Targeting MuRF1 to Combat Skeletal Muscle Wasting in Cardiac Cachexia: Mechanisms and Therapeutic Prospects

Introduction

Chronic heart failure (CHF) is a multifactorial clinical syndrome characterized by structural or functional abnormalities of the heart [1]. In its terminal stage, CHF can lead to cardiac cachexia, a condition exacerbated by multi-organ dysfunction and disordered metabolism. The term “cachexia” is derived from the Greek “kakós,” meaning bad, and “hexis,” meaning condition or appearance, which literally means “a bad condition” (Figure 1). Cardiac cachexia was first reported in 1860 by the French physician Charles Mauriac. It is now recognized as an indicator of poor prognosis and diminished quality of life in patients experiencing end-stage heart failure [2]. Cardiac cachexia affects 5–15% of individuals with CHF [3].

The clinical manifestations of cardiac cachexia include severe edema, weight loss of at least 5% irrespective of initial body weight, or a body mass index (BMI) of less than 20 kg/m2 over a period of 12 months. Patients may also experience fatigue, reduced muscle mass and strength, dyspnea, anorexia, anemia, and abnormal blood biomarker levels such as elevated CRP (C-reactive protein) (>5.0 mg/l), increased IL-6 (interleukin-6) (>4.0 pg/ml), low hemoglobin (<12 g/dl), and/or decreased serum albumin (<3.2 g/dl) [4–6] (Figure 1). In developed countries and regions like the United States, Europe, and Japan, the annual mortality rate for patients with cardiac cachexia is 20–40% [3]. Research by Anker et al demonstrated a stark contrast in mortality rates: 17% for heart failure patient without cachexia versus 50% for those with cachexia after an 18-month follow-up. Their study, employing a Cox proportional hazards model for survival analysis, further corroborated cardiac cachexia as an independent predictor of mortality in CHF patients [7].

Cachexia often occurs secondary to severe conditions such as sepsis, severe trauma, and many chronic systemic inflammatory diseases, such as chronic obstructive pulmonary disease (COPD), CHF, chronic renal failure, cancer, type 2 diabetics, and rheumatoid arthritis [8]. In cachexia, the rate of catabolism exceeds anabolism, leading primarily to a reduction in body mass dominated by skeletal muscle wasting [9]. Skeletal muscle wasting is characterized by a decline in muscle mass and dysfunction. Patients with cachexia frequently experience a reduced capacity for exercise, disability, weakness, and fatigue, which adversely affect their quality of life and increase the risk of mortality and healthcare costs [10–13]. Preserving skeletal muscle mass and function when healthy may improve outcomes for patients with cardiac cachexia. While research continues into various nutritional supplements and medications to prevent muscle wasting associated with diseases, exercise training remains the only clinically-approved intervention that is both safe and effective. However, it is not recommended for patients with severely limited capacity for exercise, compromised heart function, or those who are bedridden or immobilized for extended periods [8,14,15].

Skeletal muscle atrophy primarily results from an imbalance between protein synthesis and degradation. The ubiquitin-proteasome-system (UPS), autophagy-lysosome system, and apoptosis are implicated in increased protein breakdown, with UPS playing a predominant role in the disassembly of myofibrils [8,10,16,17]. Bodine et al identified 2 critical atrophy-related genes, Trim63 and Fbxo32, which are overexpressed in striated muscle under atrophic conditions and encode for Muscle RING-Finger 1 (MuRF1) and MAFbx/atrogin-1, respectively [18]. These genes are upregulated in various atrophy scenarios, including fasting [19,20], exposure to glucocorticoids [21,22], denervation [23,24], hindlimb suspension [25], and immobilization [26,27]. The transcription of atrogenes Trim63 and Fbxo32 is under the regulation of NF-κB, Foxo, and SMAD signaling pathways, with several inhibitors of these pathways demonstrated to indirectly reduce MuRF1 and MAFbx expression [28–30].

