1008. Prostate-Specific Oncolytic Adenovirus Based on Bidirectional TSTA System
Authors: Makoto Sato, Mai Johnson, Sanjiv S. Gambhir, Michael Carey, Lily Wu
Journal: Molecular Therapy (2005)

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Due to the exceptionally limited treatment options currently available for recurrent hormone refractory prostate cancer (HRPC), gene therapy can provide a tool to fight this Herculean disease. To develop a safe and efficient vector-based gene delivery approach, we employed a prostate-targeted gene expression system. The combination of a modified PSA promoter/enhancer and two-step transcriptional amplification (TSTA) system enabled us to achieve robust expression of transgenes in our vectors while maintaining prostate specificity. Furthermore, integration of an efficient adenovirus-mediated gene delivery approach and noninvasive optical imaging allowed us to continuously monitor and evaluate our system on human prostate cancer mouse xenografted models (Sato, et al. Mol Ther, '03). We have also shown that the TSTA system can be usable for clinically-translatable mPET radionuclide imaging, applicable to HRPC where androgen receptor (AR) is expressed (Sato, et al. Clin Cancer Res, '05), and that the suicide gene therapy approach with the TSTA system exhibited promising results, as well as remarkably lower liver toxicity compared to conventional CMV promoter (Johnson et al. submitted).

To further explore therapeutic approaches to utilize the TSTA system, we have taken an oncolytic approach to eradicate prostate cancer. Previously we had confirmed that two distinct genes can be expressed from a single construct because of the structural nature of the TSTA system. A pair of firefly luciferase (FL) and HSV-1 sr39tk genes flanking five repeats of GAL4 binding sites were both expressed from a single adenovirus vector simultaneously. We successfully obtained both optical and mPET imaging on the same animal by this bidirectional vector. Based on this, we developed an oncolytic adenovirus utilizing the TSTA system. In this new vector, both E1A and E1B driven by GAL4 binding sites will make the virus replication competent by the GAL4VP16 activator expressed in trans. Therefore, where the TSTA is active, E1A and E1B will be expressed, and this should lead to oncolytic adenoviral replication. In a preliminary study, virus replication was suggested by the increase of the FL activity from the AdTSTA-FL co-infected in the AR+ prostate cell line, LNCaP, following the transfection of the plasmid with E1A and E1B flanking two GAL4 repeats (G2). However, no increase of the FL activity was seen with the E1A and E1B plasmid with no GAL4 repeat (G0), possibly because of no virus production. The viral production was actually confirmed by a plaque formation in the transfection study with a G2 or G4 driving E1 proteins but was not for G0 in addition to the expression of E1 proteins. Viral replication was also confirmed by co-infection study using G4 adenovirus but not with G0 adenovirus. Currently we are working on analyzing G0, G2, and G4 adenovirus to see oncolytic activity and constructing all-in-one TSTA oncolytic adenoviruses. We foresee that this oncolytic approach based on the TSTA system can be another promising gene therapy strategy for the advanced stages of prostate cancer.