Abanoquil, a New Alpha-1 Adrenoceptor Antagonist. in Vitro and in Vivo Effect on Erectile Tissue

Abanoquil, a new alpha-1 adrenoceptor antagonist. In vitro and in vivo effect on erectile tissue

A Giraldi1*, M Wyllie2 and G Wagner1

1Department of Medical Physiology, University of Copenhagen, Rigshospitalet, 7121, Blegdamsvej 9, DK-2100, Copenhagen, Denmark; 2Urodoc, Kent, UK

Using an organ bath model with porcine cavernosal tissue strips and an in vivo monkey model we demonstrated that abanoquil, a novel alpha adrenoceptor antagonist, is able to relax contracted tissue strips and induce erectile response when injected intracorporally. The erectile response in the monkeys was not dose-related and compared to the effect of papaverine injections, abanoquil induced a lower level of tumescence and rigidity. Hence, abanoquil might be useful as a facilitator of erection in the pharmacological treatment of erectile dysfunction. International Journal of Impotence Research (2000) 12, Suppl 1, S37±S40

Keywords: abanoquil; alpha-1 adrenoceptor antagonist; erectile dysfunction

Introduction on corpus cavernosum smooth muscle in vitro and in vivo. It is generally accepted that ¯accidity of the penis is a consequence of contraction of the smooth muscle Material and methods cells of corpus cavernosum (cc) and the penile arteries, caused in part, by sympathetic nervous activity stimulating alpha adrenoceptors.1 The dom- In vitro studies inating alpha adrenoceptor in cc is alpha-1 subtype, Porcine corpus cavernosum was obtained from boars whereas it is alpha-2 in the cavernosal arteries.2±4 immediately after slaughtering. Tissue strips Relaxation of the trabecular smooth muscle cells of cc (~3 6 3 6 8 mm) were prepared and mounted be- is crucial to develop an erection.5,6 In the ongoing tween two L-shaped metal hooks immersed in search for agents that have the capacity of relaxing cc standard model organ baths for recording of the smooth muscle cells and thereby have a possible role isometric tension. All tissue strips that were used for in the pharmacological treatment of erectile dysfunc- experiments showed spontaneous activity and re- tion, multiple, in vivo and in vitro studies on cc sponded to addition of norepinephrine (NE) by smooth muscle have been carried out. These studies contraction. have demonstrated relaxatory response of cc smooth muscle to naturally occuring compounds such as prostaglandin,7 vasoactive intestinal polypeptide8 Pharmacological experiments and pharmacological agents such as papaverine,9 phosphodiesterase inhibitors,10 calcium channel blocking compounds,11 potassium channel open- The effect of abanoquil (UK-52,046 from P®zer, UK) ers12,13 and alpha adrenoceptor blocking agents.14,15 on spontaneous contractions was studied by addi- Abanoquil, UK-52,046 (4-amino-6,7-dimethoxy- tion of increasing concentrations to the organ bath 8 5 2(1,2,3,4-tetrahydro-6,7-dimethoxyisoquinol-2- (10 ±10 M, n 11). Dose±response curves to NE yl) quinoline methanesulphonate) (Figure 1) is a were constructed by cumulative addition of NE 7 4 novel alpha adrenoceptor antagonist with a high (10 ±10 M) to the organ bath followed by a selectivity to the alpha-1 subtype.16 The aim of the complete washout when steady-state was achieved present study was to evaluate the effect of abanoquil after addition of the highest concentration. There- after, abanoquil (10 10±10 7 M) was added to the organ baths and after 30 min incubation new NE, dose±response curves were constructed. *Correspondence: Dr A Giraldi, Department of Medical Physiology, University of Copenhagen, Rigshospitalet, To study the effect of abanoquil on precontracted Dept. 7121, Blegdamsvej 9, DK-2100, Copenhagen, Denmark. tissue-strips, the strips were precontracted with NE E-mail: [email protected] (10 6±10 4 M). When maximal contraction was Effect of abanoquil on erectile tissue AGiraldiet al S38 monkeys had in previous experiments responded to intracavernosal (ic) injections of papaverine by increasing tumescence and rigidity of the penis. The monkeys were anaesthetized with intramus- cular apozepam (0.1 mg=kg) and ketamine-chloride (20 mg=kg) and placed in a supine position with the penis stretched out. A rubber band was placed around the root of the penis as a tourniquet. The agents of use were dissolved in 0.3 ml saline and injected into one of the copora cavernosa using a 27 G needle; the tourniquet was kept in place for 3 min after the injection. Five, 10, 15, 20, 30, 60 and 120 min after the injection, tumescence (increase in volume) and rigidity of the penis were estimated visually and by palpation by the same experienced Figure 1 Chemical structure of abanoquil UK52,046. investigator. The estimations were graded on a ®ve point scale (0-4), 0 representing no change in tumescence=rigidity and 4 full tumescence=rigidity. Before and during the experiments the brachial arterial systolic pressure and pulse were measured using a neonatal cuff and an 8 MHz Doppler to localize the artery.