Evidence has mounted that MAFbx plays a role in the myocardial hypertrophy triggered by the NF-κB pathway [31]. As a result, suppressing MAFbx activity may alleviate the symptoms of pathological hypertrophy and could even help in reversing the course of heart failure. Nonetheless, it remains a significant gap in the field that no studies have yet announced the development of small-molecule inhibitors designed to specifically target and suppress MAFbx. Therefore, this article emphasizes the introduction of MuRF1-targeting inhibitors. Recent research involving MuRF1-targeting inhibitors, such as MyoMed-205, has shown promise in mitigating skeletal muscle wasting and partially reversing myocardial fibrosis in a rat model of heart failure with preserved ejection fraction (HFpEF) over a 32-week period. Targeting these pathways could therefore be beneficial for treating skeletal muscle wasting associated with cardiac cachexia [32].

Pathophysiological Mechanism of Cardiac Cachexia

Patients with CHF experience a cascade of pathophysiological changes, including significant fluctuations in neurohormonal, metabolic hormone levels, and heightened inflammatory responses. These alterations precipitate a catabolic state, characterized by reduced muscle and adipose tissue mass, bone density, and mineral content. Concurrently, these patients often have loss of appetite, increased feelings of fullness, and gastrointestinal dysfunction. This persistent catabolic and depletion state culminates in cardiac cachexia, which clinically manifests as frailty, fatigue, weight loss, and diminished physical strength (Figure 2).

Alterations in Hormone Levels

ANG-II (ANGIOTENSIN-II):

The enhanced release of Ang-II, a primary component of the renin–angiotensin–aldosterone system (RAAS), occurs in response to increased sympathetic activity. This elevation contributes to muscle wasting, diminished appetite, and weight loss upon receptor engagement [33]. Ang-II orchestrates the balance of protein synthesis and degradation by boosting NADPH oxidase activity, resulting in elevated reactive oxygen species (ROS) production [33]. Its influence extends to the hypothalamic appetite regulation centers, where it modulates the secretion of neuropeptide-Y (NPY), orexin, and corticotropin-releasing hormone (CRH), thereby reducing appetite [35].

GHRELIN-GROWTH HORMONE (GH)-AXIS:

Ghrelin, a natural agonist for the growth hormone secretagogue receptor (GHS-R) [36], is found in higher concentrations in patients with cardiac cachexia compared to those with CHF but without cachexia [37]. This hormone stimulates secretion of growth hormone (GH), which predominantly promotes muscle growth through the insulin-like growth factor-1 (IGF-1) signaling pathway [38]. However, research indicates that patients with CHF may develop ghrelin resistance due to elevated levels of this “hunger hormone” during catabolic states, alongside GH resistance, which is characterized by increased GH levels but decreased IGF-1 [36,39]. This hormonal imbalance is associated with loss of muscle mass and contributes to progression of cardiac cachexia in CHF patients.

ADIPOKINES – ADIPONECTIN AND LEPTIN:

Abnormal lipid and glucose metabolism may stem from increased levels of adiponectin and a relatively higher concentration of leptin compared to healthy individuals – likely due to weight loss and reduced adipose tissue [40–42]. Doehner et al documented that heightened leptin levels in CHF patients, irrespective of cachexia status, can contribute to decreased insulin sensitivity and development of insulin resistance [43,44]. Moreover, this increase in leptin can also amplify feelings of fullness, diminish appetite, and reduce nutrient intake in individuals with cardiac cachexia [45].

TESTOSTERONE:

The catabolic state in cardiac cachexia is marked by diminished testosterone and elevated cortisol levels, which further intensify muscle wasting, inflammatory responses, and oxidative stress [5,46,47].

