bluuebunny/crossref_metadata_2025 huggingface dataset:
Obesity-Related Dietary Behaviors among Racially and Ethnically Diverse Pregnant and Postpartum Women
Authors: Harris Ashley , Chilukuri Nymisha , West Meredith , Henderson Janice , Lawson Shari , Polk Sarah , Levine David , Bennett Wendy L.
Published: May 2016

Introduction.Obesity is common among reproductive age women and disproportionately impacts racial/ethnic minorities. Our objective was to assess racial/ethnic differences in obesity-related dietary behaviors among pregnant and postpartum women, to inform peripartum weight management interventions that target diverse populations.Methods.We conducted a cross-sectional survey of 212 Black (44%), Hispanic (31%), and White (25%) women, aged ≥ 18, pregnant or within one year postpartum, in hospital-based clinics in Baltimore, Maryland, in 2013. Outcomes were fast food or sugar-sweetened beverage intake once or more weekly. We used logistic regression to evaluate the association between race/ethnicity and obesity-related dietary behaviors, adjusting for sociodemographic factors.Results.In adjusted analyses, Black women had 2.4 increased odds of fast food intake once or more weekly compared to White women (CI = 1.08, 5.23). There were no racial/ethnic differences in the odds of sugar-sweetened beverage intake.Discussion.Compared with White or Hispanic women, Black women had 2-fold higher odds of fast food intake once or more weekly. Black women might benefit from targeted counseling and intervention to reduce fast food intake during and after pregnancy.
Abstract A124: Amino acid availability modulates vacuolar-ATPase activity and assembly
Authors: Stransky Laura A. , Forgac Michael
Published: May 2016

Mechanistic target of rapamycin complex 1 (mTORC1) activity is frequently deregulated in cancers of many etiologies. The mechanisms governing its activity in normal and pathological settings are incompletely understood, particularly with respect to amino acid sensing, an important contributor to its activation. Characterization of the processes controlling mTORC1 activation by amino acids may identify novel targets with broad application to cancer treatment.

Amino acid sensing requires the Vacuolar H+-ATPase (V-ATPase), an ATP-dependent proton pump primarily responsible for the pH control of intracellular compartments. While it is known that the intact V-ATPase is necessary for mTORC1 activation by amino acids, the mechanism of its involvement unknown. We thus sought to determine if amino acids affect activity of the V-ATPase by assessing V-ATPase-dependent fluorescence quenching in lysosomes loaded with FITC-Dextran (a pH sensitive dye). Our data demonstrate that amino acid starvation increases V-ATPase activity, and that readdition of amino acids reverses this effect. We then demonstrated that amino acid starvation increases V-ATPase assembly, a main mode of regulation of the enzyme whereby association of the two functional domains, V1 and V0, is altered to increase or decrease the amount of active V-ATPase. Similarly to the change observed in activity, the change in V-ATPase assembly is reversed by amino acid readdition. These changes do not depend on PI3K or mTORC1 activity, both of which have been linked to changes in V-ATPase activity and assembly in response to other stimuli, such as glucose and growth factor stimulation. Further, we sought to determine if specific amino acids recapitulate the effect of total amino acid starvation, and preliminary data suggest that leucine and lysine both play important roles in controlling V-ATPase activity. These studies demonstrate that amino acid sensing controls V-ATPase activity and assembly, which may be important in subsequent signaling to mTORC1.

Citation Format: Laura A. Stransky, Michael Forgac. Amino acid availability modulates vacuolar-ATPase activity and assembly. [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2015 Nov 5-9; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2015;14(12 Suppl 2):Abstract nr A124.
Abstract A147: MI130004, a new ADC with a payload of marine origin shows outstanding activity against HER2-expressing tumors
Authors: Aviles Pablo , Guillen Maria Jose , Dominguez Juan Manuel , Galmarini Carlos M. , Cuevas Carmen
Published: May 2016

PM050489 is a tubulin-binding agent originally isolated from the marine sponge Lithoplocamia lithistoides. This compound binds with very high affinity to β-tubulin at a new site, disrupting the microtubule network and impairing its function during divison, which leads to mitotic aberrations. MI130004 (a novel antibody-drug conjugate formed by PM050489, a non-hydrolysable linker and trastuzumab) was tested for its in vitro and in vivo activity against selected tumor cell lines with different levels of HER2 expression. In vitro MI130004 showed very high potency and good selectivity for tumor cells that overexpressed HER2, namely HCC-1954 (IC50, 0.036 μg/mL), SK-BR-3 (IC50, 0.017 μg/mL) and BT-474 (IC50, 0.156 μg/mL). At the cellular level, MI130004 impaired tubulin polymerization, causing disorganization and disintegration of the microtubule network which ultimately led to mitotic failure. Expressing HER2 cells of breast (BT-474 and JIMT-1), gastric (Gastric-008 and N87) and ovarian (SK-OV-3 and A2780cis) as well as negative HER2 breast (MDA-MB-231) and gastric (Hs748t) were subcutaneously implanted into immunosuppressed (SCID or athymic) mice. Tumor (ca. 115 mm3) bearing animals (N = 10/group) were randomly allocated to receive the ADC treatments (at different doses) or the appropriate control (placebo included). Treatments were administered weekly for 5

consecutive weeks. Tumor growth (as median/group) was calculated 2-3 times per week. Twenty-four hours after the first dose, representative tumors were dissected free and processed for HER2 expression (erb2) and chromatin organization (Hoescht 33258). The treatment with MI130004 induced a long lasting antitumor effect with statistically significant increases (P<0.05) in median survival time compared to placebo. The highest dose of MI130004 induced complete tumor remissions in mice bearing BT-474, JIMT-1, Gastric-008, N87, SK-OV-3 and A2780cis xenografts lasting up to 120, 60, 165, 63, 237 and 231 days, respectively. Also 24-h post-dosing, MI130004 induced a dose-dependent disappearance of HER2-expressing cells as well as mitotic aberrations, this last being consistent with the mechanism of action of PM050489. These results demonstrate that PM050489, a marine derived compound, is a novel and remarkable payload for the design of new ADCs with potential therapeutic anti-cancer properties.

