3685

ITGAV

integrin, alpha V;ITGAV;integrin, alpha V

antigen identified by monoclonal antibody L230|integrin alpha-V|integrin alphaVbeta3|integrin, alpha V (vitronectin receptor, alpha polypeptide, antigen CD51)|vitronectin receptor subunit alpha

2q31-q32

protein-coding

expression of adhesion receptor integrin alphavbeta3 in an activated functional form strongly promotes metastasis in human breast cancer cells. Here, we report that alphavbeta3 cooperates with matrix metalloproteinase type 9 (MMP-9) in breast cancer cell migration. This cooperation is regulated by the activation state of the integrin. expression of activated alphavbeta3 in metastatic variants of MDA-MB 435 human breast cancer cells and primary metastatic cells from breast cancer patients strongly enhanced migration toward vitronectin and fibrinogen. This enhancement was mediated by a soluble factor produced by breast cancer cells expressing activated alphavbeta3. When transferred, this factor also up-regulated alphavbeta3-dependent migration of breast cancer cells that express the nonactivated integrin. The factor was identified as metalloproteinase MMP-9. Whereas all tested breast cancer cell variants produced latent MMP-9, only those with activated alphavbeta3 produced the mature form of this metalloproteinase. Recombinant mature MMP-9, but not latent MMP-9 or either form of MMP-2, enhanced alphavbeta3-dependent breast cancer cell migration. The migratory response was inhibited by tissue inhibitors of metalloproteinase or when MMP-9 was depleted from the inducing supernatants. The results indicate a causal relationship between the expression of activated integrin alphavbeta3 and production of enzymatically active MMP-9 in metastatic breast cancer cells. These molecules cooperate to enhance breast cancer cell migration toward specific matrix proteins, and this may contribute to the strongly enhanced metastatic capacity of breast cancer cells that express activated alphavbeta3.

CNTO 95 is a fully human monoclonal antibody that recognizes alphav integrins. Previous studies have shown that CNTO 95 exhibits both anti-tumor and anti-angiogenic activities (Trikha M et al., Int J cancer 110:326-335, 2004). In this study we investigated the biological activities of CNTO 95 on breast tumor cells both in vitro and in vivo. In vitro treatment with CNTO 95 decreased the viability of breast tumor cells adhering to vitronectin. CNTO 95 inhibited tumor cell adhesion, migration, and invasion in vitro. CNTO 95 treatment also induced tyrosine dephosphorylation of focal adhesion kinase (FAK), and the docking protein paxillin that recruits both structural and signaling molecules to focal adhesions (Turner CE, Int J Biochem Cell Biol 30:955-959, 1998; O'Neil GM et al., Trends Cell Biol 10:111-119, 2000). These results suggest that CNTO 95 inhibits breast tumor cell growth, migration and invasion by interruption of alphav integrin mediated focal adhesions and cell motility signals. In vivo studies of CNTO 95 were conducted in an orthotopic breast tumor xenograft model. Treatment with CNTO 95 resulted in significant inhibition of both tumor growth and spontaneous metastasis of MDA-MB-231 cells to the lungs. CNTO 95 also inhibited lung metastasis in a separate experimental (tail vein injection) model of metastasis. The results presented here demonstrate the anti-tumor and anti-metastatic activities of CNTO 95 in breast cancer models and provide insight into the cellular and molecular mechanisms mediating its inhibitory effects on metastasis.

Periostin, a matricellular protein, is overexpressed in the stroma of several cancers. The aim of our study was to investigate more specifically whether periostin expression is associated with bone metastases from breast cancer and to determine its source in the affected bone. Nude mice were inoculated with human MDA-B02 breast cancer cells. Bone metastases-bearing mice were treated with zoledronic acid-an antiresorptive drug-or vehicle. Bone metastases were examined for tumor- and stroma-derived periostin expression by quantitative polymerase chain reaction with human- and mouse-specific primers and immunohistochemistry. Serum periostin and conventional bone turnover markers were also measured. MDA-B02 cells did not express periostin both in vitro and in vivo. However, mouse-derived periostin was markedly overexpressed (eightfold) in metastatic legs compared to noninoculated mice. Serum periostin levels were also markedly increased in metastatic mice and correlated with in situ expression levels. Immunostaining showed that periostin derived from the environing stromal cells of bone metastasis. Bone turnover blockade by zoledronic acid markedly decreased osteolytic lesions but only slightly modulated serum periostin levels. Bone metastases from breast cancer induce overexpression of periostin by surrounding stromal cells. Periostin could be a biochemical marker of the early stromal response associated to breast cancer bone metastasis formation.

