bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract A198: A novel regulatory circuit involving STIM1 and HIF-1 mediates hypoxia-driven hepatocarcinogenesis
Authors: Li Yongsheng
Published: May 2016

Hypoxia and intracellular Ca2+ transient are fundamental traits of cancer. Signaling cascades initiated or regulated by HIF-1 is essential for hypoxic responses. Stromal-interaction molecule 1 (STIM1) is well known as a Ca2+ sensor on endoplasmic reticulum (ER) that mediates SOCE activation and promotes tumor invasion and migration. However, there was no general consensus about the relationship between HIF-1 and STIM1. Also, the significance of STIM1, as well as the Ca2+ mobilization route in cancer cells during hypoxia remains unknown. Our results show that STIM1 correlates with elevated hypoxia-inducible factor-1 alpha (HIF-1α) in hypoxic hepatocarcinoma cells (HCC) and is upregulated during hepatocarcinoma growth. HIF-1 directly transcripts STIM1 and contributes to store-operated Ca2+ entry (SOCE), while STIM1-mediated SOCE is also required for HIF-1 accumulation in hypoxic HCC, via activating Ca2+/calmodulin-dependent protein kinase II and p300. Administration of YC-1, a HIF-1 inhibitor, or knockdown of HIF1A significantly diminishes hypoxia-enhanced STIM1 and suppresses tumorigenesis. Moreover, ectopic expression of STIM1 or HIF-1α partially reverses the impaired growth of the tumor treated with YC-1. These results suggest a mutual dependency and regulation of STIM1 and HIF-1 in controlling Ca2+ mobilization and hypoxic tumor growth, and highlight a potential target for early hypoxia-related intervention.

Citation Format: Yongsheng Li. A novel regulatory circuit involving STIM1 and HIF-1 mediates hypoxia-driven hepatocarcinogenesis. [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2015 Nov 5-9; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2015;14(12 Suppl 2):Abstract nr A198.
Abstract A33: Utility of multiplexed biomarker analysis in non-small-cell lung cancer patients with discrepancies between FISH and immunohistochemical results of ALK testing
Authors: KIM TAE-JUNG , TERAI HIDEKI
Published: May 2016

Background: The clinical implications in Non-small cell lung cancer (NSCLC) patients with discrepancy between fluorescent in situ hybridization (FISH) and immunohistochemistry (IHC) assessing ALK status are unclear.

Materials and Methods: From 1131 archived tissue samples, we identified 41 NSCLC patients with discrepant results showing positive or borderline ALK FISH and negative IHC using ALK1 antibody. Of these, 21 cases with sufficient tissue remaining for selected reaction monitoring (SRM) and an additional case within normal range FISH split (4%) were sent to confirm ALK IHC using other antibodies (5A4 and D5F3). Target protein levels measured by multiplexed SRM were correlated with records of clinical outcomes after treatment with crizotinib or cytotoxic chemotherapy.

Results: Of 22 cases with various ALK FISH results and negative IHC (ALK1), IHC using other antibodies (5A4 and D5F3) identified 4 positive cases, whereas the SRM assay identified 7 cases with tumor-specific ALK protein. IHC showed lower sensitivity in specimen with delayed fixation but SRM showed superior sensitivity in those samples as SRM is independent to modified epitope. Of 8 crizotinib-treated cases, 4 cases with detectable levels of either ALK or cMET proteins had complete or partial responses; cases without either of these proteins (n = 4) did not respond to crizotinib regardless of ALK FISH results. Cases with 3 or more predictive proteins specific to chemotherapeutic agents were found to be more responded to those agents and had a higher probability of progression-free survival.

Conclusions: The SRM assay showed superior sensitivity in poorly fixed specimens and can inform therapeutic decision-making in NCSLC patients with ALK screening discordances.

Citation Format: TAE-JUNG KIM, HIDEKI TERAI. Utility of multiplexed biomarker analysis in non-small-cell lung cancer patients with discrepancies between FISH and immunohistochemical results of ALK testing. [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2015 Nov 5-9; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2015;14(12 Suppl 2):Abstract nr A33.
Abstract A39: 3,4',5- Trismethoxybenzophenone inhibits the growth of human hepatocarcinoma cells via cell-cycle arrest at the G2/M phase
Authors: Patton Christopher J. , Khadka Pritika , Kotturi Hari
Published: May 2016

3,4',5-trismethoxybenzophenone (TMBP) is an analogue of the natural dietary polyphenolic compound resveratrol (3,5,4'-trihydro-trans-stilbene, abbreviated as RVT). Natural RVT belongs to the phytoalexin class of phytochemicals that are antibiotics produced by plants in response to stimuli such as injury, ultraviolet irradiation, stress, and fungal attack. RVT and its analogues have also been shown to prevent hepatic steatosis. In this study we focused on TMBP which is a methylated derivative of RVT. Studies have shown that TMBP exhibits added biological effects and a better pharmacokinetic profile compared to RVT due to the presence of methoxy groups. The human hepatoma cell line (FCA4 cells) that harbors a subgenomic selectable HCV replicon was used for this study. The Huh7 cell line lacking the replicon served as the corresponding control. MTS assay was used to determine the cytotoxicity of TMBP at various concentrations. Wound healing assay was performed to determine the effect of the analogue on cell migration. Flow cytometry and magnetic levitation were used to measure the effect of compound on cell cycle and 3D spheroid formation. Our data showed that treating liver cancer cells with TMBP resulted in a significant dose and time dependent growth inhibition combined with G2/M-phase cell cycle arrest at 5 μM with (IC50) at 7.5 μM. TMBP at 5μM inhibited the formation of 3D spheroids, and also repressed the migration of cancer cells. Current therapies available for treating hepatocellular carcinoma (HCC) have a high incidence of recurrence and postoperative death in patients. Exploring different strategies with high pharmaceutical function and low toxicity is of great urgency for treating HCC. Our studies show that we have identified a promising analogue for treating HCC.

