bluuebunny/crossref_metadata_2025 huggingface dataset:
Theoretical Models for Video on Demand Services on Peer-to-Peer Networks
Authors: Branca Giovanni , Schierl Thomas , Atzori Luigi
Published: October 2009

Peer-to-peer networks (P2Ps) are becoming more and more popular in video content delivery services, such as TV broadcast and Video on Demand (VoD), thanks to their scalability feature. Such characteristic allows for higher numbers of simultaneous users at a given server load and bandwidth with respect to alternative solutions. However, great efforts are still required to study and design reliable and QoS-guaranteed solutions. In this paper, within the scenario of P2P-based VoD services, we study the phenomenon of peer churns and propose four models of the peer behaviour to evaluate its impact on the system performance, which are based on the Gilbert-Elliot chain, the fluidic representation of the user behavior, and a queuing analysis of the system. The models are compared by computing the resources the system has to add on top of the P2P network to satisfy all the download requests. Simulations show important relationships between playback buffer length, peer request rate, peer average lifetime, and server upload rate.
The KM-Algorithm Identifies Regulated Genes in Time Series Expression Data
Authors: Bremer Martina , Doerge R. W.
Published: October 2009

We present a statistical method to rank observed genes in gene expression time series experiments according to their degree of regulation in a biological process. The ranking may be used to focus on specific genes or to select meaningful subsets of genes from which gene regulatory networks can be built. Our approach is based on a state space model that incorporates hidden regulators of gene expression. Kalman (K) smoothing and maximum (M) likelihood estimation techniques are used to derive optimal estimates of the model parameters upon which a proposed regulation criterion is based. The statistical power of the proposed algorithm is investigated, and a real data set is analyzed for the purpose of identifying regulated genes in time dependent gene expression data. This statistical approach supports the concept that meaningful biological conclusions can be drawn from gene expression time series experiments by focusing on strong regulation rather than large expression values.
Reverse Genetics for Functional Genomics of Phytopathogenic Fungi and Oomycetes
Authors: Bhadauria Vijai , Banniza Sabine , Wei Yangdou , Peng You-Liang
Published: October 2009

Sequencing of over 40 fungal and oomycete genomes has been completed. The next major challenge in modern fungal/oomycete biology is now to translate this plethora of genome sequence information into biological functions. Reverse genetics has emerged as a seminal tool for functional genomics investigations. Techniques utilized for reverse genetics like targeted gene disruption/replacement, gene silencing, insertional mutagenesis, and targeting induced local lesions in genomes will contribute greatly to the understanding of gene function of fungal and oomycete pathogens. This paper provides an overview on high-throughput reverse genetics approaches to decode fungal/oomycete genomes.
Expression of LGI1 Impairs Proliferation and Survival of HeLa Cells
Authors: Gabellini Nadia , Masola Valentina
Published: October 2009

The LGI1 gene was suggested to function as tumor suppressor for its ability to reduce malignant features of glioblastoma cells. In support to this proposal were the findings that overexpression of LGI1 in neuroblastoma cells inhibited proliferation and induced apoptosis. In this study we performed stable LGI1 expression in HeLa cells to examine whether the noxious effect of LGI1 might be extended to cancer cells of diverse origin. HeLa cell clones stably expressing LGI1 exhibited a significant impairment of proliferation and a consistent increase of cell death when compared with control cells lacking expression of LGI1. Expression of LGI1 increased the activity of apoptosis effectors caspase-3/7; furthermore it downregulated the antiapoptotic BCL2 gene and upregulated the proapoptotic BAX gene expression, suggesting that the cause of HeLa cells death might be an increased susceptibility to apoptosis induced by LGI1. The results suggested that LGI1 is capable to restrain growth and survival of adenocarcinoma cells such as HeLa.
Glaucoma Patients' Trust in the Physician
Authors: Muir Kelly W. , Santiago-Turla Cecilia , Stinnett Sandra S. , Herndon Leon W. , Allingham R. Rand , Challa Pratap , Lee Paul P.
Published: October 2009

Objectives. To describe glaucoma patients' trust in the physician and to test the hypothesis that increased interpersonal trust is associated with increased medication adherence.Methods. One hundred ninety-five subjects with open-angle glaucoma seen by multiple glaucoma subspecialists participated in a cross-sectional patient survey and concomitant chart review which included a test of health literacy and the Trust in Physician Scale (TPS), a scale from 1–100, with 100 indicating greatest trust. Charts were reviewed for visual acuity and visual field results. Subjects' pharmacies were contacted to ascertain medication refill rates over the preceding six months.Results. TPS scores ranged from 57.5 to 100,78.7±8.4(mean±SD,) median 75.0. When age, race, gender, baseline visual acuity and visual field status, education level, and literacy status were considered, only race was associated with TPS. Caucasians expressed slightly higher levels of trust (n=108; TPS80.1±8.2) than non-Caucasians, (n=87(82 Africans Americans); TPS77.1±8.4;P=.012). TPS score was not associated with refill rates (P=.190).Conclusions. Trust in physician is generally high in this group of glaucoma patients but varies slightly by race. Trust in physician was not associated with glaucoma medication adherence in this tertiary care population.
Bridging the Divide between Manual Gating and Bioinformatics with the Bioconductor Package flowFlowJo
Authors: Gosink John J. , Means Gary D. , Rees William A. , Su Cheng , Rand Hugh A.
Published: October 2009

