Key words
overactive bladder - benign prostatic hyperplasia - imidafenacin - oxybutynin - urinary
flow
Introduction
Antimuscarinic drugs are widely used for treatment of overactive bladder (OAB), which
is characterized by storage symptoms, such as urinary urgency. Although the clinical
efficacy of antimuscarinics has been well established, their mechanism-based adverse
reactions often result in poor compliance and in turn limiting their clinical usefulness.
The most common and bothersome of such adverse reactions include dry mouth and constipation
[1]. Another significant concern and potentially harmful reaction of antimuscarinics
is acute urinary retention (AUR) in male patients, especially those with bladder outlet
obstruction due to benign prostatic hyperplasia (BPH), and indeed OAB symptoms frequently
accompany BPH [2]
[3]
[4]. Because muscarinic receptors (mainly via M3 subtype) play a pivotal role in contracting detrusor smooth muscles, antimuscarinics
theoretically have an ability to induce AUR [5]. In addition, BPH per se is typically responsible for AUR because of intrinsically
narrowed urethra. Thus, prescribing antimuscarinics for BPH patients could further
aggravate voiding dysfunction. Nevertheless, some data from recent clinical trials
suggest that the incidence of AUR in antimuscarinic therapy is infrequent (<1%), even
in male patients with BPH [1]. However, the results reported may have underestimated the true incidence of this
adverse reaction, because these trials were of short duration and included the patients
with low post-void residual urine volume (RV) at baseline. Indeed, Radomski [6] described, according to his database study that AUR risk is somewhat higher (the
rate of 2.5%) in real life clinical practice. Furthermore, above-mentioned clinical
trials have consistently been reported to increase the post-void RV in antimuscarinic
therapy, indicating the impairment of voiding function [1]. Therefore, despite the absence of definitive evidence, it is recommended that OAB
patients, especially those associated with BPH, should be carefully monitored for
post-void RV to avoid inadvertent AUR during the antimuscarinic therapy [7].
Adverse reaction profiles of the existing antimuscarinics may be different depending
on their selectivity for specific subtypes of muscarinic receptor, their tissue selectivity
(organ specificity) for bladder, their physicochemical properties, as well as their
pharmacokinetic properties [1]. In fact, clinical studies have shown that a new generation of antimuscarinics with
higher selectivity for the M3 muscarinic receptor subtype and/or bladder tissue has a comparable efficacy with
first-generation antimuscarinic oxybutynin but has fewer adverse reactions, mainly
dry mouth [8]. On the other hand, diminishing adverse effects on voiding function was probably
considered more difficult to achieve, because antimuscarinics had been traditionally
believed to exert their therapeutic action on OAB by reducing detrusor muscle contraction
via blockade of M3 subtype [9]. More recently, however, Finney et al. [10] found that at least in a range of usual therapeutic doses, antimuscarinics primarily
act during storage phase of micturition cycle, resulting in decreasing urgency and
increasing bladder capacity (BC), whereas they have little or no influence on detrusor
muscle contraction during voiding phase. Similarly basic researches with experimental
animal models of OAB have reported that low doses of antimuscarinics significantly
increase BC without influence on voiding contractions, and their increasing effect
is significantly inhibited by sensory denervation induced by resiniferatoxin [11]. These findings suggest that some regulatory involvement of antimuscarinics in afferent
nerve activity during the storage phase contributes to their therapeutic effect, but
such involvement may be absent in efferent activity during voiding phase.
Imidafenacin, 4-(2-methyl-1H-imidazol-1-yl)-2,2-diphenylbutanamide, belongs to a new generation of antimuscarinics
with high affinity for the M1/M3 muscarinic receptor subtypes and high bladder selectivity [12]
[13], and was approved for treating OAB in Japan in 2007. A randomized, double-blind,
controlled trial (RCT) demonstrated that imidafenacin is as efficacious as a first-generation
antimuscarinic drug propiverine but has better adverse reaction profiles [14]. Antimuscarinics are currently recommended as a useful addition to drug regimens
for BPH patients with predominantly OAB symptom. Recent RCTs in male BPH patients
with OAB symptoms reported that antimuscarinics, including propiverine, tolterodine,
and solifenacin, in combination with α1-adrenoceptor antagonist, caused AUR, though the incidence was considerably low (<2%
in all of these trials) [15]
[16]
[17]. In a similar RCT no patients experienced AUR when imidafenacin was combined with
α1-adrenoceptor antagonist tamsulosin [18]. And the post-void RV in this combination treatment was statistically different
from that in tamsulosin treatment alone [18]. These clinical findings led us to hypothesize that imidafenacin may have a different
degree of inducing risk of voiding dysfunction from other existing antimuscarinics.
