BMC Cancer 2004, 4:49 http://www.biomedcentral.com/1471-2407/4/49 cFdTouimgnriutenrr-gocelion(5ucOrus)beatoiof6n-wkeitho-1P5GµFM1α ptarmodouxcifteionn(γb)yarnadt MlivEeMrc/BeSllAs Time-course of 6-keto-PGF1α production by rat liver cells during incubation with 15 µM tamoxifen (γ) and MEM/BSA control (O). Analyses were performed with duplicate and triplicate dishes; mean values are shown. retinoic acid and the tyrosine analog of thiazolidinedione, GW7845 [19]. The rapidity of stimulation (5 min) and the lack of effect of actinomycin D or ICI-182,780 suggest that the stimulation of PGI2 production by tamoxifen is non-genomic and does not require ER occupancy. LY117018 blocks PGI2 production stimulated by tamoxifen (Fig. 7). This blockade is unaffected by preincu- bation of the cells for 2-h with either actinomycin D or ICI-182,780 (data not shown). LY117018, however, does not inhibit the release of AA stimulated by tamoxifen; their combined effect is at least additive (Fig. 8). Discussion These studies demonstrate that, at µM concentrations, tamoxifen and the raloxifene analog, LY117,018 stimulate the release of AA from cells in culture. At nM levels, the release is not observed. The effectiveness of these two compounds at prevention of estrogen-dependent breast cancer reflects competition for the ER [1,2]. In addition to occupancy of the ER, I am postulating that these drugs, at µM concentrations, may also prevent breast cancer of estrogen independent tumors. Consistent with this hypothesis are the findings that even at 100 µM concen- trations, ethanol extracts of Femora ® (letrozole) or Arami- dex ® (anastrozole) do not release AA from cells in culture (unpublished data). These two drugs inhibit estrogen syn- thesis by blocking aromatase enzymes and also prevent estrogen-dependent breast cancer [20]. In view of the stimulations of AA release by tamoxifen and LY117018 from human colon carcinoma cells (Fig. 2-A), the occur- rence of colon cancer in women undergoing long term taEpFinlfoiufgdensuc(sTtrBtePoi)mAfe6u()fAfle6a)ct-ektadecotfboinIy-CoPtmIGa-m1yF8coi2nx,p7iDfr8eo0nodonuncbtbaiosanaslaslot6rim-iknuedltauotce-ePddGb(Fylatcatpmarcoyxdstiufiencn- 1α 1α Effect of (A) actinomycin D on basal or induced (lactacystin plus TPA) 6-keto-PGF1α production stimulated by tamoxifen and (B) effect of ICI-182,780 on basal 6-keto-PGF1α produc- tion stimulated by tamoxifen. In (A), rat liver cells were pre- incubated with 1 µM actinomycin for 2-h and then incubated with tamoxifen (16 µM) or lactacystin plus TPA for 6-h. In (B), rat liver cells were incubated with tamoxifen (16 µM) and tamoxifen in the presence and absence of ICI-182,780 (16 µM) for 6-h. The analyses were performed with triplicate dishes. Each bar gives the mean value and the brackets give the SEM. Parts of the experiments were repeated several times with the same results. * Statistically significant vs con- trol. ** Statistically significant vs lactacystin plus TPA. treatment with high levels of tamoxifen would be of interest. A mechanism that most simply explains the release of AA by tamoxifen and LY117018 is the ability of such com- pounds to intercalate into cell membranes and affect phospholipase activities. The release of AA from endothe- lial cells by the Ca2+ ionophore A-23187 reflects phos- pholipase activities. It is regulated by phosphorylation of the enzyme [21]. The enzymic properties of the altered membrane may impact signaling mechanisms e.g., Page 5 of 8 (page number not for citation purposes)