Pathological role of serum- and glucocorticoid-regulated kinase 1 in adverse ventricular remodeling.

BACKGROUND: Heart failure is a growing cause of morbidity and mortality. Cardiac phosphatidylinositol 3-kinase signaling promotes cardiomyocyte survival and function, but it is paradoxically activated in heart failure, suggesting that chronic activation of this pathway may become maladaptive. Here, we investigated the downstream phosphatidylinositol 3-kinase effector, serum- and glucocorticoid-regulated kinase-1 (SGK1), in heart failure and its complications. METHODS AND RESULTS: We found that cardiac SGK1 is activated in human and murine heart failure. We investigated the role of SGK1 in the heart by using cardiac-specific expression of constitutively active or dominant-negative SGK1. Cardiac-specific activation of SGK1 in mice increased mortality, cardiac dysfunction, and ventricular arrhythmias. The proarrhythmic effects of SGK1 were linked to biochemical and functional changes in the cardiac sodium channel and could be reversed by treatment with ranolazine, a blocker of the late sodium current. Conversely, cardiac-specific inhibition of SGK1 protected mice after hemodynamic stress from fibrosis, heart failure, and sodium channel alterations. CONCLUSIONS: SGK1 appears both necessary and sufficient for key features of adverse ventricular remodeling and may provide a novel therapeutic target in cardiac disease.

Head-to-Head Comparison of Tau-PET Radioligands for Imaging TDP-43 in Post-Mortem ALS Brain.

PURPOSE: In vivo detection of transactivation response element DNA binding protein-43 kDa (TDP-43) aggregates through positron emission tomography (PET) would impact the ability to successfully develop therapeutic interventions for a variety of neurodegenerative diseases, including amyotrophic lateral sclerosis (ALS).  The purpose of the present study is to evaluate the ability of six tau PET radioligands to bind to TDP-43 aggregates in post-mortem brain tissues from ALS patients. PROCEDURES: Herein, we report the first head-to-head evaluation of six tritium labeled isotopologs of tau-targeting PET radioligands, [3H]MK-6240 (a.k.a. florquinitau), [3H]Genentech Tau Probe-1 (GTP-1), [3H]JNJ-64326067(JNJ-067), [3H]CBD-2115, [3H]flortaucipir, and [3H]APN-1607, and their ability to bind to the ß-pleated sheet structures of aggregate TDP-43 in post-mortem ALS brain tissues by autoradiography and immunostaining methods. Post-mortem frontal cortex, motor cortex, and cerebellum tissues were evaluated, and binding intensity was aligned with areas of elevated phosphorylated tau (ptau), pTDP-43, and ß-amyloid. RESULTS: Negligible binding was observed with [3H]MK-6240, [3H]JNJ-067, and [3H]GTP-1. While [3H]CBD-2115 displayed marginal specific binding, this binding did not significantly correlate with the distribution of pTDP-43 and AT8 inclusions. Of the remaining ligands, the distribution of [3H]flortaucipir did not significantly correlate to pTDP-43 pathology; however, specific binding trends to a positive relationship with tau. Finally, [3H]APN-1607 relates most strongly to amyloid load and does not indicate pTDP-43 pathology as confirmed by [3H]PiB distribution in sister sections. CONCLUSIONS: Our results demonstrate the prominent nature of mixed pathology in ALS, and do not support the application of [3H]MK-6240, [3H]JNJ-067, [3H]GTP-1, [3H]CBD-2115, [3H]flortaucipir, or [3H]APN-1607 for selective imaging TDP-43 in ALS for clinical research with the currently available in vitro data. Identification of potent and selective radiotracers for TDP-43 remains an ongoing challenge.

Common variants for atrial fibrillation: results from genome-wide association studies.

Atrial fibrillation (AF) affects more than 5 million people worldwide; however, none of the anti-arrhythmic drugs available now are entirely optimal in terms of efficacy and safety. A better understanding of the molecular mechanism of AF will facilitate the process of finding new strategies to prevent AF. As the non-familial AF is the major form of AF, identifying common variants for AF in these populations by genome-wide association studies will definitely accelerate this process. This review summarizes the recently identified common AF variants on 4q25, 16q22, and 1q21 and discusses their implications for the clinic.

Characterization of neuroinflammatory positron emission tomography biomarkers in chronic traumatic encephalopathy.

