- Email: [email protected]
Antiproliferative effect of ergot alkaloids in a human breast cancer cell line
164s
D-067
ANTIPROLIFERATIVEEFFECT OF ERGOT ALKALOIDS IN A HUMAN BREAST CANCER CELL LINE Giubertoni M., Calvo M., Sica G.*, Di Carlo F. Istituto di Parmacologia e Terapia Sperimentale, Universita di Torino *Istituto di Istologia, Universita Cattolica de1 Sacro Cuore, Roma, Italia. A number of experimental data suggest that prolactin plays a prominent role as a stimulating factor not only for normal mammary gland, but also for breast cancer growth. Accordingly bromocriptine has recently been included in experimental programs for the therapy of this kind of tumors. In this study we have investigated the effect of bromocriptine and other ergot alkaloids (ergotamine, dihydrocriptine) on the CG5 cell line proliferation. In parallel the same effect of estradiol and tamoxifen has been investigated. The changes in estradiol (ER) and progesterone-receptor(PgR) levels caused by estradiol, tamoxifen and bromocriptine have also been studied. After 6 days of exposure of the cells to the drugs we have seen on the cell proliferation that: 1) low estradiol concentrations (10-g-10-7M) increase and high concentrations (1O-5-1O-4M) reduce; 2) tamoxifen and bromocriptine (lo-9-lo-5M) do have an antiproliferative action; 3) ergotamine and dihydrocriptine have no effect. As regards the ER and PgR changes: 1) estradiol and tamoxifen (lo-*M) reduce ER (70% and 15% respectively) and increase PgR (80% and 40%); 2) bromocriptine increases ER (75%). but has no effect on PgR level, We conclude that bromocriptine (differently from dihydrocriptine and ergotamine) is active as an antiproliferativedrug by acting directly on tumor cells. Its mechanism of action is not clear but seems to be different from that of estradiol or tamoxifen. D-068
EFFECT OF ESTRADIOL (E2) ON CELL CYCLE DISTRIBUTION OF MCF-7 HUMAN BREAST CANCER CELLS: ANALYSIS BY FLOW CYTOMETRY (PI) AND IMMUNOCYTOCHEMISTRY(Ki-67) Bontenbal M., Peters H.A., Sonneveld P.*, Foekens J.A. and Klijn J.G.M. The Dr. Daniel den Hoed Cancer Center and * Erasmus University Rotterdam, The Netherlands. When MCF-7 human breast cancer cells were grown in medium supplemented with DCC-treated male human serum, administration of 30 pM E2 enhanced after 20-24 h the percentage of SGZMphase cells from lo-15 to 50-60X measured by flow cytometry. We have subsequently cultured MCF-7 cells in the presence of DCC-treated FCS under "completely estrogen-deprived"conditions, i.e. in medium without phenol red. The effect of E2 (1 nM) administration for 21 h and medium refreshment was studied both with flow cytometry (PI) and with the monoclonal antibody Ki-67 as a marker for proliferating cells. In the absence of E2, daily medium refreshment resulted in decreased growth of the MCF-7 cell cultures (19% and 26% decrease in cell number 72 h and 96 h after seeding resp., p < 0.05), compared to cultures not receiving fresh medium. This decrease in growth was also apparent from the amount of cycling cells (SG2MGl) based on the Ki-67 assay. With daily medium refreshment, 60% of the cells were in the SGLMGl phase at 72 h and 96 h, whereas without medium refreshment resp. 84% and 76% of the cells were in cycle. At 96 h both groups were treated with a 21 h E2 pulse. In the group where medium was not refreshed E2 did not increase the % of cycling cells (76% Ki-67 pas., 63% SGZM-phase by flow cytometry) in cultures not having received fresh medium. On the other hand after daily medium refreshment E2 stimulation resulted in an increased % of cycling cells. The Ki-67 pos. fraction rose from 60 to 79% (p < 0.01) and the SGZM cells from 33 to 53% (p < 0.05). In conclusion: these data suggest that MCF-7 cells secrete factor(s) into the medium which can replace E2 for growth stimulation. (Supp. by Neth. Cancer Found.) D-069 MODULATION