A targeted siRNA based screen to identify calcium transporters involved in the calcium-dependent regulation of ABCC3 gene expression in a model of breast cancer epithelial-mesenchymal transition (EMT) [Poster]

Introduction: Invasion and metastasis are hallmarks of cancer and are linked to resistance to anticancer therapies. Increased expression of members of the ATP-binding cassette (ABC) transporter superfamily is also implicated in resistance to chemotherapeutics, and has been linked to invasive behaviour. We have previously identified the Ca²⁺- dependent upregulation of ABCC3 gene expression in a model of breast cancer epithelial-mesenchymal transition (EMT), a process involved in metastasis. The Ca²⁺ transporter(s) involved in the regulation of ABCC3 gene expression in this model of EMT are yet to be identified.

Aims: To identify the Ca²⁺ channels and/or pumps involved in the Ca²⁺-dependent upregulation of ABCC3 transporter gene expression in a breast cancer model of epidermal growth factor (EGF)-induced EMT.

Methods: To assess the role of Ca²⁺ transporters in ABCC3 gene expression, an siRNA-based screen targeting 24 Ca²⁺ channels and pumps using Dharmacon On-TARGETplus SMARTpool siRNAs was performed. MDA-MB-468 basal-like breast cancer cells were transfected with each siRNA SMARTpool for 48 h. Following transfection, cells were serum starved (0.5% FBS) for 24 h before induction of EMT via treatment with EGF (50 ng/mL) for 48 h. Quantitative RT-PCR was used to assess changes in ABCC3 mRNA expression following siRNA and EGF treatment.

Results: siRNA mediated silencing of the Ca²⁺ permeable ion channel, TRPM7, a previously characterised partial regulator of EMT in this model, did not inhibit ABCC3 gene expression following 24 h EGF treatment. However, siRNA screening identified other Ca²⁺ transporters that may act as potential regulators of EGF-induced ABCC3 expression at 48 h, for further characterisation.

Discussion: Induction of EMT by EGF in MDA-MB-468 breast cancer cells results in the Ca²⁺-dependent upregulation of ABCC3 transporter gene expression. Further studies are required to characterise the Ca2+ transporters involved in the regulation of ABCC3 expression in this model.