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Neuroblastoma Cell Line (neuroblastoma + cell_line)
Kinds of Neuroblastoma Cell Line Selected AbstractsFractalkine reduces N -methyl- d -aspartate-induced calcium flux and apoptosis in human neurons through extracellular signal-regulated kinase activationEUROPEAN JOURNAL OF NEUROSCIENCE, Issue 12 2004Kumaran Deiva Abstract Our purpose was to investigate in human neurons the neuroprotective pathways induced by Fractalkine (FKN) against glutamate receptor-induced excitotoxicity. CX3CR1 and FKN are expressed constitutively in the tested human embryonic primary neurons and SK-N-SH, a human neuroblastoma cell line. Microfluorometry assay demonstrated that CX3CR1 was functional in 44% of primary neurons and in 70% of SK-N-SH. Fractalkine induced ERK1/2 phosphorylation within 1 min and Akt phosphorylation after 10 min, and both phosphorylation decreased after 20 min. No p38 and SAPK/JNK activation was observed after FKN treatment. Application of FKN triggered a 53% reduction of the NMDA-induced neuronal calcium influx, which was insensitive to pertussis toxin and LY294002 an inhibitor of Akt pathway, but abolished by PD98059, an ERK1/2 pathway inhibitor. Moreover, FKN significantly reduced neuronal NMDA-induced apoptosis, which was pertussis toxin insensitive and abolished in presence of PD98059 and LY294002. In conclusion, FKN protected human neurons from NMDA-mediated excitotoxicity in at least two ways with different kinetics: (i) an early ERK1/2 activation which reduced NMDA-mediated calcium flux; and (ii), a late Akt activation associated with the previously induced ERK1/2 activation. [source] A novel method of generating neuronal cell lines from gene-knockout mice to study prion protein membrane orientationEUROPEAN JOURNAL OF NEUROSCIENCE, Issue 3 2003Andrea Holme Abstract The technology of gene knockout and transgenic mice has allowed the study of the role of genes and their proteins in animal physiology and metabolism. However, these techniques have often been found to be limited in that some genetic manipulations of mice led either to a fatal phenotype or to compensations that mask the loss of function of the target protein. The experimentation on neurons from transgenic mice is particularly critical in the study of key proteins that may be involved in neurodegeneration. The cell fusion technique has been implemented as a novel way to generate cell lines from prion protein knockout mice. Fusion between neonatal mouse neurons and a neuroblastoma cell line have led to a Prnp°/° cell line that facilitates the study of the knockout phenotype. These cells are readily transfectable and allowed us to study the expression of prion protein mutants on a PrP-knockout background. Using this cell line we have examined the effect of PrP mutations reported to alter PrPc to a transmembrane form. Our results suggest that these mutations do not create transmembrane forms of the protein, but block normal transport of PrP to the cell membrane. [source] Regulation of endogenous human NPFF2 receptor by neuropeptide FF in SK-N-MC neuroblastoma cell lineJOURNAL OF NEUROCHEMISTRY, Issue 2 2006Minna-Liisa Änkö Abstract Neuropeptide FF has many functions both in the CNS and periphery. Two G protein-coupled receptors (NPFF1 and NPFF2 receptors) have been identified for neuropeptide FF. The expression analysis of the peptide and receptors, together with pharmacological and physiological data, imply that NPFF2 receptor would be the primary receptor for neuropeptide FF. Here, we report for the first time a cell line endogenously expressing hNPFF2 receptor. These SK-N-MC neuroblastoma cells also express neuropeptide FF. We used the cells to investigate the hNPFF2 receptor function. The pertussis toxin-sensitive inhibition of adenylate cyclase activity upon receptor activation indicated coupling to Gi/o proteins. Upon agonist exposure, the receptors were internalized and the mitogen-activated protein kinase