bluuebunny/crossref_metadata_2025 huggingface dataset:
Predicting Workplace Aggression and Violence
Authors: Barling Julian , Dupré Kathryne E. , Kelloway E. Kevin
Published: September 2008

Consistent with the relative recency of research on workplace aggression and the considerable media attention given to high-profile incidents, numerous myths about the nature of workplace aggression have emerged. In this review, we examine these myths from an evidence-based perspective, bringing greater clarity to our understanding of the predictors of workplace aggression. We conclude by pointing to the need for more research focusing on construct validity and prevention issues as well as for methodologies that minimize the likelihood of mono-method bias and that strengthen the ability to make causal inferences.
The second Conceptual Assessment in the Biological Sciences workshop
Authors: Michael Joel , McFarland Jenny , Wright Ann
Published: September 2008

A second National Science Foundation-sponsored workshop on Conceptual Assessment in Biology was held in January 2008. Reports prepared for the workshop revealed that research groups working in a variety of biological sciences are continuing to develop conceptual assessment instruments for use in the classroom. Discussions at this meeting largely focused on two issues: 1) the utility of the backwards design approach of Wiggins and McTighe ( 11 ), in which identification of learning outcomes (determining what to assess) lies at the beginning of course design; and 2) the utility of defining expected learning outcomes as the building of runable mental models (and designing conceptual assessments that would test the correctness of these mental models). A third meeting is being planned that will focus on the processes involved in writing and validating conceptual assessment instruments.
Explorations in statistics: standard deviations and standard errors
Authors: Curran-Everett Douglas
Published: September 2008

Learning about statistics is a lot like learning about science: the learning is more meaningful if you can actively explore. This series in Advances in Physiology Education provides an opportunity to do just that: we will investigate basic concepts in statistics using the free software package R. Because this series uses R solely as a vehicle with which to explore basic concepts in statistics, I provide the requisite R commands. In this inaugural paper we explore the essential distinction between standard deviation and standard error: a standard deviation estimates the variability among sample observations whereas a standard error of the mean estimates the variability among theoretical sample means. If we fail to report the standard deviation, then we fail to fully report our data. Because it incorporates information about sample size, the standard error of the mean is a misguided estimate of variability among observations. Instead, the standard error of the mean provides an estimate of the uncertainty of the true value of the population mean.
Using the pathophysiology of obstructive sleep apnea to teach cardiopulmonary integration
Authors: Levitzky Michael G.
Published: September 2008

Obstructive sleep apnea (OSA) is a common disorder of upper airway obstruction during sleep. The effects of intermittent upper airway obstruction include alveolar hypoventilation, altered arterial blood gases and acid-base status, and stimulation of the arterial chemoreceptors, which leads to frequent arousals. These arousals disturb sleep architecture and cause hypersomnolence. Chronic intermittent alveolar and systemic arterial hypoxia-hypercapnia can cause pulmonary and systemic hypertension, with effects on the right and left ventricles, and even the renal system. The pathophysiology of OSA can therefore be used to review and integrate many topics in pulmonary and cardiovascular physiology in the context of problem-based learning, a guided discussion, or a formal lecture. The discussion begins with a case scenario, followed by a definition of the disorder, the common symptoms and signs of OSA, and a description of an apneic event. These are related to the physiology of the upper airway in OSA, normal alterations in the respiratory system during sleep, the effects of apnea on gas exchange and arterial blood gases, and the cardiovascular consequences of alterations in alveolar and systemic arterial Po2 and Pco2. The treatment of OSA, particularly how the use of continuous positive airway pressure relates to the pathophysiology of the disorder, is discussed briefly.
Using “spinal shrinkage” as a trigger for motivating students to learn about obesity and adopt a healthy lifestyle
Authors: Yar Talay
Published: September 2008

Obesity is a global problem; however, relatively little attention is directed toward preparing and inspiring students of medicine and allied medical sciences to address this serious matter. Students are not routinely exposed to the assessment methods for obesity, its overall prevalence, causative factors, short- and long-term consequences, and its management by lifestyle modification. This physiology laboratory exercise involving students of medicine ( n = 106) was developed to 1) introduce medical students to methods of obesity assessment and to differentiate between general and abdominal obesity, 2) generate an interest and sensitivity about obesity, and 3) stimulate thinking about modification of their lifestyle in relation to eating habits, weight control, and physical activity. Spinal shrinkage (the difference between the standing height of a person and his/her recumbent length) was used as an immediate observable parameter to demonstrate the effect of adiposity. Spinal shrinkage is recognized as an index of the compressive forces acting on the spine and is related to body mass index. A positive correlation ( r = 0.365, P < 0.05) was observed between body mass index and spinal shrinkage. A questionnaire was used to assess student responses to this exercise. Students were motivated to engage in more physical activity (74%), adopt healthier eating (63%), and enhance their knowledge about obesity (67%). They expressed keen interest in the laboratory exercise and found the sessions enjoyable (91%). The laboratory exercise proved to be a success in motivating the students to actively learn and inquire about obesity and to adopt a healthier lifestyle.
Teaching alveolar ventilation with simple, inexpensive models
Authors: DiCarlo Stephen E.
Published: September 2008

When teaching and learning about alveolar ventilation with our class of 300 first-year medical students, we use four simple, inexpensive “models.” The models, which encourage research-oriented learning and help our students to understand complex ideas, are distributed to the students before class. The students anticipate something new every day, and the models provide elements of surprise and physical examples and are designed to help students to understand 1) cohesive forces of the intrapleural space, 2) chest wall and lung dynamics, 3) alveolar volumes, and 4) regional differences in ventilation. Students are drawn into discussion by the power of learning that is associated with manipulating and thinking about objects. Specifically, the models encourage thinking about complex interactions, and the students appreciate manipulating objects and actually understanding how they work. Using models also allows us to show students how we think as well as what we know. Finally, students enjoy taking the models home to demonstrate to friends and family “how the body works” as well as use the models as future study aids.
Chronic central leptin infusion restores cardiac sympathetic-vagal balance and baroreflex sensitivity in diabetic rats
Authors: do Carmo Jussara M. , Hall John E. , da Silva Alexandre A.
Published: September 2008

