Novel off-target effect of tamoxifen--inhibition of acid ceramidase activity in cancer cells.
Acid ceramidase (AC), EC 3.5.1.23, a lysosomal enzyme, catalyzes the hydrolysis of ceramide to constituent sphingoid base, sphingosine, and fatty acid. Because AC regulates the levels of pro-apoptotic ceramide and mitogenic sphingosine-1-phosphate, it is considered an apt target in cancer therapy. The present study reveals, for the first time, that the prominent antiestrogen, tamoxifen, is a pan-effective AC inhibitor in the low, single digit micromolar range, as demonstrated in a wide spectrum of cancer cell types, prostate, pancreatic, colorectal, and breast. Prostate cancer cells were chosen for the detailed investigations. Treatment of intact PC-3 cells with tamoxifen produced time- and dose-dependent inhibition of AC activity. Tamoxifen did not impact cell viability nor did it inhibit AC activity in cell-free assays. In pursuit of mechanism of action, we demonstrate that tamoxifen induced time-, as early as 5min, and dose-dependent, as low as 5μM, increases in lysosomal membrane permeability (LMP), and time- and dose-dependent downregulation of AC protein expression. Assessing various protease inhibitors revealed that a cathepsin B inhibitor blocked tamoxifen-elicited downregulation of AC protein; however, this action failed to restore AC activity unless assayed in a cell-free system at pH4.5. In addition, pretreatment with tamoxifen inhibited PC-3 cell migration. Toremifene, an antiestrogen structurally similar to tamoxifen, was also a potent inhibitor of AC activity. This study reveals a new, off-target action of tamoxifen that may be of benefit to enhance anticancer therapies that either incorporate ceramide or target ceramide metabolism.
John Wayne Cancer Institute at Saint John's Health Center, Department of Experimental Therapeutics, Santa Monica, CA 90404, USA.
Samy A F
SA
Morad
Jonathan C
JC
Levin
Su-Fern
SF
Tan
Todd E
TE
Fox
David J
DJ
Feith
Myles C
MC
Cabot
D055573:Acid Ceramidase / Q000037:antagonists & inhibitors* / Q000235:genetics / Q000378:metabolism; D000970:Antineoplastic Agents / Q000494:pharmacology*; D017209:Apoptosis / Q000187:drug effects; D002401:Cathepsin B / Q000037:antagonists & inhibitors / Q000378:metabolism; D045744:Cell Line, Tumor; D002470:Cell Survival / Q000187:drug effects; D002474:Cell-Free System; D002518:Ceramides / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D005260:Female; D015870:Gene Expression; D006801:Humans; D006863:Hydrogen-Ion Concentration; D008246:Lysophospholipids / Q000378:metabolism; D008247:Lysosomes / Q000187:drug effects* / Q000201:enzymology; D008297:Male; D020845:Selective Estrogen Receptor Modulators / Q000494:pharmacology*; D013110:Sphingosine / Q000031:analogs & derivatives / Q000378:metabolism; D013629:Tamoxifen / Q000494:pharmacology*; D017312:Toremifene / Q000494:pharmacology
D000970:Antineoplastic Agents; D002518:Ceramides; D004791:Enzyme Inhibitors; D008246:Lysophospholipids; D020845:Selective Estrogen Receptor Modulators; D013629:Tamoxifen; C060506:sphingosine 1-phosphate; D017312:Toremifene; D002401:Cathepsin B; D055573:Acid Ceramidase; D013110:Sphingosine
AC; AO; Acid ceramidase; Antiestrogen; Ceramide; GC; LMP; Lysosomal protease; N-oleoylethanolamine; NOE; PBS; PBST; PMSF; S1-P; SK; Tamoxifen; acid ceramidase; acridine orange; glucosylceramide; lysosomal membrane permeability; phenylmethylsulfonyl fluoride; phosphate-buffered saline; phosphate-buffered saline with Tween-20; sphingosine 1-phosphate; sphingosine kinase