Cardiovascular disease (CVD) is considered a global health concern, and its prevalence is increasing, especially in older people [1]. Among the different comorbidities of CVD, heart failure (HF) is a complex clinical condition that manifests as abnormal cardiac structure or function, thereby altering normal cardiac filling and the ejection of an appropriate blood volume to maintain peripheral organ perfusion. HF is defined according to the clinical measure of left ventricular (LV) ejection fraction (EF), which may either be reduced (<40%; HFrEF or systolic HF) or preserved (>50%; HFpEF or diastolic HF) ^[2][3][2,3]. Despite the enormous therapeutic advances that have emerged in the management of CVD in the past three decades, patients diagnosed with HF still have a very poor prognosis and quality of life. It is also the most common reason for hospitalization in the aged population [4]. Traditional guideline-directed therapies primarily slow the progression of cardiovascular disorders, but there is a need to develop novel, preventive, and reparative therapeutic strategies.

2. Chemical Properties of Steroidal and Non-Steroidal MRAs

The steroidal MRA, eplerenone, is around 40-fold less potent than spironolactone toward the MR; however, it exhibits much greater selectivity than spironolactone and shows less steroidal hormone-related adverse effects ^[5][6][12,15]. Among the recently developed non-steroidal MRAs, the cardiovascular endpoint outcomes of finerenone (BAY94-8862, Bayer) and esaxerenone (CS-3150, Daiichi Sankyo) have been published in both pre-clinical and clinical studies. In terms of tissue distribution, steroidal MRAs have been suggested to likely accumulate in the kidney, which alters electrolyte homeostasis, leading to hyperkalemia ^[5][7][8][12,17,20]. In contrast, the non-steroidal MRAs finerenone and esaxerenone have a balanced distribution between the kidney and heart ^[9][10][21,22]. On the basis of the available information, non-steroidal MRAs appear to be potent and selective, and their tissue distribution and affinity are rather balanced in both the heart and kidney. The pharmacological actions of different MRAs cause distinct regulation of MR target gene promoter activity based on ligand-specific gene regulation. The distinct clinical actions of different MRAs can be explained, at least in part, by the different binding modes to the MR and the initiation of MR modulation through different molecular mechanisms. Structurally different MRAs have different MR binding modes, resulting in distinct gene expression and ligand-specific clinical actions. The molecular mechanism of steroidal and non-steroidal MRAs is quite different, even though all of the MRAs bind to the same MR ligand-binding domain (MR-LBD). Spironolactone and eplerenone are both passive MRAs that are derivatives of aldosterone. However, spironolactone promotes co-factor (SRC-1) recruitment to an MR-dependent promoter, but to a much lesser extent than aldosterone ^[11][12][13,14]. Conversely, the non-steroidal MRA finerenone is a complete antagonist, unlike spironolactone and eplerenone, which binds to the ligand-binding domain of the MR, causing a conformational change inside the MR complex and thereby altering the stability, as well as causing nuclear translocation, of the MR ^[13][23]. Esaxerenone is an enantiomer of atropisomer that binds to the MR-LBD with large side chains, which are flipped and form a relatively large and secluded binding pocket ^[14][24]. Thus, the binding mode of non-steroidal MRAs is completely different to that of classic steroidal MRAs.

3. Pharmacological Effects of Steroidal and Non-Steroidal MRAs in CVD: Evidence from Pre-Clinical Studies

3.1. Blood Pressure

Hypertension is considered a predominant risk factor for the development and progression of CVD ^[15][25]. Steroidal MRAs have been proven to be effective in reducing blood pressure (BP), either alone or as add-on therapy. Similarly, non-steroidal MRAs have antihypertensive properties. 

Among the experimental animal models of CVD, hypertension-induced HF has been repeatedly studied in Dahl salt-sensitive (DSS) hypertensive rats with high salt (HS) loading. The steroidal MRAs eplerenone ^[16][17][19,26] and spironolactone ^[16][19] reduced the BP by around 8–10% compared with their counterparts in 8% salt diet-fed DSS rats at higher doses (100 mg/kg/day), but not at lower doses (10 and 30 mg/kg). Notably, the non-steroidal MRA esaxerenone caused a reduction in systolic BP (SBP) by approximately 18% and 27% at a dose of 1 and 2 mg/kg/day, respectively, in the same model ^[16](Table 1) [19]. However, HS loading for 6 weeks prior to esaxerenone treatment (1 mg/kg/day) for 4 weeks reduced the BP by around 9% ^[18][27]. The deoxycorticosterone acetate (DOCA)-salt model is frequently considered as a model of cardiovascular remodeling due to the persistently high BP. In this model in uninephrectomized (UNX) Wistar Kyoto rats, esaxerenone treatment (3 mg/kg) for 2 weeks significantly reduced the BP (30%), while eplerenone (30 mg/kg) and spironolactone (30 mg/kg) were not effective in reducing BP (3% decrease and 2% increase, respectively) ^[19][18].