cascade was activated. Upon neuropeptide FF treatment, the actin cytoskeleton was reorganized in the cells. The expression of hNPFF2 receptor mRNA was up-regulated by neuropeptide FF. Concomitant with the receptor mRNA, the receptor protein expression was increased. The homologous regulation of hNPFF2 receptor correlates with our previous results in vivo showing that during inflammation, the up-regulation of neuropeptide FF mRNA precedes that of NPFF2 receptor. The regulation of hNPFF2 receptor by NPFF could also be important in the periphery where neuropeptide FF has been suggested to function as a hormone. [source] Activation of caspase-3 alone is insufficient for apoptotic morphological changes in human neuroblastoma cellsJOURNAL OF NEUROCHEMISTRY, Issue 6 2002Margaret M. Racke Abstract Activated caspase-3 is considered an important enzyme in the cell death pathway. To study the specific role of caspase-3 activation in neuronal cells, we generated a stable tetracycline-regulated SK-N-MC neuroblastoma cell line, which expressed a highly efficient self-activating chimeric,caspase-3, consisting of the caspase-1 prodomain fused to the caspase-3 catalytic domain. Under expression-inducing conditions, we observed a time-dependent increase of processed caspase-3 by immunostaining for the active form of the enzyme, intracellular caspase-3 enzyme activity, as well as poly(ADP-ribose) polymerase (PARP) cleavage. Induced expression of the caspase fusion protein showed predominantly caspase-3 activity without any apoptotic morphological changes. In contrast, staurosporine treatment of the same cells resulted in activation of multiple caspases and profound apoptotic morphology. Our work provides evidence that auto-activation of caspase-3 can be efficiently achieved with a longer prodomain and that neuronal cell apoptosis may require another caspase or activation of multiple caspase enzymes. [source] Regulation of relaxin 3 gene expression via cAMP-PKA in a neuroblastoma cell lineJOURNAL OF NEUROSCIENCE RESEARCH, Issue 3 2009Masaki Tanaka Abstract Relaxin 3 is expressed in neurons of the brain stem that inneravate wide areas of the forebrain. Relaxin 3 mRNA levels in these neurons are increased in response to restraint stress, and by central administration of corticotropin-releasing factor (CRF). In the present study, we observed that relaxin 3 was expressed in a mouse neuroblastoma cell line, Neuro2a, and investigated the intracellular signaling that activated relaxin 3 gene transcription in vitro. By means of a clone stably transfected with a relaxin 3 promoter-EGFP gene, we observed that dibutyryl cyclic AMP and forskolin increased the relaxin 3 promoter activity. These increases were inhibited by pretreatment with PKA inhibitors, H89 and KT5720. Moreover, the promoter activity was enhanced by CRF treatment after expression of CRF-R1 receptor on the cells. Taken together, these results indicate that relaxin 3 transcription is activated via the cAMP-PKA pathway in the downstream of CRF-R1. © 2008 Wiley-Liss, Inc. [source] Presenilin 1 is involved in the maturation of ,-site amyloid precursor protein-cleaving enzyme 1 (BACE1)JOURNAL OF NEUROSCIENCE RESEARCH, Issue 1 2007Akira Kuzuya Abstract One of the pathologic hallmarks of Alzheimer's disease is the excessive deposition of ,-amyloid peptides (A,) in senile plaques. A, is generated when ,-amyloid precursor protein (APP) is cleaved sequentially by ,-secretase, identified as ,-site APP-cleaving enzyme 1 (BACE1), and ,-secretase, a putative enzymatic complex containing presenilin 1 (PS1). However, functional interaction between PS1 and BACE1 has never been known. In addition to this classical role in the generation of A, peptides, it has also been proposed that PS1 affects the intracellular trafficking and maturation of selected membrane proteins. We show that the levels of exogenous and endogenous mature BACE1 expressed in presenilin-deficient mouse embryonic fibroblasts (PS,/,MEFs) were