Abstract P1-05-01: Chemokine-mediated nuclear translocation and novel nuclear role for LIM and SH3 Protein-1(LASP-1) in breast cancer

Abstract

Abstract Background: LASP-1 (LIM and SH3 protein-1) mediates cell migration, proliferation and survival in several breast cancer cell lines. LASP-1 is a scaffold protein that plays a critical role in cytoskeletal organization and cell motility. Silencing of LASP-1 in breast cancer cells inhibits cell migration and proliferation by 40%. We discovered that LASP-1 associates with chemokine receptors that are involved in migration of tumor cells or leukocytes in the tumor microenvironment or to the metastatic sites. LASP-1 directly binds to the chemokine receptors CXCR1, CXCR2, CXCR3 and CXCR4, suggesting that LASP-1 is a general mediator of CXC chemokine mediated migration and possibly cell survival. This established LASP-1 as a key component of the “CXC chemokine receptor chemosynapse”. This is highly significant as chemokine receptors CXCR2, CXCR4 and CXCR3 play a paramount role in primary and metastatic breast cancer. CXCR1 is involved in self-renewal of breast cancer stem cells that contribute to metastasis and drug resistance. The increase in expression level of LASP-1 with an increasing invasiveness of the tumor cells implicates a role for LASP-1 in tumor progression and lymph node metastases. Nuclear localization of LASP-1 (44% of LASP-1 positive specimens) was observed in tissue sections of aggressive and invasive breast cancer that positively correlated with poor survival rate. This is suggestive of a key role for nuclear LASP-1 in breast cancer progression. Methodology/Results: We investigated the chemokine-mediated nuclear translocation of LASP-1 in breast cancer and endothelial cell lines. Initially, cell surface expression of CXCR4 was profiled in breast cancer cell lines by FACS analysis. Medium to high CXCR4 expressers were stimulated with CXCL12 and the subcellular status of LASP-1 was examined by confocal microscopy. LASP-1 accumulated in the nucleus of the CXCL12-stimulated cells. Surprisingly, in MDA-MB-231 cells that were isolated from the mouse bone metastatic lesions had LASP-1 in the nucleus even in the absence of any stimulation by CXCL12. Additionally, the nuclear translocation of LASP-1 was observed in EGF- or heregulin-stimulated cells. Ligand activated CXCR2 in human endothelial cell line was also able to drive the nuclear accumulation of LASP-1. In order ascertain the role played by nuclear LASP-1 it was stably knocked down (KD). The control silenced (NS) and LASP-1 KD cells were grown in 3D-matrigel. Morphology of the 3D-clusters and the profile of the secreted cytokines were assessed by light microscopy and cytokine antibody microarray respectively along with additional biochemical methods and gene expression analysis by oligo microarray. Conclusions/Significance: We demonstrate here for the first time that LASP-1 translocates to the nucleus through activation of i) GPCRs -chemokine receptors CXCR2, CXCR4 and ii) Receptor tyrosine kinases - EGF receptor and HER2. Shuttling of LASP1 to the nucleus through the activation of a variety of receptors present in the tumor cells, macrophages, neutrophils and endothelial cells of the tumor microenvironment is of high significance. Silencing of LASP-1 in breast cancer cells revealed an altered profile of the 3D-clusters and the secreted cytokines. Citation Information: Cancer Res 2012;72(24 Suppl):Abstract nr P1-05-01.