bluuebunny/crossref_metadata_2025 huggingface dataset:
G-Rich Proto-Oncogenes Are Targeted for Genomic Instability in B-Cell Lymphomas
Authors: Duquette Michelle L. , Huber Michael D. , Maizels Nancy
Published: March 2007

Diffuse large B-cell lymphoma is the most common lymphoid malignancy in adults. It is a heterogeneous disease with variability in outcome. Genomic instability of a subset of proto-oncogenes, including c-MYC, BCL6, RhoH, PIM1, and PAX5, can contribute to initial tumor development and has been correlated with poor prognosis and aggressive tumor growth. Lymphomas in which these proto-oncogenes are unstable derive from germinal center B cells that express activation-induced deaminase (AID), the B-cell–specific factor that deaminates DNA to initiate immunoglobulin gene diversification. Proto-oncogene instability is evident as both aberrant hypermutation and translocation, paralleling programmed instability which diversifies the immunoglobulin loci. We have asked if genomic sequence correlates with instability in AID-positive B-cell lymphomas. We show that instability does not correlate with enrichment of the WRC sequence motif that is the consensus for deamination by AID. Instability does correlate with G-richness, evident as multiple runs of the base guanine on the nontemplate DNA strand. Extending previous analysis of c-MYC, we show experimentally that transcription of BCL6 and RhoH induces formation of structures, G-loops, which contain single-stranded regions targeted by AID. We further show that G-richness does not characterize translocation breakpoints in AID-negative B- and T-cell malignancies. These results identify G-richness as one feature of genomic structure that can contribute to genomic instability in AID-positive B-cell malignancies. [Cancer Res 2007;67(6):2586–94]
A Single Nucleotide Polymorphism in the MDM2 Gene Disrupts the Oscillation of p53 and MDM2 Levels in Cells
Authors: Hu Wenwei , Feng Zhaohui , Ma Lan , Wagner John , Rice J. Jeremy , Stolovitzky Gustavo , Levine Arnold J.
Published: March 2007

Oscillations of both p53 and MDM2 proteins have been observed in cells after exposure to stress. A mathematical model describing these oscillations predicted that oscillations occur only at selected levels of p53 and MDM2 proteins. This model prediction suggests that oscillations will disappear in cells containing high levels of MDM2 as observed with a single nucleotide polymorphism in the MDM2 gene (SNP309). The effect of SNP309 upon the p53-MDM2 oscillation was examined in various human cell lines and the oscillations were observed in the cells with at least one wild-type allele for SNP309 (T/T or T/G) but not in cells homozygous for SNP309 (G/G). Furthermore, estrogen preferentially stimulated the transcription of MDM2 from SNP309 G allele and increased the levels of MDM2 protein in estrogen-responsive cells homozygous for SNP309 (G/G). These results suggest the possibility that SNP309 G allele may contribute to gender-specific tumorigenesis through further elevating the MDM2 levels and disrupting the p53-MDM2 oscillation. Furthermore, using the H1299-HW24 cells expressing wild-type p53 under a tetracycline-regulated promoter, the p53-MDM2 oscillation was observed only when p53 levels were in a specific range, and DNA damage was found to be necessary for triggering the p53-MDM2 oscillation. This study shows that higher levels of MDM2 in cells homozygous for SNP309 (G/G) do not permit coordinated p53-MDM2 oscillation after stress, which might contribute to decreased efficiency of the p53 pathway and correlates with a clinical phenotype (i.e., the development of cancers at earlier age of onset in female). [Cancer Res 2007;67(6):2757–65]
Inactivation of Apc in the Mouse Prostate Causes Prostate Carcinoma
Authors: Bruxvoort Katia J. , Charbonneau Holli M. , Giambernardi Troy A. , Goolsby James C. , Qian Chao-Nan , Zylstra Cassandra R. , Robinson Daniel R. , Roy-Burman Pradip , Shaw Aubie K. , Buckner-Berghuis Bree D. , Sigler Robert E. , Resau James H. , Sullivan Ruth , Bushman Wade , Williams Bart O.
Published: March 2007

Alterations of the Wnt/β-catenin signaling pathway are positively associated with the development and progression of human cancer, including carcinoma of the prostate. To determine the role of activated Wnt/β-catenin signaling in mouse prostate carcinogenesis, we created a mouse prostate tumor model using probasin-Cre–mediated deletion of Apc. Prostate tumors induced by the deletion of Apc have elevated levels of β-catenin protein and are highly proliferative. Tumor formation is fully penetrant and follows a consistent pattern of progression. Hyperplasia is observed as early as 4.5 weeks of age, and adenocarcinoma is observed by 7 months. Continued tumor growth usually necessitated sacrifice between 12 and 15 months of age. Despite the high proliferation rate, we have not observed metastasis of these tumors to the lymph nodes or other organs. Surgical castration of 6-week-old mice inhibited tumor formation, and castration of mice with more advanced tumors resulted in the partial regression of specific prostate glands. However, significant areas of carcinoma remained 2 months postcastration, suggesting that tumors induced by Apc loss of function are capable of growth under conditions of androgen depletion. We conclude that the prostate-specific deletion of Apc and the increased expression of β-catenin associated with prostate carcinoma suggests a role for β-catenin in prostate cancer and offers an appropriate animal model to investigate the interaction of Wnt signaling with other genetic and epigenetic signals in prostate carcinogenesis. [Cancer Res 2007;67(6):2490–6]
Concomitant Activation of the JAK/STAT, PI3K/AKT, and ERK Signaling Is Involved in Leptin-Mediated Promotion of Invasion and Migration of Hepatocellular Carcinoma Cells
Authors: Saxena Neeraj K. , Sharma Dipali , Ding Xiaokun , Lin Songbai , Marra Fabio , Merlin Didier , Anania Frank A.
Published: March 2007

