Modulation of the pentose phosphate pathway alters phase I metabolism of testosterone and dextromethorphan in HepG2 cells

Abstract

Aim:The pentose phosphate pathway (PPP) is involved in the activity of glucose-6-phosphate dehydrogenase (G6PD) and generation of NADPH, which plays a key role in drug metabolism. The aim of this study was to investigate the effects of modulation of the PPP on drug metabolism capacity in vitro. Methods: A pair of hepatic cell lines, ie the cancerous HepG2 cells and normal L02 cells, was used. The expression of CYP450 enzymes, p53 and G6PD in the cells were analyzed. The metabolism of testosterone (TEST, 10 μmol/L) and dextromethorphan (DEM, 1 μmol/L), the two typical substrates for CYP3A4 and CYP2D6, in the cells was examined in the presence of different agents. Results: Both the expression and metabolic activities of CYP3A4 and CYP2D6 were considerably higher in HepG2 cells than in L02 cells. The metabolism of TEST and DEM in HepG2 cells was dose-dependently inhibited by the specific CYP3A4 inhibitor ketoconazole and CYP2D6 inhibitor quinidine. Addition of the p53 inhibitor cyclic PFT-α (5, 25 μmol/L) in HepG2 cells dose-dependently enhanced the metabolism of DEM and TEST, whereas addition of the p53 activator NSC 66811 (3, 10, 25 μmol/L) dose-dependently inhibited the metabolism. Furthermore, addition of the G6PD inhibitor 6-aminonicotinamide (5, 15 μmol/L) in HepG2 cells dose-dependently inhibited the metabolism of DEM and TEST, whereas addition of the PPP activity stimulator menadione (1, 5, 15 μmol/L) dosedependently enhanced the metabolism. Conclusion: Modulation of p53 and the PPP alters the metabolism of DEM and TEST, suggesting that the metabolic flux pattern of PPP may be closely involved in drug metabolism and the individual variance.

17β‐Estradiol inhibits testosterone‐induced cell proliferation in HepG2 by modulating the relative ratios of 3 estrogen receptor isoforms to the androgen receptor

“…The medium was refreshed 2 or 3 times a week and, up to approximately 80% to 90% confluence, cells were detached from the petri dish with a 0.25% trypsin‐ethylenediaminetetraacetic acid solution (Beyotime) for subculture at a 1:4 split ratio. As cell lines can be changed over time in culture, cells within 20 passages from the originally purchased vial from the American Type Culture Collection were used to control for genomic drift due to instability . To deplete endogenous steroids in the gene expression studies, the media were changed to phenol red‐free medium containing 10% charcoal/dextran‐FBS (Gemini Bio Products, Sacramento, CA) for 48 hours before experimentation …”

“…As cell lines can be changed over time in culture, cells within 20 passages from the originally purchased vial from the American Type Culture Collection were used to control for genomic drift due to instability. 14 To deplete endogenous steroids in the gene expression studies, the media were changed to phenol red-free medium containing 10% charcoal/dextran-FBS (Gemini Bio Products, Sacramento, CA) for 48 hours before experimentation. 15 For all experimental treatments, cells in the logarithmic growth phase were plated in 96-or 6-well plates with a density of 5 × 10 3 and 3 × 10 5 cell per well, respectively, according to different experimental purposes.…”

Niacin analogue, 6-Aminonicotinamide, a novel inhibitor of hepatitis B virus replication and HBsAg production

“…The inhibition of meiosis resumption by 6-AN following sperm penetration into zona-free oocytes was also associated with the PPP [26] . Furthermore, 6-AN directly modulates the PPP signal pathway to trigger metabolic disorders, especially the drug metabolism, in HepG2 and other cancer cells [27] , indicating that 6-AN generated metabolic disorders of liver cancer. However, there are no reports of 6-AN on HBV replication [28] .…”

Disrupted hepatic pentose phosphate pathway directly participates in and indirectly promotes CYP3A reduction: A new strategy for CYP3A‐mediated drug hepatotoxicity

“…A previous study found that spontaneous PPP impairment in tumours could stimulate chk2/p53/p65 expression (Wang et al, ). In addition, the modulation of p53 and PPP could alter the metabolism of CYP3A substrates in HepG2 cells and LO2 cells (Xiao et al, ). Here, we found that the chk2/p53/p65 pathway was stimulated in the liver along with the impaired PPP (Figure a–d).…”