bluuebunny/crossref_metadata_2025 huggingface dataset:
Metabolic Signatures Associated with a NAD Synthesis Inhibitor–Induced Tumor Apoptosis Identified by 1H-Decoupled-31P Magnetic Resonance Spectroscopy
Authors: Muruganandham Manickam , Alfieri Alan A. , Matei Cornelia , Chen Yuchun , Sukenick George , Schemainda Isabel , Hasmann Max , Saltz Leonard B. , Koutcher Jason A.
Published: September 2005

Purpose: Attempts to selectively initiate tumor cell death through inducible apoptotic pathways are increasingly being exploited as a potential anticancer strategy. Inhibition of NAD+ synthesis by a novel agent FK866 has been recently reported to induce apoptosis in human leukemia, hepatocarcinoma cells in vitro, and various types of tumor xenografts in vivo. In the present study, we used 1H-decoupled phosphorus (31P) magnetic resonance spectroscopy (MRS) to examine the metabolic changes associated with FK866 induced tumor cell death in a mouse mammary carcinoma.

Experimental Design: Induction of apoptosis in FK866-treated tumors was confirmed by histology and cytofluorometric analysis. FK866-induced changes in mammary carcinoma tumor metabolism in vivo were investigated using 1H-decoupled 31P MRS. To discern further the changes in metabolic profiles of tumors observed in vivo, high-resolution in vitro 1H-decoupled 31P MRS studies were carried out with perchloric acid extracts of mammary carcinoma tumors excised after similar treatments. In addition, the effects of FK866 on mammary carcinoma tumor growth and radiation sensitivity were studied.

Results: Treatment with FK866 induced a tumor growth delay and enhanced radiation sensitivity in mammary carcinoma tumors that was associated with significant increases in the 31P MR signal in the phosphomonoester region and a decrease in NAD+ levels, pH, and bioenergetic status. The 31P MRS of perchloric acid extracts of treated tumors identified the large unresolved signal in the phosphomonoester region as the resultant of resonances originating from intermediates of tumor glycolysis and guanylate synthesis in addition to alterations in pyridine nucleotide pools and phospholipid metabolism.

Conclusion: The present results suggest that FK866 interferes with multiple biochemical pathways that contribute to the increased cell death (apoptosis) and subsequent radiation sensitivity observed in the mammary carcinoma that could be serially monitored by 31P MRS.
Fractional Genomic Alteration Detected by Array-Based Comparative Genomic Hybridization Independently Predicts Survival after Hepatic Resection for Metastatic Colorectal Cancer
Authors: Mehta Kshama R. , Nakao Kentaro , Zuraek Marlene B. , Ruan Daniel T. , Bergsland Emily K. , Venook Alan P. , Moore Dan H. , Tokuyasu Taku A. , Jain Ajay N. , Warren Robert S. , Terdiman Jonathan P. , Waldman Frederic M.
Published: September 2005

Purpose: Although liver resection is the primary curative therapy for patients with colorectal hepatic metastases, most patients have a recurrence. Identification of molecular markers that predict patients at highest risk for recurrence may help to target further therapy.

Experimental Design: Array-based comparative genomic hybridization was used to investigate the association of DNA copy number alterations with outcome in patients with colorectal liver metastasis resected with curative intent. DNA from 50 liver metastases was labeled and hybridized onto an array consisting of 2,463 bacterial artificial chromosome clones covering the entire genome. The total fraction of genome altered (FGA) in the metastases and the patient's clinical risk score (CRS) were calculated to identify independent prognostic factors for survival.

Results: An average of 30 ± 14% of the genome was altered in the liver metastases (14% gained and 16% lost). As expected, a lower CRS was an independent predictor of overall survival (P = 0.03). In addition, a high FGA also was an independent predictor of survival (P = 0.01). The median survival time in patients with a low CRS (score 0-2) and a high (≥20%) FGA was 38 months compared with 18 months in patients with a low CRS and a low FGA. Supervised analyses, using Prediction Analysis of Microarrays and Significance Analysis of Microarrays, identified a set of clones, predominantly located on chromosomes 7 and 20, which best predicted survival.

Conclusions: Both FGA and CRS are independent predictors of survival in patients with resected hepatic colorectal cancer metastases. The greater the FGA, the more likely the patient is to survive.
Clinical Significance of Heparin-Binding Epidermal Growth Factor–Like Growth Factor and A Disintegrin and Metalloprotease 17 Expression in Human Ovarian Cancer
Authors: Tanaka Yoshihiro , Miyamoto Shingo , Suzuki Satoshi O. , Oki Eiji , Yagi Hiroshi , Sonoda Kenzo , Yamazaki Ayano , Mizushima Hiroto , Maehara Yoshihiko , Mekada Eisuke , Nakano Hitoo
Published: September 2005

Purpose: Lysophosphatidic acid, which is enriched in the peritoneal fluid of ovarian cancer patients, plays a key role in the progression of ovarian cancer. Lysophosphatidic acid can generate epidermal growth factor receptor (EGFR) signal transactivation involving processing of EGFR ligands by ADAM (a disintegrin and metalloprotease) family metalloproteases. We aimed to investigate the clinical significance of EGFR ligands and ADAM family in the lysophosphatidic acid–induced pathogenesis of ovarian cancer.

Experimental Design: We examined the expression of EGFR ligands and ADAM family members in 108 patients with normal ovaries or ovarian cancer, using real-time PCR, immunohistochemistry, and in situ hybridization, and analyzed the clinical roles of these molecules. Statistical analyses of these data were done using the Mann-Whitney test, Kaplan-Meier method, or Spearman's correlation analysis.

