We have reported on the design, synthesis, and biological characterization of (R)-4-[3,4-dioxo-2-(1,2,2-trimethyl-propylamino)-cyclobut-1-enylamino]-3-ethyl-benzonitrile (1), a novel, potent, and selective adenosine 5'-triphosphate-sensitive potassium (K(ATP)) channel opener with potential utility for the treatment of urge urinary incontinence (UUI). Excising the aniline-derived nitrogen atom of 1 or replacing it with an aralkyl group, led to bladder smooth muscle relaxant chemotypes 3 and 4, respectively. Prototype compounds in these series were found to produce significant increases in an iberiotoxin (IbTx)-sensitive hyperpolarizing current, thus suggesting that these relatively modest structural modifications resulted in a switch in the mechanism of action of these smooth muscle relaxants from K(ATP) channel openers to activators of the large-conductance Ca2+-activated potassium channel (BK(Ca)). We report herein the syntheses and biological evaluation of a series of substituted 3-amino-4-aryl-(and aralkyl-)cyclobut-3-ene-1,2-diones.

A series of benzofuroindoles and indenoindoles were synthesized using a, microwave-assisted Fischer-indole cyclization. Improvement in yield and purity was observed. These compounds were shown to possess biological activity.

Compounds in a structurally novel series of substituted 10H-benzo[4,5]furo[3,2-b]indole-1-carboxylic acids and related 5,10-dihydro-indeno[1,2-b]indole-1-carboxylic acids were prepared and shown to possess potent, bladder-selective smooth muscle relaxant properties and thus are potentially useful for the treatment of urge urinary incontinence. Electrophysiological studies using rat detrusor myocytes have demonstrated that prototype compound 7 produces a significant increase in hyperpolarizing current, which is iberiotoxin (IbTx)-reversed, thus consistent with activation of the large-conductance Ca(2+)-activated potassium channel (BK(Ca)).

A novel series of benzylamine, potassium channel openers (KCOs) is presented as part of our program toward designing new, bladder-selective compounds for the treatment of urge urinary incontinence (UUI). We have found that the in vitro potency of (R)-4-[3,4-dioxo-2-(1,2, 2-trimethyl-propylamino)-cyclobut-1-enylamino]-3-ethyl-benzo nitrile 1 in the relaxation of precontracted rat detrusor strips can also be obtained with cyanobenzylamine derivative 4 (IC(50) = 0.29 microM) (Figure 3). Addition of a 2-Cl substituted benzylamine moiety and changing the alkylamino substituent of 4 to a t-Bu amine gives 31 (IC(50) = 0.14 microM)-a compound with similar in vitro potency as 4 as well as relaxant activity on bladder smooth muscle in vivo when administered orally (31, ED(50) = 3 mg/kg) in a rodent model of bladder instability. Further modifications, particularly the replacement of the t-Bu amino substituent with a tert-amylamine, gave a similarly active compound 60 (IC(50) = 0.10 microM) which shows excellent in vivo efficacy (ED(50) = 0.6 mg/kg). Moreover, 60, 3-(2,4-dichloro-6-methyl-benzylamino)-4-(1, 1-dimethyl-propylamino)-cyclobut-3-ene-1,2-dione (WAY-151616), shows excellent tissue selectivity for bladder K channels over arterial tissue (60, MAP ED(20) = 100 mg/kg; selectivity: MAP ED(20)/bladder ED(50) = 166). Other manipulations of the benzylamino cyclobutenediones, acylation of the benzylamine, conversion of the benzylamine substituent to a benzamide, homologation of the benzylamine to a phenethylamine, and incorporation of a methyl group at the benzyl carbon, all led to substantial loss of in vitro activity, although some in vivo activity was maintained in the acylated analogues. Compound 60 represents an attractive candidate for development in the treatment of UUI.

A structurally novel series of adenosine 5'-triphosphate-sensitive potassium (K(ATP)) channel openers is described. As part of our efforts directed toward identifying novel, bladder-selective potassium channel openers (KCOs) targeted for urge urinary incontinence (UUI), we found that bioisosteric replacement of the N-cyanoguanidine moiety of pinacidil (1, Figure 1) with a diaminocyclobutenedione template afforded squaric acid analogue 2, the prototype of a novel series of K(ATP) channel openers with unique selectivity for bladder smooth muscle in vivo. Further modification of the heterocyclic ring to give substituted aryl derivatives (3) afforded potent KCOs that possessed the desired detrusor selectivity when administered orally. The effects of these potassium channel agonists on bladder contractile function was studied in vitro using isolated rat detrusor strips. Potent relaxants were evaluated in vivo in a rat model of bladder instability. Lead compounds were evaluated concomitantly in normotensive rats for their effects on mean arterial blood pressure (MAP) and heart rate as a measure of in vivo bladder selectivity. (R)-4-[3,4-Dioxo-2-(1,2, 2-trimethyl-propylamino)-cyclobut-1-enylamino]-3-ethyl-benzo nitrile (79) met our potency and selectivity criteria and represents an attractive development candidate for the treatment of UUI. Electrophysiological studies using isolated rat bladder detrusor myocytes have demonstrated that compound 79 produces significant hyperpolarization which is glyburide-reversed, thus consistent with the activation of K(ATP). The design, synthesis, structure-activity relationships (SAR), and pharmacological activity associated with this series of novel KCOs will be discussed.

The effects of the ATP-dependent potassium channel agonists ZD6169, celikalim, and WAY-133537 on bladder contractile function were examined in vitro on isolated bladder strips and in vivo on spontaneous bladder contractions. All three compounds produced a concentration-dependent relaxation of isolated rat detrusor strips (IC50 values = 0.93, 0.03, and 0.09 microM, respectively for ZD6169, celikalim, and WAY-133537. Contractile inhibition by all three compounds was fully reversed by 6 microM glyburide. These compounds also effectively inhibited spontaneous bladder contractions in the rat hypertrophied bladder model of detrusor instability. We also examined the electrophysiological properties of WAY-133537 on isolated rat bladder detrusor myocytes. Myocytes had an average resting membrane potential of -40 mV. Under patch current-clamp conditions, WAY-133537 (0.3 and 1.0 microM, n = 4-5) produced a significant hyperpolarization of 21 and 26 mV, respectively. Hyperpolarization was reversed by the addition of 5 microM glyburide. In patch voltage-clamp studies, WAY-133537 (0.3 microM, n = 3) significantly increased outward current in response to both voltage step and ramp protocols consistent with activation of the ATP-dependent potassium channel. In the detrusor instability model, WAY-133537 and celikalim had similar oral potencies (ED50 = 0.13 and 0.3 mg/kg, respectively), whereas ZD6169 was less potent (ED50 = 2.4 mg/kg). The antihypertensive agent celikalim exerted effects on the bladder at doses that significantly reduced systemic blood pressure. In contrast, both WAY-133537 and ZD6169 inhibited bladder hyperactivity at doses that produced minimal changes in both mean arterial blood pressure and heart rate. These data suggest that both WAY-133537 and ZD6169 may be useful in the treatment of bladder instability at doses associated with minimal hemodynamic side effects.