risk factors for Psoriasis and PsA Psoriasis and PsA are considered complex diseases in which the interaction between genetic and environmental risk factors is thought to play a major role [73]. Genetic factors cannot solely account for all cases. One of the suggested pathogenic models for PsA is that psoriasis patients who carry susceptibility genes for arthritis develop PsA after being exposed to triggering environmental factors [74]. These environmental factors are largely unknown. Several environmental factors have been associated with psoriasis including: infections, particularly streptococcal pharyngitis, smoking, trauma, stressful life events, obesity and certain medications [75]. Only a limited number of studies have investigated environmental risk factors for PsA. Pattison et al. compared the prevalence of environmental exposures among 98 British PsA and 163 psoriasis patients over a window of exposure that ranged from 5 to 10 years prior to the onset of arthritis. Information about environmental exposures was collected through questionnaires. In their study, physical trauma, rubella vaccination, oral ulcers and moving house were found to be associated with PsA [76]. Another study by Thumboo et al. used an administrative database from Rochester, Minnesota and retrospectively evaluated exposure to several environmental risk factors among 60 PsA and 120 psoriasis patients [77]. They did not define a specific window of exposure. In that study, pregnancy was found to be protective of PsA, while steroid use was associated with higher risk of the disease. Several case reports and case series have highlighted the role of physical trauma as a potential triggering environmental exposure for PsA. In these reports, patients with psoriasis developed arthritis following local trauma, such as injury, surgery or fracture [78-80]. The pathogenic process that may lead from local trauma to joint inflammation in PsA is unclear. However, the hypothesis that these two events are linked is supported by a similar process in the skin that was termed the Koebner phenomenon, in which local skin trauma induces certain skin lesions including psoriasis. This process has been reported in 24-52% of psoriasis patients [81-83]. Many different injuries can induce Koebner response including: insect bites, burns, laceration, venipuncture among others. The time period from

24 12 injury to psoriasis is diverse; in general the interval ranges between 10 and 20 days [84]. Koebnerization can trigger a cascade of events that switch the feature of the disease, from localized to generalized psoriasis. Psoriasis patients who carry the HLA-C*06 allele have a higher incidence of Koebnerization. Local trauma to the skin induces release of proliferation factors and proliferation of inflammatory cells that initiate an inflammatory process. Similar events may take place in a joint following local trauma and was termed Deep Koebner phenomenon [85]. Reactive arthritis is an episode of an acute articular and periarticular sterile inflammation occurring in a genetically predisposed individual secondary to a primary infection elsewhere in the body [86]. Urogenital and gastrointestinal infections are well established triggering events for ReA [87]. Reactive arthritis occurs in 1% to 4% of patients with preceding bacterial gastrointestinal infections with enterobacteria or urogenital infections with Chlamydia trachomatis, however, this rate increases to 20% to 30% in patients infected with one of these bacteria who are positive for HLA-B*27 [86]. Reactive arthritis belongs to the SpA group and has some common features with PsA including: similar pattern of joint involvement, psoriasiform skin lesions, enthesitis, eye involvement and an association with the HLA-B*27 allele. Infections also play a role in the pathogenesis of psoriasis. Streptococcal infection is a common trigger for guttate psoriasis particularly in children and young adults [88] and the reported incidence of streptococcal infections preceding this type of psoriasis ranges between 56% and 97% [89]. Chronic plaque psoriasis is also exacerbated after such infections and psoriasis patients were found to have a ten-fold higher frequency of streptococcal throat infections than age-matched household controls [90]. Psoriatic lesional T cells are oligoclonal, and T cells recognizing determinants common to streptococcal M- protein and keratin have been detected in patients blood [91]. It has been suggested that the association between streptococcal infection and psoriasis may reflect an autoimmune process that is triggered by the presence of a superantigen, possibly the streptococcal M- protein. However, the role of autoimmunity in the pathogenesis of psoriasis remains controversial. The association between streptococcal infection and PsA was suggested by elevated levels of streptococcal RNA in PsA peripheral blood [92, 93]. However, although

