bluuebunny/crossref_metadata_2025 huggingface dataset:
Proinflammation and Hypertension: A Population‐Based Study
Authors: Mauno Vanhala , Hannu Kautiainen , Esko Kumpusalo
Published: December 2008

There is evidence that proinflammation may be linked to the development of hypertension (HT). We examined the association of both the interleukin‐1 beta (IL‐1β) and the interleukin 1‐receptor antagonist (IL‐1ra) with future blood pressure (BP) and HT occurrence (BP ≥ 140/90 mmHg, or antihypertensive drug) in a population‐based prospective study. Our study consisted of 396 (147 men and 249 women) middle‐aged, baseline apparently healthy, normotensive subjects participating in a 6.5‐year follow‐up study. Subjects with high‐sensitivity CRP (hs‐CRP) < 10 mg/L were excluded at the initial visit. At follow‐up, the occurrence of HT was 32%. The levels of baseline IL‐1β and IL‐1ra were significantly higher for subjects who developed HT during the follow‐up than for those who did not (IL‐1β; 0.67 ± 0.62 pg/mL versus 0.56 ± 0.32 pg/mL, P = .020 and IL‐1ra; 184 ± 132 pg/mL versus 154 ± 89 pg/mL, P = .007). After adjustments for age, follow‐up time, sex, baseline systolic BP, and BMI, our results confirm a statistically significant (P = .036) linear association between the quartiles of IL‐1β and change of systolic BP during the study. After adjustments for age, follow‐up time, sex, and BMI, our results also show a linear association between incident HT and the quartiles of IL‐1ra. (P = .026). These results provide evidence that proinflammation may precede BP elevation and HT.
Vapor Sensing Using Conjugated Molecule‐Linked Au Nanoparticles in a Silica Matrix
Authors: Dirk Shawn M. , Howell Stephen W. , Price B. Katherine , Fan Hongyou , Washburn Cody , Wheeler David R. , Tour James M. , Whiting Joshua , Simonson R. Joseph
Published: December 2008

Cross‐linked assemblies of nanoparticles are of great value as chemiresistor‐type sensors. Herein, we report a simple method to fabricate a chemiresistor‐type sensor that minimizes the swelling transduction mechanism while optimizing the change in dielectric response. Sensors prepared with this methodology showed enhanced chemoselectivity for phosphonates which are useful surrogates for chemical weapons. Chemoselective sensors were fabricated using an aqueous solution of gold nanoparticles that were then cross‐linked in the presence of the silica precursor, tetraethyl orthosilicate with the α‐,
ω‐dithiolate (which is derived from the in situ deprotection of
1,4‐di(Phenylethynyl‐4′,4″‐diacetylthio)‐benzene (1)
with wet triethylamine). The cross‐linked nanoparticles and silica matrix were drop coated onto interdigitated electrodes having 8 μm spacing. Samples were exposed to a series of analytes including dimethyl methylphosphonate (DMMP), octane, and toluene. A limit of detection was obtained for each analyte. Sensors assembled in this fashion were more sensitive to dimethyl methylphosphonate than to octane by a factor of 1000.
A Feed-Forward Loop Involving Protein Kinase Cα and MicroRNAs Regulates Tumor Cell Cycle
Authors: Cohen Ezra E.W. , Zhu Hongyan , Lingen Mark W. , Martin Leslie E. , Kuo Wen-Liang , Choi Eugene A. , Kocherginsky Masha , Parker Joel S. , Chung Christine H. , Rosner Marsha Rich
Published: December 2008

Protein kinase Cα (PKCα) has been implicated in cancer, but the mechanism is largely unknown. Here, we show that PKCα promotes head and neck squamous cell carcinoma (SCCHN) by a feed-forward network leading to cell cycle deregulation. PKCα inhibitors decrease proliferation in SCCHN cell lines and xenografted tumors. PKCα inhibition or depletion in tumor cells decreases DNA synthesis by suppressing extracellular signal-regulated kinase phosphorylation and cyclin E synthesis. Additionally, PKCα down-regulates miR-15a, a microRNA that directly inhibits protein synthesis of cyclin E, as well as other cell cycle regulators. Furthermore, both PKCα and cyclin E protein expression are increased in primary tumors, and PKCα inversely correlates with miR-15a expression in primary tumors. Finally, PKCα is associated with poor prognosis in SCCHN. These results identify PKCα as a key regulator of SCCHN tumor cell growth by a mechanism involving activation of mitogen-activated protein kinase, an initiator of the cell cycle, and suppression of miR-15a, an inhibitor of DNA synthesis. Although the specific components may be different, this type of feed-forward loop network, consisting of a stimulus that activates a positive signal and removes a negative brake, is likely to be a general one that enables induction of DNA synthesis by a variety of growth or oncogenic stimuli. [Cancer Res 2009;69(1):65–74]
Interleukin-6 in the Bone Marrow Microenvironment Promotes the Growth and Survival of Neuroblastoma Cells
Authors: Ara Tasnim , Song Liping , Shimada Hiroyuki , Keshelava Nino , Russell Heidi V. , Metelitsa Leonid S. , Groshen Susan G. , Seeger Robert C. , DeClerck Yves A.
Published: December 2008

