bluuebunny/crossref_metadata_2025 huggingface dataset:
Abstract 711: The expression of human equilibrative nucleoside transporter 1 (hENT1) is associated with overall survival in advanced biliary tract carcinoma (BTC) treated with gemcitabine
Authors: Murata Akihiro , Amano Ryousuke , Kimura Kenjiro , Yamada Nobuya , Nakata Bunzo , Nagahara Hisashi , Noda Eiji , Kubo Naoshi , Tanaka Hiroaki , Muguruma Kazuya , Yashiro Masakazu , Maeda Kiyoshi , Ohira Masaichi , Ishikawa Tetsurou , Hirakawa Kosei
Published: August 2012

[Introduction] The outcome of patients with unresectable and postoperative recurrent biliary tract carcinoma (BTC) is mostly miserable and most patients are possible candidates for palliative chemotherapy. Gemcitabine is a pyrimidine nucleoside analogue that is commonly used chemotherapeutic agent for advanced BTC. Gemcitabine is transported into the cell mostly by human equilibrative nucleoside transporter 1(hENT1). The hENT1 expression has been demonstrated to play an important role in predicting clinical outcome after gemcitabine chemotherapy for several types of cancer. The aim of the present study was to investigate a predictive marker for good prognosis of gemcitabine chemotherapy for unresectable and postoperative recurrent BTC. [Materials and methods] The analysis was performed on samples from 25 patients with unresectable (6 patients) and postoperative recurrent (19 patients) BTC treated with gemcitabine at the host institute between January 1997 and January 2011. The hENT1 expression levels in tumors were evaluated by immunohistochemistry. The clinical and histopathological variables were analyzed to evaluate predictive values for survival. [Results] Of the 25 tumor specimens, 18 (72%) specimens had positive hNET1 immunostaing, while 7 (28%) specimens were classified as negative. No statistical significant differences were found between the expression of hENT1 and patient characteristics (gender, age, tumor stage, lymph node stage, lymph duct invasion, vascular invasion, perineural invasion). In the univariate analysis, hENT1 expression was significantly correlated with overall survival (OS). The median OS was 15.3 versus 4.2 months, respectively in patients with positive versus negative hENT1 staining (p=0.006). As a result of multivariate analysis, hENT1 expression was useful as a predictive marker for good prognosis with significant difference (p<0.001). [Conclusions] Patients with positive hENT1 expression in BTC have a significantly longer survival after gemcitabine chemotherapy than patients with negative hENT1 expression. Our results suggest that hENT1 expression in advanced BTC patients treated with gemcitabine is a prognostic predictive indicator and is useful to decide gemcitabine chemotherapy.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 711. doi:1538-7445.AM2012-711
Abstract 712: Parf-1A (Partner of ARF isoform 1A) promotes oxaliplatin resistance and is a new prognostic marker of survival in pancreatic ductal adenocarcinoma
Authors: Muniz Viviane P. , Zhang Xuefeng , Reed Sara M. , Tompkins Van S. , Smith Tarik , Hagen Jussara KF , Fitzgerald Matthew , Button Anna , Smith Brian , Zamba Kokou D. , Domann Frederick E. , Mezhir James J. , Weydert Jamie , Askeland Ryan A. , Quelle Dawn E.
Published: August 2012

Pancreatic ductal adenocarcinoma (PDAC) is an incurable malignancy with ineffective treatments and dismal median survival. The INK4a/ARF locus, which encodes the alternative reading frame (ARF) tumor suppressor, is commonly inactivated in PDAC tumors. We recently discovered Parf-1A, a novel “Partner of ARF” whose role in ARF signaling and tumorigenesis is not known. Parf-1A is highly expressed in the normal pancreas and microarray databases suggest Parf expression is altered (both up- and down-regulated) in human PDAC tumors. We generated Parf-1A specific antibodies and examined its protein levels by immunohistochemistry in PDAC tumors from patients who underwent resective surgery. Parf-1A expression was altered in 72% of tumors (33% reduced, 39% elevated) compared to adjacent normal ductal tissue. Tumors with the highest Parf-1A levels were significantly associated with poor patient outcome (median survival 6 months) while patients with undetectable Parf-1A in tumors had a dramatically extended lifespan (median survival 59 months, p=0.0037). Notably, Parf-1A knockdown in cultured PDAC cells increased the p53-independent growth inhibitory activity of ARF. Parf-1A loss also sensitized PDAC cells to oxaliplatin, a chemotherapeutic agent used in combination with gemcitabine to treat PDAC patients. This work identifies Parf-1A as a new inhibitor of ARF p53-independent activity that promotes PDAC chemoresistance in vitro and is a novel prognostic marker of survival in PDAC patients.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 712. doi:1538-7445.AM2012-712
Abstract 723: Detection of TIMP3 promoter hypermethylation in salivary rinse as an independent predictor of local recurrence-free survival in head and neck cancer
Authors: Sun Wenyue , Zaboli David , Wang Hao , Liu Yan , Arnaoutakis Demetri , Khan Tanbir , Khan Zubair , Koch Wayne , Califano Joseph
Published: August 2012