25 13 the immunoreactivity to streptococcal antigens is accepted, it is unclear if the infection triggers PsA or if the breakage of skin barrier because of psoriasis leads to streptococcal exposure and finally to a form of reactive arthritis. Infection with Human Immunodeficiency Virus (HIV) is also associated with the development of psoriasis and PsA particularly in HIV endemic populations [94, 95]. It has been reported that in Sub-Saharan Africa the great majority of PsA patients were HIV positive [96]. This association may hint at a viral trigger of PsA or that HIV increases the risk for other infections that may trigger PsA. Obesity is associated with psoriasis. Multiple studies have demonstrated that patients with psoriasis are more frequently overweight or obese compared with patients without psoriasis [97-99].Obesity occurs prior to the onset of psoriasis and is now thought to be a risk factor for the development of the disease. A large prospective cohort from the US demonstrated a dose-response relationship for obesity on the risk of developing incident psoriasis [100]. Obesity (BMI 30) compared with normal body weight was associated with two-fold increased risk for psoriasis onset. Obesity is also associated with more severe psoriasis [101] and poor response to therapy [102]. A recent case-control study from Utah demonstrated an association between obesity and PsA. The results showed that obesity at age 18 was associated with an increased risk and an earlier onset of PsA among psoriasis patients [103], however the study was based on a recall weight and height at age 18 and was not a prospective study. The role of smoking as a risk factor for psoriasis is well established. The first studies drew attention to the linkage between smoking and palmo-plantar pustular psoriasis [104]. Since then several case-control studies have established the association between smoking and psoriasis vulgaris [ ], the most common type of psoriasis. The most conclusive epidemiological evidence for the causative role of smoking in the pathogenesis of psoriasis was provided by the Nurses Health Study II that showed, for the first time, a strong association between smoking and incident psoriasis in a large population-based prospective cohort study [109]. In this study the relative risk of psoriasis was 1.8 for current smokers and 1.4 for past smokers. The dose-effect relation between smoking intensity and psoriasis

26 14 risk also supports its etiological role. Smoking has also been associated with more severe psoriasis and poor response to treatment [110, 111]. The independent association between smoking and psoriasis remained significant even after adjustment for another potential confounder, excessive alcohol consumption, that is also increased among psoriasis patients [112, 113]. Rakkhit et al. found a temporal association between psoriasis, PsA and smoking [114]. They reported that the duration of time from the onset of psoriasis to development of PsA decreases with a history of smoking prior to psoriasis onset and increases with smoking after psoriasis onset. Lastly, stressful life events are thought to have an effect on the course of psoriasis. In a case-control study, psoriasis patients were more likely to experience a stressful event that preceded the onset and the exacerbation of their disease compared to patients with other skin disorders [115]. There is some evidence that psychological stress may modulate immune function in humans and experimental animals [116]. It has been reported that stress induced anxiety is related to T helper type 1 response. A study that applied psychological stress prior to immunization suggested that stress exerts an adjuvant effect on dendritic cells by promoting enhanced migration to lymph nodes and resulting in increased antigen-specific T cell responses [117]. This effect may be mediated by the release of norepinephrine from the sympathetic nervous system Limitations of previous studies of environmental risk factors in PsA There were only two studies that assessed environmental exposures among PsA and psoriasis patients and one that compared PsA to rheumatoid arthritis. These studies were limited in several aspects [76, 77, 118].First, due to their small sample size these studies were underpowered to detect important associations. Secondly, the use of an administrative database as a source of information [77] may have led to an underestimation of events that are often not coded or are coded incorrectly. Lastly, as PsA is a common condition among psoriasis patients, a potential misclassification of cases and controls might have occurred

27 15 since psoriasis patients were not assessed to rule out the presence of inflammatory arthritis [76, 77]. It has been shown that PsA is underestimated among psoriasis patients, as approximately 18% of the psoriasis patients that were found to have inflammatory arthritis were unaware of their condition and had never been seen by a rheumatologist [119]. The problem of misclassification of patients and controls may significantly affect the results in case-control studies where the outcome (PsA in this case) is not rare among the control group. Misclassification may decrease the power of the study to detect significant associations

28 Genetics of Psoriasis and PsA The Approach to identification of genetic determinants of a disease A series of steps is required for the identification of susceptibility genes for a disease [120]. The first step involves determination of a familial aggregation. Segregation analysis is the next step, which examines the patterns of disease in families and determines if the patterns are indicative of traditional genetic inheritance models (such as autosomal dominant or recessive) or are more consistent with multi-factorial (polygenic and environmental) models [121]. Finally, once enough evidence of a genetic component in the cause of the disease has been obtained, the next step is to locate and identify any causative gene [122]. Two basic study designs are used to identify susceptibility loci: linkage analysis and association studies. Genetic linkage analysis relies on the tendency for short haplotypes to pass intact, without recombination events, to the next generation within families. A genetic marker that passes down though a family accompanied by the disease of interest, suggests that the susceptibility gene for the disease is located close to that marker [123, 124]. An association analysis is based on a putative functional genetic variant in the pathophysiologic mechanism of the disease of interest. Association studies look for a significant increase or decrease in frequency of a marker allele, genotype or haplotype within a disease trait than would be expected by chance if there were no association between markers and phenotype. This method can be applied to data obtained from families or populations of healthy individuals and patients with the disease of interest [125, 126]. For complex diseases, segregation and linkage analyses have not proven to be generally useful [127, 128]mainly due to low statistical power and the need to recruit large samples. Therefore, for these diseases, once familial aggregation is demonstrated, it may be appropriate to proceed with association studies, either candidate gene or genome-wide, to determine susceptibility loci/genes. Once a gene is identified, functional studies in biological systems are required to characterize the gene.

