Inhibition of cell movement and proliferation by cell-cell contact-induced interaction of Necl-5 with nectin-3.

Immunoglobulin-like Necl-5/Tage4/poliovirus receptor (PVR)/CD155, originally identified as the PVR, has been shown to be up-regulated in cancer cells and to enhance growth factor-induced cell movement and proliferation. In addition, Necl-5 heterophilically trans-interacts with nectin-3, a cell-cell adhesion molecule known to form adherens junctions in cooperation with cadherin. We show here that Necl-5 was down-regulated from cell surface upon cell-cell contacts in NIH3T3 cells. This down-regulation of Necl-5 was initiated by its interaction with nectin-3 and was mainly mediated by clathrin-dependent endocytosis. Then, the down-regulation of Necl-5 induced in this way reduced movement and proliferation of NIH3T3 cells. These results indicate that the down-regulation of Necl-5 induced by its interaction with nectin-3 upon cell-cell contacts may be at least one mechanism underlying contact inhibition of cell movement and proliferation.

Prognostic significance of beta-catenin nuclear expression in hepatocellular carcinoma.

BACKGROUND/AIMS: There have been a few clinical reports about beta-catenin nuclear expression for hepatocellular carcinoma patients. However, the clinical significance of the nuclear expression has not been fully elucidated and remains to be controversial. METHODOLOGY: We performed immunohistochemical examinations using an anti-beta-catenin monoclonal antibody for 101 hepatocellular carcinoma patients who underwent curative hepatic resection. Background factors and the disease-free survival were compared between hepatocellular carcinomas with and without beta-catenin nuclear expression. The prognostic factors influencing disease-free survival were examined by univariate and multivariate analyses. RESULTS: Beta-catenin nuclear expression was judged as positive in 24 (23.8%) and negative in 77 (76.2%). Representative 3 hepatocellular carcinomas with beta-catenin nuclear expression showed the protein accumulation by Western blotting analysis. The clinicopathological analysis proved that vascular invasion was less prevalent in hepatocellular carcinomas with beta-catenin nuclear expression than in those without the expression (p=0.034). Hepatocellular carcinoma patients with beta-catenin nuclear expression had a longer disease-free survival than patients without the expression (p=0.035). Multivariate analysis proved that beta-catenin nuclear expression was selected as one of the independent factors related to disease-free survival (p=0.0054). CONCLUSIONS: Our study demonstrated that beta-catenin nuclear expression is valuable as a prognostic factor for hepatocellular carcinoma patients who underwent hepatic resection.

[A case of multiple liver metastases from breast cancer in which we confirmed disappearance of cancer cells after hepatic resection following hepatic arterial infusion chemotherapy].

A 46-year-old woman underwent right mastectomy for stage IIA breast cancer in 1993. Six years and 8 months after this operation, she was diagnosed as having multiple liver metastasis form the breast cancer. An intra-arterial catheter was inserted percutaneously into the hepatic artery and she was given hepatic arterial infusion chemotherapy and general chemoendocrine therapy. The metastatic liver tumors were gradually reduced in size and tumor markers returned to the normal range. However, tumor size was unchanged after February 2001. After obtaining her informed consent from the patient, we performed hepatic resection in December 2001. Histopathologically, three scars were observed and no tumor cell was found. She has been free from recurrence to date. This case suggests that chemoendocrine therapy including hepatic arterial infusion chemotherapy is effective for breast cancer patients with liver metastasis.

[A case of advanced pancreatic cancer with multiple liver metastasis that improved remarkably with use of low-dose cisplatin and TS-1].

The prognosis and QOL of unresectable pancreatic cancer are very poor. A symptomless 60-year-old male was admitted for examination of a high serum CA19-9 level. Following ultrasound and abdominal CT, we diagnosed unresectable advanced pancreatic cancer with multiple liver metastasis. After we obtained his informed consent, we administered continuous infusion of 5-FU and low-dose cisplatin (CDDP) infusion (low-dose FP therapy) for 3 weeks. He then underwent combination chemotherapy with low-dose CDDP and TS-1 on an outpatient basis. During the chemotherapy, he did not experience any major adverse event and his QOL was relatively good. On follow-up CT 3 months later, the primary tumor in the pancreas was found to be stable. However, the size and number of liver tumors were remarkably reduced. The serum CA19-9 level had also remarkably decreased from 48,300 U/ml to 1,480 U/ml. In conclusion, the combination chemotherapy using low-dose CDDP and TS-1 can be effective in cases of unresectable pancreatic cancer with multiple liver metastasis.

Involvement of up-regulated Necl-5/Tage4/PVR/CD155 in the loss of contact inhibition in transformed NIH3T3 cells.

