bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract A11: Modeling drug response and resistance using patient-derived xenograft models.
Authors: Yang Guizhi , Green John , Kowal Colleen , Cogan Shawn , Velazquez Roberto , Monahan John , Korn Joshua , Keen Nicholas , Williams Juliet , Gao Hui
Published: August 2014

Metastatic melanoma is a highly lethal type of cancer and is often refractory to all traditional chemotherapeutic agents. Melanoma is associated with a high burden of genetic alteration, e.g. BRAF, NRAS or NF1 mutation. In the clinic, BRAF inhibitors are used as a targeted therapy to treat melanoma patients, but thus far a single-agent approach is not capable of achieving a durable response for this lethal disease. Very often, patients rapidly develop resistance to BRAF inhibitors. In the current study, we propose a large scale in vivo screening using patient-derived xenograft models (PDXs) for compound profiling. Since these PDX melanoma models are derived from individual patient tumors and therefore represent the genetic heterogeneity of patient tumors, we propose a “one animal per model per compound” approach (mouse clinical trial) to understand the genetic factors that contribute to drug response. In addition, we use this approach to model drug resistance after long-term compound treatment. We find that the mouse clinical trial approach accurately predicts response of both cytotoxic chemotherapeutics as well as targeted therapies. In addition, this approach successfully recapitulates the resistance profile of a novel BRAF inhibitor LGX818 in melanoma. We also find that combination of LGX818 with other targeted therapy agents including PI3K and MEK inhibitors significantly increases the response rates and delays the development of resistance. Taken together, our findings suggest that the mouse clinical trial is a valid approach to model drug response and resistance, and can be used to predict patient response in the clinic.

Citation Information: Mol Cancer Ther 2013;12(11 Suppl):A11.

Citation Format: Guizhi Yang, John Green, Colleen Kowal, Shawn Cogan, Roberto Velazquez, John Monahan, Joshua Korn, Nicholas Keen, Juliet Williams, Hui Gao. Modeling drug response and resistance using patient-derived xenograft models. [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2013 Oct 19-23; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2013;12(11 Suppl):Abstract nr A11.
Abstract A171: Antitumor effects of intervenolin, a novel natural compound with anti-Helicobacter pylori activity, and its synthetic derivatives on gastric and colorectal cancer cells.
Authors: Kawada Manabu , Inoue Hiroyuki , Ohba Shun-ichi , Abe Hikaru , Hatano Masaki , Amemiya Masahide , Hayashi Chigusa , Usami Ihomi , Watanabe Takumi , Igarashi Masayuki , Masuda Tohru , Shibasaki Masakatsu , Nomoto Akio
Published: August 2014

Tumor tissues are composed of not only tumor cells but also the surrounding stroma. While the stroma consists of various kinds of cells, among them fibroblasts-like stromal cells can regulate the growth and metastasis of tumor cells positively or negatively. Thus, the modulation of the tumor-stromal cell interactions can be a new antitumor strategy. We have been searching compounds that modulate the interactions using coculture system of tumor cells and stromal cells. In the course of our screening, we have isolated a novel compound, intervenolin, from the culture broth of Nocardia sp. ML96-86F2. Intervenolin inhibits the growth of human gastric and colorectal cancer cell lines when cocultured with the respective organ-derived stromal cells more strongly than that of the cancer cells cultured alone. To improve low yield of intervenolin from the bacterial culture and to create more potent compounds, we have synthesized intervenolin and its derivatives. Some of the derivatives also show the same effect as intervenolin in vitro. Furthermore, intervenolin and its derivatives show antitumor effects against xenograft models of human gastric cancer cells and also MDR-expressing colorectal cancer cells in vivo. Furthermore, intervenolin and its derivatives exert selective anti-Helicobacter pylori activity. We are now investigating the mechanism of its action and trying to synthesize more potent its derivatives.

Citation Information: Mol Cancer Ther 2013;12(11 Suppl):A171.

Citation Format: Manabu Kawada, Hiroyuki Inoue, Shun-ichi Ohba, Hikaru Abe, Masaki Hatano, Masahide Amemiya, Chigusa Hayashi, Ihomi Usami, Takumi Watanabe, Masayuki Igarashi, Tohru Masuda, Masakatsu Shibasaki, Akio Nomoto. Antitumor effects of intervenolin, a novel natural compound with anti-Helicobacter pylori activity, and its synthetic derivatives on gastric and colorectal cancer cells. [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2013 Oct 19-23; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2013;12(11 Suppl):Abstract nr A171.
Abstract A177: Mode of action of PM060184, a new interfacial microtubule inhibitor of marine origin.
Authors: Martínez-Diez Marta , Martínez-Leal Juan Fernando , Garcia-Fernandez Luis F. , Galmarini Carlos M.
Published: August 2014

PM060184 belongs to a new family of tubulin-binding agents originally isolated from the marine sponge Lithoplocamia lithistoides. This compound is currently produced by total chemical synthesis and is under evaluation in Phase I clinical studies in patients with advanced cancer diseases. Recently published results indicate that members of the PM060184 family are interfacial microtubule inhibitors that bind with nanomolar affinity to unassembled αβ-tubulin dimers at the beta tubulin plus end (Pera et al. 2013, ACS Chem. Biol.) In an attempt to shed light into the mechanism responsible of its antitumor activity, we have studied the effects of PM060184 in tumor cells including microtubule network structure, centrosome modifications and type of cell death induced by this compound. PM060184 rapidly internalizes into the tumor cells and shows a high intracellular retention rate, reflecting the high avidity for its cellular target. In addition, the antitumor activity of the compound seems to be rather irreversible after short (30 min) exposure times. At the cellular level, PM060184 inhibits tubulin polymerization. Interestingly, PM060184 affects interphase cells, where the compound induces disorganisation and fragmentation of the microtubule network and disturbing cell migration. PM060184 also affects mitotic cells, inducing the appearance of multipolar mitosis and lagging chromosomes at the metaphase plate. All these effects correlate with prometaphase arrest and induction of caspase-dependent apoptosis or appearance of cells in a multinucleated interphase-like state unrelated to classical apoptosis pathways. In summary, these results indicate that PM060184 represents a new tubulin binding agent with promising potential as an anticancer agent.

