bluuebunny/crossref_metadata_2025 huggingface dataset:
Effects of unique biomedical education programs for engineers: REDEEM and ESTEEM projects
Authors: Matsuki Noriaki , Takeda Motohiro , Yamano Masahiro , Imai Yohsuke , Ishikawa Takuji , Yamaguchi Takami
Published: June 2009

Current engineering applications in the medical arena are extremely progressive. However, it is rather difficult for medical doctors and engineers to discuss issues because they do not always understand one another's jargon or ways of thinking. Ideally, medical engineers should become acquainted with medicine, and engineers should be able to understand how medical doctors think. Tohoku University in Japan has managed a number of unique reeducation programs for working engineers. Recurrent Education for the Development of Engineering Enhanced Medicine has been offered as a basic learning course since 2004, and Education through Synergetic Training for Engineering Enhanced Medicine has been offered as an advanced learning course since 2006. These programs, which were developed especially for engineers, consist of interactive, modular, and disease-based lectures (case studies) and substantial laboratory work. As a result of taking these courses, all students obtained better objective outcomes, on tests, and subjective outcomes, through student satisfaction. In this article, we report on our unique biomedical education programs for engineers and their effects on working engineers.
20-kDa placental hGH-V has diminished diabetogenic and lactogenic activities compared with 22-kDa hGH-N while retaining antilipogenic activity
Authors: Vickers M. H. , Gilmour S. , Gertler A. , Breier B. H. , Tunny K. , Waters M. J. , Gluckman P. D.
Published: June 2009

Placental human growth hormone-variant (hGH-V) and pituitary human growth hormone-N (hGH-N) are of identical size (22 kDa) but differ in 13 residues scattered throughout the protein. Several isoforms of GH are produced by the hGH-N and hGH-V genes including a 20-kDa hGH-V resulting from a 45-bp deletion caused by the use of an alternative acceptor site within exon 3. To date, the biological properties of the 20-kDa GH-V have not been characterized in vivo. Using young male Wistar rats fed either chow or a high-fat (HF) diet for 4 wk postweaning, we investigated the effect of 7 days treatment with either 22-kDa hGH-N, 20-kDa hGH-V (5 ug·g−1·day−1sc), or vehicle on body composition and endocrine and metabolic profiles. Total body growth (absolute weight gain and linear growth trajectory) in the 20-kDa hGH-V-treated animals was intermediary between that of control and hGH-N-treated animals. Both 22-kDa hGH-N and 20-kDa hGH-V significantly reduced total body fat mass compared with control animals, and there were no differences between the GH isoforms in anti-lipogenic activity in animals fed the HF diet. Fasting plasma insulin and C peptide were significantly increased in animals on the HF diet and further increased by hGH-N but were unchanged in 20-kDa hGH-V-treated animals compared with saline-treated controls. Plasma volume as assessed by hematocrit was increased in hGH-N-treated animals but was unchanged in 20-kDa hGH-V-treated animals compared with controls. Furthermore, 20-kDa hGH-V had reduced lactogenic (prolactin receptor mediated) activity characteristic of hGH-N as tested in vitro compared with the 20-kDa hGH-N and 22-kDa hGH-N variants. In summary, placental 20-kDa hGH-V retains some of the growth-promoting and all antilipogenic activities of pituitary 22-kDa hGH-N but has diminished diabetogenic and lactogenic properties compared with the native 22-kDa hGH-N.
Truncation of SNAP-25 reduces the stimulatory action of cAMP on rapid exocytosis in insulin-secreting cells
Authors: Vikman Jenny , Svensson Hjalmar , Huang Ya-Chi , Kang Youhou , Andersson Sofia A. , Gaisano Herbert Y. , Eliasson Lena
Published: June 2009

Synaptosomal protein of 25 kDa (SNAP-25) is important for Ca2+-dependent fusion of large dense core vesicles (LDCVs) in insulin-secreting cells. Exocytosis is further enhanced by cAMP-increasing agents such as glucagon-like peptide-1 (GLP-1), and this augmentation includes interaction with both PKA and cAMP-GEFII. To investigate the coupling between SNAP-25- and cAMP-dependent stimulation of insulin exocytosis, we have used capacitance measurements, protein-binding assays, and Western blot analysis. In insulin-secreting INS-1 cells overexpressing wild-type SNAP-25 (SNAP-25WT), rapid exocytosis was stimulated more than threefold by cAMP, similar to the situation in nontransfected cells. However, cAMP failed to potentiate rapid exocytosis in INS-1 cells overexpressing a truncated form of SNAP-25 (SNAP-251-197) or Botulinum neurotoxin A (BoNT/A). Close dissection of the exocytotic response revealed that the inability of cAMP to stimulate exocytosis in the presence of a truncated SNAP-25 was confined to the release of primed LDCVs within the readily releasable pool, especially from the immediately releasable pool, whereas cAMP enhanced mobilization of granules from the reserve pool in both SNAP-251-197( P < 0.01) and SNAP-25WT( P < 0.05) cells. This was supported by hormone release measurements. Augmentation of the immediately releasable pool by cAMP has been suggested to act through the cAMP-GEFII-dependent, PKA-independent pathway. Indeed, we were able to verify an interaction between SNAP-25 with both cAMP-GEFII and RIM2, two proteins involved in the PKA-independent pathway. Thus we hypothesize that SNAP-25 is a necessary partner in the complex mediating cAMP-enhanced rapid exocytosis in insulin-secreting cells.
Metabolic effects of dietary cholesterol in an animal model of insulin resistance and hepatic steatosis
Authors: Basciano Heather , Miller Abigale E. , Naples Mark , Baker Christopher , Kohen Rita , Xu Elaine , Su Qiaozhu , Allister Emma M. , Wheeler Michael B. , Adeli Khosrow
Published: June 2009

