Convection-enhanced delivery improves distribution and efficacy of tumor-selective retroviral replicating vectors in a rodent brain tumor model

Abstract

In the present study, we compared the therapeutic effect of tumor-selective retroviral replicating vectors (RRV) expressing the yeast cytosine deaminase (CD) delivered by convection-enhanced delivery (CED) or simple injection, followed by systemic administration of the pro-drug, 5-fluorocytosine (5-FC). Treatment with RRV-CD and systemic 5-FC significantly increased survival in rodent U87MG glioma model in comparison with controls (P<0.01). Interestingly, CED of RRV-CD followed by 5-FC further enhanced survival in this animal model in comparison with intra-tumoral injection of RRV-CD, followed by systemic 5-FC (P<0.05). High expression levels of Ki-67 were found in untreated tumors compared with treated. Untreated tumors were also much larger than treated. CED resulted in excellent distribution of RRV while only partial distribution of RRV was obtained after injection. Furthermore, RRV-CD and CD were also found in tumors from treated rats at study end points. These results demonstrated that RRV vectors may efficiently transduce and stably propagate in malignant human glioma, thereby achieving a significant in situ amplification effect after initial administration. We conclude that delivery of RRV into the glioma by CED provides much wider vector distribution than simple injection, and this correlated with better therapeutic outcomes.

Fig. 1. Survival study in nude rats with U87MG xenografts

Thirty-two nude rats with implanted U87MG tumors were randomly divided into 3 groups: control (Black, n=10); manual injection of RRV-CD (Green, n=11); and CED of RRV-CD (Red, n=11). Five days after tumor implantation, rats received either a simple injection or CED of RRV-CD into the brain tumor whereas control animals received CED of 0.9% normal saline. Five days after RRV administration, all the animals received intraperitoneal injections of 5-FC (500 mg/kg) daily for 7 days. Antitumor efficacy was determined as increase in median survival compared to control. Results are shown by Kaplan-Meier plots.

Fig. 2. Representative tumor sizes and Ki-67 immunohistochemistry

Shown are three representative sections of brains from control, injection and CED treatment groups 5 days after 5-FC treatment (20 days after tumor implantation). Scale bar is 2,000 µm in A–C, and 100 µm in D–F.

Fig. 3. Representative GFP staining of distribution of RRV vectors in brain tumor

Panels indicate RRV-GFP delivered by CED (A) or manual injection (B).

Fig. 4. Representative gag staining of RRV vectors in brain tumor at the end-point of the efficacy study

Shown are representative sections from control (CTRL), injected (Inj) and CED rats stained with anti-gag antibodies (Methods). Lower panels are higher magnification images of injected (left) and CED (right) sections.

Fig. 5. Representative cytosine deaminase staining in the brain tumor at end-point of efficacy study

Shown are representative low-magnification images of tumors after either simple injection or CED of RRV-CD at the efficacy study end-point.