Results: Large differences in expression were found for heparin-binding EGF-like growth factor (HB-EGF) and other EGFR ligands and for ADAM 17 and other ADAM family members. HB-EGF expression was significantly increased in advanced ovarian cancer compared with that in normal ovaries (P < 0.01). HB-EGF expression was significantly associated with the clinical outcome (P < 0.01). ADAM 17 expression was significantly enhanced in both early and advanced ovarian cancer compared with that in normal ovaries (both P < 0.01), although it had no clinical significance in the progression-free survival. HB-EGF expression was significantly correlated with ADAM 17 expression (γ = 0.437, P < 0.01).

Conclusions: Our findings suggest that HB-EGF and ADAM 17 contribute to the progression of ovarian cancer and that HB-EGF plays a pivotal role in the aggressive behavior of a tumor in ovarian cancer.
Farnesyltransferase Inhibitors and Human Malignant Pleural Mesothelioma: A First-Step Comparative Translational Study
Authors: Cesario Alfredo , Catassi Alessia , Festi Luigi , Imperatori Andrea , Pericelli Andrea , Galetta Domenico , Margaritora Stefano , Porziella Venanzio , Cardaci Vittorio , Granone Pierluigi , Dominioni Lorenzo , Russo Patrizia
Published: September 2005

It is known that the potential clinical use of farnesyltransferase inhibitors (FTI) could be expanded to include cancers harboring activated receptor tyrosine kinases. Approximately 70% of malignant pleural mesotheliomas (MPM) overexpress epidermal growth factor receptors (EGFR) and a subset express both EGFR and transforming growth factor α (TGF-α), suggesting an autocrine role for EGFR in MPM. We checked on MPM cells (10 human cell lines, 11 primary cultures obtained by human biopsies, and 7 short-term normal mesothelial cell cultures) concerning the following: (a) the relative overexpression of EGFR (Western blotting, flow cytometry, immunohistochemistry), (b) the relative expression of EGFR ligands (EGF, amphiregulin, TGF-α, ELISA), (c) the relative increase of the activated form of Ras (Ras-bound GTP) after EGF stimulation (Ras activation assay), (d) the efficacy of five different FTIs (HDJ2 prenylation, cell cytotoxicity, and apoptosis using ApopTag and gel ladder). EGFR was overexpressed in MPM cells compared with normal pleural mesothelial cells in equivalent levels as in non–small cell lung cancer cells A459. MPM cells constitutively expressed EGFR ligands; however, Ras activation was attenuated at high EGF concentrations (100 ng/mL). Growth of MPM cells was substantially not affected by treatment with different FTIs (SCH66336, BMS-214662, R115777, RPR-115135, and Manumycin). Among these, BMS-214662 was the only one moderately active. BMS-214662 triggered apoptosis in a small fraction of cells (not higher than 30%) that was paralleled by a slight decrease in the levels of TGF-α secreted by treated MPM cells. Our data highlighted the concept that the same signaling pathway can be regulated in different ways and these regulations can differ between different cells of different origin.
Phase I Clinical and Pharmacokinetic Study of Kahalalide F in Patients with Advanced Androgen Refractory Prostate Cancer
Authors: Rademaker-Lakhai Jeany M. , Horenblas Simon , Meinhardt Willem , Stokvis Ellen , de Reijke Theo M. , Jimeno José M. , Lopez-Lazaro Luis , Lopez Martin José A. , Beijnen Jos H. , Schellens Jan H.M.
Published: September 2005

Purpose: The purpose is to determine the maximum tolerated dose, profile of adverse events, and dose-limiting toxicity of Kahalalide F (KF) in patients with androgen refractory prostate cancer. Furthermore, the pharmacokinetics after KF administration and preliminary antitumor activity were evaluated. KF is a dehydroaminobutyric acid–containing peptide isolated from the marine herbivorous mollusk, Elysia rufescens.

Experimental Design: Adult patients with advanced or metastatic androgen refractory prostate cancer received KF as an i.v. infusion over 1 hour, during five consecutive days every 3 weeks. The starting dose was 20 μg per m2 per day. Clinical pharmacokinetics studies were done in all patients using noncompartmental analysis. Prostate-specific antigen levels were evaluated as a surrogate marker for activity against prostate cancer.

Results: Thirty-two patients were treated at nine dose levels (20-930 μg per m2 per day). The maximum tolerated dose on this schedule was 930 μg per m2 per day. The dose-limiting toxicity was reversible and asymptomatic Common Toxicity Criteria grade 3 and 4 increases in transaminases. The recommended dose for phase II studies is 560 μg per m2 per day. Pharmacokinetics analysis revealed dose linearity up to the recommended dose. Thereafter, a more than proportional increase was observed. Elimination was rapid with a mean (SD) terminal half-life (t1/2) of 0.47 hour (0.11 hour). One patient at dose level 80 μg per m2 per day had a partial response with a prostate-specific antigen decline by at least 50% for ≥4 weeks. Five patients showed stable disease.

Conclusions: KF can be given safely as a 1-hour i.v. infusion during five consecutive days at a dose of 560 μg per m2 per day once every 3 weeks.
Caspase-Dependent Apoptosis Induction by Phenethyl Isothiocyanate, a Cruciferous Vegetable-Derived Cancer Chemopreventive Agent, Is Mediated by Bak and Bax
Authors: Xiao Dong , Zeng Yan , Choi Sunga , Lew Karen L. , Nelson Joel B. , Singh Shivendra V.
Published: September 2005

Purpose: The present study was undertaken to gain insights into the molecular mechanism of apoptosis induction by phenethyl isothiocyanate (PEITC) using prostate cancer cell lines derived from transgenic adenocarcinoma mouse prostate (TRAMP) mice (TRAMP-C1 and TRAMP-C2).

