The IL-6 cytokine family is a group of signaling molecules with wide expression and function across vertebrates. Each member of the family signals by binding to its specific receptor and at least one molecule of gp130, which is the common transmembrane receptor subunit for the whole group. Signal transduction upon stimulation of the receptor complex results in the activation of multiple downstream cascades, among which, in mammary cells, the JAK-STAT3 pathway plays a central role. In this review, we summarize the role of the IL-6 cytokine family—specifically IL-6 itself, LIF, OSM, and IL-11—as relevant players during breast cancer progression. We have compiled evidence indicating that this group of soluble factors may be used for early and more precise breast cancer diagnosis and to design targeted therapy to treat or even prevent metastasis development, particularly to the bone. Expression profiles and possible therapeutic use of their specific receptors in the different breast cancer subtypes are also described. In addition, participation of these cytokines in pathologies of the breast linked to lactation and involution of the gland, as post-partum breast cancer and mastitis, is discussed.

The mammary gland is a secretory organ, which develops as a network of growing epithelial ducts composed of luminal and basal cells that invade the surrounding adipose tissue through a series of developmental cycles. Mammary stem cells (MaSCs) maintain an accurate tissue homeostasis, and their proliferation and cell fate determination are regulated by multiple hormones and local factors. The WNT pathway plays a critical role in controlling the enormous tissue expansion and remodeling during mammary gland development through the maintenance and differentiation of MaSCs, and its deregulation has been implicated in breast cancer (BC) initiation and progression. The R-spondins (RSPOs) are four secreted proteins that strongly enhance target cell sensitivity to WNT ligands. Moreover, leucine-rich repeat-containing G-protein-coupled receptors (LGRs) 4-6 are considered obligate high-affinity receptors for RSPOs and have been described as stem cell markers. Importantly, elevated RSPO expression has been recently identified in several tumor types from patients, including BC, and it has been reported that they play a significant role in mammary tumor progression in experimental models. In this review, exploring our present knowledge, we summarize the role of the RSPO-LGR axis as a WNT-enhancing signaling cascade in the MaSC compartment and during the normal and neoplastic mammary gland development. In addition, we include an updated expression profile of the RSPOs and their action mediators at the cell membrane, the LGRs, and the ubiquitin-ligases ZNRF3/ RNF43, in different BC subtypes. Finally and based on these data, we discuss the significance of tumor-associated alterations of these proteins and their potential use as molecular targets for detection and treatment of BC.

R-spondin3 (RSPO3) is a member of a family of secreted proteins that enhance Wnt signaling pathways in diverse processes, including cancer. However, the role of RSPO3 in mammary gland and breast cancer development remains unclear. In this study, we show that RSPO3 is expressed in the basal stem cell-enriched compartment of normal mouse mammary glands but is absent from committed mature luminal cells in which exogenous RSPO3 impairs lactogenic differentiation. RSPO3 knockdown in basal-like mouse mammary tumor cells reduced canonical Wnt signaling, epithelial-to-mesenchymal transition-like features, migration capacity, and tumor formation Conversely, RSPO3 overexpression, which was associated with some LGR and RUNX factors, highly correlated with the basal-like subtype among patients with breast cancer. Thus, we identified RSPO3 as a novel key modulator of breast cancer development and a potential target for treatment of basal-like breast cancers. These findings identify RSPO3 as a potential therapetuic target in basal-like breast cancers. http://cancerres.aacrjournals.org/content/canres/78/16/4497/F1.large.jpg .

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We have recently identified R-spondin3 (RSPO3) as a novel key modulator of breast cancer development that may become a potential target for treatment of the basal subtype, which lacks efficient therapeutic options to date. RSPO3 is expressed in the basal stem cell-enriched compartment of mouse mammary glands whereas it is absent in the mature luminal cells. RSPO3 knockdown in mouse mammary tumor cells reduces canonical Wnt signaling pathway, epithelial-to-mesenchymal transition (EMT)-like features, migration capacity, and tumor formation in vivo. To evaluate the relevance of RSPO3 in human breast cancer, we analyzed samples from 75 patients with ductal infiltrating breast carcinomas, where we found a high prevalence of positive immunoreactivity for RSPO3 (70%). Expression data of The Cancer Genome Atlas Network from 1985 human breast carcinoma samples showed that RSPO3 is primarily expressed in Claudin-low breast cancer subtype, which is characterized for having stem cells features. In culture assays revealed that knockdown of RSPO3 expression reduced SOX2 expression and impaired mammary tumor cell ability to form mammospheres. Previous reports suggested the potential involvement of RUNX1-CBFβ on RSPO3 transcription in mammary tumor cells. Therefore, we evaluated if pharmacologic inhibition of RUNX1 activity would reduce RSPO3 expression and activity in the cancer stem cell compartment. Treatment of breast cancer cells with AI-10-104, a small molecule that inhibits CBFβ-RUNX interaction, reduced RSPO3 mRNA and protein levels, and reduced the migration ability of MDA-MB231 breast cancer cells, ability that was recovered upon treatment with recombinant RSPO3. In addition, AI-10-104 inhibited the expression of canonical Wnt pathway targets, EMT and stem-cell markers. Therefore, our results demonstrate that inhibition of RUNX-CBFβ transcriptional activity leads to a down-regulation of RSPO3 expression, constituting a new molecular target for therapeutic modulation in breast cancer treatment. Citation Format: Carla M. Felcher, Johanna M. Tocci, Martin E. Garcia Sola, John H. Bushweller, Lucio H. Castilla, Edith C. Kordon. Inhibition of RUNX-CBFβ binding reduces RSPO3 expression and EMT features in breast cancer cells [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 5041.