Citation Information: Mol Cancer Ther 2013;12(11 Suppl):A177.

Citation Format: Marta Martínez-Diez, Juan Fernando Martínez-Leal, Luis F. Garcia-Fernandez, Carlos M. Galmarini. Mode of action of PM060184, a new interfacial microtubule inhibitor of marine origin. [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2013 Oct 19-23; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2013;12(11 Suppl):Abstract nr A177.
Abstract A193: Transmembrane protein TMEM92 as a novel target in prostate cancer.
Authors: Morgan Richard , Feng Gui , Pandha Hardev S.
Published: August 2014

TMEM92 is a previously uncharacterized gene predicted to encode a 159 amino acid peptide with a single transmembrane domain and sequence homology to the SHISA group of proteins, some of which have a role in early development. In this study we evaluated the expression and function of TMEM92 in prostate cancer. This gene is expressed in primary prostate tumors and the prostate cancer derived cell lines PC3, DU145 and LNCaP, both at the RNA and protein level. TMEM92 expression appears to be cancer specific, as it is not expressed in normal prostate tissue or in normal adult organs. It is also absent from the WPMY-1 cell line, which is derived from normal prostate epithelium. siRNA-mediated knockdown of TMEM92 in PC3 cells induces apoptosis, as does the treatment of these cells with an anti-TMEM92 antibody, whilst neither treatment is cytotoxic for WPMY-1 cells. Taken together these findings suggest that TMEM92 is a potential therapeutic target in prostate cancer.

Citation Information: Mol Cancer Ther 2013;12(11 Suppl):A193.

Citation Format: Richard Morgan, Gui Feng, Hardev S. Pandha. Transmembrane protein TMEM92 as a novel target in prostate cancer. [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2013 Oct 19-23; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2013;12(11 Suppl):Abstract nr A193.
Abstract A203: An in vitro tumor model to characterize tumor cell metabolism in heterogeneous complex microenvironments.
Authors: Rodenhizer Darren , Cojocari Dan , Wouters Bradley G. , McGuigan Alison P.
Published: August 2014

Tumor cells often exhibit altered metabolic behaviors that are influenced by the heterogeneity of the tumor microenvironment. This heterogeneity arises from the presence of multiple cell types, variable ECM compositions, and variable protein expression due to the presence of oxygen and nutrient gradients. Conventional methods of identifying tumor therapies fail to account for these metabolic variations in the context of a heterogeneous microenvironment. To address this shortfall, we have developed a layered, three-dimensional, heterogeneous tumor model that allows spatiotemporal metabolite collection on a timescale relevant for metabolomics profiling. SKOV-3 ovarian cancer cells were mixed with collagen and infiltrated into a scaffolding material to create a thin layer. Multiple layers were then stacked to make a thick section of tumor tissue which is then combined with a customized oxygen impermeable bioreactor. EF5 immunofluorescence was used to confirm the presence of an oxygen gradient and to quantify the spatial PO2 values, which mimic those found in solid tumors. Cell viability and nutrient, and drug distribution profiles were also characterized and profiles were found to resemble those found in spheroid culture and in vivo. Cellular response to hypoxia was confirmed through qPCR analysis of mRNA of hypoxia-inducible genes such as CA9 and CHOP, and with HIF1α staining. Our technology allows for the creation of complex, yet controlled tumor microenvironments while facilitating “snap shot” data acquisition, and will allow researchers to decouple the influence of heterogeneity on metabolic response in cancerous solid tumors.

Citation Information: Mol Cancer Ther 2013;12(11 Suppl):A203.

Citation Format: Darren Rodenhizer, Dan Cojocari, Bradley G. Wouters, Alison P. McGuigan. An in vitro tumor model to characterize tumor cell metabolism in heterogeneous complex microenvironments. [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2013 Oct 19-23; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2013;12(11 Suppl):Abstract nr A203.
Abstract A235: Anti-CSF-1R antibodies reduce tumor-associated macrophages and inhibit tumor growth in preclinical models.
Authors: Li Yanxia , Mathew Sneha , Wu Lan , Wang Ying , Persaud Kris , Burtrum Douglas , Balderes Paul , Surguladze David , Haurum John , Ludwig Dale , Doody Jacqueline
Published: August 2014