Citation Format: Pablo Aviles, Maria Jose Guillen, Juan Manuel Dominguez, Carlos M. Galmarini, Carmen Cuevas. MI130004, a new ADC with a payload of marine origin shows outstanding activity against HER2-expressing tumors. [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2015 Nov 5-9; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2015;14(12 Suppl 2):Abstract nr A147.
Abstract A201: UCHL1 promotes distant tumor metastasis through its deubiquitinating effect on HIF-1α
Authors: Harada Hiroshi
Published: May 2016

A Hypoxia-inducible transcription factor, HIF-1, has been associated with distant tumor metastases; however, genes that activate HIF-1 and promote metastases remain largely unknown. Here we identified Ubiquitin C-terminal hydrolase-L1 (UCHL1) as an novel activator of HIF-1 and revealed that it directly bound to HIF-1α, the regulatory subunit of HIF-1, abrogated the von Hippel-Lindau-mediated ubiquitination of HIF-1α, upregulated HIF-1 activity, and consequently promoted metastasis in murine models of pulmonary metastasis. Blockade of the UCHL1-HIF-1 axis by silencing UCHL1 or a pharmacological UCHL1 inhibitor, LDN57444, suppressed the formation of metastatic tumors. The expression levels of UCHL1 correlated with those of HIF-1α and were strongly associated with the poor prognosis of breast and lung cancer patients. These results indicate that UCHL1 promotes metastases as a deubiquitinating enzyme for HIF-1α, which justifies exploiting it as a prognostic marker and therapeutic target of cancers.

Citation Format: Hiroshi Harada. UCHL1 promotes distant tumor metastasis through its deubiquitinating effect on HIF-1α. [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2015 Nov 5-9; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2015;14(12 Suppl 2):Abstract nr A201.
Abstract A202: Lipocalin2 stabilizes hypoxia inducible factor-1α through the iron delivery into normoxic cancer cells
Authors: Hama Susumu , Nakamura Ibuki , Nishimoto Akinori , Nishi Takayuki , Itakura Shoko , Kogure Kentaro
Published: May 2016

The level of lipocalin 2 (LCN2), a secreted protein of the lipocalin superfamily, increases in various tumors. Moreover, LCN2 is known to promote tumor progression with the hypoxia inducible factor-1α (HIF-1α)-mediated epithelial mesenchymal transition and angiogenesis. However, the mechanism of LCN2-induced HIF-1α activation is not fully understood. In the present study, we examined the effect of LCN2 on HIF-1α activation in the normoxic cancer cells that HIF-1α is immediately degraded. A mouse melanoma cell line, B16-F1, was treated with recomninant LCN2 protein under normoxic condition. After the treatment, the protein level and nuclear translocation of HIF-1α were examined by western blot and immunohistochemical staining, respectively. Moreover, the activation of HIF-1α was evaluated by reporter gene assay in the cells transfected with the plasmid DNA containing HIF-1α response element. The intracellular iron was quantified by iron colorimetric assay kit. In the B16-F1 cells treated with LCN2, HIF-1α protein was not degraded and its nuclear translocation was observed even under normoxic condition, suggesting that LCN2 stabilizes HIF-1α in normoxic cancer cells. Moreover, the HIF-1α-mediated transcriptional activity in the B16-F1 cells treated with LCN2 was significantly higher than those with non-treatment, suggesting that LCN2 activates HIF-1α in normoxic cancer cells. Next, to clarify the mechanism of LCN2-mediated HIF-1α activation, we hypothesized that LCN2, which is an Fe3+ transport protein, stabilizes HIF-1α by preventing the activity of proline hydroxylase (PHD) as a primary enzyme of HIF-1α degradation, because the increase of intracellular Fe3+ is known to inhibit PHD activity. As a result, the intracellular iron increased in the cells treated with LCN2 protein. Moreover, the level of HIF-1α by co-treatment of LCN2 with FeCl3 was higher than that only by LCN2 treatment. Collectively, LCN2-mediated Fe3+ influx stabilizes HIF-1α in normoxic cancer cells.

Citation Format: Susumu Hama, Ibuki Nakamura, Akinori Nishimoto, Takayuki Nishi, Shoko Itakura, Kentaro Kogure. Lipocalin2 stabilizes hypoxia inducible factor-1α through the iron delivery into normoxic cancer cells. [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2015 Nov 5-9; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2015;14(12 Suppl 2):Abstract nr A202.
Abstract A4: CXCR3-B-mediated signaling suppresses c-MET-mediated angiogenic signals through the down-regulation of HO-1 and VEGF expression
Authors: Balan Murugabaskar , Pal Soumitro
Published: May 2016

