bluuebunny/crossref_metadata_2025 huggingface dataset:
Taxane-Mediated Antiangiogenesis
in Vitro
Authors: Ng Sylvia S. W. , Figg William D. , Sparreboom Alex
Published: September 2005

Paclitaxel (Taxol) and docetaxel (Taxotere) have been shown to inhibit angiogenesis at low concentrations that do not affect cancer cell proliferation. Here, we used rat aortic rings and human umbilical vein endothelial cells to evaluate the influence of their formulation vehicles Cremophor EL and polysorbate 80, as well as serum binding proteins on taxane-mediated antiangiogenesis. The data show that clinically relevant concentrations of the vehicles and binding proteins nullify the antiangiogenic activity of both taxanes. It is suggested that these agents may need to be used at much higher doses than anticipated for effective antiangiogenic chemotherapy.
Microarray Analysis Reveals Differences in Gene Expression of Circulating CD8+ T Cells in Melanoma Patients and Healthy Donors
Authors: Xu Tong , Shu Chen-Tsen , Purdom Elizabeth , Dang Demi , Ilsley Diane , Guo Yaqian , Weber Jeffrey , Holmes Susan P. , Lee Peter P.
Published: September 2005

Circulating T cells from many cancer patients are known to be dysfunctional and undergo spontaneous apoptosis. We used microarray technology to determine whether gene expression differences exist in T cells from melanoma patients versus healthy subjects, which may underlie these abnormalities. To maximize the resolution of our data, we sort purified CD8+ subsets and amplified the extracted RNA for microarray analysis. These analyses show subtle but statistically significant expression differences for 10 genes in T cells from melanoma patients versus healthy controls, which were additionally confirmed by quantitative real-time PCR analysis. Whereas none of these genes are members of the classical apoptosis pathways, several may be linked to apoptosis. To additionally investigate the significance of these 10 genes, we combined them into a classifier and found that they provide a much better discrimination between melanoma and healthy T cells as compared with a classifier built uniquely with classical apoptosis-related genes. These results suggest the possible engagement of an alternative apoptosis pathway in circulating T cells from cancer patients.
Stromal Cell-Derived Factor-1α Promotes Melanoma Cell Invasion across Basement Membranes Involving Stimulation of Membrane-Type 1 Matrix Metalloproteinase and Rho GTPase Activities
Authors: Bartolomé Rubén A. , Gálvez Beatriz G. , Longo Natividad , Baleux Françoise , van Muijen Goos N. P. , Sánchez-Mateos Paloma , Arroyo Alicia G. , Teixidó Joaquin
Published: September 2005

Tissue invasion by tumor cells involves their migration across basement membranes through activation of extracellular matrix degradation and cell motility mechanisms. Chemokines binding to their receptors provide chemotactic cues guiding cells to specific tissues and organs; they therefore could potentially participate in tumor cell dissemination. Melanoma cells express CXCR4, the receptor for the chemokine stromal cell-derived factor-1α (SDF-1α). Using Matrigel as a model, we show that SDF-1α promotes invasion of melanoma cells across basement membranes. Stimulation of membrane-type 1 matrix metalloproteinase (MT1-MMP) activity by SDF-1α was necessary for invasion, involving at least up-regulation in the expression of this metalloproteinase, as detected in the highly metastatic BLM melanoma cell line. Moreover, SDF-1α triggered the activation of the GTPases RhoA, Rac1, and Cdc42 on BLM cells, and expression of dominant-negative forms of RhoA and Rac1, but not Cdc42, substantially impaired the invasion of transfectants in response to SDF-1α, as well as the increase in MT1-MMP expression. Furthermore, CXCR4 expression on melanoma cells was notably augmented by transforming growth factor-β1, a Matrigel component, whereas anti-transforming growth factor-β antibodies inhibited increases in CXCR4 expression and melanoma cell invasion toward SDF-1α. The identification of SDF-1α as a potential stimulatory molecule for MT1-MMP as well as for RhoA and Rac1 activities during melanoma cell invasion, associated with an up-regulation in CXCR4 expression by interaction with basement membrane factors, could contribute to better knowledge of mechanisms stimulating melanoma cell dissemination.
Characterization of the Interaction of Ingenol 3-Angelate with Protein Kinase C
Authors: Kedei Noemi , Lundberg Daniel J. , Toth Attila , Welburn Peter , Garfield Susan H. , Blumberg Peter M.
Published: September 2005

Ingenol 3-angelate (I3A) is one of the active ingredients in Euphorbia peplus, which has been used in traditional medicine. Here, we report the initial characterization of I3A as a protein kinase C (PKC) ligand. I3A bound to PKC-α in the presence of phosphatidylserine with high affinity; however, under these assay conditions, little PKC isoform selectivity was observed. PKC isoforms did show different sensitivity and selectivity for down-regulation by I3A and phorbol 12-myristate 13-acetate (PMA) in WEHI-231, HOP-92, and Colo-205 cells. In all of the three cell types, I3A inhibited cell proliferation with somewhat lower potency than did PMA. In intact CHO-K1 cells, I3A was able to translocate different green fluorescent protein-tagged PKC isoforms, visualized by confocal microscopy, with equal or higher potency than PMA. PKC-δ in particular showed a different pattern of translocation in response to I3A and PMA. I3A induced a higher level of secretion of the inflammatory cytokine interleukin 6 compared with PMA in the WEHI-231 cells and displayed a marked biphasic dose-response curve for the induction. I3A was unable to cause the same extent of association of the C1b domain of PKC-δ with lipids, compared with PMA or the physiological regulator diacylglycerol, and was able to partially block the association induced by these agents, measured by surface plasmon resonance. The in vitro kinase activity of PKC-α induced by I3A was lower than that induced by PMA. The novel pattern of behavior of I3A makes it of great interest for further evaluation.
Marijuana Use and Risk of Oral Squamous Cell Carcinoma
Authors: Rosenblatt Karin A. , Daling Janet R. , Chen Chu , Sherman Karen J. , Schwartz Stephen M.
Published: September 2005

