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Query: UMLS:C0406810 (NAME)
 13,345 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Adenovirus (AdV)-mediated gene expression of immune stimulators represents a valuable in vivo approach for gene therapy of human cancer. The expression level of the therapeutic gene is of crucial importance for the efficacy of this type of treatment. Entry of AdV is dependent on the primary adenovirus receptor CAR and the secondary AdV receptor identified earlier to be a member of the integrin family of surface molecules. We have analyzed 14 different human melanoma cell cultures from different stages together with one melanoma cell line for their AdV-mediated transduction and expression efficiency. Recombinant viruses at various concentrations were used for expression of the B7-1 costimulatory molecule under the control of different promoters and the expression levels of B7-1 were analyzed by flow cytometry. AdV-mediated IL-12 expression was measured using a commercial ELISA. Levels of transgene expression were compared with the expression levels of HCAR, the alpha(v)beta3 and alpha(v)beta5 integrins, and HLA class I. In 4 of 14 cell cultures tested, the presence of the primary virus receptor CAR was associated with the high transduction efficiency phenotype when using the B7-1- and IL-12-expressing viruses at a relatively low multiplicity of infection (MOI) of 50. Immunohistochemistry on cryosections from the original biopsies yielded a strong signal specific for CAR. In contrast, cell cultures expressing low or undetectable levels of CAR needed a 20- to 40-fold higher viral input to show comparable expression level of B7-1 or IL-12. Expression levels of the transgenes hardly varied when using different promoters and no association was observed with the presence or absence of HLA class I molecules or with the expression levels of integrins.
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PMID:The presence of human coxsackievirus and adenovirus receptor is associated with efficient adenovirus-mediated transgene expression in human melanoma cell cultures. 982 35

Adenovirus binds to its receptor via the head domain of its fiber protein. We have crystallized the adenovirus serotype 2 (subgroup C) receptor binding domain and solved the structure at 1.5 A resolution by the molecular replacement technique using the known adenovirus type 5 head structure. Included in the high-resolution model are 306 water molecules, five alternative side chain conformations, and individual anisotropic temperature factors for each atom. The overall structure of the serotype 2 head is very similar to its serotype 5 homologue, apart from differences in some of the flexible loops. All but subgroup B adenoviruses are believed to use the recently identified protein CAR (Coxsackievirus and adenovirus receptor) as receptor. By comparison of the two structures and sequence alignment of CAR binding and non-CAR binding serotype fiber heads, we discuss possible receptor binding sites and propose a receptor binding site in a crevice between two monomers on the side of the trimer. The structural basis of the extraordinary stability of the fiber head trimer is also discussed.
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PMID:Structure of the human adenovirus serotype 2 fiber head domain at 1.5 A resolution. 1050 12

Adenovirus (Ad) cell entry involves sequential interactions with host cell receptors that mediate attachment (CAR), internalization (alphavbeta3 and alphavbeta5), and penetration (alphavbeta5) of the endosomal membrane. These events allow the virus to deliver its genome to the nucleus. While integrins alphavbeta3 and alphavbeta5 both promote Ad internalization into cells, integrin alphavbeta5 selectively facilitates Ad-mediated membrane permeabilization and endosome rupture. In the experiments reported herein, we demonstrate that the intracellular domain of the integrin beta5 subunit specifically regulates Ad-mediated membrane permeabilization and gene delivery. CS-1 melanoma cells expressing a truncated integrin beta5 or a chimeric (beta5-beta3) cytoplasmic tail (CT) supported normal levels of Ad endocytosis but had reduced Ad-mediated gene delivery and membrane permeabilization relative to cells expressing a wild-type integrin beta5. Thin-section electron microscopy revealed that virion particles were capable of being endocytosed into cells expressing a truncated beta5CT, but they failed to escape cytoplasmic vesicles and translocate to the nucleus. Site-specific mutagenesis studies suggest that a C-terminal TVD motif in the beta5CT plays a major role in Ad membrane penetration.
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PMID:Regulation of adenovirus membrane penetration by the cytoplasmic tail of integrin beta5. 1068 89

