Cuellar Trinna L, Barnes Dwight, Nelson Christopher, Tanguay Joshua, Yu Shang-Fan, Wen Xiaohui, Scales Suzie J, Gesch Julie, Davis David, van Brabant Smith Anja, Leake Devin, Vandlen Richard, Siebel Christian W
Department of Molecular Biology, Genentech, Inc., 1 DNA Way, South San Francisco, CA 94080-0511, USA.
Department of Discovery Oncology, Genentech, Inc., 1 DNA Way, South San Francisco, CA 94080-0511, USA.
Nucleic Acids Res. 2015 Jan;43(2):1189-203. doi: 10.1093/nar/gku1362. Epub 2014 Dec 30.
Delivery of siRNA is a key hurdle to realizing the therapeutic promise of RNAi. By targeting internalizing cell surface antigens, antibody-siRNA complexes provide a possible solution. However, initial reports of antibody-siRNA complexes relied on non-specific charged interactions and have not been broadly applicable. To assess and improve this delivery method, we built on an industrial platform of therapeutic antibodies called THIOMABs, engineered to enable precise covalent coupling of siRNAs. We report that such coupling generates monomeric antibody-siRNA conjugates (ARCs) that retain antibody and siRNA activities. To broadly assess this technology, we generated a battery of THIOMABs against seven targets that use multiple internalization routes, enabling systematic manipulation of multiple parameters that impact delivery. We identify ARCs that induce targeted silencing in vitro and extend tests to target prostate carcinoma cells following systemic administration in mouse models. However, optimal silencing was restricted to specific conditions and only observed using a subset of ARCs. Trafficking studies point to ARC entrapment in endocytic compartments as a limiting factor, independent of the route of antigen internalization. Our broad characterization of multiple parameters using therapeutic-grade conjugate technology provides a thorough assessment of this delivery technology, highlighting both examples of success as well as remaining challenges.
小干扰RNA（siRNA）的递送是实现RNA干扰（RNAi）治疗前景的关键障碍。通过靶向内化细胞表面抗原，抗体-siRNA复合物提供了一种可能的解决方案。然而，关于抗体-siRNA复合物的初步报道依赖于非特异性的电荷相互作用，并未得到广泛应用。为了评估和改进这种递送方法，我们基于一种名为硫醇化抗体（THIOMABs）的治疗性抗体工业平台进行构建，该平台经过工程改造，能够实现siRNA的精确共价偶联。我们报道这种偶联产生了保留抗体和siRNA活性的单体抗体-siRNA缀合物（ARCs）。为了广泛评估这项技术，我们针对七个使用多种内化途径的靶点生成了一系列硫醇化抗体，从而能够系统地操纵影响递送的多个参数。我们鉴定出在体外诱导靶向沉默的ARCs，并将测试扩展到在小鼠模型中全身给药后靶向前列腺癌细胞。然而，最佳沉默仅限于特定条件，并且仅在一部分ARCs中观察到。转运研究表明，ARC被困在内吞小室中是一个限制因素，与抗原内化途径无关。我们使用治疗级缀合技术对多个参数进行的广泛表征，对这种递送技术进行了全面评估，突出了成功案例以及尚存的挑战。
