bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract LB-295: Combination of PI3K (SAR245408 and SAR245409) and MEK (MSC1936369B) inhibitors induce tumor apoptosis in vivo: fluorescence molecular tomography study in a human colon carcinoma xenograft model
Authors: GUERIF Stephane , D'heilly Sebastien , Lefranc Celine , Egile Coumaran , Debusshe Laurent , Vincent Loïc , Carrez Chantal , Chiron Marielle , Ma Jiango , Ogden Janet A. , Lager Joanne , Garcia-Echeverria Carlos , Lengauer Christoph
Published: August 2014

The Merck-Serono MEK1/2 allosteric inhibitor MSC1936369B (formerly known as AS703026) was combined with the sanofi-aventis pan-PI3K inhibitor SAR245408 (also known as XL147) or the pan-PI3K/mTOR inhibitor SAR24540 (also know as XL765) in mice bearing colon carcinoma HCT116 xenograft tumors. These tumors harbor both KRAS (G13D) and PIK3CA (H1047R) mutations. Apoptosis induction was monitored non-invasively in vivo using fluorescence molecular tomography (FMT) after injection of fluorescent Annexin-V. Ex vivo apoptosis was assessed on tumor lysates using assays for cleaved caspase-3 and cleaved PARP detection.

Suboptimal doses of SAR245408, SAR245409, and MSC1936369B were administered orally on a daily schedule as single agents or in combination. Single agent treatments showed marginal or no activity on HCT116 tumor growth inhibition. In contrast, robust enhanced anti-tumor activity was observed with the combination treatments, resulting in sustained tumor stasis and regression for both SAR245408/MSC1936369B (p<0.0001) and SAR245409/MSC1936369B (p=0.0009) combination regimens. This therapeutic synergy was associated with clear induction of tumor apoptosis after four days of treatment, as evidenced by markedly increased levels of cleaved caspase-3 and cleaved PARP proteins in the tumor lysates. Apoptosis monitored in vivo by FMT was induced for the SAR245408/MSC1936369B combination, with p=0.005 and p<0.0001 after three and seven days of therapy respectively.

In conclusion, the combination between the MEK1/2 inhibitor MSC1936369B with the pan-PI3K inhibitor SAR245408 or the pan-PI3K/mTOR SAR245409 resulted in significantly enhanced anti-tumor activity in a KRAS/PIK3CA mutated tumor xenograft model, with synergistic induction of tumor apoptosis as demonstrated ex vivo for both combinations and in vivo using longitudinal FMT imaging for the SAR245408/MSC1936369B combination.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr LB-295. doi:10.1158/1538-7445.AM2011-LB-295
Abstract LB-296: Involvement of T-cell immunity in changes in the plasma amino acid profiles of patients with colorectal cancer
Authors: Ohara Eiji , Kitadai Yasuhiko , Shinagawa Kei , Sumida Tomonori , Tanaka Miwako , Ohnishi Mayu , Nagai Kenta , Ueno Yoshitaka , Tanaka Shinji , Fujii Kiyomu , Nakamura Kazuki , Yoneda Junya , Chayama Kazuaki
Published: August 2014

Metabolic changes in patients with various disorders, including liver disease and cancer, lead to alterations in amino acid balance, and plasma amino acid profiles can be used to discriminate between non-malnourished patients with cancer and healthy individuals. Not with standing, the mechanisms of amino acid imbalance are not well understood. In patients with cancer, various factors including metabolic changes in the tumor itself and immune responses may affect plasma amino acid levels. To investigate involvement of the immune system, we measured plasma amino acid levels in an orthotopic transplant model of colon cancer established in both conventional BALB/c mice and athymic BALB/c nude mice (T-lymphocyte-deficient mice).

CT26 colon cancer cells (murine colon cancer cell line) were implanted into the cecal wall of the BALB/c mice (n=10) and nude mice (n=10). Plasma samples were collected from the tumor-bearing mice at 7 and 14 days after transplantation, and changes in plasma amino acid profiles were compared. Plasma was also collected from mice that underwent sham operation (control mice, n=10). Plasma amino acid concentrations were measured by LC/MS/MS.

The plasma concentrations of several amino acids changed significantly in tumor-bearing mice compared to concentrations in control mice. Although cells from the same line were injected into the cecal wall of BALB/c mice and nude mice, a difference was noted in the change in plasma amino acid levels. Plasma Val, Ile, Leu, and Met levels decreased in BALB/c mice but increased in nude mice. There was no difference in the amino acid profile of CT26 tumor tissues between BALB/c mice and nude mice. We then injected antiCD3-antibody into the peritoneal cavity of tumor-bearing BALB/c mice (n=5) to eliminate T-cell function. We also injected IgG into tumor-bearing BALB/c mice (n=5) for control. The pattern of change in amino-acid profiles differed between the two groups of mice.

