Significance of transforming growth factor β1 as a new tumor marker for colorectal cancer

Transforming growth factor β1 (TGF‐β1) is thought to be involved in cancer growth and progression. TGF‐β1 changes to its active form after being secreted in its latent form. Our aim was to clarify the significance of plasma concentrations of active and total TGF‐β1 of patients with colorectal cancer. Plasma concentrations of active and total TGF‐β1 in 45 patients with colorectal cancer and 23 healthy volunteers were measured using ELISA and the activation rate (ratio of active to total TGF‐β1) was determined. Plasma concentrations of active TGF‐β1 (21.9 ± 12.8 pg/ml) were significantly higher in patients with colorectal cancer than in healthy volunteers (9.9 ± 5.9 pg/ml; p < 0.001, Welch's t‐test). Concentration of total TGF‐β1 was also significantly higher for patients with colorectal cancer (18.0 ± 13.0 ng/ml vs. 11.1 ± 6.4 ng/ml; p < 0.01, Welch's t‐test). However, there was no significant difference in the TGF‐β1 activation rate between the 2 groups. There was a correlation between Dukes' stage and plasma concentration of active or total TGF‐β1 (p < 0.01, Spearman's rank correlation test) and on day 7 the active TGF‐β1 levels for patients recovering from curative resection were similar to those of the control group of healthy volunteers. These results suggest that active TGF‐β1 might be used as a tumor marker for colorectal cancer. © 2001 Wiley‐Liss, Inc.     

Expression of retinoic acid α and β receptor genes in liver and hepatocellular carcinoma

cDNA probes for human retinoic acid receptors α and β (RARα and RARβ) were modified for use as specific hybridization probes to study hepatocellular carcinomas (HCC) and cell lines, liver regeneration, and fetal development. RARβ mRNA was detected at low levels in adult liver and rose markedly during the early phase of liver regeneration. RARβ mRNA was present at very low levels in HCC and was not detected in fetal liver. In contrast, RARα mRNA was present at low levels in normal liver, but showed a marked elevation in several HCCs and cell lines. Growth of cell lines was altered by retinoic acid (RA), but the effects could not be predicted by the levels of either RARα or RARβ mRNA. However, the response correlated with cell phenotype. Three cell lines with an adult phenotype (high albumin and low α-fetoprotein) were inhibited by RA, two undifferentiated lines showed moderate growth stimulation, and two of three cell lines that had high levels of α-fetoprotein were markedly stimulated by RA.     

Interleukin-1α gene expression and localization of interleukin-1α protein during tumor promotion

Treatment of the dorsal epidermis of SENCAR mice with 12-O-tetradecanoylphorbol-13-acetate (TPA) induced a time- and dose-dependent stimulation of interleukin-1α (IL-1α) gene expression. Levels of IL-1α mRNA were elevated as early as 15 min and peaked at 3–4 h after a single application of TPA (2 μg or 10 μg). IL-1α gene expression increased in epidermal tissue isolated from SENCAR mice at 1, 3, 6, 10, 16, and 22 wk after a single treatment with 10 nmol 7,12-dimethylbenz[a]anthracene (DMBA) and subsequent twice-weekly application of 2 μg TPA. IL-1α-immunoreactive protein was specifically localized within suprabasal keratinocytes in cutaneous tissue isolated from mice treated with DMBA-TPA for 1–22 wk and in nonproliferating cells located within papilloma tissue isolated from SENCAR mice at 22 wk after initiation and promotion. Basal cells within hyperplastic epidermis did not produce IL-1α-immunoreactive protein. DMBA treatment alone did not induce IL-1α gene expression. Injection of IL-1α-specific antibodies (50 μg) into SENCAR mice via the tail vein 2 h before treatment with TPA (2 μg or 10 μg) significantly (P < 0.05) inhibited the skin thickening usually observed 24 h after treatment with TPA. Autoradiography studies showed that injection of anti—IL-1α antibodies inhibited incorporation of [methyl-³H]thymidine by keratinocytes within the epidermis and by cells within hair follicles. It also inhibited neutrophil infiltration into the dermis, which usually results from topical application of TPA. These data suggest that IL-1α is a pivotal cytokine produced by specific subpopulations of epidermal keratinocytes and that IL-1α primarily regulates the epidermal proliferative response of a distinctly separate population of keratinocytes after topical exposure of murine epidermis to TPA and secondarily modulates neutrophil migration into the dermis. Consequently, manipulation of IL-1α may be a way to attenuate or abrogate the cutaneous response to TPA by altering keratinocyte proliferation, the resultant hyperplasia, and a portion of the inflammatory response characterized by dermal infiltration of neutrophils.     