This study tested whether leptin restores sympathetic-vagal balance, heart rate (HR) variability, and cardiac baroreflex sensitivity (BRS) in streptozotocin (STZ)-induced diabetes. Sprague-Dawley rats were instrumented with arterial and venous catheters, and a cannula was placed in the lateral ventricle for intracerebroventricular (ICV) leptin infusion. Blood pressure (BP) and HR were monitored by telemetry. BRS and HR variability were estimated by linear regression between HR and BP responses to phenylephrine or sodium nitroprusside and autoregressive spectral analysis. Measurements were made during control period, 7 days after induction of diabetes, and 7 days after ICV leptin infusion. STZ diabetes was associated with hyperglycemia (422 ± 17 mg/dl) and bradycardia (−79 ± 4 beats/min). Leptin decreased glucose levels (165 ± 16 mg/dl) and raised HR to control values (303 ± 10 to 389 ± 10 beats/min). Intrinsic HR (IHR) and chronotropic responses to a full-blocking dose of propranolol and atropine were reduced during diabetes (260 ± 7 vs. 316 ± 6, −19 ± 2 vs. −43 ± 6, and 39 ± 3 vs. 68 ± 8 beats/min), and leptin treatment restored these variables to normal (300 ± 7, −68 ± 10, and 71 ± 8 beats/min). Leptin normalized BRS (bradycardia, −2.6 ± 0.3, −1.7 ± 0.2, and −3.0 ± 0.5; and tachycardia, −3.2 ± 0.4, −1.9 ± 0.3, and −3.4 ± 0.3 beats·min−1·mmHg−1 for control, diabetes, and leptin) and HR variability (23 ± 4 to 11 ± 1.5 ms2). Chronic glucose infusion to maintain hyperglycemia during leptin infusion did not alter the effect of leptin on IHR but abolished the improved BRS. These results show rapid impairment of autonomic nervous system control of HR after the induction of diabetes and that central nervous system actions of leptin can abolish the hyperglycemia as well as the altered IHR and BRS in STZ-induced diabetes.
PPADS does not block contraction-induced prostaglandin E2synthesis in cat skeletal muscle
Authors: McCord Jennifer L. , Hayes Shawn G. , Kaufman Marc P.
Published: September 2008

Pyridoxal-phosphate-6-azophenyl-2′-4-disulfonate (PPADS), a purinergic 2 (P2) receptor antagonist, has been shown to attenuate the exercise pressor reflex in cats. In vitro, however, PPADS has been shown to block the production of prostaglandins, some of which play a role in evoking the exercise pressor reflex. Thus the possibility exists that PPADS blocks the exercise pressor reflex through a reduction in prostaglandin synthesis rather than through the blockade of P2receptors. Using microdialysis, we collected interstitial fluid from skeletal muscle to determine prostaglandin E2(PGE2) concentrations during the intermittent contraction of the triceps surae muscle before and after a popliteal arterial injection of PPADS (10 mg/kg). We found that the PGE2concentration increased in response to the intermittent contraction before and after the injection of PPADS (both, P < 0.05). PPADS reduced the pressor response to exercise ( P < 0.05) but had no effect on the magnitude of PGE2production during contraction ( P = 0.48). These experiments demonstrate that PPADS does not block the exercise pressor reflex through a reduction in PGE2synthesis. We suggest that PGE2and P2receptors play independent roles in stimulating the exercise pressor reflex.
P-Glycoprotein–Mediated Resistance to Hsp90-Directed Therapy Is Eclipsed by the Heat Shock Response
Authors: McCollum Andrea K. , TenEyck Cynthia J. , Stensgard Bridget , Morlan Bruce W. , Ballman Karla V. , Jenkins Robert B. , Toft David O. , Erlichman Charles
Published: September 2008

Despite studies that show the antitumor activity of Hsp90 inhibitors, such as geldanamycin (GA) and its derivative 17-allylamino-demethoxygeldanamycin (17-AAG), recent reports indicate that these inhibitors lack significant single-agent clinical activity. Resistance to Hsp90 inhibitors has been previously linked to expression of P-glycoprotein (P-gp) and the multidrug resistant (MDR) phenotype. However, the stress response induced by GA treatment can also cause resistance to Hsp90-targeted therapy. Therefore, we chose to further investigate the relative importance of P-gp and the stress response in 17-AAG resistance. Colony-forming assays revealed that high expression of P-gp could increase the 17-AAG IC50 6-fold in cells transfected with P-gp compared with parent cells. A549 cells selected for resistance to GA overexpressed P-gp, but verapamil did not reverse the resistance. These cells also overexpressed Hsp27, and Hsp70 was induced with 17-AAG treatment. When the GA and 17-AAG resistant cells were transfected with Hsp27 and/or Hsp70 small interfering RNA (siRNA), the 17-AAG IC50 decreased 10-fold compared with control transfected cells. Transfection with siRNA directed against Hsp27, Hsp70, or Hsp27 and Hsp70 also increased sensitivity to EC78, a purine scaffold-based Hsp90 inhibitor that is not a P-gp substrate. We conclude that P-gp may contribute, in part, to resistance to 17-AAG, but induction of stress response proteins, such as Hsp27 and Hsp70, by Hsp90-targeted therapy plays a larger role. Taken together, our results indicate that targeting of Hsp27 and Hsp70 should be exploited to increase the clinical efficacy of Hsp90-directed therapy. [Cancer Res 2008;68(18):7419–27]
A Novel, Macrophage Migration Inhibitory Factor Suicide Substrate Inhibits Motility and Growth of Lung Cancer Cells
Authors: Winner Millicent , Meier Jason , Zierow Swen , Rendon Beatriz E. , Crichlow Gregg V. , Riggs Randall , Bucala Richard , Leng Lin , Smith Ned , Lolis Elias , Trent John O. , Mitchell Robert A.
Published: September 2008

