Metastatic Capacity Of Cancer Cells Research Articles

Multifractal analysis of cellular ATR-FTIR spectrum as a method for identifying and quantifying cancer cell metastatic levels

Cancer is a leading cause of mortality today. Sooner a cancer is detected, the more effective is the treatment. Histopathological diagnosis continues to be the gold standard worldwide for cancer diagnosis, but the methods used are invasive, time-consuming, insensitive, and still rely to some degree on the subjective judgment of pathologists. Recent research demonstrated that Attenuated Total Reflection-Fourier Transform Infrared (ATR-FTIR) spectroscopy can be used to determine the metastatic potential of cancer cells by evaluating their membrane hydration. In the current study, we demonstrate that the conversion of ATR-FTIR spectra using multifractal transformation generates a unique number for each cell line’s metastatic potential. Applying this technique to murine and human cancer cells revealed a correlation between the metastatic capacity of cancer cells within the same lineage and higher multifractal value. The multifractal spectrum value was found to be independent of the cell concentration used in the assay and unique to the tested lineage. Healthy cells exhibited a smaller multifractal spectrum value than cancer cells. Further, the technique demonstrated the ability to detect cancer progression by being sensitive to the proportional change between healthy and cancerous cells in the sample. This enables precise determination of cancer metastasis and disease progression independent of cell concentration by comparing the measured spectroscopy derived multifractal spectrum value. This quick and simple technique devoid of observer bias can transform cancer diagnosis to a great extent improving public health prognosis worldwide.

DDR1 promotes migration and invasion of breast cancer by modulating the Src-FAK signaling.

Breast cancer is the most commonly diagnosed cancer among women, causing 15% of patient deaths. The metastasis of breast cancer cells is the leading cause of death for patients. Several studies have shown that Discoidin Domain Receptor 1 (DDR1) was highly expressed in breast cancer and could influence tumor cell behaviors. However, the specific role of DDR1 in breast cancer metastasis is still elusive. In this study, we uncovered that DDR1 is significantly increased in breast cancer and inversely correlated with the prognosis of patients. Knockdown of DDR1 suppressed the migration and invasion of breast cancer cells. Additionally, overexpression of DDR1 enhanced the metastatic capacity of cancer cells. Immunoblotting revealed that activation of Src and FAK, which are involved in cancer cell metastasis, were correlated with the expression level of DDR1. Co-immunoprecipitation experiments showed that DDR1 could bind to Src and FAK. Finally, the inhibition of FAK and Src could attenuate DDR1 enhanced migration ability of breast cancer cells. In summary, our study revealed that DDR1 was highly expressed in breast cancer and negatively correlated with the prognosis of breast cancer patients. DDR1 facilitates migration and invasion in breast cancer cells via activation of the Src-FAK signaling. Accordingly, blocking DDR1/Src/FAK axis is a promising therapeutic strategy for breast cancer treatment.

Metabolic Compartmentalization at the Leading Edge of Metastatic Cancer Cells.

Despite advances in targeted therapeutics and understanding in molecular mechanisms, metastasis remains a substantial obstacle for cancer treatment. Acquired genetic mutations and transcriptional changes can promote the spread of primary tumor cells to distant tissues. Additionally, recent studies have uncovered that metabolic reprogramming of cancer cells is tightly associated with cancer metastasis. However, whether intracellular metabolism is spatially and temporally regulated for cancer cell migration and invasion is understudied. In this review, we highlight the emergence of a concept, termed “membraneless metabolic compartmentalization,” as one of the critical mechanisms that determines the metastatic capacity of cancer cells. In particular, we focus on the compartmentalization of purine nucleotide metabolism (e.g., ATP and GTP) at the leading edge of migrating cancer cells through the uniquely phase-separated microdomains where dynamic exchange of nucleotide metabolic enzymes takes place. We will discuss how future insights may usher in a novel class of therapeutics specifically targeting the metabolic compartmentalization that drives tumor metastasis.

