2014 Novel Therapeutic Targets in the Management of Atrial
Transcription
2014 Novel Therapeutic Targets in the Management of Atrial
1 iš 22 www.medscape.com Novel Therapeutic Targets in the Management of Atrial Fibrillation Abhishek Maan, Moussa Mansour, David D. McManus, Vickas V. Patel, Alan Cheng, Jeremy N. Ruskin, E. Kevin Heist Am J Cardiovasc Drugs. 2014;14(6):403-421. Abstract and Introduction Abstract Atrial fibrillation (AF) is the most common cardiac arrhythmia, contributing to increased morbidity and reduced survival through its associations with stroke and heart failure. AF contributes to a four- to fivefold increase in the risk of stroke in the general population and is responsible for 10–15 % of all ischemic strokes. Diagnosis and treatment of AF require considerable health care resources. Current therapies to restore sinus rhythm in AF are suboptimal and are limited either by their proarrhythmic effects or by their procedure-related complications. These limitations have necessitated identification of newer therapeutic targets to expand the treatment options. There has been a considerable amount of research interest in investigating the mechanisms of initiation and propagation of AF. Despite extensive research focused on the pathogenesis of AF, a thorough understanding of various pathways mediating initiation and propagation of AF still remains limited. Research efforts focused on the identification of these pathways and molecular mediators have generated a great degree of interest for developing more targeted therapies. This review discusses the potential therapeutic targets and the results from experimental and clinical research investigating these targets. Introduction Atrial fibrillation (AF) is the most common cardiac arrhythmia encountered in clinical practice, contributing to increased morbidity and reduced survival through its associations with stroke, thromboembolism and heart failure.[1–3] Current antiarrhythmic drugs remain limited in their efficacy and carry the risk of adverse effects, including proarrhythmia.[4,5] The landmark discovery by Haïssaguerre et al.[6] that arrhythmogenic triggers in the pulmonary veins contributed to some cases of AF led to the development of pulmonary vein isolation as an effective interventional therapy for symptomatic drug-refractory AF. Catheter ablation, although efficacious, remains limited because of its invasive nature and possible procedural complications.[7,8] The current medical and invasive AF rhythm control strategies necessitate research focused on the identification of newer therapeutic targets to expand available treatment options. Despite the fact that the pathophysiology of AF has been extensively investigated for almost a century, the precise mechanisms leading to and maintaining AF still remains elusive.[9] This review focuses on the ° mechanisms of AF that might represent targets for future novel AF therapies. Structural and Electrical Remodeling as Therapeutic Targets Structural and electrical remodeling appear to be the key synergistic mechanisms contributing to the arrhythmogenic substrate underlying AF. Atrial fibrosis is the hallmark of arrhythmogenic structural remodeling.[10–12] Atrial fibrosis can also be seen as a convergent pathological end point in a variety of settings, including the aging process,[13] valvular disease,[14] ischemic heart disease[15] and cardiac dysfunction.[16] Multiple pathways, including the renin-angiotensin-aldosterone system (RAAS), and proinflammatory factors are potential targets for prevention of atrial fibrosis. Increased collagen deposition has been observed in patients with lone AF as compared with control patients in sinus rhythm (SR). [17] RAAS has been shown to be a predominant pathway implicated in myocardial fibrosis in the settings of congestive heart failure (CHF), myocardial infarction (MI), cardiomyopathy and hypertensive heart disease.[18] The adverse effects of RAAS seem to be predominantly mediated by increased production of angiotensin II (Ang II) (Fig. 1). Electrical remodeling in AF refers to the changes in electrophysiological properties as a result of AF. This process is facilitated by the alteration in ion-channel expression and function. This pathophysiological process is believed to occur as a compensatory mechanism to avoid Ca++ overload which adversely affects the viability of cardiac myocytes. This maladaptive mechanism (although autoprotective to an extent) further increases the susceptibility to develop sustained episodes of AF.[19,20] 2015-04-29 08:05 2 iš 22 Figure 1. Explaining molecular pathways mediating atrial fibrosis and electrical remodeling changes leading to atrial fibrillation (AF). AGE advanced glycation end products, Ang II angiotensin II, AP-1 activator protein-1, APD action potential duration, CHF congestive heart failure, Cx connexin, DM diabetes mellitus, ERK extracellular signal-regulated kinase, HTN hypertension, JNK c-Jun NH2-terminal kinase, MAP mitogen-activated protein, miRNA microRNA, NADPH nicotinamide adenine dinucleotide phosphate, NF- B nuclear factor kappa-light-chain-enhancer of activated B cells, PKC protein kinase C, RAAS renin-angiotensin-aldosterone system, RAGE receptor for advanced glycation end products, ROS reactive oxygen species, TGF 1 transforming growth factor- 1 Electrical remodeling can be envisioned as the result of short-term and long-term processes. The short-term factors which act within minutes to hours include (a) intracellular Ca++ increase which leads to inactivation of the L-type Ca++ channel and (b) post-translation modification of ion channels and neurohumoral factors which affect channel function. The long-term factors which contribute to the electrical remodeling act over a period of days to weeks and include (a) alteration in gene transcription and protein expression, (b) disturbances in Ca++ homeostasis and (c) increases in the oxidative stress and fibrosis. Therapeutic agents which can target these short- and long-term mechanisms facilitating electrical remodeling can be of potential interest as novel treatment options for suppression of AF. Angiotensin II-mediated Atrial Fibrosis Ang II has been well characterized as a profibrotic molecule that is implicated in cardiomyocyte apoptosis and reactive interstitial fibrosis.[21] Ang II-mediated atrial dilatation with focal atrial fibrosis leading to AF has been observed in transgenic mice with cardiacrestricted angiotensin-converting enzyme (ACE) overexpression.[22] Overproduction of Ang II promotes upregulation of mitogenactivated protein (MAP) kinase signaling, an important downstream mediator that further leads to altered gap-junctional coupling and conduction properties.[23,24] Ang II has also been demonstrated to mediate cardiac hypertrophy by upregulation of c-Jun 2015-04-29 08:05 3 iš 22 NH2-terminal kinase (JNK) and extracellular signal-regulated kinase (ERK).[25] In a mouse model, an increase in the production of Ang II by rapid electrical stimulation was found to be associated with increased expression of connexin 43 (Cx43). This effect was also associated with upregulation of MAP kinases, suggesting their activation could play a key role as mediators of adverse electrical and structural remodeling in AF. The final effect of Cx43-mediated alteration of gap junctions is believed to lead to an alteration in conduction properties that is proarrhythmic.[26] A clinical study has suggested that a decrease in the ratio of atrial Cx40 to Cx43 is correlated with chronic AF, so this may be secondary to either a rise in Cx43 or loss of Cx40.