bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 2814: Inhibition of PhIP/DSS-induced colon carcinogenesis by different forms of tocopherols in CYP1A-humanized mice
Authors: Yang Chung S. , Chen Jayson X. , Liu Anna B. , Wang Hong , Lee Marlon , Yu Siyuan , Zhao Chunfang , Kuo Ying-Yi , Chi Eric , Suh Nanjoo
Published: August 2015

Tocopherols (T), which exist as α-, β-, γ-, and δ-T are important dietary antioxidants. Our previous studies have demonstrated that a mixture of tocopherols (γ-TmT), δ-T, and γ-T effectively inhibited colon carcinogenesis in AOM/DSS-treated mice or AOM-treated rats. However, the effects of different forms of tocopherols on carcinogenesis in animal models and in humans remain controversial. In the present study, we investigated the inhibitory effects of α-T, γ-T, and δ-T on colon carcinogenesis induced by a dietary carcinogen, 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP), and promoted by dextran sodium sulfate (DSS) in CYP1A-humanized (hCYP1A) mice. At 5-6 weeks age, male hCYP1A mice were fed the AIN93M diet or diet supplemented with 0.2% α-T, γ-T, or δ-T for 1 week before PhIP treatment (2x 100mg/kg b.w., 3 days apart), which was followed by DSS (1.5% in drinking water for 4 days) treatment. The mice were sacrificed 10 weeks after the first dose of PhIP. Significant inhibition of colon tumor incidences was caused by dietary δ-T and γ-T, but not by α-T. In addition, oxidative and nitrosative stress markers (i.e., 8-oxo-dG and nitrotyrosine) as well as pro-inflammatory stress markers (i.e., PGE2, LTB4, NFκB and p-STAT3) were significantly reduced by δ-T. Dietary treatment with δ-T and γ-T significantly increased the δ-T and γ-T levels, respectively, in the blood and tissues, but their levels were still lower than the levels of α-T. However, the levels of their side-chain degradation metabolites (δ-T and γ-T forms of carboxyethyl hydroxychroman and carboxymethylbutyl hydroxychroman) were much higher than their parent tocopherols in blood and tissues. Altogether, we demonstrated the inhibitory effects of δ-T and γ-T in a dietary carcinogen-induced colon carcinogenesis model. The inhibition is possibly due to the antioxidative, reactive nitrogen species-trapping, and anti-inflammatory activities δ-T and γ-T. (Supported by NIH grants F31CA168333, RO1CA120915 and RO1AT007036 as well as shared facilities funded by CA72720 and ES05022)

Citation Format: Chung S. Yang, Jayson X. Chen, Anna B. Liu, Hong Wang, Marlon Lee, Siyuan Yu, Chunfang Zhao, Ying-Yi Kuo, Eric Chi, Nanjoo Suh. Inhibition of PhIP/DSS-induced colon carcinogenesis by different forms of tocopherols in CYP1A-humanized mice. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 2814. doi:10.1158/1538-7445.AM2015-2814
Abstract 2816: 2′-Hydroxyflavanone inhibits lung cancer growth by inhibiting tumor cell proliferation and angiogenesis
Authors: Singhal Sharad S. , Figarola James , Singhal Jyotsana , Nagaprashantha Lokesh , Awasthi Sanjay
Published: August 2015

Lung cancer remains the number one cause of cancer mortality in the US. Although systemic chemotherapy and novel biologic agents have been shown to improve survival, toxicities are often severe and effective preventive therapy has not been defined. Intake of flavonoids from citrus fruits has been shown to decrease the risk of lung cancer in large scale epidemiologic studies. Hence, we studied the biologic anti-lung cancer effects of 2HF, a naturally occurring compound in citrus fruits. Our in-vitro investigations revealed that 2HF inhibits the growth of bronchogenic adeno-, squamous and small cell lung carcinoma. This was associated with G2/M arrest, collapse in mitochondrial membrane potential (Δψm), and apoptosis. We demonstrated a down-regulation of fibronectin and key signaling proteins of the PI3K pathway. RLIP76 expression was significantly reduced by 2HF, and the survival as well as 14C-DOX transport on purified reconstituted-proteoliposomes prepared from H1618 and H358 cells was significantly inhibited, an effect which was synergistic with anti-RLIP76 IgG. Our in-vivo studies in mice xenografts confirmed our in-vitro results by marked reduction of tumor growth, which was paralleled by a reduction in Ki67 and CD31, as proliferative and angiogenic markers, respectively. Importantly, 2HF also reduced the level of vimentin and increased E-cadherin, reversing the phenotype of epithelial-to-mesenchymal transition. In summary, 2HF demonstrated strong anti-proliferative, anti-angiogenic and pro-differentiation effects on in-vitro and in-vivo lung cancer models, whilst sparing normal cells. This holds not only great promise for therapeutic use but also suggests a role for 2HF in the prevention of lung cancer (Supported by NIH grant CA 77495 and Beckman Research Institute of the City of Hope).

Citation Format: Sharad S. Singhal, James Figarola, Jyotsana Singhal, Lokesh Nagaprashantha, Sanjay Awasthi. 2′-Hydroxyflavanone inhibits lung cancer growth by inhibiting tumor cell proliferation and angiogenesis. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 2816. doi:10.1158/1538-7445.AM2015-2816
Abstract 2829: Discovery and in vitro and in vivo characterization of aminopyrazoline-based SMYD2 inhibitors
Authors: Stresemann Carlo , Hartung Ingo , Stellfeld Timo , Barak Naomi , Mowat Jeffrey , Christ Clara , ter Laak Antonius , Koehr Silke , Weiske Jörg , Hillig Roman , Badock Volker , Stoeckigt Detlef , Ziegelbauer Karl , Weinmann Hilmar , Gekeler Volker
Published: August 2015

SMYD2 (SET and MYND domain-containing protein 2) is a protein lysine methyltransferase (PKMT) which was initially described as a histone H3K36 and H3K4 methyltransferase involved in transcriptional regulation. SMYD2 has recently been reported to methylate and regulate several non-histone cancer relevant proteins such as p53, retinoblastoma protein (Rb) and the estrogen receptor alpha. Given the reports that overexpression of SMYD2 is linked to poor prognosis in certain cancers, SMYD2 is proposed to be an oncogene and an attractive cancer drug target.