Although the atherogenic role of dietary cholesterol has been well established, its diabetogenic potential and associated metabolic disturbances have not been reported. Diet-induced hamster models of insulin resistance and dyslipidemia were employed to determine lipogenic and diabetogenic effects of dietary cholesterol. Metabolic studies were conducted in hamsters fed diets rich in fructose (40%), fat (30%), and cholesterol (0.05–0.25%) (FFC) and other test diets. Short-term feeding of the FFC diet induced insulin resistance, glucose intolerance, hypertriglyceridemia, and hypercholesterolemia. Prolonged feeding (6–22 wk) of the FFC diet led to severe hepatic steatosis, glucose intolerance, and mild increases in fasting blood glucose, suggesting progression toward type 2 diabetes, but did not induce β-cell dysfunction. Metabolic changes induced by the diet, including dyslipidemia and insulin resistance, were cholesterol concentration dependent and were only markedly induced on a high-fructose and high-fat dietary background. There were significant increases in hepatic and plasma triglyceride with FFC feeding, likely due to a 10- to 15-fold induction of hepatic stearoyl-CoA desaturase compared with chow levels ( P < 0.03). Hepatic insulin resistance was evident based on reduced tyrosine phosphorylation of the insulin receptor-β, IRS-1, and IRS-2 as well as increased protein mass of protein tyrosine phosphatase 1B. Interestingly, nuclear liver X receptor (LXR) target genes such as ABCA1 were upregulated on the FFC diet, and dietary supplementation with an LXR agonist (instead of dietary cholesterol) worsened dyslipidemia, glucose intolerance, and upregulation of target mRNA and proteins similar to that of dietary cholesterol. In summary, these data clearly implicate dietary cholesterol, synergistically acting with dietary fat and fructose, as a major determinant of the severity of metabolic disturbances in the hamster model. Dietary cholesterol appears to induce hepatic cholesterol ester and triglyceride accumulation, and diet-induced LXR activation (via cholesterol-derived oxysterols) may possibly be one key underlying mechanism.
Shock-induced stress induces loss of microvascular endothelial Tie2 in the kidney which is not associated with reduced glomerular barrier function
Authors: van Meurs Matijs , Kurniati Neng F. , Wulfert Francis M. , Asgeirsdottir Sigridur A. , de Graaf Inge A. , Satchell Simon C. , Mathieson Peter W. , Jongman Rianne M. , Kümpers Philipp , Zijlstra Jan G. , Heeringa Peter , Molema Grietje
Published: June 2009

Both hemorrhagic shock and endotoxemia induce a pronounced vascular activation in the kidney which coincides with albuminuria and glomerular barrier dysfunction. We hypothesized that changes in Tie2, a vascular restricted receptor tyrosine kinase shown to control microvascular integrity and endothelial inflammation, underlie this loss of glomerular barrier function. In healthy murine and human kidney, Tie2 is heterogeneously expressed in all microvascular beds, although to different extents. In mice subjected to hemorrhagic and septic shock, Tie2 mRNA and protein were rapidly, and temporarily, lost from the renal microvasculature, and normalized within 24 h after initiation of the shock insult. The loss of Tie2 protein could not be attributed to shedding as both in mice and healthy volunteers subjected to endotoxemia, sTie2 levels in the systemic circulation did not change. In an attempt to identify the molecular control of Tie2, we activated glomerular endothelial cell cultures and human kidney slices in vitro with LPS or TNF-α, but did not observe a change in Tie2 mRNA levels. In parallel to the loss of Tie2 in vivo, an overt influx of neutrophils in the glomerular compartment, which coincided with proteinuria, was seen. As neutrophil-endothelial cell interactions may play a role in endothelial adaptation to shock, and these effects cannot be mimicked in vitro, we depleted neutrophils before shock induction. While this neutrophil depletion abolished proteinuria, Tie2 was not rescued, implying that Tie2 may not be a major factor controlling maintenance of the glomerular filtration barrier in this model.
Glycinergic Inhibition Creates a Form of Auditory Spectral Integration in Nuclei of the Lateral Lemniscus
Authors: Peterson Diana Coomes , Nataraj Kiran , Wenstrup Jeffrey
Published: June 2009

For analyses of complex sounds, many neurons integrate information across different spectral elements via suppressive effects that are distant from the neurons' excitatory tuning. In the mustached bat, suppression evoked by sounds within the first sonar harmonic (23–30 kHz) or in the subsonar band (<23 kHz) alters responsiveness to the higher best frequencies of many neurons. This study examined features and mechanisms associated with low-frequency (LF) suppression among neurons of the lateral lemniscal nuclei (NLL). We obtained extracellular recordings from neurons in the intermediate and ventral nuclei of the lateral lemniscus, observing different forms of LF suppression related to the two above-cited frequency bands. To understand the mechanisms underlying this suppression in NLL neurons, we examined the roles of glycinergic and GABAergic input through local microiontophoretic application of strychnine, an antagonist to glycine receptors (GlyRs), or bicuculline, an antagonist to γ-aminobutyric acid type A receptors (GABAARs). With blockade of GABAARs, neurons showed an increase in firing rate to best frequency (BF) and/or LF tones but retained LF suppression of BF sounds. For neurons that displayed LF suppression tuned to 23–30 kHz, the suppression was eliminated or nearly eliminated by GlyR blockade. In contrast, GABAAR blockade did not eliminate nor had any consistent effect on suppression tuned to these frequencies. We conclude that LF suppression tuned in the 23- to 30-kHz range results from neuronal inhibition within the NLL via glycinergic inputs. For neurons displaying suppression tuned <23 kHz, neither GlyR nor GABAR blockade altered LF suppression. We conclude that such suppression originates at a lower auditory level, perhaps a result of cochlear mechanisms. These findings demonstrate that neuronal interactions within NLL create a particular form of LF suppression that contributes to the analysis of complex acoustic signals.
Spectral Properties of Induced and Evoked Gamma Oscillations in Human Early Visual Cortex to Moving and Stationary Stimuli
Authors: Swettenham J. B. , Muthukumaraswamy S. D. , Singh K. D.
Published: June 2009

