bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 5106: Induction of reactive oxygen species-dependent mitotic arrest and apoptosis by sulforaphane in human bladder cancer 5637 cells
Authors: Choi Yung Hyun
Published: May 2015

The present study was undertaken to determine whether sulforaphane-derived reactive oxygen species (ROS) might cause growth arrest and apoptosis in human bladder cancer 5637 cells. Our results show that the reduced viability of 5637 cells by sulforaphane is due to mitotic arrest, but not the G2 phase. The sulforaphane-induced mitotic arrest correlated with an induction of cyclin B1 and phosphorylation of Cdk1, as well as a concomitant increased complex between cyclin B1 and Cdk1. Sulforaphane-induced apoptosis was associated with the activation of caspase-8 and -9, the initiators caspases of the extrinsic and intrinsic apoptotic pathways, respectively, and activation of effector caspase-3 and cleavage of PAPR. However, blockage of caspase activation inhibited apoptosis and abrogated growth inhibition in sulforaphane-treated 5637 cells. This study further investigated the roles of ROS with respect to mitotic arrest and the apoptotic effect of sulforaphane, and the maximum level of ROS accumulation was observed at 30 min after sulforaphane treatment. However, a ROS scavenger, N-acetyl L-cysteine, notably attenuated sulforaphane-mediated apoptosis as well as mitotic arrest. Overall, these results suggest that sulforaphane induces mitotic arrest and apoptosis of 5637 cells via a ROS-dependent pathway.

Citation Format: Yung Hyun Choi. Induction of reactive oxygen species-dependent mitotic arrest and apoptosis by sulforaphane in human bladder cancer 5637 cells. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 5106. doi:10.1158/1538-7445.AM2014-5106
Abstract 5111: Nitric oxide and its role in photodynamic therapy
Authors: Monroy Marco , Tipgunlakant Pooncharas , Simonis Ursula , Esquerra Raymond
Published: May 2015

Porphyrinic pigments are used as photosensitizers (PS) in photodynamic detection (PDD) and therapy (PDT) that is a minimally invasive modality in the fight against cancer. When the PS is activated by visible light at a given wavelength, reactive oxygen species (ROS) are generated, which cause cancer cells to undergo cell death. Despite significant advances, drawbacks of the PSs in clinical use include their non-selectivity in cellular-targeting causing cell death by necrosis leading to tissue inflammation. Nitric oxide (NO) has been shown to play a key role in modulating apoptotic cell death pathways and to react with reactive oxygen species to form additional lethal reactive nitrogen species (RNS). In our efforts to enhance the effectiveness of PDT, we set out to investigate the role of NO in PDT. We hypothesized that NO delivered to cancer cells at the time that the PS was administered would enhance the efficacy of PDT by promoting mitochondria-mediated apoptosis. To this end, we incubated androgen-sensitive human prostate adenocarcinoma (LNCaP) cells with both a PS and an NO releasing agent that was most effective in NO release as was spectrophotometrically determined by its oxidation of hemoglobin. Phototoxicity experiments were carried out at 37 OC with a noncoherent light source. Cell viability, damage and death were assessed in both illuminated and non-illuminated cells and were quantified by MTT staining as well as trypan blue and propidium iodide exclusion. To corroborate the cell viability results, we assayed clonogenic recovery in response to PDT in pigmented cells both in the presence and absence of NO. Our results indicate that the effectiveness of PDT in causing cell death depends on the NO concentration. PDT with NO alone was toxic to the cancer cells at high concentration of NO, whereas at low NO concentration no significant cell damage was observed after light illumination. PDT with the PS alone was not as effective in promoting cell death as PDT in the presence of both NO and the PS. Depending on the concentrations of the PS and NO, we observed that either necrosis or apoptosis were the prevailing modes of cell death after PDT. Our results indicate that the phototoxicity of the compounds is mainly determined by their intracellular concentration. Thus, understanding the combined effects of NO and the PSs in enhancing cell phototoxicity will aid in determining the roles that NO plays in improving the efficacy of PDT and may provide an alternate regimen to enhance PDT efficacy.

Citation Format: Marco Monroy, Pooncharas Tipgunlakant, Ursula Simonis, Raymond Esquerra. Nitric oxide and its role in photodynamic therapy. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 5111. doi:10.1158/1538-7445.AM2014-5111
Abstract 5117: Cytotoxic effects of hypericum extracts on breast and colorectal tumors
Authors: Gibbs Carla , Duggans Johnette S. , Malone Erin , Myles Elbert L.
Published: May 2015

The risk of acquiring certain types of cancers has many causes which include age, environment and genetics. There have been advances in the primary treatment options available, which include surgery, radiation therapy, and chemotherapy. However, the survival prognosis for many cancer patients remains low. Cancer treatments may involve chemotherapy, which often cause harsh side effects. The Plant kingdom may provide alternative therapy that may have less side effects and possibly more effective. Hypericum perforatum possesses hypericin, which has been been shown to have antitumor effects with invitro studies on specific cell lines. In other Hypericum species, hypericin makes up a smaller percentage of the extract. However, these species contain other phytochemical compounds that may also exert an antitumor effect. The objective of the present study was to assess the cytotoxic potential of H. perforatum cv. topas, H. olympicum, and H. calycinum on breast and colorectal adenocarcinoma cell lines. MCF7, SW480, and SW620 cells were treated with varying concentrations dosages of methanolic crude extracts from the plant species. Cytotoxicity was determine by measuring the reduction of Alamar Blue after a 24 hour exposure period of 10,000 cells in 96 well fluorescent plates. The reduction was determine using the Molecular Devices fluorescent spectrophotometer. At the lowest comparable concentrations, H. perforatum cv. topas had a greater reduction in tumor growth than MCF7 and SW40 cells lines. Whereas, H. olympicum demonstrated a greater decrease in SW620 cell growth at the lowest comparable concentration. These results suggest a difference in the phytochemical composition of these plants, which should be evaluated in order to determine what metabolite(s) are having the antitumor effect and their target.

