Figure 5. Polymeric particles targeted to tumor cell surface amplify immune cytokinemediated apoptosis in vivo.

(a) Schematic of epithelial cell adhesion molecule (EpCAM)-targeted particle delivery to COLO 205 tumour cells in nude (nu/nu) mice in vivo, followed by treatment with TRAIL. Mice were inoculated with COLO 205 tumour cells via tail vein injection (2 × 10⁶ cells), followed by injection of nontargeted and EpCAM-targeted PLGA particles (500 nm diameter; ∼500 particles per tumour cell) 15 min post tumour cell injection. At 30 min post particle injection, mice were treated with TRAIL (0.1 μg ml⁻¹ plasma concentration). Tumour cells in blood were collected via submandibular bleed 90 min post TRAIL injection. Tumour cells were detected in vivo via whole-body bioluminescent imaging (BLI) at 7 and 14 days post injection. (b) Representative flow cytometry plots of GFP+ COLO 205 tumour cells removed after delivery of nontargeted particles (Particles) and EpCAM-targeted particles (t-Particles) followed by TRAIL. FSC, forward scatter; SSC, side scatter. (c) Number of viable GFP+ COLO 205 tumour cells per ml mouse blood 90 min post TRAIL treatment of tumour cells in vivo under various conditions. Cells only denotes mice treated with tumour cells followed by PBS via tail vein injection. N=5 mice for all treatments. (d) Representative whole-body BLI images of COLO 205 tumour cells in mice 7 days post injection of particles and targeted particles followed by TRAIL. (e) COLO 205 BLI signals in mice 7 and 14 days post injection of COLO 205 tumour cells under various conditions. N=5 mice for all treatments. (f) PC-3 tumour growth curves after intravenous injections of targeted particles (40 mg kg⁻¹) followed by TRAIL (15 mg kg⁻¹) 3 h post particle injection. For combination therapies, tumour-bearing nu/nu mice were also treated with the TRAIL-sensitizer resveratrol (30 mg kg⁻¹). After tumour formation (100 mm³), mice began treatment regimen and tumour volume was measured every 3 days. Blue arrows indicate days where mice were treated with targeted particles, followed 3 h later by TRAIL treatment. Green arrows indicate days where mice were treated with resveratrol via oral gavage. N=5 mice for all treatments. Data are reported as the mean±s.e. Different treatment groups were compared for statistical significance using Student's two-tailed t-test for two conditions and one-way analysis of variance (ANOVA) for multiple comparisons. *P<0.05, **P<0.01 and ***P<0.001. NS, not significant.