Late Na(+) current (INaL) is enhanced in myocytes of animals with chronic heart failure and patients with hypertrophic cardiomyopathy. To define the role of INaL in diastolic heart failure, the effects of GS-458967 (GS-967), a potent INaL inhibitor on mechanical and electrical abnormalities, were determined in an animal model of diastolic dysfunction. Dahl salt-sensitive (DSS) rats fed a high-salt (HS) diet for 8 wk, compared with a normal salt (NS) diet, had increased left ventricular (LV) mass (1,257 ± 96 vs. 891 ± 34 mg) and diastolic dysfunction [isovolumic relaxation time (IVRT): 26.8 ± 0.5 vs. 18.9 ± 0.2 ms; early transmitral flow velocity/early mitral annulus velocity (E/E') ratio: 25.5 ± 1.9 vs. 14.9 ± 0.9]. INaL in LV myocytes from HS rats was significantly increased to 0.41 ± 0.02 from 0.14 ± 0.02 pA/pF in NS rats. The action potential duration (APD) was prolonged to 136 ± 12 from 68 ± 9 ms in NS rats. QTc intervals were longer in HS vs. NS rats (267 ± 8 vs. 212 ± 2 ms). Acute and chronic treatment with GS-967 decreased the enhanced INaL to 0.24 ± 0.01 and 0.17 ± 0.02 pA/pF, respectively, vs. 0.41 ± 0.02 pA/pF in the HS group. Chronic treatment with GS-967 dose-dependently reduced LV mass, the increases in E/E' ratio, and the prolongation of IVRT by 27, 27, and 20%, respectively, at the 1.0 mg·kg(-1)·day(-1) dose without affecting blood pressure or LV systolic function. The prolonged APDs in myocytes and QTc of HS rats were significantly reduced with GS-967 treatment. These results indicate that INaL is a significant contributor to the LV diastolic dysfunction, hypertrophy, and repolarization abnormalities and thus, inhibition of this current is a promising therapeutic target for diastolic heart failure.

Background: Heart failure with preserved ejection fraction (HFpEF) is associated with high morbidity and mortality, yet there are few effective therapies for HFpEF. Previously, we have demonstrated that TN-301, a highly selective small molecule inhibitor of histone deacetylase 6 (HDAC6), works directly on the heart and on systemic metabolism and inflammation in mouse models of HFpEF. Recently, empagliflozin, a selective inhibitor of sodium-glucose transport protein (SGLT2) inhibitor, was approved in the U.S. for patients with HFpEF. Purpose: We aimed to compare the efficacy of HDAC6i and SGLT2i in a mouse model of established diastolic dysfunction with preserved ejection fraction. Methods and Results: To recapitulate systemic and cardiovascular features of HFpEF in humans, we induced diastolic dysfunction with a combination of high fat diet and inhibition of constitutive nitric oxide synthase using N ω -nitrol-arginine methyl ester (L-NAME). After the HFpEF phenotypes were established, mice received TYA-11018 (TN-301-like HDAC6i) or empagliflozin, or vehicle orally once daily for 9 weeks. We found that TYA-11018 reduced left ventricular hypertrophy and restored diastolic function with comparable efficacy to empagliflozin. In addition, TYA-11018 reduced fasting glucose and improved glucose tolerance in HFpEF mice, to levels similar to empagliflozin. Preliminary data suggested TYA-11018 corrects hyperglycemia independently of glucose urine excretion, which is distinct from SGLT2i. Using whole transriptome RNA-seq, TYA-11018 showed greater effects on correcting expression of gene sets associated with inflammation, fibrosis and mitochondrial energy production compared to empagliflozin in HFpEF hearts. Conclusion: Our results show that with its distinct HDAC6 inhibition mechanism, TYA-11018 reverses preexisting hypertrophy, diastolic dysfunction and glucose tolerance, demonstrated similar efficacy to that with a SGLT2 inhibitor in HFpEF model. The comparable efficacy seen in this HFpEF model with TYA-11018 and empagliflozin provides early but encouraging evidence of the potential translatability of these findings to clinical development. We are developing TN-301 for treating HFpEF patients.