Although chemokine and growth factor receptors are attractive and popular targets for cancer therapeutic intervention, structure-based targeting of the ligands themselves is generally not considered practical. New evidence indicates that a notable exception to this is macrophage migration inhibitory factor (MIF). MIF, an autocrine- and paracrine-acting cytokine/growth factor, plays a pivotal role in both the initiation and maintenance of neoplastic diseases. MIF possesses a nonphysiologic enzymatic activity that is evolutionarily well-conserved. Although small molecule antagonists of MIFs enzymatic active site have been reported to inhibit biological activities of MIF, universally high IC50s have limited their clinical appeal. Using a computational virtual screening strategy, we have identified a unique small molecule inhibitor that serves as a suicide substrate for MIF, resulting in the covalent modification of the catalytically active NH2-terminal proline. Our studies further reveal that this compound, 4-iodo-6-phenylpyrimidine (4-IPP), is ∼5× to 10× times more potent in blocking MIF-dependent catalysis and lung adenocarcinoma cell migration and anchorage-independent growth than the prototypical MIF inhibitor, ISO-1. Finally, using an in silico combinatorial optimization strategy, we have identified four unique congeners of 4-IPP that exhibit MIF inhibitory activity at concentrations 10× to 20× lower than that of parental 4-IPP. [Cancer Res 2008;68(18):7253–7]
Curcumin Inhibits the Proteasome Activity in Human Colon Cancer Cells In vitro and In vivo
Authors: Milacic Vesna , Banerjee Sanjeev , Landis-Piwowar Kristin R. , Sarkar Fazlul H. , Majumdar Adhip P.N. , Dou Q. Ping
Published: September 2008

Curcumin (diferuloylmethane) is the major active ingredient of turmeric (Curcuma longa) used in South Asian cuisine for centuries. Curcumin has been shown to inhibit the growth of transformed cells and to have a number of potential molecular targets. However, the essential molecular targets of curcumin under physiologic conditions have not been completely defined. Herein, we report that the tumor cellular proteasome is most likely an important target of curcumin. Nucleophilic susceptibility and in silico docking studies show that both carbonyl carbons of the curcumin molecule are highly susceptible to a nucleophilic attack by the hydroxyl group of the NH2-terminal threonine of the proteasomal chymotrypsin-like (CT-like) subunit. Consistently, curcumin potently inhibits the CT-like activity of a purified rabbit 20S proteasome (IC50 = 1.85 μmol/L) and cellular 26S proteasome. Furthermore, inhibition of proteasome activity by curcumin in human colon cancer HCT-116 and SW480 cell lines leads to accumulation of ubiquitinated proteins and several proteasome target proteins, and subsequent induction of apoptosis. Furthermore, treatment of HCT-116 colon tumor–bearing ICR SCID mice with curcumin resulted in decreased tumor growth, associated with proteasome inhibition, proliferation suppression, and apoptosis induction in tumor tissues. Our study shows that proteasome inhibition could be one of the mechanisms for the chemopreventive and/or therapeutic roles of curcumin in human colon cancer. Based on its ability to inhibit the proteasome and induce apoptosis in both HCT-116 and metastatic SW480 colon cancer cell lines, our study suggests that curcumin could potentially be used for treatment of both early-stage and late-stage/refractory colon cancer. [Cancer Res 2008;68(18):7283–92]
Regulation of Apoptosis and Caspase-8 Expression in Neuroblastoma Cells by Isoforms of the IG20 Gene
Authors: Li Liang Cheng , Sheng Jian Rong , Mulherkar Nirupama , Prabhakar Bellur S. , Meriggioli Matthew N.
Published: September 2008

The IG20 gene undergoes alternative splicing resulting in the differential expression of six putative splice variants. Four of these (IG20pa, MADD, IG20-SV2, and DENN-SV) are expressed in virtually all human tissues. However, investigations examining alternative splicing of the IG20 gene to date have been largely limited to nonneural malignant and nonmalignant cells. In this study, we investigated the expression of alternative splice isoforms of the IG20 gene in human neuroblastoma cells. We found that six IG20 splice variants (IG20-SVs) were expressed in two human neuroblastoma cell lines (SK-N-SH and SH-SY5Y), highlighted by the expression of two unique splice isoforms (i.e., KIAA0358 and IG20-SV4). Similarly, we found enriched expression of these two IG20-SVs in human neural tissues derived from cerebral cortex, hippocampus, and, to a lesser extent, spinal cord. Using gain-of-function studies and siRNA technology, we determined that these “neural-enriched isoforms” exerted significant and contrasting effects on vulnerability to apoptosis in neuroblastoma cells. Specifically, expression of KIAA0358 exerted a potent antiapoptotic effect in both the SK-N-SH and SH-SY5Y neuroblastoma cell lines, whereas expression of IG20-SV4 had proapoptotic effects directly related to the activation of caspase-8 in these cells, which have minimal or absent constitutive caspase-8 expression. These data indicate that the pattern of expression of these neural-enriched IG20-SVs regulates the expression and activation of caspase-8 in certain neuroblastoma cells, and that manipulation of IG20-SV expression pattern may represent a potent therapeutic strategy in the therapy of neuroblastoma and perhaps other cancers. [Cancer Res 2008;68(18):7352–61]
Purine-Rich Box-1–Mediated Reduced Expression of CD20 Alters Rituximab-Induced Lysis of Chronic Lymphocytic Leukemia B Cells
Authors: Mankaï Amani , Bordron Anne , Renaudineau Yves , Martins-Carvalho Christine , Takahashi Shinichiro , Ghedira Ibtissem , Berthou Christian , Youinou Pierre
Published: September 2008