Objective: To validate a panel of methylation-based salivary rinse biomarkers (P16, CCNA1, DCC, TIMP3, MGMT, DAPK, and MINT31) previously shown to be independently associated with poor overall survival and local recurrence in a larger, separate cohort of patients with head and neck squamous cell carcinoma (HNSCC). Methods: One hundred ninety-seven patients were included. All pre-treatment saliva DNA samples were evaluated for the methylation status of the gene promoters by quantitative methylation-specific PCR. The main outcome measures were overall survival, local recurrence-free survival and disease-free survival. Results: In univariate analyses, the detection of hypermethylation of CCNA1, MGMT, and MINT31 was significantly associated with poor overall survival; the detection of hypermethylation of TIMP3 was significantly associated with local recurrence-free survival; and the detection of hypermethylation of MINT31 was significantly associated with poor disease-free survival. In multivariate analyses, detection of hypermethylation at any single marker was not predictive of overall survival in patients with HNSCC; detection of hypermethylation of TIMP3 in salivary rinse had an independent, significant association with local recurrence-free survival (Hazard Ratio, 2.51, 95% CI, 1.10 to 5.68); and none of the studied markers was significantly associated with disease-free survival. Conclusions: The detection of promoter hypermethylation of the seven genes in salivary rinse as an independent prognostic indicator of overall survival in patients with HNSCC was not validated. Detection of promoter hypermethylation of TIMP3 in pretreatment salivary rinse is independently associated with local recurrence-free survival in patients with HNSCC and may be a valuable salivary rinse biomarker for HNSCC recurrence. Such a test could potentially refine our ability to identify HNSCC patients at a high risk for recurrence.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 723. doi:1538-7445.AM2012-723
Abstract 725: eIF4E/4eBP-1 as predictive factor(s) of response to EGFR targeted drug in head and neck cancer
Authors: Vergez Sébastien , Tavergnier Christelle , Rochaix Philippe , Filleron Thomas , Vagner Stéphan , Canal Pierre R. , Etienne Chatelut , Delord Jean-Pierre , Allal Ben
Published: August 2012

Purpose: There is a clinical need to identify predictive markers of the response to epidermal growth factor receptor (EGFR) inhibitors in patients with head and neck squamous cell carcinoma (HNSCC). In breast cancer, the resistance to targeted therapies against HER2 implicates the reactivation of the PI3K/Akt/mTOR pathway and eIF4E overexpression. The axis eIF4E/4eBP-1 is one of the major downstream effector of the PI3K/Akt/mTOR pathway. The aim of this study was to further investigate these markers in the response to preclinical and clinical EGFR inhibitors used in HNSCC, cell lines and patients. Experimental Design: We studied the effects of cetuximab using human HNSCC cell lines (CAL33 and CAL166) on proliferation, survival pathways, eIF4E and 4eBP activities and expression by western blot or immunohistochemistry in initial biopsies of patients treated for an HNSCC with cetuximab used in combination with radiotherapy or cisplatin (n=28). Results: The HNSCC cell lines have an opposite effect under cetuximab treatment. Whereas the CAL33 are not sensitive, the CAL166 shows a moderate cytostatic effect allowing us to determine an IC20 of 5,56 +/− 0.37 µg/mL. In the CAL33 cell lines cetuximab doesn't significantly modify the 4eBP phosphorylation level. On the contrary in CAL166 cetuximab induces the inhibition of the 4eBP phosphorylation level corroborating the cytostatic effect previously observed. More over, in patients the combination Cetuximab / radiotherapy or cisplatin shows, in the initial biopsie examination in the non-responder group, a significant higher level of eIF4E. Conclusion: The couple eIF4E expression levels and/or activities appeared to be determinant to discriminate the response to the EGFR targeted inhibitor in HNSCC.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 725. doi:1538-7445.AM2012-725
Abstract 729: Plasma EBV DNA concentration and clearance rate as good prognostic factors for metastatic nasopharyngeal carcinoma
Authors: Hsu Cheng-Lung , Huang Shiang-Fu , Chang Kai-Ping , Wang Hung-Ming
Published: August 2012

Purpose: To investigate the pretreatment copy number and the clearance rate of plasma EBV DNA as novel prognostic outcome markers for metastatic nasopharyngeal carcinoma (NPC) Experimental Design: Nineteen newly diagnosed and fifty-four relapsed metastatic NPC patients were treated at outpatient department (i.v. or i.v. plus oral chemotherapy followed by local irradiation boost where applicable). Multiple blood samples were collected before and during the first 6 weeks of chemotherapy. Plasma EBV DNA concentrations and half-life values of plasma viral clearance rates, t1/2, were determined by a real-time quantitative PCR. The association between the copy number of plasma EBV DNA, t1/2, and clinical outcome were analyzed. Results: Treatment response evaluated after 3-6 months of treatment showed the best response of 18 complete responses (CR)(24.7%), 21 partial responses (PR)(28.8%) and overall response rate was 53.5%. The pretreatment plasma EBV DNA concentrations and the t1/2 of plasma EBV DNA clearance rates had significant effects on treatment response and overall survival prediction. In the chemotherapy regimen, gemcitabine plus cisplatin had better treatment outcome than cisplatin plus oral UFT and calcium folinate based regimen. Multivariate analysis revealed both pretreatment EBV DNA load and t1/2 of plasma EBV DNA clearance rates but not chemotherapy regimen had significant effects on overall survival. Conclusions: The pretreatment plasma EBV DNA copy number and their clearance rates are significant predictors for NPC treatment outcome. In low copy number group, the t1/2 in the first 6 weeks may provide the physician to choose a tailored regimen for the patients.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 729. doi:1538-7445.AM2012-729
Abstract 732: Overexpression of the basic helix-loop-helix protein DEC1 induces cellular senescence and is related to better survival in esophageal squamous cell carcinoma
Authors: Xu Qing , Ma Peiqing , Hu Chenfei , Chen Lechuang , Xue Liyan , Wang Zaozao , Liu Mei , Zhu Hongxia , Xu Ningzhi , Lu Ning
Published: August 2012

