bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 3546: Arming a tumor targeted oncolytic herpes simplex sirus type 1 with matrix metalloproteinase 9 for enhanced vector distribution and killing activity
Authors: Li Aofei , Marzulli Marco , Zhang Mingdi , Goins William , Kaur Balveen , Bolyard Chelsea , Amankulor Nduka , Leronni Daniela , Sette Paola , Cohen Justus , Glorioso Joseph , Grandi Paola
Published: August 2015

Early phase human clinical trials using several versions of oncolytic herpes simplex virus type 1 (oHSV) have shown promise in the treatment of GBM, but efficacy has been limited. Impediments to oHSV therapy include poor virus spread due in part to the tumor extracellular matrix, and insufficient replication in tumor cells as a result of attenuating mutations. Thus, the central goal of this project is to improve oncolytic vector delivery, replication and spread while maintaining safety and tumor specificity. We have already developed a new class of two stage tumor targeted oHSV combining (i) selective infection through tumor-specific receptors and (ii) selective replication based on differential expression of microRNAs (miRs) in tumor and normal cells. We further modify our vector by arming with the matrix metalloproteinase 9 (MMP9) as a means to reduce vector trapping in the tumor extracellular matrix. Here we show that MMP9 expression (i) enhances Oncolytic vector spreading in GBM neruospheres in vitro and (ii) improves tumor killing in a xenogeneic model of primary human GBM with significant long-term survival (≥50%) comparable to the control.

Citation Format: Aofei Li, Marco Marzulli, Mingdi Zhang, William Goins, Balveen Kaur, Chelsea Bolyard, Nduka Amankulor, Daniela Leronni, Paola Sette, Justus Cohen, Joseph Glorioso, Paola Grandi. Arming a tumor targeted oncolytic herpes simplex sirus type 1 with matrix metalloproteinase 9 for enhanced vector distribution and killing activity. [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 3546. doi:10.1158/1538-7445.AM2015-3546
Abstract 3547: Systemically delivered human telomerase reverse transcriptase (hTERT)-targeting p53-laden adenovirus shows strong antitumor effects in intrahepatic Hep3B xenograft mouse model
Authors: Lee Won Sup , Kang Hye Lim , Nagappan Arulkumar , Yun Jeong Won , Kim GonSup , Hong Soon Chan , Lee Sang-Jin , Kim In-Hoo
Published: August 2015

Human telomerase reverse transcriptase (hTERT) RNA and p53 tumour suppressor were previously suggested as useful targets for cancer gene therapy. In this study, we investigated the effects of systemically delivered adenoviruses harboring trans-splicing ribozyme that could target hTERT and induce p53 transcript in targeted cancer cells in vivo using mice with Hep3B human hepatocellular carcinoma cells that metastasized to the liver. We observed that intravenous injection of this virus (0.5 X1011 Virus) produced efficient regression of Hep 3B tumor metastasized nodules. Next we performed in vitro study to know whether it has a synergistic effects with CDDP, we observed that CDDP (2.5 ug/ml) increased cytotoxic effects in vitro given with the virus (10 MOI), but in vivo study revealed that addition of CDDP or arsenic hexoxide (As4O6) did not improve the therapeutic efficacy of the virus treatment. Next we performed toxicity test with 4 doses (0.5 X1011, 1 X1011, 2 X1011, and 5 X1011, Virus), which revealed that there were no toxic death, but 2-4 fold increased in AST and ALT with no changes in bilirubin, BUN, Creatinine levels. In conclusion, this study demonstrates that the adenoviruses harboring trans-splicing ribozyme targeting hTERT and inducing p53 transcript represents a safe and promising cancer treatment for hepatocellular carcinoma. [This study was supported by a grant from the National R&D Program for Cancer Control, Ministry for Health, Welfare & Family Affairs, Republic of Korea. (0820050), and National Research Foundation of Korea (NRF) grant funded by the Korea government(MEST) (20120002631).]

Keywords: Adenovirus, trans-splicing ribozyme, p53 tumour suppressor, Human Telomerase Reverse Transcriptase, cancer

Citation Format: Won Sup Lee, Hye Lim Kang, Arulkumar Nagappan, Jeong Won Yun, GonSup Kim, Soon Chan Hong, Sang-Jin Lee, In-Hoo Kim. Systemically delivered human telomerase reverse transcriptase (hTERT)-targeting p53-laden adenovirus shows strong antitumor effects in intrahepatic Hep3B xenograft mouse model. [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 3547. doi:10.1158/1538-7445.AM2015-3547
Abstract 355: A monoclonal antibody against laminin alpha 4 inhibits both MCAM and integrin-a6b1 binding, blocks tumor cell adhesion in vitro and slows melanoma tumor growth in vivo
Authors: Tam Stephen J. , Li Lauri E. , Lorenzana Carlos , Liu Yue , Flanagan Ken , Dolan Philip J. , Alexander Lana , Salmans Josh , Barbour Robin M. , Higaki Jeffrey N. , Nijjar Tarlochan , Zago Wagner , Yednock Ted A. , Kinney Gene
Published: August 2015

