Repozytorium publikacji - Politechnika Gdańska

We examine whether CYP3A4 overexpression influences the rate and pattern of antitumor imidazoacridinone C-1311 metabolism, in relation to the impact of this overexpression on cell cycle progression and final cellular response of CHO cells following C-1311 treatment. Methods: Three CHO cell lines: CHO-WT, wild type, CHO-HR, overexpressing cytochrome P450 reductase (CPR) and CHO-HR-3A4, coexpressing CPR and CYP3A4 were applied. Metabolic transformation of C-1311 and measurement of CYP3A4 activity were performed using RP-HPLC. CYP3A4 protein level was examined using Western blot assay. Flow cytometry for phosphatydilserine externalization, caspase-3 activity and sub-G1 DNA fraction was applied to identify apoptosis. DAPI staining was performed to analyze the cellular morphology. Accelerated senescence was examined on the basis of analysis of pH 6.0-dependent β-galactosidase (SA-β-gal) expression. Results: CYP3A4 supported by CPR did not participate in C-1311 metabolism in CHO cells. However, C-1311 effectively inhibited CYP3A4 enzymatic activity, while having no impact on CYP3A4 protein level. CHO wild type cells underwent rather stable G2/M arrest following C-1311 treatment, while transfected cells only transiently accumulated in this compartment. C-1311-treated cells died by apoptosis and necrosis, induced faster and to a greater extent in CHO transfected cells than in wild type. Additionally, the surviving CYP3A4 overexpressing cells underwent accelerated senescence.

Autorzy

• Monika Pawłowska,
• dr hab. Ewa Augustin link otwiera się w nowej karcie ,
• prof. dr hab. inż. Zofia Mazerska link otwiera się w nowej karcie