bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 1787: ImmTACs: Bi-functional reagents for redirected tumour cell killing
Authors: Paston Samantha J. , Ashfield Rebecca , Liddy Nathaniel , Bossi Giovanna , Adams Katherine , McCormack Emmet , Lissina Anna , Mahon Tara , Hassan Namir , Gavarret Jessie , Bianchi Frayne , Pumphrey Nick , Ladell Kristin , Gostick Emma , Sewell Andrew K. , Lissin Nikolai , Molloy Peter , Li Yi , Cameron Brian , Sami Malkit , Baston Emma , Harper Jane , Johnson Andrew , Vuidepot Annelise , Todorov Penio , Dennis Rebecca , Kalos Michael , June Carl , Price David A. , Williams Daniel , Jakobsen Bent K.
Published: June 2012

The human immune system can theoretically identify malignant cells by inspecting cell surface Class I HLA -peptide complexes for the presence of disease-associated epitopes. Indeed, many cancer patients generate CD8 cyto-toxic T cell responses to tumour-associated antigens; the majority of patients, however, fail to clear tumours since T cell avidity for self-antigens tends to be weak, and cancer cells employ escape mechanisms for avoiding destruction by T cells. To overcome these issues, we have engineered novel, bi-functional protein therapeutics termed ImmTACs (Immune Mobilising mTCR Against Cancer) which re-direct the immune system to target and destroy tumour cells with a high degree of potency and specificity. An ImmTAC comprises a high affinity ‘monoclonal’ T cell Receptor (mTCR) targeting a cancer-associated HLA-peptide complex, fused to an anti-CD3 scFv domain which activates an anti-tumour T cell response.

We demonstrate that ImmTACs against a number of different cancer-associated antigens can target and kill tumour cells expressing as few as 10-20 epitopes per cell with pico-Molar potency. ImmTACs preferentially activate effector memory CD8 T cells, resulting in secretion of multiple cytokines and tumour cell killing; a single activated T cell can kill multiple antigen positive tumour cells. Furthermore, we demonstrate that the reagents are able to inhibit tumour growth in mouse xenograft models. In vitro ImmTAC potency translates to a dose of less than 1mg in humans, representing a significant advance over existing targeted anti-cancer therapies. Currently we are conducting a phase I dose-escalation trial and Phase 0 exploratory trial using ImmTAC-gp100 in late stage melanoma patients. In summary, ImmTACs offers a novel therapeutic approach for the treatment of various cancers with the potential to provide major benefits over current treatments including reduction in dose to sub-mg quantities and an improved safety profile.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 1787. doi:10.1158/1538-7445.AM2011-1787
Abstract 1790: Functional importance of EpCAM for the activity of tumor-initiating cancer cells and their eradication by EpCAM/CD3-bispecific antibody MT110
Authors: Münz Markus , Herrmann Ines , Friedrich Matthias , Murr Alexander , Kufer Peter , Baeuerle Patrick A. , Raum Tobias
Published: June 2012

We have recently shown that tumor-initiating cells (TICs) derived from colorectal cancer tissue express EpCAM and can be eradicated in cell culture experiments as well as established xenograft mouse models by EpCAM/CD3-bispecific BiTE antibody MT110 (Hermann et al., 2010; PLoS ONE, 5: e13474). Here we show that TICs derived from liver and breast cancer tissue grow tumors in SCID mice at cell doses as low as 100 after a latency period of 37-58 days. Like colorectal cancer tissue-derived TICs, they expressed EpCAM and grew colonies in soft agar. In the presence of human T cells, breast and liver cancer-derived TICs could be eradicated by BiTE antibody MT110 as tested in a sensitive soft agar assay.

In order to assess the role of EpCAM expression in colorectal TICs, we used short hairpin RNA molecules for specifically knocking down EpCAM expression. A shRNA molecule binding to the coding, non-translated 3’-UTR of the EpCAM mRNA showed the highest efficacy (ca 10-fold) in suppressing EpCAM expression, as shown at the mRNA level by RT-PCR and at the surface expression level by FACS analysis. Although the EpCAM knockdown was not complete and left ca 10% of the normal EpCAM expression, a strong impact on the phenotype of colorectal TICs was observed. Morphologically, TICs with reduced EpCAM expression adopted in cell culture an epithelial-like phenotype, could no longer efficiently close a scratch in a cell culture experiment, and reduced the expression of epithelial-mesenchymal transition (EMT)-associated proteins such as fibronectin, vimentin and slug. Compared to non-sense control shRNA, colony formation of TICs in soft agar was reduced by >60%. Most importantly, the tumor-initiating potential of TICs appeared to depend on a high level of EpCAM expression. TICs expressing the EpCAM-specific shRNA could no longer form a tumor at a dose of 500 cells after 116 days, whereas the non-sense shRNA-expressing TICs formed during this period a tumor of 0.5 cm3.

