bluuebunny/crossref_metadata_2025 huggingface dataset:
Anti–Transforming Growth Factor β Receptor II Antibody Has Therapeutic Efficacy against Primary Tumor Growth and Metastasis through Multieffects on Cancer, Stroma, and Immune Cells
Authors: Zhong Zhaojing , Carroll Kyla Driscoll , Policarpio Desiree , Osborn Carla , Gregory Michael , Bassi Rajiv , Jimenez Xenia , Prewett Marie , Liebisch Gregory , Persaud Kris , Burtrum Douglas , Wang Su , Surguladze David , Ng Stanley , Griffith Heather , Balderes Paul , Doody Jacqueline , Schwartz Jonathan D. , Youssoufian Hagop , Rowinsky Eric K. , Ludwig Dale L. , Witte Larry , Zhu Zhenping , Wu Yan
Published: February 2010

Purpose: Transforming growth factor β (TGFβ) is a pleiotropic cytokine that affects tumor growth, metastasis, stroma, and immune response. We investigated the therapeutic efficacy of anti–TGFβ receptor II (TGFβ RII) antibody in controlling metastasis and tumor growth as well as enhancing antitumor immunity in preclinical tumor models.

Experimental Design: We generated neutralizing antibodies to TGFβ RII and assessed the antibody effects on cancer, stroma, and immune cells in vitro. The efficacy and mechanism of action of the antibody as monotherapy and in combination with chemotherapy in suppression of primary tumor growth and metastasis were evaluated in several tumor models.

Results: Anti–TGFβ RII antibody blocked TGFβ RII binding to TGFβ 1, 2, and 3, and attenuated the TGFβ-mediated activation of downstream Smad2 kinase, invasion of cancer cells, motility of endothelial and fibroblast cells, and induction of immunosuppressive cells. Treatment with the antibody significantly suppressed primary tumor growth and metastasis and enhanced natural killer and CTL activity in tumor-bearing mice. Immunohistochemistry analysis showed cancer cell apoptosis and massive necrosis, and increased tumor-infiltrating T effector cells and decreased tumor-infiltrating Gr-1+ myeloid cells in the antibody-treated tumors. Fluorescence-activated cell sorting analysis indicated the significant reduction of peripheral Gr-1+/CD11b+ myeloid cells in treated animals. Concomitant treatment with the cytotoxic agent cyclophosphamide resulted in a significantly increased antitumor efficacy against primary tumor growth and metastasis.

Conclusions: These preclinical data provide a foundation to support using anti–TGFβ RII antibody as a therapeutic agent for TGFβ RII–dependent cancer with metastatic capacity. Clin Cancer Res; 16(4); 1191–205
Low-Molecular-Weight Cyclin E Can Bypass Letrozole-Induced G1 Arrest in Human Breast Cancer Cells and Tumors
Authors: Akli Said , Bui Tuyen , Wingate Hannah , Biernacka Anna , Moulder Stacy , Tucker Susan L. , Hunt Kelly K. , Keyomarsi Khandan
Published: February 2010

Purpose: Low-molecular-weight cyclin E (LMW-E) in breast cancer cells induces genomic instability and resistance to inhibition by p21, p27, and fulvestrant therapy. Here, we sought to determine if LMW-E renders breast cancer cells unresponsive to aromatase inhibitors (AI), elucidate the mechanism of such resistance, and ascertain if inhibitors of LMW-E–associated kinase activity could overcome this resistance.

Experimental Design: The antiproliferative effects of the AIs were examined in aromatase-overexpressing MCF-7/Ac1 cells in the presence or absence of full-length cyclin E and LMW-E. Inhibition of LMW cyclin E kinase activity by roscovitine [a cyclin-dependent kinase (CDK) inhibitor] was examined in letrozole-unresponsive MCF-7/Ac1 cells. The role of LMW-E and CDK2 in mediating recurrence following AI treatment was also assessed in breast cancer patients.

Results: Overexpression of LMW-E in postmenopausal patients was associated with a poor prognosis. Letrozole, but not exemestane or anastrozole, mediated a pronounced G1 arrest in MCF-7/Ac1 cells. Androstenedione-induced G1 exit correlated with increased cyclin E–associated kinase activity and increased CDK2 levels. Letrozole treatment inhibited cyclin E-CDK2 kinase activity by preventing the androstenedione-induced increase in CDK2. LMW-E bypassed this effect and rendered the cells resistant to letrozole inhibition. Roscovitine blocked the androstenedione-induced increase in CDK2, and LMW-E overexpression could not bypass this effect. Lastly, breast cancer patients whose tumors overexpress LMW-E were not responsive to AI treatment.

Conclusions: Roscovitine treatment can reverse intrinsic or acquired resistance to letrozole due to LMW-E expression in breast cancer cells. These data support the clinical investigation of CDK2 inhibitor therapy for postmenopausal women with estrogen receptor–positive, LMW-E–expressing breast cancer. Clin Cancer Res; 16(4); 1179–90
High Frequency of p53/MDM2/p14ARF Pathway Abnormalities in Relapsed Neuroblastoma
Authors: Carr-Wilkinson Jane , O'Toole Kieran , Wood Katrina M. , Challen Christine C. , Baker Angela G. , Board Julian R. , Evans Laura , Cole Michael , Cheung Nai-Kong V. , Boos Joachim , Köhler Gabriele , Leuschner Ivo , Pearson Andrew D.J. , Lunec John , Tweddle Deborah A.
Published: February 2010

Purpose: Most neuroblastomas initially respond to therapy but many relapse with chemoresistant disease. p53 mutations are rare in diagnostic neuroblastomas, but we have previously reported inactivation of the p53/MDM2/p14ARF pathway in 9 of 17 (53%) neuroblastoma cell lines established at relapse.

Hypothesis: Inactivation of the p53/MDM2/p14ARF pathway develops during treatment and contributes to neuroblastoma relapse.

Methods: Eighty-four neuroblastomas were studied from 41 patients with relapsed neuroblastoma including 38 paired neuroblastomas at different stages of therapy. p53 mutations were detected by automated sequencing, p14ARF methylation and deletion by methylation-specific PCR and duplex PCR, respectively, and MDM2 amplification by fluorescent in situ hybridization.