Laminin is an extracellular matrix molecule that is secreted as a trimeric protein composed of an α, β and γ chain. There are five distinct laminin α chains that impart distinct binding and signaling activities for cell adhesion, migration, and differentiation in a tissue-specific fashion. Laminin a4 (LAMA4) is expressed by vascular endothelial cells throughout the body, but is also overexpressed by cells within tumors of different origin compared to healthy human tissue. LAMA4 serves as a ligand for the cell adhesion receptors MCAM and a6b1 integrin, as well as for cell surface heparin sulfate proteoglycans. In order to probe how the MCAM/Laminin interaction contributes to tumor growth, we screened for LAMA4-specific monoclonal antibodies that block MCAM binding. We identified several such antibodies, and found that a subset also inhibits LAMA4 binding to a6b1 integrin, indicating that MCAM and a6b1 integrin recognize a similar region of LAMA4. Examining cells from multiple types of tumors, we found that these antibodies were potent inhibitors of tumor cell adhesion to LAMA4 in vitro. Finally, with in vivo administration, we demonstrated dose-dependent inhibition of human melanoma tumor growth in a mouse xenograft model, accompanied by morphologic changes in LAMA4 tumor distribution. These findings suggest that the MCAM/a6b1 integrin binding activity of LAMA4 contributes to both tumor adhesion and growth and that LAMA4 may provide a therapeutic drug target for several different oncology indications.

Citation Format: Stephen J. Tam, Lauri E. Li, Carlos Lorenzana, Yue Liu, Ken Flanagan, Philip J. Dolan, Lana Alexander, Josh Salmans, Robin M. Barbour, Jeffrey N. Higaki, Tarlochan Nijjar, Wagner Zago, Ted A. Yednock, Gene Kinney. A monoclonal antibody against laminin alpha 4 inhibits both MCAM and integrin-a6b1 binding, blocks tumor cell adhesion in vitro and slows melanoma tumor growth 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 355. doi:10.1158/1538-7445.AM2015-355
Abstract 356: Mutaome-based magnolin sensitivity in ovarian cancer cells
Authors: Song Ji-Hong , Kim Woo-Young , Cho Yong-Yeon
Published: August 2015

Signaling pathways mediated through Ras/Raf/MEK/ERK and/or PI3K/AKT/mTOR signaling pathways play a key role in cell transformation and cancer development. During carcinogenesis, genomic alterations, accumulation of mutations and amplification of growth signaling, induce cellular phenotype changes including escaping aging, metabolic change of energy source and resistance against therapeutic agents. Recently, our research group has identified a natural compound, magnolin, that inhibits ERK1 and ERK2 kinase activities with about 87 uM and 16.5 uM of IC50, respectively. However, the effect of magnolin in cancer cells has been poorly understood. To explore the effect of magnolin in cancer cells, we examined anti-proliferation effect of magnolin in pancreatic, colon and ovarian cancer cells. We found that striking differences of cancer cell proliferation was observed in ovarian cancer cells, TOV112D and SKOV3, by magnolin treatment. In TCGA mutaome data-base analysis, we found that TOV112D and SKOV3 harbor the different mutations in different genes which are known to involve in chemo-resistance. In cell cycle analysis, we found that TOV112D reduced S-phase of cell cycle and increased G1/G0-phase of cell cycle. Importantly, activation protein profiles in TOV112D and SKOV3 cells by Western blotting showed that phosphorylation of p90RSK including RSK2 was higher in SKOV3 cells compared with TOV112D cells. These results strongly suggested that alteration of p90RSK signaling may play an important role in expression of chemo-resistance in ovarian cancer cells.

Citation Format: Ji-Hong Song, Woo-Young Kim, Yong-Yeon Cho. Mutaome-based magnolin sensitivity in ovarian cancer cells. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 356. doi:10.1158/1538-7445.AM2015-356
Abstract 3569: Inhibition of the TRPM7 kinase domain inhibits breast cancer cell migration and invasion and tumor metastasis
Authors: Kaoud Tamer S. , Xie Xuemei , Park Jihyun , Tavares Clint D.J. , Mitra Shreya , Cano Micael , Sammons Rachel M. , Radwan Mohamed F. , Bartholomeusz Chandra , Dalby Kevin N.
Published: August 2015

TRPM7 (transient receptor potential melastatin 7) is a non-selective cation channel fused to protein kinase domain at the C-terminal whose activity is linked to the control of actomyosin contractility. TRPM7 mediates adhesion and migration of breast cancer cells and promotes breast tumor metastasis. The lack of cell-permeable inhibitors of the kinase domain represents a barrier to understand the kinase function. Herein, we discovered the first small molecule (KD-1) that targets TRPM7 kinase activity. Mg2+ starvation, which promotes TRPM7 kinase activity, induces phosphorylation of eEF2. Treatment of Mg2+-starved HEK293 cells with KD-1 decreased eEF2 phosphorylation, consistent with TRPM7 kinase activity suppression in-cells. KD-1 decreased the binding of Myosin IIB to TRPM7 in HEK293 and MDA-MB-231 cells. And when MDA-MB-231 cells were treated with increasing doses of KD-1, no change in cell viability was seen. Interestingly, KD-1 inhibited MDA-MB-231 cells migration and invasion that is reportedly regulated by TRPM7 kinase activity. Finally, the bioluminescent signals (to assess metastasis) were significantly lower in KD-1-treated mice (25 and 50 mg/kg/day) than in mice treated with vehicle control (P ≤ 0.05, 2-sided t-test.). in conclusion, inhibition of TRPM7 kinase activity may reduce or block breast tumor progression and/or metastasis.

