bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 5392: High-throughput screening of compounds for targeting system Xc(-) in human breast cancer cells
Authors: Lin Han-xin , Fazzari Jennifer , Linher-Melville Katja , Singh Gurmit
Published: May 2015

Breast cancer cells often metastasize to bone, causing severe and untreatable pain, which greatly compromises the quality of life of patients. We discovered that breast cancer cells secrete large amounts of glutamate via an amino acid antiporter Xc(-). Glutamate is an important neurotransmitter that has been associated with pain and it is also critical in bone metabolic remodeling. Excessive glutamate secreted by invading breast cancer cells can cause excitotoxic injury to bone neurons, disturb bone homeostasis, and activate or sensitize nociceptors, thereby causing acute and chronic pain. Another important activity of system Xc(-) is the import of cystine into cells, which is critical for antioxidant defence mechanism in cancer cells. These make system Xc(-) a potential target to overcome breast cancer induced-pain and sensitize cancer cells to oxidative damage.

A cell-based Amplex red glutamate assay was optimized and miniaturized to be compatible with high throughput screening (HTS) setting. The library for screening is the Canadian Compound Collection containing ∼30,000 compounds. We utilized the blockade of glutamate release to identify potential compounds.

We identified 320 compounds that effectively inhibit glutamate release of human breast cancer cell MDA-MB-231. However on closer scrutiny and stringent conditions we narrowed our potential compounds to 8 for further characterization. These compounds were tested in vitro for cytotoxicity and dose response effects on glutamate release.

In conclusion we have identified several lead compounds for in vivo pain monitoring tests. Information obtained from this study will not only advance our understanding of the role of system Xc(-) in breast cancer cells, but also help design novel drugs for blocking breast cancer induced-bone pain, thereby improving the quality of life for breast cancer patients.

(This project was supported by Canadian Breast Cancer Foundation-Ontario)

Citation Format: Han-xin Lin, Jennifer Fazzari, Katja Linher-Melville, Gurmit Singh. High-throughput screening of compounds for targeting system Xc(-) in human breast cancer cells. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 5392. doi:10.1158/1538-7445.AM2014-5392
Abstract 5405: Self-emulsifying nanoemulsions target intracranial glioblastoma tumors: In vivo optical imaging and fluorescence microscopy evidences
Authors: Liu Xinli , Qhattal Hussaini Syed Sha , Hye Tanvirul
Published: May 2015

Introduction: As the most aggressive form of brain tumor, glioblastoma has limited treatment options due to highly invasive nature, the largely impermeable blood-brain barrier (BBB) and presence of efflux transporters at BBB. We aimed to investigate the ability of self-emulsifying nanoemulsions to delivery BBB-impermeable molecules to the brain of orthotopic U87-luc human glioblastoma xenografts.

Methods: Oil-in-water nanoemulsions containing medium-chain triglycerides oil and co-surfactants (polysorbate-80:transcutol 6:4 w/w) were prepared by a spontaneous self-emulsification method. Luciferase transfected U87-luc human glioblastoma cells were injected intracranially as well as subcutaneously to Nu/Nu mice and tumor growth was monitored by bioluminescence imaging. Seventeen days after tumor implantation, near infrared dye DiD labeled nanoemulsions and size matched pegylated stealth liposomes were injected intravenously in U87-luc xenografts and the real-time biodistributions of formulations were monitored by whole animal imaging.

Results: Nanoemulsions had a mean particle size ∼120 nm with slightly negative charge (-10 mV). It is biocompatible with no hemolysis in vitro. In orthotopic U87-luc xenograft tumor model, intravenous administration of DiD labeled nanoemulsions revealed enhanced accumulation of the nanoemulsions in the brain and subcutaneous tumor at 1-9 hours post injection, comparing to that of the free dye and pegylated liposomes. The brain accumulations of nanoemulsions were persistent till 96 hours. The ex vivo fluorescence microscopy analysis showed that the DiD-nanoemulsions specifically accumuolated at the U87-luc tumor region, not at the surrounding normal brain tissue. In the normal CD-1 mice with intact BBB, the nanoemulsions accumulation in the brain was still significantly higher than the free dye, albeit lower than the brain level achieved in the U87-luc xenografts. The nanoemulsions were non-toxic to both CD-1 and Nu/Nu mice after daily repeated doses for three weeks.

Conclusion: Our data indicates that nanoemulsion is a promising nanocarrier to deliver imaging or therapeutic agents for the diagnosis or systemic treatment of GBM.

Citation Format: Xinli Liu, Hussaini Syed Sha Qhattal, Tanvirul Hye. Self-emulsifying nanoemulsions target intracranial glioblastoma tumors: In vivo optical imaging and fluorescence microscopy evidences. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 5405. doi:10.1158/1538-7445.AM2014-5405
Abstract 5406: The CendR pathway: A novel cell penetration and transcytosis pathway regulated by nutrient availability
Authors: Pang Hong-Bo , Braun Gary B. , Friman Tomas , Aza-Blanc Pedro , Ruidiaz Manuel E. , Sugahara Kazuki N. , Teesalu Tambet , Ruoslahti Erkki
Published: May 2015

