bluuebunny/crossref_metadata_2025 huggingface dataset:
Enhanced radiosensitization of human glioma cells by combining inhibition of poly(ADP-ribose) polymerase with inhibition of heat shock protein 90
Authors: Dungey Fiona A. , Caldecott Keith W. , Chalmers Anthony J.
Published: August 2009

Glioblastoma multiforme (GBM) are the most common primary brain tumor and are resistant to standard therapies. The nondividing nature of normal brain provides an opportunity to enhance the therapeutic ratio by combining radiation with inhibitors of replication-specific DNA repair pathways. Based on our previous findings that inhibition of poly(ADP-ribose) polymerase (PARP) increases radiosensitivity of human glioma cells in a replication-dependent manner and generates excess DNA breaks that are repaired by homologous recombination (HR), we hypothesized that inhibition of HR would amplify the replication-specific radiosensitizing effects of PARP inhibition. Specific inhibitors of HR are not available, but the heat shock protein 90 inhibitor 17-allylamino-17-demethoxygeldanamycin (17-AAG) has been reported to inhibit HR function. The radiosensitizing effects of 17-AAG and the PARP inhibitor olaparib were assessed, and the underlying mechanisms explored. 17-AAG down-regulated Rad51 and BRCA2 protein levels, abrogated induction of Rad51 foci by radiation, and inhibited HR measured by the I-Sce1 assay. Individually, 17-AAG and olaparib had modest, replication-dependent radiosensitizing effects on T98G glioma cells. Additive radiosensitization was observed with combination treatment, mirrored by increases in γH2AX foci in G2-phase cells. Unlike olaparib, 17-AAG did not increase radiation sensitivity of Chinese hamster ovary cells, indicating tumor specificity. However, 17-AAG also enhanced radiosensitivity in HR-deficient cells, indicating that its effects were only partially mediated by HR inhibition. Additional mechanisms are likely to include destabilization of oncoproteins that are up-regulated in GBM. 17-AAG is therefore a tumor-specific, replication-dependent radiosensitizer that enhances the effects of PARP inhibition. This combination has therapeutic potential in the management of GBM. [Mol Cancer Ther 2009;8(8):2243–54]
Drug ratio–dependent antitumor activity of irinotecan and cisplatin combinations in vitro and in vivo
Authors: Tardi Paul G. , Dos Santos Nancy , Harasym Troy O. , Johnstone Sharon A. , Zisman Natalia , Tsang Alan W. , Bermudes David G. , Mayer Lawrence D.
Published: August 2009

Irinotecan and cisplatin are two established anticancer drugs, which together constitute an effective combination for treating small-cell lung cancer. We investigated whether the efficacy of this combination could be improved by controlling drug ratios following in vivo administration. Irinotecan and cisplatin combinations were evaluated systematically for drug ratio–dependent synergy in vitro using a panel of 20 tumor cell lines. In vitro screening informatics on drug ratio–dependent cytotoxicity identified a consistently antagonistic region between irinotecan/cisplatin molar ratios of 1:2 to 4:1, which was bordered by two synergistic regions. Liposomal co-formulations of these two agents were developed that exhibited plasma drug half-lives of ∼6 hours and maintained a fixed drug ratio for more than 24 hours. Drug ratio–dependent antitumor activity was shown in vivo for these liposome formulations, and irinotecan/cisplatin ratios between 5:1 and 10:1 were identified as therapeutically optimal. The relationship between irinotecan/cisplatin ratio and in vivo efficacy was consistent with in vitro drug ratio dependency results. Superior antitumor activity was observed for the liposome-encapsulated 7:1 molar ratio of irinotecan/cisplatin (designated CPX-571) compared with the free-drug cocktail in all models tested. Further efficacy studies in a range of human tumor xenografts, including an irinotecan-resistant model, showed that both liposomal agents contributed to the overall efficacy in a manner consistent with in vivo synergy. These results show the ability of drug delivery technology to enhance the therapeutic activity of irinotecan/cisplatin combination treatment by maintaining synergistic ratios in vivo. CPX-571, a fixed-ratio formulation of irinotecan and cisplatin, is a promising candidate for clinical development. [Mol Cancer Ther 2009;8(8):2266–75]
Differential requirement for focal adhesion kinase signaling in cancer progression in the transgenic adenocarcinoma of mouse prostate model
Authors: Slack-Davis Jill K. , Hershey E. Daniel , Theodorescu Dan , Frierson Henry F. , Parsons J. Thomas
Published: August 2009

Increasing evidence indicates that adhesion signaling plays an important role in the tumor microenvironment, contributing to cancer progression, invasion, and metastasis. Focal adhesion kinase (FAK) is a nonreceptor protein tyrosine kinase that regulates adhesion-dependent cell signaling and has been implicated in mediating steps in cancer progression and metastasis in many human cancers, including prostate. We have investigated the role of FAK in the appearance of adenocarcinoma (atypical epithelial hyperplasia of T antigen) and neuroendocrine carcinomas in the transgenic adenocarcinoma of mouse prostate (TRAMP) model using either Cre-mediated recombination to genetically ablate FAK expression or pharmacologic inhibition of FAK activity with the small-molecule inhibitor, PF-562,271. We provide evidence that loss of FAK or its inhibition with PF-562,271 does not alter the progression to adenocarcinoma. However, continued FAK expression (and activity) is essential for the androgen-independent formation of neuroendocrine carcinoma. These data indicate that integrin signaling through FAK is an important component of cancer progression in the TRAMP model and suggest that treatment modalities targeting FAK may be an appropriate strategy for patients with castrate-resistant cancer. [Mol Cancer Ther 2009;8(8):2470–77]
Curcumin blocks prostaglandin E2 biosynthesis through direct inhibition of the microsomal prostaglandin E2 synthase-1
Authors: Koeberle Andreas , Northoff Hinnak , Werz Oliver
Published: August 2009