Objective: Human differentiated embryo chondrocyte expressed gene 1 (Dec1), also known as Bhlhb2/Stra13/Sharp-2, is a basic helix-loop-helix transcription factor that involves in cell proliferation, differentiation, apoptosis, and cellular senescence. In this study, the expression, function, diagnosis and prognostic impact of DEC1 in normal esophageal epithelia, esophageal squamous cell carcinoma (ESCC) and its precursor lesions (intraepithelial neoplasia) were investigated. Methods: ESCC combined with adjacent precursor tissues from 241 patients were collected to construct tissue microarray (TMA) blocks. DEC1 expression among normal epithelia, introepithelial neoplasia and ESCC was analyzed by immunohistochemistry (IHC), and the correlations between Dec1 and clinicopathological parameters were also analyzed statistically. In vitro study, DEC1 was transient transfected into ESCC cell line EC9706, and then cellular senescence was analyzed by senescence associated β-galactosidase (SA-α-Gal) activity. Fresh esophagectomy tissues from ESCC patients were embedded and fixed, and tissue sections were detected by immunohistochemistry of DEC1 and SA-α-Gal activity. Results: Compared with normal epithelia, DEC1 expression was significantly increased in intraepithelial neoplasia and DEC1 expression was significantly decreased in ESCC in comparison with intraepithelial neoplasia. DEC1 overexpression both induced cellular senescence in EC9706 cell line and correlated to more senescent cells in fresh tissue sections. Kaplan - Meier method analysis revealed that DEC1 expression levels were significantly correlated with the survival of ESCC patients after surgery. The expression levels of DEC1 were also correlated with the tumor embolus, depth of invasion of ESCC, lymph metastasis status and pTNMs. Conclusions: These results suggest that DEC1 overexpression is a protective mechanism by inducing cellular senescence in ESCC progression, and DEC1 is a potential prognostic marker of ESCC.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 732. doi:1538-7445.AM2012-732
Abstract 741: Sunitinib plus hormone ablation and radiation therapy for patients with high-risk localized prostate cancer: Results from a multi-institutional phase I study
Authors: Corn Paul G. , Song Danny , Heath Elisabeth , Maier Jordan , Meyn Raymond , Kuban Deborah , DiPetrillo Thomas , Mathew Paul
Published: August 2012

Background: Novel strategies are needed to improve the long-term outcomes of patients (pts) with high-risk prostate cancer treated with hormonal ablation and external beam radiation therapy (XRT). Preclinical data suggest that angiogenesis inhibitors normalize vasculature, reduce hypoxia, and improve the therapeutic index of XRT. To assess the feasibility of combined VEGFR/PDGFR inhibition in combination with XRT, a Phase I study with sunitinib was initiated. Methods: Seventeen men with adenocarcinoma of the prostate with cT2c-cT4 or Gleason 8-10 or PSA > 20ng/ml received initial hormonal ablation (leuprolide 22.5mg every 12 weeks + oral bicalutamide 50mg daily) for 4-8 weeks prior to oral sunitinib 12.5, 25, or 37.5 mg daily for 4 weeks as lead-in, then concurrently with and 4 weeks post XRT (prostate and proximal seminal vesicles received 75.6 Gy in 42 fractions with at least 50 Gy for the distal seminal vesicles). A 3+3 sequential dose-escalation design was employed to assess the frequency of dose-limiting toxicity (DLT) and establish a maximal tolerated dose (MTD) of sunitinib. Results: The first 4 pts enrolled on 37.5 mg experienced a DLT during lead-in. *With 2 pts unexpectedly experiencing Grade 3 neutropenia and a third experiencing Grade 3 thrombocytopenia, a drug-interaction with bicalutamide was suspected. The protocol was revised to replace these pts and omit concurrent bicalutamide. Of these first 4 pts, 3 were dose reduced to 25 mg and successfully completed full treatment, while 1 pt withdrew. Of the next 3 pts subsequently enrolled at 37.5 mg, 2 of 3 on concurrent therapy have experienced DLTs (G3 diarrhea with concomitant sunitinib-induced thyrotoxicosis; G3 proctitis). Conclusions: Using a daily dosing regimen and a lead-in phase, the MTD of concurrent sunitinib, hormonal ablation and XRT was 37.5mg daily. The recommended Phase 2 dose of sunitinib for further study is 25mg daily.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 741. doi:1538-7445.AM2012-741
Abstract 742: Comparison of commercially available phosphorylated ERK antibodies for immunohistochemical biomarker analysis
Authors: Jeffers Michael , Dauffenbach Lisa M. , Pena Carol , Lathia Chetan , Bhargava Ajay , Kerfoot Christopher A.
Published: August 2012