Various epidemiologic studies have shown that obesity is associated with hepatocellular carcinoma. Leptin, the key player in the regulation of energy balance and body weight control, also acts as a growth factor on certain organs in both normal and disease states. It is plausible that leptin acts to promote hepatocellular carcinogenesis directly affecting malignant properties of liver cancer cells. However, a direct role for leptin in hepatocellular carcinoma has not been shown. In this study, we analyzed the role of leptin and the mechanism(s) underlying its action in hepatocellular carcinoma cells, which express both short and long isoforms of leptin receptors. Treatment with leptin resulted in increased proliferation of both HepG2 and Huh7 cells and involves activation of signal transducers and activators of transcription 3 (STAT3), AKT, and extracellular signal-regulated kinase (ERK) signaling pathways. Leptin-induced phosphorylation of ERK and AKT was dependent on Janus-activated kinase (JAK)/STAT activation. Intriguingly, we also found that leptin potently induces invasion of hepatocellular carcinoma cells in Matrigel invasion and electric cell-substrate impedance-sensing assays. Leptin-stimulated invasion was effectively blocked by pharmacologic inhibitors of JAK/STAT and, to a lesser extent, by ERK and phosphatidylinositol 3-kinase (PI3K) inhibition. Importantly, leptin also induced the migration of both HepG2 and Huh7 cells on fibronectin matrix. Inhibition of JAK/STAT, ERK, and PI3K activation using pharmacologic inhibitors effectively blocked leptin-induced migration of HepG2 and Huh7 cells. Taken together, these data indicate that leptin promotes hepatocellular carcinoma growth, invasiveness, and migration and implicate the JAK/STAT pathway as a critical mediator of leptin action. Our findings have potential clinical implications for hepatocellular carcinoma progression in obese patients. [Cancer Res 2007;67(6):2497–507]
The Proteasome Inhibitor Bortezomib Acts Independently of p53 and Induces Cell Death via Apoptosis and Mitotic Catastrophe in B-Cell Lymphoma Cell Lines
Authors: Strauss Sandra J. , Higginbottom Karen , Jüliger Simone , Maharaj Lenushka , Allen Paul , Schenkein David , Lister T. Andrew , Joel Simon P.
Published: March 2007

Bortezomib is a proteasome inhibitor with proven efficacy in multiple myeloma and non–Hodgkin's lymphoma. This study reports the effects of bortezomib in B-cell lymphoma cell lines with differing sensitivity to bortezomib to investigate factors that influence sensitivity. Bortezomib induced a time- and concentration-dependent reduction in cell viability in five lymphoma cell lines, with EC50 values ranging from 6 nmol/L (DHL-7 cells) to 25 nmol/L (DHL-4 cells) after 72 h. Bortezomib cytotoxicity was independent of p53 function, as all cell lines exhibited mutations by sequence analysis. The difference in sensitivity was not explained by proteasome or nuclear factor-κB (NF-κB) inhibition as these were similar in the most and least sensitive cells. NF-κB inhibition was less marked than that of a specific NF-κB inhibitor, Bay 11-7082. Cell cycle analysis showed a marked G2-arrested population in the least sensitive DHL-4 line only, an effect that was not present with Bay 11-7082 treatment. Conversely, in DHL-7 cells, bortezomib treatment resulted in cells moving into an aberrant mitosis, indicative of mitotic catastrophe that may contribute to increased sensitivity to bortezomib. These studies show that although bortezomib treatment had similar effects on apoptotic and NF-κB signaling pathways in these cell lines, different cell cycle effects were observed and induction of a further mechanism of cell death, mitotic catastrophe, was observed in the more sensitive cell line, which may provide some pointers to the difference in sensitivity between cell lines. An improved understanding of how DHL-7 cells abrogate the G2-M cell cycle checkpoint may help identify targets to increase the efficacy of bortezomib. [Cancer Res 2007;67(6):2783–90]
Adrenomedullin Is Expressed in Pancreatic Cancer and Stimulates Cell Proliferation and Invasion in an Autocrine Manner via the Adrenomedullin Receptor, ADMR
Authors: Ramachandran Vijaya , Arumugam Thiruvengadam , Hwang Rosa F. , Greenson Joel K. , Simeone Diane M. , Logsdon Craig D.
Published: March 2007

The current study investigated adrenomedullin as a potential autocrine regulator of pancreatic cancer cell function. Adrenomedullin was localized in the neoplastic epithelium of 90% (43 of 48) of human pancreatic adenocarcinomas analyzed by immunohistochemistry and was expressed by 100% (8 of 8) of pancreatic cancer cell lines analyzed by reverse transcription-PCR. Pancreatic cancer cell lines also secreted adrenomedullin into the culture medium as determined by ELISA (5 of 5). Exogenous adrenomedullin treatment of Panc-1, BxPC3, and MPanc96 cells in vitro stimulated cell proliferation, invasion, and nuclear factor κB activity, indicating the ability of the cells to respond to adrenomedullin. Treatment of the cell cultures with an adrenomedullin antagonist inhibited basal levels of proliferation and nuclear factor κB activity, supporting the autocrine function of this molecule. Furthermore, increasing adrenomedullin levels by gene transfer to Panc-1 cells increased, whereas adrenomedullin small hairpin RNA silencing in MPanc96 cells inhibited tumor growth and metastasis in vivo. Adrenomedullin is able to act through at least two different receptors, adrenomedullin receptor (ADMR) and calcitonin receptor–like receptor (CRLR). Reverse transcription-PCR and Western blotting indicated that pancreatic cancer cells expressed only ADMR but not CRLR. In contrast, cells found in the tumor microenvironment, primary human pancreatic stellate and endothelial (HUVEC) cells, expressed both ADMR and CRLR. Small hairpin RNA silencing of ADMR in pancreatic cancer cells blocked adrenomedullin-induced growth and invasion, indicating that this receptor is involved in the autocrine actions of adrenomedullin. These data indicate that adrenomedullin acting via ADMR increases the aggressiveness of pancreatic cancer cells and suggests that these molecules may be useful therapeutic targets. [Cancer Res 2007;67(6):2666–75]
Long-term XPC Silencing Reduces DNA Double-Strand Break Repair
Authors: Despras Emmanuelle , Pfeiffer Petra , Salles Bernard , Calsou Patrick , Kuhfittig-Kulle Steffi , Angulo Jaime F. , Biard Denis S.F.
Published: March 2007