Citation Format: Carla Gibbs, Johnette S. Duggans, Erin Malone, Elbert L. Myles. Cytotoxic effects of hypericum extracts on breast and colorectal tumors. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 5117. doi:10.1158/1538-7445.AM2014-5117
Abstract 5118: Decursin synergistically enhances TRAIL-induced apoptosis through CHOP dependent DR5 signaling in TRAIL resistant lung cancer cells
Authors: Kwang Kim Jae
Published: May 2015

Targeting the tumor necrosis factor-related apoptosis-inducing ligand (TRAIL), which has been clinically established for anticancer therapy, is the attractive strategy to specifically kill cancer cells through the death receptors. Nonetheless, its therapeutic potential is limited due to the resistance in cancer cells. Thus, in the present study, we evaluated the therapeutic potential of Decursin, a pyranocoumarin isolated from the Korean Angelica gigas root as a TRAIL sensitizer in a variety of resistant lung cancer cells. Co-treatment of Decursin with TRAIL synergistically induced apoptosis in TRAIL resistant lung cancer cells, A545, H1299, H596 and Calu-1. This synergistic effect through the induction of DR5 in Decursin treated cells was also confirmed by immunoblotting and immunoanalysis of endogeneous and surface expression of DR5, respectively. In contrast, in DR knockdown cells, combination treatment Decursin and TRAIL did not induce apoptosis at all. Furthermore, we found that DR5 induction by Decursin was mediated by ER-stress and its byproduct CHOP. Depletion of CHOP via siRNA transfection completely blocked Decusin induced apoptosis cell death in TRAIL resistant lung cancer cells. Taken together, induction of the DR5 via upregulation of CHOP and its inducer, ER stress, by Decursin leads to synergistically increase the TRAIL mediated cell death in TRAIL resistant lung cancer cells. Our results indicate that Decursin is a useful therapeutic strategy for overcoming TRAIL resistance in lung tumor treatment.

Key words: Decursin, DR-5, ER stress, CHOP, Trail resistant, Lung cancer

Conflict of Interests: The authors disclose no potential conflicts of interest.

Acknowledgment: This work was supported by the National Research Foundation of Korea (NRF) Grant funded by the Korea Government (MEST) (no. 2012-0005755).

Citation Format: Kim Jae Kwang. Decursin synergistically enhances TRAIL-induced apoptosis through CHOP dependent DR5 signaling in TRAIL resistant lung cancer cells. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 5118. doi:10.1158/1538-7445.AM2014-5118
Abstract 5121: Omega-3 polyunsaturated fatty acids induce apoptotic and autophagic cell death through inhibition of Akt/mTOR pathway in human oral cancer cells
Authors: Lim Kyu , Shin Soyeon , Hong Tae-Hwa , Jing Kaipeng , Jeong Soyeon , Kim Soyeon , Kweon Gi-Ryang , Park Seung-Kiel , Park Jong-Il
Published: May 2015

Omega-3 polyunsaturated fatty acids (ω3-PUFAs) have been found to possess anticancer properties in a variety of cancer cell lines and animal models, but their effect in oral cancer remains unclear. This study was designed to examine the impact of ω3-PUFAs on oral cancer and the molecular mechanism of their action using in vitro and in vivo models. Exposure SCC4 and SCC9 human oral cancer cells to the ω3-PUFA docosahexaenoic acid (DHA) reduced cell viability in a dose- and time-dependent manner. The increase in cell death was due in part to apoptosis, since cells incubated with DHA exhibited elevated levels of apoptotic markers including cleaved PARP expression, subG1 DNA portion and TUNEL-positive nuclei. DHA treatment also led to autophagic vesicle formation and an increase in autophagic flux, indicating the involvement of both apoptosis and autophagy in the cytotoxic effect of ω3-PUFAs on oral cancer cells. Further experiments revealed that the concurrent activation of apoptosis and autophagy induced by DHA was linked to reactive oxygen species (ROS) overproduction and inhibition of Akt/mTOR signaling molecules. Moreover, stable SCC9 cell lines expressing the ω3-desaturase gene (SCC9-fsc) or control vector (SCC9-csc) were generated, and the effect of endogenous high level of ω3-PUFAs was investigated. The growth rate and colony-forming capacity of SCC9-fsc were remarkably decreased compared to those of SCC9-csc. Likewise, tumor size and volume of SCC9-fsc implanted into nude mice were also significantly inhibited with increases in the cell death index. Taken together, these results indicate that ω3-PUFAs induce apoptosis- and autophagy-associated cell death through ROS and Akt/mTOR signaling pathways in oral cancer cells. Thus, utilization of ω3-PUFAs may represent a promising therapeutic approach for chemoprevention and treatment of human oral cancer. [This work was supported by the National Research Foundation of Korea (NRF) grant funded by the Korea government (MEST) (No. 2011-0013263 and 2007-0054932).]

