High Metastatic Cancer Research Articles

Abstract B1-63: Prostate cancer classification using a transcriptome atlas

Abstract Background: Gene expression profiling has been used to define subclasses of cancers and in the assembly of potential prognostic signatures. Molecular categories are commonly defined by supervised or unsupervised cluster analysis of mRNA levels. However, despite efforts to profile prostate cancer using transcriptome data, the genetic alterations and biological processes that contribute to clinical progression have made disease subtype categorization difficult. In addition, most studies are limited in statistical power, and therefore do not accurately reflect the heterogeneity of the disease in the general population. The goal of this study was to generate molecular profiles of prostate cancer using an unprecedentedly large transcriptome dataset. Methods: We compiled a very large compendium of 50 human prostate cancer transcriptome datasets containing over 4,000 human prostate cancer specimens, which we named the “Prostate Cancer Transcriptome Atlas” (PCTA). From this resource, 38 datasets, containing over 2,000 specimens, were combined into a single, normalized and batch-corrected dataset using the median-centering method. Molecular categories were sought using 29 published gene expression signature profiles, including castration-resistance (CRPC), oncogene activation, AR activation, AR variants, EZH2, FOXA1, TMPRSS2–ERG fusion, stemness, PTEN inactivation, polycomb complex (PRC2) repression, cell proliferation, epithelial-mesenchymal transition, and proneural and neuroendocrine differentiation. Signature profiles were computed by the weighted Z-score method. Results: Unsupervised clustering identified subsets of tumors manifesting consistent molecular features across many specimens, including a category showing strong AR signatures coupled in concert with significantly repressed PRC2 signatures. These features were mostly seen in high Gleason score (>7) or metastatic prostate cancer. Another major group was an “inverse” signature, showing activated PRC2 and repressed AR signatures. This stratification was independently validated using two datasets: (1) from 545 formalin-fixed paraffin-embedded (FFPE) tissue samples from primary prostate cancer from the Mayo Clinic (PLoS One 2013; 8(6):e66855) and 281 prostate cancers from a watchful-waiting cohort recruited in Sweden (BMC Med Genomics 2010;3:8). This analysis also allowed us to observe other, distinct variant classes, including activation of AR-variants and EZH2, high enrichment of stemness, and proneural and neuroendocrine differentiation. Conclusion: These results show that analyzing gene signatures using an integrated collection of transcriptome profiles from multiple platforms, and thereby significantly increasing sample size, allows the assignment of provisional disease categories that are either difficult to observe or not detected even in large profiling studies. Citation Format: Sungyong You, Jayoung Kim, Michael R. Freeman. Prostate cancer classification using a transcriptome atlas. [abstract]. In: Proceedings of the AACR Special Conference on Computational and Systems Biology of Cancer; Feb 8-11 2015; San Francisco, CA. Philadelphia (PA): AACR; Cancer Res 2015;75(22 Suppl 2):Abstract nr B1-63.

Abstract A1-49: Prostate cancer classification using a transcriptome atlas

Abstract Background: Gene expression profiling has been used to define subclasses of cancers and in the assembly of potential prognostic signatures. Molecular categories are commonly defined by supervised or unsupervised cluster analysis of mRNA levels. However, despite efforts to profile prostate cancer using transcriptome data, the genetic alterations and biological processes that contribute to clinical progression have made disease subtype categorization difficult. In addition, most studies are limited in statistical power, and therefore do not accurately reflect the heterogeneity of the disease in the general population. The goal of this study was to generate molecular profiles of prostate cancer using an unprecedentedly large transcriptome dataset. Methods: We compiled a very large compendium of 50 human prostate cancer transcriptome datasets containing over 4,000 human prostate cancer specimens, which we named the “Prostate Cancer Transcriptome Atlas” (PCTA). From this resource, 38 datasets, containing over 2,000 specimens, were combined into a single, normalized and batch-corrected dataset using the median-centering method. Molecular categories were sought using 29 published gene expression signature profiles, including castration-resistance (CRPC), oncogene activation, AR activation, AR variants, EZH2, FOXA1, TMPRSS2–ERG fusion, stemness, PTEN inactivation, polycomb complex (PRC2) repression, cell proliferation, epithelial-mesenchymal transition, and proneural and neuroendocrine differentiation. Signature profiles were computed by the weighted Z-score method. Results: Unsupervised clustering identified subsets of tumors manifesting consistent molecular features across many specimens, including a category showing strong AR signatures coupled in concert with significantly repressed PRC2 signatures. These features were mostly seen in high Gleason score (>7) or metastatic prostate cancer. Another major group was an “inverse” signature, showing activated PRC2 and repressed AR signatures. This stratification was independently validated using two datasets: (1) from 545 formalin-fixed paraffin-embedded (FFPE) tissue samples from primary prostate cancer from the Mayo Clinic (PLoS One 2013; 8(6):e66855) and 281 prostate cancers from a watchful-waiting cohort recruited in Sweden (BMC Med Genomics 2010;3:8). This analysis also allowed us to observe other, distinct variant classes, including activation of AR-variants and EZH2, high enrichment of stemness, and proneural and neuroendocrine differentiation. Conclusion: These results show that analyzing gene signatures using an integrated collection of transcriptome profiles from multiple platforms, and thereby significantly increasing sample size, allows the assignment of provisional disease categories that are either difficult to observe or not detected even in large profiling studies. Citation Format: Sungyong You, Jayoung Kim, Michael R. Freeman. Prostate cancer classification using a transcriptome atlas. [abstract]. In: Proceedings of the AACR Special Conference on Translation of the Cancer Genome; Feb 7-9, 2015; San Francisco, CA. Philadelphia (PA): AACR; Cancer Res 2015;75(22 Suppl 1):Abstract nr A1-49.

