bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 3507: Facilitating pixantrone-induced DNA damage and subsequent cell kill through formaldehyde activation and improvements in drug design
Authors: Evison Benny J. , Mansour Oula C. , Sleebs Brad E. , Watson Keith G. , Rephaeli Ada , Nudelman Abraham , Phillips Don R. , Cutts Suzanne M.
Published: February 2011

Pixantrone is a novel aza-anthracenedione originally developed to improve the therapeutic profile of mitoxantrone, a DNA-interactive agent currently used in the clinical management of a range of haematological malignancies and solid tumours. Like mitoxantrone, pixantrone interacts with DNA via intercalation and stimulates topoisomerase II-mediated DNA cleavage, presumably by stabilising the cleavable complex. Despite the drug's ability to stimulate DNA cleavage via topoisomerase II impairment, this form of DNA damage does not directly correlate with drug cytotoxicity, suggesting that pixantrone may be operating by a distinct, currently undefined mechanism of cell kill. A novel form of pixantrone-DNA interaction has recently emerged in which the drug can be efficiently activated by formaldehyde to yield covalent drug-DNA adducts. In cellular systems, pixantrone exhibits a mild synergistic relationship with the formaldehyde-releasing prodrug AN-9, indicating a favourable role for formaldehyde in mediating pixantrone-induced cell kill. A variety of methods including in vitro transcription and mass spectrometry have established that formaldehyde-activated pixantrone is a monofunctional DNA alkylator that binds selectively to CpG and CpA dinucleotides via the exocyclic N2 amino group of guanine within DNA. Crucially, formaldehyde provides the methylene bridge that covalently links DNA with a single drug side-chain. A major functional limitation of the monoadduct is its poor intrinsic stability (t1/2 = 75 min). It was subsequently rationalised that an increase in drug side-chain length may permit the drug to completely bridge the two strands of duplex DNA, much like an interstrand crosslink. Accordingly, new anthracenediones were synthesised that incorporated symmetrical side chains of the nature -NH-(CH2)n-NH2 (where n = 2—5). An in vitro crosslinking assay demonstrated that side-chain extension generally conferred a remarkable increase in the temporal stability of formaldehyde-activated anthracenedione-DNA adducts. A clear and direct relationship existed between side-chain length and drug-DNA adduct half-life with MX5 (n = 5) exhibiting exceptional stability (t1/2 > 2 days). Significantly, the enhanced drug-DNA adduct stability was reflected in breast adenocarcinoma MCF-7 cells where derivatives with longer side-chains were synergistic in combination with the formaldehyde-releasing prodrug AN-9.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3507.
Abstract 3508: A diazirine-based photoaffinity etoposide probe for labeling topoisomerase II
Authors: Chee Gaik-Lean , Yalowich Jack , Hasinoff Brian B.
Published: February 2011

Etoposide is a widely used anticancer drug that targets topoisomerase II, an essential nuclear enzyme. However, despite the fact that it has been in use and studied for more than 30 years the specific site on the enzyme to which it binds is unknown. In order to identify the etoposide binding site(s) on topoisomerase II, a diazirine-based photoaffinity etoposide analog probe has been synthesized and its photoreactivity and biological activities have been characterized. Upon UV irradiation, the diazirine probe rapidly produced a highly reactive carbene species that formed covalent adducts containing stable carbon-based bonds indicating that it should also be able to form stable covalent adducts with amino acid residues on topoisomerase II. The human leukemia K562 cell growth and topoisomerase II inhibitory properties of the diazirine probe suggest that it targets topoisomerase II in a manner similar to etoposide. The diazirine probe was also shown to act as a topoisomerase II poison through its ability to cause topoisomerase IIα-mediated double-strand cleavage of DNA. Additionally, the diazirine probe significantly increased protein-DNA covalent complex formation upon photoirradiation of diazirine probe-treated K562 cells, as compared to etoposide-treated cells. This result suggests that the photoactivated probe forms a covalent adduct with topoisomerase IIα. In conclusion, the present characterization of the chemical, biochemical, and biological properties of the newly synthesized diazirine-based photoaffinity etoposide analog indicates that use of a proteomics mass spectrometry approach will be a tractable strategy for future identification of the etoposide binding site(s) on topoisomerase II through covalent labeling of amino acid residues. This work was supported by institutional grants from the University of Manitoba, Canadian Institutes of Health Research, National Institutes of Health grant CA090787 to J.C.Y. and a Canada Research Chair for Drug Development for B.B.H

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3508.
Abstract 3510: The molecular docking-based design and biological evaluation of a novel series of bisintercalating DNA-binding bisanthrapyrazole compounds containing amine linkers
Authors: Wu Xing , Zhang Rui , Guziec Lynn , Guziec Frank , Yalowich Jack C. , Hasinoff Brian B.
Published: February 2011

