Summary

Overactive bladder (OAB) affects approximately 13% of adults [1], with a major impact on quality of life, equivalent to diabetes or rheumatoid arthritis [2, 3]. There is an urgent need for objective biomarkers to help us predict occurrence, progression, and therapeutic response. beta3-adrenoceptor gene (ADRb3), transient receptor potential vanilloid 4 (TRPV4) and nerve growth factor beta polypeptide gene (NGFB) single nucleotide polymorphisms (SNPs) together represent highly plausible candidates as genetic population risk factors for OAB. The initial phase of this study will use existing DNA, banked from the ALSPAC cohort. Using mailings of a contemporary validated self-completion questionnaire, to women in the sample, the current prevalence of OAB will be established. Genotyping will be performed for all participants (including men) for 41 SNPs (see appendix), characterising all known common variation around the ADRb3, TRPV4 and NGFB loci. The study will assess the relationship of these variants to OAB prevalence, OAB onset, and OAB severity, while controlling for known risk factors for OAB. The large size of the biobank, and the detailed follow-up of the sample over the last 20 years, provides a unique opportunity to assess the relationship of genetics and the environment in the natural history of OAB. The data should allow us to predict disease susceptibility, lead to improved treatment with current agents, and provide evidence for new potential drug targets. We therefore believe the study will have dramatic impact on our ability to diagnose and treat patients suffering with OAB.

Background

Cross-sectional studies [4] and classical twin studies [5] have suggested that OAB is highly heritable, with an affected first degree relative being associated with 1.3-1.9 fold increased risk, however no candidate genes have been identified. This study proposes to investigate three potential genetic biomarkers. beta3-adrenoceptors are the principle mediator of detrusor relaxation [6], and the YM-178 beta3-adrenoceptor agonist is in development for treatment of OAB [7]. The beta3-adrenoceptor gene (ADRb3) has several common SNPs, including the rs4994 polymorphism, which is associated with diabetes and obesity. In a single small sample is has also reported to be more common in women with OAB (OR 3.0, RR 2.1, n=201) [8], indicating the possibility of a shared susceptibility to OAB and metabolic syndrome. TRPV4 functions as a mechano-sensitive channel in the urothelium, with TRPV4 agonists inducing bladder overactivity in mice [9]. It is considered a key target for future OAB drug development. Functional TRPV4 SNPs have diverse pathological associations, including obstructive airways disease [10], and hyponatraemia [11]. NGF is one link between tissue inflammation and bladder C-fiber afferent nerve excitability. Urinary NGF levels correlate with multiple measures of OAB disease severity [12], and demonstrate responsiveness to therapy. A common functional SNP of the NGFB gene, rs6330, is associated with conditions linked to OAB, including anxiety traits [13] and multiple sclerosis [14]. ADRb3, TRPV4 and NGFB SNPs therefore together represent highly plausible candidates as genetic population risk factors, and require investigation in a population based sample.

Objectives

This study will characterise all known common variation around the ADRb3, TRPV4, and NGFB loci, with particular interest in the rs4994, rs12578401 and rs6330 variants, in a population based study of OAB symptoms in women. We will identify the role of ADRb3, TRPV4, and NGF SNPs and haplotypes in the pathophysiology of OAB.

Main hypothesis:

Common functional variation of the ADRb3, TRPV4 and NGFB genes is associated with increased risk of OAB in women.

Secondary hypotheses:

Common functional variation of the ADRb3, TRPV4 and NGFB genes is associated with alterations in disease phenotype in women with OAB:

* earlier onset of adult symptoms,

* later remission of childhood enuresis

* increased subjective severity and bother of OAB symptoms

Common functional variation of the ADRb3, TRPV4 and NGFB genes modifies environmental risks for OAB, including parity, smoking, weight gain and dietary habits.

Methodology

Design: Population based genetic association study of approximately 15,000 mothers and daughters.

Genotyping: KBiosciences will genotype 41 SNPs (see appendix) around the ADRb3, TRPV4 and NGFB genes.

Case assessment: Up to 3 mailings of the 12 item validated International Consultation on Incontinence - Female Lower Urinary Tract Symptoms Questionnaire, for self completion, with OAB cases identified using item 5a: greater than = "nine to ten" voids per day, and item 3a: "sometimes" "most of the time" or "all of the time" sudden need to rush to the toilet to urinate. Existing items in the ALSPAC database on urinary symptoms in mothers, asking about incontinence and frequency will be used to assess the onset of OAB, and items about bed wetting in children will be used to assess the remission of childhood enuresis.

Assessment of confounders: The existing database includes 1-2 yearly details from 1991 for mothers and daughters of birth records, co-morbidities, medication use, dietary habits (food, alcohol, caffeine), smoking, socioeconomics, ethnicity, and weight gain.

Statistics: Genotype data will be analysed using Hardy-Weinberg Equilibrium testing, and Cochran Armitage trend test with input and advice from statistical geneticists. The sample size is powered on the basis of the minor allele frequencies for the rs4994 SNP. With the frequency estimated at 10%, and alpha set at 0.05, an estimated sample size of 10000 provides 90% power to detect a 3% difference in the allele prevalence.