Krista A. Power

Mammalian lignans enterolactone and enterodiol, alone and in combination with the isoflavone genistein, do not promote the growth of MCF‐7 xenografts in ovariectomized athymic nude mice

This study determined the effect of the mammalian lignans enterolactone (ENL) and enterodiol (END) alone and in combination with the isoflavone genistein (GEN) on the growth of MCF‐7 tumors in ovariectomized nude mice. Ovariectomized athymic nude mice with established MCF‐7 tumors were fed a basal diet (AIN‐93G) and divided into 5 groups that received daily subcutaneous injections (10 mg/kg body weight (BW)) of ENL, END, GEN, a mixture of these compounds (MIX), or vehicle as a negative control for 22 weeks. A positive control group was implanted with an estradiol pellet in order to establish an estrogenic tumor growth response. In the ENL‐ and END‐treated mice, palpable tumors regressed significantly by 91 and 83%, respectively, resulting in final tumors that were similar to the negative control tumors. However, tumor cell apoptosis was significantly enhanced by the lignans. In the GEN‐treated mice, tumors initially regressed significantly by 64% but regression ceased following prolonged treatment, resulting in final tumors that were significantly larger compared to negative control, ENL‐, and END‐treated mice, in part by increasing tumor cell proliferation. The MIX treatment significantly regressed palpable tumors by 87% similar to negative control group, with no effects on tumor cell apoptosis or proliferation. The isoflavone GEN alone promoted the growth of established MCF‐7 human breast cancer xenografts after prolonged treatment while the mammalian lignans ENL and END did not. When these phytoestrogens were given in combination, no tumor growth‐promoting effects were observed.

Ligand-induced regulation of ERα and ERβ is indicative of human breast cancer cell proliferation

Two estrogen receptors (ER), ERα and ERβ, are expressed in breast cancer but their role in treatment response is unclear. The overall objective of this study was to determine if the presence of ERβ protein in breast cancer cell lines is an indicator of a poor prognosis based on cell proliferation. In addition, we determined the effect of estradiol (E2) and selective estrogen receptor modulators (SERMs), such as tamoxifen and genistein, on ERα and ERβ protein regulation, to help in the understanding of the mechanism behind their role in modulating cell proliferation. Using western blot and immunofluorescence analysis, the ER positive cell lines, MCF-7 and T47D, were found to contain both ERα and ERβ, and thus were used as model systems. E2 and genistein, which increased cell proliferation in both cell lines, induced an up regulation of ERβ in both cell lines. This suggests that an estrogenic response in breast cancer cells is indicated by an increase in ERβ expression. Tamoxifen decreased cell proliferation in both cell lines, while up regulating ERα in both cell lines, suggesting that antiestrogenic response is indicated by an increase in ERα expression. Although a change in the ERα/ERβ ratio may play a role in the effect seen in cell proliferation, this study indicates that ERβ is a poor prognosticator of cell proliferation in breast cancer and that ERα is a positive prognosticator of responsiveness to antiestrogen treatment.

Flaxseed alone or in combination with tamoxifen inhibits MCF-7 breast tumor growth in ovariectomized athymic mice with high circulating levels of estrogen

Flaxseed (FS) is rich in mammalian lignan precursors and α-linolenic acid, which have been suggested as having anticancer effects. Previous studies have shown that 10% FS inhibits the growth of human estrogen–dependent breast cancer (MCF-7) in athymic mice, and it enhances the inhibitory effect of tamoxifen (TAM). This study determined whether the effect of FS, alone or in combination with TAM, is dose dependent, and it explored the potential mechanism of action. Ovariectomized athymic mice with estradiol (E2) supplementation (1.7 mg/pellet, 60-day release) and established MCF-7 tumors were treated with basal diet control (0FS), 5% FS (5FS), 10% FS (10FS), and TAM (TAM/ 0FS; 5 mg/pellet, 60-day release), alone or in combination (TAM/ 5FS and TAM/10FS) for 8 weeks. Compared with control, 5FS and 10FS significantly inhibited tumor growth by 26% and 38%, respectively. TAM/0FS had an effect similar to the 10FS. TAM/ 5FS and TAM/10FS, respectively, induced significant 48% and 43% reductions in tumor size compared with 0FS, and 18% and 10% reductions compared with TAM/0FS. The relative uterine weight was significantly lower in all TAM groups compared with the control. The reduction of tumor growth resulted from decreased cell proliferation and increased cell apoptosis. TAM/ 5FS caused a significantly higher expression of estrogen receptor-α (ERα) compared with 5FS and TAM/0FS, whereas TAM/10FS had a higher ERα than 10FS and TAM/0FS. Compared with the control, progesterone receptor (PgR) expression was significantly reduced in all treatment groups, but insulin-like growth factor-1 (IGF-1) expression was reduced only by 10FS, TAM/5FS and TAM/10FS. Tumor cell proliferation was significantly positively associated with expression of PgR and IGF-1 and negatively associated with apoptosis and ERα. Apoptosis was only associated with ERα. In conclusion, FS inhibited MCF-7 tumor growth in a dose-dependent manner and enhanced the inhibitory effect of TAM due to the modulation of ER and growth factor signal transduction pathways.