Anticancer drugs that bind to DNA and inhibit DNA-processing enzymes represent an important class of anticancer drugs. In order to find stronger DNA binding and more potent cytotoxic compounds, a series of bisanthrapyrazole derivatives containing amine linkers were designed and evaluated by molecular docking techniques. Because the anthrapyrazoles are unable to be reductively activated like doxorubicin and other anthracyclines, they should lack the cardiotoxicity exhibited by this class of antitumor agent. The concentration dependent increase in DNA melting temperature was used to determine the strength of DNA binding and the bisintercalation potential of the newly synthesized analogs. These compounds were also assessed for their ability to inhibit: 1) the growth of the human erythroleukemic K562 cell line; 2) the decatenation activity of DNA topoisomerase IIα; 3) the relaxation activity of DNA topoisomerase I. Finally, the compounds were evaluated as topoisomerase IIα poisons by measuring the topoisomerase IIα-mediated double strand cleavage of DNA. All of the bisanthrapyrazoles inhibited K562 cell growth in the low to submicromolar concentration range and also strongly inhibited the decatenation activity of topoisomerase IIα. However, none of the bisanthrapyrazole compounds act as topoisomerase IIα poisons. In conclusion, a novel group of bisanthrapyrazole compounds were designed, synthesized, and biologically evaluated as potential anticancer agents. Support: CIHR, a Canada Research Chair in Drug Development to B.B.H., a National Institutes of Health grant CA090787 to J.C.Y. and the Dishman Foundation at Southwestern University.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3510.
Abstract 352: Preclinical evaluation of LYS6K1, a novel, highly selective, orally bioavailable inhibitor of p70 S6 kinase currently in phase I clinical trials for cancer
Authors: Geeganage Sandaruwan , Sams Lillian L. , Joseph Sajan , Brail Leslie , Wang Tao , Rodgers George , Akunda Jackie , Huang Jianping , Perkins Everett J. , Ajamie Rose
Published: February 2011

PI3K/AKT/mTOR/S6K signaling pathway (AKT pathway) controls cell survival, cell-cycle progression, cell growth and metabolism through a cascade phosphorylation of a number of key substrates. This pathway is regulated by three well characterized tumor suppressors; pten, tsc2, and lkb1. Deletion of these genes results in activation of the AKT pathway and proliferative disorders. Similarly, activating mutations of the receptor tyrosine kinases or PI3 Kinase result in the activation of the pathway. Therefore, multiple nodes of the pathway have become drug targets. As part of a comprehensive drug discovery platform aimed at targeting the PI3K pathway, we have developed a potent small molecule inhibitor of p70 S6 kinase that is a downstream effector in this pathway.

LYS6K1 is a potent, highly selective ATP competitive inhibitor against p70 S6 kinase with an IC50 of 0.002 uM. In vitro, LYS6K1 inhibits the phosphorylation of S6 ribosomal protein in HCT116 colon cancer cells with an IC50 of 0.2 uM and similar activity is seen in a broad range of other cell lines. In vivo, LYS6K1.tosylate demonstrates potent phospho-S6 inhibition in nude mice bearing HCT116 colon carcinoma cells, with an ED50 value of 2.2 mg/kg and a ED90 value of 10 mg/kg 4 hours after a single oral dose. In these studies, LYS6K1.tosylate did not show statistically significant elevation of phospho-AKT, in spite of potent p70 S6 kinase inhibition. Proportional dose, exposure, and pharmacodynamic relationships were observed for LYS6K1 in dose- and time- dependent studies. In vivo, LYS6K1.tosylate effectively inhibits the growth of HCT116 colon carcinoma xenografts in mice and the growth of U87MG glioblastoma tumor as a single agent at 2.5 mg/kg given twice daily.

Overall, LYS6K1.tosylate has high permeability that resulted in good oral absorption and PK properties. Based on these pre-clinical observations, LYS6K1.tosylate has advanced to Phase I studies.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 352.
Abstract 3520: Pegylated nanoplatinates: A novel re-engineered cisplatin for cancer chemotherapy
Authors: Soni Shivani , Sengupta Shiladitya , Paraskar Abhimanyu , Chaudhuri Padmaparna
Published: February 2011

Cis-diamminedichloroplatinum (II) (CDDP) is a platinum based chemotherapy drug used to treat various types of cancers, including sarcomas, some carcinomas, lymphomas, and germ cell tumors. There are two major concerns of cisplatin based chemotherapy: first is relapse of cancer in majority of pateints with cisplatin resistant disease and second is cisplatin associated nephrotoxicity (kidney damage). In the present study, we have demonstrated that cisplatin can be complexed using a carboxylato bond with poly-isobutylene-maleic acid, derivatized with a polyethylene glycol moiety. We found that at unique cisplatin to polymer ratio, this conjugate self-assembles into nanoparticles, and exhibits enhanced therapeutic efficacy without any nephrotoxicity as compared with cisplatin in murine lewis lung carcinoma model. The PIMA-PEG-CDDP conjugate inhibited proliferation and induced apoptosis in Lewis lung carcinoma and 4T1 breast cancer cells in vitro. The nanoparticles were taken up by cancer cells through endocytosis within 6 hours and retained in subcellular compartment over 24 hours in 4T1 breast cancer cells. Furthermore, in an in vivo 4T1 breast cancer model, PIMA-PEG-CDDP nanoparticles reduce tumor growth drastically with no side effects, thus increasing the survival rate of the mice as compared to free cisplatin treated mice. Thus our studies showed that pegylated-PIMA-CDDP can be harnessed for lung and breast cancer chemotherapy with increased therapeutic index compared with cisplatin.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3520.
Abstract 3530: OATP1B3 in pancreatic cancer tissue as a potential target for anticancer drug transport
Authors: Obaidat Amanda , Hagenbuch Bruno
Published: February 2011

