1. ANTI-ALDOSTERONE DRUGS: WILL CINDERELLA BECOME A SUPERSTAR? Maria Rosa Costanzo, M.D. Medical Director, Midwest Heart Specialists Heart Failure Program Medical Director, Edward Hospital Center for Advanced Heart Failure Naperville, Illinois U.S.A

2. The Renin Angiotensin Aldosterone System

3. The Classic Genomic Action of Aldosterone on Epithelial Tissue Interstitium Fluid (Blood) Cortisol Aldosterone HRE Gene Nucleus Sgk1 CHIF Ki-Ras ENaCNa + Channel Na + /K + ATPase Cortisone MR K+K+ K+K+ H20H20H20H20 Na + 11β-HSD2 Endoplasmic Reticulum Na + K+K+ ATP ADP Lumen

4. Effects of Eplerenone, Enalapril and Eplerenone/Enalapril in Patients with Essential Hypertension and Left Ventricular Hypertrophy: The 4E-Left Ventricular Hypertrophy Study Hypertension with Echocardiographic Evidence of LVH 14-Day Placebo Run-IN Eplerenone 200 mg* (n =64) Enalapril 40 mg* (n =71) Eplerenone 200 mg/ Enalapril 10 mg* (n =67) Δ LV Mass by MRI Δ SBP/DBP Δ UACR Safety Pitt, B. et al. Circulation 2003;108:1831-1838 *Add-on therapy at week 8 with hydrochlorothiazide 12.5 or 25 mg and/or amlodipine 10 mg in patients with DBP 90 mm Hg or SBP >180 mm Hg

5. The 4E-LVH Study Pitt, B. et al. Circulation 2003;108:1831-1838 P<0.001 for Δ bsl for each group; *P=0.007 for epl/enal vs epl; † P=0.107 for epl/enal vs enala; ‡P=0.258 for epl vs enal Greater Change in LV Mass Despite Similar Reduction In BP!

6. Proposed Mechanisms of Aldosterone Excess, Linking Polymorphism in CYP11B2 Gene to Hypertensive Phenotype with Increased Aldosterone to Renin Ratio CYP11B2 Polymorphism in Lactic Dehydrogenase with Variants in CYP11B1 Reduced 11β-Hydroxylase activity Reduced Cortisol Production Chronic Compensatory Increase in ACTH Drive Adrenocortical Hyperplasia Increased Aldosterone Synthetic Capacity Hypertension with Increased Aldosterone to Renin Ratio MacKenzie SM and Connell JMC Current Hypertension Reports 2006; 8: 255-61

7. RALES and EPHESUS: Aldosterone blockade in HF and post-MI LV dysfunction RALES ( RALES (Randomized ALdactone Evaluation Study) N = 1633 with NYHA class III/IV HF N = 1633 with NYHA class III/IV HF Randomized to placebo or spironolactone 25 mg Randomized to placebo or spironolactone 25 mg Treatment in addition to ACE inhibitor and loop diuretic; most patients also received digoxin Treatment in addition to ACE inhibitor and loop diuretic; most patients also received digoxin EPHESUS ( EPHESUS (Eplerenone Post-Acute Myocardial Infarction Heart Failure Efficacy and SUrvival Study) N = 6632 with post-MI LV dysfunction and HFN = 6632 with post-MI LV dysfunction and HF Randomized to placebo or eplerenone 50 mg Randomized to placebo or eplerenone 50 mg Treatment in addition to ACEI or ARB,  -blockers, diuretics, aspirin Treatment in addition to ACEI or ARB,  -blockers, diuretics, aspirin Pitt B et al. N Engl J Med. 1999;341:709-17. Pitt B et al. N Engl J Med. 2003;348:1309-21.

8. Aldosterone Blockade and AT 1 Receptor Blockade: Trials in Post-MI LV Dysfunction and Heart Failure Pitt B et al. N Eng J Med. 1999;341:709-17. Pitt B et al. N Eng J Med. 2003;348:1309-21. RALES 0.75 0.60 1.00 0 Placebo Spironolactone 25 mg Months Probability of survival 2436 6 30% Risk reduction RR 0.70 (0.60–0.82) P < 0.001 30 0.00 12 18 0.90 0.45 EPHESUS 22 10 2 6 24 300 Eplerenone 50 mg Months 18 14 6 3612 15% Risk reduction RR 0.85 (0.75–0.96) P = 0.008 Cumulative incidence (%) Placebo 0 18

