bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 677: Possible role of nimbolide in prevention and therapy of prostate cancer
Authors: Zhang Jingwen , Shanmugam Muthu K. , Deivasigamani Amudha , Kumar Alan P. , Hui Kam M. , Sethi Gautam
Published: August 2015

Prostate cancer is one of the most commonly diagnosed cancers worldwide. Current therapies for metastatic prostate cancer are only marginally effective, and hence novel treatment modalities are urgently needed. Considerable evidence suggests that chronic inflammation plays a pivotal role in the development and progression of prostate cancer. Thus agents that can suppress these inflammatory mediators could form the basis of novel therapy for prostate cancer patients. In our study, we focused on analyzing the potential anticancer effects of nimbolide, a terpenoid lactone derived from Azadirachta indica (Neem tree) against prostate cancer. Data from the in vitro studies indicated that nimbolide could inhibit cell proliferation, induce apoptosis and suppress cellular invasion and migration. Interestingly, nimbolide also abrogated the activation of pro-inflammatory STAT3 transcription factor, and this effect was found to be mediated via an increased production of reactive oxygen species (ROS), whereas depletion of ROS attenuated p-STAT3 inhibitory effects of the drug. The in vivo efficacy of nimbolide was also analyzed in transgenic adenocarcinoma of mouse prostate (TRAMP) model, in which this triterpenoid significantly suppressed the tumor progression and growth without exhibiting any significant adverse effects. Overall our findings indicate that nimbolide exhibits significant anticancer effects in prostate cancer, and these effects may be mediated at least in part through the modulation of STAT3 signaling pathway.

Citation Format: Jingwen Zhang, Muthu K. Shanmugam, Amudha Deivasigamani, Alan P. Kumar, Kam M. Hui, Gautam Sethi. Possible role of nimbolide in prevention and therapy of prostate cancer. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 677. doi:10.1158/1538-7445.AM2015-677
Abstract 679: CF102 induces apoptosis of hepatocellular carcinoma
Authors: Cohen Shira , Barer Faina , Stemmer Salomon M. , Fishman Pnina
Published: August 2015

Background: The A3 Gi protein adenosine receptor (A3AR) is highly expressed in tumor compared to normal cells. A3AR agonists were shown to inhibit the growth of various tumor cell types via down-regulation of the Wnt and NF-κB signaling pathways. CF102 is a highly selective agonist of the A3AR, shown to have an anti-tumor effect in liver cancer in in vivo models. CF102 induced apoptosis via up-regulation of caspase-3 and the mitochondrial proteins Bax and Bad, demonstrating cell death in the tumor cells.

In this study we explored the molecular mechanism involved with the effect of CF102 on human hepatocellular carcinoma cells.

Methods: CF102 was added to a medium culture of Hep3b for 48 hours. CF102 effect was observed using Lactate dehydrogenase (LDH), MTT and H3 Thymidine proliferation assays. Cell mRNA and protein were extracted and were analyzed for different signaling pathways.

Results: CF102 inhibits the proliferation of Hep3b cells and showed an apoptotic activity in MTT assay. A cytotoxicity effect of CF102 was observed when high levels of LDH were measured in the culture medium of CF102 treated cells compared to the control. Moreover, cyclin D1 expression levels were down-regulated and caspase-3 levels were up-regulated upon CF102 treatment. Both mRNA and protein expression levels of A3AR were down-regulated by CF102 with a subsequent decrease in all members of the JAK-STAT signaling pathway including pSTAT-1, p-JAK-2 and p-AKT along with up-regulation of SOCS-1 and SOCS-3.

Conclusions: CF102 showed an anti-cancer activity via down-regulation of the A3AR and the JAK-STAT signaling pathway. CF102 induces cytotoxicity effect resulting in apoptosis of Hep3b cells, indicating that this drug may be considered as an anti-cancer drug for the treatment of human hepatocellular carcinoma.

Note: This abstract was not presented at the meeting.

Citation Format: Shira Cohen, Faina Barer, Salomon M. Stemmer, Pnina Fishman. CF102 induces apoptosis of hepatocellular carcinoma. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 679. doi:10.1158/1538-7445.AM2015-679
Abstract 687: Compound screen identifies PIM kinases as therapeutic targets for melanoma
Authors: Vultur Adina M. , Shannan Batool , Chen Quan , Watters Andrea , Mollin Stefan , Meggers Eric , Krepler Clemens , Perego Michela , Li Ling , Gimotty Phyllis A. , Xu Xiaowei , Herlyn Meenhard
Published: August 2015

Current treatments against malignant melanoma, including the use of small molecule inhibitors, are displaying encouraging results in the clinic. However, tumor resistance develops even for patients who initially respond favorably. To identify novel drug targets and compounds with anti-melanoma activity, we screened a panel of genetically distinct human-derived metastatic melanoma cell lines against structurally diverse organometallic kinase inhibitors. We observed a compound that preferentially inhibits melanoma cell proliferation compared to normal primary fibroblasts and melanocytes in adherent and three-dimensional (3D) cultures. We further showed that this compound (SM200) is pro-apoptotic and anti-invasive in 3D melanoma cell cultures. A kinome screen to identify the targets responsible for the anti-melanoma effects of SM200 revealed PIM kinases to be highly inhibited. PIM1 knockdown studies to validate this target in human metastatic melanoma indicate that PIM1 contributes to melanoma growth in 3D culture and in a xenograft model. We then evaluated the clinically available PIM kinase inhibitor SGI-1776 in our melanoma models and observed reduced cell proliferation in 2D, 3D, and in vivo melanoma models (as seen for SM200) suggesting an important role for PIM kinases in melanoma pathobiology. Taken together, our findings suggest that PIM kinases are valid targets for melanoma and PIM kinase inhibitors could enhance the effects of current therapies in the clinic.