Our data suggest that a certain EpCAM expression level is required for maintenance of a stem cell-like phenotype of colorectal TICs. This is consistent with a signal transducing role of EpCAM within a non-canonical wnt pathway (Maetzel et al. 2009; Nature Cell Biol. 11: 162-171), and in the process of EMT of colorectal cancer cells.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 1790. doi:10.1158/1538-7445.AM2011-1790
Abstract 1793: 5-Fluorouracil chemotherapy acts synergistically with adoptive dendritic cell immunotherapy to induce durable anti-tumor responses in vivo
Authors: Williams Patrick , Bouchentouf Manaf , Rafei Moutih , Romieu-Mourez Raphaelle , Yuan Shala , Forner Kathy A. , Birman Elena , Galipeau Jacques
Published: June 2012

Introduction: 5-Fluorouracil induces a form of cell death that facilitates the presentation of antigens by dendritic cells and depletes myeloid suppressor cells in vivo. The resulting immune modulation plays a substantial role in prolonging survival of mice with cancer. Our lab has previously demonstrated that a fusion of granulocyte-colony macrophage-stimulating-factor (GM-CSF) and Interleukin (IL) – 21, hereafter GIFT-21, could differentiate monocytes into a novel dendritic cell population (GIFT-21 DC). Characterization of these cells revealed a unique phenotype, with the expression of markers normally associated with both conventional and plasmacytoid dendritic cells. They are effective at cross-presenting antigens and secrete substantial amounts of CCL2, IL-6, TNF-α and IFN-α. When injected into tumor-bearing mice, they migrate to and sample from tumors in vivo, allowing them to educate CD8 T cells to recognize tumor antigens, which then significantly impeded the growth of B16 and D2F2 Neu tumors in mice.

Hypothesis: We propose that 5-Fluorouracil mediated immune modulation can be combined with adoptively transferred autologous GIFT-21 DCs to treat established cancer, and lead to a synergistic anticancer immune response.

Methods and Results: Twelve days following a subcutaneous tumor implantation, mice with established B16 tumors (average volume of 40mm3) were treated daily with 1.5 mg 5-Fluorouracil for 2 days and then injected on the third day with 1.5×106 syngeneic GIFT-21 DCs intraperitoneally. The treatment was repeated one week later. While 5-FU treatment alone or dendritc cell therapy alone had no significant impact on the growth of pre-established melanoma, the combination of 5-Fluorouracil with GIFT-21 DC-based immunotherapy reduced the rate of tumor growth by 75% over 5-FU and 85% over untreated mice, while eventually leading to the regression of the pre-established B16 tumors.

Conclusion: These findings demonstrate how chemotherapy can be combined with immunotherapy to target cancers for which tumors antigens are unknown or ill-defined. While chemotherapy suppresses some elements of the immune system in the short term, we speculate that modulating the endogenous immunological processes secondary to chemotherapy shows promising synergy with adoptive cell therapy and is a promising avenue of future research in cancer immunotherapy.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 1793. doi:10.1158/1538-7445.AM2011-1793
Abstract 1799: Can trastuzumab enhance the effectiveness of chemotherapy in patients with resected, HER2-positive, Stage II/III or Stage IV gastric cancer
Authors: Oshima Takashi , Kunisaki Chikara , Fujii Shoichi , Yamada Roppei , Rino Yasushi , Masuda Munetaka , Imada Toshio
Published: June 2012

Objectives: Trastuzumab, a monoclonal antibody against human epidermal growth factor receptor 2 (HER2), in combination with chemotherapy has been reported to significantly prolong survival in patients with HER2-positive, unresectable, advanced gastric cancer. However, the therapeutic usefulness of this regimen for surgically resected gastric cancer remains unclear. Adjuvant chemotherapy with S-1 has been shown to be beneficial in patients with stage II or III gastric cancer. Whether trastuzumab should be given to patients with resected, HER2-positive gastric cancer is a very interesting problem. We therefore examined whether trastuzumab can be expected to enhance the effectiveness of chemotherapy in patients with resected, HER2-positive, stage II/III or IV gastric cancer.

Methods: The study group comprised 183 patients: 116 had stage II or III gastric cancer and received adjuvant chemotherapy with S-1 after surgery from May 2003 through March 2008; the other 67 had stage IV gastric cancer and sequentially received new anticancer agents after surgery from July 1999 through September 2005. HER2 was evaluated by inmmunohistochemistry and FISH according to the protocol of a Study of Trastuzumab in Combination with Chemotherapy Compared with Chemotherapy Alone in Patients with HER2-Positive Advanced Gastric Cancer (ToGA study). Positive expression of HER2 was defined as a score of 3+ on immunohistochemical staining or as a score of 2+ on immunohistochemical staining plus relative HER2 DNA expression of >2.2 on FISH. Cumulative survival rates were compared between HER2-positive patients and HER2-negative patients.

Results: Among patients with stage II or III gastric cancer who received adjuvant chemotherapy with S-1, 20% had HER2-positive tumors; however, cumulative survival rates did not differ significantly between HER2-positive patients and HER2-negative patients. Among patients with stage IV gastric cancer who underwent surgery, 22% had HER2 positive tumors; however, cumulative survival rates did not differ significantly between HER2-positive and HER2-negative patients.

