bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 1975: AICAR induces cell death in LKB1-deficient NSCLC.
Authors: Liu Fakeng , Zhou Wei , Liu Xiuju , Sun Shi-yong , Khuri Fadlo , He Yu-long , Zhong Diansheng
Published: August 2014

AICAR is a well known AMPK activator, and AICAR-induced AMPK activation has been shown to inhibit cell proliferation or induce apoptosis through various mechanisms, such as the inactivation of metabolic enzyme involved ATP consumption, the inhibition of the of fatty acid synthesisPI3K/AKT pathway, the activation of cell cycle regulators, or the activation of the JNK pathway. LKB1 is an upstream AMPK kinase, and energetic stress-induced AMPK activation usually requires LKB1. Interestingly, AICAR has also been shown to induce caspase-3 cleavage in LKB1- null MEF and ovarian cancer cells, indicating AICAR can induce cell killing through an AMPK- independent mechanism. Because LKB1 is most frequently inactivated in non-small cell lung cancer, we evaluated the effect of AICAR in lung cancer cell lines. AICAR preferentially induced caspase-3 cleavage in LKB1- mutant NSCLC cell lines H157, A549, H460 and H1944, but not in LKB1- wild type cell lines H1299, H358, H1650 and H1792. Transient depletion of LKB1 by RNAi in H1299 and H1792 cells promoted AICAR-induced caaspase-3 cleavage in the absence of AMPK activation, supporting the notion that AICAR induces cell killing in LKB1-depleted cells through an AMPK- independent mechanism. In LKB1-null H460 cells, AICAR treatment led to a significant increase in S phase cell populations, and G1 cell cycle arrest by serum starvation is was sufficient to prevent AICAR-induced cell death. These data indicated that AICAR-induced cell killing in LKB1- deficient cells is cell cycle dependent. In summary, the ability of AICAR to preferentially induce cell death in LKB1- deficient NSCLC suggestsed that LKB1 deficiency may be a therapeutic target for the treatment of NSCLC.

Citation Format: Fakeng Liu, Wei Zhou, Xiuju Liu, Shi-yong Sun, Fadlo Khuri, Yu-long He, Diansheng Zhong. AICAR induces cell death in LKB1-deficient NSCLC. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 1975. doi:10.1158/1538-7445.AM2013-1975
Abstract 1982: TCEB3C (Elongin A3) on chromosome 18 presents a putative tumor suppressor gene of small intestine neuroendocrine tumors.
Authors: Edfeldt Katarina , Ahmad Tanvver , Åkerström Göran , Tiensuu Janson Eva , Hellman Per , Stålberg Peter , Björklund Peyman , Westin Gunnar
Published: August 2014

Small intestine neuroendocrine tumors (SI-NETs) arise from the enterochromaffin cells and produce hormones which can cause the carcinoid syndrome. The incidence is increasing and the five-year survival rate is 60-80 %. Loss of one copy of chromosome 18 is a very frequent event in these tumors, supporting the presence of a crucial tumor suppressor gene. This study investigated whether TCEB3C, encoding Elongin A3, presents a putative tumor suppressor gene in SI-NETs. This gene is currently the only known imprinted gene on chromosome 18, requiring only one gene copy inactivation event to fulfill the Knudson hypothesis.

Elongin A3 expression in SI-NETs (n=47) was heterogeneous and negative in most cells. 5-Aza-2’-deoxycytidine induced expression of Elongin A3 in a carcinoid tumor cell line and in tumor derived primary cell cultures, but not in a control cell line. Also the general histone methyltransferase inhibitor 3-deazaneplanocin A induced TCEB3C expression in the carcinoid tumor cell line, but not in control cells suggesting further that the TCEB3C gene was epigenetically suppressed in neuroendocrine tumor cells. The TCEB3C gene encompasses an CpG island and quantitative cytosine CpG methylation analysis will be presented. Furthermore, overexpression of Elongin A3 led to a decrease in clonogenic survival of carcinoid tumor cells, but not of control cells strongly supporting a growth-regulatory role. These results support a putative tumor suppressor role of TCEB3C in neuroendocrine cells of the small intestine.

Citation Format: Katarina Edfeldt, Tanvver Ahmad, Göran Åkerström, Eva Tiensuu Janson, Per Hellman, Peter Stålberg, Peyman Björklund, Gunnar Westin. TCEB3C (Elongin A3) on chromosome 18 presents a putative tumor suppressor gene of small intestine neuroendocrine tumors. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 1982. doi:10.1158/1538-7445.AM2013-1982
Abstract 1992: Is the ubiquitin ligase WWP2 playing a role in PTEN degradation in endometrial cancer.
Authors: Clements Aine , McElroy Joseph , Suarez Adrian , Majumder Sarmila , Cohn David , Leone Gustavo
Published: August 2014

Objective: The ubiquitin ligase WWP2 has recently been described to directly interact with PTEN, the major tumor suppressor gene in the PI3K pathway commonly altered in endometrial cancer. Whether alterations of WWP2 contribute to post-translational modifications of the PTEN protein in endometrial cancer are unknown. We sought to determine if WWP2 gene expression levels differ in human endometrial cancer tissues with evidence of post-translational PTEN degradation. We also wished to determine if elevated WWP2 gene expression was associated with clinical prognostic variables.

Methods: Twenty-four endometrial cancer specimens collected at hysterectomy were available for analysis. These tumors were stained for PTEN by immunohistochemistry (IHC) and scored using a scale of 0-6. Frozen tissue was then used to determine PTEN and WWP2 expression by Real time PCR (RT PCR). Gene expression was then compared with findings on immunohistochemistry. Patient clinical characteristics including stage, grade, lymph node metastases, recurrence or 2 year disease status were then compared with PTEN and WWP2 expression.