Chemokine receptor CXCR3 gene encodes two splice variants with opposite functions; CXCR3-A promotes cell growth whereas CXCR3-B is growth inhibitory. Down-regulation of CXCR3-B promotes survival of many cancer cell types including Renal Cell Carcinoma (RCC) cells. However, the molecular signaling events that mediate down-regulation of CXCR3-B are not fully understood. Growth factors mediated signaling events govern Alternative Splicing (AS) of genes. The receptor tyrosine kinase c-MET is over-expressed in RCC cells and signaling through its ligand Hepatocyte Growth Factor (HGF) can play a major role in the growth and survival of tumor. In this study, we investigated the role of c-MET-mediated signaling in the modulation of AS of CXCR3 gene and analyzed the impact of CXCR3-B-mediated growth inhibitory signals on c-MET-mediated angiogenic pathway in RCC cells. We found that HGF treatment down-regulated CXCR3-B expression in both primary renal epithelial (RPTEC) and RCC (786-O and ACHN) cells. We found that c-MET-mediated signaling increased the expression of gene AS regulating protein SAM68 (also a prognostic marker for RCC) and modulated the association of SAM68 with CXCR3 gene. Utilizing gene over expression approach, we found that CXCR3-B-mediated signaling down-regulated the expression of cytoprotective molecule Hemeoxygenease-1 (HO-1) and angiogenic factor VEGF at both transcription and protein levels. We also found that CXCR3-B-mediated signaling down-regulated c-MET-induced secretion of angiogenic factors from RCC cells, as observed by the decrease in the tube formation ability of endothelial (HUVEC) cells grown with supernatant obtained from CXCR3-B over-expressing RCC cells culture.

Together, we suggest that CXCR3-B-mediated signaling acts as a checkpoint for c-MET-mediated angiogenic signals in RCC. We propose a novel function for c-MET-mediated signaling in promoting angiogenesis in renal cancer cells through the down-regulation of CXCR3-B expression.

Citation Format: Murugabaskar Balan, Soumitro Pal. CXCR3-B-mediated signaling suppresses c-MET-mediated angiogenic signals through the down-regulation of HO-1 and VEGF expression. [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2015 Nov 5-9; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2015;14(12 Suppl 2):Abstract nr A4.
Abstract A41: Targeting the ARK5/MYC axis to develop second generation CDK4/6 inhibitors
Authors: Parekh Samir , Perumal Deepal , Kuo Pei Yu , Leshchenko Violetta , Jiang Zewei , Chari Ajai , Cho Hearn Jay , Jagannath Sundar , Reddy E. Premkumar
Published: May 2016

Inhibition of cyclin dependent kinases CDK4/6 is an emerging therapeutic approach in myeloma and other cancers. A direct link between MYC and the cell cycle machinery exists through its ability to directly transactivate the CCND1 and CCNE genes. Deregulated MYC expression is linked to increased cyclin A, cyclin E expression and activation of CDK4 via its direct target gene CDC25. We describe a novel mechanism of MYC silencing that is therapeutically targeted by small molecule inhibitors. ARK5 (AMPK-related protein kinase 5), a novel member of the human AMPK family, represses MYC and MYC-driven gene expression, leading to rapid induction of apoptosis and cell cycle arrest. RNAi loss of function studies confirm that ARK5 decreases MYC expression via post-translational de-acetylation by NAD-dependent deacetylase sirtuin 1 (SIRT1). In ARK5 over-expressing primary myeloma cells and cell lines, treatment with dual ARK5 and CDK4/6 inhibitor ON123300 resulted in down-regulation of pS6K, Rb and MYC, resulting in cell cycle arrest followed by apoptosis in vitro. Treatment of MM tumor-bearing mice with ON123300 significantly decreased the growth of tumors as opposed to control treated mice and was well tolerated, with no significant weight loss. The CDK4/6 selective inhibitor PD0332991 in contrast does not inhibit the ARK5 or mTOR/pS6K pathways. We hypothesized that the modest activity for the current class of CDK4/6 inhibitors could be the result of a narrow kinase inhibition profile and incomplete targeting of critical onco-kinases, resulting in cytostatic rather than cytotoxic effects. This is the first report showing that dual CDK4/6 and ARK5 inhibitor ON123300 has significant effects on mTOR and MYC. In summary, our results demonstrate a novel mechanistic connection between ARK5 and SIRT1 in the regulation of MYC in MM cells. Our data showing potent cytotoxicity of ARK5 inhibition along with inhibition of MYC and mTOR signaling provides the foundation for further development of ARK5 inhibitors and second generation CDK inhibitors such as ON123300 for MM and potentially other MYC over expressing cancers.

Citation Format: Samir Parekh, Deepal Perumal, Pei Yu Kuo, Violetta Leshchenko, Zewei Jiang, Ajai Chari, Hearn Jay Cho, Sundar Jagannath, E. Premkumar Reddy. Targeting the ARK5/MYC axis to develop second generation CDK4/6 inhibitors. [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2015 Nov 5-9; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2015;14(12 Suppl 2):Abstract nr A41.
Abstract A46: Exposure of quercetin, kaempferol and exogenous glutathione on the expression levels of phospho- and total-Akt in 3T3-L1 preadipocytes
Authors: Boadi William Y.
Published: May 2016

