bluuebunny/crossref_metadata_2025 huggingface dataset:
A Transforming Growth Factor-β Receptor–Interacting Protein Frequently Mutated in Human Ovarian Cancer
Authors: Ding Wei , Tang Qian , Espina Virginia , Liotta Lance A. , Mauger David T. , Mulder Kathleen M.
Published: September 2005

Ovarian carcinomas, particularly recurrent forms, are frequently resistant to transforming growth factor-β (TGF-β)–mediated growth inhibition. However, mutations in the TGF-β receptor I and receptor II (TβR-I and TβR-II) genes have only been reported in a minority of ovarian carcinomas, suggesting that alterations in TGF-β–signaling components may play an important role in the loss of TGF-β responsiveness. Using laser-capture microdissection and nested reverse-transcription-PCR, we found that km23, which interacts with the TGF-β receptor complex, is altered at a high frequency in human ovarian cancer patients. A novel form of km23, missing exon 3 (Δexon3-km23), was found in 2 of 19 tumor tissues from patients with ovarian cancer. In addition to this alteration, a stop codon mutation (TAA → CAC) was detected in two patients. This alteration results in an elongated protein, encoding 107-amino-acid residues (Δ107km23), instead of the wild-type 96-amino-acid form of km23. Furthermore, five missense mutations (T38I, S55G, T56S, I89V, and V90A) were detected in four patients, providing a total alteration rate of 42.1% (8 of 19 cases) in ovarian cancer. No km23 alterations were detected in 15 normal tissues. Such a high alteration rate in ovarian cancer suggests that km23 may play an important role in either TGF-β resistance or tumor progression in this disease. In keeping with these findings, the functional studies described herein indicate that both the Δexon3-km23 and S55G/I89V-km23 mutants displayed a disruption in binding to the dynein intermediate chain in vivo, suggesting a defect in cargo recruitment to the dynein motor complex. In addition, the Δexon3-km23 resulted in an inhibition of TGF-β–dependent transcriptional activation of both the p3TP-lux and activin responsive element reporters. Collectively, our results suggest that km23 alterations found in ovarian cancer patients result in altered dynein motor complex formation and/or aberrant transcriptional regulation by TGF-β.
Functional Epigenomics Identifies Genes Frequently Silenced in Prostate Cancer
Authors: Lodygin Dimitri , Epanchintsev Alexey , Menssen Antje , Diebold Joachim , Hermeking Heiko
Published: September 2005

In many cases, silencing of gene expression by CpG methylation is causally involved in carcinogenesis. Furthermore, cancer-specific CpG methylation may serve as a tumor marker. In order to identify candidate genes for inactivation by CpG methylation in prostate cancer, the prostate cancer cell lines LNCaP, PC3, and Du-145 were treated with 5-aza-2′ deoxycytidine and trichostatin A, which leads to reversion of epigenetic silencing. By microarray analysis of 18,400 individual transcripts, several hundred genes were found to be induced when compared with cells treated with trichostatin A. Fifty re-expressed genes were selected for further analysis based on their known function, which implied a possible involvement in tumor suppression. Twelve of these genes showed a significant degree of CpG methylation in their promoters. Six genes were silenced by CpG methylation in the majority of five analyzed prostate cancer cell lines, although they displayed robust mRNA expression in normal prostate epithelial cells obtained from four different donors. In primary prostate cancer samples derived from 41 patients, the frequencies of CpG methylation detected in the promoter regions of these genes were: GPX3, 93%; SFRP1, 83%; COX2, 78%; DKK3, 68%; GSTM1, 58%; and KIP2/p57, 56%. Ectopic expression of SFRP1 or DKK3 resulted in decreased proliferation. The expression of DKK3 was accompanied by attenuation of the mitogen-activated protein kinase pathway. The high frequency of CpG methylation detected in the promoters of the identified genes suggests a potential causal involvement in prostate cancer and may prove useful for diagnostic purposes.
Choline Kinase Is a Novel Oncogene that Potentiates RhoA-Induced Carcinogenesis
Authors: Ramírez de Molina Ana , Gallego-Ortega David , Sarmentero Jacinto , Bañez-Coronel Mónica , Martín-Cantalejo Yolanda , Lacal Juan Carlos
Published: September 2005

Choline kinase is overexpressed in human breast, lung, colorectal, and prostate tumors, a finding that suggests the involvement of this enzyme in carcinogenesis. Here we show that overexpression of choline kinase induce oncogenic transformation of human embryo kidney fibroblasts and canine epithelial Madin-Darby canine kidney cells. Choline kinase lays downstream of RhoA signaling and is activated through ROCK kinase, one of the best-characterized RhoA effectors. In keeping with this, coexpression of RhoA and choline kinase potentiates both anchorage independent growth and tumorigenesis. Finally, choline kinase–mediated transformation is sensitive to MN58b, a well-characterized specific choline kinase inhibitor. These results provide the definitive evidence that choline kinase has oncogenic properties and that choline kinase inhibition constitutes a novel valid antitumor strategy.
Impairment of Stromelysin-1 Transcriptional Activity by Promoter Mutations in High Microsatellite Instability Colorectal Tumors
Authors: Morán Alberto , Iniesta Pilar , de Juan Carmen , García-Aranda Cristina , Díaz-López Antonio , Benito Manuel
Published: September 2005

