New strategies for inhibition of non‐adrenergic prostate smooth muscle contraction by pharmacologic intervention

Abstract

Background Inhibition of prostate smooth muscle contraction by α1‐adrenoceptor antagonists (α1‐blockers) is a first‐line medical treatment of lower urinary tract symptoms suggestive of benign prostatic hyperplasia. Increased smooth muscle tone in the hyperplastic prostate may drive urethral obstruction, resulting in bladder outlet obstruction and voiding symptoms. However, efficacy of α1‐blockers is limited, as non‐adrenergic mediators including endothelin‐1 and thromboxane A2 (TXA2) increase prostate smooth muscle tension in parallel to α1‐adrenoceptors. This may maintain urethral obstruction despite therapy with α1‐blockers. Consequently, future treatment options with higher efficacy need to target α1‐adrenergic and non‐adrenergic contractions simultaneouly. Recently, several compounds were reported to inhibit adrenergic or neurogenic prostate contractions, however, their effects on non‐adrenergic contraction are unknown. Here, we examined effects of inhibitors for Rac‐GTPase, Src family kinases (SFKs), and p21‐activated kinases (PAKs) on non‐adrenergic prostate contractions. Methods Prostate tissues were obtained from radical prostatectomy. Contractions were studied in an organ bath. Viability of cultured stromal cells was assessed by CCK‐8 assay. Results Inhibition of α1‐adrenergic contractions by Rac inhibitors EHT1864 (100 μM) and NSC23766 (100 μM), and SFK inhibitors AZM475721 (10 μM) and PP2 (10 μM) was confirmed by inhibition of methoxamine‐induced contractions. No effects of the PAK inhibitors FRAX486 (30 μM) and IPA3 (300 μM) on α1‐adrenergic contraction were confirmed by absent effects on methoxamine‐inuced contractions. EHT1864 caused inhibition of endothelin‐1‐ and U46619‐induced contractions. EHT1864 reduced the viability of stromal cells concentration‐ and time‐dependently. EHT1864 attenuated KCl‐induced contractions of prostate strips only slightly, so that toxic effects may not account alone for inhibition of agonist‐induced contractions. NSC23766 inhibited U46619‐induced contractions, but not endothelin‐1‐induced contractions. AZM475271 had no effects on endothelin‐1‐ or U46619‐induced contractions, while PP2 inhibited U46619‐ but not endothelin‐1‐induced contractions. FRAX486 caused inhibition of U46619‐induced contractions. IPA3 inhibited U46619‐, but not endothelin‐1‐induced contractions. Conclusions Of all six inhibitors, EHT1864 seems to be most promising from a translational point of view, as it inhibited TXA2‐ and endothelin‐1‐induced besides α1‐adrenergic prostate contractions. This reflects divergent pharmacologic profiles of EHT1864 and NSC23766, although both are Rac‐GTPase inhibitors. In vivo, urodynamic effects of EHT1864 and possibly of FRAX486 may exceed those of α1‐blockers.

Inhibition of Female and Male Human Detrusor Smooth Muscle Contraction by the Rac Inhibitors EHT1864 and NSC23766

“…While the role of RhoA for promotion of smooth muscle contraction is well established for all smooth muscle tissues (Somlyo and Somlyo, 2000;Somlyo and Somlyo, 2003;Christ and Andersson, 2007;Puetz et al, 2009), a similar role for RacGTPases has been proposed more recently. NSC23766 and EHT1864 inhibited contractions of human prostate tissues, so that a procontractile role of Rac1 has been proposed for prostate smooth muscle (Wang et al, 2015;Yu et al, 2019). These previous and our present findings for the detrusor and prostate are in line with other studies, where Rac inhibitors inhibited cholinergic, serotonin-, and endothelin-induced contractions of airway tissues from humans, mice and rats, as well as adrenergic and serotonin-induced contractions of rodent vessels, and contractions of the mouse ileum (Staiculescu et al, 2013;Rahman et al, 2014;Shibata et al, 2015;Andre-Gregoire et al, 2017;Harvey et al, 2017;Sakai et al, 2018).…”

“…For the prostate, cytotoxic effects of NSC23766 and EHT1864 were recently excluded to account for contraction inhibition in organ bath experiments, as these effects did not occur quickly enough (Yu et al, 2019). As very similar conditions were applied here to bladder tissues, it appears unlikely, that cytotoxic effects underly the inhibitory effect on detrusor contraction in our study.…”

“…NSC23766 and EHT1864 were here applied in concentrations of 100 µM. This concentration was used, as it inhibited smooth muscle contractions of human prostate tissues in previous studies (Wang et al, 2015;Yu et al, 2019). This was paralleled by inhibition of Rac1, but not of the closely related GTPase RhoA (Wang et al, 2015).…”

“…In addition to their involvement in actin organization and cell cycle progression, a possible Rac-dependent control of smooth muscle contractions has been repeatedly suggested in recent years. Thus, contractions of human prostate tissues can be inhibited by inhibitors for RacGTPases (Wang et al, 2015;Yu et al, 2019). Other studies suggested a role of RacGTPases in smooth muscle contraction of airways, vessels, ileum, and urinary bladder in mice (Rahman et al, 2014;Andre-Gregoire et al, 2017).…”

Targeting Rac and Cdc42 GEFs in Metastatic Cancer

“…Recent studies have also demonstrated the potential of NSC23766 in combination therapy to overcome resistance to targeted as well as cytotoxic therapies (Karpel-Massler et al, 2017;Tian et al, 2018). Nonetheless, although the use of NSC23766 may look promising in some instances, the high effective concentrations (µM range) and the reported off-target effects in platelets, on alfa-1-adrenergic receptors and CXCR4 chemokine receptors, limits its therapeutic use (Dutting et al, 2015;Mills et al, 2018;Yu et al, 2019).…”