Transport of molecules across the vascular wall, through tissue, and into target cells plays an important role in various physiological and pathological processes. We previously described a novel class of tumor-targeting peptides (CendR peptides) that penetrate into cells and tumor tissue through an as yet incompletely characterized transport pathway initiated by peptide binding to neuropilin-1 (NRP1). This pathway has shown promise in enhancing drug delivery into tumors. Here we perform a genome-wide RNAi screen to identify components responsible for cell penetration of CendR peptides. We show that the binding of CendR peptides to NRP1 initiates a novel endocytic process that depends on a subset of genes and a transport route distinct from those of known endocytic pathways. Strikingly, we found the internalization of CendR peptides to be controlled by nutrient-sensing and cell growth-regulatory pathways. Nutrient deprivation stimulated the cell penetration and intercellular transport of CendR peptides, both in vitro and in vivo. These data suggest a physiological role for CendR pathway in nutrient sensing and transport. The ability to activate this pathway with peptides provides immediate applications to drug delivery.

Citation Format: Hong-Bo Pang, Gary B. Braun, Tomas Friman, Pedro Aza-Blanc, Manuel E. Ruidiaz, Kazuki N. Sugahara, Tambet Teesalu, Erkki Ruoslahti. The CendR pathway: A novel cell penetration and transcytosis pathway regulated by nutrient availability. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 5406. doi:10.1158/1538-7445.AM2014-5406
Abstract 541: The role of long noncoding RNA SChLAP1 in prostate cancer
Authors: Sahu Anirban , Iyer Matthew K. , Prensner John R. , Chinnaiyan Arul M.
Published: May 2015

Prostate cancer is the second most common epithelial cancer and second leading cause of cancer death in men. Prostate cancers remain indolent in the majority of individuals but behave aggressively in a minority of patients. The molecular basis for this clinical heterogeneity remains incompletely understood. Long noncoding RNAs (lncRNAs) have emerged as a prominent layer of transcriptional regulation implicated in various biological and disease processes, including several types of cancer. Therefore, we hypothesized that lncRNAs may play a role in mediating aggressive prostate cancer.

Previously, our lab utilized RNA sequencing across a cohort of prostate tissues to discover differentially expressed lncRNAs in cancer versus benign samples. We identifed a novel transcript termed SChLAP1 (Second Chromosome Locus Associated with Prostate-1) overexpressed in a subset of localized and metastatic cancers. Clinical analyses showed that SChLAP1 levels independently predict poor outcomes, including metastasis and prostate cancer-specific mortality. In vitro and in vivo experiments indicated that SChLAP1 is critical for cancer cell invasiveness and metastasis. Mechanistically, we found that SChLAP1 binds to and antagonizes the genome-wide localization of the tumor-suppressive SWI/SNF nucleosome-remodeling complex.

Given the scaffolding capabilities of lncRNAs, previous studies describing an inhibitory role of SWI/SNF on PRC2 function, and the known oncogenic role of PRC2 in prostate cancer, we hypothesized that SChLAP1 may mediate PRC2 activity in conjunction with SWI/SNF to promote aggressive disease. Here, we show that a SChLAP1-associated gene signature nominates PRC2-related concepts, and in vitro studies indicate an interaction between SChLAP1 and PRC2 that enhances PRC2 genome-wide localization as well as its histone methyltransferase activity. These results suggest a mechanistic model in which SChLAP1 simultaneously engages multiple epigenetic complexes, inhibiting tumor-suppressive functions of SWI/SNF while enhancing oncogenic activity of PRC2, which leads to aggressive prostate cancer.

Citation Format: Anirban Sahu, Matthew K. Iyer, John R. Prensner, Arul M. Chinnaiyan. The role of long noncoding RNA SChLAP1 in prostate cancer. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 541. doi:10.1158/1538-7445.AM2014-541
Abstract 5412: Antitumor properties of ouabain in lipid double emulsion in orthotopic canine osteosarcoma xenografted mouse model
Authors: Wu Jun , Hsieh Meng-Yin , Wang Yan , Roberts Eugene , Vallejo Derek , Lee Abraham , Ermel Richard
Published: May 2015

Canine cancers develop in immunocompetent environments and share many similar features observed in human cancers, such as long periods of latency for initiation, progression and development, metastasis, acquired drug resistance and recurrence. Osteosarcoma is a primary bone tumor that occurs in dogs. It is highly aggressive and metastatic in nature, and the therapeutic methods utilized to treat this primary canine tumor are very limited. We have successfully established canine osteosarcoma orthotopic xenograft models with NSG mice (from Jackson Lab) in vivo. Ouabain, known as Na+, K+ ATPase inhibitor, has been used for many years in human clinical applications for the treatment of congestive heart failure. Recently, ouabain has been investigated for potential anticancer properties. However, the therapeutic dose window is very narrow because the side effects of ouabain overdose can be lethal. In an attempt to mitigate such narrow therapeutic margin of safety, ouabain was formulated in lipid double emulsion by using microfluidic synthesis method or membrane emulsification method. Our preliminary results showed enhanced antitumor activities of ouabain in double emulsion or nanoemulsion against canine osteosarcoma cell lines in vitro and in orthotopic xenograft models in vivo. The safety and delivery efficiency of ouabain in vivo was also improved, suggesting a therapeutic application of ouabain double emulsion could be benefit in canine osteosarcoma targeting therapy.

