High prevalence of protein tyrosine phosphatase non-receptor N22 gene functional variant R620W in systemic lupus erythematosus patients from Kuwait: implications for disease susceptibility

Systemic lupus erythematosus (SLE; MIM 152700) is a common, complex autoimmune disease with multiple-organ involvement, characterized by the production of pathogenic autoantibodies directed against cytoplasmic and nuclear cellular components [1]. It occurs predominantly in women (> 90%) and is distinguished by a loss of tolerance to self-antigens, the deposition of immune complexes and tissue inflammation and destruction. There are variable clinical manifestations associated with SLE which include arthralgia, rashes, alopecia, serositis, leukopenia and renal involvement [2, 3]. Unrestricted hyper-activation of the immune system may lead to the overproduction of autoantibodies, immune complex deposition, inflammatory cytokine release and eventually organ damage in SLE pathogenesis [4, 5]. The causes of SLE are not completely understood at present. It is thought that the onset of SLE results from multifactorial etiology, involving hormonal factors, environmental triggers and genetic susceptibility [6]. Systemic autoimmune diseases e.g. SLE and many others are thought to share common genetic causative factors in populations of different ethnic or racial origin [7]. The involvement of major histocompatibility complex (MHC) also supported this hypothesis. However, non-MHC alleles such as a functional polymorphism (rs2476601, Arginine 620 Tryptophan, R620W) in protein tyrosine phosphatase non-receptor type 22 (PTPN22), which encodes the lymphoid PTP (LYP), was reported to be significantly associated with multiple autoimmune disorders including type 1 diabetes mellitus (T1DM; [8]), rheumatoid arthritis [9], SLE [10], Hashimoto thyroiditis [11], autoimmune thyroid disease [12] and Grave’s disease [13].

The PTPN22 gene encodes a lymphoid-specific phosphatase (LYP) which has been shown to be a negative regulator of T cell activation [14]. The LYP contribute to T cell activation by binding to the regulatory Src tyrosine kinase (Csk) [14]. It has been reported that a functional variant R620W (C1858T polymorphism) of the PTPN22 gene is located in a motif which facilitate binding of the LYP with Csk [15]. The variant form, 620 W carries a tryptophan residue in place of arginine (R) and affects the binding of LYP and Csk thereby disrupting the regulation of T cell receptor-signaling kinases e.g. Lck, Fyn and ZAP70 [14–16]. It has been postulated that the individuals carrying the variant allele (620 W) have an altered threshold for thymic selection, with increased numbers of autoreactive T cells escaping the negative selection, thus persisting in the circulation, and are prone to autoimmunity [17]. The C1858T variant has also been shown to be associated with changes in cytokine profile in SLE patients in vivo [18].

In spite of a large number of reports linking the PTPN22 gene R620W functional variant with a number of autoimmune diseases, conflicting results have appeared in the literature about its prevalence in different world populations [3, 5, 7, 19–22]. In view of these diverse findings on prevalence and relationship of the PTPN22 gene R620W functional variant with SLE, we carried out this study in SLE patients from Kuwait to investigate its possible association with susceptibility to SLE in a completely different population/ethnic group.

The SLE patients were recruited from two major teaching hospitals (Amiri and Mubarak Al-Kabeer) from Kuwait. The inclusion criteria described by the American College of Rheumatology (ACR) for the diagnosis of SLE was used [23]. For the patients included in the study, a diagnosis of SLE had been made at least 2–3 months prior to their recruitment. The information collected from the patients included age, age-at-onset, gender, clinical manifestations associated with SLE and disease severity. The evaluation of SLE severity was made as follows: the patients were considered to have a ‘mild disease’ if they presented muco-cutaneous serositis, and/or arthritis and ‘mild-severe’ disease if the patients had hematological, renal and neurological manifestations. The renal disease associated with SLE was ascertained if the 24 h urine protein excretion exceeded 500 mg or hematouria was detected (> 5 red blood cells/field). The hematological abnormalities associated with SLE included the presence of hemolytic anemia with reticulocytosis, or leucopenia (< 4000/mm³ on > two occasions), or lymphopenia (< 1500 mm³ on > two occasions) or thrombocytopenia (< 100,000/mm³ in the absence of drugs). The presence of seizures or psychosis without the use of drugs or metabolic derangement were considered as the neurological presentations associated with SLE. The frequency of PTPN22 gene polymorphism in SLE patients was compared to that in a group of 214 healthy controls, matched for age and sex with the patients. The controls subjects were unrelated to the patients; were otherwise healthy and were seen at hospital outpatient clinics for minor illnesses. The controls were thoroughly examined by a specialist to ascertain their health status before recruitment in the study.