To study the relationships between different DNA repair pathways, we established a set of clones in which one specific DNA repair gene was silenced using long-term RNA interference in HeLa cell line. We focus here on genes involved in either nucleotide excision repair (XPA and XPC) or nonhomologous end joining (NHEJ; DNA-PKcs and XRCC4). As expected, XPAKD (knock down) and XPCKD cells were highly sensitive to UVC. DNA-PKcsKD and XRCC4KD cells presented an increased sensitivity to various inducers of double-strand breaks (DSBs) and a 70% to 80% reduction of in vitro NHEJ activity. Long-term silencing of XPC gene expression led to an increased sensitivity to etoposide, a topoisomerase II inhibitor that creates DSBs through the progression of DNA replication forks. XPCKD cells also showed intolerance toward acute γ-ray irradiation. We showed that XPCKD cells exhibited an altered spectrum of NHEJ products with decreased levels of intramolecular joined products. Moreover, in both XPCKD and DNA-PKcsKD cells, XRCC4 and ligase IV proteins were mobilized on damaged nuclear structures at lower doses of DSB inducer. In XPC-proficient cells, XPC protein was released from nuclear structures after induction of DSBs. By contrast, silencing of XPA gene expression did not have any effect on sensitivity to DSB or NHEJ. Our results suggest that XPC deficiency, certainly in combination with other genetic defects, may contribute to impair DSB repair. [Cancer Res 2007;67(6):2526–34]
Genetic Alterations in Mouse Medulloblastomas and Generation of TumorsDe novofrom Primary Cerebellar Granule Neuron Precursors
Authors: Zindy Frederique , Uziel Tamar , Ayrault Olivier , Calabrese Christopher , Valentine Marc , Rehg Jerold E. , Gilbertson Richard J. , Sherr Charles J. , Roussel Martine F.
Published: March 2007

Mice lacking p53 and one or two alleles of the cyclin D–dependent kinase inhibitor p18Ink4c are prone to medulloblastoma development. The tumor frequency is increased by exposing postnatal animals to ionizing radiation at a time when their cerebella are developing. In irradiated mice engineered to express a floxed p53 allele and a Nestin-Cre transgene, tumor development can be restricted to the brain. Analysis of these animals indicated that inactivation of one or both Ink4c alleles did not affect the time of medulloblastoma onset but increased tumor invasiveness. All such tumors exhibited complete loss of function of the Patched 1 (Ptc1) gene encoding the receptor for sonic hedgehog, and many exhibited other recurrent genetic alterations, including trisomy of chromosome 6, amplification of N-Myc, modest increases in copy number of the Ccnd1 gene encoding cyclin D1, and other complex chromosomal rearrangements. In contrast, medulloblastomas arising in Ptc1+/− mice lacking one or both Ink4c alleles retained p53 function and exhibited only limited genomic instability. Nonetheless, complete inactivation of the wild-type Ptc1 allele was a universal event, and trisomy of chromosome 6 was again frequent. The enforced expression of N-Myc or cyclin D1 in primary cerebellar granule neuron precursors isolated from Ink4c−/−, p53−/− mice enabled the cells to initiate medulloblastomas when injected back into the brains of immunocompromised recipient animals. These “engineered” tumors exhibited gene expression profiles indistinguishable from those of medulloblastomas that arose spontaneously. These results underscore the functional interplay between a network of specific genes that recurrently contribute to medulloblastoma formation. [Cancer Res 2007;67(6):2676–84]
Chemoresistance of Endothelial Cells Induced by Basic Fibroblast Growth Factor Depends on Raf-1–Mediated Inhibition of the Proapoptotic Kinase, ASK1
Authors: Alavi Alireza S. , Acevedo Lisette , Min Wang , Cheresh David A.
Published: March 2007

Genotoxic stress induced by anticancer drugs can lead to apoptosis of both angiogenic endothelial cells (ECs) and proliferating tumor cells. However, growth factors such as basic fibroblast growth factor (bFGF) and vascular endothelial cell growth factor (VEGF) present within the tumor microenvironment can promote chemoresistance by suppressing apoptotic mechanisms in these cells. Here, we have identified apoptosis signal-regulating kinase 1 (ASK1), a proapoptotic member of the MAP3K family, as a target of bFGF-mediated survival signaling in ECs. Evidence is provided that ASK1 is required for EC apoptosis in response to the genotoxic chemotherapeutic agent doxorubicin, and that bFGF, but not VEGF, neutralizes the death-promoting activity of ASK1. Specifically, bFGF stimulation promotes the formation of a Raf-1/ASK1 complex at the mitochondria, inhibits ASK1 kinase activity, and protects ECs from genotoxic stress. Mutation of the Raf-1 activation domain (SS338/9AA) not only prevents Raf-1/ASK1 complex formation but abolishes bFGF-mediated EC protection from genotoxic stress. In line with these observations, bFGF, but not VEGF, neutralizes the antiangiogenic effects of doxorubicin in vivo. These findings reveal a new pathway of EC survival signaling and define a molecular mechanism for chemoresistance induced by bFGF. [Cancer Res 2007;67(6):2766–72]
Integrin αvβ3 Controls Activity and Oncogenic Potential of Primed c-Src
Authors: Huveneers Stephan , van den Bout Iman , Sonneveld Petra , Sancho Ana , Sonnenberg Arnoud , Danen Erik H.J.
Published: March 2007