Results: Three tumors had elevated PTEN gene expression (at least 2 fold) but low (score of less than 4) PTEN protein on IHC, indicating post-translational modification of PTEN. In these tumors, however, WWP2 expression was not elevated (range 0.004-0.09). There were three tumors with elevation in WWP2 expression (1.9-4.7). PTEN was low in these tumors on IHC, but was also low by RT-PCR indicating genomic, epigenetic or post-transcriptional modification of PTEN. Neither PTEN nor WWP2 expression on RT PCR was associated with stage, grade, lymph node metastases, recurrence or 2 year disease status.

Conclusions: In this cohort of tumors, WWP2 did not contribute to post-translational degradation of PTEN. WWP2 expression was not associated with stage, grade, lymph node metastases, recurrence, or 2 year disease status.

Citation Format: Aine Clements, Joseph McElroy, Adrian Suarez, Sarmila Majumder, David Cohn, Gustavo Leone. Is the ubiquitin ligase WWP2 playing a role in PTEN degradation in endometrial cancer. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 1992. doi:10.1158/1538-7445.AM2013-1992
Abstract 2005: Analysis of gene expression and copy number variation in breast tumors using both sequencing and hybridization-based platforms.
Authors: Norton Nadine , Perez Edith A. , Asmann Yan W. , Carr Jennifer M. , Necela Brian M. , Kachergus Jennifer M. , Jen Jin , Eckloff Bruce W. , Thompson E Aubrey
Published: August 2014

Next Generation Sequencing (NGS) technologies provide rapid genomic analyses of single nucleotide variants, RNA expression and DNA copy number. Application of these technologies to material isolated from formalin fixed paraffin embedded (FFPE) tissue and even degraded frozen material could provide powerful replication samples but remains challenging. We tested the nanoString platform to validate deep sequence analysis of gene expression and DNA copy number in degraded and FFPE material. Firstly, RNA from the Universal Human Reference RNA and a breast cancer cell line (MDA-MB-436) was artificially degraded to different degrees (RIN 1.2-6.8). We used the nanoString platform to simultaneously measure RNA expression across 226 genes in each degraded sample and the corresponding undegraded RNA. Secondly we isolated RNA and DNA from matched fresh frozen and FFPE tissues from nine breast cancer patients (3 HER2+/ER+/PR+, 2 HER2+/ER-/PR-, 2 HER2+/ER+/PR-, 2 HER2-/ER+/PR+) using the nanoString platform to compare expression and copy number across 226 and 86 genes respectively. Finally, we correlated expression and copy number data generated by nanoString with Illumina transcriptome and whole genome sequencing (WGS). NanoString log2 expression fold-change between all artificially degraded samples and their undegraded counterpart showed extremely high correlation (r2>0.91). NanoString DNA copy number between matched fresh-frozen and FFPE showed a high degree of correlation (r2=0.71). All gene amplifications with copy number ≥ 5 in DNA from fresh-frozen material (N=9) were successfully identified in DNA from FFPE material. We also observed good correlation of gene expression between whole transcriptome sequencing and the nanoString platform (r2 0.59 - 0.72) in FFPE and artificially degraded material and for DNA copy number between WGS and nanoString in DNA isolated from cancer cell lines (r2=0.96). The nanoString platform provides reliable data from highly degraded and FFPE material and correlates with sequence analysis of both expression and copy number from NGS platforms demonstrating potential for large-scale replication studies in FFPE material.

Citation Format: Nadine Norton, Edith A. Perez, Yan W. Asmann, Jennifer M. Carr, Brian M. Necela, Jennifer M. Kachergus, Jin Jen, Bruce W. Eckloff, E Aubrey Thompson. Analysis of gene expression and copy number variation in breast tumors using both sequencing and hybridization-based platforms. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 2005. doi:10.1158/1538-7445.AM2013-2005
Abstract 2007: The NF-kB pathway is constitutively active in endometrial cancer.
Authors: Noonan Anne M. , Casablanca Yovanni , Maxwell George L. , Annunziata Christina M.
Published: August 2014

The NF-κB Pathway is Constitutively Active in Endometrial Cancer

Metastatic endometrial cancer (EC) is a disease for which therapeutic options are limited and prognosis is poor. Although 75% of endometrial cancers are diagnosed at an early stage and cured by surgery, 15-20% develop metastases and die from their disease. Uterine papillary serous carcinoma (UPSC) occurs in less than 10% of patients, but has a worse clinical outcome. Prior studies suggest that NF-kB signaling may play a role in UPSC. We therefore proceeded with a detailed investigation of the NF-κB pathway in EC.

Methods: A set of 11 endometrial cancer cell lines, representing endometrioid EC and UPSC, were tested for sensitivity to a specific inhibitor of IKKβ after 3 days of drug treatment using XTT assay. Western blot was used to assess changes in NF-κB proteins after treatment of sensitive UPSC cell lines ARK-1 and ACI-126 with IKKβ inhibitor over a time series ranging from 15 min to 24h. RNA was extracted from ARK-1 and ACI-126 cell lines after 48h treatment with IKK-2 inhibitor IV or DMSO control, and after transfection of the same cell lines with IκBα super-repressor or empty vector control. RNA was hybridized to Affymetrix U133 Plus2.0 microarray platform for gene expression profiling and the data was analyzed using mAdb. Gene and protein expression was examined in patient samples to verify biologic relevance.