Chronic traumatic encephalopathy is a neurological disorder associated with head trauma and is confirmed upon autopsy. PET imaging of chronic traumatic encephalopathy may provide a means to move towards ante-mortem diagnosis and therapeutic intervention following brain injuries. Characterization of the neuroinflammatory PET biomarkers, 18 kDa translocator protein and monoamine oxidase-B was conducted using [3H]PBR-28 and [3H]L-deprenyl, respectively, in post-mortem chronic traumatic encephalopathy brain tissue. [3H]PBR-28 displayed high specific binding in both chronic traumatic encephalopathy (95.40 ± 1.87%; n = 11 cases) and healthy controls (89.89 ± 8.52%, n = 3 cases). Cell-type expression of the 18 kDa translocator protein was confirmed by immunofluorescence to microglia, astrocyte and macrophage markers. [3H]L-deprenyl also displayed high specific binding in chronic traumatic encephalopathy (96.95 ± 1.43%; n = 12 cases) and healthy controls (93.24 ± 0.43%; n = 2 cases), with the distribution co-localized to astrocytes by immunofluorescence. Saturation analysis was performed to quantify the target density of the 18 kDa translocator protein and monoamine oxidase-B in both chronic traumatic encephalopathy and healthy control tissue. Using [3H]PBR-28, the target density of the 18 kDa translocator protein in healthy controls was 177.91 ± 56.96 nM (n = 7 cases; mean ± standard deviation); however, a highly variable target density (345.84 ± 372.42 nM; n = 11 cases; mean ± standard deviation) was measured in chronic traumatic encephalopathy. [3H]L-deprenyl quantified a monoamine oxidase-B target density of 304.23 ± 115.93 nM (n = 8 cases; mean ± standard deviation) in healthy control tissue and is similar to the target density in chronic traumatic encephalopathy tissues (365.80 ± 128.55 nM; n = 12 cases; mean ± standard deviation). A two-sample t-test determined no significant difference in the target density values of the 18 kDa translocator protein and monoamine oxidase-B between healthy controls and chronic traumatic encephalopathy (P > 0.05), albeit a trend towards increased expression of both targets was observed in chronic traumatic encephalopathy. To our knowledge, this work represents the first in vitro characterization of 18 kDa translocator protein and monoamine oxidase-B in chronic traumatic encephalopathy and reveals the variability in neuroinflammatory pathology following brain injuries. These preliminary findings will be considered when designing PET imaging studies after brain injury and for the ultimate goal of imaging chronic traumatic encephalopathy in vivo.

In Vitro and In Vivo Evaluation of GSK-3 Radioligands in Alzheimer's Disease: Preliminary Evidence of Sex Differences.

Glycogen synthase kinase-3 (GSK-3) is a positron emission tomography (PET) imaging target with implications in the pathogenesis of Alzheimer's disease (AD). This preliminary study evaluates human AD and transgenic P301L mouse brain tissues using the GSK-3-targeting radiotracers [3H]PF-367 and [3H]OCM-44 in radioligand binding assays. A saturation analysis showed decreased GSK-3 density in female human AD compared to a normal healthy brain. Equivalence in density (B max), affinity (K d), and apparent affinity (K i) of both radiotracers was demonstrated to enable their interchangeability for in vitro evaluations of GSK-3 expression. An evaluation of P301L mouse brain by [3H]/[11C]OCM-44 delineated differences in the B max of GSK-3 between the control and transgenic mice within male subjects. PET imaging showed similar trends to those observed in vitro. Sex differences are revealed as a potential parameter to consider in the development of GSK-3-targeted diagnostics and therapeutics and could guide recruitment for clinical studies.

In Vitro Evaluation of [3H]CPPC as a Tool Radioligand for CSF-1R.

Microglia play a role in several central nervous system (CNS) diseases and are a highly sought target for positron emission tomography (PET) imaging and therapeutic intervention. 5-Cyano-N-(4-(4-[11C]methylpiperazin-1-yl)-2-(piperidin-1-yl)phenyl)furan-2-carboxamide ([11C]CPPC) is a radiopharmaceutical designed to selectively target microglia via macrophage colony stimulating factor-1 receptor (CSF-1R) in the CNS. Herein, we report the first preclinical evaluation of [3H]CPPC using radioligand binding methods for the evaluation of putative CSF-1R inhibitors in rodent models of neuroinflammation. The distribution of [3H]CPPC by autoradiography did not align with 18 kDa translocator protein (TSPO) distribution using [3H]PBR28 and IBA-1 staining for microglia. In the CNS, [3H]CPPC had considerable nonspecific binding, as indicated by a low displacement of the tritiated ligand by unlabeled CPPC and the known CSF1R inhibitors BLZ-945 and PLX3397. Spleen was identified as a tissue that provided an adequate signal-to-noise ratio to enable screening with [3H]CPPC and a library of 20 novel PLX3397 derivatives. However, unlabeled CPPC lacked selectivity and showed off-target binding to a substantial number of kinase targets (204 out of 403 tested) at a concentration relevant to in vitro radioligand binding assays (10 µM). These findings suggest that, while [3H]CPPC may have utility as a radioligand tool for the evaluation of peripheral targets and screening of CSF-1R inhibitors, it may have limited utility as an in vivo CNS imaging probe on the basis of the current evaluation.

