Hepatic CYP3A4 Enzyme Compensatively Maintains Endogenous Geranylgeranoic Acid Levels in -Knockout Human Hepatoma Cells.

Geranylgeranoic acid (GGA), developed as a preventive agent against second primary hepatoma, has been reported to be biosynthesized via the mevalonate pathway in human hepatoma-derived cells. Recently, we found that monoamine oxidase B (MAOB) catalyzed the oxidation of geranylgeraniol (GGOH) to produce geranylgeranial (GGal), a direct precursor of endogenous GGA in hepatoma cells, using tranylcypromine, an inhibitor of MAOs, and knockdown by siRNA. However, endogenous GGA level was unexpectedly unchanged in -knockout (KO) cells established using the CRISPR-Cas9 system, suggesting that some other latent metabolic pathways maintain endogenous GGA levels in the -KO cells. Here, we investigated the putative latent enzymes that oxidize GGOH in Hep3B/-KO cells. First, the broad-specific cytochrome P450 enzyme inhibitors decreased the amount of endogenous GGA in Hep3B/-KO cells in a dose-dependent manner. Second, among the eight members of superfamily that have been suggested to be involved in the oxidation of isoprenols and/or retinol in previous studies, only the gene significantly upregulated its cellular mRNA level in Hep3B/-KO cells. Third, a commercially available recombinant human CYP3A4 enzyme was able to oxidize GGOH to GGal, and fourth, the knockdown of by siRNA significantly reduced the amount of endogenous GGA in Hep3B/-KO cells. These results indicate that CYP3A4 can act as an alternative oxidase for GGOH when hepatic MAOB is deleted in the human hepatoma-derived cell line Hep3B, and that endogenous GGA levels are maintained by a multitude of enzymes.