Excessive Inflammatory Responses

In the setting of cardiac cachexia, levels of inflammatory mediators, particularly TNF-α, IL-1 and IL-6, are significantly elevated [48]. These cytokines are predominantly released by monocytes and macrophages. Additionally, cardiomyocytes contribute to cytokine release due to hypoxic conditions induced by CHF, which serves as a primary pathway for release of these key inflammatory agents [49]. An increase in neurohormonal secretion, observed in CHF patients both with and without cardiac cachexia, further augments the release of inflammatory factors, including cortisol, catecholamines, and RAAS-related hormones such as aldosterone and Ang-II [48,50]. Notably, TNF-α concentrations are reported to be significantly higher in patients with cardiac cachexia compared to those without [51], and research by Anker et al has identified an association between TNF-α levels and weight loss in these patients [52]. Previous studies have demonstrated that TNF-α, IL-1, and IL-6 are critical upstream triggers for UPS activity, leading to increased protein degradation and skeletal muscle wasting [48]. Furthermore, the adverse effects of TNF-α on vascular endothelial cell function and capillary density reduction impede blood supply to the skeletal muscles, thereby disrupting energy metabolism and nutritional status, and exacerbating the progression of cardiac cachexia [5,40,53].

Unbalanced Protein Synthesis and Degradation

INHIBITION OF PROTEIN SYNTHESIS:

Both IGF-1 and insulin stimulate protein synthesis via the PI3K-Akt-mTOR pathway. However, in the catabolic state characteristic of CHF, protein synthesis is impeded due to diminished levels of circulating IGF-1 and prevalent insulin resistance in patients [38,54,55]. Ang-II exacerbates this condition by inhibiting the release of IGF-1 into the bloodstream, further hindering protein synthesis, as noted by Sukhanov [56]. Moreover, Breitbart et al found that myostatin, a potent inhibitor of muscle growth, is secreted into the circulation by the heart during failure, playing a pivotal role in the muscle wasting seen in cardiac cachexia [57]. Morissette et al reported that the inhibitory effect of myostatin on protein synthesis could be due to its interference with IGF-1-mediated phosphorylation of Akt, also known as protein kinase B, as a result of increased myostatin level [58].

ACCELERATION OF PROTEIN DEGRADATION:

In vertebrates, protein catabolism is predominantly facilitated by 3 key systems: the UPS, the autophagic lysosomal system, and the caspase-dependent apoptotic system.

UBIQUITIN-PROTEASOME SYSTEM (UPS):

UPS is hyperactivated by atrophic stimuli, leading to breakdown of key skeletal muscle proteins, including myosin, myosin binding protein C (MyBP-C), and α-actin [59]. The ubiquitination process involves a series of enzyme-catalyzed reactions in which the ubiquitin-activating enzyme (E1) forms high-energy thioester bonds between cysteine residues of E1 and the C-terminal glycine residues of ubiquitin. This activated ubiquitin is then conveyed to the ubiquitin-conjugating enzyme (E2), creating an E2-ubiquitin complex. Following this, the ubiquitin ligase (E3) facilitates the transfer of ubiquitin to lysine residues on the target protein, generating polyubiquitin chains. These polyubiquitinated proteins are then recognized and disassembled by the 26S proteasome complex [60].

In CHF patients, a cascade of pathophysiological changes alters levels of various hormones and inflammatory factors, which can upregulate atrophy-related genes such as Trim63 and Fbxo32. This upregulation enhances protein degradation while suppressing synthesis, ultimately leading to skeletal muscle atrophy. Ang-II, for instance, indirectly influences the UPS by generating an excess of ROS, which in turn activates the NF-κB signaling pathway [56]. Elevated levels of inflammatory cytokines like TNF-α, IL-1, and IL-6 can increase the transcription of atrogenes encoding muscle-specific E3 ligases, such as MuRF1 and MAFbx/atrogin-1, via NF-κB pathway, [59,61] and AMPK-Foxo pathway [62]. The suppression of Akt-mediated phosphorylation of Foxo transcription factor also contributes to the enhanced transcription of MuRF1 and MAFbx/atrogin-1 in cachexia [54,63]. Myostatin plays a dual role by promoting the transcription of Trim63/MuRF1 and Fbxo32 and proteolysis through the activin A receptor type IIB (ActRIIB) and the subsequent phosphorylation of SMAD2/3 in both skeletal and cardiac muscles [10]; it also inhibits muscle growth by preventing satellite cell proliferation and differentiation [64]. Furthermore, increased glucocorticoid receptor engagement also contributes to upregulation of Trim63 and Fbxo32, leading to intensified protein degradation [65].

