Therapeutic targeting of tetraspanin8 in epithelial ovarian cancer invasion and metastasis.

Epithelial ovarian cancer (EOC) invasion and metastasis are complex phenomena that result from the coordinated action of many metastatic regulators and must be overcome to improve clinical outcomes for patients with these cancers. The identification of novel therapeutic targets is critical because of the limited success of current treatment regimens, particularly in advanced-stage ovarian cancers. In this study, we found that tetraspanin 8 (TSPAN8) is overexpressed in about 52% (14/27) of EOC tissues and correlates with poor survival. Using small interfering RNA-mediated TSPAN8 knockdown and a competition assay with purified TSPAN8 large extracellular loop (TSPAN8-LEL) protein, we identified TSPAN8-LEL as a key regulator of EOC cell invasion. Furthermore, monotherapy with TSPAN8-blocking antibody we developed shows that antibody-based modulation of TSPAN8-LEL can significantly reduce the incidence of EOC metastasis without severe toxicity in vivo. Finally, we demonstrated that the TSPAN8-blocking antibody promotes the internalization and concomitant downregulation of cell surface TSPAN8. Collectively, our data suggest TSPAN8 as a potential novel therapeutic target in EOCs and antibody targeting of TSPAN8 as an effective strategy for inhibiting invasion and metastasis of TSPAN8-expressing EOCs.

Human antibodies targeting the C-type lectin-like domain of the tumor endothelial cell marker clec14a regulate angiogenic properties in vitro.

It has been suggested that clec14a may be involved in tumor angiogenesis. However, a molecular mechanism has not been clearly identified. In this study, we show for the first time that C-type lectin-like domain (CTLD) of clec14a may be important for regulating cell migration and filopodia formation. Using phage display technology, recombinant human antibodies specific to the CTLDs of human and mouse clec14a (clec14a-CTLD (immunoglobulin G) IgG) were selected. Functional assays using the antibodies showed that clec14a-CTLD IgGs specifically blocked endothelial cell migration and tube formation without affecting cell viability or activation. Further, clec14a-CTLD IgGs inhibited clec14a-mediated cell-cell contact by blocking interaction between CTLDs. Finally, clec14a cross-linking by the clec14a-CTLD IgGs significantly downregulated clec14a expression on the surface of endothelial cells. These results strongly suggest that the clec14a-CTLD may be a key domain in angiogenesis, and that clec14a-CTLD IgGs specifically inhibit angiogenesis by modulating CTLD-mediated cell interactions and clec14a expression on the surface of endothelial cells.

Alzheimer's disease amyloid-beta peptide modulates apolipoprotein E isoform specific receptor binding.

The major protein component of the extracellular deposits in Alzheimer's disease (AD) is a 4 kDa peptide termed amyloid-beta (Abeta). This peptide is known to bind apolipoprotein E (apoE), a key mediator of lipoprotein transport, in an isoform specific manner. Whilst these isoform specific effects on apoE are well recognized, the functional significance of this interaction is poorly understood. Here, we investigated the influence of Abeta on apoE-mediated lipoprotein binding to cells using fluorescently tagged lipoprotein-like emulsions. Using this approach, we demonstrate that Abeta enhanced the normally poor binding of apoE2 lipoprotein-like particles to fibroblasts in culture, whilst markedly reducing the binding of apoE3 and apoE4. This suggests that the action of apoE isoforms on cellular lipoprotein or cholesterol metabolism is differentially modulated by Abeta. This also suggests that Abeta may also compromise apoE function in the Alzheimer disease affected brain.

Hormone interactions to Leu-rich repeats in the gonadotropin receptors. III. Photoaffinity labeling of human chorionic gonadotropin with receptor Leu-rich repeat 4 peptide.

Human chorionic gonadotropin (hCG) binds to the extracellular N-terminal domain, exodomain, of its receptor, and the resulting hCG-exodomain complex is thought to modulate the membrane associated domain, endodomain, of the receptor to generate hormone signal. The bulk of the exodomain is speculated to assume a crescent structure consisting of eight to nine Leu-rich repeats (LRRs), which may provide the hormone contact sites. Unfortunately, little experimental evidence is available for the precise hormone contact points in the exodomain and the endodomain. The two preceding articles (Song, Y., Ji, I., Beauchamp, J., Isaacs, N., and Ji, T. (2001) J. Biol. Chem. 276, 3426-3435; Song, Y., Ji, I., Beauchamp, J., Isaacs, N., and Ji, T. (2001) J. Biol. Chem. 276, 3436-3442) show that putative LRR2 and LRR4 are crucial for hormone binding. In particular, the N-terminal region of LRR4 assumes the hydrophobic core of the LRR4 loop, whereas the C-terminal region is crucial for signal generation. However, it is unclear whether LRR4 interacts hCG and the endodomain and how it might be involved in signal generation. In this article, our affinity labeling results present the first evidence that the N-terminal region of LRR4 interacts with hCG, preferentially the hCGalpha subunit and that the hCG/LRR4 complex interacts with exoloop 2 of the endodomain. This interaction offers a mechanism to generate hormone signal.