29 Familial aggregation of Psoriasis and PsA A study on familial aggregation is the first step in pursuing a possible genetic etiology to a disease. Familial aggregation is assessed by the recurrence risk ratio (λ). The recurrence risk ratio (λ R ) is defined as the prevalence of the disease in relatives of type R of affected cases divided by the prevalence in the general population [ ]. Family and twin studies have clearly demonstrated that psoriasis has a strong genetic basis. Two large scale epidemiological studies revealed a substantially higher incidence of psoriasis in relatives of patients with psoriasis compared to the general population. The recurrence risk for affected siblings (λ s ) was estimated to be between 4 and 10 [ ]. Twin studies reveal a concordance rate for monozygotic twins to be between 62 to 70% compared to 21 to 23% for dizygotic twins [ ]. A segregation study concluded that a polygenic or a multifactorial pattern is the most likely mode of inheritance [138, 139]. The genetics of PsA has been much less investigated compared to that of psoriasis. However, epidemiological studies have implicated a strong genetic basis to PsA. Four family studies have investigated the recurrence risk of PsA within families. The first study that demonstrated familial aggregation of PsA was published in 1973 and found that the prevalence of PsA among first-degree relatives of probands with PsA was 5.5% compared to the estimated prevalence in the UK population of 0.1%. The calculated recurrence risk ratio in first degree relatives (λ 1 ) was 55, compared with estimates ranging from 5 10 in cutaneous psoriasis [140]. Another study from the UK, reported a PsA prevalence of 14.3% among siblings and recurrence risk ratio (λ s ) of 47 (based on estimated PsA prevalence of 0.3% in the general population) [141]. Recently, the risk ratio for PsA was estimated using the Icelandic genealogical database. First degree relatives to fourth-degree relatives of patients with PsA had risk ratios of 39, 12, 3.6 and 2.3, respectively (all P-values < ), reflecting a strong familial aggregation, whereas the fifth-degree relatives had an RR of 1.2 (P = 0.236) [142]. Finally, our PsA cohort was assessed for familial aggregation. All

30 18 available first degree relatives of 100 PsA patients were assessed for evidence of PsA and psoriasis. Among the 289 first degree relatives that were evaluated, 7.6% had PsA leading to a calculated recurrence risk ratio in first degree relatives (λ 1 ) of 30.4 [143]. There is only one twin study in PsA from Denmark that evaluated 36 complete twins that included one proband with PsA. In this study no difference was found in the concordance rate between mono- and dizygotic twins [144]. However, this study may have been underpowered to detect a genetic effect Mode of inheritance Psoriasis and PsA are consistent with a multifactorial pattern of inheritance [145]. Several studies have suggested an autosomal dominant [146] and autosomal recessive pattern [147] of inheritance for psoriasis, however, even in these studies; a multifactorial model could not be ruled out. Genomic imprinting, a non-mendelian mode of transmission, has been suggested to play a role in the inheritance of psoriasis and PsA [148, 149]. Genomic imprinting refers to an epigenetic effect that causes differential expression of a gene depending on the sex of the transmitting parent. The imprinting process allows gene expression from only the maternally or paternally derived chromosome [150]. A family study from the Faroe Island reported a higher penetrance of psoriasis if the father was affected or a presumed gene carrier [151]. A similar phenomenon has also been reported in PsA patients. In our cohort, the proportion of PsA patients with an affected father was significantly higher than the expected proportion (0.65 vs. 0.5 p=0.001) [148]. These findings were supported by a linkage study in PsA that noted a significant linkage on chromosome 16q only after conditioning for paternal transmission [152]. Thus, there are some evidence that this epigenetic phenomenon may play a role in the inheritance pattern of psoriasis and PsA.

