A Novel Mode of Action of the Putative Sphingosine Kinase Inhibitor 2-(p-hydroxyanilino)-4-(p-chlorophenyl) Thiazole (SKI II): Induction of Lysosomal Sphingosine Kinase 1 Degradation

作者：Ren Shuyu; Xin Cuiyan; Pfeilschifter Josef; Huwiler Andrea^*

来源：Cellular Physiology and Biochemistry, 2010, 26(1): 97-104.

DOI：10.1159/000315110

• SKI II
• Sphingosine kinase 1
• Protein degradation
• Lysosome
• Chloroquine
• Cathepsin B

Background: Sphingosine kinase 1 (SK1) is a key enzyme in the generation of sphingosine 1-phosphate (S1P) which critically regulates a variety of important cell responses such as proliferation and migration. Therefore, inhibition of SK-1 has been suggested to be an attractive approach to treat tumor growth and metastasis formation. Results: We show here that the previously developed putative SK-1 inhibitor 2-(phydroxyanilino)4-(p-chlorophenyl) thiazole (SKI II) displays an additional facet of action complementary to the known inhibition of enzymatic SK-1 activity. In various human cell lines including glomerular podocytes and mesangial cells, the human endothelial cell line EA. hy 926, and the lung cancer cell line NCI H358, SKI II reduced TGF beta- and TPA-stimulated cellular SK1 activity by downregulating SK-1 protein expression without affecting SK-1 mRNA expression. By using cycloheximide to block the de novo protein synthesis, the protein expression of SK-1 under untreated conditions was stable over 24h. Under SKI II treatment, the half-live drastically decreased to approximately 0.8h. Mechanistically, this degradation occurred through a lysosomal pathway and involved cathepsin B since the general lysosomal inhibitor chloroquine and the specific cathepsin B inhibitor CA-074ME were able to reverse the effect of SKI II. Surprisingly, in vitro SK-1 activity assays revealed only a very weak direct inhibitory effect of SKI II on SK-1 overexpressed HEK293 cell lysates. Conclusion: These data show for the first time that the previously developed SK inhibitor SKI II hardly inhibits SK-1 directly but rather acts by triggering the lysosomal degradation of SK-1 in various cell types. This finding discloses a new mode of action of SKI II and strongly suggests that additional direct targets of SKI II may exist other than SK-1.

• 出版日期2010