Expression of the gene encoding secreted placental alkaline phosphatase (SEAP) by a nondefective adenovirus vector

Doronin, Konstantin; Zakharchuk, A. N.; Grinenko, N. F.; Yurov, Gleb K.; Krougliak, Valeri; Naroditsky, Boris S.

doi:10.1016/0378-1119(93)90374-c

Cited by 10 publications

(6 citation statements)

References 13 publications

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“…Determination of the tightness of the TetOn system using secreted embryonic alkaline phosphatase To examine the functionality and tightness of the new TetOn system we used secreted embryonic alkaline phosphatase (SEAP), previously employed by others for the detection of baculovirus and adenovirus expression (Davis et al 1992;Chung et al 1993;Doronin et al 1993;Joshi et al 2000). Synthesis and secretion of SEAP were examined in infected glioma cell lines using a chemiluminescence kit.…”

Section: Modification Of the Teton Systemmentioning

confidence: 99%

Endogenous GD3 ganglioside induces apoptosis in U‐1242 MG glioma cells

Saqr

Omran

Dasgupta

et al. 2006

Journal of Neurochemistry

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GD3 ganglioside induces apoptosis in several cell types, but the molecular events through which this occurs are largely unknown. We investigated the apoptotic effects of GD3 expression using U-1242 MG glioblastoma cells, as these cells synthesize almost exclusively GM3 and GM2 but not GD3. To express GD3 under the control of the TetOn system with minimum leakage, we modified the system by constructing a single tri-cistronic retrovirus vector containing three genes separated by two internal ribosome entry sites: (a) transcriptional silencer, tTS; (b) mutant of reverse transcriptional activator, rtTA2(S)-M2 (provided by H. Bujard, Heidelberg, Germany); and (c) enhanced green fluorescent protein (EGFP), as an indicator of the tri-cistronic gene expression. Using flow cytometry, we selected glioma cells (U1242MG-GD3 clone) that express high levels of GD3 in response to doxycycline. Expression of GD3 was associated with apoptosis as verified by annexin-V binding, TdT-mediated dUTPnick end-labelling assay (TUNEL), and EGFP degradation. GD3-induced apoptosis occurred via caspase-8 activation, as GD3 caused cleavage of caspase-8 and inhibition of caspase-8 activation by zlETD-fmk minimized GD3-induced apoptosis.

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Section: Modification Of the Teton Systemmentioning

confidence: 99%

Endogenous GD3 ganglioside induces apoptosis in U‐1242 MG glioma cells

Saqr

Omran

Dasgupta

et al. 2006

Journal of Neurochemistry

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“…21). To generate the enzyme (phosphatase) input under the control of these cells, secreted embryonic alkaline phosphatase (SEAP)2021 was expressed in HEK293- seap+ cells (using Xfect-mediated transfection by a pBluescript vector containing CMV- seap , Addgene, plasmid 24595). SEAP was chosen as a form of huPLAP that lacks the membrane-anchoring domain, and is therefore secreted by cells, potentially allowing extracellular control.…”

Section: Resultsmentioning

confidence: 99%

Creation of a gated antibody as a conditionally functional synthetic protein

et al. 2014

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The ability to conditionally direct antibodies is a potentially powerful application for Synthetic Biology in Medicine. Here we show that control of antibody binding through site-specific, chemical phosphorylation of a recognition domain creates a ‘gated’ antibody (Ab). This displays a crude Boolean logic where binding is induced in an enzyme-AND-antigen dependent manner. This ‘AND-Ab’ is therefore active only in the presence of two biomarker inputs: the simultaneous expression of a (cell surface) antigen and secreted enzyme to generate function in vitro, on cells and in mammalian tissue. Such gated Abs, either alone or in combination, could allow the application of logic strategies to enhance precision in biological interrogation, modulation and in therapy.

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“…The E3 region in these vectors is identical to the E3 region in KD3 allowing for overexpression of ADP in the absence of immunomodulatory E3 genes. 32 The TetON-EGFP and TetON-SEAP vectors were constructed for visualization 33 and quantification 30 of transgene expression, respectively.…”

Section: Discussionmentioning

confidence: 99%

“…5,30 Trans-complementing binary vector system allows for regulatable expression of functionally active TRAIL We studied the expression of TRAIL in Hep3B cells infected with the TetON-TRAIL vector alone or in combination with KD3 or KD3-IFN. TRAIL was detected by western blotting only in cells coinfected with TetON-TRAIL and either RC vector, and only in the presence of Dox (Figure 3a) implying that the antibody against TRAIL had a sensitivity threshold preventing detection of TRAIL at lower concentrations.…”

Section: Design Of the Vectors Kd3 (mentioning

confidence: 99%

Anticancer activity of oncolytic adenovirus vector armed with IFN-α and ADP is enhanced by pharmacologically controlled expression of TRAIL

et al. 2007

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We have previously described oncolytic adenovirus (Ad) vectors KD3 and KD3-interferon (IFN) that were rendered cancer-specific by mutations in the E1A region of Ad; these mutations abolish binding of E1A proteins to p300/CBP and pRB. The antitumor activity of the vectors was enhanced by overexpression of the Adenovirus Death Protein (ADP, E3-11.6K) and by replication-linked expression of IFN-a. We hypothesized that the anticancer efficacy of the KD3-IFN vector could be further improved by expression of tumor necrosis factor-related apoptosis-inducing ligand (TRAIL). E1-deleted Ad vectors were constructed carrying reporter genes for enhanced green fluorescent protein or secreted placental alkaline phosphatase (SEAP) and a therapeutic gene for TRAIL under control of the TetON system. Expression of the genes was increased in the presence of a helper virus and the inducer doxycycline such that up to 231-fold activation of expression for the TetON-SEAP vector was obtained. Coinfection with TetON-TRAIL augmented oncolytic activity of KD3 and KD3-IFN in vitro. Induction of TRAIL expression did not reduce the yield of progeny virus. Combination of TetON-TRAIL and KD3-IFN produced superior antitumor activity in vivo as compared with either vector alone demonstrating the efficacy of a four-pronged cancer gene therapy approach, which includes Ad oncolysis, ADP overexpression, IFN-a-mediated immunotherapy, and pharmacologically controlled TRAIL activity.

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Expression of the gene encoding secreted placental alkaline phosphatase (SEAP) by a nondefective adenovirus vector

Cited by 10 publications

References 13 publications

Endogenous GD3 ganglioside induces apoptosis in U‐1242 MG glioma cells

Endogenous GD3 ganglioside induces apoptosis in U‐1242 MG glioma cells

Creation of a gated antibody as a conditionally functional synthetic protein

Anticancer activity of oncolytic adenovirus vector armed with IFN-α and ADP is enhanced by pharmacologically controlled expression of TRAIL

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