The anti-CD20 monoclonal antibody rituximab has been less successful in treating chronic lymphocytic leukemia (CLL) than lymphoma, possibly due to the lower density of CD20 on B lymphocytes from CLL patients than on those from lymphoma patients. This lowering may result from insufficiency of one of the transcription factors of cd20. Of these, purine-rich box-1 (PU.1) is poorly expressed in CLL. To estimate its weight in CD20 expression, pu.1 cDNA was transfected into CLL B cells and shown to raise the membrane expression of CD20 and to improve the rituximab-induced lysis of transfected cells. Granulocyte macrophage colony-stimulating factor and all-trans-retinoic acids were not involved in the defective expression of PU.1 or the excessive methylation of the pu.1 gene, because 6 of 14 CLL samples tested were normally methylated. This was confirmed by the failure of DNA methyltransferase inhibitors to restore pu.1 transcription in hypermethylated CLL, and, in fact, the expression of PU.1 was down-regulated by excessive expression of the FMS proto-oncogene–like tyrosine kinase 3 (Flt3) receptor. This abnormality is consistent with our finding of elevated levels of Flt3 ligand (FL) in 20 of 23 CLL sera tested. We propose that FL-dependent increased Flt3 signaling prevents the expression of PU.1, which down-regulates that of CD20, and accounts for resistance of leukemic B cells to rituximab-induced lysis. [Cancer Res 2008;68(18):7512–9]
A Novel Gene Expression Profile in Lymphatics Associated with Tumor Growth and Nodal Metastasis
Authors: Clasper Steven , Royston Daniel , Baban Dilair , Cao Yihai , Ewers Stephan , Butz Stefan , Vestweber Dietmar , Jackson David G.
Published: September 2008

Invasion of lymphatic vessels is a key step in the metastasis of primary tumors to draining lymph nodes. Although the process is enhanced by tumor lymphangiogenesis, it is unclear whether this is a consequence of increased lymphatic vessel number, altered lymphatic vessel properties, or both. Here we have addressed the question by comparing the RNA profiles of primary lymphatic endothelial cells (LEC) isolated from the vasculature of normal tissue and from highly metastatic T-241/vascular endothelial growth factor (VEGF)-C fibrosarcomas implanted in C57BL/6 mice. Our findings reveal significant differences in expression of some 792 genes (i.e., ≥2-fold up- or down-regulated, P ≤ 0.05) that code for a variety of proteins including components of endothelial junctions, subendothelial matrix, and vessel growth/patterning. The tumor LEC profile, validated by immunohistochemical staining, is distinct from that of normal, inflammatory cytokine, or mitogen-activated LEC, characterized by elevated expression of such functionally significant molecules as the tight junction regulatory protein endothelial specific adhesion molecule (ESAM), the transforming growth factor-β coreceptor Endoglin (CD105), the angiogenesis-associated leptin receptor, and the immunoinhibitory receptor CD200, and reduced expression of subendothelial matrix proteins including collagens, fibrillin, and biglycan. Moreover, we show similar induction of ESAM, Endoglin, and leptin receptor within tumor lymphatics in a series of human head and neck and colorectal carcinomas, and uncover a dramatic correlation between ESAM expression and nodal metastasis that identifies this marker as a possible prognostic indicator. These findings reveal a remarkable degree of phenotypic plasticity in cancer lymphatics and provide new insight into the processes of lymphatic invasion and lymph node metastasis. [Cancer Res 2008;68(18):7293–303]
Curcumin Inhibits Lung Cancer Cell Invasion and Metastasis through the Tumor Suppressor HLJ1
Authors: Chen Huei-Wen , Lee Jen-Yi , Huang Ji-Ying , Wang Chi-Chung , Chen Wan-Jiun , Su Sheng-Fang , Huang Chia-Wen , Ho Chao-Chi , Chen Jeremy J.W. , Tsai Meng-Feng , Yu Sung-Liang , Yang Pan-Chyr
Published: September 2008

Curcumin (diferuloylmethane) is an active component of the spice turmeric and has a diversity of antitumor activities. In this study, we found that curcumin can inhibit cancer cell invasion and metastasis through activation of the tumor suppressor DnaJ-like heat shock protein 40 (HLJ1). Human lung adenocarcinoma cells (CL1-5) treated with curcumin (1–20 μmol/L) showed a concentration-dependent reduction in cell migration, invasion, and metastatic ability, and this was associated with increased HLJ1 expression. Knockdown of HLJ1 expression by siRNA was able to reverse the curcumin-induced anti-invasive and antimetastasis effects in vitro and in vivo. The HLJ1 promoter and enhancer in a luciferase reporter assay revealed that curcumin transcriptionally up-regulates HLJ1 expression through an activator protein (AP-1) site within the HLJ1 enhancer. JunD, one of the AP-1 components, was significantly up-regulated by curcumin (1–20 μmol/L) in a concentration- and time-dependent manner. Knockdown of JunD expression could partially reduce the curcumin-induced HLJ1 activation and diminish the anti-invasive effect of curcumin, indicating that JunD would seem to be involved in curcumin-induced HLJ1 expression. Curcumin was able to induce c-Jun NH2-kinase (JNK) phosphorylation, whereas the JNK inhibitor (SP-600125) could attenuate curcumin-induced JunD and HLJ1 expression. Activation of HLJ1 by curcumin further leads to up-regulation of E-cadherin and a suppression of cancer cell invasion. Our results show that curcumin induces HLJ1, through activation of the JNK/JunD pathway, and inhibits lung cancer cell invasion and metastasis by modulating E-cadherin expression. This is a novel mechanism and supports the application of curcumin in anti–cancer metastasis therapy. [Cancer Res 2008;68(18):7428–38]
Cyp1b1 Exerts Opposing Effects on Intestinal Tumorigenesis via Exogenous and Endogenous Substrates
Authors: Halberg Richard B. , Larsen Michele Campaigne , Elmergreen Tammy L. , Ko Alex Y. , Irving Amy A. , Clipson Linda , Jefcoate Colin R.
Published: September 2008