BACKGROUND: Although integrins have been implicated in the progression of breast cancer, the exact mechanism whereby they exert this regulation is clearly not understood. To understand the role of integrins in breast cancer, we examined the expression levels of several integrins in mouse breast cancer cell lines by flow cytometry and the data were validated by Western and RT-PCR analysis. The importance of integrins in cell migration and cell invasion was examined by in vitro assays. Further the effect of integrins on metastasis was investigated by in vivo experimental metastasis assays using mouse models. RESULTS: Integrin alpha5 subunit is highly expressed in the nonmetastatic cell line 67NR and is significantly low in the highly invasive cell line 4T1. In contrast, expression levels of integrin alpha6 subunit are high in 4T1 cells and low in 67NR cells. In vitro data indicated that overexpression of alpha5 subunit and knockdown of alpha6 integrin subunit inhibited cell proliferation, migration, and invasion. Our in vivo findings indicated that overexpression of integrin alpha5 subunit and knockdown of alpha6 subunit decreased the pulmonary metastasis property of 4T1 cells. Our data also indicated that overexpression of alpha 5 integrin subunit and suppression of alpha6 integrin subunit inhibited cells entering into S phase by up-regulating p27, which results in downregulation of cyclinE/CDK2 complexes, This suggests that these integrins regulate cell growth through their effects on cell-cycle-regulated proteins. We also found that modulation of these integrins upregulates E2F, which may induce the expression of chk1 to regulate cdc25A/cyclin E/CDK2/Rb in a feedback loop mechanism. CONCLUSION: This study indicates that Integrin alpha5 subunit functions as a potential metastasis suppressor, while alpha6 subunit functions as a metastasis promoter. The modulation of integrins reduces cdc25 A, another possible mechanism for downregulation of CDK2. Taken together we demonstrate a link between integrins and the chk1-cdc25-cyclin E/CDK2-Rb pathway.

Cyclin D1b, a splice variant of the cell cycle regulator cyclin D1, holds oncogenic functions in human cancer. However, the mechanisms underlying cyclin D1b function remain poorly understood. Here we introduced wild-type cyclin D1a or cyclin D1b variant into non-metastatic MCF-7 cells. Our results show that ectopic expression of cyclin D1b promotes invasiveness of the cancer cells in a cyclin D1a independent manner. Specifically, cyclin D1b is found to modulate the expression of alphavbeta3, which characterizes the metastatic phenotype, and enhance tumor cell invasive potential in cooperating with HoxD3. Notably, cyclin D1b promotes alphavbeta3-mediated adhesion and invasive migration, which are associated with invasive potential of breast cancer cells. Further exploration indicates that cyclin D1b makes breast cancer cells more sensitive to toll-like receptor 4 ligand released from damaged tumor cells. These findings reveal a role of cyclin D1b as a possible mediator of alphavbeta3 transcription to promote tumor metastasis.

Detachment from the extracellular matrix induces a form of programmed cell death termed anoikis. Resistance to anoikis permits cancer cells to survive in systemic circulation and facilitates their metastasis to distant organs. It is well known that S100A4 is overexpressed in many tumors and involved in tumor metastasis, but the mechanisms of the metastasis-promoting function of S100A4 are not fully understood. We hypothesized that S100A4 might play a role in anoikis of gastric cancer cells and further affects their metastasis. To test this hypothesis, we changed the expression of S100A4 by means of RNA interference or experimental overexpression and investigated the effect on anoikis. We found that knockdown of S100A4 by RNA interference led to significantly increased anoikis, whereas overexpression of S100A4 resulted in inhibition of anoikis. Furthermore, we provide evidence that inhibition of S100A4 resulted in the downregulation of alpha5 and alphav integrin expression. These findings suggest that S100A4 protects gastric cancer cells from anoikis by regulation of alphav and alpha5 integrin expression, which sheds a novel light for the role of S100A4 in cancer metastasis.

