Levels of SCS7/FA2H-Mediated Fatty Acid 2-Hydroxylation Determine the Sensitivity of Cells to Antitumor PM02734

Abstract

PM02734 is a novel synthetic antitumor drug that is currently in phase I clinical trials. To gain some insight into its mode of action, we used the yeast Saccharomyces cerevisiae as a model system. Treatment of S. cerevisiae with PM02734 rapidly induced necrosis-like cell death, as also found for mammalian cells treated with its close analogue kahalalide F. We have screened the complete set of 4,848 viable S. cerevisiae haploid deletion mutants to identify genes involved in sensitivity or resistance to PM02734. Forty-five percent of the 40 most sensitive strains identified had a role in intracellular vesicle trafficking, indicating that the drug severely affects this process. A mutant strain lacking the sphingolipid fatty acyl 2-hydroxylase Scs7 was found to be the most resistant to PM02734, whereas overexpression of Scs7 rendered the cells hypersensitive to PM02734. To validate these findings in human cells, we did small interfering RNA experiments and also overexpressed the Scs7 human homologue FA2H in human cancer cell lines. As in yeast, FA2H silencing turned the cells resistant to the drug, whereas FA2H overexpression led to an increased sensitivity. Moreover, exogenous addition of the 2-hydroxylated fatty acid 2-hydroxy palmitic acid to different human cell lines increased their sensitivity to the cytotoxic compound. Taken together, these results suggest that the cell membrane and, in particular, 2-hydroxy fatty acidcontaining ceramides are important for PM02734 activity. These findings may have important implications in the development of PM02734 because tumor cells with high FA2H expression are expected to be particularly sensitive to this drug. [Cancer Res 2008;68(23):9779-87]

Fatty Acid 2-Hydroxylase Mediates Diffusional Mobility of Raft-associated Lipids, GLUT4 Level, and Lipogenesis in 3T3-L1 Adipocytes

“…FA2H catalyzes the 2-hydroxylation of straight chain FAs and is responsible for the formation of 2-hydroxylated sphingolipids (14,40). Saccharomyces cerevisiae yeast strains with defective SCS7/FA2H are resistant to the antifungal syringomycin E (30) and the antitumor compound PM02734 (46), suggesting that 2-hydroxylated sphingolipids could influence the physical properties of some plasma membrane microdomains/lipid rafts and their internalization. This is supported by biophysical studies of membranes in reconstituted systems (47,48).…”

Biochemistry and genetics of inherited disorders of peroxisomal fatty acid metabolism

“…Given extracellularly, the majority of 2OH-FA appear to be degraded, with few molecules being incorporated into structural lipids ( 19,59,60 ). The steps involved in the degradation of 2OH-FA are an activation, uptake of the…”

Stereospecificity of fatty acid 2-hydroxylase and differential functions of 2-hydroxy fatty acid enantiomers

“…However, there is lack of information on which enantiomer is generated by FA2H and on whether both enantiomers are biologically active. Moreover, due to the lack of stereochemistry information on FA2H, racemic 2-OH FAs, but not 2-OH FA enantiomers were used to explore its potential role or to reverse effects of FA2H deficiency ( 7,13 ). In the current study, we examined the stereospecifi city of FA 2-hydroxylation catalyzed by FA2H.…”