Results: Six of 11 cell lines were sensitive to IKKβ inhibitor with IC50s ranging between 4 and 10uM. The highest sensitivity occurred in two UPSC cell lines ARK-1 and ACI-126. Following pharmacologic IKKβ inhibition, nuclear levels of NF-κB p65 decreased by 3h and began to increase again from 6h to 24h. Cytoplasmic NF-κB p65 increased from 15 min to 3h and then began to fall. Gene expression profiling after IKKβ inhibition, either by drug treatment or by mutant IkBa super-repressor, revealed an overlapping gene signature in the two UPSC cell lines. The genes were co-expressed across the gene expression profiles of patients with EC, and were significantly highest in African American women with UPSC.

Conclusions: There is aberrant signaling through the NF-κB pathway in a subset of EC. An IKKβ target gene signature shows that this pathway is most relevant in African American women with UPSC. Therefore, targeting NF-kB may provide a novel treatment strategy for this underserved population with a rare variant of EC.

Citation Format: Anne M. Noonan, Yovanni Casablanca, George L. Maxwell, Christina M. Annunziata. The NF-kB pathway is constitutively active in endometrial cancer. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 2007. doi:10.1158/1538-7445.AM2013-2007
Abstract 201: (-)-Epigallocatechin-3-gallate inhibits the development of hepatic preneoplastic lesions in obese and hypertensive rats, a novel model of non-alcoholic steatohepatitis-related liver tumorigenesis.
Authors: Kochi Takahiro , Shimizu Masahito , Tanaka Takuji , Moriwaki Hisataka
Published: August 2014

Background: Non-alcoholic steatohepatitis (NASH), which involves hepatic inflammation and fibrosis, is associated with liver carcinogenesis. The activation of the renin-angiotensin system (RAS), which plays a key role in blood pressure regulation, promotes hepatic fibrogenesis. In this study, we investigated the effects of (-)-epigallocatechin-3-gallate (EGCG), a major component of green tea catechins, on the development of hepatic preneoplastic lesions in SHRSP.Z-Leprfa/IzmDmcr (SHRSP-ZF) obese and hypertensive rats. Methods: Male 7-week-old SHRSP-ZF rats and control non-obese and normotensive WKY/Izm rats were fed a high fat diet and received intraperitoneal injections of carbon tetrachloride twice a week for 8 weeks. The rats were also provided tap water containing 0.1% EGCG during the experiment. Results: SHRSP-ZF rats presented with obesity, insulin resistance, dyslipidemia, an imbalance of adipokines in the serum, and hepatic steatosis. The development of hepatic preneoplastic lesions and liver fibrosis was markedly accelerated in SHRSP-ZF rats compared to that in control rats. Additionally, in SHRSP-ZF rats, RAS was activated and inflammation and oxidative stress were induced. Administration of EGCG, however, inhibited the development of hepatic premalignant lesions by improving liver fibrosis, inhibiting RAS activation, and attenuating inflammation and oxidative stress in SHRSP-ZF rats. Conclusions: Obese and hypertensive SHRSP-ZF rats treated with a high fat diet and carbon tetrachloride displayed the histopathological and pathophysiological characteristics of NASH and developed hepatic premalignant lesions, suggesting the model might be useful for the evaluation of NASH-related liver tumorigenesis. EGCG might also be able to prevent NASH-related liver fibrosis and tumorigenesis.

Citation Format: Takahiro Kochi, Masahito Shimizu, Takuji Tanaka, Hisataka Moriwaki. (-)-Epigallocatechin-3-gallate inhibits the development of hepatic preneoplastic lesions in obese and hypertensive rats, a novel model of non-alcoholic steatohepatitis-related liver tumorigenesis. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 201. doi:10.1158/1538-7445.AM2013-201
Abstract 2012: Ultra-deep sequencing of circulating free DNA to identify predictive, mutated HSP90 clients in the GALAXY TrialTM (NCT01348126): a randomized phase IIB/III study of ganetespib (STA-9090) in combination with docetaxel versus docetaxel alone in subjects wit
Authors: Fennell Dean Anthony , Martins L. Miguel , El-Hariry Iman , Vukovic Vojo , Teofilovici Florentina , Reichert Vienna L. , Shaw Jacqui
Published: August 2014

Background: Non-small cell lung cancer is a mosaic, comprising several distinct molecular subtypes driven by somatic mutations in so-called cancer genes. Several of these oncogenic mutations confer dependence on HSP90 e.g. EML4-ALK, BRAF, KRAS, KIT. Inhibition of Hsp90 induces apoptosis and can exploit this dependence. Ganetespib (G) is an Hsp90 inhibitor with single agent activity in molecularly defined disease, including EML4-ALK rearrangement, KRAS mutations, HER2 amplification and BRAF mutations. Circulating free DNA (cfDNA) is present at low levels in plasma of healthy individuals allowing detection of somatic mutations by deep sequencing. The aim of this work was to determine the mutational spectrum of patients enrolled into the GALAXY trial using this liquid biopsy strategy.

Methods: We performed a prospective exploratory analysis to identify plasma-borne somatic mutations as predictors of clinical outcome with G in GALAXY, a randomized trial, comparing G + docetaxel (D), to D alone in 2nd line advanced NSCLC patients. Plasma samples were collected from approximately 200 patients with adenocarcinoma at baseline prior to initiation of treatment, and during cycles 1 and 2. cfDNA samples were evaluated using the Ion AmpliSeq™ Cancer Panel on the Ion Torrent |PGM platform to survey 739 amplicons in 46 cancer genes at up to 6000x depth.