Our findings suggest that T-lymphocyte-mediated immunoreactions are involved in changes in the plasma amino acid profile in patients with colorectal cancer.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr LB-296. doi:10.1158/1538-7445.AM2011-LB-296
Abstract LB-302: Inhibition of STAT3 following systemic delivery of STAT3 antisense oligonucleotide results in target inhibition in tumor and tumor-associated stromal cells and dramatic reduction in circulating pro-tumorigenic cytokines
Authors: Kim Youngsoo , Hsu Jeff , Zhou Tianyuan , He Guobin , May Chris , Monia Brett P. , MacLeod Robert
Published: August 2014

Constitutive activation of STAT3 has been documented in a broad range of neoplastic tissues. STAT3 is a transcription factor that regulates the expression of numerous genes involved in the proliferation, survival, and metastasis of tumor cells. In addition, STAT3 contributes to tumorigenesis by modulating immune responses mediated by tumor stromal cells in the tumor microenvironment. The immuno-suppressive, pro-tumorigenic effects of STAT3 are regulated by induction or inhibition of several key cytokines. However STAT3 remains a difficult target to drug with conventional approaches. Here, we have studied the effects of selective downregulation of STAT3 in various cancer settings by optimized antisense oligonucleotides (ASOs). We assessed the effects of STAT3 downregulation on the expression of cytokines and the progression of cancer. Initially, the safety of inhibiting STAT3 was demonstrated in tolerability studies, where no notable toxicities were observed despite near complete abrogation of STAT3 expression in the liver following systemic delivery of ASO in both rodents and in non-human primate. ASO treatment led to STAT3 inhibition in tumor cells and several non-tumor stromal cell populations of the tumor. In addition, dramatic reduction in key pro-tumorigenic cytokines including IL-6 and IL-1b was observed in C26 colon cancer-bearing animals, which correlated with the decrease of STAT3 level in tumor and stromal cells. This result suggests that STAT3 might not only transmit the IL-6 signal from upstream pathway, but regulates IL-6 level possibly in an auto-feedback manner. Downregulation of STAT3 also led to the inhibition of IL-6-induced C-reactive protein (CRP) expression, which was recently reported to play a key role in the progression of multiple myeloma (MM). Moreover, inhibition of STAT3 resulted in a significant delay in tumor growth in STAT3-dependent MDA-MB-231 breast cancer and U251 glioblastoma xenograft models following systematic ASO treatment. The results from this study collectively suggest that STAT3 expression can be specifically modulated by ASOs in vivo tumor models and that in addition to effects in tumor cells STAT3 ASOs regulate the balance of tumorigenic cytokines in the tumor microenvironment.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr LB-302. doi:10.1158/1538-7445.AM2011-LB-302
Abstract LB-362: Both canonical and non-canonical autocrine Wnt signaling regulates clonogenic growth and epithelial-mesenchymal transition in colon cancer cells lacking mutational activation of Wnt pathway
Authors: Planutis Andryus , Planutiene Marina , Holcombe Randall F. , Planutis Kestutis
Published: August 2014

Introduction: Wnt signaling in cells with mutationally-induced constitutive pathway activation promotes clonogenic growth and epithelial-mesenchymal transition (EMT). The role of autocrine Wnt signals in colon cancer cells lacking a Wnt pathway activating mutation is less clear.

Hypothesis: Extracellular Wnt inhibitors will reduce clonogenic growth and EMT in colon cancer cell lines lacking mutational Wnt pathway activation by inhibiting basal levels of autocrine Wnt signaling. Methods: Two cells lines were utilized: HCT116B is a colon cancer cell line in which an abnormal beta-catenin allele has been removed by homologous recombination; RKO is a colon cancer cell line with no known Wnt pathway activation. The Wnt inhibitors DKK1, which blocks canonical Wnt signals by interaction with LRP5 and sFRP1 which blocks both canonical and non-canonical Wnt signals were utilized and transfected into each cell line. Canonical Wnt throughput was measured with a luciferase based reporter construct, clonogenic growth by colony formation in soft agar and EMT by morphology.

Results: Untransfected HCT116B, but not RKO expressed DKK1. Neither cell line expressed sFRP1. Both cell lines expressed significant quantities of DKK1 or sFRP1 after transfection. Canonical Wnt signaling was suppressed in RKO by both DKK1 and sFRP1 but only by DKK1 in HCT116B. DKK1 inhibited clonogenic growth in RKO but not in HCT116B, while sFRP1 inhibited it in both cell lines. EMT was clearly suppressed by these inhibitors in RKO; such an effect was less visible in HCT116B.

Discussion: Colon cancer cells lacking mutational activation of the Wnt pathway are susceptible to the actions of extracellular Wnt inhibitors, presumably via inhibition of autocrine Wnt signals and canonical Wnt signaling. However, cells with basal expression of a canonical Wnt inhibitor such as DKK1 (HCT116B) rely preferentially on non-canonical pathways for clonogenic growth and EMT which can be blocked by global Wnt ligand antagonists such as sFRP1. The importance of autocrine Wnt signaling and contributions by non-canonical pathways to clonogenic growth and EMT has important implications for the development of Wnt-targeted therapeutic agents for colon cancer.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr LB-362. doi:10.1158/1538-7445.AM2011-LB-362
Abstract LB-366: Patient-derived EBV-immortalized B-lymphocytes are a dominant contaminant of in vitro cultured human neuroblastoma tumor-initiating cells isolated from bone marrow
Authors: Påhlman Sven , Johnsson Sofie A. , Pietras Alexander , Wigerup Caroline , Øra Ingrid , Andäng Michael , Nilsson Kenneth , Olofsson Tor , Gisselsson David
Published: August 2014