Obesity has been reported to be a risk factor for several types of cancer. The Akt pathway or PI3K-Akt pathway is a signal transduction pathway that promotes survival and growth in response to extracellular signals. Key proteins involved are phosphatidylinositol 3-kinase (PI3K) and Akt, or Protein Kinase B. Activated Akt mediates downstream responses, including cell survival, growth, proliferation, cell migration and angiogenesis, by phosphorylating a range of intracellular proteins. The effects of the exposure of several doses each of the flavonoids quercetin and kaempferol and that of exogenous glutathione (GSH) (at 0, 5 10, 15, 20 and 25 μM doses) for 24 hr on the expression levels of phospho- and total-Akt levels in 3T3-L1 preadipocytes were investigated. Phospho-, total-Akt and the expression levels of glyceraldehyde 3-phosphate dehydrogenase (GAPDH, which was used as an internal standard) were assayed by the MILLIPLEX® MAP mates protocol, based on the Luminex® xMAP® technology (Millipore Corp. St. Charles, MI). There were no significant differences in the expression levels of GAPDH for the tested compounds and at the different concentrations. Significant (p<0.05) decreases however, in the espression levels of phospho-Akt in cells for the quercetin, kaempferol and GSH treated samples were observed. Total-Akt levels remained the same and, were not significantly different from each other for the tested compounds and at the various concentrations. The results may indicate that exposure of 3T3-L1 preadipocytes to quercetin, kaempferol and GSH did not result in the activation of Akt suggesting the role of such compounds in blocking cell differentiation in 3T3-L1 cells. Thus, quercetin, kaempferol and GSH may significantly cause a decrease in the proliferation of 3T3-L1 preadipocytes, suggesting an association with the low phosphorylation of the Akt protein.

Citation Format: William Y. Boadi. Exposure of quercetin, kaempferol and exogenous glutathione on the expression levels of phospho- and total-Akt in 3T3-L1 preadipocytes. [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2015 Nov 5-9; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2015;14(12 Suppl 2):Abstract nr A46.
Abstract A60: Mcl-1 confers protection of Her2-positive breast cancer cells to hypoxia: Therapeutic implications
Authors: Bashari Muhammad Hasan , Fan Fengjuan , Vallet Sonia , Arn M. , Cardone Michael H. , Beckhove Philipp , Schneeweiss Andreas , Sattler Martin , Opferman Joseph T. , Jäger Dirk , Podar Klaus
Published: May 2016

Molecular mechanisms leading to the adaptation of breast cancer (BC) cells to hypoxia are largely unknown. The anti-apoptotic Bcl-2 family member Myeloid cell leukemia-1 (Mcl-1) is frequently amplified in BC; and elevated Mcl-1 levels have been correlated with poor prognosis. Here, we demonstrate for the first time strong correlation between Mcl-1 protein levels and hypoxia, predominantly in Her2-positive BC cells. Surprisingly, genetic depletion of Mcl-1 decreased Her2 and Hif-1α levels followed by inhibition of BC cell survival. In contrast, Mcl-1 protein levels were not downregulated after genetic depletion of Her2 indicating a regulatory role of Mcl-1 upstream of Her2. Indeed, Mcl-1 and Her2 co-localize within the mitochondrial fraction and form a Mcl-1/Her2- protein complex. Similar to genetically targeting Mcl-1 the novel small molecule Mcl-1 inhibitor EU-5346 induced cell death and decreased spheroid formation in Her2-positive BC cells. Of interest, EU-5346 induced ubiquitination of Mcl-1- bound Her2 demonstrating a previously unknown role for Mcl-1 to stabilize Her2 protein levels. Importantly, targeting Mcl-1 was also active in Her2-positive BC cells resistant to Her2 inhibitors, including a brain-primed Her2-positive cell line. In summary, our data demonstrate a critical role of Mcl-1 in Her2-positive BC cell survival under hypoxic conditions and provide the preclinical framework for the therapeutic use of novel Mcl-1- targeting agents to improve patient outcome in BC.

Citation Format: Muhammad Hasan Bashari, Fengjuan Fan, Sonia Vallet, M. Arn, Michael H. Cardone, Philipp Beckhove, Andreas Schneeweiss, Martin Sattler, Joseph T. Opferman, Dirk Jäger, Klaus Podar. Mcl-1 confers protection of Her2-positive breast cancer cells to hypoxia: Therapeutic implications. [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2015 Nov 5-9; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2015;14(12 Suppl 2):Abstract nr A60.
Abstract A65: A novel regulatory mechanism involving Ras-mediated activation of the zinc-finger transcription factor, SAF-1/MAZ induces EGFR/HER1 expression in breast cancer cells
Authors: Ray Alpana , Havis Brett , Ray Bimal
Published: May 2016

Tumor microenvironment (TME) plays a critical role in tumor growth, invasion and metastasis. In TME, epidermal growth factor receptor (EGFR) family members, including HER1, HER2, HER3 and HER4, are involved in determining aggressive growth of breast cancer due to their ability to transduce the growth promoting functions of growth factors. This activity is potentiated by the over-expression of these receptor molecules in cancer cells. To reduce the activity of EGFR molecules, various inhibitors have been developed. EGFR/HER1 tyrosine kinase inhibitors (TKIs), gefitinib and erlotinib, show antitumor activity but these drugs have not meet their primary goal of improved survival in the overall patient population. A similar TKI, lapatinib, has shown some limited success in breast cancer. While these therapeutic options focus on reducing the functional activity of EGFR protein, the fundamental and major problem of over-expression of EGFR in cancer cells has not been addressed.

In two-thirds of aggressive breast cancer patients, transcriptional induction of EGFR causes high EGFR/HER1 level. Therefore, the goal of this study is to identify a molecular mechanism by which EGFR is over-expressed in breast cancer cells. To understand how transcriptional induction might occur, we have explored a novel biosynthetic pathway for EGFR over-expression. Our findings reveal that EGFR promoter is significantly more active in MDA-MB-231 cells in comparison to MDA-MB-468 cells. When transcription factor SAF-1 was ectopically expressed in these cells, EGFR promoter activity was further increased in MDA-MB-231 cells. Since MDA-MB-231 cells contain a highly active form of Ras, the data suggested a possible Ras-mediated activation of SAF-1 which in turn induces EGFR expression. Consistent with these findings, inhibition of K- and H-Ras, by using both siRNA and CRISPR/Cas9-mediated knock-out systems, reduced the expression of EGFR in MDA-MB-231 cells. The effect was more profound when K-Ras was targeted for inhibition. Since SAF-1/MAZ is seen to be activated by Ras, our data implicates K-Ras - SAF-1/MAZ - EGFR axis in breast cancer cell growth. Our findings may provide new targets for breast cancer therapy.