Conclusions: The survival benefit of adding trastuzumab to standard chemotherapy was considered minimal in patients with resected, HER2-positive stage II/III or stage IV gastric cancer.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 1799. doi:10.1158/1538-7445.AM2011-1799
Abstract 180: Suppressor of cytokine signaling 6 (SOCS6) induces apoptosis through promoting mitochondrial fragmentation and Bax activation
Authors: Lin Huan-Yu , Lin Chung-Chih , Lee Hsin-Chen , Wang Fung-Fang , Chen Jeou-Yuan
Published: June 2012

Mitochondria are highly motile organelles that constantly undergo fission and fusion. Impairment of mitochondrial dynamics is associated with mitochondrial dysfunction and frequently linked to the pathogenesis of neurodegenerative diseases and cancer. We have previously shown that loss-of-function of suppressor of cytokine signaling 6 (SOCS6) is a frequent event in human gastric cancer. In this study, we recapitulated the event of loss of SOCS6 in cancer-derived cell lines where endogenous SOCS6 was expressed, and we showed that elimination of SOCS6 expression delays UV-induced apoptosis process. By contrast, overexpression of SOCS6 in cancer cell lines that express endogenous SOCS6 at minimal levels promotes apoptosis. SOCS6 induces intrinsic apoptotic process and facilitates cytochrome c release by enhancing Bax conformational change, mitochondrial targeting and oligomerization. We further demonstrated that SOCS6 is a novel mitochondrial protein that regulates mitochondrial dynamics. By confocal microscopy, we showed that ectopically expressed SOCS6 is predominantly localized in mitochondria, and the carboxyl-terminal SOCS box is important for its mitochondrial localization. Localization of SOCS6 in mitochondria is accompanied with concomitant mitochondrial fragmentation, both of which can be reversed by coexpression of mitochondrial fusion proteins (Mfn1 and Mfn2) or a dominant-negative form of Drp1. Ectopic expression of SOCS6 severely impairs the dynamics and function of mitochondria. Taken together, our data suggest that SOCS6 is a novel mitochondrial protein that induces intrinsic apoptosis pathway through perturbing mitochondrial dynamics and facilitating Bax activation.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 180. doi:10.1158/1538-7445.AM2011-180
Abstract 1805: Integration of 17-(Dimethylaminoethylamino)-17-demethoxygeldanamycin (17-DMAG) in combinational immunotherapy targeting tumor antigen EphA2
Authors: Rao Aparna , Kawabe Mayumi , Storkus Walter J.
Published: June 2012

Introduction: EphA2 and HSP90 play important roles in tumor growth, survival and metastasis. An effective therapeutic approach would promote decreased expression of EphA2 on tumor cells, while increasing presentation of EphA2-derived peptides on tumor MHC class I complexes, thereby enhancing the recognition of tumors by anti-EphA2 CD8+ T effector cells. Aim: We are assessing the safety and effectiveness of the HSP90 inhibitor 17-DMAG in enhancing anti-tumor immunotherapy with EphA2-specific T cells in murine tumor models. Methods: We tested the effect of 17-DMAG on tumor cell expression of EphA2 and the MHC I antigen-processing machinery in vitro. Efficacy of 17-DMAG in an MCA205 sarcoma model was also evaluated in vivo. Results: 17-DMAG facilitated a dose-dependent reduction in EphA2 expression, but had no adverse effects on the antigen-processing machinery of the tumors. Similar effects were seen in an MCA205 sarcoma model in vivo, where reduction in EphA2 levels in tumors corresponded with increased recognition by EphA2-specific CD8+ T cells in vitro. 17-DMAG treatment also facilitated increased infiltration of CD4 and CD8 T cells in the tumor, compared to untreated cohorts. Conclusion: 17-DMAG treatment promotes immunologically-preferred alterations tumor cells in vitro as well as In vivo. Our preliminary in vivo studies suggest that decreased EphA2 levels in tumors is accompanied by increased presentation of EphA2 peptides in tumor MHC class I complexes. Further in vivo studies are warranted to assess the net anti-tumor impact of combinational immunotherapies integrating 17-DMAG.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 1805. doi:10.1158/1538-7445.AM2011-1805
Abstract 183: Loss of Bad phosphorylation and Mcl-1 expression is necessary for rapid apoptosisinduced by combination of TGFα-PE and ZSTK474 in prostate cancer cells
Authors: Baiz Daniele , Yancey Dana , Choi Young A. , Baurin Vitaliy , Karpova Yelena , Hassan Sazzad , Debinski Waldemar , Kulik George
Published: June 2012

Advanced androgen-independent prostate cancer is notoriously resistant to conventional systemic therapies, and at the moment there is no effective protocol for hormone-refractory advanced prostate cancer affected patients.

One of possible mechanisms of such resistance is activation of the anti-apoptotic signaling PI3K/Akt pathway that is constitutively active in the 60% of advanced prostate cancers. However, multiple inhibitors of PI3K are just well studied and some have just gone on to clinical trials, but unfortunately they showed limited efficacy against prostate cancer alone.

Our recent experiments has shown remarkable synergy in inducing rapid and massive apoptosis when PI3K inhibitor ZSTK474 is used in combination with Pseudomonas aeruginosa exotoxin A fragment fused with Transforming Growth Factor Alpha (TGFα-PE38).

Time lapse video microscopy of prostate cancer C4-2 cell line has shown the substantial cell death within 4-6 hours with combined administration, compared to limited cell death in single agents-treated cells.

