Cort S. Madsen

(3R,5S,E)-7-(4-(4-Fluorophenyl)-6-isopropyl-2-(methyl(1-methyl-1H-1,2,4-triazol-5-yl)amino)pyrimidin-5-yl)-3,5-dihydroxyhept-6-enoic Acid (BMS-644950): A Rationally Designed Orally Efficacious 3-Hydroxy-3-methylglutaryl Coenzyme-A Reductase Inhibitor with Reduced Myotoxicity Potential

3-hydroxy-3-methylglutaryl coenzyme-A reductase (HMGR) inhibitors, more commonly known as statins, represent the gold standard in treating hypercholesterolemia. Although statins are regarded as generally safe, they are known to cause myopathy and, in rare cases, rhabdomyolysis. Statin-dependent effects on plasma lipids are mediated through the inhibition of HMGR in the hepatocyte, whereas evidence suggests that myotoxicity is due to inhibition of HMGR within the myocyte. Thus, an inhibitor with increased selectivity for hepatocytes could potentially result in an improved therapeutic window. Implementation of a strategy that focused on in vitro potency, compound polarity, cell selectivity, and oral absorption, followed by extensive efficacy and safety modeling in guinea pig and rat, resulted in the identification of compound 1b (BMS-644950). Using this discovery pathway, we compared 1b to other marketed statins to demonstrate its outstanding efficacy and safety profile. With the potential to generate an excellent therapeutic window, 1b was advanced into clinical development.

The Guinea Pig as a Preclinical Model for Demonstrating the Efficacy and Safety of Statins

Statins, because of their excellent efficacy and manageable safety profile, represent a key component in the current armamentarium for the treatment of hypercholesterolemia. Nonetheless, myopathy remains a safety concern for this important drug class. Cerivastatin was withdrawn from the market for myotoxicity safety concerns. BMS-423526 [{(3R,5S)-7-[4-(4-fluorophenyl)-6,7-dihydro-2-(1-methylethyl)-5H-benzo[6,7]cyclohepta[1,2-b]pyridin-3-yl]-3,5-dihydroxy-heptenoic acid} sodium salt], similar to cerivastatin in potency and lipophilicity, was terminated in early clinical development due to an unacceptable myotoxicity profile. In this report, we describe the guinea pig as a model of statin-induced cholesterol lowering and myotoxicity and show that this model can distinguish statins with unacceptable myotoxicity profiles from statins with acceptable safety profiles. In our guinea pig model, both cerivastatin and BMS-423526 induced myotoxicity at doses near the ED50 for total cholesterol (TC) lowering in plasma. In contrast, wide differences between myotoxic and TC-lowering doses were established for the currently marketed, more hydrophilic statins, pravastatin, rosuvastatin, and atorvastatin. This in vivo model compared favorably to an in vitro model, which used statin inhibition of cholesterol synthesis in rat hepatocytes and L6 myoblasts as surrogates of potential efficacy and toxicity, respectively. Our conclusion is that the guinea pig is a useful preclinical in vivo model for demonstrating whether a statin is likely to have an acceptable therapeutic safety margin.

Pyrazole-based sulfonamide and sulfamides as potent inhibitors of mammalian 15-lipoxygenase.

A series of inhibitors of mammalian 15-lipoxygenase (15-LO) based on a 3,4,5-tri-substituted pyrazole scaffold is described. Replacement of a sulfonamide functionality in the lead series with a sulfamide group resulted in improved physicochemical properties generating analogs with enhanced inhibition in cell-based and whole blood assays.

Genomic footprinting of mitochondrial DNA: II. In vivo analysis of protein-mitochondrial DNA interactions in Xenopus laevis eggs and embryos.

Publisher Summary This chapter describes a simple footprinting protocol, which yields reproducible, high-resolution genomic footprints from small amounts of mtDNA templates. This is generally applicable to study mitochondrial gene regulation in a wide variety of experimental samples where mitochondrial metabolism can be modified at will. Hence, genomic footprinting of mitochondrial DNA is an important tool in understanding mitochondrial gene expression. One important factor in obtaining useful footprint information is the uniformity of the cell or tissue sample. Heterogeneity in cell lineage or cell stage may lead to masking or overrepresentation of footprint data, leading to inaccurate conclusions. The advantage of this technique is that it is readily adaptable to other systems where the amount of DNA available does not warrant developing a litigation mediated polymerase chain reaction (LMPCR) protocol for genomic footprinting. This primer extension footprinting (PEF) protocol has been used to perform in vitro dimethyl sulfate (DMS) footprinting using recombinant plasmids and mitochondrial protein fractions.

Genomic footprinting of mitochondrial DNA: I. In organello analysis of protein-mitochondrial DNA interactions in bovine mitochondria.

Publisher Summary In organello footprinting involves the use of dimethyl sulfate (DMS) in a methylation protection assay, first used to analyze protein–nuclear DNA interactions in vivo . DMS is a small molecule that methylates guanine residues at the N-7 position and adenines at the N-3 position, making them sensitive to subsequent cleavage at alkaline pH and elevated temperatures. The ability of DMS to permeate mitochondrial membranes readily allows detection of protein–mtDNA interaction within the organelle. During development of the in organello technique, bovine brain cortex mitochondria is used to monitor protein–mtDNA interactions at several domains within the mitochondrial genome considered important in regulating transcriptional and replicative processes.