Organic Anion Transporting Polypeptides (OATPs) are multispecific transporters that are expressed in normal and cancerous tissue (e.g. colon, prostate, breast and bone cancers). Several anticancer drugs have been shown to be transported by certain OATPs. The aim of the study was to determine the expression of OATPs in pancreatic cancer tissue and functionally characterize anticancer drug transport via the expressed OATPs. Ten pancreas tissue samples were obtained for analysis: 6 normal pancreas, 3 pancreatic cancer and 1 metastatic tissue. Expression of OATPs was analyzed at the mRNA level using the Quantigene assay. Protein expression and localization were determined by immunofluorescence with OATP specific antibodies. Transport of the radiolabeled anticancer drugs paclitaxel and docetaxel was measured in an OATP-expressing Chinese Hamster Ovary (CHO) cell system. Among the 11 human OATPs, expression in pancreatic cancer tissue at the mRNA level was highest for OATP4A1 followed by OATP3A1, OATP2A1 and OATP1B3. Expression of OATP4A1, OATP3A1 and OATP1B3 was confirmed at the protein level. In addition, transport of paclitaxel and docetaxel was observed in OATP1B3-expressing CHO cells with Km values of 2.2 μM for both substrates. In summary, OATP4A1, OATP3A1 and OATP1B3 are expressed in pancreatic cancer. OATP1B3 transports the anticancer drugs paclitaxel and docetaxel with relatively high affinities. In conclusion, given that OATP1B3 is exclusively expressed in normal human hepatocytes, which are generally resistant to the effects of anticancer therapy, transport of anticancer drugs via this transporter might be a feasible approach to target the killing of OATP1B3-expressing cancer cells.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3530.
Abstract 3535: Retinoblastoma cerebrospinal fluid metastases treatable by intensive intraventricular and multidrug resistance-reversal systemic chemotherapy without radiation
Authors: Chan Helen S. L. , Héon Elise , Doyle John J. , Dimaras Helen , Gallie Brenda L.
Published: February 2011

PURPOSE: Despite combining chemotherapy with peripheral stem cell transplant, cerebrospinal fluid (CSF) metastasis is the most difficult type of retinoblastoma (RB) metastasis to cure, since most drugs that can be given safely into the CSF are anti-leukemia agents that are ineffective against solid tumors. Most metastatic RB cells also express P-glycoprotein causing multidrug resistance (MDR). Drugs do not readily enter into the central nervous system because of the blood-brain barrier formed by P-glycoprotein-rich blood vessels. High-dose craniospinal radiation is too morbid for treatment of young children. Radiation also greatly increases the secondary malignancy risk for patients with germline RB1 mutations. To cure CSF metastases without craniospinal radiation, we designed an intensive multimodality chemotherapy regimen.

METHOD: Since 2000, we treated 4 children with CSF metastases: Child A, with germline bilateral RB, and optic nerve involvement (1 eye enucleated but no radiation); Child B, with germline trilateral RB (no enucleation and no surgery or radiation for the huge suprasellar tumor); Child C, with germline trilateral RB (1 eye enucleated but no surgery or radiation for the huge suprasellar tumor); and Child D, with nongermline unilateral RB with orbital and intracranial recurrence (only localized orbital radiation). All children were treated with 6 or 7-cycle high-dose carboplatin and etoposide, standard-dose vincristine, and high-dose/3-hour-infusion cyclosporine to inhibit both tumor-cell and blood-brain barrier P-glycoprotein. Intraventricular drugs, cytarabine, which is not a substrate of P-glycoprotein but effective against RB cell line in vitro, and topotecan, which is less susceptible to MDR, contributed to treatment of the CSF metastases. On achieving significant responses, supralethal-dosage carboplatin, etoposide and cyclophosphamide were used for consolidation, with the bone marrow rescued with autologous peripheral blood stem cells.

RESULTS: Child A and C are relapse-free 9.1 and 2.3 years post-diagnosis (8.6 and 1.8 years post-transplant). Child B and D died of disease 2.7 and 1.8 years post-diagnosis (2 and 1.4 years post-transplant).

CONCLUSION: Two of the 4 children with CSF metastases, who in the past would have died, are now surviving after intensive multimodality chemotherapy without radiation.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3535.
Abstract 354: PI-103 and TCN as proliferative inhibitors of the RTK/PI3K/ATK signaling pathways in astrocytomas
Authors: Connell-Albert Yvette , Gursel Demir , Tuskan Robert , Anastassiadis Theonie , Hawes Jessica , Van Schaick Jessica , Reilly Karlyne
Published: February 2011

INTRODUCTION: Glioblastomas (GBMs), the most aggressive of malignant astrocytomas, are recurrent, infiltrative, and fatal. In GBMs, receptor tyrosine kinases (RTKs) are often mutated and activated, leading to the dysregulation of normal cell signaling pathways. The PI3K/Akt1/mTOR pathway, which is important in normal biological events such as proliferation, survival, invasion, migration and angiogenesis, is often deregulated in malignant cancers, resulting from the simultaneous loss of tumor suppressor PTEN, and the hyperactivation of the Akt pathway. Therefore, pharmacological inhibition of the PI3K/Akt1/mTOR may prove beneficial in arresting the proliferation of astrocytomas and glioblastomas.