In two experiments, magnetoencephalography (MEG) was used to investigate the effects of motion on gamma oscillations in human early visual cortex. When presented centrally, but not peripherally, stationary and moving gratings elicited several evoked and induced response components in early visual cortex. Time-frequency analysis revealed two nonphase locked gamma power increases—an initial, rapidly adapting response and one sustained throughout stimulus presentation and varying in frequency across observers from 28 to 64 Hz. Stimulus motion raised the sustained gamma oscillation frequency by a mean of ∼10 Hz. The largest motion-induced frequency increases were in those observers with the lowest gamma response frequencies for stationary stimuli, suggesting a possible saturation mechanism. Moderate gamma amplitude increases to moving versus stationary stimuli were also observed but were not correlated with the magnitude of the frequency increase. At the same site in visual cortex, sustained alpha/beta power reductions and an onset evoked response were observed, but these effects did not change significantly with the presence of motion and did not correlate with the magnitude of gamma power changes. These findings suggest that early visual areas encode moving and stationary percepts via activity at higher and lower gamma frequencies, respectively.
Skeletal muscle gene expression after myostatin knockout in mature mice
Authors: Welle Stephen , Cardillo Andrew , Zanche Michelle , Tawil Rabi
Published: June 2009

There is much interest in developing anti-myostatin agents to reverse or prevent muscle atrophy in adults, so it is important to characterize the effects of reducing myostatin activity after normal muscle development. For assessment of the effect of loss of myostatin signaling on gene expression in muscle, RNA from mice with postdevelopmental myostatin knockout was analyzed with oligonucleotide microarrays. Myostatin was undetectable in muscle within 2 wk after Cre recombinase activation in 4-month-old male mice with floxed myostatin genes. Three months after myostatin depletion, muscle mass had increased 26% (vs. 2% after induction of Cre activity in mice with normal myostatin genes), at which time the expression of several hundred genes differed in knockout and control mice at nominal P < 0.01. In contrast to previously reported effects of constitutive myostatin knockout, postdevelopmental knockout did not downregulate expression of genes encoding slow isoforms of contractile proteins or genes encoding proteins involved in energy metabolism. Several collagen genes were expressed at 20–50% lower levels in the myostatin-deficient muscles, which had ∼25% less collagen than normal muscles as reflected by hydroxyproline content. Most of the other genes affected by myostatin depletion have not been previously linked to myostatin signaling. Gene set enrichment analysis suggested that Smads are not the only transcription factors with reduced activity after myostatin depletion. These data reinforce other evidence that myostatin regulates collagen production in muscle and demonstrate that many of the previously reported effects of constitutive myostatin deficiency do not occur when myostatin is knocked out in mature muscles.
Targeting Angiogenesis via a c-Myc/Hypoxia-Inducible Factor-1α–Dependent Pathway in Multiple Myeloma
Authors: Zhang Jing , Sattler Martin , Tonon Giovanni , Grabher Clemens , Lababidi Samir , Zimmerhackl Alexander , Raab Marc S. , Vallet Sonia , Zhou Yiming , Cartron Marie-Astrid , Hideshima Teru , Tai Yu-Tzu , Chauhan Dharminder , Anderson Kenneth C. , Podar Klaus
Published: June 2009

Bone marrow angiogenesis is associated with multiple myeloma (MM) progression. Here, we report high constitutive hypoxia-inducible factor-1α (Hif-1α) expression in MM cells, which is associated with oncogenic c-Myc. A drug screen for anti-MM agents that decrease Hif-1α and c-Myc levels identified a variety of compounds, including bortezomib, lenalidomide, enzastaurin, and adaphostin. Functionally, based on transient knockdowns and overexpression, our data delineate a c-Myc/Hif-1α–dependent pathway mediating vascular endothelial growth factor production and secretion. The antiangiogenic activity of our tool compound, adaphostin, was subsequently shown in a zebrafish model and translated into a preclinical in vitro and in vivo model of MM in the bone marrow milieu. Our data, therefore, identify Hif-1α as a novel molecular target in MM and add another facet to anti-MM drug activity. [Cancer Res 2009;69(12):5082–90]
Long-term Cultures of Bone Marrow–Derived Human Mesenchymal Stem Cells Frequently Undergo Spontaneous Malignant Transformation
Authors: Røsland Gro Vatne , Svendsen Agnete , Torsvik Anja , Sobala Ewa , McCormack Emmet , Immervoll Heike , Mysliwietz Josef , Tonn Joerg-Christian , Goldbrunner Roland , Lønning Per Eystein , Bjerkvig Rolf , Schichor Christian
Published: June 2009

Human mesenchymal stem cells (hMSC) aid in tissue maintenance and repair by differentiating into specialized cell types. Due to this ability, hMSC are currently being evaluated for cell-based therapies of tissue injury and degenerative diseases. However, extensive expansion ex vivo is a prerequisite to obtain the cell numbers required for human cell-based therapy protocols. Recent studies indicate that hMSC may contribute to cancer development and progression either by acting as cancer-initiating cells or through interactions with stromal elements. If spontaneous transformation ex vivo occurs, this may jeopardize the use of hMSC as therapeutic tools. Whereas murine MSC readily undergo spontaneous transformation, there are conflicting reports about spontaneous transformation of hMSC. We have addressed this controversy in a two-center study by growing bone marrow–derived hMSC in long-term cultures (5–106 weeks). We report for the first time spontaneous malignant transformation to occur in 45.8% (11 of 24) of these cultures. In comparison with hMSC, the transformed mesenchymal cells (TMC) showed a significantly increased proliferation rate and altered morphology and phenotype. In contrast to hMSC, TMC grew well in soft agar assays and were unable to undergo complete differentiation. Importantly, TMC were highly tumorigenic, causing multiple fast-growing lung deposits when injected into immunodeficient mice. We conclude that spontaneous malignant transformation may represent a biohazard in long-term ex vivo expansion of hMSC. On the other hand, this spontaneous transformation process may represent a unique model for studying molecular pathways initiating malignant transformation of hMSC. [Cancer Res 2009;69(13):5331–9]
ROC1/RBX1 E3 Ubiquitin Ligase Silencing Suppresses Tumor Cell Growth via Sequential Induction of G2-M Arrest, Apoptosis, and Senescence
Authors: Jia Lijun , Soengas Maria S. , Sun Yi
Published: June 2009

