bluuebunny/crossref_metadata_2025_split huggingface dataset:
Gene therapy withE2F-1up-regulates the protein kinase PKR and inhibits growth of leiomyosarcomain vivo
Authors: Vorburger Stephan A. , Hetrakul Nophadol , Xia Weiya , Wilson-Heiner Marcia , Mirza Nadeem , Pollock Raphael E. , Feig Barry , Swisher Stephen G. , Hunt Kelly K. 
Published: March 2006

AbstractOverexpression of the transcription factor E2F-1 induces apoptosis in a variety of carcinoma cells and inactivates murine double minute protein 2, a factor associated with poor prognosis in soft tissue sarcomas. We have shown previously that the double-stranded RNA-activated protein kinase PKR plays an important role in mediating this apoptotic response in carcinoma cells to E2F-1. We sought to evaluate the potential of E2F-1 gene therapy in soft tissue sarcomas and to study the involvement of PKR in the response to E2F-1 overexpression in mesenchymal cells. A replication-deficient adenovirus carrying the E2F-1 gene (Ad5E2F) was used to induce E2F-1 overexpression in the p53 mutated leiomyosarcoma cell line, SKLMS-1. Western blot analysis confirmed E2F-1 overexpression and up-regulation of the antiapoptotic factor Bcl-2 48 hours following infection with Ad5E2F. Apoptosis in Ad5E2F-treated cells was confirmed by fluorescence-activated cell sorting analysis and by poly(ADP-ribose) polymerase cleavage and DNA fragmentation assays. Vector-dependent up-regulation of PKR correlated with the amount of Ad5E2F-induced apoptosis. In vivo treatment of SKLMS-1 tumor-bearing BALB/c mice with intratumoral injections of Ad5E2F at a dose of 2 × 1010 viral particles resulted in significant inhibition in tumor growth compared with control-treated animals (P &lt; 0.016). Complete disappearance of all tumors was seen in two of seven mice in the Ad5E2F-treated animals. Immunohistochemical analysis of tumor specimens showed overexpression of E2F-1 and up-regulation of PKR in Ad5E2F-treated tumors. These findings show that adenovirus-mediated overexpression of E2F-1 results in up-regulation of PKR and significant growth suppression of leiomyosarcomas in vivo. Taken together, these data suggest that E2F-1 gene therapy and PKR modulation might be a promising treatment strategy for these tumors that are highly resistant to conventional therapies.
Imatinib binding and cKIT inhibition is abrogated by the cKIT kinase domain I missense mutation Val654Ala
Authors: McLean Sean R. , Gana-Weisz Mali , Hartzoulakis Basil , Frow Richard , Whelan Jeremy , Selwood David , Boshoff Chris 
Published: March 2006

Abstract
Several activating mutations in the cKIT receptor tyrosine kinase are associated with the development and progression of gastrointestinal stromal tumors (GIST). Treatment of GIST with the tyrosine kinase inhibitor imatinib (Gleevec, STI571; Novartis, Basel, Switzerland) increases patient survival. However, many patients develop resistance to imatinib following initial responses. We sequenced cKIT exons from two patients with GIST after the development of imatinib resistance, revealing a point mutation in kinase domain I (exon 13), Val654Ala, which has been associated previously with relapse and resistance. Molecular modeling of cKIT-imatinib complexes shows that this residue is located in the drug-binding site and that the Val654Ala mutation disrupts drug binding by removing hydrophobic contacts with the central diaminophenyl ring of imatinib. Loss of these contacts results in a destabilizing effect on two key hydrogen bonds between imatinib and Asp310 and Thr670 of cKIT. Calculations based on published crystallography data show an estimated destabilization energy of 2.25 kcal/mol in the Val654Ala cKIT compared with wild type. When present on the same cKIT allele as an oncogenic mutation, the Val654Ala mutation abolishes imatinib-mediated inhibition of cKIT phosphoactivation in vitro. These results highlight some of the structural and functional consequences of the Val654Ala mutation in relapsing imatinib-resistant GIST and emphasize the importance of tumor genetics in drug development and patient-specific cancer treatment regimens. [Mol Cancer Ther 2005;4(12):2005–15]
Novel angiogenic inhibitor DN-9693 that inhibits post-transcriptional induction of connective tissue growth factor (CTGF/CCN2) by vascular endothelial growth factor in human endothelial cells
Authors: Kondo Seiji , Tanaka Noriko , Kubota Satoshi , Mukudai Yoshiki , Yosimichi Gen , Sugahara Toshio , Takigawa Masaharu 
Published: March 2006

