bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 5007: A novel polyisoprenylated cysteinyl amide inhibitor, NSL-BA-055, selectively inhibits proliferation of hepatocellular carcinoma cells
Authors: Naidoo Michelle K. , Lamango Nazarius , Ogunwobi Olorunseun O.
Published: August 2015

The monomeric GTP-ase, Ras, is mutated in over 30% of cancers where it promotes uncontrolled proliferation of cancer cells. Ras signaling is one of many pathophysiological pathways implicated in hepatocellular carcinoma that results in cell survival, differentiation, proliferation, and angiogenesis. Ras is bound to the cytoplasmic face of the plasma membrane where it relays intracellular signals. Polyisoprenylated cysteinyl amide inhibitors (PCAIs) effectively disrupt the functions of polyisoprenylated proteins such as Ras. Many hepatocellular carcinomas (HCCs) have dysregulated Ras signaling. However, the effects of PCAIs on hepatocellular carcinoma (HCC) are unknown. The aim of this study was to investigate the effect of a novel PCAI, NSL-BA-055, on HCC. Concentrations of NSL-BA-055 ranging from 0 to 10 μM were used for experimentation. Effects on proliferation of BNL CL.2 (non-tumorigenic mouse hepatocytes) and BNL 1ME A.7R.1 (mouse HCC cells derived from BNL CL.2 hepatocytes) were analyzed. Next, the effects of NSL-BA-055 on Akt protein expression, a downstream component of Ras signaling, were examined by western blotting. NSL-BA-055 significantly inhibited the proliferation of BNL 1ME A. 7R.1 HCC cells, but not the non-tumorigenic BNL CL.2 hepatocytes. Also, concentrations of the NSL-BA-055 compound as low as 4 μM significantly inhibited the proliferation of BNL 1ME A.7R.1 HCC cells. Western blotting revealed that NSL-BA-055 inhibited Akt protein expression at 4μM. Therefore PCAIs such as NSL-BA-055 is a novel class of agents that selectively inhibit the proliferation of HCC cells, but not normal hepatocytes. This effect may be via inhibition of Akt, a downstream intermediate of Ras signaling. Consequently, NSL-BA-055 is a novel candidate agent for HCC therapy.

Note: This abstract was not presented at the meeting.

Citation Format: Michelle K. Naidoo, Nazarius Lamango, Olorunseun O. Ogunwobi. A novel polyisoprenylated cysteinyl amide inhibitor, NSL-BA-055, selectively inhibits proliferation of hepatocellular carcinoma 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 5007. doi:10.1158/1538-7445.AM2015-5007
Abstract 5018: HLA class I expression in a tumor is higher than that out of a tumor: Promising new findings for antigen-specific cancer immunotherapy
Authors: Nobuoka Daisule , Takahashi Mari , Yoshikawa Toshiaki , Yagi Takahito , Fujiwara Toshiyoshi , Nakatsura Tetsuya
Published: August 2015

Background: We have demonstrated that intratumoral peptide injection is an attractive strategy that enhances tumor cell antigenicity by means of additional peptide loading onto HLA class I molecules (Nobuoka D, et al. Cancer Immunol Immunother, 62: 639-652, 2013). However, HLA class I expression has been reported to be down-regulated in various malignancies, which may explain why most studies on antigen-specific cancer immunotherapy did not demonstrate a remarkable benefit.

Methods: The tissue samples of histologically confirmed hepatocellular carcinoma were obtained from 20 patients who underwent hepatectomy. The paraffin-embedded blocks of tumor area and nontumorous liver were analyzed using monoclonal anti-HLA class I antibody, EMR 8-5. The fresh tissue samples were analyzed using another monoclonal anti-HLA class I antibody, W6/32. Moreover, HLA class I expression in a tumor and that out of a tumor were compared in various types of cancer.

Results: Among 20 patients with hepatocellular carcinoma, 14 (70%) were graded as having high expression (++) of HLA class I molecules using EMR 8-5 in cancer cells, whereas 15 patients (75%) were graded as having negative expression (-) in nontumorous hepatocytes. The immunoreactivity level in cancer cells was significantly higher than that in nontumorous hepatocytes (P < 0.05). Moreover, similar observations were obtained using W6/32. In other types of cancer including intrahepatic cholangiocarcinoma, ampullary carcinoma, gallbladder carcinoma, pancreatic carcinoma, esophageal carcinoma, colon carcinoma, and breast carcinoma, most cancer cells has higher expression of HLA class I than surrounding normal cells.

Conclusions: HLA class I expression is not so down-regulated as previously reported. Intratumoral peptide injection is applicable in various malignancies.

Citation Format: Daisule Nobuoka, Mari Takahashi, Toshiaki Yoshikawa, Takahito Yagi, Toshiyoshi Fujiwara, Tetsuya Nakatsura. HLA class I expression in a tumor is higher than that out of a tumor: Promising new findings for antigen-specific cancer immunotherapy. [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 5018. doi:10.1158/1538-7445.AM2015-5018
Abstract 5035: Effect of FXR agonist on colitis and colorectal carcinogenesis in mice
Authors: Miyazaki Tsuneyuki , Kochi Takahiro , Sakai Hiroyasu , Shirakami Yohei , Shimizu Masahito , Tanaka Takuji , Moriwaki Hisataka
Published: August 2015

Background & Aims: Farnesoid X receptor (FXR) is a member of the nuclear receptor superfamily and bile acids are endogenous ligand of FXR. FXR is highly expressed in the intestine and liver where FXR is essential for maintaining bile acid homeostasis. FXR activation has recently reported to inhibit intestinal inflammation and tumor development. The aim of this study was to investigate whether FXR agonist prevents colitis and colorectal carcinogenesis in mice.