reduced significantly compared to those in wild-type MEFs. Moreover, the levels of mature BACE1 were increased in human neuroblastoma cell line, SH-SY5Y, stably expressing wild-type PS1, compared to native cells. Conversely, the maturation of BACE1 was compromised under the stable expression of dominant,negative mutant PS1 overexpression. Immunoprecipitation assay showed that PS1 preferably interacts with proBACE1 rather than mature BACE1, indicating that PS1 can be directly involved in the maturation process of BACE1. Further, endogenous PS1 was immunoprecipitated with endogenous BACE1 in SH-SY5Y cells and mouse brain tissue. We conclude that PS1 is directly involved in the maturation of BACE1, thus possibly functioning as a regulator of both ,- and ,-secretase in A, generation. © 2006 Wiley-Liss, Inc. [source] Iron accelerates the conversion of dopamine-oxidized intermediates into melanin and provides protection in SH-SY5Y cellsJOURNAL OF NEUROSCIENCE RESEARCH, Issue 1 2005Yasuhiko Izumi Abstract Parkinson's disease (PD) is characterized by the selective loss of dopaminergic neurons in the substantia nigra (SN), and it has been suggested that dopamine is one of the main endogenous toxins in the genesis of PD. We demonstrated that thiol antioxidants (the reduced form of glutathione, N -acetyl-L-cysteine, and L-cysteine), which conjugate with one dopamine oxidation intermediate, o -quinone, provided almost complete protection from dopamine-mediated toxicity in SH-SY5Y, a human neuroblastoma cell line. In contrast, catalase partially provided protection against cell death caused by dopamine. These data suggest that the generation of dopamine oxidation intermediates, rather than hydrogen peroxide, plays a pivotal role in dopamine-induced toxicity. Iron accumulated in the SN of patients with PD can cause dopaminergic neuronal degeneration by enhancing oxidative stress. However, we found that iron reduced the total amounts of dopamine oxidation intermediates and enhanced the formation of melanin, a final product of dopamine oxidation. Also, addition of iron inhibited dopamine-induced cytotoxicity. These results suggest that iron can provide protection when it accelerates the conversion of dopamine oxidation intermediates. © 2005 Wiley-Liss, Inc. [source] p -quinone mediates 6-hydroxydopamine-induced dopaminergic neuronal death and ferrous iron accelerates the conversion of p -quinone into melanin extracellularlyJOURNAL OF NEUROSCIENCE RESEARCH, Issue 6 2005Yasuhiko Izumi Abstract Parkinson's disease (PD) is characterized by the selective loss of dopaminergic neurons in the substantia nigra (SN). 6-Hydroxydopamine (6-OHDA), a dopaminergic neurotoxin, is detected in human brains and the urine of PD patients. Using SH-SY5Y, a human neuroblastoma cell line, we demonstrated that 6-OHDA toxicity was determined by the amount of p -quinone produced in 6-OHDA auto-oxidation rather than by reactive oxygen species (ROS). Glutathione (GSH), which conjugated with p -quinone, provided significant protection whereas catalase, which detoxified hydrogen peroxide and superoxide anions, failed to block cell death caused by 6-OHDA. Although iron accumulated in the SN of patients with PD can cause dopaminergic neuronal degeneration by enhancing oxidative stress, we found that extracellular ferrous iron promoted the formation of melanin and reduced the amount of p -quinone. The addition of ferrous iron to the culture medium inhibited caspase-3 activation and apoptotic nuclear morphologic changes and blocked 6-OHDA-induced cytotoxicity in SH-SY5Y cells and primary cultured mesencephalic dopaminergic neurons. These data suggested that generation of p -quinone played a pivotal role in 6-OHDA-induced toxicity and extracellular iron in contrast to intracellular iron was protective rather than harmful because it accelerated the conversion of p -quinone into melanin. © 2005 Wiley-Liss, Inc. [source] Proliferation and apoptosis in a neuroblastoma cell line