Abstract
Connective tissue growth factor (CTGF/CCN2) is a potent angiogenic factor. In this report, we describe for the first time that vascular endothelial growth factor (VEGF)–mediated induction of the ctgf/ccn2 gene was a post-transcriptional event that was inhibited by a novel angiogenic inhibitor, DN-9693, in human umbilical vein endothelial cells. Steady-state mRNA levels of ctgf/ccn2 were remarkably increased by VEGF in a concentration-dependent manner, whereas the activity of the ctgf/ccn2 promoter was not responsive to VEGF as confirmed by a reporter gene assay and quantitative real-time PCR analysis. By employing a RNA degradation assay, we eventually found that the observed increase in the ctgf/ccn2 mRNA level was due to an increased stability of the mRNA induced by VEGF. DN-9693 at a dose of 0.1 to 2 ng/mL did not affect basal levels of ctgf/ccn2 mRNA; however, enhancement of ctgf/ccn2 mRNA expression by VEGF was specifically inhibited by DN-9693. Of importance, the inhibitory effects could be also ascribed to post-transcriptional regulation, because the VEGF-mediated increase in stability of ctgf/ccn2 mRNA was suppressed by DN-9693. Furthermore, we investigated the effects of DN-9693 on VEGF-induced activation of three subgroups of mitogen-activated protein kinase pathways and found that DN-9693 blocked the activation of these pathways by VEGF. These results suggest that VEGF increases ctgf/ccn2 mRNA stability through mitogen-activated protein kinase–mediated intracellular signaling cascade(s), which can be inhibited posttranscriptionally by a novel angiogenic inhibitor, DN-9693, in human umbilical vein endothelial cells. [Mol Cancer Ther 2006;5(1):129–37]
Homozygous deletions of methylthioadenosine phosphorylase in human biliary tract cancers
Authors: Karikari Collins A. , Mullendore Michael , Eshleman James R. , Argani Pedram , Leoni Lorenzo M. , Chattopadhyay Shrikanta , Hidalgo Manuel , Maitra Anirban 
Published: March 2006

Abstract
The p16INK4A/CDKN2A gene on chromosome 9p21 is a site of frequent allelic loss in human cancers, and in a subset of cases, homozygous deletions at this locus encompass the telomeric methylthioadenosine phosphorylase (MTAP) gene. The MTAP gene product is the principal enzyme involved in purine synthesis via the salvage pathway, such that MTAP-negative cancers are solely dependent on de novo purine synthesis mechanisms. Inhibitors of the de novo pathway can then be used to selectively blockade purine synthesis in cancer cells while causing minimal collateral damage to normal cells. In this study, we determine that 10 of 28 (35%) biliary tract cancers show complete lack of Mtap protein expression. In vitro analysis using a selective inhibitor of the de novo purine synthesis pathway, l-alanosine, shows robust growth inhibition in MTAP-negative biliary cancer cell lines CAK-1 and GBD-1 accompanied by striking depletion of intracellular ATP and failure to rescue this depletion via addition of exogenous methylthioadenosine, the principal substrate of the MTAP gene product; in contrast, no significant effects were observed in MTAP-expressing HuCCT1 and SNU308 cell lines. Colony formation studies confirmed that l-alanosine reduced both number and size of CAK-1 colonies in soft agar assays. Knockdown of Mtap protein by RNA interference in l-alanosine-resistant HuCCT1 cells conferred sensitivity to this agent, confirming that intracellular Mtap protein levels determine response to l-alanosine. Inhibitors of de novo purine synthesis can be a potential mechanism-based strategy for treatment of biliary tract cancers, one third of which show complete loss of MTAP function. [Mol Cancer Ther 2005;4(12):1860–6]
Enhancement of DNA topoisomerase I inhibitor–induced apoptosis by ursodeoxycholic acid
Authors: Ikegami Tadashi , Matsuzaki Yasushi , Al Rashid Maryam , Ceryak Susan , Zhang Yining , Bouscarel Bernard 
Published: March 2006