Methods: We achieved two experiments. One experiment (Ex.1) used mouse colitis model with short term administration of FXR agonist, Nelumal A, and the other (Ex.2) employed mouse carcinogenesis model with long term administration. In Ex.1, male 5 week old A/J mice received drinking water containing 1.5% DSS for one week and were fed diets containing 100 or 400 ppm Nelumal A for 4 weeks. In Ex.2, male 5 week old A/J mice were administered once intraperitoneal injec¬]]tions of AOM (20 mg/kg body weight) and after one week, mice received drinking water containing 1.5% DSS for one week and were fed diets containing 100 or 400 ppm Nelumal A for 15 weeks.

Results: In Ex.1, DSS-treated mice all had colitis and multiple colonic mucosal ulcers. Both mucosal ulcer and inflammation grade in colon were significantly reduced in 400 ppm Nelumal A-treated mice, and also tended to be reduced in 100 ppm Nelumal A-treated mice. Body weight loss and colon length had no significant difference. The results of Ex.2 showed that the number of colonic mucosal ulcer, adenoma, and adenocarcinoma and total tumor number were decreased in 400 ppm Nelumal A-treated mice, but there was no statistically significant difference. The expression levels of FXR mRNA in colonic mucosa were increased by Nelumal A treatment. Moreover, administration of Nelumal A significantly decreased the expression levels of tumor necrosis factor α, monocyte chemoattractant protein 1, and proliferating cell nuclear antigen, and increased the level of Bax in colonic epithelium.

Conclusion: FXR agonist prevents chemically induced intestinal inflammation and carcinogenesis by inhibition of proinflammatory cytokine production and cell proliferation and promotion of apotosis in colonic mucosa.

Citation Format: Tsuneyuki Miyazaki, Takahiro Kochi, Hiroyasu Sakai, Yohei Shirakami, Masahito Shimizu, Takuji Tanaka, Hisataka Moriwaki. Effect of FXR agonist on colitis and colorectal carcinogenesis in mice. [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 5035. doi:10.1158/1538-7445.AM2015-5035
Abstract 5087: Precise characterization of macrophage secretory exosomes can lead to novel therapeutic approaches
Authors: Bednarczyk Robert Bronislaw , Kitadai Yuki , Tuli Neha Y. , Hanly Elyse K. , Benrahoma Ghada , Mittelman Abraham , Tiwari Raj K.
Published: August 2015

Multiple cell types interact within the tumor microenvironment (TME) through crosstalk events that are mediated by many biomolecules including cytokines, mRNA, and miRNA-containing exosomes that can influence cancer progression as well as suppression. Inflammation within the TME is a major determinant in tumor progression. Since tumor associated macrophages (TAMs) are a major cellular factor in tumor inflammation, we hypothesized that crosstalk events initiated by macrophage secreted factors such as cytokines and exosomes will modulate breast cancer phenotype. We evaluated the cytokine profile of a human monocytic cell line, THP-1, following stimulation with a phorbol ester TPA (12-O-Tetradecanoylphorbol-13-acetate) in vitro. Activated THP-1 cells exhibited an M1 macrophage phenotype. In addition, we assessed the morphological and cellular growth changes of luminal (T47D, MCF-7) and basal-like (MDA-MB 231) breast cancer cell lines in vitro following treatment with activated THP-1 conditioned medium (CM). Each cell line displayed mesenchymal-like features and decreased cellular growth following activated THP-1 CM treatment. MCF-7 and MDA-MB 231 displayed increased MEK/ERK signaling, while all three cell lines showed increased p-AKT expression. Furthermore, epithelial-like T47D and MCF-7 cells exhibited an increase in EMT transcription factor expression. Moreover, although T47D and MCF-7 did not show drastic changes in E-cadherin expression via western blot analysis, immunofluorescence examination showed a decrease in E-cadherin following activated THP-1 CM treatments emphasizing a possible partial EMT event. Lastly, activated THP-1 exosomes were isolated and used to treat MDA-MB 231 to determine the effects on breast cancer cell proliferation and senescence. MDA-MB 231 exhibited significantly decreased cellular growth and increased senescence following incubation with activated THP-1 exosomes. The variety of secretory factors within the TME as a result of inflammation may contribute to tumor suppression as well as tumor progression via a secretion of cytokines and exosomes.

