Chronic rhinosinusitis (CRS) is a frequent and important chronic disease, which causes significant patient discomfort and morbidity. Patients with CRS report a significantly lower quality of life index in measures of bodily pain and social functioning than do patients with congestive heart failure, angina, chronic obstructive pulmonary disease, and back pain 1 . In health surveys, CRS affect more than 10% of individuals in western countries, with an overall direct cost of CRS in the United States annually estimated at $4.3 billion 2 3 .

Despite the frequency of CRS and the costs associated with its care, there is limited knowledge about the initial development of disease. Our current conception of CRS is as of a chronic inflammation of the paranasal sinus membrane, which is colonised with nasal and exogenous bacteria that may contribute to the disorder. Histologically, CRS is characterized by an accumulation of inflammatory cells that are mainly eosinophils, with a neutrophilic infiltrate only during acute infections. Numerous pro-inflammatory cytokines and chemokines, both Th-1 and Th-2 profiles, are over expressed in CRS. These include IL-2, IL-4, IL-5, IL-6, IL8, IL-13 and IL-16 4 .

Current knowledge of the disease is limited to that garnered by study of biopsy samples obtained from patients affected with well-established disease, with little information available on early events or environmental factors associated with development of disease. Genetic studies may thus offer a better insight into the pathogenesis of the disorder. Recently, the primary interest in genetics has changed from simple Mendelian diseases, for which genotypes of some gene are the cause of disease, to more complex diseases, for which genotypes of some set of genes together with environmental factors merely alter the probability that an individual gets the disease, although individual factors are typically insufficient to cause the disease outright 5 .

Reports of familial clustering of CRS support a genetic basis for the disease 6 7 8 . The high prevalence of patients with CRS in well-defined genetic disorders such as cystic fibrosis (CF) and primary cilia dyskinesia (Kartagener's syndrome) also supports the concept that mutated genes play a role in the pathogenesis of the disease 9 . However, it is suspected that CRS generally encountered in clinical practice is due to a complex interaction of multiple genes rather than the single gene anomalies as observed in CF and Kartagener's syndrome. A polygenic model for nasal polyps was recently confirmed by segregation analysis 10 . The same study also confirms that nasal polyp is a highly heritable disease with heritability coefficients above 60%. Interestingly, the quantitative contribution of genetic factors seems to be higher in young and recurrent patients 6 10 . A number of genetic association studies have been conducted to elucidate the genetic loci conferring risk of CRS with or without nasal polyposis. Significant difference in allele frequencies between patients and controls were observed for polymorphisms located in the HLA 11 12 13 14 , CFTR 15 , LTA 16 , TGFB1 17 , IL1RN 18 , and ADRB2 19 . However, all these findings remain to be confirmed in larger populations.

It is possible that cysteinyl leukotrienes (cysLTs) may play a disease-regulating role in rhinosinusitis. CysLTs, described primarily as lipid mediators involved in pathogenesis of airway inflammation and related symptoms such as bronchoconstriction, mucus secretion, and airway hyperresponsiveness, are recognized now as important mediators of cell trafficking and innate immune responses 20 . CysLTs are synthesized through the lipoxygenase (LO) pathway of arachidonic acid metabolism. The LO pathway for synthesizing cysLTs has several steps, and distinct enzymes are involved in each steps, namely arachidonate 5-lipoxygenase (ALOX5), arachidonate 5-lipoxygenase-activating protein (ALOX5AP or FLAP), and leukotriene C4 synthase (LTC4S) 21 . Effects at the cellular level are mediated via two G-protein coupled receptors termed cysteinyl leukotriene receptor 1 (CYSLTR1) and cysteinyl leukotriene receptor 2 (CYSLTR2) (Figure 1).

In this initial study, we propose to study the genes regulating the leukotriene pathway to identify potential polymorphisms involved in disease susceptibility. We hope that identification of genetic polymorphisms involved will lead to improved comprehension of disease pathogenesis, and may eventually lead to more effective treatment, screening, and prevention of CRS.

210 patients with severe CRS were recruited. Four were not included in the analysis, as phenotypic questionnaires were incomplete. Table 2 shows the characteristics of cases and controls. For cases, initial diagnosis was recurrent rhinosinusitis in 12.1%, CRS without polyposis in 13.1%, and CRS with polyposis in 74.8%. Average number of previous surgeries was 3.15 (range 1 – 20; median 2.0), with an average age at first surgery of 38.3 years (range 1 – 76; median 38). Medication use in this group was high, with 73.4% of patients requiring medication beyond an inhaled nasal corticosteroid for disease control and 23.3% remaining uncontrolled despite medication. Atopy was present in 65.5% and asthma requiring treatment in 53.9%. Active smoking was present in 11.2%. Patients reported blood relatives affected with CRS in 37.7% of instances.

Measured serum biomarkers showed mean circulating eosinophilia of 4.4% (range 0 – 25%; median 4.0%) with 33.5% of patients demonstrating > 5% eosinophilia. Mean total serum IgE was 155.9 IU (range 2 – 1460, median = 85.0) with 41.1% having levels IgE > 120 IU/ml.