Abstract
Certain hydrophobic bile acids, including deoxycholic acid and chenodeoxycholic acid, exert toxic effects not only in the liver but also in the intestine. Moreover, ursodeoxycholic acid (UDCA), which has protective actions against apoptosis in the liver, may have both protective and toxic effects in the intestine. The goal of the present study was to clarify the mechanisms responsible for the toxic effect of UDCA in intestinal HT-29 cells. Here, we show that UDCA potentiated both phosphatidylserine externalization and internucleosomal DNA fragmentation induced by SN-38, the most potent metabolite of the DNA topoisomerase I inhibitor, CPT-11. Furthermore, the loss of mitochondrial membrane potential as well as mitochondrial membrane permeability transition induced by SN-38 was enhanced in the presence of UDCA, resulting in an increased lethality determined by colony-forming assay. This UDCA-induced increased apoptosis was not due to alteration of either intracellular accumulation of SN-38 or cell cycle arrest by SN-38. The increased apoptosis was best observed when UDCA was present after SN-38 stimulation and was independent of caspase-8 but dependent on caspase-9 and caspase-3 activation. Furthermore, UDCA enhanced SN-38-induced c-Jun NH2-terminal kinase activation. In conclusion, UDCA increases the apoptotic effects while decreasing the necrotic effects of SN-38 when added after the topoisomerase I inhibitor, showing potential clinical relevance as far as targeted cell death and improved wound healing are concerned. However, the use of this bile acid as an enhancer in antitumor chemotherapy should be further evaluated clinically. [Mol Cancer Ther 2006;5(1):68–79]
Isothiazolones as inhibitors of PCAF and p300 histone acetyltransferase activity
Authors: Stimson Lindsay , Rowlands Martin G. , Newbatt Yvette M. , Smith Nicola F. , Raynaud Florence I. , Rogers Paul , Bavetsias Vassilios , Gorsuch Stephen , Jarman Michael , Bannister Andrew , Kouzarides Tony , McDonald Edward , Workman Paul , Aherne G. Wynne 
Published: March 2006

Abstract
Histone acetylation plays an important role in regulating the chromatin structure and is tightly regulated by two classes of enzyme, histone acetyltransferases (HAT) and histone deacetylases (HDAC). Deregulated HAT and HDAC activity plays a role in the development of a range of cancers. Consequently, inhibitors of these enzymes have potential as anticancer agents. Several HDAC inhibitors have been described; however, few inhibitors of HATs have been disclosed. Following a FlashPlate high-throughput screen, we identified a series of isothiazolone-based HAT inhibitors. Thirty-five N-substituted analogues inhibited both p300/cyclic AMP–responsive element binding protein–binding protein–associated factor (PCAF) and p300 (1 to &gt;50 μmol/L, respectively) and the growth of a panel of human tumor cell lines (50% growth inhibition, 0.8 to &gt;50 μmol/L). CCT077791 and CCT077792 decreased cellular acetylation in a time-dependent manner (2–48 hours of exposure) and a concentration-dependent manner (one to five times, 72 hours, 50% growth inhibition) in HCT116 and HT29 human colon tumor cell lines. CCT077791 reduced total acetylation of histones H3 and H4, levels of specific acetylated lysine marks, and acetylation of α-tubulin. Four and 24 hours of exposure to the compounds produced the same extent of growth inhibition as 72 hours of continuous exposure, suggesting that growth arrest was an early event. Chemical reactivity of these compounds, as measured by covalent protein binding and loss of HAT inhibition in the presence of DTT, indicated that reaction with thiol groups might be important in their mechanism of action. As one of the first series of small-molecule inhibitors of HAT activity, further analogue synthesis is being pursued to examine the potential scope for reducing chemical reactivity while maintaining HAT inhibition.
A role for PKCζ in potentiation of the topoisomerase II activity and etoposide cytotoxicity by wortmannin
Authors: Reis Caroline , Giocanti Nicole , Hennequin Christophe , Mégnin-Chanet Frédérique , Fernet Marie , Filomenko Rodolphe , Bettaieb Ali , Solary Eric , Favaudon Vincent 
Published: March 2006

AbstractEnhanced cytotoxicity of etoposide by wortmannin, an inhibitor of enzymes holding a phosphatidylinositol 3-kinase domain, was investigated in eight cell lines proficient or deficient for DNA double-strand break repair. Wortmannin stimulated the decatenating activity of topoisomerase II, promoted etoposide-induced accumulation of DNA double-strand breaks, shifted the specificity for cell killing by etoposide from the S to G1 phase of the cell cycle, and potentiated the cytotoxicity of etoposide through two mechanisms. (a) Sensitization to high, micromolar amounts of etoposide required integrity of the nonhomologous end-joining repair pathway. (b) Wortmannin dramatically increased the susceptibility to low, submicromolar amounts of etoposide in a large fraction of the cell population irrespective of the status of ATM, Ku86, and DNA-PKCS. It is shown that this process correlates depression of phosphatidylinositol 3-kinase–dependent phosphorylation of the atypical, ζ isoform of protein kinase C (PKCζ). Stable expression of a dominant-negative, kinase-dead mutant of PKCζ in a tumor cell line reproduced the hypersensitivity pattern induced by wortmannin. The results are consistent with up-regulation of the topoisomerase II activity in relation to inactivation of PKCζ and indicate that PKCζ may be a useful target to improve the efficiency of topoisomerase II poisons at low concentration.
Transcriptional targeting modalities in breast cancer gene therapy using adenovirus vectors controlled by α-lactalbumin promoter
Authors: Li Xiong , Zhang Jie , Gao Huanling , Vieth Edyta , Bae Kyung-Hee , Zhang Yan-Ping , Lee Sang-Jin , Raikwar Sudhanshu , Gardner Thomas A. , Hutchins Gary D. , VanderPutten Dale , Kao Chinghai , Jeng Meei-Huey 
Published: March 2006