Citation Format: Robert Bronislaw Bednarczyk, Yuki Kitadai, Neha Y. Tuli, Elyse K. Hanly, Ghada Benrahoma, Abraham Mittelman, Raj K. Tiwari. Precise characterization of macrophage secretory exosomes can lead to novel therapeutic approaches. [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 5087. doi:10.1158/1538-7445.AM2015-5087
Abstract 5091: Inhibiting endothelium directed tumor cell streaming by targeting the HGF/C-Met and EGF/CSF-1 signaling pathways
Authors: Leung Edison , Wang Yarong , Smith Bryan , Flynn Daniel , Condeelis John
Published: August 2015

In vivo, tumor cells need to migrate towards blood vessels in order to metastasize. Streaming, a specialized mode of migration involving chemotactic signaling between macrophages and tumor cells, allows linear streams of tumor cells to migrate directionally towards blood vessels on collagen I-containing ECM fibers in vivo. We have successfully reconstructed tumor cell streaming in vitro by co-plating tumors cells, macrophages, and endothelial cells on 2.5 um micro-patterned substrates. We found that tumor cells and macrophages do not demonstrate directional migration in only one direction but random walk. Directional streaming migration of tumor cells in only one direction was established when beads coated with human umbilical vein endothelial cells (HUVEC) were placed at one end of the micro-patterned “collagen fibers” within the assay. Using this in vitro reconstituted streaming system, we found that directional streaming can be blocked by inhibiting multiple signaling pathways - notably the HGF/C-Met signaling pathway between endothelial cells and tumor cells and the EGF/CSF-1 paracrine loop between macrophages and tumor cells. These pathways were blocked using gefitinib (EGFRi), DCC-3014 (CSFRi), and altiratinib (DCC-2701)(c-Meti), which led to a marked decrease in endothelium directed streaming of the tumor cells. The inhibitors showed different efficacies of inhibition of streaming at different distances from the endothelium indicating that HGF from the endothelium organized the random walk of paired macrophages and tumor cells into a directional migration. Key observations made with the in vitro reconstituted system were reproduced in mammary tumors using the in vivo invasion assay. These results highlight a promising role for combinations of these inhibitors in blocking tumor cell streaming and metastasis in vivo and for use in human trials.

Citation Format: Edison Leung, Yarong Wang, Bryan Smith, Daniel Flynn, John Condeelis. Inhibiting endothelium directed tumor cell streaming by targeting the HGF/C-Met and EGF/CSF-1 signaling pathways. [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 5091. doi:10.1158/1538-7445.AM2015-5091
Abstract 5104: Isorhapontigenin (ISO) inhibits cancer invasion by enhancing FOXO1 transcription through targeting STAT1 phosphorylation at Tyr701
Authors: Xue Lei , Jiang Guosong , Li Jingxia , Huang Chuanshu
Published: August 2015

Muscle invasion of bladder cancer can progress to life threatening metastases. Isorhapontigenin (ISO) is a novel derivative of stilbene that isolated from a Chinese herb Gnetum cleistostachyum, which has been used for centuries as treatment for several cancers including bladder cancer. However, nothing is known whether ISO possesses an inhibitory effect on invasion in human cancers. Here, we found that 10 to 20 mmol/L of ISO, which were relevant to in vivo study, inhibited human bladder cancer cell invasion accompanied by up-regulation of the forkhead box class O1 (FOXO1) mRNA transcription. Accordingly, FOXO1 was significantly down-regulated in invasive human bladder cancer tissues, and forced expression of FOXO1 specifically suppressed invasion of human bladder cancer cells. Moreover, knockdown of FOXO1 expression abolished the ISO inhibition of invasion. Further studies showed that increased FOXO1 transcription was accompanied by reduction of STAT1 phosphorylation at Tyr701, and repressing STAT1 phosophorylation by the overexpression of dominant negative STAT1 could mimic the effect of ISO treatment on FOXO1 transcription, while the mutation of STAT1's binding site in the FOXO1 promoter profoundly impaired ISO-induced up-regulation of FOXO1 transcription. Furthermore, this study revealed that metalloproteinase-2 (MMP-2) was a FOXO1 downstream effector mediating its effect on bladder cancer invasion. These findings not only provide a novel insight into the understanding of underlying mechanism of bladder cancer's propensity to invasion, but also identify a new role and mechanisms underlying the natural compound ISO that specifically suppresses such invasion through targeting the STAT1-FOXO1-MMP2 axis.

Citation Format: Lei Xue, Guosong Jiang, Jingxia Li, Chuanshu Huang. Isorhapontigenin (ISO) inhibits cancer invasion by enhancing FOXO1 transcription through targeting STAT1 phosphorylation at Tyr701. [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 5104. doi:10.1158/1538-7445.AM2015-5104
Abstract 5106: Influence of chloride flux on cell motility in head and neck squamous cell carcinoma
Authors: Khan Nayel , Steehler Kevin , Kemp Carolyn , Kulkarni Sucheta , Davis Kara , Duvvuri Umamaheswar
Published: August 2015

Purpose: To assess the specific involvement of chloride flux through TMEM16A in the regulation of tumor cell motility.

Background: Head and neck squamous cell carcinoma (HNSCC) often features tumor metastases and is associated with decreased patient survival. A key feature of the metastatic process is the ability of the cancer cells to migrate. TMEM16A, a calcium-activated chloride channel (CaCC), has been shown to play a role in cell migration and tumor metastasis, most recently as a potential node between cell proliferation versus cell migration. However, the role of chloride flux in this phenomenon has yet to be elucidated.

Methods: HNSCC cell lines were engineered to stably overexpress or knock-down TMEM16A. Cell migration was assessed via wound healing and transwell migration assays in cell culture media with varying chloride concentrations and in the presence of a TMEM16A specific inhibitor. The impact of chloride manipulation on EMT markers (E-cadherin, vimentin, ZO-1) was assessed using immunoblotting.