Exploitation of the chick embryo chorioallantoic membrane (CAM) as a platform for anti-metastatic drug testing

The establishment of clinically relevant models for tumor metastasis and drug testing is a major challenge in cancer research. Here we report a physiologically relevant assay enabling quantitative analysis of metastatic capacity of tumor cells following implantation into the chorioallantoic membrane (CAM). Engraftment of as few as 103 non-small cell lung cancer (NSCLC) and prostate cancer (PCa) cell lines was sufficient for both primary tumor and metastasis formation. Standard 2D-imaging as well as 3D optical tomography imaging were used for the detection of fluorescent metastatic foci in the chick embryo. H2228- and H1975-initiated metastases were confirmed by genomic analysis. We quantified the inhibitory effect of docetaxel on LNCaP, and that of cisplatin on A549- and H1299-initiated metastatic growths.The CAM assay also mimicked the sensitivity of ALK-rearranged H2228 and EGFR-mutated H1975 NSCLC cells to tyrosine kinase inhibitors crizotinib and gefitinib respectively, as well as sensitivity of LNCaP cells to androgen-dependent enzalutamide therapy. The assay was suggested to reconstitute the bone metastatic tropism of PCa cells. We show that the CAM chick embryo model may be a powerful preclinical platform for testing and targeting of the metastatic capacity of cancer cells.

Mechanophenotyping of B16 Melanoma Cell Variants for the Assessment of the Efficacy of (-)-Epigallocatechin Gallate Treatment Using a Tapered Microfluidic Device.

Metastatic cancer cells are known to have a smaller cell stiffness than healthy cells because the small stiffness is beneficial for passing through the extracellular matrix when the cancer cells instigate a metastatic process. Here we developed a simple and handy microfluidic system to assess metastatic capacity of the cancer cells from a mechanical point of view. A tapered microchannel was devised through which a cell was compressed while passing. Two metastasis B16 melanoma variants (B16-F1 and B16-F10) were examined. The shape recovery process of the cell from a compressed state was evaluated with the Kelvin–Voigt model. The results demonstrated that the B16-F10 cells showed a larger time constant of shape recovery than B16-F1 cells, although no significant difference in the initial strain was observed between B16-F1 cells and B16-F10 cells. We further investigated effects of catechin on the cell deformability and found that the deformability of B16-F10 cells was significantly decreased and became equivalent to that of untreated B16-F1 cells. These results addressed the utility of the present system to handily but roughly assess the metastatic capacity of cancer cells and to investigate drug efficacy on the metastatic capacity.

Intravital imaging reveals systemic ezrin inhibition impedes cancer cell migration and lymph node metastasis in breast cancer

BackgroundLimited understanding of the cancer biology of metastatic sites is a major factor contributing to poor outcomes in cancer patients. The regional lymph nodes are the most common site of metastasis in most solid cancers and their involvement is a strong predictor of relapse in breast cancer (BC). We have previously shown that ezrin, a cytoskeletal–membrane linker protein, is associated with lymphovascular invasion and promotes metastatic progression in BC. However, the efficacy of pharmacological inhibition of ezrin in blocking cancer cell migration and metastasis remains unexplored in BC.MethodsWe quantified ezrin expression in a BC tissue microarray (n = 347) to assess its correlation with risk of relapse. Next, we developed a quantitative intravital microscopy (qIVM) approach, using a syngeneic lymphatic reporter mouse tumor model, to investigate the effect of systemic ezrin inhibition on cancer cell migration and metastasis.ResultsWe show that ezrin is expressed at significantly higher levels in lymph node metastases compared to matched primary tumors, and that a high tumor ezrin level is associated with increased risk of relapse in BC patients with regional disease. Using qIVM, we observe a subset of cancer cells that retain their invasive and migratory phenotype at the tumor-draining lymph node. We further show that systemic inhibition of ezrin, using a small molecule compound (NSC668394), impedes the migration of cancer cells in vivo. Furthermore, systemic ezrin inhibition leads to reductions in metastatic burden at the distal axillary lymph node and lungs.ConclusionsOur findings demonstrate that the tumor ezrin level act as an independent biomarker in predicting relapse and provide a rationale for therapeutic targeting of ezrin to reduce the metastatic capacity of cancer cells in high-risk BC patients with elevated ezrin expression.