exposed to 900 MHz modulated radiofrequency fieldBIOELECTROMAGNETICS, Issue 3 2006P. Merola Abstract The aim of this study was to examine whether a modulated radiofrequency of the type used in cellular phone communications at a specific absorption rate (SAR) higher than International Commission on Non-ionizing Radiation Protection (ICNIRP) reference level for occupational exposure, could elicit alterations on proliferation, differentiation, and apoptosis processes in a neuroblastoma cell line. The cell line was exposed for 24, 48, and 72 h to 900 MHz radiofrequency and proliferation and differentiation were tested by WST-I assay and by a molecular analysis of specific markers, two oncogenes and a cytoskeleton protein, in exponential growth phase and in synchronized cell cultures. Apoptosis was evaluated by caspase activation analysis and by molecular detection of Poly (ADP-ribose) polimerase (PARP) cleavage. Combined exposures to radiofrequency and to the differentiative agent retinoic acid or to the apoptotic inducer camptothecin were carried out to test possible interference between electromagnetic field and chemical agents. Overall our data suggest that 900 MHz radiofrequency exposure up to 72 h does not induce significant alterations in the three principal cell activities in a neuroblastoma cell line. Bioelectromagnetics 27:164,171, 2006. © 2006 Wiley-Liss, Inc. [source] Cytocompatibility, interactions, and uptake of polyethyleneimine-coated boron nitride nanotubes by living cells: Confirmation of their potential for biomedical applicationsBIOTECHNOLOGY & BIOENGINEERING, Issue 4 2008Gianni Ciofani Abstract Boron nitride nanotubes (BNNTs) have unique physical properties, which can be exploited in the biomedical field. Hence, the surprising lack of reported studies on their biocompatibility and interactions with living cells, addressed by the present paper which deals the results of such an investigation based on 72 h culture of human neuroblastoma cell line (SH-SY5Y) in the presence of an aqueous suspension of polyethyleneimine (PEI)-coated BNNTs. BNNTs conjugated with fluorescent markers (quantum dots) are employed to enable tracking of their uptake by living cells. The results demonstrate good cytocompatibility together with unequivocal BNNT cellular uptake by an energy-dependent endocytic process. Biotechnol. Bioeng. 2008;101: 850,858. © 2008 Wiley Periodicals, Inc. [source] Expression profiles and clinical relationships of ID2, CDKN1B, and CDKN2A in primary neuroblastomaGENES, CHROMOSOMES AND CANCER, Issue 4 2004Sigrun Gebauer Despite considerable research into the etiology of neuroblastoma, the molecular basis of this disease has remained elusive. In contrast to the absence of expression of the known tumor suppressor CDKN2A (also known as p16 and INK4A) in a wide variety of tumor types we have found in previous studies that CDKN2A protein is paradoxically highly expressed in many advanced stage neuroblastomas and unrelated to RB1 status. In the present study, we sought to identify the mechanistic relationships that might influence CDKN2A expression and negate its influence on tumor cell proliferation. In this regard, we examined the role of the tumor-suppressor gene CDKN1B (also known as p27 and Kip1) and the oncogene ID2 in relationship to CDKN2A expression, MYCN amplification, and neuroblastoma pathogenesis in 17 neuroblastoma cell lines and 129 samples of primary tumors of all stages. All neuroblastoma cell lines expressed the ID2 transcript and protein. However, although the majority of primary neuroblastomas also expressed the ID2 transcript, expression of the ID2 protein was undetectable or only barely detectable, regardless of transcript expression. In both cell lines and primary tumors, ID2 expression was independent of both CDKN2A and MYCN expression. In primary neuroblastomas, CDKN1B protein was expressed in significantly fewer advanced-stage neuroblastomas than early-stage neuroblastomas, but its expression had