Results: Data obtained by both cell migration assays showed that in conditions of physiologically normal chloride, OSC19 TMEM16A overexpressors migrated less when compared to OSC19 vector controls. However, in conditions of low chloride, the migration rates between both cell lines were similar. Furthermore, inhibition of chloride flux using chloride depleted media, or pharmacologic inhibition of TMEM16A lead to a decrease in cell motility. As expected, an increase in markers of epithelial phenotype (i.e., Zo-1) and decrease in markers of mesenchymal phenotype (i.e., alpha-smooth muscle actin) in cells incubated in low chloride media was observed.

Conclusion: This data further supports the evidence that increased TMEM16A expression is correlated with decreased cell motility and therefore potential for cell metastasis. In addition, we have shown a role of chloride flux in TMEM16A's ability to modulate cell motility with a correlated change in EMT.

Citation Format: Nayel Khan, Kevin Steehler, Carolyn Kemp, Sucheta Kulkarni, Kara Davis, Umamaheswar Duvvuri. Influence of chloride flux on cell motility in head and neck squamous 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 5106. doi:10.1158/1538-7445.AM2015-5106
Abstract 5112: In vivo molecular imaging of HIF-1α and septin 9 interaction by bimolecular fluorescence complementation
Authors: Golan Maya , Mabjeesh Nicola J.
Published: August 2015

Hypoxia is a common feature of many solid tumors that promotes tumor progression and leads to radiation and chemotherapy resistance. Hypoxia-inducible factor 1 (HIF-1), as one of the major mediators of the hypoxic response, has been shown to activate hypoxia-responsive genes, which are involved in multiple aspects of tumorigenesis and cancer progression.

Previously, we have described an interaction between septin 9 isoform 1 (SEPT9_i1) protein and HIF-1α, the oxygen regulated subunit of HIF-1. SEPT9_i1 is a member of the conserved family of septins, which are GTP-binding proteins that form cytoskeleton-like filaments essential for many functions in eukaryotic organisms including neoplasia. The interaction of SEPT9_i1 with HIF-1α increases protein stability and HIF-1 transcriptional activity in vitro and promotes proliferation, tumor growth and angiogenesis in vivo. Recent data indicated that SEPT9_i1 was also involved in the trafficking of cytoplasmic HIF-1α into the nucleus by direct interaction to the nuclear transporter importin-α. Herein, we utilized split YFP bimolecular fluorescence complementation (BiFC) methodology in order to monitor HIF-1α/SEPT9_i1 interactions in vivo. First, we generated split YFP protein chimeras fused to HIF-1α and to SEPT9_i1 on both their amino and carboxyl termini. Different pairs of HIF-1α and SEPT9_i1 chimeras were tested to functional complementation of the split YFP fragments using transient transfection in HEK293 cells. The complemented proteins capable of fluorescence were identified as SEPT9_i1-YN (the amino terminus of YFP fused to the carboxyl terminal of SEPT9_i1) and YC-HIF-1α (the carboxyl terminus of YFP fused to the N-terminal of HIF-1α). This pair was selected for further studies in PC-3 human prostate cancer cells. YFP complementation fluorescence derived from these chimeras was increased in the presence of hypoxia mimicking agents, such as CoCl2 and the iron chelators deferoxamine (DFO) and dibenzoylmethane (DBM). The chimera, YCΔHLH lacking the HLH domain that is essential for the interaction with SEPT9_i1 gave a significant reduction in YFP complementation fluorescence. Furthermore, expression of SEPT9_i1 252-379aa fragment, the domain essential for the interaction with HIF-1α, abolished almost completely the YFP complementation fluorescence. These results reconfirm our previous studies on HIF-1α/SEPT9_i1 interactions and make this system attractive for identifying new compounds capable of disrupting this complex in order to inhibit tumor growth and angiogenesis.

Citation Format: Maya Golan, Nicola J. Mabjeesh. In vivo molecular imaging of HIF-1α and septin 9 interaction by bimolecular fluorescence complementation. [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 5112. doi:10.1158/1538-7445.AM2015-5112
Abstract 517: The role of MYST4 in ovarian carcinoma
Authors: Mao Tsui Lien , Lin Hsuan-Ping , Sheu Jim
Published: August 2015

Histone acetyltransferases (HATs) play critical roles in the process of cell growth and development. However, little is known about the relationship between MYST4, a known HAT, and human tumors. In our previous studies, we found that MYST4 was overexpressed in ovarian high-grade serous carcinomas and patients with tumors overexpressing MYST4 had significantly worse survival. To explore the role of MYST4 in ovarian carcinoma, we built up stable ovarian cancer cell lines with MYST4 knockdown by shRNA. By cell proliferation assay and flow cytometry, we found that MYST4 enhanced cancer cell growth and regulated cell cycle progression. In addition, the migration distance was significantly decreased in MYST4 knockdown cell lines, suggesting that MYST4 promotes cellular migration. Further cDNA microarray revealed several target genes that were down-regulated by MYST4 knockdown, some have been reported to be involved in tumorigenesis and aggressive tumor behavior. Together, our study suggests that MYST4 promotes cancer cell growth and migration. Overexpression of MYST4 in cancer cells may contribute to tumors toward a more aggressive behavior. Thus, MYST4 may serve as a potential target in treating ovarian carcinomas overexpressing MYST4.

