Old questions and new approaches

<p class="article-intro">Overactive bladder (OAB) is a clinical syndrome of urinary urgency, frequency, with or without urinary incontinence. Clinically, OAB can be diagnosed based on the symptoms reported by a patient; although subjective, these symptoms can be bothersome and negatively affect the quality of life. By 85 years of age, approximately 90 % of men will have benign prostatic hyperplasia (BPH) and benign prostatic obstruction (BPO) with lower urinary tract symptoms (LUTS) including OAB and urinary retention. </p> <p class="article-content"><div id="keypoints"> <h2>Keypoints</h2> <ul> <li>Since none of the potential urinary biomarkers assessed to date seem to have enough specificity and sensitivity to accurately diagnose OAB, we need a better understanding of the molecular mechanisms, including the key regulators and activated pathways characteristic of various types of OAB (obstructive, neuro&shy;genic or idiopathic; with and without DO).</li> <li>A combination of proteins rather than single molecules needs to be evaluated in order to increase the relia&shy;bility and discriminative potential of biomarkers.</li> <li>MiRNAs and small molecules need to be included into biomarker signat&shy;ures and validated for different types of OAB.</li> </ul> </div> <p>OAB is often associated with detrusor overactivity (DO) which is an objective, measurable characteristic of bladder dysfunction. DO is the urodynamic diagnosis of involuntary spontaneous bladder contractions which can cause the symptoms of OAB. However, the symptomatic diagnosis of OAB does not always correlate with DO, since fewer than half of patients with urgency are found to have DO and the remainder have symptoms but no such activity. In BPO-induced LUTS, DO is present in over 60 % of patients, and its prevalence increases with age and grade of obstruction to the estimated 83 % in men with severe bladder outlet ob&shy;struction.</p> <h2>OAB biomarkers</h2> <p>A biomarker is &ldquo;a characteristic that is objectively measured and evaluated as an indicator of normal biological processes, pathogenic processes, or pharmacological responses to a therapeutic intervention&rdquo;.¹ Accurate assessment of OAB is complicat&shy;ed due to the multifactorial aetiology of the disease, which can be obstructive, neurogenic or idiopathic. Identification and validation of a biomarker should be hypothesis-driven rather than serendipitous, implicating the need for better understanding of the molecular mechanisms of OAB. It is advantageous for a biomarker to be easily detectable in body fluids, such as urine. Urine is not subjected to homeostatic mechanisms and its contents reflect many changes of the body, such as pregnancy, aging, and dis&shy;ease. Urine is easy to collect non-invasively, and recently a number of studies examined urinary inflammatory mediators, ATP and neurotrophins as potential biomarkers of OAB. <br />Inflammatory mediators have been pos&shy;tulated to be involved in the pathogen&shy;esis of LUTS, and urinary PGE2 is significantly increased in OAB.² In a different study, urinary levels of cytokines MCP and the soluble fraction of the CD40 lig&shy;and were raised in OAB whereas IL-5, IL-6, IL-7, and GM-CSF were down-regulated.³ ATP is released by the urothelium in response to stretch and other stimuli, and has been suggested as a molecule transducing external stress of the detrusor. The amount of released ATP is signi&shy;f&shy;i&shy;cantly higher and the ATP catabolism by ectonucleotidases slower on the luminal side of the urothelium, which favours accumulation of ATP in the urine. In&shy;deed, the amount of ATP in the instilled fluid inversely correlated with the first desire to void in OAB patients.⁴ Neurotrophins are a protein family, which includes nerve growth factor (NGF), brain-derived neurotrophic factor (BDNF) and neurotrophins NT3 and NT4/5. NTP and BDNF in particular have been in focus of urologists seeking a reliable and sensitive biomarker for bladder pain, overactivity and related LUTS. Elevated NGF is detected in the urine of patients with OAB, BPO, bladder pain and DO, and its levels decrease upon treatment with anti-muscarinic agents and detrusor botulinum toxin injections (reviewed in^{5, 6}). The urinary NGF levels seem to correlate with DO or symptomatic OAB: the urinary creatinine-normalized NGF/Cr levels were very low in the control group and in patients with BOO/non-OAB and signif&shy;icantly greater in patients with BOO/OAB and BOO/DO.⁷ Expression of BDNF is affected by a variety of stimuli, including neuronal activity, inflammat&shy;ion, NGF-mediated signalling, ischemia and hypoglycemic insult. Similar to NGF, the levels of BDNF rapidly increase in the inflamed bladder, together with its high affinity TrkB and general p75 receptors, implicating its release in peripheral organs in the neurogenic inflammatory state.⁸ Acute intrathecal administration of BDNF induced bladder hyperactivity, and BDNF has been shown to be an important modulator of neurogenic DO.⁹ Urinary BDNF levels in pain and OAB patients behave similar to NGF, being elevated in the patients&rsquo; groups and re&shy;duced following treatment.