In flow cytometry, different cell types are usually selected or “gated” by a series of 1- or 2-dimensional geometric subsets of the measurements made on each cell. This is easily accomplished in commercial flow cytometry packages but it is difficult to work computationally with the results of this process. The ability to retrieve the results and work with both them and the raw data is critical; our experience points to the importance of bioinformatics tools that will allow us to examine gating robustness, combine manual and automated gating, and perform exploratory data analysis. To provide this capability, we have developed a Bioconductor package called flowFlowJo that can import gates defined by the commercial package FlowJo and work with them in a manner consistent with the other flow packages in Bioconductor. We present this package and illustrate some of the ways in which it can be used.
ARTEMIS Nuclease Facilitates Apoptotic Chromatin Cleavage
Authors: Britton Sébastien , Frit Philippe , Biard Denis , Salles Bernard , Calsou Patrick
Published: October 2009

One hallmark of apoptosis is DNA degradation that first appears as high molecular weight fragments followed by extensive internucleosomal fragmentation. During apoptosis, the DNA-dependent protein kinase (DNA-PK) is activated. DNA-PK is involved in the repair of DNA double-strand breaks (DSB) and its catalytic subunit is associated with the nuclease ARTEMIS. Here, we report that, on initiation of apoptosis in human cells by agents causing DNA DSB or by staurosporine or other agents, ARTEMIS binds to apoptotic chromatin together with DNA-PK and other DSB repair proteins. ARTEMIS recruitment to chromatin showed a time and dose dependency. It required DNA-PK protein kinase activity and was blocked by antagonizing the onset of apoptosis with a pan-caspase inhibitor or on overexpression of the antiapoptotic BCL2 protein. In the absence of ARTEMIS, no defect in caspase-3, poly(ADP-ribose) polymerase-1, and XRCC4 cleavage or in H2AX phosphorylation was observed and DNA-PK catalytic subunit was still phosphorylated on S2056 in response to staurosporine. However, DNA fragmentation including high molecular weight fragmentation was delayed in ARTEMIS-deficient cells compared with cells expressing ARTEMIS. In addition, ARTEMIS enhanced the kinetics of MLL gene cleavage at a breakage cluster breakpoint that is frequently translocated in acute or therapy-related leukemias. These results show a facilitating role for ARTEMIS at least in early, site-specific chromosome breakage during apoptosis. [Cancer Res 2009;69(20):8120–6]
Azacytidine Inhibits RNA Methylation at DNMT2 Target Sites in Human Cancer Cell Lines
Authors: Schaefer Matthias , Hagemann Sabine , Hanna Katharina , Lyko Frank
Published: October 2009

The cytosine analogues azacytidine and decitabine are currently being developed as drugs for epigenetic cancer therapy. Although various studies have shown that both drugs are effective in inhibiting DNA methylation, it has also become clear that their mode of action is not limited to DNA demethylation. Because azacytidine is a ribonucleoside, the primary target of this drug may be cellular RNA rather than DNA. We have now analyzed the possibility that azacytidine inhibits the RNA methyltransferase DNMT2. We found that DNMT2 is variably expressed in human cancer cell lines. RNA bisulfite sequencing showed that azacytidine, but not decitabine, inhibits cytosine 38 methylation of tRNAAsp, a major substrate of DNMT2. Azacytidine caused a substantially stronger effect than decitabine on the metabolic rate of all the cancer cell lines tested, consistent with an effect of this drug on RNA metabolism. Of note, drug-induced loss of RNA methylation seemed specific for DNMT2 target sites because we did not observe any significant demethylation at sites known to be methylated by other RNA methyltransferases. Our results uncover a novel and quantifiable drug activity of azacytidine and raise the possibility that tRNA hypomethylation might contribute to patient responses. [Cancer Res 2009;69(20):8127–32]
Phosphorylation and Degradation of MdmX Is Inhibited by Wip1 Phosphatase in the DNA Damage Response
Authors: Zhang Xinna , Lin Lin , Guo Huarong , Yang Jianhua , Jones Stephen N. , Jochemsen Aart , Lu Xiongbin
Published: October 2009

MdmX and Mdm2 regulate p53 tumor suppressor functions by controlling p53 transcriptional activity and/or stability in cells exposed to DNA damage. Accumulating evidence indicates that ATM-mediated phosphorylation and degradation of Mdm2 and MdmX may be the initial driving force that induces p53 activity during the early phase of the DNA damage response. We have recently determined that a novel protein phosphatase, Wip1 (or PPM1D), contributes to p53 regulation by dephosphorylating Mdm2 to close the p53 activation loop initiated by the ATM/ATR kinases. In the present study, we determine that Wip1 directly dephosphorylates MdmX at the ATM-targeted Ser403 and indirectly suppresses phosphorylation of MdmX at Ser342 and Ser367. Wip1 inhibits the DNA damage–induced ubiquitination and degradation of MdmX, leading to the stabilization of MdmX and reduction of p53 activities. Our data suggest that Wip1 is an important component in the ATM-p53-MdmX regulatory loop. [Cancer Res 2009;69(20):7960–8]
Increased Expression of Androgen Receptor Sensitizes Prostate Cancer Cells to Low Levels of Androgens
Authors: Waltering Kati K. , Helenius Merja A. , Sahu Biswajyoti , Manni Visa , Linja Marika J. , Jänne Olli A. , Visakorpi Tapio
Published: October 2009