Colorectal tumorigenesis is characterized by the sequential inactivation of a series of tumor suppressor genes (microsatellite-stable tumors) and genetic or epigenetic alterations in mismatch repair genes in nonpoliposic hereditary tumours and 13% to 15% of sporadic colorectal cancer [high microsatellite instability (MSI-H) tumors]. We hypothesized a molecular mechanism for MSI-H colorectal tumors related to matrix metalloproteinase 3 (MMP-3) promoter mutations, down-regulation of MMP-3 expression, and impairment of MMP-9 activation. We have now analyzed the 2.2-kb full MMP-3 promoter to assess the mutation distribution. The mutations found are restricted to the polymorphic region that includes the zinc-binding protein (ZBP-89) binding element. To show that these alterations were the cause of the low expression of this gene, we have generated three constructs with different MMP-3 promoters (wild type and two mutants) and we have expressed them in SW480 human colorectal cells. The basal transcriptional activity of wild-type MMP-3 promoter was much higher than the mutants activity. In addition, 12-O-tetradecanoylphorbol-13-acetate (TPA)–induced transcriptional activity of wild-type MMP-3 promoter was 10-fold higher than the mutants activity. Dexamethasone inhibited the basal transcriptional activity of wild-type MMP-3 promoter and of the two mutants found in the MSI-H subgroup of colorectal tumors. Significantly, dexamethasone almost completely blunted the TPA-induced effect on wild-type MMP-3 promoter transcriptional activity and on the mutants, even below their basal activity. Our data show that mutations found in the polymorphic region of the MMP-3 promoter from MSI-H colorectal tumors impair its basal and induced transcriptional activity, which may contribute to their better clinical outcome.
Enforced Expression of Superoxide Dismutase 2/Manganese Superoxide Dismutase Disrupts Autocrine Interleukin-6 Stimulation in Human Multiple Myeloma Cells and Enhances Dexamethasone-Induced Apoptosis
Authors: Hodge David R. , Xiao Weihua , Peng Benjamin , Cherry James C. , Munroe David J. , Farrar William L.
Published: September 2005

Autocrine pathways of proliferative and antiapoptotic growth factors represent a serious impediment to the treatment of many types of tumors. In particular, interleukin-6 (IL-6), a pleiotropic cytokine known to play a critical role in the survival and growth of multiple myeloma cells, participates in an autocrine stimulation loop that serves to inhibit the induction of apoptosis during chemotherapy. Manganese superoxide dismutase (MnSOD) is an important antioxidant enzyme encoded by the SOD2 gene that attenuates oxidative free radicals in the mitochondria by catalyzing the formation of hydrogen peroxide from superoxide radicals. Transcription factor activity and binding is influenced by the oxidative state of cells, and dysregulation of MnSOD levels can result in abnormal patterns of gene expression. In the human multiple myeloma cell line IM-9, an autocrine IL-6 loop exists, which enables the cell to resist the effects of dexamethasone, a common treatment for multiple myeloma. Here, we show that SOD2 expression is epigenetically silenced in IM-9 cells, and replacement of MnSOD reduces cell proliferation and partially restores susceptibility to dexamethasone. The restoration of MnSOD also serves to decrease the expression levels of IL-6 by reducing the ability of activator protein-1, an important mediator of IL-6 expression in multiple myeloma cells, to bind to its enhancer site. These results show the importance of free radical–mediated dysregulation of autocrine growth factor loops in tumor cells and their effect on cell growth and response to chemotherapy.
Detection of Prostate Cancer with a Blood-Based Assay for Early Prostate Cancer Antigen
Authors: Paul Barbara , Dhir Rajiv , Landsittel Douglas , Hitchens Moira R. , Getzenberg Robert H.
Published: September 2005

Prostate-specific antigen lacks specificity for prostate cancer, so the identification and characterization of a unique blood-based marker for the disease would provide for a more accurate diagnosis, reducing both unnecessary biopsies and patient uncertainty. We previously identified a novel biomarker for prostate cancer, early prostate cancer antigen (EPCA). EPCA antibodies positively stained the negative biopsies of men who, as much as 5 years later, were diagnosed with prostate cancer. The goal of this study was to determine whether EPCA antibodies could be used in a clinically applicable plasma-based immunoassay to specifically detect prostate cancer. Using an EPCA-based ELISA, the protein was measured in the plasma of 46 individuals, including prostate cancer patients, healthy individuals, other cancer patients, spinal cord injury victims, and patients with prostatitis. With a predetermined cutoff value of 1.7 absorbance at 450 nm, only the prostate cancer population, as a whole, expressed plasma-EPCA levels above the cutoff. Statistical analysis showed a significant difference in EPCA levels between the prostate cancer population and each of the other groups, specifically the healthy donors (P < 0.0001), bladder cancer patients (P = 0.03), and spinal cord injury patients (P = 0.001). Sensitivity of the EPCA assay for prostate cancer patients was 92% whereas the overall specificity was 94%. Specificity for the healthy donors was 100%. Although larger trials are required, this initial study shows the potential of EPCA to serve as a highly specific blood-based marker for prostate cancer. EPCA, when coupled with prostate-specific antigen, may help reduce the number of both unnecessary biopsies and undetected prostate tumors.
The Retinoic Acid Synthesis GeneALDH1a2Is a Candidate Tumor Suppressor in Prostate Cancer
Authors: Kim Hanna , Lapointe Jacques , Kaygusuz Gulsah , Ong David E. , Li Chunde , van de Rijn Matt , Brooks James D. , Pollack Jonathan R.
Published: September 2005