Osteopontin production by TM4SF4 signaling drives a positive feedback autocrine loop with the STAT3 pathway to maintain cancer stem cell-like properties in lung cancer cells.

Transmembrane 4 L6 family proteins have been known to promote cancer. In this study, we demonstrated that transmembrane 4 L6 family member 4 (TM4SF4), which is induced by γ-radiation in non-small cell lung cancer (NSCLC) cells, is involved in epithelial-to-mesenchymal transition (EMT) and cancer stem cell (CSC) properties of NSCLC through the regulation of osteopontin (OPN). Forced TM4SF4 overexpression in A549 cells increased the secretion of OPN, which activates CD44 or integrin signaling and thus maintains EMT-associated CSC-like properties. OPN, known as a downstream target of β-catenin/T-cell factor 4 (TCF-4), was induced by up-regulated β-catenin via TM4SF4-driven phosphorylation of glycogen synthase kinase 3b (GSK3β). TCF4 complexed to promoter regions of OPN in TM4SF4-overexpressing A549 cells was also confirmed by chromatin immunoprecipitation. Knockout of either β-catenin or TCF4-suppressed OPN expression, demonstrating that both factors are essential for OPN expression in NSCLC cells. OPN secreted by TM4SF4/GSK3β/β-catenin signaling activated the JAK2/STAT3 or FAK/STAT3 pathway, which also up-regulates OPN expression in an autocrine manner and consequently maintains the self-renewal and metastatic capacity of cancer cells. Neutralizing antibody to OPN blocked the autocrine activation of OPN expression, consequently weakened the metastatic and self-renewal capacity of cancer cells. Collectively, our findings indicate that TM4SF4-triggered OPN expression is involved in the persistent reinforcement of EMT or cancer stemness by creating a positive feedback autocrine loop with JAK2/STAT3 or FAK/STAT3 pathways.

Arsenic trioxide inhibits CXCR4-mediated metastasis by interfering miR-520h/PP2A/NF-κB signaling in cervical cancer.

Arsenic apparently affects numerous intracellular signal transduction pathways and causes many alterations leading to apoptosis and differentiation in malignant cells. We and others have demonstrated that arsenic inhibits the metastatic capacity of cancer cells. Here we present additional mechanistic studies to elucidate the potential of arsenic as a promising therapeutic inhibitor of metastasis. The effects of arsenic trioxide (ATO) on human cervical cancer cell lines migration and invasion were observed by transwell assays. In experimental metastasis assays, cancer cells were injected into tail veins of severe combined immunodeficient mice for modeling metastasis. The mechanisms involved in ATO regulation of CXCR4 were analyzed by immunoblot, real-time polymerase chain reaction, and luciferase reporter assays. Immunohistochemistry was utilized to identify PP2A/C and CXCR4 protein expressions in human cervical cancer tissues. ATO inhibited CXCR4-mediated cervical cancer cell invasion in vitro and distant metastasis in vivo. We determined that ATO modulates the pivotal nuclear factor-kappa B (NF-κB)/CXCR4 signaling pathway that contributes to cancer metastasis. Substantiating our findings, we demonstrated that ATO activates PP2A/C activity by downregulating miR-520h, which results in IKK inactivation, IκB-dephosphorylation, NF-κB inactivation, and, subsequently, a reduction in CXCR4 expression. Furthermore, PP2A/C was reduced during cervical carcinogenesis, and the loss of PP2A/C expression was closely associated with the nodal status of cervical cancer patients. Our results indicate a functional link between ATO-mediated PP2A/C regulation, CXCR4 expression, and tumor-suppressing ability. This information will be critical in realizing the potential for synergy between ATO and other anti-cancer agents, thus providing enhanced benefit in cancer therapy.

Abstract 3959: MMP13 contributes to metastatic tumor growth in the steatotic liver microenvironment.