Androgen receptor (AR) is known to be overexpressed in castration-resistant prostate cancer. To interrogate the functional significance of the AR level, we established two LNCaP cell sublines expressing in a stable fashion two to four times (LNCaP-ARmo) and four to six times (LNCaP-ARhi) higher level of AR than the parental cell line expressing the empty vector (LNCaP-pcDNA3.1). LNCaP-ARhi cell line grew faster than the control line in low concentrations, especially in 1 nmol/L 5α-dihydrotestosterone (DHT). Microarray-based transcript profiling and subsequent unsupervised hierarchical clustering showed that LNCaP-ARhi cells clustered together with VCaP cells, containing endogenous AR gene amplification and overexpression, indicating the central role of AR in the overall regulation of gene expression in prostate cancer cells. Two hundred forty genes showed >2-fold changes on DHT treatment in LNCaP-ARhi at 4 h time point, whereas only 164 and 52 showed changes in LNCaP-ARmo and LNCaP-pcDNA3.1, respectively. Many androgen-regulated genes were upregulated in LNCaP-ARhi at 10-fold lower concentration of DHT than in control cells. DHT (1 nmol/L) increased expression of several cell cycle–associated genes in LNCaP-ARhi cells. ChIP-on-chip assay revealed the presence of chromatin binding sites for AR within ±200 kb of most of these genes. The growth of LNCaP-ARhi cells was also highly sensitive to cyclin-dependent kinase inhibitor, roscovitine, at 1nmol/L DHT. In conclusion, our results show that overexpression of AR sensitizes castration-resistant prostate cancer cells to the low levels of androgens. The activity of AR signaling pathway is regulated by the levels of both ligand and the receptor. [Cancer Res 2009;69(20):8141–9]
Runt-Related Transcription Factor RUNX3 Is a Target of MDM2-Mediated Ubiquitination
Authors: Chi Xin-Zi , Kim Jiyeon , Lee Yong-Hee , Lee Jung-Won , Lee Kyeong-Sook , Wee Heejun , Kim Wun-Jae , Park Woo-Yoon , Oh Byung-Chul , Stein Gary S. , Ito Yoshiaki , van Wijnen Andre J. , Bae Suk-Chul
Published: October 2009

The p14ARF-MDM2-p53 pathway constitutes an effective mechanism for protecting cells from oncogenic stimuli such as activated Ras and Myc. Importantly, Ras activation induces p14ARF and often occurs earlier than p53 inactivation during cancer development. Here, we show that RUNX3, a tumor suppressor in various tumors including stomach, bladder, colon, and lung, is stabilized by Ras activation through the p14ARF-MDM2 signaling pathway. RUNX3 directly binds MDM2 through its Runt-related DNA-binding domain. MDM2 blocks RUNX3 transcriptional activity by interacting with RUNX3 through an acidic domain adjacent to the p53-binding domain of MDM2 and ubiquitinates RUNX3 on key lysine residues to mediate nuclear export and proteasomal degradation. Our data indicate that the lineage-specific tumor suppressor RUNX3 and the ubiquitous p53 protein are both principal responders of the p14ARF-MDM2 cell surveillance pathway that prevents pathologic consequences of abnormal oncogene activation. [Cancer Res 2009;69(20):8111–9]
Enhanced Leukemia Cell Detection Using a Novel Magnetic Needle and Nanoparticles
Authors: Jaetao Jason E. , Butler Kimberly S. , Adolphi Natalie L. , Lovato Debbie M. , Bryant Howard C. , Rabinowitz Ian , Winter Stuart S. , Tessier Trace E. , Hathaway Helen J. , Bergemann Christian , Flynn Edward R. , Larson Richard S.
Published: October 2009

Acute leukemia is a hematopoietic malignancy for which the accurate measurement of minimal residual disease is critical to determining prognosis and treatment. Although bone marrow aspiration and light microscopy remain the current standard of care for detecting residual disease, these approaches cannot reliably discriminate less than 5% lymphoblast cells. To improve the detection of leukemia cells in the marrow, we developed a novel apparatus that utilizes antibodies conjugated to superparamagnetic iron oxide nanoparticles (SPION) and directed against the acute leukemia antigen CD34, coupled with a “magnetic needle” biopsy. Leukemia cell lines expressing high or minimal CD34 were incubated with anti-CD34–conjugated SPIONs. Three separate approaches including microscopy, superconducting quantum interference device magnetometry, and in vitro magnetic needle extraction were then used to assess cell sampling. We found that CD34-conjugated nanoparticles preferentially bind high CD34-expressing cell lines. Furthermore, the magnetic needle enabled identification of both cell line and patient leukemia cells diluted into normal blood at concentrations below those normally found in remission marrow samples. Finally, the magnetic needle enhanced the percentage of lymphoblasts detectable by light microscopy by 10-fold in samples of fresh bone marrow aspirate approximating minimal residual disease. These data suggest that bone marrow biopsy using antigen-targeted magnetic nanoparticles and a magnetic needle for the evaluation of minimal residual disease in CD34-positive acute leukemias can significantly enhance sensitivity compared with the current standard of care. [Cancer Res 2009;69(21):8310–6]
Aldehyde Dehydrogenase–Expressing Colon Stem Cells Contribute to Tumorigenesis in the Transition from Colitis to Cancer
Authors: Carpentino Joseph E. , Hynes Mark J. , Appelman Henry D. , Zheng Tong , Steindler Dennis A. , Scott Edward W. , Huang Emina H.
Published: October 2009

Patients with chronic ulcerative colitis are at increased risk of developing colorectal cancer. Although current hypotheses suggest that sporadic colorectal cancer is due to inability to control cancer stem cells, the cancer stem cell hypothesis has not yet been validated in colitis-associated cancer. Furthermore, the identification of the colitis to cancer transition is challenging. We recently showed that epithelial cells with the increased expression of aldehyde dehydrogenase in sporadic colon cancer correlate closely with tumor-initiating ability. We sought to determine whether ALDH can be used as a marker to isolate tumor-initiating populations from patients with chronic ulcerative colitis. We used fluorescence-activated cell sorting to identify precursor colon cancer stem cells from colitis patients and report both their transition to cancerous stem cells in xenografting studies as well as their ability to generate spheres in vitro. Similar to sporadic colon cancer, these colitis-derived tumors were capable of propagation as sphere cultures. However, unlike the origins of sporadic colon cancer, the primary colitic tissues did not express any histologic evidence of dysplasia. To elucidate a potential mechanism for our findings, we compared the stroma of these different environments and determined that at least one paracrine factor is up-regulated in the inflammatory and malignant stroma compared with resting, normal stroma. These data link colitis and cancer identifying potential tumor-initiating cells from colitic patients, suggesting that sphere and/or xenograft formation will be useful to survey colitic patients at risk of developing cancer. [Cancer Res 2009;69(20):8208–15]
A Novel PTEN-Dependent Link to Ubiquitination Controls FLIPS Stability and TRAIL Sensitivity in Glioblastoma Multiforme
Authors: Panner Amith , Crane Courtney A. , Weng Changjiang , Feletti Alberto , Parsa Andrew T. , Pieper Russell O.
Published: October 2009