Citation Format: Tamer S. Kaoud, Xuemei Xie, Jihyun Park, Clint D.J. Tavares, Shreya Mitra, Micael Cano, Rachel M. Sammons, Mohamed F. Radwan, Chandra Bartholomeusz, Kevin N. Dalby. Inhibition of the TRPM7 kinase domain inhibits breast cancer cell migration and invasion and tumor metastasis. [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 3569. doi:10.1158/1538-7445.AM2015-3569
Abstract 3571: Furin activity: A driver of rhabdomyosarcoma progression
Authors: Jaaks Patricia AIM , Meier Gianmarco , Schäfer Beat W. , Bernasconi Michele
Published: August 2015

Rhabdomyosarcoma (RMS) is the most common soft tissue sarcoma in children. The prognosis and survival rate is often very poor and therefore new therapy approaches are required. Thus, we focus on 1) employing RMS-specific peptides for targeted delivery of chemotherapeutics to the tumor site and 2) identifying novel therapeutic targets. In a previous study we discovered that RMS-homing peptides bind preferably to the proprotein convertase furin. Initial investigations confirmed a high expression of furin throughout different pediatric soft tissue sarcoma types and hinted that furin promotes the tumorigenic phenotype of RMS cells in vitro as well as in corresponding xenografts in vivo. Here we present a novel approach of tetracycline-induced shRNA-based silencing of furin in vitro and in vivo in order to investigate in depth the role of furin in RMS progression. Furin depletion was confirmed at mRNA, protein and activity level and led to impaired maturation furin substrates insulin like growth factor 1 receptor (IGF1R) and transforming growth factor beta 1 (TGFβ-1). We found that loss of furin activity suppresses the malignant phenotype of Rh30 cells by decreasing proliferation and affecting formation of colonies. Furthermore, we observed increased caspase 3/7 activity and enrichment of nucleosomes in the cytoplasm upon furin depletion, thus hinting initiation of apoptotic processes. Induction of furin silencing in RMS xenografts in NOD/SCID mice delayed tumour growth, indicating a crucial role of furin in early phases of tumour growth. Taken together, our data underscore the importance of furin for RMS progression and therefore targeting the activity of furin represents a promising tool for treatment of RMS.

Citation Format: Patricia AIM Jaaks, Gianmarco Meier, Beat W. Schäfer, Michele Bernasconi. Furin activity: A driver of rhabdomyosarcoma progression. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 3571. doi:10.1158/1538-7445.AM2015-3571
Abstract 3636: Targeting the MLL complex in castration resistant prostate cancer
Authors: Malik Rohit , Khan Amjad P. , Asangani Irfan A. , Cieślik Marcin , Prensner John R. , Wang Xiaoju , Iyer Matthew K. , Jiang Xia , Borkin Dmitry , Escara-Wilke June , Stender Rachell , Wu Yi-Mi , Cao Xuhong , Feng Felix Y. , Grembecka Jolanta , Cierpicki Tomasz , Chinnaiyan Arul M.
Published: August 2015

Resistance to androgen deprivation therapies and increased androgen receptor (AR) activity are major drivers of castration resistant prostate cancer (CRPC). Substantial prior work has focused on targeting AR directly; however, the identification and therapeutic targeting of co-activators of AR signaling remains an underexplored area of potential clinical significance. Here we demonstrate that the MLL (mixed-lineage leukemia) complex, a well-known contributor in MLL-fusion-positive leukemia, acts as a co-activator of AR signaling. AR directly interacts with the MLL complex via its critical subunit, menin. Small molecule inhibition of the menin-MLL interaction blocks AR signaling and inhibits the growth of castration resistant tumors in vivo. Furthermore, we find that menin is up-regulated in castration resistant prostate cancer and high expression correlates with poor overall survival. Taken together, our study identifies the MLL complex as a critical co-activator of AR that can be targeted in advanced prostate cancer.

Citation Format: Rohit Malik, Amjad P. Khan, Irfan A. Asangani, Marcin Cieślik, John R. Prensner, Xiaoju Wang, Matthew K. Iyer, Xia Jiang, Dmitry Borkin, June Escara-Wilke, Rachell Stender, Yi-Mi Wu, Xuhong Cao, Felix Y. Feng, Jolanta Grembecka, Tomasz Cierpicki, Arul M. Chinnaiyan. Targeting the MLL complex in castration resistant prostate cancer. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 3636. doi:10.1158/1538-7445.AM2015-3636
Abstract 365: Mobilizing prostate cancer cells from the endosteal niche by targeting the SDF-1/CXCR4 axis
Authors: Valkenburg Kenneth C. , Pienta Kenneth J.
Published: August 2015