METHODS: In order to determine if the selected inhibitors PIA-6, OSU03012, Rapamycin, Tricribine (TCN), and PI-103 had an effect on the PI3K/Akt-1/mTOR pathway, we examined their inhibitory effects on mouse astrocytoma cell lines K1861-10 Grade II; KR158 Grade III; K130G#3 Grade IV; human astrocytoma cell lines, U87MG and SF295, both Grades IV; and normal proliferating mouse primary astrocytes using the Alamar blue assay as a measure of metabolic activity and proliferation. Inhibitors with low IC50 values in tumor cells as compared to normal primary astrocytes were favored.

RESULTS: From the select panel of inhibitors, only the AKT inhibitor TCN and dual PI3K/mTor inhibitor PI-103 showed low IC50 values in the nM or pM range, suggesting strong inhibition of proliferation in the astrocytoma cell lines tested. Rapamycin, an mTOR inhibitor, showed inconsistent inhibition of cell proliferation on the cell lines tested, although mTor activity was effectively inhibited, suggesting that inhibitory effects of rapamycin at higher concentrations may be acting via different pathways.

CONCLUSIONS: Our results support the hypothesis that the PI3K/Akt1/mTOR pathway is a potential druggable target in astrocytomas and glioblastomas.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 354.
Abstract 3541: Pharmacological chaperones increase membrane expression of ABCG2 harboring the Q141K single nucleotide polymorphism
Authors: Tamaki Akina , Caterina Ierano , Shukla Suneet , Robey Robert W. , Ambudkar Suresh V. , Bates Susan E.
Published: February 2011

ABCG2 is an ATP-binding cassette half-transporter that functions as a xenobiotic transporter. Since ABCG2's discovery in multidrug-resistant cancer cells, the transporter has garnered interest in pharmacogenomics as a component of the blood-brain barrier and the maternal-fetal barrier as well as a modulator of oral drug absorption. A single nucleotide polymorphism (SNP) C421A in ABCG2, resulting in a glutamic acid to leucine mutation at amino acid 141 (Q141K), has been extensively studied and has been found to affect protein expression and ATPase activity, but may also affect protein folding. As misfolded ABCG2 has been shown to be degraded through the endoplasmic reticulum associated protein degradation (ERAD) system (T. Furukawa et al., Pharm Res., 26(2): 469-479, 2008), we postulated that pharmacological chaperones such as mitoxantrone, a known substrate of ABCG2, might increase surface expression of Q141K ABCG2. In this study, we used HEK293 stably transfected with Q141K ABCG2 to examine whether incubating cells expressing the mutated transporter with mitoxantrone or other ABCG2 substrates would serve to “rescue” Q141K ABCG2 and improve trafficking to the cell surface, as we have demonstrated in other mutations that impair trafficking (O. Polgar et al., Biochemistry, 43(29): 9448-9456, 2004). Expression of ABCG2 in cells transfected with wild-type or Q141K ABCG2 was measured by immunoblot analysis and flow cytometry. After treatment with 5 μM of mitoxantrone, cell surface expression of ABCG2 in cells transfected with vector containing Q141K ABCG2 increased, while cells transfected with wild-type ABCG2 showed no change. Increased ABCG2 expression was also observed in membrane preparations from mitoxantrone-treated cells transfected with Q141K ABCG2 compared to wild-type ABCG2 transfected cells. Interestingly, immunoblot analysis showed that when cells transfected with Q141K ABCG2 were incubated with 50 μM of the dietary compounds tert-butyl hydroquinone (TBHQ) and dibenzoylmethane (DBM), increased expression of ABCG2 was observed in membrane preparations, while no change was observed for cells expressing wild-type ABCG2. These results suggest that some dietary compounds may promote proper folding and membrane localization of mutant forms of ABCG2.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3541.
Abstract 3542: Analysis of miRNA mediated Cisplatin resistant mechanisms
Authors: Suzuki Toshihiro , NIshio Kazuto , Togawa Tadayasu , Sugawara Kanako , Sakuraba Hitoshi
Published: February 2011

Cisplatin is an effective anticancer agent that is widely used in the treatment of testicular, ovarian, bladder and lung cancers. However, the development of resistance to cisplatin is a major obstacle to treatment. To understand the clinically relevant mechanisms of resistance, many studies have been aimed at clarifying the biochemical/molecular alterations of cisplatin-resistant cells. However, these studies did not conclusively identify the basis of cellular resistance to cisplatin.

Recently some studies have shown that micro RNAs play an important role in cancer cell resistance to chemotherapeutic agents and cancer development.

In this study, micro RNA expression profiles of cisplatin resistant non small cell lung cancer cell lines were analyzed using microarray and real time RT-PCR. We found some of micro RNAs were similarly regulated in couple of cisplatin resistant cell lines. These finding are useful for understanding the miRNA mediated cisplatin resistance mechanisms. Now, functional analyses on miRNAs involving cisplatin resistance are in progress.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3542.
Abstract 3545: Alterations of molecular mechanisms in newly selected 5-fluorouracil resistant pancreatic cancer cell line and the effect of biochanin A thereon
Authors: Bhardwaj Vikas , Lai James C.K. , Bhushan Alok
Published: February 2011

