Monitoring viral decontamination procedures with green fluorescent protein-expressing adenovirus

Losert, Annemarie; Mauritz, Ilse; Erlach, Natascha; Herbacek, Irene; Schulte‐Hermann, Rolf; Holzmann, Klaus; Grusch, Michael

doi:10.1016/j.ab.2006.04.034

Cited by 8 publications

(6 citation statements)

References 12 publications

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“…An adenovirus-expressing green fluorescent protein (GFP) was used as control. 27 Virus titers were determined with the Adeno X Rapid Titer Kit (Clontech, Mountain View, CA). Cells were exposed to the constructs at the lowest multiplicity of infection leading to 100% cell infection (titrated by the GFP adenovirus).…”

Section: Methodsmentioning

confidence: 99%

Fibroblast growth factor receptor 3 isoforms: Novel therapeutic targets for hepatocellular carcinoma?

et al. 2015

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Fibroblast growth factor receptors (FGFRs) are frequently up-regulated in subsets of hepatocellular carcinoma (HCC). Here, we provide mechanistic insight that FGFR3 splice variants IIIb and IIIc impact considerably on the malignant phenotype of HCC cells. The occurrence of FGFR3 variants was analyzed in human HCC samples. In hepatoma/hepatocarcinoma cell lines, FGFR3 isoforms were overexpressed by lentiviral constructs or down-modulated by small interfering RNA (siRNA; affecting FGFR3-IIIb and -IIIc) or an adenoviral kinase-dead FGFR3-IIIc construct (kdFGFR3). Elevated levels of FGFR3-IIIb and/or -IIIc were found in 53% of HCC cases. FGFR3-IIIb overexpression occurred significantly more often in primary tumors of large (pT2-4) than of small size (pT1). Furthermore, one or both isoforms were enhanced mostly in cases with early tumor infiltration and/or recurrence at the time of surgery or follow-up examinations. In hepatoma/hepatocarcinoma cells, up-regulated FGFR3-IIIb conferred an enhanced capability for proliferation. Both FGFR3-IIIb and FGFR3-IIIc suppressed apoptotic activity, enhanced clonogenic growth, and induced disintegration of the blood/lymph endothelium. The tumorigenicity of cells in severe combined immunodeficiency mice was augmented to a larger degree by variant IIIb than by IIIc. Conversely, siRNA targeting FGFR3 and kdFGFR3 reduced clonogenicity, anchorage-independent growth, and disintegration of the blood/lymph endothelium in vitro. Furthermore, kdFGFR3 strongly attenuated tumor formation in vivo. Conclusions: Deregulated FGFR3 variants exhibit specific effects in the malignant progression of HCC cells. Accordingly, blockade of FGFR3-mediated signaling may be a promising therapeutic approach to antagonize growth and malignant behavior of HCC cells. (HEPATOLOGY 2015;62:1767-1778 H epatocellular carcinoma (HCC), the most frequent primary malignant liver tumor entity, is estimated to be the second-leading cause for cancer mortality worldwide. 1 Owing to the high inherent chemoresistance of this tumor type, standard chemotherapy is insufficient and only 15% of the patients are candidates for surgical resection or liver transplantation. As a result, HCC patients are usually left with an infaust prognosis. Thus, new therapeutic approaches are eagerly needed. 2,3 Up-regulation of growth/survival factors and their respective receptors is known to drive the formation, progression, and dissemination of HCC cells within the body. 2-4 Recently, we and others found overexpression of fibroblast growth factor (FGF) receptor (FGFR) 3 and/or FGFR4 in the majority of HCC cases and relatively rare up-regulation of FGFR1 and/or FGFR2, which was confirmed in the present study (Supporting Fig. 1). 5,6 Meanwhile, it is known that FGFR4 is involved in the

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Section: Methodsmentioning

confidence: 99%

Fibroblast growth factor receptor 3 isoforms: Novel therapeutic targets for hepatocellular carcinoma?

et al. 2015

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“…The resulting FGFR4-cyan fluorescent protein chimera was transferred into pAd/CCMV/V5-Dest by Gateway recombination (Invitrogen). Virus amplification was done as described and an adenovirus-expressing GFP was used as control (Losert et al , 2006). Virus titers were determined with the Adeno-X Rapid Titer Kit (Clontech, Mountain View, CA).…”

