bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 2619: Potential advantages of CUDC-101, a multi-targeted HDAC, EGFR and HER2 inhibitor, on preventing drug resistance and tumor metastasis
Authors: Wang Jing , Selmi Kader , Samson Maria , Forrester Jeffrey , Atoyan Ruzanna , Qu Hui , Yin Ling , Wang Da-Gong , Zifcak Brian , Borek Mylissa , DellaRocca Steven , Bao Rudi , Zhai Hai-Xiao , Cai Xiong , Qian Changgeng , Savagner Pierre , Lai Cheng-Jung
Published: June 2012

CUDC-101 is a novel small-molecule multi-target anti-cancer agent currently in Phase Ib clinical trial, which targets histone deacetylase (HDAC), epidermal growth factor receptor (EGFR) and human epidermal growth factor receptor2 (HER2). Previously, we reported that CUDC-101 has potent anti-proliferative and pro-apoptotic activities in cultured tumor cells and in vitro xenograft models when compared to other single-target agents. Additionally, we demonstrated that CUDC-101 reduces the levels of phosphorylated and total MET. MET amplification and secondary EGFR mutation are two validated mechanisms responsible for EGFR tyrosine kinase inhibitor (TKI) drug resistance in patients. In an effort to expand on our previous findings, we further substantiate here that cancer cells harboring MET amplification are sensitive to CUDC-101. This result, together with the finding that cancer cells containing the EGFR-T790M mutation are sensitive to CUDC-101, suggests that RTK inhibitor-resistant tumor cells are sensitive to treatment with CUDC-101. Additionally, erlotinib-resistant HCC827 cells, which lost their EGFR dependence, are still sensitive to treatment with CUDC-101. Interestingly, this drug-resistant cell line shows neither secondary EGFR mutation nor c-Met amplification, suggesting that CUDC-101 has the potential to overcome drug resistance through other mechanisms. Because c-Met plays an important role in metastasis, we further investigated the effect of CUDC-101 in regulating cell motility, and demonstrate that CUDC-101 can induce E-cadherin accumulation in MDA-MB-231 cells and inhibit EGF induced epithelial-mesenchymal transition (EMT) in in vitro cultured cell and in vivo animal models. In vitro functional study also shows that CUDC-101 reduces tumor cell migration and invasion. Together, our findings suggest that CUDC-101, an investigational anti-cancer drug, may provide great potential for simultaneously overcoming tumor growth, metastasis, and drug resistance.

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 2619. doi:10.1158/1538-7445.AM2011-2619
Abstract 2623: ST3595, a new histone deacetylase (HDAC) inhibitor endowed with a peculiar mechanism of action
Authors: Pisano Claudio , Vesci Loredana , Milazzo Ferdinando M. , Guglielmi Mario B. , Barbarino Marcella , Foderà Rosanna , Zuco Valentina , Zunino Franco
Published: June 2012

The novel hydroxamic acid-based compound ST3595, was identified within an in vitro screening for its inhibitory activity on histone deacetylase's (HDAC's) (Pisano C. et al., Clin Cancer Res 2010, 16 (15): 3944-53; Dalla Valle S et al., Eur J Med Chem 2009, 44 (5): 1900-12). With aim to define its antitumor potential, we investigated the in vivo activity of ST3595 in a variety of tumor models. In addition, we performed a comparative study of ST3595 and vorinostat (SAHA) to better understand the cellular/molecular basis of the antiproliferative activity. In in vivo experiments, ST3595, delivered orally or i.p. at well-tolerated doses (100 mg/kg) induced a significant tumor growth inhibition (ranging from 57 to 87%) in a variety of human tumor xenografts (H460 non-small cell lung, HCT116 colon and A2780 ovarian carcinoma). In in vitro studies, ST3595 exhibited arrest of H460 cells (wt p53) in S phase associated with a large extent of apoptosis. Under conditions producing comparable antiproliferative effects, treatment with vorinostat resulted in a G2/M arrest with a marginal induction of apoptosis. In HT29 cells (mutant p53) ST3595 caused arrest in S and G2/M phases with apoptosis induction, whereas vorinostat induced G1 accumulation without evidence of apoptosis, thus suggesting only antiproliferative effects. To explore the basis of the different cellular effects, we investigated the modulation of various factors implicated in DNA damage response and repair, cell cycle control and regulation of apoptosis. In H460 cells, we found that, in contrast to vorinostat, ST3595 was an effective inducer of p53 acetylation, supporting a role of this p53 modification in the proapoptotic activity of ST3595. In addition, in contrast to vorinostat, ST3595 strongly affected the expression of critical proteins involved in cell cycle control and DNA damage response. In particular, ST3595 induced activation and phosphorylation of H2AX, and of the checkpoint kinases chk1 and chk2, thus supporting an interference of the HDAC inhibitor in the processes controlling DNA integrity. In the light of the use of broad-specificity HDAC inhibitors in cancer therapy and in combination with other antitumor agents, the ability of ST3595 to modulate DNA damage response and it proapoptotic efficacy may have relevant implications in enhancing antitumor activity of well-established cytotoxic agents. Preclinical studies of ST3595 with genotoxic agents support this interpretation.