Pharmacological experiments

In order to investigate the effect of abanoquil on corporal smooth muscle the monkeys were injected with 0.7 mg(n 12), 1.8 mg(n 8) or 3.5 mg(n 11) abanoquil. Six monkeys were injected with papaverine (9 mg, SAD, DK) in order to compare the effect of papaverine to the effect of abanoquil. A Figure 2 Concentration response curve showing norepine- further six monkeys served as controls and were phrine-induced contraction of porcine corpus cavernosum after injected with saline 0.3 ml. In order to elucidate the 10 7 incubation with abanoquil (10 ±10 M). Calculated as percent receptor speci®city and reversibility of the effect of contraction of NE-induced contraction before abanoquil incubation. Control tissues were not incubated with abanoquil. abanoquil, seven monkeys were injected with 10 mg abanoquil and tumescence and rigidity were evaluated. Thereafter, the monkeys were injected with obtained, abanoquil was added to the organ bath in NE (10 mg, n 3) or phenylephrine (PE, 10 mg, n 4, increasing concentrations (10 9±10 7 M). Sigma, USA). Ensuing tumescence and rigidity were evaluated 5 and 10 min after the NE=PE injection. Using the same procedure, six monkeys were Statistical analysis injected with 25 mg abanoquil followed by 50 mg clonidine. The in vitro experiments were analysed using the paired Student's t-test, P 0.05 was considered as Statistical analysis signi®cant.

The data in each series of experiments were In vivo monkey studies analysed using the non-paired-parametric Fried- mann test and the Page test for trend. To analyse differences between the maximal obtained tumes- In accordance with local ethical guidelines, each cence and rigidity with each agent the non-paired, monkey participated in only one experiment a day non-parametrical Kruskal-Wallis test was used. In with an interval of at least two weeks between each order to compare the values obtained with abanoquil experiment. to those obtained with papaverine, the Wilcoxon- In these series of experiments, 12 male monkeys Pratt test was used. All these statistical calculations were used (Maccaca Fascicularis, weight 4±6 kg). All were performed with the MEDSTAT program.17