Citation Format: Tsui Lien Mao, Hsuan-Ping Lin, Jim Sheu. The role of MYST4 in ovarian 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 517. doi:10.1158/1538-7445.AM2015-517
Abstract 523: Decrease of ARNT promotes cancer metastasis by activating the fibronectin/integrin β1/FAK axis
Authors: Huang Chi-Ruei , Lee Chung-Ta , Chang Kwang-Yu , Chang Wen-Chang , Chen Ben-Kuen
Published: August 2015

The aryl hydrocarbon receptor nuclear translocator (ARNT) is widely involved in regulating the tumorigenesis process by inducing genes that are involved in tumor growth and angiogenesis. Tumorigenesis usually involves normoxic conditions. However, the role of ARNT in tumor metastasis during normoxia remains unclear. Here, we demonstrated that ARNT protein levels were decreased in late stage human colorectal cancer using an immunohistochemical analysis. Stably silence of the ARNT protein promoted cancer cell migration and invasion, which was mediated by the activation of the fibronectin/integrin β1/FAK signaling axis. In addition, ARNT knockdown-induced migration and invasion was inhibited when ARNT was restored in cells. In a xenograft analysis of severe combined immunodeficiency mice, ARNT-knockdown inhibited tumor growth. However, when ARNT expression recovered, the tumor growth of ARNT-knockdown-induced metastatic lung colonies was significantly enhanced. Interestingly, chemotherapeutic drugs inhibited ARNT expression and promoted invasion of residual tumor cells. These results suggested that ARNT may play a positive role during tumor growth (either in early stage tumor growth or in metastatic organs), but plays a negative role in tumor migration and invasion. Therefore, the efficiency of ARNT-targeted therapy in different cancer stages should be carefully evaluated.

Citation Format: Chi-Ruei Huang, Chung-Ta Lee, Kwang-Yu Chang, Wen-Chang Chang, Ben-Kuen Chen. Decrease of ARNT promotes cancer metastasis by activating the fibronectin/integrin β1/FAK axis. [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 523. doi:10.1158/1538-7445.AM2015-523
Abstract 5241: Clinical relevance of circulating tumor DNA in plasma from pancreatic cancer patients
Authors: Tjensvoll Kjersti , Lapin Morten , Buhl Tove , Oltedal Satu , Berry Katrine Steen-Ottosen , Gilje Bjørnar , Søreide Jon Arne , Javle Millind , Nordgård Oddmund , Smaaland Rune
Published: August 2015

Background: Circulating tumor DNA (ctDNA) may originate from necrotic or apoptotic tumor cells in the primary tumor, metastatic lesions or in the circulation. There is evidence that the ctDNA level may reflect the total tumor burden in a patient. We wanted to investigate whether changes in the ctDNA level might be used to monitor disease progression in pancreatic cancer patients with locally or advanced disease. Methods: Blood samples (9 mL in EDTA tubes) were collected from 15 prospectively recruited patients with locally advanced and/or metastatic pancreatic cancer before initiation of treatment, and subsequently every month during chemotherapy. The samples were processed by LymphoprepTM (Axis Shield) density centrifugation before plasma DNA isolation using the QIAamp Circulating Nucleic Acid kit (Qiagen). A high-fidelity polymerase-based PNA clamp PCR method (Gilje et al., 2008, Oltedal et al., 2010) was then used for the detection of KRAS mutations in exon 12 and 13, as a surrogate marker for ctDNA. KRAS mutations have previously been reported to be present in >90% of the pancreatic cancers. Plasma samples from 29 healthy individuals were also analysed as a reference group for the PNA clamp PCR method. The results were compared with conventional biochemical and radiological monitoring measures. Results: The majority of the patients (80%) had metastatic disease and they were treated either by gemcitabine or FOLFORINOX. Nine (60%) patients had a positive KRAS status in the plasma samples obtained before initiation of chemotherapy, indicating presence of ctDNA. Moreover, 11 of the 15 included patients had ≥2 follow-up samples, and in several of these patients the ctDNA level changed substantially during the course of chemotherapy. In total, 17/38 (44.7%) patient samples were positive for plasma DNA KRAS mutations during chemotherapy. Changes in the ctDNA level seemed to correspond both to radiological follow-up data and changes in CA19-9 for several patients. Analyses of the total plasma DNA fraction with regard to disease monitoring, gave more inconclusive results in this small pilot study. Conclusion: Our pilot study gives support to the hypothesis that ctDNA may be used as a marker for monitoring treatment efficacy and disease progression in pancreatic cancer patients. This will be further investigated.

Citation Format: Kjersti Tjensvoll, Morten Lapin, Tove Buhl, Satu Oltedal, Katrine Steen-Ottosen Berry, Bjørnar Gilje, Jon Arne Søreide, Millind Javle, Oddmund Nordgård, Rune Smaaland. Clinical relevance of circulating tumor DNA in plasma from pancreatic cancer patients. [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 5241. doi:10.1158/1538-7445.AM2015-5241
Abstract 5257: Immunohistochemical analysis and comprehensive hormone receptor profiling of uterine adenosarcomas at a single institution
Authors: Zechmeister Jenna R. , Huang Gloria S. , Karabakhtsian Rouzan , Schwartz Melissa , Prossnitz Eric R. , Goldberg Gary L. , Klobocista Merieme
Published: August 2015

