bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 2619: Whole genome and transcriptome sequencing identifies an activating mutation of LPAR1 in a patient with metastatic neuroblastoma.
Authors: Wei Jun S. , Johansson Peter , Chen Li , Song Young K. , Tolman Catherine , Li Samuel , Hurd Laura , Patidar Rajesh , Wen Xinyu , Badgett Thomas C. , Cheuk Adam , Marshall Jean-Claude , Steeg Patricia , Vaqué Díez José Pedro , Gutkind J Silvio , Khan Javed
Published: August 2014

Neuroblastoma is the most common extra-cranial solid tumor of childhood, and is one of the most genomically heterogeneous malignances studied to date. Despite advance of multimodal therapies, the current overall survival rate for patient with metastatic neuroblastoma is <40%, and the molecular events that drive the disease are not fully understood. To systematically identify expressed driver oncogenes, we performed whole genome and transcriptome sequencing on an index liver metastasis of neuroblastoma from a young adult patient. Additional transcriptome sequencing was performed on tumor cells from bone marrow taken at diagnosis and from the adrenal primary taken at the time of primary surgical resection. Whole genome sequencing of the index tumor identified 44 somatic small variant mutations in 42 genes and a large hemizygous deletion in the ATRX gene which has been recently implicated in neuroblastoma. Of these 45 somatic alterations, 15 genes including ATRX were found commonly mutated in all three tumors and transcriptome sequencing showed 3 of them (LPAR1, GATA2, and NUFIP1) had high level of expression of the mutant alleles, suggesting potential oncogenic driver roles for these mutated genes. Among these commonly mutated expressed genes, the druggable G-protein coupled receptor LPAR1 was expressed at the highest level in all tumors. Cells expressing the LPAR1 mutant resulted in a significant increase in motility, but had no effect on growth. Further biochemical characterization demonstrated that elevated Rho signaling was responsible for the increased cell mobility mediated by the LPAR1 mutant. Therefore, parallel whole genome and transcriptome sequencing identified a cell motility driver mutation in the LPAR1 gene, and this combinatorial approach may be leveraged for identification of expressed driver mutations in cancer patients for precision therapy.

Citation Format: Jun S. Wei, Peter Johansson, Li Chen, Young K. Song, Catherine Tolman, Samuel Li, Laura Hurd, Rajesh Patidar, Xinyu Wen, Thomas C. Badgett, Adam Cheuk, Jean-Claude Marshall, Patricia Steeg, José Pedro Vaqué Díez, J Silvio Gutkind, Javed Khan. Whole genome and transcriptome sequencing identifies an activating mutation of LPAR1 in a patient with metastatic neuroblastoma. [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 2619. doi:10.1158/1538-7445.AM2013-2619
Abstract 263: Identification of subgroup of triple negative breast cancer by cancer stem cell markers.
Authors: Kida Kumiko , Ishikawa Takashi , Yamada Akimitsu , Shimada Kazuhiro , Narui Kazutaka , Sugae Sadatoshi , Shimizu Daisuke , Tanabe Mikiko , Sasaki Takeshi , Ichikawa Yasushi , Endo Itaru
Published: August 2014

[Background]

Anti-cancer agents are essential for treating triple negative breast cancer (TNBC). The cases resistant to them showed unfavorable clinical courses and thus may require novel agents. It is crucial to predict the efficacy of existing agents in TNBC. Recently, cancer stem cell (CSC) theory has drawn an attention in relation to TNBC and chemoresistance. Aldehyde dehydrogenase 1 (ALDH1) is a biomarker of breast cancer stem cell.

[Aims]

In order to subdivide TNBC and finally find subgroup resistant to existing anti-cancer agents and better markers for stem cell phenotype.

[Methods]

1)The expression of ALDH1 was examined immunohistochemically in each subtype of 447 breast cancer patients.

2)The expression of ALDH1 and its association with Ki-67,P53, CK5/6 or EGFR were examined in TNBC.

3)Correlation between ALDH1 and pathological response by neoadjuvant chemotherapy was examined.

4)ALDH1 positive and negative cells were collected by microdissection from 5 ALDH1 positive breast cancer cases. The expression profile was assessed by Affymetrix cDNA microarray. The difference of profile between ALDH-1 positive and negative cells was assessed.

[Results]

1)The expression rate of ALDH1 was 4.5% in Luminal type, 11.1% in Luminal-HER2 type, 23.8% in HER2-enriched type and 33.6% in triple negative type. ALDH1 was expressed highly in TNBC.

2)In 79 TNBC with neoadjuvant chemotherapy, 29 cases(36.7%) was positive for ALDH1.The averages of NG, Ki67 and p53 were 2.7, 37.8% and 37.7% respectively in ALDH1 positive TNBC. 80% of them were positive for CK5/6 or EGFR. There was no significant correlation between ALDH1 and these pathological markers.

3) Poorly responsiveness to NAC (0-1b) was found in 17 cases (21.5%). Only ALDH1 was associated with chemoresistance to NAC (P=0.033), suggesting ALDH1 positive TNBC were resistant to anticancer agents.

4)63 and 41 genes were up- and down-regulated respectively in ALDH1 positive cells. These potential related genes are now under investigation.

[Conclusions]

ALDH1 could be a marker for chemoresistance in TNBC. As ALDH1 identifies some specific subpopulation of breast cancer, further studies will provide important information on subdividing TNBC and individualizing treatment of this intractable subtype.

