Department of Biochemistry, Indian Institute of Science, Biological Sciences Building, Bangalore, 560012, India.
Molecular Biophysics Unit, Indian Institute of Science, Bangalore, 560012, India.
Arch Virol. 2020 May;165(5):1163-1176. doi: 10.1007/s00705-020-04581-y. Epub 2020 Mar 30.
Monoclonal antibodies have attracted wide attention in therapeutics owing to their high efficacy, low toxicity, and specific targeting. However, antibodies cannot cross the cell membrane barrier. Therefore, their therapeutic potential is limited to surface-exposed antigens or secreted proteins. In the present investigation, we have developed a chimeric virus-like particle (VLP) of pepper vein banding virus (PVBV) and explored the possibility of using it as a delivery vehicle for antibodies against intracellular antigens as well as for future applications in immunodiagnostics. The chimeric PVBV particles were generated by genetically engineering the B domain of Staphylococcus aureus protein A (SpA) at the N-terminus of the PVBV coat protein (CP). The chimeric VLPs purified by sucrose density gradient centrifugation had ~440-fold higher affinity towards IgG antibody when compared to SpA. Interestingly, the unassembled chimeric CP with the B-domain at the N-terminus (BCP) purified by Ni-NTA chromatography was a monomer, and it had ~45-fold higher affinity towards antibodies compared to SpA. Additionally, the chimeric particles were able to bind and deliver antibodies against both intracellular (α-tubulin) and surface-exposed antigens (CD 20). However, the BCP monomer failed to enter mammalian cells. Thus, for the first time, we have demonstrated that the assembled VLPs are essential for internalization. These results demonstrate the potential of the use of chimeric PVBV VLPs in diagnostics and, more importantly, as nanocarriers for intracellular delivery of antibodies.
单克隆抗体由于其高效、低毒和特异性靶向等特点，在治疗学中引起了广泛关注。然而，抗体不能穿过细胞膜屏障。因此，它们的治疗潜力仅限于表面暴露的抗原或分泌蛋白。在本研究中，我们开发了一种辣椒叶脉坏死病毒（PVBV）的嵌合病毒样颗粒（VLP），并探索了将其用作针对细胞内抗原的抗体的递送载体的可能性，以及在免疫诊断学中的未来应用。嵌合 PVBV 颗粒是通过在 PVBV 外壳蛋白（CP）的 N 末端基因工程改造金黄色葡萄球菌蛋白 A（SpA）的 B 结构域而产生的。通过蔗糖密度梯度离心纯化的嵌合 VLP 与 SpA 相比，对 IgG 抗体的亲和力高约 440 倍。有趣的是，用 Ni-NTA 色谱法纯化的具有 N 末端 B 结构域的未组装嵌合 CP（BCP）是一个单体，与 SpA 相比，它对抗体的亲和力高约 45 倍。此外，嵌合颗粒能够结合并递送至细胞内（α-微管蛋白）和表面暴露的抗原（CD20）的抗体。然而，BCP 单体不能进入哺乳动物细胞。因此，我们首次证明了组装的 VLP 对于内化是必需的。这些结果表明，嵌合 PVBV VLP 具有用于诊断的潜力，更重要的是，作为细胞内抗体递送的纳米载体。