Abstract
The breast-specific antigen α-lactalbumin is expressed in &gt;60% of breast cancer tissues. To evaluate the effect of gene therapy for breast cancer by controlling adenovirus replication with human α-lactalbumin promoter, we investigated the activity of a 762-bp human α-lactalbumin promoter. α-Lactalbumin promoter showed significantly higher activity in MDA-MB-435S and T47D breast cancer cells than in normal breast cell lines or other tumor cell lines. We then developed two novel breast cancer–restricted replicative adenoviruses, AdALAE1a and AdE1aALAE1b. In AdALAE1a, expression of adenoviral E1a gene is under the control of α-lactalbumin promoter, and in AdE1aALAE1b, expression of both E1a and E1b genes is under the control of a single α-lactalbumin promoter. Both breast cancer–restricted replicative adenoviruses showed viral replication efficiency and tumor cell-killing capability similar to wild-type adenovirus in MDA-MB-435S and T47D cells. The replication efficiency and tumor cell-killing capability of both viruses were attenuated significantly in cells that did not support α-lactalbumin promoter. AdE1aALAE1b showed better breast cancer–restricted replication than AdALAE1a, suggesting that a transcriptional targeting modality with α-lactalbumin promoter controlling both E1a and E1b gene expression is superior to α-lactalbumin promoter controlling only E1a gene expression. Importantly, we found that AdE1aALAE1b could be used to target hormone-independent breast tumors in vivo by inhibiting the growth of MDA-MB-435S s.c. tumors. These data showed that α-lactalbumin promoter could regulate the replication of adenovirus to target hormone-independent breast cancers, suggesting that α-lactalbumin promoter can be used to develop a novel therapeutic modality for hormone-independent breast cancer. [Mol Cancer Ther 2005;4(12):1850–9]
Angiogenesis and tumor growth inhibition by a matrix metalloproteinase inhibitor targeting radiation-induced invasion
Authors: Kaliski Alexandre , Maggiorella Laurence , Cengel Keith A. , Mathe Denis , Rouffiac Valerie , Opolon Paule , Lassau Nathalie , Bourhis Jean , Deutsch Eric 
Published: March 2006

AbstractIn this study, we have evaluated the interactions between ionizing radiation and a matrix metalloproteinase (MMP) inhibitor. Using Matrigel invasion assays, we show that ionizing radiation induced a dose-dependent increase in the invasive phenotype of cultured B16 melanoma cells and that conditioned medium from these irradiated B16 cells promoted endothelial cell [human microvascular endothelial cells (HMEC)] invasiveness. To determine whether the radiation-induced changes in invasive phenotype could be due to changes in MMP activation, we have tested the ability of the MMP inhibitor Metastat to modulate the ionizing radiation–induced invasive phenotype using both an in vitro melanoma model and a mouse s.c. tumor model. In these studies, Metastat inhibited the ionizing radiation–induced invasive phenotype in cultured B16 cells and similarly inhibited the increase in HMEC invasion induced by conditioned medium from irradiated B16 cells. Conversely, ionizing radiation increased B16 MMP-2 activity and the conditioned medium from irradiated B16 induced HMEC MMP-2 activity. To further investigate the interaction between ionizing radiation and MMP activation, we then studied the effects of ionizing radiation on downstream effectors of the MMP system. We found that ionizing radiation induced vascular endothelial growth factor (VEGF) secretion by B16 melanoma cells and that this secretion was inhibited by Metastat. Similarly, conditioned medium from irradiated B16 was also able to increase VEGF secretion in HMECs. Moreover, ionizing radiation–induced melanoma cell invasiveness was partially inhibited by an anti-VEGF monoclonal antibody. In vivo, ionizing radiation plus concomitant Metastat yielded the greatest growth inhibition of melanoma s.c. tumors and this effect correlated with inhibition of angiogenesis as measured by both Doppler ultrasonography and platelet/endothelial cell adhesion molecule-1 staining. Finally, ionizing radiation modulated MMP-2, VEGF, and VEGF receptor expression in these tumor samples using immunohistochemistry. Taken together, these results suggest that there is an ionizing radiation–induced tumor survival pathway and a possible paracrine ionizing radiation–induced stimulatory pathway emanating from tumor cells toward the endothelial bed that is impeded when Metastat is given simultaneously. This model could provide in vivo evidence of the antitumor efficacy of combining a MMP inhibitor with ionizing radiation to target radiation-induced invasion and angiogenesis.
Zinc ribbon domain-containing 1 (ZNRD1) mediates multidrug resistance of leukemia cells through regulation of P-glycoprotein and Bcl-2
Authors: Hong Liu , Piao Ying , Han Yu , Wang Jun , Zhang Xiaoyin , Du Yulei , Cao Shanshan , Qiao Taidong , Chen Zhen , Fan Daiming 
Published: March 2006