Prostaglandin E2 (PGE2) plays a crucial role in the apparent link between tumor growth and chronic inflammation. Cyclooxygenase (COX)-2 and microsomal PGE2 synthase-1, which are overexpressed in many cancers, are functionally coupled and thus produce massive PGE2 in various tumors. Curcumin, a polyphenolic β-diketone from tumeric with anti-carcinogenic and anti-inflammatory activities, was shown to suppress PGE2 formation and to block the expression of COX-2 and of microsomal PGE2 synthase-1. Here, we identified microsomal PGE2 synthase-1 as a molecular target of curcumin and we show that inhibition of microsomal PGE2 synthase-1 activity is the predominant mechanism of curcumin to suppress PGE2 biosynthesis. Curcumin reversibly inhibited the conversion of PGH2 to PGE2 by microsomal PGE2 synthase-1 in microsomes of interleukin-1β–stimulated A549 lung carcinoma cells with an IC50 of 0.2 to 0.3 μmol/L. Closely related polyphenols (e.g., resveratrol, coniferyl alcohol, eugenol, rosmarinic acid) failed in this respect, and isolated ovine COX-1 and human recombinant COX-2 were not inhibited by curcumin up to 30 μmol/L. In lipopolysaccharide-stimulated human whole blood, curcumin inhibited COX-2–derived PGE2 formation from endogenous or from exogenous arachidonic acid, whereas the concomitant formation of COX-2–mediated 6-keto PGF1α and COX-1–derived 12(S)-hydroxy-5-cis-8,10-trans-heptadecatrienoic acid was suppressed only at significant higher concentrations. Based on the key function of PGE2 in inflammation and carcinogenesis, inhibition of microsomal PGE2 synthase-1 by curcumin provides a molecular basis for its anticarcinogenic and anti-inflammatory activities. [Mol Cancer Ther 2009;8(8):2348–55]
An oncogenic isoform of HER2 associated with locally disseminated breast cancer and trastuzumab resistance
Authors: Mitra Doyel , Brumlik Michael J. , Okamgba Stella U. , Zhu Yun , Duplessis Tamika T. , Parvani Jenny G. , Lesko Samuel M. , Brogi Edi , Jones Frank E.
Published: August 2009

The HER2-targeted therapy trastuzumab is widely used for the treatment of patients with metastatic breast tumors overexpressing HER2. However, an objective response is observed in only 12% to 24% of patients treated with trastuzumab as a single agent and initial responders regress in <6 months (1–3). The reason for the clinical failure of trastuzumab in this setting remains unclear. Here we show that local lymph node–positive disease progression in 89% of breast cancer patients with HER2-positive tumors involves the HER2 oncogenic variant HER2Δ16. We further show that ectopic expression of HER2Δ16, but not wild-type HER2, promotes receptor dimerization, cell invasion, and trastuzumab resistance of NIH3T3 and MCF-7 tumor cell lines. The potentiated metastatic and oncogenic properties of HER2Δ16 were mediated through direct coupling of HER2Δ16 to Src kinase. Cotargeting of HER2Δ16 and Src kinase with the single-agent tyrosine kinase inhibitor dasatinib resulted in Src inactivation, destabilization of HER2Δ16, and suppressed tumorigenicity. Activated Src kinase was also observed in 44% of HER2Δ16-expressing breast carcinomas underscoring the potential clinical implications of coupled HER2Δ16 and Src signaling. Our results suggest that HER2Δ16 expression is an important genetic event driving trastuzumab-refractory breast cancer. We propose that successful targeted therapeutics for intervention of aggressive HER2-positive breast cancers will require a strategy to suppress HER2Δ16 oncogenic signaling. One possibility involves a therapeutic strategy employing single-agent tyrosine kinase inhibitors to disengage the functionally coupled oncogenic HER2Δ16 and Src tyrosine kinase pathways. [Mol Cancer Ther 2009;8(8):2152–62]
Microtubule inhibitors: Differentiating tubulin-inhibiting agents based on mechanisms of action, clinical activity, and resistance
Authors: Perez Edith A.
Published: August 2009

Microtubules are important cellular targets for anticancer therapy because of their key role in mitosis. Microtubule inhibitors (MTI) such as taxanes, vinca alkaloids, and epothilones stabilize or destabilize microtubules, thereby suppressing microtubule dynamics required for proper mitotic function, effectively blocking cell cycle progression and resulting in apoptosis. In spite of their antitumor activity, innate or acquired drug resistance to MTIs such as the taxanes is common, limiting their overall clinical efficacy. Further insight into the mechanisms of action of microtubule-targeting drugs has lead to the discovery of novel agents that may provide higher efficacy with limited toxicity and help overcome resistance to conventional MTIs. This review will focus on the different mechanisms of action of MTIs, potential factors related to resistance and tolerability, and will discuss the recent approval as well as the development of new antineoplastic agents. [Mol Cancer Ther 2009;8(8):2086–95]
BAX/BAK–Independent Mitoptosis during Cell Death Induced by Proteasome Inhibition?
Authors: Lomonosova Elena , Ryerse Jan , Chinnadurai G.
Published: August 2009