Citation Format: Christopher J. Patton, Pritika Khadka, Hari Kotturi. 3,4',5- Trismethoxybenzophenone inhibits the growth of human hepatocarcinoma cells via cell-cycle arrest at the G2/M phase. [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2015 Nov 5-9; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2015;14(12 Suppl 2):Abstract nr A39.
Abstract A7: Drug resistant murine tumor models facilitate development of next generation anticancer therapeutics
Authors: Jin Ying , Zhang Juan , Zhu Dandan , Sun Yanmei , Mao Guanping , Wang Minxia , Shi Qian
Published: May 2016

Despite the success in the development of new anti-cancer drugs, all cancer therapeutics, either conventional cytotoxic chemo- or targeted therapeutics, are limited by the drug resistance. Resistance may be divided into two broad categories: intrinsic/de novo or acquired, involving various mechanisms. Certain patients are de novo resistant to a specific therapy, while others initially responded, but eventually recurred with resistant tumors that no longer respond to the original therapy. Even with the newly developed immunotherapies that promise to have long lasting tumor inhibitory effect in patients, a large percentage of patients still do not respond to the therapy. To overcome the resistance to the existing drugs, effective new strategies, e.g. next-generation selective inhibitors, rational drug combinations, and new drug delivery or formulation designs, are developed and under active preclinical and clinical investigations. To enable a more informative selection of resistant models for in vivo preclinical drug evaluation, here we set out to identify a series of animal models, which are intrinsically resistant to either chemotherapy, targeted therapy, or immunotherapy, from more than 200 CrownBio validated cell line xenograft and syngeneic tumor models. On another hand, we also validated a panel of acquired drug resistant models, covering the resistance to Doxorubicin, Imatinib, Elotinib, Quizartinib, Crizotinib or Androgen ablation (castration), etc. The possible mechanisms of some resistance were discussed. In conclusion, these resistant models provide a valuable platform for the development of new strategies to overcome the drug resistance.

Citation Format: Ying Jin, Juan Zhang, Dandan Zhu, Yanmei Sun, Guanping Mao, Minxia Wang, Qian Shi. Drug resistant murine tumor models facilitate development of next generation anticancer therapeutics. [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2015 Nov 5-9; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2015;14(12 Suppl 2):Abstract nr A7.
Abstract A84: Intravital microscopy reveals biopsy induced local wound-healing response
Authors: Alieva Maria , Colak Burcin , Boquetale Carla , Van Rheenen Jacco
Published: May 2016

A biopsy is for most types of cancer the golden standard for a definitive cancer diagnosis, as it provides the most accurate analysis of tissues. However taking biopsies creates a wound thereby potentially changing the composition and architecture of the tissue. In non-pathological situations, wounded tissue ‘repairs’ itself by a cascade of cellular events, including the induction of an inflammatory response that promotes cell proliferation and migration of surrounding cells to close the wound. Proliferation and migration are two malignant processes that promote tumor progression and metastatic disease. Thus it is vital to identify the potential effect of biopsy on the behavior of cells that remain in the tumor.

By making use of repeated high-resolution intravital microscopy (IVM) we studied how the behavior of tumor cells locally changes upon biopsy. Our results showed that biopsy-like injury triggers a wound-healing response observed as increased tumor cell migration and proliferation. 24 hours after biopsy-like injury there was a 1.75 fold increase in migratory cells and a 1.52 fold increase in proliferating cells compared to the control group that did not received an injury. IVM showed that the observed effects were mediated by a CCL-2- dependent inflammation response. Interestingly, immune cell ablation by dexamethasone, a glucocorticoid commonly used to prevent inflammation resulted in decreased tumor cell motility and proliferation and suppressed the observed wound healing response. This result opens new therapeutic possibilities to prevent biopsy undesired effects on tumor cell behavior.

Taken together, our data show that biopsy induces a local wound-healing response in the tumor through an immune system mediated mechanism and illustrate that IVM is a powerful technique to study local and temporal changes in cell behavior that cannot be assessed by other techniques (e.g. immunohistochemistry) that only provide a snapshot of a dynamic tissue. Furthermore this tool can be useful to determine local and dynamic responses of (tumor) cells to other more aggressive clinical interventions, such as surgical resection or chemotherapeutic treatment.

Citation Format: Maria Alieva, Burcin Colak, Carla Boquetale, Jacco Van Rheenen. Intravital microscopy reveals biopsy induced local wound-healing response. [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2015 Nov 5-9; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2015;14(12 Suppl 2):Abstract nr A84.
Abstract B121: A novel oncogenic BTK isoform is overexpressed in colon cancers and required for RAS-mediated transformation
Authors: Grassilli Emanuela , Pisano Fabio , Cialdella Annamaria , Bonomo Sara , Missaglia Carola , Cerrito Maria Grazia , Masiero Laura , Ianzano Leonarda , Narloch Robert , D'Amato Filomena , Noli Barbara , Ferri Gian Luca , Leone Biagio E. , Stanta Giorgio , Bonin Serena , Helin Kristian , Giovannoni Roberto , Lavitrano Marialuisa
Published: May 2016

It is known that Bruton's tyrosine kinase (BTK) is essential for B-cell proliferation/differentiation and it is generally believed that its expression and function are limited to bone marrow-derived cells. Here, we report the identification and characterization of p65BTK, a novel oncogenic isoform expressed in colon carcinomas.