Cytochrome P450 1B1 (Cyp1b1) metabolism contributes to physiologic functions during embryogenesis but also to carcinogenic activation of polycyclic aromatic hydrocarbons (PAH). We generated Cyp1b1-deficient mice carrying the Min allele of the adenomatous polyposis coli gene. These Cyp1b1-deficient Min mice developed twice as many tumors as Min controls, which, however, remained similar in size and histology. Tumors from older (130 days) Cyp1b1-deficient Min mice selectively exhibited focal areas of nuclear atypia associated with less organized epithelia. The metabolism of endogenous substrates by Cyp1b1, therefore, suppresses tumor initiation but also affects progression. Treatment of Min mice with 7,12-dimethylbenzanthracene (DMBA) doubled both tumor multiplicity and size within 20 days but not when mice lacked Cyp1b1. This was paralleled by an abnormal staining of crypts with β-catenin, phospho-IκB kinase, and RelA, which may represent an early stage of tumorigenesis similar to aberrant crypt formation. Cyp1b1 deletion did not affect circulating DMBA and metabolites. Cyp1b1 expression was higher in the tumors compared with normal small intestines. Increased tumorigenesis may, therefore, arise from generation of DMBA metabolites by Cyp1b1 in the developing tumors. Benzo(a)pyrene (BP), which is similarly activated by Cyp1b1 in vitro, did not affect tumorigenesis in Min mice. By contrast, BP and DMBA each suppressed tumor multiplicity in the absence of Cyp1b1. Cyp1b1 metabolism of DMBA and endogenous oxygenation products may each affect a tumor-promoting nuclear factor-κB activation, whereas Ah receptor activation by PAH affects suppression. Tumorigenesis may, therefore, depend on activation of PAH by Cyp1b1 and on offsetting suppression by Cyp1b1 of endogenous tumor-enhancing substrates. [Cancer Res 2008;68(18):7394–402]
HSF1 as a Mitotic Regulator: Phosphorylation of HSF1 by Plk1 Is Essential for Mitotic Progression
Authors: Lee Yoon-Jin , Kim Eun-Ho , Lee Jae Seon , Jeoung Dooil , Bae Sangwoo , Kwon Seung Hae , Lee Yun-Sil
Published: September 2008

Previously, heat shock factor 1 (HSF1) had been reported to induce genomic instability and aneuploidy by interaction with Cdc20. Here, we have further examined the functions of HSF1 in the regulation of mitosis. A null mutant or knockdown of HSF1 caused defective mitotic progression. By monitoring chromosomes in living cells, we determined that HSF1 was localized to the centrosome in mitosis and especially to the spindle poles in metaphase. HSF1 was phosphorylated by Plk1 at Ser216 of the DSGXXS motif during the timing of mitosis and a phospho-defective mutant form of HSF1 inhibited mitotic progression. Phosphorylated HSF1 during spindle pole localization underwent ubiquitin degradation through the SCFβ-TrCP pathway. However, binding of HSF1 with Cdc20 stabilized the phosphorylation of HSF1. Moreover, SCFβ-TrCP–mediated degradation only occurred when phosphorylated HSF1 was released from Cdc20. HSF1 phosphorylation at Ser216 occurred in the early mitotic period with simultaneous binding of Cdc20. The interaction of HSF1 with SCFβ-TrCP was followed and then the interaction of APC/Cdc20 was subsequently observed. From these findings, it was shown that Plk1 phosphorylates HSF1 in early mitosis and that the binding of phosphorylated HSF1 with Cdc20 and ubiquitin degradation by SCFβ-TrCP regulates mitotic progression. [Cancer Res 2008;68(18):7550–60]
Growth Inhibition and Radiosensitization of Glioblastoma and Lung Cancer Cells by Small Interfering RNA Silencing of Tumor Necrosis Factor Receptor–Associated Factor 2
Authors: Zheng Min , Morgan-Lappe Susan E. , Yang Jie , Bockbrader Katrina M. , Pamarthy Deepika , Thomas Dafydd , Fesik Stephen W. , Sun Yi
Published: September 2008

Radiotherapy combined with chemotherapy is the treatment of choice for glioblastoma and locally advanced lung cancer, but radioresistance of these two types of cancer remains a significant therapeutic hindrance. To identify molecular target(s) for radiosensitization, we screened a small interfering RNA (siRNA) library targeting all protein kinases and E3 ubiquitin ligases in the human genome and identified tumor necrosis factor receptor–associated factor 2 (TRAF2). Silencing of TRAF2 using siRNA caused a significant growth suppression of glioblastoma U251 cells and moderately sensitized these radioresistant cells to radiation. Overexpression of a really interesting new gene (RING)–deleted dominant-negative TRAF2 mutant also conferred radiosensitivity, whereas overexpression of wild-type (WT) TRAF2 significantly protected cells from radiation-induced killing. Likewise, siRNA silencing of TRAF2 in radioresistant lung cancer H1299 cells caused growth suppression and radiosensitization, whereas overexpression of WT TRAF2 enhanced radioresistance in a RING ligase-dependent manner. Moreover, siRNA silencing of TRAF2 in UM-SCC-1 head and neck cancer cells also conferred radiosensitization. Further support for the role of TRAF2 in cancer comes from the observations that TRAF2 is overexpressed in both lung adenocarcinoma tissues and multiple lung cancer cell lines. Importantly, TRAF2 expression was very low in normal bronchial epithelial NL20 cells, and TRAF2 silencing had a minimal effect on NL20 growth and radiation sensitivity. Mechanistically, TRAF2 silencing blocks the activation of the nuclear factor-κB signaling pathway and down-regulates several G2-M cell cycle control proteins, resulting in enhanced G2-M arrest, growth suppression, and radiosensitization. Our studies suggest that TRAF2 is an attractive drug target for anticancer therapy and radiosensitization. [Cancer Res 2008;68(18):7570–8]
Transient Inhibition of ATM Kinase Is Sufficient to Enhance Cellular Sensitivity to Ionizing Radiation
Authors: Rainey Michael D. , Charlton Maura E. , Stanton Robert V. , Kastan Michael B.
Published: September 2008

