UCIBIO, REQUIMTE, Departamento de Ciências Químicas, Faculdade de Farmácia, Universidade do Porto, Porto, Portugal.
Institute of Organic Chemistry and Biochemistry, Academy of Sciences of the Czech Republic, Flemingovo nám. 2, 16610, Prague 6, Czech Republic.
Sci Rep. 2017 Jul 24;7(1):6343. doi: 10.1038/s41598-017-06445-z.
Despite doxorubicin being commonly used in chemotherapy there still remain significant holes in our knowledge regarding its delivery efficacy and an observed resistance mechanism that is postulated to involve the cell membrane. One possible mechanism is the efflux by protein P-gp, which is found predominantly in cholesterol enriched domains. Thereby, a hypothesis for the vulnerability of doxorubicin to efflux through P-gp is its enhanced affinity for the ordered cholesterol rich regions of the plasma membrane. Thus, we have studied doxorubicin's interaction with model membranes in a cholesterol rich, ordered environment and in liquid-disordered cholesterol poor environment. We have combined three separate experimental protocols: UV-Vis spectrophotometry, fluorescence quenching and steady-state anisotropy and computational molecular dynamics modeling. Our results show that the presence of cholesterol induces a change in membrane structure and doesn't impair doxorubicin's membrane partitioning, but reduces drug's influence on membrane fluidity without directly interacting with it. It is thus possible that the resistance mechanism that lowers the efficacy of doxorubicin, results from an increased density in membrane regions where the efflux proteins are present. This work represents a successful approach, combining experimental and computational studies of membrane based systems to unveil the behavior of drugs and candidate drug molecules.
尽管多柔比星在化疗中被广泛应用，但我们对其输送效果仍存在显著的认识空白，并且观察到一种被认为涉及细胞膜的耐药机制。一种可能的机制是蛋白 P-gp 的外排，它主要存在于富含胆固醇的区域。因此，推测多柔比星易通过 P-gp 外排的假设是其与质膜中有序富含胆固醇区域的增强亲和力。因此，我们研究了多柔比星在富含胆固醇的有序环境和液体无序胆固醇缺乏环境中与模型膜的相互作用。我们结合了三种独立的实验方案：紫外可见分光光度法、荧光猝灭和稳态各向异性以及计算分子动力学建模。我们的结果表明，胆固醇的存在会引起膜结构的变化，不会损害多柔比星的膜分配，但会降低药物对膜流动性的影响，而不会直接与之相互作用。因此，降低多柔比星疗效的耐药机制可能是由于存在外排蛋白的膜区域密度增加所致。这项工作代表了一种成功的方法，结合了基于膜系统的实验和计算研究，揭示了药物和候选药物分子的行为。