Standard procedures were used for cloning the full length p65BTK-encoding mRNA and raise anti-p65BTK specific polyclonal antibodies. Quantitative PCR, western blot, RNA immunoprecipitation, silencing experiments and fluorescence assay upon transfection with bi-cistronic vectors were used to demonstrate post-transcriptional regulation of p65BTK expression. Immunohystochemistry was employed to study p65BTK expression in colon cancer patients specimens. Soft agar assay and foci assay were carried out to assess p65BTK oncogenic properties. Cell growth, cell viability and colony assays were performed to study the effects of p65BTK inhibition (by a specific kinase inhibitor) on colon cancer cells biology.

We found that p65BTK differs from the already known 77 kDa isoform for the lack of the N-terminal PH domain and is translated - through an IRES-dependent mechanism -from a transcript containing an alternative first exon in the 5'UTR. Moreover, p65BTK mRNA translation requires phosho-hnRNPK binding to its cognate sites (located in the alternative first exon) and is post-transcriptionally regulated, via hnRNPK, by the MAPK pathway. We demonstrate that p65BTK is endowed with strong transforming activity that depends on active ERK1/2 and its inhibition abolishes RAS transforming activity. Accordingly, p65BTK overexpression in colon cancer tissues correlates with ERK1/2 activation. Finally we show that p65BTK inhibition affects growth and survival of colon cancer cells.

In conclusion, our data reveal that BTK, via p65BTK expression, is a novel and powerful oncogene acting downstream of the RAS/MAPK pathway and suggest that its targeting may be a promising therapeutic approach.

Citation Format: Emanuela Grassilli, Fabio Pisano, Annamaria Cialdella, Sara Bonomo, Carola Missaglia, Maria Grazia Cerrito, Laura Masiero, Leonarda Ianzano, Robert Narloch, Filomena D'Amato, Barbara Noli, Gian Luca Ferri, Biagio E. Leone, Giorgio Stanta, Serena Bonin, Kristian Helin, Roberto Giovannoni, Marialuisa Lavitrano. A novel oncogenic BTK isoform is overexpressed in colon cancers and required for RAS-mediated transformation. [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2015 Nov 5-9; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2015;14(12 Suppl 2):Abstract nr B121.
Abstract B142: A “moneyball” approach to predicting clinical trial failures and successes
Authors: Gayvert Kaitlyn , Madhukar Neel , Elemento Olivier
Published: May 2016

Over the past decade, significant strides have been made in the management and treatment of various diseases. Despite this progress, drug attrition rates due to clinical trial failure have continued to substantially rise. Clinical trials can fail for various reasons, ranging from design issues to drug efficacy and safety problems. Drug-likeness approaches, as first proposed by Lipinski almost two decades ago, have become a key tool for the pre-selection of compounds that are likely to have manageable toxicity in clinical studies. However all these methods consider molecular properties of the drug itself alone. In general, these approaches struggle to simultaneously well-characterize the properties of both FDA approved drugs (sensitivity) (and drugs that fail clinical trials (specificity).

We introduce a novel data-driven approach (PrOCTOR) that integrates chemical properties of a compound, along with that of its targets, to provide a new measure, the PrOCTOR score, that helps predict whether drugs in clinical trials will fail for toxicity reasons. When trained on failed Phase I clinical trials and FDA approved drugs, the PrOCTOR score performs at a high accuracy, specificity and sensitivity (∼0.75), as well as high area under the ROC curve (>0.80). In comparison, none of the drug-likeness approaches were able to successfully maintain both high sensitivity and specificity. Robust feature analysis of the model indicates that it is critical to consider properties of the drug target, with the target's network connectivity and liver toxicity as two of the most important features. The approach was further evaluated by testing the predictions of the trained model on an established independent dataset. We found that our method was able to significantly distinguish a representative set of bioavailable drugs from a representative set of toxic drugs (D = 0.2133, p<2.2e-16, Kolmogorov-Smirnov test). Additionally, we found that the measure is strongly correlated with severe toxicity events, such as pleural effusion (ρ = −0.9792) and neutropenia (ρ = −0.9613). Altogether, our method provides a novel, data-driven and broadly applicable strategy that is able to identify drugs likely to possess manageable toxicity in clinical trials.

Citation Format: Kaitlyn Gayvert, Neel Madhukar, Olivier Elemento. A “moneyball” approach to predicting clinical trial failures and successes. [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2015 Nov 5-9; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2015;14(12 Suppl 2):Abstract nr B142.
Abstract B146: Preclinical pharmacokinetic studies of RPT835, an allosteric FGFR2 inhibitor
Authors: Tsimafeyeu Ilya , Byakhov Mikhail , Bencetić Mihaljević Vlatka , Padovan Jasna , Murillo Genoveva , Tjulandin Sergei
Published: May 2016

RPT835 (recommended INN, alofanib) is a novel allosteric FGFR2 inhibitor with activity in FGFR2-expressing women's cancers. Administration of RPT835 intravenously (iv) daily demonstrated dramatic effect in ovarian cancer xenografts compared with RPT835 orally daily [S. Tjulandin et al. AACR 2015]. Here we explore the pharmacokinetic (PK) profile of RPT835.