In cancer, increased infiltration of macrophages within and surrounding the tumor mass correlates with increased tumor invasiveness and growth. In addition, presence of tumor-associated macrophages (TAMs) has been shown to correlate with poor prognosis, particularly in breast, prostate, ovarian, and cervical cancer. TAM proliferation, differentiation, and survival is dependent on CSF-1R activation, a type III integral membrane tyrosine kinase receptor selectively expressed on cells of the mononuclear phagocyte lineage. Given that TAMs enhance tumor growth and that activation of the CSF-1R pathway is required for TAM function, an antibody against mouse CSF-1R was generated for proof-of-principle studies. CS7, a monoclonal anti-mouse CSF-1R antibody inhibited both CSF-1 and IL-34 binding to mouse CSF-1R, leading to inactivation of the receptor and downstream signaling molecules. In addition, CS7 prevented monocyte proliferation and macrophage differentiation with IC50s of 0.1 nM and 0.75 nM, respectively. In murine models of breast cancer using CSF-1-secreting MDA-MB-231, Hcc1954, 4T1 or EMT6 tumor cells, CS7 treatment led to a marked reduction in TAMs and an associated decrease in tumor growth. In contrast, breast tumor xenografts with CSF-1 non-secreting breast tumor cell lines JimT1 and MCF-7 had limited or no decrease in tumor volume following CS7 treatment. In prostate models, DU145 tumor cell line xenografts (CSF-1-secreting) but not PC3 (CSF-1 non-secreting) tumor growth was inhibited by CS7, recaptitulating the results seen with breast xenografts. Thus, in breast and prostate preclinical models, CSF-1 secretion by tumor cells is a prerequisite for sensitivity to anti-CSF-1R treatment. Taken together, targeting CSF-1R with a monoclonal antibody inhibits CSF-1R signaling via CSF-1 and IL-34, prevents monocytic to macrophage differentiation, and reduces tumor volume in preclinical models, validating CSF-1R as a target for therapeutic application in cancer.

Citation Information: Mol Cancer Ther 2013;12(11 Suppl):A235.

Citation Format: Yanxia Li, Sneha Mathew, Lan Wu, Ying Wang, Kris Persaud, Douglas Burtrum, Paul Balderes, David Surguladze, John Haurum, Dale Ludwig, Jacqueline Doody. Anti-CSF-1R antibodies reduce tumor-associated macrophages and inhibit tumor growth in preclinical models. [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2013 Oct 19-23; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2013;12(11 Suppl):Abstract nr A235.
Abstract A246: Development of an IGF2 super-antagonist by directed evolution of IGF2R.
Authors: Frago Susana , Strickland Madeleine , Hughes Jennifer , Williams Christopher , Nicholls Ryan , Garner Lee , Rezgui Dellel , Crump Matthew P. , Hassan A. Bassim
Published: August 2014

Insulin-like growth factor 2 (IGF2) plays an important role both in human and mouse embryonic development and in tumour growth. IGF2 causes IGF1R and insulin receptor isoform A, IR(A), dependent activation of proliferation (MAPK) and survival pathways (AKT-PI3K). IGF2 bioavailability is tightly regulated by six IGF binding proteins and by the IGF2/mannose 6-phosphate receptor (IGF2R). IGF2R is a 15-domain extracellular receptor, frequently mutated in cancer, which clears IGF2 via binding to domain 11 and internalising it for lysosomal degradation. Increased supply of IGF2 occurs in hepatocellular, breast, prostate, colorectal and ovarian tumours. Excessive amounts of IGF2 secreted by some tumours, mainly sarcoma, can lead to activation of IR(A) and cause non-islet cell tumour induced hypoglycaemia. Approaches aiming to counteract the activity of IGF2 by targeting IGF1R often fail due to IGF2-mediated activation of IR(A). A ligand trap that exploits the high specificity and affinity of IGF2R for IGF2 has been designed by the fusion of a mutated form of human domain 11 of IGF2R to a C-terminal human IgG1 Fc domain (Fc-dom11). For the ligand trap to function as a clinically useful IGF2 super-antagonist, the binding affinity of domain 11 to IGF2 was further improved. The binding affinity optimisation strategy consisted of random mutagenesis of domain 11 loops involved in IGF2 binding, followed by yeast surface display in P. Pastoris, FACS and SPR screening, as well as structure-directed mutagenesis, complemented with NMR studies. Several mutants showing enhanced IGF2 binding have been identified, the combination of separately identified mutations improving affinity up to 100 fold. In particular, the mutant domain 113-4D shows an increase in affinity through both on- and off-rate modification attributed to the decrease in flexibility of one of the loops involved in IGF2 binding, as suggested by NMR results. The ability of Fc-dom113-4D to function as an IGF2 antagonist has been tested in vitro, using hepatocellular carcinoma cell lines, and in vivo, where it counteracts IGF2-induced hypoglycaemia in mice.

Citation Information: Mol Cancer Ther 2013;12(11 Suppl):A246.

Citation Format: Susana Frago, Madeleine Strickland, Jennifer Hughes, Christopher Williams, Ryan Nicholls, Lee Garner, Dellel Rezgui, Matthew P. Crump, A. Bassim Hassan. Development of an IGF2 super-antagonist by directed evolution of IGF2R. [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2013 Oct 19-23; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2013;12(11 Suppl):Abstract nr A246.
Abstract A259: Characterization of next generation (Gen2.5) KRAS antisense inhibitors in NSCLC, PDAC, and CRC models.
Authors: Revenko Alexey S. , Bhattachrjee Gourab , May Chris , Monia Brett P. , MacLeod A. Robert
Published: August 2014

KRAS is among the first described oncogenes and one of the most frequently mutated genes in human solid tumors. Despite substantial efforts to identify specific KRAS inhibitors, no effective therapeutic strategies to target KRAS directly have been identified. Here we describe the selective inhibition of KRAS with next generation (Gen2.5) antisense oligonucleotides.