Prostate Cancer Heterogeneous High-Metastatic Multi-Organ-Colonizing Chemo-Resistant Variants Selected by Serial Metastatic Passage in Nude Mice Are Highly Enriched for Multinucleate Giant Cells.

In order to further understand the role of tumor heterogeneity in metastasis and chemo-resistance, high metastatic PC-3 human prostate cancer variants were selected by injecting parental PC-3 cells, expressing green fluorescent protein (GFP) in the footpad of nude mice, which then metastasize to inguinal lymph nodes. The PC-3-GFP cells which metastasized to the inguinal lymph nodes were collected and were re-injected to the footpad. After 6 such cycles, the PC-3-GFP cells collected from inguinal lymph nodes (PC-3-GFP-LN) were again injected to the footpad. PC-3-GFP-LN showed 100% metastasis to major lymph nodes (popliteal, inguinal, axillary, and cervical), and 100% metastasis to bone and lung. The percent of giant cell variants was enriched in PC-3-GFP-LN-6 compared to parental cells and increased with each cycle of selection, which in turn had increased metastasis. PC-3-GFP-LN-6 cells were resistant to 5-fluorouracil, doxorubicin and cisplatinum, compared to parental PC-3. However, PC-3-GFP-LN-6 was sensitive to the traditional Chinese medicine (TCM) herbal mixture LQ, similar to the parental cells. These results suggest that PC-3 tumors are heterogenous and that subpopulations of highly metastatic, drug-resistant cells can be step-wise selected using a mouse model of tumor progression.

Grifolin directly targets ERK1/2 to epigenetically suppress cancer cell metastasis.

Grifolin, a secondary metabolite isolated from the fresh fruiting bodies of the mushroom Albatrellus confluens, has been reported by us and others to display potent antitumor effects. However, the molecular target of grifolin has not been identified and the underlying mechanism of action is not fully understood. Here, we report that the ERK1/2 protein kinases are direct molecular targets of grifolin. Molecular modeling, affinity chromatography and fluorescence quenching analyses showed that grifolin directly binds to ERK1/2. And in vitro and ex vivo kinase assay data further demonstrated that grifolin inhibited the kinase activities of ERK1/2. We found that grifolin suppressed adhesion, migration and invasion of high-metastatic cancer cells. The inhibitory effect of grifolin against tumor metastasis was further confirmed in a metastatic mouse model. We found that grifolin decreased phosphorylation of Elk1 at Ser383, and the protein as well as the mRNA level of DNMT1 was also down-regulated. By luciferase reporter and ChIP assay analyses, we confirmed that grifolin inhibited the transcription activity of Elk1 as well as its binding to the dnmt1 promoter region. Moreover, we report that significant increases in the mRNA levels of Timp2 and pten were induced by grifolin. Thus, our data suggest that grifolin exerts its anti-tumor activity by epigenetic reactivation of metastasis inhibitory-related genes through ERK1/2-Elk1-DNMT1 signaling. Grifolin may represent a promising therapeutic lead compound for intervention of cancer metastasis, and it may also be useful as an ERK1/2 kinase inhibitor as well as an epigenetic agent to further our understanding of DNMT1 function.

The role of miR-205 in the VEGF-mediated promotion of human ovarian cancer cell invasion

ObjectiveOur objective was to investigate a miRNA pathway that acts downstream of VEGF-induced invasion of ovarian cancer cells. MethodWe used two paired high and low metastatic serous ovarian cancer cells to demonstrate the role of miR-205 in VEGF-induced invasion of ovarian cancer cells and to investigate the gene targets of miR-205. ResultsOur previous comparative proteomics studies showed that VEGF decreased the expression of Ezrin and Lamin A/C, and this result was validated in the present study using qPCR and Western blotting. Then we found that VEGF enhanced the invasiveness of and inhibited apoptosis in ovarian cancer cells as assessed by transwell invasion assays and Annexin V-FITC immunostaining, respectively. VEGFR was also expressed in ovarian cancer cells, as assessed by immunocytochemical staining. Furthermore, using the dual-luciferase report assay system, we demonstrated that miR-205 targeted Ezrin and Lamin A/C. MiR-205 was up-regulated in ovarian cancer cells exposed to VEGF, as determined by miRNA microarray analysis and verified by qPCR. MiR-205 promoted the invasion and proliferation of ovarian cancer cells. ConclusionOur data reveal a new potential pathway in which VEGF promotes the invasion of ovarian cancer cells, partially via the down-regulation of Ezrin and Lamin A/C caused by increased expression of miR-205.

