PK/PD Disconnect Observed with a Reversible Endothelial Lipase Inhibitor

Screening of a small set of nonselective lipase inhibitors against endothelial lipase (EL) identified a potent and reversible inhibitor, <i>N</i>-(3-(3,4-dichlorophenyl)­propyl)-3-hydroxy-1-methyl-2-oxo-1,2-dihydropyridine-4-carboxamide (<b>5</b>; EL IC₅₀ = 61 nM, EL_HDL IC₅₀ = 454 nM). Deck mining identified a related hit, <i>N</i>-(3-(3,4-dichlorophenyl)­propyl)-4-hydroxy-1-methyl-5-oxo-2,5-dihydro-1<i>H</i>-pyrrole-3-carboxamide (<b>6a</b>; EL IC₅₀ = 41 nM, EL_HDL IC₅₀ = 1760 nM). Both compounds were selective against lipoprotein lipase (LPL) but nonselective versus hepatic lipase (HL). Optimization of compound <b>6a</b> for EL inhibition using HDL as substrate led to <i>N</i>-(4-(3,4<b>-</b>dichlorophenyl)­butan-2-yl)-1-ethyl-4-hydroxy-5-oxo-2,5-dihydro-1<i>H</i>-pyrrole-3-carboxamide (<b>7c</b>; EL IC₅₀ = 148 nM, EL_HDL IC₅₀ = 218 nM) having improved PK over compound <b>6a</b>, providing a tool molecule to test for the ability to increase HDL-cholesterol (HDL-C) levels in vivo using a reversible EL inhibitor. Compound <b>7c</b> did not increase HDL-C in vivo despite achieving plasma exposures targeted on the basis of enzyme activity and protein binding demonstrating the need to develop more physiologically relevant in vitro assays to guide compound progression for in vivo evaluation

Triphenylethanamine Derivatives as Cholesteryl Ester Transfer Protein Inhibitors: Discovery of <i>N</i>‑[(1<i>R</i>)‑1-(3-Cyclopropoxy-4-fluorophenyl)-1-[3-fluoro-5-(1,1,2,2-tetrafluoroethoxy)­phenyl]-2-phenylethyl]-4-fluoro-3-(trifluoromethyl)­benzamide (BMS-795311)

Cholesteryl ester transfer protein (CETP) inhibitors raise HDL-C in animals and humans and may be antiatherosclerotic by enhancing reverse cholesterol transport (RCT). In this article, we describe the lead optimization efforts resulting in the discovery of a series of triphenylethanamine (TPE) ureas and amides as potent and orally available CETP inhibitors. Compound <b>10g</b> is a potent CETP inhibitor that maximally inhibited cholesteryl ester (CE) transfer activity at an oral dose of 1 mg/kg in human CETP/apoB-100 dual transgenic mice and increased HDL cholesterol content and size comparable to torcetrapib (<b>1</b>) in moderately-fat fed hamsters. In contrast to the off-target liabilities with <b>1</b>, no blood pressure increase was observed with <b>10g</b> in rat telemetry studies and no increase of aldosterone synthase (CYP11B2) was detected in H295R cells. On the basis of its preclinical profile, compound <b>10g</b> was advanced into preclinical safety studies

Triphenylethanamine Derivatives as Cholesteryl Ester Transfer Protein Inhibitors: Discovery of <i>N</i>‑[(1<i>R</i>)‑1-(3-Cyclopropoxy-4-fluorophenyl)-1-[3-fluoro-5-(1,1,2,2-tetrafluoroethoxy)­phenyl]-2-phenylethyl]-4-fluoro-3-(trifluoromethyl)­benzamide (BMS-795311)

Cholesteryl ester transfer protein (CETP) inhibitors raise HDL-C in animals and humans and may be antiatherosclerotic by enhancing reverse cholesterol transport (RCT). In this article, we describe the lead optimization efforts resulting in the discovery of a series of triphenylethanamine (TPE) ureas and amides as potent and orally available CETP inhibitors. Compound <b>10g</b> is a potent CETP inhibitor that maximally inhibited cholesteryl ester (CE) transfer activity at an oral dose of 1 mg/kg in human CETP/apoB-100 dual transgenic mice and increased HDL cholesterol content and size comparable to torcetrapib (<b>1</b>) in moderately-fat fed hamsters. In contrast to the off-target liabilities with <b>1</b>, no blood pressure increase was observed with <b>10g</b> in rat telemetry studies and no increase of aldosterone synthase (CYP11B2) was detected in H295R cells. On the basis of its preclinical profile, compound <b>10g</b> was advanced into preclinical safety studies

Identification of regulatory deletions telomeric to <i>PAX6</i>.