We recently demonstrated preclinical and clinical activity of a next-generation constrained-ethyl (cEt) modified (Gen 2.5) antisense oligonucleotides (ASOs) targeting STAT3 mRNA (STAT3Rx/AZD9150). Systemic delivery of unformulated STAT3Rx/AZD9150 produced robust target RNA and protein reductions in a broad range of xenograft models (AACR 2013). More importantly, STAT3Rx/AZD9150 showed striking clinical activity in the phase I dose escalation study, producing anti-tumor activity in 4 out of 6 patients with advanced treatment-refractory lymphoma (ASCO 2013). In this study, we generated a panel of high affinity Gen2.5 ASO targeting KRAS. Anti-proliferative effects of the KRAS ASOs were evaluated on a panel of colorectal carcinoma (CRC), non-small cell lung carcinoma (NSCLC) and pancreatic ductal adenocarcinoma (PDAC) cell lines. KRAS ASOs potently and selectively inhibited growth of KRAS mutant CRC lines but did not affect the growth of KRAS WT CRC cells in 2D growth assays. Evaluation of KRAS depletion in NSCLC and PDAC lines revealed that sensitivity of these lines to growth inhibition in 2D assays required mutant KRAS status and the presence of a previously described “KRAS addiction” gene expression signature (Singh et al 2009). Surprisingly, further evaluation of KRAS ASO mediated depletion of NSCLC and PDAC lines in 3D growth assays (e.g. soft agar colony formation and polystyrene sponge (Alvetex)) resulted in strong inhibition of colony formation in all mutant KRAS lines, but not WT KRAS lines, and the sensitivity to KRAS depletion was independent of the expression of the “KRAS addiction” signature.

Collectively, we have shown that KRAS mutational status confers specific sensitivity to the ASO-mediated KRAS knockdown (KRAS oncogene addiction). Our data also highlights that antisense-mediated inhibition of KRAS may represent a promising and translatable approach to target KRAS therapeutically.

Citation Information: Mol Cancer Ther 2013;12(11 Suppl):A259.

Citation Format: Alexey S. Revenko, Gourab Bhattachrjee, Chris May, Brett P. Monia, A. Robert MacLeod. Characterization of next generation (Gen2.5) KRAS antisense inhibitors in NSCLC, PDAC, and CRC models. [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2013 Oct 19-23; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2013;12(11 Suppl):Abstract nr A259.
Abstract A265: L-methionine restriction sensitizes triple-negative breast cancer cells to a TRAIL receptor-2 agonist by reducing the expression of the melanoma-associated antigen MAGE-D2.
Authors: Strekalova Elena , Malin Dmitry , Good David , Cryns Vincent
Published: August 2014

The prognosis for most women with metastatic breast cancer remains poor due to a dearth of effective adjuvant therapies for this advanced stage of the disease. We have previously demonstrated that targeting TNF-related apoptosis-inducing ligand receptor-2 (TRAIL-R2) suppressed metastases in a murine model of triple-negative breast cancer (TNBC). However, many TNBCs are resistant to TRAIL receptor agonists. Restriction of the essential amino acid L-methionine has been reported to inhibit growth of transformed cells in preclinical models. We postulated that methionine restriction would sensitize TNBC cells to TRAIL-R2 agonists such as the humanized monoclonal antibody lexatumumab. Here we demonstrate that methionine restriction sensitized TNBC cells to lexatumumab-induced caspase activation and apoptosis. Intriguingly, methionine restriction also sensitized transformed MCF10A-RasV12 cells, but not untransformed MCF10A-Vector cells, to lexatumumab. Methionine restriction enhanced the cell surface expression of TRAIL-R2. To delineate the underlying mechanism(s), we performed proteomics analyses of TNBC cells grown in methionine restricted media and observed that methionine restriction dramatically reduced the expression of the melanoma-associated antigen MAGE-D2, which has recently been reported to inhibit TRAIL-R2 expression. siRNAs targeting MAGE-D2 mimicked the actions of methionine restriction by augmenting cell surface expression of TRAIL-R2 and sensitizing TNBC cells to lexatumumab. Overall, our results demonstrate that methionine restriction sensitizes TNBC cells to TRAIL-R2 agonists and suggest that dietary methionine restriction may be a strategy to augment the antitumor effects of TRAIL receptor agonists in vivo.

Citation Information: Mol Cancer Ther 2013;12(11 Suppl):A265.

Citation Format: Elena Strekalova, Dmitry Malin, David Good, Vincent Cryns. L-methionine restriction sensitizes triple-negative breast cancer cells to a TRAIL receptor-2 agonist by reducing the expression of the melanoma-associated antigen MAGE-D2. [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2013 Oct 19-23; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2013;12(11 Suppl):Abstract nr A265.
Abstract A268: TAS-119, a selective Aurora A inhibitor, enhanced the antitumor efficacy of taxanes in multiple human tumor cell lines including paclitaxel-resistant cells.
Authors: Miura Akihiro , Sootome Hiroshi , Ishihara Keiko , Masuko Norio , Hirai Hiroshi , Utsugi Teruhiro
Published: August 2014

Background: Aurora A is a serine/ threonine kinase that plays a critical role in regulating multiple mitosis processes. Aberrant expression of Aurora A has been associated with spindle checkpoint dysfunction and increased resistance to the microtubule-polymerizing agents, paclitaxel and docetaxel (1). Aurora A has been identified as one of the genes associated with enhancement of paclitaxel cytotoxicity in a siRNA library screen (2). Moreover, Aurora A overexpression/ gene amplification has been reported in various types of human cancers. In this report, we demonstrate that a selective Aurora kinase A inhibitor, TAS-119, synergistically enhances the activity of paclitaxel in multiple cell lines in vitro.