Suppression of hepatocellular carcinoma invasion and metastasis by Rho-kinase inhibitor Fasudil through inhibition of BTBD7-ROCK2 signaling pathway

To explore the eff ect of Fasudil on the invasion and metastatic abilities of human high metastatic liver cancer cells (HCCLM3) and the underlying mechanisms. HCCLM3 cells were incubated with 100 μmol/L Fasudil. Fluorescence staining for F-actin and Transwell assay were performed to observe the invasion ability of HCCLM3 cells. HCCLM3 cells were divided into 3 groups: a negative control group, a Fasudil group and a BTB/ POZ domain containing 7 (BTBD7)-siRNA group. Western blot assay was performed to detect the expression levels of BTBD7, ras homolog family member C (RhoC) and Rho-associated, coiled-coil containing protein kinase 2 (ROCK2), matrix metalloproteinases 2 (MMP2) and MMP9. Zymogram analysis method was performed to detect the expression activities of MMP2 and MMP9. The BTBD7-siRNA group was served as a positive control. In HCCLM3 cells treated with Fasudil, the invasion ability was significant decreased compared with the control group, concomitant with the down-regulated expression levels of BTBD7, RhoC and ROCK2 protein as well as the decreased activities of MMP2 and MMP9. Fasudil plays an important role in interfering BTBD7-ROCK2 signaling pathway and suppressing the invasion and metastasis of hepatocellular carcinoma.

Highlights from Recent Cancer Literature

Highlights from Recent Cancer Literature

Editorial introductions

Current Opinion in Oncology was launched in 1989. It is one of a successful series of review journals whose unique format is designed to provide a systematic and critical assessment of the literature as presented in the many primary journals. The field of psychiatry is divided into 15 sections that are reviewed once a year. Each section is assigned a Section Editor, a leading authority in the area, who identifies the most important topics at that time. Here we are pleased to introduce the Journal's Section Editors for this issue. SECTION EDITORS Gilberto de Castro Jr.Gilberto de Castro Jr.Gilberto de Castro Jr. received his MD degree from the University of São Paulo Medical School, Brazil in 1995, and completed his training in Internal Medicine and Clinical Oncology in 2000 at Hospital das Clínicas, University of São Paulo Medical School, where he also received his PhD degree in 2009. He is an ex fellow of the Medical Oncology Clinic at the Institut Jules Bordet, Brussels, Belgium. His current position is Chief at the Unit of Head and Neck and Thoracic Tumors of the Clinical Oncology Service of the Instituto do Câncer do Estado de São Paulo, Brazil - the largest cancer center in the South America. His areas of interest are the medical management of head and neck cancer and thoracic malignancies, focusing on the development of molecular targeted therapies, predictive factors of response to chemotherapy and quality of life studies. Dr Castro has more than 35 publications in peer-reviewed journals and he is also involved in cancer education, being a member of the European School of Oncology team of Discussants (e-ESO). Arif HussainArif HussainArif Hussain, MD is Professor of Medicine with Tenure in the Department of Medicine, School of Medicine, at the University of Maryland in Baltimore, USA, where he is also Professor of Biochemistry in the Department of Biological Chemistry and Professor of Pathology in the Department of Pathology. He also serves as Co-Leader of the Hormone Responsive Cancers Program in the Greenebaum Cancer Center, University of Maryland. Dr Hussain's clinical interest is in genitourinary cancers, and he serves as Principal Investigator on several clinical trials that focus on therapies for high risk, recurrent and metastatic prostate cancer, as well as renal cell cancer. In addition, he has a research laboratory focusing on drug resistance, intracellular calcium pumps, and pre-clinical models of prostate cancer. Ahmad AwadaAhmad AwadaProfessor Ahmad Awada studied Medicine at the Free University in Brussels (ULB), Belgium. He did a specialisation in Internal Medicine and Medical Oncology at Jules Bordet Institute (under the supervision of Professor Jean Klastersky), in Brussels, until 1992 (“La plus grande distinction”). During his specialisation, he also followed trainings in the clinical development of new therapies and new anticancer drugs, under the supervision of Professor Martine Piccart. To deepen his training, he stayed in the Netherlands (New Drug Development Office, Free University, Amsterdam) and in San Antonio, USA (Institute for Drug Development, under the direction of Professor D. Von Hoff). He focused on the clinical development of new anticancer agents. Back from the USA at the beginning of 1994, Doctor Awada became Assistant Head of Medical Oncology Clinic, and Head of the New Drugs Development Unit at Jules Bordet Institute, Brussels. Since April 2005, he is the Head of the Medical Oncology Clinic. In addition and from 1st March 2011, Dr Awada was hold Associate Head of Medicine Department. He has an important clinical activity in the treatment of solid tumors. Doctor Awada took an active part in the development of new anticancer drugs, some of them already widely used. Thanks to the recent progress in technology and computer sciences, and consequently in molecular biology, we can understand better now how a cancer cell works. Many drugs are now studied to block this process. Those are the molecular-targeted therapies, a subject studied extensively in clinical research by Doctor Awada and were the basis for this thesis obtained from the ULB, Brussels, in 2004. The aim of Doctor Awada, as far as research is concerned, is to look for new active anticancer drugs in solid tumors (new cytotoxics and molecular targeted therapies), and to individualize the treatments of cancer patients.

