Efficacy and safety of CDR132L in patients with reduced left ventricular ejection fraction after myocardial infarction: Rationale and design of the HF-REVERT trial.

AIM: Inhibition of microRNA (miR)-132 effectively prevents and reverses adverse cardiac remodelling, making it an attractive heart failure (HF) target. CDR132L, a synthetic antisense oligonucleotide selectively blocking pathologically elevated miR-132, demonstrated beneficial effects on left ventricular (LV) structure and function in relevant preclinical models, and was safe and well tolerated in a Phase 1b study in stable chronic HF patients. Patients with acute myocardial infarction (MI) and subsequent LV dysfunction and remodelling have limited therapeutic options, and may profit from early CDR132L treatment. METHODS: The HF-REVERT (Phase 2, multicenter, randomized, parallel, 3-arm, placebo-controlled Study to Assess Efficacy and Safety of CDR132L in Patients with Reduced Left Ventricular Ejection Fraction after Myocardial Infarction) evaluates the efficacy and safety of CDR132L in HF patients post-acute MI (n = 280), comparing the effect of 5 and 10 mg/kg CDR132L, administered as three single intravenous doses 28 days apart, in addition to standard of care. Key inclusion criteria are the diagnosis of acute MI, the development of systolic dysfunction (LV ejection fraction ≤45%) and elevated N-terminal pro-B-type natriuretic peptide. The study consists of a 6-month double-blinded treatment period with the primary endpoint LV end-systolic volume index and relevant secondary endpoints, followed by a 6-month open-label observation period. CONCLUSION: The HF-REVERT trial may underpin the concept of miR-132 inhibition to prevent or reverse cardiac remodelling in post-MI HF. The results will inform the design of subsequent outcome trials to test CDR132L in HF.

Classical Xanthinuria in Nine Israeli Families and Two Isolated Cases from Germany: Molecular, Biochemical and Population Genetics Aspects.

Classical xanthinuria is a rare autosomal recessive metabolic disorder caused by variants in the XDH (type I) or MOCOS (type II) genes. Thirteen Israeli kindred (five Jewish and eight Arab) and two isolated cases from Germany were studied between the years 1997 and 2013. Four and a branch of a fifth of these families were previously described. Here, we reported the demographic, clinical, molecular and biochemical characterizations of the remaining cases. Seven out of 20 affected individuals (35%) presented with xanthinuria-related symptoms of varied severity. Among the 10 distinct variants identified, six were novel: c.449G>T (p.(Cys150Phe)), c.1434G>A (p.(Trp478*)), c.1871C>G (p.(Ser624*)) and c.913del (p.(Leu305fs*1)) in the XDH gene and c.1046C>T (p.(Thr349Ileu)) and c.1771C>T (p.(Pro591Ser)) in the MOCOS gene. Heterologous protein expression studies revealed that the p.Cys150Phe variant within the Fe/S-I cluster-binding site impairs XDH biogenesis, the p.Thr349Ileu variant in the NifS-like domain of MOCOS affects protein stability and cysteine desulfurase activity, while the p.Pro591Ser and a previously described p.Arg776Cys variant in the C-terminal domain affect Molybdenum cofactor binding. Based on the results of haplotype analyses and historical genealogy findings, the potential dispersion of the identified variants is discussed. As far as we are aware, this is the largest cohort of xanthinuria cases described so far, substantially expanding the repertoire of pathogenic variants, characterizing structurally and functionally essential amino acid residues in the XDH and MOCOS proteins and addressing the population genetic aspects of classical xanthinuria.

CDR132L improves systolic and diastolic function in a large animal model of chronic heart failure.