Here we report the discovery of a novel potent and selective SMYD2 inhibitor, SMYD2-BAY-01, by high throughput screening and extensive biophysical validation. The co-crystal structure revealed that SMYD2-BAY-01 binds to the substrate binding site and occupies the hydrophobic pocket for lysine binding using an unprecedented hydrogen bond pattern. The competitive behavior of the inhibitor in biochemical assays is consistent with the binding mode observed in the crystal structure.

Further optimization generated SMYD2-BAY-02, which shows improved low nanomolar potency and is selective against kinases and other PKMTs. Furthermore, SMYD2-BAY-02 specifically inhibits SMYD2 methylation activity in a cellular assay with similar potency and reduces methylation of the tumor suppressor protein p53. Based on promising in vitro and in vivo DMPK data, SMYD2-BAY-02 was further characterized in vivo for SMYD2-specific methylation inhibition. In vivo activity could be shown upon in vivo administration at doses as low as 30 mg/kg p.o. in a SMYD2 overexpressing esophageal squamous cell carcinoma model. In summary, SMYD2-BAY-02 is a promising selective and potent SMYD2 inhibitor in vitro and in vivo and may represent a new treatment option for cancers overexpressing SMYD2.

Citation Format: Carlo Stresemann, Ingo Hartung, Timo Stellfeld, Naomi Barak, Jeffrey Mowat, Clara Christ, Antonius ter Laak, Silke Koehr, Jörg Weiske, Roman Hillig, Volker Badock, Detlef Stoeckigt, Karl Ziegelbauer, Hilmar Weinmann, Volker Gekeler. Discovery and in vitro and in vivo characterization of aminopyrazoline-based SMYD2 inhibitors. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 2829. doi:10.1158/1538-7445.AM2015-2829
Abstract 2875: p53 regulation of PDL1 is mediated through miR-34a
Authors: Cortez Maria A. , Valdecanas David , Wang Xiaohong , Ivan Cristina , Peltier Heidi , Ye Huiping , Araujo Luiz , Carbone David , Giri Dipak K. , Komaki Ritsuko , Krishnan Sunil , Skoulidis Ferdinandos , Heymach John , Calin George , Bader Andreas G. , Welsh James W.
Published: August 2015

Background Although clinical studies have shown promise for targeting PD1/PDL1 signaling in non-small cell lung cancer (NSCLC), little is known of how PDL1 expression is regulated. We previously found that miR-200s directly regulate PDL1; here we show that PDL1 is regulated by miR-34a and p53.

Methods We confirmed that PDL1 is a direct target of miR-34a with western blotting and luciferase assays. We then used in vitro models (p53-/- and p53+/+ HCT116 cells, p53-inducible H1299 cells, and p53-knockdown H460 cells) to determine if p53 regulates PDL1 via miR-34a. Next, we analyzed p53, PDL1, and miR-34a expression in formalin-fixed paraffin-embedded specimens from patients with NSCLC. Finally, we used a p53R172HΔg/+K-rasLA1/+ syngeneic mouse model to deliver miR-34a-loaded liposomes (MRX34) plus radiotherapy and assessed PDL1 expression and tumor-infiltrating lymphocytes (TILs).

Results We found that p53 regulates PDL1 via miR-34a, which directly binds to the PDL1 3′ untranslated region, in NSCLC. Delivery of MRX34 (previously tested in a phase I clinical trial) promoted CD8+ TILs and reduced CD8+PD1+ cells in vivo. Further, MRX34 plus radiotherapy increased CD8+ cell numbers more than either therapy alone. Finally, we showed that miR-34a delivery reduced the numbers of radiation-induced macrophages and T regulatory cells.

Conclusions We identified a novel mechanism by which tumor immune evasion is regulated by p53 and miR-34a via PDL1. Our results suggest that delivery of miR-34a combined with standard therapies, such as radiotherapy, may represent a novel therapeutic approach for lung cancer.

Citation Format: Maria A. Cortez, David Valdecanas, Xiaohong Wang, Cristina Ivan, Heidi Peltier, Huiping Ye, Luiz Araujo, David Carbone, Dipak K. Giri, Ritsuko Komaki, Sunil Krishnan, Ferdinandos Skoulidis, John Heymach, George Calin, Andreas G. Bader, James W. Welsh. p53 regulation of PDL1 is mediated through miR-34a. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 2875. doi:10.1158/1538-7445.AM2015-2875
Abstract 2882: Calorie-induced silencing of the tumor suppressive guanylin-GUCY2C paracrine axis underlies colorectal cancer in obesity
Authors: Blomain Erik S. , Lin Jieru E. , Colón-González Francheska , Kim Gilbert W. , Hyslop Terry , Zhan Tingting , Snook Adam E. , Waldman Scott A.
Published: August 2015