Radiosynthesis, In Vitro and In Vivo Evaluation of [18F]CBD-2115 as a First-in-Class Radiotracer for Imaging 4R-Tauopathies.

CBD-2115 was selected from a library of 148 compounds based on a pyridinyl-indole scaffold as a first-in-class 4R-tau radiotracer. In vitro binding assays showed [3H]CBD-2115 had a KD value of 6.9 nM and a nominal Bmax of 500 nM in 4R-tau expressing P301L transgenic mouse tissue. In binding assays with human brain tissue homogenates, [3H]CBD-2115 has a higher affinity (4.9 nM) for progressive supranuclear palsy specific 4R-tau deposits than [3H]flortaucipir (45 nM) or [3H]MK-6240 (>50 nM). [18F]CBD-2115 was reliably synthesized (3-11% radiochemical yield with molar activity of 27-111 GBq/µmol and >97% radiochemical purity). Dynamic PET imaging was conducted in mice, rats, and nonhuman primates, and all species showed initial brain uptake of 0.5-0.65 standardized uptake value with fast clearance from normal tissues. [3H]CBD-2115 could be a useful lead radioligand for further research in 4R-tauopathies, and PET radiotracer development will focus on improving brain uptake and binding affinity.

DDiT4L promotes autophagy and inhibits pathological cardiac hypertrophy in response to stress.

Physiological cardiac hypertrophy, in response to stimuli such as exercise, is considered adaptive and beneficial. In contrast, pathological cardiac hypertrophy that arises in response to pathological stimuli such as unrestrained high blood pressure and oxidative or metabolic stress is maladaptive and may precede heart failure. We found that the transcript encoding DNA damage-inducible transcript 4-like (DDiT4L) was expressed in murine models of pathological cardiac hypertrophy but not in those of physiological cardiac hypertrophy. In cardiomyocytes, DDiT4L localized to early endosomes and promoted stress-induced autophagy through a process involving mechanistic target of rapamycin complex 1 (mTORC1). Exposing cardiomyocytes to various types of pathological stress increased the abundance of DDiT4L, which inhibited mTORC1 but activated mTORC2 signaling. Mice with conditional cardiac-specific overexpression of DDiT4L had mild systolic dysfunction, increased baseline autophagy, reduced mTORC1 activity, and increased mTORC2 activity, all of which were reversed by suppression of transgene expression. Genetic suppression of autophagy also reversed cardiac dysfunction in these mice. Our data showed that DDiT4L may be an important transducer of pathological stress to autophagy through mTOR signaling in the heart and that DDiT4L could be therapeutically targeted in cardiovascular diseases in which autophagy and mTOR signaling play a major role.

A Novel Transgenic Mouse Model of Cardiac Hypertrophy and Atrial Fibrillation.

Cardiac hypertrophy is a major risk factor for the development of atrial fibrillation (AF). However, there are few animal models of AF associated with cardiac hypertrophy. In this study, we describe the in vivo electrophysiological characteristics and histopathology of a mouse model of cardiac hypertrophy that develops AF. Myostatin is a well-known negative regulator of skeletal muscle growth that was recently found to additionally regulate cardiac muscle growth. Using cardiac-specific expression of the inhibitory myostatin pro-peptide, we generated transgenic (TG) mice with dominant-negative regulation of MSTN (DN-MSTN). One line (DN-MSTN TG13) displayed ventricular hypertrophy, as well as spontaneous AF on the surface electrocardiogram (ECG), and was further evaluated. DN-MSTN TG13 had normal systolic function, but displayed atrial enlargement on cardiac MRI, as well as atrial fibrosis histologically. Baseline ECG revealed an increased P wave duration and QRS interval compared with wild-type littermate (WT) mice. Seven of 19 DN-MSTN TG13 mice had spontaneous or inducible AF, while none of the WT mice had atrial arrhythmias (p<0.05). Connexin40 (Cx40) was decreased in DN-MSTN TG13 mice, even in the absence of AF or significant atrial fibrosis, raising the possibility that MSTN signaling may play a role in Cx40 down-regulation and the development of AF in this mouse model. In conclusion, DN-MSTN TG13 mice represent a novel model of AF, in which molecular changes including an initial loss of Cx40 are noted prior to fibrosis and the development of atrial arrhythmias.