Citation Format: Jun Wu, Meng-Yin Hsieh, Yan Wang, Eugene Roberts, Derek Vallejo, Abraham Lee, Richard Ermel. Antitumor properties of ouabain in lipid double emulsion in orthotopic canine osteosarcoma xenografted mouse model. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 5412. doi:10.1158/1538-7445.AM2014-5412
Abstract 5426: Assessing factors predictive of response to ADCs for companion diagnostic strategies
Authors: Krueger Joseph S. , Lange Holger , Potts Steve , Young David
Published: May 2015

One premise of antibody-drug conjugates (ADC) is that the bound mAb-antigen complex on the cell surface will internalize and be metabolized by lysosomal proteases to release the free drug. Thus, the efficacy of an ADC is dependent not only on the presence of cell surface antigens, but also in-tact delivery of the conjugated drug, and an active pathway of receptor-mediated endocytosis. On a cellular level, the biological mechanisms behind receptor internalization and turnover have not been elucidated for novel therapeutic targets. On a tissue level, vascularization and hypoxic profile will affect delivery of the antibody into the tissue. Varying expression of the antibody target within the tissue (antigen density) will also affect the uptake of the ADC. Furthermore, the extracellular stability of the ADC may be affected by the activity of the various proteases in the tumor microenvironment (TME), outside of the lysozome. These concepts are critical for efficacy, and thus are especially important for companion diagnostic approaches (CDx) meant to predict response to ADCs. Thus, a predictive assay which would account not only for the degree of cell surface expression of the target, but also receptor internalization and potential effects of tumor microenvironment is required. To answer this need, Flagship Biosciences has invented several proprietary approaches for measuring critical properties of the therapeutic target on the cell surface or inside the cell, as well as properties of the TME which could be used to understand and predict efficacy to an ADC using FFPE biopsies. These quantitative pathology approaches are based on image analysis approaches which been designed specifically for ADC CDx programs to answer these critical questions: 1) Defining cell surface target expression independent of cytoplasmic expression; 2) Estimating receptor flux (turnover, internalization) based on staining profile; 3) Assessing critical factors in the TME which may affect delivery of the in-tact drug to the intracellular target; 4) Assaying vascular properties which may affect delivery of the drug; and 5) Heterogeneity of the target within a tumor. We are able to provide these as discrete evaluations or multiplex these evaluations for an integrative answer which can be derived from a typical clinical biopsy. Incorporation of these novel tools will enable ADC developers to create efficacy and patient stratification paradigms which incorporate the critical biological endpoints unique to ADCs.

Citation Format: Joseph S. Krueger, Holger Lange, Steve Potts, David Young. Assessing factors predictive of response to ADCs for companion diagnostic strategies. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 5426. doi:10.1158/1538-7445.AM2014-5426
Abstract 5428: Inhibition of collagen receptor discoidin domain receptor-1 (DDR1) tyrosine kinase enhances cytotoxicity of anti-mesothelin immunotoxin
Authors: Ali-Rahmani Fatima , Fitzgerald David , Martin Scott , Patel Paresma , Thomas Craig , Pastan Ira
Published: May 2015

Discoidin domain receptor 1 (DDR1) is an emerging anti-cancer target that belongs to the family of tyrosine kinases that are activated by collagen, the most abundant component of extracellular matrix (ECM). Collagen mediated induction of DDR1 facilitates cell adhesion, migration, proliferation and matrix remodeling. The collagen/DDR1 axis is also thought to modulate tumor-stromal interaction and potentially can affect tumor response to therapy. Mesothelin is a cell-surface tumor-associated antigen over-expressed in several human cancers including mesothelioma, ovarian, lung, breast, and pancreatic cancers with limited expression on normal cells. RG7787 is a second-generation less immunogenic recombinant immunotoxin (RIT) consisting of an Fv portion of an anti-mesothelin antibody attached to a portion of Pseudomonas exotoxin A in which immunogenic B cell epitopes were silenced. Activation of tyrosine kinase mediated signaling in tumor cells has been shown to modulate the activity of RITs. Therefore, we hypothesized that DDR1 regulates RIT action and its inhibition might enhance the activity of RG7787. Knockdown of DDR1 by siRNA or its inhibition with a novel and specific inhibitor, ‘7rh’, synergistically enhanced the cytotoxic activity of RG7787 in several cancer cell lines. Investigation into the mechanism of action showed increased inhibition of protein synthesis in cells treated with combination of ‘7rh’ and RG7787. Furthermore, stimulation of DDR1 activity by collagen treatment protected cancer cells from killing by RG7787. In conclusion, we report that collagen protects cells from killing by RITs through DDR1 and that lowering DDR1 protein or inhibiting DDR1 kinase activity enhances the cytotoxic activity of RITs. Our data suggest that the combination of ‘7rh’ and RG7787 represents a novel therapeutic strategy to target mesothelin-expressing cancers.