AUTOPHAGY-LYSOSOME SYSTEM (ALS):

Concurrent with cachexia, the maintenance of skeletal muscle protein homeostasis involves cathepsin L-dependent degradation processes [66]. Within skeletal myocytes, the ALS and UPS are activated by signaling pathways such as AMPK, as well as Foxo3 and Foxo1. An accumulation of autophagy signals under catabolic conditions is associated with ensuing muscle atrophy [38,67].

APOPTOSIS:

Cardiac cachexia is associated not only with increased apoptosis in cardiac myocytes [68], but also, as Adams et al have identified [69], with heightened apoptosis in the skeletal muscles of over half of patients with congestive heart failure. This condition correlates with a reduction in exercise endurance. During this state, cells encounter intrinsic apoptotic stimuli, leading to mitochondrial damage and the release of cytochrome c mediated by the BCL-2 protein family. Concurrently, there is an upregulation of pro-apoptotic BAX (BCL2-Associated X) and a downregulation of anti-apoptotic BCL-2. These alterations trigger a chain reaction that promotes apoptosis through activation of caspase-9 and caspase-3 [69–71].

Structure and Functions of MuRF1

STRUCTURE:

E3 ubiquitin ligases are categorized into 3 types based on their structural domains: those with a RING-finger domain, those homologous to E6-associated protein C terminus (HECT), and those with an in-between RING (RING-in-Between-RING) domain that exhibit characteristics of both RING-finger and HECT-type E3s [73]. MuRF1, a RING-finger type E3 within the RING-B-box-Coiled-Coil (RBCC) family – also known as the Tripartite Motif (Trim) family [74] – serves as a connector, facilitating the transfer of ubiquitin from E2 enzymes to target proteins without direct ubiquitin binding [73].

Centner et al were the first to suggest MuRF1’s interaction with titin at the M-line and Z-line of muscle sarcomeres, regulating titin’s stability [75]. MuRF1 is predominantly found in striated muscles, including skeletal [18] and cardiac muscles, and it is also present in certain smooth muscles, potentially playing a role in postpartum uterine atrophy [76].

In skeletal muscles, MuRF1 selectively binds to and targets substrates such as myosin heavy chain (MyHC) [77,78], myosin light chain (MyLC) [79], myosin binding protein C (MyBP-C) [80], α-actin [81], and titin-cap protein (Titin-cap, or TCAP, or telethonin) [82] for ubiquitination and subsequent degradation. In cardiac myocytes, MuRF1’s localization includes the M-line and specific sequences within titin [83]. It targets MyHC [84], MyBP-C [85], and troponin I for ubiquitination [86].

FUNCTIONS:

The initial step in understanding MuRF1’s role in skeletal and cardiac muscle tissues is to manipulate the expression of its encoding gene, Trim63. This research is crucial for laying the groundwork for development of inhibitors targeting MuRF1. Studies on MuRF1-KO mice have demonstrated protection against muscle wasting across different atrophy models regardless of the muscle fiber type. Bodine et al found that MuRF1-KO mice retained 36% more muscle mass in the gastrocnemius after 14 days of denervation compared to wild-type mice [18]. In atrophy models induced by amino acid deprivation, Koyama et al observed enhanced protein synthesis in the quadriceps muscles of MuRF1-KO [87]. Labeit et al reported almost complete resistance to atrophy in the soleus muscles of MuRF1-KO mice after 10 days in a hindlimb suspension model [88]. Additionally, Baehr et al discovered that MuRF1-KO mice were resistant to glucocorticoid (dexamethasone)-induced muscle atrophy, maintaining muscle fiber cross-sectional area (CSA) over 2 weeks [21].