In response to DNA damage, the ATM protein kinase activates signal transduction pathways essential for coordinating cell cycle progression with DNA repair. In the human disease ataxia-telangiectasia, mutation of the ATM gene results in multiple cellular defects, including enhanced sensitivity to ionizing radiation (IR). This phenotype highlights ATM as a potential target for novel inhibitors that could be used to enhance tumor cell sensitivity to radiotherapy. A targeted compound library was screened for potential inhibitors of the ATM kinase, and CP466722 was identified. The compound is nontoxic and does not inhibit phosphatidylinositol 3-kinase (PI3K) or PI3K-like protein kinase family members in cells. CP466722 inhibited cellular ATM-dependent phosphorylation events and disruption of ATM function resulted in characteristic cell cycle checkpoint defects. Inhibition of cellular ATM kinase activity was rapidly and completely reversed by removing CP466722. Interestingly, clonogenic survival assays showed that transient inhibition of ATM is sufficient to sensitize cells to IR and suggests that therapeutic radiosensitization may only require ATM inhibition for short periods of time. The ability of CP466722 to rapidly and reversibly regulate ATM activity provides a new tool to ask questions about ATM function that could not easily be addressed using genetic models or RNA interference technologies. [Cancer Res 2008;68(18):7466–74]
Tube Travel: The Role of Proteases in Individual and Collective Cancer Cell Invasion
Authors: Friedl Peter , Wolf Katarina
Published: September 2008

Recent advances in high-resolution multimodal microscopy reveal how MT1-matrix metalloproteinase (MMP)/MMP-14 and other cell surface proteases degrade and remodel the extracellular matrix (ECM) to drive the dissemination of cancer cells into normal adjacent tissue. By cleaving collagen fibers and repatterning them into parallel bundles, individual cells reorient the ECM to permit movement in tube-like microtracks. Cells along the edge of these tubes can excavate ECM outward, generating macrotracks through which collective mass movement of cancer cells can occur. These findings develop our understanding of invasive processes in cancer and how to attack them by interfering with MMP-14 activity. [Cancer Res 2008;68(18):7247–9]
One-Hit Effects in Cancer: Altered Proteome of Morphologically Normal Colon Crypts in Familial Adenomatous Polyposis
Authors: Yeung Anthony T. , Patel Bhavinkumar B. , Li Xin-Ming , Seeholzer Steven H. , Coudry Renata A. , Cooper Harry S. , Bellacosa Alfonso , Boman Bruce M. , Zhang Tao , Litwin Samuel , Ross Eric A. , Conrad Peggy , Crowell James A. , Kopelovich Levy , Knudson Alfred
Published: September 2008

We studied patients with Familial Adenomatous Polyposis (FAP) because they are virtually certain to develop colon cancer, and because much is known about the causative APC gene. We hypothesized that the inherited heterozygous mutation itself leads to changes in the proteome of morphologically normal crypts and the proteins that changed may represent targets for preventive and therapeutic agents. We determined the differential protein expression of morphologically normal colon crypts of FAP patients versus those of individuals without the mutation, using two-dimensional gel electrophoresis, mass spectrometry, and validation by two-dimensional gel Western blotting. Approximately 13% of 1,695 identified proteins were abnormally expressed in the morphologically normal crypts of APC mutation carriers, indicating that a colon crypt cell under the one-hit state is already abnormal. Many of the expression changes affect pathways consistent with the function of the APC protein, including apoptosis, cell adhesion, cell motility, cytoskeletal organization and biogenesis, mitosis, transcription, and oxidative stress response. Thus, heterozygosity for a mutant APC tumor suppressor gene alters the proteome of normal-appearing crypt cells in a gene-specific manner, consistent with a detectable one-hit event. These changes may represent the earliest biomarkers of colorectal cancer development, potentially leading to the identification of molecular targets for cancer prevention. [Cancer Res 2008;68(18):7579–86]
Development of a Small-Molecule Serum- and Glucocorticoid-Regulated Kinase-1 Antagonist and Its Evaluation as a Prostate Cancer Therapeutic
Authors: Sherk Andrea B. , Frigo Daniel E. , Schnackenberg Christine G. , Bray Jeffrey D. , Laping Nicholas J. , Trizna Walter , Hammond Marlys , Patterson Jaclyn R. , Thompson Scott K. , Kazmin Dmitri , Norris John D. , McDonnell Donald P.
Published: September 2008