[27] The role of matrix metalloproteinases (MMPs) is also being increasingly recognized in the extracellular matrix (ECM) remodeling which leads to atrial dilatation and contributes to the pathogenesis of AF. MMP expression and regulation, which are regulated at various steps, are relevant in matrix turnover and structural remodeling further contributing to AF. MMP-1 is primarily responsible for degradation of collagen components, and a decrease in its level is associated with an increased degree of atrial fibrosis.[28] Overexpression of MMP-9 is also observed to be another key process in the pathogenesis of AF via increasing atrial fibrosis.[29] Ang II has been demonstrated to mediate an increase in the level of MMP-9.[30] Similarly, the profibrotic role of Ang II is further substantiated by its inhibition in the levels of MMP-1.[31] Myeloperoxidases (MPOs) have also generated a keen degree of interest in the pathogenesis of atrial fibrosis. The potency of MPOs to generate reactive oxygen species (ROS) and free radicals has been believed to be central to their role in perpetuation of AF. The mechanistic link between MPOs and AF development was clarified by Rudolph et al.[32] in their experiments. The investigators observed that the angiotensin II-treated mpo mice were protected against the fibrotic remodeling of the atria as compared with the wild-type mice. The role of MPOs was further confirmed by a dose-dependent increase in the AF vulnerability when the wild-type and mpo mice were exposed to the intravenous infusion of MPOs.[32] In addition to the effects of Ang II leading to structural remodeling of atrial tissue, there is further evidence that it also contributes to electrical remodeling.[33–35] Ang II has been demonstrated to modulate the inward calcium (ICa) current density by activation of protein kinase C (PKC) and induction of phosphoinositide-3-kinase (PI3K)-dependent pathways.[36,37] Another recently published study found that Ang II also increased the L-type calcium channel (LCC) density by upregulating the transcription of LCC messenger RNA (mRNA) via PKC, ROS and cyclic adenosine monophosphate response element-binding protein pathways.[36] Therapeutic Interventions Targeting the Renin-angiotensin-aldosterone System On the basis of the established relationship of RAAS, atrial fibrosis and AF, angiotensin-converting enzyme inhibitors (ACEIs) and angiotensin receptor blockers (ARBs) have been investigated in the context of primary and secondary AF prevention. Li et al.[37] investigated the mechanistic effect of ACEIs on atrial remodeling in a canine ventricular tachypacing (VTP)-induced CHF model and observed that the administration of enalapril was associated with a significant decrease in tissue concentration of Ang II and also reduced the expression of the phosphorylated form of ERK. The administration of enalapril was also associated with attenuation of atrial fibrosis and conduction heterogeneity.[37] Similar experiments by Shi et al.[38] also investigated the role of enalapril in attenuation of atrial fibrosis in a VTP-induced CHF canine model. Atrial fibrosis was estimated with left atrium (LA) functional area shortening (FAS) (measured on transthoracic as systolic area – diastolic area/systolic area × 100), and AF inducibility was measured by the effective refractory period (ERP) and AF duration. The treatment with enalapril was observed with attenuation of atrial fibrosis which further correlated with histological evidence as well as echocardiographic measurement of FAS.[38] Another study, by Saygili et al.,[39] observed that, in cultured atrial myocytes exposed to stretch, treatment with the ARB losartan prevented stretch-induced and consequently angiotensinmediated atrial hypertrophy. In addition, losartan also attenuated stretch-induced alterations in I K1, IKur, and I to density and thereby prevented stretch-induced decreases in action potential duration (APD). In addition to these salutatory effects of ACEIs/ARBs on atrial fibrosis, Chen et al. [40] demonstrated that these agents also mitigated local Ang II-induced automaticity in isolated rabbit pulmonary vein tissue preparations and isolated cardiomyocytes. Further investigations by Shetty and DelGrande [41] also observed that the ARBs irbesartan and valsartan reduced local Ang II-mediated norepinephrine spillover in rat atria, and thereby led to a decrease in conduction heterogeneity and inducibility of AF. Endothelial dysfunction also has a pertinent role in the pathogenesis of AF; an increased expression of adhesion molecules mediated by Ang II has been demonstrated in the fibrillating atria. Goette et al.[42,43] used a rapid atrial pacing model in pigs to show treatment with ACEIs abrogated the increased expression of adhesion molecules, further supporting the role of ACE inhibition in mitigating the endothelial dysfunction. Clinical Studies A recently published meta-analysis of 14 clinical trials including 92,817 patients demonstrated that RAAS inhibition with an ACEI was associated with a modest decrease in the incidence of new-onset AF as compared with conventional therapy or placebo [relative risk (RR) 0.79, 95 % confidence interval (CI) 0.62–1.00, p = 0.05]. ARBs showed a similar effect in reduction of AF (RR 0.78, 95 %CI 0.66–0.92, p = 0.009).[44] The authors of this meta-analysis acknowledged that although there was a modest degree of benefit of RAAS inhibition in the prevention of AF, the overall quality of evidence of these findings was low. 2015-04-29 08:05 4 iš 22 Data from the Losartan Intervention For Endpoint Reduction in Hypertension (LIFE) study further support the benefit of RAAS antagonism for prevention of AF. The LIFE investigators showed that, despite similar reductions in blood pressure, patients in the losartan arm had a lower incidence of new-onset AF than patients in the atenolol arm (6.8 vs. 10.1 per 1,000 patient-years; RR 0.67, 95 % CI 0.55–0.83, p<0.001).[45] Further analyses from this study also showed that a greater degree of regression of left ventricular hypertrophy (LVH) with the use of losartan as compared with atenolol might also be responsible for prevention of AF. A greater degree of LVH regression might contribute to greater improvement in left atrial hemodynamics and dilatation. These additional mechanisms also seem pertinent in the beneficial effects of losartan in the LIFE study. Similarly, in a broad CHF patient population, candesartan was found to be more effective in prevention of new-onset AF as compared with placebo [5.55 % patients developed new-onset AF in the candesartan arm vs. 6.74 % in the placebo arm; odds ratio (OR) 0.81, 95 % CI 0.66–0.99, p = 0.048] in the Candesartan in Heart Failure: Assessment of Reduction in Mortality and Morbidity (CHARM) trial.[46] Contrary to these beneficial effects of ACE inhibition particularly for prevention of new-onset AF, the results of the Gruppo Italiano per lo Studio della Sopravvivenzanell'Infarto Miocardico-Atrial Fibrillation (GISSI-AF) trial showed that valsartan did not reduce the recurrence rates for AF in patients with pre-existing cardiovascular diseases.[47] The Atrial Fibrillation Clopidogrel Trial with Irbesartan for Prevention of Vascular Events (ACTIVE-I) trial investigated the impact of irbesartan on a composite of adverse cardiovascular events (stroke, MI, death from vascular causes as the primary outcome). The rate of this outcome was observed to be similar for both irbesartan and placebo [5.4 % per 100 patient-years in each; hazard ratio (HR) with irbesartan 0.99, 95 % CI 0.91–1.08, p = 0.85]. The use of irbesartan also did not have a significant effect in reducing the rate of hospitalization due to heart failure in this study.