Materials and Methods: Evaluation of synergistic effects in combination was conducted by curve-shift analysis and Bliss additivity model, based on cell viability. Cell viability was assayed by using CellTiter-Glo™ which measures cellular ATP. Selectivity of TAS-119 for three isotypes of Aurora kinases was determined via monitoring the auto-phosphorylation of kinases by Western blot. Apoptosis induction was assessed by cleaved-PARP detection. As paclitaxel-resistant cell lines, both A549.T12 (3) and PTX10 (4) were used in combination experiments. A549.T12 and PTX10 were reported to have altered expression of β-tubulin isotypes and β-tubulin mutation, respectively.

Results: TAS-119 selectively inhibited cellular auto-phosphorylation of Aurora A without affecting the activity of both Aurora B and Aurora C. TAS-119 showed a synergistic effect with paclitaxel or docetaxel in multiple tumor cell lines from different origins, even in cells insensitive to TAS-119 treatment alone. Furthermore, TAS-119 also showed synergistic effects in paclitaxel-resistant cell lines. Cleaved-PARP analysis revealed that the cytotoxic synergism in combination with paclitaxel was induced by promoting the caspase-dependent apoptotic cell death.

Conclusion: The unique characteristic of TAS-119 as a highly selective Aurora A inhibitor contributed to the synergistic enhancement of antiproliferative effects of taxanes in multiple cell lines, possibly mediated by apoptotic pathways. These findings provide a rationale for the clinical development of TAS-119 as a combination therapy with taxanes.

Citation Information: Mol Cancer Ther 2013;12(11 Suppl):A268.

Citation Format: Akihiro Miura, Hiroshi Sootome, Keiko Ishihara, Norio Masuko, Hiroshi Hirai, Teruhiro Utsugi. TAS-119, a selective Aurora A inhibitor, enhanced the antitumor efficacy of taxanes in multiple human tumor cell lines including paclitaxel-resistant cells. [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2013 Oct 19-23; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2013;12(11 Suppl):Abstract nr A268.
Abstract A287: Discovery of c-Met kinase inhibitors for anticancer therapeutics.
Authors: Yun Chang-Soo , Cho Sung Yun , Kim Hyoung Rae , Jung Hee Jung , Ha Jae D. , Lee Kwangho , Kim Pilho , Park Chi Hoon , Lee Chong Ock
Published: August 2014

c-Met recepter tyrosine kinase (RTK) and its ligand, hepatocyte growth factor (HGF), have been reported to be involved in tumorigenesis and metastatic progression. Also, aberrant c-Met signaling has been reported in a wide variety of cancers such as gastric, lung, colon, breast, bladder, head and neck, ovarian, prostate, thyroid, pancreatic, hematological malignancies, and central nervous system tumors. We have synthesized a series of triazolopyridazine derivatives and their c-Met (mesenchymal-epithelial transition factor) inhibitory activities have been evaluated. A potent c-Met kinase inhibitor by structural modification of the parent pyrimidine scaffold with particular focus on the aryl substituent on the triazolopyridazines will be discussed. Several triazolopyridazine derivatives were found to be potent as c-Met inhibitors in enzyme assays and cell-based assays (c-Met addicted cell lines). We found KRC-00831 is a highly potent and selective c-Met receptor tyrosine kinase inhibitor over other kinases (c-Met enzyme, IC50=3 nM) and exhibit excellent cellular activities in c-Met driven cell lines (Hs746T, GI50=0.1 nM; H1993, GI50=27 nM, MKN45, GI50=15 nM; SNU-5, GI50=8 nM). KRC-00831 is chemically and metabolically very stable and showed no CYP inhibition, hERG binding activity. KRC-00831 strongly suppressed the growth of c-Met over-expressed cancer cells, while not in c-Met absent cancer cell lines. In xenograft animal models, we observed not only dose dependent tumor growth inhibitions but complete tumor regressions. In summary, we suggest that KRC-00831 wil be a novel drug candidate with the therapeutic potential of targeting c-Met in human cancer.

Citation Information: Mol Cancer Ther 2013;12(11 Suppl):A287.

Citation Format: Chang-Soo Yun, Sung Yun Cho, Hyoung Rae Kim, Hee Jung Jung, Jae Du Ha, Kwangho Lee, Pilho Kim, Chi Hoon Park, Chong Ock Lee. Discovery of c-Met kinase inhibitors for anticancer therapeutics. [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2013 Oct 19-23; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2013;12(11 Suppl):Abstract nr A287.
Abstract A291: A highly selective dual insulin receptor (IR)/insulin-like growth factor 1 receptor (IGF-1R) inhibitor derived from an ERK inhibitor.
Authors: Duong-Ly Krisna C. , Anastassiadis Theonie , Deacon Sean W. , Lafontant Alec , Ma Haiching , Devarajan Karthik , Dunbrack Roland L. , Wu Jinhua , Peterson Jeffrey R.
Published: August 2014

Dual inhibitors of the closely related receptor tyrosine kinases insulin-like growth factor 1 receptor (IGF-1R) and insulin receptor (IR) are promising therapeutic agents in cancer. Here we report an unusually selective class of dual inhibitors of IGF-1R and IR identified in a parallel screen of known kinase inhibitors against a panel of 300 human protein kinases. Biochemical and structural studies indicate that this class achieves its high selectivity by binding to the ATP-binding pocket of inactive, unphosphorylated IGF-1R/IR and stabilizing the activation loop in a native-like inactive conformation. One member of this compound family was originally reported as an inhibitor of the serine/threonine kinase ERK, a kinase that is distinct in the structure of its unphosphorylated/inactive form from IR/IGF-1R. Remarkably, this compound binds to the ATP-binding pocket of ERK in an entirely different conformation to that of IGF-1R/IR, explaining the potency against these two structurally distinct kinase families. These findings suggest a novel approach to polypharmacology in which two or more unrelated kinases are inhibited by a single compound that targets different conformations of each target kinase.