9. Zannad, F. et al. Circulation 2000;102:2700-2706 30-Month Mortality by treatment groups and PIIINP baseline levels Data from RALES

10. Relationship Between Transcardiac Extraction of Aldosterone and LV Remodeling in Patients with First AMI Hayashi, M. et al. J Am Coll Cardiol 2001;38:1375-1382

11. Antiarrhythmic Effects of Aldosterone Blockade* Yee, K.-M. et al. J Am Coll Cardiol 2001;37:1800-1807 Heart Rate VariabilityQT C Dispersion ○ = placebo ▲= spironolactone *Direct modulation of of the activity of voltage-dependent K+ channels Delpon E et al. Trends Pharmachol Sci 2005; 26:155-61

12. Extraadrenal Production of Aldosterone by Endothelial and Vascular Smooth Muscle Cells

13. Mechanisms of Aldosterone-Induced Vascular Fibrosis Extracellular Matrix Fibroblast Transformation monocyte T-cell Aldosterone Ang II genomic Brown, N. J. Hypertension 2008;51:161-7

14. Potential Mechanisms of Aldosterone-Induced Myocardial Fibrosis Renal Dependent –Increase in total body sodium –Hypertension –Potassium Deficiency Renal Independent (local vascular and cardiac effects) –Direct Myocardial Effects Increase in sodium influx into myocardial fibroblasts Activation of transcription factors activator protein-1 and NFkB Increase in collagen synthesis (perivascular and interstitial) Increase in expression of procollagen mRNA –Vasculitis Enhanced Vasoconstriction –Decrease in NO synthesis Coronary endothelium-independent dysfunction Activation of proinflammatory molecules –Cyclooxygenase-2 –Osteopontin –Macrophage chemoattractant protein-1 –IL-1β, IL-6 –Reactive oxygen species Formation of microthrombi –Increase in PAI-1

15. Mechanisms of Aldosterone-Induced Oxidative Stress and Endothelial Dysfunction Marney AM and Brown NJ Clinical Science 2007; 113: 267-78 G6PD NADPH NAD(P)H Oxidases NADP + ˙O2-˙O2- Nitric Oxide Synthase L-Arginine L-citrulline + ˙ NO ONOO - Aldosterone PP2A Ser 1177 P BH4 O2O2

16. Vidal, A. et al. Am J Physiol Heart Circ Physiol 2006; 290: H286-H294 Aldosterone-Induced Oxidative Stress as Demonstrated by Immunohistochemical Study of Activation of gp91 phox in Coronal Cryostat Sections of Ventricle Control gp91 phox, A Subunit of NADPH Oxidase Activity During Aldosterone Administration Aldosterone-Induced gp91 phox Activity Attenuated, but Not Prevented by Parathyroidectomy

17. Potential Pharmacological Interventions for the Proinflammatory/Fibrogenic Phenotype Vascular Fibrosis Tissue Invasion + Fibroblasts Proinflammatory Phenotype (ICAM-1. MCP-1, TNF-α) PMBC Activation Oxi/nitrosative Stress Ca 2+ Overload Ang II ALDO ET-1 PPARs HGM-CoA PDEI MMF Antioxidant CCB Receptor Antagonist ↓[Mg 2+] I ↑ NADPH Oxidase ROS, RNS NFkB

18. Martinez, D. V. et al. Hypertension 2002;39:614-618 Protection Against Myocardial Damage by Eplerenone or Low-Salt Diet in the L -NAME/Angiotensin II Model

19. *P<0.05 **P<0.001 * P<0.05 endothelium-dependent vasodilator endothelium-independent vasodilator competitive NOS inhibitor vasoconstriction only through conversion to Ang II Forearm Blood Flow Responses Farquharson, C. A. J. et al. Circulation 2000;101:594-7 ■ = placebo ▲= spironolactone *P<0.05

20. Sleep Apnea, Aldosterone, & Resistant Hypertension Pratt-Ubunama M. N. et.al. Chest 2007;131:453-459 Pimenta E et al. Progr Cardivasc Dis 2009; 51: 371-80 ρ = 0.44: p = 0.0002 Links between Aldosterone and OSA  Increased edema of nasopharingeal tissues  Oxidative Stress  Endothelial Dysfunction  Enhanced Endothelin-1 activity