AIMS: Cardiac miR-132 activation leads to adverse remodelling and pathological hypertrophy. CDR132L is a synthetic lead-optimized oligonucleotide inhibitor with proven preclinical efficacy and safety in heart failure (HF) early after myocardial infarction (MI), and recently completed clinical evaluation in a Phase 1b study (NCT04045405). The aim of the current study was to assess safety and efficacy of CDR132L in a clinically relevant large animal (pig) model of chronic heart failure following MI. METHODS AND RESULTS: In a chronic model of post-MI HF, slow-growing pigs underwent 90 min left anterior descending artery occlusion followed by reperfusion. Animals were randomized and treatment started 1-month post-MI. Monthly intravenous (IV) treatments of CDR132L over 3 or 5 months (3× or 5×) were applied in a blinded randomized placebo-controlled fashion. Efficacy was evaluated based on serial magnetic resonance imaging, haemodynamic, and biomarker analyses. The treatment regime provided sufficient tissue exposure and CDR132L was well tolerated. Overall, CDR132L treatment significantly improved cardiac function and reversed cardiac remodelling. In addition to the systolic recovery, diastolic function was also ameliorated in this chronic model of HF. CONCLUSION: Monthly repeated dosing of CDR132L is safe and adequate to provide clinically relevant exposure and therapeutic efficacy in a model of chronic post-MI HF. CDR132L thus should be explored as treatment for the broad area of chronic heart failure.

Novel antisense therapy targeting microRNA-132 in patients with heart failure: results of a first-in-human Phase 1b randomized, double-blind, placebo-controlled study.

AIMS: Cardiac microRNA-132-3p (miR-132) levels are increased in patients with heart failure (HF) and mechanistically drive cardiac remodelling processes. CDR132L, a specific antisense oligonucleotide, is a first-in-class miR-132 inhibitor that attenuates and even reverses HF in preclinical models. The aim of the current clinical Phase 1b study was to assess safety, pharmacokinetics, target engagement, and exploratory pharmacodynamic effects of CDR132L in patients on standard-of-care therapy for chronic ischaemic HF in a randomized, placebo-controlled, double-blind, dose-escalation study (NCT04045405). METHODS AND RESULTS: Patients had left ventricular ejection fraction between ≥30% and <50% or amino terminal fragment of pro-brain natriuretic peptide (NT-proBNP) >125 ng/L at screening. Twenty-eight patients were randomized to receive CDR132L (0.32, 1, 3, and 10 mg/kg body weight) or placebo (0.9% saline) in two intravenous infusions, 4 weeks apart in four cohorts of seven (five verum and two placebo) patients each. CDR132L was safe and well tolerated, without apparent dose-limiting toxicity. A pharmacokinetic/pharmacodynamic dose modelling approach suggested an effective dose level at ≥1 mg/kg CDR132L. CDR132L treatment resulted in a dose-dependent, sustained miR-132 reduction in plasma. Patients given CDR132L ≥1 mg/kg displayed a median 23.3% NT-proBNP reduction, vs. a 0.9% median increase in the control group. CDR132L treatment induced significant QRS narrowing and encouraging positive trends for relevant cardiac fibrosis biomarkers. CONCLUSION: This study is the first clinical trial of an antisense drug in HF patients. CDR132L was safe and well tolerated, confirmed linear plasma pharmacokinetics with no signs of accumulation, and suggests cardiac functional improvements. Although this study is limited by the small patient numbers, the indicative efficacy of this drug is very encouraging justifying additional clinical studies to confirm the beneficial CDR132L pharmacodynamic effects for the treatment of HF.

The Pyrazolo[3,4-d]pyrimidine Derivative, SCO-201, Reverses Multidrug Resistance Mediated by ABCG2/BCRP.

ATP-binding cassette (ABC) transporters, such as breast cancer resistance protein (BCRP), are key players in resistance to multiple anti-cancer drugs, leading to cancer treatment failure and cancer-related death. Currently, there are no clinically approved drugs for reversal of cancer drug resistance caused by ABC transporters. This study investigated if a novel drug candidate, SCO-201, could inhibit BCRP and reverse BCRP-mediated drug resistance. We applied in vitro cell viability assays in SN-38 (7-Ethyl-10-hydroxycamptothecin)-resistant colon cancer cells and in non-cancer cells with ectopic expression of BCRP. SCO-201 reversed resistance to SN-38 (active metabolite of irinotecan) in both model systems. Dye efflux assays, bidirectional transport assays, and ATPase assays demonstrated that SCO-201 inhibits BCRP. In silico interaction analyses supported the ATPase assay data and suggest that SCO-201 competes with SN-38 for the BCRP drug-binding site. To analyze for inhibition of other transporters or cytochrome P450 (CYP) enzymes, we performed enzyme and transporter assays by in vitro drug metabolism and pharmacokinetics studies, which demonstrated that SCO-201 selectively inhibited BCRP and neither inhibited nor induced CYPs. We conclude that SCO-201 is a specific, potent, and potentially non-toxic drug candidate for the reversal of BCRP-mediated resistance in cancer cells.