[48] The Angiotensin II-Antagonist in Paroxysmal Atrial Fibrillation (ANTIPAF) trial aimed to investigate the impact of olmesartan on the AF burden (measured by transtelephonic electrocardiograms) in patients with paroxysmal AF without any coexisting structural heart disease. During a 12-month follow-up period, the use of olmesartan was not observed to have any beneficial effect in decreasing AF burden (p = 0.77). Similarly, the secondary outcomes of the trial (quality of life, time to develop persistent AF, and rate of cardiovascular hospitalizations) were also not affected by the use of olmesartan as compared with placebo.[49] The lack of clinically beneficial effects with the use of ARBs for prevention of AF suppression was further substantiated by the results of the Japanese Rhythm Management Trial II for Atrial Fibrillation (J-RHYTHM II). In this open-labeled randomized trial, symptomatic and asymptomatic recurrences of AF episodes were assessed in patients with paroxysmal AF who received either candesartan or amlodipine. At a follow-up period of 1-year, the frequency of total AF was 2.1 ? 3.8 days/month in the candesartan group as compared with 2.4 ± 4.4 days/month in the amlodipine group, p = 0.51. The use of candesartan was also not found to have a significant effect on the frequency of cardiovascular events, left atrial dimensions and subsequent development of persistent AF.[50] Aldosterone Antagonists Aldosterone antagonists have also been investigated in both pre-clinical and clinical studies as potential therapeutic antagonists of the RAAS system. The role of these agents might be even more relevant in AF, considering the evidence of intracardiac production of aldosterone, which has been shown to upregulate the gene expression of ACE[51] and may also have a direct role in fibrosis[52] and electrophysiological changes.[53] There is also evidence from clinical studies which supports the finding that patients with AF have a relatively higher serum level of aldosterone as compared with patients in SR,[54] and these levels decrease after SR is restored in AF patients.[55] Milliez et al.[56] investigated the role of a 1-month treatment regimen of spironolactone alone and in combination with lisinopril and atenolol in a 3-month post-MI rat model using echocardiographic and P-wave measurements on electrocardiogram. Spironolactone was observed to mitigate the atrial fibrosis measured by increased LA mass and also decreased the post-MI elevation in left ventricular end-diastolic pressure, which was measured as a covariate in this study. The authors in this study also speculated a superior efficacy of spironolactone, particularly in reducing the atrial fibrosis once it had occurred.[56] This observation could be potentially explained by the fact that once atrial fibrosis has ensued, it might become relatively less sensitive to the hemodynamic alterations and to the activity of the RAAS agents.[57] Another study, by Shroff et al.,[58] further elucidated the role of eplerenone in a VTP-induced CHF model. The results from this study indicated that selective aldosterone blockade using eplerenone was associated with improvement in diastolic dysfunction and reduced AF inducibility by prolongation of atrial ERP. The investigators in this study speculate that the prolongation of atrial ERP with the use of eplerenone could be due to its additional ionic-modulation properties.[58] Clinical Studies These findings were further supported by a retrospective analysis of the Eplerenone in Mild Patients Hospitalization and Survival Study in Heart Failure (EMPHASIS-HF) study. In this study, randomization to eplerenone was observed to be associated with a decreased incidence of new-onset AF as compared with placebo (2.7 vs. 4.5 %, HR 0.58, 95 % CI 0.35–0.96, p = 0.034).[59] Another recently published study, by Ito et al.,[60] investigated the efficacy of eplerenone on clinical outcomes of 161 patients with long-standing persistent AF who underwent radio-frequency ablation; after a follow-up period of 24 months, eplerenone was observed to be associated with a greater incidence of freedom from AF (60 vs. 40 % in the non-eplerenone group, p = 0.011). 2015-04-29 08:05 5 iš 22 Although these data are from small observational studies, spironolactone may be potentially useful in reducing AF recurrences,[61] AF burden and hospitalizations necessitated by cardioversion.[62] Further studies focused on investigation of additional mechanisms of aldosterone antagonists and larger clinical studies validating these findings are still needed to make a compelling recommendation for encouraging the use of these agents for prevention of AF ( ). Table 1. Pre-clinical mechanisms and data from clinical studies on ACEIs, ARBs and aldosterone antagonists Agent Mechanism of action Clinical evidence References ACEIs Decrease Ang II production Meta-analysis demonstrated benefit in reducing new-onset AF [44] ARBs Attenuates Ang II-mediated downstream effects (a) Beneficial effects seen in LIFE and CHARM trials, negative evidence from GISSI-AF trial [45–49] (b) Lack of beneficial effect in reducing cardiovascular outcomes in patients with AF in the ACTIVE I trial (irbesartan) (c) Lack of efficacy in suppressing AF in the ANTIPAF trial (olmesartan) Aldosterone antagonists Decrease cardiac production of aldosterone Reduction in new-onset AF with eplerenone in EMPHASIS-HF [59, 60] trial, decreased AF recurrences after catheter ablation ACEI angiotensin-converting enzyme inhibitor, AF atrial fibrillation, Ang II angiotensin II, ARB angiotensin receptor blocker Effect of Pioglitazone on Atrial Fibrosis Recently, there has been a great degree of interest in the role of peroxisome proliferator-activated receptor (PPAR- ) agonists. The activation of these receptors has been observed to reduce the expression of pro-inflammatory cytokines such as interleukin (IL)-1 and IL-6, inducible nitric oxide, tumor necrosis factor- (TNF- ) and MMP-9 by inhibition of transcription factors which bind to nuclear factor kappa-light-chain-enhancer of activated B cells (NF- B) in monocytes and macrophages.[63,64] Similarly, this receptor also leads to a decreased production of TNF- by downregulating the function of NF- B.[65] Shimano et al.[66] investigated the role of pioglitazone in comparison and combination with candesartan for its role in mitigation of atrial fibrosis and prevention of AF based on a VTP-induced CHF rabbit model. The investigators observed that the treatment with pioglitazone was associated with attenuation of atrial fibrosis, significant reductions in the interatrial activation time and duration of AF as compared with controls. These changes in the pioglitazone group were comparable with those in the candesartan group. In the group which received the combination of pioglitazone and candesartan, no additional reduction in attenuation of atrial fibrosis and reduction in AF was observed. The findings that both pioglitazone and candesartan decreased the expression of transforming growth factor- 1 (TGF- 1), TNF- and the phosphorylated form of ERK suggest that they might have a common underlying antifibrotic mechanism.[66] Another study, by Xu et al.,[67] showed that pioglitazone reduced age-related atrial fibrosis and AF propagation by upregulating the gene expression of antioxidant molecules such as superoxide dismutase (SOD-2) and heat shock protein (hsp 70) and by preventing hyperactivity of nicotinamide adenine dinucleotide phosphate (NADPH) oxidase responsible for free-radical generation. This study also highlighted the role of pioglitazone in decelerating cardiac apoptosis which was predominantly mediated by the activation of bad signaling pathway molecule via the phosphoinositide-3 kinase pathway. Pioglitazone also led to inhibition of caspases-3 and 9, which are also relevant molecules implicated in cardiac apoptosis.