Abstract The expanding obesity epidemic has led to an escalation in the incidence of non-alcoholic fatty liver disease (NAFLD) and steatohepatitis, which are associated with an increase in incidence of primary and metastatic liver cancer. As NAFLD progresses, it is characterized by marked inflammation in the liver described as non-alcoholic steatohepatitis (NASH). The mechanisms by which NASH increases risk of liver cancer are not fully known but complete understanding of these mediators is necessary for effective therapeutic and preventive strategies. Importantly, metastasis represents the final stage in cancer progression and is the most significant cause of cancer related mortality. Using our mouse model of diet induced steatosis, coupled with a model of experimental liver metastasis, we have previously shown a significant increase in the number of metastatic foci in the steatotic liver compared to the normal livers. Microarray analysis revealed that MMP13, a member of the Matrix Metalloproteinase (MMP) family recognized to be involved in liver disease and cancer progression, is significantly upregulated in the steatotic liver compared to normal livers. Additionally, MMP13 is expressed in several cancer cell types. Emerging data suggests that MMP13 may play a key role in priming the microenvironment for the establishment of liver metastases, by modulation of cytokine profiles and recruitment of inflammatory cells. . We hypothesize that MMP13 is a key modulator of the steatotic microenvironment, and that increased MMP13 levels favor primary tumor growth as well as the establishment of metastases. To determine the role of stromal MMP13 on metastasis to the liver, we are using the experimental splenic injection model of liver metastasis with syngeneic MC38 colon cancer cells in mice genetically deficient in MMP13. Additionally, since the tumor cells themselves express MMP13, wildtype mice are injected with either shRNA control or MMP13 knockdown MC38 colon cancer cells to determine the role of tumor cell derived MMP13. Preliminary data shows localization of MMP13 expression to the hepatic stellate cells as well as CD11b positive myeloid cells from livers with NASH. Further, invitro analysis of migration and invasion ability of parental and MMP13 shRNA knockdown MC38 cells show that MMP13 promotes the invasive and migratory properties of the metastatic cancer cells themselves. MMP13 may thus be a potential mediator of inflammation in the tumor microenvironment as well as of the metastatic capacity of cancer cells to the liver. Citation Format: Alisha M. Mendonsa, Michael VanSaun, David L. Gorden. MMP13 contributes to metastatic tumor growth in the steatotic liver microenvironment. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 3959. doi:10.1158/1538-7445.AM2013-3959

Ezrin mRNA target site selection for DNAzymes using secondary structure and hybridization thermodynamics

Ezrin, a membrane organizer and linker between plasma membrane and cytoskeleton, is well documented to play an important role in the metastatic capacity of cancer cells especially for osteosarcoma cells. It has provided an ideal target for cancer gene therapy. RNA-cleaving 10-23 DNAzymes, consisting of a 15-nucleotide catalytical domain flanked by two target-specific complementary arms, can cleave the target mRNA at purine-pyrimidine dinucleotide effectively. In the present study, we designed and screened the target sites for 10-23 DNAzymes against ezrin mRNA by using multiple computational methods with combination of secondary structural and hybridization thermodynamic parameters. Then, we testified the activities of the DNAzymes directed against these selected target sites in vitro. Our results show that AU1751 is the most effective target site of ezrin mRNA for DNAzymes because of its ideal secondary structure and hybridization thermodynamics. So, there is a significant correlation between the multiple computational methods and the efficacy of the corresponding DNAzymes. These provide a rational, efficient way for DNAzymes selection.

Expression change of ezrin as a prognostic factor in primary osteosarcoma

Osteosarcoma is a life-threatening malignancy that most often occurs in teenagers. Ezrin is a cytoskeleton linker protein that is actively involved in the metastatic capacity of cancer cells. In this study, we investigated the expression change of ezrin after preoperative chemotherapy and its prognostic value in patients with primary osteosarcoma. Ezrin mRNA expression level of initial biopsy specimens and resected tumor specimens after preoperative chemotherapy of 25 patients who had primary osteosarcoma with ezrin expression was detected by semiquantitative reverse transcription polymerase chain reaction (RT-PCR) to evaluate the expression change of ezrin. In addition, the prognostic factors and the relationship between the expression change of ezrin and the clinical characteristics were analyzed. Expression change of ezrin was found in 64% of all 25 patients. The expression change of ezrin had good relation with histology (P = 0.037), grade (P = 0.006), chemotherapy response (P = 0.017), and metastasis or recurrence (P = 0.041). The 2-year overall survival and event-free survival were associated with expression change of ezrin (P = 0.001 and P = 0.002, respectively) and response to preoperative chemotherapy (P = 0.001 and P = 0.012, respectively). Our findings suggest that expression change of ezrin is an independent positive prognostic factor in non-metastatic osteosarcoma.