Citation Format: Kyu Lim, Soyeon Shin, Tae-Hwa Hong, Kaipeng Jing, Soyeon Jeong, Soyeon Kim, Gi-Ryang Kweon, Seung-Kiel Park, Jong-Il Park. Omega-3 polyunsaturated fatty acids induce apoptotic and autophagic cell death through inhibition of Akt/mTOR pathway in human oral cancer cells. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 5121. doi:10.1158/1538-7445.AM2014-5121
Abstract 5127: Reprogramming of EMT and stemness by human recombinant CCN5 in triple negative breast cancer cells
Authors: Das Amlan , De Archana , Banerjee Snigdha , Banerjee Sushanta K.
Published: May 2015

Epithelial to Mesenchymal transition (EMT) is an important and coordinated series of events associated with tumor metastasis and invasion. Recent studies from our group had shown the importance of CCN5/ WISP-2 in the suppression of breast and pancreatic cancer progression through the regulation of the invasive phenotypes. In our study we have observed that exposure of triple negative human breast cancer cells (TNBC), MDA-MB-231 and HCC-70 to recombinant CCN5 (hrCCN5), resulted in a dose-dependent inhibition of cell-proliferation through the induction of apoptotic cell death. Anchorage-independent growth and colony forming ability of MDA-MB-231 cells were also found to be drastically inhibited in the presence of hrCCN5. Distinct up regulation of the epithelial markers such as CD24, and cytokeratin19 and down regulation of the mesenchymal markers such as Notch-1, vimentin, and CD44 were observed in CCN5 treated TNBC cells. CCN5 treatment also resulted in the significant reduction of stemness of these cells, as confirmed by mammosphere assay. Collectively, the studies indicate CCN5 could be a re-programmer of EMT and stemness and thus hrCCN5 can be considered as a potential therapeutic agent for TNBC.

Note: This abstract was not presented at the meeting.

Citation Format: Amlan Das, Archana De, Snigdha Banerjee, Sushanta K. Banerjee. Reprogramming of EMT and stemness by human recombinant CCN5 in triple negative breast cancer cells. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 5127. doi:10.1158/1538-7445.AM2014-5127
Abstract 5129: Role of PRKD2 in HSP90 inhibition-mediated suppression of cancer growth
Authors: Azoitei Ninel , Diepold Kristina , Genze Felicitas , Rouhi Arefeh , Brobovich Susanne , Froehling Stefan , Chiosis Gabriela , Brunner Cornelia , Lint Johan van , Cronauer Marcus , Scholl Claudia , Seufferlein Thomas
Published: May 2015

In various types of malignancies, conventional forms of therapy such as surgery, radiation and chemotherapy are often ineffective. In the past decade, a convergence of scientific and technological advances has enabled the identification of molecular targets and signaling pathways specific to cancer cells, resulting in therapies with enhanced selectivity and efficacy and reduced toxicity. Protein kinases represent such molecular targets. We have recently shown that PRKD2 is a crucial regulator of tumor cell - endothelial cell communication in gastrointestinal tumors and demonstrated the kinase to be a novel regulator of glioblastoma growth. Here we report that HSP90/Cdc37 chaperone complex binds to and stabilizes PRKD2 in human cancer cells. RNAi-mediated ablation of HSP90 expression but also the pharmacological inhibition of the chaperone with two independent inhibitors induced degradation of PRKD2 in a broad range of human cancer cell lines and in vivo. Treatment of various cancer cell lines with proteasome inhibitors such as MG-132 or Bortezomib, followed by incubation with HSP90 inhibitors rescued the PRKD2 levels to the detergent-insoluble fraction suggesting that degradation of the kinase follows the proteasomal pathway. Kinetic experiments conducted with chaperone inhibitors demonstrated that PRKD2 was degraded in a time- and dose-dependent manner and this degradation was associated with the activation of the apoptotic programme both in vitro and in vivo. Furthermore, ectopic expression of the kinase protected cancer cells from the apoptotic effects of HSP90 abrogation suggesting PRKD2 as a protein whose depletion mediates the sensitivity of tumor cells to chaperone inhibition. Altogether, these findings 1) propose PRKD2 ablation through HSP90 inhibition as a potential therapeutic strategy with immediate implementation, 2) reveal the kinase as a crucial modulator of HSP90-mediated apoptotic effects and 3) promote the intensification of designing specific PRKD2 inhibitors and their implementation in targeted therapy in human cancer.

