bluuebunny/crossref_metadata_2025 huggingface dataset:
Transforming Growth Factor β Induces Clustering of HER2 and Integrins by Activating Src-Focal Adhesion Kinase and Receptor Association to the Cytoskeleton
Authors: Wang Shizhen Emily , Xiang Bin , Zent Roy , Quaranta Vito , Pozzi Ambra , Arteaga Carlos L.
Published: January 2009

It has been proposed that cross talk between integrin and growth factor receptor signaling such as ErbB2 (HER2) is required for activation of downstream effectors and ErbB2-mediated mammary tumorigenesis. Here we show that transforming growth factor β (TGF-β) induced focal adhesion kinase (FAK)–dependent clustering of HER2 and integrins α6, β1, and β4 in HER2-overexpressing mammary epithelial cells without altering the total and surface levels of HER2 receptors. This effect was mediated by ligand-induced epidermal growth factor receptor (EGFR) activation and the subsequent phosphorylation of Src and FAK. We have previously reported that TGF-β up-regulates EGFR ligand shedding through a mechanism involving the phosphorylation of tumor necrosis factor-α–converting enzyme (TACE/ADAM17). Knockdown of TACE, FAK, or integrin α6 by siRNA or inhibition of EGFR or Src by specific inhibitors abrogated TGF-β–induced receptor clustering and signaling to phosphatidylinositol 3-kinase-Akt. Finally, inhibition of Src-FAK reversed TGF-β–induced resistance to the therapeutic HER2 inhibitor trastuzumab in HER2-overexpressing breast cancer cells. Taken together, these data suggest that, by activating Src-FAK, TGF-β integrates ErbB receptor and integrin signaling to induce cell migration and survival during breast cancer progression. [Cancer Res 2009;69(2):475–82]
Pretreatment with Statin Attenuates the Cardiotoxicity of Doxorubicin in Mice
Authors: Riad Alexander , Bien Sandra , Westermann Dirk , Becher Peter M. , Loya Komal , Landmesser Ulf , Kroemer Heyo K. , Schultheiss Heinz P. , Tschöpe Carsten
Published: January 2009

Cardiotoxicity, which may result from intense cardiac oxidative stress and inflammation, is the main limiting factor of the anticancer therapy using doxorubicin. Because statins might exert beneficial pleiotropic cardiovascular effects, among other things, by anti-inflammatory and antioxidative mechanisms, we investigated whether or not fluvastatin pretreatment can attenuate doxorubicin-induced cardiotoxicity. Five days after a single injection of doxorubicin (20 mg/kg; i.p.), left ventricular (LV) function was measured in fluvastatin-treated (DoxStatin; 100 mg/kg/day, p.o.) and saline-treated (doxorubicin) mice (n = 8 per group) by a micro conductance catheter. Untreated mice served as controls (placebo; n = 8 per group). After measurement of cardiac function, LV tissues were analyzed by molecular biological and immunohistologic methods. Injection resulted in significantly impaired LV function (LV pressure, −29%; dp/dtmax, −45%; cardiac output, −68%; P < 0.05) when compared with placebo. This was associated with a significant increase in cardiac oxidative stress, inflammation and apoptotic mechanisms, as indicated by significant increased cardiac lipid peroxidation activity, protein expression of nitrotyrosine, tumor necrosis factor α and Bax (P < 0.05). In contrast, DoxStatin mice showed improved LV function (LV pressure, +24%; dp/dtmax, +87%; cardiac output, +87%; P < 0.05) when compared with untreated doxorubicin mice. This was associated with reduced cardiac expression of nitrotyrosine, enhanced expression of the mitochondrial located antioxidative SOD 2, attenuated mitochondrial apoptotic pathways, and reduced cardiac inflammatory response. Statin pretreatment attenuates doxorubicin-induced cardiotoxicity via antioxidative and anti-inflammatory effects. [Cancer Res 2009;69(2):695–9]
Mammary Epithelial-Specific Ablation of the Focal Adhesion Kinase Suppresses Mammary Tumorigenesis by Affecting Mammary Cancer Stem/Progenitor Cells
Authors: Luo Ming , Fan Huaping , Nagy Tamas , Wei Huijun , Wang Chenran , Liu Suling , Wicha Max S. , Guan Jun-Lin
Published: January 2009

Focal adhesion kinase (FAK) has been implicated in the development of cancers, including those of the breast. Nevertheless, the molecular and cellular mechanisms by which FAK promotes mammary tumorigenesis in vivo are not well understood. Here, we show that targeted deletion of FAK in mouse mammary epithelium significantly suppresses mammary tumorigenesis in a well-characterized breast cancer model. Ablation of FAK leads to the depletion of a subset of bipotent cells in the tumor that express both luminal marker keratin 8/18 and basal marker keratin 5. Using mammary stem/progenitor markers, including aldehyde dehydrogenase, CD24, CD29, and CD61, we further revealed that ablation of FAK reduced the pool of cancer stem/progenitor cells in primary tumors of FAK-targeted mice and impaired their self-renewal and migration in vitro. Finally, through transplantation in NOD-SCID mice, we found that cancer stem/progenitor cells isolated from FAK-targeted mice have compromised tumorigenicity and impaired maintenance in vivo. Together, these results show a novel function of FAK in maintaining the mammary cancer stem/progenitor cell population and provide a novel mechanism by which FAK may promote breast cancer development and progression. [Cancer Res 2009;69(2):466–74]
Combined Transductional Untargeting/Retargeting and Transcriptional Restriction Enhances Adenovirus Gene Targeting and Therapy for Hepatic Colorectal Cancer Tumors
Authors: Li Hua-Jung , Everts Maaike , Yamamoto Masato , Curiel David T. , Herschman Harvey R.
Published: January 2009