Despite a well-characterized epidemiological association, mechanisms underlying the relationship between obesity and colorectal cancer remain incompletely defined. An emerging paradigm suggests that colorectal cancer arises through silencing of the GUCY2C tumor suppressor by loss of the hormone guanylin, a gene product universally lost in colorectal cancer. The present studies demonstrate that obesity is linked to colorectal cancer through suppression of guanylin expression, resulting in silencing of the GUCY2C tumor suppressor and epithelial dysfunction underlying tumorigenesis. In mice, obesogenic diets suppress the expression of guanylin protein, silencing GUCY2C and increasing epithelial dysfunction including DNA damage, crypt hyperproliferation and metabolic reprogramming, associated with amplification of colorectal tumorigenesis. Mouse models uncoupling caloric intake and weight revealed that suppression of guanylin by obesogenic diets reflected ingested calories, rather than the endocrine, adipokine and inflammatory milieu associated with obesity. Further, suppression of guanylin expression by obesogenic diets in wild type mice could be reversed by switching to a lean diet. Importantly, genetically enforced tissue-specific expression of guanylin eliminated epithelial dysfunction and colorectal tumorigenesis in transgenic mice. Taken together, the present studies highlight the guanylin-GUCY2C axis as a principle mechanistic link between colorectal cancer and the increased caloric intake characterizing obesity. Moreover, they highlight a unique opportunity to prevent colorectal cancer in obese patients by oral replacement with FDA-approved GUCY2C ligands.

Citation Format: Erik S. Blomain, Jieru E. Lin, Francheska Colón-González, Gilbert W. Kim, Terry Hyslop, Tingting Zhan, Adam E. Snook, Scott A. Waldman. Calorie-induced silencing of the tumor suppressive guanylin-GUCY2C paracrine axis underlies colorectal cancer in obesity. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 2882. doi:10.1158/1538-7445.AM2015-2882
Abstract 2904: Induction of autophagy reveals a cytoprotective mechanism by which DNA polymerase gamma (pol-γ) prevents UVB-induced skin cancer
Authors: Dhar Sanjit K. , Bakthavachalu Vasu , Izumi Tadahide , Chen Jing , Zhu Haining , St. Clair Daret K.
Published: August 2015

DNA polymerase γ (polγ), a major mitochondrial DNA repair enzyme, is essential for replication and repair of the mitochondrial genome. It is well-established that mitochondrial genomic stability is important for healthy cell survival; however, whether and how polγ plays a role in autophagic response remain unknown. Our previous studies demonstrated that polγ is inactivated upon exposure to ultraviolet B (UVB) irradiation by a peroxynitrite-mediated mechanism. In this study, we identified tyrosine 964, which is near the active site of polγ, as the target of nitration-induced inactivation by the use of LC/MS/MS mass spectroscopy coupled to immunoprecipitation, Western blotting and a 3` to 5` exonuclease activity assay. Treatment of mouse skin epithelial cells with UVB radiation validated the presence of polγ nitration and reduction of its enzymatic activity in vivo. To determine the effect of UVB in autophagic responses, mouse skin epithelial (JB6) cells were transfected with GFP-labeled LC-3 cDNA and exposed to UVB radiation, or wild-type C57BL/6 mice were exposed to UVB radiation and examined for LC-3 cleavage products. UVB radiation of JB6 cells and mouse skin increased LC-3 cleavage products and beclin levels, suggesting increased autophagic response to UVB treatment both in vitro and in vivo. To elucidate the role of Polγ in autophagic responses, the expression of polγ in JB6 cells was inhibited by siRNA for polγ. Consistent with the role of polγ in preventing UVB-induced autophagy, knockdown of polγ led to an increase LC-3 cleavage products. Knockdown of polγ induced prosurvival autophagic responses via the mTOR-signaling-complex-1(mTORC1) and complex-2(mTORC2). Interestingly, the mTORC2 complex plays a prominent role in the prosurvival autophagy responses based on the bromodeoxyuridine uptake assay, following knockdown and overexpression approaches. Importantly, suppression of polγ alone leads to transformation and invasion ofJB6 cells, which are increased following UVB treatment. The results reveal a unique role of Polγ in autophagy and support the role of Polγ in the suppression of prosurvival autophagic response to UVB- mediated carcinogenesis.

Citation Format: Sanjit K. Dhar, Vasu Bakthavachalu, Tadahide Izumi, Jing Chen, Haining Zhu, Daret K. St. Clair. Induction of autophagy reveals a cytoprotective mechanism by which DNA polymerase gamma (pol-γ) prevents UVB-induced skin cancer. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 2904. doi:10.1158/1538-7445.AM2015-2904
Abstract 2907: AMPK induction, lysosomal acidification and melanoma survival increased by p53 dysfunction are counteracted by inhibiting autophagy
Authors: Chavez-Perez Valery , Strasberg-Rieber Mary , Rieber Manuel
Published: August 2015

BACKGROUND. In contrast to the majority of human tumors which harbour p53 mutations, melanomas preferentially have BRAF V600E or NRAS(Q61K) mutations accompanied by dysfunction of the p53 tumor suppressor gene, which may help progression from nevi to invasive melanoma. Although ultraviolet radiation accelerates BRAF-driven melanomagenesis by targeting TP53 (Viros A. et al, Nature. 2014;511:478-82), some evidences for an anomalous wt p53 response to radiation in melanomas are a dysfunctional pro-apoptotic DNA damage response which can be enhanced by radiation sensitizers like halogenated pyrimidines (Rieber M, Rieber MS.Cancer Biol Ther. 2008;7:1561-6), and a report that p53 knockdown in melanoma cells resulted in decreased proliferation (Avery-Kiejda KA1, Bowden NA, Croft AJ, Scurr LL, Kairupan CF, Ashton KA, Talseth-Palmer BA, Rizos H, Zhang XD, Scott RJ, Hersey P.BMC Cancer. 2011;11:203) .PURPOSE. Since activation of AMPK by AMP inhibits ATP-consuming metabolic pathways during metabolic stress which may cause autophagy, we investigated the possible role of melanoma p53 status in modulating response to AMPK activators and autophagy inhibitors, in physiological 5mM glucose. EXPERIMENTAL. Genetically-matched NRAS-mutant human melanoma cells with unequal p53 mutational status were exposed to different glucose levels in the presence of AMPK inducers including autophagy modulators. Formation of autophagic vesicles were quantitated by cytometric red/gree acridine orange fluorescence Apoptosis-associated PARP cleavage was studied by specific immune blotting. RESULTS. AMPK induction in response to AICAR was greater than that of metformin and was increased in mutant p53 cells, which also showed greater resistance to autophagy inhibition. Greater AMPK activation correlated with increased lysosomal acidification in mutant p53 cells. CONCLUSIONS. Our results identify a role for a) p53 dysfunction in enhancing the response to AMPK activators and autophagy to favour tumor cell survival, b) autophagy inhibitors in counteracting pro-survival AMPK signaling.

