The design and generation of complex multifunctional macromolecular structures by bioconjugation is a hot topic due to increasing interest in conjugates with therapeutic applications. In this regard, the development of efficient, selective, and safe conjugation methods is a major objective. In this report, we describe the use of the bis(bromomethyl)benzene scaffold as a linker for bioconjugation with special emphasis on antibody conjugation. We first performed the monothioalkylation of 1,3,5-tris(bromomethyl)benzene, which rendered the reactive dibromotrimethylbenzyl derivatives to be used in thiol bis-alkylation. Next, we introduced into the linker either a bis(Cys)-containing peptide or anti-CD4 and -CD13 monoclonal antibodies, previously subjected to partial reduction of disulfide bonds. Mass spectrometry, UV–vis spectra, and SDS-PAGE experiments revealed that this bis-alkylating agent for bioconjugation preserved both antibody integrity and antibody–antigen binding affinity, as assessed by flow cytometry. Taken together, our results show that the mesitylene scaffold is a suitable linker for thiol-based bioconjugation reactions. This linker could be applicable in the near future for the preparation of antibody drug conjugates.

中文翻译：

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均三甲苯硫醇烷基化的生物共轭

由于对结合物的治疗兴趣日益浓厚，通过生物共轭作用设计和生成复杂的多功能大分子结构是一个热门话题。在这方面，开发有效，选择性和安全的缀合方法是一个主要目标。在此报告中，我们描述了使用双（溴甲基）苯支架作为生物偶联的连接子，并特别强调抗体偶联。我们首先进行了1,3,5-三（溴甲基）苯的单硫代烷基化反应，这使反应性二溴三甲基苄基衍生物可用于硫醇双烷基化反应。接下来，我们将含有bis（Cys）的肽或抗CD4和-CD13单克隆抗体引入连接子，这些抗体先前已部分还原了二硫键。质谱，紫外可见光谱，和SDS-PAGE实验表明，通过流式细胞术评估，这种用于生物偶联的双烷基化剂既保留了抗体的完整性，又保留了抗体-抗原的结合亲和力。综上所述，我们的结果表明，均三甲苯基支架是基于硫醇的生物缀合反应的合适连接体。该接头可在不久的将来应用于抗体药物偶联物的制备。