The therapeutic potential of heat shock proteins in cardiomyopathies due to mutations in cardiac structural proteins

Journal of Molecular and Cellular Cardiology 49 (2010) 904–907

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Journal of Molecular and Cellular Cardiology j o u r n a l h o m e p a g e : w w w. e l s e v i e r. c o m / l o c a t e / y j m c c

Editorial

The therapeutic potential of heat shock proteins in cardiomyopathies due to mutations in cardiac structural proteins

Over the past 12 years, a new group of cardiomyopathies have emerged from the recognition of the myofibrillar myopathies (MFM). The myofibrillar myopathies (MFM) are a group of morphologically similar muscular dystrophies characterized by the accumulation of abnormal proteins that make up critical structural components of muscle including desmin, dystrophin, myotilin, ubiquitin, sarcoglycans, and (co-) chaperone proteins such as αB-crystallin (CryAB), heat shock protein 27, and DNAJB2 [1–8]. Mutations in at least 7 proteins cause MFMs, including desmin [9], myotilin [10], ZASP [3], Trp2710X [5], Bag3 [7], FHL1 [11–13], and αB-crystallin [14]. The myofibillar myopathies were first described in 1998 with the identification of disease associated with mutations in desmin (see Table 1). What the myofibrillar myopathies have in common is their association with severe cardiomyopathies and variable skeletal muscle involvement which presents in both childhood and in adults. Since only a relatively few number of patients with MFMs have been reported to date, the prevalence of these diseases is not entirely clear. Mutations in desmin and αB-crystallin (CryAB) have been the most studied of the myofibrillar myopathies and are sub-categorized as desmin-related myopathies (DRM). Desmin is expressed in cardiac, skeletal muscle, and smooth muscle and is the main intermediate filament that forms a continuous cytoskeletal structure between the contractile apparatus, the cell membrane, and the nucleus. It is more abundant in the heart (2%) compared to skeletal muscle (0.35%) [22]. Desmin is prominent in the specialized conduction system of the heart, including the Purkinje fibers that regulate coordinated cardiac contraction [22]. CryAB is a chaperone protein that interacts with desmin to prevent its aggregation during stress [23]. Therefore, it is not surprising that mutations in desmin and CryAB result in significant cardiac impairment given the critical role of the structural proteins in the heart. The cardiac phenotype of DRM is variable and includes restrictive cardiomyopathy, atrioventricular block, arrhythmias, and sudden death [24,25]. Presently, there is no specific treatment for DRMs, although premature death can be prevented by identifying lifethreatening arrhythmias and conduction defects and treating appropriately [25–27]. In the present issue of the Journal of Molecular and Cellular Cardiology, Zhang et al. [55] tested the hypothesis that enhancing the expression of the small heat shock protein HSPB1 (aka Hsp27) would enhance the cardiomyocyte's ability to handle mis-folded CryAB mutant proteins. To do this, they tested three distinct CryAB mutations: R120G, 450delA, and 464delCT in cardiac-derived cells. Despite the common aggregation-prone nature of these mutations, their human disease phenotypes are distinct. The 450delA mutation is 0022-2828/$ – see front matter © 2010 Elsevier Ltd. All rights reserved. doi:10.1016/j.yjmcc.2010.09.022

generally restricted to the lens while the 464delCT and R120G affects the muscle and are associated with cardiomyopathy [28]. In the article by Zheng et al., the investigators identified that increasing HSPB1 expression significantly increased the solubility of the 464delCT mutations, while having negligible effects on the solubility of the R120G and the 450delA mutant proteins. Conversely, knocking down HSPB1 decreased the solubility of all three CryAB mutant proteins, indicating a role for endogenous HSPB1 in clearing these mutant proteins. Four lines of evidence are presented that the ubiquitin proteasome system is involved in the clearance of R120G, 450delA, and 464delCT via HSPB1. Pharmacologic proteasome inhibition increased the protein levels of all three CryAB mutant proteins, while increasing the expression of mutant-specific ubiquitin conjugates in vitro. Knocking down the expression of HSPB1 increased poly-ubiquitin conjugated proteins. Lastly, these studies identified that the attenuated 464delCT expression mediated by increasing HSPB1 could be prevented by proteasome inhibition. One role HSPB1 may play is in clearing specific mutant CryAB proteins, which may be due in part, to their ability to shuttle proteins to the ubiquitin proteasome system for rapid degradation, resulting in less accumulated toxic aggregated protein. The differences in how HSPB1 interacts with these different CryAB mutant proteins may also lie in the structural aspects of the mutant protein itself. In contrast to the missense R120G CryAB mutation, which does not differ in size to the wild type protein [14], the frameshift 450delA results in a elongated CryAB protein [29], while the 464delCT results in a truncated CryAB protein [1]. While the studies by Zhang et al. [55] did not delineate the specific physical interaction between the small heat shock protein HSPB1 and the ubiquitin proteasome system, cross talk between the two has previously been described. One of the more clear examples is CHIP (co-chaperone carboxyl terminus of the HSC70-interacting protein), which has both co-chaperone activity with its interaction with HSC70 and its own ubiquitin ligase activity [30,31]. During normal wear and tear that occurs in proteins, this duo of CHIP and HSC70 mediate substrate refolding. If the protein damage is too much and refolding is not possible, the duo recognizes this and ubiquitinates the damaged/misfolded protein to target it for degradation by the proteasome [25]. In the presence of mainly misfolded proteins, as in the case of mutant proteins, a similar complex made up of small heat shock proteins and ubiquitin ligases may rapidly target the mutant proteins for rapid degradation by the ubiquitin proteasome system, as has been described in other cardiomyopathies [32,33]. It is likely that HSPB1 may similarly bridge the gap between the attempt to refold mutant proteins and