Citation Format: Kumiko Kida, Takashi Ishikawa, Akimitsu Yamada, Kazuhiro Shimada, Kazutaka Narui, Sadatoshi Sugae, Daisuke Shimizu, Mikiko Tanabe, Takeshi Sasaki, Yasushi Ichikawa, Itaru Endo. Identification of subgroup of triple negative breast cancer by cancer stem cell markers. [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 263. doi:10.1158/1538-7445.AM2013-263
Abstract 2638: The BCAR3/Cas/c-Src signaling node in breast cancer progression.
Authors: Batties Allison M. , Guerrero Michael S. , Wilson Ashley L. , Thomas Keena S. , Bouton Amy H.
Published: August 2014

Despite advances in early detection and improved adjuvant therapies, breast cancer remains the second leading cause of cancer death among women mainly due to metastatic spread of the disease. The ability of cancer cells to migrate and invade is important in the process of metastasis. Studies have shown that the adaptor protein Breast Cancer Antiestrogen Resistance 3 (BCAR3) is highly expressed in invasive breast cancer cell lines and functions as a positive regulator of migration and invasion. To date, BCAR3 has only one clearly established binding partner in breast cancer cells, the adaptor protein, p130Cas (Cas). Studies of human breast tumors have shown a positive correlation between high levels of Cas protein expression and poor clinical outcome in patients (Dorssers LC et al, 2004). Additionally, the protein tyrosine kinase c-Src is a C-terminal binding partner of Cas. c-Src kinase activity promotes enhanced Cas signaling and is associated with breast cancer progression and poor clinical outcome (Blscardi JS et al, 1999). These data suggest a critical role for the BCAR3/Cas/c-Src signaling node in breast tumor progression. We hypothesize that enhanced signaling through the BCAR3/Cas/c-Src network in breast epithelial cells is critical for the transition to an invasive cancer cell phenotype. In this work, we compared the relationship between these proteins in non-tumorigenic mammary epithelial cells (MCF10A) and invasive breast cancer cells (BT549 and MDA-MB-231). As is the case for invasive breast cancer cells, we found that the majority of BCAR3 in MCF10A breast epithelial cells is in complex with Cas. However, less Cas is found in association with c-Src in MCF10A cells compared to more invasive breast cancer cells. These finding suggest that despite some similarities in BCAR3, Cas, and c-Src signaling between non-tumorigenic and invasive breast cancer cells, both the physical interplay between these proteins and the signaling pathways activated downstream become dysregulated as non-tumorigenic cells progress to a fully oncogenic state. A better understanding of BCAR3/Cas/c-Src signaling in non-tumorigenic versus invasive breast tumor cells is necessary to determine whether this signaling network contributes to breast tumor progression and metastasis.

Citation Format: Allison M. Batties, Michael S. Guerrero, Ashley L. Wilson, Keena S. Thomas, Amy H. Bouton. The BCAR3/Cas/c-Src signaling node in breast cancer progression. [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 2638. doi:10.1158/1538-7445.AM2013-2638
Abstract 2643: Regulation of stem cell differentiation by p53.
Authors: Li Mangmang , He Yunlong , Dubois Wendy , Huang Jing
Published: August 2014

Differentiation is one of the barriers for tumorigenesis and reprogramming. In addition, embryonic stem (ES) and induced pluripotent stem (iPS) cells utilize differentiation to maintain their genomic stability by removing cells with mutation burden from the population in response to DNA damage. The tumor suppressor p53 is critical for regulating the differentiation of ES and iPS cells. However, the mechanism of how p53 regulates the differentiation of ES and iPS cells is unclear. To systematically study the underlying mechanisms, we employ an integrated genome-wide study in mouse ES cells. Systems analyses reveal that p53-activated genes and p53-repressed genes have distinct regulatory mechanisms and functions in mouse ES cells. Binding of p53 at promoter region significantly correlates with gene activation but not with repression. In addition, we unexpectedly identify a novel regulatory mode for p53-mediated repression through interfering with distal enhancer activity. This regulatory mode explains some but not all of the p53-mediated repressive events. Importantly, many ES cell-enriched core transcription factors are p53-repressed genes. Further analyses demonstrate that p53-repressed genes are functionally associated with ES and iPS cell status while p53-activated genes are linked to differentiation. Genome-wide analyses also show that p53-activated genes and -repressed genes have distinguishable features of expression levels and epigenetic markers, such as H3K4me3, H3K27me3 and H3K79me2. Upon DNA damage, p53 regulates the self-renewal and pluripotency of ES cells. Together, these results support a model that, in response to DNA damage, p53 affects the status of ES cells through activating differentiation-associated genes and repressing ES cell-enriched genes. Because many poorly differentiated tumors express ES-cell gene signature, our results may also have far-reaching implications in how p53 restricts the origin of cancer stem cells in adult tissues.

Citation Format: Mangmang Li, Yunlong He, Wendy Dubois, Jing Huang. Regulation of stem cell differentiation by p53. [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 2643. doi:10.1158/1538-7445.AM2013-2643

Note: This abstract was not presented at the AACR Annual Meeting 2013 because the presenter was unable to attend.
Abstract 2648: Self-renewing cellular compartments in embryonal rhabdomyosarcoma are modulated by Hedgehog signaling.
Authors: Satheesha Sampoorna , Dogan Nurhak , Casanova Elisa A. , Walter Dagmar , Schäfer Beat W.
Published: August 2014

Rhabdomyosarcoma (RMS) is a heterogeneous group of malignancies with features of impaired skeletal muscle differentiation and represents the majority of the soft tissue cancers in the pediatric age group. Current treatment regimens offer some clinical respite but at a significant risk of averse side effects and secondary tumors later in life. Identifying and targeting sub-populations with higher tumorigenic potential and self renewal capacity not only decreases tumor burden but also lowers the probability of relapse.