Phosphatase and tensin homologue (PTEN) loss and activation of the Akt-mammalian target of rapamycin (mTOR) pathway increases mRNA translation, increases levels of the antiapoptotic protein FLIPS, and confers resistance to tumor necrosis factor–related apoptosis-inducing ligand (TRAIL)–induced apoptosis in glioblastoma multiforme (GBM). In PTEN-deficient GBM cells, however, the FLIPS protein also exhibited a longer half-life than in PTEN mutant GBM cells, and this longer half-life correlated with decreased FLIPS polyubiquitination. FLIPS half-life in PTEN mutant GBM cells was reduced by exposure to an Akt inhibitor, but not to rapamycin, suggesting the existence of a previously undescribed, mTOR-independent linkage between PTEN and the ubiquitin-dependent control of protein stability. Total levels of the candidate FLIPS E3 ubiquitin ligase atrophin-interacting protein 4 (AIP4) were comparable in PTEN wild-type (WT) and PTEN mutant GBM cells, although in PTEN-deficient cells, AIP4 was maintained in a stable polyubiquitinated state that was less able to associate with FLIPS or with the FLIPS-containing death inducing signal complex. Small interfering RNA–mediated suppression of AIP4 levels in PTEN WT cells decreased FLIPS ubiquitination, prolonged FLIPS half-life, and increased TRAIL resistance. Similarly, the Akt activation that was previously shown to increase TRAIL resistance did not alter AIP4 levels, but increased AIP4 ubiquitination, increased FLIPS steady-state levels, and suppressed FLIPS ubiquitination. These results define the PTEN-Akt-AIP4 pathway as a key regulator of FLIPS ubiquitination, FLIPS stability, and TRAIL sensitivity and also define a novel link between PTEN and the ubiquitin-mediated control of protein stability. [Cancer Res 2009;69(20):7911–6]
Apolipoprotein E/C1 Locus Variants Modify Renal Cell Carcinoma Risk
Authors: Moore Lee E. , Brennan Paul , Karami Sara , Menashe Idan , Berndt Sonja I. , Dong Linda M. , Meisner Allison , Yeager Meredith , Chanock Stephen , Colt Joanne , Schwartz Kendra , Davis Faith , Zaridze David , Mattveev Vsevolod , Janout Vladimir , Kollarova Hellena , Bencko Vladimir , Navratilova Marie , Szeszenia-Dabrowska Neonilia , Mates Dana , Holcatova Ivana , Boffetta Paolo , Chow Wong-Ho , Rosenberg Philip S. , Rothman Nathaniel
Published: October 2009

Lipid peroxidation is considered a unifying mechanistic pathway through which known risk factors induce renal cell carcinoma (RCC). We hypothesized that genes selected a priori for their role in lipid peroxidation would modify cancer risk. We genotyped 635 single nucleotide polymorphisms (SNP) in 38candidate genes in 777 Caucasian RCC cases and 1,035 controls enrolled in a large European case-control study. Top candidate SNPs were confirmed among 718 Caucasian cases and 615 controls in a second study in the United States. Two of the three SNPs (rs8106822 and rs405509) that replicated in the U.S. study were within a regulatory region of the APOE promoter. The OR for rs8106822 A>G variant was 1.22AG and 1.41GG (Ptrend = 0.01) in the European study, 1.05AG and 1.51GG (Ptrend = 0.03) in the U.S. study, and 1.15AG and 1.44GG (Ptrend = 0.001) among 1,485 cases and 1,639 controls combined. The rs405509 G>T variant was associated with risk in the European (OR, 0.87TG; OR, 0.71TT; Ptrend = 0.02), the U.S. (OR, 0.68TG; OR, 0.71TT; Ptrend = 0.02), and both studies combined (ORTG, 0.79; ORTT, 0.71; Ptrend = 0.001), as was the G-G haplotype (r2 = 0.64; P= 4.7 × 10−4). This association is biologically plausible as SNP rs405509 was shown to modify protein binding and transcriptional activity of the APOE protein in vitro and is in linkage disequilibrium with key known variants defining the e2, e3, and e4 alleles that modify risk of atherosclerosis, Alzheimer's disease risk, and progression to AIDS. In two large case-control studies, our findings further define a functional region of interest at the APOE locus that increases RCC susceptibility. [Cancer Res 2009;69(20):8001–8]
Tumor Antigen–Specific FOXP3+ CD4 T Cells Identified in Human Metastatic Melanoma: Peptide Vaccination Results in Selective Expansion of Th1-like Counterparts
Authors: Jandus Camilla , Bioley Gilles , Dojcinovic Danijel , Derré Laurent , Baitsch Lukas , Wieckowski Sébastien , Rufer Nathalie , Kwok William W. , Tiercy Jean-Marie , Luescher Immanuel F. , Speiser Daniel E. , Romero Pedro
Published: October 2009