Unresectable hepatic colorectal cancer (CRC) metastases are a leading cause of cancer mortality. These tumors and other epithelial tumors often express both cyclooxygenase-2 (COX-2) and carcinoembryonic antigen (CEA). Because adenovirus (Ad) vectors infect the liver and lack tumor tropism, they cannot be used for systemic therapy of hepatic metastases. We used COX-2 transcriptional restriction, in combination with transductional Ad hepatic untargeting and tumor retargeting by a bispecific adapter, sCARhMFE, composed of sCAR [the coxsackie/Ad receptor (CAR) ectodomain] and MFE-23 (a single-chain anti-CEA antibody), to untarget liver after i.v. administration of Ad vectors expressing firefly luciferase and to retarget virus to hepatic colorectal tumor xenografts and non–small cell lung tumor xenografts. To improve both liver untargeting and tumor retargeting, we developed sCARfMFE, a trimerized sCARhMFE adapter. Trimerization greatly improves both untargeting of CAR-dependent Ad infection and CEA-dependent virus retargeting in culture and in vivo. Combining sCARfMFE bispecific adapter transductional liver untargeting and transductional tumor retargeting with COX-2 transcriptional tumor-restricted transgene expression increases systemically administered Ad therapeutic efficacy for hepatic CRC tumors, using herpes virus type 1 thymidine kinase (HSV1-tk) as a therapeutic gene in conjunction with the prodrug ganciclovir (GCV). Both transductional untargeting and COX-2 transcriptional restriction also reduce HSV1-tk/GCV hepatic toxicity. In addition, transductional sCARfMFE untargeting reduces the innate immune response to systemic Ad administration. Combined transductional liver Ad untargeting, transductional tumor retargeting, and transcriptional transgene restriction suggests a means to engineer practical, effective therapeutic agents for hepatic CRC metastases in particular, as well as hepatic metastases of other epithelial cancers. [Cancer Res 2009;69(2):554–64]
Host CXCR2-Dependent Regulation of Melanoma Growth, Angiogenesis, and Experimental Lung Metastasis
Authors: Singh Seema , Varney Michelle , Singh Rakesh K.
Published: January 2009

Crucial steps in tumor growth and metastasis are proliferation, survival, and neovascularization. Previously, we have shown that receptors for CXCL-8, CXCR1, and CXCR2 are expressed on endothelial cells and CXCR2 has been shown to be a putative receptor for angiogenic chemokines. In this report, we examined whether tumor angiogenesis and growth of CXCL-8–expressing human melanoma cells are regulated in vivo by a host CXCR2–dependent mechanism. We generated mCXCR2−/−, mCXCR2+/−, and wild-type nude mice following crosses between BALB/c mice heterozygous for nude+/− and heterozygous for mCXCR2+/−. We observed a significant inhibition of human melanoma tumor growth and experimental lung metastasis in mCXCR2−/− mice as compared with wild-type nude mice. Inhibition in tumor growth and metastasis was associated with a decrease in melanoma cell proliferation, survival, inflammatory response, and angiogenesis. Together, these studies show the importance of host CXCR2–dependent CXCL-8–mediated angiogenesis in the regulation of melanoma growth and metastasis. [Cancer Res 2009;69(2):411–5]
Genome-Wide Promoter Analysis of the SOX4 Transcriptional Network in Prostate Cancer Cells
Authors: Scharer Christopher D. , McCabe Colleen D. , Ali-Seyed Mohamed , Berger Michael F. , Bulyk Martha L. , Moreno Carlos S.
Published: January 2009

SOX4 is a critical developmental transcription factor in vertebrates and is required for precise differentiation and proliferation in multiple tissues. In addition, SOX4 is overexpressed in many human malignancies, but the exact role of SOX4 in cancer progression is not well understood. Here, we have identified the direct transcriptional targets of SOX4 using a combination of genome-wide localization chromatin immunoprecipitation–chip analysis and transient overexpression followed by expression profiling in a prostate cancer model cell line. We have also used protein-binding microarrays to derive a novel SOX4-specific position-weight matrix and determined that SOX4 binding sites are enriched in SOX4-bound promoter regions. Direct transcriptional targets of SOX4 include several key cellular regulators, such as EGFR, HSP70, Tenascin C, Frizzled-5, Patched-1, and Delta-like 1. We also show that SOX4 targets 23 transcription factors, such as MLL, FOXA1, ZNF281, and NKX3-1. In addition, SOX4 directly regulates expression of three components of the RNA-induced silencing complex, namely Dicer, Argonaute 1, and RNA Helicase A. These data provide new insights into how SOX4 affects developmental signaling pathways and how these changes may influence cancer progression via regulation of gene networks involved in microRNA processing, transcriptional regulation, the TGFβ, Wnt, Hedgehog, and Notch pathways, growth factor signaling, and tumor metastasis. [Cancer Res 2009;69(2):709–17]
Unlike Esophageal Squamous Cells, Barrett's Epithelial Cells Resist Apoptosis by Activating the Nuclear Factor-κB Pathway
Authors: Hormi-Carver Kathy , Zhang Xi , Zhang Hui Ying , Whitehead Robert H. , Terada Lance S. , Spechler Stuart J. , Souza Rhonda F.
Published: January 2009