no relationship with CDKN2A expression or MYCN amplification. We concluded that the paradoxical expression of CDKN2A in neuroblastoma cannot be explained by inactivation of the tumor-suppressor gene CDKN1B or overexpression of the oncogene ID2. We further concluded that ID2 is not a target of MYCN regulation nor is it a prognostic factor for neuroblastoma. Finally, the loss of CDKN1B in advanced-stage neuroblastoma suggests this protein may play a role in the neuroblastoma disease process. © 2004 Wiley-Liss, Inc. [source] Oncostatin M enhances the expression of prostaglandin E2 and cyclooxygenase-2 in astrocytes: Synergy with interleukin-1,, tumor necrosis factor-,, and bacterial lipopolysaccharideGLIA, Issue 4 2003Pavle Repovic Abstract Oncostatin M (OSM), a cytokine of the interleukin-6 family, is expressed in rheumatoid arthritis, multiple sclerosis, multiple myeloma, and other inflammatory and neoplastic conditions. Prostaglandin E2 (PGE2), an eicosanoid also associated with inflammation and cancer, has recently been shown to induce OSM expression. We report here that OSM in turn induces PGE2 production by astrocytes and astroglioma cells. More importantly, in combination with the inflammatory mediators IL-1,, tumor necrosis factor-,, and lipopolysaccharide, OSM exhibits a striking synergy, resulting in up to 50-fold higher PGE2 production by astrocytes, astroglioma, and neuroblastoma cell lines. Enhanced PGE2 production by OSM and IL-1, treatment is explained by their effect on cyclooxygenase-2 (COX-2), an enzyme that catalyzes the committed step in PGE2 synthesis. Of the enzymes involved in PGE2 biosynthesis, only COX-2 mRNA and protein levels are synergistically amplified by OSM and IL-1,. Nuclear run-on assays demonstrate that OSM and IL-1, synergistically upregulate transcription of the COX-2 gene, and the mRNA stability assay indicates that COX-2 mRNA is posttranscriptionally stabilized by OSM and IL-1,. To effect synergy on the PGE2 level, OSM signals in part through its gp130/OSMR, receptor, since neutralizing antibodies against gp130 and OSMR,, but not LIFR,, decrease PGE2 production in response to OSM plus IL-1,. SB202190 and U0126, inhibitors of p38 MAPK and ERK1/2 activation, respectively, inhibit IL-1, and OSM upregulation of COX-2 and PGE2, indicating that these MAPK cascades are utilized by both stimuli. This mechanism of PGE2 amplification may be active in brain pathologies where both OSM and IL-1, are present, such as glioblastomas and multiple sclerosis. GLIA 42:433,446, 2003. © 2003 Wiley-Liss, Inc. [source] Protein kinase B modulates the sensitivity of human neuroblastoma cells to insulin-like growth factor receptor inhibitionINTERNATIONAL JOURNAL OF CANCER, Issue 11 2006Ana S. Guerreiro Abstract The potential of the novel insulin-like growth factor receptor (IGF-IR) inhibitor NVP-AEW541 as an antiproliferative agent in human neuroblastoma was investigated. Proliferation of a panel of neuroblastoma cell lines was inhibited by NVP-AEW541 with IC50 values ranging from 0.15 to 5 ,M. Experiments using an IGF-IR neutralizing antibody confirmed that the IGF-IR was essential to support growth of neuroblastoma cell lines. The expression levels of the IGF-IR in individual neuroblastoma cell lines did not correlate with the sensitivities to NVP-AEW541, while coexpression of the IGF-IR and the insulin receptor (IR) correlated with lower sensitivity to the inhibitor in some cell lines. Intriguingly, high levels of activation of Akt/protein kinase B (PKB) and phosphorylation of the ribosomal S6 protein were observed in neuroblastoma cell lines with decreased sensitivities to NVP-AEW541. Inhibition of Akt/PKB activity restored the sensitivity of neuroblastoma cells to the IGF-IR inhibitor. Transfection of neuroblastoma cells with activated Akt or ribosomal protein S6 kinase (S6K) decreased the sensitivity of the cells to NVP-AEW541. IGF-I-stimulated proliferation of neuroblastoma cell lines was completely blocked by