Citation Format: Adina M. Vultur, Batool Shannan, Quan Chen, Andrea Watters, Stefan Mollin, Eric Meggers, Clemens Krepler, Michela Perego, Ling Li, Phyllis A. Gimotty, Xiaowei Xu, Meenhard Herlyn. Compound screen identifies PIM kinases as therapeutic targets for melanoma. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 687. doi:10.1158/1538-7445.AM2015-687
Abstract 688: Activating JAK1-S703I mutation may predict the sensitivity of JAK-STAT inhibition in hepatocellular carcinoma patient-derived xenograft tumor models
Authors: Yang Shuqun , Luo Chonglin , Gu Qingyang , Xu Qiang , Sun Hongye , Qian Ziliang , Tan Yexiong , Wu Hao , Qin Yuxin , Shen Yuhong , Xu Xiaowei , Chen Shu-Hui , Chan Chi-Chung , Wang Hongyang , Mao Mao , Fang Douglas D.
Published: August 2015

Hepatocellular carcinoma (HCC) is the fifth most common type of cancer worldwide. However, current therapeutic approaches for this epidemic disease are limited and its 5-year survival rate has not been improved in the past decades. Patient-derived xenograft (PDX) tumor models have become an excellent in vivo system for understanding of disease biology and drug discovery. In order to identify new therapeutic targets for HCC, whole-exome sequencing (WES) was performed on more than 60 HCC PDX models. Among them, four models exhibited point mutations in JAK1 (Janus Kinase) gene, which were confirmed by Sanger sequencing. To explore the transforming capability, these mutations were then introduced into 293T and Ba/F3 cells by transient transfection and retroviral infection, respectively. The results demonstrated that JAK1S703I mutation was able to activate JAK-STAT (Signal Transducer and Activator of Transcription) signaling pathway and drive cell proliferation in the absence of cytokine stimulation in vitro. Furthermore, a modest sensitivity to the treatment of a JAK1/2 inhibitor, Ruxolitinib, was observed in JAK1S703I mutant PDX model, but not in other mutant or wild type models. Pharmacodynamic analysis showed that phosphorylation of STAT3 in the Ruxolitinib-treated tumor tissues was significantly inhibited. Collectively, our results suggested that JAK1S703I is an activating mutation for JAK-STAT signaling pathway in vitro and in vivo. JAK-STAT pathway might represent a new therapeutic opportunity for HCC. Monotherapy using a more potent inhibitor and combinatory therapy may be further explored in JAK1S703I mutant PDX model.

Citation Format: Shuqun Yang, Chonglin Luo, Qingyang Gu, Qiang Xu, Hongye Sun, Ziliang Qian, Yexiong Tan, Hao Wu, Yuxin Qin, Yuhong Shen, Xiaowei Xu, Shu-Hui Chen, Chi-Chung Chan, Hongyang Wang, Mao Mao, Douglas D. Fang. Activating JAK1-S703I mutation may predict the sensitivity of JAK-STAT inhibition in hepatocellular carcinoma patient-derived xenograft tumor models. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 688. doi:10.1158/1538-7445.AM2015-688
Abstract 69: Identification of rare and novel alleles in FFPE tumor samples using laser capture microdissection (LCM) and Ampliseq™ sequencing technologies
Authors: Jackson Stephen , Choppa Paul , Kulvar Kulvinda , Schmidt Kristin
Published: August 2015

Tumors are becoming recognized as genetically heterogeneous masses of cells with different clonal histories. Identifying the mutations present in these heterogeneous masses can lead to important insights into the future behavior of the tumor and possible intervention mechanisms. However, the rarity of pathogenic mutations in small subsets of cells can make identification of such alleles difficult. In this study, we demonstrate a complete workflow that facilitates the identification of rare and novel alleles from FFPE tumor sections. We collected small regions with different cellular morphologies from lung tumor samples using laser capture microdissection, extracted both DNA and RNA from these regions, and characterized mutations present and transcript abundances by using Ampliseq™ targeted sequencing. We show that LCM facilitates the detection of alleles that are not detectable in macrodissected tissue scrapes. We also show that different regions of a tumor have very different patterns of alleles detectable and have a great deal of genetic diversity. Finally, we show that RNA expression patterns are also clearly different in the different regions. Interestingly, dissected regions with similar gross tissue morphologies display differences in alleles present and RNA expression patterns. These results suggest the best way to analyze mutations present in a tumor is to microdissect different subregions of the tumor, and using Ampliseq™ panels to identify the alleles present in those subregions.

