bluuebunny/crossref_metadata_2025 huggingface dataset:
Variable expression of protein kinase Cε in human melanoma cells regulates sensitivity to TRAIL-induced apoptosis
Authors: Gillespie Susan , Zhang Xu Dong , Hersey Peter
Published: September 2005

Protein kinase C (PKC) activation is believed to protect against apoptosis induced by death receptors. We have found however that the effect of activation of PKC on tumor necrosis factor–related apoptosis-inducing ligand (TRAIL)–induced apoptosis of melanoma differs between cell lines. Pretreatment with phorbol 12-myristate 13-acetate (PMA) led to inhibition of apoptosis in the majority of the melanoma cell lines, but those with relatively low PKCε expression were sensitized to TRAIL-induced apoptosis. Introduction of PKCε into PKCε-low cell lines reversed sensitization of the cells to TRAIL-induced apoptosis by PMA. In contrast, a dominant-negative form of PKCε caused an increase in sensitivity. The changes in sensitivity to TRAIL-induced apoptosis were reflected in similar changes in conformation of Bax and its relocation from the cytosol to mitochondria. Similarly, there were concordant increases or decreases in mitochondrial release of second mitochondria-derived activator of caspase/DIABLO, activation of caspase-3, and processing of its substrates. Activation of PKC seemed to mediate its effects upstream of mitochondria but downstream of caspase-8 and Bid in that pretreatment with PMA did not cause significant changes in the expression levels of TRAIL death receptors, alterations in the levels of caspase-8 activation, or cleavage of Bid. PKC activated the antiapoptotic extracellular signal-regulated kinase 1/2 pathway, but inhibitors of this pathway only partially reversed the protective effect of PKC against TRAIL-induced apoptosis. These results provide further insights into the variable responses of melanoma to TRAIL-induced apoptosis and may help define responsive phenotypes to treatment of melanoma with TRAIL.
ZM336372, a Raf-1 activator, suppresses growth and neuroendocrine hormone levels in carcinoid tumor cells
Authors: Van Gompel Jamie J. , Kunnimalaiyaan Muthusamy , Holen Kyle , Chen Herbert
Published: September 2005

Neuroendocrine tumors, such as carcinoids, are highly metastatic neoplasms that secrete bioactive hormones resulting in carcinoid syndrome. Few curative treatments exist outside of surgical resection. We have previously shown that activation of the Raf-1 signaling pathway can suppress hormone production in carcinoid tumor cells. In this study, we investigated a novel treatment for carcinoid tumor cell growth based on pharmacologic Raf-1 activation using the compound ZM336372. Treatment of carcinoid tumor cells with ZM336372 resulted in progressive phosphorylation of Raf-1, mitogen-activated protein kinase 1/2, and extracellular signal–regulated kinase 1/2. Importantly, exposure to ZM336372 resulted in a significant reduction of bioactive hormone levels as well as the transcription factor, human achaete-scute homologue-1 in carcinoid tumor cells. Furthermore, treatment with ZM336372 led to a marked suppression of cellular proliferation and induction of the cell cycle inhibitors p21 and p18. In summary, ZM336372 targets both proliferation and palliative issues associated with carcinoid tumor cells, and therefore, warrants further investigation as a possible therapeutic strategy for patients with carcinoid tumors.
Proteasome inhibitor therapy in multiple myeloma
Authors: Chauhan Dharminder , Hideshima Teru , Mitsiades Constantine , Richardson Paul , Anderson Kenneth C.
Published: September 2005

Multiple myeloma remains incurable despite available therapies, and novel therapies that target both tumor cell and bone marrow microenvironment are urgently needed. Preclinical in vitro and in vivo studies show remarkable anti–multiple myeloma activity of the proteasome inhibitor bortezomib/PS-341 even in multiple myeloma cells refractory to multiple prior therapies, including dexamethasone, melphalan, and thalidomide. Based on these findings, the U.S. Food and Drug Administration recently approved the first proteasome inhibitor bortezomib (Velcade), formerly known as PS-341, for the treatment of relapsed/refractory multiple myeloma. Bortezomib therapy has set an outstanding example of translational research in the field of oncology. Genomics and proteomic studies further provide rationale for combining bortezomib with conventional and novel agents to inhibit multiple myeloma growth, overcome drug resistance, reduce attendant toxicity, and improve patient outcome in multiple myeloma.
Sapphyrins induce apoptosis in hematopoietic tumor–derived cell lines and show in vivo antitumor activity
Authors: Naumovski Louie , Ramos Jason , Sirisawad Mint , Chen Jun , Thiemann Patti , Lecane Philip , Magda Darren , Wang Zhong , Cortez Cecilia , Boswell Garry , Gyu Cho Dong , Sessler Jonathan , Miller Richard
Published: September 2005