Androgens, through their actions on the androgen receptor (AR), are required for the development of the prostate and contribute to the pathologic growth dysregulation observed in prostate cancers. Consequently, androgen ablation has become an essential component of the pharmacotherapy of prostate cancer. In this study, we explored the utility of targeting processes downstream of AR as an alternate approach for therapy. Specifically, we show that the serum and glucocorticoid-regulated kinase 1 (SGK1) gene is an androgen-regulated target gene in cellular models of prostate cancer. Furthermore, functional serum- and glucocorticoid-regulated kinase 1 (SGK1) protein, as determined by the phosphorylation of its target Nedd4-2, was also increased with androgen treatment. Importantly, we determined that RNA interference–mediated knockdown of SGK1 expression attenuates the androgen-mediated growth of the prostate cancer cell line LNCaP. Given these findings, we explored the utility of SGK1 as a therapeutic target in prostate cancer by developing and evaluating a small-molecule inhibitor of this enzyme. From these studies emerged GSK650394, a competitive inhibitor that quantitatively blocks the effect of androgens on LNCaP cell growth. Thus, in addition to androgen ablation, inhibition of pathways downstream of AR is likely to have therapeutic utility in prostate cancer. [Cancer Res 2008;68(18):7475–83]
The Peptidyl-Isomerase Pin1 Regulates p27kip1 Expression through Inhibition of Forkhead Box O Tumor Suppressors
Authors: Brenkman Arjan B. , de Keizer Peter L.J. , van den Broek Niels J.F. , van der Groep Petra , van Diest Paul J. , van der Horst Armando , Smits Alida M.M. , Burgering Boudewijn M.T.
Published: September 2008

The Forkhead box O (FOXO) protein family is an evolutionarily conserved subclass of transcription factors recently identified as bona fide tumor suppressors. Preventing the accumulation of cellular damage due to oxidative stress is thought to underlie its tumor-suppressive role. Oxidative stress, in turn, also feedback controls FOXO4 function. Regulation of this process, however, is poorly understood but may be relevant to the ability of FOXO to control tumor suppression. Here, we characterize novel FOXO4 phosphorylation sites after increased cellular oxidative stress and identify the isomerase Pin1, a protein frequently found to be overexpressed in cancer, as a critical regulator of p27kip1 through FOXO4 inhibition. We show that Pin1 requires these phosphorylation events to act negatively on FOXO4 transcriptional activity. Consistent with this, oxidative stress induces binding of Pin1 to FOXO, thereby attenuating its monoubiquitination, a yet uncharacterized mode of substrate modulation by Pin1. We have previously shown that monoubiquitination is involved in controlling nuclear translocation in response to cellular stress, and indeed, Pin1 prevents nuclear FOXO4 accumulation. Interestingly, Pin1 acts on FOXO through stimulation of the activity of the deubiquitinating enzyme HAUSP/USP7. Ultimately, this results in decreased transcriptional activity towards target genes, including the cell cycle arrest gene p27kip1. Notably, in a primary human breast cancer panel, low p27kip1 levels inversely correlated with Pin1 expression. Thus, Pin1 is identified as a novel negative FOXO regulator, interconnecting FOXO phosphorylation and monoubiquitination in response to cellular stress to regulate p27kip1. [Cancer Res 2008;68(18):7597–605]
An Immortalization-Dependent Switch in Integrin Function Up-regulates MMP-9 to Enhance Tumor Cell Invasion
Authors: Lamar John M. , Pumiglia Kevin M. , DiPersio C. Michael
Published: September 2008

Integrins, the major receptors for cell adhesion to the extracellular matrix, play important roles during tumor progression. However, it is still unclear whether genetic lesions that occur during carcinoma development can lead to altered integrin function, and how changes in integrin function contribute to subsequent carcinoma progression. Loss-of-function mutations in p53 and activating mutations in H-Ras, which immortalize and transform epithelial cells, respectively, are common causal events in squamous cell carcinoma (SCC). Phenotypes resulting from these two genetic lesions promote SCC progression and are, therefore, potential targets for anticancer therapies. We developed a model system of keratinocyte transformation that has allowed us to investigate the individual roles of p53 mutation and oncogenic Ras mutation in the acquisition of integrin α3β1-regulated phenotypes that promote SCC progression. Using this model, we show that keratinocyte immortalization by p53-null mutation causes a switch in α3β1 function that induces matrix metalloproteinase (MMP)-9 gene expression in tumorigenic cells. This acquired α3β1-dependent regulation of MMP-9 was maintained during subsequent transformation by oncogenic Ras, and it promoted invasion of tumorigenic keratinocytes. Our results show that loss of p53 function leads to changes in integrin-mediated gene regulation that occur during SCC progression and play a critical role in tumor cell invasion. [Cancer Res 2008;68(18):7371–9]
Cyclin-Dependent Kinase 3–Mediated Activating Transcription Factor 1 Phosphorylation Enhances Cell Transformation
Authors: Zheng Duo , Cho Yong-Yeon , Lau Andy T.Y. , Zhang Jishuai , Ma Wei-Ya , Bode Ann M. , Dong Zigang
Published: September 2008

Cyclin-dependent kinase (cdk)-3, a member of the cdk family of kinases, plays a critical role in cell cycle regulation and is involved in G0-G1 and G1-S cell cycle transitions. However, the role of cdk3 in cell proliferation, as well as cell transformation, is not yet clearly understood. Here, we report that the protein expression level of cdk3 is higher in human cancer cell lines and human glioblastoma tissue compared with normal brain tissue. Furthermore, we found that cdk3 phosphorylates activating transcription factor 1 (ATF1) at serine 63 and enhances the transactivation and transcriptional activities of ATF1. Results also indicated that siRNA directed against cdk3 (si-cdk3) suppresses ATF1 activity, resulting in inhibition of proliferation and growth of human glioblastoma T98G cells in soft agar. Importantly, we showed that cdk3 enhances epidermal growth factor–induced transformation of JB6 Cl41 cells and si-cdk3 suppresses RasG12V/cdk3/ATF1–induced foci formation in NIH3T3 cells. These results clearly showed that the cdk3-ATF1 signaling axis is critical for cell proliferation and transformation. [Cancer Res 2008;68(18):7650–60]
Coupling of Endoplasmic Reticulum Stress to CDDO-Me–Induced Up-regulation of Death Receptor 5 via a CHOP–Dependent Mechanism Involving JNK Activation
Authors: Zou Wei , Yue Ping , Khuri Fadlo R. , Sun Shi-Yong
Published: September 2008