The mechanisms leading to squamous cell carcinoma of the head and neck (SCCHN) metastasis are incompletely understood. Although evidence shows that the chemokine receptor 7 (CCR7) and its ligand CCL19 may regulate tumor dissemination, their role in SCCHN is not clearly defined. CCR7 has been shown to regulate integrins, which facilitate adhesion of cancer cells to and/or migration through the extracellular matrix (ECM). To investigate the relationship between CCR7 and integrin alphavbeta3 in metastatic SCCHN, we used adhesion and migration assays, immunofluorescence staining and western blotting to determine whether integrin alphavbeta3 can be activated by CCL19 in the metastatic SCCHN cell line PCI-37B, which was pre-incubated with CCL19 or the integrin alphavbeta3 inhibitor, IS201. Our results demonstrate that CCR7 favors PCI-37B cell adhesion and migration, induces reorganization of the actin cytoskeleton and induces integrin alphavbeta3 phosphorylation. The integrin alphavbeta3 inhibitor, IS201, blocked all of these effects. CCR7 and integrin alphavbeta3 expression significantly and positively correlated with tumor size, clinical stage and nodal metastasis. Taken together, our data indicate that CCR7 regulates cell adhesion and migration via integrin alphavbeta3 in metastatic SCCHN. These results should provide the groundwork for new strategies aimed at preventing SCCHN metastasis.

OBJECTIVE: To investigate the expression of integrin alpha-v subunit in laryngeal and hypopharyngeal squamous cell carcinomas (LHSCC) and to correlate the expression ratio with clinic and pathologic features of LHSCC, meanwhile, to investigate the relationship between the expression of integrin alpha-v subunit and tumor angiogenesis. METHODS: A tissue microarray of LHSCC was designed and made. Using this microarray, the expression of integrin alpha-v subunit in LHSCC was studied by immunohistochemistry, and the expression disparity in different clinic and pathologic staging of LHSCC was analyzed. The immunohistochemical staining of CD105 was done in the same microarray, the intratumor microvessel density (IMVD) was calculated by CD105 staining. The relationship between integrin alpha-v subunit expression and the IMVD was analyzed. RESULTS: In primary cancer tissue, the expression ratio of integrin alpha-v subunit was 68.0% (51/75), significantly higher than normal tissue beside cancer (10.3%, 3/29, chi2 = 28.68, P < 0.001); the expression ratio of integrin alpha-v subunit in lymph node metastatic carcinoma was 100.00% (20/20), significantly higher than normal tissue (chi2 = 38.77, P < 0.001) and primary cancer tissue (chi2 = 12.69, P < 0.05); in group with lymph node metastasis, the expression ratio of integrin alpha-v subunit was significantly higher than group without lymph node metastasis (chi2 = 10.87, P < 0.001); the IMVD of the group with integrin alpha-v subunit positive expression was significantly higher than the group with integrin alpha-v subunit negative expression (P < 0.001). CONCLUSIONS: There was significant relationship between integrin alpha-v subunit expression and lymphatic metastasis of LHSCC. Overexpression of the integrin alpha-v subunit may have contributed to the tumor angiogenesis and lead to lymphatic metastasis. Integrin alpha-v subunit may become a novel lymphatic metastasis marker of LHSCC.

The objective of this study was to investigate the role of alpha v-integrin subunit (ITGAV, CD51) in invasion and metastasis of the laryngeal and hypopharyngeal squamous cell carcinomas and to evaluate whether an antisense oligonucleotide sequence (ASONs) targeting ITGAV gene can result in proliferative inhibition and induce to apoptosis of laryngeal carcinoma cell lines (Hep-2). Firstly, a tissue microarray contained 75 primary carcinomas, 29 non-cancerous normal tissues and 20 metastatic lymph nodes was constructed and used to detect the expression of ITGAV by immunohistochemistry. The changes of ITGAV expression from each group were assessed and correlated to the clinical parameters of the patients. Secondly, the ASONs targeting ITGAV gene was transfected into Hep-2 cells in vitro. The proliferative ability of the cells after transfection was measured by MTT assay and the apoptosis was detected using flow cytometry. Results showed that the expression of ITGAV was significantly correlated with differentiation and lymph node metastasis of these cancers. In vitro test showed that the proliferative ability of Hep-2 cells was significantly inhibited by ASONs in a way of concentration- and time-depending mode, and a significant apoptosis of Hep-2 cells was also observed after ASONs transfection. In conclusion, the expression of ITGAV was significantly correlated with differentiation and metastasis of the laryngeal and hypopharyngeal carcinomas; down-regulation of ITGAV gene could inhibit proliferation of Hep-2 cells and induce to its apoptosis. These results suggest that ITGAV gene may become a promising prognostic marker and new treatment target for these cancers.