MuRF1 plays a multifaceted protective role in cardiac muscle tissue, distinct from its function in skeletal muscle. Its contributions to myocardial health are evident in 4 main areas: (1) Regulation of pathological myocardial remodeling: MuRF1-KO mice exhibit more pronounced cardiac hypertrophy post-Transverse Aortic Constriction (TAC) compared to wild-type mice, indicating that MuRF1 can mitigate compensatory myocardial changes during early heart failure and thus improve prognosis [89,90]. MuRF1 appears to suppress protein synthesis linked to hypertrophy, as evidenced by the exacerbation of left ventricular hypertrophy following pressure overload in MuRF1-KO mice [91]. Molecular pathways through which MuRF1 exerts this effect include downregulation of the CnA (Calcineurin A)-NFAT pathway, inhibition of the JNK/AP-1 signaling pathway, and interactions with serum response factor (SRF) that modulate hypertrophic gene expression [90,92–94]. (2) Association with hypertrophic cardiomyopathy: Chen et al sequenced the Trim63 gene in 302 hypertrophic cardiomyopathy patients and identified mutations that cause mislocalization of MuRF1 in cardiomyocytes, impair its ubiquitination activities, and lead to myocardial hypertrophy [84]. (3) Response to drug-induced myocardial atrophy: In MuRF1-KO mice, myocardial atrophy induced by drugs like dexmedetomidine (a2-adrenoceptor agonists), dexamethasone (glucocorticoids), and Adriamycin (antibiotic) was not observed to be beneficial [89,90,95]. (4) Protection against myocardial ischemia-reperfusion (I/R) injury: Overexpression of MuRF1 may protect against I/R damage by reducing cardiac myocyte apoptosis through the inhibition of the JNK signaling pathway and downstream AP-1 signaling [96].

Development of Related Inhibitors Targeting MuRF1

P013222:

Eddins et al were the pioneers in identifying the MuRF1 inhibitor P013222, first reported in 2011 [104]. Utilizing an E3 ligase screening platform, they assessed a diverse library of small molecules for MuRF1 activity inhibition and found that P013222 impeded MuRF1’s autoubiquitination in a dose-dependent manner. Furthermore, the inhibitor was shown to protect MyHC from degradation in dexamethasone-treated C2C12 myotubes, suggesting its ability to block MuRF1-dependent substrate ubiquitination. Despite these promising in vitro results, in vivo and clinical trial data for P013222 are not yet available.

ID#704946:

Bowen et al developed a small-molecule compound, ID#704946, which targets the MuRF1-Titin complex [105]. In myotube culture experiments, this compound exhibited minimal toxicity and effectively protected myotubes from atrophy induced by synthetic glucocorticoids such as dexamethasone. Animal trials involved C57/BL6 mice, which were divided into 3 groups to establish a cardiac cachexia model via weekly injections of 600 mg/kg monocrotaline (MCT), leading to right ventricular insufficiency due to pulmonary hypertension. The experimental group received the inhibitor starting 1 week before MCT administration. After 6 weeks, the mice were euthanized for skeletal muscle analysis. Tissue studies revealed that ID#704946 specifically inhibited the upregulation of MuRF1 expression induced by cardiac cachexia without affecting MAFbx/atrogin-1 expression. Furthermore, it impeded proteasome activity and the ubiquitination of contractile proteins, reducing actin depletion. In a rat model of cardiac cachexia, the compound significantly mitigated contractile dysfunction in the tibialis anterior (TA) and diaphragm muscles, although it did not positively affect weight loss or ventricular hypertrophy development.

In continued research with compound ID#709496, Bowen et al [106] addressed diaphragm myopathy and systolic dysfunction induced by CHF. They induced CHF in female C57/BL6 mice by ligating the anterior descending branch of the left coronary artery to stimulate myocardial infarction. The mice that subsequently developed diaphragm malfunction were divided into 2 groups: one receiving normal feed and the other receiving feed supplemented with the compound, in addition to a sham-operated group with normal feeding. By week 10, the experimental group exhibited signs of diaphragmatic disease, marked by a 21% decrease in cross-sectional area (CSA) and diminished maximum diaphragmatic contractility. Treatment with the compound significantly reversed the impairment in diaphragmatic contractility, although it did not have a notable effect on CSA.