Neuroblastoma, the second most common solid tumor in children, frequently metastasizes to the bone marrow and the bone. Neuroblastoma cells present in the bone marrow stimulate the expression of interleukin-6 (IL-6) by bone marrow stromal cells (BMSC) to activate osteoclasts. Here we have examined whether stromal-derived IL-6 also has a paracrine effect on neuroblastoma cells. An analysis of the expression of IL-6 and its receptor, IL-6R, in 11 neuroblastoma cell lines indicated the expression of IL-6 in 4 cell lines and of IL-6R in 9 cell lines. Treatment of IL-6R–positive cells with recombinant human IL-6 resulted in signal transducer and activator of transcription-3 and extracellular signal–regulated kinase-1/2 activation. Culturing IL-6R–positive neuroblastoma cells in the presence of BMSC or recombinant human IL-6 increased proliferation and protected tumor cells from etoposide-induced apoptosis, whereas it had no effect on IL-6R–negative tumor cells. In vivo, neuroblastoma tumors grew faster in the presence of a paracrine source of IL-6. IL-6 induced the expression of cyclooxygenase-2 in neuroblastoma cells with concomitant release of prostaglandin-E2, which increased the expression of IL-6 by BMSC. Supporting a role for stromal-derived IL-6 in patients with neuroblastoma bone metastasis, we observed elevated levels of IL-6 in the serum and bone marrow of 16 patients with neuroblastoma bone metastasis and in BMSC derived from these patients. Altogether, the data indicate that stromal-derived IL-6 contributes to the formation of a bone marrow microenvironment favorable to the progression of metastatic neuroblastoma. [Cancer Res 2009;69(1):329–37]
Macrophage Inhibitory Cytokine-1: Possible Bridge Molecule of Inflammation and Prostate Cancer
Authors: Karan Dev , Holzbeierlein Jeffrey , Thrasher J. Brantley
Published: December 2008

There is emerging evidence that inflammation may lead to prostate cancer development. Although inflammation is an essential response to injury or infection, chronic inflammation is harmful and causes tissue damage. Increasing evidence suggests that inflammation leads to the development of epithelial cancers; however, studies on inflammation-targeted genes that might contribute to the development of cancer are at the beginning stage. Here, we describe macrophage inhibitory cytokine-1, which provides a potential link between inflammation and prostate cancer. Understanding the regulation of macrophage inhibitory cytokine-1 in response to inflammation may have potential for novel therapeutic strategies. [Cancer Res 2009;69(1):2–5]
Indibulin, a Novel Microtubule Inhibitor, Discriminates between Mature Neuronal and Nonneuronal Tubulin
Authors: Wienecke Anke , Bacher Gerald
Published: December 2008

Microtubule inhibitors interfere with microtubule dynamics, causing cell cycle arrest and apoptosis. These effects are responsible for the chemotherapeutic activities of members of the taxane and Vinca alkaloid families in oncology. Unfortunately, a major side effect of the taxanes and Vinca alkaloids is the development of peripheral neuropathies. Indibulin (N-[pyridin-4-yl]-[1-(4-chlorbenzyl)-indol-3-yl]-glyoxyl-amid; D-24851; ZIO-301), a novel synthetic small molecule microtubule inhibitor, destabilizes microtubules and has antitumor activity but does not exhibit neurotoxicity in preclinical animal studies. In the present study, it has been found that indibulin is able to discriminate between highly posttranslationally modified tubulin present in mature neuronal microtubules, and less-modified tubulin present in immature neuronal or nonneuronal microtubules. Vincristine and colchicine act on either tubulin equally well. The binding site of indibulin on mature neuronal microtubules seems to be inaccessible due to the posttranslational modifications, a theory that is supported by the observation that indibulin did not disrupt the integrity of highly modified microtubules present in neurites of pheochromocytoma (PC12) cells. The specificity of indibulin for unmodified microtubules seems to be dependent on the pyridyl moiety of indibulin because derivatives that have the pyridyl moiety replaced are not able to discriminate between highly and less-modified tubulins. The observed broad antitumor activity of indibulin and the lack of central and peripheral nervous system toxicity in preclinical studies make it a promising candidate for development as a cancer treatment. Indibulin is currently in phase I clinical trials. [Cancer Res 2009;69(1):171–7]
Epigenetic Profiles Distinguish Pleural Mesothelioma from Normal Pleura and Predict Lung Asbestos Burden and Clinical Outcome
Authors: Christensen Brock C. , Houseman E.A. , Godleski John J. , Marsit Carmen J. , Longacker Jennifer L. , Roelofs Cora R. , Karagas Margaret R. , Wrensch Margaret R. , Yeh Ru-Fang , Nelson Heather H. , Wiemels Joe L. , Zheng Shichun , Wiencke John K. , Bueno Raphael , Sugarbaker David J. , Kelsey Karl T.
Published: December 2008

Mechanisms of action of nonmutagenic carcinogens such as asbestos remain poorly characterized. As pleural mesothelioma is known to have limited numbers of genetic mutations, we aimed to characterize the relationships among gene-locus–specific methylation alterations, disease status, asbestos burden, and survival in this rapidly fatal asbestos-associated tumor. Methylation of 1505 CpG loci associated with 803 cancer-related genes were studied in 158 pleural mesotheliomas and 18 normal pleura. After false-discovery rate correction, 969 CpG loci were independently associated with disease status (Q < 0.05). Classifying samples based on CpG methylation profile with a mixture model approach, methylation classes discriminated tumor from normal pleura (permutation P < 0.0001). In a random forests classification, the overall misclassification error rate was 3.4%, with <1% (n = 1) of tumors misclassified as normal (P < 0.0001). Among tumors, methylation class membership was significantly associated with lung tissue asbestos body burden (P < 0.03), and significantly predicted survival (likelihood ratio P < 0.01). Consistent with prior work, asbestos burden was associated with an increased risk of death (hazard ratio, 1.4; 95% confidence interval, 1.1–1.8). Our results have shown that methylation profiles powerfully differentiate diseased pleura from nontumor pleura and that asbestos burden and methylation profiles are independent predictors of mesothelioma patient survival. We have added to the growing body of evidence that cellular epigenetic dysregulation is a critical mode of action for asbestos in the induction of pleural mesothelioma. Importantly, these findings hold great promise for using epigenetic profiling in the diagnosis and prognosis of human cancers. [Cancer Res 2009;69(1):227–34]
K-ras and Wnt Signaling Synergize to Accelerate Prostate Tumorigenesis in the Mouse
Authors: Pearson Helen B. , Phesse Toby J. , Clarke Alan R.
Published: December 2008