Citation Format: Fatima Ali-Rahmani, David Fitzgerald, Scott Martin, Paresma Patel, Craig Thomas, Ira Pastan. Inhibition of collagen receptor discoidin domain receptor-1 (DDR1) tyrosine kinase enhances cytotoxicity of anti-mesothelin immunotoxin. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 5428. doi:10.1158/1538-7445.AM2014-5428
Abstract 5433: Binding of multivalent CD147 extracellular domain on U937 cells induces apoptosis via activation of caspase-9 and downregulation of MCT-1
Authors: Intasai Nutjeera , Pata Supansa , Tragoolpua Khajornsak , Tayapiwatana Chatchai
Published: May 2015

CD147 is highly expressed in activated lymphocytes and cancer cells. CD147 plays many important roles in cellular functions. We have previously reported that binding of multivalent CD147 extracellular domain displayed on phage particles (multivalent CD147Ex phages) induced apoptosis in a monocytic cell line, U937, by triggering caspase-3 activation. Activation of caspase-8, caspase-9 and MCT-1 was investigated in this study. Levels of intracellular procaspase-9 and membrane MCT-1 were noticeably decreased in multivalent CD147Ex phage-induced U937 cells in comparison to controls. However, activation of caspase-8 was not obvious. Multivalent CD147Ex phages also induced DNA fragmentation and growth arrest of U937 cells. Taken together with previous studies, we hypothesize that the CD147 extracellular domain on phage particles may bind to its ligand(s) on the cell surface of U937 leading to inhibition of MCT-1 cell surface expression, intracellular activation of caspase-9, followed by caspase-3 activation, DNA fragmentation and growth arrest. We identified a mechanism of CD147 in triggering its ligand(s) on U937 cells leading to U937 apoptotic cell death. This may contribute to the conceptual development of novel cancer drugs based on CD147 polymers.

Citation Format: Nutjeera Intasai, Supansa Pata, Khajornsak Tragoolpua, Chatchai Tayapiwatana. Binding of multivalent CD147 extracellular domain on U937 cells induces apoptosis via activation of caspase-9 and downregulation of MCT-1. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 5433. doi:10.1158/1538-7445.AM2014-5433
Abstract 5435: Molecular subclasses of human hepatocellular carcinoma predict sensitivity to fibroblast growth factor receptor inhibition
Authors: Schmidt Benjamin , Wei Lan , DePeralta Danielle K. , Hoshida Yujin , Lanuti Michael , Tanabe Kenneth K. , Fuchs Bryan C.
Published: May 2015

Background: A recent gene expression classification of hepatocellular carcinoma (HCC) includes two poor survival subclasses, termed S1 and S2. S1 cells are more locally invasive, while S2 cells express E-cadherin and secrete AFP. As the expression of fibroblast growth factor receptors (FGFRs) differs between the S1 and S2 gene signatures, this study investigated whether molecular subclasses of HCC could predict sensitivity to FGFR inhibition. Methods: The effects of AZD4547 and BGJ398, two small molecule inhibitors of FGFR1-4, were assessed in four S1 and five S2 human HCC cell lines. Western blotting was performed for FGFR1-4 and phosphorylated p42/44 as a marker of MAPK signaling. Small interfering RNA (siRNA) transfection was used to assess the effects of individual FGFR knockdown. Results: Cell proliferation was inhibited at least two-fold more in S2 than in S1 cell lines at all doses tested above 1 uM of BGJ398 and AZD4547 (p<0.002). BGJ398 also decreased MAPK signaling in S2 but not in S1 cell lines. In xenografts from the S2 cell line HuH-7, BGJ398 reduced tumor volume by 70% over 14 days and this effect was associated with decreased MAPK signaling and a 36% decline in the number of Ki67 positive cells. BGJ398 had no effect on xenografts from the S1 cell line SK-Hep. All cell lines expressed FGFR1 and FGFR2, but only S2 cell lines expressed FGFR3 and FGFR4. FGFR4 siRNA decreased proliferation by 59 ± 12% in all five S2 cell lines (p<0.00001), a significantly greater decrease than seen with knockdown of FGFR1, FGFR2 or FGFR3. FGFR4 knockdown also decreased MAPK signaling in S2 cell lines. Conclusions: The S2 molecular subclass of HCC is sensitive to FGFR inhibition. FGFR4-MAPK signaling plays an important role in driving proliferation of S2 cells. This classification system may help identify those patients who are most likely to benefit from inhibition of the FGFR pathway.

Citation Format: Benjamin Schmidt, Lan Wei, Danielle K. DePeralta, Yujin Hoshida, Michael Lanuti, Kenneth K. Tanabe, Bryan C. Fuchs. Molecular subclasses of human hepatocellular carcinoma predict sensitivity to fibroblast growth factor receptor inhibition. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 5435. doi:10.1158/1538-7445.AM2014-5435
Abstract 5438: Resistance to β1 integrin inhibition in HER2 amplified cancers is mediated by HER3
Authors: Park Catherine C. , Amin Dhara , Zhang Hui , Moasser Mark
Published: May 2015

Our previous work focused on investigating β1 integrin as a target for BC, alone and with radiation therapy. It is well recognized that there is significant cross talk between the EGFR family of receptors and β1 integrin signaling, and a relative resistance of HER2+ BC to β1 integrin inhibitory agents. In 2006, we published our findings that β1 integrin inhibition enhanced the effect of trastuzumab in HER2+ BC. Subsequently, others and we reported that β1 integrin inhibition enhanced the cytostatic effect of multiple HER2-targeted therapies in HER2+ BC. However, the nature of cross talk between β1 integrins and HER2, and the role of β1 integrin in HER2+BC resistance remains unknown. In addition, HER2+BCs have higher rates of local recurrence after radiotherapy, consistent with previous work showing relative radioresistance in HER2+BC. The most potent mitogenic HER2 signaling in HER2+BC occurs upon heterodimerization with HER3. We hypothesized that HER3 could mediate significant resistance to β1 integrin targeting in HER2+BC.