Immunohistochemical (IHC) analysis of ERK phosphorylation is commonly performed on pre- and post-treatment cancer biopsies as part of pharmacodynamic biomarker assessments for targeted oncology therapeutics. The current study was performed to assess the specificity and sensitivity of four commercially-available pERK antibodies for IHC analysis, which included three monoclonal antibodies (clones 20G11, MAPK-YT, D13.14.4E) and one polyclonal antibody. To facilitate this analysis, the HEPG2 liver cancer cell line was grown in the presence or absence of inhibitors of the RAF/MEK/ERK pathway (U0126 or sorafenib). Cell lysates were prepared for analysis via Western blotting and formalin-fixed, paraffin-embedded cell pellets were prepared for analysis via IHC. All antibodies produced bands at 44 and 42 kDa, the expected molecular weights of ERK1/2, when used for Western analysis of untreated cells and demonstrated a reduction or ablation of these bands following treatment of cells with inhibitors of the RAF/MEK/ERK pathway. When examined via IHC, the monoclonals demonstrated nuclear and cytoplasmic staining in HEPG2 cells with increased sensitivity relative to the polyclonal antibody. The monoclonals also demonstrated greatly reduced staining in HEPG2 cells treated with inhibitors of the RAF/MEK/ERK pathway. These antibodies were further characterized for reactivity in human tumor specimens representing breast cancer, colon cancer, lung cancer, melanoma, and ovarian cancer. Melanoma tissues were stained with a red chromogen; all others were stained with DAB. Differences in the frequency of expression, percentage of expression, and protein localization were identified between the three monoclonals. In summary, this work has identified commercially-available antibodies that appear to be of suitable specificity and sensitivity for use in IHC assays.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 742. doi:1538-7445.AM2012-742
Abstract 752: The effect of food on the pharmacokinetics of tivozanib
Authors: Cotreau Monette M. , King Thomas , Massmanian Lisa , Strahs Andrew , Slichenmyer William , Vargo Dennis
Published: August 2012

Background: Tivozanib, a potent, selective, long-half-life tyrosine kinase inhibitor of vascular endothelial growth factor receptors 1, 2, and 3, has demonstrated antitumor activity in a Phase II study in patients with renal cell carcinoma (RCC), and is currently being studied in clinical trials in patients with RCC and other solid tumors. The goal of this study was to investigate the effect of food on the pharmacokinetics (PK) of a single 1.5 mg dose of tivozanib in healthy subjects. Methods: This was a single-center, open-label, randomized, two-period, crossover Phase I trial. Subjects were admitted to the clinical research unit (CRU) 1 day before dosing, fasted ∼10 hours, and were randomized to fed (standard high-fat breakfast)/fasted or fasted/fed sequence. In each phase of the sequence, subjects received a single oral dose of tivozanib 1.5 mg with a 6-week washout period between doses. Subjects remained at the CRU for at least 48 hours post dose for blood sample collection and safety monitoring, and were assessed on an outpatient basis for up to 504 hours post dose. PK data were analyzed by non-compartmental methods. The effect of food on PK was assessed using standard criteria for bioequivalence based on the exposure parameters AUC0-α and Cmax. If the 90% confidence intervals for the fed/fasted AUC0-α and Cmax fell within the range of 80% to 125%, it was concluded that food had no effect on exposure. Results: Thirty healthy volunteers were enrolled (19M/11F; mean age 39 years [range 22-53 years]). There was no significant difference in AUC0-α of serum tivozanib between the fed and fasted states (107.4%; Table 1). Food caused a significant decrease in serum tivozanib levels vs fasted state (Cmax: 77.5%). Conclusions: These results indicate that dosing tivozanib with food decreases maximal concentrations by ∼ 23%, but does not affect overall exposure. As tivozanib is dosed chronically in oncology patients and accumulates ∼6 to 7 times single-dose levels when at steady-state, these results are not likely to affect dosing.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 752. doi:1538-7445.AM2012-752
Abstract 768: MCL1 and glycolysis synergistically induce prednisolone resistance, which can be therapeutically reversed
Authors: Ariës Ingrid M. , van den Dungen Rosanna , Pieters Rob , Boer Monique L. Den
Published: August 2012

Although cure rates for acute lymphoblastic leukemia (ALL) have greatly improved, contemporary treatment is still ineffective in 20% of patients. Unsuccessful treatment can be mainly ascribed to resistance towards antileukemic drugs. In particular prednisolone is highly associated with poor prognosis. Overcoming drug resistance is therefore necessary to increase current survival rates. Microarray analysis of precursor B-ALL patients revealed MCL1 as an important gene involved in prednisolone resistance (Holleman, N Engl J Med 2004). We demonstrated sensitization towards prednisolone after lentiviral silencing of MCL1 in prednisolone resistant leukemic cell lines (Stam, Blood 2009). These results point to MCL1 as an interesting candidate for targeted therapy. We also discovered that cells upregulate their glucose consumption level to protect from prednisolone induced apoptosis. 2-Deoxyglucose (2DG), an inhibitor of glycolysis, sensitized for prednisolone in leukemic cell lines and patients’ cells (Hulleman, Blood 2009). We here provide evidence for an inter-connection between MCL1, glycolysis and prednisolone resistance. A clear increase in glucose consumption was observed after targeting MCL1 in leukemic cells by both lentiviral silencing and mRNA antagonists. This suggests that cells upregulate glycolysis to prevent apoptosis induced upon MCL1 knockdown. Indeed, co-exposure to 2DG substantially decreased the proliferation rate of MCL1-silenced cells, and most importantly, synergized with silencing of MCL1 in reversing resistance to prednisolone. Altogether these data provide evidence for concomitant causes of resistance and indicate that MCL1 silencing and glycolysis inhibition can serve as potent ways to reverse resistance to prednisolone in acute leukemia.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 768. doi:1538-7445.AM2012-768
Abstract 777: GW583340 and GW2974, human EGFR and HER-2 inhibitors, reverse ABCG2- and ABCB1-mediated drug resistance
Authors: Sodani Kamlesh S. , Tiwari Amit K. , Singh Satyakam , Patel Atish , Xiao Zhijie , Chen Junjiang , Sun Yueli , Talele Tanaji T. , Chen Zhe-Sheng
Published: August 2012