Aberrant Ras and Wnt signaling are emerging as key events in the multistep nature of prostate tumorigenesis and progression. Here, we report the generation of a compound model of prostate cancer to define the synergism of activated K-ras (K-ras+/V12) and dominant stabilized β-catenin (Catnb+/lox(ex3)) in the murine prostate. Recombination of floxed alleles and subsequent expression of oncogenic transgenes was mediated by Cre recombinase expression governed by the composite Probasin (PB) promoter (termed PBCre). Concomitant with elevated mitogen-activated protein kinase (MAPK) signaling, PBCre+K-ras+/V12 mice developed AH at 100 days (100% incidence) and low-grade prostate intraepithelial neoplasia and adenocarcinoma (60% and 7% incidence) by 500 days. PBCre+Catnb+/lox(ex3) mice showed reduced longevity (average 428 days) and were predisposed to PIN-like keratinized squamous metaplasia at 100 days (100% incidence) and adenocarcinoma (100% incidence) at end-point. These lesions displayed elevated Wnt signaling and basal levels of MAPK signaling. Synchronous activation of K-ras and β-catenin significantly reduced survival (average 189 days), reflecting accelerated tumorigenesis and the development of invasive carcinoma that displayed activated Wnt and MAPK signaling. Notably, expression of the basal cell marker p63 negatively correlated with tumor grade, resembling human prostate adenocarcinoma. Taken together, our data show that combinatorial oncogenic mutations of K-ras and β-catenin drive rapid progression of prostate tumorigenesis to invasive carcinoma, characterized by the synergistic elevation of androgen receptor, cyclooxygenase-2, and c-Myc. [Cancer Res 2009;69(1):94–101]
Genetic Regulation of the Response to Her-2 DNA Vaccination in Human Her-2 Transgenic Mice
Authors: Radkevich-Brown Olga , Jacob Jennifer , Kershaw Michael , Wei Wei-Zen
Published: December 2008

Genetic regulation of immune reactivity to Her-2 vaccination and the consequent antitumor effect was tested in human Her-2 transgenic (Tg) mice of C57BL/6 (B6), BALB/c (BALB), and (B6x BALB) F1 (F1) background. Mice were electrovaccinated with Her-2 DNA with or without pretreatment with CD25 monoclonal antibody to remove CD25hi regulatory T cells. When CD25+ T cells were intact, BALB Her-2 Tg mice were more responsive than the other two strains in both humoral and cellular immunities, and their tumor growth was significantly delayed. B6 Her-2 Tg mice responded poorly and F1 mice showed modest immune reactivity, but tumor growth did not change in either strain. Depletion of CD25hi T cells before vaccination significantly improved protection from tumor challenge in F1 Her-2 Tg mice. This was associated with elevated levels of Her-2 IgG1, IgG2a, and IgG2c antibodies, and some mice also showed IFN-γ producing T-cell response. The same treatment induced modest improvement in B6 Her-2 Tg mice. In BALB Her-2 Tg mice, however, depletion of CD25hi T cells did not further improve antitumor efficacy. Although their Her-2–specific IgG1 and interleukin-5–secreting T cells increased, the levels of IgG2a and IFN-γ–secreting T cells did not change. These results are the first to show genetic regulation of the response to a cancer vaccine and an unequal effect of removing CD25hi T cells on antitumor immunity. These results warrant individualized treatment plans for patients with heterogeneous genetic backgrounds and possibly differential intrinsic immune reactivity to tumor antigens. [Cancer Res 2009;69(1):212–8]
Cyclin-Dependent Kinase-3–Mediated c-Jun Phosphorylation at Ser63 and Ser73 Enhances Cell Transformation
Authors: Cho Yong-Yeon , Tang Faqing , Yao Ke , Lu Chengrong , Zhu Feng , Zheng Duo , Pugliese Angelo , Bode Ann M. , Dong Zigang
Published: December 2008

c-Jun is a component of the activator protein-1 (AP-1) complex, which plays a crucial role in the regulation of gene expression, cell proliferation, and cell transformation, as well as cancer development. Herein, we found that cyclin-dependent kinase (Cdk)-3, but not Cdk2 or c-Jun NH2-terminal kinase, is a novel kinase of c-Jun induced by stimulation with growth factors such as epidermal growth factor (EGF). Cdk3 was shown to phosphorylate c-Jun at Ser63 and Ser73 in vitro and ex vivo. EGF-induced Cdk3 activation caused c-Jun phosphorylation at Ser63 and Ser73, resulting in increased AP-1 transactivation. Ectopic expression of Cdk3 resulted in anchorage-independent cell transformation of JB6 Cl41 cells induced by EGF and foci formation stimulated by constitutively active Ras (RasG12V), which was mediated by AP-1 in NIH3T3 cells. These results showed that the Cdk3/c-Jun signaling axis plays an important role in EGF-stimulated cell proliferation and cell transformation. [Cancer Res 2009;69(1):272–81]
High Frequency of LMAN1 Abnormalities in Colorectal Tumors with Microsatellite Instability
Authors: Roeckel Nina , Woerner Stefan M. , Kloor Matthias , Yuan Yan-Ping , Patsos Georgios , Gromes Roland , Kopitz Juergen , Gebert Johannes
Published: December 2008

