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(Circulation. 2003;107:378.)
© 2003 American Heart Association, Inc.

Editorial

Treatment of Heart Failure Through Stabilization of the Cardiac Ryanodine Receptor

Gerd Hasenfuss, MD; Tim Seidler, MD

From the Department of Cardiology and Pneumology, University of Göttingen, Göttingen, Germany.

Correspondence to Gerd Hasenfuss, MD, University of Göttingen, Cardiology and Pneumology, Robert-Koch-Straße 40, 37075 Göttingen, Germany. E-mail hasenfus{at}med.uni-goettingen.de

Key Words: Editorials • heart failure • calcium • sarcoplasmic reticulum

The key event in cardiac excitation-contraction coupling is calcium-induced calcium release from the sarcoplasmic reticulum (SR) after activation of cardiac ryanodine receptors (RyRs).¹ Two distinct genes encode the cardiac (RyR2) and skeletal muscle (RyR1) specific receptors.^2,3 A third type of ryanodine receptor (RyR3) exhibits functional properties distinct from those of RyR1 and RyR2.⁴ The RyR channels form a tetrameric structure composed of 4 monomeric subunits, each of about 565 000 Daltons. These channels are about 10 times larger than the voltage-gated Ca²⁺ and Na⁺ channels.⁵ RyR2 opening increases cytosolic Ca²⁺ from approximately 100 nmol/L to 1 µmol/L with consecutive activation of contractile proteins. The RyR was recently shown to be a macromolecular complex including protein kinase A (PKA) and its anchoring protein (mAKAP), the protein phosphatases PP1 and PP2A, sorcin, calmodulin, FK506 binding protein (FKBP), and other proteins.⁵ FKBP12.0 and its isoform FKBP12.6 seem to play an important role for the regulation of the RyR. The immunophilin FKBP12 was originally identified as the cytosolic receptor for the immunosuppressant FK506 in T-cells. FKBP12 is a cis/trans peptidyl-prolyl isomerase. However, it is unlikely that this enzymatic activity is directly involved in RyR regulation. FKBP12 has been postulated to bind to the RyR with a stoichiometry of 4 or less FKBP12 molecules per single functional channel.⁶ FKBP12.0 is tightly bound to and regulates the function of the skeletal RyR1, whereas RyR2 has been found to co-purify selectively with FKBP12.6 despite a higher expression level of FKBP12.0 in cardiac myocytes.⁷ However, considerable species differences in binding preferences of FKBPs to RyR isoforms have been observed.⁸

See p 477

Marx et al⁹ recently suggested that phosphorylation of RyR2 dissociates the regulatory protein FKBP12.6 from the channel, resulting in pronounced altered channel function. First, activity of the channel (open probability) is increased, which results from a leftward shift in the Ca²⁺ dependence for channel activation. This increased sensitivity is associated with destabilization of the channel, resulting in subconductance states. Second, the dissociation of FKBP12.6 from the RyR functionally uncouples multiple RyRs and disturbs coupled gating. Coupled gating enables neighboring RyRs to open and close in a coordinated manner. Under uncoupling conditions, not all RyRs in a given region of the SR open together during systole and close together during diastole. Some RyRs remain open, resulting in a diastolic leak of Ca²⁺ from the SR.¹⁰

Marx et al⁹ furthermore reported that in failing human hearts, the RyR is hyperphosphorylated, resulting in dissociation of FKBP12.6 from RyR. Accordingly, increased Ca²⁺ sensitivity, subconductance states, uncoupled gating of RyRs, and, in consequence, SR Ca²⁺ leak were described. Hyperphosphorylation may result from decreased levels of phosphatases (PP1 and PP2A) associated with the RyRs. Reduced FKBP12.6 to RyR binding in the failing heart was confirmed by Ono et al¹¹ using [³H]dihydro-FK506–binding. They showed that in SR vesicles from dogs with pacing-induced heart failure, abundance of FKBP12.6 was decreased by 83% when compared with normal hearts.

The concept of hyperphosphorylation-induced RyR dysfunction in heart failure is still somewhat controversial. Recent studies in phospholamban knockout mice indicated that PKA-dependent RyR phosphorylation does not influence Ca²⁺ leak and SR Ca²⁺ release and that PKA-dependent effects on SR Ca²⁺ release result from phospholamban phosphorylation, with subsequently increased SR Ca²⁺.¹²

However, independent from the concept of RYR hyperphosphorylation in heart failure, there is considerable evidence that SR leak may be relevant for disturbed SR function in heart failure in addition to reduced SR Ca²⁺ uptake, and FKBPs are important regulators for SR Ca²⁺ handling. There is general agreement that disturbed SR function is the dominant mechanism that causes altered excitation-contraction coupling in human heart failure. It has been shown that frequency-dependent increase of SR Ca²⁺ content, as it is observed in non-failing human myocardium, is blunted or absent in the failing heart, which results in lower SR Ca²⁺ load and Ca²⁺ availability for systolic release.¹³ Quantitatively, the contribution of Ca²⁺ leak through RyRs to disturbed SR Ca²⁺ accumulation is unknown, and altered SR Ca²⁺ uptake may still be the major component. Ca²⁺ uptake in heart failure is reduced because of decreased SR Ca²⁺ ATPase activity due to decreased pump levels and reduced phospholamban phosphorylation. In addition, it has been suggested that increased Na⁺-Ca²⁺ exchanger activity may result in increased transsarcolemmal Ca²⁺ elimination and SR Ca²⁺ depletion. The relevance of FKBPs for SR Ca²⁺ handling has been demonstrated in a recent study. It was shown that overexpression of FKBP12.6 in rabbit myocytes results in reduced spontaneous SR Ca²⁺ leak, increased SR Ca²⁺ content, and increased isotonic shortening of isolated myocytes. These findings confirm the postulated effects of FKBP12.6 to reduce Ca²⁺ leak from the SR through the RyR and promote Ca²⁺ release during activation.¹⁴

