Barry Kaye

Measurement of the class III antidysrhythmic drug, UK-68,798, in plasma by radioimmunoassay.

A sensitive radioimmunoassay (RIA) for the specific determination of 1-(4-methanesulphonamidophenoxy)-2-[N-(4-methanesulphonamido -phenethyl)-N- methylamino]ethane (UK-68,798), a novel class III antidysrhythmic agent, in human plasma is described. Specific antisera were raised in sheep using desmesyl-UK-68,798-succinate-ovalbumin conjugate as the antigenic hapten carrier protein. The antisera produced exhibited high specificity for UK-68,798 compared with known metabolites from animals, other antidysrhythmic agents and co-administered drugs. Good correlation was found in a comparison of the RIA method with a high-performance liquid chromatography (HPLC) method (r = 0.997) and a 10-fold lower limit of determination was observed for the RIA method compared with the HPLC method (0.05 and 0.5 ng ml-1, respectively). The RIA method was applied to the analysis of UK-68,798 in plasma obtained from human volunteers receiving the compound.

Clinical analysis of sampatrilat, a combined renal endopeptidase and angiotensin-converting enzyme inhibitor I : Assay in plasma of human volunteers by atmospheric-pressure ionisation mass-spectrometry following derivatisation with BF3-methanol

Sampatrilat is a dual inhibitor of angiotensin converting enzyme (ACE) and neutral endopeptidase (NEP) under development for the treatment of hypertension and congestive heart failure. In order to support the early clinical development (with oral administration and an expected low bioavailability), a sensitive and selective assay was required. A method for plasma was developed and validated employing HPLC APCI MS-MS. The plasma samples were extracted on solid-phase extraction cartridges, derivatised with BF3-methanol, diluted, extracted again and then subjected to HPLC APCI-MS-MS. Derivatisation was necessary because the two carboxyl group in the molecule prevented efficient ionisation in the heated nebuliser source. The calibration range was from 0.5 to 20 ng ml(-1) and the lower limit of quantification was 0.5 ng ml(-1). Imprecision and inaccuracy were determined on three separate occasions at three concentrations (0.5, 5 and 20 ng ml[-1]) and shown to be lower than 10% in every case.

Clinical analysis of sampatrilat, a combined renal endopeptidase and angiotensin-converting enzyme inhibitor: II: Assay in the plasma and urine of human volunteers by dissociation enhanced lanthanide fluorescence immunoassay (DELFIA)

Sampatrilat is a dual inhibitor of angiotensin converting enzyme (ACE) and neutral endopeptidase (NEP) under development for the treatment of hypertension and congestive heart failure. In order to support the early clinical development (with oral administration and an expected low bioavailability), a sensitive and selective assay was required. An HPLC-atmospheric-pressure chemical ionisation mass-spectrometric (HPLC-APCI-MS-MS) assay had been already validated (R.F. Venn et al., J. Pharm. Biomed. Anal., in press), but due to its low throughput an alternative method was sought. As the molecule is peptide-like and not metabolised, we believed the immunoassay approach was appropriate. Thus we developed an immunoassay for the compound using time-resolved fluorescence as an end-point (DELFIA) with lower limits of quantification of 0.2 ng ml(-1) for the plasma assay and 5 ng ml(-1) for the assay in urine. This assay is a 96-well plate based competitive immunoassay; the end-point is the determination of a (non-radioactive) europium label by time-resolved fluorimetry. Sampatrilat is labelled with chelated europium via isothiocyanate chemistry. The advantage of this assay is its extremely high throughput, allowing rapid analysis of many thousands of samples. The DELFIA method was successfully cross-validated with the HPLC-APCI-MS-MS method.