Opposing effects of HIF1α and HIF2α on chromaffin cell phenotypic features and tumor cell proliferation: Insights from MYC‐associated factor X

Pheochromocytomas and paragangliomas (PPGLs) are catecholamine‐producing chromaffin cell tumors with diverse phenotypic features reflecting mutations in numerous genes, including MYC‐associated factor X (MAX). To explore whether phenotypic differences among PPGLs reflect a MAX‐mediated mechanism and opposing influences of hypoxia‐inducible factor (HIF)s HIF2α and HIF1α, we combined observational investigations in PPGLs and gene‐manipulation studies in two pheochromocytoma cell lines. Among PPGLs from 140 patients, tumors due to MAX mutations were characterized by gene expression profiles and intermediate phenotypic features that distinguished these tumors from other PPGLs, all of which fell into two expression clusters: one cluster with low expression of HIF2α and mature phenotypic features and the other with high expression of HIF2α and immature phenotypic features due to mutations stabilizing HIFs. Max‐mutated tumors distributed to a distinct subcluster of the former group. In cell lines lacking Max, re‐expression of the gene resulted in maturation of phenotypic features and decreased cell cycle progression. In cell lines lacking Hif2α, overexpression of the gene led to immature phenotypic features, failure of dexamethasone to induce differentiation and increased proliferation. HIF1α had opposing actions to HIF2α in both cell lines, supporting evolving evidence of their differential actions on tumorigenic processes via a MYC/MAX‐related pathway. Requirement of a fully functional MYC/MAX complex to facilitate differentiation explains the intermediate phenotypic features in tumors due to MAX mutations. Overexpression of HIF2α in chromaffin cell tumors due to mutations affecting HIF stabilization explains their proliferative features and why the tumors fail to differentiate even when exposed locally to adrenal steroids.     

Phosphorylated form of pyruvate dehydrogenase α1 mediates tumor necrosis factor α-induced glioma cell migration

Cell migration is an important factor influencing the treatment outcomes of high-grade glioma (World Health Organization grades III–IV). Using immunohistochemical staining, the present study demonstrated that the protein levels of phosphorylated pyruvate dehydrogenase α1 (p-PDHA1) were increased according to the grade of glioma. Moreover, p-PDHA1 mediated tumor necrosis factor-α (TNF-α)-induced cell migration in glioma cells. Phalloidin staining and western blot analysis were used to detect the protein level of p-PDHA1 in U251 glioma cells stimulated by TNF-α at different time points. Phalloidin staining was used to observe the cytoskeletal structure. The effects on the expression of specific migration markers and on the cytoskeletal structure were also detected. Dichloroacetic acid is an inhibitor of PDK. These results indicated that p-PDHA1 served an important role in the migration of glioma cells, and consequently in the development of glioma.     

Overexpression of leucine‐rich α2‐glycoprotein‐1 is a prognostic marker and enhances tumor migration in gastric cancer

Gastric cancer is one of the most common malignant tumors. Although improvement in chemotherapy has been achieved, the clinical prognosis of advanced gastric cancer remains poor. Therefore, it is increasingly important to predict the prognosis and determine whether patients should or should not receive neoadjuvant or adjuvant chemotherapy. Leucine‐rich α2‐glycoprotein‐1 (LRG1) is overexpressed during inflammation and is associated with various malignancies. In this study, we assessed LRG1 expression in cancer specimens and in the sera of patients with cancer to clarify the usefulness of LRG1 as a biomarker in gastric cancer. This study enrolled 239 (for immunohistochemical staining; IHC) and 184 (for ELISA) patients with gastric cancer. Results of IHC showed that LRG1 expression was significantly associated with histological type, lymphatic and venous invasion, tumor and node factors, and disease stage. Overall survival was significantly worse in the high LRG1 expression group than in the low LRG1 group (P = 0.0003). Cox multivariate analysis of overall survival revealed that LRG1 expression was an independent prognostic factor (P = 0.0258). Serum LRG1 was significantly higher in gastric cancer patients than in healthy volunteers, and increased as the pathological stage progressed. Furthermore, a significant correlation was revealed between serum LRG1 level and LRG1 expression with IHC (P < 0.0001). Inhibition of LRG1 significantly decreased cell proliferation in vitro (migratory and invasive capacity of gastric cancer cells). These results suggest that LRG1 expression in tumors and serum may be a useful prognostic marker in gastric cancer patients.     