Editorial introductions

Current Opinion in Oncology was launched in 1989. It is one of a successful series of review journals whose unique format is designed to provide a systematic and critical assessment of the literature as presented in the many primary journals. The field of psychiatry is divided into 15 sections that are reviewed once a year. Each section is assigned a Section Editor, a leading authority in the area, who identifies the most important topics at that time. Here we are pleased to introduce the Journal's Section Editors for this issue. SECTION EDITORS Gilberto de Castro Jr.Gilberto de Castro Jr.Gilberto de Castro Jr. received his MD degree from the University of Sao Paulo Medical School, Brazil in 1995, and completed his training in Internal Medicine and Clinical Oncology in 2000 at Hospital das Clínicas, University of Sao Paulo Medical School, where he also received his PhD degree in 2009. He is an ex fellow of the Medical Oncology Clinic at the Institut Jules Bordet, Brussels, Belgium. His current position is Chief at the Unit of Head and Neck and Thoracic Tumors of the Clinical Oncology Service of the Instituto do Cancer do Estado de Sao Paulo, Brazil - the largest cancer center in the South America. His areas of interest are the medical management of head and neck cancer and thoracic malignancies, focusing on the development of molecular targeted therapies, predictive factors of response to chemotherapy and quality of life studies. Dr Castro has more than 35 publications in peer-reviewed journals and he is also involved in cancer education, being a member of the European School of Oncology team of Discussants (e-ESO). Arif HussainArif HussainArif Hussain, MD is Professor of Medicine with Tenure in the Department of Medicine, School of Medicine, at the University of Maryland in Baltimore, USA, where he is also Professor of Biochemistry in the Department of Biological Chemistry and Professor of Pathology in the Department of Pathology. He also serves as Co-Leader of the Hormone Responsive Cancers Program in the Greenebaum Cancer Center, University of Maryland. Dr Hussain's clinical interest is in genitourinary cancers, and he serves as Principal Investigator on several clinical trials that focus on therapies for high risk, recurrent and metastatic prostate cancer, as well as renal cell cancer. In addition, he has a research laboratory focusing on drug resistance, intracellular calcium pumps, and pre-clinical models of prostate cancer. Ahmad AwadaAhmad AwadaProfessor Ahmad Awada studied Medicine at the Free University in Brussels (ULB), Belgium. He did a specialisation in Internal Medicine and Medical Oncology at Jules Bordet Institute (under the supervision of Professor Jean Klastersky), in Brussels, until 1992 (“La plus grande distinction”). During his specialisation, he also followed trainings in the clinical development of new therapies and new anticancer drugs, under the supervision of Professor Martine Piccart. To deepen his training, he stayed in the Netherlands (New Drug Development Office, Free University, Amsterdam) and in San Antonio, USA (Institute for Drug Development, under the direction of Professor D. Von Hoff). He focused on the clinical development of new anticancer agents. Back from the USA at the beginning of 1994, Doctor Awada became Assistant Head of Medical Oncology Clinic, and Head of the New Drugs Development Unit at Jules Bordet Institute, Brussels. Since April 2005, he is the Head of the Medical Oncology Clinic. In addition and from 1st March 2011, Dr Awada was hold Associate Head of Medicine Department. He has an important clinical activity in the treatment of solid tumors. Doctor Awada took an active part in the development of new anticancer drugs, some of them already widely used. Thanks to the recent progress in technology and computer sciences, and consequently in molecular biology, we can understand better now how a cancer cell works. Many drugs are now studied to block this process. Those are the molecular-targeted therapies, a subject studied extensively in clinical research by Doctor Awada and were the basis for this thesis obtained from the ULB, Brussels, in 2004. The aim of Doctor Awada, as far as research is concerned, is to look for new active anticancer drugs in solid tumors (new cytotoxics and molecular targeted therapies), and to individualize the treatments of cancer patients.

Abstract 410: Elevated Jagged-1 and Notch-1 expression in high grade and metastatic prostate cancers.

Abstract Background: Emerging evidence has suggested that Notch signaling pathway may be involved in the development, progression and metastasis of prostate cancer (PCa). In the present study, we investigated the expression levels of Jagged-1 and Notch-1 in human prostate tumors and their associations with prostate cancer progression and metastasis. Methods: Immunohistochemistry (IHC) for Jagged-1 and Notch-1 was performed on tissue microarray (TMA) slides containing 286 formalin-fixed and paraffin-embedded tissue specimens with various prostatic disorders, including benign changes, high grade prostatic intraepithelial neoplasia (HGPIN), low and high grade prostate cancers as well as metastatic prostate cancers. The expression levels of Jagged-1 and Notch-1 were reported as a final IHC score calculated as staining extent score (0-3) multiplied by intensity score (0-3); with a maximal score equal to 9. ANOVA and t-test were employed to analyze the difference of IHC scores among and between different pathologic categories Results: Jagged-1 cytoplasmic IHC scores in both metastatic PCa (5.92 ± 2.12, Mean ± SD) and high grade PCa (5.74 ± 2.45) were significantly higher than those in low grade PCa (3.55 ± 2.02, p = 9.02E-07 and 4.65E-08, respectively), in HGPIN (4.50 ± 2.34, p = 0.023 and 0.030, respectively), and in prostatic tissues with benign changes (3.39 ± 1.85, p = 7.96E-07 and 2.47E-07, respectively). In addition, Jagged-1 membranous IHC scores in both metastatic PCa (3.05 ± 3.52) and high grade PCa (2.41 ± 2.90) were also significantly higher than those in low grade PCa (0.71 ± 1.42, p = 1.48E-05 and 3.40E-06, respectively) and in prostatic tissues with benign changes (0.82 ± 1.28, p = 3.19E-04 and 0.001, respectively). Similarly, Notch-1 cytoplasmic IHC scores in both metastatic PCa (2.22 ± 2.11) and high grade PCa (1.10 ± 1.68) were significantly elevated when compared with the IHC scores observed in low grade PCa (0.51 ± 1.15, p = 3.63E-06 and 0.023, respectively) and in prostate tissues with benign changes (0.48 ± 1.01, p = 3.37E-06 and 0.017, respectively). Furthermore, significantly more highly expressed Jagged-1 in membrane was observed in Caucasians (vs. African Americans) and in positive surgical resection margin group (vs. negative surgical resection margin group). Conclusions: Our results provide strong evidence that dysregulation of Jagged 1-Notch1 signaling plays a role in PCa progression and metastasis and suggest that Jagged-1 and Notch-1 may be useful markers in distinguishing indolent and aggressive prostate cancers. Citation Format: He Zhu, Xinchun Zhou, Samantha Redfield, Jack Lewin, Lucio Miele. Elevated Jagged-1 and Notch-1 expression in high grade and metastatic prostate cancers. [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 410. doi:10.1158/1538-7445.AM2013-410