Three preclinical PK studies were conducted. In Study 1, RPT835 was administered to 49 male BALB/C mice via two routes (oral and iv). Mice in group 1 received a single iv injection of the RPT835 (30 mg/kg), while mice in group 2 received a single dose of RPT835 (30 mg/kg) via oral gavage. The aim of the Study 2 was to evaluate the PK for RPT835 in 24 male Sprague Dawley rats after a single iv dose of 22 mg/kg and oral dosing in capsules at three dose levels (22, 110 and 220 mg/kg). In Study 3, PK of pharmaceutical form of RPT835 in male Sprague Dawley rats after a single iv dose of 55.3, 113.8 and 218.7 mg/kg was evaluated. Six animals per dose group were dosed, with plasma samples collected from the tail vein up to 24 hours. Plasma concentrations were quantified by LC-MS/MS using a research qualified method. The PK data are summarized in Table.

Following oral administration, RPT835 appeared rapidly in plasma (within 30 min), but could not be detected after 2 hours. Bioavailability for oral administration could not be calculated but is estimated to be low (<1%). Following single iv bolus dosing of RPT835, animals showed a moderate intra-individual variability in plasma levels. RPT835 plasma levels were quantifiable up to 8 hours post dose in all animals in Study 3. The compound resulted in a moderate and low clearance (32-12% of liver blood flow), small volume of distribution (0.26-0.5 L/kg) and short half-life (0.4-0.9 h). Based on results of PK, protocol of Phase IB study of continuous iv infusion of RPT835 via pump in patients with ovarian cancer was developed at the ECCO-AACR-EORTC-ESMO Workshop on Methods in Clinical Cancer Research (“Flims17”).

PK resultsStudyGroup (dose, mg/kg)C0/Cmax, ng/mlTmax, ht1/2AUC, h*ng/mLCL, mL/min/kgVss, L/kgF, %1iv, 3057739-0.93580386.76.9NC - Parameter cannot be calculatedoral/gavage, 3043.61-2NCNCNCNCNC2iv, 2279323-0.441423427.10.47-oral/caps, 2222.02.0NC68.0--0.36oral/caps, 11082.32.5NC1.0--0.18oral/caps, 220110.01.0NC1.0--0.183iv, 55.3246499-0.863710523.80.3-iv, 113.8432342-0.7313078814.80.29-iv, 218.7730496-0.6836314510.10.26-

Citation Format: Ilya Tsimafeyeu, Mikhail Byakhov, Vlatka Bencetić Mihaljević, Jasna Padovan, Genoveva Murillo, Sergei Tjulandin. Preclinical pharmacokinetic studies of RPT835, an allosteric FGFR2 inhibitor. [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2015 Nov 5-9; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2015;14(12 Suppl 2):Abstract nr B146.
Abstract B170: Discovery of tubulysin payloads for antibody drug conjugates with potent in vitro activity and in vivo efficacy in solid tumor models
Authors: Toader Dorin , Harper Jay , Lloyd Chris , Marwood Rose , Bannister David , Mao Shenlan , Chen Cui (Tracy) , Zhong Haihon (Helen) , Bedian Vahe , Wang Fengjiang , Gingipalli Lakshmaiah , Vasbinder Melisa , Thompson Pamela , Fleming Ryan , Bezabeh Byniam , Dimasi Nazzareno , Gao Changshou , Kamal Adeela
Published: May 2016

Antibody drug conjugates (ADCs) combine the specificity of antibodies with the potency of small molecule cytotoxic drugs and have the potential to provide significant efficacy as a treatment for cancer. The objective of this work was to identify potent new cytotoxic ADC payloads that can be used to target diverse tumor types. Here we report for the first time the discovery of fully synthetic tubulysin payloads which belong to a class of highly cytotoxic natural products that disrupt the cellular microtubule network leading to apoptosis of tumor cells. Our fully synthetic tubulysin payloads are comprised of: (i) a tubulysin warhead that displays pM potency, (ii) a protease cleavable amino-acid sequence and (iii) a tether bearing a reactive maleimide group. Tubulysin-based ADCs were generated via site-specific conjugation of these payloads to cysteines engineered into antibodies against cancer antigen target oncofetal protein 5T4. The resulting ADCs showed potent in vitro cell killing and in vivo efficacy in multiple solid tumor xenograft models including prostate cancer, non-small cell lung adenocarcinoma, breast cancer and gastric carcinoma. Furthermore, specific structural features of the tubulysin warhead, linker design and antibody engineering were shown to impact the overall in vitro and in vivo properties of the ADCs. Thus, these synthetic tubulysin payloads represent novel microtubule network disrupting compounds that display potent preclinical anti-tumor activity as an ADC that could be advanced to the clinic.

Citation Format: Dorin Toader, Jay Harper, Chris Lloyd, Rose Marwood, David Bannister, Shenlan Mao, Cui (Tracy) Chen, Haihon (Helen) Zhong, Vahe Bedian, Fengjiang Wang, Lakshmaiah Gingipalli, Melisa Vasbinder, Pamela Thompson, Ryan Fleming, Byniam Bezabeh, Nazzareno Dimasi, Changshou Gao, Adeela Kamal. Discovery of tubulysin payloads for antibody drug conjugates with potent in vitro activity and in vivo efficacy in solid tumor models. [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2015 Nov 5-9; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2015;14(12 Suppl 2):Abstract nr B170.
Abstract B186: ITRI-2531 - A novel kinase inhibitor for treating hepatocellular carcinoma
Authors: Kuo Tsung-Keng , Lee On , Lin Mai-Wei , Lin Li-Zong , Liu Chun-Min , Hsieh Tai-ju , Wang Chun-Chung , Lin Shyh-Horng , Chang Chia-Ni , Hsu Hui-Chun , Chou Nien-Tzu , Lai Chin-Pen , Chen Chih-Hung , Tu Chia-Mu , Chen Shih-Ta , Tsai Yuan-Jang , Liu Chih-Peng , Hwang Jenn-Tsang , Huang Jui-Wen , Chen Yen-Chun , Hwang Chrong-Shiong , Tseng Hsiang-Wen
Published: May 2016

