Framework for the quality assurance of 'omics technologies considering GLP requirements

‘Omics technologies are gaining importance to support regulatory toxicity studies. Prerequisites for performing ‘omics studies considering GLP principles were discussed at the European Centre for Ecotoxicology and Toxicology of Chemicals (ECETOC) Workshop Applying ‘omics technologies in Chemical Risk Assessment. A GLP environment comprises a standard operating procedure system, proper pre-planning and documentation, and inspections of independent quality assurance staff. To prevent uncontrolled data changes, the raw data obtained in the respective ‘omics data recording systems have to be specifically defined. Further requirements include transparent and reproducible data processing steps, and safe data storage and archiving procedures. The software for data recording and processing should be validated, and data changes should be traceable or disabled. GLP-compliant quality assurance of ‘omics technologies appears feasible for many GLP requirements. However, challenges include (i) defining, storing, and archiving the raw data; (ii) transparent descriptions of data processing steps; (iii) software validation; and (iv) ensuring complete reproducibility of final results with respect to raw data. Nevertheless, ‘omics studies can be supported by quality measures (e.g., GLP principles) to ensure quality control, reproducibility and traceability of experiments. This enables regulators to use ‘omics data in a fit-for-purpose context, which enhances their applicability for risk assessment

Preparation and Evaluation of Poly(Ethylene Glycol)–Poly(Lactide) Micelles as Nanocarriers for Oral Delivery of Cyclosporine A

A series of monomethoxy poly(ethylene glycol)–poly(lactide) (mPEG–PLA) diblock copolymers were designed according to polymer–drug compatibility and synthesized, and mPEG–PLA micelle was fabricated and used as a nanocarrier for solubilization and oral delivery of Cyclosporine A (CyA). CyA was efficiently encapsulated into the micelles with nanoscaled diameter ranged from 60 to 96 nm with a narrow size distribution. The favorable stabilities of CyA-loaded polymeric micelles were observed in simulated gastric and intestinal fluids. The in vitro drug release investigation demonstrated that drug release was retarded by polymeric micelles. The enhanced intestinal absorption of CyA-loaded polymeric micelles, which was comparable to the commercial formulation of CyA (Sandimmun Neoral®), was found. These suggested that polymeric micelles might be an effective nanocarrier for solubilization of poorly soluble CyA and further improving oral absorption of the drug

Screening of Tanzanian plant extracts for their potential inhibitory effect on P-glycoprotein mediated efflux

For years, many efforts have been made to discover new drugs using plants as natural screening libraries. In this study, extracts of 43 Tanzanian medicinal plants were screened for their potential inhibitory effect on P-gp, using the secretory transport of Cyclosporin A (CsA) in the Caco-2 system as a measure of the functionality of P-gp efflux. Two out of these 43 plant extracts (extracts of Annickia kummeriae and Acacia nilotica) appeared to have a modulatory effect on P-gp related efflux carriers. In presence of the extract of Annickia kummeriae, a concentration dependent decrease on the polarity in transport of CsA was observed; the inhibitory effect of this extract on P-gp was comparable to that of valspodar, a known P-gp inhibiting agent. The exact nature of the active components of these botanicals remains to be identified.status: publishe

Simulated intestinal fluid as transport medium in the Caco-2 cell culture model

The Caco-2 model is widely used as a predictive tool for the oral absorption of drug candidates. Presently. transport experiments in the Caco-2 system are usually performed in 'HBSS-like' buffers. In this paper, we investigate the possibility of using simulated intestinal buffers as donor solvent during Caco-2 experiments. Toxicity assessment of these buffers on the monolayer showed that FASSIF was compatible with the Caco-2 model for at least 2 h. On the other hand, FESSIF was toxic to the monolayer. The functionality of the Caco-2 cells was assessed by determination of the transport of model compounds and the metabolic activity of hydrolases in presence of these buffers. Similar P-app values for the (passive) theophyllin transport as well as for the (active) phenylalanine transport were obtained in TM and FASSIF It was demonstrated that NaTC (present in FASSIF) had a P-gp inhibitory activity, as inclusion of NaTC in the apical compartment resulted in an increased absorptive and decreased secretory transport of CsA. The activity of the aminopeptidase enzyme was similar in both models. These results suggest that FASSIF can be used as an apical medium in the Caco-2; system. Since bile salts are also present in physiological conditions, the use of FASSIF may increase the relevance for the prediction of oral absorption using Caco-2 experiments. (C) 2002 Elsevier Science B.V. All rights reserved

Simulated intestinal fluid as transport medium in the Caco-2 cell culture model

The Caco-2 model is widely used as a predictive tool for the oral absorption of drug candidates. Presently, transport experiments in the Caco-2 system are usually performed in 'HBSS-like' buffers. In this paper, we investigate the possibility of using simulated intestinal buffers as donor solvent during Caco-2 experiments. Toxicity assessment of these buffers on the monolayer showed that FASSIF was compatible with the Caco-2 model for at least 2 h. On the other hand, FESSIF was toxic to the monolayer. The functionality of the Caco-2 cells was assessed by determination of the transport of model compounds and the metabolic activity of hydrolases in presence of these buffers. Similar P(app) values for the (passive) theophyllin transport as well as for the (active) phenylalanine transport were obtained in TM and FASSIF. It was demonstrated that NaTC (present in FASSIF) had a P-gp inhibitory activity, as inclusion of NaTC in the apical compartment resulted in an increased absorptive and decreased secretory transport of CsA. The activity of the aminopeptidase enzyme was similar in both models. These results suggest that FASSIF can be used as an apical medium in the Caco-2 system. Since bile salts are also present in physiological conditions, the use of FASSIF may increase the relevance for the prediction of oral absorption using Caco-2 experiments.status: publishe

Intestinal absorption enhancement of the ester prodrug tenofovir disoproxil fumarate through modulation of the biochemical barrier by defined ester mixtures

The effect of discrete esters and ester mixtures on the intestinal stability and absorption of tenofovir disoproxil fumarate (tenofovir DF, an esterase-sensitive prodrug of the antiviral tenofovir) was compared with the effect of strawberry extract, which has been shown to enhance the absorption of the prodrug across Caco-2 monolayers and in rat ileum. In addition, the mechanism of absorption enhancement was investigated. In rat intestinal homogenates, complete inhibition of the conversion of tenofovir DF (as obtained by strawberry extract) could only be obtained at relatively high concentrations of the discrete esters or by using mixtures of esters (e.g., propyl p-hydroxybenzoate 0.02%, octyl acetate 0.02%, ethyl caprylate 0.01%). Coincubation of tenofovir DF with this mixture also resulted in an enhancement of its absorption in the in vitro Caco-2 system as well as in rat ileum. As tenofovir DF is a substrate for P-glycoprotein (P-gp)-related efflux carriers in the Caco-2 model, the modulatory effect of the ester mixtures was studied on the functionality of P-gp using cyclosporin A (CsA) as a model substrate. Strawberry extract as well as the mixture of three esters interfered with the absorptive transport of CsA across Caco-2 monolayers, illustrating that both mixtures interfere with both esterase-activity and P-gp functionality. This concerted barrier was not observed in rat ileum, suggesting differential functional activities of the biochemical barrier toward tenofovir DF in different absorption systems. Overall, our results illustrate that modulation of the biochemical barrier (metabolism and efflux) of tenofovir DF by ester mixtures can be used to increase the intestinal absorption of tenofovir DF in an in vitro and an in situ absorption model; the mechanism of action appears to be a complex interplay of different systems; the differential expression of carriers and enzymes in different systems illustrates the difficulty of extrapolating observations between different systems/species.status: publishe