bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 1968: Nanoparticle mediated measurement of target-drug binding in cancer cells
Authors: Ullal Adeeti V. , Reiner Thomas , Yang Katherine S. , Gorbatov Rostic , Min Changwook , Issadore David , Lee Hakho , Weissleder Ralph
Published: August 2012

Responses to molecularly targeted therapies can be highly variable and depend on mutations, fluctuations in target protein levels in individual cells and drug delivery. The ability to rapidly quantitate drug response in cells harvested from patients in a point-of-care setting would have far reaching implications. Capitalizing on recent developments with miniaturized diagnostic NMR (DMR) technologies, we have developed a magnetic nanoparticle based approach to directly measure both target expression and drug binding in freshly harvested human cancer cells. The method involves covalent conjugation of a small molecule drug to a magnetic nanoparticle that is then used as a read-out for target expression and drug binding affinity. Using poly(ADP-ribose) polymerase (PARP) inhibition as a model system, we developed an approach to distinguish differential expression of PARP across various cell lines with excellent correlation of DMR measurements to gold standards such as flow cytometry (r2 = 0.97) and western blotting (r2 = 0.92). We also sought to mimic drug pharmacodynamics ex-vivo through competitive target-drug binding and could quantify the relative binding affinities of several PARP inhibitors (e.g. Olaparib, Velaparib, AG-014699, and 3-aminobenzamide) in whole cells. Finally, we demonstrate the potential to perform such measurements directly in clinical samples. Further applications of the assay could result in a drug development platform to probe drug binding in cells, the identification of resistant cancer cells, and the ability to determine whether a drug has reached its target.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 1968. doi:1538-7445.AM2012-1968
Abstract 1970: Liposomal nanoparticle siRNA delivery systems for in vivo silencing of the androgen receptor in human prostate cancer
Authors: Zhang Kevin X. , Lee Justin B. , Tam Yuen Yi C. , Tam Ying K. , Cullis Pieter R. , Rennie Paul S.
Published: August 2012

Prostate cancer is the most commonly diagnosed non-skin cancer in men and one of the leading causes of cancer-death. While frequently curable by surgical or radiation ablation, recurrent or metastatic disease is usually managed with androgen withdrawal therapies (medical castration), which block the growth promoting effects of androgens on the androgen receptor (AR). Unfortunately, the cancers eventually progress to the lethal castration-resistant state. Previously, we reported that knocking down the AR by RNA interference using small hairpin RNA (shRNA) inhibits this progression and, in about 50% of tumours, causes complete regression. In this study, we investigated the ability of small interfering RNA (siRNA) encapsulated in lipid nanoparticle (LNP) to silence AR in human prostate cancer cell lines and xenograft tumours following intravenous injections. In vitro screening studies with prostate cancer cell lines using a panel of cationic lipids demonstrated that siRNA formulated in LNPs containing the ionizable cationic lipid 2,2-dilinoleyl-4-(2-dimethylaminoethyl)-[1,3]-dioxolane (DLin-KC2-DMA) exhibited the most potent AR silencing effects. This is attributed to an optimized ability of DLin-KC2-DMA-containing LNP to enter into cells and to release the siRNA into the cell cytoplasm following endocytotic uptake. DLin-KC2-DMA LNPs were also effective in silencing the AR in a wild-type AR expressing cell line, LAPC-4, and a variant AR expressing cell line, CWR22Rv1. Importantly, we found that LNP AR-siRNA systems containing DLin-KC2-DMA could silence AR gene expression in distal LNCaP xenograft tumors and decrease serum PSA levels following intravenous injection. The 5′RACE experiments further confirmed that siRNA-induced specific cleavage of AR mRNA was the primary mechanism of action. In conclusion, these results indicate that in vivo delivery of siRNA to knockdown AR using our LNP formulations causes a decrease in serum PSA and provides proof in principle that this approach may offer a therapeutic strategy for treating advanced prostate cancer. (Funded by Canadian Institutes for Health Research and a PCF-STAR Project from Prostate Cancer Canada with the support of Safeway Canada)

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 1970. doi:1538-7445.AM2012-1970
Abstract 1972: Active targeting, fluorescence imaging, and NIR photothermal therapy of malignant tumors
Authors: Green Hadiyah-Nicole , Grizzle William E. , Rosenthal Eben L. , Mirov Sergey B. , McNally Lacey R.
Published: August 2012