Apoptosis is an important mechanism for maintaining tissue homeostasis and for preventing the proliferation of cells with mutations that could result in malignancy. Barrett's epithelium has been reported to be more resistant to apoptosis than normal esophageal squamous epithelium. We have explored the contribution of the nuclear factor-κB (NF-κB) pathway to apoptotic resistance in non-neoplastic, telomerase-immortalized esophageal squamous (NES) and Barrett's (BAR-T) epithelial cell lines. We exposed these cells to UV-B irradiation in doses known to cause DNA damage and to induce apoptosis in normal cells, and studied apoptosis as well as the expression of phospho-H2AX, NF-κB, Bcl-2, XIAP, cIAP-1, and survivin proteins. We also used Bay 11-7085 and siRNAs to NF-κB and Bcl-2 to assess the effects of NF-κB and Bcl2 inhibition on apoptosis. UV-B irradiation at low doses (50 and 100 J/m2) caused DNA damage in both NES and BAR-T cells but significantly increased apoptosis only in NES cells. UV-B irradiation caused a decrease in the levels of NF-κB, Bcl-2, cIAP-1, XIAP, and survivin in NES cells but increased the levels of those proteins in BAR-T cells. The resistance of BAR-T cells to apoptosis induced by low-dose UV-B irradiation was abolished by Bay 11-7085 and by siRNA for NF-κB and was decreased significantly by siRNA for Bcl-2. We conclude that the ability of Barrett's epithelial cells to activate the NF-κB pathway when they have sustained DNA damage allows them to resist apoptosis. This capacity to avoid apoptosis despite genotoxic damage may underlie the persistence and malignant predisposition of Barrett's metaplasia. [Cancer Res 2009;69(2):672–7]
On the Role of Cell Signaling Models in Cancer Research
Authors: Ventura Alejandra C. , Jackson Trachette L. , Merajver Sofia D.
Published: January 2009

The main objective of this review is to emphasize the role and importance of the careful mathematical/computational modeling of signaling networks for the understanding of aberrant signaling in cancer and for the development of targeted therapies. [Cancer Res 2009;69(2):400–2]
Prospective Internal Validation of Mathematical Models to Predict Malignancy in Adnexal Masses: Results from the International Ovarian Tumor Analysis Study
Authors: Van Holsbeke Caroline , Van Calster Ben , Testa Antonia C. , Domali Ekaterini , Lu Chuan , Van Huffel Sabine , Valentin Lil , Timmerman Dirk
Published: January 2009

Purpose: To prospectively test the mathematical models for calculation of the risk of malignancy in adnexal masses that were developed on the International Ovarian Tumor Analysis (IOTA) phase 1 data set on a new data set and to compare their performance with that of pattern recognition, our standard method.

Methods: Three IOTA centers included 507 new patients who all underwent a transvaginal ultrasound using the standardized IOTA protocol. The outcome measure was the histologic classification of excised tissue. The diagnostic performance of 11 mathematical models that had been developed on the phase 1 data set and of pattern recognition was expressed as area under the receiver operating characteristic curve (AUC) and as sensitivity and specificity when using the cutoffs recommended in the studies where the models had been created. For pattern recognition, an AUC was made based on level of diagnostic confidence.

Results: All IOTA models performed very well and quite similarly, with sensitivity and specificity ranging between 92% and 96% and 74% and 84%, respectively, and AUCs between 0.945 and 0.950. A least squares support vector machine with linear kernel and a logistic regression model had the largest AUCs. For pattern recognition, the AUC was 0.963, sensitivity was 90.2%, and specificity was 92.9%.

Conclusion: This internal validation of mathematical models to estimate the malignancy risk in adnexal tumors shows that the IOTA models had a diagnostic performance similar to that in the original data set. Pattern recognition used by an expert sonologist remains the best method, although the difference in performance between the best mathematical model is not large.
Safety, Pharmacokinetics, and Efficacy of CPX-1 Liposome Injection in Patients with Advanced Solid Tumors
Authors: Batist Gerald , Gelmon Karen A. , Chi Kim N. , Miller Wilson H. , Chia Stephen K.L. , Mayer Lawrence D. , Swenson Christine E. , Janoff Andrew S. , Louie Arthur C.
Published: January 2009

Purpose: CPX-1 is a novel, liposome-encapsulated formulation of irinotecan and floxuridine designed to prolong in vitro optimized synergistic molar ratios of both drugs postinfusion. This open-label, single-arm, dose-escalating phase I study was designed to determine the maximum tolerated dose and pharmacokinetics of CPX-1 in patients with advanced solid tumors.

Experimental Design: Patients received CPX-1 at 30, 60, 100, 150, 210, or 270 units/m2 (1 unit = 1 mg irinotecan + 0.36 mg floxuridine) infused over 90 minutes every 14 days in 28-day cycles. Pharmacokinetic samples were collected on days 1 and 15 of cycle 1.

Results: Thirty-three patients were enrolled, treated, and evaluated for safety; 30 patients were evaluated for response. A 1:1 plasma irinotecan to floxuridine molar ratio was maintained for 8 to 12 hours. Grade 3/4 toxicities included diarrhea (24.2%), neutropenia (12.1%), and hypokalemia (12.1%); 1 patient (270 units/m2) died of persistent diarrhea, which led to dehydration and renal failure (grade 5). Partial response occurred in 3 (12%) of the 25 subjects evaluated through Response Evaluation Criteria in Solid Tumors. Progression-free survival lasting >6 months occurred in 9 patients, 6 with colorectal cancer. Among 15 colorectal cancer patients (10 with prior irinotecan), the calculated median progression-free survival was 5.4 months; 11 patients (72.7%) achieved disease control and 2 patients (13%) had partial response.

