Cardiovascular Drug Development Research Articles

Natural AMPK Activators in Cardiovascular Disease Prevention.

Cardiovascular diseases (CVD), as a life-threatening global disease, is receiving worldwide attention. Seeking novel therapeutic strategies and agents is of utmost importance to curb CVD. AMP-activated protein kinase (AMPK) activators derived from natural products are promising agents for cardiovascular drug development owning to regulatory effects on physiological processes and diverse cardiometabolic disorders. In the past decade, different therapeutic agents from natural products and herbal medicines have been explored as good templates of AMPK activators. Hereby, we overviewed the role of AMPK signaling in the cardiovascular system, as well as evidence implicating AMPK activators as potential therapeutic tools. In the present review, efforts have been made to compile and update relevant information from both preclinical and clinical studies, which investigated the role of natural products as AMPK activators in cardiovascular therapeutics.

Can machine learning augment clinician adjudication of events in cardiovascular trials? A case study of major adverse cardiovascular events (MACE) across CVRM trials

Abstract Background and introduction Accurate identification of clinical outcome events is critical to obtaining reliable results in cardiovascular outcomes trials (CVOTs). Current processes for event adjudication are expensive and hampered by delays. As part of a larger project to more reliably identify outcomes, we evaluated the use of machine learning to automate event adjudication using data from the SOCRATES trial (NCT01994720), a large randomized trial comparing ticagrelor and aspirin in reducing risk of major cardiovascular events after acute ischemic stroke or transient ischemic attack (TIA). Purpose We studied whether machine learning algorithms could replicate the outcome of the expert adjudication process for clinical events of ischemic stroke and TIA. Could classification models be trained on historical CVOT data and demonstrate performance comparable to human adjudicators? Methods Using data from the SOCRATES trial, multiple machine learning algorithms were tested using grid search and cross validation. Models tested included Support Vector Machines, Random Forest and XGBoost. Performance was assessed on a validation subset of the adjudication data not used for training or testing in model development. Metrics used to evaluate model performance were Receiver Operating Characteristic (ROC), Matthews Correlation Coefficient, Precision and Recall. The contribution of features, attributes of data used by the algorithm as it is trained to classify an event, that contributed to a classification were examined using both Mutual Information and Recursive Feature Elimination. Results Classification models were trained on historical CVOT data using adjudicator consensus decision as the ground truth. Best performance was observed on models trained to classify ischemic stroke (ROC 0.95) and TIA (ROC 0.97). Top ranked features that contributed to classification of Ischemic Stroke or TIA corresponded to site investigator decision or variables used to define the event in the trial charter, such as duration of symptoms. Model performance was comparable across the different machine learning algorithms tested with XGBoost demonstrating the best ROC on the validation set for correctly classifying both stroke and TIA. Conclusions Our results indicate that machine learning may augment or even replace clinician adjudication in clinical trials, with potential to gain efficiencies, speed up clinical development, and retain reliability. Our current models demonstrate good performance at binary classification of ischemic stroke and TIA within a single CVOT with high consistency and accuracy between automated and clinician adjudication. Further work will focus on harmonizing features between multiple historical clinical trials and training models to classify several different endpoint events across trials. Our aim is to utilize these clinical trial datasets to optimize the delivery of CVOTs in further cardiovascular drug development. Funding Acknowledgement Type of funding sources: Private company. Main funding source(s): AstraZenca Plc

Current state and future of 3D bioprinted models for cardiovascular research and drug development.

In the last decade, 3D bioprinting technology has emerged as an innovative tissue engineering approach for regenerative medicine and drug development. This article aims at providing an overview about the most commonly used bioengineered tissues, focusing on 3D bioprinted cardiac cells and how they have been utilized for drug discovery and development. The review describes that, while this field is still developing, cardiovascular research may benefit from laboratory-engineered heart tissues built of specific cell types with precise 3D architecture mimicking the native cardiac microenvironment. It also describes the role played by regulatory agencies and potential commercialization pathways for direct translation from the bench to the bedside of studies using 3D bioprinted cardiac tissues.