OF PLASMINCGEN ACTIVATOR LEVELS BY MATRIX COMPONENTS IN BREAST CANCER CELL LINES. BOUTIERE B., POURREAU-SCHNEIDERIi.,ARNOUX D., SAMPOL J. and MARTIN P.M. C.N.R.S. U.A. 1175, Faculti?de EledecineNord, 13015 Marseille, France Plasminogen activators (PA) are thought to be involved in the invasive growth of tumors. Both tissue-type (t-PA) and urokinase-like (u-PA) PAS have been characterized in breast adenocarcinomas: t-PA secretion has been implicated in the maintenance of the fluidity of the extracellular matrix; u-PA remains largely cell bound and its association with the cell membrane may facilitate cell migration. Plasninogen binds to fibronectin and to laminin as does t-PA, u-PA to a lesser extent. In addition, hormones (ca. estradiol, insulin) stiolulate PA production in target cells. In an attempt to precise the interrelation between tunor cells, basement membrane components, hormones and PA activities, we assayed u-PA and t-PA contents in cellular extracts and conditioned media from MCF-7 and MDA-MD-231 cells grown in the presence of purified matrix components. T-PA was measured using a solid phase fibrin assay, U-PA using a liquid phase assay, both being quantitated using a chromogenic substrate. We found high u-PA activity in the MDA-MB-231 cell line mostly recovered in cellular extracts, and moderate t-PA activity predominately secreted into the culture medium. Enhancement of u-PA occurred when the plastic growth surface was precoated with laminin or fibronectin. With estrogen-sensitive MCF-7 cells, the association of estradiol and laminin enhanced u-PA. These results support the association of tumor cells, matrix components and horuones in the production of PA. Further studies have been undertaken to evaluate the role of membrane binding of matrix proteins, hormones, plasminogen and its activators, as markers of the invasive capacity of tumor cells and the expression of the malignant phenotype. This research received financial support from INSERM and ARC.
D-067
ANTIPROLIFERATIVEEFFECT OF ERGOT ALKALOIDS IN A HUMAN BREAST CANCER CELL LINE Giubertoni M., Calvo M., Sica G.*, Di Carlo F. Istituto di Parmacologia e Terapia Sperimentale, Universita di Torino *Istituto di Istologia, Universita Cattolica de1 Sacro Cuore, Roma, Italia. A number of experimental data suggest that prolactin plays a prominent role as a stimulating factor not only for normal mammary gland, but also for breast cancer growth. Accordingly bromocriptine has recently been included in experimental programs for the therapy of this kind of tumors. In this study we have investigated the effect of bromocriptine and other ergot alkaloids (ergotamine, dihydrocriptine) on the CG5 cell line proliferation. In parallel the same effect of estradiol and tamoxifen has been investigated. The changes in estradiol (ER) and progesterone-receptor(PgR) levels caused by estradiol, tamoxifen and bromocriptine have also been studied. After 6 days of exposure of the cells to the drugs we have seen on the cell proliferation that: 1) low estradiol concentrations (10-g-10-7M) increase and high concentrations (1O-5-1O-4M) reduce; 2) tamoxifen and bromocriptine (lo-9-lo-5M) do have an antiproliferative action; 3) ergotamine and dihydrocriptine have no effect. As regards the ER and PgR changes: 1) estradiol and tamoxifen (lo-*M) reduce ER (70% and 15% respectively) and increase PgR (80% and 40%); 2) bromocriptine increases ER (75%). but has no effect on PgR level, We conclude that bromocriptine (differently from dihydrocriptine and ergotamine) is active as an antiproliferativedrug by acting directly on tumor cells. Its mechanism of action is not clear but seems to be different from that of estradiol or tamoxifen. D-068