Results: Abnormalities in the p53 pathway were identified in 20 of 41 (49%) cases. Downstream defects due to inactivating missense p53 mutations were identified in 6 of 41 (15%) cases, 5 following chemotherapy and/or at relapse and 1 at diagnosis, postchemotherapy, and relapse. The presence of a p53 mutation was independently prognostic for overall survival (hazard ratio, 3.4; 95% confidence interval, 1.2–9.9; P = 0.02). Upstream defects were present in 35% of cases: MDM2 amplification in 3 cases, all at diagnosis and relapse and p14ARF inactivation in 12 of 41 (29%) cases: 3 had p14ARF methylation, 2 after chemotherapy, and 9 had homozygous deletions, 8 at diagnosis and relapse.

Conclusions: These results show that a high proportion of neuroblastomas which relapse have an abnormality in the p53 pathway. The majority have upstream defects suggesting that agents which reactivate wild-type p53 would be beneficial, in contrast to those with downstream defects in which p53-independent therapies are indicated. Clin Cancer Res; 16(4); 1108–18
Comprehensive MicroRNA Profiling for Head and Neck Squamous Cell Carcinomas
Authors: Hui Angela B.Y. , Lenarduzzi Michelle , Krushel Tiffaney , Waldron Levi , Pintilie Melania , Shi Wei , Perez-Ordonez Bayardo , Jurisica Igor , O'Sullivan Brian , Waldron John , Gullane Pat , Cummings Bernard , Liu Fei-Fei
Published: February 2010

Purpose: The objective of this study is to investigate the significance of microRNAs (miRNA) in patients with locally advanced head and neck squamous cell carcinoma (HNSCC).

Experimental Design: A global miRNA profiling was done on 51 formalin-fixed archival HNSCC samples using quantitative reverse transcription-PCR approach, correlated with patients' clinical parameters. Functional characterization of HNSCC-associated miRNAs was conducted on three HNSCC cell lines. Cell viability and proliferation were investigated using MTS and clonogenic assays, respectively; cell cycle analyses were assessed using flow cytometry.

Results: Thirty-eight of the 117 (33%) consistently detected miRNAs were significantly differentially expressed between malignant versus normal tissues. Concordant with previous reports, overexpression of miR-21, miR-155, let-7i, and miR-142-3p and underexpression of miR-125b and miR-375 were detected. Upregulation of miR-423, miR-106b, miR-20a, and miR-16 as well as downregulation of miR-10a were newly observed. Exogenous overexpression of miR-375 in HNSCC cell lines reduced proliferation and clonogenicity and increased cells in sub-G1. Similar cellular effects were observed in knockdown studies of the miR-106b-25 cluster but with accumulation of cells in G1 arrest. No major difference was detected in miRNA profiles among laryngeal, oropharyngeal, or hypopharyngeal cancers. miR-451 was found to be the only significantly overexpressed miRNA by 4.7-fold between nonrelapsed and relapsed patients.

Conclusion: We have identified a group of aberrantly expressed miRNAs in HNSCC and showed that underexpression of miR-375 and overexpression of miR-106b-25 cluster might play oncogenic roles in this disease. Further detailed examinations of miRNAs will provide opportunities to dissect the complex molecular abnormalities driving HNSCC progression. Clin Cancer Res; 16(4); 1129–39
Metastatic Renal Cell Carcinoma Treated with Sunitinib: Early Evaluation of Treatment Response Using Dynamic Contrast-Enhanced Ultrasonography
Authors: Lassau Nathalie , Koscielny Serge , Albiges Laurence , Chami Linda , Benatsou Baya , Chebil Mohamed , Roche Alain , Escudier Bernard J.
Published: February 2010

Purpose: To determine the utility of dynamic contrast-enhanced ultrasonography (DCE-US) as a prognostic tool for metastatic renal cell carcinoma patients receiving sunitinib and to identify DCE-US parameters that correlate with early treatment response.

Experimental Design: Thirty-eight patients received 50 mg/d sunitinib on schedule 4/2 (4 weeks on followed by 2 weeks off treatment). After two cycles, response evaluation criteria in solid tumors were used to classify patients as responders or nonresponders. DCE-US evaluations were done before treatment and at day 15; variations between days 0 and 15 were calculated for seven DCE-US functional parameters and were compared for responders and nonresponders. The correlation between DCE-US parameters and disease-free survival (DFS) and overall survival (OS) was assessed.

Results: The ratio between DCE-US examinations at baseline and day 15 significantly correlated with response in five of the seven DCE-US parameters. Two DCE-US parameters (time to peak intensity and slope of the wash-in) were significantly associated with DFS; time to peak intensity was also significantly associated with OS.

Conclusions: DCE-US is a useful tool for predicting the early efficacy of sunitinib in metastatic renal cell carcinoma patients. Robust correlations were observed between functional parameters and classic assessments, including DFS and OS. Clin Cancer Res; 16(4); 1216–25
MutS Homologue 2 and the Long-term Benefit of Adjuvant Chemotherapy in Lung Cancer
Authors:   for the International Adjuvant Lung Trial-Bio investigators, Kamal Nermine S. , Soria Jean-Charles , Mendiboure Jean , Planchard David , Olaussen Ken A. , Rousseau Vanessa , Popper Helmut , Pirker Robert , Bertrand Pascale , Dunant Ariane , Le Chevalier Thierry , Filipits Martin , Fouret Pierre
Published: February 2010

Purpose: We sought to determine the long-term (median follow-up, 7.5 years) predictive power of human MutS homologue 2 (MSH2) immunohistochemical expression in patients who enrolled in the International Adjuvant Lung Trial.

Experimental design: We tested the interaction between MSH2 and the allocated treatment (chemotherapy versus observation) in a Cox model adjusted on clinicopathologic variables. The significance level was set at 0.01.