Organs-on-chip models for cardiovascular drug development.

Organs-on-chip models for cardiovascular drug development.

Integrating genomics with biomarkers and therapeutic targets to invigorate cardiovascular drug development.

Drug development in cardiovascular disease is stagnating, with lack of efficacy and adverse effects being barriers to innovation. Human genetics can provide compelling evidence of causation through approaches such as Mendelian randomization, with genetic support for causation increasing the probability of a clinical trial succeeding. Mendelian randomization applied to quantitative traits can identify risk factors for disease that are both causal and amenable to therapeutic modification. However, important differences exist between genetic investigations of a biomarker (such as HDL cholesterol) and a drug target aimed at modifying the same biomarker of interest (such as cholesteryl ester transfer protein), with implications for the methodology, interpretation and application of Mendelian randomization to drug development. Differences include the comparative nature of the genetic architecture - that is, biomarkers are typically polygenic, whereas protein drug targets are influenced by either cis-acting or trans-acting genetic variants - and the potential for drug targets to show disease associations that might differ from those of the biomarker that they are intended to modify (target-mediated pleiotropy). In this Review, we compare and contrast the use of Mendelian randomization to evaluate potential drug targets versus quantitative traits. We explain how genetic epidemiological studies can be used to assess the aetiological roles of biomarkers in disease and to prioritize drug targets, including designing their evaluation in clinical trials.

Distinct circulating cytokine levels in patients with angiography-proven coronary artery disease compared to disease-free controls

Abstract Background Systemic inflammation has a critical role in the development of coronary artery disease (CAD). Identification of inflammatory pathways may provide a platform for novel therapeutic approaches. Purpose We sought to determine whether there are differences in circulating cytokine profiles between patients with CAD and disease-free controls. Methods Study population consisted of 458 patients who underwent diagnostic invasive coronary angiography for clinical indications. Altogether 312 patients had angiographically significant CAD whereas 146 had no angiographically-detected coronary atherosclerosis. We measured the serum concentrations of 48 circulating cytokines. Results Patients with CAD had increased levels of interleukin-1 receptor antagonist (IL-1-RA), interleukin-2 receptor alpha (IL-2Rα), IL-3, IL-4, IL-6, IL-9, IL-10, IL-16, IL-17, IL-18, granulocyte-macrophage colony-stimulating factor (GM-CSF), platelet-derived growth factor-BB (PDGF-BB), macrophage inflammatory protein-1-beta (MIP-1-β), tumor necrosis factor-alpha (TNF-α), TNF-β, cutaneous T-cell-attracting chemokine (CTACK), growth regulated oncogene alpha (GRO-α), hepatocyte growth factor (HGF), interferon alpha-2 (IFNα-2), leukemia inhibitory factor (LIF), macrophage colony-stimulating factor (M-CSF), macrophage migration inhibitory factor (MIF), monokine induced by gamma interferon (MIG), beta-nerve growth factor (β-NGF), stem cell factor (SCF) and stromal cell-derived factor 1 alfa (SDF1α). On a logistic multivariate regression model adjusted with age, sex, hypertension and treatment for diabetes, increased levels of IL-4 (p=0.027, OR 1.090), IL-9 (p=0.000, OR 1.013), IL-17 (p=0.011, OR 1.005), CTACK (p=0.008, OR 1.001), MIP-1-β (p=0.004, OR 1.006), GRO-α (p=0.008, OR 1.004), TNF-α (p=0.019, OR 1.011) were independently associated with atherosclerosis. Conclusions Patients with CAD have distinct circulating cytokine profiles compared to disease-free controls. Based on these findings, certain cytokines may have a pivotal role in the development of atherosclerotic cardiovascular disease and cytokine-mediated pathway appear as a promising target for cardiovascular drug development. Funding Acknowledgement Type of funding source: Foundation. Main funding source(s): The Finnish Medical Foundation, Helsinki, Finland; The Finnish Foundation for Cardiovascular Research, Helsinki, Finland

The Effect of Salvianolic Acid on Vascular Protection and Possible Mechanisms.