Metastatic prostate cancer cells preferentially home to bone, and skeletal metastases are present in approximately 90% of patients upon autopsy. Our lab was the first to show that prostate cancer cells compete with hematopoietic stem cells (HSCs) for space within the endosteal niche. The HSC niche is located on the surface of endosteal osteoblasts, where HSCs are kept in a slow-cycling, protective state. The niche also provides prostate cancer cells with a chemoprotective environment in which they can remain dormant for an extended period of time, even decades. C-X-C chemokine receptor 4 (CXCR4) is required for HSC recruitment to the HSC niche. Osteoblasts secrete stromal cell derived factor 1 (SDF-1, also known as CXCL12), the ligand for CXCR4, which attracts HSCs to the niche via a chemotactic gradient. CXCR4 is also expressed by disseminated prostate cancer cells and is required for prostate cancer cells homing to bone. HSCs can be mobilized from the niche into the bloodstream physiologically via infection or stress and pharmacologically via proteins like granulocyte-colony stimulating factor (G-CSF) or small molecule inhibitors like AMD3100 (plerixafor). AMD3100, which antagonizes CXCR4, is capable of mobilizing both HSCs and prostate cancer cells from the niche. When combined with AMD3100, docetaxel (the standard of care for prostate cancer) is more effective at killing tumor cells than when given as a single agent. This indicates that mobilization of disseminated tumor cells (DTCs) from the endosteal niche by targeting the SDF-1/CXCR4 axis might cause these cells to be more sensitive to chemotherapy in patients. This strategy would theoretically kill DTCs, which, when they become metastatic lesions, lead to most of the morbidity and mortality that affects prostate cancer patients. We are engaged in a Phase 0 clinical trial in metastatic prostate cancer patients to test this strategy in human patients. To complement the human trial, we are testing several CXCR4 inhibitors combined with G-CSF and docetaxel. We are able to detect CTCs in human patients, as well as CTCs and DTCs in mice via several techniques, and will use this technology as a readout for the clinical trial, as well as the preclinical studies.

Citation Format: Kenneth C. Valkenburg, Kenneth J. Pienta. Mobilizing prostate cancer cells from the endosteal niche by targeting the SDF-1/CXCR4 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 365. doi:10.1158/1538-7445.AM2015-365
Abstract 3658: Structure-guided de novo design of selective Mcl-1 Inhibitors: Synthesis, structural and biochemical characterization
Authors: Mady Ahmed , Miao Lei , Perdih Andrej , Liao Chenzhong , Bajwa Naval , Stuckey Jeanne , Nikolovska-Coleska Zaneta
Published: August 2015

Myeloid cell leukemia-1 (Mcl-1) is a potent anti-apoptotic protein, member of the prosurvival Bcl-2 family. Amplification of the gene encoding the Mcl-1 protein is a common genetic aberration in human cancer. Overexpression of Mcl-1 is associated with high tumor grade, resistance to chemotherapy and poor prognosis in many types of cancers. Thus Mcl-1 is emerging as a critical survival factor in a broad range of human cancers and represents an attractive molecular target for development of a new class of cancer therapy. There is still need for developing BH3 mimetics that can efficiently and selectively target Mcl-1 protein.

Employing de-novo structure-based drug-design, we have designed a new class of potent small-molecule Mcl-1 inhibitors based on the initial lead compound discovered by high-throughput screening. Analyzing the binding model of this compound with Mcl-1 through computational docking supported by HSQC NMR studies, and using the structural information, we have designed and optimized a class of small-molecule inhibitors of Mcl-1 using a 2-(phenylsulfonamido) benzoic acid as a scaffold to reproduce the key interactions with Mcl-1 such as the conserved hydrogen bond between the aspartate in pro-apoptotic proteins and Arg263 in Mcl-1. Several co-crystal structures of this class inhibitors in complex with Mcl-1 have provided a basis for their further optimization which ultimately led to the discovery of ligands with low-nanomolar potency (Ki = 8 nM) and selectivity for Mcl-1. These compounds bind in the canonical BH3 binding groove. Interestingly, depending on the particular substitution pattern in the core scaffold, a “flip’ binding mode was observed where the substituents bind in a reverse orientation. These findings were confirmed by X-ray crystallography. The most potent compounds were further biologically characterized with a series of complementary biochemical and functional assays.

Citation Format: Ahmed Mady, Lei Miao, Andrej Perdih, Chenzhong Liao, Naval Bajwa, Jeanne Stuckey, Zaneta Nikolovska-Coleska. Structure-guided de novo design of selective Mcl-1 Inhibitors: Synthesis, structural and biochemical characterization. [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 3658. doi:10.1158/1538-7445.AM2015-3658
Abstract 3659: Target validation and structural optimization of selective small molecule survivin inhibitors as potential anti-cancer agents
Authors: Wang Jin , Xiao Min , Miller Duane D. , Li Wei
Published: August 2015

The anti-apoptotic protein survivin is highly expressed in most human cancer cells, but has very low expression in normal differentiated cells. Thus survivin is widely considered as an attractive cancer drug target. Herein we report the in vitro and in vivo study on a series of novel survivin inhibitors modified from our recently identified hit compound UC-112. NCI 60-cell line screening assay showed that three new analogs with oxyquinoline scaffold had GI50 values within the nanomolar range (513 nM, 646 nM and 891 nM). These new compounds can overcome the P-gp mediated multidrug-resistance in cancer cell lines. Western blot analyses demonstrated the high selectivity for survivin inhibition among other members in the inhibitor of apoptosis protein family. Mechanistic studies such as isothermal calorimetry measurements, survivin protein pull-down assay and fluorescence polarization assay confirm that these novel compounds indeed target survivin and inhibit its function. The most active compound showed excellent aqueous solubility of 148.7 μM (PBS pH7.4), reasonable metabolic stability (half-life) of 51 minutes as tested in human liver microsomes, and no significant inhibitions to major CYP enzymes except one of CYPs. Rat pharmacokinetic studies indicated good pharmacokinetic profiles for this novel scaffold. When tested in an A375 human melanoma xenograft model, this compound effectively suppressed tumor growth and strongly induced cancer cell apoptosis in tumor tissues. Overall, our study has demonstrated that these cell-permeable small molecular survivin inhibitors possessing a novel scaffold could be developed as potent anti-tumor therapeutic agents.