Sapphyrins are pentapyrrolic, metal-free, expanded porphyrins. In the present study, the activity of sapphyrins as anticancer agents in hematopoietic-derived tumor cells was explored. It was found that a dihydroxylated water-soluble sapphyrin derivative (PCI-2000) is a potent inducer of apoptosis in a wide variety of tumor cell lines including lymphoma (Ramos, DHL-4, and HF-1), leukemia (Jurkat and HL-60), and myeloma (8226/S, 1-310, C2E3, and 1-414). PCI-2000 triggers an apoptotic pathway in these tumor cells as shown by release of cytochrome c from mitochondria; activation of caspases 9, 8, and 3; cleavage of the caspase substrate poly(ADP-ribose) polymerase; and Annexin V binding. Apoptosis can be partially inhibited by overexpression of the antiapoptotic protein Bcl-2 or treatment with benzyloxycarbonyl-valine-alanine-aspartic acid-fluoromethylketone, a cell-permeable caspase inhibitor. Both PCI-2000 and PCI-2010, a tetrahydroxy bis-carbamate derivative of PCI-2000, result in increased levels of phosphorylated p38 mitogen-activated protein kinase. Inhibition of p38 mitogen-activated protein kinase phosphorylation resulted in a synergistic increase of PCI-2000 cytotoxicity. PCI-2010 showed less toxicity in mice than PCI-2000 and was active in slowing the growth of Ramos and HL-60 tumor xenografts in nude mice. These results provide preclinical rationale for the further study of sapphyrins for potential use in the treatment of hematopoietic-derived tumors.
Matrix metalloproteinase–activated doxorubicin prodrugs inhibit HT1080 xenograft growth better than doxorubicin with less toxicity
Authors: Albright Charles F. , Graciani Nilsa , Han Wei , Yue Eddy , Stein Ross , Lai Zhihong , Diamond Melody , Dowling Randine , Grimminger Lisa , Zhang Shu-Yun , Behrens Davette , Musselman Amy , Bruckner Robert , Zhang Mingzhu , Jiang Xiang , Hu Daniel , Higley Anne , DiMeo Susan , Rafalski Maria , Mandlekar Sandya , Car Bruce , Yeleswaram Swamy , Stern Andrew , Copeland Robert A. , Combs Andrew , Seitz Steve P. , Trainor George L. , Taub Rebecca , Huang Pearl , Oliff Allen
Published: September 2005

Matrix metalloproteinase (MMP)–activated prodrugs were formed by coupling MMP-cleavable peptides to doxorubicin. The resulting conjugates were excellent in vitro substrates for MMP-2, -9, and -14. HT1080, a fibrosarcoma cell line, was used as a model system to test these prodrugs because these cells, like tumor stromal fibroblasts, expressed several MMPs. In cultured HT1080 cells, simple MMP-cleavable peptides were primarily metabolized by neprilysin, a membrane-bound metalloproteinase. MMP-selective metabolism in cultured HT1080 cells was obtained by designing conjugates that were good MMP substrates but poor neprilysin substrates. To determine how conjugates were metabolized in animals, MMP-selective conjugates were given to mice with HT1080 xenografts and the distribution of doxorubicin was determined. These studies showed that MMP-selective conjugates were preferentially metabolized in HT1080 xenografts, relative to heart and plasma, leading to 10-fold increases in the tumor/heart ratio of doxorubicin. The doxorubicin deposited by a MMP-selective prodrug, compound 6, was more effective than doxorubicin at reducing HT1080 xenograft growth. In particular, compound 6 cured 8 of 10 mice with HT1080 xenografts at doses below the maximum tolerated dose, whereas doxorubicin cured 2 of 20 mice at its maximum tolerated dose. Compound 6 was less toxic than doxorubicin at this efficacious dose because mice treated with compound 6 had no detectable changes in body weight or reticulocytes, a marker for marrow toxicity. Hence, MMP-activated doxorubicin prodrugs have a much higher therapeutic index than doxorubicin using HT1080 xenografts as a preclinical model.
Protein kinase C-α mediates epidermal growth factor receptor transactivation in human prostate cancer cells
Authors: Stewart Jubilee R. , O'Brian Catherine A.
Published: September 2005

Progression of human prostate cancer to a malignancy that is refractory to androgen-ablation therapy renders the disease resistant to available treatment options and accounts for the high prostate cancer mortality rate. Epidermal growth factor receptor (EGFR) expression in human prostate cancer specimens increases with disease progression to androgen-refractory prostate cancer, and experimental models implicate EGFR-dependent signaling to Erk1/2 activation in the androgen-refractory prostate cancer phenotype. 12-O-Tetradecanoylphorbol-13-acetate (TPA)-induced Erk1/2 activation in human prostate cancer PC-3 cells is a paradigm of diacylglycerol-induced EGFR transactivation in androgen-independent prostate cancer. In this report, we establish an obligatory role for TPA-induced protein kinase C (PKC)-α activation in EGFR transactivation and signaling to Erk1/2 activation in PC-3 cells. TPA-regulated molecules include PKCs, PKDs, and Ras guanyl nucleotide-releasing proteins. The PKC-selective inhibitors GF109203X and Gö6983 each blocked TPA-induced EGFR transactivation, indicating a requirement for PKC. PC-3 cells express four PKC isozymes. Prolonged bryostatin 1 treatment abrogated PKCα expression without altering expression levels of the other PKC isozymes. Pharmacologic PKCα “knockdown” abrogated TPA-induced Erk1/2 activation without affecting the EGF/EGFR-induced response, indicating that PKCα was required for EGFR transactivation but dispensable for signaling of ligand-activated EGFR to Erk1/2 activation. We corroborated this by showing that Gö6976, which is a PKCα-selective inhibitor in PC-3 cells, likewise abolished TPA-induced Erk1/2 activation and did not inhibit EGF/EGFR-induced Erk1/2 activation. Gö6976 had similar effects in DU145 cells, providing evidence for a common PKCα-dependent Erk1/2 activation mechanism in androgen-independent human prostate cancer cells of distinct genetic origin. These results constitute a rational basis for selective PKCα inhibition as a modality of prostate cancer therapy.
The immunocytokine scFv23/TNF sensitizes HER-2/neu–overexpressing SKBR-3 cells to tumor necrosis factor (TNF) via up-regulation of TNF receptor-1
Authors: Lyu Mi-Ae , Rosenblum Michael G.
Published: September 2005

