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. 2012 Sep;23(9):951-9.
doi: 10.1089/hum.2012.047. Epub 2012 Aug 22.

Characterization of infectivity-enhanced conditionally replicating adenovectors for prostate cancer radiovirotherapy

Michael J Oneal  1 , Miguel A Trujillo, Julia Davydova, Samantha McDonough, Masato Yamamoto, John C Morris 3rd
Affiliations

Characterization of infectivity-enhanced conditionally replicating adenovectors for prostate cancer radiovirotherapy

Michael J Oneal et al. Hum Gene Ther. 2012 Sep.

Abstract

Prostate cancer (PCa) is the second most commonly diagnosed and sixth leading cause of cancer death in American men and one for which no curative therapy exists after metastasis. To meet this need for novel therapies, our laboratory has previously generated conditionally replicating adenovirus (CRAd) vectors expressing the sodium iodide symporter (hNIS). This virus transduced PCa cells and induced functional NIS expression, allowing for noninvasive tumor imaging and combination therapy with radioiodide, referred to as radiovirotherapy. We have now generated two new modified vectors to further improve efficacy. Ad5/3PB-ADP-hNIS and Ad5/3PB-hNIS include a hybrid Ad5/3 fiber knob to improve transduction efficiency, and express NIS from the endogenous major late promoter to restrict NIS expression to target cells. Additionally, Ad5/3PB-ADP-hNIS includes the adenovirus death protein (ADP), which hastens the release of viral particles after assembly. These two vectors specifically induce radioisotope uptake, cytopathic effect, and viral replication in androgen receptor-expressing PCa cell lines with Ad5/3PB-ADP-hNIS showing earlier (131)I uptake and cytolysis at low multiplicity of infection. SPECT-CT imaging of xenograft tumors infected with Ad5/3PB-hNIS showed steady uptake, whereas infection with Ad5/3PB-ADP-hNIS led to increasing uptake, indicating viral spread. Radiovirotherapy of xenograft LNCaP tumors with Ad5/3PB-ADP-hNIS showed the most significant survival extension versus control tumors (p=0.001), but the benefit of radiovirotherapy was not statistically significant compared with virotherapy alone in this model. These results show the potential of Ad5/3PB-ADP-hNIS as a vector for treatment of prostate cancer.

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Figures

FIG. 1.
FIG. 1.
Structures and cytopathic effect assays. (A) Diagrams of the Ad5/3PB-ADP-hNIS and Ad5/3PB-hNIS structures. (B) MTS assays: 5,000 cells were infected at the MOIs shown, and absorbance at 490 nm was read in triplicate at each time point. (C) Crystal violet stains of cells infected with CRAds at the MOIs shown.
FIG. 2.
FIG. 2.
125I uptake kinetics. (A) Cells (105) from permissive and nonpermissive lines were infected with the CRAd vectors and measured daily for radioisotope uptake as described. (B) Comparison of uptake in low-dose infection. Cells were infected with either Ad5/3PB-ADP-hNIS or Ad5/3PB-hNIS at MOI 0.1.
FIG. 3.
FIG. 3.
Burst assay: 4×106 of the described cells were infected at MOI 20, and the medium was titered for pfu at 2 and 4 days post infection by plaque assay on HEK 293A cells. *Below detectable threshold of 1,000 pfu.
FIG. 4.
FIG. 4.
In vivo radioisotope uptake of LNCaP xenografts. Tumor-bearing mice (n=4) were imaged at the time points shown after injection of 109 pfu of virus. At each time point, mice were injected intraperitoneally with 0.5 mCi of 99Tc and imaged by SPECT-CT 1 hr later. Intratumoral uptake was recorded by volume-of-interest analysis. (A) Cross-sectional views of intratumoral isotope concentration. (B) Radioisotope uptake kinetics over 3 weeks of imaging. p.i., post infection.
FIG. 5.
FIG. 5.
Efficacy of radiovirotherapy treatment. LNCaP-xenografted mice were injected intratumorally with 109 pfu of the CRAd vectors or CRAd injection plus an intraperitoneal injection of 2 mCi of 131I 4 days later (n=10). (A) Tumor growth curves comparing mice treated with Ad5/3PB-ADP-hNIS and 131I versus control. (B) Kaplan-Meier curves of treated mice. Censored animals are indicated by tick marks. Significance of survival extension was determined by z tests of coefficients and standard errors generated by Cox Proportional Hazard Regression using controls as baseline.
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