Analysis of PARP and cleaved-caspase 3 fragment by Western blotting confirmed that cell death occurs via apoptosis. Quantitation of caspase 3 and 7 activity with luminescent AFC-DEVD substrate confirmed synergy in apoptosis activation by combination of TGFα-PE38 and ZSTK474, while single agents even in higher concentration did not induce substantial apoptosis within 6 hours time period.

Analysis of the mechanisms underlying this synergy has shown that PI3K inhibitor ZSTK474 triggers Bad dephosphorylation, while TGFα-PE38 reduces expression levels of Mcl-1. A slight increase of Bim protein was detected and related with ZSTK474 administration. No major variations were detected in the expression levels of pro- and anti-apoptotic proteins Bax, Bcl-2 and Bcl-XL.

To address the role of Bad phosphorylation and Mcl-1 expression in apoptosis induction by PI3K inhibitor ZSTK474 and TGFα-PE38, we examined apoptosis in C4-2 cells that express phosphorylation-deficient mutant Bad (BAD2SA). Results indicated that the expression of BAD2SA sensitized C4-2 cells to apoptosis induced by TGFα-PE38. Experiments that address the role of Mcl-1 loss in apoptosis induced by TGFα-PE38 are ongoing.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 183. doi:10.1158/1538-7445.AM2011-183
Abstract 1831: Using narrative to transform cancer communication: Impact of a cervical cancer storyline on primetime television
Authors: Murphy Sheila T. , de Castro Buffington Sandra , Hether Heather , Baezconde-Garbanati Lourdes A.
Published: June 2012

Epidemiological evidence reveals that Hispanic women lag behind all other ethnic groups in their rate of cervical cancer screening (National Health Interview Survey, 2006). Can stories succeed where more traditional cancer campaigns have faltered? Funding by the National Cancer Institute enabled researchers at Hollywood, Health & Society, the Annenberg School for Communication and Journalism, and the Institute for Prevention Research at the University of Southern California to encourage writers of primetime programming to include accurate information regarding cervical cancer.

Purpose of study: This study examined whether a popular Spanish language telenovela could impact Hispanic viewers’ knowledge, attitudes and behavior with respect to cervical cancer screening.

Experimental procedures: This was assessed by a quantitative survey of viewers before and after the fictional character of Dora in a Telemundo telenovela, El Clon, had an abnormal pap test.

Results: Statistical comparisons between the pretest and posttest surveys revealed that after seeing cervical cancer storyline viewers were 6 times more likely to know that a pap test detects cervical cancer; 4 times more likely to know that pap tests are available for free or at low cost; 80 percent more likely to know pap women should have a pap test at least every 2 years; and 80 percent more likely to intend to have pap in next 2 years. This shift was strongest among viewers who identified with Dora and reported being absorbed or “transported” while viewing.

Conclusions: A cervical cancer storyline on a popular telenovela was significantly associated with greater level of knowledge and intention to have pap test in the next 2 years. Implications of these results and the use of entertainment education in cancer communication more generally will be discussed.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 1831. doi:10.1158/1538-7445.AM2011-1831
Abstract 1843: An unintended consequence of Nrf2-directed cytoprotection against a major human bladder carcinogen and a counteracting solution
Authors: Ding Yi , Paonessa Joseph D. , Randall Kristen L. , Argoti Dayana , Munday Rex , Vouros Paul , Zhang Yuesheng
Published: June 2012

NF-E2 related factor 2 (Nrf2) plays a pivotal role in cytoprotection against carcinogens and oxidants. Activating Nrf2 has been widely suggested as a key chemopreventive strategy against cancer and other diseases. Here, we show that Nrf2 faces a serious dilemma in dealing with 4-aminobiphenyl (ABP), a major human bladder carcinogen from tobacco smoke and other sources. ABP is metabolized/activated primarily in the liver. While Nrf2 significantly protected mouse liver against ABP in vivo, bladder DNA damage was approximately 2.6 fold higher in Nrf2+/+ mice than in Nrf2-/- mice after ABP exposure, as measured by dG-C8-ABP, the predominant ABP-DNA adduct formed in bladder cells and tissues. Interestingly, Nrf2 significantly protected bladder cells against ABP in vitro. Subsequent experiments revealed that UDP-glucuronosyltransferase (UGT) might be the principal, if not the only mediator of the dichotomous effects of Nrf2. Nrf2 upregulated UGT, promoted glucuronide conjugation of ABP in the liver, and increased urinary excretion of the conjugate. While glucuronidation of ABP and its metabolites is generally a detoxification process, these conjugates, which are excreted in urine, are known to be unstable in acidic urine and deliver the parent compounds to bladder. Hence, higher liver UGT activity may protect liver against ABP but increases bladder exposure to ABP. We further show that 5,6-dihydrocyclopenta[c][1,2]-dithiole-3(4H)-thione (CPDT), a promising cancer chemopreventive dithiolethione, was able to significantly activate Nrf2 cytoprotective signaling and to inhibit dG-C8-ABP formation in the bladder in vivo, but did not modulate liver UGT expression. Thus, CPDT exemplifies a counteracting solution to the dilemma faced by Nrf2 with regard to ABP. In conclusion, our study shows that Nrf2 increases ABP genotoxicity in the bladder in vivo and that UGT may be the key mediator. Using CPDT, we also demonstrate how the adverse effect of Nrf2 may be mitigated.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 1843. doi:10.1158/1538-7445.AM2011-1843
Abstract 2146: Role of apoptosis-inducing factor in the death of MDA-MB-231 breast carcinoma cells after DNA damage
Authors: Feng Xiaoxing , Zhou Yiran , Koh David W.
Published: June 2012

