Abstract 1125: Secretory pathway calcium ATPase-2 (SPCA2) regulates metastasis by suppressing mesenchymal markers in triple negative breast cancer cell lines

Abstract

Abstract Introduction: Over 90% of cancer deaths in breast cancer are associated with metastasis. Epithelial-mesenchymal transition (EMT) is the hallmark of metastasis. Dysregulation of the Ca2+ toolkit has profound consequences for tumor growth and metastasis, raising hopes for novel avenues of therapeutic intervention. The Secretory Pathway Ca2+-ATPase Isoform 2 (SPCA2) transports Ca2+ from cytoplasm to the Golgi, and elicits Ca2+ influx by interacting with plasma membrane Ca2+ channels. Previously, we showed that SPCA2 is implicated in breast cancer progression (Feng et al., Cell 2010). Furthermore, low SPCA2 expression is associated with triple negative breast cancers (TNBC), which are highly metastatic. Therefore, we investigated if ectopic expression of SPCA2 modulates EMT in TNBC cell lines. Methods: TCGA invasive breast carcinoma project datasets were accessed through cBioPortal. Gene expression was determined by qPCR and protein expression by Immunoblotting and confocal microscopy. Live cell calcium imaging was performed using Fura-2 AM dye. NSG mice were used for in vivo studies. Results: Low SPCA2 expression was associated with poor survival in TNBC patients (n=255, P < 0.05). TNBC cell lines show low SPCA2 expression compared to receptor positive cell lines, when normalized to non-tumorigenic epithelial cell line MCF-10A. Similarly, significantly low SPCA2 expression (P < 0.001) was observed in TNBC patients compared to receptor positive subtypes. Interestingly, we did not observe these results in SPCA1, the housekeeping SPCA isoform, revealing isoform-specific function of Golgi calcium pumps in breast cancer subtypes. Ectopic expression of SPCA2 in TNBC cell line MDA-MB-231 significantly increased baseline intracellular Ca2+ levels (P < 0.001) and uptake of extracellular Ca2+ (P < 0.001) through store independent calcium entry. Increased SPCA2 expression suppressed mesenchymal gene markers (CDH2, SNAI1, SNAI2, VIM and ZEB1, P < 0.05), and decreased cell migration in vitro (P < 0.05) in TNBC cell lines. Overexpression of SPCA2 in MDA-MB-231-luc-D3H2LN cell line reduced metastasis in vivo (quantified 5-weeks after injection in mammary fat pad, P < 0.01) (n=6) compared to control group. Treatment of metastatic TNBC patients with histone deacetylase (HDAC) inhibitors is currently being evaluated in clinical trials. We found treatment of TNBC cell lines with FDA-approved HDAC inhibitors vorinostat and romidepsin elevated SPCA2 expression in a dose-dependent manner (P < 0.05), simultaneously decreasing mesenchymal gene expression (P < 0.05). Conclusion: These novel findings point to a causal link between low SPCA2 levels, poor prognosis, and the epithelial-mesenchymal transition required for breast cancer metastasis. Restoration of SPCA2 expression in TNBC by HDAC inhibitors may have therapeutic potential. Citation Format: Monish Ram Makena, Donna K. Dang, Myungjun Ko, Manuj Bandral, Rajini Rao. Secretory pathway calcium ATPase-2 (SPCA2) regulates metastasis by suppressing mesenchymal markers in triple negative breast cancer cell lines [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 1125.