Establishment of a novel hepatocyte model that expresses four cytochrome P450 genes stably via mammalian-derived artificial chromosome for pharmacokinetics and toxicity studies.

The utility of HepG2 cells to assess drug metabolism and toxicity induced by chemical compounds is hampered by their low cytochrome P450 (CYP) activities. To overcome this limitation, we established HepG2 cell lines expressing major CYP enzymes involved in drug metabolism (CYP2C9, CYP2C19, CYP2D6, and CYP3A4) and CYP oxidoreductase (POR) using the mammalian-derived artificial chromosome vector. Transchromosomic HepG2 (TC-HepG2) cells expressing four CYPs and POR were used to determine time- and concentration-dependent inhibition and toxicity of several compounds by luminescence detection of CYP-specific substrates and cell viability assays.

Correlation between luminescence intensity and cytotoxicity in cell-based cytotoxicity assay using luciferase.

The luciferase reporter assay has become one of the conventional methods for cytotoxicity evaluation. Typically, the decrease of luminescence expressed by a constitutive promoter is used as an index of cytotoxicity. However, to our knowledge, there have been no reports of the correlation between cytotoxicity and luminescence intensity.

Intra-/inter-laboratory validation study on reactive oxygen species assay for chemical photosafety evaluation using two different solar simulators.

A previous multi-center validation study demonstrated high transferability and reliability of reactive oxygen species (ROS) assay for photosafety evaluation. The present validation study was undertaken to verify further the applicability of different solar simulators and assay performance. In 7 participating laboratories, 2 standards and 42 coded chemicals, including 23 phototoxins and 19 non-phototoxic drugs/chemicals, were assessed by the ROS assay using two different solar simulators (Atlas Suntest CPS series, 3 labs; and Seric SXL-2500V2, 4 labs).

Establishment and intra-/inter-laboratory validation of a standard protocol of reactive oxygen species assay for chemical photosafety evaluation.

A reactive oxygen species (ROS) assay was previously developed for photosafety evaluation of pharmaceuticals, and the present multi-center study aimed to establish and validate a standard protocol for ROS assay. In three participating laboratories, two standards and 42 coded chemicals, including 23 phototoxins and 19 nonphototoxic drugs/chemicals, were assessed by the ROS assay according to the standardized protocol. Most phototoxins tended to generate singlet oxygen and/or superoxide under UV-vis exposure, but nonphototoxic chemicals were less photoreactive.

Suitable top concentration for tests with mammalian cells: mouse lymphoma assay workgroup.

The Mouse Lymphoma Expert Workgroup of the International Workshop for Genotoxicity Tests (IWGT) met in Basel, Switzerland in August of 2009. The Workgroup (WG) was tasked with discussing the appropriate top concentration for non-pharmaceuticals that would be required for the conduct of the mouse lymphoma assay (MLA) when sufficient cytotoxicity [to between 10 and 20% relative total growth (RTG)] has not been attained. The WG approached this task by (1) enumerating the various regulatory decisions/use for MLA data, (2) discussing the appropriate assays to which MLA data and assay performance should be compared and (3) discussing all the proposals put forth concerning the top concentration for non-pharmaceuticals.