Reported herein are a series of reverse indoles that represent novel non-steroidal mineralocorticoid receptor (MR) antagonists. The key structure-activity relationships (SAR) are presented below. This reverse indole series is exemplified by a compound that demonstrated efficacy in an acute natriuresis rodent model comparable to marketed MR antagonists, spironolactone and eplerenone.

The renal outer medullary potassium channel (ROMK, KCNJ1) mediates potassium recycling and facilitates sodium reabsorption through the Na(+)/K(+)/2Cl(-) cotransporter in the loop of Henle and potassium secretion at the cortical collecting duct. Human genetic studies indicate that ROMK homozygous loss-of-function mutations cause type II Bartter syndrome, featuring polyuria, renal salt wasting, and hypotension; humans heterozygous for ROMK mutations identified in the Framingham Heart Study have reduced blood pressure. ROMK null mice recapitulate many of the features of type II Bartter syndrome. We have generated an ROMK knockout rat model in Dahl salt-sensitive background by using zinc finger nuclease technology and investigated the effects of knocking out ROMK on systemic and renal hemodynamics and kidney histology in the Dahl salt-sensitive rats. The ROMK(-/-) pups recapitulated features identified in the ROMK null mice. The ROMK(+/-) rats, when challenged with a 4% salt diet, exhibited a reduced blood pressure compared with their ROMK(+/+) littermates. More importantly, when challenged with an 8% salt diet, the Dahl salt-sensitive rats with 50% less ROMK expression showed increased protection from salt-induced blood pressure elevation and signs of protection from renal injury. Our findings in ROMK knockout Dahl salt-sensitive rats, together with the previous reports in humans and mice, underscore a critical role of ROMK in blood pressure regulation.

We investigated the effects of chronic mineralocorticoid receptor blockade with eplerenone on the development and progression of hypertension and end organ damage in Dahl salt-sensitive rats. Eplerenone significantly attenuated the progressive rise in systolic blood pressure (SBP) (204 ± 3 vs. 179±3 mmHg, p < 0.05), reduced proteinuria (605.5 ± 29.6 vs. 479.7 ± 26.1 mg/24h, p < 0.05), improved injury scores of glomeruli, tubules, renal interstitium, and vasculature in Dahl salt-sensitive rats fed a high-salt diet. These results demonstrate that mineralocorticoid receptor antagonism provides target organ protection and attenuates the development of elevated blood pressure (BP) in a model of salt-sensitive hypertension.

Aldosterone (Aldo) is a steroid hormone that promotes Na reabsorption by binding to the mineralocorticoid receptor (MR). MR antagonists, spironolactone (SPL) and eplerenone (EPL) are used for the treatment of hypertension and heart failure. One measure of anti‐MR activity is urinary Na/K ratio derived from adrenalectomized rats infused with Aldo. Our goal was to develop an improved approach to measure anti‐MR activity in rats with intact adrenal glands. Plasma Aldo levels and anti‐MR activity of SPL were determined at different times of the day in WKY rats. Circulating Aldo levels and urinary Na, K excretion showed diurnal dependence. During the 24 hr cycle (light period 6AM: 6PM), plasma Aldo was significantly higher in the active phase than in the inactive phase. Na excretion was significantly lower during the period of 4PM:10PM than from 9AM:3PM. In contrast, kaliuresis was inversely correlated with natriuresis. The anti‐MR activity of SPL within 6hrs post dose was much greater when dosed prior to the active phase than when dosed during the early inactive phase. Results from 4PM dosing showed that natriuresis continuously increased up to 14hrs but declined during 15–17hrs post dose. These findings demonstrate that the anti‐MR activity of SPL varies with its time of administration. In summary, the use of a chronopharmacological approach proved effective in defining an improved natriuresis model of MR antagonism.