Supported by grants from College of Veterinary Medicine Faculty Research Award Grant and MU Center for Botanical Interaction Studies Pilot Project Grant.

Citation Format: Alpana Ray, Brett Havis, Bimal Ray. A novel regulatory mechanism involving Ras-mediated activation of the zinc-finger transcription factor, SAF-1/MAZ induces EGFR/HER1 expression in breast cancer cells. [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2015 Nov 5-9; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2015;14(12 Suppl 2):Abstract nr A65.
Abstract A68: Design and development of HDAC6-selective inhibitors for hematological cancer treatment and solid tumor immunotherapy
Authors: Shuttleworth Stephen J.
Published: May 2016

Despite its name, HDAC6 is primarily a tubulin deacetylase, not a histone deacetylase, and it has emerged as a particularly interesting - and unique - cancer target as it plays an essential role in stress response and in aggresomal protein degradation: specifically, HDAC6 binds polyubiquitinated misfolded proteins and recruits misfolded protein cargo to dynein motors for transport to aggresomes. Cells deficient in HDAC6 fail to clear these misfolded protein aggregates from the cytoplasm, cannot form aggresomes properly, and are hypersensitive to the accumulation of misfolded proteins, which leads to activation of apoptosis. Further, recent evidence has emerged of HDAC6 modulating immune-related pathways in solid tumors, notably regulating PD-L1 expression through STAT3 modulation. In order to explore the clinical potential of HDAC6 in both hematological and solid tumor immunotherapy, we have de novo-designed a class of highly-potent, ultra-selective and orally-active HDAC6 inhibitors with potential in the treatment of multiple tumor types, including through immunotherapeutic mechanisms. Additionally, owing to the selectivity we have achieved for HDAC6 with our lead compounds, much greater opportunities for broader combination modalities can be realized, and we have established potentiation of our leads with other tumor immunotherapeutics in in vivo syngeneic models. The profile of our preclinical development candidate will be presented.

Citation Format: Stephen J. Shuttleworth. Design and development of HDAC6-selective inhibitors for hematological cancer treatment and solid tumor immunotherapy. [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2015 Nov 5-9; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2015;14(12 Suppl 2):Abstract nr A68.
Abstract A70: Immunmodulatory effects of the novel HDAC inhibitor resminostat
Authors: Kohlhof Hella , Prenzel Tanja , Hamm Sevetlana , Baumgartner Roland , Borgmann Matthias , Vitt Daniel
Published: May 2016

Introduction:

HDAC inhibitors mediate their anti-cancer activity at least in part via modulation of the immunogenicity of cancer cells and reactivation of effector cells of the immune system. Evaluating possible immunomodulating properties of resminostat which potentially contribute to its overall anti-tumoral effect should be investigated.

Experiments:

Resminostat was tested for its effects on mechanisms affecting the anti-tumoral immune response such as immunosuppression, immunogenicity, immunogenic cell death (ICD) and natural killer (NK) cell recognition in several cancer cell lines.

Results:

Resminostat has the potency to affect the immune response at different levels on tumor as well as immune cells. Resminostat strongly reduced the expression of immunosuppressive enzymes, IDO1 and ARG1, which deplete the tumor microenvironment of amino acids essential for T cell activity. Further, resminostat enhanced the expression of various tumor associated antigens and MHC class I molecules leading to enhanced tumor immunogenicity. This was further enhanced by upregulation of the ICD marker and dendritic cell engulfment signal Calreticulin. The additional upregulation of MHC class II molecules and co-stimulatory molecules by resminostat lead to convertion of tumor cells into so called unprofessional antigen presenting cells. Furthermore, by upregulation of NK cell activating ligands on tumor cells, resminostat increased NK cell-mediated tumor cell cytotoxicity.

Conclusion:

Resminostat displayed promising immunomodulatory effects and immune priming capacity. Thus, this novel HDAC inhibitor demonstrates potential synergistic effects with immunotherapeutics such as opsonizing antibodies (e.g. rituximab), immunostimulating agents (e.g. cytokines and TLR ligands) and immune checkpoint blockers (such as PD1/PDL1 and CTLA-4 inhibitors) which will be analyzed on functional assays and in vivo models.

Citation Format: Hella Kohlhof, Tanja Prenzel, Sevetlana Hamm, Roland Baumgartner, Matthias Borgmann, Daniel Vitt. Immunmodulatory effects of the novel HDAC inhibitor resminostat. [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2015 Nov 5-9; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2015;14(12 Suppl 2):Abstract nr A70.
Abstract A73: Antisense oligonucleotides targeting ENO2 have selective anti-proliferative effects in glioblastoma cell with 1p36 genomic loss
Authors: Shahmoradgoli Maria , Mazur Curt , Kim Youngsoo , Monia Brett P , Macleod A. Robert , Muller Florian M , Hart Christopher E
Published: May 2016