Regulator of Cullins-1 (ROC1) or Ring Box Protein-1 (RBX1) is a RING component of SCF (Skp-1, cullins, F-box proteins) E3 ubiquitin ligases, which regulate diverse cellular processes by targeting a variety of substrates for degradation. However, little is known about the role of ROC1 in human cancer. Here, we report that ROC1 is ubiquitously overexpressed in primary human tumor tissues and human cancer cell lines. ROC1 silencing by siRNA significantly inhibited the growth of multiple human cancer cell lines via induction of senescence and apoptosis as well as G2-M arrest. Senescence induction is coupled with DNA damage in p53/p21- and p16/pRB-independent manners. Apoptosis is associated with accumulation of Puma and reduction of Bcl-2, Mcl-1, and survivin; and G2-M arrest is associated with accumulation of 14-3-3σ and elimination of cyclin B1 and Cdc2. In U87 glioblastoma cells, these phenotypic changes occur sequentially upon ROC1 silencing, starting with G2-M arrest, followed by apoptosis and senescence. Thus, ROC1 silencing triggers multiple death and growth arrest pathways to effectively suppress tumor cell growth, suggesting that ROC1 may serve as a potential anticancer target. [Cancer Res 2009;69(12):4974–82]
Harnessing Competing Endocytic Pathways for Overcoming the Tumor-Blood Barrier: Magnetic Resonance Imaging and Near-Infrared Imaging of Bifunctional Contrast Media
Authors: Migalovich Helena Sheikhet , Kalchenko Vyacheslav , Nevo Nava , Meir Gila , Kohen Fortune , Neeman Michal
Published: June 2009

Ovarian cancer is the most lethal gynecologic malignancy, often diagnosed at advanced stage leading to poor prognosis. In the study reported here, magnetic resonance imaging and near-infrared reflectance imaging were applied for in vivo analysis of two competing endocytic pathways affecting retention of bifunctional daidzein-bovine serum albumin (BSA)–based contrast media by human epithelial ovarian carcinoma cells. Suppression of caveolae-mediated uptake using nystatin or by BSA competition significantly enhanced daidzein-BSA-GdDTPA/CyTE777 uptake by tumor cells in vitro. In vivo, perivascular myofibroblasts generated an effective perivascular barrier excluding delivery of BSA-GdDTPA/CyTE777 to tumor cells. The ability to manipulate caveolae-mediated sequestration of albumin by perivascular tumor myofibroblasts allowed us to effectively overcome this tumor-stroma barrier, increasing delivery of daidzein-BSA-GdDTPA/CyTE777 to the tumor cells in tumor xenografts. Thus, both in vitro and in vivo, endocytosis of daidzein-BSA-GdDTPA/CyTE777 by ovarian carcinoma cells was augmented by albumin or by nystatin. In view of the cardinal role of albumin in affecting the availability and pharmacokinetics of drugs, this approach could potentially also facilitate the delivery of therapeutics and contrast media to tumor cells. [Cancer Res 2009;69(13):5610–7]
Subcellular Localization of Cyclic AMP-Responsive Element Binding Protein-Regulated Transcription Coactivator 2 Provides a Link between Obesity and Breast Cancer in Postmenopausal Women
Authors: Brown Kristy A. , McInnes Kerry J. , Hunger Nicole I. , Oakhill Jonathan S. , Steinberg Gregory R. , Simpson Evan R.
Published: June 2009

Epidemiologic evidence supports a correlation between obesity and breast cancer in women. AMP-activated protein kinase plays an important role in energy homeostasis and inhibits the actions of cyclic AMP-responsive element binding protein-regulated transcription coactivator 2 (CRTC2). In postmenopausal women, the cyclic AMP-responsive element binding protein-dependent regulation of aromatase is a determinant of breast tumor formation through local production of estrogens. The present work aimed to examine the effect of adipokines on aromatase expression and identify additional mechanisms by which prostaglandin E2 causes increased aromatase expression in human breast adipose stromal cells. Treatment of human adipose stromal cells with forskolin and phorbol 12-myristate 13-acetate (PMA), to mimic prostaglandin E2, resulted in nuclear translocation of CRTC2. Aromatase promoter II (PII) activity assays showed that CRTC2 in addition to forskolin/PMA treatment significantly increased PII-induced activity. CRTC2 binding to PII was examined by chromatin immunoprecipitation, and forskolin/PMA treatment was associated with increased binding to PII. Treatment of human adipose stromal cells with leptin significantly up-regulated aromatase expression associated with nuclear translocation of CRTC2 and increased binding of CRTC2 to PII. Adiponectin treatment significantly decreased forskolin/PMA-stimulated aromatase expression, consistent with the decreased nuclear translocation of CRTC2 and the decreased binding of CRTC2 to PII. The expression and activity of the AMP-activated protein kinase LKB1 was examined and found to be significantly decreased following either forskolin/PMA or leptin treatment. In contrast, adiponectin significantly increased LKB1 expression and activity. In conclusion, the regulation of aromatase by CRTC2, in response to the altered hormonal milieu associated with menopause and obesity, provides a critical link between obesity and breast cancer. [Cancer Res 2009;69(13):5392–9]
Infrequent Detection of Germline Allele-Specific Expression of TGFBR1 in Lymphoblasts and Tissues of Colon Cancer Patients
Authors: Guda Kishore , Natale Leanna , Lutterbaugh James , Wiesner Georgia L. , Lewis Susan , Tanner Stephan M. , Tomsic Jerneja , Valle Laura , de la Chapelle Albert , Elston Robert C. , Willis Joseph , Markowitz Sanford D.
Published: June 2009