Previous laboratory investigations, case reports, and a hospital-based case-control study have suggested that marijuana use may be a risk factor for squamous cell head and neck cancer. We conducted a population-based case-control study to determine whether marijuana use is associated with the development of oral squamous cell carcinoma (OSCC). Case subjects (n = 407) were 18–65-year-old residents of three counties in western Washington State who were newly diagnosed with OSCC from 1985 through 1995. Control subjects (n = 615), who were similar to the cases with respect to age and sex, were selected from the general population using random-digit telephone dialing. Lifetime histories of marijuana use and exposure to known OSCC risk factors were ascertained using a structured questionnaire. Information on genetic polymorphisms in glutathione S-transferase enzymes was obtained from assays on participant DNA. Odds ratios for associations with features of marijuana use were adjusted for sex, education, birth year, alcohol consumption, and cigarette smoking. A similar proportion of case subjects (25.6%) and control subjects (24.4%) reported ever use of marijuana (adjusted odds ratio, 0.9; 95% confidence interval, 0.6–1.3). There were no trends in risk observed with increasing duration or average frequency of use or time since first or last use. No subgroup defined by known or suspected OSCC risk factors (age, cigarette smoking, alcohol consumption, and genetic polymorphisms) showed an increased risk. Marijuana use was not associated with OSCC risk in this large, population-based study.
Recent Advances in Stress Signaling in Cancer
Authors: Morin Chantale I. , Huot Jacques
Published: September 2005

This meeting brought together some of the world’s leading scientists in the field of stress signaling, apoptosis, and cancer. This resulted in a productive interaction that updated our current knowledge on “Stress Signaling in Cancer.” It comes out that subtle disturbances in cell signaling can be associated with and even lead to cancer. As a corollary, it appears that correcting the signaling defects associated with cancer constitutes a new approach to the treatment and control of neoplastic diseases. The meeting also raised several questions that should be addressed. In particular, it is of the utmost importance to better understand the mechanisms that underlie the specificity of the cellular response with regard to different ligands. For example, why does Gadd45b prevent apoptotic cell death in response to tumor necrosis factor α, whereas it favors apoptosis after transforming growth factor β (E. De Smaele et al., Nature (Lond.), 414: 308–313, 2001; J. Yoo et al., J. Biol. Chem., 278: 43001–43007, 2003). Other questions concern the understanding of the cross-talk mechanisms between different stress and apoptotic pathways and how the strength and the position and timing of a signal may affect different pathways. The next few years of research in this field should be enlightening and fruitful.
Caffeine Inhibits Cell Proliferation by G0/G1 Phase Arrest in JB6 Cells
Authors: Hashimoto Takashi , He Zhiwei , Ma Wei-Ya , Schmid Patricia C. , Bode Ann M. , Yang Chung S. , Dong Zigang
Published: September 2005

Caffeine is a major biologically active constituent in coffee and tea. Because caffeine has been reported to inhibit carcinogenesis in UVB-exposed mice, the cancer-preventing effect of caffeine has attracted considerable attention. In the present study, the effect of caffeine in quiescent (G0 phase) cells was investigated. Pretreatment with caffeine suppressed cell proliferation in a dose-dependent manner 36 h after addition of fetal bovine serum as a cell growth stimulator. Analysis by flow cytometry showed that caffeine suppressed cell cycle progression at the G0/G1 phase, i.e., 18 h after addition of fetal bovine serum, the percentages of cells in G0/G1 phase in 1 mm caffeine-treated cells and in caffeine-untreated cells were 61.7 and 29.0, respectively. The percentage of cells in G0/G1 phase at 0 h was 75.5. Caffeine inhibited phosphorylation of retinoblastoma protein at Ser780 and Ser807/Ser811, the sites where retinoblastoma protein has been reported to be phosphorylated by cyclin-dependent kinase 4 (cdk4). Furthermore, caffeine inhibited the activation of the cyclin D1-cdk4 complex in a dose-dependent manner. However this compound did not directly inhibit the activity of this complex. In addition, caffeine did not affect p16INK4 or p27Kip1 protein levels, but inhibited the phosphorylation of protein kinase B (Akt) and glycogen synthase kinase 3β. Our results showed that caffeine suppressed the progression of quiescent cells into the cell cycle. The inhibitory mechanism may be due to the inhibition of cell growth signal-induced activation of cdk4, which may be involved in the inhibition of carcinogenesis in vivo.
Transcriptional Abnormality of the
hsMAD2
Mitotic Checkpoint Gene Is a Potential Link to Hepatocellular Carcinogenesis
Authors: Jeong Sook-Jung , Shin Hyun-Jin , Kim So-Jung , Ha Geun-Hyoung , Cho Bok-Im , Baek Kwan-Hyuck , Kim Chang-Min , Lee Chang-Woo
Published: September 2005

MAD2 is localized to kinetochores of unaligned chromosomes, where it inactivates the anaphase-promoting complex/cyclosome, thus contributing to the production of a diffusible anaphase inhibitory signal. Disruption of MAD2 expression leads to defects in the mitotic checkpoint, chromosome missegregation, and tumorigenesis. However, the mechanism by which deregulation and/or abnormality of hsMAD2 expression remains to be elucidated. Here, we clone and analyze a ∼0.5 kb fragment upstream of hsMAD2 and show that this fragment acts as a strong promoter. Transcriptional dysfunction of hsMAD2 is frequently observed in hepatocellular carcinoma cells, and down-regulation of hsMAD2 protein expression is correlated with transcriptional silencing of the hsMAD2 promoter by hypermethylation. These results imply a relationship between transcriptional abnormality of this mitotic checkpoint gene and mitotic abnormality in human cancers.
BPR0L075, a Novel Synthetic Indole Compound with Antimitotic Activity in Human Cancer Cells, Exerts Effective Antitumoral Activityin Vivo
Authors: Kuo Ching-Chuan , Hsieh Hsing-Pang , Pan Wen-Yu , Chen Ching-Ping , Liou Jing-Ping , Lee Shiow-Ju , Chang Yi-Ling , Chen Li-Tzong , Chen Chiung-Tong , Chang Jang-Yang
Published: September 2005