Citation Format: Ninel Azoitei, Kristina Diepold, Felicitas Genze, Arefeh Rouhi, Susanne Brobovich, Stefan Froehling, Gabriela Chiosis, Cornelia Brunner, Johan van Lint, Marcus Cronauer, Claudia Scholl, Thomas Seufferlein. Role of PRKD2 in HSP90 inhibition-mediated suppression of cancer growth. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 5129. doi:10.1158/1538-7445.AM2014-5129
Abstract 513: Investigating stress-response pathways in pancreatic cancer cells using novel PERK inhibitors
Authors: Park Jihyun , Wang Qiantao , Kaoud Tamer S. , Tavares Clint D. J , Edupuganti Ramakrishna , Ren Pengyu , Dalby Kevin N.
Published: May 2015

An adaptive stress response mechanism, termed the unfolded protein response (UPR), is highly activated in a variety of tumor types to cope with ER stress and maintain protein homeostasis. PERK (PKR-like endoplasmic reticulum kinase) is one of the key components of the UPR. PERK is a Ser/Thr protein kinase whose active homodimer phosphorylates and inactivates eukaryotic initiation factor 2α (eIF2α), thereby inhibiting protein translation globally and reducing the load of newly synthesized proteins entering the ER. Previous studies demonstrated that cells with a compromised PERK signaling pathway are sensitive to hypoxic stress in vitro and form tumors that grow more slowly in vivo. Thus, selective inhibitors of PERK activity may be useful therapeutic candidates for the treatment of cancer. Using an HTS approach a number of lead compounds were identified with demonstrated ability to inhibit PERK kinase activity in vitro. The potent PERK inhibitors were further tested in pancreatic cancer cells to evaluate the modulation of the PERK-eIF2α-ATF4-CHOP signaling pathway. The effect of PERK inhibitors on cell viability was determined by an MTS assay. PERK inhibitors had dose-dependent effects on suppression of pancreatic cancer cell proliferation. Inhibitor-mediated PERK inhibition is associated with the down-regulation of PERK signaling and with the suppression of pancreatic cancer cell proliferation, suggesting their potential anticancer activities. Here we present new findings implicating additional inputs into this stress response.

Citation Format: Jihyun Park, Qiantao Wang, Tamer S. Kaoud, Clint D. J Tavares, Ramakrishna Edupuganti, Pengyu Ren, Kevin N. Dalby. Investigating stress-response pathways in pancreatic cancer cells using novel PERK inhibitors. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 513. doi:10.1158/1538-7445.AM2014-513
Abstract 5133: Identification of FC101-interacting proteins by immunoprecipitation
Authors: Washington Dominique C. , Williams Amber D. , Williams-Hart Tara
Published: May 2015

Fusarochromanone (FC 101) is a small toxic metabolite produced by Fusarium equiseti, a fungus found on decaying cereal plants and food grains. FC101 has been shown to exhibit anti-angiogenic and anti-cancer activity on various mammalian cells. Previous studies have revealed that FC101 inhibits growth of 35 human cancer cells and reduces mouse tumor size almost two-fold with increased caspase3 expression in FC101-treated tumors. cDNA microarray analysis and RT-qPCR were used to identify differentially expressed genes in human bladder cancer cell lines (UM-UC14) and Saccharomyces cerevisiae (budding yeast) exposed to different concentrations of FC101. These genes encode proteins that represent possible molecular targets of FC101. The objective of this study is to identify proteins that interact with FC101 by immunopercipitation using an FC101 antibody in UM-UC14 cells. We predict FC101 molecular targets will be verified in future animal studies and eventually FC101 may be used as a chemotherapeutic agent.

Citation Format: Dominique C. Washington, Amber D. Williams, Tara Williams-Hart. Identification of FC101-interacting proteins by immunoprecipitation. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 5133. doi:10.1158/1538-7445.AM2014-5133
Abstract 5135: A chromatin modifier from the 8p11 amplicon in luminal breast cancer
Authors: Ivey Brittany Turner , Guest Steve , Ethier Stephen P.
Published: May 2015

The chromosome 8p11-p12 amplicon is present in 12-15% of breast cancers, resulting in an increase in both copy number and expression of several chromatin modifiers in these tumors, including KAT6A, WHSC1L1, and ASH2L. Previous analyses in SUM52 breast cancer cells showed amplification and over-expression of KAT6A, and subsequent RNAi screening identified KAT6A as a potential driving oncogene. Also known as MYST3, KAT6A is a histone acetyltransferase (HAT) with intrinsic HAT activity towards itself and lysine residues on histone 2B, histone 3, and histone 4. Previously identified as a fusion partner with CREB binding protein (CREBBP) in acute myeloid leukemia, KAT6A is amplified in 8.7% of invasive breast cancers according to the Cancer Genome Atlas (TCGA). Knockdown of KAT6A with several shRNAs in SUM52 cells, a luminal breast cancer cell line harboring the 8p11 amplicon, resulted in reduced growth rate compared to non-silencing controls and profound loss of clonogenic capacity both in mono-layer and in soft agar. The normal cell line MCF10A, however, did not exhibit slower growth with knockdown of KAT6A, nor did other breast cancer cell lines without KAT6A amplfied. Interestingly, SUM52 cells with KAT6A knockdown formed fewer mammospheres in culture compared to non-silencing controls, suggesting a possible role for KAT6A in breast cancer stem cell activity and self-renewal. Further analysis of cell surface markers CD24 and CD44 in KAT6A knockdown cells revealed an increase in cells double positive for CD24 and CD44 expression, a characteristic of increased cell differentiation. Previous work from our laboratory identified FGFR2 as a driving oncogene in SUM52 cells. While FGFR inhibition can completely arrest the growth of these cells, it is completely reversible. Thus, exposure of SUM52 cells to FGFR inhibitors does not significantly affect the colony forming ability of the cells. By contrast, the colony forming efficiency of SUM52 KAT6A knockdown cells in the presence of the FGFR inhibitor PD170374 was significantly reduced compared to cells with KAT6A knockdown only. These data suggest that KAT6A may be a potential novel oncogene in breast cancers bearing the 8p11 amplicon and that it interacts with FGFR2 to regulate expression of transformed phenotypes. While there are other putative oncogenes in the amplicon, the identification of KAT6A as a driving oncogene suggests that chromatin-modifying enzymes are a key class of oncogenes in these cancers, and play an important role in the selection of this amplicon in luminal B breast cancers.