Results: MSH2 levels were low in 257 (38%) and high in 416 (62%) tumors. The benefit from chemotherapy was likely different according to MSH2 (interaction test, P = 0.06): there was a trend for chemotherapy to prolong overall survival when MSH2 was low [hazard ratio (HR), 0.76; 95% confidence interval (95% CI), 0.59-0.97; P = 0.03], but not when MSH2 was high (HR, 1.12; 95% CI, 0.81-1.55; P = 0.48). In the control arm, the HR was 0.66 (95% CI, 0.49-0.90; P = 0.01) when MSH2 was high. When combining MSH2 with excision repair cross-complementing group 1 (ERCC1) into four subgroups, the benefit of chemotherapy decreased with the number of markers expressed at high levels (P = 0.01). A similar decrease was noted when combining MSH2 and P27 (P = 0.01). Chemotherapy prolonged overall survival in the combined low MSH2/low ERCC1 subgroup (HR, 0.65; 95% CI, 0.47-0.91; P = 0.01) and in the combined low MSH2/low P27 subgroup (HR, 0.65; 95% CI, 0.46-0.93; P = 0.01).

Conclusions: MSH2 expression is a borderline significant predictor of a long-term benefit from adjuvant cisplatin-based chemotherapy in patients with completely resected lung cancer. MSH2 combined with ERCC1 or P27 may identify patients most likely to benefit durably from chemotherapy. Clin Cancer Res; 16(4); 1206–15
Tobacco Use in Human Papillomavirus–Positive Advanced Oropharynx Cancer Patients Related to Increased Risk of Distant Metastases and Tumor Recurrence
Authors: Maxwell Jessica H. , Kumar Bhavna , Feng Felix Y. , Worden Francis P. , Lee Julia S. , Eisbruch Avraham , Wolf Gregory T. , Prince Mark E. , Moyer Jeffrey S. , Teknos Theodoros N. , Chepeha Douglas B. , McHugh Jonathan B. , Urba Susan G. , Stoerker Jay , Walline Heather M. , Kurnit David M. , Cordell Kitrina G. , Davis Samantha J. , Ward Preston D. , Bradford Carol R. , Carey Thomas E.
Published: February 2010

Purpose: The goal of this study was to examine the effect of tobacco use on disease recurrence (local/regional recurrence, distant metastasis, or second primary) among patients with human papillomavirus (HPV)–positive squamous cell carcinoma of the oropharynx (SCCOP) following a complete response to chemoradiation therapy.

Experimental Design: Between 1999 and 2007, 124 patients with advanced SCCOP (86% with stage IV) and adequate tumor tissue for HPV analysis who were enrolled in one of two consecutive University of Michigan treatment protocols were prospectively included in this study. Patients were categorized as never-, former, or current tobacco users. The primary end points were risk of disease recurrence and time to recurrence; secondary end points were disease-specific survival and overall survival.

Results: One hundred and two patients (82.3%) had HPV-positive tumors. Over two thirds (68%) of patients with HPV-positive tumors were tobacco users. Among HPV-positive patients, current tobacco users were at significantly higher risk of disease recurrence than never-tobacco users (hazard ratio, 5.2; confidence interval, 1.1-24.4; P = 0.038). Thirty-five percent of HPV-positive ever tobacco users recurred compared with only 6% of HPV-positive never users and 50% of HPV-negative patients. All HPV-negative patients were tobacco users and had significantly shorter times to recurrence (P = 0.002), and had reduced disease-specific survival (P = 0.004) and overall survival (P < 0.001) compared with HPV-positive patients. Compared with HPV-positive never-tobacco users, those with a tobacco history showed a trend for reduced disease-specific survival (P = 0.064) but not overall survival (P = 0.221).

Conclusions: Current tobacco users with advanced, HPV-positive SCCOP are at higher risk of disease recurrence compared with never-tobacco users. Clin Cancer Res; 16(4); 1226–35
Dasatinib Inhibits the Growth of Molecularly Heterogeneous Myeloid Leukemias
Authors: Guerrouahen Bella S. , Futami Muneyoshi , Vaklavas Christos , Kanerva Jukka , Whichard Zakary L. , Nwawka Kenechi , Blanchard Elisabeth G. , Lee Francis Y. , Robinson Lisa J. , Arceci Robert , Kornblau Steven M. , Wieder Eric , Cayre Yvon E. , Corey Seth J.
Published: February 2010

Purpose: Dasatinib is a dual Src/Abl inhibitor recently approved for Bcr-Abl+ leukemias with resistance or intolerance to prior therapy. Because Src kinases contribute to multiple blood cell functions by triggering a variety of signaling pathways, we hypothesized that their molecular targeting might lead to growth inhibition in acute myeloid leukemia (AML).

Experimental Design: We studied growth factor–dependent and growth factor–independent leukemic cell lines, including three cell lines expressing mutants of receptor tyrosine kinases (Flt3 or c-Kit) as well as primary AML blasts for responsiveness to dasatinib.

Results: Dasatinib resulted in the inhibition of Src family kinases in all cell lines and blast cells at ∼1 × 10−9 mol/L. It also inhibited mutant Flt3 or Kit tyrosine phosphorylation at ∼1 × 10−6 mol/L. Mo7e cells expressing the activating mutation (codon 816) of c-Kit were most sensitive to growth inhibition with a GI50 of 5 × 10−9 mol/L. Primary AML blast cells exhibited a growth inhibition of <1 × 10−6 mol/L. Cell lines that showed growth inhibition at ∼1 × 10−6 mol/L showed a G1 cell cycle arrest and correlated with accumulation of p21 and p27 protein. The addition of rapamycin or cytotoxic agents enhanced growth inhibition. Dasatinib also caused the apoptosis of Mo7e cells expressing oncogenic Kit.

Conclusions: Although all of the precise targets for dasatinib are not known, this multikinase inhibitor causes either growth arrest or apoptosis in molecularly heterogeneous AML. The addition of cytotoxic or targeted agents can enhance its effects. Clin Cancer Res; 16(4); 1149–58
X-Linked Ectodermal Dysplasia Receptor Is Downregulated in Breast Cancer via Promoter Methylation
Authors: Punj Vasu , Matta Hittu , Chaudhary Preet M.
Published: February 2010

Purpose: The X-linked ectodermal dysplasia receptor (XEDAR) is a novel receptor of the tumor necrosis factor receptor family that binds to ectodysplasin-A2 (EDA-A2) and induces cell death. The purpose of this study was to determine the tumor-suppressive potential of XEDAR in the development of breast cancer.

