VPC-18005 inhibits migration and invasion of prostate cell lines in vitro and in vivo. (A) PNT1B-Mock cells and (B) PNT1B-ERG cells were seeded in the upper chamber of a real-time cell analysis system (xCelligence) and treated with 5 ?M VPC-18005 (red line), YK-4-279 (blue line) or 0.01% DMSO (control; black line) at 24 h. The normalized cell index is a measure of the migration of the cells through the pores of the upper chamber and is used as the migration index. Dotted lines represent standard deviations (n = 3). The horizontal dotted red line indicates the level of migration the PNT1B-MOCK cells reached at 48 h in comparison to -ERG cells. (C) Rates of migration were determined by the slopes of the curves between 24?48 h for VPC-18005 (red) (p = 0.031, unpaired t-test) and YK-4-279 (blue) (p < 0.001, unpaired t-test) relative to DMSO control (black). (D) Quantitative analysis of PNT1B-ERG spheroid invasion into the surrounding matrix in the presence or absence of VPC-18005 (red line), YK-4-279 (blue line), or 0.01% DMSO (black line) over the period of 6 days. The rate of invasion between day 2 and 6 was significantly reduced in those cells treated with VPC-18005 (p = 0.02, unpaired t-test) and YK-4-279 (p = 0.005, unpaired t-test) compared to vehicle control. Error bars indicate standard error of the mean (n = 3). (E) Pre-stained PNT1B-Mock and PNT1B-ERG cells were microinjected into the yolk sac (green arrows) of the zebrafish, and the metastatic capability of the cells (white arrows) were detected using confocal microscope at day 2 and day 5. (F) Evaluation of compound toxicity to zebrafish embryos. Zebrafish embryos were treated with increasing concentration of VPC-18005 and YK-4-279 in their water. After 4 days, surviving embryos were counted. (G) Following 5 days of daily treatment, VPC-18005 reduced occurrence of metastasis in zebrafish grafted with PNT1B-ERG and VCaP cells. DMSO versus 1 ?M (p = 0.03/0.03, chi square) and 10 ?M (p = 0.002/<0.001, chi square) VPC-18005 (PNT1B/VCaP). YK-4-279 was significant only at 10 ?M (p = 0.02/0.04, chi square).