It is practically difficult, however, to directly compare the clinical data of different
antimuscarinics each other, because of differences in various conditions, such as
the treatment protocol/design and the baseline severity of BPH. Therefore, to prove
our hypothesis, we evaluated the effectiveness (increasing the BC) and the voiding
dysfunction (decreasing the maximum urinary flow rate (Qmax)) in rats when treated
with imidafenacin and oxybutynin each alone, and compared their relative risk of voiding
dysfunction in terms of relative therapeutic index. Oxybutynin was selected for comparison
because it was a representative of the first-generation of antimuscarinic drugs that
has been intensively investigated for the AUR [19].
Materials and Methods
Animals
Female Sprague-Dawley rats (Charles River Laboratories Japan, Kanagawa, Japan), weighing
210–280 g, were housed in a room maintained under controlled conditions 23±3°C, 55±15%
RH, and 12:12-h light-dark cycle. The rats had free access to food pellets and tap
water. All animal care/experiments and the study protocol complied with the guidelines
of the Committee for the Purpose of Control and Supervision of Experiments on Animals
and were approved, prior to the study, by the Institutional Animal Ethics Committee
at Kyorin Pharmaceutical Co., Ltd. Our facility is certified as an animal testing
research institute by the Japan Health Sciences Foundation, established under the
jurisdiction of the Japanese Ministry of Health, Labor and Welfare.
Pressure-flow measurement in urethane-anesthetized rats with functional urethral obstruction
Pressure-flow study (a simultaneous recording of urinary flow rate and intravesical
pressure) was performed by a slight modification of the method reported by Streng
et al. [20]. Although Streng et al. used the continuous cystometry in their study, we used the
intermittent cystometry, because the continuous cystometry does not allow to measure
the residual urine volume. In brief, under urethane anesthesia (1.2 g/kg, s.c.), a
lower abdominal midline incision was made to expose the bladder. A saline-filled polyethylene
catheter (PE50; Japan Becton Dickinson, Tokyo, Japan) was inserted into the bladder
dome. The catheter was connected via a 3-way cock to a pressure transducer (Nihon
Kohden, Tokyo, Japan) for measurement of the intravesical pressure and to a syringe
pumps (Harvard Apparatus, Holliston, MA, USA) for saline infusion. The bone of symphysis
pubis was cut vertically to expose the urethra. An ultrasonic flow probe (MC2.5PSB;
Transonic Systems, Ithaca, NY, USA) was placed around the distal urethra and connected
to a transit-time flow meter (TS420, Transonic Systems) for the measurement of actual
urinary flow rate. The intravesical pressure and the urinary flow rate were recorded
at a sampling rate of 400 Hz using PowerLab system (AD Instruments Pty Ltd, Castle
Hill, Australia). The influence of antimuscarinics on the urinary flow rate was assessed
under a partial urethral obstruction induced by a selective α1-adrenoceptor agonist A-61603 (i. e., functional urethral obstruction), because in
the preliminary examination, treatment with each antimuscarinic drug alone even if
at higher doses resulted in too much residue of Qmax (approximately 50% to Qmax in
normal rats) to evaluate the effects of antimuscarinics on voiding function. The functional
urethral obstruction was induced according to a modification of the method of Nagabukuro
et al. [21]. In the present study, A-61603, a potent selective α1A-adrenoceptor agonist [22], instead of phenylephrine, was used in order to induce the urethral obstruction,
because A-61603 affected less on blood pressure than phenylephrine at the dose causing
the similar-level of Qmax decrease by phenylephrine in urethane anesthetized rat in
our preliminary experiments. In brief, at least 30-min after surgery, saline was infused
into the bladder at a rate of 0.06 mL/min in order to confirm the recovery of micturition
reflexes (priming). And then, A-61603 (0.03 μg/kg/min) was continuously infused into
the jugular vein at a rate of 0.01 mL/min. After several micturitions occurred, the
saline infusion was stopped and residual urine was removed from the bladder. 10 min
later, saline was reinfused into the bladder until micturition was evoked and then
the residual urine was removed again. The procedures were repeated at 10-min intervals
until 2 similar consecutive responses (cystometrogram and flowmetrogram patterns)
were obtained (pre-treatment value). 5 min after the last micturition, each antimuscarinic
drug was intravenously administered to the rat via the femoral vein. 5 min after drug
treatment, the same procedure as described in the measurement of pre-treatment value
was repeated twice. Changes in the second patterns each of cystometrogram and flowmetrogram
during this period was used for analysis of post-treatment value.