We combine active targeting, fluorescent imaging, and subsequent photothermal treatment of malignant cells as a viable approach for the treatment of a variety of cancer types that over-express the epidermal growth factor including head and neck, colorectal, ovarian, cervical, breast, bladder, pancreatic, and prostate cancers. In our photothermal therapy experiments, the use of a near infrared (NIR) laser enables us to take advantage of both the plasmon resonance absorption peak of gold nanorods (GNR), ∼785 nm with aspect ratio of ∼ 4, as well as the absorption peak of the fluorescent dye, ∼774 nm. We used a 785 nm NIR laser with fluence of ∼9.5 W/cm2 and a 4 minute exposure time. Anti- epidermal growth factor receptor (EGFR) antibody, which targets the epidermal growth factor receptor over-expressed on malignant head and neck cancer cells, was labeled with a fluorescent dye and conjugated to GNR. We have studied the conjugation parameters for GNR and anti-EGFR antibodies and used nearest neighbor approximations to analyze the nanorod-to-antibody ratio from transmission electron microscope images. The optimal conjugation of dye:antibody:GNR facilitates active targeting of the nanorods to the tumor site. This active targeting is more efficient than previously reported results that rely solely on the enhanced permeability and retention effect. We demonstrate that PEGylating the GNR before conjugation to antibodies improves the quality of the conjugation. We also show that binding ability of the conjugated antibody to the epidermal growth factor receptor has not been affected. In addition, cell death by NIR photothermal therapy and gold nanorods is not compromised following conjugation of gold nanorods to a fluorescently labeled antibody. NIR photothermal treatment with the dye-antibody-nanorod conjugate selectively heated the GNR, excited fluorescence, and was sufficient to kill nearly 90% of tumor cells comparable to photothermal therapy with the GNR alone.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 1972. doi:1538-7445.AM2012-1972
Abstract 1991: Effects of isoliquiritigenin (ISL) on VEGF secretion in Human breast cancer cell line MDA-MB-231
Authors: Hsia Shih-Min , Shieh Tzong-Ming , Shih Yin-Hua
Published: August 2012

Breast cancer is the most frequent cause of mortality among females in the world. The death reasons of most breast cancer patients were due to the tumor metastasis from breast to other organs. The principal mechanisms involved in metastasis are migration and invasion. Unfortunately, there are almost no efficacious therapeutic modalities. Thus, it is imperative to develop novel targets and agents to prevent invasion and metastasis of breast cancer cells.This purpose of this study is to investigate the anti-metastatic potency of ISL via modulation of the expression of VEGF and MMP in the highly metastatic human breast cancer cell line MDA-MB-231. Based on ELISA and western blot analysis, ISL treatment (0.1-10 μM) for 24 h and 48 h reduced secretions and protein levels of VEGF in a time- and dose-dependent manner. ISL decreased the hypoxia-inducible factor 1-alpha (HIF-1α) protein expression, which is the up-regulator of VEGF. According to chamber migration assay and wound migration assay, ISL significantly suppressed the migration of MDA-MB-231 cells at 24 h and 48 h. Among human MMPs, MMP-2 and MMP-9 have been of particular interest because they efficiently cleave the major components of ECM. As confirmed by western blot and gelatin zymography assay, ISL inhibited the protein expression and gelatinolytic activity of MMP-2/-9 at 48 h. To further clarify the signal pathway responsible for ISL-downregulated cell migration in MDA-MB-231 cells, we examine the effect of ISL on PI3K/Akt pathway and MAPK pathway, as well as the NF-κB. The expression of PI3K and phosphorylation of p38 mitogen-activated protein kinase and Akt kinase were suppressed after ISL treatment. In addition, NF-κB DNA binding assay demonstrated that NF-κB DNA binding activity was inhibited after ISL treatment. Taken together, these findings suggest the ISL inhibits p38, PI3K/Akt, and NF-κB signaling, resulting in the inhibition of VEGF, HIF-1α, MMP-9, and MMP-2, and thus suppresses migration of MDA-MB-231 cells. We believe that these findings will provide the potential application of ISL on the treatment of metastatic breast cancer.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 1991. doi:1538-7445.AM2012-1991
Abstract 1992: Ganoderma lucidum (Reishi) induces autophagy in inflammatory breast cancer by regulation of the mTOR signaling pathway
Authors: Rosario-Acevedo Raysa , Suárez-Arroyo Ivette , Aguilar-Pérez Alexandra , Cubano Luis A. , Martínez-Montemayor Michelle M.
Published: August 2012

The control of cell growth by the mammalian target of rapamycin (mTOR) signaling pathway and autophagy are relevant to disease, as altered regulation of either pathway may result in tumorigenesis. Autophagy is a regulated pathway involving the lysosomal degradation of cytoplasmic components and organelles. Activation of the human autophagy-initiation kinase (ULK1) is directly controlled by mTOR and is a critical part of the autophagy machinery. In this study we test the autophagy inducing effects of the medicinal mushroom Ganoderma lucidum (Reishi) in the patient derived inflammatory breast cancer (IBC) cell line SUM-149 and in the non-cancerous mammary epithelial cell line MCF10A. IBC is a rare, lethal and aggressive form of locally advanced breast cancer that has a poor prognosis and no targeted therapy. Herein, we demonstrate that Reishi modulates the expression of autophagy related molecules depending on treatment time. At 2h post-Reishi treatment SUM-149 cells show that ULK1, Atg5, Atg12, Beclin1 and the microtubule-associated protein 1 light chain 3 (LC3) are α1.2 fold upregulated. Furthermore, autophagosomal structures were identified in SUM-149 but not in MCF10A Reishi treated cells. At 4h post-Reishi Atg5 and Atg12 (required for phagophore formation) are ∼1.5 fold upregulated, while at 6h LC3 expression (involved in autophagosome formation) is ∼2 fold upregulated compared to vehicle. We also show that autophagy induction in SUM-149 cells is mediated via downregulation of the PI3K/mTOR signaling pathway. Reishi downregulates PI3K (p85 and p110), mTOR phosphorylation and reduces the expression of Raptor, which is the regulatory subunit of mTORC1, the complex that directly regulates proteins involved in autophagy. Rictor, the regulatory subunit of the mTORC2 complex, which inhibits autophagy indirectly through Akt is also downregulated by Reishi. We demonstrate that Reishi has great potential to be used as an anti-IBC-therapeutic. This project was sponsored by Title V PPOHA grant number P031M105050 from the US Dept of Education to UCC, and NIH/RCMI 2G12RR003035 to UCC.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 1992. doi:1538-7445.AM2012-1992
Abstract 1996: Anticancer efficacy and mechanism of action of novel ginsenoside 25-OCH3-PPD, a natural product MDM2 inhibitor
Authors: Wang Wei , Wang Hui , Zhang Ruiwen
Published: August 2012