Overexpression of HER-2/neu confers cellular resistance to tumor necrosis factor (TNF)–mediated cytotoxicity to SKBR-3 breast cancer cell lines. To understand the correlation between HER-2/neu expression and TNF resistance, we examined the unique signaling pathways associated with the cytotoxic effects of the immunocytokine scFv23/TNF, recombinant single-chain antibody fusion constructs containing TNF and targeting HER-2/neu, in TNF-resistant SKBR-3-LP cells. We found that treatment of HER-2/neu–overexpressing SKBR-3-LP cells with scFv23/TNF resulted in a 5- to 7-fold higher level of TNF receptor-1 expression 48 hours after exposure. In addition, treatment of SKBR-3-LP cells with scFv23/TNF resulted in down-regulation of Akt phosphorylation and induced apoptosis through cleavage of caspase-8, caspase-3, and poly(ADP-ribose) polymerase. ScFv23/TNF-induced cytotoxicity was inhibited by blocking of the binding of the TNF component of scFv23/TNF to TNF receptor-1 and was dependent on activation of caspase-8 and caspase-3. These results indicate that the immunocytokine scFv23/TNF sensitizes TNF-resistant HER-2/neu–overexpressing SKBR-3-LP cells to TNF-induced apoptosis via the overexpression of TNF receptor-1 and suggest that the overexpression of TNF receptor-1 plays a crucial role in TNF sensitivity in HER-2/neu–overexpressing cancer cells. ScFv23/TNF targeting the HER-2/neu may be an effective cytotoxic agent against HER-2/neu–overexpressing cancer cells, which are inherently resistant to TNF.
Effects of a novel immunomodulating agent, FTY720, on tumor growth and angiogenesis in hepatocellular carcinoma
Authors: Ho Joanna W.Y. , Man Kwan , Sun Chris K. , Lee Terence K. , Poon Ronnie T.P. , Fan Sheung Tat
Published: September 2005

In this study, we aimed to evaluate the potential anticancer and antiangiogenic effects of FTY720 on hepatocellular carcinoma. In vitro, chemosensitivity was tested on hepatoma cells, nontumorigenic, immortalized hepatocyte cells, as well as human umbilical vein endothelial cells (HUVEC). Moreover, effect of FTY720 on cell cycle and apoptosis was analyzed. In addition, a number of angiogenesis-associated assays were carried out. The in vivo effect of the drug on hepatocellular carcinoma tumor growth on nude mice was studied. Tissues obtained were analyzed in terms of proliferation, apoptosis, tumor microvessel density, and tumor vascular permeability. Compared with the MIHA cells, the hepatoma cell lines as well as HUVECs were found to be highly sensitive to the drugs in the aspect that FTY720 could induce G1 arrest and apoptosis in the hepatoma cells. Furthermore, FTY720 significantly decreased invasion, migration, and capillary tube formation of HUVECs at very low doses. In vivo study showed that tumor growth was significantly suppressed in the FTY720-treated animals, and staining of the tissue sections showed decreased tumor cell proliferation and increased tumor cell apoptosis in the treatment groups. Interestingly, significant reductions in tumor microvessel density and tumor vascular permeability were also found in the FTY720-treated groups. In conclusion, FTY720 not only shows potent antiangiogenic effects but is also cytotoxic toward hepatoma cells. Results from our preclinical study suggest that FTY720 can be selected as a good candidate for the treatment of hepatocellular carcinoma.
The sesquiterpene lactone parthenolide in combination with docetaxel reduces metastasis and improves survival in a xenograft model of breast cancer
Authors: Sweeney Christopher J. , Mehrotra Sanjana , Sadaria Miral R. , Kumar Suresh , Shortle Nicholas H. , Roman Yaritzabel , Sheridan Carol , Campbell Robert A. , Murry Daryl J. , Badve Sunil , Nakshatri Harikrishna
Published: September 2005

Parthenolide, a sesquiterpene lactone, shows antitumor activity in vitro, which correlates with its ability to inhibit the DNA binding of the antiapoptotic transcription factor nuclear factor κB (NF-κB) and activation of the c-Jun NH2-terminal kinase. In this study, we investigated the chemosensitizing activity of parthenolide in vitro as well as in MDA-MB-231 cell–derived xenograft metastasis model of breast cancer. HBL-100 and MDA-MB-231 cells were used to measure the antitumor and chemosensitizing activity of parthenolide in vitro. Parthenolide was effective either alone or in combination with docetaxel in reducing colony formation, inducing apoptosis and reducing the expression of prometastatic genes IL-8 and the antiapoptotic gene GADD45β1 in vitro. In an adjuvant setting, animals treated with parthenolide and docetaxel combination showed significantly enhanced survival compared with untreated animals or animals treated with either drug. The enhanced survival in the combination arm was associated with reduced lung metastases. In addition, nuclear NF-κB levels were lower in residual tumors and lung metastasis of animals treated with parthenolide, docetaxel, or both. In the established orthotopic model, there was a trend toward slower growth in the parthenolide-treated animals but no statistically significant findings were seen. These results for the first time reveal the significant in vivo chemosensitizing properties of parthenolide in the metastatic breast cancer setting and support the contention that metastases are very reliant on activation of NF-κB.
Matrix metalloproteinase inhibitor MMI-166 inhibits lymphogenous metastasis in an orthotopically implanted model of lung cancer
Authors: Fujino Haruhiko , Kondo Kazuya , Ishikura Hisashi , Maki Hideo , Kinoshita Hidetaka , Miyoshi Takanori , Takahashi Yuji , Sawada Naruhiko , Takizawa Hiromitsu , Nagao Taeko , Sakiyama Shoji , Monden Yasumasa
Published: September 2005