BPR0L075 is a novel synthetic compound discovered through research to identify new microtubule inhibitors. BPR0L075 inhibits tubulin polymerization through binding to the colchicine-binding site of tubulin. Cytotoxic activity of BPR0L075 in a variety of human tumor cell lines has been ascertained, with IC50 values in single-digit nanomolar ranges. As determined by flow cytometry, human cervical carcinoma KB cells are arrested in G2-M phases in a time-dependent manner before cell death occurs. Terminal deoxynucleotidyl transferase-mediated nick end labeling assay indicates that cell death proceeds through an apoptotic pathway. Additional studies indicate that the effect of BPR0L075 on cell cycle arrest is associated with an increase in cyclin B1 levels and a mobility shift of Cdc2 and Cdc25C. The changes in Cdc2 and Cdc25C coincide with the appearance of phosphoepitopes recognized by a marker of mitosis, MPM-2. Furthermore, phosphorylated forms of Bcl-2, perturbed mitochondrial membrane potential, and activation of the caspase-3 cascade may be involved in BPR0L075-induced apoptosis. Notably, several KB-derived multidrug-resistant cell lines overexpressing P-gp170/MDR and MRP are resistant to vincristine, paclitaxel, and colchicine but not to BPR0L075. Moreover, BPR0L075 shows potent activity against the growth of xenograft tumors of the gastric carcinoma MKN-45, human cervical carcinoma KB, and KB-derived P-gp170/MDR-overexpressing KB-VIN10 cells at i.v. doses of 50 mg/kg in nude mice. These findings indicate BPR0L075 is a promising anticancer compound with antimitotic activity that has potential for management of various malignancies, particularly for patients with drug resistance.
Vitamin D Receptor and p21/WAF1 Are Targets of Genistein and 1,25-Dihydroxyvitamin D3 in Human Prostate Cancer Cells
Authors: Rao Anuradha , Coan April , Welsh Jo-Ellen , Barclay Wendy W. , Koumenis Constantinos , Cramer Scott D.
Published: September 2005

We investigated mechanisms by which genistein and 1,25-dihydroxyvitamin D3 [1,25(OH)2D3] act synergistically to inhibit the growth of the human prostate cancer cell line LNCaP. We demonstrate that 1,25(OH)2D3 and genistein cooperate to up-regulate the vitamin D receptor protein by increasing the stability of the vitamin D receptor. Genistein and 1,25(OH)2D3 also cooperate to up-regulate the levels of p21/WAF1 (p21). Small interfering RNA-mediated knockdown of p21 expression showed that p21 is essential for significant growth inhibition of LNCaP cells in response to either compound or their combination. We conclude that one mechanism of synergism between genistein and 1,25(OH)2D3 is through genistein modulation of vitamin D signaling.
Prostate Cancer Cell Adhesion to Bone Marrow Endothelium
Authors: Romanov Victor I. , Whyard Terry , Adler Howard L. , Waltzer Wayne C. , Zucker Stanley
Published: September 2005

Bone metastasis is the most frequent complication of prostate cancer (PC). Elucidation of the biological basis of this specificity is required for the development of approaches for metastatic inhibition. We investigated the possibility that the preferential attachment of PC cells to bone marrow endothelium (as opposed to endothelium from other organs) affects this specificity. We selected, from peptide phage-displayed libraries, peptide ligands to surfaces of PC cells (C4-2B) attenuated (30–40%) binding of C4-2B cells to bone marrow endothelial cells (BMECs). We then determined the molecules on the surface of C4-2B cells interacted with the selected peptides using column affinity chromatography and a cDNA expression phage-displayed library generated from C4-2B cells in T7 phage. We identified a phage from the cDNA library that specifically bound to one of the selected peptides-L11. This phage displayed the amino acid sequence homologous for the COOH-terminal portion of prostate-specific antigen (PSA). To examine the possible direct involvement of PSA in the interactions between PC and BMECs, we performed a cell–cell adhesion assay. Antibodies to PSA attenuated PC cells adhesion to BMECs. In addition, exogenous proteolytically active PSA modulated this adhesion. Finally, inactivation of mRNA coding PSA by a small interfering RNA (siRNA) diminished C4-2B cell adhesion to BMECs. These results indicate that PSA expressed as secreted and surface-associated molecules in C4-2B cells is involved in cell–cell interactions and/or digests components of bone marrow endothelium for preferential adhesion and penetration of PC cells. The suggested experimental approach is a promising strategy for identification of cell surface molecules involved in intercellular interactions.
Constitutive and Induced CD44 Shedding by ADAM-Like Proteases and Membrane-Type 1 Matrix Metalloproteinase
Authors: Nakamura Hiroyuki , Suenaga Naoko , Taniwaki Kaori , Matsuki Hirokazu , Yonezawa Kayoko , Fujii Masato , Okada Yasunori , Seiki Motoharu
Published: September 2005