Conclusions: Outpatient CPX-1 was well tolerated and antitumor activity was shown in patients with advanced solid tumors. The recommended dose for future studies is 210 units/m2. This is the first clinical evaluation of fixed drug ratio dosing designed to maintain synergistic molar ratios for enhanced therapeutic benefit.
Polyphenon E Inhibits the Growth of Human Barrett's and Aerodigestive Adenocarcinoma Cells by Suppressing Cyclin D1 Expression
Authors: Song Shumei , Krishnan Koyamangalath , Liu Kaifeng , Bresalier Robert S.
Published: January 2009

Purpose: Green tea consumption has been shown to exhibit cancer-preventive activities in preclinical studies. Polyphenon E (Poly E) is a well-defined green tea–derived catechin mixture. This study was designed to determine the effects of Poly E on the growth of human Barrett's and aerodigestive adenocarcinoma cells and the mechanisms involved in growth regulation by this agent.

Experimental Design: Human adenocarcinoma cells and immortalized Barrett's epithelial cells were used as model systems.

Results: Poly E inhibited the proliferation of immortalized Barrett's cells as well as various adenocarcinoma cells, and this was associated with the down-regulation of cyclin D1 protein expression. Inhibition of cyclin D1 led to dephosphorylation of the retinoblastoma protein in a dose-dependent manner; these changes were associated with G1 cell cycle arrest. Poly E down-regulated cyclin D1 promoter activity and mRNA expression, suggesting transcriptional repression, and this correlated with decreased nuclear β-catenin and β-catenin/TCF4 transcriptional activity. MG132, an inhibitor of 26S proteosome, blocked the Poly E–induced down-regulation of cyclin D1, and Poly E promoted cyclin D1 polyubiquitination, suggesting that Poly E also inhibits cyclin D1 expression by promoting its degradation.

Conclusion: Poly E inhibits growth of transformed aerodigestive epithelial cells by suppressing cyclin D1 expression through both transcriptional and posttranslational mechanisms. These results provide insight into the mechanisms by which Poly E inhibits growth of Barrett's and adenocarcinoma cells, and provides a rationale for using this agent as a potential chemopreventive and therapeutic strategy for esophageal adenocarcinoma and its precursor, Barrett's esophagus.
Inhibition of mTOR Radiosensitizes Soft Tissue Sarcoma and Tumor Vasculature
Authors: Murphy James D. , Spalding Aaron C. , Somnay Yash R. , Markwart Sonja , Ray Michael E. , Hamstra Daniel A.
Published: January 2009

Purpose: The PI3K/Akt/mTOR prosurvival pathway is frequently up-regulated in soft tissue sarcoma. Mammalian target of rapamycin (mTOR) inhibitors, such as rapamycin, have recently shown clinical benefit in soft tissue sarcoma, and mTOR inhibition has also been associated with radiosensitization of carcinoma and endothelial cells. This study tested the hypothesis that rapamycin radiosensitizes soft tissue sarcoma and endothelial cells in vitro and in vivo through the inhibition of mTOR.

Experimental Design: Colony formation assays were done to determine the radiosensitizing properties of rapamycin on three human soft tissue sarcoma cell lines (SK-LMS-1, SW-872, and HT-1080) and human dermal microvascular endothelial cells (HDMEC). The functional effects of rapamycin and radiation on the endothelial compartment were evaluated with microvascular sprouting assays. The in vivo radiosensitizing activity of rapamycin was assessed with s.c. SK-LMS-1 nude mice xenografts treated with concurrent daily rapamycin, radiation, or both for three weeks.

Results: In vitro radiosensitization was shown in all three soft tissue sarcoma cell lines with minimally cytotoxic doses of rapamycin. SK-LMS-1 xenografts displayed significant tumor growth delay with rapamycin and radiation compared with either treatment alone. Radiation resulted in transient increased mTOR function, whereas rapamycin abolished this signaling in irradiated and unirradiated samples. In HDMEC, rapamycin and radiation reduced microvessel sprouting, but did not alter colony formation.

Conclusions: Minimally cytotoxic concentrations of rapamycin inhibited the mTOR cascade in culture and in vivo while radiosensitizing soft tissue sarcoma, and produced synergistic effects with radiation on HDMEC microvessel formation. By targeting both tumor and endothelial compartments, rapamycin produced potent radiosensitization of soft tissue sarcoma xenografts. Clinical trials combining rapamycin and radiotherapy in soft tissue sarcoma are warranted.
Clinical and Immunohistochemical Features Associated with a Response to Bortezomib in Patients with Multiple Myeloma
Authors: Dawson Mark A. , Opat Stephen S. , Taouk Yamna , Donovan Mark , Zammit Michele , Monaghan Katherine , Horvath Noemi , Roberts Andrew W. , Prince H. Miles , Hertzberg Mark , McLean Catriona A. , Spencer Andrew
Published: January 2009

Purpose: Multiple myeloma is an incurable disease with heterogeneous clinical behavior. Bortezomib has offered some patients with relapsed and refractory disease an opportunity for prolonged survival. However, there remains a paucity of data in patients treated with bortezomib that accurately delineates and identifies such patients. This information is crucial to guide management.

Experimental Design: In this study, we aimed to identify the patients most likely to respond to bortezomib salvage therapy. We analyzed the baseline clinical variables and profiled the baseline expression of a broad range of immunohistochemical markers of cell cycle activity, apoptosis, and angiogenesis in a large cohort of multiply relapsed myeloma patients recruited to one of two prospective multicentre trials assessing the efficacy of bortezomib salvage therapy.