Flaxseed attenuates the tumor growth stimulating effect of soy protein in ovariectomized athymic mice with MCF-7 human breast cancer xenografts

In several epidemiological studies, a phytoestrogen‐rich diet containing lignans and isoflavones is associated with reduced breast cancer risk, but experimental findings are controversial. In postmenopausal mammary cancer xenograft model, flaxseed (FS), a rich source of plant lignans, reduced breast cancer growth, while soy protein (SP), a rich source of isoflavones, enhanced it. The intake of phytoestrogens is increasing particularly among postmenopausal women, emphasizing the importance of elucidating their interactive effects on breast cancer. Our study determined the effect of FS and SP diets, alone and in combination, on the established human breast cancer MCF‐7 tumor growth in ovariectomized athymic nude mice. Tumor bearing mice were divided into 4 groups and fed for 25 weeks either the basal diet (BD), or BD supplemented with 10% FS, 20% SP or 10% FS and 20% SP. After estrogen deprivation, FS regressed the tumor size similar to that of control. SP initially regressed the tumors but starting at week 13, the tumors regressed significantly less than in control and 43% of the tumors were regrowing until the end of the experiment and were significantly larger in size than in control. The combination of SP with FS reduced the tumor growth similar to that of control, as suggested also by the reduced tumor cell proliferation index. In conclusion, dietary FS did not stimulate the growth of estrogen responsive MCF‐7 cancers in ovariectomized mice, while long‐term consumption of SP did. Furthermore, FS reduced the tumor growth stimulating effect of SP to the same level as control, suggesting tumor growth attenuating effect of FS.

Dietary flaxseed interaction with tamoxifen induced tumor regression in athymic mice with MCF-7 xenografts by downregulating the expression of estrogen related gene products and signal transduction pathways

Abstract Our previous short-term study has shown that 10% flaxseed (FS) inhibits the growth of human estrogen dependent estrogen receptor positive breast tumors (MCF-7) xenografts in ovariectomized (OVX) athymic mice and enhances the tumor inhibitory effect of tamoxifen (TAM). This study determined the long-term effect of 5% and 10% FS, with or without TAM, on the growth of MCF-7 xenografts in athymic mice and the potential mechanisms of actions. OVX mice with established MCF-7 tumors were treated with basal diet (control), 5% FS (5FS), 10% FS (10FS), and TAM (5 mg/pellet, 60-day release), alone or in combination, for 16 wk without estrogen supplementation. Tumor growth was monitored weekly. At sacrifice, the tumors were analyzed by immunohistochemistry for cell proliferation, apoptosis, and expression of estrogen-related genes and signal transduction pathways. Both 5FS and 10FS regressed the pretreatment tumor size by over 90% similar to control. TAM initially regressed the tumors but then induced a regrowth; thus, only a final 6% reduction from pretreatment tumor size was achieved, which was attenuated by combining TAM with 10FS but not with 5FS. TAM combined with 10FS regressed tumors to 55% of pretreatment tumor size due to decreased cell proliferation and increased apoptosis. The expressions of cyclin D1, estrogen receptor α, human epidermal growth factor receptor 2, and insulin-like growth factor I receptor in the TAM group were significantly reduced when TAM was combined with 5FS or 10FS. In conclusion, after long-term treatment, FS did not stimulate tumor growth and combined with TAM, regressed tumor size in part due to downregulation of the expression of estrogen-related gene products and signal transduction pathways.

Changes in biomarkers of estrogen receptor and growth factor signaling pathways in MCF-7 tumors after short- and long-term treatment with soy and flaxseed.