Preclinical development of a miR-132 inhibitor for heart failure treatment.

Despite proven efficacy of pharmacotherapies targeting primarily global neurohormonal dysregulation, heart failure (HF) is a growing pandemic with increasing burden. Treatments mechanistically focusing at the cardiomyocyte level are lacking. MicroRNAs (miRNA) are transcriptional regulators and essential drivers of disease progression. We previously demonstrated that miR-132 is both necessary and sufficient to drive the pathological cardiomyocytes growth, a hallmark of adverse cardiac remodelling. Therefore, miR-132 may serve as a target for HF therapy. Here we report further mechanistic insight of the mode of action and translational evidence for an optimized, synthetic locked nucleic acid antisense oligonucleotide inhibitor (antimiR-132). We reveal the compound's therapeutic efficacy in various models, including a clinically highly relevant pig model of HF. We demonstrate favourable pharmacokinetics, safety, tolerability, dose-dependent PK/PD relationships and high clinical potential for the antimiR-132 treatment scheme.

Identification of persulfide-binding and disulfide-forming cysteine residues in the NifS-like domain of the molybdenum cofactor sulfurase ABA3 by cysteine-scanning mutagenesis.

The Moco (molybdenum cofactor) sulfurase ABA3 from Arabidopsis thaliana catalyses the sulfuration of the Moco of aldehyde oxidase and xanthine oxidoreductase, which represents the final activation step of these enzymes. ABA3 consists of an N-terminal NifS-like domain that exhibits L-cysteine desulfurase activity and a C-terminal domain that binds sulfurated Moco. The strictly conserved Cys430 in the NifS-like domain binds a persulfide intermediate, which is abstracted from the substrate L-cysteine and finally needs to be transferred to the Moco of aldehyde oxidase and xanthine oxidoreductase. In addition to Cys4³°, another eight cysteine residues are located in the NifS-like domain, with two of them being highly conserved among Moco sulfurase proteins and, at the same time, being in close proximity to Cys4³°. By determination of the number of surface-exposed cysteine residues and the number of persulfide-binding cysteine residues in combination with the sequential substitution of each of the nine cysteine residues, a second persulfide-binding cysteine residue, Cys²°6, was identified. Furthermore, the active-site Cys4³° was found to be located on top of a loop structure, formed by the two flanking residues Cys4²8 and Cys4³5, which are likely to form an intramolecular disulfide bridge. These findings are confirmed by a structural model of the NifS-like domain, which indicates that Cys4²8 and Cys4³5 are within disulfide bond distance and that a persulfide transfer from Cys4³° to Cys²°6 is indeed possible.

Comparative Genomics and Evolution of Molybdenum Utilization.

The trace element molybdenum (Mo) is the catalytic component of important enzymes involved in global nitrogen, sulfur, and carbon metabolism in both prokaryotes and eukaryotes. With the exception of nitrogenase, Mo is complexed by a pterin compound thus forming the biologically active molybdenum cofactor (Moco) at the catalytic sites of molybdoenzymes. The physiological roles and biochemical functions of many molybdoenzymes have been characterized. However, our understanding of the occurrence and evolution of Mo utilization is limited. This article focuses on recent advances in comparative genomics of Mo utilization in the three domains of life. We begin with a brief introduction of Mo transport systems, the Moco biosynthesis pathway, the role of posttranslational modifications, and enzymes that utilize Mo. Then, we proceed to recent computational and comparative genomics studies of Mo utilization, including a discussion on novel Moco-binding proteins that contain the C-terminal domain of the Moco sulfurase and that are suggested to represent a new family of molybdoenzymes. As most molybdoenzymes need additional cofactors for their catalytic activity, we also discuss interactions between Mo metabolism and other trace elements and finish with an analysis of factors that may influence evolution of Mo utilization.