Salvia miltiorrhiza (Danshen), as an important traditional Chinese medicinal plant, has been used in China for the treatment of cardiovascular diseases for hundreds of years. Salvianolic acids (salvianolic acid A and salvianolic acid B) as the most abundant water-soluble component extracted from Salvia miltiorrhiza have attracted more and more attention from cardiovascular scientists due to its comprehensive cardiovascular actions. In vivo and in vitro studies have rendered salvianolic acid an excellent drug candidate for the treatment and prevention of cardiovascular diseases. In this review, we surveyed the protective effects of salvianolic acid A and salvianolic acid B against cardiovascular diseases and the pharmacological basis, providing a strong scientific rationale for elucidating the important role of Salvia miltiorrhiza in cardiovascular therapy. More importantly, we also hope to provide new inspiration and perspectives on the development and innovation of small-molecule cardiovascular drugs based on salvianolic acid.

Inflammageing in the cardiovascular system: mechanisms, emerging targets, and novel therapeutic strategies.

In the elderly population, pathological inflammation has been associated with ageing-associated diseases. The term 'inflammageing', which was used for the first time by Franceschi and co-workers in 2000, is associated with the chronic, low-grade, subclinical inflammatory processes coupled to biological ageing. The source of these inflammatory processes is debated. The senescence-associated secretory phenotype (SASP) has been proposed as the main origin of inflammageing. The SASP is characterised by the release of inflammatory cytokines, elevated activation of the NLRP3 inflammasome, altered regulation of acetylcholine (ACh) nicotinic receptors, and abnormal NAD+ metabolism. Therefore, SASP may be 'druggable' by small molecule therapeutics targeting those emerging molecular targets. It has been shown that inflammageing is a hallmark of various cardiovascular diseases, including atherosclerosis, hypertension, and adverse cardiac remodelling. Therefore, the pathomechanism involving SASP activation via the NLRP3 inflammasome; modulation of NLRP3 via α7 nicotinic ACh receptors; and modulation by senolytics targeting other proteins have gained a lot of interest within cardiovascular research and drug development communities. In this review, which offers a unique view from both clinical and preclinical target-based drug discovery perspectives, we have focused on cardiovascular inflammageing and its molecular mechanisms. We have outlined the mechanistic links between inflammageing, SASP, interleukin (IL)-1β, NLRP3 inflammasome, nicotinic ACh receptors, and molecular targets of senolytic drugs in the context of cardiovascular diseases. We have addressed the 'druggability' of NLRP3 and nicotinic α7 receptors by small molecules, as these proteins represent novel and exciting targets for therapeutic interventions targeting inflammageing in the cardiovascular system and beyond.

Improving Cardiovascular Drug and Device Development and Evidence Through Patient-Centered Research and Clinical Trials: A Call to Action From the Value in Healthcare Initiative's Partnering With Regulators Learning Collaborative.

The pipeline of new cardiovascular drugs is relatively limited compared with many other clinical areas. Challenges causing lagging drug innovation include the duration and expense of cardiovascular clinical trials needed for regulatory evaluation and approvals, which generally must demonstrate noninferiority to existing standards of care and measure longer-term outcomes. By comparison, there has been substantial progress in cardiovascular device innovation. There has also been progress in cardiovascular trial participation equity in recent years, especially among women, due in part to important efforts by Food and Drug Administration, National Institutes of Health, American Heart Association, and others. Yet women and especially racial and ethnic minority populations remain underrepresented in cardiovascular trials, indicating much work ahead to continue recent success. Given these challenges and opportunities, the multistakeholder Partnering with Regulators Learning Collaborative of the Value in Healthcare Initiative, a collaboration of the American Heart Association and the Robert J. Margolis, MD, Center for Health Policy at Duke University, identified how to improve the evidence generation process for cardiovascular drugs and devices. Drawing on a series of meetings, literature reviews, and analyses of regulatory options, the Collaborative makes recommendations across four identified areas for improvement. First, we offer strategies to enhance patient engagement in trial design, convenient participation, and meaningful end points and outcomes to improve patient recruitment and retention (major expenses in clinical trials). Second, new digital technologies expand the potential for real-world evidence to streamline data collection and reduce cost and time of trials. However, technical challenges must be overcome to routinely leverage real-world data, including standardizing data, managing data quality, understanding data comparability, and ensuring real-world evidence does not worsen inequities. Third, as trials are driven by evidence needs of regulators and payers, we recommend ways to improve their collaboration in trial design to streamline and standardize efficient and innovative trials, reducing costs and delays. Finally, we discuss creative ways to expand the minuscule proportion of sites involved in cardiovascular evidence generation and medical product development. These actions, paired with continued policy research into better ways to pay for and equitably develop therapies, will help reduce the cost and complexity of drug and device research, development, and trials.