Editorial

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Table 1 Protein mutations giving rise to myofibrillar myopathies. First described

Cardiac, other phenotype

References

Desmin

1998

[9,15]

αB-crystallin (CryAB)

2003

Myotilin Filamin

2000 2005

ZASP

2005

Bag3

2009

FHL1

2008

Arrhythomogenic cardiomyopathy common, distal, limb-girdle, scapuloperoneal distribution of muscle weakness. Most patients present at ages 10-61 with syncopal issues. Hypertrophic cardiomyopathy, symmetric proximal and distal muscle weakness, atrophy and respiratory involvement (Cataracts). Affected patients reported in adulthood. Cardiomyopathy, distal myopathy, peripheral neuropathy (= Limb-girdle muscular dystrophy) Cardiomyopathy, respiratory insufficiency, peripheral neuropathy, The age of onset has been reported to be 24-60 (~ 5 families). Cardiac involvement without signs of CAD, predates muscle involvement in some cases, distal myopathy. The mean age of onset is the 6th decade. Cardiomyopathy, severe muscle weakness, diaphragm paralysis by 2nd decade. Three patients described presented in childhood. Heart failure, slowly progressive muscle weakness and wasting initially in a humeroperoneal muscle distribution, extending to scapular and pelvic girdle muscles. Heart palpitations, presyncope, syncope, and poor exercise tolerance may also occur due to arrhythmias and conduction defects. Age of onset generally in childhood.

interacting with ubiquitin ligases, yet to be identified, to target misfolded proteins for degradation. Several questions remain to be answered in the Zhang et al. [55] studies. First, does the enhanced clearance of the 464delCT by increasing HSPB1 improve the cardiomyocyte performance in vivo? This is an anticipated outcome, given the toxicity of some mutant proteins in the heart. Also, does the apparent lack of improvement in solubility of the R120G CryAB mutant proteins with increased HSPB1 have no effect on cardiac function? One recent publication suggests that increasing small heat shock proteins may improve R120G CryAB function experimentally in vivo. In this study, R120G cardiac transgenic mice were treated with geranylgeranylacetone (GGA), a potent inducer of HSPs, including HSPB8 and HSPB1 [34]. Treatment with GGA was reported to reduce amyloid oligomer levels and aggregates, improve cardiac hypertrophy and interstitial fibrosis, in addition to improving cardiac function and survival [34]. It remains to be determined if specific heat shock proteins interact with specific CryAB mutations, and if increasing one or more of the heat shock proteins may more uniformly improve cardiac function, protein aggregation, and long term survival in vivo. The mechanisms by which accumulating aggregated proteins impair cardiac function have begun to be delineated, mainly in mouse models of R120G CryAB. In mice with R120G CryAB, a greater than two fold increase in autophagic activity was identified [35]. Autophagy is the cellular process responsible for the removal or turnover of long lived proteins and organelles [36]. Autophagy may also play a role in clearing the aggregates formed in desminopathies as well. Experimentally inhibiting autophagy increased protein aggregates and decreased the time to heart failure onset, increased interstitial fibrosis, and the presence of poly-ubiquitinated proteins [35]. Apoptosis is increased in R120G CryAB transgenic hearts and plays a role in the dilated cardiomyopathy and heart failure that ensues [37]. In additional studies, R120G CryAB transgenic mice were crossed to mice with sustained cardiac overexpression of the anti-apoptotic Bcl-2 [37]. Increasing Bcl-2 prolonged the survival of R120G CryAB mice by 20% [37]. A decrease in mitochondrial abnormalities, restoration of cardiac function, and a prevention of cardiac hypertrophy were also identified when apoptosis was attenuated [37]. Surprisingly, the protein aggregation was significantly reduced when apoptosis was inhibited in R120G CryAB mice with the Bcl-2 transgene [37]. However, with the reduction of apoptosis came the enhancement of autophagy and an increase in the activation of alternative death pathways resulting in necrosis [37]. These findings suggest that R120G CryAB mutations enhance autophagy and apoptosis, whereby there is cross talk between autophagy signaling, apoptosis, and necrosis. The universality of these mechanisms to desmin-related cardiomyopathies