Uterine adenosarcomas (AS) are rare uterine tumors with benign epithelium and a sarcomatous component which is typically low-grade. Those displaying sarcomatous overgrowth (AS+SO) behave in a more aggressive fashion and are associated with worse clinical outcomes. Immunohistochemistry performed on AS and AS+SO have shown a greater preponderance for estrogen receptor alpha (ERα) and progesterone receptor (PR) positivity in those with a low-grade sarcomatous component. Further studies showed an increased proliferative index (Ki-67) in AS+SO and confirmed loss of expression in ER/PR in addition to loss of CD10. Expression of estrogen receptor beta (ERβ), a classic estrogen receptor as well as G protein-coupled estrogen receptor (GPER), a receptor that interacts with estrogen independently of the classic receptors, has been reported in uterine carcinosarcomas and they are associated with advanced stage disease. We conducted this study to analyze the receptor status of ERβ and GPER in cases of uterine AS. An H-score was calculated as the product of intensity and the percentage of cells staining positive (0-100%), higher scores indicate increased intensity and a greater number of positive cells. Unpaired t-test was used to compare the mean H-scores of receptor expression between low-grade AS (LG-AS) and high-grade AS (HG-AS). Mean age of diagnosis was 59.4 ± 11.9 years, mean BMI of 34.2 ± 8.5, and median disease free survival was 18 months (range 2-113 months). There were 7 cases of LG-AS and 3 cases of HG-AS, 2 cases displayed SO. Seven cases were stage IA, two were IB and one stage IV. There was a significant difference (p = 0.01) between the mean H-score for ERα between LG-AS (203) and HG-AS (1.7), There was also a significant difference (p = 0.04) in mean H-score for PR between LG-AS (188.6) and HG-AS (18.3). ERβ and GPER expression were retained in the HG-AS at similar levels as in the LG-AS, thus there was no difference in mean H-scores. In one patient with HG-AS and recurrent disease after 2 years, the recurrent tumor remained negative for ERα and PR, and showed somewhat decreased ERβ expression. Although the sample size is small, this is the first report of comprehensive estrogen receptor profiling in this uncommon uterine tumor.

Citation Format: Jenna R. Zechmeister, Gloria S. Huang, Rouzan Karabakhtsian, Melissa Schwartz, Eric R. Prossnitz, Gary L. Goldberg, Merieme Klobocista. Immunohistochemical analysis and comprehensive hormone receptor profiling of uterine adenosarcomas at a single institution. [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 5257. doi:10.1158/1538-7445.AM2015-5257
Abstract 5258: Expression analysis of STEAP1 in breast cancer patients as therapeutic target
Authors: Mudalagiriyappa Chidananda , Pillai Sreeraj , Watson Mark , Aft Rebecca
Published: August 2015

Objective: The six transmembrane epithelial antigen of the prostate (STEAP1) is predominantly overexpressed in human prostate cancer. STEAP1 was first identified as a prostate-specific cell-surface antigen and found to be up-regulated in multiple cancer cell lines and in various cancers, including lung, bladder, colon and ovarian cancers. However no significant expression in normal tissue has been reported suggesting its potential use as a target for anti-STEAP1 immunotherapy. An anti-STEAP1 monoclonal antibody linked to an antimitotic agent is in Phase I clinical trials for prostate cancer patients. Microarray data from our lab suggested that STEAP1 is also highly expressed in human breast cancers. In this study we evaluate expression STEAP1 in primary tumors and the bone marrow from breast cancer patients as a therapeutic target for minimal residual disease.

Experimental procedures: RNA was isolated from primary tumor and non-malignant breast tissue. RNA was also isolated from bone marrow of state II and III breast cancer patients, healthy volunteers and breast cancer cell lines. STEAP1 expression was analyzed by nanostring nCounter and real time qRT-PCR using human specific probes. STEAP1 immunohistochemical (IHC) staining of human tissue was performed using standard IHC Protocol.

Results: STEAP1 mRNA was up-regulated in 77% of tumors (28/36) compared to the corresponding normal tissue. STEAP1 protein was expressed in 100% of tumors (8/8) and was absent in non-malignant breast tissue (7/7) by IHC staining. STEAP1 mRNA was not expressed normal BM, but was detected in 8% (6/74) of BM from patients with early stage breast cancer. Of patients with STEAP1 positive BM, 50% (3/6) had triple negative disease and 66% (4/6) developed recurrent disease (p = .028).

Conclusion: STEAP1 is expressed in human breast tumors and the BM of breast cancer patients. Expression of STEAP1 in the BM is significantly associated with the development of metastatic disease. Our data indicate that STEAP1 could serve as a therapeutic target for the treatment of minimal residual disease in breast cancer.

Citation Format: Chidananda Mudalagiriyappa, Sreeraj Pillai, Mark Watson, Rebecca Aft. Expression analysis of STEAP1 in breast cancer patients as therapeutic 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 5258. doi:10.1158/1538-7445.AM2015-5258
Abstract 5365: Alternating electric fields (TTFields) in combination with paclitaxel are therapeutically effective against ovarian cancer cells in vitro and in vivo
Authors: Munster Mijal , Roberts Christopher P. , Schmelz Eva M. , Giladi Moshe , Blat Roni , Schneiderman Rosa S. , Porat Yaara , Bomzon Zeev , Urman Noa , Itzhaki Aviran , Voloshin Sela Tali , Cahal Shay , Kirson Eilon D. , Weinberg Uri , Palti Yoram
Published: August 2015

Tumor Treating Fields (TTFields) are low intensity intermediate frequency alternating electric fields that disrupt mitosis. Our previous in vitro studies suggest that ovarian cancer cells are highly sensitive to TTFields treatment.