Citation Information: Mol Cancer Ther 2013;12(11 Suppl):A291.

Citation Format: Krisna C. Duong-Ly, Theonie Anastassiadis, Sean W. Deacon, Alec Lafontant, Haiching Ma, Karthik Devarajan, Roland L. Dunbrack, Jinhua Wu, Jeffrey R. Peterson. A highly selective dual insulin receptor (IR)/insulin-like growth factor 1 receptor (IGF-1R) inhibitor derived from an ERK inhibitor. [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2013 Oct 19-23; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2013;12(11 Suppl):Abstract nr A291.
Abstract A297: Microtubule targeting antimitotic drugs induce a lower mitotic arrest than clinically less effective antimitotic Eg5 inhibitors in mouse xenografts.
Authors: Florian Stefan , Chittajallu Deepak R. , Kohler Rainer H. , Orth James D. , Sorger Peter K. , Weissleder Ralph , Danuser Gaudenz , Mitchison Timothy J.
Published: August 2014

Despite immense progress in targeted cancer drug design, to date, the most important drugs in chemotherapy remain traditional cytotoxic drugs which are thought to kill proliferating cells, no matter if malignant of not. This "antiproliferative" hypothesis is based on results from the 1970s coming from mouse leukemia models or cell culture experiments. Microtubule binding drugs are clinically important antiproliferative drugs which arrest cultured cancer cells in mitosis. As opposed to empirically found microtubule binding drugs, newer drugs designed from scratch to induce mitotic arrest have repeatedly failed in clinical trials, suggesting that the antitumor effect of microtubule binding drugs is still not well understood. Alas, tissue culture assays are not predictive of their respective clinical success, suggesting that more realistic models are needed to understand why there are such huge differences in antitumor efficacy. Therefore, in this project, we used in vivo microscopy of mouse xenografts to compare the cell cycle effects of microtubule drugs to those of newer, targeted antimitotic drugs in the form of inhibitors of the mitotic kinesin Eg5. Image analysis with a custom developed software for automated three dimensional segmentation and cell cycle classification revealed that the microtubule interacting drugs Paclitaxel and Eribulin which are in wide clinical use induce a much lower mitotic arrest than the Eg5 inhibitor Ispinesib which failed clinical trials. This suggests that, rather than mitotic arrest, a different mechanism, intrinsically coupled to microtubule binding, might be essential for the antitumor effect of microtubule drugs in vivo.

Citation Information: Mol Cancer Ther 2013;12(11 Suppl):A297.

Citation Format: Stefan Florian, Deepak R. Chittajallu, Rainer H. Kohler, James D. Orth, Peter K. Sorger, Ralph Weissleder, Gaudenz Danuser, Timothy J. Mitchison. Microtubule targeting antimitotic drugs induce a lower mitotic arrest than clinically less effective antimitotic Eg5 inhibitors in mouse xenografts. [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2013 Oct 19-23; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2013;12(11 Suppl):Abstract nr A297.
Abstract A47: Study of promoter methylation pattern of 14-3-3 sigma gene in serum of patients with breast cancer: A potential biomarker for diagnostic anf prognostic factor.
Authors: Joaquina Martínez-Galán , Delgado Juan Ramón , Torres-Torres Blanca , Del Moral Rosario , Rios Sandra , Núñez M. Isabel
Published: August 2014

Background: Expression of 14-3-3 σ is a tumor suppressor gene induced in response to DNA damage, and has been implicated in G2/M cell cycle arrest by p53. Hypermethylation of CpG islands located in the promoter regions of tumor suppressor genes is now firmly established as an important mechanism for gene inactivation.

Objective: To correlation methylation levels of promoter 14-3-3σ with association prognostic factors in breast cancer.

Material and Methods: This is a prospective study we quantified methylation levels of promoter 14-3-3σ gene in 107 women with breast cancer and 108 control subjects by real time QMS-PCR SYBR green and analyzed association with prognostics factor in breast cancer.

Results: Median age was 58 years (32-88); 69% were postmenopausal women.Nodal involvement N0;63%,N1;30%,N2;7%), tumor size (T1;58%,T2;35%,T3;4%,T4;4%) and grade G1; 20%, G2; 37%, G3; 30%). The methylation of 14-3-3σ were 60% of sporadic breast cancer patients and were 34% of normal breast (p=0.0047).The methylation of 14-3-3σ gene in serum was markedly related with T3-4 stage (p<0.05),nodal positive status (p<0.05) and poor outcome. With a median follow up 6 years we saw more probability of developing distance metastasis in patients with methylation 14-3-3σ (p>0.05).

Conclusions: Hypermethylation of the 14-3-3σ a promoter is an early and frequent event in breast neoplastic transformation, leading to the suggestion that silencing of 14-3-3σ may be an important event in tumor progression and particularly in breast carcinogenesis.Therefore, it is possible that loss of σ expression contributes to malignant transformation by impairing the G2 cell cycle checkpoint function, thus allowing an accumulation of genetic defects. Perhaps in the detection of CpG methylation of 14-3-3σ may be used for diagnostic and prognostic purposes.

Citation Information: Mol Cancer Ther 2013;12(11 Suppl):A47.