Glycosyl epitopes have been identified as tumor-specific markers in colorectal tumors and various lines of evidence indicate the significance of altered synthesis, transport, and secretion of glycoproteins in tumorigenesis. However, aberrant glycosylation has been largely ignored in microsatellite unstable (MSI-H) colorectal tumors. Therefore, we analyzed mutation frequencies of genes of the cellular glycosylation machinery in MSI-H tumors, focusing on frameshift mutations in coding MNRs (cMNRs). Among 28 candidate genes, LMAN1/ERGIC53, a mannose-specific lectin mediating endoplasmatic reticulum (ER)-to-Golgi transit of glycosylated proteins, showed high mutation frequency in MSI-H colorectal cancer cell lines (52%; 12 of 23), carcinomas (45%; 72 of 161), and adenomas (40%; 8 of 20). Biallelic mutations were observed in 17% (4 of 23) of MSI-H colorectal cancer cell lines. LMAN1 was found to be transcribed but truncated protein remained undetectable in these LMAN1-mutant cell lines. Immunohistochemical and molecular analysis of LMAN1-mutated carcinomas and adenomas revealed regional loss of LMAN1 expression due to biallelic LMAN1 cMNR frameshift mutations. In LMAN1-deficient colorectal cancer cell lines, secretion of the LMAN1 client protein α-1-antitrypsin (A1AT), an inhibitor of angiogenesis and tumor growth, was significantly impaired but could be restored upon LMAN1 re-expression. These results suggest that LMAN1 mutational inactivation is a frequent and early event potentially contributing to MSI-H tumorigenesis. [Cancer Res 2009;69(1):292–9]
BRCA1-Associated Protein 1 Interferes with BRCA1/BARD1 RING Heterodimer Activity
Authors: Nishikawa Hiroyuki , Wu Wenwen , Koike Ayaka , Kojima Ryoko , Gomi Hiromichi , Fukuda Mamoru , Ohta Tomohiko
Published: December 2008

The breast and ovarian tumor suppressor BRCA1 constitutes a RING heterodimer E3 ligase with BARD1. BRCA1-associated protein 1 (BAP1) is a ubiquitin COOH-terminal hydrolase that was initially identified as a protein that bound to the RING finger domain of BRCA1. However, how BAP1 contributes to the E3 activity of BRCA1/BARD1 is unclear. Here, we report that BAP1 interacts with BARD1 to inhibit the E3 ligase activity of BRCA1/BARD1. Domains comprised by residues 182-365 of BAP1 interact with the RING finger domain of BARD1, and surface plasmon resonance spectroscopy (BIAcore) analyses showed that BAP1 interferes with the BRCA1/BARD1 association. The perturbation resulted in inhibition of BRCA1 autoubiquitination and NPM1/B23 ubiquitination by BRCA1/BARD1. Although BAP1 was capable of deubiquitinating the polyubiquitin chains mediated by BRCA1/BARD1 in vitro, a catalytically inactive mutant of BAP1, C91S, still inhibited the ubiquitination in vitro and in vivo, implicating a second mechanism of action. Importantly, inhibition of BAP1 expression by short hairpin RNA resulted in hypersensitivity of the cells to ionizing irradiation and in retardation of S-phase progression. Together, these results suggest that BAP1 and BRCA1/BARD1 coordinately regulate ubiquitination during the DNA damage response and the cell cycle. [Cancer Res 2009;69(1):111–9]
GEMS (Gene Expression Metasignatures), a Web Resource for Querying Meta-analysis of Expression Microarray Datasets: 17β-Estradiol in MCF-7 Cells
Authors: Ochsner Scott A. , Steffen David L. , Hilsenbeck Susan G. , Chen Edward S. , Watkins Christopher , McKenna Neil J.
Published: December 2008

With large amounts of public expression microrray data being generated by multiple laboratories, it is a significant task for the bench researcher to routinely identify available datasets, and then to evaluate the collective evidence across these datasets for regulation of a specific gene in a given system. 17β-Estradiol stimulation of MCF-7 cells is a widely used model in the growth of breast cancer. Although myriad independent studies have profiled the global effects of this hormone on gene expression in these cells, disparate experimental variables and the limited power of the individual studies have combined to restrict the agreement between them as to the specific gene expression signature elicited by this hormone. To address these issues, we have developed a freely accessible Web resource, Gene Expression MetaSignatures (GEMS) that provides the user a consensus for each gene in the system. We conducted a weighted meta-analysis encompassing over 13,000 genes across 10 independent published datasets addressing the effect of 17β-estradiol on MCF-7 cells at early (3–4 hours) and late (24 hours) time points. In a literature survey of 58 genes previously shown to be regulated by 17β-estradiol in MCF-7 cells, the meta-analysis combined the statistical power of the underlying datasets to call regulation of these genes with nearly 85% accuracy (false discovery rate–corrected P < 0.05). We anticipate that with future expression microarray dataset contributions from investigators, GEMS will evolve into an important resource for the cancer and nuclear receptor signaling communities. [Cancer Res 2009;69(1):23–6]
The Origins of Breast Cancer Prognostic Gene Expression Profiles
Authors: Lukes Luanne , Crawford Nigel P.S. , Walker Renard , Hunter Kent W.
Published: December 2008

Recent high profile clinical trials show that microarray-based gene expression profiling has the potential to become an important tool for predicting prognosis in breast cancer. Earlier work in our laboratory using mouse models and human breast cancer populations has enabled us to show that metastasis susceptibility is an inherited trait. This same combined approach facilitated the identification of a number of candidate genes that, when dysregulated, have the potential to induce prognostic gene expression profiles in human data sets. To investigate if these gene expression signatures were of somatic or germline origin and to assess the contribution of different cell types to the induction of these signatures, we have performed a series of expression profiling experiments in a mouse model of metastatic breast cancer. These results show that both the tumor epithelium and invading stromal tissues contribute to the development of prognostic gene signatures. Furthermore, analysis of normal tissues and tumor transplants suggests that prognostic signatures result from both somatic and inherited components, with the inherited components being more consistently predictive. [Cancer Res 2009;69(1):310–8]
Phase I and Pharmacokinetic Study of Pemetrexed plus Cisplatin in Chemonaive Patients with Locally Advanced or Metastatic Malignant Pleural Mesothelioma or Non–Small Cell Lung Cancer
Authors: Dickgreber Nicolas J. , Fink Thomas H. , Latz Jane E. , Hossain Anwar M. , Musib Luna C. , Thomas Michael
Published: December 2008

Purpose: Pemetrexed is approved as monotherapy and in combination with cisplatin. The established combination dose was identified before the addition of folic acid and vitamin B12 to the treatment regimen. We evaluated the toxicity and pharmacokinetics (PK) of higher pemetrexed doses with cisplatin and vitamin supplementation.

