Biology subjects

Kapiloff, M. S.

Publications and source records attributed to Kapiloff, M. S..

5 recordsLinked to original sources

bioRxiv2026

Gene therapy targeting of AKAP6β-CaMKII signalosomes improves myocardial inflammation and heart failure in a swine model of cardiometabolic syndrome

BackgroundCardiometabolic heart failure with preserved ejection fraction (HFpEF) is associated with systemic and cardiac inflammation and diastolic dysfunction. A-kinase anchoring protein 6{beta} (AKAP6{beta}) is a scaffold protein located at the cardiomyocyte outer nuclear membrane that promotes pathological cardiac remodeling via the recruitment of multiple regulatory proteins including protein kinases. In mice, adeno-associated virus (AAV) mediated expression of a peptide based upon a kinase binding domain (KBD) within AKAP6{beta} inhibited the development of heart failure due to chronic pressure overload. Whether KBD expression can also inhibit the development of cardiometabolic heart failure is unknown, and if so, the mechanism of KBD action in HFpEF has yet to be explored. MethodsThe efficacy of a cardiotropic self-complementary AAV gene therapy that expresses the AKAP6{beta} KBD peptide (AAV9sc.KBD) was tested in a female Ossabaw swine model of cardiometabolic syndrome and HFpEF. Single nucleus and bulk RNA sequencing of swine heart tissue and immunoprecipitation-mass spectrometry, live cell imaging, and biochemical assays using primary rat cardiomyocytes were employed to study KBD mechanism of action. ResultsAAV9sc.KBD inhibited the development of diastolic dysfunction and heart failure in the Ossabaw model, without negatively impacting systolic function. The improvement in cardiac phenotype was associated with decreased T-cell myocardial infiltrates and partial reversal of pathological gene expression. An unbiased interactome study revealed that the KBD peptide binds Ca2+/calmodulin-dependent protein kinase II (CaMKII), identifying CaMKII as a new AKAP6{beta} binding partner. Perinuclear CaMKII activity detected by live cell imaging required AKAP6{beta} expression and was inhibited by KBD expression. In addition, the CaMKII substrate Inhibitor of NF-{kappa}B Kinase {beta} (IKK{beta}) bound AKAP6{beta}. IKK phosphorylation in the Ossabaw model and in myocytes was inhibited by KBD expression, and NF-{kappa}B nuclear translocation in myocytes was dependent upon AKAP6{beta}-CaMKII protein complex formation. AAV9sc.KBD treatment inhibited cardiomyocyte NF-{kappa}B-dependent gene expression in the Ossabaw model. ConclusionsRegulated by perinuclear AKAP6{beta}-CaMKII signalosomes, NF-{kappa}B pro-inflammatory gene expression in cardiomyocytes participates in a positive feedback loop with cardiac inflammation promoting HFpEF. Proof-of-concept is provided in a large animal model that gene therapy-based cardiomyocyte expression of the KBD peptide will prevent cardiac dysfunction in cardiometabolic syndrome. Clinical PerspectiveO_ST_ABSWhat is newC_ST_ABSO_LIThe cardiomyocyte-selective gene therapy AAV9sc.KBD, which targets signalosomes organized by the scaffold protein AKAP6{beta}, is shown to inhibit myocardial T-cell infiltration and improve cardiac structure and function in a large animal model of cardiometabolic HFpEF. C_LIO_LIThe AKAP6{beta} KBD peptide is shown to bind and inhibit the function of CaMKII. C_LIO_LICaMKII and IKK{beta} are shown to participate in perinuclear AKAP6{beta} signalosomes, where they regulate activation of the NF-{kappa}B pro-inflammatory gene regulatory pathway. C_LI Clinical implicationsO_LIProof-of-concept for a novel strategy for the treatment of HFpEF is provided, intracellular expression by a cardiomyocyte-selective gene therapy vector of an inhibitory peptide, which will inhibit compartmentalized intracellular signal transduction. C_LIO_LIIn conjunction with previous studies in small rodents, the new data obtained in Ossabaw swine support clinical translation of the AAV9sc.KBD gene therapy. C_LI

physiology↗

bioRxiv2026

Regulation of the Balance between Concentric and Eccentric Cardiac Hypertrophy by a CDC14A-KMT5A Signaling Pathway

