In vivo silencing of a molecular target by short interfering RNA electroporation: tumor vascularization correlates to delivery efficiency.
Screening for a molecular target for cancer therapy requires multiple steps, of which an important one is evaluation of the knockdown effect of the target molecule on pregrown xenograft tumors. However, methods currently used for local administration of knockdown reagents, such as short interfering RNA (siRNA), are not satisfactory as to simplicity and efficiency. We established an electroporation method involving a constant voltage and "plate and fork" type electrodes and used it for in vivo delivery of siRNA. The delivery efficiency correlated to the electric current. The electric current correlated to the microvascular density and vascular endothelial growth factor (VEGF) expression and exhibited a threshold that guaranteed efficient delivery. Consequently, we showed that the vascularization and VEGF expression in tumors determined the efficiency of delivery of siRNA by electroporation. VEGF was chosen as a model target. VEGF siRNA electroporation suppressed the growth of tumors exhibiting high VEGF expression to less than 10% of the control level, but it had no effect on low VEGF-expressing tumors. Notably, a long interval (20 days) of electroporation was enough to obtain a satisfactory effect. Systemically injected siRNA could also be delivered into tumors by this method. Our data will provide the technical basis for in vivo electroporation, and this simple and efficient siRNA delivery method is applicable to in vivo comprehensive screening for a molecular target.
Takei
Yoshifumi
Y
Department of Biochemistry, Nagoya University Graduate School of Medicine, 65 Tsurumai-cho, Showa-ku, Nagoya 466-8550, Japan. takei@med.nagoya-u.ac.jp
Nemoto
Toshio
T
Mu
Ping
P
Fujishima
Tatsuya
T
Ishimoto
Takuji
T
Hayakawa
Yasuhiko
Y
Yuzawa
Yukio
Y
Matsuo
Seiichi
S
Muramatsu
Takashi
T
Kadomatsu
Kenji
K
D000818:Animals; D045744:Cell Line, Tumor; D018274:Electroporation / Q000379:methods*; D020868:Gene Silencing; D006801:Humans; D051379:Mice; D008819:Mice, Nude; D009369:Neoplasms / Q000098:blood supply* / Q000235:genetics* / Q000378:metabolism / Q000473:pathology; D009389:Neovascularization, Pathologic / Q000235:genetics* / Q000378:metabolism / Q000473:pathology*; D034741:RNA, Small Interfering / Q000008:administration & dosage* / Q000235:genetics*; D042461:Vascular Endothelial Growth Factor A / Q000378:metabolism; D023041:Xenograft Model Antitumor Assays
D034741:RNA, Small Interfering; D042461:Vascular Endothelial Growth Factor A