Proteasome inhibitors induce rapid death of cancer cells. We show that in epithelial cancer cells, such death is associated with dramatic and simultaneous up-regulation of several BH3-only proteins, including BIK, BIM, MCL-1S, NOXA, and PUMA, as well as p53. Elevated levels of these proteins seem to be the result of direct inhibition of their proteasomal degradation, induction of transcription, and active translation. Subsequent cell death is independent of BAX, and probably BAK, and proceeds through the intrinsic mitochondrial apoptosis pathway. We identify the cascade of molecular events responsible for cell death induced by a prototypical proteasome inhibitor, MG132, starting with rapid accumulation of BH3-only proteins in the mitochondria, proceeding through mitochondrial membrane permeabilization and subsequent loss of ΔΨm, and leading to irreversible changes of mitochondrial ultrastructure, degradation of mitochondrial network, and detrimental impairment of crucial mitochondrial functions. Our results also establish a rationale for the broader use of proteasome inhibitors to kill apoptosis-resistant tumor cells that lack functional BAX/BAK proteins. (Mol Cancer Res 2009;7(8):1268–84)
MDM2 Controls the Timely Expression of Cyclin A to Regulate the Cell Cycle
Authors: Frum Rebecca , Ramamoorthy Mahesh , Mohanraj Lathika , Deb Sumitra , Deb Swati Palit
Published: August 2009

Overexpression of MDM2 has been related to oncogenesis. In this communication, we present evidence to show that MDM2 controls the cell cycle–dependent expression of cyclin A by using a pathway that ensures its timely expression. MDM2 does not inhibit cyclin D or E expression. Silencing of endogenous MDM2 expression elevates cyclin A expression. The p53-binding domain of MDM2 harbors a SWIB region homologous to a conserved domain of a chromosome remodeling factor BRG1-associated protein. The SWIB domain of MDM2 inhibits cyclin A expression in a p53- and BRG1-dependent fashion, suggesting that MDM2 interferes with p53 binding of the BRG1 complex freeing it to repress cyclin A expression. Silencing of cyclin-dependent kinase (cdk) inhibitor p16 prevents MDM2-mediated inhibition of cyclin A expression, implicating its role in the process. MDM2-mediated repression of cyclin A expression induces G1-S arrest, which can be rescued by ectopic expression of cyclin A. Cancer cells lacking p53, p16, or BRG1 escape MDM2-mediated repression of cyclin A expression and growth arrest. Our data propose a novel mechanism by which MDM2 controls the cell cycle in normal cells and how cancer cells may escape this important safety barrier. (Mol Cancer Res 2009;7(8):1253–67)
Cadherin-Cadherin Engagement Promotes Cell Survival via Rac1/Cdc42 and Signal Transducer and Activator of Transcription-3
Authors: Arulanandam Rozanne , Vultur Adina , Cao Jun , Carefoot Esther , Elliott Bruce E. , Truesdell Peter F. , Larue Lionel , Feracci Hélène , Raptis Leda
Published: August 2009

Signal transducer and activator of transcription-3 (Stat3) is activated by a number of receptor and nonreceptor tyrosine kinases, whereas a constitutively active form of Stat3 alone is sufficient to induce neoplastic transformation. In the present report, we show that Stat3 can also be activated through homophilic interactions by the epithelial (E)-cadherin. Indeed, by plating cells onto surfaces coated with fragments encompassing the two outermost domains of this cadherin, we clearly show that cadherin engagement can activate Stat3, even in the absence of direct cell-to-cell contact. Most importantly, our results also reveal for the first time an unexpected and dramatic surge in total Rac1 and Cdc42 protein levels triggered by cadherin engagement and an increase in Rac1 and Cdc42 activity, which is responsible for the Stat3 stimulation observed. Inhibition of cadherin interactions using a peptide, a soluble cadherin fragment, or genetic ablation induced apoptosis, points to a significant role of this pathway in cell survival signaling, a finding that could also have important therapeutic implications. (Mol Cancer Res 2009;7(8):1310–27)
Small Interfering RNA–Directed Knockdown of Uracil DNA Glycosylase Induces Apoptosis and Sensitizes Human Prostate Cancer Cells to Genotoxic Stress
Authors: Pulukuri Sai Murali Krishna , Knost James A. , Estes Norman , Rao Jasti S.
Published: August 2009

Uracil DNA glycosylase (UNG) is the primary enzyme responsible for removing uracil residues from DNA. Although a substantial body of evidence suggests that DNA damage plays a role in cancer cell apoptosis, the underlying mechanisms are poorly understood. In particular, very little is known about the role of base excision repair of misincorporated uracil in cell survival. To test the hypothesis that the repair of DNA damage associated with uracil misincorporation is critical for cancer cell survival, we used small interfering RNA (siRNA) to target the human UNG gene. In a dose-dependent and time-dependent manner, siRNA specifically inhibited UNG expression and modified the expression of several genes at both mRNA and protein levels. In LNCaP cells, p53, p21, and Bax protein levels increased, whereas Bcl2 levels decreased. In DU145 cells, p21 levels were elevated, although mutant p53 and Bax levels remained unchanged. In PC3 cells, UNG inhibition resulted in elevated p21 and Bax levels. In all three cell lines, UNG inhibition reduced cell proliferation, induced apoptosis, and increased cellular sensitivity to genotoxic stress. Furthermore, an in vitro cleavage experiment using uracil-containing double-stranded DNA as a template has shown that siRNA-mediated knockdown of UNG expression significantly reduced the uracil-excising activity of UNG in human prostate cancer cells, which was associated with DNA damage analyzed by comet assay. Taken together, these findings indicate that RNA interference–directed targeting of UNG is a convenient, novel tool for studying the biological role of UNG and raises the potential of its application for prostate cancer therapy. (Mol Cancer Res 2009;7(8):1285–93)
FOXO Transcription Factors Enforce Cell Cycle Checkpoints and Promote Survival of Hematopoietic Cells after DNA Damage
Authors: Lei Hong , Quelle Frederick W.
Published: August 2009