Experimental Design and Results: The viability of TRAMP-C1 and TRAMP-C2 cells was reduced significantly in the presence of PEITC in a concentration-dependent manner as determined by sulforhodamine B and trypan blue dye exclusion assays. Treatment of TRAMP-derived cells with PEITC revealed features characteristic of apoptosis induction, including appearance of subdiploid cells (determined by flow cytometry), cytoplasmic histone-associated DNA fragmentation (determined by an ELISA assay), and cleavage of caspase-3 (determined by immunoblotting). The PEITC-induced apoptosis in TRAMP-derived cells was associated with a marked increase in the level of proapoptotic protein Bak and/or a decrease in the levels of antiapoptotic protein Mcl-1 or Bcl-xL and disruption of mitochondrial membrane potential. The SV40 immortalized mouse embryonic fibroblasts derived from Bak and Bax double knockout mice were significantly more resistant to PEITC-induced DNA fragmentation compared with wild-type or Bak−/− mouse embryonic fibroblasts. The PEITC-induced apoptosis in both cell lines was significantly attenuated in the presence of caspase inhibitors zVAD-fmk, zLEHD-fmk, and zIETD-fmk. Oral administration of PEITC (9 or 12 μmol PEITC/d, Monday-Friday) significantly retarded growth of TRAMP-C1 xenografts in nude mice without causing weight loss or any other side effects.

Conclusion: The results of the present study indicate that caspase-dependent apoptosis by PEITC is mediated by Bak and Bax proteins.
Risk Assessment in Patients with Acute Myeloid Leukemia and a Normal Karyotype
Authors: Bienz Marianne , Ludwig Madleina , Mueller Beatrice U. , Oppliger Leibundgut Elisabeth , Ratschiller Daniel , Solenthaler Max , Fey Martin F. , Pabst Thomas
Published: September 2005

Purpose: The recognition of a number of leukemia-specific cytogenetic abnormalities and their role as independent prognostic factors have provided considerable insights into leukemia pathogenesis and have paved the way to adopt risk-adapted treatment. However, ∼50% of newly diagnosed acute myeloid leukemia (AML) have a normal karyotype. There has therefore been much interest in identifying molecular markers that could help to improve the prognostic stratification of patients with normal-karyotype AML.

Experimental Design: Consecutive untreated AML patients (n = 67) from a single institution all with normal karyotype were analyzed for the presence of mutations in the myeloid transcription factor gene CEBPA (for CCAAT/enhancer binding protein-α), for internal tandem duplications (ITD) of the tyrosine kinase receptor gene FLT3 (for fms-like tyrosine kinase 3), and for expression of the BAALC gene (for brain and acute leukemia, cytoplasmic).

Results: 17.9% of normal-karyotype AML had mutations in the CEBPA gene, and 28.4% had FLT3-ITD; 65.7% (44 of 67) had high BAALC expression and 34.3% (23 of 67) had low BAALC expression. Patients with CEBPA mutations had a very favorable course of their disease. Median disease-free survival (DFS) and overall survival (OS) were 33.5 and 45.5 months, respectively, compared with 10 (e.g., 12 months in patients without CEBPA mutations; P = 0.0017; P = 0.0007). AML patients with FLT3-ITD had significantly shorter median DFS (P = 0.0328) and OS (P = 0.0148) than patients without FLT3-ITD. High BAALC expression predicted for a shorter DFS (P = 0.0152) and OS (P = 0.0210) compared with AML with low BAALC expression; 53.7% of normal-karyotype AML had neither FLT3-ITD nor CEBPA mutations. We found that high BAALC expression in normal-karyotype AML with neither FLT3-ITD nor CEBPA mutations (18 of 67) indicates adverse prognosis for both DFS and OS (P = 0.0001; e.g., P = 0.0001) compared with the group with low BAALC expression and absent FLT3-ITD and CEBPA mutations (18 of 67). Thus, BAALC expression represents a novel prognostic marker particularly for normal-karyotype AML patients with neither FLT3-ITD nor CEBPA mutations.

Conclusions: Assessment of CEBPA mutations, FLT3-ITD, and BAALC expression permits to split normal-karyotype AML into clinically distinct subgroups.
Myc Down-Regulation Sensitizes Melanoma Cells to Radiotherapy by Inhibiting MLH1 and MSH2 Mismatch Repair Proteins
Authors: Bucci Barbara , D'Agnano Igea , Amendola Donatella , Citti Arianna , Raza Giorgio H. , Miceli Roberto , De Paula Ugo , Marchese Rodolfo , Albini Sonia , Felsani Armando , Brunetti Ercole , Vecchione Aldo
Published: September 2005

Purpose: Melanoma patients have a very poor prognosis with a response rate of <1% due to advanced diagnosis. This type of tumor is particularly resistant to conventional chemotherapy and radiotherapy, and the surgery remains the principal treatment for patients with localized melanoma. For this reason, there is particular interest in the melanoma biological therapy.

Experimental Design: Using two p53 mutant melanoma models stably expressing an inducible c-myc antisense RNA, we have investigated whether Myc protein down-regulation could render melanoma cells more susceptible to radiotherapy, reestablishing apoptotic p53-independent pathway. In addition to address the role of p53 in the activation of apoptosis, we studied the effect of Myc down-regulation on radiotherapy sensitivity also in a p53 wild-type melanoma cell line.

Results: Myc down-regulation is able per se to induce apoptosis in a fraction of the cell population (∼40% at 72 hours) and in combination with γ radiation efficiently enhances the death process. In fact, ∼80% of apoptotic cells are evident in Myc down-regulated cells exposed to γ radiation for 72 hours compared with ∼13% observed after only γ radiation treatment. Consistent with the enhanced apoptosis is the inhibition of the MLH1 and MSH2 mismatch repair proteins, which, preventing the correction of ionizing radiation mismatches occurring during DNA replication, renders the cells more prone to radiation-induced apoptosis.