Neuroblastoma is one of the most common solid tumors of infancy and is believed to originate from immature sympathetic nervous system progenitor cells. Tumor stem cells from different cancer forms have during the last decade been isolated and propagated in vitro in stem cell-promoting medium. These cells have given the opportunity to study the population within the tumor that is believed to be responsible for relapse and metastatic disease. Three years ago, putative neuroblastoma tumor-initiating cells (TICs) were isolated from bone marrow of patients with high stage disease. Several findings concerning these cells have been difficult to explain in a neuroblastoma context, i.e. that initially only cells from MYCN non-amplified tumors could be established as proliferating cultures in vitro, that they have a B-cell phenotype with VDJ rearrangement as presented at the 2010 Advances in Neuroblastoma Research (ANR) meeting, and as presented here, that they lack genomic imbalances and fail to recapitulate neuroblastoma morphology when xenografted. We show that these widely studied patient-derived TIC cultures (NB12, NB88 and NB122R), also at low passages, are highly contaminated by proliferating cells of a B-lymphocyte phenotype, with the absolute majority of cells expressing multiple B-cell antigens while lacking expression of neuroblastoma-associated markers. We further show that these lymphoblastoid cells are Epstein-Barr virus (EBV)-infected with robust expression of the two EBV nuclear antigens EBNA2 and EBNA3A. In addition, orthotopic and subcutaneous TIC-derived xenograft tumors all have EBNA2 and B-cell marker CD19 positive cells at a high frequency. Published and unpublished results based on the use of the proposed TIC lines as a model for neuroblastoma development should be considered with caution, as these lines, at least with time in culture, mainly consist of EBV-transformed B-lymphoblastoid cells.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr LB-366. doi:10.1158/1538-7445.AM2011-LB-366
Abstract LB-367: New evidence supports the potential use of methotrexate in children with MYCN-amplified neuroblastoma.
Authors: Lau Diana T. , Norris Murray D. , Marshall Glenn M. , Haber Michelle , Ashton Lesley J.
Published: August 2014

Background

Folate is essential for cell growth and differentiation. Cancer cells frequently up-regulate folate receptors to increase folate uptake to meet their elevated need for nucleotides to support DNA synthesis and growth. SLC19A1 encodes for a major folate or anti-folate transporter and has been reported to be regulated by oncogenic transcription factors. In childhood neuroblastoma, amplification of the MYCN oncogene is a strong prognostic marker for predicting unfavourable outcome. In light of previous findings showing that overexpression of SLC19A1 increases folate influx and methotrexate (MTX) sensitivity, we aimed to investigate the relationship between SLC19A1 expression, MYCN amplification and expression, to determine if this relationship modified MTX sensitivity in neuroblastoma.

Methods

Expression levels of SLC19A1 and MYCN were examined in 42 neuroblastoma patient samples and in two independent in vitro MYCN functional systems. The impact of MYCN amplification/expression and SLC19A1 expression on MTX sensitivity were evaluated on 13 neuroblastoma cell lines. SLC19A1 G80A polymorphism was also examined to determine its impact on SLC19A1 gene expression and MTX sensitivity.

Results

Expression levels of SLC19A1 and MYCN were positively correlated in neuroblastoma tumour samples (p=0.0001). Induction of MYCN in vitro increased the mRNA expression levels of SLC19A1 by 12-fold (p=0.0003) while silencing of MYCN reduced the expression levels of SLC19A1 by 7-fold (p=0.0017). Neuroblastoma cell lines with MYCN overexpression or amplification were at least 2000-fold more sensitive to MTX than those without (expression: p=0.0002; amplification: p<0.0001). However, SLC19A1 G80A polymorphism did not appear to impact on SLC19A1 gene expression or MTX sensitivity.

Conclusions

Our data indicate a potential functional link between MYCN and the regulation of anti-folate uptake. Our findings suggest that MYCN amplification confers MTX sensitivity through the regulation of the folate receptor; SLC19A1. Targeting SLC19A1 mediated anti-folate uptake in patients with MYCN amplification may provide a new therapeutic approach in children with neuroblastoma.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr LB-367. doi:10.1158/1538-7445.AM2011-LB-367
Abstract LB-368: The reduction of Bmi-1 expression increases the sensitivity of prostate cancer the cell line PC3 to X-ray irradiation
Authors: Jason Gonzales C. , Fink Louis M.
Published: August 2014

Bmi-1 is a proto-oncogene that has been implicated in the polycomb complex that is required for cell proliferation and the maintenance of stem cells in a variety of tissues and cancers. Bmi-1 expression has been found to be elevated in a variety of tumors and its abundance has been correlated with survival. Previous studies have implicated Bmi-1 as having a role in radioresistance of cancer cells. In the present studies, we have shown that a stably integrated siRNA, which is conditionally regulated by doxycycline, can be used to study the susceptibility of prostate cancer cell line PC3 to radiation injury following a reduction of Bmi-1 expression.