BackgroundDepending upon the type of pathological stress, the heart undergoes concentric or eccentric remodeling. This structural change is associated with diastolic and/or systolic ventricular dysfunction reflecting differentially altered cardiomyocyte morphology, ultrastructure, metabolism, contractility, and survival, as well as interstitial myocardial fibrosis. Despite an association of both concentric and eccentric remodeling with heart failure and sudden death, the molecular mechanisms resulting in abnormal cardiac geometry remain poorly understood. A better understanding of the basic mechanisms conferring these contrasting forms of remodeling should inform novel approaches to preserve normal cardiac structure and function in cardiovascular disease. The protein phosphatase Cell Division Cycle 14A (CDC14A) and its substrate the lysine methyltransferase KMT5A are identified herein as key regulators of the balance between concentric and eccentric pathological cardiac remodeling. MethodsThe regulation of adult rat ventricular myocyte morphology by CDC14A and KMT5A was studied in vitro following gain and loss of function by expression of wild-type and mutant proteins and RNA interference (RNAi). Epigenomic regulation by KMT5A was studied by mapping histone 4 lysine 20 mono-methylation (H4K20me1) modified chromatin sites and correlating them with gene transcription. Regulation of pathological cardiac remodeling in vivo was demonstrated by CDC14A and KMT5A RNAi using adeno-associated virus (AAV) mediated cardiomyocyte-specific small hairpin RNA (shRNA) expression in mice. ResultsCDC14A inhibited the growth in width of cultured adult myocytes stimulated by -adrenergic receptor activation or by serum response factor. KMT5A was downregulated by CDC14A in cardiomyocytes and was required for myocyte growth in width. -adrenergic stimulation of KMT5A-dependent H4K20 mono-methylation across transcription units correlated with regulation of gene transcription. Accordingly, AAV-expressed KMT5A shRNA induced eccentric remodeling and cardiac dysfunction in wild-type mice. Conversely, expression of Cdc14A shRNA improved systolic function and cardiac structure and inhibited pathological gene expression in the Tpm1 E54K mouse with Dilated Cardiomyopathy. ConclusionsCDC14A-KMT5A-dependent epigenomic regulation of gene transcription constitutes a molecular switch that determines concentric versus eccentric cardiac remodeling. These findings identify CDC14A as a potential therapeutic target for the treatment of dilated cardiomyopathy and other forms of heart failure with reduced ejection fraction. Clinical PerspectiveO_ST_ABSWhat is newC_ST_ABSO_LIA function is identified for the first time for the protein phosphatase CDC14A in the heart, regulation of cardiomyocyte morphology and overall cardiac geometry in pathological cardiac remodeling. C_LIO_LIThe lysine methyltransferase KMT5A is shown to mediate the effects of CDC14A in the adult cardiomyocyte by regulating H4K20 mono-methylation, such that reduced KMT5A expression promotes a phenotype resembling Dilated Cardiomyopathy. C_LIO_LIH4K20me1 epigenomic modification is identified as a regulator of cardiac structure and function. C_LI Clinical implicationsO_LICDC14A loss of function experimentation in vivo, resulting in improved cardiac structure and function in a mouse model of Dilated Cardiomyopathy, suggests that CDC14A is a novel therapeutic target for heart failure with reduced ejection fraction. C_LI

cell biology↗

bioRxiv2025

Targeting of CIP4-Calcineurin Signalosomes Improves Cardiac Structure and Function After Myocardial Infarction