Apoptosis-inducing factor (AIF) is a flavoprotein that plays a cytoprotective role within mitochondria and induces cell death when it translocates to the nucleus in response to apoptotic signals. The objective of this study was to pursue the relevance of AIF in cancer. We showed that AIF translocated to the nucleus and mediated caspase-independent cell death in MDA-MB-231 breast cancer cells following treatment with the DNA alkylating agent N-methyl-N’-nitro-N-nitrosoguanidine (MNNG). We utilized small interfering RNA (siRNA) to down-regulate AIF expression in various human cancer cell lines and mouse trophoblast stem (TS) cells. AIF levels were reduced approximately 90% compared to untransfected cells. Analysis of AIF levels after subcellular fractionations demonstrated that nuclear translocation was compromised in these AIF knockdown cells. FACS analysis showed that AIF knockdown protected MDA-MB-231 against apoptosis induced by the chemotherapeutic agent cyclophosphamide. Furthermore, siRNA knockdown of AIF increased MDA-MB-231 cell survival following UV radiation. Taken together, the results demonstrate a role for AIF in cancer cell death following radio- or chemotherapeutic treatments. Thus, the activation of AIF mediated cell death is a promising method to improve the treatment of breast cancer patients.

This research was supported by funding from the American Cancer Society and the Pharmaceutical Research Manufacturers of America.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 2146. doi:10.1158/1538-7445.AM2011-2146
Abstract 2152: Paroxetine, a selective serotonin re-uptake inhibitor, induces growth inhibition and apoptosis in prostate cancer cells
Authors: Shibuya Tadamasa
Published: June 2012

Introduction and Objective: Recently, neuroendocrine cells is suggested to be involved in prostate cancer (PC) progression through the secretion of a variety of neuroendocrine products, such as serotonin, which have mitogenic effects on adjacent cancer cells and thus can contribute to the androgen independent proliferation of PC cells. With having experience that a patient with hormone refractory prostate cancer responded to paroxetine, a selective serotonin re-uptake inhibitor (SSRI), in serum prostate specific antigen (PSA) level, we investigated the effect of paroxetine on the growth and apoptosis induction in PC cell lines.

Methods: Paroxetine was administered to four patients with hormone refractory, metastatic PC and their serum PSA levels were determined. The effects of paroxetine on the growth and apoptosis induction in PC cell lines (PC3, DU145, 22RV1 and LNCaP) were confirmed. Cell growth inhibition was assessed in vitro by cell number count. The induction of apoptosis was examined by flow cytometry analysis and caspase activations were confirmed by western blot analysis.

Results: Serum PSA levels decreased soon after the administration of paroxetine but increased again in all four patients but one in whom it decreased from 0.485 ng/ml to undetectable level (less than 0.008 ng/ml) for over three years. In vitro study, paroxetine inhibited cell growth in a dose and time dependent manner. The 50% inhibitory concentration (IC50 values) of paroxetine in PC3, 22RV1, DU145 and LNCaP cells were 38.8, 30.1, 35.9 and 21µM, respectively. Apoptosis induction following the treatment of paroxetine was revealed in prostate cancer cells by the increase of sub-G1 population in flow cytometry analysis and a cleavage of procaspase-3 in western blot analysis.

Conclusions: A potent SSRI, paroxetine, induces growth inhibition and apoptosis in PC cells. These results suggest that SSRIs have the potential of anti-tumor effect against PC.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 2152. doi:10.1158/1538-7445.AM2011-2152
Abstract 2154: Simultaneous analysis of cell death mechanisms and oxidative stress using live cell fluorescence microscopy
Authors: Yan Michelle , Clarke Scott T. , Mandavilli Bhaskar S. , Huang Shih-Jung , Singh Upinder , Janes Michael S. , Dolman Nicholas J.
Published: June 2012

Reactive oxygen species (ROS) can directly damage DNA, proteins and lipids, while increased oxidative stress has been reported to be associated with apoptotic and autophagic cell death. Importantly, increased oxidative stress and dysregulation of cell death mechanisms have been linked with cancer development. To better understand the connection between increased oxidative stress and cell death pathways, it is important to be able to monitor both of these events simultaneously in the same cell and amongst mixed populations of cells. The goal of this study was to utilize multi-parametric fluorescence microscopy to examine the temporal progression of oxidative stress relative to cell death. By using a fluorogenic caspase substrate in combination with a near infrared fluorescent probe for ROS, we observed that increased oxidative stress was followed by activation of caspase-3/7 after induction by several agonists. We examined the effect of various agonists in both primary cells as well as cancer cell lines. Loss of mitochondrial membrane potential, nuclear condensation, and cell proliferation were also measured to establish a more comprehensive assessment of the cellular response to stress in these models. By simultaneously examining multiple parameters over time, we were able to define the temporal progression of apoptosis relative to the onset of oxidative stress. Furthermore, we were able to characterize the mechanism of cell death by discriminating between cells which were apoptotic (active caspase-3/7), autophagic (LC3B-positive autophagosomes), or both. This multi-parametric approach provided detailed spatial information at the cellular level so that correlations and temporal resolution could be determined between oxidative stress and cell death mechanisms.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 2154. doi:10.1158/1538-7445.AM2011-2154
Abstract 2156: c-Myb promotes growth, invasion and androgen-independence of prostate cancer cells
Authors: Srivastava Sanjeev K. , Bhardwaj Arun , Singh Seema , Reed Eddie , Grizzle William , Singh Ajay P.
Published: June 2012