Results: Using the European Group for Bone Marrow Transplantation criteria, response (complete or partial) to bortezomib salvage therapy was associated with a previous history of complete response to alternative antimyeloma treatment. Patients who expressed cyclin D1 were more likely to achieve a response. In contrast, patients who expressed p16INK4A, cytoplasmic p53, and the highest intensity of Bcl-2 staining had a poor response. Patients who achieved a response to bortezomib and those patients who expressed cyclin D1 at baseline showed a significant survival advantage. Patients who expressed FGFR3, a poor prognostic marker, responded equally well and had similar outcomes with bortezomib compared with FGFR3-negative patients.

Conclusions: Baseline clinical variables and selective immunohistochemical markers expressed by patients may be used effectively to identify patients that are most likely to achieve a meaningful clinical response to bortezomib salvage therapy.
Dominant Effect of Antiangiogenesis in Combination Therapy Involving Cyclophosphamide and Axitinib
Authors: Ma Jie , Waxman David J.
Published: January 2009

Purpose: Antiangiogenic drug treatment inhibits tumor growth by decreasing blood supply, which can also reduce the delivery of other therapeutic agents. Presently, we investigated the effect of the vascular endothelial growth factor receptor tyrosine kinase inhibitor axitinib (AG-013736) on tumor vascular patency and chemotherapeutic drug uptake. Furthermore, the effect of axitinib on the antitumor activity of combination treatments with cyclophosphamide was examined.

Experimental Design: Prostate cancer PC-3 xenografts were used to evaluate the effect of axitinib treatment on tumor vascular morphology, fluorescent dye perfusion, hypoxia, and uptake of 4-hydroxycyclophosphamide, the active metabolite of the chemotherapeutic prodrug cyclophosphamide. Sequential or simultaneous schedules for axitinib and cyclophosphamide administration were evaluated in both PC-3 tumors and 9L gliosarcoma xenograft models.

Results: Axitinib monotherapy induced sustained growth stasis in PC-3 tumors in association with extensive apoptotic cell death. A substantial decrease in tumor vascular patency was observed, exemplified by a near complete loss of Hoechst 33342 perfusion and the absence of pimonidazole staining in the increasingly hypoxic tumors. Antitumor activity was significantly enhanced in both PC-3 and 9L tumors treated using an optimized schedule of sequential, intermittent axitinib-cyclophosphamide combination therapy despite a 40% to 70% decrease in tumor tissue uptake of 4-hydroxycyclophosphamide.

Conclusions: In axitinib-cyclophosphamide combination therapy, enhanced anticancer activity can be achieved when the reduced tumor cell exposure to the cancer chemotherapeutic agent is compensated by antiangiogenesis-induced tumor cell starvation. This intrinsic antitumor effect was particularly evident in PC-3 tumor xenografts, where tumor blood flow deprivation dominates the overall therapeutic response.
Obstructing Shedding of the Immunostimulatory MHC Class I Chain–Related Gene B Prevents Tumor Formation
Authors: Wu Jennifer D. , Atteridge Catherine L. , Wang Xuanjun , Seya Tsukasa , Plymate Stephen R.
Published: January 2009

Purpose: Clinical observations have suggested that shedding of the MHC class I chain–related molecule (MIC) may be one of the mechanisms by which tumors evade host immunosurveillance and progress. However, this hypothesis has never been proven. In this study, we tested this hypothesis using a prostate tumor model and investigated the effect of shedding of MIC on tumor development.

Experimental Design: We generated a shedding-resistant noncleavable form of MICB (MICB.A2). We overexpressed MICB.A2, the wild-type MICB, and the recombinant soluble MICB (rsMICB) in mouse prostate tumor TRAMP-C2 (TC2) cells and implanted these cells into severe combined immunodeficient mice.

Results: No tumors were developed in animals that were implanted with TC2-MICB.A2 cells, whereas all the animals that were implanted with TC2, TC2-MICB, or TC2-rsMICB cells developed tumors. When a NKG2D-specific antibody CX5 or purified rsMICB was administered to animals before tumor implantation, all animals that were implanted with TC2-MICB.A2 cells developed tumors. In vitro cytotoxicity assay revealed the loss of NKG2D-mediated natural killer cell function in these prechallenged animals, suggesting that persistent levels of soluble MICB in the serum can impair natural killer cell function and thus allow tumor growth.

Conclusions: These data suggest that MIC shedding may contribute significantly to tumor formation by transformed cells and that inhibition of MIC shedding to sustain the NKG2D receptor-MIC ligand recognition may have potential clinical implication in targeted cancer treatment.
Bortezomib, Dexamethasone, and Fibroblast Growth Factor Receptor 3–Specific Tyrosine Kinase Inhibitor in t(4;14) Myeloma
Authors: Bisping Guido , Wenning Doris , Kropff Martin , Gustavus Dirk , Müller-Tidow Carsten , Stelljes Matthias , Munzert Gerd , Hilberg Frank , Roth Gerald J. , Stefanic Martin , Volpert Sarah , Mesters Rolf M. , Berdel Wolfgang E. , Kienast Joachim
Published: January 2009

Purpose: Novel drugs including targeted approaches have changed treatment paradigms for multiple myeloma (MM) and may also have therapeutic potential in the poor-prognosis t(4;14) subset; t(4;14) results in overexpressed and activated fibroblast growth factor receptor 3 (FGFR3). Blocking this receptor tyrosine kinase (RTK) induces apoptosis in t(4;14)+ MM cells and decreases adhesion to bone marrow stromal cells (BMSC). Using combinations of novel drugs, we investigated potential enhancement of single-agent activities within the tumor cells, targeting of the marrow micromilieu, or circumvention of drug resistance in t(4;14)+ MM.

Experimental Design: We tested effects on apoptosis and related signaling pathways in the t(4;14)+ MM subset, applying drug combinations including a FGFR3 tyrosine kinase inhibitor (RTKI), the proteasome inhibitor bortezomib, and dexamethasone.