Resistance to drugs is one of the major contributor to the high mortality rate of pancreatic cancer. The molecular mechanisms of pancreatic cancer drug resistance are not well understood. We generated and characterized a 5FU resistant pancreatic cancer cell line (AsPC1/R). 5FU resistant pancreatic cancer cells (AsPC1/R) were selected with the increasing concentration of the drug till 6-fold resistance was achieved. Our dose response studies show that the resistant cells were cross-resistant to the gefitinib; however, the sensitivity to gemcitabine was not altered. Western blot analysis shows lower level of EGFR in the AsPC1/R as compared to AsPC1 cells. In addition, our results indicate that the resistant cells have high levels of activated Erk 1 and Akt. Levels of c-fos and c-jun in the nuclear fraction were upregulated in the resistant cells. Furthermore, invasive mechanisms such as uPAR and MMP activity are upregulated in the AsPC1/R cells compared to the parent cells. We also determined the effect of biochanin A on 5FU resistance and found that it reverses the 5FU resistance in AsPC1/R cells. Our previous studies have shown that biochanin A reduces the levels of thymidylate synthase and the levels of activated Erk and Akt. We conclude that biochanin A may overcome 5FU resistance in pancreatic cancer and could be used in combination with other chemotherapeutic agents in treating pancreatic cancer.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3545.
Abstract 3548: Role of synuclein-gamma (SNCG) in paclitaxel resistance and mitogen activated protein kinase activation in uterine papillary serous carcinoma(UPSC)
Authors: Chapman-Davis Eloise , Patel Parin A. , Kim J. Julie , Buttin Barbara
Published: February 2011

Objectives: SNCG is a novel prognostic biomarker in UPSC. We have shown that its overexpression correlates with aggressive cellular properties and resistance to paclitaxel in a UPSC cell line (SPEC2) as well as poor prognosis in a pilot set of UPSC patients. The goal of this study was to determine the mechanism of SNCG-associated taxane resistance by evaluating the effect of SNCG on the mitotic checkpoint and its propensity to modulate extracellular signal-regulated protein kinase 1/2 (ERK 1/2) signaling pathways.

Methods: A stably transfected SPEC2 cell line was created using shSNCG versus shControl oligonucleotides. Similarly, Ishikawa cells were transiently transfected with an SNCG expression vector versus empty vector. Differences in cell proliferation were measured using a cell viability assay and BrdU incorporation assay after treatment with paclitaxel. Cell cycle analysis was used to study the effect of SNCG on the mitotic checkpoint. Differences in expression of the checkpoint kinase BubR1 due to SNCG were studied using real time PCR, Western blot, 26S ubiquitin-proteasome inhibition, and siRNA to BubR1. ERK and phospho-ERK levels were measured in SPEC2 cells with and without SNCG expression by Western blot. SPEC2 cells were treated with paclitaxel with and without the MEK inhibitor UO126 and apoptosis was measured using cleaved PARP analysis.

Results: SNCG overexpression lead to acquired, reversible paclitaxel resistance in endometrial cancer cell lines. SNCG was noted to interfere with mitotic checkpoint function by causing degradation of the critical checkpoint protein BubR1 thereby allowing cells to escape paclitaxel-induced mitotic arrest. SPEC2 cells exhibited sustained activation of ERK which was attenuated by silencing SNCG. When SPEC2 cells were exposed to paclitaxel combined with the MEK inhibitor UO126, decreased cell viability and increased apoptosis was noted compared to either agent alone. SNCG knockdown further enhanced paclitaxel and UO126 sensitivity. Cell cycle analysis revealed that silencing SNCG in SPEC2 cells and adding UO126 acted synergistically in overcoming paclitaxel resistance.

Conclusions: SNCG overexpression in UPSC leads to taxane resistance by interacting with the mitotic checkpoint kinase BubR1 causing its degradation. SNCG is also associated with sustained ERK activation. Targeting SNCG in conjunction with MEK inhibition may represent a novel therapeutic strategy to overcome taxane resistance in these cells.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3548.
Abstract 3549: miR-451 links CRC stem-like cells with chemoresistance
Authors: Bitarte Nerea , Boni Valentina , Zarate Ruth , Gonzalez-Huarriz Marisol , Lopez Ines , Rodriguez Javier , Bandres Eva , Garcia-Foncillas Jesus
Published: February 2011

Introduction: In recent years, an emerging role for microRNAs (miRNAs) in human cancer has been proposed. Our group has previously identified miR-451 as a potential tumor suppressor microRNA involved in cell cycle progression and radiation response. We hypothesized whether miRNAs may be involved in drug resistance mechanisms among cytotoxic agents commonly used in metastatic colorectal cancer (mCRC).

Experimental Design: miR-451 was overexpressed by ectopically synthetic pre-miR-451 transient transfection and by generating stable-miR-451 overexpressed cells through retroviral vectors. The role of miR-451 on chemosensitivity to 5-fluorouracil (5FU), oxaliplatin (LOHP) and SN38, an active metabolite of irinotecan, was characterized by MTS. We sought to determine links between miR-451 and cancer stem cells (CSCs) by colonspheres generation, soft-agar cell invasion and xenograft in vivo experiments. Down-stream targets involved in drug-resistance mechanisms were also identified.