Citation Format: Jin Wang, Min Xiao, Duane D. Miller, Wei Li. Target validation and structural optimization of selective small molecule survivin inhibitors as potential anti-cancer agents. [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 3659. doi:10.1158/1538-7445.AM2015-3659
Abstract 3665: Reconstructing the itraconazole scaffold towards improved specificity for hedeghog signaling inhibition
Authors: DeBerardinis Albert M. , Hadden M. Kyle
Published: August 2015

Inappropriate regulation of the hedgehog (Hh) signaling pathway contributes to the development and progression of a number of human cancers; most notably, basal cell carcinoma (BCC) and medulloblastoma (MB). Clinical validation of small molecule mediated silencing of the Hh pathway was recently demonstrated by the FDA approval of vismodegib for the treatment of locally advanced BCC, representing a first-in-class inhibitor of the Hh signaling pathway. Vismodegib antagonizes Hh signaling by directly binding smoothened (SMO), a critical regulator of the pathway and target of numerous small molecule preclinical and clinical drug candidates. Vismodegib treatments in MB patients revealed novel SMO mutants that demonstrated resistance to vismodegib therapy, which underscores the need for continued development of novel Hh pathway inhibitors. Recently, itraconazole (ITZ) was identified as an inhibitor of Hh signaling in vitro and in vivo. Characterization indicated ITZ, a clinically used antifungal agent, maintains Hh inhibition against vismodegib-resistant SMO mutants. Ongoing structure-activity analysis for the ITZ scaffold is currently being investigated to identify analogues with improved potency and reduced non-specific toxicity. Results from these studies to identify candidates with improved pharmacological profiles and selectivity for Hh signaling modulation will be described herein.

Citation Format: Albert M. DeBerardinis, M. Kyle Hadden. Reconstructing the itraconazole scaffold towards improved specificity for hedeghog signaling inhibition. [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 3665. doi:10.1158/1538-7445.AM2015-3665
Abstract 368: Phage display derived antibodies for the detection of mesenchymal CTCs in TNBC
Authors: Kalscheuer Stephen , Panyam Jayanth
Published: August 2015

Enumeration of circulating tumor cells (CTCs) in peripheral blood of patients allows clinicians to obtain immediate evidence of metastatic dissemination. CELLSEARCH® is the only method currently approved for CTC detection (1). The critical assumption of the CELLSEARCH® platform is that CTCs will express EpCAM, based on the fact that the cell of origin in carcinoma is epithelial. However, the key cellular event that gives rise to CTCs is epithelial-to-mesenchymal transition (EMT) and the acquisition of the mesenchymal phenotype within the primary tumor. Recent studies have demonstrated EMT marker gene expression in CTCs, especially during disease progression. (2, 3). Thus, the full complement of CTCs may not be effectively captured with existing CTC technologies that rely solely on epithelial marker expression.

We have developed a competition based cell panning procedure which employs a commercially available single chain Fv (scFv) phage display library to identify antibodies with selective affinity to EMT phenotypic cells. HMLE cells as well as a sub-clone stably expressing Twist1 (HMLE-Twist1; a transcription factor known to induce EMT) were used for the panning. A two-color fluorescent scheme was employed to selectively sort the target cells from mixed populations containing the control cells, in the presence of phage library, via fluorescence activated cell sorting. Repeated iterations of this procedure yielded a scFv (‘clone 6′) with selective affinity to HMLE-Twist1cells. We then engineered a full length human IgG1 antibody in which the variable region sequences were derived from clone 6 scFv. The clone 6 IgG showed high affinity for metastatic cell lines (MDA-MB-231-LM2, LCC6, and HS578T). The antibody was also able to detect the presence of CTCs in an orthotopic MDA-MB-231-LM2 mouse tumor model. Intra-sample comparison showed distinct cell populations captured with anti-EpCAM antibody and Clone 6 IgG. Unlike those captured by anti-EpCAM antibody, the cells captured by Clone 6 were negative for EpCAM expression and were mesenchymal in phenotype. Future studies will examine the usefulness of clone 6 IgG in recognizing CTCs in patient samples.

References

1. M. Cristofanilli et al., New Engl J Med 351, 781 (2004).

2. B. Aktas et al., Breast Cancer Res : BCR 11, R46 (2009).

3. G. Kallergi et al., Breast Cancer Research : BCR 13, R59 (2011).

Citation Format: Stephen Kalscheuer, Jayanth Panyam. Phage display derived antibodies for the detection of mesenchymal CTCs in TNBC. [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 368. doi:10.1158/1538-7445.AM2015-368
Abstract 3796: Synergistic effect of targeted combination therapy in bladder cancer model using HSP90 inhibitors
Authors: Chelluri Raju , Agarwal Piyush K. , Neckers Leonard M. , Smith Armine K.
Published: August 2015

Background

In order to survive bladder cancer (BCa) cells deregulate key pathways - including the inhibition of apoptosis - while developing “oncogene addiction” to molecular chaperones such as Heat Shock Protein 90 (HSP90). While successful in vitro, monotherapies targeting such pathways have failed in clinical trials. We aim to show the superiority of combination therapy using proteostatic and targeted cytotoxic agents. We hypothesize that these combinations will synergize to increase cytotoxicity (CT) in BCa by amplifying the apoptosis via simultaneous activation of both intrinsic and extrinsic apoptotic pathways.