[67] A recently published study by Gu et al.[68] on cultured atrial myocytes demonstrated that pioglitazone attenuated Ang II-induced connective tissue growth factor (CTGF) expression and proliferation of atrial fibroblasts. This study also demonstrated these effects were mediated by at least partial if not complete inhibition of the TGF- 1/Smad2/3 pathway.[68] Further investigations, by Takahashi et al.,[69] also demonstrated the beneficial effect of pioglitazone in attenuating hypertrophy-mediated atrial fibrosis by repression of monocyte chemoattractant protein-1, which was observed to be a key profibrotic molecule involved in atrial fibrosis. Clinical Studies Gu et al.[70] also investigated the role of pioglitazone in 150 patients with type 2 diabetes mellitus (DM) who underwent catheter ablation for drug-refractory paroxysmal AF. After a mean follow-up duration of 22.9 ± 5.1 months following index ablation, SR was maintained in 44/51 patients (86.3 %) in the pioglitazone group versus 70/99 patients (70.7 %) in the control group (p = 0.034). Transforming Growth Factor- 1 (TGF- 1) TGF- 1 is another key mediator of atrial fibrosis which is also observed to be a primary downstream mediator of Ang II effects. [71,72] Pre-clinical studies based on treatment of human atrial tissue with Ang II have demonstrated an overexpression of TGF- 1 mRNA.[73,74] Furthermore, there is accumulating evidence that Ang IImediated overexpression of TGF- 1 could also be facilitated by multiple mechanisms such as overexpression of TGF- 1 mRNA[74] and activation of MAP kinase by either Ca++- or PKC-dependent pathways.[75] TGF- 1 predominantly acts through the mothers against decapentaplegic (Drosophila protein) (SMAD) signaling pathway to stimulate collagen production.[76,77] An increased atrial expression of TGF- 1 has been observed in 2015-04-29 08:05 6 iš 22 the VTPinduced CHF animal model.[78] Targeted overexpression of TGF- 1 in mice has been observed as a key mediator of selective atrial fibrosis, conduction heterogeneity further leading to AF propagation.[79,80] Therapeutic Interventions on TGF- 1 Considering the central role of TGF- 1 as a mediator of atrial fibrosis, it is intuitive to believe that anti-TGF- 1 therapies could be potentially useful to inhibit atrial fibrosis and hence prevent the development of AF. Pirfenidone, 5-methyl-1-phenyl-2 (1H)-pyridone, was investigated by Lee et al.[81] in a VTP-induced CHF canine model. Induction of CHF was found to be associated with increased degree of atrial fibrosis and atrial conduction heterogeneity as compared with controls, whereas the treatment with pirfenidone resulted in decreased atrial fibrosis, conduction heterogeneity and hence reduced AF vulnerability. Furthermore, pirfenidone was also found to reduce the TGF- 1-mediated downstream expression of various MAP kinases studied on cytokine profile in this study.[81] Treatment with pirfenidone was also observed to be associated with a decrease in MMP-9 and tissue inhibitor of metalloproteinase-4 levels, further supporting its potential as an antifibrotic agent. Yamazaki et al.[82] also demonstrated the beneficial effects of pirfenidone in an Ang II-mediated cardiac hypertrophy model in mice and demonstrated its efficacy in mitigating Ang II-mediated perivascular and interstitial fibrosis and TGF- 1 expression; additionally, pirfenidone also attenuated the expression of mineralocorticoid receptors. Various other models investigating pulmonary fibrosis have also studied the antifibrotic role of pirfenidone.[83–85] In the future, more robust pre-clinical and clinical studies are likely to help clarify the relevance of pirfenidone as a novel agent for prevention of AF ( ). Table 2. Role of various potentially therapeutic agents based on experimental studies Therapeutic agent Mechanism of action Comments References Pioglitazone PPAR- agonist Decreases pro-inflammatory cytokine (TNF- , IL-6 and MMP-9) production [63–70] Pirfenidone Anti-TGF- 1 agent Decreases TGF- 1-mediated activation of kinases and attenuates atrial fibrosis [81–85] Statins Antioxidant, pleiotropic effects Decrease Ang II-mediated free-radical production and inhibit rac1-mediated activation of NADPH oxidase [151] PUFAs Modulates membrane Counteract arrhythmogenic effect of atrial stretch by modulating fluidity membrane proteins and regulate MAP kinase activity [152–155] Ang II angiotensin II, IL-6 interleukin-6, MAP mitogen-activated protein, MMP matrix metalloproteinase, NADPH nicotinamide adenine dinucleotide phosphate, PPAR- peroxisome proliferator-activated receptor, PUFA polyunsaturated fatty acid, TGF- 1 transforming growth factor- 1, TNF- tumor necrosis factorRecent investigations by Barter et al.[86] have also identified histone deacetylases (HDACs) as regulatory mediators of the profibrotic function of TGF- 1 at the gene-expression level. Several genes (such as smad2, 3 and 7) which regulate matrix turnover in response to TGF- 1 are selectively regulated by HDACs. Selective inhibition of HDACs was demonstrated to suppress the induction of profibrotic signals (disintegrin and metalloproteinase-12) which are driven by TGF- 1.[86] In addition, Liu et al.[87] showed that global HDAC inhibition could reverse atrial fibrosis and inducible atrial arrhythmias. In this study, mice that were ~3 months old already had established atrial fibrosis and inducible atrial arrhythmias. After treatment with the global HDAC inhibitor trichostatin A daily for 2 weeks, the percentage of atrial fibrosis and arrhythmia burden were reduced in the experimental group when compared with wild-type littermate mice. Interestingly, the degree of ventricular hypertrophy and diastolic dysfunction were not affected by the HDAC inhibitor, but along with a reduction in atrial fibrosis, there was an increase in Cx40 gap junctions in the atrium of mice treated with HDAC inhibitor. These observations support the role of HDACs as key mediators of profibrotic proliferation signal in fibroblasts which are activated by TGF- 1. While it is intuitive to be optimistic about the role of antifibrotic therapy in AF, it is also important to be cognizant about the limitations of the role of atrial fibrosis in pathogenesis of AF. There is also a further need to better understand the relationship between the extent of atrial fibrosis and the threshold for AF initiation. Knowledge of a precise mechanism by which atrial fibrosis leads to conduction heterogeneity and development of AF will help clarify whether atrial fibrosis is the central event in the pathogenesis of AF. MicroRNAs Recently there has been a rapid evolution in the field of microRNAs (miRNAs) as novel mediators of various cardiovascular conditions at the molecular level. miRNAs are non-coding RNAs that are 19–25 nucleotides in length that repress gene expression at the post-transcriptional level by acting on the 30-untranslated region of RNAs.[88,89] 2015-04-29 08:05 7 iš 22 As discussed in the previous sections, structural atrial remodeling results from an imbalance between the anti- and profibrotic pathways; similarly, certain miRNAs can be visualized as mediators of antifibrotic and others as mediators of profibrotic processes. Xu et al.[90] investigated the expression profile of various miRNAs in three groups of dogs (adult and aged dogs in SR and aged dogs with persistent AF induced by rapid atrial pacing) and observed that, as compared with adult dogs, the expression of miRNA-21 and 29 was significantly increased in the older dogs whereas the expression of miRNA-1 and 133 was downregulated; when the aged SR group was compared with the aged AF group, miRNA-1, 21 and 29 showed an increased expression, while miRNA-133 showed downregulation in the aged AF group. The investigators in this study also observed that miRNA-1 was also implicated in increased apoptosis by repressing the levels of hsp 60 and hsp 70 and this proapoptotic effect of miRNA-1 was attenuated by miRNA-133 through deactivation of caspase-9.