¹⁰</p> <h2>New approaches to study OAB pathogenesis and biomarker discovery</h2> <p>Although considerable progress has been made characterizing OAB biomark&shy;ers, their sensitivity and specificity has been repeatedly questioned, citing the lack of standardisation of testing, no clear definition of &ldquo;normal&rdquo; range of urinary content, and absent normalisation to urinary creatinine in many studies. In order to gain a better understanding of the molecular mechanisms of DO-related OAB in BPO patients, we recently completed the quantitative transcriptome and miRNA profiling of different urodyna&shy;m&shy;ically-defined states of BPO in the human patients using comprehensive next-generation sequen&shy;cing (NGS-)derived gene expression data.¹¹ The gene expression profiles correlated well with the functional characteristics of the bladder, identified by urodynamics in BPO pa&shy;tients with detrusor overactivity (DO group), BPO without DO (BO group) obstructed patients with detrusor underactivity (UA group). Importantly, DO patients had symptomatic characteristics of OAB. <br />Gene expression changes at mRNA and miRNA level are indicative of activation of cell signalling pathways, leading to the morphological and functional changes in the bladder, and for BPO these included cytokine and immune response pathways, growth factor signalling (TGF-&beta;, HGF and IGF-1), activation of GPCRs (endothelin and cholecytokinin/gastrin pathways), NO signalling and hypertrophy-relevant PI3K/AKT signalling path&shy;ways. Interes&shy;tingly, BPO patients with DO and OAB symptoms showed activa&shy;&shy;t&shy;ion of neurotransmitters and other nervous system signalling, contrary to the other groups without overactivity (Fig. 1). In addition, smooth muscle contractility pathways were specifically and significantly in&shy;creased based on the mRNAs, altered in DO patients but not in the other groups without overac&shy;tivity.</p> <p><img src="/custom/img/files/files_datafiles_data_Zeitungen_2017_Leading Opinions_Uro_1701_Weblinks_s16.jpg" alt="" width="2150" height="1403" /></p> <p><br />We applied statistical tools for the unbiased identification of biomarkers of each functional phenotype, including patients with OAB (DO group). Three-mRNA (NRXN3, BMP7 and UPK1A) and three-miRNA (miR-103a-3p, miR-10a-5p and miR-199a-3p) signatures were identified based on ROC curve analysis of NGS data and validated by QPCR in an independent patient population. These signatures are sufficient to discriminate DO, BO and UA groups.¹¹ A blinded study was performed using an independent group of patients (n=21), including sam&shy;ples taken from the same patients before and after TURP (n=6), to assess the predictive performance of the signatures. A patient grouping based on the expression profiles of signature miRNAs and mRNAs in a blinded study was significantly correlated with the phenotypes confirming the findings. Importantly, both the three-mRNA and three-miRNA signatures in repeated biopsies of the same patients after deobstruction were in agreement with the changes in uro&shy;dynamic parameters. <br />Circulating miRNAs have been detected in the extracellular space and body fluids and hailed as biomarkers. Our results indicate that miRNAs play an important role in BPO, and identification of a three-miRNA miR-103a-3p, miR-10a-5p and miR-199a-3p signature of DO and other urodynamic phenotypes opens up a possibility of exploiting their potential as urinary OAB biomarkers.</p></p> <p class="article-footer"> <a class="literatur" data-toggle="collapse" href="#collapseLiteratur" aria-expanded="false" aria-controls="collapseLiteratur" >Literatur</a> <div class="collapse" id="collapseLiteratur"> <p><strong>1</strong> Biomarkers Definitions Working Group: Biomarkers and surrogate endpoints: preferred definitions and conceptual framework. Clin Pharmacol Ther 2001; 69: 89-95 <strong>2</strong> Kim JC et al: Changes of urinary nerve growth factor and prostaglandins in male patients with overactive bladder symptom. Int J Urol 2005; 12: 875-80 <strong>3</strong> Ghoniem G et al: Differential profile analysis of urinary cytokines in patients with overactive bladder. Int Urogynecol J 2011; 22: 953-61 <strong>4</strong> Cheng Y et al: Correlation between cystometric volumes, ATP release, and pH in women with overactive bladder versus controls. Neurourol Urodyn 2013; 32: 969-73 <strong>5</strong> Fry CH et al: What is the role for biomarkers for lower urinary tract disorders? ICI-RS 2013. Neurourol Urodyn 2014; 33: 602-5 <strong>6</strong> Seth JH et al: Nerve growth factor (NGF): a potential urinary biomarker for overactive bladder syndrome (OAB)? BJU Int 2013; 111: 372-80 <strong>7</strong> Liu HT, Kuo HC: Urinary nerve growth factor levels are increased in patients with bladder outlet obstruction with overactive bladder symptoms and re&shy;duced after successful medical treatment. Urology 2008; 72: 104-8; discussion 108 <strong>8</strong> Qiao LY et al: Inflammation and activity augment brain-derived neurotrophic factor peripheral release. Neuroscience 2016; 318: 114-21 <strong>9</strong> Frias B et al: The role of brain-derived neurotrophic factor (BDNF) in the development of neurogenic detrusor overactivity (NDO). J Neurosci 2015; 35: 2146-60 <strong>10</strong> Antunes-Lopes T et al: Urinary neurotrophic factors in healthy individuals and patients with overactive bladder. J Urol 2013; 189: 359-65 <strong>11</strong> Gheinani AH et al: Characterization of miRNA-regulated networks, hubs of signaling, and biomarkers in obstruction-induced bladder dysfunction. JCI Insight 2017; 2: e89560</p> </div> </p>