The overexpression of ATP-binding cassette (ABC) transporters often lead to the development of multidrug resistance (MDR), resulting in chemotherapy failure. Previously, we reported that lapatinib (GW572016), a human epidermal growth factor receptor (EGFR) and HER-2 tyrosine kinase inhibitor (TKI), modulates ABC subfamily B member 1 (ABCB1)- and ABC subfamily G member 2 (ABCG2)-mediated MDR in cancer cells. In the present study, we conducted experiments to determine if GW583340 and GW2974; structural analogues of lapatinib; could reverse ABCB1- and ABCG2-mediated MDR. Our results showed that both GW583340 and GW2974 significantly sensitized MDR cells overexpressing ABCB1 to their anticancer substrates as well as ABCG2 overexpressing cells to its substrates. Subsequently, both GW583340 and GW2974 significantly increased intracellular accumulation of [3H]-paclitaxel in ABCB1 overexpressing cells. Similarly, both GW583340 and GW2974 significantly increased accumulation of [3H]-mitoxantrone in ABCG2 overexpressing cells. In addition, GW583340 and GW2974 significantly inhibited ABCG2-mediated transport of methotrexate in ABCG2 overexpressing membrane vesicles. Moreover, there was no significant change in the expression levels of ABCB1 and ABCG2 in the cell lines exposed to 5 μM of either GW583340 or GW2974 for 3 days. In addition, docking model predicted the binding conformation of GW583340 and GW2974 within the large cavity of the transmembrane region of homology modeled human ABCB1. We conclude that GW583340 and GW2974, at clinically achievable concentrations reverse ABCB1- and ABCG2-mediated MDR by directly blocking the drug efflux function of these transporters. These findings may be useful in developing cancer combinational therapy with EGFR TKIs in the future.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 777. doi:1538-7445.AM2012-777
Abstract 78: BRCA1/2 mutation analysis in 43 ovarian cell lines reveals only one functionally deleterious BRCA1 mutation
Authors: Stordal Britta , Timms Kirsten , Renaud Mickaël , Thery Julien , Farelly Angela , Williams Deborah , Potter Jennifer , Tran Thanh , Korpanty Greg , Cremona Mattia , Carey Mark , Li Jie , Li Yang , Aslan Ozlem , O'Leary John J. , Mills Gordon B. , Hennessy Bryan T.
Published: August 2012

Introduction: Germline mutations in DNA repair genes BRCA1/2 increase the risk of developing breast and ovarian cancer. Germline BRCA1/2 mutations occur in 8.6-13.7% of unselected epithelial ovarian cancers, somatic mutations are also frequent. BRCA1/2 mutated or dysfunctional cells may be sensitive to PARP inhibition by synthetic lethality, the inhibition of two major DNA repair pathways. Therefore, there is a need to characterize ovarian cell lines for their BRCA1/2 status for in vitro studies of novel drugs. Methods: The BRCA1/2 genes were sequenced in 43 ovarian cell lines, mRNA expression of BRCA1/2 and gene methylation of BRCA1 were also examined. The cytotoxicity of PARP inhibitors olaparib and veliparib was examined in 10 cell lines. Results: Sequencing of BRCA1/2 revealed 18 and 33 different alterations respectively, mostly non-pathogenic polymorphisms. The cell line SNU-251 has a deleterious BRCA1 mutation at 5564G>A and is the only deleterious BRCA1/2 mutant in the panel. Two cell lines (UPN-251 and PEO1) had deleterious mutations as well as additional reversion mutations which restore the function of the protein. BRCA1 was methylated in three cell lines (A1847, OVCAR8, EF021) and there was a corresponding decrease in gene expression in these cell lines. The deleterious mutant SNU-251 was relatively resistant to PARP inhibition, compared to wild-type cells. However, the methylated cell lines A1847 and OVCAR8 were more sensitive to PARP inhibition than wild-type cells. Conclusions: The incidence of BRCA1/2 deleterious mutations 1/43 cell lines (2.3%) is much lower than the population incidence. The reversion mutations suggest that there is a selective pressure against BRCA1/2 mutations in cell culture. PARP inhibitors may be useful in patients with BRCA1 methylation and reduced gene expression in their ovarian tumour.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 78. doi:1538-7445.AM2012-78
Abstract 811: VAL083, a novel N7 alkylating agent, surpasses temozolomide activity and inhibits cancer stem cells providing a new potential treatment option for glioblastoma multiforme
Authors: Hu Kaiji , Fotovati Abbas , Chen James , Triscott Joanna , Bacha Jeffery , Brown Dennis , Dunn Sandra E.
Published: August 2012