The aforementioned studies not only underscored ID#704946’s effectiveness in preventing skeletal muscle wasting but also explored its mechanisms of action: (1) In the skeletal muscle of control mice, an increase in cardiac ankyrin repeat protein (CARP) expression was observed, a protein associated with the structural domain where MuRF1 acts on Titin and is present in both cardiac and skeletal muscle tissues [107]. Typically inactive, CARP overexpression is induced by pathogenic stimuli and is associated with muscular atrophy [108]. The experimental results suggest that ID#704946 can inhibit CARP overexpression, which is linked to MuRF1 activity in cardiac cachexia [105]. (2) Additionally, the studies proposed that ID#704946 maintains protein synthesis and reduces apoptosis in skeletal muscle cells, with an upregulation of ELF2B (E47-like factor 2) and downregulation of BAX being observed. However, further research is needed to verify the specificity of these effects in skeletal muscle [105]. (3) A subsequent study hypothesized that ID#704946 can affect mitochondrial metabolic balance [106]. Molecular analyses indicated that the activity and concentration of key mitochondrial enzymes – such as citrate synthase (CS), succinodehydrogenase (SUH), and oxidative respiratory chain complex I (Complex I) – were significantly reduced in CHF conditions. Treatment with ID#704946 improved these mitochondrial changes, suggesting that MuRF1 acts as a negative regulator of mitochondrial metabolic functions. While the study confirmed the beneficial impact on mitochondrial, the precise mechanisms by which ID#704946 alters mitochondrial metabolism were not extensively investigated. (4) Transcriptome analysis of C2C12 myotubes treated with ID#704946 revealed a decrease in MuRF1 expression by 4–10%, further supporting the compound’s potential role as a MuRF1 inhibitor [106].

In conclusion, further investigation is essential to clarify the precise mechanisms behind ID#704946’s therapeutic actions. This includes determining whether its efficacy stems from a suitable reduction in MuRF1 expression and activity, the disruption of interactions with target proteins, or broader effects such as those on mitochondrial metabolism, protein synthesis, and apoptosis. Additionally, the compound’s potential regulatory impact on MuRF2 and the possibility of functional redundancy should be validated through further studies. Clinical trials for ID#704946 have not been conducted, as there is insufficient clarity regarding its bioavailability, metabolic system toxicity, and impact on cardiac remodeling, and there is a lack of comprehensive data from animal models.

MYOMED-946 AND MYOMED-205:

Building on the success of compound ID#709496 in mitigating skeletal muscle wasting associated with cardiac cachexia [105] and diaphragm dysfunction in CHF [106], Adams et al synthesized 2 new compounds: MyoMed-946, which is structurally identical to ID#709496 but produced via a different synthesis method, and MyoMed-205, a chemically modified variant of ID#709496 designed to enhance serum stability [109]. These compounds were independently administered to mice with advanced melanoma to assess their impact on skeletal muscle function. Both MyoMed-946 and MyoMed-205 were reported to reduce weight loss in the TA, soleus, and extensor digitorum longus (EDL) muscles, indicating an improvement in muscle wasting conditions. Additionally, the mice showed protection against tumor-induced weight loss. Further analysis of EDL muscle tissue revealed that treatment with the inhibitors increased the expression of SMDT1, a calcium-regulatory subunit of the mitochondrial single transporter, potentially affecting mitochondrial apoptosis via the BAX pathway [109–111]. The compounds were also shown to prevent depletion of citrate synthase and maintain Complex I enzyme activity, suggesting that their beneficial effects on skeletal muscle wasting are due to a combination of mechanisms involving apoptosis regulation, mitochondrial energy metabolism, and MuRF1 inhibition.