Matrix metalloproteinases (MMP) are considered to be critically involved in tumor invasion and the metastasis of various cancers. MMI-166 is a selective inhibitor of matrix metalloproteinase (MMP-2, MMP-9, and MMP-14). The purpose of this study was to evaluate the effects of MMI-166 on both the growth of the implanted tumor and the lymph node metastasis of the mediastinum and prolonging the life span, using an orthotopic implantation model of the Ma44-3 cancer cell line. We examined the anti-invasive effect of MMI-166 in lung cancer cell lines using an in vitro invasion assay. Next, we examined the anticancer effect of MMI-166 in vivo. MMI-166 (200 mg/kg body weight) or a vehicle was administered orally to the orthotopically implanted lung cancer model. MMI-166 dose-dependently inhibited the invasion of cancer cell lines with expressions of MMP-2 and/or MMP-9 in vitro. In vivo, MMI-166 significantly inhibited mediastinal lymph node metastasis in this orthotopic model (weight of the mediastinum: control, 0.089 ± 0.009 versus MMI-166, 0.069 ± 0.008 mg; P = 0.005; metastatic area: control, 93,495 ± 55,747 versus MMI-166, 22,747 ± 17,478 pixels; P = 0.045). MMI-166 prolonged the life span by 6 days in median survival time in the orthotopically implanted model (P = 0.039). These results showed that MMI-166 could possibly inhibit lymph node metastasis and prolong the life span in lung cancer patients.
Identification of genes and molecular pathways involved in the progression of premalignant oral epithelia
Authors: Banerjee Abhijit G. , Bhattacharyya Indraneel , Vishwanatha Jamboor K.
Published: September 2005

An early interventional effort in oral premalignancy requires novel molecular targets and diagnostic biomarkers to delay or reverse incidences of malignant progression. Microarray-based transcriptional profiling in disease states provides global insight into the causal biomolecular processes and novel pathways involved. In this study, we investigated transcript profiles in precancerous oral lesions to identify nearly 1,700 genes as significantly overexpressed or underexpressed and a primarily affected metabolic pathway that may be responsible for irreversible transition to progressive stages of oral cancer. For the first time, we show a convergence of several genes and pathways known for their oncogenic capabilities, in progression of premalignant oral epithelial tissues. This study consequently provides a molecular basis for persistent proinflammatory conditions in oral premalignant tissues. We found that lipocalin-type prostaglandin D2 synthase (PTGDS), a key enzyme in the arachidonic acid metabolism pathway, as repressed in premalignant stages. We show the protective role of these enzyme-derived metabolites in inhibiting cell proliferation using an in vitro oral cancer progression model. We have also confirmed the overexpression of two invasion-related biomarkers, psoriasin (PSOR1) and versican (CSPG2), in oral premalignant and malignant archival tissues. Our results clearly indicate that pharmacologic intervention with anti-inflammatory prostaglandin D2–like analogues may help prevent or delay oral epithelial carcinogenesis because of metabolic restoration of a negative feedback regulatory loop through its several cognate receptors or target molecules. Further studies directed toward a multitude of possible protective mechanisms of this lipocalin-type enzyme or its products in oral cancer progression are warranted.
Identification of compounds that inhibit growth of 2-amino-1-methyl-6-phenylimidazo(4,5-b)pyridine–resistant cancer cells
Authors: Bachman Kurtis E. , Sager Jason , Cheong Ian , Catto Marco , Bardelli Alberto , Park Ben Ho , Vogelstein Bert , Carotti Angelo , Kinzler Kenneth W. , Lengauer Christoph
Published: September 2005

The dietary carcinogen 2-amino-1-methyl-6-phenylimidazo(4,5-b)pyridine (PhIP) is a heterocyclic amine and is a common byproduct of cooked meat and fish. Although most cells undergo apoptosis when exposed to this mutagen, subsets develop resistance. Rather than die, these resistant cells persist and accumulate mutations, thereby driving tumorigenesis of exposed organs within the gastrointestinal tract. By applying a high-throughput cell-based screen of 32,000 small molecules, we have identified a family of compounds that specifically inhibit the growth of PhIP-resistant cancer cells. These compounds may prove useful for the treatment or prevention of gastrointestinal tumors arising after exposure to PhIP and related carcinogens.
Reversal of cisplatin resistance with a BH3 mimetic, (−)-gossypol, in head and neck cancer cells: role of wild-type p53 and Bcl-xL
Authors: Bauer Joshua A. , Trask Douglas K. , Kumar Bhavna , Los Gerrit , Castro Jason , Shin-Jung Lee Julia , Chen Jianyong , Wang Shaomeng , Bradford Carol R. , Carey Thomas E.
Published: September 2005

Organ preservation protocols in head and neck squamous cell carcinoma (HNSCC) are limited by tumors that fail to respond. We observed that larynx preservation and response to chemotherapy is significantly associated with p53 overexpression, and that most HNSCC cell lines with mutant p53 are more sensitive to cisplatin than those with wild-type p53. To investigate cisplatin resistance, we studied two HNSCC cell lines, UM-SCC-5 and UM-SCC-10B, and two resistant sublines developed by cultivation in gradually increasing concentrations of cisplatin. The cisplatin-selected cell lines, UM-SCC-5PT and UM-SCC-10BPT, are 8 and 1.5 times more resistant to cisplatin than the respective parental cell lines, respectively. The parental lines overexpress p53 and contain p53 mutations but the cisplatin-resistant cell lines do not, indicating that cells containing mutant p53 were eliminated during selection. Bcl-xL expression increased in the cisplatin-resistant lines relative to the parental lines, whereas Bcl-2 expression was high in the parental lines and decreased in the cisplatin-resistant lines. Thus, cisplatin selected for wild-type p53 and high Bcl-xL expression in these cells. We tested a small-molecule BH3 mimetic, (−)-gossypol, which binds to the BH3 domain of Bcl-2 and Bcl-xL, for activity against the parental and cisplatin-resistant cell lines. At physiologically attainable levels, (−)-gossypol induces apoptosis in 70% to 80% of the cisplatin-resistant cells but only in 25% to 40% of the parental cells. Thus, cisplatin-resistant cells seem to depend on wild-type p53 and Bcl-xL for survival and BH3 mimetic agents, such as (−)-gossypol, may be useful adjuncts to overcome cisplatin resistance in HNSCC.
Inhibition of invasion, angiogenesis, tumor growth, and metastasis by adenovirus-mediated transfer of antisense uPAR and MMP-9 in non–small cell lung cancer cells
Authors: Rao Jasti S. , Gondi Christopher , Chetty Chandramu , Chittivelu Subramanyam , Joseph Pushpa A. , Lakka Sajani S.
Published: September 2005