Conclusions: Data herein reported show that Myc down-regulation lowers the apoptotic threshold in melanoma cells by inhibiting MLH1 and MSH2 proteins, thus increasing cell sensitivity to γ radiation in a p53-independent fashion. Our results indicate the basis for developing new antitumoral therapeutic strategy, improving the management of melanoma patients.
Combined GADD45A and Thymidine Phosphorylase Expression Levels Predict Response and Survival of Neoadjuvant-Treated Gastric Cancer Patients
Authors: Napieralski Rudolf , Ott Katja , Kremer Markus , Specht Katja , Vogelsang Holger , Becker Karen , Müller Martina , Lordick Florian , Fink Ulrich , Rüdiger Siewert Jörg , Höfler Heinz , Keller Gisela
Published: September 2005

Purpose: We evaluated the expression of seven therapy-related genes to predict the clinical outcome of advanced gastric cancer patients treated with a neoadjuvant chemotherapeutic protocol.

Experimental Design: Pretherapeutic, formalin-fixed, and paraffin-embedded biopsies of 61 patients, who received a 5-fluorouracil (5-FU)– and cisplatin-based chemotherapy were studied. The expressions of the 5-FU–related genes TS, DPD, and TP and of the cisplatin-related genes ERCC1, ERCC4, KU80, and GADD45A were analyzed by quantitative real-time PCR. The expression levels of single genes and of various combinations were tested for an association with response and overall survival.

Results: High DPD levels were more frequently found in nonresponding patients and were associated with worse survival. GADD45A and TP levels showed weak associations with response, but GADD45A expression correlated with survival. There was no association with response for TS expression, but tumors with a high TS level were associated with worse survival. The combination of GADD45A and TP revealed the strongest predictive effect. High expression values of TP and/or GADD45A were exclusively found in nonresponding patients (P = 0.002) and were associated with a significantly poorer survival (P = 0.04).

Conclusions: Combined gene expression levels of TP and GADD45A represent a new variable to predict the clinical outcome after neoadjuvant chemotherapy in gastric cancer. The association of DPD expression with response and survival underlines a predominant role of DPD to predict 5-FU sensitivity. The association of TS expression levels with survival but not with response suggests an importance of this gene for tumor progression.
The Clinical Significance of Aurora-A/STK15/BTAK Expression in Human Esophageal Squamous Cell Carcinoma
Authors: Tanaka Eiji , Hashimoto Yosuke , Ito Tetsuo , Okumura Tomoyuki , Kan Takatsugu , Watanabe Go , Imamura Masayuki , Inazawa Johji , Shimada Yutaka
Published: September 2005

Purpose: Aurora-A/STK15/BTAK (Aurora-A) encodes a Serine/Threonine kinase associated with chromosomal distribution, and its up-regulation induces chromosomal instability thereby leading to aneuploidy and cell transformation in several types of cancer. In this study, we investigated the role of Aurora-A in human esophageal squamous cell carcinoma (ESCC).

Experimental Design: The expression levels of Aurora-A mRNA were compared in 33 ESCC tissues with that in corresponding normal esophageal epithelium by semiquantitative reverse transcription-PCR, and the distribution patterns and expression levels of Aurora-A protein were immunohistochemically investigated in the ESCC tumors of 142 patients. The results were then separately compared with the clinicopathologic findings of the patients, and the expression of Aurora-A was examined in nine ESCC cell lines and a normal esophageal epithelial cell line using Western blot analysis.

Results: The up-regulation of Aurora-A mRNA was found in 30% (10 of 33) of the tumors by semiquantitative reverse transcription-PCR, and protein up-regulation was found in 53% (75 of 142) of the patients by immunohistochemistry. mRNA and protein up-regulation of Aurora-A were correlated with distant lymph node metastasis (P = 0.05 and P = 0.04, respectively), and patients with Aurora-A mRNA or protein up-regulation had a poorer prognosis (P = 0.003 and P = 0.0009, respectively). Furthermore, multivariate analysis revealed that up-regulation of the Aurora-A protein was an independent prognostic factor. In addition, Aurora-A expression in all ESCC cell lines was higher than that in a normal esophageal epithelial cell line.

Conclusions: The up-regulation of Aurora-A expression may reflect the malignant behavior of ESCC and may prove useful information as a prognostic factor for ESCC patients.
In vivo Radioiodide Imaging and Treatment of Breast Cancer Xenografts after MUC1-Driven Expression of the Sodium Iodide Symporter
Authors: Dwyer Roisin M. , Bergert Elizabeth R. , O'Connor Michael K. , Gendler Sandra J. , Morris John C.
Published: September 2005

Purpose: Expression of the sodium iodide symporter (NIS) in the thyroid gland provides for effective imaging and treatment of thyroid cancer using radiolabeled iodide. Transfer of NIS into other tumors would expand the utility of this treatment to tumors of nonthyroid origin. MUC1 is a transmembrane glycoprotein that is overexpressed in many tumor types, including breast, pancreatic, and ovarian. The aim of this study was to create a construct containing NIS under the control of the MUC1 promoter to target expression specifically to MUC1-positive breast cancer cells.

Experimental Design: A replication-deficient adenoviral construct was created containing the MUC1 promoter followed by the human NIS gene. Iodide uptake assays, Western blot, and immunohistochemistry were used to confirm NIS expression and function. Breast cancer xenografts in mice were infected with Ad5/MUC1/NIS and then imaged and treated using radioiodide.

Results: A 58-fold increase in iodide uptake was observed in infected MUC1-positive T47D cells with no significant increase observed in MUC1-negative MDA-MB-231 cells or in cells infected with the control virus. The in vivo study yielded clear images of Ad/MUC1/NIS-infected tumor xenografts using 123I. Administration of a therapeutic dose of 131I resulted in an 83% reduction in tumor volume, whereas control tumors continued to increase in size (P < 0.01).