Citation Format: Martínez-Galán Joaquina, Juan Ramón Delgado, Blanca Torres-Torres, Rosario Del Moral, Sandra Rios, M. Isabel Núñez. Study of promoter methylation pattern of 14-3-3 sigma gene in serum of patients with breast cancer: A potential biomarker for diagnostic anf prognostic factor. [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2013 Oct 19-23; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2013;12(11 Suppl):Abstract nr A47.
Abstract A50: Epstein-Barr virus in the putative stem cell fraction of some multiple myeloma cell lines.
Authors: Biswas Sunetra , Shamay Meir , Shirley Courtney M. , Yonescu Raluca , Matsui William H. , Borrello Ivan , Jones Richard J. , Ambinder Richard F.
Published: August 2014

Epstein-Barr virus (EBV) associated cancers have traditionally been thought to harbor the viral genomes as nuclear plasmids in all or nearly all of a patient's tumor cells as determined by in situ hybridization (ISH). We discovered EBV in 4 out of 7 standard multiple myeloma (MM) cell lines. In these positive cell lines, EBV was found in only a subpopulation of cells. For example, in MM.1S (a MM cell line) less than 1% of the cells harbored viral DNA as determined by limiting dilution PCR. These rare infected cells carried approximately 20 EBV copies. Fluorescence in situ hybridization (FISH) confirmed the presence of multiple copies of the EBV genome in rare cells. Immunofluorescence similarly detected the presence of viral genomes expressing EBV nuclear antigen (EBNA) - 1 and 2 in rare cells. Reverse transcriptase PCR confirmed EBNA1 and EBNA2 viral RNA expression. The cells that harbor virus are phenotypically distinct CD19+CD138− B cells. The phenotype overlaps with that identified as a MM cancer stem cell population. Viral genomes were not detected in the plasma cell-like population (CD19−CD138+). The results are consistent with the possibility that EBV is present in MM cancer stem cells in some MM cell lines and that virus is lost as these MM cancer stem cells with a B cell-like phenotype differentiate into plasma cells. Introduction of a dominant negative shRNA of the EBV EBNA-1, a protein essential for viral replication, decreased the proliferation of only the MM cell lines which were positive for EBV. Insertion of the dominant negative EBNA-1 shRNA had no effect on EBV negative MM cell lines, suggesting that EBV plays a role in the growth kinetics of cell lines even when present in a tiny percentage of cells. These results suggest that in some MM lines, EBV is harbored in a B cell-like progenitor population that overlaps with the putative cancer stem cell fraction. The loss of EBV inhibits cell proliferation and that EBV is lost during the B-cell to plasma cell differentiation process. The results presented here suggest that identifying the EBV status of MM patients prior to treatment might lead to a more efficient course of therapy.

Citation Information: Mol Cancer Ther 2013;12(11 Suppl):A50.

Citation Format: Sunetra Biswas, Meir Shamay, Courtney M. Shirley, Raluca Yonescu, William H. Matsui, Ivan Borrello, Richard J. Jones, Richard F. Ambinder. Epstein-Barr virus in the putative stem cell fraction of some multiple myeloma cell lines. [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2013 Oct 19-23; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2013;12(11 Suppl):Abstract nr A50.
Abstract A90: Feedback activation of EGFR drives resistance to BRAF(V600E) inhibition in breast cancer.
Authors: Yang Yj , Na HJ , Kim Ws , Bong JP , Beak SJ , Kim Jw , Park Jk , Hong Hj , Shin Dy , Kwon Jh , Lee Ej , Koh Yw , Choi Ec
Published: August 2014

Purpose: The BRAFV600E mutation has shown significant activity in the several tumor types, including breast, melanoma, and colorectal cancer. The selective inhibitor of BRAFV600, PLX4032 (vemurafenib), has shown clinical response in patients with BRAFV600Emelanoma, especially but not in 20-40% of patients despite the presence of a BRAFV600E. Thus, it is important to understand the molecular mechanisms of resistance to improve outcomes in patient with BRAF mutation. We aimed to investigate the cause of the limited therapeutic effect of PLX4032 in cancers harboring the mutant BRAFV600E gene.

Experiment Design: We investigated the correlation between RTK pathway activation and sensitivity to PLX4032 in several types of cancer cell lines harboring the BRAFV600E. Cell lines were exposed to PLX4032 and pharmacologic inhibitors and in combination with each other to inhibit RTK signaling. Cell survival and apoptosis were measured using MTT assay, Annexin V/PI staining, and colony forming assay. Real-time PCR, Western blots were performed to determine possible mechanisms of resistance and predictors of synergy.Results: In a comparison with BRAFV600E cancer cells, we show that PLX4032-resistant MDA-MB breast cancer cells have the enhanced level of p-EGFR in the presence of PLX4032. PLX4032 treatment causes a rapid feedback activation of EGFR and up-regulation of AKT in MDA-MB cells, which supports continued proliferation in the presence of BRAFV600E inhibition. We show that EGFR activation acquired resistance to BRAF inhibition using EGFR inhibitor. Gefitinib, EGFR inhibitor, shows strong synergy with a BRAFV600E inhibitor in short-time and long-time treatment. In addition, we investigated that rapid feedback activation of EGFR regulates the expression of epithelial to mesenchymal transition (EMT) regulators by

Real-time PCR, which resulted in high migratory potentials in invasion assay and wound healing assay. These results provide rational therapeutic strategies for clinical studies in this poor prognosis subtype of BRAFV600E inhibitor.

Conclusions: Our data suggest that BRAFV600E mutant breast cancers, for which there are currently no targeted treatment options available, might benefit from combination therapy consisting of BRAF and EGFR inhibitors.

Citation Information: Mol Cancer Ther 2013;12(11 Suppl):A90.