Results: CfDNA targeted sequence analysis of the first 38 patients revealed multiple concurrent mutations in client proteins including the HSP90 client proteins BRAF, PDGFR and KIT, demonstrating the feasibility of this method. Sequencing of a larger cohort is underway. Longitudinal sampling of plasma has been conducted to monitor temporal evolution of the penetrance of mutations.

Summary: Ultra-deep re-sequencing of multiple somatic mutations in circulating cfDNA is feasible, and can potentially enable identification of G sensitive subgroups. This represents a new approach to biomarker discovery in the context of phase II trials.

Citation Format: Dean Anthony Fennell, L. Miguel Martins, Iman El-Hariry, Vojo Vukovic, Florentina Teofilovici, Vienna L. Reichert, Jacqui Shaw. Ultra-deep sequencing of circulating free DNA to identify predictive, mutated HSP90 clients in the GALAXY TrialTM (NCT01348126): a randomized phase IIB/III study of ganetespib (STA-9090) in combination with docetaxel versus docetaxel alone in subjects wit [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 2012. doi:10.1158/1538-7445.AM2013-2012
Abstract 2017: Exome sequencing reveals novel mutations in a GIST and a paraganglioma occurring simultaneously in a patient with complete form of Carney triad.
Authors: Haller Florian , Agaimy Abbas , Moskalev Evgeny A. , Jäger Natalie , Broers Benedikt , Wiemann Stefan , Hartmann Arndt
Published: August 2014

Carney triad is a rare syndrome which is defined by the occurrence of multicentric gastric GIST, paraganglioma and pulmonary chondroma. In contrast to the closely-related Carney-Stratakis syndrome (CSS), Carney syndrome is not inherited. In both syndromes, the tumors are characterized by a loss of Succinate Dehydrogenase B protein (so-called SDHB-deficient tumors). Only recently, inactivating mutations in the SDH subunits A, B, C, and D have been found to be present in the majority of SDHB-deficient GISTs of CSS, but not in GISTs in the setting of Carney triad. We performed exome sequencing of DNA samples derived from blood as well as of the GIST and the paraganglioma from a female patient with a complete Carney triad. Additionally, the genome from the blood DNA was sequenced at high coverage, and low coverage genomes of both tumor DNAs were evaluated for copy number variations. SDHA and B mRNA and protein expression were analysed by qRT-PCR and Western Blot analysis, respectively. Both the GIST and the paraganglioma from the Carney patient were negative for SDHB by immunohistochemistry, and did not display noticeable levels of SDHB in the Western Blot analysis. In contrast, SDHB mRNA levels were equal to SDHA levels, and also to GISTs from sporadic GISTs with KIT mutations. Exome sequencing revealed 10 and 18 somatic missense mutations in the DNA from the GIST and the paraganglioma, which were not present in the DNA from the blood. Additionally, one somatic insertion/deletion was found in each tumor. These novel mutations will provide better insights in our understanding of this rare albeit fascinating disease, and might also be suitable as targets for future therapies.

Citation Format: Florian Haller, Abbas Agaimy, Evgeny A. Moskalev, Natalie Jäger, Benedikt Broers, Stefan Wiemann, Arndt Hartmann. Exome sequencing reveals novel mutations in a GIST and a paraganglioma occurring simultaneously in a patient with complete form of Carney triad. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 2017. doi:10.1158/1538-7445.AM2013-2017
Abstract 2018: Genome-wide analysis of DNA methylation and copy number changes in breast cancers.
Authors: Stefansson Olafur A. , Moran Sebastian , Gomez Antonio , Puig Sergi Sayols , Eyfjord Jorunn , Esteller Manel
Published: August 2014

Epigenetic modifications are known to be critical in regulating normal developmental processes and cellular differentiation. DNA methylation of CpG dinucleotides is a well-characterized epigenetic marker frequently associated with transcriptional silencing. CpG dinucleotides are non-randomly distributed throughout the genome with regions enriched for CpGs being known as CpG islands. It is now well established that epigenetic inactivation of tumor suppressor genes can occur by CpG island promoter methylation. For example, BRCA1 promoter methylation occurs in approximately 10 - 15% of all breast cancers and is associated with gene silencing. Recent technological advances have enabled DNA methylation to be profiled genome-wide. Using the Infinium 450 thousand probe arrays, we have studied DNA methylation and copy number changes to discover new markers of potential clinical relevance in breast cancer. Expression analyses by immunohistochemisty on tissue arrays for subtype-specific markers (ER, PR, HER2, Ki-67, EGFR, CK5/6, CK8, CK18, MUC1 and Claudin-3) carried out previously were used to establish DNA methylation markers of breast cancer subtypes. This integrated approach is presented in clinical context with available data on histological grade, disease stage and breast cancer-specific survival outcomes.

Citation Format: Olafur A. Stefansson, Sebastian Moran, Antonio Gomez, Sergi Sayols Puig, Jorunn Eyfjord, Manel Esteller. Genome-wide analysis of DNA methylation and copy number changes in breast cancers. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 2018. doi:10.1158/1538-7445.AM2013-2018
Abstract 2020: Next-generation sequencing for mutation discovery in Ewing sarcoma.
Authors: Brohl Andrew S. , Song Young , Hurd Laura , Liao Hongling , Chen Li , Patidar Rajesh , Shern John F. , Wei Jun S. , Khan Javed
Published: August 2014

Background: Ewing sarcoma (EWS) is the second most common type of primary bone tumor to affect children and adolescent and accounts for 2.9% of all childhood cancers. Despite advances in multidisciplinary treatment leading to improved outcomes over time, long term survival still remains less than 50% for patients older than 15 years of age and only 65% in younger patients. In an effort to identify novel prognostic and therapeutic targets, we utilized next generation sequencing techniques to evaluate EWS samples for somatic mutation.

