Bortolotti Massimo, Biscotti Francesco, Zanello Andrea, Bolognesi Andrea, Polito Letizia
Department of Medical and Surgical Sciences-DIMEC, General Pathology Section, Alma Mater Studiorum University of Bologna, 40126 Bologna, Italy.
Biomedicines. 2023 Apr 19;11(4):1214. doi: 10.3390/biomedicines11041214.
Saporin is a type 1 ribosome-inactivating protein widely used as toxic payload in the construction of targeted toxins, chimeric molecules formed by a toxic portion linked to a carrier moiety. Among the most used carriers, there are large molecules (mainly antibodies) and small molecules (such as neurotransmitters, growth factors and peptides). Some saporin-containing targeted toxins have been used for the experimental treatment of several diseases, giving very promising results. In this context, one of the reasons for the successful use of saporin lies in its resistance to proteolytic enzymes and to conjugation procedures. In this paper, we evaluated the influence of derivatization on saporin using three heterobifunctional reagents, namely 2-iminothiolane (2-IT), N-succinimidyl 3-(2-pyridyldithio)propionate (SPDP) and 4-succinimidyloxycarbonyl-α-methyl-α-[2-pyridyldithio]toluene (SMPT). In order to obtain the highest number of inserted -SH groups with the lowest reduction of saporin biological activities, we assessed the residual ability of saporin to inhibit protein synthesis, to depurinate DNA and to induce cytotoxicity after derivatization. Our results demonstrate that saporin maintains an excellent resistance to derivatization processes, especially with SPDP, and permit us to define reaction conditions, in which saporin biological properties may not be altered. Therefore, these findings provide useful information for the construction of saporin-based targeted toxins, especially with small carriers.
皂草素是一种1型核糖体失活蛋白，在构建靶向毒素（即由与载体部分相连的毒性部分形成的嵌合分子）时被广泛用作毒性载荷。在最常用的载体中，有大分子（主要是抗体）和小分子（如神经递质、生长因子和肽）。一些含皂草素的靶向毒素已被用于几种疾病的实验性治疗，并取得了非常有前景的结果。在这种情况下，成功使用皂草素的原因之一在于其对蛋白水解酶和偶联程序具有抗性。在本文中，我们使用三种异双功能试剂，即2-亚氨基硫醇（2-IT）、N-琥珀酰亚胺基3-（2-吡啶二硫基）丙酸酯（SPDP）和4-琥珀酰亚胺氧基羰基-α-甲基-α-[2-吡啶二硫基]甲苯（SMPT），评估了衍生化对皂草素的影响。为了在皂草素生物活性降低最少的情况下获得最多的插入-SH基团，我们评估了衍生化后皂草素抑制蛋白质合成、使DNA脱嘌呤和诱导细胞毒性的残留能力。我们的结果表明，皂草素对衍生化过程保持着优异的抗性，尤其是对SPDP，这使我们能够确定不会改变皂草素生物学特性的反应条件。因此，这些发现为构建基于皂草素的靶向毒素，尤其是与小载体结合的靶向毒素，提供了有用的信息。