[90] Another recently published study, by Dawson et al.,[91] using a VTP-induced HF model in dogs, observed that the atrial expression of miRNA-29 was downregulated in AF as compared with SR. The mechanisms linking miRNA-29 to AF were further clarified in a knockdown model where the absence of miRNA-29 was associated with upregulation of collagen producing genes (COL1A1 and COL3A1).[91] Upregulation of miRNA-21 in rat atria was associated with atrial fibrosis and subsequent development of AF. These findings were confirmed by the lesser amount of atrial fibrosis in an atrial miRNA-21 knockdown model.[92] miRNA-21 promotes cardiac fibrosis by upregulating the activity of cardiac ERK and MAP kinases through the inhibition of sprouty homologue 1.[93] The role of miRNAs has also been thought to be particularly relevant in mediating the process of electrical remodeling. From a conceptual standpoint, it is now widely agreed upon that electrical remodeling of atrial tissue results in shortening of atrial ERP, which further favors the generation of AF due to re-entry. A simplified way to visualize this process could be an imbalance mediated by two important alterations. The first change is the reduction of L-type Ca++ (I CaL) and transient outward K+ (Ito) currents which are primarily responsible for decreasing the plateau phase of the atrial action potential.[94,95] The second relevant process involved with shortening of the terminal phase of the action potential is an increase in the inward rectifier K + current (I K1).[96] In a study by Yue et al.[97] investigating the ionic mechanisms underlying propagation of AF in a canine model, the alteration in current density was found to be related to the reduction in functional channels without any alterations in the other properties of ion channels such as voltage, time and frequency dependence. These findingswere further supported by another study, by Gaborit et al.,[98] which compared the ion-channel gene expression profiles in patients with AF and SR and observed that AF patients had a disproportionate upregulation in functional Kir 2.1 current compared with the overexpression of its codingmRNA(twofold increase vs. 20 %increase), suggesting that post-transcriptional modification of the Kir 2.1 channel subunit may be responsible for these changes. The results of these two key hypothesis-generating studies were further validated in a mouse model of AF by Yu et al.,[99] who reported an increased expression of miR-328 in AF as compared with SR and forced overexpression of miR-328 led to an increased propensity to develop AF mediated by diminished L-type Ca++ current, which was mediated by downregulation of the genes coding (CACNA1C and CACNB1) the cardiac L-type Ca++ channel by miR-328. Clinical Studies A recently published investigation by McManus et al.[100] based on analysis of the Framingham study reported that circulating miRNA-328 levels were significantly lower in the patients with prevalent AF as compared with patients without AF (8.76 vs. 7.75, OR 1.21, 95 % CI 1.09–1.33, p<0.001). The mechanistic basis of miRNAs leading to ionic remodeling and thereby shortening of atrial APD was investigated by Luo et al.,[101] who observed that miR-26 was downregulated in samples from animals and patients with AF accompanied by an upregulation of I K1/KIR 2.1 protein. Investigations by Ling et al.[102] have also implicated the role of miRNA-499 in the pathogenesis of AF; on the basis of the comparison of miRNA expression profile in AF versus SR patients, the levels of miRNA-499 were observed to be upregulated in AF as compared with SR patients. Upregulation of miRNA-499 was also accompanied by downregulation of the small conductance calcium-activated potassium channel-3 (SK3), which was further confirmed by transfection experiments with miRNA-499. While the exact role of the SK3 channel is still under investigation, it is believed to be involved with electrical remodeling changes mediating the development of AF. Apart from being implicated in these two fundamental pathways of structural and electrical remodeling leading to AF, miRNAs are also being investigated as mediators in the cardiac apoptotic pathway in response to oxidative stress (miRNA-133).[103] Based on data from these studies, it is conceivable that various miRNA molecules could very well turn out to be not only therapeutic targetsbut also potential biomarkers in AF ( ). Table 3. Role of various miRNAs in the pathogenesis of atrial remodeling leading to AF miRNA Function and role in AF References miRNA-1 Upregulated in AF, increases apoptosis in cardiomyocytes by repressing heat shock proteins (hsp 60 and hsp 70) [90] miRNA-21 Repression of sprouty-1 and upregulation of phosphorylated ERKs [92, 93] miRNA-29 Downregulated in AF leading to increased expression of collagen genes (COL1A1 and COL3A1) [91] 2015-04-29 08:05 8 iš 22 miRNA-133 Downregulated in AF, attenuates the proapoptotic function of miRNA-1 by deactivation of caspase-9, some relevance in oxidative stress-mediated AF miRNA-328 Increased expression of miR-328 in AF, diminished L-type Ca++ current mediated by [99, 100] downregulation of the genes coding (CACNA1C and CACNB1) decreasing APD and promoting AF miRNA-499 Increased expression in AF leading to downregulation of function of SK3 channels leading to decrease in repolarization currents [103] [102] AF atrial fibrillation, APD action potential duration, ERK extracellular signal-regulated kinase, miRNA micro ribonucleic acid, SK3 small conductance calcium-activated potassium channel-3 Role of Ion Channels as Potential Targets in Atrial Fibrillation (AF) Normal cardiac electrical activity (reflected by APD) is facilitated by the proper functioning of the ion channels. Electrical remodeling reflected by a reduction in APD is a key process in promoting initiation and propagation of AF. The intracellular resting potential of atrial cells is maintained at a negative value by a large resting K+ permeability, which in turn is governed by the principal cardiac inward-rectifier current I K1. I K1 is composed of Kir 2.1 subunits which are overexpressed in chronic AF.[104,105] Another inwardrectifier K+ current, I KACh, is upregulated by vagal activation and mediates the effects of acetylcholine. The upregulation of both these currents leads to shortening of APD and thereby provides an electrical basis for AF pathogenesis.[106] The increased activation of IKACh is mediated by its reduced inhibition by the protein kinase (PKC isoform ).[107] Upon activation, the cells are depolarized by the rapid entry of Na+ through the Na+ channels, which generates a large inward-flowing current (I Na). Following this phase, the cells undergo brief repolarization, which is mediated by the outward flow of K+ via the Ito channel. This phase is followed by the plateau phase, during which there is a balance between the inward current I CaL (mediated by Ca++ entry through the L-type Ca++ channel). Reduction in ICaL contributes to shortening of APD and is of particular relevance in the pathogenesis of re-entry-mediated AF.[108] The plateau phase of APD is also of key interest in the pathogenesis of AF mediated by altered impulse formation, which is believed to be caused by ''afterdepolarization.'' Early afterdepolarizations (EADs) are a result of an impairment of action potential repolarization which leads to a prolongation of the plateau phase. These EADs are mediated by either an excess of inward currents (ICaL or late INa) or decreased activity of I to which mediates early repolarization.