Citation Format: Yj Yang, HJ Na, Ws Kim, JP Bong, SJ Beak, Jw Kim, Jk Park, Hj Hong, Dy Shin, Jh Kwon, Ej Lee, Yw Koh, Ec Choi. Feedback activation of EGFR drives resistance to BRAF(V600E) inhibition in breast cancer. [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2013 Oct 19-23; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2013;12(11 Suppl):Abstract nr A90.
Abstract B109: The role of SIRT6 in drug resistance and cellular senescence in cancer.
Authors: Khongkow Mattaka , Lam Eric
Published: August 2014

Introduction: The Sirtuin family consists of 7 members. They comprize highly conserved NAD+-dependent deacetylases that play an important role in a board spectrum of cellular processes, including metabolism, differentiation, survival and cellular senescence. It has also been suggested that these proteins are central to cancer development and drug resistance. Recent studies have documented that SIRT6, a nuclear chromatin-associated protein, has a key role in genomic stability and degenerative pathologies associated with ageing. Moreover, SIRT6 has been reported to have a function in the regulation of DNA damage repair. Therefore, this research aims to investigate the role of SIRT6 in drug resistance and cellular senescence in cancer cells.

Methods: Cell lines that we have used in this research consist of sensitive and resistant breast cancer cells, Hela cells and mouse embryonic fibroblasts (MEFs). To investigate the role of SIRT6 in drug resistance, we treated cells with either epirubicin or paclitaxel, then performed immunofluorescent, comet and repair assays to investigate DNA damage repair. Clonogenic and beta-galactosidase assays were also used to investigate long-term survival and cellular senescence.

Results: We found that SIRT6 is elevated in drug resistant cells compared to drug sensitive parental cells and that SIRT6 mediates drug resistance by increasing DNA damage repair. Sirt6-/- MEFs have lower levels of γH2AX accumulation compared to WT MEFs. These higher levels of SIRT6 are also associated with the lower activities of p53 and FOXO3a which are two important cell cycle/survival regulators. Here, we show that depletion of SIRT6 in cancer cells results in a decrease in long-term survival and lower HR and NHEJ DNA repair activity. SIRT6-deficeint cells also exhibit the induction of senescence-associated β-galactosidase activity and flat cell morphology.

Conclusion: Our results suggest a novel role of SIRT6 in epirubicin and paclitaxel treatment resistance and cellular senescence. SIRT6 could be a potential biomarker for identifying tumors that are likely to be resistant to these chemotherapeutic agents. It might be also a useful therapeutic target for the prevention of acquired drug resistance and for improving the efficiency of drug treatment.

Citation Information: Mol Cancer Ther 2013;12(11 Suppl):B109.

Citation Format: Mattaka Khongkow, Eric Lam. The role of SIRT6 in drug resistance and cellular senescence in cancer. [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2013 Oct 19-23; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2013;12(11 Suppl):Abstract nr B109.
Abstract B11: In vivo ADCC-mediated antitumor activity study with new preclinical humanized models.
Authors: Mignard Caroline , Duchamp Olivier , Mirjolet Jean-François , Ohnishi Yasuyuki , KATANO Ikumi , ITO Mamoru
Published: August 2014

Experimental systems to effectively evaluate therapeutic antibodies are urgently needed. One of the mechanisms used by antibodies to kill tumor cells is antibody-dependent cellular cytotoxicity (ADCC) in which natural killer cells (NK) are the main mediator.

The generation of humanized mice as immunodeficient mice (such as NOG) engrafted with hematopoietic stem cells (HSC) demonstrated their usefulness for studying the human immune system. Nevertheless, one issue of this humanized model is the incomplete maturation of NK cells.

To circumvent this lack of functionality the Central Institute for Experimental Animals (CIEA) has developed the second generation of transgenic NOG mice expressing human IL2 or IL15 (NK-NOG). hIL-2 and hIL-15 NOG mice were generated by microinjection of DNA fragments containing human IL-2 cDNA or human IL-15 cDNA replaced with a hIL-2 signal peptide, under the control of a CMV-promoter, into fertilized eggs of NOD-IL-2Rgnull mice. The serum levels of hIL-2 and hIL-15 in mice were 1-2 and 0.1-0.2 ng/mL, respectively.

Humanization of NK-NOG mice was validated using freshly collected or frozen hematopoietic stem cells (HSCs) in transgenic NK-NOG mice. The characterization of the reconstituted NK-NOG mice was done in peripheral blood as well as central lymphoid organs such as bone marrow and spleen using flow cytometry. Moreover, the human NK cell function was evidenced using an ex vivo 51Cr assay demonstrating lysis properties of NK cells from collected spleen.

Finally in vivo NK-mediated ADCC was evaluated in IL2-NOG mice simultaneously engrafted with human immune system and human Burkitt's lymphoma. Reconstituted mice were treated with Rituximab to examine immune system-related antitumor activity.

Genetic manipulation of the host continue to improve the ability of humanized mice to more accurately recapitulate the in vivo function of human immune cells such as NK cells.

Prediction of ADCC-mediated antitumor activity of humanized antibodies is now routinely feasible for preclinical evaluation.

Citation Information: Mol Cancer Ther 2013;12(11 Suppl):B11.

Citation Format: Caroline Mignard, Olivier Duchamp, Jean-François Mirjolet, Yasuyuki Ohnishi, Ikumi KATANO, Mamoru ITO. In vivo ADCC-mediated antitumor activity study with new preclinical humanized models. [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2013 Oct 19-23; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2013;12(11 Suppl):Abstract nr B11.