Increased activity of the proto-oncogene c-Src and elevated levels of integrin αvβ3 are found in melanomas and multiple carcinomas. Regulation of c-Src involves “priming” through disruption of intramolecular interactions followed by “activation” through phosphorylation in the kinase domain. Interactions with overexpressed receptor tyrosine kinases or mutations in the SRC gene can induce priming of c-Src in cancer. Here, we show that αvβ3 promotes activation of primed c-Src, causing enhanced phosphorylation of established Src substrates, survival, proliferation, and tumor growth. The β3 cytoplasmic tail is required and sufficient for integrin-mediated stimulation of all these events through a mechanism that is independent of β3 tyrosine phosphorylation. Instead, experiments using Src variants containing the v-Src Src homology 3 (SH3) domain and using mutant β3 subunits indicate that a functional interaction of the β3 cytoplasmic tail with the c-Src SH3 domain is required. These findings delineate a novel integrin-controlled oncogenic signaling cascade and suggest that the interaction of αvβ3 with c-Src may represent a novel target for therapeutic intervention. [Cancer Res 2007;67(6):2693–700]
Oncolytic Treatment and Cure of Neuroblastoma by a Novel Attenuated Poliovirus in a Novel Poliovirus-Susceptible Animal Model
Authors: Toyoda Hidemi , Yin Jiang , Mueller Steffen , Wimmer Eckard , Cello Jeronimo
Published: March 2007

Neuroblastoma is one of the most common solid tumors in children. Treatment is of limited utility for high-risk neuroblastoma and prognosis is poor. Resistance of neuroblastoma to conventional therapies has prompted us to search for a novel therapeutic approach based on genetically modified polioviruses. Poliovirus targets motor neurons leading to irreversible paralysis. Neurovirulence can be attenuated by point mutations or by exchange of genetic elements between different picornaviruses. We have developed a novel and stable attenuated poliovirus, replicating in neuroblastoma cells, by engineering an indigenous replication element (cre), copied from a genome-internal site, into the 5′-nontranslated genomic region (mono-crePV). An additional host range mutation (A133G) conferred replication in mouse neuroblastoma cells (Neuro-2aCD155) expressing CD155, the poliovirus receptor. Crossing immunocompetent transgenic mice susceptible to poliovirus (CD155 tg mice) with A/J mice generated CD155 tgA/J mice, which we immunized against poliovirus. Neuro-2aCD155 cells were then transplanted into these animals, leading to lethal tumors. Despite preexisting high titers of anti-poliovirus antibodies, established lethal s.c. Neuro-2aCD155 tumors in CD155 tgA/J mice were eliminated by intratumoral administrations of A133Gmono-crePV. No signs of paralysis were observed. Interestingly, no tumor growth was observed in mice cured of neuroblastoma that were reinoculated s.c. with Neuro-2aCD155. This result indicates that the destruction of neuroblastoma cells by A133Gmono-crePV may lead to a robust antitumor immune response. We suggest that our novel attenuated oncolytic poliovirus is a promising candidate for effective oncolytic treatment of human neuroblastoma or other cancer even in the presence of present or induced antipolio immunity. [Cancer Res 2007;67(6):2857–64]
Neuronal Differentiation and Synaptogenesis in Retinoblastoma
Authors: Johnson Dianna A. , Zhang Jiakun , Frase Sharon , Wilson Matthew , Rodriguez-Galindo Carlos , Dyer Michael A.
Published: March 2007

Retinoblastomas initiate in the developing retina in utero and are diagnosed during the first few years of life. We have recently generated a series of knockout mouse models of retinoblastoma that recapitulate the timing, location, and progression of human retinoblastoma. One of the most important benefits of these preclinical models is that we can study the earliest stages of tumor initiation and expansion. This is not possible in human retinoblastoma because tumors initiate in utero and are not diagnosed until they are at an advanced stage. We found that mouse retinoblastoma cells exhibit a surprising degree of differentiation, which has not been previously reported for any neural tumor. Early-stage mouse retinoblastoma cells express proteins found normally in retinal plexiform layers. They also extend neurites and form synapses. All of these features, which were characterized by immunostaining, Golgi-Cox staining, scanning electron microscopy, and transmission electron microscopy, suggest that mouse retinoblastoma cells resemble amacrine/horizontal cells from the retina. As late-stage retinoblastoma cells expand and invade the surrounding tissue, they lose their differentiated morphology and become indistinguishable from human retinoblastomas. Taken together, our data suggest that neuronal differentiation is a hallmark of early-stage retinoblastoma and is lost as cells become more aggressive and invasive. We also show that rosette formation is not a hallmark of retinoblastoma differentiation, as previously believed. Instead, rosette formation reflects extensive cell-cell contacts between retinoblastoma cells in both early-stage (differentiated) and late-stage (dedifferentiated) tumors. [Cancer Res 2007;67(6):2701–11]
Inhibitor of Growth 4 Suppresses Cell Spreading and Cell Migration by Interacting with a Novel Binding Partner, Liprin α1
Authors: Shen Jiang-Cheng , Unoki Motoko , Ythier Damien , Duperray Alain , Varticovski Lyuba , Kumamoto Kensuke , Pedeux Remy , Harris Curtis C.
Published: March 2007