[109] The plateau phase is followed by the late repolarization which is mediated by the rapid (IKr) and slow (I Ks ) K+ currents. Additionally, this phase is also mediated by the ultrarapid delayed rectifier current (I Kur) in the atrial cells. The activity of I Kur in turn is mediated by the Kv 1.5 channel. This phase is also of relevance in the role of delayed afterdepolarizations (DADs), which are initiated by the activity of Na+-Ca++ exchanger (NCX) and an abnormal Ca++ leak from the subcellular stores (i.e., sarcoplasmic reticulum). The DADs are further mediated by the dysfunction of ryanodine receptor 2 (RyR2) in the patients with AF.[110] In the experimental models of AF, hyperphosphorylation of RyR2 by protein kinase A (PKA) seems to contribute to both an increased activation as well as its increased sensitivity to cytosolic Ca++ [111–113] (Fig. 2, ). Table 4. Role of various ion-channels in various phases of action potential and pathogenesis of AF Ion channel Role in action potential Relevance in pathogenesis of AF IK1 Maintains the negative resting membrane potential Increased in AF because of increased expression of Kir 2.1, which could be mediated by decreased expression of miR-26 and miR-101 IKACh Inward current, mediates vagal effects Increased expression detected in AF, believed to play a relevant role in vagally mediated AF Increase mediated by enhanced open probability of Kir 3.1 (GIRK1) and Kir 3.4 (GIRK2), which is mediated by reduced inhibition by PKC (PKC ) and increased activation of PKC INa Responsible for rapid action potential depolarization (Phase 0) Decreased INa leads to slowing of conduction and promotes re-entry-mediated AF Ito Mediates brief rapid repolarization phase (Phase 1) Definite role in the pathogenesis of AF remains unclear, speculated to be atrialselective target, also believed to be a mediator in AF-related ionic remodeling ICaL Mediates the plateau phase, inward Ca++ current (Phase 2) Phosphorylated form observed to hyperactive and hypersensitive to cytosolic Ca++ PKA-mediated hyperphosphorylation of RyR2 is seen in AF models Reduction favors shortening of APD and AF perpetuation 2015-04-29 08:05 9 iš 22 IKs Slow component of delayed rectifier current (Phase 3) Alterations in delayed-rectifier current speculated to be involved in AF, reduction in IKs could lead to shortening of APD and promote AF IKr Rapid component of delayed rectifier current (Phase 3) Contributes to the delayed-rectifier current and thus could promote development of AF If Contributes to Phase 4 Limited data, unclear role NCX Contributes to Phase 4 and also Mediates the initiation of DADs, increased expression is reported in AF implicated in Ca++ homeostasis AF atrial fibrillation, APD action potential duration, DADs delayed afterdepolarizations, GIRK G-protein coupled inwardly-rectifying potassium channels, NCX Na+-Ca++ exchanger, PKA protein kinase A, PKC protein kinase C, RyR2 dysfunction of ryanodine receptor 2 Figure 2. Role of ion channels in action potential duration and pathogenesis of atrial fibrillation (AF). PartArepresents the role of various ion channels in various phases of action potential. Afterdepolarizations occurring after full repolarization [delayed afterdepolarizations (DADs); part B] and incomplete repolarization [early afterdepolarizations (EADs); part C] mediate the pathogenesis of AF by causing premature beats. Conditions which promote spontaneous re-entry of electrical impulses are shown in the figure D. Re-entry is 2015-04-29 08:05 10 iš 22 initiated by a premature beat (indicated by a yellow star), acting on interconnected zones of tissue with two alternate conduction pathways that have different refractory periods. The numbers 1, 2, and 3 in part D represent different sites of electrical activation. (Modified and reproduced with permission from Dobrev et al. [110]) Abnormal release of Ca++ from sarcoplasmic reticulum has also been demonstrated to be a local trigger generator and also mediates re-entry leading to AF.[114] This release of intracellular Ca++ is dynamically regulated by the frequency of open state of RyR2. The probability of open state of RyR2 is modulated by the accessory binding proteins (FK506 binding protein 12.6, calmodulin, sorcin) as well as posttranslational modifications (especially phosphorylation). Ca++/calmodulin-dependent protein kinase II (CaMKII) is an important mediator of phosphorylation of RyR2. The phosphorylated form of RyR2 facilitates an open state of RyR2 and thereby governs the abnormal Ca++ release and re-entry driven AF.[115] Azimilide Azimilide is a class III multichannel blocker which has been demonstrated to have electrophysiological effects on the repolarization phase of cardiac APD. At varying concentrations, it was observed to have inhibitory action on I Kr, IKs and I KACh.[116,117] Azimilide showed initial promising results in suppressing AF in the four randomized, placebo-controlled clinical trials [referred to as the Supraventricular Arrhythmia Study (SVA 1–4)].[118] However, the subsequent studies such as the Azimilide-CardiOversion Maintenance Trial-II (A-COMETII)[119] and the clinical trial by Page et al.[120] did not show a significant benefit of azimilide in suppressing AF. Considering its lack of beneficialfindings inAFprevention, the future development of azimilide has been discontinued. JTV-519/K201/Multichannel Blocker JTV-519/K201 is a multichannel blocker which inhibits the activity of I Kr and I KACh [121] and also stabilizes RyR2.[122] In a pre-clinical study by Nakaya et al.,[123] JTV-519 was observed to decrease the prolongation in APD induced experimentally by carbachol. The addition of JTV-519 to the guinea-pig atrial cells in this study also increased the carbachol-induced shortening in ERP. Both these mechanisms contribute to the efficacy of JTV-519 in mitigating the perpetuation of AF.[123] The antiarrhythmic potential of JTV-519 was further demonstrated on the activity of pulmonary veins in isolated cardiomyocytes by Chen et al..[124] This was mediated by its inhibition of I CaL and NCX currents in the pulmonary veins [125] ( ). Table 5. Summary of various investigational agents, respective targets and their developmental status Investigational agent Mode of action/target channels Development status Comments Multi-channel blocker JTV-519/K201 (Aetas Pharma) [122–125] Ranolazine [128, 129] Multi-channel blocker, also Phase II stabilizes RyRs. Blocks I Kr Phase II study has been completed Believed to have antiarrhythmic action by INa blockade Currently being investigated in combination with dronedarone in the HARMONY trial Phase II Atrial-selective blocker AVE-0118 (Sanofiaventis) Kv 1.5 blocker, also blocks Phase I [126] IKur Non-selective blocker of Kv 1.5, also blocks other Kv channels XEN D 0101 (Xention/Galapagos NV) [127] Kv 1.5 blocker Phase I Atrial-selective blocker of Kv 1.5, prolongs atrial APD, lack of effect on QTc NyK 1001 (Nyken) [156, 157] hsp 70 inducer Phase II Promotes anti-apoptotic and anti-inflammatory actions of hsp ISIS-CRPRx (Isis Pharmaceuticals) [159] C-reactive protein inhibitor Phase II Anti-inflammatory action Potentially anti-inflammatory in action, could have relevant role in post-operative AF Mechanism remains investigational 2015-04-29 08:05 11 iš 22 BMS 914392 Unknown mechanism (Bristol-Myers Squibb) [175] Phase II Currently being investigated for pharmacokinetics properties and drug interaction with metoprolol and diltiazem F 373280 (Pierre Fabre) [176] Unknown mechanism Phase II Being investigated for maintenance of SR after electric cardioversion of patients with persistent AF and cardiac failure OPC-108459 (Otsuka Pharmaceutical) [177] Unknown mechanism Phase II Currently being investigated in the CADENCE 215 study for its efficacy in paroxysmal and persistent AF patients AF atrial fibrillation, APD action potential duration, hsp heat shock protein, QTc corrected QT interval, RyRs ryanodine receptors, SR sinus rhythm Atrial-selective Agents An important limitation of multichannel blockers for the treatment of AF is the risk of ventricular arrhythmias. This limitation has necessitated the development of ''atrialselective'' agents with a promising efficacy and a better safety profile. Blockage of I KACh and I Kur has been considered to be a promising atrial-selective approach for the management of AF. AVE-0118 is a potent inhibitor of I KACh and prolongs the ERP, which contributes to its efficacy in suppressing AF. Since I KACh is constitutively expressed in AF and also tends to be atrial selective, AVE-0118 offers the benefit of being a selective target drug without affecting ventricular depolarization.