RESUMO
We recently reported that SF2312 ((1,5-dihydroxy-2-oxopyrrolidin-3-yl)phosphonic acid), a phosphonate antibiotic with a previously unknown mode of action, is a potent inhibitor of the glycolytic enzyme, Enolase. SF2312 can only be synthesized as a racemic-diastereomeric mixture. However, co-crystal structures with Enolase 2 (ENO2) have consistently shown that only the (3S,5S)-enantiomer binds to the active site. The acidity of the alpha proton at C-3, which deprotonates under mildly alkaline conditions, results in racemization; thus while the separation of four enantiomeric intermediates was achieved via chiral High Performance Liquid Chromatography (HPLC) of the fully protected intermediate, deprotection inevitably nullified enantiopurity. To prevent epimerization of the C-3, we designed and synthesized MethylSF2312, ((1,5-dihydroxy-3-methyl-2-oxopyrrolidin-3-yl)phosphonic acid), which contains a fully-substituted C-3 alpha carbon. As a racemic-diastereomeric mixture, MethylSF2312 is equipotent to SF2312 in enzymatic and cellular systems against Enolase. Chiral HPLC separation of a protected MethylSF2312 precursor resulted in the efficient separation of the four enantiomers. After deprotection and inevitable re-equilibration of the anomeric C-5, (3S)-MethylSF2312 was up to 2000-fold more potent than (3R)-MethylSF2312 in an isolated enzymatic assay. This observation strongly correlates with biological activity in both human cancer cells and bacteria for the 3S enantiomer of SF2312. Novel X-ray structures of human ENO2 with chiral and racemic MethylSF2312 show that only (3S,5S)-enantiomer occupies the active site. Enolase inhibition is thus a direct result of binding by the (3S,5S)-enantiomer of MethylSF2312. Concurrent with these results for MethylSF2312, we contend that the (3S,5S)-SF2312 is the single active enantiomer of inhibitor SF2312.
Assuntos
Inibidores Enzimáticos/farmacologia , Organofosfonatos/farmacologia , Fosfopiruvato Hidratase/antagonistas & inibidores , Fosfopiruvato Hidratase/química , Pirrolidinonas/farmacologia , Sítios de Ligação , Ativação Enzimática/efeitos dos fármacos , Inibidores Enzimáticos/química , Modelos Moleculares , Conformação Molecular , Estrutura Molecular , Organofosfonatos/química , Ligação Proteica , Pirrolidinonas/química , Análise Espectral , Estereoisomerismo , Relação Estrutura-AtividadeRESUMO
Despite being crucial for energy generation in most forms of life, few if any microbial antibiotics specifically inhibit glycolysis. To develop a specific inhibitor of the glycolytic enzyme enolase 2 (ENO2) for the treatment of cancers with deletion of ENO1 (encoding enolase 1), we modeled the synthetic tool compound inhibitor phosphonoacetohydroxamate (PhAH) into the active site of human ENO2. A ring-stabilized analog of PhAH, in which the hydroxamic nitrogen is linked to Cα by an ethylene bridge, was predicted to increase binding affinity by stabilizing the inhibitor in a bound conformation. Unexpectedly, a structure-based search revealed that our hypothesized backbone-stabilized PhAH bears strong similarity to SF2312, a phosphonate antibiotic of unknown mode of action produced by the actinomycete Micromonospora, which is active under anaerobic conditions. Here, we present multiple lines of evidence, including a novel X-ray structure, that SF2312 is a highly potent, low-nanomolar inhibitor of enolase.
Assuntos
Inibidores Enzimáticos/farmacologia , Organofosfonatos/farmacologia , Fosfopiruvato Hidratase/antagonistas & inibidores , Pirrolidinonas/farmacologia , Relação Dose-Resposta a Droga , Inibidores Enzimáticos/química , Humanos , Modelos Moleculares , Estrutura Molecular , Organofosfonatos/química , Fosfopiruvato Hidratase/metabolismo , Pirrolidinonas/química , Relação Estrutura-AtividadeRESUMO
This study aimed to describe and evaluate the type, frequency and patterns of congenital heart diseases (CHDs) in patients with Down Syndrome (DS) in Sétif, Algeria. Down Syndrome, or trisomy 21, is the most common genetic disorder in the world. Data were collected and followed from January 2009 to December 2013. Parental consanguinity documenting pedigree analyzing, chromosome analysis and clinical examination were carried out for all cases. Results have shown that 22 (15.4%; ± 0.06) of the total 143 known cases of DS from DS centres have CHDs and 88 (10.6%; ± 2.2) of the total 770 patients with CHDs collected from public departments at the child and maternity teaching hospital, Sétif, have DS. Among the 110 cases, 75 (68%) have single cardiac abnormalities and 35 (32%) have multiple cardiac abnormalities. The most frequent CHDs were Atrioventricular Septal Defect (AVSD). In conclusion, our study will be helpful to demonstrate the current status of DS and to identify the distribution of CHD in patients with DS in Sétif, Algeria, for further study.
