Background— Current therapies are ineffective in preventing the development of cardiac phenotype in young carriers of mutations associated with hypertrophic cardiomyopathy (HCM). Ranolazine, a late Na+ current blocker, reduced the electromechanical dysfunction of human HCM myocardium in vitro. Methods and Results— To test whether long-term treatment prevents cardiomyopathy in vivo, transgenic mice harboring the R92Q troponin-T mutation and wild-type littermates received an oral lifelong treatment with ranolazine and were compared with age-matched vehicle-treated animals. In 12-months-old male R92Q mice, ranolazine at therapeutic plasma concentrations prevented the development of HCM-related cardiac phenotype, including thickening of the interventricular septum, left ventricular volume reduction, left ventricular hypercontractility, diastolic dysfunction, left-atrial enlargement and left ventricular fibrosis, as evaluated in vivo using echocardiography and magnetic resonance. Left ventricular cardiomyocytes from vehicle-treated R92Q mice showed marked excitation–contraction coupling abnormalities, including increased diastolic [Ca2+] and Ca2+ waves, whereas cells from treated mutants were undistinguishable from those from wild-type mice. Intact trabeculae from vehicle-treated mutants displayed inotropic insufficiency, increased diastolic tension, and premature contractions; ranolazine treatment counteracted the development of myocardial mechanical abnormalities. In mutant myocytes, ranolazine inhibited the enhanced late Na+ current and reduced intracellular [Na+] and diastolic [Ca2+], ultimately preventing the pathological increase of calmodulin kinase activity in treated mice. Conclusions— Owing to the sustained reduction of intracellular Ca2+ and calmodulin kinase activity, ranolazine prevented the development of morphological and functional cardiac phenotype in mice carrying a clinically relevant HCM-related mutation. Pharmacological inhibitors of late Na+ current are promising candidates for an early preventive therapy in young phenotype-negative subjects carrying high-risk HCM-related mutations.

BackgroundIn cardiomyocytes from patients with hypertrophic cardiomyopathy, mechanical dysfunction and arrhythmogenicity are caused by mutation‐driven changes in myofilament function combined with excitation‐contraction (E‐C) coupling abnormalities related to adverse remodeling. Whether myofilament or E‐C coupling alterations are more relevant in disease development is unknown. Here, we aim to investigate whether the relative roles of myofilament dysfunction and E‐C coupling remodeling in determining the hypertrophic cardiomyopathy phenotype are mutation specific.Methods and ResultsTwo hypertrophic cardiomyopathy mouse models carrying the R92Q and the E163R TNNT2 mutations were investigated. Echocardiography showed left ventricular hypertrophy, enhanced contractility, and diastolic dysfunction in both models; however, these phenotypes were more pronounced in the R92Q mice. Both E163R and R92Q trabeculae showed prolonged twitch relaxation and increased occurrence of premature beats. In E163R ventricular myofibrils or skinned trabeculae, relaxation following Ca2+ removal was prolonged; resting tension and resting ATPase were higher; and isometric ATPase at maximal Ca2+ activation, the energy cost of tension generation, and myofilament Ca2+ sensitivity were increased compared with that in wild‐type mice. No sarcomeric changes were observed in R92Q versus wild‐type mice, except for a large increase in myofilament Ca2+ sensitivity. In R92Q myocardium, we found a blunted response to inotropic interventions, slower decay of Ca2+ transients, reduced SERCA function, and increased Ca2+/calmodulin kinase II activity. Contrarily, secondary alterations of E‐C coupling and signaling were minimal in E163R myocardium.ConclusionsIn E163R models, mutation‐driven myofilament abnormalities directly cause myocardial dysfunction. In R92Q, diastolic dysfunction and arrhythmogenicity are mediated by profound cardiomyocyte signaling and E‐C coupling changes. Similar hypertrophic cardiomyopathy phenotypes can be generated through different pathways, implying different strategies for a precision medicine approach to treatment.