Using the sphere formation assay we could select for self-renewing cells from the heterogeneous monolayer cultures of embryonal RMS (eRMS) cell lines, RD and Rh36, which had multipotent differentiation ability in vitro and greater tumorigenic potential in vivo. These cells also showed increased expression of embryonic stem cell genes. Interestingly, quantitative gene expression analysis revealed increased expression of various components of the Hedgehog pathway, a developmental pathway important in eRMS biology. Inhibition or activation of the pathway in vitro by small molecule agents led to increased or decreased self renewing ability respectively. Furthermore we observed that eRMS adherent monolayer cultures contained a sub-population of GLI1-positive cells which could be modulated by pathway manipulation. Importantly, inhibition of the hedgehog pathway in vitro led to significantly delayed tumor initiation in vivo. Ongoing experiments aim at confirming these results on the genetic level.

We postulate that the Hedgehog pathway sustains a primitive cellular compartment necessary for tumor initiation within embryonal RMS cell lines.

Citation Format: Sampoorna Satheesha, Nurhak Dogan, Elisa A. Casanova, Dagmar Walter, Beat W. Schäfer. Self-renewing cellular compartments in embryonal rhabdomyosarcoma are modulated by Hedgehog signaling. [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 2648. doi:10.1158/1538-7445.AM2013-2648
Abstract 2649: Human adipose tissue-derived mesenchymal stem cells can target brain tumor initiating cells.
Authors: Choi Seung Ah , Wang Kyu-Chang , Phi Ji Hoon , Lee Ji Yeoun , Choi Jung Won , Kim Hyung Ah , Kim Se Hee , Kim Yong Hwy , Kwon Sung Eun , Kim Young-Hoon , Kim Seung-Ki
Published: August 2014

Abstract

The discovery of neural stem cells offers paradigm shift in brain tumor research. In neuro-oncology, the biology of neural stem cells has been pursued in two ways: as cancer stem cells to understand the origin and maintenance of brain tumors and as a potential cell-based vehicle for gene therapy. In addition to neural stem cells, mesenchymal stem cells (MSCs) have been reported to possess tumor-tropic migratory capacities. However, there is scanty data on migratory capacity of MSC toward brain tumor initiating cells (BTICs). This study focuses on investigating the ability of human adipose tissue derived MSCs (hAT-MSCs) to target BTICs and their cross-talk in the microenvironment.

BTICs were isolated from three different kinds of brain tumors (medulloblastoma, atypical teratoid/rhabdoid tumor (ATRT), glioblastoma). The migration capacities of hAT-MSCs toward BTICs were examined both in vitro transwell assay and in vivo bioluminescence imaging analysis. To investigate the crosstalk between hAT-MSCs and BTICs, various cytokines were analyzed. Using co-culture system of hAT-MSCs and BTICs, we analyzed the mRNA expression patterns of cytokine receptors (CCR2, CCR4, CCR5, CCR7, CCR9, CCR10, XCR1, CXCR1, IL-8R, CXCR4, CX3CR1, IL1R1, IL6R, MET, PDGFRB, KDR, CD44, IFNAR1 and TEK) by qRT-PCR and protein level of their ligand in co-cultured media by Bio-Plex human cytokine ELISA.

We confirmed the migratory capacity of t hAT-MSCs toward BTICs in vitro and BTICs-derived xenograft brain tumors in vivo live imaging. mRNA expression of receptors (CCR4, CCR5, CCR10, XCR1, CXCR1, IL-8R, CXCR4, PDGFRB, KDR, CD44, IFNAR1 and TEK) increased two- to eighteen-fold higher levels. The cytokine analysis revealed that the ligand levels of hAT-MSCs (medulloblastoma: VEGF, IL6, IL8, CCL3, CCL2 and TEK; ATRT: VEGF, CCL3 and TEK; glioblastoma: CCL2, PDGF, TEK and IGF1) and the ligand levels BTICs (medulloblastoma: CCL5 and CXCL4; ATRT: IL-8 and CXCL4; glioblastoma: CXCL4 and IGF1) were elevated in the co-cultured media.

Our findings demonstrated that hAT-MSCs can target BTICs. Cross-talk between hAT-MSCs cytokine receptor and BTICs ligand (medulloblastoma: CCR5/CCL5 and CXCR4/CXCL4; ATRT: IL-8R/IL-8 and CXCR4/CXCL4; glioblastoma: CXCR4/CXCL4 and IGF1R/IGF1) play cardinal role in this process.

