Dynamic regulation of ROCK in tumor cells controls CXCR4-driven adhesion events.

CXCR4 is a chemokine receptor often found aberrantly expressed on metastatic tumor cells. To investigate CXCR4 signaling in tumor cell adhesion, we stably overexpressed CXCR4 in MCF7 breast tumor cells. Cell attachment assays demonstrate that stimulation of the receptor with its ligand, CXCL12, promotes adhesion of MCF7-CXCR4 cells to both extracellular matrix and endothelial ligands. To more closely mimic the conditions experienced by a circulating tumor cell, we performed the attachment assays under shear stress conditions. We found that CXCL12-induced tumor cell attachment is much more pronounced under flow. ROCK is a serine/threonine kinase associated with adhesion and metastasis, which is regulated by CXCR4 signaling. Thus, we investigated the contribution of ROCK activity during CXC12-induced adhesion events. Our results demonstrate a biphasic regulation of ROCK in response to adhesion. During the initial attachment, inhibition of ROCK activity is required. Subsequently, re-activation of ROCK activity is required for maturation of adhesion complexes and enhanced tumor cell migration. Interestingly, CXCL12 partially reduces the level of ROCK activity generated by attachment, which supports a model in which stimulation with CXCL12 regulates tumor cell adhesion events by providing an optimal level of ROCK activity for effective migration.

Glyceollin I, a novel antiestrogenic phytoalexin isolated from activated soy.

Glyceollins, a group of novel phytoalexins isolated from activated soy, have recently been demonstrated to be novel antiestrogens that bind to the estrogen receptor (ER) and inhibit estrogen-induced tumor progression. Our previous publications have focused specifically on inhibition of tumor formation and growth by the glyceollin mixture, which contains three glyceollin isomers (I, II, and III). Here, we show the glyceollin mixture is also effective as a potential antiestrogenic, therapeutic agent that prevents estrogen-stimulated tumorigenesis and displays a differential pattern of gene expression from tamoxifen. By isolating the individual glyceollin isomers (I, II, and III), we have identified the active antiestrogenic component by using competition binding assays with human ERalpha and in an estrogen-responsive element-based luciferase reporter assay. We identified glyceollin I as the active component of the combined glyceollin mixture. Ligand-receptor modeling (docking) of glyceollin I, II, and III within the ERalpha ligand binding cavity demonstrates a unique type II antiestrogenic confirmation adopted by glyceollin I but not isomers II and III. We further compared the effects of glyceollin I to the antiestrogens, 4-hydroxytamoxifen and ICI 182,780 (fulvestrant), in MCF-7 breast cancer cells and BG-1 ovarian cancer cells on 17beta-estradiol-stimulated expression of progesterone receptor and stromal derived factor-1alpha. Our results establish a novel inhibition of ER-mediated gene expression and cell proliferation/survival. Glyceollin I may represent an important component of a phytoalexin-enriched food (activated) diet in terms of chemoprevention as well as a novel therapeutic agent for hormone-dependent tumors.