Results: Overexpression of miR-451 significantly increased the sensitivity of CRC cell lines to SN38. The IC50 values were significantly lower in pre-miR-451-transfected cells, including DLD1, LoVo and RKO than in controls (950 nM versus 460 nM for DLD1; 10 nM versus 1.4 nM for LoVo and 20 nM versus 5.9 nM for RKO). No changes were detected for 5-FU and LOHP response. ATP-binding cassette transporter protein 1 (ABCB1) proved to be miR-451-target directly associated with SN38 sensitivity. Moreover, restoring miR-451 levels translated into a reduced tumorigenicity in a xenograft model and a decreased ability of CRC cells to growth under anchorage-independent conditions in soft agar.

These results suggest that miR-451 down-regulation in CRC cells may be involved in CSCs generation. Indeed, overexpression of miR-451 was able to reduce the number and size of CRC cells-derived colonspheres. The phenotype of formed colonspheres was confirmed evaluating overexpression of several putative CSCs markers (CD133, CD44, EpCam and CD166). We also found colonspheres to be more resistant to SN38 treatment and to express higher levels of ABCB1 than parental cells. Finally, miR-451 restoration levels reverted CSCs resistance to SN38 through ABCB1 down-regulation.

Conclusions: Our findings suggest that miR-451 is involved in SN38 resistance through suppression of ABCB1 and regulation of CRC stem-like cell phenotype. Therefore, miR-451 may represent a novel potential candidate target to circumvent drug resistance in CRC.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3549.
Abstract 355: CIP2A mediates effects of bortezomib on phospho-Akt and apoptosis in hepatocellular carcinoma cells
Authors: Chen Kuen-Feng , Liu Chun-Yu , Lin Yu-Chin , Yu Hui-Chuan , Liu Tsung-Hao , Hou Duen-Ren , Chen Pei-Jer , Cheng Ann-Lii
Published: February 2011

Previously, we reported that Akt inactivation determines the sensitivity of hepatocellular carcinoma (HCC) cells to bortezomib (Chen et al, Cancer Res 2008). Here, we report that cancerous inhibitor of protein phosphatase 2A (CIP2A), a cellular inhibitor of protein phosphatase 2A (PP2A), mediates the apoptotic effect of bortezomib in HCC. Silencing PP2A by small interference RNA (siRNA) abolishes bortezomib-induced down-regulation of phospho-Akt and apoptosis. Bortezomib increases PP2A activity in sensitive HCC cells, including Sk-Hep1, Hep3B and Huh-7 but not in resistant PLC5 cells. Bortezomib down-regulates CIP2A in a dose- and time-dependent manner in all sensitive HCC cells whereas no alterations in CIP2A were found in resistant PLC5 cells. Knockdown of CIP2A by siRNA restored bortezomib's effects on apoptosis and PP2A activity in PLC5 cells. Moreover, over-expression of CIP2A up-regulated phospho-Akt and protected Sk-Hep1 cells from bortezomib-induced apoptosis. Importantly, ectopic expression of CIP2A decreased Akt-related PP2A activity whereas silencing CIP2A increased this activity, indicating that CIP2A negatively regulates Akt-related PP2A activity in HCC cells, Furthermore, our in vivo data showed that bortezomib down-regulates CIP2A and up-regulates PP2A activity in Huh-7 tumors but not in PLC5 tumors. In conclusion, inhibition of CIP2A determines effects of bortezomib on apoptosis and PP2A-dependent Akt inactivation in HCC. CIP2A may be a biomarker for predicting clinical response of bortezomib in HCC treatment. Supported by NTUH98P03, NTUH98FTN21, NSC97-2314-B-002-182, NSC98-2314-B-002-067-MY3, and NSC98-3112-B-002-037.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 355.
Abstract 3550: Role of TNFα in the cytotoxicity of docetaxel and in docetaxel resistance in MCF-7 cells
Authors: Sprowl Jason A. , Parissenti Amadeo
Published: February 2011

Taxanes such as paclitaxel and docetaxel are widely used in cancer chemotherapy regimens due to their ability to stabilize microtubules and to arrest cell division in mitosis (Chazard, 1994). While the events leading to mitotic arrest are well understood, the mechanism by which the drug induces cytotoxicity is unclear. Paclitaxel has previously been shown to promote secretion of TNFα in macrophages; however this has not been observed in other cell lines (Bogdan, 1992). TNFα induces cell death in many cell lines through interaction with its receptor TNFR1 (Hsu, 1996). TNFα also binds to a second receptor, TNFR2, which does not possess a death domain. TNFR2 does however retain the ability to induce expression of NFκB-dependent survival genes.

Our laboratory group recently selected MCF-7 cells for survival in increasing concentrations (doses) of paclitaxel or docetaxel (MCF-7TAX and MCF-7TXT, respectively). Resistance was only achieved at or above selection dose 9 (3.33nM docetaxel) (Hembruff et al., 2008). MCF-7 cells were also “selected” in the absence of these drugs to control for changes simply associated with extended cell propagation. The ABC transporter inhibitor cyclosporin A either had no effect on docetaxel sensitivity at selection dose 9, or partially restored sensitivity at selection dose 12 (Hembruff, 2008). These observations suggest there are likely additional mechanisms of docetaxel resistance at selection doses 9 through 12.

