title: The anti-tumor alkylphospholipid perifosine is internalized by an ATP-dependent translocase activity across the plasma membrane of human KB carcinoma cells.
abstract: Perifosine is a promising anticancer alkylphospholipid (ALP) that induces apoptosis in tumor cells. Here we report evidences against a role of endocytosis in perifosine uptake by human KB carcinoma cells. We have generated a KB cell line resistant to perifosine (KB PER(R) clone10), which shows cross-resistance to the ALPs miltefosine and edelfosine, a marked impairment in the uptake of (14)C-perifosine at both 37 degrees C and 4 degrees C, and no signs for active efflux of the drug. KB PER(R) clone10 cells show a similar rate of raft-dependent endocytosis with respect to the parental cells, and silencing of both clathrin and dynamin in the latter causes only minor changes in the rate of perifosine uptake. Perifosine uptake is a temperature- and ATP-dependent, N-ethylmaleimide- and orthovanadate-sensitive process in parental cells. Accumulation of (14)C-perifosine and the fluorescent phospholipid analogue 6-[(7-nitrobenz-2-oxa-1,3-diazol-4-yl)aminocaproyl]-phosphatidylethanolamine (NBD-PE) is inhibited by perifosine in a concentration-dependent manner in parental cells. Moreover, NBD-PE accumulation is slower in PER(R) clone10 cells and correlated with phosphatidylserine exposure in their plasma membrane surface. Together, all these data suggest a role of plasma membrane translocation by a putative phospholipid translocase, rather than endocytosis, as the true mechanism for ALPs uptake in KB carcinoma cells.
authors.affiliation: Instituto de Parasitología y Biomedicina López-Neyra, Consejo Superior de Investigaciones Científicas, Parque Tecnológico de Ciencias de la Salud, Avda. del Conocimiento s/n, 18100 Armilla, Granada, Spain.
authors.forename: Francisco
authors.initials: F
authors.lastname: Muñoz-Martínez
authors.forename: Cristina
authors.initials: C
authors.lastname: Torres
authors.forename: Santiago
authors.initials: S
authors.lastname: Castanys
authors.forename: Francisco
authors.initials: F
authors.lastname: Gamarro
mesh_terms: D000255:Adenosine Triphosphate / Q000378:metabolism*; D000970:Antineoplastic Agents / Q000378:metabolism*; D015153:Blotting, Western; D045744:Cell Line, Tumor; D002462:Cell Membrane / Q000378:metabolism; D002966:Clathrin / Q000378:metabolism; D034284:Dynamin I / Q000378:metabolism; D004705:Endocytosis; D006801:Humans; D049294:Phospholipid Transfer Proteins / Q000378:metabolism*; D010767:Phosphorylcholine / Q000031:analogs & derivatives* / Q000378:metabolism; D034741:RNA, Small Interfering
chemical_list: D000970:Antineoplastic Agents; D002966:Clathrin; D049294:Phospholipid Transfer Proteins; D034741:RNA, Small Interfering; D010767:Phosphorylcholine; C105905:perifosine; D000255:Adenosine Triphosphate; D034284:Dynamin I