Tamoxifen-resistance remains a significant clinical problem in breast cancer treatment. Currently, hormonal therapies can achieve 50-60% breast cancer remission rate in ERα+ patients, who will mostly develop resistance. We have hypothesized that fat-1 gene therapy, which introduce an n-3 fatty acid desaturase to convert n-6 to n-3 polyunsaturated fatty acids (PUFA), as an adjuvant treatment will enhance hormonal therapy of breast cancer. We have previously reported that C57BL/6 fat-1 transgenic mice suppress breast cancer growth and the inflammation/chemically induced lung metastasis of E0771 murine mammary tumors. This corroborates the experimental reports about the tumor suppression effect of n-3 PUFA, a main component of fish or fish oil. In this report, we will present experimental evidence that fat-1 gene expression suppresses tamoxifen-resistant human breast cancer in vitro; in addition, expression of n-3 fatty acid desaturase encoded by fat-1 gene in transgenic mice significantly suppresses murine tamoxifen-resistant E0771TamR mammary tumor growth in vivo.

Methods: The tamoxifen-resistant human breast cancer cells MCF-7TamR and MDA-MB-231 (ER-), as well as the tamoxifen-sensitive human breast cancer T47D cells, were transfected with pST180 plasmid carrying fat-1 gene and the control empty pST128 plasmid. These cells were characterized in proliferation and caspase-3 apoptosis assays. The non-tumorigenic MCF-10A cells were also transfected with fat-1 gene to investigate the n-3 fatty acid desaturase effect on non-malignant human epithelial mammary cells. Two experimental groups of the fat-1 transgenic female C57BL/6 mice and wild type C57BL/6 control mice, were injected with tamoxifen-resistant E0771TamR tumor cells, or the parental E0771 cells, into breast fat pad of the 4th nipple and flank subcutaneously. Tumor sizes were measured daily until tumors reach 2 cm, when mice were sacrificed and tumor tissues and internal organs were harvested for fatty acid analysis and histology staining.

Results: In cell culture studies, introduction of fat-1 gene significantly reduced the proliferation rate of the tamoxifen-resistant MCF-7TamR cells (37.3%), MDA-MB-231 cells (27.5%), and T47D cells (67.7%). The MCF-10A cells transfected with fat-1 gene did not show any change of cell proliferation rate compared to the control cells. The caspase-3 activities in MCF-7TamR cells, MDA-MB-231 cells, and T47D cells were respectively increased by 1.3-, 2.1- and 5.6-fold when expressing fat-1 gene, as compared to the control cells. In vivo, n-3 fatty acid desaturase expressed in fat-1 mice significantly suppressed growth of tamoxifen-resistant murine E0771TamR mammary tumors grown in flank and breast fat pad, as compared to the control mice. The fat-1 growth inhibition effect is more dramatic for E0771TamR tumors than the parental E0771 tumors in fat-1 mice. Fatty acid analysis has shown a significant decrease of long chain n-6/n-3 PUFA ratio in breast tumor tissues (33.5%), flank tumor tissues (33.9%), kidney (32.7%), muscle (43.8%), and adipose tissues (69.1%) in fat-1 mice compared to wild type mice.

Conclusions: Introducing an n-3 fatty acid desaturase in tumor cells and transgenic animals has reduced long chain n-6/n-3 PUFA ratio and imposed an anti-tumor effect against tamoxifen-resistant breast cancer growth. Tamoxifen-sensitive breast cancer were also inhibited in Fat-1 gene therapy.

Citation Format: Raghunatha Reddy, Ian Bayles, Qiuhong He. Expression of n-3 fatty acid desaturase suppresses tamoxifen-resistant breast cancer in vitro and in fat-1 transgenic mice. [abstract]. In: Proceedings of the Eleventh Annual AACR International Conference on Frontiers in Cancer Prevention Research; 2012 Oct 16-19; Anaheim, CA. Philadelphia (PA): AACR; Cancer Prev Res 2012;5(11 Suppl):Abstract nr B103.