Human HEK293 cells were transfected with lentiviral packaging plasmids and the pTRIPZ vector containing an RFP reporter gene upstream of the Bmi-1 siRNA. PC3 cells were stably transduced with the Bmi-1 siRNA via lentivirus and were initially selected for the puromycin resistance. The RFP reporter gene and Bmi-1 siRNA is under control of a tetracycline response element (TRE) and, following the administration of doxycycline, RFP and Bmi-1 siRNA are concomitantly expressed. Studies on the expression of RFP, Bmi-1 RNA expression by RT-PCR and Bmi-1 measurements by western blots confirm the stable conditional expression of Bmi-1 in prostate cancer PC3 cells.

The cell cycle analysis, the apoptotic rate and growth rates of the Bmi-1 expressing and non-expressing cell cultures were not significantly different. However, after treatment with 8 Gy x-irradiation, there was a marked increase in cells in the G2/M and in DNA fragmentation in the PC3 cells when Bmi-1 was reduced. There also was a marked decrease in the plating efficacy and in the cloning efficacy in methylcellulose.

Perhaps the delineation of the Bmi-1 status of the cells in a biopsy or in the circulating tumor cells of patients with PC can be used to predict the biological behavior and therapeutic response of these cancers. This study was supported by grant DE-FG02–08ER64608 from the DOE.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr LB-368. doi:10.1158/1538-7445.AM2011-LB-368
Abstract LB-37: Inducible knockout of GRP78/BiP in the hematopoietic system suppresses Pten-null leukemogenesis and AKT oncogenic signaling
Authors: Wey Shiuan , Luo Biquan , Ni Min , Zhou Hui , Fu Yong , Bhojwani Deepa , Carroll William L. , Lee Amy S.
Published: August 2014

Traditionally GRP78 is regarded as protective against hypoxia and nutrient starvation prevalent in the microenvironment of solid tumors, thus, its role in hematologic malignancies remains to be determined. In this study, elevated GRP78 expression was detected in leukemic blasts of adult patients, leukemia cell lines and inversely correlates with time to relapse in childhood acute lymphocytic leukemia. To directly elucidate the requirement of GRP78 in leukemogenesis, we created a biallelic conditional knockout mouse model of GRP78 and PTEN in the hematopoietic system. Strikingly, heterozygous knockdown of GRP78 in PTEN null mice is sufficient to restore the hematopoietic stem cell (HSC) population back to the normal percentage and suppress leukemic blast cell expansion. AKT/mTOR activation in PTEN null bone marrow cells is potently inhibited by Grp78 heterozygosity, corresponding with suppression of AKT phosphorylation by GRP78 knockdown in leukemia cell lines. This is the first demonstration that GRP78 is a critical effector of leukemia progression, at least in part through control of oncogenic AKT/mTOR signaling. Furthermore, overexpression of GRP78 renders human leukemic cells more resistant to AraC-induced apoptosis whereas knockdown of GRP78 sensitizes them, suggesting GRP78 is a novel potent therapeutic target for leukemia.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr LB-37. doi:10.1158/1538-7445.AM2011-LB-37
Abstract LB-379: Combating trastuzumab resistance by targeting Src, a common node downstream of multiple resistance pathways
Authors: Zhang Siyuan , Huang Wen-Chien , Li Ping , Guo Hua , Poh Say-Bee , Brady Samuel , Xiong Yan , Tseng Ling-Ming , Li Shau-Hsuan , Ding Zhaoxi , Sahin Aysegul , Esteva Francisco , Hortobagyi Gabriel , Yu Dihua
Published: August 2014

Objective of the study: Trastuzumab is a highly successful example of rationally designed ERBB2-targeted therapy. However, about half of patients with ERBB2-overexpressing breast cancer do not respond to trastuzumab-based therapies due to various resistance mechanisms. Clinically applicable regimens for overcoming trastuzumab resistance of different mechanisms are not yet available. The objective of the study is to find a common key node of multiple trastuzumab resistance mechanisms and design a clinical applicable strategy to target this common node to more effectively overcome trastuzumab resistance. Methodology: First, we evaluated Src activity in multiple acquired and de novo trastuzumab-resistant cell lines models. Cells with modulated Src activity were examined for trastuzumab resistance in vitro and in an orthotopic xenograft model. Moreover, tumors from patients receiving trastuzumab therapy were also analyzed retrospectively for clinical correlation of Src activity with trastuzumab response. Src activated trastuzumab-resistant cells were further treated with Src inhibitor (saracatinib), trastuzumab, or a combination of both in vitro and in vivo. Results: We demonstrate that the non-receptor tyrosine kinase Src serves as a critical modulator of trastuzumab response and a common node downstream of multiple trastuzumab resistance pathways. Src is activated in both acquired and de novo trastuzumab-resistant cells. Increased Src activation conferred significant trastuzumab resistance (P = 0.011) in breast cancer cells and correlated with trastuzumab resistance in patients. Targeting the common node Src in combination with trastuzumab universally sensitized multiple lines of trastuzumab-resistant cells to trastuzumab and eliminated trastuzumab-resistant tumors in vivo. Targeting Src, a critical convergence point of divergent trastuzumab resistance mechanisms, represents a highly effective and clinically convenient strategy to overcome trastuzumab resistance.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr LB-379. doi:10.1158/1538-7445.AM2011-LB-379
Abstract LB-38: Selective Smoothened antagonists for anti-tumor therapy
Authors: Tunici Patrizia , Mancuso Mariateresa , Giordano Cinzia , Ferruzzi Pietro , Mennillo Federica , Tanori Mirella , Saran Anna , Thomas Russell ,   Gal. La Pericot Mohr, Bakker Annette
Published: August 2014