Short hairpin RNA targeting FOXQ1 inhibits invasion and metastasis via the reversal of epithelialmesenchymal transition in bladder cancer

The epithelial-mesenchymal transition (EMT) promotes cancer invasion and metastasis, however, the integrative mechanisms that coordinate the process are incompletely understood. In this study, we defined a pivotal functional role for the Forkhead transcription factor FOXQ1 in regulating EMT in bladder cancer. We initially investigated the expression of FOXQ1, TGF-β1 and EMT biomarkers E-cadherin, Vimentin in 65 cases of bladder transitional cell carcinoma (BTCC) specimens by reverse transcription-polymerase chain reaction (RT-PCR), western blot analysis and immunohistochemistry. Search results indicated that FOXQ1 expression was inversely correlated to E-cadherin, but positively to TGF-β1 and Vimentin in patients with BTCC (P<0.05). Furthermore, we aimed to construct short hairpin RNA (shRNA) expression plasmids against the FOXQ1 gene and transfect shRNAs into high metastatic potential human bladder cancer T24 cells with Lipofectamine 2000. RNAi-mediated suppression of FOXQ1 expression reversed the EMT process accompanied by upregulation of E-cadherin, as well as a loss expression of Vimentin in highly invasive T24 cells (P<0.05). The inhibition of FOXQ1 expression with shRNA vector also led T24 cells to acquire an epithelial cobblestone phenotype, significantly reduced motility and subsequent invasiveness of bladder cancer cells (P<0.05). In conclusion that FOXQ1 may be a novel EMT-inducing transcription factor through controlling the expression of E-cadherin and aggressiveness of cancer cells and targeting the transcription factor FOXQ1 could hence serve as a novel therapeutic strategy for cancer patients.

Expression of metadherin/AEG-1 gene is positively related to orientation chemotaxis and adhesion of human hepatocellular carcinoma cell lines of different metastatic potentials

Metastasis contributes to the poor prognosis of hepatocellular carcinoma (HCC). However, the mechanism through which a primary HCC cell develops into a metastatic phenotype is not well understood. The purpose of this study was to examine the correlation between metadherin (MTDH)/astrocyte elevated gene-1 (AEG-1) expression in HCC cell lines of different metastatic potentials and such metastatic phenotypes as orientation chemotaxis and adhesion. MTDH/AEG-1 expression was detected by RT-PCR and western blotting in HCC cell lines (HepG2, Huh7, Sk-HEP-1, MHCC-97H). Distribution of MTDH/AEG-1 was observed by immunofluorescence staining and confocal laser scanning microscopy. The abilities of orientation chemotaxis and adhesion and the index of interaction between HCC cell lines and microvascular endothelial cell lines (MVECs, including HUVECs and HPMECs) were measured by chemotaxis assay and adhesion assay, respectively. The results showed that MTDH/AEG-1 protein expression was significantly higher in high metastatic potential cancer cell lines (Sk-HEP-1, MHCC-97H) than in low metastatic potential cell lines (HepG2, Huh7) (P<0.05). The MTDH/AEG-1 protein was localized in the perinuclear region of HCC cells. Furthermore, the abilities of orientation chemotaxis and adhesion of HCC cells to HPMECs were increased as compared with those of HCC cells to HUVECs (P<0.05). The abilities of orientation chemotaxis and adhesion were much stronger in Sk-HEP-1 and MHCC-97H cells with MTDH/AEG-1 highly expressed than in HepG2 and Huh7 cells with MTDH/AEG-1 lowly expressed (P<0.05). These results suggested that the expression of MTDH/AEG-1 gene in HCC cell lines of different metastatic potentials was closely positively related to the abilities of orientation chemotaxis and adhesion of HCC cells. It was deduced that MTDH/AEG-1 might play a pivotal role in the lung-specific metastasis of HCC, which may be mediated through orientation chemotaxis and adhesion abilities of HCC cells. MTDH/AEG-1 may serve as a potential therapeutic target for HCC.