CD44 is a receptor for hyaluronan and mediates signaling that regulates complex cell behavior including cancer cell migration and invasion. Shedding of the extracellular portion of CD44 is the last step in the regulation of the molecule-releasing interaction between the ligand and cell. However, highly glycosylated forms of CD44 have hampered the identification of the exact cleavage sites for shedding and the responsible proteases. In this study, we found that expression of membrane-type 1 matrix metalloproteinase (MT1-MMP) increased shedding of the 65–70 kDa CD44H (standard form) fragments and generated two additional smaller fragments. We purified the shed fragments and identified the cleaved sites by mass spectrometry. Specific antibodies that recognize the newly exposed COOH terminus by cleavage were prepared and used to analyze shedding at each site. Shedding of the 65–70 kDa fragments was inhibited by tissue inhibitor of metalloproteinase 3 (TIMP-3) but not by TIMP-1 and TIMP-2, suggesting involvement of a disintegrin and metalloproteinase (ADAM)-like proteases, although shedding is affected by MT1-MMP. Conversely, shedding of the two smaller fragments was inhibited by TIMP-2 and TIMP-3 but not TIMP-1, suggesting involvement of MT1-MMP itself. Shed fragments cleaved at these sites were also detected in human tumor tissues. Increased shedding at one of the MT1-MMP-sensitive sites was observed in the tumor compared with the surrounding normal tissue. However, no significant difference was observed with shedding by ADAM-like proteases. Thus, the cleavage sites for the shedding of CD44H were identified for the first time, and the results provide a basis for exploring the unknown biologic roles of shedding at different sites.
HER-2, gp100, and MAGE-1 Are Expressed in Human Glioblastoma and Recognized by Cytotoxic T Cells
Authors: Liu Gentao , Ying Han , Zeng Gang , Wheeler Christopher J. , Black Keith L. , Yu John S.
Published: September 2005

It has recently been demonstrated that malignant glioma cells express certain known tumor-associated antigens, such as HER-2, gp100, and MAGE-1. To further determine the possible utilization of these antigens for glioma immunotherapy and as surrogate markers for specific tumor antigen cytotoxicity, we characterized the presence of mRNA and protein expression in 43 primary glioblastoma multiforme (GBM) cell lines and 7 established human GBM cell lines. HER-2, gp100, and MAGE-1 mRNA expression was detected in 81.4%, 46.5%, and 39.5% of the GBM primary cell lines, respectively. Using immunoreactive staining analysis by flow cytometry, HER-2, gp100, and MAGE-1 protein expression was detected in 76%, 45%, and 38% of the GBM primary cell lines, respectively. HLA-A1-restricted epitope specific for MAGE-1 peptide (EADPTGHSY) CTL clone B07 and HLA-A2-restricted epitope specific for HER-2 peptide (KIFGSLAFL) CTL clone A05 and gp100 peptide (ITDQVPFSV) CTL clone CK3H6 were used in this study. The specificity of CTL clone was verified by HLA/peptide tetramer staining. Three CTL clones could efficiently recognize GBM tumor cells in an antigen-specific and MHC class I-restricted manner. IFN-γ treatment can dramatically increase MHC class I expression of GBM tumor cells and significantly increase CTL recognition of tumor cells. Treatment with the DNA hypomethylating agent 5-aza-2′-deoxycytidine induced and up-regulated the mRNA expression of MAGE-1 and epitope presentation by autologous MHC. These data indicate that HER-2, gp100, and MAGE-1 could be used as tumor antigen targets for surrogate assays for antigen-specific CTLs or to develop antigen-specific active immunotherapy strategies for glioma patients.
Protein Kinase C α/β Inhibitor Go6976 Promotes Formation of Cell Junctions and Inhibits Invasion of Urinary Bladder Carcinoma Cells
Authors: Koivunen Jussi , Aaltonen Vesa , Koskela Sanna , Lehenkari Petri , Laato Matti , Peltonen Juha
Published: September 2005

Changes in activation balance of different protein kinase C (PKC) isoenzymes have been linked to cancer development. The current study investigated the effect of different PKC inhibitors on cellular contacts in cultured high-grade urinary bladder carcinoma cells (5637 and T24). Exposure of the cells to isoenzyme-specific PKC inhibitors yielded variable results: Go6976, an inhibitor of PKCα and PKCβ isoenzymes, induced rapid clustering of cultured carcinoma cells and formation of an increased number of desmosomes and adherens junctions. Safingol, a PKCα inhibitor, had similar but less pronounced effects. In contrast, a PKCδ inhibitor, rottlerin, had an opposite effect on cell clustering and caused dissociation of cell junctions. A broad-spectrum PKC inhibitor bisindolylmaleimide I did not have any apparent effect on the morphology of the cultures or on the number of cell junctions. Additional studies with Go6976 demonstrated that inhibition of PKCα and β isoenzymes induced translocation of β1-integrin from the cell-matrix junctions and that β4-integrin was translocated to face the culture substratum. Go6976 was also highly effective in inhibiting migration of carcinoma cells and inhibited invasion through artificial basement membrane. Our results on urinary bladder carcinoma cells emphasize that Go6976 is a potential anticancer drug due to its effects on cell-cell and cell-matrix junctions, migration, and invasion. Furthermore, the results may be explained by changes in PKC activation balance promoted by inhibition of PKCα/β.
Choice of Oxygen-Conserving Treatment Regimen Determines the Inflammatory Response and Outcome of Photodynamic Therapy of Tumors
Authors: Henderson Barbara W. , Gollnick Sandra O. , Snyder John W. , Busch Theresa M. , Kousis Philaretos C. , Cheney Richard T. , Morgan Janet
Published: September 2005