Lung cancer is currently the leading cause of cancer deaths in the United States. Conventional therapeutic treatments, including surgery, chemotherapy, and radiation therapy, have achieved only limited success. The overexpression of proteases, such as urokinase-type plasminogen activator (uPA), its receptor (uPAR), and matrix metalloproteinases (MMP), is correlated with the progression of lung cancer. In the present study, we used a replication-deficient adenovirus capable of expressing antisense uPAR and antisense MMP-9 transcripts to simultaneously down-regulate uPAR and MMP-9 in H1299 cells. Ad-uPAR-MMP-9 infection of H1299 cells resulted in a dose- and time-dependent decrease of uPAR protein levels and MMP-9 activity as determined by Western blotting and gelatin zymography, respectively. Corresponding immunohistochemical analysis also showed that Ad-uPAR-MMP-9 infection inhibited uPAR and MMP-9 expression. As shown by Boyden chamber assay, Ad-uPAR-MMP-9 infection significantly decreased the invasive capacity of H1299 cells compared with mock and Ad-CMV (empty vector)–infected cells in vitro. Furthermore, Ad-uPAR-MMP-9 infection inhibited capillary-like structure formation in H1299 cells cocultured with endothelial cells in a dose-dependent manner compared with mock- and Ad-CMV-infected cells. Ad-uPAR-MMP-9 injection caused the regression of s.c. induced tumors after s.c. injection with H1299 lung cancer cells and inhibited lung metastasis in the metastatic model with A549 cells. These data suggest that Ad-uPAR-MMP-9 shows its antitumor activity against both established and early phases of lung cancer metastases by causing the destruction of the tumor vasculature. In summary, adenovirus-mediated inhibition of uPA-uPAR interaction and MMP-9 on the cell surface may be a promising anti-invasion and antimetastatic strategy for cancer gene therapy.
T315I-mutated Bcr-Abl in chronic myeloid leukemia and imatinib: insights from a computational study
Authors: Pricl Sabrina , Fermeglia Maurizio , Ferrone Marco , Tamborini Elena
Published: September 2005

The early stage of chronic myeloid leukemia is triggered by the tyrosine kinase Bcr-Abl. Imatinib mesylate, a selective inhibitor of Bcr-Abl, has been successful in chronic myeloid leukemia clinical trials, but short-lived remissions are usually observed in blast crisis patients. Sequencing of the BCR-ABL gene in relapsed patients revealed a set of mutants that mediate drug resistance. Previously reported work postulated that the missense T315I mutation both alters the three-dimensional structure of the protein binding site, thus decreasing the protein sensitivity for the drug, and does not feature a fundamental hydrogen bond that is critical for binding with imatinib. These speculations, however, were not supported by investigations at the molecular modeling level. Here, we present the results obtained from the application of molecular dynamics simulations to the study of the interactions between T315I Bcr-Abl and imatinib. For the first time, we show that, with respect to the wild-type system, the absence of the supposedly critical H-bond is not the only cause for the failure of receptor inhibition by imatinib, but also a plethora of other protein/drug interactions are drastically and unfavorably changed in the mutant protein.
Modulation of cell cycle and gene expression in pancreatic tumor cell lines by methionine deprivation (methionine stress): implications to the therapy of pancreatic adenocarcinoma
Authors: Kokkinakis Demetrius M. , Liu XiaoYan , Neuner Russell D.
Published: September 2005

The effect of methionine deprivation (methionine stress) on the proliferation, survival, resistance to chemotherapy, and regulation of gene and protein expression in pancreatic tumor lines is examined. Methionine stress prevents successful mitosis and promotes cell cycle arrest and accumulation of cells with multiple micronuclei with decondensed chromatin. Inhibition of mitosis correlates with CDK1 down-regulation and/or inhibition of its function by Tyr15 phosphorylation or Thr161 dephosphorylation. Inhibition of cell cycle progression correlates with loss of hyperphosphorylated Rb and up-regulation of p21 via p53 and/or transforming growth factor-β (TGF-β) activation depending on p53 status. Although methionine stress–induced toxicity is not solely dependent on p53, the gain in p21 and loss in CDK1 transcription are more enhanced in wild-type p53 tumors. Up-regulation of SMAD7, a TGF-β signaling inhibitor, suggests that SMAD7 does not restrict the TGF-β-mediated induction of p21, although it may prevent up-regulation of p27. cDNA oligoarray analysis indicated a pleiotropic response to methionine stress. Cell cycle and mitotic arrest is in agreement with up-regulation of NF2, ETS2, CLU, GADD45α, GADD45β, and GADD45γ and down-regulation of AURKB, TOP2A, CCNA, CCNB, PRC1, BUB1, NuSAP, IFI16, and BRCA1. Down-regulation of AREG, AGTR1, M-CSF, and EGF, IGF, and VEGF receptors and up-regulation of GNA11 and IGFBP4 signify loss of growth factor support. PIN1, FEN1, and cABL up-regulation and LMNB1, AREG, RhoB, CCNG, TYMS, F3, and MGMT down-regulation suggest that methionine stress sensitizes the tumor cells to DNA-alkylating drugs, 5-fluorouracil, and radiation. Increased sensitivity of pancreatic tumor cell lines to temozolomide is shown under methionine stress conditions and is attributed in part to diminished O6-methylguanine-DNA methyltransferase and possibly to inhibition of the cell cycle progression.
BRAF status and mitogen-activated protein/extracellular signal-regulated kinase kinase 1/2 activity indicate sensitivity of melanoma cells to anthrax lethal toxin
Authors: Abi-Habib Ralph J. , Urieto Jeffrey O. , Liu Shihui , Leppla Stephen H. , Duesbery Nicholas S. , Frankel Arthur E.
Published: September 2005