The goal of the present study was to evaluate the efficacy of the combined treatment of TTFields and paclitaxel against ovarian cancer cells in vitro and in vivo.

For in vitro studies, TTFields (1.75 V/cm) were applied for 72 hours using the inovitro system. The in vivo efficacy of the combined treatment was tested in C57Bl/6 mice, orthotopically injected with MOSE-L FFL luciferase positive cells. Finite Element Mesh (FEM) simulations were performed using the Sim4life software package (ZMT, Zurich, Switzerland) for the calculations of the electric fields intensities around the ovaries.

Our results demonstrate that in vitro application of 200 kHz TTFields led to a significant reduction in both the number of viable cells (44.6%) and the clonogenic potential (23.8%) as compared to untreated cells (p<0.001). Further reduction in the number of viable cells was achieved when TTFields were combined with paclitaxel. In vivo, the combined treatment of TTFields and Paclitaxel led to a significant reduction in tumor luminescence (40%, p<0.01) and in the tumor weight (55%, p<0.05) as compared to untreated tumor bearing mice. FEM simulations demonstrated that electric fields intensities inside and in the vicinity of the ovaries of a real human anatomy model are about 1 and 2 V/cm RMS which is above the minimal threshold required for TTFields response.

Taken together these results demonstrate that the combined treatment of TTFields and paclitaxel could serve as an effective treatment against ovarian cancer. A clinical trial testing the efficacy of the combined modalities is now underway.

Citation Format: Mijal Munster, Christopher P. Roberts, Eva M. Schmelz, Moshe Giladi, Roni Blat, Rosa S. Schneiderman, Yaara Porat, Zeev Bomzon, Noa Urman, Aviran Itzhaki, Tali Voloshin Sela, Shay Cahal, Eilon D. Kirson, Uri Weinberg, Yoram Palti. Alternating electric fields (TTFields) in combination with paclitaxel are therapeutically effective against ovarian cancer cells in vitro and in vivo. [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 5365. doi:10.1158/1538-7445.AM2015-5365
Abstract 5395: Anti-tumor activity of BMS-595, a novel CK2 kinase inhibitor
Authors: Rupnow Brent A. , Yu Chiang , Pabalan Jonathan G. , Roongta Urvashi V. , Lippy Jonathan S. , Dongre Ashok R. , Obermeier Mary T. , Fura Aberra , Elzinga Paul A. , Henley Benjamin J. , Fargnoli Joseph , Lee Francis Y. , Foster William R. , Tarby Christine M. , Fink Brian E. , Tokarski John S. , Gavai Ashvinikumar V. , Wong Tai W. , Hunt John T. , Vite Gregory D. , Purandare Ashok V.
Published: August 2015

The CK2 protein kinases are a small family of two highly related serine/threonine kinases composed of two catalytic subunits, α and α’, and a single β subunit. Numerous substrates have been reported for CK2 and these proteins are known to participate in diverse cellular processes, including cell signaling, transcription, DNA repair, apoptosis regulation and tumor suppression. Elevated CK2 expression and kinase activity has been observed in many cancer types. Further, mRNA knockdown and enzyme inhibition studies have demonstrated that many cancer cell lines are dependent on CK2 for growth and survival. To further evaluate CK2 kinases as targets for therapeutic intervention in cancer, we identified BMS-595, a potent and selective, ATP-competitive CK2 inhibitor. BMS-595 inhibits the in vitro proliferation of human colorectal and lung cancer cell lines with IC50s ranging from less than 10 nM to greater than 1 μM. In sensitive cell lines, anti-proliferative effects of BMS-595 and structurally related analogs strongly correlated with cellular CK2 kinase inhibition. Oral administration of BMS-595 to mice bearing colorectal cancer and lung cancer xenografts demonstrated pharmacodynamic effects and robust efficacy at tolerated doses. These studies confirm the dependence of a subset of human colon and lung cancer cell lines on CK2 activity for growth and demonstrate that pharmacologic inhibition of CK2 can produce anti-tumor efficacy at tolerated doses.

Citation Format: Brent A. Rupnow, Chiang Yu, Jonathan G. Pabalan, Urvashi V. Roongta, Jonathan S. Lippy, Ashok R. Dongre, Mary T. Obermeier, Aberra Fura, Paul A. Elzinga, Benjamin J. Henley, Joseph Fargnoli, Francis Y. Lee, William R. Foster, Christine M. Tarby, Brian E. Fink, John S. Tokarski, Ashvinikumar V. Gavai, Tai W. Wong, John T. Hunt, Gregory D. Vite, Ashok V. Purandare. Anti-tumor activity of BMS-595, a novel CK2 kinase inhibitor. [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 5395. doi:10.1158/1538-7445.AM2015-5395
Abstract 5436: Targeting oncoproteins via disruption of proteostasis: Identification of oncoprotein destabilizing agents using luciferase tagged oncoproteins
Authors: Middleton Richard E. , Olsen Greg , McSweeney Russell , Lu Gang , Gao Wenhua , Roberts Justin , McKeown Michael R. , Bittinger Mark A. , Wong Kwok-Kin , Bradner James E. , Kaelin William G.
Published: August 2015