EFFECT OF ESTRADIOL (E2) ON CELL CYCLE DISTRIBUTION OF MCF-7 HUMAN BREAST CANCER CELLS: ANALYSIS BY FLOW CYTOMETRY (PI) AND IMMUNOCYTOCHEMISTRY(Ki-67) Bontenbal M., Peters H.A., Sonneveld P.*, Foekens J.A. and Klijn J.G.M. The Dr. Daniel den Hoed Cancer Center and * Erasmus University Rotterdam, The Netherlands. When MCF-7 human breast cancer cells were grown in medium supplemented with DCC-treated male human serum, administration of 30 pM E2 enhanced after 20-24 h the percentage of SGZMphase cells from lo-15 to 50-60X measured by flow cytometry. We have subsequently cultured MCF-7 cells in the presence of DCC-treated FCS under "completely estrogen-deprived"conditions, i.e. in medium without phenol red. The effect of E2 (1 nM) administration for 21 h and medium refreshment was studied both with flow cytometry (PI) and with the monoclonal antibody Ki-67 as a marker for proliferating cells. In the absence of E2, daily medium refreshment resulted in decreased growth of the MCF-7 cell cultures (19% and 26% decrease in cell number 72 h and 96 h after seeding resp., p < 0.05), compared to cultures not receiving fresh medium. This decrease in growth was also apparent from the amount of cycling cells (SG2MGl) based on the Ki-67 assay. With daily medium refreshment, 60% of the cells were in the SGLMGl phase at 72 h and 96 h, whereas without medium refreshment resp. 84% and 76% of the cells were in cycle. At 96 h both groups were treated with a 21 h E2 pulse. In the group where medium was not refreshed E2 did not increase the % of cycling cells (76% Ki-67 pas., 63% SGZM-phase by flow cytometry) in cultures not having received fresh medium. On the other hand after daily medium refreshment E2 stimulation resulted in an increased % of cycling cells. The Ki-67 pos. fraction rose from 60 to 79% (p < 0.01) and the SGZM cells from 33 to 53% (p < 0.05). In conclusion: these data suggest that MCF-7 cells secrete factor(s) into the medium which can replace E2 for growth stimulation. (Supp. by Neth. Cancer Found.) D-069 MODULATION OF PLASMINCGEN ACTIVATOR LEVELS BY MATRIX COMPONENTS IN BREAST CANCER CELL LINES. BOUTIERE B., POURREAU-SCHNEIDERIi.,ARNOUX D., SAMPOL J. and MARTIN P.M. C.N.R.S. U.A. 1175, Faculti?de EledecineNord, 13015 Marseille, France Plasminogen activators (PA) are thought to be involved in the invasive growth of tumors. Both tissue-type (t-PA) and urokinase-like (u-PA) PAS have been characterized in breast adenocarcinomas: t-PA secretion has been implicated in the maintenance of the fluidity of the extracellular matrix; u-PA remains largely cell bound and its association with the cell membrane may facilitate cell migration. Plasninogen binds to fibronectin and to laminin as does t-PA, u-PA to a lesser extent. In addition, hormones (ca. estradiol, insulin) stiolulate PA production in target cells. In an attempt to precise the interrelation between tunor cells, basement membrane components, hormones and PA activities, we assayed u-PA and t-PA contents in cellular extracts and conditioned media from MCF-7 and MDA-MD-231 cells grown in the presence of purified matrix components. T-PA was measured using a solid phase fibrin assay, U-PA using a liquid phase assay, both being quantitated using a chromogenic substrate. We found high u-PA activity in the MDA-MB-231 cell line mostly recovered in cellular extracts, and moderate t-PA activity predominately secreted into the culture medium. Enhancement of u-PA occurred when the plastic growth surface was precoated with laminin or fibronectin. With estrogen-sensitive MCF-7 cells, the association of estradiol and laminin enhanced u-PA. These results support the association of tumor cells, matrix components and horuones in the production of PA. Further studies have been undertaken to evaluate the role of membrane binding of matrix proteins, hormones, plasminogen and its activators, as markers of the invasive capacity of tumor cells and the expression of the malignant phenotype. This research received financial support from INSERM and ARC.