Recently, germline allele-specific expression (ASE) of the gene encoding for transforming growth factor-β type I receptor (TGFBR1) has been proposed to be a major risk factor for cancer predisposition in the colon. Germline ASE results in a lowered expression of one of the TGFBR1 alleles (>1.5-fold), and was shown to occur in ∼20% of informative familial and sporadic colorectal cancer (CRC) cases. In the present study, using the highly quantitative pyrosequencing technique, we estimated the frequency of ASE in TGFBR1 in a cohort of affected individuals from familial clusters of advanced colon neoplasias (cancers and adenomas with high-grade dysplasia), and also from a cohort of individuals with sporadic CRCs. Cases were considered positive for the presence of ASE if demonstrating an allelic expression ratio <0.67 or >1.5. Using RNA derived from lymphoblastoid cell lines, we find that of 46 informative Caucasian advanced colon neoplasia cases with a family history, only 2 individuals display a modest ASE, with allelic ratios of 1.65 and 1.73, respectively. Given that ASE of TGFBR1, if present, would likely be more pronounced in the colon compared with other tissues, we additionally determined the allele ratios of TGFBR1 in the RNA derived from normal-appearing colonic mucosa of sporadic CRC cases. We, however, found no evidence of ASE in any of 44 informative sporadic cases analyzed. Taken together, we find that germline ASE of TGFBR1, as assayed in lymphoblastoid and colon epithelial cells of colon cancer patients, is a relatively rare event. [Cancer Res 2009;69(12):4959–61]
Cancer Genome Sequencing—An Interim Analysis
Authors: Fox Edward J. , Salk Jesse J. , Loeb Lawrence A.
Published: June 2009

With the publishing of the first complete whole genome of a human cancer and its paired normal, we have passed a key milestone in the cancer genome sequencing strategy. The generation of such data will, thanks to technical advances, soon become commonplace. As a significant number of proof-of-concept studies have been published, it is important to analyze now the likely implications of these data and how this information might frame cancer research in the near future. The diversity of genes mutated within individual tumor types, the most striking feature of all studies reported to date, challenges gene-centric models of tumorigenesis. Although cancer genome sequencing will revolutionize certain aspects of personalized care, the value of these studies in facilitating the development of new therapies, their primary goal, seems less promising. Most significantly, however, the cancer genome sequencing strategy, as currently applied, fails to characterize the most relevant genomic features of cancer—the mutational heterogeneity within individual tumors. [Cancer Res 2009;69(12):4948–4950]
Proposal and Validation of Prognostic Scoring Systems for IgG and IgA Monoclonal Gammopathies of Undetermined Significance
Authors: Rossi Francesca , Petrucci Maria Teresa , Guffanti Andrea , Marcheselli Luigi , Rossi Davide , Callea Vincenzo , Vincenzo Federico , De Muro Marianna , Baraldi Alessandra , Villani Oreste , Musto Pellegrino , Bacigalupo Andrea , Gaidano Gianluca , Avvisati Giuseppe , Goldaniga Maria , DePaoli Lorenzo , Baldini Luca
Published: June 2009

Purpose: The presenting clinico-hematologic features of 1,283 patients with IgG and IgA monoclonal gammopathies of undetermined significance (MGUS) were correlated with the frequency of evolution into multiple myeloma (MM).

Experimental Design: Two IgG MGUS populations were evaluated: a training sample (553 patients) and a test sample (378 patients); the IgA MGUS population consisted of 352 patients.

Results: Forty-seven of the 553 training group patients and 22 of 378 test group IgG patients developed MM after a median follow-up of 6.7 and 3.6 years, respectively. Multivariate analysis showed that serum monoclonal component (MC) levels of ≤1.5 g/dL, the absence of light-chain proteinuria and normal serum polyclonal immunoglobulin levels defined a prognostically favorable subset of patients, and could be used to stratify the patients into three groups at different 10-year risk of evolution (hazard ratio, 1.0, 5.04, 11.2; P < 0.001). This scoring system was validated in the test sample. Thirty of the 352 IgA patients developed MM after a median follow-up of 4.8 years, and multivariate analysis showed that hemoglobin levels of <12.5 g/dL and reduced serum polyclonal immunoglobulin correlated with progression. A pooled statistical analysis of all of the patients confirmed the validity of Mayo Clinic risk model showing that IgA class, serum MC levels, and light-chain proteinuria are the most important variables correlated with disease progression.

Conclusions: Using simple variables, we validated a prognostic model for IgG MGUS. Among the IgA cases, the possible prognostic role of hemoglobin emerged in addition to a decrease in normal immunoglobulin levels.
Suppression of Androgen-Independent Prostate Cancer Cell Aggressiveness by FTY720: Validating Runx2 as a Potential Antimetastatic Drug Screening Platform
Authors: Chua Chee-Wai , Chiu Yung-Tuen , Yuen Hiu-Fung , Chan Kwok-Wah , Man Kwan , Wang Xianghong , Ling Ming-Tat , Wong Yong-Chuan
Published: June 2009

Purpose: Previously, FTY720 was found to possess potent anticancer effects on various types of cancer. In the present study, we aimed to first verify the role of Runx2 in prostate cancer progression and metastasis, and, subsequently, assessed if FTY720 could modulate Runx2 expression, thus interfering downstream events regulated by this protein.

Experimental Design: First, the association between Runx2 and prostate cancer progression was assessed using localized prostate cancer specimens and mechanistic investigation of Runx2-induced cancer aggressiveness was then carried out. Subsequently, the effect of FTY720 on Runx2 expression and transcriptional activity was investigated using PC-3 cells, which highly expressed Runx2 protein. Last, the involvement of Runx2 in FTY720-induced anticancer effects was evaluated by modulating Runx2 expression in various prostate cancer cell lines.