Abstract 3310: Ezrin is an independent prognostic factor in extremity soft tissue sarcomas

Abstract Ezrin is a cytoskeleton protein actively involved in regulating the growth and metastatic capacity of cancer cells. It has recently been reported to be to be associated with poor prognosis high grade soft tissue sarcoma (STS) and to mediate growth and survival through AKT pathway. To further validate the prognostic value of ezrin in STS, we evaluated 229 extremity primary STS using tissue microarrays and immunohistochemistry. Ezrin expression was correlated with metastasis during follow-up for at least 5 years or until death.185 of the 229 cases analyzed (80%) showed ezrin immunoreactivity in the membrane and cytoplasm of the tumor cells. Multivariate analysis with other known risk factors, including size, vascular invasion and necrosis showed that ezrin expression was significantly associated with development of distant metastasis during follow-up (P = 0.01, 95%CI 1.4-3.4). Ezrin expression was also correlated with AKT pathway changes, evaluated by gene expression. Our findings suggest that ezrin is a promissing prognostic marker in highly malignant STS and support an interaction between ezrin and AKT in the genesis of STS metastasis. Note: This abstract was not presented at the AACR 101st Annual Meeting 2010 because the presenter was unable to attend. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3310.

Evaluation of potential interactions between the metastasis-associated protein S100A4 and the tumor suppressor protein p53

Metastasis is a complex cascade of events involving a finely tuned interplay between malignant cells and multiple host factors. The transition from benign tumor growth to malignancy is manifested by the ability of tumor cells to traverse tissue barriers and invade surrounding tissues. Among a multitude of factors playing a role, the small calcium-binding protein S100A4 has been found to add to the invasive and metastatic capacity of cancer cells. However, the exact molecular function or mechanism by which S100A4 exerts its putative metastasis-promoting effects has not been fully elucidated, and the protein is most likely involved in several aspects of tumor progression. Several studies have recently described a direct interaction and/or reciprocal influence between S100A4 and the tumor suppressor protein p53. This corresponds to reports linking p53 to other S100-family members, especially S100B. The consequences are intriguing, connecting the metastasis-promoting protein S100A4 to the large set of important p53-mediated functions, with broad potential importance in cancer development and metastasis. In this review we emphasize the studies involving p53 and S100A4, elucidating and comparing reported results and conclusions.

Differential expression of ezrin in epithelial skin tumors: cytoplasmic ezrin immunoreactivity in squamous cell carcinoma

Ezrin is a cytoskeleton linker protein that is actively involved in regulating the growth and metastatic capacity of cancer cells. The purpose of the study was to assess the expression pattern of ezrin in normal skin and various epithelial neoplasms. We used immunohistochemical techniques to examine the expression of ezrin in paraffin-embedded tissues of squamous cell carcinoma (n = 23), basal cell carcinoma (n = 10), Bowen's disease (n = 10), actinic keratosis (n = 10), keratoacanthoma (n = 9), seborrheic keratosis (n = 5), psoriasis vulgaris (n = 5), and normal control skin (n = 5). In Bowen's disease, actinic keratosis, keratoacanthoma, and seborrheic keratosis, ezrin was dominantly expressed in the cell membrane except for the cornified layer. In squamous cell carcinoma (SCC) specimens, the percentage of ezrin-positive cells was increased compared with Bowen's disease, actinic keratosis, keratoacanthoma, and seborrheic keratosis. Especially in SCC samples, ezrin expression was markedly expressed in the cytoplasm. In addition, there was a correlation between the pattern of ezrin expression and tumor differentiation in SCC. Basal cell carcinoma showed intense and diffuse staining especially in the solid growth pattern. Our findings suggest that dysregulation of ezrin may be important in the development of cutaneous epithelial malignancies and tumor grade. We suggest that the cytoplasmic localization of ezrin may be useful in the diagnosis of skin SCC.