Conclusions: These results show that the MUC1 promoter is capable of directing efficient and selective expression of the NIS gene in MUC1-positive breast tumor cells. This could potentially have applications for both imaging and therapy in a range of MUC1-positive tumor types.
Phase I Study of the Farnesyltransferase Inhibitor BMS-214662 Given Weekly in Patients with Solid Tumors
Authors: Papadimitrakopoulou Vali , Agelaki Sofia , Tran Hai T. , Kies Merrill , Gagel Robert , Zinner Ralph , Kim Edward , Ayers Gregory , Wright John , Khuri Fadlo
Published: September 2005

Purpose: A phase I trial of BMS-214662, a selective farnesyltransferase inhibitor with significant preclinical antitumor activity in which drug was given as a weekly 1-hour infusion for four of six weeks, was conducted to evaluate the tolerability, pharmacokinetics, and pharmacodynamic effect on farnesyltransferase activity in peripheral blood mononuclear cells.

Experimental Design: BMS-214662 was given to 27 patients with solid tumors at 10 escalating dose levels (28-220 mg/m2) allowing intrapatient dose escalation; pharmacokinetics and pharmacodynamics were done at the first seven dose levels.

Results: Grade 4 neutropenia (four patients) was the most common dose-limiting toxicity followed by aminotransferase elevation (grade 3 alanine aminotransferase and grade 4 aspartate aminotransferase) and grade 3 dehydration. Most frequent toxicities were neutropenia in 11 (14%), anemia in 15 (19%), fatigue in 9 (12%), and nausea and diarrhea in 6 (8%) of courses, respectively. One minor response lasting 18 weeks in a patient with non–small cell lung cancer, serum calcitonin level reduction accompanied by disease stabilization in two of four patients with medullary thyroid carcinoma, and stable disease in 16 of 25 evaluable patients was seen. No correlation was observed between dose and Cmax, total body clearance (mean, 26.15 ± 10.88 L per hour per m2), volume of distribution at steady state (mean, 39.51 ± 17.91 L/m2), or half-life (mean, 2.63 ± 1.81 hours); a moderate correlation existed between dose given and systemic drug exposure (AUC). Substantial inhibition of peripheral blood mononuclear cell farnesyltransferase activity but near complete recovery by 24 hours was seen.

Conclusion: BMS-214667 was well tolerated as a weekly 1-hour i.v. infusion for four of six weeks with evidence of pharmacodynamic effect. The study was terminated before maximum tolerated dose was reached. Alternative schedules of drug administration might result in improved pharmacodynamic profile.
4-Hydroxytamoxifen Inhibits Proliferation of Multiple Myeloma Cells In vitro through Down-Regulation of c-Myc, Up-Regulation of p27Kip1, and Modulation of Bcl-2 Family Members
Authors: Gauduchon Juliette , Gouilleux Fabrice , Maillard Sébastien , Marsaud Véronique , Renoir Jack-Michel , Sola Brigitte
Published: September 2005

Purpose: Multiple myeloma is an incurable B-cell malignancy requiring new therapeutic strategies. Our approach was to analyze the in vitro effects of a selective estrogen receptor modulator, 4-hydroxytamoxifen (4-OHT), on six multiple myeloma cell lines.

Experimental Design: Cultured multiple myeloma cells were treated with various 4-OHT concentrations and the cellular response was studied: cell proliferation, cell viability, induction of apoptosis, caspase activities, and expression of signaling proteins.

Results: We found that pharmacologic concentrations of 4-OHT inhibit cell proliferation (4 of 6 cell lines). This inhibition is achieved by two independent events: a block at the G1 phase of the cell cycle and the induction of apoptotic death. The cellular response to 4-OHT depends on the presence of functional estrogen receptors. 4-OHT treatment activates an intrinsic mitochondrial caspase-9-dependent pathway but not the Fas/FasL death pathway. Signaling pathways known to be involved in the survival and/or proliferation of multiple myeloma cells are not affected by 4-OHT treatment. 4-OHT-induced G1 arrest is accompanied by the up-regulation of the cell cycle inhibitor p27Kip1 and the down-regulation of c-Myc. Among the Bcl-2 family members tested, the proapoptotic BimS protein is induced whereas the antiapoptotic protein Mcl-1 is decreased.

Conclusions: Although the effects of 4-OHT are observed at micromolar concentrations, cellular mechanisms responsible for G1 arrest, as well as apoptosis induction, are similar to those observed in breast cancer cells. Our data support the concept that 4-OHT may represent an alternative approach to inhibit proliferation and induce apoptosis of multiple myeloma cells.
Overexpression of 5-Lipoxygenase and Cyclooxygenase 2 in Hamster and Human Oral Cancer and Chemopreventive Effects of Zileuton and Celecoxib
Authors: Li Ning , Sood Sandeep , Wang Su , Fang Mingzhu , Wang Peng , Sun Zheng , Yang Chung S. , Chen Xiaoxin
Published: September 2005

Purpose: Previous studies have suggested an important role of aberrant arachidonic acid metabolism, especially the cyclooxygenase (Cox) pathway, in oral carcinogenesis. However, it is unknown whether the 5-lipoxygenase (5-Lox) pathway contributes to oral carcinogenesis, and whether combination of inhibitors of both pathways may have synergistic or additive effects of chemoprevention.

Experimental Design: 5-Lox expression was examined in 7,12-dimethylbenz[a]anthracene (DMBA)–induced hamster and human oral cancer tissues by immunohistochemistry, and Cox2 expression was investigated in hamster oral tissues using in situ hybridization. Zileuton (a specific 5-Lox inhibitor) and celecoxib (a specific Cox2 inhibitor), either alone or in combination, were investigated for their chemopreventive effects on the DMBA-induced hamster model at the post-initiation stage through topical application.