To test this, we correlated β1 integrin and HER3 expression levels in SKBR3 cells over several time points using WB. Inhibitions of β1 integrin activity or shRNA knockdown of HER3 lead to upregulation of HER3 and β1 integrin, respectively. To test whether HER3 knockdown enhanced β1 integrin inhibition, we estimated proliferation by quantitating Ki-67 expression in SKBR3 cells in response to treatment. β1 integrin inhibition lead to a ∼50% reduction in Ki-67 positive cells (48%-22%), which was further enhanced by 50% with HER3 knockdown (12%), p<0.05. In addition, HER3 knockdown enhanced the cytostatic effect of ionizing radiation (IR) by ∼50% (32% to 14%), p<0.05. Enhanced cytostasis was correlated with a significant down modulation of p-HER2 and p-473 Akt on WB. In conclusion, our data indicate that HER3 plays a role in mediating resistance to β1 integrin inhibition in HER2+BC. Further mechanistic and in vivo validation are on going.

Citation Format: Catherine C. Park, Dhara Amin, Hui Zhang, Mark Moasser. Resistance to β1 integrin inhibition in HER2 amplified cancers is mediated by HER3. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 5438. doi:10.1158/1538-7445.AM2014-5438
Abstract 544: Recurrent DICER1 hotspot mutations in endometrial cancer and their impact on microRNA biogenesis
Authors: Chen Jiamin , Wang Yemin , McConechy Melissa , Anglesio Michael , Senz Janine , Yang Winnie , Rosner Jamie , Chu Andy , Cheng Grace , Morin Gregg , Huntsman David
Published: May 2015

Alternation in genes associated with microRNA (miRNA) biogenesis pathway may lead to miRNA dysregulation, and is implicated in a variety of human malignancies. Previously our group identified recurrent somatic “hotspot” mutations (E1705, D1709, D1810, E1813) in a critical miRNA-processing gene, DICER1, in rare sex cord-stromal tumors. During miRNA biogenesis, the two RNase III domains of DICER1 form an intramolecular dimer, which leads to the cleavage of the precursor miRNA (pre-miRNA) hairpin and generate mature 5p and 3p miRNAs from 5’ and 3’ arms of the precursor hairpin respectively. Studies have shown that the hotspot mutations in the RNase IIIb metal binding domain could impair DICER1's ability to generate mature 5p miRNAs, leading to global loss of 5p miRNAs.

Recently, in collaboration with The Cancer Genome Atlas (TCGA), we identified DICER1 hotspot mutations in a small subset of endometrial cancer from TCGA cohort (6/248) as well as our own tumor bank (6/307), suggesting disruption of DICER1 is implicated in a common malignancy. We also found an additional recurrent mutation G1809R and demonstrated that it has similar detrimental effects on miRNA biogenesis as hotspot mutations through deep sequencing and realtime PCR. Using Illumina Miseq targeted resequencing and Sanger sequencing, we observed biallelic DICER1 mutations in RNase IIIb domain in some but not all cases. miRNA deep sequencing confirmed that 5p miRNAs are decreased in both cell line models and endometrial tumors with hotspot mutations. Bioinformatic analysis of RNA sequencing profiles from TCGA dataset predicted hotspot DICER1 mutations to have greater functional impact than non-hotspot DICER1 mutations on gene expression. The oncogenic properties of DICER1 hotspot mutations are currently under investigation.

Citation Format: Jiamin Chen, Yemin Wang, Melissa McConechy, Michael Anglesio, Janine Senz, Winnie Yang, Jamie Rosner, Andy Chu, Grace Cheng, Gregg Morin, David Huntsman. Recurrent DICER1 hotspot mutations in endometrial cancer and their impact on microRNA biogenesis. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 544. doi:10.1158/1538-7445.AM2014-544
Abstract 5448: PTK7 as a potential therapeutic target in ovarian cancer
Authors: Ding Zhihu , Lennon Amanda , Perron Keli , Harper David , Su Hui , Wolfram Meredith , Murtie Joshua , Licht Stuart , Pinckney Jason , Simonds-Mannes Helene , Bishop Kimberly , Gavigan Julie-Ann , Bangari Dinesh , Magnay Maureen , Weber William , Reczek David , Brondyk William , Drewett Vicky , Trombe Marc , Hoffmann Dietmar , Baffa Raffaele , Silver Serena , Richon Victoria , Winter Christopher , Reddy Venkat , Gregory Richard C.
Published: May 2015