Essential cellular processes are often supported by redundant genes and pathways. Cancer cells frequently harbor deletions or mutations in essential genes and survive because of this functional redundancy. Identification of recurrent events that leave cancer cells selectively sensitive to inhibition of a particular pathway or gene can provides the opportunity to develop therapeutics with a high degree of selectivity for cancer cells over normal cells. In Glioblastoma Multiform (GBM), one of the most malignant brain cancers, roughly 3-5% of tumors have loss of the chromosome locus 1p36. Among the genes lost at this locus, is enolase1 (ENO1). Loss of ENO1 is tolerated by the cancer cells and is not required for cancer initiation or progression; likely because of functional redundancy with the closely related paralog, ENO2. However, it has been recently demonstrated that cancer cells with homozygous ENO1 loss are selectively sensitive to ENO2 inhibition. Here we investigate the potential of Antisense Oligonucleotides (ASO) targeted to ENO2 as a therapeutic approach to treat GBM with 1p36 loss of ENO1.We show that constrained ethyl (cET) modified ASOs targeting ENO2 produced dose dependent inhibition of ENO2 RNA and protein levels in GBM cells when delivered by free uptake. ASO-mediated ENO2 depletion led to the selective inhibition of ENO1-null cells and not the isogenic cells engineered to express ENO1. Moreover, cell cycle analysis is demonstrated that ENO2 depletion resulted in impaired proliferation due to unsuccessful transition of cells from G1 to S phase of the cell cycle. These findings support the application of ASO to elicit synthetic lethality in GBM tumors with 1p36 loss.

Citation Format: Maria Shahmoradgoli, Curt Mazur, Youngsoo Kim, Brett P Monia, A. Robert Macleod, Florian M Muller, Christopher E Hart. Antisense oligonucleotides targeting ENO2 have selective anti-proliferative effects in glioblastoma cell with 1p36 genomic loss. [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2015 Nov 5-9; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2015;14(12 Suppl 2):Abstract nr A73.
Abstract A75: Pan-cancer identification of fusion genes as therapeutic targets in cancer
Authors: Edgren Henrik , Ojala Kalle , Ruusulehto Anja
Published: May 2016

Fusion genes are a well established category of oncogenic mutations, in which a chromosomal rearrangement brings together parts of two different genes to create a novel gene with oncogenic potential. Originally identified mostly in hematologic malignancies (e.g. BCR-ABL1 in CML), over the last years fusion genes have been shown to occur in practically all types of cancers examined. Some of the best known examples of these are ETS-family gene fusions (e.g. TMPRSS2-ERG) in prostate cancer and EML4-ALK fusions in non-small cell lung cancer. Although the prevalence of a specific gene fusion in a single cancer is often low, the example of treating e.g. EML4-ALK fusion positive NSCLC cancers with ALK inhibitors, such as crizotinib and ceritinib, has shown gene fusions to have significant value as targets of drug development.

In this study, we describe a comprehensive survey of gene fusions in 7428 cancer samples from 27 different cancer types, using paired-end RNA-seq data created by the TCGA project. At least one fusion gene was identified in 3801/7428 samples, with over 9000 genes taking part in a total of ∼11000 fusion gene events. The number of gene fusions per tumor is highly variable, with the largest number of fusions typically identified in tumors with high level amplifications. This suggests many of the fusion genes identified here may arise as byproducts of amplicon formation, making them likely passenger events.

Several potentially targetable genes take part in gene fusions across different cancer types. For instance, we identified a total of 12 gene fusions involving ALK in 5 different cancer types. Strikingly, only 5 of these involve EML4 as 5' partner gene. NTRK fusions were identified in 24 samples from 9 cancer types, FGFR fusions in 39 samples from 14 cancer types and MAST family kinase fusions in 14 samples from 7 cancer types.

In summary, our analysis of fusion gene occurrence across a wide variety of cancer types identifies several interesting and potentially targetable gene fusions. Furthermore, it illustrates that many of these are found across several different cancer types, as well as involve a large number of different fusion partner genes, suggesting this may need to be taken into account in diagnostics of fusion genes as well as selection of patients for clinical trials.

Citation Format: Henrik Edgren, Kalle Ojala, Anja Ruusulehto. Pan-cancer identification of fusion genes as therapeutic targets in cancer. [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2015 Nov 5-9; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2015;14(12 Suppl 2):Abstract nr A75.
Abstract A90: Combination therapy of chemokine receptor inhibition plus PDL-1 blockade potentiates anti-tumor effects in a murine model of breast cancer
Authors: Jung Heiyoun
Published: May 2016

Trafficking and expansion of myeloid derived suppressor cells (MDSCs) plays a major role in immune suppression of tumor. MDSCs express chemokine receptors which likely mediate their recruitment to the tumor microenvironment. Suppression of T cells is also mediated by the interaction of programmed death-1 (PD-1) and its ligands which are abundantly expressed in cancer cells and immune infiltrates, including MDSCs. Here, we show that targeting both pathways through administration of a small molecule chemokine receptor antagonist (CCX9588 which blocks CCR1) and an anti-PDL1 antibody significantly reduced tumor burden in an orthotopic breast cancer mouse model. Primary tumor growth and lung metastasis were modestly reduced by either agent (anti-PDL-1 or CCR1 inhibitor) alone, but the combination of CCR1 inhibitor plus anti-PDL1 antibody resulted in significantly reduced tumor burden as compared to either of the single agents. Orthotropic breast cancer cell engraftment induces robust expansion of CD11b+Ly6Ghi Ly6Chi MDSCs, a subpopulation of which express CCR1. Analysis of the tumor infiltrating cells revealed that CCX9588 significantly reduced the number of MDSCs in primary tumors, showing that CCR1 blockade of MDSC trafficking translates into reduced tumor burden. Analysis of human breast cancer patient samples from The Cancer Genome Atlas (TCGA) database revealed that the CCR1 ligands, MCP-7 (CCL7) and RANTES (CCL5) are present at significantly higher levels in breast cancers as compared to normal breast tissue. Furthermore, CCR1 and its ligands, as well as PDL-1 are significantly higher in triple negative breast cancer samples than in the other breast cancer subtypes. These data are the first to show that CCR1 chemokine receptor antagonists can act synergistically with PDL-1 inhibitors, and suggest a novel approach for potentiating the activity of immune cell checkpoint inhibitors in the most aggressive forms of breast cancer.