Prostate cancer is the most common cancer among men in the United States, and aberrant DNA methylation is known to be an early molecular event in its development. Here, we have used expression profiling to identify novel hypermethylated genes whose expression is induced by treatment of prostate cancer cell lines with the DNA methyltransferase inhibitor 5-Aza-2′-deoxycytidine (5-aza-dC). Of the 271 genes that were induced by 5-aza-dC treatment, 25 also displayed reduced expression in primary prostate tumors compared with normal prostate tissue, and the decreased expression of only one gene, aldehyde dehydrogenase 1 family, member A2 (ALDH1a2), was also associated with shorter recurrence-free survival. ALDH1a2 encodes an enzyme responsible for synthesis of retinoic acid (RA), a compound with prodifferentiation properties. By immunohistochemistry, we observed that ALDH1a2 was expressed in epithelia from normal prostate but not prostate cancer. Using bisulfite sequencing, we determined that the ALDH1a2 promoter region was significantly hypermethylated in primary prostate tumors compared with normal prostate specimens (P = 0.01). Finally, transfection-mediated reexpression of wild-type ALDH1a2 (but not a presumptive catalytically dead mutant) in the prostate cancer cell line DU145 resulted in decreased colony growth (P < 0.0001), comparable with treatment with either 5-aza-dC or RA. Taken together, our findings implicate ALDH1a2 as a candidate tumor suppressor gene in prostate cancer and further support a role of retinoids in the prevention or treatment of prostate cancer.
Intratumoral T-Cell Infiltrates and MHC Class I Expression in Patients with Stage IV Melanoma
Authors: Al-Batran Salah-Eddin , Rafiyan Mohammad-Reza , Atmaca Akin , Neumann Antje , Karbach Julia , Bender Armin , Weidmann Eckhart , Altmannsberger Hans-Michael , Knuth Alexander , Jäger Elke
Published: September 2005

The infiltration of tumors by T cells has been shown to correlate with prolonged patients' survival. However, it remains unclear why only some tumors are infiltrated with T cells. This study was designed to investigate possible correlations between intratumoral T-cell infiltrates and the expression of cancer-associated antigens and MHC class I and II molecules in patients with melanoma. Fresh frozen samples from 124 stage IV melanoma patients were analyzed by immunohistochemistry for the expression of Melan-A/MART-1, tyrosinase, gp100, NY-ESO-1, and MHC class I and II. Intratumoral T-cell and B-cell infiltrates were detected by staining with anti-CD4, anti-CD8, anti-CD3, and L26 antibodies. The NY-ESO-1 serum antibody status was assessed by Western blot analysis. Intratumoral CD8+ and CD4+ T cells were detected in 63.9% and 71.3% of patients, respectively. We observed a significant heterogeneity of the expression of the melanocyte differentiation antigens, NY-ESO-1, and MHC class I and II molecules. The only significant correlation was found between the expression of MHC class I and the presence of CD4+ and CD8+ T cells (P < 0.0001). There was a strong association between these two variables with respect to the density and distribution of infiltrating T cells and the pattern of MHC class I expression (focal versus homogenous). Intratumoral T-cell infiltration is closely correlated with the MHC class I expression but not with the expression of differentiation antigens, cancer-associated antigens, or MHC class II molecules. These results may have implications for the definition of prognostic variables and for the identification of patients who may benefit from antigen-specific cancer immunotherapy.
Cloning and Characterization of the Novel Chimeric Gene TEL/PTPRR in Acute Myelogenous Leukemia with inv(12)(p13q13)
Authors: Nakamura Fumihiko , Nakamura Yuichi , Maki Kazuhiro , Sato Yuko , Mitani Kinuko
Published: September 2005

We have cloned a novel TEL/protein tyrosine phosphatase receptor-type R (PTPRR) chimeric gene generated by inv(12)(p13q13). PTPRR is the first protein tyrosine phosphatase identified as a fusion partner of TEL. The chimeric gene fused exon 4 of the TEL gene with exon 7 of the PTPRR gene, and produced 10 isoforms through alternative splicing. Two isoforms that were expressed at the highest level in the leukemic cells could have been translated into COOH-terminally truncated TEL protein possessing the helix-loop-helix domain (tTEL) and TEL/PTPRR chimeric protein linking the helix-loop-helix domain of TEL to the catalytic domain of PTPRR. These two mutant proteins exerted a dominant-negative effect over transcriptional repression mediated by wild-type TEL, although they themselves did not show any transcriptional activity. Heterodimerization with wild-type TEL might be an underlying mechanism in this effect. TEL/PTPRR did not exhibit any tyrosine phosphatase activity. Importantly, overexpression of TEL/PTPRR in granulocyte macrophage colony-stimulating factor–dependent UT7/GM cells resulted in their factor-independent proliferation, whereas overexpression of tTEL did not. After cytokine depletion, phosphorylated signal transducers and activators of transcription 3 (STAT3) significantly declined in mock cells, but remained in both tTEL- and TEL/PTPRR-overexpressing cells. Loss of tumor suppressive function of wild-type TEL and maintenance of STAT3-mediated signal could at least partly contribute to the leukemogenesis caused by inv(12)(p13q13).
Differences in Glycosylation Patterns of Heat Shock Protein, gp96: Implications for Prostate Cancer Prevention
Authors: Suriano Robert , Ghosh Salil K. , Ashok Badithe T. , Mittelman Abraham , Chen Yuangen , Banerjee Asesh , Tiwari Raj K.
Published: September 2005

Heat shock protein gp96 induces a tumor-specific protective immunity in a variety of experimental tumor models. Because the primary sequences of the glycoprotein, gp96 are identical between tumor and normal tissues, the peptides associated with gp96 and/or the posttranslational modifications of gp96, determine its immunogenicity. Gp96-associated peptides constitute the antigenic repertoire of the source tissue; thus, purified gp96-peptide complexes have clinical significance as autologous cancer vaccines. However, the role of altered glycosylation and its contribution in the biological as well as immunologic activity of gp96 still remains uncharacterized. We examined the cancer-specific glycosylation patterns of gp96. To this end, monosaccharide compositions of gp96 were compared between normal rat prostate and two cancerous rat prostate tissues, nonmetastatic/androgen-dependent Dunning G and metastatic/androgen-independent MAT-LyLu, as well as two human nonmetastatic prostate cancer cell lines, androgen-dependent LnCaP and androgen-independent DU145. Marked differences were observed between the gp96 monosaccharide compositions of the normal and cancerous tissues. Furthermore, gp96 molecules from more aggressive cellular transformations were found to carry decreasing quantities of several monosaccharides as well as sum total content of neutral and amino sugars. We believe that the unique glycosylation patterns contribute to cellular phenotype and that the posttranslational modifications of gp96 may affect its functional attributes.
BRCA2 Suppresses Cell Proliferation via Stabilizing MAGE-D1
Authors: Tian Xin-xia , Rai Deepak , Li Jun , Zou Chaozhong , Bai Yujie , Wazer David , Band Vimla , Gao Qingshen
Published: September 2005