Bladder capacity measurement in urethane-anesthetized rats
Effects of antimuscarinics on BC were evaluated using a separate group of urethane-anesthetized
rats without partial urethral obstruction, because the preliminary examination confirmed
that A-61603 per se induced a 2.4-fold extension of intercontraction intervals in
urethane anesthesia rats, and because of this extension, BC-increasing effects of
the antimuscarinics failed to detect. The BC was measured by the method Yamazaki et
al. reported [13]. In brief, under urethane anesthesia (1.2 g/kg, s.c.), a lower abdominal midline
incision was made to expose the bladder. A saline-filled polyethylene catheter (PE50)
was inserted into the bladder dome. The catheter was connected via a 3-way cock to
a pressure transducer for measurement of the intravesical pressure and to a syringe
pumps for saline infusion. The intravesical pressure was recorded at a sampling rate
of 400 Hz using PowerLab system. At least 30-min after surgery, saline was infused
into the bladder at a rate of 0.06 mL/min in order to confirm the recovery of micturition
reflexes (priming). After several micturitions occurred, the saline infusion was stopped
and the residual urine was removed from the bladder. 10 min later, saline was reinfused
into the bladder until micturition was evoked and then the residual urine was removed
again. The procedures were repeated at 10-min intervals until 2 similar consecutive
responses were obtained (pre-treatment value). 5 min after the last micturition, each
antimuscarinic drug was intravenously administered to an individual rat via the femoral
vein. 5 min after drug treatment, the same procedure as described in the measurement
of pre-treatment value was repeated twice. Change in the second pattern of cystometrogram
was used for analysis of post-treatment value. The post-void RV was measured by gravimetric
method.
Drugs
Imidafenacin was synthesized by Kyorin Pharmaceutical Co., Ltd (Tokyo, Japan). Oxybutynin
chloride and A-61603 hydrate were purchased from Sigma-Aldrich Japan (Tokyo, Japan).
Imidafenacin was dissolved in saline with 1 mol/L hydrochloric acid, then neutralized
with 1 mol/L sodium hydroxide, and serially diluted with saline to desired concentrations.
Oxybutynin chloride and A-61603 hydrate were dissolved in saline and diluted with
saline to desired concentrations, when necessary. The dose levels of the drugs used
in this study were set up based on the results of preliminary studies.
Data analysis
All data are expressed as mean±S.E.M. In the pressure-flow study, the numerical variables
relating to the voiding, Qmax, maximum intravesical pressure (Pves max), and intravesical
pressure at Qmax (Pves Qmax) were derived from data obtained in pressure-flow measurement.
The influences of antimuscarinics on these parameters are expressed as a ratio of
post- to pre-treatment value. In BC measurement, BC, the variable relating to bladder
function during the storage phase, and maximum intravesical pressure (IVPmax), the
variable relating to the voiding, were derived from data obtained by cystometry. The
voiding efficiency (VE) was calculated from a formula of (BC-RV)/BC×100. The influences
of antimuscarinics on these parameters are expressed as a ratio of post- to pre-treatment
value. Influence index was calculated as the ratio of minimum effective dose in Qmax
to that in BC. Statistical analysis was performed by paired, 2-tailed Student’s t-test
for comparison between 2 groups or by one-way ANOVA followed by Dunnett’s post-hoc
test for multiple group comparison. In all comparisons, P<0.05 was considered statistically significant.
Results
Pressure-flow measurement in rats with partial urethral obstruction
To evaluate the inhibitory effects of antimuscarinics on the urinary flow rate, the
dose of A-61603 (0.03 μg/kg/min, i. v. infusion) that induces a modest level of functional
urethral obstruction was determined on the basis of the preliminary data of dose-response
relationship. This dose of A-61603 caused a significant partial reduction of Qmax
to approximately 24% (P<0.001), accompanied with a significant increase in Pves max and Pves Qmax to approximately
21 and 9%, respectively ([Fig. 1a, b]).
Fig. 1 A typical trace of intravesical pressure and urinary flow rate a, and changes in Qmax b between pre- and post-injection of A-61603 in urethane-anesthetized rats. A-61603
was infused continuously (0.03 μg/kg/min) into jugular vein. Each column and bar presents
the mean±standard error of the mean of 45 animals. *** P<0.001 vs. pre value (student’s paired t-test).
A61603; A-61603 hydrate.