Anthrax lethal toxin, composed of protective antigen and lethal factor, was tested for cytotoxicity to human melanoma cell lines and normal human cells. Eleven of 18 melanoma cell lines were sensitive to anthrax lethal toxin (IC50 < 400 pmol/L) and 10 of these 11 sensitive cell lines carried the V599E BRAF mutation. Most normal cell types (10 of 15) were not sensitive to anthrax lethal toxin and only 5 of 15 normal human cell types were sensitive to anthrax lethal toxin (IC50 < 400 pmol/L). These cells included monocytes and a subset of endothelial cells. In both melanoma cell lines and normal cells, anthrax toxin receptor expression levels did not correlate with anthrax lethal toxin cytotoxicity. Furthermore, an anthrax toxin receptor–deficient cell line (PR230) did not show any enhanced sensitivity to anthrax lethal toxin when transfected with anthrax toxin receptor. Anthrax lethal toxin toxicity correlated with elevated phosphorylation levels of mitogen-activated protein/extracellular signal-regulated kinase kinase (MEK) 1/2 in both melanoma cell lines and normal cells. Anthrax lethal toxin–sensitive melanoma cell lines and normal cells had higher phospho-MEK1/2 levels than anthrax lethal toxin–resistant melanoma cell lines and normal tissue types. U0126, a specific MEK1/2 inhibitor, was not toxic to anthrax lethal toxin–resistant melanoma cell lines but was toxic to 8 of 11 anthrax lethal toxin–sensitive cell lines. These results show that anthrax lethal toxin toxicity correlates with elevated levels of active MEK1/2 pathway but not with anthrax toxin receptor expression levels in both normal and malignant tissues. Anthrax lethal toxin may be a useful therapeutic for melanoma patients, especially those carrying the V599E BRAF mutation with constitutive activation of the mitogen-activated protein kinase pathway.
The phosphatidylinositol-3-kinase inhibitor PX-866 overcomes resistance to the epidermal growth factor receptor inhibitor gefitinib in A-549 human non–small cell lung cancer xenografts
Authors: Ihle Nathan T. , Paine-Murrieta Gillian , Berggren Margareta I. , Baker Amanda , Tate Wendy R. , Wipf Peter , Abraham Robert T. , Kirkpatrick D. Lynn , Powis Garth
Published: September 2005

Epidermal growth factor receptor (EGFR) inhibitors such as gefitinib show antitumor activity in a subset of non–small cell lung cancer (NSCLC) patients having mutated EGFR. Recent work shows that phosphatidylinositol-3-kinase (PI3-K) is coupled to the EGFR only in NSCLC cell lines expressing ErbB-3 and that EGFR inhibitors do not inhibit PI3-K signaling in these cells. The central role PI3-K plays in cell survival suggests that a PI3-K inhibitor offers a strategy to increase the antitumor activity of EGFR inhibitors in resistant NSCL tumors that do not express ErbB-3. We show that PX-866, a PI3-K inhibitor with selectivity for p110α, potentiates the antitumor activity of gefitinib against even large A-549 NSCL xenografts giving complete tumor growth control in the early stages of treatment. A-549 xenograft phospho-Akt was inhibited by PX-866 but not by gefitinib. A major toxicity of PX-866 administration was hyperglycemia with decreased glucose tolerance, which was reversed upon cessation of treatment. The decreased glucose tolerance caused by PX-866 was insensitive to the AMP-activated protein kinase inhibitor metformin but reversed by insulin and by the peroxisome proliferator-activated receptor-γ activator pioglitazone. Prolonged PX-866 administration also caused increased neutrophil counts. Thus, PX-866, by inhibiting PI3-K signaling, may have clinical use in increasing the response to EGFR inhibitors such as gefitinib in patients with NSCLC and possibly in other cancers who do not respond to EGFR inhibition.
Vascular endothelial growth factor selectively targets boronated dendrimers to tumor vasculature
Authors: Backer Marina V. , Gaynutdinov Timur I. , Patel Vimal , Bandyopadhyaya Achintya K. , Thirumamagal B.T.S. , Tjarks Werner , Barth Rolf F. , Claffey Kevin , Backer Joseph M.
Published: September 2005

Tumor neovasculature is a potential but, until very recently, unexplored target for boron neutron capture therapy (BNCT) of cancer. In the present report, we describe the construction of a vascular endothelial growth factor (VEGF)–containing bioconjugate that potentially could be used to target up-regulated VEGF receptors (VEGFR), which are overexpressed on tumor neovasculature. A fifth-generation polyamidoamine dendrimer containing 128 reactive amino groups was reacted with 105 to 110 decaborate molecules to produce a macromolecule with 1,050 to 1,100 boron atoms per dendrimer. This was conjugated to thiol groups of VEGF at a 4:1 molar ratio using the heterobifunctional reagent sulfo-LC-SPDP. In addition, the boronated dendrimer was tagged with a near-IR Cy5 dye to allow for near-IR fluorescent imaging of the bioconjugate in vitro and in vivo. As would be predicted, the resulting VEGF-BD/Cy5 bioconjugate was not cytotoxic to HEK293 cells engineered to express 2.5 × 106 VEGFR-2 per cell. Furthermore, it showed binding and activation of VEGFR-2 comparable with that of native VEGF. Internalization of VEGF-BD/Cy5 by PAE cells expressing 2.5 × 105 VEGFR-2 per cell was inhibited by excess VEGF, indicating a VEGFR-2-mediated mechanism of uptake. Near-IR fluorescent imaging of 4T1 mouse breast carcinoma revealed selective accumulation of VEGF-BD/Cy5, but not BD/Cy5, particularly at the tumor periphery where angiogenesis was most active. Accumulation of VEGF-BD/Cy5 in 4T1 breast carcinoma was diminished in mice pretreated with a toxin-VEGF fusion protein that selectively killed VEGFR-2-overexpressing endothelial cells. Our data lay the groundwork for future studies using the VEGF-BD/Cy5 bioconjugate as a targeting agent for BNCT of tumor neovasculature.
The rice bran constituent tricin potently inhibits cyclooxygenase enzymes and interferes with intestinal carcinogenesis in ApcMin mice
Authors: Cai Hong , Al-Fayez Mohammad , Tunstall Richard G. , Platton Sharon , Greaves Peter , Steward William P. , Gescher Andreas J.
Published: September 2005