Germ line mutations in BRCA2 gene predispose women to early-onset familial breast and ovarian cancer. BRCA2 is a protein of multiple functions. In addition to its role in DNA double-strand break repair, BRCA2 also plays a role in stabilization of stalled DNA replication forks, cytokinesis, transcription regulation, mammalian gametogenesis, centrosome duplication, and suppression of cell proliferation. However, how BRCA2 mutations predispose women specifically to breast and ovarian cancer remains undefined. Here we found that BRCA2 binds and stabilizes MAGE-D1, a member of the MAGE gene family of proteins. Expression of BRCA2 and MAGE-D1 synergistically suppresses cell proliferation independently of the p53 pathway. Using two MAGE-D1 RNA interferences and two cell lines expressing low or undetectable levels of MAGE-D1, we further showed that the expression of MAGE-D1 is required for BRCA2-mediated suppression of cell proliferation, indicating that MAGE-D1 is a downstream target of BRCA2 and that BRCA2 suppresses cell proliferation via stabilizing MAGE-D1. Importantly, MAGE-D1 protein expression was reduced in 6 of 16 breast carcinoma cell lines tested as compared with untransformed immortal mammary epithelial cell lines, suggesting that suppression of MAGE-D1 expression may be involved in the tumorigenesis of a subset of sporadic breast cancers.
A Monoclonal Antibody that Binds Anionic Phospholipids on Tumor Blood Vessels Enhances the Antitumor Effect of Docetaxel on Human Breast Tumors in Mice
Authors: Huang Xianming , Bennett Mary , Thorpe Philip E.
Published: September 2005

Anionic phospholipids, principally phosphatidylserine, become exposed on the external surface of viable vascular endothelial cells in tumors, providing an excellent marker for tumor vascular targeting. We recently raised an IgG monoclonal antibody, 3G4, which binds to anionic phospholipids in a β2-glycoprotein I–dependent manner. It inhibited tumor growth in a variety of rodent tumor models by stimulating antibody-dependent cellular cytotoxicity toward tumor vessels. In the present study, we tested the hypothesis that docetaxel, which is known to have antivascular effects on tumors, might induce exposure of anionic phospholipids on tumor vasculature and, thus, enhance the antitumor activity of 3G4. Treatment of human umbilical vascular endothelial cells with subtoxic concentrations of docetaxel (20 pmol/L) in vitro caused anionic phospholipids to be externalized without inducing apoptosis. Docetaxel treatment of mice increased the percentage of tumor vessels that expose anionic phospholipids from 35% to 60%. No induction of phosphatidylserine was observed on vessels in normal tissues even after systemic treatment with docetaxel. Treatment of mice bearing orthotopic MDA-MB-435 human breast tumors with 3G4 plus docetaxel inhibited tumor growth by 93%. Treatment of mice bearing disseminated MDA-MB-435 tumors with 3G4 plus docetaxel reduced the average number of tumor colonies in the lungs by 93% and half the animals did not develop tumors. In both tumor models, the antitumor effect of the combination was statistically superior (P < 0.01) to that of docetaxel or 3G4 alone. Combination therapy reduced the tumor vessel density and plasma volume in tumors to a greater extent than did the individual drugs. The combination therapy was no more toxic to the mice than was docetaxel alone. These results indicate that, as an adjuvant therapy, 3G4 could enhance the therapeutic efficacy of docetaxel in breast cancer patients.
The Ability of E1A to Rescue ras-Induced Premature Senescence and Confer Transformation Relies on Inactivation of Both p300/CBP and Rb Family Proteins
Authors: Deng Qingdong , Li Yilei , Tedesco Donato , Liao Rong , Fuhrmann Gerhard , Sun Peiqing
Published: September 2005

In primary cells, oncogenic ras induces a stable growth arrest known as premature senescence. Ras-induced premature senescence is considered as a tumor-suppressing defense response that needs to be bypassed before oncogenic potential ras can be revealed. To gain insights into the mechanism of senescence bypass during oncogenic transformation, we dissected the activities of an adenoviral oncoprotein E1A, which is capable of overcoming ras-induced senescence. Our results have indicated that the senescence bypassing activity resides in the NH2 terminus and requires both Rb-binding and p300/CBP-binding functions of E1A. Although interference with the p16INK4A/Rb pathway or inactivation of p300/CBP alone did not lead to senescence bypass, these two types of genetic alterations complemented the Rb-binding defective and the p300/CBP-binding defective mutants of E1A, respectively, to rescue premature senescence. Therefore, genetic alterations disrupting the p16INK4A/Rb pathway or the p300/CBP functions both contribute to the bypass of senescence. We further showed that p300/CBP were essential for ras-induced p53 activity, providing a potential mechanism underlying the important role of p300/CBP in senescence. Furthermore, p300/CBP inactivation led to cellular transformation in cooperation with the p300/CBP-binding defective E1A mutants, MDM2 and Ha-RasV12. These results have shown that p300 and CBP are integral components of the pathway that mediates ras-induced senescence. The critical role of p300 and CBP in the senescence response that limits the oncogenic potential of ras has provided a mechanistic basis for the tumor-suppressing function of these proteins.
1′-Acetoxychavicol Acetate Is a Novel Nuclear Factor κB Inhibitor with Significant Activity against Multiple Myeloma In vitro and In vivo
Authors: Ito Keisuke , Nakazato Tomonori , Xian Ming Ji , Yamada Taketo , Hozumi Nobumichi , Murakami Akira , Ohigashi Hajime , Ikeda Yasuo , Kizaki Masahiro
Published: September 2005