Citation Format: Stephen Jackson, Paul Choppa, Kulvinda Kulvar, Kristin Schmidt. Identification of rare and novel alleles in FFPE tumor samples using laser capture microdissection (LCM) and Ampliseq™ sequencing technologies. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 69. doi:10.1158/1538-7445.AM2015-69
Abstract 695: Synergistic antitumor effects of polo like kinase inhibitor volasertib in combination with ionizing radiation in glioblastoma
Authors: Dong Jianwen , Nguyen Nghi , Ezhilarasan Ravesanker , Wu Shaofang , Piao Yuji , Park Soon Young , Tiao Ningyi , Stephan Clifford , Sulman Erik P. , de Groot John F.
Published: August 2015

Despite the availability of hundreds of drugs, there is little data on the efficacy of these agents in the extremely heterogeneous populations of tumor cells observed in glioblastoma. In this study, a high-throughput compound-screening (HTS) assay was used to identify drug sensitivities of a panel of 15 glioblastoma stem cell (GSC) lines, which are representative of the classic the cancer genome atlas (TCGA) molecular subtypes, to 21 compounds in Published Protein Kinase Inhibitor Set (PKSI). This HTS screen identified sensitivity of GSCs to inhibition of polo like kinase-1 (PLK-1), a key regulator of mitosis. Given that PLK-1 is often overexpressed in a broad spectrum of cancers, and with highest expression levels being correlated with poor prognosis in several cancer types as well, we further verified the HTS result with the second-generation PLK1 inhibitor volasertib as a single agent or in combination with ionizing radiation in GSCs. Efficacy of volasertib was analyzed by Cell-Titer Glo 5 days after treatment. Morphological and molecular changes were approached by immunoblotting and flow cytometry after volasertib treatment, and in combination with ionizing radiation at 2 Gy. In vitro studies showed that volasertib inhibited cell viability with an IC50 ranging from 44.3nM to 4.36μM. Volasertib induced G2/M arrest accompanied by high levels of PLK-1, Aurora B and phosphor-histone 3 and prominent cleaved poly ADP ribose polymerase (PARP) in a dose- and time-dependent manner. Colony formation assay demonstrated that volasertib and ionizing radiation had synergistic effects on colony formation inhibition, suggesting that GSCs arrested in M phase by volasertib are more sensitive to ironizing radiation. The promising in vitro results of volasertib in GSCs led to further investigate the drug efficacy in intracranial xenograft models by volasertib alone, and in combination with radiation. Taken together, our results reinforce the potential therapeutic candidate of volasertib as a single agent and in combination with ionizing radiation in glioblastoma.

Citation Format: Jianwen Dong, Nghi Nguyen, Ravesanker Ezhilarasan, Shaofang Wu, Yuji Piao, Soon Young Park, Ningyi Tiao, Clifford Stephan, Erik P. Sulman, John F. de Groot. Synergistic antitumor effects of polo like kinase inhibitor volasertib in combination with ionizing radiation in glioblastoma. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 695. doi:10.1158/1538-7445.AM2015-695
Abstract 697: c-MET as a potential target in ovarian clear-cell carcinoma
Authors: Lee Jeong-Won , Kim Byoung-Gie , Bae Duk-Soo , Lee Yoo Young
Published: August 2015

Purpose: Recent reports revealed that c-MET activation is associated with epithelial ovarian carcinoma (EOC) and is association with poor prognosis. In this study, we investigated the effects of therapeutic targeting for c-MET in ovarian clear cell carcinoma (OCCC).

Experimental Design: Expression levels of c-MET in the epithelial ovarian carcinomas and normal ovarian tissues were evaluated using real-time PCR. To test c-MET inhibitors in CCC cell lines, we performed in vitro experiments including MTT and apoptosis assay. We performed Western blots to evaluate the c-MET expression and down-stream pathway. In addition, we performed in vivo therapy experiments in orthotopic ovarian cancer mice model (RMG1) and patient-derived tumor xenograft (PDX) models of CCC to confirm these effects.

Results: The c-MET expression was significantly increased in CCCs compared with serous carcinomas and normal ovarian tissues (P < 0.05). CCC cells treated with c-MET inhibitors (SU11274 or crizotinib) demonstrated significantly decreased cell viability and increased apoptosis. Western blot assay showed that the protein expressions for c-MET signaling pathway were decreased by c-MET inhibitor. Moreover, tumor weight was significant decreased in both in vivo RMG1 and PDX models receiving SU11274 treatment compared with control (P < 0.05).

Conclusion: These results show that c-MET inhibitor has the significant anti-tumor effects in ovarian CCC, and suggested that c-MET could have the potential agent in the therapy for this cancer.

Citation Format: Jeong-Won Lee, Byoung-Gie Kim, Duk-Soo Bae, Yoo Young Lee. c-MET as a potential target in ovarian clear-cell carcinoma. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 697. doi:10.1158/1538-7445.AM2015-697
Abstract 698: CPI-based Crizotinib systemic study on discovery of phenotypic off-target
Authors: Sun Jiazhi , Yang Lun , Li Minghua , Zhang Shimin , Heller Ian E. , Samuelson Johnathan , Cindric Steven , Mishra Kunal , Mohamed Eslam , Cheng Feng , Sneed Kevin B. , He Lin , Zhou Shu-Feng
Published: August 2015