Biased allosteric modulators as a novel approach to induce biased signalling in G‐protein coupled receptors: The case of beta 2 adrenergic receptor in cardiovascular diseases

G protein‐coupled receptors (GPCRs) are the therapeutic target of nearly half of the current drugs in the market. It has been now well established that GPCRs can signal through multiple transducers, including G proteins and beta‐arrestins. These signalling pathways can be activated or blocked by “balanced” agonists or antagonists, but also, they can also be selectively activated in a “biased” response. Until now, biased signalling has been induced by biased ligands and biased receptors, any of which can result in preferential signalling through G proteins or beta‐arrestins. However, the discovery and development of GPCR biased agonists has been quite challenging, since in most GPCRs there is no structure activity relationship studies available, moreover, different active conformations for each pathway are likely very similar. Traditionally, biased agonists have been developed to target only the orthosteric site which is the binding site of the endogenous ligands. Development of biased agonists targeting the orthosteric site might be extremely difficult, especially in the absence of structure‐activity relationship information. Biased agonism holds great promise as a mechanism to significantly reduce the side effects that current drugs in used in the clinic, as well as develop drugs for use on their own. Also, ligands that produce biased signalling will serve as valuable tools for elucidation of the molecular mechanisms underlying GPCR‐signalling. Allosteric modulators are ligands which bind to a receptor at a site distinct from that of the endogenous agonist and they do not activate the receptor rather than stabilize or induce intermediate active or inactive structural conformations modulating signalling of the orthosteric ligands. We tested the hypothesis that biased signalling of the receptor could be achieved by targeting allosteric domains which show high diversity in structure and amino acid sequence among the GPCRs. The rationale is that the development of a novel series of allosteric modulators will regulate GPCR signalling in such way that the endogenous ligand would produce biased signalling only when the target receptor is bound to an allosteric modulator. In this study, we targeted the intracellular domains of the beta 2 adrenergic receptor (b2‐AR) to produce beta‐arrestin biased signalling when activated by traditional orthosteric ligands. A highly selective b2‐AR biased allosteric modulator would be highly desirable to treat a number of cardiovascular diseases. Thus, we used a combination of state of art drug discovery platforms, in silico calculations and peptidomimetics to develop a beta‐arrestin biased allosteric modulator AR1981. This allosteric modulator increased the beta‐arrestin recruitment efficacy of isoproterenol by two‐fold. Also, AR1981 was able to increase the potency of isoproterenol in beta‐arrestin recruitment by more than ten‐fold. Thus, AR1981 is a promising tool to access a novel pharmacological profile stimulating cardioprotective signalling through the b2‐AR and can serve as a model for the next generation of cardiovascular drug development. The outcome of the proposed work will facilitate the development of a new generation of allosteric modulators for GPCRs.