In rats with the partial urethral obstruction, imidafenacin and oxybutynin significantly
decreased Qmax in a dose-dependent manner, with minimum influence doses of 0.03 (P<0.01 vs. pre-treatment value, approx. 37% inhibition) and 1 mg/kg (P<0.01 vs. pre-treatment value, approx. 38% inhibition), respectively ([Fig. 2a]). The maximum inhibition of Qmax induced by imidafenacin and oxybutynin were 49%
and 38%, and these values were not significantly different each other. In addition
to these, imidafenacin and oxybutynin dose-dependently decreased Pves max (up to 31%
and 33% inhibition, respectively) and Pves Qmax (up to 15% and 15% inhibition, respectively),
both in almost the same dose range as in decreasing the Qmax ([Fig. 2b,c]). These results suggest that both of the antimuscarinics reduce Qmax by inhibiting
detrusor contractions during voiding phase. Under the same experimental conditions,
vehicle treatment showed no influence on these values.
Fig. 2 Effects of imidafenacin and oxybutynin on maximum flow rate a, maximum intravesical pressure b, and intravesical pressure at Qmax c in rats with functional urethral obstruction. Vehicle, imidafenacin, and oxybutynin
were administered intravenously. Data are expressed as a ratio of post- to pre-treatment
value. Each column and bar represents the mean±standard error of the mean of 5 animals.
*P<0.05, **P<0.01, ***P<0.001 vs. vehicle group (Dunnett’s multiple comparison test).
Oxybutynin; oxybutynin chloride.
Bladder capacity measurement in rats
Imidafenacin and oxybutynin significantly caused a dose-dependent increase in BC,
with minimum influence doses of 0.01 (P<0.01 vs. vehicle group, 57% increase) and 3 (P<0.01 vs. vehicle group, 53% increase) mg/kg, respectively ([Fig. 3a]). The maximum increases of BC by imidafenacin and oxybutynin were 57% and 56%, these
values were not significantly different. Under the same experimental conditions, vehicle
treatment showed no influence on these values. Furthermore, both imidafenacin and
oxybutynin dose-dependently influenced voiding-related variables such as RV and VE.
Specifically, at the minimum influence dose to increase BC, oxybutynin tended to influence
on IVPmax (P=0.08, decrease in 25%) and resulted in significant influence on RV (P<0.01,
increase in 411%) and VE (P<0.001, decrease in 51%), whereas imidafenacin had no influence
on these parameters ([Fig. 3b,c,d]). Under the same experimental conditions, vehicle treatment showed no influence
on these values.
Fig. 3 Effects of imidafenacin and oxybutynin on bladder capacity a, maximum intravesical pressure b, residual urine volume c, and voiding efficiency d in urethane-anesthetized rats. Vehicle, imidafenacin and oxybutynin were administered
intravenously. Data are expressed as a ratio of post- to pre-treatment value. Each
column and bar represents the mean±standard error of the mean of 5 animals. *P<0.05, **P<0.01, ***P<0.001 vs. vehicle group (Dunnett’s multiple comparison test).
Oxybutynin; oxybutynin chloride.
The resultant relative influence index (i. e., minimum effective dose ratio of Qmax/BC)
of imidafenacin was 3 vs. 0.3 for oxybutynin, and thus10 fold higher than that of
oxybutynin.
Discussion
The present study revealed that imidafenacin has less influence on voiding function
than oxybutynin in rats. These results provide new information that antimuscarinics
may have varying degrees of impact on voiding function.
Indeed, many studies have evaluated the functional bladder selectivity of antimuscarinics
in various animals, mainly by focusing on dry mouth and constipation. In relation
to AUR, some studies have compared and reported the influences of antimuscarinics
on the IVP max (or RV) and on the effectiveness such as the BC increasing effect.
Yamazaki et al. [13] reported that imidafenacin increased BC without inhibiting IVP max or increasing
RV in urethane-anesthetized rats. Ney et al. [23] reported that oxybutynin increased BC with increasing RV in conscious rats. These
findings are in agreement with our present results. To our knowledge, however, there
have been no studies reported so far from viewpoint of uroflowmetric indices such
as Qmax, which reflects more directly the voiding function for comparative evaluation
of the effectiveness among antimuscarinics. Probably, the present study is the first
to investigate the risk of voiding dysfunction of antimuscarinics using such as uroflowmetric
index.