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Síndrome de Down/epidemiologia , Cardiopatias Congênitas/epidemiologia , Adolescente , Argélia/epidemiologia , Criança , Pré-Escolar , Consanguinidade , Feminino , Humanos , Lactente , Masculino , Fatores de Risco , Adulto JovemRESUMO
3ß,16ß,17α-Trihydroxycholest-5-en-22-one 16-O-(2-O-4-methoxybenzoyl-ß-D-xylopyranosyl)-(1â3)-2-O-acetyl-α-L-arabinopyranoside (OSW-1) is a natural product with potent antitumor activity against various types of cancer cells, but the exact mechanisms of action remain to be defined. In this study, we showed that OSW-1 effectively killed leukemia cells at subnanomolar concentrations through a unique mechanism by causing a time-dependent elevation of cytosolic Ca(2+) prior to induction of apoptosis. A mechanistic study revealed that this compound inhibited the sodium-calcium exchanger 1 on the plasma membrane, leading to an increase in cytosolic Ca(2+) and a decrease in cytosolic Na(+). The elevated cytosolic Ca(2+) caused mitochondrial calcium overload and resulted in a loss of mitochondrial membrane potential, release of cytochrome c, and activation of caspase-3. Furthermore, OSW-1 also caused a Ca(2+)-dependent cleavage of the survival factor GRP78. Inhibition of Ca(2+) entry into the mitochondria by the uniporter inhibitor RU360 or by cyclosporin A significantly prevented the OSW-1-induced cell death, indicating the important role of mitochondria in mediating the cytotoxic activity. The extremely potent activity of OSW-1 against leukemia cells and its unique mechanism of action suggest that this compound may be potentially useful in the treatment of leukemia.
Assuntos
Produtos Biológicos/farmacologia , Cálcio/metabolismo , Colestenonas/farmacologia , Homeostase/efeitos dos fármacos , Leucemia/metabolismo , Leucemia/patologia , Saponinas/farmacologia , Canais de Cálcio/metabolismo , Calpaína/metabolismo , Caspase 3/metabolismo , Morte Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Ciclosporina/farmacologia , Citocromos c/metabolismo , Citosol/efeitos dos fármacos , Citosol/metabolismo , Ensaios de Seleção de Medicamentos Antitumorais , Retículo Endoplasmático/efeitos dos fármacos , Retículo Endoplasmático/metabolismo , Chaperona BiP do Retículo Endoplasmático , Ativação Enzimática/efeitos dos fármacos , Espaço Extracelular/efeitos dos fármacos , Espaço Extracelular/metabolismo , Proteínas de Choque Térmico/metabolismo , Humanos , Leucemia/enzimologia , Linfoma/enzimologia , Linfoma/patologia , Potencial da Membrana Mitocondrial/efeitos dos fármacos , Mitocôndrias/efeitos dos fármacos , Mitocôndrias/metabolismo , Trocador de Sódio e Cálcio/antagonistas & inibidores , Trocador de Sódio e Cálcio/metabolismo , Tapsigargina/farmacologia , Fatores de TempoRESUMO
INTRODUCTION: Triple-negative breast cancer (TNBC) is a subtype of highly malignant breast cancer with poor prognosis. TNBC is not amenable to endocrine therapy and often exhibit resistance to current chemotherapeutic agents, therefore, further understanding of the biological properties of these cancer cells and development of effective therapeutic approaches are urgently needed. METHODS: We first investigated the metabolic alterations in TNBC cells in comparison with other subtypes of breast cancer cells using molecular and metabolic analyses. We further demonstrated that targeting these alterations using specific inhibitors and siRNA approach could render TNBC cells more sensitive to cell death compared to other breast cancer subtypes. RESULTS: We found that TNBC cells compared to estrogen receptor (ER) positive cells possess special metabolic characteristics manifested by high glucose uptake, increased lactate production, and low mitochondrial respiration which is correlated with attenuation of mTOR pathway and decreased expression of p70S6K. Re-expression of p70S6K in TNBC cells reverses their glycolytic phenotype to an active oxidative phosphorylation (OXPHOS) state, while knockdown of p70S6K in ER positive cells leads to suppression of mitochondrial OXPHOS. Furthermore, lower OXPHOS activity in TNBC cells renders them highly dependent on glycolysis and the inhibition of glycolysis is highly effective in targeting TNBC cells despite their resistance to other anticancer agents. CONCLUSIONS: Our study shows that TNBC cells have profound metabolic alterations characterized by decreased mitochondrial respiration and increased glycolysis. Due to their impaired mitochondrial function, TNBC cells are highly sensitive to glycolytic inhibition, suggesting that such metabolic intervention may be an effective therapeutic strategy for this subtype of breast cancer cells.