Editorial introductions

Current Opinion in Oncology was launched in 1989. It is one of a successful series of review journals whose unique format is designed to provide a systematic and critical assessment of the literature as presented in the many primary journals. The field of psychiatry is divided into 15 sections that are reviewed once a year. Each section is assigned a Section Editor, a leading authority in the area, who identifies the most important topics at that time. Here we are pleased to introduce the Journal's Section Editors for this issue. SECTION EDITORS Gilberto de CastroGilberto de CastroGilberto de Castro Jr. received his MD degree from the University of Sao Paulo Medical School, Brazil in 1995, and completed his training in Internal Medicine and Clinical Oncology in 2000 at Hospital das Clínicas, University of Sao Paulo Medical School, where he also received his PhD degree in 2009. He is an ex fellow of the Medical Oncology Clinic at the Institut Jules Bordet, Brussels, Belgium. His current position is Chief at the Unit of Head and Neck and Thoracic Tumors of the Clinical Oncology Service of the Instituto do Cancer do Estado de Sao Paulo, Brazil - the largest cancer center in the South America. His areas of interest are the medical management of head and neck cancer and thoracic malignancies, focusing on the development of molecular targeted therapies, predictive factors of response to chemotherapy and quality of life studies. Dr Castro has more than 25 publications in peer-reviewed journals and he is also involved in cancer education, being a member of the European School of Oncology team of Discussants (e-ESO). Arif HussainArif HussainArif Hussain, MD is Professor of Medicine with Tenure in the Department of Medicine, School of Medicine, at the University of Maryland in Baltimore, USA, where he is also Professor of Biochemistry in the Department of Biological Chemistry and Professor of Pathology in the Department of Pathology. He also serves as Co-Leader of the Hormone Responsive Cancers Program in the Greenebaum Cancer Center, University of Maryland. Dr Hussain's clinical interest is in genitourinary cancers, and he serves as Principal Investigator on several clinical trials that focus on therapies for high risk, recurrent and metastatic prostate cancer, as well as renal cell cancer. In addition, he has a research laboratory focusing on drug resistance, intracellular calcium pumps, and pre-clinical models of prostate cancer. Ahmad AwadaAhmad AwadaProfessor Ahmad Awada was born in Lebanon and studied Medicine at the Free University in Brussels (ULB), Belgium. He did a specialisation in Internal Medicine and Medical Oncology at Jules Bordet Institute (under the supervision of Professor Jean Klastersky), in Brussels, until 1992 (“La plus grande distinction”). During his specialisation, he completed training in the clinical development of new therapies and new anticancer drugs, under the supervision of Professor Martine Piccart. To further his training, he stayed in the Netherlands (New Drug Development Office, Free University, Amsterdam) and in San Antonio, USA (Institute for Drug Development, under the direction of Professor D. Von Hoff). He focused on the clinical development of new anticancer agents. Back from the USA at the beginning of 1994, Professor Awada became Assistant Head of Medical Oncology Clinic, and Head of the New Drugs Development Unit at Jules Bordet Institute, Brussels. Since April 2005, he is the Head of the Medical Oncology Clinic. He has an important clinical activity in the treatment of solid tumors and in particular head & neck cancers. Doctor Awada took an active part in the development of new drugs, some of them already widely used. Thanks to the recent progress in technology and computer sciences, and consequently in molecular biology, we can understand better now how a cancer cell works and how it becomes a cancer cell. Many drugs are now studied to block this process. Those are the molecular-targeted therapies, a subject studied extensively in clinical research by Professor Awada and were the basis for his thesis obtained from the ULB, Brussels, in 2004. Professor Awada's research aims to look for new active anticancer drugs in solid tumors (new cytotoxics and molecular targeted therapies), and to individualize the treatments according to the tumor characteristics.

Identification and chromosomal localizations of signal transduction genes associated with human ovarian cancer metastasis

Gene chip technology can be used to identify and localize signal transduction genes associated with metastasis. We used the human genome U133A gene chip to detect differences in gene expression profiles among high (H) and low (L) metastatic human ovarian cancer cell lines (HO-8910PM, HO-8910), and normal ovarian tissues (C), to identify metastasis-associated signal transduction genes and determine their chromosomal localizations. A total of 37 signal transduction genes showed more than twofold differences in expression levels between the H and L metastatic ovarian cancer cell lines; of these, 21 genes were up-regulated [signal log ratio (SLR)≥1], and 16 genes were down-regulated (SLR≤-1). Most genes were located on chromosome 1 (7 genes, 18.9%), followed by chromosome 8 (5 genes, 13.5%), then chromosomes 6, 11, and 17 (3 genes each, 8.1%). A total of 21 of the differentially expressed genes (56.7%) were localized on the short arm of the chromosome (q). The disruption of signal transduction gene expression may be an important factor associated with ovarian cancer metastasis. The affected signal transduction genes were localized to chromosomes 1, 8, 6, 11, and 17.

