Article Figures & Data
Figures
- Fig. 1.
Xenograft experiments in nude mice. Animals (n = 5 per group) were treated with different anti-VEGF compounds or a control antibody. In A, 5 × 106 human A673 rhabdomyosarcoma cells were injected s.c. in the dorsal area. Treatment was started after day 5 by i.p. administration of Mab A.4.6.1 or control Mab at 10 mg/kg, twice weekly. Intratumoral injection of mFlt(1-3)-IgG, Fab, or control Mab was at 25 mg/kg, every other day. In B, mice were treated as indicated. Mice receiving Mab A.4.6.1 plus mFlt(1-3)-IgG were treated until day 33. C, systemic administration of VEGF inhibitors. A673 cells were injected as described above. Animals were injected i.p. with Mab A.4.6.1 at 10 mg/kg twice weekly or with mFlt(1-3)-IgG daily at the doses indicated. Control Mab was administered at the dose of 25 mg/kg. After 2 weeks, the animals were killed, and the tumor weight was determined.
- Fig. 2.
Effects of anti-VEGF treatments on the histopathology of A673 xenografts. Serial sections of tumors harvested from mice treated with control antibody (A–D), systemic Mab 4.6.1 and intratumoral FAb (E–H), or systemic Mab 4.6.1 and intratumoral mFlt(1-3)-IgG (I–L) were stained with H&E (A, B, E, F, I, J) or by immunoperoxidase for the endothelial cell markers Flk-1 (C, G, K) and CD31 (D, H, L). In A, control tumors showed regions of geographic necrosis that were often most pronounced in deeper areas (bottom). In B, entrapped host-derived elements such as skeletal muscle fibers (arrows) were frequently seen in the invasive tumors. In C and D, tumor microvessels were often present in increased numbers near host elements such as entrapped muscle fibers (arrows). In E, Mab A.4.6.1-treated tumors also showed geographic necrosis. In some areas, the tumor-necrosis interface stained more deeply than the viable tumor (arrows); this is attributable to nuclear pyknosis, an early necrotic change (see J also ). In other areas, the tumor-necrosis interface lacked pyknotic changes (arrowheads and box). The interface region within the box (E) is shown at higher magnification in F, G, and H. In F, areas of previous necrosis began to resolve with ingrowth of granulation tissue stroma (upper right). The adjacent tumor (lower left) appeared to be viable and proliferative, without nuclear pyknosis and karyorrhexis in the border zone. In G and H, immunostains for endothelial cell markers demonstrated the high vascularity of the stromal ingrowth. In I, mFlt(1-3)IgG/Mab A4.6.1-treated tumors were largely necrotic. A thin peripheral zone of viable tumor remains adjacent to the border of the tumor mass and surrounding host tissues. In J, the tumor-necrosis interface in mFlt-1 (1-3)IgG-treated tumors showed progression from viable tumor through pyknosis and karyorrhexis, indicating continuing cell death. In K and L, immunostains for endothelial cell markers revealed a complete lack of neovascularization at the tumor-necrosis interface. Nonspecific staining of the necrotic regions as shown here was also seen in nonspecific isotype-matched primary antibody controls. A single vessel is present within the viable tumor (arrow). It is notable that in control tumors, immunoreactivity for Flk-1 (C) was more intense than for CD31 (D), whereas in the anti-VEGF-treated tumors, this relationship was reversed (G and K versus H and L). Scale bars: 1 mm (A, E, and I); 100 μm (B–D, F–H, and J–L).
- Fig. 3.
RT-PCR gene expression analysis of control tumors and tumors treated with Mab A4.6.1/Flt-IgG. A, comparison of relative RNA levels of human genes between total RNA isolated from tumors harvested from mice treated with Mab A.4.6.1/Flt(1-3)-IgG or control/control during a 12-day treatment period. Black bars, relative changes in percentage to control-treated tumors. Statistically significant changes: ∗, P < 0.05; ∗∗, P < 0.005 (n = 5). Expression levels have been standardized to the probe/primer sets specific for human or murine GAPDH, respectively. B, comparison of RNA levels of mouse genes with total RNA isolated from tumors as defined in A. ∗, P < 0.05; ∗∗, P < 0.005 (n = 5).
Tables
Volume 60, Issue 22