While brown rice is a staple dietary constituent in Asia, rice consumed in the Western world is generally white, obtained from brown rice by removal of the bran. Rice bran contains the flavone tricin, which has been shown to inhibit colon cancer cell growth. We tested the hypothesis that tricin interferes with adenoma formation in the ApcMin mouse. Mice received tricin (0.2%) in their American Institute of Nutrition 93G diet throughout their postweaning life span (4–18 weeks). Consumption of tricin reduced numbers of intestinal adenomas by 33% (P < 0.05) compared with mice on control diet. We explored whether tricin may exert its effect via inhibition of cyclooxygenase (COX) enzymes. Its effect on COX activity was assessed in purified enzyme preparations in vitro and its ability to reduce prostaglandin E2 (PGE2) levels in human colon–derived human colon epithelial cell (HCEC) and HCA-7 cells in vitro and in ApcMin mice in vivo. Tricin inhibited activity of purified COX-1 and COX-2 enzyme preparations with IC50 values of ∼1 μmol/L. At 5 μmol/L, it reduced PGE2 production in HCEC or HCA-7 cells by 36% (P < 0.01) and 35% (P < 0.05), respectively. COX-2 expression was reduced by tricin weakly in HCEC and unaffected in HCA-7 cells. PGE2 levels in the small intestinal mucosa and blood of ApcMin mice that had received tricin were reduced by 34% (P < 0.01) and 40% (P < 0.05), respectively, compared with control mice. The results suggest that tricin should be further evaluated as a putative colorectal cancer chemopreventive agent.
Inhibition of human tumor cell growth in vivo by an orally bioavailable inhibitor of CDC25 phosphatases
Authors: Brezak Marie-Christine , Quaranta Muriel , Contour-Galcera Marie-Odile , Lavergne Olivier , Mondesert Odile , Auvray Pierrïck , Kasprzyk Philip G. , Prevost Gregoire P. , Ducommun Bernard
Published: September 2005

Cell cycle regulators, such as the CDC25 phosphatases, are potential targets for the development of new anticancer drugs. Here we report the identification and the characterization of BN82685, a quinone-based CDC25 inhibitor that is active in vitro and in vivo. BN82685 inhibits recombinant CDC25A, B, and C phosphatases in vitro. It inhibits the growth of human tumor cell lines with an IC50 in the submicromolar range, independently of their resistance to chemotherapeutic agents. This inhibitory effect is irreversible on both the purified CDC25 enzyme in vitro and on tumor cell proliferation. The specificity of BN82685 towards the CDC25 phosphatases is shown by an increase in cyclin-dependent kinase 1 tyrosine 15 phosphorylation, by the reversion of the mitosis-inducing effect of CDC25B overexpression in HeLa cells, and by the lack of a growth inhibitory effect in an assay based on the use of a CDC25-independent fission yeast model. Finally, when administered p.o., BN82685 is shown to inhibit the growth of the human pancreatic tumor Mia PaCa-2 xenografted in athymic nude mice. BN82685 is therefore a promising new compound targeting CDC25, which confirms the interest of the inhibition of these enzymes as an anticancer therapeutic strategy.
The Candidate Tumor Suppressor Gene ZAC Is Involved in Keratinocyte Differentiation and Its Expression Is Lost in Basal Cell Carcinomas
Authors: Basyuk Eugenia , Coulon Vincent , Le Digarcher Anne , Coisy-Quivy Marjorie , Moles Jean-Pierre , Gandarillas Alberto , Journot Laurent
Published: September 2005

ZAC is a zinc finger transcription factor that induces apoptosis and cell cycle arrest in various cell lines. The corresponding gene is maternally imprinted and localized on chromosome 6q24-q25, a region harboring an unidentified tumor suppressor gene for a variety of solid neoplasms. ZAC expression is lost or down-regulated in some breast, ovary, and pituitary tumors and in an in vitro model of ovary epithelial cell transformation. In the present study, we examined ZAC expression in normal skin and found a high expression level in basal keratinocytes and a lower, more heterogeneous, expression in the first suprabasal differentiating layers of epidermis. In vitro, ZAC was up-regulated following induction of keratinocyte differentiation. Conversely, ZAC expression triggered keratinocyte differentiation as indicated by induction of involucrin expression. Interestingly, we found a dramatic loss of ZAC expression in basal cell carcinoma, a neoplasm characterized by a relatively undifferentiated morphology. In contrast, ZAC expression was maintained in squamous cell carcinomas that retain the squamous differentiated phenotype. Altogether, these data suggest a role for ZAC at an early stage of keratinocyte differentiation and further support its role in carcinogenesis.
Development of New EBV-Based Vectors for Stable Expression of Small Interfering RNA to Mimick Human Syndromes: Application to NER Gene Silencing
Authors: Biard Denis S.F. , Despras Emmanuelle , Sarasin Alain , Angulo Jaime F.
Published: September 2005