The Hedgehog (Hh) pathway is an important signaling pathway during cellular proliferation, differentiation and embryonic morphogenesis. Aberrant pathway activation has been linked to tumorigenesis in several cancers, (including pancreatic, prostate, lung, colorectal, bladder and ovarian) and therefore the inhibition of the pathway could be an attractive approach for anticancer therapy. The pathway is either activated by the binding of one of the three Hh ligands (sonic, Indian and desert) to the Patched transmembrane protein or by the mutation of inhibitory pathway components (e.g. Pached, SUFU,…). Pathway activation by ligand binding to Patched releases patched repression of the smoothened (Smo) receptor. Activated Smo initiates a cascade of events leading to the activation of transcription factors of the Gli family. Pathway activation by mutation has been reported in diseases such as medulloblastoma or basal cell carcinoma in which the Patched co-receptor is mutated, thereby functionally inactivated and unable to repress the Smo receptor. We report the characterization of a novel Smoothened receptor antagonist (SEN450), showing Hedgehog pathway specific activity in hedgehog responsive cellular models. Finally, we show a synergistic effect of SEN450 with temozolomide in a subcutaneous CD-1 nude mice xenograft of DBTRG-05MG neurospheres. Our data indicate that growing conditions can affect hedgehog responsive cells and that a combined therapy with temozolomide and hedgehog inhibitors could be helpful in the prevention/delay of GBM recurrence. Moreover, we have obtained extremely promising in vivo data using two compounds structurally related to SEN450 in a xenograft tumour model that have been generated from irradiated heterozygous Patched mice. In the current study, the Ptc+/− mice were used to generate a tumor model by transplanting medulloblatoma fragments subcutaneously in nude mice. Upon tumor development two Smo antagonists administered orally were tested in this model. Both compounds showed a significant reduction in tumor volume that was maintained over the entire dosing period. Histological examination showed very low mitotic index in treated tumors versus controls. The expression of the target gene Gli1 revealed a significant decrease of the gene expression in treated samples. The results corroborated the specificity of action of both molecules in targeting the Hh pathway always activated in this tumor model. Further investigation is in progress in order to verify the antitumor capacity of the compounds in other tumor models.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr LB-38. doi:10.1158/1538-7445.AM2011-LB-38
Abstract LB-382: Inhibition of CD95 signalling by APG-101 enhances efficacy of radiotherapy (RT) and reduces RT-induced tumor satellite formation
Authors: Pfenning Philipp N. , Weiler Markus , Merz Christian , Jugold Manfred , Gieffers Christian , Platten Michael , Fricke Harald , Wick Wolfgang
Published: August 2014

CD95 is a prototype death receptor that regulates the induction of apoptosis, upon binding of its ligand CD95L. Therefore, activation of the CD95L/CD95 system has been regarded an excellent target for treatment of various malignancies. In contrast to the aim of promoting CD95 activation, there is compelling evidence that CD95 is able to promote tumor growth through intracellular non-apoptotic signalling mechanisms. This mechanism has recently been described for glioblastoma, in which activation of CD95 by CD95L leads to invasive growth and glioma cell migration. This is signalled through increased activity of pivotal glioblastoma invasion-related proteases, that is matrix metalloproteinases (MMP). With this shift of paradigms, blocking of the CD95L mediated invasion of tumor cells and subsequent tumor progression may therefore be a promising approach in anti-glioma therapy. APG101 is a fusion protein, consisting of the extracellular domain of human CD95 and the Fc-region of human immunoglobulin G. Hence, it acts as a soluble CD95 receptor trapping the CD95 ligand.

The presented project aims at analyzing the clinical relevance of blocking CD95 signalling, given the fact that glioma patients with upregulated CD95 expression have worse survival rates than those with intermediate expression levels. In proof-of principle experiments APG101 inhibits CD95L-mediated invasion of glioma cells. More importantly, APG101-treatment (100 mg/kg body weight) resulted in significantly prolonged survival of SMA560-tumor bearing Vm/Dk mice, less glioma cell satellites in the surrounding tissue and reduced activity of MMP. APG101 in combination with focal irradiation at 6 Gy demonstrated a remarkable reduction of tumor growth with a significantly prolonged survival compared with irradiation treatment alone and inhibition of the proinvasive properties of radiotherapy as demonstrated by magnetic resonance imaging and histology. Surprisingly, APG-101 added to the vascular endothelial growth factor receptor (VEGFR) inhibitor cediranib (AZD2171) did not increase the survival of SMA-560-tumor bearing mice as compared to cediranib alone nor did it impair the proinvasive consequences of cediranib.