Automated high-throughput heartbeat quantification in medaka and zebrafish embryos under physiological conditions

Accurate quantification of heartbeats in fish models is an important readout to study cardiovascular biology, disease states and pharmacology. However, dependence on anaesthesia, laborious sample orientation or requirement for fluorescent reporters have hampered the use of high-throughput heartbeat analysis. To overcome these limitations, we established an efficient screening assay employing automated label-free heart rate determination of randomly oriented, non-anesthetized medaka (Oryzias latipes) and zebrafish (Danio rerio) embryos in microtiter plates. Automatically acquired bright-field data feeds into an easy-to-use HeartBeat software with graphical user interface for automated quantification of heart rate and rhythm. Sensitivity of the assay was demonstrated by profiling heart rates during entire embryonic development. Our analysis revealed rapid adaption of heart rates to temperature changes, which has implications for standardization of experimental layout. The assay allows scoring of multiple embryos per well enabling a throughput of >500 embryos per 96-well plate. In a proof of principle screen for compound testing, we captured concentration-dependent effects of nifedipine and terfenadine over time. Our novel assay permits large-scale applications ranging from phenotypic screening, interrogation of gene functions to cardiovascular drug development.

Clinical Prescription-Protein-Small Molecule-Disease Strategy (CPSD), A New Strategy for Chinese Medicine Development: A Case Study in Cardiovascular Diseases.

Chinese medicine is a national treasure that has been passed down for thousands of years in China. According to the statistics of the World Health Organization, there are currently four billion people in the world who use Chinese medicine to treat diseases, accounting for 80% of the world’s total population. However, the obscurity of its theory, its unmanageable quality, its complex compositions, and the unknown effective substances and mechanisms are great obstacles to the internationalization of Chinese medicine. Here, we propose a new strategy for the development of Chinese medicine: the clinical prescription (C)-protein (P)-small-molecule (S)-disease (D) strategy, namely the CPSD strategy. The strategy uses clinical prescriptions as the source of medicine and uses computer simulation technology to find small-molecule drugs targeting therapeutic proteins for treating specific diseases so as to deepen awareness of the value of Chinese medicine. At the same time, this article takes cardiovascular drug development as an example to introduce the application of CPSD, which will be instrumental in the further development, modernization, and internationalization of Chinese medicine.

Sanofi exits diabetes, heart research

After just 100 days on the job, Sanofi CEO Paul Hudson has unveiled his plan for transforming the French pharmaceutical company over the next 3 years. Among Hudson’s goals for Sanofi are devoting resources to six specific drug candidates, creating a stand-alone consumer health unit, and saving roughly $2.2 billion. What the future does not include is diabetes and cardiovascular disease drug development. Sanofi markets the industry’s top-selling insulin treatment and for years relied on sales of Plavix, a now-generic blood thinner. But Hudson, who took over as CEO in September, told investors last week that while “our diabetes and cardiovascular portfolio is significant, we also have to recognize that it’s declining.” As part of that decision, Sanofi won’t launch efpeglenatide, a long-acting GLP-1 agonist for diabetes that is in late-stage studies. Hudson noted that many of the diabetes and cardiovascular compounds in development don’t offer enough benefits. “We don’t

Atherosclerosis imaging using PET: Insights and applications.

PET imaging is able to harness biological processes to characterise high-risk features of atherosclerotic plaque prone to rupture. Current radiotracers are able to track inflammation, microcalcification, hypoxia, and neoangiogenesis within vulnerable plaque. 18 F-fluorodeoxyglucose (18 F-FDG) is the most commonly used radiotracer in vascular studies and is employed as a surrogate marker of plaque inflammation. Increasingly, 18 F-FDG and other PET tracers are also being used to provide imaging endpoints in cardiovascular interventional trials. The evolution of novel PET radiotracers, imaging protocols, and hybrid scanners are likely to enable more efficient and accurate characterisation of high-risk plaque. This review explores the role of PET imaging in atherosclerosis with a focus on PET tracers utilised in clinical research and the applications of PET imaging to cardiovascular drug development.

Generation of two induced pluripotent stem cell lines (NHLBIi001-A and NHLBIi001-B) from a healthy Caucasian female volunteer with normal cardiac function.

Human-derived induced pluripotent stem cells (iPSCs) have proven to be indispensable in cardiovascular drug development, disease modeling, and developmental biology research. For this reason, it is particularly useful to develop wild-type iPSC lines to be used in experimental or control conditions. Here, we present two such cell lines generated from a sample of peripheral blood mononuclear cells (PBMCs) from a healthy patient with normal cardiac function.