3.2. Myocardial Structural Remodeling

During CVD progression, complex alterations in myocardial structure are commonly associated with cardiomyocyte apoptosis and interstitial and perivascular collagen fiber accumulation ^[20][38]. MRAs have consistently shown beneficial effects on pathological myocardial remodeling ^[21][39]. Here, evidence from pre-clinical studies is discussed to compare the beneficial effects of steroidal and non-steroidal MRAs on myocardial remodeling. In HS-loaded DSS rats, LV weight and LV end-diastolic diameter were significantly reduced, and LV end-systolic elastance was increased, in response to eplerenone, while BP was not altered (10 mg/kg) ^[17][26]. Spironolactone reduced LV weight at a higher dose (30 mg/kg) ^[16][19]. In contrast, a significant reduction in LV weight and LV internal diameter was evident with esaxerenone at doses as low as 0.5–1 mg/kg ^[16][19]. In the same study, the plasma concentration of brain natriuretic peptide (BNP), which is a well-accepted biomarker of cardiac remodeling, was not significantly reduced, even with a high dose of eplerenone (100 mg/kg). However, it was significantly reduced by esaxerenone at a dose as low as 1 mg/kg in salt-loaded DSS rats ^[16][19]. In the DOCA/salt model, finerenone caused a significant reduction in heart weight by 1 mg/kg, but structural heart injury was reduced by 10 mg/kg ^[9][21]. 

3.3. Myocardial Function

Kobayashi et al. ^[17][26] demonstrated that LV percent fractional shortening (%FS), which is an index of systolic function, was significantly reduced with HS loading for 12 weeks in DSS rats, while treatment with eplerenone attenuated these changes independent of changes in BP. In the same animal model, HS loading for 10 weeks caused systolic dysfunction with an increase in LV end-systolic diameter (LVIDs) and a reduction in EF, %FS, stroke volume (SV), and cardiac output (CO) ^[18][27]. Esaxerenone treatment (1 mg/kg) significantly improved systolic dysfunction with a 13% reduction in LVIDs and a significant increase in EF (16%), %FS (25%), SV (21%), and CO (26%) ^[18][27]. These data indicate that the non-steroidal MRA esaxerenone has cardioprotective effects in salt-sensitive hypertensive animals.

The ischemia-driven chronic myocardial infarction model induces both systolic and diastolic dysfunction. An alteration in myocardial contractility and relaxation was evidenced by a decrease in dp/dtmax and a deterioration in dp/dtmin following 8 weeks of myocardial infarction in rats. These characteristic parameters of LV systolic function in HF were significantly ameliorated by finerenone (1 mg/kg) but not eplerenone (100 mg/kg). In contrast, LV end-diastolic pressure and LV relaxation time constant (tau), which are indices of diastolic function, were significantly increased in mice with myocardial infarction. In the case of finerenone treatment (1 mg/kg), a decreasing trend was obvious for these parameters of LV diastolic dysfunction; however, there was no observable change in the case of eplerenone (100 mg/kg). In mice with TAC-induced LV hypertrophy, altered cardiac remodeling was improved by finerenone; however, cardiac function was not significantly changed either by TAC intervention or by finerenone/eplerenone treatment. Above all, the TAC-induced increase in heart rate was reduced by finerenone but not eplerenone ^[22][31].

3.4. Vascular Remodeling and Function

The MR is expressed in vascular endothelial and VSM cells, and it is crucial for the contractile and relaxation responses of the vasculature. Genetic inactivation of the MR in endothelial cells ameliorated DOCA/salt-induced cardiac remodeling in a previous study ^[23][42]. Furthermore, finerenone treatment accelerated re-endothelialization, as well as attenuated VSM cell proliferation and neointimal lesion formation, reducing inflammation and exerting a protective effect against vascular remodeling in C57BL/6 mice ^[24][32]. Pharmacological blockade of the MR with finerenone reduced focal vasculopathy and vascular fibrosis in DOCA/salt-loaded UNX SD rats ^[9][21] and L-NAME-treated (mRen2)27 transgenic rats ^[25][28]. Intervention with finerenone enhanced acetylcholine-induced relaxation in the coronary artery in ovariectomized (OVX) mice ^[26][30], while it reduced the noradrenaline and angiotensin II-induced contractile response in the thoracic artery ^[27][29]. These data indicate that blockade of the MR with non-steroidal MRAs attenuates vascular injury during CVD development.

