Numerous studies on the biological mechanism of breast cancer have identified a number of potential therapeutic molecular targets. In this context, one type of potential candidates appears to be agents that target the actin cytoskeleton of cancer cells or regulate actin cytoskeleton dynamics. The aim of the present study was to study the impact of altered actin transport between the cytoplasm and nucleus by the downregulation of importin-9 (IPO9) in breast adenocarcinoma MCF-7 cells exposed to an apoptosis-inducing combination of garlic-derived S-allyl-L-cysteine sulfoxide (alliin) and paclitaxel (PTX). The expression of IPO9 was downregulated by the transfection of non-aggressive breast cancer MCF-7 cells with siRNA against IPO9. The altered expression of IPO9 and cofilin-1 (CFL1) was examined using western blotting. Moreover, the effect of the downregulation of IPO9 on cell death induced by the combination of PTX and alliin was also investigated. The alterations of IPO9 and CFL1 levels were also related with F-actin organizational changes and F-actin fluorescence intensity in the nuclear/perinuclear area of the cells. The results presented here indicate that alliin and PTX act synergistically to promote and potentiate apoptosis in MCF-7 cells. Furthermore, using RNA interference technique, we showed that downregulation of IPO9 expression was correlated with a significant reduction in the apoptotic cell population as well as with a decrease in F-actin content in whole cells, and in the cortical and nuclear/perinuclear areas of the cells. Simultaneously, the downregulation of IPO9 was also accompanied by the increased post-translational expression of CFL1. Furthermore, the data obtained in the present study allow us to conclude that CFL1 itself does not translocate actin into the cell nucleus but this transport requires the functional expression of IPO9.