Glioblastoma (GBM) remains one of the most difficult tumors to treat in part because many new agents fail to cross the blood brain barrier (BBB) and secondly due to intrinsic drug resistance. Temozolomide (TMZ) is a front-line therapy for the treatment of GBM, however, it is often ineffective due to drug inactivation by O6-methylguanine-DNA methyltransferase (MGMT). Cancer stem cells (CSC) are a subpopulation of the tumor that resist therapy and give rise to relapse. Here we described VAL083 a novel alkylating agent that creates N7 methylation on DNA, which was initially intriguing because it crosses the BBB. We addressed how it compared to TMZ, whether it could be used to overcome MGMT-driven drug resistance and if it has activity against CSCs. Addressing these questions provides further preclinical support for VAL083, which is currently undergoing human clinical trials in the USA against refractory GBM. VAL083 inhibited U251 and SF188 cell growth in monolayer and as neurospheres better then TMZ and caused apoptosis after 72 hrs. In a 10-day colony formation assay, VAL083 (5uM) suppressed SF188 growth by ∼95%. T98G cells are classically TMZ resistant and express MGMT yet VAL083 inhibited their growth in monolayer after 72 hrs in a dose-dependent manner (IC50<5 uM). VAL083 also inhibited the growth of CSCs by 100% in neurosphere growth assays. In summary, VAL083 has better in vitro efficacy than TMZ against brain tumor cells, can overcome resistance associated with MGMT, and targets brain tumor CSCs demonstrating that it has the potential to surpass the standard-of-care.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 811. doi:1538-7445.AM2012-811
Abstract 813: Mechanism of cellular uptake of Pt drugs: The role of hCTR1
Authors: Ivy Kristin D. , Kaplan Jack H.
Published: August 2012

Cisplatin is an anti-cancer drug used to treat various human malignancies including breast, testicular, ovarian, prostrate, lung, head, neck, and colon cancers. While this drug has been shown to successfully treat tumors, its use is limited by the rapid development of resistance. This drug has been used since the 70's to treat different forms of cancer and recent studies have attempted to understand the mechanism of resistance by examining how this anti-cancer drug is taken into the cell, distributed within the cell, and exported. It has been claimed that hCTR1, the human high affinity Cu transporter, is the major entry pathway for cis-Pt and related anti-cancer drugs in human cells. Furthermore, that the entry mechanism via hCTR1 mimics that of Cu. Since hCTR1 is a very selective transporter for Cu(I), its function as a cisplatin entry pathway is unexpected. We will describe the results of our recent studies on HEK293 cells that over-express wild-type hCTR1 and a number of mutant Cu transporters, ovarian tumor cells that are sensitive or resistant to cis-Pt, and a number of human breast cell tumor lines. We have compared the relative entry rates of Cu and Pt in these cells and, via cell surface biotinylation and imaging, the effects of extracellular Cu, which causes regulatory endocytosis of the transporter, to that of the cis-Pt drugs. Our results suggest that hCTR1 is not the major entry route of Pt drugs into human cells and that the response of the transporter, in a number of these cells, to the presence of extracellular drug is different from its response to Cu. Supported by NIH Grant P01 GM 067166.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 813. doi:1538-7445.AM2012-813
Abstract 818: Targeting the redox-adaptive response in tumors to potentiate cancer therapy
Authors: Hanigan Marie H. , Wickham Stephanie , Reagan Nicholas , West Matthew B. , Thai Justin , Cook Paul F. , Li Pui Kai
Published: August 2012

Resistance to chemotherapy is a major impediment to successful cancer treatment. To enhance therapeutic intervention, we are developing novel compounds that target the redox-adaptive response in tumors, a process that increases intracellular glutathione levels. Elevated levels of intracellular glutathione render tumors resistant to chemotherapy and radiation therapy by protecting against reactive oxygen species, inactivating electrophilic metabolites of chemotherapy drugs, and blocking Bcl-2-mediated apoptosis. Glutathione synthesis requires free intracellular cysteine. Cells under oxidative stress synthesize high levels of glutathione and the supply of cysteine becomes rate limiting for glutathione synthesis. We have developed a novel strategy to deplete the free cysteine available to tumor cells, thereby blocking the drug resistance in tumor cells that have undergone redox-adaption. To accomplish this in humans, two enzymes must be inhibited, gamma-glutamyl transpeptidase (aka gamma-glutamyl transferase, GGT1) and gamma-glutamyl leukotrienase (aka, GGT-rel, GGT5). In humans, both GGT1 and GGT5 cleave the gamma-glutamyl bond of extracellular glutathione initiating the breakdown of glutathione into its constituent amino acids, glutamate, cysteine and glycine. Inhibiting GGT1 and GGT5 depletes the body of cysteine by blocking the catabolism of glutathione in the kidney resulting in the excretion of glutathione in the urine. Inhibiting these enzymes also blocks the tumor's ability to use extracellular glutathione as a source of cysteine.Our approach not only reduces the ability of the tumor to synthesize glutathione, it also reduces the cysteine (and the total amount of reduced thiol) in the tumor. Previously we used high-throughput screening and identified OU749 (N-[5-(4-methoxybenzyl)-1,3,4-thiadiazol-2-yl]benzenesulfonamide), a novel inhibitor of GGT1. We have subsequently developed a new assay system, which provides an unambiguous method to evaluate the inhibition of the physiologic reactions catalyzed by both GGT1 and GGT5. Kinetic data obtained from the assay can be used to directly evaluate the potency and mechanism of action of each inhibitor. Synthesis of an extensive array of OU749 analogues and evaluation of the inhibitory activity of these analogues in our new assay system has resulted in the identification of two new lead compounds, HL-6 and HL-7. These new leads are uncompetitive inhibitors of both human GGT1 and GGT5. These compounds are orders of magnitude less toxic than the currently available GGT1 inhibitors. Continued optimization of these lead compounds will provide new drugs for potentiating cancer therapy.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 818. doi:1538-7445.AM2012-818
Abstract 835: Epigenetic biomarkers of chemoresistance in colorectal cancer
Authors: Moutinho Cátia , Martinez-Cardús Anna , Martínez-Balibrea Eva , Abad Albert , Esteller Manel
Published: August 2012