Citation Format: Brittany Turner Ivey, Steve Guest, Stephen P. Ethier. A chromatin modifier from the 8p11 amplicon in luminal breast cancer. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 5135. doi:10.1158/1538-7445.AM2014-5135
Abstract 5139: MOF cooperates with DNA polymerase eta to repair intrastrand break induced by crosslinking agents
Authors: Singh Mayank , Gupta Arun , Komal Komal , Pandita Tej K.
Published: May 2015

The human MOF gene encodes a protein that specifically acetylates histone H4 at lysine 16 (H4K16ac). Reduced levels of H4K16ac correlate with a defective DNA damage response (DDR) and double-strand break (DSB) repair post ionizing radiation exposure. MOF overexpression increases cell survival to intrastrand crosslink inducing agents like cisplatin. DNA polymerase eta (Pol H), the product of the xeroderma pigmentosum variant (XPV) gene and a Y-family DNA polymerase, plays a pivotal role in translesion DNA synthesis and was found to interact with MOF. Its interaction with MOF increases in a dose dependent manner after exposure to intrastrand DNA damage induced by cisplatin. Pol H colocalized with phosphorylated MOF and γ-H2AX foci in cisplatin treated cells. Overexpression of MOF enhances recruitment of Pol H to laser induced DNA damage. This correlated with monoubiquitination of PCNA which has been shown to facilitate the switch between the replicative DNA polymerase with the low-fidelity polymerase eta to bypass UV or cisplatin-induced DNA lesions during replication. MOF was found to interact with PCNA and regulating the ubiquitination of PCNA thereby facilitating Pol H recruitment to DNA damage sites.

Note: This abstract was not presented at the meeting.

Citation Format: Mayank Singh, Arun Gupta, Komal Komal, Tej K. Pandita. MOF cooperates with DNA polymerase eta to repair intrastrand break induced by crosslinking agents. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 5139. doi:10.1158/1538-7445.AM2014-5139
Abstract 514: Heat shock transcription factor-1 and heat shock protein 60 are important for lung carcinoma anoikis resistance
Authors: Carter Jack D. , Murphy Suzanne K.
Published: May 2015

Metastasis of cancer cells to distant sites leads to poor prognosis and comprises approximately 90% of cancer mortalities. It is evident that metastasis requires resistance to anoikis, an apoptotic program induced by detachment from the extracellular matrix. Heat shock proteins (HSPs) are molecular chaperones whose major functions include protein folding and suppression of apoptosis, and are transcriptionally regulated by heat shock transcription factor-1 (HSF1). HSF1 has been linked to metastasis and promotes anchorage-independent proliferation of breast cancer cells. HSP60 is a chaperone found in both the mitochondria and cytosol and is linked to metastasis and regulation of tumor cell apoptosis. To investigate whether HSF1 and HSP60 are necessary for anoikis resistance, anchorage-dependent lung carcinoma (A549, H460) and normal lung epithelial (Beas-2b) cells were suspended by plating on poly-2-hydroxyethyl-methacrylate-coated culture dishes. Anoikis induction was measured by executioner caspase-7 cleavage and PARP cleavage by western blot. Protein expression was measured by western blot. Small-interfering RNAs were used to inhibit hsf1 and hsp60. We show here that upon detachment for 24-48h, lung cancer cells resisted caspase-dependent anoikis in contrast to normal lung epithelial cells, with A549 cells showing the highest level of anoikis resistance. After 24h detachment there was increased phosphorylation of HSF1 on activating serine 326 in lung cancer cells. Antisense inhibition of hsf1 and hsp60 in attached lung cancer cells had minimal effects on caspase-7 and PARP cleavage. In contrast, hsf1 and hsp60 knockdown in cancer cells that were suspended for 24-48h initiated caspase-dependent anoikis. Furthermore, knockdown of hsf1 resulted in inhibition of HSP72, HSP40, and HSP27. HSP60 was not significantly inhibited with loss of HSF1. This data suggests that HSF1 and HSP60 are necessary for lung carcinoma anoikis resistance. Our work adds to evidence suggesting HSF1 and HSP60 are promising therapeutic targets for metastasis prevention. Although more detailed mechanistic evaluation is necessary, a therapeutic approach to induce anoikis in circulating tumor cells by modulating HSF1 or HSP60 may be promising.