Most prostate cancer (PCa) patients treated with first line androgen-deprivation therapy (ADT) initially exhibit a dramatic regression of the androgen-dependent (AD) cancer cells; however, the tumors eventually progress to an androgen-independent (AI) stage. The resulting hormone-refractory tumors are highly aggressive and resistant to conventional therapies. The causal mechanisms for such a transition remain largely unknown; however, a number of genetic and epigenetic alterations are believed to be involved in the process of AI progression of PCa. In an earlier study, c-Myb was identified among the genes that are amplified at higher frequency in hormone-refractory PCa. The c-Myb proto-oncogene is a cellular progenitor of v-Myb oncogenes, which encodes for a transcription factor and confers its oncogenic activity by regulating the expression of several target genes. In the present study, we have investigated the functional role of c-Myb in prostate cancer cells. Our data showed that c-Myb was overexpressed in AI (C4-2) PCa cells as compared to AD (LNCaP) PCa cells of the same genotypic lineage. Furthermore, we found that c-Myb was regulated by androgens in a biphasic manner and its expression correlated with the effect of androgens on prostate cancer cell growth. Short-hairpin RNA (shRNA)-mediated silencing of c-Myb in C4-2 cells reduced their growth, clonogenicity, motility and invasion, whereas its ectopic expression in LNCaP cells had opposite effects. Moreover, we observed that the overexpression of c-Myb in LNCaP and C4-2 cells was associated with their growth in androgen-depleted media, suggesting its role in androgen-independence. Immunoblot analysis demonstrated loss of mesenchymal markers N-cadherin, vimentin, Slug, Snail, and Twist, and gain of epithelial marker, E-cadherin in c-Myb-silenced C4-2 cells. In contrast, c-Myb overexpressing LNCaP cells exhibited a loss of epithelial and gain of mesenchymal markers, thus indicating a role of c-Myb in epithelial to mesenchymal transition (EMT) of PCa cells. Altogether, our studies provide first experimental evidence for a functional role of c-Myb in altered growth, androgen-independence and malignant behavior of the prostate cancer cells.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 2156. doi:10.1158/1538-7445.AM2011-2156
Abstract 2159: Molecular characterization of ERG mediated CXCR4 transcriptional regulation
Authors: Singareddy Rajareddy , John Jason Saint , Chinni Sreenivasa R.
Published: June 2012

CXCR4 is a chemokine receptor, which has been shown to be expressed in several types of tumor cells, and mediate invasion and metastasis. CXCR4 expression is transcriptionally regulated in cancer cells and is associated with aggressive phenotypes of prostate cancer. Previously, we and others have shown that ERG transcription factor regulates CXCR4 expression in prostate cancer cells. In prostate cancer patients ERG is expressed via chromosomal alterations resulting in fusion of androgen responsive TMPRSS2 promoter with the coding sequence of ERG transcription factor and systemic androgens regulate ERG expression. We further show that in TMPRSS2-ERG fusion positive prostate cancer cells androgens regulate CXCR4 expression via activating ERG transcription factor expression. The CXCR4 promoter contains several putative ERG/Ets transcription factor binding sites. We hypothesize that ERG binds to selective ERG/Ets binding sites in the CXCR4 promoter and activates CXCR4 expression. Using electrophoretic mobility shift assay experiments with each of the individual ERG/Ets binding sites with VCaP cell nuclear extracts and in vitro translated ERG protein, sequential ERG/Ets binding site deletions in CXCR4 promoter reporters, chromatin immunoprecipitation experiments with anti-ERG antibodies in TMPRSS2-ERG positive VCaP cells, and chemoinvasion studies with VCaP cells we show that: (a) ERG expressed in VCaP cells selectively interacts with specific ERG/Ets bindings sites in the CXCR4 promoter; (b) several Ets factor family members are expressed in VCaP cells; (c) in vitro translated ERG binding with ERG/Ets elements in CXCR4 promoter further confirms that selective ERG/Ets sites in CXCR4 promoter are active in binding with ERG; (d) ERG binds with CXCR4 gene promoter in VCaP cells; (e) androgens regulate CXCR4 expression in TMPRSS2-ERG fusion positive cells; and (f) androgen induced CXCR4 is active in chemoinvasion of prostate cancer cells. These data suggest that androgens regulate CXCR4 expression via ERG transcription factor expression and ERG factor may regulate CXCR4 expression by binding to the specific ERG/Ets responsive elements and activating CXCR4 transcription. These findings may provide a link between TMPRSS2-ERG translocations and enhanced metastasis of tumor cells via CXCR4 function in PC cells.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 2159. doi:10.1158/1538-7445.AM2011-2159
Abstract 2164: High fat diet-induced intraprostatic inflammation involves association between Stat-3 and NF-kappaB: Role in pathogenesis of prostate cancer
Authors: Shankar Eswar , Vykhovanets Eugene V. , Vykhovanets Olena V. , Bhaskaran Natarajan , Shukla Sanjeev , Gupta Sanjay
Published: June 2012