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Mitocôndrias/metabolismo , Serina-Treonina Quinases TOR/metabolismo , Neoplasias de Mama Triplo Negativas/metabolismo , Trifosfato de Adenosina/metabolismo , Linhagem Celular Tumoral , Complexo de Proteínas da Cadeia de Transporte de Elétrons/metabolismo , Metabolismo Energético/efeitos dos fármacos , Feminino , Glucose/metabolismo , Glutationa/metabolismo , Humanos , Hidrocarbonetos Bromados/farmacologia , Ácido Láctico/metabolismo , NADP/metabolismo , Oxirredução , Consumo de Oxigênio , Propionatos/farmacologia , Espécies Reativas de Oxigênio/metabolismo , Receptor ErbB-2/metabolismo , Receptores de Estrogênio/metabolismo , Receptores de Progesterona/metabolismo , Transdução de Sinais , Neoplasias de Mama Triplo Negativas/tratamento farmacológicoRESUMO
Metabolically labile prodrugs can experience stark differences in catabolism incurred by the chosen route of administration. This is especially true for phosph(on)ate prodrugs, in which successive promoiety removal transforms a lipophilic molecule into increasingly polar compounds. We previously described a phosphonate inhibitor of enolase (HEX) and its bis-pivaloyloxymethyl ester prodrug (POMHEX) capable of eliciting strong tumor regression in a murine model of enolase 1 (ENO1)-deleted glioblastoma following parenteral administration. Here, we characterize the pharmacokinetics and pharmacodynamics of these enolase inhibitors in vitro and in vivo after oral and parenteral administration. In support of the historical function of lipophilic prodrugs, the bis-POM prodrug significantly improves cell permeability of and rapid hydrolysis to the parent phosphonate, resulting in rapid intracellular loading of peripheral blood mononuclear cells in vitro and in vivo. We observe the influence of intracellular trapping in vivo on divergent pharmacokinetic profiles of POMHEX and its metabolites after oral and parenteral administration. This is a clear demonstration of the tissue reservoir effect hypothesized to explain phosph(on)ate prodrug pharmacokinetics but has heretofore not been explicitly demonstrated.
RESUMO
Cancers harboring homozygous deletion of the glycolytic enzyme enolase 1 (ENO1) are selectively vulnerable to inhibition of the paralogous isoform, enolase 2 (ENO2). A previous work described the sustained tumor regression activities of a substrate-competitive phosphonate inhibitor of ENO2, 1-hydroxy-2-oxopiperidin-3-yl phosphonate (HEX) (5), and its bis-pivaloyoxymethyl prodrug, POMHEX (6), in an ENO1-deleted intracranial orthotopic xenograft model of glioblastoma [Nature Metabolism 2020, 2, 1423-1426]. Due to poor pharmacokinetics of bis-ester prodrugs, this study was undertaken to identify potential non-esterase prodrugs for further development. Whereas phosphonoamidate esters were efficiently bioactivated in ENO1-deleted glioma cells, McGuigan prodrugs were not. Other strategies, including cycloSal and lipid prodrugs of 5, exhibited low micromolar IC50 values in ENO1-deleted glioma cells and improved stability in human serum over 6. The activity of select prodrugs was also probed using the NCI-60 cell line screen, supporting its use to examine the relationship between prodrugs and cell line-dependent bioactivation.