Our data strongly support the potential use and clinical evaluation of APG101 in combination with radiotherapy in the treatment of malignant glioma.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr LB-382. doi:10.1158/1538-7445.AM2011-LB-382
Abstract LB-39: Phenotypic switching and molecular anatomy of malignant progression (E-M-T/M-E-T).
Authors: Gao Chong-Feng , Vande woude George F.
Published: August 2014

Proliferation and invasion are two fundamental phenotypes required for malignancy while phenotypic switching is required for malignant progression. Thus, during malignant progression primary tumor cells switch from proliferative to invasive phenotypes and switch again to form proliferating metastases. The acquisition of the invasive phenotype through epithelial-mesenchymal-transition (E-M-T) is a crucial step in tumor progression, as is mesenchymal-epithelial-transition (M-E-T) or switching to become tumor metastases. Using a reductionist approach, we have been able to reproduce these steps through a combination of in vitro and in vivo methods: Beginning with prostate and ovarian carcinoma cells, we show that the majority are epithelial (E) cells, grow as acini in three dimensional (3D) Matrigel and are nonresponsive to hepatocyte growth factor/scatter factor (HGF/SF). These E cells grow in soft agar and are tumorigenic in immunocompromised mice. However, there is a second cell population which also grows as acini in 3D Matrigel, but display extensive branching morphogenesis when HGF/SF is present. These subclones are mesenchymal (M), are highly invasive in response to HGF/SF, but do not grow in soft agar and display low or no tumorigenicity in vivo. We show that M cells can originate in vitro from the non-invasive E subclones. However, M subclones undergo M-E-T only in vivo, at low frequency mimicking tumor progression to proliferating metastases.

We show that E-M-T switching events occur with loss of E-Cadherin and the gain of c-MET expression. Expression of β-Catenin does not change with phenotypic switching, but with the loss of E-Cadherin, its localization shifts from the cell membrane to nuclear/cytoplasmic location, activating β-Catenin dependent expression. Moreover, depleting E-Cadherin in E cells triggers nuclear/cytoplasmic shift of β-Catenin and enhanced HGF/SF-c-Met signaling, while ectopic expression of E-Cadherin in M cells restores E phenotype and decreases HGF/SF-Met signaling. Importantly, we show that phenotypic switching occurs with distinct changes in karyotype that dictate changes in the chromosome transcriptome and the expression of genes that account for changes to M or to E phenotypes, thereby implicating chromosome instability as a key mechanism for phenotypic switching. We conclude that chromosome instability generates the transcriptome diversity in vitro or in vivo, which in the appropriate environment, leads to expansion of clonal variants responsible for phenotypic switching.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr LB-39. doi:10.1158/1538-7445.AM2011-LB-39
Abstract LB-391: Aurora kinase inhibitor AT9283 potently inhibits the proliferation and migration of medulloblastoma cells
Authors: Macdonald Tobey J. , Liu Jingbo , Lyons John
Published: August 2014

Aurora kinases play an important role in regulating cell mitotic division. It has been found that they are over expressed in many human cancers, and decreased expression in either Aurora kinase A or B levels lead to cell apoptosis. It has been suggested that Aurora kinase inhibitors might be an attractive molecular target of anticancer therapy. AT9283, a multi-targeted kinase inhibitor with potent activity against both Aurora kinases A and B, is currently being evaluated in clinical trails in patients with advanced hematological malignancies including AML. However to date, this compound has not been evaluated in medulloblastoma, one of the most common malignant brain tumors in children. In this study, AT9283 was investigated in a panel of human medulloblastoma cell lines (DAOY and D556) and the SmoA1 transgenic mouse model of medulloblastoma cells (PS125) to assess its ability to inhibit medulloblastoma cell survival, growth and migration. Western blotting demonstrated significant inhibition of both Aurora kinases A and B in each cell type by AT9283. MTT assay demonstrated that AT9283 significantly inhibited cell growth and proliferation in a dose-dependent manner in all 3 cell lines. Furthermore, the compound potently reduced phospho-histone H3 levels, and induced caspase-3 and cleaved PARP in all cells. AT9283 treatment at concentrations as low as 0.01–0.05μM significantly inhibited cell migration, as demonstrated by the Scratch assay without affecting caspase-3 induction, cleaved PARP or phospho-histone H3 levels. Thus taken together, our results indicate that AT983 has a dual effect on inhibiting cell growth and migration in medulloblastoma cells, suggesting it may be a valuable therapy in medulloblastoma treatment and warrants further preclinical in vivo investigation in this disease model.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr LB-391. doi:10.1158/1538-7445.AM2011-LB-391
Abstract LB-396: FGFR4-antagonistic CovX-Body™ inhibits tumor growth
Authors: Kaspar Allan A. , Liu Dingguo , Do Janet , Betancourt Oscar , Liu Bin , Desharnais Joel , Del Rosario Joselyn , Li Lingna , Pirie-Shepherd Steven , Levin Nancy , Bhat Abhijit , Woodnutt Gary , Lappe Rodney
Published: August 2014

FGFR4 is the receptor for FGF19. Published data has shown FGF19 transgenic animals develop hepatocellular carcinoma (HCC), suggesting that the FGFR4:FGF19 axis may play a role in HCC. FGFR4 upregulation has been shown to be a negative prognostic indicator in multiple tumor types.

CVX-63, an FGFR4-specific CovX-Body, was produced by fusing a chemically modified peptide to the specific Fab binding site of a specially designed antibody. CVX-63 was shown to inhibit FGF19 binding to FGFR4 in a competition ELISA with an IC50 of 10 nM, and had no cross reactivity with FGFR1–3.

