This article was originally published in Proteomics 2010, 10, 1463–1473, DOI 10.1002/pmic.200900785

A comparative proteomic analysis of HepG2 cells

incubated by S(�) and R(1) enantiomers of

anti-coagulating drug warfarin

Jing Bai, Laleh Sadrolodabaee, Chi Bun Ching, Balram Chowbay and Wei Ning Chen

Keywords:

Cell biology / ERp 57 / iTRAQ-coupled LC-MS/MS proteomics / Protein DJ-1 / 14-3-3 Sigma / Warfarin

enantiomers

Warfarin is a commonly prescribed oral anti-coagulant with

narrow therapeutic index. It interferes with vitamin K cycle to

achieve anti-coagulating effects. Warfarin has two enantio-

mers, S(�) and R(1) and undergoes stereoselective metabo-

lism, with the S(�) enantiomer being more effective. We

reported that the intracellular protein profile in HepG2 cells

incubated with S(�) and R(1) warfarin, using iTRAQ-

coupled 2-D LC-MS/MS. In samples incubated with S(�) and

R(1) warfarin alone, the multi-task protein Protein SET

showed significant elevation in cells incubated with S(�)

warfarin but not in those incubated with R(1) warfarin. In

cells incubated with individual enantiomers of warfarin in the

presence of vitamin K, protein disulfide isomerase A3 which

is known as a glucose-regulated protein, in cells incubated

with S(�) warfarin was found to be down-regulated compared

to those incubated with R(1) warfarin. In addition, Protein

DJ-1 and 14-3-3 Proteins were down-regulated in cells incu-

bated with either S(�) or R(1) warfarin regardless of the

presence of vitamin K. Our results indicated that Protein DJ-1

may act as an enzyme for expression of essential enzymes in

vitamin K cycle. Taken together, our findings provided

molecular evidence on a comprehensive protein profile on

warfarin–cell interaction, which may shed new lights on

future improvement of warfarin therapy.ROS level of HepG2 cells incubated with drugs.

Proteomics Clin. Appl. 2010, 4, 745–771 763