Effect of AA861 combined with celecoxib on metastatic colon carcinoma cell

Objective To investigate the effects of cyclooxygenase-2 (COX-2) selective inhibitor combined with 5-lipoxygenase (5-LOX) inhibitor docebenone (AA861) on cell proliferation and migration of human colon cancer cell line HT-29. Methods Cultured in vitro of HT-29 cells in high metastatic colon cancer, A A861 and celecoxib on colon cancer cell drugs for 48 h, the cell proliferation was assayed by MTT; the migration of cell was detected by Transwell; immunofluorescence staining of intracellular changes in ICAM-1 protein, RT-PCR detect the gene expression of ICAM-1 and VEGF. Results MTT and Transwell experiments showed that the inhibition on celecoxib and AA861 on colon cancer was dose-dependent and time-dependent. And the inhibition of AA861 and celecoxib 100 μmol/L alone on colon cancer cells proliferation were 43.2 % and 42.8%, and when the two drugs 100 μmol/L combined on colon cancer cells the inhibition was 53.8%, and the difference between alone group and combined group was statistical (P ＜0.001). The inhibition of AA861 and celecoxib 100 μmol/L alone on colon cancer cells migration were 32.0 % and 29.3%, and when the two drugs 100 μ,mol/L combined on colon cancer cells the inhibition is 57.8 %, and the difference of between alone group and combined group was statistical (P ＜0.001). Immunefluorescence staining and RT-PCR results suggested that the two drugs can inhibit ICAM-1 and VEGF protein and gene expression, and when the two drugs combined, a stronger inhibition effect appeared than used alone (P＜0.001). Conclusion Low-dose celecoxib and AA861 combined has a synergistic inhibited effect on colon cancer cells invasion and metastasis, and the mechanism relates with the VEGF and ICAM-1 expression. Key words: Neoplasms, experimental; Colonic neoplasms; Drug therapy, combination; Neoplasms metastasis

Secretory leukocyte protease inhibitor is associated with MMP-2 and MMP-9 to promote migration and invasion in SNU638 gastric cancer cells

Secretory leukocyte protease inhibitor (SLPI) protects tissue from proteases, and promotes cell proliferation and healing during inflammatory response. SLPI is also overexpressed in gastric, lung and ovarian cancers, which accelerates the metastasis of cancer cells. Matrix metalloproteinases-2, -9 (MMP-2 and MMP-9) are overexpressed in high metastatic cancers, and promote the migration of cancer cells through collagen degradation. SLPI and MMP-2, -9 are critical factors in stimulating the metastatic processes but there are no reports of a direct correlation between these molecules. Therefore, this study examined the role of SLPI related to MMP-2 and MMP-9 using two gastric cancer cell lines, such as characterized non-metastatic SNU484 and highly metastatic SNU638 cells. SLPI, MMP-2 and MMP-9 mRNA and protein expression were higher in SNU638 cells than in SNU484 cells. In addition, the rate of cell migration and invasion was higher in the SNU638 cells than in SNU484 cells. Interestingly, after treatment with SLPI, the rate of migration and invasion was higher in the SNU484 cells than in the positive control (PC) SNU484 cells. The rate of migration was also higher in the SNU638 cells after SLPI treatment than in the SNU638 cells (PC) but the invasion rate was not changed. The expression and secretion of MMP-2 and MMP-9 as well the rate of cell migration and invasion were significantly lower in SLPI-siRNA transfected SNU638 cells (si-SLPI/SNU638) but higher in SLPI-treated SNU484 cells (SNU484 + SLPI). Strong Elk-1 phosphorylation was detected in SNU484 + SLPI and SNU638 cells but was barely detectable in SNU484 and si-SLPI/SNU638 cells. These results show that SLPI promotes the metastasis of SNU638 gastric cancer cells by increasing MMP-2 and MMP-9 expression through Elk-1 signaling, indicating its role as a signaling molecule not a protease inhibitor.

An inhibitory effect of miR-22 on cell migration and invasion in ovarian cancer

Objectives Aberrant expression of microRNAs (miRNAs) has been implicated in ovarian carcinoma. However, roles of miRNAs in ovarian caner metastasis have not been comprehensively addressed. This work is aimed to identify selected miRNAs involved in ovarian cancer metastasis. Methods We examined the distinct miRNA expression profiles between paired high-metastatic human serous ovarian cancer cell SKOV-3ip and low-metastatic human serous ovarian cell SKOV-3 using miRNA microarray. Subsequently, a validation with Real-time RT-PCR was performed for miR-22 expression level, and a functional study was carried out for miR-22. Results Through a screen with microarray, we found there were a variety of miRNAs differentially expressed between paired high and low metastatic serous ovarian cancer cells. Particularly, miR-22 was identified as a potential metastasis-inhibitor in ovarian cancer. There was a negative correlation between miR-22 expression and the metastatic potential in ovarian cancer cells. Furthermore, both gain-of-function and loss-of-function studies displayed an inhibitory effect of miR-22 on cell migration and invasion in vitro without significantly affecting cell viability and apoptosis. Subsequent bioinformatics analysis revealed that miR-22 might regulate multiple pro-metastatic genes, which could provide an explanation to the inhibitory effects of miR-22 on cell migration and invasion. Conclusions Taken together, our findings suggested that miR-22 might be involved in inhibiting ovarian cancer metastasis.