PD‐L1 expression enhancement by infiltrating macrophage‐derived tumor necrosis factor‐α leads to poor pancreatic cancer prognosis

Immunotherapy using anti‐PD‐1/PD‐L1 antibodies for several types of cancer has received considerable attention in recent decades. However, the molecular mechanism underlying PD‐L1 expression in pancreatic ductal adenocarcinoma (PDAC) cells has not been clearly elucidated. We investigated the clinical significance and regulatory mechanism of PD‐L1 expression in PDAC cells. Among the various cytokines tested, tumor necrosis factor (TNF)‐α upregulated PD‐L1 expression in PDAC cells through NF‐κB signaling. The induction of PD‐L1 expression was also caused by co‐culture with activated macrophages, and the upregulation was inhibited by neutralization with anti‐TNF‐α antibody after co‐culture with activated macrophages. PD‐L1 expression in PDAC cells was positively correlated with macrophage infiltration in tumor stroma of human PDAC tissues. In addition, survival analysis revealed that high PD‐L1 expression was significantly associated with poor prognosis in 235 PDAC patients and especially in patients harboring high CD8‐positive T‐cell infiltration. These findings indicate that tumor‐infiltrating macrophage‐derived TNF‐α could be a potential therapeutic target for PDAC.     

Endometrial cancer invasion depends on cancer‐derived tumor necrosis factor‐α and stromal derived hepatocyte growth factor

Cancer invasion is an outcome of interactions of the cancer and the host cell. It is now becoming increasingly clear that ovarian hormones have a huge influence on such intercommunications in various types of cancers. Estrogen is known to aggravate the aggressiveness of the endometrial cancer whereas progesterone seems to act as a negative factor. Insight into the mode of ovarian hormonal actions could come from the studies of its regulation of the paracrine interactions between the endometrial cancer and the normal stromal cells during the cancer invasion. In this context, we report here that estrogen promotes the endometrial cancer invasion by inducing humoral interactions between the cancer and the stromal cells, i.e., estrogen stimulates tumor necrosis factor‐α expression from the endometrial cancer cells, which, in turn, induces the stromal expression of hepatocyte growth factor (HGF), conferring the enhanced NK4 (HGF‐antagonist/angiogenesis inhibitor)‐sensitive invasion characteristic of the endometrial cancer cells. Additionally, we demonstrate a close correlation of the invasion of endometrial cancer cells with the expression and dimerization of integrin α_vβ₅ as well as the activation of focal adhesion kinase as the consequences of paracrine interactions. Thus, understanding of paracrine interactions of cancer cells with host stromal cells can yield new insight into the architecture and function of cancer invasion and metastasis, leading to a development of a new cancer therapeutic intervention. © 2008 Wiley‐Liss, Inc.     

Phenotypic characterization and prognostic impact of circulating γδ and αβ T‐cells in metastatic malignant melanoma

Human T cells carrying γδ T‐cell receptors (TCRs) represent a minor population relative to those with αβ TCRs. There has been much interest recently in the possibility of using these γδ T‐cells in cancer therapy because they can kill tumor cells in vitro in an MHC‐unrestricted manner, and possess potential regulatory capability and antigen‐presenting capacity. The presence of γδ T‐cells in late‐stage melanoma patients and their relationship with survival has not been extensively explored, although relatively lower percentages of total γδ T‐cells and Vδ2+ cells have been reported. Here, we present a detailed analysis of associations of γδ T‐cell subsets and differentiation stages with survival in Stage IV patients, compared with CD4+ and CD8+ αβ T‐cells. We found an increased Vδ1:Vδ2‐ratio and a decreased CD4:CD8‐ratio in patients compared to healthy controls, on the basis both of relative frequencies and absolute cell counts per μL blood. Nonetheless, Kaplan–Meier analyses showed that a higher than median frequency of Vδ1+ cells was negatively associated with survival, whereas there were no positive or negative associations with frequencies of Vδ2+ cells. Correlations of cell differentiation status with survival revealed a negative association of early‐differentiated Vδ1+ T cells with survival, both on the basis of relative frequencies and absolute counts. There was also a positive correlation between the frequencies of early‐differentiated CD8+ αβ T‐cells and survival. Our findings suggest peripheral blood frequencies of Vδ1+ T‐cells as a potential prognostic marker in melanoma. The mechanisms by which higher abundance of Vδ1+ cells are associated with poorer survival require determination.     