1′-Acetoxychavicol acetate (ACA) is a component of a traditional Asian condiment obtained from the rhizomes of the commonly used ethno-medicinal plant Languas galanga. Here, we show for the first time that ACA dramatically inhibits the cellular growth of human myeloma cells via the inhibition of nuclear factor κB (NF-κB) activity. In myeloma cells, cultivation with ACA induced G0-G1 phase cell cycle arrest, followed by apoptosis. Treatment with ACA induced caspase 3, 9, and 8 activities, suggesting that ACA-induced apoptosis in myeloma cells mediates both mitochondrial- and Fas-dependent pathways. Furthermore, we showed that ACA significantly inhibits the serine phosphorylation and degradation of IκBα. ACA rapidly decreased the nuclear expression of NF-κB, but increased the accumulation of cytosol NF-κB in RPMI8226 cells, indicating that ACA inhibits the translocation of NF-κB from the cytosol to the nucleus. To evaluate the effects of ACA in vivo, RPMI8226-transplanted NOD/SCID mice were treated with ACA. Tumor weight significantly decreased in the ACA-treated mice compared with the control mice. In conclusion, ACA has an inhibitory effect on NF-κB, and induces the apoptosis of myeloma cells in vitro and in vivo. ACA, therefore, provides a new biologically based therapy for the treatment of multiple myeloma patients as a novel NF-κB inhibitor.
Preclinical Antilymphoma Activity of a Humanized Anti-CD40 Monoclonal Antibody, SGN-40
Authors: Law Che-Leung , Gordon Kristine A. , Collier John , Klussman Kerry , McEarchern Julie A. , Cerveny Charles G. , Mixan Bruce J. , Lee Wyne P. , Lin Zhonghau , Valdez Patricia , Wahl Alan F. , Grewal Iqbal S.
Published: September 2005

SGN-40 is a humanized IgG1 antihuman CD40 that is currently in a phase I clinical trial for the treatment of multiple myeloma. As surface CD40 expression on B-lineage cells is maintained from pro-B cells to plasma cells, SGN-40 may be applicable to treatment of other B-cell neoplasias, including non-Hodgkin's lymphoma. In this study, we examined potential in vitro and in vivo anti–B-lineage lymphoma activity of SGN-40. Recombinant SGN-40 was expressed and purified from Chinese hamster ovary cells and characterized based on binding affinity, specificity, and normal B-cell stimulation. The ability of SGN-40 to target neoplastic B cells was examined in vitro by proliferation inhibition, cytotoxicity, and antibody-dependent cell cytotoxicity assays and in vivo by human lymphoma xenograft models. Recombinant SGN-40 showed high affinity, Kd of ∼1 nmol/L, and specific binding to CD40. Whereas SGN-40 was a weak agonist in stimulating normal B-cell proliferation in the absence of IL-4 and CD40L, it delivered potent proliferation inhibitory and apoptotic signals to, and mediated antibody-dependent cytotoxicity against, a panel of high-grade B-lymphoma lines. These in vitro antilymphoma effects were extended to disseminated and s.c. xenograft CD40 tumor models. In these xenograft models, the antitumor activity of SGN-40 was comparable with that of rituximab. The preclinical in vitro and in vivo antilymphoma activity of SGN-40 observed in this study provides a rationale for the clinical testing of SGN-40 in the treatment of CD40+ B-lineage lymphomas.
Implication of Galectin-3 in Wnt Signaling
Authors: Shimura Tatsuo , Takenaka Yukinori , Fukumori Tomoharu , Tsutsumi Soichi , Okada Kohji , Hogan Victor , Kikuchi Akira , Kuwano Hiroyuki , Raz Avraham
Published: September 2005

Galectin-3 (gal-3), a member of the β-galactoside–binding proteins family, was identified as a binding partner of β-catenin. Analysis of the human gal-3 sequence reveled a structural similarity to β-catenin as it also contains the consensus sequence (S92XXXS96) for glycogen synthase kinase-3β (GSK-3β) phosphorylation and can serve as its substrate. In addition, Axin, a regulator protein of Wnt that complexes with β-catenin, also binds gal-3 using the same sequence motif identified here by a deletion mutant analysis. The data presented here give credence to the suggestion that gal-3 is a key regulator in the Wnt/β-catenin signaling pathway and highlight the functional similarities between gal-3 and β-catenin.
MicroRNA-21 Is an Antiapoptotic Factor in Human Glioblastoma Cells
Authors: Chan Jennifer A. , Krichevsky Anna M. , Kosik Kenneth S.
Published: September 2005