Citation Format: Seung Ah Choi, Kyu-Chang Wang, Ji Hoon Phi, Ji Yeoun Lee, Jung Won Choi, Hyung Ah Kim, Se Hee Kim, Yong Hwy Kim, Sung Eun Kwon, Young-Hoon Kim, Seung-Ki Kim. Human adipose tissue-derived mesenchymal stem cells can target brain tumor initiating cells. [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 2649. doi:10.1158/1538-7445.AM2013-2649
Abstract 267: MYC maintains the self-renewal of cancer stem cells through the regulation of the glutathione redox system.
Authors: Das Bikul , Tsuchida Rika , Bayat-Mokhtari Reza , Gotlib Jason R. , Reijo-Pera Renee , Yeger Herman , Dean Felsher W.
Published: August 2014

Cancer stem cells (CSCs) are a rare population of self-renewing cells that can persist even after extensive therapeutic intervention. The molecular mechanism of CSC self-renewal is not yet clear. Here we show that MYC is causally involved in CSC self-renewal through the regulation of glutathione (GSH) redox system. In a MYC-driven conditional transgenic mouse model of T-cell acute lymphoblastic leukemia (T-ALL), we identified Sca-1+ CSCs. We found that these CSCs express the glutathione (GSH) redox system related genes including glutamate cysteine ligase (GCL) and Pyruvate Dehydrogenase Kinase 1 (PDK1) but low levels of expression of the mitochondrial redox enzyme, Superoxide Dismutase 2 (SOD2). Even a brief period of MYC inactivation in the Sca1+ CSCs led to their apoptosis associated with the downregulation of GCL and PDK1, but the upregulation of SOD2, in turn resulting in ROS-generation and mitochondrial damage. Moreover, suppression of SOD2 by siRNA silencing or the use of the peroxide scavenger, ascorbic acid, prevented MYC inactivation from inducing apoptosis in the Sca-1+ CSCs. Finally, in human CSCs, including primary human Chronic Myeloid Leukemia (CML), and human colon cancers, we confirmed that MYC expression was required for high GCL, PDK1 and low SOD2. Even the brief inactivation of MYC in human CSCs resulted in the loss of their self-renewal, as well as the global chance in redox state by decrease in GCL, PDK1 and increase in SOD2. We conclude that high levels of continued MYC expression are essential for CSC self-renewal through regulation of redox state.

Citation Format: Bikul Das, Rika Tsuchida, Reza Bayat-Mokhtari, Jason R. Gotlib, Renee Reijo-Pera, Herman Yeger, Felsher W. Dean. MYC maintains the self-renewal of cancer stem cells through the regulation of the glutathione redox system. [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 267. doi:10.1158/1538-7445.AM2013-267

Note: This abstract was not presented at the AACR Annual Meeting 2013 because the presenter was unable to attend.
Abstract 2685: Regulation of melanoma metastasis by malignant melanoma-initiating cell marker ABCB5.
Authors: Ma Jie , Alloo Allireza , Wilson Brian J. , Frank Markus H.
Published: August 2014

Cancer stem cells drive tumor growth, including metastatic disease progression. ATP-binding cassette member B5 (ABCB5) expression identifies malignant melanoma initiating cells (MMICs), which are distinguishable from cancer bulk populations by their self-renewal and differentiation capacities, and pro-oncogenic interactions with the microenvironment. ABCB5 confers drug resistance in multiple human malignancies through its actions as a drug efflux transporter. Here, we demonstrate that ABCB5 also regulates melanoma metastasis. ABCB5 monoclonal antibody (mAb) treatment significantly inhibited metastasis development in highly immune-compromised NOD-scid IL2Rγnull mice carrying fully established human melanoma xenografts, independent of previously reported antibody-dependent cell-mediated cytotoxicity (ADCC) effects. ABCB5 mAb treatment was well tolerated, as indicated by equal body weights of treated mice and untreated controls. Moreover, ABCB5 mAb also inhibited melanoma cell invasive capacity, suggesting that ABCB5 regulates intravasation and/or extravasation of metastatic tumor cells. Furthermore, the average size of metastatic nodules in ABCB5 mAb-treated mice was significantly smaller than those in isotype control mAb-treated or untreated mice, indicating that ABCB5 also regulates proliferation of disseminated tumor cells. Expression profile analysis revealed that multiple known metastasis-associated genes are regulated by ABCB5. Rescue of their expression could reverse the anti-metastatic effects induced by ABCB5 inhibition. Thus, ABCB5 is not only a marker for MMICs, but functionally drives metastatic melanoma progression.

Citation Format: Jie Ma, Allireza Alloo, Brian J. Wilson, Markus H. Frank. Regulation of melanoma metastasis by malignant melanoma-initiating cell marker ABCB5. [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 2685. doi:10.1158/1538-7445.AM2013-2685

Note: This abstract was not presented at the AACR Annual Meeting 2013 because the presenter was unable to attend.
Abstract 2687: Krüppel-like factor 8 targets matrix metalloproteinase-14 to promote breast cancer invasion and metastasis .
Authors: Lu Heng , Hu Liu , Yu Lin , Wang Xianhui , Shen Chao , Li Tianshu , Mukherjee Debarati , Lahiri Satadru , Wason Melissa , Zhao Jihe
Published: August 2014

Krüppel-like factor 8 (KLF8) regulates critical gene transcription and invasion events associated with cancer, including epithelial to mesenchymal transition (EMT) and extracellular matrix degradation. Here, we report a novel KLF8-to- matrix metalloproteinase-14 (MMP14) signaling that promotes human breast cancer cell EMT, invasion and metastasis. Cell-surface protein biotinylation and streptavidin pull-down showed that overexpression of KLF8 induced cell-surface accumulation of MMP14 in MCF-10A cells in a FAK-dependent manner, which was correlated with increased MMP14 activity and the ectodomain shedding of E-cadherin. On the other hand, knockdown of KLF8 in MDA-MB-231 cells reduced expression and activity of MMP14 on the cell surface. Promoter reporter assays and chromatin and oligonucleotide precipitations determined that KLF8 activated the human MMP14 gene promoter both directly and indirectly via beta-catenin. Importantly, Matrigel invasion assays showed that MMP14 was required for the KLF8-induced MCF-10A cell invasion in vitro. Experiments using nude mice demonstrated that MMP-14 overexpression downstream of KLF8 in the MDA-MB-231 cells makes a significant contribution to the invasion and angiogenesis in the local primary tumors and lung metastasis. Taken together, this work identified a novel role and mechanisms for MMP14 downstream of KLF8 in the progression of breast cancer metastasis.

