Design and synthesis of endocannabinoid enzyme inhibitors for ocular indications.

A small library of FAAH and dual FAAH/MAGL inhibitors designed for peripheral selectivity were targeted. Of these compounds, three were identified to have desirable FAAH inhibition and reduced permeability in a PAMPA assay. Those three compounds were advanced into a MAGL inhibitor assay and one was found to be a relative selective FAAH inhibitor, FAAH to MAGL IC50 ratio of 1:27, and one was found to be more characteristic of a true dual enzyme inhibitor, FAAH to MAGL IC50 ratio of 1:4. Both compounds showed activity in an ABPP assay, blockage of TAMRA-FP labeling of FAAH and MAGL in rat eye homogenate.

Multisite analytical evaluation of the Abbott ARCHITECT cyclosporine assay.

The objective of this study was to evaluate the analytical performance of the Abbott ARCHITECT Cyclosporine (CsA) immunoassay in 7 clinical laboratories in comparison to liquid chromatography/tandem mass spectrometry (LC/MS/MS), Abbott TDx, Cobas Integra 800, and the Dade Dimension Xpand immunoassay. The ARCHITECT assay uses a whole blood specimen, a pretreatment step with organic reagents to precipitate proteins and extract the drug, followed by a 2-step automated immunoassay with magnetic microparticles coated with anti-CsA antibody and an acridinium-CsA tracer. Imprecision testing at the 7 evaluation sites gave a range of total % coefficient of variations of 7.5%-12.2% at 87.5 ng/mL, 6.6%-14.3% at 411 ng/mL, and 5.2%-10.7% at 916 ng/mL. The lower limit of quantification ranged from 12 to 20 ng/mL. Purified CsA metabolites AM1, AM1c, AM4N, AM9, and AM19 were tested in whole blood by the ARCHITECT assay and showed minimal cross-reactivity at all 7 sites. In particular, AM1 and AM9 cross-reactivity in the ARCHITECT assay, ranged from -2.5% to 0.2% and -0.8% to 2.2%, respectively, and was significantly lower than for the TDx assay, in which the values were 3.2% and 16.1%, respectively. Comparable testing of metabolites in the Dade Dimension Xpand assay at 2 evaluation sites showed cross-reactivity to AM4N (6.4% and 6.8%) and AM9 (2.6% and 3.6%) and testing on the Roche Integra 800 showed cross-reactivity to AM1c (2.4%), AM9 (10.7%), and AM19 (2.8%). Cyclosporine International Proficiency Testing Scheme samples, consisting of both pooled specimens from patients receiving CsA therapy as well as whole-blood specimens supplemented with CsA, were tested by the ARCHITECT assay at 6 sites and showed an average bias of -24 to -58 ng/mL versus LC/MSMS CsA and -2 to -37 ng/mL versus AxSYM CsA. Studies were performed with the ARCHITECT CsA assay on patient specimens with the following results: ARCHITECT CsA assay versus LC/MSMS, average bias of 31 ng/mL; ARCHITECT versus the Dade Dimension assay (4 sites), average biases of -7 to -228 ng/mL; ARCHITECT versus AxSYM and TDx, average biases of -4 and -53 ng/mL, respectively. Spearman correlation coefficients were >or=0.89. The ARCHITECT CsA assay has significantly reduced CsA metabolite interference relative to other immunoassays and is a convenient and sensitive semiautomated method to measure CsA in whole blood.