International Journal of Impotence Research Effect of abanoquil on erectile tissue A Giraldi et al S39 Blood pressure and pulse were expressed as mean time showed no statistically signi®cant difference Æ SE and analysed using the paired Student's t-test. between the responses induced by abanoquil and P < 0.05 was considered as signi®cant. papaverine using the Wilcoxon-Pratt test (P 0.063). If the responses were compared through the whole time period (0±120 min) signi®cant higher Results erectile responses (both tumescence and rigidity) were observed after papaverine injections compared to abanoquil. Saline injections had no effect on In vitro experiments penile tumescence and rigidity. Injection of NE or PE subsequent to abanoquil injection resulted in a signi®cant decrease in tumescence and rigidity Abanoquil, in concentrations ranging from 10 7 to during the following 10 min after the injection. In 10 4 M, did not affect the spontaneous activity of contrast, clonidine did not reverse the effect of the smooth muscle tissue strips. Addition of abanoquil, since no signi®cant decrease in tumes- abanoquil (10 10±10 7 M) prior to NE-induced cence and rigidity was observed after the injection. contraction inhibited the NE-induced contraction in a dose dependent manner as illustrated by the rightward shift of the NE-dose±response curve shown in Figure 2. Abanoquil 10 10 M had no Discussion signi®cant effect compared to control tissue, whilst abanoquil 10 7 M almost completely inhibited contraction of the tissue strips. Addition of abanoquil to In the present study it was shown that abanoquil NE-precontracted porcine tissue strips resulted in a induced an erectile response when injected intra- dose-related relaxation of the tissue with a minimal corporally in monkeys. Abanoquil induced relaxa- relaxation using abanoquil 10 9 M and an almost tion of pre-contracted porcine corporal smooth complete relaxation when the dose of abanoquil is muscle strips in vitro, but had no effect on the increased to 10 7 M (data not shown). spontaneous activity. The penile response to intracavernosal injection achieved in monkeys was characterized by a greater effect on tumescence than In vivo experiments on rigidity. Most of the monkeys developed an erection after abanoquil injection, however, most of the erections were scored lower than the attainable Injections of 0.7, 1.8 and 3.5 mg abanoquil resulted in maximum (4=4). When compared to the effect of a signi®cant change in tumescence and rigidity of intracavernosally administered papaverine a signi®- the penis in the monkeys (P < 0.001 for all three cantly lower attainable erectile response was ob- concentrations). Judged by the rank sums (Fried- served with abanoquil, even though there was no mann test) the responses for all three concentrations difference when comparing the attained level at peaked 15±20 min after the intracavernosal injec- peaking time, probably due to the small numbers of tions. The responses then levelled off, but were still monkeys used. present in 27 of the 31 trials after 120 min. On this The achieved erectile responses were not dose- basis we applied the Page-test for trend on the related. This may be attributed to the high doses period of 0±20 min and 20±120 min. This showed utilized or because the responses were induced by that a steady increase in tumescence and rigidity a non-speci®c reaction caused by abanoquil. The occurred in all three groups from 0 to 20 min after latter does not seem to be the case, since the alpha-1 the injection (P < 0.001), whereas there were no agonists NE and PE were able to reverse the effect of signi®cant changes from 20 to 120 min. The high concentrations of abanoquil in vivo and the monkeys were controlled 4 h after the injections alpha-2 agonist, clonidine, had no effect on the and none of them developed long lasting erections. response. These ®ndings indicate that abanoquil Furthermore there was no signi®cant difference in primarily exerts its effect by alpha-1 antagonism. the obtained maximum values for tumescence or This is in agreement with the ®ndings that alpha-1 rigidity when comparing the effect of three different receptors are the predominant alpha-adrenoceptor abanoquil concentrations. Arterial blood pressure type in the corpus cavernosum. and pulse were unaffected during the experiments. In the search for an erectogenic agent, it is crucial Papaverine injections resulted in a signi®cant that the agent does not cause priapism. In the increase in both penile tumescence and rigidity present in vivo studies, no long lasting erections from the time of intracavernosal injection to time were observed in the monkeys. It should, however, of peak erectile response (15 min) whereafter the be emphasized that all studies were done under response levelled off with a signi®cant decrease in anaesthesia and the number of experiments were tumescense but not in rigidity after 120 min. Four limited. Previous studies of alpha-adrenoceptor h after the experiment, no lasting erections were antagonists have shown that not all compounds observed. Comparison of the attained values at peak are useful in the treatment of erectile dysfunction.

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