<p>Regulatory deletions telomeric to <i>PAX6</i> were identified in individual 1514 (chr11:30,874,642–31,654,833), individual 753 (chr11:30,967,000–31,704,000), individual 555 (chr11:31,108,579–31,649–842), individual 2014 (chr11:31,234,395–31,751,815) and individual 659 (chr11:31,379,000–31,708,000). The schematic diagram shows how the ‘critical region’ (delimited by grey dotted lines) required for <i>PAX6</i> transcriptional activation was delineated by combining our data with published deletions with known coordinates [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0153757#pone.0153757.ref055" target="_blank">55</a>,<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0153757#pone.0153757.ref067" target="_blank">67</a>,<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0153757#pone.0153757.ref068" target="_blank">68</a>]. <i>PAX6</i> regulatory deletions from the present study are shown by red blocks. Genes transcribed on the forward strand are in blue and those transcribed on the reverse strand are in green, also indicated by arrows. Genomic coordinates are based on the Human Genome Assembly hg18.</p

Identification of a potential <i>PITX2</i> regulatory deletion.

<p>Genome-wide array CGH identified a deletion of approximately 3.5 Mb in individual 1194 (chr4:111,994,000–115,504,000) (red bar). The deletion is located telomeric to the <i>PITX2</i> gene on chromosome 4. The positions of conserved elements (CE) in the deleted region, as identified by Volkmann et al., 2011 [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0153757#pone.0153757.ref047" target="_blank">47</a>] are marked by orange ellipses. Genes transcribed on the forward strand are in blue and those transcribed on the reverse strand are in green, also indicated by arrows. Genomic coordinates are shown on the x-axis and are based on the Human Genome Assembly hg18.</p

Details of the clinical diagnoses and genetic pathology identified in individuals in this study.

<p>Details of the clinical diagnoses and genetic pathology identified in individuals in this study.</p

Mutation analysis of the <i>FOXC1</i> locus.

<p><b>(A)</b> Genome-wide array CGH identified two deletions encompassing the <i>FOXC1</i> gene in individuals 1449 (chr6:1,543,591–1,675,085) and 1246 (chr6:1,543,591–1,675,085). <b>(B)</b> Direct sequencing of the <i>FOXC1</i> coding region identified a heterozygous substitution in individual 1839 (c.235C>A, p.(Pro79Thr)) and another in individual 1634 (c.302T>C, p.(Leu101Pro)). <i>FOXC1</i> mutation screening in unaffected parents of both patients showed that the mutations had occurred <i>de novo</i>. The locations of both mutations within the fork-head domain of the FOXC1 protein are indicated by vertical arrows. Genes transcribed on the forward strand are in blue and those transcribed on the reverse strand are in green, also indicated by arrows. Genomic coordinates are based on the Human Genome Assembly hg18. The genomic sequence identifier for <i>FOXC1</i> is NG_009368.</p

Identification of <i>PAX6</i> whole-gene deletions.

<p>Genome-wide array CGH analysis identified a 650 kb deletion in individual 2193 (chr11:31,199,000–31,849,000), a 520 kb deletion in individual 377 (chr11:31,394,000–31,914,000), a 154 kb deletion in individual 1510 (chr11:31,779,000–31,933,000) and a 96 kb deletion in individual 1977 (chr11:31,698,271–31,794,414), all involving <i>PAX6</i>. Red bars show the position of the deletions. Genes transcribed on the forward strand are in blue and those transcribed on the reverse strand are in green, also indicated by arrows. Genomic coordinates are based on the Human Genome Assembly hg18.</p