Assuntos
Glioblastoma , Glioma , Organofosfonatos , Pró-Fármacos , Humanos , Pró-Fármacos/uso terapêutico , Pró-Fármacos/farmacocinética , Organofosfonatos/farmacologia , Homozigoto , Deleção de Sequência , Fosfopiruvato Hidratase/genética , Fosfopiruvato Hidratase/metabolismo , Glioblastoma/tratamento farmacológico , Ésteres , Lipídeos , Proteínas de Ligação a DNA , Biomarcadores Tumorais , Proteínas Supressoras de Tumor/genéticaRESUMO
Cancer metabolism has emerged as an important area of research in recent years. Elucidation of the metabolic differences between cancer and normal cells and the underlying mechanisms will not only advance our understanding of fundamental cancer cell biology but also provide an important basis for the development of new therapeutic strategies and novel compounds to selectively eliminate cancer cells by targeting their unique metabolism. This article reviews several important metabolic alterations in cancer cells, with an emphasis on increased aerobic glycolysis (the Warburg effect) and glutamine addiction, and discusses the mechanisms that may contribute to such metabolic changes. In addition, metabolic alterations in cancer stem cells, mitochondrial metabolism and its influence on drug sensitivity, and potential therapeutic strategies and agents that target cancer metabolism are also discussed.
Assuntos
Glutamina/metabolismo , Glicólise/efeitos dos fármacos , Mitocôndrias/metabolismo , Neoplasias/metabolismo , Neoplasias/patologia , Antineoplásicos/uso terapêutico , Resistencia a Medicamentos Antineoplásicos , Genes Supressores de Tumor/fisiologia , Glicólise/fisiologia , Humanos , Mitocôndrias/genética , Mutação , Neoplasias/tratamento farmacológico , Células-Tronco Neoplásicas/metabolismo , Oncogenes/fisiologia , Fosforilação OxidativaRESUMO
Inhibiting glycolysis remains an aspirational approach for the treatment of cancer. We have previously identified a subset of cancers harbouring homozygous deletion of the glycolytic enzyme enolase (ENO1) that have exceptional sensitivity to inhibition of its redundant paralogue, ENO2, through a therapeutic strategy known as collateral lethality. Here, we show that a small-molecule enolase inhibitor, POMHEX, can selectively kill ENO1-deleted glioma cells at low-nanomolar concentrations and eradicate intracranial orthotopic ENO1-deleted tumours in mice at doses well-tolerated in non-human primates. Our data provide an in vivo proof of principle of the power of collateral lethality in precision oncology and demonstrate the utility of POMHEX for glycolysis inhibition with potential use across a range of therapeutic settings.
Assuntos
Antineoplásicos/uso terapêutico , Biomarcadores Tumorais/genética , Proteínas de Ligação a DNA/genética , Inibidores Enzimáticos/uso terapêutico , Neoplasias/tratamento farmacológico , Neoplasias/genética , Fosfopiruvato Hidratase/antagonistas & inibidores , Proteínas Supressoras de Tumor/genética , Animais , Linhagem Celular Tumoral , Feminino , Glioma/tratamento farmacológico , Glicólise/efeitos dos fármacos , Humanos , Macaca fascicularis , Masculino , Camundongos , Camundongos SCID , Fosfopiruvato Hidratase/genética , Medicina de Precisão , Deleção de Sequência , Relação Estrutura-Atividade , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
BACKGROUND: Atrial fibrillation (AF) is the most common cardiac arrhythmia, with substantial public health and economic impact on healthcare systems due to the prevention and management of thromboembolic and hemorrhagic complications. In Algeria, stroke is a leading cause of death, representing 15.6% of all deaths in 2012. Current data on the epidemiology and costs associated with non-valvular AF (NVAF) in Algeria are not available. METHODS: A three-step approach was undertaken to estimate the economic burden of NVAF in Algeria. First, a literature review identified the epidemiological burden of the disease. Second, expert clinicians practicing in Algerian hospitals were surveyed on consumed resources and unit costs of treatment and management of complications and prevention. Finally, these data were combined with event probabilities in an economic model to estimate the annual cost of NVAF prevention and complications for the Algerian healthcare system. RESULTS: Based on literature and demographics data, it was estimated that there are currently 187,686 subjects with NVAF in Algeria. Seventy per cent of this population was treated for prevention, half of which were controlled. Cost of prevention was estimated at 203 million DZD (1.5 million) for drugs and 349 million DZD (2.6 million) for examinations. Mean hospitalization costs for complications ranged between 123,500 and 435,500 DZD (910-3,209), according to the type and severity of complications. Hospitalization costs for thromboembolic and hemorrhagic complications were estimated at 8,313 million DZD (62 million), half of which was for untreated patients. Finally, the economic burden of NVAF was estimated at 8,865 million DZD (>65 million) annually. CONCLUSION: The economic burden of NVAF is important in Algeria, largely driven by untreated and INR-uncontrolled patients. There is a lack of information on the Algerian healthcare system that could increase uncertainty around this assessment, but it clearly establishes the importance of NVAF as a public health concern.
