A guanylate kinase/HSV-1 thymidine kinase fusion protein enhances prodrug-mediated cell killing

Willmon, Candice; Krabbenhoft, Elizabeth A.; Black, Margaret E.

doi:10.1038/sj.gt.3302794

Cited by 22 publications

(27 citation statements)

References 18 publications

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“…The most extensively evaluated of these is mutant SR39, which is 43-fold more sensitive to GCV and 20-fold more sensitive to ACV than the parental HSVtk (Black et al, 1996(Black et al, , 2001. Applying a completely different strategy to improve the therapeutic effect of the HSVtk/GCV system, Willmon et al succeeded in decreasing the IC 50 of GCV by 175-fold by using a fusion protein of HSVtk and guanylate kinase (GMK), the cellular enzyme converting GCV-DP to GCV-TP following phosphorylation of GCV by HSVtk, when compared to HSVtk alone (Willmon et al, 2006).…”

Section: Thymidine Kinasementioning

confidence: 97%

Suicide genes for cancer therapy

Portsmouth

Hlavatý

Renner³

2007

Molecular Aspects of Medicine

139

155

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Section: Thymidine Kinasementioning

confidence: 97%

Suicide genes for cancer therapy

Portsmouth

Hlavatý

Renner³

2007

Molecular Aspects of Medicine

139

155

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“…In particular, poor prodrug activation kinetics, 15,16 limited cell toxicity mechanisms, 17 evolution of drug resistance, 18,19 and prophylactic use of GCV in the clinic 20 impact broad implementation of that approach. Furthermore, the poor lipophilicity of the hydrophilic GCV and its reduced ability to penetrate the blood --brain barrier are considered limiting in HSV-tk/GCV-based applications targeted to the brain in the clinic.…”

Section: Introductionmentioning

confidence: 99%

Bystander killing of malignant cells via the delivery of engineered thymidine-active deoxycytidine kinase for suicide gene therapy of cancer

Neschadim

Wang²,

Lavie

et al. 2012

Cancer Gene Ther

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Activity and specificity of chemotherapeutic agents against solid tumors can be augmented via the targeted or localized delivery of 'suicide' genes. Selective activation of specific prodrugs in cells expressing the 'suicide' gene drives their elimination by apoptosis, while also enabling the killing of adjacent bystander cells. Strong bystander effects can compensate for poor 'suicide' gene delivery, and depend on the prodrugs used and mechanisms for the acquisition of activated drug by the bystander population, such as the presence of gap junctional intercellular communications. Although a number of 'suicide' gene therapies for cancer have been developed and characterized, such as herpes simplex virus-derived thymidine kinase (HSVtk)-based activation of ganciclovir, their limited success highlights the need for the development of more robust approaches. Limiting activation kinetics and evolution of chemoresistance are major obstacles. Here we describe 'suicide' gene therapy of cancer based on the lentivirus-mediated delivery of a thymidine-active human deoxycytidine kinase variant. This enzyme possesses substrate plasticity that enables it to activate a multitude of prodrugs, some with distinct mechanisms of action. We evaluated the magnitude and mechanisms of bystander effects induced by different prodrugs, and show that when used in combination, they can synergistically enhance the bystander effect while avoiding off-target toxicity.

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“…While BmGK K m for dGMP was found to be similar to humans (Auvynet et al 2009), differences were found with K m of GMP as compared with humans, mouse, yeast, P. falciparum and Arabidopsis thaliana (Li et al 1996;Kumar et al 2001;Willmon et al 2006;Kandeel et al 2008;Auvynet et al 2009). Lower K m value for GMP as compared with the human enzyme indicates a higher affinity for GMP and thus a tendency to convert substrate into product at a higher rate.…”

Section: Discussionmentioning

confidence: 87%

Purification and characterization of guanylate kinase, a nucleoside monophosphate kinase ofBrugia malayi

et al. 2014

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Guanylate kinase, a nucleoside monophosphate kinase of Brugia malayi which is involved in reversible transfer of phosphate groups from ATP to GMP, was cloned, expressed and characterized. The native molecular mass of BmGK was found to be 45 kDa as determined by size exclusion chromatography and glutaraldehyde cross-linking which revealed that the protein is homodimer in nature. This is a unique characteristic among known eukaryotic GKs. GMP and ATP served as the most effective phosphate acceptor and donor, respectively. Recombinant BmGK utilized both GMP and dGMP, as substrates showing Km values of 30 and 38 μ m, respectively. Free Mg+2 (un-complexed to ATP) and GTP play a regulatory role in catalysis of BmGK. The enzyme showed higher catalytic efficiency as compared with human GK and showed ternary complex (BmGK-GMP-ATP) formation with sequential substrate binding. The secondary structure of BmGK consisted of 45% α-helices, 18% β-sheets as revealed by CD analysis. Homology modelling and docking with GMP revealed conserved substrate binding residues with slight differences. Differences in kinetic properties and oligomerization of BmGK compared with human GK can provide the way for design of parasite-specific inhibitors.

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A guanylate kinase/HSV-1 thymidine kinase fusion protein enhances prodrug-mediated cell killing

Cited by 22 publications

References 18 publications

Suicide genes for cancer therapy

Suicide genes for cancer therapy

Bystander killing of malignant cells via the delivery of engineered thymidine-active deoxycytidine kinase for suicide gene therapy of cancer

Purification and characterization of guanylate kinase, a nucleoside monophosphate kinase ofBrugia malayi

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