Non-synonymous single nucleotide alterations found in the CYP2C8 gene result in reduced in vitro paclitaxel metabolism.

Abstract

By sequencing genomic DNA from 73 established cell lines derived from Japanese individuals, we detected 9 single nucleotide polymorphisms (SNPs) in the CYP2C8 gene. Of them, 3 exonic SNPs resulted in amino acid alterations (g416a, R139K; a1196g, K399R; c1210g, P404A). The first two alterations were detected concurrently in one cell line and thought to be the same as CYP2C8*3. To examine the effects of these amino acid alterations on CYP2C8 function, wild-type and four types of variant CYP2C8 cDNA constructs (R139K, K399R, R139K/K399R and P404A) were transfected into Hep G2 cells and their paclitaxel 6a-hydroxylase activities were determined in vitro. Km values were not significantly different from that of the wild-type in any of the variants studied. The variant R139K/K399R showed reduced values for Vmax and clearance (Vmax/Km) similar to those of its single variant, R139K. The variant P404A also showed a significantly lowered clearance due to reduced level of protein expression. These results suggest that not only the double variant (R139K/K399R, CYP2C8*3) but also our novel variant P404A in the CYP2C8 gene are less efficient in paclitaxel metabolism.