Methods: Whole genome paired-end sequencing using the Complete Genomics method was performed on the DNA from 6 EWS matched tumor-normal samples. Somatic mutations were identified and classified using standard bioinformatics techniques and mutations of interest were verified using capillary sequencing. To extend discoveries and evaluate for mutational recurrence, targeted sequencing was performed on a validation cohort of 61 EWS tumors and 22 EWS cell lines using a custom multiplex PCR design followed by high-coverage sequencing on the Ion Torrent system.

Results: In the discovery cohort, we detected an average of 361 somatic mutations per tumor in non-repetitive regions and an average of 6 somatic mutations per tumor in protein coding regions (=0.15 mutations/Mb of coding sequence). The well-established FLI1-EWSR1 gene fusion was detected by the whole genome sequencing in all 6 samples. Additional previously reported genetic events were also seen including frequent chromosomal trisomy and deletion of the CDKN2A containing locus on chromosomal region 9p21 in 2 of 6 samples. In the validation cohort of tumors, known recurrent mutations such as in TP53 (3/61 = 4.9%) were seen at rates similar to that previously reported in EWS. Continued efforts are underway to verify novel recurrent mutations.

Conclusion: Next-generation sequencing provides a powerful tool to characterize the genetic landscape and evaluate for recurrent mutations in cancer. We utilize this tool in Ewing sarcoma in a project that provides both a comprehensive genomic evaluation from whole genome sequencing data and as well evaluates for recurrent mutations using a targeted sequencing approach.

Citation Format: Andrew S. Brohl, Young Song, Laura Hurd, Hongling Liao, Li Chen, Rajesh Patidar, John F. Shern, Jun S. Wei, Javed Khan. Next-generation sequencing for mutation discovery in Ewing sarcoma. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 2020. doi:10.1158/1538-7445.AM2013-2020
Abstract 2021: Characterization of myeloma tumors from a multi-ethnic cohort using cytogenetics and genomic analysis.
Authors: Baker Angela S. , Braggio Esteban , Jacobus Susana , Jung Sungwon , Larson Dirk , Therneau Terry , Dispenzieri Angela , Van Wier Scott A. , Ahmann Gregory , Levy Joan , Perkins Louise , Kim Seungchan , Henderson Kim , Vesole David , Rajkumar S. Vincent , Jelinek Dianne F. , Carpten John , Fonseca Rafael
Published: August 2014

The plasma cell malignancy Multiple Myeloma (MM) is a rare but deadly form of cancer with 5 year survival rates of ∼30%. Epidemiological studies have suggested that MM exacts an especially heavy burden on African American (AA) patients. We examined a multi-ethnic cohort of MM tumors to assess potential differences in the frequency of molecular events in tumors derived from either African American (AA) or European American (EA) patients. The frequency of 14q32 translocation breakpoints at the IgH locus was compared among a series of data from 115 AA MM patients from three studies and EA MM from the Eastern Cooperative Oncology Group (ECOG). In addition, we analyzed matching genome-wide copy number and transcriptional data among 45 AA MM tumors and 196 EA MM tumors to determine differences in the frequency of molecular events in tumors from these populations.

No differences were found for specific translocation subtypes; however, we detected a statistically significant difference in the overall frequency of IgH translocations between the two groups, which occurred more frequently in EA patients (52% vs. 40%; p = 0.032). When assessing genomic copy number events previously shown to be associated with poor outcome in MM, frequencies of alterations were not significant after correcting for multiple testing. Furthermore, there was not a significant difference in the association of high-risk disease using expression profiling. Our study represents the first comprehensive assessment of the frequency and distribution of molecular alterations in MM tumors from both AA and EA patients and could help shed light on possible biological features associated with population differences in incidence and outcome in MM.

Citation Format: Angela S. Baker, Esteban Braggio, Susana Jacobus, Sungwon Jung, Dirk Larson, Terry Therneau, Angela Dispenzieri, Scott A. Van Wier, Gregory Ahmann, Joan Levy, Louise Perkins, Seungchan Kim, Kim Henderson, David Vesole, S. Vincent Rajkumar, Dianne F. Jelinek, John Carpten, Rafael Fonseca. Characterization of myeloma tumors from a multi-ethnic cohort using cytogenetics and genomic analysis. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 2021. doi:10.1158/1538-7445.AM2013-2021
Abstract 2036: Overcoming platinum-resistance in ovarian cancer via targeting basonuclin1 (BNC1).
Authors: Wu Sherry Y. , Unruh Anna K. , Rupaimoole Rajesha , Ivan Cristina , Ruder Dennis , Pradeep Sunila , Sehgal Vasudha , Rodriguez-Aguayo Cristian , Pecot Chad , Miyake Takahito , Lopez-Berestein Gabriel , Ram Prahlad , Baggerly Keith A. , Sood Anil K.
Published: August 2014