The rate of light delivery (fluence rate) plays a critical role in photodynamic therapy (PDT) through its control of tumor oxygenation. This study tests the hypothesis that fluence rate also influences the inflammatory responses associated with PDT. PDT regimens of two different fluences (48 and 128 J/cm2) were designed for the Colo 26 murine tumor that either conserved or depleted tissue oxygen during PDT using two fluence rates (14 and 112 mW/cm2). Tumor oxygenation, extent and regional distribution of tumor damage, and vascular damage were correlated with induction of inflammation as measured by interleukin 6, macrophage inflammatory protein 1 and 2 expression, presence of inflammatory cells, and treatment outcome. Oxygen-conserving low fluence rate PDT of 14 mW/cm2 at a fluence of 128 J/cm2 yielded ∼70–80% tumor cures, whereas the same fluence at the oxygen-depleting fluence rate of 112 mW/cm2 yielded ∼10–15% tumor cures. Low fluence rate induced higher levels of apoptosis than high fluence rate PDT as indicated by caspase-3 activity and terminal deoxynucleotidyl transferase-mediated nick end labeling analysis. The latter revealed PDT-protected tumor regions distant from vessels in the high fluence rate conditions, confirming regional tumor hypoxia shown by 2-(2-nitroimidazol-1[H]-yl)-N-(3,3,3-trifluoropropyl) acetamide staining. High fluence at a low fluence rate led to ablation of CD31-stained endothelium, whereas the same fluence at a high fluence rate maintained vessel endothelium. The highest levels of inflammatory cytokines and chemokines and neutrophilic infiltrates were measured with 48 J/cm2 delivered at 14 mW/cm2 (∼10–20% cures). The optimally curative PDT regimen (128 J/cm2 at 14 mW/cm2) produced minimal inflammation. Depletion of neutrophils did not significantly change the high cure rates of that regimen but abolished curability in the maximally inflammatory regimen. The data show that a strong inflammatory response can contribute substantially to local tumor control when the PDT regimen is suboptimal. Local inflammation is not a critical factor for tumor control under optimal PDT treatment conditions.
Gene Expression Profile of Gastric Carcinoma
Authors: Oue Naohide , Hamai Yoichi , Mitani Yoshitsugu , Matsumura Shunji , Oshimo Yasuhiro , Aung Phyu Phyu , Kuraoka Kazuya , Nakayama Hirofumi , Yasui Wataru
Published: September 2005

Gastric carcinoma (GC) is one of the most common malignancies worldwide. To better understand the genetic basis of this disease, we performed serial analysis of gene expression (SAGE) on four primary GC samples and one associated lymph node metastasis. We obtained a total of 137,706 expressed tags (Gene Expression Omnibus accession number GSE 545, SAGE Hiroshima gastric cancer tissue), including 38,903 that were unique. Comparing tags from our GC libraries containing different stages and different histologies, we found several genes and tags that are potentially involved in invasion, metastasis, and carcinogenesis. Among these, we selected 27 genes and measured mRNA expression levels in an additional 46 GC samples by quantitative reverse transcription-PCR. Frequently overexpressed genes (tumor/normal ratio > 2) were COL1A1 (percentage of cases with overexpression, 78.3%), CDH17 (73.9%), APOC1 (67.4%), COL1A2 (58.7%), YF13H12 (52.2%), CEACAM6 (50.0%), APOE (50.0%), REGIV (47.8%), S100A11 (41.3%), and FUS (41.3%). Among these genes, mRNA expression levels of CDH17 and APOE were associated with depth of tumor invasion (P = 0.0060 and P = 0.0139, respectively), and those of FUS and APOE were associated with degree of lymph node metastasis (P = 0.0416 and P = 0.0006, respectively). In addition, mRNA expression levels of FUS, COL1A1, COL1A2, and APOE were associated with stage (P = 0.0414, P = 0.0156, P = 0.0395, and P = 0.0125, respectively). Quantitative reverse transcription-PCR analysis also showed a high level of REGIV expression (>100 arbitrary units) in 14 of 46 GC samples (30.4%) but not in noncancerous tissues. We detected V5-tagged RegIV protein in the culture media of cells transfected with pcDNA-RegIV-V5 by Western blot. Our results provide a list of candidate genes that are potentially involved in invasion, metastasis, and carcinogenesis of GC. REGIV may serve as a specific biomarker for GC.
Progressive Loss of Syk and Abnormal Proliferation in Breast Cancer Cells
Authors: Moroni Maria , Soldatenkov Viatcheslav , Zhang Li , Zhang Ying , Stoica Gerald , Gehan Edmund , Rashidi Banafsheh , Singh Baljit , Ozdemirli Metin , Mueller Susette C.
Published: September 2005

The tumor suppressor gene Syk tyrosine kinase is absent or reduced in invasive breast cancer tissues and cell lines; its loss in breast tissues is linked to poor prognosis and metastasis. Also, evidence shows that in vitro Syk is involved in regulating proliferation. Here, we show by in situ hybridization on breast tissue sections that the loss of Syk expression is progressive during tumor development. Strikingly, Syk is already partially lost in normal epithelial tissue adjacent to the cancer lesion. In vivo, cell proliferation (as measured by the proliferative index Ki67) increased from normal to ductal carcinoma in situ to invasive, whereas Syk in situ staining in the same tissues decreased. In vitro, the presence of Syk was associated with reduced cell proliferation in an epidermal growth factor receptor-overexpressing breast cancer cell line, BT549, whereas changes in apoptosis were undetected. Concomitantly, the kinase activity of the proto-oncogene Src was reduced by ∼30%. A 5-fold increase in abnormal mitoses was observed in the Syk-transfected cells compared with vector control. We propose that Syk is involved in the regulation of cell proliferation, possibly by controlling mechanisms of mitosis and cytokinesis via Src signal transduction pathway(s). Because of its progressive and early loss during tumor onset and development, monitoring of Syk loss in breast epithelial cells by noninvasive techniques such as ductal lavage may be a powerful tool for screening purposes.
Immediate Early Response Gene X-1, a Stress-Inducible Antiapoptotic Gene, Encodes Cytotoxic T-Lymphocyte (CTL) Epitopes Capable of Inducing Human Leukocyte Antigen-A33-Restricted and Tumor-Reactive CTLs in Gastric Cancer Patients
Authors: Sasada Tetsuro , Takedatsu Hiroko , Azuma Koichi , Koga Makoto , Maeda Yoshiaki , Shichijo Shigeki , Shoumura Hiroki , Hirai Tatsuya , Takabayashi Arimichi , Itoh Kyogo
Published: September 2005