CVX-63 has a beta half life of 55 hrs in the mouse. The anti-tumor activity of CVX-63 was demonstrated in a staged Colo-205 xenograft model, with CVX-63 administered i.v. weekly. Maximal activity of 30% tumor growth inhibition was seen at dosing 10 mg/kg. Immunohistochemistry analysis of treated tumors shows a >90% reduction in phospho-ERK and 50% downregulation of FGFR4 on tumor cells. Ki67, a marker of proliferation was reduced, but active caspase 3 was not found in treated tumors.

A patient-derived HCC xenograft was identified which expresses both FGFR4 and FGF19. When tested in this HCC model, CVX-63 did not show any effect inhibiting tumor growth. These data demonstrate that CVX-63 effectively inhibits FGFR4 activity in xenografts. FGFR4 inhibition has a modest effect on human tumor progression and FGFR4 does not seem to be a critical driver of HCC.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr LB-396. doi:10.1158/1538-7445.AM2011-LB-396
Abstract LB-400: STAT3 mediates resistance to MEK inhibitor through microRNA miR-17
Authors: Dai Bingbing , Meng Jieru , Peyton Michael , Girard Luc , Bornmann William G. , Ji Lin , Minna John D. , Fang Bingliang , Roth Jack A.
Published: August 2014

AZD6244 is a small-molecule-based inhibitor targeting the MEK pathway that is currently in clinical trials. However, the mechanisms mediating intrinsic resistance to MEK inhibition remain to be completely characterized. To identify the molecular mechanism of MEK inhibitor resistance, we analyzed responses of 38 lung cancer cell lines following AZD6244 treatment and their genome-wide gene expression profiles and identified a panel of genes correlated with sensitivity or resistance to AZD6244 treatment. Ingenuity pathway analysis revealed that activation of the signal transducer and activator of transcription 3 (STAT3) pathway was associated with MEK inhibitor resistance. Inhibition of the STAT3 pathway using JSI-124, a STAT3-specific inhibitor, sensitized lung cancer cells to AZD6244 treatment and induced apoptosis in vitro and in vivo. Moreover, combining a STAT3 inhibitor with AZD6244 induced the expression of BIM and poly (ADP-ribose) polymerase (PARP) cleavage, whereas activation of the STAT3 pathway inhibited BIM expression and thus induced MEK inhibitor resistance. In addition, we found that miR-17, which is regulated by STAT3, played an important role in STAT3-mediated MEK inhibitor resistance by targeting BIM. Inhibition of miR-17 with anti-miR-17 sensitized resistant cells to AZD6244 by inducing BIM and PARP cleavage. We conclude that STAT3-mediated overexpression of miR-17 blocked BIM expression, which resulted in resistance to AZD6244. These results implicate novel alternative approaches for overcoming the MEK inhibitor resistance by combining AZD6244 with STAT3 inhibitors or miR-17 antagomirs.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr LB-400. doi:10.1158/1538-7445.AM2011-LB-400
Abstract LB-42: Inducible mouse models for studying liver stem cell activation and tumor development
Authors: Endo Yoko , Zhang Mingjun , Yamaji Sachie , CANG YONG
Published: August 2014

In contrast to skin and intestine, liver does not undergo constant renewal, but it can regenerate itself after massive damage or volume loss. Liver regeneration is driven by proliferation of parenchymal hepatocytes, but when hepatocyte proliferation is compromised, quiescent liver stem cells (LSCs) can be activated to differentiate to transit-amplifying oval cells and eventually to hepatocytes and cholangiocytes. Currently no specific biomarkers have been defined for LSCs, making it difficult to characterize this rare population of cells. Conventional rodent models, i.e., mice or rats treated with liver damaging chemicals followed by partial hepatectomy, are not ideal for studying LSCs due to the promiscuous toxicity of the chemicals and the lack of distinguishing damaged and non-damaged cells.

Here we demonstrate that hepatocyte-specific deletion of Damaged DNA Binding protein 1 (DDB1), an E3 ubiqutin ligase required for cell cycle progression targeted by oncogenic hepatic B virus X protein, abolishes self-renewal capacity of hepatocytes. This defect leads to compensatory activation of DDB1-expressing and EpCam-positive oval cells. These DDB1 mutant mice exhibit very minor liver damage, compared with mice that are treated with diethyldithiocarbamate (DDC). These EpCam-positive oval cells, purified by fluorescent-activated cell sorting, express both hepatotcyte and cholangiocyte markers, can form colonies in vitro, and differentiate to hepatocytes after transplantation. Microarray analysis was performed to compare gene expression profiles in EpCam-positive cells isolated from DDB1 mutant mice and from DDC-treated mice. Several previously unrecognized genes are found to be specifically upregulated in DDB1 mutant mice. LGR5, a stem cell marker found in intestine, colon, stomach, and hair follicles, is demonstrated to be not expressed in facultative LSCs or oval cells in both our genetic models and DDC models. Interestingly, DDB1 mutant mice develop liver tumors in an age-dependent manner, and the onset of tumorigenesis is positively correlated to the extent of LSC activation. We are currently investigating the possible origin of these tumors from activated LSCs.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr LB-42. doi:10.1158/1538-7445.AM2011-LB-42
Abstract 1008: Concomitant HDAC and PI3K-AKT pathway inhibition as a new therapeutic strategy for prostate cancer.
Authors: Ku Sheng-Yu , Pili Roberto , Ellis Leigh
Published: August 2014