Inhibitor of growth 4 (ING4) is a candidate tumor suppressor that plays a major role in gene regulation, cell cycle control, apoptosis, and angiogenesis. ING4 expression is down-regulated in glioblastoma cells and head and neck squamous cell carcinoma. Here, we identified liprin α1/PPFIA1, a cytoplasmic protein necessary for focal adhesion formation and axon guidance, as a novel interacting protein with ING4. ING4 and liprin α1 colocalized at lamellipodia in the vicinity of vinculin. Overexpressed ING4 suppressed cell spreading and cell migration. In contrast, overexpressed liprin α1 enhanced cell spreading and cell migration. Knockdown of endogenous ING4 with RNA interference induced cell motility, whereas knockdown of endogenous liprin α1 suppressed cell motility. ING4 also suppressed cell motility that was enhanced by liprin α1. However, ING4 did not further suppress cell motility when liprin α1 was suppressed with RNA interference, suggesting a functional and mechanistic interdependence between these proteins. In addition to its nuclear functions, cytoplasmic ING4 interacts with liprin α1 to regulate cell migration and, with its known antiangiogenic function, may prevent invasion and metastasis. [Cancer Res 2007;67(6):2552–8]
Galectin-7 in Lymphoma: Elevated Expression in Human Lymphoid Malignancies and Decreased Lymphoma Dissemination by Antisense Strategies in Experimental Model
Authors: Demers Mélanie , Biron-Pain Katherine , Hébert Josée , Lamarre Alain , Magnaldo Thierry , St-Pierre Yves
Published: March 2007

Galectin-7 is found mainly in stratified squamous epithelia as well as in various other types of cancer cells. As with other members of the galectin family, the expression of galectin-7 has been shown to negatively regulate the development of some tumors while correlating with the progression of other tumor types. For example, up-regulation of galectin-7 is associated with rat mammary carcinomas and with progression to T-cell malignancy. Here, we provide evidence indicating that galectin-7 functions as an important molecule in the dissemination of lymphoma cells in vivo. We found that stable transfection of lymphoma cells with a plasmid encoding antisense galectin-7 cDNA significantly inhibited the dissemination and invasion of lymphoma cells to peripheral organs, thereby increasing the survival of mice. We also found that inhibition of galectin-7 in aggressive lymphoma cells correlated with a decreased invasion of tumor cells in target organs and a reduced expression of matrix metalloproteinase-9, a gene associated with a poor prognosis in non–Hodgkin's lymphoma. We finally examined the expression of galectin-7 in 50 specimens of different mature B-cell neoplasms and found high galectin-7 expression levels in a significant proportion of mature B-cell neoplasms but not in normal B cells. Taken together, these findings suggest that galectin-7 is a potential therapeutic target in the treatment of lymphoid malignancies. [Cancer Res 2007;67(6):2824–9]
Warm Hepatic Ischemia-Reperfusion Promotes Growth of Colorectal Carcinoma Micrometastases in Mouse Liver via Matrix Metalloproteinase-9 Induction
Authors: Nicoud Ian B. , Jones Christopher M. , Pierce Janene M. , Earl T. Mark , Matrisian Lynn M. , Chari Ravi S. , Gorden D. Lee
Published: March 2007

Surgical resection remains the best treatment for colorectal metastases isolated to the liver; however, 5-year survival rates following liver resection are only 40% to 50%, with liver recurrence being a significant reason for treatment failure. The ischemia-reperfusion (I/R) injury incurred during liver surgery can lead to cellular dysfunction and elevations in proinflammatory cytokines and matrix metalloproteinases (MMP). In rodents, I/R injury to the liver has been shown to accelerate the outgrowth of implanted tumors. The mechanism for increased tumor growth in the setting of liver I/R injury is unknown. To investigate the effect of I/R on tumor growth, an experimental model was used whereby small hepatic metastases form after 28 days. Mice subjected to 30 min of 70% liver ischemia at the time of tumor inoculation had significantly larger tumor number and volume, and had elevated MMP9 serum and liver tissue MMP9 as evidenced by zymography and quantitative real-time PCR. Mice treated with doxycycline, a broad-spectrum MMP inhibitor, had reduced MMP9 levels and significantly smaller tumor number and volume in the liver. MMP9-null mice were used to determine if the effects of doxycycline were due to the absence of stromal-derived MMP9. The MMP9-null mice, with or without doxycycline treatment, had reduced tumor number and volume that was equivalent to wild-type mice treated with doxycycline. These findings indicate that hepatic I/R-induced elevations in MMP9 contribute to the growth of metastatic colorectal carcinoma in the liver and that postresection MMP9 inhibition may be clinically beneficial in preventing recurrence following hepatic surgery. [Cancer Res 2007;67(6):2720–8]
Functional Neoangiogenesis Imaging of Genetically Engineered Mouse Prostate Cancer Using Three-Dimensional Power Doppler Ultrasound
Authors: Xuan Jim W. , Bygrave Michael , Jiang Hongyi , Valiyeva Fatma , Dunmore-Buyze Joy , Holdsworth David W. , Izawa Jonathan I. , Bauman Glenn , Moussa Madeleine , Winter Scott F. , Greenberg Norman M. , Chin Joseph L. , Drangova Maria , Fenster Aaron , Lacefield James C.
Published: March 2007

