Gap junction-mediated bystander effect in primary cultures of human malignant gliomas with recombinant expression of the HSVtk gene

Asklund, Thomas; Appelskog, Ioulia B.; Ammerpohl, Ole; Langmoen, Iver A.; Dilber, M. Siraç; Aints, Alar; Ekström, Tomas J.; Almqvist, Per

doi:10.1016/s0014-4827(02)00052-6

Cited by 74 publications

(63 citation statements)

References 27 publications

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“…22 In HSV-TK/ GCV therapy, bystander cytotoxicity has been effective at killing cocultures of HSV-TK-expressing and HSV-TKnon-expressing cells in vitro and at reducing tumor size in vivo. 10,11,[29][30][31][32][33][34][35][36] HSV-TK expression in as few as 10% of tumor cells in vivo resulted in tumor regression or a significant reduction in tumor growth when GCV was administered. Bystander cytotoxicity has been attributed to the transfer of phosphorylated GCV metabolites from HSV-TK-expressing cells to non-HSV-TK-expressing bystander cells, presumably via protein channels known as gap junctions.…”

Section: Introductionmentioning

confidence: 99%

GCV phosphates are transferred between HeLa cells despite lack of bystander cytotoxicity

Gentry

Boucher

et al. 2005

Gene Ther

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The role of gap junctional intercellular communication (GJIC) in bystander killing with herpes simplex virus thymidine kinase (HSV-TK) and ganciclovir (GCV) was evaluated in U251 cells expressing a dominant-negative connexin 43 cDNA (DN14), and in HeLa cells, reportedly devoid of connexin protein. These cell lines both exhibited 0% GJIC when assayed by Lucifer Yellow fluorescent dye microinjection. Bystander cytotoxicity was still apparent in 50:50 cocultures of DN14 and HSV-TK-expressing U251 cells, but not in 50:50 cocultures of HeLa cells. However, the sensitivity of HeLa HSV-TK-expressing cells to GCV decreased nearly 100-fold (IC 90 ¼ 109 mM) when cocultured with bystander cells compared to results in 100% cultures of HSV-TK-expressing cells (IC 90 ¼ 1.2 mM). A more sensitive flow cytometry technique to measure GJIC over 24 h revealed that the DN14 and HeLa cells exhibited detectable levels of communication (29 and 23%, respectively). Transfer of phosphorylated GCV to HeLa bystander cells occurred within 4 h after drug addition, and GCV triphosphate (GCVTP) accumulated to 213784 pmol/10 6 cells after 24 h. In addition, GCVTP levels were decreased in HSV-TK-expressing cells in coculture (867733 pmol/10 6 cells) compared to 100% cultures of HSV-TK-expressing cells (17737188 pmol/10 6 cells). The half-life of GCVTP in the HSV-TK-expressing cells was approximately four times that measured in the bystander cells (12.3 and 3.1 h, respectively). These data suggest that the lack of bystander cytotoxicity in HeLa cocultures is due to low transfer of phosphorylated GCV and a rapid half-life of GCVTP in the bystander cells. Thus, GCV phosphate transfer to non-HSV-TK-expressing bystander cells may mediate either bystander cell killing or sparing of HSV-TK-positive cells, depending upon the cell specific drug metabolism.

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Section: Introductionmentioning

confidence: 99%

GCV phosphates are transferred between HeLa cells despite lack of bystander cytotoxicity

Gentry

Boucher

et al. 2005

Gene Ther

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“…Interestingly, the effects of this type of vectors are amplified by the bystander effect, caused by the passage of the toxic metabolites to cells connected by gap junctions. [23][24][25] This effect increases the therapeutic capacity of this approach; however, a limitation to this system is that the level of intercellular communication varies in different tumors, and may even be blocked in the later stages of carcinogenesis. 24,26,27 We consider, as we will discuss in more detail presently, that the procedure we put forward in this paper shows improvements with respect to the transfer of HSV-tk, because it affects neighboring cells that need not to be joined by gap junctions, thus increasing its range of therapeutic activity.…”

Section: Discussionmentioning

confidence: 99%

Lentiviral transfer of an inducible transgene expressing a soluble form of Gas1 causes glioma cell arrest, apoptosis and inhibits tumor growth

et al. 2010

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Gliomas are the most frequent primary tumors of the central nervous system, and their clinical prognosis remains very poor. Because of the characteristics of gliomas, gene therapy appears as a potentially relevant strategy for their treatment. However, the inability of viral vectors to transfer the therapeutic genes to a significantly high number of tumor cells, due to their limited diffusion and distribution, remains a critical obstacle for their application treating gliomas. We have demonstrated that the overexpression of growth arrest specific1 (Gas1) induces cell arrest and apoptosis and eliminates glioma cells in vitro and when implanted in mice. To improve the therapeutic range of Gas1, we generated lentiviral vectors coding for a soluble form of Gas1. Here, we show that cells infected with this virus produce the mutant protein, that acting both in autocrine and paracrine manners, causes death of infected and neighbor cells, thus importantly enhancing the effect of Gas1. Furthermore, the administration of this vector, or cells expressing it, inhibit the growth of tumors inoculated in mice. We present a gene therapy strategy that increases the effect of the therapeutic molecule by eliminating not just the infected cells that express Gas1, but neighbor non-infected cells.

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“…The GBM-29 cell line was developed from a tumor explant of glioblastoma multiforme as described (36). The cell lines U-343 MG and MGa Cl.…”

Section: Methodsmentioning

confidence: 99%

HDAC inhibitors effectively induce cell type-specific differentiation in human glioblastoma cell lines of different origin

Svechnikova

Almqvist²,

Ekström³

2008

Int J Oncol

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Abstract. The anti-neoplastic effects of histone deacetylase inhibitors (HDACi), Trichostatin A (TSA) and 4-phenylbutyrate (4-PB) on the human glioblastoma cell lines GBM-29, U-343 MG and U-343 MGa Cl. 2:6 were investigated. TSA and 4-PB induced apoptosis in the three cell lines in a doseand time-dependent manner. Whereas caspase-3 activation was detected in all three cell lines, U-343 MG cells were more sensitive to the apoptotic effect of HDACi compared with U-343 MGa Cl. 2:6. TSA and 4-PB induced differentiation in the three cell lines, each cell line developing unique phenotypic characteristics. During long-term treatment with a low dose of HDACi U-343 MGa Cl. 2:6 cells developed an astrocytic morphology with expression of glial fibrillary acidic protein (GFAP). GFAP-negative U-343 MG cells changed their morphology in response to HDACi and down-regulated their expression of vimentin. The nestin and vimentin positive GBM-29 cells also showed a morphological differentiation, while the expression of the two malignancy markers decreased. In summary, our results showed that these three glioblastoma cell lines display unique phenotypes and differentiation patterns in response to HDACi.

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Gap junction-mediated bystander effect in primary cultures of human malignant gliomas with recombinant expression of the HSVtk gene

Cited by 74 publications

References 27 publications

GCV phosphates are transferred between HeLa cells despite lack of bystander cytotoxicity

GCV phosphates are transferred between HeLa cells despite lack of bystander cytotoxicity

Lentiviral transfer of an inducible transgene expressing a soluble form of Gas1 causes glioma cell arrest, apoptosis and inhibits tumor growth

HDAC inhibitors effectively induce cell type-specific differentiation in human glioblastoma cell lines of different origin

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