Vascular targeting of anti-CD40 antibodies and IL-2 into autochthonous tumors enhances immunotherapy in mice
J. Clin. Invest. Juliana Hamzah, et al. 118:1691 doi:10.1172/JCI33201 [Go to this article.]

Figure 3
Intratumoral anti-CD40/IL-2 treatment reduces tumor vascularity. (A) Representative anti-CD31 vessel staining in 30-week-old RIP1-Tag5 tumors in anti-CD40–RGR/IL-2–RGR (left panel) or anti-CD40/IL-2 (right panel) treatment groups after 7 weeks of treatment and vessel counts. Vessel density was quantified by counting the number of CD31⁺ vessels within randomly selected fields (n = 5 mice; 5 fields per tumor; *P = 0.005 compared with control mice treated with IgG/RGR peptide). (B) Corresponding hypoxia stain (hypoxyprobe-1 kit; Chemicon) and quantification of hypoxic tumor areas in different treatment groups (n = 5; *P = 0.005 compared with control mice). (C) CD8⁺ T cells in anti–CD40-RGR/IL-2–RGR (left panel) or anti-CD40/IL-2 (right panel) treated tumors and quantification of CD8⁺ and CD4⁺ T cell infiltrates (n = 5; no statistically significant difference was observed between treatment groups). (D) Schematic representation of treatment with anti-CD4⁺ and anti-CD8⁺ depletion for a period of 5 weeks. (E) Survival analysis of anti-CD4⁺ and anti-CD8⁺ depleted RIP1-Tag5 mice (control) and RIP1-Tag5 mice treated with anti-CD40–RGR/IL-2–RGR combination therapy and depleting antibodies or IgG control (n = 8; P = 0.005). (F) Quantification of vessel density in depletion groups (n = 3 mice; 5 fields per tumor; *P = 0.005, **P = 0.005 compared with CD4⁺/CD8⁺ depleted control mice). Original magnification, ×10 (A); ×20 (B and C). Scale bar: 100 μm (A); 50 μm (B and C).