31 Genetic model for the relationship between PsA and Psoriasis There is no doubt that there is a close relationship between psoriasis and PsA however the nature of the relationship is not completely clear. Several models have been suggested to describe the genetic and clinical association between psoriasis and PsA [153]. The first model suggests that PsA and psoriasis should be viewed as two distinct conditions with different risk factors with the psoriasiform skin manifestation common to each. The model explains the families with large numbers of psoriasis cases without any PsA, on the other hand families with cases of both PsA and psoriasis or only PsA. This model predicts that case series ascertained by their psoriasis will show genetic heterogeneity. However case series ascertained by their PsA will share certain susceptibility genes and all PsA will be genetically distinct from psoriasis. The second model is based on the hypothesis that the genetic determinants of the skin and the joint diseases are independent. However, the presence of one of these factors leads to a lower threshold for the development of the other. This model explains the presence of undifferentiated spondyloarthritis which is very similar to PsA but does not involve the skin. This model predicts that case series ascertained by their psoriasis will be genetically heterogeneous and those ascertained by the presence of PsA will be homogeneous and share only some of the genes with psoriasis. The third model views PsA as a disease within a disease with psoriasis as the parent disease. PsA is considered a more severe phenotype of psoriasis that occurs due to a greater number of susceptibility genes or environmental factors. The additional genes or environmental risk factors on the background of psoriasis will lead to development of PsA. This model predicts that case series ascertained by psoriasis will be genetically homogenous and that case series ascertained by PsA will completely overlap those of psoriasis apart from several additional distinct genes.

32 Gene identification studies - Linkage analysis studies Genetic linkage analysis can be used to identify regions of the genome that contain genes that predispose to disease. The aim of this method is to isolate the disease gene by its chromosomal location without any prior knowledge of the position or function of the gene. Linkage analysis requires collection of families with multiple affected individuals. There are two methods for linkage analysis: parametric (model-based) linkage analysis and nonparametric linkage analysis. Parametric linkage analysis requires constructing a model to explain the inheritance of a disease in the pedigrees and then estimating the recombination fraction for a given pedigree. For multifactorial diseases, where several genes (and environmental factors) might contribute to disease risk, there is no clear mode of inheritance [123]. Methods to investigate linkage have therefore been developed that do not require specification of a disease model. Such methods are referred to as non-parametric, or model-free. In this method, affected sibling pairs are assessed for allele sharing. The premise for the non-parametric method is the fact that in the presence of linkage between a marker and disease, sets of relatives who share the same disease status are more likely to share alleles at the marker locus than the value of 50% that would be predicted by chance [154] Linkage analysis in psoriasis Both parametric and non-parametric methods have been used to investigate the genetics of psoriasis. The psoriasis susceptibility loci that have been mapped using linkage methods include: PSORiasis Susceptibility locus (PSORS)1 on 6p21.3 [151, 152], PSORS2 on 17q [155, 156],PSORS3 on 4q [157], PSORS4 on 1q21 [158], PSORS5 on 3q21 [159], PSORS6 on 19p [160], PSORS7 on 1p [161], PSORS8 on 16q [162], PSORS9 on 4q [163] and PSORS10 on 18p11 [164]. By far the strongest association is with a locus within the MHC on chromosome 6p21 (PSORS1). Additional putative psoriasis candidate loci have been reported on 16q and 20p [165]. The loci on 6p and 17q have been replicated with independent linkage studies and a meta-analysis of previous studies has found an increased

33 21 allele sharing for 16q [155, 163, 165]. However, the rest of the loci were found to be difficult to replicate. With regard to PsA, however, only one genome-wide linkage scan has been conducted [152]. This study identified a locus on 16q close to the PSORS8 locus identified for psoriasis, but only when conditioned on paternal inheritance. Thus, it is unclear whether there are distinct susceptibility loci for psoriasis and PsA. Maximum LOD score (MLS) analysis of affected sibling pairs yielded allele sharing of 60% for markers within the MHC region. These results emphasize the importance of the MHC region as a candidate susceptibility region in psoriasis. In that study, evidence of allele sharing was also found on 16q and 10q22-q Gene identification studies - Association Studies Genetic association studies aim to detect associations between one or more genetic polymorphisms and a trait. Traditionally, linkage analysis studies were performed for coarse mapping as they have a limited genetic resolution of 1 cm. Association studies were the next step for fine mapping. However, association studies have several advantages over linkage studies. No assumption about the mode of inheritance is required, they have greater power to detect small effects and they do not require large affected families. Therefore, association studies are often used initially for the investigation of complex diseases [126]. There are two basic approaches. The first one is to determine the association between a candidate gene and a trait. This approach is based on prior knowledge from linkage studies or the biology of the disease. The other approach is to perform hypothesis free Genome Wide Association Study (GWAS), where thousands of genetic markers, single nucleotide polymorphisms (SNPs), throughout the genome are tested individually for their association with the disease. The association between genotype and phenotype can be explained either by direct biological action of the polymorphism or by allelic association between the marker and a susceptibility gene. The term Linkage Disequilibrium (LD) is used to refer to allelic association between linked loci. Linkage disequilibrium is the hallmark of association