Experimental Design: Patients with malignant pleural mesothelioma or non–small cell lung cancer received pemetrexed doses from 500 to 900 mg/m2 + 75 mg/m2 cisplatin once every 21 days. Folic acid and vitamin B12 were administered per label recommendations.

Results: Twenty-one patients received a combined total of 84 cycles. The maximum tolerated dose was 900 mg/m2 pemetrexed + 75 mg/m2 cisplatin. Dose-limiting toxicities at this dose included grade 3 anemia, bronchopneumonia, and neutropenia, and 1 death from sepsis secondary to grade 4 febrile neutropenia, considered possibly related to study drugs. The recommended dose was 800 mg/m2 pemetrexed + 75 mg/m2 cisplatin. Pemetrexed PK were consistent across doses; pemetrexed did not seem to affect total or free platinum PK.

Conclusions: Pemetrexed with vitamin supplementation was safe and well tolerated at higher doses than the currently established 500 mg/m2 + 75 mg/m2 cisplatin. Based on this study, the recommended dose would be 800 mg/m2 pemetrexed + 75 mg/m2 cisplatin. However, recent studies showed a lack of improved efficacy for 900 or 1,000 mg/m2 single-agent pemetrexed versus 500 mg/m2 and a lack of PK/pharmacodynamic exposure-response relationship for the pemetrexed/cisplatin combination across pemetrexed exposures corresponding to this dose range. Based on currently available evidence, we recommend retaining the established dose.
rNAPc2 Inhibits Colorectal Cancer in Mice through Tissue Factor
Authors: Zhao Jingsong , Aguilar Gerard , Palencia Servando , Newton Elizabeth , Abo Arie
Published: December 2008

Purpose: Recombinant nematode anticoagulant protein c2 (rNAPc2) is a specific inhibitor of tissue factor (TF)/factor VIIa complex with novel antithrombotic activity. TF is highly expressed in human colorectal tumors, and levels are positively correlated with disease progression.

Experimental Design: To explore the therapeutic potential and mechanism of action of rNAPc2 during tumor growth and metastasis, we tested rNAPc2 in several experimental colorectal cancer models in mice.

Results: Administration of rNAPc2 inhibited pulmonary metastasis in mice systemically disseminated with CT26 murine colon carcinoma cells in a dose-dependent fashion. Combining rNAPc2 with the cytotoxic agent 5-fluorouracil or bevacizumab (humanized anti-vascular endothelial growth factor monoclonal antibody) resulted in additive growth inhibition and simultaneous reduction of microvessel density in HCT116 human colorectal tumor xenografts in nude mice. Furthermore, rNAPc2 potentiated CPT-11 in inhibiting hepatic metastasis in nude mice with portal vein injection of HCT116 tumor cells. Long-term administration of rNAPc2 significantly suppressed spontaneous formation of intestinal tumors in ApcMin/+ mice. Using a RNA interference approach, we showed that TF expression is necessary for rNAPc2-mediated inhibition of HCT116 human colorectal tumor xenograft growth in nude mice, indicating that the antitumor effect of rNAPc2 may be transduced through TF that is expressed on tumor cells.

Conclusions: rNAPc2 is a potent anticancer agent when used in combination with chemotherapy or antiangiogenic therapy in mouse models of colorectal cancer, and TF positivity appears to be required for its activity.
Histone Deacetylase 8 in Neuroblastoma Tumorigenesis
Authors: Oehme Ina , Deubzer Hedwig E. , Wegener Dennis , Pickert Diana , Linke Jan-Peter , Hero Barbara , Kopp-Schneider Annette , Westermann Frank , Ulrich Scott M. , von Deimling Andreas , Fischer Matthias , Witt Olaf
Published: December 2008

Purpose: The effects of pan–histone deacetylase (HDAC) inhibitors on cancer cells have shown that HDACs are involved in fundamental tumor biological processes such as cell cycle control, differentiation, and apoptosis. However, because of the unselective nature of these compounds, little is known about the contribution of individual HDAC family members to tumorigenesis and progression. The purpose of this study was to evaluate the role of individual HDACs in neuroblastoma tumorigenesis.

Experimental Design: We have investigated the mRNA expression of all HDAC1-11 family members in a large cohort of primary neuroblastoma samples covering the full spectrum of the disease. HDACs associated with disease stage and survival were subsequently functionally evaluated in cell culture models.

Results: Only HDAC8 expression was significantly correlated with advanced disease and metastasis and down-regulated in stage 4S neuroblastoma associated with spontaneous regression. High HDAC8 expression was associated with poor prognostic markers and poor overall and event-free survival. The knockdown of HDAC8 resulted in the inhibition of proliferation, reduced clonogenic growth, cell cycle arrest, and differentiation in cultured neuroblastoma cells. The treatment of neuroblastoma cell lines as well as short-term-culture neuroblastoma cells with an HDAC8-selective small-molecule inhibitor inhibited cell proliferation and clone formation, induced differentiation, and thus reproduced the HDAC8 knockdown phenotype. Global histone 4 acetylation was not affected by HDAC8 knockdown or by selective inhibitor treatment.