Results: Runx2 nuclear expression was found to be up-regulated in prostate cancer and its expression could be used as a predictor of metastasis in prostate cancer. Further mechanistic studies indicated that Runx2 accelerated prostate cancer aggressiveness through promotion of cadherin switching, invasion toward collagen I, and Akt activation. Subsequently, we found that FTY720 treatment down-regulated Runx2 expression and its transcriptional activity, as well as inhibited its regulated downstream events. More importantly, silencing Runx2 in PC-3 enhanced FTY720-induced anticancer effects as well as cell viability inhibition, whereas overexpressing Runx2 in 22Rv1 that expressed very low endogenous Runx2 protein conferred resistance in the same events.

Conclusion: This study provided a novel mechanism for the anticancer effect of FTY720 on advanced prostate cancer, thus highlighting the therapeutic potential of this drug in treating this disease.
Stereotyped B-Cell Receptor Is an Independent Risk Factor of Chronic Lymphocytic Leukemia Transformation to Richter Syndrome
Authors: Rossi Davide , Spina Valeria , Cerri Michaela , Rasi Silvia , Deambrogi Clara , De Paoli Lorenzo , Laurenti Luca , Maffei Rossana , Forconi Francesco , Bertoni Francesco , Zucca Emanuele , Agostinelli Claudio , Cabras Antonello , Lucioni Marco , Martini Maurizio , Magni Michele , Deaglio Silvia , Ladetto Marco , Nomdedeu Josep F. , Besson Caroline , Ramponi Antonio , Canzonieri Vincenzo , Paulli Marco , Marasca Roberto , Larocca Luigi M. , Carbone Antonino , Pileri Stefano A. , Gattei Valter , Gaidano Gianluca
Published: June 2009

Purpose: Few biological prognosticators are useful for prediction of Richter syndrome (RS), representing the transformation of chronic lymphocytic leukemia (CLL) to aggressive lymphoma. Stereotyped B-cell receptors (BCR) may have prognostic effect in CLL progression. We tested the prognostic effect of stereotyped BCR for predicting RS transformation.

Experimental Design: The prevalence of stereotyped BCR was compared in RS (n = 69) versus nontransformed CLL (n = 714) by a case-control analysis. Subsequently, the effect of stereotyped BCR at CLL diagnosis on risk of RS transformation was actuarially assessed in a consecutive CLL series (n = 753).

Results: RS (n = 69) displayed a higher prevalence of stereotyped BCR (P < 0.001) compared with nontransformed CLL. The actuarial risk of RS transformation was significantly higher in CLL carrying stereotyped BCR (P < 0.001). Among BCR subsets most represented in CLL, subset 8 using IGHV4-39/IGHD6-13/IGHJ5 carried the highest risk of RS transformation [hazard ratio (HR), 24.50; P < 0.001]. Multivariate analysis selected stereotyped BCR (HR, 3.33; P = 0.001) and IGHV4-39 usage (HR, 4.03; P = 0.004) as independent predictors of RS transformation. The combination of IGHV4-39 usage and stereotyped BCR in the same patient identified CLL with a very high risk of RS transformation (5-year risk, 68.7%). The risk carried by stereotyped BCR and IGHV4-39 usage was specific for RS transformation and had no effect on CLL progression without transformation.

Conclusions: Analysis of BCR features may help identify CLL patients at risk of RS. A close monitoring and a careful biopsy policy may help early recognition of RS in CLL patients using stereotyped BCR, particularly if combined with IGHV4-39.
Antitumor Activity of NK012 Combined with Cisplatin against Small Cell Lung Cancer and Intestinal Mucosal Changes in Tumor-Bearing Mouse after Treatment
Authors: Nagano Tatsuya , Yasunaga Masahiro , Goto Koichi , Kenmotsu Hirotsugu , Koga Yoshikatsu , Kuroda Jun-ichiro , Nishimura Yoshihiro , Sugino Takashi , Nishiwaki Yutaka , Matsumura Yasuhiro
Published: June 2009

Purpose: To investigate the advantages of treatment with the SN-38–incorporating polymeric micelles NK012 over CPT-11 in combination with cisplatin [cis-dichlorodiammineplatinum (II) (CDDP)] in mice bearing a small cell lung cancer xenograft in terms of antitumor activity and toxicity, particularly intestinal toxicity.

Experimental Design: Cytotoxic effects were evaluated in human small cell lung cancer cell lines [H69, H82, and vascular endothelial growth factor (VEGF)–secreting cells (SBC-3/VEGF and its mock transfectant SBC-3/Neo)]. In vivo antitumor effects were evaluated in SBC-3/Neo–bearing and SBC-3/VEGF–bearing mice after NK012/CDDP or CPT-11/CDDP administration on days 0, 7, and 14. Drug distribution was analyzed by high-performance liquid chromatography or fluorescence microscopy, and the small intestine was pathologically examined.

Results: The in vitro growth-inhibitory effects of NK012 were 198- to 532-fold more potent than those of CPT-11. A significant difference in the relative tumor volume on day 30 was found between NK012/CDDP and CPT-11/CDDP treatments (P = 0.0058). Inflammatory changes in the small intestinal mucosa were rare in all NK012-treated mice but were commonly observed in CPT-11–treated mice. Moreover, a large amount of CPT-11 was excreted into the feces and high CPT-11 concentration was detected in the small intestinal epithelium. On the other hand, a small amount of NK012 was found in the feces and NK012 was weakly and uniformly distributed in the mucosal interstitium.

Conclusions: NK012/CDDP combination may be a promising candidate regimen against lung cancer without severe diarrhea toxicity and therefore warrants further clinical evaluation.
Immune-Mediated Antitumor Activity of Reovirus Is Required for Therapy and Is Independent of Direct Viral Oncolysis and Replication
Authors: Prestwich Robin J. , Ilett Elizabeth J. , Errington Fiona , Diaz Rosa M. , Steele Lynette P. , Kottke Tim , Thompson Jill , Galivo Feorillo , Harrington Kevin J. , Pandha Hardev S. , Selby Peter J. , Vile Richard G. , Melcher Alan A.
Published: June 2009

Purpose: Reovirus is a naturally occurring oncolytic virus in clinical trials. Although tumor infection by reovirus can generate adaptive antitumor immunity, its therapeutic importance versus direct viral oncolysis is undefined. This study addresses the requirement for viral oncolysis and replication, and the relative importance of antitumor immunity and direct oncolysis in therapy.