Resistance acquisition to chemotherapeutic agents is one of the main problems that come up during cancer treatment. The mechanisms of developing resistance are poorly understood, and the causes underlying the loss of sensitivity to common chemotherapy drugs are largely unknown. Promoter hypermethylation of DNA repair genes has been demonstrated to be an excellent biomarker of sensitivity to chemotherapy agents, being the epigenetic silencing of MGMT in gliomas the most remarkable example. In this study, we have focused our research on one of the most common cancer type, the colorectal cancer (CRC). Thus, we propose to assess how DNA methylation alterations can help to explain acquired resistance to oxaliplatin (OXA) in this type of tumours. To address this issue we used LoVo OXA sensitive and resistant cell lines (LoVoS_OXA vs. LoVoR_OXA). Using DNA genomic bisulfite sequencing we found that CpG islands located at some genes promoters (candidate genes) are unmethylated in LoVoS_OXA but hypermethylated in LoVoR_OXA, being these phenomena correlated with lower mRNA and protein expression in the resistant cell line. DNA promoter methylation of theses candidate genes was then checked by Methylation Specific PCR (MSP) in 25 human CRC tumour samples obtained from patients treated with FOLFOX (5-fluorouracil; leucovorin and oxaliplatin)(blind study). Remarkably, one candidate gene promoter hypermethylation was related with worse response to chemotherapy, poor overall survival and poor progression- free survival. In contrast, promoter hypermethylation of a second candidate gene was related with improved treatment response and survival rates. Being this an ongoing project, we are performing in vitro studies in order to check if changes in promoter hypermethylation can modulate cell sensitivity to OXA drug. The results obtained in this study can help to understand the biological processes leading to chemoresistance and also to the future management of cancer patients.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 835. doi:1538-7445.AM2012-835
Abstract 847: Foretinib, a multi-kinase inhibitor of cMET and PDGFRα, in the treatment of disseminated medulloblastoma
Authors: Faria Claudia C. , Golbourn Brian J. , Smith Christian A. , Rutka James T.
Published: August 2012

Medulloblastoma (MB) is the most common malignant brain tumor in childhood accounting for around 10% of all pediatric cancer deaths. Dissemination occurs in 30% of the patients at the time of diagnosis and essentially defines children with incurable disease given that metastatic disease is refractory to current treatments. Therefore, there is an urgent need for new therapies to treat metastatic MB. The hepatocyte growth factor (HGF)/cMET signaling pathway has been recently implicated in the pathogenesis of MB. Overexpression of cMET and HGF is associated with group C tumors which have high incidence of metastasis and poor outcome. Moreover, overexpression of the platelet-derived growth factor receptor (PDGFR) was also identified in metastatic MB. Using foretinib, a multi-kinase inhibitor of cMET and PDGFRα, we aimed to target two important pathways involved in MB dissemination. Using three MB cell lines (Daoy, ONS - 76 and D425) that express different amounts of the receptors (cMET and PDGF), we performed dose-response experiments with foretinib, measuring downstream targets of cMET and PDGFRα activation. We showed that foretinib inhibits proliferation, migration, invasion and anchorage independent growth of MB cell lines in a dose-dependent manner. By immunofluorecence, we observed that foretinib induces polyploidy and using flow cytometry analysis we found that it also induces apoptosis and cell cycle arrest in G2-M phase. To test the efficacy of foretinib in vivo we have created a disseminated mouse model of MB injecting MB cells in the fourth ventricle of nude mice, mimicking the process of dissemination in children. The cells were previously transfected with a luciferase expressing vector allowing weekly monitoring of tumor growth and dissemination by bioluminescence imaging. Foretinib was able to reduce tumor growth and metastasis in xenografts, increasing survival when compared to controls. Altogether these results suggest that small molecule inhibitors of cMET and PDGFR may represent a new therapeutic strategy in the treatment of disseminated MB. Given that foretinib is already being tested in clinical trials for other forms of cancer, the findings from our experiments may lead to the design of identical trials in children with metastatic medulloblastoma.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 847. doi:1538-7445.AM2012-847
Abstract 85: RUNX1 and ELK1 directly regulate the transcription of EVI1 during megakaryocytic differentiation
Authors: Maicas Miren , Vazquez Iria , Vicente Carmen , Cortes-Lavaud Xabier , Marcotegui Nerea , Urquiza Leyre , Calasanz Maria J. , Odero Maria D.
Published: August 2012