Tissue invasive macrophage density is correlated with prognosis in cholangiocarcinoma

Cholangiocarcinoma (CCA) is a high metastatic cancer with no effective treatment. Here, the pro-metastatic action of tissue macrophages in CCA is demonstrated and suggested as a prognostic marker and novel target for the therapeutic intervention of CCA. Fifty CCA tissues were immunohistochemically stained with a marker for reactive/infiltrating monocytes/macrophages (MAC387) and matrix metalloproteinase (MMP)-9. The antigenic densities in positively-stained cells along the leading edge of tumors were scored. Correlations between the densities of MAC387, MMP-9-positive cells, clinicopathological features and patient survival were investigated. High densities of MAC387-positive cells were detected in more than 60% of the CCA tissues. This was significantly associated with poor prognosis parameters (non-papillary and mass-forming type CCA). Overall survival was worst in patients with high-density MAC387-positive cells. Double immunofluorescent staining indicated that MAC387-positive cells co-expressed MMP-9. Immunohistochemical staining of MMP-9 in serial sections of CCA tissues indicated that MMP-9 was rarely expressed in CCA tumor cells, but highly expressed in MAC387-positive cells and polymorphonucleated infiltrating cells. Patients with high tissue expression levels of MAC387 in combination with MMP-9-expressing cells had the worst survival. These factors were found to be independent predictors of the post-resectional survival of CCA patients. Since CCA tumor cells rarely expressed MMP-9, it is likely that tissue macrophages are critical for degrading the extracellular matrix and for facilitating tumor metastasis. They may therefore serve as a prognostic marker for poor clinical outcome, and represent novel targets for the therapeutic intervention of CCA.

Abstract 3295: The type I insulin-like growth factor receptor (IGF1R) pathway driven metastasis signature correlates with poor prognosis and decreased distant metastasis-free survival in human breast cancer

Abstract The insulin-like growth factors (IGFs) acting via the type I IGF receptor (IGF1R) regulate growth and metastasis of multiple types of cancers. Agents targeting IGF1R are currently being tested in phase II/III clinical trials. We have previously shown that IGF1R can regulate metastasis independently of primary tumor growth. In a model of high risk metastatic cancer using the MDA435/LCC6 metastatic cancer cells, we have shown that disruption of IGF signaling either with an antibody against IGF1R, AVE1642, or a dominant negative IGF-1R construct, does not affect growth in the mammary fat pad but inhibits pulmonary metastases and colonization of the lungs. Regulation of such a phenotype will not be measured in clinical trials and these findings imply that clinical trials of this therapeutic strategy may need to adjust their endpoints for assessing benefit by taking into account that inhibition of IGF1R signaling may inhibit metastasis but not primary tumor growth. Here, we sought to develop an IGF1R driven metastasis signature that may be useful in selecting patients that could benefit from IGF1R targeted therapy and determine if changes in expression of the genes in the signature can be useful in monitoring response. Serum starved LCC6-WT cells (with wild-type functional IGF1R) and LCC6-DN (cells with dominant negative IGF1R) were treated with IGF-I, AVE1642 (antibody against IGF1R that inhibits metastasis but not tumor growth in the mammary fat pad), or AVE1642+ IGF-I for 4 and 24 hours and microarray analysis was performed using the U133 Plus 2.0 human arrays with 47,000 transcripts. Array data were analyzed to determine genes regulated by IGF1R signaling in these cells and if disruption of signaling with AVE1642 affected genes whose expression was regulated by IGF-I signaling. A total of 206 transcripts were regulated by IGF signaling in LCC6-WT cells and AVE1642 treatment reversed the effects of IGF1R signaling. 53 of the most significantly regulated genes were used to query two different human breast tumor datasets to determine if genes regulated by activation of IGF1R are associated with lymph node metastasis and poor prognosis in human tumors. IGF1R regulated metastasis signature genes were expressed in human tumors and were correlated to decreased survival and decreased distant metastasis free survival in breast cancer patients. Our results suggest that presence of IGF1R metastasis signature in patients confers prognostic value and identifying IGF1R driven metastasis signatures may be useful to identify and monitor patients who could benefit from IGF1R targeted therapy for inhibition of metastatic disease. 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 3295.

Abstract B239: Profiling the type I insulin-like growth factor receptor (IGF1R) pathway regulated metastasis signature

Abstract The insulin-like growth factors (IGFs) acting via the type I IGF receptor (IGF1R) regulate growth and metastasis of multiple types of cancers. Agents targeting IGF1R are currently being tested in phase II/III clinical trials. We have previously shown that IGF1R can regulate metastasis independently of primary tumor growth in a model of high risk metastatic cancer using the MDA435/LCC6 metastatic cancer cells. Here we sought to develop an IGF1R driven metastasis signature that may be useful in selecting patients that could benefit from IGF1R targeted therapy and determine if changes in expression of the genes in the signature can be useful in predicting response. Serum starved LCC6-WT cells (with wild-type functional IGF1R) and LCC6-DN (cells with dominant negative IGF1R) were treated with IGF-I, AVE1642 (antibody against IGF1R that inhibits metastasis but not tumor growth in the mammary fat pad), or AVE1642+ IGF-I for 4 and 24 hours and microarray analysis was performed using the U133 Plus 2.0 human arrays with 47,000 transcripts. Array data were analyzed to determine genes regulated by IGF1R signaling in these cells and if disruption of signaling with AVE1642 affected genes whose expression was regulated by IGF-I signaling. A total of 91 transcripts were regulated by IGF signaling in LCC6-WT cells and AVE1642 treatment reversed the effects of IGF1R signaling. We are currently querying human tumor databases to determine if genes regulated by activation of IGF1R are associated with lymph node metastasis and poor prognosis in human tumors. Our results suggest that identifying IGF1R driven metastasis signatures may be useful to identify and monitor patients who could benefit from IGF1R targeted therapy for inhibition of metastatic disease and that patients with this IG Citation Information: Mol Cancer Ther 2009;8(12 Suppl):B239.