We have previously shown that vaccination of HLA-A2 metastatic melanoma patients with the analogue Melan-A26-35(A27L) peptide emulsified in a mineral oil induces ex vivo detectable specific CD8 T cells. These are further enhanced when a TLR9 agonist is codelivered in the same vaccine formulation. Interestingly, the same peptide can be efficiently recognized by HLA-DQ6–restricted CD4 T cells. We used HLA-DQ6 multimers to assess the specific CD4 T-cell response in both healthy individuals and melanoma patients. We report that the majority of melanoma patients carry high frequencies of naturally circulating HLA-DQ6–restricted Melan-A–specific CD4 T cells, a high proportion of which express FOXP3 and proliferate poorly in response to the cognate peptide. Upon vaccination, the relative frequency of multimer+ CD4 T cells did not change significantly. In contrast, we found a marked shift to FOXP3-negative CD4 T cells, accompanied by robust CD4 T-cell proliferation upon in vitro stimulation with cognate peptide. A concomitant reduction in TCR diversity was also observed. This is the first report on direct ex vivo identification of antigen-specific FOXP3+ T cells by multimer labeling in cancer patients and on the direct assessment of the impact of peptide vaccination on immunoregulatory T cells. [Cancer Res 2009;69(20):8085–93]
Tpl2 Is a Key Mediator of Arsenite-Induced Signal Transduction
Authors: Lee Kyung Mi , Lee Ki Won , Bode Ann M. , Lee Hyong Joo , Dong Zigang
Published: October 2009

Arsenite is a well-known human carcinogen that especially targets skin. The tumor progression locus 2 (Tpl2) gene encodes a serine/threonine protein kinase that is overexpressed in various cancer cells. However, the relevance of Tpl2 in arsenite-induced carcinogenesis and the underlying mechanisms remain to be explored. We show that arsenite increased Tpl2 kinase activity and its phosphorylation in mouse epidermal JB6 P+ cells in a dose- and time-dependent manner. Exposure to arsenite resulted in a marked induction of cyclooxygenase-2 (COX-2) and prostaglandin E2 (PGE2), important mediators of inflammation and tumor promotion. Treatment with a Tpl2 kinase inhibitor or Tpl2 short hairpin RNA suppressed COX-2 expression and PGE2 production induced by arsenite treatment, suggesting that Tpl2 is critical in arsenite-induced carcinogenesis. We also found that arsenite-induced phosphorylation of extracellular signal-regulated kinases (ERK) or c-Jun NH2-terminal kinases (JNK) was markedly suppressed by Tpl2 kinase inhibitor or Tpl2 short hairpin RNA. Inhibition of arsenite-induced ERK or JNK signaling using a pharmacologic inhibitor of ERK or JNK substantially blocked COX-2 expression. Furthermore, inhibition of Tpl2 reduced the arsenite-induced promoter activity of NF-κB and activator protein-1 (AP-1), indicating that NF-κB and AP-1 are downstream transducers of arsenite-triggered Tpl2. Our results show that Tpl2 plays a key role in arsenite-induced COX-2 expression and PGE2 production and further elucidate the role of Tpl2 in arsenite signals that activate ERK/JNK and NF-κB/AP-1 in JB6 P+ cells. [Cancer Res 2009;69(20):8043–9]
Recent Advances in Cancer Therapy Targeting Proteins Involved in DNA Double-Strand Break Repair
Authors: Bolderson Emma , Richard Derek J. , Zhou Bin-Bing S. , Khanna Kum Kum
Published: October 2009

Damage to genetic material represents a persistent and ubiquitous threat to genomic stability. Once DNA damage is detected, a multifaceted signaling network is activated that halts the cell cycle, initiates repair, and in some instances induces apoptotic cell death. In this article, we will review DNA damage surveillance networks, which maintain the stability of our genome, and discuss the efforts underway to identify chemotherapeutic compounds targeting the core components of DNA double-strand breaks (DSB) response pathway. The majority of tumor cells have defects in maintaining genomic stability owing to the loss of an appropriate response to DNA damage. New anticancer agents are exploiting this vulnerability of cancer cells to enhance therapeutic indexes, with limited normal tissue toxicity. Recently inhibitors of the checkpoint kinases Chk1 and Chk2 have been shown to sensitize tumor cells to DNA damaging agents. In addition, the treatment of BRCA1- or BRCA2-deficient tumor cells with poly(ADP-ribose) polymerase (PARP) inhibitors also leads to specific tumor killing. Due to the numerous roles of p53 in genomic stability and its defects in many human cancers, therapeutic agents that restore p53 activity in tumors are the subject of multiple clinical trials. In this article we highlight the proteins mentioned above and catalog several additional players in the DNA damage response pathway, including ATM, DNA-PK, and the MRN complex, which might be amenable to pharmacological interventions and lead to new approaches to sensitize cancer cells to radio- and chemotherapy. The challenge is how to identify those patients most receptive to these treatments. (Clin Cancer Res 2009;15(20):6314–20)
Therapy-induced PML/RARA Proteolysis and Acute Promyelocytic Leukemia Cure
Authors: Nasr Rihab , Lallemand-Breitenbach Valérie , Zhu Jun , Guillemin Marie-Claude , de Thé Hugues
Published: October 2009

Acute promyelocytic leukemia (APL) is characterized by a specific t(15;17) chromosomal translocation that yields the PML/RARA fusion gene. Clinically, besides chemotherapy, two drugs induce clinical remissions: retinoic acid (RA) and arsenic trioxide (As). Both agents directly target PML/RARA-mediated transcriptional repression and protein stability, inducing to various extent promyelocyte differentiation and clinical remission of APL patients. RA targets the RARA moiety of the fusion, whereas arsenic targets its PML part. PML/RARA expression in the mouse is sufficient to initiate APL. The RA-As association, which synergizes for PML/RARA degradation but not for differentiation, rapidly clears leukemia initiating cells (LIC), resulting in APL eradication in murine APL models, but also in several APL clinical trials. Cyclic AMP triggered PML/RARA phosphorylation also enhances RA-induced APL regression, PML/RARA degradation, and LIC clearance, raising new options for therapy-resistant patients. Although differentiation has a major role in debulking of the tumor, PML/RARA degradation seems to be the primary basis for APL eradication by the RA-As association. Oncoprotein degradation could be a general therapeutic strategy that may be extended beyond APL. (Clin Cancer Res 2009;15(20):6321–6)
Polysomy for Chromosomes 1 and 19 Predicts Earlier Recurrence in Anaplastic Oligodendrogliomas with Concurrent 1p/19q Loss
Authors: Snuderl Matija , Eichler April F. , Ligon Keith L. , Vu Quynh U. , Silver Michael , Betensky Rebecca A. , Ligon Azra H. , Wen Patrick Y. , Louis David N. , Iafrate A. John
Published: October 2009

