Abstract P2-05-08: An assessment of the potential role of intracellular Ca2+ store regulators in breast cancer cells

Abstract

Abstract Background: Calcium (Ca2+) regulates many crucial cellular processes including cell survival, proliferation and death. Endoplasmic reticulum (ER) Ca2+ store levels are critical in the death-inducing effects of some anti-cancer agents. Modulators of ER Ca2+ signalling, such as neuronal calcium sensor-1 (NCS-1) may therefore represent new therapeutic opportunities to enhance the effect of some anti-cancer agents. NCS-1 is associated with poorer survival in breast cancer patients. However, the expression of NCS-1 in specific breast cancer molecular subtypes and its potential role in intracellular Ca2+ signalling in breast cancer cells has not been fully explored. Aim: To assess expression of NCS-1 in breast cancer molecular subtypes and assess the effect of silencing NCS-1 on intracellular Ca2+ homeostasis and on sensitivity to doxorubicin treatment in MDA-MB-231 breast cancer cells. Methods: NCS-1 levels were assessed in breast cancer molecular subtypes based on PAM50 subtyping using the TCGA public breast cancer database. Intracellular Ca2+ changes as a consequence of siRNA-mediated silencing of NCS-1 were evaluated using a Fluorescence Imaging Plate Reader (FLIPR) in MDA-MB-231 cells expressing the genetically-encoded Ca2+ indicator GCaMP6m. The effect of NCS-1 silencing on MDA-MB-231 cells treated with doxorubicin (0.03 and 1 μM, 24 h) was evaluated by cell nuclear enumeration (Hoechst 33342 staining) and the percentage of dead cells (propidium iodide staining). Images were acquired using an automated epifluorescence microscope (ImageXpress Micro). Results: Levels of NCS-1 were higher in the basal molecular subtype compared to other molecular subtypes. NCS-1 silencing promoted cell death induced by 1 μM doxorubicin. NCS-1 silencing had no major effect on cytosolic free Ca2+ levels as a result of either IP3-mediated Ca2+ store release with the purinergic receptor activator ATP or the protease activated receptor activator trypsin. However, NCS-1 silencing suppressed constitutive Ca2+ influx in MDA-MB-231 breast cancer cells. The expression of NCS-1 was positively correlated with the Ca2+ entry channel Orai1 in breast cancers on the TCGA database. Orai1 is associated with increased migration and invasiveness in some breast cancers. Discussion: These studies suggest that elevation in NCS-1 is a feature of breast cancers of the basal molecular subtype and that NCS-1 is a regulator of the cytotoxic effects of doxorubicin. Further studies are required to determine if this effect is related to changes in constitutive Ca2+ influx that may be mediated via Orai1. Citation Format: Bong AH, Roberts-Thomson SJ, Milevskiy MJ, Monteith GR. An assessment of the potential role of intracellular Ca2+ store regulators in breast cancer cells [abstract]. In: Proceedings of the 2018 San Antonio Breast Cancer Symposium; 2018 Dec 4-8; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2019;79(4 Suppl):Abstract nr P2-05-08.