Hormonal Therapy Resistance and Breast Cancer: Involvement of Adipocytes and Leptin.

Obesity, a recognized risk factor for breast cancer in postmenopausal women, is associated with higher mortality rates regardless of menopausal status, which could in part be explained by therapeutic escape. Indeed, adipose microenvironment has been described to influence the efficiency of chemo- and hormonal therapies. Residual cancer stem cells could also have a key role in this process. To understand the mechanisms involved in the reduced efficacy of hormonal therapy on breast cancer cells in the presence of adipose secretome, human adipose stem cells (hMAD cell line) differentiated into mature adipocytes were co-cultured with mammary breast cancer cells and treated with hormonal therapies (tamoxifen, fulvestrant). Proliferation and apoptosis were measured (fluorescence test, impedancemetry, cytometry) and the gene expression profile was evaluated. Cancer stem cells were isolated from mammospheres made from MCF-7. The impact of chemo- and hormonal therapies and leptin was evaluated in this population. hMAD-differentiated mature adipocytes and their secretions were able to increase mammary cancer cell proliferation and to suppress the antiproliferative effect of tamoxifen, confirming previous data and validating our model. Apoptosis and cell cycle did not seem to be involved in this process. The evaluation of gene expression profiles suggested that STAT3 could be a possible target. On the contrary, leptin did not seem to be involved. The study of isolated cancer stem cells revealed that their proliferation was stimulated in the presence of anticancer therapies (tamoxifen, fulvestrant, doxorubicine) and leptin. Our study confirmed the role of adipocytes and their secretome, but above all, the role of communication between adipose and cancer cells in interfering with the efficiency of hormonal therapy. Among the pathophysiological mechanisms involved, leptin does not seem to interfere with the estrogenic pathway but seems to promote the proliferation of cancer stem cells.

Adipocyte/breast cancer cell crosstalk in obesity interferes with the anti-proliferative efficacy of tamoxifen.

BACKGROUND: Obesity is a well-known risk factor of breast cancer in post-menopausal women that also correlates with a diminished therapeutic response. The influence of adipocytes and their secretome, i.e. adipokines, on the efficacy of hormone therapy has yet to be elucidated. METHODS: We investigated, ex vivo, whether mature adipocytes, differentiated from adipose stem cells of normal-weight (MA20) or obese (MA30) women, and their secretions, were able to counteract the effects of tamoxifen (Tx) which is known to decrease neoplastic cell proliferation. RESULTS: In a tridimensional model and in a model of co-culture, the anti-proliferative effect of Tx on MCF-7 cancer cells was counteracted by MA30. These two models highlighted two different specific gene expression profiles for genes encoding cytokines or involved in angiogenesis based on the adipocyte microenvironment and the treatment. Thus it notably showed altered expression of genes such as TNFα that correlated with IL-6. In addition, leptin, IL-6 and TNFα, at concentrations reflecting plasma concentrations in obese patients, decreased the anti-proliferative efficacy of 4-hydroxytamoxifen (a major active metabolite of Tx). CONCLUSIONS: These findings bring insights on adipocytes and mammary cancer cell interactions in Tx therapy, particularly in overweight/obese people. Indeed, patient' adipokine status would give valuable information for developing individual strategies and avoid resistance to treatment.

Supernatants of Adipocytes From Obese Versus Normal Weight Women and Breast Cancer Cells: In Vitro Impact on Angiogenesis.

Breast cancer is correlated with a higher risk of metastasis in obese postmenopausal women. Adipokines, whose plasma concentrations are modulated in obese subjects and adipocytes surround mammary cells, suggesting that adipocyte secretome affect mammary tumorogenesis. We hypothesize that mature adipocyte secretions from obese women conditioned or not by breast neoplasic cells, increase changes on the angiogenesis stages. Supernatants of human mature adipocytes, differentiated from stem cells of either adipose tissue of normal weight (MA20) or obese (MA30) women or obtained from co-cultures between MA20 and MA30 and breast cancer cell line MCF-7, were collected. The impact of these supernatants was investigated on proliferation, migration, and tube formation by endothelial cells (HUVEC). MA20 and MA30 showed a preservation of their "metabolic memory" (increase of Leptin, ObR, VEGF, CYP19A1, and a decrease of Adiponectin expression in MA30 compared to MA20). Supernatants from obese-adipocytes increased HUVEC proliferation, migration, and sprouting like with supernatants obtained from co-cultures of MA/MCF-7 regardless the women's BMI. Additional analyses such as the use of neutralizing antibodies, analysis of supernatants (Milliplex®) and variations in gene expression (qRT-PCR), strongly suggest an implication of IL-6, or a synergistic action among adipokines, probably associated with that of VEGF or IL-6. As a conclusion, supernatants from co-cultures of MA30 and MCF-7 cells increase proliferation, migration, and sprouting of HUVEC cells. These results provide insights into the interaction between adipocytes and epithelial cancer cells, particularly in case of obesity. The identification of synergistic action of adipokines would therefore be a great interest in developing preventive strategies. J. Cell. Physiol. 232: 1808-1816, 2017. © 2016 Wiley Periodicals, Inc.

Wnt and Neuregulin1/ErbB signalling extends 3D culture of hormone responsive mammary organoids.

The development of in vitro culture systems quantitatively and qualitatively recapitulating normal breast biology is key to the understanding of mammary gland biology. Current three-dimensional mammary culture systems have not demonstrated concurrent proliferation and functional differentiation ex vivo in any system for longer than 2 weeks. Here, we identify conditions including Neuregulin1 and R-spondin 1, allowing maintenance and expansion of mammary organoids for 2.5 months in culture. The organoids comprise distinct basal and luminal compartments complete with functional steroid receptors and stem/progenitor cells able to reconstitute a complete mammary gland in vivo. Alternative conditions are also described that promote enrichment of basal cells organized into multiple layers surrounding a keratinous core, reminiscent of structures observed in MMTV-Wnt1 tumours. These conditions comprise a unique tool that should further understanding of normal mammary gland development, the molecular mechanism of hormone action and signalling events whose deregulation leads to breast tumourigenesis.