Citation Format: Heiyoun Jung. Combination therapy of chemokine receptor inhibition plus PDL-1 blockade potentiates anti-tumor effects in a murine model of breast cancer. [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2015 Nov 5-9; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2015;14(12 Suppl 2):Abstract nr A90.
Abstract A91: BGB-283 exhibits potentials in enhancing tumor immune sensitization and lymphocyte function
Authors: Yuan Xi , Lian Mi , Wu Xiaoran , Luo Lusong
Published: May 2016

Oncogenic B-RAF, which drives cell transformation and proliferation, has been detected in a variety of human malignancies. Prior studies suggested that B-RAF-targeting therapies could increase antigen expression, decrease immune suppressive factors in tumor microenvironment, and improve homing of T effector cells to the tumors. Thus, selective BRAF inhibitors were suggested to possess immunosensitization properties and could be used in combination with immunotherapies to achieve more durable disease control/response in treating BRAF V600E melanomas. BGB-283 is a novel inhibitor of the RAF dimer with potent and reversible inhibitory activities against RAF family kinases including wild type A-RAF, B-RAF, C-RAF and B-RAF V600E. In addition, BGB-283 has potent inhibitory activity against EGFR (Mol. Can. Ther. onlinefirst 2015; doi: 10.1158/1535-7163). BGB-283 is currently under Phase 1 clinical investigations. In the present study, BGB-283 showed significant stimulation of melanocyte-differentiation antigens and HLA-A expression in BRAF V600E melanoma cell lines. Intermediate level of BGB-283 was found to stimulate proliferation of human peripheral blood mononuclear cell (PBMC) and isolated human CD4+ and CD8+ T cells. In addition, BGB-283 potentiated IFNγ releasing from PBMCs at concentration range of 30∼300 nM. In newly developed the 3D tumor spheroid/PBMC co-culture system, BGB-283 improved PBMC function as measured by cell proliferation and IFNγ production. The presence of PBMC also enhanced the anti-proliferation activity of BGB-283 on A375 BRAF V600E mutant melanoma cells in the co-culture system. Further studies have shed light on the mechanistic underpinning of the immunomodulation effect of BGB-283 and its potential differentiation from vemurafenib. These findings support BGB-283 as a promising clinical stage antitumor drug that could potentially combine with immunotherapies by enhancing immune sensitization and T cell function.

Citation Format: Xi Yuan, Mi Lian, Xiaoran Wu, Lusong Luo. BGB-283 exhibits potentials in enhancing tumor immune sensitization and lymphocyte function. [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2015 Nov 5-9; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2015;14(12 Suppl 2):Abstract nr A91.
Abstract A92: Effects of lenvatinib on tumor-associated macrophages enhance antitumor activity of PD-1 signal inhibitors
Authors: Kato Yu , Tabata Kimiyo , Hori Yusaku , Tachino Sho , Okamoto Kiyoshi , Matsui Junji
Published: May 2016

[Background]

Lenvatinib mesylate (lenvatinib) is an orally administrated multiple receptor tyrosine kinase inhibitor that selectively inhibits the kinase activities of VEGFR1-3, FGFR1-4, KIT, PDGF-Rα and RET, which are involved in tumor angiogenesis and proliferation of several cancer types such as thyroid cancer. In order to investigate whether lenvatinib has effects on immune cell population in tumors, we examined antitumor effects and immune modulating abilities of lenvatinib in syngeneic murine tumor models.

[Materials and methods]

We examined antitumor activity of lenvatinib against BNL 1ME A.7R.1 murine hepatocellular carcinoma cell line in syngeneic and immune-deficient mice model. Combination treatment with lenvatinib at 10mg/kg (qd) plus anti-PD-1 mAb at 500μg/ml (twice weekly) was conducted in LL/2 murine Lewis lung carcinoma cell line syngeneic model. For immune population analyses, lenvatinib at 10mg/kg was orally administrated in BNL 1ME A.7R.1 or LL/2 mice model. On Day 7 after starting treatment, percentages of immune populations were analyzed by flow cytometer. For cytokine analysis, IFNγ in CD8+ T cells was intracellularly stained and analyzed by flow cytometer. The expression of immune related molecules in tumors was analyzed by quantitative PCR.

[Results]

Lenvatinib showed more potent inhibition of BNL 1ME A.7R.1 tumor growth in immune functional mice than in immune-deficient mice when given the same dose. A flow-cytometry analysis of immune populations in both BNL 1ME A.7R.1 and LL/2 tumors indicated that lenvatinib significantly decreased the percentage of tumor associated macrophages (TAM) with CD45+CD11b+Ly6GlowLy6ClowF4/80+ markers. In addition, immune inhibitory molecules were downregulated in tumors. Lenvatinib significantly increased the percentage of IFNγ-secreting CD8+ T cells in regional lymph node. In LL/2 tumor model, the combination effect of lenvatinib with anti-PD-1 mAb showed a superior antitumor activity to each monotherapy.

[Conclusion]

These results indicate that lenvatinib has more potent antitumor effect in immune functional environment. It was noteworthy that lenvatinib decreased TAM population in tumors. Furthermore, lenvatinib enhanced antitumor activity of PD-1 signal inhibitors with combination treatment.