Interestingly, we observed that docetaxel concentrations between 1.11nM and 3.33nM induced a >100-fold increase in TNFα secretion from MCF-7 cells, which likely contributes to cytotoxicity. Consistent with this view, a neutralizing TNFR1 antibody decreased cytotoxicity of docetaxel in MCF-7 cells. We also discovered that MCF-7TXT and MCF-7TAX cells at or above selection dose 9 were resistant to TNFα-induced cell death. Moreover, MCF-7TXT cells displayed downregulation of TNFR1 upon the acquisition of resistance to taxanes. Increased levels of TNFα were observed within the media of MCF-7TXT cells selected to dose levels 9 and 10, but not dose levels 11 or 12. We further provide evidence that the increased levels of TNFα in the medium of MCF-7TXT cells contributes to increased cell survivability through a TNFR2-dependent activation of NFκB and its resulting increase in the expression of cell survival genes.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3550.
Abstract 3556: New curcumin analogues are cytotoxic towards estrogen receptor negative human breast cancer cell lines
Authors: Yadav Babasaheb D. , Taurin Sebastien , Larsen Lesley , Rosengren Rhonda J.
Published: February 2011

There is an urgent need for the development of a safe and efficacious drug therapy for the treatment of estrogen receptor negative breast cancers due to their high malignant nature and poor prognosis. In this study, a series of new heterocyclic monoketo curcumin analogues were synthesized and screened for anticancer activity. Among twenty eight analogues screened in estrogen receptor negative human breast cancer cell-lines MDA-MB-231 (ER-/HER2-), MDA-MB-468 (ER-/HER2-) and SKBr3(ER-/HER2+), the two most potent compounds were designated as RL-71 and RL-66. Cytotoxicity was assessed by sulforhodamine B (SRB) assay. Furthermore, the compounds were studied for their ability to modulate cell cycle progression and induce apoptosis. The mechanism of these drugs was also examined by determining protein expression by Western blot analysis. The results showed that RL-71 and RL-66 had sub micromolar EC50 values in all the cell lines and the cytotoxicity was concentration and time dependent. Mechanistic analyses demonstrated that RL-71 at 1µM and RL-66 at 2µM increased the proportion of MDA-MB-231 cells in G2/M-phase by 162.02 ± 1.53% and 134.12 ± 2.17% over control (P<0.001) respectively. In addition, the treatment with RL-71 (1µM) and RL-66 (2µM) increased the proportion of apoptotic MDA-MB-231 cells by 10-fold and 4-fold over control (p<0.001), respectively. Similar results were also produced in SKBr3 and MDA-MB-468 cells. Furthermore, Western blot analysis confirmed the effect of drugs on different cancer related genes involved in cell proliferation and apoptosis. Thus, these data suggest that the novel curcumin analogues RL-71 and RL-66 are potent antitumor drugs and have significant potential for further drug development for estrogen receptor negative breast cancer.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3556.
Abstract 3560: Evaluation of RNA-stress granules formation as an indicator of response to Darinaparsin in cancer cell lines
Authors: Kolobova Elena , Larocca M. Cecilia , Goldenring James R.
Published: February 2011

Darinaparsin (ZIO101) is a novel organic arsenic compound demonstrating clinical efficacy and a favorable safety profile in hematological cancers and solid tumors. Its mechanism of action includes disruption of mitochondrial functions and induction of apoptosis. We studied the ability of darinaparsin to induce RNA stress granules, a critical survival adaptation mechanism for cells under stress. Darinaparsin caused a stress granule response similar to sodium arsenite, but at lower concentrations. Treatment of cells with darinaparsin induced the formation of granules containing a number of proteins specifically associated with stress granules, including G3BP (the Ras-GAP SH3 domain binding protein), CCAR1 (Cell Cycle and Apoptosis Regulatory Protein-1), caprin-1 (Cytoplasmic Activation/Proliferation-associated protein1) and AKAP350A (AKAP9). However, darinaparsin led to development of a greater number of smaller granules compared to sodium arsenite-induced stress.

We evaluated the effect of 0.5 mM arsenite or 0.05 mM darinaparsin on poorly differentiated uterine sarcoma (MES-SA) and the multiple drug resistant cell line MES-SA/Dx5 that was established from MES-SA cells. MES-SA/Dx5 cells express high levels of mdr-1 mRNA and P-glycoprotein. Both sodium arsenite and darinaparsin led to stress granule formation in both cell lines. However, MES-SA/Dx5 cells were extremely sensitive to darinaparsin treatment and exhibited a prominent loss of cell contact. The amount of cleaved PARP (a marker of apoptosis) determined at 48h and 72h after darinaparsin treatment was consistent with the immunofluorescence findings in that cleaved PARP was significantly elevated in MES-SA/Dx5 cells compared to MES-SA cells. This indicates enhanced cytotoxic effects of darinaparsin on this otherwise chemotherapeutic resistant cell line.