Assuming that SR Ca²⁺ leak is relevant for the pathophysiology and progression of heart failure, restoration of RYR function could be an important target for the treatment of heart failure. A leak of Ca²⁺ from the SR decreases SR Ca²⁺ content and thus availability of systolic Ca²⁺ to be released; increased Ca²⁺ leak may promote diastolic dysfunction because of diastolic activation of contractile proteins; increased Ca²⁺ leak from the RyR may be a trigger for arrhythmias; altered cytosolic Ca²⁺ may contribute to altered gene expression and myocardial remodeling; and an increased leak would have unfavorable energetic consequences because ATP consumption for SR Ca²⁺ cycling would be increased for any given level of SR Ca²⁺ content. In this regard, reducing the leak seems to be advantageous over increasing SR Ca²⁺ uptake because increased SR Ca²⁺ content by itself increases Ca²⁺ leak in a nonlinear relation.¹⁵

In this issue of Circulation, Yano et al¹⁶ reported that the agent JTV519, a 1,4-benzothiazepine, restores disturbed RyR function and preserves contractile performance in hearts from dogs with 4 weeks rapid right ventricular pacing-induced heart failure.

In this heart failure model, alterations of excitation-contraction coupling, as previously analyzed by Yano et al, are similar to those reported by Marx et al⁹ in humans, ie, RYR hyperphosphorylation, decreased FKBP12.6 binding to RyRs, and increased SR leak. The authors suggest that JTV519 acts by exerting an FKBP12.6-like channel stabilizing effect either directly or by catalyzing re-association of FKBP12.6. The data presented support the hypothesis. It is shown that JTV519 reduces Ca²⁺ leak from SR vesicles of failing hearts. The drug is reported to restore binding of FKBP12.6 to RyR toward the stoichiometry seen in normal hearts. It decreases the effects of FK506 on FKBP12.6 dissociation from the RyR in a dose-dependent manner in immunoprecipitation experiments. Furthermore, it prevents cAMP-dependent reduction of FKBP-binding to RyRs. These effects were associated with preservation of FK506-induced Ca²⁺ leak in both normal and failing hearts.

The molecular mechanisms by which JTV519 may induce its actions are not completely understood. Obviously, there are no direct effects on RyR phosphorylation as shown by back-phosphorylation experiments. Interestingly, JTV519 as a benzothiazepine derivative, has structural similarities with the L-type Ca²⁺ channel blocker diltiazem and decreases L-type Ca²⁺ current by about 20%.¹⁷ Accordingly, diltiazem itself was found to partially inhibit SR Ca²⁺ leak. However, it can be excluded that the observed effect on SR leak results from L-type Ca²⁺ channel inhibition because measurements were predominantly performed in isolated vesicles. However, it cannot by excluded that part of the long-term effects on ventricular function (see below) may be due to L-type Ca²⁺ channel blockade or other reported effects of JTV519, such as influence on potassium currents¹⁸ or interaction with annexin V.¹⁹ On the other hand, the observed structural relationship to diltiazem may help to identify other molecules with even more specific effects to RyRs.

The chronic effects of intravenous JTV519 treatment are quite remarkable. When JTV519 was continuously infused by means of an infusion pump, it partially preserved systolic and diastolic function and prevented development of heart failure in chronically paced dogs. In addition, chronic administration of JTV519 reduced the levels of plasma norepinephrine, angiotensin II and atrial natriuretic peptide (ANP). Prevention of heart failure was associated with higher SR Ca²⁺ATPase levels, higher phospholamban phosphorylation, and thus increased SR Ca²⁺ uptake. In addition, PKA phosphorylation of RyR was lower compared with untreated pacing induced failing myocardium. The latter findings are most likely not primarily related to the effects of JTV519 on RyRs, and rather reflect prevention of left ventricular dysfunction and development of heart failure. Accordingly, in a recent article,²⁰ propranolol treatment of dogs during tachycardia pacing prevented development of heart failure and of SR Ca²⁺ leak. This was associated with restoration of FKBP12.6 binding to RyRs. Furthermore, it was shown that ß-blocker treatment reduced the level of PKA-dependent RyR phosphorylation and normalized single-channel RyR function in another study with pacing induced heart failure.²¹

In summary, there is cumulating evidence that increased leak of Ca²⁺ through RyRs is a significant component of altered excitation-contraction coupling in heart failure. Accordingly, reducing SR leak is a promising approach to improve Ca²⁺ cycling of the failing heart that theoretically should be advantageous over strategies to increase SR Ca²⁺ uptake. The present study¹⁶ is the first to show the feasibility of pharmacologically improving RyR function and SR leak in isolated vesicles and to show that long-term application of this agent is associated with improved ventricular function and prevention of heart failure in a dog model. However, it cannot be excluded that a variety of other unspecific effects of this drug contributed to its beneficial effects in heart failure animals, in particular because the molecular mechanism of its effects on RyRs is not completely understood. However, the present work should stimulate research to better understand the effects of JVT519 on SR function and, even more importantly, to search for other interventions to reduce SR leak and improve Ca²⁺ cycling in heart failure.

Footnotes

The opinions expressed in this editorial are not necessarily those of the editors or of the American Heart Association.

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