Imidafenacin and oxybutynin decreased Qmax dose-dependently ([Fig. 2a]). Moreover, imidafenacin did not affect Qmax at the minimum dose of increasing BC,
whereas oxybutynin affected Qmax at such a minimum dose ([Fig. 2a]). These results suggest that the relative risk of potentially inducing the voiding
dysfunction is lower in imidafenacin than in oxybutynin. One of the reasons for this
difference is that imidafenacin may act more selectively on bladder afferent pathway
than detrusor muscle, when compared with oxybutynin. Indeed, in urethane-anesthetized
rats of this study, imidafenacin did not affect IVPmax, RV, and VE at the minimum
dose of increasing BC, but oxybutynin affected RV and VE at the dose levels lower
than the minimum dose of increasing BC ([Fig. 3b,c,d]). Similar results have been reported using urethane-anesthetized rats and cerebral
infarction rats [11]
[13]
[24]. As one of putative mechanisms behind these, Yamada et al. [25] indicated that imidafenacin excreted in urine may contribute to its selective and
long-lasting distribution in the bladder, because imidafenacin showed a significant
binding to bladder muscarinic receptors in rats following intravesical injection at
concentration levels similar to its urine concentrations after oral administration
in rats as well as healthy volunteers. Mansfield [26] suggested that antimuscarinic drugs excreted in urine may have therapeutic potential.
The urinary excretion rate (percentage of doses) of oxybutynine in humans is less
than 5% [26]. On the other hand, the urinary excretion rate of imidafenacin in humans is relatively
high (7.8%) [27]. Because any antimuscarinic drug excreted in the urine may have direct access to
urothelial muscarinic receptors [28], antimuscarinic drugs excreted in the urine have the potential enough to act on
afferent nerve more easily than on detrusor muscle. Therefore, the above difference
in excretion rates between imidafenacin and oxybutynin may produce differences in
the relative influence index found in the present study. Another probable reason is
that oxybutynin has a stronger inhibitory effect on detrusor contractility than imidafenacin
because of the former’s Ca2+ channel antagonist actions [29].
Imidafenacin has a greater ratio of the minimum influence dose in decreasing Qmax
to the minimum influence dose in increasing BC than oxybutynin ([Table 1]). In some clinical studies, urinary retention has been reported in patients with
OAB during oxybutynin treatment [19]
[30]. On the other hand, no urinary retention has been reported in patients with OAB
during imidafenacin treatment [14]. Furthermore, combination therapy of imidafenacin with tamsulosin in BPH patients
induced no urinary retention in any of the patients in the ADDITION study, unlike
other antimuscarinics [18]. Considering these clinical reports, imidafenacin may have less influence on voiding
function than oxybutynin. But since these clinical studies were performed under limited
conditions, such as defined study period and selected patients with a certain baseline
severity, it is still unclear whether imidafenacin actually has less risk of inducing
AUR compared with other antimuscarinics. The present study with rats, however, shows
that antimuscarinics may have varying degrees of impact on voiding difficulty. The
present authors, therefore, are very interested in whether imidafenacin actually has
little influence on risk of AUR in BPH patients during its long term administration
or in those with a severe condition. In addition, this study suggests that new generation
antimuscarinic imidafenacin is considered more beneficial than first-generation antimuscarinic
oxybutynin, and thus the authors are also interested in whether new generation of
antimuscarinics are different in their clinical benefit. As the first step, it is
intriguing challenges to compare influences on voiding function among the new generation
of antimuscarinics, and furthermore, to investigate whether the receptor selectivity
is related to the difference in the influence on voiding function in animal models
of BPH.
Table 1 The minimum doses of decreasing the maximum urinary flow rate (Qmax) and increasing
the bladder capacity (BC).
|
Drug
|
Minimum effective dose
|
Ratio (C)
|
|
Qmax (A)
|
BC (B)
|
|
|
Imidafenacin
|
0.03
|
0.01
|
3
|
|
Oxybutynin
|
1
|
3
|
0.3
|
The dose ratio (C) was calculated by dividing the minimum dose (A) of decreasing Qmax
by that (B) of increasing BC
Oxybutynin; oxybutynin chloride
In conclusion, this study demonstrated that imidafenacin has a lower relative risk
of voiding difficulty than oxybutynin in comparing the relative effects of these 2
antimuscarinics on the Qmax obtained from rats with functional urethral obstruction
and on the bladder capacity obtained from the urethane-anesthetized rats. These results
provide new information that antimuscarinics may have varying degrees of impact on
voiding difficulty. Thus, more comparative studies will be needed to further clarify
such differences in degrees of impact on the voiding phase among more antimuscarinic
drugs.