Since many oncoproteins are considered undruggable targets, we have developed and validated a new screening strategy to identify small molecules that inhibit oncoprotein activity by causing their degradation. Recently, the mechanism of action for the class of drugs known as immunomodulators (IMiDs; e.g. thalidomide, lenalidomide and pomalidomide) has uncovered a novel oncogene targeting strategy. IMiDs commandeer an E3 ligase complex that contains the E3 recognition subunit cereblon and divert its activity toward two transcription factors IKZF1 and IKZF3, which play critical roles in hematopoietic cells. This mechanism explains the efficacy of IMiDs in multiple myeloma and may translate into a general approach to target traditionally undruggable oncoproteins. We generated cell lines stably expressing IKZF1as a firefly luciferase fusion protein with coexpression of Renilla luciferase. A pilot HTS campaign was run to uncover novel drugs that can mimic the effects of IMiDs. A cell line expressing the MYC-firefly fusion protein was used for counter-screening. We used pomalidomide as our positive control and it reduced firefly luciferase signal ≈75% with no change in the renilla luciferase signals. This control resulted in a Z prime value typically >0.5. The pilot screen of 3,850 compounds resulted in 39 “hits” that reduced firefly luciferase >35%. More importantly, both thalidomide and lenalidomide were found to be active in the screen. Confirmation testing revealed that 75% of “hits” repeated, but also had activity in the MYC-firefly cell line. Actives were tested in western blots for their ability to decrease fusion protein abundance. Many active compounds did not reduce protein levels in western blots and represent nonspecific hits (e.g. luciferase inhibitor). In conclusion, we present a screening paradigm with the potential to uncover novel proteostasis disruptors by expressing luciferase tagged oncoproteins.

Citation Format: Richard E. Middleton, Greg Olsen, Russell McSweeney, Gang Lu, Wenhua Gao, Justin Roberts, Michael R. McKeown, Mark A. Bittinger, Kwok-Kin Wong, James E. Bradner, William G. Kaelin. Targeting oncoproteins via disruption of proteostasis: Identification of oncoprotein destabilizing agents using luciferase tagged oncoproteins. [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 5436. doi:10.1158/1538-7445.AM2015-5436
Abstract 5445: Dual-color immunohistochemistry assays for measuring Aiolos and Ikaros proteins in multiple myeloma patient samples
Authors: Ren Yan , Wang Maria , Couto Suzana , Hansel Donna , Gandhi Anita K. , Hagner Patrick , Thakurta Anjan , Chopra Rajesh , Breider Mike
Published: August 2015

IMiDs® immunomodulatory drugs Lenalidomide (LEN) and Pomalidomide (POM) are antineoplastic agents that have a significant clinical impact in the treatment of patients with multiple myeloma (MM). Recent studies indicate that the anti-proliferative activity of LEN and POM in MM cells is associated with ubiquitination and degradation of the Ikaros family zinc finger protein transcription factors, Aiolos (IKZF3) and Ikaros (IKZF1), the substrates of the Cereblon (CRBN)-dependent Cullin 4 E3-ligase complex. Measurement of Aiolos and Ikaros protein levels in MM patient samples is critical for future correlative studies. Here we describe the development and validation of dual IHC Aiolos/CD138 and Ikaros/CD138 assays using the highly specific Aiolos rabbit monoclonal antibody CGN-9-9-7, and a commercial Ikaros rabbit polyclonal antibody H-100 in combination with a commercial CD138 antibody. Aiolos and Ikaros immunoreactivity in CD138+ MM cells is scored using the semi-quantitative H-score method. H-scores range from 0 to 300 and are the sum of the products of the percentage of tumor cells (0-100%) and the intensity of staining (0-3). The assays were specific and detected high Aiolos and Ikaros expression in the positive control OCI-Ly10 lymphoma cell line, and low Aiolos and Ikaros levels in POM treated OCI-Ly10 cells. These results were confirmed via Western Blot analysis of Aiolos and Ikaros expression data in these cells lines. Assay precision was examined via staining serial sections of the same MM tumor across three different days and deemed acceptable. The dual IHC assays were used to evaluate bone marrow core biopsies or aspirate clots from 22 MM patients and H-scores were determined by a board-certified pathologist. The dual IHC assays detected a wide range of Aiolos or Ikaros immunoreactivity in the 22 MM cases, with nuclear H-scores ranging from 10 to 260 for Aiolos and 40 to 240 for Ikaros. In conclusion, the dual Aiolos/CD138 and Ikaros/CD138 IHC assays were developed to provide a specific and reliable semi-quantitative method to evaluate Aiolos and Ikaros protein levels in both bone marrow core biopsies and clots from MM patients. These assays are currently being used in clinical trials to assess the importance of Aiolos and Ikaros as biomarkers in MM patients treated with IMiDs® immunomodulatory drugs.

Citation Format: Yan Ren, Maria Wang, Suzana Couto, Donna Hansel, Anita K. Gandhi, Patrick Hagner, Anjan Thakurta, Rajesh Chopra, Mike Breider. Dual-color immunohistochemistry assays for measuring Aiolos and Ikaros proteins in multiple myeloma patient samples. [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 5445. doi:10.1158/1538-7445.AM2015-5445