Abstract
Here, we investigated the role of zinc ribbon domain-containing 1 (ZNRD1) in multidrug resistance (MDR) of leukemia cells and the possible underlying mechanisms. ZNRD1 was found overexpressed in the vincristine-induced MDR leukemia cell HL-60/vincristine moreso than its parental cell HL-60. Up-regulation of ZNRD1 expression could confer resistance of both P-glycoprotein (P-gp)-related and P-gp-nonrelated drugs on HL-60 cells and suppress Adriamycin-induced apoptosis accompanied by decreased accumulation and increased releasing amount of Adriamycin. ZNRD1 could significantly up-regulate the expression of P-gp, Bcl-2, and the transcription of the MDR1 gene but not alter the expression of MDR-associated protein, glutathione S-transferase activity, or intracellular glutathione content in leukemia cells. In addition, inhibition of ZNRD1 expression by RNA interference or P-gp inhibitor could partially reverse ZNRD1-mediated MDR. The further study of the biological functions of ZNRD1 may be helpful for understanding the mechanisms of MDR of leukemia and developing possible strategies to treat leukemia. [Mol Cancer Ther 2005;4(12):1936–42]
Paclitaxel-resistant cells have a mutation in the paclitaxel-binding region of β-tubulin (Asp26Glu) and less stable microtubules
Authors: Hari Malathi , Loganzo Frank , Annable Tami , Tan Xingzhi , Musto Sylvia , Morilla Daniel B. , Nettles James H. , Snyder James P. , Greenberger Lee M. 
Published: March 2006

AbstractResistance to paclitaxel-based therapy is frequently encountered in the clinic. The mechanisms of intrinsic or acquired paclitaxel resistance are not well understood. We sought to characterize the resistance mechanisms that develop upon chronic exposure of a cancer cell line to paclitaxel in the presence of the P-glycoprotein reversal agent, CL-347099. The epidermoid tumor line KB-3-1 was exposed to increasing concentrations of paclitaxel and 5 μmol/L CL-347099 for up to 1 year. Cells grown in 15 nmol/L paclitaxel plus CL-347099 (KB-15-PTX/099) developed 18-fold resistance to paclitaxel and were dependent upon paclitaxel for maximal growth. They grew well and retained resistance to paclitaxel when grown in athymic mice. Cross-resistance (3- to 5-fold) was observed in tissue culture to docetaxel, the novel taxane MAC-321, and epothilone B. Collateral sensitivity (∼3-fold) was observed to the depolymerizing agents vinblastine, dolastatin-10, and HTI-286. KB-15-PTX/099–resistant cells did not overexpress P-glycoprotein nor did they have an alteration of [14C]paclitaxel accumulation compared with parental cells. However, a novel point mutation (T to A) resulting in Asp26 to glutamate substitution in class I (M40) β-tubulin was found. Based on an electron crystallography structure of Zn-stabilized tubulin sheets, the phenyl ring of C-3′ NHCO-C6H5 of paclitaxel makes contact with Asp26 of β-tubulin, suggesting a ligand-induced mutation. Optimized model complexes of paclitaxel, docetaxel, and MAC-321 in β-tubulin show a novel hydrogen bonding pattern for the glutamate mutant and rationalize the observed resistance profiles. However, a mutation in the paclitaxel binding pocket does not explain the phenotype completely. KB-15-PTX/099 cells have impaired microtubule stability as determined by a reduced percentage of tubulin in microtubules and reflected by less acetylated tubulin. These results suggest that a mutation in tubulin might affect microtubule stability as well as drug binding and contribute to the observed resistance profile. [Mol Cancer Ther 2006;5(2):270–8]