[1,14,16] [10,17] [5,18] [3,19,20] [7] [11,21]

has not been determined. Nor has the role of these mechanisms in human disease. Investigators are just beginning to recognize desmin-related cardiomyopathies, including those caused by mutations in CryAB, in a relatively few patients. Conceptually, however, the underlying pathophysiology shown in the article by Zhang et al. [55] may possibly be applicable to a wider array of more common cardiomyopathies, which are also due to mutations in structural sarcomere proteins. However, this is currently an untested hypothesis. These cardiomyopathies include familial hypertrophic cardiomyopathy (found in 1:500 people), dilated cardiomyopathy (found in 1:2500 people), and arrhythmogenic right ventricular cardiomyopathy (found in 1:5000 people) [38]. If we can determine how heat shock proteins are regulated and could be therapeutically modulated, this type of therapy may be useful in a number of related, fairly common, cardiomyopathies. The regulation of heat shock protein expression is by the HSF family of transcription factors (HSF1–4), which bind HSF binding elements to induce expression of many of the known heat shock proteins. In the heart, regulation of HSF1 and HSF2 have recently been described to increase in response to increased pre-load and mechanical stress, in addition to heart failure and ischemia reperfusion injury [39–43]. HSPB1 has been reported to be under the transcriptional control of HSF1 [44]. HSPB1 has been reported to accumulate in inclusion bodies in a number of conformational diseases, including models of amyotrophic lateral sclerosis [45]. Over the past two decades, the protective role of heat shock proteins in neurodegenerative diseases, including Alzheimer disease, Huntington's disease, and prion disease has been studied extensively. The pathophysiology of these diseases is primarily due to the excess amount of mis-folded proteins, much like the desmin-related cardiomyopathies, which leave the cell with an increasing amount of toxicity. At least four types of drugs, including herbal products, have been reported to increase heat shock proteins and could potentially be beneficial in patients with myofibrillar cardiomyopathies, although this has yet to be proven. These compounds include arimoclomol, celastrol, statins, and geranylgeranylacetone. Arimoclomol, a small molecule that induces HSF-1 and downstream HSP70 and HSP90, is currently in phase II/III clinical trials for treating amyotrophic lateral sclerosis (ALS) [46,47]. Celastrol is an extract from traditional Chinese medicine that potently induces HSF-1 and HSP70 to protect against the neurodegeneration in SOD1 mutant mice (due to misfolded protein accumulation) [47]. Among their other beneficial effects, both simvastatin and lovastatin induce HSPB1, HSF1, and HSP70 and HSP90 [48–52]. Lastly, GGA is a cyclic polyisoprenoid gastric ulcer drug that protects the intestine by inducing HSF-1 and HSP70 and has been shown to be cardioprotective in ischemia reperfusion injury experimentally, in part, by inducing HSP72 [53,54]. Recent studies have also shown that GGA

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Editorial

protects against the R120G mutation in CryAB experimentally, by inducing HSPB8 [34]. So given the novel findings by Zheng et al. for some types of CryAB mutations, the therapeutic realization of these findings may soon be possible should they have applicability to in vivo function and survival.

[25] [26]

[27]

Disclosures [28]

There are no potential conflicts of interest to disclose (J.E.R., M.S.W.).

[29]

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[30]

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Editorial against ischemia/reperfusion injury in rat heart. Circulation 2001 Oct 9;104(15): 1837–43. [54] Yamagami K, Yamamoto Y, Ishikawa Y, Yonezawa K, Toyokuni S, Yamaoka Y. Effects of geranyl–geranyl–acetone administration before heat shock preconditioning for conferring tolerance against ischemia–reperfusion injury in rat livers. J Lab Clin Med 2000 Jun;135(6):465–75. [55] Zhang H, et al. Selective degradation of aggregate-prone CryABmutants by HSPB1 is mediated by ubiquitin–proteasome pathways. J Mol Cell Cardiol in press. doi:10.1016/j.yjmcc.2010.09.004.

Jessica E. Rodríguez Department of Pathology & Laboratory Medicine, University of North Carolina, Chapel Hill, NC 27599-7525, USA

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Monte S. Willis Department of Pathology & Laboratory Medicine, University of North Carolina, Chapel Hill, NC 27599-7525, USA McAllister Heart Institute, University of North Carolina, Chapel Hill, NC 27599-7525, USA E-mail address: [email protected]. Corresponding author. 103 Mason Farm Road, 2340 Medical Biomolecular Research Building, Chapel Hill, NC 27599-7525, USA. Tel.: +1 919 843-1938; fax: +1 919 843 4585. 21 September 2010