In their subsequent research, Adams et al extended their investigation to the impact of MyoMed-205 on various skeletal muscle wasting disorders and cardiac function in HFpEF rats [32]. They divided 30 ZSF1 obese rats into 2 groups: one group received MyoMed-205 and the other served as a placebo group, with healthy mice acting as controls. After feeding the rats until 20 32 weeks to progressively induce the HFpEF model, the following conclusions were drawn from comprehensive tissue studies: (1) MyoMed-205 significantly reduced skeletal muscle atrophy to normal levels, particularly in the TA muscle, where it also increased muscle mass and CSA by 26%. The treatment led to decreased ubiquitination of total muscle proteins and a reduction in MuRF1 expression in skeletal muscle. (2) The compound mitigated mitochondrial stress effects from heart failure, as shown by proteomic analysis, which indicated differential expression of 8 proteins in the placebo group’s muscle tissue, of which 5 are related to mitochondrial metabolism and autophagy. (3) Treatment with MyoMed-205 resulted in an upregulation of protein synthesis for mitochondrial Complex I–V and significantly higher citrate synthase activity in the TA muscle, supporting the notion that MuRF1 affects mitochondrial oxidative phosphorylation, as demonstrated by a MuRF1-KO model. (4) MyoMed-205 caused greater improvement in skeletal muscle wasting compared to ID#704946, potentially attributable to its enhanced serum stability.

Recently, the research team led by Ribeiro has utilized MyoMed-205 in the therapeutic intervention of contractile impairments after unilateral phrenic nerve denervation, reaffirming the muscle-protective role of MuRF1 inhibitors concerning atrophy and muscle fiber cross-sectional dimensions. Expanding on this, they meticulously crafted experiments to assess the toxicity and dose-response profile of this MuRF1-targeting small molecule, with no adverse toxic effects noted within the dosage spectrum of 300 to 2000 mg/kg. Upon subjecting the diaphragm to denervation for 12 hours, a range of MyoMed-205 dosages (from 12.5 to 250 mg/kg body weight) was applied. Notably, the 50 mg/kg body weight dosage of MyoMed-205 demonstrated significant prevention of diaphragmatic contractile dysfunction, while higher dosages, such as 100 and 150 mg/kg body weight, provided partial amelioration of strength degradation. Conversely, the maximum dosage of 250 mg/kg body weight failed to manifest any protective impact on diaphragmatic strength. Western blotting analysis revealed that MyoMed-205, through the modulation of muscle atrophy-associated factors (HDAC4, FoxO1, and MuRF2) coupled with the activation of the Akt signaling cascade, confers a protective influence on the early-stage diaphragmatic dysfunction and atrophy [112].

Discussion

Effective treatment for cardiac cachexia with skeletal muscle wasting should encompass a multimodal approach, including pharmaceutical therapy, exercise training, and nutritional counseling. Recent advances in MuRF1 research have led to development of several small-molecule inhibitors targeted at this protein, such as ID#709496, MyoMed-946, and MyoMed-205, created by Adams’ team. These inhibitors have demonstrated potential in reducing skeletal muscle wasting in cachectic states.

Notably, In their investigation of the HFpEF rat model, Adams et al [32] examined the direct effects of MyoMed-205 on myocardial diastolic function. The compound decreased the severity of left ventricular sclerosis and improved cardiac diastolic function in the experimental rats, suggesting a potential reversal of myocardial alterations in HFpEF. Myocardial fibrosis, characterized by an excessive accumulation of extracellular matrix, plays a crucial role in myocardial remodeling in HFpEF. This process is regulated by an imbalance between matrix metalloproteinases (MMPs) and their tissue inhibitors, with MMP activation leading to reduced tensile strength of cardiac tissue and consequent diastolic dysfunction [113–116]. Treatment with MyoMed-205 resulted in reduced myocardial fibrosis in the experimental group compared to the placebo group, as indicated by lower MMP levels in protein blotting assays [32]. Furthermore, previous studies have linked low phosphorylation levels of Titin and an altered collagen-Titin ratio with the development of the HFpEF myocardium [117,118]. Adams et al found that in obese HFpEF mice, myocardial Titin phosphorylation levels and myocardial tissue collagen were low, although the experimental group did not show significant differences from the control group in myocardial phosphorylation levels [32]. They hypothesized that MyoMed-205 maintains myosin flexibility by promoting cGMP-PK (cyclic guanosine monophosphate-protein kinase G) phosphorylation modifications, a hypothesis that warrants further research. In summary, the study suggests that the beneficial effects of MyoMed-205, a small molecule targeting HFpEF, on cardiac diastolic function may be linked to reduced myocardial fibrosis and modifications in actin phosphorylation. The proposed mechanisms involve inhibition of MuRF1 expression, which reduces myogenic fibronectin degradation in the UPS pathway, balancing protein synthesis and degradation to prevent muscle wasting. Additionally, modulating actin and reducing fibrosis may benefit cardiac muscle during HFpEF.