Adenovirus is a widely used vector for cancer gene therapy because of its high infection efficiency and capacity for transgene expression in both dividing and nondividing cells. However, neutralisation of adenovirus by pre-existing antibodies can lead to inefficient delivery, and the wide tissue distribution of the coxsackie and adenovirus receptor (CAR, the primary receptor for adenovirus type 5) precludes target selectivity. These limitations have largely restricted therapeutic use of adenovirus to local or direct administration. A successful viral gene therapy vector would be protected from neutralising antibodies and exhibit a preferential tropism for target cells. We report here the development of a covalent coating and retargeting strategy using a multivalent hydrophilic polymer based on poly-[N-(2-hydroxypropyl)methacrylamide] (pHPMA). Incorporation of targeting ligands such as basic fibroblast growth factor and vascular endothelial growth factor on to the polymer-coated virus produces ligand-mediated, CAR-independent binding and uptake into cells bearing appropriate receptors. Retargeted virus is resistant to antibody neutralisation and can infect receptor-positive target cells selectively in mixed culture, and also in xenografts in vivo. Multivalent polymeric modification of adenovirus is an effective way of changing its tropism and interaction with the immune system. As a non-genetic one-step process, the technology is simple, versatile and should yield vectors with an improved safety profile.
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PMID:Polymer-coated adenovirus permits efficient retargeting and evades neutralising antibodies. 1131 9

In this study we analyzed two ways of retargeting of Ad-vectors to human pancreatic carcinoma with the aim of enhancing the gene transfer efficiency. First, we analyzed the expression of the epidermal growth factor receptor (EGFR) on primary, as well as established pancreatic carcinoma cells by flow cytometry which revealed high expression levels of EGFR on the surface of these cells. We showed that EGFR-retargeted entry pathway using a bispecific fusion protein formed by a recombinant soluble form of truncated Coxsackie and Adenovirus Receptor (sCAR) genetically fused with human EGF (sCAR-EGF) redirects them to the EGFR leading to an enhanced gene transfer efficiency to pancreatic carcinoma cells. Since flow cytometry revealed absence of CAR expression, but the presence of at least one of both alphav integrins on the pancreatic carcinoma cells, a second way of targeting was investigated using a genetically modified Ad vector which has an RGD (Arg-Gly-Asp)-containing peptide inserted into the HI-loop of the fiber knob. This RGD targeted Ad (AdlucRGD) revealed efficient CAR-independent infection by allowing binding to cellular integrins resulting in a dramatic enhancement of gene transfer. These findings have direct relevance for Ad-vector based gene therapy strategies for pancreatic carcinoma.
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PMID:Improved gene transfer efficiency to primary and established human pancreatic carcinoma target cells via epidermal growth factor receptor and integrin-targeted adenoviral vectors. 1143 31

Adenovirus type 5 (Ad) based vectors transduce vascular endothelial cells (EC) and have been widely used for vascular gene transfer. However, many cell types express the Ad receptor (cox-sackievirus adenovirus receptor; CAR), preventing selective EC infection and precluding clinical use. We previously isolated the human EC-binding peptides SIGYPLP and LSNFHSS by phage display and demonstrated by means of a bispecific antibody that SIGYPLP directs efficient, high-level, EC-selective Ad-mediated gene transfer. We now generate genetically modified Ad fiber proteins with selective EC tropism by engineering these peptides into the HI loop of the Ad fiber. SIGYPLP, but not LSNFHSS, enhanced EC selectivity, demonstrating maintenance of peptide-cell binding fidelity upon incorporation into virions. Combining fiber mutations that block CAR binding (detargeting) with SIGYPLP insertion (retargeting) generated a novel Ad vector, AdKO1SIG, in a single component system. AdKO1SIG demonstrated efficient and selective tropism for EC compared with control Ad vectors. This is the first demonstration of genetic incorporation of a novel, mammalian, cell-selective ligand that retains its targeting fidelity in the Ad fiber HI loop, in combination with point mutations that abolish fiber-CAR interaction. This study demonstrates the potential for improving the cell-selectivity and safety of adenoviral vectors.
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PMID:Ablating adenovirus type 5 fiber-CAR binding and HI loop insertion of the SIGYPLP peptide generate an endothelial cell-selective adenovirus. 1173 37