Ginseng and related herbs have been used for various health benefits in over 2000 years. However, their mechanisms of action are not fully understood. In our laboratory, we have been isolating and characterizing ginsenosides as anticancer agents. 20(S)-25-methoxyl-dammarane-3α, 12α, 20-triol (25-OCH3-PPD), a newly identified natural product from Panax notoginseng, has shown activities against a variety of cancer cells in vitro and in vivo. However, the mechanisms of action for the novel anticancer agent are largely unknown. In this study, we discovered a novel mode of action for 25-OCH3-PPD: direct inhibition of the MDM2 oncoprotein. It is well established that the MDM2 oncogene plays a major role in human cancer development and progression. Its tumorigenic properties are associated with both p53-dependent and -independent pathways. We first demonstrated that, in a dose-dependent manner, 25-OCH3-PPD inhibited MDM2 expression in human breast cancer cell lines with different p53 backgrounds, including MCF-7 (p53 wild type and p53 knockout) and MDA-MB-468 (p53 mutate) cells. We further found that 25-OCH3-PPD inhibited MDM2 expression at the transcriptional level, affecting MDM2 promoter activity. We also observed that 25-OCH3-PPD decreased the survival, inhibited proliferation, induced apoptosis and led to G1 cell cycle arrest in the test breast cancer cells. The anti-cancer activities of 25-OCH3-PPD were reduced in MCF7 inducible MDM2-overexpression cells, further supporting the relevance of the MDM2 inhibitory effect to its anti-cancer activity. Moreover, 25-OCH3-PPD inhibited the growth of breast cancer xenograft tumors in vivo which was associated with its MDM2 inhibitory effects. 25-OCH3-PPD demonstrated low toxicity to non-cancer cells, and did not have any observable toxicity in animals. In summary, our preclinical data indicate that 25-OCH3-PPD is a potential therapeutic agent and that further preclinical and clinical development of this agent is warranted. These results also provide an example of a natural product/oncogene interaction as a novel mechanism of action for chemotherapeutic agents. (This work was supported by NIH/NCI grants R01 CA112029 and R01 CA121211 and a grant (BCTR070731) from Susan G Komen for the Cure.)

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 1996. doi:1538-7445.AM2012-1996
Abstract 2: FGFR3 as a therapeutic target in squamous cell lung carcinoma
Authors: Liao Rachel , Hammerman Peter , Sivachenko Andrey , Greulich Heidi , Getz Gad , Liu Qingsong , Gray Nathanael , Meyerson Matthew
Published: August 2012

Squamous cell lung carcinoma (lung SCC) is the second most common subtype of non-small cell lung cancer with, 40,000 new cases diagnosed every year in the United States. Unlike lung adenocarcinoma, few targetable genomic events are known drivers of lung SCC, and therefore, therapeutic options are limited for patients with this disease. As part of The Cancer Genome Atlas (TCGA) project, we have analyzed the entire coding sequence of 196 lung SCCs for mutated genes which may be amenable to targeted therapeutics. In the 108 patient samples analyzed to date, we have observed mutations in the Fibroblast Growth Factor Receptor 3 (FGFR3) kinase gene in six cases at four sites, or 3% of samples. The observed FGFR3 mutations are present in both the extracellular domain and the kinase domain of the protein, an observation that is consistent with activating mutations in FGFR3 that are known to drive bladder cancer, and in fact, two of the observed mutations are at sites previously observed in bladder cancer. We therefore hypothesized that these mutations are oncogenic and may be driving transformation in a subset of lung SCC patients. Expression of mutated FGFR3 in NIH-3T3 cells led to anchorage-independent colony formation, which was inhibited by treatment with a pan-FGFR inhibitor. A known mechanism of FGFR3 activation is the constitutive dimerization of receptors with mutated extracellular domains via intermolecular disulfide bond formation, and we have observed this phenomenon in our ECD mutants as well. These data suggest that mutations in the FGFR3 gene are sufficient to transform cells. This finding, along with recently published data suggesting that FGFR1 amplification is also able to drive lung SCC development, demonstrates that the FGFR family may be a promising therapeutic target in lung SCC.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 2. doi:1538-7445.AM2012-2
Abstract 2010: Development of a cytotoxic peptide based on the C-terminal domain of Bax
Authors: Boohaker Rebecca , Zhang Ge , Nemec Kathleen , Khaled Annette R.
Published: August 2012