We report the first application of high-frequency three-dimensional power Doppler ultrasound imaging in a genetically engineered mouse (GEM) prostate cancer model. We show that the technology sensitively and specifically depicts functional neoangiogenic blood flow because little or no flow is measurable in normal prostate tissue or tumors smaller than 2–3 mm diameter, the neoangiogenesis “switch-on” size. Vascular structures depicted by power Doppler were verified using Microfil-enhanced micro-computed tomography (micro-CT) and by correlation with microvessel distributions measured by immunohistochemistry and enhanced vascularity visualized by confocal microscopy in two GEM models [transgenic adenocarcinoma of the mouse prostate (TRAMP) and PSP94 gene-directed transgenic mouse adenocarcinoma of the prostate (PSP-TGMAP)]. Four distinct phases of neoangiogenesis in cancer development were observed, specifically, (a) an early latent phase; (b) establishment of a peripheral capsular vascular structure as a neoangiogenesis initiation site; (c) a peak in tumor vascularity that occurs before aggressive tumor growth; and (d) rapid tumor growth accompanied by decreasing vascularity. Microsurgical interventions mimicking local delivery of antiangiogenesis drugs were done by ligating arteries upstream from feeder vessels branching to the prostate. Microsurgery produced an immediate reduction of tumor blood flow, and flow remained low from 1 h to 2 weeks or longer after treatment. Power Doppler, in conjunction with micro-CT, showed that the tumors recruit secondary blood supplies from nearby vessels, which likely accounts for the continued growth of the tumors after surgery. The microsurgical model represents an advanced angiogenic prostate cancer stage in GEM mice corresponding to clinically defined hormone-refractory prostate cancer. Three-dimensional power Doppler imaging is completely noninvasive and will facilitate basic and preclinical research on neoangiogenesis in live animal models. [Cancer Res 2007;67(6):2830–9]
Association of CYP1B1 Germ Line Mutations with Hepatocyte Nuclear Factor 1α–Mutated Hepatocellular Adenoma
Authors: Jeannot Emmanuelle , Poussin Karine , Chiche Laurence , Bacq Yannick , Sturm Nathalie , Scoazec Jean-Yves , Buffet Catherine , Nhieu Jeanne Tran Van , Bellanné-Chantelot Christine , de Toma Claudia , Laurent-Puig Pierre , Bioulac-Sage Paulette , Zucman-Rossi Jessica
Published: March 2007

Biallelic somatic mutations of TCF1 coding for hepatocyte nuclear factor 1α (HNF1α) are found in 50% of the hepatocellular adenoma (HCA) cases usually associated with oral contraception. In rare cases, HNF1α germ line mutations could also predispose to familial adenomatosis. In order to identify new genetic factors predisposing to HNF1α-mutated HCA, we searched for mutations in genes involved in the metabolism of estrogen. For 10 genes (CYP1A1, CYP1A2, CYP3A4, CYP3A5, COMT, UGT2B7, NQO1, GSTM1, GSTP1, and GSTT1), we did not find mutations nor differences in the allele distribution among 32 women presenting HNF1α-mutated adenomas compared with 58 controls. In contrast, we identified a CYP1B1 germ line heterozygous mutation in 4 of 32 women presenting HNF1α-mutated adenomas compared with none in 58 controls. We confirmed these results with the identification of four additional CYP1B1 mutations in a second series of 26 cases. No mutations were found in the control group, which was extended to 98 individuals, and only a known rare genetic variant was observed in two controls (P = 0.0003). We did an ethoxyresorufin O-deethylase assay to evaluate the functional consequence of the CYP1B1 mutations. We found reduced enzymatic activity in each CYP1B1 variant. In addition, an E229K CYP1B1 mutation was found in a woman with a germ line HNF1α mutation in a familial adenomatosis context. In this large family, all three patients with adenomatosis bore both HNF1 and CYP1B1 germ line mutations. In conclusion, our data suggested that CYP1B1 germ line–inactivating mutations might increase the incidence of HCA in women with HNF1α mutations. [Cancer Res 2007;67(6):2611–6]
Lens Epithelium-Derived Growth Factor Is an Hsp70-2 Regulated Guardian of Lysosomal Stability in Human Cancer
Authors: Daugaard Mads , Kirkegaard-Sørensen Thomas , Ostenfeld Marie Stampe , Aaboe Mads , Høyer-Hansen Maria , Ørntoft Torben Falck , Rohde Mikkel , Jäättelä Marja
Published: March 2007

Heat shock protein 70-2 (Hsp70-2) is a chaperone protein essential for the growth of spermatocytes and cancer cells. Here, we show that Hsp70-2 depletion triggers lysosomal membrane permeabilization and cathepsin-dependent cell death and identify lens epithelium-derived growth factor (LEDGF) as an Hsp70-2–regulated guardian of lysosomal stability in human cancer. Knockdown of LEDGF in cancer cells induces destabilization of lysosomal membranes followed by caspase-independent and Bcl-2–resistant cell death. Accordingly, ectopic LEDGF stabilizes lysosomes and protects cancer cells against cytotoxicity induced by anticancer agents that trigger the lysosomal cell death pathway. Remarkably, ectopic LEDGF also increases the tumorigenic potential of human cancer cells in immunodeficient mice, and LEDGF expression is increased in human breast and bladder carcinomas correlating with that of Hsp70-2 in invasive bladder cancer. Taken together, these data reveal LEDGF as an oncogenic protein that controls a caspase-independent lysosomal cell death pathway. [Cancer Res 2007;67(6):2559–67]
The Telomerase Reverse Transcriptase (hTERT) Gene Is a Direct Target of the Histone Methyltransferase SMYD3
Authors: Liu Cheng , Fang Xiaolei , Ge Zheng , Jalink Marit , Kyo Satoru , Björkholm Magnus , Gruber Astrid , Sjöberg Jan , Xu Dawei
Published: March 2007

Recent evidence has accumulated that the dynamic histone methylation mediated by histone methyltransferases and demethylases plays key roles in regulation of chromatin structure and transcription. In the present study, we show that SET and MYND domain-containing protein 3 (SMYD3), a histone methyltransferase implicated in oncogenesis, directly trans-activates the telomerase reverse transcriptase (hTERT) gene that is essential for cellular immortalization and transformation. SMYD3 occupies its binding motifs on the hTERT promoter and is required for maintenance of histone H3-K4 trimethylation, thereby contributing to inducible and constitutive hTERT expression in normal and malignant human cells. Knocking down SMYD3 in tumor cells abolished trimethylation of H3-K4, attenuated the occupancy by the trans-activators c-MYC and Sp1, and led to diminished histone H3 acetylation in the hTERT promoter region, which was coupled with down-regulation of hTERT mRNA and telomerase activity. These results suggest that SMYD3-mediated trimethylation of H3-K4 functions as a licensing element for subsequent transcription factor binding to the hTERT promoter. The present findings provide significant insights into regulatory mechanisms of hTERT/telomerase expression; moreover, identification of the hTERT gene as a direct target of SMYD3 contributes to a better understanding of SMYD3-mediated cellular transformation. [Cancer Res 2007;67(6):2626–31]
Oncogenic Activation of the RAS/RAF Signaling Pathway Impairs the Response of Metastatic Colorectal Cancers to Anti–Epidermal Growth Factor Receptor Antibody Therapies
Authors: Benvenuti Silvia , Sartore-Bianchi Andrea , Di Nicolantonio Federica , Zanon Carlo , Moroni Mauro , Veronese Silvio , Siena Salvatore , Bardelli Alberto
Published: March 2007

