Modified vaccinia virus Ankara as a vector for suicide gene therapy

Abstract

Modified vaccinia virus Ankara (MVA) has been used successfully to express various antigens for the development of vaccines. Here we show that MVA can also be used as an efficient vector for the transfer of suicide genes to cancer cells. We have generated a new and highly potent suicide gene, FCU1, which encodes a fusion protein derived from the yeast cytosine deaminase and uracil phosphoribosyltransferase genes. We now describe the therapeutic benefit of using MVA to deliver and express the FCU1 gene in cancer cells. MVA-mediated transfer of the FCU1 gene to various human tumor cells results in the production of a bifunctional intracellular enzyme, such that exposure to the prodrug 5-FC suppresses the growth of the tumor cells both in vitro and in vivo. Moreover, we report a more potent tumor growth delay at lower doses of 5-FC using MVA-FCU1 in comparison to adenovirus encoding FCU1. Prolonged therapeutic levels of cytotoxic 5-FU were detected in tumors in mice treated with both MVA-FCU1 and 5-FC while no detectable 5-FU was found in the circulation. This original combination between MVA and FCU1 represents a potentially safe and attractive therapeutic option to test in man.

Targeted delivery of a suicide gene to human colorectal tumors by a conditionally replicating vaccinia virus

“…These results are comparable to those obtained with the modified vaccinia virus Ankara (MVA) strain. 16 Expression of the FCU1 fusion protein in the vaccinia context was confirmed by western blot using the mouse monoclonal antibody directed against FCU1 (Figure 1b). Western blot shows that VV-FCU1 expressed the expected 42-kDa FCU1 protein (Figure 1b).…”

“…15 A systemic infusion of the prodrug 5-FC, a Food and Drug Administration-approved antifungal agent, is relatively nontoxic and results in direct intracellular conversion to 5-FU in VV-FCU1-infected tumor cells. 16 This strategy has the potential to be more effective and less toxic than conventional chemotherapy with systemic 5-FU, and in the clinical setting, could enhance the therapeutic index of combination regimens that currently include 5-FU. Additionally, 5-FC administration may also selectively kill any normal actively dividing cells that might have been inadvertently transduced by the vector expressing FCU1.…”

“…Thus the enhanced cell killing by the combination of VV-FCU1 and 5-FC is due to the expression of FCU1 able to activate the prodrug. In comparison, using the nonpropagative MVA-FCU1 16 at an MOI of 0.0001, no cytotoxicity was observed in presence or absence of 5-FC (data not shown). Despite the inhibitory influence of FCU1/5-FC system on VV propagation, VV-FCU1/5-FC combination therapy was still more effective than either treatment alone.…”

“…16 The CDase and UPRTase activities were determined 48 h postinfection by the analysis of the enzymatic conversions of 5-FC to 5-FU and 5-FU to 5-fluorouridine monophosphate (5-FUMP), respectively. This was determined using lysates prepared from human LoVo tumor cells infected at an MOI of 0.0001 by VVTK À and VV-FCU1.…”

“…14,15 Expression of the fusion yeast CDaseHUPRTase gene, designated FCU1, has been reported to increase the antitumor effect in experimental animals in vivo. 15,16 The success of any gene therapy strategy relies on efficient delivery of the transgene specifically to target cells to achieve high therapeutic efficacy whereas limiting unwanted systemic side effects. The use of replication competent viruses is an attractive strategy for tumor therapy because the virus can replicate and spread in situ, exhibiting oncolytic activity through a direct cytopathic effect (CPE).…”