Citation Format: Yu Kato, Kimiyo Tabata, Yusaku Hori, Sho Tachino, Kiyoshi Okamoto, Junji Matsui. Effects of lenvatinib on tumor-associated macrophages enhance antitumor activity of PD-1 signal inhibitors. [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2015 Nov 5-9; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2015;14(12 Suppl 2):Abstract nr A92.
Abstract B111: Efficacy of AKT inhibitor ARQ 092 compared with Sorafenib in a cirrhotic rat model with hepatocellular carcinoma
Authors: Roth Gael S. , Zeybek Ayca , Macek-Jilkova Zusana , Abbadessa Giovanni , Yu Yi , Marche Patrice , Leroy Vincent , Decaens Thomas
Published: May 2016

Background and aim: Hepatocellular carcinoma (HCC) is the third cause of cancer-related death worldwide. PI3K/AKT pathway is activated in 50% of HCC. The aim of this study was to assess toxicity and efficacy of a new AKT inhibitor (ARQ 092) compared to sorafenib.

Method: In vitro experiments were realized with viability, apoptosis and migration assays in 5 different cell lines to analyze the effects of ARQ 092 and sorafenib on the different mechanisms of carcinogenesis. Furthermore, an in vivo study with a diethylnitrosamine (DEN)-induced rat model of HCC on cirrhosis was performed. After 14 weeks of DEN, rats were randomized in 3 groups: control, sorafenib, or ARQ 092 and treated during 6 weeks. Tumor volume and progression were assessed by MRI before treatment, after 3 and 6 weeks of treatment. After euthanasia, pathological analysis of livers was performed assessing tumor numbers and volumes.

Results: ARQ 092 showed a dose-dependent decrease in cell viability, induction of apoptosis, and inhibition of migration. In vivo study on 26 rats showed a significantly lower tumor volume in ARQ 092 group: 45.9% of the control versus 60.4% of the control with sorafenib (p = 0.022). Number of tumors per liver was also significantly lower in ARQ 092 group with a mean of 53.9 tumors versus 96.3 tumors in control group (p = 0.001) and 96.8 tumors in sorafenib group (p = 0.015). MRI analysis showed a lower tumor progression in ARQ 092 group with 57.0% versus 155.3% in control group (p<0.0001), and 80.2% in sorafenib group (p = 0.08).

Conclusion: ARQ 092 was more significantly control tumor progression in a DEN-induced cirrhotic rat model with HCC compared to sorafenib.

Citation Format: Gael S. Roth, Ayca Zeybek, Zusana Macek-Jilkova, Giovanni Abbadessa, Yi Yu, Patrice Marche, Vincent Leroy, Thomas Decaens. Efficacy of AKT inhibitor ARQ 092 compared with Sorafenib in a cirrhotic rat model with hepatocellular carcinoma. [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2015 Nov 5-9; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2015;14(12 Suppl 2):Abstract nr B111.
Abstract B131: Chemical array screen identifies a novel MMP inhibitor NPDK-18 that suppresses tumor cell migration
Authors: Kawatani Makoto , Kondoh Yasumitsu , Honda Kaori , Sekine Tomomi , Osada Hiroyuki
Published: May 2016

Background and Purpose: Matrix metalloproteinases (MMPs) are zinc-dependent endopeptidases that degrade extracellular matrix components and that have been implicated in a wide range of pathological processes including tumor metastasis. Although many small-molecule MMP inhibitors have been developed, there are some problems related to specificity and side effects. In addition, recent studies have shown that several MMPs have nonproteolytic and tumor-suppressing functions depending upon the stage of cancer progression. Thus, much work remains for the elucidation of each MMP function and the development of new MMP inhibitors. Small-molecule microarrays represent a powerful new platform for facilitating target-based screens. Thousands of small molecules are arrayed on a slide glass and used to screen for protein-small molecule interactions in a high-throughput manner. We have developed unique photo-cross-linked chemical arrays, which enable the immobilization of a variety of small molecules on a solid support in a functional group-independent manner using a photoaffinity reaction. In this study, we screened MMP-9 inhibitors using the photo-cross-linked chemical arrays.

Methods and Results: The array slides with compounds of the RIKEN NPDepo chemical library were probed sequentially with His-Myc-tagged human recombinant MMP-9, anti-His antibody, and a Cy5-labeled secondary antibody, and then scanned with a microarray scanner. The fluorescence signals from two slides treated with and without His-Myc-tagged MMP-9 were quantified. We obtained 27 hit compounds among 24,275 compounds; 2 of the identified compounds (isoxazole compound NPDK-1 and naphthofluorescein) inhibited MMP-9 enzyme activity in vitro. We further explored 17 analogs of NPDK-1 and found that NPDK-18 showed the strongest inhibitory activity against MMP-9, with an IC50 value of 3.8 μM. NPDK-18 inhibited the enzyme activities of MMP-2, MMP-12 and MMP-13, as well as MMP-9 and MMP-9/ΔHemopexin, with almost identical potency, but showed no inhibition against MMP-1 and MMP-7. The biological potency of NPDK-18 was confirmed in a wound-healing scratch assay, where NPDK-18 significantly inhibited cell migration of human fibrosarcoma HT1080 cells with no cell toxicity at 10 μM.

Conclusion: These results suggest that NPDK-18 is a broad-spectrum MMP inhibitor. Our study has shown that the photo-cross-linked chemical arrays are useful for high-throughput drug screening.

Citation Format: Makoto Kawatani, Yasumitsu Kondoh, Kaori Honda, Tomomi Sekine, Hiroyuki Osada. Chemical array screen identifies a novel MMP inhibitor NPDK-18 that suppresses tumor cell migration. [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2015 Nov 5-9; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2015;14(12 Suppl 2):Abstract nr B131.