The Bcl-2 family proteins maintain the balance of life and death in a cell. Bax, a pro-apoptotic member, translocates to the mitochondrial membrane in response to diverse death stimuli, initiating a cascade of programmed events that lead to cell death. Tumor progression involves deregulation of this process, in part, through mutations that cause aberrant expression of anti-apoptotic members of the Bcl-2 family that normally sequester and inhibit Bax. This study seeks to identify functional domains of Bax that can be developed into therapeutic peptides to promote the death of cancer cells resistant to existing chemotherapeutics. Our goal was to generate a peptide, much like antimicrobial peptides, that could target and disrupt mitochondrial membranes. To this end, we discovered that the C-terminal, alpha-9 helix of Bax (CT20), when tagged to EGFP or a destabilization domain (DD), could bind mitochondria and cause cell death, mimicking the activity of full-length Bax. Cell death was measured by morphological changes, such as membrane blebbing, as well as DNA fragmentation. Targeted mutagenesis of two adjacent lysines at the carboxyl end of the CT20 peptide demonstrated that these residues enabled mitochondrial membrane association. Expression of the CT20 peptide in the presence or absence of endogenous Bax resulted in cytotoxicity, indicating that the death mechanism engaged could function independently of the Bcl-2 family. These findings suggest the CT20 peptide has the potential to be developed into a viable therapeutic agent in the treatment of a broad range of cancer types.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 2010. doi:1538-7445.AM2012-2010
Abstract 2011: A novel bcl-2/xL inhibitor induces tumor cell apoptosis and potentiates the activity of standard chemotherapeutics
Authors: MacIntyre Terry , Powell Francoise , Lewis Paula , Adam Ammar , Lawson Deborah , Wu Jiaquan , Larsen Nicholas , Repik Galina , Parmentier Julie , Secrist Paul , Ioaniddis Stephanos , Byth Kate
Published: August 2012

Tumor cells often evade apoptosis by tipping the balance of Bcl-2 family and BH3-only protein expression towards a more anti-apoptotic environment, thus providing a survival advantage as well as a means of resistance to anti-cancer therapies. Indeed, over-expression of Bcl-2 family members has been shown to confer chemo-resistance in multiple tumor types. One potential result of this survival adaptation, however, is that the tumor cells become dependent on said Bcl-2 family member and hence increasingly vulnerable to targeted pro-apoptotic BH3 mimetics. Clinical validation of this concept has been demonstrated with the Bcl-2/xL inhibitor Navitoclax, where as a single agent in phase I/II trials in relapsed, refractory CLL patients, a 20% response rate was observed. Due to the potential that directly targeting the apoptotic machinery holds in treating cancer, developing BH3 mimetics that easily and effectively combine with standard chemotherapies is an attractive proposition. With this in mind, we have discovered and characterized a novel small molecule BH3 mimetic with nanomolar affinity for Bcl-2 and Bcl-xL. Here we characterize the in vitro and in vivo activity of this inhibitor in a variety of cell types. In addition to exhibiting potent single agent anti-cancer activity, it also potentiates the effectiveness of standard chemotherapeutic agents. Importantly, it also induces regression of sensitive tumors in vivo after a single IV bolus dose. Overall, this novel Bcl-2/xL inhibitor represents an exciting tool to further explore the mechanistic rationale for using potent BH3 mimetics as a foundation therapy across a broad range of tumor types where resistance to standard therapies arises from deregulation of the Bcl-2 family of anti-apoptotic proteins.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 2011. doi:1538-7445.AM2012-2011
Abstract 2014: IRF8 regulates Bax transcription in primary myeloid cells differentiation and pathogenesis in CML
Authors: Hu Xiaolin , Yang Jine , Liu Kebin
Published: August 2012

Chronic myelogeneous leukemia (CML) is a myeloid proliferative disease featured in a chronic stage of clonal expansion of undifferentiated myeloid cells followed by an accelerated stage with uncontrolled clonal expansion of blastocytes that quickly progresses to blast crisis. IFN regulatory factor 8 (IRF8) is a key transcription factor for myeloid cell differentiation and its expression is frequently lost in hematopoietic cells of human myeloid leukemia patients. IRF8-deficient mice exhibit similar symptoms to human CML patients with accumulation of undifferentiated myeloid cells in the bone marrow and periphery blood that can progress to a fatal blast crisis, thus IRF8 is a myeloid leukemia suppressor. Although the role of IRF8 function in CML has highly appreciated, the molecular mechanisms underlying IRF8 function in CML are still largely unknown. In this study, we use IRF8 knockout mice as proof of study. We observed that Bax expression level is low in bone marrow progenitor cells and increases dramatically in primary myeloid cells in wt mice. In contrast, Bax expression level remained at a low level in primary myeloid cells in IRF8 KO mice. However, in vitro IRF8 KO bone marrow-differentiated myeloid cells expressed Bax at a level as high as that in wild type myeloid cells. Furthermore, we demonstrated that IRF8 specifically binds to the Bax promoter region in primary myeloid cells. Functional analysis showed that IRF8 deficiency results in increased resistance of the primary myeloid cells to Fas-mediated apoptosis, which indicates that CML cells could use the strategy of silencing IRF8 to downregulate pro-apoptotic gene Bax to acquire apoptosis resistance. To summarize, our findings show that IRF8 directly regulates Bax transcription in vivo, but not in vitro during myeloid cell lineage differentiation and silencing IRF8 to disrupt apoptosis related genes expression could result in uncontrolled clonal expansion of myeloid cells and leukemia development.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 2014. doi:1538-7445.AM2012-2014
Abstract 2015: Lysophosphatidylcholine induces apoptosis via suppressing STAT3 and HDAC3 activities and binding to the Bcl-xL promoter in K562 chronic myelogenous leukemia cells
Authors: hoon Jung Ji , Jeong Soo-Jin , Kim Ji-Hyun , Lee Hyo-Jung , Lee Hyo-Jeong , Lee Eun-Ok , Kim Junghyo , Kim Sung-Hoon
Published: August 2012