The most significant finding in this study is that a relatively high frequency of PTPN22 gene functional variant 620 W was detected in the SLE patients (17.4%) compared to 0.9% in the controls. When taken together, the homozygous and heterozygous combinations of this variant allele were detected in 35.7% of the SLE patients from Kuwait. This is significantly higher than previous reports from other populations/ethnic groups. Machado-Contreras et al. [5], did not find homozygous TT genotype in any of their Mexican SLE patients, however heterozygous CT genotype was detected in 6% Mexican SLE patients. Similarly, in a Greek study on the SLE patients from the Island of Crete, TT genotype was not detected at all while CT genotype was detected in 5.39% cases. In a report on SLE patients from Egypt [26], the authors could not detect a homozygous TT genotype but did find heterozygous CT genotype in 52.5% SLE cases. A comprehensive PTPN22 gene association study [3], reported minor allele frequency (MAF) of 11.9% in European Americans, 2.1% in African Americans and 7.6% in Hispanics. This study also reported that the association of the 1858 T allele with SLE was greater in European American patients with familial SLE (11.9%) compared to the sporadic SLE (8.2%). A study from Poland [21] found a positive association between the variant allele of the PTPN22 gene and SLE. Aksoy et al. [22] did not find the TT genotype in Turkish SLE patients while the CT genotype was detected in 7% cases. Another study, on white Americans [27], reported that the TT genotype was detected in 2.5% SLE patients while the CT genotype was found in 20.4% SLE patients. In this study, the overall risk-allele frequency was found to be 12.67% compared to 8.64% in the white American controls. Positive association between PTPN22 gene polymorphism and SLE susceptibility has been reported from Sweden [28] and Spain [29]. Namjou et al. [29] reported findings from genotyping of ten SNPs in four large multi-ethnic populations and using their results in conjunction with data from the Hap-Map project, concluded that SLE association with PTPN22 was largely accounted for by the R620W variant (rs2476601) in individuals of European ancestry [21–26, 30, 31]. A strong North-South gradient in the risk-allele frequency has also been reported in Europe [32].

Recent reports provide information on the mechanism of R620W functional variant of the PTPN22 gene [33–38]. In the light of postulated scheme for involvement of R620W variant in the molecular mechanism of autoimmunity [33, 34], our data from SLE patients from Kuwait supports and highlights its role as a significant determinant of the SLE susceptibility. However, it may also be appropriate to mention that the genetic factors which contribute to susceptibility/protection to develop SLE most likely involve multiple genes. Kuwait is a small country located in the North of Arabian Gulf. The population of Kuwait is quite diverse; Kuwaiti Arabs constitute nearly 45% of the population. There is a high incidence of consanguinity (54%; [39]), which often results in familial clustering of common chronic disorders. The incidence of diseases with autoimmune etiology such as type 1 diabetes mellitus is amongst the highest in the region and bordering with the high-incidence countries of the world [40, 41]. The original settlers of Kuwait were immigrants from Najd, an area that now constitute eastern and central Saudi Arabia. The ethnic origin of Kuwaiti Arabs is quite varied; 50% are of Arab origin, some are Bedouins and nearly 50% are immigrants [42]. The Arabs residing in most of the Gulf countries are a result of an admixture with other populations such as Persians, Turks, South Asians, Europeans and Africans [42]. The genotype frequency of variant 620 W reported in our SLE patient group from Kuwait is amongst the highest (TT, 17.4% and combined TT and CT in 35.7% patients) compared to any of the other populations/ethnic groups. This can possibly be due to a cumulative effect of unique genetic/ethnic background along with a very high rate of consanguinity (54%) in the Kuwaiti population and can at least in part, explain the high incidence of autoimmune diseases including SLE in the country.

The frequency of PTPN22 gene functional variant R620W reported in this study is amongst the highest compared to other world populations. A high prevalence of this variant in SLE patients in comparison to controls suggests its significant contribution in conferring susceptibility to SLE along with other factors.