B7-H3, an immunoregulatory protein, is known to play a role in tumor progression. In many cancer types, observed correlations between high B7-H3 expression and poor prognosis have been attributed to involvement in antitumor immunity.However, here we demonstrate a nonimmunological alternative function of B7-H3 in cancer metastasis. Since advanced malignant melanoma is a disease with a poor survival rate and a broad pattern of metastasis, we used this disease as a model in our studies. We found that shRNA silencing of B7-H3 reduced the in vitro migratory potential and matrigel invasiveness of MDA-MB-435 and FEMX-I melanoma cells. In an experimental metastasis model in vivo, B7-H3 silencing of MDA-MB-435 cells resulted in reduced metastatic capacity and significantly increased the median symptom-free survival of nude mice (147 vs. 65 days, p < 0.001) and rats (53 vs. 42 days, p 5 0.025) injected with MDA-MB-435 cells. Furthermore, a smaller fraction of mice had microscopically detectable metastases compared to control animals, and the pattern of metastases was slightly different between the two groups but with the brain as the predominant organ. Immunohistochemistry on samples from two melanoma patients showed strong B7-H3 staining in both a primary tumor and metastases. Notably, the metastasis-associated proteins, matrix metalloproteinase (MMP)-2, signal transducer and activator of transcription 3 (Stat3), and the level of secreted interleukin-8 (IL-8) were reduced in the B7-H3 knock-down cell variants, whereas tissue inhibitor of metalloproteinase (TIMP)-1 and-2 levels were increased. Taken together, our findings indicate a novel role for B7-H3 in the regulation of the metastatic capacity of melanoma cells and it might be a potential therapeutic target for anti-metastasis therapy.B7-H3 (CD276) is a type I transmembrane protein with immunoglobulin-like structure. Two different isoforms are described having either two or four immunoglobulin domains, the latter being dominant in humans. 1,2 B7-H3 is a member of the B7/CD28 immunoglobulin superfamily which consists of several ligands that have inhibitory and stimulatory effects on the regulation of immune responses to transformed cells, thus being involved in tumor immunity, 3 and the protein is known to both activate and inhibit T-cell responses. 4,5 This is reflected in the conflicting reports on its role in cancer describing both beneficial and adverse effects. Thus, several immunohistochemical studies have demonstrated a correlation between high expression of the protein and tumor progression, 6-11 whereas other studies suggest a positive effect on the clinical outcome. 12,13 Although the effects of B7-H3 in cancer have been attributed to its involvement in antitumor immunity, its exact function remains unclear.In contrast to previous reports, we have investigated the B7-H3 protein in nonimmunological systems and conclude that it also plays a direct role in cancer progression. In vitro studies on cancer cells showed that siRNA down-regulation of B7-H3 reduced cel...

Background:B7-H3, an immunoregulatory protein, is overexpressed in several cancers and is often associated with metastasis and poor prognosis. Here, our aim was to identify microRNAs (miRNAs) regulating B7-H3 and assess their potential prognostic implications in breast cancer.Methods:MicroRNAs targeting B7-H3 were identified by transfecting two breast cancer cell lines with a library of 810 miRNA mimics and quantifying changes of B7-H3 protein levels using protein lysate microarrays. For validations we used western immunoblotting and 3′-UTR luciferase assays. Clinical significance of the miRNAs was assayed by analysing whether their expression levels correlated with outcome in two cohorts of breast cancer patients (142 and 81 patients).Results:We identified nearly 50 miRNAs that downregulated B7-H3 protein levels. Western immunoblotting validated the impact of the 20 most effective miRNAs. Thirteen miRNAs (miR-214, miR-363*, miR-326, miR-940, miR-29c, miR-665, miR-34b*, miR-708, miR-601, miR-124a, miR-380-5p, miR-885-3p, and miR-593) targeted B7-H3 directly by binding to its 3′-UTR region. Finally, high expression of miR-29c was associated with a significant reduced risk of dying from breast cancer in both cohorts.Conclusions:We identified miRNAs efficiently downregulating B7-H3 expression. The expression of miR-29c correlated with survival in breast cancer patients, suggesting a tumour suppressive role for this miRNA.

SummaryAcute lymphoblastic leukaemia (ALL) is the most common malignancy in children. Recently, there has been a growing interest in Wnt signalling in several aspects of cellular development, including cancer formation. Little is known about Wnt signalling in B-ALL. We investigated whether activation of canonical Wnt signalling could occur in B-ALL cells and thereby play a potential role in cellular growth and/or survival. This study found that Wnt3A induced b-catenin accumulation in both primary B-ALL cells and B-ALL leukaemia cell lines. Further, Wnt3A was shown to induce nuclear translocation of b-catenin and TCF/Lef-1 dependent transcriptions in the B-ALL cell line Nalm-6. Examination of the mRNA expression pattern of WNT ligands, FZD receptors and WNT antagonists in Nalm-6 cells identified a set of ligands and receptors available for signalling, as well as antagonists potentially available for modulating the response. Functional analyses showed that Wnt3A inhibited the proliferation of several, but not all, B-ALL cell lines studied. Finally, microarray analysis was used to identify several Wnt3A target genes involved in a diverse range of cellular activities, which are potential mediators of the Wnt3A-restrained proliferation.

B7-H3 is a transmembrane glycoprotein and a member of the B7 family of proteins. It was previously known as an immunoregulatory molecule, shown in recent years to be of clinical significance in different types of cancer. In some tumor types high expression of B7-H3 has been linked to a poor prognosis, whereas in other cancers the opposite effect has been observed. Taken together, the precise role of B7-H3 in tumor immunity is unclear and further investigations are needed. Another aspect of B7-H3 that so far has received little interest is its role in non-immunological systems. We have demonstrated that knockdown of B7-H3 in melanoma and breast cancer cells results in both increased chemosensitivity and decreased metastatic potential. This has been observed in both in vitro and in vivo experiments. Several different signaling pathways seems to be involved, as B7-H3 knockdown can be linked to both higher expression of apoptotic markers and increased phosphorylation of Stat3. Increased knowledge of also the non-immunological role of B7-H3 protein is therefore of great biological and putative therapeutic interest.

No abstract