Citation Format: Valery Chavez-Perez, Mary Strasberg-Rieber, Manuel Rieber. AMPK induction, lysosomal acidification and melanoma survival increased by p53 dysfunction are counteracted by inhibiting autophagy. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 2907. doi:10.1158/1538-7445.AM2015-2907
Abstract 2950: Epigenetic silencing BCL6B inactivates p53 signaling and causes human hepatocellular carcinoma cell resist to 5-FU
Authors: Li Xin , Yu Jie , Brock Malcolm , Tao Qian , Herman James , Liang Ping , Guo Mingzhou
Published: August 2015

BCL6B is a potential tumor suppressor in human gastric cancer, but the regulation and mechanism of BCL6B in human hepatocellular carcinogenesis remain unclear. This study is to explore the epigenetic change and mechanism of BCL6B in human hepatocellular carcinoma (HCC). 14 hepatic cancer cell lines, 8 cases of normal hepatic tissue and 149 cases of HCC were employed. Semi-quantitative RT-PCR, methylation specific PCR (MSP), gene expression array, flow cytometry assay, colony formation, western blot and immunohistochemistry were used. BCL6B is methylated in 100% (14/14) of human HCC cell lines and 86.6% (129/149) of primary cancer samples. Methylation of BCL6B is associated with HBV positive (p<0.05). But no association was found with age, sex, tumor size, differentiation, TNM stage, recurrence and survival. Loss of BCL6B expression was found in 14 of completely methylated HCC cell lines. BCL6B was re-expressed after 5-aza-2′-deoxycytidine (5-AZA) treatment. Restoration of BCL6B expression suppressed cell proliferation, induced apoptosis and G1/S arrest in HCC cells. The expression of EGR1, a key component of p53 signaling, was increased after re-expression BCL6B in HCC cells. Re-expression of BCL6B activated P53 signaling and sensitized HepG2 and SNU449cells to 5-fluorouracil (5-FU). BCL6B is frequently methylated in human HCC and the expression of BCL6B is regulated by promoter region hypermethylation. BCL6B suppresses cell proliferation and induces apoptosis in HCC cells. BCL6B activates P53 signaling by increasing EGR1 expression in HCC. BCL6B sensitizes HCC cells to 5-FU by activating P53 signaling.

Note: This abstract was not presented at the meeting.

Citation Format: Xin Li, Jie Yu, Malcolm Brock, Qian Tao, James Herman, Ping Liang, Mingzhou Guo. Epigenetic silencing BCL6B inactivates p53 signaling and causes human hepatocellular carcinoma cell resist to 5-FU. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 2950. doi:10.1158/1538-7445.AM2015-2950
Abstract 2996: Hypoxia pathway activation in HPV-associated OSCC
Authors: Klussmann Jens P. , Knuth JeJennifer , Wagner Steffen
Published: August 2015

Introduction: HIF1A induces hypoxia dependent transcription of its target genes, resulting e.g. in increased glycolysis. However, increased glycolysis is found in a lot of tumors despite presence of oxygen, known as Warburg effect. Under normoxia HIF1A is hydroxylated by prolyl hydroxylases (PHDs) and degraded by the proteasome. According to literature, HIF1A protein level increase in the presence of HPV-oncoproteins. Nevertheless, HPV dependent induction of HIF1A and resulting downstream effects are still ambiguous for cervix carcinoma and even more for OSCC. We propose HPV-associated HIF1A induction in OSCC to be responsible for a Warburg-like effect, to gain nutrient resources for tumor growth and to be related to immune escape and metastasis. We investigated characteristic effects of HPV on the hypoxia signaling cascade and glycolysis for testing these hypotheses.

Experimental Procedure: HPV-positive and -negative cell lines were cultured under normoxic and hypoxic conditions or in the presence or absence of hypoxia mimics. We analyzed components of the hypoxia signaling pathway with molecular- and cell biological methods. Furthermore we analyzed HIF1A and its target gene expression in a cohort of 40 HPV-positive and -negative OSCC by immunohistochemistry.

Data: Immunoblot analysis showed 2.6 times higher HIF1A protein level (normalized relative expression) in normoxia and increased accumulation under hypoxic conditions in HPV-positive compared to HPV-negative (4.5 vs. 1.6 times more) cell lines. HPV-positive and -negative cell lines showed comparable PHD2 expression level under normoxia. Interestingly, PHD2 protein level increased more in HPV-positive than in HPV-negative cell lines (5.5 vs. 1.6 times) under hypoxia. Immunofluorescence also showed increased basal HIF1A expression and higher up-regulation after incubation with deferoxamine in HPV-positive vs. -negative cell lines. Immunohistochemistry of GLUT1 in HPV-associated OSCC showed a distinct “HPV-pattern” with a more evenly distributed expression compared to HPV-negative OSCC.

