Abstract
Prodrug activator gene therapy mediated by murine leukemia virus (MLV)-based retroviral replicating vectors (RRV) was previously shown to be highly effective in killing glioma cells both in culture and in vivo. To avoid receptor interference and enable dual vector co-infection with MLV-RRV, we have developed another RRV based on gibbon ape leukemia virus (GALV) that also shows robust replicative spread in a wide variety of tumor cells. We evaluated the potential of GALV-based RRV as a cancer therapeutic agent by incorporating yeast cytosine deaminase (CD) and E. coli nitroreductase (NTR) prodrug activator genes into the vector. The expression of CD and NTR genes from GALV-RRV achieved highly efficient delivery of these prodrug activator genes to RG-2 glioma cells, resulting in enhanced cytotoxicity after administering their respective prodrugs 5-fluorocytosine and CB1954 in vitro. In an immune-competent intracerebral RG-2 glioma model, GALV-mediated CD and NTR gene therapy both significantly suppressed tumor growth with CB1954 administration after a single injection of vector supernatant. However, NTR showed greater potency than CD, with control animals receiving GALV-NTR vector alone (i.e., without CB1954 prodrug) showing extensive tumor growth with a median survival time of 17.5 days, while animals receiving GALV-NTR and CB1954 showed significantly prolonged survival with a median survival time of 30 days. In conclusion, GALV-RRV enabled high-efficiency gene transfer and persistent expression of NTR, resulting in efficient cell killing, suppression of tumor growth, and prolonged survival upon CB1954 administration. This validates the use of therapeutic strategies employing this prodrug activator gene to arm GALV-RRV, and opens the door to the possibility of future combination gene therapy with CD-armed MLV-RRV, as the latter vector is currently being evaluated in clinical trials.
Highlights
Glioblastoma multiforme (GBM) is the most frequent form of primary malignant brain tumor in adults [1], and it tends to aggressively invade the surrounding brain tissue so as to make complete surgical resection virtually impossible
Persistent nonlytic infection of tumor cells by retroviral replicating vectors (RRV) facilitates the widespread seeding of prodrug activator genes, thereby allowing synchronized cell killing triggered by prodrug administration
Using gibbon ape leukemia virus (GALV)-based RRV expressing the cytosine deaminase (CD) and NTR suicide genes followed by the administration of the prodrugs 5-FC and CB1954, we have achieved highly efficient killing of glioma cells both in culture and in vivo, resulting in significantly prolonged survival in an immune-competent
Summary
Glioblastoma multiforme (GBM) is the most frequent form of primary malignant brain tumor in adults [1], and it tends to aggressively invade the surrounding brain tissue so as to make complete surgical resection virtually impossible. Clinical trials of GBM gene therapies using a conventional replication-defective retroviral vector encoding Herpes simplex virus thymidine kinase with subsequent administration of the prodrug ganciclovir did not achieve any improvement in patient survival [2], which was attributed to extremely low levels of tumor transduction. We and others have previously shown that amphotropic murine leukemia virus (MLV)-based retroviral replicating vectors (RRV) achieve highly efficient and tumor-selective gene transfer to glioma cells both in culture and in vivo [9,10,11]. Phase clinical studies of RRV-mediated prodrug activator gene therapy have shown highly promising results in recurrent high-grade glioma patients [15,16], and an international Phase III trial is currently on-going
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