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 2623. doi:10.1158/1538-7445.AM2011-2623
Abstract 2626: Thiazole antibiotic/proteasome inhibitor thiostrepton against cancer
Authors: Pandit Bulbul , Wang Ming , Halasi Marianna , Gartel Andrei L.
Published: June 2012

Thiazole antibiotic, thiostrepton was identified as an inhibitor of oncogenic transcription factor FoxM1, later demonstrated to exhibit proteasome inhibitory activity. We found that polymeric micelle-encapsulated thiostrepton reduced tumor growth rate in MDA-MB-231-luc breast cancer xenografts. Encapsulation of thiostrepton into polymeric micelles can aid its solubilization and increase its accumulation into tumor sites. Furthermore, its anti-cancer effects on breast cancer xenografts were found to be through reducing cell proliferation and inducing cell death. Thiostrepton is sulfur containing highly modified macrocyclic antibiotic with a central pyridine/tetrapyridine/dehydropiperidine ring with up to three thiazole substituents on positions 2, 3 and 6, which has macrocyclic loop connecting thiazole rings at position 2 and 3 described as ring A. In addition, it has a quinaldic acid macrocycle also connected to thiazole on position 2 described as ring B. Structural modification of thiostrepton to thiostrepton methyl ester (with open B ring) did not demonstrate proteasome inhibitory activity. These data suggest that B ring of thiostrepton and similar thiazole antibiotic Siomycin A that is absent in other thiazole antibiotics determines the proteasome inhibitory activity of these drugs. We investigated the therapeutic potential of the combination of thiostrepton and proteasome inhibitor bortezomib (Velcade) on various human tumor cell lines. Combination of sublethal concentrations of thiostrepton and bortezomib induced potent apoptosis and inhibition of long-term colony formation in a wide variety of human cancer cell lines. The synergistic realtionship between thiostrepton and bortezomib combination was also quantitatively demonstrated by low combination index between 0.1 and 0.8. The synergy between these drugs was based on their proteasome inhibitory activities of both drugs, because structurally similar thiostrepton modification, thiostrepton methyl ester that did not have proteasome inhibitory activity failed to increase apoptosis in combination with bortezomib

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 2626. doi:10.1158/1538-7445.AM2011-2626
Abstract 263: MYC enforces overexpression of EZH2 in early prostatic neoplasia via transcriptional and posttranscriptional mechanisms
Authors: Koh Cheryl M. , Iwata Tsuyoshi , Zheng Qizhi , Bethel Carlise , Yegnasubramanian Srinivasan , De Marzo Angelo M.
Published: June 2012

EZH2 is part of the PRC2 polycomb repressive complex that is overexpressed in multiple cancer types and has been implicated in prostate cancer initiation and progression. Here we identify EZH2 as a target of the MYC oncogene in prostate cancer and show that MYC coordinately regulates EZH2 through transcriptional and post-transcriptional means. Although prior studies in prostate cancer revealed a number of possible mechanisms of EZH2 upregulation, these changes cannot account for the overexpression of EZH2 in many primary prostate cancers, nor in most cases of high grade PIN. We report that upregulation of MYC in the mouse prostate results in overexpression of EZH2 mRNA and protein which coincides with reductions in miR-26a and miR-26b, known regulators of EZH2 in some non-prostate cell types, albeit not in others. Further, in human prostate cancer cells, MYC negatively regulates miR-26a and miR-26b via direct binding to their parental Pol II gene promoters, and forced overexpression of miR-26a and miR-26b in prostate cancer cells results in decreased EZH2 levels and suppressed proliferation. In human clinical samples, miR-26a and miR-26b are downregulated in most prostate cancers. In a novel mechanism of EZH2 mRNA upregulation, we find that MYC binds directly to and activates the transcription of the EZH2 promoter. These results link two major pathways in prostate cancer by providing two additional and complementary MYC-regulated mechanisms by which EZH2 upregulation occurs and is enforced during prostatic carcinogenesis. Further, the results implicate EZH2-driven mechanisms by which MYC may stimulate prostate tumor initiation and disease 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 263. doi:10.1158/1538-7445.AM2011-263
Abstract 2633: Histone deacetylase inhibitors induce CXCR4 mRNA but antagonize CXCR4 migration
Authors: Ierano Caterina , To Kin Wah , Robey Robert W. , Bates Susan E. , Scala Stefania
Published: June 2012

The stromal cell-derived factor-1α SDF-1α (CXCL12)/CXCR4 axis has a critical role, particularly in renal cancer where CXCR4 expression has been linked to poor prognosis. Histone deacetylase inhibitors (HDIs) exert antitumor effects by targeting both histone and nonhistone proteins affecting cell migration, apoptosis, cell proliferation and angiogenesis. Thus, the aim of the present work was to evaluate the effects of the HDIs apicidin, vorinostat, MS-275 and romidepsin on the expression and function of CXCR4 in human cancer cell lines including renal, non-small cell lung, breast, colon and glioma cell lines. After treatment with romidepsin or apicidin, CXCR4 mRNA expression increased 12-fold in UOK121 and 2.5-fold in UOK143 renal cancer cell lines; treatment with vorinostat or MS-275 yielded similar effects. CXCR4 induction was also observed in H460 non-small cell lung cancer cells, and SF-295 glioblastoma cells, but not in MCF-7 breast cancer cells or SW620 colon cancer cells. To evaluate the corresponding CXCR4 functional increase, CXCL12-induced ERK1/2, STAT-3, and c-SRC activation and cell migration were examined in UOK121, SF295 and H460 cells. In general, the HDIs increased pERK1/2, while reducing pSTAT-3 and pSRC. Following CXCL12 pERK1/2 induction was maintained but STAT3 and SRC phosphorylation was impaired. These findings resulted in reduced basal and CXCL12-mediated cell migration. In conclusion, HDIs up-regulated CXCR4 mRNA expression but impaired signalling cascades CXCL12-dependent on STAT3 and c-SRC, suggesting a role for HDI in delaying or preventing metastatic process in solid tumors.