PURPOSE: Osteopontin (OPN) is a secreted phosphoprotein, which functions as a cell attachment protein and cytokine that signals through two cell adhesion molecules, integrin alphavbeta3 and CD44, to regulate cancer growth and metastasis. The aim of this study was to investigate the expression of OPN and integrin alphav (ITGAV, main receptor of the OPN) in laryngeal and hypopharyngeal squamous cell carcinoma (LHSCC) and correlate the expression quantity with tumor biological behavior. METHODS: We have made a tissue microarray of LHSCC and detected the OPN and ITGAV expression by immunohistochemistry in the microarray. The expression quantity of OPN and ITGAV was determined by a professional pathological image processing software (Image-Pro plus 5.02), and the clinical significances of the expression quantity in LHSCC were analyzed. RESULTS: The expression quantity of OPN and ITGAV in primary and metastatic carcinomas is significantly higher than in normal tissues. The expression of OPN and ITGAV in the well-differentiated group is significantly lower than in moderately and poorly differentiated group; the expression quantity of OPN and ITGAV in group with lymph node metastasis is significantly higher than in group without lymph node metastasis. CONCLUSIONS: The expression of OPN and ITGAV was significantly influenced to the differentiation and metastasis of the LHSCC. Overexpression of the OPN and ITGAV may have contributed to invasion and metastasis of the LHSCC, and therefore, OPN and ITGAV may have value as a target for chemotherapy in LHSCC treatment.

Early metastasis is the primary cause of death in melanoma patients. The adhesion receptor integrin alpha v beta 3 contributes to tumor cell functions that are potentially involved in melanoma growth and metastasis. We tested whether integrin alpha v beta 3 supports metastasis of human melanoma cells when injected into the bloodstream of immune deficient mice. Comparing variants of the same melanoma cell type that expressed either alpha v beta 3, alpha IIb beta 3 or no beta 3 integrin, we found that only alpha v beta 3 strongly supported metastasis. Inhibition of tumor cell alpha v beta 3 function reduced melanoma metastasis significantly and prolonged animal survival. To understand mechanisms that allow alpha v beta 3, but not alpha IIb beta 3 to support melanoma metastasis, we analyzed proteolytic and migratory activities of the melanoma cell variants. Melanoma cells expressing alpha v beta 3, but not those expressing alpha IIb beta 3 or no beta 3 integrin, produced the active form of metalloproteinase MMP-2 and expressed elevated mRNA levels of MT1-MMP and TIMP-2. This indicates an association between alpha v beta 3 expression and protease processing. Furthermore, alpha v beta 3 expression was required for efficient melanoma cell migration toward the matrix proteins fibronectin and vitronectin. The results suggest that expression of integrin alpha v beta 3 promotes the metastatic phenotype in human melanoma by supporting specific adhesive, invasive and migratory properties of the tumor cells and that the related integrin alpha IIb beta 3 cannot substitute for alpha v beta 3 in this respect.

To clarify the function of integrin alpha(v)beta3 in the early stage of liver metastasis, we investigated the interactions of metastatic cells with their target organ under the actual blood flow by using positron emission tomography (PET). The cells used were CHO-K1 cells and their transfectants bearing human integrin alpha(v)beta3 cDNA (alpha(v)beta3-CHO-K1 cells). The liver accumulation of alpha(v)beta3-CHO-K1 cells was significantly higher than that of CHO-K1 cells after injection via the portal vein, whereas no significant difference was observed in the lung accumulation after tail vein injection, suggesting a specific interaction of alpha(v)beta3-CHO-K1 cells with the hepatic sinusoids. Furthermore, to clarify the precise location of each cell in the liver, i.e., to determine whether individual cells were intravascularly localized or had extravasated, we performed intravital fluorescence microscopy (IVM) on the liver by using stable transfectants bearing the green fluorescent protein (GFP) gene, namely, GFP-CHO-K1 and GFP-alpha(v)beta3-CHO-K1 cells. Both types of cells remained in the hepatic blood vessels 1 h after injection via the portal vein. On the other hand, expression of integrin alpha(v)beta3 promoted the cells to reach the extravascular region after 24 h. These results suggest the possibility that the specific accumulation of alpha(v)beta3-CHO-K1 cells in the liver is followed by migration of the cells into the extravascular region. Interestingly, the adhesion of the two types of cells to hepatic sinusoidal endothelial cells in vitro did not correspond to in vivo accumulation of these cells. Therefore, integrin alpha(v)beta3 may function to promote extravasation of integrin alpha(v)beta3-expressing tumor cells in liver through a process possibly mediated by vitronectin produced by this organ.