Heart failure affects an estimated 38 million people worldwide and is typically caused by cardiomyocyte (CM) loss or dysfunction. Although CMs have limited ability to regenerate, a large pool of non-myocytes, including cardiac fibroblasts (CFs), exist in the postnatal heart. In vivo reprogramming of non-myocytes into functional CMs is emerging as a potential new approach to treat heart failure and substantial proof-of-concept has been achieved in this new field. However, challenges remain in terms of clinical application. First, reported human reprogramming cocktails often consist of five to seven factors that require multiple AAV vectors for delivery. Thus, a less complex cocktail that is able to fit into one AAV vector is needed for this technology to impact human health. Second, the lack of specificity in AAV tropism further complicates the safety and regulatory landscape. A means to limit the expression of reprogramming factors to target cells is critical for maximizing long-term safety. Lastly, although promising studies in small animals have already been reported, safety and efficacy results in large animal MI models are critical to justify cardiac reprogramming in human clinical trials. We have developed a novel human cardiac reprogramming cocktail that consists of only two transcription factors and one miRNA. This new cocktail has been engineered into a single AAV cassette to efficiently reprogram human CFs into cardiomyocytes. We also substantially improved transduction of hCFs through AAV capsid engineering and eliminated CMs expression through a microRNA de-targeting method. Moreover, our novel cardiac reprogramming gene therapy improved cardiac function in both rat and swine MI models upon delivery at various time-points after MI without inducing arrhythmias. Given these promising safety and efficacy results in larger animals, we endeavor to translate direct cardiac reprogramming for clinical application.

Introduction: It has been shown that activated CaMKII can phosphorylate cardiac Na+ channels that leads to an enhanced late Na current (INaL). We hypothesized that enhanced INaL plays an important role in arrhythmogenesis linked to increased CaMKII activity. To test this hypothesis we determined the effect of a selective INaL inhibitor GS-967 on spontaneously occurring ventricular arrhythmias in mice overexpressing CaMKIIδc (TG). Methods: TG (n=6) and wild type (WT, n=3) mice at age 8 week (wk) were instrumented with telemetry transmitters to record ECG. ECG were recorded continuously starting 1 wk post- surgery (age 9 wk) until age 19 wk. Incidence and burden of arrhythmias were evaluated at ages 9, 11, 13, 15, 17 and 19 wks using DSI ECG Pro software. At age 17 wk, when arrhythmia burden was relatively stable, mice were treated with a single dose of GS-967 (1 mg/kg, i.p.) or vehicle. After 4 days of washout, the treatment was repeated in a cross-over manner. Arrhythmia burden was quantified over a duration of 15-hours post-treatment. Using patch clamp technique, INaL was measured in ventricular myocytes isolated from both TG and WT mice at 17 wk age. Results: Incidence and burden of spontaneous ventricular arrhythmias increased progressively with age in TG mice. Between age 9 and 11 wks, 50% of TG mice had an arrhythmia burden ≥ 5 minutes/24 hours. The incidence increased to 83% at wk 13, 15, 17 and to 100% at 19 wks and the burden increased to 71 ± 35 min/24 hr at 17 wk. A single dose of GS-967 significantly decreased arrhythmia burden from 42.5 ± 9.6 min (treated with vehicle) to 9.4 ± 4.3 min (p<0.05) during a 15-hr period. No arrhythmias were observed in WT mice during the course of the study. INaL was enhanced by more than 2-fold in myocytes isolated from TG mice compared to WT (0.2±0.03 vs. 0.08±0.02 pA/pF, n=16 each, p<0.05). GS-967 caused a concentration-dependent reduction of INaL (50.5±3.4% at 0.3 μM, n=7; and 91±5.2% at 1 μM, n=4). Conclusions: This is the first report to show a reduction of spontaneously occurring ventricular arrhythmias by inhibition of INaL indicating a key role for INaL in CaMKII associated arrhythmogenesis.