Discontinued Drugs for the Treatment of Cardiovascular Disease from 2016 to 2018

Cardiovascular drug research and development (R&D) has been in active state and continuously attracts attention from the pharmaceutical industry. However, only one individual drug can eventually reach the market from about the 10,000 compounds tested. It would be useful to learn from these failures when developing better strategies for the future. Discontinued drugs were identified from a search performed by Thomson Reuters Integrity. Additional information was sought through PubMed, ClinicalTrials.gov, and pharmaceutical companies search. Twelve compounds discontinued for cardiovascular disease treatment after reaching Phase I–III clinical trials from 2016 to 2018 are detailed in this manuscript, and the reasons for these failures are reported. Of these, six candidates (MDCO-216, TRV027, ubenimex, sodium nitrite, losmapimod, and bococizumab) were dropped for lack of clinical efficacy, the other six for strategic or unspecified reasons. In total, three candidates were discontinued in Phase I trials, six in Phase II, and three in Phase III. It was reported that the success rate of drug R&D utilizing selection biomarkers is higher. Four candidate developments (OPC-108459, ONO-4232, GSK-2798745, and TAK-536TCH) were run without biomarkers, which could be used as surrogate endpoints in the 12 cardiovascular drugs discontinued from 2016 to 2018. This review will be useful for those involved in the field of drug discovery and development, and for those interested in the treatment of cardiovascular disease.

Impact of Drug Metabolism/Pharmacokinetics and their Relevance Upon Traditional Medicine-based Cardiovascular Drug Research.

The representative cardiovascular herbs, i.e. Panax, Ligusticum, Carthamus, and Pueraria plants, are traditionally and globally used in the prevention and treatment of various cardiovascular diseases. Modern phytochemical studies have found many medicinal compounds from these plants, and their unique pharmacological activities are being revealed. However, there are few reviews that systematically summarize the current trends of Drug Metabolism/Pharmacokinetic (DMPK) investigations of cardiovascular herbs. Here, the latest understanding, as well as the knowledge gaps of the DMPK issues in drug development and clinical usage of cardiovascular herbal compounds, was highlighted. The complicated herb-herb interactions of cardiovascular Traditional Chinese Medicine (TCM) herb pair/formula significantly impact the PK/pharmacodynamic performance of compounds thereof, which may inspire researchers to develop a novel herbal formula for the optimized outcome of different cardiovascular diseases. While the Absorption, Distribution, Metabolism, Excretion and Toxicity (ADME/T) of some compounds has been deciphered, DMPK studies should be extended to more cardiovascular compounds of different medicinal parts, species (including animals), and formulations, and could be streamlined by versatile omics platforms and computational analyses. In the context of systems pharmacology, the DMPK knowledge base is expected to translate bench findings to clinical applications, as well as foster cardiovascular drug discovery and development.

Antiarrhythmic effects of ginsenoside Rg2 on calcium chloride–induced arrhythmias without oral toxicity

BackgroundMalignant arrhythmias require drug therapy. However, most of the currently available antiarrhythmic drugs have significant side effects. Ginsenoside Rg2 exhibits excellent cardioprotective effects and appears to be a promising candidate for cardiovascular drug development. So far, the oral toxicity and antiarrhythmic effects of Rg2 have not been evaluated. MethodsAcute oral toxicity of Rg2 was assessed by the Limit Test method in mice. Subchronic oral toxicity was determined by repeated dose 28-day toxicity study in rats. Antiarrhythmic activities of Rg2 were evaluated in calcium chloride–induced arrhythmic rats. Antiarrhythmic mechanism of Rg2 was investigated in arrhythmic rats and H9c2 cardiomyocytes. ResultsThe results of toxicity studies indicated that Rg2 exhibited no single-dose (10 g/kg) acute oral toxicity. And 28-day repeated dose treatment with Rg2 (1.75, 3.5 and 5 g/kg/d) demonstrated minimal, if any, subchronic toxicity. Serum biochemical examination showed that total cholesterol in the high-dose cohort was dramatically decreased, whereas prothrombin time was increased at Day 28, suggesting that Rg2 might regulate lipid metabolism and have a potential anticoagulant effect. Moreover, pretreatment with Rg2 showed antiarrhythmic effects on the rat model of calcium chloride induced arrhythmia, in terms of the reduced duration time, mortality, and incidence of malignant arrhythmias. The antiarrhythmic mechanism of Rg2 might be the inhibition of calcium influx through L-type calcium channels by suppressing the phosphorylation of Ca2+/calmodulin-dependent protein kinase II. ConclusionOur findings support the development of Rg2 as a promising antiarrhythmic drug with fewer side effects for clinical use.