Lysophosphatidylcholine (LPC) is a class of chemical compound which are derived from phosphatidylcholines usually isolated from the surface of wheat starch granules. In the present study, we investigated the molecular mechanisms responsible for LPC-induced apoptosis in chronic myeloid leukemia K562 cells. LPC exerted the inhibitory effect on the viability of K562 cells as well as other leukemic cell lines HL-60 and U937. Nonetheless, LPC significantly increased the acetylation of histone H3 at K18, 9 and 23 in K562 cells, but not in HL-60 and U937 cells. Consistently, LPC reduced histone deacetylase (HDAC) transcriptional activity and the expression of HDAC3 at protein level. LPC suppressed phosphorylation of mTOR and AKT, the upstream kinases of HDACs and induced p27 expression. We also found that LPC mediated inactivation of signal transducer and activator of transcription 3 (STAT3)-related signaling pathway in K562 cells, but not HL-60 and U937. LPC significantly inhibited phosphorylation and the DNA binding activity of STAT3, phosphorylation of Src and Janus activated kinase 2 (JAK2) as well as the expression of STAT3 target genes Cyclin D1, Cyclin E, Bcl-xL, Bcl-2, and Survivin at both protein and mRNA levels. LPC stimulated protein tyrosin phosphatases Src homology 2 domain-containing phosphatase 1 (SHP-1), but not SHP-2, SOCS-1 and PTEN. Blocking SHP-1 using sodium pervanadate or SHP-1 specific siRNA reversed LPC-mediated inactivation of STAT3 and HDAC and induction of apoptosis. Of importance, LPC blocked the nuclear translocation and interaction of HDAC3 and STAT3 as well as their binding onto the Bcl-xL promoter. Overall, our findings demonstrate that LPC has a potent anti-cancer activity through inducing apoptosis by suppressing HDAC3/STAT3 activation and binding to the Bcl-xL promoter in K562 cells.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 2015. doi:1538-7445.AM2012-2015
Abstract 2020: Suppression of Twist expression sensitizes human non-small cell lung cancer (NSCLC) cells to cisplatin
Authors: Park In-Chul
Published: August 2012

Twist, a highly conserved basic helix-loop-helix transcriptional factor, is a major inducer of epithelial-mesenchymal transition (EMT) and involved in different phases of tumorigenicity. The elevated Twist expression in cancer cells has been shown to promote metastasis and induce chemoresistance. In this study, we have investigated the mechanism of action by which Twist depletion induces chemosensitivity of lung cancer cells to chemotherapeutic agent. We found that knock-down of Twist expression by siRNAs sensitized H1299 cells to cisplatin by inhibiting mTOR activity. Inhibition of mTOR pathway using mTOR or S6K siRNAs led to potentiate cell death in cisplatin resistant H1299 cells. Our results suggest that suppression of Twist expression sensitized NSCLC cells to cisplatin treatment by down-regulating mTOR pathway.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 2020. doi:1538-7445.AM2012-2020
Abstract 2026: Targeted delivery of therapeutic NefM1 peptide with single-walled carbon nanotubes (SWNTs) suppresses growth of 3-D cultures of breast and colon cancers
Authors: Janagama Dasharatham G. , Basson Marc D. , Bumpers Harvey L.
Published: August 2012

