Designer gene therapy using an Escherichia coli purine nudeloside phosphorylase/prodrug system
Designer gene therapy using an Escherichia coli purine nudeloside phosphorylase/prodrug system
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DOI：
10.1016/j.chembiol.2003.11.008
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发表时间：
2003-12-01
影响因子：
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通讯作者：
Ealick, SE
中科院分区：
文献类型：
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作者：
Bennett, EM;Anand, R;Ealick, SE
大肠杆菌嘌呤核苷磷酸化酶(PNP)激活前药提供了一种选择性杀死表达转染的PNP基因的肿瘤细胞的方法。这种基因治疗方法需要将前药与仅存在于肿瘤细胞中的已知酶活性相匹配。该方法的特异性依赖于避免前药被宿主细胞或肠道菌群中已经存在的酶裂解。以晶体学和计算机建模方法为指导，我们重新设计了大肠杆菌 PNP，能够比野生型酶更有效地裂解新的前药底物。特别是，M64V PNP 突变体裂解 9-(6-脱氧-α-L-呋喃糖基)-6-甲基嘌呤，其 k(cat)/K-m 比天然大肠杆菌 PNP 高 100 倍以上。在异种移植肿瘤实验中，该化合物引起表达 M64V PNP 基因的肿瘤消退。
Activation of prodrugs by Escherichia coli purine nucleoside phosphorylase (PNP) provides a method for selectively killing tumor cells expressing a transfected PNP gene. This gene therapy approach requires matching a prodrug and a known enzymatic activity present only in tumor cells. The specificity of the method relies on avoiding prodrug cleavage by enzymes already present in the host cells or the intestinal flora. Using crystallographic and computer modeling methods as guides, we have redesigned E. coli PNP to cleave new prodrug substrates more efficiently than does the wild-type enzyme. In particular, the M64V PNP mutant cleaves 9-(6-deoxy-alpha-L-talofuranosyl)-6-methyipurine with a k(cat)/K-m over 100 times greater than for native E. coli PNP. In a xenograft tumor experiment, this compound caused regression of tumors expressing the M64V PNP gene.