Conclusions: Our data point toward an important role of HDAC8 in neuroblastoma pathogenesis and identify this HDAC family member as a specific drug target for the differentiation therapy of neuroblastoma.
Cyclooxygenase-2 Is an Independent Prognostic Factor in Gastric Carcinoma Patients Receiving Adjuvant Chemotherapy and Is Not Associated with EBV Infection
Authors: Park Eun Soo , Do In Gu , Park Cheol Keun , Kang Won Ki , Noh Jae Hyung , Sohn Tae Sung , Kim Sung , Kim Min-Ji , Kim Kyoung-Mee
Published: December 2008

Purpose: Cyclooxygenase-2 (COX-2) is believed to be involved in carcinogenesis in patients with chronic gastritis with Helicobacter pylori infection. EBV is detected in ∼10% of gastric carcinomas and H. pylori induces EBV reactivation in the gastric epithelium. We aimed to evaluate significance of COX-2 in gastric carcinoma occurred in EBV and H. pylori prevalent area.

Experimental Design: Tissue microarray samples from 457 gastric carcinoma patients who underwent gastrectomy and adjuvant chemotherapy were studied with EBER1 in situ hybridization for EBV and immunohistochemistry for COX-2 and other gastric carcinoma-related proteins (hMLH1, E-cadherin, c-erbB, and cyclin D1).

Results: EBV infection was observed in 10.9% of gastric carcinomas and was associated with proximal tumor location, increased numbers of lymph node, and E-cadherin expression (P < 0.01). COX-2 overexpression was closely associated with intestinal histologic type and lower tumor stage (P = 0.01). Univariate analysis showed that pT, pN, lymph node ratio, American Joint Committee on Cancer stage, numbers of negative lymph nodes, and resection margin <1 cm were significant prognostic factors. The Cox proportional hazards regression analysis indicated that lack of COX-2 expression and resection margin <1 cm were independent prognostic factors for disease-free survival (P = 0.008 and 0.03, respectively) and overall survival (P = 0.01 and 0.007, respectively).

Conclusions: EBV infection is not associated with COX-2 expression or survival in gastric carcinoma. Lack of COX-2 expression is an independent prognostic factor in both overall and disease-free survival in gastric carcinoma. Our results indicate that COX-2 may play a role in the progression of gastric carcinoma regardless of EBV infection and is closely associated with histologic differentiation and prognosis.
RRR-α-Vitamin E Succinate Potentiates the Antitumor Effect of Calcitriol in Prostate Cancer without Overt Side Effects
Authors: Yin Yi , Ni Jing , Chen Ming , Guo Yinglu , Yeh Shuyuan
Published: December 2008

Purpose: To determine the antitumor efficacy of using calcitriol combined with RRR-α-vitamin E succinate (VES) on prostate cancer.

Experimental Design: The effects of VES or VES in combination with calcitriol on the calcitriol target genes were evaluated by Western blot and real-time PCR. The antiproliferation effect of the combination in prostate cancer cells was evaluated by the combination index method. The role of the vitamin D3 receptor (VDR) in the enhanced antitumor effects of the combination was confirmed by small interfering RNA knockdown strategy. Xenograft-bearing mice were used to reaffirm the antitumor efficacy of this combination. Pathohistology analyses and expressions of VDR and its target genes were analyzed in untreated and treated tumors.

Results: VES selectively increased VDR protein in different prostate cancer cells. Low doses of calcitriol combined with VES were significantly superior to the additive effect of individual treatments against prostate cancer cell proliferation. The expression of VDR target genes involved in antiproliferation were further sensitized in the presence of VES. Knockdown of VDR expression abolished the combination benefits in LNCaP and PC3 cells. Consistently, in prostate cancer xenograft models, VES enhanced the therapeutic efficacy of a tolerated dose of calcitriol yet without overt evidence of systemic toxicity and hypercalcemia. This notable in vivo effect was also accompanied by up-regulation of VDR target genes.

Conclusions: Low-dose calcitriol combined with vitamin E analogue could be a solution to the calcemic side effect. The demonstration of superior antitumor activity of low-dose calcitriol plus VES provides the preclinical basis for developing a useful therapeutic strategy for prostate cancer.
S100A2 Induces Metastasis in Non–Small Cell Lung Cancer
Authors: Bulk Etmar , Sargin Bülent , Krug Utz , Hascher Antje , Jun Yu , Knop Markus , Kerkhoff Claus , Gerke Volker , Liersch Ruediger , Mesters Rolf M. , Hotfilder Marc , Marra Alessandro , Koschmieder Steffen , Dugas Martin , Berdel Wolfgang E. , Serve Hubert , Müller-Tidow Carsten
Published: December 2008

Purpose: S100 proteins are implicated in metastasis development in several cancers. In this study, we analyzed the prognostic role of mRNA levels of all S100 proteins in early stage non–small cell lung cancer (NSCLC) patients as well as the pathogenetic of S100A2 in the development of metastasis in NSCLC.

Experimental Design: Microarray data from a large NSCLC patient cohort was analyzed for the prognostic role of S100 proteins for survival in surgically resected NSCLC. Metastatic potential of the S100A2 gene was analyzed in vitro and in a lung cancer mouse model in vivo. Overexpression and RNAi approaches were used for analysis of the biological functions of S100A2.

Results: High mRNA expression levels of several S100 proteins and especially S100A2 were associated with poor survival in surgically resected NSCLC patients. Upon stable transfection into NSCLC cell lines, S100A2 did not alter proliferation. However, S100A2 enhanced transwell migration as well as transendothelial migration in vitro. NOD/SCID mice injected s.c. with NSCLC cells overexpressing S100A2 developed significantly more distant metastasis (64%) than mice with control vector transfected tumor cells (17%; P < 0.05). When mice with S100A2 expressing tumors were treated i.v. with shRNA against S100A2, these mice developed significantly fewer lung metastasis than mice treated with control shRNA (P = 0.021).