Repressive role of stabilized hypoxia inducible factor 1α expression on transforming growth factor β‐induced extracellular matrix production in lung cancer cells

Activation of transforming growth factor β (TGF‐β) combined with persistent hypoxia often affects the tumor microenvironment. Disruption of cadherin/catenin complexes induced by these stimulations yields aberrant extracellular matrix (ECM) production, characteristics of epithelial‐mesenchymal transition (EMT). Hypoxia‐inducible factors (HIF), the hallmark of the response to hypoxia, play differential roles during development of diseases. Recent studies show that localization of cadherin/catenin complexes at the cell membrane might be tightly regulated by protein phosphatase activity. We aimed to investigate the role of stabilized HIF‐1α expression by protein phosphatase activity on dissociation of the E‐cadherin/β‐catenin complex and aberrant ECM expression in lung cancer cells under stimulation by TGF‐β. By using lung cancer cells treated with HIF‐1α stabilizers or carrying doxycycline‐dependent HIF‐1α deletion or point mutants, we investigated the role of stabilized HIF‐1α expression on TGF‐β‐induced EMT in lung cancer cells. Furthermore, the underlying mechanisms were determined by inhibition of protein phosphatase activity. Persistent stimulation by TGF‐β and hypoxia induced EMT phenotypes in H358 cells in which stabilized HIF‐1α expression was inhibited. Stabilized HIF‐1α protein expression inhibited the TGF‐β‐stimulated appearance of EMT phenotypes across cell types and species, independent of de novo vascular endothelial growth factor A (VEGFA) expression. Inhibition of protein phosphatase 2A activity abrogated the HIF‐1α‐induced repression of the TGF‐β‐stimulated appearance of EMT phenotypes. This is the first study to show a direct role of stabilized HIF‐1α expression on inhibition of TGF‐β‐induced EMT phenotypes in lung cancer cells, in part, through protein phosphatase activity.     

Evaluation of the safety,pharmacokinetics and treatment effects of an ανβ3 integrin inhibitor on bone turnover and disease activity in men with hormone‐refractory prostate cancer and bone metastases

Aim: This study aimed to evaluate the safety, pharmacokinetics and treatment effects of an α_νβ₃ integrin inhibitor on bone turnover and disease activity in men with hormone‐refractory prostate cancer (HRPC) and bone metastases. Methods: A total of 21 patients with bone metastases and HRPC were randomized to receive MK‐0429 200 mg b.i.d. or 1600 mg b.i.d. for 4 weeks. Toxicity, pharmacokinetics and markers of bone turnover and tumor activity were examined. Results: Nausea was the most common adverse event: one (200‐mg group) and 11 (1600‐mg group) patients. At 4 weeks, mean AUC_{0–12 h} was 210 mmol*h (200‐mg group) and 673 mmol*h (1600‐mg group); mean C_max values were 42 mmol/L (200‐mg group) and 154 mmol/L (1600‐mg group). Urinary cross‐linked N‐telopeptides of type I collagen to creatinine ratio (uNTx), a bone turnover biomarker, showed a change from baseline of ?43.4 percent (200‐mg group) and ?34.1 percent (1600‐mg group). There was an increase in serum prostate specific antigen (PSA), a marker for disease activity, of 54.1 percent (200‐mg group) and 44.5 percent (1600‐mg group). Conclusion: MK‐0429 was generally well tolerated, with the most common side‐effect being nausea. There was some evidence of an early reduction of bone turnover, indicating a potential for clinical use in the treatment of MBD although serum PSA was unexpectedly increased during the study.     