4. Molecular Mechanisms Underlying the Beneficial Effects of Non-Steroidal MRAs on CVD: Evidence from Pre-Clinical Studies

4.1. Oxidative Stress

Cumulative evidence demonstrates the impact of MR activation and associated oxidative stress in cardiac tissues, which contribute to the development of LV remodeling and dysfunction ^[28][29][43,44]. Of note, activation of the MR in a ligand-dependent manner positively regulates nicotinamide adenine dinucleotide phosphate (NADPH) oxidase subunits, an important component for reactive oxygen species (ROS) generation ^[30][45]. HS-loaded DSS rats exhibited high expression of 4-hydroxynonenal, increased expression of the components of NADPH oxidase gp47phox and p22phox, and elevated lipid peroxidation, as evidenced by an increased level of malondialdehyde in cardiac tissues ^[18][27]. However, esaxerenone treatment significantly improved all of these indicators of oxidative stress ^[18][27]. In contrast, ligand-independent activation of the MR is associated with ROS generation through the Rac1-dependent pathway ^[31][46]. Importantly, finerenone treatment attenuated NADPH oxidase activity in RacET transgenic mice with constitutively active Rac1 in the myocardium ^[32][34]. 

4.2. Inflammation

Cardiomyocyte-specific MR knockdown leads to a reduced inflammatory burden in abdominal aortic constriction in mice ^[33][47] and an L-NAME-treated hypertensive animal model ^[34][48], as well as a dramatic decrease in the infiltration of inflammatory cells in DOCA/salt-loaded mice ^[35][49]. In contrast, activation of the MR contributed to the maintenance of inflammation through activation of the nuclear factor kappa B (NF-κB) pathway, as well as the initiation of pro-inflammatory cytokine expression ^[36][50]. These data suggest the critical role of the MR in CV remodeling, dysfunction, and target end-organ damage. HS-loaded DSS rats exhibited very high levels of CXCL8, which is a key regulator of inflammatory cell influx, while esaxerenone treatment downregulated the abundance of CXCL8 in ventricular tissues. Additionally, inflammatory cytokines, specifically tumor necrosis factor (TNF)-α and interleukin (IL)-6, were reduced by esaxerenone treatment ^[18][27]. In a wire-induced femoral artery injury mouse model, finerenone significantly reduced intimal and medial leukocyte content, as evidenced by a decrease in the abundance of CD45+ cells ^[24][32]. Moreover, activation of the macrophage MR modulated their status toward the pro-inflammatory M1 phenotype, while MR knockdown in macrophages induced an anti-inflammatory M2 phenotype ^[34][48].

4.3. Interstitial Fibrosis

Cardiac fibrosis is caused by elevated accumulation and cross-linking of extracellular matrix proteins, leading to increased myocyte stiffening, which can impair ventricular relaxation and contraction of the myocardium ^[37][56]. Numerous studies have shown the critical role of MR activation in diffuse interstitial and perivascular collagen deposition in the process of cardiac fibrosis ^[38][39][57,58]. HS-loaded DSS rats exhibited interstitial and perivascular fibrosis in LV tissues. Treatment with esaxerenone reduced cardiac fibrosis in association with a reduction in transforming growth factor (TGF)-β, which is a pleiotropic mediator of cardiac fibrosis; collagen type I and III, which are major extracellular matrix proteins; and PAI-1, which is an inducer of regional cytokine production ^[18][27]. Serum and glucocorticoid-regulated kinase (SGK)-1 plays an important role in angiotensin or MR-induced cardiac fibrosis ^[40][59]. In HS-loaded DSS rats, esaxerenone treatment blunted the elevation in SGK-1 ^[18][27]. In neonatal rat cardiac fibroblasts, finerenone treatment attenuated aldosterone-induced nuclear MR translocation in association with the downregulation in connective tissue growth factor (CTGF), which is a central pro-fibrotic mediator that induces collagen production and subsequent pro-fibrotic enzymes; lysyl oxidase (LOX), which is a mediator of collagen cross-linking to form stable collagen fibers; TGF-β; and fibronectin, which is an extracellular matrix protein that regulates proliferation, differentiation, migration, and adhesion ^[32][34].

4.4. Vascular Injury

Aggravated activation of the MR in vascular endothelial and VSM cells has been indicated to contribute to the pathophysiology of HF ^[41][60]. Recent in vitro studies have demonstrated the preventive effects of finerenone in aldosterone-induced human umbilical vein endothelial cell apoptosis and coronary artery smooth muscle cell proliferation ^[24][32]. Furthermore, endothelial dysfunction in the coronary arteries is efficiently reduced by treatment with finerenone in OVX mice ^[26][30]. Aldosterone promotes atherosclerosis ^[42][61], where low-density lipoprotein cholesterol (LDL)-C plays a central role ^[43][62]. However, finerenone treatment reduces intrinsic arterial stiffness in Munich Wistar Frömter (MWF) rats accompanied by changes in elastin organization, normalization of matrix metalloproteinases, and reduction of oxidative stress ^[44][63].