Inactivation of the WASF3 gene in prostate cancer cells leads to suppression of tumorigenicity and metastases

Abstract

Background:The WASF3 protein is involved in cell movement and invasion, and to investigate its role in prostate cancer progression we studied the phenotypic effects of knockdown in primary tumors and cell lines.Methods:ShRNA was used to knockdown WASF3 function in prostate cell lines. Cell motility (scratch wound assay), anchorage independent growth and in vivo tumorigenicity and metastasis were then compared between knockdown and wild-type cells.Results:Increased levels of expression were seen in high-grade human prostate cancer and in the PC3 and DU145 cell lines. Inactivation of WASF3 using shRNAs reduced cell motility and invasion in these cells and reduced anchorage independent growth in vitro. The loss of motility was accompanied by an associated increase in stress fiber formation and focal adhesions. When injected subcutaneously into severe combined immunodeficiency (SCID) mice, tumor formation was significantly reduced for PC3 and DU145 cells with WASF3 knockdown and in vivo metastasis assays using tail vain injection showed a significant reduction for PC3 and DU145 cells. The loss of the invasion phenotype was accompanied by down-regulation of matrix metalloproteinase 9.Conclusions:Overall, these observations demonstrate a critical role for WASF3 in the progression of prostate cancer and identify a potential target to control tumorigenicity and metastasis.

HSP90 and HSP70 Proteins Are Essential for Stabilization and Activation of WASF3 Metastasis-promoting Protein

“…These observations support the idea that ABL is a client protein for HSP90. 3 When PC3 cells were treated with 50 M STI-571 (Gleevec or imatinib), which inhibits ABL, for 24 h, the phosphorylation level of WASF3 was reduced significantly (Fig. 2F).…”

“…HSP90 has emerged as a promising target for the treatment of cancer and inhibition of its function by the 17-AAG drug leads to decreased cancer cell motility and invasion (15)(16)(17). We have recently demonstrated that WASF3 is important in controlling cell motility and invasion in breast and prostate cancer cells (2,3,5). To determine whether the interaction between WASF3 and HSP90 maintains protein stability, we inactivated HSP90 function using 17-AAG, which binds directly to the essential ATP/ADP binding pocket in the N-terminal region of HSP90 (8,18).…”

“…3C). We have recently demonstrated that WASF3 is important in controlling cell motility and invasion in breast cancer cells (2,3) and IP of WASF3 from MDA-MB-231 and SkBr3 also identified HSP70 in the immunocomplex (Fig. 3D).…”

“…We have demonstrated that knockdown of WASF3 in breast and prostate cancer cells has a profound effect on their ability to migrate and invade in vitro (2,3) and to metastasize in vivo (3,4). Specifically, WASF3 appears to coordinate the development of lamellipodia (2) at the leading edges of cells, and loss of its function prevents motility.…”

“…Motility and Invasion Assays-The wound-closure assay was performed as described previously (3). Invasion assays were performed in Matrigel-coated modified Boyden chambers with 8-m pore size filters (BD Biosciences).…”