We developed and characterized replicative small interfering RNA (siRNA) vectors for efficient, specific, and long-term gene silencing in human cells. We created stable XPAKD and XPCKD (knockdown) syngeneic cell lines to mimic human cancer-prone syndromes. We also silenced HSAKIN17. Several clones displaying undetectable protein levels of XPA, XPC, or HSAkin17 were grown for more than 300 days. This stability of gene silencing over several months of culture allows us to assess the specific involvement of these proteins in UVC sensitivity in syngeneic cells. Unlike XPA, HSAKIN17, and XPC gene silencing dramatically impeded HeLa cell growth for several weeks after transfection. As expected, XPAKD and XPCKD HeLa cells were highly UVC sensitive. They presented an impaired unscheduled DNA synthesis after UVC irradiation. Interestingly, XPCKD HeLa clones were more sensitive to UVC than their XPAKD or KIN17KD counterparts. Hygromycin B withdrawal led to the total disappearance of EBV vectors and the resumption of normal XPA or XPC protein levels. Whereas reverted XPAKD cells recovered a normal UVC sensitivity, XPCKD cells remained highly sensitive, suggestive of irreversible damage following long-term XPC silencing. Our results show that in HeLa cells, HSAkin17 participates indirectly in early events following UVC irradiation, and XPC deficiency strongly affects cell physiology and contributes to UVC sensitivity to a greater extent than does XPA. EBV-based siRNA vectors improve the interest of siRNA by permitting long-term gene silencing without the safety concerns inherent in viral-based siRNA vehicles.
Genomic Profiling Maps Loss of Heterozygosity and Defines the Timing and Stage Dependence of Epigenetic and Genetic Events in Wilms' Tumors
Authors: Yuan Eric , Li Chi-Ming , Yamashiro Darrell J. , Kandel Jessica , Thaker Harshwardhan , Murty Vundavalli V. , Tycko Benjamin
Published: September 2005

To understand genetic and epigenetic pathways in Wilms' tumors, we carried out a genome scan for loss of heterozygosity (LOH) using Affymetrix 10K single nucleotide polymorphism (SNP) chips and supplemented the data with karyotype information. To score loss of imprinting (LOI) of the IGF2 gene, we assessed DNA methylation of the H19 5′ differentially methylated region (DMR). Few chromosomal regions other than band 11p13 (WT1) were lost in Wilms' tumors from Denys-Drash and Wilms' tumor-aniridia syndromes, whereas sporadic Wilms' tumors showed LOH of several regions, most frequently 11p15 but also 1p, 4q, 7p, 11q, 14q, 16q, and 17p. LOI was common in the sporadic Wilms' tumors but absent in the syndromic cases. The SNP chips identified novel centers of LOH in the sporadic tumors, including a 2.4-Mb minimal region on chromosome 4q24-q25. Losses of chromosomes 1p, 14q, 16q, and 17p were more common in tumors presenting at an advanced stage; 11p15 LOH was seen at all stages, whereas LOI was associated with early-stage presentation. Wilms' tumors with LOI often completely lacked LOH in the genome-wide analysis, and in some tumors with concomitant 16q LOH and LOI, the loss of chromosome 16q was mosaic, whereas the H19 DMR methylation was complete. These findings confirm molecular differences between sporadic and syndromic Wilms' tumors, define regions of recurrent LOH, and indicate that gain of methylation at the H19 DMR is an early event in Wilms' tumorigenesis that is independent of chromosomal losses. The data further suggest a biological difference between sporadic Wilms' tumors with and without LOI.
Radiofrequency for the Treatment of Liver Tumours
Authors: Ruers T.J.M. , de Jong K.P. , IJzermans J.N.M.
Published: September 2005

Resection should still be considered the gold standard for many liver tumours. There is, however, growing interest in the use of radiofrequency (RFA) for the treatment of liver tumours. By RFA, tumour tissue can be destructed selectively without significant damage to vascular structures in the remaining liver. The technique is safe and feasible for many types of liver tumours. RFA appears to be beneficial in the treatment of hepatocellular carcinoma. In the Netherlands, single-centre reports suggest that RFA may be used successfully to control hepatocellular carcinoma in those patients awaiting liver transplantation. RFA is being increasingly used for colorectal liver metastases as an adjunct to surgical resection in case of unresectable lesions. However, to date, there are still no data showing that such an approach is beneficial. For this reason, in the Netherlands, RFA for unresectable colorectal liver metastases is mainly used within a clinical trial. Within the multimodality treatment of neuroendocrine metastases, RFA may deserve a place for either intention to cure (rare) or debulking with the aim of reduction of symptoms or prolongation of life.
Translational Research in Prognostic Profiling in Colorectal Cancer
Authors: de Noo M.E. , Liefers G.J. , Tollenaar R.A.E.M.
Published: September 2005

There is a widening gap between basic research and clinical practice, particularly for colorectal cancer. In recent years, many have expressed concerns regarding the disconnection between the promises of basic science and the delivery of better individual health. In this paper we describe some of our research in serum proteomics, microarrays and minimal residual disease dedicated to this field and discuss some of the roadblocks ahead in translational research. We conclude that translational medicine should be a collective effort for the medical community as a whole with adequate financial support and sound, measurable outcome. Since extensive validation of the above mentioned research fields is necessary, adequate funding is required. This may require some adjustments in the current funding policy because it involves non-innovative studies. Furthermore, the pool of researchers/clinicians capable of performing translational research must be increased. Additionally, there should be an enhanced participation of patients in clinical trials and an optimization of the efficiency of these trials using validated surrogate markers. Only when these conditions are fulfilled will the ‘post-genomic` era of biomedical research have unprecedented opportunities to innovate and improve therapy for cancer.
Recent Insights into the Cell Biology of Bladder Smooth Muscle
Authors: Adam Rosalyn M.
Published: September 2005

Much of current biomedical research is focused on the development of ‘targeted therapies’ based on detailed knowledge about the signals that mediate aberrant cellular behavior in a given disease. Although this concept has been used most widely in cancer treatment, the same strategy applies to nonmalignant conditions such as pathologic tissue expansion in the genitourinary tract. A rigorous understanding of the key molecular events and pathways that underlie normal and pathologic activity of the bladder would allow us to identify potential targets for rational drug design. In this review, I will summarize our current understanding of cell signaling in bladder smooth muscle and highlight potential targets for drug-based treatment of tissue remodeling in the lower urinary tract.