Experimental Design: We analyzed the expression of XEDAR in breast cancer cell lines and tumor samples using quantitative real-time PCR analysis and immunoblotting. We analyzed the human XEDAR gene promoter for the presence of any CpG island and examined its methylation status using methylation-specific real-time PCR. We examined the effect of 5-aza-2′-deoxycytidine on the expression of XEDAR and sensitivity to EDA-A2–induced apoptosis in breast cancer cell lines.

Results: Expression of XEDAR, but not EDA-A2, was downregulated in most tumorigenic breast cancer cell lines and tumor samples. Loss of XEDAR expression correlated with the hypermethylation of its promoter. Ectopic expression of XEDAR in MDA-MB-231 cells resulted in significant induction of apoptosis and reduction in colony formation. Treatment with 5-aza-2′-deoxycytidine restored XEDAR expression in breast cancer cell lines with methylated XEDAR promoter and sensitized them to EDA-A2–induced cell death.

Conclusions: Our results suggest that XEDAR expression is downregulated in most breast cancers via promoter methylation, which may contribute to accelerated tumor development by blocking EDA-A2–induced cell death. XEDAR may represent a novel breast tumor suppressor gene, and restoration of its expression by treatment with DNA demethylating agents may represent an attractive approach for the treatment of breast cancer. Clin Cancer Res; 16(4); 1140–8
Frequent Downregulation of miR-34 Family in Human Ovarian Cancers
Authors: Corney David C. , Hwang Chang-Il , Matoso Andres , Vogt Markus , Flesken-Nikitin Andrea , Godwin Andrew K. , Kamat Aparna A. , Sood Anil K. , Ellenson Lora H. , Hermeking Heiko , Nikitin Alexander Yu.
Published: February 2010

Purpose: The miR-34 family is directly transactivated by tumor suppressor p53, which is frequently mutated in human epithelial ovarian cancer (EOC). We hypothesized that miR-34 expression would be decreased in EOC and that reconstituted miR-34 expression might reduce cell proliferation and invasion of EOC cells.

Experimental Designs: miR-34 expression was determined by quantitative reverse transcription-PCR and in situ hybridization in a panel of 83 human EOC samples. Functional characterization of miR-34 was accomplished by reconstitution of miR-34 expression in EOC cells with synthetic pre-miR molecules followed by determining changes in proliferation, apoptosis, and invasion.

Results: miR-34a expression is decreased in 100%, and miR-34b*/c in 72%, of EOC with p53 mutation, whereas miR-34a is also downregulated in 93% of tumors with wild-type p53. Furthermore, expression of miR-34b*/c is significantly reduced in stage IV tumors compared with stage III (P = 0.0171 and P = 0.0029, respectively). Additionally, we observed promoter methylation and copy number variations at mir-34. In situ hybridization showed that miR-34a expression is inversely correlated with MET immunohistochemical staining, consistent with translational inhibition by miR-34a. Finally, miR-34 reconstitution experiments in p53 mutant EOC cells resulted in reduced proliferation, motility, and invasion, the latter of which was dependent on MET expression.

Conclusions: Our work suggests that miR-34 family plays an important role in EOC pathogenesis and reduced expression of miR-34b*/c may be particularly important for progression to the most advanced stages. Part of miR-34 effects on motility and invasion may be explained by regulation of MET, which is frequently overexpressed in EOC. Clin Cancer Res; 16(4); 1119–28
A 3′-Untranslated Region Polymorphism in IGF1 Predicts Survival of Non–Small Cell Lung Cancer in a Chinese Population
Authors: Zhang Mingfeng , Hu Zhibin , Huang Jinlin , Shu Yongqian , Dai Juncheng , Jin Guangfu , Tang Rong , Dong Jing , Chen Yijiang , Xu Lin , Huang Xinen , Shen Hongbing
Published: February 2010

Purpose: Disruption of the balance of insulin-like growth factor I (IGF-I) and IGF-binding protein 3 (IGFBP3) has been implicated in the etiology and progression of lung and other cancers. Single nucleotide polymorphisms (SNP) in IGF1 and IGFBP3 have been reported to be associated with the expression of the IGF-I/IGFBP3 axis. Therefore, we hypothesized that SNPs in these two genes were associated with lung cancer survival.

Experimental Design: We selected and genotyped 21 tagging and potentially functional SNPs in IGF1 and IGFBP3 by using Illumina Goldengate Genotyping Chip in a case cohort of 568 patients diagnosed with non–small cell lung cancer (NSCLC) in a Chinese population. Log-rank test and Cox proportional hazard models were used for the survival analyses.

Results: We found that rs5742714C/G in the 3′-untranslated region of IGF1 was associated significantly with NSCLC survival after adjustment for demographic and clinicopathologic factors, showing an improved median survival time in patients carrying variant CG/GG genotypes [median survival time, 28.5 months for CG/GG and 23.0 for CC; crude hazard ratio (HR), 0.74; 95% confidence interval (95% CI), 0.57-0.95, and adjusted HR, 0.77; 95% CI, 0.60-0.99]. This protective effect was more predominant for patients receiving surgical operation (HR, 0.58; 95% CI, 0.40-0.85; P for heterogeneity test = 0.045), along with a significant multiplicative interaction between variant genotypes and operation status (P = 0.028).