21. Crosstalk Between Insulin and RAAS: Effects on Glucose Metabolism Activation of the RAAS Metabolic Effects of A II ↓ IRS protein content and phosphorilation ↑ TG secretion and accumulation ↑ gluconeogenesis ↓ glucose-mediated Insulin release ↑ MCP-1 expression in islets ↑ ROS Production ↓ adipocyte differentiation ↓ adiponectin production ↓ IRS-1 content and phosphorilation ↓ GLUT-4 translocation ↓ Glucose uptake Eplerenone Src PPI inhibitor Antioxidants Eplerenone Hitomi H et al. 2007; 50:750-5

22. Bidirectional Counter-Regulation Between the Hypothalamic- Pituitary-Adrenal Axis and the Immune System Hypothalamus Corticotropin- Releasing Hormone Anterior Pituitary Adrenal Cortex ACTH MC GC AT II K +, Mg 2+ NE, E ET-1 (+) (-) Immune System Cytokines Blood SNS and Vagal Afferents Weber KT Herz 2003; 28: 692-701

23. Renal Actions of Aldosterone Wenzel U Curr Opin Nephrol Hypertens 2008; 17: 44-50 Reabsorbed Na is pumped out of the cell by the Na-K-ATPase pump in the basolateral (peritubular) membrane. Spironolactone and eplerenone act by competing with aldosterone. Triamterene and amiloride function as indirect aldosterone antagonists by closing the epithelial sodium channels.

24. Aldosterone and Progression of Renal Disease Ramipril Ramipril+ Ibersartan Ramipril+ Spironolactone Ramipril + Ibersartan + Spironolactone Chrysostomu A et al. Clin J Am Soc Nephrol 2006; 1: 256-62 Bianch S et al. Kidney Int 2006; 2116-23

25. Mechanisms of Aldosterone-Dependent Renal Injury Kiyomoto H et al. J Pharmacol Sci 2008; 108: 399-405 1% NaCl 1% NaCl + Aldosterone Vehicle +1% NaCl 1% NaCl + Aldosterone 1% NaCl + Aldosterone +Eplerenone 1% NaCl +Aldosterone + Tempol MR in Mesangial cells Podocyte Abnormality Glomerular Sclerosis Superoxide Dismutase Mimetic Renally-Produced Aldosterone Increased MR in Mesangial Cells Increased transcription of MC-Responsive Genes ROS Production Podocyte Injury PROTEINURIA

26. Calcium Paradox of Aldosteronism and Its Consequences*** Vidal A et al. Am J Physiol Circ Physiol 2006; H286-H294 CELLACTIVATIONCELLACTIVATION Sequestration of Cytosolic Ca2+ by Mitochondria  H 2 O 2 -(1-2 wks) ↓α1AP–(1-6 wks)  gp91- (4wk) Parathyroidectomy +Ca2+ Suppl. Increased Aldosterone and Na + Hypercalciuria (6 wks) Hypermagnesuria (6 wks) Loop Diuretics Spironolactone Decreased Plasma Ca 2+ Concentration Decreased Plasma Mg 2+ Concentration Ca2+ Suppl. or Vitamin D 3 *** ≈ Zinc  PTH & ET-1 Parathyroidectomy Increased Cellular Expression of Ca 2+ Channels PMBC Platelets Vascular Cells Heart Muscle Cells Skin Skeletal Muscle CCM Parathyroidectomy

27. Human Counterparts of the Calcium Paradox of Parathyroidism ↑ urinary Ca 2+ excretion, ↓plasma ionized Ca 2+, ↑ plasma levels of PTH, and ↑ cytosolic free [Ca 2+ ] i found in pts. with low-renin HTN (Brickman AS et al. Hypertension 1990; 16: 515) High dietary Na +, which suppresses renin and aldosterone, and elevated aldosterone, inappropriate for 1% dietary NaCl, are each accompanied by hypercalciuria. (Ahokas RA et al. Circulation 2005; 111: 51-7 Immune cell activation accompanies secondary HPT of CRF (Alexievic JM et al. Kidney Int. 1996; 50: 1249-54) Secondary HPT (elevated plasma PTH and ET-1 levels, osteopenia and osteoporosis) found in HF pts. (Anker SD et al. Am J Cardiol 1999;83: 612-5) Hypovitaminosis D common in HF pts. (Shane E et al. Am J Med 1997; 103: 197-207 The cytokine profile of primary & secondary HPT resembles that of HF (Mann DL et al. Chest 1994; 105:897-904) (Mann DL et al. Chest 1994; 105:897-904) Loop diuretics exaggerate urinary Ca 2+ and Mg 2+ excretion and promote further PTH release and greater bone loss (Law PH at al. J Am Coll Cardiol 2005; 46: 142-6 The combination of thiazide diuretics and spironolactone reverses these losses (Runyan AL et al. Am J Med Sci 2005; 330: 1-7)