The PI3K/AKT signaling pathway contributes to cell cycle progression of cytokine-dependent hematopoietic cells under normal conditions, and it is absolutely required to override DNA damage–induced cell cycle arrest checkpoints in these cells. Phosphatidylinositol-3-kinase (PI3K)/AKT activity also correlates with Cdk2 activity in hematopoietic cells, suggesting that Cdk2 activation may be a relevant end point for this signaling pathway. However, mediators downstream of AKT in this pathway have not been defined. The forkhead transcription factor O (FOXO) family are negatively regulated by AKT-dependent phosphorylation and are known regulators of genes affecting cell cycle progression. We show that enhanced FOXO activity replicates the effect of PI3K inhibitors in enforcing G1 and G2 phase arrest after DNA damage. Conversely, knockdown of endogenous FOXO proteins increased Cdk2 activity and overrode DNA damage checkpoints in cells lacking PI3K activity. Moreover, loss of FOXO activity caused an increase in sensitivity to cisplatin-induced cell death, which was associated with failure to arrest cell cycle progression in the face of DNA damage caused by this chemotherapeutic agent. These cell cycle arrests were dependent on p27 expression when mediated by FOXO3a alone, but also involve p27-independent mechanisms when promoted by endogenous FOXO proteins. Together, these observations show that FOXO proteins enforce DNA damage–induced cell cycle arrest in hematopoietic cells. Inhibition of FOXO activity by cytokine-induced PI3K/AKT signaling is sufficient to override these DNA damage–induced cell cycle checkpoints, but may negatively impact hematopoietic cell viability. (Mol Cancer Res 2009;7(8):1294–303)
NGFI-B Nuclear Orphan Receptor Nurr1 Interacts with p53 and Suppresses Its Transcriptional Activity
Authors: Zhang Tao , Wang Pingping , Ren Haigang , Fan Jun , Wang Guanghui
Published: August 2009

Nurr1 is a member of the NGFI-B nuclear orphan receptor family which includes two other members, Nur77 and Nor-1. Nurr1 is essential for the development and survival of dopaminergic neurons. It was reported that Nurr1 has antiapoptotic functions, however, the mechanisms by which Nurr1 mediates these effects remain unknown. Here, we show that overexpression of Nurr1 decreases Bax expression whereas knockdown of Nurr1 increases Bax expression. Nurr1 also interacts with p53 and represses its assembly. Furthermore, Nurr1 represses p53 transcriptional activity in interaction-dependent and dose-dependent manners. Moreover, Nurr1 protects cells from doxorubicin-induced apoptosis. These findings provide evidence that Nurr1 promotes cell survival through its interacting with and repressing p53, thus implicating that Nurr1 may play an important role in carcinogenesis and other diseases. (Mol Cancer Res 2009;7(8):1408–15)
A Novel Function of CXCL13 to Stimulate RANK Ligand Expression in Oral Squamous Cell Carcinoma Cells
Authors: Yuvaraj Sambandam , Griffin Alfred C. , Sundaram Kumaran , Kirkwood Keith L. , Norris James S. , Reddy Sakamuri V.
Published: August 2009

Oral squamous cell carcinomas (OSCC) are malignant tumors with a potent activity of local bone invasion/osteolysis. The chemokine ligand, CXCL13, has been identified as a prognostic marker for OSCC development and progression. Here in, we show that recombinant hCXCL13 treatment of OSCC cells stimulates (5-fold) RANK ligand (RANKL), a critical bone resorbing osteoclastogenic factor expression. Anti-CXCR5 chemokine receptor antibody abrogates CXCL13-induced RANKL expression in these cells. Also, CXCL13 stimulated (3.0-fold) hRANKL gene promoter activity in SCC14a cells. SuperArray screening for transcription factors by real-time RT-PCR identified significant increase in the levels of c-Jun and NFATc3 mRNA expression in CXCL13-stimulated OSCC cells. CXCL13 treatment significantly increased (3.5-fold) phospho-c-Jun levels in these cells and a c-Jun-NH2-kinase inhibitor abolished CXCL13-stimulated RANKL expression. Furthermore, we show that CXCL13 stimulation induced nuclear translocation of NFATc3 in OSCC cells. Chromatin-immune precipitation assay confirmed NFATc3 binding to the RANKL promoter region. We also show that overexpression of NFATc3 stimulates RANKL expression/promoter activity and that siRNA suppression of NFATc3 abolished CXCL13-stimulated RANKL expression. Thus, our results suggest that NFATc3 is a downstream target of the CXCL13/CXCR5 axis to stimulate RANKL expression in OSCC cells and implicates CXCL13 as a potential therapeutic target to prevent OSCC bone invasion/osteolysis. (Mol Cancer Res 2009;7(8):1399–407)
Cyclooxygenase-2 (COX-2) Negatively Regulates Expression of Epidermal Growth Factor Receptor and Causes Resistance to Gefitinib in COX-2–Overexpressing Cancer Cells
Authors: Kim Young Mee , Park Soo-Yeon , Pyo Hongryull
Published: August 2009