Peptide-based vaccine therapy, which is designed to elicit T-cell immunity against tumors, is an attractive approach for the treatment of cancer patients. To provide a scientific basis for peptide therapy, an increasing number of CTL-directed peptides have been identified, and some of them have been tried as antigen-specific immunotherapy in the past decade. Only a few studies, however, have been performed on such peptides restricted with alleles other than HLA-A2 and −A24. In the present study, we show that immediate early response gene X-1 (IEX-1), a stress-inducible protein associated with the regulation of cell proliferation and apoptosis, produces antigenic epitopes recognized by 850B-CTLs, HLA-A33-restricted CTLs newly established from T cells infiltrating into gastric adenocarcinoma. The IEX-1 gene was highly expressed in most cell lines and tissues from various types of cancer at both the mRNA and protein levels. However, it was not expressed at the protein level in any normal epithelium or connective tissues tested. Three IEX-1-derived peptides at positions 47–56, 61–69, and 65–73, which were recognized by the 850B-CTLs, could induce CD8+ peptide-specific CTL reaction to tumor cells from HLA-A33+ gastric cancer patients and other epithelial cancer patients, but not from healthy donors, in an HLA class I-restricted manner. Because increased expression of IEX-1 is suggested to be involved in the resistance to apoptosis and in the proliferation of cancer cells, these antigenic peptides could be potent candidates for peptide-based specific immunotherapy against HLA-A33+ gastric cancer and other epithelial cancers.
Interferon Regulatory Factor-1 Mediates the Proapoptotic but Not Cell Cycle Arrest Effects of the Steroidal Antiestrogen ICI 182,780 (Faslodex, Fulvestrant)
Authors: Bouker Kerrie B. , Skaar Todd C. , Fernandez David R. , O’Brien Kerry A. , Riggins Rebecca B. , Cao Donghua , Clarke Robert
Published: September 2005

Antiestrogens induce both cytostasis (cell cycle arrest) and apoptosis, but the relationship between these end points and the signaling that regulates their induction are unclear. We have previously implicated the transcription factor and putative tumor suppressor IFN regulatory factor-1 (IRF-1) in acquired antiestrogen resistance (Gu et al., Cancer Res, 62: 3428–3437, 2002). We now show the functional significance of IRF-1 in affecting antiestrogen responsiveness in estrogen receptor-positive antiestrogen-sensitive models (MCF-7, T47D, and ZR-75-1), a model of acquired antiestrogen resistance (MCF7/LCC9; estrogen receptor positive), and a model of de novo antiestrogen resistance (MDA-MB-231; estrogen receptor negative). Basal IRF-1 mRNA expression is lower in MCF7/LCC9 cells when compared with MCF-7, T47D, and ZR-75-1 cells. IRF-1 transcriptional activity in MCF-7/LCC9 cells is 18-fold lower than that seen in the parental cells (MCF-7/LCC1) and is comparable with that in MDA-MB-231 cells. Although IRF-1 mRNA expression is induced by ICI 182,780 in sensitive cells, this regulation is lost in MCF-7/LCC9 and is absent in MDA-MB-231 cells. Loss of IRF-1 regulation appears specific to antiestrogen resistance—resistant cells induce IRF-1 mRNA in response to the cytotoxic drug doxorubicin. A dominant-negative IRF-1 eliminates the ICI 182,780-induced apoptotic response (reduced >4-fold) and reduces MCF-7 and T47D cell sensitivity to the antiproliferative effects of ICI 182,780. This effect is not mediated by changes in cell cycle distribution; rather, dominant-negative IRF-1 reduces ICI 182,780-induced apoptosis. These data identify a novel mechanism of antiestrogen resistance and implicate IRF-1 as a key component in signaling some ER-mediated effects on apoptosis/cell survival.
Characterization of the t(14;18)
BCL2-IGH
Translocation in Farmers Occupationally Exposed to Pesticides
Authors: Roulland Sandrine , Lebailly Pierre , Lecluse Yannick , Briand Mélanie , Pottier Didier , Gauduchon Pascal
Published: September 2005

Increasing incidence of non-Hodgkin’s lymphoma have been associated repeatedly with farming occupation and particular attention focused on the role of pesticide exposure to potentially explain part of this trend. A genetic hallmark of non-Hodgkin’s lymphoma is the presence of recurrent chromosomal translocations involving the immunoglobulin heavy chain gene. Of these, the t(14;18), which deregulates BCL2 expression and inhibits apoptosis, is the most frequent in follicular lymphoma and has been detected consistently in peripheral blood lymphocytes of healthy individuals. As BCL2-IGH translocation represents an early step of the malignant process, we evaluated the occurrence and molecular characteristics of BCL2-IGH translocation in 56 individuals occupationally exposed to pesticides in open field farming They were selected from a representative cohort of farmers with a well-defined assessment of pesticide exposure taking into account potential confounding factors, smoking, sunlight, and age. Our results suggest that occupational exposure to pesticides would increase BCL2-IGH prevalence together with the frequency of BCL2-IGH-bearing cells especially during the high pesticide use period. Distribution of BCL2 or IGH breakpoint positions seemed to be independent of pesticide exposure and was similar to those found in other healthy populations or lymphoma patients. Finally, these results provide additional evidence that BCL2-IGH translocation measurements could be a measure of acquired genetic instability in relation to genotoxic exposure in a gene directly relevant in term of lymphomagenesis.
RETRACTED: Mechanisms by which SGN-40, a Humanized Anti-CD40 Antibody, Induces Cytotoxicity in Human Multiple Myeloma Cells: Clinical Implications
Authors: Tai Yu-Tzu , Catley Laurence P. , Mitsiades Constantine S. , Burger Renate , Podar Klaus , Shringpaure Reshma , Hideshima Teru , Chauhan Dharminder , Hamasaki Makoto , Ishitsuka Kenji , Richardson Paul , Treon Steven P. , Munshi Nikhil C. , Anderson Kenneth C.
Published: September 2005