Conclusions: Our findings suggest that rs5742714 in IGF1 may be a genetic modifier for NSCLC prognosis in this Chinese population, especially among patients with surgical operation. Clin Cancer Res; 16(4); 1236–44
Finding the Right Dose for Cancer Therapeutics—Can We Do Better?
Authors: Rubin Eric H. , Anderson Keaven M.
Published: February 2010

Unlike other diseases, dose-selection for cancer therapeutics is often based on the maximum-tolerated dose in phase 1 studies involving relatively few patients. In this issue of Clinical Cancer Research, Jain and colleagues provide evidence that lower doses may be as effective as maximum-tolerated doses in the treatment of cancer patients. Clin Cancer Res; 16(4); 1085–7
The Mismatch Repair System Modulates Curcumin Sensitivity through Induction of DNA Strand Breaks and Activation of G2-M Checkpoint
Authors: Jiang Zhihua , Jin ShunQian , Yalowich Jack C. , Brown Kevin D. , Rajasekaran Baskaran
Published: February 2010

The highly conserved mismatch (MMR) repair system corrects postreplicative errors and modulates cellular responses to genotoxic agents. Here, we show that the MMR system strongly influences cellular sensitivity to curcumin. Compared with MMR-proficient cells, isogenically matched MMR-deficient cells displayed enhanced sensitivity to curcumin. Similarly, cells suppressed for MLH1 or MSH2 expression by RNA interference displayed increased curcumin sensitivity. Curcumin treatment generated comparable levels of reactive oxygen species and the mutagenic adduct 8-oxo-guanine in MMR-proficient and MMR-deficient cells; however, accumulation of γH2AX foci, a marker for DNA double-strand breaks (DSB), occurred only in MMR-positive cells in response to curcumin treatment. Additionally, MMR-positive cells showed activation of Chk1 and induction of G2-M cell cycle checkpoint following curcumin treatment and inhibition of Chk1 by UCN-01 abrogated Chk1 activation and heightened apoptosis in MMR-proficient cells. These results indicate that curcumin triggers the accumulation of DNA DSB and induction of a checkpoint response through a MMR-dependent mechanism. Conversely, in MMR-compromised cells, curcumin-induced DSB is significantly blunted, and as a result, cells fail to undergo cell cycle arrest, enter mitosis, and die through mitotic catastrophe. The results have potential therapeutic value, especially in the treatment of tumors with compromised MMR function. Mol Cancer Ther; 9(3); 558–68
Urinary-Type Plasminogen Activator Receptor/α3β1 Integrin Signaling, Altered Gene Expression, and Oral Tumor Progression
Authors: Ghosh Supurna , Koblinski Jennifer , Johnson Jeffrey , Liu Yueying , Ericsson Aaron , Davis J. Wade , Shi Zonggao , Ravosa Matthew J. , Crawford Susan , Frazier Shellaine , Stack M. Sharon
Published: February 2010

Oral squamous cell carcinoma (OSCC) has 50% 5-year survival rate, highlighting our limited understanding of the molecular events that contribute to disease progression. Microarray analyses of primary oral tumors have identified urinary-type plasminogen activator (uPA) and its receptor (uPAR) as key genes associated with human OSCC progression. The uPAR functions as both a proteinase receptor and an integrin ligand, modifying proteolysis, migration, integrin signaling, and cellular transcription. In the current study, uPAR expression levels were modified in OSCC cells followed by analysis of tumor growth in an in vivo orthotopic xenograft model and by transcriptional profiling. Overexpression of uPAR resulted in more infiltrative and less differentiated tumors, with ill-defined borders, cytologic atypia, and enhanced vascularity. Analysis of serial sections of both murine experimental tumors and microarrayed human OSCC showed a statistically significant association between uPAR and α3 integrin colocalization in areas exhibiting extracellular signal-regulated kinase phosphorylation, suggesting that uPAR/α3 integrin interaction potentiates extracellular signal-regulated kinase signaling in vivo. This is supported by cDNA microarray analysis, which showed differential expression of 148 genes (113 upregulated and 35 downregulated). Validation of gene expression changes in human OSCC using immunohistochemistry and quantitative real-time PCR showed increased growth factors, proteinases/inhibitors, and matrix components in uPAR-overexpressing tumors. Together, these results support a model wherein increased uPAR expression promotes α3β1 integrin association, resulting in increased mitogen-activated protein kinase signaling and transcriptional activation, leading to the formation of more aggressive tongue tumors. This combined approach has efficacy to identify additional biomarkers and/or prognostic indicators associated with aggressive human OSCC. Mol Cancer Res; 8(2); 145–58
FEN1 Functions in Long Patch Base Excision Repair Under Conditions of Oxidative Stress in Vertebrate Cells
Authors: Asagoshi Kenjiro , Tano Keizo , Chastain Paul D. , Adachi Noritaka , Sonoda Eiichiro , Kikuchi Koji , Koyama Hideki , Nagata Kenji , Kaufman David G. , Takeda Shunichi , Wilson Samuel H. , Watanabe Masami , Swenberg James A. , Nakamura Jun
Published: February 2010

From in vitro studies, flap endonuclease 1 (FEN1) has been proposed to play a role in the long patch (LP) base excision repair (BER) subpathway. Yet the role of FEN1 in BER in the context of the living vertebrate cell has not been thoroughly explored. In the present study, we cloned a DT40 chicken cell line with a deletion in the FEN1 gene and found that these FEN1-deficient cells exhibited hypersensitivity to H2O2. This oxidant produces genotoxic lesions that are repaired by BER, suggesting that the cells have a deficiency in BER affecting survival. In experiments with extracts from the isogenic FEN1 null and wild-type cell lines, the LP-BER activity of FEN1 null cells was deficient, whereas repair by the single-nucleotide BER subpathway was normal. Other consequences of the FEN1 deficiency were also evaluated. These results illustrate that FEN1 plays a role in LP-BER in higher eukaryotes, presumably by processing the flap-containing intermediates of BER. Mol Cancer Res; 8(2); 204–15
Phosphorylation of p53 Serine 18 Upregulates Apoptosis to Suppress Myc-Induced Tumorigenesis
Authors: Sluss Hayla K. , Gannon Hugh , Coles Andrew H. , Shen Qichang , Eischen Christine M. , Jones Stephen N.
Published: February 2010