Purpose This study aims to interrogate if basonuclin1 (BNC1), a key gene identified through integrative TCGA-based functional genomic analysis, is a relevant target for overcoming chemo-resistance in high-grade serous ovarian cancer (HGS-OvCa). BNC1 is a transcription factor which can regulate ribosomal biogenesis and cell proliferation. Methods TCGA mRNA expression and clinical response data were used to systematically identify genes associated with chemo-resistance in patients with HGS-OvCa. Gene-specific effects were subsequently studied for selected targets in vitro. BNC1 targeted siRNA was used to study the functional role of BNC1 in chemo-resistance. Apoptosis and cell cycle analyses were carried out following BNC1 silencing in ovarian cancer cell lines with or without the presence of cisplatin. The in vivo effects of silencing BNC1 on the chemosensitivity were tested using an orthotopic mouse model (A2780CP20) of ovarian cancer. Results The expression of BNC1 was found to be increased by more than two-fold (Agilent platform; p<0.001) in chemo-resistant tumors in the TCGA dataset. A 5-fold increase in its expression level was also observed in cisplatin-resistant A2780-CP20 and IGROV-CP20 cells compared to parental cell lines. BNC1 silencing increased the sensitivity of A2780CP20 cells to cisplatin treatment (IC50) with a 30% increase in the percentage of apoptotic cells present at 72 hours following cisplatin treatment, when compared to cells treated with control siRNA. Profound G2-cell cycle arrest was also noted following combined siBNC1 and cisplatin (IC30) treatment (68% vs. 28%; siBNC1 plus cisplatin vs. siControl plus cisplatin; p<0.0001). Importantly, combinational treatment of siBNC1 and cisplatin resulted in a 90% reduction in tumor burden in chemo-resistant A2780CP20 orthotopic mouse model when compared to control siRNA plus cisplatin treatment group (p<0.01). Conclusion Collectively, BNC1 represents an important target for enhancing chemosensitivity in ovarian tumors and its down-regulation could lead to better clinical outcome.

Citation Format: Sherry Y. Wu, Anna K. Unruh, Rajesha Rupaimoole, Cristina Ivan, Dennis Ruder, Sunila Pradeep, Vasudha Sehgal, Cristian Rodriguez-Aguayo, Chad Pecot, Takahito Miyake, Gabriel Lopez-Berestein, Prahlad Ram, Keith A. Baggerly, Anil K. Sood. Overcoming platinum-resistance in ovarian cancer via targeting basonuclin1 (BNC1). [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 2036. doi:10.1158/1538-7445.AM2013-2036
Abstract 2040: Indication switching of the FDA-approved anticancer tyrosine kinase inhibitors to treat metabolic diseases.
Authors: Sun Jiazhi , Yang Lun , Li Minghua , Wang Zhixin , Haynes Amy , Wang Qian , Raplee Isaac , Melchisedek Jean , Prasad Anand , Sneed Kevin B. , He Lin , Zhou Shufeng
Published: August 2014

Drug switching is a unique way to identify and extend new indications for approved drugs in drug discovery & development. Tyrosine kinase inhibitors (TKIs) are used in biologically targeted cancer therapy and so far 12 TKIs have been approved by FDA. We hypothesize that these TKIs act on other molecular targets in addition to tyrosine kinases and may manage metabolic diseases given that there is a complex network of kinases that work together to regulate a number of important cellular processes. Employing a comprehensive docking method with our established chemical-protein interactome (CPI) and 11 FDA-approved TKIs, we have discovered301 PDB-deposited proteins corresponding to 353 ligand binding pockets among a total of 1,780 PDB-deposited human protein entries. Notably, sorafenib, dasatinib and crizotinib had a CPI binding score (ZZ_score) of -1.2903, -1.0278 and -1.5384 against histone deacetylase 7A, respectively. In addition, those TKIs achieved high ZZ scores against B-Raf, PPAR and VDR, suggesting a high binding affinity of sorafenib, dasatinib and crizotinib with these proteins. Our preliminary studies have showed that the acetylated-lysine in α-tublin and oncogenic Akt and Raf-signaling was inhibited significantly in human multiple melanoma cells by these TKIs. Interestingly, dasatinib increased VDRE and HDLR luciferase reporter activity in the human lung adenocarcinoma A-549 cells, and dasatinib and crizotinib induced autophage by activated LC3 in vitro. Furthermore, the erlotinib and sorafenib decreased glucose levels in Wistar STZ-D rats. These TKIs are predicted to act on a series of therapeutics targets associated with metabolic diseases (such as PPAR and Sirt1). Collectively, FDA-approved TKIs may be switched to become a “magic bullet” concurrently targeting tyrosine kinase, HDAC, PPAR, VDR and B-Raf, shedding a light for future anti-metabolic disorder and anticancer drug discovery & development. Further validation of additional “hot targets” besides tyrosine kinase such as HDAC, B-Raf, PPAR, Sirt1, Akt and VDR, and in vivo evaluation of anti-metabolic disorder by TKIs are undergoing at our laboratory. These findings suggest that drug switching may represent a new and effective approach to expanding the application of existing drugs.

Citation Format: Jiazhi Sun, Lun Yang, Minghua Li, Zhixin Wang, Amy Haynes, Qian Wang, Isaac Raplee, Jean Melchisedek, Anand Prasad, Kevin B. Sneed, Lin He, Shufeng Zhou. Indication switching of the FDA-approved anticancer tyrosine kinase inhibitors to treat metabolic diseases. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 2040. doi:10.1158/1538-7445.AM2013-2040
Abstract 2047: Discovery and molecular characterization of AGS-15/SLITRK6 as a novel target for antibody-mediated therapeutic development in bladder cancer.
Authors: Shostak Yuriy , Said Suzanne , Russell Deanna L. , Mattie Michael D. , Chang Mi Sook , Christensen Ashley , Morrison Karen , Morrison Kendall , Stover David , Challita-Eid Pia
Published: August 2014