The identification of proteins that selectively discriminate between tumor cells and normal adult cells allows for the specific targeting of diseased cells with antibody therapeutics. One such recently identified protein, PTK7, is an onco-fetal membrane protein which exhibits limited expression and function in adults. PTK7 was identified as a member of the RTK super family but lacks a functional kinase domain. Normally, PTK7 is expressed early in development and its loss is associated with severe defects in neural tube closure and sensory hair cell bundle formation. Functionally, little is known about the signaling involving PTK7, but it has been linked to both the canonical and noncanonical WNT pathways. Recently, PTK7 expression has been shown to be upregulated in a number of cancers including: ovarian, melanoma, leukemia, lung, pancreatic, colon, renal and breast. In vitro and in vivo studies support a role in regulating angiogenesis, invasion & survival. To further validate PTK7 as a potential cancer target that may be required for tumor maintenance and progression, we analyzed the expression of PTK7 in normal and tumor samples, and validated an in vitro and in vivo role of PTK7 on cell growth in ovarian cancer cell lines using both genetic tools and polyclonal antibodies. Silencing PTK7 with stably expressed inducible shRNAs is shown to inhibit the growth of ovarian cancer cell lines in vitro and to lead to delayed tumor growth upon PTK7 knockdown in murine tumor xenograft models. Further supporting the role of PTK7 as a potential antibody target, polyclonal antibodies to PTK7 are shown to inhibit the growth of SKOV3 and OVCAR8 cells in vitro. Although 4 human antibodies derived from phage display failed to inhibit in vitro cell growth, these results suggest that functionally blocking PTK7 may lead to the inhibition of ovarian tumor growth and is a potential target for antibody therapies.

Citation Format: Zhihu Ding, Amanda Lennon, Keli Perron, David Harper, Hui Su, Meredith Wolfram, Joshua Murtie, Stuart Licht, Jason Pinckney, Helene Simonds-Mannes, Kimberly Bishop, Julie-Ann Gavigan, Dinesh Bangari, Maureen Magnay, William Weber, David Reczek, William Brondyk, Vicky Drewett, Marc Trombe, Dietmar Hoffmann, Raffaele Baffa, Serena Silver, Victoria Richon, Christopher Winter, Venkat Reddy, Richard C. Gregory. PTK7 as a potential therapeutic target in ovarian cancer. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 5448. doi:10.1158/1538-7445.AM2014-5448
Abstract 545: Frameshift mutations of chromosome cohesion-related genes SGOL1 and PDS5B in gastric and colorectal cancers with high microsatellite instability
Authors: Kim Min Sung , Choi Youn Jin , Gwak Min , Oh Hyerim , Choi Mi Ryoung , Hwang Sun Yong , Yoo Nam Jin , Lee Sug Hyung
Published: May 2015

Cohesin is a protein complex that regulates chromatid cohesion and plays a role in preventing aneuploidy and maintaining chromosomal stability. SGOL1 encodes a cohesin protector, and PDS5B encodes a regulatory cohesion factor. Both SGOL1 and PDS5B are considered putative tumor suppressor genes. The aim of this study was to explore whether SGOL1 and PDS5B genes are mutated and expressionally altered in gastric and colorectal cancers. A genome database indicated that both genes possessed mononucleotide repeats in coding sequences, which could be mutation targets in cancers with microsatellite instability. We analyzed mutations in 91 gastric cancers and 100 colorectal cancers with high microsatellite instability or stable/low microsatellite instability by single-strand conformation polymorphism analysis and DNA sequencing. We also analyzed SGOL1 and PDS5B expression by immunohistochemistry. Overall, we found 21 SGOL1 frameshift mutations in 21 cases and 18 PDS5B frameshift mutations in 16 cases. SGOL1 and PDS5B frameshift mutations were detected in 26.6% and 20.3%, respectively, of high microsatellite instability but not in stable/low microsatellite instability (0/112). By immunohistochemistry, losses of SGOL1 and PDS5B were identified in 19% to 47% of the gastric and colorectal cancers irrespective of microsatellite instability status. The losses were more common in those with frameshift mutations or high microsatellite instability than those without mutations or high microsatellite instability. The data indicate that frameshift mutations of SGOL1 and PDS5B and the loss of their expression may be a feature of gastric and colorectal cancers with high microsatellite instability. In addition, the data suggest that these alterations might contribute to cancer pathogenesis by deregulating cohesin-related functions.

Citation Format: Min Sung Kim, Youn Jin Choi, Min Gwak, Hyerim Oh, Mi Ryoung Choi, Sun Yong Hwang, Nam Jin Yoo, Sug Hyung Lee. Frameshift mutations of chromosome cohesion-related genes SGOL1 and PDS5B in gastric and colorectal cancers with high microsatellite instability. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 545. doi:10.1158/1538-7445.AM2014-545
Abstract 5463: ICG-001 induces cell cycle arrest to inhibit in vitro and in vivo pancreatic cancer growth
Authors: Arensman Michael D. , Lay Anna R. , Dawson David W.
Published: May 2015