Bone sialoprotein‐αvβ3 integrin axis promotes breast cancer metastasis to the bone

The underlying mechanisms of breast cancer cells metastasizing to distant sites are complex and multifactorial. Bone sialoprotein (BSP) and αvβ3 integrin were reported to promote the metastatic progress of breast cancer cells, particularly metastasis to bone. Most theories presume that BSP promotes breast cancer metastasis by binding to αvβ3 integrin. Interestingly, we found the αvβ3 integrin decreased in BSP silenced cells (BSPi), which have weak ability to form bone metastases. However, the relevance of their expression in primary tumor and the way they participate in metastasis are not clear. In this study, we evaluated the relationship between BSP, αvβ3 integrin levels, and the bone metastatic ability of breast cancer cells in patient tissues, and the data indicated that the αvβ3 integrin level is closely correlated to BSP level and metastatic potential. Overexpression of αvβ3 integrin in cancer cells could reverse the effect of BSPi in vitro and promote bone metastasis in a mouse model, whereas knockdown of αvβ3 integrin have effects just like BSPi. Moreover, The Cancer Genome Atlas data and RT‐PCR analysis have also shown that SPP1, KCNK2, and PTK2B might be involved in this process. Thus, we propose that αvβ3 integrin is one of the downstream factors regulated by BSP in the breast cancer‐bone metastatic cascade.     

γ‐Glutamyltransferase and cancer risk: The Korean cancer prevention study

Elevated serum γ‐glutamyltransferase (GGT) is a marker of hepatic injury and is associated with risk of chronic disease. However, the value of GGT as a biomarker for cancer risk remains unclear. Therefore, we evaluated the association of serum GGT with cancer incidence among more than 1.6 million Koreans. We included 1,662,087 Koreans (1,108,121 men and 553,966 women aged 20–95 years) who received health insurance from the National Health Insurance Service and had a biennial medical evaluation between 1995 and 1998. Follow‐up was through December 2012. Using Cox proportional hazards models, we adjusted for age, smoking status, alcohol consumption, exercise and body mass index after exclusion of early cases (cancer diagnosis or death within 5 years of starting follow‐up) and estimated hazard ratios (HRs) of overall and organ‐specific cancer incidence by GGT quintiles. During the 17‐year follow‐up, 129,087 new cancer cases occurred among the participants. Across levels of GGT, there was a positive gradient of HR and the highest quintile of GGT (≥60 IU/L) had the highest HR for all cancers in both men and women. By cancer site, the association was strongest for liver cancer, comparing the highest and lowest strata in men [HR, 6.67; 95% confidence interval (95%CI), 5.88–7.57] and in women (HR, 7.57; 95%CI, 6.41–8.94). Significant associations were also observed for cancers of the esophagus, larynx, stomach, colorectal, bile duct and lung in men and of the bile duct in women. Increased serum GGT level is independently associated with risk of cancer.     

The combination therapy of α‐galactosylceramide and 5‐fluorouracil showed antitumor effect synergistically against liver tumor in mice

α‐Galactosylceramide (α‐GalCer) has been reported to be therapeutic against metastatic liver tumors in mice. However, little is known regarding the efficacy of combined chemo‐immunotherapy using α‐GalCer and anticancer drugs. In this study, we evaluated the antitumor effect of the combination therapy of α‐GalCer and 5‐fluorouracil (5‐FU) against liver tumors of MC38 colon cancer cells. The liver weights of tumor‐bearing mice treated with the combination were significantly lower than those of nontreated mice and of mice treated with 5‐FU or α‐GalCer alone. No toxic effects on the liver and renal functions were observed in any of the treatment groups. α‐GalCer treatment induced significant activation of liver NK cells in vivo, but 5‐FU treatment did not. 5‐FU treatment resulted in a significant upregulation of NKG2D activating molecules (Rae‐1 and H60) and DNAM‐1 ligands (CD112 and CD155) on MC38 cells, but α‐GalCer did not. The cytolytic activity of α‐GalCer‐activated liver mononuclear cells against 5‐FU‐treated MC38 cells was significantly higher than that against nontreated cells. The increase of the cytolytic activity induced by 5‐FU partially depended on NKG2D‐Rae‐1 or H60 signals. Depletion of NK cells significantly inhibited the antitumor efficacy of 5‐FU against MC38 liver tumors, which suggested that the antitumor effect of 5‐FU partially depended on the cytolytic activity of NK cells. These results demonstrated that the combination therapy of α‐GalCer and 5‐FU produced synergistic antitumor effects against liver tumors by increasing the expression of NK activating molecules on cancer cells. This study suggests a promising new chemo‐immunotherapy against metastatic liver cancer.     