Purpose: Loss of chromosome arms 1p and 19q is a molecular feature of oligodendroglial tumors characterized by responsiveness to chemotherapy and a favorable prognosis. The purpose of this study was to evaluate the prognostic significance of polysomy of chromosomes 1 and 19 in the setting of 1p/19q codeletion.

Experimental Design: We analyzed 64 anaplastic oligodendrogliomas with 1p/19q loss or maintenance diagnosed at Massachusetts General Hospital and Brigham and Women's Hospital from 1996 to 2005; fluorescence in situ hybridization for 1p/19q and Ki-67 immunohistochemistry was done. Polysomy was defined as more than two 1q and 19p signals in >30% of the cells with concurrent 1p/19q deletion. Tumors were divided into groups based on their 1p/19q status and compared for progression-free survival, overall survival, and 5-year survival probabilities.

Results: Forty-six tumors (72%) in our cohort had 1p/19q loss and 18 (28%) had 1p/19q maintenance. Of those with loss, 19 (41%) had concurrent polysomy and 27 (59%) lacked polysomy. In agreement with previous studies, the group of anaplastic oligodendrogliomas with 1p/19q loss had significantly better progression-free survival and overall survival than anaplastic oligodendrogliomas with 1p/19q maintenance (P = 0.0009 and P < 0.0003, respectively). Among anaplastic oligodendrogliomas with 1p/19q loss, those with polysomy showed shorter progression-free survival than those with 1p/19q loss without polysomy (P = 0.0048). Overall survival was similar in tumors with and without polysomy. The Ki-67 labeling index was not associated with polysomy and did not have prognostic significance.

Conclusion: The presence of polysomy in anaplastic oligodendrogliomas with deletion of 1p/19q is a marker of earlier recurrence. (Clin Cancer Res 2009;15(20):6430–7)
Oral Poly(ADP-Ribose) Polymerase-1 Inhibitor BSI-401 Has Antitumor Activity and Synergizes with Oxaliplatin against Pancreatic Cancer, Preventing Acute Neurotoxicity
Authors: Melisi Davide , Ossovskaya Valeria , Zhu Cihui , Rosa Roberta , Ling Jianhua , Dougherty Patrick M. , Sherman Barry M. , Abbruzzese James L. , Chiao Paul J.
Published: October 2009

Purpose: Development of novel agents and drug combinations are urgently needed for treatment of pancreatic cancer. Oxaliplatin belongs to an important class of DNA-damaging organoplatinum agents, useful in pancreatic cancer therapy. However, increased ability of cancer cells to recognize and repair DNA damage enables resistance to these agents. Poly (ADP ribose) polymerase-1 is a sensor of DNA damage with key roles in DNA repair. Here, we report the therapeutic activity of the poly (ADP ribose) polymerase-1 inhibitor BSI-401, as a single agent and in combination with oxaliplatin in orthotopic nude mouse models of pancreatic cancer, and its effect on oxaliplatin-induced acute neurotoxicity.

Experimental Design: We determined in vitro the effect of BSI-401 and its synergism with oxaliplatin on the growth of pancreatic cancer cells. Activity of different dosages of parenteral and oral BSI-401, alone and in combination with oxaliplatin, was evaluated in orthotopic nude mouse models with luciferase-expressing pancreatic cancer cells. The effect of BSI-401 in preventing oxaliplatin-induced acute cold allodynia was measured in rats using a temperature-controlled plate.

Results: BSI-401 alone and in synergism with oxaliplatin significantly inhibited the growth of pancreatic cancer cells in vitro. In nude mice, i.p. [200 mg/kg once a week (QW) × 4] and oral [400 mg/kg days 1-5 of each week (QD5 + R2) × 4] administration of BSI-401 significantly reduced tumor burden and prolonged survival (46 versus 144 days, P = 0.0018; 73 versus 194 days, P = 0.0017) compared with no treatment. BSI-401 combined with oxaliplatin had potent synergistic antitumor activity (46 versus 132 days, P = 0.0063), and significantly (P = 0.0148) prevented acute oxaliplatin-induced neurotoxicity.

Conclusions: BSI-401, alone or in combination with oxaliplatin, is a promising new therapeutic agent that warrants further evaluation for treatment of pancreatic cancer. (Clin Cancer Res 2009;15(20):6367–77)
Loss of SNF5 Expression Correlates with Poor Patient Survival in Melanoma
Authors: Lin Hanyang , Wong Ronald P.C. , Martinka Magdalena , Li Gang
Published: October 2009

Purpose: Aberrant expression of SWI/SNF chromatin remodeling complex is involved in cancer development. The tumor suppressor SNF5, the core subunit of SWI/SNF complex, has been shown to regulate cell differentiation, cell cycle control, and apoptosis. To investigate the role of SNF5 in the development of melanoma, we examined the expression of SNF5 in melanocytic lesions at different stages and analyzed the correlation between SNF5 expression and clinicopathologic variables and patient survival.

Experimental Design: Using tissue microarry and immunohistochemistry, we evaluated SNF5 staining in 51 dysplastic nevi, 88 primary melanomas, and 48 metastatic melanomas. We studied chemosensitivity of melanoma cells with reduced SNF5 expression by siRNA using cell survival and apoptosis assays.