Results: 5-Lox was overexpressed during oral carcinogenesis in hamsters and humans, as well as Cox2 in the hamster tissues. In a chemoprevention study using the post-initiation DMBA model, incidence of hamster oral squamous cell carcinoma was reduced from 76.9% (20 of 26) to 45.8% (11 of 24, P < 0.05) and 32.1% (9 of 28, P < 0.01) by 3% and 6% topical zileuton, respectively; and to 57.6% (15 of 26, P > 0.05) and 50% (12 of 24, P < 0.05) by 3% and 6% topical celecoxib, respectively. When used in combination, celecoxib and zileuton (3% of each) had an additive inhibitory effect on the incidence of squamous cell carcinoma (36%, 9 of 25, P < 0.01). Other pathologic variables and the levels of leukotriene B4 and prostaglandin E2 of the hamster tissues were reduced as well.

Conclusions: The results clearly showed that both 5-Lox and Cox2 played important roles in oral carcinogenesis. Zileuton and celecoxib prevented oral carcinogenesis at the post-initiation stage through their inhibitory effects on arachidonic acid metabolism.
Frequent Immune Responses to a Cancer/Testis Antigen, CAGE, in Patients with Microsatellite instability–Positive Endometrial Cancer
Authors: Iwata Takashi , Fujita Tomonobu , Hirao Nobumaru , Matsuzaki Yuriko , Okada Tsutomu , Mochimaru Hiroshi , Susumu Nobuyuki , Matsumoto Eri , Sugano Kokichi , Yamashita Naohide , Nozawa Shiro , Kawakami Yutaka
Published: September 2005

Purpose: Identification of cancer/testis antigens useful for diagnosis or immunotherapy of cancers was attempted by cDNA expression cloning with patients' sera (SEREX).

Experimental Design: cDNA expression libraries made from testis or endometrial cancer cell lines were screened using sera from patients with endometrial cancer or melanoma patients immunized with dendritic cells pulsed with autologous tum or lysates. Tissue-specific expression by RT-PCR and immunogenicity by Western blotting of the bacterial recombinant antigen with sera from cancer patients were evaluated.

Results: A cancer/testis antigen, CAGE, was isolated by two independently performed SEREX. CAGE was expressed in various cancer cell lines including endometrial cancer, colon cancer, and melanoma in 7 of 10 endometrial cancer tissues and in 1 of 3 atypical endometrial hyperplasia, but not in normal tissues including the endometrium and testis. The protein expression on cancer cells was confirmed by Western blot analysis with the recombinant CAGE protein, anti-CAGE IgG antibody was detected in sera from 5 of 45 endometrial cancer, 2 of 24 melanoma, and 2 of 33 colon cancer patients, but not in sera from healthy individuals. By ELISA analysis, anti-CAGE antibody was detected in 12 of 45 endometrial cancer, 2 of 20 melanoma, and 4 of 33 colon cancer patients. Intriguingly, anti-CAGE antibody was highly positive in 7 of the 13 (53.8%) microsatellite instability (MSI)-H patients with endometrial cancer, but negative in 20 non–MSI-H patients (P = 0.001).

Conclusion: CAGE may be useful for immunotherapy and diagnosis of various cancers particularly MSI-positive endometrial cancer.
Effects of Imatinib on Monocyte-Derived Dendritic Cells Are Mediated by Inhibition of Nuclear Factor-κB and Akt Signaling Pathways
Authors: Appel Silke , Rupf Anette , Weck Markus M. , Schoor Oliver , Brümmendorf Tim H. , Weinschenk Toni , Grünebach Frank , Brossart Peter
Published: September 2005

Dendritic cells are the most powerful antigen-presenting cells playing a decisive role for the initiation and maintenance of primary immune responses. However, signaling pathways involved in the differentiation of these cells have not been fully determined. Imatinib is a novel tyrosine kinase inhibitor effective against Abl kinases, c-Kit, and platelet-derived growth factor receptor. Using this compound, we show that human monocyte-derived dendritic cells generated in the presence of therapeutic concentrations of imatinib show a reduced expression of CD1a, MHC class I and II, and costimulatory molecules as well as decreased secretion of chemokines and cytokines resulting in an impaired capacity of dendritic cells to elicit primary T-cell responses. Using Western blot analyses, we found that these effects are mediated by inhibition of phosphatidylinositol 3-kinase/Akt pathways and a pronounced down-regulation of nuclear localized protein levels of nuclear factor-κB family members. Importantly, using blocking antibodies and tyrosine kinase inhibitors, we show that the inhibitory effects of imatinib on dendritic cell differentiation are not mediated via platelet-derived growth factor receptor and c-Kit. Taken together, our study reveals that imatinib inhibits dendritic cell differentiation and function via Akt and nuclear factor-κB signal transduction. Importantly, we show that imatinib can inhibit the function of normal, nonmalignant cells that may result in immunosuppression of these patients.
Cyclosporin A Is a Broad-Spectrum Multidrug Resistance Modulator
Authors: Qadir Misbah , O'Loughlin Kieran L. , Fricke Stacy M. , Williamson Nicole A. , Greco William R. , Minderman Hans , Baer Maria R.
Published: September 2005

Purpose: Overexpression of the multidrug resistance proteins P-glycoprotein (Pgp), multidrug resistance protein (MRP-1), breast cancer resistance protein (BCRP), and lung resistance protein (LRP) is associated with treatment failure in acute myeloid leukemia (AML) and other malignancies. The Pgp modulator cyclosporin A has shown clinical efficacy in AML, whereas its analogue PSC-833 has not. Cyclosporin A is known to also modulate MRP-1, and we hypothesized that broad-spectrum multidrug resistance modulation might contribute to its clinical efficacy.

