Department of Radiation Oncology, School of Medicine, Washington University in St Louis, St Louis, MO, USA.
Department of Infectious Disease and Global Health, Tufts Cummings School of Veterinary Medicine, North Grafton, MA, USA.
Lab Invest. 2014 Aug;94(8):893-905. doi: 10.1038/labinvest.2014.82. Epub 2014 Jun 16.
The unique ability of human adenovirus serotype 5 (Ad5) to accomplish efficient transduction has allowed the use of Ad5-based vectors for a range of gene therapy applications. Several strategies have been developed to alter tropism of Ad vectors to achieve a cell-specific gene delivery by using fiber modifications via genetic incorporation of targeting motifs. In this study, we have explored the utility of novel anti-human carcinoembryonic antigen (hCEA) single variable domains derived from heavy chain (VHH) camelid family of antibodies to achieve targeted gene transfer. To obtain anti-CEA VHHs, we produced a VHH-display library from peripheral blood lymphocytes RNA of alpacas at the peak of immune response to the hCEA antigen (Ag). We genetically incorporated an anti-hCEA VHH into a de-knobbed Ad5 fiber-fibritin chimera and demonstrated selective targeting to the cognate epitope expressed on the membrane surface of target cells. We report that the anti-hCEA VHH used in this study retains Ag recognition functionality and provides specificity for gene transfer of capsid-modified Ad5 vectors. These studies clearly demonstrated the feasibility of retargeting of Ad5-based gene transfer using VHHs.
人类腺病毒血清型 5（Ad5）独特的高效转导能力使得基于 Ad5 的载体能够应用于多种基因治疗。已经开发了几种策略来改变 Ad 载体的趋向性，通过遗传整合靶向基序来实现对特定细胞的基因传递。在这项研究中，我们探索了新型抗人癌胚抗原（hCEA）单可变结构域的用途，这些结构域来源于重链（VHH）骆驼科抗体，以实现靶向基因传递。为了获得抗 CEA VHH，我们在免疫反应高峰期从羊驼外周血淋巴细胞 RNA 中产生了一个 VHH 展示文库针对 hCEA 抗原（Ag）。我们将抗 hCEA VHH 基因整合到去 knob 化的 Ad5 纤维-纤维蛋白嵌合体中，并证明了对靶细胞表面表达的同源表位的选择性靶向。我们报告说，本研究中使用的抗 hCEA VHH 保留了 Ag 识别功能，并为壳修饰的 Ad5 载体的基因转移提供了特异性。这些研究清楚地证明了使用 VHH 重新靶向 Ad5 基因为基础的基因转移的可行性。