Hepatocellular carcinoma (HCC) is one of the most common cancers and is also a leading cause of cancer-related deaths worldwide. Until now, treatments for HCC remain limited, especially for the shortage of promising systemic therapies to replace systemic chemotherapy. Sorafenib, the first drug applied in targeted therapy for HCC, was approved and drew considerable attention. However, the efficacy and side effects of sorafenib are still unsatisfied, which make sorafenib far from ideal for treatment of HCC. Thus, developing novel therapeutics for HCC is necessary. A novel compound, ITRI-2531, is developed to improve the efficacy from sorafenib. ITRI-2531 shows cytotoxicity in HCC cell lines (Huh-7 and PLC/PRF/5) and patient-derived HCC tissue cell lines. In studies of kinase activities, ITRI-2531 suppresses the phosphorylation of MEK and ERK in PLC/PRF/5 cells and shows inhibition of multiple kinases (FLT3, FLT4, PDGFRA, PGDFRB, and VEGFR2) in KINOMEscanTM study. In in vivo studies, oral treatment of ITRI-2531 repressed subcutaneous Huh-7 and PLC/PRF/5 tumor growth in severe combined immunodeficient (SCID) mice and shows better efficacy than sorafinib. Moreover, ITRI-2531 prolongs the survival of SCID mice with orthotopic patient-derived xenograft tumor and suppresses alpha-fetoprotein in serum. On the other hand, ITRI-2531 also has good pharmacokinetic profiles. According to these results, we believe that ITRI-2531 warrants further evaluation as a new anti-HCC drug.

Citation Format: Tsung-Keng Kuo, On Lee, Mai-Wei Lin, Li-Zong Lin, Chun-Min Liu, Tai-ju Hsieh, Chun-Chung Wang, Shyh-Horng Lin, Chia-Ni Chang, Hui-Chun Hsu, Nien-Tzu Chou, Chin-Pen Lai, Chih-Hung Chen, Chia-Mu Tu, Shih-Ta Chen, Yuan-Jang Tsai, Chih-Peng Liu, Jenn-Tsang Hwang, Jui-Wen Huang, Yen-Chun Chen, Chrong-Shiong Hwang, Hsiang-Wen Tseng. ITRI-2531 - A novel kinase inhibitor for treating hepatocellular carcinoma. [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2015 Nov 5-9; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2015;14(12 Suppl 2):Abstract nr B186.
Abstract B205: EphA10 targeted antibody therapy is a potential as a novel option for management of breast cancers including triple negative breast cancer
Authors: Takahide Mori , Kazuya Nagano , Tsunoda Shin-ichi , Tsutsumi Yasuo
Published: May 2016

Introduction: Triple negative breast cancer (TNBC) is defined by the absence of receptors for Her2, estrogen and progesterone and is often referred to as a refractory disease, because it is resistant to treatment with anti-Her2 antibody and anti-hormone drugs. Thus, new targets for therapeutic agents are urgently required. Using a proteomic approach, we have identified EphA10, which is more highly expressed in breast cancer, including TNBC, than in normal breast tissues. Moreover, we previously revealed that EphA10 is expressed in testis, but not in other normal tissues. This suggests that EphA10 could become a promising therapeutic target in breast cancer patients. Here, we describe a novel monoclonal antibody (mAb) against the extracellular region of EphA10 showing tumor accumulation and anti-tumor effects against EphA10-positive breast cancer.

Materials and Methods: Tumor accumulation analysis of anti-EphA10 mAb; Alexa647-conjugated anti-EphA10 mAb or control mAb were administrated intravenously and observed daily using an in vivo imaging system. Tumor accumulation was quantified as the fluorescence intensity ratio of the tumor compared to that of the contralateral side. Anti-EphA10 mAb treatment in a mouse xenograft model; A mouse model was constructed by orthotopic transplantation of EphA10 stably expressing MDA-MB435 cells. When the tumor size reached approximately 100 mm3, anti-EphA10 mAb and control mAb were administrated intraperitoneally twice a week.

Results and Discussion: Anti-EphA10 mAb we have developed by hybridoma methods displayed an affinity against EphA10 in the nanomolar range and specificity for EphA family proteins. Administration of the fluorescein-labeled mAb to EphA10-positive breast tumor-bearing mice showed that the mAb specifically accumulated in tumors, but not in normal organs. Furthermore, tumor growth was significantly suppressed in the mAb-treated mice in a dose-dependent manner. These data indicate that EphA10 targeted antibody therapy have a potential as a novel option for management of EphA10-positive breast cancer. We are currently generating human anti-EphA10 mAb by phage human antibody library method toward the clinical application.

Conclusion: EphA10 targeted antibody therapy is effective for a mouse xenograft model. We hope that these findings will contribute to the development of a novel drug for refractory breast cancer patients.