Results: RTKI, bortezomib, and dexamethasone were active as single agents in t(4;14)+ MM. RTK inhibition triggered complementary proapoptotic pathways (e.g., decrease of Mcl-1, down-regulation of p44/42 mitogen-activated protein kinase, and activation of proapoptotic stress-activated protein/c-Jun NH2-terminal kinases). Synergistic or additive effects were found by combinations of RTKI with dexamethasone or bortezomib. In selected cases of t(4;14)+ MM, triple combinations were superior to dual combinations tested. Prevention from MM cell apoptosis by BMSC or exogenous interleukin-6 was circumvented by drug combinations. In t(4;14)+, N-ras–mutated NCI-H929 cells, resistance to RTKI was overcome by addition of dexamethasone. Notably, the combination of RTKI and dexamethasone showed additive proapoptotic effects in bortezomib-insensitive t(4;14)+ MM.

Conclusions: Combining novel drugs in poor-prognosis t(4;14)+ MM should take into account at least bortezomib sensitivity and probably Ras mutational status.
Functional Analysis of Epidermal Growth Factor Receptor (EGFR) Mutations and Potential Implications for EGFR Targeted Therapy
Authors: Kancha Rama Krishna , von Bubnoff Nikolas , Peschel Christian , Duyster Justus
Published: January 2009

Purpose: Epidermal growth factor receptor (EGFR) mutations in non–small cell lung cancer (NSCLC) might be predictive for clinical response to EGFR inhibitor treatment. However, retrospective analyses of EGFR mutations in clinical trials have shown inconclusive results and the effect of EGFR sequencing in NSCLC is still controversial. Because the vast majority of EGFR mutations described have not been functionally characterized, simple correlation of mutational status and treatment response may not provide reliable information about the predictive value of EGFR mutations. Thus, we aimed to characterize a comprehensive panel of clinically observed EGFR mutations.

Experimental Design and Results: A panel of 30 EGFR mutations was cloned and characterized for kinase activity and the ability to confer growth factor independence. Interestingly, 4 of 30 EGFR mutations showed no kinase activity even after ligand stimulation and were not able to confer growth factor independence. Ba/F3 cells expressing activating EGFR mutants were then used to test the efficacy of EGFR inhibitors in a cell proliferation assay. IC50 values were calculated for gefitinib, erlotinib, and AEE788. We show that the sensitivity of EGFR mutations toward different inhibitors varies significantly, thus establishing a comprehensive sensitivity profile for each inhibitor.

Conclusions: EGFR mutations identified in NSCLC patients display distinct biological features. The variability in kinase activity, transforming potential, and sensitivity to EGFR inhibitors has to be considered in clinical studies aiming to correlate mutational status and drug response. The identification of comprehensive drug resistance profiles opens the opportunity to test alternative EGFR inhibitors in vitro.
Molecular Alterations at 9q33.1 and Polyploidy in Asbestos-Related Lung Cancer
Authors: Nymark Penny , Kettunen Eeva , Aavikko Mervi , Ruosaari Salla , Kuosma Eeva , Vanhala Esa , Salmenkivi Kaisa , Pirinen Risto , Karjalainen Antti , Knuutila Sakari , Wikman Harriet , Anttila Sisko
Published: January 2009

Purpose: Asbestos causes DNA damage and the fibers, together with tobacco smoke, have a synergistic effect on lung cancer risk. We recently identified 18 chromosomal regions that showed differences in DNA copy number between the lung tumors of asbestos-exposed and nonexposed patients. One of the previously identified asbestos-associated chromosomal regions at 9q was further analyzed for allelic imbalance and DNA copy number alterations (CNA) in the lung tumors of asbestos-exposed and nonexposed patients. In addition, the ploidy level of the tumors was studied.

Experimental Design: Allelic imbalance was analyzed at 9q31.3-34.3 with 15 microsatellite markers in 52 lung tumor samples from asbestos-exposed and nonexposed patients. CNA at 9q32-34.3 were characterized by fluorescent in situ hybridization (FISH) with six bacterial artificial chromosome probes in 95 lung tumors. The ploidy level was analyzed in 100 lung tumors with FISH using three to five centromere probes.

Results: Allelic imbalance at 9q31.3-q34.3 was found in all asbestos-exposed patient tumors (100%, 17 of 17) compared with 64% (14 of 22) in the nonexposed cases (P = 0.005). The most significant difference was detected at 9q33.1 (P = 0.002). FISH results showed that also CNA were more frequent at 9q33.1 in the three major histologic types of non–small-cell lung tumors of exposed patients, and the association showed a dose-dependent trend (P = 0.03). Furthermore, we detected more frequent polyploidy among the exposed (48%, 28 of 58) than among the nonexposed (29%, 12 of 42) patient tumors (P < 0.05).

Conclusions: These results provide a basis for the development of a method to identify asbestos-related lung cancer on a molecular level.
Sp-1 and c-Myc Mediate Lysophosphatidic Acid–Induced Expression of Vascular Endothelial Growth Factor in Ovarian Cancer Cells via a Hypoxia-Inducible Factor-1–Independent Mechanism
Authors: Song Yuanda , Wu Jinhua , Oyesanya Regina A. , Lee Zendra , Mukherjee Abir , Fang Xianjun
Published: January 2009

Purpose: Lysophosphatidic acid (LPA), which is present in ascites of ovarian cancer patients, stimulates expression of vascular endothelial growth factor (VEGF). VEGF is essential for the development and abdominal dissemination of ovarian cancer. We examined how LPA drives VEGF expression to gain a better understanding of tumor angiogenesis under normoxic conditions.