Crizotinib, a first ever ALK/MET multi-targeted receptor tyrosine kinase inhibitor for the treatment of ALK-rearranged NSCLC patients. We hypothesize that crizotinib acts on other molecular targets in addition to tyrosine kinases ALK/MET and may play critical role given that there is a complex network of kinases that work together to regulate a number of important cellular processes and different disease signature. Approaching with a comprehensive docking method with our established chemical-protein interactome (CPI) and crizotinib, we have discovered 301 PDB-deposited proteins corresponding to 353 ligand binding pockets among a total of 1,780 PDB-deposited human protein entries. Systemic pharmacology approach also applied including validation the molecular target(s) of TKIs in vitro. Especially multiple human cellular models (including NSCLC and melanoma) and signaling study, luc-reporter, autophagy/apoptosis, proteomics study and NGS analysis applied. Interestingly, crizotinib besides had a high CPI binding score (ZZ_score) of -2.2778 against tyrosine kinase Lck, also had -1.4672, -1.1242, -0.7033 and -1.5384 against MAPK3, S6K, GSK-3β and HDAC 7A, respectively, which indicated crizotinib had potential broad range interaction with both TK and ser/thr kinases. Our data have showed that the crizotinib induced autophagy by activated LC3 in vitro and inhibited oncogenic Akt/mTOR signaling in human multiple melanoma cells resulting in G2/M cell cycle arrest. Furthermore, crizotinib increased p21Waf1/cip1 and Foxo3A expression and tumor suppressor miR-146a in human melanoma cells. Taken together, crizotinib induces miR-146a, autophagy and p21waf1/cip1 may potentially treat other type of human cancer by concurrently targeting tyrosine kinase, Akt/mTOR/Foxo axis and nuclear receptor, shedding a light for future both anti-cancer and anti-metabolic disorder drug discovery and development. This is the first CPI-based systemic study on leading discovery of crizotinib off-target phenotypic importance. These findings suggest that non-tyrosine kinase target of crizotinib may play critical role(s) for drug repositioning may represent a new and effective approach to expanding the application of existing drugs.

Citation Format: Jiazhi Sun, Lun Yang, Minghua Li, Shimin Zhang, Ian E. Heller, Johnathan Samuelson, Steven Cindric, Kunal Mishra, Eslam Mohamed, Feng Cheng, Kevin B. Sneed, Lin He, Shu-Feng Zhou. CPI-based Crizotinib systemic study on discovery of phenotypic off-target. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 698. doi:10.1158/1538-7445.AM2015-698
Abstract 699: PIK3CA/BRAF mutations negatively affect outcome of patients with KRAS wild-type metastatic colorectal cancer treated with front line anti-EGFR monoclonal antibodies: Meta-analysis results
Authors: Mohamed Amr , Schrapp Kelly , Attumi Taraq , Saba Nabil F. , El-Rayes Bassel F.
Published: August 2015

BACKGROUND:The aim of this meta-analysis is to evaluate the effect of PIK3CA and BRAF mutations in patients with KRAS wild-type metastatic colorectal cancer receiving an EGFR monoclonal antibody (anti-EGFR) inhibitor in the first line therapy.

METHODS:literature search was performed to identify studies exploring the association between PIK3CA/BRAF mutations and clinical outcomes of KRAS wild mCRC patients treated with anti-EGFR as a first line therapy. The primary clinical outcome is response rate (RR). The secondary outcomes include progression-free survival (PFS), and overall survival (OS). The pooled relative risk (RR) or hazard ratio (HR) was estimated by using fixed effect model or random effect model according to heterogeneity between studies.

RESULTS:Ten studies with 1,470 mCRC patients (357 for PI3K studies and 1,113 from BRAF studies) met selection criteria. We observed a trend towards lower RR in patients with PIK3CA mutations (2 studies, 137 patients; RR = 14.3% versus 52.4%; 95% confidence interval (CI) -0.12-0.02; P = 0.13). Patients with mutant-type PI3K have significant shorter PFS (3.8 vs 4.15 mos, HR = 1.36; 95% CI 1.04-1.77; P = 0.02), and OS (14.17 Vs 16.3 mos., HR = 1.50; 95% CI 1.14-1.97; P = 0.004) compared to those with wild PI3K. For BRAF, patients with mutant type have significant lower RR [4 studies, 416 patients; RR = 33% versus 39%; 95% confidence interval (CI) -0.16- 0.01; P = 0.03], shorter PFS (6 studies, 893 pts., 3.9 Vs 5.7 mos., HR = 1.72; 95% CI 1.47- 2.01; P = 0.00001), and shorter OS (6 studies, 1065 pts., 9.1 Vs 18.9 mos., HR = 1.22; 95% CI 1.04-1.44; P = 0.01) compared to those with wild type.

CONCLUSION:These analyses suggest that patients with colorectal cancer and either PIK3CA or BRAF mutation have worse outcome when treated with anti-EGFR in the first line. Given their worse outcome, trials evaluating novel agents are needed in this group of patients.

Meta-analyses on BRAFF and PIK3CA mutations in KRAS wild mCRCBRAF WildBRAF MutantHR (95% CI)p-valuePI3K WildPI3K MutantHR (95% CI)p-valueRR (%)39%33%-0.16 - 0.010.0352.4%14.3%-0.12 - 0.020.13PFS (mos)5.73.91.47 - 2.010.000014.153.81.04 - 1.770.02OS (mos)18.99.11.04 - 1.440.0116.314.71.14 - 1.970.004

Citation Format: Amr Mohamed, Kelly Schrapp, Taraq Attumi, Nabil F. Saba, Bassel F. El-Rayes. PIK3CA/BRAF mutations negatively affect outcome of patients with KRAS wild-type metastatic colorectal cancer treated with front line anti-EGFR monoclonal antibodies: Meta-analysis results. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 699. doi:10.1158/1538-7445.AM2015-699
Abstract 70: Targeted sequencing analyses of uterine and ovarian carcinosarcoma
Authors: Mori Seiichi , Gotoh Osamu , Amino Sayuri , Nakai Megumi , Takazawa Yutaka , Sugiyama Yuko , Noda Tetsuo
Published: August 2015