Monoclonal antibodies (mAbs) against the extracellular domain of the epidermal growth factor receptor (EGFR) have been introduced for the treatment of metastatic colorectal cancer (mCRC). We have reported recently that increased copy number of the EGFR can predict response to anti-EGFR mAbs and that patients might be selected for treatment based on EGFR copy number. Here, we show that mutations activating the RAS/RAF signaling pathway are also predictive and prognostic indicators in mCRC patients, being inversely correlated with response to anti-EGFR mAbs. In cellular models of CRCs, activation of the RAS signaling pathway by introduction of an activated K-RAS allele (Gly12Val) impairs the therapeutic effect of anti-EGFR mAbs. In cancer cells carrying constitutively active RAS, the pharmacologic inhibition of the mitogen-activated protein kinase (MAPK) signaling cascade improves anti-EGFR treatment based on mAbs. These results have implications for the identification of patients who are likely to respond to anti-EGFR treatment. They also provide the rationale for combination therapies, targeted simultaneously to the EGFR and RAS/RAF/MAPK signaling pathways in CRC patients. [Cancer Res 2007;67(6):2643–8]
The Obesity-Cancer Link: Lessons Learned from a Fatless Mouse
Authors: Hursting Stephen D. , Nunez Nomeli P. , Varticovski Lyuba , Vinson Charles
Published: March 2007

Current dogma suggests that the positive correlation between obesity and cancer is driven by white adipose tissue that accompanies obesity, possibly through excess secretion of adipokines. Recent studies in fatless A-Zip/F1 mice, which have undetectable adipokine levels but display accelerated tumor formation, suggest that adipokines are not required for the enhanced tumor development. The A-Zip/F-1 mice are also diabetic and display elevated circulating levels of other factors frequently associated with obesity (insulin, insulin-like growth factor-1, and proinflammatory cytokines) and activation of several signaling pathways associated with carcinogenesis. In view of this information, the risk factors underlying the obesity-cancer link need to be revisited. We postulate that the pathways associated with insulin resistance and inflammation, rather than adipocyte-derived factors, may represent key prevention and therapeutic targets for disrupting the obesity-cancer link. [Cancer Res 2007;67(6):2391–3]
RAD001 (Everolimus) Delays Tumor Onset and Progression in a Transgenic Mouse Model of Ovarian Cancer
Authors: Mabuchi Seiji , Altomare Deborah A. , Connolly Denise C. , Klein-Szanto Andres , Litwin Samuel , Hoelzle Matthew K. , Hensley Harvey H. , Hamilton Thomas C. , Testa Joseph R.
Published: March 2007

The mammalian target of rapamycin (mTOR) is thought to play a critical role in regulating cell growth, cell cycle progression, and tumorigenesis. Because the AKT-mTOR pathway is frequently hyperactivated in ovarian cancer, we hypothesized that the mTOR inhibitor RAD001 (Everolimus) would inhibit ovarian tumorigenesis in transgenic mice that spontaneously develop ovarian carcinomas. We used TgMISIIR-TAg transgenic mice, which develop bilateral ovarian serous adenocarcinomas accompanied by ascites and peritoneal dissemination. Fifty-eight female TgMISIIR-TAg mice were treated with 5 mg/kg RAD001 or placebo twice weekly from 5 to 20 weeks of age. To monitor tumor development, mice were examined biweekly using magnetic resonance microimaging. In vivo effects of RAD001 on Akt-mTOR signaling, tumor cell proliferation, and blood vessel area were analyzed by immunohistochemistry and Western blot analysis. RAD001 treatment markedly delayed tumor development. Tumor burden was reduced by ∼84%. In addition, ascites formation, together with peritoneal dissemination, was detected in only 21% of RAD001-treated mice compared with 74% in placebo-treated animals. Approximately 30% of RAD001-treated mice developed early ovarian carcinoma confined within the ovary, whereas all placebo-treated mice developed advanced ovarian carcinoma. Treatment with RAD001 diminished the expression of vascular endothelial growth factor in tumor-derived cell lines and inhibited angiogenesis in vivo. RAD001 also attenuated the expression of matrix metalloproteinase-2 and inhibited the invasiveness of tumor-derived cells. Taken together, these preclinical findings suggest that mTOR inhibition, alone or in combination with other molecularly targeted drugs, could represent a promising chemopreventive strategy in women at high familial risk of ovarian cancer. [Cancer Res 2007;67(6):2408–13]
Global Expression Analysis of Cancer/Testis Genes in Uterine Cancers Reveals a High Incidence of BORIS Expression
Authors: Risinger John Ian , Chandramouli Gadisetti V.R. , Maxwell G. Larry , Custer Mary , Pack Svetlana , Loukinov Dmitri , Aprelikova Olga , Litzi Tracy , Schrump David S. , Murphy Susan K. , Berchuck Andrew , Lobanenkov Victor , Barrett J. Carl
Published: March 2007

Purpose: Cancer/testis (CT) genes predominantly expressed in the testis (germ cells) and generally not in other normal tissues are aberrantly expressed in human cancers. This highly restricted expression provides a unique opportunity to use these CT genes for diagnostics, immunotherapeutic, or other targeted therapies. The purpose of this study was to identify those CT genes with the greatest incidence of expression in uterine cancers.