MicroRNAs (miRNAs) are small noncoding RNA molecules that regulate protein expression by targeting the mRNA of protein-coding genes for either cleavage or repression of translation. The roles of miRNAs in lineage determination and proliferation as well as the location of several miRNA genes at sites of translocation breakpoints or deletions has led to the speculation that miRNAs could be important factors in the development or maintenance of the neoplastic state. Here we show that the highly malignant human brain tumor, glioblastoma, strongly overexpresses a specific miRNA, miR-21. Our studies show markedly elevated miR-21 levels in human glioblastoma tumor tissues, early-passage glioblastoma cultures, and in six established glioblastoma cell lines (A172, U87, U373, LN229, LN428, and LN308) compared with nonneoplastic fetal and adult brain tissues and compared with cultured nonneoplastic glial cells. Knockdown of miR-21 in cultured glioblastoma cells triggers activation of caspases and leads to increased apoptotic cell death. Our data suggest that aberrantly expressed miR-21 may contribute to the malignant phenotype by blocking expression of critical apoptosis-related genes.
Pancreatic Cancer Cell Radiation Survival and Prenyltransferase Inhibition: The Role of K-Ras
Authors: Brunner Thomas B. , Cengel Keith A. , Hahn Stephen M. , Wu Junmin , Fraker Douglas L. , McKenna W. Gillies , Bernhard Eric J.
Published: September 2005

Activating K-ras mutations are found in ∼90% of pancreatic carcinomas and may contribute to the poor prognosis of these tumors. Because radiotherapy is frequently used in pancreatic cancer treatment, we assessed the contribution of oncogenic K-ras signaling to pancreatic cancer radiosensitivity. Seven human pancreatic carcinoma lines with activated K-ras and two cell lines with wild-type ras were used to examine clonogenic cell survival after Ras inhibition. Ras inhibition was accomplished by small interfering RNA (siRNA) knockdown of K-ras expression and by blocking Ras processing using a panel of prenyltransferase inhibitors of differing specificity for the two prenyltransferases that modify K-Ras.

K-ras knockdown by siRNA or inhibition of prenyltransferase activity resulted in radiation sensitization in vitro and in vivo in tumors with oncogenic K-ras mutations. Inhibition of farnesyltransferase alone was sufficient to radiosensitize most K-ras mutant tumors, although K-Ras prenylation was not blocked. These results show that inhibition of activated K-Ras can promote radiation killing of pancreatic carcinoma in a superadditive manner. The finding that farnesyltransferase inhibition alone radiosensitizes tumors with K-ras mutations implies that a farnesyltransferase inhibitor–sensitive protein other than K-Ras may contribute to survival in the context of mutant K-ras. Farnesyltransferase inhibitors could therefore be of use as sensitizers for pancreatic carcinoma radiotherapy.
Silencing of Epidermal Growth Factor Receptor Suppresses Hypoxia-Inducible Factor-2–Driven VHL−/− Renal Cancer
Authors: Smith Karlene , Gunaratnam Lakshman , Morley Melissa , Franovic Aleksandra , Mekhail Karim , Lee Stephen
Published: September 2005

Inactivating mutations in the von Hippel-Lindau (VHL) tumor suppressor gene are associated with clear cell renal cell carcinoma (VHL−/− RCC), the most frequent malignancy of the human kidney. The VHL protein targets the α subunits of hypoxia-inducible factor (HIF) transcription factor for ubiquitination and degradation. VHL−/− RCC cells fail to degrade HIF resulting in the constitutive activation of its target genes, a process that is required for tumorigenesis. We recently reported that HIF activates the transforming growth factor-α/epidermal growth factor receptor (TGF-α/EGFR) pathway in VHL-defective RCC cells. Here, we show that short hairpin RNA (shRNA)–mediated inhibition of EGFR is sufficient to abolish HIF-dependent tumorigenesis in multiple VHL−/− RCC cell lines. The 2α form of HIF (HIF-2α), but not HIF-1α, drives in vitro and in vivo tumorigenesis of VHL−/− RCC cells by specifically activating the TGF-α/EGFR pathway. Transient incubation of VHL−/− RCC cell lines with small interfering RNA directed against EGFR prevents autonomous growth in two-dimensional culture as well as the ability of these cells to form dense spheroids in a three-dimensional in vitro tumor assay. Stable expression of shRNA against EGFR does not alter characteristics associated with VHL loss including constitutive production of HIF targets and defects in fibronectin deposition. In spite of this, silencing of EGFR efficiently abolishes in vivo tumor growth of VHL loss RCC cells. These data identify EGFR as a critical determinant of HIF-2α-dependent tumorigenesis and show at the molecular level that EGFR remains a credible target for therapeutic strategies against VHL−/− renal carcinoma.
Rho GDP Dissociation Inhibitor Protects Cancer Cells against Drug-Induced Apoptosis
Authors: Zhang Baolin , Zhang Yaqin , Dagher Marie-Claire , Shacter Emily
Published: September 2005

Rho GDP dissociation inhibitor (RhoGDI) plays an essential role in control of a variety of cellular functions through interactions with Rho family GTPases, including Rac1, Cdc42, and RhoA. RhoGDI is frequently overexpressed in human tumors and chemoresistant cancer cell lines, raising the possibility that RhoGDI might play a role in the development of drug resistance in cancer cells. We found that overexpression of RhoGDI increased resistance of cancer cells (MDA-MB-231 human breast cancer cells and JLP-119 lymphoma cells) to the induction of apoptosis by two chemotherapeutic agents: etoposide and doxorubicin. Conversely, silencing of RhoGDI expression by DNA vector–mediated RNA interference (small interfering RNA) sensitized MDA-MB-231 cells to drug-induced apoptosis. Resistance to apoptosis was restored by reintroduction of RhoGDI protein expression. The mechanism for the antiapoptotic activity of RhoGDI may derive from its ability to inhibit caspase-mediated cleavage of Rac1 GTPase, which is required for maximal apoptosis to occur in response to cytotoxic drugs. Taken together, the data show that RhoGDI is an antiapoptotic molecule that mediates cellular resistance to these chemotherapy agents.
Overexpression of the Tumor Suppressor Gene Phosphatase and Tensin Homologue Partially Inhibits Wnt-1–Induced Mammary Tumorigenesis
Authors: Zhao Hong , Cui Yongzhi , Dupont Joelle , Sun Hui , Hennighausen Lothar , Yakar Shoshana
Published: September 2005