The synthetic triterpenoid methyl-2-cyano-3,12-dioxoolean-1,9-dien-28-oate (CDDO-Me) is in phase I clinical trials as a novel cancer therapeutic agent. We previously showed that CDDO-Me induces c-Jun NH2-terminal kinase (JNK)–dependent death receptor 5 (DR5) expression and augments death receptor–induced apoptosis. The current study focused on addressing how CDDO-Me induces JNK-dependent DR5 expression. Analysis of DR5 promoter regions defines that the CCAAT/enhancer binding protein homologous protein (CHOP) binding site is responsible for CDDO-Me–induced transactivation of the DR5 gene. Consistently, CDDO-Me induced DR5 expression and parallel CHOP up-regulation. Blockade of CHOP up-regulation also abrogated CDDO-Me–induced DR5 expression. These results indicate that CDDO-Me induces CHOP-dependent DR5 up-regulation. Moreover, the JNK inhibitor SP600125 abrogated CHOP induction by CDDO-Me, suggesting a JNK-dependent CHOP up-regulation by CDDO-Me as well. Importantly, knockdown of CHOP attenuated CDDO-Me–induced apoptosis, showing that CHOP induction is involved in CDDO-Me–induced apoptosis. Additionally, CDDO-Me increased the levels of Bip, phosphorylated eukaryotic translation initiation factor 2α, inositol requiring kinase 1α, and activating transcription factor 4, all of which are featured changes during endoplasmic reticulum (ER) stress. Furthermore, salubrinal, an inhibitor of ER stress–induced apoptosis, inhibited JNK activation and up-regulation of CHOP and DR5 by CDDO-Me and protected cells from CDDO-Me–induced apoptosis. Thus, ER stress seems to be important for CDDO-Me–induced JNK activation, CHOP and DR5 up-regulation, and apoptosis. Collectively, we conclude that CDDO-Me triggers ER stress, leading to JNK-dependent, CHOP-mediated DR5 up-regulation and apoptosis. [Cancer Res 2008;68(18):7484–92]
Combination Therapy with Cisplatin and Anti–4-1BB: Synergistic Anticancer Effects and Amelioration of Cisplatin-Induced Nephrotoxicity
Authors: Kim Young H. , Choi Beom K. , Kim Kwang H. , Kang Sang W. , Kwon Byoung S.
Published: September 2008

Anti–4-1BB and cisplatin showed synergistic anticancer effects in the CT-26 colon carcinoma model, producing complete regression in >60% of mice with either preventive or therapeutic treatment. The tumor-free mice formed long-lasting CD8+ T cell–dependent tumor-specific memory. Anti–4-1BB induced rapid repopulation of T and B cells from cisplatin-mediated lymphopenia and differentiation and expansion of IFN-γ+CD11c+CD8+ T cells. Cisplatin facilitated expansion of naïve, effector, and memory CD8+ T cells; combination therapy produced almost twice as many lymphoid cells as anti–4-1BB alone. Cisplatin increased 4-1BB on antigen-primed T cells and induced 4-1BB de novo on kidney tubular epithelium. Cross-linking of 4-1BB protected the T cells and kidney epithelium from cisplatin-mediated apoptosis by increasing expression of antiapoptotic molecules. Thus, cisplatin-induced 4-1BB provided a mechanism for amelioration of the lymphopenia and nephrotoxicity inherent in cisplatin treatment. We concluded that chemoimmunotherapy with anti–4-1BB and cisplatin is synergistic in tumor killing and prevention of organ-specific toxicity. [Cancer Res 2008;68(18):7264–9]
LKB1 Is Necessary for Akt-Mediated Phosphorylation of Proapoptotic Proteins
Authors: Zhong Diansheng , Liu Xiuju , Khuri Fadlo R. , Sun Shi-Yong , Vertino Paula M. , Zhou Wei
Published: September 2008

LKB1 plays the role of tumor suppressor, opposite to Akt, by negatively regulating mammalian target of rapamycin through the activation of AMP-activated protein kinase and TSC signaling. We have discovered a novel, potentially oncogenic role for LKB1 as a supporter of Akt-mediated phosphorylation of proapoptotic proteins. We found that Akt activation led to increased phosphorylation of FoxO3a at Thr32 in LKB1 wild-type cells but not in LKB1-null cells. Depletion of LKB1 in the cells with wild-type LKB1 resulted in attenuation of that phosphorylation of FoxO3a by activated Akt, whereas the restoration of LKB1 function in LKB1-null cells reestablished Akt-mediated FoxO3a phosphorylation. On expanding our analysis to other Akt targets, using isogenic LKB1 knockdown cell line pairs and a phospho-specific antibody microarray, we observed that there was a requirement for LKB1 in the phosphorylation of other Akt downstream targets, including Ask1 (Ser83), Bad (Ser136), FoxO1 (Ser319), FoxO4 (Ser197), and glycogen synthase kinase 3β (GSK3β; Ser9). Because the phosphorylation of these sites by Akt suppresses apoptosis, the requirement of LKB1 suggests that LKB1 may have an antiapoptotic role in tumor cells with constitutively active Akt. Indeed, we found that the suppression of LKB1 expression led to apoptosis in three cell lines in which Akt is constitutively active but not in two cell lines without Akt activation. This observation may explain the lack of LKB1 somatic mutations in brain, breast, and colon cancers, where Akt is frequently activated due to mutations in phosphatidylinositol 3-kinase, PTEN, or Akt itself. [Cancer Res 2008;68(18):7270–7]