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 2633. doi:10.1158/1538-7445.AM2011-2633
Abstract 2634: Methionine metabolites specifically protect anticancer effect of 5-FU but not for cisplatin in human lung cancer cells
Authors: Ham Myeong-Sun , Jeong Jong-Woo , Lee Ju-Kyung , Park Jin-Ah , Kim Kyeong-Ah , Kim Keun-Cheol
Published: June 2012

It has been known that cellular methylation is associated with stabilization of chromatin structure. S-adenosylmethionine (SAM) and S-adenosylhomocysteine (SAH), as metabolites of methionine metabolism, are the substrate and end product of essential cellular methyltransferase reactions, respectively. In this study, we are to explain that these metabolites specifically regulate anticancer effect of 5-FU but not for cisplatin. Using MTT assay, we have found that combination treatment of SAM or SAH with 5-FU specifically protect the anticancer effect of 5-FU in time or dose dependent manner, whereas combination treatment with cisplatin had no effect. This result was also consistent in cell cycle distribution by FACS analysis. Induction of tumor suppressor p53 by anticancer drugs was decreased in combination treatment of those metabolites with 5-FU. We hypothesized what cellular methylation related proteins involve in the protection effect. SUV39H1 and G9a, typical HMTases for H3K9, also have no change in combination treatment of those metabolites with 5-FU. However, expression of DNMT3A and DNMT3B were increased at the combination treatment of SAM and 5-FU. Interestingly, there were no change on expression of DNMT3A and DNMT3B in combination treatment of SAH with 5-FU, indicating protection effect of SAM and SAH on 5-FU differently act at the molecular level. Together, we suggest that cellular methylation can specifically modulate anticancer effect of DNA damage agent 5-FU and this can be modulated by aberrant DNA methylation.

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 2634. doi:10.1158/1538-7445.AM2011-2634
Abstract 2639: RanGAP1·SUMO1 hyperphosphorylation (RGSH) and mitotic arrest by ON 01910.Na, okadaic acid or tubulin agents
Authors: Oussenko Irina , Holland James F. , Reddy E. Premkumar , Ohnuma Takao
Published: June 2012

INTRODUCTION: RanGAP1·SUMO1 is phosphorylated before nuclear envelope breakdown when cells enter mitosis. Previous work from our laboratory showed that a new anticancer antimitotic agent, ON 01910.Na (abbreviated 1910), a benzyl styryl sulfone analog, induced RGSH in all four human tumor cell lines tested. RGSH was not observed in the presence of 1911, inactive analog of 1910. However, RGSH was observed in the presence of tubulin depolymerizing agent nocodazole. Tubulin polymerization assay revealed that, unlike nocodazole, neither 1910 nor 1911 had effects on tubulin polymerization in vitro. In this study, we examined whether other tubulin agents induce RGSH and whether known mitotic phosphatases are involved. METHODS and RESULTS: We used two cell lines, MOLT-3 ALL and DU145 prostate cancer cell lines. We examined RGSH in the presence of known tubulin toxins including paclitaxel, vinblastine, vincristine, colchicine and nocodazole at a concentration of 1 µM in 4 h and 24 h. Doxorubicin and camptothecin served as negative controls. Following treatment with drugs, cells were analyzed by Western blotting. Drug concentrations that inhibited cell growth by 50% (IC50) or 90% (IC90) were determined in 3-day drug exposure experiments. All tubulin agents in the study were capable of inducing RGSH after 4h and 24 h exposure in both cell lines. The drug-induced RGSH was inhibited by caffeine, an ATM and ATR kinase inhibitor, and roscovitine, a Cdk kinase inhibitor. Okadaic acid (OA), known primarily as PP1 and PP2A phosphatase inhibitor, was also found to be capable of RGSH, therefore making it plausible by analogy that 1910 is RanGAP1·SUMO1 phosphatase inhibitor. The concentrations of 1910, OA or tubulin agents sufficient to elicit RGSH in 24 h, also caused 50 to 90% growth inhibition, respectively. 1910, as well as all tubulin agents in the study, induced phosphorylation of the phosphatase Cdc25C (aka “mitotic” Cdc25C) and down-regulation of interphase form of Cdc25C. CONCLUSION: 1910 and selected tubulin agents caused RGSH at IC50 – IC90 concentrations. Similar to tubulin agents, 1910 produced Cdc25C phosphorylation and down-regulation of its interphase form. OA inhibition of PP1 phosphatase and activation of RGSH suggest that 1910 could also serve as a phosphatase inhibitor.

