An Jaeyeon, Kim Sungwan, Shrinidhi Annadka, Kim Junghyun, Banna Hasanul, Sung Gihyun, Park Kyeng Min, Kim Kimoon
Center for Self-Assembly and Complexity (CSC), Institute for Basic Science (IBS), Pohang, Republic of Korea.
Department of Chemistry, Pohang University of Science and Technology (POSTECH), Pohang, Republic of Korea.
Nat Biomed Eng. 2020 Nov;4(11):1044-1052. doi: 10.1038/s41551-020-0589-7. Epub 2020 Jul 20.
Efficient purification is crucial to providing large quantities of recombinant therapeutic proteins, such as monoclonal antibodies and cytokines. However, affinity techniques for manufacturing protein therapeutics that use biomolecule-conjugated agarose beads that harness specific biomolecular interactions suffer from issues related to protein denaturation, contamination and the need to maintain biomolecule-specific conditions for efficient protein capture. Here, we report a versatile and scalable method for the purification of recombinant protein therapeutics. The method exploits the high-affinity and controllable host-guest interactions between cucurbit[7]uril (CB[7]) and selected guests such as adamantylammonium. We show that the Herceptin (the brand name of trastuzumab, a monoclonal antibody drug used to treat breast cancer) and the much smaller cytokine interferon α-2a can be purified by site-specifically tagging them with adamantylammonium using the enzyme sortase A, followed by high-affinity binding with CB[7]-conjugated agarose beads and the recovery of the protein using a guest with a stronger affinity for CB[7]. The thermal and chemical stability of CB[7] beads and their scalability, recyclability and low cost may also make them advantageous for the manufacturing of biosimilars.
高效纯化对于大量生产重组治疗性蛋白质至关重要，例如单克隆抗体和细胞因子。然而，用于制造蛋白质治疗药物的亲和技术，即使用利用特定生物分子相互作用的生物分子偶联琼脂糖珠，存在与蛋白质变性、污染以及为有效捕获蛋白质而需维持生物分子特异性条件相关的问题。在此，我们报告了一种用于纯化重组蛋白质治疗药物的通用且可扩展的方法。该方法利用了葫芦[7]脲（CB[7]）与选定客体（如金刚烷基铵）之间的高亲和力和可控的主客体相互作用。我们表明，赫赛汀（曲妥珠单抗的商品名，一种用于治疗乳腺癌的单克隆抗体药物）以及小得多的细胞因子干扰素α - 2a可以通过使用分选酶A用金刚烷基铵对它们进行位点特异性标记来纯化，随后与CB[7]偶联的琼脂糖珠进行高亲和力结合，并使用对CB[7]具有更强亲和力的客体回收蛋白质。CB[7]珠的热稳定性和化学稳定性及其可扩展性、可回收性和低成本也可能使其在生物类似药的生产中具有优势。