Background and Objectives: In vitro studies with NefM1 peptide demonstrated apoptotic effects on a number of cancer cell lines containing CXCR4 receptors. This suggests that NefM1 apoptotic peptides can be used as a therapeutic agent for breast and colon cancers. In this study we hypothesized these cancers cultured as 3-D tumor spheres will undergo greater regression and destruction after treatment with NefM1 peptide- linked-carbon nanotubes (SWNTs) particles than with NefM1 peptide alone. Material and Methods: Human breast cancer (MDA-MB-231, MDA-MB-468) and colon cancer (HT-29, WiDr, SW 480) cells were cultured with 1% Sea Prep hydrogel in supplemented RPMI1640 medium. After 3 weeks the 3-D cultures developed as tumor spheres. Like in vivo conditions these 3-D cultures had cell-cell interactions and the sphere is held together by an extracellular matrix. The cultures were treated with NefM1 peptide and NefM1 carbon nanotubes (SWNTs) to compare their relative effects on the growth of the tumor spheres. The viability status of the tumor spheres was established using ethidium bromide and acridine stain and fluorescent microscopy. The Nef peptide was linked to SWNTs using thionine and binding was verified by gel electrophoresis. Transmission electron microscopy (TEM) showed internalization of NefM1-SWNTs nanoparticles. Results: In 3-D cultures, the SeaPrep hydrogel facilitated the growth of tumor spheres. All breast (MDA-MB-231, MDA-MB-468) and colon (HT-29, WiDr, SW-480) tumor spheres showed reduction and disintegration with nefM1 administration. This was enhanced with the NefM1- SWNTs conjugate which destroyed all cells in the culture. This indicated effective delivery of NefM1 into the cells via SWNTs. This is novel in that not only were these growing tumor spheres killed by the peptide binding to the CXCR-4 receptors, but the nanoparticles allowed for increased internalization of the peptide into the cells. The NefM1-SWNT's will undergo further study regarding its ability to enhance suppression of these tumors in SCID mice models. Conclusion: An apoptotic peptide (NefM1) can be linked to a nanoparticle which enhances its cytotoxicity to breast and colorectal cancers in vitro. Using 3-D cultured tumor spheres to evaluate the suppression of cancer growth is convenient and cost effective for drug delivery and drug discovery.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 2026. doi:1538-7445.AM2012-2026
Abstract 2029: Rictor regulates FBXW7-dependent c-Myc and cyclin E degradation in colorectal cancer
Authors: Guo Zheng , Zhou Yuning , Evers Mark , Wang Qingding
Published: August 2012

Rictor (Rapamycin-insensitive companion of mTOR) forms a complex with mTOR and phosphorylates and activates Akt. Activation of Akt induces expression of c-Myc and cyclin E, which are overexpressed in colorectal cancer and play an important role in colorectal cancer cell proliferation. Here, we show that rictor associates with FBXW7 to form an E3 complex participating in the regulation of c-Myc and cyclin E degradation. The Rictor-FBXW7 complex is biochemically distinct from the previously reported mTORC2 and can be immunoprecipitated independently of mTORC2. Moreover, disruption of the interaction between rictor and FBXW7 by knockdown of rictor increased FBXW7 ubiquitination and degradation. Knocking down of rictor in serum-deprived colorectal cancer cells results in the decreased ubiquitination and increased protein levels of c-Myc and cyclin E while overexpression of rictor induces the degradation of c-Myc and cyclin E proteins. Genetic knockout of FBXW7 blunts the effects of rictor, suggesting that rictor regulation of c-Myc and cyclin E requires FBXW7. Our findings identify rictor as an important component of FBXW7 E3 ligase complex participating in the regulation of c-Myc and cyclin E protein ubiquitination and degradation. Importantly, our results suggest that elevated growth factor signaling may contribute to decrease Rictor/FBXW7-mediated ubiquitination of c-Myc and cyclin E, thus leading to accumulation of cyclin E and c-Myc in colorectal cancer cells.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 2029. doi:1538-7445.AM2012-2029
Abstract 2030: Mechanisms of E2F-mediated centrosome amplification in HER2 positive breast cancer
Authors: Lee Miyoung , Saavedra Harold I.
Published: August 2012

Centrosome amplification associates with most human cancers and has been suggested to trigger tumor initiation and progression. Centrosomes are the major microtubule organizing centers in animal cells. They are duplicated once in each cell cycle and duplicated centrosomes establish mitotic spindle bipolarity. Centrosome duplication is tightly regulated because deregulated centrosome duplication leads to centrosome amplification, multipolar and pseudobipolar mitotic spindles that potentiate chromosome instability and may trigger tumorigenesis. One of the tumor types in which the relationship between centrosome amplification and tumorigenesis is well known are breast cancers, as most breast tumors exhibit centrosome amplification. Centrosome amplification arises through various mechanisms, such as deregulation of cell cycle regulators, oncogenes and tumor suppressor genes. Our laboratory discovered that HER2 positive breast cancer cells and K-RasG12D pre-malignant mouse mammary lesions displayed overexpressed E2F1-3 transcriptional activators. Although E2Fs are involved in various cellular processes, including the regulation of the cell and centrosome duplication cycles and differentiation, the role of E2Fs in centrosome amplification in breast tumors is unknown. To study the role of the E2F activators in centrosome amplification, we first assayed centrosome amplification in the E2F-overexpressing HER2 positive breast cancer cells, HCC1954, JIMT1 and SKBR3. There was increased centrosome amplification in the HER2 positive cell lines HCC1954, JIMT1 or SKBR3 relative to MCF10A control mammary epithelial cells. To find if centrosome amplification shown in HER2 positive cells overexpressing E2F1-3 was E2F-specific, the siRNA-mediated silencing technique was applied to transiently knock-down the E2F activators (E2F1-3a) and cells were assayed for centrosome amplification. In addition, to permanently knockdown E2Fs in HER2 positive cells, shRNA-mediated silencing technique will be applied. Western blots confirmed that E2Fs knockdown was efficient, followed by decreased centrosome amplification in HER2 positive cells downregulated for E2F1 and E2F3a; results for E2F2 are still awaiting. These data suggested that centrosome amplification detected in HER2 positive cells is specific to the E2F activators. Future experiments will involve the identification of E2F-dependent transcriptional targets mediating centrosome amplification in HER2 positive cells.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 2030. doi:1538-7445.AM2012-2030
Abstract 2033: Protection of quiescent fibroblasts from proteasome inhibition-mediated proteotoxicity
Authors: Legesse-Miller Aster , Raitman Irene , Haley Erin M. , Liao Albert , Sun Lova L. , Wang David J. , Lemons Johanna M.S. , Suh Eric J. , Johnson Elizabeth L. , Laund Benjamin A. , Coller Hilary A.
Published: August 2012