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 2639. doi:10.1158/1538-7445.AM2011-2639
Abstract 265: c-Myc and NFκB are reciprocal and contextual transcriptional regulators of glioma cell proliferation or invasion
Authors: Dhruv Harshil D. , McDonough Wendy S. , Tran Nhan , Armstrong Brock , Tuncali Serdar , Eschbacher Jenny , Kislin Kerri , Berens Michael E.
Published: June 2012

Classic histological features of glioblastoma include dense proliferative areas rich in angiogenesis as well as centripetal dissemination of neoplastic cells into adjacent brain tissue (most frequently white matter). The infiltrative dispersion patterns of malignant glioma preclude complete tumor resection; growth of satellite lesions causes significant neurological morbidity and mortality, and accounts for much of the post-treatment, recurrent (fatal) disease. Distinct transcriptomes are discernable between GBM cells at the tumor core and invasive rim, and many of the differentially-expressed genes are co-associated with migration and the collateral phenotype of cell survival; reciprocal downregulation of genes in invasive glioma cells are ontologically associated with proliferation. Our studies of glioma cells from paired core and rim human biopsy specimens reveal a higher proliferative index (Ki67 Mib-1 IHC score) at the core as compared to the rim (19 out of 35 specimens) p < 0.002. Analysis of activation states of transcription factors (Luminex multiplex assay) revealed that nuclear c-myc activity is up in the tumor core while nuclear NFκB activity is up at the invasive rim of the tumor. Depletion of c-myc (siRNA oligonucleotides) resulted in an increase in the migration rate of glioma cells in vitro, whereas inhibition of NFκB by SN50 resulted in a decrease in migration rate both in vitro and ex vivo (rat brain slice). Immunohistochemical validation using a glioma tissue microarray containing paired core and rim biopsy specimens showed that phosphorylated c-myc staining was higher in the core than in the rim for 23 out of 39 biopsy specimens scored, and that phosphorylated NFκB was higher in the rim than the core for 30 out of 43 biopsy specimens scored. The Go vs Grow hypothesis suggests cell proliferation and cell migration are temporally exclusive behaviors and tumor cells postpone cell division for migration. Our findings argue that differential suppression/activation of c-myc and NFκB underlie the shift of glioma cells from growing to going.

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 265. doi:10.1158/1538-7445.AM2011-265
Abstract 2658: Phase II study of chemoradiation therapy and erlotinib for stage III non-small cell lung cancer
Authors: Komaki Ritsuko U. , Wistuba Ignacio , Tang Xi Ming , Meyn Raymond E. , Lee J. Jack , Wei Xiong , Allen Pamela K. , O'Reilly Michael S. , Welsh James , Hong Wuan Ki
Published: June 2012

Preclinical evidence that the epidermal growth factor receptor (EGFR) inhibitor erlotinib has radiosensitizing properties in vitro led us to test its use, in combination with standard chemoradiation therapy (ChRT), in a prospective phase II study of patients with locally advanced non-small cell lung cancer. Endpoints were radiographic response, survival rates, and toxicity. We also tested if EGFR copy number or protein expression and mutations in EGFR exons 19-21 or KRAS codons 12-13, all measured in formalin-fixed paraffin-embedded biopsy specimens, predicted response to this therapy. Patients were to receive ChRT (RT=63 Gy in 35 fractions in 7 wks; Ch=weekly paclitaxel 45 mg/m2+carboplatin AUC 2) with erlotinib 150 mg po daily (except on Ch days) followed by two cycles of consolidation Ch. Of the 48 patients enrolled (30 male, 18 female; median age 64 yr; all with good performance status), 38 were evaluable for response. At a median follow-up of 11 months, the response rate was 79% (29% complete and 50% partial); 10% had stable disease and 11% progressive disease. The overall survival rate at 1 yr was 85%; 42% were alive without evidence of disease, 34% alive with disease, and 24% dead. Severe acute toxicity (grade 3 in the NCI Common Terminology Criteria v3) was manageable and included pneumonia (n=7), skin reactions (n=4), pneumonitis (n=3), and dehydration (n=3); several hematologic toxicity included leukopenia (n=13), neutropenia (n=7), and thrombocytopenia (n=1). No patients died of treatment-related causes. Of the 34 patients with biopsy specimens, only 3 showed EGFR mutation and 1 a KRAS mutation, and none correlated with response. However, complete response was more likely among current or former smokers (n=10) than never-smokers (n=0, P=0.023). We conclude that the 1-yr overall survival rate was promising but the complete response rate was disappointing. Erlotinib may have had a radiosensitizing effect in vivo, but additional patients and longer follow-up are needed.