Overexpression of cyclooxygenase-2 (COX-2) and epidermal growth factor receptor (EGFR) has been detected in many types of cancer. Although COX-2 and EGFR are closely related to each other, the exact mechanism of COX-2 in tumors has not been well understood. In this study, we investigated the relationship between COX-2 and EGFR in cancer cells. Using two cell lines stably overexpressing COX-2 (HCT-116-COX-2 and H460-COX-2) and a stable line of COX-2 knockdown MOR-P cells, we analyzed patterns of COX-2 and EGFR expression. To observe the effects of COX-2 on EGFR expression and activity, we did comparative analyses after treatment with various drugs (EGF, celecoxib, prostaglandin E2, gefitinib, Ro-31-8425, PD98059, and SP600125) in HCT-116-Mock versus HCT-116-COX-2 cells and H460-Mock versus H460-COX-2 cells. Overexpression of COX-2 specifically down-regulated EGFR expression at the level of transcription. COX-2–overexpressing cells have a decreased sensitivity to gefitinib. COX-2 induced activation of extracellular signal-regulated kinase (ERK) and c-Jun NH2-terminal kinase (JNK) but suppressed Akt activation. JNK inhibition by SP600125, a specific JNK inhibitor, resulted in restoration of EGFR levels in COX-2–overexpressing cells, whereas ERK inhibition by PD98059 did not. Overexpressed COX-2 negatively regulates EGFR expression via JNK activation, leading to gefitinib resistance. COX-2 may also regulate ERK activity independently of EGFR. Therefore, resistance of COX-2–overexpressing cells to gefitinib may be due to decreased expression of EGFR by JNK activation and EGFR-independent elevation of ERK activity by COX-2. The ability of COX-2 to inhibit EGFR expression and gefitinib effects may have significance in clinical cancer therapy. (Mol Cancer Res 2009;7(8):1367–77)
Cathepsin G–Mediated Activation of Pro–Matrix Metalloproteinase 9 at the Tumor-Bone Interface Promotes Transforming Growth Factor-β Signaling and Bone Destruction
Authors: Wilson Thomas J. , Nannuru Kalyan C. , Singh Rakesh K.
Published: August 2009

Increased transforming growth factor-β (TGF-β) signaling has been observed at the tumor-bone interface of mammary tumor–induced osteolytic lesions despite no observed transcriptional up-regulation of TGF-β. To this point, the mechanism for enhanced TGF-β signaling remains unclear. The bulk of TGF-β that is released at the tumor-bone interface is in an inactive form secondary to association with β-latency–associated protein and latency TGF-β binding protein. We hypothesized that the observed increase in TGF-β signaling is due to increased cathepsin G–dependent, matrix metalloproteinase 9 (MMP9)–mediated activation of latent TGF-β. MMP9 is capable of activating latent TGF-β, and we observed that decreased production of MMP9 was associated with reduced TGF-β signaling. Similar to TGF-β, MMP9 is released in an inactive form and requires proteolytic activation. We showed that cathepsin G, which we have previously shown to be up-regulated at the tumor-bone interface, is capable of activating pro-MMP9. Inhibition of cathepsin G in vivo significantly reduced MMP9 activity, increased the ratio of latent TGF-β to active TGF-β, and reduced the level of TGF-β signaling. Our proposed model based on these results is that cathepsin G is up-regulated through tumor-stromal interactions and activates pro-MMP9, active MMP9 cleaves and releases active TGF-β, and active TGF-β can then promote tumor growth and enhance osteoclast activation and subsequent bone resorption. Thus, for the first time, we have identified cathepsin G and MMP9 as proteases involved in enhanced TGF-β signaling at the tumor-bone interface of mammary tumor–induced osteolytic lesions and have identified these proteases as potential therapeutic targets. (Mol Cancer Res 2009;7(8):1224–33)
Utility of DNA Repair Protein Foci for the Detection of Putative BRCA1 Pathway Defects in Breast Cancer Biopsies
Authors: Willers Henning , Taghian Alphonse G. , Luo Chen-Mei , Treszezamsky Alejandro , Sgroi Dennis C. , Powell Simon N.
Published: August 2009

The DNA damage response pathway controlled by the breast cancer and Fanconi anemia (FA) genes can be disrupted by genetic or epigenetic mechanisms in breast cancer. Defects in this pathway may render the affected tumors hypersensitive to DNA-damaging agents. The identification of these defects poses a challenge because of the large number of genes involved in the FA/BRCA pathway. Many pathway components form subnuclear repair protein foci upon exposure to ionizing radiation in vitro, but it was unknown whether foci can be detected in live cancer tissues. Thus, the goal of this pilot study was to identify pathway defects by using a novel ex vivo foci biomarker assay on tumor biopsies. Fresh pretreatment biopsy specimens from patients with locally advanced sporadic breast cancer were irradiated or mock-treated in the laboratory (ex vivo). Foci formation of DNA repair proteins BRCA1, FANCD2, and RAD51 was detected by immunofluorescence microscopy. Three out of seven tumors showed intact radiation-induced foci formation, whereas the other four tumors exhibited a defective foci response. Notably, three of the foci-defective tumors were estrogen receptor/progesterone receptor/HER2–negative (triple-negative), a phenotype that has been associated with BRCA1 deficiency. In conclusion, in this pilot study, we report the successful detection of BRCA1, FANCD2, and RAD51 foci in breast cancer biopsies irradiated ex vivo. Our approach represents a potentially powerful biomarker assay for the detection of pre-existing and functionally important defects within the complex FA/BRCA pathway, which may ultimately allow us to tailor cancer treatment to the DNA repair profile of individual tumors. (Mol Cancer Res 2009;7(8):1304–9)
Detection and Typing of Human Papillomavirus Nucleic Acids in Biological Fluids
Authors: Coutlée François , Mayrand Marie-Hélène , Roger Michel , Franco Eduardo L.
Published: August 2009