Experimental Design: We studied the effects of cyclosporin A and PSC-833 on in vitro drug retention and cytotoxicity in resistant cell lines overexpressing Pgp, MRP-1, and BCRP and on nuclear-cytoplasmic drug distribution and cytotoxicity in cells overexpressing LRP. Cellular drug content was assessed by flow cytometry and nuclear-cytoplasmic drug distribution by confocal microscopy.

Results: Cyclosporin A enhanced retention of the substrate drug mitoxantrone in cells overexpressing Pgp (HL60/VCR), MRP-1 (HL60/ADR), and BCRP (8226/MR20, HEK-293 482R) and increased cytotoxicity 6-, 4-, 4-, and 3-fold, respectively. Moreover, cyclosporin A enhanced nuclear distribution of doxorubicin in 8226/MR20 cells, which also express LRP, and increased doxorubicin cytotoxicity 12-fold without an effect on cellular doxorubicin content, consistent with expression of wild-type BCRP, which does not efflux doxorubicin. Cyclosporin A also enhanced nuclear doxorubicin distribution in a second cell line with LRP overexpression, HT1080/DR4. PSC-833 enhanced mitoxantrone retention and cytotoxicity in cells overexpressing Pgp, but had no effect in cells overexpressing MRP-1, BCRP, or LRP.

Conclusions: Cyclosporin A modulates Pgp, MRP-1, BCRP, and LRP, and this broad-spectrum activity may contribute to its clinical efficacy.
Combined Evaluation of Expressions of Cyclin E and p53 Proteins as Prognostic Factors for Patients with Gastric Cancer
Authors: Bani-Hani Kamal E. , Almasri Nidal M. , Khader Yousef S. , Sheyab Fawzi M. , Karam Hanan N.
Published: September 2005

Background: There is a lack of consistency regarding the prognostic value of cyclin E overexpression in gastric cancer (gastric cancer). Our aim was to report on this overexpression and to analyze its correlations with the clinicopathologic variables. Another aim was to examine if aberrant expression of both cyclin E and p53 might increase the malignant potential of gastric cancer.

Methods: Specimens from 89 patients with gastric cancer treated with “curative” intent were evaluated for cyclin E and p53 expressions using immunohistochemical method. The correlations between cyclin E overexpression alone or in combination with p53 expression and the patient's clinicopathologic variables were analyzed.

Results: Cyclin E overexpression and p53 expression were shown in 35 (39.3%) and 46 (51.7%) tumors, respectively. The incidence of cyclin E overexpression was significantly higher in deeply invasive cancers (P < 0.0001), in cancers with lymph node metastasis (P = 0.003), and in cancers with advanced stages (P < 0.0001). There were no significant correlations with other clinicopathologic variables. Patients in whom their tumors showed cyclin E overexpression alone or in combination with p53 survived less than patients with negative cyclin E tumors. Multivariate analysis revealed that combined cyclin E overexpression and p53 expression was significantly associated with poor survival after adjusting for other variables (hazard ratio, 3.12; P = 0.009).

Conclusions: Cyclin E overexpression is a common event in gastric cancer. Gastric cancer with cyclin E overexpression exhibit increased aggressiveness in the presence of aberrant p53. The combination of cyclin E overexpression with the p53 expression in gastric cancer further distinguished a subgroup of patients with poor prognosis.
Allelotypes and Fluorescence In situ Hybridization Profiles of Poorly Differentiated Endocrine Carcinomas of Different Sites
Authors: Furlan Daniela , Bernasconi Barbara , Uccella Silvia , Cerutti Roberta , Carnevali Ileana , Capella Carlo
Published: September 2005

Purpose: The aim of this work was to investigate the genotypic profiles of 36 poorly differentiated endocrine carcinoma (PDEC) of different sites to verify if their very similar phenotype may reflect similar pattern of genetic anomalies and if useful diagnostic or prognostic markers may be pointed out.

Experimental Design: All tumors were microallelotyped at 57 microsatellite on 11 autosomes and the allelotypes of a selected panel of tumors were validated by interphasic fluorescence in situ hybridization with centromeric probes for chromosomes 1, 3, 6, 11, 17, and 18 and a probe specific for p53.

Results: Regardless of the primary sites, PDECs exhibit very complex allelotypes (86%) and TP53 allelic imbalance (89%). Among these cases, fluorescence in situ hybridization analysis confirmed the presence of multiple aneusomies and a chromosome instability phenotype. Very low percentage of allelic imbalance (AI) and few aneuploidies were detected in only five PDECs for which an overall longer survival was observed. We found recurrent AI on 3p, 5, and 11q13 in lung PDECs, on 5q21, 8p, and 18q21 in colorectal PDECs and on 7 and 11q22 in gastric PDECs. Significantly better outcome was observed in patients with PDEC exhibiting 8q AIs and absence of AI at chromosome regions 6q25 and 6p.

Conclusions: The concurrence of p53 inactivation and aneuploidies or chromosome instability are the main features of PDECs. However, the specific allelotypes observed in relation to primary site support the hypothesis that PDECs and exocrine carcinomas of all sites may share early pathogenetic mechanisms. Molecular markers of potential diagnostic and prognostic values for PDECs of different sites have been identified.
Effect of YM529 on a Model of Mandibular Invasion by Oral Squamous Cell Carcinoma in Mice
Authors: Cui Nianhui , Nomura Takeshi , Noma Hiroyasu , Yokoo Keiko , Takagi Ryo , Hashimoto Sadamitsu , Okamoto Masato , Sato Mitsunobu , Yu Guangyan , Guo Chuanbin , Shibahala Takahiko
Published: September 2005

Purpose: This study examined the mechanisms of osteoclast-mediated bone invasion in a model of oral squamous cell carcinoma (OSCC). C3H/HeN mice were inoculated with SCC VII cells into the masseter region to establish an animal model of mandibular invasion by OSCC.