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 2658. doi:10.1158/1538-7445.AM2011-2658
Abstract 2668: Radiosensitization of solid tumor cell lines by an MDM2 inhibitor
Authors: Cerna David , LI Hongyun , Flaherty Siobhan , Carter Donna , Coleman C. Norman , Yoo Stephen
Published: June 2012

The tumor suppressor p53 is a key molecule in the prevention of cancer. More than half of all human cancers have mutation or loss of the p53 gene. A significant percentage of the other half contain overexpression of Hdm2 which serves as a ubiquitin ligase (E3) to target p53 for degradation. While it has been established that an inhibitor of Hdm2 ubiquitin E3 ligase activity HLI373 (5-(3-dimethylaminopropylamino)-3,10-dimethyl-10H-pyrimido(4,5-b)quinoline-2,4-dione, NSC373989), selectively kills tumor cells harboring wild-type p53, we investigated the effect of the Hdm2 inhibitor HLI373, in the response to ionizing radiation in solid tumor lines expressing wild-type, mutant or null p53 in vitro and in vivo. We now report HLI373 enhances radiosensitivity in a panel of human tumor cell lines in p53 deficient and competent cell lines using clonogenic assay. HCT116 cells exposed to HLI373 (5-7.5 uM) for 16 hours before and maintained in the medium after irradiation had an increase in radiosensitivity. Cell cycle distribution of treated cells after 16 hours was measured using flow cytometry. In a comparison of wildtype and null cell lines, cell cycle arrest in G1 was only detected in cells with active p53 at 5uM of HLI373. In contrast, cells with a null or mutated p53 progressed to and arrested in G2. In addition, wild type p53 cells pretreated with 5 uM HLI373 and irradiated continued to be arrested in G1 and did not progress to a G2. However, cells without p53 treated with HLI373 and radiation increased in accumulation in G2. Treatment with higher dosages of HLI373 (>10uM) resulted in the accumulation of cells in G2 regardless of p53 status. These results indicate that HLI373 enhances tumor cell radiosensitivity in vitro and agents such as HLI373 may have potential for use in the clinics.

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 2668. doi:10.1158/1538-7445.AM2011-2668
Abstract 2708: Functional characterization of a fluorescent highly tumorigenic ovarian cancer line to test cellular therapy in experimental models
Authors: Ingersoll Susan Blaydes , Ahmad Sarfraz , Stoltzfus Gregory P. , Merchant Mohammed H. , Radi Michael J. , Finkler Neil J. , Edwards John R. , Holloway Robert W.
Published: June 2012

Introduction: Because current therapies for ovarian cancer have little impact on the long-term survival, there is a compelling need to develop innovative strategies. Our aim was to functionally characterize a fluorescent highly tumorigenic ovarian cancer line to test cellular therapy in combination with cytokines or chemotherapies in experimental models.

Methods: Two fluorescent highly tumorigenic sub-lines (SKOV3-AF1 and SKOV3-AF2) were derived from the SKOV-3 ovarian cancer cell line. Peripheral blood mononuclear cells (PBMC)-mediated cytotoxicity in combination with IL-2 and IFNα-2b of SKOV3-AF1 and -AF2 was assayed by lactate dehydrogenase release. Sensitivity of SKOV3-AF2 cells to polyethylene glycol (PEG)-IFNα-2b or IL-2 was assayed in a xenograph nude mouse model. Histopathology was performed on tumors harvested from mice to determine necrosis and tumor infiltrating lymphocytes in the solid tumors.

Results: SKOV3-AF1 and -AF2 sub-lines exhibit increased cytotoxicity mediated by PBMC, IL-2, and IFNα-2b compared to parental SKOV-3 cells. The average cytotoxicity was 59% for both SKOV3-AF1 and -AF2 compared to 28% for SKOV-3 parental cells (p<0.05). SKOV3-AF1 and -AF2 cells are more tumorigenic in vivo compared to SKOV-3 cells as indicated by tumor incidence (SKOV3 66%, SKOV3-AF1 100%, SKOV3-AF2 100%), time to sacrifice, tumor weight, and ascitic fluid production. In addition, the SKOV3-AF2 cells continue to fluoresce when harvested from the mice. SKOV3-AF2 tumor growth was inhibited by PEG-IFNα-2b but not low-dose IL-2. The average tumor weight was 0.96 g in the control treatment group that did not receive PEG-IFNα-2b versus 0.23 g for PEG-IFNα-2b (12,000 U per week); p < 0.05). There was no significant difference in any of the IL-2 treatment groups (0-4,000 U, thrice weekly). Both PEG-IFNα-2b and IL-2 were well tolerated by the mice as demonstrated by no significant weight loss during the treatment. Histopathology revealed that the tumors consisted of poorly differentiated surface epithelial carcinoma and there was no significant difference in the percent necrosis or TIL score in any of the treatment groups.

Conclusions: Characterization of SKOV3-AF2 cells revealed that these cells are more susceptible to PBMC-mediated cytotoxicity than SKOV-3 cells, are highly tumorigenic in a xenograph model, and continue to fluoresce when harvested from mice. Our data suggests that the SKOV3-AF2 sub-line will be a useful tool to test cellular therapy for the treatment of ovarian cancer utilizing experimental models.

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 2708. doi:10.1158/1538-7445.AM2011-2708
Abstract 272: Transcriptional regulation of REG4, a novel diagnostic marker of gastrointestinal cancer, by CDX2
Authors: Naito Yutaka , Oue Naohide , Sentani Kazuhiro , Sakamoto Naoya , Anami Katsuhiro , Hayashi Tetsutaro , Uraoka Naohiro , Oo Htoo Zarni , Wakamatsu Yuta , Yasui Wataru
Published: June 2012

Regenerating gene family, member 4 (REG4, encodes Reg IV), was originally identified from cDNA library of inflammatory bowel disease tissues, and is specifically expressed in gastrointestinal tract. We have identified REG4 as an overexpressed gene in gastric cancer from SAGE analysis in gastric cancer tissues and noticed as a novel serum marker of cancer. The expression of Reg IV is associated with intestinal mucin differentiation, and RegIV is expressed in CDX2-positive cells. CDX2, the intestine-specific caudal-related homeobox transcription factor, plays a key role in intestinal development and differentiation. In the present study, we investigated whether CDX2 induce RegIV expression.