Carcinosarcoma (CS) of the uterus or the ovary, known as malignant mixed Muellerian tumor, is a biphasic neoplasm composed of malignant epithelial (carcinoma) and mesenchymal (sarcoma) elements. CS is a rare disease (two per 100,000 women) and exhibits unfavorable outcomes (5-year survival rates: uterus and ovary; 31 and 7.5%, respectively). CS has been considered as an aggressive form of high-grade endometrioid adenocarcinoma (EC), therefore, the treatment regimen for CS follows to that for EC. Nevertheless, the fact that CS shows substantially poorer prognosis than EC implies a need to identify therapeutic targets for CS. Toward this aim, we have been performing targeted resequencing with a panel of 521 genes (including 30 introns), which were previously implicated as critical not only for uterine, ovarian or sarcomatus oncogenesis but also for molecular targets in therapeutics and the panel covers 2.161 Mbp with 38,105 probes on Agilent's SureSelect. Genomic DNA was extracted from matched tumor and normal pairs derived from more than 70 cases of uterine and ovarian CS patients and then used to construct pooled libraries, which were subsequently subjected to sequencing analysis with an Illumina HiSeq sequencer. Consensus clustering with genomic changes such as mutational spectrum and number of somatic SNVs revealed several subtypes that correlate with histopathology and patient outcomes. Moreover, potentially actionable mutations or copy number aberrations were identified in a number of genes including PIK3CA, MTOR, KRAS and CCNE1. This study would provide deep insights into the development of novel diagnostic and personalized treatment for CS patients.

Citation Format: Seiichi Mori, Osamu Gotoh, Sayuri Amino, Megumi Nakai, Yutaka Takazawa, Yuko Sugiyama, Tetsuo Noda. Targeted sequencing analyses of uterine and ovarian carcinosarcoma. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 70. doi:10.1158/1538-7445.AM2015-70
Abstract 702: BRAF V600E mutation in benign and malignant epithelial odontogenic tumors
Authors: Diniz Marina Goncalves , Gomes Carolina Cavalieri , Guimaraes Bruna Viana Antonini , Duarte Alessandra Pires , Gomez Ricardo Santiago
Published: August 2015

The classification of ameloblastoma in multicystic or unicystic variants is associated with its clinical behaviour. Recently, BRAF and SMO mutations have been reported in ameloblastomas. However, it is not clear if such mutations are shared by the multi- and unicystic variants of ameloblastoma or by odontogenic carcinomas. We assessed BRAFV600E and SMOF412E in multicystic, unicystic and desmoplastic ameloblastomas. In addition, we investigated whether the BRAFV600E mutation occurs in odontogenic carcinomas. A total of 28 formalin-fixed paraffin-embedded samples, comprising 17 ameloblastomas and 11 odontogenic carcinomas, were included. The BRAFV600E mutation was assessed by real-time PCR with a specific TaqMan probe and confirmed by Sanger sequencing. The SMOF412E mutation was assessed by Sanger sequencing. Fourteen out of 17 (82%) ameloblastomas showed the BRAFV600E mutation, specifically, 5/6 (83%) unicystic, 7/9 (78%) multicystic and 2/2 desmoplastic ameloblastomas. BRAFV600E mutation was detected in 4/11 (36%) malignant tumours, specifically, 3/8 (38%) ameloblastic carcinomas and 1/1 clear cell odontogenic carcinoma, while the two ghost cell odontogenic carcinomas did not harbour this mutation. The SMOF412E mutation was not detected in ameloblastoma. The BRAFV600E mutation was less frequent in odontogenic carcinomas than in ameloblastomas. In ameloblastomas, this mutation occurred at similar proportions in cases from the mandible and maxilla, as well as among unicystic, multicystic and desmoplastic variants, suggesting that there is no difference in their molecular pathogenesis. Our findings support the possibility for personalised, molecular targeted therapy for ameloblastomas and ameloblastic carcinomas harbouring the BRAFV600E mutation. Supported by: FAPEMIG (Fundação de Amparo à Pesquisa de Minas Gerais), CNPq (National Council for Scientific and Technological Development) and CAPES (Coordination for Improvement of Higher Education Personnel), Brazil

Citation Format: Marina Goncalves Diniz, Carolina Cavalieri Gomes, Bruna Viana Antonini Guimaraes, Alessandra Pires Duarte, Ricardo Santiago Gomez. BRAF V600E mutation in benign and malignant epithelial odontogenic tumors. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 702. doi:10.1158/1538-7445.AM2015-702
Abstract 704: Scaffold protein KSR1 is negatively regulated by merlin and promotes tumor development in merlin deficient tumors
Authors: Hanemann Clemens O. , Zhou Lu , Ammoun Sylwia , Lasonder Edwin , Sharma Vikram , Muller Juergen , Ercolano Emanuela
Published: August 2015

Accumulated evidence suggests that the Ras/Raf/MEK/ERK pathway plays a crucial role in the development of Merlin-deficient tumours. However, the mechanism in which Merlin precisely suppresses this mitogenic signalling pathway has remained poorly understood. In addition, drug specificity, side effects and drug resistance are serious problems when treating patients with MEK inhibitors and therefore a more specific therapeutic target needs to be identified. Using our primary human in vitro model and three complementary approaches including lentiviral shRNA, active mutants and use of a clinically approved inhibitor, we report that Ras/Raf/MEK/ERK scaffold protein Kinase Suppressor of Ras 1 (KSR1) plays a vital role in promoting schwannoma proliferation, multipolar morphology and other pathological phenotypes. Proteomic analysis suggests that merlin shares multiple binding partners with KSR1 incl DCAF1 from the E3 Ubiquitin Ligase CRL4DCAF1, which is directly associated with and functionally inhibited by the tumour suppressor Merlin in the nucleus. Functional study suggests that KSR1 and DCAF1 might co-operate and/or complement to regulate schwannoma's proliferation. These findings demonstrate that Merlin disrupts Ras/Raf/MEK/ERK activity at the scaffold protein level and also suggest that KSR1 is a specific therapeutic target for Merlin-deficient tumours.