Normal cells show contact inhibition of cell movement and proliferation, but this is lost following transformation. We found that Necl-5, originally identified as a poliovirus receptor and up-regulated in many cancer cells, enhances growth factor-induced cell movement and proliferation. We showed that when cells contact other cells, Necl-5 interacts in trans with nectin-3 and is removed by endocytosis from the cell surface, resulting in a reduction of cell movement and proliferation. We show here that up-regulation of the gene encoding Necl-5 by the oncogene V12-Ki-Ras causes enhanced cell movement and proliferation. Upon cell-cell contact, de novo synthesis of Necl-5 exceeds the rate of Necl-5 endocytosis, eventually resulting in a net increase in the amount of Necl-5 at the cell surface. In addition, expression of the gene encoding nectin-3 is markedly reduced in transformed cells. Thus, up-regulation of Necl-5 following transformation contributes to the loss of contact inhibition in transformed cells.

Necl-5/poliovirus receptor interacts in cis with integrin alphaVbeta3 and regulates its clustering and focal complex formation.

Integrin alphavbeta3, which forms focal complexes at leading edges in moving cells, is up-regulated in cancer cells and so is implicated in their invasiveness. Necl-5, originally identified as a poliovirus receptor and also up-regulated in cancer cells, colocalizes with integrin alphavbeta3 at leading edges in moving cells and enhances growth factor-induced cell movement. Here, we show that Necl-5 interacts directly, in cis, with integrin alphavbeta3, and enhances integrin alphavbeta3 clustering and focal complex formation at leading edges in NIH3T3 cells. The extracellular region of Necl-5, but not the cytoplasmic region, is necessary for its interaction with integrin alphavbeta3; however, both regions are necessary for its action. An interaction between integrin alphavbeta3 and vitronectin and PDGF-induced activation of Rac are also necessary for integrin alphavbeta3 clustering. The interaction between Necl-5 and integrin alphavbeta3 enhances PDGF-induced Rac activation, facilitating integrin alphavbeta3 clustering presumably in a feedback amplification manner. Thus, Necl-5 has a critical role in integrin alphavbeta3 clustering and focal complex formation.

Direct binding of cell polarity protein PAR-3 to cell-cell adhesion molecule nectin at neuroepithelial cells of developing mouse.

PAR-3 is a cell polarity protein that localizes at tight junctions (TJs) by direct binding to an immunoglobulin (Ig)-like cell-cell adhesion molecule JAM-1 in mammalian epithelial cells. Another Ig-like cell-cell adhesion molecule nectin plays a role in the localization of JAM-1 at TJs in epithelial cells. Nectin furthermore plays a role in the organization of adherens junctions (AJs) and TJs. Nectin comprises a family of four members, nectin-1, -2, -3, and -4. Nectins are associated with the actin cytoskeleton through afadin, of which the PDZ domain binds to nectins through their C-terminal four amino acids. We show here that PAR-3 binds to nectin-1 and -3 in neuroepithelial cells of the embryonic telencephalon, which are equipped with AJs, but not with typical TJs. Nectin-1, -2, -3, and afadin, but not JAM-1, were concentrated at AJs in neuroepithelial cells of the embryonic telencephalon at E13.5 and PAR-3 co-localized with nectins. PAR-3 was co-immunoprecipitated with nectin-1 and -3, but not with nectin-2 or JAM-1, from the mouse whole brain at E13.5. Recombinant PAR-3 stoichiometrically bound to recombinant nectin-1 and -3. The first one of the three PDZ domains of PAR-3 bound to the C-terminal four amino acids of nectin-1 and -3. The affinities of PAR-3 and afadin for nectin-1 and -3 were similar. Cadherin-deficient L cells expressing nectin-1 and -3 formed nectin-1- and -3-based cell-cell junctions, respectively, where PAR-3 as well as afadin was recruited. These results indicate that nectin-1 and -3 are involved in the localization of PAR-3 at AJs in the neuroepithelial cells of the embryonic telencephalon.

Enhancement of serum- and platelet-derived growth factor-induced cell proliferation by Necl-5/Tage4/poliovirus receptor/CD155 through the Ras-Raf-MEK-ERK signaling.

Necl-5/Tage4/poliovirus receptor/CD155 has been shown to be the poliovirus receptor and to be up-regulated in rodent and human carcinoma. We have found previously that mouse Necl-5 regulates cell motility. We show here that mouse Necl-5 is furthermore involved in the regulation of cell proliferation. Studies using a specific antibody against Necl-5 and a dominant negative mutant of Necl-5 revealed that Necl-5 enhanced the serum-induced proliferation of NIH3T3, Swiss3T3, and mouse embryonic fibroblast cells. Necl-5 enhanced the serum-induced activation of the Ras-Raf-MEK-ERK signaling, up-regulated cyclins D2 and E, and down-regulated p27(Kip1), eventually shortening the period of the G(0)/G(1) phase of the cell cycle in NIH3T3 cells. Necl-5 similarly enhanced the platelet-derived growth factor-induced activation of the Ras-Raf-MEK-ERK signaling and shortened the period of the G(0)/G(1) phase of the cell cycle in NIH3T3 cells. Necl-5 acted downstream of the platelet-derived growth factor receptor and upstream of Ras. Moreover, up-regulated Necl-5 was involved at least partly in the enhanced proliferation of transformed cells including NIH3T3 cells transformed by an oncogenic Ras or v-Src. These results indicate that Necl-5 plays roles not only in cell motility but also in cell proliferation.