Citation Format: Heng Lu, Liu Hu, Lin Yu, Xianhui Wang, Chao Shen, Tianshu Li, Debarati Mukherjee, Satadru Lahiri, Melissa Wason, Jihe Zhao. Krüppel-like factor 8 targets matrix metalloproteinase-14 to promote breast cancer invasion and metastasis . [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 2687. doi:10.1158/1538-7445.AM2013-2687
Abstract 2694: SRC-1 promotes tumor angiogenesis through up-regulation of HIF1α-mediated VEGFa expression.
Authors: Qin Li , Xu Yixiang , Feng Zhen , Wu Yelin , Xu Yan , Liao Lan , Xu Jianming
Published: August 2014

Steroid receptor coactivator 1 (SRC-1) belongs to the p160 coactivator family which also contains SRC-2 (TIF2) and SRC-3 (AIB1/pCIP/ACTR/RAC3). These coactivators interact with nuclear receptors and other transcription factors to up-regulate gene expression. Accumulated evidence shows that SRC-1 exerts its promotive role in breast cancer progression and metastasis by up-regulating transcription of multiple genes important for tumor cell epithelial-mesenchymal transition (EMT), migration, invasion and endocrine therapy resistance. However, its role in tumor angiogenesis has not been examined. In our recent studies using SRC-1 KO/PyMT and WT/PyMT mammary tumor cell lines established in our lab, we found that injection of SRC-1 KO/PyMT cells to matrigel plug formed under the back skin in SCID mice induced less blood vessel formation compared with that of WT/PyMT tumor cells. Accordingly, injection of human breast cancer MDA-MB-231 cells with stable knockdown of SRC-1 by shRNA to the matrigel plug also induced less neovascularization. Meanwhile, VEGFa mRNA level is significantly reduced in mammary tumors from SRC-1 KO/PyMT transgenic mice. On the contrary, in SRC-1-overexpressed mammary tumors from MMTV/TVA/PyMT mice, the expression of VEGFa was dramatically increased. In agreement with these findings, in vitro studies revealed that VEGFa mRNA levels are much lower in two of the SRC-1 KO/PyMT tumor cell lines compared with WT/PyMT cell lines. Knockdown of SRC-1 in WT/PyMT cells and human MDA-MB-231 cells reduced VEGFa levels, while adenovirus-mediated re-expression of SRC-1 in SRC-1 KO/PyMT cells significantly potentiated VEGFa expression. Furthermore, promoter analyze revealed that SRC-1 could interact with HIF-1α and AP-1 to potentiate the VEGFa promoter activity. We also found that both SRC-1 and HIF-1α are associated with a VEGFa promoter region by ChIP assays. These results indicate that SRC-1 can promote breast cancer progression and metastasis through enhancing tumor angiogenesis. Targeting SRC-1 may be a feasible strategy for inhibiting breast tumor angiogenesis and metastasis.

Citation Format: Li Qin, Yixiang Xu, Zhen Feng, Yelin Wu, Yan Xu, Lan Liao, Jianming Xu. SRC-1 promotes tumor angiogenesis through up-regulation of HIF1α-mediated VEGFa expression. [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 2694. doi:10.1158/1538-7445.AM2013-2694

Note: This abstract was not presented at the AACR Annual Meeting 2013 because the presenter was unable to attend.
Abstract 2696: TMPRSS4 regulates levels of integrin alpha-5 in NSCLC through miR-205 activity.
Authors: Larzabal Leyre , Redrado Miriam , Lopez de Aberasturi Arrate , Rueda Paloma , Rodriguez Maria J. , Lopez Lissette , Montuenga Luis , Calvo Alfonso
Published: August 2014

TMPRSS4 is a membrane-anchored protease involved in cell migration and invasion in different cancer types including lung cancer. We have previously shown that TMPRSS4 expression is increased in NSCLC and that its inhibition through shRNA reduces lung metastasis. The goal of the present study was to elucidate molecular mechanisms involved in the protumorigenic role of TMPRSS4 in NSCLC. A knock down strategy and microarray analyses were established to identify genes regulated by TMPRSS4. Among other genes, miR-205 was identified as an overexpressed gene upon TMPRSS4 downregulation. Levels of miR-205 were found to be reduced in NSCLC cells compared to non-malignant cells. miR-205 overexpression promoted the acquisition of an epithelial phenotype with increased E-cadherin and reduced fibronectin expression, probably through the regulation of transcription factors ZEB1 and ZEB2. Furthermore, miR-205 inhibited very significantly (p<0.01) tumor cell migration, invasion and metastasis formation in vivo. A further transcriptional and luciferase assays in miR-205-overexpressing cells identified integrin alpha-5 (a proinvasive protein overexpressed in tumors) as a new miR-205 target in NSCLC. We have therefore demonstrated for the first time a new molecular connection between TMPRSS4 and integrin alpha-5 through miR-205.