P/Q-type Ca(2+) channels, which are postulated to play major roles in synaptic transmission, are regulated in a variety of ways. Ca(2+) currents through P/Q-type Ca(2+) channels (Ca(v)2.1/beta(1a)/alpha(2)delta) heterologously expressed in mammalian cells were recorded using the whole-cell patch clamp method. The oxidant H(2)O(2) increased the current amplitude and the effect was reversed by the reducing agent dithiothreitol (DTT). The stimulatory effect of H(2)O(2) on the Ca(2+) current was mimicked by the NO donors, SNAP, and diethylamine NONOate, and reversed by the reducing agent DTT. The presence of a soluble guanylate cyclase inhibitor did not abolish the ability of SNAP to increase the Ca(2+) current. Adenovirus-mediated overexpression of nitric oxide synthase in combination with application of the Ca(2+) ionophore A23187 also increased the Ca(2+) current amplitude and the effect was again reversed by DTT. The NOS inhibitor L-NAME abolished the stimulatory effect of A23187, and A23187 did not change the Ca(2+) currents in the cells treated with control adenovirus particles. The time course of the decline of the Ca(2+) current, but not of the Ba(2+) current, in response to repeated depolarization was markedly slowed by adenovirus-mediated overexpression of nitric oxide synthase. The results demonstrate that nitric oxide enhances the channel activity by promoting oxidation and suggest that Ca(2+), nitric oxide synthase, and nitric oxide could constitute a positive feedback loop for regulation of voltage-gated P/Q-type Ca(2+) channels.
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PMID:Nitric oxide augments voltage-gated P/Q-type Ca(2+) channels constituting a putative positive feedback loop. 1190 98

Adenovirus-based gene therapy may provide an alternative mode of treatment for prostate cancer, especially for late-stage and androgen-independent disease for which there is currently no effective treatment. Efficient adenovirus infection of target cells depends upon the presence of the coxsackie adenovirus cell surface receptor, CAR, which is the primary receptor for group C adenoviruses and is important for the attachment of adenovirus to the cell membrane. To evaluate the potential efficacy of adenoviral therapy for prostate cancer, we evaluated CAR expression in normal prostate tissue and in prostate carcinoma of increasing Gleason grades in paraffin-embedded, archival tissues using a polyclonal antibody raised against human CAR. Immunohistochemical analysis of benign prostate epithelia demonstrated intense luminal and lateral cell membrane staining. There was a statistically significant difference in CAR membrane expression with respect to Gleason score. In addition, metastatic prostate specimens demonstrated strong membrane staining for CAR. Adenovirus therapy may, therefore, provide an alternate modality in the treatment of prostate cancer and may be especially efficacious in the treatment of metastatic disease.
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PMID:Expression of the coxsackie adenovirus receptor in normal prostate and in primary and metastatic prostate carcinoma: potential relevance to gene therapy. 1209 94

Adenovirus binds its receptor (CAR), enters cells, and replicates. It must then escape to the environment to infect a new host. We found that following infection, human airway epithelia first released adenovirus to the basolateral surface. Virus then traveled between epithelial cells to emerge on the apical surface. Adenovirus fiber protein, which is produced during viral replication, facilitated apical escape. Fiber binds CAR, which sits on the basolateral membrane where it maintains tight junction integrity. When fiber bound CAR, it disrupted junctional integrity, allowing virus to filter between the cells and emerge apically. Thus, adenovirus exploits its receptor for two important but distinct steps in its life cycle: entry into host cells and escape across epithelial barriers to the environment.
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PMID:Adenovirus fiber disrupts CAR-mediated intercellular adhesion allowing virus escape. 1257 37

Adenovirus (Ad) serotype 5 (Ad5) continues to be the predominant vector used for cancer gene therapy. However, many tumor types are reported to be relatively refractory to Ad5 infection because of low surface expression of the native Ad5 receptor, CAR. The observation that many tumor cells are CAR deficient has necessitated the development of CAR-independent infection strategies, including the introduction of heterologous ligand sequences into the virus fiber gene and immunological or chemical modifications of the capsid proteins. Alternatively, native Ad5 tropism can be modified by substituting the knob region from other Ad serotypes such as Ad type 3 (Ad3) into the Ad5 knob region. To date, the effect(s) of tropism modification on the replication and oncolytic capacity of these chimeric Ad vectors has not been fully evaluated. To address this issue, Ad5 vectors and isogenically matched chimeric vectors with Ad3 tropism (Ad5/3) were compared in this study. Various parameters of virus infection were compared, including binding, nuclear translocation, E1A transcription, transgene expression, de novo virus production, and oncolysis. Overall, the chimeric Ad5/3 virus was progressively more efficient at each step of the replication cycle compared with its Ad5 counterpart. The higher replication efficiency of the chimeric Ad5/3 vector translated into improved therapeutic efficacy in a murine in vivo tumor rejection model. These findings suggest that in addition to the initial target cell interaction, multiple mechanisms contribute to the enhanced replication of the chimeric Ad5/3 vector. Furthermore, the data demonstrate that alternative Ad serotype receptors can be used to improve infection and subsequent oncolytic replication, which is particularly relevant in gene therapy applications for tumors that are inefficiently infected with Ad5.
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PMID:Substitution of the adenovirus serotype 5 knob with a serotype 3 knob enhances multiple steps in virus replication. 1264 86


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