Owczarzy Aleksandra, Rogóż Wojciech, Kulig Karolina, Zięba Andrzej, Maciążek-Jurczyk Małgorzata
Department of Physical Pharmacy, Faculty of Pharmaceutical Sciences in Sosnowiec, Medical University of Silesia, 40-055, Katowice, Poland.
Department of Organic Chemistry, Faculty of Pharmaceutical Sciences in Sosnowiec, Medical University of Silesia, 40-055, Katowice, Poland.
Naunyn Schmiedebergs Arch Pharmacol. 2025 Feb 13. doi: 10.1007/s00210-025-03884-8.
The aim of this work was to extend the existing knowledge of the interaction between newly synthetized substances with anticancer properties (5-methyl-12(H)-chino[3,4-b]-[1,4]-benzothiazine chloride (Salt1), 9-fluoro-5-alkyl-12(H)-quino[3,4-b][1,4]benzothiazine chloride (Salt2), and 9-amino-5-alkyl-12(H)-quino[3,4-b][1,4]benzothiazine chloride (Salt3) with model plasma carrier proteins. The thermodynamic profile of ligand-protein complexation and the contribution of bonds responsible for complex formation have been studied using calorimetry technique. The research has theoretical and experimental nature, but from a scientific point of view is novelty due to promising biological properties of Salt1, Salt2, and Salt3 and provide an important basis for further in vitro and in vivo studies. All measurements were conducted using nanoITC calorimeter (TA Instruments, New Castle, USA). The results were analyzed using Launch NanoAnalyze program (TA Instruments, New Castle, USA). Based on the obtained data, it is safe to consider the bonds within Salt1-HSA, Salt3-HSA, Salt1-AGP, Salt3-AGP, and Salt3-HGG complexes to be predominantly hydrophobic (ΔH > 0 and ΔS > 0) with K values: (1.95 ± 0.59)·10, (34.6 ± 0.06)·10, (3.34 ± 0.35)·10, (0.45 ± 0.14)·10, and (0.56 ± 0.09)·10 (L·mol), respectively. In contrast, complexes of Salt2 with proteins were stabilized by hydrogen bonds and/or van der Waals interaction (ΔH < 0 and ΔS < 0) and K values (25.50 ± 9.20)·10, (1.37 ± 0.37)·10, and (1.17 ± 0.01)·10 (L·mol) for HSA, AGP, and HGG, respectively, have been obtained. In turn, the reaction of Salt1-HGG complex formation was accompanied by ionic bonds (ΔH ≅ 0, ΔS > 0, and K = (0.64 ± 0.45)·10 (L·mol)). Regardless of the involvement of bonds and interaction between the ligands and proteins, the reactions occurred spontaneously (ΔG < 0). By comparing the binding parameters obtained using nanocalorimetric measurements and previously obtained spectroscopic data, due to the characteristic of complex formation, Salt2 was selected for further analysis. In addition, it was found that, despite the many advantages of the nanoITC technique, it still requires coupling with other techniques that allow analysis of the complexes formed at the molecular level and complementing spectroscopic analysis. Therefore, the use of these two techniques should be considered simultaneously.
这项工作的目的是扩展关于新合成的具有抗癌特性的物质（5-甲基-12(H)-喹诺[3,4-b]-[1,4]-苯并噻嗪氯化物（盐1）、9-氟-5-烷基-12(H)-喹诺[3,4-b][1,4]苯并噻嗪氯化物（盐2）和9-氨基-5-烷基-12(H)-喹诺[3,4-b][1,4]苯并噻嗪氯化物（盐3））与模型血浆载体蛋白之间相互作用的现有知识。利用量热技术研究了配体 - 蛋白质络合的热力学概况以及负责络合物形成的键的贡献。该研究具有理论和实验性质，但从科学角度来看具有新颖性，因为盐1、盐2和盐3具有有前景的生物学特性，并为进一步的体外和体内研究提供了重要依据。所有测量均使用纳米ITC量热仪（TA仪器公司，美国特拉华州纽卡斯尔）进行。结果使用Launch NanoAnalyze程序（TA仪器公司，美国特拉华州纽卡斯尔）进行分析。基于获得的数据，可以认为盐1 - 人血清白蛋白（HSA）、盐3 - HSA、盐1 - 血清类粘蛋白（AGP）、盐3 - AGP和盐3 - 人γ-球蛋白（HGG）络合物中的键主要是疏水的（ΔH＞0且ΔS＞0），其K值分别为：(1.95 ± 0.59)·10、(34.6 ± 0.06)·10、(3.34 ± 0.35)·10、(0.45 ± 0.14)·10和(0.56 ± 0.09)·10（L·mol）。相比之下，盐2与蛋白质的络合物通过氢键和/或范德华相互作用得以稳定（ΔH＜0且ΔS＜0），并且分别获得了人血清白蛋白、血清类粘蛋白和人γ-球蛋白的K值为(25.50 ± 9.20)·10、(1.37 ± 0.37)·10和(1.17 ± 0.01)·10（L·mol）。反过来，盐1 - HGG络合物的形成反应伴随着离子键（ΔH≅0，ΔS＞0，且K = (0.64 ± 0.45)·10（L·mol））。无论配体与蛋白质之间键的参与情况以及相互作用如何，反应都是自发发生的（ΔG＜0）。通过比较使用纳米量热测量获得的结合参数和先前获得的光谱数据，由于络合物形成的特性，选择盐2进行进一步分析。此外，还发现尽管纳米ITC技术有许多优点，但它仍然需要与其他能够在分子水平分析形成的络合物并补充光谱分析的技术相结合。因此，应同时考虑使用这两种技术。