In this study, significantly higher expression of Gal3ST-2 (Gal: 3-O-sulfotransferase-2) and 3'-sulfated glycoconjugates were observed in highly metastastic cancer cells and in larynx cancer tissues with lymph node metastasis than those in lowly metastatic cancer cells and larynx cancer tissues without metastasis (P<0.01, n=42). These results indicated that there was a marked correlation between the expression of Gal3ST-2 and tumor metastasis potential. After RNAi transfection, a striking morphological change of SMMC7721 hepatoma cells from polygon to shuttle shape and significant decrease in adhesion to sL-selectin and HUVEC were observed. Interestingly, the expression of integrin subunit alphaV was markedly downregulated and 3'-sulfated subunit alphaV almost disappeared in the transfectants, but integrin subunit beta3 almost had no change. These results suggested that Gal3ST-2 was involved in tumor metastasis process by regulation of adhesion ability to selectins and expression of integrins.

tumor metastasis is a multistep pathological process involved in the final phase of tumor development. During this process, epithelium-derived tumor cells undergo fibroblast-like transformation, referred to as epithelial-mesenchymal transition (EMT), which contributes to aggressive behavior of tumors. We identify periostin, a mesenchyme-specific gene product, as a contributor to EMT and metastatic potential. Stable expression of a periostin transgene in tumorigenic but non-metastatic 293T cells caused cells to undergo fibroblast-like transformation accompanied by increased expressions of vimentin, epidermal growth factor receptor (EGFR), and matrix metalloproteinase-9. The cells expressing ectopic periostin increased cell migration, invasion, and adhesion by 2-9-fold. Invasive characteristics required signaling through integrin alpha(v)beta5 and EGFR. In addition, periostin-engineered 293T cells formed metastases in immunodeficient mice following either cardiac inoculation or injection into mammary fat pad. These data demonstrate an active role for periostin in EMT and metastasis that requires cross-talk between integrin and EGFR signaling pathways.

Transendothelial migration is a key step in the extravasation of tumor cells during metastasis formation. Here, we have classified 45 human tumor cell lines derived from various tissues according to their capacity for transendothelial migration in vitro. We could distinguish cell lines showing strong transmigration (TEM+ cell lines) from others that did not transmigrate (TEM- cell lines). By DNA microarray analysis we could cluster TEM+ and TEM- cell lines according to their gene expression pattern and identify genes differentially expressed between the 2 groups. Among these we found the integrin beta3 subunit to be highly expressed in TEM+ cell lines as compared to TEM- cell lines. Cell surface localization of alphavbeta3 integrin receptors was exclusively found in TEM+ cell lines. Transendothelial migration of TEM+ cells but not their adhesion to the endothelial cells, or invasion into collagen gels could be blocked with an antibody against alphavbeta3 integrin and by RNAi mediated knock-down of the integrin beta3 subunit. These data establishes alphavbeta3 integrin as one key component of the transendothelial migration process of tumor cells, and as a potential target for anti-metastatic therapy. Our gene expression analysis of a defined collection of tumor cell lines can be used as a starting point to identify further genes functionally involved in transendothelial migration.

