Functional characterisation of VAV-interacting Krüppel-like factor (VIK) in breast cancer

Abstract

Background. VAV interacting Krüppel-like factor (VIK) is a novel transcription factor. Previously our lab reported a series of breast cancer tumour samples where VIK methylation was associated with an increased risk of recurrence in tamoxifen-treated patients, indicating a role for VIK in ER positive breast cancer and endocrine resistance. Additionally VIK has been shown to be involved in cell cycle regulation, interacting with CDK4 and VAV1. The cyclin D-CDK4/6-Rb pathway is frequently dysregulated in ER positive breast cancer. Combined treatment of palbociclib, a highly selective CDK4/6 inhibitor, with endocrine therapy substantially improved outcome of patients with ER positive metastatic breast cancer. Increasing clinical use means acquired resistance to palbociclib is emerging as a major clinical challenge. Results. VIK was confirmed to be subject to regulation by DNA methylation in breast cancer. VIK methylation correlated to transcriptional silencing of mRNA in both cancer cell lines and primary tumours. To determine the functional significance of loss of VIK expression, VIK was knocked down in unmethylated breast cancer cell lines and a normal breast epithelial cell line. Knockdown resulted in cell death via induction of apoptosis. VIK knockdown altered cell cycle progression from G1 to S phase. Expression of multiple regulatory cell cycle proteins was altered, differentially in normal and tumour cells. Treatment with the CDK4/6 inhibitor, palbociclib, in cells with reduced VIK expression resulted in decreased sensitivity to the drug, inducing a shift in IC50 value towards resistance. In a model of acquired resistance, T47D cells were cultured long-term with palbociclib generating resistant clones. VIK was significantly downregulated in all resistant clones to barely detectable mRNA levels, suggesting a role for VIK in resistance to CDK4/6 inhibition. Conclusion. This PhD has confirmed VIK is a novel epigenetically regulated gene in breast cancer. VIK expression is essential to both normal and tumour breast cell survival and is involved in the regulation of the G1/S phase transition in the cell cycle. Loss of VIK expression potentially contributes to the development of acquired resistance to CDK4/6 inhibitors.