Methods

The proteostatic agents tested were the HSP90 inhibitor STA9090 (9090) and the proteasome inhibitor Marizomib (NPI). The cytotoxic agent used was a synthetic TRAIL ligand. These agents were tested in multiple ATCC BCa cell lines and the NCI patient tumor-derived BCa cell line UOBL 101 (UOBL). CT was determined via MTT assay. Mechanism of cytotoxic response was investigated using a series of immunoblots targeting the components of intrinsic and extrinsic apoptotic pathways and nuclear degradation proteins. All the assays were performed in triplicates for validation.

Results

9090 (IC50 7.4-10 nM) was effective in all cell lines, as was NPI (IC50 10-20 nM). TRAIL caused 100% CT in UOBL (IC50 2.5 ng/mL, 95% CI [1.0, 7.0]) and a maximal 50% CT in T24, however the cytotoxicity of individual agents was not sustained. Combination effects of 9090 + TRAIL and NPI + TRAIL demonstrated an increase from 50% CT to 100% CT at the IC50 of both 9090 and NPI, and this effect was sustained over the course of the experiment. The increased cytotoxicity of combination therapy had corresponded to rapid cytoplasmic release of cytochrome c, additive increase in cleaved caspase 3 and caspase 8, with the corresponding increase in cleaved PARP and γ-H2ax.

Conclusions

Combination of a pro-apoptotic agent with a molecular chaperone or proteasome inhibitor holds promise in BCa therapyby affecting intracellular anti-apoptotic signaling and compromising efficient proteostasis in cancer cells lines, and should be investigated further.

Citation Format: Raju Chelluri, Piyush K. Agarwal, Leonard M. Neckers, Armine K. Smith. Synergistic effect of targeted combination therapy in bladder cancer model using HSP90 inhibitors. [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 3796. doi:10.1158/1538-7445.AM2015-3796
Abstract 38: ERK5 inhibitor XMD8-92 for malignant mesothelioma treatment: A preclinical study
Authors: Shukla Arti , Thompson Joyce , Shukla Anurag , Leggett Alan , MacPherson Maximilian , Beuschel Stacie
Published: August 2015

Malignant mesothelioma (MM) is an aggressive cancer in dire need of successful treatment. We are first to demonstrate that Extracellular signal regulated kinase 5 (ERK5) plays an important role in MM pathogenesis (Clinical Cancer Research, 19, 2013). In this study we used genetically manipulated (ERK5 inhibited) MM cell lines to show that MM tumorigenesis is influenced by ERK5. In the present study we used recently discovered specific ERK5 inhibitor (XMD8-92) to show that it regulates MM tumor growth both in vitro and in vivo. First, we show that XMD8-92 inhibits ERK5 phosphorylation in 3 out of 4 MM cell lines showing constitutive activation of ERK5. Next, time and dose dependent study demonstrated significant cytotoxicity of XMD8-92 on 3 different MM cell lines. We also showed that soft agar colony forming efficiency of H2373 MM cells was significantly reduced in response to XMD8-92 treatment. Furthermore, in vivo studies using syngeneic mouse model of intraperitoneal mesothelioma showed that XMD8-92 treatment (50 mg/kg, ip, 1x daily for 3 weeks) resulted in significant decreases in tumor weights as compared to vehicle treated mice. Total and differential cell counts in peritoneal lavage fluid (PLF) were also inhibited by XMD8-92 suggesting the role of inflammation in MM tumor growth and their inhibition by ERK5 inhibitor. Cytokine profile assessment in PLF demonstrated the reduced levels of VEGF and IL-6 in XMD8-92 treated animals as compared to control animals receiving saline. FGF2, MCP-1, G-CSF, KC and RAGE levels were, however, not significantly impacted by the ERK5 inhibition. Taken together these data suggest that ERK5 inhibitor/ERK5 inhibition could be used for inhibiting the growth of MM tumors. Currently we are exploring the role of XMD8-92 alone as well as in combination with chemotherapeutic drugs for the treatment of MM using various mouse models. We are also exploring the role(s) of ERK5 inhibitor in inflammasome regulation by chemotherapeutics. This work is supported by NIEHS RO1 ES021110 and VCC/LCCRO grants.

Citation Format: Arti Shukla, Joyce Thompson, Anurag Shukla, Alan Leggett, Maximilian MacPherson, Stacie Beuschel. ERK5 inhibitor XMD8-92 for malignant mesothelioma treatment: A preclinical study. [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 38. doi:10.1158/1538-7445.AM2015-38
Abstract 3800: Targeting PI3K signaling pathway for therapeutic enhancement of verteporfin-mediated photodynamic therapy
Authors: Kraus Daniel E. , Palasuberniam Pratheeba , Chen Bin
Published: August 2015

Photodynamic therapy (PDT) induces cell injury and death through generation of reactive oxygen species (ROS) after light activation. Verteporfin is a photosensitizer that has been approved for the treatment of age-related macular degeneration and is under investigation for targeting tumor vasculature for therapy. Being primarily localized in mitochondria, verteporfin-mediated PDT induced a rapid apoptotic cell death in SVEC mouse endothelial cells by activating mitochondria-initiated cell death pathways. However, activation of phosphatidylinositol 3-kinase (PI3K) signaling pathway was also observed after verteporfin-PDT, especially at a sub-lethal dose. Furthermore, activation of PI3K signaling was correlated with cell growth after PDT. Thus, the goal of this study is to test the hypothesis that therapeutic outcome of verteporfin-PDT can be enhanced by targeting PDT-induced pro-survival PI3K signaling pathway.