Experimental Design: The mice were divided into three groups: a control group, given daily s.c. injections of saline; group 1, given 2 μg per mouse per day of the bisphosphonate YM529; and group 2, given 10 μg per mouse per day of YM529. After 3 weeks of treatment, the lesions were studied by micro-computed tomography. After tartrate-resistant acid phosphatase (TRAP) staining, the osteoclasts were easily identified, and the percentages of the area occupied by osteoclasts were calculated by computer for each sample. The tumors were analyzed by RT-PCR to determine the mRNA expression of interleukin-6 (IL-6), parathyroid hormone–related protein (PTHrP), tumor necrosis factor-α (TNF-α), receptor activator of nuclear factor-κB (RANK), RANK ligand (RANKL), and osteoprotegerin.

Results: SCC VII cells rapidly multiplied in the masseter muscle of the mice. Bone invasion was evident only in the control group on micro-computed tomography. On TRAP-stained slices, the percentages of osteoclasts in groups 1 and 2 were significantly lower than that in the control group. The mRNA expressions of IL-6, PTHrP, THF-α, and RANK decreased as the concentration of YM529 increased.

Conclusions: We conclude that various cancer-derived cytokines play important roles in the invasion of bone by OSCC. YM529, a third-generation bisphosphonate, can suppress osteoclast-mediated bone invasion by OSCC. The mechanism of this effect might involve inhibition of cytokines such as IL-6, PTHrP, TNF-α, and RANK by YM529.
Mcm2, Geminin, and KI67 Define Proliferative State and are Prognostic Markers in Renal Cell Carcinoma
Authors: Dudderidge Tim J. , Stoeber Kai , Loddo Marco , Atkinson Geraldine , Fanshawe Thomas , Griffiths David F. , Williams Gareth H.
Published: September 2005

Purpose: The origin licensing factors minichromosome maintenance 2 (Mcm2) and Geminin have recently been identified as critical regulators of growth and differentiation. Here we have investigated the regulation of these licensing factors together with Ki67 to further elucidate the cell cycle kinetics of renal cell carcinoma (RCC). Furthermore, we have examined the role of Ki67, Mcm2, and Geminin in disease-free survival after nephrectomy in patients with localized RCC.

Experimental Design: Tissue sections from 176 radical nephrectomy specimens were immunohistochemically stained with Mcm2, Geminin, and Ki67 antibodies. Labeling indices (LI) for these markers were compared with clinicopathologic parameters (median follow-up 44 months).

Results: In RCC, Mcm2 is expressed at much higher levels than Ki-67 and Geminin, respectively [medians 41.6%, 7.3%, and 3.5% (P < 0.001)] and was most closely linked to tumor grade (P < 0.001). For each marker, Kaplan-Meier survival curves provided strong evidence that increased expression is associated with reduced disease-free survival time (P < 0.001). Additionally, an Mcm2 − Ki67 LI identified a unique licensed but nonproliferating population of tumor cells that increased significantly with tumor grade (P = 0.004) and was also of prognostic value (P = 0.01). On multivariate analysis, grade, vascular invasion, capsular invasion, Ki67 LI >12%, and age were found to be independent prognostic markers.

Conclusions: Although Ki67 is identified as an independent prognostic marker, semiquantitative assessment is difficult due to the very low proliferative fraction identified by this marker. In contrast, Mcm2 identifies an increased growth fraction that is closely linked to grade, provides prognostic information, and is amenable to semiquantitative analysis in routine pathologic assessment.
2-Methoxyestradiol Is an Estrogen Receptor Agonist That Supports Tumor Growth in Murine Xenograft Models of Breast Cancer
Authors: Sutherland Tara E. , Schuliga Michael , Harris Trudi , Eckhardt Bedrich L. , Anderson Robin L. , Quan Lilly , Stewart Alastair G.
Published: September 2005

Purpose: 2-Methoxyestradiol (2MEO) is being developed as a novel antitumor agent based on its antiangiogenic activity, tumor cell cytotoxicity, and apparent lack of toxicity. However, pharmacologic concentrations of 2MEO bind to estrogen receptors (ER). We have therefore examined the ER activity of 2MEO.

Experimental Design: Estrogenic actions of 2MEO were evaluated by changes in gene expression of the ER-positive (MCF7) breast tumor cell line and, in vivo, estrogenicity was assessed in breast tumor xenograft models and by measuring endocrine responses in uterus and liver.

Results: In the ER-positive breast tumor cell line (MCF7), microarray experiments revealed that 269 of 279 changes in gene expression common to 2MEO and estradiol were prevented by the ER antagonist, ICI 182,780. Changes in the expression of selected genes and their sensitivity to inhibition by ICI 182,780 were confirmed by quantitative reverse transcription–PCR measurement. Activation of ER in MCF7 cells by 2MEO was further confirmed by stimulation of an estrogen response element–dependent reporter gene that was blocked by ICI 182,780 (1 μmol/L). Doses of 2MEO (15-150 mg/kg) that had no antitumor efficacy in either nu/nu BALB/c or severe combined immunodeficient mice bearing ER-negative MDA-MB-435 tumors had uterotropic and hepatic estrogen-like actions. In female nu/nu BALB/c mice inoculated with the estrogen-dependent MCF7 tumor cells, 2MEO (50 mg/kg/d) supported tumor growth.

Conclusions: Tumor growth enhancement by 2MEO at doses generating serum levels (100-500 nmol/L) that have estrogenic activity suggests that a conservative approach to the further clinical evaluation of this agent should be adopted and that its evaluation in breast cancer is inappropriate.