Christoph Hirschhäuser

Paraoxonase-1 is a major determinant of clopidogrel efficacy

Clinical efficacy of the antiplatelet drug clopidogrel is hampered by its variable biotransformation into the active metabolite. The variability in the clinical response to clopidogrel treatment has been attributed to genetic factors, but the specific genes and mechanisms underlying clopidogrel bioactivation remain unclear. Using in vitro metabolomic profiling techniques, we identified paraoxonase-1 (PON1) as the crucial enzyme for clopidogrel bioactivation, with its common Q192R polymorphism determining the rate of active metabolite formation. We tested the clinical relevance of the PON1 Q192R genotype in a population of individuals with coronary artery disease who underwent stent implantation and received clopidogrel therapy. PON1 QQ192 homozygous individuals showed a considerably higher risk than RR192 homozygous individuals of stent thrombosis, lower PON1 plasma activity, lower plasma concentrations of active metabolite and lower platelet inhibition. Thus, we identified PON1 as a key factor for the bioactivation and clinical activity of clopidogrel. These findings have therapeutic implications and may be exploited to prospectively assess the clinical efficacy of clopidogrel.

Intramolecular 1,6-Addition to 2 Pyridones. Mechanism and Synthetic Scope

The scope and limitations of the intramolecular 1,6-addition of an enolate to a 2-pyridone moiety, a reaction that has found application in the synthesis of the lupin alkaloids, have been probed. This nucleophilic addition process has been shown to be reversible and favored in the case of (less stabilized) amide and lactam enolates, which readily form five- and six-membered bi-/tricyclic products. Alternative enolates (ketone, ester, thiolactam) and a variety of different acceptors (isoquinolinone, pyrimidinone, pyrazinone, pyridopyrazinone) have been evaluated, and a range of competing side reactions have been identified and characterized using various techniques, including in situ IR.

Organometallic nucleosides induce non-classical leukemic cell death that is mitochondrial-ROS dependent and facilitated by TCL1-oncogene burden

BackgroundRedox stress is a hallmark of the rewired metabolic phenotype of cancer. The underlying dysregulation of reactive oxygen species (ROS) is interconnected with abnormal mitochondrial biogenesis and function. In chronic lymphocytic leukemia (CLL), elevated ROS are implicated in clonal outgrowth and drug resistance. The pro-survival oncogene T-cell leukemia 1 (TCL1) is causally linked to the high threshold towards classical apoptosis in CLL. We investigated how aberrant redox characteristics and bioenergetics of CLL are impacted by TCL1 and if this is therapeutically exploitable.MethodsBio-organometallic chemistry provided compounds containing a cytosine nucleobase, a metal core (ferrocene, ruthenocene, Fe(CO)3), and a 5’-CH2O-TDS substituent. Four of these metal-containing nucleoside analogues (MCNA) were tested for their efficacy and mode of action in CLL patient samples, gene-targeted cell lines, and murine TCL1-transgenic splenocytes.ResultsThe MCNA showed a marked and selective cytotoxicity towards CLL cells. MCNA activity was equally observed in high-risk disease groups, including those of del11q/del17p cytogenetics and of clinical fludarabine resistance. They overcame protective stromal cell interactions. MCNA-evoked PARP-mediated cell death was non-autophagic and non-necrotic as well as caspase- and P53-independent. This unconventional apoptosis involved early increases of ROS, which proved indispensible based on mitigation of MCNA-triggered death by various scavengers. MCNA exposure reduced mitochondrial respiration (oxygen consumption rate; OCR) and induced a rapid membrane depolarization (∆ΨM). These characteristics distinguished the MCNA from the alkylator bendamustine and from fludarabine. Higher cellular ROS and increased MCNA sensitivity were linked to TCL1 expression. The presence of TCL1 promoted a mitochondrial release of in part caspase-independent apoptotic factors (AIF, Smac, Cytochrome-c) in response to MCNA. Although basal mitochondrial respiration (OCR) and maximal respiratory capacity were not affected by TCL1 overexpression, it mediated a reduced aerobic glycolysis (lactate production) and a higher fraction of oxygen consumption coupled to ATP-synthesis.ConclusionsRedox-active substances such as organometallic nucleosides can confer specific cytotoxicity to ROS-stressed cancer cells. Their P53- and caspase-independent induction of non-classical apoptosis implicates that redox-based strategies can overcome resistance to conventional apoptotic triggers. The high TCL1-oncogenic burden of aggressive CLL cells instructs their particular dependence on mitochondrial energetic flux and renders them more susceptible towards agents interfering in mitochondrial homeostasis.

Reply to: "Paraoxonase-1 and clopidogrel efficacy"

Jordi Camps1, Jorge Joven1, Bharti Mackness1, Michael Mackness1, Dan Tawfik2, Dragomir Draganov3, Lucio G Costa4, György Paragh5, Ildikó Seres5, Sven Horke6, Richard James7, Antonio Hernández8, Srinivasa Reddy9, Diana Shih9, Mohamed Navab9, Daniel Rochu10 & Michael Aviram11 1Centre de Recerca Biomèdica, Institut d’Investigació Sanitària Pere Virgili, Universitat Rovira i Virgili, C. Sant Joan, Reus, Spain. 2Department of Biological Chemistry, Weizmann Institute of Science, Rehovot, Israel. 3WIL Research Laboratories, Ashland, Ohio, USA. 4Department of Environmental and Occupational Health Sciences, University of Washington, Seattle, Washington, USA. 5Department of Internal Medicine, University of Debrecen, Hungary. 6Institute of Pharmacology, University Medical Center, Meinz, Obere Zahlbacher, Meinz, Germany. 7Department of Internal Medicine, University of Geneva, Switzerland. 8Department of Toxicology, University of Granada, Granada, Spain. 9Division of Cardiology, University of California–Los Angeles, Los Angeles, California, USA. 10Département de Toxicologie, Centre de Recherches du Service de Santé des Armées, La Tronche Cedex, France. 11The Lipid Research Laboratory, Technion Faculty of Medicine, Rambam Medical Center, Haifa, Israel. e-mail: jcamps@grupsagessa.cat

Protein Surface Recognition by Synthetic Molecules

Rational design of synthetic molecules with the aim of inhibiting protein–protein interactions (PPIs) that are key players in human diseases and biology, opens a new perspective in drug discovery. Here we describe synthetic molecules including α-helical mimitics, peptide foldamers, dendrimers, molecular tweezers, metal complexes, calixarene, and porphyrin-based small molecules that efficiently bind to protein surfaces and inhibit PPI.