Background: Prostate cancer (PCa) is the most prevalent cancer in men in the United States. Prostate cancer patients that progress to a lethal phenotype are currently treated with chemotherapy, second generation androgen therapies and immunotherapy. Unfortunately, patient response is not often durable; therefore novel treatments are urgently needed. Histone deacetylases (HDACs) have shown to be up-regulated in PCa. Preclinical data indicates that HDAC inhibitors induce apoptosis in androgen receptor (AR)-positive cells. Unfortunately, HDACi display minimal clinical benefit. From this, we propose that combination of HDACi with novel targeted therapies will result in greater therapeutic efficacy. Methods: PC3, PC3AR and PtenCaP8 PCa cell lines treated with panobinostat (HDACi) and BEZ235 (PI3K-mTORC1 inhibitor) as single treatment or in combination for 48 hours. Cell death/apoptosis was assessed by loss of cell membrane integrity, phosphotidylserine exposure, cell cycle and caspase activation by flow cytometry. Expression levels of activated AKT were assessed by immunoblot. Results: Panobinostat induces cell death/apoptosis greater in PC3AR than parental PC3 cells by increasing subG1 population (47.8±9.3 vs. 11.1±2.8,), phosphotidylserine exposeure (41.4±3.0 vs. 21±7.2,) and caspase activation (57.3±0.9 vs. 12.0 ±4.1,) respectively. Moreover, PC3AR cells treated with panobinostat results in loss of AKT activation, but not in PC3 cells. Further, combination of panobinostat with BEZ235 increases greater cell apoptosis in PC3AR cells compared to panobinostat single agent (30.6±1.9 vs. 19.9±1.4) and in PtenCaP8 cells (42.0±6.5 vs. 29.4±2.2). Western blotting also shows that combination treatment leads to loss of AKT activation compared to panobinostat and BEZ235 as a single agent. Conclusions: Overall, our results suggest that inhibition of HDACs induces greater cell death associated with loss of AKT activation in AR-positive cells, but not in AR-null cells. Also, combination of HDAC inhibitor and PI3K-mTORC1 inhibitor has more effective cell cytotoxicity compared to single agent, suggesting that inhibition of HDACs and PI3K-mTORC1 will exhibit greater therapeutic efficacy. In future work, combination of panobinostat with BEZ235 will be investigated in preclinical animal models to propose this therapy combination for clinical trials in patients with lethal PCa.

Citation Format: Sheng-Yu Ku, Roberto Pili, Leigh Ellis. Concomitant HDAC and PI3K-AKT pathway inhibition as a new therapeutic strategy for prostate 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 1008. doi:10.1158/1538-7445.AM2013-1008
Abstract 1016: Combination low dose 5-azacytidine (AZA) and romidepsin (FK228) therapy re-sensitizes ovarian cancer cells to cisplatin.
Authors: Wilson Andrew , Sinaise Lanzi , Readus Jade M. , Park Michelle , Lalani Anum S. , Saskowski Jeanette , Khabele Dineo
Published: August 2014

Ovarian cancer is the deadliest gynecologic malignancy. Current options for treating platinum-resistant disease are limited by drug choices and toxicities. Epigenetic therapy is emerging as a strategy for reversing tumor resistance and a recent encouraging clinical trial in platinum-resistant ovarian cancer suggests a potential unique application in ovarian cancer. The epigenetic drugs histone deacetylase inhibitors (HDACi) and DNA methyltransferase inhibitors (DNMTi) cause chromatin relaxation and multiple anti-tumor effects on cell growth, cell cycle, cell death and deregulation of DNA damage response and repair (DDR) pathways. Among HDACi we have tested in a panel of ovarian cancer cells, the depsipeptide FK228 exerts the most potent growth inhibitory effects in the majority of cells. Therefore, we aimed to determine whether the combination of FK228 and the DNMTi 5-azacytadine (AZA) can re-sensitize ovarian cancer cells to cisplatin more effectively than either agent alone. We found that FK228 synergized with low doses of the DNMTi 5-azacytadine (AZA) in SRB cell growth assays in vitro, and that low dose combinations of the drugs enhanced the growth inhibitory effects of cisplatin. Similar effects were observed when ovarian cancer cells were grown as xenografts in immunocompromised mice in vivo. Furthermore, a key DDR sensor, phosphorylated gamma-H2AX (pH2AX) activation was associated with the enhanced cytotoxic effects of the combination of the epigenetics and may be a marker of response. In conclusion, we have identified a novel combination of epigenetic drugs that can re-sensitize ovarian cancer cells to cisplatin treatment, at least in part through stimulation of pH2AX. Targeting DDR pathways in DDR-deficient ovarian cancer cells with low dose epigenetic drugs represents a promising strategy in ovarian cancer treatment.

Citation Format: Andrew Wilson, Lanzi Sinaise, Jade M. Readus, Michelle Park, Anum S. Lalani, Jeanette Saskowski, Dineo Khabele. Combination low dose 5-azacytidine (AZA) and romidepsin (FK228) therapy re-sensitizes ovarian cancer cells to cisplatin. [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 1016. doi:10.1158/1538-7445.AM2013-1016