Conclusion: Our findings suggest a difference between HPV-induced and HPV-unrelated carcinogenesis in OSCC referring to up-regulation of HIF1A and activation of its target genes under normoxia as well as under hypoxia. This was shown for HPV-positive cell lines in vitro and indicated for clinical samples. Current analyzes focus on HPV-oncoprotein interactions with cellular factors related to HIF1A protein degradation and stabilization.

Citation Format: Jens P. Klussmann, JeJennifer Knuth, Steffen Wagner. Hypoxia pathway activation in HPV-associated OSCC. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 2996. doi:10.1158/1538-7445.AM2015-2996
Abstract 3: Mechanistic rationale for MCL1 inhibition during androgen deprivation therapy
Authors: Santer Frédéric R. , Erb Holger H.H. , Oh Su Jung , Handle Florian , Feiersinger Gertrud E. , Luef Birgit , Bu Huajie , Schäfer Georg , Ploner Christian , Egger Martina , Rane Jayant K. , Maitland Norman J. , Klocker Helmut , Eder Iris E. , Culig Zoran
Published: August 2015

The cellular consequences of androgen deprivation therapy (ADT), a first-line therapy for locally advanced and metastatic prostate cancer (PCa), are induction of apoptosis or G1 cell cycle arrest. Inability of PCa cells to induce apoptosis is the starting point of development of castration resistance. Hence, an improved therapy should target the cell cycle-arrested cells in a combinatorial approach together with currently applied ADT. Here we set out to analyze whether MCL1, a pro-survival member of the BCL2 family and known mediator of chemotherapy resistance regulates the cellular response to androgen withdrawal. Analysis of MCL1 protein and mRNA expression in PCa tissue and primary cell culture specimens of luminal and basal origin, respectively, reveals higher expression in cancerous tissue compared to benign origin. Using PCa cellular models in vitro and in vivo we show that MCL1 expression is regulated through the action of androgens and upregulated in androgen-sensitive PCa cells when grown under steroid-deprived conditions. Analysis of the underlying mechanism suggests that regulation of MCL1 through the AR signaling axis is indirectly mediated via a cell cycle-dependent mechanism. Using constructs downregulating or overexpressing MCL1 we demonstrate that expression of MCL1 prevents induction of apoptosis when androgen-sensitive PCa cells are grown under steroid-deprived conditions. The BH3-mimetic Obatoclax induces apoptosis and decreases MCL1 expression in androgen-sensitive PCa cells, while castration-resistant PCa cells are less sensitive and react with an upregulation of MCL1 expression. Synergistic effects of Obatoclax with androgen receptor inactivation can be observed in androgen-sensitive cells. In addition, Obatoclax efficiently inhibits clonogenicity of primary basal PCa cells. Altogether, our results suggest that MCL1 is a key molecule deciding over the fate of PCa cells upon inactivation of androgen receptor signaling and provide a mechanistic rationale for a clinical assessment of a MCL1-targeting therapy adjuvant to ADT.

Citation Format: Frédéric R. Santer, Holger H.H. Erb, Su Jung Oh, Florian Handle, Gertrud E. Feiersinger, Birgit Luef, Huajie Bu, Georg Schäfer, Christian Ploner, Martina Egger, Jayant K. Rane, Norman J. Maitland, Helmut Klocker, Iris E. Eder, Zoran Culig. Mechanistic rationale for MCL1 inhibition during androgen deprivation therapy. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 3. doi:10.1158/1538-7445.AM2015-3
Abstract 30: Medi3622, a monoclonal antibody to ADAM17, inhibits tumor growth by inhibiting EGFR- and non-EGFR-mediated pathways
Authors: Sabol Darrin , RiosDoria Jonathan , Chesebrough Jon , Stewart David , Schifferli Kevin , Rothstein Raymond , Leow Ching Ching , Heidbrink-Thompson Jenny , Cheng Li , Du Qun , Xu Linda , Jin Xiaofang , Tammali Ravinder , Gao Chanshou , Friedman Jay , Wilkinson Brandy , Damschroder Melissa , Pierce Andrew , Patnaik MunMun , Zeng Rong , Wu Yuling , Spitz Susan , Robbie Gabriel , Roskos Lorin , Hollingsworth Robert , Tice David , Michelotti Emil
Published: August 2015

ADAM17 is the primary sheddase for HER pathway ligands. We report the discovery of a potent and specific ADAM17 inhibitory antibody, MEDI3622, which induces tumor regression or stasis in many EGFR-dependent tumor models. The inhibitory activity of MEDI3622 correlated with EGFR activity both in a series of tumor models across several indications as well as in a focused set of head and neck patient derived xenograft models. Cynomolgus monkey and rat PK/PD assays showed MEDI3622 inhibited TNFα shedding. Toxicity observed in cynomolgus monkey and rat was similar to EGFR inhibitor-induced rash. However, the antitumor activity of MEDI3622 was superior to that of EGFR/HER pathway inhibitors in OE21 head and neck and COLO205 colorectal xenograft models suggesting additional activity outside of the EGFR pathway. Combination of MEDI3622 and cetuximab in the OE21 model was additive and eradicated tumors. Proteomics analysis revealed novel ADAM17 substrates which function outside of the HER pathways and may contribute towards the antitumor activity of the monoclonal antibody.