Discovery efforts utilizing suppression subtractive hybridization identified AGS-15/SLITRK6 as a differentially expressed gene in bladder cancer. SLITRK6 encodes a cell surface type I transmembrane protein, a member of the SLITRK family of factors with neuronal function activities. Comprehensive SLITRK6 RNA expression analysis in patient specimens revealed limited restricted normal tissue expression and high frequent expression in bladder cancers, as well as expression in subsets of breast, lung and several other cancer types. Tissue immunohistochemical analysis validated SLITRK6 protein limited expression in normal tissues and high expression levels in cancers. SLITRK6 expression profiling in cancer cell lines and Agensys proprietary patient-derived xenografts (PDXs) identified model systems in relevant cancer indications for further target characterization and antibody therapeutics development. Genomic analyses, including whole exome next generation sequencing, chromosomal aberrations and gene copy number assessments, were performed to evaluate SLITRK6 gene status in model cell lines and PDXs. Thus, we discovered and characterized AGS-15/SLITRK6 as a novel target with high levels and selective expression in tumors, particularly in bladder cancer. This cell surface antigen's attractive expression profile is a major determinant of AGS-15/SLITRK6 as a suitable and preferential candidate for antibody drug conjugate therapeutic targeting in bladder cancer treatment and, potentially, in several other cancer indications.

Citation Format: Yuriy Shostak, Suzanne Said, Deanna L. Russell, Michael D. Mattie, Mi Sook Chang, Ashley Christensen, Karen Morrison, Kendall Morrison, David Stover, Pia Challita-Eid. Discovery and molecular characterization of AGS-15/SLITRK6 as a novel target for antibody-mediated therapeutic development in bladder cancer. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 2047. doi:10.1158/1538-7445.AM2013-2047
Abstract 2049: A novel, potent, Cyp17 inhibitor with favorable pharmacokinetics and oral bioavailability for potential treatment of human castration resistant prostate cancer (CRPC).
Authors: Feng Ying , Wang Bing , Chu Daniel , Post Leonard , Shen Yuqiao
Published: August 2014

Cytochrome P450 17α-hydroxylase/C17,20-lyase (CYP17) is a key enzyme in androgen bio-synthesis in adrenals and testes. It has been validated clinically as an effective target for CRPC through successful clinical trials with abiraterone acetate. Abiraterone acetate is a selective, potent inhibitor of Cyp17, and was recently approved as a treatment for CRPC. We have designed a novel Cyp17 inhibitor, BMN680, with improved pharmacokinetic (PK) profile and oral bioavailability while maintaining potent Cyp17 inhibition. BMN680 inhibits Cyp17 lyase activity with IC50 of 0.8 nM and reduces testosterone production in human adrenal cancer cell line H295R cells with IC50 of 3.87 nM. in vitro metabolic stability studies indicate that BMN680 has a half-life of 5-hr in human liver microsome and 8-hr half-life in human hepatocytes. Animal PK profiling in rats and monkeys demonstrated BMN680 has in vivo half-life of >4 hr and oral bioavailability of >30%. Single oral dosing of BMN680 at 10 mg/kg in normal, intact, Cynomolgus monkeys resulted in suppression of plasma testosterone level for up to 24 hour. Pharmacodynamic (PD) effect of single and multi-day oral dosing of BMN680 in monkey will be presented. In human liver microsome Cyp panel screen, BMN680 exhibited IC50 greater than 1.5 uM for all Cyp enzymes tested including 1A2, 2C9, 2C19, 2D6, 3A4, indicating low potential of drug-drug interaction. In conclusion, BMN680 is a potent Cyp17 inhibitor with low interaction with Cyp450 isoforms, and desirable PK and oral bioavailability. BMN680 can effectively suppress testosterone production in vivo, and will be further evaluated in preclinical development.

Citation Format: Ying Feng, Bing Wang, Daniel Chu, Leonard Post, Yuqiao Shen. A novel, potent, Cyp17 inhibitor with favorable pharmacokinetics and oral bioavailability for potential treatment of human castration resistant prostate cancer (CRPC). [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 2049. doi:10.1158/1538-7445.AM2013-2049
Abstract 2059: Kinase library siRNA screen identifies KSR1 as a synergistic therapeutic target in combination with TIC10.
Authors: Allen Joshua E. , El-Deiry Wafik S.
Published: August 2014

Apo2L/TRAIL is an endogenous antitumor protein that possesses efficacy-limiting properties such as short serum half-life, stability, cost, and limitations in delivery when used as a recombinant protein therapeutic. TIC10 is a first-in-class small molecule inducer of the human TRAIL gene that improves these unfavorable drug properties to ultimately increase the efficacy and utility of TRAIL. TIC10 upregulates TRAIL gene transcription by activating the pro-apoptotic transcription factor Foxo3a, which is achieved through dual inhibition of Akt and MEK/ERK. We conducted a siRNA screen using a kinase library to identify kinases that affect TIC10 response in cancer cells assayed by cell viability assays to gain mechanistic insight into the molecular mechanisms of TIC10 and potentially identify a combinatorial target to improve the activity of TIC10. Execution and validation of the siRNA screen revealed 4 kinases (DGKD, SGKL, STK23, KSR) that improved the therapeutic response to TIC10 upon siRNA-mediated knockdown at 36 hours, but not 12 hours, post-treatment in HCT116 human colon cancer cells. We next assayed the effect of knocking down these 4 kinases on cancer cell cytotoxic response to recombinant human TRAIL. These studies revealed no effect with kinase knockdowns, suggesting that the mechanisms of sensitization lie upstream of TIC10-mediated TRAIL production. Network analysis of the “hit” kinases and the previously described mechanism of TIC10 revealed rational overlap of signaling pathways and kinetics. Among the 4 identified kinases, KSR1 possessed the most direct connection to the putative mechanism of TIC10 that involves the dual inhibition of Akt and the MAPK pathway. KSR1 is a MAPK scaffold protein that positively regulates the MAPK signaling pathway. Based on these observations, we are testing the hypothesis that inhibiting KSR1 in combination with TIC10 cooperatively inhibits MAPK signaling to increase TRAIL production. Together, these results suggest that KSR1 is a key player in the cytotoxic response to TIC10 and that combining TIC10 and an inhibitor of KSR1 may result in improved therapeutic response.