Results: SNF5 expression was reduced in metastatic melanoma compared with dysplastic nevi (P = 0.005), in advanced primary melanoma (Clark's level V) compared with low risk Clark's level II melanoma (P = 0.019), and in melanoma at sun-exposed sites compared with sun-protected sites (P = 0.044). Furthermore, we showed a strong correlation between negative SNF5 expression and a worse 5-year survival in melanoma patients (P = 0.016). Multivariate Cox regression analysis revealed that negative SNF5 expression is an independent prognostic factor to predict patient outcome in primary melanomas (P = 0.031). Finally, we showed that knockdown of SNF5 in melanoma cell lines resulted in significant chemoresistance.

Conclusions: Our data indicate that SNF5 may be an important marker for human melanoma progression and prognosis as well as a potential therapeutic target. (Clin Cancer Res 2009;15(20):6404–11)
Quantification of endothelial cell–targeted anti–Bcl-2 therapy and its suppression of tumor growth and vascularization
Authors: Jain Harsh Vardhan , Nör Jacques Eduardo , Jackson Trachette Levon
Published: October 2009

Proapoptotic and antiapoptotic proteins in the Bcl family are key regulators of programmed cell death. It is the interaction between these molecules that determines cellular response to apoptotic signals, making them attractive targets for therapeutic intervention. In recent experiments designed to study tumor angiogenesis, Bcl-2 upregulation in endothelial cells was shown to be a critical mediator of vascular development. In this article, we develop a mathematical model that explicitly incorporates the response of endothelial cells to variations in proapoptotic and antiapoptotic proteins in the Bcl family, as well as the administration of specific antiangiogenic therapies targeted against Bcl-2. The model is validated by comparing its predictions to in vitro experimental data that reports microvessel density prior to and following the administration of 0.05 to 5.0 μmol/L of BL193, a promising small molecule inhibitor of Bcl-2. Numerical simulations of in vivo treatment of tumors predict the existence of a threshold for the amount of therapy required for successful treatment and quantify how this threshold varies with the stage of tumor growth. Furthermore, the model shows how rapidly the least effective dosage of BL193 decreases if an even moderately better inhibitor of Bcl-2 is used and predicts that increasing cell wall permeability of endothelial cells to BL193 does not significantly affect this threshold. A critical challenge of experimental therapeutics for cancer is to decide which drugs are the best candidates for clinical trials. These results underscore the potential of mathematical modeling to guide the development of novel antiangiogenic therapies and to direct drug design. [Mol Cancer Ther 2009;8(10):2926–36]
A peptide fragment of azurin induces a p53-mediated cell cycle arrest in human breast cancer cells
Authors: Yamada Tohru , Mehta Rajeshwari R. , Lekmine Fatima , Christov Konstantin , King Marissa L. , Majumdar Dibyen , Shilkaitis Anne , Green Albert , Bratescu Laura , Beattie Craig W. , Das Gupta Tapas K.
Published: October 2009

We report that amino acids 50 to 77 of azurin (p28) preferentially enter the human breast cancer cell lines MCF-7, ZR-75-1, and T47D through a caveolin-mediated pathway. Although p28 enters p53 wild-type MCF-7 and the isogenic p53 dominant-negative MDD2 breast cancer cell lines, p28 only induces a G2-M-phase cell cycle arrest and apoptosis in MCF-7 cells. p28 exerts its antiproliferative activity by reducing proteasomal degradation of p53 through formation of a p28:p53 complex within a hydrophobic DNA-binding domain (amino acids 80-276), increasing p53 levels and DNA-binding activity. Subsequent elevation of the cyclin-dependent kinase inhibitors p21 and p27 reduces cyclin-dependent kinase 2 and cyclin A levels in a time-dependent manner in MCF-7 cells but not in MDD2 cells. These results suggest that p28 and similar peptides that significantly reduce proteasomal degradation of p53 by a MDM2-independent pathway(s) may provide a unique series of cytostatic and cytotoxic (apoptotic) chemotherapeutic agents. [Mol Cancer Ther 2009;8(10):2947–58]
Triptolide is an inhibitor of RNA polymerase I and II–dependent transcription leading predominantly to down-regulation of short-lived mRNA
Authors: Vispé Stéphane , DeVries Luc , Créancier Laurent , Besse Jérome , Bréand Sophie , Hobson David J. , Svejstrup Jesper Q. , Annereau Jean-Philippe , Cussac Didier , Dumontet Charles , Guilbaud Nicolas , Barret Jean-Marc , Bailly Christian
Published: October 2009

Triptolide, a natural product extracted from the Chinese plant Tripterygium wilfordii, possesses antitumor properties. Despite numerous reports showing the proapoptotic capacity and the inhibition of NF-κB–mediated transcription by triptolide, the identity of its cellular target is still unknown. To clarify its mechanism of action, we further investigated the effect of triptolide on RNA synthesis in the human non–small cell lung cancer cell line A549. Triptolide inhibited both total RNA and mRNA de novo synthesis, with the primary action being on the latter pool. We used 44K human pan-genomic DNA microarrays and identified the genes primarily affected by a short treatment with triptolide. Among the modulated genes, up to 98% are down-regulated, encompassing a large array of oncogenes including transcription factors and cell cycle regulators. We next observed that triptolide induced a rapid depletion of RPB1, the RNA polymerase II main subunit that is considered a hallmark of a transcription elongation blockage. However, we also show that triptolide does not directly interact with the RNA polymerase II complex nor does it damage DNA. We thus conclude that triptolide is an original pharmacologic inhibitor of RNA polymerase activity, affecting indirectly the transcription machinery, leading to a rapid depletion of short-lived mRNA, including transcription factors, cell cycle regulators such as CDC25A, and the oncogenes MYC and Src. Overall, the data shed light on the effect of triptolide on transcription, along with its novel potential applications in cancers, including acute myeloid leukemia, which is in part driven by the aforementioned oncogenic factors. [Mol Cancer Ther 2009;8(10):2780–90]
Apomab, a fully human agonistic antibody to DR5, exhibits potent antitumor activity against primary and metastatic breast cancer
Authors: Zinonos Irene , Labrinidis Agatha , Lee Michelle , Liapis Vasilios , Hay Shelley , Ponomarev Vladimir , Diamond Peter , Zannettino Andrew C.W. , Findlay David M. , Evdokiou Andreas
Published: October 2009