In this study, we have analyzed SNPs covering 5 genes implicated in the cysLT pathway in a case-control series of patients with severe CRS. After adjustment of multiple testing, there were no significant associations observed between severe CRS and genetic variants within these genes. Rather than suggest that there is no role for polymorphisms in the leukotriene cascade, it suggests that the effect of polymorphisms in cysLTs is less than the power detectable by our population size, which is well-powered to detect only polymorphisms in common alleles which confer a relative risk (RR) of 3.0 or greater, and only moderately powered to detect polymorphisms with a RR of 2.0. In this study, the minor allele of the most significant polymorphisms conferred a RR of 1.45 (ALOX5-rs3780894), 1.67 (ALOX5AP-rs17612127) and 0.69 (CYSLTR1-rs321090). Most genes for complex diseases that have been described have an effect in this range or less.

Although SNP CYSLTR1-rs321090 is not significant after multiple testing correction, the results observed in the current study replicated the previous association reported by Hao et al 27 . In fact, the polymorphism, labeled 927T/C (rs320995) was significantly associated with atopy severity in a group of asthmatic families. Interestingly, this polymorphism is in tight linkage disequilibrium (LD) with rs321090 (r² = 0.82 based on the CEU genotyping dataset of the International HapMap project 23 ). In both studies, the common allele is more frequent in cases than controls. In contrast, the current study does not corroborate the previous association between SNP LTC4S-rs730012 (also known as -444A/C) and allergic rhinitis 28 . However, a number of controversies exist concerning the role of this polymorphism in allergic-related diseases 29 30 . Accordingly, larger and well-powered studies will be required to determine the role of this polymorphism in pathogenesis of these diseases.

Beside the lack of power, other factors may explain the absence of significant results. First, we have studied what is a relatively heterogeneous population, with several different initial diagnoses, some unresponsive to therapy effective in others and different levels of serum biomarkers of inflammation. It is possible that the effect of cysLTs may be different within these groups and less noticeable in the group as a whole. Analysis of subgroups within our population may be of benefit but will be limited by sample size. Secondly, the control group utilized must also be considered. The group used has been selected for comparability in terms of environment rather than for absence of the disease. With the prevalence of self-reported chronic sinusitis being reported as high a 16% and the incidence of allergies being over 20%, confusion with these other disorders may make it hard to differentiate from these other disorders.

Leukotrienes have nevertheless been implicated in CRS in other models and several studies have shown that leukotrienes may be involved in the development of the disorder. CysLT metabolites, leukotriene C4 (LTC4), leukotriene D4 (LTD4), and leukotriene E4 (LTE4) likely contribute to the pathogenesis of chronic rhinosinusitis through their effects on microvascular leakage, epithelial cell activation, elevated mucus secretion, and mucosal inflammation 31 . CysLTs also appear to be related to the severity of eosinophilic inflammation 32 .

CysLTs appear related to nasal polyposis. In an examination of 27 patients by Kaplan et al. 33 patients with sinonasal polyps were noted by radioimmunoassay to have elevated levels of LTC4. Patients with sinonasal polyposis with recurrences within 18 months of surgery had higher levels of LTC4 when compared with those patients that did not have recurrences. In another study of 58 individuals, Steinke et al. 34 reported that chronic hyperplastic eosinophilic sinusitis is characterized by the increased presence of CysLTs when compared with concentrations seen in tissue from patients with chronic inflammatory sinusitis or healthy sinus tissue. Specifically implicating the polyp tissue as the source of these cysLTs, Higashi et al. 35 found that there were significant decreases in the urinary LTE4 concentrations before and after the sinus surgery for patients with chronic hyperplastic rhinosinusitis with nasal polyposis. In support of this, a recent study showed that urinary LTE4 concentration is highly significant in 10 patients with aspirin-intolerant asthma (AIA) with nasal polyps compare to patients with 24 AIA without polyps 36 .

Despite the evidence supporting a role for leukotrienes other authors have failed to demonstrate a role for cysLTs in CRS. A study by Kountakis et al 37 showed that mean cysLTs levels were similar in CRS patients with eosinophilic versus non eosinophilic and in patients with polyps versus without polyp. Also cysLTs levels did not correlate with severity of CRS according to CT findings, endoscopy findings, or symptom scores.

While these initial results do not support that polymorphisms in genes assessed involved in the leukotriene pathways are contributing to the pathogenesis of CRS, this initial study was not powered to detect polymorphisms with relative risk of 2.0 or less, where we could expect many gene effects for complex diseases to occur. Thus, despite this lack of significant association noted in this study, we believe that validation with external populations and the use of better-powered studies in the future may allow more conclusive findings.

AIA: aspirin-intolerant asthma, ALOX5: arachidonate 5-lipoxygenase, ALOX5AP: arachidonate 5-lipoxygenase-activating protein, CF: cystic fibrosis, CRS: Chronic rhinosinusitis, COX: cyclo-oxygenase pathway, CYSLTR1: cysteinyl leukotriene receptor 1, CYSLTR2: cysteinyl leukotriene receptor 2, cysLTs: cysteinyl leukotrienes, HWE: Hardy-Weinberg equilibrium, LD: Linkage disequilibrium, LO: lipoxygenase pathway, LTB4: leukotriene B4, LTC4: leukotriene C4, LTC4S: leukotriene C4 synthase, LTE4: leukotriene E4, RR: relative risk, SNP: single nucleotide polymorphism

The author(s) declare that they have no competing interests.

HAS was involved in sample collection and draft the first version of the manuscript. YB carried out SNP selection and genotyping, performed statistical analyses, and finalized the manuscript. TJH participated in the design of the study. MC, MD, SVS, SF were involved in sample collection and coordination of the case-control population. ML calculated the p-value threshold to ensure a project-wide type 1 error rate of 0.05. MD conceived the study, acquired the funding, and provided general supervision of the research group. All authors read and approved the final manuscript.