Trace amine associated receptor 1 (TAAR1) is a G protein coupled receptor (GPCR) expressed in brain and periphery activated by a wide spectrum of agonists that include, but are not limited to, trace amines (TAs), amphetamine-like psychostimulants, and endogenous thyronamines such as thyronamine (T0AM) and 3-iodothyronamine (T1AM). Such polypharmacology has made it challenging to understand the role and the biology of TAAR1. In an effort to understand the molecular basis of TAAR1 activation, we rationally designed and synthesized a small family of thyronamine derivatives. Among them, compounds 2 and 3 appeared to be a good mimic of the parent endogenous thyronamine, T0AM and T1AM, respectively, both in vitro and in vivo. Thus, these compounds offer suitable tools for studying the physiological roles of mouse TAAR1 and could represent the starting point for the development of more potent and selective TAAR1 ligands.

a b s t r a c tChemotherapy-Induced Peripheral Neuropathy (CIPN) is the most frequent adverse effect of pharmacological cancer treatments. The occurrence of neuropathy prevents the administration of fully-effective drug regimen, affects negatively the quality of life of patients, and may lead to therapy discontinuation. CIPN is currently treated with anticonvulsants, antidepressants, opioids and non-opioid analgesics, all of which are flawed by insufficient anti-hyperalgesic efficacy or addictive potential. Understandably, developing new drugs targeting CIPN-specific pathogenic mechanisms would dramatically improve efficacy and tolerability of anti-neuropathic therapies. Neuropathies are associated to aberrant excitability of DRG neurons due to the alteration in the expression or function of a variety of ion channels. In this regard, Hyperpolarization-activated Cyclic Nucleotide-gated (HCN) channels are overexpressed in inflammatory and neuropathic pain states, and HCN blockers have been shown to reduce neuronal excitability and to ameliorate painful states in animal models. However, HCN channels are critical in cardiac action potential, and HCN blockers used so far in pre-clinical models do not discriminate between cardiac and non-cardiac HCN isoforms. In this work, we show an HCN current gain of function in DRG neurons from oxaliplatin-treated rats. Biochemically, we observed a downregulation of HCN2 expression and an upregulation of the HCN regulatory beta-subunit MirP1. Finally, we report the efficacy of the selective HCN1 inhibitor MEL57A in reducing hyperalgesia and allodynia in oxaliplatin-treated rats without cardiac effects. In conclusion, this study strengthens the evidence for a disease-specific role of HCN1 in CIPN, and proposes HCN1-selective inhibitors as new-generation pain medications with the desired efficacy and safety profile.

3-Iodothyronamine (T1AM) is an endogenous high-affinity ligand of the trace amine-associated receptor 1 (TAAR1), detected in mammals in many organs, including the brain. Recent evidence indicates that pharmacological TAAR1 activation may offer a novel therapeutic option for the treatment of a wide range of neuropsychiatric and metabolic disorders. To assess potential neuroprotection by TAAR1 agonists, in the present work, we initially investigated whether T1AM and its corresponding 3-methylbiaryl-methane analog SG-2 can improve learning and memory when systemically administered to mice at submicromolar doses, and whether these effects are modified under conditions of MAO inhibition by clorgyline. Our results revealed that when i.p. injected to mice, both T1AM and SG-2 produced memory-enhancing and hyperalgesic effects, while increasing ERK1/2 phosphorylation and expression of transcription factor c-fos. Notably, both compounds appeared to rely on the action of ubiquitous enzymes MAO to produce the corresponding oxidative metabolites that were then able to activate the histaminergic system. Since autophagy is key for neuronal plasticity, in a second line of experiments we explored whether T1AM and synthetic TAAR1 agonists SG1 and SG2 were able to induce autophagy in human glioblastoma cell lines (U-87MG). After treatment of U-87MG cells with 1 μM T1AM, SG-1, SG-2 transmission electron microscopy (TEM) and immunofluorescence (IF) showed a significant time-dependent increase of autophagy vacuoles and microtubule-associated protein 1 light chain 3 (LC3). Consistently, Western blot analysis revealed a significant increase of the LC3II/LC3I ratio, with T1AM and SG-1 being the most effective agents. A decreased level of the p62 protein was also observed after treatment with T1AM and SG-1, which confirms the efficacy of these compounds as autophagy inducers in U-87MG cells. In the process to dissect which pathway induces ATG, the effects of these compounds were evaluated on the PI3K-AKT-mTOR pathway. We found that 1 μM T1AM, SG-1 and SG-2 decreased pAKT/AKT ratio at 0.5 and 4 h after treatment, suggesting that autophagy is induced by inhibiting mTOR phosphorylation by PI3K-AKT-mTOR pathway. In conclusion, our study shows that T1AM and thyronamine-like derivatives SG-1 and SG-2 might represent valuable tools to therapeutically intervene with neurological disorders.