Citation Format: Jack D. Carter, Suzanne K. Murphy. Heat shock transcription factor-1 and heat shock protein 60 are important for lung carcinoma anoikis resistance. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 514. doi:10.1158/1538-7445.AM2014-514
Abstract 5142: Probing the molecular mechanism of EZH2 in rhabdoid tumors
Authors: Lira Michelle , Chan Ho Man , Gibaja Veronica
Published: May 2015

Proteins that regulate chromatin dynamics and post-translational modifications of histones are crucial transcriptional regulators, and are implicated in diverse cellular processes including proliferation, differentiation and genome integrity control. The Polycomb repressive complex 2 (PRC2) plays a key role in the maintenance of transcriptional repression of genes necessary for normal embryonic development. Enhancer of Zeste homolog 2 (EZH2) is the catalytic subunit of PRC2 and methylates lysine 27 on histone 3 (H3K27).Overexpression and mutations of EZH2 have been reported to promote tumorigenesis [1,2]. Malignant Rhabdoid Tumors (MRTs) are highly aggressive tumors of soft tissues showing high sensitivity to an EZH2 inhibitor [1,3,4].Treatment with a highly selective and potent EZH2 inhibitor (EI1) leads to induction of p16 expression, sustained growth arrest and senescence in G401, a MRT cell line model (Fig. 1) (3, 4). Taking advantage of the strong anti-proliferative effect of EI1 in G401, we performed an shRNA screen to identify molecular targets that will confer resistance to EI1-mediated growth arrest. We hypothesized that such targets will provide novel insight of EZH2 biology in rhabdoid cancers, and may be relevant to potential mechanisms of resistance.

Citation Format: Michelle Lira, Ho Man Chan, Veronica Gibaja. Probing the molecular mechanism of EZH2 in rhabdoid tumors. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 5142. doi:10.1158/1538-7445.AM2014-5142
Abstract 5143: KDM3A promotes cell growth and cisplatin-resistance in ovarian cancer cells
Authors: Ramadoss Sivakumar , Sen Suvajit , Chaudhuri Gautam , Farias-Eisner Robin
Published: May 2015

Background: Ovarian cancer is the most lethal gynecological cancer affecting the women worldwide. One of the main challenging hurdles in ovarian cancer treatment is the high recurrence rate due to therapeutic resistance. Recent evidences showed that JmjC-domain (JMJD) containing histone demethylases plays a critical role in controlling gene expression during normal development and cancer progression.

Aims: To determine whether histone demethylase plays a role in ovarian cancer progression and therapeutic resistance.

Methods: We developed cisplatin-resistance ovarian cancer cell lines SKOV-3 and OVCAR-5 and isolated total RNA to screen the expression pattern of JMJD family genes by real-time RT-PCR. We performed stable knock-down of KDM3A by lentiviral mediated ShRNA transfection and studied its effects on cell proliferation and apoptosis by MTT assay and western blot analysis (PARP, caspase-3), respectively. We also determined the effect of KDM3A knock-down on cell cycle and apoptosis regulators by western blot.

Results: Our observations indicated the differential expression of various JMJD family proteins between cisplatin-resistance and sensitive cell lines. To further evaluate our results, we also tested the expression of those differentially expressed JMJD proteins in a widely used cisplatin-resistance and sensitive A2780 cell lines. We observed that KDM3A/JMJD1A is consistently overexpressed in all the cisplatin-resistance cell lines studied. Next, we stably knock-down KDM3A in cisplatin-resistance OVCAR-5 and A2780 cell lines. More interestingly, we found that KDM3A depletion inhibited cell proliferation, induced apoptosis and enhanced the sensitivity to cisplatin treatment. Further, we confirmed that the important regulators of cell cycle and apoptosis including p21 and Bcl-2 were modulated by KDM3A knock-down.

Conclusions: Our results indicate that KDM3A is a critical epigenetic factor required for the ovarian cancer growth by promoting cell proliferation and cisplatin-resistance. Our findings might have important clinical implications in targeting ovarian cancer cells by using specific histone demethylase inhibitors.

Grant support: This work was supported by Carl and Roberta Deutsch, and Kelly Day Foundations.

Citation Format: Sivakumar Ramadoss, Suvajit Sen, Gautam Chaudhuri, Robin Farias-Eisner. KDM3A promotes cell growth and cisplatin-resistance in ovarian cancer cells. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 5143. doi:10.1158/1538-7445.AM2014-5143
Abstract 5152: Mechanism and therapeutic potential of the histone demethylase GASC1 in castration-resistant prostate cancer
Authors: Labbe Roselyne M. , Ye Qin , Holowatyj Andreana , Zhang Lihong , Yang Zeng Quan
Published: May 2015