Experimental Design: ELISA, Northern blotting, immunoblotting, quantitative PCR, and promoter reporter analysis in combination with small interfering RNA and pharmacologic inhibitors were used to examine LPA-induced VEGF expression and the underlying mechanisms.

Results: LPA stimulated expression of multiple VEGF variants. A 123-bp fragment proximal to the transcriptional initiation site was identified to be functional promoter region responsible for the response to LPA. The fragment harbors consensus sites for several transcription factors including c-Myc and Sp-1 but not hypoxia-inducible factor-1. Blockade of Rho, ROCK, or c-Myc reduced LPA-dependent VEGF production and promoter activation, suggesting that the G12/13-Rho-ROCK-c-Myc cascade partially contributes to VEGF induction by LPA. More significantly, the multiple Sp-1 sites within the responsive region of the VEGF promoter were essential for LPA-mediated transcription. LPA induced Sp-1 phosphorylation and DNA-binding and transcriptional activities. The silencing of Sp-1 expression with small interfering RNA or inhibition of Sp-1 with pharmacologic inhibitors blocked VEGF production induced by LPA.

Conclusions: LPA stimulates hypoxia-inducible factor-1-independent VEGF expression to promote tumor angiogenesis through activation of the c-Myc and Sp-1 transcription factors.
In vitro and In vivo Radiosensitization of Glioblastoma Cells by the Poly (ADP-Ribose) Polymerase Inhibitor E7016
Authors: Russo Andrea L. , Kwon Hyuk-Chan , Burgan William E. , Carter Donna , Beam Katie , Weizheng Xu , Zhang Jie , Slusher Barbara S. , Chakravarti Arnab , Tofilon Philip J. , Camphausen Kevin
Published: January 2009

Purpose: Poly (ADP-ribose) polymerase (PARP) inhibitors are undergoing clinical evaluation for cancer therapy. Because PARP inhibition has been shown to enhance tumor cell sensitivity to radiation, we investigated the in vitro and in vivo effects of the novel PARP inhibitor E7016.

Experimental Design: The effect of E7016 on the in vitro radiosensitivity of tumor cell lines was evaluated using clonogenic survival. DNA damage and repair were measured using γH2AX foci and neutral comet assay. Mitotic catastrophe was determined by immunostaining. Tumor growth delay was evaluated in mice for the effect of E7016 on in vivo (U251) tumor radiosensitivity.

Results: Cell lines exposed to E7016 preirradiation yielded an increase in radiosensitivity with dose enhancement factors at a surviving fraction of 0.1 from 1.4 to 1.7. To assess DNA double-strand breaks repair, γH2AX measured at 24 hours postirradiation had significantly more foci per cell in the E7016/irradiation group versus irradiation alone. Neutral comet assay further suggested unrepaired double-strand breaks with significantly greater DNA damage at 6 hours postirradiation in the combination group versus irradiation alone. Mitotic catastrophe staining revealed a significantly greater number of cells staining positive at 24 hours postirradiation in the combination group. In vivo, mice treated with E7016/irradiation/temozolomide had an additional growth delay of six days compared with the combination of temozolomide and irradiation.

Conclusions: These results indicate that E7016 can enhance tumor cell radiosensitivity in vitro and in vivo through the inhibition of DNA repair. Moreover, enhanced growth delay with the addition of E7016 to temozolomide and radiotherapy in a glioma mouse model suggests a potential role for this drug in the treatment of glioblastoma multiforme.
Suppression of Anoikis by SKP2 Amplification and Overexpression Promotes Metastasis of Esophageal Squamous Cell Carcinoma
Authors: Wang Xiao-Chun , Wu Yu-Peng , Ye Bo , Lin De-Chen , Feng Yan-Bin , Zhang Zi-Qiang , Xu Xin , Han Ya-Ling , Cai Yan , Dong Jin-Tang , Zhan Qi-Min , Wu Min , Wang Ming-Rong
Published: January 2009

The gene of SKP2, located on chromosome 5p13, plays a critical role in cell cycle progression, especially at the G1-S transition, putatively through its control of several cell cycle regulator proteins including p27kip1, p21cip1, p57kip2, p130, cyclin E, and c-Myc. Previous studies in this laboratory revealed that gain of chromosome 5p was often seen in esophageal squamous cell carcinoma (ESCC). In the present study, we examined the amplification status and expression level of SKP2 in ESCC and investigated its clinicopathologic significance. Amplification and elevated expression of SKP2 correlated significantly with tumor stage and positive lymph node metastasis (P < 0.05). The SKP2 protein expression level as determined by immunohistochemical staining showed a significant inverse correlation with p27 protein. In vivo assay showed that inhibition of SKP2 expression also decreased tumor growth and lung metastasis of ESCC cells. At the molecular level, knockdown of SKP2 by RNA interference inhibited cell migration and invasion ability. Knockdown of SKP2 expression sensitized cancer cells to anoikis, and a wobble mutant of SKP2 that is resistant to SKP2 small interfering RNA can rescue this effect. Expression level of pAkt decreased after SKP2 knockdown. Treatment of cells with phosphoinositidyl 3-kinase inhibitor (LY294002) and constitutively activator (insulin-like growth factor I) had significant effects on the anoikis of SKP2 RNA interference cells. These results show for the first time that SKP2 is amplified and overexpressed in ESCC. Elevated expression of SKP2 protected cancer cells from anoikis, and this effect was mediated, at least in part, by the phosphoinositidyl 3-kinase-Akt pathway. (Mol Cancer Res 2009;7(1):12–22)
Two Tumor Suppressors, p27Kip1 and Patched-1, Collaborate to Prevent Medulloblastoma
Authors: Ayrault Olivier , Zindy Frederique , Rehg Jerold , Sherr Charles J. , Roussel Martine F.
Published: January 2009