We found that PI3K-mTOR pathway inhibitor NVP-BEZ235 (BEZ235) significantly inhibited PDT-induced PI3K signaling activation and combination of BEZ235 and verteporfin-PDT resulted in more cell apoptosis and greater inhibition in cell proliferation in SVEC cells. The combination therapy led to significantly more tumor growth inhibition than each individual treatment in the PC-3 prostate tumor model. Examination of tumor samples for PI3K signaling activity and functional vasculature revealed that the combination treatment caused more inhibition in PI3K signaling and reduction in functional blood vessels than verteporfin-PDT or BEZ235 treatment alone. These results indicate that targeting pro-survival PI3K signaling pathway is an effective approach for enhancing tumor response to verteporfin-PDT.

Citation Format: Daniel E. Kraus, Pratheeba Palasuberniam, Bin Chen. Targeting PI3K signaling pathway for therapeutic enhancement of verteporfin-mediated photodynamic therapy. [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 3800. doi:10.1158/1538-7445.AM2015-3800
Abstract 3827: Histone methytransferase SMYD3 promotes invasion in ovarian carcinoma spheroids
Authors: Lyu Tianjiao , Jia Nan , Wang Jieyu , Bast Robert C. , Baak Jan Pa , Hua Keqin , Feng Weiwei
Published: August 2015

Epithelial ovarian cancer (EOC) metastasis is a direct contributor to high recurrence and low survival for patients with this disease. The ovarian carcinoma spheroids are cells shed from primary tumors and aggregate in ascites, bearing the main responsibility for metastasis. “Epigenetic plasticity” refers to the capability of mammalian cells to alter their differentiation status via chromatin remodeling-associated alterations in gene expression. In the current study, we extend this attribute of epigenetic plasticity to aggressive ovarian cancer cells. Upon a simple alteration in the microenvironment, transition from two- to three-dimensional culture, we mimicked the in-vivo spheroids formation and found that a batch of histone markers and their enzymes experienced a marvelous elevation, among which, H3K4Me3 and its methyltransferase SMYD3 emerged as focal points for their concordant change. Using siRNA-SMYD3-depletion, we proved SMYD3 was attributed to the enhanced invasion and adhesion abilities in spheroids compared to monolayer cells. Gene array results further discovered that integrin-related molecules were suppressed after SMYD3 depletion. By ChIP assay, we confirmed SMYD3 and H3K4me3 binding increased at ITGA8, ITGB6 and ITGAM's promoter, indicating that SMYD3 regulated those downstream genes through methylating H3K4. At last, we reconfirmed the SMYD3 and integrin-mediated adhesion molecules were both activated in in vitro model, in vivo intraperitoneal xenograft model and EOC patients’ spheroids. Overall, our results indicated that ovarian cancer cell epigenetic plasticity reflects an inherent transcriptional flexibility for context-responsive alterations in phenotype and SMYD3 could be a promising epigenetic marker for anti-tumor treatment.

Citation Format: Tianjiao Lyu, Nan Jia, Jieyu Wang, Robert C. Bast, Jan Pa Baak, Keqin Hua, Weiwei Feng. Histone methytransferase SMYD3 promotes invasion in ovarian carcinoma spheroids. [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 3827. doi:10.1158/1538-7445.AM2015-3827
Abstract 3853: Unique pattern of copy number changes including chromothripsis in pulmonary adenocarcinoma with EML4-ALK fusion
Authors: Ninomiya Hironori , Kato Motohiro , Ogawa Seishi , Motoi Noriko , Takeuchi Kengo , Kodama Tatsushi , Sakamoto Hiroshi , Ishii Nobuya , Fujiwara Mutsunori , Ishikawa Yuichi
Published: August 2015

Background: A subset of lung adenocarcinomas harbors an ALK fusion gene resulting in dominant oncogenic activity. Gene inversion may play a main role in forming the fusion, not producing copy number changes. But its detailed mechanism leading to gene fusion remains unsolved.

MATERIAL AND METHODS: To elucidate the underlying mechanisms resulting in gene fusion, two cancer cell lines positive for EML4-ALK fusion and surgically resected frozen specimens (33 ALK fusion-positive and 95 ALK fusion-negative adenocarcinomas) were analyzed using Affymetrix CytoScan HD Array with Chromosome Analysis Suite (ChAS) software for cell lines (JFCR-LC649 and JFCR-LC654) and Affymetrix GeneChip Mapping 250K arrays for surgical specimens.

RESULTS: One (JFCR-LC649) of the two cell lines with gene fusion showed an extremely frequent change of copy number, or “copy number oscillation”, which we thought was chromothripsis. In surgical materials, 30% (10/33) of adenocarcinomas with ALK fusion harbored frequent copy number changes at various degrees, but not like the “oscillation” pattern. None of the 95 resected cases without ALK fusion showed similar genomic aberrations. These results from the JFCR-LC649 line and the 10 surgical cases indicated that genomic changes resulting in copy number alterations (including chromothripsis) as well as a simple inversion may cause gene fusion.

CONCLUSION: As well as genome inversion, copy number alterations such as chromothripsis contribute to gene rearrangements resulting in the EML4-ALK fusion.