The histone lysine demethylase, GASC1 (Gene Amplified in Squamous Cell Carcinoma 1) is an epigenetic regulatory protein both highly expressed during mammalian embryogenesis and overexpressed in several types of cancer, including aggressive Castration Resistant Prostate Cancer (CRPC). GASC1 mainly catalyzes demethylation of tri- and di-methylated forms of histone H3 lysine 9 (H3K9me3/me2) epigenetic repressive marks, regulating gene expression and chromatin architecture. This research aims to elucidate the fundamental mechanism by which GASC1 dysregulation contributes to CRPC progression, as well as to evaluate its potential as a therapeutic target against aggressive prostate cancer. Previously, our lab and others have shown that GASC1 interacts with the androgen receptor (AR) and functions as a co-activator of AR-induced transcription in prostate cancer. We have used an shRNA approach to determine whether GASC1 knockdown affects the proliferation and transformation of CRPC cells. We found that knocking down GASC1 inhibits the growth and colony formation of C4-2B and CWR22Rv1 cells in vitro. Knockdown also reduced transcript levels of classical and CRPC-specific AR target genes in either the presence or absence of the AR ligand. As a result, we are evaluating the therapeutic potential of novel and highly specific lysine demethylase inhibitors towards abating GASC1 function in prostate cancer cells in vitro. We have recently determined that one of these novel inhibitors is highly effective in modulating the survival and proliferation of several prostate cancer cell lines including CRPC lines with IC50 values, all well under 1 uM. Together, our data demonstrates that GASC1 is a therapeutically relevant target in controlling and preventing the emergence of prostate cancers, with particular application against aggressive CRPC subtypes.

Citation Format: Roselyne M. Labbe, Qin Ye, Andreana Holowatyj, Lihong Zhang, Zeng Quan Yang. Mechanism and therapeutic potential of the histone demethylase GASC1 in castration-resistant prostate cancer. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 5152. doi:10.1158/1538-7445.AM2014-5152
Abstract 5156: The oncogenic polycomb histone methyltransferase EZH2 methylates lysine 120 on histone H2B and competes ubiquitination in human cancer
Authors: Hamamoto Ryuji , Nakamura Yusuke
Published: May 2015

The histone methyltransferase EZH2 is known to be a polycomb protein homologus to Drosophila enhancer of zeste and catalyzes the addition of methyl groups to histone H3 at lysine 27 (H3K27). We previously reported that EZH2 was overexpressed in various types of cancer and plays a crucial role in the cell cycle regulation of cancer cells. In this study, we demonstrated that EZH2 has the function to mono-methylated lysine 120 on histone H2B (H2BK120). EZH2-dependent H2BK120 methylation in cancer cells was confirmed with an H2BK120 methylation-specific antibody. Overexpression of EZH2 significantly attenuated the ubiquitination of H2BK120, a key post-translational modification of histones for transcriptional regulation. Concordantly, knockdown of EZH2 increased the ubiquitination level of H2BK120, suggesting that the methylation of H2BK120 by EZH2 may competitively inhibit the ubiquitination of H2BK120. Subsequent ChIP-Seq and microarray analyses identified downstream candidate genes regulated by EZH2 through the methylation of H2BK120. This is the first report to describe a novel substrate of EZH2, H2BK120, unveiling a new aspect of EZH2 functions in human carcinogenesis.

Citation Format: Ryuji Hamamoto, Yusuke Nakamura. The oncogenic polycomb histone methyltransferase EZH2 methylates lysine 120 on histone H2B and competes ubiquitination in human cancer. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 5156. doi:10.1158/1538-7445.AM2014-5156
Abstract 517: MiR-29b is a novel potential prognostic marker and inhibits cell proliferation in colorectal cancer
Authors: Inoue Akira , Yamamoto Hirofumi , Uemura Mamoru , Nishimura Junichi , Hata Taishi , Takemasa Ichiro , Mizushima Tsunekazu , Doki Yuichiro , Mori Masaki
Published: May 2015

Introduction Accumulating evidence indicates that microRNAs (miRNAs) are aberrantly expressed in human cancers and contribute to the tumorigenesis. Recent studies have suggested that microRNA-29 (miR-29) family members in human may play an important role in many types of cancer by regulating cell proliferation, differentiation, apoptosis, migration and invasion. MiR-29 is considered good candidate for miRNA-based therapeutics. In colorectal cancer (CRC), however, few studies have shown the clinical significance and biological function of miR-29. The aim of the present study was to examine the prognostic impact of miR-29b and the biological significance of miR-29b in CRC.

Materials and methods Expression of miR-29b was quantified by quantitative real-time PCR in 127 CRC patients who underwent curative resection. The impact of miR-29b expression on disease free survival (DFS) and overall survival (OS) was assessed in these 127 CRC patients. To investigate the functional role of miR-29b in vitro, tumor suppressive activity of miR-29b was assessed by proliferation assay, TUNEL assay and Ki-67 staining in CRC-derived cell lines (DLD1, HT29 and SW480).

Results MiR-29b expression was significantly decreased in tumor tissues compared to corresponding normal tissue (P = 0.014). The DFS and OS were significantly longer in the high miR-29b expression group compared to those in the low miR-29b expression group (P < 0.05). Univariate and multivariate analyses indicated that expression of miR-29b was an independent prognostic factor for OS (P = 0.003), together with lymph node metastasis (P = 0.009), and serosal invasion (P = 0.002). In vitro assays revealed that forced expression of miR-29b was associated with a reduction of proliferative activity. TUNEL assay and Ki-67 staining revealed that miR-29b could induce apoptosis and cell cycle arrest in CRC cell lines.

Conclusions Our findings suggested that miR-29b is a useful prognostic marker for CRC, and could be a promising therapeutic target against CRC.

