Carol Berry

Preclinical pharmacology of lumiracoxib: a novel selective inhibitor of cyclooxygenase-2

1 This manuscript presents the preclinical profile of lumiracoxib, a novel cyclooxygenase‐2 (COX‐2) selective inhibitor. 2 Lumiracoxib inhibited purified COX‐1 and COX‐2 with Ki values of 3 and 0.06 μM, respectively. In cellular assays, lumiracoxib had an IC50 of 0.14 μM in COX‐2‐expressing dermal fibroblasts, but caused no inhibition of COX‐1 at concentrations up to 30 μM (HEK 293 cells transfected with human COX‐1). 3 In a human whole blood assay, IC50 values for lumiracoxib were 0.13 μM for COX‐2 and 67 μM for COX‐1 (COX‐1/COX‐2 selectivity ratio 515). 4 Lumiracoxib was rapidly absorbed following oral administration in rats with peak plasma levels being reached between 0.5 and 1 h. 5 Ex vivo, lumiracoxib inhibited COX‐1‐derived thromboxane B2 (TxB2) generation with an ID50 of 33 mg kg−1, whereas COX‐2‐derived production of prostaglandin E2 (PGE2) in the lipopolysaccharide‐stimulated rat air pouch was inhibited with an ID50 value of 0.24 mg kg−1. 6 Efficacy of lumiracoxib in rat models of hyperalgesia, oedema, pyresis and arthritis was dose‐dependent and similar to diclofenac. However, consistent with its low COX‐1 inhibitory activity, lumiracoxib at a dose of 100 mg kg−1 orally caused no ulcers and was significantly less ulcerogenic than diclofenac (P<0.05). 7 Lumiracoxib is a highly selective COX‐2 inhibitor with anti‐inflammatory, analgesic and antipyretic activities comparable with diclofenac, the reference NSAID, but with much improved gastrointestinal safety.

Non-peptidic inhibitors of neutral endopeptidase 24.11 1. Discovery and optimization of potency

Abstract Although based on a single α-amino acid residue, N -phosphonomethyl-(S)-(4-phenyl)phenylalanine ( 2 ) was discovered to produce strong inhibition of the zinc metalloprotease, neutral endopeptidase (NEP 24.11). Structural optimization of this new lead culminated with the design of the phosphonic acid tetrazole 17 (CGS 26303), a non-peptidic and extremely potent NEP inhibitor.

Non-peptidic inhibitors of neutral endopeptidase 24.11 2. Design and pharmacology of orally active phosphonate prodrugs

Abstract Prodrugs of 1 (CGS 26303), a novel non-peptidic neutral endopeptidase (NEP 24.11) inhibitor, have been designed. In particular, the diphenylphosphonate 6 (CGS 26393) was found to be a potent, orally bioavailable and long-acting NEP inhibitor. Accordingly, 6 protected atrial natriuretic peptide from enzymatic degradation and displayed a significant antihypertensive activity in the DOCA-salt rat.

Dual inhibition of neutral endopeptidase and angiotensin-converting enzyme by N-phosphonomethyl and N-carboxyalkyl dipeptides

Abstract Structural modifications have been performed on the selective α-amino phosphonic acid NEP inhibitor CGS 24592, to achieve dual ACE/NEP inhibition in vitro. (S)-N-Phosphonomethyl-valyl-(S)-(4-phenyl)phenylalanine (19) is representative of a new type of phosphorus-containing ACE/NEP inhibitors, approaching the in vitro potency of the sulfhydryl ACE inhibitor captopril and the NEP inhibitor thiorphan.

Characterisation of Neutral Endopeptidase 3.4.24.11 (NEP) in the Kidney: Comparison Between Normotensive, Genetically Hypertensive and Experimentally Hypertensive Rats

Neutral endopeptidase 3.4.24.11 (NEP) has been identified as the major atrial natriuretic factor (ANF) degrading enzyme in rat kidney, therefore, suggesting a possible role for this enzyme in blood volume and pressure regulation. Various experimentally induced and genetically hypertensive rat models have been used to test NEP inhibitors. The presence of different isoforms of NEP in the various hypertensive rat models would have relevance when searching for novel NEP inhibitors. Therefore, we compared the properties of NEP in kidney cortex homogenates in order to test for possible differences in the following hypertensive rat models and their appropriate controls: spontaneously hypertensive rats (SHR), Wistar Kyoto strain (WKY), DOCA-salt hypertensive rats, and Sprague Dawley control rats (SD). No relevant differences were found when comparing the following parameters: (1) specific activity (mean: 204 U/mg protein), (2) Michaelis constant (mean: 280 microM), (3) IC50 of thiorphan (mean: 6.5 nM) and phosphoramidon (mean: 54 nM), (4) pH profiles (optimum at pH 8.0), (5) heat inactivation profiles (half-life 20 min at 65 degrees C), (6) immunotitration of kidney cortex homogenates, (7) molecular weight as determined by gel filtration (92,000 Dalton) and (8) affinity chromatography with concanavalin A. Without evidence for the presence of different NEP isoforms, it is unlikely that divergent findings in DOCA-salt rats and SHR using a given NEP inhibitor are due to isoforms of NEP.

α-Mercaptoacyl dipeptides that inhibit angiotensin converting enzyme and neutral endopeptidase 24.11

Abstract α-Mercaptoacyl dipeptides were prepared and found to inhibit angiotensin converting enzyme (ACE) and neutral endopeptidase 24.11 (NEP). Compounds with a proline as the C-terminal amino acid, and possessing the S-stereochemistry at the thiol bearing carbon were preferred for optimal ACE inhibition while, R-stereochemistry was preferred for optimal NEP inhibition. Compounds containing alternative amino acid residues at the C-terminal position displayed optimal dual ACE/NEP inhibition when the stereochemistry was ‘S’ at this carbon atom.

Dual inhibition of angiotensin-converting enzyme and neutral endopeptidase by tricyclic benzazepinone thiols §

Abstract Various thioacyl analogs of CGS 28106, a tricyclic dual inhibitor of angiotensin-converting enzyme and neutral endopeptidase, have been synthesized and their inhibitory potencies evaluated in vitro. The structure-activity relationship supports the proposed hypothesis that, despite its conformational constraints, CGS 28106 can inhibit the two distinct metalloproteases by adopting different binding modes. In addition, the structural features of CGS 28106 confer remarkable oral activity to this dual inhibitor, as measured by its ability to block the angiotensin-I pressor response and to potentiate plasma levels of atrial natriuretic peptide.

Differential Cleavage of Urodilatin and Atrial Natriuretic Factor by Thrombin and Protease 3.4.24.11

AbstractHuman urodilatin (residues 95–126) and atrial natriuretic factor (residues 99–126, based on ANF prohor-mone sequence) were incubated separately with three proteases, thrombin, angiotensin converting enzyme (ACE), and neutral endopeptidase 3.4.24.11 (NEP). Thrombin cleaved urodilatin on the carboxyl side of arginine98 to yield ANF but under the same conditions did not cleave h-ANF. Neither urodilatin nor ANF was cleaved by ACE. ANF was rapidly degraded by NEP resulting in a major product cleaved between amino acid residues Cysl05 and Phe106. Urodilatin was also cleaved by NEP and the amino acid sequencing of the cleaved product revealed the site of cleavage to be the same Cys105-Phe106 site as for ANF with a second cleavage site at Gly118-Leu119. However, cleavage of urodilatin by NEP proceeded much more slowly when compared to ANF. A comparison of the affinities of ANF and urodilatin for purified NEP from rabbit kidney revealed Km values of 11.7 and 3.1 μM, respectively. The turnover rates (kcat/K...