28. Reduction of Fracture Risk by Spironolactone in Men with CHF Fracture Site CasesControls Adjusted Odds Ratio (95% CI) p Value Total1676680.575(0.346-0.955)0.0324 Hip361440.848(0.255-2.8250.7889 Wrist1040-- Vertebral692760.581(0.252-1.343)0.2041 Other522080236(0.077-0.726)0.0118 Carbone LD et al. J Am Coll Cardiol 2008; 52: 135-8

29. Jorde, U. P. et al. Circulation 2002;106:1055-1057 Aldosterone Escape During Therapy with ACEIs N = 11 Upper Limit of Normal Range for Plasma Aldosterone 7/ 11 (64%) had AII/AI ratio 0.05, indicating complete inhibition of the vascular ACE

30. Juurlink et al. NEJM 2004;351:543 after RALES: RX

31. Juurlink et al. NEJM 2004;351:543 after RALES:Death

32. Guidelines for Minimizing the Risk of Hyperkalemia in Patients Treated With Aldosterone Antagonists 1.Impaired renal function is a risk factor for hyperkalemia during treatment with aldosterone antagonists. The risk of hyperkalemia increases progressively when sCr exceeds 1.6 mg/dL. * In elderly patients or others with low muscle mass in whom sCr does not accurately reflect GFR, determination that GFR or Cr clearance exceeds 30 ml/min is recommended. 2. Aldosterone antagonists should not be administered to patients with baseline serum potassium > 5.0 mEq/L. 3. An initial dose of spironolactone of 12.5 mg or eplerenone 25 mg is recommended, following which the dose may be increased to spironolactone 25 mg or eplerenone 50 mg if appropriate. 4. The risk of hyperkalemia is increased with concomitant use of higher doses of ACEIs (captopril ≥ 75 mg daily; enalapril or lisinopril ≥ to 10 mg daily. 5. NSAIDs and COX-2 inhibitors should be avoided. 6. Potassium supplements should be discontinued or reduced. 7. Close monitoring of serum potassium is required; potassium levels and renal function should be checked in 3 days and at 1 week after initiating therapy and at least monthly for the first 3 months. 8. Diarrhea or other causes of dehydration should be addressed emergently. Hunt SA et al.2009 Focused Update Incorporated Into the ACC/AHA 2005 Guidelines for the Diagnosis and Management of Heart Failure in Adults J Am Coll Cardiol, 2009; 53:1-90

33. Ezekowitz, J. A. et al. Eur Heart J 2009 30:469-77 Effects of Aldosterone Blockers on All-Cause Mortality in All Randomized Trials 01-Heart Failure 02-Myocardial Infarction

34. Ezekowitz, J. A. et al. Eur Heart J 2009 30:469-77 01-Heart Failure 02-Myocardial Infarction Effect of Aldosterone Blockers on LVEF in All Randomized Trials

35. Deleterious Actions of High Levels of Aldosterone in the Cardiovascular System Increased Aldosterone Level Kidney Heart Vasculature Na + reabsorption and water retention K + and Mg 2+ excretion Volume expansion and edema Electrolyte imbalance ↓ NE uptake Inflammation And tissue injury ↓ Myocardial function Ventricular arrhythmias Myocardial fibrosis Perivascular fibrosis ↓ Arterial compliance Baroreceptor dysfunction Endothelial dysfunction ↓Vascular reserve End-organ damage and cardiovascular disease

36. Conclusions Aldosterone has been shown to have deleterious cardiovascular and renal effects due to his genomic and non-genomic actions Aldosterone escape occurs in humans treated with ACEIs and ARBs; the addition of aldosterone antagonists can reduce BP and proteinuria, and importantly, reduce morbidity and mortality in HF pts. The risks of hyperkalemia in pts. treated with RAAS inhibitors and aldosterone antagonists are not insignificant, mandating close F/U, especially in pts. with underlying kidney disease Given the effects of aldosterone on fibrosis, inflammation, oxidative stress and endothelial dysfunction, aldosterone antagonists are headed for “Super Star” status!