Section: Methodsmentioning

confidence: 99%

Fibroblast Growth Factor Receptors as Therapeutic Targets in Human Melanoma: Synergism with BRAF Inhibition

Metzner

Bedeir

Held

et al. 2011

Journal of Investigative Dermatology

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Cutaneous melanoma is a tumor with rising incidence and a very poor prognosis at the disseminated stage. Melanomas are characterized by frequent mutations in BRAF and also by overexpression of fibroblast growth factor 2 (FGF2), offering opportunities for therapeutic intervention. We investigated inhibition of FGF signaling and its combination with dacarbazine or BRAF inhibitors as an antitumor strategy in melanoma. The majority of melanoma cell lines displayed overexpression of FGF2 but also FGF5 and FGF18 together with different isoforms of FGF receptors (FGFRs) 1-4. Blockade of FGF signals with dominant-negative receptor constructs (dnFGFR1, 3, or 4) or small-molecule inhibitors (SU5402 and PD166866) reduced melanoma cell proliferation, colony formation, as well as anchorage-independent growth, and increased apoptosis. DnFGFR constructs also significantly inhibited tumor growth in vivo. Combination of FGF inhibitors with dacarbazine showed additive or antagonistic effects, whereas synergistic drug interaction was observed when combining FGFR inhibition with the multikinase/BRAF inhibitor sorafenib or the V600E mutant-specific BRAF inhibitor RG7204. In conclusion, FGFR inhibition has antitumor effects against melanoma cells in vitro and in vivo. Combination with BRAF inhibition offers a potential for synergistic antimelanoma effects and represents a promising therapeutic strategy against advanced melanoma.

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“…Discs were dried and autoclaved before use. Purified replication‐deficient recombinant human adenovirus Ad5GFP, an E1/E3 gene deleted vector expressing green fluorescent protein (GFP), a reporter gene providing a simple and rapid measure of viral transduction and gene expression in HEK 293 cells (Losert et al. 2006) (obtained from Professor Hill’s group, University of Oxford), was grown in HEK 293 cells (ECACC) and purified using standard caesium chloride density centrifugation to a titre of approximately 10 12 TCID 50 ml −1 .…”

Section: Negative Log Of Tcid50 Endpoint For the Control Untreated 5mentioning

confidence: 99%

“…It was possible to clearly see a single cell expressing GFP. This detection technique has been shown to be a quick and effective way of detecting viable adenovirus when assessing decontamination procedures (Losert et al 2006). The TCID 50 of each resuspended virus was estimated using the Karber method (Karber 1931):…”

Section: Introductionmentioning

confidence: 99%

Hydrogen peroxide vapour (HPV) inactivation of adenovirus

Berrie

Andrews

Yezli

et al. 2011

Letters in Applied Microbiology

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Aims: Adenovirus contamination can be problematic in various settings including life science laboratories and during pharmaceutical manufacturing processes. Stringent and effective decontamination procedures are necessary to minimize the risk of personnel exposure or product cross‐contamination in these settings. Hydrogen peroxide vapour (HPV) is sporicidal, tuberculocidal and fungicidal with proven efficacy against some viruses. We investigate the efficacy of HPV for the inactivation of a recombinant adenovirus. Methods and Results: In this study, the survival of a dried recombinant adenovirus (Ad5GFP) was tested before and after HPV exposure to determine the efficacy of HPV at inactivating adenovirus. A > 8‐log TCID50 reduction resulted from 45‐min exposure to HPV in a microbiological safety cabinet. Conclusions: HPV is effective for the inactivation of a recombinant adenovirus. Significance and impact of the study: The results suggest that HPV may be useful for adenovirus decontamination in life science laboratories or in manufacturing facilities.

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Monitoring viral decontamination procedures with green fluorescent protein-expressing adenovirus

Cited by 8 publications

References 12 publications

Fibroblast growth factor receptor 3 isoforms: Novel therapeutic targets for hepatocellular carcinoma?

Fibroblast growth factor receptor 3 isoforms: Novel therapeutic targets for hepatocellular carcinoma?

Fibroblast Growth Factor Receptors as Therapeutic Targets in Human Melanoma: Synergism with BRAF Inhibition

Hydrogen peroxide vapour (HPV) inactivation of adenovirus

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