Watanabe Masami, Nasu Yasutomo, Kashiwakura Yuji, Kusumi Norihiro, Tamayose Kenji, Nagai Atsushi, Sasano Tetsuo, Shimada Takashi, Daida Hiroyuki, Kumon Hiromi
Department of Urology, Okayama University Graduate School of Medicine and Dentistry, Okayama 700-8558, Japan.
Hum Gene Ther. 2005 Jun;16(6):699-710. doi: 10.1089/hum.2005.16.699.
Maspin is a member of the serine protease inhibitors and the maspin gene, a tumor suppressor gene, is down-regulated in a large fraction of prostate cancers. We evaluated the use of adeno-associated virus (AAV, serotype 2) vector encoding maspin as a means for in vivo gene therapy for human prostate cancer. TUNEL assay of subcutaneously formed LNCaP or DU145 tumors in nude mice showed that intratumoral AAV-mediated maspin expression significantly upregulated the number of apoptotic cells compared with AAV-LacZ treatment. Immunofluorescence double staining for maspin protein and apoptosis in LNCaP tumors showed that the percentage of apoptotic cells in AAV-maspin-mediated maspin-expressing cells was significantly high compared with that in AAV-GFP-mediated GFP-expressing cells. Moreover, significantly fewer CD31-positive microvessels were observed in AAV-maspin-treated tumors compared with the control tumors. These therapeutic responses were highly correlated to persistent maspin expression in tumors, confirmed by Western blot analysis until at least day 56 after treatment. Finally, intratumoral delivery of AAV-maspin significantly suppressed growth of LNCaP and DU145 tumors and improved survival of mice. We conclude that AAV-mediated prolonged maspin expression efficiently suppresses human prostate tumor growth in vivo by apoptosis induction and inhibition of angiogenesis.
Maspin是丝氨酸蛋白酶抑制剂家族的一员，maspin基因作为一种肿瘤抑制基因，在大部分前列腺癌中表达下调。我们评估了使用编码maspin的腺相关病毒（AAV，2型）载体作为人前列腺癌体内基因治疗手段的效果。对裸鼠皮下形成的LNCaP或DU145肿瘤进行TUNEL检测显示，与AAV-LacZ治疗相比，瘤内AAV介导的maspin表达显著上调了凋亡细胞的数量。对LNCaP肿瘤中maspin蛋白和凋亡进行免疫荧光双重染色显示，与AAV-GFP介导的GFP表达细胞相比，AAV-maspin介导的maspin表达细胞中的凋亡细胞百分比显著更高。此外，与对照肿瘤相比，在AAV-maspin治疗的肿瘤中观察到的CD31阳性微血管明显更少。这些治疗反应与肿瘤中maspin的持续表达高度相关，Western印迹分析证实至少在治疗后56天仍有表达。最后，瘤内注射AAV-maspin显著抑制了LNCaP和DU145肿瘤的生长并提高了小鼠的存活率。我们得出结论，AAV介导的maspin长期表达通过诱导凋亡和抑制血管生成在体内有效抑制人前列腺肿瘤生长。