Experimental melanoma metastasis in lungs of mice with congenital coagulation disorders

Experimental animal studies as well as clinical trials have shown that interventions targeting the blood coagulation cascade inhibit cancer cell metastasis. These data support the hypothesis that congenital prothrombotic disorders, like factor V Leiden, facilitate metastasis whereas bleeding disorders, like haemophilia impede metastasis. To test this hypothesis, we subjected factor V Leiden and factor VIII deficient mice to a murine model of experimental lung metastasis. In this model, B16F10 murine melanoma cells are injected into the tail vein resulting in multiple lung metastases within 20 days. Both hemi- and homozygous factor VIII deficient mice were protected against lung metastasis compared to wild-type littermate controls. In contrast, homozygous factor V Leiden mice developed more metastases than wild-type littermates, whereas heterozygous carriers showed an intermediate number of pulmonary foci. Overall, these data show that a congenital susceptibility to either bleeding or thrombosis modifies the metastatic capacity of cancer cells in the bloodstream and suggest that procoagulant phenotypes are a risk factor for tumour metastasis.

High Level of Ezrin mRNA Expression in an Osteosarcoma Biopsy Sample With Lung Metastasis

Osteosarcoma (OS) remains a life-threatening malignancy and its molecular character is not fully understood. Ezrin is a cytoskeleton linker protein involved in regulating the growth and metastatic capacity of cancer cells. However, the correlation between ezrin mRNA expression and clinical severity has not yet been examined in OS biopsy samples. Furthermore, recent evidence has demonstrated that the level of glyceraldehyde-3-phosphate dehydrogenase (GAPDH) expression is increased in human cancers of various origins, but this has not yet been examined in OS cells. To clarify the correlation between the clinical severity and the levels of ezrin and GAPDH mRNA expression, we quantified these mRNA levels in 4 pediatric OS biopsy samples using real-time quantitative polymerase chain reaction. Among these 4 samples, ezrin mRNA expression was approximately 5-fold higher in a case with lung metastasis compared with the other cases without metastasis, suggesting an association between the ezrin mRNA expression level and metastasis. On the other hand, the GAPDH mRNA expression level was not related to the clinical severity. This is the first report to demonstrate a high level of ezrin mRNA expression in an OS biopsy sample with lung metastasis.

Ezrin in osteosarcoma: Comparison between conventional high-grade and central low-grade osteosarcoma

Ezrin is a cytoskeleton linker protein that is actively involved in the regulation of growth and metastatic capacity of cancer cells. Recently, it has been demonstrated that a significant correlation exists between high ezrin expression levels and the poor outcome of pediatric osteosarcoma patients. The expression of ezrin was compared in conventional high-grade and central low-grade osteosarcoma lesions to investigate the role of ezrin overexpression in the metastasis of osteosarcoma. We compared the expression levels of the ezrin protein in 32 cases of high-grade osteosarcomas and 21 cases of low-grade osteosarcomas using immunohistochemistry. Ezrin protein expression levels were examined in three different human osteosarcoma cell lines by Western blotting. In addition, the mRNA expression levels of ezrin in these osteosarcoma cell lines and control fibroblasts were evaluated by real-time quantitative PCR. Ezrin immunoreactivity was present in 43.7% of high-grade osteosarcoma specimens. All low-grade osteosarcomas were negative for ezrin. The expression of ezrin was detected by Western blotting in all three osteosarcoma cell lines. The tested osteosarcoma cell lines showed marked amplification of ezrin mRNA compared to control cells. Taken together, ezrin appears to play a role in the progression of tumors, such as the metastasis of osteosarcoma. However, further data are needed before ezrin can be considered in clinical decision-making about osteosarcoma patients.