The incidence of brain metastasis is rising and poses a severe clinical problem, as we lack effective therapies and knowledge of mechanisms that control metastatic growth in the brain. Here we demonstrate a crucial role for high-affinity tumor cell integrin alpha(v)beta(3) in brain metastatic growth and recruitment of blood vessels. Although alpha(v)beta(3) is frequently up-regulated in primary brain tumors and metastatic lesions of brain homing cancers, we show that it is the alpha(v)beta(3) activation state that is critical for brain lesion growth. Activated, but not non-activated, tumor cell alpha(v)beta(3) supports efficient brain metastatic growth through continuous up-regulation of vascular endothelial growth factor (VEGF) protein under normoxic conditions. In metastatic brain lesions carrying activated alpha(v)beta(3), VEGF expression is controlled at the post-transcriptional level and involves phosphorylation and inhibition of translational respressor 4E-binding protein (4E-BP1). In contrast, tumor cells with non-activated alpha(v)beta(3) depend on hypoxia for VEGF induction, resulting in reduced angiogenesis, tumor cell apoptosis, and inefficient intracranial growth. Importantly, the microenvironment critically influences the effects that activated tumor cell alpha(v)beta(3) exerts on tumor cell growth. Although it strongly promoted intracranial growth, the activation state of the receptor did not influence tumor growth in the mammary fat pad as a primary site. Thus, we identified a mechanism by which metastatic cells thrive in the brain microenvironment and use the high-affinity form of an adhesion receptor to grow and secure host support for proliferation. Targeting this molecular mechanism could prove valuable for the inhibition of brain metastasis.

AIMS: Coxsackie and adenovirus receptor (CAR) is a tumor suppressor and a primary receptor for adenovirus type 5 (Ad5). Our study aims to examine the influence of forced expression of CAR in rhabdomyosarcoma cells (RD) on expression levels of integrins implicated in Ad5 entry, and the effect of CAR on cell-extracellular matrix adhesion and migration. MAIN METHODS: CAR expressing clones were established from RD cells by stable transfection. Flow cytometry was used to evaluate the expression of CAR and integrins. Adhesion was measured in plates previously coated with vitronectin or fibronectin. Boyden chambers were used to investigate migration. Transfection of cells with siRNA was used to achieve integrin silencing. Ad5-mediated transgene expression was measured by beta-gal staining. KEY FINDINGS: Increased expression of CAR in RD cells reduces the expression of alphavbeta3 and alphavbeta5 integrins. Cells overexpressing CAR exhibit significantly reduced adhesion to vitronectin and fibronectin, and reduced cell migration. Specifically silencing alphavbeta3 integrin in RD cells reduced cell migration indicating that reduced migration could be the consequence of alphavbeta3 integrin downregulation. This study also demonstrates the negative effect of reduced levels of alphavbeta3 and alphavbeta5 integrins on Ad5-mediated transgene expression with Ad5 retargeted to alphav integrins. SIGNIFICANCE: The pharmacological upregulation of CAR aimed to increase Ad5-mediated transgene expression may actually downregulate alphavbeta3 and alphavbeta5 integrins and thus alter Ad5-mediated gene transfer. The mechanism of decreased cell migration, a prerequisite for metastasis and invasion, due to increased CAR expression may be explained by reduced alphavbeta3 integrin expression.

The growth of metastatic prostate cancer cells in the bone involves an intimate interaction between the tumor cells and various elements of the bone microenvironment, resulting in increased rate of bone turnover and rapid tumor growth. The alpha(v)beta3 integrin has been shown to play an important role in tumor growth and angiogenesis, and is known to be critical to osteoclast formation and activity. This study was designed to examine the role of alpha(v)beta3 expressed by cells native to the bone in the growth and pathogenesis of prostate cancer bone metastases. Human prostate cancer cells which do not express alpha(v)beta3 or alpha(IIb)beta3 integrins were injected directly into human bone fragments previously implanted subcutaneously in SCID mice (SCID-human-bone model). At the same time treatment with anti-beta3 antibody fragment (m7E3 F(ab')2) i.p. at 300 microg/dose 3 x per week was initiated and continued for 2 weeks. In this system, m7E3 F(ab')2 only recognizes human bone-derived alpha(v)beta3. Antibody inhibition of alpha(v)beta3 integrin in vivo resulted in a specific reduction in the proportion of antigenically-human blood vessels within tumor-bearing bone implants (from 73.5% +/- 3.93 in controls to 17.74% +/- 5.64 in treated animals). Proliferation of the alpha(v)beta3-negative tumor cells was also reduced, although the overall vessel density was maintained by compensating mouse vasculature. Blockage of human bone-derived alpha(v)beta3 also significantly reduced the recruitment of osteoclasts in response to tumor cells, as well as degradation of calcified bone tissue. Together these observations confirm the importance of alpha(v)beta3 in bone metabolism and angiogenesis, and point to the role of these processes in controlling growth of metastatic prostate cancer cells in the bone.