The tumor suppressor phosphatase and tensin homologue (PTEN) is involved in cell proliferation, adhesion, and apoptosis. PTEN overexpression in mammary epithelium leads to reduced cell number and impaired differentiation and secretion. In contrast, overexpression of the proto-oncogene Wnt-1 in mammary epithelium leads to mammary hyperplasia and subsequently focal mammary tumors. To explore the possibility that PTEN intersects with Wnt-induced tumorigenesis, mice that ectopically express PTEN and Wnt-1 in mammary epithelium were generated. PTEN overexpression resulted in an 11% reduction of Wnt-1–induced tumors within a 12-month period and the onset of tumors was delayed from an average of 5.9 to 7.7 months. The rate of tumor growth, measured from 0.5 cm diameter until the tumors reached 1.0 cm diameter, was increased from 8.4 days in Wnt-1 mice to 17.7 days in Wnt-1 mice overexpressing PTEN. Here we show for the first time in vivo that overexpression of PTEN in the Wnt-1 transgenic mice resulted in a marked decrease in the insulin-like growth factor (IGF)-I receptor levels leading to a reduced IGF-I–mediated mitogenesis. Moreover, the percentage of BrdUrd-positive epithelial nuclei was decreased by 48%. β-Catenin immunoreactivity was significantly decreased and the percentage of signal transducer and activator of transcription 5a (stat5a)–positive mammary epithelial cells was increased by 2-fold in Wnt-1 mice overexpressing PTEN. The present study shows that PTEN can partially inhibit the Wnt-1–induced mammary tumorigenesis in early neoplastic stages by blocking the AKT pathway and by reducing the IGF-I receptor levels in mammary gland. This study identifies the PTEN as a therapeutic target for the treatment of mammary cancer and presumably other types of cancer.
A Novel Peptide Specifically Binding to Interleukin-6 Receptor (gp80) Inhibits Angiogenesis and Tumor Growth
Authors: Su Jen-Liang , Lai Kuo-Pao , Chen Chi-An , Yang Ching-Yao , Chen Pei-Sheng , Chang Chiao-Chia , Chou Chia-Hung , Hu Chi-Lun , Kuo Min-Liang , Hsieh Chang-Yao , Wei Lin-Hung
Published: September 2005

Experimental and clinical findings support the essential role of interleukin (IL)-6 in the pathogenesis of various human cancers and provide a rationale for targeted therapeutic investigations. A novel peptide, S7, which selectively binds to IL-6 receptor (IL-6R) α chain (gp80) and broadly inhibits IL-6-mediated events, was identified using phage display library screening. The synthetic S7 peptide specifically bound to soluble IL-6R as well as cognate human IL-6Rα, resulting in a dose-dependent blockade of the interaction between IL-6 and IL-6Rα. S7 peptide prevents IL-6–mediated survival signaling and sensitizes cervical cancer cells to chemotherapeutic compounds in vitro. The in vitro analysis of antiangiogenic activity showed that S7 peptide substantially inhibits IL-6–induced vascular endothelial growth factor-A expression and angiogenesis in different cancer cell lines. Furthermore, S7 peptide was bioavailable in vivo, leading to a significant suppression of IL-6–induced vascular endothelial growth factor–mediated cervical tumor growth in severe combined immunodeficient mice. These observations show the feasibility of targeting IL-6/IL-6R interaction using the small peptide and highlight its potential in the clinical applications.
A Five-Amino-Acid Peptide Blocks Met- and Ron-Dependent Cell Migration
Authors: Matzke Alexandra , Herrlich Peter , Ponta Helmut , Orian-Rousseau Véronique
Published: September 2005

Various human cancers express elevated levels of the receptor tyrosine kinases Met or Ron and v6-containing isoforms of CD44. The activation of Met and Ron requires the presence of such CD44 v6-containing isoforms that act as coreceptors. Three amino acids within the v6 sequence were identified by mutational analysis to be essential for the coreceptor function: EWQ in the rat sequence and RWH in human. Peptides comprising these three amino acids (the smallest containing only five amino acids) efficiently act as competitors and block ligand-dependent activation of Met or Ron and subsequent cell migration.
Targeted Inhibition of the KLF6 Splice Variant, KLF6 SV1, Suppresses Prostate Cancer Cell Growth and Spread
Authors: Narla Goutham , DiFeo Analisa , Yao Shen , Banno Asoka , Hod Eldad , Reeves Helen L. , Qiao Rui F. , Camacho-Vanegas Olga , Levine Alice , Kirschenbaum Alexander , Chan Andrew M. , Friedman Scott L. , Martignetti John A.
Published: September 2005

Prostate cancer is a leading cause of cancer death in men. Risk prognostication, treatment stratification, and the development of rational therapeutic strategies lag because the molecular mechanisms underlying the initiation and progression from primary to metastatic disease are unknown. Multiple lines of evidence now suggest that KLF6 is a key prostate cancer tumor suppressor gene including loss and/or mutation in prostate cancer tumors and cell lines and decreased KLF6 expression levels in recurrent prostate cancer samples. Most recently, we identified a common KLF6 germ line single nucleotide polymorphism that is associated with an increased relative risk of prostate cancer and the increased production of three alternatively spliced, dominant-negative KLF6 isoforms. Here we show that although wild-type KLF6 (wtKLF6) acts as a classic tumor suppressor, the single nucleotide polymorphism-increased splice isoform, KLF6 SV1, displays a markedly opposite effect on cell proliferation, colony formation, and invasion. In addition, whereas wtKLF6 knockdown increases tumor growth in nude mice >2-fold, short interfering RNA–mediated KLF6 SV1 inhibition reduces growth by ∼50% and decreases the expression of a number of growth- and angiogenesis-related proteins. Together, these findings begin to highlight a dynamic and functional antagonism between wtKLF6 and its splice variant KLF6 SV1 in tumor growth and dissemination.
Potent Modulation of Intestinal Tumorigenesis in Apcmin/+ Mice by the Polyamine Catabolic Enzyme Spermidine/Spermine N1-acetyltransferase
Authors: Tucker Jody M. , Murphy John T. , Kisiel Nicholas , Diegelman Paula , Barbour Karen W. , Davis Celestia , Medda Moussumi , Alhonen Leena , Jänne Juhani , Kramer Debora L. , Porter Carl W. , Berger Franklin G.
Published: September 2005