ATM and p53 are critical regulators of the cellular DNA damage response and function as potent tumor suppressors. In cells undergoing ionizing radiation, ATM is activated by double-strand DNA breaks and phosphorylates the NH2 terminus of p53 at serine residue 18. We have previously generated mice bearing an amino acid substitution at this position (p53S18A) and documented a role for p53 phosphorylation in DNA damage–induced apoptosis. In this present study, we have crossed Eμmyc transgenic mice with our p53S18A mice to explore a role for ATM-p53 signaling in response to oncogene-induced tumorigenesis. Similar to DNA damage induced by ionizing radiation, expression of c-Myc in pre–B cells induces p53 serine18 phosphorylation and Puma expression to promote apoptosis. Eμmyc transgenic mice develop B-cell lymphoma more rapidly when heterozygous or homozygous for p53S18A alleles. However, Eμmyc-induced tumorigenesis in p53S18A mice is slower than that observed in Eμmyc mice deficient for either p53 or ATM, indicating that both p53-induced apoptosis and p53-induced growth arrest contribute to the suppression of B-cell lymphoma formation in Eμmyc mice. These findings further reveal that oncogene expression and DNA damage activate the same ATM-p53 signaling cascade in vivo to regulate apoptosis and tumorigenesis. Mol Cancer Res; 8(2); 216–22
Bcl-2 Overexpression Induces a Partial Epithelial to Mesenchymal Transition and Promotes Squamous Carcinoma Cell Invasion and Metastasis
Authors: Zuo Jianhong , Ishikawa Tohru , Boutros Shadi , Xiao Zhiqiang , Humtsoe Joseph O. , Kramer Randall H.
Published: February 2010

Evidence shows that Bcl-2 family members play a direct role in the development of some human malignancies. However, the mechanism by which Bcl-2 may influence tumor cell invasion and metastasis remains unclear. Ectopic overexpression of Bcl-2 in the human squamous carcinoma cell line HSC-3 enhanced tumorigenicity and experimental pulmonary metastasis. Interestingly, Bcl-2–expressing cells showed morphologic changes that resembled that of cells with an epithelial-mesenchymal transition phenotype. Analysis revealed increased N-cadherin and vimentin expression in parallel with attenuated E-cadherin level, along with enhanced migration and invasive behavior. Zymography studies confirmed elevated levels of matrix metalloproteinase-9 (MMP-9) in media of Bcl-2–expressing cells. siRNA-mediated suppression of N-cadherin expression not only prevented the enhanced invasion but also blocked the increased MMP-9 expression induced by elevated Bcl-2 expression. Accordingly, pharmacologic inhibition of MMP-9 abrogated the increased tumor cell invasion. Furthermore, the Bcl-2–mediated increase in MMP-9 expression and tumor cell invasion was dependent on fibroblast growth factor receptor-1 or extracellular signal–regulated kinase signaling. Collectively, the data establish that Bcl-2 overexpression in squamous carcinoma cells induces a partial epithelial to mesenchymal transition that promotes not only survival but also invasion and metastasis through the N-cadherin/fibroblast growth factor receptor/extracellular signal–regulated kinase pathway. Mol Cancer Res; 8(2); 170–82
Cyclooxygenase-2 Is a Novel Transcriptional Target of the Nuclear EGFR-STAT3 and EGFRvIII-STAT3 Signaling Axes
Authors: Lo Hui-Wen , Cao Xinyu , Zhu Hu , Ali-Osman Francis
Published: February 2010

Emerging evidence indicates a novel mode of epidermal growth factor receptor (EGFR) signaling, notably, one involves EGFR nuclear translocalization and subsequent gene activation. To date, however, the significance of the nuclear EGFR pathway in glioblastoma (GBM) is unknown. Here, we report that EGFR and its constitutively activated variant EGFRvIII undergo nuclear translocalization in GBM cells, in which the former event requires EGF stimulation and the latter is constitutive. To gain insights into the effect of nuclear EGFR on gene expression in GBM, we created isogenic GBM cell lines, namely, U87MG-vector, U87MG-EGFR, and U87MG-EGFRdNLS that, respectively, express the control vector, EGFR, and nuclear entry–defective EGFR with a deletion of the nuclear localization signal (NLS). Microarray analysis shows that 19 genes, including cyclooxygenase-2 (COX-2), to be activated in U87MG-EGFR cells but not in U87MG-EGFRdNLS and U87MG-vector cells. Subsequent validation studies indicate that COX-2 gene is expressed at higher levels in cells with EGFR and EGFRvIII than those with EGFRdNLS and EGFRvIIIdNLS. Nuclear EGFR and its transcriptional cofactor signal transducer and activator of transcription 3 (STAT3) associate with the COX-2 promoter. Increased expression of EGFR/EGFRvIII and activated STAT3 leads to the synergistic activation of the COX-2 promoter. Promoter mutational analysis identified a proximal STAT3-binding site that is required for EGFR/EGFRvIII-STAT3–mediated COX-2 gene activation. In GBM tumors, an association exists between levels of COX-2, EGFR/EGFRvIII, and activated STAT3. Together, these findings indicate the existence of the nuclear EGFR/EGFRvIII signaling pathway in GBM and its functional interaction with STAT3 to activate COX-2 gene expression, thus linking EGFR-STAT3 and EGFRvIII-STAT3 signaling axes to proinflammatory COX-2 mediated pathway. Mol Cancer Res; 8(2); 232–45
Elevated Phosphate Activates N-ras and Promotes Cell Transformation and Skin Tumorigenesis
Authors: Camalier Corinne E. , Young Matthew R. , Bobe Gerd , Perella Christine M. , Colburn Nancy H. , Beck George R.
Published: February 2010