Citation Format: Clemens O. Hanemann, Lu Zhou, Sylwia Ammoun, Edwin Lasonder, Vikram Sharma, Juergen Muller, Emanuela Ercolano. Scaffold protein KSR1 is negatively regulated by merlin and promotes tumor development in merlin deficient tumors. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 704. doi:10.1158/1538-7445.AM2015-704
Abstract 705: Combinatorial therapeutic targeting of BMP2 and MEK in NF1-null malignant peripheral nerve sheath tumors
Authors: Ahsan Sidra , Sun Daochun , Tainsky Michael A.
Published: August 2015

Our lab has identified and validated bone morphogenetic protein 2 (BMP2) signaling pathways as a therapeutic target in malignant peripheral nerve sheath tumors (MPNSTs), independent of the NRAS and MEK1/2 regulation. BMP2/SMAD signaling is associated with enhanced motility and invasion, a hallmark feature of the biological aggressiveness of MPNSTs. We have shown that inhibition of BMP2 leads to decreased motility and invasiveness in MPNST cell lines. Consistent with our data, gene expression studies from NF1 patient tissue samples demonstrated that increased expression of BMP2 is associated with malignancy in NF1 patients. Ras-GAP related domain (GRD) is the most widely studied functional target of Nf1 implicated in tumorigenesis, however, therapeutic interventions targeting Ras activity have met with limited success. Our objective is to target the BMP2/SMAD pathway, involved in migration and invasion, in combination with MEK inhibitors to develop a novel therapeutic approach for treatment of MPNSTs.

The mechanism by which NF1-deficiency leads to activation of BMP2 signaling pathways, and consequently increased cellular motility and invasion is unknown. Using NF1(+/-) and NF1 knockdown MPNST cell lines, we are investigating the transcriptional regulation of BMP2 by NF1 to identify signaling mechanisms downstream of NF1 mediating increase in BMP2 levels. We will present data showing that BMP2 is regulated at the level of transcript stability upon NF1 deficiency. We will identify and design a rational approach of targeting of the molecular components involved in stability of the BMP2 3′ UTR in MPNSTs. Furthermore, we will present a combinatorial approach that targets the growth and proliferative potential of MPNSTs by RAS-MEK inhibitors, along with motility and invasive capability by inhibition of the BMP2-Smad axis for better targeting of these tumors. The overall goal of our research is to use the regulation of BMP2 transcriptional control to define mechanism-based novel druggable targets that can be exploited in combinatorial therapies with RAS-MEK inhibitors to treat NF1-related tumors.

Citation Format: Sidra Ahsan, Daochun Sun, Michael A. Tainsky. Combinatorial therapeutic targeting of BMP2 and MEK in NF1-null malignant peripheral nerve sheath tumors. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 705. doi:10.1158/1538-7445.AM2015-705
Abstract 708: Androgen receptor activation inhibits papillary thyroid carcinoma cell cycle progression
Authors: Jones Melanie Elizabeth , O'Connell Timmy , Gupta Anvita , Zhao Hong , Iacob Codrin , Moscatello Augustine , Shin Edward , Darzynkiewicz Zbigniew , Tiwari Raj K. , Geliebter Jan
Published: August 2015

The American Cancer Society predicts 62,980 new cases of thyroid cancer and 1,890 deaths due to the disease in 2014, making thyroid cancer the most prevalent endocrine malignancy. This is particularly concerning for women who have an approximately three-fold higher incidence of thyroid cancer than men. Significant research has been done to elucidate the role of estrogen as a driver of thyroid cancer. Taking the reverse approach, that men have a much lower incidence of thyroid cancer, led us to hypothesize that androgens and/or androgen receptors play a protective role and that PTC represents the escape from androgen-mediated cell regulation. Analysis of AR expression in 24 PTC patient tissue samples indicated a 2.7 fold reduction in AR expression compared to matched normal tissue (P<0.005). This suggests that the reduction of AR levels in the neoplastic process of thyroid tissue may be one mechanism of escape from androgen regulation. To more fully understand the function of androgen/AR in thyroid cells, the AR was stably transfected into 8505C anaplastic/PTC cells. Addition of DHT to the 8505C-transfected clone, 84E7, resulted in AR translocation into the nucleus and a 70% reduction in proliferation, as well as a shift in the cell cycle toward G1 arrest. Transcription profiling using RNA-Seq and gene ontology analysis revealed significant changes in genes associated with proliferation, cell cycle, and cell cycle regulation confirming the proliferation assay and cell cycle analysis above. Further, when we examined G1 associated cell cycle proteins, we found significant decreases in cell cycle progression proteins cdc25a, CDK6, CDK4, and CDK2 as well as significant increases in the inhibitors p27 and p21. Thus, protein, RNA and cell cycle analysis data are concordant and demonstrate an accumulation of cells in G1 when treated with DHT. Together these data demonstrate that expression of androgen/AR in normal thyroid cells may play a protective, anti-proliferative role that is lost in PTC resulting in dysregulation of genes normally under AR control. The differential expression of androgen/AR in males verses females may provide new insight into the different clinical presentations and outcomes between women and men.

