**5. The Rationale for BoNT-A Application in Chronic Prostatitis**

BoNT-A causes muscle relaxation by blocking the release of acetylcholine.

BoNT-A has been well known to have paralyzing effects via the blockage of acetylcholine release at the presynaptic cholinergic neuromuscular junction. The inhibitory effects of BoNT-A on both somatic and autonomic nerves have been used to treat a variety of conditions associated with muscularhypercontractility. Intramuscular injection of BoNT-A could lead to temporary chemodenervation and muscle relaxation in both striated and smooth muscles [29,30]. In 1988, Dykstra et al. first treated 11 patients with DSD due to spinal cord injury (SCI) by using urethral sphincter injection of BoNT-A [11]. In a prospective study using 100 U of BoNT-A injection into the urethral sphincter to treat patients with spinal cord lesions and DSD, decreased voiding detrusor

pressure and increased maximum flow rate were observed [12]; 60.6% of patients felt satisfied with the outcomes but post-injection urinary incontinence was another concern.

Huang et al. conducted a prospective multicentre trial with a total of 59 SCI patients having both detrusor overactivity (DO) and DSD. All the patients simultaneously received intravesical (200 U) and urethral sphincter (100 U) injections of BoNT-A [32]. This trial demonstrated significant reductions in maximum detrusor pressure, urinary incontinence episode, and improvement in voiding volume after treatment. All patients could achieve complete dryness at the follow-up point of 2 weeks. In view of the effect of BoNT-A on smooth muscle relaxation, the therapeutic response in DO has been widely studied. In 2000, Schurch et al. first reported the injection of BoNT-A (200 to 300 U) into detrusor muscle to treat patients with neurogenic DO [33]. At the 6-week follow up, 17 of 19 (89%) patients achieved complete continence. Significantly increased maximum bladder capacity and decreased maximum detrusor voiding pressure were observed after injection. To date, intravesical BoNT-A injection has become the preferred option for refractory OAB and neurogenic DO.

In addition to the role in blocking motor neurons, BoNT-A also has effects on the modulation of sensory nerves and inflammation as evinced by animal studies (Figure 1). A large body of evidence now supports that BoNT-A achieves analgesic effect by hindering the release of mediators responsible for painful sensation, including nerve growth factor (NGF), substance P, calcitonin gene-related peptide (CGRP), glutamate, as well as adenosine triphosphate (ATP) involved in afferent neurotransmission [22,23]. Therefore, it is postulated that BoNT-A may relieve the pain associated with chronic prostatitis by inhibiting the abnormal nociceptive neurotransmission conveyed by prostatic afferent nerves. Sensory dysfunction may be one of the postulated causes driving the symptoms of chronic prostatitis [6].

**Figure 1.** Conceivable mechanisms of BoNT-A in the treatment of CP/CPPS.

Transient Receptor Potential Vanilloid 1 (TRPV1), a non-selective cation channel, is expressed in some primary afferent neurons [34], especially in small and medium diameters (e.g., C-fibers) that are responsible for neurogenic pain and inflammation development [35]. In humans, TRPV1 activation might result in a burning pain sensation in the lower urinary tract [36]. Dinis et al. have demonstrated that abundant TRPV1 innervation are located on the prostatic urethral mucosa, verumontanum, and ejaculatory ducts [37]. Using rats in an adjuvant-arthritis pain model, Fan et al. reported that BoNT-A exerts its antinociceptive effect by reducing TRPV1 protein expression via the inhibition of plasma membrane trafficking [38]. Taken together, BoNT-A might play an important role in inhibiting TRPV1 expression in human prostate, which provides an alternative therapeutic strategy for CP/CPPS.

Regarding the anti-inflammatory effect, Chuang et al. reported that BoNT-A significantly decreased painful behavior, inflammatory cell accumulation, and cyclooxygenase (COX)-2 expression in the prostate and L6 spinal cord in dose-dependent fashion in a rat model of capsaicininduced prostatitis [24,25]. This finding demonstrates the potential of BoNT-A in the treatment of prostate inflammation.