Recent results suggest a paradigm shift from viewing inorganic phosphate as a passive requirement for basic cell functions to an active regulator of cell behavior. We have previously shown that elevated concentrations of phosphate increased cell proliferation and expression of protumorigenic genes such as Fra-1 and osteopontin in a preosteoblast cell line. Therefore, we hypothesized that elevated phosphate concentrations would promote cell transformation in vitro and tumorigenesis in vivo. Supplementation of medium with phosphate increased anchorage-independent transformation and proliferation of BALB/c mouse JB6 epidermal cells, activation of N-ras, ERK1/2, and activator protein-1, and increased gene expression of Fra-1, COX-2, and osteopontin in a dose-dependent manner. These in vitro results led to the hypothesis that varying the levels of dietary inorganic phosphate would alter tumorigenesis in the mouse model of skin carcinogenesis. Female FVB/N mice were treated with 7,12-dimethylbenz(a)anthracene/12-O-tetradecanoylphorbol-13-acetate and fed high- or low-phosphate diets (1.2% versus 0.2% of the diet) for 19 weeks. The high-phosphate diet increased skin papilloma number by ∼50% without changing feed intake and body weights. High dietary phosphate increased serum concentrations of phosphate, parathyroid hormone, and osteopontin and decreased serum concentrations of calcium. Thus, we conclude that elevated phosphate promotes cell transformation and skin tumorigenesis partly by increasing the availability of phosphate for activation of N-ras and its downstream targets, which defines reducing dietary phosphate as a novel target for chemoprevention. Cancer Prev Res; 3(3); 359–70
Parity-Induced Decrease in Systemic Growth Hormone Alters Mammary Gland Signaling: A Potential Role in Pregnancy Protection from Breast Cancer
Authors: Dearth Robert K. , Delgado David A. , Hiney Jill K. , Pathiraja Thushangi , Oesterreich Steffi , Medina Dan , Dees W. Les , Lee Adrian V.
Published: February 2010

Early full-term pregnancy is an effective natural protection against breast cancer in both humans and experimental rodents. The protective effect of an early pregnancy is, in part, linked to changes in circulating hormones that are involved in both normal breast development and breast cancer. For example, a reduction in circulating growth hormone (GH) has been shown to protect rats from carcinogen-induced mammary tumors. We examined the ability of a full-term pregnancy to alter the endocrine GH/insulin-like growth factor-I (IGF-I) axis and how this change affected normal mammary gland function in two commonly used rat models (Sprague-Dawley and Wistar Furth). Circulating GH and IGF-I were measured in blood drawn every 30 minutes from parous and age-matched virgin female rats. Mean serum GH levels were significantly decreased (P < 0.01) in parous compared with age-matched virgin rats for both strains. Changes in GH levels were independent of estrous cycle, indicated by a significant (P < 0.05) reduction in circulating levels of GH during estrus and diestrus in both parous strains. Despite the decrease in circulating GH, pituitary GH mRNA levels were unaltered in parous rats. Circulating IGF-I and hepatic IGF-I mRNA were also unaltered by parity in either rat strain. Immunoblot analysis of mammary glands showed decreases in phosphorylation of signal transducer and activator of transcription 5A and Janus-activated kinase 2, suggesting reduced action of GH in the mammary gland. Therefore, although the parity reduction in circulating GH does not affect circulating IGF-I levels, it is possible that reduced GH acts directly at the mammary gland and may play a role in pregnancy protection from breast cancer. Cancer Prev Res; 3(3); 312–21
Null Activity of Selenium and Vitamin E as Cancer Chemopreventive Agents in the Rat Prostate
Authors: McCormick David L. , Rao K.V.N. , Johnson William D. , Bosland Maarten C. , Lubet Ronald A. , Steele Vernon E.
Published: February 2010

To evaluate the potential efficacy of selenium and vitamin E as inhibitors of prostate carcinogenesis, four chemoprevention studies using a common protocol were done in a rat model of androgen-dependent prostate cancer. After stimulation of prostate epithelial cell proliferation by a sequential regimen of cyproterone acetate followed by testosterone propionate, male Wistar-Unilever rats received a single i.v. injection of N-methyl-N-nitrosourea (MNU) followed by chronic androgen stimulation via subcutaneous implantation of testosterone pellets. At 1 week post-MNU, groups of carcinogen-treated rats (39-44/group) were fed either a basal diet or a basal diet supplemented with l-selenomethionine (3 or 1.5 mg/kg diet; study 1), dl-α-tocopherol (vitamin E, 4,000 or 2,000 mg/kg diet; study 2), l-selenomethionine + vitamin E (3 + 2,000 mg/kg diet or 3 + 500 mg/kg diet; study 3), or selenized yeast (target selenium levels of 9 or 3 mg/kg diet; study 4). Each chemoprevention study was terminated at 13 months post-MNU, and prostate cancer incidence was determined by histopathologic evaluation. No statistically significant reductions in prostate cancer incidence were identified in any group receiving dietary supplementation with selenium and/or vitamin E. These data do not support the hypotheses that selenium and vitamin E are potent cancer chemopreventive agents in the prostate, and when considered with the recent clinical data reported in the Selenium and Vitamin E Cancer Prevention Trial (SELECT), show the predictive nature of this animal model for human prostate cancer chemoprevention. Cancer Prev Res; 3(3); 381–92
Traumatic Brain Injury May Increase the Risk for Frontotemporal Dementia through Reduced Progranulin
Authors: Jawaid Ali , Rademakers Rosa , Kass Joseph S. , Kalkonde Yogeshwar , Schulz Paul E.
Published: February 2010

Frontotemporal lobar degeneration with TAR-DNA-binding protein inclusions (FTLD-TDP) is the most common pathological subtype of frontotemporal dementia (FTD). Mutations leading to a loss of function in the progranulin gene (PGRN) are the most common known cause of FTLD-TDP. In agreement with the proposed loss of function disease mechanism, several groups have reported decreased plasma levels of PGRN in patients carrying PGRN mutations compared to individuals without PGRN mutations. We propose that traumatic brain injury (TBI), an environmental factor, may also increase the risk of FTD by altering PGRN metabolism. TBI may lead to an increase in the central nervous system levels of microglial elastases, which proteolyze PGRN into proinflammatory products called granulins causing a reduction in PGRN levels. Hence, inhibiting microglial activation may have an important implication for the prevention of FTD in patients with TBI.
A Computer Model of Dementia Prevalence in Australia: Foreseeing Outcomes of Delaying Dementia Onset, Slowing Disease Progression, and Eradicating Dementia Types
Authors: Vickland Victor , McDonnell Geoff , Werner Joel , Draper Brian , Low Lee-Fay , Brodaty Henry
Published: February 2010