Human papillomaviruses (HPV) are the etiologic agents of cancer of the uterine cervix and several other neoplasias. Detection of HPV infection will improve the sensitivity of primary and secondary screening of cervical cancer. The clinical indications for the use of HPV tests will have to consider the natural history of HPV infection and diseases, and the multiplicity of types involved. Signal amplification HPV DNA tests detect several high-risk HPV types, are standardized, commercially available and approved for clinical use. Nucleic acid amplification techniques are ideal methods for epidemiologic purposes since they minimize misclassification of HPV infection status and allow detection of infection with low viral burden. They are currently under evaluation for clinical use. PCR is the most widespread method for HPV typing, especially with the use of consensus primers and typing with reverse hybridization techniques. Novel promising HPV detection strategies are now proposed, such as HPV mRNA detection, and suspension or solid phase arrays. These novel techniques will have to be evaluated as stringently as actual assays in clinical studies. Although assays have been developed for the evaluation of viral load, viral integration and HPV polymorphism in molecular epidemiological studies, their role in clinical practice is not currently defined.
Economic Evaluation of Human Papillomavirus Vaccination in Developed Countries
Authors: Brisson Marc , van de Velde Nicolas , Boily Marie-Claude
Published: August 2009

Background: With promising efficacy results from randomized control trials of human papillomavirus (HPV) vaccines and the availability of new screening paradigms, policymakers are being asked to make recommendations and decisions regarding the optimal strategies to reduce HPV infection and disease. Such decisions are increasingly being made with significant input from mathematical and economic models. The demand for modeling has resulted in the publication of numerous mathematical models looking at the cost-effectiveness of HPV vaccination. Objective: To review published models that have been used to evaluate the cost-effectiveness of HPV vaccination in developed countries and highlight points of consensus and disagreement in methods and findings. Methods: This review consists of cost-effectiveness studies published in the peer-reviewed literature before August 2008. Results: Despite variations in methods, modeling studies are producing consistent conclusions: (1) vaccinating young girls against HPV is likely to be cost- effective; (2) vaccinating boys will most likely not be cost- effective in countries that can reach high coverage rates in girls, and (3) results are most sensitive to the duration of vaccine protection. However, results from analyses examining the effectiveness and cost-effectiveness of vaccinating boys when coverage rates are low (≤80%) and catch-up strategies have reached conflicting conclusions.
The Case for Vaccinating Boys against Human Papillomavirus
Authors: Hull Sarah C. , Caplan Arthur L.
Published: August 2009

Vaccination policy in the case of human papillomavirus (HPV) has remained a constant source of controversy ever since Gardasil, Merck’s vaccine against HPV, received US Food and Drug Administration approval in the summer of 2006. This controversy has centered on the risks and benefits of vaccinating girls and women in rich and poor nations alike. However, despite all of the attention created by this important policy question, relatively little has been focused on another key public health question: should boys be vaccinated against HPV as well? If herd immunity against the most carcinogenic strains of HPV could be more rapidly and efficiently achieved by vaccinating everyone at risk for being a carrier, it logically follows that vaccine policy should expand to include boys and men.
From Causality to Prevention – The Example of Cervical Cancer: My Personal Contribution to This Fascinating History
Authors: Muñoz Nubia
Published: August 2009

The history of the generation of an hypothesis linking human papillomavirus to cervical cancer, as well as a summary of the various molecular epidemiological studies undertaken to confirm such hypothesis are described.
Evolutionary Diversity and Turn-Over of Sex Determination in Teleost Fishes
Authors: Mank J.E. , Avise J.C.
Published: August 2009

Sex determination, due to the obvious association with reproduction and Darwinian fitness, has been traditionally assumed to be a relatively conserved trait. However, research on teleost fishes has shown that this need not be the case, as these animals display a remarkable diversity in the ways that they determine sex. These different mechanisms, which include constitutive genetic mechanisms on sex chromosomes, polygenic constitutive mechanisms, environmental influences, hermaphroditism, and unisexuality have each originated numerous independent times in the teleosts. The evolutionary lability of sex determination, and the corresponding rapid rate of turn-over among different modes, makes the teleost clade an excellent model with which to test theories regarding the evolution of sex determining adaptations. Much of the plasticity in sex determination likely results from the dynamic teleost genome, and recent advances in fish genetics and genomics have revealed the role of gene and genome duplication in fostering emergence and turn-over of sex determining mechanisms.
Sex Determination and Sex Chromosome Evolution: Insights from Medaka
Authors: Kondo M. , Nanda I. , Schmid M. , Schartl M.
Published: August 2009

Among fish that exhibit a variety of sex-determining systems, medaka (Oryzias latipes) has been used as an ideal model to study sex determination and differentiation. Medaka sex differentiation starts with germ cell migration and proliferation during development, and the earliest sexual difference can be seen in germ cell number at stage 38 (1 day before hatching). Differentiation continues in young larvae. Medaka has an XX-XY genetic system, and the male sex-determining gene dmrt1bY (dmy) has been identified. Along with a sister species O. curvinotus, these species are the only 2 fish species whose sex-determining gene has been identified so far. The medaka sex chromosome is of recent origin, and the Y chromosome was formed through generation of the Y-specific region by duplication and insertion of an autosomal region into the proto-Y chromosome. Thus, the dmrt1bY gene is a duplicate of the autosomal dmrt1a gene. This event is estimated to have occurred about 10 million years ago. Sex-determining systems of other Oryzias species are under investigation, and a variety of sex-determining systems seem to exist. These findings may provide new insights into the sex-determining system of medaka.
Comparative Aspects of Gonadal Sex Differentiation in Medaka: A Conserved Role of Developing Oocytes in Sexual Canalization
Authors: Saito D. , Tanaka M.
Published: August 2009

