NO1, a New Sigma 2 Receptor/TMEM97 Fluorescent Ligand, Downregulates SOCE and Promotes Apoptosis in the Triple Negative Breast Cancer Cell Lines

Abstract

(1) Background: The structure of the Sigma 2 receptor/TMEM97 (σ2RTMEM97) has recently been reported. (2, 3) Methods and results: We used genetic and biochemical approaches to identify the molecular mechanism downstream of σ2R/TMEM97. The novel σ2R/TMEM97 fluorescent ligand, NO1, reduced the proliferation and survival of the triple negative breast cancer cell lines (TNBC: MDA-MB-231 and MDA-MB-468 cell lines), due to NO1-induced apoptosis. Greater bioaccumulation and faster uptake of NO1 in MDA-MB-231 cells compared to MCF10A or MCF7 cell lines were also shown. Accordingly, elevated σ2R/TMEM97 expression was confirmed by Western blotting. In contrast to NO1, other σ2R/TMEM97 ligands, such as SM21 and PB28, enhanced MDA-MB-231 cell proliferation and migration. Store-operated calcium entry (SOCE) is crucial for different cancer hallmarks. Here, we show that NO1, but not other σ2R/TMEM97 ligands, reduced SOCE in MDA-MB-231 cells. Similarly, TMEM97 silencing in MDA-MB-231 cells also impaired SOCE. NO1 administration downregulated STIM1-Orai1 interaction, probably by impairing the positive regulatory effect of σ2R/TMEM97 on STIM1, as we were unable to detect interaction with Orai1. (4) Conclusion: σ2R/TMEM97 is a key protein for the survival of triple negative breast cancer cells by promoting SOCE; therefore, NO1 may become a good pharmacological tool to avoid their proliferation.

Arachidonic Acid Attenuates Cell Proliferation, Migration and Viability by a Mechanism Independent on Calcium Entry

“…Therefore, we incubated the MDA-MB-231 cells for 24 h with 8 µM of AA, and subsequently, upon loading cell with Fura-2, they were stimulated with AA (8µM) in the presence of extracellular CaCl 2 (1 mM), which did not evoke changes in the [Ca 2+ ] c ( Figure 1D). We have further explored whether treatment with AA might alter SOCE, a major Ca 2+ entry mechanism in non-excitable cells, whose regulation results are crucial for MDA-MB-231 cell proliferation [5,6]. As depicted in Figure 1E,F, preincubation of MDA-MB-231 cells for 5 min or 24 h with 8 µM of AA had no effect neither in TG-evoked release nor in SOCE in these cells.…”

“…Changes in the cytosolic Ca 2+ concentration ([Ca 2+ ] c ) due to Ca 2+ entry across the plasma membrane have been reported to be involved in cell proliferation [1][2][3][4]. In fact, several cancer cell lines present altered [Ca 2+ ] c due to dysregulation of Ca 2+ entry mechanisms, among others [3,[5][6][7][8][9]. The Ca 2+ currents I SOC , I CRAC , and I ARC were described as relevant inward Ca 2+ currents in non-excitable cells including several cancer cell types [10,11].…”

PGRMC1 Inhibits Progesterone-Evoked Proliferation and Ca2+ Entry Via STIM2 in MDA-MB-231 Cells

“…Nonetheless, sigma-2R has been recently claimed to be different from PGRMC1 based on genetic and biochemical evidence [ 57 , 58 , 59 , 60 ]. In MDA-MB-231 cells, we demonstrated that inhibition of sigma-2R with NO1 reported a different effect on TG-evoked SOCE to the one presented here by silencing PGRMC1 [ 26 ]. The different physiological roles of both proteins would confirm the previous observations done by other groups [ 55 ]; however, we could not discard that both proteins would be part of a macromolecular complex targeted by P 4 .…”

“…Considering the relevant role of the Ca 2+ - homeostasis on cell proliferation, even in MDA-MB-231 cells, as previously demonstrated by our research group [ 17 , 25 , 26 ], and the controversy regarding the role of P 4 in calcium homeostasis aforementioned, we analyzed the effect of P 4 on the Ca 2+ homeostasis in MDA-MB-231 cells [ 18 ]. MDA-MB-231 cells suspended in a Ca 2+ -free medium exhibited a small Ca 2+ entry when it was added back to the extracellular medium (1 mM CaCl 2 ; Figure 1 B, dotted line).…”

MRP1-Collateral Sensitizers as a Novel Therapeutic Approach in Resistant Cancer Therapy: An In Vitro and In Vivo Study in Lung Resistant Tumor

“…Therefore, we gathered inspiration from our library of sigma-2 receptors' ligands, that have been recently associated with CS [28,29,33,34]. Sigma-2 receptors, lately identified as TMEM97 proteins, are overexpressed in diverse types of cancers and their modulation leads to cell death upon the activation of a number of still investigated pathways [35][36][37][38][39][40][41][42][43][44][45]. Several sigma-2 receptor high-affinity ligands have been developed during recent decades (and lately reviewed), revealing cytotoxic activity through pathways that appear to be molecule-dependent and cell-type-dependent, with mechanisms that span from caspase-dependent apoptosis, autophagy, increase in ROS, and mithocondrial superoxide production [36,37,40,44].…”