Citation Format: Darrin Sabol, Jonathan RiosDoria, Jon Chesebrough, David Stewart, Kevin Schifferli, Raymond Rothstein, Ching Ching Leow, Jenny Heidbrink-Thompson, Li Cheng, Qun Du, Linda Xu, Xiaofang Jin, Ravinder Tammali, Chanshou Gao, Jay Friedman, Brandy Wilkinson, Melissa Damschroder, Andrew Pierce, MunMun Patnaik, Rong Zeng, Yuling Wu, Susan Spitz, Gabriel Robbie, Lorin Roskos, Robert Hollingsworth, David Tice, Emil Michelotti. Medi3622, a monoclonal antibody to ADAM17, inhibits tumor growth by inhibiting EGFR- and non-EGFR-mediated pathways. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 30. doi:10.1158/1538-7445.AM2015-30
Abstract 3008: CEP55 is a determinant of genomic instability in aneuploid breast cancer cells and facilitates anti-mitotic drugs resistance by interacting directly with HSF1
Authors: Kalimutho Murugan , Waddell Nicola , Jeffry Jessie , Srihari Sriganesh , Khanna Kum Kum
Published: August 2015

Centrosomal proteins have long been known to act as a scaffolds, spindle checkpoint regulators as well as key regulating elements of microtubule organizing centre, hence as an essential component of cell cycle progression. CEP55 is a centrosomal protein originally identified as an indispensable regulator of cytokinesis, the final stage of cell division that divides cytoplasm equally amongst two daughter cells. Failure to faithfully complete cytokinesis due to loss of CEP55 resulted in primary genetic lesion leading to multinucleated cells, thus promotes tumourigenesis and aneuploidy. CEP55 has been to shown elevated in wide range of cancers including breast cancer, however the molecular mechanisms on how CEP55 mediated genomic instability in cancers are still not well understood. To decipher the functional role of CEP55 in regulating genomic instability in breast cancer, through a series of in-vitro and in-vivo experiments we found that high levels of CEP55 is significantly associates with basal-like breast cancer subtype with poor overall survival, relapse free survival and distant-metastasis free survival. Furthermore, we found that depletion of CEP55 decreased cell viability due to induction of apoptosis and hamper primary tumour growth due to clearance of aneuploid cells that are particularly important in oncogenic transformation. In addition, depletion of CEP55 hindered premature exit of mitotic arrested cells and this is partly mediated through CEP55-dependent interaction with HSF1 during mitosis to facilitate development of aneuploidy. Strikingly, we found that CEP55 mediates anti-mitotic drug resistance to docetaxel and PLK1 inhibition. Depletion of CEP55 increased sensitivity to anti-mitotic drugs in particular docetaxel or PLK1 inhibitor, providing a rationale to target CEP55 and its dependent-pathways in aggressive and aneuploid breast cancer.

Citation Format: Murugan Kalimutho, Nicola Waddell, Jessie Jeffry, Sriganesh Srihari, Kum Kum Khanna. CEP55 is a determinant of genomic instability in aneuploid breast cancer cells and facilitates anti-mitotic drugs resistance by interacting directly with HSF1. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 3008. doi:10.1158/1538-7445.AM2015-3008
Abstract 3010: Meiotic cohesin REC8 associates with chromosome instability in melanoma
Authors: Escandon Julia , Lindsey Scott , Eller Mark S , Grichnik James M
Published: August 2015

Chromosome instability (CIN) is an important component of multiple malignancies. CIN is linked to drug resistance, tumor grade and aggressiveness. Melanoma is a highly chromosome instable tumor and CIN is a differentiating feature between melanoma and benign nevi. Meiotic proteins are present in melanoma and previous work in our lab suggests a role for these proteins in CIN and that melanoma cell replication has attributes as chromosome pairing and DNA bridging that can be explained by the presence of meiosis proteins. REC8 is a meiotic cohesin which holds sister chromatids together during the first meiotic division, therefore we sought to study the potential role for REC8 in CIN in Melanoma. A model of chromosome instability developed in melanocytic cell lines showed association of REC8 as CIN progressed strengthening the hypothesis that REC8 may be involved in CIN. We overexpressed REC8 in melanoma lines and in E6E7 melanocytes with a TET-inducible system and examined the change in the polyploid population by flow cytometry, cell count and FISH. Results suggest that overexpression of REC8 alone is not sufficient to drive polyploidy in these cell lines. Nevertheless, the association of REC8 with CIN is intriguing and further research in the role of meiosis proteins in melanoma is warranted. Future work will aim at dissecting coexisting meiotic proteins; specially the proteins of the cohesin ring and its regulatory proteins that are also present at high levels in melanoma and could be responsible of chromosome number abnormalities.

Citation Format: Julia Escandon, Scott Lindsey, Mark S Eller, James M Grichnik. Meiotic cohesin REC8 associates with chromosome instability in melanoma. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 3010. doi:10.1158/1538-7445.AM2015-3010
Abstract 3013: RNF126 promotes homologous recombination via regulating E2F1-mediated BRCA1 expression
Authors: Wang Ying , Deng Ou , Feng Zhihui , Du Zhanwen , Xiong Xiahui , Chen Ceshi , Ma Zhefu , Zhang Junran
Published: August 2015

Abstract: Homologous recombination (HR)-mediated DNA double-strand break repair is important for cell proliferation, cancer prevention and cancer therapy resistance. HR protein BRCA1 is one of the most important breast/ovarian cancer suppressor genes. Lower expression of BRCA1 has been frequently observed in a broad spectrum of human cancers, particularly sporadic breast/ovarian cancers. However, the mechanisms governing BRCA1 expression is elusive. RNF126 is an E3 ubiquitin ligase and is mapped to chromosome 19 p13.3 which is a commonly deleted region in ovarian cancers. In addition, high-resolution 19p13.2-13.3 allelotyping of breast carcinomas demonstrates frequent loss of heterozygosity. The Cancer Genome Atlas (TCGA) data suggested that RNF126 depletion present in a broad spectrum of human tumors, particularly ovarian cancers. Although a recent study suggested that RNF126 promote cell growth, the function of RNF126 remains largely unknown. Here, we demonstrate that RNF126 facilitates HR by promoting the expression of BRCA1, in a manner independent of its E3 ligase activity but depending on E2F1, a well-known transcription factor of BRCA1 promoter. In support of this result, RNF126 promotes transactivation of BRCA1 promoter by directly binding to E2F1. Most importantly, an RNF126 mutant lacking 10 amino acids that is responsible for the interaction with E2F1 has a dominant-negative effect on BRCA1 expression by suppressing E2F1-mediated transactivation of BRCA1 promoter and blocking the enrichment of E2F1 on BRCA1 promoter. Last, RNF126 depletion leads to the increased sensitivity to ionizing radiation (IR) and poly (ADP-ribose) polymerase (PARP) inhibitor. Collectively, our results suggest a novel role of RNF126 in promoting HR-mediated repair through the positive regulation on BRCA1 expression by binding to E2F1. This study not only offers novel insights into our current understanding of the biological functions of RNF126 but also provides a potential therapeutic target for the cancer treatment.