Wnt/β-catenin signaling is an important mediator of pancreatic ductal adenocarcinoma (PDAC) initiation and progression, warranting its exploration as a target of therapeutic intervention. Wnt modulators with acceptable in vivo bioavailability and efficacy as Wnt/β-catenin transcriptional inhibitors have only recently become available and include the small molecule compound ICG-001. Discovered in a library screen for inhibitors of Wnt/β-catenin reporter activity, ICG-001 interacts with cyclic AMP-response element-binding protein (CBP) to block its function as a co-transcriptional activator of β-catenin/TCF-mediated transcription. ICG-001 inhibits both in vitro and in vivo colon cancer growth and has demonstrated efficacy in a variety of other in vivo Wnt-driven disease models. Having recently shown subsets of PDAC tumors are dependent on increased levels of Wnt ligand-dependent autocrine signaling, we addressed the efficacy and function of ICG-001 on in vitro and in vivo PDAC growth. As a single agent, ICG-001 significantly inhibited anchorage-dependent and -independent growth of PDAC cell lines in vitro and increased survival in an orthotopic xenograft model of PDAC (HR=0.207, 95%CI: 0.079-0.543, P=0.001). ICG-001 also synergized with gemcitabine to inhibit anchorage-dependent growth in vitro and improve survival in the orthotopic xenograft model (HR= 0.074, 95%CI: 0.026, 0.212, P<0.001). Mechanistically, ICG-001 treatment did not significantly increase apoptosis in PDAC cell lines, but instead dramatically inhibited cell cycle progression via G1 arrest. Although ICG-001 inhibited β-catenin/TCF-mediated transcription as measured by luciferase-based reporter assays and endogenous target gene expression, these changes occurred at concentrations above those seen to induce G1 cell cycle arrest, suggesting ICG-001 growth inhibitory effects may be unrelated to its specific effects as a Wnt pathway inhibitor. Gene expression microarray analysis revealed ICG-001 treatment alters the expression of known Wnt target genes, as well as a much larger number of genes involved in DNA replication and cell cycle control. In conclusion, this study indicates ICG-001 is a promising small-molecule therapeutic for pancreatic adenocarcinoma and identifies numerous target genes whose altered expression provide insights into mechanism of action and candidate biomarkers for following therapeutic response.

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Citation Format: Michael D. Arensman, Anna R. Lay, David W. Dawson. ICG-001 induces cell cycle arrest to inhibit in vitro and in vivo pancreatic cancer growth. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 5463. doi:10.1158/1538-7445.AM2014-5463
Abstract 5476: Simultaneous targeting of multiple cancer-related pathways in HPV-positive head and neck cancer with demethylating agents
Authors: Biktasova Asel , Gubanova Evgenia , Zou Mi , Yarbrough Wendell G. , Issaeva Natalia
Published: May 2015

Head and neck cancer is the 6th most common cancer worldwide with the vast majority being squamous cell carcinoma (SCC). The human papillomavirus (HPV) has recently been shown to be the etiologic agent in approximately 60% of head and neck SCC (HNSCC) arising from the oropharynx. Patients with HPV-positive HNSCC are clinically distinct from those with HPV-negative cancer being younger, and having decreased exposure to the established risk factors for HNSCC of tobacco and alcohol. Although HPV-positive HNSCC patients have higher cure rates compared to HPV-negative HNSCC, lifelong side effects of treatment are severe, and upon recurrence, treatment options and survival are limited. Therefore, new therapies with better side effect profiles that can also offer therapeutic options for patients with recurrent HPV-associated HNSCC are needed.

Recently, we found that HPV-positive head and neck cancer cell lines and primary cells were more sensitive than HPV-negative cells to the demethylating agent 5-azacytidine. Importantly, even low doses of 5-azacytidine delayed HPV-positive tumor growth in mouse xenograft model. Cancer-related pathways that we found to be altered in HPV-positive, but not in HPV-negative head and neck cancer cells, upon demethylation included: 1) restoration of tumor suppressor p53 levels and associated apoptotic activity; 2) activation of cytotoxic type I interferon response; and, 3) downregulation of matrix metalloproteinase (MMP1 and MMP10 ) expression.

Demethylation as a therapy for HPV-associated HNSCC is particularly interesting, since simultaneous targeting of multiple molecular pathways that are important for cancer development and progression, may increase efficacy and reduce acquisition of resistance. Based on our data, a small clinical trial with 5-azacytide on patients with HNSC C was initiated at Yale Cancer Center.

Citation Format: Asel Biktasova, Evgenia Gubanova, Mi Zou, Wendell G. Yarbrough, Natalia Issaeva. Simultaneous targeting of multiple cancer-related pathways in HPV-positive head and neck cancer with demethylating agents. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 5476. doi:10.1158/1538-7445.AM2014-5476
Abstract 5477: mTOR inhibitors protect from apoptosis in B-ALL cells treated with chemotherapeutic drugs
Authors: Vo Thanh-Trang , Fruman David A.
Published: May 2015

B-cell acute lymphocytic leukemia (B-ALL) is the most common pediatric cancer. Although most children can be cured by combination chemotherapy, 10-20% are refractory or have early relapses. These high-risk B-ALLs include Philadelphia chromosome (Ph+) and Ph-like with activated tyrosine kinases that drive elevated signaling through the PI3K/AKT/mTOR pathway. High mTOR signaling correlates with poor prognosis. These findings have raised interest in targeting mTOR in B-ALL and other leukemias. Novel inhibitors of the PI3K/AKT/mTOR pathway are in clinical development, but little is known about how these agents modulate responses to the standard of care in B-ALL. Here we have tested inhibitors of various nodes in the PI3K/AKT/mTOR pathway on B-ALL cells in combination with chemotherapeutic drugs. The data show that PI3K inhibition sensitizes B-ALL cell lines to chemotherapy but, surprisingly, mTOR inhibition induces resistance to several drugs. This chemotherapy resistance is observed when using rapamycin as well as active-site mTOR inhibitors (MLN0128 and AZD8055). The degree of drug resistance conferred by mTOR inhibition varies with different chemotherapeutic agents. mTOR inhibition best protects cells from killing by methotrexate and is least protective when combined with daunorubicin. In Ph+ cell lines, dasatinib also confers resistance to methotrexate that correlates with mTOR inhibition. We designed a simple in vitro model of relapse in which B-ALL cells were treated for 3 days with chemotherapy alone or with mTOR inhibitor, followed by 9 days of culture without drugs. Including mTOR inhibitor in the first culture period resulted in a striking increase in outgrowth of viable cells compared to chemotherapy alone. Mechanistic studies show that resistance does not correspond with effects on cell cycle. Preliminary data indicate that mTOR inhibition enhances DNA repair in B-ALL cells treated with etoposide or AraC. Further studies to define the resistance mechanisms and reverse the protective effect could potentially augment the effectiveness of inhibiting the PI3K/AKT/mTOR pathway. Our results will lead to a better understanding of how to best utilize PI3K pathway inhibitors in combination with standard chemotherapy as an option for high-risk B-ALL cases driven by oncogenic PI3K/AKT/mTOR signaling.