The striated muscle groups in cardiac and skeletal muscle tissues are not the same, and MuRF1 performs distinct functions and has different domains of action in these 2 muscles, as previously described. Although MuRF1 plays a part in the atrophy of skeletal muscle, its function in cardiac muscle seems to be more complex. Further research into MuRF1’s function and mechanism in cardiac tissue is needed, particularly its influence on myocardial remodeling, ischemia-reperfusion injury, and myocardial hypertrophy [84,119,120].

Developing MuRF1-targeting inhibitors has several challenges: (1) There is a lack of comprehensive data from animal models and clinical trials, despite the demonstrated efficacy of these inhibitors in treating muscle wasting. (2) All inhibitors have not shown concentration-related negative effects, which could be due to the short duration of studies and their dissimilarity from typical clinical treatment courses. (3) The impact of MuRF1 suppression on the development of myocardial hypertrophy, a condition that worsens heart failure, is yet to be determined. Furthermore, MuRF1 plays a protective role against myocardial ischemia-reperfusion injury [96], raising concerns about potential exacerbation of reperfusion injury if MuRF1 expression is reduced. (4) Incomplete pharmacological, toxicological, and pharmacokinetic studies have hindered the progression and clinical application of MuRF1 inhibitors. (5) There are various subtypes within the MuRF protein family. In normal conditions, MuRF1 and MuRF2 co-knockout animals display cardiac and skeletal muscle hypertrophy [121], while MuRF3-KO mice do not show a distinct phenotype under ordinary conditions but have increased susceptibility to heart rupture after myocardial infarction. Co-knockout of MuRF1 and MuRF3 results in an abnormal accumulation of MyHC fragments in skeletal and cardiac muscle [122]. This suggests that MuRF1 and other family members share functions, and due to MuRF1’s complex transcriptional regulatory system, the development of inhibitors must be refined to enhance specificity towards MuRF1.

Despite the current absence of effective pharmacological treatments for patients with skeletal muscle wasting in cardiac cachexia, the identification of MuRF1-targeting inhibitors has opened up new therapeutic possibilities. While these inhibitors have limitations, their role in treating skeletal muscle wasting is promising, and using them as a novel target is currently a viable concept. Concurrently, establishing reliable diagnostic criteria for cardiac cachexia with skeletal muscle wasting is essential. A proactive approach in preventing skeletal muscle wasting in CHF patients, with a focus on MuRF1 as a key target, could potentially improve both the prognosis and the quality of life for these individuals. Traditional Chinese medicine has also explored this class of E3 ligases. Compounds such as ginsenoside Rg1 have shown potential to suppress MuRF1 expression in C2C12 cells [123], and formulations like the heated kidney and blood activation formula have improved cardiac function and exercise tolerance while reducing MAFbx/atrogin-1 and MuRF1 mRNA expression in heart failure models [124]. The E3 ligase encoded by the atrophy gene MAFbx/atrogin-1, which shares a similar transcriptional signaling pathway with MuRF1 [17,125–127], is another potential target against skeletal muscle wasting and myocardial injury. Investigating atrogenes like MuRF1/Trim63, MAFbx/atrogin-1/Fbxo32, and their homologs [128] histologically will not only elucidate their tissue action mechanisms but will also assist in developing new biomarkers for diagnosis, risk stratification, prognosis assessment, and monitoring treatment efficacy.