Key words: Homologous recombination, RNF126, E2F1, BRCA1

Citation Format: Ying Wang, Ou Deng, Zhihui Feng, Zhanwen Du, Xiahui Xiong, Ceshi Chen, Zhefu Ma, Junran Zhang. RNF126 promotes homologous recombination via regulating E2F1-mediated BRCA1 expression. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 3013. doi:10.1158/1538-7445.AM2015-3013
Abstract 3031: Replication stress and DNA damage promote genomic instability in near-tetraploid colorectal cancer cells
Authors: Quintanilla Isabel , Wangsa Darawalee , Brown Markus , Ercilla Amaia , Klus Greg , Vila Maria , Lozano Juan José , Szallsi Zoltan , Agell Neus , Castells Antoni , Ried Thomas , Camps Jordi
Published: August 2015

Aneuploidy represents a hallmark of most solid tumours, and potentially has a causal role in carcinogenesis. Also, cancer cells exhibit high rates of chromosome missegregation, which leads to chromosomal instability (CIN). However, a large amount of tumours present near-triploid karyotypes likely to go through a tetraploid transient stage. Here, by using DLD-1 isogenic diploid and tetraploid cell lines generated by single cell FACS-sorting, we aimed at exploring how polyploidization affects cellular functions and whether tetraploid per se generates CIN in a genomically stable background. First, gene expression and subsequent gene set enrichment analysis revealed that genes involved in the machinery of the DNA synthesis and replication, such as MCM2 and RRM2, and genes involved in cell cycle, were significantly upregulated in near-tetraploid cells compared to their diploid counterparts. Functionally, polyploidy cells exhibited replication stress, as indicated by higher levels of pCHK1, RPA and 53BP1 foci, which resulted in increased DNA damage in S- and M-phase. In addition, tetraploid cells displayed impaired proliferative capabilities as a consequence of a cell cycle delay confirmed by BrdU pulse and flow cytometry. Furthermore, near-tetraploid clones showed a higher amount of intracellular karyotypic heterogeneity due to the higher frequency of micronuclei formation compared to their diploid counterparts. In fact, polyploid cells displayed an increased tendency of abnormal anaphase events, including lagging chromosomes and acentric fragments. Interestingly, these heterogeneous cellular populations showed strengthen migratory capabilities and preliminary experiments suggested an increment in tumor invasiveness too. Taken together, our data suggest that near-tetraploid cells systematically undergo replication stress, which can be responsible for the increased levels of genomic instability.

Citation Format: Isabel Quintanilla, Darawalee Wangsa, Markus Brown, Amaia Ercilla, Greg Klus, Maria Vila, Juan José Lozano, Zoltan Szallsi, Neus Agell, Antoni Castells, Thomas Ried, Jordi Camps. Replication stress and DNA damage promote genomic instability in near-tetraploid colorectal cancer cells. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 3031. doi:10.1158/1538-7445.AM2015-3031
Abstract 3040: NANOG metabolically reprograms tumor-initiating stem-like cells in oxidative phosphorylation and fatty acid metabolism
Authors: Chen Chia-Lin , Uthaya Kumar Dinesh Babu , Punj Vasu , Xu Jun , Sher Linda , Hess Sonja , Machida Keigo
Published: August 2015

Stem cell markers such as NANOG have been implicated in various cancers; however, the functional contribution of pluripotency transcription factor NANOG to cancer pathogenesis has remained unclear. Gene profiling, proteomics and metabolomics approaches were combined to identify the pathway(s) altered in tumors. Here, we show that Toll-like receptor 4 (TLR4) signaling phosphorylates E2F1 to transactivate NANOG via TAK1/TBK1. CD133+/CD49f+ TICs isolated from human HCC have tumorigenic activities dependent on TLR4 and NANOG. Down-regulation of Nanog significantly reduces tumor development in obesity-associated HCC mouse model. In the search for the NANOG-dependent mechanisms underlying growth of tumor-initiating stem-like cells (TICs), NANOG ChIP-seq identifies the genes associated with mitochondrial metabolic pathways. The specific pathways, which were examined: oxidative phosphorylation (OXPHOS) and fatty acid oxidation were identified as novel NANOG-mediated oncogenic pathways by NANOG ChIP-seq analysis and metabolomics. The causal roles of NANOG in mitochondrial metabolic reprogramming occurs through the inhibition of oxidative phosphorylation (OXPHOS) and levels of mitochondrial ROS and activation of fatty acid oxidation (FAO), which are required for both the self-renewal of TICs and their drug resistance. PPARδ physically interacts with NANOG. Restoration of OXPHOS and inhibition of FAO rendered TICs susceptible to drug. Our data showed that NANOG reprogramming of mitochondrial metabolism was responsible for human TIC oncogenicity and chemo-resistance. Together, NANOG-mediated metabolic reprogramming through dysregulation of mitochondrial functions generates TICs and drives tumorigenesis, providing insight into mechanisms of NANOG-mediated metabolic reprogramming and its roles in TIC oncogenic activity and chemo-resistance. NANOG suppresses OXPHOS and activates FAO, thus inhibiting OCR and ROS production, conferring a tumor chemoresistant state. These novel pathways are potential drug targets for the highly malignant TICs found in cancer patients. Reversal of NANOG-dependent effects on OXPHOS and FAO gene may offer a noteworthy strategy of countering therapeutic drug resistance associated with NANOG activation.