Experimental Design: Nonantigen specific T cells loaded with reovirus were delivered i.v. to C57BL/6 and severe combined immunodeficient mice bearing lymph node and splenic metastases from the murine melanoma, B16ova, with assessment of viral replication, metastatic clearance by tumor colony outgrowth, and immune priming. Human cytotoxic lymphocyte priming assays were done with dendritic cells loaded with Mel888 cells before the addition of reovirus.

Results: B16ova was resistant to direct oncolysis in vitro, and failed to support reovirus replication in vitro or in vivo. Nevertheless, reovirus purged lymph node and splenic metastases in C57BL/6 mice and generated antitumor immunity. In contrast, reovirus failed to reduce tumor burden in severe combined immunodeficient mice bearing either B16ova or reovirus-sensitive B16tk metastases. In the human system, reovirus acted solely as an adjuvant when added to dendritic cells already loaded with Mel888, supporting priming of specific antitumor cytotoxic lymphocyte, in the absence of significant direct tumor oncolysis; UV-treated nonreplicating reovirus was similarly immunogenic.

Conclusion: The immune response is critical in mediating the efficacy of reovirus, and does not depend upon direct viral oncolysis or replication. The findings are of direct relevance to fulfilling the potential of this novel anticancer agent.
Rapid Immune Recovery and Graft-versus-Host Disease–like Engraftment Syndrome following Adoptive Transfer of Costimulated Autologous T Cells
Authors: Rapoport Aaron P. , Stadtmauer Edward A. , Aqui Nicole , Vogl Dan , Chew Anne , Fang Hong-Bin , Janofsky Stephen , Yager Kelly , Veloso Elizabeth , Zheng Zhaohui , Milliron Todd , Westphal Sandra , Cotte Julio , Huynh Hong , Cannon Andrea , Yanovich Saul , Akpek Gorgun , Tan Ming , Virts Kristen , Ruehle Kathleen , Harris Carolynn , Philip Sunita , Vonderheide Robert H. , Levine Bruce L. , June Carl H.
Published: June 2009

Purpose: Previously, we showed that adoptive transfer of in vivo vaccine-primed and ex vivo (anti-CD3/anti-CD28) costimulated autologous T cells (ex-T) at day +12 after transplant increased CD4 and CD8 T-cell counts at day +42 and augmented vaccine-specific immune responses in patients with myeloma. Here, we investigated the safety and kinetics of T-cell recovery after infusing ex-T at day +2 after transplant.

Experimental Design: In this phase I/II two-arm clinical trial, 50 patients with myeloma received autografts after high-dose melphalan followed by infusions of ex-T at day +2 after transplant. Patients also received pretransplant and posttransplant immunizations using a pneumococcal conjugate vaccine only (arm B; n = 24) or the pneumococcal conjugate vaccine plus an HLA-A2–restricted multipeptide vaccine for HLA-A2+ patients (arm A; n = 26).

Results: The mean number of T cells infused was 4.26 × 1010 (range, 1.59-5.0). At day 14 after transplant, the median CD3, CD4, and CD8 counts were 4,198, 1,545, and 2,858 cells/μL, respectively. Interleukin (IL)-6 and IL-15 levels increased early after transplant and IL-15 levels correlated significantly to day 14 T-cell counts. Robust vaccine-specific B- and T-cell responses were generated. T-cell infusions were well tolerated with no effect on hematopoietic recovery. Eight patients (16%) developed a T-cell “engraftment syndrome” characterized by diarrhea and fever that was clinically and histopathologically indistinguishable from grade 1 to 3 acute graft-versus-host disease (GVHD) of the gastrointestinal tract (seven patients) and/or grade 1 to 2 cutaneous GVHD (four patients).

Conclusions: Adoptive T-cell transfers achieve robust T-cell recovery early after transplant and induce moderate-to-severe autologous GVHD in a subset of patients.
Inflexinol inhibits colon cancer cell growth through inhibition of nuclear factor-κB activity via direct interaction with p50
Authors: Ban Jung Ok , Oh Ju Hoon , Hwang Bang Yeon , Moon Dong Cheul , Jeong Heon-Sang , Lee Seram , Kim Soyoun , Lee Hyosung , Kim Kyung-Bo , Han Sang Bae , Hong Jin Tae
Published: June 2009

Kaurane diterpene compounds have been known to be cytotoxic against several cancer cells through inhibition of nuclear factor-κB (NF-κB) activity. Here, we showed that inflexinol, a novel kaurane diterpene compound, inhibited the activity of NF-κB and its target gene expression as well as cancer cell growth through induction of apoptotic cell death in vitro and in vivo. These inhibitory effects on NF-κB activity and on cancer cell growth were suppressed by the reducing agents DTT and glutathione and were abrogated in the cells transfected with mutant p50 (C62S). Sol-gel biochip and surface plasmon resonance analysis showed that inflexinol binds to the p50 subunit of NF-κB. These results suggest that inflexinol inhibits colon cancer cell growth via induction of apoptotic cell death through inactivation of NF-κB by a direct modification of cysteine residue in the p50 subunit of NF-κB. [Mol Cancer Ther 2009;8(6):1613–24]
Consensus transcriptome signature of perineural invasion in pancreatic carcinoma
Authors: Abiatari Ivane , DeOliveira Tiago , Kerkadze Vachtang , Schwager Christian , Esposito Irene , Giese Nathalia A. , Huber Peter , Bergman Frank , Abdollahi Amir , Friess Helmut , Kleeff Jörg
Published: June 2009