CD40 is expressed on B-cell malignancies, including human multiple myeloma (MM) and a variety of carcinomas. We examined the potential therapeutic utility of SGN-40, the humanized anti-CD40 monoclonal antibody, for treating human MM using MM cell lines and patient MM cells (CD138++, CD40+). SGN-40 (0.01–100 μg/ml) induces modest cytotoxicity in MM cell lines and patient MM cells. In the presence of de novo protein synthesis inhibitor cycloheximide, SGN-40 significantly induced apoptosis in Dexamethasone (Dex)-sensitive MM.1S and Dex-resistant MM.1R cells and in patient MM cells. SGN-40-mediated cytotoxicity is associated with up-regulation of cytotoxic ligands of the tumor necrosis factor family (Fas/FasL, tumor necrosis factor-related apoptosis-inducing ligand, and tumor necrosis factor α). SGN-40 treatment also induces a down-regulation of CD40 dependent on an endocytic pathway. Consequently, pretreatment of MM cells with SGN-40 blocked sCD40L-mediated phosphatidylinositol 3′-kinase/AKT and nuclear factor κB activation. Importantly, pretreatment of MM.1S and MM.1R cells with SGN-40 inhibited proliferation triggered by interleukin 6 (IL-6) but not by insulin-like growth factor-I. In addition, SGN-40 pretreatment of MM.1S cells blocked the ability of IL-6 to protect against Dex-induced inhibition of DNA synthesis. This was associated with a 2–4-fold reduction of IL-6 receptor at protein and mRNA levels in SGN-40-treated MM.1S cells and patient MM cells. Taken together, these results provide the preclinical rationale for the evaluation of SGN-40 as a potential new therapy to improve patient outcome in MM.
Treatment-Induced Changes in Tumor Oxygenation Predict Photodynamic Therapy Outcome
Authors: Wang Hsing-Wen , Putt Mary E. , Emanuele Michael J. , Shin Daniel B. , Glatstein Eli , Yodh Arjun G. , Busch Theresa M.
Published: September 2005

Photodynamic therapy (PDT) requires oxygen to cause tumor damage, yet therapy itself can deplete or enhance tumor oxygenation. In the present work we measured the PDT-induced change in tumor oxygenation and explored its utility for predicting long-term response to treatment. The tissue hemoglobin oxygen saturation (SO2) of murine tumors was noninvasively measured by broadband diffuse reflectance spectroscopy. In initial validation studies, the oxyhemoglobin dissociation curve for mouse blood was accurately recreated based on measurements during deoxygenation of a tissue phantom of mouse erythrocytes. In vivo studies exhibited excellent correlation between carbogen-induced changes in SO2 and pO2 of radiation-induced fibrosarcoma tumors measured by reflectance spectroscopy and the Eppendorf pO2 histograph, respectively. In PDT studies radiation-induced fibrosarcoma tumor SO2 was measured immediately before and after Photofrin-PDT (135 J/cm2, 38 mW/cm2). Animals were subsequently followed for tumor growth to a volume of 400 mm3 (time-to-400 mm3) or the presence of tumor cure (no tumor growth at 90 days after treatment). In animals that recurred, the PDT-induced change in tumor SO2, i.e., relative-SO2 (SO2 after PDT/SO2 before PDT) was positively correlated with treatment durability (time-to-400 mm3). The predictive value of relative-SO2 was confirmed in a second group of animals with enhanced pre-PDT oxygenation due to carbogen breathing. Furthermore, when all of the animals were considered (those that recurred and those that were cured) a highly significant association was found between increasing relative-SO2 and increasing probability of survival, i.e., absence of recurrence. As independent variables, the SO2 after PDT, the pre-PDT tumor volume, and light penetration depth all failed to predict response. As an independent variable, the SO2 before PDT demonstrated a weak negative association with treatment durability; this association was driven by a correlation between decreasing pre-PDT SO2 and increasing relative-SO2. These data suggest that monitoring of PDT-induced changes in tumor oxygenation may be a valuable prognostic indicator.
Inverse PCR-Based RFLP Scanning Identifies Low-Level Mutation Signatures in Colon Cells and Tumors
Authors: Liu Wei-Hua , Kaur Manjit , Wang Gang , Zhu Penny , Zhang Yuzhi , Makrigiorgos G. Mike
Published: September 2005

Detecting the presence and diversity of low-level mutations in human tumors undergoing genomic instability is desirable due to their potential prognostic value and their putative influence on the ability of tumors to resist drug treatment and/or metastasize. However, direct measurement of these genetic alterations in surgical samples has been elusive, because technical hurdles make mutation discovery impractical at low-mutation frequency levels (<10−2). Here, we describe inverse PCR-based amplified restriction fragment length polymorphism (iFLP), a new technology that combines inverse PCR, RFLP, and denaturing high-performance liquid chromatography to allow scanning of the genome at several thousand positions per experiment for low-level point mutations. Using iFLP, widespread, low-level mutations at mutation frequency 10−2–10−4 were discovered in genes located on different chromosomes, e.g., OGG1, MSH2, PTEN, β-catenin, Bcl-2, P21, ATK3, and Braf, in human colon cancer cells that harbor mismatch repair deficiency whereas mismatch repair-proficient cells were mutation free. Application of iFLP to the screening of sporadic colon cancer surgical specimens demonstrated widespread low-level mutations in seven out of 10 samples, but not in their normal tissue counterparts, and predicted the presence of millions of diverse, low-incidence mutations in tumors. Unique low-level mutational signatures were identified for each colon cancer cell line and tumor specimen. iFLP allows the high-throughput discovery and tracing of mutational signatures in human cells, precancerous lesions, and primary or metastatic tumors and the assessment of the number and heterogeneity of low-level mutations in surgical samples.
The Kaposi’s Sarcoma-Associated Herpesvirus (KSHV/HHV-8) K1 Protein Induces Expression of Angiogenic and Invasion Factors
Authors: Wang Ling , Wakisaka Naohiro , Tomlinson Christine C , DeWire Scott M , Krall Stuart , Pagano Joseph S , Damania Blossom
Published: September 2005