Background: A computer model was designed to test hypothetical scenarios regarding dementia prevalence in Australia (2001–2040). Methods: The study implemented 3 scenarios: delaying dementia onset, slowing disease progression and, in a previously unpublished experiment, eradicating dementia types. Sensitivity analysis and parameter variation were the main methods of experimentation. Results: The model predicts that delaying dementia onset by 5 years will reduce the 2040 prevalence by 37%. An onset delay of 2 years, introduced in 2010, will reduce the 2040 prevalence by 16%. Slowing disease progression increases the 2040 prevalence by 4–7%. Total eradication of Alzheimer’s disease (currently approximately 50% of all dementia cases) in 2020 will decrease the 2040 prevalence by 42%. Conclusion: Computer modeling of future scenarios and interventions helps health and aged care planners understand the likely challenges society will face with the ageing of the world’s population.
Transpontine Connection Fibers between Corticospinal Tracts in Hemiparetic Patients with Intracerebral Hemorrhage
Authors: Yeo Sang Seok , Choi Byung Yeon , Chang Chul Hoon , Jang Sung Ho
Published: February 2010

Objectives: A connection of fibers between corticospinal tracts (CSTs) at the pons, originating from the CST of the affected hemisphere, has been observed in hemiparetic patients with stroke. The authors investigated the incidence and the clinical significance of transpontine connection of fibers (TCFs) in hemiparetic patients with intracerebral hemorrhage (ICH), using diffusion tensor tractography (DTT). Subjects and Methods: Forty-two patients with ICH with weakness of the affected extremities at the time of DTI scanning and 41 age-matched control subjects were recruited. TCFs were classified into three types according to severity: type A – no TCF extending to the opposite hemisphere, type B – a TCF crossing to the opposite hemisphere and ending at the subcortical level, and type C – a TCF crossing the pons and ascending to the cortex of the opposite hemisphere. Results: TCFs originating from the CST in affected and unaffected hemispheres were significantly more prevalent among patients than controls (both p < 0.05). In addition, TCF severity was found to be closely related to motor function reduction in affected extremities (p < 0.05) and to extent of CST injury in affected hemispheres (p < 0.05). Conclusions: TCF appears to represent a compensatory mechanism associated with motor weakness or CST injury in patients with ICH.
Prostaglandins and Radical Oxygen Species Are Involved in Microvascular Effects of Hyperoxia
Authors: Rousseau A. , Tesselaar E. , Henricson J. , Sjöberg F.
Published: February 2010

Hyperoxia causes vasoconstriction in most tissues, by mechanisms that are not fully understood. We investigated microvascular effects of breathing 100% oxygen in healthy volunteers, using iontophoresis to deliver acetylcholine (ACh) and sodium nitroprusside (SNP). Aspirin and vitamin C were used to test for involvement of prostaglandins and radical oxygen species. Forearm skin perfusion was measured using laser Doppler perfusion imaging. Results were analysed using dose-response modelling. The response to ACh was reduced by 30% during oxygen breathing compared to air breathing [0.98 (0.81–1.15) PU vs. 1.45 (1.30–1.60) PU, p < 0.001]. ED50 values were unchanged [2.25 (1.84–2.75) vs. 2.21 (1.79–2.74), not significant]. Aspirin pre-treatment abolished the difference in response between oxygen breathing and air breathing [maximum: 1.03 (0.90–1.16) vs. 0.89 (0.77–1.01), not significant; ED50: 1.83 (1.46–2.30) vs. 1.95 (1.65–2.30), not significant]. ACh-mediated vasodilatation during 100% oxygen breathing was partially restored after pre-treatment with vitamin C. Breathing 100% oxygen did not change the microvascular response to SNP [1.45 (1.28–1.62) vs. 1.40 (1.26–1.53), not significant]. These results favour the hypothesis that hyperoxic vasoconstriction is mediated by inhibition of prostaglandin synthesis. Radical oxygen species may be involved as vitamin C, independently of aspirin, partially restored ACh-mediated vasodilatation during hyperoxia.
Molecular Cloning and Immunochemical Characterization of a Novel Major Japanese Cedar Pollen Allergen Belonging to the Aspartic Protease Family
Authors: Ibrahim Ahmed Ragaa Nour , Kawamoto Seiji , Aki Tsunehiro , Shimada Yayoi , Rikimaru Satoshi , Onishi Nobukazu , Babiker Elfadil Elfadl , Oiso Isao , Hashimoto Kunihiko , Hayashi Takaharu , Ono Kazuhisa
Published: February 2010

Background:Japanese cedar (Cryptomeria japonica) pollen is a major cause of seasonal pollinosis in Japan. Protease activity in the pollen grains may trigger pro-allergic responses but no such proteases have yet been identified as pollen allergens. Objectives:We report the molecular cloning and immunochemical characterization of a novel C. japonica pollen allergen belonging to the aspartic protease family. Methods:We focused on the C. japonica pollen allergen spot No. 63 (CPA63, 47.5% IgE binding frequency) on our 2-dimensional IgE immunoblot map. The internal amino acid sequences were determined using time-of-flight mass spectrometry. Full-length cpa63 cDNA was cloned by rapid amplification of cDNA ends (RACE)-PCR. Recombinant CPA63 (r-CPA63) was expressed using the baculovirus-insect cell culture system and its IgE binding capacity was analyzed by enzyme-linked immunosorbent assay (ELISA). The proteolytic activity of r-CPA63 was also assessed using a putative mature enzyme produced upon autolysis. Results: cpa63 cDNA encoded a 472 amino acid polypeptide showing about 40% sequence identity to members of the plant atypical aspartic protease family. ELISA showed that r-CPA63 was recognized by IgE antibodies in the serum of 58% (18/31) of Japanese cedar pollinosis patients. We also demonstrated an aspartic protease-like enzyme activity of the putative mature r-CPA63. Conclusions:We have identified the first plant aspartic protease allergen from Japanese cedar pollen. The availability of the CPA63 sequence and its recombinant allergen production system are useful not only for pharmaceutical applications but also for further examination of the role of protease activity in the pathogenesis of cedar pollinosis.