Prognostic Impact of Immunohistochemical Expression of Ezrin in Highly Malignant Soft Tissue Sarcomas

Ezrin is a cytoskeleton linker protein that is actively involved in regulating the growth and metastatic capacity of cancer cells. It has recently been reported to be involved in dissemination of pediatric soft tissue sarcoma (STS). To further evaluate the prognostic value of ezrin in STS progression, we screened 50 primary STSs of high malignancy grade using immunohistochemistry. At the initial surgery, all patients were without local or distant metastasis. The expression was then compared with the outcome during follow-up for at least 4 years or until the patients' death. Twenty-five of the 50 STSs analyzed (50%) showed ezrin immunoreactivity in the membrane and cytoplasm of the tumor cells. A significant association was shown between positive expressions of ezrin and death in disease as well as overall survival (P = 0.014 and 0.007, respectively). Similarly, ezrin expression was significantly associated with development of distant metastasis during follow-up (P = 0.031), also excluding locally recurrent disease (P = 0.049). The relative abundance of metastasis in ezrin-positive cases was observed both over time and irrespective of time. In comparison with clinical, histopathologic, and genetic characteristics of the STSs, ezrin expression was found to correlate significantly with an infiltrative growth pattern outside the tumor capsule as well as with copy number gain of chromosomal region 9cen-q22. Our findings suggest that ezrin immunoreactivity could be valuable as an additional prognostic marker in highly malignant STSs and support a causative role of ezrin in STS tumor dissemination.

Core 3 synthase is down-regulated in colon carcinoma and profoundly suppresses the metastatic potential of carcinoma cells

The core 3 structure of the O-glycan, GlcNAcbeta1-3GalNAcalpha1-Ser/Thr, an important precursor in the biosynthesis of mucin-type glycoproteins, is synthesized by beta1,3-N-acetylglucosaminyltransferase 6 (beta3Gn-T6; core 3 synthase). We generated an anti-beta3Gn-T6 mAb (G8-144 mAb) and performed immunohistochemical analyses. In normal stomach and colon, beta3Gn-T6 was strongly expressed in the Golgi region of epithelia. In contrast, its expression was markedly down-regulated in gastric and colorectal carcinomas. Tissue specimens from a familial adenomatous polyposis patient showed a clear correlation between the down-regulation of beta3Gn-T6 expression and the degree of dysplasia/neoplasia. In vitro, the level of beta3Gn-T6 transcript was increased according to the differentiation of Caco-2 cells. These results suggested that the expression of beta3Gn-T6 is closely regulated during differentiation and dedifferentiation. beta3Gn-T6 would be a useful marker for distinguishing between benign adenomas and premalignant lesions. HT1080 FP-10 cells stably transfected with the beta3Gn-T6 gene showed a decrease in the core 1 structure, Galbeta1,3GalNAcalpha1-Ser/Thr, probably due to competition between the core 1 synthase and core 3 synthase. The migration activity of the transfectants was markedly lower than that of mock transfectants in vitro, and lung metastasis after i.v. injection of the transfectants into nude mice was significantly suppressed. These findings indicated that the core structures of O-glycans are profoundly involved in the metastatic capacity of cancer cells.

Expression of ezrin in prostatic intraepithelial neoplasia

Objectives To study the protein expression and gene copy number of ezrin in a set of high-grade prostatic intraepithelial neoplasia (HGPIN) samples with concomitant prostate cancer. Ezrin is a cytoskeleton linker protein that is actively involved in regulating the growth and metastatic capacity of cancer cells. Methods Nineteen HGPIN samples obtained from radical prostatectomy specimens were used for the study. Among them, 13 samples also contained invasive prostate cancer. The expression of ezrin was studied by immunohistochemistry. The same samples were also used for fluorescence in situ hybridization to study the gene copy number of ezrin. Results Immunoreactivity for ezrin was absent or weak in benign prostatic epithelial cells. Weak or moderate immunostaining was detected in 11 of 13 prostate cancer specimens. However, the immunostaining was moderate or strong in all HGPIN samples. In addition, whenever HGPIN and prostate cancer were found in the same sample, the staining was always more intense in the HGPIN cells than in the cancer cells. No alteration was found in the gene copy number detected by fluorescence in situ hybridization. Conclusions We have shown that ezrin is overexpressed in HGPIN and prostate cancer compared with adjacent benign prostatic epithelium. In addition, HGPIN has a greater expression level of ezrin compared with that of prostate cancer. Our results indicate that aberrant expression of ezrin might be involved in the pathogenesis of prostate cancer, and ezrin expression may be useful for the diagnosis of HGPIN.