[126] XEN D 0101 is another ''atrial-selective'' agent which increases the APD by its inhibition of K v 1.5 and I Kur.[127] Both these agents are in phase I of their development. Recently, there has been a renewed interest in the role of ranolazine as an antiarrhythmic agent for AF suppression. Ranolazine, although a multichannel blocker, has also been shown to be a potent inhibitor of the activity of I Na in the atrial myocytes.[128] The clinical efficacy of ranolazine as a primary agent as well as in combination with dronedarone for treatment of paroxysmal AF is being investigated in the HARMONY (a study to evaluate the effect of ranolazine and dronaderone when given alone and in combination in patients with paroxysmal atrial fibrillation) trial.[129] Receptor for Advanced Glycation End Products (RAGE) The role of receptor for advanced glycation end products (RAGE) is implicated in the pathogenesis of AF inDM. Both the presences of DM as well as the degree of glycemic control in diabetes have been shown to be relevant risk factors for the pathogenesis of AF.[130,131] There is emerging evidence that the activation of the advanced glycation end products (AGE)-RAGE axis has a relevant role in oxidative stress and inflammation-mediated pathogenesis of AF.[132] In a study by Kato et al.,[133] the atrial tissue from diabetic rats revealed a remarkably increased degree of fibrosis and an increased expression of RAGE. In this study, the development of atrial fibrosis was further accompanied by upregulation of CTGF, suggesting its role as a mediator of atrial fibrosis. Another study, by Candido et al.,[134] also found that atrial tissue from diabetic rats demonstrated increased production of AGE with increased expression of RAGE and RAGE-3; these changes were also accompanied by upregulation in the gene and protein expression of CTGF. The profibrotic role of AGE was further supported by the AGE-RAGE axis-mediated upregulation of MMPs production and alteration in the cross-linking of type 1 collagen and laminin. [135–138] AGE-RAGE interaction has also been demonstrated to have a pro-inflammatory role by activation of pleiotropic transcription factor NF- B.[139] Data from the animal studies, epidemiological studies and various clinical trials suggest that inflammation plays a pivotal role in the pathophysiology of AF,[140,141] and there is also interesting evidence that inflammation could also partly be mediated by the arrhythmia itself.[142] The link between the activation of AGE-RAGE axis is particularly relevant as a therapeutic target for antioxidant molecules. Yamagishi et al.[143] demonstrated that N-acetylcysteine (NAC) was found to inhibit the AGE-induced apoptosis, which in turn was mediated by AGE-induced upregulation of p53 protein in a mesangial cell-based model. Clinical Studies Ozaydin et al.[144] investigated the role of NAC in a double-blind prospective trial in 115 patients undergoing coronary artery bypass grafting (CABG) and/or valve surgery who were randomized to receive either placebo or NAC; the patients in NAC arm had a significantly lower incidence of post-operative AF (5.2 %) as compared with the placebo arm (21.1 %, p = 0.019). A meta-analysis of eight randomized controlled trials including 578 patients also demonstrated beneficial effects of NAC in reducing the incidence of post-operative AF (OR 0.62, 95 % CI 0.41–0.93, p = 0.021) as compared with controls.[145] Oxidative Stress Pathway and Targets in AF Oxidative stress is another key area of interest that appears to have a relevant role in the pathogenesis of AF.[141] Mihm et al.[146] 2015-04-29 08:05 12 iš 22 were some of the first investigators to observe a significantly greater degree of oxidative damage in AF as compared with SR in the patients who underwent cardiovascular operations. The results from this study also identified myofibrillar creatine kinase (MM-CK) to be a potential target of oxidative injury, with decreased activity levels in AF as compared with SR. Further results of this study also demonstrated higher levels of 3-nitrotyrosine (peroxynitrite biomarker) and protein carbonyl (hydroxy radical biomarker), implicating their role in oxidation-mediated injury in the atrial tissue of AF patients. These findings were further supported by the study of Kim et al.,[147] which investigated transcriptional profiles from human atrial tissue obtained from AF patients undergoing Maze procedure. They found an increased expression of genes associated with the production of ROS, which was accompanied by a simultaneous decreased expression of genes related to antioxidant protection (glutathione peroxidase 1 and heme oxygenase 2). These results supporting the role of increased oxidative damage in the atrium of patients with AF were further supported by Lenaerts et al.,[148] who reported an increased NADPH oxidase-mediated superoxide production in a sheep model of persistent AF. These changes were also paralleled by increased mRNA expression of the NADPH regulator protein Rac1, suggesting its role as a key mediator for increased NAPH oxidase activity.[148] Based on the mechanistic link between oxidative stress and AF, the role of antioxidants has been investigated for both preventive and treatment purposes, particularly in the setting of post-operative AF. A protective role of vitamin C on atrial remodeling has been observed to be mediated by its potency to decrease the production of peroxynitrite[149] and attenuation of the oxidative effects of peroxide on the atrial tissue.[150] Statins have also been investigated for their additional role as antioxidants in suppression of AF which is mediated by oxidative stress. Yagi et al.[151] demonstrated that treatment with pitavastatin was protective against Ang II-mediated atrial superoxide production and rac1-mediated activation of NADPH oxidase. Similar to the pleiotropic actions of statins, polyunsaturated fatty acids (PUFAs) have also been demonstrated to have multiple antiarrhythmic effects. PUFAs have been demonstrated to counteract the arrhythmogenic effects of atrial stretch by regulating membrane fluidity and modulating the properties of various membrane proteins.[152] PUFAs have also been demonstrated to have direct electrophysiological effects on various ion channels [153,154] and a regulatory effect on MAP kinase function.[155] Heat shock protein 70, C-reactive protein (CRP) and adenosine are some other relevant mediators of therapeutic interest in inflammation-mediated AF. NyK 1001, which is an inducer of hsp 70, is currently being investigated for its anti-inflammatory properties,[156,157] Increased levels of CRP after catheter ablation have been found to be associated with a greater recurrence of AF.