Citation Format: Joshua E. Allen, Wafik S. El-Deiry. Kinase library siRNA screen identifies KSR1 as a synergistic therapeutic target in combination with TIC10. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 2059. doi:10.1158/1538-7445.AM2013-2059
Abstract 2081: Evaluation of Src as a therapeutic target in biliary tract cancer using bosutinib, a c-Src/Abl kinase inhibitor.
Authors: Nam Ah-Rong , Min Ahrum , Han Sae-Won , Song Sang-hyun , Oh Do-Youn , Im Seock-Ah , Kim Tae-You , Bang Youg-Jue
Published: August 2014

Background: Src is a nonreceptor tyrosine kinase involved in the crosstalk and mediation of many signaling pathways that promote cell proliferation, invasion and angiogenesis. Elevation of Src activity has been reported in many types of cancers including biliary tract cancer (BTC). In BTC, no molecular therapeutic target has been validated so far. Therefore, the prognosis of advanced BTC is limited and there is a huge unmet medical need in this area. The purpose of this study is to evaluate of Src as a therapeutic target in BTC and to elucidate the therapeutic potential and action mechanism of bosutinib, an orally active small molecule c-Src/Abl kinase inhibitor, alone or in combination with chemotherapy for the treatment of BTC.

Methods: Experiments were conducted using 8 human biliary tract cancer cell lines (SNU-245, 308, 478, 869, 1079, 1196, HuCCT1, TFK1). MTT assay, colony formation assay and 3D culture were done for determining growth inhibitory effect of bosutinib, alone or in combination with chemotherapeutic agents (gemcitabine, cisplatin). Cell cycle distribution was analyzed by FACS Calibur flow cytometer. Matrigel invasion assay and wound healing assay were done. The methods described by Chou and Talalay were used to determine whether a synergistic effect existed between drug combination.

Results: pSrc was highly expressed in most BTC cells. Bosutinib inhibited the growth of BTC cells, especially SNU308, SNU478 and HuCCT1 (IC50, 0.6-0.8 μmol/L). Bosutinib induced G1 arrest (decrease of cyclin D, cyclin E, cyclin A, cyclin B and increase of p27) in SNU308 and SNU478. It decreased pSrc, pFAK, pAKT and pERK in these cells. Bosutinib also inhibited the cell migration and invasion in parallel with decrease of snail, not only in SNU308, SNU478 cells but also in several other cells. Interestingly, bosutinib showed synergistic antiproliferative effects (Combination Index value <1) with gemcitabine in 4 out of 8 BTC cells. Bosutinib was downregulated TS (thymidine synthase) expression, one of resistance mechanisms to gemcitabine. Bosutinib also showed synergistic anti-invasion activity in combination of gemcitabine or cisplatin.

Conclusion: Taken together, these preclinical evaluations reveal that Src could be a potential therapeutic target in biliary tract cancer and support clinical development of bosutinib alone or in combination with chemotherapy in biliary tract cancer.

Citation Format: Ah-Rong Nam, Ahrum Min, Sae-Won Han, Sang-hyun Song, Do-Youn Oh, Seock-Ah Im, Tae-You Kim, Youg-Jue Bang. Evaluation of Src as a therapeutic target in biliary tract cancer using bosutinib, a c-Src/Abl kinase inhibitor. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 2081. doi:10.1158/1538-7445.AM2013-2081
Abstract 2098: Synergistic lethality of mifepristone and LY294002 in ovarian cancer cells.
Authors: Wempe Stacy L. , Gamarra-Luques Carlos D. , Telleria Carlos M.
Published: August 2014

We have previously shown that the antiprogestin/antiglucocorticoid mifepristone inhibits the growth of ovarian cancer cells. In this work, we hypothesized that cellular stress caused by mifepristone is limited to cytostasis and that cell killing is avoided because of a compensatory activation of the PI3K/Akt survival pathway.To investigate the role of this pathway in mifepristone-induced growth inhibition, human ovarian cancer cells of various histological subtypes and genetic backgrounds were exposed to cytostatic doses of mifepristone in the presence or absence of the PI3K inhibitor, LY294002. Mifepristone-induced growth arrest of ovarian cancer cells was associated with phosphorylation of Akt (Ser473), which was prevented by treatment with LY294002. The combination mifepristone/LY294002, but not the individual drugs, killed ovarian cancer cells via apoptosis, as attested by genomic DNA fragmentation and cleavage of caspase-3, and the concomitant downregulation of anti-apoptotic proteins Bcl-2 and XIAP. From a pharmacological standpoint, when assessing cell growth inhibition using a median-dose analysis algorithm, the interaction between mifepristone and LY294002 was synergistic. The lethality caused by the combination mifepristone/LY294004 in two dimensional cell cultures was recapitulated in organized, tri-dimensional spheroids. This study demonstrates that mifepristone and LY294002, when used individually cause cell growth arrest, yet when combined, they cause lethality. This work further provides proof-of-concept that in combination, targeted cytostatic therapies can be an efficient tool to engage cancer cells in a death program.

Citation Format: Stacy L. Wempe, Carlos D. Gamarra-Luques, Carlos M. Telleria. Synergistic lethality of mifepristone and LY294002 in ovarian cancer cells. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 2098. doi:10.1158/1538-7445.AM2013-2098