Medaka is a differentiated gonochoristic species with a male heterogametic sex determination. Here, we review recent studies on gonadal sex differentiation in medaka, as well as the experimental sex-reversal models available for this organism. The accumulated literature on teleost gonadal development facilitates comparative interspecies studies. Among these, comparison between medaka and zebrafish is of special interest, because zebrafish is an undifferentiated gonochorist that, nevertheless, shares many similarities with medaka and is also used as a small fish model for developmental biology. Accordingly, here we focus on the comparative aspects of gonadal development in medaka and zebrafish. In medaka, oogenesis begins in female gonads, whereas oogenesis is suppressed and germ cells remain in an undifferentiated state in male gonads. In zebrafish, oogenesis begins in all individuals, regardless of their future sex, while actual sex differentiation begins later in gonadal development, which means that degeneration of immature oocytes occurs in presumptive males, while oogenesis proceeds to completion in presumptive females. Despite these apparent differences between medaka and zebrafish, the process of gonadal development comprises similar stages: early oogenesis, early aromatase expression, later oocyte development or loss, sexually dimorphic expression of somatic genes, and spermatogenesis. We propose the concept of canalization as a key to gaining a comprehensive understanding of gonadal sex differentiation. In this respect, the possible role of the male-determining gene DMY/dmrt1bY is also discussed.
Molecular Aspects of Gonadal Differentiation in a Teleost Fish, the Nile Tilapia
Authors: Kobayashi T. , Nagahama Y.
Published: August 2009

In contrast to many developmental processes, sex-determining mechanisms show no clear evolutionary conservation among phyla. However, recent studies indicate that some downstream products of sex determination genes are functionally similar in diverse species. To date, numerous conserved genes involving gonadal sex differentiation have been examined in the teleost fish Nile tilapia (Oreochromis niloticus). Morphogenesis during gonadal differentiation is also conserved, as is evident in the differentiation and development of parenchyma/medullary cells (testis cord) and follicles. Therefore, it is important to understand the mechanisms of gonadal sex differentiation from the perspective of the relationship between conserved gene expression cascades and morphogenesis during gonadal sex differentiation. This article reviews the expression profiles of male- and female-related genes involved in histogenesis during sex differentiation in tilapia and discusses gene function in gonadal sex differentiation, especially the role of endogenous estrogens for ovarian differentiation.
Environmental Effects on Fish Sex Determination and Differentiation
Authors: Baroiller J.F. , D’Cotta H. , Saillant E.
Published: August 2009

Environmental factors affect the sex ratio of many gonochoristic fish species. They can either determine sex or influence sex differentiation. Temperature is the most common environmental cue affecting sex but density, pH and hypoxia have also been shown to influence the sex ratio of fish species from very divergent orders. Differential growth or developmental rate is suggested to influence sex differentiation in sea bass. Studies in most fish species used domestic strains reared under controlled conditions. In tilapia and sea bass, domestic stocks and field-collected populations showed similar patterns of thermosensitivity under controlled conditions. Genetic variability of thermosensitivity is seen between populations but also between families within the same population. Furthermore, in the Nile tilapia progeny testing of wild male breeders has strongly suggested the existence of XX males in 2 different natural populations. Tilapia and Atlantic silverside studies have shown that temperature sensitivity is a heritable trait which can respond to directional (tilapia) or frequency dependent selection. In tilapia, transitional forms within a genetic sex determination (GSD) and environmental sex determination (ESD) continuum seem to exist. Temperature regulates the expression of the ovarian-aromatase cyp19a1 which is consistently inhibited in temperature masculinized gonads. Foxl2 issuppressed before cyp19a1. Recent in vitro studies have shown that foxl2 activates cyp19a1, suggesting that temperature acts directly on foxl2 or further upstream. Dmrt1 up-regulation is correlated with temperature-induced male phenotypes. Temperature through apoptosis or germ cell proliferation could be a critical threshold for male or female sex differentiation.
Evolutionary Perspectives on Hermaphroditism in Fishes
Authors: Avise J.C. , Mank J.E.
Published: August 2009

Hermaphroditism is a derived and polyphyletic condition in fishes, documented in about 2% of all extant teleost species scattered across more than 20 taxonomic families in 9 orders. It shows a variety of expressions that can be categorized into sequential and synchronous modes. Among the sequential hermaphrodites are protogynous species in which an individual begins reproductive life as a female and later may switch to male, protandrous species in which a fish starts as a male and later may switch to female, and serial bi-directional sex changers. Among the synchronous hermaphrodites (in which an individual can simultaneously produce eggs and sperm) are several outcrossing and one predominantly selfing species. A few species also consist of mixtures of hermaphroditic and single-sex individuals. All of these reproductive categories have been the subject of numerous theoretical and empirical treatments from an evolutionary perspective. Here we highlight some of the major conclusions from these studies, which collectively have been informative on a variety of biological topics related to reproductive modes, gender allocations, sexual conflict and gamesmanship, mating systems, and life-history tradeoffs.
Sexual Development in Fish, Practical Applications for Aquaculture
Authors: Cnaani A. , Levavi-Sivan B.
Published: August 2009