Perineural invasion, the growth of tumor cells along nerves, is a key feature of pancreatic cancer. The cardinal symptom of pancreatic cancer, abdominal pain often radiating to the back, as well as the high frequency of local tumor recurrence following resection are both attributed to the unique ability of pancreatic tumor cells to invade the neuronal system. The molecular mechanisms underlying the neuroaffinity of pancreatic tumors are not completely understood. In this study, we developed a novel method to monitor ex vivo perineural invasion into surgically resected rat vagal nerves by different human pancreatic tumor cell lines. Genome-wide transcriptional analyses were employed to identify the consensus set of genes differentially regulated in all highly nerve-invasive (nerve invasion passage 3) versus less invasive (nerve invasion passage 0) pancreatic tumor cells. The critical involvement of kinesin family member 14 (KIF14) and Rho-GDP dissociation inhibitor β (ARHGDIβ) in perineural invasion was confirmed on RNA and protein levels in human pancreatic tumor specimens. We found significant up-regulation of KIF14 and ARHGDIβ mRNA levels in patients with pancreatic cancer, and both proteins were differentially expressed in tumor cells invading the perineural niche of pancreatic cancer patients as detected by immunohistochemistry. Moreover, functional knockdown of KIF14 and ARHGDIβ using small interfering RNA resulted in altered basal and/or perineural invasion of pancreatic tumor cells. Our work provides novel insights into the molecular determinants of perineural invasion in pancreatic cancer. The established nerve invasion model and the consensus signature of perineural invasion could be instrumental in the identification of novel therapeutic targets of pancreatic cancer as exemplified by KIF14 and ARHGDIβ. [Mol Cancer Ther 2009;8(6):1494–1504]
SHP and Sin3A expression are essential for adamantyl-substituted retinoid-related molecule–mediated nuclear factor-κB activation, c-Fos/c-Jun expression, and cellular apoptosis
Authors: Farhana Lulu , Dawson Marcia I. , Xu Liping , Dannenberg Jan-Hermen , Fontana Joseph A.
Published: June 2009

We previously found that the adamantyl-substituted retinoid-related molecules bind to the small heterodimer partner (SHP) as well as the Sin3A complex. In this report, we delineated the role of SHP and the Sin3A complex in 4-[3′-(1-adamantyl)-4′-hydroxyphenyl]-3-chlorocinnamic acid (3-Cl-AHPC)–mediated inhibition of cell growth and apoptosis. We examined the effect of loss of SHP and Sin3A expression in a number of cell types on 3-Cl-AHPC–mediated growth inhibition and apoptosis induction, 3-Cl-AHPC–mediated nuclear factor-κB (NF-κB) activation, and 3-Cl-AHPC–mediated increase in c-Fos and c-Jun expression. We found that loss of SHP or Sin3A expression, while blocking 3-Cl-AHPC–mediated apoptosis, had little effect on 3-Cl-AHPC inhibition of cellular proliferation. We have previously shown that 3-Cl-AHPC–mediated NF-κB activation is necessary for apoptosis induction. We have now shown that 3-Cl-AHPC–enhanced c-Fos and c-Jun expression is also essential for maximal 3-Cl-AHPC–mediated apoptosis. 3-Cl-AHPC induction of c-Fos and c-Jun expression as well as NF-κB activation was dependent on SHP protein levels. In turn, SHP levels are regulated by Sin3A because ablation of Sin3A resulted in a decrease in SHP expression. Thus, SHP and Sin3A play an important role in adamantyl-substituted retinoid-related induction of cellular apoptosis. [Mol Cancer Ther 2009;8(6):1625–35]
Luteinizing hormone-releasing hormone receptor–targeted deslorelin-docetaxel conjugate enhances efficacy of docetaxel in prostate cancer therapy
Authors: Sundaram Sneha , Durairaj Chandrasekar , Kadam Rajendra , Kompella Uday B.
Published: June 2009

Docetaxel, a chemotherapeutic agent currently used for improving survival of prostate cancer patients, suffers from low therapeutic index. The objective of this study was to prepare a new docetaxel derivative conjugated to deslorelin, a luteinizing hormone-releasing hormone (LHRH) superagonist, and to determine whether it enhances docetaxel potency in vitro and in vivo. Because docetaxel is not amenable for conjugation with peptides, we introduced a -COOH group in docetaxel, forming docetaxel-hemiglutarate, and subsequently conjugated this to serine in deslorelin, forming deslorelin-docetaxel. Fourier-transform IR, 1H-nuclear magnetic resonance, and liquid chromatography-mass spectrometry analyses confirmed deslorelin-docetaxel formation. Antiproliferative efficacy in LNCaP and PC-3 cell lines over 24, 48, and 72 hours exhibited the order deslorelin-docetaxel > docetaxel, whereas deslorelin alone had no effect, with deslorelin-docetaxel potency being 15-fold greater than docetaxel at 72 h. Further, cells pretreated with antisense oligonucleotide against LHRH receptor exhibited decreased deslorelin-docetaxel efficacy, without any change in docetaxel efficacy. Thus, deslorelin-docetaxel efficacy is likely mediated via LHRH receptor. Cell cycle analysis showed that docetaxel treatment led to arrest in G2-M phase, whereas deslorelin-docetaxel treatment allowed greater progression to apoptosis in both cell lines, with deslorelin-docetaxel exerting 5-fold greater apoptosis compared with docetaxel in prostate cancer cell lines. Antitumor efficacy studies in PC-3 prostate xenograft-bearing mice indicated the efficacy order deslorelin-docetaxel > docetaxel ≫ deslorelin > PBS, with deslorelin-docetaxel exerting ∼5.5-fold greater tumor growth inhibition than docetaxel alone. Thus, deslorelin-docetaxel prepared in this study retains pharmacologic effects of both docetaxel and deslorelin while enhancing the antiproliferative, apoptotic, and antitumor efficacy of docetaxel by several folds in prostate cancer therapy. [Mol Cancer Ther 2009;8(6):1655–65]