NVP-AEW541, or by a combination of an inhibitor of phosphoinositide 3-kinase and rapamycin. In addition to its antiproliferative effects, NVP-AEW541 sensitized neuroblastoma cells to cisplatin-induced apoptosis. Together, our data demonstrate that NVP-AEW541 in combination with Akt/PKB inhibitors or chemotherapeutic agents may represent a novel approach to target human neuroblastoma cell proliferation. © 2006 Wiley-Liss, Inc. [source] Oxidative modulation of nuclear factor-,B in human cells expressing mutant fALS-typical superoxide dismutasesJOURNAL OF NEUROCHEMISTRY, Issue 5 2002Arianna Casciati Abstract Previous evidence supports the notion of a redox regulation of protein phosphatase calcineurin that might be relevant for neurodegenerative processes where an imbalance between generation and removal of reactive oxygen species occurs. We have recently observed that calcineurin activity is depressed in human neuroblastoma cells expressing Cu,Zn superoxide dismutase (SOD1) mutant G93A and in brain areas from G93A transgenic mice, and that mutant G93A-SOD1 oxidatively inactivates calcineurin in vitro. We have studied the possibility that, by interfering directly with calcineurin activity, mutant SOD1 can modulate pathways of signal transduction mediated by redox-sensitive transcription factors. In this paper, we report a calcineurin-dependent activation of nuclear factor-,B (NF-,B) induced by the expression of familial amyotrophic lateral sclerosis (fALS)-SOD1s in human neuroblastoma cell lines. Alteration of the phosphorylation state of I,B, (the inhibitor of NF-,B translocation into the nucleus) and induction of cyclooxygenase 2 are consistent with the up-regulation of this transcription factor in this system. All of these modifications might be relevant to signaling pathways involved in the pathogenesis of fALS. [source] Evidence for a role of the N-terminal domain in subcellular localization of the neuronal connexin36 (Cx36)JOURNAL OF NEUROSCIENCE RESEARCH, Issue 4 2002G. Zoidl Abstract The expression and functional properties of connexin36 (Cx36) have been investigated in two neuroblastoma cell lines (Neuro2A, RT4-AC) and primary hippocampal neurons transfected with a Cx36-enhanced green fluorescent protein (EGFP) expression vector. Transfected cells express Cx36-EGFP mRNA, and Cx36-EGFP protein is localized in the perinuclear area and cell membrane. Upon differentiation of cell lines, Cx36-EGFP protein was detectable in processes with both axonal and dendritic characteristics. Small gap junction plaques were found between adjacent cells, and electrophysiological recordings demonstrated that the electrical properties of these gap junctions were virtually indistinguishable from those reported for native Cx36. Mutagenesis of Cx36 led to the identification of a structural element that interferes with normal protein localization. In contrast, site directed mutagenesis of putative protein phosphorylation motifs did not alter subcellular localization. This excludes phosphorylation/dephosphorylation as a major regulatory step in Cx36 protein transport. © 2002 Wiley-Liss, Inc. [source] Structural studies and model membrane interactions of two peptides derived from bovine lactoferricinJOURNAL OF PEPTIDE SCIENCE, Issue 7 2005Leonard T. Nguyen Abstract The powerful antimicrobial properties of bovine lactoferricin (LfcinB) make it attractive for the development of new antimicrobial agents. An 11-residue linear peptide portion of LfcinB has been reported to have similar antimicrobial activity to lactoferricin itself, but with lower hemolytic activity. The membrane-binding and membrane-perturbing properties of this peptide were studied together with an amidated synthetic version with an added disulfide bond, which was designed to confer increased stability and possibly activity. The antimicrobial and cytotoxic properties of the peptides were measured against Staphylococcus aureus and Escherichia coli and by hemolysis assays. The peptides