Citation Format: Leyre Larzabal, Miriam Redrado, Arrate Lopez de Aberasturi, Paloma Rueda, Maria J. Rodriguez, Lissette Lopez, Luis Montuenga, Alfonso Calvo. TMPRSS4 regulates levels of integrin alpha-5 in NSCLC through miR-205 activity. [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 2696. doi:10.1158/1538-7445.AM2013-2696
Abstract 2699: Organ-specific signatures of melanoma metastasis.
Authors: Sundstrøm Terje , Prestegarden Lars , Tepper Clifford , Ingham Elizabeth , Even Lisa , Johnson Sarah , Skaftnesmo Kai Ove , Lund-Johansen Morten , Bjerkvig Rolf , Ferrara Katherine , Thorsen Frits
Published: August 2014

We have previously reported on a reproducible and predictive animal model that can be used to elucidate the enigmatic biology of melanoma metastasis, a key prerequisite for therapeutic advances. Here, we have used this model to investigate the molecular drivers responsible for metastasis formation in different organs, with a special emphasis on brain-specific features. A human melanoma brain metastasis cell line (H1) was developed in our laboratory, transduced with the genes for green fluorescent protein and luciferase, and injected into the left cardiac ventricle of seven immunodeficient mice. Whole-body bioluminescence imaging (BLI) and brain magnetic resonance imaging was performed weekly to follow the development of metastases. The animals invariably developed both intracranial and extracranial tumors. At about seven weeks, the animals were euthanized and organ involvement was examined in detail by ex vivo BLI. The brain, femurs, adrenals, and ovaries from five random animals were enzymatically dissociated using organ-tailored protocols. Sample yield was validated by fluorescence microscopy prior to fluorescence-activated cell sorting of tumor cells. Three tumor cell samples from the brain, femurs, adrenals, and ovaries of different mice were then analyzed using next-generation RNA-sequencing (NGS) technology. We also performed baseline NGS on the H1 cell line before injection. Taken together, bioinformatics analysis revealed unique molecular signatures of organ-specific metastasis, validating some previously reported metastasis drivers and introducing other genomic features critical to the detrimental momentum of metastatic melanoma.

Citation Format: Terje Sundstrøm, Lars Prestegarden, Clifford Tepper, Elizabeth Ingham, Lisa Even, Sarah Johnson, Kai Ove Skaftnesmo, Morten Lund-Johansen, Rolf Bjerkvig, Katherine Ferrara, Frits Thorsen. Organ-specific signatures of melanoma metastasis. [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 2699. doi:10.1158/1538-7445.AM2013-2699
Abstract 2713: The Pten and p53 tumor suppressors cooperate to constrain the PI3K/mTOR pathway activation.
Authors: Liu Pixu , Cheng Hailing , Symonds Lynn , Xu Erbo , Maira Sauveur-Michel , di Tomaso Emmanuelle , Roberts Thomas , Zhao Jean
Published: August 2014

Loss of Pten expression and mutational inactivation of p53 are frequent molecular events in liver cancer associated with advanced disease stage and poor prognosis. The tumor suppressor PTEN is a phosphoinositide phosphatase that antagonizes the class IA PI3K-engaged oncogenic signaling by dephosphorylating PtdIns(3,4,5)P(3); however, the specific roles of class IA PI3K isoforms, p110a and p110b, have not been assessed in genetically engineered mouse models of liver cancer with Pten inactivation. Furthermore, effective treatment strategies for Pten and p53 deficient liver malignancy remain to be defined. In this study, we showed that combined deletion of Pten and p53 in liver led to rapid development of hepatocellular carcinoma and cholangiocellular carcinoma, the two most common types of liver cancer, with complete penetrance. While a single ablation of p110a or p110b markedly reduced the tumor burden, inactivation of both p110a and p110b completely blocked the liver tumorigenesis. Although liver tumors driven by inactivation of Pten and p53 showed a good response to GDC-0941, a pan-PI3K inhibitor, the combined treatment with both GDC-0941 and Rad001, an mTOR inhibitor or single-agent BEZ235, a dual pan-PI3K and mTOR inhibitor, led to marked tumor regression. These studies suggest that Pten and p53 cooperate to constrain the PI3K/mTOR pathway activation and inhibitors targeting both PI3K and mTOR may effectively treat Pten and p53 deficient liver cancers.

Citation Format: Pixu Liu, Hailing Cheng, Lynn Symonds, Erbo Xu, Sauveur-Michel Maira, Emmanuelle di Tomaso, Thomas Roberts, Jean Zhao. The Pten and p53 tumor suppressors cooperate to constrain the PI3K/mTOR pathway activation. [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 2713. doi:10.1158/1538-7445.AM2013-2713
Abstract 2718: Spheroid culture of colorectal cancer cells from patient tumor tissue preserves important cellular characteristics of the parental tumor.
Authors: Jeppesen Maria , Hagel Grith , Glenthoj Anders , Thastrup Ole , Jorgensen Lars Nannestad , Thastrup Jacob
Published: August 2014

At the time of diagnosis more than 65% of patients with colorectal cancer have advanced disease and require chemotherapeutic treatment. However, colorectal tumors demonstrate pronounced heterogeneity, leading to significant individual variation in sensitivity to standard chemotherapeutic drugs. Tools for assessing chemosensitivity and tailoring therapeutic regimens to the individual patient are therefore urgently needed. A promising approach is to isolate viable cancer cells from patient tissue and to test the chemosensitivity of the individual tumor in vitro.

