Tissue Inhibitor of Metalloproteinase-3 Expression from an Oncolytic Adenovirus Inhibits Matrix Metalloproteinase Activity In vivo without Affecting Antitumor Efficacy in Malignant Glioma

Abstract
Oncolytic adenoviruses exhibiting tumor-selective replication are promising anticancer agents. Insertion and expression of a transgene encoding tissue inhibitor of metalloproteinase-3 (TIMP-3), which has been reported to inhibit angiogenesis and tumor cell infiltration and induce apoptosis, may improve the antitumor activity of these agents. To assess the effects of TIMP-3 gene transfer to glioma cells, a replication-defective adenovirus encoding TIMP-3 (Ad.TIMP-3) was employed. Ad.TIMP-3 infection of a panel of glioma cell cultures decreased the proliferative capacity of these cells and induced morphologic changes characteristic for apoptosis. Next, a conditionally replicating adenovirus encoding TIMP-3 was constructed by inserting the TIMP-3 expression cassette into the E3 region of the adenoviral backbone containing a 24-bp deletion in E1A. This novel oncolytic adenovirus, AdΔ24TIMP-3, showed enhanced oncolytic activity on a panel of primary cell cultures and two glioma cell lines compared with the control oncolytic virus AdΔ24Luc. In vivo inhibition of matrix metalloproteinase (MMP) activity by AdΔ24TIMP-3 was shown in s.c. glioma xenografts. The functional activity of TIMP-3 was imaged noninvasively using a near-IR fluorescent MMP-2–activated probe. Tumoral MMP-2 activity was significantly reduced by 58% in the AdΔ24TIMP-3–treated tumors 24 hours after infection. A study into the therapeutic effects of combined oncolytic and antiproteolytic therapy was done in both a s.c. and an intracranial model for malignant glioma. Treatment of s.c. (U-87MG) or intracranial (U-87δEGFR) tumors with AdΔ24TIMP-3 and AdΔ24Luc both significantly inhibited tumor growth and prolonged survival compared with PBS-treated controls. However, expression of TIMP-3 in the context of AdΔ24 did not significantly affect the antitumor efficacy of this oncolytic agent.

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