were also tested in an anti-cancer assay against neuroblastoma cell lines. Vesicle disruption caused by these LfcinB derivatives was studied using the fluorescent reporter molecule calcein. The extent of burial of the two Trp residues in membrane mimetic environments were quantitated by fluorescence. Finally, the solution NMR structures of the peptides bound to SDS micelles were determined to provide insight into their membrane bound state. The cyclic peptide was found to have greater antimicrobial potency than its linear counterpart. Consistent with this property, the two Trp residues of the modified peptide were suggested to be embedded deeper into the membrane. Although both peptides adopt an amphipathic structure without any regular ,-helical or ß-sheet conformation, the 3D-structures revealed a clearer partitioning of the cationic and hydrophobic faces for the cyclic peptide. Copyright © 2004 European Peptide Society and John Wiley & Sons, Ltd. [source] Prosaposin-derived peptides enhanced sprouting of sensory neurons in vitro and induced sprouting at motor endplates in vivoJOURNAL OF THE PERIPHERAL NERVOUS SYSTEM, Issue 3 2000W. Marie Campana Abstract Prosaposin exhibits neurotrophic factor properties that are localized to a 12-amino acid sequence located in the amino terminal portion of the saposin C domain. Prosaptides are peptides derived from the neurotrophic portion of prosaposin; these have been previously reported to be bioactive in neuroblastoma cell lines in vitro. We report that prosaptides were also bioactive in explants of adult primary sensory neurons by dose-dependently increasing both the number (3- to 4-fold) and elongation of these neurites by 50%. Local injection of prosaptides into the gluteus muscle of adult mice also induced sprouting at the motor endplate. Our results indicate that prosaptides are potent neuritogenic factors for both sensory and motor neurons of adult peripheral nerve. [source] Signaling of ERBB receptor tyrosine kinases promotes neuroblastoma growth in vitro and in vivoCANCER, Issue 13 2010Kristen N. Richards MS Abstract BACKGROUND: ERBB receptor tyrosine kinases can mediate proliferation, migration, adhesion, differentiation, and survival in many types of cells and play critical roles in many malignancies. Recent reports suggest a role for EGFR signaling in proliferation and survival of neuroblastoma, a common form of pediatric cancer that often has an extremely poor outcome. METHODS: The authors examined ERBB family expression in neuroblastoma cell lines and patient samples by flow cytometry, western blot, and quantitative real time polymerase chain reaction (Q-PCR). Response to ERBB inhibition was assessed in vitro by cell-cycle analysis and western blot and in vivo by serial tumor-size measurements. RESULTS: A panel of neuroblastoma cell lines and primary patient tumors expressed EGFR, HER-3, and HER-4, with HER-2 in some tumors. HER-4 mRNA was expressed predominantly in cleavable isoforms. Whereas EGFR inhibition with erlotinib and pan-ERBB inhibition with CI-1033 inhibited EGF-induced phosphorylation of EGFR, AKT, and ERK1/2, only CI-1033 induced growth inhibition and dose-dependent apoptosis in vitro. Both CI-1033 and erlotinib treatment of neuroblastoma xenograft tumors resulted in decreased tumor growth in vivo, although CI-1033 was more effective. In vivo expression of EGFR was observed predominantly in vascular endothelial cells. CONCLUSIONS: Pan-ERBB inhibition is required for ERBB-related neuroblastoma apoptosis in vitro, although EGFR contributes indirectly to tumor growth in vivo. Inhibition of EGFR in endothelial cells may be an important aspect of erlotinib's impact on neuroblastoma growth in vivo. Our results suggest that non-EGFR ERBB family members contribute directly to neuroblastoma growth and survival, and pan-ERBB inhibition represents a potential therapeutic target for treating neuroblastoma. Cancer 2010. © 2010 American Cancer Society. [source] | |||||||||||||