The aim of the current study was to establish primary colorectal cancer cell cultures from patient tissue and investigate if the cultured cells retain important characteristics of the original tumor. Small cell clusters were isolated from colorectal tumor tissue and allowed to form cell spheroids in suspension culture. The cell type composition of patient spheroids was characterized by immunostaining with cell type specific antibodies. Spheroid size was monitored over time and growth curves established for each patient. The distribution of proliferating and apoptotic cells in the spheroids was assessed by staining for specific markers. The 3D culture system was verified as a tumor model by comparing the results obtained for the spheroids to paraffin-embedded sections of the original tumors.

Successful spheroid culture was obtained from more than 80% of tissue samples. Immunostaining confirmed that the primary cultures were composed of colorectal cancer cells, without significant contaminations by other cell types. Growth could be observed as an increase in spheroid size. The dominating cell populations of the original tumor could generally be identified in the derived spheroids as well. In summary, spheroid cultures of colorectal cancer cells can be established from tumor tissue of individual patients with a high success rate and the formed spheroids retain important cellular characteristics of the original tumor. Culture of colorectal cancer cells as spheroids represents a promising method for establishing primary cultures for chemotherapeutic screening of the individual patient.

Citation Format: Maria Jeppesen, Grith Hagel, Anders Glenthoj, Ole Thastrup, Lars Nannestad Jorgensen, Jacob Thastrup. Spheroid culture of colorectal cancer cells from patient tumor tissue preserves important cellular characteristics of the parental tumor. [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 2718. doi:10.1158/1538-7445.AM2013-2718
Abstract 2722: Characteristics of pancreatic cancer spheroids as a model for anticancer efficacy test.
Authors: Yeon Sang-Eun , Lee Sang-Hoon , Kuh Hyo-Jeong
Published: August 2014

Compared to the conventional monolayers or suspension cultures, 3D culture models have drawn attention as a vivo-mimic model which can produce clinically relevant data. Among the several types of 3D culture models, multicellular spheroid (MCS) is a appropriate model for study of penetration and efficacy of anticancer agents. Purpose of this study was to establish MCS of human pancreatic cancer cell (Panc-1) and evaluate its usefulness as anticancer efficacy test. Panc-1 MCS was produced successfully by using poly-dimethylsiloxane(PDMS) microwell instead of agarose-coated 96-well system. MCS with average diameter of 375 μm was prepared in 700 μm microwells after 4 days of culture. Spherical morphology with rather rough surface and junctional structures in outer cell layers were observed with no notable necrotic area. Extracellular matrix proteins such as collagen I, fibronectin, and laminin were detected in the interstitial space of MCS. Stem cell population with CD44+/CD24+/ESA+ expression increased from 0.1% to 2.1% when cultured as MCS. miRNA expression profiling showed a difference between monolayers and MCS, i.e., 36 and 85 genes were up- and down-regulated in MCS, respectively. Imaging of drug penetration and viability indicators including MitoSOX, Edu incorporation, and Ki-67 expression were optimized for microwell format. Colorimetric assays of MTS and APH were performed after transferring MCS to 96-well plates. Overall, we demonstrated that PDMS microwell is an appropriate platform for preparation of MCS of Panc-1 cells and MCS showed different biological features from monolayers, suggesting that Panc-1 MCS prepared using PDMS microwell may be a useful 3D model for anticancer efficacy test.

Citation Format: Sang-Eun Yeon, Sang-Hoon Lee, Hyo-Jeong Kuh. Characteristics of pancreatic cancer spheroids as a model for anticancer efficacy test. [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 2722. doi:10.1158/1538-7445.AM2013-2722

Note: This abstract was not presented at the AACR Annual Meeting 2013 because the presenter was unable to attend.
Abstract 2737: Spontaneous genomic alterations identified in a chimeric model of colorectal cancer guide effective combinatorial therapy.
Authors: Zhou Yinghui , Rideout William M. , Bressel Angela , Yalavarthi Sireesha , Zi Tong , Monti Anthony , Bottega Steve , Feng Bin , Chiu M. Isabel , Bosenberg Marcus , Heyer Joerg
Published: August 2014

Colon cancer is the second most common cause of cancer mortality in the western world. We generated a mouse model of colon cancer in which a conditional mutation of p53 is combined with inducible delta131-beta-Catenin over-expression in the intestinal epithelium. After 4-12 months of beta-Catenin induction, small adenomas and large solitary nodules of invasive adenocarcinomas developed in either the upper or lower intestinal tract, while the remainder of the intestine appeared normal. Molecular analysis revealed that EGFR and MET were expressed in advanced adenocarcinomas but not early stage adenomas. Although propagating mouse colon tumors has been very challenging, we nevertheless successfully established one tumor line through direct in vivo propagation in the kidney capsule. This tumor line, CB42, grew aggressively and metastasized to ancillary lymph nodes as well as the lung in a subcutaneous metastasis assay and grew in the liver when seeded there by intra splenic injection. Treatment with a MET inhibitor Crizotinib, had no effect on either primary tumor growth, or metastasis. By comparing the genome of CB42 with other colon tumors that did not propagate, we discovered an amplicon on chromosome five containing genes that correlated with propagation and metastasis. In addition, whole genome sequencing revealed that CB42 harbored a mutation in KRAS. Results from combination therapies against key genetic alterations will be presented.