Two cyclin-dependent kinase inhibitors, p18Ink4c and p27Kip1, are required for proper cerebellar development. Loss of either of these proteins conferred a proliferative advantage to granule neuron progenitors, although inactivation of Kip1 exerted a greater effect. Mice heterozygous for Patched-1 (Ptc1+/−) that are either heterozygous or nullizygous for Kip1 developed medulloblastoma rapidly and with high penetrance. All tumors from Ptc1+/−;Kip1+/− or Ptc1+/−;Kip1−/− mice failed to express the wild-type Ptc1 allele, consistent with its role as a canonical “two-hit” tumor suppressor. In contrast, expression of the wild-type p27Kip1 protein was invariably maintained in medulloblastomas arising in Ptc1+/−;Kip1+/− mice, indicating that Kip1 is haploinsufficient for tumor suppression. Although medulloblastomas occurring in Ptc1+/− mice were histopathologically heterogeneous and contained intermixed regions of both rapidly proliferating and nondividing more differentiated cells, tumors that also lacked Kip1 were uniformly less differentiated, more highly proliferative, and invasive. Molecular analysis showed that the latter medulloblastomas exhibited constitutive activation of the Sonic hedgehog signaling pathway without loss of functional p53. Apart from gains or losses of single chromosomes, with gain of chromosome 6 being the most frequent, no other chromosomal anomalies were identified by spectral karyotyping, and half of the medulloblastomas so examined retained a normal karyotype. In this respect, this mouse medulloblastoma model recapitulates the vast majority of human medulloblastomas that do not sustain TP53 mutations and are not aneuploid. (Mol Cancer Res 2009;7(1):33–40)
`Better the Devil You Know': Submerged Consciousness and Tenant Participation in Housing Stock Transfers
Authors: McCormack John
Published: January 2009

To what extent does the Freirean concept of submerged consciousness contribute towards understanding of tenant participation in housing stock transfers? This paper describes the policy and process of housing stock transfer, noting both its contested nature and the role of tenants in the process. It explores Freire's concept of submerged consciousness, relating it to a cliché common amongst tenants experiencing stock transfer—`better the devil you know'—before applying a Freirean analytical framework to an ethnographic study of a housing stock transfer. The paper concludes that Freire does offer a valuable analytical framework in respect of tenant participation in housing stock transfers, before offering some suggestions for further, related research.
Goal and Personality Trait Development in a Transitional Period: Assessing Change and Stability in Personality Development
Authors: Lüdtke Oliver , Trautwein Ulrich , Husemann Nicole
Published: January 2009

This longitudinal study examined continuity and change in the Big Five personality traits and in the importance of life goals from eight domains (Personal Growth, Relationships, Community, Health, Wealth, Fame, Image, and Hedonism) in 2,141 students in a 2-year period at the transition from school to college or employment. Both personality traits and life goals demonstrated high levels of rank—order and structural stability and showed significant individual differences in individual change. Moreover, mean-level changes were in line with the maturity principle: Scores on Agreeableness, Conscientiousness, and Openness increased over time, whereas Neuroticism decreased. However, the importance of life goals decreased in all domains except health. Reciprocal effects models revealed that there were effects of prior personality traits on subsequent life goal importance but almost no effects of prior life goal importance on subsequent personality traits. Separate analyses by gender showed that the findings were almost invariant across gender.
Ontogeny of Circadian Organization in the Rat
Authors: Yamazaki Shin , Yoshikawa Tomoko , Biscoe Elizabeth W. , Numano Rika , Gallaspy Lauren M. , Soulsby Stacy , Papadimas Evagelia , Pezuk Pinar , Doyle Susan E. , Tei Hajime , Sakaki Yoshiyuki , Block Gene D. , Menaker Michael
Published: January 2009

The mammalian circadian system is orchestrated by a master pacemaker in the brain, but many peripheral tissues also contain independent or quasi-independent circadian oscillators. The adaptive significance of clocks in these structures must lie, in large part, in the phase relationships between the constituent oscillators and their micro- and macroenvironments. To examine the relationship between postnatal development, which is dependent on endogenous programs and maternal/environmental influences, and the phase of circadian oscillators, the authors assessed the circadian phase of pineal, liver, lung, adrenal, and thyroid tissues cultured from Period 1-luciferase ( Per1-luc ) rat pups of various postnatal ages. The liver, thyroid, and pineal were rhythmic at birth, but the phases of their Per1-luc expression rhythms shifted remarkably during development. To determine if the timing of the phase shift in each tissue could be the result of changing environmental conditions, the behavior of pups and their mothers was monitored. The circadian phase of the liver shifted from the day to night around postnatal day (P) 22 as the pups nursed less during the light and instead ate solid food during the dark. Furthermore, the phase of Per1-luc expression in liver cultures from nursing neonates could be shifted experimentally from the day to the night by allowing pups access to the dam only during the dark. Peak Per1-luc expression also shifted from midday to early night in thyroid cultures at about P20, concurrent with the shift in eating times. The phase of Per1-luc expression in the pineal gland shifted from day to night coincident with its sympathetic innervation at around P5. Per1-luc expression was rhythmic in adrenal cultures and peaked around the time of lights-off throughout development; however, the amplitude of the rhythm increased at P25. Lung cultures were completely arrhythmic until P12 when the pups began to leave the nest. Taken together, the data suggest that the molecular machinery that generates circadian oscillations matures at different rates in different tissues and that the phase of at least some peripheral organs is malleable and may shift as the organ's function changes during development.