Objectives: The aim of this study is to investigate quiescence-related resistance to proteasome-inhibition mediated proteotoxicity, and its regulatory mechanisms. Introduction: Quiescence, the reversible arrest of cellular proliferation, is critical for normal cellular development and tissue maintenance. Moreover, while proteasome inhibitors (PIs) are used to treat specific cancers, it has been shown that subsets of cells within a tumor, through quiescence, avoid PI-induced apoptosis and survive. Thus, deciphering the mechanisms through which quiescent cells resist proteasome inhibition is of basic and clinical relevance. Methods: Proliferating and quiescent primary human fibroblasts were used as a model system to study the effects of PIs (MG132, epoxomycin and Bortezomib). Flow cytometry, Western blots, microscopy and microarrays were used to identify specific pathways activated by proliferating and quiescent fibroblasts in response to proteasome inhibition. Results: We found that the proteasome-inhibited quiescent fibroblasts maintain viability while proliferating fibroblasts rapidly undergo apoptosis. We further found that proteasome-inhibited proliferating and quiescent fibroblasts up-regulate multiple protective pathways, including authophagy, chaperones and antioxidants, and evidence no observable difference in proteasome-mediated degradation pathway activity. However, we found that in proliferating fibroblasts, proteasome inhibition leads to increased reactive oxygen species (ROS), peri-nuclear protein aggresomes, and apoptotic cell death. By comparison, proteasome-inhibited quiescent cells display selective up-regulation of MnSOD (a ROS detoxifying enzyme), dispersed protein aggregates, and less apoptotic cell death. Knockdown of p62/SQSTM1 (a protein important for aggresome formation) or treatment with an antioxidant N-acetylcysteine results in reduced aggresome formation and decreased apoptosis in response to proteasome inhibition. Moreover, inhibition of autophagy sensitizes serum-starved quiescent cells to proteasome inhibition induced apoptosis. This suggests that multiple mechanisms, including autophagy, ROS detoxification and differential subcellular trafficking of ubiquitinated proteins are implicated in quiescence-related resistance to PI-induced apoptosis. These results point to possible strategies for improving the effectiveness of PIs in cancer treatment.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 2033. doi:1538-7445.AM2012-2033
Abstract 2034: The Fbw7 tumor suppressor: Genetic and biochemical approaches to study a dimeric ubiquitin ligase
Authors: Larimore Elizabeth A. , Welcker Markus , Clurman Bruce E.
Published: August 2012

Fbw7, the human ortholog of yeast Cdc4, is the substrate-recognition component of a cullin-RING ubiquitin ligase and regulates a number of oncoprotein substrates including cyclin E, c-Myc, Notch, and c-Jun. A growing number of additional pro-growth and pro-survival factors have been more recently described as Fbw7 targets. Accordingly, Fbw7 is a tumor suppressor and is mutated in approximately ten percent of human malignancies. Its loss leads to chromosomal instability and tumorigenesis. Fbw7 binds its substrates after they become phosphorylated within a consensus sequence called a Cdc4 phospho-degron (CPD). CPD phosphorylation creates a high-affinity binding site for Fbw7, which brings the ligase machinery into proximity with the substrate, leading to ubiquitylation and proteasomal degradation of the substrate. Interestingly, Fbw7 contains a conserved dimerization domain and we can show the protein primarily forms dimers in cells. One Fbw7 dimer contains two substrate-binding interfaces and may make two independent CPD contacts, but the physiologic function of Fbw7 dimerization is still elusive. Using gene-targeting, we studied the effects of ablating Fbw7 dimerization in vivo and found endogenous cyclin E and c-Myc regulation are moderately affected. Using purified Fbw7 monomers or dimers in in vitro ubiquitylation assays, we observed that a dimeric ligase is able to conjugate ubiquitin moieties onto a wider variety of substrate lysine residues than can the monomeric ligase. This is likely due to the greater spatial reach of the dimer. Additionally, preliminary data shows that the dimer may be capable of catalyzing the formation of non-K48-linked polyubiquitin chains, whereas the monomer is not. Cancer-associated mutations in Fbw7 most often target three arginine residues that form a pocket for binding phosphorylated substrate CPDs. Mutation of any of these arginines render Fbw7 unable to bind substrates and properly regulate their abundance. However, these mutations occur typically in only one allele. Dimerization of Fbw7 may explain why such heterozygous mutations are sufficient for tumorigenesis: the mutant form may dominantly inhibit enzyme function of the wild-type allele through dimerization. We have generated hetero-typic Fbw7 dimers containing one wild-type and one mutant protein in each dimer and are currently studying its binding and ubiquitylation capabilities. Given the importance of Fbw7 and its substrates in tumor-progression pathways, the results of this study will further our understanding of human tumor suppression.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 2034. doi:1538-7445.AM2012-2034
Abstract 2037: Cyclin D1 overexpression supports stable EBV infection in nasopharyngeal epithelial cells
Authors: Tsao George Sai-Wah , Tsang C M. , Yip Y L. , Zhang G T. , Deng W , Lo K W. , Lau Victoria , Hau P M. , Lung M
Published: August 2012