Citation Format: Hironori Ninomiya, Motohiro Kato, Seishi Ogawa, Noriko Motoi, Kengo Takeuchi, Tatsushi Kodama, Hiroshi Sakamoto, Nobuya Ishii, Mutsunori Fujiwara, Yuichi Ishikawa. Unique pattern of copy number changes including chromothripsis in pulmonary adenocarcinoma with EML4-ALK fusion. [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 3853. doi:10.1158/1538-7445.AM2015-3853
Abstract 386: Rapid in situ RNA analysis of circulating tumor cells using magnetic micropore-based sorting and Turbo FISH
Authors: Ko Jin A , Bhagwat Neha , Yee Stephanie S. , Carpenter Erica , Stanger Ben , Issadore David
Published: August 2015

Pancreatic cancer is the fourth most common cause of cancer-related death in the US, with a five year survival rate of only 6%. Recently, we have shown that circulating pancreatic cells can be detected at the onset of the disease cycle (pre-Stage 1). Thus, there is both a great need for early diagnosis and an opportunity, due to the early presence of circulating tumor cells (CTCs). 

Currently, the gold standard for CTC detection cannot resolve these extremely sparse, heterogeneous CTCs, which do not have any single surface marker that allows them to be targeted. Additionally, conventional downstream analyses are limited to either protein expression of individual cells on-chip, which has limited sensitivity to relevant biomarkers, or genetic testing off-chip, which has limited practical clinical use.

To address these challenges, we have developed a rapid (< 40 minute) clinical diagnostic for pancreatic cancer detection. Our approach combines magnetic micropore based negative selection of immunomagnetically labeled leukocytes with on-chip RNA fluorescence in situ hybridization (Turbo RNA FISH) to profile individual cells. With the high-specificity of FISH and its ability to be multiplexed, combined with the high sensitivity of track-etched magnetic micropore (TEMPO) to rare cells, we have demonstrated the potential to diagnose pancreatic cancer early in the disease cycle.

A key innovation is the development of our magnetic micropore based sorting technique, which preserves the low cell-loss and specificity of microfluidic sorting but at >10x greater flow rates and an insensitivity to clogging, enabling use directly on whole blood. The TEMPO filter consists of an electroplated 1.25 μm NiFe, 30 μm pores, and 5.1×10E5 pores total. 

Using this chip, we have demonstrated extremely high sensitivity to rare cells (LOD ∼ 1 cell / 7.5 mL blood) and exponentially improved sorting efficiency by stacking multiple TEMPO filters in series. Furthermore, single molecules of targeted RNA in each individual CTC could be resolved.

Citation Format: Jin A Ko, Neha Bhagwat, Stephanie S. Yee, Erica Carpenter, Ben Stanger, David Issadore. Rapid in situ RNA analysis of circulating tumor cells using magnetic micropore-based sorting and Turbo FISH. [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 386. doi:10.1158/1538-7445.AM2015-386
Abstract 3876: Actionable somatic mutations in head and neck cancer prognosis
Authors: Nelson Omar L. , Wu Evan S. , Park Jong Y. , Zeitouni Joseph , Gomez Carmen R. , Reis Isildinha M. , Zhao Wei , Lee Eunkyung , Lin Hui-Yi , Franzman Elizabeth J. , Savell Jason , McCaffrey Thomas V. , Goodwin W. Jarrard , Hu Jennifer J.
Published: August 2015

Head and neck squamous cell carcinoma (HNSCC) is a devastating illness with racial/ethnic disparities in survival. Using a retrospective study design, we evaluated 210 somatic mutation sties of 10 cancer genes (i.e., CDKN2A, EGFR, FGFR3, HRAS, KRAS, MET, NOTCH1, PIK3CA, STK11, and TP53) in 214 HNSCC patients: 98 Non-Hispanic Whites (NHW; 46%), 72 Hispanic Whites (HW; 34%), and 44 African Americans (AA; 20%). HW and AA had significantly higher mutation rates for EGFR, HRAS, KRAS, and TP53 compared to that in NHW. Significantly higher proportions of HW (30%) and AA (18%) had at least 3 mutations compared to that in NHW (p<0.001). In logistic regression models using 2-year as the cut-off for early death, we evaluated the overall and racial/ethnic-specific somatic mutations that are associated with early death. Although there was no significant association between somatic mutation and early death in the total population, NOTCH1 mutations were significantly associated with early death in NHW (odds ratio [OR] = 5.51, 95% confidence internal [CI] = 1.22, 24.83). More importantly, the NOTCH1 mutation C6814T (P2272S) within the PEST domain was associated with early death (OR = 13.13 (95%CI = 1.61, 107.36). TP53 mutations were significantly associated with early death in HW (OR = 3.84; 95%CI = 1.08, 13.68) in multivariate analysis after adjustment for age, gender, tumor site, and tumor stage. With limited sample size, no association was observed in AA. This is the first pilot study evaluating the association between somatic mutations and HNSCC early death in a tri-racial/ethnic population. Our data suggest that HW and AA have higher EGFR mutations and these patients may benefit from mutation-targeted precision medicine to improve survival.

Citation Format: Omar L. Nelson, Evan S. Wu, Jong Y. Park, Joseph Zeitouni, Carmen R. Gomez, Isildinha M. Reis, Wei Zhao, Eunkyung Lee, Hui-Yi Lin, Elizabeth J. Franzman, Jason Savell, Thomas V. McCaffrey, W. Jarrard Goodwin, Jennifer J. Hu. Actionable somatic mutations in head and neck cancer prognosis. [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 3876. doi:10.1158/1538-7445.AM2015-3876