Experimental Design: We queried the expression of known and putative CT gene transcripts (representing 79 gene loci) using whole genome gene expression arrays. Specifically, the global gene expressions of uterine cancers (n = 122) and normal uteri (n = 10) were determined using expression data from the Affymetrix HG-U133A and HG-U133B chips. Additionally, we also examined the brother of the regulator of imprinted sites (BORIS) transcript by reverse transcription-PCR and quantitative PCR because its transcript was not represented on the array.

Results: Global microarray analysis detected many CT genes expressed in various uterine cancers; however, no individual CT gene was expressed in more than 25% of all cancers. The expression of the two most commonly expressed CT genes on the arrays, MAGEA9 (24 of 122 cancers and 0 of 10 normal tissues) and Down syndrome critical region 8 (DSCR8)/MMA1 (16 if 122 cancers and 0 of 10 normal tissues), was confirmed by reverse transcription-PCR methods, validating the array screening approach. In contrast to the relatively low incidence of expression of the other CT genes, BORIS expression was detected in 73 of 95 (77%) endometrial cancers and 24 of 31 (77%) uterine mixed mesodermal tumors.

Conclusions: These data provide the first extensive survey of multiple CT genes in uterine cancers. Importantly, we detected a high frequency of BORIS expression in uterine cancers, suggesting its potential as an immunologic or diagnostic target for these cancers. Given the high incidence of BORIS expression and its possible regulatory role, an examination of BORIS function in the etiology of these cancers is warranted.
A Nonfucosylated Anti-HER2 Antibody Augments Antibody-Dependent Cellular Cytotoxicity in Breast Cancer Patients
Authors: Suzuki Eiji , Niwa Rinpei , Saji Shigehira , Muta Mariko , Hirose Makiko , Iida Shigeru , Shiotsu Yukimasa , Satoh Mitsuo , Shitara Kenya , Kondo Masahide , Toi Masakazu
Published: March 2007

Purpose: Removal of fucose residues from the oligosaccharides of human antibody is a powerful approach to enhance antibody-dependent cellular cytotoxicity (ADCC), a potential important antitumor mechanism of therapeutic antibodies. To provide clinically relevant evidence of this mechanism, we investigated ADCC of a fucose-negative version of trastuzumab [anti–human epidermal growth factor receptor 2 (HER2) humanized antibody] using peripheral blood mononuclear cells (PBMC) from breast cancer patients as effector cells.

Experimental Design: Thirty volunteers, including 20 breast cancer patients and 10 normal healthy control donors, were recruited randomly, and aliquots of peripheral blood were collected. ADCC of commercial trastuzumab (fucosylated) and its fucose-negative version were measured using PBMCs drawn from the volunteers as effector cells and two breast cancer cell lines with different HER2 expression levels as target cells. Relationships between cytotoxicity and characteristics of the patients, such as content of natural killer cells in PBMCs, type of therapy, FCGR3A genotypes, etc. were also analyzed.

Results: ADCC was significantly enhanced with the fucose-negative antibody compared with the fucose-positive antibody using PBMCs from either normal donors or breast cancer patients. Enhancement of ADCC was observed irrespective of the various clinical backgrounds of the patients, even in the chemotherapy cohort that presented with a reduced number of natural killer cells and weaker ADCC.

Conclusions: This preliminary study suggests that the use of fucose-negative antibodies may improve the therapeutic effects of anti-HER2 therapy for patients independent of clinical backgrounds.
Phase I Trial of Sorafenib in Combination with IFN α-2a in Patients with Unresectable and/or Metastatic Renal Cell Carcinoma or Malignant Melanoma
Authors: Escudier Bernard , Lassau Nathalie , Angevin Eric , Soria Jean Charles , Chami Linda , Lamuraglia Michele , Zafarana Eric , Landreau Veronique , Schwartz Brian , Brendel Eric , Armand Jean-Pierre , Robert Caroline
Published: March 2007

Purpose: To determine the safety, maximum tolerated dose, pharmacokinetics, and efficacy, and to evaluate biomarkers, of the multikinase inhibitor sorafenib plus IFN α-2a in advanced renal cell carcinoma (RCC) or melanoma.

Experimental Design: Patients received 28-day cycles of continuous, oral sorafenib twice daily and s.c. IFN thrice weekly: sorafenib 200 mg twice daily plus IFN 6 million IU (MIU) thrice weekly (cohort 1); and sorafenib 400 mg twice daily plus IFN 6 MIU thrice weekly (cohort 2); or plus IFN 9 MIU thrice weekly (cohort 3). Tumor response was assessed by Response Evaluation Criteria in Solid Tumors and dynamic contrast-enhanced ultrasonography.

Results: Thirteen patients received at least one dose of sorafenib plus IFN (12 RCC; one melanoma). The maximum tolerated dose was not reached [only one dose-limiting toxicity (grade 3 asthenia)]. Most frequently reported drug-related adverse events were grade 2 or less in severity, including fatigue, diarrhea, nausea, alopecia, and hand-foot skin reaction. One (7.7%) RCC patient achieved partial response and eight (61.5%) had stable disease (including the melanoma patient). Good responders assessed by dynamic contrast-enhanced ultrasonography had increased progression-free survival and overall survival, relative to poor responders. IFN had no effect on the pharmacokinetics of sorafenib. There were no significant changes in absolute values of lymphocytes, levels of proangiogenic cytokines, or inhibition of phosphorylated extracellular signal-regulated kinase in T cells or natural killer cells, with combination therapy.

Conclusions: This sorafenib combination was well tolerated, with preliminary antitumor activity in advanced RCC and melanoma patients. There were no drug-drug interactions and the recommended dose for future studies is sorafenib 400 mg twice daily plus IFN 9 MIU.