Citation Format: Chia-Lin Chen, Dinesh Babu Uthaya Kumar, Vasu Punj, Jun Xu, Linda Sher, Sonja Hess, Keigo Machida. NANOG metabolically reprograms tumor-initiating stem-like cells in oxidative phosphorylation and fatty acid metabolism. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 3040. doi:10.1158/1538-7445.AM2015-3040
Abstract 3053: Knockout of UCP2 suppresses skin carcinogenesis
Authors: Sreedhar Annapoorna , Zhang Chunjing , Zhao Yunfeng
Published: August 2015

Mitochondria are important and necessary components of cell organelles. In recent times, mitochondrial metabolism has regained attention due to the growing evidence suggesting that dysfunction of mitochondria could play a major role in cancer.

Cancer cells that are actively proliferating have great demand for ATP. This huge demand for ATP is met by glycolysis, which is called the ‘Warburg Effect’.

Although the Warburg effect has lately been challenged, dysfunction of the mitochondrial dependent processes is a characteristic feature of almost all tumor cells, which lead to higher oxidative stress, inhibition of apoptosis and enhanced cell proliferation; which are the hallmarks of tumor cells.

Mitochondrial uncoupling proteins have gained enormous interest in the field of cancer metabolism due to their impact on mitochondrial dependent processes.

Uncoupling proteins are a family of mitochondrial proteins present in the inner mitochondrial membrane whose physiological role is to decrease membrane potential and hence ATP production. One of the most interesting functions of UCPs is to decrease the generation of ROS.

However, because of altered mitochondrial dependent processes, cancer cells have elevated levels of ROS production leading to a phenomenon called ‘oxidative stress’. To fight against this oxidative stress, UCP2 (a homologue of UCP family) has shown to be up-regulated in a number of human cancers. However, there is no direct evidence supporting UCP2 contributes to carcinogenesis.

Considering this ability of UCP2 to modulate ROS, membrane potential and mitochondrial bioenergetics, an interesting question to us, was whether UCP2 is involved in the progression of cancer.

Using UCP2 homozygous knockout mice and UCP2 overexpressed cells; our data have demonstrated that knockout of UCP2 suppressed skin carcinogenesis. Overexpression of UCP2 enhanced skin cell transformation and oncogenic activation. In addition, genipin, a known UCP2 inhibitor, was able to inhibit oncogenic activation during skin cell transformation, even in UCP2 overexpressed cells. Finally, although knockout of UCP2 increased the basal levels of protein oxidation, it did not enhance protein oxidation in mouse skin epidermal tissues during skin carcinogenesis.

In summary, our data have provided direct evidence that UCP2 promotes carcinogenesis, and targeting UCP2 may serve as a translational approach for chemoprevention.

Citation Format: Annapoorna Sreedhar, Chunjing Zhang, Yunfeng Zhao. Knockout of UCP2 suppresses skin carcinogenesis. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 3053. doi:10.1158/1538-7445.AM2015-3053
Abstract 3085: MicroRNA-186 inhibition alters cell proliferation and colony formation in prostate cancer
Authors: Jones Dominique Zilpha , Hobbing Katharine R. , Schmidt M. L. , Clark Geoffrey J. , Kidd LaCreis R.
Published: August 2015

MicroRNAs (miRs), key regulators of cancer-related genes, are dysregulated in many solid tumors, including prostate cancer. However, the role of miR-186 in prostate cancer (PCA) requires further elucidation. Previously, we observed mature miR-186-5p was significantly up-regulated in the serum collected from PCA patients (n = 15) diagnosed with aggressive PCA (stage III/IV) disease compared to disease-free men (n = 5). In addition, miR-186 expression was significantly up-regulated in metastatic PCA cells (PC3) compared to a normal prostate epithelial cell line (RWPE1). Putative and predicted miR-186 gene targets in various cancers include pro-apoptotic and tumor suppression-related genes (e.g., P2×7, FOXO1, AKAP12). We hypothesized that miR-186 inhibition will reduce the aggressive PCA phenotype.

To test this hypothesis, PC3 cells were transiently transfected with miR-186 inhibitor and scramble control for 24hrs. Cell viability and colony formation of transfected cells were evaluated via the ATPlite Luminescence assay and soft agar assay. We also evaluated expression profiles of predicted miR-186 gene targets in PCA cell lines and transiently transfected PC3 cells using qRT-PCR.

Transient inhibition of miR-186 repressed cellular proliferation and colony formation by 48-58% in 72-96hrs and 30% after 21 days, respectively. We observed a 1.6-2.24 fold up-regulation in ROCK1 gene expression in PC3 cells treated with miR-186 inhibitor. However, additional studies are needed to assess whether stable inhibition of miR-186 will increase PCA invasiveness by up-regulating ROCK1 using in vitro assays and murine models. Moreover, predicted miR-186 gene targets require validation using qRT-PCR, western blots and luciferase reporter assays. Such efforts may lead to the identification of novel biomarkers to improve diagnostic, prognostic and clinical management strategies.

Citation Format: Dominique Zilpha Jones, Katharine R. Hobbing, M. L. Schmidt, Geoffrey J. Clark, LaCreis R. Kidd. MicroRNA-186 inhibition alters cell proliferation and colony formation in prostate cancer. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 3085. doi:10.1158/1538-7445.AM2015-3085