High expression of EVI1 is a negative prognostic indicator of survival in patients with acute myeloid leukemia (AML). The EVI1 gene (3q26) codes for a transcription factor indispensable for the proliferation and self-renewal of HSCs during embryogenesis and in the adult, and is downregulated upon differentiation; however, the regulation of EVI1 in megakaryocytic differentiation should be different, as it is highly expressed in megakaryocytes and platelets, but not in other committed hematopoietic cells. Our aim was to identify transcription factors involved in the regulation of EVI1. Site-directed mutagenesis and ChIP assays identified RUNX1 and ELK1 as putative transcription factors of EVI1. Furthermore, knockdown of RUNX1 and ELK1 led to EVI1 downregulation, and their overexpression to upregulation of EVI1. Of note, in a series of patient samples with AML at diagnosis (n=46), we found a significant positive correlation between EVI1 and RUNX1, and EVI1 and ELK1 mRNA expression levels. RUNX1 has been reported as a key transcription factor in megakaryocytic development. To investigate whether RUNX1 could regulate EVI1 in this process, we treated K562 cells with TPA. After the treatment, we observed an increase in RUNX1, as previously described, and in ELK1 and EVI1. Knockdown of RUNX1 resulted in a marked decrease of the megakaryocytic markers ITGA2B and ITGB3, and of EVI1. Moreover, EVI1 knockdown led to the same effects by blocking the differentiation process, suggesting that the role of EVI1 in megakaryocytic differentiation would be through RUNX1. Interestingly, chromatin immunoprecipitation assay (ChIP) showed that RUNX1 regulates the transcription of EVI1 by acetylation of the histone H3 on its promoter region. In conclusion, we demonstrate that RUNX1 and ELK1, two proteins with essential functions in hematopoiesis, regulate EVI1 in AML. Moreover, our results suggest that these transcription factors could regulate EVI1 expression during megakaryocytic differentiation, and that RUNX1 would activate EVI1 expression through acetylation of the histone H3 on its promoter region. This study opens new directions to further understand the mechanisms of EVI1 overexpressing leukemias.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 85. doi:1538-7445.AM2012-85
Abstract 857: Development of a Bruton's tyrosine kinase (Btk) inhibitor - ONO-WG-307, a potential treatment for B-cell malignancies
Authors: Kozaki Ryohei , Yoshizawa Toshio , Yasuhiro Tomoko , Mirjolet Jean-François , Birkett Joseph , Narita Masami , Kawabata Kazuhito
Published: August 2012

Purpose: Signals from B cell receptors (BCR) play a central role in signal transduction pathways regulating survival, activation, proliferation, and differentiation of B-lineage lymphoid cells. BCR signaling is implicated in the survival of malignant B cells and recent studies indicate that targeting Btk, an essential component of the BCR pathway, may be effective in the treatment of B-cell lymphoma. ONO-WG-307 is a highly potent and selective Btk inhibitor with an IC50 in the sub-nmol/L range. We evaluated the inhibitory effect of ONO-WG-307 alone and in combination with the chimeric type I anti-CD20 antibody rituximab, using in vitro tumor growth assays and mouse xenograft models. Methods: Two types of tumor cell lines (follicular lymphoma (FL) and activated B-cell-like (ABC) sub-type of diffuse large B cell lymphoma (DLBCL)) wereplated on to micro-titration plates (at doses up to 200 umol/L for ONO-WG-307 and 100 ug/mL for rituximab) for in vitro cytotoxic assays. The same cells were also implanted subcutaneously (SC) into female SCID mice to explore ONO-WG-307 anti-tumor activity in vivo (orally at doses up to 50 mg/kg, bid for ONO-WG-307 and intraperitoneally at doses up to 10 mg/kg, q7d). The IC50 of the in vitro cytotoxic activity was determined using a MTS assay 72 and 96 hours after incubation. Anti-tumor activity was defined as the ratio of the median tumor volume of treatment groups versus control group. The determination of combination index (CI) was calculated by the median-effect method. The CI was used to express synergism (<1), additivity (=1) or antagonism (>1). Results: In the MTS assay, ABC-DLBCL cells were much more sensitive to ONO-WG-307 given as single agent compared with FL cell lines, however, the activity of rituximab single agent had no impact on ABC-DLBCL cell lines in contrast to its inhibitory activity against FL cell lines. When ONO-WG-307 was combined with rituximab, a moderate antagonism was observed in vitro on FL cell lines. In an ABC-DLBCL xenograft model, treatment with single agent ONO-WG-307 resulted in a dose-dependent inhibition of tumor growth and also showed moderate anti-tumor activity in a FL xenograft model. Conclusion: ONO-WG-307 is a highly potent and selective oral Btk inhibitor with evidence of efficacy in both the ABC-DLBCL and the FL cell lines and in xenograft models. These preliminary results suggest that ONO-WG-307 may be an effective therapy in the treatment of B-cell malignancies. To determine whether combination of ONO-WG-307 and rituximab is more efficacious, additional in vivo combination work is underway.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 857. doi:1538-7445.AM2012-857