BackgroundCalcineurin in a pleiotropic signaling enzyme that promotes pathological cardiac remodeling but also cardioprotection in ischemia-reperfusion injury. In addition, calcineurin inhibitors are immunosuppressants. This pleiotropy has precluded the use of calcineurin inhibitors as treatments for heart failure. Cdc42-interacting protein 4 (CIP4/TRIP10) is an endosomal scaffold protein that organizes a calcium and calcineurin A{beta}2 (CaNA{beta}2) signaling compartment activated by G-protein coupled receptors independently of contractile calcium. CIP4 binds CaNA{beta}2 via the CaNA{beta}-specific N-terminal polyproline (PP) domain. We previously showed that targeting of CIP4-CaNA{beta}2 signalosomes inhibited pathological cardiac hypertrophy and the development of heart failure induced by chronic pressure overload in mice. It is unknown whether CIP4-CaNA{beta}2 signalosomes contribute to cardioprotection and/or cardiac remodeling in ischemic heart disease. MethodsCIP4 conditional knock-out (CKO) mice were studied by echocardiography with strain analysis and histology following ischemia-reperfusion (I/R) injury and permanent left coronary artery (LCA) ligation to induce myocardial infarction. Wildtype C57BL/6NJ mice were transduced with adeno-associated virus (AAV) engineered for cardiomyocyte-specific expression of either a CaNA{beta}2 shRNA to inhibit CaNA{beta}2 expression, a VIVIT peptide to inhibit CaN-NFAT signaling, or a CaNA{beta}2 PP peptide to block CIP4-CaNA{beta}2 binding. AAV-transduced mice were studied by I/R injury. Additional mice were subjected to permanent LCA ligation and subsequently treated with AAV to test the effects of CaN inhibition in chronic ischemic cardiomyopathy. The effects of CaNA{beta}2 PP-GFP expression on primary T-cell activation were studied in vitro. ResultsCIP4 CKO mice and mice expressing the PP anchoring disruptor peptide exhibited preserved cardiac function after I/R injury and decreased infarct size and preserved cardiac function 8 weeks after myocardial infarction by permanent LCA ligation. In contrast, cardiomyocyte-specific depletion of CaNA{beta}2 and VIVIT peptide expression worsened outcome after I/R injury and in chronic ischemic cardiomyopathy. In addition, in contrast to cardiomyocytes, PP-mediated CaNA{beta} anchoring inhibition had no effect on T-cell activation and cytokine expression in vitro. ConclusionsCIP4-CaNA{beta}2 signalosomes promote adverse cardiac remodeling and are not cardioprotective. Proof-of-concept is provided for the treatment of ischemic cardiomyopathy by a PP anchoring disruptor gene therapy. Targeting these complexes may be beneficial in cardiovascular diseases, including ischemic cardiomyopathy and acute myocardial infarction. Clinical PerspectiveO_ST_ABSWhat is New?C_ST_ABSO_LITargeting CIP4, which is a scaffold protein for the phosphatase calcineurin, improves cardiac function in mice after acute myocardial infarction due to ischemia-reperfusion injury and in chronic ischemic cardiomyopathy. C_LIO_LIGene therapy-based expression of a calcineurin A{beta}-derived polyproline peptide, which can compete CIP4-calcineurin binding, is beneficial in acute and chronic myocardial infarction. C_LI What Are the Clinical Implications?O_LIThis study establishes CIP4 signalosomes as a new drug target for the treatment of ischemia-reperfusion injury and chronic pathological cardiac remodeling. C_LIO_LIThis study provides proof-of-concept for a new gene therapy approach to treating acute myocardial infarction and chronic ischemic cardiomyopathy. C_LI

physiology↗

bioRxiv2024

The Unique Role of Intracellular Perinuclear β-Adrenergic Receptors in defining Signaling Compartmentation and Pathological Cardiac Remodeling

The {beta}-adrenergic receptor is a prototypical G-protein coupled receptor that initiates signaling from the plasma membrane. However, active receptors have been detected within intracellular compartments. The functional significance of these intracellular receptors remains unclear, including whether they regulate distinct cellular processes or function independently of plasma membrane receptors. We show using live cell imaging of primary cardiomyocytes that {beta}-adrenergic receptors localized to Golgi apparatus opposing the outer nuclear membrane are sufficient and necessary for the stimulation of cAMP and calcium signaling within a nanometer scale compartment independent of receptors at other sites. Using compartment-specific activators and inhibitors, we show Golgi {beta}-adrenergic receptors associated with the scaffold protein AKAP6{beta} and the outer nuclear membrane protein nesprin-1 are responsible for pathological gene transcription and the induction of cardiomyocyte hypertrophy. The functional significance of Golgi-localized receptors is demonstrated in mice models of cardiomyopathy, providing proof-of-concept for a compartment-specific therapeutic intervention in Dilated Cardiomyopathy.

cell biology↗

bioRxiv2021

FGF21-FGFR4 signaling in cardiac myocytes promotes concentric cardiac hypertrophy in mouse models of diabetes

Fibroblast growth factor (FGF) 21, a hormone that increases insulin sensitivity, has shown promise as a therapeutic agent to improve metabolic dysregulation. Here we report that FGF21 directly targets cardiac myocytes by binding {beta}-klotho and FGF receptor (FGFR) 4. In combination with high glucose, FGF21 induces cardiac myocyte growth in width mediated by extracellular signal-regulated kinase 1/2 (ERK1/2) signaling. While short-term FGF21 elevation can be cardio-protective, we find that in type 2 diabetes (T2D) in mice, where serum FGF21 levels are elevated, FGFR4 activation induces concentric cardiac hypertrophy. As T2D patients are at risk for heart failure with preserved ejection fraction (HFpEF), we propose that induction of concentric hypertrophy by elevated FGF21-FGFR4 signaling constitutes a novel mechanism promoting T2D-associated HFpEF and that FGFR4 blockade might serve as a cardio-protective therapy in T2D. In addition, potential adverse cardiac effects of FGF21 mimetics currently in clinical trials should be investigated.

molecular biology↗