The phospholipidosis-lnducing potential of the chemopotentiating drug, N,N-Diethyl-2-[4-(phenylmethyl)phenoxy]ethanamine (DPPE, tesmilifene) correlates with its stimulation of phosphatidylserine synthesis and exposure on the plasma membrane in MCF-7 breast cancer cells.

N,N-Diethyl-2-[4-(phenylmethyl)phenoxy]ethanamine (DPPE, tesmilifene), a potent chemopotentiating drug currently in Phase III clinical trials of metastatic breast cancer, increases cytotoxicity of anthracyclines and taxanes in a variety of multi-drug resistance expressing (MDR+) tumor cell lines in vitro; inhibits binding of histamine to CYP3A4, a lipid/prostanoid-regulatory P450; and modulates serum levels of HDL/LDL cholesterol and phospholipids in vivo. Since increased exposure of phosphatidylserine (PS) on the outer cell membrane leaflet is associated with apoptosis, increased clearance of dead cells by phagocytes and inhibition of the P-glycoprotein pump, the effect of DPPE on PS synthesis was assessed in vitro in a human breast cancer cell line. MCF-7 cells were incubated with 5 microM DPPE for 24 hr or 5 days, followed by addition of [1-(14)C]arachidonic acid for 4 hr; or [3H]serine for 8 hr. Compared to untreated cells, a 27-42% (p < 0.05) increase in [1-(14)C]arachidonic acid incorporated into all phospholipids, including a 1.9-fold increase (p < 0.05) in PS was observed in DPPE-treated cells. [3H]Serine incorporation into PS was elevated 37%, while the pool size of PS was elevated 23% (p < 0.05) in DPPE-treated cells, indicating elevated de novo PS biosynthesis. Annexin-5 binding studies indicated an elevation in exposure of PS on the surface of the plasma membrane in DPPE-treated cells. DPPE-treatment also resulted in N-(7-nitrobenz-2-oxa-1,3-diazol-4-yl)-1,2-dihexadecanoyl-sn-glycero-3-phosphoethanolamine accumulation indicative of phospholipidosis-inducing potential. Thus, the chemopotentiating effect of DPPE may be due to its phospholipidosis-inducing potential and stimulation of PS synthesis leading to an increased exposure of PS on the cell surface which could potentially enhance cancer cell clearance by phagocytes.

Displacement of histamine from liver cells and cell components by ligands for cytochromes P450.

Intracellular histamine (HA) and cytochrome P450 monooxygenases (P450) each have been proposed as mediators of cell function, growth, and proliferation. The P450 family of heme enzymes is found in virtually all cells and generates, transforms, or inactivates steroids and other lipids that participate in cell regulation. We previously demonstrated a second messenger role for HA in blood platelets and the formation of a HA-P450 heme complex when exogenous HA was added to microsomes isolated from rat liver cells or to purified human P450 isozymes. Employing a radioimmunoassay, we now demonstrate that rat liver slices, microsomes derived from the livers of adult male rats and mast cell-deficient mice, and hepatoma cells, all contain endogenous HA. HA release from microsomes into the incubation medium, as determined by radioimmunoassay, is enhanced in the presence of carbon monoxide, steroids, and certain drugs, all agents that unite either directly with the iron atom or bind elsewhere within the heme cavity. Rat liver slices preincubated with (3)H-HA release labeled amine into the medium in the presence of those same ligands. These findings provide evidence of an in situ HA-P450 complex and offer further support that the imidazole, HA, is a physiological, intracellular modulator of cytochromes P450 in liver cells, and perhaps of these and other heme proteins in tissues in general.