### **6. Clinical Studies in BoNT-A for CP**/**CPPS Treatment**

Table 1 summarized the clinical studies of BoNT-A intraprostatic injection for chronic prostatitis. Maria el al. first reported the transperineal injection of 30 U BoNT-A into prostatic apex to treat the voiding problem in patients with chronic prostatitis in 1998 [39]. In their series, three of four patients obtained improvement in voiding without urinary incontinence with a mean follow-up duration of 12 months.



CP/CPPS, Chronic prostatitis/chronic pelvic pain syndrome; NIH-CPSI, National Institutes of Health Chronic Prostatitis Symptom Index; AUA-SS, American Urological Association-symptom score; VAS, visual analogue scale; QoL, quality of life; "—" indicates "not available"

Park et al. reported intraprostatic BoNT-A injection in 84 patients with CPPS using transrectal (40 U, *n* = 78) or transperineal (200 U, *n* = 6) route [40]. Symptom improvement was found in 59% of patients with transrectal injection and in 50% with transperineal route. By the National Institutes of Health Chronic Prostatitis Symptom Index (NIH-CPSI) questionnaire, the most significant improvement was in the pain domain, followed by ejaculation-related pain. The therapeutic effect was sustained for 6~18 months. The NIH-CPSI is a 13-item questionnaire developed to assess symptoms and quality of life (QoL) in men with CP/CPPS. This questionnaire includes three domains evaluating pain, urinary complaints, and QoL.

Gottsch el al. conducted a randomized placebo-controlled study to evaluate the effect of BoNT-A (100 U) injection into perineal body and bulbospongiosus muscle for CPPS treatment [41]. One month after treatment, the response rate assessed by Global Response Assessment (GRA) was significantly better in the BoNT-A group than the placebo group (30% vs. 13%, *p* = 0.0002). Although the NIH-CPSI pain subdomain was also significantly improved in the BoNT-A group, the total NIH-CPSI score did not improve.

In a prospective, randomized, double-blind, placebo-controlled trial on transurethral intraprostatic injection of BoNT-A (100 U) for men with chronic prostatitis, Falahatkar et al. demonstrated that BoNT-A treatment group significantly improved the NIH-CPSI total and subscale scores, the American Urological Association-symptom score (AUA-SS), visual analogue scale (VAS), quality of life, and frequencies of diurnal and nocturnal urinations when compared to baseline. The most noticeable improvement was the NIH-CPSI pain subdomain and the VAS scores, which decreased by 79.9% and 82.1% at 6-month follow-up, respectively [42].

Recently, another prospective controlled study using transurethral intraprostatic injection of BoNT-A (200 U) in 43 patients also showed encouraging response rates for refractory nonbacterial CP/CPPS [43]. However, the therapeutic effect gradually declined at 9–12 months.

As for the optimal injection route into the prostate, including transurethral, transperineal, and transrectal approaches, there is no definite suggestion so far. El-Enen et al. compared the transurethral route with transrectal intraprostatic BoNT-A injection for refractory CP/CPPS [44]. More significant improvement was observed in the transrectal group during the follow-up points. Their data also showed better results for BoNT-A injection in men with small prostates. Controversially, the aforementioned study showed that activation of TRPV1 might be one of the possible etiologies in CP/CPPS [36]. TRPV1-immunoreactive nerve fibers are distributed throughout the prostatic urethra mucosa, verumontanum, ejaculatory duct, and periurethral prostatic acini [37]. It is plausible that instead of transrectal injection, transurethral BoNT-A injection may be able to target TRPV1-immunoreactive nerve fibers as not only TRPV1-immunoreactive nerve fibers but other nerve fibers distributed in the transitional and peripheral zones may affect the efficacy of different injection routes. In addition, the drug extravasation out of the target site after injection is also an interesting issue to be discussed [45]. Further randomized and large-scale comparative studies are needed to evaluate the efficacy of different injection routes. The potential injection sites of BoNT-A for CP/CPPS treatment are shown in Figure 2.

**Figure 2.** The potential injection sites of BoNT-A for CP/CPPS treatment: intraprostatic, prostatic apex, external urethral sphincter, and pelvic floor muscles. (**A**) MRI, axial view (**B**) MRI, coronal view.

BoNT-A injection targeting at prostate and surrounding tissue is a safe procedure when administered by an experienced injector. Side effects are always transient, like local pain, hematuria, and urinary tract infection, and in the majority of cases they are mild and tolerable [17,19,20].