Western blot analysis indicated that forced expression of CDX2 in HT29 colon cancer cell line induced Reg IV expression. In silico analysis of 2-kilobase upstream region of REG4 revealed at least four putative CDX2 consensus sequences. To assess the role of these putative CDX2 consensus sequences in regulating REG4 transcription, we performed luciferase assay into HSC39 gastric cancer cell line, in which CDX2 is highly expressed. Reporter gene constructs containing 2-kilobase of 5’-flanking region from the REG4 gene showed strong reporter activity. Significant change of reporter activity was not observed between 2.0 and 0.65-kilobase upstream region of REG4 constructs, apart from that significant lower expression level at 0.1-kilobase. These results suggest that expression of REG4 is regulated by CDX2, and upstream region of REG4 has enhancer sequences at proximal region at 0.1-kilobase upstream region. Based on the analysis of single mutation in the putative CDX2 consensus sequences in the 5’-flanking region of the REG4 gene, two putative CDX2 consensus sequences were found to activate REG4 expressions. Therefore, RegIV may be a key factor mediating CDX2 function in intestinal determination. To identify the CDX2-binding site in detail, ChIP assay and mutagenesis with multiple mutations in the putative CDX2 consensus sequences must be carried out.

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 272. doi:10.1158/1538-7445.AM2011-272
Abstract 2724: Intravesical treatment with Vesicular Stomatitis virus decreases the size of bladder tumors without adversely affecting the health and well-being of immune-competent mice
Authors: Skinner Lisa A. , Zhang Kevin , Jia William , Rennie Paul , So Alan
Published: June 2012

Recent studies have shown that wild-type Vesicular Stomatitis virus (VSV) and the Δ51M recombinant IFN-inducing mutant of wild-type VSV (AV3) have powerful oncolytic activity against bladder cancer cells in an immune-compromised bladder cancer model. The objective of the current study was to determine the safety and efficacy of the wild-type VSV and AV3 when intravesically administered to an immune-competent bladder cancer model. Using an established orthotopic bladder cancer model, 100 µL of 108 pfu mL−1 wild-type VSV, AV3, or UV-inactivated wild-type VSV (UVI) was instilled into the urinary bladders of female C3H mice. Half of the mice had normal, healthy bladders and were instilled with the oncolytic virus on day 0, 2, and 4. The remaining mice were given non-muscle invasive bladder tumors through the transurethral instillation of luciferase expressing MBT-2 mouse bladder carcinoma cells. On day 5 after initial tumor inoculation, tumor-bearing mice were imaged for bioluminescence and tumor size was quantified. On days 6, 8, and 10 after the initial tumor inoculation, tumor-bearing mice were randomized for treatment, and wild-type VSV, AV3, or UVI was instilled into each mouse. Levels of tumor bioluminescence (a correlate of tumor size), and body weight (a key variable of animal health and indicator of treatment toxicity) were monitored for 22 days. Following three consecutive intravesical treatments of an oncolytic virus, there were no differences in weight between the treated mice (wild-type VSV and AV3) and the control mice (UV-inactivated wild-type VSV), regardless of tumor presence. In addition, tumor bioluminescence in the AV3 treated mice was slightly decreased (30%) compared to the control mice, but not different from the wild-type VSV treated mice. These data correspond to in vitro data which show that wild-type VSV and AV3 are inhibitors of proliferation in MBT-2 mouse bladder carcinoma cells. In conclusion, our data strongly suggest that oncolytic viruses such as wild-type VSV and its attenuated form, AV3, can be used to treat superficial bladder cancer in immune-competent mice, without adversely affecting the health and well-being of the animal.

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 2724. doi:10.1158/1538-7445.AM2011-2724
Abstract 2732: Short inverted repeats are hot spots for genetic instability in mammalian cells
Authors: Lu Steve , Wang Guliang , Vasquez Karen M.
Published: June 2012

Analyses of chromosomal aberrations in genetic disorders such as leukemia and lymphoma have provided compelling evidence that segments of the human genome are prone to breakage that lead to DNA rearrangements. Many of these hot spot areas contain sequences that have the potential to form non-B DNA such as Z-DNA, H-DNA or cruciforms. Although research has shown that large palindromes are mutagenic in mammalian cells, the instability of short inverted repeat sequences (30 bp), which are mapped near breakpoints and quite abundant in the eukaryotic genome, has not been investigated. In order to study the effects of short cruciform-forming sequences on genetic instability, we subcloned short AT- and CG-rich inverted repeats into a lacZ’ mutation-reporter shuttle vector. The cruciform structures that formed at the palindromic sequences were probed using S1 nuclease to confirm the presence of the non-B DNA structure. Cruciform-induced mutagenesis was measured in mammalian COS-7 cells via transfection then blue-white screening in DH5α cells. Ligation-mediated PCR provided direct evidence that cruciform-forming sequences induced double-stranded breaks in mammalian cells. In vitro replication assays using SV40- HeLa cell-free extracts were employed to establish the connection between replication and cruciform-induced genetic instability. Results from our study demonstrate for the first time that cruciform structures that form at short inverted repeats are mutagenic in mammalian cells. The majority of the mutants induced by cruciforms were large-scale deletions that spanned the inverted repeat sequence. Analyses of junctions revealed that >80% of the mutants contained microhomologies which are characteristic of error-prone nonhomologous end-joining repair. This suggests that cruciform structure is cleaved by replication-independent, structure-specific nucleases in mammalian cells that result in DNA double-stranded breaks at positions directly adjacent to the inverted repeat. These results indicate that cruciform-forming sequences may play a role in genetic instability and contribute to human diseases associated with gene translocations.