Kaposi’s sarcoma-associated herpesvirus (KSHV/HHV-8) has been linked to Kaposi’s sarcoma, primary effusion lymphoma, and multicentric Castleman’s disease. In addition to endothelial cells and B lymphocytes, KSHV also has been shown to infect epithelial cells and keratinocytes. The transmembrane glycoprotein K1, encoded by the first open reading frame of KSHV, is a signaling protein capable of eliciting B-cell activation. We show that KSHV K1 can induce expression and secretion of vascular endothelial growth factor (VEGF) in epithelial and endothelial cells. Up-regulation of VEGF was mediated at the transcriptional level because expression of K1 resulted in VEGF promoter activation. We also show that K1 induces expression of matrix metalloproteinase-9 (MMP-9) in endothelial cells. Additional analyses with K1 mutant proteins revealed that the SH2 binding motifs present in the K1 cytoplasmic tail are necessary for VEGF secretion and MMP-9 induction. These results indicate that K1 signaling may contribute to KSHV-associated pathogenesis through a paracrine mechanism by promoting the secretion of VEGF and MMP-9 into the surrounding matrix.
Effect of Mutational Inactivation of Tyrosine Kinase Activity on BCR/ABL-Induced Abnormalities in Cell Growth and Adhesion in Human Hematopoietic Progenitors
Authors: Ramaraj Pandurangan , Singh Harjeet , Niu Ning , Chu Su , Holtz Melissa , Yee Jiing Kuan , Bhatia Ravi
Published: September 2005

Chronic myelogenous leukemia (CML) results from transformation of a primitive hematopoietic cell by the BCR/ABL gene. The specific BCR/ABL signaling mechanisms responsible for transformation of primitive human hematopoietic cells are not well defined. Previous studies have suggested that constitutively activated tyrosine kinase activity plays an important role for in abnormal proliferation of CML progenitors but has not clearly defined its role in abnormal adhesion and migration. We established a human progenitor model of CML by ectopic expression of BCR/ABL in normal CD34+ cells using retrovirus-mediated gene transfer. CD34+ cells expressing BCR/ABL demonstrated several features characteristic of primary CML progenitors including increased proliferation in committed and primitive progenitor culture, reduced adhesion to fibronectin, and reduced chemotaxis to stroma-derived factor-1α. We expressed a kinase-inactive BCR/ABL gene to directly investigate the role of kinase activity in abnormal progenitor function. Abnormalities in proliferation were completely reversed, whereas defects in adhesion and migration were significantly improved but not completely reversed in cells expressing a kinase-inactive BCR/ABL. Furthermore, the BCR/ABL kinase inhibitor imatinib mesylate markedly inhibited proliferation of BCR/ABL-expressing progenitors but did not fully correct the adhesion and migration defects. Expression of BCR/ABL genes with deletions of either the COOH-terminal actin binding or proline-rich domains resulted in enhanced adhesion and chemotaxis compared with wild-type BCR/ABL but did not affect progenitor proliferation. We conclude that abnormal kinase activity is essential for abnormal proliferation and survival of CML progenitors but that abnormal adhesion and migration result from both kinase-dependent and -independent mechanisms.
Nitrosative Stress in Rotated Three-Dimensional Colorectal Carcinoma Cell Cultures Induces Microtubule Depolymerization and Apoptosis
Authors: Laguinge Luciana M. , Lin Shuling , Samara Raed N. , Salesiotis Anastasios N. , Jessup J. Milburn
Published: September 2005

Malignant cells undergo anoikis as they encounter fluid shear stress during transit to a metastatic site. We postulated that intracellular nitric oxide (NO) contributes to this cell death by comparing the growth of human colorectal carcinoma cells in low fluid shear stress rotated three-dimensional (Rotated 3-D) cultures with growth in static three-dimensional (Static 3-D) cultures on nonadherent surfaces and with two-dimensional monolayer (Monolayer 2-D) cultures. NO, loss of microtubules, and apoptosis increased significantly in Rotated 3-D cultures within 10 min and persisted at 24 h, whereas inhibition of NO synthase decreased apoptosis and intracellular NO and prevented tubulin degradation. Thus, fluid shear stress and three-dimensional growth increases NO synthase and NO to cause tubulin breakdown and induce anoikis. Intracellular NO in malignant cells entering the circulation may be a novel target for metastasis by colorectal carcinoma.
The Human Cervical Cancer Oncogene Protein Is a Biomarker for Human Hepatocellular Carcinoma
Authors: Yoon Seung Kew , Lim Nam Kyu , Ha Seon-Ah , Park Yong Gyu , Choi Jong Young , Chung Kyu Won , Sun Hee Sik , Choi Myung Ja , Chung Junho , Wands Jack R. , Kim Jin Woo
Published: September 2005

Human cervical cancer oncogene (HCCR) was identified and appeared to function as a negative regulator of p53 gene. The objective of this study was to validate HCCR expression as a candidate marker for human hepatocellular carcinoma. HCCR epitope was identified as Y355LGTRR360. According to immunofluorescence study, HCCR was predominantly localized in the plasma membrane and cytoplasm of hepatocellular carcinoma. HCCR proteins were overexpressed in the tumorous compared with the nontumorous cirrhosis tissues. However, HCCR was not detected in normal liver tissue. Concentration of HCCR protein in the serum was measured in a total of 570 subjects, and comparisons were made to α-fetoprotein. Serological studies revealed 78.2% sensitivity of HCCR (cutoff value, 15 μg/ml), which was significantly higher than 64.6% of α-fetoprotein (P = 0.0098) and 95.7% specificity for hepatocellular carcinoma. Forty of 52 (76.9%) patients with carcinoma negative for α-fetoprotein showed positive values for HCCR. A positive rate of 69.2% in carcinoma patients with tumor sizes <2 cm was found to be a higher rate than measurement of α-fetoprotein. Furthermore, HCCR expression was also detected in liver cirrhosis at an intermediate level between carcinoma and normal groups, which gave 88.1% sensitivity and 79.0% specificity using 8 μg/ml as a cutoff value. In summary, the HCCR assay may have an advantage over the α-fetoprotein assay in that it is elevated according to disease progression from liver cirrhosis to carcinoma, and it is more frequently positive in patients with early, small hepatocellular carcinoma.