Fig. 5.

Human soluble FMS-like tyrosine kinase 3 ligand (Flt3L) and herpes simplex virus type 1–thymidine kinase (TK)-mediated gene therapy induces tumor regression in the multifocal, intracranial glioblastoma multiforme (GBM) model. (A and B) Representative Nissl stain of multifocal tumors, in which the primary tumor mass had been treated with AdFlt3L and AdTK followed by GCV administration. Animals received 5,000 CNS-1 rat glioma cells into the left striatum followed 10 days later by intratumoral injection of adenoviral (Ad) vector expressing Flt3L (AdFlt3L) and an Ad vector expressing TK (AdTK). Immediately after intratumoral viral vector administration, a second distant GBM was implanted (5,000 CNS-1 cells) in the contralateral striatum. No treatment was administered to the second tumor. Animals receiving AdTK began receiving ganciclovir (GCV) 24 h after Ad vector injection (25 mg/kg, twice daily for 7 days). Animals were euthanized as they became moribund or at 60 days postimplantation (long-term survivors) for neuropathological analysis. (A) Representative Nissl stain of an animal that required euthanasia at day 20 after implantation of the primary tumor. Boxes represent approximate location of the higher magnification images of the primary large tumor mass (1) and the site in the contralateral hemisphere where the second tumor was implanted (2). Scale bar = 1,000 μ m. Immunocytochemistry for cells immunoreactive for vimentin (CNS-1 cells and reactive astrocytes), major histocompatibility complex II (MHC II; immune cells), and CD68 (macrophages/activated microglia) within the primary (1) and secondary (2) tumors. Boxes represent approximate area shown in the higher magnification images to the right. Scale bars = 500 μ m and 50 μ m, respectively. (B) Representative image from the brain of a rat euthanized at the end of the experiment (day 60 after primary CNS-1 rat glioma cell implantation). Arrowheads indicate location of primary and secondary tumor injection sites. Boxes represent approximate location of the higher magnification images of site of primary tumor injection (1) and the site in the contralateral hemisphere were the second tumor was implanted (2). Immunocytochemistry for cells immunoreactive for vimentin (CNS-1 tumor cells and reactive astrocytes), MHC II (immune cells), and CD68 (macrophages/activated microglia) within the primary (1) and secondary (2) tumor sites are shown. Scale bars represent 500 μ m and 50 μ m, respectively. (C) Naive rats were unilaterally injected into the striatum with Ad 0 or AdFlt3L and AdTK. GCV administration began 24 h after viral vector injection in all animals. Animals were euthanized 60 days later and evaluated for neuropathology (Nissl) and immune infiltrates (CD68, CD8). Box in the Nissl stain indicates the approximate location of higher magnification images. Scale bars = 1,000 μ m and 50 μ m, respectively.