Figure 6: Effects of in vivo ZNF304 gene silencing on tumour growth and vasculature.

(a) Effect of ZNF304 siRNA-DANP treatment on tumour weight (left panel) and number of nodules (right panel) in HeyA8 orthotopic murine model (P values obtained with Student’s t-test; *P<0.05; **P<0.01; ***P<0.001 or ****P<0.0001; compared with control siRNA-treated group; bars and error bars represent mean values and the corresponding s.e.m). (b) Knockdown of ZNF304 by DANP-ZNF304 siRNA and the effect of treatment in SKOV3 tumour weight (left) and number of nodules (right). Data are presented as mean± s.e.m. (n=10 per group). *P<0.05; **P<0.01; ***P<0.001 or ****P<0.0001 (Student’s t-test). (c) Immunohistochemical stainings for tumour proliferation (Ki67) and microvessel density (CD31) in SKOV3 orthotopic murine model of OC. Scale bar, 100 μm. Quantification of Ki67-positive and CD31-positive cells in control siRNA and ZNF304 siRNA-treated groups are shown in Supplementary Fig. 14. (d) Kaplan–Meier survival curve illustrating the effects of DANP-ZNF304 siRNA treatment versus control siRNA treatment for the in vivo OVCA-432 survival model. Survival curves indicate that biweekly treatment of DANP-ZNF304 siRNA improves survival in vivo (n=8 per group, P=0.01 (Control siRNA versus ZNF304 siRNA), Log-rank (Mantel-Cox) test). (e) Viability of epithelial cells in ascites of mice. DANP-siRNA was administered intravenously when ascites was detectable. Ascites was removed 7 days after a single administration and viability of epithelial cells were detected by FITC-Epcam and PI staining followed by flow cytometry (n=3, P<0.0001, Student’s t-test). NS, not significant.