The critical role of calcium signalling in processes related to cancer cell proliferation and invasion has seen a focus on pharmacological inhibition of overexpressed ion channels in specific cancer subtypes as a potential therapeutic approach. However, despite the critical role of calcium in cell death pathways, pharmacological activation of overexpressed ion channels has not been extensively evaluated in breast cancer. Here we define the overexpression of transient receptor potential vanilloid 4 (TRPV4) in a subgroup of breast cancers of the basal molecular subtype. We also report that pharmacological activation of TRPV4 with GSK1016790A reduced viability of two basal breast cancer cell lines with pronounced endogenous overexpression of TRPV4, MDA-MB-468 and HCC1569. Pharmacological activation of TRPV4 produced pronounced cell death through two mechanisms: apoptosis and oncosis in MDA-MB-468 cells. Apoptosis was associated with PARP-1 cleavage and oncosis was associated with a rapid decline in intracellular ATP levels, which was a consequence of, rather than the cause of, the intracellular ion increase. TRPV4 activation also resulted in reduced tumour growth in vivo. These studies define a novel therapeutic strategy for breast cancers that overexpress specific calcium permeable plasmalemmal ion channels with available selective pharmacological activators.

Processes that are important in cancer progression, such as sustained cell growth, invasion to other organs, and resistance to cell death inducers, have a clear overlap with pathways regulated by Ca 2+ signaling. It is therefore not surprising that proteins important in Ca 2+ signaling, sometimes referred to as the "Ca 2+ signaling toolkit," can contribute to cancer cell proliferation and invasiveness, and the ability of agents to induce cancer cell death. Ca 2+ signaling is also critical in other aspects of cancer progression, including events in the tumor microenvironment and processes involved in the acquisition of resistance to anticancer therapies. This review will consider the role of Ca 2+ signaling in tumor progression and highlight areas in which a better understanding of the interplay between the Ca 2+ -signaling toolkit and tumorigenesis is still required.

Cancer is a major cause of death. The diversity of cancer types and the propensity of cancers to acquire resistance to therapies, including new molecularly targeted and immune-based therapies, drives the search for new ways to understand cancer progression. The remodelling of calcium (Ca) signalling and the role of the Ca signal in controlling key events in cancer cells such as proliferation, invasion and the acquisition of resistance to cell death pathways is well established. Most of the work defining such changes has focused on Ca permeable Transient Receptor Potential (TRP) Channels and some voltage gated Ca channels. However, the identification of ORAI channels, a little more than a decade ago, has added a new dimension to how a Ca influx pathway can be remodelled in some cancers and also how calcium signalling could contribute to tumour progression. ORAI Ca channels are now an exemplar for how changes in the expression of specific isoforms of a Ca channel component can occur in cancer, and how such changes can vary between cancer types (e.g. breast cancer versus prostate cancer), and even subtypes (e.g. oestrogen receptor positive versus oestrogen receptor negative breast cancers). ORAI channels and store operated Ca entry are also highlighting the diverse roles of Ca influx pathways in events such as the growth and metastasis of cancers, the development of therapeutic resistance and the contribution of tumour microenvironmental factors in cancer progression. In this review we will highlight some of the studies that have provided evidence for the need to deepen our understanding of ORAI Ca channels in cancer. Many of these studies have also suggested new ways on how we can exploit the role of ORAI channels in cancer relevant processes to develop or inform new therapeutic strategies.

The remodeling of specific calcium-permeable ion channels is a feature of some breast cancer subtypes. ORAI1 is a protein that forms a calcium-permeable ion channel responsible for store-operated calcium entry (SOCE) in a variety of cell types. ORAI3, a related isoform, is not a regulator of SOCE in most cell types. However, ORAI3 does control SOCE in many estrogen receptor-positive breast cancer cell lines, where it also controls proliferation. ORAI1 is a well-characterized regulator of the proliferation and migration of many basal breast cancer cells; however, the role of ORAI3 in these types of breast cancer cells remains unclear. Here, we sought to define ORAI1 and ORAI3 expression in breast cancer cell lines of different molecular subtypes and assess the potential role and regulation of ORAI3 in basal breast cancer cells. Our study demonstrates that elevated ORAI1 is a feature of basal-like breast cancers, while elevated ORAI3 is a feature of luminal breast cancers. Intriguingly, we found that ORAI3 is over-expressed in the mesenchymal subtype of triple-negative breast cancer. Given this, we assessed ORAI3 levels in the presence of two inducers of the mesenchymal phenotype, hypoxia and epidermal growth factor (EGF). Hypoxia induced ORAI3 levels in basal breast cancer cell lines through a pathway involving hypoxia-inducible factor-1 alpha (HIF1α. The silencing of ORAI3 attenuated hypoxia-associated phosphorylation of the EGF receptor (EGFR) and the expression of genes associated with cell migration and inflammatory/immune responses in the MDA-MB-468 model of basal breast cancer. Although elevated ORAI3 levels were not associated with survival; basal, estrogen receptor-negative and triple-negative breast cancers with high ORAI3 and low ORAI1 levels were associated with poorer clinical outcomes. This study defines ORAI3 as a potential fine-tuner for processes relevant to the progression of basal breast cancers.

Store-operated calcium (Ca 2+ ) entry is an important homeostatic mechanism in cells, whereby the release of Ca 2+ from intracellular endoplasmic reticulum stores triggers the activation of a Ca 2+ influx pathway. Mediated by Orai1, this Ca 2+ influx has specific and essential roles in biological processes as diverse as lactation to immunity. Although pharmacological inhibitors of this Ca 2+ influx mechanism have helped to define the role of store-operated Ca 2+ entry in many cellular events, the lack of isoform specific modulators and activators of Orai1 has limited our full understanding of these processes.Here we report the identification and synthesis of an Orai1 activity enhancer that concurrently potentiated Orai1 Ca 2+ -dependent inactivation (CDI). This unique enhancer of Orai1 had only a modest effect on Orai3 with weak inhibitory effects at high concentrations in intact MCF-7 breast cancer cells. The Orai1 enhancer heightened vascular smooth muscle cell migration induced by platelet-derived growth factor and the unique store-operated Ca 2+ entry pathway present in skeletal muscle cells. These studies show that IA65 is an exemplar for the translation and development of Orai isoform selective agents. The ability of IA65 to activate CDI demonstrates that agents can be developed that can enhance Orai1-mediated Ca 2+ influx but avoid the cytotoxicity associated with sustained Orai1 activation. IA65 and/or future analogues with similar Orai1-and CDIactivating properties could function to fine-tune physiological processes important in specific disease states, such as cellular migration and immune cell function.