We also evaluated the effect of darinaparsin and sodium arsenite on different leukemia cell lines including Jurkat (T-cell lymphoblastic leukemia derived), Oci and THP1 (acute myeloid leukemia derived). THP1 cells express wild type nucleophosmin (NPM), while Oci cells are heterozygous for its mutant form NPMc+. Interestingly, we found that Jurkat cells did not developed stress granules in response to darinaparsin treatment and this cell line showed accelerated toxicity compared with THP1 and Oci cells, which did form stress granules. These studies demonstrate that darinaparsin has a different spectrum of activity than sodium arsenite and can elicit differential toxicity on specific classes of tumor cells.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3560.
Abstract 3562: Characterization of a novel small molecule inhibitor with potent anticancer activity
Authors: Chun Gina , Lee Young Bok , Kim Deog Joong , Cho Won-Jea , Lee Sang Kook , Chung Hwa-Jin , Ahn Chang-Ho
Published: February 2011

A series of isoquinolineamine small molecule derivatives were synthesized and screened as potential anticancer agents. Among them, we identified one isoquinolineamine analogue with potential anticancer activity. This compound inhibited the proliferation of a variety of cancer cells derived from many human solid tumors with IC50 values ranging from 14 nM to 71 nM. The compound also was more effective against gemcitabine and cisplatin-resistant cancer cell lines when compared to the original cytotoxic cancer drugs. In mice bearing tumor xenografts, treatment with the compound significantly inhibited the growth of tumors, enhanced tumor regression, and also prevented the growth of tumors in a paclitaxel-resistant xenograft model. This compound is well tolerated and had no effects on body weight compared to control animals. Mechanistic studies showed that cancer cells treated with this compound resulted in the elevation of cytochrome c in the cytosol, the decrease of Bcl-2 and the activation of Caspase 9. Further studies showed inhibition of the expression of several signaling molecules important for cell proliferation (p-Erk, p-p38 and p-Akt) and also decreased protein levels of p-GSK3α/β and β-catenin. Therefore, our result suggests that this compound could be a promising antitumor agent.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3562.
Abstract 3564: Anti-breast cancer activity of NIM811, a nonimmunosuppressive cyclosporine A analogue
Authors: Zeng Xianke , Harrington Katie , Clevenger Charles
Published: February 2011

Cyclophilin A (CypA) is a member of the immunophilin family of peptidyl prolyl isomerases (PPI) that catalyze the cis-trans interconversion of proline imide bonds of peptides. In this role, cyclophilins have been found to function as signaling switches, regulating the activity of receptors, kinases and transcription factors. The PPI activity of CypA is inhibited by the immunosuppressive drug cyclosporine A (CsA), and in turn the CypA-CsA complex inhibits calcineurin-mediated NFAT activation. Our laboratory has recently demonstrated that CypA is necessary for the prolactin (PRL)-induced activation of Jak2/Stat5 signaling and gene expression (Ca Res 68:7769, 2008). In addition, we have also shown that CsA inhibits the in vitro and in vivo growth and progression of both ER+ and ER- breast cancer cell lines. Here, we show that the non-immunosuppressive CsA analog NIM811 blocked PRL-stimulated activation of the Jak2/Stat5 pathway, and the kinases AKT and MAPK in T47D cells. NIM811 also inhibited ER+, ER-, and Her2+ breast cancer cell proliferation, survival and anchorage independent growth in a dose-dependent manner. NIM811 also blocked the PRL-induced association between Stat5 and cMyb, and at higher doses, triggered PARP cleavage. In summary, these results indicate that non-immunosuppressive cyclophilin inhibitor NIM811 has significant potential as a novel for breast cancer therapeutic.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3564.
Abstract 3565: Novel inhibitors of mPGES-1 exhibit anti-tumor activity in colorectal and lung cancers
Authors: Chang Hui-Hua , Song Zuohe , Tripp Tina , Wisner Lee , Gokhale Vijay , Hulme Chris , Meuillet Emmanuelle J.
Published: February 2011

Chronic inflammation is associated with 25% of all cancers. In the inflammation-cancer axis, prostaglandin E2 (PGE2) is one of the major players involved. Prostaglandin E synthase (PGES) is an enzyme downstream of the cyclooxygenases (COXs) in the PGE2 biosynthesis pathway. Among the three PGES isoforms, microsomal prostaglandin E synthase-1 (mPGES-1) is inducible by proinflammatory stimuli and constitutively expressed in colon, lung, and gastric cancers. The potential role for this enzyme in tumorigenesis has also been reported. Since inhibition of COXs may lead to some side effects due to the global reduction of other key prostaglandins, targeting downstream mPGES-1 might be a better therapeutic or chemopreventive strategy for cancers. In order to identify novel small molecule inhibitors of mPGES-1, we have screened in silico a small library of compounds. As a result, 14 lead compounds were identified by molecular docking simulations of the chemicals based on the crystal structure of mPGES-1. These compounds were tested for their ability to inhibit cell growth, induce apoptosis and decrease cytokine-stimulated PGE2 production in various colorectal and lung cancer cell lines. Using surface plasmon resonance (SPR) spectroscopy, we show the binding of the compounds PGE001 and PGE011 to human mPGES-1 recombinant protein with good affinity (KD at low micromolar range). These two compounds also showed the ability to reduce the IL-1beta-induced PGE2 production in HT29 and SW837 colon cancer cells and A549 lung cancer cell line with EC50 at low micromolar range. Furthermore, the compounds exhibit significant anti-tumor activity in the SW837 xenograft mouse model with T/C (relative size of treated and control tumors)=40.3% for compound PGE001 and T/C=68.7% for compound PGE011 (P<0.05), when injected i.p. at 200 mg/kg for 5 and 10 days, respectively. When tested daily i.p. for 5 days at 100mg/kg in A549 lung cancer xenografts, the PGE001 showed anti-tumor activity with T/C=67% (P<0.05). In summary, our data suggest that the identified compounds exert their anti-tumor activity through selective binding and inhibition of the enzyme mPGES-1 in the PGE2 synthesis pathway.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3565.