Protein-Polymer Conjugates Synthesized Using Water-Soluble Azlactone-Functionalized Polymers Enable Receptor-Specific Cellular Uptake toward Targeted Drug Delivery.
Protein-Polymer Conjugates Synthesized Using Water-Soluble Azlactone-Functionalized Polymers Enable Receptor-Specific Cellular Uptake toward Targeted Drug Delivery.
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DOI：
10.1021/acs.bioconjchem.9b00155
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发表时间：
2019-03
影响因子：
4.7
通讯作者：
Julia S Kim;Allison R. Sirois;Analia J. Vazquez Cegla;Eugenie Jumai’an;Naomi Murata;M. E. Buck;Sarah J Moore
中科院分区：
文献类型：
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作者：
Julia S Kim;Allison R. Sirois;Analia J. Vazquez Cegla;Eugenie Jumai’an;Naomi Murata;M. E. Buck;Sarah J Moore
将蛋白质偶联到载药聚合物结构上是一种有吸引力的策略，可以促进多种临床需要的靶向药物递送。目前可用于结合蛋白质的聚合物通常具有有限的化学通用性，用于随后的药物装载。许多具有用于药物装载的化学功能的聚合物通常不溶于水，这使得合成用于靶向药物递送的功能性蛋白质-聚合物偶联物变得困难。在这项工作中，我们证明了反应性的，氮杂内酯功能化的聚合物可以接枝到蛋白质上，结合到小分子荧光团上，然后以受体特异性的方式内化到细胞中。采用可逆加成-断裂链转移聚合法制备了聚（2-乙烯基-4,4-二甲基内酯），经亚化学计量当量的三甘醇单甲醚修饰后聚合得到活性水溶性氮杂内酯功能化共聚物。然后将这些活性聚合物偶联到全转铁蛋白和卵转铁蛋白上。蛋白质凝胶分析证实了蛋白质与聚合物的成功偶联，随后使用尺寸排除色谱法从未反应的蛋白质和聚合物中纯化蛋白质-聚合物偶联物。在乳腺癌细胞株表面过表达转铁蛋白受体的内化实验表明，全转铁蛋白-聚合物偶联物被细胞成功内化，其机制与受体介导的内吞作用一致。蛋白质-聚合物缀合物的内在化表明，蛋白质配体在与聚合物缀合后保持了其整体结构和功能。我们的蛋白质-聚合物偶联物合成方法为制备广泛的生物医学应用感兴趣的生物偶联物提供了一种简单，可定制的策略。
Conjugation of proteins to drug-loaded polymeric structures is an attractive strategy for facilitating target-specific drug delivery for a variety of clinical needs. Polymers currently available for conjugation to proteins generally have limited chemical versatility for subsequent drug loading. Many polymers that do have chemical functionality useful for drug loading are often insoluble in water, making it difficult to synthesize functional protein-polymer conjugates for targeted drug delivery. In this work, we demonstrate that reactive, azlactone-functionalized polymers can be grafted to proteins, conjugated to a small-molecule fluorophore, and subsequently internalized into cells in a receptor-specific manner. Poly(2-vinyl-4,4-dimethylazlactone), synthesized using reversible addition-fragmentation chain transfer polymerization, was modified post-polymerization with substoichiometric equivalents of triethylene glycol monomethyl ether to yield reactive water-soluble, azlactone-functionalized copolymers. These reactive polymers were then conjugated to proteins holo-transferrin and ovotransferrin. Protein gel analysis verified successful conjugation of proteins to polymer, and protein-polymer conjugates were subsequently purified from unreacted proteins and polymers using size exclusion chromatography. Internalization experiments using a breast cancer cell line that overexpresses the transferrin receptor on its surface showed that the holo-transferrin-polymer conjugate was successfully internalized by cells in a mechanism consistent with receptor-mediated endocytosis. Internalization of protein-polymer conjugate demonstrated that the protein ligand maintained its overall structure and function following conjugation to polymer. Our approach to protein-polymer conjugate synthesis offers a simple, tailorable strategy for preparing bioconjugates of interest for a broad range of biomedical applications.