Conclusions: These findings identify S100A2 as a strong metastasis inducer in vivo. S100A2 might be a potential biomarker as well as a novel therapeutic target in NSCLC metastasis.
Mechanism and Management of AKT Inhibitor-Induced Hyperglycemia
Authors: Crouthamel Ming-Chih , Kahana Jason A. , Korenchuk Susan , Zhang Shu-Yun , Sundaresan Gobalakrishnan , Eberwein Derek J. , Brown Kathleen K. , Kumar Rakesh
Published: December 2008

Purpose: Insulin-like growth factor-I receptor and phosphoinositide 3-kinase/AKT/mammalian target of rapamycin pathways are among the most active areas of drug discovery in cancer research. However, due to their integral roles in insulin signaling, inhibitors targeting these pathways often lead to hyperglycemia and hyperinsulinemia. We investigated the mechanism of hyperglycemia induced by GSK690693, a pan-AKT kinase inhibitor in clinical development, as well as methods to ameliorate these side effects.

Experimental Design: The effect of GSK690693 on blood glucose, insulin, and glucagon levels was characterized in mice. We then evaluated the effects of commonly prescribed antidiabetic agents on GSK690693-induced hyperglycemia. The mechanism of blood glucose increase was evaluated using fasting and tracer uptake studies and by measuring liver glycogen levels. Finally, approaches to manage AKT inhibitor-induced hyperglycemia were designed using fasting and low carbohydrate diet.

Results: We report that treatment with antidiabetic agents does not significantly affect GSK690693-induced hyperglycemia in rodents. However, administration of GSK690693 in mice significantly reduces liver glycogen (∼90%), suggesting that GSK690693 may inhibit glycogen synthesis and/or activate glycogenolysis. Consistent with this observation, fasting before drug administration reduces baseline liver glycogen levels and attenuates hyperglycemia. Further, GSK690693 also inhibits peripheral glucose uptake and introduction of a low-carbohydrate (7%) or 0% carbohydrate diet after GSK690693 administration effectively reduces diet-induced hyperglycemia in mice.

Conclusions: The mechanism of GSK690693-induced hyperglycemia is related to peripheral insulin resistance, increased gluconeogenesis, and/or hepatic glycogenolysis. A combination of fasting and low carbohydrate diet can reduce the magnitude of hyperglycemia induced by an AKT inhibitor.
An Immune Response Enriched 72-Gene Prognostic Profile for Early-Stage Non–Small-Cell Lung Cancer
Authors: Roepman Paul , Jassem Jacek , Smit Egbert F. , Muley Thomas , Niklinski Jacek , van de Velde Tony , Witteveen Anke T. , Rzyman Witold , Floore Arno , Burgers Sjaak , Giaccone Giuseppe , Meister Michael , Dienemann Hendrik , Skrzypski Marcin , Kozlowski Miroslaw , Mooi Wolter J. , van Zandwijk Nico
Published: December 2008

Purpose: Current staging methods are imprecise for predicting prognosis of early-stage non–small-cell lung cancer (NSCLC). We aimed to develop a gene expression profile for stage I and stage II NSCLC, allowing identification of patients with a high risk of disease recurrence within 2 to 3 years after initial diagnosis.

Experimental Design: We used whole-genome gene expression microarrays to analyze frozen tumor samples from 172 NSCLC patients (pT1-2, N0-1, M0) from five European institutions, who had undergone complete surgical resection. Median follow-up was 89 months (range, 1.2-389) and 64 patients developed a recurrence. A random two thirds of the samples were assigned as the training cohort with the remaining samples set aside for independent validation. Cox proportional hazards models were used to evaluate the association between expression levels of individual genes and patient recurrence-free survival. A nearest mean analysis was used to develop a gene-expression classifier for disease recurrence.

Results: We have developed a 72-gene expression prognostic NSCLC classifier. Based on the classifier score, patients were classified as either high or low risk of disease recurrence. Patients classified as low risk showed a significantly better recurrence-free survival both in the training set (P < 0.001; n = 103) and in the independent validation set (P < 0.01; n = 69). Genes in our prognostic signature were strongly enriched for genes associated with immune response.

Conclusions: Our 72-gene signature is closely associated with recurrence-free and overall survival in early-stage NSCLC patients and may become a tool for patient selection for adjuvant therapy.
Alemtuzumab Therapy for Hypereosinophilic Syndrome and Chronic Eosinophilic Leukemia
Authors: Verstovsek Srdan , Tefferi Ayalew , Kantarjian Hagop , Manshouri Taghi , Luthra Raja , Pardanani Animesh , Quintás-Cardama Alfonso , Ravandi Farhad , Ault Pat , Bueso-Ramos Carlos , Cortes Jorge E.
Published: December 2008

Purpose: Patients with hypereosinophilic syndrome (HES) or chronic eosinophilic leukemia (CEL) that are refractory to standard therapies are difficult to manage and have significantly shortened life expectancy.

Experimental Design: CD52 is a surface glycoprotein highly expressed on eosinophils. We treated 11 patients with advanced HES/CEL with alemtuzumab, a humanized anti-CD52 monoclonal antibody. Alemtuzumab was administered, in general, first in escalating doses (5, 10, 30 mg i.v. on days 1-3), then at the tolerated dose thrice per week for a total of 12 doses. Patients with complete hematologic response (CHR; normal percent and absolute eosinophil count) were allowed to continue therapy once weekly as maintenance.

Results: Ten patients (91%) achieved CHR after a median of 2 weeks (0.5-5 weeks) of therapy. Bone marrow eosinophilia resolved in four of seven evaluable patients. The median duration of CHR was 3 months (1.5-17+ months). Seven of the 10 CHR patients relapsed, five while off-therapy. Two patients achieved second CHR upon alemtuzumab rechallenge. Three patients experienced mild infusion-related symptoms, two developed cytomegalovirus reactivation requiring therapy, and one developed orbital lymphoma that was successfully treated.

