In vivo antitumor activity of DNA-specific ADP-ribosyltransferase, pierisin-1

Pierisin-1是一种在甘蓝蝴蝶中发现的98 kDa的蛋白质，对癌细胞具有细胞毒性。为了验证该蛋白在体内是否具有抗癌活性，对小鼠的急性毒性和对移植瘤的抑制作用进行了分析。小鼠腹腔注射胡椒素-1的半数致死量为5μg/kg体重。6周龄BALB/c nu/nu雌性裸鼠接种1×10~(-7)HeLa细胞，次日按3μg/kg剂量腹腔注射菌毛菌素-1。此后，动物在第80天被宰杀。10只未注射胡椒素-1的小鼠均成瘤，平均瘤重为1.16g/只。在接受Pierisin-1治疗的小鼠中，每只小鼠的平均肿瘤重量为0.56g，其值明显较低，在接受测试的10只小鼠中，有3只没有发现肿瘤。我们的结果表明，Pierisin-1在体内具有抗肿瘤活性。接下来，我们从1,340只菜粉蝶蛹中提纯了47.6 mg的菜青素-1。酶学分析表明，N末端多肽和胰酶处理的Pierisin-1的最大周转次数分别为每秒55次和25次。在某些条件下，相关的C末端片段对N末端的酶结构域有明显的抑制作用。长期的反应表明，一个Pierisin-1分子能够产生超过10^6个ADP-核糖化的DNA加合物，这可能会导致哺乳动物细胞的死亡。

Pierisin-1, a 98-kDa protein found in cabbage butterfly, exhibits cytotoxicity to cancer cells. To examine whether this protein has anti-cancer activity in viva, acute toxicity to mice and suppression effects to transplanted tumors were analyzed. The LD_<50> value for pierisin-1 was around 5 μg/kg body weight on intraperitoneal injection into BALB/c mice. BALB/c nu/nu female 6-week-old nude mice inoculated 1 x 10^7 HeLa cells on the first day were subjected for intraperitoneal injection of pierisin-1 at a dose of 3 μg/kg on the next day. Thereafter, animals were killed at day 80. Tumors were formed in all ten mice treated without pierisin-1 and the mean tumor weight was 1.16 g per mouse. In mice treated with pierisin-1, the mean tumor weight was 0.56 g per mouse, and its value was significantly lower and no tumors were found in three of ten mice tested. Our results indicate that pierisin-1 has antitumor activity in viva. Next, we purified 47.6 mg pierisin-1 from 1,340 pupae of Pieris rapae. Enzymatic characterization demonstrated that the maximum turnover number of the N-terminal polypeptide and trypsin-treated pierisin-1 were 55 and 25 per second, respectively. Inhibition of the N-terminal enzyme domain by the associated C-terminal fragment was evident under certain conditions. Long-term reactions indicated that a single molecule of pierisin-1 has the capacity to generate more than 10^6 ADP-ribosylated DNA adducts, which could cause the death of a mammalian cell.