Western style high-fat diet (HFD) is a risk factor for obesity, diabetes, and cancer. In our previous studies we have demonstrated that HFD increases oxidative stress and inflammation in the prostate of NF-kappaB-luciferase transgenic reporter mice (Vykhovanets et al., Prostate 2010). However, the molecular mechanism(s) underlying HFD-induced inflammation in the prostate largely remains unknown. Signal transducer and activator of transcription (Stat-3) and nuclear factor-kappa B are the important signaling pathways that are activated during inflammation. The pro-inflammatory cytokines induced are the transcriptional targets of Stat-3 and NF-kappaB and are known to be involved in their activation. We examined the role of HFD in increasing the nuclear activation of Stat-3 and NF-kappaB and the mechanism(s) involved leading to inflammation of the prostrate. Male 16 week old C57BL/6J mice were either fed with regular diet (RD) or HFD for 4 and 8 weeks. The animals were sacrificed at the end of each time period. HFD intake caused significant increase in the plasma levels of IL-6 and IL-17, after 4 weeks of feeding that further increased after 8 weeks, compared to mice fed with RD. These cytokines have been reported to induce Stat-3. HFD intake also caused marked increase in the levels of IL-1beta and TNFalpha which are known activators of NF-kappaB. A significant increase in the levels of PKCepsilon was observed after 4 weeks of HFD feeding in these mice. Elevated levels of PDK1 and p-Akt (Ser473) were observed in the prostate tissues of HFD group. Increase in the levels of kinases corroborated with the nuclear translocation of Stat-3 and NF-kappaB/p65/RelA with significantly increased expression of Tyr705, Ser727 Stat-3 and Ser536 p65/RelA in the prostrate of HFD mice. An association between Ser727 Stat-3 and Ser 536 p65/RelA was observed in the nucleus of prostate tissue of HFD group. Furthermore, increased DNA binding of p65/RelA and Stat-3 as well as their association was observed in HFD group. Taken together, HFD increases inflammation in the prostate through activation of Stat-3 and NF-kappaB and their association may play a key role in the pathogenesis of prostate cancer.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 2164. doi:10.1158/1538-7445.AM2011-2164
Abstract 2173: Negative transcriptional regulation of the human type III TGFβ receptor in ccRCC
Authors: Cooper Simon J. , Roemeling Christina Von , Marlow Laura , Tun Han W. , Copland John A.
Published: June 2012

Renal cell carcinoma (RCC) accounts for 3% of all malignancies reported each year making it the sixth leading cause of cancer death in the United States. Within this year more than 50,000 individuals will be diagnosed with RCC and around 12,000 will die from this disease. Out of these RCCs the most frequently diagnosed form is clear cell renal cell carcinoma (ccRCC) which accounts for approximately 80% of all RCCs. We have previously identified through genomic profiling that loss of the type II and III TGFβ receptors in the progression of this disease lead to loss of TGFβ responsiveness and suggests a role for dysregulated TGFβ signaling in RCC progression and metastasis. Restoration of the TGFβ receptor type III (TβRIII) leads to apoptosis in RCC cell lines (Margulis et al. 2007) suggesting that re-expression of TβRIII is an attractive method for intervention of ccRCC. Recently we identified that loss of TβRIII is an early event in ccRCC and that this loss is not through epigenetic silencing but due to the loss of transcriptional regulation. The transcription factor GATA3 was identified to be a positive regulator of TβRIII expression through its actions on the proximal promoter of TβRIII. Through deletion analysis we have now identified a region within the proximal promoter that contains a number of putative transcription factor binding sites that are capable of negatively regulating the expression of TβRIII. We have identified that one of these transcription factors is over-expressed in ccRCC patient matched renal tissue samples and that silencing of this transcription factor through lentiviral shRNA, leads to a three fold induction of endogenous TβRIII in ccRCC cell lines. Furthermore, loss of this transcription factor in ccRCC cell lines leads to decreased cell proliferation when compared to non-target infected controls. Further evaluation of these putative negatively regulating transcription factors and their actions upon the proximal promoter of TβRIII are being analyzed utilizing EMSA and ChIP assays. These investigations will identify novel molecular targets that through their dysregulation will have the potential to up-regulate TβRIII in ccRCC inducing apoptosis within these cells. Together these findings will allow for a deeper molecular understanding of the events taking place in ccRCC.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 2173. doi:10.1158/1538-7445.AM2011-2173
Abstract 2174: Progesterone receptor, AP-1 and Stat3 cooperative transcriptional complex in breast cancer
Authors: Díaz Flaqué María C. , Béguelin Wendy , Vicario Rocio , Proietti Cecilia , Rivas Martín , Tkach Mercedes , Izzo Franco , Charreau Eduardo , Schillaci Roxana , Elizalde Patricia
Published: June 2012