Conclusions: Our limited experience suggests alemtuzumab to be a valuable therapy for advanced HES or CEL, refractory to standard therapies, and supports the clinical evaluation of alemtuzumab in a larger trial.
Induction of Apoptosis in Human Leukemia Cells by Grape Seed Extract Occurs via Activation of c-Jun NH2-Terminal Kinase
Authors: Gao Ning , Budhraja Amit , Cheng Senping , Yao Hua , Zhang Zhuo , Shi Xianglin
Published: December 2008

Purpose: To characterize the functional role of c-Jun NH2-terminal kinase (JNK) and other apoptotic pathways in grape seed extract (GSE)-induced apoptosis in human leukemia cells by using pharmacologic and genetic approaches.

Experimental Design: Jurkat cells were treated with various concentrations of GSE for 12 and 24 h or with 50 μg/mL GSE for various time intervals, after which apoptosis, caspase activation, and cell signaling pathways were evaluated. Parallel studies were done in U937 and HL-60 human leukemia cells.

Results: Exposure of Jurkat cells to GSE resulted in dose- and time-dependent increase in apoptosis and caspase activation, events associated with the pronounced increase in Cip1/p21 protein level. Furthermore, treatment of Jurkat cells with GSE resulted in marked increase in levels of phospho-JNK. Conversely, interruption of the JNK pathway by pharmacologic inhibitor (e.g., SP600125) or genetic (e.g., small interfering RNA) approaches displayed significant protection against GSE-mediated lethality in Jurkat cells.

Conclusions: The result of the present study showed that GSE induces apoptosis in Jurkat cells through a process that involves sustained JNK activation and Cip1/p21 up-regulation, culminating in caspase activation.
BH3 Mimetic Obatoclax Enhances TRAIL-Mediated Apoptosis in Human Pancreatic Cancer Cells
Authors: Huang Shengbing , Okumura Kenji , Sinicrope Frank A.
Published: December 2008

Purpose: Prosurvival Bcl-2 proteins inhibit the mitochondrial and death receptor-mediated apoptotic pathways. Obatoclax is a small-molecule antagonist of the BH3-binding groove of Bcl-2 proteins that may enhance tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) sensitivity and efficacy.

Experimental Design: Human pancreatic cancer cell lines were incubated with obatoclax and/or TRAIL and cell viability, Annexin V labeling, caspase cleavage, and cytochrome c release were measured. In drug-treated cell lines, protein-protein interactions were studied by immunoprecipitation. Bax/Bak activation was analyzed using conformation-specific antibodies. Lentiviral short hairpin RNA was used to knockdown Bim, Bid, and apoptosis-inducing factor (AIF) expression.

Results: Obatoclax reduced the viability of PANC-1 and BxPC-3 cell lines and synergistically enhanced TRAIL-mediated cytotoxicity. Obatoclax enhanced TRAIL-mediated apoptosis, as shown by Annexin V labeling, which was accompanied by caspase activation (caspase-8, -9, and -3) and cleavage of Bid. Obatoclax potentiated TRAIL-mediated Bax/Bak activation and the release of mitochondrial cytochrome c, Smac, and AIF. Mechanisms underlying the apoptotic effect of obatoclax include displacement of Bak from its sequestration by Bcl-xL or Mcl-1 and release of Bim from Bcl-2 or Mcl-1. Bid knockdown by short hairpin RNA attenuated caspase cleavage and cytotoxicity of obatoclax plus TRAIL. Bim knockdown failed to inhibit the cytotoxic effect of obatoclax alone or combined with TRAIL yet attenuated TRAIL-mediated cytotoxicity. AIF knockdown attenuated cytotoxicity of the drug combination.

Conclusions: Obatoclax potentiates TRAIL-mediated apoptosis by unsequestering Bak and Bim from Bcl-2/Bcl-xL or Mcl-1 proteins. This drug combination enhances Bid-mediated cross-talk between the mitochondrial and death receptor-mediated apoptotic pathways and may represent a novel therapeutic strategy against pancreatic cancer.
Characterization of Genetically Modified T-Cell Receptors that Recognize the CEA:691-699 Peptide in the Context of HLA-A2.1 on Human Colorectal Cancer Cells
Authors: Parkhurst Maria R. , Joo Jayne , Riley John P. , Yu Zhiya , Li Yong , Robbins Paul F. , Rosenberg Steven A.
Published: December 2008

Purpose: Carcinoembryonic antigen (CEA) is a tumor-associated protein expressed on a variety of adenocarcinomas. To develop an immunotherapy for patients with cancers that overexpress CEA, we isolated and genetically modified a T-cell receptors (TCRs) that specifically bound a CEA peptide on human cancer cells.

Experimental Design: HLA-A2.1 transgenic mice were immunized with CEA:691-699. A CEA-reactive TCR was isolated from splenocytes of these mice and was genetically introduced into human peripheral blood lymphocytes via RNA electroporation or retroviral transduction. Amino acid substitutions were introduced throughout the complementarity determining regions (CDR1, CDR2, and CDR3) of both TCR α and β chains to improve recognition of CEA.

Results: Murine lymphocytes bearing the CEA-reactive TCR specifically recognized peptide-loaded T2 cells and HLA-A2.1+ CEA+ human colon cancer cells. Both CD8+ and CD4+ human lymphocytes expressing the murine TCR specifically recognized peptide-loaded T2 cells. However, only gene-modified CD8+ lymphocytes specifically recognized HLA-A2.1+ CEA+ colon cancer cell lines, and tumor cell recognition was weak and variable. We identified two substitutions in the CDR3 of the α chain that significantly influenced tumor cell recognition by human peripheral blood lymphocytes. One substitution, T for S at position 112 (S112T), enhanced tumor cell recognition by CD8+ lymphocytes, and a second dually substituted receptor (S112T L110F) enhanced tumor cell recognition by CD4+ T cells.

Conclusions: The modified CEA-reactive TCRs are good candidates for future gene therapy clinical trials and show the power of selected amino acid substitutions in the antigen-binding regions of the TCR to enhance desired reactivities.