Citation Format: Akira Inoue, Hirofumi Yamamoto, Mamoru Uemura, Junichi Nishimura, Taishi Hata, Ichiro Takemasa, Tsunekazu Mizushima, Yuichiro Doki, Masaki Mori. MiR-29b is a novel potential prognostic marker and inhibits cell proliferation in colorectal cancer. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 517. doi:10.1158/1538-7445.AM2014-517
Abstract 5171: An integrated analysis of copy number alteration and global gene expression reveals potential oncogenes underlying stomach cancer
Authors: Song Kyoung , Kim Ryong Nam , Jeon Sinyoung , Kim Hae I. , Choi Yoon-La , Shin Young Kee
Published: May 2015

Although several studies had been carried out to elucidate causative copy number alteration (CNA) underlying stomach carcinogenesis, deeper understanding of CNA accumulations in stomach cancer genome in association with age and histo-pathological subtypes remains still mysterious. Here, we report an integrated analysis of genome-wide copy number alteration and global mRNA expression of 40 stomach cancer tissues from 19 old and 21 young patients using aCGH and microarray. We identified that stomach cancer genomes showed recurrent amplifications at 17q12 (ERBB2 amplicon) and 7q31.1-7q31.31 (MET amplicon), and homozygous deletions at 9p21.3 and 17p12 regions harboring tumor suppressor genes. The most frequent gained regions were 3q26.2 (45%), 7p21.1 (>50%), 7p12.2 (∼50%), 8q21.11-8q21.12 (45%), 8q22.3 (45%), 8q24.21 (45%), 8q24.22 (45%), 8q24.3 (45%), 20q12 (45%) and 20q13.2 (45%). The most frequent deleted regions were 21p11.1 (∼50%), 19p13.2 (45%), 19p13.3 (45%), 4q12 (40%), 3p21.31 (40%) and 1p35.3 (40%). Whereas young stomach cancer genomes showed more frequent gains at 7q21-7q31 than old cancer genomes, old stomach cancer genomes exhibited more recurrent gains at 8q12-8q24 and 1p12-1q42 than young cancer genomes, providing a clue about evolution of CNA accumulation in stomach cancer genome. Hierarchical expression clustering of 506 (in amplified regions) and 60 (in regions showing homozygous deletion) genes across the 40 stomach cancer samples reclassified pathological and histological subtypes based on expression pattern. Our integrated analysis of aCGH and microarray gene expression identified a pool of potential candidate stomach oncogenes and tumor suppressor genes. SiRNA-based interference of genes in MET amplicon and ERBB2 amplicon in stomach cancer cell lines significantly suppressed cell proliferation and migration, suggesting that they could be novel potential oncogenes in stomach cancer. Taken together, this study improves our understanding of stomach cancer genome evolution during CNA accumulation process and provides a new gene set of novel potential oncogenes and tumor suppressor genes that are essential for developing novel diagnostic biomarker and therapeutics for stomach cancer.

Citation Format: Kyoung Song, Ryong Nam Kim, Sinyoung Jeon, Hae In Kim, Yoon-La Choi, Young Kee Shin. An integrated analysis of copy number alteration and global gene expression reveals potential oncogenes underlying stomach cancer. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 5171. doi:10.1158/1538-7445.AM2014-5171
Abstract 5174: Toward understanding the genomic alterations in human gastric cancer
Authors: Cui Juan , Xu Ying
Published: May 2015

Gastric cancer is one of the most prevalent and aggressive cancers worldwide, and its molecular mechanism remains largely elusive. Although recent genome sequencing studies have uncovered rich information on cancer evolutions of many types, little is known about the genetic basis of gastric cancer due to its complexity. Here we report genomic mutations in primary gastric adenocarcinoma of human, based on the whole genome sequences of five pairs of cancer and matching normal samples. In total, 407 non-synonymous somatic point mutations have been identified in these genomes and the most recurrent mutations were harbored by genes such as Mucins (MUC3A and MUC12) and transcription factors (ZNF717, ZNF595 and TP53). 679 genomic rearrangements were detected which affect 355 protein-coding genes; and 76 genes show copy number changes. The focus of the functional analysis is to validate the biological relevance of each variant to cancer formation and progression, particularly to explore the to-be-determined causal relationships between the observed mutations in cancer genomes and cancer development. Additionally, we have attempted to investigate the mechanism of how these variants form in each tumor and discuss about multiple possible factors that may affect the susceptibility to each specific chromosomal changes. For examples, structural correlation between two boundaries of most rearrangements was observed based on the Hi-C data analysis, which indicate these rearrangements may happen among DNA fragments in close spatial proximity, especially when two endpoints stay in a similar replication phase. Sequence analysis on the flacking region of those rearrangements also showed clear evidences that about half of these genomic changes may be involved in the mechanism of microhomology-mediated break induced replication (MMBIR). Furthermore, DNA integrations from Helicobacter pylori have been detected for the first time, for instance, in the gene region of PREX2, which may reflect a new mechanism about how H. pylori induce cancer development. Overall, the in-depth analyses on these sequenced genomes lead to the detection of a myriad of novel genomic variations in gastric cancer, which in turn bring new insights in our understanding of the genetic basis and indicate new molecular mechanism of the gastric tumorigenesis.

Citation Format: Juan Cui, Ying Xu. Toward understanding the genomic alterations in human gastric cancer. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 5174. doi:10.1158/1538-7445.AM2014-5174