Citation Format: Thanh-Trang Vo, David A. Fruman. mTOR inhibitors protect from apoptosis in B-ALL cells treated with chemotherapeutic drugs. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 5477. doi:10.1158/1538-7445.AM2014-5477
Abstract 5478: A novel chloroquine derivative inhibits proteasome function, deregulates centrosome cycle and has selective anti-tumor activity
Authors: Pundir Sheetal
Published: May 2015

The use of chloroquine as a lead compound in anticancer drug discovery is being widely explored because of its low toxicity and ability to sensitize cancer cell killing when used in combination with radiotherapy. Therefore we designed and screened chloroquine analogs using hybrid pharmacophore approach. We report here anti-cancer activity and characterization of a novel anti-cancer compound VR23 derived from chloroquine. Biological investigation revealed that the derivative VR23 inhibits proteasome function, selectively kills cancer cells, causes the formation of supernumerary centrosomes, tri- and multi-polar mitosis, and slows down the progression of cells through cell cycle. In addition VR23 has shown promising anti-cancer activity in animal work.

Further, proteasome inhibition deregulates the translocation of cyclin E and other centrosomal proteins including plk4 and plk1 on centrosomes. This, in turn, leads to formation of supernumerary centrosomes, mitotic catastrophe, p38 stress kinase mediated activation of apoptosis, and eventually cancer cell death.

The work elucidates the consequences of the use of proteasome inhibitor on centrosome duplication cycle which, in turns also delays cell cycle progression and leads to the formation of irregular microtubule asters. Interestingly such phenotype was absent from non-malignant cell line.

Citation Format: Sheetal Pundir. A novel chloroquine derivative inhibits proteasome function, deregulates centrosome cycle and has selective anti-tumor activity. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 5478. doi:10.1158/1538-7445.AM2014-5478
Abstract 5483: Implications of the type of BRCA1 germline mutation in the treatment of patients with hereditary breast cancer
Authors: Vaclova Tereza , Woods Nicholas T. , Setién Fernando , Bueno José María García , Macías José Antonio , Barroso Alicia , Urioste Miguel , Esteller Manel , Monteiro Alvaro N.A. , Benítez Javier , Osorio Ana
Published: May 2015

Carrying an inherited mutation in BRCA1 or BRCA2 genes significantly increases individual´s lifetime risk to develop breast or ovarian cancer. However, there are considerable differences in disease manifestation among mutation carriers, suggesting the existence of other factors which could modify the risk of cancer development.

Since BRCA1 is an important component of DNA repair, BRCA1-deficient cells have to rely on other members of the DNA repair machinery. This makes them sensitive to drugs which target specific repair pathways, such as inhibitors of poly (ADP-ribose) polymerase (PARP). Several PARP inhibitors are currently being tested in preclinical or clinical trials, either as a monotherapy or in combination with chemo- or radiotherapy. Taking into account the promising results in patients with germline mutation in BRCA1 or BRCA2 genes, we aimed to investigate which BRCA1 mutation carriers, depending on the mutation type, might benefit from treatment with PARP inhibitors the most. In addition, we were also interested in identifying markers of sensitivity/resistance to PARP inhibitors.

We have used lymphoblastoid cell lines (LCLs) derived from lymphocytes of healthy women heterozygous for different BRCA1 mutations and healthy women from the same families with no mutation. In total, 23 LCLs were characterized according to their DNA repair capacity, growth rate, gene/miRNA expression and sensitivity to PARP inhibitors. Our preliminary results have shown significantly higher sensitivity to PARP inhibitor olaparib in cells harbouring missense mutation in comparison to cells with truncating or no mutation. The ongoing functional studies on these BRCA1 missense variants indicate that these mutations could act in a dominant negative manner. Importantly, the observed differences in sensitivity to olaparib among mutation carriers could be also relevant for recently proposed use of PARP inhibitors as prophylactic agent for mutation carriers.

Our ongoing work may help us to better understand the mechanisms of action of PARP inhibitors and lead to improvement of therapeutic strategies for BRCA1-associated cancers with extension to other tumors.

Citation Format: Tereza Vaclova, Nicholas T. Woods, Fernando Setién, José María García Bueno, José Antonio Macías, Alicia Barroso, Miguel Urioste, Manel Esteller, Alvaro N.A. Monteiro, Javier Benítez, Ana Osorio. Implications of the type of BRCA1 germline mutation in the treatment of patients with hereditary breast cancer. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 5483. doi:10.1158/1538-7445.AM2014-5483