We have previously demonstrated that progestin induces the rapid activation of AP-1 transcription factor and its nuclear interaction with the progesterone receptor (PR) by tethering in C4HD cells an experimental model of hormonal carcinogenesis in which the synthetic progestin medroxyprogesterone acetate (MPA) induced mammary adenocarcinomas in female Balb/c mice and in the human breast cancer line T47D. In this work, we demonstrated that MPA induces AP-1 transcriptional activation through the activation of MAPKs signaling pathways. Our recent findings have demonstrated that MPA induces the recruitment of Stat3 and PR to the GAS site in the cyclin D1 promoter. Here, we identified a novel PR/AP-1/Stat3 transcriptional complex in breast cancer cells. Cyclin D1 is a cancer-related protein whose promoter contains AP-1 binding sites (TRE sites) and Stat3 binding sites (GAS sites) in a very close proximity and lacks progesterone response elements (PRE sites). By using quantitative chromatin immunoprecipitation (ChIP) and sequential ChIP, we demonstrated that progestin induces the recruitment of c-jun, c-fos and PR to the TRE site and this recruitment is abolished when we pretreat the cells with the MAPKs inhibitor U0126. When cells were transfected with c-jun and c-fos Dominant Negatives Forms (TAM-67 and A-Fos respectively) none of these proteins are able to activate cyclin D1 promoter or are recruited to the TRE site and there is not MPA-induced Stat3 recruitment to the GAS site. In the same way when we pre-treat the cells with siRNAs against Stat3 neither c-jun, c-fos, PR or Stat3 are recruited to the AP-1 or GAS siteS in the cyclin D1 promoter. Inhibition of MPA-induced AP-1 transcriptional activation by transfection of breast cancer cells with TAM-67 and A-Fos resulted in complete abrogation of progestin-induced in vitro and in vivo breast cancer growth. Our findings reveal a novel MPA-induced PR/AP-1/Stat3 cooperative transcriptional complex in breast cancer cells. Interestingly the blockage of any of the members of the complex prevents its assembly and inhibits the MPA-induced cyclin D1 up-regulation and cell proliferation.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 2174. doi:10.1158/1538-7445.AM2011-2174
Abstract 2186: Characterization of a novel ubiquitin E3 ligase that targets tumor suppressor RASSF1A for degradation during mitosis
Authors: Jiang Lingyan , Rong Rong , Sheikh M. Saeed , Huang Ying
Published: June 2012

Ras association domain family 1 isoform A (RASSF1A) is an important tumor suppressor. Loss or altered expression of RASSF1A has been associated with pathogenesis of a variety of cancers. RASSF1A is known to play an important role in regulation of mitosis, however, little is known about how RASSF1A is regulated during mitotic-phase of the cell cycle. In the present study, we have identified Cul4A as a RASSF1A E3 ligase and DDB1 serves as an adaptor protein to bring RASSF1A into the Cul4A-DDB1 complex. We have found that depletion of Cul4A E3 ligase results in increased levels of RASSF1A protein due to increased half-life; whereas overexpression of the Cul4A ligase markedly enhances RASSF1A protein ubiquitination resulting in reduced RASSF1A levels. We have further shown that Cul4A-mediated RASSF1A degradation occurs during mitosis and depletion of the E3 ligase markedly reverses mitotic-phase-stimulated RASSF1A degradation. Thus, our present study demonstrates that Cul4A is important for regulation of RASSF1A during mitosis and it may contribute to inactivation of RASSF1A thereby promoting cell cycle progression.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 2186. doi:10.1158/1538-7445.AM2011-2186
Abstract 2187: Protocadherin 10, a novel tumor suppressor, suppresses tumorigenesis and liver metastasis of colorectal cancer cells in mice
Authors: Jao Tzu-Ming , Tsai Ming-Hong , Weng Wei-Ting , Chang Chia-Yun , Lio Hoi-Yan , Lai Yen-Chun , Yu Sung-Liang , Yang Ya-Chein
Published: June 2012

Colorectal cancer (CRC) is one of the leading causes of cancer death in the world. By loss of heterozygosity approach, we identified Protocadherin 10 (PCDH10) gene with a high frequency of allelic deletion in CRC. Furthermore, the genetic alteration was associated with presence of distant metastasis and poorer overall survival of patients. Importantly, the expression of PCDH10 mRNA was silenced or downregulated in 37 (84.1%) out of 44 colorectal tumors as compared with their paired normal mucosa. We proposed that PCDH10 might be a CRC-associated tumor suppressor gene, and then investigated the biological functions of PCDH10 in CRC cells. First of all, re-expression of PCDH10 in HCT116 cells reduced cell proliferation, migration, invasion and anchorage-independent growth in vitro. Meanwhile, stable PCDH10- expressing cells (HCT116/PCDH10) exhibited suppression of tumorigenicity and liver metastasis in xenograft tumor models. In SCID mice, HCT116/PCDH10 cells with subcutaneous injection showed reduced tumor growth, and, with intrasplenic injection, showed reduced metastatic incidence and nodules in the liver, as compared with mock control cells. Taken together, these results indicate that PCDH10 is a tumor suppressor gene associated with CRC, and its downregulation might promote tumor progression and distant metastasis. In addition, allelic loss of PCDH10 gene could serve as a molecular predictor of metastasis and poor prognosis in patients with CRC.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 2187. doi:10.1158/1538-7445.AM2011-2187