Intracellular polyamine pools are homeostatically maintained by processes involving biosynthesis, catabolism, and transport. Although most polyamine-based anticancer strategies target biosynthesis, we recently showed that activation of polyamine catabolism at the level of spermidine/spermine N1-acetyltransferase-1 (SSAT) suppresses tumor outgrowth in a mouse prostate cancer model. Herein, we examined the effects of differential SSAT expression on intestinal tumorigenesis in the ApcMin/+ (MIN) mouse. When MIN mice were crossed with SSAT-overproducing transgenic mice, they developed 3- and 6-fold more adenomas in the small intestine and colon, respectively, than normal MIN mice. Despite accumulation of the SSAT product, N1-acetylspermidine, spermidine and spermine pools were only slightly decreased due to a huge compensatory increase in polyamine biosynthetic enzyme activities that gave rise to enhanced metabolic flux. When MIN mice were crossed with SSAT knock-out mice, they developed 75% fewer adenomas in the small intestine, suggesting that under basal conditions, SSAT contributes significantly to the MIN phenotype. Despite the loss in catabolic capability, tumor spermidine and spermine pools failed to increase significantly due to a compensatory decrease in biosynthetic enzyme activity giving rise to a reduced metabolic flux. Loss of heterozygosity at the Apc locus was observed in tumors from both SSAT-transgenic and -deficient MIN mice, indicating that loss of heterozygosity remained the predominant oncogenic mechanism. Based on these data, we propose a model in which SSAT expression alters flux through the polyamine pathway giving rise to metabolic events that promote tumorigenesis. The finding that deletion of SSAT reduces tumorigenesis suggests that small-molecule inhibition of the enzyme may represent a nontoxic prevention and/or treatment strategy for gastrointestinal cancers.
Meningioma Transcript Profiles Reveal Deregulated Notch Signaling Pathway
Authors: Cuevas Ileana C. , Slocum Alison L. , Jun Peter , Costello Joseph F. , Bollen Andrew W. , Riggins Gregory J. , McDermott Michael W. , Lal Anita
Published: September 2005

Meningiomas constitute the second most common central nervous system tumor, and yet relatively little is known about the molecular events that are important for the pathogenesis and malignant progression of these tumors. We have used serial analysis of gene expression to compare the transcriptomes of nonneoplastic meninges and meningiomas of all malignancy grades. A novel finding from this screen is the induction of three components of the Notch signaling pathway: the transcription factor, hairy and enhancer of Split1 (HES1) and two members of the Groucho/transducin-like enhancer of Split family of corepressors, TLE2 and TLE3. TLE corepressors interact and modulate the activity of a wide range of transcriptional regulatory systems, one of which is HES1. We have shown that the transcript and protein levels of HES1, the Notch2 and Notch1 receptors and the Jagged1 ligand are induced in meningiomas of all grades, whereas induction of TLE2 and TLE3 occurs specifically in higher-grade meningiomas. Meningioma cell lines express components of the Notch signaling pathway and an inhibitor of this pathway suppresses meningioma cell survival. These results suggest that deregulated expression of the Notch pathway is a critical event in meningioma pathogenesis and that modulation of this and potentially other signaling pathways by TLE corepressors leads to a more malignant phenotype.
In vitro Activity of Bcr-Abl Inhibitors AMN107 and BMS-354825 against Clinically Relevant Imatinib-Resistant Abl Kinase Domain Mutants
Authors: O'Hare Thomas , Walters Denise K. , Stoffregen Eric P. , Jia Taiping , Manley Paul W. , Mestan Jürgen , Cowan-Jacob Sandra W. , Lee Francis Y. , Heinrich Michael C. , Deininger Michael W.N. , Druker Brian J.
Published: September 2005

Imatinib, a Bcr-Abl tyrosine kinase inhibitor, is a highly effective therapy for patients with chronic myelogenous leukemia (CML). Despite durable responses in most chronic phase patients, relapses have been observed and are much more prevalent in patients with advanced disease. The most common mechanism of acquired imatinib resistance has been traced to Bcr-Abl kinase domain mutations with decreased imatinib sensitivity. Thus, alternate Bcr-Abl kinase inhibitors that have activity against imatinib-resistant mutants would be useful for patients who relapse on imatinib therapy. Two such Bcr-Abl inhibitors are currently being evaluated in clinical trials: the improved potency, selective Abl inhibitor AMN107 and the highly potent dual Src/Abl inhibitor BMS-354825. In the current article, we compared imatinib, AMN107, and BMS-354825 in cellular and biochemical assays against a panel of 16 kinase domain mutants representing >90% of clinical isolates. We report that AMN107 and BMS-354825 are 20-fold and 325-fold more potent than imatinib against cells expressing wild-type Bcr-Abl and that similar improvements are maintained for all imatinib-resistant mutants tested, with the exception of T315I. Thus, both inhibitors hold promise for treating imatinib-refractory CML.