Citation Format: Yinghui Zhou, William M. Rideout, Angela Bressel, Sireesha Yalavarthi, Tong Zi, Anthony Monti, Steve Bottega, Bin Feng, M. Isabel Chiu, Marcus Bosenberg, Joerg Heyer. Spontaneous genomic alterations identified in a chimeric model of colorectal cancer guide effective combinatorial therapy. [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 2737. doi:10.1158/1538-7445.AM2013-2737
Abstract 274: Neuropilin-2 is upregulated during EMT in lung cancer.
Authors: Nasarre Patrick , Roche Joelle , Potiron Vincent A. , Nair-Menon Joyce , Gemmill Robert M. , Drabkin Harry A.
Published: August 2014

Neuropilins (NRP1 and NRP2) are high affinity receptors for the class-3 semaphorins, cell guidance molecules involved in tissue development, immune responses, angiogenesis and cancer. Semaphorins can inhibit proliferation, affect cell protrusion, spreading and adhesion, and trigger migratory responses often in a repulsive manner depending on the cellular context.However, neuropilins can also serve as receptors for galectin-1 and growth promoting factors including VEGF, PlGF, EGF and TGFβ.

We previously cloned the SEMA3F gene from chromosome 3p21.3, which undergoes homozygous deletion and frequent loss of heterozygosity in lung cancer. We found that SEMA3F levels were inversely correlated with tumor aggressiveness in human lung cancer and confirmed its tumor suppressor activity in experimental xenograft models. We subsequently discovered that SEMA3F is directly downregulated by ZEB1, a transcription factor involved in the epithelial to mesenchymal transition (EMT). In the present study, we asked whether NRP2, the SEMA3F-specific receptor, was also regulated during the EMT process.TGFβ, a physiologic EMT inducer, rapidly increased NRP2 expression in NSCLC cell lines. Mechanistically, this increase resulted primarily from enhanced translation with some contribution from increased mRNA levels. However, TGFβ did not stabilize NRP2 mRNA or protein. Furthermore, we found out that a substantial amount of protein at the cell surface directly depended on the presence of TGFβ in the tumor microenvironment. The pathways regulating NRP2 induction include ERK1/2 and Akt but are Smad independent. In addition, forced expression of ZEB1 also induced NRP2. Conversely, ZEB1 inhibition reduced NRP2 upregulation by TGFβ. Importantly, inhibiting NRP2 affected TGFβ-induced morphologic changes, migration and ERK1/2 activation. Interestingly, some EMT target genes were also affected by NRP2 knockdown. Preliminary data in xenograft models indicate that NRP2 inhibition counteracts tumor growth-promoting effects of TGFβ .In a lung tumor tissue microarray, we observed that higher tumor grades were characterized by positive staining for NRP2 and negative staining for E-cadherin while lower tumor grades often presented the opposite profile. Since NRP2 is the highest affinity receptor for SEMA3F, our results suggest that loss of SEMA3F coupled with increased NRP2 would facilitate the binding of growth factors to NRP2 to further promote EMT and metastasis. Therefore, targeting NRP2 could be an important therapeutic approach against EMT in lung cancer.

Citation Format: Patrick Nasarre, Joelle Roche, Vincent A. Potiron, Joyce Nair-Menon, Robert M. Gemmill, Harry A. Drabkin. Neuropilin-2 is upregulated during EMT in lung cancer. [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 274. doi:10.1158/1538-7445.AM2013-274
Abstract 2741: NF-κB activation by IL-1α and p62 feedforward loops is required for Kras-induced pancreatic ductal adenocarcinoma.
Authors: Ling Jianhua , Kang Ya'an , Chiao Paul
Published: August 2014

Pancreatic ductal adenocarcinoma (PDAC) is the fourth leading cause of cancer death in the United States with a less than six month mean survival period and less than five percent of an overall five-year survival rate. Despite developments in the management of pancreatic cancer, the survival rate has remained unchanged in the past 50 years. Thus, a better understanding of the mechanism of pancreatic cancer inception and development is the key to identifying prevention strategies and therapeutic targets for increasing patient survival rates, which is one of the greatest challenges in pancreatic cancer research.

The mutational activation of Kras is an early event in PDAC development and has been detected in 90% of PDAC and 50% of pancreatic intraepithelial neoplasia (PanIN). The genetic mutation of oncogenic Kras in the pancreas induces chronic pancreatitis and pancreatic cancer. However, the key signaling pathway of the downstream Kras remains unidentified.

Here, we report that pancreas-specific targeted IKK2/β inactivation inhibited NF-κB activation and PDAC development in KrasG12D and KrasG12D; Ink4a/ArfF/F mice, demonstrating a mechanistic link between IKK2/β/NF-kB signaling pathway and KrasG12D signaling pathways in PDAC inception. Our findings reveal that KrasG12D-activated AP-1 induces IL-1α, which, in turn, activates NF-κB and its target genes IL-1α and p62, to initiate IL-1α/p62 feedforward loops for inducing and sustaining NF-κB activity. Furthermore, IL-1α overexpression correlates with Kras mutation, NF-κB activity, and poor survival in PDAC patients. Knock-down of IL-1α in mouse PDAC cell lines and lung adenocarcinoma cell lines significantly reduces tumorigenesis in orthotopic xenograft mouse and subcutaneous xenograft mouse models. Therefore, our findings demonstrate the new mechanism by which IKK2/β/NF-κB is activated by KrasG12D through dual feedforward loops of IL-1α/p62 to promote development of PDAC. This suggests that the interruption of the dual feedforward loops of IL-1α/p62 could be the therapeutic targets for inhibiting oncogenic Kras signaling in PDAC.

Citation Format: Jianhua Ling, Ya'an Kang, Paul Chiao. NF-κB activation by IL-1α and p62 feedforward loops is required for Kras-induced pancreatic ductal adenocarcinoma. [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 2741. doi:10.1158/1538-7445.AM2013-2741