Apomab, a fully human agonistic DR5 monoclonal antibody, triggers apoptosis through activation of the extrinsic apoptotic signaling pathway. In this study, we assessed the cytotoxic effect of Apomab in vitro and evaluated its antitumor activity in murine models of breast cancer development and progression. MDA-MB-231-TXSA breast cancer cells were transplanted into the mammary fat pad or directly into the tibial marrow cavity of nude mice. Apomab was administered early, postcancer cell transplantation, or after tumors progressed to an advanced stage. Tumor burden was monitored progressively using bioluminescence imaging, and the development of breast cancer–induced osteolysis was measured using microcomputed tomography. In vitro, Apomab treatment induced apoptosis in a panel of breast cancer cell lines but was without effect on normal human primary osteoblasts, fibroblasts, or mammary epithelial cells. In vivo, Apomab exerted remarkable tumor suppressive activity leading to complete regression of well-advanced mammary tumors. All animals transplanted with breast cancer cells directly into their tibiae developed large osteolytic lesions that eroded the cortical bone. In contrast, treatment with Apomab following an early treatment protocol inhibited both intraosseous and extraosseous tumor growth and prevented breast cancer–induced osteolysis. In the delayed treatment protocol, Apomab treatment resulted in the complete regression of advanced tibial tumors with progressive restoration of both trabecular and cortical bone leading to full resolution of osteolytic lesions. Apomab represents a potent immunotherapeutic agent with strong activity against the development and progression of breast cancer and should be evaluated in patients with primary and metastatic disease. [Mol Cancer Ther 2009;8(10):2969–80]
The Role of MET Activation in Determining the Sensitivity to Epidermal Growth Factor Receptor Tyrosine Kinase Inhibitors
Authors: Rho Jin Kyung , Choi Yun Jung , Lee Jin Kyung , Ryoo Baek-Yeol , Na Im Il , Yang Sung Hyun , Lee Seung Sook , Kim Cheol Hyeon , Yoo Young Do , Lee Jae Cheol
Published: October 2009

The development of resistance to epidermal growth factor receptor tyrosine kinase inhibitors (EGFR-TKI) seems almost inevitable, even in patients with lung cancer that initially respond well to EGFR-TKIs. MET amplification was recently found to be a mechanism of escape from the anticancer effect of EGFR inhibitors. In the present study, we investigated the means whereby MET affects sensitivity to EGFR-TKIs in PC-9 cells. Gefitinib- or erlotinib-resistant sublines were established by exposing the parental PC-9 cell line to chronic, repeated treatments with these drugs. These resistant sublines showed more than 100-fold more resistance to gefitinib and erlotinib and acquired cross-resistance to other EGFR-TKIs. The T790M EGFR mutation was found by pyrosequencing, and this seemed to be the cause of drug resistance. Resistant cells also showed MET activation, although gene amplification was not detected. Furthermore, the induction of MET activity was not found to be associated with sensitivity to EGFR-TKIs. Interestingly, increased passage number without exposure to gefitinib or erlotinib caused MET activation, but this did not affect sensitivity to EGFR-TKIs. In addition, hepatocyte growth factor was found to block the ability of EGFR-TKIs to inhibit MET activation. However, sustained MET activation by hepatocyte growth factor did not modulate the cellular effects of gefitinib or erlotinib. Rather, activated MET enhanced migration and invasion abilities. Summarizing, MET activation may be acquired during cancer cell proliferation and enhances migratory and invasive abilities without affecting cellular sensitivity to EGFR-TKIs. Accordingly, the present study suggests that MET activation caused by factors other than MET gene amplification is not a suitable surrogate marker of resistance to EGFR-TKIs. (Mol Cancer Res 2009;7(10):1736–43)
Integrin (α6β4) Signals Through Src to Increase Expression of S100A4, a Metastasis-Promoting Factor: Implications for Cancer Cell Invasion
Authors: Kim Tae Hyong , Kim Hong Im , Soung Young Hwa , Shaw Leslie A. , Chung Jun
Published: October 2009

Integrin α6β4 is linked to cancer cell motility and invasion in aggressive and metastatic cancer cells. In this study, we showed that expression of the β4 integrin in MDA-MB-435 cancer cells (MDA-MB-435/β4) leads to a dramatic increase in expression of a metastasis-promoting factor, S100A4, as determined by affymetrix gene chip microarray, quantitative real-time PCR, and Western blot analysis. Alternatively, knocking down β4 integrin expression in MDA-MB-231 breast carcinoma cells by shRNA reduced the level of S100A4 expression. The mechanism by which α6β4 enhances S100A4 expression involves Src, Akt, and NFAT. We have further shown that Y1494, a tyrosine residue of the ITIM motif in the cytoplasmic domain of the β4 integrin subunit, is essential for α6β4-dependent S100A4 expression. Reduction of S100A4 expression by shRNA blocked migration, invasion, and anchorage-independent growth of MDA-MB-435/β4, SUM-159, and MDA-MB-231 cells. These studies define a novel mechanism by which integrin α6β4 promotes cancer cell motility and invasion, and provides insight into how S100A4 expression is regulated in cancer cells. (Mol Cancer Res 2009;7(10):1605–12)