Citation Format: Melanie Elizabeth Jones, Timmy O'Connell, Anvita Gupta, Hong Zhao, Codrin Iacob, Augustine Moscatello, Edward Shin, Zbigniew Darzynkiewicz, Raj K. Tiwari, Jan Geliebter. Androgen receptor activation inhibits papillary thyroid carcinoma cell cycle progression. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 708. doi:10.1158/1538-7445.AM2015-708
Abstract 71: Epigenetic control of signature gene expression in the inducible differentiation of acute promyelocytic leukemia cells
Authors: Song Chunhua , Pan Xiaokang , Li Zhanjun , Ding Yali , Mudhusami Sunil , Gowda Chandrika , Ge Zheng , Steffens Sadie , Tan Bi-hua , Dovat Sinisa
Published: August 2015

[Background] Epigenetic changes in DNA and chromatin are regarded as emerging major players for hematopoietic stem cells development and lineage differentiation. Epigenetic deregulation of gene expression leads to leukemia and reversibility of epigenetic modifications makes DNA and chromatin changes attractive targets for therapeutic intervention. The promyelocytic leukemia HL-60 cell can differentiate into microphage, granulocyte and monocyte with stimulation of phorbol myristic acid (PMA), all-trans retinoic acid (ATRA) or dimethylsulfoxide (DMSO), and Vitamin D3, respectively, by affecting the gene expression on cell cycle and cell differentiation. However, how epigenetic regulation on gene expression in the inducible differentiation is still not well determined. [Methods and Results] We did the microarray analysis to identify the genome-wide gene expression profile of HL-60 cells in various time points(0h, 6h, 1d,2d,4d and 6d) under the treatment of PMA, ATRA/DMSO and D3 and found that around 3000 genes are significantly altered commonly under those treatments in the process. The percentage of down-regulated genes in the all commonly altered genes is significantly higher than that of up-regulated genes, and the significantly altered genes showed significantly physical clustering on chromosome loci, indicating the epigenetic regulation involved in the regulation of the gene expression. We did observed the expression changes of epigenetic enzymes in the process and further did the ChIP-on-ChIP analysis by using the customer array tiling the selected changed genes and 5 genomic loci hybridizing with ChIP’d DNA of histone markers (H3K4me3, H3K9me3 and H3K27me3). We found that the obvious decrease of H3K4me3 binding and increase of H3K9me3 and H3K27me3 are observed in the promoter of common down-regulated genes. These data indicated histone H3K4, H3K9 and H3K27 marker play important roles in shaping the chromatin state and regulating gene expression. [Conclusion] We identified the signature genes controlling the inducible differentiation of promyelocytic leukemia cells and found the epigenetic mechanism regulating the gene expression in the process.

Citation Format: Chunhua Song, Xiaokang Pan, Zhanjun Li, Yali Ding, Sunil Mudhusami, Chandrika Gowda, Zheng Ge, Sadie Steffens, Bi-hua Tan, Sinisa Dovat. Epigenetic control of signature gene expression in the inducible differentiation of acute promyelocytic leukemia cells. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 71. doi:10.1158/1538-7445.AM2015-71
Abstract 715: Regulation of the ABCG2 drug efflux transporter in breast cancer cells
Authors: Chua Vivian YL , Harvey Jennet , Bentel Jacqueline
Published: August 2015

The ABC transporters, including ABCG2 or breast cancer resistance protein (BCRP) are drug efflux transporters. Their overexpression in cancers including breast cancer has been associated with the development of multidrug resistance (MDR) due to ABC transporter-mediated export from tumour cells of therapeutic drugs such as chemotherapeutic (eg. mitoxantrone, doxorubicin) and targeted agents (eg. tyrosine kinase inhibitors). PCR microarray analysis identified that ABCG2 expression was downregulated in the breast cancer cell lines, MCF-7 and T-47D following treatment of cells with the androgen, 5α-dihydrotestosterone (DHT), the Hedgehog pathway inhibitor, cyclopamine, or a combination of 10-8M DHT and 2μM cyclopamine. RT-qPCR analysis confirmed that DHT and cyclopamine treatments of MCF-7 cells rapidly downregulated ABCG2 mRNA levels by 24 hours. ABCG2 protein levels were also progressively decreased over 8 days of treatment, with earlier reductions in ABCG2 levels observed in cells co-treated with DHT and cyclopamine. As the ABC transporters are primarily active on membrane surfaces as drug exporters, the intracellular localisation of ABCG2 was investigated by immunofluorescence microscopy. In MCF-7 cells, DHT and/or cyclopamine decreased ABCG2 accumulation in cell-to-cell membrane junction complexes and interestingly, accumulation of ABCG2 into cytoplasmic vesicles which resembled aggresomes was observed in cells exposed to cyclopamine. Consistent with the reduced levels of membrane-bound ABCG2, export of the fluorescent ABCG2-specific substrate, mitoxantrone from MCF-7 cells was delayed following treatment of cultures with DHT and cyclopamine, thereby indicating diminished ABCG2 efflux activities. Moreover, these DHT- and cyclopamine-treated cells were also more responsive to the cytotoxic effects of mitoxantrone as the IC50 for mitoxantrone was decreased by ∼80% in treated cells compared to controls. Resistance to drug therapies, which is often facilitated by the ABC transporters, is a major cause of treatment failure and recurrence of breast cancer. As such, development and implementation of adjuvant compounds such as androgens and Hedgehog inhibitors, which inhibit expression and function of ABCG2 and other efflux transporters in breast cancer cells, may improve treatment outcomes and prolong recurrence-free survival of patients.