Previously we have shown that MCF-7 human breast tumor growth is stimulated after prolonged treatment with dietary soy protein isolate (SPI). However, the effects are attenuated when SPI is combined with flaxseed (FS). This study determined the changes that occur in tumor growth biomarkers, after both short- and long-term treatment with SPI, FS or their combination, to help identify signaling pathways potentially involved in SPI-stimulated tumor growth. Ovariectomized mice with established MCF-7 tumors were fed basal diet (control), 20%SPI, 10%FS, or SPI+FS for 2 or 25 weeks. After 2 weeks, there were no differences in tumor size, however, compared with control, SPI-treated tumors had higher IGF-IR and cyclin D1 while FS and SPI+FS-fed mice had lower pMAPK expression. After 25 weeks, SPI-treated tumors were larger, had higher proliferation, ERalpha, cyclin D1, IGF-IR, and pMAPK and lower ERbeta and HER2 levels. When combined with FS, however, the effects on these tumor biomarkers induced by SPI were attenuated. This study demonstrates that SPI and FS differently modulate tumor biomarkers of estrogen and growth factor signaling pathways, after both short- and long-term treatment, which may indicate a role of these pathways in the tumor stimulatory effects of SPI and the tumor inhibitory effects of FS.

Lignans are Accessible to Human Breast Cancer Xenografts in Athymic Mice

Lignan-rich diet has been linked with reduced breast cancer risk, and experimental studies have supported the hypothesis of lignans as cancer growth inhibiting compounds. However, it has not been clear if these compounds are accessible in the mammary tumor tissue in vivo. In this study, the accessibility and accumulation of lignans to breast cancer tissue was determined after oral administration of tritium labeled dietary lignan secoisolariciresinol diglucoside ( 3 H-SDG) to athymic mice bearing MCF-7 tumors. The 3 H-SDG administration increased tumor tissue radioactivity to the level similar to that in brain, skin, spleen, kidney, uterus, and lungs. The tumor tissue radioactivity was up to 92% of that found in serum, with the highest concentrations found in small (< 0.5 g) tumors. Accessibility of lignans to tumor tissue suggests that part of the anticancer activity of lignans may be due to their direct local effects on the breast cancer tissues.

Flaxseed and soy protein isolate, alone and in combination, differ in their effect on bone mass, biomechanical strength, and uterus in ovariectomized nude mice with MCF-7 human breast tumor xenografts.

In our previous study, flaxseed (FS) reduced while soy protein isolate (SPI) stimulated MCF-7 breast tumor growth in ovariectomized mice. In addition, combining SPI and FS resulted in a negation of SPI-induced tumor growth. In this study, the effects of SPI, FS, and their combination were further examined on mouse bone and uterus to further ensure overall safety of the breast cancer treatments. Ovariectomized mice with established MCF-7 xenografts were fed either a basal diet (control), or a basal diet supplemented with 10% FS, 20% SPI, or SPI + FS for 25 wk. Mouse bones were analyzed for mineral and biomechanical strength properties, and uterus weight was measured. The SPI group had a higher femur bone mineral density and biomechanical strength parameters (yield load, stiffness, and peak load) compared to control, while the FS group significantly increased femur stiffness and peak load. The SPI + FS group did not affect femur mineral, but significantly reduced whole femur area and length and increased femur yield load, stiffness, and peak load. Uterus weight was significantly increased by the SPI + FS group, while SPI alone induced an intermediate effect. In conclusion, all dietary treatments induced beneficial effects on bone in a preclinical mouse model of postmenopausal breast cancer. Although the SPI + FS and SPI groups exerted stimulatory effects on uterus weight, other histological parameters need to be measured to determine the overall safety of these breast cancer treatments on the uterus.

Interaction of Sesame Seed and Tamoxifen on Tumor Growth and Bone Health in Athymic Mice

Some premenopausal breast cancer patients use phytoestrogen-rich soy and flaxseed to alleviate side effects induced by drugs such as tamoxifen (TAM). Lignan-rich flaxseed protects against breast cancer and increases the effectiveness of TAM. This study determined the interactive effects of lignan-rich sesame seed (SS) and TAM on estrogen-responsive MCF-7 breast tumor growth and bone health in ovariectomized athymic mice under premenopausal-simulated conditions. Ovariectomized mice with an estrogen implant and established MCF-7 tumors were treated for 8 weeks as follows: (i) positive control fed basal diet (BD), (ii) SS group fed BD supplemented with 10% ground SS, (iii) TAM group with TAM implant fed BD, (iv) SS + TAM group with TAM implant fed BD supplemented with 10% SS, and (v) negative control fed BD with no estrogen implant. Palpable tumor data, adjusted for body weight, showed that SS does not inhibit MCF-7 tumor growth and tends to negate the tumor inhibitory effect of TAM by increasing cell proliferation and reducing apoptosis. SS alone and combined with TAM enhanced femur biomechanical strength but caused no differences in bone mineral content or bone mineral density in either the femur or lumbar vertebrae. SS is not protective and interacts adversely with TAM in MCF-7 breast tumors but induces beneficial effects on bone both alone and when combined with TAM.