TNF‐α enhances TGF‐β‐induced endothelial‐to‐mesenchymal transition via TGF‐β signal augmentation

The tumor microenvironment (TME) consists of various components including cancer cells, tumor vessels, cancer‐associated fibroblasts (CAFs), and inflammatory cells. These components interact with each other via various cytokines, which often induce tumor progression. Thus, a greater understanding of TME networks is crucial for the development of novel cancer therapies. Many cancer types express high levels of TGF‐β, which induces endothelial‐to‐mesenchymal transition (EndMT), leading to formation of CAFs. Although we previously reported that CAFs derived from EndMT promoted tumor formation, the molecular mechanisms underlying these interactions remain to be elucidated. Furthermore, tumor‐infiltrating inflammatory cells secrete various cytokines, including TNF‐α. However, the role of TNF‐α in TGF‐β‐induced EndMT has not been fully elucidated. Therefore, this study examined the effect of TNF‐α on TGF‐β‐induced EndMT in human endothelial cells (ECs). Various types of human ECs underwent EndMT in response to TGF‐β and TNF‐α, which was accompanied by increased and decreased expression of mesenchymal cell and EC markers, respectively. In addition, treatment of ECs with TGF‐β and TNF‐α exhibited sustained activation of Smad2/3 signals, which was presumably induced by elevated expression of TGF‐β type I receptor, TGF‐β2, activin A, and integrin αv, suggesting that TNF‐α enhanced TGF‐β‐induced EndMT by augmenting TGF‐β family signals. Furthermore, oral squamous cell carcinoma‐derived cells underwent epithelial‐to‐mesenchymal transition (EMT) in response to humoral factors produced by TGF‐β and TNF‐α‐cultured ECs. This EndMT‐driven EMT was blocked by inhibiting the action of TGF‐βs. Collectively, our findings suggest that TNF‐α enhances TGF‐β‐dependent EndMT, which contributes to tumor progression.     

Multipotent mesenchymal stromal cells promote tumor growth in distinct colorectal cancer cells by a β1‐integrin‐dependent mechanism

Tumor–stroma interactions play an essential role in the biology of colorectal carcinoma (CRC). Multipotent mesenchymal stromal cells (MSC) may represent a pivotal part of the stroma in CRC, but little is known about the specific interaction of MSC with CRC cells derived from tumors with different mutational background. In previous studies we observed that MSC promote the xenograft growth of the CRC cell‐line DLD1. In the present study, we aimed to analyze the mechanisms of MSC‐promoted tumor growth using various in vitro and in vivo experimental models and CRC cells of different mutational status. MSC specifically interacted with distinct CRC cells and supported tumor seeding in xenografts. The MSC–CRC interaction facilitated three‐dimensional spheroid formation in CRC cells with dysfunctional E‐cadherin system. Stable knock‐downs revealed that the MSC‐facilitated spheroid formation depended on β1‐integrin in CRC cells. Specifically in α‐catenin‐deficient CRC cells this β1‐integrin‐dependent interaction resulted in a MSC‐mediated promotion of early tumor growth in vivo. Collagen I and other extracellular matrix compounds were pivotal for the functional MSC–CRC interaction. In conclusion, our data demonstrate a differential interaction of MSC with CRC cells of different mutational background. Our study is the first to show that MSC specifically compared to normal fibroblasts impact early xenograft growth of distinct α‐catenin deficient CRC cells possibly through secretion of extracellular matrix. This mechanism could serve as a future target for therapy and metastasis prevention.     