Poorly or undifferentiated nasopharyngeal carcinoma (NPC) is regularly associated with Epstein-Barr virus (EBV) infection. EBV infection of premalignant nasopharyngeal epithelial cells has been postulated to play an important role in the pathogenesis of nasopharyngeal carcinoma. We have previously reported that genetic alterations could be detected in premalignant nasopharyngeal epithelial tissues. It is not clear if these genetic alterations may associate with the establishment of EBV infection. In this study, we have examined the role of cyclin D1 in supporting EBV infection in nasopharyngeal epithelial cells. While cyclin D1 is commonly expressed in nasopharyngeal carcinoma tissues we found that overexpression of cycline D1 is readily detected in dysplastic nasopharyngeal epithelial closely associated with EBV infection. Furthermore, using a panel of immortalized nasopharyngeal epithelial cells, we demonstrated that overexpression of cyclin D1 or CKD4 supports EBV infection and result in the establishment of stable EBV infected nasopharyngeal cell lines. We also observed that EBV infection induced growth inhibition and senescence in nasopharyngeal epithelial cells, which may be the underlying reason for low detection rate of EBV infection in healthy nasopharyngeal epithelial tissues. However, cyclin D1 overexpression suppressed senescence and differentiation in the immortalized nasopharyngeal epithelial cells, suggesting cyclin D1 may play an important role in supporting EBV infection in these cells. We postulated that the altered expression of cyclin D1 or CDK4 in premalignant nasopharyngeal epithelial cells may over-ride the growth suppression effects of EBV infection and support EBV latent infection in premalignant nasopharyngeal epithelial cells. Acknowledgement: This study is sponsored by grant support received from the Hong Kong Research Grant Council (Grant number: GRF780911 and AoE /M-06/08)

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 2037. doi:1538-7445.AM2012-2037
Abstract 2047: The interaction of ZSCAN4 with TRF1: Effects on regulation of telomere elongation in cancer cells
Authors: Lee Kyungwoo , Gollahon Lauren
Published: August 2012

Telomeres are repetitive sequences at the ends of chromosomes protected by DNA binding proteins of shelterin complex that form capping structures. One of these proteins, TRF1, plays a role in the regulation of telomere length through its interaction with other proteins in the shelterin complex. High levels of TRF1 expression have been shown to accelerate telomere shortening, whereas dominant-negative inhibition of TRF1 leads to telomere elongation. Recently, the newly identified Zinc finger and SCAN domain containing4 gene (ZSCAN4) plays a key role in genomic stability by regulating telomere elongation and was shown to co-localize with TRF1 foci. Here, we show that ZSCAN4 is a TRF1 association protein. We hypothesized that ZSCAN4 would affect levels of TRF1expression and correspondingly control the function of TRF1 through negative regulation and dominant-negative inhibition. Using Co-immunoprecipitation, we demonstrate that ZSCAN4 binds with TRF1 in vitro. Pull-down assay results show that ZSCAN4 directly interacts with TRF1. Using PCR mutagenesis, we have identified the binding site between ZSCAN4 and TRF1. Furthermore, over-expression of ZSCAN4 expression acts as an activator of telomere elongation. The ZSCAN4 mediated cooperative binding of TRF1 to telomeric DNA has important implications for understanding the mechanism of telomere extension.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 2047. doi:1538-7445.AM2012-2047
Abstract 2050: Plk1 phosphorylation of Orc2 promotes DNA replication under conditions of stress
Authors: Song Bing , Liu X. Shawn , Davis Korbin , Liu Xiaoqi
Published: August 2012

Polo-like kinase 1 (Plk1) plays pivotal roles in mitosis, however, little is known about its function in S phase. In this study, we show that inhibition of Plk1 impairs DNA replication and results in slow S-phase progression in cultured cancer cells. We have identified Origin Recognition Complex 2 (Orc2), a member of the DNA replication machinery, as a Plk1 substrate and have shown that Plk1 phosphorylates Orc2 at Ser188 in vitro and in vivo. Furthermore, Orc2-S188 phosphorylation is enhanced when DNA replication is under challenge induced by UV, hydroxyurea, gemcitabine, or aphidicolin treatment; cells expressing the unphosphorylatable mutant (S188A) of Orc2 had defects in DNA synthesis under stress, suggesting that this phosphorylation event is critical to maintain DNA replication under stress. To dissect the mechanism pertinent to this observation, we showed that Orc2-S188 phosphorylation associates with DNA replication origin and that cells expressing Orc2-S188A mutant fail to maintain the functional pre-replicative complex (pre-RC) under DNA replication stress. Furthermore, the intra-S phase checkpoint is activated in Orc2-S188A-expressing cells to cause delay of S-phase progress. Our study suggests a novel role of Plk1 in facilitating DNA replication under conditions of stress to maintain genomic integrity.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 2050. doi:1538-7445.AM2012-2050