[158] ISIS-CRPRx is currently being investigated as a novel CRP inhibitor in the phase II studies.[159] Increased release of adenosine has also been found to be implicated in the pathogenesis of post-operative AF by shortening of atrial ERP. Trabodenoson is being investigated as an antagonist of these arrhythmogenic effects of adenosine.[160] Clinical Studies Currently available clinical data on the efficacy of vitamin C and E as antioxidants is limited because of the absence of larger randomized controlled trials and are mostly based on observations from smaller retrospective studies. In a relatively recent meta-analysis of five randomized controlled trials incorporating 567 patients, Harling et al.[161] showed that the prophylactic use of vitamin C and E prior to cardiac surgery significantly reduced the incidence of post-operative AF (OR 0.43, 95 % CI 0.21–0.89). The role of statins in the prevention of post-operative AF was supported by the finding from the Atrial Fibrillation Suppression Trials (AFIST I–III). On the basis of a multivariate analysis of patients enrolled in these studies, the use of statins was found to be associated with a reduction in post-operative AF (adjusted OR 0.60, 95 % CI 0.37–0.99), and this effect was suggested to be dose dependent (adjusted OR 0.45, 95 % CI 0.21–0.99 for a dose of atorvastatin 40 mg).[162] These findings were further supported by the results of the Atorvastatin for Reduction of MYocardial Dysrhythmia After (ARMYDA-3) trial based on 200 patients without prior history of AF and statin intake undergoing elective cardiac surgery who were randomized to receive statins or placebo for 7 days prior to the procedure. After a follow-up of 30 days, the patients in the atorvastatin arm had a reduced incidence of AF (35 vs. 57 %, p = 0.003) as compared with the placebo group.[163] Current clinical evidence for the beneficial effects of PUFAs for prevention of AF remains limited and contradictory on the basis of available data. In a randomized controlled trial based on 160 patients, a 5-day treatment course of N-3 fatty acids prior to CABG and continued until the day of discharge was found to be associated with a lower incidence of new-onset AF (15.2 vs. 33.3 %, OR 0.35, 95 % CI 0.16–0.76, p = 0.013) as compared with controls.[164] These beneficial effect of PUFAs for preventing AF were not observed in the recently concluded larger, multicenter Omega-3 Fatty Acids for Prevention of Post-operative Atrial Fibrillation (OPERA)[165] and Omega-3 Fatty Acid for Inhibition of Supraventricular Arrhythmia (FISH) trials.[166] The lack of efficacy of PUFAs for prevention of AF was further substantiated by the results of the OM8 (efficacy and safety of prescription omega-3 fatty acids for the prevention of recurrent symptomatic atrial fibrillation: a randomized controlled trial) trial, which compared a regimen of omega-3 fatty acids (prescription omega-3 at 8 g/day for a week, followed by 4 g/day thereafter until 24 weeks) with placebo in reducing the symptomatic recurrence of AF in paroxysmal and persistent AF patients. After a follow-up interval of 24 weeks, the use of omega-3 fatty acids was not observed to reduce the study end point (HR 1.22, 95 % CI 0.98–1.52, p = 0.08) in both paroxysmal and persistent AF patients.[167] Connexins as Molecular Targets 2015-04-29 08:05 13 iš 22 Connexins are ubiquitous proteins which have also generated a great degree of interest as mediators of electrical remodeling. Each connexin consists of four -helical transmembrane domains (TMI–TM4), two extracellular loops (EL1 and EL2), a cytoplasmic loop (CL) between TM2 and TM3, and cytoplasmic amino-terminal (NT) and carboxy-terminal (CT) domains.[168] Six connexins join together to form a hemichannel, which docks with a hemichannel on an adjacent cell to form a gap junction. Alteration in the expression of connexins leads to an impaired conduction across gap junctions and contributes to the development of AF.[169] Two connexin molecules, namely, Cx40 and Cx43, are believed to be the main mediators of conduction heterogeneity and disturbances as part of atrial electrical remodeling.[169–171] Cx40 is of particular interest in the pathogenesis of AF because of its selective expression in the atrium.[169] Early investigations by Bagwe et al.[172] reported altered atrial activation and increased P-wave duration in Cx40 knockout mice, and the authors speculated these changes in atrial conduction could be due to alteration of intercellular coupling and possibly secondary to the ion-channel independent effects of connexin alteration. In addition to reduced quantitative expression of Cx40, an alteration in regional distribution and lateralization of Cx40 was also found to be associated with regional heterogeneity in atrial conduction and an increased tendency to develop re-entry mediated AF.[173] Almeida et al.[174] observed increased upregulation of Cx40 mRNA in a mouse model with hyperthyroidism, which was further associated with increased atrial depolarization velocity and an upregulation of Cx40 expression suggesting that Cx40 could be a relevant mediator of interest in the pathogenesis of AF in the setting of hyperthyroidism. Some other agents which are also being investigated for their potential efficacy for suppressing AF are BMS 914392,[175] F373280[176] and OPC-108459.[177] Currently, the mechanism of these agents remains to be elucidated. Future Directions Limitations in the current drug- and catheter ablation-based therapies for AF have sparked great interest in identification of various molecular pathways mediating structural and electrical atrial remodeling leading to AF. Future research efforts will help clarify the relationship between the degree of atrial fibrosis, conduction heterogeneity and predisposition to develop AF. Further data from trials investigating ACE and aldosterone inhibition as ''upstream'' therapies will yield useful information regarding the use of these agents for prevention of AF. In the coming years, further pre-clinical studies will also help uncover new pathways of potential interest. Research in the field of miRNA is rapidly evolving and is of considerable interest with respect to a biomarker-based diagnostic approach in the patients with AF; pre-clinical studies supplemented with data from clinical trials will help advance the role of miRNAs from ''bench to bedside.'' A better understanding of oxidative stress, ion-channel remodeling and alterations of connexins will not only help identify the relevance of these individual pathways in the pathogenesis of AF, but will also help uncover key interrelationships between these pathways that ultimately result in the substrates responsible for driving AF. Sidebar Key Points Limitations of current therapeutic options for atrial fibrillation (AF) necessitate research and development of newer agents. Modification of atrial fibrosis has generated considerable interest as a potential target for several agents as ''upstream therapies''. Atrial selective ion channel blockers are also of potential as promising agents for suppression of AF without an increased risk of development of ventricular arrhythmias. This review discusses the role of various therapeutic agents in development as novel agents in the management of AF. References 1. Benjamin EJ, Chen PS, Bild DE, et al. 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Acknowledgements Supported in part by The Deane Institute for Integrative Research in Atrial Fibrillation at Massachusetts General Hospital. 2015-04-29 08:05 22 iš 22 Am J Cardiovasc Drugs. 2014;14(6):403-421. © 2014 Adis Springer International Publishing AG 2015-04-29 08:05