Citation Format: Vivian YL Chua, Jennet Harvey, Jacqueline Bentel. Regulation of the ABCG2 drug efflux transporter in breast cancer cells. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 715. doi:10.1158/1538-7445.AM2015-715
Abstract 718: The effects of trastuzumab, paclitaxel, and carboplatin on HER2-positive cancer stem cells that are isolated from primary breast cancer cultures: a preliminary report
Authors: Artac Mehmet , Kayadibi Gozde , Ceylan Ayca , Kars Meltem Demirel , Artac Hasibe , Cakır Murat , Boruban Cem , Tekin Ahmet , Tavlı Lema , Kartal Adil
Published: August 2015

Introduction:

It is generally accepted that trastuzumab has only effective in HER2+ breast cancer patients who have HER2 gene amplification or HER2 over expression. On the other hand, trastuzumab resistance is a problem in HER2+ patients. Our aim is to investigate the effects of trastuzumab, paclitaxel and carboplatin on cancer stem cells which is isolated from primary breast cancer cultures.

Methods:

Tumor tissue was obtained in the routine surgery of a breast cancer patient who was HER2+ by immunohistochemistry. Primary breast cancer culture were established. CD44+CD24- and ALDH+ breast cancer stem cells were sorted by flowcytometry from these culture. HER2 receptor status of the sorted cells was also determined by flowcytometry. HER2+ and HER2- breast cancer stem cells were treated with trastuzumab, paclitaxel and carboplatin, and combination of these drugs by checker board microplate method for 96 h. For all of these breast cancer stem cell subgroups, the responses to the drugs were determined.

Results:

Breast cancer stem cells were isolated from a HER2+ breast cancer patient. The cell sorting analysis results demonstrates that 43% of the population is CD44+CD24- and 22% of the population is ALDH+. CD44+CD24- breast cancer stem cell subpopulation contains 84% HER2+ and 16% HER2- cells. Carboplatin has the highest antiproliferative effect on cancer stem cells among the three drugs which means carboplatin application resulted in the lowest IC50 values in all subgroups (IC50: 0.26 μM- 0.41 μM). HER2- cancer stem cells are more resistant to the drugs (1.5- 2 fold resistance) than HER2+ subcultures. Although combination of drugs exerted synergistic effect on the subcultures, ‘breast cancer stem cells’ were found to be more resistant to the combination therapy by exhibiting higher proliferation in drug combinations.

Conclusions:

HER2- cancer stem cells may be one of reason for trastuzumab resistance in HER2+ breast cancer patients. The result of this experimental study may be used to suggest new treatment strategies in HER2+ breast cancer patients.

Citation Format: Mehmet Artac, Gozde Kayadibi, Ayca Ceylan, Meltem Demirel Kars, Hasibe Artac, Murat Cakır, Cem Boruban, Ahmet Tekin, Lema Tavlı, Adil Kartal. The effects of trastuzumab, paclitaxel, and carboplatin on HER2-positive cancer stem cells that are isolated from primary breast cancer cultures: a preliminary report. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 718. doi:10.1158/1538-7445.AM2015-718
Abstract 722: Caveolin-1-dependent endocytosis enhanced chemosensitivity of TDM-1 in HER-2-positive breast cancer cells
Authors: Chung Yuan-Chiang , Wei Wan-Chen , Chang King-Jen , Chao Wei-Ting
Published: August 2015

Trastuzumab emtansine (TDM-1, kadcyla) is the antibody drug conjugate (ADC) was approved to treat HER-2 positive metastatic breast cancer. Although its effectiveness, however, the biomarker of drug resistance of TDM-1 has not been identified. The efficacy of ADC is restricted to the antibody internalization in the cell; therefore, the drug sensitivity may be associated with cellular vesicle trafficking system. Caveolin-1, which is the 21 KD membrane protein that is required for caveolae structure formation and membrane endocytic transport. In this study, the relationship of caveolin-1 expression and the chemosensitivity of TDM-1 in HER-2 positive breast cancer cells was investigated. Immunohistochemical staining of 32 specimens of breast cancer patients were evaluated for caveolin-1 expression. For in vitro study, BT-474 and SKBR-3 breast cancer cells which represent different caveolin-1 expression were treated with trastuzumab or T-DM1. Cell viability and molecular localization were examined. Cellular caveolin-1 expression was also manipulated by transfection of GFP tagged caveolin-1 or caveolin-1 shRNA plasmid, cell viability was then determined after drug treatment. In the results, Immunohistochemical study showed 68% (22/32) overexpression of Caveolin-1 in breast cancerous specimens. And eight cases with high expression of Caveolin-1 were found in HER-2 positive breast cancerous patients (72.1%, 8/11). In cell biology study, T-DM1 was found colocalized with caveolin-1 by confocal microscopy. SKBR-3 which represents higher caveolin-1 expression than BT-474 was more sensitive to T-DM1 treatment. After molecular transfection, BT-474 cells with overexpression of caveolin-1 increased chemosensitivity in the treatment of TDM-1, instead, SKBR-3 cells transfected with caveolin-1 shRNA decreased T-DM1 sensitivity. Expression of caveolin-1 could mediate endocytosis and promoting the internalization of TDM-1 into HER-2 positive cancer cells. Thereafter, caveolin-1 may be an effective predictor for TDM-1 treatment in the future.

Citation Format: Yuan-Chiang Chung, Wan-Chen Wei, King-Jen Chang, Wei-Ting Chao. Caveolin-1-dependent endocytosis enhanced chemosensitivity of TDM-1 in HER-2-positive breast cancer cells. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 722. doi:10.1158/1538-7445.AM2015-722