Assuntos
Fibrilação Atrial/complicações , Fibrilação Atrial/economia , Hemorragia/induzido quimicamente , Acidente Vascular Cerebral/etiologia , Tromboembolia/etiologia , Idoso , Idoso de 80 Anos ou mais , Argélia/epidemiologia , Anticoagulantes/efeitos adversos , Fibrilação Atrial/epidemiologia , Feminino , Hemorragia/economia , Humanos , Coeficiente Internacional Normatizado , Masculino , Pessoa de Meia-Idade , Modelos Econométricos , Medicina Estatal/economia , Medicina Estatal/estatística & dados numéricos , Acidente Vascular Cerebral/economia , Acidente Vascular Cerebral/prevenção & controle , Tromboembolia/economiaRESUMO
The trileaflet mitral valve is a very rare congenital malformation with three equal size leaflets and three papillary muscles. In this article, we report the first case of trileaflet mitral valve associated with a bicuspid aortic valve in a patient referred for management of infective endocarditis.
RESUMO
Inhibition of glycolysis is of great potential for the treatment of cancer. However, inhibitors of glycolytic enzymes with favorable pharmacological profiles have not been forthcoming. Due to the nature of their active sites, most high-affinity transition-state analogue inhibitors of glycolysis enzymes are highly polar with poor cell permeability. A recent publication reported a novel, non-active site inhibitor of the glycolytic enzyme Enolase, termed ENOblock (N-[2-[2-2-aminoethoxy)ethoxy]ethyl]4-4-cyclohexylmethyl)amino]6-4-fluorophenyl)methyl]amino]1,3,5-triazin-2-yl]amino]benzeneacetamide). This would present a major advance, as this is heterocyclic and fully cell permeable molecule. Here, we present evidence that ENOblock does not inhibit Enolase enzymatic activity in vitro as measured by three different assays, including a novel 31P NMR based method which avoids complications associated with optical interferences in the UV range. Indeed, we note that due to strong UV absorbance, ENOblock interferes with the direct spectrophotometric detection of the product of Enolase, phosphoenolpyruvate. Unlike established Enolase inhibitors, ENOblock does not show selective toxicity to ENO1-deleted glioma cells in culture. While our data do not dispute the biological effects previously attributed to ENOblock, they indicate that such effects must be caused by mechanisms other than direct inhibition of Enolase enzymatic activity.
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Benzamidas/farmacologia , Glicólise , Fosfopiruvato Hidratase/antagonistas & inibidores , Triazinas/farmacologia , Linhagem Celular Tumoral , Humanos , Fosfopiruvato Hidratase/metabolismoRESUMO
Deletion of chromosome 17p with a loss of p53 is an unfavorable cytogenetic change in chronic lymphocytic leukemia (CLL) with poor clinical outcome. Since p53 affects mitochondrial function and integrity, we examined possible mitochondrial changes in CLL mice with TCL1-Tg/p53-/- and TCL1-Tg/p53+/+ genotypes and in primary leukemia cells from CLL patients with or without 17p-deletion. Although the expression of mitochondrial COX1, ND2, and ND6 decreased in p53-/-CLL cells, there was an increase in mitochondrial biogenesis as evidenced by higher mitochondrial mass and mtDNA copy number associated with an elevated expression of TFAM and PGC-1α. Surprisingly, the overall mitochondrial respiratory activity and maximum reserved capacity increased in p53-/- CLL cells. Our study suggests that leukemia cells lacking p53 seem able to maintain respiratory function by compensatory increase in mitochondrial biogenesis.