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 2732. doi:10.1158/1538-7445.AM2011-2732
Abstract 2751: Fanconi anemia pathway and DNA repair-based prognostic signatures in ovarian cancer
Authors: Kang Josephine , D'Andrea Alan , Kozono David
Published: June 2012

Introduction: Of the 21,000 patients diagnosed annually in the United States with epithelial ovarian cancer, approximately 75% present with Stage III-IV disease. The vast majority of these patients are treated with cytotoxic chemotherapy regimens, where clinical response may be associated with activity of DNA repair pathways in tumors. Characterizing these repair pathways should help predict clinical response to specific chemotherapeutic agents.

Methods: We devised a set of 150 DNA repair genes with documented functional importance in key DNA repair pathways including base excision repair (BER), Fanconi Anemia (FA), homologous recombination (HR), mismatch repair (MMR), non-homologous end joining (NHEJ), nucleotide excision repair (NER) and translesion synthesis (TLS). We extracted microarray expression data for these genes for 511 tumors and 8 normal specimens using data from Agilent Custom 244K Gene Expression and Affymetrix Human Genome U133A arrays. We performed Significance Analysis of Microarrays (SAM) to determine which of the 150 DNA repair genes were significantly more expressed in tumor vs. normal specimens. We then generated Kaplan-Meier survival curves using overall survival data.

Results: Using the Affymetrix data, we identified 57 genes that were significantly up-regulated with a median fold change of 3.15 (range, 2.03-6.07) and 6 genes that were significantly down-regulated with a median fold change of 2.67 (range, 2.11-4.74) among tumor vs. normal tissue samples by SAM (delta 1.013, false discovery rate 0%). SAM using the Agilent data showed similar results; a total of 46 genes were significant hits in both datasets. A DNA repair signature consisting of genes significant in both platforms was defined and revealed a correlation between high expression values and decreased survival (median overall survival 4.5 vs. 5.2 years, log-rank p = 0.0086). DNA repair signatures specific to individual repair pathways were defined and demonstrated that high FA pathway (BRCA2, FAAP24, FANCA, FANCE, FANCG, FANCI and PALB2) expression in particular was inversely correlated with survival (median overall survival 4.1 vs. 5.2 years, log-rank p = 0.029 and 0.019 for the Affymetrix and Agilent datasets, respectively).

Conclusion: Evidence from literature supports the critical role of the Fanconi Anemia (FA) pathway in repairing DNA damage caused by platinum-based agents such as cisplatin and carboplatin, commonly utilized to treat ovarian cancer patients. We show that a DNA repair signature specific to FA can be utilized to identify patients with high FA expression, whom we hypothesize may be more resistant to traditional platinum-based therapy.

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 2751. doi:10.1158/1538-7445.AM2011-2751
Abstract 2757: Characterization of the glucuronidation of the major nicotine metabolite, 3’-hydroxycotinine: Potential role for knock-out polymorphisms in UGTs 2B17 and 2B10
Authors: Chen Gang , Giambrone Nino E. , Muscat Joshua E. , Lazarus Philip
Published: June 2012

3’-Hydroxycotinine (3HC) and its glucuronide (3HC-Gluc) conjugate are major nicotine metabolites excreted in the urine of smokers and other tobacco or nicotine users. Previous studies have demonstrated that human liver microsomes (HLM) were active in catalyzing the formation of both 3HC-O-Gluc and 3HC-N-Gluc. While not all human UDP-glucuronosyltransferases (UGTs) were previously screened, UGTs 2B7, 1A9, 2B4 and 2B15 were shown to be active in the formation of 3HC-O-Gluc and UGT1A4 active in 3HC-N-Gluc formation. In the present study, all known human UGTs except for UGT2B28 were screened for glucuronidation activity against 3HC. Homogenates from cell lines over-expressing UGTs 2B17 and 2B10 exhibited the highest O- and N-glucuronidation activities, respectively for the UGTs tested. Kinetic analysis for the formation of 3HC-O-Gluc demonstrated that the KM for UGT2B17 is 3.7- and 4.2-fold lower than that observed for UGTs 1A9 (p<0.005) and 2B7 (p<0.005); the KM of 3HC-N-Gluc formation was 4.4-fold lower (p<0.05) for UGT2B10 than that observed for UGT1A4. Kinetic analysis demonstrated that HLM from subjects exhibiting the homozygous null UGT2B17 deletion [UGT2B17 (*2/*2)] exhibited a KM for 3HC-O-Gluc formation that was 3.1-fold higher (p<0.001) than that observed in HLM from subjects exhibiting the wild-type UGT2B17(*1/*1) genotype. The KM of 15.6 μM for 3HC-O-Gluc formation in UGT2B17 (*2/*2) HLM was close to that observed for UGT2B17 (8.3 μM). 3HC-N-Gluc formation activity was decreased by 91% (p<0.001) in HLM from subjects with the UGT2B10(*2/*2) knock-out genotype and 39% (p<0.001) in HLM from subjects with the UGT2B10(*1/*2) genotype as compared to that observed for UGT2B10(*1/*1) HLMs. The KM of 13 μM for 3HC-N-Gluc formation in UGT2B10 (*1/*1) HLM (n=6) was close to that observed for UGT2B10 (13 μM). These results demonstrate that UGTs 2B17 and 2B10 play key roles in the hepatic glucuronidation of 3HC and that functional polymorphisms in UGT2B17 and UGT2B10 are associated with significantly reduced glucuronidation activities against 3HC in HLM.