Aquaculture is one of the fastest rising sectors of world food production. Hundreds of fish species are cultured, providing an affordable, high quality food source. Two aspects of sexual development are critically important for the continued improvement of cultured fish stocks: sexual dimorphism and control of reproduction. In this paper, we review the main methods used to control sex determination in fish and their application in some of the most widely cultured species. Specifically, we review the techniques available for the production of all-male, all-female, and sterile populations. Techniques for endocrinological control of reproduction are also discussed.
Does Inflammation after Stroke Affect the Developing Brain Differently than Adult Brain?
Authors: Vexler Zinaida S. , Yenari Midori A.
Published: August 2009

The immature brain is prone to hypoxic-ischemic encephalopathy and stroke. The incidence of arterial stroke in newborns is similar to that in the elderly. However, the pathogenesis of ischemic brain injury is profoundly affected by age at the time of the insult. Necrosis is a dominant type of neuronal cell death in adult brain, whereas widespread neuronal apoptosis is unique for the early postnatal synaptogenesis period. The inflammatory response, in conjunction with excitotoxic and oxidative responses, is the major contributor to ischemic injury in both the immature and adult brain, but there are several areas where these responses diverge. We discuss the contribution of various inflammatory mechanisms to injury and repair after cerebral ischemia in the context of CNS immaturity. In particular, we discuss the role of lower expression of selectins, a more limited leukocyte transmigration, undeveloped complement pathways, a more rapid microglial activation, differences in cytokine and chemokine interplay, and a different threshold to oxidative stress in the immature brain. We also discuss differences in activation of intracellular pathways, especially nuclear factor κB and mitogen-activated protein kinases. Finally, we discuss emerging data on both the supportive and adverse roles of inflammation in plasticity and repair after stroke.
Erythropoietin Attenuates Hyperoxia-Induced Cell Death by Modulation of Inflammatory Mediators and Matrix Metalloproteinases
Authors: Sifringer Marco , Genz Kerstin , Brait Daniela , Brehmer Felix , Löber Rebekka , Weichelt Ulrike , Kaindl Angela M. , Gerstner Bettina , Felderhoff-Mueser Ursula
Published: August 2009

Oxygen toxicity appears to contribute to the pathogenesis of adverse neurological outcome in survivors of preterm birth. In infant rodent brains, hyperoxia triggers widespread apoptotic neurodegeneration, induces proinflammatory cytokines and inhibits growth factor signaling cascades. Since a tissue-protective effect has been observed for recombinant erythropoietin (rEpo), we hypothesized that rEpo would influence the expression of proinflammatory cytokines and matrix metalloproteinase (MMP)-2 and MMP-9. Six-day-old Wistar rats were exposed to 80% oxygen for 2–48 h and received 20,000 IU rEpo i.p. Sex-matched littermates kept in room air and injected with normal saline or rEpo served as controls. Treatment with rEpo significantly reduced hyperoxia-induced upregulation of the proinflammatory cytokines IL-1β and IL-18 in infant rodent brains on the mRNA and protein levels. In parallel, gelatin zymography in hyperoxia-treated immature rat brains revealed an upregulation of active MMP-2 which was reduced by concomitant rEpo treatment. Furthermore, hyperoxia induced upregulation of MMP-9 following 12 h of oxygen exposure and this was attenuated by rEpo treatment. Our results suggest that rEpo generates its protective effect against oxygen toxicity through a reduction of proinflammatory mediator levels.
Neonatal Medial Frontal Cortex Lesions Disrupt Circadian Activity Patterns
Authors: Phillips Catherine I. , Smith Victoria M. , Antle Michael C. , Dyck Richard H.
Published: August 2009

The medial frontal cortex (MFC) is involved in the temporal organization of behaviour. It receives timing information from the master circadian clock in the suprachiasmatic nucleus (SCN), and exhibits daily oscillations in gene expression itself. In this study, we evaluate various properties of circadian rhythms of locomotor activity following neonatal or adult MFC aspiration lesions. Mice with neonatal lesions were more active during the day than mice with adult lesions and less active during the early night than both mice with adult lesions and control mice. Compared to controls, mice with neonatal lesions exhibited smaller phase delays to an early-night light pulse and marginally larger phase advances to a late-night light pulse. Mice with adult lesions did not differ from controls on either measure. The results suggest that the timing of behaviour is determined by an interaction between the MFC and the SCN and that injury early in life has a significant effect on the ability of animals to organize such behaviours.
Role of Mixed Lineage Kinase Inhibition in Neonatal Hypoxia-Ischemia
Authors: Carlsson Ylva , Leverin Anna-Lena , Hedtjärn Maj , Wang Xiaoyang , Mallard Carina , Hagberg Henrik
Published: August 2009

Hypoxic-ischemic brain injury is often delayed and involves both apoptotic and immunoregulatory mechanisms. In this study, we used a neonatal model of hypoxia-ischemia to examine the effect of the mixed lineage kinase (MLK) inhibitor CEP-1347 on brain damage, apoptosis and inflammation. The tissue volume loss was reduced by 28% (p = 0.019) in CEP-1347-treated versus vehicle-treated rats and CEP-1347 significantly attenuated microgliosis at 7 days (p = 0.038). CEP-1347 decreased TUNEL-positive staining as well as cleaved caspase 3 immunoreactivity. CEP-1347 did not affect the expression of pro-inflammatory cytokines IL-1β, IL-6 and MCP-1, nor did it affect the expression of OX-42 (CR3) and OX-18 (MHC I) 24 h after the insult. In conclusion, the MLK inhibitor CEP-1347 has protective effects in a neonatal rat model of hypoxia-ischemia, which is mainly related to reduced apoptosis.