Citation Format: Mori Takahide, Nagano Kazuya, Shin-ichi Tsunoda, Yasuo Tsutsumi. EphA10 targeted antibody therapy is a potential as a novel option for management of breast cancers including triple negative breast cancer. [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2015 Nov 5-9; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2015;14(12 Suppl 2):Abstract nr B205.
Abstract B36: Human serum albumin and affibody fusion proteins for targeted drug delivery to HER2 positive cells
Authors: Dong Daoyuan , Li Zhijun , Li Zhiyu
Published: May 2016

Human Epidermal growth factor Receptor 2 (HER2) has been reported to be over-expressed in 20-30% of the breast cancer patients and related to poor prognosis. Affibody ZHER2 is a small protein which has high affinity to HER2 screened by phage display. Human Serum Albumin (HSA), as the main protein in plasma, has been commonly used to extend the small peptides serum half-life. Its high solubility, stability and excellent ability to carry multiple ligands in blood stream make it a good candidate for drug delivery. In this presentation, we report the development of albumin-affibody fusion proteins in the aim of targeted delivering chemotherapeutics to HER2 positive cells.

Two HSA fusion proteins have been successfully constructed, expressed and purified, one with a single ZHER2 domain at the albumin C terminus (rHSA-ZHER2) and the other has two tandem copies of ZHER2 (rHSA-(ZHER2)2). Both of the fusion proteins maintained the HER2 and fatty acid (FA) binding ability inherited respectively from the two parental proteins demonstrated by in vitro assays. Upon binding to the HER2, rHSA-ZHER2 only showed mild effect on HER2 positive SK-BR-3 cells, while rHSA-(ZHER2)2 significantly inhibited the proliferation of SK-BR-3 cells. More interestingly, a strong activation of both MAPK and AKT signaling was observed. Our results prove the feasibility of the fusion proteins to be used as drug carriers. Currently two formulations are under investigation, one is to complex the fusion protein with FA modified chemo drugs, and the other is to make them into nanoparticles. These two proteins not only can be explored as a potential drug delivery vehicle, but also may provide us with a good molecular tool to study the HER2 signaling pathway.

Citation Format: Daoyuan Dong, Zhijun Li, Zhiyu Li. Human serum albumin and affibody fusion proteins for targeted drug delivery to HER2 positive cells. [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2015 Nov 5-9; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2015;14(12 Suppl 2):Abstract nr B36.
Abstract B64: Schlafen 11 (SLFN11) irreversibly blocks cell cycle recovery independently of ATR following replicative damage by poly(ADPribose) polymerase inhibitors
Authors: Murai Junko , Pommier Yves
Published: May 2016

PARP inhibitors kill cancer cells by trapping PARP-DNA complexes, with talazoparib the inhibitor with the highest PARP trapping activity. Genomic analyses across the 60 cancer cell lines of the National Cancer Institute (NCI-60) revealed a highly significant correlation between SLFN11 expression and sensitivity to talazoparib. The causal role of SLFN11 was established after generating four SLFN11-knockout cell lines (prostate DU145, Ewing EW8, leukemia CCRF-CEM and MOLT4) by CRISPR/Cas9 system, and finding that SLFN11 inactivation conferred high resistance to both talazoparib and olaparib. Resistance was caused neither by impaired drug penetration nor by hyperactivation of homologous recombination, as demonstrated in BRCA2-inactivated cells. SLFN11-proficient cells were irreversibly arrested in S phase by talazoparib treatment. Although ATR is the key regulator of the intra-S phase checkpoint, the addition of the ATR inhibitor (VE-821) did not release the replication arrest in SLFN11-proficient cells. By contrast, SLFN11-deficient cells treated with talazoparib reached G2, and their cell cycle and replication arrest was reversed by ATR inhibition. The combination of VE-821 with talazoparib or olaparib was highly synergistic with the greater synergy in the SLFN11-deficient cells. Our study reveals that SLFN11 acts as a novel DNA damage response gene for replicative DNA damage, and that SLFN11 inactivation, which is common in cancer cells, is a novel mechanism of resistance for PARP inhibitors, which can be mitigated by combining PARP and ATR inhibitors.

Citation Format: Junko Murai, Yves Pommier. Schlafen 11 (SLFN11) irreversibly blocks cell cycle recovery independently of ATR following replicative damage by poly(ADPribose) polymerase inhibitors. [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2015 Nov 5-9; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2015;14(12 Suppl 2):Abstract nr B64.
Abstract B72: Cell-based DNA demethylation detection system for screening of epigenetic drugs in 2D, 3D and xenograft models
Authors: Agrawal Khushboo , Das Vishwanath , Otmar Miroslav , Krecmerova Marcela , Dzubak Petr , Hajduch Marian
Published: May 2016

Aberrant DNA methylation that turns ‘off’ the gene expression has remained a significant interest in cancer research. Despite major advances, the success of epigenetic therapy is prevented due to narrow therapeutic window. A wide variety of naturally occurring epigenetic agents and artificially synthesized molecules which can alter methylation patterns exists, but their usefulness in epigenetic therapy remains unknown. This underlines the need for large scale screening of epigenetic drugs with potent hypomethylation activity. In this ‘quest’ for more potent drugs targeting methylation, we present a cell-based DNA demethylation detection system, suitable for high content screening of epigenetic drug libraries in two-dimensional and three-dimensional culture systems. The detection system is fluorescence based and exploits the dominant ability of DNA methylation to inhibit gene transcription, and utilizes FLJ32130 gene which was discovered to be silenced on account of promoter hypermethylation in human colorectal cancer. In addition to demethylation screening, the detection system further incorporates the ability to monitor induced cytostatic/cytotoxic drug effects, in live cells, in a time dependent manner. Furthermore, our detection system also supports the in vivo determination of demethylation drug efficacy of potential hits selected on in vitro screening. Here, we describe the development of our cell-based DNA demethylation detection system, the anticipated results on screening of well-established DNA methylation inhibitors in two-dimensional versus three-dimensional cell cultures, and as well the efficacy of the system for monitoring demethylation in xenograft models.