Assuntos
Deleção Cromossômica , Leucemia Linfocítica Crônica de Células B/patologia , Biogênese de Organelas , Proteína Supressora de Tumor p53/deficiência , Animais , Respiração Celular , Humanos , Camundongos , Camundongos KnockoutRESUMO
INTRODUCTION: Cancer stem cells (CSCs) possess characteristics associated with normal stem cells, specifically the abilities to renew themselves and to give rise to all cell types (differentiation). It is assumed that induction of differentiation in CSCs would reduce their ability to form tumors. What triggers CSC differentiation and the role of "differentiation" in tumorigenesis remain elusive. METHODS: Glioma stem cell (GSC) lines and subcutaneous as well as orthotopic xenografts established from fresh surgical specimens of glioblastoma multiforme were used. RESULTS: Exposure of GSCs to serum activates mitochondrial respiration and causes an increase in mitochondrial reactive oxygen species (ROS) as well as oxidative stress responses, leading to the appearance of differentiation morphology and a deceased expression of CSC markers. Chemical perturbation of the mitochondrial electron transport chain causes ROS increase and further downregulation of stem cell markers, while antioxidant N-acetyl-cysteine reduces ROS and suppresses the differentiation of GSCs. Surprisingly, the serum-induced differentiated GSCs exhibit greater ability to form tumor in both orthotopic and subcutaneous xenograft models, which can be suppressed by N-acetyl-cysteine. Mitochondrial ROS from the serum-stimulated cells triggered the activation of nuclear factor-kappa-B (NFκB) pathway, which is a potential mechanism for the promotion of tumorigenesis. CONCLUSION: This study suggests that ROS generated from active mitochondrial respiration in the presence of serum is critical in CSCs activation, which promotes tumor development in vivo.
Assuntos
Glioma/metabolismo , Mitocôndrias/metabolismo , Células-Tronco Neoplásicas/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Ativação Metabólica , Animais , Antioxidantes/farmacologia , Neoplasias Encefálicas/metabolismo , Neoplasias Encefálicas/patologia , Diferenciação Celular , Linhagem Celular Tumoral , Meios de Cultura , Transporte de Elétrons , Glioblastoma/metabolismo , Glioma/patologia , Humanos , Camundongos , Camundongos Nus , Camundongos SCID , Mitocôndrias/efeitos dos fármacos , NF-kappa B/metabolismo , Células-Tronco Neoplásicas/efeitos dos fármacos , Células-Tronco Neoplásicas/patologia , Estresse Oxidativo , Transdução de SinaisRESUMO
AIMS: The transcription factor Islet-1 (ISL1) is a marker of cardiovascular progenitors and is essential for mammalian cardiogenesis. An ISL1 haplotype has recently been associated with congenital heart disease. In this study we evaluated whether ISL1 variants are associated with hypertrophic (HCM), dilated (DCM), arrhythmogenic right ventricular cardiomyopathy (ARVC), or with Emery-Dreifuss muscular dystrophy (EDMD). METHODS AND RESULTS: The six exon and intron boundaries of ISL1 were screened for genetic variants in a cohort of 454 index cases. Eleven exonic variants were identified in HCM, DCM, ARVC, and/or EDMD. Out of the five novel variants, two are located in the 5'-untranslated region, two are silent (p.Arg171Arg and p.Asn189Asn), and one is a missense (p.Asn252Ser). The latter was identified in the homozygous state in one DCM patient, and in the heterozygous state in 11 relatives, who did not present with DCM but often with cardiovascular features. This variant was found in one HCM patient also carrying a MYH7 mutation and in 3/96 North-African Caucasian control individuals, but was absent in 138 European Caucasian control individuals. We investigated the effect of the ISL1 wild type and p.Asn252Ser mutant on myocyte enhancer factor 2C (Mef2c) promoter activity, an established ISL1 target. Mef2c promoter activity was â¼4-fold higher in the presence of wild-type and â¼6-fold higher in the presence of mutant ISL1 in both HEK and CHO cells. CONCLUSION: This study describes a new gain-of-function p.Asn252Ser variant in the human ISL1 gene, which could potentially lead to greater activation of downstream targets involved in cardiac development, dilation, and hypertrophy.