CDH6‐activated αIIbβ3 crosstalks with α2β1 to trigger cellular adhesion and invasion in metastatic ovarian and renal cancers

Cadherin 6 (CDH6) is significantly overexpressed in advanced ovarian and renal cancers. However, the role of CDH6 in cancer metastasis is largely unclear. Here, we investigated the impact of CDH6 expression on integrin‐mediated metastatic progression. CDH6 preferentially bound to αIIbβ3 integrin, a platelet receptor scarcely expressed in cancer cells, and this interaction was mediated through the cadherin Arginine–glycine–aspartic acid (RGD) motif. Furthermore, CDH6 and CDH17 were found to interact with α2β1 in αIIbβ3^low cells. Transient silencing of CDH6, ITGA2B, or ITGB3 genes caused a significant loss of proliferation, adhesion, invasion, and lung colonization through the downregulation of SRC, FAK, AKT, and ERK signaling. In ovarian and renal cancer cells, integrin αIIbβ3 activation appears to be a prerequisite for proper α2β1 activation. Interaction of αIIbβ3 with CDH6, and subsequent αIIbβ3 activation, promoted activation of α2β1 and cell adhesion in ovarian and renal cancer cells. Additionally, monoclonal antibodies specific to the cadherin RGD motif and clinically approved αIIbβ3 inhibitors could block pro‐metastatic activity in ovarian and renal tumors. In summary, the interaction between CDH6 and αIIbβ3 regulates α2β1‐mediated adhesion and invasion of ovarian and renal cancer metastatic cells and constitutes a therapeutic target of broad potential for treating metastatic progression.     

Retinoid X receptor α enhances human cholangiocarcinoma growth through simultaneous activation of Wnt/β‐catenin and nuclear factor‐κB pathways

Retinoid X receptor α (RXRα) plays important roles in the malignancy of several cancers such as human prostate tumor, breast cancer, and thyroid tumor. However, its exact functions and molecular mechanisms in cholangiocarcinoma (CCA), a chemoresistant carcinoma with poor prognosis, remain unclear. In this study we found that RXRα was frequently overexpressed in human CCA tissues and CCA cell lines. Downregulation of RXRα led to decreased expression of mitosis‐promoting factors including cyclin D1and cyclin E, and the proliferating cell nuclear antigen, as well as increased expression of cell cycle inhibitor p21, resulting in inhibition of CCA cell proliferation. Furthermore, RXRα knockdown attenuated the expression of cyclin D1 through suppression of Wnt/β‐catenin signaling. Retinoid X receptor α upregulated proliferating cell nuclear antigen expression through nuclear factor‐κB (NF‐κB) pathways, paralleled with downregulation of p21. Thus, the Wnt/β‐catenin and NF‐κB pathways account for the inhibition of CCA cell growth induced by RXRα downregulation. Retinoid X receptor α plays an important role in proliferation of CCA through simultaneous activation of Wnt/β‐catenin and NF‐κB pathways, indicating that RXRα might serve as a potential molecular target for CCA treatment.     

Tumor vasculature‐targeted delivery of tumor necrosis factor‐α*

BACKGROUND: Recently, considerable efforts have been directed toward antivascular therapy as a new modality to treat human cancers. However, targeting a therapeutic gene of interest to the tumor vasculature with minimal toxicity to other tissues remains the objective of antivascular gene therapy. Tumor necrosis factor‐α (TNF‐α) is a potent antivascular agent but has limited clinical utility because of significant systemic toxicity. At the maximum tolerated doses of systemic TNF‐α, there is no meaningful antitumor activity. Hence, the objective of this study was to deliver TNF‐α targeted to tumor vasculature by systemic delivery to examine its antitumor activity. METHODS: A hybrid adeno‐associated virus phage vector (AAVP) was used that targets tumor endothelium to express TNF‐α (AAVP‐TNF‐α). The activity of AAVP‐TNF‐α was analyzed in various in vitro and in vivo settings using a human melanoma tumor model. RESULTS: In vitro, AAVP‐TNF‐α infection of human melanoma cells resulted in high levels of TNF‐α expression. Systemic administration of targeted AAVP‐TNF‐α to melanoma xenografts in mice produced the specific delivery of virus to tumor vasculature. In contrast, the nontargeted vector did not target to tumor vasculature. Targeted AAVP delivery resulted in expression of TNF‐α, induction of apoptosis in tumor vessels, and significant inhibition of tumor growth. No systemic toxicity to normal organs was observed. CONCLUSIONS: Targeted AAVP vectors can be used to deliver TNF‐α specifically to tumor vasculature, potentially reducing its systemic toxicity. Because TNF‐α is a promising antivascular agent that currently is limited by its toxicity, the current results suggest the potential for clinical translation of this strategy. Cancer 2009. Published 2008 by the American Cancer Society.