Acknowledgements

The authors are grateful to Dr. Toshikazu Ushijima, Carcinogenesis Division, National Cancer Center Research Institute for his kind gift of FLJ32130 targeting vector. This study was supported by the grants awarded by the Ministry of Industry and Trade of the Czech Republic (MPO TIP FR-TI4/625), Ministry of Education, Youth and Sports (CZ09 / 7F14009 and LO1304; CZ.1.07/2.3.00/30.0041 to VD), Grant Agency of the Czech Republic (14-02652S and 14-03636S), and the Internal Grant Agency of Palacky University (IGA UP LF-2013/016).

Citation Format: Khushboo Agrawal, Vishwanath Das, Miroslav Otmar, Marcela Krecmerova, Petr Dzubak, Marian Hajduch. Cell-based DNA demethylation detection system for screening of epigenetic drugs in 2D, 3D and xenograft models. [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2015 Nov 5-9; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2015;14(12 Suppl 2):Abstract nr B72.
Abstract B84: Novel and selective inhibitors of histone deacetylases (HDAC) 1 and 2 significantly enhance the activity of the DNA methyltransferase inhibitor azacitidine in acute myeloid leukemia (AML)
Authors: Min Chengyin , Quayle Steven N. , Huang Pengyu , Shearstone Jeffrey R. , Jones Simon S. , Yang Min
Published: May 2016

AML is a heterogeneous group of hematopoietic stem cell disorders characterized by defects in myeloid differentiation and increased proliferation of neoplastic hematopoietic precursor cells. There is a great need for novel approaches to treat AML due to limited improvements in the treatment of patients over the past several decades. Aberrant epigenetic regulation plays an important role in the pathogenesis of AML, and the DNA methyltransferase inhibitor azacitidine is approved for the treatment of myelodysplastic (MDS), which frequently progresses to AML, and is commonly used in AML patients. Numerous clinical studies are ongoing to investigate the benefit of combining azacitidine with other investigational agents in AML.

HDAC inhibitors are promising agents for the treatment of AML. Selective HDAC inhibitors are predicted to reduce combination drug toxicity and other side effects observed with pan-HDAC inhibitors, while still enabling therapeutic benefits. In this work, we demonstrate that potent and selective inhibitors of HDAC1/2 are highly efficacious both as single agents and in combination with azacitidine in preclinical models of AML, including established AML cell lines and bone marrow samples freshly derived from AML patients. Combination treatment in vivo demonstrated good tolerability, and ongoing efficacy studies in AML xenograft models will be discussed. At the molecular level, gene expression profiling and DNA methylation mapping were performed on MV4-11 cells treated with an HDAC1/2 inhibitor, azacitidine, or the combination. Molecular signatures were analyzed by GSEA. Significantly more genes and signatures were upregulated than downregulated by the single agent and combination treatments, consistent with the transcription activation mechanism of these compounds. Signatures and genes involved in myeloid cell differentiation, apoptosis and cell cycle arrest were identified as potential mediators of the combinatorial effects of HDAC1/2 inhibition with azacitidine. Together, these findings support the clinical evaluation of selective HDAC1/2 inhibitors in combination with azacitidine in AML and potentially MDS patients.

Citation Format: Chengyin Min, Steven N. Quayle, Pengyu Huang, Jeffrey R. Shearstone, Simon S. Jones, Min Yang. Novel and selective inhibitors of histone deacetylases (HDAC) 1 and 2 significantly enhance the activity of the DNA methyltransferase inhibitor azacitidine in acute myeloid leukemia (AML). [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2015 Nov 5-9; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2015;14(12 Suppl 2):Abstract nr B84.
Abstract B90: HSP90 inhibitor NVP-AUY922 sensitizes intrinsic MEK inhibitor Trametinib-resistant NSCLC cells harboring KRAS mutation
Authors: Lee Dae Ho , Park Kang-Seo , Oh Bora , Lee Mi-Hee , Yang Hannah
Published: May 2016

Introduction: Mutant KRAS is feature of more than 25% of non-small cell lung cancers (NSCLC) and exemplifies one of the most popular oncogenic drivers. Ras-driven tumors are often hard to cure with conventional therapies and are necessary novel treatment strategies.

Experiment: Using the HSP90 inhibitor NVP-AUY922, we investigated the potential of HSP90 inhibition to overcome the resistance to Trametinib, MEK inhibitor, in lung cancers harboring K-RAS mutant cell lines (H23, H358, H647, H1944, and A549). The synergistic effect of AUY-922 and trametinib was determined by analysis of the combination index (CI). After performing western blotting, we underwent following xenograft study with 6-week-old Balb/c-nu/nu female using H649 and H1944 cell lines. The mice were randomized into groups of 5, and NVP-AUY922 (5 mg/kg) was administered 3 days/week and GSK1120212 (300 μg/kg) was administered 5 days/week for up to 21 days in total.

Result: NSCLC cells harboring G13D KRAS mutation showed intrinsic MEK inhibitor resistance via PI3K activation by highly expressed AKT as a bypass pathway. And then, HSP90 inhibitor AUY-922 suppresses both PI3K-AKT-mTOR and RAF-MEK-ERK reactivated by MEK inhibitor GSK1120212 and finally AUY-922 renders intrinsic MEK inhibitor resistant NSCLC cells sensitive to GSK1120212 with significant synergy. Furthermore, we showed this synergistic effect is occurred on sub-effective dose of either drug.