BACKGROUND: Stromal cell-derived factor-1 (SDF-1 or CXCL12) and CXCR4 are key elements in the metastasis of prostate cancer cells to bone--but the mechanisms as to how it localizes to the marrow remains unclear. METHODS: Prostate cancer cell lines were stimulated with SDF-1 and evaluated for alterations in the expression of adhesion molecules using microarrays, FACs, and Western blotting to identify alpha(v)beta(3) receptors. Cell-cell adhesion and invasion assays were used to verify that activation of the receptor is responsive to SDF-1. RESULTS: We demonstrate that SDF-1 transiently regulates the number and affinity of alpha(v)beta(3) receptors by prostate cancer cells to enhance their metastatic behavior by increasing adhesiveness and invasiveness. SDF-1 transiently increased the expression of beta(3) receptor subunit and increased its phosphorylation in metastatic but not nonmetastatic cells. CONCLUSIONS: The transition from a locally invasive phenotype to a metastatic phenotype may be primed by the elevated expression of alpha(v)beta(3) receptors. Activation and increased expression of alpha(v)beta(3) within SDF-1-rich organs may participate in metastatic localization.

AIMS: Coxsackie and adenovirus receptor (CAR) is a tumor suppressor and a primary receptor for adenovirus type 5 (Ad5). Our study aims to examine the influence of forced expression of CAR in rhabdomyosarcoma cells (RD) on expression levels of integrins implicated in Ad5 entry, and the effect of CAR on cell-extracellular matrix adhesion and migration. MAIN METHODS: CAR expressing clones were established from RD cells by stable transfection. Flow cytometry was used to evaluate the expression of CAR and integrins. Adhesion was measured in plates previously coated with vitronectin or fibronectin. Boyden chambers were used to investigate migration. Transfection of cells with siRNA was used to achieve integrin silencing. Ad5-mediated transgene expression was measured by beta-gal staining. KEY FINDINGS: Increased expression of CAR in RD cells reduces the expression of alphavbeta3 and alphavbeta5 integrins. Cells overexpressing CAR exhibit significantly reduced adhesion to vitronectin and fibronectin, and reduced cell migration. Specifically silencing alphavbeta3 integrin in RD cells reduced cell migration indicating that reduced migration could be the consequence of alphavbeta3 integrin downregulation. This study also demonstrates the negative effect of reduced levels of alphavbeta3 and alphavbeta5 integrins on Ad5-mediated transgene expression with Ad5 retargeted to alphav integrins. SIGNIFICANCE: The pharmacological upregulation of CAR aimed to increase Ad5-mediated transgene expression may actually downregulate alphavbeta3 and alphavbeta5 integrins and thus alter Ad5-mediated gene transfer. The mechanism of decreased cell migration, a prerequisite for metastasis and invasion, due to increased CAR expression may be explained by reduced alphavbeta3 integrin expression.

The mechanisms leading to squamous cell carcinoma of the head and neck (SCCHN) metastasis are incompletely understood. Although evidence shows that the chemokine receptor 7 (CCR7) and its ligand CCL19 may regulate tumor dissemination, their role in SCCHN is not clearly defined. CCR7 has been shown to regulate integrins, which facilitate adhesion of cancer cells to and/or migration through the extracellular matrix (ECM). To investigate the relationship between CCR7 and integrin alphavbeta3 in metastatic SCCHN, we used adhesion and migration assays, immunofluorescence staining and western blotting to determine whether integrin alphavbeta3 can be activated by CCL19 in the metastatic SCCHN cell line PCI-37B, which was pre-incubated with CCL19 or the integrin alphavbeta3 inhibitor, IS201. Our results demonstrate that CCR7 favors PCI-37B cell adhesion and migration, induces reorganization of the actin cytoskeleton and induces integrin alphavbeta3 phosphorylation. The integrin alphavbeta3 inhibitor, IS201, blocked all of these effects. CCR7 and integrin alphavbeta3 expression significantly and positively correlated with tumor size, clinical stage and nodal metastasis. Taken together, our data indicate that CCR7 regulates cell adhesion and migration via integrin alphavbeta3 in metastatic SCCHN. These results should provide the groundwork for new strategies aimed at preventing SCCHN metastasis.