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 2757. doi:10.1158/1538-7445.AM2011-2757
Abstract 2785: Modulating the interaction between cdk2 and cyclin A: Screening, synthesis and characterization of a quinoline-based inhibitor
Authors: Windsor William T. , Deng Yongqi , Shipps Gerald W. , Zhao Lianyun , Siddiqui Arshad , Popovici-Muller Janeta , Curran Patrick , Fischmann Thierry , Duca Jose S. , Hruza Alan , Zhang Rumin , McNemar Charles , Mayhood Todd , Syto Rosalinda , Parry David A. , Lees Emma , Kirschmeier Paul
Published: June 2012

Cyclin dependent kinase 2 (CDK2) is aberrantly regulated in a wide range of cancers and small molecule inhibitors should be beneficial in the treatment of cancer. Preclinical and clinical CDK2 inhibitors are ATP competitive and bind at the ATP site in a canonical mode. A novel binding mode for a new class of quinoline-based inhibitors of CDK2 has been discovered using the affinity-based Automated Ligand Identification System (ALIS) technology. While the inhibitors do bind to the ATP site the key mechanism of action for inhibition is the induction of a novel protein conformational change that enables the compound to bind into the core of the kinase domain. Global changes to CDK2 upon compound binding result in a conformation that can no longer bind to its activating protein cyclin A. Therefore, these inhibitors are both ATP competitive and are protein-protein interaction disrupters. The key strategy for discovering this class of protein-protein disrupter compounds was to screen the unphosphorylated monomer CDK2 in ALIS and to perform secondary binding and enzyme mechanism of action assays that identified quinoline-based compounds that only bound to the inactive CDK2 monomer and not the active CDK2/cyclin A heterodimer. Through a series of chemical modifications the affinity (Ki) of the original hit improved from 0.9 μM to 0.005 μM. The screening strategy utilized and the discovery of this new kinase inhibitor binding mode may be applicable to other kinase programs.

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 2785. doi:10.1158/1538-7445.AM2011-2785
Abstract 2795: An onco-protein axis linking polycomb repressive complex 2 and polycomb repressive complex 1 through miRNAs in cancer
Authors: Cao Qi , Mani Ram , Ateeq Bushra , Dhanasekaren Saravana M. , Asangani Irfan , Yu Jindan , Prensner John , Kim Jung J. , Brenner J. Chad , Cao Xuhong , Jing Xiaojun , Wang Rui , Li Yong , Dahiya Arun , Wang Lei , Lonigro Robert , Tomlins Scott , Palanisamy Nallasivam , Maher Christopher , Varambally Sooryanarayana , Chinnaiyan Arul M.
Published: June 2012

Enhancer of Zeste Homolog 2 (EZH2) is the catalytic histone methyltransferase subunit of the Polycomb Repressive Complex 2 (PRC2), that trimethylates histone H3 at lysine 27 (H3K27me3) resulting in the silencing of target genes. PRC2 plays a critical role in many basic cellular processes including cell proliferation, differentiation, early embryogenesis, and X chromosome inactivation. In cancer, EZH2 upregulation is implicated in metastasis and tumor aggressiveness of prostate and breast cancer and several other solid tumors. Recently our lab reported the genomic loss of miR-101 microRNA accompanying EZH2 overexpression in tumor cells. Here we identified several microRNAs that were downregulated by EZH2 and their levels were restored upon EZH2 depletion in cancer cell lines, and expression levels of these microRNAs were negatively correlated with EZH2 in human prostate tumors. Additionally, H3K27me3 modification was observed in the upstream regions of the miRNAs, suggesting a direct role for EZH2 in their regulation. Ectopic overexpression of the miRNAs suppressed cell proliferation, invasion, anchorage-independent growth, sphere formation and xenograft tumor growth of aggressive prostate and breast cancer cell lines. Finally, our investigations showed that the miRNAs also repress the expression of Polycomb Repressive Complex 1 (PRC1) members BMI1 and RING2, leading to a global decrease in the epigenetic marker, ubiquityl-H2A-K119 (uH2A) in cells, a key step in PRC1-mediated silencing. Our findings provide compelling argument for a regulatory axis joining PRC2 and PRC1 through miRNAs. This novel link between PRC2 and PRC1 indicates a coordinated mechanism by polycomb group proteins to promote an aggressive cancer phenotype.

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 2795. doi:10.1158/1538-7445.AM2011-2795