Conclusion: The dual inhibition of HSP90 and MEK signaling pathways on sub-effective dose may constitute a potent therapeutic strategy to treat intrinsic MEK inhibitor resistant G13DKRAS mutant NSCLCs for resolving toxicity problem of dual inhibition of AKT and MEK in clinical trials.

Citation Format: Dae Ho Lee, Kang-Seo Park, Bora Oh, Mi-Hee Lee, Hannah Yang. HSP90 inhibitor NVP-AUY922 sensitizes intrinsic MEK inhibitor Trametinib-resistant NSCLC cells harboring KRAS mutation. [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2015 Nov 5-9; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2015;14(12 Suppl 2):Abstract nr B90.
Abstract C110: Evaluation of immune cell populations in checkpoint inhibitor responsive syngeneic models
Authors: Barnes Sheri , Lasaro Marcio , Hauser Jacob , Mullin Robert J. , Synnott Aidan
Published: May 2016

Given the increasing success of cancer immunotherapies, we recognized a growing need for well-characterized preclinical models to enable drug discovery efforts. Syngeneic mouse tumor models offer advantages over conventional xenograft models for immunotherapy studies due to the presence of a fully functional immune system. We recently completed a comparative evaluation of the responsiveness of a collection of syngeneic models to two antibody based immunocheckpoint inhibitor therapeutics (anti-CTLA4, anti-PD1). This body of work resulted in the identification of several models responsive to checkpoint inhibition; CT26, Colon26, MC38, EMT-6, and A20. Our results clearly show differential tumor growth responses across this set of models following treatment with CTLA-4 and PD-1 targeted therapy. To further characterize these responses, we have performed a series of studies to identify and track immune cell populations by flow cytometry in blood, spleen, and tumor samples from mice harboring each responsive model. In these studies, we have investigated whether the modulation of CD4+, CD8+, regulatory CD4+ T cells (Treg), and myeloid derived suppressor cells (MDSC) populations correlates with the impact of immunocheckpoint inhibitors on tumor growth over time in untreated and challenged animals. These correlations of flow analyses with our antitumor data can point to the utility of cell-based endpoints as additional tools for evaluating combination outcomes.

Citation Format: Sheri Barnes, Marcio Lasaro, Jacob Hauser, Robert J. Mullin, Aidan Synnott. Evaluation of immune cell populations in checkpoint inhibitor responsive syngeneic models. [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2015 Nov 5-9; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2015;14(12 Suppl 2):Abstract nr C110.
Abstract C115: Targeting glutamine metabolism in colorectal cancers with PIK3CA mutations
Authors: Wang Zhenghe John , Hao Yujun
Published: May 2016

Glutamine addiction is a major metabolic reprogramming event that occurs in cancer cells. Many tumors exhibit oncogene-dependent addiction to glutamine. PIK3CA, which encodes the p110α catalytic subunit of phosphatidylinositol 3-kinase α, is the most frequently mutated oncogene in human cancers. However, whether PIK3CA mutations reprogram cancer metabolism is an important unaddressed question. Here we show that oncogenic PIK3CA mutations render colorectal cancers (CRCs) more dependent on glutamine to growth. As a metabolite, glutamine is first converted glutamate by glutaminase (GLS) and then to α-ketoglutarate (α-KG) by either a transaminase or a glutamate dehydrogenase. Calithera Biosciences recently developed a potent GLS inhibitor called CB-839, which is currently in phase I clinical trials in cancer patients. Using isogenic colorectal cancer cell lines with either WT or mutant PIK3CA allele knockout, we demonstrated that CRCs with PIK3CA mutations are more sensitive to growth inhibition by CB-839. Remarkably, combination of CB-839 with 5-FU induces regression xenograft tumors from a CRC with a PIK3CA mutation, suggesting that this combinational therapy may be effective approach to treat CRC patients whose tumors harbor PIK3CA mutations.

Mechanistically, mutant p110α up-regulates gene expression of glutamate pyruvate transaminase 2 (GPT2) in CRC cells, thereby facilitate conversion of glutamate to α-KG. Using [13C5-]glutamine isotope-tracing technology, we showed that PIK3CA mutant CRCs produce more α-KG from glutamine to α-KG to replenish the tricarboxylic acid (TCA) cycle to generate ATP. Moreover, aminooxyacetate, which inhibits enzymatic activity of transaminases including GPT2, suppresses xenograft tumor growth of CRCs with PIK3CA mutations, but not CRCs with WT PIK3CA. Mutant p110α up-regulates the transcription of GPT2 through an AKT-independent PDK1-RSK2-ATF4 signaling axis. We showed that ATF4 is a transcription factor that activates GPT2 gene expression. We further demonstrated that mutant p110α activates RSK2 kinase through PDK1. Activated RSK2 then phosphorylates ATF4 at the serine residue 245, which in turn recruits deubiquitinase USP8 and protects ATF4 from ubiquitin-mediated degradation. Together, our data establish oncogenic PIK3CA mutations as a cause of glutamine addiction in CRCs and that targeting glutamine metabolism may be effective approach to treat CRCs with PIK3CA mutations.

Citation Format: Zhenghe John Wang, Yujun Hao. Targeting glutamine metabolism in colorectal cancers with PIK3CA mutations. [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2015 Nov 5-9; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2015;14(12 Suppl 2):Abstract nr C115.