Quantitative Systems Pharmacology: An Exemplar Model-Building Workflow With Applications in Cardiovascular, Metabolic, and Oncology Drug Development.

Quantitative systems pharmacology (QSP), a mechanistically oriented form of drug and disease modeling, seeks to address a diverse set of problems in the discovery and development of therapies. These problems bring a considerable amount of variability and uncertainty inherent in the nonclinical and clinical data. Likewise, the available modeling techniques and related software tools are manifold. Appropriately, the development, qualification, application, and impact of QSP models have been similarly varied. In this review, we describe the progressive maturation of a QSP modeling workflow: a necessary step for the efficient, reproducible development and qualification of QSP models, which themselves are highly iterative and evolutive. Furthermore, we describe three applications of QSP to impact drug development; one supporting new indications for an approved antidiabetic clinical asset through mechanistic hypothesis generation, one highlighting efficacy and safety differentiation within the sodium‐glucose cotransporter‐2 inhibitor drug class, and one enabling rational selection of immuno‐oncology drug combinations.

Protective effects of five compounds from Livistona chinensis R. Brown leaves against hypoxia/reoxygenation, H2O2, or adriamycin-induced injury in H9c2 cells.

Purpose: Discovering new antimyocardial ischemia drug candidates that are highly efficient, have low toxicity, and originate from natural products is a popular trend for new cardiovascular drug development at present. The ethanol extract of Livistona chinensis leaves showed a favorable antioxidant activity in our preliminary screening test. This study aims to screen out antioxidants from the herb leaves further and evaluate their efficacy in acute myocardial ischemia treatment at the cellular level.Materials and methods: Guided with online 1, 1-diphenyl-2-picrylhydrazyl (DPPH)–high-performance liquid chromatography (HPLC) screening, antioxidants were first separated and isolated from the ethanol extract of L. chinensis leaves by preparative-HPLC. Subsequently, offline DPPH approach was used to validate the free radical scavenging activity of the components. Ultimately, the resulting antioxidants were evaluated against the hypoxia/reoxygenation (H/R)-, H2O2-, or adriamycin (ADM)-induced injury in H9c2 cells to verify their cardioprotective effects in vitro.Results: Five antioxidant ingredients, namely, orientin, isoorientin, vitexin, isovitexin, and tricin, were quickly distinguished and isolated from L. chinensis leaves. The IC50 values of these ingredients were further examined by offline DPPH assay, as follows: 15.51±0.22, 6.64±0.38, 11.86±0.24, 8.89±0.66, and 31.86±0.24 μg/mL, respectively. Out of these ingredients, isoorientin showed the strongest antioxidation, which was equivalent to that of the positive control drug (vitamin C, IC50: 6.99±0.62 μg/mL). Using H/R-, H2O2-, and ADM-induced H9c2 cell injury models, the five ingredients had different extents of cardioprotective effects in vitro. In particular, isoorientin showed the strongest protection. All the five ingredients also showed insignificant cytotoxic effect to normal H9c2 cells.Conclusion: The ethanol extract of L. chinensis leaves contained five antioxidants with low cardiac cytotoxicity. Isoorientin possessed the strongest antioxidation, which can predominantly account for the myocardial protection effects within the extract.