Jugel Willi, Tietze Stefanie, Daeg Jennifer, Appelhans Dietmar, Broghammer Felix, Aigner Achim, Karimov Michael, Schackert Gabriele, Temme Achim
Department of Neurosurgery, Section Experimental Neurosurgery and Tumor Immunology, University Hospital Carl Gustav Carus, TU Dresden, Fetscherstraße 74, 01307 Dresden, Germany.
Leibniz Institute of Polymer Research Dresden e.V., Mailbox 120411, 01069 Dresden, Germany.
Cancers (Basel). 2022 Apr 11;14(8):1925. doi: 10.3390/cancers14081925.
Among non-viral vectors, cationic polymers, such as poly(propylene imine) (PPI), play a prominent role in nucleic acid delivery. However, limitations of polycationic polymer-based DNA delivery systems are (i) insufficient target specificity, (ii) unsatisfactory transgene expression, and (iii) undesired transfer of therapeutic DNA into non-target cells. We developed single-chain antibody fragment (scFv)-directed hybrid polyplexes for targeted gene therapy of prostate stem cell antigen (PSCA)-positive tumors. Besides mono-biotinylated PSCA-specific single-chain antibodies (scFv(AM1-P-BAP)) conjugated to neutravidin, the hybrid polyplexes comprise β-cyclodextrin-modified PPI as well as biotin/maltose-modified PPI as carriers for minicircle DNAs encoding for Sleeping Beauty transposase and a transposon encoding the gene of interest. The PSCA-specific hybrid polyplexes efficiently delivered a GFP gene in PSCA-positive tumor cells, whereas control hybrid polyplexes showed low gene transfer efficiency. In an experimental gene therapy approach, targeted transposition of a codon-optimized p53 into p53-deficient HCT116 cells demonstrated decreased clonogenic survival when compared to mock controls. Noteworthily, p53 transposition in PTEN-deficient H4 glioma cells caused nearly complete loss of clonogenic survival. These results demonstrate the feasibility of combining tumor-targeting hybrid polyplexes and Sleeping Beauty gene transposition, which, due to the modular design, can be extended to other target genes and tumor entities.
在非病毒载体中，阳离子聚合物，如聚（丙烯亚胺）（PPI），在核酸递送中发挥着重要作用。然而，基于聚阳离子聚合物的DNA递送系统存在以下局限性：（i）靶向特异性不足；（ii）转基因表达不理想；（iii）治疗性DNA意外转移到非靶细胞中。我们开发了用于前列腺干细胞抗原（PSCA）阳性肿瘤靶向基因治疗的单链抗体片段（scFv）导向的混合多聚体。除了与中性抗生物素蛋白偶联的单生物素化PSCA特异性单链抗体（scFv(AM1-P-BAP)）外，混合多聚体还包含β-环糊精修饰的PPI以及生物素/麦芽糖修饰的PPI，作为编码睡美人转座酶的微型环DNA和编码目的基因的转座子的载体。PSCA特异性混合多聚体在PSCA阳性肿瘤细胞中有效地递送了GFP基因，而对照混合多聚体显示出低基因转移效率。在一种实验性基因治疗方法中，与模拟对照相比，将密码子优化的p53靶向转座到p53缺陷的HCT116细胞中显示出克隆形成存活率降低。值得注意的是，在PTEN缺陷的H4胶质瘤细胞中进行p53转座导致克隆形成存活率几乎完全丧失。这些结果证明了将肿瘤靶向混合多聚体与睡美人基因转座相结合的可行性，由于其模块化设计，该方法可扩展到其他靶基因和肿瘤实体。
